*5.2.1. Enzymatic antioxidants*

The enzymatic antioxidants include superoxide dismutase (SOD), catalase, glutathione reductase (GRx) and glutathione peroxidase (GPx).

**Superoxide dismutase (SOD):** SOD is an enzymatic antioxidant that exists in three forms in mammalian tissues and differs on their cofactor, subcellular location and tissue distribution. 1. Copper zinc superoxide dismutase (CuZnSOD) is present in the cytoplasm and organelles of almost all mammalian cells [43]. This enzyme has a molecular mass of about 32,000 kDa with two protein subunits, each containing a catalytically active copper and zinc atom. 2. Manganese superoxide dismutase (MnSOD) has a molecular mass of 40,000 kDa and is found in the mitochondria of almost all cells [44]. It consists of four protein subunits, each containing a single manganese atom. 3. Extracellular superoxide dismutase (ECSOD) is a secretory copper and zinc containing SOD which is different from CuZnSOD [45]. It is synthesized only in fibroblasts and endothelial cells and expressed on the cell surface where it binds to heparan sulfates. Following its release from heparin, it is secreted into extracellular fluids and enters into the circulation. Superoxide dismutase catalyzes the dismutation of superoxide to hydrogen peroxide:

$$\rm O\_2^- + O\_2^- + 2H^+ \rightarrow H\_2O\_2 + O\_2.\tag{11}$$

The hydrogen peroxide can then be removed by catalase or glutathione peroxidase.

**Catalase:** Catalase was the first antioxidant enzyme to be characterized. It is located mostly within the peroxisomes of cells which contain most of the enzymes capable of generating hydrogen peroxide. It consists of four protein subunits, each containing a haem group and a molecule of NADPH [46]. Catalase is mostly present in liver and erythrocytes showing the greatest activities but is found in other tissues. It catalyzes the conversion of hydrogen peroxide to water and oxygen in two stages:

Stage 1: Catalase–Fe(III) <sup>+</sup> H2 O2 <sup>→</sup> compound <sup>I</sup>

Stage 2: Compound <sup>I</sup> <sup>+</sup> H2 O2 <sup>→</sup> catalase–Fe(III) <sup>+</sup> 2H2 <sup>O</sup> <sup>+</sup> O2

**Glutathione peroxidases (GPx):** Glutathione peroxidase is an enzyme which is synthesized mainly in the kidney and found in almost all tissues although it is highly found in the liver [47]. Its subcellular location is usually the cytosol and mitochondria. Selenium serves as its cofactor located at the active site of the enzyme and deficiency of selenium greatly affects the activity of the enzyme [48]. Glutathione peroxidases catalyze the oxidation of reduced glutathione (GSH) decomposing hydrogen peroxide or another species such as a lipid hydroperoxide:

$$\text{ROOH} + 2\text{GSH} \rightarrow \text{GSSG} + \text{H}\_2\text{O} + \text{ROH}.\tag{12}$$

Competing pathway that utilizes NADPH such as the aldose reductase pathway may lead to a deficiency of reduced glutathione thereby limiting the action of glutathione peroxidase.

Free Radicals and the Role of Plant Phytochemicals as Antioxidants Against Oxidative Stress-Related Diseases

http://dx.doi.org/10.5772/intechopen.76719

59

The non-enzymatic antioxidants are usually low-molecular-weight antioxidant (LMWA) compounds capable of preventing oxidative damage either by directly interacting with ROS or indirectly by chelating metals [50]. Transition metals are directly chelated by some of this LMWA thereby preventing them from participating in metal-mediated Haber-Weiss reaction [51]. Other direct acting LMWA molecules scavenge free radicals by donating electrons to free radicals to make them stable thereby preventing attacks of biological targets. These LMWA molecules also called scavengers may be advantageous over enzymatic antioxidants as they can penetrate cellular membranes and be localized in close proximity to the biological target due to their small size. More so, these non-enzymatic antioxidants can interact together to scavenge free radicals and their scavenging activity may be synergic. Most scavengers originate from endogenous sources, such as biosynthetic processes and waste-product generation by the cell. However, the number of LMWA synthesized by the living cell or generated as waste products such as histidine dipeptides, glutathione, uric acid, lipoic acid and bilirubin is limited [52]. More so, the concentration of scavenger must be sufficiently high to compete with the biological target on the deleterious species [50]. As such, exogenous sources of nonenzymatic antioxidants especially from plant diet and phytochemicals are needed to supplement the endogenous non-enzymatic antioxidants. The oxidative stress defense mechanism

Plants have long been consumed as food which is rich in vitamins and other nutrients that are useful for the body. Also, various plants were used in folk medicine for various therapeutic purposes. Though these uses, the notion of plant as a source of antioxidant became more evident in recent time as oxidative stress was considered a major attribute to most diseases in humans and the antioxidant defense system in human was usually not sufficient to overcome the free radical level in the body. As such, plants have gained considerable interest as a source of antioxidants and so much research has been done to identify plants substances with

Like other humans, plants do have enzymatic and non-enzymatic antioxidant defense systems to protect them against free radicals. The enzymatic system includes catalase, SOD, glutathione peroxidase(GPx), and glutathione reductase (GRx) [7], while non-enzymatic systems consist of low molecular weight antioxidants (LMWA) such as ascorbic acid, proline, glutathione, carotenoids, flavonoids, phenolic acids, etc. and the high molecular weight antioxidants (HMWA) which are mostly secondary metabolites such as tannins [53]. The possible reason for the presence of these antioxidants in plants is that plants lack an immune system unlike animals thus, utilize the antioxidant defense system to protect them against microbial pathogens and animal herbivores. Also, these phytochemicals serve as a defense system against environmental stress.

*5.2.2. Non-enzymatic antioxidants*

in humans is summarized in **Figure 2**.

antioxidant activities.

**6. Plants as source of antioxidants**

The fact that GPx also acts on lipid hydroperoxides suggest it may be involved in repairing cellular damages due lipid peroxidation [49]. The activity of GPx is dependent on the constant availability of reduced glutathione which is regenerated from oxidized glutathione (GSSG).

**Glutathione reductase** (GRx): GRx is a flavine nucleotide dependent enzyme and has a similar tissue distribution to glutathione peroxidase [49]. The role of GRx is to generate GSH from GSSG using NADPH in order to increase the ratio of reduced to oxidized glutathione:

$$\text{GSSG} + \text{NADPH} + \text{H}^\* \rightarrow 2\text{GSH} + \text{NADP}^\*.\tag{13}$$

The NADPH required by this enzyme to replenish the supply of reduced glutathione is provided by Glucose-6-phosphate dehydrogenase (G-6-PD) in the pentose phosphate pathway.

**Figure 2.** Oxidative stress defence mechanism.

Competing pathway that utilizes NADPH such as the aldose reductase pathway may lead to a deficiency of reduced glutathione thereby limiting the action of glutathione peroxidase.
