Mamta, Kshipra Misra, Gurpreet Singh Dhillon, Satinder Kaur Brar, and Mausam Verma


Antioxidants are the molecules that prevent cellular damage caused by oxidation of other molecules. Oxidation is a chemical reaction that transfers electrons from one molecule to an oxidizing agent. Oxidation reactions are known to produce free radicals. These free radicals are highly reactive species which contains one or more unpaired electrons in their outermost shell. Once they are formed, the chain reaction starts. Antioxidant reacts with these free radicals and terminates this chain reaction by removing free radical intermediates and inhibits other oxidation reactions by oxidizing themselves.

Though oxidation reactions are crucial for life, they can also be damaging. Plants and animals have a complex system of multiple types of antioxidants, such as vitamin C and vitamin E, as well as enzymes, such as catalase (CAT), superoxide dismutase (SOD), and various peroxidases (Hamid et al. 2010). Oxidative stress plays a key role in causing various human diseases, such as cellular necrosis, cardiovascular disease, cancer, neurological disorder, Parkinson's dementia, Alzheimer's disease, infl tory disease, muscular dystrophy, liver disorder, and even aging (Amit and Priyadarsini 2011). Besides, there are some antioxidants in the form of micronutrients which cannot be manufactured by the body itself such as vitamin E, β-carotene, and vitamin C, and hence these must be supplemented in the normal diet (Teresa et al. 2011).

Antioxidants can also act as prooxidants when these are not present at the right place at the right concentration at the right time (Touriño et al. 2008). The relative importance of the antioxidant and prooxidant activities is not yet explored fully and needs further research.

In this chapter, authors have tried to discuss the various types, sources, synthesis, uses, and protective efficacy of antioxidant with examples.

Classification of Antioxidants

Antioxidants can be classified into two major types based on their source, i.e., natural and synthetic antioxidants (schematic representation of the classification of antioxidants is shown in Fig. 6.1).

Natural Antioxidants

Natural antioxidants either are synthesized in human body through metabolic process or are supplemented from other natural sources, and their activity very much depends upon their physical and chemical properties and mechanism of action. This can be further divided into two categories, i.e., enzymatic antioxidants and nonenzymatic antioxidants. Enzymatic Antioxidants

Enzymatic antioxidants are uniquely produced in the human body and can be subdivided into primary and secondary antioxidant.

Primary Antioxidants

Primary antioxidants mainly include superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) as described below.

Superoxide Dismutase Superoxide dismutase (SOD) enzyme is found in both the dermis and the epidermis. It removes the superoxide radical (O .−) and repairs the body cells damaged by free radical. SOD catalyzes the reduction of superoxide anions to hydrogen peroxide (6.1). SOD is also known to compete with nitric oxide (NO) for superoxide anion, which inactivates NO to form peroxynitrite. Therefore, by scavenging superoxide anions, it promotes the activity of NO (Chakraborty et al. 2009).

Fig. 6.1 Schematic representation of classification of antioxidants

Catalase Catalase enzyme (CAT) is found in the blood and most of the living cells and decomposes H2O2 into water and oxygen (6.2). Catalase with glucose peroxidase is also used commercially for the preservation of the fruit juices, cream consisting of egg yolk, and salad by removing the oxygen (Chakraborty et al. 2009).

Glutathione Peroxidase Glutathione peroxidase (GPx) is a group of seleniumdependentenzymes, anditconsistsofcytosolic, plasma, phospholipidhydroperoxide, and gastrointestinal glutathione peroxidase (Chakraborty et al. 2009). GPx (cellular and plasma) catalyzes the reaction of H2O2 by reduced glutathione (GSH); as a

Fig. 6.2 Outline of the mechanism of enzymatic antioxidants in the removal of free radical

result, oxidized glutathione (GSSG) is produced (6.3) and it is again recycled to its reduced form by glutathione reductase (GR) and reduced nicotinamide adenine dinucleotide phosphate (NADPH).

Secondary Antioxidant

Secondary antioxidant includes glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH). G6PDH generates NADPH. GR is required to recycle the reduced glutathione (GSH) using secondary enzyme GR and NADPH (6.4).

GSSG + NADPH ¾G¾R ® NADP + 2GSH (6.4)

Glutathione is a cysteine containing peptide-type antioxidant and is synthesized in the body cells. The thiol group in its cysteine moiety is a reducing agent and can be reversibly oxidized and reduced. A high level of glutathione is found in the cells (~3,100 μg/g of tissue) (Hissin and Hilf 1976), maintained in the reduced form (GSH) by the enzyme GR, and in turn reduces other metabolites and enzyme systems, such as ascorbate. Due to its high concentration and its role in maintaining redox state in the cells, it is considered one of the most important cellular antioxidants. (Outline of the mechanism of enzymatic antioxidants in the removal of free radical is shown in Fig. 6.2.) Nonenzymatic Antioxidants

They are a class of the antioxidants which are not found in the body naturally but are required to be supplemented for the proper metabolism (Raygani et al. 2007). Some of the known nonenzymatic antioxidants are minerals, vitamins, carotenoids, polyphenols, and other antioxidants as listed below.


Minerals are required in the body cells for the proper functioning of the enzymes. Their absence is known to affect the metabolism of many macromolecules. They include selenium, copper, iron, zinc, and manganese. They act as cofactors for the enzymatic antioxidants.

Iron (Fe) Iron is the most abundant trace metal found to bound with protein in the biological system. Normally the concentration of free iron is very low and the low concentrations of iron-binding proteins promote ROS production, lipid peroxidation, and oxidative stress (Dabbagh et al. 1984). Hence iron supplementation helps in reducing the oxidative stress.

Magnesium (Mg) Magnesium is a cofactor for glucose-6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD) involved in pentose cycle which catalyzes the production of NADPH from NADP during the glucose metabolism and hence maintains the normal ratio of GSH to GSSG and a normal redox state in cells. Deficiency of magnesium reduces GR activity and GSSG does not reduce to GSH, hence causing oxidative damage to the cells (Fang et al. 2002).

Selenium (Se) Selenium is also a very important component of enzymatic antioxidant. In the presence of selenium (Se), glutathione peroxidase (GPx) plays a protective role against oxidation of lipid and protects the cell membrane and takes part in H2O2 and lipids' hydroxyperoxide metabolism. Hence, Se behaves like vitamin E and can be substituted in place of vitamin E and is used to prevent the risk of cancer and cardiovascular diseases (Sikora et al. 2008).

Copper (Cu), Zinc (Zn), and Manganese (Mn) SOD is a class of enzyme that consists of different types of SODs, depending upon their metal cofactor such as Cu–Zn and Mn. Cu–Zn SOD is found in the cytosol having Cu and Zn at their active sites which helps in proton conduction, whereas Mn-SOD is found in mitochondria and has Mn at its active site. These metals are responsible for SOD's antioxidant activities.


Vitamins form the class of micronutrients required for the proper functioning of the body's antioxidant enzyme system, such as vitamin A, vitamin C, vitamin E, and vitamin B. They cannot be synthesized in our body and hence need to be supplemented in the diet.

Vitamin A Vitamin A is helpful in night vision and in maintenance of epithelial cells in mucus membranes and skin. Because of its antioxidant properties, it assists immune system also and is found in three main forms: retinol, 3,4-didehydroretinol, and 3-hydroxyretinol. The main sources of this include sweet potatoes, carrots, milk, egg yolks, and mozzarella cheese.

Vitamin C Vitamin C is water soluble and is also called as ascorbic acid. It is found in fruits (mainly citrus), vegetables, cereals, beef, poultry, fish, etc. It is helpful in preventing some of the DNA damage caused by free radicals, which may contribute to the aging process and the development of diseases, such as cancer, heart disease, and arthritis.

Vitamin E Vitamin E is a lipid-soluble vitamin. This consists of eight different forms such as α-, β-, γ-, and δ-tocopherol and α-, β-, γ-, and δ-tocotrienol. Most abundantly found in almonds, safflower oil, soybean oils, oil of wheat germs, nuts, broccoli, fish oil, etc., α-tocopherol possesses highest bioavailability and is the most important lipid-soluble antioxidant which reacts with the lipid radical and protects the membranes from lipid peroxidation; as a result, oxidized α-tocopheroxyl radicals are produced that can be recycled to the reduced form through reduction by other antioxidants, such as ascorbate and retinol.


Carotenoid consists of β-carotene, lycopene, lutein, and zeaxanthin. They are fatsoluble colored compounds found in fruits and vegetables. β-Carotene is found mostly in radish-orange-green color food items including carrots, sweet potatoes, apricots, pumpkin, mangoes, and cantaloupe along with some green and leafy vegetables, including collard greens, spinach, and kale. Lutein is abundant in green leafy vegetables such as collard greens, spinach, and kale (Hamid et al. 2010). Lutein is best known for its role in protection of retina against harmful action of free radicals and also prevents atherosclerosis (Sikora et al. 2008).

Although lycopene, lutein, canthaxanthin, and zeaxanthin do not possess provitamin A activity, β-carotene is known as a precursor for vitamin A (Fang et al. 2002). Tomato is a good source of lycopene and spinach is a good source of zeaxanthin. It has been shown that lycopene is a potent antioxidant and is the most effective compound in removing singlet oxygen found in tomatoes, watermelon, guava, papaya, apricots, pink grapefruit, and other foods.


Polyphenols is a class of the phytochemicals that possess marked antioxidant activities. Their antioxidant activities depend on their chemical and physical properties which in turn regulates the metabolism depending on their molecular structures (Ajila et al. 2011). These consist of phenolic acids, flavonoids, gingerol, curcumin, etc. (Amit and Priyadarsini 2011).

Flavonoid is a major class of polyphenolic compound and is mostly found in vegetables, fruits, grains, seeds, leaves, flower, bark, etc. Some of the spices, such as ginger and turmeric, are also good sources of polyphenolic compound, e.g., gingerol is obtained from the rhizomes of ginger, whereas curcumin (diferuloylmethane) is the main bioactive component of turmeric and is known to possess good antioxidant activity. Curcumin is an excellent scavenger of ROS, such as O2 radicals, lipid peroxyl radicals (LO2 ), OH radicals, and nitrogen dioxide (NO2 ) radicals, which induced oxidative stress. Curcumin has been shown to inhibit lipid peroxidation and has been shown to increase GSH levels also in epithelial cells which lead to lower ROS production (Biswas et al. 2005).

Other Antioxidants

Transition Metal-Binding Proteins Albumin, ceruloplasmin, hepatoglobin, and transferrin are the transition metal-binding proteins found in human plasma, bind with transition metals, and control the production of metal catalyzed free radicals. Albumin and ceruloplasmin are the copper ion sequesters, hepatoglobin is hemoglobin sequester, and transferrin acts as free iron sequester.

Nonprotein Antioxidants Bilirubin, uric acids, and ubiquinol are nonprotein antioxidants which inhibit the oxidation processes by scavenging free radicals (Papas 1998).

Bilirubin Bilirubin is an end product of heme catabolism. It is a lipid-soluble cytotoxic product that needs to be excreted. However, bilirubin efficiently scavenges peroxyl radical at micromolar concentrations in in vitro model (Stocker et al. 1987) and is regarded as the best antioxidant against lipid peroxidation.

Uric Acid Uric acid is a powerful antioxidant and is a scavenger of singlet oxygen and radicals. Urate reduces the oxo-heme oxidant formed by peroxide reaction with hemoglobin and protects erythrocytes from peroxidative damage. The plasma-urate levels in humans are about 300 μM, making it one of the major antioxidants in humans (Ames et al. 1981).

Coenzyme Q Coenzyme Q is also known as ubiquinol (Co Q) and is an oil-soluble antioxidant. This is produced in the body through monovalent pathway, in heart, liver, kidney, pancreas, etc. The mechanism of the action may occur in two ways:

In the first mechanism, reduced form of ubiquinol (CoQH) acts as chain-breaking antioxidant and reduces peroxyl (ROO.) and alcoxyl radicals (LO.) (Papas 1998) (6.5 and 6.6).

CoQH + ROO. ® Q. + ROOH (6.5)

In the second mechanism, it reacts with vitamin E radical (TO.) and regenerating vitamin E.

CoQH + TO. ® Q. + ROOH (6.6)

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