1 INTRODUCTION. Page 1

Transcription

1 INTRODUCTION

2 1 INTRODUCTION You are what you eat. Diet is an integral part of human life which indirectly reflects the physical and emotional state of a person. There is a huge diversity in food patterns worldwide. Although, these diversities are reflected in the typical diets of different states of India, any traditional Indian meal includes all the nutrients required to make a balanced meal. Moreover, it also comprises of other non-nutrient beneficial elements like antioxidants. Indian diet is thus a wholesome and powerful diet in terms of nutrition, medicinal as well as antioxidant properties. These antioxidant properties protect us from the chronic diseases which are gift of globalization and urbanization that has made undesirable changes in environment, diet as well as lifestyle. The term antioxidant refers to any molecule capable of stabilizing or deactivating free radicals before they attack cells. Halliwell and Gutteridge, (1989) have defined antioxidants as substances that are able, at relatively low concentrations, to compete with other oxidizable substrates and, thus, to significantly delay or inhibit the oxidation of these substrates. Humans have evolved highly complex antioxidant systems (enzymatic and non-enzymatic), which work synergistically, and in combination with each other. The most efficient enzymatic antioxidants involve glutathione peroxidase, catalase and superoxide dismutase (Mates et al, 1999). Non-enzymatic antioxidants include Vitamin E and C, thiol antioxidants (glutathione, thioredoxin and lipoic acid), melatonin, carotenoids and phenolic compounds (McCall and Frei, 1999). Antioxidants act at the levels of prevention, interception and repair. Preventive antioxidants attempt to stop the formation of ROS. These include superoxide dismutase (SOD) that catalyzes the dismutation of superoxide to H 2 O 2 and catalase that breaks it down to water (Sies, 1996; Cadenas and Packer, 1996). Interception of free radicals is mainly by radical scavenging, while at the secondary level scavenging of peroxyl radicals are affected. The Page 1

3 effectors include various antioxidants like vitamins C and E, glutathione, other thiol compounds, carotenoids, flavonoids, etc. At the repair and reconstitution level, mainly repair enzymes are involved (Sies, 1996; Cadenas and Packer, 1996; Halliwell and Aruoma, 1993). Antioxidant systems work together to protect the cells and organ systems of the body against free radical damage. Free radicals can be defined as reactive chemical species having a single unpaired electron in an outer orbit (Riley, 1994). This unstable configuration creates energy which is released through reactions with adjacent molecules, such as proteins, lipids, carbohydrates, and nucleic acids. Free radicals damaging biological systems are reactive oxygen species (ROS) which are oxygen-free radicals. These are the main byproducts formed in the cells of aerobic organisms, and can initiate autocatalytic reactions so that molecules to which they react are themselves converted into free radicals to propagate the chain of damage. ROS are produced both endogenously and exogenously. The endogenous sources of ROS include mitochondria, cytochrome P450 metabolism, peroxisomes, and inflammatory cell activation (Inoue et al, 2003). ROS can be (i) generated during UV light irradiation and by X-rays and gamma rays (ii) produced during metal catalyzed reactions (iii) are present in the atmosphere as pollutants (iv) are produced by neutrophils and macrophages during inflammation and (iv) are by-products of mitochondrial catalyzed electron transport reactions, and various other mechanisms (Cadenas, 1989). Reactive oxygen species can be either free radicals, such as the superoxide anion radical (O 2 ) and the hydroxyl radical (HO ), or singlet molecular oxygen ( 1 O 2 ) (Halliwell and Gutteridge, 1993). Singlet molecular oxygen produced as a consequence of absorbed light energy is not a free radical (chemical species capable of independent existence with one or more unpaired electrons), but instead has an excited unpaired electron with antiparallel spin in its outer orbital. It is, however, a very reactive oxygen species and may be especially significant source of potential oxidative damage Page 2

4 in people exposed to long periods of intense sunlight [ultraviolet (UV) light exposure] during work in an outdoor environment (Halliwell and Gutteridge, 1993). Hydrogen peroxide (H 2 O 2 ) is not a free radical, but a two-electron reduction product of oxygen that in the presence of iron (Fenton reaction) can give rise to the highly reactive hydroxyl radical (HO ) (Halliwell and Gutteridge, 1993). Reactive oxygen species can interact destructively with many cellular structures, especially cell and organelle membrane structures (Halliwell and Gutteridge, 1993; Jenkins and Goldfarb, 1993) causing lipid peroxidation (Hochstein and Ernster, 1963), DNA damage (Kasai et al, 1986), and protein degradation (Griffith et al, 1988). Excess of free radicals in a system leads to condition called oxidative stress. Oxidative stress is the sum of all the oxidative reactions resulting from such diverse biological processes as aerobic metabolism, inflammation, radiation damage, aging, carcinogenesis, and photobiological effects (DiMascio et al, 1991). Oxidative damage to proteins, DNA and lipids increases with age (Sohal and Weindruch 1996). Free radicals are also considered to play a casual role in the process of cardiovascular diseases also. (Schachinger and Zeiher, 2002). ROS lead to the oxidation of low density lipoprotein (OxLDL), and this accumulates within plaques, and contributes to the inflammatory state of atherosclerosis and plays a key role in its pathogenesis (Galle et al, 2006). Oxidized-LDL leads to endothelial dysfunction, and can result in either cell growth or apoptotic cell death and can cause vasoconstriction. Oxidative damage may also play a role in amyloid deposition in Alzheimer s disease. Parkinson s Disease has also been found to be associated with increased oxidative damage to DNA (Migliore et al, 2002) proteins (Choi et al, 2006) and lipids (Agil et al, 2006). Considering the above mentioned health hazards caused by oxidative stress, the role of antioxidant becomes very crucial in human health for protection Page 3

5 against life style diseases. Therefore, the measurement of antioxidant compounds and their antioxidant capacity has become an interesting area of research in food science. Hundreds of food items have been measured for their antioxidant capacity using various methods. Several extraction techniques have been reported regarding chemical extraction of dietary antioxidants. Methanol, acetone, ethyl acetate and chloroform are the main solvents used for extraction of low molecular weight polyphenols (Nakamura et al, 1998; Rubilar et al., 2006; Vrhovsek et al, 2004). Carotenoids and vitamin E are principally extracted with high apolar solvents (Castan et al, 2005; Larsen & Christensen, 2005), while vitamin C is extracted with polar solvents (Frenich et al, 2005). Because of the complexity and diversity of antioxidant and food matrices, nowadays there is no optimal method or solvent used to extract all antioxidants present in foods. Since dietary antioxidants may have different solvent affinities, some studies analyze lipophilic and hydrophilic antioxidants separately to assess antioxidant capacity but may not assess the synergistic effects of these two kinds of antioxidants. However, these methods do not imply to the fate of antioxidants in physiological conditions and chemical environment. Vast difference exists between the antioxidant potential of the food as extracted by conventional methods and the actually bioavailable antioxidants from food. Also, most research papers estimate the antioxidant properties of raw food material though they are seldom consumed so. Cooking brings about a number of changes in the physical characteristics and chemical composition of foods. Food processing steps such as dehulling, peeling, thermal processing, mashing, etc. contribute to degradation and loss of phenolic compounds (Howard et al, 1999; Hunter and Fletcher, 2002). Cooking sets the phenolic compounds free from these linkages to make them more bioaccessible. Reports have been found that thermal processing disrupted the plant cell membranes and cell walls and released phytochemicals from the insoluble Page 4

6 portion of the broccoli, which increased the pool of phenolic compounds (Roy et al, 2009). During digestion, phenolic compounds are released from the matrix after mastication. In the stomach, gastric glands secrete pepsin, gastrin hormone and hydrochloric acid for protein digestion. Pancreatic juice contains proteolytic enzymes, amylase for starch and glycogen hydrolysis, lipase for neutral fat digestion, phospholipases which split fatty acids from phospholipids, and cholesterol esterase. Bile salts and sodium bicarbonate are secreted into the duodenum from the biliary gland of the liver (Guyton and Hall 1996b). All these factors help the release of phenolic compounds which are present in bound form with carbohydrates and proteins in food. The biological properties of antioxidants depends on the release of phenolic compounds from the food matrix during the digestion process (bioaccessibility) and may differ quantitatively and qualitatively from those produced by the chemical extraction employed in most studies (Serrano et al, 2007). Perez-Jimenez and Saura-Calixto (2005) have stated that the antioxidant capacity of foods may be underestimated in the literature because the extraction solvents usually used do not allow a complete release of antioxidant compounds and additionally nonextractable polyphenols with a high antioxidant capacity are ignored. In vitro GI models have been developed for bioaccessibility assessment, allowing the study of changes in dietary components during the gastric and the intestinal stage, due to factors impacting their availability discussed above (Cilla et al, 2009; McDougall et al., 2005). The transition from the acidic gastric to the mild alkaline intestinal environment caused a decrease in the amount of bioaccessible total polyphenols, flavonoids and especially anthocyanins. Tagliazucchi et al, (2010) have also shown that radical-scavenging activities of polyphenols may be ph-dependent, suggesting greater scavenging capacity in Page 5

7 the intestine than in the stomach. Much data has been generated on polyphenol constituents in a variety of food items measuring total phenolics, flavonoids and related total antioxidant activities based on chemical extraction, typically using methanol or water/methanol mixtures (Lamperi et al, 2008; Lee et al, 2003; Neveu et al, 2010; Wojdylo et al, 2008). However, under in vivo conditions, polyphenols from the diet have to be extracted following gastrointestinal (GI) digestion. The nature of extractable phytochemicals, their stability and their antioxidant activity depend on many factors, such as the food matrix, the ph, the temperature, presence of inhibitors or enhancers of absorption, presence of enzymes, host, and other related factors (McDougall et al, 2005; Saura-Calixto et al, 2007; Tagliazucchi et al, 2010). The GI tract may be considered as an efficient extractor, where part of the phytochemicals contained in food matrices is extracted and becomes available for uptake in the intestine (Saura-Calixto et al, 2007; Tagliazucchi et al, 2010). All the above mentioned factors make chemical extraction a non-implacable and non-reliable method for the estimation of antioxidant capacity and their bioaccessibility. Spanish and Mediterranean diet have been thoroughly studied for their antioxidant potential, not only by conventional extraction but also by in vitro gastrointestinal digestion. No study has been so far done to evaluate the antioxidant potential of traditional Indian diet either by conventional or physiological extraction. The present study is thus an investigation of the traditional Indian diet for its antioxidant compounds as well as antioxidant capacity with following objectives: i. To determine the Total Antioxidant Capacity of the raw and cooked staple foods of India by different chemical methods. Page 6

8 ii. iii. To determine the Total Antioxidant Capacity of the raw and cooked staple foods of India by using a physiological approach i.e. after extraction using an in vitro gastrointestinal model. To calculate the Total Antioxidant Intake of the population of India. Page 7