Development and application of novel enzyme inhibitors for the preservation of freshness of marine products using a library of marine microorganisms

众所周知，适应冷的酶是海洋产品降解的原因。为了保持海产品的新鲜度，本研究旨在对这些酶进行表征，并从天然来源中寻找它们的抑制剂。为了获得冷适应酶，我们从日本不同地区的海洋动物肠道内容物中校正了3000多种海洋细菌。我们筛选了具有脂肪酶活性和磷脂降解酶活性（磷脂酶A1、磷脂酶C、溶磷脂酶C和鞘磷脂酶活性）的有价值菌株。最后，我们在培养液中确认了约100株具有脂肪酶活性的菌株，约10株具有磷脂降解酶活性的菌株。我们计划对这些酶进行表征，并从之前分离的所有海洋微生物中寻找这些海洋脂质降解酶的特异性抑制剂。本研究还尝试从不同地区的海水和海洋沉积物样品中分离产生酪氨酸酶抑制剂的海洋微生物。其中一株产酪氨酸酶抑制剂的菌株从Off-Niijima Islands的100 m深度分离得到，并被选中进行进一步的鉴定和生产抑制剂的研究。根据分类学特征，鉴定菌株为木霉（Trichoderma sp.）。菌株在海水培养基中振荡培养时产生抑菌剂。活性物质为低分子量肽。原油抑制剂可以防止甲壳类动物（如虾）的变黑反应，这表明该抑制剂可能适用于渔业。

Cold-adapted enzymes are known to be cause for debasement of marine products. To preserve freshness of marine products, our present study was designed to characterize the enzymes and to search their inhibitors from natural sources.In order to obtain the cold-adapted enzymes, we have corrected over 3,000 strains of marine bacteria from the intestinal contents of marine animals in various regions in Japan. We have screened valuable strains indicating lipase activity and phospholipid-degrading enzyme activities (phospholipase A1, phaspholipase C, lyzophospholipase C and sphingomyelinase activities). Finally, we have confirmed around 100 strains with lipase activity, and around 10 strains with phospholipids-degrading enzyme activities in the culture solution. We are planning to characterize these enzymes and to search their specific inhibitors for these marine lipid-degrading enzymes from all marine microorganisms isolated before at once.In the present study, an attempt was also made to isolate tyrosinase inhibitor-producing marine microorganisms from seawater and marine sediment samples of various areas. One strain which produced a tyrosinase inhibitor was isolated from Off-Niijima Islands at a depth of 100 m and was selected for further studies on the identification of it and production of the inhibitor. Base on the taxonomical characteristics, the strain was identified as Trichoderma sp. The strain produced the inhibitor when cultivated in seawater medium with shaking. The active substance was found to be a low molecular weight peptide. The crude inhibitor prevented the darkening reaction of crustaceans such as shrimp they are stored, suggesting that this inhibitor might be applicable to fisheries.