**Meet the editors**

Professor Antonio Valero has obtained a degree in Biology and a PhD in Food Science and Technology at the University of Cordoba. He has currently more than 10 years' experience in managing projects, training courses, academic teaching, and events in food and related sectors. Professor Valero has participated in different national and international research projects related to

predictive modelling and risk assessment (USDA-Fresh Express, Selection of fit-for-purpose sampling procedures for specific foods and risks, BASE-LINE, COST Action 920). He has also been involved in collaborative work with EFSA through participation in working groups as an external expert (Public health risks represented by certain composite products containing food of animal origin, 2012–2013 and Self-tasking mandate for scientific opinion on the requirements for the development of microbiological criteria). He has participated in two recently awarded contracts from EFSA (OC/EFSA/BIOCONTAM/2014/02 and NP/EFSA/BIOCONTAM/2015/04) in relation to the development of a risk assessment for *Listeria monocytogenes* in selected RTE food categories. He has strong involvement in national and international networks regarding food safety (SICURA), being a member of the Spanish Microbiology Society and International Association for Food Protection. His research activity is widely known as he has published over 70 peer-reviewed papers and book chapters mainly related to the application of predictive models in foods, microbial risk assessment, and management and derivation of microbiological criteria in selected foods. Additionally, he has presented more than 80 communications in different congresses and symposia since 2003. He also belongs to the editorial boards of international peer-reviewed journals.

During his career, he has participated in different international events and training workshops supported by different programs/initiatives (Erasmus+, Spanish Ministries, etc.) in the EU and worldwide. In light of his position at the Department of Food Science and Technology (University of Cordoba), Antonio Valero has a broad experience (5 years+) in academic teaching within the food science and technology and veterinary degrees. In addition, he participates as an invited teacher in national and international masters related to food quality and food safety.

Professor Dr. Rosa Mª García-Gimeno is a full professor at the Department of Food Science and Technology at the University of Cordoba (lecturer 1996–present) in the subjects of Food Hygiene and Inspection and Descriptive Food Science of the degrees of Veterinary Medicine and Food Science. She has a broad research background focused on quantitative risk assessment of biologi-

Contents

**Preface VII**

**and Quality 3**

Saskia M. van Ruth

**Compounds 43**

**Liver Diseases 93**

Adámez

Chapter 1 **The Dairy Industry: Process, Monitoring, Standards,**

Michael P. Ryan and Catherine C. Adley

Chapter 3 **Agrifood By-Products as a Source of Phytochemical**

Petravić Tominac and Ana Mornar

**Section 2 Effects of Technologies on Food Quality 113**

Uzma Latief and Riaz Ahmad

Chapter 2 **Food Authenticity: Provenancing. A Case Study of Fish 27**

Chapter 4 **Chemical Constituents of Fruit Wines as Descriptors of their**

Chapter 5 **Role of Dietary Carotenoids in Different Etiologies of Chronic**

Chapter 6 **Quality of Reduced-Fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying Methods 115**

Simin Hedayatnia and Hamed Mirhosseini

Niamh Burke, Krzysztof A. Zacharski, Mark Southern, Paul Hogan,

Annemieke M. Pustjens, Rita Boerrigter-Eenling, Alex H. Koot and

Sara Martillanes, Javier Rocha-Pimienta and Jonathan Delgado-

**Nutritional, Sensorial and Health-Related Properties 59** Darko Velić, Daniela Amidžić Klarić, Natalija Velić, Ilija Klarić, Vlatka

**Section 1 Quality of Food 1**

cal hazards and predictive microbiology activities through the study of diverse pathogen behavior in different food matrices (leafy green salads, fish, and cooked and cured meat products) and shelf-life estimations. She has authored more than 67 indexed scientific articles and 24 chapters of books. She has directed or taken part in 23 projects and competitive research contracts and 14 projects with companies.

## Contents

## **Preface XI**



#### Chapter 7 **Effects of Radiation Technologies on Food Nutritional Quality 137**

Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de Souza

Preface

ty and safety.

cialization to meet consumer expectations.

their role in the prevention of liver diseases.

The increasing consumer demand for healthier and more convenient foods has led to the implementation of novel strategies and food management systems at an industrial level. Knowledge of the main food types—their physical and chemical properties—as well as transformation processes could provide a better understanding of how to improve commer‐

*Descriptive Food Science* is based on the principles of food composition and quality, its nutri‐ tional value, and the potential application of food technology in relation to a selected range of food categories. Different aspects in relation to the engineering, biological, and physical sciences are illustrated to study the nature of foods, the causes of product deterioration, the principles underlying food processing, and the improvement of foods for the consuming public. Throughout the book, definitions of different food types provide basic knowledge and understanding of the relationship between structural food composition and food quali‐ ty criteria (nutritional and commercial value). Current food legislation, standards, and the

*Descriptive Food Science* presents a structured series of chapters dealing with this subject. It stresses the importance of assessing food quality parameters and understanding how proc‐ essing conditions and food formulation can influence final product properties. The book is

The first part deals with food quality and comprises five chapters. *The Dairy Industry: Proc‐ esses, Monitoring, Standards and Quality* describes the whole milk processing chain, stressing quality and safety aspects, as well as reinforcing the idea of implementing new technologies into an "Industry 4.0" concept. *Geographical Origin of Foods: Fish Provenance* highlights the importance of food authenticity to assure consumers' confidence and to increase food quali‐

*Agrifood By-products as a Source of Phytochemical Compounds* explains the classification of phy‐ tochemical compounds and the analytical methods for their quantification. Their relevance in the food industry is also discussed in this chapter. A step forward in this topic is descri‐ bed in the chapter *Chemical Constituents of Fruit Wines as Descriptors of Their Nutritional, Sen‐ sorial and Health-related Properties* where the authors give an overview of various factors used to evaluate the quality and functional properties of fruit wines. To finalize this part, the chapter *Role of Dietary Carotenoids in Etiologies of Chronic Liver Diseases* provides a compre‐ hensive review of dietary carotenoids and includes updated information with respect to

application of novel processing methodologies are also emphasized.

organized into two macro parts, and subdivided into eight chapters in total.

#### Chapter 8 **Ultrasound Application to Improve Meat Quality 153** Alma D. Alarcon-Rojo, Esmeralda Peña-González, Iván García-Galicia, Luis Carrillo-López, Mariana Huerta-Jiménez, Raúl Reyes-

Villagrana and Hector Janacua-Vidales

## Preface

Chapter 7 **Effects of Radiation Technologies on Food Nutritional Quality 137**

Chapter 8 **Ultrasound Application to Improve Meat Quality 153**

Villagrana and Hector Janacua-Vidales

Souza

**VI** Contents

Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de

Alma D. Alarcon-Rojo, Esmeralda Peña-González, Iván García-Galicia, Luis Carrillo-López, Mariana Huerta-Jiménez, Raúl Reyes-

> The increasing consumer demand for healthier and more convenient foods has led to the implementation of novel strategies and food management systems at an industrial level. Knowledge of the main food types—their physical and chemical properties—as well as transformation processes could provide a better understanding of how to improve commer‐ cialization to meet consumer expectations.

> *Descriptive Food Science* is based on the principles of food composition and quality, its nutri‐ tional value, and the potential application of food technology in relation to a selected range of food categories. Different aspects in relation to the engineering, biological, and physical sciences are illustrated to study the nature of foods, the causes of product deterioration, the principles underlying food processing, and the improvement of foods for the consuming public. Throughout the book, definitions of different food types provide basic knowledge and understanding of the relationship between structural food composition and food quali‐ ty criteria (nutritional and commercial value). Current food legislation, standards, and the application of novel processing methodologies are also emphasized.

> *Descriptive Food Science* presents a structured series of chapters dealing with this subject. It stresses the importance of assessing food quality parameters and understanding how proc‐ essing conditions and food formulation can influence final product properties. The book is organized into two macro parts, and subdivided into eight chapters in total.

> The first part deals with food quality and comprises five chapters. *The Dairy Industry: Proc‐ esses, Monitoring, Standards and Quality* describes the whole milk processing chain, stressing quality and safety aspects, as well as reinforcing the idea of implementing new technologies into an "Industry 4.0" concept. *Geographical Origin of Foods: Fish Provenance* highlights the importance of food authenticity to assure consumers' confidence and to increase food quali‐ ty and safety.

> *Agrifood By-products as a Source of Phytochemical Compounds* explains the classification of phy‐ tochemical compounds and the analytical methods for their quantification. Their relevance in the food industry is also discussed in this chapter. A step forward in this topic is descri‐ bed in the chapter *Chemical Constituents of Fruit Wines as Descriptors of Their Nutritional, Sen‐ sorial and Health-related Properties* where the authors give an overview of various factors used to evaluate the quality and functional properties of fruit wines. To finalize this part, the chapter *Role of Dietary Carotenoids in Etiologies of Chronic Liver Diseases* provides a compre‐ hensive review of dietary carotenoids and includes updated information with respect to their role in the prevention of liver diseases.

The second part of the book moves towards food technological aspects, and is comprised of the final three chapters. *Quality of Reduced-fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying Methods* investigates different analytical methods and formulations of the physicochemical properties of regular and instant reduced-fat dairy creamers. Novel technologies are studied in the chapter *Effects of Radiation Technologies on Food Nutritional Quality*, aimed at discussing retention or loss of vitamins and minerals in several food prod‐ ucts submitted to a radiation process. Finally, the chapter *Ultrasound Application to Improve Meat Quality* shows the recent potential applications of ultrasound in meat systems, as well as physical and chemical effects of ultrasound treatments on the preservation and modifica‐ tion of processed meat foods.

*Descriptive Food Science* is oriented to both academic and research purposes and provides a substantial contribution to the food science and technology field.

#### **Prof. Antonio Valero Díaz and Prof. Rosa Mª García-Gimeno**

Department of Food Science and Technology Faculty of Veterinary University of Cordoba, Spain **Quality of Food**

**Section 1**

**Section 1**

## **Quality of Food**

The second part of the book moves towards food technological aspects, and is comprised of the final three chapters. *Quality of Reduced-fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying Methods* investigates different analytical methods and formulations of the physicochemical properties of regular and instant reduced-fat dairy creamers. Novel technologies are studied in the chapter *Effects of Radiation Technologies on Food Nutritional Quality*, aimed at discussing retention or loss of vitamins and minerals in several food prod‐ ucts submitted to a radiation process. Finally, the chapter *Ultrasound Application to Improve Meat Quality* shows the recent potential applications of ultrasound in meat systems, as well as physical and chemical effects of ultrasound treatments on the preservation and modifica‐

*Descriptive Food Science* is oriented to both academic and research purposes and provides a

**Prof. Antonio Valero Díaz and Prof. Rosa Mª García-Gimeno**

Department of Food Science and Technology

Faculty of Veterinary

University of Cordoba, Spain

substantial contribution to the food science and technology field.

tion of processed meat foods.

VIII Preface

**Chapter 1**

**Provisional chapter**

**The Dairy Industry: Process, Monitoring, Standards,**

**The Dairy Industry: Process, Monitoring, Standards,** 

Sampling and analysis occur along the milk processing train: from collection at farm level, to intake at the diary plant, the processing steps, and the end products. Milk has a short shelf life; however, products such as milk powders have allowed a global industry to be developed. Quality control tests are vital to support activities for hygiene and food standards to meet regulatory and customer demands. Multiples of chemical and microbiological contamination tests are undertaken. Hazard analysis testing strategies are necessary, but some tests may be redundant; it is therefore vital to identify product optimization quality control strategies. The time taken to undergo testing and turnaround time are rarely measured. The dairy industry is a traditional industry with a low margin commodity. Industry 4.0 vision for dairy manufacturing is to introduce the aspects of operational excellence and implementation of information and communications technologies. The dairy industries' reply to Industry 4.0 is represented predominantly by proactive maintenance and optimization of production and logistical chains, such as robotic milking machines and processing and packaging line automation reinforced by sensors for rapid chemical and microbial analysis with improved and real-time data management. This chapter reviews the processing trains with suggestions for improved optimization.

**Keywords:** dairy, processing, hygiene, analytical tests, automation

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

The implementation of strategies to improve and strengthen milk process optimization is of vital importance within the dairy industry. The rapid deterioration of milk products forces

DOI: 10.5772/intechopen.80398

**and Quality**

**and Quality**

Catherine C. Adley

Catherine C. Adley

**Abstract**

**1. Introduction**

Niamh Burke, Krzysztof A. Zacharski,

Niamh Burke, Krzysztof A. Zacharski,

http://dx.doi.org/10.5772/intechopen.80398

Mark Southern, Paul Hogan, Michael P. Ryan and

Mark Southern, Paul Hogan, Michael P. Ryan and

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

#### **The Dairy Industry: Process, Monitoring, Standards, and Quality The Dairy Industry: Process, Monitoring, Standards, and Quality**

DOI: 10.5772/intechopen.80398

Niamh Burke, Krzysztof A. Zacharski, Mark Southern, Paul Hogan, Michael P. Ryan and Catherine C. Adley Niamh Burke, Krzysztof A. Zacharski, Mark Southern, Paul Hogan, Michael P. Ryan and Catherine C. Adley

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.80398

#### **Abstract**

Sampling and analysis occur along the milk processing train: from collection at farm level, to intake at the diary plant, the processing steps, and the end products. Milk has a short shelf life; however, products such as milk powders have allowed a global industry to be developed. Quality control tests are vital to support activities for hygiene and food standards to meet regulatory and customer demands. Multiples of chemical and microbiological contamination tests are undertaken. Hazard analysis testing strategies are necessary, but some tests may be redundant; it is therefore vital to identify product optimization quality control strategies. The time taken to undergo testing and turnaround time are rarely measured. The dairy industry is a traditional industry with a low margin commodity. Industry 4.0 vision for dairy manufacturing is to introduce the aspects of operational excellence and implementation of information and communications technologies. The dairy industries' reply to Industry 4.0 is represented predominantly by proactive maintenance and optimization of production and logistical chains, such as robotic milking machines and processing and packaging line automation reinforced by sensors for rapid chemical and microbial analysis with improved and real-time data management. This chapter reviews the processing trains with suggestions for improved optimization.

**Keywords:** dairy, processing, hygiene, analytical tests, automation

#### **1. Introduction**

The implementation of strategies to improve and strengthen milk process optimization is of vital importance within the dairy industry. The rapid deterioration of milk products forces

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

dairy processors to critically optimize and plan their production schedules. The business model is to look at the work force, to reduce or eliminate any time or/and resource wastage, unnecessary costs, bottlenecks, and mistakes while attaining the process objective of creating a quality product [1].

The global dairy sector is currently going through change. The Food and Agricultural Organization of the United Nations (FAO-UN), dairy price index shows prices 26% below its peak from February 2014 [2]. The demand for milk products from China is beginning to slow, trade sanctions on Russia and the end of "milk quotas" within the European Union (EU) has caused a period of excess supply and low prices [3]. Notwithstanding this, the dairy sector is expanding and projected to grow at a rate of 1.8% per year over the next 10 years, to 177 million tons of powdered milk by 2025 [4]. This increase is mainly due to rising urbanization and growing incomes in emerging markets [5]. In the EU, however, dairy farmers have used intervention stocks to shield themselves from poorer international prices. In September 2017, for instance, EU farmers consigned 16,597 tons of skimmed milk powder (SMP) to the interventions stock at Euro €1.698 [6].

The milk proteins contain the nine essential amino acids required by humans, making it an important human food. The caseins are easily digested, while the whey proteins are relatively

\*The moisture content does not include water of crystallization of the lactose, the milk solids-not-fat content includes the

**Whole milk powder** 

**Acid whey powder** 

5

**(WP) %**

http://dx.doi.org/10.5772/intechopen.80398

The Dairy Industry: Process, Monitoring, Standards, and Quality

**(WMP) %**

The milk fat content varies within the same dairy products and between different dairy products. Raw farm milk, full-fat milk, semi-skimmed milk, and skimmed milk have their own percentage of fat. Raw milk normally has a fat content of ~4.4 g of milk fat per 100 g. This can be skimmed to obtain lower fat varieties. Full-fat milk is standardized to 3.5% of fat and semi-skimmed milk contains ~1.5% fat. Skimmed milk and buttermilk are very low in fat and, on average, contain 0.1 or 0.2% fat, respectively. The fat content of milk and cream is also known as butterfat, an important factor in determining the price to be paid for milk supplied by farmers in many countries. Milk sold to the consumer is standardized with a range of different fat content choices. However, international variances in standardization mean that the fat percentage for (semi)-skimmed, whole milk, and buttermilk can differ between countries. Modifications in the composition of milk are allowed, if they are indicated on the packing of the product, so that it can be easily seen and read, complying with the obligation as regards nutrition labeling, laid down by the countries regulations. In the case of the EU, regulation No. 1169/2011 applies on the provision of food information to consumers [9], plus providing an indication of origin, is considered of particular interest. The US Public Health Service (USPHS) Milk Ordinance and Code recommends a minimum of 3.25% butterfat in farm milk,

Milk is not necessarily a local product and has developed into a global trade with the development of milk powders. In particular, whole milk powder (WMP) and skimmed milk powder (SMP) are the most traded agricultural commodities globally, as percentage of production traded, while fresh dairy products, with less than 1% of production traded are the least traded agricultural commodity [4]. The dairy industry, however, has been targeted in the climate change debate as it has been estimated that 14.5% of greenhouse gas emissions come from livestock with beef and milk production the main culprits [13, 14]. Extreme changes in climate can affect the microbiological safety of food. Wet conditions are favorable to pathogen growth and may result in increased risk of food contamination, including mycotoxin. Aflatoxin M1 is the most studied mycotoxin in milk and levels exceeding the EU maximum level (0.050 μg/kg) have been found [15]. Climate effects on animal diseases lead to increased use of veterinary

less digestible in the intestine.

**Cow's milk %**

water of crystallization of the lactose (Source: [11, 12]).

**Table 1.** Average composition of milk and milk powders.

**Skim milk powder** 

Moisture 85.5–89.5 3.0–4.0 2.0–4.5\* 3.5–5.0 Fat 2.5–6.0 0.6–1.5 26.0–42.0 1.0–1.5 Protein 2.9–5.0 34.0–37.00 24.5–27 11.0–14.5 Lactose 3.6–5.5 49.5–52.0 36.0–38.5 63.0–75.0 Minerals (ash) 0.8–0.9 8.2–8.6 5.5–6.5 8.2–8.8

**(SMP) %**

as the official national standard [10].

In addition, changing consumer demand patterns are affecting food production. The "Traditional" value drivers of price, taste, and convenience have been complemented by newer and "Evolving" drivers such as health and wellness, safety, social impact, and experience. Central to all of these drivers is a need for transparency from food companies [5]. Given the ever-changing nature of the consumer food value drivers, dairy producers must look to their production processes to innovate with new products and to optimize output without compromising on quality and safety.

The world's milk is predominantly cow's milk, followed by buffalo milk. The leading producers include, Asia (30%), followed by the EU (28%), North and Central America (18%), South America (9%), other European countries (9%), Africa (5%), and Oceania (5%) [7]. To be named a dairy product, food must be produced from the milk of cows, buffalo, goats, etc. The dairy sector includes food such as liquid milk, milk powders, cheese, butter, and yogurt, as well as ice cream. Several factors including genetics, and breed of animal, environment, stages of lactation, parity, and nutrition, together determine the final composition of milk [8]. Milk and dairy products are significant sources of protein, essential minerals (calcium, potassium, magnesium, phosphorous, sodium, iodine) and several vitamins, (the fat-soluble vitamins A, D, E, K, and B1, B3, B6, B12). In a Western diet, dairy products provide between 40 and 70% of the recommended daily calcium intake. Cow's milk consists of about 87% water (**Table 1**), and 12–13% total solids. The solids consist of fat ~4% and solids-not-fat (SNF) ~9%, such as proteins, lactose, and various minerals and vitamins. Milk proteins consist of whey and caseins; caseins have four different species (αS1, αS2, β, and κ-caseins) which are separate molecules, but they do possess similarity in structure and they comprise around 80% of total milk protein. The major whey proteins in cow and sheep's milk are β-lactoglobulin and α-lactalbumin; the other proteins are serum albumin and immunoglobulins. Minor proteins include lactoferrin (LF), an iron binding protein and β<sup>2</sup> -Microglobulin—part of the Major Histocompatibility Complex II (MHC II), the rest are mostly enzymes including; lactoperoxidase, an enzyme that breaks down hydrogen peroxide, lysozyme which breaks down bacterial cell walls and has low activity in cow milk, proteases, protease activators, nucleases, glycosidases, and others.


\*The moisture content does not include water of crystallization of the lactose, the milk solids-not-fat content includes the water of crystallization of the lactose (Source: [11, 12]).

**Table 1.** Average composition of milk and milk powders.

dairy processors to critically optimize and plan their production schedules. The business model is to look at the work force, to reduce or eliminate any time or/and resource wastage, unnecessary costs, bottlenecks, and mistakes while attaining the process objective of creating

The global dairy sector is currently going through change. The Food and Agricultural Organization of the United Nations (FAO-UN), dairy price index shows prices 26% below its peak from February 2014 [2]. The demand for milk products from China is beginning to slow, trade sanctions on Russia and the end of "milk quotas" within the European Union (EU) has caused a period of excess supply and low prices [3]. Notwithstanding this, the dairy sector is expanding and projected to grow at a rate of 1.8% per year over the next 10 years, to 177 million tons of powdered milk by 2025 [4]. This increase is mainly due to rising urbanization and growing incomes in emerging markets [5]. In the EU, however, dairy farmers have used intervention stocks to shield themselves from poorer international prices. In September 2017, for instance, EU farmers consigned 16,597 tons of skimmed milk powder (SMP) to the inter-

In addition, changing consumer demand patterns are affecting food production. The "Traditional" value drivers of price, taste, and convenience have been complemented by newer and "Evolving" drivers such as health and wellness, safety, social impact, and experience. Central to all of these drivers is a need for transparency from food companies [5]. Given the ever-changing nature of the consumer food value drivers, dairy producers must look to their production processes to innovate with new products and to optimize output without

The world's milk is predominantly cow's milk, followed by buffalo milk. The leading producers include, Asia (30%), followed by the EU (28%), North and Central America (18%), South America (9%), other European countries (9%), Africa (5%), and Oceania (5%) [7]. To be named a dairy product, food must be produced from the milk of cows, buffalo, goats, etc. The dairy sector includes food such as liquid milk, milk powders, cheese, butter, and yogurt, as well as ice cream. Several factors including genetics, and breed of animal, environment, stages of lactation, parity, and nutrition, together determine the final composition of milk [8]. Milk and dairy products are significant sources of protein, essential minerals (calcium, potassium, magnesium, phosphorous, sodium, iodine) and several vitamins, (the fat-soluble vitamins A, D, E, K, and B1, B3, B6, B12). In a Western diet, dairy products provide between 40 and 70% of the recommended daily calcium intake. Cow's milk consists of about 87% water (**Table 1**), and 12–13% total solids. The solids consist of fat ~4% and solids-not-fat (SNF) ~9%, such as proteins, lactose, and various minerals and vitamins. Milk proteins consist of whey and caseins; caseins have four different species (αS1, αS2, β, and κ-caseins) which are separate molecules, but they do possess similarity in structure and they comprise around 80% of total milk protein. The major whey proteins in cow and sheep's milk are β-lactoglobulin and α-lactalbumin; the other proteins are serum albumin and immunoglobulins. Minor proteins include lactofer-

Complex II (MHC II), the rest are mostly enzymes including; lactoperoxidase, an enzyme that breaks down hydrogen peroxide, lysozyme which breaks down bacterial cell walls and has low activity in cow milk, proteases, protease activators, nucleases, glycosidases, and others.


a quality product [1].

4 Descriptive Food Science

ventions stock at Euro €1.698 [6].

compromising on quality and safety.

rin (LF), an iron binding protein and β<sup>2</sup>

The milk proteins contain the nine essential amino acids required by humans, making it an important human food. The caseins are easily digested, while the whey proteins are relatively less digestible in the intestine.

The milk fat content varies within the same dairy products and between different dairy products. Raw farm milk, full-fat milk, semi-skimmed milk, and skimmed milk have their own percentage of fat. Raw milk normally has a fat content of ~4.4 g of milk fat per 100 g. This can be skimmed to obtain lower fat varieties. Full-fat milk is standardized to 3.5% of fat and semi-skimmed milk contains ~1.5% fat. Skimmed milk and buttermilk are very low in fat and, on average, contain 0.1 or 0.2% fat, respectively. The fat content of milk and cream is also known as butterfat, an important factor in determining the price to be paid for milk supplied by farmers in many countries. Milk sold to the consumer is standardized with a range of different fat content choices. However, international variances in standardization mean that the fat percentage for (semi)-skimmed, whole milk, and buttermilk can differ between countries. Modifications in the composition of milk are allowed, if they are indicated on the packing of the product, so that it can be easily seen and read, complying with the obligation as regards nutrition labeling, laid down by the countries regulations. In the case of the EU, regulation No. 1169/2011 applies on the provision of food information to consumers [9], plus providing an indication of origin, is considered of particular interest. The US Public Health Service (USPHS) Milk Ordinance and Code recommends a minimum of 3.25% butterfat in farm milk, as the official national standard [10].

Milk is not necessarily a local product and has developed into a global trade with the development of milk powders. In particular, whole milk powder (WMP) and skimmed milk powder (SMP) are the most traded agricultural commodities globally, as percentage of production traded, while fresh dairy products, with less than 1% of production traded are the least traded agricultural commodity [4]. The dairy industry, however, has been targeted in the climate change debate as it has been estimated that 14.5% of greenhouse gas emissions come from livestock with beef and milk production the main culprits [13, 14]. Extreme changes in climate can affect the microbiological safety of food. Wet conditions are favorable to pathogen growth and may result in increased risk of food contamination, including mycotoxin. Aflatoxin M1 is the most studied mycotoxin in milk and levels exceeding the EU maximum level (0.050 μg/kg) have been found [15]. Climate effects on animal diseases lead to increased use of veterinary medicines resulting in drug resistance and anthropogenic (synthetic) chemicals with the potential for transmission of chemical residues into the food chain. The more frequent and intense rainfall that is predicted could encourage the spread of perchlorate through surface runoff with the potential to enter the food chain via cow's milk [16]. Perchlorate reduces thyroid hormone production in the thyroid gland [17].

## **2. Milk processing**

The flow diagram for milk processing is presented in **Figure 1**. Milk arrives at the milk dairy processing plant over the weighbridge and the weight of milk is automatically recorded. At the same time, data from an on-board computer is downloaded wirelessly to a data capture system, which holds the records of the temperature and volumes of milk collected from each farm. The temperature should be at 4–6°C. Milk samples using sterile containers are collected automatically from each supplier at source and are delivered to a laboratory technician for detailed analysis. Milk that deviates in composition, taste, and smell from normal milk receives a lower quality rating. The technician also takes a composite sample, from each compartment in the refrigerated truck, which is compartmentalized to reduce sloshing of the milk. The samples from each compartment are tested for acidity, antibiotics, added water, fat, and protein content. These analytical tests and methods are determined by international standards as outlined in **Table 2**.

The ISO standards catalog ISO/TC34/SC5 [18] lists all milk and milk products standards, while other standard sets include, microbiology of the food chain, microbiological quality of milk, etc. The bacterial quality of the milk is also measured and these specify tests are outlined later.

The titratable acid test measures the acidity of the milk. Both titratable acidity (TA) and pH are measures of acid. TA is a more reliable indicator because relative to pH measurement, it is more sensitive to small changes in milk acidity, especially important in cheese making. The acidity of milk is of two types; natural acidity due to citrates and phosphates present in the milk and dissolved CO2 during the processing of milking. The second is the developed acidity due to lactic acid produced by bacteria using the lactose in the milk as a nutrient, converting it to lactic acid. The acidity of milk measures the total acidity (natural acidity of milk and developed acidity). The International Standard Method for titratable acid is ISO 6091:2010 [19]. Titratable acidity is a measure of the buffering of milk between pH 6.6 and 8.3 (phenolphthalein endpoint) [26]. The appearance of a faint pink color, which signals the endpoint and the number of ml of NaOH used to reach the endpoint, is recorded. This value is called the "titer," titratable acidity is reported as percent lactic acid and is dependent on the volume of sample. As this test is dependent on the analyst reading eye measurement of the color change, it is prone to human error causing incorrect and unpredictable recording of results.

The antibiotic test uses kits known as Charm and Delvo tests. The Charm test is made by Charm Science Inc., e.g., one kit, the Charm Rosa TET–SL (www.charm.com), detects chlortetracycline, oxytetracycline, and tetracycline residues in raw milk in the initial assay at or below 100 ppb, which complies with the EU regulation No 2377/90 [27], ISO 26844:2006 [20], and Codex Alimentarius regulations CAC/MRL 2-2015 [28]. The Delvo test (a rapid kit developed by DSM, food and beverage section; see, https://www.dsm.com) can be specific for residues of β-lactam or broad-spectrum antibiotics. Considerable concerns regarding antibiotics in the

**Quality tests Acceptable limits Standards Reference number**

The Dairy Industry: Process, Monitoring, Standards, and Quality

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Acidity (Titratable) ≤0.18% ISO 6091:2010 [19] Antibiotic residues Absent/0.1 g ISO 26844:2006c [20] Freezing point (added water) −0.54°C ISO 5764:2009 [21] Fat 0.8% ISO 1736:2008 [22] Protein 34% ISO 8968–1/2:2014 and ISO 14891:2002 [23, 24] Lactose >4.2% ISO 22662:2007 [25]

**Figure 1.** Milk processing stages.

**Table 2.** Quality analytical tests for raw milk.

The Dairy Industry: Process, Monitoring, Standards, and Quality http://dx.doi.org/10.5772/intechopen.80398 7

**Figure 1.** Milk processing stages.

medicines resulting in drug resistance and anthropogenic (synthetic) chemicals with the potential for transmission of chemical residues into the food chain. The more frequent and intense rainfall that is predicted could encourage the spread of perchlorate through surface runoff with the potential to enter the food chain via cow's milk [16]. Perchlorate reduces

The flow diagram for milk processing is presented in **Figure 1**. Milk arrives at the milk dairy processing plant over the weighbridge and the weight of milk is automatically recorded. At the same time, data from an on-board computer is downloaded wirelessly to a data capture system, which holds the records of the temperature and volumes of milk collected from each farm. The temperature should be at 4–6°C. Milk samples using sterile containers are collected automatically from each supplier at source and are delivered to a laboratory technician for detailed analysis. Milk that deviates in composition, taste, and smell from normal milk receives a lower quality rating. The technician also takes a composite sample, from each compartment in the refrigerated truck, which is compartmentalized to reduce sloshing of the milk. The samples from each compartment are tested for acidity, antibiotics, added water, fat, and protein content. These analytical tests and methods are determined by international

The ISO standards catalog ISO/TC34/SC5 [18] lists all milk and milk products standards, while other standard sets include, microbiology of the food chain, microbiological quality of milk, etc. The bacterial quality of the milk is also measured and these specify tests are outlined later. The titratable acid test measures the acidity of the milk. Both titratable acidity (TA) and pH are measures of acid. TA is a more reliable indicator because relative to pH measurement, it is more sensitive to small changes in milk acidity, especially important in cheese making. The acidity of milk is of two types; natural acidity due to citrates and phosphates present in the

due to lactic acid produced by bacteria using the lactose in the milk as a nutrient, converting it to lactic acid. The acidity of milk measures the total acidity (natural acidity of milk and developed acidity). The International Standard Method for titratable acid is ISO 6091:2010 [19]. Titratable acidity is a measure of the buffering of milk between pH 6.6 and 8.3 (phenolphthalein endpoint) [26]. The appearance of a faint pink color, which signals the endpoint and the number of ml of NaOH used to reach the endpoint, is recorded. This value is called the "titer," titratable acidity is reported as percent lactic acid and is dependent on the volume of sample. As this test is dependent on the analyst reading eye measurement of the color change, it is

The antibiotic test uses kits known as Charm and Delvo tests. The Charm test is made by Charm Science Inc., e.g., one kit, the Charm Rosa TET–SL (www.charm.com), detects chlortetracycline, oxytetracycline, and tetracycline residues in raw milk in the initial assay at or below

prone to human error causing incorrect and unpredictable recording of results.

during the processing of milking. The second is the developed acidity

thyroid hormone production in the thyroid gland [17].

**2. Milk processing**

6 Descriptive Food Science

standards as outlined in **Table 2**.

milk and dissolved CO2


**Table 2.** Quality analytical tests for raw milk.

100 ppb, which complies with the EU regulation No 2377/90 [27], ISO 26844:2006 [20], and Codex Alimentarius regulations CAC/MRL 2-2015 [28]. The Delvo test (a rapid kit developed by DSM, food and beverage section; see, https://www.dsm.com) can be specific for residues of β-lactam or broad-spectrum antibiotics. Considerable concerns regarding antibiotics in the food chain and antibiotic resistance transfer to human has been postulated, but in most cases, there is not sufficient evidence to demonstrate this conclusively [29].

previously. The determination of protein content of milk and milk products underpins the international trade in dairy products. There are different analytical approaches for the determination of protein quality for nutrition purposes and chemically defined protein. These are divided into three broad categories: (i) determination of total nitrogen, (ii) direct protein determination, and (iii) indirect protein determination [40]. The Kjeldahl method ISO 8968/1:2014 [23] and Dumas method ISO 14891:2002 [24] are the current international standards, and use chemical digestion and combustion approaches respectfully. The advantage of these methods is that they have high reliability and accuracy. Using these methods, around 95% of nitrogen in milk is found to be present as proteins, with the remainder as nonprotein nitrogen sources such as urea. Together these tests and values form the basis for testing the quality of milk and

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The raw milk in the milk container truck, having passed the preliminary analytical tests, proceeds to whole milk intake bays and the milk hoses are connected up by the driver. The milk is pumped into bulk storage tanks called milk silos (capacity can be up to 300,000 l, plus). The driver enters the trucks identification number on the pump's control panel or uses a key fob (a passive wireless electronic device that usually uses radio frequency ID technology) to start pumping into the whole milk silos. Unloaded milk is cooled automatically to 4–6°C with a heat plate exchanger (HPE) while pumped into the silo. The offload time and setup time taken to couple and decouple the milk intake hoses are areas where processing monitoring can be implemented. The pumping time can be variable, indicating performance specific to each pump and the flow rate represents a reasonable performance indicator. Other significant factors that can influence pumping time include the volume of milk in the receiving silo, the number of bends and valves in each pipeline, and the associated backpressure variations. At milk offload, process optimization can be achieved by ensuring pumps are working effectively, efficiently, and planning truck supply due to

Different milk processing plants have their own process trains. In many cases, milk must be clarified on reception at the dairy, to remove particles of dirt such as sand, soil, dust, and precipitated protein, which will protect downstream processing equipment. In addition, removal of bacteria, spores, and somatic cells from milk can be achieved with centrifugation and microfiltration techniques [41]. Somatic cells such as leucocytes are removed, which will reduce the presence of Listeria trapped inside the leucocyte [42]. Reduction in the microbial load at this point can decrease the burden of biofilms [43], which leads to more efficient work of the HPE [44]. Milk bacterial clarification also avoids problems during cheese aging, and improves shelf life and organoleptic properties of the dairy products. A clarifier is a type of centrifugal separator, but clarifiers and milk separators serve slightly different duties. All

milk products.

**3. Dairy processing stages**

intelligent time slot management.

**3.1. Separation, clarification, and centrifugation**

Added water can be measured by changes in the freezing point of milk from its normal values, the current official freezing point limit is −0.525° Horvet or −0.505°C and was designed for whole-herd, bulk-tank samples, or processed milk samples. The freezing point of milk is the constant physical-chemical property of milk, which is determined only by its watersoluble components such as lactose, and salts, which in accordance with the Wigner law are held in milk at an approximately constant concentration. However, the mineral composition of milk depends on lactation, nutritional status of the animal, and environmental and genetic factors [30].

Adulteration of milk with water will cause a measureable rise of the freezing point of milk. The freezing point is also lowered by acidification of milk, which leads to protein denaturation. The freezing point is considered as an accurate and sensitive method, most laboratories use a cryoscopy, method that is the ISO reference method ISO 5764:2009 [21].

The average fat content of raw milk is ~4.4 g of milk fat per 100 g; with more than 400 various fatty acids (FA) being present in milk [31]. The milk fatty acids are derived almost equally from two sources, the feed and the microbial activity in the rumen of the cow [32]. A study of Swedish bovine milk found that the milk contained substantial quantities of unsaturated fatty acids with 4–10 carbon chains (C4:0–C10:0), about 2% each of saturated C18:2 and trans-C18:1, and almost no other long-chain polyunsaturated fatty acids. The most important fatty acid from a quantitative viewpoint was palmitic acid (C16:0), which accounted for approximately 30% by weight of the total fatty acids. Myristic acid (C14:0) and stearic acid (C18:0), made up 11 and 12% by weight, respectively [31]. Fatty acid composition can show rapid and significant variation in response to changes in diet. The ISO standard for fat determination is ISO 1736:2008 [22].

The fatty acid make-up of the milk can be altered by changes in diet [33], but are also affected by a number of factors, including diet composition, nutrient utilization, lactation cycle, breed of cows, with dietary variations changes up to 3% units, been reported [34]. Specific fatty acids produced during microbial fermentation of dietary fats in the rumen of the cows are responsible for low milk fat. 2–3 g of these fatty acids can decrease milk fat by 0.5% or more [35]. The Gerber method is a historic method still used today to find the fat content of milk in particular in milk powders. By using specific butyrometers designed especially for the different dairy products, e.g., for cream, ice cream, whole milk, or cheese butyrometers, with method modifications [36]. There are many suppliers of such analytical tools, e.g., Gerber instruments (http://www.gerber-instruments.com) or Brouwland instruments (https://www. brouwland.com). Infrared analysis (IR) is a commonly used method for the examination of milk and its components such as solid fat, milk proteins, or carbohydrates [37, 38]. An ISO standard 9622:2013 is available for IR applications for milk and milk products [39].

The protein fat and lactose content of milk has a bearing on the price the farmer achieves for its milk. Liquid milk contains around 3.4% protein. The proteins in milk were described previously. The determination of protein content of milk and milk products underpins the international trade in dairy products. There are different analytical approaches for the determination of protein quality for nutrition purposes and chemically defined protein. These are divided into three broad categories: (i) determination of total nitrogen, (ii) direct protein determination, and (iii) indirect protein determination [40]. The Kjeldahl method ISO 8968/1:2014 [23] and Dumas method ISO 14891:2002 [24] are the current international standards, and use chemical digestion and combustion approaches respectfully. The advantage of these methods is that they have high reliability and accuracy. Using these methods, around 95% of nitrogen in milk is found to be present as proteins, with the remainder as nonprotein nitrogen sources such as urea. Together these tests and values form the basis for testing the quality of milk and milk products.

## **3. Dairy processing stages**

food chain and antibiotic resistance transfer to human has been postulated, but in most cases,

Added water can be measured by changes in the freezing point of milk from its normal values, the current official freezing point limit is −0.525° Horvet or −0.505°C and was designed for whole-herd, bulk-tank samples, or processed milk samples. The freezing point of milk is the constant physical-chemical property of milk, which is determined only by its watersoluble components such as lactose, and salts, which in accordance with the Wigner law are held in milk at an approximately constant concentration. However, the mineral composition of milk depends on lactation, nutritional status of the animal, and environmental and genetic

Adulteration of milk with water will cause a measureable rise of the freezing point of milk. The freezing point is also lowered by acidification of milk, which leads to protein denaturation. The freezing point is considered as an accurate and sensitive method, most laboratories

The average fat content of raw milk is ~4.4 g of milk fat per 100 g; with more than 400 various fatty acids (FA) being present in milk [31]. The milk fatty acids are derived almost equally from two sources, the feed and the microbial activity in the rumen of the cow [32]. A study of Swedish bovine milk found that the milk contained substantial quantities of unsaturated fatty acids with 4–10 carbon chains (C4:0–C10:0), about 2% each of saturated C18:2 and trans-C18:1, and almost no other long-chain polyunsaturated fatty acids. The most important fatty acid from a quantitative viewpoint was palmitic acid (C16:0), which accounted for approximately 30% by weight of the total fatty acids. Myristic acid (C14:0) and stearic acid (C18:0), made up 11 and 12% by weight, respectively [31]. Fatty acid composition can show rapid and significant variation in response to changes in diet. The ISO standard for fat determination is ISO

The fatty acid make-up of the milk can be altered by changes in diet [33], but are also affected by a number of factors, including diet composition, nutrient utilization, lactation cycle, breed of cows, with dietary variations changes up to 3% units, been reported [34]. Specific fatty acids produced during microbial fermentation of dietary fats in the rumen of the cows are responsible for low milk fat. 2–3 g of these fatty acids can decrease milk fat by 0.5% or more [35]. The Gerber method is a historic method still used today to find the fat content of milk in particular in milk powders. By using specific butyrometers designed especially for the different dairy products, e.g., for cream, ice cream, whole milk, or cheese butyrometers, with method modifications [36]. There are many suppliers of such analytical tools, e.g., Gerber instruments (http://www.gerber-instruments.com) or Brouwland instruments (https://www. brouwland.com). Infrared analysis (IR) is a commonly used method for the examination of milk and its components such as solid fat, milk proteins, or carbohydrates [37, 38]. An ISO

standard 9622:2013 is available for IR applications for milk and milk products [39].

The protein fat and lactose content of milk has a bearing on the price the farmer achieves for its milk. Liquid milk contains around 3.4% protein. The proteins in milk were described

use a cryoscopy, method that is the ISO reference method ISO 5764:2009 [21].

there is not sufficient evidence to demonstrate this conclusively [29].

factors [30].

8 Descriptive Food Science

1736:2008 [22].

The raw milk in the milk container truck, having passed the preliminary analytical tests, proceeds to whole milk intake bays and the milk hoses are connected up by the driver. The milk is pumped into bulk storage tanks called milk silos (capacity can be up to 300,000 l, plus). The driver enters the trucks identification number on the pump's control panel or uses a key fob (a passive wireless electronic device that usually uses radio frequency ID technology) to start pumping into the whole milk silos. Unloaded milk is cooled automatically to 4–6°C with a heat plate exchanger (HPE) while pumped into the silo. The offload time and setup time taken to couple and decouple the milk intake hoses are areas where processing monitoring can be implemented. The pumping time can be variable, indicating performance specific to each pump and the flow rate represents a reasonable performance indicator. Other significant factors that can influence pumping time include the volume of milk in the receiving silo, the number of bends and valves in each pipeline, and the associated backpressure variations. At milk offload, process optimization can be achieved by ensuring pumps are working effectively, efficiently, and planning truck supply due to intelligent time slot management.

#### **3.1. Separation, clarification, and centrifugation**

Different milk processing plants have their own process trains. In many cases, milk must be clarified on reception at the dairy, to remove particles of dirt such as sand, soil, dust, and precipitated protein, which will protect downstream processing equipment. In addition, removal of bacteria, spores, and somatic cells from milk can be achieved with centrifugation and microfiltration techniques [41]. Somatic cells such as leucocytes are removed, which will reduce the presence of Listeria trapped inside the leucocyte [42]. Reduction in the microbial load at this point can decrease the burden of biofilms [43], which leads to more efficient work of the HPE [44]. Milk bacterial clarification also avoids problems during cheese aging, and improves shelf life and organoleptic properties of the dairy products. A clarifier is a type of centrifugal separator, but clarifiers and milk separators serve slightly different duties. All centrifuges can act as clarifiers; however, in general, only centrifuges with a high hydraulic capacity are used in this way. The clarifier can function with either cold (below 8°C) or hot milk (50–60°C).

The EU Center for Disease Control and Infection (ECDC) reported Listeriosis cases of 2536 in 2016, of which *L. monocytogenes* was most frequently detected in both soft and semi-soft cheeses prepared from raw milk (2.5%), while 0.7% of raw milk (*n* = 968) samples tested positive [52]. These pathogens can also be found in multiple food products including meat [53]. However, postpasteurization contamination has been found to be the most causative factor in

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The PasLite test is an internationally accepted method used by dairies and food manufacturers to verify pasteurization for many types of dairy products. The PasLite test verifies the completeness of milk pasteurization by detecting alkaline phosphatase, a natural enzyme in milk that is destroyed by the heat and hold time of pasteurization. The test takes 3 min and multiple samples can be run simultaneously, however only one sample can be read at

When a dairy sample is mixed with PasLite reagents and incubated, the resulting solution emits light in an amount directly proportional to the phosphatase enzyme present. The Charm nova LUM ATP detection system is used to measure the light emitted and coverts light readings to enzyme units. Phosphatase readings greater than 350 mU/L indicate product pasteurization issues, according to US and EU pasteurization requirements. The PasLite test detection limit for liquid dairy products is 20 milliunits per liter (mU/L) phosphatase (~0.002% raw milk). This is much lower than the 350 mU/L level (0.1% raw milk) mandated by nearly

The development of milk powders has revolutionized the dairy industry and allowed for a highly nutritional foodstuff to be exported safely around the world. Milk contains 85–90% water (**Table 1**); it is reduced by removing the water and can reduce the milk weight to 12% w/v, allowed for cheaper and easier transport. History tells us that in the thirteenth century, Marco Polo reported that soldiers of Kublai Khan carried sun-dried milk on their expeditions [57]. In 2013, the world's largest dairy spray dryer was installed by Fonterra Dairy Co-Op in New Zealand that has a capacity to produce 30 tons of milk powder per hour, converting four and a half million liters of fresh milk each day [58]. Three years later, in 2016, a "second of its kind," world's largest spray dryer started production at another Fonterra milk powder plant, which

Milk powders can include whole milk powder (WMP), skim milk powder (SMP), fat filled milk powder (FFMP), infant formula, and milk protein concentrate, which is 85% pure milk protein. Its uses include in bakery, confectionary, ice cream, and in fermented food such as yogurt. Many are advertised as nutritional supplements and are fortified with vitamins, folic

illustrates that, the trend is toward maximized production of dairy powders.

microbial outbreaks due to milk products [54, 55].

**3.3. Verifying the pasteurization process**

a time [56].

all public health agencies.

**4. Milk powders**

acid, and iron.

The main use for centrifuges in diary processing plants is hot milk separation. The aim is to separate the globular milk fat from the serum, the skim milk. This process is known as skimming. This process is generally combined into the pasteurization line and joined with an in-line fat standardization system for both milk and cream. Separation normally takes place at 122–140°F (50–60°C). The fat content of the cream discharged from the separator can be controlled to a level of between 20 and 70%. The terminology for separation in the dairy industry includes continuous centrifugal separation of solid particles (Clarifier), separation of cream (Separator), or separation of bacteria (Bactofuge). The microbial quality of milk powders is highly significant and it is possible at this early phase of processing to remove 99.9% of the spore-forming bacteria by either bacto-fugation or microfiltration preceding heat treatment.

Standardization of milk is the alteration of fat and solids-not-fat (SNF) levels, i.e., raising or lowering of these levels. This is regularly carried out for the consumer market milk supply and in the production of other milk products including: condensed milk, milk powder, ice cream and cheese, etc. Standardization is typically carried out to create a uniform milk fat content in the final dairy product [45].

#### **3.2. Pasteurization**

Pasteurization was originally introduced to control *Mycobacterium bovis*, which causes tuberculosis (TB), which is no longer problematic as cows are tested for TB annually and removed from herds if they test positive for the disease [46]. The TB bacillus is a highly heat resistant microorganism; however, *Coxiella burnetii*, the cause of Q fever in humans [47], required pasteurization of 161°F (71.7°C) for 15 s, and is the current official standard for milk pasteurization [48], the standard vat pasteurization is 63°C (145°F) for 30 min. However, heat processing can result in the loss of subtle aroma and flavors components, loss of vitamins and natural antioxidants, the loss of texture and freshness, and the denaturation of proteins. The US Grade A pasteurization milk ordinance (PMO) is managed by the Departments of Health and Human Services and Public Health, and the Food and Drug Administration and gives the criteria concerning the milk parlor and processing plant design, milking practices, milk handling, sanitation, and standards for the pasteurization of Grade A milk products. Regulation of milk processing is controlled on each US state basis; however, all dairy products must meet the regulations outlined in the PMO for products that will be sold outside of that state [10].

The center for disease control (CDC) in the US, reported that unpasteurized milk is 150 times more likely to cause foodborne illness and results in 13 times more hospitalizations than illnesses involving pasteurized dairy products [49]. The dangerous bacteria include *Salmonella* spp., *Escherichia coli*, and *Listeria monocytogenes*; it is also for this reason milk is pasteurized. *E coli* 0157 emerged in 1982, while multidrug-resistant *Salmonella typhimurium* DT104 was reported in 1990 [50, 51] with some *E. coli* and *Salmonella* isolates resistant to seven antibiotics. The EU Center for Disease Control and Infection (ECDC) reported Listeriosis cases of 2536 in 2016, of which *L. monocytogenes* was most frequently detected in both soft and semi-soft cheeses prepared from raw milk (2.5%), while 0.7% of raw milk (*n* = 968) samples tested positive [52]. These pathogens can also be found in multiple food products including meat [53]. However, postpasteurization contamination has been found to be the most causative factor in microbial outbreaks due to milk products [54, 55].

#### **3.3. Verifying the pasteurization process**

centrifuges can act as clarifiers; however, in general, only centrifuges with a high hydraulic capacity are used in this way. The clarifier can function with either cold (below 8°C) or hot

The main use for centrifuges in diary processing plants is hot milk separation. The aim is to separate the globular milk fat from the serum, the skim milk. This process is known as skimming. This process is generally combined into the pasteurization line and joined with an in-line fat standardization system for both milk and cream. Separation normally takes place at 122–140°F (50–60°C). The fat content of the cream discharged from the separator can be controlled to a level of between 20 and 70%. The terminology for separation in the dairy industry includes continuous centrifugal separation of solid particles (Clarifier), separation of cream (Separator), or separation of bacteria (Bactofuge). The microbial quality of milk powders is highly significant and it is possible at this early phase of processing to remove 99.9% of the spore-forming bacteria by either bacto-fugation or microfiltration preceding heat treatment. Standardization of milk is the alteration of fat and solids-not-fat (SNF) levels, i.e., raising or lowering of these levels. This is regularly carried out for the consumer market milk supply and in the production of other milk products including: condensed milk, milk powder, ice cream and cheese, etc. Standardization is typically carried out to create a uniform milk fat

Pasteurization was originally introduced to control *Mycobacterium bovis*, which causes tuberculosis (TB), which is no longer problematic as cows are tested for TB annually and removed from herds if they test positive for the disease [46]. The TB bacillus is a highly heat resistant microorganism; however, *Coxiella burnetii*, the cause of Q fever in humans [47], required pasteurization of 161°F (71.7°C) for 15 s, and is the current official standard for milk pasteurization [48], the standard vat pasteurization is 63°C (145°F) for 30 min. However, heat processing can result in the loss of subtle aroma and flavors components, loss of vitamins and natural antioxidants, the loss of texture and freshness, and the denaturation of proteins. The US Grade A pasteurization milk ordinance (PMO) is managed by the Departments of Health and Human Services and Public Health, and the Food and Drug Administration and gives the criteria concerning the milk parlor and processing plant design, milking practices, milk handling, sanitation, and standards for the pasteurization of Grade A milk products. Regulation of milk processing is controlled on each US state basis; however, all dairy products must meet the regulations outlined in the PMO for products that will be sold outside of

The center for disease control (CDC) in the US, reported that unpasteurized milk is 150 times more likely to cause foodborne illness and results in 13 times more hospitalizations than illnesses involving pasteurized dairy products [49]. The dangerous bacteria include *Salmonella* spp., *Escherichia coli*, and *Listeria monocytogenes*; it is also for this reason milk is pasteurized. *E coli* 0157 emerged in 1982, while multidrug-resistant *Salmonella typhimurium* DT104 was reported in 1990 [50, 51] with some *E. coli* and *Salmonella* isolates resistant to seven antibiotics.

milk (50–60°C).

10 Descriptive Food Science

content in the final dairy product [45].

**3.2. Pasteurization**

that state [10].

The PasLite test is an internationally accepted method used by dairies and food manufacturers to verify pasteurization for many types of dairy products. The PasLite test verifies the completeness of milk pasteurization by detecting alkaline phosphatase, a natural enzyme in milk that is destroyed by the heat and hold time of pasteurization. The test takes 3 min and multiple samples can be run simultaneously, however only one sample can be read at a time [56].

When a dairy sample is mixed with PasLite reagents and incubated, the resulting solution emits light in an amount directly proportional to the phosphatase enzyme present. The Charm nova LUM ATP detection system is used to measure the light emitted and coverts light readings to enzyme units. Phosphatase readings greater than 350 mU/L indicate product pasteurization issues, according to US and EU pasteurization requirements. The PasLite test detection limit for liquid dairy products is 20 milliunits per liter (mU/L) phosphatase (~0.002% raw milk). This is much lower than the 350 mU/L level (0.1% raw milk) mandated by nearly all public health agencies.

### **4. Milk powders**

The development of milk powders has revolutionized the dairy industry and allowed for a highly nutritional foodstuff to be exported safely around the world. Milk contains 85–90% water (**Table 1**); it is reduced by removing the water and can reduce the milk weight to 12% w/v, allowed for cheaper and easier transport. History tells us that in the thirteenth century, Marco Polo reported that soldiers of Kublai Khan carried sun-dried milk on their expeditions [57]. In 2013, the world's largest dairy spray dryer was installed by Fonterra Dairy Co-Op in New Zealand that has a capacity to produce 30 tons of milk powder per hour, converting four and a half million liters of fresh milk each day [58]. Three years later, in 2016, a "second of its kind," world's largest spray dryer started production at another Fonterra milk powder plant, which illustrates that, the trend is toward maximized production of dairy powders.

Milk powders can include whole milk powder (WMP), skim milk powder (SMP), fat filled milk powder (FFMP), infant formula, and milk protein concentrate, which is 85% pure milk protein. Its uses include in bakery, confectionary, ice cream, and in fermented food such as yogurt. Many are advertised as nutritional supplements and are fortified with vitamins, folic acid, and iron.

#### **4.1. Evaporation and drying**

Milk powder is manufactured by spray-drying precondensed milk. A falling film evaporator is commonly used in the dairy industry to concentrate the milk from ~13% total solids (TS) to a target of up to 52%. Evaporation is simply the removal of a solvent from a solution or slurry. Milk itself is defined as a colloid with the solvent being the water. Other methods of removing water can include freeze-drying [59]. The constituents of milk can be seen in **Table 1**. As some products are sensitive to heat, the design of evaporators with respect to temperature and holding time is vital in order to achieve the desired effects on the one hand, but without causing heat damage and denaturation to the milk proteins. To minimize the thermal impact on the products from the heat applied, evaporation takes place in a vacuum at pressures of 160–320 hPa, equivalent to water boiling temperatures of 55–70°C. Energy efficiency is the main driving force in improved design and technologies in evaporation [60, 61]. Inside the evaporator are a bundle of tubes for the exchange of heat and these are enclosed in another steel cylinder, in evaporation parlance called a "calandria." The vaporized solvent is cooled to condensate, which is then removed. It can go to storage, be recirculated, recovered for heat transfer, or filtrated but this is secondary to the evaporation process itself. The main unit of an evaporator is called an "effect." Generally, more than one "effect" is used, to increase efficiency by using the heat from the vapor from the previous "effect" to heat the feed in the next. Steam economy is a term used to quantify how much original steam is used in ratio to vapor steam. If 1 Kg of steam produces 1 Kg of vapor in a single "effect" system, 1 Kg of steam will produce ~2 Kg of vapor in a two "effect" system. The specific steam consumption of the former is 100%, while it is 50% in the latter case. This cuts down the cost of generating original steam feed. A subsequent "effect" must have a lower pressure than the previous "effect," and a step-wise vacuum is applied to the whole evaporation process to achieve this. There are three main elements in evaporation: heat transfer, vapor-liquid separation, and energy efficiency [62].

Milk powders can be classified accordingly to the heat treatment they receive. There are five levels of heat classification: ultra-low (<70°C/15 s) common low (70°C/15 s), medium (85–90°C/20–30 s), high (110–135°C/30 s), and high-heat stable (~135°C/30 s). The whey protein nitrogen index (WPNI) expresses the content of un-denatured whey protein (mg WPNI per gram of powder) and demonstrates the severity of the heat treatment. Low-heat WPNI >6.0 mg, while high-heat WPNI is <1.5 mg, values are expected [65]. An alternative heat classification of milk powder is by casein number (CN—total nitrogen precipitated at pH 4.7), this measure was introduced as the protein concentration in milk changes with the seasons and feeding patterns [66]. The CN number is not linked to the overall protein content of the milk. The CN value of high quality raw milk is in the range 80–82, expressed in percentages. The CN values in excess of 82 indicate that the denaturation of whey protein has taken place.

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The composition of and additives allowed in milk powders are regulated by the Codex Alimentarius Commission—Milk and Milk Products [67, 68]. The Codex standard stipulates that only milk and cream may be allowed in milk powders; though the protein content can be altered by adding lactose. Milk proteins include casein complexes and whey protein fractions. Casein is the most abundant with whey proteins in lower concentrations. The casein concentration in cow milk is 2.46–2.80/100 g and whey proteins in the range 0.55–0.70/100 g. The composition of milks from various animal species is well reviewed in Barłowska et al. [69].

The moisture content of milk powder must be controlled during milk processing, as it is a factor in the long-term quality of the product, and it influences the cost of production. The method for determination of the moisture content includes the ISO 5537: 2004 reference method [70] and IDF Provisional Standard 26A:1993 [71] and EU commission Directive method (79/1067) [72]. A test portion of milk powder is dried at 102 ± 2°C until constant mass is obtained, but this measurement can be affected by the relative humidity of the air in the laboratory where the test is carried out. Rapid methods and newly designed equipment are always being introduced to avoid air humidity interference in the measurement and one new

Each step along the milk processing train can be contaminate by the air [74] and the water [75], used in the milk processing stages. Hygiene control at all stages, including hygienic

The microbial quality of milk starts a farm level. Milk is sterile at secretion in the udder but is colonized by bacteria before it leaves the udder [76]. The temperature of milk expelled from the udder is approximately 35°C; to prevent microbial growth, rapid cooling, and storage to 4°C is necessary. The dairy farmer has the responsibility of managing and maintaining a

method is by using a microwave cavity perturbation technique [73].

design of the manufacturing equipment, is critically important.

**5. Microbiological quality analyses**

**5.1. Microbial quality analysis at farm level**

Completely denatured milk has a CN value of 92.

When milk leaves the evaporator (**Figure 1**), it is passed through the spray dryer through small nozzles, which make small droplets or atomizing the liquid, the smaller the better. The drying chamber has a temperature of 160–205°C, the droplets are swirled around (1 l of concentrate is atomized to 1.2 × 1011 droplets with a diameter of 50 micron with a total surface of 120 m2 .). For effective drying, the air should be hot, dry (low humidity) and moving. The powder falls to the bottom where it is collected in a "fluid" bed under the cone of the drying chamber, where fine powder behave in an analogous manner to a liquid and it can be conveyed without forming clusters. Fluid beds permit mild second stage drying and cooling of delicate products. Agglomeration changes the bulk density of the product [12].

The bulk density of the powder can dictate how the milk powder dissolves in hot beverages including for tea, coffee, and chocolate. The particle size of the milk powders determines its reconstitution properties. Powders consisting of particles of <100 μm are difficult to wet with water and form lumps [63], in the case of full-fat milk powder (FFMP), which is difficult to wet, it is sprayed with lecithin or oils (e.g., palm) to improve reconstitution characteristics. The standard method for measuring bulk density is ISO 8966:2005 [64].

Milk powders can be classified accordingly to the heat treatment they receive. There are five levels of heat classification: ultra-low (<70°C/15 s) common low (70°C/15 s), medium (85–90°C/20–30 s), high (110–135°C/30 s), and high-heat stable (~135°C/30 s). The whey protein nitrogen index (WPNI) expresses the content of un-denatured whey protein (mg WPNI per gram of powder) and demonstrates the severity of the heat treatment. Low-heat WPNI >6.0 mg, while high-heat WPNI is <1.5 mg, values are expected [65]. An alternative heat classification of milk powder is by casein number (CN—total nitrogen precipitated at pH 4.7), this measure was introduced as the protein concentration in milk changes with the seasons and feeding patterns [66]. The CN number is not linked to the overall protein content of the milk. The CN value of high quality raw milk is in the range 80–82, expressed in percentages. The CN values in excess of 82 indicate that the denaturation of whey protein has taken place. Completely denatured milk has a CN value of 92.

The composition of and additives allowed in milk powders are regulated by the Codex Alimentarius Commission—Milk and Milk Products [67, 68]. The Codex standard stipulates that only milk and cream may be allowed in milk powders; though the protein content can be altered by adding lactose. Milk proteins include casein complexes and whey protein fractions. Casein is the most abundant with whey proteins in lower concentrations. The casein concentration in cow milk is 2.46–2.80/100 g and whey proteins in the range 0.55–0.70/100 g. The composition of milks from various animal species is well reviewed in Barłowska et al. [69].

The moisture content of milk powder must be controlled during milk processing, as it is a factor in the long-term quality of the product, and it influences the cost of production. The method for determination of the moisture content includes the ISO 5537: 2004 reference method [70] and IDF Provisional Standard 26A:1993 [71] and EU commission Directive method (79/1067) [72]. A test portion of milk powder is dried at 102 ± 2°C until constant mass is obtained, but this measurement can be affected by the relative humidity of the air in the laboratory where the test is carried out. Rapid methods and newly designed equipment are always being introduced to avoid air humidity interference in the measurement and one new method is by using a microwave cavity perturbation technique [73].

## **5. Microbiological quality analyses**

**4.1. Evaporation and drying**

12 Descriptive Food Science

efficiency [62].

of 120 m2

Milk powder is manufactured by spray-drying precondensed milk. A falling film evaporator is commonly used in the dairy industry to concentrate the milk from ~13% total solids (TS) to a target of up to 52%. Evaporation is simply the removal of a solvent from a solution or slurry. Milk itself is defined as a colloid with the solvent being the water. Other methods of removing water can include freeze-drying [59]. The constituents of milk can be seen in **Table 1**. As some products are sensitive to heat, the design of evaporators with respect to temperature and holding time is vital in order to achieve the desired effects on the one hand, but without causing heat damage and denaturation to the milk proteins. To minimize the thermal impact on the products from the heat applied, evaporation takes place in a vacuum at pressures of 160–320 hPa, equivalent to water boiling temperatures of 55–70°C. Energy efficiency is the main driving force in improved design and technologies in evaporation [60, 61]. Inside the evaporator are a bundle of tubes for the exchange of heat and these are enclosed in another steel cylinder, in evaporation parlance called a "calandria." The vaporized solvent is cooled to condensate, which is then removed. It can go to storage, be recirculated, recovered for heat transfer, or filtrated but this is secondary to the evaporation process itself. The main unit of an evaporator is called an "effect." Generally, more than one "effect" is used, to increase efficiency by using the heat from the vapor from the previous "effect" to heat the feed in the next. Steam economy is a term used to quantify how much original steam is used in ratio to vapor steam. If 1 Kg of steam produces 1 Kg of vapor in a single "effect" system, 1 Kg of steam will produce ~2 Kg of vapor in a two "effect" system. The specific steam consumption of the former is 100%, while it is 50% in the latter case. This cuts down the cost of generating original steam feed. A subsequent "effect" must have a lower pressure than the previous "effect," and a step-wise vacuum is applied to the whole evaporation process to achieve this. There are three main elements in evaporation: heat transfer, vapor-liquid separation, and energy

When milk leaves the evaporator (**Figure 1**), it is passed through the spray dryer through small nozzles, which make small droplets or atomizing the liquid, the smaller the better. The drying chamber has a temperature of 160–205°C, the droplets are swirled around (1 l of concentrate is atomized to 1.2 × 1011 droplets with a diameter of 50 micron with a total surface

powder falls to the bottom where it is collected in a "fluid" bed under the cone of the drying chamber, where fine powder behave in an analogous manner to a liquid and it can be conveyed without forming clusters. Fluid beds permit mild second stage drying and cooling of

The bulk density of the powder can dictate how the milk powder dissolves in hot beverages including for tea, coffee, and chocolate. The particle size of the milk powders determines its reconstitution properties. Powders consisting of particles of <100 μm are difficult to wet with water and form lumps [63], in the case of full-fat milk powder (FFMP), which is difficult to wet, it is sprayed with lecithin or oils (e.g., palm) to improve reconstitution characteristics.

delicate products. Agglomeration changes the bulk density of the product [12].

The standard method for measuring bulk density is ISO 8966:2005 [64].

.). For effective drying, the air should be hot, dry (low humidity) and moving. The

Each step along the milk processing train can be contaminate by the air [74] and the water [75], used in the milk processing stages. Hygiene control at all stages, including hygienic design of the manufacturing equipment, is critically important.

#### **5.1. Microbial quality analysis at farm level**

The microbial quality of milk starts a farm level. Milk is sterile at secretion in the udder but is colonized by bacteria before it leaves the udder [76]. The temperature of milk expelled from the udder is approximately 35°C; to prevent microbial growth, rapid cooling, and storage to 4°C is necessary. The dairy farmer has the responsibility of managing and maintaining a clean and hygienic milking parlor with a good milking and storage routine. The farmer can detect early signs of mastitis infection by using a somatic cell count (SSC) test. Low levels of SCC (<200,000/ml) are wanted to guarantee good extraction of protein from milk. High levels of SCC also reduce other levels of milk constituent including lactose. The California Mastitis Test (CMT) offers a quick and easy on-farm test; the test does not provide a specific SCC, but will give a positive result once a cow's SCC goes over 400,000 cells/mL. The addition of the CMT solution to milk samples with a high number of leukocytes/white blood cells causes the solution to become mucous like. This reaction is caused by the release of DNA from somatic cells, which are now higher due to the immune response of the cow to infections. Mastitis is caused by the microorganism *Staphylococcus aureus.* CMT test are available commercially from many companies.

The common bacteria in milk are lactic acid bacteria (LAB), which can produce enough acid to reduce the pH of milk, and cause the coagulation of proteins, thus fermenting the milk [82]. The density test as previously described should be introduced at milk intake, as it can determine the degree of LAB growth. LAB can be divided into rods (*Lactobacillus* and

The Dairy Industry: Process, Monitoring, Standards, and Quality

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15

Psychrotrophic microorganisms are also present up to 80% in fresh collected milk, they are able to grow quickly below 7°C, and some contain heat-stable enzymes, which cause spoilage, including many Gram-negative bacteria, such as *Pseudomonas fluorescens*, *Pseudomonas fragi*, *Pseudomonas putida*, *Achromobacter*, *Aeromonas*, *Alcaligenes*, *Chromobacterium*, *Flavo-*

Thermoduric bacteria can survive pasteurization. They do this by forming spores, which can then carry over into the final product. This can cause quality defects in milk products such as decreasing the shelf life of pasteurized milk. They are represented mainly by Gram-positive bacteria, e.g., *Bacillus* and *Clostridium* spp., and the nonspore-forming genera, e.g., *Micrococcus*, *Streptococcus*, and *Corynebacterium.* Levels of greater than 1000 cfu/ml are normally the result of poor cow hygiene and milking equipment (particularly in the case of ineffective hot wash routines). Potential sources of thermoduric bacteria include silage, faces, animal bedding, and soil [84]. Thermophilic bacteria grow in milk held at raised temperatures (55°C or higher), including pasteurization, 62.8°C, they include the *Bacillus* spp. Thermophilic bacteria are monitored by standard plate count methods with incubation at 55°C [85]. However, obligate thermophiles, such as *Geobacillus stearothermophilus* and *Anoxybacillus flavithermus* tend to grow to high numbers in milk powder manufacturing plants [86]. Although these microorganisms generally are not pathogenic, there is evidence to show that they cause human diseases [87], their growth results in high bacterial numbers and their presence can be interpreted as an indicator of poor plant hygiene. Spores of *G. stearothermophilus* are also able to survive ultra-high-temperature (UHT; 134–145°C for 1–10 s) treatment [88]. A recent study outlined the prevalence of contaminated milk processing samples with spore-forming bacteria, which

increased from 23% on farm, to up to 58% post pasteurization stage [89].

The total viable count (TVC), or total bacterial count (TBC), is used to indicate the overall level of microorganism in milk; *E. coli* and *coliforms* to indicate any fecal contamination; and *Pseudomonas* spp., to indicate any nonfecal contamination. EU legislation, describing precise hygiene rules for foods from animal origins (amended in 2017) lays down comprehensive criteria for milk quality [90]. The ruling indicates that TBC in raw milk should be less than 100,000 cfu/ml; however, a TBC of less than 15,000 cfu/ml is desired. A standard to aim for is <1000 cfu/ml as milk leaves the udder; <3000 cfu/ml as milk leaves the milking machine; and <5000 cfu/ml in the bulk tank. Further contamination takes place during storage and

New technologies in the dairy industry are slowly integrating both at farm level and in the dairy processing plant. At farm level, the introduction of robotics such as automated milking parlors developed by Lely and introduced in 1992 by Delaval (Sweden). The cows enter

*Carnobacterium*) and cocci (all other genera).

*bacterium*, *Serratia*, and *Enterobacter* [83].

preprocessing activities.

**7. Industry 4.0 in the dairy industry**

The milk tanker driver can perform a few tests at the farm, but this is not often practical. The collector will also take a sample of raw milk and label it with a bar code identifier, to be brought back to the dairy processing plant. Composite samples are taken for the detection of inhibitory substances (e.g., antibiotics, antiseptics) to be tested later at the processing plant and if positive the individual suppliers samples are then analyzed.

#### **5.2. Microbial quality testing at milk intake**

At the milk intake point, the milk is tested before acceptance into the processing train. One such historic test described in 1929 [77, 78] is the Resazurin test, which determines the microbiological quality of the milk. The theory of this test is that Resazurin, a blue dye, is reduced in an oxidation-reduction reaction, as bacteria grow in the milk they use up oxygen and this can reduce the Resazurin dye to a pink color. All that is required is 10 ml of milk, 1 ml of resazurin solution (0.05%), mix well and incubate at 37°C for 2 min. The color changes from blue to mauve to purple to pink and lastly colorless and is compared to standardized color disks or measured in an instrument called a Comparator (developed by Lovibond, originally) which is a short path length instrument (up to 40 mm) for visually matching samples with relatively dark colors. A reading of ≥4, which is comparable to an estimate of a total bacterial count of 0.1–2 million cfu/ml, is a satisfactory milk quality result.

The milk density is another rapid test to determine adulteration of the milk and an indication for the deviations from the normal milk composition, for example, if it has been watered down or skimmed. In this test, a dipping lactodensimeter combined with a thermometer is used (Gerber instruments; Brouwland instruments), lactometers/milk hydrometers are calibrated in either grams per milliliter (g/cm3 ), degrees specific gravity (SG), or Degrees Quevenne. 1° Quevenne = 0.001°SG. Density ranges for standard milk are between 1.026 and 1.034 g/cm3 . The adding of 10% water to milk will end up decreasing milk density by ~0.003 g/cm3 .

## **6. Microorganisms and milk**

A wide variety of bacteria grow and survive in milk, including problematic spore-forming bacteria [79] and pathogens such as nontyphoid *Salmonella*, *Campylobacter*, *Listeria monocytogenes,* and Shiga toxin-producing *Escherichia coli* are also found [80]. In addition, *Cronobacter sakazakii* has been found in milk powder producing plants and is a particular risk to infants [81].

The common bacteria in milk are lactic acid bacteria (LAB), which can produce enough acid to reduce the pH of milk, and cause the coagulation of proteins, thus fermenting the milk [82]. The density test as previously described should be introduced at milk intake, as it can determine the degree of LAB growth. LAB can be divided into rods (*Lactobacillus* and *Carnobacterium*) and cocci (all other genera).

Psychrotrophic microorganisms are also present up to 80% in fresh collected milk, they are able to grow quickly below 7°C, and some contain heat-stable enzymes, which cause spoilage, including many Gram-negative bacteria, such as *Pseudomonas fluorescens*, *Pseudomonas fragi*, *Pseudomonas putida*, *Achromobacter*, *Aeromonas*, *Alcaligenes*, *Chromobacterium*, *Flavobacterium*, *Serratia*, and *Enterobacter* [83].

Thermoduric bacteria can survive pasteurization. They do this by forming spores, which can then carry over into the final product. This can cause quality defects in milk products such as decreasing the shelf life of pasteurized milk. They are represented mainly by Gram-positive bacteria, e.g., *Bacillus* and *Clostridium* spp., and the nonspore-forming genera, e.g., *Micrococcus*, *Streptococcus*, and *Corynebacterium.* Levels of greater than 1000 cfu/ml are normally the result of poor cow hygiene and milking equipment (particularly in the case of ineffective hot wash routines). Potential sources of thermoduric bacteria include silage, faces, animal bedding, and soil [84]. Thermophilic bacteria grow in milk held at raised temperatures (55°C or higher), including pasteurization, 62.8°C, they include the *Bacillus* spp. Thermophilic bacteria are monitored by standard plate count methods with incubation at 55°C [85]. However, obligate thermophiles, such as *Geobacillus stearothermophilus* and *Anoxybacillus flavithermus* tend to grow to high numbers in milk powder manufacturing plants [86]. Although these microorganisms generally are not pathogenic, there is evidence to show that they cause human diseases [87], their growth results in high bacterial numbers and their presence can be interpreted as an indicator of poor plant hygiene. Spores of *G. stearothermophilus* are also able to survive ultra-high-temperature (UHT; 134–145°C for 1–10 s) treatment [88]. A recent study outlined the prevalence of contaminated milk processing samples with spore-forming bacteria, which increased from 23% on farm, to up to 58% post pasteurization stage [89].

The total viable count (TVC), or total bacterial count (TBC), is used to indicate the overall level of microorganism in milk; *E. coli* and *coliforms* to indicate any fecal contamination; and *Pseudomonas* spp., to indicate any nonfecal contamination. EU legislation, describing precise hygiene rules for foods from animal origins (amended in 2017) lays down comprehensive criteria for milk quality [90]. The ruling indicates that TBC in raw milk should be less than 100,000 cfu/ml; however, a TBC of less than 15,000 cfu/ml is desired. A standard to aim for is <1000 cfu/ml as milk leaves the udder; <3000 cfu/ml as milk leaves the milking machine; and <5000 cfu/ml in the bulk tank. Further contamination takes place during storage and preprocessing activities.

#### **7. Industry 4.0 in the dairy industry**

clean and hygienic milking parlor with a good milking and storage routine. The farmer can detect early signs of mastitis infection by using a somatic cell count (SSC) test. Low levels of SCC (<200,000/ml) are wanted to guarantee good extraction of protein from milk. High levels of SCC also reduce other levels of milk constituent including lactose. The California Mastitis Test (CMT) offers a quick and easy on-farm test; the test does not provide a specific SCC, but will give a positive result once a cow's SCC goes over 400,000 cells/mL. The addition of the CMT solution to milk samples with a high number of leukocytes/white blood cells causes the solution to become mucous like. This reaction is caused by the release of DNA from somatic cells, which are now higher due to the immune response of the cow to infections. Mastitis is caused by the microorganism *Staphylococcus aureus.* CMT test are available commercially from

The milk tanker driver can perform a few tests at the farm, but this is not often practical. The collector will also take a sample of raw milk and label it with a bar code identifier, to be brought back to the dairy processing plant. Composite samples are taken for the detection of inhibitory substances (e.g., antibiotics, antiseptics) to be tested later at the processing plant

At the milk intake point, the milk is tested before acceptance into the processing train. One such historic test described in 1929 [77, 78] is the Resazurin test, which determines the microbiological quality of the milk. The theory of this test is that Resazurin, a blue dye, is reduced in an oxidation-reduction reaction, as bacteria grow in the milk they use up oxygen and this can reduce the Resazurin dye to a pink color. All that is required is 10 ml of milk, 1 ml of resazurin solution (0.05%), mix well and incubate at 37°C for 2 min. The color changes from blue to mauve to purple to pink and lastly colorless and is compared to standardized color disks or measured in an instrument called a Comparator (developed by Lovibond, originally) which is a short path length instrument (up to 40 mm) for visually matching samples with relatively dark colors. A reading of ≥4, which is comparable to an estimate of a total bacterial count of

The milk density is another rapid test to determine adulteration of the milk and an indication for the deviations from the normal milk composition, for example, if it has been watered down or skimmed. In this test, a dipping lactodensimeter combined with a thermometer is used (Gerber instruments; Brouwland instruments), lactometers/milk hydrometers are calibrated

Quevenne = 0.001°SG. Density ranges for standard milk are between 1.026 and 1.034 g/cm3

A wide variety of bacteria grow and survive in milk, including problematic spore-forming bacteria [79] and pathogens such as nontyphoid *Salmonella*, *Campylobacter*, *Listeria monocytogenes,* and Shiga toxin-producing *Escherichia coli* are also found [80]. In addition, *Cronobacter sakazakii* has been found in milk powder producing plants and is a particular risk to infants [81].

The adding of 10% water to milk will end up decreasing milk density by ~0.003 g/cm3

), degrees specific gravity (SG), or Degrees Quevenne. 1°

.

.

and if positive the individual suppliers samples are then analyzed.

**5.2. Microbial quality testing at milk intake**

0.1–2 million cfu/ml, is a satisfactory milk quality result.

in either grams per milliliter (g/cm3

**6. Microorganisms and milk**

many companies.

14 Descriptive Food Science

New technologies in the dairy industry are slowly integrating both at farm level and in the dairy processing plant. At farm level, the introduction of robotics such as automated milking parlors developed by Lely and introduced in 1992 by Delaval (Sweden). The cows enter the parlor without prompting and some cows are milked three times a day, with increased milk product for the farmer. The tags on the cows allow for integration into the machines which collect vast amounts of data, including number of steps, chewing the curd, etc. Robotic milking machines have a life span of approx. 13 years and then required further investment. Determination of when a cow is in heat for efficient reproduction is available with MooCow developed by Dairy Master (Ireland), together with MooMonitor to guide cows in the parlor. A separate company created MooCall, a sensor attached to the cow tail, which can monitor contractions during calf birth and send a SMS message to the farmer, the sensor can determine as close as 1 h to delivery [91]. Some of the more recent analytical instruments for milk analysis that has been introduced, but are not yet standard and include: Fourier transform MIR spectroscopy for milk-based quantitative, qualitative phenotypic and genomic analysis. Flow CYTOMETRY is a well-established technique for bacteria and somatic cells counting and differentiation [92], and companies making these include: Bentley (https://bentleyinstruments.com/), Foss (https://www.fossanalytics.com/en#) and Delta (http://www.deltainstruments.com/); ELISA (enzyme linked immunosorbent assay) for milk amyloid; milk amyloidA (MAA) biomarker is an early detection method for clinical and subclinical mastitis. MAA is the only acute phase protein produced in response to bacterial infection in the udder and is therefore an immediate and direct indicates of infection [93]. PCR (polymerase chain reaction) is used for bacteria identification and differentiation [94, 95]. Gas chromatography/mass spectrometry can be used for analysis for metabolomics characterization of milk [96, 97]. Liquid chromatography/electrospray ionization and mass spectroscopy can be used for the detection of peptides and glycopeptides. ICP-MS (Induction coupled plasma mass spectroscopy) for mineral and trace element analysis. Industry 4.0 technologies can be introduced at all stages of the plant manufacturing systems, including environment monitoring [98].

Day, an animal-free milk made by using yeast and fermentation techniques to produce a product with equivalent dairy proteins (http://www.perfectdayfoods.com/). It is unlikely that nondairy products will overtake real-natural product in the short term. Milk powders are still a big business and the optimization approach to change the paradigm from inspect and reject to predict and prevent is developing increased interest for the factory of the future [101], and is an approach that cannot be ignored. Regardless of developments, a quality milk product

The Dairy Industry: Process, Monitoring, Standards, and Quality

http://dx.doi.org/10.5772/intechopen.80398

17

This work was supported by the Irish State through funding from the Technology Centre's

1 Dairy Processing Technology Centre (DPTC), University of Limerick, Limerick, Ireland

3 Microbiology Laboratory, School of Natural Sciences, University of Limerick, Limerick,

[1] Ahern J.Applying lean techniques to dairy farming. Acountance Plus. 2017;**03**(Sept):26-27 [2] FAO-UN. Food and Drug Organisation of the United Nations [Internet]. 2017. Milk and Milk Products: Price and Trade Update Price Index. Available from: http://www.fao.

[3] DeLoitte. The Global Dairy Sector—Trends and Opportunities [Internet]. 2017. Deloitte Development LLC. Available from: https://www2.deloitte.com/ie/en/pages/consumer-indu

strial-products/articles/global-dairy-trends-2017.html [Accessed: February 24, 2018]

2 Enterprise Research Centre, University of Limerick, Limerick, Ireland

, Mark Southern1,2, Paul Hogan<sup>1</sup>

, Michael P. Ryan<sup>3</sup>

must be the result. There can be no food security without food safety.

**Acknowledgements**

**Conflict of interest**

**Author details**

and Catherine C. Adley3

Niamh Burke1

Ireland

**References**

There is no conflict of interest.

program—Grant Number TC/2014/0016.

, Krzysztof A. Zacharski1

\*Address all correspondence to: catherine.adley@ul.ie

org/3/a-i8326e.pdf [Accessed: April 3, 2018]

\*

## **8. Conclusion**

The milk processing chain demands accurate and quality products from farm to plate and for all of its products, e.g., fluid milk, milk powders, etc. It must start with the raw material at farm level including; dairy herd improvement testing, to payment parameters, and quality control of the raw milk. Optimization is important in the processing of milk in the dairy chain as 73 plus tests are carried out including chemical physical and microbiological tests, set against ISO standards, EU, USFDA regulations, and most countries internal regulations. Advances are slowly being made to have modern and optimized methodologies approved. The regulatory bodies are setting new standards from verified inter-laboratory studies, targeting the advancement in instrumentation and for at-line and in-line production analysis for improved predictability and control of manufacturing processes. The finished product must be safe and comply with regulatory requirements.

At a conference in Glasgow (Semex Dairy Conference, Jan 2018), it was questioned whether the dairy industry could cease to exist after approximately 10 years, due to the interest in vegan alternatives and the increased population who are lactose intolerant [99, 100]. A business model to address this alternative has resulted in a cow-free milk product called Perfect Day, an animal-free milk made by using yeast and fermentation techniques to produce a product with equivalent dairy proteins (http://www.perfectdayfoods.com/). It is unlikely that nondairy products will overtake real-natural product in the short term. Milk powders are still a big business and the optimization approach to change the paradigm from inspect and reject to predict and prevent is developing increased interest for the factory of the future [101], and is an approach that cannot be ignored. Regardless of developments, a quality milk product must be the result. There can be no food security without food safety.

## **Acknowledgements**

the parlor without prompting and some cows are milked three times a day, with increased milk product for the farmer. The tags on the cows allow for integration into the machines which collect vast amounts of data, including number of steps, chewing the curd, etc. Robotic milking machines have a life span of approx. 13 years and then required further investment. Determination of when a cow is in heat for efficient reproduction is available with MooCow developed by Dairy Master (Ireland), together with MooMonitor to guide cows in the parlor. A separate company created MooCall, a sensor attached to the cow tail, which can monitor contractions during calf birth and send a SMS message to the farmer, the sensor can determine as close as 1 h to delivery [91]. Some of the more recent analytical instruments for milk analysis that has been introduced, but are not yet standard and include: Fourier transform MIR spectroscopy for milk-based quantitative, qualitative phenotypic and genomic analysis. Flow CYTOMETRY is a well-established technique for bacteria and somatic cells counting and differentiation [92], and companies making these include: Bentley (https://bentleyinstruments.com/), Foss (https://www.fossanalytics.com/en#) and Delta (http://www.deltainstruments.com/); ELISA (enzyme linked immunosorbent assay) for milk amyloid; milk amyloidA (MAA) biomarker is an early detection method for clinical and subclinical mastitis. MAA is the only acute phase protein produced in response to bacterial infection in the udder and is therefore an immediate and direct indicates of infection [93]. PCR (polymerase chain reaction) is used for bacteria identification and differentiation [94, 95]. Gas chromatography/mass spectrometry can be used for analysis for metabolomics characterization of milk [96, 97]. Liquid chromatography/electrospray ionization and mass spectroscopy can be used for the detection of peptides and glycopeptides. ICP-MS (Induction coupled plasma mass spectroscopy) for mineral and trace element analysis. Industry 4.0 technologies can be introduced at all stages

of the plant manufacturing systems, including environment monitoring [98].

be safe and comply with regulatory requirements.

The milk processing chain demands accurate and quality products from farm to plate and for all of its products, e.g., fluid milk, milk powders, etc. It must start with the raw material at farm level including; dairy herd improvement testing, to payment parameters, and quality control of the raw milk. Optimization is important in the processing of milk in the dairy chain as 73 plus tests are carried out including chemical physical and microbiological tests, set against ISO standards, EU, USFDA regulations, and most countries internal regulations. Advances are slowly being made to have modern and optimized methodologies approved. The regulatory bodies are setting new standards from verified inter-laboratory studies, targeting the advancement in instrumentation and for at-line and in-line production analysis for improved predictability and control of manufacturing processes. The finished product must

At a conference in Glasgow (Semex Dairy Conference, Jan 2018), it was questioned whether the dairy industry could cease to exist after approximately 10 years, due to the interest in vegan alternatives and the increased population who are lactose intolerant [99, 100]. A business model to address this alternative has resulted in a cow-free milk product called Perfect

**8. Conclusion**

16 Descriptive Food Science

This work was supported by the Irish State through funding from the Technology Centre's program—Grant Number TC/2014/0016.

## **Conflict of interest**

There is no conflict of interest.

## **Author details**

Niamh Burke1 , Krzysztof A. Zacharski1 , Mark Southern1,2, Paul Hogan<sup>1</sup> , Michael P. Ryan<sup>3</sup> and Catherine C. Adley3 \*

\*Address all correspondence to: catherine.adley@ul.ie

1 Dairy Processing Technology Centre (DPTC), University of Limerick, Limerick, Ireland

2 Enterprise Research Centre, University of Limerick, Limerick, Ireland

3 Microbiology Laboratory, School of Natural Sciences, University of Limerick, Limerick, Ireland

## **References**


[4] FAO-UN. Global Dairy Sector: Facts. 2016. Food and Agriculture Organisation of the United Nations. Available from: https://www.fil-idf.org/wp-content/uploads/2016/12/ FAO-Global-Facts-1.pdf [Accessed: February 24, 2018]

[16] EPA. US Environmental Protection Agency. Technical Fact Sheet—Perchlorate [Internet]. 2014. Available from: https://www.epa.gov/sites/production/files/2014-03/documents/

The Dairy Industry: Process, Monitoring, Standards, and Quality

http://dx.doi.org/10.5772/intechopen.80398

19

[17] Leung AM, Pearce EN, Braverman LE. Perchlorate, iodine and the thyroid. Best Practice & Research. Clinical Endocrinology & Metabolism. 2010;**24**(1):133-141. DOI: 10.1016/j.beem.

[18] ISO/TC34/SC5. Milk and Milk Products, (Calalogue of Standards). Geneva, Switzerland:

[19] ISO 6091. Dried Milk. Determination of Titratable Acidity (Reference Method). Geneva,

[20] ISO 26844. Milk and Milk Products. Determination of Antimicrobial Residues. Tube Diffusion Test. Geneva, Switzerland: International Organization for Standardization;

[21] ISO 5764. Milk—Determination of Freezing Point—Thermistor Cryoscopy Method (Reference Method). Geneva, Switzerland: International Organization for Standardi-

[22] ISO 1736. Dried Milk and Dried Milk Products. Determination of Fat Content. Gravimetric Method (Reference Method). Geneva, Switzerland: International Organization for

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**Chapter 2**

**Provisional chapter**

**Food Authenticity: Provenancing. A Case Study of Fish**

Authentication of food products is of ongoing interest to consumers in developed countries. Recently, a general interest in the sustainability of food productions, from both societal and environmental perspectives, developed and added a new dimension. Fish and fish products are common targets for food adulteration. The most important issue is fish management, e.g., the environmental impact of overfishing. Analytical means would be helpful for verification. The aim of the present study was to evaluate various marker groups for the distinction of European plaice from the North Sea from European plaice from other geographical origins: volatile organic compounds (VOCs), fatty acids (FA), and isotope ratios. VOCs were analyzed using proton transfer reaction mass spectrometry (PTR-MS); the FA composition was analyzed using gas chromatography with a flame ionization detector, and carbon, hydrogen, nitrogen; and sulfur isotope ratios were analyzed using isotope ratio mass spectrometry. In a principal component analysis, FA profiling appeared the best option to distinguish European plaice from the North Sea

**Keywords:** European plaice, fatty acid composition, geographical origin, isotope ratio,

In food science and technology, considerable attention to the preservation and quality of foods has been paid throughout history. Food authenticity is a typical food quality issue and is not new either. Some food authenticity infringements were reported over 2000 years ago, and the first related food standards appeared 150 years ago. The classical form of food authenticity violations is the removal or addition of undeclared ingredients or constituents. However,

**Food Authenticity: Provenancing. A Case Study of Fish**

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.80470

Annemieke M. Pustjens, Rita Boerrigter-Eenling,

Annemieke M. Pustjens, Rita Boerrigter-Eenling,

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

from those originating from other seas.

PCA, *Pleuronectes platessa*, PTR-MS

Alex H. Koot and Saskia M. van Ruth

Alex H. Koot and Saskia M. van Ruth

http://dx.doi.org/10.5772/intechopen.80470

**Abstract**

**1. Introduction**

#### **Food Authenticity: Provenancing. A Case Study of Fish Food Authenticity: Provenancing. A Case Study of Fish**

DOI: 10.5772/intechopen.80470

Annemieke M. Pustjens, Rita Boerrigter-Eenling, Alex H. Koot and Saskia M. van Ruth Annemieke M. Pustjens, Rita Boerrigter-Eenling, Alex H. Koot and Saskia M. van Ruth

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.80470

#### **Abstract**

Authentication of food products is of ongoing interest to consumers in developed countries. Recently, a general interest in the sustainability of food productions, from both societal and environmental perspectives, developed and added a new dimension. Fish and fish products are common targets for food adulteration. The most important issue is fish management, e.g., the environmental impact of overfishing. Analytical means would be helpful for verification. The aim of the present study was to evaluate various marker groups for the distinction of European plaice from the North Sea from European plaice from other geographical origins: volatile organic compounds (VOCs), fatty acids (FA), and isotope ratios. VOCs were analyzed using proton transfer reaction mass spectrometry (PTR-MS); the FA composition was analyzed using gas chromatography with a flame ionization detector, and carbon, hydrogen, nitrogen; and sulfur isotope ratios were analyzed using isotope ratio mass spectrometry. In a principal component analysis, FA profiling appeared the best option to distinguish European plaice from the North Sea from those originating from other seas.

**Keywords:** European plaice, fatty acid composition, geographical origin, isotope ratio, PCA, *Pleuronectes platessa*, PTR-MS

#### **1. Introduction**

In food science and technology, considerable attention to the preservation and quality of foods has been paid throughout history. Food authenticity is a typical food quality issue and is not new either. Some food authenticity infringements were reported over 2000 years ago, and the first related food standards appeared 150 years ago. The classical form of food authenticity violations is the removal or addition of undeclared ingredients or constituents. However,

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

more recently, a general interest in the sustainability of food productions, from both societal and environmental perspectives, developed and added a new dimension. With advancing technology and increased awareness of social corporate responsibility, new questions have emerged in the food authenticity arena. How are production in terms of raw materials and processing mirrored in unique product characteristics? Do special production (farming management systems, geographical origin, etc.) result in special traits? How to discern and measure these relevant characteristics? And, can these characteristics be used for discriminatory purposes and/or substantiate the produce quality?

**1.1. VOC profile in the verification of geographical origin**

**1.2. FA profile in the verification of geographical origin**

**1.3. IR in the verification of geographical origin**

cal origin of different fish species [20, 21].

**1.4. Case study: geographical origin of European plaice**

higher latitude, marine organisms contain more n-3 long-chain PUFAs.

the diet they consumed [14].

VOCs have shown to satisfactorily predict the country of origin of olive oil, truffles, Grana Padano cheese, as reviewed by Luykx et al. [5] and crude palm oil, butter, cumin cheese, dry cured hams, and coffees, as listed by Pustjens et al. [6]. More recently, VOCs have also shown to successfully verify geographical origin of Chinese cabbage [7], plant-related liquors [8], saffron [9], rosemary [10], teas [11], and capers [12]. For plant products, the difference in VOC profile was ascribed to differences in climate and soil type [13]. This can be translated to fish, since the sea bed and climate will vary for fish from different geographical origins. The VOC profile of fish has been linked to lipid type and source and would therefore be determined by

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470 29

FAs have shown to satisfactorily predict the country of origin of milk and olive oils, as reviewed by Luykx et al. [5] and chocolate and wheat, as listed by Pustjens et al. [6]. More recently, FAs have also shown to successfully verify geographical origin of berries [15], cockles [16], and almonds [17]. FAs in the fish feed are incorporated into its tissues; therefore, the FA profile of the fish tissue reflects the diet over a longer period of time [18]. Especially the content of polyunsaturated fatty acids (PUFAs) seems to be dependent on the geographical origin, i.e., the latitude, and thus broadly related to temperature [19]. They found that at a

All food products have their own isotopic composition, which is determined by the animal's diet (carbon and nitrogen), climate (hydrogen and oxygen), and soil composition (sulfur) [6]. IRMS analysis combined with or without other techniques and/or chemometric models has been successfully applied to determine the geographical origin of dairy and animal products, vegetables, natural flavors, honey, wines, coffee, and fruit, as reviewed by Luykx et al. [5] and cereals, fish and crustaceans, and olive oil and wine, as listed by Pustjens et al. [6]. IRMS analysis of carbon, hydrogen, nitrogen, and oxygen has shown to be able to verify geographi-

European plaice (*Pleuronectes platessa*) is the principal commercial flatfish in Europe [22]. European plaice live at the bottom of the sea not far from the shore, predominantly in the North Sea, but extend to the Baltic Sea, the Barents Sea, and the sea around Ireland and Iceland [23]. Juvenile European plaice feed on a variety of microbenthic species [24], whereas larger European plaice mainly feed on Polychaeta and mollusks [25]. The habitat of European plaice is dependent on: (a) their life stages: they gradually move from shallow coastal nurseries into deeper water [26, 27] and (b) climate change: European plaice

Adulteration has been defined as "the fraudulent addition of non-authentic substances or removal or replacement of authentic substances without the purchaser's knowledge for economic gain of the seller" by the United States Pharmacopeial Convention (USP). Fish and fish products are common targets for food adulteration [1]. There are different issues in regard to the traceability of fish: (1) the species of the fish, (2) geographical origin, and (3) the production method [2]. During production, fish types can be completely interchanged or mixed with cheaper or less sustainable fish types. According to European law (Regulation 1379/2013), information concerning the commercial designation, the catch area, and the production method shall be available at each stage of marketing of the species concerned [3]. Next to the fact that geographical origin of food products is laid down in legislation, consumers find it important because of economic, safety, and sustainability reasons. Different types of food adulteration can be distinguished: most types influence the product composition by, e.g., substitution or dilution, while other issues involve cultivar and variety of products of plant origin, species of products of animal origin, geographical origin of foods, production system, and processing. Geographical origin has become a more important issue in recent years for economic, safety, and sustainability reasons [4]. Counterfeiting occurs, for example, with the European Union's protected designation of origin, like certain cheeses or olive oils.

Authenticity and traceability research aims to combine different complementary analytical strategies to determine the authenticity of materials and commodities. Various techniques have been studied based on organic constituents, mineral contents or composition, light- or heavy-element isotope ratios, or combinations thereof. If the components have sufficient discriminatory power, the set of their concentrations will form a characteristic pattern or "fingerprint" relating to the geographical origin of the sample. Chemometrics provides the ability to detect these patterns, and is essentially helpful when the number of components necessary to differentiate samples from different geographical origins increases [5]. The three major groups of analytical techniques are: mass spectrometry techniques, spectroscopic techniques, and separation techniques.

Markers that have been successfully used for the verification of the geographical origin of food and feed products are volatile organic compounds (VOCs), the fatty acid (FA) profile, and isotope ratios. These markers can be analyzed using mass spectrometry (like isotope ratio mass spectrometry—IRMS; and proton transfer reaction mass spectrometry—PTR-MS), spectroscopic techniques (like NMR and infrared spectroscopy), and separation techniques (such as liquid chromatography—LC; and gas chromatography—GC) [6].

#### **1.1. VOC profile in the verification of geographical origin**

more recently, a general interest in the sustainability of food productions, from both societal and environmental perspectives, developed and added a new dimension. With advancing technology and increased awareness of social corporate responsibility, new questions have emerged in the food authenticity arena. How are production in terms of raw materials and processing mirrored in unique product characteristics? Do special production (farming management systems, geographical origin, etc.) result in special traits? How to discern and measure these relevant characteristics? And, can these characteristics be used for discriminatory

Adulteration has been defined as "the fraudulent addition of non-authentic substances or removal or replacement of authentic substances without the purchaser's knowledge for economic gain of the seller" by the United States Pharmacopeial Convention (USP). Fish and fish products are common targets for food adulteration [1]. There are different issues in regard to the traceability of fish: (1) the species of the fish, (2) geographical origin, and (3) the production method [2]. During production, fish types can be completely interchanged or mixed with cheaper or less sustainable fish types. According to European law (Regulation 1379/2013), information concerning the commercial designation, the catch area, and the production method shall be available at each stage of marketing of the species concerned [3]. Next to the fact that geographical origin of food products is laid down in legislation, consumers find it important because of economic, safety, and sustainability reasons. Different types of food adulteration can be distinguished: most types influence the product composition by, e.g., substitution or dilution, while other issues involve cultivar and variety of products of plant origin, species of products of animal origin, geographical origin of foods, production system, and processing. Geographical origin has become a more important issue in recent years for economic, safety, and sustainability reasons [4]. Counterfeiting occurs, for example, with the

European Union's protected designation of origin, like certain cheeses or olive oils.

Authenticity and traceability research aims to combine different complementary analytical strategies to determine the authenticity of materials and commodities. Various techniques have been studied based on organic constituents, mineral contents or composition, light- or heavy-element isotope ratios, or combinations thereof. If the components have sufficient discriminatory power, the set of their concentrations will form a characteristic pattern or "fingerprint" relating to the geographical origin of the sample. Chemometrics provides the ability to detect these patterns, and is essentially helpful when the number of components necessary to differentiate samples from different geographical origins increases [5]. The three major groups of analytical techniques are: mass spectrometry techniques, spectroscopic techniques,

Markers that have been successfully used for the verification of the geographical origin of food and feed products are volatile organic compounds (VOCs), the fatty acid (FA) profile, and isotope ratios. These markers can be analyzed using mass spectrometry (like isotope ratio mass spectrometry—IRMS; and proton transfer reaction mass spectrometry—PTR-MS), spectroscopic techniques (like NMR and infrared spectroscopy), and separation techniques (such

as liquid chromatography—LC; and gas chromatography—GC) [6].

purposes and/or substantiate the produce quality?

28 Descriptive Food Science

and separation techniques.

VOCs have shown to satisfactorily predict the country of origin of olive oil, truffles, Grana Padano cheese, as reviewed by Luykx et al. [5] and crude palm oil, butter, cumin cheese, dry cured hams, and coffees, as listed by Pustjens et al. [6]. More recently, VOCs have also shown to successfully verify geographical origin of Chinese cabbage [7], plant-related liquors [8], saffron [9], rosemary [10], teas [11], and capers [12]. For plant products, the difference in VOC profile was ascribed to differences in climate and soil type [13]. This can be translated to fish, since the sea bed and climate will vary for fish from different geographical origins. The VOC profile of fish has been linked to lipid type and source and would therefore be determined by the diet they consumed [14].

#### **1.2. FA profile in the verification of geographical origin**

FAs have shown to satisfactorily predict the country of origin of milk and olive oils, as reviewed by Luykx et al. [5] and chocolate and wheat, as listed by Pustjens et al. [6]. More recently, FAs have also shown to successfully verify geographical origin of berries [15], cockles [16], and almonds [17]. FAs in the fish feed are incorporated into its tissues; therefore, the FA profile of the fish tissue reflects the diet over a longer period of time [18]. Especially the content of polyunsaturated fatty acids (PUFAs) seems to be dependent on the geographical origin, i.e., the latitude, and thus broadly related to temperature [19]. They found that at a higher latitude, marine organisms contain more n-3 long-chain PUFAs.

#### **1.3. IR in the verification of geographical origin**

All food products have their own isotopic composition, which is determined by the animal's diet (carbon and nitrogen), climate (hydrogen and oxygen), and soil composition (sulfur) [6]. IRMS analysis combined with or without other techniques and/or chemometric models has been successfully applied to determine the geographical origin of dairy and animal products, vegetables, natural flavors, honey, wines, coffee, and fruit, as reviewed by Luykx et al. [5] and cereals, fish and crustaceans, and olive oil and wine, as listed by Pustjens et al. [6]. IRMS analysis of carbon, hydrogen, nitrogen, and oxygen has shown to be able to verify geographical origin of different fish species [20, 21].

#### **1.4. Case study: geographical origin of European plaice**

European plaice (*Pleuronectes platessa*) is the principal commercial flatfish in Europe [22]. European plaice live at the bottom of the sea not far from the shore, predominantly in the North Sea, but extend to the Baltic Sea, the Barents Sea, and the sea around Ireland and Iceland [23]. Juvenile European plaice feed on a variety of microbenthic species [24], whereas larger European plaice mainly feed on Polychaeta and mollusks [25]. The habitat of European plaice is dependent on: (a) their life stages: they gradually move from shallow coastal nurseries into deeper water [26, 27] and (b) climate change: European plaice have shifted toward deeper and more northern areas [28]. This information can be used to find specific groups of markers potentially useful for the verification of their geographical origin.

**2.2. Chemicals**

otherwise.

*2.3.1. PTR-MS*

*2.3.3. IR-MS*

**2.3. Analytical methods**

All chemicals used were of analytical grade purchased from Merck (Darmstadt, Germany), Sigma-Aldrich (St. Louis, MO, USA), or VWR International (Radnor, PA, USA), unless stated

The samples were thawed to room temperature in their package and 1 gram of finely sliced material was placed in a 250-mL glass bottle. The headspace above the samples was equilibrated in a waterbath at 25°C for 30 minutes and kept at 25°C for the duration of the measurement. Two independent replicates of each sample were analyzed. The headspace was drawn from the sample flask at a rate of 60 ml/min, which was led through a heated transfer line into a high-sensitivity PTR-MS system (Ionicon GmbH, Innsbruck, Austria). A constant drift voltage of 600 V and a pressure of 2.20 ± 0.02 mbar were maintained in the reaction chamber. Data were collected for the mass range m/z 20–160 using a dwell time of 0.2 s/mass, resulting in a duration of 28 s/cycle. The instrument was operated at a standard E/N (ratio of electric field strength across the drift tube, E,

O+

and detected as ion count/s, resulting in a fingerprint of the volatiles. Each sample was analyzed for five full mass cycles. The headspace concentrations of the compounds during the second, third, and fourth cycles were calculated as described by Hansel et al. [29]. Blank measurements (empty bottle) were performed for five cycles, with the average of the third, fourth, and fifth mass cycles subtracted from the samples' spectra as background correction. These corrected headspace

The samples were freeze-dried. Fat was extracted from the samples (2 g) using chloroform:methanol (2:1, v/v) as described by Tres et al. [30]. Fatty acids were methyl esterified according to Guardiola et al. [31]. The obtained methyl esters were analyzed in duplicate using an Agilent 7890A GC system, fitted with a flame ionization detector, using a CP7419 50 m × 0.25 mm FAME column (Agilent, Santa Clara, CA, USA). Temperature was kept at 100°C for 1 minute and then increased to 230°C with 5°C/min, after which the temperature was kept at 230°C for 9 min. Helium was used as a carrier gas using a flow of 0.9 ml per min and a split ratio of 1:50. Fatty acids were identified by comparing retention times with those of standard mixtures (Supelco 37 component FAME mix (Supelco, St. Louis, MO, USA)). Results

The dried defatted samples (1 g) were placed in a paper filter and washed three times with 10 ml of distilled water for removal of sodium sulfate, then three times with 10 ml of acetone to accelerate drying, and then placed into glass containers and dried in vacuum overnight.

). Inlet and drift chamber tempera-

ionization in a quadrupole mass spectrometer

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470 31

to buffer gas density, N) of 138 Td (1Td = 10−<sup>17</sup> cm2 V molecule−<sup>1</sup>

concentrations (ppbv) were subsequently used for further statistical analysis.

were expressed as normalized peak areas (% of total fatty acids).

tures were 60°C. Masses are analyzed after H3

*2.3.2. GC flame ionization detection (GC-FID)*

The aim of this study was to examine how well VOCs, FAs, and isotope ratios are able to distinguish European plaice from the North Sea from European plaice from other provenance.

## **2. Materials and methods**

#### **2.1. Samples**

A reference set of 49 European plaice were collected by local fishermen from five different locations in the Northeast Atlantic ocean—FAO area 27: the North Sea, subarea IV a, b, c (n = 31); other research institutes from the Baltic Sea, subarea IIId (n = 3); the Barents Sea, subarea I (n = 5); the sea around Ireland, subarea Va (n = 5); and the sea around Iceland, subarea VIIj2 (n = 5). Locations are labeled as NS, BalS, BarS, SIr, and SIc, respectively, in **Figure 1**. European plaice from the North Sea were both collected in 2013 and 2014; all other samples were collected in 2014. Samples were transported on dry ice to our institute, filleted and stored at −18°C until further analyses. In 2014, a set of market samples were also collected: 11 non-breaded and non-marinated frozen European plaice fillets from Italian (n = 6), German (n = 4), and Dutch (n = 1) supermarkets. According to the package, they originated from the Northeast Atlantic Ocean (FAO zone 27). For some fish, the North Sea, which is part of this FAO catch area, was mentioned specifically.

**Figure 1.** Habitat of European plaice, with the sample locations marked as NS—North Sea, BalS—Baltic Sea, BarS—Barents Sea, SIr—sea around Ireland, and SIc—sea around Iceland. Original figure from FAO Fish Finder (23).

#### **2.2. Chemicals**

have shifted toward deeper and more northern areas [28]. This information can be used to find specific groups of markers potentially useful for the verification of their geographi-

The aim of this study was to examine how well VOCs, FAs, and isotope ratios are able to distinguish European plaice from the North Sea from European plaice from other

A reference set of 49 European plaice were collected by local fishermen from five different locations in the Northeast Atlantic ocean—FAO area 27: the North Sea, subarea IV a, b, c (n = 31); other research institutes from the Baltic Sea, subarea IIId (n = 3); the Barents Sea, subarea I (n = 5); the sea around Ireland, subarea Va (n = 5); and the sea around Iceland, subarea VIIj2 (n = 5). Locations are labeled as NS, BalS, BarS, SIr, and SIc, respectively, in **Figure 1**. European plaice from the North Sea were both collected in 2013 and 2014; all other samples were collected in 2014. Samples were transported on dry ice to our institute, filleted and stored at −18°C until further analyses. In 2014, a set of market samples were also collected: 11 non-breaded and non-marinated frozen European plaice fillets from Italian (n = 6), German (n = 4), and Dutch (n = 1) supermarkets. According to the package, they originated from the Northeast Atlantic Ocean (FAO zone 27). For some fish, the North Sea, which is part

**Figure 1.** Habitat of European plaice, with the sample locations marked as NS—North Sea, BalS—Baltic Sea, BarS—Barents Sea, SIr—sea around Ireland, and SIc—sea around Iceland. Original figure from FAO Fish Finder (23).

cal origin.

30 Descriptive Food Science

provenance.

**2.1. Samples**

**2. Materials and methods**

of this FAO catch area, was mentioned specifically.

All chemicals used were of analytical grade purchased from Merck (Darmstadt, Germany), Sigma-Aldrich (St. Louis, MO, USA), or VWR International (Radnor, PA, USA), unless stated otherwise.

#### **2.3. Analytical methods**

#### *2.3.1. PTR-MS*

The samples were thawed to room temperature in their package and 1 gram of finely sliced material was placed in a 250-mL glass bottle. The headspace above the samples was equilibrated in a waterbath at 25°C for 30 minutes and kept at 25°C for the duration of the measurement. Two independent replicates of each sample were analyzed. The headspace was drawn from the sample flask at a rate of 60 ml/min, which was led through a heated transfer line into a high-sensitivity PTR-MS system (Ionicon GmbH, Innsbruck, Austria). A constant drift voltage of 600 V and a pressure of 2.20 ± 0.02 mbar were maintained in the reaction chamber. Data were collected for the mass range m/z 20–160 using a dwell time of 0.2 s/mass, resulting in a duration of 28 s/cycle. The instrument was operated at a standard E/N (ratio of electric field strength across the drift tube, E, to buffer gas density, N) of 138 Td (1Td = 10−<sup>17</sup> cm2 V molecule−<sup>1</sup> ). Inlet and drift chamber temperatures were 60°C. Masses are analyzed after H3 O+ ionization in a quadrupole mass spectrometer and detected as ion count/s, resulting in a fingerprint of the volatiles. Each sample was analyzed for five full mass cycles. The headspace concentrations of the compounds during the second, third, and fourth cycles were calculated as described by Hansel et al. [29]. Blank measurements (empty bottle) were performed for five cycles, with the average of the third, fourth, and fifth mass cycles subtracted from the samples' spectra as background correction. These corrected headspace concentrations (ppbv) were subsequently used for further statistical analysis.

#### *2.3.2. GC flame ionization detection (GC-FID)*

The samples were freeze-dried. Fat was extracted from the samples (2 g) using chloroform:methanol (2:1, v/v) as described by Tres et al. [30]. Fatty acids were methyl esterified according to Guardiola et al. [31]. The obtained methyl esters were analyzed in duplicate using an Agilent 7890A GC system, fitted with a flame ionization detector, using a CP7419 50 m × 0.25 mm FAME column (Agilent, Santa Clara, CA, USA). Temperature was kept at 100°C for 1 minute and then increased to 230°C with 5°C/min, after which the temperature was kept at 230°C for 9 min. Helium was used as a carrier gas using a flow of 0.9 ml per min and a split ratio of 1:50. Fatty acids were identified by comparing retention times with those of standard mixtures (Supelco 37 component FAME mix (Supelco, St. Louis, MO, USA)). Results were expressed as normalized peak areas (% of total fatty acids).

#### *2.3.3. IR-MS*

The dried defatted samples (1 g) were placed in a paper filter and washed three times with 10 ml of distilled water for removal of sodium sulfate, then three times with 10 ml of acetone to accelerate drying, and then placed into glass containers and dried in vacuum overnight.

For analysis of δ<sup>2</sup> H, 250–300 μg of dried sample material was weighed into a silver capsule. δ2 H was determined using a high-temperature pyrolysis system working at 1450°C on the basis of ceramic tubes with glassy carbon reactor filling coupled with a Thermo Finnigan Delta-Plus XL IRMS system (Thermo Fisher Inc.), designed for continuous flow isotope ratio mass spectrometry (CF-IRMS) of hydrogen isotopes in helium carrier gas.

For the other isotopes (for δ<sup>13</sup>C, δ<sup>15</sup>N, and δ34S), 3–4 mg of sample material was weighed into tin capsules. The measurements of δ<sup>13</sup>C, δ<sup>15</sup>N, and δ<sup>34</sup>S were carried out on an elemental analyzer (Vario EL, Elementar, Hanau, Germany) coupled with an isotope-ratio mass spectrometer, IRMS (IsoPrime, GV Instruments, Manchester, UK). All isotopes were analyzed in duplicate.

The values of the isotope ratios are expressed in δ (‰) and correspond to international standards (V-SMOW for δ<sup>2</sup> H, V-PDB for δ<sup>13</sup>C, air for δ15N, and V-CDT for δ<sup>34</sup>S) according to the relation:

$$\delta\left(\%\right) = 1000 \ge \frac{R\_{sample} - R\_{standard}}{R\_{standard}} \tag{1}$$

the first dimension. Samples from the other seas demonstrated high negative scores in this dimension. Remarkable is the fact that there is more variation in European plaice from the North Sea than in European plaice from the other seas. This might be caused by the fact that the European plaice from the North Sea have been sampled at six different locations within the North Sea, while the other seas have been sampled only on one location. For this reference set, distinction of the North Sea European plaice from other European plaice by their VOCs

However, the VOC profiles are affected not only by the fish diet [14] but also by the freshness of the fish [32, 33]. This factor may interfere in the analysis. VOCs formed during storage are alcohols, aldehydes, esters, organic acids, and sulfur compounds, which can be either of microbial or nonmicrobial origin [34]. Their non-protonated masses range from m/z 46 for ethanol to m/z 176 for 3-hydroxy-2-butanone. This is a complicating factor and can influence

 398 ± 623 43 ± 14 60 ± 15 30 ± 17 26 ± 3 0.000 3 ± 1 3 ± 0 4 ± 0 4 ± 0 4 ± 0 0.000 10 ± 1 10 ± 1 12 ± 1 11 ± 2 10 ± 1 0.063 14 ± 7 12 ± 4 11 ± 2 12 ± 3 7 ± 4 0.044 39 ± 43 2 ± 1 1 ± 0 0 ± 0 0 ± 0 0.000 23 ± 12 13 ± 4 10 ± 1 7 ± 1 16 ± 5 0.001 301 ± 240 204 ± 161 69 ± 34 101 ± 29 579 ± 239 0.044 7 ± 5 4 ± 4 2 ± 1 2 ± 1 13 ± 6 0.051 6 ± 8 14 ± 12 3 ± 1 2 ± 1 55 ± 25 0.793 6 ± 3 4 ± 1 6 ± 1 5 ± 1 5 ± 1 0.983 10 ± 10 4 ± 1 3 ± 1 25 ± 13 2 ± 1 0.049 168 ± 112 79 ± 8 53 ± 17 45 ± 13 66 ± 52 0.000 6 ± 4 3 ± 0 2 ± 1 2 ± 0 2 ± 2 0.000 1 ± 1 2 ± 2 1 ± 0 1 ± 0 2 ± 0 0.001 7 ± 2 3 ± 2 4 ± 1 3 ± 1 4 ± 2 0.000 4 ± 2 1 ± 0 0 ± 0 1 ± 0 3 ± 1 0.000 7 ± 2 8 ± 1 9 ± 4 6 ± 1 6 ± 3 0.369 3 ± 2 1 ± 1 2 ± 0 1 ± 0 1 ± 0 0.000

**Table 1.** Average protonated VOC composition (ppbv) of European plaice sampled from the North Sea (NS), the Baltic Sea (BalS), the Barents Sea (BarS), the sea around Ireland (SIr), and the sea around Iceland (SIc); mean intensity, standard deviation, and significant differences between European plaice from the North Sea and European plaice caught in other

**NS (n = 31) BalS (n = 3) BarS (n = 5) SIr (n = 5) SIc (n = 5) P-value NS** 

**versus other seas**

33

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470

is a promising approach.

the robustness of the method.

**Protonated VOC** 

seas (Kruskal-Wallis).

**(m/z)**

where R represents the ratio of the higher mass to the lower mass isotopes measured in the sample (Rsample) and in the standard (Rstandard).

#### **2.4. Data analysis**

Normality of the distribution of VOC profiles, FA profiles, and isotope ratios within the groups of geographical origin was checked using the Shapiro-Wilk test. Distributions appeared to be nonnormal (Shapiro-Wilk P < 0.05). Therefore, Kruskal-Wallis test for group comparisons was performed among the provenance groups using SPSS version 23.0.0.2 (IBM Corp., Armonk, NY, USA). Principal component analysis (PCA) was performed using the Unscrambler (Version X 10.3). Raw data were auto-scaled prior to PCA.

## **3. Results and discussion**

#### **3.1. Distinction of the provenance of European plaice**

#### *3.1.1. VOC profiles analyzed by PTR-MS*

The reference set of 49 European plaice samples was subjected to PTR-MS analyses. Most abundant masses of protonated VOCs of European plaice collected from the North Sea are m/z 33, 45, and 59. Subsequently, significant differences in mass intensities between European plaice originating from the North Sea versus other provenance were examined (Kruskal-Wallis P < 0.05). Results are presented in **Table 1**.

In order to explore the data, PCA was carried out. PCA (**Figure 2A**) revealed grouping of samples, with two-third of the samples from the North Sea showing high positive scores in the first dimension. Samples from the other seas demonstrated high negative scores in this dimension. Remarkable is the fact that there is more variation in European plaice from the North Sea than in European plaice from the other seas. This might be caused by the fact that the European plaice from the North Sea have been sampled at six different locations within the North Sea, while the other seas have been sampled only on one location. For this reference set, distinction of the North Sea European plaice from other European plaice by their VOCs is a promising approach.

For analysis of δ<sup>2</sup>

32 Descriptive Food Science

in duplicate.

relation:

dards (V-SMOW for δ<sup>2</sup>

**2.4. Data analysis**

**3. Results and discussion**

*3.1.1. VOC profiles analyzed by PTR-MS*

*δ* (‰) = 1000 *x*

sample (Rsample) and in the standard (Rstandard).

(Version X 10.3). Raw data were auto-scaled prior to PCA.

**3.1. Distinction of the provenance of European plaice**

Wallis P < 0.05). Results are presented in **Table 1**.

δ2

H, 250–300 μg of dried sample material was weighed into a silver capsule.

H, V-PDB for δ<sup>13</sup>C, air for δ15N, and V-CDT for δ<sup>34</sup>S) according to the

(1)

*<sup>R</sup>sample* <sup>−</sup> *<sup>R</sup>* \_\_\_\_\_\_\_\_\_\_\_ *standard Rstandard*

H was determined using a high-temperature pyrolysis system working at 1450°C on the basis of ceramic tubes with glassy carbon reactor filling coupled with a Thermo Finnigan Delta-Plus XL IRMS system (Thermo Fisher Inc.), designed for continuous flow isotope ratio

For the other isotopes (for δ<sup>13</sup>C, δ<sup>15</sup>N, and δ34S), 3–4 mg of sample material was weighed into tin capsules. The measurements of δ<sup>13</sup>C, δ<sup>15</sup>N, and δ<sup>34</sup>S were carried out on an elemental analyzer (Vario EL, Elementar, Hanau, Germany) coupled with an isotope-ratio mass spectrometer, IRMS (IsoPrime, GV Instruments, Manchester, UK). All isotopes were analyzed

The values of the isotope ratios are expressed in δ (‰) and correspond to international stan-

where R represents the ratio of the higher mass to the lower mass isotopes measured in the

Normality of the distribution of VOC profiles, FA profiles, and isotope ratios within the groups of geographical origin was checked using the Shapiro-Wilk test. Distributions appeared to be nonnormal (Shapiro-Wilk P < 0.05). Therefore, Kruskal-Wallis test for group comparisons was performed among the provenance groups using SPSS version 23.0.0.2 (IBM Corp., Armonk, NY, USA). Principal component analysis (PCA) was performed using the Unscrambler

The reference set of 49 European plaice samples was subjected to PTR-MS analyses. Most abundant masses of protonated VOCs of European plaice collected from the North Sea are m/z 33, 45, and 59. Subsequently, significant differences in mass intensities between European plaice originating from the North Sea versus other provenance were examined (Kruskal-

In order to explore the data, PCA was carried out. PCA (**Figure 2A**) revealed grouping of samples, with two-third of the samples from the North Sea showing high positive scores in

mass spectrometry (CF-IRMS) of hydrogen isotopes in helium carrier gas.

However, the VOC profiles are affected not only by the fish diet [14] but also by the freshness of the fish [32, 33]. This factor may interfere in the analysis. VOCs formed during storage are alcohols, aldehydes, esters, organic acids, and sulfur compounds, which can be either of microbial or nonmicrobial origin [34]. Their non-protonated masses range from m/z 46 for ethanol to m/z 176 for 3-hydroxy-2-butanone. This is a complicating factor and can influence the robustness of the method.


**Table 1.** Average protonated VOC composition (ppbv) of European plaice sampled from the North Sea (NS), the Baltic Sea (BalS), the Barents Sea (BarS), the sea around Ireland (SIr), and the sea around Iceland (SIc); mean intensity, standard deviation, and significant differences between European plaice from the North Sea and European plaice caught in other seas (Kruskal-Wallis).

**FA NS (n = 31) BalS (n = 3) BarS (n = 5) SIr (n = 5) SIc (n = 5) P-value NS** 

C12:0 0.03 ± 0.04 0.02 ± 0.01 0.02 ± 0.01 0.06 ± 0.03 0.01 ± 0.01 0.436 isoC14:0 0.03 ± 0.05 0.00 ± 0.00 0.02 ± 0.01 0.01 ± 0.02 0.00 ± 0.01 0.146 C14:0 2.02 ± 0.72 2.01 ± 0.28 2.54 ± 0.24 3.10 ± 0.91 2.69 ± 0.83 0.002 C14:1n5 0.15 ± 0.13 0.06 ± 0.01 0.06 ± 0.04 0.04 ± 0.04 0.03 ± 0.02 0.000 isoC15:0 0.06 ± 0.04 0.00 ± 0.00 0.03 ± 0.02 0.01 ± 0.02 0.01 ± 0.01 0.000 C15:0 0.74 ± 0.16 0.95 ± 0.05 0.70 ± 0.04 0.76 ± 0.11 0.50 ± 0.10 0.332 isoC16:0 0.26 ± 0.19 0.38 ± 0.00 0.35 ± 0.07 0.28 ± 0.09 0.26 ± 0.06 0.004 C15:1n5 0.02 ± 0.04 0.01 ± 0.01 0.03 ± 0.02 0.00 ± 0.00 0.05 ± 0.03 0.279 C16:0 17.22 ± 2.06 17.57 ± 0.23 15.89 ± 1.83 18.89 ± 2.84 20.02 ± 0.68 0.134 C16:1n9 0.43 ± 0.17 0.65 ± 0.06 0.53 ± 0.06 0.70 ± 0.16 0.42 ± 0.04 0.006 C16:1n7 4.28 ± 1.97 7.64 ± 2.21 16.74 ± 3.10 4.88 ± 1.24 6.45 ± 1.53 0.000 isoC17:0 0.43 ± 0.16 0.36 ± 0.05 0.39 ± 0.08 0.33 ± 0.11 0.28 ± 0.06 0.033 anteisoC17:0 0.40 ± 0.29 0.52 ± 0.05 0.49 ± 0.12 0.34 ± 0.14 0.35 ± 0.13 0.140 C17:1n7 0.31 ± 0.17 0.65 ± 0.03 0.28 ± 0.05 0.34<sup>a</sup> ± 0.15 0.25 ± 0.08 0.371 C18:0 4.53 ± 0.64 4.07 ± 0.04 3.16 ± 0.43 4.15 ± 0.42 3.94 ± 0.73 0.001 Trans C18:1 + C18:T2 0.38 ± 0.33 0.49 ± 0.10 0.40 ± 0.15 0.91 ± 0.40 0.65 ± 0.23 0.001 C18:1n9 6.38 ± 2.41 8.60 ± 1.06 8.81 ± 1.41 8.46 ± 1.62 8.21 ± 1.23 0.000 C18:1n7 3.31 ± 0.93 4.50 ± 0.47 4.17 ± 0.62 3.03 ± 0.66 4.23 ± 0.53 0.018 C18:2n6t 0.09 ± 0.07 0.14 ± 0.01 0.08 ± 0.02 0.09 ± 0.06 0.13 ± 0.06 0.253 c9t12C18:2 0.01 ± 0.03 0.02 ± 0.03 0.01 ± 0.02 0.02 ± 0.03 0.03 ± 0.05 0.189 t9c12C18:2 0.21 ± 0.11 0.14 ± 0.00 0.09 ± 0.02 0.23 ± 0.09 0.10 ± 0.02 0.001 C18:2n6c 0.84 ± 0.95 0.85 ± 0.08 1.21 ± 0.29 1.20 ± 0.57 0.41 ± 0.12 0.095 C18:3n6 0.07 ± 0.05 0.13 ± 0.03 0.16 ± 0.03 0.09 ± 0.02 0.06 ± 0.01 0.019 C18:3n3 + C20:0 0.38 ± 0.18 0.41 ± 0.02 0.57 ± 0.18 0.63 ± 0.19 0.29 ± 0.08 0.057 C20:1n9 1.16 ± 0.52 1.99 ± 0.07 2.45 ± 0.71 3.49 ± 1.36 1.98 ± 0.57 0.000 C21:0 0.07 ± 0.06 0.12 ± 0.02 0.08 ± 0.01 0.04 ± 0.02 0.09 ± 0.03 0.482 C20:2n6 0.35 ± 0.19 0.53 ± 0.01 0.40 ± 0.09 0.69 ± 0.33 0.30 ± 0.11 0.054 C20:3n6 0.14 ± 0.05 0.17 ± 0.04 0.09 ± 0.06 0.13 ± 0.05 0.05 ± 0.01 0.044 C20:3n3 + C20:4n6 + C22:0 6.00 ± 2.20 3.44 ± 0.26 3.28 ± 1.20 4.24 ± 0.75 2.44 ± 0.91 0.000 C22:1n9 0.19 ± 0.26 0.41 ± 0.04 0.41 ± 0.07 0.64 ± 0.17 0.48 ± 0.15 0.000 C20:5n3 17.70 ± 3.27 13.72 ± 1.45 13.19 ± 3.05 15.63 ± 3.33 19.44 ± 2.70 0.075 C22:2n6 0.07 ± 0.03 0.06 ± 0.00 0.10 ± 0.04 0.04 ± 0.05 0.05 ± 0.02 0.991

**versus other seas**

35

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470

**Figure 2.** Principal component analysis plot of (A) PTR-MS data, (B) fatty acid profile, and (C) all data combined, on European plaice from the North Sea (1—box), the Baltic Sea (2—dot), the Barents Sea (3—triangle), the sea around Ireland (4—diamond), and the sea around Iceland (5—inverted triangle).


**Figure 2.** Principal component analysis plot of (A) PTR-MS data, (B) fatty acid profile, and (C) all data combined, on European plaice from the North Sea (1—box), the Baltic Sea (2—dot), the Barents Sea (3—triangle), the sea around

Ireland (4—diamond), and the sea around Iceland (5—inverted triangle).

34 Descriptive Food Science


originating from other seas. This was expected since the FA composition of the fish tissue is reflected by their diets and these will vary for the different geographical locations [18].

Finally, all reference samples were subjected to IR-MS for analysis of carbon, hydrogen, nitrogen, and sulfur isotope ratios. Results are visualized in a scatter plot (**Figure 3**). Hydrogen iso-

(versus Vienna Standard Mean Ocean Water). This can be explained by the fact that organic tissue is synthesized from metabolized products and their isotopic composition is referenced to local food chains rather than dissolved water isotopes [36]. Sulfur isotope ratio (δ<sup>34</sup>S) is enriched in European plaice, ranging from +4.6 to +20.1‰. This is typical for marine fish, whereas freshwater fish typically shows depleted δ<sup>34</sup>S [37]. Carbon isotope ratio (δ<sup>13</sup>C) is depleted, ranging from −14.6 to −21.1‰. Nitrogen isotope ratio (δ<sup>15</sup>N) is enriched, ranging from +8.9 to +13.0‰. Both δ<sup>13</sup>C and δ<sup>15</sup>N ratios are in accordance with previous research on flatfish from the southern North Sea and claimed to reflect the European plaices' diet [38].

In both scatter plots, grouping of the samples according to their geographical origin is clearly visible. Only samples from the seas around Ireland and Iceland seem to group together. Nitrogen and hydrogen isotope ratios in the North Sea European plaice were significantly different from the ratios in European plaice from other seas (Kruskal-Wallis P < 0.05). This was expected based on the results previously described on the carbon, hydrogen, nitrogen, and

All VOC, FA, and isotope ratio data acquired in this study were also combined and subjected to PCA plot (**Figure 2C**). Again, groups of samples from the same geographical origin cluster together, with 80% of the European plaice from the North Sea demonstrating high positive

plaice from the North Sea (box), the Baltic Sea (dot), the Barents Sea (triangle), the sea around Ireland (diamond), and

H versus δ<sup>34</sup>S (right) on European

H) is severely depleted compared to sea water, ranging from −43.7 to −112.5‰

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470 37

*3.1.3. Isotope ratios by IR-MS*

oxygen isotope ratios of various fish species [20, 21].

**Figure 3.** Scatter plot of IRMS data on European plaice: δ<sup>13</sup>C versus δ15N (left) and δ<sup>2</sup>

tope ratio (δ<sup>2</sup>

*3.1.4. All data combined*

the sea around Iceland (plus).

\* SFA = saturated fatty acids, MUFA = monounsaturated fatty acids, PUFA = polyunsaturated fatty acids, OFA = other fatty acids.

**Table 2.** Average fatty acid composition of European plaice sampled from the North Sea (NS), the Baltic Sea (BalS), the Barents Sea (BarS), the sea around Ireland (SIr), and the sea around Iceland (SIc); mean concentration, standard deviation, and significant differences between European plaice from the North Sea and European plaice caught in other seas (Kruskal-Wallis).

#### *3.1.2. FA profiles by GC-FID*

Subsequently, the reference set of European plaice was subjected to GC-FID analysis. European plaice contains around 0.8% fat [35]. This was extracted and its FA composition was analyzed. The most abundant FAs in European plaice were palmitic acid (C16:0), eicosapentaenoic acid (EPA, C20:5n3), and docosahexaenoic acid (DHA, C22:6n3) (**Table 2**). Karl et al. [35] found a similar FA profile for European plaice from the North Sea.

When comparing the origins for each FA, 19 out of 35 FAs revealed significant differences between European plaice from the North Sea and European plaice from other seas (Kruskal-Wallis P < 0.05; **Table 2**). European plaice from the Barents Sea seems to have a rather distinct FA profile compared to the other seas. When considering groups of FAs instead of individual FAs, contents of MUFA (monounsaturated FA), PUFA (polyunsaturated FA), OFA (other FA), omega-3 FAs, and omega-6 FAs were significantly different in North Sea European plaice from European plaice originating from other seas (Kruskal-Wallis P < 0.05). Colombo et al. [19] suggested that fish originating from a higher latitude would contain more PUFA. However, this was not confirmed in our study. European plaice from the sea with the highest latitude, the Barents Sea, showed to have the lowest PUFA content, namely 25%. The sea around Ireland, which has the lowest latitude, showed a rather average PUFA content, namely 33%.

PCA reveals clustering of samples (**Figure 2B**). Generally, the samples from the North Sea and those from the other seas are separated in the second dimension. Therefore, the FA profile seems to be a suitable option to distinguish North Sea European plaice from those originating from other seas. This was expected since the FA composition of the fish tissue is reflected by their diets and these will vary for the different geographical locations [18].

#### *3.1.3. Isotope ratios by IR-MS*

Finally, all reference samples were subjected to IR-MS for analysis of carbon, hydrogen, nitrogen, and sulfur isotope ratios. Results are visualized in a scatter plot (**Figure 3**). Hydrogen isotope ratio (δ<sup>2</sup> H) is severely depleted compared to sea water, ranging from −43.7 to −112.5‰ (versus Vienna Standard Mean Ocean Water). This can be explained by the fact that organic tissue is synthesized from metabolized products and their isotopic composition is referenced to local food chains rather than dissolved water isotopes [36]. Sulfur isotope ratio (δ<sup>34</sup>S) is enriched in European plaice, ranging from +4.6 to +20.1‰. This is typical for marine fish, whereas freshwater fish typically shows depleted δ<sup>34</sup>S [37]. Carbon isotope ratio (δ<sup>13</sup>C) is depleted, ranging from −14.6 to −21.1‰. Nitrogen isotope ratio (δ<sup>15</sup>N) is enriched, ranging from +8.9 to +13.0‰. Both δ<sup>13</sup>C and δ<sup>15</sup>N ratios are in accordance with previous research on flatfish from the southern North Sea and claimed to reflect the European plaices' diet [38].

In both scatter plots, grouping of the samples according to their geographical origin is clearly visible. Only samples from the seas around Ireland and Iceland seem to group together. Nitrogen and hydrogen isotope ratios in the North Sea European plaice were significantly different from the ratios in European plaice from other seas (Kruskal-Wallis P < 0.05). This was expected based on the results previously described on the carbon, hydrogen, nitrogen, and oxygen isotope ratios of various fish species [20, 21].

#### *3.1.4. All data combined*

*3.1.2. FA profiles by GC-FID*

seas (Kruskal-Wallis).

36 Descriptive Food Science

\*

fatty acids.

similar FA profile for European plaice from the North Sea.

Subsequently, the reference set of European plaice was subjected to GC-FID analysis. European plaice contains around 0.8% fat [35]. This was extracted and its FA composition was analyzed. The most abundant FAs in European plaice were palmitic acid (C16:0), eicosapentaenoic acid (EPA, C20:5n3), and docosahexaenoic acid (DHA, C22:6n3) (**Table 2**). Karl et al. [35] found a

SFA = saturated fatty acids, MUFA = monounsaturated fatty acids, PUFA = polyunsaturated fatty acids, OFA = other

**Table 2.** Average fatty acid composition of European plaice sampled from the North Sea (NS), the Baltic Sea (BalS), the Barents Sea (BarS), the sea around Ireland (SIr), and the sea around Iceland (SIc); mean concentration, standard deviation, and significant differences between European plaice from the North Sea and European plaice caught in other

**FA NS (n = 31) BalS (n = 3) BarS (n = 5) SIr (n = 5) SIc (n = 5) P-value NS** 

C24:0 0.15 ± 0.08 0.19 ± 0.05 0.23 ± 0.12 0.15 ± 0.11 0.07 ± 0.05 0.889 C24:1n9 0.23 ± 0.29 0.47 ± 0.07 0.25 ± 0.08 0.52 ± 0.15 0.41 ± 0.12 0.000 C22:6n3 15.78 ± 4.22 13.31 ± 2.72 6.35 ± 1.30 10.60 ± 4.34 11.45 ± 3.24 0.000 SFA\* 26.51 ± 1.62 25.77 ± 0.08 23.50 ± 1.83 28.15 ± 2.00 28.05 ± 0.77 0.788 MUFA 16.31 ± 4.99 24.42 ± 3.93 32.07 ± 4.75 20.10 ± 3.57 21.52 ± 2.78 0.000 PUFA 41.52 ± 6.35 32.75 ± 4.23 25.31 ± 4.74 33.14 ± 6.83 34.53 ± 2.89 0.000 OFA 15.66 ± 3.89 17.06 ± 0.38 19.12 ± 1.90 18.60 ± 5.64 15.90 ± 1.24 0.016 Omega 3 39.05 ± 7.85 27.32 ± 4.17 20.01 ± 3.83 26.58 ± 6.68 31.05 ± 2.66 0.000 Omega 6 1.38 ± 1.04 2.05 ± 0.20 2.12 ± 0.36 2.48 ± 0.74 1.14 ± 0.21 0.002 Omega 3/omega 6 ratio 38.09 ± 18.72 13.51 ± 3.33 9.90 ± 3.66 11.75 ± 5.44 28.21 ± 6.26 0.000

**versus other seas**

When comparing the origins for each FA, 19 out of 35 FAs revealed significant differences between European plaice from the North Sea and European plaice from other seas (Kruskal-Wallis P < 0.05; **Table 2**). European plaice from the Barents Sea seems to have a rather distinct FA profile compared to the other seas. When considering groups of FAs instead of individual FAs, contents of MUFA (monounsaturated FA), PUFA (polyunsaturated FA), OFA (other FA), omega-3 FAs, and omega-6 FAs were significantly different in North Sea European plaice from European plaice originating from other seas (Kruskal-Wallis P < 0.05). Colombo et al. [19] suggested that fish originating from a higher latitude would contain more PUFA. However, this was not confirmed in our study. European plaice from the sea with the highest latitude, the Barents Sea, showed to have the lowest PUFA content, namely 25%. The sea around Ireland,

which has the lowest latitude, showed a rather average PUFA content, namely 33%.

PCA reveals clustering of samples (**Figure 2B**). Generally, the samples from the North Sea and those from the other seas are separated in the second dimension. Therefore, the FA profile seems to be a suitable option to distinguish North Sea European plaice from those All VOC, FA, and isotope ratio data acquired in this study were also combined and subjected to PCA plot (**Figure 2C**). Again, groups of samples from the same geographical origin cluster together, with 80% of the European plaice from the North Sea demonstrating high positive

**Figure 3.** Scatter plot of IRMS data on European plaice: δ<sup>13</sup>C versus δ15N (left) and δ<sup>2</sup> H versus δ<sup>34</sup>S (right) on European plaice from the North Sea (box), the Baltic Sea (dot), the Barents Sea (triangle), the sea around Ireland (diamond), and the sea around Iceland (plus).

relevant information regarding the provenance of European plaice. VOC analysis by PTR-MS allows rapid analysis, but may be influenced by factors like freshness and packaging. FA and isotope ratio analyses are more robust from that perspective, and both allow automated routine analyses. More samples should be collected to provide insight into differences between seasons and for building chemometric classification models. Nevertheless, the approaches are promising for future provenance verification of European plaice. The results may help to underpin sustainable fishing. Sustainable fishing is an important topic for governmental and

Food Authenticity: Provenancing. A Case Study of Fish http://dx.doi.org/10.5772/intechopen.80470 39

In a broader perspective, provenance of food products has become an important topic in food authenticity because of: (a) a quality perspective: the geographical origin of a food product influences its composition and thereby its quality and (b) an environmental perspective: some production locations are more sustainable than others. Consumer awareness on these topics is growing rapidly; they want their food to be of high quality and with a clear geographical

The authors gratefully acknowledge the European Fisheries Fund for financial support. We would also like to thank Francisco de la Granda from Thünen Institute of Baltic Sea Fisheries in Germany, Cat Smith from Daithi O'Murchu Marine Research Station in Ireland, Olafur S. Astthorsson from the Marine Research Institute in Iceland, and André de Vries from the North Sea Fish Centre for collecting the samples. Furthermore, we would like to thank Leon de Jonge for the Animal Nutrition Group from Wageningen University & Research for the analysis of isotope ratios and Martijn Staats from RIKILT for leading the overall project.

The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manu-

, Alex H. Koot1

and Saskia M. van Ruth1,2

\*, Rita Boerrigter-Eenling1

1 RIKILT, Wageningen University and Research, Wageningen, The Netherlands 2 Food Quality and Design, Wageningen University and Research, Wageningen,

\*Address all correspondence to: annemieke.pustjens@wur.nl

possible certification organizations.

identity.

**Acknowledgements**

**Conflict of interest**

**Author details**

The Netherlands

Annemieke M. Pustjens1

script; and in the decision to publish the results.

**Figure 4.** PCA plot of fatty acid profile of European plaice from the North Sea (1—box), the Baltic Sea (2—dot), the Barents Sea (3—triangle), the sea around Ireland (4—diamond), the sea around Iceland (5—inverted triangle), with projection of retail European plaice samples (6—star).

scores in the first dimension and samples from other seas clustering together with the rest from the North Sea and showing negative scores in the first PCA dimension. Distinction of North Sea European plaice from others appears less effective with all data combined in comparison to some of the individual datasets.

#### *3.1.5. Retail European plaice samples*

Eventually, a set of 11 non-breaded and non-marinated European plaice fillets from German, Italian, and Dutch supermarkets were analyzed for their FA profiles and compared with the reference set, since FA profiling showed a good distinction and is a robust method. They were projected on the PCA plot of the FA profiles as a sixth class (**Figure 4**). All retail samples cluster in the same region as European plaice samples from the Baltic Sea, the Barents Sea, and the sea around Ireland. According to their label, they originated from the North Sea specifically (n = 4), or more generally from FAO zone 27 (n = 7). In this phase of the research, it cannot be explained why all retail European plaice samples would cluster together away from the samples from the North Sea. This is possibly caused by the fact that samples were collected in a different year and season. However, more research would be needed to evaluate the seasonal effect on the FA composition.

## **4. Conclusion**

This case study showed that the geographical origin of food products is reflected in its product characteristics, which can be measured by several analytical techniques. From the techniques evaluated in this study, it appears that VOC, FA, and isotope ratios all provide relevant information regarding the provenance of European plaice. VOC analysis by PTR-MS allows rapid analysis, but may be influenced by factors like freshness and packaging. FA and isotope ratio analyses are more robust from that perspective, and both allow automated routine analyses. More samples should be collected to provide insight into differences between seasons and for building chemometric classification models. Nevertheless, the approaches are promising for future provenance verification of European plaice. The results may help to underpin sustainable fishing. Sustainable fishing is an important topic for governmental and possible certification organizations.

In a broader perspective, provenance of food products has become an important topic in food authenticity because of: (a) a quality perspective: the geographical origin of a food product influences its composition and thereby its quality and (b) an environmental perspective: some production locations are more sustainable than others. Consumer awareness on these topics is growing rapidly; they want their food to be of high quality and with a clear geographical identity.

## **Acknowledgements**

scores in the first dimension and samples from other seas clustering together with the rest from the North Sea and showing negative scores in the first PCA dimension. Distinction of North Sea European plaice from others appears less effective with all data combined in com-

**Figure 4.** PCA plot of fatty acid profile of European plaice from the North Sea (1—box), the Baltic Sea (2—dot), the Barents Sea (3—triangle), the sea around Ireland (4—diamond), the sea around Iceland (5—inverted triangle), with

Eventually, a set of 11 non-breaded and non-marinated European plaice fillets from German, Italian, and Dutch supermarkets were analyzed for their FA profiles and compared with the reference set, since FA profiling showed a good distinction and is a robust method. They were projected on the PCA plot of the FA profiles as a sixth class (**Figure 4**). All retail samples cluster in the same region as European plaice samples from the Baltic Sea, the Barents Sea, and the sea around Ireland. According to their label, they originated from the North Sea specifically (n = 4), or more generally from FAO zone 27 (n = 7). In this phase of the research, it cannot be explained why all retail European plaice samples would cluster together away from the samples from the North Sea. This is possibly caused by the fact that samples were collected in a different year and season. However, more research would be needed to evaluate the

This case study showed that the geographical origin of food products is reflected in its product characteristics, which can be measured by several analytical techniques. From the techniques evaluated in this study, it appears that VOC, FA, and isotope ratios all provide

parison to some of the individual datasets.

projection of retail European plaice samples (6—star).

38 Descriptive Food Science

*3.1.5. Retail European plaice samples*

seasonal effect on the FA composition.

**4. Conclusion**

The authors gratefully acknowledge the European Fisheries Fund for financial support. We would also like to thank Francisco de la Granda from Thünen Institute of Baltic Sea Fisheries in Germany, Cat Smith from Daithi O'Murchu Marine Research Station in Ireland, Olafur S. Astthorsson from the Marine Research Institute in Iceland, and André de Vries from the North Sea Fish Centre for collecting the samples. Furthermore, we would like to thank Leon de Jonge for the Animal Nutrition Group from Wageningen University & Research for the analysis of isotope ratios and Martijn Staats from RIKILT for leading the overall project.

## **Conflict of interest**

The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; and in the decision to publish the results.

## **Author details**

Annemieke M. Pustjens1 \*, Rita Boerrigter-Eenling1 , Alex H. Koot1 and Saskia M. van Ruth1,2

\*Address all correspondence to: annemieke.pustjens@wur.nl

1 RIKILT, Wageningen University and Research, Wageningen, The Netherlands

2 Food Quality and Design, Wageningen University and Research, Wageningen, The Netherlands

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[10] Li G, Cervelli C, Ruffoni B, Shachter A, Dudai N. Volatile diversity in wild populations of rosemary (*Rosmarinus officinalis* L.) from the Tyrrhenian Sea vicinity cultivated under homogeneous environmental conditions. Industrial Crops and Products. 2016;**84**:381-390

[11] Yener S, Sánchez-López JA, Granitto PM, Cappellin L, Märk TD, Zimmermann R, et al. Rapid and direct volatile compound profiling of black and green teas (*Camellia sinensis*)

[12] Condurso C, Mazzaglia A, Tripodi G, Cincotta F, Dima G, Maria Lanza C, et al. Sensory analysis and head-space aroma volatiles for the characterization of capers from different

from different countries with PTR-ToF-MS. Talanta. 2016;**152**:45-53

geographic origin. Journal of Essential Oil Research. 2016;**28**(3):185-192

geographical origin of food products. Food Chemistry. 2008;**107**(2):897-911

Reviews in Food Science and Nutrition. 2016;**56**(2):306-317

and Repealing Council Regulation (EC) No 104/2000; 2013

Authenticity Testing. Woodhead Publishing; 2016. pp. 339-367

Science and Biotechnology. 2017;**26**(3):603-609


[26] Engelhard GH, Pinnegar JK, Kell LT, Rijnsdorp AD. Nine decades of North Sea sole and plaice distribution. ICES Journal of Marine Science. 2011;**68**(6):1090-1104

**Chapter 3**

**Provisional chapter**

are being used for the

**Agrifood By-Products as a Source of Phytochemical**

**Agrifood By-Products as a Source of Phytochemical** 

In last years, food by-products and waste valorization practices have gained importance because these processes are sustainable and can increase the profit for local economies. Many compound families of phytochemicals like carotenoids, tocopherols, glucosinolates and phenolic compounds can be obtained through plant by-products coming from agroindustries, such as citric peels, tomato wastes or wine pomace. A number of novel

extraction of compounds, affecting them in different ways. Phytochemicals obtained can be used in cosmetics, medical uses and dietary supplements or reused in agrifood indus-

**Keywords:** phytochemical, valorization, by-products, extraction methods, antioxidant,

It is estimated that one-third of all food produced for human consumption is lost or wasted. The annual global volume of food wastage is estimated to be 1.6 Gtonnes. It seems clear that a reduction of food wastage would have a substantial positive effect on natural and societal resources and that food wastage represents a missed opportunity to improve global food

Nowadays, the agricultural processing industries produce substantial quantities of by-products, which are currently generally treated as waste of industry. This practice is not only a waste of

methods like pressured liquid, microwaves or supercritical CO2

tries among others, as natural pigments, antioxidants or antimicrobials.

security and to mitigate environmental impact generated by agriculture [1].

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.79434

**Compounds**

**Abstract**

antimicrobial

**1. Introduction**

**Compounds**

Jonathan Delgado-Adámez

Jonathan Delgado-Adámez

http://dx.doi.org/10.5772/intechopen.79434

Sara Martillanes, Javier Rocha-Pimienta and

Sara Martillanes, Javier Rocha-Pimienta and

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter


#### **Agrifood By-Products as a Source of Phytochemical Compounds Agrifood By-Products as a Source of Phytochemical Compounds**

DOI: 10.5772/intechopen.79434

Sara Martillanes, Javier Rocha-Pimienta and Jonathan Delgado-Adámez Sara Martillanes, Javier Rocha-Pimienta and Jonathan Delgado-Adámez

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.79434

#### **Abstract**

[26] Engelhard GH, Pinnegar JK, Kell LT, Rijnsdorp AD. Nine decades of North Sea sole and

[27] van Keeken OA, van Hoppe M, Grift RE, Rijnsdorp AD. Changes in the spatial distribution of North Sea plaice (*Pleuronectes platessa*) and implications for fisheries manage-

[28] Petitgas P, Rijnsdorp AD, Dickey-Collas M, Engelhard GH, Peck MA, Pinnegar JK, et al. Impacts of climate change on the complex life cycles of fish. Fisheries Oceanography.

[29] Hansel A, Jordan A, Holzinger R, Prazeller P, Vogel W, Lindinger W. Proton transfer reaction mass spectrometry: On-line trace gas analysis at the ppb level. International

[30] Tres A, Van Ruth SM. Verification of organic feed identity by fatty acid fingerprinting.

[31] Guardiola F, Codony R, Rafecas M, Boatella J, López A. Fatty acid composition and nutritional value of fresh eggs, from large- and small-scale farms. Journal of Food Com-

[32] Olafsdottir G, Lauzon HL, Martinsdóttir E, Oehlenschláuger J, Kristbergsson K. Evaluation of shelf life of superchilled cod (*Gadus morhua*) fillets and the influence of temperature fluctuations during storage on microbial and chemical quality indicators. Journal

[33] Phan N-T, Kim K-H, Jeon E-C, Kim U-H, Sohn JR, Pandey SK.Analysis of volatile organic compounds released during food decaying processes. Environmental Monitoring and

[34] Parlapani FF, Mallouchos A, Haroutounian SA, Boziaris IS. Volatile organic compounds of microbial and non-microbial origin produced on model fish substrate un-inoculated and inoculated with gilt-head sea bream spoilage bacteria. LWT- Food Science and

[35] Karl H, Manthey-Karl M, Ostermeyer U, Lehmann I, Wagner H. Nutritional composition and sensory attributes of Alaskan flatfishes compared to plaice (*Pleuronectes platessa*).

[36] Trueman CN, Mackenzie KM, Palmer MR. Identifying migrations in marine fishes

[37] Sayle KL, Cook GT, Ascough PL, Hastie HR, Einarsson T, McGovern TH, et al. Application of 34S analysis for elucidating terrestrial, marine and freshwater ecosystems: Evidence of animal movement/husbandry practices in an early Viking community around Lake

[38] Fuller BT, Müldner G, Van Neer W, Ervynck A, Richards MP. Carbon and nitrogen stable isotope ratio analysis of freshwater, brackish and marine fish from Belgian archaeological sites (1st and 2nd millennium AD). Journal of Analytical Atomic Spectrometry.

International Journal of Food Science and Technology. 2013;**48**(5):962-971

through stable-isotope analysis. Journal of Fish Biology. 2012;**81**(2):826-847

Mývatn, Iceland. Geochimica et Cosmochimica Acta. 2013;**120**:531-544

Journal of Agricultural and Food Chemistry. 2011;**59**(16):8816-8821

plaice distribution. ICES Journal of Marine Science. 2011;**68**(6):1090-1104

ment. Journal of Sea Research. 2007;**57**(2):187-197

Journal of Mass Spectrometry. 1995;**149-150**:609-619

position and Analysis. 1994;**7**(3):171-188

of Food Science. 2006;**71**(2):S97-S109

Assessment. 2012;**184**(3):1683-1692

Technology. 2017;**78**:54-62

2012;**27**(5):807-820

2013;**22**(2):121-139

42 Descriptive Food Science

In last years, food by-products and waste valorization practices have gained importance because these processes are sustainable and can increase the profit for local economies. Many compound families of phytochemicals like carotenoids, tocopherols, glucosinolates and phenolic compounds can be obtained through plant by-products coming from agroindustries, such as citric peels, tomato wastes or wine pomace. A number of novel methods like pressured liquid, microwaves or supercritical CO2 are being used for the extraction of compounds, affecting them in different ways. Phytochemicals obtained can be used in cosmetics, medical uses and dietary supplements or reused in agrifood industries among others, as natural pigments, antioxidants or antimicrobials.

**Keywords:** phytochemical, valorization, by-products, extraction methods, antioxidant, antimicrobial

#### **1. Introduction**

It is estimated that one-third of all food produced for human consumption is lost or wasted. The annual global volume of food wastage is estimated to be 1.6 Gtonnes. It seems clear that a reduction of food wastage would have a substantial positive effect on natural and societal resources and that food wastage represents a missed opportunity to improve global food security and to mitigate environmental impact generated by agriculture [1].

Nowadays, the agricultural processing industries produce substantial quantities of by-products, which are currently generally treated as waste of industry. This practice is not only a waste of

> © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

resource, but also causes environmental pollution and they gradually permeant and release odors. The valorization of agricultural and food residues via green chemistry technologies is among the most important objectives of contemporary chemical research. The environmental benefits reduce waste production, reduce energy demand economical grows, thanks to the significant economic value of many bioproducts new revenues made available to farmers and job creation.

have been used in perfumery from the earliest times. The main terpenoids shown in nature are: myrcene, ocimene, citral, geraniol, eugenol, carvacrol, linanool, citronellal, carvone, limonene, terpinenes, menthol and menthone, the carane group and the pinane group, among others [3]. The main by-product source of essential oils is the citrus juice processing residues. Tetraterpenoids (carotenoids): the carotenoids are a large group of pigments widely in animal and vegetable kingdoms; they produce colors ranging from yellow to deep red. Chemically, carotenoids are divided into two groups: the carotenes, which are hydrocarbons, and the xanthophylls, their oxygenated derivatives. Among the carotenes, very soluble in no-polar solvents, are α, β and γ-carotenes, and lycopene. The xanthophylls may present as acids, aldehydes or alcohols. Examples of these compounds are the cryptoxanthin, fucoxanthin, lutein and zeaxanthin. Currently, a high proportion of carotenoids is obtained synthetically, since it is cheaper; however, increasingly those of natural origin are used more [4]. Carotenes can be obtained from orange and reddish sources, as carrot waste or tomato pomace, while xantho-

Agrifood By-Products as a Source of Phytochemical Compounds

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45

Polyphenols or phenol compounds are organic compounds whose molecular structures contain at least one phenol group; an aromatic ring at least joined a hydroxyl group. Many are classified as secondary metabolites of plants, products synthesized in plants, which possess the biological characteristic of being secondary products of their metabolism. They are generally synthesized by one of two biosynthetic pathways: the path of the shikimic acid or via

Phenolic compounds in plants are a heterogeneous group of products with more than 10,000 compounds. Some are soluble in solvents organic, others are glycosides or carboxylic acids

This group also plays a very heterogeneous range of roles in plants, roles that are generally attributed to the by-products of plants: many are products of defense against herbivores and pathogens; others provide mechanical support to the plant, other they attract pollinators or dispersers of fruits, some of them absorb the ultraviolet radiation or act as allelopathic agents. In humans, these compounds are well known to possess healthy benefits, and the interest in food phenolics has increased due to their antioxidant and free radical-scavenging abilities [6]. Polyphenols can be structurally classified into several groups: flavonoids, lignans, stilbenes,

Flavonoids are one of the most studied families, since about 4000 different structures are known. Being a numerous group, they can be subdivided into: flavonols, flavanones, flavanols, flavones, and isoflavones anthocyanidins. Flavonoids have antioxidant and other livingbody modulating activities [7]. For its part, phenolic acids can be classified as hydroxybenzoic acids and hydroxycinnamic acids, which are derived from non-phenolic molecules of benzoic and cinnamic acid, respectively. Phenols are present in numerous products, and collection has been studied in by-products of the obtaining of different oils, wine, juices, coffee, as well

and therefore soluble in water, and others are very large and insoluble polymers.

phylls can be extracted from spinach residues [5].

**2.2. Phenolic compounds**

and phenolic acids.

as saved cereal among others.

malonic acid (or two, e.g., flavonoids).

## **2. Classification of phytochemicals and sources**

Phytochemicals are natural chemical compounds, which are found in foods derived from plants. In these, phytochemical substances act as systems of natural defenses for their plants, protecting them from infections and microbial invasions and guests giving them color, aroma and flavor. Phytochemicals are not considered essential for our organism; however, most of them have beneficial properties for health. The main sources of these compounds in foods are fruits, vegetables, legumes, whole grains, nuts, seeds, mushrooms, herbs and spices.

There are more than 2000 phytochemicals in plants, which can be grouped according to their structural characteristics in four large groups: terpenoids, phenolic compounds, nitrogen compounds and sulfur compounds.

#### **2.1. Terpenoids**

Terpenoids also referred as isoprenoids are a large and diverse family of organic compounds similar to terpenes. Terpenoids are made up of five-carbon isoprene units, assembled and modified in many different ways, always based on the skeleton of the isopentane. Majority of terpenoids have multicyclical structures, which differ from each other not only in functional group but also in their basic carbon skeletons.

These compounds are found in all kinds of living beings, and are biosynthesized in plants, where they are important in numerous biotic interactions [2]. In plants, the terpenoids fulfill many primary functions: some carotenoid pigments are formed by terpenoids; they are also part of the chlorophyll and gibberellin hormones among others. Steroids and sterols are produced from terpenoids precursors.

Terpenoids of the plants are widely used for their aromatic qualities. They play an important role in traditional medicine and herbal remedies, and their possible antibacterial effects and other pharmaceutical uses are being investigated. They are present, for example, in the essences of eucalyptus, the flavors of clove and ginger. The biosynthesis of terpenoids in plants is through the mevalonic acid pathway.

Terpenoids family comprises very different compounds, of which, the most interesting in food can be classified in the following subgroups:

Mono- and sesquiterpenoids: are the chief constituents of the essential oils; these are the volatile oils obtained from the sap and tissues of certain plants and trees. The essential oils have been used in perfumery from the earliest times. The main terpenoids shown in nature are: myrcene, ocimene, citral, geraniol, eugenol, carvacrol, linanool, citronellal, carvone, limonene, terpinenes, menthol and menthone, the carane group and the pinane group, among others [3]. The main by-product source of essential oils is the citrus juice processing residues.

Tetraterpenoids (carotenoids): the carotenoids are a large group of pigments widely in animal and vegetable kingdoms; they produce colors ranging from yellow to deep red. Chemically, carotenoids are divided into two groups: the carotenes, which are hydrocarbons, and the xanthophylls, their oxygenated derivatives. Among the carotenes, very soluble in no-polar solvents, are α, β and γ-carotenes, and lycopene. The xanthophylls may present as acids, aldehydes or alcohols. Examples of these compounds are the cryptoxanthin, fucoxanthin, lutein and zeaxanthin. Currently, a high proportion of carotenoids is obtained synthetically, since it is cheaper; however, increasingly those of natural origin are used more [4]. Carotenes can be obtained from orange and reddish sources, as carrot waste or tomato pomace, while xanthophylls can be extracted from spinach residues [5].

#### **2.2. Phenolic compounds**

resource, but also causes environmental pollution and they gradually permeant and release odors. The valorization of agricultural and food residues via green chemistry technologies is among the most important objectives of contemporary chemical research. The environmental benefits reduce waste production, reduce energy demand economical grows, thanks to the significant economic value of many bioproducts new revenues made available to farmers and job

Phytochemicals are natural chemical compounds, which are found in foods derived from plants. In these, phytochemical substances act as systems of natural defenses for their plants, protecting them from infections and microbial invasions and guests giving them color, aroma and flavor. Phytochemicals are not considered essential for our organism; however, most of them have beneficial properties for health. The main sources of these compounds in foods are

There are more than 2000 phytochemicals in plants, which can be grouped according to their structural characteristics in four large groups: terpenoids, phenolic compounds, nitrogen

Terpenoids also referred as isoprenoids are a large and diverse family of organic compounds similar to terpenes. Terpenoids are made up of five-carbon isoprene units, assembled and modified in many different ways, always based on the skeleton of the isopentane. Majority of terpenoids have multicyclical structures, which differ from each other not only in functional

These compounds are found in all kinds of living beings, and are biosynthesized in plants, where they are important in numerous biotic interactions [2]. In plants, the terpenoids fulfill many primary functions: some carotenoid pigments are formed by terpenoids; they are also part of the chlorophyll and gibberellin hormones among others. Steroids and sterols are pro-

Terpenoids of the plants are widely used for their aromatic qualities. They play an important role in traditional medicine and herbal remedies, and their possible antibacterial effects and other pharmaceutical uses are being investigated. They are present, for example, in the essences of eucalyptus, the flavors of clove and ginger. The biosynthesis of terpenoids in

Terpenoids family comprises very different compounds, of which, the most interesting in

Mono- and sesquiterpenoids: are the chief constituents of the essential oils; these are the volatile oils obtained from the sap and tissues of certain plants and trees. The essential oils

fruits, vegetables, legumes, whole grains, nuts, seeds, mushrooms, herbs and spices.

**2. Classification of phytochemicals and sources**

compounds and sulfur compounds.

duced from terpenoids precursors.

group but also in their basic carbon skeletons.

plants is through the mevalonic acid pathway.

food can be classified in the following subgroups:

**2.1. Terpenoids**

creation.

44 Descriptive Food Science

Polyphenols or phenol compounds are organic compounds whose molecular structures contain at least one phenol group; an aromatic ring at least joined a hydroxyl group. Many are classified as secondary metabolites of plants, products synthesized in plants, which possess the biological characteristic of being secondary products of their metabolism. They are generally synthesized by one of two biosynthetic pathways: the path of the shikimic acid or via malonic acid (or two, e.g., flavonoids).

Phenolic compounds in plants are a heterogeneous group of products with more than 10,000 compounds. Some are soluble in solvents organic, others are glycosides or carboxylic acids and therefore soluble in water, and others are very large and insoluble polymers.

This group also plays a very heterogeneous range of roles in plants, roles that are generally attributed to the by-products of plants: many are products of defense against herbivores and pathogens; others provide mechanical support to the plant, other they attract pollinators or dispersers of fruits, some of them absorb the ultraviolet radiation or act as allelopathic agents. In humans, these compounds are well known to possess healthy benefits, and the interest in food phenolics has increased due to their antioxidant and free radical-scavenging abilities [6]. Polyphenols can be structurally classified into several groups: flavonoids, lignans, stilbenes, and phenolic acids.

Flavonoids are one of the most studied families, since about 4000 different structures are known. Being a numerous group, they can be subdivided into: flavonols, flavanones, flavanols, flavones, and isoflavones anthocyanidins. Flavonoids have antioxidant and other livingbody modulating activities [7]. For its part, phenolic acids can be classified as hydroxybenzoic acids and hydroxycinnamic acids, which are derived from non-phenolic molecules of benzoic and cinnamic acid, respectively. Phenols are present in numerous products, and collection has been studied in by-products of the obtaining of different oils, wine, juices, coffee, as well as saved cereal among others.


type of molecule extracted, as well as the use that will be given to extract thus obtained. The conventional techniques used include maceration, infusion, decoction or Soxhlet extraction among others [10]. However, since the main objective of the extraction is the reuse of waste and reduce environmental impact, green extraction techniques are necessary for this purpose. The objective of these techniques is to improve extraction performance, decreasing the use of

Agrifood By-Products as a Source of Phytochemical Compounds

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47

Ultrasound is a mechanical wave that propagates in an elastic medium and their frequency is above the audible sounds, this is more than 20,000 Hz. The ultrasound-assisted extraction is a technique that uses ultrasonic waves to agitate an immersed sample in an organic solvent. The major effects of ultrasound in a liquid medium are attributed to the cavitation phenomena, which comes from the physical processes that create, enlarge, and implode microbubbles of

Since the extraction is carried out in a medium, the temperature, time and solvent-type can affect not only the extraction yield but also the composition of the extract and should thus be taken into consideration. Furthermore, after extraction, compounds with organic solvent are separated from the matrix by centrifugation or filtration like in a conventional extraction. The greatest advantage of using this technique against conventional techniques is that this technology shortens the total time of procedure, together with a decrease of consumed energy and pollution. For that, the procedures using ultrasound assistance have a production cost and a functioning cost much lower than the cost for conventional procedures with a high purity final product. On the other hand, as the temperature reached is not very high, sonication is suitable for extracting thermolabile compounds [12]. However, and despite the fact that this technology can be easily integrated into the industry, its use implies initial costs,

The application of this technology for the extraction of phytochemicals from by-product has been widely researched, but its application differs according to the various matrices and analytes to extract. Thus, terpenoids have been obtained by UAE from different byproducts: The industrial extraction of terpenoids limonin, nomolin and obakunone were performed from lemon seeds with UAE, obtaining high extraction yields of limonoids [13]. In case of lycopene, tomato paste processing wastes are an important source of obtainment; the UAE has demonstrated to decrease the solvent-solid ratio and to require less time and lower temperature than conventional organic solvent extraction [14]. Papaya processing waste is also studied to lycopene extraction. In this case, results indicated that UAE was the most effective extracting method among the conventional and Soxhlet method [15]. In addition, the extraction of carotenoids without solvent also can be improved with the use of ultrasound, as is the case with the extraction of carotenoids from pomegranate peel using

Different wastes have been used for the ultrasound-assisted extraction of phenolic compounds. Mandarin peel has been utilized for the extraction of flavonoids and hesperidin comparing the results with maceration extraction. These showed greater extraction efficiency

organic solvents or the energy used. These techniques include:

**3.1. Ultrasound-assisted extraction (UAE)**

gases dissolved in the solvent [11].

sunflower oil [16].

which range between 10,000 and 200,000 euros [11].

**Table 1.** Classification of phytochemical and main sources of obtainment in agroindustrial by-products.

#### **2.3. Nitrogen alkaloids**

They are a large family of secondary metabolites of plants, which chemically have three characteristics in common: they are soluble in water, contain at least one atom of nitrogen and have biological activity [8]. In plants, alkaloids seem to offer protection from insects and herbivores. In humans, the alkaloids produce physiological and psychological responses, most of them because of their interaction with neurotransmitters. The alkaloids are classified according to the rings present in the molecule. Some of the alkaloids present in foods are of interest as piperine, caffeine and theobromine, from the pepper, coffee and cocoa, respectively.

#### **2.4. Organosulfur compounds**

The organosulfur compounds or thiols are chemical compounds containing sulfur in their structure. They are present in garlic and vegetables of genus cruciferous (cabbage, turnips and members of the mustard family). They include the glucosinolates and allyl sulphides.

Glucosinolates are metabolites characteristic of the *Brassicales* order. Many of us are familiar with the characteristic pungent flavor of wasabi and mustard. Chemically, glucosinolates are organic anions with an amino acid-derived side chain and oxime moiety with an S-linked thioglucose and an O-linked sulfate group [9]. Allyl sulphides are a very important phytochemical group present in garlic.

Main sources of obtainment of these compounds in agrifood by-products are summarized in **Table 1**.

#### **3. Extraction processes**

Numerous methods of extraction have been developed for obtaining phytochemical compounds. The use of one or other will depend on both type of matrix (solid or liquid) and type of molecule extracted, as well as the use that will be given to extract thus obtained. The conventional techniques used include maceration, infusion, decoction or Soxhlet extraction among others [10]. However, since the main objective of the extraction is the reuse of waste and reduce environmental impact, green extraction techniques are necessary for this purpose. The objective of these techniques is to improve extraction performance, decreasing the use of organic solvents or the energy used. These techniques include:

#### **3.1. Ultrasound-assisted extraction (UAE)**

**2.3. Nitrogen alkaloids**

46 Descriptive Food Science

**Group Subgroup Sources** Terpenoids Mono- and sesquiterpenoid Citric peels

**2.4. Organosulfur compounds**

chemical group present in garlic.

**3. Extraction processes**

**Table 1**.

They are a large family of secondary metabolites of plants, which chemically have three characteristics in common: they are soluble in water, contain at least one atom of nitrogen and have biological activity [8]. In plants, alkaloids seem to offer protection from insects and herbivores. In humans, the alkaloids produce physiological and psychological responses, most of them because of their interaction with neurotransmitters. The alkaloids are classified according to the rings present in the molecule. Some of the alkaloids present in foods are of interest as piperine, caffeine and theobromine, from the pepper, coffee and cocoa, respectively.

Carotenoids Carrot, tomato and spinach wastes

soybean flour

Phenolic acids Pomegranate and grape by-products, rice bran

Phenolic compounds Flavonoids Onion skin waste, citric peels, grape skins, olive leaf,

Lignans Sesame cake Stilbenes Wine pomace

Organosulfur compounds Glucosinolates Cauliflower and broccoli by-product Allyl sulphides Garlic

**Table 1.** Classification of phytochemical and main sources of obtainment in agroindustrial by-products.

Nitrogen alkaloids — Cocoa and coffee wastes

The organosulfur compounds or thiols are chemical compounds containing sulfur in their structure. They are present in garlic and vegetables of genus cruciferous (cabbage, turnips and members of the mustard family). They include the glucosinolates and allyl sulphides.

Glucosinolates are metabolites characteristic of the *Brassicales* order. Many of us are familiar with the characteristic pungent flavor of wasabi and mustard. Chemically, glucosinolates are organic anions with an amino acid-derived side chain and oxime moiety with an S-linked thioglucose and an O-linked sulfate group [9]. Allyl sulphides are a very important phyto-

Main sources of obtainment of these compounds in agrifood by-products are summarized in

Numerous methods of extraction have been developed for obtaining phytochemical compounds. The use of one or other will depend on both type of matrix (solid or liquid) and Ultrasound is a mechanical wave that propagates in an elastic medium and their frequency is above the audible sounds, this is more than 20,000 Hz. The ultrasound-assisted extraction is a technique that uses ultrasonic waves to agitate an immersed sample in an organic solvent. The major effects of ultrasound in a liquid medium are attributed to the cavitation phenomena, which comes from the physical processes that create, enlarge, and implode microbubbles of gases dissolved in the solvent [11].

Since the extraction is carried out in a medium, the temperature, time and solvent-type can affect not only the extraction yield but also the composition of the extract and should thus be taken into consideration. Furthermore, after extraction, compounds with organic solvent are separated from the matrix by centrifugation or filtration like in a conventional extraction.

The greatest advantage of using this technique against conventional techniques is that this technology shortens the total time of procedure, together with a decrease of consumed energy and pollution. For that, the procedures using ultrasound assistance have a production cost and a functioning cost much lower than the cost for conventional procedures with a high purity final product. On the other hand, as the temperature reached is not very high, sonication is suitable for extracting thermolabile compounds [12]. However, and despite the fact that this technology can be easily integrated into the industry, its use implies initial costs, which range between 10,000 and 200,000 euros [11].

The application of this technology for the extraction of phytochemicals from by-product has been widely researched, but its application differs according to the various matrices and analytes to extract. Thus, terpenoids have been obtained by UAE from different byproducts: The industrial extraction of terpenoids limonin, nomolin and obakunone were performed from lemon seeds with UAE, obtaining high extraction yields of limonoids [13]. In case of lycopene, tomato paste processing wastes are an important source of obtainment; the UAE has demonstrated to decrease the solvent-solid ratio and to require less time and lower temperature than conventional organic solvent extraction [14]. Papaya processing waste is also studied to lycopene extraction. In this case, results indicated that UAE was the most effective extracting method among the conventional and Soxhlet method [15]. In addition, the extraction of carotenoids without solvent also can be improved with the use of ultrasound, as is the case with the extraction of carotenoids from pomegranate peel using sunflower oil [16].

Different wastes have been used for the ultrasound-assisted extraction of phenolic compounds. Mandarin peel has been utilized for the extraction of flavonoids and hesperidin comparing the results with maceration extraction. These showed greater extraction efficiency and lower times of extraction with UAE method [17]. On the other hand, the pomace formed in the wine or juice making from blueberry contains many phenolic and other bioactive compounds. These can be retrieved through ultrasound-assisted extraction obtaining the same compounds than in a conventional solvent extraction but with higher yields of extraction [18]. Spent coffee grounds are a valuable source of phenolic compounds, of which, the main are chlorogenic and protocatechuic acids. These can be extracted with an ultrasound-assisted solid-liquid extraction with mild temperatures and short times [19].

based on prior treatment of the matrix with the corresponding enzyme followed by a process of extraction solvent [28]. This non-traditional and environmentally friendly technology improve the yield of target compounds while reduces the use of solvents in the process. This is possible because enzymes can catalyze reactions in aqueous solutions under mild conditions. The use of enzymes for the extraction is based on their ability to degrade cell walls and membranes, thereby increasing cell wall permeability and enabling targeted compounds

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49

This involves the use of hydrolytic enzymes to alter the cell, mainly composed of large polymer walls highly complex, such as cellulose, hemicellulose, lignin, and pectin [27]. Specific use of enzymes can increase use pre-treatment effects or decrease the amount of solvent, as well as increase the yield of extracted compounds. Enzymes such as cellulases, pectinases or hemicellulases are widely used in juice processing and clarification of beer for cell wall degradation. When matrix have been previously treated with enzymes, it is gotten the cell walls break, thus increasing the performance of the extraction of bioactive

The use of EAE can be tested and optimized on the laboratory scale, and the use of common food-grade enzymes makes it a low-cost technique for extraction purposes. However, some technical limitations on the industry application are a higher relative cost for processing and

EAE has been tested for improve the extractions of carotenoids from tomato peels [29]. Therefore, the use of hydrolytic enzymes can improve the recovery of bound and free phenols from pomegranate peels [30]. EAE has been used successfully for the recovery of polyphenols in citrus peel and ginger [31] winemaking by-products [32]; underutilized watermelon rind

It is a technique that uses a solvent in supercritical conditions. A substance at a pressure higher than its critical pressure and temperature higher than its critical temperature is known as supercritical fluid. Supercritical fluids have properties intermediate between a gas and a liquid, favoring its penetration in different matrices, and therefore the solubilization of solutes, having the possibility of extracting thermolabile compounds. Nowadays, the most important application of SFE to food industry is the extraction of caffeine from

The supercritical fluid extraction consists of four stages [10]: pressurization step, temperature

Advantages of using this technology are the easy separation between solvent and matrix that avoids to increase the temperature to remove the solvent; the possibility to work with nontoxic solvent and the reuse of this, and a relatively low-cost obtainment of high pressure or temperature. However, its practical application is limited to processes that are not affected by

the difficulty to optimize the process at industrial scale [10].

[33] or cauliflower outer leaves [34]; among others.

adjustment stage, extraction stage and separation step.

the relatively high temperature necessaries and the initial costs are high.

**3.4. Supercritical fluid extraction (SFE)**

release into the medium.

compounds [28].

coffee.

For its part, isothiocyanates extraction has been optimized from cauliflower by-products using ultrasound-assisted extraction. These extracts can be added to an apple beverage until 10% preserving well the sensorial properties to obtain a new functional drink [20].

#### **3.2. Microwave-assisted extraction (MAE)**

Microwave-assisted extraction (MAE) is a relatively new extraction technique that combines microwave irradiation and traditional solvent extraction. Microwave energy can be used to improve the extraction of compounds soluble in specific fluids (liquids or gases) as a result of changes in the cell structure caused by electromagnetic waves. It provides rapid and selective techniques with best recoveries than the obtained in conventional extraction processes [21]. Also offers other advantages over common technologies, such as lower energy consumption, volume of solvent and toxicity of the solvents used and, in general, less waste. Another advantage of MAE is that the quicker heating occurring inside the solids where the dissolution of the extract components takes place.

In general, the use of microwave is widespread at laboratory scale. However, its use in the industry offers some disadvantages such as the high initial cost, maintenance, or safety aspects [22]. Moreover, since its use is based on the warming of the matrix and many of the phytochemicals of interest are thermolabile, its use for this purpose is limited. However, there are numerous studies that have been carried out using MAE, both in the pre-treatment of the sample and to facilitate the extraction. For example, the extraction of β-carotene and other carotenoids contained in carrot residues has been studied using intermittent microwave radiation. The thermal degradation caused by MAE in β-carotene was investigated using for that the measurement of antioxidant activity [23]. For the extraction of phenolic compounds from dried waste grape skins, MAE achieves a savings of 83% with respect to conventional in extraction time [24]. Microwave-assisted extraction has also been investigated for the extraction of phenolics from other different matrix as *Eucalyptus robusta* [25] or green tea leaves [21].

The combined use of microwave-assisted and ultrasound-assisted extraction has also been developed for the extraction of phytochemicals. For example, it has been proposed the extraction of essential oils, polyphenols and pectin from the orange peels waste in a solvent-free process [26].

#### **3.3. Enzymatic-assisted extraction (EAE)**

A procedure widely used to improve the efficiency of the extraction of compounds from a plant matrix is based on the enzymatic assisted extraction (EAE) [27]. This procedure is based on prior treatment of the matrix with the corresponding enzyme followed by a process of extraction solvent [28]. This non-traditional and environmentally friendly technology improve the yield of target compounds while reduces the use of solvents in the process. This is possible because enzymes can catalyze reactions in aqueous solutions under mild conditions. The use of enzymes for the extraction is based on their ability to degrade cell walls and membranes, thereby increasing cell wall permeability and enabling targeted compounds release into the medium.

This involves the use of hydrolytic enzymes to alter the cell, mainly composed of large polymer walls highly complex, such as cellulose, hemicellulose, lignin, and pectin [27]. Specific use of enzymes can increase use pre-treatment effects or decrease the amount of solvent, as well as increase the yield of extracted compounds. Enzymes such as cellulases, pectinases or hemicellulases are widely used in juice processing and clarification of beer for cell wall degradation. When matrix have been previously treated with enzymes, it is gotten the cell walls break, thus increasing the performance of the extraction of bioactive compounds [28].

The use of EAE can be tested and optimized on the laboratory scale, and the use of common food-grade enzymes makes it a low-cost technique for extraction purposes. However, some technical limitations on the industry application are a higher relative cost for processing and the difficulty to optimize the process at industrial scale [10].

EAE has been tested for improve the extractions of carotenoids from tomato peels [29]. Therefore, the use of hydrolytic enzymes can improve the recovery of bound and free phenols from pomegranate peels [30]. EAE has been used successfully for the recovery of polyphenols in citrus peel and ginger [31] winemaking by-products [32]; underutilized watermelon rind [33] or cauliflower outer leaves [34]; among others.

#### **3.4. Supercritical fluid extraction (SFE)**

and lower times of extraction with UAE method [17]. On the other hand, the pomace formed in the wine or juice making from blueberry contains many phenolic and other bioactive compounds. These can be retrieved through ultrasound-assisted extraction obtaining the same compounds than in a conventional solvent extraction but with higher yields of extraction [18]. Spent coffee grounds are a valuable source of phenolic compounds, of which, the main are chlorogenic and protocatechuic acids. These can be extracted with an ultrasound-assisted

For its part, isothiocyanates extraction has been optimized from cauliflower by-products using ultrasound-assisted extraction. These extracts can be added to an apple beverage until

Microwave-assisted extraction (MAE) is a relatively new extraction technique that combines microwave irradiation and traditional solvent extraction. Microwave energy can be used to improve the extraction of compounds soluble in specific fluids (liquids or gases) as a result of changes in the cell structure caused by electromagnetic waves. It provides rapid and selective techniques with best recoveries than the obtained in conventional extraction processes [21]. Also offers other advantages over common technologies, such as lower energy consumption, volume of solvent and toxicity of the solvents used and, in general, less waste. Another advantage of MAE is that the quicker heating occurring inside the solids where the dissolution of

In general, the use of microwave is widespread at laboratory scale. However, its use in the industry offers some disadvantages such as the high initial cost, maintenance, or safety aspects [22]. Moreover, since its use is based on the warming of the matrix and many of the phytochemicals of interest are thermolabile, its use for this purpose is limited. However, there are numerous studies that have been carried out using MAE, both in the pre-treatment of the sample and to facilitate the extraction. For example, the extraction of β-carotene and other carotenoids contained in carrot residues has been studied using intermittent microwave radiation. The thermal degradation caused by MAE in β-carotene was investigated using for that the measurement of antioxidant activity [23]. For the extraction of phenolic compounds from dried waste grape skins, MAE achieves a savings of 83% with respect to conventional in extraction time [24]. Microwave-assisted extraction has also been investigated for the extraction of phenolics from other different matrix as *Eucalyptus robusta* [25] or green tea leaves [21]. The combined use of microwave-assisted and ultrasound-assisted extraction has also been developed for the extraction of phytochemicals. For example, it has been proposed the extraction of essential oils, polyphenols and pectin from the orange peels waste in a solvent-free process [26].

A procedure widely used to improve the efficiency of the extraction of compounds from a plant matrix is based on the enzymatic assisted extraction (EAE) [27]. This procedure is

10% preserving well the sensorial properties to obtain a new functional drink [20].

solid-liquid extraction with mild temperatures and short times [19].

**3.2. Microwave-assisted extraction (MAE)**

48 Descriptive Food Science

the extract components takes place.

**3.3. Enzymatic-assisted extraction (EAE)**

It is a technique that uses a solvent in supercritical conditions. A substance at a pressure higher than its critical pressure and temperature higher than its critical temperature is known as supercritical fluid. Supercritical fluids have properties intermediate between a gas and a liquid, favoring its penetration in different matrices, and therefore the solubilization of solutes, having the possibility of extracting thermolabile compounds. Nowadays, the most important application of SFE to food industry is the extraction of caffeine from coffee.

The supercritical fluid extraction consists of four stages [10]: pressurization step, temperature adjustment stage, extraction stage and separation step.

Advantages of using this technology are the easy separation between solvent and matrix that avoids to increase the temperature to remove the solvent; the possibility to work with nontoxic solvent and the reuse of this, and a relatively low-cost obtainment of high pressure or temperature. However, its practical application is limited to processes that are not affected by the relatively high temperature necessaries and the initial costs are high.

In the majority of cases, the supercritical fluid used is CO<sup>2</sup> , for being generally recognized as safe (GRAS). The application of supercritical CO<sup>2</sup> has high performances for the extraction of nonpolar substances. However, the CO<sup>2</sup> under critical conditions is a poor solvent for polar compounds. This limitation can be solved by adding co-solvents that alter the polarity of the CO2 , but they may become contained, requiring a subsequent separation operation.

SFE has been used for the extraction of lycopene β-carotene from tomato peels [35] and carrot peels [36]. For its part, the recovery of phenol by SFE has been tested in apple pomace [37], cacao pod husk [38], mango by-products [39] or sour cherry pomace [40].

## **3.5. Pressurized liquid extraction (PLE)**

The so-called pressurized solvent extraction is a technique that combines solvent extraction at temperatures (50–200°C) and high pressures (1500–2000 psi) to quickly and efficiently extract compounds from solid matrices. The use of liquid solvents at high temperatures and pressures improves the performance of the extractions since the solubility is increased and the mass transfer is improved and the rupture of the composite-matrix superficial equilibrium is facilitated [41]. PLE can be considered as a green extraction process especially when a nontoxic solvent is used.

**4. Industry phytochemical residue applications**

Combined extraction processes The possible synergic effect of techniques

**Table 2.** Overview of the improvement of the main green extraction techniques.

**Extraction method Improvement of method**

plants with more concentration of these compounds.

some of the most important applications in detail.

**4.1. Phytochemicals in medicine and pharmacy**

is viewed as a form of traditional medicine.

for food industry producers, as it generates an economic advantage [47].

permeability

Pressurized liquid extraction (PLE) The solubility is increased and the mass transfer is improved

As stated in previous sections, phytochemicals are present in foods of plant origin. The food industry is considered a source of extraction of phytochemical compounds, which can be obtained from waste generated in the processing of fruit and vegetables. This fact is beneficial

Ultrasound-assisted extraction (UAE) Cavitation phenomena, which comes from the physical processes that create,

Microwaves-assisted extraction (MAE) The quicker heating occurring inside the solids where the dissolution of the extract components takes place Enzyme-assisted extraction (EAE) Ability of enzymes to degrade cell walls and membranes, increasing cell wall

Supercritical fluid extraction (SFE) Supercritical fluids have properties intermediate between a gas and a liquid, favoring its penetration in different matrices

enlarge, and implode microbubbles of gases dissolved in the solvent

Agrifood By-Products as a Source of Phytochemical Compounds

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51

It is worth mentioning that in its elaboration process, the less handling and processing of food generate a more retain of these compounds. For this reason, processes like a cereal refining, intense and long heating, cocking in broths which are then discarded and some of its compounds are lost, reduce the food phytochemical content. In nature, phytochemicals are in many foods, but in the future, the development of bioengineering will allow us to create

In reference to its possible applications, the great phytochemicals properties allow us to open a wide range of possibilities. For example, due to its antioxidant activity, these could be used in creams, functional foods and fortified foods formulation, or due to its antimicrobial activity, these could be used in food conservation and medicine. In the following sections, we see

The phytochemical application in medicine and pharmacy is directly related with the phytotherapy (use of plant-derived medications in the treatment and prevention of disease). The large quantity of properties that the plants has allowed us to apply them for treatments of diseases, and this is due to their phytochemical content. Numerous trials and pharmacological studies of specific phytotherapeutic preparations exist but, in some countries, phytotherapy

A wide range of organic nonpolar to polar solvent and their mixtures have been used in pressurized liquid extraction of phytochemical. However, it is a technology that modifies solvent properties. This fact makes water suitable for the extraction to polar and nonpolar organic compounds.

PLE could be carried out in static and dynamic modes. Static extraction is considered more efficient because of the greater penetration of the solvent into the pores of the source [10]. The advantages of this method with respect to conventional ones are the short extraction times, the yields obtained and the reduced usage of solvents. However, this method is not suitable for thermolabile compounds because the high temperature can alter the structure of these. For that, the use of this technique for the extraction of phytochemical is very limited.

Some researches have been carried out with pressurized liquid for the extraction of carotenoids from shrimp waste [42] or phenolic acid in potato peels [43].

#### **3.6. Other technologies**

In addition to the above, other extraction technologies have been developed as the instant controlled pressure drop or pulsed electric field. The latter has been tested for the extraction of phenolic compounds from orange peel [44] or grape pomace [45] and alkaloids form potato peels [46]. Finally, there are numerous investigations in which the combination of these techniques has been used to take advantage of a possible synergy between them. Thus, for example, enzymes-assisted extraction often complemented with ultrasonic extraction or supercritical fluid extraction.

An overview of the improvement of all these methods can be observed in **Table 2**.


**Table 2.** Overview of the improvement of the main green extraction techniques.

In the majority of cases, the supercritical fluid used is CO<sup>2</sup>

compounds. This limitation can be solved by adding co-solvents that alter the polarity of the

SFE has been used for the extraction of lycopene β-carotene from tomato peels [35] and carrot peels [36]. For its part, the recovery of phenol by SFE has been tested in apple pomace [37],

The so-called pressurized solvent extraction is a technique that combines solvent extraction at temperatures (50–200°C) and high pressures (1500–2000 psi) to quickly and efficiently extract compounds from solid matrices. The use of liquid solvents at high temperatures and pressures improves the performance of the extractions since the solubility is increased and the mass transfer is improved and the rupture of the composite-matrix superficial equilibrium is facilitated [41]. PLE can be considered as a green extraction process especially when a non-

A wide range of organic nonpolar to polar solvent and their mixtures have been used in pressurized liquid extraction of phytochemical. However, it is a technology that modifies solvent properties. This fact makes water suitable for the extraction to polar and nonpolar organic

PLE could be carried out in static and dynamic modes. Static extraction is considered more efficient because of the greater penetration of the solvent into the pores of the source [10]. The advantages of this method with respect to conventional ones are the short extraction times, the yields obtained and the reduced usage of solvents. However, this method is not suitable for thermolabile compounds because the high temperature can alter the structure of these. For

Some researches have been carried out with pressurized liquid for the extraction of carot-

In addition to the above, other extraction technologies have been developed as the instant controlled pressure drop or pulsed electric field. The latter has been tested for the extraction of phenolic compounds from orange peel [44] or grape pomace [45] and alkaloids form potato peels [46]. Finally, there are numerous investigations in which the combination of these techniques has been used to take advantage of a possible synergy between them. Thus, for example, enzymes-assisted extraction often complemented with ultrasonic extraction or

that, the use of this technique for the extraction of phytochemical is very limited.

An overview of the improvement of all these methods can be observed in **Table 2**.

enoids from shrimp waste [42] or phenolic acid in potato peels [43].

, but they may become contained, requiring a subsequent separation operation.

cacao pod husk [38], mango by-products [39] or sour cherry pomace [40].

safe (GRAS). The application of supercritical CO<sup>2</sup>

nonpolar substances. However, the CO<sup>2</sup>

**3.5. Pressurized liquid extraction (PLE)**

toxic solvent is used.

**3.6. Other technologies**

supercritical fluid extraction.

compounds.

CO2

50 Descriptive Food Science

, for being generally recognized as

has high performances for the extraction of

under critical conditions is a poor solvent for polar

## **4. Industry phytochemical residue applications**

As stated in previous sections, phytochemicals are present in foods of plant origin. The food industry is considered a source of extraction of phytochemical compounds, which can be obtained from waste generated in the processing of fruit and vegetables. This fact is beneficial for food industry producers, as it generates an economic advantage [47].

It is worth mentioning that in its elaboration process, the less handling and processing of food generate a more retain of these compounds. For this reason, processes like a cereal refining, intense and long heating, cocking in broths which are then discarded and some of its compounds are lost, reduce the food phytochemical content. In nature, phytochemicals are in many foods, but in the future, the development of bioengineering will allow us to create plants with more concentration of these compounds.

In reference to its possible applications, the great phytochemicals properties allow us to open a wide range of possibilities. For example, due to its antioxidant activity, these could be used in creams, functional foods and fortified foods formulation, or due to its antimicrobial activity, these could be used in food conservation and medicine. In the following sections, we see some of the most important applications in detail.

#### **4.1. Phytochemicals in medicine and pharmacy**

The phytochemical application in medicine and pharmacy is directly related with the phytotherapy (use of plant-derived medications in the treatment and prevention of disease). The large quantity of properties that the plants has allowed us to apply them for treatments of diseases, and this is due to their phytochemical content. Numerous trials and pharmacological studies of specific phytotherapeutic preparations exist but, in some countries, phytotherapy is viewed as a form of traditional medicine.

For example, against respiratory diseases are effective the consumption of essentials oils of *Eucalyptus globulus Labill.* [48], *Origanum vulgare* L. [49], *Pinus* sp. [50] or *Thymus vulgaris* [51], which due to its high concentration of tannins, phenolic acids, terpenes and flavonoids, among others, cause an antiseptic and expectorant action.

cell damage, and this can generate to appear early signs of aging. If plant extract or directly phytochemicals compounds are adding to cosmetic products these can combat these toxic substances to cells and prevent the appearance of wrinkles, skin blemishes, and so on [60].

Currently, plant extracts rich in phytochemicals are also being evaluated in the oral cavity to prevent the development of microorganisms and prevent the appearance of dental caries and plaque. Specifically, the green tea extract has been evaluated against one of the main microorganisms that develop oral infections, Streptococcus mutants, obtaining great antimicrobial activity when applying the ethanolic extract of *C. sinensis* [61]. These results are of great importance because they show us that plant extracts rich in phytochemical compounds can be applied, for example, to dental prosthesis or toothpaste to prevent buccal infections.

Sara Martillanes thanks the sponsorship of Valhondo Calaff Foundation for the predoctoral fellowship. The publication of this chapter was supported by Government of Extremadura

and Jonathan Delgado-Adámez<sup>2</sup>

\*

Agrifood By-Products as a Source of Phytochemical Compounds

http://dx.doi.org/10.5772/intechopen.79434

53

**Acknowledgements**

and FEDER (GR100006).

, Javier Rocha-Pimienta<sup>2</sup>

\*Address all correspondence to: jonathan.delgado@juntaex.es

1 Agricultural Engineering School, University of Extremadura, Badajoz, Spain

2 Center for Scientific and Technological Research of Extremadura (CICYTEX-INTAEX),

[1] The Food and Agricultural Organization (FAO). Food Wastage Footprint: Impacts on Natural Resources. Summary Report. Rome, Italy: The Food and Agricultural Organization

[2] Goodwin TW. Distribution of carotenoids. Chemistry and Biochemistry of Plant Pigments.

[3] Singh G. Terpenoids. In: Chemistry of Terpenoids and Carotenoids. Vol. 3. Discovery

[4] Badui Dergal S. Química de los alimentos. 4th ed. Pearson Education; 2006. 736 p

**Author details**

Sara Martillanes<sup>1</sup>

Badajoz, Spain

**References**

(FAO); 2013

1976;**1**:225-261

Publishing House; 2007

Against circulatory diseases, the application of *Vitis vinifera* L. due to its high concentration of polyphenols, mainly resveratrol, in its fruits generate a lot of antioxidant activity and its anthocyanins do a veinotonic and vasoprotective action [52].

Other plant species which are of great importance is *Camellia sinensis* from which green tea is obtained. Some studies have reported the use of this specie rich in catechins against cancer chemoprevention, hypercholesterolemia, atherosclerosis, Parkinson's disease, Alzheimer's disease, and other aging-related disorders [53].

Concretely, an agroindustry residue, olive leaf, is a great source of phytochemicals. These contain a high concentration of oleuropein, tyrosol and hydroxytyrosol, three important phenolic compounds which have bioactive properties like antioxidant capacity [54]. The European Food Safety Association (EFSA) determined officially that oleuropein present in olive leaf extract generate a better glucose tolerance in humans [55]. These are some applications and uses in medicine and pharmacy of phytochemical compounds present in some plants but the number or these is increased day by day.

#### **4.2. Phytochemicals in food industry**

The food and agricultural products industries are an important source of phenolics-rich byproducts, which have been a good source of natural antioxidant. The application of these by-products in other foods like oil, fish and meat has shown antioxidant values similar to synthetic antioxidant, particularly the flavonoids and hydroxycinnamic acids [56]. The way to consume these by-products and add to our diet is directly, for example, the consumption of plants in tea form, or indirectly, for example, like an alimentary additive or supplement.

Another application of phytochemicals in food is the creation of active packaging for them. The films and coatings of different origin (proteins, polysaccharides, etc.) incorporate a wide variety of essential oils or plant extracts with the aim of creating active packaging with antimicrobial/antioxidant properties that improve the conservation of various types of food, for example, for fruit conservation [57], meat [58] and fish [59].

As we have already mentioned, the consumption of phytochemicals and adding these to our diet generate a lot of benefits to our health due to the large number of biological activities that possess.

#### **4.3. Other phytochemical application fields**

A great number of phytochemicals bioactivities open a wide range of possibilities and applications of these important food plants compounds. Other field is the cosmetic industry due to the antioxidant properties of phytochemical compounds. The reactive oxygen species generate cell damage, and this can generate to appear early signs of aging. If plant extract or directly phytochemicals compounds are adding to cosmetic products these can combat these toxic substances to cells and prevent the appearance of wrinkles, skin blemishes, and so on [60].

Currently, plant extracts rich in phytochemicals are also being evaluated in the oral cavity to prevent the development of microorganisms and prevent the appearance of dental caries and plaque. Specifically, the green tea extract has been evaluated against one of the main microorganisms that develop oral infections, Streptococcus mutants, obtaining great antimicrobial activity when applying the ethanolic extract of *C. sinensis* [61]. These results are of great importance because they show us that plant extracts rich in phytochemical compounds can be applied, for example, to dental prosthesis or toothpaste to prevent buccal infections.

## **Acknowledgements**

For example, against respiratory diseases are effective the consumption of essentials oils of *Eucalyptus globulus Labill.* [48], *Origanum vulgare* L. [49], *Pinus* sp. [50] or *Thymus vulgaris* [51], which due to its high concentration of tannins, phenolic acids, terpenes and flavonoids,

Against circulatory diseases, the application of *Vitis vinifera* L. due to its high concentration of polyphenols, mainly resveratrol, in its fruits generate a lot of antioxidant activity and its

Other plant species which are of great importance is *Camellia sinensis* from which green tea is obtained. Some studies have reported the use of this specie rich in catechins against cancer chemoprevention, hypercholesterolemia, atherosclerosis, Parkinson's disease, Alzheimer's

Concretely, an agroindustry residue, olive leaf, is a great source of phytochemicals. These contain a high concentration of oleuropein, tyrosol and hydroxytyrosol, three important phenolic compounds which have bioactive properties like antioxidant capacity [54]. The European Food Safety Association (EFSA) determined officially that oleuropein present in olive leaf extract generate a better glucose tolerance in humans [55]. These are some applications and uses in medicine and pharmacy of phytochemical compounds present in some plants but the

The food and agricultural products industries are an important source of phenolics-rich byproducts, which have been a good source of natural antioxidant. The application of these by-products in other foods like oil, fish and meat has shown antioxidant values similar to synthetic antioxidant, particularly the flavonoids and hydroxycinnamic acids [56]. The way to consume these by-products and add to our diet is directly, for example, the consumption of plants in tea form, or indirectly, for example, like an alimentary additive or supplement.

Another application of phytochemicals in food is the creation of active packaging for them. The films and coatings of different origin (proteins, polysaccharides, etc.) incorporate a wide variety of essential oils or plant extracts with the aim of creating active packaging with antimicrobial/antioxidant properties that improve the conservation of various types of food, for

As we have already mentioned, the consumption of phytochemicals and adding these to our diet generate a lot of benefits to our health due to the large number of biological activities that

A great number of phytochemicals bioactivities open a wide range of possibilities and applications of these important food plants compounds. Other field is the cosmetic industry due to the antioxidant properties of phytochemical compounds. The reactive oxygen species generate

among others, cause an antiseptic and expectorant action.

anthocyanins do a veinotonic and vasoprotective action [52].

example, for fruit conservation [57], meat [58] and fish [59].

**4.3. Other phytochemical application fields**

disease, and other aging-related disorders [53].

52 Descriptive Food Science

number or these is increased day by day.

**4.2. Phytochemicals in food industry**

possess.

Sara Martillanes thanks the sponsorship of Valhondo Calaff Foundation for the predoctoral fellowship. The publication of this chapter was supported by Government of Extremadura and FEDER (GR100006).

## **Author details**

Sara Martillanes<sup>1</sup> , Javier Rocha-Pimienta<sup>2</sup> and Jonathan Delgado-Adámez<sup>2</sup> \*

\*Address all correspondence to: jonathan.delgado@juntaex.es

1 Agricultural Engineering School, University of Extremadura, Badajoz, Spain

2 Center for Scientific and Technological Research of Extremadura (CICYTEX-INTAEX), Badajoz, Spain

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**Chapter 4**

**Provisional chapter**

**Chemical Constituents of Fruit Wines as Descriptors of**

Functional foods are foods that provide positive health effects apart from the provision of essential nutrients. Along with nutraceuticals, they represent the top trends in the food industry. Fruit wines are considered functional foods. When assessing the fruit wine quality, a wide range of descriptors are taken into consideration, namely physicochemical and sensorial properties of fruit wine. Furthermore, within the context of the new food products development (e.g. functional products), functional properties of fruit wines are also taken into consideration. Functional properties are determined by the content of the biologically active components, such as polyphenols, vitamins and micro- and macrominerals. It is also important to consider the food-safety issues regarding the fruit wines consummation, that is, the presence of pesticides, mycotoxins and biogenic amines in different fruit wines. This chapter aims to give an overview of various factors used to

evaluate the quality and the functional properties of fruit wines.

compounds, mineral composition, volatile compounds, food safety

**Keywords:** fruit wine, functional food, biologically active compounds, phenolic

The past decade has seen the rapid development of fruit wine production in many countries, which may be attributed to the enhanced consumers' interest in functional foods that may help to promote health and help reduce the risk of disease. However, there is literature gap data

**Chemical Constituents of Fruit Wines as Descriptors** 

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.78796

**their Nutritional, Sensorial and Health-Related**

**of their Nutritional, Sensorial and Health-Related** 

Darko Velić, Daniela Amidžić Klarić, Natalija Velić, Ilija Klarić, Vlatka Petravić Tominac and Ana Mornar

Darko Velić, Daniela Amidžić Klarić, Natalija Velić, Ilija Klarić, Vlatka Petravić Tominac and Ana Mornar

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.78796

**Properties**

**Abstract**

**1. Introduction**

**Properties**


#### **Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial and Health-Related Properties Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial and Health-Related Properties**

DOI: 10.5772/intechopen.78796

Darko Velić, Daniela Amidžić Klarić, Natalija Velić, Ilija Klarić, Vlatka Petravić Tominac and Ana Mornar Darko Velić, Daniela Amidžić Klarić, Natalija Velić, Ilija Klarić, Vlatka Petravić Tominac and Ana Mornar

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.78796

#### **Abstract**

[52] Bhat KP, Kosmeder JW, Pezzuto JM. Biological effects of resveratrol. Antioxidants and

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58 Descriptive Food Science

1924/2006. EFSA Journal. 2014;**12**(5):3655

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987-1000. DOI: 10.1002/ptr.1363

Research. 2016;**30**(7):1172-1179. DOI: 10.1002/ptr.5625

Functional foods are foods that provide positive health effects apart from the provision of essential nutrients. Along with nutraceuticals, they represent the top trends in the food industry. Fruit wines are considered functional foods. When assessing the fruit wine quality, a wide range of descriptors are taken into consideration, namely physicochemical and sensorial properties of fruit wine. Furthermore, within the context of the new food products development (e.g. functional products), functional properties of fruit wines are also taken into consideration. Functional properties are determined by the content of the biologically active components, such as polyphenols, vitamins and micro- and macrominerals. It is also important to consider the food-safety issues regarding the fruit wines consummation, that is, the presence of pesticides, mycotoxins and biogenic amines in different fruit wines. This chapter aims to give an overview of various factors used to evaluate the quality and the functional properties of fruit wines.

**Keywords:** fruit wine, functional food, biologically active compounds, phenolic compounds, mineral composition, volatile compounds, food safety

#### **1. Introduction**

The past decade has seen the rapid development of fruit wine production in many countries, which may be attributed to the enhanced consumers' interest in functional foods that may help to promote health and help reduce the risk of disease. However, there is literature gap data

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

regarding the production, properties and health benefits of different fruit wines, compared to grape wines. With the ageing of the population, the incidence of chronic diseases that can be linked to caloric excess and the cost of health-care both increase, and so is the consumers' preference towards self-care, instead of pharmaceutical management of disease [1]. The utilisation of functional foods for health-care can lead to the reduction of the use of pharmaceuticals or their replacement. According to the Functional Food and Nutraceuticals market report [2], many consumers were prompted to turn to preventive or alternative health-care practices due to the increase of health-care costs [3]. Consequently, the consummation of functional foods and nutraceuticals increased substantially. Moderate consumption of grape wine (mainly red) has a beneficial health effects when combined with balanced diet, which is confirmed by many studies [4–9]. Some of the reported effects regarding health effects include the protection of the incidence of cardiovascular diseases, ischaemic stroke, hypertension, diabetes, dietary cancers, peptic ulcers, kidney stones and macular degeneration, as well as stimulation of resistance to infection and bone density retention [10–14]. However, apart from grapes, a whole range of other fruits that differ in shape, taste, colour and nutritive value can also be utilised for the production of wine [9], which are nowadays gaining the full acceptance at the market [15]. Fruit wines have proved to be an excellent dietary source of antioxidants, phytonutrients and minerals [16]. According to the European regulations, fruit wines must be obtained by the fermentation of fruits (or respective juices) other than grape. Based on the carbon dioxide retention and content, fruit wines can be classified either as still or sparkling, while their permitted alcoholic strength lies between 1.2 and 14% by volume [14]. Winemaking technology of fruit wines is mostly similar to that of grape wines, except for the variations based on the fruit used, that usually involves the adjustment of fruit juice for winemaking, for example, sugar content or acidity. Geographical area, that is, the fruit cultivars typically grown in that specific area, mainly determines the production and consumption of different fruit wines. Non-grape fruits successfully used for the production of fruit wines in different parts of the world include blackberries, strawberries, currants, apples, wild apricots, pears, kiwifruit, plums, peaches, cherries, bananas, pineapples, cashew nuts, pomegranates, lemons, tangerines, oranges, dates, figs, and so on [17, 18]. There is a large volume of published studies describing chemical composition, with the particular emphasis on various bioactive compounds, of different fruits and fruit juices [19–32]. However, comprehensive reviews of respective fruit wines are still scarce, as well as the comparison of different fruit wines regarding their bioactive compounds. The primary focus of this chapter includes three most popular fruit wines in Croatia, produced from three different groups of fruit, namely berries (blackberry wine), stone fruit (sour cherry wine) and pome fruit (apple wine), in respect to chemical constituents affecting their nutritional, sensorial and health-related properties. Furthermore, food-safety issues, that is, the presence of pesticides, mycotoxins and biogenic amines, are also tackled.

Fermentation is an anaerobic process carried out by yeast in which yeast converts the relatively high level of sugars present in fruits into ethanol and carbon dioxide [14]. Ethanol production is controlled by three main factors: sugar content, fermentation temperature and yeast strain. Particular types of yeast are used in wine production that can both tolerate and produce high alcohol levels (in some cases up to 14%, or even higher). The alcohol content is usually expressed as a volume percentage. Under standard fermentation conditions, ethanol can accumulate to up to 14–15%, while higher ethanol levels can be achieved by the sequential addition of sugar during fermentation. However, to achieve ethanol concentrations above 15%, the wine has to be fortified [33]. The role of ethanol in wine is multiple: it acts as an essential cosolvent (along with water) in extracting fruits constituents, it serves as a reactant in the generation of important volatile compounds (e.g. ethyl esters) and is crucial to the sensory properties, stability and ageing of wine [33]. The secondary products of yeast metabolism include higher alcohols (containing more than two carbons), also known as fusel alcohols or fusel oils [34]. Some of the representatives of higher alcohols are *n*-propanol, isobutyl alcohol (2-methyl-1-propanol), 2-methyl butanol (optically active amyl alcohol), isoamyl alcohol and 2-phenyl ethanol. Glycerol is the primary fermentation product of yeast, besides ethanol and carbon dioxide, and can indirectly contribute to the sensory character of the wine [35]. It is a colourless, odourless, non-volatile compound, without aromatic properties. However, it contributes significantly to the sweetness, as well as to the full and round mouth-feel of wine and wine texture [36]. There is a difference in the levels of glycerol formed by various yeast strains, and therefore glycerol production should be considered in the selection of wine yeast strains [37, 38]. Glycerol is mainly produced during glyceropyruvic fermentation at the start of alcoholic fermentation, and its degradation can be detrimental to the wine quality—partly because of the decrease of its concentration and partly because of the resulting metabolic products [39]. Glycerol concentrations of 5.976 and 4.491 were determined in two blackberry wines, produced on a small scale (in microfermentation experiments) by two commercial wine yeasts [40]. The reported glycerol concentrations in cider are 3–6 g/L [41]. Furthermore, glycerol can be metabolised by lactic acid bacteria during cider maturation, decreasing its final sensorial quality. Selection of yeast strains for secondary fermentation of sparkling ciders has been made by Suárez Valles et al. [42]. Glycerol level was 4.7 g/L in base cider while sparkling ciders produced using four different yeast strains for secondary fermentation contained 5.1–5.2 g/L. Besides ethanol and glycerol, methanol can also be present in measurable concentrations in some types of fruit wines (e.g. plum wine). However, being toxic to humans, methanol content of commercially available fruit wines should not exceed 200–250 mg/L, as regulated in different countries [14]. Fructose, glucose and, in some fruits, sucrose are the major sugars present in fruits. They are fermented by yeast during fruit wine fermentation, producing previously mentioned major metabolites: ethanol, carbon dioxide and glycerol. However, fruits other than grapes are often much lower in sugar content compared to grapes, which makes them unsuitable for winemaking unless sugar content is adjusted by the addition of (most often) sucrose [14]. Sugars may also be metabolised to higher alcohols, fatty acid esters, and aldehydes, which mostly define the primary aromatic character of wine [33]. The sugars that remain unfermented (i.e. the sugars in wine) are referred to as residual sugars. Amidžić Klarić et al. [43] reported the wide range of residual (reducing) sugars from 13.5 to 177.6 g/L in investigated blackberry wines. The authors noted that sugar (mostly sucrose) is often added during different stages of the blackberry wine production, so the wide range

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial…

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61

#### **2. Chemical constituents of fruit wines**

#### **2.1. Alcohol, sugars and organic acids**

Fruit wine consists of two primary ingredients—water and ethanol, the latter being the primary product of alcoholic fermentation, essential for both flavour and stability of wine. Fermentation is an anaerobic process carried out by yeast in which yeast converts the relatively high level of sugars present in fruits into ethanol and carbon dioxide [14]. Ethanol production is controlled by three main factors: sugar content, fermentation temperature and yeast strain. Particular types of yeast are used in wine production that can both tolerate and produce high alcohol levels (in some cases up to 14%, or even higher). The alcohol content is usually expressed as a volume percentage. Under standard fermentation conditions, ethanol can accumulate to up to 14–15%, while higher ethanol levels can be achieved by the sequential addition of sugar during fermentation. However, to achieve ethanol concentrations above 15%, the wine has to be fortified [33]. The role of ethanol in wine is multiple: it acts as an essential cosolvent (along with water) in extracting fruits constituents, it serves as a reactant in the generation of important volatile compounds (e.g. ethyl esters) and is crucial to the sensory properties, stability and ageing of wine [33]. The secondary products of yeast metabolism include higher alcohols (containing more than two carbons), also known as fusel alcohols or fusel oils [34]. Some of the representatives of higher alcohols are *n*-propanol, isobutyl alcohol (2-methyl-1-propanol), 2-methyl butanol (optically active amyl alcohol), isoamyl alcohol and 2-phenyl ethanol. Glycerol is the primary fermentation product of yeast, besides ethanol and carbon dioxide, and can indirectly contribute to the sensory character of the wine [35]. It is a colourless, odourless, non-volatile compound, without aromatic properties. However, it contributes significantly to the sweetness, as well as to the full and round mouth-feel of wine and wine texture [36]. There is a difference in the levels of glycerol formed by various yeast strains, and therefore glycerol production should be considered in the selection of wine yeast strains [37, 38]. Glycerol is mainly produced during glyceropyruvic fermentation at the start of alcoholic fermentation, and its degradation can be detrimental to the wine quality—partly because of the decrease of its concentration and partly because of the resulting metabolic products [39]. Glycerol concentrations of 5.976 and 4.491 were determined in two blackberry wines, produced on a small scale (in microfermentation experiments) by two commercial wine yeasts [40]. The reported glycerol concentrations in cider are 3–6 g/L [41]. Furthermore, glycerol can be metabolised by lactic acid bacteria during cider maturation, decreasing its final sensorial quality. Selection of yeast strains for secondary fermentation of sparkling ciders has been made by Suárez Valles et al. [42]. Glycerol level was 4.7 g/L in base cider while sparkling ciders produced using four different yeast strains for secondary fermentation contained 5.1–5.2 g/L. Besides ethanol and glycerol, methanol can also be present in measurable concentrations in some types of fruit wines (e.g. plum wine). However, being toxic to humans, methanol content of commercially available fruit wines should not exceed 200–250 mg/L, as regulated in different countries [14]. Fructose, glucose and, in some fruits, sucrose are the major sugars present in fruits. They are fermented by yeast during fruit wine fermentation, producing previously mentioned major metabolites: ethanol, carbon dioxide and glycerol. However, fruits other than grapes are often much lower in sugar content compared to grapes, which makes them unsuitable for winemaking unless sugar content is adjusted by the addition of (most often) sucrose [14]. Sugars may also be metabolised to higher alcohols, fatty acid esters, and aldehydes, which mostly define the primary aromatic character of wine [33]. The sugars that remain unfermented (i.e. the sugars in wine) are referred to as residual sugars. Amidžić Klarić et al. [43] reported the wide range of residual (reducing) sugars from 13.5 to 177.6 g/L in investigated blackberry wines. The authors noted that sugar (mostly sucrose) is often added during different stages of the blackberry wine production, so the wide range

regarding the production, properties and health benefits of different fruit wines, compared to grape wines. With the ageing of the population, the incidence of chronic diseases that can be linked to caloric excess and the cost of health-care both increase, and so is the consumers' preference towards self-care, instead of pharmaceutical management of disease [1]. The utilisation of functional foods for health-care can lead to the reduction of the use of pharmaceuticals or their replacement. According to the Functional Food and Nutraceuticals market report [2], many consumers were prompted to turn to preventive or alternative health-care practices due to the increase of health-care costs [3]. Consequently, the consummation of functional foods and nutraceuticals increased substantially. Moderate consumption of grape wine (mainly red) has a beneficial health effects when combined with balanced diet, which is confirmed by many studies [4–9]. Some of the reported effects regarding health effects include the protection of the incidence of cardiovascular diseases, ischaemic stroke, hypertension, diabetes, dietary cancers, peptic ulcers, kidney stones and macular degeneration, as well as stimulation of resistance to infection and bone density retention [10–14]. However, apart from grapes, a whole range of other fruits that differ in shape, taste, colour and nutritive value can also be utilised for the production of wine [9], which are nowadays gaining the full acceptance at the market [15]. Fruit wines have proved to be an excellent dietary source of antioxidants, phytonutrients and minerals [16]. According to the European regulations, fruit wines must be obtained by the fermentation of fruits (or respective juices) other than grape. Based on the carbon dioxide retention and content, fruit wines can be classified either as still or sparkling, while their permitted alcoholic strength lies between 1.2 and 14% by volume [14]. Winemaking technology of fruit wines is mostly similar to that of grape wines, except for the variations based on the fruit used, that usually involves the adjustment of fruit juice for winemaking, for example, sugar content or acidity. Geographical area, that is, the fruit cultivars typically grown in that specific area, mainly determines the production and consumption of different fruit wines. Non-grape fruits successfully used for the production of fruit wines in different parts of the world include blackberries, strawberries, currants, apples, wild apricots, pears, kiwifruit, plums, peaches, cherries, bananas, pineapples, cashew nuts, pomegranates, lemons, tangerines, oranges, dates, figs, and so on [17, 18]. There is a large volume of published studies describing chemical composition, with the particular emphasis on various bioactive compounds, of different fruits and fruit juices [19–32]. However, comprehensive reviews of respective fruit wines are still scarce, as well as the comparison of different fruit wines regarding their bioactive compounds. The primary focus of this chapter includes three most popular fruit wines in Croatia, produced from three different groups of fruit, namely berries (blackberry wine), stone fruit (sour cherry wine) and pome fruit (apple wine), in respect to chemical constituents affecting their nutritional, sensorial and health-related properties. Furthermore, food-safety issues, that is, the presence

of pesticides, mycotoxins and biogenic amines, are also tackled.

Fruit wine consists of two primary ingredients—water and ethanol, the latter being the primary product of alcoholic fermentation, essential for both flavour and stability of wine.

**2. Chemical constituents of fruit wines**

**2.1. Alcohol, sugars and organic acids**

60 Descriptive Food Science

of reducing sugars concentration found in investigated samples is probably the result of the applied technological procedure. Sun et al. [44] studied the suitability of different yeast strains for the production of cherry wine and reported the concentration of total reducing sugars in wine ranging from 0.7 to 1 g /L. Initial analysis of must revealed the initial concentration of total sugars of 165 g/L, achieved by the addition of sucrose to the crushed and deseeded cherries. The discrepancy in the residual sugar content between the blackberry wines and cherry wines can be attributed to the production technology and consumers' preferences—low acidity and high sugar fruit wines seem to be the most acceptable [14]. Fruit species, climate and geomorphological character of soil determine the content of organic acids in fruit. Consequently, the acidity of fruits affects the fruit wine acidity. Total acidity is the wine-quality parameter that gives the measure of the wine acidity, a parameter influencing the wine taste and the overall quality [43]. The total acidity of blackberry wines reported by Amidžić Klarić et al. [43] ranged from 6.7 to 18.1 g/L (as tartaric acid), while the reported total acidities of cherry wines ranged from 5.94 to 6.71 g/L [44]. When it comes to grape wine tasting, a high level of acidity refers to excessively tart, sour and sharp wine attributes, while a low total acidity results in a flat-tasting wine that is more susceptible to infection and spoilage by microorganisms [45]. Organoleptic properties of musts and fruit wines are strongly influenced by organic acids [46]. The major organic acid present in blackberry, cherry and apple wines is malic acid, with reported concentrations of 3.5, 6.8 and 6.2 g/L, respectively. The second most abundant is citric acid [14], while the predominant volatile acid is acetic acid, often expressed as the wine-quality parameter called volatile acidity. Acetic acid is a secondary metabolite derived from a pyruvic acid, which is always formed during alcoholic fermentation. Its accumulation in wine is usually the result of the secondary infection of the fruit, that is, the acetic acid bacteria activity, which is promoted by excessive oxygen uptake. The secondary infection can occur during the vinification process or after bottling [47]. Higher concentrations of acetic acid can detrimentally affect the organoleptic properties of wine, because of bitter taste and smell-like vinegar [39]. Besides acetic acid, as already mentioned, malic acid also contributes to the acidity of fruit wine. However, its degradation by malolactic fermentation (MLF) can reduce the acidity of the wine. MLF can occur spontaneously during or at the end of alcoholic fermentation or can be induced by the addition of lactic acid bacteria starter cultures, namely *Oenococcus oeni* as the primary species used in MLF. Compared to malic acid, lactic acid has a softer flavour, which results in the more desirable flavour profile of wine [48]. Furthermore, the conversion of malic acid to lactic acid results in reduced wine acidity and improved stability and quality of high-acid wines [49].

volatiles, there are also many minor volatile and non-volatile compounds adding to the aroma of fruit wines, such as aldehydes, ketones, lactones, terpenes and phenols [51]. A fast, selective and sensitive method for the determination of volatile compounds in blackberry wine samples using an HSS-GC-FID method was presented by Mornar et al. [52]; still, the study encompassed a limited number of compounds as well as samples. Therefore, Amidžić Klarić et al. [53] analysed volatile compounds in the 15 blackberry wine samples produced from conventionally and organically grown blackberries using a GC-FID method. The amount of ethyl acetate in the investigated samples ranged from 53.8 to 188.4 mg/L, while propane-1-ol was found in three organic samples. Although isoamyl alcohol was found in all samples, the measured values ranged from 56.7 to 226.9 mg/L. A GC–MS technique was used by Wang et al. [54] for the investigation of the fermentation process influence (primary and secondary) on volatile compounds of blackberry wine. Fifty-five volatile compounds were detected in blackberry fruit juice before fermentation, while nine new aroma components such as octanoate, benzenepropanoic acid ethyl ester, ethyl benzoate, dodecyl ethyl, *n*-propanol, *n*-butanol, D-citronellol, benzaldehyde and cedrol were detected in natural ageing wine which appeared during secondary fermentation. These findings emphasise the importance of natural ageing for the formation of aroma components of blackberry wine. Due to the high popularity of cherry wine, the volatile compounds of this fruit wine were in focus of several investigations as indicators of aroma profile of fruit wines. Niu et al. [55] have applied descriptive sensory analysis to describe the aroma attributes of different cherry wines by GC-olfactometry, an analytical technique that uses human assessors as a sensitive and selective detector for odour-active compounds. Fifty-one compounds were detected and subsequently quantified by GC–MS. The conducted research revealed that the aroma-active compound profiles were dominated by ethyl 2-methyl propionate, 2,3-butanedione, ethyl butyrate, ethyl pentanoate, 3-methyl-1-butanol, ethyl hexanoate, 3-hydroxy-2-butanone, ethyl lactate, 1-hexanol, (Z)-3 hexen-1-ol, ethyl hydroxyacetate, acetic acid, furfural, 2-ethyl-1-hexanol, benzaldehyde, propanoic acid, butanoic acid, guaiacol, beta-citronellol, hexanoic acid, 2-methoxy-4-methylphenol, 2-ethyl-3-hydroxy-4H-pyran-4-one, ethyl cinnamate and 2-methoxy-4-vinylphenol. Afterwards, Xiao et al. [56] have determined 75 volatiles in 9 cherry wine samples using an improved sample preparation technique HS-SPME. The cluster analysis results suggested that esterification reactions and fermentation process were more extended during the ageing period and the production process could have a significant influence on volatile profile of cherry wines. The similar conclusion was obtained by Xiao et al. [57] using multivariate classification of cherry wines. The research revealed the major volatile components of investigated cherry wines: isoamylol, ethyl acetate, benzyl alcohol, benzaldehyde and diethyl succinate. More recently, the research of Xiao et al. [58] have shown that the Chinese cherry wines from different price segments have the different aroma-active compounds profile as well as various aroma attributes. Furthermore, the correlation between the composition of aroma-active compounds and cherry wine from different regions was established by Xiao et al. [59]. Wang et al. [60] have proposed a new, rapid method for analysis of volatile compounds in apple wine using HS-SPME-GC-MS technique. Forty-three volatile compounds (alcohols, esters, lower fatty acids, carbonyls, alkenes, terpenes and phenols) were quantified in `Fuji´ apple wine. The most abundant aroma compounds were esters, alcohols and lower fatty acids; total concentrations were 242.1, 479.3 and 297.4 mg/mL, respectively. Still, the

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#### **2.2. Volatile (aroma) compounds**

Knowledge of the volatile composition of wine is of great interest; since these compounds are responsible for the quality of wine aroma. The aroma of fruit wines is mainly determined by volatile compounds produced by the fruit itself (varietal wine aroma), as by-products of alcoholic and malolactic fermentation (fermentative wine aroma), and formed during bottling, ageing and storing (post-fermentative wine aroma) [50]. Esters, higher alcohols, acetates, organic acids and other compounds are the groups of volatile compounds that most commonly contribute to the flavour and/or aroma profile of fruit. Apart from the listed groups of volatiles, there are also many minor volatile and non-volatile compounds adding to the aroma of fruit wines, such as aldehydes, ketones, lactones, terpenes and phenols [51]. A fast, selective and sensitive method for the determination of volatile compounds in blackberry wine samples using an HSS-GC-FID method was presented by Mornar et al. [52]; still, the study encompassed a limited number of compounds as well as samples. Therefore, Amidžić Klarić et al. [53] analysed volatile compounds in the 15 blackberry wine samples produced from conventionally and organically grown blackberries using a GC-FID method. The amount of ethyl acetate in the investigated samples ranged from 53.8 to 188.4 mg/L, while propane-1-ol was found in three organic samples. Although isoamyl alcohol was found in all samples, the measured values ranged from 56.7 to 226.9 mg/L. A GC–MS technique was used by Wang et al. [54] for the investigation of the fermentation process influence (primary and secondary) on volatile compounds of blackberry wine. Fifty-five volatile compounds were detected in blackberry fruit juice before fermentation, while nine new aroma components such as octanoate, benzenepropanoic acid ethyl ester, ethyl benzoate, dodecyl ethyl, *n*-propanol, *n*-butanol, D-citronellol, benzaldehyde and cedrol were detected in natural ageing wine which appeared during secondary fermentation. These findings emphasise the importance of natural ageing for the formation of aroma components of blackberry wine. Due to the high popularity of cherry wine, the volatile compounds of this fruit wine were in focus of several investigations as indicators of aroma profile of fruit wines. Niu et al. [55] have applied descriptive sensory analysis to describe the aroma attributes of different cherry wines by GC-olfactometry, an analytical technique that uses human assessors as a sensitive and selective detector for odour-active compounds. Fifty-one compounds were detected and subsequently quantified by GC–MS. The conducted research revealed that the aroma-active compound profiles were dominated by ethyl 2-methyl propionate, 2,3-butanedione, ethyl butyrate, ethyl pentanoate, 3-methyl-1-butanol, ethyl hexanoate, 3-hydroxy-2-butanone, ethyl lactate, 1-hexanol, (Z)-3 hexen-1-ol, ethyl hydroxyacetate, acetic acid, furfural, 2-ethyl-1-hexanol, benzaldehyde, propanoic acid, butanoic acid, guaiacol, beta-citronellol, hexanoic acid, 2-methoxy-4-methylphenol, 2-ethyl-3-hydroxy-4H-pyran-4-one, ethyl cinnamate and 2-methoxy-4-vinylphenol. Afterwards, Xiao et al. [56] have determined 75 volatiles in 9 cherry wine samples using an improved sample preparation technique HS-SPME. The cluster analysis results suggested that esterification reactions and fermentation process were more extended during the ageing period and the production process could have a significant influence on volatile profile of cherry wines. The similar conclusion was obtained by Xiao et al. [57] using multivariate classification of cherry wines. The research revealed the major volatile components of investigated cherry wines: isoamylol, ethyl acetate, benzyl alcohol, benzaldehyde and diethyl succinate. More recently, the research of Xiao et al. [58] have shown that the Chinese cherry wines from different price segments have the different aroma-active compounds profile as well as various aroma attributes. Furthermore, the correlation between the composition of aroma-active compounds and cherry wine from different regions was established by Xiao et al. [59]. Wang et al. [60] have proposed a new, rapid method for analysis of volatile compounds in apple wine using HS-SPME-GC-MS technique. Forty-three volatile compounds (alcohols, esters, lower fatty acids, carbonyls, alkenes, terpenes and phenols) were quantified in `Fuji´ apple wine. The most abundant aroma compounds were esters, alcohols and lower fatty acids; total concentrations were 242.1, 479.3 and 297.4 mg/mL, respectively. Still, the

of reducing sugars concentration found in investigated samples is probably the result of the applied technological procedure. Sun et al. [44] studied the suitability of different yeast strains for the production of cherry wine and reported the concentration of total reducing sugars in wine ranging from 0.7 to 1 g /L. Initial analysis of must revealed the initial concentration of total sugars of 165 g/L, achieved by the addition of sucrose to the crushed and deseeded cherries. The discrepancy in the residual sugar content between the blackberry wines and cherry wines can be attributed to the production technology and consumers' preferences—low acidity and high sugar fruit wines seem to be the most acceptable [14]. Fruit species, climate and geomorphological character of soil determine the content of organic acids in fruit. Consequently, the acidity of fruits affects the fruit wine acidity. Total acidity is the wine-quality parameter that gives the measure of the wine acidity, a parameter influencing the wine taste and the overall quality [43]. The total acidity of blackberry wines reported by Amidžić Klarić et al. [43] ranged from 6.7 to 18.1 g/L (as tartaric acid), while the reported total acidities of cherry wines ranged from 5.94 to 6.71 g/L [44]. When it comes to grape wine tasting, a high level of acidity refers to excessively tart, sour and sharp wine attributes, while a low total acidity results in a flat-tasting wine that is more susceptible to infection and spoilage by microorganisms [45]. Organoleptic properties of musts and fruit wines are strongly influenced by organic acids [46]. The major organic acid present in blackberry, cherry and apple wines is malic acid, with reported concentrations of 3.5, 6.8 and 6.2 g/L, respectively. The second most abundant is citric acid [14], while the predominant volatile acid is acetic acid, often expressed as the wine-quality parameter called volatile acidity. Acetic acid is a secondary metabolite derived from a pyruvic acid, which is always formed during alcoholic fermentation. Its accumulation in wine is usually the result of the secondary infection of the fruit, that is, the acetic acid bacteria activity, which is promoted by excessive oxygen uptake. The secondary infection can occur during the vinification process or after bottling [47]. Higher concentrations of acetic acid can detrimentally affect the organoleptic properties of wine, because of bitter taste and smell-like vinegar [39]. Besides acetic acid, as already mentioned, malic acid also contributes to the acidity of fruit wine. However, its degradation by malolactic fermentation (MLF) can reduce the acidity of the wine. MLF can occur spontaneously during or at the end of alcoholic fermentation or can be induced by the addition of lactic acid bacteria starter cultures, namely *Oenococcus oeni* as the primary species used in MLF. Compared to malic acid, lactic acid has a softer flavour, which results in the more desirable flavour profile of wine [48]. Furthermore, the conversion of malic acid to lactic acid results in reduced wine

acidity and improved stability and quality of high-acid wines [49].

Knowledge of the volatile composition of wine is of great interest; since these compounds are responsible for the quality of wine aroma. The aroma of fruit wines is mainly determined by volatile compounds produced by the fruit itself (varietal wine aroma), as by-products of alcoholic and malolactic fermentation (fermentative wine aroma), and formed during bottling, ageing and storing (post-fermentative wine aroma) [50]. Esters, higher alcohols, acetates, organic acids and other compounds are the groups of volatile compounds that most commonly contribute to the flavour and/or aroma profile of fruit. Apart from the listed groups of

**2.2. Volatile (aroma) compounds**

62 Descriptive Food Science

dominant aroma component was isoamyl alcohol (232.0 mg/mL). The same technique was used for the investigation of the temperature effect during apple winemaking on both the critical aroma compounds and sensory properties of wine. The concentration of all aroma compounds was changed with a temperature increase, and sensory analysis showed the highest acceptance of apple wine fermented at 20°C [61]. The influence of apple variety (Šampion, Idared and Gloster) harvested from the orchard in Poland on the volatile composition, and sensory characteristics of apple wine were investigated by Satora et al. [62]. The high concentration of acetaldehyde, ethyl acetate and methanol was found in Šampion wines, while Gloster wines contained a higher concentration of fusel alcohols. The Idared wines had the best results of the sensory evaluation and high levels of butanol and acetic acid. As mentioned earlier, the majority of investigations were performed using sample preparation procedures such as HS and/or SPME, while gas chromatography with FID and MS detectors was shown to be the analytical technique of choice for the determination of volatile compounds in fruit wine. Ye et al. [63] have developed and applied a new rapid method for the identification of volatile compounds in apple wine using FT-NIR spectroscopy. To provide the greater insight into yeast metabolism and flavour formation of mulberry wine, Butkhup et al. [64] have developed a new HS-SPME-GC-MS method to characterise various volatile compounds of in-house-made mulberry wine. Eighty volatile compounds belonging to groups of higher alcohols, fatty acids, esters and phenols were quantified. These compounds were present in various amounts from 0.1 mg/L (benzene carboxaldehyde) to 138.4 mg/mL (isoamyl alcohol). Feng et al. [65] have extended their research on other fruit wines produced from mulberry as well as raspberry and strawberry. Alcohols formed the most abundant group, followed by esters and acids. Comparing to investigated grape wine, two alcohols, 4-methyl-2-pentanol and 2,3-butanediol, were not found in the three fruit wines. While the number of esters in raspberry (1.5%) and mulberry (2.1%) wines were higher than those of strawberry (0.8%) wine, there were no significant differences in acid content. Song et al. [66] have used the same sample preparation and analytical techniques to determine 78 volatile compounds in wild strawberry wine sample from a southern region of China. Odour activity values were detected for 21 compounds while 6 of them were identified as the particular aroma substances for wild strawberry wine, in particular, methyl 2-methylbutyrate, ethyl 2-methylbutyrate, methyl 3-methylbutyrate, (E)-3-hexen-1-ol, 1-octen-3-ol and phenylacetaldehyde.

**2.3. Phenolic compounds**

Wine quality is significantly influenced by phenolic compounds, namely anthocyanins, flavonols, catechins and other flavonoids since they have an enormous impact on the sensory characteristics of wines, mainly colour and astringency [69]. Furthermore, they exhibit a range of antioxidant and pharmacological effects. Phenolic compounds can be classified in different ways because they are constituted at a large number of heterogeneous structures that range from simple molecules to highly polymerised compounds. A review of the composition and content of phenolic compounds in various fruit wines is presented in **Table 1**. Based on the total phenolic content, wines are commonly categorised to three major groups: (i) high in total phenolics, (ii) moderately high in total phenolics and (iii) low in total phenolics. The total phenolic content of cherry, raspberry, black currant, bilberry, elderberry and sea buckthorn fruit wines proved to be comparable or even higher than that of grape red wines, while apple, plum and peach fruit wines had a lower total phenolic content than red grape wines [20, 87–89]. Among the phenolic compounds with known antioxidant activity,

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*Flavonoids*, the most studied phenolics, are important components of the human diet. They are also the most widely present in plants and are among the most potent antioxidants in plants. Different members of certain classes of flavonoids can be either colourless or coloured (e.g. anthocyanins), such as pigments of flowers and other plant parts [26]. Important subclasses of flavonoids are, among others, anthocyanins and flavanols. Anthocyanins are important polyphenolic components of fruits, especially berries [90]. The anthocyanins are conjugated anthocyanidins, and they provide the unique colours of dark berries. There are six anthocyanidins distributed throughout the plant kingdom: cyanidin, malvidin, delphinidin, peonidin, pelargonidin and petunidin. The significant anthocyanins in blackberries are cyanidine derivatives. In nature, anthocyanins are mainly found as heterosides. The aglycone form of anthocyanin, known as anthocyanidin, is structurally based on the flavilium or 2-phenylbenzopyrilium cation, with methoxyl and hydroxyl groups present at different positions of the basic structure. Numerous studies have shown that anthocyanins are absorbed in their original glycosylated forms in humans [14, 91, 92]. Anthocyanins are predominant phenolic components or comprise a considerable portion of the total polyphenol content of blackberry as well as cherry juice [76, 84, 93–95]. They are found in berry wines and some fruit wines and presented as total anthocyanin content (TAC). The TAC values found in some blackberry and blueberry wines were 75.56 and 20.82 mg/L (expressed as ellagic acid equivalents), respectively [79]. Mudnić et al. [96] determined high TAC values in blackberry wines, comparable to that measured in red grape wine and ranging from 134 to 164 mg/L (expressed as malvidin 3-glucoside equivalents). Flavonols, another important class of flavonoids, are characteristic for the cool climate fruits and their respective wines [14]. Myricetin and quercetin have been recognised as the first flavonols detected in red berry wines [5]. The results of

a study conducted by Mudnić et al. [73] that compared flavanol and procyanidin B<sup>2</sup>

were determined in two blackberry wines. The content of procyanidin B<sup>2</sup>

of several grape wines (red and white), with four commercially available blackberry wines, revealed that the highest concentrations of catechin (45.2 mg/L) and epicatechin (34.7 mg/L)

contents

(77.1 mg/L) of the

flavonoids, phenolic acids and tannins are highlighted [26].

#### *2.2.1. Volatile compounds containing sulphur*

The formation of off-flavours represents the core problem of high-quality fruit wine production. The main compounds responsible for off-flavours of fruit wines are sulphur-containing volatiles, acetic acids and free amino nitrogen [14]. The volatile compounds containing sulphur, formed as a part of sulphur metabolism, include both molecules positively correlated to the aromatic profile of the wine (volatile thiols), as well as those responsible for wine defects, such as notes described as cabbage, onion, rotten egg, garlic, sulphur and rubber. Hydrogen sulphide and mercaptans (e.g. dimethyl sulphide, dimethyl disulphide, dimethyl trisulphide) are the most often linked to the formation of off-odours in wine, and they are usually present in very low concentration [67]. However, their perception threshold is also low [68].

#### **2.3. Phenolic compounds**

dominant aroma component was isoamyl alcohol (232.0 mg/mL). The same technique was used for the investigation of the temperature effect during apple winemaking on both the critical aroma compounds and sensory properties of wine. The concentration of all aroma compounds was changed with a temperature increase, and sensory analysis showed the highest acceptance of apple wine fermented at 20°C [61]. The influence of apple variety (Šampion, Idared and Gloster) harvested from the orchard in Poland on the volatile composition, and sensory characteristics of apple wine were investigated by Satora et al. [62]. The high concentration of acetaldehyde, ethyl acetate and methanol was found in Šampion wines, while Gloster wines contained a higher concentration of fusel alcohols. The Idared wines had the best results of the sensory evaluation and high levels of butanol and acetic acid. As mentioned earlier, the majority of investigations were performed using sample preparation procedures such as HS and/or SPME, while gas chromatography with FID and MS detectors was shown to be the analytical technique of choice for the determination of volatile compounds in fruit wine. Ye et al. [63] have developed and applied a new rapid method for the identification of volatile compounds in apple wine using FT-NIR spectroscopy. To provide the greater insight into yeast metabolism and flavour formation of mulberry wine, Butkhup et al. [64] have developed a new HS-SPME-GC-MS method to characterise various volatile compounds of in-house-made mulberry wine. Eighty volatile compounds belonging to groups of higher alcohols, fatty acids, esters and phenols were quantified. These compounds were present in various amounts from 0.1 mg/L (benzene carboxaldehyde) to 138.4 mg/mL (isoamyl alcohol). Feng et al. [65] have extended their research on other fruit wines produced from mulberry as well as raspberry and strawberry. Alcohols formed the most abundant group, followed by esters and acids. Comparing to investigated grape wine, two alcohols, 4-methyl-2-pentanol and 2,3-butanediol, were not found in the three fruit wines. While the number of esters in raspberry (1.5%) and mulberry (2.1%) wines were higher than those of strawberry (0.8%) wine, there were no significant differences in acid content. Song et al. [66] have used the same sample preparation and analytical techniques to determine 78 volatile compounds in wild strawberry wine sample from a southern region of China. Odour activity values were detected for 21 compounds while 6 of them were identified as the particular aroma substances for wild strawberry wine, in particular, methyl 2-methylbutyrate, ethyl 2-methylbutyrate,

methyl 3-methylbutyrate, (E)-3-hexen-1-ol, 1-octen-3-ol and phenylacetaldehyde.

in very low concentration [67]. However, their perception threshold is also low [68].

The formation of off-flavours represents the core problem of high-quality fruit wine production. The main compounds responsible for off-flavours of fruit wines are sulphur-containing volatiles, acetic acids and free amino nitrogen [14]. The volatile compounds containing sulphur, formed as a part of sulphur metabolism, include both molecules positively correlated to the aromatic profile of the wine (volatile thiols), as well as those responsible for wine defects, such as notes described as cabbage, onion, rotten egg, garlic, sulphur and rubber. Hydrogen sulphide and mercaptans (e.g. dimethyl sulphide, dimethyl disulphide, dimethyl trisulphide) are the most often linked to the formation of off-odours in wine, and they are usually present

*2.2.1. Volatile compounds containing sulphur*

64 Descriptive Food Science

Wine quality is significantly influenced by phenolic compounds, namely anthocyanins, flavonols, catechins and other flavonoids since they have an enormous impact on the sensory characteristics of wines, mainly colour and astringency [69]. Furthermore, they exhibit a range of antioxidant and pharmacological effects. Phenolic compounds can be classified in different ways because they are constituted at a large number of heterogeneous structures that range from simple molecules to highly polymerised compounds. A review of the composition and content of phenolic compounds in various fruit wines is presented in **Table 1**. Based on the total phenolic content, wines are commonly categorised to three major groups: (i) high in total phenolics, (ii) moderately high in total phenolics and (iii) low in total phenolics. The total phenolic content of cherry, raspberry, black currant, bilberry, elderberry and sea buckthorn fruit wines proved to be comparable or even higher than that of grape red wines, while apple, plum and peach fruit wines had a lower total phenolic content than red grape wines [20, 87–89]. Among the phenolic compounds with known antioxidant activity, flavonoids, phenolic acids and tannins are highlighted [26].

*Flavonoids*, the most studied phenolics, are important components of the human diet. They are also the most widely present in plants and are among the most potent antioxidants in plants. Different members of certain classes of flavonoids can be either colourless or coloured (e.g. anthocyanins), such as pigments of flowers and other plant parts [26]. Important subclasses of flavonoids are, among others, anthocyanins and flavanols. Anthocyanins are important polyphenolic components of fruits, especially berries [90]. The anthocyanins are conjugated anthocyanidins, and they provide the unique colours of dark berries. There are six anthocyanidins distributed throughout the plant kingdom: cyanidin, malvidin, delphinidin, peonidin, pelargonidin and petunidin. The significant anthocyanins in blackberries are cyanidine derivatives. In nature, anthocyanins are mainly found as heterosides. The aglycone form of anthocyanin, known as anthocyanidin, is structurally based on the flavilium or 2-phenylbenzopyrilium cation, with methoxyl and hydroxyl groups present at different positions of the basic structure. Numerous studies have shown that anthocyanins are absorbed in their original glycosylated forms in humans [14, 91, 92]. Anthocyanins are predominant phenolic components or comprise a considerable portion of the total polyphenol content of blackberry as well as cherry juice [76, 84, 93–95]. They are found in berry wines and some fruit wines and presented as total anthocyanin content (TAC). The TAC values found in some blackberry and blueberry wines were 75.56 and 20.82 mg/L (expressed as ellagic acid equivalents), respectively [79]. Mudnić et al. [96] determined high TAC values in blackberry wines, comparable to that measured in red grape wine and ranging from 134 to 164 mg/L (expressed as malvidin 3-glucoside equivalents). Flavonols, another important class of flavonoids, are characteristic for the cool climate fruits and their respective wines [14]. Myricetin and quercetin have been recognised as the first flavonols detected in red berry wines [5]. The results of a study conducted by Mudnić et al. [73] that compared flavanol and procyanidin B<sup>2</sup> contents of several grape wines (red and white), with four commercially available blackberry wines, revealed that the highest concentrations of catechin (45.2 mg/L) and epicatechin (34.7 mg/L) were determined in two blackberry wines. The content of procyanidin B<sup>2</sup> (77.1 mg/L) of the


**Phenolic compound Blackberry wine (no.** 

*Anthocyanins*

*Phenolic acids*

**of samples) [Ref.]**

**Cherry wine (no. of samples) [Ref.]**

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial…

<LOQ-3.78 (5) [84]

1.3–10.2 (5) [84]

7.4–361.3 (9) [74] 10.1–44.6 (5) [84]

1.7–46.3 (9) [74] 0.6–2.5 (3) [62]

Cyanidin <LOD-3.2 (15) [43] ND-4.2 (5) [84] NF <0.4–12.3 [86]

Cyanidin 3-glucoside 0.39–25.51 (13) [74] 0.36–0.68 (6) [44] NF NF 8–18 (10) [77] 0.78–12.07 (9) [74]

Cyanidin 3-rutinoside 1.9–119.3 (13) [74] 10.7–15.1 (6) [44] NF NF 25–69 (10) [77] 3.2–91.4 (9) [74]

Cyanidin 3-glucosylrutinoside NF 17.6–25.4 (6) [44] NF NF

Pelargonidin <LOD-1.46 (15) [43] NF NF NF

10.8–52.3 (13) [74] 1.1 (1) [78]

Chlorogenic acid 1.0–3.9 (17) [70] 21.7–29.6 (6) [44] 13.3–25.7 (3) [74] NF

Neochlorogenic acid NF 12.8–18.5 (6) [44] NF NF

*p*-Hydroxybenzoic acid NF 4.0–4.7 (6) [44] NF 0.4–0.7 (6) [81]

*p*-Coumaric acid 0.1–79.7 (13) [74] 3.3–7.0 (6) [44] <LOQ-0.2 (3) [74] 0.2–14.2 [80–83]

28.1–122.4 (17) [70]

Peonidin 3-rutinosid NF 1.06–1.64 (6) [44] NF 11.4–128.0

Gallic acid 45.4–59.0 (4) [73] <LOQ–5.7 (9) [74] 0.1–1.4 (3) [74] 6.7–104.8 [80–83]

Caffeic acid 2.0–4.8 (17) [70] 0.4–1.7 (6) [44] 1.1–5.4 (3) [74] 0.2–30.8 [80–83]

13.9 (1) [78] 16.3–25.6 (5) [76]

3.6 (1) [78] 27.3–81.0 (5) [76]

6.7 (1) [78]

1.0–4.4 (17) [70] 0.2–22.2 (9) [74] 0.1–0.9 (3) [62] 23.4 (1) [78] 0.6–15.2 (5) [76]

12.2–110. 5 (9) [74]

44.5–71.4 (5) [76]

**Apple wine (no. of samples) [Ref.]**

http://dx.doi.org/10.5772/intechopen.78796

**Wine [Ref.]**

67

(67) [82]


0.6–15.2 (5) [76]

**Phenolic compound Blackberry wine (no.** 

66 Descriptive Food Science

**Individual phenolic compounds**

Quercetin-3-O-rutinoside

*Flavonol*

(Rutin)

*Flavan-3-ol*

*Flavanone*

*Flavone*

**of samples) [Ref.]**

52–105 (19) [77]

**Cherry wine (no. of samples) [Ref.]**

1697–22,789 (4) [73] 1081–2711 (9) [74] 225–645 (3) [74] 190–1215 [75]

NF 0.2 (1) [78] NF 6.8–20.2 (67) [82]

Total phenolics\* 733–2698 (17) [70] 584–743 (6) [44] 160–470 (4) [71] 932–1055 [72]

Total anthocyanins 1.3–125.3 (17) [70] NF 61–125 [72] 13–164 (4) [73] 17.9–27.9 (6) [44] 23–217 (13) [74] 55–483 (9) [74] 18.5–192.0 (6) [79] 120 (1) [78]

Flavonoids\* 924–1417 (4) [73] NF NF NF

0.8–21.7 (15) [43] ND (1) [78] Quercetin-4-glucoside ND-2.6 (4) [73] NF <LOD-0.1 (3) [62] NF

Tannin\* 1010–1312 (10) [77] 741–901 (6) [44] NF 266–414 [72]

Quercetin 0.4–17.9 (13) [74] 0.3–3.9 (9) [74] 0.4–0.9 (3) [74] <LOD-13.4

(+)-Catechin 1.7–10.3 (13) [74] 0.4–11.2 (9) [74] 1.5–15.2 (3) [74] 13.4–95.8 (6) [81]

(−)-Epicatechin ND-34.7 (4) [73] 0.5–24.1 (9) [74] 4.1–25.9 (3) [74] 4.4–68.5 [80–83]

Epigallocatechin gallate ND-148.8 (4) [73] ND (1) [78] NF <LOD-15.6 [85]

Procyanidin B2 6.1–77.1 (4) [73] 24.7–69.1 (4) [84] 2.9–7.4 (3) [62] 3.0–83.2 (67) [82]

(−)-Epigallocatechin NF 1.01 (1) [78] NF NF

Naringenin NF 0.15 (1) [78] NF NF

Apigenin NF 0.06 (1) [78] NF NF

9.1–45.2 (4) [73] 12.0–15.2 (4) [84] 1.6–3.5 (3) [79] 1.3 (1) [78]

0.1–61.4 (13) [74] 10.3–20.91 (4) [84] 3.5–6.6 (3) [62] 3.91 (1) [78]

1055–2704 (13) [74] 850–1300 (5) [76] 451 (6) [20] 1052–3621 (6) [73] 991 (6) [20] 228–639 (3) [62] 1122–1400 (10) [77] 1940 (1) [78] 471–801 (40) [77]

**Apple wine (no. of samples) [Ref.]** **Wine [Ref.]**

[80–83]


and often contradictory. Amidžić Klarić et al. [43] evaluated the quercetin content, colour and selected physicochemical quality parameters of Croatian blackberry wines produced using organically and conventionally grown blackberries. Quercetin content of organic wine samples group was slightly higher than that of conventional wine samples group. No significant overall differences were found in case of colour. Phenolic compounds, in general, are not well absorbed, and it is supposed that the presence of ethanol enhances their bioavailability. This can explain the reported results, indicating that (grape) wines offer somewhat higher health benefits than the preparation of isolated phenolic extracts alone or dealcoholised wine [101, 102]. Similar investigations of fruit wines are still lacking. A progressive loss of phenolic compounds can occur during the winemaking process, as a result of chemical reactions, such as degradation, oxidation, precipitation of phenolics with polysaccharides, condensation with tannins and the formation of other stable anthocyanin-derived pigments. All these reactions can result in significant changes in colour and flavour of red (grape) wines [103, 104]. The same could be applied to other fruit wines. Rommel et al. [93] investigated the processing and storage effects on the composition of anthocyanin, colour and appearance of blackberry juice and wine. They reported 85–100% degradation of juice total anthocyanins, caused by

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69

Vitamins are compounds essential for both plant and animal metabolism, and their deficiencies in humans often lead to different disorders and diseases. Because of their ability to synthesise vitamins, plants (namely fruits and vegetables) are an excellent source of the vitamins in the diet. The stability of vitamins in different foods is often at risk due to various technological practices used during the processing of food, namely changes in temperature regimes (e.g. thermal treatments) and oxygen levels [105, 106]. Since fruits are a good source of vitamins, consequently the vitamins can also be found in fruit juices. However, the concentration of vitamins decreases during the winemaking (fermentation and ageing), so their levels in wine are inadequate to be of significance in human nutrition. On the other hand, their levels are usually enough or more than enough to support microbial growth. Biotin (vitamin H) and

of vitamins determined in grape wines are probably the reason for the absence of relevant studies regarding the vitamins in fruit wines. Grape wines are reported to contain some B vitamins (thiamine, riboflavin and vitamin B12) and very small amounts of vitamin C and fatsoluble vitamins A, D and K. Some vitamins, such as vitamin C, may be primarily destroyed during processing and storage, whereas others, for example, B vitamins, are also produced by the action of yeast and may be present in measurable quantities in the final product [107].

light [33]. Free riboflavin (RF) is naturally present in raw and processed fruits and fermented beverages. Flavin-adenine dinucleotide (FAD) and sometimes flavin mononucleotide (FMN) are present together with RF in significant amounts in fruit juices, while RF is the only form of riboflavin present in important quantities in wine [108]. Since some yeasts synthesise RF

nificant. In addition, thiamine levels are lowered by reaction with SO<sup>2</sup>

as well as the absorption by bentonite. Riboflavin (vitamin B<sup>2</sup>

) contents are adequate for most yeast strains [33]. Low levels

during fermentation

) is oxidised on exposure to

) is utilised by yeast during fermentation, so its levels in wine are insig-

fermentation in combination with depectinising and storage time.

**2.4. Vitamins**

nicotinic acid (niacin, vitamin B<sup>3</sup>

Thiamine (vitamin B<sup>1</sup>

All concentrations are expressed in mg/L.

ND, not detected; NF, not found; LOD, limit of detection; LOQ, limit of quantification.

\* Values are expressed as gallic acid equivalents (milligrams of gallic acid per litre of fruit wine).

**Table 1.** A review of the composition and content of phenolic compounds in various fruit wines.

same wines was sevenfold higher than in red grape wine (12.3 mg/L). Cherry wines proved to be a much more potent source of flavonols than grape wines, with almost 10 times higher values (from 24.2 to 69.08 g/L for procyanidin B<sup>2</sup> ) [84]. The flavanol profile of cherry wine is similar to its grape counterparts, with values up to 3.92 mg/kg for epicatechin, followed by smaller amounts of catechin and epigallocatechin [78]. The major flavanol in apple wines is epicatechin, followed by catechin and their polymerisation product procyanidin B<sup>2</sup> [62].

*Phenolic acids.* P (Phenolic acids) are usually divided into hydroxybenzoic acid derivatives (such as gallic, hydroxybenzoic, salicylic and protocatechuic) and diverse hydroxycinnamic acids and their derivatives. Blackberry and wine made from this edible fruit are known to have high concentrations of benzoic and cinnamic acids [19, 21], especially gallic acid, chlorogenic acid, *p*-coumaric acid and caffeic acid [70]. Cherry wines have a significantly higher content of phenolic acids in comparison to grape wine and almost 10 times higher content of caffeic acid. The prevalent hydroxycinnamic acids in apple wines are chlorogenic and *p*-coumaroylquinic acid. However, they are susceptible to very fast oxidation catalysed by enzyme polyphenol oxidase, giving *o*-quinone, a compound responsible for the browning of apple must, as the principal product [62]. It has been reported in the literature that organically produced fruits may have a higher level of phenolic compounds. This can probably be explained by the enhanced synthesis of endogenous natural defence substances (phenolics) enriching plant defence mechanisms, that is, produced as a response to biotic and abiotic stress [97], in the absence of fertilisers and pesticides, commonly used during conventional production [98–100]. Even though there are generally recognised differences between the conventionally and organically grown food, namely the yield, the size and the pesticide residues, the data on the impact of the practices mentioned earlier on nutritional quality are scarce and often contradictory. Amidžić Klarić et al. [43] evaluated the quercetin content, colour and selected physicochemical quality parameters of Croatian blackberry wines produced using organically and conventionally grown blackberries. Quercetin content of organic wine samples group was slightly higher than that of conventional wine samples group. No significant overall differences were found in case of colour. Phenolic compounds, in general, are not well absorbed, and it is supposed that the presence of ethanol enhances their bioavailability. This can explain the reported results, indicating that (grape) wines offer somewhat higher health benefits than the preparation of isolated phenolic extracts alone or dealcoholised wine [101, 102]. Similar investigations of fruit wines are still lacking. A progressive loss of phenolic compounds can occur during the winemaking process, as a result of chemical reactions, such as degradation, oxidation, precipitation of phenolics with polysaccharides, condensation with tannins and the formation of other stable anthocyanin-derived pigments. All these reactions can result in significant changes in colour and flavour of red (grape) wines [103, 104]. The same could be applied to other fruit wines. Rommel et al. [93] investigated the processing and storage effects on the composition of anthocyanin, colour and appearance of blackberry juice and wine. They reported 85–100% degradation of juice total anthocyanins, caused by fermentation in combination with depectinising and storage time.

#### **2.4. Vitamins**

same wines was sevenfold higher than in red grape wine (12.3 mg/L). Cherry wines proved to be a much more potent source of flavonols than grape wines, with almost 10 times higher

**Cherry wine (no. of samples) [Ref.]**

23.9 (1) [78]

1.9–13.4 (9) [74]

Ferulic acid NF ND-4.2 (5) [76] NF 0.1–0.2 (6) [81]

*trans*-Resveratrol 0.5–0.9 (4) [73] NF ND (1) [78] 0.02–1.46 (6) [81]

similar to its grape counterparts, with values up to 3.92 mg/kg for epicatechin, followed by smaller amounts of catechin and epigallocatechin [78]. The major flavanol in apple wines is

*Phenolic acids.* P (Phenolic acids) are usually divided into hydroxybenzoic acid derivatives (such as gallic, hydroxybenzoic, salicylic and protocatechuic) and diverse hydroxycinnamic acids and their derivatives. Blackberry and wine made from this edible fruit are known to have high concentrations of benzoic and cinnamic acids [19, 21], especially gallic acid, chlorogenic acid, *p*-coumaric acid and caffeic acid [70]. Cherry wines have a significantly higher content of phenolic acids in comparison to grape wine and almost 10 times higher content of caffeic acid. The prevalent hydroxycinnamic acids in apple wines are chlorogenic and *p*-coumaroylquinic acid. However, they are susceptible to very fast oxidation catalysed by enzyme polyphenol oxidase, giving *o*-quinone, a compound responsible for the browning of apple must, as the principal product [62]. It has been reported in the literature that organically produced fruits may have a higher level of phenolic compounds. This can probably be explained by the enhanced synthesis of endogenous natural defence substances (phenolics) enriching plant defence mechanisms, that is, produced as a response to biotic and abiotic stress [97], in the absence of fertilisers and pesticides, commonly used during conventional production [98–100]. Even though there are generally recognised differences between the conventionally and organically grown food, namely the yield, the size and the pesticide residues, the data on the impact of the practices mentioned earlier on nutritional quality are scarce

epicatechin, followed by catechin and their polymerisation product procyanidin B<sup>2</sup>

) [84]. The flavanol profile of cherry wine is

**Apple wine (no. of samples) [Ref.]** **Wine [Ref.]**

[62].

values (from 24.2 to 69.08 g/L for procyanidin B<sup>2</sup>

All concentrations are expressed in mg/L.

**Phenolic compound Blackberry wine (no.** 

*Stilbene*

68 Descriptive Food Science

\*

**of samples) [Ref.]**

ND, not detected; NF, not found; LOD, limit of detection; LOQ, limit of quantification.

Values are expressed as gallic acid equivalents (milligrams of gallic acid per litre of fruit wine).

**Table 1.** A review of the composition and content of phenolic compounds in various fruit wines.

*p*-Coumarylquinic acid NF NF 0.3–4.3 (3) [62] NF Protocatechuic acid 13.8–62.6 (13) [74] <LOQ–15.4 (9) [74] NF NF

*cis*-Resveratrol ND-1.5 (4) [73] NF NF NF Astringin ND-12.1 (4) [73] NF NF NF (E)-Piceid 2.0–6.5 (4) [73] NF NF NF

> Vitamins are compounds essential for both plant and animal metabolism, and their deficiencies in humans often lead to different disorders and diseases. Because of their ability to synthesise vitamins, plants (namely fruits and vegetables) are an excellent source of the vitamins in the diet. The stability of vitamins in different foods is often at risk due to various technological practices used during the processing of food, namely changes in temperature regimes (e.g. thermal treatments) and oxygen levels [105, 106]. Since fruits are a good source of vitamins, consequently the vitamins can also be found in fruit juices. However, the concentration of vitamins decreases during the winemaking (fermentation and ageing), so their levels in wine are inadequate to be of significance in human nutrition. On the other hand, their levels are usually enough or more than enough to support microbial growth. Biotin (vitamin H) and nicotinic acid (niacin, vitamin B<sup>3</sup> ) contents are adequate for most yeast strains [33]. Low levels of vitamins determined in grape wines are probably the reason for the absence of relevant studies regarding the vitamins in fruit wines. Grape wines are reported to contain some B vitamins (thiamine, riboflavin and vitamin B12) and very small amounts of vitamin C and fatsoluble vitamins A, D and K. Some vitamins, such as vitamin C, may be primarily destroyed during processing and storage, whereas others, for example, B vitamins, are also produced by the action of yeast and may be present in measurable quantities in the final product [107]. Thiamine (vitamin B<sup>1</sup> ) is utilised by yeast during fermentation, so its levels in wine are insignificant. In addition, thiamine levels are lowered by reaction with SO<sup>2</sup> during fermentation as well as the absorption by bentonite. Riboflavin (vitamin B<sup>2</sup> ) is oxidised on exposure to light [33]. Free riboflavin (RF) is naturally present in raw and processed fruits and fermented beverages. Flavin-adenine dinucleotide (FAD) and sometimes flavin mononucleotide (FMN) are present together with RF in significant amounts in fruit juices, while RF is the only form of riboflavin present in important quantities in wine [108]. Since some yeasts synthesise RF

during cell growth as an extracellular by-product of the fermentation process, its content can increase in wine. *p*-aminobenzoic acid (PABA), classified into a group of vitamin-like substances, although often referred to as a vitamin [109], is the only compound to increase substantially during fermentation [33]. Folic acid (pteroylglutamic acid) is a water-soluble B vitamin, essential for healthy growth and development of humans, can be found in significant amounts in berries [110] and can reach up to 1 mg/100 mL in some berry wines [14]. During the process of alcoholic fermentation, this vitamin is not subject to change because it has no significant effect on yeast, while for lactic acid bacteria, it is essential [111]. Vitamin C is one of the essential nutritional quality factors of fruits and has many vital biological roles in the human body, such as cell division and proliferation, photosynthesis, hormone biosynthesis and signalling [112, 113]. It is a crucial antioxidant that is produced by plants response to various biotic and abiotic stresses. Vitamin C plays a fundamental role in scavenging reactive oxygen species (ROS) due to its potent antioxidant properties [113]. Fruits and vegetables provide more than 90% of the vitamin C in human diets [112]. The reported vitamin C concentration ranges in blackberries, cherries and apples are 87–696 mg/L [114], 43–177 mg/L [115] and 3–25 mg/100 mg of fresh weight [116], respectively. Previous investigation has indicated that the content of ascorbic acid varied for organically and conventionally cultivated fruits and vegetables [117], as well as for different environmental conditions under which the plant was grown, such as temperature, water availability, pathogenic attack and nutrients [118]. Taken together, the facts listed earlier lead to a conclusion that moderate wine consumption is very far from providing an adequate intake of vitamins for maintaining optimal nutritional status and overall health [14].

cherry juice and cherry wines and concluded that cherry wine had enhanced levels of the mentioned enzymes. Superoxide dismutases (SODs) are the major antioxidant defence systems against superoxide anion, the most common free radical in the body [123]. SODs reduce the cellular damage caused by superoxide anion. When comparing different fruit wines, it has been reported that berry fruit wines (blackberry, elderberry, blueberry and raspberry) in general had higher total antioxidant capacities than apple and pear wines (as representatives

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Minerals are inorganic ingredients of food, required by the body in small amounts for a variety of different metabolic processes important for the functioning of human body. Since some minerals are needed in more substantial quantities than others, they can be divided to three groups, based on their recommended daily intake: macrominerals (over 50 mg/day), trace minerals and ultratrace minerals (lower than 50 mg/day) [124–126]. Nowadays, a mineral deficiency occurs very often without its noticeable signs. The sufficient intake of minerals depends on their quantities consumed with food, but even more on their bioavailability, which depends on the food composition and properties. The mineral content of fruits can vary significantly depending on the genetic and climate factors, cultivation procedures, soil composition, ripeness and many other factors. Furthermore, changes in mineral content usually occur as a result of applied technological procedures, for example, the removal of a portion of the raw material [125, 126]. The sources of minerals (metals) in alcoholic beverages are various: raw materials, process equipment, applied technological procedures, fermentation, bottling, ageing, storage and adulteration [127]. The soil on which fruits are grown is a source of metals of natural origin (primary metals) present in wine. The concentration of primary metals is characteristic and comprises the most significant part of the total metal content in wine [128, 129]. External impurities present in fruits, as well as those that occur during different stages of winemaking, are the source of metals of secondary origin. For example, the prolonged contact of wine and the construction materials used for the oenological equipment (stainless steel, aluminium, brass, glass and wood) can be a source of contamination of wine with Al, Cd, Cr, Cu, Fe and Zn [128–130]. Fining and clarifying substances (e.g. bentonites) used during winemaking can be a source of contamination with Na, Ca or Al [129–132]. Furthermore, individual metals have a significant effect on the organoleptic properties of wine (i.e. Al, Zn, Cu and Fe) and, therefore, their concentration must be monitored [133]. The total amount of minerals in a sample of fruits and fruit wines can be expressed as the ash content. Ash is the inorganic residue remaining after the water, and organic matter has been removed. A higher ash content in a fruit wine implies a higher amount of minerals and a higher quality of the wine [134]. The literature regarding grape wines proposes that the mineral profile of wines could be used as a 'fingerprint', used for the characterisation of wines based on their geographical origin [20, 135]. While there is a significant volume of available data on the mineral content of blackberries, cherries and apples, far too little attention has been paid to the mineral composition of respective fruit wines. A review of metal concentrations in fruit wines is given in **Table 2**. As it could be seen from the table,

of pome wines) [14, 20].

**2.6. Mineral composition**

#### **2.5. Antioxidant capacity**

Fruits contain various dietary phytonutrients with strong antioxidant capacities, such as phenolics (which include flavonoids and phenolic acids), carotenoids and vitamins. Many phenolic substances present in fruits influence sensory properties of fruit wine such as taste, astringency, bitterness as well as colour. Since fruit wines are processed in the same way as wine made from grapes, significant compositional changes take part in winemaking [119]. Both fermentation and ageing result in the transformation of starting compounds present in musts into secondary metabolites that determine the quality of the final product [119, 120]. Berries, grape and their processed products such as wines contain a wide range of flavanoids and other phenolic compounds that possess antioxidant activity. Heinonen et al. [71] evaluated the antioxidant activity of over 44 different fruit wines, mainly berry wines, but also apple wines. The results of the study showed that all the investigated wines possessed a significant antioxidant activity. However, the total phenolic content did not correlate with the antioxidant activity. On the other hand, several other studies confirmed a strong positive correlation between the total antioxidant activity of fruit wines and total phenolics [26, 28, 31, 69, 95, 121]. When compared to red (grape) wines, blackberry, blueberry and sour cherry wines proved to have 30–40% more superoxide radical-scavenging activity [122], which makes them very potent natural antioxidants beneficial to human health [74, 76, 84, 94]. Yoo et al. [12] compared the levels of enzymes superoxide dismutase (SOD) and catalase in fresh cherries, cherry juice and cherry wines and concluded that cherry wine had enhanced levels of the mentioned enzymes. Superoxide dismutases (SODs) are the major antioxidant defence systems against superoxide anion, the most common free radical in the body [123]. SODs reduce the cellular damage caused by superoxide anion. When comparing different fruit wines, it has been reported that berry fruit wines (blackberry, elderberry, blueberry and raspberry) in general had higher total antioxidant capacities than apple and pear wines (as representatives of pome wines) [14, 20].

#### **2.6. Mineral composition**

during cell growth as an extracellular by-product of the fermentation process, its content can increase in wine. *p*-aminobenzoic acid (PABA), classified into a group of vitamin-like substances, although often referred to as a vitamin [109], is the only compound to increase substantially during fermentation [33]. Folic acid (pteroylglutamic acid) is a water-soluble B vitamin, essential for healthy growth and development of humans, can be found in significant amounts in berries [110] and can reach up to 1 mg/100 mL in some berry wines [14]. During the process of alcoholic fermentation, this vitamin is not subject to change because it has no significant effect on yeast, while for lactic acid bacteria, it is essential [111]. Vitamin C is one of the essential nutritional quality factors of fruits and has many vital biological roles in the human body, such as cell division and proliferation, photosynthesis, hormone biosynthesis and signalling [112, 113]. It is a crucial antioxidant that is produced by plants response to various biotic and abiotic stresses. Vitamin C plays a fundamental role in scavenging reactive oxygen species (ROS) due to its potent antioxidant properties [113]. Fruits and vegetables provide more than 90% of the vitamin C in human diets [112]. The reported vitamin C concentration ranges in blackberries, cherries and apples are 87–696 mg/L [114], 43–177 mg/L [115] and 3–25 mg/100 mg of fresh weight [116], respectively. Previous investigation has indicated that the content of ascorbic acid varied for organically and conventionally cultivated fruits and vegetables [117], as well as for different environmental conditions under which the plant was grown, such as temperature, water availability, pathogenic attack and nutrients [118]. Taken together, the facts listed earlier lead to a conclusion that moderate wine consumption is very far from providing an adequate intake of vitamins for maintaining optimal nutritional

Fruits contain various dietary phytonutrients with strong antioxidant capacities, such as phenolics (which include flavonoids and phenolic acids), carotenoids and vitamins. Many phenolic substances present in fruits influence sensory properties of fruit wine such as taste, astringency, bitterness as well as colour. Since fruit wines are processed in the same way as wine made from grapes, significant compositional changes take part in winemaking [119]. Both fermentation and ageing result in the transformation of starting compounds present in musts into secondary metabolites that determine the quality of the final product [119, 120]. Berries, grape and their processed products such as wines contain a wide range of flavanoids and other phenolic compounds that possess antioxidant activity. Heinonen et al. [71] evaluated the antioxidant activity of over 44 different fruit wines, mainly berry wines, but also apple wines. The results of the study showed that all the investigated wines possessed a significant antioxidant activity. However, the total phenolic content did not correlate with the antioxidant activity. On the other hand, several other studies confirmed a strong positive correlation between the total antioxidant activity of fruit wines and total phenolics [26, 28, 31, 69, 95, 121]. When compared to red (grape) wines, blackberry, blueberry and sour cherry wines proved to have 30–40% more superoxide radical-scavenging activity [122], which makes them very potent natural antioxidants beneficial to human health [74, 76, 84, 94]. Yoo et al. [12] compared the levels of enzymes superoxide dismutase (SOD) and catalase in fresh cherries,

status and overall health [14].

**2.5. Antioxidant capacity**

70 Descriptive Food Science

Minerals are inorganic ingredients of food, required by the body in small amounts for a variety of different metabolic processes important for the functioning of human body. Since some minerals are needed in more substantial quantities than others, they can be divided to three groups, based on their recommended daily intake: macrominerals (over 50 mg/day), trace minerals and ultratrace minerals (lower than 50 mg/day) [124–126]. Nowadays, a mineral deficiency occurs very often without its noticeable signs. The sufficient intake of minerals depends on their quantities consumed with food, but even more on their bioavailability, which depends on the food composition and properties. The mineral content of fruits can vary significantly depending on the genetic and climate factors, cultivation procedures, soil composition, ripeness and many other factors. Furthermore, changes in mineral content usually occur as a result of applied technological procedures, for example, the removal of a portion of the raw material [125, 126]. The sources of minerals (metals) in alcoholic beverages are various: raw materials, process equipment, applied technological procedures, fermentation, bottling, ageing, storage and adulteration [127]. The soil on which fruits are grown is a source of metals of natural origin (primary metals) present in wine. The concentration of primary metals is characteristic and comprises the most significant part of the total metal content in wine [128, 129]. External impurities present in fruits, as well as those that occur during different stages of winemaking, are the source of metals of secondary origin. For example, the prolonged contact of wine and the construction materials used for the oenological equipment (stainless steel, aluminium, brass, glass and wood) can be a source of contamination of wine with Al, Cd, Cr, Cu, Fe and Zn [128–130]. Fining and clarifying substances (e.g. bentonites) used during winemaking can be a source of contamination with Na, Ca or Al [129–132]. Furthermore, individual metals have a significant effect on the organoleptic properties of wine (i.e. Al, Zn, Cu and Fe) and, therefore, their concentration must be monitored [133]. The total amount of minerals in a sample of fruits and fruit wines can be expressed as the ash content. Ash is the inorganic residue remaining after the water, and organic matter has been removed. A higher ash content in a fruit wine implies a higher amount of minerals and a higher quality of the wine [134]. The literature regarding grape wines proposes that the mineral profile of wines could be used as a 'fingerprint', used for the characterisation of wines based on their geographical origin [20, 135]. While there is a significant volume of available data on the mineral content of blackberries, cherries and apples, far too little attention has been paid to the mineral composition of respective fruit wines. A review of metal concentrations in fruit wines is given in **Table 2**. As it could be seen from the table,


the mineral content of blackberry, cherry and apple wine is in accordance with that of grape wines. Predominant metal in all reviewed fruit wines is K, its concentration being the highest in blackberry wine. Na is a widespread metal in food since the industrially processed food is a particularly important source of this macroelement. The sodium concentrations of blackberry wines varied over a wide range when compared to other fruit wines (**Table 2**). Still, the lowsodium/high-potassium levels found in all three types of fruit wine indicate that fruit wines can be considered as effective sources of potassium. Due to irregular diet and large intake of refined sugars, coffee and alcohol, the insufficient intake of magnesium may occur. Amidžić Klarić et al. [141] investigated the mineral composition of commercially available Croatian blackberry wines. The results indicated that 17 investigated Croatian blackberry wines could be considered as an excellent additional source of magnesium, manganese and potassium. When compared to cherry and apple wines (**Table 2**), blackberry wine seems to be a better source of Ca, Mg, Fe and Mn. The literature data show that calcium and phosphorus absorption is closely related, but due to an unbalanced diet dominated by a high phosphorus intake, homeostasis is compromised and contributes to low serum calcium levels (hypocalcaemia). For this reason, it has been suggested that the calcium-phosphorus ratio in foods should be between 1:1 and 2:1. As can be seen from **Table 2**, the calcium-phosphorus ratio of reviewed blackberry wine samples was higher than 1 in favour of calcium. Iron deficiency is one of the most common nutritional deficiencies, characterised by signs and symptoms such as unusual tiredness, paleness, shortness of breath and anaemia. In Croatia, blackberry wine is traditionally called 'ferrous wine', and it has been used as a popular medicine for anaemia and iron deficiency. Some of the metals exhibit not only beneficial effects on the human body but also deleterious ones that made them a cause of concern for wine producers and consumers for years now. Heavy metals are some of the most critical chemicals found in the environment,

**Cherry wine (no. of samples) [Ref]**

136 μg/g (6) [20]

Se <1.5 μg/L (15) [134] 0.011 mg/kg (1) [78] NF NF

0.3 mg/kg (1) [78] 0.1 μg/g (6) [20]

**Table 2.** A review of metal concentrations in blackberry, cherry and apple wines.

Sb NF 0.093 μg/kg (1) [78] NF 0.18–29.7 μg/L (31) [128]

Si 1.82–8.24 mg/L (15) [134] NF NF 5.30–33.3 mg/L (68) [137] Sn 12.7–22.0 μg/L (15) [134] NF <32.5 μg/L (3) [9] 0.02–1.0 mg/L (68) [137] Sr 165–1445 μg/L (22) [136] NF NF 0.12–3.22 μg/mL [129]

V NF 6.86 μg/kg (1) [78] NF ND-0.40 μg/mL [129]

**Apple wine (no. of samples) [Ref]**

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial…

36–105 μg/L (3) [9] 0.2 μg/g (6) [20]

80 μg/g (6) [20] NF

**Grape wine [Ref]**

73

http://dx.doi.org/10.5772/intechopen.78796

0.27–1.84 mg/L (54) [140]

ND-8.9 μg/mL [129]

**Metal Blackberry wine (no. of samples) [Ref]**

Zn 0.25–6.65 mg/L (32) [134, 138] 0.30–1.96 mg/L (22) [136]

ND, not detected; NF, not found.

S NF 0.089 g/kg (1) [78]

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial… http://dx.doi.org/10.5772/intechopen.78796 73


**Table 2.** A review of metal concentrations in blackberry, cherry and apple wines.

**Metal Blackberry wine (no. of samples) [Ref]**

72 Descriptive Food Science

Al 6–16 mg/L (15) [134]

37–1110 μg/L (22) [136]

Ca 86–457 mg/L (32) [134, 138] 115–555 mg/L (22) [136]

Co 1.3–11.9 μg/L (32) [134, 138] 2–40 μg/L (22) [136]

Cu 0.06–0.77 mg/L (32) [134, 138] 0.05–3.83 mg/L (22) [136]

Fe 0.082–8.4 mg/L (32) [134, 138] 0.093–5.49 mg/L (22) [136]

K 564–2014 mg/L (32) [134, 138] 615–1760 mg/L (22) [136]

Mg 706–381 mg/L (32) [134, 138] 77–238 mg/L (22) [136]

Mn 0.7–11.5 mg/L (32) [134, 138] 0.5–11.3 mg/L (22) [136]

Na 12–213 mg/L (32) [134, 138] 3–13 mg/L (22) [136]

Pb 13.6–52.8 μg/L (15) [134] 17–54 μg/L (22) [136]

P 32–119 mg/L (15) [43] 0.179 g/kg (1) [78]

Li 13–21 μg/L (15) [134] 1–40 μg/L (22) [136]

**Cherry wine (no. of samples) [Ref]**

0.084 g/kg (1) [78] 69 μg/g (6) [20]

Ag 0.34–2.76 μg/L (15) [134] NF NF 0.008–2.77 μg/L (31) [128]

As <0.9 μg/L (15) [134] 0.093 μg/kg (1) [78] NF 1.7–15.2 μg/L (31) [128] B NF 2.760 mg/kg (1) [78] NF 2.1–12.1 mg/L (68) [137] Ba 0.48–1.46 mg/L (15) [134] NF NF 0.01–0.48 μg/mL [129]

Cd 0.3–9.9 μg/L (32) [134, 138] 0.093 μg/kg (1) [78] <0.026–3.72 μg/L (3) [9] ND-0.054 μg/mL [129]

Cr 2.3–18.7 μg/L (32) [134, 138] 0.016 mg/kg (1) [78] NF ND-0.2 μg/mL [129]

2.192 mg/kg (1) [78] 1.2 μg/g (6) [20]

Hg NF NF <0.036 μg/L (3) [9] NF

1.373 g/kg (1) [78] 834 μg/g (6) [20]

0.072 g/kg (1) [78] 50 μg/g (6) [20]

0.632 mg/kg (1) [78] 0.2 μg/g (6) [20]

1.65 mg/kg (1) [78] 38 μg/g (6) [20]

54 μg/g (6) [20]

Mo NF 0.093 μg/kg (1) [78] NF 0.7–64 μg/L (88) [139]

Ni 60–278 μg/L (15) [134] 0.054 mg/kg (1) [78] NF ND-0.5 μg/mL [129]

Rb 90–1470 μg/L (22) [136] NF NF 0.03–9.90 μg/mL [129]

**Apple wine (no. of samples) [Ref]**

8–28 mg/L (3) [9] 45 μg/g (6) [20]

0.577 μg/kg (1) [78] NF ND-0.045 μg/mL [129]

0.030 mg/kg (1) [78] NF ND-2.60 μg/mL [129]

<0.027–0.508 mg/L (3) [9]

0.4 μg/g (6) [20]

233–353 mg/L (3) [9] 958 μg/g (6) [20]

13–19 mg/L (3) [9] 38 μg/g (6) [20]

4.4 μg/kg (1) [78] 75.3–116.3 μg/L (3) [9] ND-1.1 μg/mL [129]

0.678 μg/kg (1) [78] NF 0.002–0.098 μg/mL [129]

0.2 μg/g (6) [20] ND-5.5 μg/mL [129]

31 μg/g (6) [20] ND-310 μg/mL [129]

68 μg/g (6) [20] 0.3–47.3 mg/L (31) [128]

0.200 mg/kg (1) [78] NF 0.017–14.3 μg/mL [129]

**Grape wine [Ref]**

7–241 μg/mL [129]

0.06–23.7 μg/mL [129]

265–3056 μg/mL [129]

7–718 μg/mL [129]

96–2470 μg/L (31) [128]

the mineral content of blackberry, cherry and apple wine is in accordance with that of grape wines. Predominant metal in all reviewed fruit wines is K, its concentration being the highest in blackberry wine. Na is a widespread metal in food since the industrially processed food is a particularly important source of this macroelement. The sodium concentrations of blackberry wines varied over a wide range when compared to other fruit wines (**Table 2**). Still, the lowsodium/high-potassium levels found in all three types of fruit wine indicate that fruit wines can be considered as effective sources of potassium. Due to irregular diet and large intake of refined sugars, coffee and alcohol, the insufficient intake of magnesium may occur. Amidžić Klarić et al. [141] investigated the mineral composition of commercially available Croatian blackberry wines. The results indicated that 17 investigated Croatian blackberry wines could be considered as an excellent additional source of magnesium, manganese and potassium. When compared to cherry and apple wines (**Table 2**), blackberry wine seems to be a better source of Ca, Mg, Fe and Mn. The literature data show that calcium and phosphorus absorption is closely related, but due to an unbalanced diet dominated by a high phosphorus intake, homeostasis is compromised and contributes to low serum calcium levels (hypocalcaemia). For this reason, it has been suggested that the calcium-phosphorus ratio in foods should be between 1:1 and 2:1. As can be seen from **Table 2**, the calcium-phosphorus ratio of reviewed blackberry wine samples was higher than 1 in favour of calcium. Iron deficiency is one of the most common nutritional deficiencies, characterised by signs and symptoms such as unusual tiredness, paleness, shortness of breath and anaemia. In Croatia, blackberry wine is traditionally called 'ferrous wine', and it has been used as a popular medicine for anaemia and iron deficiency. Some of the metals exhibit not only beneficial effects on the human body but also deleterious ones that made them a cause of concern for wine producers and consumers for years now. Heavy metals are some of the most critical chemicals found in the environment, and the exposure through the consumption of food is frequent, regardless of the mode of food production (conventional or organic) [142]. Amidžić Klarić et al. [134] examined mineral and heavy metal content of blackberry wines made from conventionally and organically grown blackberries. The comparison between these two groups of investigated blackberry wines showed the statistically significant difference in the content of Si and Li, whereas the organic wines group contained higher levels of these compounds. Toxic metals (As, Cd, Hg, Pb) are frequently found to be the primary food contaminants [126], and they can be poisonous and harmful. Therefore, the metal content of grape wine is regulated according to the national legislation and the European Union legislation [141, 143]. Above optimal level, elements, such as Al, Cu, K, Fe, Mn, Zn, may have detrimental effects on wine stability and its commercial acceptability [144].

**3.2. Mycotoxins**

Mycotoxins are secondary metabolites of filamentous fungi that naturally occur in food. Mycotoxins can be present in food when fungi are no longer present, which means that the fungal growth is not necessarily associated with the mycotoxins synthesis [152]. Fruits are susceptible to fungal growth, whose development occurs in between harvest and fermentation process. During fermentation, the fungal growth is inhibited by ethanol and the anaerobic conditions [153]. Organic acids present in fruits (malic, citric and tartaric acids) slow down the bacterial spoilage of fruits by lowering the pH. However, the acidic pH of fruit that varies from <2.5 to 5.0 is tolerable for many fungal species [152]. Different practices applied in fruit cultivation (such as the use of pesticides and different cultivars) and in winemaking (period and conditions of storage, type of maceration, time and temperature of fermentation) can have an impact on mycotoxins accumulation [154]. It seems that organic production could be more affected by mycotoxins since no synthetic fungicides are applied [142]. Several mycotoxins have been reported in fruit juices and wine (the list, however, is not exhaustive): *Alternaria* mycotoxins are produced by *A. alternata*, byssochlamic acid produced by *Byssochlamys nivea* in prune, grape and apple juices, citrinin produced by *Penicillium expansum* in apple juice, ochratoxin A, the most crucial mycotoxin in wine, secreted by *Aspergillus carbonarius* present in harvested grapes and juices and finally patulin (PAT), the most critical mycotoxin in fruit juices, produced by several species such as *B. fulva*, *B. nivea* and *P. expansum* [155]. The most critical mycotoxin in grape wine is the ochratoxin A (OTA), which is produced by *A. carbonarius* and is not degraded during winemaking, fermentation process and storage [155]. OTA has been detected in cherries and strawberries and their associated juices. Since the fungus grows on the exterior surfaces of the fruit, the concentration of OTA tends to be higher in wines that are produced by the increased skin contact that is necessary to extract pigments and tannins. OTA concentration in wine is highest after maceration and tends to diminish during the yeast and malolactic fermentations, probably due to adsorption to yeast cells or degradation by lactic acid bacteria [156]. Toxicological studies have determined that OTA may have several effects on health, such as genotoxicity, carcinogenicity, immunosuppressive properties and nephrotoxicity, and for that reason, the EU regulation (EC no. 1881/2006) [157] has set the acceptable limit for OTA in wine to 2 μg/L. Patulin (PAT), which is the most important mycotoxin in fruit juices, has been studied mainly in fruit juices and particularly in apple juices, although the presence of the compound was also described in the brown rot of other fruits [155]. It is produced by the fungi belonging to the genera *Penicillium*, *Aspergillus* and *Byssochlamys,* and its presence in foodstuffs may be a health hazard since this mycotoxin can cause severe acute (convulsions, nausea and ulceration) and chronic (carcinogenic, genotoxic and immunotoxic) effects in humans [158]. The removal of the damaged or rotten fruit could help reduce the levels of PAT in the juices, but the complete elimination of PAT is not possible since PAT diffuses into the healthy parts of the fruit [152]. Stinson et al. [159] reported the 99% reduction of PAT levels as a result of alcoholic fermentation of apple juice. The Codex Alimentarius [160] and the EU Commission Regulation (EC no. 1881/2006) [157] have established limits for patulin maximum level of 50 μg/kg for apple products intended for human consumption. When it comes to winemaking, it seems that the use of sound manufacturing practices (e.g. fruit selection, handling, sorting, storage, culling and washing) could keep the residual mycotoxin levels of the fruit juices and wine under the maximum allowed limits [155].

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial…

http://dx.doi.org/10.5772/intechopen.78796

75

## **3. Food-safety issues regarding fruit wines**

#### **3.1. Pesticides residues**

Pesticides comprise a numerous and diverse group of chemical compounds with a common characteristic of eliminating pest in agriculture and households. They are widely used in agricultural practice, not only during cultivation but also in post-harvest storage. Pesticide residues may end up in fruit pulp and juices since they can penetrate plant tissues. However, their concentrations are lower than those in the intact fruit [145]. Pesticide residues in fruit wine, like those found in juice, can also be introduced by planting and preservation process [146]. In winemaking, the activity of yeast can be affected by pesticides. The presence of pesticides has been associated with stuck and sluggish fermentation [147]. Most of the pesticides can pass from fruit/grape to must and wine, but the extent to which they pass is different [148, 149]. Organic products, compared to those produced by conventional practices, present some advantages in respect to well-known toxicants, such as pesticides and nitrates [142]. However, caution is needed concerning the problem of pesticide because the number of chemicals to trace is very high [142]. The research, which included the analysis of 25 pesticides in Croatian grape wines, conducted by Vitali Čepo et al. [150], indicated significantly lower total pesticide concentrations and the average number of pesticides per sample of organic grape wines, compared to conventional wines. Pesticide residues not only result in potential health risks for the consumers but also lead to a decrease in the fruit wine quality. Because of the health risk of pesticide residues in juice and fruit wine, it is of particular importance to provide the precise, accurate and reliable result of residues as the scientific basis for ensuring food safety. Determination of pesticides is challenging, because of their chemical diversity and lack of collective analytical methods, which probably explains the fact that the reports regarding the pesticide residues in different fruit wines are still missing. Since the diet is the primary source of exposure of the general population to pesticide residues, the regulatory controls on pesticides by different organisations aim at minimisation of exposure to pesticide residues in food. The great variety of applied pesticides, both within European Union countries (EU) and non-EU countries, as well as the arrival of new plant protectors and chemicals, calls for an ever-expanding list of pesticides along with their accompanying maximum residue limits (MRLs) [151].

#### **3.2. Mycotoxins**

and the exposure through the consumption of food is frequent, regardless of the mode of food production (conventional or organic) [142]. Amidžić Klarić et al. [134] examined mineral and heavy metal content of blackberry wines made from conventionally and organically grown blackberries. The comparison between these two groups of investigated blackberry wines showed the statistically significant difference in the content of Si and Li, whereas the organic wines group contained higher levels of these compounds. Toxic metals (As, Cd, Hg, Pb) are frequently found to be the primary food contaminants [126], and they can be poisonous and harmful. Therefore, the metal content of grape wine is regulated according to the national legislation and the European Union legislation [141, 143]. Above optimal level, elements, such as Al, Cu, K, Fe, Mn, Zn, may have detrimental effects on wine stability and its commercial

Pesticides comprise a numerous and diverse group of chemical compounds with a common characteristic of eliminating pest in agriculture and households. They are widely used in agricultural practice, not only during cultivation but also in post-harvest storage. Pesticide residues may end up in fruit pulp and juices since they can penetrate plant tissues. However, their concentrations are lower than those in the intact fruit [145]. Pesticide residues in fruit wine, like those found in juice, can also be introduced by planting and preservation process [146]. In winemaking, the activity of yeast can be affected by pesticides. The presence of pesticides has been associated with stuck and sluggish fermentation [147]. Most of the pesticides can pass from fruit/grape to must and wine, but the extent to which they pass is different [148, 149]. Organic products, compared to those produced by conventional practices, present some advantages in respect to well-known toxicants, such as pesticides and nitrates [142]. However, caution is needed concerning the problem of pesticide because the number of chemicals to trace is very high [142]. The research, which included the analysis of 25 pesticides in Croatian grape wines, conducted by Vitali Čepo et al. [150], indicated significantly lower total pesticide concentrations and the average number of pesticides per sample of organic grape wines, compared to conventional wines. Pesticide residues not only result in potential health risks for the consumers but also lead to a decrease in the fruit wine quality. Because of the health risk of pesticide residues in juice and fruit wine, it is of particular importance to provide the precise, accurate and reliable result of residues as the scientific basis for ensuring food safety. Determination of pesticides is challenging, because of their chemical diversity and lack of collective analytical methods, which probably explains the fact that the reports regarding the pesticide residues in different fruit wines are still missing. Since the diet is the primary source of exposure of the general population to pesticide residues, the regulatory controls on pesticides by different organisations aim at minimisation of exposure to pesticide residues in food. The great variety of applied pesticides, both within European Union countries (EU) and non-EU countries, as well as the arrival of new plant protectors and chemicals, calls for an ever-expanding list of

pesticides along with their accompanying maximum residue limits (MRLs) [151].

acceptability [144].

74 Descriptive Food Science

**3.1. Pesticides residues**

**3. Food-safety issues regarding fruit wines**

Mycotoxins are secondary metabolites of filamentous fungi that naturally occur in food. Mycotoxins can be present in food when fungi are no longer present, which means that the fungal growth is not necessarily associated with the mycotoxins synthesis [152]. Fruits are susceptible to fungal growth, whose development occurs in between harvest and fermentation process. During fermentation, the fungal growth is inhibited by ethanol and the anaerobic conditions [153]. Organic acids present in fruits (malic, citric and tartaric acids) slow down the bacterial spoilage of fruits by lowering the pH. However, the acidic pH of fruit that varies from <2.5 to 5.0 is tolerable for many fungal species [152]. Different practices applied in fruit cultivation (such as the use of pesticides and different cultivars) and in winemaking (period and conditions of storage, type of maceration, time and temperature of fermentation) can have an impact on mycotoxins accumulation [154]. It seems that organic production could be more affected by mycotoxins since no synthetic fungicides are applied [142]. Several mycotoxins have been reported in fruit juices and wine (the list, however, is not exhaustive): *Alternaria* mycotoxins are produced by *A. alternata*, byssochlamic acid produced by *Byssochlamys nivea* in prune, grape and apple juices, citrinin produced by *Penicillium expansum* in apple juice, ochratoxin A, the most crucial mycotoxin in wine, secreted by *Aspergillus carbonarius* present in harvested grapes and juices and finally patulin (PAT), the most critical mycotoxin in fruit juices, produced by several species such as *B. fulva*, *B. nivea* and *P. expansum* [155]. The most critical mycotoxin in grape wine is the ochratoxin A (OTA), which is produced by *A. carbonarius* and is not degraded during winemaking, fermentation process and storage [155]. OTA has been detected in cherries and strawberries and their associated juices. Since the fungus grows on the exterior surfaces of the fruit, the concentration of OTA tends to be higher in wines that are produced by the increased skin contact that is necessary to extract pigments and tannins. OTA concentration in wine is highest after maceration and tends to diminish during the yeast and malolactic fermentations, probably due to adsorption to yeast cells or degradation by lactic acid bacteria [156]. Toxicological studies have determined that OTA may have several effects on health, such as genotoxicity, carcinogenicity, immunosuppressive properties and nephrotoxicity, and for that reason, the EU regulation (EC no. 1881/2006) [157] has set the acceptable limit for OTA in wine to 2 μg/L. Patulin (PAT), which is the most important mycotoxin in fruit juices, has been studied mainly in fruit juices and particularly in apple juices, although the presence of the compound was also described in the brown rot of other fruits [155]. It is produced by the fungi belonging to the genera *Penicillium*, *Aspergillus* and *Byssochlamys,* and its presence in foodstuffs may be a health hazard since this mycotoxin can cause severe acute (convulsions, nausea and ulceration) and chronic (carcinogenic, genotoxic and immunotoxic) effects in humans [158]. The removal of the damaged or rotten fruit could help reduce the levels of PAT in the juices, but the complete elimination of PAT is not possible since PAT diffuses into the healthy parts of the fruit [152]. Stinson et al. [159] reported the 99% reduction of PAT levels as a result of alcoholic fermentation of apple juice. The Codex Alimentarius [160] and the EU Commission Regulation (EC no. 1881/2006) [157] have established limits for patulin maximum level of 50 μg/kg for apple products intended for human consumption. When it comes to winemaking, it seems that the use of sound manufacturing practices (e.g. fruit selection, handling, sorting, storage, culling and washing) could keep the residual mycotoxin levels of the fruit juices and wine under the maximum allowed limits [155].

#### **3.3. Biogenic amines**

Fermentation by-products, biogenic amines, are low molecular nitrogen compounds formed mainly by decarboxylation of amino acids or by amination and transamination of aldehydes and ketones. In food and beverages, they are synthesised by the enzymes from raw material or are generated by decarboxylation of amino acids by microorganisms [161, 162]. Biogenic amines are undesirable in all foods and beverages because if absorbed at too high concentrations, they may cause adverse physiological effects in sensitive humans, especially in the presence of alcohol and acetaldehyde. The most studied biogenic amine is histamine [33, 163]. Biogenic amines, particularly histamine, the most frequently found in wines, often reduce the sensory quality of grape wines. Different microorganisms present in wine can synthesise biogenic amines from their respective amino acid precursors, at any stage of wine production, ageing or storage. Therefore, their presence in wines can serve as an indicator of spoilage and authenticity [164, 165]. Biogenic amines associated with grape wine are putrescine, histamine, tyramine and cadaverine, followed by phenylethylamine, spermidine, spermine, agmatine and tryptamine [162]. Lactic acid bacteria present in wine (e.g. *Lactobacillus*, *Leuconostoc* and *Oenococcus* spp.) are mainly responsible for the occurrence of biogenic amines in wine (and in other fermented products) since they produce enzymes that decarboxylate the respective precursor amino acids [166]. To prevent the problem of a high biogenic amine concentration in wines, the length of the processes of maceration and the contact of wine and lees should be reduced to a minimum, because those are the processes that incorporate amino acids to must or wine. This is hard to achieve when age wines are produced. Another way of solving the problem is to inhibit the growth of indigenous lactic acid bacteria and use the selected *O. oeni* strains unable to produce biogenic amine [167]. Biogenic amines present in fruits also contribute to their level in the wine. It has been reported that fruit cultivation conditions and management technique play essential roles in the accumulation of biogenic amines in fruits, for example, the accumulation of putrescine in grapes as a response to a potassium deficiency in soil [168]. While data are available on the concentrations of biogenic amines in grape wines, rare investigations have been conducted to evaluate their concentrations in fruit wines. Ouyang et al. [169] observed a noticeable difference in the profile of biogenic amines in the wines made from different fruits. A high level of the total biogenic amines (28.11–67.48 mg L−1) was detected in red grape wines, followed by the strawberry wine (14.60 mg L−1) and the raspberry wine (8.75 mg L−1). The total content of BA in the white grape wines ranged from 5.42 to 7.21 mg L−1, while the level of total biogenic amines of other fruit wines was below 2.5 mg L−1. When individual biogenic amines were analysed, the results revealed that spermidine was present in most of the wines, while putrescine was present in all the grape wine samples. However, the levels of putrescine in blueberry, raspberry, schisandrae and strawberry wines were higher than that in the grape wines. Blueberry wine and raspberry wine also contained tyramine.

total SO2

**4. Conclusions**

**Author details**

and Ana Mornar2

Darko Velić<sup>1</sup>

Herzegovina

**References**

in grape wine is set between 150 and 300 mg/L depending on the wine type and the


http://dx.doi.org/10.5772/intechopen.78796

investigated so

77

[171]. The alternatives to SO<sup>2</sup>

Chemical Constituents of Fruit Wines as Descriptors of their Nutritional, Sensorial…

level of reducing substances in wine. One of the challenges for scientists and winemakers who

far include treatments with lysozyme, dimethyl dicarbonate (DMDC), the addition of reduced glutathione (up to 20 mg/L) and the addition of commercial oenological tannins [170, 172–176].

The role of fruits in the human diet is well established due to various important nutritive and biologically active components intrinsic to fruits. Consequently, fruit wines tend to preserve these components, along with developing new desirable ones in the final product. Fruit wines proved to be a good source of different antioxidants, phytonutrients and minerals, which classify them with the functional food. The production of fruit wines is growing steadily in recent years, probably driven by the demand for new functional products. However, having in mind the fact that most fruits can be used for fruit wines production, it is evident that they comprise a vast and diverse group of fermented beverages. Compared to grape wines, the comprehensive reviews of specific fruit wines are still scarce, as well as the comparison of different fruit wines regarding their production, composition, nutritional, functional and health-related properties. Hopefully, this chapter will contribute to the body of knowledge, as well as to diversification, standardisation and, consequently, commercialisation of quality fruit wine production.

, Natalija Velić<sup>1</sup>

1 Faculty of Food Technology Osijek, University of Osijek, Osijek, Croatia

2 Faculty of Pharmacy and Biochemistry, University of Zagreb, Zagreb, Croatia 3 Department of Health Ecology, Public Health Brčko DC, Brčko DC, Bosnia and

4 Faculty of Food Technology and Biotechnology, University of Zagreb, Zagreb, Croatia

foods. Journal of the American Dietetic Association. 2004;**104**(5):814-826 [2] Global Functional Food and Nutraceuticals Market 2016-2020; March 2016

[1] Hasler C, Bloch A, Thomson C. Position of the American dietetic association: Functional

, Ilija Klarić<sup>3</sup>

, Vlatka Petravić Tominac<sup>4</sup>

are striving to meet today's consumer demands for SO<sup>2</sup>

tion of new technologies to replace the use of SO2

\*, Daniela Amidžić Klarić<sup>2</sup>

\*Address all correspondence to: dvelic@ptfos.hr

#### **3.4. Sulphur dioxide**

The most common preservative in wine production used to control both oxidative processes and unwanted spontaneous fermentations is sulphur dioxide (SO<sup>2</sup> ) [39]. This is due to its potent antioxidant and antimicrobial properties. However, when present in high concentrations in wine, SO2 can have unfavourable health effects, for example, diarrhoea, urticaria and abdominal pain [170]. The International Organisation of Vine and Wine (OIV) has been gradually reducing the maximum advisable levels of total SO<sup>2</sup> in wines [143]. The maximum acceptable limit for total SO2 in grape wine is set between 150 and 300 mg/L depending on the wine type and the level of reducing substances in wine. One of the challenges for scientists and winemakers who are striving to meet today's consumer demands for SO<sup>2</sup> -free high-quality wines is the application of new technologies to replace the use of SO2 [171]. The alternatives to SO<sup>2</sup> investigated so far include treatments with lysozyme, dimethyl dicarbonate (DMDC), the addition of reduced glutathione (up to 20 mg/L) and the addition of commercial oenological tannins [170, 172–176].

## **4. Conclusions**

**3.3. Biogenic amines**

76 Descriptive Food Science

**3.4. Sulphur dioxide**

wine, SO2

Fermentation by-products, biogenic amines, are low molecular nitrogen compounds formed mainly by decarboxylation of amino acids or by amination and transamination of aldehydes and ketones. In food and beverages, they are synthesised by the enzymes from raw material or are generated by decarboxylation of amino acids by microorganisms [161, 162]. Biogenic amines are undesirable in all foods and beverages because if absorbed at too high concentrations, they may cause adverse physiological effects in sensitive humans, especially in the presence of alcohol and acetaldehyde. The most studied biogenic amine is histamine [33, 163]. Biogenic amines, particularly histamine, the most frequently found in wines, often reduce the sensory quality of grape wines. Different microorganisms present in wine can synthesise biogenic amines from their respective amino acid precursors, at any stage of wine production, ageing or storage. Therefore, their presence in wines can serve as an indicator of spoilage and authenticity [164, 165]. Biogenic amines associated with grape wine are putrescine, histamine, tyramine and cadaverine, followed by phenylethylamine, spermidine, spermine, agmatine and tryptamine [162]. Lactic acid bacteria present in wine (e.g. *Lactobacillus*, *Leuconostoc* and *Oenococcus* spp.) are mainly responsible for the occurrence of biogenic amines in wine (and in other fermented products) since they produce enzymes that decarboxylate the respective precursor amino acids [166]. To prevent the problem of a high biogenic amine concentration in wines, the length of the processes of maceration and the contact of wine and lees should be reduced to a minimum, because those are the processes that incorporate amino acids to must or wine. This is hard to achieve when age wines are produced. Another way of solving the problem is to inhibit the growth of indigenous lactic acid bacteria and use the selected *O. oeni* strains unable to produce biogenic amine [167]. Biogenic amines present in fruits also contribute to their level in the wine. It has been reported that fruit cultivation conditions and management technique play essential roles in the accumulation of biogenic amines in fruits, for example, the accumulation of putrescine in grapes as a response to a potassium deficiency in soil [168]. While data are available on the concentrations of biogenic amines in grape wines, rare investigations have been conducted to evaluate their concentrations in fruit wines. Ouyang et al. [169] observed a noticeable difference in the profile of biogenic amines in the wines made from different fruits. A high level of the total biogenic amines (28.11–67.48 mg L−1) was detected in red grape wines, followed by the strawberry wine (14.60 mg L−1) and the raspberry wine (8.75 mg L−1). The total content of BA in the white grape wines ranged from 5.42 to 7.21 mg L−1, while the level of total biogenic amines of other fruit wines was below 2.5 mg L−1. When individual biogenic amines were analysed, the results revealed that spermidine was present in most of the wines, while putrescine was present in all the grape wine samples. However, the levels of putrescine in blueberry, raspberry, schisandrae and strawberry wines were higher than that in the grape wines. Blueberry wine and raspberry wine also contained tyramine.

The most common preservative in wine production used to control both oxidative processes

antioxidant and antimicrobial properties. However, when present in high concentrations in

pain [170]. The International Organisation of Vine and Wine (OIV) has been gradually reducing

can have unfavourable health effects, for example, diarrhoea, urticaria and abdominal

) [39]. This is due to its potent

in wines [143]. The maximum acceptable limit for

and unwanted spontaneous fermentations is sulphur dioxide (SO<sup>2</sup>

the maximum advisable levels of total SO<sup>2</sup>

The role of fruits in the human diet is well established due to various important nutritive and biologically active components intrinsic to fruits. Consequently, fruit wines tend to preserve these components, along with developing new desirable ones in the final product. Fruit wines proved to be a good source of different antioxidants, phytonutrients and minerals, which classify them with the functional food. The production of fruit wines is growing steadily in recent years, probably driven by the demand for new functional products. However, having in mind the fact that most fruits can be used for fruit wines production, it is evident that they comprise a vast and diverse group of fermented beverages. Compared to grape wines, the comprehensive reviews of specific fruit wines are still scarce, as well as the comparison of different fruit wines regarding their production, composition, nutritional, functional and health-related properties. Hopefully, this chapter will contribute to the body of knowledge, as well as to diversification, standardisation and, consequently, commercialisation of quality fruit wine production.

## **Author details**

Darko Velić<sup>1</sup> \*, Daniela Amidžić Klarić<sup>2</sup> , Natalija Velić<sup>1</sup> , Ilija Klarić<sup>3</sup> , Vlatka Petravić Tominac<sup>4</sup> and Ana Mornar2

\*Address all correspondence to: dvelic@ptfos.hr

1 Faculty of Food Technology Osijek, University of Osijek, Osijek, Croatia

2 Faculty of Pharmacy and Biochemistry, University of Zagreb, Zagreb, Croatia

3 Department of Health Ecology, Public Health Brčko DC, Brčko DC, Bosnia and Herzegovina

4 Faculty of Food Technology and Biotechnology, University of Zagreb, Zagreb, Croatia

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[174] Lužar J, Košmerl T, Jamnik P, Korošec M. Wine ageing as alternative to addition SO<sup>2</sup>

[175] Santos MC, Nunes C, Saraiva JA, Coimbra MA. Chemical and physical methodologies for the replacement/reduction of sulfur dioxide use during winemaking: Review of their potentialities and limitations. European Food Research and Technology. 2012;**234**(1):1-12.

[176] Sonni F, Chinnici F, Natali N, Riponi C. Pre-fermentative replacement of sulphur dioxide by lysozyme and oenological tannins: Effect on the formation and evolution of volatile compounds during the bottle storage of white wines. Food Chemistry.

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**Chapter 5**

**Provisional chapter**

**Role of Dietary Carotenoids in Different Etiologies of**

**Role of Dietary Carotenoids in Different Etiologies of** 

Carotenoids are tetraterpenoid organic pigments synthesized by a variety of plants and microorganisms. Dietary carotenoids, taken by animals through food, play an essential role in cell differentiation, morphogenesis, vision, prevention of cancer, atherosclerosis, and age-related macular degeneration in humans due to their potential to suppress oxidative stress. As reactive oxygen species and oxidative damage to biomolecules have been found to be involved in the causation and progression of chronic liver diseases (CLDs), including hepatocellular carcinoma (HCC), which is one of the major causes of morbidity and mortality worldwide. Therefore, dietary antioxidants, which inactivate reactive oxygen species and obstruct oxidative damage, are considered as vital prophylactic strategic molecules. Data from various epidemiological studies and clinical trials strongly validate the observation that adequate carotenoid supplementation may significantly reduce the risk of several liver disorders. This chapter, thus, provides a comprehensive account of dietary carotenoids and includes the recent information with respect to their role in pre-

**Keywords:** β-carotene, lycopene, lutein, β-cryptoxanthin, oxidative stress, chronic liver

There are unambiguous evidences that regular consumption of vegetables and fruits decreases the prevalence of chronic liver disease (CLD) [1, 2]. One of the main reasons of the health organizations to increase the consumption of vegetables and fruits is that these are good sources of carotenoids and other biologically active phytochemicals. Carotenoids are naturally occurring tetraterpenoids and represented by approximately 700 different structural

> © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.80650

**Chronic Liver Diseases**

**Chronic Liver Diseases**

Uzma Latief and Riaz Ahmad

Uzma Latief and Riaz Ahmad

**Abstract**

http://dx.doi.org/10.5772/intechopen.80650

vention of liver diseases.

diseases

**1. Introduction**

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

#### **Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases**

DOI: 10.5772/intechopen.80650

Uzma Latief and Riaz Ahmad Uzma Latief and Riaz Ahmad

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.80650

#### **Abstract**

Carotenoids are tetraterpenoid organic pigments synthesized by a variety of plants and microorganisms. Dietary carotenoids, taken by animals through food, play an essential role in cell differentiation, morphogenesis, vision, prevention of cancer, atherosclerosis, and age-related macular degeneration in humans due to their potential to suppress oxidative stress. As reactive oxygen species and oxidative damage to biomolecules have been found to be involved in the causation and progression of chronic liver diseases (CLDs), including hepatocellular carcinoma (HCC), which is one of the major causes of morbidity and mortality worldwide. Therefore, dietary antioxidants, which inactivate reactive oxygen species and obstruct oxidative damage, are considered as vital prophylactic strategic molecules. Data from various epidemiological studies and clinical trials strongly validate the observation that adequate carotenoid supplementation may significantly reduce the risk of several liver disorders. This chapter, thus, provides a comprehensive account of dietary carotenoids and includes the recent information with respect to their role in prevention of liver diseases.

**Keywords:** β-carotene, lycopene, lutein, β-cryptoxanthin, oxidative stress, chronic liver diseases

#### **1. Introduction**

There are unambiguous evidences that regular consumption of vegetables and fruits decreases the prevalence of chronic liver disease (CLD) [1, 2]. One of the main reasons of the health organizations to increase the consumption of vegetables and fruits is that these are good sources of carotenoids and other biologically active phytochemicals. Carotenoids are naturally occurring tetraterpenoids and represented by approximately 700 different structural

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

variants, but only 50 have been reported to play an important role in human diet [3]. They are synthesized by plants, fungi, algae, and bacteria [4]. They are classified into two groups, carotenes containing only carbon and hydrogen atoms and xanthophylls containing at least one oxygen atom [5]. These carotenoids are further paired into two classes, provitamin A carotenoids (β-carotene and β-cryptoxanthin) and non-provitamin A carotenoids (lycopene and lutein) [6]. In animals and human beings, carotenoids particularly β-carotene, lycopene, lutein, and β-cryptoxanthin play an important role in protection against photooxidative damage by acting as singlet molecular oxygen and peroxy radical scavenger [7, 8]. There are increasing evidences that an alteration of the cellular redox state involving production of reactive oxygen species (ROS) plays a central role in different steps that initiate and regulate the progression of various liver diseases irrespective of the cause. Liver damage caused by reactive oxygen species is induced by alcohol, alteration of lipid, viruses, carbohydrate metabolism, and xenobiotics [9]. In this context, provitamin A activity of these carotenoids [10, 11] has received considerable interest by researchers and health professionals to prevent chronic liver diseases.

The major source of ROS in hepatocytes is the mitochondria. Hepatocytes contain many mitochondria and thus have high production of ROS. An imbalance between the production of ROS and antioxidant defense generates various pathophysiological alterations in the hepatic cells, such as activation of hepatic stellate cells (HSCs), initiation of collagen formation, and beginning of proliferative processes [17–21]. HSCs are responsible for storage of more than 90% of the body's vitamin A content as retinyl esters in normal liver. Chronic liver injuries may cause these cells to undergo activation and lose their capacity to store vitamin A while acquiring pro-inflammatory and proliferative properties, responsible for fibrogenesis [22]. Fibrosis if not treated may lead to end stage of liver damage, that is, liver cancer (**Figure 1**)

Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases

http://dx.doi.org/10.5772/intechopen.80650

95

In this context, dietary carotenoids being an important part of antioxidant defense system and precursor of vitamin A play an important role in the prevention of chronic liver diseases. Nowadays, due emphasis is given on the search of alternative therapeutics or medicines as these have least side effects and are cost effective. In this context, this chapter focuses on the use of dietary carotenoids in chronic liver disease prevention which have been documented

β-Carotene is the major carotenoid present in human diet. It is mainly present in plant sources of human diet such as carrots, pumpkin, spinach, sweet potato, cantaloupe, papaya, and mangoes [25]. It is a potent quencher of singlet oxygen, scavenges peroxy radicals, reduces levels of reactive oxygen species, and augments the investigated antioxidant enzyme activities [26, 27]. It is also a precursor of vitamin A and is reportedly converted to vitamin A by the action of

Various studies have demonstrated the hepatoprotective potential of β-carotene carried out in animal models, cell lines, and human beings. Patel and Sail reported that β-carotene protects the physiological antioxidants against aflatoxin-B1-induced carcinogenesis in albino rats [28]. β-Carotene supplementation increases the levels of vitamin C, glutathione, and glutathione-related enzymes which function as free radical scavenger and, thus, reduces the toxicity of aflatoxin-B1 in rats. In another study, it was found that various antioxidants including β-carotene modulate the hepatotoxicity induced by aflatoxin-B1 [29]. Both HBV and HCV have been found to elevate inflammatory and oxidative conditions in hepatocytes. Moreover, in chronic liver diseases, hepatic stellate cells, which store almost 70–90% of vitamin A, become activated, lose their retinoid content, and produce extracellular matrix, which is responsible for liver fibrosis [30]. Hepatic fibrosis can lead to cirrhosis and in some cases hepatocellular carcinoma (HCC). Through its provitamin A activity and roles in inhibiting reactive oxygen species, β-carotene has been shown to ameliorate the development and progression of HBVand HCV-induced HCC. In a human study, it was found that the oxidative stress increased in patients with chronic hepatitis C (CHC). Various antioxidants such as retinol, α- and γ-tocopherol, β-cryptoxanthin, lycopene, and α- and β-carotene were decreased in serum of CHC patients, and levels in liver tissue seem to reflect serum levels [31]. In Chinese patients,

[15, 23, 24].

extensively in the recent literature.

β-carotene 15,15′-monooxygenase [10].

**2. β-Carotene and chronic liver diseases**

Chronic liver diseases (CLDs) are the major concern throughout the world because of the increasing death rate due to them. CLDs such as alcoholic liver disease (ALD), nonalcoholic fatty liver disease (NAFLD), and viral hepatitis (B and C) progress to fibrosis, cirrhosis, and ultimately hepatocellular carcinoma (HCC) [12, 13]. HCC is the fifth most common type of cancer and third most common cause of cancer mortality throughout the world [14]. Globally, liver cancer is responsible for causing more than 700,000 deaths annually [15]. Oxidative stress resulting from various sources is a major mechanism for hepatic fibrosis and cirrhosis [16].

**Figure 1.** Functions of various dietary carotenoids against chronic liver diseases.

The major source of ROS in hepatocytes is the mitochondria. Hepatocytes contain many mitochondria and thus have high production of ROS. An imbalance between the production of ROS and antioxidant defense generates various pathophysiological alterations in the hepatic cells, such as activation of hepatic stellate cells (HSCs), initiation of collagen formation, and beginning of proliferative processes [17–21]. HSCs are responsible for storage of more than 90% of the body's vitamin A content as retinyl esters in normal liver. Chronic liver injuries may cause these cells to undergo activation and lose their capacity to store vitamin A while acquiring pro-inflammatory and proliferative properties, responsible for fibrogenesis [22]. Fibrosis if not treated may lead to end stage of liver damage, that is, liver cancer (**Figure 1**) [15, 23, 24].

In this context, dietary carotenoids being an important part of antioxidant defense system and precursor of vitamin A play an important role in the prevention of chronic liver diseases. Nowadays, due emphasis is given on the search of alternative therapeutics or medicines as these have least side effects and are cost effective. In this context, this chapter focuses on the use of dietary carotenoids in chronic liver disease prevention which have been documented extensively in the recent literature.

## **2. β-Carotene and chronic liver diseases**

**Figure 1.** Functions of various dietary carotenoids against chronic liver diseases.

chronic liver diseases.

94 Descriptive Food Science

variants, but only 50 have been reported to play an important role in human diet [3]. They are synthesized by plants, fungi, algae, and bacteria [4]. They are classified into two groups, carotenes containing only carbon and hydrogen atoms and xanthophylls containing at least one oxygen atom [5]. These carotenoids are further paired into two classes, provitamin A carotenoids (β-carotene and β-cryptoxanthin) and non-provitamin A carotenoids (lycopene and lutein) [6]. In animals and human beings, carotenoids particularly β-carotene, lycopene, lutein, and β-cryptoxanthin play an important role in protection against photooxidative damage by acting as singlet molecular oxygen and peroxy radical scavenger [7, 8]. There are increasing evidences that an alteration of the cellular redox state involving production of reactive oxygen species (ROS) plays a central role in different steps that initiate and regulate the progression of various liver diseases irrespective of the cause. Liver damage caused by reactive oxygen species is induced by alcohol, alteration of lipid, viruses, carbohydrate metabolism, and xenobiotics [9]. In this context, provitamin A activity of these carotenoids [10, 11] has received considerable interest by researchers and health professionals to prevent

Chronic liver diseases (CLDs) are the major concern throughout the world because of the increasing death rate due to them. CLDs such as alcoholic liver disease (ALD), nonalcoholic fatty liver disease (NAFLD), and viral hepatitis (B and C) progress to fibrosis, cirrhosis, and ultimately hepatocellular carcinoma (HCC) [12, 13]. HCC is the fifth most common type of cancer and third most common cause of cancer mortality throughout the world [14]. Globally, liver cancer is responsible for causing more than 700,000 deaths annually [15]. Oxidative stress resulting from various sources is a major mechanism for hepatic fibrosis and cirrhosis [16].

β-Carotene is the major carotenoid present in human diet. It is mainly present in plant sources of human diet such as carrots, pumpkin, spinach, sweet potato, cantaloupe, papaya, and mangoes [25]. It is a potent quencher of singlet oxygen, scavenges peroxy radicals, reduces levels of reactive oxygen species, and augments the investigated antioxidant enzyme activities [26, 27]. It is also a precursor of vitamin A and is reportedly converted to vitamin A by the action of β-carotene 15,15′-monooxygenase [10].

Various studies have demonstrated the hepatoprotective potential of β-carotene carried out in animal models, cell lines, and human beings. Patel and Sail reported that β-carotene protects the physiological antioxidants against aflatoxin-B1-induced carcinogenesis in albino rats [28]. β-Carotene supplementation increases the levels of vitamin C, glutathione, and glutathione-related enzymes which function as free radical scavenger and, thus, reduces the toxicity of aflatoxin-B1 in rats. In another study, it was found that various antioxidants including β-carotene modulate the hepatotoxicity induced by aflatoxin-B1 [29]. Both HBV and HCV have been found to elevate inflammatory and oxidative conditions in hepatocytes. Moreover, in chronic liver diseases, hepatic stellate cells, which store almost 70–90% of vitamin A, become activated, lose their retinoid content, and produce extracellular matrix, which is responsible for liver fibrosis [30]. Hepatic fibrosis can lead to cirrhosis and in some cases hepatocellular carcinoma (HCC). Through its provitamin A activity and roles in inhibiting reactive oxygen species, β-carotene has been shown to ameliorate the development and progression of HBVand HCV-induced HCC. In a human study, it was found that the oxidative stress increased in patients with chronic hepatitis C (CHC). Various antioxidants such as retinol, α- and γ-tocopherol, β-cryptoxanthin, lycopene, and α- and β-carotene were decreased in serum of CHC patients, and levels in liver tissue seem to reflect serum levels [31]. In Chinese patients, it was observed in Chinese patients that the serum levels of retinoids are low in HBV-induced HCC [32, 33]. In another study, it was shown that a high percentage of patients have vitamin A deficiency in serum in chronic hepatitis C [34, 35]. This condition reflects no response to antiviral therapy, indicating that serum levels of vitamin A could regulate the responsiveness to interferon-based antiviral therapy [35].

with low prevalence of NAFLD [45]. These carotenoids mediate the protective effects against NAFLD through antioxidant mechanism, enhancing gap junction communication, reducing inflammation, and modulation of gene expression. In another human study, it was found that NAFLD has reverse relationship with vitamin A nutritional status in individuals with class III obesity. Retinol and β-carotene serum levels were evaluated as biochemical indicators. The researchers observed low serum retinol and β-carotene serum levels in the patients with

Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases

http://dx.doi.org/10.5772/intechopen.80650

97

In one study, β-carotene and vanadium inhibit the diethylnitrosamine (DEN)-induced hepatic carcinogenesis in rats [47]. It was observed that treatment of β-carotene and vanadium reduced the number and size of the hyperplastic nodules significantly, while the combination treatment proved as an additive effect, decreasing number and size of the hyperplastic nodules from 89 to 22%. Further, it significantly reduced the level of cytosolic glutathione and glutathione-S-transferase (GST) activity and stabilized the aerobic metabolism and hepatic architecture of the cells. In another study involving cell lines, it was found that acyclic retinoid (synthetic analog of retinoids) retards overexpression of Ras/Erk signaling system, thereby declining the progression of HCC [48]. In a human study, it was found that greater intake of retinol, total vitamin A, and carotenes decreases the risk of primary liver cancer at an intake of 1000 μg retinol equivalent (RE)/day or greater from food sources [49]. In another human study in the presence of hepatitis B virus, levels of dietary and serum vitamin A and β-carotene were significantly lower in HCC patients than in the

Lycopene, like other carotenoids, is a natural pigment mainly present in tomato and products of tomato. It is also present in watermelon, apricot, pink guava, pink grapefruit, and papaya [50]. It does not show provitamin A activity since it lacks the β-ionone ring structure which is characteristic in other carotenes that are precursors of vitamin A [50, 51]. Various studies showed that lycopene possesses antioxidant, anticancer, anti-cardiovascular disease, and detoxification abilities in many epidemiological and animal experiments with few side effects

As a dietary phytochemicals, lycopene has been demonstrated to mitigate AFB1-induced adverse effects in vitro and in vivo. The carcinogenicity of aflatoxin B1 (AFB1) in HepG2 cells was prevented by lycopene through decreasing DNA damage and AFB1-N7-guanine (AFB1-N7-Gua) adduct formation [55]. In another study, it was found that lycopene, because of its high antioxidant activity and free radical scavenging capacity, has been shown to be effective against oxidative stress due to aflatoxin. Lycopene blocks phase 1 metabolic enzymes of AFB such as 3A4, 2A6, and 1A2 [56]. In another study, lycopene relieves AFB1 induced liver injury through enhancing hepatic antioxidation and detoxification potential with Nrf2 activation [57]. Lycopene, a nutritional antioxidant, has also been shown for its hepatoprotective potential in D-galactosamine/lipopolysaccharide (D-GalN/LPS)-induced

NAFLD [46].

control subjects [32].

[52, 53, 54].

**3. Lycopene and chronic liver diseases**

The generation of very high levels of oxidative stress during the metabolism of alcohol may exceed the antioxidant defense ability of the body and cause the development of liver dysfunction. It has been reported that β-carotene exerts protective effect on chronic ethanol-fed rats [36]. It was shown that ethanol treatment causes increase in oxidative stress which could stimulate apoptosis in the liver, thus leading to liver injury. However, the lower dose of β-carotene supplementation (0.52 mg/kg BW/day), which acted as an antioxidant, decreased the ROS level by downregulating lipid peroxidation and CYP2E1 expression. Moreover, it prevented ethanol-induced liver damage by impeding hepatic apoptosis via inhibiting caspase-9 and caspase-3 expressions and increasing Bcl-xL expression in the liver. In addition, the higher dose of β-carotene supplementation (6.2 mg/kg BW/day) possibly halted ethanol-induced liver damage through inhibiting TNF-α secretion and lipid peroxidation in ethanol-fed rats. Literature revealed that β-carotene supplementation can prevent liver damage in rats with chronic alcohol consumption [37, 38]. β-carotene supplement is known to attenuate ethanol-induced liver damage, decreased oxidative stress and increased GSH concentrations in erythrocytes and the liver. β-Carotene may act as an antioxidant, scavenging lipophilic radicals produced by ethanol within the membranes [39]. In a human study, it was found that in alcoholic patients with liver damage, the plasma β-carotene level was found to be lowered than in control subjects [40]. ALD is also associated with depleted levels of hepatic vitamin A [41]. As β-carotene is the precursor of vitamin A, thus, the supplementation of this carotenoid tends to regain the hepatic vitamin A content, thus facilitating attenuation of the disease.

In a study, it was found that dietary intake of apricot reduced the risks of hepatic steatosis and damage induced by CCl4 in Wistar rats [42]. Apricot is believed to have high content of carotenoids, especially β-carotene. Markers of oxidative stress such as malondialdehyde, total GSH levels, catalase, SOD, and GSH peroxidase activities were significantly altered by CCl<sup>4</sup> indicating increased oxidative stress. Hepatic damage and steatosis imposed by high concentration of ROS were ameliorated by β-carotene-rich apricot intake. Consumption of tomato juice which contains lycopene and β-carotene as the major components reduces plasmatic triglycerides, steatosis, and very low-density lipoproteins. Also, it elevates lipid metabolism by inducing the overexpression of genes involved in more efficient fatty acid oxidation in rats [43]. It is also shown that Campari tomato, which contains more β-carotene and lycopene than regular tomato, ameliorated diet-induced obesity, dyslipidemia, and hepatosteatosis via downregulation of gene expression related to lipogenesis in the zebra fish model. It decreased sterol regulatory element-binding transcription factor 1 (SREBF1) mRNA by increasing the forkhead box O1 (foxo1) gene expression. This may be due to high percentage of β-carotene in this strain of tomato which is responsible for downregulating the expression of SREBF1 [44]. In Chinese population greater levels of carotenoids such as β-carotene in serum have been correlated with low prevalence of NAFLD [45]. These carotenoids mediate the protective effects against NAFLD through antioxidant mechanism, enhancing gap junction communication, reducing inflammation, and modulation of gene expression. In another human study, it was found that NAFLD has reverse relationship with vitamin A nutritional status in individuals with class III obesity. Retinol and β-carotene serum levels were evaluated as biochemical indicators. The researchers observed low serum retinol and β-carotene serum levels in the patients with NAFLD [46].

In one study, β-carotene and vanadium inhibit the diethylnitrosamine (DEN)-induced hepatic carcinogenesis in rats [47]. It was observed that treatment of β-carotene and vanadium reduced the number and size of the hyperplastic nodules significantly, while the combination treatment proved as an additive effect, decreasing number and size of the hyperplastic nodules from 89 to 22%. Further, it significantly reduced the level of cytosolic glutathione and glutathione-S-transferase (GST) activity and stabilized the aerobic metabolism and hepatic architecture of the cells. In another study involving cell lines, it was found that acyclic retinoid (synthetic analog of retinoids) retards overexpression of Ras/Erk signaling system, thereby declining the progression of HCC [48]. In a human study, it was found that greater intake of retinol, total vitamin A, and carotenes decreases the risk of primary liver cancer at an intake of 1000 μg retinol equivalent (RE)/day or greater from food sources [49]. In another human study in the presence of hepatitis B virus, levels of dietary and serum vitamin A and β-carotene were significantly lower in HCC patients than in the control subjects [32].

## **3. Lycopene and chronic liver diseases**

it was observed in Chinese patients that the serum levels of retinoids are low in HBV-induced HCC [32, 33]. In another study, it was shown that a high percentage of patients have vitamin A deficiency in serum in chronic hepatitis C [34, 35]. This condition reflects no response to antiviral therapy, indicating that serum levels of vitamin A could regulate the responsiveness

The generation of very high levels of oxidative stress during the metabolism of alcohol may exceed the antioxidant defense ability of the body and cause the development of liver dysfunction. It has been reported that β-carotene exerts protective effect on chronic ethanol-fed rats [36]. It was shown that ethanol treatment causes increase in oxidative stress which could stimulate apoptosis in the liver, thus leading to liver injury. However, the lower dose of β-carotene supplementation (0.52 mg/kg BW/day), which acted as an antioxidant, decreased the ROS level by downregulating lipid peroxidation and CYP2E1 expression. Moreover, it prevented ethanol-induced liver damage by impeding hepatic apoptosis via inhibiting caspase-9 and caspase-3 expressions and increasing Bcl-xL expression in the liver. In addition, the higher dose of β-carotene supplementation (6.2 mg/kg BW/day) possibly halted ethanol-induced liver damage through inhibiting TNF-α secretion and lipid peroxidation in ethanol-fed rats. Literature revealed that β-carotene supplementation can prevent liver damage in rats with chronic alcohol consumption [37, 38]. β-carotene supplement is known to attenuate ethanol-induced liver damage, decreased oxidative stress and increased GSH concentrations in erythrocytes and the liver. β-Carotene may act as an antioxidant, scavenging lipophilic radicals produced by ethanol within the membranes [39]. In a human study, it was found that in alcoholic patients with liver damage, the plasma β-carotene level was found to be lowered than in control subjects [40]. ALD is also associated with depleted levels of hepatic vitamin A [41]. As β-carotene is the precursor of vitamin A, thus, the supplementation of this carotenoid tends to regain the hepatic vitamin A content, thus facilitating

In a study, it was found that dietary intake of apricot reduced the risks of hepatic steatosis and

enoids, especially β-carotene. Markers of oxidative stress such as malondialdehyde, total GSH

ing increased oxidative stress. Hepatic damage and steatosis imposed by high concentration of ROS were ameliorated by β-carotene-rich apricot intake. Consumption of tomato juice which contains lycopene and β-carotene as the major components reduces plasmatic triglycerides, steatosis, and very low-density lipoproteins. Also, it elevates lipid metabolism by inducing the overexpression of genes involved in more efficient fatty acid oxidation in rats [43]. It is also shown that Campari tomato, which contains more β-carotene and lycopene than regular tomato, ameliorated diet-induced obesity, dyslipidemia, and hepatosteatosis via downregulation of gene expression related to lipogenesis in the zebra fish model. It decreased sterol regulatory element-binding transcription factor 1 (SREBF1) mRNA by increasing the forkhead box O1 (foxo1) gene expression. This may be due to high percentage of β-carotene in this strain of tomato which is responsible for downregulating the expression of SREBF1 [44]. In Chinese population greater levels of carotenoids such as β-carotene in serum have been correlated

levels, catalase, SOD, and GSH peroxidase activities were significantly altered by CCl<sup>4</sup>

in Wistar rats [42]. Apricot is believed to have high content of carot-

indicat-

to interferon-based antiviral therapy [35].

96 Descriptive Food Science

attenuation of the disease.

damage induced by CCl4

Lycopene, like other carotenoids, is a natural pigment mainly present in tomato and products of tomato. It is also present in watermelon, apricot, pink guava, pink grapefruit, and papaya [50]. It does not show provitamin A activity since it lacks the β-ionone ring structure which is characteristic in other carotenes that are precursors of vitamin A [50, 51]. Various studies showed that lycopene possesses antioxidant, anticancer, anti-cardiovascular disease, and detoxification abilities in many epidemiological and animal experiments with few side effects [52, 53, 54].

As a dietary phytochemicals, lycopene has been demonstrated to mitigate AFB1-induced adverse effects in vitro and in vivo. The carcinogenicity of aflatoxin B1 (AFB1) in HepG2 cells was prevented by lycopene through decreasing DNA damage and AFB1-N7-guanine (AFB1-N7-Gua) adduct formation [55]. In another study, it was found that lycopene, because of its high antioxidant activity and free radical scavenging capacity, has been shown to be effective against oxidative stress due to aflatoxin. Lycopene blocks phase 1 metabolic enzymes of AFB such as 3A4, 2A6, and 1A2 [56]. In another study, lycopene relieves AFB1 induced liver injury through enhancing hepatic antioxidation and detoxification potential with Nrf2 activation [57]. Lycopene, a nutritional antioxidant, has also been shown for its hepatoprotective potential in D-galactosamine/lipopolysaccharide (D-GalN/LPS)-induced hepatitis in rats [58, 59]. It is able to affect the lipoprotein metabolism by restoring the altered levels of lipid-metabolizing enzymes and stabilizing the arrangement of lipoprotein levels during experimentally induced hepatitis. Another study demonstrated that the regular use of our carotenoid-based functional food minimizes the severity of ribavirin-induced anemia in patients with CHC and improves tolerance to the full dose of antiviral therapy [60]. It was found that a mixture of various carotenoids particularly lycopene seems to be promising for the prevention of liver cancer in hepatitis virus-infected patients with cirrhosis [61, 62].

Lutein decreases inflammation and oxidative stress in the liver and eyes of guinea pigs fed with hypercholesterolemic diet [78]. This carotenoid could prevent degenerative conditions of the liver by decreasing the free cholesterol pool and attenuating lipid peroxidation and proinflammatory cytokine production. Further, attenuated inflammatory state in the liver could be explained by decreased NF-κB DNA-binding activity. Another study demonstrated that lutein possesses ameliorative effect against NAFLD [79]. It suggests that lutein supplementation could protect against hepatic lipid accumulation and insulin resistance induced by highfat diet, possibly via the activation of the expression of sirtuin 1 (SIRT1) and, subsequently, peroxisome proliferator activated receptor (PPAR)-α and other key factors in insulin signaling [80]. Sirtuin 1 is reported to have therapeutic potential in NAFLD and play a key role in insulin sensitivity. SIRT1 regulates the expression of PPAR-α, a key factor in the regulation of lipid metabolism [81, 82]. Lutein has also been found to have anticarcinogenetic effects against NDEA-induced HCC in rats [83]. Inhibition of carcinogenesis by this carotenoid could be because of the combined effect of its antioxidant activity along with the inhibition of cytochrome P450 enzymes, inducing detoxifying enzymes such as glutathione-S-transferase and

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99

β-Cryptoxanthin is an oxygenated carotenoid usually present in squash, pepper, papaya, sweet pickles, carrots, and orange juice [84]. It is a xanthophyll carotenoid specifically found in the Satsuma mandarin (*Citrus unshiu* Marc.). Similar to other carotenoids, β-cryptoxanthin has an antioxidant activity [85, 86]. β-Cryptoxanthin is readily absorbed and relatively abundant in human plasma, together with β-carotene, lycopene, lutein, and zeaxanthin [87].

It has been shown that β-cryptoxanthin ameliorates diet-induced nonalcoholic steatohepatitis by repressing inflammatory gene expression in mice [88]. β-Cryptoxanthin suppressed the expression of lipopolysaccharide (LPS)-inducible and TNFα-inducible genes in NASH.Elevated levels of the oxidative stress marker thiobarbituric acid-reactive substances (TBARS) were lowered by β-cryptoxanthin in NASH. Thus, it represses inflammation and the resulting fibrosis probably by primarily repressing the increase and activation of macrophages and other immune cells. Reducing reactive oxygen species is likely to be a major mechanism of inflammation and injury suppression in the livers of mice with NASH. Another study revealed that β-cryptoxanthin reversed steatosis, inflammation, and fibrosis progression in preexisting NASH in mice [89]. Thus, β-cryptoxanthin prevents and reverses insulin resistance and steatohepatitis through decreasing activation of macrophages or Kupffer cells in a lipotoxic model of NASH. It was found that plasma levels of carotenoids such as β-carotene, lycopene, lutein, and β-cryptoxanthin were decreased in patients with NASH [90]. This study suggests that antioxidant supplementation may be a rational option for the treatment of NASH. In patients of NAFLD, it was found that β-cryptoxanthin treatment inhibits its progression [91]. β-Cryptoxanthin supplementation is very effective in raising antioxidant and anti-inflamma-

UDP glucuronyl transferase.

**5. β-Cryptoxanthin and chronic liver diseases**

tion activities in patients of NAFLD (**Table 1**).

It is found that tomato powder which is rich in lycopene acts as a novel candidate for prevention against alcohol-related hepatic injury in rodents [63]. It provides protection against alcohol-induced liver injury by suppressing CYP450 2E1 induction. It has been reported that lycopene prevents nonalcoholic steatohepatitis induced by high-fat diet [64, 65, 66]. It can reduce high-fat diet-induced steatohepatitis by reducing oxidative stress to the cells. It has been also shown that incorporation of lycopene in balanced diet prevents NAFLD [67, 68]. Lycopene may be a useful functional compound for treating NAFLD by regulating hepatic lipid metabolism [69]. Lycopene inhibits the downregulation of miR-21, which led to the downregulation of fatty acid-binding protein 7 (FABP7) at both the transcriptional and translational levels, thus inhibiting hepatic steatosis induced by high-fat diet.

Lycopene also showed beneficial effects against hepatocellular carcinoma. It provides protection against HCC by modulation of cellular proliferation, glycolysis, and ultrastructure of hepatic cells [70]. Lycopene supplementation also prevents high-fat diet-induced HCC incidence in mice. It suppressed oncogenic signals, including methionine mRNA, β-catenin protein, and mammalian target of rapamycin (mTOR) complex 1 activation. These results provide novel experimental evidence that dietary lycopene and its metabolites can be used to prevent liver cancer and reduce cancer risk in patients with NAFLD [71].

## **4. Lutein and chronic liver diseases**

Lutein is a non-provitamin A carotenoid that belongs to the oxycarotenoid family. It is present in dark green leafy vegetables, such as kale and spinach and eggs [72, 73]. It is commercially prepared from the marigold flower (*Tagetes erecta* L.) in which it occurs at 1.5–1.8%. It directly quenches free radicals, especially singlet oxygen species [74].

Lutein possesses an antiviral activity against hepatitis B. It exerts its antivirus effects through inhibition of hepatitis B virus transcription [75]. Because of its strong antioxidant potential, lutein has been shown to provide protection against ethanol-induced hepatic damage [76]. It increases levels of antioxidant enzymes, like superoxide dismutase, catalase, glutathione peroxidase, and glutathione, and decreases levels of hydroxyproline. Another study demonstrated that lutein attenuates alcohol-induced liver damage in rats by regulating inflammation and oxidative stress [77]. Its supplementation downregulated inflammatory proteins and cytokines with collateral upregulation of Nrf2 levels and antioxidant enzymatic activities. Lutein decreases inflammation and oxidative stress in the liver and eyes of guinea pigs fed with hypercholesterolemic diet [78]. This carotenoid could prevent degenerative conditions of the liver by decreasing the free cholesterol pool and attenuating lipid peroxidation and proinflammatory cytokine production. Further, attenuated inflammatory state in the liver could be explained by decreased NF-κB DNA-binding activity. Another study demonstrated that lutein possesses ameliorative effect against NAFLD [79]. It suggests that lutein supplementation could protect against hepatic lipid accumulation and insulin resistance induced by highfat diet, possibly via the activation of the expression of sirtuin 1 (SIRT1) and, subsequently, peroxisome proliferator activated receptor (PPAR)-α and other key factors in insulin signaling [80]. Sirtuin 1 is reported to have therapeutic potential in NAFLD and play a key role in insulin sensitivity. SIRT1 regulates the expression of PPAR-α, a key factor in the regulation of lipid metabolism [81, 82]. Lutein has also been found to have anticarcinogenetic effects against NDEA-induced HCC in rats [83]. Inhibition of carcinogenesis by this carotenoid could be because of the combined effect of its antioxidant activity along with the inhibition of cytochrome P450 enzymes, inducing detoxifying enzymes such as glutathione-S-transferase and UDP glucuronyl transferase.

## **5. β-Cryptoxanthin and chronic liver diseases**

hepatitis in rats [58, 59]. It is able to affect the lipoprotein metabolism by restoring the altered levels of lipid-metabolizing enzymes and stabilizing the arrangement of lipoprotein levels during experimentally induced hepatitis. Another study demonstrated that the regular use of our carotenoid-based functional food minimizes the severity of ribavirin-induced anemia in patients with CHC and improves tolerance to the full dose of antiviral therapy [60]. It was found that a mixture of various carotenoids particularly lycopene seems to be promising for the prevention of liver cancer in hepatitis virus-infected patients with cir-

It is found that tomato powder which is rich in lycopene acts as a novel candidate for prevention against alcohol-related hepatic injury in rodents [63]. It provides protection against alcohol-induced liver injury by suppressing CYP450 2E1 induction. It has been reported that lycopene prevents nonalcoholic steatohepatitis induced by high-fat diet [64, 65, 66]. It can reduce high-fat diet-induced steatohepatitis by reducing oxidative stress to the cells. It has been also shown that incorporation of lycopene in balanced diet prevents NAFLD [67, 68]. Lycopene may be a useful functional compound for treating NAFLD by regulating hepatic lipid metabolism [69]. Lycopene inhibits the downregulation of miR-21, which led to the downregulation of fatty acid-binding protein 7 (FABP7) at both the transcriptional and trans-

Lycopene also showed beneficial effects against hepatocellular carcinoma. It provides protection against HCC by modulation of cellular proliferation, glycolysis, and ultrastructure of hepatic cells [70]. Lycopene supplementation also prevents high-fat diet-induced HCC incidence in mice. It suppressed oncogenic signals, including methionine mRNA, β-catenin protein, and mammalian target of rapamycin (mTOR) complex 1 activation. These results provide novel experimental evidence that dietary lycopene and its metabolites can be used to

Lutein is a non-provitamin A carotenoid that belongs to the oxycarotenoid family. It is present in dark green leafy vegetables, such as kale and spinach and eggs [72, 73]. It is commercially prepared from the marigold flower (*Tagetes erecta* L.) in which it occurs at 1.5–1.8%. It directly

Lutein possesses an antiviral activity against hepatitis B. It exerts its antivirus effects through inhibition of hepatitis B virus transcription [75]. Because of its strong antioxidant potential, lutein has been shown to provide protection against ethanol-induced hepatic damage [76]. It increases levels of antioxidant enzymes, like superoxide dismutase, catalase, glutathione peroxidase, and glutathione, and decreases levels of hydroxyproline. Another study demonstrated that lutein attenuates alcohol-induced liver damage in rats by regulating inflammation and oxidative stress [77]. Its supplementation downregulated inflammatory proteins and cytokines with collateral upregulation of Nrf2 levels and antioxidant enzymatic activities.

lational levels, thus inhibiting hepatic steatosis induced by high-fat diet.

prevent liver cancer and reduce cancer risk in patients with NAFLD [71].

quenches free radicals, especially singlet oxygen species [74].

**4. Lutein and chronic liver diseases**

rhosis [61, 62].

98 Descriptive Food Science

β-Cryptoxanthin is an oxygenated carotenoid usually present in squash, pepper, papaya, sweet pickles, carrots, and orange juice [84]. It is a xanthophyll carotenoid specifically found in the Satsuma mandarin (*Citrus unshiu* Marc.). Similar to other carotenoids, β-cryptoxanthin has an antioxidant activity [85, 86]. β-Cryptoxanthin is readily absorbed and relatively abundant in human plasma, together with β-carotene, lycopene, lutein, and zeaxanthin [87].

It has been shown that β-cryptoxanthin ameliorates diet-induced nonalcoholic steatohepatitis by repressing inflammatory gene expression in mice [88]. β-Cryptoxanthin suppressed the expression of lipopolysaccharide (LPS)-inducible and TNFα-inducible genes in NASH.Elevated levels of the oxidative stress marker thiobarbituric acid-reactive substances (TBARS) were lowered by β-cryptoxanthin in NASH. Thus, it represses inflammation and the resulting fibrosis probably by primarily repressing the increase and activation of macrophages and other immune cells. Reducing reactive oxygen species is likely to be a major mechanism of inflammation and injury suppression in the livers of mice with NASH. Another study revealed that β-cryptoxanthin reversed steatosis, inflammation, and fibrosis progression in preexisting NASH in mice [89]. Thus, β-cryptoxanthin prevents and reverses insulin resistance and steatohepatitis through decreasing activation of macrophages or Kupffer cells in a lipotoxic model of NASH. It was found that plasma levels of carotenoids such as β-carotene, lycopene, lutein, and β-cryptoxanthin were decreased in patients with NASH [90]. This study suggests that antioxidant supplementation may be a rational option for the treatment of NASH. In patients of NAFLD, it was found that β-cryptoxanthin treatment inhibits its progression [91]. β-Cryptoxanthin supplementation is very effective in raising antioxidant and anti-inflammation activities in patients of NAFLD (**Table 1**).


**S. No. Carotenoid investigated Object of study Results References**

lowered serum malondialdehyde and tumor necrosis factor (TNF-α) levels and elevated liver GSH level

Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases

in liver weight, serum low-density lipoproteins, total hepatic cholesterol, and activity of hepatic SOD, catalase,

downregulated TNF-α and CYP2E1 expression and decreased infiltration

NAFLD by alleviating amino acid depletion, recovery of the redox balance in liver, and incrementing

proliferation-associated genes (PCNA, cyclin D1, and p21) and glycolytic

Lutein inhibited the activity of HBV

glutathione, and hepatic hydroxyproline content were restored by lutein

COX-2, and iNOS were downregulated along with the upregulation of Nrf2 levels and activities of antioxidant

serum and hepatic cholesterol and triglyceride. It also increased the expression of key factors involved in

hepatic insulin signaling

glucuronyl transferase

P450 enzymes and increases detoxifying enzymes such as glutathione-S-transferase and UDP

TBARS and suppressed the expression of lipopolysaccharide and TNF-α inducible gene. It further suppressed the activation of macrophages, T helper, and cytotoxic cells

and glutathione peroxidase

of liver fats

L-carnitine levels

cancer in NAFLD

full-length promoter

enzymes

enzymes

[64]

101

http://dx.doi.org/10.5772/intechopen.80650

[68]

[67]

[65]

[70]

[71]

[75]

[76]

[77]

[79]

[83]

[88]

14 Lycopene Rat liver Supplementation with lycopene

16 Lycopene Rat liver Lycopene supplementation

15 Lycopene Rat liver Lycopene treatment reverted changes

17 Lycopene Rat liver Lycopene provides protection against

18 Lycopene Mouse liver It decreases the expression of cell

19 Lycopene Mouse liver Lycopene and its metabolites reduce

21 Lutein Rat liver SOD, catalase, glutathione peroxidase,

22 Lutein Rat liver Inflammatory proteins such as NF-κB,

23 Lutein Rat liver Lutein administration decreased

24 Lutein Rat liver Lutein treatment inhibits cytochrome

25 β-Cryptoxanthin Mouse liver β-Cryptoxanthin reduced the levels of

hepatoblastoma cells

20 Lutein Human


**S. No. Carotenoid investigated Object of study Results References**

glutathione peroxidase, glutathione-S-transferase, catalase, and vitamin C

of hepatic glutathione, RNA, and serum protein thiols relative to control

CYP2E1 expression and lipid peroxidation. β-Carotene also inhibited hepatic apoptosis via inhibiting caspase-3 and caspase-9 and increasing

Bcl-xL expression in the liver

in serum and liver tissue

activities were restored

Human serum Serum carotenoid levels were inversely

Human liver Dietary consumption of retinol,

cancer risk

Rat liver Carotenoids exert protective effect

Increased oxidative stress is present in patients with chronic hepatitis C. Antioxidants were severely depleted

lipid peroxidation in hepatic tissues

levels and catalase, superoxide dismutase, and glutathione peroxidase

associated with the risk of NAFLD

carotenes, and total vitamin A decreases the risk of primary liver

against aflatoxin B1-induced liver preneoplastic foci and DNA damage

cholesterol, triglycerides, free fatty acids, and phospholipids in serum and

density lipoproteins, decrease in high-density lipoproteins, and lipid-

peroxisome proliferator-activated receptor-α, inflammatory gene expression, and reticulum stress markers were restored

detoxification potential

liver were restored

metabolizing enzymes

[28]

[29]

[36]

[31]

[38]

[42]

[45]

[49]

[55]

[57]

[58]

[59]

[63]

were elevated

Rat liver β-Carotene intake restored the levels

animals

1 β-Carotene Rat liver Liver antioxidant enzymes such as

3 β-Carotene Rat liver Oxidative stress is decreased by

Human liver and

5 β-Carotene Rat liver β-Carotene prevented ethanol-induced

6 β-Carotene Rat liver Malondialdehyde, total glutathione

10 Lycopene Rat liver Enhanced hepatic antioxidant and

12 Lycopene Rat liver It restored the increase in very-low-

13 Lycopene Rat and mouse liver Hepatic CYP2E1 protein levels,

11 Lycopene Rat liver Liver function test enzymes,

serum

2 β-Carotene, vitamin E, selenium, silymarin, and

100 Descriptive Food Science

coenzyme Q10

4 Retinol, α- and

γ-tocopherol, lutein, β-cryptoxanthin, lycopene, and α- and β-carotene

7 α- and β-Carotene, lutein, and zeaxanthin

8 Dietary carotenes and vitamin A

9 β-Carotene, β-apo-8′-

carotenal, canthaxanthin, astaxanthin, and lycopene


**Table 1.** Effects of carotenoids on chronic liver diseases in cell line, human, and animal models.

## **6. Other carotenoids and chronic liver diseases**

Other carotenoids such as α-carotene, fucoxanthin, and zeaxanthin also show promising effects against chronic liver injury. It has been found that α-carotene has an inhibitory effect on spontaneous liver carcinogenesis in male mice [92]. α-Carotene significantly decreases the mean number of hepatomas per mouse. In another study, it was found that fucoxanthin promoted weight loss, reduced body and liver fat content, and improved liver function tests in obese nondiabetic women [93]. Zeaxanthin also shows protective effects against nonalcoholic steatohepatitis [94]. It significantly prevented NASH progression by decreasing oxidative stress and liver fibrosis. It has been found also that zeaxanthin showed therapeutic effects against alcoholic liver diseases [95]. Zeaxanthin show protective effects through the lower expression level of cytochrome P450 2E1 (CYP2E1), diminished activity of nuclear factor kappa B (NF-κB) through the restoration of its inhibitor kappa B alpha (IκBα), and the modulation of MAPK pathways including p38 MAPK, JNK, and ERK.

## **7. Conclusions and future directions**

Various epidemiological studies investigate the effects of dietary carotenoids on various markers of oxidative stress and inflammation indicating their preventive role in chronic liver disease prevention. Oxidative stress is clearly associated with the etiology of chronic liver diseases. Thus, the use of carotenoid-rich fruits and vegetables should be the part of diet (**Tables 2** and **3**). Although several pathways related to inhibition of oxidative stress by carotenoids have been uncovered, many aspects remain poorly understood and warrant further research. As many carotenoids have been found to have provitamin A activity, the mechanism related to their regaining of vitamin A content by hepatic stellate cells and restoring their normal functions should be properly understood (**Figure 2**).

There is further need of long-term controlled trials in normal and diseased groups to study the dose response of each dietary carotenoid. Future research areas may include their bioavailability, metabolism, safety, and mechanism of action. Studies on the type of carotenoid and its metabolites which may act as a suitable regulator to alter pathways related to oxidative

stress, inflammation, and carcinogenicity shall also be among the priority areas. However, studying the molecular targets of these dietary carotenoids cannot be ignored and be given

**S. no. Carotenoid Recommended daily dose (mg) References** β-Carotene 7 [98] Lycopene 35 [99] Lutein 1–4 [100] β-Cryptoxanthin 3 [91]

**Dietary sources β-Carotene Lycopene Lutein β-Cryptoxanthin**

Role of Dietary Carotenoids in Different Etiologies of Chronic Liver Diseases

http://dx.doi.org/10.5772/intechopen.80650

103

Apricot (dried) 17.6\$ 0.9\$ — — Carrot (raw) 7.9\$ — — — Spinach (raw) 4.1\$ — 11.9\$ — Kale 4.7\$ — 15.8\$ — Tomato juice — 8.6\$ — — Avocado (raw) 0.053# — — 0.036# Grape fruit, pink (raw) — 3.4\$ — — Guava (raw) — 5.4\$ — — Corn, sweet (cooked) — — 1.8\$ — Papaya (raw) 0.276# — — 0.761# Orange (raw) 0.051# — 0.187# 0.122# Sweet potato (raw) 8.8\$ — — — Lettuce (raw) 1.27# — 2.6# — Watermelon (raw) 0.295# 4.8# 0.017# 0.105# Cabbage (raw) 0.065# — 0.310# — Broccoli (raw) 2.4# — 0.78# — Brussels sprouts (raw) 0.45# — 1.5# — Peas, green (raw) 0.48# — — — Tangerine (raw) 0.071# — 0.243# 0.485# Pepper, sweet red (raw) 2.379# — — 2.205#

full consideration.

#

\$

Derived from Arscott [96].

Derived from Johnson [97].

**Table 2.** Dietary sources of major carotenoids (mg/100 g).

**Table 3.** Ingestion levels of dietary carotenoids to prevent liver diseases.


\$ Derived from Johnson [97].

**6. Other carotenoids and chronic liver diseases**

102 Descriptive Food Science

lation of MAPK pathways including p38 MAPK, JNK, and ERK.

normal functions should be properly understood (**Figure 2**).

**7. Conclusions and future directions**

Other carotenoids such as α-carotene, fucoxanthin, and zeaxanthin also show promising effects against chronic liver injury. It has been found that α-carotene has an inhibitory effect on spontaneous liver carcinogenesis in male mice [92]. α-Carotene significantly decreases the mean number of hepatomas per mouse. In another study, it was found that fucoxanthin promoted weight loss, reduced body and liver fat content, and improved liver function tests in obese nondiabetic women [93]. Zeaxanthin also shows protective effects against nonalcoholic steatohepatitis [94]. It significantly prevented NASH progression by decreasing oxidative stress and liver fibrosis. It has been found also that zeaxanthin showed therapeutic effects against alcoholic liver diseases [95]. Zeaxanthin show protective effects through the lower expression level of cytochrome P450 2E1 (CYP2E1), diminished activity of nuclear factor kappa B (NF-κB) through the restoration of its inhibitor kappa B alpha (IκBα), and the modu-

**S. No. Carotenoid investigated Object of study Results References**

macrophage content

NAFLD patients

and anti-inflammatory activities in

and improved liver function tests

[89]

[91]

[93]

26 β-Cryptoxanthin Mouse liver β-Cryptoxanthin reduced total hepatic

27 β-Cryptoxanthin Human liver β-Cryptoxanthin induced antioxidant

28 Fucoxanthin Human liver It reduced liver and body fat content

**Table 1.** Effects of carotenoids on chronic liver diseases in cell line, human, and animal models.

Various epidemiological studies investigate the effects of dietary carotenoids on various markers of oxidative stress and inflammation indicating their preventive role in chronic liver disease prevention. Oxidative stress is clearly associated with the etiology of chronic liver diseases. Thus, the use of carotenoid-rich fruits and vegetables should be the part of diet (**Tables 2** and **3**). Although several pathways related to inhibition of oxidative stress by carotenoids have been uncovered, many aspects remain poorly understood and warrant further research. As many carotenoids have been found to have provitamin A activity, the mechanism related to their regaining of vitamin A content by hepatic stellate cells and restoring their

There is further need of long-term controlled trials in normal and diseased groups to study the dose response of each dietary carotenoid. Future research areas may include their bioavailability, metabolism, safety, and mechanism of action. Studies on the type of carotenoid and its metabolites which may act as a suitable regulator to alter pathways related to oxidative **Table 2.** Dietary sources of major carotenoids (mg/100 g).


**Table 3.** Ingestion levels of dietary carotenoids to prevent liver diseases.

stress, inflammation, and carcinogenicity shall also be among the priority areas. However, studying the molecular targets of these dietary carotenoids cannot be ignored and be given full consideration.

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2003.12.006

(01)00192-1

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**Figure 2.** Hypothesis of progression of chronic liver diseases and their prevention by dietary carotenoids.

## **Acknowledgements**

The authors sincerely thank the Chairman of the Department of Zoology for necessary facilities. UL acknowledges the financial assistance from UGC-Maulana Azad National Fellowship.

## **Conflict of interest**

The authors declare no potential conflict of interest and are responsible for the writing and content of the chapter.

## **Author details**

Uzma Latief and Riaz Ahmad\*

\*Address all correspondence to: ahmadriaz2013@gmail.com

Section of Genetics, Department of Zoology, Aligarh Muslim University, Aligarh, UP, India

## **References**

**Figure 2.** Hypothesis of progression of chronic liver diseases and their prevention by dietary carotenoids.

The authors sincerely thank the Chairman of the Department of Zoology for necessary facilities. UL acknowledges the financial assistance from UGC-Maulana Azad National Fellowship.

The authors declare no potential conflict of interest and are responsible for the writing and

Section of Genetics, Department of Zoology, Aligarh Muslim University, Aligarh, UP, India

**Acknowledgements**

104 Descriptive Food Science

**Conflict of interest**

content of the chapter.

Uzma Latief and Riaz Ahmad\*

\*Address all correspondence to: ahmadriaz2013@gmail.com

**Author details**


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**Section 2**

**Effects of Technologies on Food Quality**


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10718953

**Chapter 6**

**Provisional chapter**

**Quality of Reduced-Fat Dairy Coffee Creamer: Affected**

This work aims to investigate the effects of inulin (0, 2.5, 5 and 7.5%, w/w) and maltodextrin (0, 15, 20 and 25%, w/w) as wall materials and fat replacers and drying techniques (i.e. spray drying and fluidized-bed drying) on physicochemical properties of regular and instant reduced-fat dairy creamers. The regular reduced-fat dairy creamer was produced by one-stage drying (i.e. spray drying), while the instant reduced-fat dairy creamer was produced by two-stage drying (i.e. spray drying followed by fluidized-bed drying). In this study, control (0% inulin and 0% maltodextrin) and two commercial regular and instant coffee creamers (A and B) were also considered for comparison purposes. The results showed that the regular creamer containing 25% maltodextrin and 7.5% inulin had the largest particle size, highest viscosity and most desirable wettability among all formulated regular creamers. The yield of reduced-fat coffee creamer was significantly increased from 43.55 to 94.60% by increasing the amount of fat replacers to the maximum level (25% maltodextrin and 7.5% inulin). The current study revealed that the application of fluidized-bed drying for agglomeration led to significantly improve the wettability and instant properties of the instant creamer. In this study, the formulated instant creamer containing 25% maltodextrin and 7.5% inulin was the most desirable product as

**Keywords:** reduced-fat dairy creamer, inulin, maltodextrin, spray drying, fluidized-bed

Coffee is one of the most vastly consumed beverages. Coffee drink is usually consumed in black or white form, depending on the taste of the consumer. Coffee creamers, also known as "coffee whitener" or "coffee sweetener", are liquid or granular substances intended to substitute for

**Quality of Reduced-Fat Dairy Coffee Creamer: Affected** 

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.76367

**by Different Fat Replacer and Drying Methods**

**by Different Fat Replacer and Drying Methods**

Simin Hedayatnia and Hamed Mirhosseini

Simin Hedayatnia and Hamed Mirhosseini

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.76367

compared to all creamers.

drying

**1. Introduction**

**Abstract**

#### **Quality of Reduced-Fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying Methods Quality of Reduced-Fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying Methods**

DOI: 10.5772/intechopen.76367

Simin Hedayatnia and Hamed Mirhosseini Simin Hedayatnia and Hamed Mirhosseini

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.76367

#### **Abstract**

This work aims to investigate the effects of inulin (0, 2.5, 5 and 7.5%, w/w) and maltodextrin (0, 15, 20 and 25%, w/w) as wall materials and fat replacers and drying techniques (i.e. spray drying and fluidized-bed drying) on physicochemical properties of regular and instant reduced-fat dairy creamers. The regular reduced-fat dairy creamer was produced by one-stage drying (i.e. spray drying), while the instant reduced-fat dairy creamer was produced by two-stage drying (i.e. spray drying followed by fluidized-bed drying). In this study, control (0% inulin and 0% maltodextrin) and two commercial regular and instant coffee creamers (A and B) were also considered for comparison purposes. The results showed that the regular creamer containing 25% maltodextrin and 7.5% inulin had the largest particle size, highest viscosity and most desirable wettability among all formulated regular creamers. The yield of reduced-fat coffee creamer was significantly increased from 43.55 to 94.60% by increasing the amount of fat replacers to the maximum level (25% maltodextrin and 7.5% inulin). The current study revealed that the application of fluidized-bed drying for agglomeration led to significantly improve the wettability and instant properties of the instant creamer. In this study, the formulated instant creamer containing 25% maltodextrin and 7.5% inulin was the most desirable product as compared to all creamers.

**Keywords:** reduced-fat dairy creamer, inulin, maltodextrin, spray drying, fluidized-bed drying

#### **1. Introduction**

Coffee is one of the most vastly consumed beverages. Coffee drink is usually consumed in black or white form, depending on the taste of the consumer. Coffee creamers, also known as "coffee whitener" or "coffee sweetener", are liquid or granular substances intended to substitute for

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

milk or cream as an additive to coffee or other beverages [18, 28]. As stated by Tuot et al. [51], a desired coffee creamer should have specific physicochemical and functional characteristics particularly in terms of solubility, viscosity and stability. In addition, it should provide a good whitening effect after adding to hot coffee or similar hot beverages [41]. During the past few years, the demand of consumers for healthier products has significantly increased, lowering the tendency for consumption of high-fat foods. Hence, one of the main health issues for coffee drinkers is the presence of high percentage of fat in creamer formulation. As reported by Lambert [33], coffee creamer mainly contains high amount of vegetable fat (37–51%) and corn syrup (41–46%). Sudha et al. [47] suggested that the replacement of fat with various fat replacers led to the reduction of fat content and calories in food products.

the reduced-fat creamers. Inulin and maltodextrin have been used as a proper drying agent, fat replacer and wall material in powder technology and processing. It was hypothesized that there is a possibility to produce the reduced-fat coffee creamer with more nutritional benefit by partial replacement of its fat with proper fat replacer (like inulin and maltodextrin). In this study, water activity (aw), wettability, apparent viscosity, solubility, particle size and color of differently formulated regular and instant reduced-fat creamers were examined. The one-stage drying (i.e. spray drying) was applied to produce the regular reduced-fat dairy creamer (RRDC), while two-stage drying (i.e. spray drying followed by fluidized-bed drying) was employed to manufacture the instant reduced-fat dairy creamer (IRDC). All formulated creamers were compared with the properties of control (0% inulin and 0% maltodextrin) and commercial creamers (A and B). To the best of our knowledge, non-data of the different dry-

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The following components were used in creamer formulation: Maltodextrin (DE = 10, Roquette Freres Co, Lestrem, France), long-chain inulin (Fibruline Xl, Warcoing, Warcoing, Belgium), silicon dioxide (Sigma Aldrich, St. Louis, MO, USA), dipotassium hydrogen phosphate (Nacalai Tesque Co, Kyoto, Japan) and soy lecithin (Kordel's Co, CA, USA). Other ingredients such as commercial skim milk powder, an instant coffee (Brazilian freeze-dried Gold Bon CAFÉ), regular commercial coffee creamer (A) and instant commercial coffee creamer (B), hydrogenated palm kernel oil, sugar and vanilla were purchased from the supermarket (Kuala Lumpur, Malaysia). **Table 1** shows the composition of regular and instant commercial

Reduced-fat creamer emulsions were produced according to a method described by Hedayatnia et al. [22] with minor modification (**Figure 1**). Initially, the dispersed phase (**A**) containing the hydrogenated palm kernel oil (8% w/w) and soy lecithin (emulsifier, 0.5% w/w) was mixed in a 100-mL beaker and kept in a thermo-controlled water bath (70°C and rotated at 100 rpm for 20 min). The aqueous phase (**B**) which consists of sodium caseinate (2.5% w/w),

**Composition Regular commercial creamer A Instant commercial creamer B**

**Table 1.** The composition of regular and instant commercial creamers applied for comparison purposes.

Fat (%) 34.0 34.6 Carbohydrate (%) 57.1 56.9 Protein (%) 2.0 1.3

ing process and components were reported about reduced-fat dairy creamer.

**2. Materials and methods**

creamers applied for comparison purposes.

**2.2. Creamer preparation**

**2.1. Materials**

There are some difficulties for the drying of coffee creamer due to the low glass transition temperatures (T<sup>g</sup> ) of the components such as high sugar and caseinate that leads to stickiness problems. In order to prevent stickiness and caking issue during storage, specific drying aids or wall materials (such as gum arabic, xanthan gum, maltodextrin, inulin, etc.) with high glass transition temperature (T<sup>g</sup> ) are used [21]. Maltodextrin is a carbohydrate polymer made up of D-glucose units with dextrose-equivalent (DE) of under 20 [40, 52]. Maltodextrin is used as a dispersing aid, flavor carrier, bulking agent, fat replacer, volume enhancer, texture modifier, encapsulating agent and wall material [12]. It has many advantages such as highly soluble, relatively low cost, neutral aroma, taste, mouth feel and good protection of flavors against oxidation as compared to other drying aids [50].

Inulin is a non-digestible prebiotic soluble carbohydrate with very low energy value [17]. It is used as a sweetener component, especially in combination with high-intensity sweeteners, texture modifier and fat replacer [23]. Dietary fibers such as inulin are functional ingredients which are commonly used in different food products in order to modify physical and structural properties of hydration, viscosity, texture, sensory characteristics and oil holding capacity and also prolong the shelf life of products [30, 37].

The final characteristics of the dried products are broadly affected by the drying type and condition. The spray drying techniques are one of the most commonly applied techniques for manufacturing creamer [4]. Spray drying involves the transformation of feed from a liquid or slurry form to dry powder [34]. Spray-dried powders may have small particles with low bulk density, leading to inadequate flowability and poor reconstitution properties, thus causing difficulties in handling, transportation and storage. Manufacturers require free-flowing powders without any dust, and these requirements are met just by applying agglomeration process [42]. Agglomeration is a combination of wetting and nucleation, consolidation and growth and attrition and breakage [25]. Fluidization is a promising alternative technology, which allows the simultaneous drying, encapsulation and agglomeration in a single stream, reducing operation costs, saving time, simultaneously reducing the caking issue and improving the physicochemical properties (i.e. flowability, density, dissolution and dispersion characteristics) of the powder [2, 5, 11, 39].

The present study was conducted to investigate the effect of inulin (0, 2.5, 5 and 7.5%, w/w) and maltodextrin (0, 15, 20 and 25%, w/w) and fluidized-bed drying on the characteristics of the reduced-fat creamers. Inulin and maltodextrin have been used as a proper drying agent, fat replacer and wall material in powder technology and processing. It was hypothesized that there is a possibility to produce the reduced-fat coffee creamer with more nutritional benefit by partial replacement of its fat with proper fat replacer (like inulin and maltodextrin). In this study, water activity (aw), wettability, apparent viscosity, solubility, particle size and color of differently formulated regular and instant reduced-fat creamers were examined. The one-stage drying (i.e. spray drying) was applied to produce the regular reduced-fat dairy creamer (RRDC), while two-stage drying (i.e. spray drying followed by fluidized-bed drying) was employed to manufacture the instant reduced-fat dairy creamer (IRDC). All formulated creamers were compared with the properties of control (0% inulin and 0% maltodextrin) and commercial creamers (A and B). To the best of our knowledge, non-data of the different drying process and components were reported about reduced-fat dairy creamer.

## **2. Materials and methods**

#### **2.1. Materials**

milk or cream as an additive to coffee or other beverages [18, 28]. As stated by Tuot et al. [51], a desired coffee creamer should have specific physicochemical and functional characteristics particularly in terms of solubility, viscosity and stability. In addition, it should provide a good whitening effect after adding to hot coffee or similar hot beverages [41]. During the past few years, the demand of consumers for healthier products has significantly increased, lowering the tendency for consumption of high-fat foods. Hence, one of the main health issues for coffee drinkers is the presence of high percentage of fat in creamer formulation. As reported by Lambert [33], coffee creamer mainly contains high amount of vegetable fat (37–51%) and corn syrup (41–46%). Sudha et al. [47] suggested that the replacement of fat with various fat replac-

There are some difficulties for the drying of coffee creamer due to the low glass transition

problems. In order to prevent stickiness and caking issue during storage, specific drying aids or wall materials (such as gum arabic, xanthan gum, maltodextrin, inulin, etc.) with high glass

D-glucose units with dextrose-equivalent (DE) of under 20 [40, 52]. Maltodextrin is used as a dispersing aid, flavor carrier, bulking agent, fat replacer, volume enhancer, texture modifier, encapsulating agent and wall material [12]. It has many advantages such as highly soluble, relatively low cost, neutral aroma, taste, mouth feel and good protection of flavors against

Inulin is a non-digestible prebiotic soluble carbohydrate with very low energy value [17]. It is used as a sweetener component, especially in combination with high-intensity sweeteners, texture modifier and fat replacer [23]. Dietary fibers such as inulin are functional ingredients which are commonly used in different food products in order to modify physical and structural properties of hydration, viscosity, texture, sensory characteristics and oil holding capac-

The final characteristics of the dried products are broadly affected by the drying type and condition. The spray drying techniques are one of the most commonly applied techniques for manufacturing creamer [4]. Spray drying involves the transformation of feed from a liquid or slurry form to dry powder [34]. Spray-dried powders may have small particles with low bulk density, leading to inadequate flowability and poor reconstitution properties, thus causing difficulties in handling, transportation and storage. Manufacturers require free-flowing powders without any dust, and these requirements are met just by applying agglomeration process [42]. Agglomeration is a combination of wetting and nucleation, consolidation and growth and attrition and breakage [25]. Fluidization is a promising alternative technology, which allows the simultaneous drying, encapsulation and agglomeration in a single stream, reducing operation costs, saving time, simultaneously reducing the caking issue and improving the physicochemical properties (i.e. flowability, density, dissolution and dispersion char-

The present study was conducted to investigate the effect of inulin (0, 2.5, 5 and 7.5%, w/w) and maltodextrin (0, 15, 20 and 25%, w/w) and fluidized-bed drying on the characteristics of

) of the components such as high sugar and caseinate that leads to stickiness

) are used [21]. Maltodextrin is a carbohydrate polymer made up of

ers led to the reduction of fat content and calories in food products.

temperatures (T<sup>g</sup>

116 Descriptive Food Science

transition temperature (T<sup>g</sup>

oxidation as compared to other drying aids [50].

ity and also prolong the shelf life of products [30, 37].

acteristics) of the powder [2, 5, 11, 39].

The following components were used in creamer formulation: Maltodextrin (DE = 10, Roquette Freres Co, Lestrem, France), long-chain inulin (Fibruline Xl, Warcoing, Warcoing, Belgium), silicon dioxide (Sigma Aldrich, St. Louis, MO, USA), dipotassium hydrogen phosphate (Nacalai Tesque Co, Kyoto, Japan) and soy lecithin (Kordel's Co, CA, USA). Other ingredients such as commercial skim milk powder, an instant coffee (Brazilian freeze-dried Gold Bon CAFÉ), regular commercial coffee creamer (A) and instant commercial coffee creamer (B), hydrogenated palm kernel oil, sugar and vanilla were purchased from the supermarket (Kuala Lumpur, Malaysia). **Table 1** shows the composition of regular and instant commercial creamers applied for comparison purposes.

#### **2.2. Creamer preparation**

Reduced-fat creamer emulsions were produced according to a method described by Hedayatnia et al. [22] with minor modification (**Figure 1**). Initially, the dispersed phase (**A**) containing the hydrogenated palm kernel oil (8% w/w) and soy lecithin (emulsifier, 0.5% w/w) was mixed in a 100-mL beaker and kept in a thermo-controlled water bath (70°C and rotated at 100 rpm for 20 min). The aqueous phase (**B**) which consists of sodium caseinate (2.5% w/w),


**Table 1.** The composition of regular and instant commercial creamers applied for comparison purposes.

drying chamber. In the present study, spray drying procedure was set at the following condition: inlet temperature, 180 ± 5°C; outlet temperature, 80 ± 5°C; pressure, 552 kPa; and feed

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In this study, a laboratory scale fluidized-bed dryer (Aeromatic-Fielder AG, GEA Co, Copenhagen, Denmark) was used for agglomeration process under the following experimental condition: 50°C (inlet fluidizing air temperature), 5 mL/min (solution feed rate) and 1.5 m/s (atomizing air pressure) for 30 min. In this study, the creamer powder (150 g) was placed in a container. Then, 30-mL aqueous solution of lecithin concentration (2%, w/w) was fed by a peristaltic pump and sprayed from a spray nozzle, which was located at the top of the chamber. The lecithin solution acts as a binder during the drying process as recommended for fluidization process by Dhanalakshmi et al. [13]. The solution droplets fell down on the creamer powders, while the filtrated hot air from the bottom of the chamber flowed throughout the chamber to reduce the moisture content and dustiness of particles. The atomization of the feed solution was stopped for 5 min every 10 min during fluidization, and the gas flow rate was increased steadily to ensure the proper flow pattern of the solids, and the balance between the coating and agglomeration mechanisms (layering and particle coalescence) could be reached. This helps to compensate the moisture and prevent further stickiness in the drying chamber. Vanilla (5% w/w) was added at the final drying stage because of thermal sensitivity of aromatic compounds. Additional flavors could be added to enhance the overall

Water activity (aw) of all regular and instant creamers was measured in triplicate by using an AquaLab water activity metre (Series 3TE, Decagon Devices Inc., Pullman, WA, USA) with

Average particle size (D[4,3]) was determined by measuring the volume-weighted mean diameter (de Brouckere mean diameter, D4,3) in triplicate for each sample. The experiments was performed by means of a particle size analyzer with powder feeder unit (Model 2000 hydro S, Malvern Instrument, Worcestershire, UK) equipped with a Mastersizer software 2000 (Version 5.13). The volume-weighted mean diameter is estimated by the following equation:

[15].

is the number of particles with diameter D<sup>i</sup>

rate, 10 mL/min.

**2.4. Fluidized-bed drying**

flavor of the reduced-fat products [53].

**3. Analytical tests**

**3.1. Water activity (aw)**

±0.001 sensitivity at 21°C.

**3.2. Average particle size**

where ni

**Figure 1.** Schematic description of the preparation of reduced-fat creamer.

silicon dioxide (1.0% w/w), dipotassium hydrogen phosphate (2.5% w/w), skim milk powder 7% (w/w) and corn syrup solid (15% w/w) was prepared by gradually dispersing them into 100-mL hot distilled water (70 ± 5°C) and stirred consequently with a magnetic at 100 rpm for 5 min. Subsequently, different concentrations of inulin (0.0, 2.5, 5.0 and 7.5% w/w) and maltodextrin (0, 15, 20 and 25%) (**C**) were gradually added to the aqueous phase (**B**) to prepare the emulsion continuous phase. Hot distilled water (70 ± 5°C) was added to each creamer formulation up to 100%. In the final stage, upon mixing the ingredients, dispersed phase (**A**) was gradually added to the premix and gently stirred for 10 min. Subsequently, the coarse creamer emulsion was homogenized by a high-pressure homogenizer (at 200- and 180-MPa pressure for 2 cycles) prior to the drying process. All formulated reduced-fat creamers and control were prepared under the same drying condition depending on the regular or instant case.

#### **2.3. Spray drying procedure**

After homogenization, the creamer emulsion was fed into a lab scale mini spray dryer (BÜCHI model B-290, Flawil, Switzerland). The samples were atomized with a rotary atomizer into the drying chamber. In the present study, spray drying procedure was set at the following condition: inlet temperature, 180 ± 5°C; outlet temperature, 80 ± 5°C; pressure, 552 kPa; and feed rate, 10 mL/min.

#### **2.4. Fluidized-bed drying**

In this study, a laboratory scale fluidized-bed dryer (Aeromatic-Fielder AG, GEA Co, Copenhagen, Denmark) was used for agglomeration process under the following experimental condition: 50°C (inlet fluidizing air temperature), 5 mL/min (solution feed rate) and 1.5 m/s (atomizing air pressure) for 30 min. In this study, the creamer powder (150 g) was placed in a container. Then, 30-mL aqueous solution of lecithin concentration (2%, w/w) was fed by a peristaltic pump and sprayed from a spray nozzle, which was located at the top of the chamber. The lecithin solution acts as a binder during the drying process as recommended for fluidization process by Dhanalakshmi et al. [13]. The solution droplets fell down on the creamer powders, while the filtrated hot air from the bottom of the chamber flowed throughout the chamber to reduce the moisture content and dustiness of particles. The atomization of the feed solution was stopped for 5 min every 10 min during fluidization, and the gas flow rate was increased steadily to ensure the proper flow pattern of the solids, and the balance between the coating and agglomeration mechanisms (layering and particle coalescence) could be reached. This helps to compensate the moisture and prevent further stickiness in the drying chamber. Vanilla (5% w/w) was added at the final drying stage because of thermal sensitivity of aromatic compounds. Additional flavors could be added to enhance the overall flavor of the reduced-fat products [53].

## **3. Analytical tests**

## **3.1. Water activity (aw)**

silicon dioxide (1.0% w/w), dipotassium hydrogen phosphate (2.5% w/w), skim milk powder 7% (w/w) and corn syrup solid (15% w/w) was prepared by gradually dispersing them into 100-mL hot distilled water (70 ± 5°C) and stirred consequently with a magnetic at 100 rpm for 5 min. Subsequently, different concentrations of inulin (0.0, 2.5, 5.0 and 7.5% w/w) and maltodextrin (0, 15, 20 and 25%) (**C**) were gradually added to the aqueous phase (**B**) to prepare the emulsion continuous phase. Hot distilled water (70 ± 5°C) was added to each creamer formulation up to 100%. In the final stage, upon mixing the ingredients, dispersed phase (**A**) was gradually added to the premix and gently stirred for 10 min. Subsequently, the coarse creamer emulsion was homogenized by a high-pressure homogenizer (at 200- and 180-MPa pressure for 2 cycles) prior to the drying process. All formulated reduced-fat creamers and control were

**Figure 1.** Schematic description of the preparation of reduced-fat creamer.

prepared under the same drying condition depending on the regular or instant case.

After homogenization, the creamer emulsion was fed into a lab scale mini spray dryer (BÜCHI model B-290, Flawil, Switzerland). The samples were atomized with a rotary atomizer into the

**2.3. Spray drying procedure**

118 Descriptive Food Science

Water activity (aw) of all regular and instant creamers was measured in triplicate by using an AquaLab water activity metre (Series 3TE, Decagon Devices Inc., Pullman, WA, USA) with ±0.001 sensitivity at 21°C.

#### **3.2. Average particle size**

Average particle size (D[4,3]) was determined by measuring the volume-weighted mean diameter (de Brouckere mean diameter, D4,3) in triplicate for each sample. The experiments was performed by means of a particle size analyzer with powder feeder unit (Model 2000 hydro S, Malvern Instrument, Worcestershire, UK) equipped with a Mastersizer software 2000 (Version 5.13). The volume-weighted mean diameter is estimated by the following equation:

$$\mathbf{D} \text{ [}4, 3\text{]} = \Sigma \text{nħ} \text{D}\_{\text{i}}^{4} / \Sigma \text{nġ} \text{D}\_{\text{i}}^{3}$$

where ni is the number of particles with diameter D<sup>i</sup> [15].

#### **3.3. Wettability determination**

The wettability of creamers was determined according to the method described by Gong et al. [19] with minor modification. In this experiment, 100-mL hot distilled water (70 ± 5°C) was poured into a 250-mL glass beaker; then 10 g of creamer powder was poured into the beaker. The time required for the powder to completely become wet was recorded as wetting time. This measurement was carried out in triplicate for each sample.

#### **3.4. Apparent viscosity measurement**

The apparent viscosity of all creamers was measured with a rheometer (RheolabQC Rheometer, Anton Paar Co, Österreich, Austria) at room temperature (25 ± 1°C). The experiment was conducted by reconstituting 20-g creamer with100-mL hot distilled water (70 ± 5°C). Then, 25 ml of prepared solution (20%, w/w) was shaken prior to analysis. Prior to shearing test, all samples were left 5 min to reach the equilibrium condition. Apparent viscosity was measured in triplicate for each sample.

#### **3.5. Color evaluation**

The color intensity of all creamers was measured by a Hunter Lab colorimeter (Model A60– 1012-402, Fairfax County, VA, USA). The color intensity was expressed in the CIELAB space as L\* (lightness; 0 = black, 100 = white) and b\* (+b = yellowness, −b = blueness) values [24]. For color measurement, 10 mg of sample was placed in a transparent polypropylene bag for analysis. The color measurement was done in triplicate for each sample.

#### **3.6. Yield determination**

The drying yield was measured according to the method described by Koocheki et al. [31]. The averages of three individual measurements were considered for each sample:

$$Y = 100 \times \left(^{\text{M}\_{\text{M}\_z}}\right)$$

M1 = mass of initial ingredients (g); M2 = mass of final powders (g).

#### **3.7. Experimental design and statistical analysis**

A full factorial design technique was considered to prepare different samples (**Table 2**). Oneway analysis of variance (ANOVA) and Fisher's multiple comparison tests were used to find out the significant (*p* < 0.05) or insignificant (*p* > 0.05) difference among all samples. Then, the data were subjected to two-way analysis of variance (ANOVA) to determine the main and interaction effects of inulin and maltodextrin on the creamer characteristics. The degree of significance of all independent variables could be determined with F-ratio. The factor or independent variable with higher F-ratio represents the factor with more significant powerful effect and vice versa [38]. Also, the t-test was applied to analyze the significant (*p* < 0.05) difference among mean values of samples before and after agglomeration process. Minitab

**Components** **CS**

Maltodextrin (%)

Inulin (%) HPKO (%) Sodium caseinate (%)

Skim milk powder (%)

DPHP (%) Silicon dioxide (%)

Lecithin (%) Solid corn (%)

Vanilla (%)

**Table 2.**

The compositions of different reduced-fat creamer slurries.

15.0

5.0

5.0

5.0

5.0 HPKO, hydrogenated palm kernel oil; DPHP, dipotassium hydrogen phosphate; final volume was adjusted up to 100

5.0

5.0

5.0

5.0

5.0

5.0

5.0

ml by distilled water.

5.0

5.0

5.0

5.0

5.0

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15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

15.0

1.0 2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

7.0 2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

7.0

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

2.5

0 0 8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

8.0

2.5

5.0

7.5

0

2.5

5.0

7.5

0

2.5

5.0

7.5

0

2.5

5.0

7.5

0

0

0

15

15

15

15

20

20

20

20

25

25

25

25

**A**

**B**

**C**

**D**

**E**

**F**

**G**

**H**

**I**

**J**

**K**

**L**

**M**

**N**

**O**

**Formulations**


**3.3. Wettability determination**

120 Descriptive Food Science

**3.4. Apparent viscosity measurement**

in triplicate for each sample.

**3.5. Color evaluation**

**3.6. Yield determination**

The wettability of creamers was determined according to the method described by Gong et al. [19] with minor modification. In this experiment, 100-mL hot distilled water (70 ± 5°C) was poured into a 250-mL glass beaker; then 10 g of creamer powder was poured into the beaker. The time required for the powder to completely become wet was recorded as wetting time.

The apparent viscosity of all creamers was measured with a rheometer (RheolabQC Rheometer, Anton Paar Co, Österreich, Austria) at room temperature (25 ± 1°C). The experiment was conducted by reconstituting 20-g creamer with100-mL hot distilled water (70 ± 5°C). Then, 25 ml of prepared solution (20%, w/w) was shaken prior to analysis. Prior to shearing test, all samples were left 5 min to reach the equilibrium condition. Apparent viscosity was measured

The color intensity of all creamers was measured by a Hunter Lab colorimeter (Model A60– 1012-402, Fairfax County, VA, USA). The color intensity was expressed in the CIELAB space as L\* (lightness; 0 = black, 100 = white) and b\* (+b = yellowness, −b = blueness) values [24]. For color measurement, 10 mg of sample was placed in a transparent polypropylene bag for

The drying yield was measured according to the method described by Koocheki et al. [31].

A full factorial design technique was considered to prepare different samples (**Table 2**). Oneway analysis of variance (ANOVA) and Fisher's multiple comparison tests were used to find out the significant (*p* < 0.05) or insignificant (*p* > 0.05) difference among all samples. Then, the data were subjected to two-way analysis of variance (ANOVA) to determine the main and interaction effects of inulin and maltodextrin on the creamer characteristics. The degree of significance of all independent variables could be determined with F-ratio. The factor or independent variable with higher F-ratio represents the factor with more significant powerful effect and vice versa [38]. Also, the t-test was applied to analyze the significant (*p* < 0.05) difference among mean values of samples before and after agglomeration process. Minitab

*<sup>M</sup>*<sup>2</sup> )

The averages of three individual measurements were considered for each sample:

analysis. The color measurement was done in triplicate for each sample.

M1 = mass of initial ingredients (g); M2 = mass of final powders (g).

*Y* = 100 × (*M*1⁄

**3.7. Experimental design and statistical analysis**

This measurement was carried out in triplicate for each sample.

**Table 2.** The compositions of different reduced-fat creamer slurries. version 16 (Minitab Inc., State College, PA, USA) was used to run the statistical analysis. All formulated creamers were compared with control and two commercial creamers (A and B) to investigate the impact of different fat replacers and drying techniques.

0.26 with increase in maltodextrin and inulin contents. The samples containing higher maltodextrin and inulin contents had lower water activity, while control sample (CS) had the highest water activity among all formulated creamers (**Figure 2**a). The control sample is probably more hygroscopic than the other formulated creamers. The result showed that the control creamer adsorbed more water than other formulated creamers. It had the highest stickiness and very poor reconstitution properties as well. In this case, Kumar and Mishra [32] explained that the proper water activity of powder should be between 0.20 and 0.25. Water activity (aw) of instant spray-dried samples also varied from 0.36 to 0.24 (**Figure 2**b). It was concluded that the single and interaction effects of inulin and maltodextrin significantly (*p* < 0.05) affected the water activity of regular and instant spray-dried creamers (**Table 3**). Maltodextrin has highly hygroscopic effect with the extensive ability to capture the free moisture, so aw reduction can

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123

**Table 4** also shows that the water activity of the creamer was significantly influenced by fluidized-bed drying. This difference was analyzed by comparing the water activity of the regular and instant creamer before and after fluidized-bed drying, respectively. Maltodextrin showed more significant (*p* < 0.05) effects than inulin as indicated by its higher F-ratio. It was observed that the agglomeration induced by fluidized-bed drying significantly (*p* < 0.05) decreased the water activity of instant creamers. This might be attributed to long residence time (about '30 min) and hot air (50°C) injected throughout fluidized-bed the dryer. The hot air caused

**Creamer Creamer characteristics Linear effect Interaction effect R2**

Regular Water activity 0.000 52 0.000 793 0.000 1 0.971

Instant Water activity 0.000 585 0.000 1389 0.000 12 0.998

particle size 0.000 14 0.000 1744 0.000 74 0.994 Apparent viscosity 0.000 48 0.000 1442 0.278<sup>a</sup> 1 0.990 Wettability 0.000 22 0.000 832 0.006 4 0.988 Lightness (L\*) 0.000 158 0.000 5181 0.000 40 0.997 Yellowness (b\*) 0.000 134 0.000 8691 0.000 22 0.999 Yield 0.367<sup>a</sup> 1 0.002 8 0.477<sup>a</sup> 1 0.690

particle size 0.000 108 0.000 2259 0.001 7 0.995 Apparent viscosity 0.000 859 0.000 38,957 0.000 95 0.999 Wettability 0.000 550 0.000 800 0.000 5 0.988 Lightness (L\*) 0.000 7609 0.000 265,995 0.000 206 0.999 Yellowness (b\*) 0.000 474 0.000 15,137 0.000 32 0.997 Yield 0.000 23 0.000 4009 0.000 12 0.998

**Table 3.** Two-way ANOVA showing the single main effect and interaction effect of inulin and maltodextrin on

**Inulin Maltodextrin Inulin\* Maltodextrin p-value F-ratio p-value F-ratio p-value F-ratio**

be due to such maltodextrin function.

a

Non-significant (*p* > 0.05).

characteristics of regular and instant reduced-fat dairy creamers.

#### **4. Results and dissociation**

#### **4.1. Effect of different fat replacers and drying techniques on water activity**

**Figure 2** shows that the water activity (aw) of the formulated creamers was significantly (*p* < 0.05) affected by the type and content of fat replacer as well as drying technique. As shown in **Figure 2**a, water activity of regular creamers significantly decreased from 0.36 to

**Figure 2.** Water activity of differently formulated regular creamers (a) and instant creamers (b) as compared to the control (CS) and two regular and instant commercial creamers (CO) (a and B). a–m Significant differences at the confidence level of *p* ≤ 0.05 (mean ± SD, n = 3); A–C: Creamers with 0% maltodextrin and 2.5, 5.0 and 7.5% inulin, respectively. D–G: Creamers with 15% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. H–K: Creamers with 20% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. L–O: Creamers with 25% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively.

0.26 with increase in maltodextrin and inulin contents. The samples containing higher maltodextrin and inulin contents had lower water activity, while control sample (CS) had the highest water activity among all formulated creamers (**Figure 2**a). The control sample is probably more hygroscopic than the other formulated creamers. The result showed that the control creamer adsorbed more water than other formulated creamers. It had the highest stickiness and very poor reconstitution properties as well. In this case, Kumar and Mishra [32] explained that the proper water activity of powder should be between 0.20 and 0.25. Water activity (aw) of instant spray-dried samples also varied from 0.36 to 0.24 (**Figure 2**b). It was concluded that the single and interaction effects of inulin and maltodextrin significantly (*p* < 0.05) affected the water activity of regular and instant spray-dried creamers (**Table 3**). Maltodextrin has highly hygroscopic effect with the extensive ability to capture the free moisture, so aw reduction can be due to such maltodextrin function.

**Table 4** also shows that the water activity of the creamer was significantly influenced by fluidized-bed drying. This difference was analyzed by comparing the water activity of the regular and instant creamer before and after fluidized-bed drying, respectively. Maltodextrin showed more significant (*p* < 0.05) effects than inulin as indicated by its higher F-ratio. It was observed that the agglomeration induced by fluidized-bed drying significantly (*p* < 0.05) decreased the water activity of instant creamers. This might be attributed to long residence time (about '30 min) and hot air (50°C) injected throughout fluidized-bed the dryer. The hot air caused


**Table 3.** Two-way ANOVA showing the single main effect and interaction effect of inulin and maltodextrin on characteristics of regular and instant reduced-fat dairy creamers.

**Figure 2.** Water activity of differently formulated regular creamers (a) and instant creamers (b) as compared to the control (CS) and two regular and instant commercial creamers (CO) (a and B). a–m Significant differences at the confidence level of *p* ≤ 0.05 (mean ± SD, n = 3); A–C: Creamers with 0% maltodextrin and 2.5, 5.0 and 7.5% inulin, respectively. D–G: Creamers with 15% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. H–K: Creamers with 20% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. L–O: Creamers with 25% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin,

version 16 (Minitab Inc., State College, PA, USA) was used to run the statistical analysis. All formulated creamers were compared with control and two commercial creamers (A and B) to

**Figure 2** shows that the water activity (aw) of the formulated creamers was significantly (*p* < 0.05) affected by the type and content of fat replacer as well as drying technique. As shown in **Figure 2**a, water activity of regular creamers significantly decreased from 0.36 to

investigate the impact of different fat replacers and drying techniques.

**4.1. Effect of different fat replacers and drying techniques on water activity**

**4. Results and dissociation**

122 Descriptive Food Science

respectively.


a Insignificant at *p* > 0.05.

**Table 4.** Student *t-*test for significant comparison of the regular and instant reduced-fat dairy creamers.

more water evaporation from the surface of the particles, thus providing less stickiness (wall deposition). The results showed that the regular creamers had higher water activity than the instant creamers, indicating a higher amount of freely available water for the undesirable biological reactions and consequently inducing shorter shelf life.

#### **4.2. Effect of different fat replacers and drying techniques on wettability and apparent viscosity**

**Table 5** shows the wettability and apparent viscosity of different reduced-fat dairy creamer as compared to two commercial creamers and control. The wettability and viscosity of the regular and instant dairy creamer were significantly (*p* < 0.05) affected by the different compositions and drying techniques. Wettability is an important instant property of powder, and it is defined as the ability of particles to overcome the surface tension between the liquid (solvent) and themselves [14]. It is also directly affected by the interactions between two phases [9]. In general, wettability is considered to be the rate-controlling step of the reconstitution process [29] with the surface content strongly affecting the wettability [16]. In the current study, the regular (75 s) and instant (63 s) control creamers showed the longest wetting time among all formulated creamers (**Table 5**). This might be explained by the effects of fat replacer and total solid contents on the wettability of the reduced-fat creamer (**Table 6**). The presence of free fat on the particle surface could reduce wettability of control sample due to its hydrophobicity, making it difficult for water to penetrate into the powder.

As shown in **Table 5**, the regular spray-dried creamers exhibited different levels of wettability (40–75.50 s), while the instant spray-dried creamers lower faster wettability (13–61 s) than regular creamers with similar formulations. This was comparable with the wettability of the regular commercial creamer A (43 s) and instant commercial creamer B (17 s), respectively. The results showed that the wetting time of regular and instant creamers was decreased by increasing the particle size and decreasing the water activity of creamers. Jakubczyk et al. [26] reported that the wettability of the apple puree powder was improved from '45 to '33 (s) by increasing maltodextrin from 6 to 15% (w/w). The amount of wall materials significantly affected the wettability of the final powder. The results showed that there was a reverse relationship between wetting time and the content of wall materials (i.e. inulin and maltodextrin).

**Sample Regular creamers Instant creamers**

**Yield (%) Wettability (s)**

Quality of Reduced-Fat Dairy Coffee Creamer: Affected by Different Fat Replacer and Drying...

66.50 ± 0.70bc 5.37 ± 0.02b 55.03 ± 0.04<sup>i</sup> 44.00 ± 2.12c 6.01 ± 0.01<sup>k</sup> 55.03 ± 0.04<sup>i</sup>

61.50 ± 0.83d 5.54 ± 0.01bc 61.45 ± 0.63<sup>h</sup> 39.50 ± 3.53d 6.18 ± 0.01j 61.45 ± 0.63<sup>h</sup>

58.50 ± 0.75<sup>e</sup> 5.81 ± 0.02cd 66.00 ± 1.41<sup>g</sup> 36.00 ± 1.41<sup>e</sup> 6.56 ± 0.02f 66.00 ± 1.41<sup>g</sup>

50.50 ± 0.65<sup>g</sup> 6.04 ± 0.03d 75.66 ± 0.48<sup>e</sup> 30.00 ± 0.00<sup>g</sup> 6.75 ± 0.02<sup>e</sup> 75.66 ± 0.48<sup>e</sup>

44.00 ± 1.40<sup>h</sup> 6.35 ± 0.01d 85.06 ± 0.09c 24.00 ± 0.73<sup>h</sup> 7.12 ± 0.01c 85.06 ± 0.09c

40.50 ± 0.77ij 6.53 ± 0.03d 94.60 ± 0.56<sup>a</sup> 13.00 ± 1.40j 7.32 ± 0.01b 94.60 ± 0.56<sup>a</sup>

43.00 ± 1.41hi 5.82 ± 0.02d — 17.00 ± 0.00<sup>i</sup> 7.29 ± 0.00b —

\*Control (0% inulin and maltodextrin); mean values ± standard deviation with different lowercase letters in the same

**Table 5.** Significant differences (*p* < 0.05) among different regular and instant reduced-fat dairy creamers as compared to

column indicating significant difference (*P* < 0.05); control (0%MA & 0%IN); MA, maltodextrin; IN, inulin.

two regular and instant commercial creamers in terms of wettability, apparent viscosity and yield.

Control\* 75.00 ± 0.82<sup>a</sup> 4.75 ± 0.19<sup>a</sup> 43.00 ± 1.83l 63.00 ± 0.00<sup>a</sup> 4.85 ± 0.01<sup>n</sup> 43.30 ± 1.20l MA0%, IN2.5% 75.50 ± 0.70<sup>a</sup> 4.88 ± 0.02ab 43.55 ± 0.62l 61.00 ± 1.4<sup>a</sup> 4.84 ± 0.01<sup>n</sup> 43.55 ± 0.62l MA 0%, IN 5% 75.50 ± 0.70<sup>a</sup> 5.04 ± 0.06b 47.00 ± 0.00<sup>k</sup> 56.50 ± 0.70b 5.21 ± 0.01<sup>m</sup> 47.00 ± 0.31<sup>k</sup> MA0%, IN7.5% 76.00 ± 0.93<sup>a</sup> 5.06 ± 0.05b 47.77 ± 0.31<sup>k</sup> 55.50 ± 0.70b 5.29 ± 0.01l 47.77 ± 0.31<sup>k</sup> MA15%, IN0% 69.00 ± 0.97b 5.32 ± 0.02b 53.35 ± 0.50 j 54.00 ± 1.41b 6.03 ± 0.02<sup>k</sup> 53.35 ± 0.50 j

MA15%, IN 5% 64.00 ± 1.06cd 5.46 ± 0.02b 60.55 ± 0.78<sup>h</sup> 40.00 ± 0.00d 6.21 ± 0.01<sup>i</sup> 60.55 ± 0.78<sup>h</sup>

MA20%, IN0% 58.50 ± 0.70<sup>e</sup> 5.76 ± 0.02cd 65.80 ± 1.13<sup>g</sup> 35.00 ± 3.53ef 6.49 ± 0.01<sup>h</sup> 65.80 ± 1.13<sup>g</sup>

MA20%, IN 5% 54.00 ± 1.21f 5.89 ± 0.01cd 70.60 ± 0.55f 32.50 ± 0.70fg 6.52 ± 0.00<sup>g</sup> 70.60 ± 0.55f

MA25%, IN0% 45.50 ± 2.05<sup>h</sup> 6.29 ± 0.00d 79.77 ± 0.48d 24.00 ± 1.41<sup>h</sup> 7.09 ± 0.01d 79.77 ± 0.48d

MA25%, IN 5% 40.00 ± 0.09j 6.41 ± 0.01d 89.45 ± 0.77b 19.00 ± 1.40<sup>i</sup> 7.35 ± 0.01<sup>a</sup> 89.45 ± 0.77b

**Apparent viscosity** 

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**Yield (%)**

125

**(mPa.s)**

**Apparent viscosity (mPa.s)**

**Wettability (s)**

MA15%, IN2.5%

MA15%, IN7.5%

MA20%, IN2.5%

MA20%, IN7.5%

MA25%, IN2.5%

MA25%, IN7.5%

Commercial creamers


\*Control (0% inulin and maltodextrin); mean values ± standard deviation with different lowercase letters in the same column indicating significant difference (*P* < 0.05); control (0%MA & 0%IN); MA, maltodextrin; IN, inulin.

more water evaporation from the surface of the particles, thus providing less stickiness (wall deposition). The results showed that the regular creamers had higher water activity than the instant creamers, indicating a higher amount of freely available water for the undesirable

**Table 5** shows the wettability and apparent viscosity of different reduced-fat dairy creamer as compared to two commercial creamers and control. The wettability and viscosity of the regular and instant dairy creamer were significantly (*p* < 0.05) affected by the different compositions and drying techniques. Wettability is an important instant property of powder, and it is defined as the ability of particles to overcome the surface tension between the liquid (solvent) and themselves [14]. It is also directly affected by the interactions between two phases [9]. In general, wettability is considered to be the rate-controlling step of the reconstitution process [29] with the surface content strongly affecting the wettability [16]. In the current study, the regular (75 s) and instant (63 s) control creamers showed the longest wetting time among all formulated creamers (**Table 5**). This might be explained by the effects of fat replacer and total solid contents on the wettability of the reduced-fat creamer (**Table 6**). The presence of free fat on the particle surface could reduce wettability of control sample due to its hydrophobicity,

biological reactions and consequently inducing shorter shelf life.

**Creamers Mean p-value T-value Test**

RSRC 0.33 0.000 5.87 Water activity (aw)

RSRC 5.66 0.000 −12.84 Apparent viscosity

RSRC 58.68 0.000 29.55 Wettability

RSRC 66.05 0.000 −9.96 Particle size

RSRC 84.33 0.000 12.61 Lightness (L\*)

RSRC 13.79 0.000 −14.99 Yellowness (b\*)

RSRC 64.40 0.360<sup>a</sup> −0.92 Yield

making it difficult for water to penetrate into the powder.

**apparent viscosity**

ISRC 0.29

124 Descriptive Food Science

ISRC 37.97

ISRC 6.30

ISRC 120.15

ISRC 81.01

ISRC 15.70

ISRC 66.59

Insignificant at *p* > 0.05.

a

RSRC: Regular spray-dried reduced-fat creamer. ISRC: Instant spray-dried reduced-fat creamer.

**4.2. Effect of different fat replacers and drying techniques on wettability and** 

**Table 4.** Student *t-*test for significant comparison of the regular and instant reduced-fat dairy creamers.

**Table 5.** Significant differences (*p* < 0.05) among different regular and instant reduced-fat dairy creamers as compared to two regular and instant commercial creamers in terms of wettability, apparent viscosity and yield.

As shown in **Table 5**, the regular spray-dried creamers exhibited different levels of wettability (40–75.50 s), while the instant spray-dried creamers lower faster wettability (13–61 s) than regular creamers with similar formulations. This was comparable with the wettability of the regular commercial creamer A (43 s) and instant commercial creamer B (17 s), respectively. The results showed that the wetting time of regular and instant creamers was decreased by increasing the particle size and decreasing the water activity of creamers. Jakubczyk et al. [26] reported that the wettability of the apple puree powder was improved from '45 to '33 (s) by increasing maltodextrin from 6 to 15% (w/w). The amount of wall materials significantly affected the wettability of the final powder. The results showed that there was a reverse relationship between wetting time and the content of wall materials (i.e. inulin and maltodextrin).


creamers was significantly (*p* < 0.05) increased by increasing the concentration of maltodextrin and inulin in the creamer formulation. This could be due to increasing the total solids of samples and positive effect of inulin and maltodextrin on the formation of a stable particle gel with tridimensional network in the present of aqueous phase which led to enhance the viscosity of creamer. Debon et al. [10] also reported the similar observation. They reported that the addition of 5% inulin to the formulation significantly increased the apparent viscosity of low-fat fermented milk. It might be due to several different factors such as (i) the interaction between inulin and milk protein (i.e. sodium caseinate) leading to enhance the viscosity, (ii) the high inulin capacity to retain water [46] and (iii) the capacity to retain water by formation of small aggregates of inulin microcrystal [20]. This might be due to the increase in the

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127

The apparent viscosity of the spray-dried creamers was greatly enhanced after agglomeration via fluidized-bed drying (**Table 5**). This could be explained by the significant (*p* < 0.05) effects of the agglomeration on the intermolecular interactions among creamer particles which resulted in higher viscosity. In addition, the lower moisture content and higher total solid content can be also responsible for the higher viscosity of the instant creamers. Water acts as a mobility enhancer, resulting in a larger free volume and a reduction in the viscosity [48].

In the current study, the average particle size of different creamers was determined by measuring the volume-weighted mean. The particle size of the powder can significantly affect its appearance, flowability, wettability and dispensability [43]. **Figure 4** showed a significantly increase in the particle size of different formulated creamers. The results showed that the creamer O containing 25% maltodextrin and 7.5% inulin exhibited the largest particle size (101.45 μm), while the control sample had the smallest particle size among all regular spray-dried dairy creamers, respectively (**Figure 4a**). As stated by Master [36], the particle size is highly influenced by the viscosity of the feed. Similar observations were previously reported by Jinapong et al. [27] wherein increasing the solids content of instant spray-dried soymilk powders from 5.2 to 13.0%

**Figure 4b** shows that agglomeration process by fluidized-bed drying technique significantly (*p* < 0.05) increased the volume-weighted mean (D4, 3) or particle size of the instant spray creamers compared to the regular creamers. As stated by Chen and Özkan [8], agglomeration results in larger particles, bigger particle clusters and better flow characteristics than one-stage drying. The instant spray-dried creamers exhibited different particle size, ranging from 46.45 to 193.26 μm compared to control creamer (47.35 μm) and the commercial instant creamer B (190.94 μm) (**Figure 4b**). As stated by Carić [7], the most suitable particle size for rapid dispersion is around 150–200 μm. Binder in the wet agglomeration process leads to the enlargement of particle size, thereby improving the flowability of the final powder [3, 44]. Jinapong et al. [27] reported that the particle size of the instant spray-dried soymilk was increased from 25 to 260 μm after subjecting the sample to fluidized-bed drying. This finding was also reported also by Machado et al. [35] wherein agglomerated soy protein had much larger particles than non-agglomerated regular creamer. In addition, it was found that the agglomeration through fluidized-bed drying also caused stickiness reduction and flowability

**4.3. Effect of different fat replacers and drying techniques on average particle size**

significantly resulted in enlargement of the particle size from 14.54 to 23.59 (μm).

amount of soluble materials and reduction in the moisture content.

r = r Pearson correlation; significant at *p* < 0.05; r > 0.9 represents positive strong correlation, while and r > −0.9 represents negative strong correlation.

**Table 6.** Correlation analysis between total solid content, inulin and maltodextrin contents and creamer characteristics.

**Figure 3.** Dissolution timeline for regular and agglomerated (instant) powder showing the overlaps between different phases with time [14].

The creamer containing the highest maltodextrin (25%) and inulin (7.5%) showed the shortest wetting time among all formulated creamers (**Table 5**).

The instant creamer exhibited significantly higher wettability (shorter wettability) than the regular creamer (**Table 5**) due to large particles exhibiting more empty spaces among themselves, resulting in easier penetration by the liquid (i.e. water) [15]. Lecithin can modify the flowability and wettability of dried powders due to its potential surface active properties with higher porosity and better wettability [13]. **Figure 3** clearly shows the schematic of dissolution timeline for standard and agglomerated powder [14] by correlation between the wettability, dispersibility and solubility in the regular (non-agglomerated) and instant creamers.

As shown in **Table 5**, the apparent viscosity of regular spray-dried creamers varied from 4.88 to 6.53 (mPa.s) as compared to the control (4.75 mPa.s) and regular commercial creamer A (5.82 mPa.s). In addition, the viscosity of the instant spray-dried creamers varied from 4.84 to 7.35 (mPa.s) compared to the commercial instant creamer B (7.29 mPa.s) and control sample (4.85 mPa.s) (**Table 5**). The result showed that the apparent viscosity of regular and instant creamers was significantly (*p* < 0.05) increased by increasing the concentration of maltodextrin and inulin in the creamer formulation. This could be due to increasing the total solids of samples and positive effect of inulin and maltodextrin on the formation of a stable particle gel with tridimensional network in the present of aqueous phase which led to enhance the viscosity of creamer. Debon et al. [10] also reported the similar observation. They reported that the addition of 5% inulin to the formulation significantly increased the apparent viscosity of low-fat fermented milk. It might be due to several different factors such as (i) the interaction between inulin and milk protein (i.e. sodium caseinate) leading to enhance the viscosity, (ii) the high inulin capacity to retain water [46] and (iii) the capacity to retain water by formation of small aggregates of inulin microcrystal [20]. This might be due to the increase in the amount of soluble materials and reduction in the moisture content.

The apparent viscosity of the spray-dried creamers was greatly enhanced after agglomeration via fluidized-bed drying (**Table 5**). This could be explained by the significant (*p* < 0.05) effects of the agglomeration on the intermolecular interactions among creamer particles which resulted in higher viscosity. In addition, the lower moisture content and higher total solid content can be also responsible for the higher viscosity of the instant creamers. Water acts as a mobility enhancer, resulting in a larger free volume and a reduction in the viscosity [48].

#### **4.3. Effect of different fat replacers and drying techniques on average particle size**

The creamer containing the highest maltodextrin (25%) and inulin (7.5%) showed the shortest

**Figure 3.** Dissolution timeline for regular and agglomerated (instant) powder showing the overlaps between different

The instant creamer exhibited significantly higher wettability (shorter wettability) than the regular creamer (**Table 5**) due to large particles exhibiting more empty spaces among themselves, resulting in easier penetration by the liquid (i.e. water) [15]. Lecithin can modify the flowability and wettability of dried powders due to its potential surface active properties with higher porosity and better wettability [13]. **Figure 3** clearly shows the schematic of dissolution timeline for standard and agglomerated powder [14] by correlation between the wettability,

As shown in **Table 5**, the apparent viscosity of regular spray-dried creamers varied from 4.88 to 6.53 (mPa.s) as compared to the control (4.75 mPa.s) and regular commercial creamer A (5.82 mPa.s). In addition, the viscosity of the instant spray-dried creamers varied from 4.84 to 7.35 (mPa.s) compared to the commercial instant creamer B (7.29 mPa.s) and control sample (4.85 mPa.s) (**Table 5**). The result showed that the apparent viscosity of regular and instant

dispersibility and solubility in the regular (non-agglomerated) and instant creamers.

wetting time among all formulated creamers (**Table 5**).

**Characteristics Inulin Maltodextrin Total solid** 

0.180 0.506

−0.156 0.564

0.656 0.045\* 0.938 0.000\*

−0.925 0.000\*

0.907 0.000\*

r = r Pearson correlation; significant at *p* < 0.05; r > 0.9 represents positive strong correlation, while and r > −0.9 represents

**Table 6.** Correlation analysis between total solid content, inulin and maltodextrin contents and creamer characteristics.

Viscosity r-correlation

126 Descriptive Food Science

Wettability r-correlation

Particle size r-correlation

negative strong correlation.

phases with time [14].

p-value

p-value

p-value

**content**

0.950 0.000\*

−0.931 0.000\*

0.924 0.000\* **Wettability Viscosity**

— —

—

0.941 0.000\*

−0.988 0.000\*

−0.903 0.001\*

> In the current study, the average particle size of different creamers was determined by measuring the volume-weighted mean. The particle size of the powder can significantly affect its appearance, flowability, wettability and dispensability [43]. **Figure 4** showed a significantly increase in the particle size of different formulated creamers. The results showed that the creamer O containing 25% maltodextrin and 7.5% inulin exhibited the largest particle size (101.45 μm), while the control sample had the smallest particle size among all regular spray-dried dairy creamers, respectively (**Figure 4a**). As stated by Master [36], the particle size is highly influenced by the viscosity of the feed. Similar observations were previously reported by Jinapong et al. [27] wherein increasing the solids content of instant spray-dried soymilk powders from 5.2 to 13.0% significantly resulted in enlargement of the particle size from 14.54 to 23.59 (μm).

> **Figure 4b** shows that agglomeration process by fluidized-bed drying technique significantly (*p* < 0.05) increased the volume-weighted mean (D4, 3) or particle size of the instant spray creamers compared to the regular creamers. As stated by Chen and Özkan [8], agglomeration results in larger particles, bigger particle clusters and better flow characteristics than one-stage drying. The instant spray-dried creamers exhibited different particle size, ranging from 46.45 to 193.26 μm compared to control creamer (47.35 μm) and the commercial instant creamer B (190.94 μm) (**Figure 4b**). As stated by Carić [7], the most suitable particle size for rapid dispersion is around 150–200 μm. Binder in the wet agglomeration process leads to the enlargement of particle size, thereby improving the flowability of the final powder [3, 44]. Jinapong et al. [27] reported that the particle size of the instant spray-dried soymilk was increased from 25 to 260 μm after subjecting the sample to fluidized-bed drying. This finding was also reported also by Machado et al. [35] wherein agglomerated soy protein had much larger particles than non-agglomerated regular creamer. In addition, it was found that the agglomeration through fluidized-bed drying also caused stickiness reduction and flowability

The low yield was observed for the control. This could be due to the stickiness of this sample to the spray drying chamber and cyclone wall. This might be because the control did not contain inulin and very low percentage of maltodextrin as a drying aid. Stickiness is one of the main technological issues in the production of powders such as coffee creamer because it results in a reduction of the yield and stability. The result showed significant improvement in the production yield by increasing the maltodextrin and inulin content in the formulation (**Table 5**). This was in agreement with the previous finding reported by Shrestha et al. [45] for spray-dried tomato pulp. In fact, the addition of drying aids such as maltodextrin with high glass transition temperature (>145°C) to the premix is one of the most suitable ways to increase the stability, decrease the stickiness and improve the yield [21, 45]. In addition, maltodextrin can help to shorten the drying time, thus reducing the input energy required for spray drying process. According to Adhikari et al. [1], the improvement of yield (recovery) might be due to the formation of a thin protein-rich membrane at the particle-air interface. The high glass transition temperature of this surface layer causes the conversion of this thin membrane into a glassy state, which prevents particles from sticking to each other and to the walls of the dryer which resulted in the decrease of the wall deposition during drying and increase of the yield. As shown in **Table 3**, maltodextrin with higher F-ratio had higher significant effects than inulin on the yield. There was no significant (*p* > 0.05) difference between the yields of single- or double-step drying processes for regular and instant creamer, respectively.

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**Figure 5.** The appearance of regular commercial creamer (a) as compared to the control creamer (b) and regular

formulated-reduced-fat creamer containing 25% maltodextrin and 7.5% inulin.

**Figure 4.** Volume-weighted mean (or average particle size) of differently formulated regular creamers (a) and instant creamers (b) as compared to the control (CS) and two regular and instant commercial creamers (CO) (a and B). a–m Significant differences at the confidence level of *p* ≤ 0.05 (mean ± SD, n = 3); A–C: Creamers with 0% maltodextrin and 2.5, 5.0 and 7.5% inulin, respectively. D–G: Creamers with 15% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. H–K: Creamers with 20% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. L–O: Creamers with 25% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively.

improvement in the creamer. As the particle size increases, the adhesive forces decrease due to the attractive forces (i.e. van der Waals) among creamer particles [54]. However, the application of the spray drying process followed by the agglomeration process resulted in a significant (*p* < 0.05) better flowability and reconstitution properties than the spray drying alone. Dhanalakshmi et al. [13] indicated that the powder flowability and reconstitution are highly affected by the particle size, shape and distribution, the particle surface properties as well as the geometry of the system. As shown in **Table 3**, the linear and interaction effects of the inulin and maltodextrin had significant (*p* < 0.05) effect on the volume-weighed mean (or particle size) of spray-dried creamers.

#### **4.4. Effect of different fat replacers and drying techniques on yield**

**Table 5** shows the yield of regular and instant dairy creamers compared to the control sample. The control sample had remarkably lower yield (43%) than other regular spray-dried creamers. The low yield was observed for the control. This could be due to the stickiness of this sample to the spray drying chamber and cyclone wall. This might be because the control did not contain inulin and very low percentage of maltodextrin as a drying aid. Stickiness is one of the main technological issues in the production of powders such as coffee creamer because it results in a reduction of the yield and stability. The result showed significant improvement in the production yield by increasing the maltodextrin and inulin content in the formulation (**Table 5**). This was in agreement with the previous finding reported by Shrestha et al. [45] for spray-dried tomato pulp. In fact, the addition of drying aids such as maltodextrin with high glass transition temperature (>145°C) to the premix is one of the most suitable ways to increase the stability, decrease the stickiness and improve the yield [21, 45]. In addition, maltodextrin can help to shorten the drying time, thus reducing the input energy required for spray drying process. According to Adhikari et al. [1], the improvement of yield (recovery) might be due to the formation of a thin protein-rich membrane at the particle-air interface. The high glass transition temperature of this surface layer causes the conversion of this thin membrane into a glassy state, which prevents particles from sticking to each other and to the walls of the dryer which resulted in the decrease of the wall deposition during drying and increase of the yield. As shown in **Table 3**, maltodextrin with higher F-ratio had higher significant effects than inulin on the yield. There was no significant (*p* > 0.05) difference between the yields of single- or double-step drying processes for regular and instant creamer, respectively.

improvement in the creamer. As the particle size increases, the adhesive forces decrease due to the attractive forces (i.e. van der Waals) among creamer particles [54]. However, the application of the spray drying process followed by the agglomeration process resulted in a significant (*p* < 0.05) better flowability and reconstitution properties than the spray drying alone. Dhanalakshmi et al. [13] indicated that the powder flowability and reconstitution are highly affected by the particle size, shape and distribution, the particle surface properties as well as the geometry of the system. As shown in **Table 3**, the linear and interaction effects of the inulin and maltodextrin had significant (*p* < 0.05) effect on the volume-weighed mean (or particle

**Figure 4.** Volume-weighted mean (or average particle size) of differently formulated regular creamers (a) and instant creamers (b) as compared to the control (CS) and two regular and instant commercial creamers (CO) (a and B). a–m Significant differences at the confidence level of *p* ≤ 0.05 (mean ± SD, n = 3); A–C: Creamers with 0% maltodextrin and 2.5, 5.0 and 7.5% inulin, respectively. D–G: Creamers with 15% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. H–K: Creamers with 20% maltodextrin and 0, 2.5, 5.0 and 7.5% inulin, respectively. L–O: Creamers with 25% maltodextrin and

**Table 5** shows the yield of regular and instant dairy creamers compared to the control sample. The control sample had remarkably lower yield (43%) than other regular spray-dried creamers.

**4.4. Effect of different fat replacers and drying techniques on yield**

size) of spray-dried creamers.

0, 2.5, 5.0 and 7.5% inulin, respectively.

128 Descriptive Food Science

**Figure 5.** The appearance of regular commercial creamer (a) as compared to the control creamer (b) and regular formulated-reduced-fat creamer containing 25% maltodextrin and 7.5% inulin.

It means that there was not any material loss or stickiness to the wall during fluidized-bed dying process. This could be attributed to the high efficiency of such drying technique.

creamer A (**Table 7**). According to Roland et al. (1999), reduced-fat ice cream made by only maltodextrin exhibited similar whiteness to 10% fat ice cream. Moreover, the control and creamer containing 0% maltodextrin and 2.5% inulin had the highest b\*among all samples. On the other hand, the regular and instant samples containing 25% maltodextrin exhibited similar yellowness (b\*) as compared to regular and instant commercial samples (A and B). The instant commercial creamer B and the formulated creamer containing 25% maltodextrin and 7.5% inulin had the lowest b\* among all instant creamers (**Table 7**). The lightness (L\*) and yellowness (b\*) of the instant creamers were slightly decreased after applying the agglomeration process. This might be attributed to the effects of binder solution and Maillard reaction on the lightness of the instant creamer. Szulc and Lenart [49] also reported similar findings for the agglomerated dairy powders. They explained that Maillard reaction was responsible for the color changes during agglomeration (i.e. fluidize-bed drying). Color can change during the drying process due to several chemical reactions. Most of the time, enzymatic activity is not desirable, because it affects the amount of nutrients in food (e.g. hydrolysis of lecithin by phospholipase) or the color of the products. The yellowish color of regular and instant creamer is mainly due to the non-enzymatic reaction, either by caramelization or by Maillard reactions. The caramelization process is a complex series of chemical reactions promoted by the direct heating of sugars [6].

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The present work describes the possibility of producing regular and instant reduced-fat dairy creamers by spray and fluid-bed drying and the changes in some of the physical, chemical and powder properties of the creamer powders depending on the maltodextrin and inulin levels. A significant effect of the type and concentration of the fat replacers (wall materials) on the process yield, wettability, viscosity, solubility, color, water activity and particle size was found. The results showed that the process has some difficulties for drying of control samples. The use of wall materials (maltodextrin and inulin) significantly improved the drying process and leads to improve the physicochemical properties of reduced-fat dairy creamer functionality. The highest wettability, viscosity, solubility, yield and lightness and lowest water activities were obtained from the samples containing the highest contents of maltodextrin (25% w/w) and inulin (7.5%). As a result, the current study also revealed that the instant formulated-reduced-fat creamers from two-stage drying (spray drying followed by fluidizedbed drying) showed significantly (*p* < 0.05) better quality than regular creamer from one-stage drying (spray dying only). The current study suggests optimizing the fluidized-bed drying condition for preparation and commercialization of instant reduced-fat dairy creamer.

Faculty of Food Science and Technology, Department of Food Technology, University Putra

**5. Conclusions**

**Author details**

Simin Hedayatnia and Hamed Mirhosseini\*

Malaysia, Serdang, Selangor, Malaysia

\*Address all correspondence to: hamedmi@upm.edu.my

#### **4.5. Effect of different fat replacers and drying techniques on color**

**Figure 5** shows the appearance of the regular creamer as compared to the control and commercial creamer. The results indicated that differently formulated creamers, control and commercial creamer had significant (*p* < 0.05) different color intensity in terms of L\* and b\* (**Table 7**). In general, the lightness (L\*) of all formulated regular and instant creamers was enhanced by increasing maltodextrin and inulin concentrations in the creamer formulation. The result indicated that the addition of maltodextrin and inulin to the creamer formulation led to decrease the stickiness and increase the lightness of samples. Similar observation was reported by Shrestha et al. [45] on spray-dried orange juice. The regular and instant creamer with 25% maltodextrin and 7.5% inulin had the highest L\*, while the control showed the lowest L\*among all samples due to the caramelization reaction (**Table 7**). The regular creamer containing the highest maltodextrin content (25%) exhibited almost similar color to the regular commercial


\*Control (0% inulin and maltodextrin); mean values ± standard deviations with different lowercase letters in the same column indicating significant difference (*P* < 0.05); MA, maltodextrin; IN, inulin.

**Table 7.** Significant different (*p* < 0.05) color intensity of different regular and instant reduced-fat dairy creamers as compared to two regular and instant commercial creamers.

creamer A (**Table 7**). According to Roland et al. (1999), reduced-fat ice cream made by only maltodextrin exhibited similar whiteness to 10% fat ice cream. Moreover, the control and creamer containing 0% maltodextrin and 2.5% inulin had the highest b\*among all samples. On the other hand, the regular and instant samples containing 25% maltodextrin exhibited similar yellowness (b\*) as compared to regular and instant commercial samples (A and B). The instant commercial creamer B and the formulated creamer containing 25% maltodextrin and 7.5% inulin had the lowest b\* among all instant creamers (**Table 7**). The lightness (L\*) and yellowness (b\*) of the instant creamers were slightly decreased after applying the agglomeration process. This might be attributed to the effects of binder solution and Maillard reaction on the lightness of the instant creamer. Szulc and Lenart [49] also reported similar findings for the agglomerated dairy powders. They explained that Maillard reaction was responsible for the color changes during agglomeration (i.e. fluidize-bed drying). Color can change during the drying process due to several chemical reactions. Most of the time, enzymatic activity is not desirable, because it affects the amount of nutrients in food (e.g. hydrolysis of lecithin by phospholipase) or the color of the products. The yellowish color of regular and instant creamer is mainly due to the non-enzymatic reaction, either by caramelization or by Maillard reactions. The caramelization process is a complex series of chemical reactions promoted by the direct heating of sugars [6].

## **5. Conclusions**

It means that there was not any material loss or stickiness to the wall during fluidized-bed dying process. This could be attributed to the high efficiency of such drying technique.

**Figure 5** shows the appearance of the regular creamer as compared to the control and commercial creamer. The results indicated that differently formulated creamers, control and commercial creamer had significant (*p* < 0.05) different color intensity in terms of L\* and b\* (**Table 7**). In general, the lightness (L\*) of all formulated regular and instant creamers was enhanced by increasing maltodextrin and inulin concentrations in the creamer formulation. The result indicated that the addition of maltodextrin and inulin to the creamer formulation led to decrease the stickiness and increase the lightness of samples. Similar observation was reported by Shrestha et al. [45] on spray-dried orange juice. The regular and instant creamer with 25% maltodextrin and 7.5% inulin had the highest L\*, while the control showed the lowest L\*among all samples due to the caramelization reaction (**Table 7**). The regular creamer containing the highest maltodextrin content (25%) exhibited almost similar color to the regular commercial

**Regular creamers Instant creamers**

95.44 ± 0.40<sup>a</sup> 7.00 ± 0.07l 88.59 ± 0.00d 8.20 ± 0.02<sup>m</sup>

\*Control (0% inulin and maltodextrin); mean values ± standard deviations with different lowercase letters in the same

**Table 7.** Significant different (*p* < 0.05) color intensity of different regular and instant reduced-fat dairy creamers as

column indicating significant difference (*P* < 0.05); MA, maltodextrin; IN, inulin.

compared to two regular and instant commercial creamers.

Sample L\* b\* L\* b\*

Commercial creamers (Regular and instant)

130 Descriptive Food Science

Control\* 71.07 ± 0.05l 22.49 ± 0.21<sup>a</sup> 70.73 ± 0.21<sup>o</sup> 24.57 ± 0.05<sup>a</sup> MA0%, IN2.5% 73.28 ± 0.33<sup>k</sup> 22.04 ± 0.06b 71.20 ± 0.11<sup>n</sup> 23.71 ± 0.21b MA0%, IN5% 72.67 ± 0.27<sup>k</sup> 21.09 ± 0.13c 72.23 ± 0.00<sup>m</sup> 23.62 ± 0.11b MA0%, IN7.5% 71.48 ± 0.55l 20.14 ± 0.04d 70.46 ± 0.02p 22.50 ± 0.47c MA15%, IN0% 78.91 ± 0.07j 18.37 ± 0.36<sup>e</sup> 74.05 ± 0.00l 20.69 ± 0.09d MA15%, IN2.5% 82.30 ± 1.47<sup>i</sup> 18.09 ± 0.07<sup>e</sup> 76.38 ± 0.03<sup>k</sup> 20.42 ± 0.17d MA15%, IN 5% 84.63 ± 0.21<sup>h</sup> 17.46 ± 0.53f 79.13 ± 0.02j 17.47 ± 0.00<sup>e</sup> MA15%, IN7.5% 85.57 ± 0.19<sup>g</sup> 15.15 ± 0.00<sup>g</sup> 80.48 ± 0.01<sup>i</sup> 17.44 ± 0.00<sup>e</sup> MA20%, IN0% 87.46 ± 0.24f 11.63 ± 0.02<sup>h</sup> 82.11 ± 0.00<sup>h</sup> 13.92 ± 0.00f MA20%, IN2.5% 87.12 ± 0.01f 11.03 ± 0.02<sup>i</sup> 84.05 ± 0.11<sup>g</sup> 13.32 ± 0.00<sup>g</sup> MA20%, IN 5% 89.28 ± 0.17<sup>e</sup> 10.28 ± 0.01j 84.30 ± 0.02f 12.61 ± 0.02<sup>h</sup> MA20%, IN7.5% 89.80 ± 0.00de 10.83 ± 0.08<sup>i</sup> 86.73 ± 0.02<sup>e</sup> 11.76 ± 0.03<sup>i</sup> MA25%, IN0% 90.90 ± 0.50c 7.73 ± 0.00<sup>k</sup> 88.73 ± 0.02c 10.79 ± 0.11j MA25%, IN2.5% 90.51 ± 0.00cd 7.22 ± 0.30l 88.71 ± 0.01c 9.21 ± 0.00<sup>k</sup> MA25%, IN 5% 92.56 ± 0.57b 6.99 ± 0.00l 89.17 ± 0.19b 8.56 ± 0.01l MA25%, IN7.5% 95.32 ± 0.00<sup>a</sup> 6.85 ± 0.07l 90.15 ± 0.01<sup>a</sup> 8.08 ± 0.07<sup>m</sup>

**4.5. Effect of different fat replacers and drying techniques on color**

The present work describes the possibility of producing regular and instant reduced-fat dairy creamers by spray and fluid-bed drying and the changes in some of the physical, chemical and powder properties of the creamer powders depending on the maltodextrin and inulin levels. A significant effect of the type and concentration of the fat replacers (wall materials) on the process yield, wettability, viscosity, solubility, color, water activity and particle size was found. The results showed that the process has some difficulties for drying of control samples. The use of wall materials (maltodextrin and inulin) significantly improved the drying process and leads to improve the physicochemical properties of reduced-fat dairy creamer functionality. The highest wettability, viscosity, solubility, yield and lightness and lowest water activities were obtained from the samples containing the highest contents of maltodextrin (25% w/w) and inulin (7.5%). As a result, the current study also revealed that the instant formulated-reduced-fat creamers from two-stage drying (spray drying followed by fluidizedbed drying) showed significantly (*p* < 0.05) better quality than regular creamer from one-stage drying (spray dying only). The current study suggests optimizing the fluidized-bed drying condition for preparation and commercialization of instant reduced-fat dairy creamer.

## **Author details**

Simin Hedayatnia and Hamed Mirhosseini\*

\*Address all correspondence to: hamedmi@upm.edu.my

Faculty of Food Science and Technology, Department of Food Technology, University Putra Malaysia, Serdang, Selangor, Malaysia

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[38] Mirhosseini H, Tan CP, Hamid NS, Yusof S, Chern BH. Characterization of the influence of main emulsion components on the physicochemical properties of orange beverage emulsion using response surface methodology. Food Hydrocolloids. 2009;**23**:271-280 [39] Mukherjee S, Bhattacharya S. Characterization of agglomeration process as a function of moisture content using a model food powder. Journal of Texture Studies. 2006;**37**:35-48

[40] Nep EI, Conway BR. Physicochemical characterization of grewia polysaccharide gum:

[41] Oldfield D, Singh H. Functional properties of milk powders. Food Science and

[42] Pietsch W. Readily engineer agglomerates with special properties from micro-and nano-

[43] Reineccius GA. The spray drying of food flavors. Drying Technology. 2004;**22**:1289-1324 [44] Shittu TA, Lawal MO. Factors affecting instant properties of powdered cocoa beverages.

[45] Shrestha AK, Howes T, Adhikari BP, Bhandari BR. Water sorption and glass transition properties of spray dried lactose hydrolysed skim milk powder. LWT- Food Science and

[46] Soukoulis C, Lebesi D, Tzia C. Enrichment of ice cream with dietary fibre: Effects on rheological properties, ice crystallisation and glass transition phenomena. Food Chemistry.

Effect of drying method. Carbohydrate Polymers. 2011;**84**:446-453

sized particles. Chemical Engineering Progress. 1999;**95**:67-81

Technology-New York-Marcel Dekker. 2005;**146**:365

Food Chemistry. 2007;**100**:91-98

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1998;**60**:301-309

2014;**254**:248-255


**Chapter 7**

**Provisional chapter**

**Effects of Radiation Technologies on Food Nutritional**

**Effects of Radiation Technologies on Food Nutritional** 

Generally, foods are thermally processed to destroy the vegetative microorganisms for food preservation. However, only thermal treatment triggers many unwanted biochemical reactions, which leads to undesirable sensorial and nutritional effects. Therefore, a number of nontraditional preservation techniques are being developed to satisfy consumer demand with regard to nutritional and sensory aspects of foods. Ensuring food safety and at the same time meeting such demands for retention of nutrition and quality attributes has resulted in increased interest in emerging preservation techniques. These emerging food preservation technologies can extend the shelf life of unprocessed or processed foods by inactivating the enzymes, reducing the food spoiling microbial growth rate or viability without altering the food quality attributes including flavor, odor, color, texture, and nutritional value. On the emerging technology, sphere promising results have been attained by the radiation processes. Ionizing radiation has been widely used in industrial processes, especially in the sterilization of medicals, pharmaceuticals, cosmetic products, and in food processing. Nonionizing radiation has been used in surface decontamination, as on the packaging sector. Although radiation techniques, as the other traditional or emerging techniques, can impair alterations that can modify the chemical composition and the nutritional value of foods, these changes depend on the food composition, the irradiation dose and factors such as temperature and presence or absence of oxygen in the irradiating environment. The aim of this chapter is to discuss retention or loss of vitamins and minerals in several food products submitted to a radiation process (gamma irradiation, short wave ultraviolet, microwave, and pulsed electric fields). **Keywords:** nutritional, nonionizing radiation, ionizing radiation, micronutrient,

> © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.80437

Fabiana Lima, Kássia Vieira, Miriam Santos and

Fabiana Lima, Kássia Vieira, Miriam Santos

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

**Quality**

**Quality**

Poliana Mendes de Souza

**Abstract**

and Poliana Mendes de Souza

http://dx.doi.org/10.5772/intechopen.80437

macronutrient, emerging technologies

#### **Effects of Radiation Technologies on Food Nutritional Quality Effects of Radiation Technologies on Food Nutritional Quality**

DOI: 10.5772/intechopen.80437

Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de Souza Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de Souza

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.80437

#### **Abstract**

Generally, foods are thermally processed to destroy the vegetative microorganisms for food preservation. However, only thermal treatment triggers many unwanted biochemical reactions, which leads to undesirable sensorial and nutritional effects. Therefore, a number of nontraditional preservation techniques are being developed to satisfy consumer demand with regard to nutritional and sensory aspects of foods. Ensuring food safety and at the same time meeting such demands for retention of nutrition and quality attributes has resulted in increased interest in emerging preservation techniques. These emerging food preservation technologies can extend the shelf life of unprocessed or processed foods by inactivating the enzymes, reducing the food spoiling microbial growth rate or viability without altering the food quality attributes including flavor, odor, color, texture, and nutritional value. On the emerging technology, sphere promising results have been attained by the radiation processes. Ionizing radiation has been widely used in industrial processes, especially in the sterilization of medicals, pharmaceuticals, cosmetic products, and in food processing. Nonionizing radiation has been used in surface decontamination, as on the packaging sector. Although radiation techniques, as the other traditional or emerging techniques, can impair alterations that can modify the chemical composition and the nutritional value of foods, these changes depend on the food composition, the irradiation dose and factors such as temperature and presence or absence of oxygen in the irradiating environment. The aim of this chapter is to discuss retention or loss of vitamins and minerals in several food products submitted to a radiation process (gamma irradiation, short wave ultraviolet, microwave, and pulsed electric fields).

**Keywords:** nutritional, nonionizing radiation, ionizing radiation, micronutrient, macronutrient, emerging technologies

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **1. Introduction**

The first food preservation methods were developed by men in the beginning of its existence. Since ancient times, radiation has been used as a preserve food method, when used to preserve meats, fish, fruits, and vegetables from solar energy. In the drying method, the men realized that some fruits and vegetables were still edible and the meat lasted longer without spoiling, once this process consists in removal of part of the water from the food to a final concentration. The microorganisms present in food need water for their vital functions, and by reducing the water content, it is possible to decrease considerably the microbiological growth in the food [1].

The growing demand from the consumers for high quality products reveals the need to apply of new food preservation techniques that provide microbiological safety in the production, increasing the commercial validity, and still provide minimal biochemical alterations, promoting the maintenance of the nutritional and sensorial quality of the products [2].

Some of the currently proposed methods inhibit microbial growth, even in some cases, they cause enzyme inactivation, without the considerable increase of the product temperature. Moreover, these processes promote little damage to pigments, flavor compounds, and vitamins; and in contrast to conventional processes that apply high temperatures, the original sensory and nutritional characteristics of the food are maintained, with no significant loss of fresh product quality [3].

Throughout the centuries, the food preservation techniques have been improving with the increase of scientific development. These methods include freezing, drying, canning, pickling preparation, pasteurization, fermentation, cooling, controlled atmosphere storage, and application of preservatives, and among the emerging technologies, the highlight is the radiation technique [4].

wavelengths in the visible range. Sir Isaac Newton was the first to realize this fact in the late 1660s by passing the sunlight into a rainbow using a prism and turning it back into white light

Effects of Radiation Technologies on Food Nutritional Quality

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139

In search for new and better food preservation methods, the research has turned to the possibility of using radiations in different ranges of frequencies, such as nonionizing radiations. This type of radiation does not have sufficient energy to break molecular bonds or remove electrons from atoms; in other words, perform ionization. The types of frequencies characterized as nonionizing are ultraviolet, infrared, radiofrequency, laser, microwave, and visible light [9, 10]. However, the use of nonionizing radiation has low interest as a food preservation

Electromagnetic energy in the microwave region (300 MHz–300 GHz) has been extensively studied as an alternative energy source for sterilization. Microwave radiation has enormous applications in our daily routine, from communication systems, medical applications, to the

Microwave radiation does not carry enough energy to ionize food molecules or to generate free radicals [13]. The basic principle of the process is the baking by molecular vibration that

with a second prism [8].

**Figure 1.** Electromagnetic spectrum.

**2.1. Microwave radiation**

**2. Nonionizing radiation**

method, in contrast to ionizing radiations [7].

level of industrial application [11, 12].

The terminology radiation refers to the physical processes of emission and propagation of energy, either by means of wave phenomena, or by the kinetic energy from the particles. In a simpler way, radiation is energy that spreads from one point to another in space or in a material environment. Irradiation is the process of applying this energy to a material, such as food, with the purpose of sterilizing or preserving them through the destruction of microorganisms, parasites, insects, and other pests [5, 6].

There are various forms of radiant energy emitted in various ways that belong to the electromagnetic radiation spectrum, and these include radio waves, microwaves, infrared radiation, visible light, ultraviolet, X-rays, and gamma rays. These forms of energy differ in the wavelength, frequency, force of penetration, and other effects that they exert on biological systems. As can be observed in **Figure 1**, in general, it can be said that in the electromagnetic spectrum, there are two types of radiation: ionizing and nonionizing [7].

Of all the bands of the electromagnetic spectrum, the best known is the narrow band of visible light, shown in the expanded form in the **Figure 1**. The human brain recognizes these wavelengths as the different colors ranging from violet to red. The sunlight, as the human being is able to see with the naked eye, is the sum of multiple electromagnetic waves with different

**Figure 1.** Electromagnetic spectrum.

**1. Introduction**

138 Descriptive Food Science

growth in the food [1].

fresh product quality [3].

ganisms, parasites, insects, and other pests [5, 6].

there are two types of radiation: ionizing and nonionizing [7].

technique [4].

The first food preservation methods were developed by men in the beginning of its existence. Since ancient times, radiation has been used as a preserve food method, when used to preserve meats, fish, fruits, and vegetables from solar energy. In the drying method, the men realized that some fruits and vegetables were still edible and the meat lasted longer without spoiling, once this process consists in removal of part of the water from the food to a final concentration. The microorganisms present in food need water for their vital functions, and by reducing the water content, it is possible to decrease considerably the microbiological

The growing demand from the consumers for high quality products reveals the need to apply of new food preservation techniques that provide microbiological safety in the production, increasing the commercial validity, and still provide minimal biochemical alterations, pro-

Some of the currently proposed methods inhibit microbial growth, even in some cases, they cause enzyme inactivation, without the considerable increase of the product temperature. Moreover, these processes promote little damage to pigments, flavor compounds, and vitamins; and in contrast to conventional processes that apply high temperatures, the original sensory and nutritional characteristics of the food are maintained, with no significant loss of

Throughout the centuries, the food preservation techniques have been improving with the increase of scientific development. These methods include freezing, drying, canning, pickling preparation, pasteurization, fermentation, cooling, controlled atmosphere storage, and application of preservatives, and among the emerging technologies, the highlight is the radiation

The terminology radiation refers to the physical processes of emission and propagation of energy, either by means of wave phenomena, or by the kinetic energy from the particles. In a simpler way, radiation is energy that spreads from one point to another in space or in a material environment. Irradiation is the process of applying this energy to a material, such as food, with the purpose of sterilizing or preserving them through the destruction of microor-

There are various forms of radiant energy emitted in various ways that belong to the electromagnetic radiation spectrum, and these include radio waves, microwaves, infrared radiation, visible light, ultraviolet, X-rays, and gamma rays. These forms of energy differ in the wavelength, frequency, force of penetration, and other effects that they exert on biological systems. As can be observed in **Figure 1**, in general, it can be said that in the electromagnetic spectrum,

Of all the bands of the electromagnetic spectrum, the best known is the narrow band of visible light, shown in the expanded form in the **Figure 1**. The human brain recognizes these wavelengths as the different colors ranging from violet to red. The sunlight, as the human being is able to see with the naked eye, is the sum of multiple electromagnetic waves with different

moting the maintenance of the nutritional and sensorial quality of the products [2].

wavelengths in the visible range. Sir Isaac Newton was the first to realize this fact in the late 1660s by passing the sunlight into a rainbow using a prism and turning it back into white light with a second prism [8].

## **2. Nonionizing radiation**

In search for new and better food preservation methods, the research has turned to the possibility of using radiations in different ranges of frequencies, such as nonionizing radiations. This type of radiation does not have sufficient energy to break molecular bonds or remove electrons from atoms; in other words, perform ionization. The types of frequencies characterized as nonionizing are ultraviolet, infrared, radiofrequency, laser, microwave, and visible light [9, 10]. However, the use of nonionizing radiation has low interest as a food preservation method, in contrast to ionizing radiations [7].

#### **2.1. Microwave radiation**

Electromagnetic energy in the microwave region (300 MHz–300 GHz) has been extensively studied as an alternative energy source for sterilization. Microwave radiation has enormous applications in our daily routine, from communication systems, medical applications, to the level of industrial application [11, 12].

Microwave radiation does not carry enough energy to ionize food molecules or to generate free radicals [13]. The basic principle of the process is the baking by molecular vibration that penetrates superficially in the foods, and the depth of penetration ranges from 2 to 4 cm. The process makes the food vibrates and consequently heating the water, fat, and sugar molecules. However, microwave heating leads to uneven distribution of the temperature in the product. In addition, the products do not show nonenzymatic darkening and surface crust formation, which might be advantageous in some applications [10].

UV-light is a physical process that offers several advantages, such as it does not produce by-products that could change the characteristics of the food, does not generate chemical residues, and it is a cold process, dry, simple, effective, and low cost in relation to other steril-

UV-light has been applied in water purification and has been used in beverage industries; in industries, it has been applied in final cleaning, in the rinsing water of CIP (Clean-in-Place)

The pulsed electric field (PEF) technology, also known as high-intensity electric field pulses

between two electrodes [22]. The inactivation of pathogenic and spoilage bacteria, yeasts, and some enzymes related to food quality have been demonstrated by several authors, although

) in a product situated

Effects of Radiation Technologies on Food Nutritional Quality

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141

ization processes, besides not generating any type of ionizing radiation [21].

systems and in the disinfection of packaging surfaces and storage tanks [20].

(HELP), refers to the application of high voltage pulses (20–80 kV cm−<sup>1</sup>

**2.3. Pulsed electric field**

**Figure 2.** Outline of pulsed electric field.

The use of microwave energy has been reported in the literature as a method used for improving microbiological quality and extending shelf life. The use of microwave irradiation resulted in the conclusion that microbial cell death occurs due to the heat produced by the irradiation and due to the electric field created by the microwaves, which promotes a change in the secondary and tertiary structure of the microorganism proteins [12, 14]. Currently, microwave energy has been applied in the pharmaceutical industry, and in drying, thawing and sterilization processes in the food industry.

Despite the limitations of this technology, the application of microwave energy has advantages in the decontamination of utensils and in the process time, which entails the reduction of energy costs and also eliminates the chemicals commonly used in some of the conventional methods. All these reasons make this technology economically and environmentally viable, to be implemented in industrial systems, provided that its limitations are reviewed, such as the lack of uniformity in the distribution of heating in systems without temperature and power control [14, 15].

#### **2.2. Ultraviolet radiation**

Ultraviolet (UV) radiation was discovered in 1801 by the German scientist Johan Ritter, who perceived an invisible form of light beyond violet capable of oxidizing silver halides, which was named ultraviolet light at the end of the nineteenth century [16]. The UV-light occupies a wide wavelength range in the nonionizing region of the electromagnetic spectrum, between X-rays (100 nm) and visible light (400 nm) [17]. The wavelength of UV-light is divided into three bands—UV-A, UV-B, and UV-C.

The long (UV-A, 400–320 nm) and middle (UV-B, 320–280 nm) wavelengths are present in sunlight and have some germicidal value. However, the short wavelengths or UV-C (280–100 nm) have high germicidal capacity and do not naturally exist, having to be produced by the conversion of electric energy [18].

The UV-light can be applied as a continuous or pulsed mode. The continuous mode is the conventional method, the light being emitted continuously without interruption. In pulsed UV-light mode, the UV-light is released as intermittent pulses using a capacitor, which allow to increase the energy intensity per pulse. Therefore, the pulsed mode is more effective for microbiological inactivation and the most used method [19].

The use of UV-light as a nonchemical disinfection method is increasingly gaining acceptance by its germicidal power. The equipment used in the UV method has a low maintenance cost, besides being considered an "eco-friendly" technology that eliminates the need to use many chemical treatments, while is able to guarantee high levels of security [20].

UV-light is a physical process that offers several advantages, such as it does not produce by-products that could change the characteristics of the food, does not generate chemical residues, and it is a cold process, dry, simple, effective, and low cost in relation to other sterilization processes, besides not generating any type of ionizing radiation [21].

UV-light has been applied in water purification and has been used in beverage industries; in industries, it has been applied in final cleaning, in the rinsing water of CIP (Clean-in-Place) systems and in the disinfection of packaging surfaces and storage tanks [20].

### **2.3. Pulsed electric field**

penetrates superficially in the foods, and the depth of penetration ranges from 2 to 4 cm. The process makes the food vibrates and consequently heating the water, fat, and sugar molecules. However, microwave heating leads to uneven distribution of the temperature in the product. In addition, the products do not show nonenzymatic darkening and surface crust

The use of microwave energy has been reported in the literature as a method used for improving microbiological quality and extending shelf life. The use of microwave irradiation resulted in the conclusion that microbial cell death occurs due to the heat produced by the irradiation and due to the electric field created by the microwaves, which promotes a change in the secondary and tertiary structure of the microorganism proteins [12, 14]. Currently, microwave energy has been applied in the pharmaceutical industry, and in drying, thawing and steriliza-

Despite the limitations of this technology, the application of microwave energy has advantages in the decontamination of utensils and in the process time, which entails the reduction of energy costs and also eliminates the chemicals commonly used in some of the conventional methods. All these reasons make this technology economically and environmentally viable, to be implemented in industrial systems, provided that its limitations are reviewed, such as the lack of uniformity in the distribution of heating in systems without temperature and power

Ultraviolet (UV) radiation was discovered in 1801 by the German scientist Johan Ritter, who perceived an invisible form of light beyond violet capable of oxidizing silver halides, which was named ultraviolet light at the end of the nineteenth century [16]. The UV-light occupies a wide wavelength range in the nonionizing region of the electromagnetic spectrum, between X-rays (100 nm) and visible light (400 nm) [17]. The wavelength of UV-light is divided into

The long (UV-A, 400–320 nm) and middle (UV-B, 320–280 nm) wavelengths are present in sunlight and have some germicidal value. However, the short wavelengths or UV-C (280–100 nm) have high germicidal capacity and do not naturally exist, having to be pro-

The UV-light can be applied as a continuous or pulsed mode. The continuous mode is the conventional method, the light being emitted continuously without interruption. In pulsed UV-light mode, the UV-light is released as intermittent pulses using a capacitor, which allow to increase the energy intensity per pulse. Therefore, the pulsed mode is more effective for

The use of UV-light as a nonchemical disinfection method is increasingly gaining acceptance by its germicidal power. The equipment used in the UV method has a low maintenance cost, besides being considered an "eco-friendly" technology that eliminates the need to use many

formation, which might be advantageous in some applications [10].

tion processes in the food industry.

control [14, 15].

140 Descriptive Food Science

**2.2. Ultraviolet radiation**

three bands—UV-A, UV-B, and UV-C.

duced by the conversion of electric energy [18].

microbiological inactivation and the most used method [19].

chemical treatments, while is able to guarantee high levels of security [20].

The pulsed electric field (PEF) technology, also known as high-intensity electric field pulses (HELP), refers to the application of high voltage pulses (20–80 kV cm−<sup>1</sup> ) in a product situated between two electrodes [22]. The inactivation of pathogenic and spoilage bacteria, yeasts, and some enzymes related to food quality have been demonstrated by several authors, although

**Figure 2.** Outline of pulsed electric field.

bacterial spore is not eliminated through this treatment [23, 24]. Regarding to this limitation, other methods are used in combination with irradiation technologies for the elimination of bacterial spores.

source) to produce gamma rays, or less frequently, by an equipment which emits high electron-beam energy [32, 33]. Gamma radiation is produced from the excited nucleus of radioac-

Effects of Radiation Technologies on Food Nutritional Quality

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143

The process of irradiation by gamma rays has several advantages: low heat generation, low energy requirements, irradiation of packaged or frozen products, and cause changes in the food nutritional value similar or inferior to other methods of conservation [36]. Nevertheless, as disadvantages of the gamma radiation are the high initial cost and difficulty in establishing the necessary doses, because certain doses can cause the death of some insects, however, others still can be found alive [37]. Despite all the benefits, a number of barriers still persist and prevent irradiated foods from being widely marketed, mainly related to the cost of their use and the resistance from the consumers due to a lack of information on food induced

In order to ensure the quality of a food, from the microbiological point of view and the nutritional quality, several methods are available for the food industries, including the use of radiation. In view of the requirements of consumers for assurance of microbiological safety and maintaining the natural characteristics of the product, the present chapter focuses on nutritional variation in food products submitted to radiation processes, such as gamma irra-

The radiation techniques have been widely studied and, like most food processing techniques, can induce some changes that are able to modify the chemical and nutritional characteristics on foods. These changes are dependent on some factors such as the radiation dose, the constitution of the irradiated food, the type of packaging, and how it was processed, besides the variables of the process as temperature and oxygen saturation on the

Activists and consumers have been questioned about the nutritional value of foods submitted to these processing techniques. The main questions and hypotheses about the use of irradiation are related to the excessive denaturation of nutrients, the conception that there is generation of toxic substances in irradiated foods, and they became radioactive [39, 40]. However, research conducted since the 1950s does not confirm these hypotheses of radioactivity in food, demonstrating there is no induction [21, 41]. Thus, the radiation methods used in food are considered safe and effective, according to several agencies such as the Food and Drug

Among the advantages cited by studies related to food irradiation are the minimal changes caused in food constituents [44]. According to research, macronutrients—carbohydrates, proteins, and lipids—are relatively stable when they come in contact with doses of ionizing

Regarding to micronutrients, in particular vitamins are perceived since they are sensitive to any processing technique. Thus, vitamins are unstable in food and dependent on specific

tive elements, such as Cobalt 60 (60Co) and Cesium 137 (<sup>137</sup>Cs) [34, 35].

diation, UV-C, microwave, and pulsed electric fields.

**4. Effects of radiation on food nutritional composition**

Administration (FDA) and the United Nations (UN) [42, 43].

radioactivity [38].

atmospheric [39].

radiation up to 10 kGy [39, 45].

The technology consists of subjecting the product to repeated high-intensity fields with short-time electrical pulses, (ms or μs), with the purpose of causing the inactivation of enzymes and destruction of microorganisms [25]. The outline of pulsed electric field technology is shown in **Figure 2**.

Regarding food quality attributes, PEF is considered superior to conventional heat treatment because it avoids or reduces changes in sensory and physicochemical properties [26]. Many studies have reported the advantage of the application of PEF for the conservation of the micronutrient content of foods, once that is expect the thermolabile vitamins to be conserved, since the electric pulse method does not constitute a heat treatment. There are several factors that influence the results from the pulsed electric field on the microorganisms. These factors are related with the parameters of the process (time, field intensity, temperature, and number of pulses), to the characteristics of the product, and the characteristics of the microorganisms present in the product [27].

The application of PEF for food preservation has shown to be a promising alternative to traditional thermal methods. Although there are not products processed by pulsed electric fields on the market, the great number of works related to such technology and its advantages claim to believe that soon the market will experience the emergence of this and other nonthermal food technologies [28, 29].

## **3. Ionizing radiation**

Nowadays, the types of radiations commonly used in the treatment of foods are the ionizing radiations. Ionizing radiation is classified as ionizing because its energy is high enough to displace electrons from atoms and molecules and to convert them into electrical charges, called ions [5].

Ionizing radiation is transmitted by high-energy particles (alpha, protons, electrons, and neutrons) or electromagnetic waves (X-rays and gamma rays). Only electromagnetic radiation (X-rays and gamma rays) and electron-beam radiation are the two forms of ionizing radiations applied on food among the various existing forms of ionizing radiation. While the propagation of electromagnetic radiation is in the form of waves, the radiation by emission of electrons (corpuscular radiation) is based on the transmission of energy by the acceleration of the electrons [30, 31].

The irradiation of food, physical method of treatment, is based on submit of the product, already packaged or in bulk, to controlled doses of ionizing radiation for sanitary, phytosanitary and/or technological purposes [30].

#### **3.1. Gamma radiation**

The frequency of the gamma rays is above 1019 Hz, which implies in wavelengths below 10–12 m. Radiation equipment is composed of one high energy radiation source (isotope source) to produce gamma rays, or less frequently, by an equipment which emits high electron-beam energy [32, 33]. Gamma radiation is produced from the excited nucleus of radioactive elements, such as Cobalt 60 (60Co) and Cesium 137 (<sup>137</sup>Cs) [34, 35].

The process of irradiation by gamma rays has several advantages: low heat generation, low energy requirements, irradiation of packaged or frozen products, and cause changes in the food nutritional value similar or inferior to other methods of conservation [36]. Nevertheless, as disadvantages of the gamma radiation are the high initial cost and difficulty in establishing the necessary doses, because certain doses can cause the death of some insects, however, others still can be found alive [37]. Despite all the benefits, a number of barriers still persist and prevent irradiated foods from being widely marketed, mainly related to the cost of their use and the resistance from the consumers due to a lack of information on food induced radioactivity [38].

In order to ensure the quality of a food, from the microbiological point of view and the nutritional quality, several methods are available for the food industries, including the use of radiation. In view of the requirements of consumers for assurance of microbiological safety and maintaining the natural characteristics of the product, the present chapter focuses on nutritional variation in food products submitted to radiation processes, such as gamma irradiation, UV-C, microwave, and pulsed electric fields.

## **4. Effects of radiation on food nutritional composition**

bacterial spore is not eliminated through this treatment [23, 24]. Regarding to this limitation, other methods are used in combination with irradiation technologies for the elimination of

The technology consists of subjecting the product to repeated high-intensity fields with short-time electrical pulses, (ms or μs), with the purpose of causing the inactivation of enzymes and destruction of microorganisms [25]. The outline of pulsed electric field technology is shown in **Figure 2**. Regarding food quality attributes, PEF is considered superior to conventional heat treatment because it avoids or reduces changes in sensory and physicochemical properties [26]. Many studies have reported the advantage of the application of PEF for the conservation of the micronutrient content of foods, once that is expect the thermolabile vitamins to be conserved, since the electric pulse method does not constitute a heat treatment. There are several factors that influence the results from the pulsed electric field on the microorganisms. These factors are related with the parameters of the process (time, field intensity, temperature, and number of pulses), to the characteristics of the product, and the characteristics of the microorganisms

The application of PEF for food preservation has shown to be a promising alternative to traditional thermal methods. Although there are not products processed by pulsed electric fields on the market, the great number of works related to such technology and its advantages claim to believe that soon the market will experience the emergence of this and other nonthermal

Nowadays, the types of radiations commonly used in the treatment of foods are the ionizing radiations. Ionizing radiation is classified as ionizing because its energy is high enough to displace electrons from atoms and molecules and to convert them into electrical charges,

Ionizing radiation is transmitted by high-energy particles (alpha, protons, electrons, and neutrons) or electromagnetic waves (X-rays and gamma rays). Only electromagnetic radiation (X-rays and gamma rays) and electron-beam radiation are the two forms of ionizing radiations applied on food among the various existing forms of ionizing radiation. While the propagation of electromagnetic radiation is in the form of waves, the radiation by emission of electrons (corpuscular radiation) is based on the transmission of energy by the acceleration of

The irradiation of food, physical method of treatment, is based on submit of the product, already packaged or in bulk, to controlled doses of ionizing radiation for sanitary, phytosani-

The frequency of the gamma rays is above 1019 Hz, which implies in wavelengths below 10–12 m. Radiation equipment is composed of one high energy radiation source (isotope

bacterial spores.

142 Descriptive Food Science

present in the product [27].

food technologies [28, 29].

**3. Ionizing radiation**

called ions [5].

the electrons [30, 31].

**3.1. Gamma radiation**

tary and/or technological purposes [30].

The radiation techniques have been widely studied and, like most food processing techniques, can induce some changes that are able to modify the chemical and nutritional characteristics on foods. These changes are dependent on some factors such as the radiation dose, the constitution of the irradiated food, the type of packaging, and how it was processed, besides the variables of the process as temperature and oxygen saturation on the atmospheric [39].

Activists and consumers have been questioned about the nutritional value of foods submitted to these processing techniques. The main questions and hypotheses about the use of irradiation are related to the excessive denaturation of nutrients, the conception that there is generation of toxic substances in irradiated foods, and they became radioactive [39, 40]. However, research conducted since the 1950s does not confirm these hypotheses of radioactivity in food, demonstrating there is no induction [21, 41]. Thus, the radiation methods used in food are considered safe and effective, according to several agencies such as the Food and Drug Administration (FDA) and the United Nations (UN) [42, 43].

Among the advantages cited by studies related to food irradiation are the minimal changes caused in food constituents [44]. According to research, macronutrients—carbohydrates, proteins, and lipids—are relatively stable when they come in contact with doses of ionizing radiation up to 10 kGy [39, 45].

Regarding to micronutrients, in particular vitamins are perceived since they are sensitive to any processing technique. Thus, vitamins are unstable in food and dependent on specific parameters such as the cooking time or time of exposure to the method, pH, temperature, and among others. The water-soluble vitamins are labile to any process technique, and the fat-soluble vitamins are destroyed by the radiation methods [46–48].

method, there was no interference in the fiber contents, but it was observed that the samples that were produced by the method of organic cultivation with application of UV-C light

Effects of Radiation Technologies on Food Nutritional Quality

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145

The effects of different thermal treatment methods were conducted in beef and chicken burgers cooked in microwave, conventional oven, and fry in oil. Samples of the two types of burgers submitted to microwave process obtained the highest loss of moisture, weight, and greater degree of retraction. In the chicken burger, the loss of moisture resulted in higher percentages of fat, protein, and ashes. In the beef hamburger, the highest percentage of proteins and ashes were observed in the microwave treatment, while the highest percentage of lipids was found

Studies on the application of UV-C light to tilapia fillets demonstrated that this type of treatment was not able to increase lipid oxidation due to the low fat content in this type of food. Regarding the protein content, UV-C light provided an increase in the carbonyl content with changes directly related to the doses of applied radiation, presenting a pro-oxidant action [57]. Action that can be explained by the capacity of stimulus of generation and reactivity of oxygen species, causing the protein oxidation, raises carbonyl formation [51]. In the literature, the oxidative effect of proteins and lipids has a direct connection and might vary according to the protein and lipid composition and pro-oxidant and antioxidant compounds of a food [58, 59, 60, 61]. Therefore, these chemical modifications promoted by UV-C light

At commercial doses, gamma irradiation causes no higher nutrient loss than the other methods used in food processing. The destruction of vitamins from the use of ionizing radiation applying doses of up to 60 kGy does not differ from the degradation generated from the cooking process [63]. The degradation of vitamins presents varied sensitivity to gamma radiation, since this level depends on the doses used and the state of matter from the food analyzed. The fat-soluble vitamins in descending order of sensitivity to gamma radiation are Vitamin E > Vitamin A > Vitamin D > Vitamin K. While the order of sensitivity of water-soluble vitamins is thiamine > ascorbic acid > pyridoxine > riboflavin > cobalamin > nicotinic acid [50, 64]. However, studies related to the effects of radiation on the content of vitamins are still inconclusive. In order to minimize nutrition losses and negative effects on sensory quality, some factors are applied, such as the use of low doses (less than 10 kGy) of gamma radiation and controlled conditions of process such as temperature and presence

Among the nonthermal technologies used in food, UV-light treatment presents great commercial potential as an alternative for thermal treatments in liquid foods, such as pasteurization. However, as well the other methods used in food preservation, UV-light causes changes in the vitamin content. UV-light treatment has been shown to be able to significantly alter vitamin content in milk samples when compared to the traditional pasteurizing process. The vitamins present in the cow's milk in descending order of sensitivity are C > E > A > B2. However, the number of sample passes through the UV-C system and the initial vitamin concentration in a

sample are important factors to affect the level of loss in vitamin content [65].

depend on the nutritional composition and the dose to be used [62].

showed a higher protein content [55].

in the oil frying treatment [56].

**4.2. Micronutrients**

of oxygen [49, 50].

#### **4.1. Macronutrients**

When food is irradiated, its components may be subject to significant changes. Among the existing macronutrients, the carbohydrates are less sensitive to radiation. Some of the glycosidic bonds linking the monosaccharides break when exposed to radiation, reducing the degree of polymerization and subsequently increasing the viscosity of polysaccharide solutions.

Regarding proteins, the amino acid chains can be altered in the presence of water, due to electron transfer, and might accelerate the denaturation protein process by altering the secondary and tertiary structures before destroying the amino acid chains. Nevertheless, the denaturation is less intense than in a thermal processing. In the case of lipids, in the presence of oxygen, radiation processes can accelerate oxidation, and other reactions such as the production of free radicals, the formation of hydrogen peroxides, and the destruction of compounds as carboxylic acids and antioxidants. Thus, radiation processes are not recommended for products with a high fat content [49, 50].

The effects of UV-C on the composition of macronutrients in eggs were evaluated, demonstrating that cholesterol is oxidized to cholesterol oxides in contact with UV-C light, presenting reduction in doses higher than 5.910 J cm−<sup>2</sup> . In addition, UV-C light seems to be able to generate vitamin D3 (cholecalciferol), from the precursor 7-dehydrocholesterol. However, further testing may be required to identify oxidation products and to evaluate the production of vitamin D3 [51]. Unlike the results found for UV-C light applied in eggs, in pasteurization, cholesterol levels remained almost unchanged in the temperature range of 110–120°C, and less than 10% was oxidized after 80 hours of heating, corroborating with the fact that cholesterol is quite resistant to heat up to that temperature. However, cholesterol levels are extensively oxidized by exceeding the temperature range [52]. In relation to the protein content, it was observed that in the presence of oxygen, the total amount of protein decreased, occurring an aggregation of the proteins, and after the UV-C treatment, this situation indicates that some sulfhydryl groups were oxidized to the formation of new disulfide bonds [51].

Analyzing the effects of UV-C light on cow's milk and comparing the results with the pasteurization treatments, it is possible to visualize there was no change in fat concentration. On the protein content, there were no significant changes when compared to the samples treated with ultraviolet against the raw milk [53]. Therefore, it is suggested that ultraviolet radiation is an alternative to traditional heat treatments, for better preserving the nutrients, unlike pasteurization that causes nutrient loss. The effects of gamma radiation on milk proteins demonstrate that solubility decreases, probably due to the denaturation and agglomeration of proteins [54].

A study was conducted on the effects of agricultural production systems (organic and conventional), UV-C radiation and different types of drying (greenhouse and lyophilization) in grape waste and flour formulated from the wastes. The results showed, regardless of the method, there was no interference in the fiber contents, but it was observed that the samples that were produced by the method of organic cultivation with application of UV-C light showed a higher protein content [55].

The effects of different thermal treatment methods were conducted in beef and chicken burgers cooked in microwave, conventional oven, and fry in oil. Samples of the two types of burgers submitted to microwave process obtained the highest loss of moisture, weight, and greater degree of retraction. In the chicken burger, the loss of moisture resulted in higher percentages of fat, protein, and ashes. In the beef hamburger, the highest percentage of proteins and ashes were observed in the microwave treatment, while the highest percentage of lipids was found in the oil frying treatment [56].

Studies on the application of UV-C light to tilapia fillets demonstrated that this type of treatment was not able to increase lipid oxidation due to the low fat content in this type of food. Regarding the protein content, UV-C light provided an increase in the carbonyl content with changes directly related to the doses of applied radiation, presenting a pro-oxidant action [57]. Action that can be explained by the capacity of stimulus of generation and reactivity of oxygen species, causing the protein oxidation, raises carbonyl formation [51]. In the literature, the oxidative effect of proteins and lipids has a direct connection and might vary according to the protein and lipid composition and pro-oxidant and antioxidant compounds of a food [58, 59, 60, 61]. Therefore, these chemical modifications promoted by UV-C light depend on the nutritional composition and the dose to be used [62].

#### **4.2. Micronutrients**

parameters such as the cooking time or time of exposure to the method, pH, temperature, and among others. The water-soluble vitamins are labile to any process technique, and the

When food is irradiated, its components may be subject to significant changes. Among the existing macronutrients, the carbohydrates are less sensitive to radiation. Some of the glycosidic bonds linking the monosaccharides break when exposed to radiation, reducing the degree of polymerization and subsequently increasing the viscosity of polysaccharide

Regarding proteins, the amino acid chains can be altered in the presence of water, due to electron transfer, and might accelerate the denaturation protein process by altering the secondary and tertiary structures before destroying the amino acid chains. Nevertheless, the denaturation is less intense than in a thermal processing. In the case of lipids, in the presence of oxygen, radiation processes can accelerate oxidation, and other reactions such as the production of free radicals, the formation of hydrogen peroxides, and the destruction of compounds as carboxylic acids and antioxidants. Thus, radiation processes are not recommended

The effects of UV-C on the composition of macronutrients in eggs were evaluated, demonstrating that cholesterol is oxidized to cholesterol oxides in contact with UV-C light,

be able to generate vitamin D3 (cholecalciferol), from the precursor 7-dehydrocholesterol. However, further testing may be required to identify oxidation products and to evaluate the production of vitamin D3 [51]. Unlike the results found for UV-C light applied in eggs, in pasteurization, cholesterol levels remained almost unchanged in the temperature range of 110–120°C, and less than 10% was oxidized after 80 hours of heating, corroborating with the fact that cholesterol is quite resistant to heat up to that temperature. However, cholesterol levels are extensively oxidized by exceeding the temperature range [52]. In relation to the protein content, it was observed that in the presence of oxygen, the total amount of protein decreased, occurring an aggregation of the proteins, and after the UV-C treatment, this situation indicates that some sulfhydryl groups were oxidized to the formation of new disulfide

Analyzing the effects of UV-C light on cow's milk and comparing the results with the pasteurization treatments, it is possible to visualize there was no change in fat concentration. On the protein content, there were no significant changes when compared to the samples treated with ultraviolet against the raw milk [53]. Therefore, it is suggested that ultraviolet radiation is an alternative to traditional heat treatments, for better preserving the nutrients, unlike pasteurization that causes nutrient loss. The effects of gamma radiation on milk proteins demonstrate that solubility decreases, probably due to the denaturation and agglomeration of proteins [54]. A study was conducted on the effects of agricultural production systems (organic and conventional), UV-C radiation and different types of drying (greenhouse and lyophilization) in grape waste and flour formulated from the wastes. The results showed, regardless of the

. In addition, UV-C light seems to

fat-soluble vitamins are destroyed by the radiation methods [46–48].

**4.1. Macronutrients**

144 Descriptive Food Science

for products with a high fat content [49, 50].

presenting reduction in doses higher than 5.910 J cm−<sup>2</sup>

solutions.

bonds [51].

At commercial doses, gamma irradiation causes no higher nutrient loss than the other methods used in food processing. The destruction of vitamins from the use of ionizing radiation applying doses of up to 60 kGy does not differ from the degradation generated from the cooking process [63]. The degradation of vitamins presents varied sensitivity to gamma radiation, since this level depends on the doses used and the state of matter from the food analyzed. The fat-soluble vitamins in descending order of sensitivity to gamma radiation are Vitamin E > Vitamin A > Vitamin D > Vitamin K. While the order of sensitivity of water-soluble vitamins is thiamine > ascorbic acid > pyridoxine > riboflavin > cobalamin > nicotinic acid [50, 64]. However, studies related to the effects of radiation on the content of vitamins are still inconclusive. In order to minimize nutrition losses and negative effects on sensory quality, some factors are applied, such as the use of low doses (less than 10 kGy) of gamma radiation and controlled conditions of process such as temperature and presence of oxygen [49, 50].

Among the nonthermal technologies used in food, UV-light treatment presents great commercial potential as an alternative for thermal treatments in liquid foods, such as pasteurization. However, as well the other methods used in food preservation, UV-light causes changes in the vitamin content. UV-light treatment has been shown to be able to significantly alter vitamin content in milk samples when compared to the traditional pasteurizing process. The vitamins present in the cow's milk in descending order of sensitivity are C > E > A > B2. However, the number of sample passes through the UV-C system and the initial vitamin concentration in a sample are important factors to affect the level of loss in vitamin content [65].

The minerals present in foods (i.e., iron, phosphorus, and calcium) are not affected by technologies that employ radiation [42]. No more data were reported in the literature relating the effects of irradiation on vitamins and minerals, and more studies are needed.

macronutrients and minerals are stable against radiation techniques. Vitamins have a higher degree of sensitivity than macronutrients when exposed to treatments that use some type of radiation. However, nutrient degradation can be reduced by using lower radiation doses and

Effects of Radiation Technologies on Food Nutritional Quality

http://dx.doi.org/10.5772/intechopen.80437

147

HELP technology, also known as high-intensity electric field pulses

Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de Souza\*

Universidade Federal dos Vales do Jequitinhonha e Mucuri/Universidade Federal de Lavras,

[1] Tezotto-Uliana JV, Silva PPM, Kluge RA, Spoto MHF. Radiação gama em produtos de origem vegetal. Revista Virtual de Química. 2015;**7**:267-277. DOI: 10.5935/1984-6835.20150013

[2] Leinstner L, Gorris LGM. Food preservation by hurdle technology. Trends in Food

[3] Vega-Mercado H et al. Non-thermal food preservation: Pulsed electric fields. Trends in Food Science & Technology. 1997;**8**:151-157. DOI: 10.1016/S0924-2244(97)01016-9

[4] Polizel GG. O uso da radiação no controle microbiológico dos alimentos de origem ani-

[5] Franco BDGM, Landgraf M. Microbiologia dos Alimentos. São Paulo: Atheneu; 1996. p. 182

[6] Spolaore AJG, Germano PML, Germano MIS.Irradiação de Alimentos. In: Germano PML, Germano MIS, editors. Higiene e Vigilância Sanitária de Alimentos. 2nd ed. São Paulo:

Science & Technology. 1995;**6**:41-45. DOI: 10.1016/S0924-2244(00)88941-4

mal [monografia]. Rio de Janeiro: Universidade Castelo Branco; 2006

Livraria Varela; 2001. pp. 421-442

\*Address all correspondence to: poliana.souza@ict.ufvjm.edu.br

using an oxygen saturated atmosphere.

**Acronyms and abbreviations**

PEF pulsed electric field

UN United Nations

UV ultraviolet

**Author details**

Diamantina, Brazil

**References**

FDA Food and Drug Administration

CIP clean-in-place

#### **4.3. Antioxidant activity**

The relationship between the treatments that use radiation and the antioxidant content of foods is still inconclusive. Several studies discuss that radiation treatments have demonstrated the decrease or increase in antioxidant activity of foods, and these variations are related to the type of treatment, the dose of radiation used, the exposure time, and the composition and state of matter of the raw material [66].

With positive effect, it is proven that the ultraviolet radiation, when applied in appropriate conditions, is able to increase the antioxidant capacity of fruits and vegetables, being a technology with commercial potential to be used in post-harvest stages in order to extend shelf life and increase quality, such as papaya, grape, and pineapple [67, 68]. Supposedly, UV-C radiation is able to increase phenolic content and antioxidant enzymatic activities as a defense mechanism against oxidative stress, thus increasing the antioxidant activity of vegetables [67, 69, 70].

In commercial doses, it is not possible to obtain conclusive answers regarding the effect of gamma radiation on the content of the antioxidant activity in foods. However, most of the studies mention gamma radiation as a conservation method related to the increase or maintenance to the potential of the antioxidant activity of foods such as, soybean, olive, starch, and safflower powder [66, 71–74]. However, it is possible to find studies that report the reduction of antioxidant activity when the radiation process is mediated with another parameter, as changes in moisture and application of different doses of gamma radiation in mung bean [75].

#### **4.4. Food allergens**

Food processing methods using radiation, ionizing, and nonionizing can be used to reduce the potential of food allergenicity. The UV-light, gamma irradiation, and ultrasound have the capacity to reduce the allergenicity of food proteins from the formation of protein aggregation, protein crosslinking, and/or amino acid sequence alteration. Studies carried out with the application of gamma radiation on cow's milk proteins have demonstrated the reduction of the allergenicity and antigenicity of the proteins when submitted to the irradiation process [54]. However, to date, the processes that involve radiation are not capable of completely inactivating the allergenic compounds [76].

## **5. Conclusions**

The technologies that involve radiation are pointed out as promising methods to be used in food preservation. Some methods used cause the elimination or decrease the growth of microorganisms, without significant nutritional losses. When applied in a controlled manner, the radiation processes promote equal or inferior damages in the nutritional quality of the food when compared to the conventional processes. At commercial doses of up to 10 kGy, macronutrients and minerals are stable against radiation techniques. Vitamins have a higher degree of sensitivity than macronutrients when exposed to treatments that use some type of radiation. However, nutrient degradation can be reduced by using lower radiation doses and using an oxygen saturated atmosphere.

## **Acronyms and abbreviations**


## **Author details**

The minerals present in foods (i.e., iron, phosphorus, and calcium) are not affected by technologies that employ radiation [42]. No more data were reported in the literature relating the

The relationship between the treatments that use radiation and the antioxidant content of foods is still inconclusive. Several studies discuss that radiation treatments have demonstrated the decrease or increase in antioxidant activity of foods, and these variations are related to the type of treatment, the dose of radiation used, the exposure time, and the composition and

With positive effect, it is proven that the ultraviolet radiation, when applied in appropriate conditions, is able to increase the antioxidant capacity of fruits and vegetables, being a technology with commercial potential to be used in post-harvest stages in order to extend shelf life and increase quality, such as papaya, grape, and pineapple [67, 68]. Supposedly, UV-C radiation is able to increase phenolic content and antioxidant enzymatic activities as a defense mechanism against oxidative stress, thus increasing the antioxidant activity of vegetables [67, 69, 70].

In commercial doses, it is not possible to obtain conclusive answers regarding the effect of gamma radiation on the content of the antioxidant activity in foods. However, most of the studies mention gamma radiation as a conservation method related to the increase or maintenance to the potential of the antioxidant activity of foods such as, soybean, olive, starch, and safflower powder [66, 71–74]. However, it is possible to find studies that report the reduction of antioxidant activity when the radiation process is mediated with another parameter, as changes in moisture and application of different doses of gamma radiation in mung bean [75].

Food processing methods using radiation, ionizing, and nonionizing can be used to reduce the potential of food allergenicity. The UV-light, gamma irradiation, and ultrasound have the capacity to reduce the allergenicity of food proteins from the formation of protein aggregation, protein crosslinking, and/or amino acid sequence alteration. Studies carried out with the application of gamma radiation on cow's milk proteins have demonstrated the reduction of the allergenicity and antigenicity of the proteins when submitted to the irradiation process [54]. However, to date, the processes that involve radiation are not capable of completely

The technologies that involve radiation are pointed out as promising methods to be used in food preservation. Some methods used cause the elimination or decrease the growth of microorganisms, without significant nutritional losses. When applied in a controlled manner, the radiation processes promote equal or inferior damages in the nutritional quality of the food when compared to the conventional processes. At commercial doses of up to 10 kGy,

effects of irradiation on vitamins and minerals, and more studies are needed.

**4.3. Antioxidant activity**

146 Descriptive Food Science

**4.4. Food allergens**

**5. Conclusions**

inactivating the allergenic compounds [76].

state of matter of the raw material [66].

Fabiana Lima, Kássia Vieira, Miriam Santos and Poliana Mendes de Souza\*

\*Address all correspondence to: poliana.souza@ict.ufvjm.edu.br

Universidade Federal dos Vales do Jequitinhonha e Mucuri/Universidade Federal de Lavras, Diamantina, Brazil

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2013.09.006

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**Chapter 8**

**Provisional chapter**

**Ultrasound Application to Improve Meat Quality**

**Ultrasound Application to Improve Meat Quality**

DOI: 10.5772/intechopen.77973

High-intensity ultrasound offers an alternative to traditional methods of food preservation and is regarded as a green, versatile, popular, and promising emerging technology. Ultrasound generates acoustic cavitation in a liquid medium, developing physical forces that are considered the main mechanism responsible for the observed changes in exposed materials. In meat, ultrasound has been successfully used to improve processes such as mass transfer and marination, tenderization of meat ,and inactivation of microorganisms. It is also an alternative to traditional meat aging methods for improving the quality properties of meat. Moreover, the combination of ultrasonic energy with a sanitizing agent can improve the effect of microbial reduction in foods. This review describes recent potential applications of ultrasound in meat systems, as well as physical and chemical effects of ultrasound treatments on the conservation and modification of processed meat foods. **Keywords:** ultrasound, cavitation, emerging technology, minimal processing, meat

Evolution of food processes is driven by changes in consumer preferences and the need to produce safe and high-quality foods. Nonthermal or intermediate technologies have great potential to achieve the characteristics desired by both the industry and consumers, especially regarding the desire to avoid altering the flavor or nutritional content during production. These technologies, which include the use of high pressure, electrical pulses, microfiltration,

> © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

Alma D. Alarcon-Rojo, Esmeralda Peña-González,

Alma D. Alarcon-Rojo, Esmeralda Peña-González,

Mariana Huerta-Jiménez, Raúl Reyes-Villagrana and

Mariana Huerta-Jiménez, Raúl Reyes-Villagrana and

Iván García-Galicia, Luis Carrillo-López,

Iván García-Galicia, Luis Carrillo-López,

http://dx.doi.org/10.5772/intechopen.77973

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

Hector Janacua-Vidales

Hector Janacua-Vidales

**Abstract**

quality

**1. Introduction**


#### **Ultrasound Application to Improve Meat Quality Ultrasound Application to Improve Meat Quality**

DOI: 10.5772/intechopen.77973

Alma D. Alarcon-Rojo, Esmeralda Peña-González, Iván García-Galicia, Luis Carrillo-López, Mariana Huerta-Jiménez, Raúl Reyes-Villagrana and Hector Janacua-Vidales Alma D. Alarcon-Rojo, Esmeralda Peña-González, Iván García-Galicia, Luis Carrillo-López, Mariana Huerta-Jiménez, Raúl Reyes-Villagrana and Hector Janacua-Vidales

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.77973

#### **Abstract**

[65] Guneser O, Yuceer YK. Effect of ultraviolet light on water- and fat-soluble vitamins in cow and goat milk. Journal of Dairy Science. 2012;**95**:6230-6241. DOI: 10.3168/jds.2011-5300

[66] Hussain PR et al. Influence of postharvest gamma irradiation treatment on the content of bioactive compound and antioxidant activity of fenegreek (*Trigonella foenum-graceum L.*) and spinach (*Spinacia oleracea L.*) leaves. Innovative Food Science & Emerging Techno-

[67] Rivera-Pastrana DM et al. Effect of UV-C irradiation and low temperature storage on bioactive compounds, antioxidant enzymes and radical scavenging activity of papaya fruit. Journal of Food Science and Technology. 2014;**51**(12):3821-3829. DOI: 10.1007/

[68] Jiang T. Influence of UV-C treatment on antioxidant capacity, antioxidant enzyme activity and texture of postharvest hiitake (*Lentinus edodes*) mushrooms during storage. Postharvest

[69] Sheng K. Comparison of postharvest UV-B and UV-C treatments on table grape: Changes in phenolic compounds and their transcription of biosynthetic genes during storage. Postharvest Biology and Technology. 2018;**128**:74-81. DOI: 10.1016/j.postharvbio.2018.

[70] Sari LK, Setha S, Naradisorn M. Effect of UV-C irradiation on postharvest quality of 'Phulae' pineapple. Scientia Horticulturae. 2016;**213**:314-320. DOI: 10.1016/j.scienta.2016.

[71] Popovic MB et al. Enhancement of antioxidant and isoflavones concentration in gamma irradiated soybean. The Scientific World Journal. 2013:383574. DOI: 10.1155/2013/383574

[72] Aouidi F et al. Gamma irradiation of air-dried olive leaves: Effective decontamination and impact on the antioxidative properties and on phenolic compounds. Food

[73] Mukhtar R et al. γ- Irradiation of oat grain—Effect on physici-chemical, structural, thermal, and antioxidant properties of extracted starch. International Journal of Biological

[74] Esmaeili S et al. Effect of gamma irradiation under various atmospheres of packing on the microbial and physicochemical properties of turmeric powder. Radiation Physics

[75] Verma M, Sharma P, Gour VS, Kothari SL. Moisture-mediated effects of γ-irradiation on antioxidant properties of mung bean (*Vigna radiate L.*) cultivars. Innovative Food Science

[76] Ekezie FC, Cheng J, Sun D. Effects of nonthermal food processing technologies on food allergens: A review of recent research advances. Trends in Food Science & Technology.

Chemistry. 2011;**127**:1105-1113. DOI: 10.1016/j.foodchem.2011.01.109

Macromolecules. 2017;**104**:1313-1320. DOI: 10.1016/j.ijbiomac.2017.05.092

and Chemistry. 2018;**148**:60-67. DOI: 10.1016/j.radphyschem.2018.02.028

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2018;**64**:12-25. DOI: 10.1016/j.tifs.2018.01.007

Biology and Technology. 2010;**56**:209-251. DOI: 10.1016/j.postharvbio.2010.01.011

logies. 2016;**33**:268-281. DOI: 10.1016/j.ifset.2015.11.017

s13197-013-0942-x

01.002

152 Descriptive Food Science

09.049

High-intensity ultrasound offers an alternative to traditional methods of food preservation and is regarded as a green, versatile, popular, and promising emerging technology. Ultrasound generates acoustic cavitation in a liquid medium, developing physical forces that are considered the main mechanism responsible for the observed changes in exposed materials. In meat, ultrasound has been successfully used to improve processes such as mass transfer and marination, tenderization of meat ,and inactivation of microorganisms. It is also an alternative to traditional meat aging methods for improving the quality properties of meat. Moreover, the combination of ultrasonic energy with a sanitizing agent can improve the effect of microbial reduction in foods. This review describes recent potential applications of ultrasound in meat systems, as well as physical and chemical effects of ultrasound treatments on the conservation and modification of processed meat foods.

**Keywords:** ultrasound, cavitation, emerging technology, minimal processing, meat quality

#### **1. Introduction**

Evolution of food processes is driven by changes in consumer preferences and the need to produce safe and high-quality foods. Nonthermal or intermediate technologies have great potential to achieve the characteristics desired by both the industry and consumers, especially regarding the desire to avoid altering the flavor or nutritional content during production. These technologies, which include the use of high pressure, electrical pulses, microfiltration,

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

and ultrasonication, are especially designed for economy, simplicity, and energy efficiency. Ultrasound is an acoustic energy [1], and therefore, it is a nonionizing, noninvasive, and nonpolluting form of mechanical energy [2]. These properties lead to a wide range of applications in the food industry. It is considered an emerging method with a great potential to control, improve, and accelerate processes without damaging the quality of food and other products [3, 4]. A low-power and high-frequency method is used to monitor the composition and physicochemical properties of food components and products during processing. Therefore, it contributes to control the properties that improve food quality. In recent years, research studies have been focused on assessing the effects of ultrasound on processes including mass transfer or marinating, meat tenderizing, crystallization, freezing, drying, degassing, filtration, foam production and reduction, emulsification, homogenization, and inactivation of microorganisms and enzymes [2]. Ultrasound has also been employed to optimize physicochemical characteristics, preparation processes for meat products, microbiological content, and sensory characteristics in fresh and processed meat [5]. Although ultrasound waves have been used to improve a wide variety of characteristics for a variety of matrices and processes, the appropriate conditions for scaling ultrasonic methods up to industrial levels have been established for a relatively small number of processes [4].

are used to apply ultrasound to products: (a) direct application; (b) coupling to the device, and

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973 155

Low-intensity, high-frequency ultrasound has analytical applications that provide information about the physicochemical properties of foods such as composition, structure, and condition [10]. Furthermore, unlike conventional analytical techniques, it is noninvasive and nondestructive [11] and the measurements are fast, automated, and easy to use in both laboratories and production lines. High-power ultrasound, also known as high-intensity ultrasound, may cause changes in the physical, chemical, or mechanical properties of foods. In the field of biochemistry, ultrasound was initially used to rupture cell walls, releasing their contents. Subsequent studies showed that high-power ultrasound can be used to activate the immobilized enzymes by increasing the rate of transport of substrates to enzymes [12].

Power or high-intensity ultrasound has emerged as a new and complementary technology with a high number of potential applications. Its effects are primarily mechanical: alternate cycles of expansion and compression are produced, causing the growth or formation of new bubbles in the medium [13]. When they reach a volume in which they can no longer absorb more energy, the bubbles implode violently, causing microcurrents and the collapse of liquid molecules, a phenomenon known as cavitation. The quantity of energy released by the cavitation depends on the kinetics of the bubble's growth and collapse. This energy increases with increasing surface tension at the bubble interface and decreases with increasing vapor pressure of the liquid [14]. The results of ultrasound in liquid media depends on variables such as the characteristics of the treatment medium (viscosity, surface tension, vapor pressure, nature and concentration of dissolved gas, presence of solid particles, and temperature), efficiency of the ultrasound generator

(frequency and input power), and the size and geometry of the treatment container [15].

Ultrasound has potentially a wide range of applications in the food industry. Researchers have identified various areas in which ultrasound can be used effectively, such as in the modification and control of crystallization processes, liquid food degassing, enzyme inactivation, drying, filtration, and oxidation induction. [12].Ultrasound methods have also been used in emulsion preparation in fruit and vegetable dehydration, enzyme inhibition, microbial inactivation, and crystallization of fats and sugars. Another example of a successful application of ultrasound technology is acoustic drying. It can be performed at lower temperatures than those used in conventional methods because the heat transfer between a solid surface and a liquid surface increases by approximately 30–60%, reducing the probability of oxidation or degradation of the material [15]. In addition, studies have shown that ultrasound is an effective method for food freezing, and the acceleration of ice nucleation and freezing leads to better control of the crystal size distribution in frozen products [16]. Ultrasound can not only

(c) immersion in an ultrasound bath [2].

**3. Power ultrasound**

**4. Applications in food**

As emphasized by Chemat et al.[6], a key goal of ultrasound research is to study and analyze both desirable and undesirable degradation phenomena in foods resulting from ultrasonic treatment (e.g., ultrasonic processing may affect the texture and chemical composition of foods). For this reason, many research questions in the meat sector are yet to be elucidated. Although multiple reports have been published, still inconsistent results have been reported, maybe because of the specific nature of meat tissues and various factors of ultrasound application possibly involved in, affecting food properties. This review aims to identify the effect of ultrasound on the major quality characteristics of fresh meat. We believe the results will help establish a methodology to enable the scaling-up of ultrasonic technology to the industrial level.

## **2. Ultrasound overview**

Ultrasound is a form of energy generated by a longitudinal mechanical wave whose vibration frequency is greater than 20,000 cycles per second (20 kHz), which is above the audible limit for humans. Sound is considered a pressure wave with one-dimensional propagation. The speed of an ultrasonic pulse depends on the acoustic properties of the medium, and the speed of sound propagation is greater in solids than in liquids and higher in liquids than in gases [7]. In an ultrasound system, the electrical energy is transformed into vibrational energy, which is mechanical energy [8] that has been transmitted through a sonicated medium. Part of the input energy is lost through conversion to heat, and the rest can produce cavitation. A fraction of the cavitation energy produces chemical, physical, or biological effects, while other fractions are reflected and consumed in the reemission of sound. Ultrasound ranges from 20 kHz to 10 MHz and is divided into three categories: (1) high-power (>5 Wcm−2or 10–1000 Wcm−2) and lowfrequency (20–100 kHz); (2) medium-power and intermediate-frequency (100 kHz–1 MHz); and (3) low-power (<1 Wcm−2) and high-frequency (1–10 MHz) [9]. Three different methods are used to apply ultrasound to products: (a) direct application; (b) coupling to the device, and (c) immersion in an ultrasound bath [2].

Low-intensity, high-frequency ultrasound has analytical applications that provide information about the physicochemical properties of foods such as composition, structure, and condition [10]. Furthermore, unlike conventional analytical techniques, it is noninvasive and nondestructive [11] and the measurements are fast, automated, and easy to use in both laboratories and production lines. High-power ultrasound, also known as high-intensity ultrasound, may cause changes in the physical, chemical, or mechanical properties of foods. In the field of biochemistry, ultrasound was initially used to rupture cell walls, releasing their contents. Subsequent studies showed that high-power ultrasound can be used to activate the immobilized enzymes by increasing the rate of transport of substrates to enzymes [12].

## **3. Power ultrasound**

and ultrasonication, are especially designed for economy, simplicity, and energy efficiency. Ultrasound is an acoustic energy [1], and therefore, it is a nonionizing, noninvasive, and nonpolluting form of mechanical energy [2]. These properties lead to a wide range of applications in the food industry. It is considered an emerging method with a great potential to control, improve, and accelerate processes without damaging the quality of food and other products [3, 4]. A low-power and high-frequency method is used to monitor the composition and physicochemical properties of food components and products during processing. Therefore, it contributes to control the properties that improve food quality. In recent years, research studies have been focused on assessing the effects of ultrasound on processes including mass transfer or marinating, meat tenderizing, crystallization, freezing, drying, degassing, filtration, foam production and reduction, emulsification, homogenization, and inactivation of microorganisms and enzymes [2]. Ultrasound has also been employed to optimize physicochemical characteristics, preparation processes for meat products, microbiological content, and sensory characteristics in fresh and processed meat [5]. Although ultrasound waves have been used to improve a wide variety of characteristics for a variety of matrices and processes, the appropriate conditions for scaling ultrasonic methods up to industrial levels have been

As emphasized by Chemat et al.[6], a key goal of ultrasound research is to study and analyze both desirable and undesirable degradation phenomena in foods resulting from ultrasonic treatment (e.g., ultrasonic processing may affect the texture and chemical composition of foods). For this reason, many research questions in the meat sector are yet to be elucidated. Although multiple reports have been published, still inconsistent results have been reported, maybe because of the specific nature of meat tissues and various factors of ultrasound application possibly involved in, affecting food properties. This review aims to identify the effect of ultrasound on the major quality characteristics of fresh meat. We believe the results will help establish a methodology to enable the scaling-up of ultrasonic technology to the industrial level.

Ultrasound is a form of energy generated by a longitudinal mechanical wave whose vibration frequency is greater than 20,000 cycles per second (20 kHz), which is above the audible limit for humans. Sound is considered a pressure wave with one-dimensional propagation. The speed of an ultrasonic pulse depends on the acoustic properties of the medium, and the speed of sound propagation is greater in solids than in liquids and higher in liquids than in gases [7]. In an ultrasound system, the electrical energy is transformed into vibrational energy, which is mechanical energy [8] that has been transmitted through a sonicated medium. Part of the input energy is lost through conversion to heat, and the rest can produce cavitation. A fraction of the cavitation energy produces chemical, physical, or biological effects, while other fractions are reflected and consumed in the reemission of sound. Ultrasound ranges from 20 kHz to 10 MHz and is divided into three categories: (1) high-power (>5 Wcm−2or 10–1000 Wcm−2) and lowfrequency (20–100 kHz); (2) medium-power and intermediate-frequency (100 kHz–1 MHz); and (3) low-power (<1 Wcm−2) and high-frequency (1–10 MHz) [9]. Three different methods

established for a relatively small number of processes [4].

**2. Ultrasound overview**

154 Descriptive Food Science

Power or high-intensity ultrasound has emerged as a new and complementary technology with a high number of potential applications. Its effects are primarily mechanical: alternate cycles of expansion and compression are produced, causing the growth or formation of new bubbles in the medium [13]. When they reach a volume in which they can no longer absorb more energy, the bubbles implode violently, causing microcurrents and the collapse of liquid molecules, a phenomenon known as cavitation. The quantity of energy released by the cavitation depends on the kinetics of the bubble's growth and collapse. This energy increases with increasing surface tension at the bubble interface and decreases with increasing vapor pressure of the liquid [14]. The results of ultrasound in liquid media depends on variables such as the characteristics of the treatment medium (viscosity, surface tension, vapor pressure, nature and concentration of dissolved gas, presence of solid particles, and temperature), efficiency of the ultrasound generator (frequency and input power), and the size and geometry of the treatment container [15].

## **4. Applications in food**

Ultrasound has potentially a wide range of applications in the food industry. Researchers have identified various areas in which ultrasound can be used effectively, such as in the modification and control of crystallization processes, liquid food degassing, enzyme inactivation, drying, filtration, and oxidation induction. [12].Ultrasound methods have also been used in emulsion preparation in fruit and vegetable dehydration, enzyme inhibition, microbial inactivation, and crystallization of fats and sugars. Another example of a successful application of ultrasound technology is acoustic drying. It can be performed at lower temperatures than those used in conventional methods because the heat transfer between a solid surface and a liquid surface increases by approximately 30–60%, reducing the probability of oxidation or degradation of the material [15]. In addition, studies have shown that ultrasound is an effective method for food freezing, and the acceleration of ice nucleation and freezing leads to better control of the crystal size distribution in frozen products [16]. Ultrasound can not only increase the speed of freezing fresh foods, but also improve the quality of frozen products. Currently, the pasteurization and conventional thermal sterilization are the most commonly used techniques for removing the threat posed by microorganisms in food products. Heat treatment destroys the vegetative microorganisms and some spores; however, its effectiveness depends on the treatment temperature and time. The magnitude, temperature, and time of treatment are also proportional to nutrient loss, development of undesirable flavors, and deterioration of functional properties of food products [17, 18]. Studies have shown that high-power ultrasound substantially reduces microbial loads because cavitation disrupts cell walls, resulting in the destruction of living cells and thereby contributing to food preservation. Unfortunately, scarce is still known about the mechanism of inactivation.

**Sample Application (intensity/**

Beef (*semimembranosus*) 24 h postmortem and matured for 24,

Beef longissimus lumborum et thoracis and semitendinosus aged up to 8.5 days

Hen breast meat stored for 0, 1, 3,

Beef (*semitendinosus*) 1500 W, 40 kHz,

Beef *semitendinosus* 40 kHz, 11 w cm-2 for 0,

48, 72 or 96 h at 2°C

or 7 d at 4°C

Pork *biceps femoris 24 h post mortem*

Beef (*semimembranosus*) 24 h postmortem and matured for 24,

Beef *longissimus thoracic* and deep

Matured 14 d at 2°C Cooked at

Holstein bulls (*longissimus* 

48, 72 or 96 h at 2 °C

*pectoralis*

62°C or 70°C

*lumborum)*

**freq/time)**

to 240 s

period

12 W cm−2 , 24 kHz, for up

12 W cm−2, 24 kHz, 15 s

10, 20, 30, 40, 50, or 60 min

150 W, 1 MHz and 500 W,

25 kHz, 40 min

60 and 90 min.

10 min

1000 W, 20 kHz, 0, 5 or

20 kHz, 100 and 300 W for

10, 20 or 30 min

**Effect of ultrasound Authors**

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973

Stadnik et al. [31]

157

Jayasooriya et al. [34]

Xiong et al. [53]

Chang et al. [61]

Jørgensen et al. [60]

Stadnik et al. [31]

Caraveo et al. [21]

Pohlman et al. [44]

Barekat and Soltanizadeh [55]

Fragmentation of protein structures. Increase WHC.

Reduced WBS force and hardness. Increased pH. No interaction between ultrasound and aging. No changes in meat color and drip loss. Ultrasound reduced

cook and total loss.

red color.

yellow.

diameter.

collagen.

shear force.

alone.

collagen.

parameters.

2 W cm−2, 45kHz, 2 min Slightly less stable color.

Reduced shear force. No change in cooking loss.

No effect on brightness and

Decreased the tendency to

Decreased the muscle fiber

No effect on heat-insoluble

Weaken collagen stability.

Ultrasound did not change in

Ultrasound combined with actinidin decreased shear force more than actinidin

No change in oxidative stability at 4 d storage.

Increases luminosity and reduces redness up to 8 d of storage. No effect on water holding capacity of meat. Decreased coliforms and psychrophilic bacterial load.

Faster cooking, higher water retention, decreased cooking loss, shear force and soluble

Higher sensory tenderness.

Improved meat tenderness, decreased shear force, filtering residue and textural

2 W cm−2, 45 kHz, 2 min Acceleration of aging process.

## **5. Applications in meat**

The use of ultrasound in the meat industry, which began with the evaluation of live cattle fat and muscle, has been conducted since the 1950s. Nowadays, low-intensity ultrasound is routinely used to improve quality, taste, and tenderness, which represent the most important quality attributes in consumer satisfaction.

Many recent studies have reported potential uses of high-intensity ultrasound on fresh meat. Applications have been published with interesting advantages in freezing [19], thawing [20], meat brining [14], cooking [2], bacterial inhibition [21], and tenderizing [22]. The resulting changes of the application of ultrasound to fresh meat are summarized in **Table 1**.



increase the speed of freezing fresh foods, but also improve the quality of frozen products. Currently, the pasteurization and conventional thermal sterilization are the most commonly used techniques for removing the threat posed by microorganisms in food products. Heat treatment destroys the vegetative microorganisms and some spores; however, its effectiveness depends on the treatment temperature and time. The magnitude, temperature, and time of treatment are also proportional to nutrient loss, development of undesirable flavors, and deterioration of functional properties of food products [17, 18]. Studies have shown that high-power ultrasound substantially reduces microbial loads because cavitation disrupts cell walls, resulting in the destruction of living cells and thereby contributing to food preserva-

The use of ultrasound in the meat industry, which began with the evaluation of live cattle fat and muscle, has been conducted since the 1950s. Nowadays, low-intensity ultrasound is routinely used to improve quality, taste, and tenderness, which represent the most important

Many recent studies have reported potential uses of high-intensity ultrasound on fresh meat. Applications have been published with interesting advantages in freezing [19], thawing [20], meat brining [14], cooking [2], bacterial inhibition [21], and tenderizing [22]. The resulting

62 W cm−2, 20 kHz, 15 s No effect on mastication

2 W cm−2, 45kHz, 2 min No effect on meat color.

2 W cm−2, 45kHz, 2 min No effect on pH or color.

proteolysis.

**Effect of ultrasound Authors**

Lyng et al. [58]

Got et al. [30]

Dolatowski et al. [32]

Dolatowski and Stadnik [16]

Stadnik and Dolatowski [35]

force, sensory traits, solubility of collagen or myofibrillar

Larger sarcomeres, Z-line disruption, increased calcium. No effect on collagen.

Lower loss of water after cooling, thawing, and heating. No effect on pH. Higher water

Increased free calcium. Changes in protein structure. Improved WHC at 4 d

holding capacity.

*postmortem*.

Reduced hardness.

changes of the application of ultrasound to fresh meat are summarized in **Table 1**.

tion. Unfortunately, scarce is still known about the mechanism of inactivation.

**5. Applications in meat**

156 Descriptive Food Science

Beef (*longissimus thoracis* and *lumborum,* and *semimembranosus*)

*Semimembranosus pre- and* 

Beef (*semimembranosus*) matured for 24, 48, 72 or 96 h at 2°C

Beef (*semimembranosus*) 24 h postmortem and matured for 24,

48, 72 or 96 h at 2°C

*post-rigor*

quality attributes in consumer satisfaction.

**Sample Application (intensity/**

Beef (*semimembranosus*) 2 W cm−2, 25 kHz, 1 or

**freq/time)**

x15 s

2 min

10 W cm−2, 2.6 MHz, 2


**6. Effects on meat quality properties**

complementary or assistive technology [28].

as a result of ultrasound treatment (2.6 MHz, 10 W/cm<sup>2</sup>

**6.1. Potential of hydrogen (pH)**

**6.2. Water holding capacity (WHC)**

The majority of quality parameters assessed 24 h *postmortem* in meat is physicochemical in nature [23]. The potential of hydrogen (pH), water holding capacity (WHC), drip loss (DL), color (L\*, a\*, b\*, C\*, and h\*), and shear force are quality indicators for the raw meat. Physicochemical characteristics are evaluated to control the quality, assess the efficiency of production and treatment processing, compare results between laboratories, and aid research [24, 25]. The effect of a number of nutritional, breed, and production factors such as genetics, management, and slaughter on specific quality attributes should be considered when meat quality is analyzed [26]. Key markers of meat quality, including raw materials, which have a strong impact on the industry, are pH, water holding capacity, texture, and oxidative stability. In addition, aroma, color, flavor, and tenderness are the most important sensory components to the meat-consuming population [27]. When considering the use of emerging technologies in meat, both the mechanisms of action and the effects on food transformation, preservation, and integrity must be known. Ultrasound application may be an efficient alternative technology to increase meat tenderness. It is used in meat processing and preservation as a

pH is one of the most important indicators of raw meat quality, because it directly affects protein stability and properties. As reported in the literature, all product quality attributes depend on the ultimate pH. Water holding capacity, drip loss, and color are among the most important product quality attributes [29]. pH greatly affects the quality and functionality of muscle proteins, and decrease in pH reduces their water holding capacity, leading to economic losses. On the other hand, increases in pH increase the water holding capacity of the meat because of changes in the electrical charges within muscle proteins that occur when the pH is above the isoelectric point. Ultrasound effect on pH has been analyzed in various studies and conditions. Some authors have reported that initial pH of meat can be increased

differences in final pH, while other reported small differences [31] or no differences in pH because of ultrasound [32–35].In a study, bovine muscles (*longissimus lumborum et thoracis* and *semitendinosus*) sonicated (24 kHz, 12 Wcm−2) for a maximum of 4 min and subsequently stored them for 8 days led to increase tenderness and pH without a significant correlation between ultrasound and aging time [34]. The increase of pH was attributed to the release of ions from the cellular structure into the cytoplasm or to changes in protein structure, which could lead to changes in the position of ionic functionalities that could lower the muscle pH.

Water holding capacity (WHC) may be defined as the ability of meat or muscle proteins to immobilize their own or added water during an applied force [25]. A decrease in pH causes a shrink of the network of polypeptide chains, which decreases the water holding capacity.

) [30] prior to *rigor mortis*, with no

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973 159

**Table 1.** Effects of ultrasound on meat.

## **6. Effects on meat quality properties**

**Sample Application (intensity/**

Beef *longissimus dorsi* 40 kHz, 11 W cm-2 for

Chicken breast 22 W cm−2 40 kHz,

Pork *longissimus dorsi* 2-4 W cm−2, 20 kHz,

Pork *longissimus dorsi* 40 kHz; 37.5 W dm−3, 15,

Pork meat and skin surface High-intensity ultrasound,

Chicken wing surface 2.5 W cm−2, 40 kHz,

**Table 1.** Effects of ultrasound on meat.

Pure culture suspensions 20 kHz, 3, 6 or 9 min, 20,

0.5 to 2 s

3 or 6 min

40 and 60 °C

Chicken carcasses SonoSteam® *Campylobacter* and viable total

Chicken breast and soybean gels,

Pork *longissimus thoracis and* 

*lumborum*

4°C to 8°C

158 Descriptive Food Science

**freq/time)**

60 min.

s pulses)

15 or 30 min

450 W, 20 kHz,

0, 3, 6, 9 or 12 min (4 or 2

Pork *longissimus dorsi* 100 W and 20 kHz, 45 min Increased salt gain and water

30, 90 or 180 min

30, 45, 60, 90 or 120 min

4.2, 11 or 19 W cm−2, 20 kHz, 10, 25 or 40 min

Chicken breast Ultrasonic bath, 20 min No effect on water retention

**Effect of ultrasound Authors**

Peña-González et al.

Zhao et al. [54]

Leal-Ramos et al.

Cárcel et al. [14]

Siró et al. [15]

Ozuna et al. [63]

Smith et al. [43]

Kordowska-Wiater and Stasiak [82]

Herceg et al. [84]

Musavian et al. [83]

McDonnell et al. [59]

[22]

[62]

Reduces shear force. Produces more tender and juice meat. No effect on meat color.

Homogeneous fine network

Increased mass transfer and higher meat weight

Higher mass transfer at higher ultrasound intensity.

Higher salt diffusion. Diffusion coefficient increases with ultrasound intensity.

Higher salt and water

No effect on water holding capacity and structure of meat. Higher mass transfer and protein extraction. Myosin denaturation at higher

capacity, shear force and cooking loss. No changes in *Salmonella* and *E. coli*.

Microorganism reduction. Higher reduction with higher time. *E. coli* more sensible to

Bacteria inactivation is higher at higher time and

Less skin and surface bacteria Morild et al. [80]

diffusion.

intensities.

ultrasound.

temperature.

count reduction.

More viscoelastic gel Improved WFB and textural

properties

loss.

microstructures

The majority of quality parameters assessed 24 h *postmortem* in meat is physicochemical in nature [23]. The potential of hydrogen (pH), water holding capacity (WHC), drip loss (DL), color (L\*, a\*, b\*, C\*, and h\*), and shear force are quality indicators for the raw meat. Physicochemical characteristics are evaluated to control the quality, assess the efficiency of production and treatment processing, compare results between laboratories, and aid research [24, 25]. The effect of a number of nutritional, breed, and production factors such as genetics, management, and slaughter on specific quality attributes should be considered when meat quality is analyzed [26]. Key markers of meat quality, including raw materials, which have a strong impact on the industry, are pH, water holding capacity, texture, and oxidative stability. In addition, aroma, color, flavor, and tenderness are the most important sensory components to the meat-consuming population [27]. When considering the use of emerging technologies in meat, both the mechanisms of action and the effects on food transformation, preservation, and integrity must be known. Ultrasound application may be an efficient alternative technology to increase meat tenderness. It is used in meat processing and preservation as a complementary or assistive technology [28].

#### **6.1. Potential of hydrogen (pH)**

pH is one of the most important indicators of raw meat quality, because it directly affects protein stability and properties. As reported in the literature, all product quality attributes depend on the ultimate pH. Water holding capacity, drip loss, and color are among the most important product quality attributes [29]. pH greatly affects the quality and functionality of muscle proteins, and decrease in pH reduces their water holding capacity, leading to economic losses. On the other hand, increases in pH increase the water holding capacity of the meat because of changes in the electrical charges within muscle proteins that occur when the pH is above the isoelectric point. Ultrasound effect on pH has been analyzed in various studies and conditions. Some authors have reported that initial pH of meat can be increased as a result of ultrasound treatment (2.6 MHz, 10 W/cm<sup>2</sup> ) [30] prior to *rigor mortis*, with no differences in final pH, while other reported small differences [31] or no differences in pH because of ultrasound [32–35].In a study, bovine muscles (*longissimus lumborum et thoracis* and *semitendinosus*) sonicated (24 kHz, 12 Wcm−2) for a maximum of 4 min and subsequently stored them for 8 days led to increase tenderness and pH without a significant correlation between ultrasound and aging time [34]. The increase of pH was attributed to the release of ions from the cellular structure into the cytoplasm or to changes in protein structure, which could lead to changes in the position of ionic functionalities that could lower the muscle pH.

#### **6.2. Water holding capacity (WHC)**

Water holding capacity (WHC) may be defined as the ability of meat or muscle proteins to immobilize their own or added water during an applied force [25]. A decrease in pH causes a shrink of the network of polypeptide chains, which decreases the water holding capacity. Therefore, WHC is directly related to pH. The speed to reach the final pH also affects the WHC. When the drop in pH is relatively rapid, the changes in myofibrillar and sarcoplasmic proteins result in a decreased water holding capacity [29]. Some meat characteristics linked to WHC include color, texture, firmness, juiciness, and tenderness. Meat WHC is affected by factors including *rigor mortis*, ATP loss, and changes in the myofibrillar structure partly associated with proteolytic activity. Many other characteristics, including drip loss, are closely related to or depend on WHC.

the meat [44, 45]. The variability seen in the literature could result from differences in the ultrasound methods; the authors employed various times and intensities, which hindered

Color is a key factor in meat quality because it is the first sensory characteristic assessed by the consumer [46]. In red meat, a bright red color is related to freshness and therefore consumer rejection or acceptance [47]. Meat color results from the quantity and chemical state of myoglobin in the muscle. Deoxymyoglobin and myoglobin are responsible for the purple color of fresh meat. When meat is exposed to air for several minutes, deoxymyoglobin is oxygenated into oxymyoglobin, which is responsible for a cherry red color in meat. When meat is exposed to air for several hours or days, it turns brown due to the oxidation of oxymyoglobin into metmyoglobin. Meat contains other pigments, some derived from external sources, sometimes in insignificant amounts, which commonly indicate deterioration. Meat color and exterior appearance may be associated with aging time, shelf life, hardness, and juiciness. Some studies suggest that ultrasound has no effect on meat color because the heat generated is insufficient to denature proteins and pigments [48, 49]. Conversely,

the color changed to a lighter, less red, and more yellow-orange color (greater hue angle), which was less bright than control meat [43]. Ultrasound accelerates total changes in color, limits the formation of oxymyoglobin, and slows down the formation of metmyoglobin [48]. Nevertheless, when meat is cooked, meat panelists do not detect differences between ultra-

Tenderness in meat is determined by its texture. Tenderness is one of the most important attributes of meat quality because it is perhaps the most appreciated feature by consumers. Inconsistencies in this characteristic have been considered one of the major problems that the meat packing industry faces [50]. Tenderness is affected by the composition, structural organization, and integrity of the skeletal muscle. The two structural components that determine the intrinsic muscle strength are myofibrillar proteins and connective tissues [51] and the nature of these two components makes difficult to achieve tenderness. Tenderness depends on the size of the longitudinally arranged fiber bundles in muscles, which are delimited by the connective tissue septa forming the perimysium [25]. Myofibrillar tenderness can be controlled by manipulating conditions *pre-* and *postmortem*. Some methods and procedures used to increase tenderness include electrical stimulation, pressurization, calcium infusion, enzymatic treatment, and marination. All these methods are invasive, cause deformation, and affect the appearance of meat. In addition, some methods may contaminate the meat (e.g., brine injection with unclean needles). Currently, aging is the foremost industrial process used to increase the tenderness of meat. Aging tenderization mechanism is well known nowadays, consisting of biochemical processes driven by endogenous proteases. Nevertheless, aging is a time-consuming process, and it can be variable among animals. Therefore, various physical methods, such as electrical stimulation and chemical methods, have been used trying to

) on meat, it has been found that

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973 161

in an assessment of the effect of ultrasound (22 W/cm<sup>2</sup>

sonicated and control meat [22].

**6.5. Tenderness**

improve tenderness.

direct comparison.

**6.4. Color**

Approximately, three-quarter of meat is water, and about 10% of the water in live animal muscle is bound to muscle proteins compared to the 5–10% of water located in small channels between adjacent cells, or extracellular space. However, most of the water content is located in spaces between thin and thick filaments of myofibrils. In any muscle, WHC is minimal at low pH. Because of aging, it tends to increase owing to protein degradation and changes in electric charges induced by intramolecular reorganization [23]. When the WHC is low, moisture or weight loss during storage is greater owing to surface evaporation and exudation of cuts because the WHC is related to several physicochemical characteristics of the protein and myofibrillar components. WHC is the main indicator of the suitability of a given meat for preparing a product. The effects of ultrasonic treatment on WHC of meat are variable. Variation of effects on WHC is described on the next section, due to the relation between WHC and drip loss.

#### **6.3. Drip loss (DL)**

The release of water droplets from the muscle originated from the extracellular water is known as DL. It is the easiest water content to extract. DL depends on the state of contraction after *rigor mortis* because of reduction in the filamentary space and changes in the cellular membrane, which causes the release of water to the extracellular space in the form of drops through the cutting surfaces [35]. These drops consist of an aqueous red solution that largely contains proteins and water-soluble minerals, some of which are highly nutritious. Drip loss is strictly related to pH and WHC. When WHC increases, DL decreases and vice versa [36]. Several factors increase the WHC during meat aging, including pH, Z-line disintegration by protease activity, and changes in membrane permeability with diffusion and ionic redistribution, which results in substitution of divalent ions and weakening of intermolecular forces between protein chains. DL is primarily an economic problem for retailers, because weight losses during cutting cause accumulation of liquid around the product, which leads to consumer rejection [37, 38]. WHC is a key indicator of meat quality that affects the economic sector. Therefore, analyzing the effect of ultrasound on WHC is important [39]. Assessment of DL is used to identify the best conditions for the refrigeration, freezing, packaging, and storage conditions of meat. Consequently, DL measurements also make possible to determine WHC. The effects of ultrasonic treatments on WHC and drip loss are highly variable. Some authors report that ultrasound increases the rates of meat exudation and water loss [40, 41]. Whereas, other authors found no effect on the water holding capacity [42] or drip loss [21, 34] in beef (24–40 kHz, 11–12 W cm−2). In contrast, some reports indicate that ultrasonicated meat has a higher WHC [16, 40, 43], similar to that of meat at an advanced *postmortem* stage. This could be explained by structural changes in myofibrillar proteins caused by ultrasound; the above is confirmed by microstructure photographs [31]. Recently, when high-intensity ultrasound is applied during brining of beef, higher WHC was found, possibly by a higher diffusion of salt into the tissue, which can increase the capacity to hold the water before cooking the meat [44, 45]. The variability seen in the literature could result from differences in the ultrasound methods; the authors employed various times and intensities, which hindered direct comparison.

#### **6.4. Color**

Therefore, WHC is directly related to pH. The speed to reach the final pH also affects the WHC. When the drop in pH is relatively rapid, the changes in myofibrillar and sarcoplasmic proteins result in a decreased water holding capacity [29]. Some meat characteristics linked to WHC include color, texture, firmness, juiciness, and tenderness. Meat WHC is affected by factors including *rigor mortis*, ATP loss, and changes in the myofibrillar structure partly associated with proteolytic activity. Many other characteristics, including drip loss, are closely related to

Approximately, three-quarter of meat is water, and about 10% of the water in live animal muscle is bound to muscle proteins compared to the 5–10% of water located in small channels between adjacent cells, or extracellular space. However, most of the water content is located in spaces between thin and thick filaments of myofibrils. In any muscle, WHC is minimal at low pH. Because of aging, it tends to increase owing to protein degradation and changes in electric charges induced by intramolecular reorganization [23]. When the WHC is low, moisture or weight loss during storage is greater owing to surface evaporation and exudation of cuts because the WHC is related to several physicochemical characteristics of the protein and myofibrillar components. WHC is the main indicator of the suitability of a given meat for preparing a product. The effects of ultrasonic treatment on WHC of meat are variable. Variation of effects on WHC is described on the next section, due to the relation between WHC and drip loss.

The release of water droplets from the muscle originated from the extracellular water is known as DL. It is the easiest water content to extract. DL depends on the state of contraction after *rigor mortis* because of reduction in the filamentary space and changes in the cellular membrane, which causes the release of water to the extracellular space in the form of drops through the cutting surfaces [35]. These drops consist of an aqueous red solution that largely contains proteins and water-soluble minerals, some of which are highly nutritious. Drip loss is strictly related to pH and WHC. When WHC increases, DL decreases and vice versa [36]. Several factors increase the WHC during meat aging, including pH, Z-line disintegration by protease activity, and changes in membrane permeability with diffusion and ionic redistribution, which results in substitution of divalent ions and weakening of intermolecular forces between protein chains. DL is primarily an economic problem for retailers, because weight losses during cutting cause accumulation of liquid around the product, which leads to consumer rejection [37, 38]. WHC is a key indicator of meat quality that affects the economic sector. Therefore, analyzing the effect of ultrasound on WHC is important [39]. Assessment of DL is used to identify the best conditions for the refrigeration, freezing, packaging, and storage conditions of meat. Consequently, DL measurements also make possible to determine WHC. The effects of ultrasonic treatments on WHC and drip loss are highly variable. Some authors report that ultrasound increases the rates of meat exudation and water loss [40, 41]. Whereas, other authors found no effect on the water holding capacity [42] or drip loss [21, 34] in beef (24–40 kHz, 11–12 W cm−2). In contrast, some reports indicate that ultrasonicated meat has a higher WHC [16, 40, 43], similar to that of meat at an advanced *postmortem* stage. This could be explained by structural changes in myofibrillar proteins caused by ultrasound; the above is confirmed by microstructure photographs [31]. Recently, when high-intensity ultrasound is applied during brining of beef, higher WHC was found, possibly by a higher diffusion of salt into the tissue, which can increase the capacity to hold the water before cooking

or depend on WHC.

160 Descriptive Food Science

**6.3. Drip loss (DL)**

Color is a key factor in meat quality because it is the first sensory characteristic assessed by the consumer [46]. In red meat, a bright red color is related to freshness and therefore consumer rejection or acceptance [47]. Meat color results from the quantity and chemical state of myoglobin in the muscle. Deoxymyoglobin and myoglobin are responsible for the purple color of fresh meat. When meat is exposed to air for several minutes, deoxymyoglobin is oxygenated into oxymyoglobin, which is responsible for a cherry red color in meat. When meat is exposed to air for several hours or days, it turns brown due to the oxidation of oxymyoglobin into metmyoglobin. Meat contains other pigments, some derived from external sources, sometimes in insignificant amounts, which commonly indicate deterioration. Meat color and exterior appearance may be associated with aging time, shelf life, hardness, and juiciness. Some studies suggest that ultrasound has no effect on meat color because the heat generated is insufficient to denature proteins and pigments [48, 49]. Conversely, in an assessment of the effect of ultrasound (22 W/cm<sup>2</sup> ) on meat, it has been found that the color changed to a lighter, less red, and more yellow-orange color (greater hue angle), which was less bright than control meat [43]. Ultrasound accelerates total changes in color, limits the formation of oxymyoglobin, and slows down the formation of metmyoglobin [48]. Nevertheless, when meat is cooked, meat panelists do not detect differences between ultrasonicated and control meat [22].

#### **6.5. Tenderness**

Tenderness in meat is determined by its texture. Tenderness is one of the most important attributes of meat quality because it is perhaps the most appreciated feature by consumers. Inconsistencies in this characteristic have been considered one of the major problems that the meat packing industry faces [50]. Tenderness is affected by the composition, structural organization, and integrity of the skeletal muscle. The two structural components that determine the intrinsic muscle strength are myofibrillar proteins and connective tissues [51] and the nature of these two components makes difficult to achieve tenderness. Tenderness depends on the size of the longitudinally arranged fiber bundles in muscles, which are delimited by the connective tissue septa forming the perimysium [25]. Myofibrillar tenderness can be controlled by manipulating conditions *pre-* and *postmortem*. Some methods and procedures used to increase tenderness include electrical stimulation, pressurization, calcium infusion, enzymatic treatment, and marination. All these methods are invasive, cause deformation, and affect the appearance of meat. In addition, some methods may contaminate the meat (e.g., brine injection with unclean needles). Currently, aging is the foremost industrial process used to increase the tenderness of meat. Aging tenderization mechanism is well known nowadays, consisting of biochemical processes driven by endogenous proteases. Nevertheless, aging is a time-consuming process, and it can be variable among animals. Therefore, various physical methods, such as electrical stimulation and chemical methods, have been used trying to improve tenderness.

Numerous studies have been conducted to develop methods to improve tenderness. Among these, ultrasound application methods have been used at various sonication times, frequencies, and intensities. Most authors agree that ultrasound increases meat tenderness [22, 40, 48] and shortens the aging period without compromising other quality parameters [16, 33]. The potential of low-frequency, low-intensity ultrasound application to improve meat tenderization is remarkable. Several authors report an important reduction of shear force after treatment with ultrasound [52]. Benefits of ultrasound treatment on beef have been observed in *longissimus lumborum* and *semitendinosus* (24 kHz and 12 W/cm<sup>2</sup> for 240 s) [34], *M. Semimembranosus* (45 kHz and 2 W/cm<sup>2</sup> for 2 min) [48], and *semitendinosus* (40 kHz, 1500 W for 10, 20, 30, 40, 50 or 60 min) [40]. Benefits to the texture of poultry (24 kHz, 12 W/cm<sup>2</sup> for 4 min after 7 d of storage) [53, 54] and pork (2.5–3 W/cm<sup>2</sup> for 180 min) [15] are also reported. It has also been observed a significant decrease in the shear force of *Bovine L. dorsi* with the application of ultrasound, both, fresh and aged [22]. More recently, it was reported a reduction of shear force values in muscle semitendinosus when it was ultrasonicated and aged for 3–7 d [45]. The effect was mainly attributed to an increase of desmin and troponin-T degradation, and myofiber fracture along Z-lines and I-bands.

of power ultrasound on pork during wet marination depends on the ultrasound intensity applied [62]. Ultrasound causes bubble formation that hits the tissue, which may lead to microinjection of brine into the sample. This effect may help to explain the observed increase

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973 163

Ultrasonic treatment (low-intensity and low-frequency) and the use of vacuum caused favorable microstructural changes in pork loins marinated in sodium chloride [15] and these effects are highly dependent on the intensity of ultrasound treatment. Some of the critical factors in food processing warrant consideration because ultrasound generates rapid changes in temperature and pressure (109°C/s) over short time periods. Furthermore, cavitation generates shock waves, which contribute to this effect. Factors that modulate the effects of ultrasound application include time of exposure, processing volume, and sample composition [12, 14].

Bacteria are the most important microorganisms in food processing. While most are harmless and many are beneficial, some indicate the likely presence of contamination and deterioration and may cause diseases. While thousands of bacterial species have been identified, all are unicellular and fall under three basic forms: spherical, rod-shaped, and spiral. Some rod-shaped bacteria can take two forms: latent spores and active vegetative cells. The vegetative cells form spores under adverse conditions to survive. Most sporulating bacteria that grow in the presence of air belong to the *Bacillus* genus, and most of those that grow only in the absence

Meat is susceptible to the growth of some pathogenic microorganisms such as *E. coli, Campylobacter jejuni, Salmonella spp., Staphylococcus aureus,* and *Listeria monocytogenes*, which recurrently affect the properties of the meat and present serious problems during packing, processing, and storage. Several methods are used to avoid microbial growth in meat. The most commonly used methods involve heating, dehydration, and addition of preservatives [64]. The most common types of mesophilic bacteria that are pathogenic to humans include *Staphylococcus aureus*, *Salmonella*, and *Listeria*. Although it may survive without damage in the intestinal tract of humans, salmonella is a common cause of food poisoning. Another common mesophilic bacterium, *Listeria monocytogenes*, is more often distributed through contaminated foods such as raw meats or unpasteurized cheeses [64]. Animals, including humans, may transport *Listeria*, but it primarily threatens those with weakened immune systems. Some *E. coli* strains found in human feces are pathogenic, causing infection and disease. These are

*Staphylococcus* is nonsporulated bacteria without mobility, but because they are resistant to drying, they are easily dispersible by dust particles through air and surfaces [65]. *S. aureus* is usually found in the skin and in mucous membranes of humans and other animals. It is almost always present in small quantities in raw meats and foods extensively handled by humans. Maintaining food that is completely free of contamination with *Staphylococcus* is often difficult or impossible. Pasteurizing or cooking destroys the organism but not its toxin [66].Meat is one of the most perishable foods consumed by humans—it is easily damaged by bacteria. One of the most commonly used preservation methods is refrigeration, including freezing. However, certain bacteria are able to grow at 4°C; these are collectively known as psychrophiles. This

of NaCl content in the ultrasonicated meat [14, 63].

**6.7. Microbiological properties**

of air belong to the genus *Clostridium*.

called enteropathogenic bacteria.

It is suggested that acoustic cavitation may induce mechanical rupture of myofibrillar protein structures [31], fragmentation of collagen macromolecules, migration of proteins, minerals and other compounds, thereby accelerating proteolysis or protein denaturation. High-intensity ultrasound can cause degradation of cells and some subcellular components, because periodic oscillation of acoustic pressure softens cell membranes. Research has also evinced tissue disruption in the migration of proteins, minerals, and other components; accelerating enzymatic activity and degradation of collagen macromolecules when meat is exposed to high-intensity ultrasound [11, 55]. In addition to tenderize, high-intensity ultrasound can also improve meat sensory properties [22]. After applying ultrasound, the quantity of ATP available in muscles at pre-*rigor* stage may change [49], accelerating the start of *rigor mortis* [56]. Indirectly, ultrasound may induce tenderization because of the activation of proteolysis by the release of lysosome cathepsins and/or intracellular calcium ions that activate calpains. This mechanism may lead to a weaker cellular structure [48] through protein denaturation, which in turn causes muscle tissue disruption that results in increased tenderness [40] and a shortened aging period [57]. It should be noted that some reports also indicate that ultrasound has no effect on shear force when it is applied at 62 W/cm<sup>2</sup> [58], 22 W/cm<sup>2</sup> [43], 4–19 W/cm<sup>2</sup> [59], or 150 and 500 W [60]. The data available thus far indicate that ultrasound does indeed exert a key effect on meat tenderization, although the application parameters must be established before the method can be scaled to industrial levels. The effect of ultrasound on the physicochemical characteristics and *semitendinosus* muscle collagen has been studied [61]. Their results suggest that ultrasound affects denaturation and aggregation of collagen fibers in the extracellular space. These changes contribute to benefit the quality and texture of the meat. Besides, meat luminosity and tendency to redden remained unaffected.

#### **6.6. Marination to improve meat quality**

High-intensity ultrasound application during meat marination has been frequently studied. Meat marinades may contain salt in two forms: dry or wet [14]. High-intensity ultrasound application resulted favorable for salt diffusion when used in wet marinades. The effect of power ultrasound on pork during wet marination depends on the ultrasound intensity applied [62]. Ultrasound causes bubble formation that hits the tissue, which may lead to microinjection of brine into the sample. This effect may help to explain the observed increase of NaCl content in the ultrasonicated meat [14, 63].

Ultrasonic treatment (low-intensity and low-frequency) and the use of vacuum caused favorable microstructural changes in pork loins marinated in sodium chloride [15] and these effects are highly dependent on the intensity of ultrasound treatment. Some of the critical factors in food processing warrant consideration because ultrasound generates rapid changes in temperature and pressure (109°C/s) over short time periods. Furthermore, cavitation generates shock waves, which contribute to this effect. Factors that modulate the effects of ultrasound application include time of exposure, processing volume, and sample composition [12, 14].

#### **6.7. Microbiological properties**

Numerous studies have been conducted to develop methods to improve tenderness. Among these, ultrasound application methods have been used at various sonication times, frequencies, and intensities. Most authors agree that ultrasound increases meat tenderness [22, 40, 48] and shortens the aging period without compromising other quality parameters [16, 33]. The potential of low-frequency, low-intensity ultrasound application to improve meat tenderization is remarkable. Several authors report an important reduction of shear force after treatment with ultrasound [52]. Benefits of ultrasound treatment on beef have been observed in *longis-*

a significant decrease in the shear force of *Bovine L. dorsi* with the application of ultrasound, both, fresh and aged [22]. More recently, it was reported a reduction of shear force values in muscle semitendinosus when it was ultrasonicated and aged for 3–7 d [45]. The effect was mainly attributed to an increase of desmin and troponin-T degradation, and myofiber fracture

It is suggested that acoustic cavitation may induce mechanical rupture of myofibrillar protein structures [31], fragmentation of collagen macromolecules, migration of proteins, minerals and other compounds, thereby accelerating proteolysis or protein denaturation. High-intensity ultrasound can cause degradation of cells and some subcellular components, because periodic oscillation of acoustic pressure softens cell membranes. Research has also evinced tissue disruption in the migration of proteins, minerals, and other components; accelerating enzymatic activity and degradation of collagen macromolecules when meat is exposed to high-intensity ultrasound [11, 55]. In addition to tenderize, high-intensity ultrasound can also improve meat sensory properties [22]. After applying ultrasound, the quantity of ATP available in muscles at pre-*rigor* stage may change [49], accelerating the start of *rigor mortis* [56]. Indirectly, ultrasound may induce tenderization because of the activation of proteolysis by the release of lysosome cathepsins and/or intracellular calcium ions that activate calpains. This mechanism may lead to a weaker cellular structure [48] through protein denaturation, which in turn causes muscle tissue disruption that results in increased tenderness [40] and a shortened aging period [57]. It should be noted that some reports also indicate that ultrasound has no effect on shear force

[58], 22 W/cm<sup>2</sup>

The data available thus far indicate that ultrasound does indeed exert a key effect on meat tenderization, although the application parameters must be established before the method can be scaled to industrial levels. The effect of ultrasound on the physicochemical characteristics and *semitendinosus* muscle collagen has been studied [61]. Their results suggest that ultrasound affects denaturation and aggregation of collagen fibers in the extracellular space. These changes contribute to benefit the quality and texture of the meat. Besides, meat luminosity

High-intensity ultrasound application during meat marination has been frequently studied. Meat marinades may contain salt in two forms: dry or wet [14]. High-intensity ultrasound application resulted favorable for salt diffusion when used in wet marinades. The effect

for 2 min) [48], and *semitendinosus* (40 kHz, 1500 W for 10, 20, 30, 40, 50 or

[43], 4–19 W/cm<sup>2</sup>

for 180 min) [15] are also reported. It has also been observed

for 240 s) [34], *M. Semimembranosus*

for 4 min after 7 d of storage)

[59], or 150 and 500 W [60].

*simus lumborum* and *semitendinosus* (24 kHz and 12 W/cm<sup>2</sup>

60 min) [40]. Benefits to the texture of poultry (24 kHz, 12 W/cm<sup>2</sup>

(45 kHz and 2 W/cm<sup>2</sup>

162 Descriptive Food Science

[53, 54] and pork (2.5–3 W/cm<sup>2</sup>

along Z-lines and I-bands.

when it is applied at 62 W/cm<sup>2</sup>

and tendency to redden remained unaffected.

**6.6. Marination to improve meat quality**

Bacteria are the most important microorganisms in food processing. While most are harmless and many are beneficial, some indicate the likely presence of contamination and deterioration and may cause diseases. While thousands of bacterial species have been identified, all are unicellular and fall under three basic forms: spherical, rod-shaped, and spiral. Some rod-shaped bacteria can take two forms: latent spores and active vegetative cells. The vegetative cells form spores under adverse conditions to survive. Most sporulating bacteria that grow in the presence of air belong to the *Bacillus* genus, and most of those that grow only in the absence of air belong to the genus *Clostridium*.

Meat is susceptible to the growth of some pathogenic microorganisms such as *E. coli, Campylobacter jejuni, Salmonella spp., Staphylococcus aureus,* and *Listeria monocytogenes*, which recurrently affect the properties of the meat and present serious problems during packing, processing, and storage. Several methods are used to avoid microbial growth in meat. The most commonly used methods involve heating, dehydration, and addition of preservatives [64]. The most common types of mesophilic bacteria that are pathogenic to humans include *Staphylococcus aureus*, *Salmonella*, and *Listeria*. Although it may survive without damage in the intestinal tract of humans, salmonella is a common cause of food poisoning. Another common mesophilic bacterium, *Listeria monocytogenes*, is more often distributed through contaminated foods such as raw meats or unpasteurized cheeses [64]. Animals, including humans, may transport *Listeria*, but it primarily threatens those with weakened immune systems. Some *E. coli* strains found in human feces are pathogenic, causing infection and disease. These are called enteropathogenic bacteria.

*Staphylococcus* is nonsporulated bacteria without mobility, but because they are resistant to drying, they are easily dispersible by dust particles through air and surfaces [65]. *S. aureus* is usually found in the skin and in mucous membranes of humans and other animals. It is almost always present in small quantities in raw meats and foods extensively handled by humans. Maintaining food that is completely free of contamination with *Staphylococcus* is often difficult or impossible. Pasteurizing or cooking destroys the organism but not its toxin [66].Meat is one of the most perishable foods consumed by humans—it is easily damaged by bacteria. One of the most commonly used preservation methods is refrigeration, including freezing. However, certain bacteria are able to grow at 4°C; these are collectively known as psychrophiles. This group includes some pathogens such as *Yersinia enterocolitica*, *Listeria monocytogenes*, nonproteolytic strains of *C. botulinum*, and some strains of enterotoxigenic *E. coli* and *Aeromonas hydrophila*. Several other organisms that can cause foodborne diseases and grow at refrigeration temperatures include: *Vibrio parahaemolyticus*, *Bacillus cereus*, *Staphylococcus aureus,* and some *Salmonella* strains [64].

Gram-positive bacteria are likely less susceptible to ultrasound because of their thicker cell walls, which contain an adhesive peptidoglycan layer [77, 78]. In general, microorganisms

Ultrasound Application to Improve Meat Quality http://dx.doi.org/10.5772/intechopen.77973 165

A considerable amount of data on the impact of ultrasound on microbial inactivation is available. One study demonstrated the effects on the microbiological environment of bacterial suspensions by inoculating the skin of broilers with *Salmonella*; the Salmonella population decreased with ultrasound treatment in peptone at 20 kHz for 30 min [16]. Studies have shown that the intensity of traditional heat treatments can be reduced by 50% when they are combined with power ultrasound. For this reason, a new method for antimicrobial treatment could feature the combined effects of pressure and ultrasound (manosonication), ultrasound and heat (thermosonication), or ultrasound, heat, and pressure (manothermosonication) [79]. These are likely the best microbial inactivation methods because they are more energy-efficient and effective in inhibiting microorganisms than conventional methods. The effectiveness of ultrasonic treatments requires prolonged exposure to high temperatures, which may deteriorate functional properties, sensory characteristics, and the nutritional content of foods [73]. In combination with heat, ultrasound can accelerate the rate of food sterilization, thereby decreasing the necessary duration and intensity of heat treatment and the resulting damage. The inactivation of *Salmonella typhimurium, Salmonella derby, Salmonella infantis, Yersinia enterocolitica,* and a pathogenic strain of *Escherichia coli* was studied in inoculated samples treated for 0.5–2.0 s. The total viable bacterial counts decreased by 1.1 log CFU cm−2 after a 1-second treatment and by 3.3 log CFU cm−2 after a 4-second treatment [80]. The reduction of the population in the skin was significantly greater than that in the meat, although no significant differences were observed between the types of bacteria. However, the study by Smith *et al.* [81] stands out. They reported no effect after ultrasound on *Salmonella* or on *E. coli* in marinated

that produce spores exhibit a greater resistance to heat and ultrasound [74, 75].

chicken, likely because ultrasound alone is not fully effective in bacterial inhibition.

may be a suitable method for decontaminating poultry carcass skins.

were combined with high temperature and amplitude.

Some authors [82] have studied the elimination of Gram-negative bacteria (*Salmonella anatum, Escherichia coli, Proteus* sp., and *Pseudomonas fluorescens*) on the surface of chicken skin after ultrasonic treatment (40 kHz and 2.5 Wcm−2 for 3 or 6 min) in water and in 1% aqueous lactic acid. Sonication in water alone or lactic acid solutions for 3 min resulted in a decrease in the number of microorganisms on the surface of the skin of 1.0 CFU cm−2. Other reports show that treating chicken carcasses in the process line with steam and ultrasonic treatments significantly reduces the population of *Campylobacter* in contaminated poultry. The total viable content decreased by approximately three logarithmic units when steam and ultrasound were applied immediately after slaughter [83]. Ultrasound treatments combined with lactic acid

Ultrasound effects depend on frequency, amplitude, time, and temperature [84] as it was demonstrated on the inactivation of suspensions containing *Escherichia coli, Staphylococcus aureus, Salmonella* sp.*, Listeria monocytogenes*, and *Bacillus cereus* treated with a 12.7-mm ultrasound probe at 20 kHz and 60, 90, and 120 mm amplitudes for 3, 6, and 9 min at 20, 40, and 60°C. These three parameters affected the inactivation of bacteria in pure cultures. The results showed increased microbial inactivation for longer treatment periods, particularly when they

When refrigeration is extended, *Pseudomonas*, *Acinetobacter*, and *Moraxella* species may grow and damage fresh meat [67]. Gram-negative organisms are known to survive less frequently compared to their Gram-positive counterparts [68–70]. However, recent studies have shown higher survival rates among Gram-negative bacteria, especially *Pseudomonas* species, which account for the majority of bacteria responsible for refrigerated meat deterioration [67]. Refrigerated foods, such as processed meat, should be stored as close as possible to 0°C. However, in most cases, they remain close to 4–8°C. This fluctuation in temperature reduces the useful life of the products and can lead to major public health problems. The fresh meat industry must incorporate as many treatments as possible that reduce the microbial population and minimize reproduction. Some of these treatments include heat, acidification, preservatives, reduced water activity, and packaging under modified atmospheres. Although modified atmospheres are included as a potential barrier, it should be noted that reduced oxygen atmospheres can actually favor anaerobic pathogens. For many products, the modified atmosphere actually helps improve product quality rather than safety.

Yeast and molds grow on most foods, equipment, and building surfaces with small amounts of nutrients and moisture [71]. Because bacteria grow faster, they greatly outgrow yeasts and molds in most foods. Fungi and yeasts grow well in low-pH, humid, and temperature environments with high concentrations of salt and sugar. Therefore, they can pose a problem in dry foods, such as dried meat and salted fish [72].

Effective microbial destruction is of paramount importance for food processing; a single report of microbial contamination could question the reputation of a manufacturer and jeopardize their future success. To minimize the bacterial load of a product, the manufacturer must reduce the initial contamination, inactivate microorganisms present in the food, and implement procedures to prevent or slow the growth of microbial populations that have not been inactivated. Conventional methods of bacterial inactivation involve thermal treatments, such as pasteurization. These treatments generally result in undesirable flavors and the loss of nutrients. Ultrasonic treatment has been used to inactivate bacterial populations [73]. This is due to cavitation effects: pressure changes produced by the ultrasonic waves cause microbiological inactivation [3, 73]. The microbiological damage resulting from the application of various ultrasound wave amplitudes depends on factors such as contact time with the microorganism, microorganism type, food quantity, composition, and treatment temperature [74]. Microbial resistance varies among microorganisms, i.e., some are more susceptible than others to the ultrasound process. Studies have shown that larger or longer cells are more susceptible to ultrasound because they have more a larger contact surface and are therefore more exposed to the pressure produced by cavitation [75]. Gram-positive bacteria are less susceptible to ultrasound compared to Gram-negative bacteria, although results have shown that rod-shaped (bacillus) microorganisms tend to be more susceptible than cocci [76]. Gram-positive bacteria are likely less susceptible to ultrasound because of their thicker cell walls, which contain an adhesive peptidoglycan layer [77, 78]. In general, microorganisms that produce spores exhibit a greater resistance to heat and ultrasound [74, 75].

group includes some pathogens such as *Yersinia enterocolitica*, *Listeria monocytogenes*, nonproteolytic strains of *C. botulinum*, and some strains of enterotoxigenic *E. coli* and *Aeromonas hydrophila*. Several other organisms that can cause foodborne diseases and grow at refrigeration temperatures include: *Vibrio parahaemolyticus*, *Bacillus cereus*, *Staphylococcus aureus,* and

When refrigeration is extended, *Pseudomonas*, *Acinetobacter*, and *Moraxella* species may grow and damage fresh meat [67]. Gram-negative organisms are known to survive less frequently compared to their Gram-positive counterparts [68–70]. However, recent studies have shown higher survival rates among Gram-negative bacteria, especially *Pseudomonas* species, which account for the majority of bacteria responsible for refrigerated meat deterioration [67]. Refrigerated foods, such as processed meat, should be stored as close as possible to 0°C. However, in most cases, they remain close to 4–8°C. This fluctuation in temperature reduces the useful life of the products and can lead to major public health problems. The fresh meat industry must incorporate as many treatments as possible that reduce the microbial population and minimize reproduction. Some of these treatments include heat, acidification, preservatives, reduced water activity, and packaging under modified atmospheres. Although modified atmospheres are included as a potential barrier, it should be noted that reduced oxygen atmospheres can actually favor anaerobic pathogens. For many products, the modi-

Yeast and molds grow on most foods, equipment, and building surfaces with small amounts of nutrients and moisture [71]. Because bacteria grow faster, they greatly outgrow yeasts and molds in most foods. Fungi and yeasts grow well in low-pH, humid, and temperature environments with high concentrations of salt and sugar. Therefore, they can pose a problem in

Effective microbial destruction is of paramount importance for food processing; a single report of microbial contamination could question the reputation of a manufacturer and jeopardize their future success. To minimize the bacterial load of a product, the manufacturer must reduce the initial contamination, inactivate microorganisms present in the food, and implement procedures to prevent or slow the growth of microbial populations that have not been inactivated. Conventional methods of bacterial inactivation involve thermal treatments, such as pasteurization. These treatments generally result in undesirable flavors and the loss of nutrients. Ultrasonic treatment has been used to inactivate bacterial populations [73]. This is due to cavitation effects: pressure changes produced by the ultrasonic waves cause microbiological inactivation [3, 73]. The microbiological damage resulting from the application of various ultrasound wave amplitudes depends on factors such as contact time with the microorganism, microorganism type, food quantity, composition, and treatment temperature [74]. Microbial resistance varies among microorganisms, i.e., some are more susceptible than others to the ultrasound process. Studies have shown that larger or longer cells are more susceptible to ultrasound because they have more a larger contact surface and are therefore more exposed to the pressure produced by cavitation [75]. Gram-positive bacteria are less susceptible to ultrasound compared to Gram-negative bacteria, although results have shown that rod-shaped (bacillus) microorganisms tend to be more susceptible than cocci [76].

fied atmosphere actually helps improve product quality rather than safety.

dry foods, such as dried meat and salted fish [72].

some *Salmonella* strains [64].

164 Descriptive Food Science

A considerable amount of data on the impact of ultrasound on microbial inactivation is available. One study demonstrated the effects on the microbiological environment of bacterial suspensions by inoculating the skin of broilers with *Salmonella*; the Salmonella population decreased with ultrasound treatment in peptone at 20 kHz for 30 min [16]. Studies have shown that the intensity of traditional heat treatments can be reduced by 50% when they are combined with power ultrasound. For this reason, a new method for antimicrobial treatment could feature the combined effects of pressure and ultrasound (manosonication), ultrasound and heat (thermosonication), or ultrasound, heat, and pressure (manothermosonication) [79]. These are likely the best microbial inactivation methods because they are more energy-efficient and effective in inhibiting microorganisms than conventional methods. The effectiveness of ultrasonic treatments requires prolonged exposure to high temperatures, which may deteriorate functional properties, sensory characteristics, and the nutritional content of foods [73]. In combination with heat, ultrasound can accelerate the rate of food sterilization, thereby decreasing the necessary duration and intensity of heat treatment and the resulting damage.

The inactivation of *Salmonella typhimurium, Salmonella derby, Salmonella infantis, Yersinia enterocolitica,* and a pathogenic strain of *Escherichia coli* was studied in inoculated samples treated for 0.5–2.0 s. The total viable bacterial counts decreased by 1.1 log CFU cm−2 after a 1-second treatment and by 3.3 log CFU cm−2 after a 4-second treatment [80]. The reduction of the population in the skin was significantly greater than that in the meat, although no significant differences were observed between the types of bacteria. However, the study by Smith *et al.* [81] stands out. They reported no effect after ultrasound on *Salmonella* or on *E. coli* in marinated chicken, likely because ultrasound alone is not fully effective in bacterial inhibition.

Some authors [82] have studied the elimination of Gram-negative bacteria (*Salmonella anatum, Escherichia coli, Proteus* sp., and *Pseudomonas fluorescens*) on the surface of chicken skin after ultrasonic treatment (40 kHz and 2.5 Wcm−2 for 3 or 6 min) in water and in 1% aqueous lactic acid. Sonication in water alone or lactic acid solutions for 3 min resulted in a decrease in the number of microorganisms on the surface of the skin of 1.0 CFU cm−2. Other reports show that treating chicken carcasses in the process line with steam and ultrasonic treatments significantly reduces the population of *Campylobacter* in contaminated poultry. The total viable content decreased by approximately three logarithmic units when steam and ultrasound were applied immediately after slaughter [83]. Ultrasound treatments combined with lactic acid may be a suitable method for decontaminating poultry carcass skins.

Ultrasound effects depend on frequency, amplitude, time, and temperature [84] as it was demonstrated on the inactivation of suspensions containing *Escherichia coli, Staphylococcus aureus, Salmonella* sp.*, Listeria monocytogenes*, and *Bacillus cereus* treated with a 12.7-mm ultrasound probe at 20 kHz and 60, 90, and 120 mm amplitudes for 3, 6, and 9 min at 20, 40, and 60°C. These three parameters affected the inactivation of bacteria in pure cultures. The results showed increased microbial inactivation for longer treatment periods, particularly when they were combined with high temperature and amplitude.

It has been observed that treating fresh beef with a power ultrasound method decreased its bacterial load, particularly of coliforms and psychrophilic bacteria, when a frequency of 40 kHz and intensity of 60 W/cm<sup>2</sup> were applied for 60 and 90 min. Meat treated for the longer period showed the largest reduction of microorganisms during storage [21].

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## **7. Conclusions**

Selected and potential applications of ultrasound mainly in the field of food preservation and product modification were discussed. High-intensity ultrasound generates acoustic cavitation in a liquid medium, developing physical forces that are considered the main mechanism responsible for the observed changes in exposed materials. These forces include acoustic streaming, cavitation, shear, micro-jet, and shockwaves. The quantity of energy released by the cavitation depends on many factors such as treatment medium and ultrasound frequency. Ultrasound has a wide range of applications in the food industry. It can be used as a processing aid in extraction, crystallization, freezing, emulsification, filtration, and drying. Applications of ultrasound in meat have been reported with interesting advantages in freezing, thawing, meat brining, and tenderizing. Ultrasound has also been shown to improve physicochemical characteristics, preparation processes for meat products, microbiological content, and sensory characteristics in fresh and processed meat. Acoustic cavitation may induce the mechanical rupture of the myofibrillar protein structure with significant effect on collagen characteristics and meat textural properties. High-intensity ultrasound reduces microbial loads in meat, resulting in the destruction of living cells and this effect remains during cold storage. Like most innovative food processing technologies, high-power ultrasonics needs to be developed and scaled up for each application.

## **8. Conflict of interest**

The authors declare that they have no interest or benefit arising from the direct applications of this chapter.

## **Author details**

Alma D. Alarcon-Rojo<sup>1</sup> \*, Esmeralda Peña-González<sup>1</sup> , Iván García-Galicia<sup>1</sup> , Luis Carrillo-López<sup>2</sup> , Mariana Huerta-Jiménez<sup>2</sup> , Raúl Reyes-Villagrana<sup>2</sup> and Hector Janacua-Vidales<sup>3</sup>

\*Address all correspondence to: aalarcon@uach.mx

1 Autonomous University of Chihuahua, Chihuahua, Mexico

2 CONACYT - Autonomous University of Chihuahua, Chihuahua, Mexico

3 Autonomous University of Ciudad Juarez, Chihuahua, Mexico

## **References**

It has been observed that treating fresh beef with a power ultrasound method decreased its bacterial load, particularly of coliforms and psychrophilic bacteria, when a frequency of

Selected and potential applications of ultrasound mainly in the field of food preservation and product modification were discussed. High-intensity ultrasound generates acoustic cavitation in a liquid medium, developing physical forces that are considered the main mechanism responsible for the observed changes in exposed materials. These forces include acoustic streaming, cavitation, shear, micro-jet, and shockwaves. The quantity of energy released by the cavitation depends on many factors such as treatment medium and ultrasound frequency. Ultrasound has a wide range of applications in the food industry. It can be used as a processing aid in extraction, crystallization, freezing, emulsification, filtration, and drying. Applications of ultrasound in meat have been reported with interesting advantages in freezing, thawing, meat brining, and tenderizing. Ultrasound has also been shown to improve physicochemical characteristics, preparation processes for meat products, microbiological content, and sensory characteristics in fresh and processed meat. Acoustic cavitation may induce the mechanical rupture of the myofibrillar protein structure with significant effect on collagen characteristics and meat textural properties. High-intensity ultrasound reduces microbial loads in meat, resulting in the destruction of living cells and this effect remains during cold storage. Like most innovative food processing technologies,

high-power ultrasonics needs to be developed and scaled up for each application.

\*, Esmeralda Peña-González<sup>1</sup>

2 CONACYT - Autonomous University of Chihuahua, Chihuahua, Mexico

, Mariana Huerta-Jiménez<sup>2</sup>

1 Autonomous University of Chihuahua, Chihuahua, Mexico

3 Autonomous University of Ciudad Juarez, Chihuahua, Mexico

\*Address all correspondence to: aalarcon@uach.mx

The authors declare that they have no interest or benefit arising from the direct applications

, Iván García-Galicia<sup>1</sup>

, Raúl Reyes-Villagrana<sup>2</sup>

,

and

period showed the largest reduction of microorganisms during storage [21].

were applied for 60 and 90 min. Meat treated for the longer

40 kHz and intensity of 60 W/cm<sup>2</sup>

**7. Conclusions**

166 Descriptive Food Science

**8. Conflict of interest**

of this chapter.

**Author details**

Alma D. Alarcon-Rojo<sup>1</sup>

Hector Janacua-Vidales<sup>3</sup>

Luis Carrillo-López<sup>2</sup>


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## *Edited by Antonio Valero Díaz and Rosa María García-Gimeno*

Deep knowledge of the chemical composition, nutrients, physical properties, toxicology, and microbiological composition of food allows for the production of safe, high-quality foods. This knowledge is fundamental when producing, preserving, manipulating, and distributing food substances, especially to reduce the risks to consumer health. The full extent of the effects on the composition of foods treated by new technologies is still unknown and it must be considered to guarantee that food is produced safely.

*Descriptive Food Science* gives an in-depth insight into this field. Section 1 focuses on the quality of various foods and Section 2 centers on how different technological treatments affect the quality of food.

Published in London, UK © 2018 IntechOpen © zhaojiankang / iStock

Descriptive Food Science

Descriptive Food Science

*Edited by Antonio Valero Díaz* 

*and Rosa María García-Gimeno*