Abstract: More than 70% people in developing countries depend on vegetables and fruits further regular dietary needs. Vegetables prevents human from several sever and chronic diseases. It is necessary to consume daily at least 400 g of vegetables including pulses, nuts and seeds. Since vegetables have antioxidant properties they prevents human from cancer, cardio vascular diseases, diabetes, hyper tension, leprosy, rheumatism, epilepsy, liver and urinary disorder, stroke, paralysis etc. The vegetables contain several phytochemicals possessing antioxidant activity. The major groups of phytochemicals include vitamin A, C, E and K, carotenpoid, terpenoid, flavonoids, polyphenols, saponins, enzymes and minerals. The present paper describes the relationship between phytochemicals and antioxidant activity. More than 50 vegetables and leafy vegetables are identified for their antioxidant activity in terms of DPPH, FRAR, IC50, ORAC values. Correlation between antioxidant activities was both positive and negative in vegetables. The present review emphasizes on vegetables as a strong source of antioxidants and a power house of nutrition. The many uncommon vegetables used by the tribals of under developed countries need to be studied for their antioxidant activity and medicinal properties.
INTRODUCTION
Recent researches have identified a vast majority of antioxidants from vegetables and fruits. Antioxidants like vitamin A, C, E, ß-carotene, glutathione precursors like selenium, vitamin B2 (Riboflavin), B3 (Niacin), B6 (Pyridoxin), B9 (Folic acid), B12 (Cynacobalmin), Bioflavanoids which are very rich all seasonal fruits and vegetables with vivid colors stand as prophylactic (Godber, 1990). Selective intake of the foods containing the above antioxidants can prevent the onset of the degenerative diseases like particularly cardiovascular diseases cancers and diabetes. Some very important micronutrients like Chromium and Venadium which improve insulin sensitivity, magnesium preventing retinopathy and vitamin E improving antioxidant defence and insulin sensitivity have become deficient in diabetics. A number of dietary antioxidants exist beyond the traditional vitamins collectively known as phytonutrients or phytochemicals which are being increasingly appreciated for their antioxidant activity. Regular consumption of fruit and vegetables is associated with reduced risks of cancer, cardiovascular disease, stroke, alzheimer disease, cataracts and some of the functional declines associated with aging. Prevention is a more effective strategy than is treatment of chronic diseases. Functional foods that contain significant amounts of bioactive components may provide desirable health benefits beyond basic nutrition and play important roles in the prevention of chronic diseases.
ETHNO-MEDICINAL USES OF VEGETABLES
It is well known that many plants and foods have been and continue to be ingested because of their perceived medicinal and health-benefiting characteristics. Providing modern healthcare to rural people in India is still a far reaching goal due to economic constraints (Grover et al., 2002). Hence, people mainly depend on the locally available plant materials to cure various health disorders (Chopra et al., 1956; Grover and Vats, 2001). Plants possess components, which render beneficial properties (Tanabe et al., 2002). Hence, currently attention is being drawn towards exploring plant sources for substances that provide nutritional and pharmaceutical advantages to humans. Green leafy vegetables (GLVs) are a good source of minerals and vitamins. The ethno-botanical reports offer information on medicinal properties of GLVs like antidiabetic (Kesari et al., 2005) anti-histaminic (Yamamura et al., 1998), anti-carcinogenic (Rajeshkumar et al., 2002), hypolipidemic (Khanna et al., 2002) and anti-bacterial activity (Kubo et al., 2004) (Table 1).
| Table 1: | Traditional uses of some vegetables |
| Table 2: | Antioxidant defence system |
PROTECTIVE ACTION OF ANTIOXIDANTS
Increased consumption of fruits and vegetables has been associated with protection against various age-related diseases (Ames et al., 1993; Steinberg, 1991). What dietary constituents are responsible for this association is not known, but well-characterized antioxidants, including vitamins C and E, or β-carotene, are often assumed to contribute to the observed protection (Ames, 1983; Buring and Hennekens, 1997; Gey et al., 1991; Stahelin et al., 1991; Steinberg, 1991; Willett, 1994). However, the results from intervention trials have not been conclusive regarding the protection following supplementation with such antioxidants (Hennekens et al., 1996; Omenn et al., 1996; Prieme et al., 1997; Van Poppel et al., 1995). Recent epidemiological evidence indicates that the putative beneficial effects of a high intake of fruits and vegetables on the risk of diseases of aging may not be exclusively due to these antioxidants (Hertog et al., 1992; Knekt et al., 1997), but other antioxidant phytochemicals contained in fruits and vegetables may be equally important (Table 2 and 3).
| Table 3: | Vegetables containing nutrients |
Over the years, consumers have been paying more and more attention to the health and nutritional aspect of horticultural products. Having a diet rich in fruits and vegetables will be able to provide some protection against the common diseases such as cardiovascular diseases, cancers and other agerelated degenerative diseases (Scalzo et al., 2005). Evidence shows that free radicals are responsible for the damage of lipids, proteins and nucleic acid in cells could lead to these common diseases (Alothman et al., 2009). Recent studies showed that frequent consumption of fruits and vegetables can reduce the risk of stroke and cancer which is related to the antioxidant microconstituents contained on the plant parts. Different vegetables may exhibit different capacities due to the presence of different dietary antioxidants, such as vitamin C and E, carotenoids, flavanoids and other phenolic compounds (Saura-Calixto and Goni, 2006).
ANTIOXIDANT METABOLITES IN VEGETABLES
Antioxidants ‘Antioxidants’ are substances that neutralize free radicals or their actions (Sies, 1996). Nature has endowed each cell with adequate protective mechanisms against any harmful effects of free radicals: Superoxide Dismutase (SOD), glutathione peroxidase, glutathione reductase, thioredoxin, thiols and disulfide bonding are buffering systems in every cell. α-Tocopherol (vitamin E) is an essential nutrient which functions as a chain-breaking antioxidant which prevents the propagation of free radical reactions in all cell membranes in the human body. Ascorbic acid (vitamin C) is also part of the normal protecting mechanism. Other non-enzymatic antioxidants include carotenoids, flavonoids and related polyphenols, α-lipoic acid, glutathione etc., (Sies, 1996; Cadenas and Packer, 1996; Halliwell and Aruoma, 1993). Antioxidant compounds like phenolic acids, polyphenols and flavonoids scavenge free radicals such as peroxide, hydroperoxide or lipid peroxyl and thus inhibit the oxidative mechanisms that lead to degenerative diseases. There are a number of clinical studies suggesting that the antioxidants in fruits, vegetables, tea and red wine are the main factors for the observed efficacy of these foods in reducing the incidence of chronic diseases including heart diseases and some cancers (Table 2 and 3).
Vitamin C: Ascorbic acid (vitamin C), a powerful, water-soluble antioxidant as scavenger of ROS (Smirnoff, 2000). To prevent or at least alleviate deleterious effects caused by ROS. It has the abitlity to donate electrons in a number of enzymatic and non-enzymatic reactions. As a directly scavenge 1O2, O2*¯ and "OH¯ and regenerate tocopherol from tocopheroxyl radical, thus, providing membrane protection (Thomas et al., 1992). Ascorbic acid also acts as co-factor of violaxanthin de-epoxidase, thus, sustaining dissipation of excess excitation energy (Smirnoff, 2000). Ascorbic acid plays an important role in minimizing the damage caused by oxidative process. This is performed by its synergistic action with other antioxidants (Smirnoff, 2005; Athar et al., 2008). Ascorbate-Glutathione cycle, it plays a role in preserving the activities of enzymes that contain prostheritic transition metal ions (Noctor and Foyer, 1998). Both vitamins C and E show very apparent effects protecting the human body from free radicals and disease. These super vitamins have been shown to have positive effects on tumors activated by ultraviolet light (Table 3).
Vitamin E: Tocochromanols, known as vitamin E, are essential components of biological membranes, exclusively located in the plastid or thylakoid membranes (Munne-Bosch, 2005), where they have both antioxidant and non-antioxidant functions (Kagan, 1989). They occur in four different types or isomers, namely, α, β, γ and δ-tocopherols. All four types of tocopherols structurally consist of a chromanol head group attached to the phytyl tail, which together giving vitamin E compounds amphipathic character (Kamal-Eldin and Appelqvist, 1996). Among others, α-tocopherol with its three methyl substituents has the highest antioxidant activity of tocopherols (Kamal-Eldin and Appelqvist, 1996) and relative antioxidant activity of the tocopherol isomers in vivo is α>β>γ>δ, due to the methylation pattern and the amount of methyl groups attached to the phenolic ring (Blokhina et al., 2003). α-tocopherols prevent the chain propagation step in lipid auto-oxidation and this makes it an effective free radical trap. α-tocopherols found in green parts of plants scavenge lipid peroxy radicals through the concerted action of other antioxidants (Kiffin et al., 2006; Hare et al., 1998). Further, tocopherols are also known to protect lipids and other membrane components by physically quenching and chemically reacting with oxygen in chloroplasts, thus, protecting the structure and function of PSII (Igamberdiev and Hill, 2004). The main function of tocopherol lies in its fatty acyl chain-breaking activity, which scavenges Reactive Oxygen Species (ROS) resulting from photosynthesis, thus protecting polyunsaturated fatty acid chains (PUFAs) from lipid peroxidation. Increasing evidence suggests that in higher plants, vitamin E may play a protective role in cell membrane systems, thus maintaining the integrity and normal function of the photosynthetic apparatus (Havaux et al., 2005; Collin et al., 2008) (Table 3).
Vitamin K: This is a group of structurally similar, fat-soluble vitamins that the human body needs for posttranslational modification of certain proteins required for blood coagulation and in metabolic pathways in bone and other tissue. They are 2-methyl-1,4-naphthoquinone (3-) derivatives (Stafford, 2005). This group of vitamins includes two natural vitamers:vitamin K1 and vitamin K2. Vitamin K1, also known as phylloquinone, phytomenadione, or phytonadione, is synthesized by plants and is found in highest amounts in green leafy vegetables because it is directly involved in photosynthesis (Newman et al., 2008). It may be thought of as the "plant form" of vitamin K. It is active in animals and may perform the classic functions of vitamin K in animals, including its activity in the production of blood clotting proteins. Animals may also convert it to vitamin K2. Vitamin K2, the main storage form in animals, has several subtypes, which differ in isoprenoid chain length. These vitamin K2 homologs are called menaquinones and are characterized by the number of isoprenoid residues in their side chains. Vitamin K1 is found chiefly in leafy green vegetables such as dandelion greens, spinach, swiss chard and Brassica (e.g., cabbage, kale, cauliflower, broccoli and brussels sprouts) and often the absorption is greater when accompanied by fats such as butter or oils (Table 3). Some vegetable oils, notably soybean, contain vitamin K, but at levels that would require relatively large calorific consumption to meet the USDA recommended levels (Weber, 2001).
Folate: Folic acid (also known as folate, vitamin M, vitamin B9, vitamin Bc (orfolacin), pteroyl-L-glutamic acid and pteroyl-L-glutamate are forms of thewater-soluble vitamin B9. Folate is composed of the aromatic pteridine ring linked to para-aminobenzoic acid and one or more glutamate residues. Folic acid is itself not biologically active, but its biological importance is due to tetrahydrofolate and other derivatives after its conversion to dihydrofolic acid in the liver (Bailey and Ayling, 2009). It is present in a wide variety of foods, such as green-leafy vegetables and fruits (Table 3). They very rich in Leafy vegetables such as beets, corn, tomato, broccoli, brussels sprouts, romaine lettuce and bok choy and some of Asian vegetables (Houlihan et al., 2011; Hoffbrand and Weir, 2001). Folate is important for the synthesis of DNA, transfer RNA and the amino acids cysteine and methionine. DNA synthesis plays an important role in germ cell development and therefore, it is obvious that folate is important for reproduction. It has also been reported that folic acid, the synthetic form of folate, effectively scavenges oxidizing free radicals and as such can be regarded as an antioxidant (Joshi et al., 2001). Despite its water-soluble character, folic acid inhibits lipid peroxidation (LPO). Therefore, folic acid can protect bio-constituents such as cellular membranes or DNA from free radical damage (Joshi et al., 2001). Only limited knowledge is available on the impact of dietary folate and synthetic folic acid on (sub) fertility.
Fibers: Dietary fiber is largely composed of complex carbohydrates that are somewhat resistant to digestion. One major component of soluble fibers is pectin, which is largely composed of uronic acid residues such as galacturonic acid. Pectin and other soluble polysaccharides may undergo some metabolism in the small intestine and especially in the large intestine through bacterial enzymes, converting it to products that contribute to maintaining the colonic microflora, which is beneficial to digestion (Weisburger et al., 1993; Cummings et al., 1979; Holloway et al., 1983). Insoluble fiber like cellulose, found in plant cell walls, can aid in waste and toxin removal through several mechanisms (Weisburger et al., 1993). Dietary fibers in foods are also beneficial for good health. Physiological impacts of insufficient dietary fiber intake are constipation, increased risk of coronary heart disease and increased fluctuation of blood glucose and insulin levels (AACC, 2001; Jenkins et al., 1998). Including fruits and vegetables in the human diet may be beneficial, based on their dietary fiber content, with regard to some cancers (Weisburger et al., 1993; Harris and Ferguson, 1993) (Table 3).
SECONDARY METABOLITES IN VEGETABLES IN FREE RADICAL SCAVENGING ACTIVITY
Secondary constituents are the remaining plant chemicals such as alkaloids (derived from amino acids), terpenes (a group of lipids) and phenolics (derived from carbohydrates). Antioxidants are secondary constituents or metabolites found naturally in the body and in plants such as fruits and vegetables. Plants produce a very impressive array of antioxidant compounds that includes carotenoids, flavonoids, cinnamic acids, benzoic acids, folic acid, ascorbic acid, tocopherols and tocotrienols to prevent oxidation of the susceptible substrate (Hollman, 2001). Common antioxidants include vitamin A, vitamin C, vitamin E and certain compounds called carotenoids (like lutein and beta-carotene) (Hayek, 2000). These plant-based dietary antioxidants are believed to have an important role in the maintenance of human health because our endogenous antioxidants provide insufficient protection against the constant and unavoidable challenge of reactive oxygen species (ROS; oxidants) (Fridovich, 1998).
PHENOLIC COMPOUNDS
Phenolic compounds comprise a large group of organic substances and flavonoids are an important subgroup. This group constitutes the majority of dietary antioxidants. There are four major groups of flavonoids, i.e., anthocyanins, flavones, flavonols and the isoflavonols. Anthocyanins, which are coloured flavonoids responsible for a wide range of colours in plants, can scavenge free radicals, particularly singlet oxygen due to their reversible oxidation-reduction properties. Flavonoids, mainly present as colouring pigments in plants also function as potent antioxidants at various levels (Sies, 1996; Cadenas and Packer, 1996). Flavonols include quercetin found in onion and to a lesser extent in French beans and broad beans. The isoflavonoids (isoflavones) are a group of flavonoids in which the position of one aromatic ring is shifted. Isoflavoids may also be responsible for the anticancer benefits of food prepared from soybeans. Food sources that are especially rich in polyphenols include, among others, potato, plums, leafy vegetables, whole grain products and coffee (Souci et al., 2000) (Table 3). Polyphenols scavenge free radicals (R*) possessing an unpaired electron either by donation of hydrogens or electrons, resulting in comparatively stable phenoxyl (PhO*) radicals (neutral (PhO*) or cationic (PhO+*) molecules, respectively), which are stabilized by delocalization of unpaired electrons around the aromatic ring (Rice-Evans et al., 1996; Bouayed et al., 2011a, b). Polyphenols have been reported to be more efficient than vitamin C, vitamin E and carotenoids (concentration ranges between high micromolar and low millimolar in human plasma and organs) against oxidative stress at tissue levels (Scalbert et al., 2002; Manach et al., 2004; Yang et al., 2008; Pandey and Rizvi, 2009; Bouayed and Bohn, 2010; Bouayed et al., 2011b, c).
Carotenoids: Among the various natural pigments, carotenoids comprise a large family of more than 700 structures (Britton et al., 2004) and are synthesized in plants and other photosynthetic organisms, as well as in some non-photosynthetic bacteria, fungi, algae, yeasts and moulds (Stahl and Sies, 2003, 2005). Most carotenoids can be derived from a 40-carbon basal structure, which includes a polyene chain contains 3 to 15 conjugated double bonds. The pattern of conjugated double bonds in the polyene backbone of carotenoids determines their light absorbing properties and influences the antioxidant activity of carotenoids (Goodwin, 1980; Britton et al., 1998). The best documented antioxidant action of carotenoids is their ability to quench singlet oxygen (Paiva and Russell, 1999) via physical or chemical quenching (Stahl and Sies, 2003). Food items rich in carotenoids includes: corn, yellow pepper, apricots, spinach, pumpkin and sweet potato, tomatoes and carrots (Biehler et al. 2012). According to their chemical composition carotenoids are categorized as either carotenes or xanthophylls (oxocarotenoids) (Olson and Krinsky, 1995). While the carotene group, such as β-carotene, α-carotene and lycopene, composed only of carbon and hydrogen atoms, xanthophylls, such as zeaxanthin, lutein, α and β-cryptoxanthin, carry at least one oxygen atom (Stahl and Sies, 2005; Chaudhary et al., 2010).
Minerals: Some minerals are essential for human nutrition (e.g., Fe, Cu, Se and Zn), others such as Cr, Cd, Ni, As and Pb (Hartwig, 1995; Valko et al., 2006), possibly through the formation of Reactive Oxygen Species (ROS) (Rojas et al., 1999; Linder, 2001). The trace elements essentially act as cofactors for antioxidant enzymes involved in the destruction of toxic free radicals produced in the body as a normal consequence of the metabolic processes. Three key trace elements whose roles in antioxidant defence are gradually gaining attention are zinc, selenium and iron. Apparently, Fe, Cu, Zn and Se are necessary to maintain genetic stability and nutritional well-being of humans and animals (Rojas et al., 1999). Recent studies suggest that ultra-trace elements, such as As and Ni, may also play a role both in animal and human nutrition (Rojas et al., 1999). In the last 20 years, a substantial body of evidence has been accumulated to support the role of zinc as a cellular antioxidant (Powell, 2000). Although zinc does not react directly with ROS, a number of indirect mechanisms have been described (Powell, 2000; DiSilvestro, 2000). One of the ways in which zinc acts as an antioxidant is through the induction of the metallothioneins, a group of low-molecular-weight amino acid residues, the production of which is induced by zinc in many tissues including the liver, gut and kidney. For instance, cadmium is present in spinach and cauliflower, while lead is found in Brussels sprouts and Chinese beets ( 3).
| Table 4: | ROS scavenging and detoxifying enzymes and reactions catalyzed |
| *R may be an aliphatic, aromatic or heterocyclic group, X may be a sulfate, nitrite or halide group; †Reaction with H2O2 is slow, (a) Gechev et al. (2006) is used as reference for localization of enzymes. The abbreviations are: Apo, apoplast, Cyt, cytosol; Chl, chloroplasts; CW, cell wall; ER, endoplasmatic reticulum; Gly, glyoxysomes; Mit, mitochondria; Nuc, nucleus; Per, peroxisomes; Vac, vacuole | |
ENZYMATIC ANTIOXIDANTS IN VEGTABLES
Detoxification constitutes the second line of defence against the maleficent effects of ROS. Therefore, once formed, the ROS must be detoxified as effectively as possible to minimize damage (Moller, 2001). Efficient destruction of ROS requires the action of several antioxidant enzymes acting in synchronicity with each other (Noctor and Foyer, 1998). Plants have also developed very efficient enzymatic antioxidant scavenging system to protect themselves against destructive ROS reactions (Mittler et al., 2004). In the cell, toxic effects of ROS are counteracted by several scavenging and detoxifying enzymes, such as superoxide dismutase (SOD), catalase (CAT), Ascorbate Peroxidase (APX), glutathione peroxidases (GPX), glutathione-S-transferases (GST), monodehydroascorbate reductases (MDHAR), dehydroascorbate reductases (DHAR) and Glutathione Reductases (GR) ( 4). While some of these enzymes are entirely dedicated to ROS homeostasis, others are involved also in other processes related to control of development, redox regulation of target proteins and detoxification reactions (Gechev et al., 2006). In cells, different scavenging enzymes encoded by the ROS network can be found in almost every subcellular compartment. Besides, each cellular compartment contains more than one enzymatic activity that detoxifies a particular ROS (Mittler et al., 2004). For example, the cytosol contains at least three different enzymatic activities that scavenge H2O2 (Suzuki and Mittler, 2006). It is clear that the presence of different enzymes in various cellular compartments disclosures their significant role in ROS detoxification for the survival of the plant (Mittler et al., 2004). In Table 4, ROS scavenging enzymes, their abbreviations, EC numbers, catalyzed reactions and cellular localizations are given.
MECHANISM OF ANTIOXIDANT ACTION
The harmful action of ROS and free radicals is normally blocked by antioxidant substances, which scavenge the free radicals and detoxify the organism (Kumaran and Karunakaran, 2006). Antioxidants are compounds that can delay or inhibit the oxidation of lipid or other molecules by inhibiting the initiation or propagation of oxidizing chain reactions (Cakir et al., 2006). All aerobic organisms have antioxidant defense systems (Cakir et al., 2006). These systems are able to scavenge free radicals and increase shelf life of processes foods by retarding the process of lipid peroxidation, the major cause of food and pharmaceutical deterioration (Halliwell, 1996; Gordon, 1996). Antioxidants can protect the human body from free radicals and ROS effects. They retard the progress of many chronic diseases as well as lipid peroxidation (Gulcin, 2007). Being enzymatic or non enzymatic species, antioxidant molecules are classified in different categories (Table 2). Both enzymatic (superoxide dismutase and catalase) and non-enzymatic (antocyanins and tochopherols) antioxidants are able to turn ROS into stable, harmless molecules. They are used in various applications such as pharmaceutical, food, cosmetic and chemical industries in order to act as preservative and extend efficiency and economical value of the products. Considering that antioxidant capacity of an organism is limited, excess production of various oxidizing compounds produces a condition called oxidative stress. Oxidative stress contributes to the pathogenesis of various diseases (e.g., cancers, neurodegenerative disorders and diabetes). Defense against oxidative stress through production of antioxidants and repair processes may constitute important allocations to somatic effort and is particularly relevant for species with low extrinsic mortality.
Antioxidants are major compounds that protect the quality of life by retarding the oxidation process through scavenging free radical produced during many natural events. Although their ultimate aim is removal of ROS, they may use different mechanism depending on their structure and site of action. Antioxidants are also able to act by up-regulating the expression of the genes encoding the antioxidant enzymes, repairing oxidative damage caused by radicals and increasing elimination of damaged molecules (Wood et al., 2006). The use of antioxidants in food industry is inevitable as they can increase shelf life and prevent oxidation. Synthetic antioxidants such as butylated hydroxytoluene (BHT), sodium benzoate and butylated hydroxyanisole (BHA) are widely used in food products. However, their use must be controlled due to possible hazards such as carcinogenicity and toxicity (Ito et al., 1983). Phenolic compounds from plant sources may act as antioxidants by scavenging lipid radicals. Over the last few years, an increasing interest in the search for naturally occurring antioxidants is ongoing. A large number of plants including fruits and vegetables are known as rich sources of antioxidant. The type of plant and its antioxidant activity depends entirely to the region that plant grows and natural vegetation present.
METHODS USED IN ANTIOXIDANT STUDIES: PRINCIPLE AND APPLICATIONS
A variety of in-vitro chemical methods are being used to determine the antioxidant activity of products and ingredients but questions regarding whether the results have any bearing on effectiveness in the human body are leading to development of additional methods that may be more appropriate for screening potential antioxidant ingredients. Several assays have been frequently used to estimate antioxidant capacities in fresh fruits and vegetables and their products and foods for clinical studies including 2,2-azinobis (3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS) (Leong and Shui, 2002; Miller and Rice-Evans, 1997), 2,2-diphenyl-1-picrylhydrazyl (DPPH) (Brand-Williams et al., 1995; Gil et al., 2002), ferric reducing antioxidant power (FRAP) (Benzie and Strain, 1999; Guo et al., 2003; Jimenez-Escrig et al., 2001) and the oxygen radical absorption capacity (ORAC) (Cao et al., 1993; Ou et al., 2001; Prior et al., 2003). The ORAC assay is said to be more relevant because it utilizes a biologically relevant radical source (Prior et al., 2003). These techniques have shown different results among crop species and across laboratories. Ou et al. (2002) reported no correlation of antioxidant activity between the FRAP and ORAC techniques among most of the 927 freeze-dried vegetable samples, whereas these methods revealed high correlation in blueberry fruit (Connor et al., 2002). Similarly, Awika et al. (2003) observed high correlation between ABTS, DPPH and ORAC among sorghum and its products.
Various methods have been developed and applied in different systems but many available methods result in inconsistent results. There is no simple universal method by which antioxidant capacity can be assessed accurately and quantitatively. In this review article, the available methods are critically reviewed on the basis of the mechanisms and dynamics of antioxidant action and the methods are proposed to assess the capacity of radical scavenging and inhibition of lipid peroxidation both in vitro and in vivo. It is emphasized that the prevailing competition methods such as Oxygen Radical Absorption Capacity (ORAC) using a reference probe may be useful for assessing the capacity for scavenging free radicals but that such methods do not evaluate the characteristics of antioxidants and do not necessarily show the capacity to suppress the oxidation, that is, antioxidation. It is recommended that the capacity of antioxidant compounds and their mixtures for antioxidation should be assessed from their effect on the levels of plasma lipid peroxidation in vitro and biomarkers of oxidative stress in vivo.
ANTIOXIDANT ACTIVITY OF VEGETABLES
In search for sources of novel antioxidants and other important nutrients, a large number of plants have been extensively studied (Dasgupta and De, 2007; Indrayan et al., 2005; Iqbal et al., 2006; Joyeux et al., 1995; Odhav et al., 2007; Xin et al., 2004) during last few years.
DPPH FREE RADICAL SCAVENGING ACTIVITY
According to their reducing/antioxidantive power the antioxidantive effect of these vegetables and leafy vegetables can be divided into four groups: (a) low (0-20%), moderate (20-40%), good (40-80%) and very good (80-100%).
The results summarized in Table 1 indicate that the vegetable interact with DPPH radicals and there by stabilize their hyper activity. The lowest groups of vegetables exhibiting antioxidant activity in the range of 0-20% are as follows: Allium sativum, Capsicum annuum Linn. Var. grossam (Willd.) Momordica balsamina L., Solanum tuberosom L. and Spinacia oleraceae Linn.
The average groups of vegetables exhibiting antioxidant activity in the range of 20-40% are as follows: Trigona foenum-graecum Linn., Lycopersicon esculentum, Rumex vesicarius Linn., Cucurbita pepo, Cucumis sativus, Rumex vesicarius Linn., Petroselinum crispum, Cucumis sativus and Raphanus sativus Linn. (Table 5).
The good groups of vegetables exhibiting antioxidant activity in the range of 40-80% are as follows: Brassica oleraceae Linn. Var. capitat, Anethum graveolens Linn., Brassica oleraceae Linn. Var. botrytis, Raphanus sativus Linn., Cucurbita maxima Duch., Lepidium meyenii, Cyamopsis tetragonoloba Linn., Lycopersicon esculentum, Capsicum annuum, Curcuma amada Roxb., Curcuma zedoaria, Macrolepiota mastoidea and Abelmoschus esculenus (Linn.) (Table 5).
The very good groups of vegetables exhibiting antioxidant activity in the range of 80-100% are as follows: Zinger officinale, Leucas aspera, Corchorus olitorius, Crotalaria ochroleuca, Solanum scabrum, Cleome gynandra, Achras sapota Linn., Amaranthus dubius, Asystasia gangetica, Amaranthus hybridus, Amaranthus spinosus, Leucas aspera, Corchorus olitorius, Crotalaria ochroleuca, Solanum scabrum, Cleome gynandra, Achras sapota Linn., Amaranthus dubius, Asystasia gangetica, Amaranthus hybridus, Amaranthus spinosus, Bidens pilosa, Chenopodium album, Centella asiatica, Cleome monophylla, Ceratotheca triloba, Justicia flava, Momordica balsamina, Oxygonum sinuatum, Portulaca oleracea, Physalis viscose, Solanum nigru, Senna occidentalis, Olax psittacorum, Cassia tora, Ipomoea cairica, Leucas aspera, Commelina benghalensis (Table 5).
Vegetables with highest anticancer activity are Allium cepa, Curcuma amada, Curcuma zedoaria, Citrus limon, Brassica oleraceae, Lycopersicon esculentum etc. and the Cucumis sativus is with low level of anticancer activity (La Vecchia and Tavani, 1998; Nahak and Sahu, 2011; Kaur and Kapoor, 2004; Rao et al., 2004). Green leafy vegetables are rich sources of antioxidant vitamins (Gupta et al., 2005). The ascorbic acid, total carotene, β-carotene and total phenolic content of the green leafy vegetables, viz., Amaranthus sp., Centella asiatica, Murraya koenigii and Trigonella foenum graecum etc., are showing highest antioxidant activity among leafy vegetables (Gupta and Prakash, 2009). Spinach and kale are also rich sources of carotenoids and polyphenols. Spinach has an exceptionally high total polyphenol and flavonoid content. The high level of polyphenol acids and flavonoids in spinach leaves influences the high antioxidant activity (Ligor et al., 2012). Majority of leafy vegetables like Brassica, Coriandrum, Carrot, mint, Curry leaf, Radish, Methy contains phytochemicals which shows antioxidant activities include vitamins (A, C, E and K), carotenoid, terpinoid, flavonoid, polyphenols, saponins, enzymes and minerals. All these compounds prevent cancers and other diseases (Govind and Madhuri, 2011).
FRAR VALUE
The FRAP assay determines the capacity of antioxidants as reductants in a redox-linked colorimetric reaction of the reduction of Fe3+-2,4,6-tripyridyl-S-triazine to a blue-coloured Fe2+ complex at low pH that is measured spectrophotometrically at 593 nm (Benzie and Strain, 1996). The extracts were incubated at room temperature with the FRAP reagent and the absorbance recorded after 1 h (Fasahat et al., 2012). The reducing power is expressed as μmol FeSO4 g-1. Here in this review some of the vegetables showing very good antioxidant activity in terms of FRAP (FRAR (Mg-1) values and these are as follows: Daucus carota L. Var. sativa DC. (0.61), Cucumis sativus (1.69), Cucumis melo var. flexuosus Lagenaria siceraria (6.57), Luffa cylindrical L. (2.93), Luffa acutangula (L.) Roxb (2.58), Trichosanthes dioica Roxb. (3.78), Carica papaya L. (4.63), Beta vulgaris L. (5.45) Citrus limon L., Aegle marmelos Correa ex roxb. (6.36), Solanum melongena L. (3.69) and Musa paradisica L. (unripe) (13.18) (Table 5).
IC50 VALUE
IC50 value is defined as the concentration of substrate that 50% loss of DPPH* activity and was calculated by linear regression method of plots of the percentage of antiradical activity against the concentration of the tested compounds. The results summarized in Table 3 and 4 indicate that the IC50 values of Brassica oleraceae (Broccoli) with 7.53 mg mL-1, Musa canvendish (10.93 mg mL-1), Zinger officinale (1.8 mg mL-1) and Phoenix dactylifera (1.65 mg mL-1), Apium nodiflorum (0.07 mg mL-1), Foeniculum vulgare (2.75 mg mL-1), Montia fontana (1.49 mg mL-1) and Anethum graveolens Linn. (3.31 mg mL-1) showed significant lower IC50 values which refer the good antioxidant potential sources (Table 5).
| Table 5: | Antioxidant activity of some vegetables |
ORAC VALUE
Based on widely reported rich antioxidant activities of vegetables, the scientists at United States Department of Agriculture (USDA) have developed a rating scale that measures the antioxidant content of various vegetables. The scale is called Oxygen Radical Absorbance Capacity (ORAC), which stands for oxygen radical absorbance capacity (Table 5). Ou et al. (2002) reported that green pepper, spinach, purple onion, broccoli, beet and cauliflower are the leading sources of antioxidant activities against the peroxyl radicals. Cao et al. (1996) reported that garlic had the highest antioxidant activity (μmol of Trolox equiv g-1) against peroxyl radicals Allium sativum (1939) followed by kale (1770), spinach (1260), Brussels sprouts, alfalfa sprouts, broccoli flowers, beets, red bell pepper, onion, corn, eggplant (980-390), cauliflower, potato, sweet potato, cabbage, leaf lettuce, string bean, carrot, yellow squash, iceberg lettuce, celery and cucumber (380-50). Kale had the highest antioxidant activity against hydroxyl radicals followed by Brussels sprouts, alfalfa sprouts, beets, spinach, broccoli flowers and the others. Lee et al. (2007) reported that among various cruciferous vegetables studied, red cabbage had the highest radical scavenging activity followed by Chinese white cabbage, green cabbage and mustard cabbage (Table 5).
CORRELATION BETWEEN ANTIOXIDANT ACTIVITY AND TOTAL PHENOLIC CONTENT
It is generally believed that plants which are having more phenolic content show good antioxidant activity that is there is direct correlation between total phenol content and antioxidant activity (Qusti et al., 2010; Zhou and Yu, 2006). Typically phenols that possess antioxidant activity are known to be phenolic acids and flavonoid, the major classes of phenolic compounds occurring widely in the kingdom especially in fruits and vegetables (Wojdylo et al., 2007). The amount of total phenols and antioxidanta are, by and large, with certain expections, within the range of values reported by several workers (Chu et al., 2000; Lee, 1992; Anese et al., 1999; Pellegrini et al., 2003) but they are lower than those reported by (Kaur and Kapoor, 2001, 2004; Kahkonen et al., 1999; Prior and Cao, 2000). It has been observed in some cases having no correlation between antioxidant activity and total phenolic content in extracts of some vegetables such as chick pea, drum stick, radish, mustard, purslane, white goose fruits, mountain abony, Caralluma, Carrot, Terminalia cattapa, Banana (unripe) determined by squared regression co-efficient (R2). These plants showed high phenol contents but comparatively low DPPH activity (Khattak, 2011; Chanda et al., 2013). Environmental factors such as differences in light, season, climate and temperature conditions on the one hand and production, optimal extraction factors and genotype on the other may have contributed to the differences in total phenols and antioxidant activities of various vegetables (Kalt, 2005; Rababah et al., 2010). It can be stated that phenolic content of the plant may be a good indicator of its antioxidant capacity (Govind and Madhuri, 2011).
CONCLUSION
Regardless of these advantages, unfortunately the majority of traditional vegetable plants are usually uncultivated and underutilized. It is critical to create awareness regarding diet related health benefits of these neglected precious crops. Further, with reference to food security there is a need to find out every potential source of safe and health-promoting nutrients. Findings of the study indicate that all the studied plants are excellent sources of micronutrients and free radical scavenging activity. The consumption of these traditional vegetables may have many beneficial health attributes. These data may also be helpful in allowing better food choices and improvement in nutritional and health status.
ACKNOWLEDGMENTS
The authors are thankful to University Grants Commission, New Delhi for providing Rajiv Gandhi Fellowship to one of the author and Director School of Biotechnology, KIIT University, Bhubaneswar, Odisha for providing necessary facilities for the research work.