**4.3 Emerging technologies for extraction**

Due to the disadvantages of traditional techniques, there is an interest in the development of new extraction techniques. Some of the most sought-after features include shortened extraction time, automation or reduced organic solvent consumption.

#### *4.3.1 Hydrostatic high-pressure extraction*

Hydrostatic high pressure (HHP) is considered an emerging technology that have been applied in the preservation of food since the end of eighties [128]. The first food products treated with this method began commercialization in Japan in 1990 [129]. This method is established on the application of high pressures (100–900 MPa) to the product of interest.

Among the advantages of this type of extraction there is the possibility to conduct it at room temperature, meaning no thermal degradation and derived bioactivity loss from extracted components. It also does not modify chemical structures of the different compounds of interest's independent form of molecular weight [130]. Moreover, in comparison with conventional techniques it is faster,


#### **Table 9.**

*Different experimental conditions carried out in* C. annuum *by enzyme-assisted extraction.*

Capsicum *Seeds as a Source of Bioactive Compounds: Biological Properties… DOI: http://dx.doi.org/10.5772/intechopen.91785*

gives higher extraction yields and fewer impurities [131] giving, unlike other preservation technologies as thermal treatment, uniform and nearly instantaneous effects throughout the foodstuff and thus independent of foodstuff geometry and equipment size which makes an easy scale-up from laboratory findings to full-scale production possible [132].

This method can be applied to multiple matrices for the extraction of natural compounds [133] like fruit and vegetables for different targets as for example carotenoids [134], antioxidants [131] and pigments [135].

The main parameters to be taken into account are pressure, time, temperature and type and quantity of solvent, studying each parameter for each variety in particular.

HHP can change physiological and biochemical properties of pepper based on a study carried out with *C. annuum* [136]. In this research, different pressures (50, 100, 200 and 300 MPa) where applied for 5 min at 25°C observing that it produces remarkable changes in seedlings, but no or very little physiological and biochemical variations, creating an antioxidant system that is positive for the plant itself in the process. Therefore, HHP (500 MPa) can be used as a preservation treatment similar to pasteurization which was the ttraditional thermal treatment for sweet pepper preservation [137].

#### *4.3.2 Supercritical-fluid extraction*

The factors to take into account in this technique are fundamentally temperature, pH, and type of enzyme. A range of enzymes (lipases, carbohydrase, celluloses, proteases, pectinases) have been widely used as specific catalysts. Each enzyme has different substrates. For example, cellulase and hemi-cellulase have their greater hydrolysing activity on the cellulose found in plant cell walls, hence their name. This enzymatic processing increases the permeability of the cell wall, resulting in a better recovery of some compounds like volatile oil and resin, which are prone to degradation when extracted with more disruptive methods [101]. Several studies (**Table 9**) show that viscozyme L is an enzyme with a superior recovery rate of bioactive compounds fractions like total carotenoid content, total phenolic content, total flavonoids and total antioxidant activity with high total suspended solids (TSS). It also has at this moment the better extract yield [95, 96]. Among its applications is the extraction of pigments or capsaicinoids. In almost all cases the pH of the medium is 4.5 as it is the pH of the sample and it is in the range

Due to the disadvantages of traditional techniques, there is an interest in the development of new extraction techniques. Some of the most sought-after features include shortened extraction time, automation or reduced organic solvent con-

Hydrostatic high pressure (HHP) is considered an emerging technology that have been applied in the preservation of food since the end of eighties [128]. The first food products treated with this method began commercialization in Japan in 1990 [129]. This method is established on the application of high pressures

Among the advantages of this type of extraction there is the possibility to conduct it at room temperature, meaning no thermal degradation and derived bioactivity loss from extracted components. It also does not modify chemical structures of the different compounds of interest's independent form of molecular weight [130]. Moreover, in comparison with conventional techniques it is faster,

**Target Enzyme Conditions Yield Ref.**

Carotenoids Viscozyme L 60 4.5 1 87 [95]

Capsaicinoids Viscozyme L 50 4.5 5 88.8 [96]

Oleoresin Viscozyme L 45 4.5 1 6 [101]

*Different experimental conditions carried out in* C. annuum *by enzyme-assisted extraction.*

**Temp pH Time**

**°C h %**

Pectinase 60 4.5 1 80 [95] Cellulase 60 4.5 1 low [95] Viscozyme L 50 4.5 5 78 [96]

Viscozyme L 45 4.5 1 22 [101] Celluclast 45 4.5 1 20 [101]

of optimal activity of the different enzymes [95].

**4.3 Emerging technologies for extraction**

*4.3.1 Hydrostatic high-pressure extraction*

(100–900 MPa) to the product of interest.

sumption.

Capsicum

**Table 9.**

**144**

In supercritical-fluid extraction (SFE), the fluid must reach temperature and pressure above the critical point, so as the fluid behaves like liquid and gas simultaneously which makes extraction easier. This technology has been used in a wide diversity of fields (food, pharmaceutical, chemical and fuel industries) due to its advantages, there is an absence of toxic residue in the final product which allows not only the extraction valuable active compounds (fatty acids, pigments, polyphenols and vitamins) free of solvents, but also to remove undesirable compounds (pollutants, toxins and pesticides) [138]. Additional advantages are great extraction selectivity, short processing times, requirement of minimal solvents, low degradability of the extracted product and the fact that the remaining biomass can be treated with other techniques in order to continue extraction.

The most important conditions are temperature, pressure and co-solvent. The selection of each parameter will depend on the specific compound searched [138]. The most used solvent is CO2 due to its thermodynamics and heat transfer properties. Moreover, it has a low critical point (31°C, 73 bar). Furthermore, the polarity of CO2 can be modified by the use of co-solvents such as ethanol, and in this way also extract polar components.

Studies demonstrate that the extracts obtained with this technique are better than natural spice for flavoring purposes as SFE could reduce aflatoxin in the final products [139]. In addition, numerous bibliographic references show that carotenoid extraction is better at higher pressures [113].

Furthermore, when analyzing the extraction of β-carotene and capsaicin at the same time, capsaicin shows lower solubility, yet the solubility of β-carotene did not change in the presence of capsaicin, a factor to bear in mind when designing the separation process of coloring and hot components from paprika [140]. Several pepper species have been used to obtain natural compounds of interest using supercritical fluid extraction including capsaicinoids, oleoresins, pigments, tocopherols and even aflatoxins (**Table 10**) observing large variations between the pressures applied in each of the methods referred to in the bibliography. The greatest amount of studies is related to the specie *C. annuum* since it is the best known, widespread and cultivated specie of the genus *Capsicum*.

#### Capsicum


air bubbles make the process less effective and the efficiency of the method depends

Further research is necessary since there is no data on the extraction of compounds with this technique. However, there are studies of the application of PEF for the production of dehydrated products or juices. As for drying, a study [143] done with red bell pepper at 320 J/kg, 2.0 kV/cm, 1 Hz and 30 pulses of 400 μs reported good results. Another study [144] with the same aim used different extraction conditions, which were 2.5 kV/cm, 100 Hz and 30 μs as the pulse time. As for juice production, *C. annuum* was studied with extracting conditions of 500 J/kg, 1.7 kV/ cm, 1 Hz and 30 pulses of 300 μs demonstrating that juice from PEF treated paprika

As it is being already done by some factories and companies in pepper growing countries, seeds, stalks or peels are extracted through whole fruit processing with or without semi-automated equipment. This is mainly done by the food industry in order to take away the non-edible parts of the vegetable to elaborate commodities as pepper powders (e.g., paprika), sauces, jams or pickled peppers. This way, stalks,

But in many cases, placenta and seeds are subjected to extraction techniques in order to obtain essential oils to treat chronic ailments via topical administration in traditional remedies [146]. Even so, it is clear that pepper seeds are a multivalent source of many important compounds that could be used in different applications. The elevated phenolic concentrations found in seeds make them an excellent target for extraction and use of these phenolic compounds, whether it is as antioxidant supplements, as food coating in order to increase shelf-life or as a potential source of functional animal feed. Moreover, the implementation of an extraction process in the food industry is dependent on the type of compounds of interest, as the yields of phytochemicals obtained may differ greatly. It should be noted that highly efficient and sustainable extraction methods continue to be a focus of

Also, antimicrobial potential of phytochemicals present in pepper should be considered. Even though this would involve highly specialized equipment and maintain strict aseptic conditions, there is promise in their use as food preservers from common food-borne pathogenic microorganisms besides their antioxidative capability. Furthermore, their use as alternative antibiotics has hardly been researched and the safety of their administration or clinical use has not yet been

Regarding their nutrition attributes, the high linoleic acid and palmitic acid present in the seeds could lead to use them as a primary source of cooking oil, or as a main component in foods such as margarine, or preserver in certain pickled products [48]. Also, as they are a rich natural source of vitamins E and C, extracts of these vitamins could also be incorporated as constituents of many juices and other foods. Pepper-processing industry could find very rewarding to investigate on the yet unknown beneficial properties of its by-products and obtain additional profit by integrating existing extraction and purification technologies into their process chain.

It is clear that the wide spectrum of historical applications of peppers is supported by the latest findings on the properties of peppers. The diversity among

on electric field strength and electrode gap [142].

*DOI: http://dx.doi.org/10.5772/intechopen.91785*

Capsicum *Seeds as a Source of Bioactive Compounds: Biological Properties…*

compared well in quality with enzymatic treatment [145].

**5. Seed valorisation and industrial application**

seeds and placenta are usually removed and discarded.

research both in industry and academia [65].

assessed.

**6. Conclusions**

**147**

#### **Table 10.**

*Different experimental conditions carried out in peppers by supercritical-fluid extraction.*

#### *4.3.3 Pulse electric field extraction*

Pulsed electric field (PEF) consists of a non-thermal method that is extensively used in food processing applications due to, among other things, its ability to kill microorganisms in liquid foods. This method involves the application of short duration electric pulses, making several pores on the cell membrane in what is called electropermeabilization or electroporation. This makes it possible for the selective recovery of intracellular components with low energy consumption as the dielectric breakdown theory explains. According to this theory, the membrane of the cell has a low-dielectric constant that when exposed to a strong electric field provokes ion migration, forming free charges of the opposite sign which accumulate at both membrane sides, generating a potential difference across the membrane which depends on the size and shape of the cells and the concentration of cells in suspension. This difference in charges makes cell walls undergo a compression, reducing the membrane thickness that results in the formation of micropores and increasing permeability (electroporation) [119, 141]. It can be used directly or as a pre-treatment prior to solvent extraction [142]. The parameters to take into account with this method are number of pulses, length of the pulses, energy input and biomass concentration.

Among the advantages of PEF is the short extraction time (usually under 1 s), efficiency at low temperatures, decreased energy losses and a successful cell wall breakdown [120]. The disadvantages are that membrane changes can be reversible, Capsicum *Seeds as a Source of Bioactive Compounds: Biological Properties… DOI: http://dx.doi.org/10.5772/intechopen.91785*

air bubbles make the process less effective and the efficiency of the method depends on electric field strength and electrode gap [142].

Further research is necessary since there is no data on the extraction of compounds with this technique. However, there are studies of the application of PEF for the production of dehydrated products or juices. As for drying, a study [143] done with red bell pepper at 320 J/kg, 2.0 kV/cm, 1 Hz and 30 pulses of 400 μs reported good results. Another study [144] with the same aim used different extraction conditions, which were 2.5 kV/cm, 100 Hz and 30 μs as the pulse time. As for juice production, *C. annuum* was studied with extracting conditions of 500 J/kg, 1.7 kV/ cm, 1 Hz and 30 pulses of 300 μs demonstrating that juice from PEF treated paprika compared well in quality with enzymatic treatment [145].

## **5. Seed valorisation and industrial application**

As it is being already done by some factories and companies in pepper growing countries, seeds, stalks or peels are extracted through whole fruit processing with or without semi-automated equipment. This is mainly done by the food industry in order to take away the non-edible parts of the vegetable to elaborate commodities as pepper powders (e.g., paprika), sauces, jams or pickled peppers. This way, stalks, seeds and placenta are usually removed and discarded.

But in many cases, placenta and seeds are subjected to extraction techniques in order to obtain essential oils to treat chronic ailments via topical administration in traditional remedies [146]. Even so, it is clear that pepper seeds are a multivalent source of many important compounds that could be used in different applications.

The elevated phenolic concentrations found in seeds make them an excellent target for extraction and use of these phenolic compounds, whether it is as antioxidant supplements, as food coating in order to increase shelf-life or as a potential source of functional animal feed. Moreover, the implementation of an extraction process in the food industry is dependent on the type of compounds of interest, as the yields of phytochemicals obtained may differ greatly. It should be noted that highly efficient and sustainable extraction methods continue to be a focus of research both in industry and academia [65].

Also, antimicrobial potential of phytochemicals present in pepper should be considered. Even though this would involve highly specialized equipment and maintain strict aseptic conditions, there is promise in their use as food preservers from common food-borne pathogenic microorganisms besides their antioxidative capability. Furthermore, their use as alternative antibiotics has hardly been researched and the safety of their administration or clinical use has not yet been assessed.

Regarding their nutrition attributes, the high linoleic acid and palmitic acid present in the seeds could lead to use them as a primary source of cooking oil, or as a main component in foods such as margarine, or preserver in certain pickled products [48]. Also, as they are a rich natural source of vitamins E and C, extracts of these vitamins could also be incorporated as constituents of many juices and other foods. Pepper-processing industry could find very rewarding to investigate on the yet unknown beneficial properties of its by-products and obtain additional profit by integrating existing extraction and purification technologies into their process chain.

#### **6. Conclusions**

It is clear that the wide spectrum of historical applications of peppers is supported by the latest findings on the properties of peppers. The diversity among

*4.3.3 Pulse electric field extraction*

*sm, semi-continuous; —, not described. <sup>a</sup>*

*Propane.*

Capsicum

**Table 10.**

biomass concentration.

**146**

Pulsed electric field (PEF) consists of a non-thermal method that is extensively used in food processing applications due to, among other things, its ability to kill microorganisms in liquid foods. This method involves the application of short duration electric pulses, making several pores on the cell membrane in what is called electropermeabilization or electroporation. This makes it possible for the selective recovery of intracellular components with low energy consumption as the dielectric breakdown theory explains. According to this theory, the membrane of the cell has a low-dielectric constant that when exposed to a strong electric field provokes ion migration, forming free charges of the opposite sign which accumulate at both membrane sides, generating a potential difference across the membrane which depends on the size and shape of the cells and the concentration of cells in suspension. This difference in charges makes cell walls undergo a compression, reducing the membrane thickness that results in the formation of micropores and increasing permeability (electroporation) [119, 141]. It can be used directly or as a pre-treatment prior to solvent extraction [142]. The parameters to take into account with this method are number of pulses, length of the pulses, energy input and

*Different experimental conditions carried out in peppers by supercritical-fluid extraction.*

**Target Species Pressure Temp. Flow rate Time Pepper Yield Ref.**

Capsaicinoids *C. frutescens* 20.5 40 0.064 10 50 5.2 [108]

Oleoresins *C. frutescens* 21.5 40 0.071 10 50 0.3 [108]

Pigments *Capsicum* spp. 47.5 80 sm — — 6 [90]

Aflatoxins *C. annuum* 30 50 — 2 7.8 16.2 [139] Tocopherols *C. annuum* 5 25 0.9–1.2a — — 84 [93]

**MPa °C cm s<sup>1</sup> min g %**

*C. frutescens* 7.84 <sup>10</sup><sup>3</sup> 55 1 — 70 7.1 <sup>10</sup><sup>4</sup> [103] *C. annuum* 55 40 — —— 8.6 [92] *C. annuum* 12 40 — 390 — — [102] *C. annuum* 40 55 0.9–1.2<sup>a</sup> — 93 [93] *C. annuum* 7.84 <sup>10</sup><sup>3</sup> 55 1 — 250 7.1 <sup>10</sup><sup>4</sup> [103] *C. chinense* 15 60 2.15 <sup>10</sup><sup>3</sup> 90 2.5 0.5 [111]

*C. annuum* 30 40 — 360 — 8.2 [102] *C. annuum* 40 35 1–1.5 — 7.4 [94] *C. annuum* 43–54 40 1 — 25–30 — [113]

*C. annuum* 36 45 — —— 17.4 [92] *C. annuum* 20 35 0.9–1.2 — — 93 [93] *C. annuum* 40 35 1–1.5 — — 7.2 [94]

*C. annuum* 40 35 1–1.5 — — 100 [94]

Among the advantages of PEF is the short extraction time (usually under 1 s), efficiency at low temperatures, decreased energy losses and a successful cell wall breakdown [120]. The disadvantages are that membrane changes can be reversible, *Capsicum* species properties have been studied over the years, although many of the possible bioactivities have only been characterized in the whole fruit and there are few *in vivo* experiments reported testing phytochemicals of this genus or assessing its safety. Moreover, research on produce by-products has acquired weight within the scientific community and industry as of lately. The exploitation of food processing by-products means not only an affordable raw material for the extraction of valuable phytochemicals, but also a reduction of ecological and economic impact of agricultural and industrial activities. Further studies are necessary to explore in depth more efficient bio-active compounds from the underutilized and non-edible biomass of *Capsicum* spp. Furthermore, the great genotypic diversity of the many pepper varieties grown in different regions, have a high variety of bioactive compounds, some still to be discovered. Currently available knowledge about the medicinal phytochemicals found in the genus *Capsicum* should be considered to develop novel drugs to treat many diseases and ailments.

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Capsicum *Seeds as a Source of Bioactive Compounds: Biological Properties…*

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