Page2239 Indo American Journal of Pharmaceutical Research, 2013 ISSN NO: 2231-6876 Journal home page: http:///index.php/en/ INDO AMERICAN JOURNAL OF PHARMACEUTICAL RESEARCH PHYTONUTRIENT COMPOSITION AND ANTIOXIDANT ACTIVITY OF PROCESSED FLAXSEED Dharshini¹, Dr.S.Sumayaa², Dr.T.Thirunalasundari ³ 1 Research Scholar, Department of Food Science and Nutrition, Thassim Beevi Abdul Kader College for Women, Kilakarai, Tamil Nadu, India. Email:shivibio@gmail.com. 2 Head of the Department, Department of Biotechnology, Bharathidasan University, Tiruchirappali, Tamil Nadu, India. 3 Principal, Thassim Beevi Abdul Kader College for Women, Kilakarai, Tamil Nadu, India. ARTICLE INFO Article history Received 21/03/2013 Available online 02/02/2013 Keywords Flaxseed (FS), antioxidant, phytonutrient Corresponding author Ms.Dharshini ABSTRACT Flax is a nutraceutical with many nutritional benefits. Aside from the vitamins, minerals, fiber and protein it contains some primary and secondary metabolites. Flaxseed is a good source of omega-3 fatty acids. Through its high concentration of alpha-linolenic acid, flax provides more omega-3 fatty acids than any other grain. (57% of its fatty acids are omega-3s.) As the health revolution continues to evolve, look for it to be added to more and more processed foods to attain the health smart claim. Flaxseed (FS), a whole grain used as a nutritional supplement, has gained popularity because it is a rich source of natural antioxidants. Dietary intake of polyphenols has been associated to reduce risk of chronic diseases, such as heart disease, stress induced diabetes and cancer. This is probably due to their antioxidant properties. Many phenolic compounds have been detected in Flaxseed. Hence with the objectives to analyze the phytonutrient composition, assess the non-antioxidant enzymes vitamin A, E and C and find the antioxidant activity of processed flaxseed the study was carried out. The flaxseed was subjected to various methods of food processing like roasting, pressure cooking and soaking. The processed flaxseed was further analysed the primary and secondary metabolites and their antioxidants activity. This study does not show much variation in primary and secondary metabolites between the raw and roasted seeds. The acceptance and palatability was high in the case of roasted soybean and flaxseed because of their nutrients nutty flavour, colour and texture than the unprocessed seeds. Nonantioxidant enzymes, vitamin A, vitamin E and vitamin C content was high in both soaked and roasted flaxseed. Antioxidant activity was high in roasted and soaked flaxseed. Thus the presence of these phytonutrient and antioxidant activity of processed namely roasted and soaked flaxseeds indicates that, it has a potential of being used in formulations prepared for food consumption. Flaxseed could also be a remedy for the major killer diseases like cancer, diabetes and cardiovascular diseases. Research Scholar, Department of Food Science and Nutrition, Thassim Beevi Abdul Kader College for Women, Kilakarai, Tamil Nadu, India. Email:shivibio@gmail.com. contact Number :+91 9786674700. Please cite this article in press as Ms.Dharshini. et.al. PHYTONUTRIENT COMPOSITION AND ANTIOXIDANT ACTIVITY OF PROCESSED FLAXSEED. Indo American Journal of Pharm Research.2013:3(2). Copy right 2013 This is an Open Access article distributed under the terms of the Indo American journal of Pharmaceutical Research, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Page2240 INTRODUCTION The health care of the world s population is likely to undergo dramatic change due to the ongoing demographic transition. The prevalence of degenerative diseases is on rise in an exponential rate. The prevalence of type II diabetes in India will increase to 87 million by 2030 [1] and prevalence of coronary heart disease and cardiovascular diseases in India will increase to 2.58 million and 4.77 million by 2020 respectively [2]. The risk of cancer in India has increased from 40 to 70 percent, due to various factors like diet, pollution and life style [3].There is a way out to solve this increasing risk of degenerative diseases through modification in life style and simple changes in diet by inclusion of natural food. The prevalence of these diseases can be reduced to a greater extent through the incorporation of natural plant sources, rich in bioactive compounds. One such natural resource is flaxseeds which is rich in bioactive compounds and is nutritionally beneficial to human body in controlling various degenerative disorders. There are different methods by which flaxseeds are processed and consumed. Traditional processing methods such as germination and roasting could effectively improve the nutritional value of seeds [4]. The use of some processing methods such as soaking, boiling, pressure cooking and autoclaving are known to achieve reduction or elimination of the anti-nutritional factors, which affect the nutritional, food and acceptability quality of seeds [5].Roasted flaxseeds are considered to be one of the few health foods that have phenomenal nutritional value. Roasted flaxseeds are usually the refined version of natural seeds that are healthier and better in taste [6]. Therefore the best method of processing which will enhance proximate nutrient, phytonutrient composition, antioxidant activity and sensory acceptability can be used in food formulations. Hence with the objectives to estimate the proximate nutrients, analyze the phytonutrient composition, assess the nonantioxidant enzymes vitamin A, E and C and find the antioxidant activity of processed flaxseed, the study was carried out. Methodology Flaxseeds were purchased from local markets of Coimbatore. The purchased seeds were cleaned in shade dried and stored in air-tight containers. Processing of Flaxseed The flaxseed was subjected to various methods of food processing like roasting, pressure cooking and soaking. Flaxseeds were finely powdered using mechanical blender and used as raw flour. Dry heat method was adopted to roast flaxseed until a desired colour and flavour was achieved. Soaked flaxseeds were pressure cooked for 20 minutes and then they were made as a paste using mechanical blender. Flaxseeds were soaked in soft water for 12 hours and the excess water was decanted. Analysis of the phytonutrient composition of processed flaxseed The samples were used for phytonutrient screening, comprising chemical tests to detect the presence of flavonoid, alkaloid, saponin, tannin, glycoside, terpenoid, steroid, phlobatannin and anthroquninone using Harborne, (1973).The samples were used for phytonutrient screening comprising chemical tests to the quantify the of flavonoid, alkaloid, total phenol and tannin content using standard procedures. The procedures used for the analysis are given in Table-1. Determination of non-antioxidant enzymes, Vitamin-A, E and Vitamin C in the processed flaxseed Processed flaxseed was subjected to analysis of non antioxidant enzymes vitamin A, E and vitamin C using standard procedures. Methods used are given in Table-2. Determination of Antioxidant activity of the processed flaxseed Antioxidant activity was studied for processed flaxseed powder using 1,1Diphenyl-β-picrylhydrazyl (DPPH) radical scavenging method. DPPH method is extensively used for investigating the free radical activities of
Page2241 compounds. In DPPH test the processed flaxseed extracts were potentially able to reduce the alcoholic DPPH solution in the presence, of a hydrogen donating antioxidant due to the formation of the non radical form of DPPH-H by its reaction. Ascorbic acid was used as standard. Brand williams et al (2001) method was adopted for the study. Table 1: Methods followed for analysis of quantitative phytonutrients & non-antioxidant enzymes S.NO Phytonutrients and non Methods Followed antioxidant enzymes 1 Flavonoid (Bohm and Kocipai Abyasan, 1994) 2 Alkaloid (Harborne, 1973) 3 Tannin Spectrophotmetry (Schanderl (1970) 4 Total phenol Spectrophotometry, Abu Baker et al,2009) 5 Vitamin-A(BetaCarotene) Spectrophotometry (Bayfiedl, & Cole (1980) 6 Vitamin E(Tochopherol) Spectrophotometry (Rosenberg. (1992) 7 Vitamin-C Tirymetric method (Sadasivam & Theymoli, 1987) Results and discussion Phytonutrients Analysis of Processed Flaxseed Phytonutrient screening was done to understand the presence of alkaloids, flavonoids, saponins, terpenoids, steroids, tannins, cardio glycosides and glycosides [13]. The results of the qualitative analysis of phytonutrients are presented below in Table 2. Table 2: Qualitative phytonutrients Analysis of Processed Flaxseed Methods of Processed Flaxseed S.No Phytonutrients Raw Roasted Pressure cooked Soaked 1 Flavonoid ++ ++ + + 2 Alkaloid ++ ++ + + 3 Saponin ++ + - - 4 Tannin ++ + - - 5 Glycoside + + + + 6 Terpenoid ++ ++ + + 7 Steriod ++ ++ + + 8 Anthoquninone - - - -
Page2242 Qualitative phytonutrients analysis of the processed flaxseed revealed the presence of flavonoid, alkaloid, terpenoid, steroid and glycoside and they are present in all the category of processed flaxseeds. Whereas saponin and tannin was seen only in raw and roasted flaxseed and absent in pressure cooked and soaked flaxseed powder. It was also clear that phlobatannin and anthroquninone were not present in all forms of processed flaxseeds. These phytonutrients namely phenolic compounds, flavonoids, alkaloids and tannin possess diverse beneficial bioactivities including antiallergic, antiviral, antinflammatory and antimutagenic property [14]. Phytonutrient Composition of Processed Flaxseed The results of the quantitative phytonutrient analysis of processed flaxseed powder are presented below. The results of the phytonutrient composition namely alkaloid and flavonoids of raw, roasted, pressure cooked and soaked flaxseed powder are presented below in figure -1 Figure 1: Composition of flavonoids and alkaloid in processed flaxseed The quantitative estimation of the total flavonoid and alkaloid levels in raw, roasted, pressure cooked and soaked flaxseed powder is found to be 0.11mg & 0.28mg, 0.23mg & 0.12mg, 0.021mg & 0.05mg and 0.05 mg & 0.08 mg respectively. Phytochemicals derived from plant foods in the diet interfere disease progression by acting directly on cells as well as by modifying the cells microenvironment and creating physiologic conditions. Polyphenolic compounds, dietary flavonoids and tannin are recognized for their antioxidant and antiproliferative properties and also play an important role in reducing the risk of cancer, heart and neurodegenerative diseases [15]. Composition of total phenol and tannin in processed flaxseed The results of the phytonutrients namely total phenol and tannin composition of raw, roasted, pressure cooked and raw flaxseed powder are given below in figure 2. Figure 2: Phytonutrient Composition of Roasted Flaxseed
Page2243 The total phenolic and tannin content were found to be 47.0 mg/gallic acid equivalent & 20.2µg, 7.25mg/gallic acid equivalent/g & tannin 16µg, 6.34 mg gallic acid equivalent and 16.4 µg respectively in all the processed flax seeds. The amount of phytonutrients in roasted flaxseed is comparatively higher than the raw flaxseed powder. In general it is reported that tannin and total phenol are found to be high in seeds coats and contain very high amounts of tannins and flavonoids and such seeds possess a high antioxidant activity [16]. The quantitative phytonutrient estimation of the pressure cooked flaxseed reveals the contained total phenol. The amount of phytonutrients in processed pressure cooked has concededly decreased in the raw and roasting process. Poly phenols have been identified as important compounds in fighting cancer and heart disease. Pressure cooking of flaxseed has reduced phytonutrient content, yet they are not totally devoid of the phytonutrients. The total phenol and tannin content are higher than the flavonoids and alkaloids with respectively. The amount of phytonutrient have increased in soaking process than in pressure cooking [17].When flaxseeds were analyzed for the presence of qualitative phytonutrients constituents showed the presence of phytonutrients in methods of processing like soaking and roasting. Flavonoids are a group of compounds responsible for many of the flavor characteristics of foods. It reduces constriction of the arteries and lowers the risk of stroke. Thus from the qualitative estimation, it is clear that roasting and soaking are best methods of processing flaxseed and has considerably high amounts of flavonoids, total phenol and tannin. Non-Antioxidant Enzymes, Vitamin A, E and Vitamin C Content of Processed Flaxseed Vitamin A, E and vitamin C are regarded as antioxidants and play a vital role in overcoming oxidation and degenerative diseases. The presence of these vitamins in foods indicates that they can help in prevention of many diseases. Non-Antioxidant Enzymes, Vitamin A, E and Vitamin C Content of Raw Flaxseed Antioxidants are compounds that will quickly react with oxygen to form new substances. Antioxidants will react with the oxygen before lipids do. Important dietary antioxidants are vitamin A, E and vitamin C. The result of the non-antioxidant enzymes, vitamin A, E and C content in processed flaxseed are presented in figure-3. Figure 3: Non antioxidant Enzymes in Processed flaxseed It is clear from the figure-3 that considerable amount of non-antioxidant enzymes Vitamin A, E and vitamin C are present in raw flaxseed powder. It was found that raw flaxseed has 10.12 IU of vitamin A, 2.18 µg of Vitamin E and 0.013 mg of Vitamin C [18].Flaxseed is rich beta-carotene, vitamin E and vitamin C. It is also
Page2244 reported that supplementation of flaxseed based mid-morning snacks on school children increased brain development and activities, due to the presence of these vitamins. Natural antioxidant such as a- tochopherol, b- carotene and vitamin C found in higher plants are being used in the food industry to inhibit lipid per oxidation and they can protect the human body from free radical and retard progress of many chronic disease [19]. Antioxidants in food contribute to disease prevention, which are essential nutrients or precursors of nutrients. Each of these antioxidant nutrients has specific activities and they often work synergistically to enhance the antioxidant capability of the body. The amount of the roasted flaxseed powder reveals that it had 7.915 IU of vitamin A, 1.772 µg of vitamin E and 0.009mg of vitamin C. It is also clear that roasting has increased the vitamin contents of the flaxseed [20].The synergistic action of wide spectrum of antioxidant is better than the activity of a single antioxidant and that antioxidants from natural source have a higher bioavailability and therefore higher protective efficacy against oxidative stress than synthetic antioxidants. Thus the mixtures of all three antioxidant enzymes are present in roasted flaxseed and may be believed to play a vital role in human body. The amount of non antioxidant enzymes, Vitamin A was found to be 5-51 IU, vitamin E was 1.652µg and vitamin C was 0.002 mg in pressure cooked flaxseed powder. The levels have been greatly decreased in pressure cooking, whereas roasting it had increased. Several compounds from the seeds show pharmacological activities and discovery of bioactive compounds, primarily for deadly diseases like cancer and diabetes, while other compounds have been developed. To treat inflammation [21-22].Dietary antioxidants, especially β-carotene, may be important in the prevention of stomach and breast cancer. From the data it is clear that considerable amount of non-antioxidant enzyme vitamins are present in soaked flaxseed. It was found that soaked flaxseed has 8.066 IU of vitamin A, 2.02µg of vitamin E and 0.005 of vitamin C. The levels of non-antioxidant vitamins are better than pressure cooked flaxseeds. Vitamin C increases HDL levels and may also lower cholesterol in the blood, the reducing the risk of cardiovascular diseases. Vitamin E is associated with improvements in arthritis. Antioxidants (Vitamin C, vitamin E and Carotenoids ) have been associated with decreased cataract risk through the retardation of lens opacity. Antioxidant Activity of Processed Flaxseed Antioxidants have many industrial uses, such as preservation in food and medicinal value. Antioxidants are manufactured within the body and can also be extracted from the food. The antioxidant activity of processed flaxseed are presented in Figure-4 Figure 4: Non-Antioxidant Enzymes, Vitamin A Content of Processed Flaxseed
Page2245 The DPPH assay was used as a rapid method to detect the processed flaxseeds for their antioxidant capacity. The results also indicate a positive DPPH radical scavenging activity for roasted flaxseed, pressure cooked, soaked and raw flaxseed. The radical scavenging activity was 35 percent for raw flaxseed powder, 76 percent for roasted flaxseed powder, 69 percent for pressure cooked flaxseed powder and 76 percent for soaked flaxseed powder. In the present study it is clear that the concentration of phytonutrient is increasing the antioxidant capacity. The phytonutrient content of roasted and soaked flaxseeds is comparatively high and also their antioxidant activities are high. The antioxidant activity of the standard ascorbic acid had a high scavenging capacity of 94 percent, which is higher than flaxseed. Phytonutrients that have generated great interest due to their antioxidant capacity, have chemopreventive effect in humans have a great influence on the stability of oxidation present in food. Phenolic compounds and other phytonutrients have high antioxidant power and also influence the organoleptic properties of food [23].Thus it can be concluded from the study that roasted and soaked method of processing can be adopted for food formulation. Summary and conclusion Eating trends of natural food, sources have been augmented to bring health and vitality, one such natural resource is flaxseeds. Flaxseeds offer untapped plethora of health benefits. The long recognized traditional health benefits of flaxseed are now being confirmed by modern scientific research. Flaxseeds are being used for its rich phytonutrient content and antioxidant property in treating major degenerative and deficiency diseases. Qualitative phytonutrient Analysis of Processed Flaxseed It was found that flavonoid, alkaloid, terpenoid, steroid and glycoside are present in all the category of processed flaxseed. Whereas saponin and tannin was seen only in raw and roasted flaxseed and absent in others. Phlabatannin and antharoquinone were not present in the processed flaxseeds. Quantitative phytonutrient Analysis of Processed Flaxseed Flavonoid content of the processed flaxseed ranged from 0.005 0.12mg with roasted flaxseed having the high amount of flavonoid (0.12 mg) and with soaked flaxseed having the low amount of flavonoid (0.005 mg). Alkaloid content was high in raw flaxseed with (0.23mg) and least amount in pressure cooked with 0.05 mg. High content of total phenol was found in roasted and raw flaxseed with 47mg gallic acid equivalent and low content of total phenol was found in soaked flaxseed, with 6.34 mg gallic acid equivalent. Non Antioxidant enzymes, Vitamin A, E and vitamin C Content of Processed Flaxseed Vitamin A content was high in raw flaxseed with 10.12 IU and low amount in pressure cooked flaxseed with 1.652 IU. Vitamin E content of the flaxseed was found to be in the range of 1.652 2.18 µg with the least amount in pressure cooked flaxseed (1.652 µg) and the highest amount in (2.18µg) raw flaxseed powder. Vitamin C content of the processed flaxseed ranged from 0.002-0.013 mg with raw flaxseed having the high amount of vitamin C (0.013mg) and (0.002mg) in pressure cooked having the least amount of vitamin C. Antioxidant activity of Processed Flaxseed: The results of the antioxidant activity of the flaxseeds showed a positive DPPH radical scavenging activity. Raw flaxseed extracts had 35 percent; pressure cooked flaxseed
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