83A-6 |
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S. TUNGKAWACHARA1, J. W. Park1, and Y. J. Choi2. (1) Dept. of Food Science & Technology, Oregon State Univ., Astoria Seafood Lab., 2001 Marine Dr., Rm. 253, Astoria, OR 97103-3420, (2) Division of Marine Bioscience, Gyeongsang National University, Tong Yeong, South Korea Fish intestines comprise approximately 1% of fish weight and are a rich source of proteases. The fish sauce process can be accelerated by the addition of such enzyme-rich organs. Considering the optimum pH of tryptic enzymes are neutral to slightly alkaline, the introduction of enzyme-rich intestines to fish sauce processing could therefore accelerate proteolytic degradation. In our continuous efforts to develop fish sauce from enzyme-rich Pacific whiting, the characterization of tryptic enzymes isolated from Pacific whiting intestines is needed. Our objective was to investigate the characteristics of protease isolated from Pacific whiting intestines. Fish intestinal organs were randomly collected from over 300 fresh fish obtained from a surimi processing line. They were kept on ice and transported to the Oregon State University Seafood Laboratory. They were frozen at -18 ºC until utilized. Tryptic enzyme from the intestines of Pacific whiting was isolated by treating with ammonium sulfate (30-70%) and column chromatography (Benzamidine FF, 1st Superdex 200, Q-Sepharose, 2nd Superdex 200). The proteinase was concentrated 436.0 folds compared to the crude extract in the ammonium sulfate fraction. The specific activity at 25ºC was 2.18 U/mg. Enzyme was characterized according to the method of Simpson and Haard (1985). The enzyme consisted of single polypeptide chains with a molecular weight of about 24 KDa on the SDS PAGE. Its optimum pH was 8.0 and showed maximum activity at 30-40ºC. The electrophoresis pattern of protease on SDS PAGE was not affected by treatment with mercaptoethanol, suggesting that it was composed of a single polypeptide chain. The K’m at 25 ºC was 0.37 mM. This enzyme was also inhibited by phenyl methane sulphonyl fluoride (PMSF), aprotinin, soybean trypsin inhibitor (SBTI), and tosyl-L-lysine chloromethylketone (TLCK). A partially purified enzyme from Pacific whiting intestine was trypsin-like. Characterization of this enzyme could maximize its applications.
Session 83A, Aquatic Food Products: Byproducts, mince and surimi
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