NATURAL ANTIOXIDANTS FROM RESIDUAL SOURCES

Contents:

1. Introduction

1.1. Biological activity of antioxidants

2. Natural sources of antioxidant compounds

3. Crude extracts from materials of residual origin.

3.1. Composition of the crude extracts

3.2. Factors affecting antioxidant activity (of extracts from materials of residual origin)

                        3.2.1. Relation between phenolic content and antioxidant activity.

                        3.2.2. Variety, plant and maturation stage.

            3.3. Processing conditions. 

            3.4. Factors affecting stability of extracts from materials of residual origin

            3.5. Effect of extract concentration

            3.6. Combined action (synergistic, additive or antagonist)

4. Potential antioxidant activity from residual wastes

 

The growing interest in the substitution of synthetic food antioxidants by natural ones has fostered research on vegetable sources and the screening of raw materials for identifying new antioxidants. Oxidation reactions are not an exclusive concern for the food industry, and antioxidants are widely needed to prevent deterioration of other oxidisable goods, such as cosmetics, pharmaceuticals and plastics. Polyphenols are the major plant compounds with antioxidant activity, although they are not the only ones. In addition, other biological properties such as anticarcinogenicity, antimutagenicity, antiallergenicity and antiaging activity have been reported for natural and synthetic antioxidants. Special attention is focussed on their extraction from inexpensive or residual sources from agricultural industries. The aim of this review, after presenting general aspects about natural antioxidants, is to focus on the extraction of antioxidant compounds (mainly polyphenols) from agricultural and industrial wastes, as well as to summarize available data on the factors affecting their antioxidant activity and stability, and, in some cases, the reported major active compounds identified.

 

1. Introduction

            The oxidative deterioration of fats and oils in foods is responsible for rancid odours and avours, with a consequent decrease in nutritional quality and safety caused by the formation of secondary, potentially toxic, compounds. The addition of antioxidants is required to preserve avour and colour and to avoid vitamin destruction. Among the synthetic types, the most fre quently used to preserve food are butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), propyl gallate (PG) and tert-butyl hydroquinone (TBHQ). Tocopherols are also used as antioxidants for food, the order of antioxidant effectiveness being d>g>b>a. Reports revealing that BHA and BHT could be toxic, and the higher manufacturing costs and lower efficiency of natural antioxidants such as tocopherols, together with the increasing consciousness of consumers with regard to food additive safety, created a need for identifying alternative natural and probably safer sources of food antioxidants. The replacement of synthetic antioxidants by natural ones may have benefits due to health implications and functionality such as solubility in both oil and water, of interest for emulsions, in food systems. However, some of them such as those from spices and herbs (oregano,thyme, dittany, marjoram, lavender, rosemary) have limited applications in spite of their high antioxidant activity, as they impart a characteristic herb avour to the food, and deodorization steps are required. Naturally occurring antioxidant substances also need safety testing. Caution regarding an assumption of safety of natural antioxidants has been repeatedly advised, since the fact than an antioxidant comes from a natural source does not prove its assumed safety. Hattori, Yamaji-Tsukamoto, Kimagai, Feng and Takahashi(1998) summarises the requirements that antioxidants must satisfy for use as food additives. Vegetable materials contain many compounds with antioxidant activity. Several plants have been studied as sources of potentially safe natural antioxidants for the food industry; various compounds have been isolated, many of them being polyphenols. A large range of low and high molecular weight plant polyphenolics presenting antioxidant properties has been studied and proposed for protection against lipid oxidation. Polyphenolic compounds affect the functional and nutritional values of vegetable proteins, reducing the nutritional values of foodstuffs, and contributing to the sensory and organoleptic properties of fruits and vegetables (colour, taste, astringency). Polyphenols have other undesirable effects in food systems such as the formation of strong complexes with dietary proteins and with salivary proteins, with digestive enzymes, and protein-polyphenol haze in beverages. Their identification has been extensively reported in seeds that are sources of both food or feed-grade protein.Polyphenol polymerization, due to autoxidation, is responsible for colour loss in processed vegetables tannins also affect the yields of protein extraction. The presence of phenolic compounds, associated with the soluble-pectin fraction, can contribute to changes in cell adherence leading to textural defects such as hard-to-cook beans. Therefore, the potential of tannins to diminish nutrient availability should be considered when using them as biological antioxidants

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1.1. Biological activity of antioxidants

 

Oxidative stress is involved in the pathology of cancer, arteriosclerosis, malaria and rheumatoid arthritis,and could play a role in neurodegenerative diseases and ageing processes. The protection that fruits and vegetables provide against several diseases has been attributed to the various antioxidants, vitamin C, vitamin E, a-tocopherol, bcarotene and polyphenolic compounds  Biological antioxidants, especially vitamin E, were the first studied. In living systems, dietary anti-oxidants (a-tocopherol, b-carotene, ascorbic acid) and endogenous enzymes (superoxide dismutase, glu- tathione peroxidase, catalase) protect against oxidative damage. Several studies have shown that phenolic compounds reduce in vitro oxidation of low density lipoprotein; particularly those phenolics with multiple hydroxyl groups which are generally the most effcient for preventing lipid and low density lipoproteins (LDL) oxidation and therefore, by inference, atherogenesis. Regeneration of a-tocopherol in human LDL was observed in the presence of tea catechins in a dose- dependent manner, although inhibition of LDL oxidation did not reduce arteriosclerotic lesions.Recent scientific studies have proved that antioxidants are capable of protecting cells from free radical damage. Furthermore, other physiological activities of natural antioxidants have been described, such as antibacterial, antiviral, antimutagenic, antiallergic, anticarcinogenic effects, antimetastasis activity, platelet agregation inhibition, blood-pressure increase inhibition, antiulcer activity and anticariogenicity.

 

            Their use as chemopreventive agents by inhibiting radical generation has been suggested since free radicals are responsible for DNA damage and radical scavengers are probably important in cancer prevention. Other studies have reported antimicrobial and antifungal properties of the polyphenolic extracts from Sempervivum tectorum. potato peel vanillin and liquid smoke. During in vivo antioxidant assays with red wine polyphenols, observed that these compounds could play a co-antioxidant role, similar to that described for vitamin C and a sparing role toward vitamin E, which increases due to supplementation with phenols. However, a pro-oxidant effect of phenolics has also been reported. More research is needed in order to establish the activity, bioavailability and other in vivo effects of natural antioxidants.

                                                                                                               

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2. Natural sources of antioxidant compounds

 

            Many of the antioxidants other than vitamin C, vitamin E and carotenoids, occur as dietary constituents. Some studies are published works about strong antioxidant compounds found in fruits. For example, antioxidants with important activity have been found in berries, cherries , citrus and in kiwi fruit, prunes  and olives. High activity antioxidants were found in olive oil and also in fruit juices. Recently a comprehensive review summarised the role of phenolic compounds in the oxidative process of fruits.The effects of processing and storage were evaluated on the changes and content of polyphenols in strawberry, plum, olive oil, grape juice, onions, beans and peas.

           

         Several studies have analysed the antioxidant potential of a wide variety of vegetables, and particularly, of cacao beans, potato, tomato , spinach , legumes such as Phaseolus vul-

garis, or vegetables such as paprika . Both natural extracts and commercial products from garlic and ginger, from rosemary, from dietary supplements, from smoke avourings containing lignin or from drinks  were evaluated for antioxidant activity.

 

            Also antioxidants from seashore plants and seaweeds were studied. Wines contain a variety of polyphenolic compounds, the most abundant being anthocyanins; antioxidant activity was also reported in whiskeys, sake, Jerez-Sherries and cavas.

 

            Green and black teas have been extensively studied for antioxidant properties since they can contain up to 30% of the dry weight as phenolic compounds. Among studies of the antioxidant activity and identification of polyphenols in green and in fermented teas are those of correlated the antioxidant activity with total phenolics content of the tea and found higher activity for green tea than for long or black tea.

Among the major components;

 

 (¡)  epigallocatechin 3 - gallate,

 (¡i ) epigallocatechin,

(¡ii) epicatechin 3-gallate,

(¡v) epicatechin, (+) gallocatechin and (+) catechin were also identified.

 

            Some of the active principles of some medicinal products are polyphenolic compounds. Thus, favones that possess antimutagenic activity, favanones and xanthones, that exhibit antiviral, antimicrobial and antiinflamatory activities, and isoflavones and coumestans that present important physiological effects in humans, have antioxidant action. A number of studies deal with the antioxidant activity of extracts from herbs, medicinal plants and spices. The antioxidant activity of sage components has been widely studied. Also ginger, Ganoderma species , dittany, green pepper , Chrysanthemum ,Honeybush or drugs are antioxidants. Selection of clonal lines with high polyphenols content was studied for lavender.A number of studies focused on the composition of rosemary due to its potent antioxidant action applied either to the retarded oxidation in oil or to the reduction of the loss of colour of carotenoids. The elucidation of the antioxidant mechanisms of its components has also been addressed.

 

            Other potential vegetable sources, such as trees, have been evaluated for antioxidant compounds. among the different parts of the plants, leaves deserve especial attention, e.g. those from green barley, Pelargonium sp., Thalictrum favum, Nerium oleander several willow species , mulberry or avocado. Roots, buckwheat groats ,cork from Quercus suber , bark from Fraxinus ornus and sprouts from mung beans were also reported to contain antioxidants.

 

            Seeds are another source of antioxidants as reported for tamarind, canola ,sesame evening primrose flaxseeds , lupinus seed, buckwheat, sunflower  and Rosa rubiginosa and Gevuina avellana. Also in cereals, as in a recent study on corn kernel, antioxidant activity was detected. Hulls contain compounds with antioxidant activity. Active compounds were detected in hulls from peanut, mung bean and buckwheat. During the extraction of oil from oilseeds, the antioxidant compounds present in the hulls could be incorporated in the oil, as reported for peanut oil extracted from the coated seeds, which contained higher oxidative stability than the oil from dehulled seeds. The bran fraction has been reported to have more antioxidant activity than other fractions, as observed for durum wheat or in the coat of tamarind seeds, with strong oxidation-inhibiting activity, whereas no activity was detected in the germ. Also, in red and black bean seed coat of Phaseolus vulgaris, pro-oxidant species were found in the germ and not in the hulls. That is found; higher inhibition of iron-accelerated oxidation of phosphatidylcholine liposomes for the water-solubles from high-branwheat than for refined wheat. The outer layers usually contain a greater amount of polyphenolic compounds, as expected from their protective function in the plants.

 

            Agricultural and industrial residues are attractive sources of natural antioxidants. Potato peel waste, rapeof olive, olive mill waste waters, grape seeds and grape pomace peels  have been studied as cheap sources of antioxidants and recently increased antioxidant activity in rat plasma after oral administration of grape seed extracts was reported.Identification of polyphenolic compounds from apple pomace, grape pomace, citrus seeds and peels, carrot pulp waste, old tea leaves, cocoa by-products, non-volatile residue from orange essential oil, and soybean molasses has also been reported. Spent ground coffee oil from the residue from the production of instant coffee was used to obtain an antioxidant product useful for food preservation and for aroma stabilisation, the antioxidant activity being due to the 5 hydroxytryptamide carboxylic acids (10± 75% dry wt. of the product)

 

            Scarce literature exists on studies with by-products other than those of plant origin, e.g. shrimp shell waste. Other compounds such as the dipeptide carnosine (beta-alanyl l-histidine) showed antioxidant potential. Protein, protein hydrolysates, soluble elastin peptides, water-soluble proteins and pressure treated b- lactoglobulin   were also reported as antioxidant agents. Essential oils, conjugated linoleic acids and phospholipids present antioxidant activity. Palm oil β-carotene and capsaicin, responsible for the pungent effect of hot chilli peppers are antioxidants, although this latter study evaluates the purified compound. Maillard reaction products were also reported as antioxidant agents.

 

            The derivation of natural products with antioxidant activity from brewing seeds, grains and/or germs has been claimed. Also some microorganisms can produce antioxidants. Few studies deal with the antioxidant activity of the bound phenolic compounds, linked to lignin or arabinoxylans, even though their antioxidant activity in barley and malt is reported to be two-fold higher than that of free phenolic compounds. However, other authors have found that the antioxidant activity of citrus peels and seed extracts is not directly related to the free or bound phenolic compounds.

 

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3. Crude extracts from materials of residual origin

 

3.1. Composition of the crude extracts:

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

TABLE 6.  Summarizes  the main polyphenolic compounds detected in extracts from residual sources. When available, the total extractable polyphenol (T.E.P.) content is also indicated. Due to the wide nature of the agricultural and industrial wastes, a wide variety in the

phenolic components exists. That is reviewed the main phenolic compounds of some studied

fruits.

 

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3.2. Factors affecting antioxidant activity (of extracts from materials of residual origin)

           

     The quality of natural extracts and their antioxidative performances depends not only on the quality of the original plant, the geographic origin, climatic condition, harvesting date and storage, but also environmental and technological factors affect the activities of antioxidants from residual sources.

 

3.2.1. Relation between phenolic content and antioxidant activity

          

      Different results were reported on this aspect; whereas some authors found correlation between the polyphenols content and the antioxidant activity, others found no such relationship. Some of them found a parallel increase between phenol content and antioxidant activity during germination of Pangium edule and some of them observed an increase in the antioxidant activity of lupinseed flour with the different compounds responsible for this activity such as phenolic compounds, peptides/ amino acids and phospholipids. No correlation between antioxidant activity and phenolic content was found in malts, since other compounds are responsible for the antioxidant activity, nor was this relationship between antioxidant activity and phenolic composition found in citrus residues, fruit berry, fruit wines or in plant extracts.

 

3.2.2. Variety, plant and maturation stage

 

            Total polyphenol content and antoxidant activity was found to be different for different parts (leaf, phloem, bark, cork, needle) of trees (pine, birch, spruce, aspen). The superoxide radical-scavenging activities of flavonoids extracted from different parts (leaves, tender leaves, branches and bark) of mulberry trees were also different . The extraction yield and the anti- oxidant activity differs among fractions of the milled durum wheat bran (bran, head shorts, tail shorts, low-quality flour and low-grade flour), but slight and non-significant differences were observed for diferent varieties. It is  found that  a slight reduction in the amounts of the total polyphenol (as gallic acid equivalents) content from young and mature leaves of different varieties of Avocado, but no differences in the mesocarp content.

 

            It is reported that  phenol mobilization during seed germination and increased content and antioxidant activity during the production of precursors for the synthesis of lignin. Also it is studied that the effect of the maturity of peanuts on both the polyphenols content of the hulls and the antioxidant activity of their methanolic extracts. Luteolin and total phenols increased with maturity, but a maximum antioxidant activity (92.9±94.8% inhibition of linoleic acid peroxidation) was detected at a total polyphenol content of 1.67 mg/g hulls, the luteolin content being more dependent on the maturity than on the variety. It is  found that the total phenol content differed significantly among peanut cultivars, although the specific antilipoperoxidant activity was similar. In paprika, harvesting at different ripening stages affected the content of ascorbic acid and tocopherols . Other factors, such as insect infestation, were reported to increase polyphenol content in maize, wheat and sorghum and, in the latter two, phytic acid content also.

 

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3.3. Processing conditions

 

3.3.1. Effect of the extracting solvent

 

            Solvent extraction is more frequently used for isolation of antioxidants and both extraction yield and antioxidant activity of extracts are strongly dependent on the solvent, due to the different antioxidant potential of compounds with different polarity. Apolar solvents are among the most employed solvents for removing polyphenols from water. Ethyl acetate and diethyl ether have been used for extraction of low molecular weight phenols from oak wood and the polyphenols extracted with ethyl acetate from natural materials were reported to have strong antioxidant activity. Ethanol and water are the most widely employed solvents for hygienic and abundance reasons, respectively. Since the activity depends on the polyphenol compounds and the antioxidant assay, comparative studies for selecting the optimal solvent providing maximum antioxidant activity are required for each substrate. Less polar solvents such as ethyl acetate, provided slightly more active extracts than mixtures with ethanol or methanol, or methanol alone for tamarind seed coats although ethanol and methanol extracts also presented high lipid peroxidation-inhibiting activity, comparable to a-tocopherol. Selective extraction of more apolar compounds was reported to enhance the antioxidant activity of lentil husk extracts.Lower IC50 values for the DPPH radical (amount of antioxidant required for causing a 50% reduction in the absorbance of DPPH) were observed for butanol extracts, followed by those in ethyl acetate. Those obtained with methanol-water were less efficient.

 

    That is found; a different behaviour in the extraction of different compounds and total extractable polyphenols (TEP). Maximum total phenolics extraction yields were attained with methanol, whereas 50% acetone extracted more selectively leucoanthocyanins and no significant effects were observed in the extraction of glycosides. Also, for extracts from burdock roots, water (regardless of the temperature used) yielded the greatests amount of extract and exhibited the strongest antioxidant activity . it is reported that maximum antioxidant activity from cocoa by-products (cocoa powder, cocoa nib, cocoashell) in the methanol, followed by mixtures of chloroform, ether and dichloroethane or chloroform, methanol and dichloroethane. The polyphenol extraction yield was higher for the more polar solvents for extracts from Gevuina avellana hulls. It is  reported that the antioxidant activity of buck-wheat extracts varied with the polarity of the solvent, those extracted with methanol being the most active. The effect of the extraction pH has also been reported. It is reported that maximum solubility of polyphenols from olive rape at pH4 in the organic phase. Also it is reported that  increased antioxidant activity of aqueous fractions from wheat bran after treatment at acidic conditions, probably due to altered phenol composition. The pH has also been considered for the aqueous extraction of antioxidants from oat fibre, the highest yield being attained at pH6 and the highest antioxidant activity at pH10. At alkaline pH, the fractions with high protein and fatty acid contents are solubilized; and due to contradictory data on the higher antioxidant activity of carbohydrate or proteins, the antioxidant activity was probably carried by the protein-rich fraction. The physical state of the fibre rather than the total concentration of some specific fibre compound were suggested as responsible for the higher antioxidant activity. That is reported selective extraction of flavan-3ol monomers, catechin and flavonols from grape marc, preferentially in the organic phase, whereas procyanidins were extracted in the aqueous phase. Reduction in particle size flavours solvent extraction of polyphenols and both mechanical crushing and enzyme demolition were reported on grape marc. Enzyme-aided extraction of antioxidants from grape pomace has been reported. The yield of extracted phenols was correlated with the plant cell wall breakdown caused by pectinases and cellulases, although these latter did not cause the degradation of grape pomace polysaccharides. Particle size reduction significantly increased the antioxidant activity as a result of both increased extractability and enhanced enzymatic degradation of polysaccharides.

 

    Increased polyphenol recovery from rosemary and sage, during enzyme-assisted ensiling with cellulases, hemicelulases and pectinases, was reported.

 

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3.3.1.1. Temperature:

 

             The temperature, during drying and extraction, affects the compound stability due to chemical and enzymatic degradation, losses by volatilization or thermal decomposition these latter have been suggested to be the main mechanism causing the reduction in polyphenol content. Also, for synthetic antioxidants, evaporation and decomposition were the main mechanisms for the loss of activity. In addition to thermal decomposition, phenols can react with other plant components, impeding their extraction. Decomposition to more active compounds has also been described; it  is  observed that mild pyrolysis of some polyphenolic acids increased the antioxidant activity over that of the original compounds, especially in the case of cafeic acid. Degradation, caused by other agents, has been observed; demonstrated ring-opening during alkaline oxidative conditions, as in non-enzymatic reactions involving polyphenolic compounds in food systems, and identified the resulting products as compounds analogous to natural lignans and neolignans. Prolonged exposure at moderate temperatures can also cause phenolic degradation during their enzyme-assisted extraction from grape pomace for 48 h hydrolysis (40_C and pH 5), whereas at 1±8 h, no degradation was observed. Also is  observed 20% reduction in antioxidant activity during kilning at 900C for bound and free polyphenols. The temperature during extraction can affect the extractable compounds differently: boiling and resting increases the total phenol content in Quercus suber cork; however, proanthocyanidin content decreased. Milder extraction temperatures are desirable in those cases where some compound can be degraded, e.g. carnosic acid, and, for these reasons, supercritical fluid extraction was reported to provide extracts with higher antioxidant activity. The effect of temperature has been studied in spray-drying of carrot pulp waste, but the drying method also affects the retention and preservation of β-carotene, drum drying being the best preservation method due to the particle size and surface carotenoid content. It is found that  a significant reduction in extractable polyphenols and condensed tannins when red grape pomace peels were dried with air at 1000C or higher. The antioxidant activity of samples dried with air at 1000C was reduced by 28% and, at 1400C by half, with respect to drying at 600C, that did not significantly affect either the extractable polyphenols or condensed tannins, with respect to freeze-drying. Drying at 1000C caused a reduction of 18.6% and at 1400C of 32.6% in the TEP s , which in this material are a complex group of different substances (phenolic acids, anthocyanins, flavonols, flavan-3-ols, and flavanonols). Anthocyanins were also probably degraded since the visible spectrum showed both a reduction in the peak at 400±500 nm and reduction in red colour. The reduction in antioxidant activity was higher than that expected from the reduction in polyphenols content, probably due to the synergistic effect of natural phenols. The amount of flavonoids in fresh Mulberry leaves was higher for air-dried than for oven-dried, probably due to decomposition after storage or to lowered extractability due to modification of the matrix. Both thermal decomposition and losses by volatilizing have been suggested as the main causes for lowered yields. Also, is reported that  a maximum yield of total willow leaf polyphenols when the drying temperature was below 500C; increasing the temperature above 600C significantly lowered the phenols, the leucoanthocyanins content being the most affected by temperature.

 

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3.4. Factors affecting stability of extracts from materials of residual origin

 

    Temperature and light are the major factors influencing antioxidant activity during storage. These factors affect different compounds to different extents. The reduction in the free radical scavenging activity, caused by exposure at high temperature, was more marked for red grape pomace peel (28.5) than white grape pomace peel (22.9) and these latter more than BHA (15.3), but all of them were lower than for α-tocopherol. The stability of different extracts from the same material was dependent on the extracting solvent used for the solubilization and removal of the polyphenolic compounds; methanol extracts from cocoa by- products were stable up to 500C and in a wide range of pH (3±11), whereas other extracts (chloroform, methanol, dichloroethane) were less stable.

          

      It is  found that neither autoclaving nor storage at 250C caused changes in potato peel polyphenol concentrations, whereas those exposed to light suffered complete degradation of chlorogenic acid after 7 days, and an increase in caffeic acid slightly higher than the 60% of the disappeared chlorogenic acid, the remaining portion probably being degraded into another compound or compounds. The caffeic acid disappeared completely in 20 days. No stability loss was noticed in freeze-dried samples during storage, but degradation of caffeic acid and increase of gallic acid during freeze-drying were observed as a result of the freeze-drying process. This extract was found to be stable for 3 years when stored, tightly capped, in plastic vials at room temperature (230C), since non-significant changes in both total phenols and antioxidant activity were found when measured as inhibition of sunflower oil oxidation.

 

            It is found that, in the darkness at 40C after 6 months, the ethanolic and aqueous extracts from G. avellana hulls were stable, but those extracted with acetone showed a 97% reduction in the β-carotene bleaching activity and 43% in DPPH radical-scavenging activity with respect to the freshly prepared ones. It is reported that increased stability of the antioxidant activity from cocoa by-products with increasing pH from 3 to 11.

 

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3.5. Effect of extract concentration

 

            The antioxidant activity depends on the extract concentration. As a general trend, increased antioxidant activity was found with increasing extract concentration, but the concentration leading to maximum antioxidant activity is closely dependent on the extracts and, for the same extract, is dependent on the antioxidant activity test. Dose-response curves are different for different antioxidants. It is compared that grape seed extract with natural antioxidants, such as tocopherol and ascorbic acid and observed different effectiveness, depending on the assay. The superoxide anion-scavenging activity was found to be dependent on the flavanol concentration. It is observed that  absence of linearity in the dependence of stabilization factor on esculetin concentration, probably due to the participation of the antioxidant in reactions other than in chain termination. As a general trend, the antioxidant activity increases with the antioxidant concentration, but only up to a certain level, which depends on both the antioxidant and the test. In liposomes, the optimal concentration of grape seed and rose hip extracts was 0.1 mM, whereas for BHT it was 0.02mM and for catechin a steady state was observed in the range 0.05±0.2 mM. For most tests and natural extracts, maximum antioxidant activity was achieved using a 0.05% concentration. Acetone extracts from G. avellana hulls when used at concentrations under 1000 mg/l, showed prooxidant activity, but increased antioxidant activity was observed with increased concentration.

 

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3.6. Combined action (synergistic, additive or antagonist)

 

            Synergistic actions between synthetic only, natural and synthetic, and natural antioxidants have been observed . This effect is defined as the combined action which results in increased antioxidant potential more than that expected from a mere additive effect. It is observed that α- tocopherol and ascorbic acid acted highly synergistically with each other in a fish oil/lecithin/water system, requiring a minimum of 0.01±0.02% ascorbic acid.

 

    It is observed that  this effect between p-coumaric and ferulic acids with ratios between 0.14± 0.22 for the expected/observed antioxidant activity measured as percentage increase (respect to a control) inthe half-life during accelerated oxidation of methyl linoleate. Protective effects against caffeic acid autoxidation in the presence of ascorbic acid were also observed. Synergistic effects of phenols from grape seeds and pomace polyphenols have been reported. Mixtures of tocopherol andcarotene, as well as mixtures with other substances (ascorbic acid, lecithin), which have been reported to enhance the antioxidant activity found interactive effects between flavonoids and phenolic acids. However, the simultaneous presence of some compounds may present lower antioxidant activity than expected; in this way antagonist effects were observed between ellagic acid and catechin. The authors suggested the possible existence of hydrogen-bonding between carbonyls in ellagic acid and o-dihydroxyl groups in catechin. Synergistic antioxidant effects between the compounds found in natural extracts are probably responsible for the higher antioxidant activities observed for the crude extracts than that measured in simulated extracts. Synergistic antioxidant effects were observed for mixtures of crude extracts of burdock and tocopherol and of grape seed extracts and ascorbic acid.

 

            Other substances could also act synergistically with the phenols; therefore, these compounds could not be the only ones responsible for the antioxidant activity. These authors reported a PV value of 37 meq/kg for soy oil after 9 h active oxygen method (AOM) treated with durum wheat bran extract, but a significantly higher value of 46.0 meq/kg for the simulated extract with the authentic standards in the proportions found in the extracts. The PV for the oil, subjected to oxidation in the presence of added pure polyphenols, ranged from 84 meq/kg when p-coumaric acid was added to 39 meq/kg when protocatechuic acid was used. Other plausible reasons could be the synergistic effects of the different phenolic compounds. İt is  reported that  higher antioxidant activity for freeze-dried potato peel extract during sunflower oil oxidation than for the synthetic mixture of individual compounds. Similarly, is reported significantly higher peroxyl-radical scavenging activity of some of the purified fractions of buckwheat hull extracts over the crude extracts. Synergy among the different classes of polyphenols was observed and reported as hypothetically existing in red wine.

 

            However, an antagonist effect of the methanolic extract with peanut hulls and tocopherol and with BHA, both at 48 and 120 ratios was reported. Since there is no single antioxidant that can scavenge all kinds of radicals or that performs optimally for all lipid products, mixtures of antioxidants resulting in a synergistic e€ffect are preferred for preventing free radical-induced diseases. Combined use of antioxidants will probably be desirable, as observed for model compounds. The use of synergistic mixtures of antioxidants allows a reduction in the concentration of each and also increases the antioxidant effectiveness with respect to the activity of the separate components although, even in widely used and commercialized extracts, such as rosemary, the antioxidative behaviour and synergistic actions of most of the compounds remain unknown. The author observed a two or three times higher oxidative stability index for peanut oil when mixtures of antioxidants were used. The beneficial effects of using mixtures of antioxidants were summarized as:

 

(1) advantages of their di€erent e€ectiveness;

(2) minimalisation of solubility or colour problems presented by individual compounds;

 (3) better control and accuracy of application;

(4) complete distribution or solution of antioxidants and chelating agents.

 

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4. Potential antioxidant activity from residual wastes

 

            Table 7. summarizes the antioxidant activity of extracts from residual sources and, when reported, the antioxidant activity of natural or synthetic antioxidants is given for comparative purposes. The concentrations tested, for both the natural extracts and for the standard

compounds used for comparative purposes, are also indicated. As a general rule the extracts from vegetable materials of residual origin showed antioxidant activity, in some cases comparable to that of synthetic antioxidants, and their extraction and use could be an alternative for obtaining natural antioxidants. Even when the natural extracts are less effcient, the use of some of them as food antioxidants can be advantageous. Maximum levels established for synthetic food additives need not be applicable to naturally occurring compounds, e.g. those from grape marc.

 TABLE.7.

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