**1.3 Olive-pomace oil**

After VOO extraction, the residual oil, or crude olive-pomace oil (COPO), is extracted by organic solvent extraction or centrifugation from olive oil wastes. After the COPO refining step, the refined olive-pomace oil (ROPO) is blended with VOO, obtaining OPO for human consumption. Currently, the growing interest in OPO is due to its biological active minor constituents (Ruiz-Gutiérrez et al., 2009). The concentration of these components in OPO is higher than the concentration in VOO, with the exception of polar phenols (Perez-Camino & Cert, 1999). Today, new processes for COPO refining are studied in order to diminish the loss of minor components (Antonopoulos et al., 2006). Some of these components are recovered in the refining steps.

Alperujo is treated by the OPO extractors for crude olive-pomace oil extraction (**Figure 1**). First, the major part of the stone present is removed, with the initial stone concentration of about 45% and, after the stone extraction, less than 15%. The stone is easily commercialised for numerous uses, such as in combustion materials, activated carbon, liquid and gas production from stone pyrolysis, an abrasive for surface preparation or for cosmetics, in

with a thick sludge consistency that contains 80 % of the olive fruit, including skin, seed, pulp and pieces of stones, which is later separated and usually used as solid fuel (Vlyssides et al., 2004). In Spain, over 90 % of olive oil mills use this system, which means that the annual production of this by-product is approximately 2,5-6 million tons, depending on the

Alperujo presents many environmental problems due to its high organic content and the presence of phytotoxic components that make its use in further bioprocesses difficult (Rodríguez et al., 2007a). Most of these components mainly phenolic compounds, confer bioactive properties, to olive oil. The extraction of the phenolic compounds has a double benefit: the detoxification of wastes and the potential utilisation as functional ingredients in foods or cosmetics, or for pharmacological applications (Rodríguez et al., 2007a). Although olive mill wastes represent a major disposal problem and potentially a severe pollution problem for the industry, they are also a promising source of substances of high value. In the olive fruits, there is a large amount of bioactive compounds, many of them known to have beneficial health properties. During olive oil processing, most of the bioactive compounds remain in the wastes or alperujo (Lesage- Meessen et al., 2001).Therefore, new strategies are needed for the utilisation of this by-product to make possible the bioprocess

Until now, efforts focussed on detoxifying these wastes prior to disposal, feeding, or fertilisation/composting, because they are not easy degradable by natural processes, or even used in combustion as biomass or fuel (Vlyssides et al.*,* 2004). However, the recovery of high value compounds or the utilisation of these wastes as raw matter for new products is a particularly attractive way to reuse them, provided that the recovery process is of economic and practical interest. This, added to the alternative proposals to diminish the environmental impact, will allow the placement of the olive market in a highly competitive position, and

After VOO extraction, the residual oil, or crude olive-pomace oil (COPO), is extracted by organic solvent extraction or centrifugation from olive oil wastes. After the COPO refining step, the refined olive-pomace oil (ROPO) is blended with VOO, obtaining OPO for human consumption. Currently, the growing interest in OPO is due to its biological active minor constituents (Ruiz-Gutiérrez et al., 2009). The concentration of these components in OPO is higher than the concentration in VOO, with the exception of polar phenols (Perez-Camino & Cert, 1999). Today, new processes for COPO refining are studied in order to diminish the loss of minor components (Antonopoulos et al., 2006). Some of these components are

Alperujo is treated by the OPO extractors for crude olive-pomace oil extraction (**Figure 1**). First, the major part of the stone present is removed, with the initial stone concentration of about 45% and, after the stone extraction, less than 15%. The stone is easily commercialised for numerous uses, such as in combustion materials, activated carbon, liquid and gas production from stone pyrolysis, an abrasive for surface preparation or for cosmetics, in

these wastes should be considered as by-products (Niaounakis & Halvadakis, 2004).

season (Aragon & Palancar, 2001).

**1.2 Utilisation of olive oil wastes** 

applications and the phase separation of alperujo.

**1.3 Olive-pomace oil** 

recovered in the refining steps.

addition to others (Rodríguez et al., 2008). The pitted alperujo is frequently centrifuged in the OPO extractor because the new decanter technology allows treating low-fat material for oil extraction, through which crude olive-pomace oil is obtained. After this mechanical extraction, a partially defatted and pitted alperujo is obtained, with a humidity close to 50%. This material is dried to no more than 10% humidity for both solvent extraction and combustion. Drying consumes much energy, therefore attempts are continuously to reduce energy costs and to avoid the appearance of undesirable compounds in pomace-olive oil formed by the high temperatures (up to 500 ºC) such as polycyclic aromatic hydrocarbons (PAHs) (León -Camacho et al., 2003) or oxidised compounds (Gomes and Caponio, 1997).

Fig. 1. General scheme of industrial olive oil and olive-pomace oil extraction and by-product processing.

New Olive-Pomace Oil Improved by Hydrothermal Pre-Treatments 253

and cellular proliferation and apoptosis in skin and intestinal cancers (Rao, 1998). After being absorbed by the human skin surface, squalene acts as a defence against oxidative

Aliphatic alcohols with long-chain fatty alcohols (C26 or hexacosanol, C28 or octacosanol and C30 or triacontanol) obtained from OPO have shown activity in reducing the release of different inflammatory mediators (Fernández-Arche et al., 2009), reducing platelet

Uvaol and erythrodiol are the triterpenic alcohol fraction present in OPO. They are active antioxidant agents in the microsomal membranes of rat liver (Perona et al., 2005), with positive effects on the inflammatory process (Márquez-Martín et al., 2006), or in the

Alperujo is a high humidity solid that needs special pre-treatments to obtain a viable utilisation of all its phases. Only a few pre-treatments have been proposed for the total utilisation of alperujo, extracting the main interesting fractions. One of the more attractive processes is based on thermal pre-treatments that allow the recovery of all of the bioactive compounds and valuable fractions, making possible the utilisation of alperujo (Fernández-Bolaños et al., 2004). Thermal treatments produce the solubilisation of bioactive compounds to the liquid phase, leaving a final solid enriched in oil, cellulose and proteins. From the liquid, it is possible to extract and purify the bioactive compounds that confer healthy properties to olive oil, mainly phenols such as hydroxytyrosol (HT). HT is one of the more important phenols in the olive oil and fruit because it has excellent activities as a pharmacological and antioxidant agent (Fernández-Bolaños et al., 2002). HT has been recently commercialised by a patented system (Fernández-Bolaños et al., 2005). In addition to other important compounds, a novel phenol has been isolated and purified for the first time: 3,4-dihydroxyphenylglycol (DHPG). DHPG has never been studied as a natural antioxidant or functional compound with a higher antiradical activity and reducing power than the potent HT (Rodríguez et al., 2007b). After the thermal treatment and the solidliquid separation, a solid that is rich in cellulose and oil is obtained. The cellulose is easy to extract and use as a source of fermentable sugar, animal feed or fertiliser (Rodríguez et al., 2007a). The thermal reaction improves the concentration in oil of minor components with functional activities. In addition, phenols increase in the liquid due to the solubilisation, with this fraction a rich source of interesting phenols, sugar and oligosaccharides, all of

This alternative pre-treatment not only increases the concentration of oil in the final solid, but also the content of minor components in COPO prior to the refining process. The thermal treatment improves the functional profile, enhancing the quality and healthy properties of this oil (Lama-Muñoz et al., 2011). The application of thermal pre-treatment to alperujo makes the extraction of olive-pomace oil easier, improving its functional composition. On the other hand, it is important to note that all chemical changes of fats and oils at elevated temperatures result in oxidation, hydrolysis, polymerisation, isomerisation or cyclisation reactions (Quiles et al., 2002, Valavanidis et al., 2004). All of these reactions may be promoted by oxygen, moisture, traces of metal and free radicals (Quiles et al., 2002). Several factors, such as contact with the air, the temperature and the length of heating, the

stress due to the exposure to ultraviolet (UV) radiation from sunlight.

them with a potential use in the food or nutraceutical industry.

**1.5 Thermal pre-treatments** 

aggregation and lowering cholesterol (Taylor et al., 2003, Singh et al., 2006).

prevention and treatment of brain tumours and other cancers (Martín et al., 2009).

The drying process is usually carried out in rotary heat dryers (Espínola-Lozano, 2003) in which alperujo and hot gases obtained from orujillo, olive stones or exhausted gases from co-generation systems (Sánchez & Ruiz, 2006) are introduced at 400 to 800 ºC. The high temperatures have negative effects on the final composition of COPO. After drying, the pitted and partially de-fatted alperujo, with humidity close to 10%, is extracted with organic solvents. After the extraction, the organic solvent is removed for COPO production. The COPO obtained by physical or chemical methods has to be refined for human consumption. The final solid, called orujillo, is commonly used as a biomass for energy production.

The apparition of alperujo was supposed to be a great advantage for olive oil mills because the liquid waste (alpechin) was removed, but it was a serious inconvenience for COPO extractions with regard to the high humidity of the new semi-solid waste, or alperujo. Previous to the two-phase extraction system, the solid waste, or orujo, from the three-phase extraction system was treated with lower humidity (50%) than the alperujo (70%). Nowadays, despite the use of the final solid as biomass, the extraction of olive-pomace oil does not, in many cases, have economic advantages. In addition, the olive oil mills are improving the centrifugation systems in order to increase the quantity of olive oil, producing alperujo with lower oil concentrations. Consequently, the higher humidity in addition to the high organic content of alperujo complicate the COPO extraction, higher temperatures in heat dryers or alperujo with lower oil content. Therefore, pre-treatment alternatives are necessary to properly process the alperujo in the OPO extractor and improve the oil extraction balance and quality, while at the same time obtain new components and add value to the product.

#### **1.4 Minor components in OPO**

Interest in olive-pomace oil is growing due to its economic advantages. It is cheaper than olive oil, and contains many minor components with bioactivities. OPO contains all of the functional compounds found in virgin olive oil, except for the polyphenols, in addition to other biologically active components (De la Puerta et al., 2009; Ruiz-Gutiérrez et al., 2009) that could be solubilised from leaves, skin or seeds of olives, depending on the extraction systems.

Phytosterols, tocopherols, aliphatic alcohols, squalene and triterpenic acid are some of the most important compounds that make the minor components an interesting fraction from the point of view of bioactive compounds that are agents for disease prevention.

The phytosterol´s structure is similar to cholesterol, and they are a powerful agent in the cholesterol-lowering effects in human blood (Jiménez-Escrig et al., 2006) and as a cytostatic agent in inflammatory and tumoral diseases (Sáenz et al., 1998).

Tocopherols (α-, β-, and γ-form) are present in high concentrations in OPO. α-tocopherol is an essential micronutrient involved in several oxidative stress processes related to atherosclerosis, Alzheimer's disease, accelerated aging and cancer (Mardones & Rigotti, 2004). Recently, biological activities against diseases like cancer in animal models have been also attributed to γ-form (Ju et al., 2010).

There is also squalene in olive-pomace oil. This compound has a beneficial effect on atherosclerotic lesions (Guillén et al., 2008, Bullon et al., 2009), dermatitis (Kelly et al., 1999) and cellular proliferation and apoptosis in skin and intestinal cancers (Rao, 1998). After being absorbed by the human skin surface, squalene acts as a defence against oxidative stress due to the exposure to ultraviolet (UV) radiation from sunlight.

Aliphatic alcohols with long-chain fatty alcohols (C26 or hexacosanol, C28 or octacosanol and C30 or triacontanol) obtained from OPO have shown activity in reducing the release of different inflammatory mediators (Fernández-Arche et al., 2009), reducing platelet aggregation and lowering cholesterol (Taylor et al., 2003, Singh et al., 2006).

Uvaol and erythrodiol are the triterpenic alcohol fraction present in OPO. They are active antioxidant agents in the microsomal membranes of rat liver (Perona et al., 2005), with positive effects on the inflammatory process (Márquez-Martín et al., 2006), or in the prevention and treatment of brain tumours and other cancers (Martín et al., 2009).

### **1.5 Thermal pre-treatments**

252 Olive Oil – Constituents, Quality, Health Properties and Bioconversions

The drying process is usually carried out in rotary heat dryers (Espínola-Lozano, 2003) in which alperujo and hot gases obtained from orujillo, olive stones or exhausted gases from co-generation systems (Sánchez & Ruiz, 2006) are introduced at 400 to 800 ºC. The high temperatures have negative effects on the final composition of COPO. After drying, the pitted and partially de-fatted alperujo, with humidity close to 10%, is extracted with organic solvents. After the extraction, the organic solvent is removed for COPO production. The COPO obtained by physical or chemical methods has to be refined for human consumption.

The final solid, called orujillo, is commonly used as a biomass for energy production.

components and add value to the product.

also attributed to γ-form (Ju et al., 2010).

**1.4 Minor components in OPO** 

systems.

The apparition of alperujo was supposed to be a great advantage for olive oil mills because the liquid waste (alpechin) was removed, but it was a serious inconvenience for COPO extractions with regard to the high humidity of the new semi-solid waste, or alperujo. Previous to the two-phase extraction system, the solid waste, or orujo, from the three-phase extraction system was treated with lower humidity (50%) than the alperujo (70%). Nowadays, despite the use of the final solid as biomass, the extraction of olive-pomace oil does not, in many cases, have economic advantages. In addition, the olive oil mills are improving the centrifugation systems in order to increase the quantity of olive oil, producing alperujo with lower oil concentrations. Consequently, the higher humidity in addition to the high organic content of alperujo complicate the COPO extraction, higher temperatures in heat dryers or alperujo with lower oil content. Therefore, pre-treatment alternatives are necessary to properly process the alperujo in the OPO extractor and improve the oil extraction balance and quality, while at the same time obtain new

Interest in olive-pomace oil is growing due to its economic advantages. It is cheaper than olive oil, and contains many minor components with bioactivities. OPO contains all of the functional compounds found in virgin olive oil, except for the polyphenols, in addition to other biologically active components (De la Puerta et al., 2009; Ruiz-Gutiérrez et al., 2009) that could be solubilised from leaves, skin or seeds of olives, depending on the extraction

Phytosterols, tocopherols, aliphatic alcohols, squalene and triterpenic acid are some of the most important compounds that make the minor components an interesting fraction from

The phytosterol´s structure is similar to cholesterol, and they are a powerful agent in the cholesterol-lowering effects in human blood (Jiménez-Escrig et al., 2006) and as a cytostatic

Tocopherols (α-, β-, and γ-form) are present in high concentrations in OPO. α-tocopherol is an essential micronutrient involved in several oxidative stress processes related to atherosclerosis, Alzheimer's disease, accelerated aging and cancer (Mardones & Rigotti, 2004). Recently, biological activities against diseases like cancer in animal models have been

There is also squalene in olive-pomace oil. This compound has a beneficial effect on atherosclerotic lesions (Guillén et al., 2008, Bullon et al., 2009), dermatitis (Kelly et al., 1999)

the point of view of bioactive compounds that are agents for disease prevention.

agent in inflammatory and tumoral diseases (Sáenz et al., 1998).

Alperujo is a high humidity solid that needs special pre-treatments to obtain a viable utilisation of all its phases. Only a few pre-treatments have been proposed for the total utilisation of alperujo, extracting the main interesting fractions. One of the more attractive processes is based on thermal pre-treatments that allow the recovery of all of the bioactive compounds and valuable fractions, making possible the utilisation of alperujo (Fernández-Bolaños et al., 2004). Thermal treatments produce the solubilisation of bioactive compounds to the liquid phase, leaving a final solid enriched in oil, cellulose and proteins. From the liquid, it is possible to extract and purify the bioactive compounds that confer healthy properties to olive oil, mainly phenols such as hydroxytyrosol (HT). HT is one of the more important phenols in the olive oil and fruit because it has excellent activities as a pharmacological and antioxidant agent (Fernández-Bolaños et al., 2002). HT has been recently commercialised by a patented system (Fernández-Bolaños et al., 2005). In addition to other important compounds, a novel phenol has been isolated and purified for the first time: 3,4-dihydroxyphenylglycol (DHPG). DHPG has never been studied as a natural antioxidant or functional compound with a higher antiradical activity and reducing power than the potent HT (Rodríguez et al., 2007b). After the thermal treatment and the solidliquid separation, a solid that is rich in cellulose and oil is obtained. The cellulose is easy to extract and use as a source of fermentable sugar, animal feed or fertiliser (Rodríguez et al., 2007a). The thermal reaction improves the concentration in oil of minor components with functional activities. In addition, phenols increase in the liquid due to the solubilisation, with this fraction a rich source of interesting phenols, sugar and oligosaccharides, all of them with a potential use in the food or nutraceutical industry.

This alternative pre-treatment not only increases the concentration of oil in the final solid, but also the content of minor components in COPO prior to the refining process. The thermal treatment improves the functional profile, enhancing the quality and healthy properties of this oil (Lama-Muñoz et al., 2011). The application of thermal pre-treatment to alperujo makes the extraction of olive-pomace oil easier, improving its functional composition. On the other hand, it is important to note that all chemical changes of fats and oils at elevated temperatures result in oxidation, hydrolysis, polymerisation, isomerisation or cyclisation reactions (Quiles et al., 2002, Valavanidis et al., 2004). All of these reactions may be promoted by oxygen, moisture, traces of metal and free radicals (Quiles et al., 2002). Several factors, such as contact with the air, the temperature and the length of heating, the

New Olive-Pomace Oil Improved by Hydrothermal Pre-Treatments 255

The system scheme is shown in F**igure 3**. A lower range of pressure and temperatures (3-9 Kg/cm2 and 140-180 ºC) than SES is applied for a longer period of time (15-90 min) in the novel system. The conditions and the contact of steam with the sample have been successfully improved, avoiding the technical complications and the high costs of the SES. This treatment has been recently patented to treat olive oil wastes, and the first tests have been carried out to assess its industrial viability for alperujo utilisation (Fernández-Bolaños

Fig. 3. New steam treatment (ST) reactor scheme designed in the Instituto de la Grasa (Seville, Spain) with: 1) sample entrance, 2) reactor chamber (100 L), 3) water steam, 4) sample exit, 5) cold water for refrigeration system, 6) vacuum and 7) solid-liquid separation.

M

**4**

PI

**6**

**1**

**5**

**2**

process and the undesirable compounds that are formed at high temperatures.

environmental impact (Rodríguez et al., 2007a).

The sample is introduced into the reaction chamber together with water steam. The sample temperature is increased up to 190ºC for 30-60 minutes. After the reaction time, the sample is cooled with indirect water as a refrigerant. The liquid and solid phases of the treated sample are separated and the solid is finally extracted to obtain the crude olive-pomace oil. The advantages of both systems are based on the important solubilisation of the initial solid to the liquid phase that occurs during the thermal treatment, leaving a final solid in which several components like oil, cellulose and protein are concentrated. The humidity of the final solid is also easier to remove by centrifugation or filtration, simplifying the drying

NITROGEN

**6**

**7**

**3**

In addition, the liquid phase is rich in bioactive compounds that are easy to extract. All these factors make possible the total utilisation of olive oil wastes, diminishing their

**1.5.2 New steam treatment (ST)** 

et al., 2011).

type of vessel, the degree of oil unsaturation and the presence of pro-oxidants or antioxidants, affect the overall performance of oil (Andrikopoulos et al., 2002). In this work, the effect of two different thermal pre-treatments on COPO composition has been individually studied to balance the positive and negative factors in the final COPO.

#### **1.5.1 Steam explosion system (SES)**

The SES is commonly used as a hydrolytic process for lignocellulosic material utilisation (McMillan, 1994). This process (**Figure 2**) combines chemical and physical effects on lignocellulosic materials. The material is treated with high-pressure saturated steam for a few minutes and then the pressure is swiftly reduced, causing the materials to undergo an explosive decompression. The process causes hemicellulose degradation and lignin transformation due to high temperature, increasing the solubilisation of interesting compounds not only into the aqueous phase but also into the oil fraction. It is used mainly for the treatment of bagasse, such as wheat or rice straw, sugar cane, etc. The pre-treatment can enhance the bio-digestibility of the wastes for bioprocess applications to obtain, for instance, ethanol or biogas, and to increase the accessibility of the enzymes to the materials (De Bari et al., 2004; Palmarola-Adrados et al., 2004; Kurabi et al., 2005). High pressures (10- 40 Kg/cm2) and temperatures (180-240 ºC) are applied with or without the addition of acid in a short period of time, followed by explosive depressurisation. The SES makes it possible to obtain a final solid that is rich in COPO from alperujo. The thermal treatment solubilises a high proportion of solid, leaving behind components such as oil, proteins and cellulose. All these components are concentrated in the final solid.

Fig. 2. Steam Explosion System scheme. Laboratory pilot unit designed in the Instituto de la Grasa (Seville, Spain), equipped with: 1) steam generator, 2) steam accumulator, 3) 2 L reactor stainless-steel and 4) expansion chamber.

#### **1.5.2 New steam treatment (ST)**

254 Olive Oil – Constituents, Quality, Health Properties and Bioconversions

type of vessel, the degree of oil unsaturation and the presence of pro-oxidants or antioxidants, affect the overall performance of oil (Andrikopoulos et al., 2002). In this work, the effect of two different thermal pre-treatments on COPO composition has been

The SES is commonly used as a hydrolytic process for lignocellulosic material utilisation (McMillan, 1994). This process (**Figure 2**) combines chemical and physical effects on lignocellulosic materials. The material is treated with high-pressure saturated steam for a few minutes and then the pressure is swiftly reduced, causing the materials to undergo an explosive decompression. The process causes hemicellulose degradation and lignin transformation due to high temperature, increasing the solubilisation of interesting compounds not only into the aqueous phase but also into the oil fraction. It is used mainly for the treatment of bagasse, such as wheat or rice straw, sugar cane, etc. The pre-treatment can enhance the bio-digestibility of the wastes for bioprocess applications to obtain, for instance, ethanol or biogas, and to increase the accessibility of the enzymes to the materials (De Bari et al., 2004; Palmarola-Adrados et al., 2004; Kurabi et al., 2005). High pressures (10- 40 Kg/cm2) and temperatures (180-240 ºC) are applied with or without the addition of acid in a short period of time, followed by explosive depressurisation. The SES makes it possible to obtain a final solid that is rich in COPO from alperujo. The thermal treatment solubilises a high proportion of solid, leaving behind components such as oil, proteins and cellulose. All

Fig. 2. Steam Explosion System scheme. Laboratory pilot unit designed in the Instituto de la Grasa (Seville, Spain), equipped with: 1) steam generator, 2) steam accumulator, 3) 2 L

individually studied to balance the positive and negative factors in the final COPO.

**1.5.1 Steam explosion system (SES)** 

these components are concentrated in the final solid.

reactor stainless-steel and 4) expansion chamber.

The system scheme is shown in F**igure 3**. A lower range of pressure and temperatures (3-9 Kg/cm2 and 140-180 ºC) than SES is applied for a longer period of time (15-90 min) in the novel system. The conditions and the contact of steam with the sample have been successfully improved, avoiding the technical complications and the high costs of the SES. This treatment has been recently patented to treat olive oil wastes, and the first tests have been carried out to assess its industrial viability for alperujo utilisation (Fernández-Bolaños et al., 2011).

Fig. 3. New steam treatment (ST) reactor scheme designed in the Instituto de la Grasa (Seville, Spain) with: 1) sample entrance, 2) reactor chamber (100 L), 3) water steam, 4) sample exit, 5) cold water for refrigeration system, 6) vacuum and 7) solid-liquid separation.

The sample is introduced into the reaction chamber together with water steam. The sample temperature is increased up to 190ºC for 30-60 minutes. After the reaction time, the sample is cooled with indirect water as a refrigerant. The liquid and solid phases of the treated sample are separated and the solid is finally extracted to obtain the crude olive-pomace oil.

The advantages of both systems are based on the important solubilisation of the initial solid to the liquid phase that occurs during the thermal treatment, leaving a final solid in which several components like oil, cellulose and protein are concentrated. The humidity of the final solid is also easier to remove by centrifugation or filtration, simplifying the drying process and the undesirable compounds that are formed at high temperatures.

In addition, the liquid phase is rich in bioactive compounds that are easy to extract. All these factors make possible the total utilisation of olive oil wastes, diminishing their environmental impact (Rodríguez et al., 2007a).

New Olive-Pomace Oil Improved by Hydrothermal Pre-Treatments 257

Tocopherols were evaluated using the IUPAC 2.432 method. Results were expressed as

The wax and squalene compositions were determined according to the European Regulation EEC/183/93, by separation on a silica gel 60 (70-230 mesh ASTM) chromatographic column (Merck KGaA, Darmstadt, Germany) using hexane/ether (98:2) as the eluent with a few drops of Sudan I as a colorant. Dodecyl arachidate (Sigma) and squalane (Fluka) were

Polar compounds, triglycerides and their derivatives oxidise and hydrolyse were prepared using solid-phase extraction and size-exclusion chromatography and monostearin as internal standard (Márquez -Ruiz et al., 1996). An aliquot (20 μL) of the final solution was injected into a Hewlett Packard Series 1050 HPLC system equipped with a refractive index detector (LaChrom L-7490 Merck) and a 100-Å PL gel column (5 μm) (Agilent). Elution was

Determination of fatty acid, free acidity and peroxide value (PV) was carried out according to the Official Methods described in the European Community Regulation EEC/2568/91. The results were expressed as the percentage of oleic acid. The peroxide value was expressed in milliequivalents of active oxygen per kilogram of oil (mequiv O2/kg oil).

The indexes K232, K270 and *∆K* were determined using the European Communities official methods (European Union Commission, 1991). Oil samples were diluted in isooctane and placed into a 1 cm quartz cuvette; for values calculation, each solution was analysed at 270

Both systems allow the utilisation of the final solid for crude olive-pomace oil extraction. These COPOs have been characterised to assess the positive and negative effects of both treatments on its composition. The SES was studied as a commonly used method for lignocellulosic materials, and the ST was designed to simplify the first system and to diminish the negative effects of SES on crude olive-pomace oil. The lipid fraction of POO extracted from solids treated with either treatment was evaluated, and the minor

An average temperature of 200 ºC and a time of 5 minutes were used with or without acid impregnation of alperujo. The acid increases the severity of the treatment, enhancing the oxidation of the samples. A vacuum was applied to one of the treatments, with acid addition in order to diminish the possible oxidative effect of oxygen at high temperatures and pressures. The results showed (**Table 1**) an important solubilisation of the solid in all treatment. In addition, the oil was concentrated in the final solid from 8,3 up to 19,9 % with respect to the dry final solid. Despite the high level of acidity in the initial sample after the

K232 and K270 are simple and useful parameters for assessing the state of oxidation of olive oil. The coefficient of specific extinction at 232 nm is related to the presence of products of

added as internal standards. The results were expressed as mg/kg of oil.

performed at 0,6 mL/min, with tetrahydrofuran as the mobile phase.

and 232 nm, with isooctane as a blank.

**3.1 Steam explosion system (SES)** 

treatment, these values decreased.

components were also characterised, in the case of the ST.

**3. Results and discussion** 

mg/kg of oil.

First, the application of SES on alperujo in order to obtain pomace olive oil was studied with or without acid as a catalytic agent. Due to the technical disadvantages of SES and to make the use of thermal pre-treatment in OPO mills easier and more convenient low severities, no depressurisation or acid addition were used in the new system (ST).
