PhytochemicalChange and Antioxidant Activities of Cereals Miss Waramporn Vongsudin Division of Biochemical Technology School of Bioresources and Technology King Mongkut s University of Technology Thonburi http://en.wikipedia.org/wiki/cereal Cereals Cereals were germinated by soaking in water, and are then halted from germinating further by drying with hot air. cereals is popular in Food&Beverage and Cosmetics industry to value-added of products. 1
Oral Presentation CRDC 5 22/07/54 Cereals Products Office of Agricultural Economics, 29 2
Cereals Cereal is an annual crops in the grass family (Gramineae) Seeds being utilized. http://en.wikipedia.org/wiki/cereal Endosperm Carbohydrates Proteins Essential fatty acid Bran Fiber Minerals Germ Vitamins Minerals Phytonutrient Structure of Cereals http://www.cemmitrading.com/article1.html.html 3
Phytonutrient Phytochemicals are a natural bioactive compound that occur naturally in plants that plant produce these chemicals to protect itself. Primary metabolite Secondary metabolite http:// ://en.wikipedia.org/wiki/phytochemical Primary metabolite http://www.maceducation.com 4
Secondary metabolite Nutritional content of cereal grains Primary metabolite contents Cereals Brown rice Wheat Barley Rye Oats Sorghum Soybean Sesame Protein (%) 9.2 9.1 12.0 11.5 13.4 17.1 11.0 34.0 20.0 Lipid (%) 2.5 4.4 0.8 2.2 1.8 6.4 3.2 18.0 45.0 Carbohydrate (%) 85.9 81.8 78.4 77.8 80.7 62.0 81.3 30.0 13.5 Sowa et, al., 1992; ; Tester et, al., 24 5
Nutritional content of cereal grains Secondary metabolite contents Cereals Brown Wheat Barley Rye Oats Sorghum Soybean Sesame rice Phenolic compound γ - oryzanol - - - GABA - - - Isoflavone - - - - - - - - Ferulic acid Antioxidants : anti-cancer, anti-aging aging Sowa et, al., 1992; ; Tester et, al., 24 Rice Bran Fiber Minerals Endosperm Carbohydrates Proteins Essential fatty acid Germ Minerals GABA γ - oryzanol Charalampopoulos et, al., 22 6
Carbohydrates Proteins Fiber Minerals Essential fatty acid Minerals Ferulic acid Endosperm Bran Germ Ou et, al., 24 Soybean Seed coat Fiber Minerals Embryo Minerals Isoflavone Endosperm Reference:kidney beans Carbohydrates Proteins Essential fatty acid Ohta et, al., 20 7
Sesame Endosperm Carbohydrates Proteins Essential fatty acid Seed coat Fiber Minerals Embryo Minerals http://learntanii.blogspot.com Anti-Nutrient content of cereal grains Cereals Brown rice Wheat Barley Rye Oats Sorghum Soybean Sesame Phytic acid Trypsin inhibitor Oxalate - - - 8
Germination Epigeal Germination Cotyledon http://theseedsite.co.uk/germinating.html 9
Hypogeal Germination Germination Cotyledon http://theseedsite.co.uk/germinating.html When the seed imbibes water, hydrolytic enzymes are activated which break down these stored food resources into metabolically useful chemicals. Water content (%) Plants/Grains Author 30-35% Rice, soybean, oat Upreti et, al. (24) 50-55% sesame, peanut Menacho et, al. (2010) Oxygen Oxygen is required by the germinating seed for metabolism. Oxygen is used in aerobic respiration, the main source of the seedling's energy until it grows leaves. 10
Temperature Temperature affects cellular metabolic and growth rate. Temperature ( C) Plants/Grains Author 10-35 C Rice, wheat Wang et, al. (24) 27-30 C sesame Guiga et, al., (28) 30-34 C corn, soybean Li et, al. (28) Light or darkness Light or darkness can be an environmental trigger for germination and is a type of physiological dormancy. http://en.wikipedia.org/wiki/germination Related Research Author Sample Conditions Influence to sorghum at 25 C, 72 h Tannin increased Moneim et, al. (1995) Lupinus albus L. at 20 C, 9 days Vitamin E, vitamin C increased and high antioxidant activity Frias et, al. (25) soybean at 20 C, at 2, 4 and 6 days Total phenolic comnpound increasing and high antioxidant activity Lopez-Amoros et, al. (26) Corn, soybean at 30 C, 8 days Phytohormone increased Cassan et, al. (29) Sesame seed at 35 C, 4 days Fat contents decreased but the minerals such as Ca, P, Mg increased Hahm et, al. (29) Rice at 30 C, 5 days Free amino acid and soluble protein increased Veluppillai et, al. (29) barley at 37 C, 24 h Protein, fat decreased but reducing sugar increased Arora et, al. (2010) Soybean at 25 C, at 24, 48, 72, 96 and 120 h Protein, reducing sugar, riboflavin and ascorbic acid increased Kaushik et, al. (2010) 11
Germ GABA γ - oryzanol Antioxidants Embryo Isoflavone Antioxidants Antioxidants act as "free radical scavengers" and hence prevent oxidation by free radicals. Free radicals are atoms or groups of atoms with an odd (unpaired) number of electrons. Examples: O 2- Superoxide anion OH - Hydroxyl radical ROO Peroxy radical H 2 O 2 Hydrogen Peroxide Free radicals electrons http://www www.natural natural-plus plus-healthy healthy.com com/antioxodant antioxodant-facts facts..html.html 12
Mechanisms of Antioxidants The free radical will take the extra electron from antioxidant molecule and become neutralized. The free radical is stabilized without creating a new free radical or damage to a healthy cell. Antioxidants The free radical is then flushed out through the kidneys, so it can do no more harm to cell membrane. http://www www.natural natural-plus plus-healthy healthy.com com/index index.html Objective Study on the bioactive compounds change. Study on the antioxidant activities of germinated cereals. 13
Methods Part 1 65 C, 6 h. AOAC, 20 14
cereals Soaked at 30 C, 48 h Soaked at 30 C, 6 h at 48 h at 24 h rice Soybean Sesame Proximate analysis Protein Lipid Fiber Ash Moisture Carbohydrate AOAC, 20 Phytochemical analysis GABA (Liu et, al., 1995) Vitamin B1 B (Moongngarm et, al., 2010) Phenolic compounds (Iqbal, 25 ) Phytic acid (Febles et, al., 22 ) DPPH free radical scavenging (Iqbal, 25 ) Scavenging activity of ABTS (Yang et, al., 29) 15
GABA 3 g +70+ 70% Ethanol 30 ml. Centrifuged 6,0 rpm 30 min. Placed to ice bath 2 min. Pipette 0.2 ml. + Borate buffer ph.9.0 0.2 0 ml. + 6% Phenol reagent 1 ml. Boiling 10 min. Evaporated. Added 7.5% NaOCl 0.4 ml.and mixing Placed into + distilled water ice bath 5 min. 3 ml Placed into ice bath 30 min. Absorbance at 630 nm. Centrifuged 6,0 rpm 2 min. Liu et. al, 1995 Vitamin B1 0.1 g + distilled water 10 ml Mixing and incubated at room temperature 30 min. Pipette 1 ml. + NH 4 Cl-NH 3 H 2 O buffer 1.5 ml. + 1% tween 20 1 ml Adjusted into volumetric flask 10 ml 0.05 05% phenol red 1-2 droplet Filter Mixing and incubated at room temperature 10 min. Pipette 0.3 ml. adjusted 10 ml. Absorbance at 427 nm. Moongngarm et. al, 2010 16
Phenolic compound Mixing 3 g +80+ 80% Methanol 30 ml Added 7.5% Na 2 CO 3 4 ml. Incubated at 25 C, 24 h Centrifuged 6,0 rpm 20 min. Incubated at room temperature 2 h. Absorbance at 760 nm. Pipette 50 µl. + Distilled water 950 µl. + 10% Folin-Ciocalteu phenol reagent 5 ml. Iqbal, 25 Phytic acid Boiling 30 min. Placed to ice bath 15 min. 0.25 g +0.2N + HCl 25 ml Mixing 1 h and Centrifuged 6,0 rpm 30 min. Pipette 0.5 ml. + 0.2 mol/l FeCl 3 1 ml. Centrifuged 6,0 rpm 30 min. Pipette 1 ml. + 2,2'-bipyridine 1.5 ml. Mixing Absorbance at 519 nm. Fabels, et. al, 22 17
Antioxidants activity DPPH free radical scavenging 3g +80+ 80% Ethanol 30 ml Placed into dark room 30 min Shaking at room temperature, 24 h Absorbance at 517 nm. Centrifuged 6,0 rpm 20 min Pipette the supernatant 40 µl. + 0.1 mm DPPH solution 4 ml. Iqbal, et. al, 25 Antioxidants activity Scavenging activity of ABTS 3g +80+ 80% Ethanol 30 ml Shaking at room temperature, 24 h Placed into dark room 6 min Absorbance at 734 nm. Centrifuged 6,0 rpm 20 min Pipette the supernatant 40 µl. + 0.1 mm ABTS solution 4 ml. Yang, et. al, 29 18
RESULTS 45. 40. Figure 1. Protein contents change of Cereals Nutrition Values (%) 35. 30. 25. 20. 15. 10. 5. Control 0. Rice Soy bean Black sesame 19
45. 40. Figure 2. Lipid contents change of Cereals Nutrition Values (%) 35. 30. 25. 20. 15. 10. 5. Control 0. Rice Soy bean Black sesame 3. Figure 3. Fiber contents change of Cereals Nutrition Values (%) 2.50 2. 1.50 1. 0.50 Control 0. Rice Soybean Black sesame 20
Figure 4. Carbohydrate contents change of Cereals 90. 80. Nutrition Values (%) 70. 60. 50. 40. 30. 20. 10. Control 0. Rice Soybean Black sesame 25. Figure 5. GABA contents change of Cereals Phytonutrient Values (mg/ mg/1 1g dry) 20. 15. 10. 5. Glutamate decarboxylase Control Geminated 0. Rice Soy bean Black sesame 21
Figure 6. Vitamin B1 contents change of Cereals 1.40 1.20 Phytonutrient Values (mg/1 1g dry) 1. 0.80 0.60 0.40 0.20 Control Geminated 0. Rice Soy bean Black sesame Figure 7. Phenolic compound contents change of Cereals 60. Total Phenolic contents (g GAE/ g dry) 50. 40. 30. 20. 10. Nongermination Germination 0. Rice Soybean Black Sesame 22
Figure 8. Phytic acid contents change of Cereals 10. 9. Phytonutrient Values (mg/ mg/1 1g dry) 8. 7. 6. 5. 4. 3. 2. 1. Control Geminated 0. Rice Soy bean Black sesame Tables 1. Antioxidants activity of Cereals Cereals (Antioxidants activity, %) Rice Rice Soybean Soybean Sesame Sesame DPPH scavenging activity ABTS scavenging activity 42.7 40.2 57.1 48.6 34.3 25.9 46.8 36.5 46.3 50.2 56.5 55.9 37.1 47.6 46.9 52.2 23
Conclusions Cereals have enhancing the nutrition values and antioxidants activity by germination. The four germinated cereals have protein contents increased but lipid, fiber and carbohydrate contents decreased. Vitamin B1, GABA and phenolic compounds contents of four germinated cereals were increased but anti-nutrition contents, phytic acid decreased. Moreover, the antioxidants activity of germinated cereals were increased due to the phenolic compounds increased. ขอขอบค ณ ขอขอบค ณส าน กงานกองท นสน บสน นการว จ ย ท นว จ ยมหาบ ณฑ ต สกว. สาขาว ทยาศาสตร และเทคโนโลย ภายใต โครงการเช อมโยงภาคการผล ตก บ งานว จ ย ท น สกว. อ ตสาหกรรม (MAG Window I) ขอขอบค ณ รองศาสตราจารย ดร. ณ ฏฐา เลาหก ลจ ตต และรอง ศาสตราจารย ดร.อรพ น เก ดช ช น คณะทร พยากรช วภาพและเทคโนโลย มหาว ทยาล ยเทคโนโลย พระจอมเกล าธนบ ร 24
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