A heat-stable trypsin inhibitor in adzuki bean (Vigna angularis): effect of extraction media, purification and biochemical characteristics |
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Authors: | Sappasith Klomklao ,Soottawat Benjakul,Hideki Kishimura,Kazufumi Osako,& Munehiko Tanaka |
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Affiliation: | Department of Food Science and Technology, Faculty of Technology and Community Development, Thaksin University, Phattalung Campus, Phattalung, 93110, Thailand; Department of Food Technology, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, Songkhla, 90112, Thailand; Research Faculty of Fisheries Sciences, Hokkaido University, Hakodate, Hokkaido, 041-8611, Japan; Department of Food Science and Technology, Tokyo University of Marine Science and Technology, Konan 4, Minato, Tokyo 108-8477, Japan |
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Abstract: | Trypsin inhibitor from adzuki bean ( Vigna angularis ) seed was isolated and characterised. Extraction of seed with NaCl at the concentration of 0.15 m showed a higher recovery of trypsin inhibitor than other solvents tested ( P < 0.05). Optimal extraction time for the recovery trypsin inhibitor from adzuki bean seed was 30 min ( P < 0.05). Purification of inhibitor was achieved by heat-treatment at 90 °C for 10 min, followed by ammonium sulphate precipitation with 30–65% saturation and size exclusion chromatography on Sephacryl S-200, presenting a yield and purification of 53.9% and 10.91-fold, respectively. The apparent molecular weight of trypsin inhibitor was estimated to be 14 kDa based on SDS-PAGE and inhibitor activity of zones separated by electrophoresis. The purified inhibitor was stable over a broad pH range and retained high inhibitory activity toward trypsin after incubation at 90 °C for 60 min. NaCl, at 0–3% concentration, did not affect the inhibitory activity of purified trypsin inhibitor, however, the activity was lost when sample was treated with β-mercaptoethanol prior to electrophoresis. |
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Keywords: | Inhibitor isolation legume proteinase trypsin |
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