罗非鱼碎肉酶法制备蛋白胨的工艺研究

为探讨罗非鱼碎鱼肉酶法制备蛋白胨的加工工艺,采用正交试验的方法,分别研究了木瓜蛋白酶、AS.1398中性蛋白酶、复合蛋白酶水解罗非鱼碎肉制备蛋白胨的工艺条件。结果表明:木瓜蛋白酶的最佳水解工艺条件为pH6.5,温度65℃,水解时间4h,加酶量1250 U/g,蛋白胨得率达12.63%;AS.1398中性蛋白酶最佳水解工艺条件为pH 7.5,温度55℃,水解时间4 h,加酶量750 U/g,蛋白胨得率达13.25%;复合蛋白酶的最佳水解工艺条件为pH 7.0,温度50℃,水解时间4.5 h,加酶量850 U/g,蛋白胨得率达11.43%。     

蛋白酶水解牛蛙排的工艺探讨

本文叙述了以生产冷冻牛蛙腿的下脚料牛蛙排作为原料，选用不同种类的蛋白酶进行水解比较试验，以挥发性盐基氮作为指标，认为选择木瓜蛋白酶较合适。同时对木瓜蛋白酶的用量，原料加水量，水妥时间及水解度进行了探讨，确定了木瓜蛋白酶水解牛蛙排较适宜的水解条件：加水量为原料的２倍，水解时间２小时，蛋白酶帮量为０．０９，水解度为５３．９％。     

酶法制备脱脂鱼蛋白的初步研究

采用5种酶对脱脂鲢肉进行水解制备水解鲢蛋白。脱脂鲢肉是分别采用异丙醇、正己烷、乙酸乙酯3种溶剂萃取法进行脱脂后得到。分别用木瓜蛋白酶、枯草杆菌中性蛋白酶、枯草杆菌碱性蛋白酶、胰蛋白酶和复合蛋白酶对3种脱脂鲢肉及未脱脂鲢肉进行水解。试验结果表明，乙酸乙酯对鲢肉脱脂率最高，脱脂率为96．33％。复合蛋白酶为最适酶，乙酸乙酯脱脂鱼肉为最佳底物；在最佳水僻条件下：pH6．5，温度55℃，酶量3500U／g蛋白质，底物浓度（乙酸乙酯脱脂鲢肉：水）1：5，其水解度达到最大，为44．00％。     

罗非鱼活性肽分离及抗氧化能力研究

利用正交试验L9(34),以清除超氧自由基能力为指标,分别对木瓜蛋白酶、中性蛋白酶水解罗非鱼肉的水解条件进行优化,并对最佳清除率下的两种酶解物进行Sephadex G-50凝胶柱分离,检测了各组分对超氧自由基清除率及活性肽分子量分布情况.结果表明:中性蛋白酶在45 ℃、酶的质量分数2.0%、水解105 min及肉水比1∶3的水解条件下对超氧自由基有较好清除作用;木瓜蛋白酶在60 ℃、酶的质量分数2.0%、时间150 min及肉水比1∶2的水解条件下对超氧自由基有较好清除效果.木瓜蛋白酶酶解物在分子量为660 Da的多肽洗脱峰具有最大超氧自由基清除率,中性蛋白酶酶产物在分子量为1320 Da多肽洗脱峰具有最大超氧自由基清除率.     

蛋白酶对文蛤肉水解效果的研究

本文探讨了木瓜蛋白酶、胰蛋白酶、精制中性蛋白酶、酸性蛋白酶、复合蛋白酶和风味蛋白酶等6种蛋白酶对文蛤肉的水解效果,以文蛤肉水解液的水解度、氮收率及水解得率为指标,并对酶解液进行感官鉴评和风味评分,结果表明精制中性蛋白酶、胰蛋白酶和风味蛋白酶为适用于文蛤肉水解的蛋白酶。     

酶法提取鱿鱼皮胶原蛋白工艺条件的研究

研究了酶法提取鱿鱼皮胶原蛋白的工艺条件。根据5种蛋白酶水解液中羟脯氨酸（HYP）的含量,确定了胰蛋白酶和木瓜蛋白酶这2种提取率高的酶为水解用酶,采用L16（4^5）正交试验确定了这2种酶水解鱿鱼皮以制备胶原蛋白的最佳酶解条件。结果表明,胰蛋白酶和木瓜蛋白酶水解鱿鱼皮以制备胶原蛋白的最佳温度、加酶量、底物浓度、pH、时间分别为55℃、1200U·g^-1、1：20、pH8.0、4h和50℃、3200U·g^-1、1：20、pH6.0、6h。2种蛋白酶提取的胶原蛋白含量分别为11.08%和11.36%,提取率分别为95.16%和97.56%。     

双酶法在鲨鱼肉水解液制备中的工艺改良研究

采用同时添加枯草杆菌中性蛋白酶与木瓜蛋白酶的改良酶解技术，对鲨鱼肉进行水解。正交试验结果表明，上述二酶同时水解的最适条件为：温度45℃，pH7．0，时间3h，酶浓度200U／ml（枯草杆菌中性蛋白酶及木瓜蛋白酶各100U／ml），原料：水＝1．5。蛋白质水解率达83．5％，所得水认的氨基酸含量为45．4mg/ml，其中必需氨基酸总量为19．8mg/ml。     

鳕碎肉酶解物清除羟自由基作用研究

通过测定鳕碎肉酶解物对Fenton体系产生的羟自由基的清除效果,从海洋蛋白酶YS-894、海洋蛋白酶YS-80、胃蛋白酶和木瓜蛋白酶中,筛选出海洋蛋白酶YS-894作为酶解鳕碎肉制备具有较高清除羟自由基活性酶解物的水解酶;用正交试验L9(34)对该酶的水解条件进行优化,并确定了水解的最佳肉水比。最终工艺条件为40℃、酶解45 m in、pH 9.0、酶解质量分数0.25%、底物∶水=1∶3的条件下进行水解,水解物对羟自由基的清除效果较好,清除率为68.88%。     

酶解罗非鱼鱼皮胶制备降血压肽的研究

为探讨罗非鱼鱼皮胶制备降血压肽的酶解工艺,研究选用Neutrase中性蛋白酶、Alcalase碱性内切蛋白酶、胰蛋白酶、木瓜蛋白酶和复合蛋白酶分别对罗非鱼鱼皮胶进行酶解,对其酶解液的水解度进行比较。结果表明复合蛋白酶的酶解效果最好。对复合蛋白酶的酶解工艺进行单因素优化,确定其最佳酶解工艺为:温度50℃、pH值7.0、料液比1∶5、酶量为底物量的1.5%。在此条件下水解6h时,水解度为60%,酶解产物对血管紧张素转化酶(ACE)的抑制率最高,达到78.96%。     

响应面优化酶法提取牡蛎糖原工艺条件研究

为研究制备牡蛎(Crassostrea angulata)糖原的最佳工艺条件,通过比较分析了碱性蛋白酶、木瓜蛋白酶、胃蛋白酶、中性蛋白酶和胰蛋白酶等5种不同蛋白酶对牡蛎进行酶解提取糖原的效果,以碱性蛋白酶为最优酶制剂。在对碱性蛋白酶酶解工艺条件进行单因素实验的基础上,应用Box-Bhenken中心组合设计3因素3水平试验方案,通过响应面分析,建立数学模型优化牡蛎糖原的提取条件,以p H、酶解温度和加酶量为自变量,牡蛎糖原提取率为指标,获得碱性蛋白酶水解牡蛎提取糖原的最佳工艺条件,即加酶量4%、p H9、水解温度50℃,此时牡蛎糖原的提取率可达(48.55±0.01)%。试验结果表明,响应面法对牡蛎糖原提取条件的优化方案合理可行,该研究为牡蛎糖原的开发利用提供了理论依据。     

Epidermal proteases of the Japanese eel

A fluorescent-sensitive assay was used to demonstrate the protease activity in the dorsal skin of Japanese eel (Anguilla japonica). Two distinct extracts were separately prepared from skin mucus and epidermal cell layers, with no mutual contamination. The epidermal extract was sensitive to various substrates, whereas there was no, or only marginal, susceptibility to the same substrates for the mucous extract. Optimum hydrolysis pHs of the epidermal extract was variable and below pH 7.0, and the optimum hydrolysis temperatures were between 40 and 50 °C. In addition, Tos-Phe-Ch₂Cl, chymostatin, CdCl₂, CuCl₂, HgCl₂ and ZnCl₂ inhibited protease activities to different extents. Several other reagents specifically affected the protease activities, and their induced effects were useful for the identification of epidermal proteases. The findings indicate that a proteolytic factor, exhibiting various enzymological specificities, is retained within epidermal cell layers of Japanese eel. This factor is composed of 4 distinct proteases, such as cathepsins L and B-like proteases, a serine protease and an aminopeptidase.     

Lysis of pathogenic bacteria by epidermal cathepsins L and B in the Japanese eel

Cysteine proteases, cathepsins L and B, in Japanese eel epidermis are suspected bacteriolysins against aquatic pathogens. This work examines the lysis of three species of gram-negative pathogenic bacteria by the eel epidermal extract according to the assay method for cysteine proteases. The optimum pH for the lysis of the three species of bacteria were commonly determined to be 5.0, where both of eel epidermal cathepsins L and B caused active proteolysis. Four kinds of specific inhibitors for cysteine proteases strongly inhibited these catheptic proteolyses and bacteriolyses. The activities for the lysis of three types of bacteria were at a similar level, but the effects on the bacterium which infects predominantly eel skin largely varied with individual fish. These results suggest that epidermal cathepsins L and B are responsible for lysis of any pathogenic bacteria in the nonspecific defense of Japanese eel.     

病毒蛋白酶的研究概况

大量的研究结果表明，病毒编码的蛋白酶参与病毒多肽前体的切割、组装和成熟，巳成为开发抗病毒药物的目标酶。本文依据近年病毒蛋白酶的研究成果，就蛋白酶种类、病毒蛋白酶、病毒蛋白酶在病毒复制中的作用及病毒蛋白酶的研究方法进行综述。     

产胞外蛋白酶菌株的筛选及环境因素对其产酶活性的影响

采集渔-稻复合系统中的水稻根际淤泥,筛选出有较强产胞外蛋白酶能力的菌株并研究环境因素对其产酶活性的影响。结果显示,通过初筛、蛋白酶复筛等筛选得到一株菌株(编号PRO9);此菌株所产产胞外蛋白酶能够促进水体中有机物质发生氨化反应,减少有机氮含量。此菌还具有产胞外碱性磷酸酶的能力。菌株鉴定结果显示此菌为芽孢杆菌。PRO9菌株所产的胞外蛋白酶在67℃下酶活性最为活跃,最适pH为7,属于中性蛋白酶。     

Characterization of protease activity in developing discus Symphysodon aequifasciata larva

The activity of different protease classes was monitored in developing discus (Symphysodon spp.) larvae using a combination of biochemical assays and substrate SDS–PAGE techniques. Results showed the presence of alkaline proteases of serine proteases such as trypsin with a significant increase in activity levels detected beginning 3 days after hatching. Other alkaline proteases such as metallo‐proteases and chymotrypsin, a type of serine protease, were only detected in older larval stages, at around 20–30 days after hatching. Acidic protease activity was very low during the first 20–25 days of development before showing a significant (P < 0.01) rise. This is despite the formation of a stomach observed 10 days after hatching. Based on the development of the protein digestive system observed, the use of microdiets to replace Artemia should be considered 25 days after hatching.     

Characterization of Gelatinolytic Activity in Seminal Plasma of Some Teleost Fish

Gelatin-containing SDS-PAGE combined with the incubation of gels in buffers containing protease inhibitors was performed for the visualization and characterization of proteolytic activity in teleost fish seminal plasma. To demonstrate the class of detected enzymes we used serine protease inhibitor – benzamidine or EDTA which inhibits metalloproteases activity. Additionally the effects of calcium ions on protease activity were investigated. Multiple gelatinolytic activities in seminal plasma of 10 teleost fish species from three orders (Cypriniformes, Salmoniformes, Perciformes) were found. Most proteases were either stimulated by Ca²⁺ and inhibited by EDTA or inhibited by benzamidine. This suggests that metalloproteases and serine proteases are major gelatinolytic proteases of fish seminal plasma. In cyprinid species we found a common profile of two gelatinolytic activities; the first band (60–66 kDa) belonged to metalloproteases, and the second one (76–81 kDa) belonged to serine proteases. Other bands were also visible and they represented mostly serine protease activity. Species from the Salmoniformes order showed a similarity in metalloproteases with molecular weights of about 64 and 75 kDa. Salmonid species also had similar serine proteases with molecular weights of about 102 and 165 kDa. In European grayling seminal plasma we found metalloproteases with molecular weight of 51, 57, 64, 70 kDa and two serine proteases activities of 35 and 125 kDa. Percid species had metalloproteases activities of 53 and 63 kDa and serine protease activity of 100 kDa. Protease of other, presently unknown classes were also found in seminal plasma of asp, chub, European grayling and pikeperch. The physiological role of seminal plasma proteases is still unknown.     

日本鳗鲡人工催产后亲鱼恢复培养与再催产效果

日本鳗鲡(Anguillajaponica)产后亲鱼的恢复培养全过程包括:产后亲鱼的海水淡化、诱导开口以及亲鱼培育。对2种不同淡化方式的结果比较发现,日本鳗鲡亲鱼的海水淡化方式以缓慢连续的方式较好,即每天换水1次,每次淡化量为总盐度的3%～4%,总淡化时间为1个月。淡化后的亲鱼存活率分别为:自然产卵的产后亲鱼达到100%,人工授精的产后亲鱼为86.3%,而难产的产后亲鱼则为81.5%。在各种开口驯养方式中,以水蚯蚓为开口饵料的驯养效果较好。诱导亲鱼开始摄食所需时间最短,为18d;摄食量也最大,达到平均体重的2.5%。从水蚯蚓逐步转为全人工饵料后,进行日常亲鱼培育。经18个月的恢复培养后,使产后亲鱼的平均体重雌鳗330g、雄鳗150g分别恢复到765g和470g。但2次繁殖的各项指标中,催熟率、成熟系数和性腺发育情况远远低于野生鳗,而催产率和幼苗存活率则略高于野生鳗,其原因尚有待进一步研究。     

三种微生物制剂对有机物废水中氨氮及亚硝酸盐的影响

实验采用复合净水菌、复合芽孢杆菌、光合细菌与空白对照比较了三种微生物制剂对有机物废水中氨氮及亚硝酸盐的处理效果。结果显示：清除氨氮方面：利用微生态制剂微生物制剂净化7d,三种微生物制剂对有机物污染水体中氨氮的清除率均极显著大于对照组（P〈0.01）,处理组之间复合芽孢杆菌的氨氮降解率56.58%,显著大于降解率分别为42.64%和44.02%的复合净水菌和光合细菌组（0.01〈P〈0.05）。清除亚硝酸盐方面：三种微生物制剂对亚硝酸的降解率均极显著大于对照组（P〈0.01）,而复合芽孢杆菌对亚硝酸的降解率68.84%,显著大于复合净水菌47.70%及光合细菌组37.17%（0.01〈P〈0.05）。结果表明：三种微生物制剂对有机物污染的水体中的氨氮及亚硝酸盐均具有降解效果,其中复合芽孢杆菌对于降低水体中氨氮和亚硝酸盐效果最佳。     

地锦草对水产病原菌体外抗活性的研究

摘要：用95％乙醇提取地锦草中的化合物，又分别用石油醚、乙酸乙酯、正丁醇和水对乙醇总提物进行分极性段萃取。探究地锦草各极性段二甲基亚砜溶液对嗜水气单胞菌、温和气单胞菌、柱状屈挠杆菌、鳗弧菌等4种常见水产动物病原菌的抑菌效果。     

扇贝裙边氨基酸营养粉的制备工艺研究

为了对扇贝裙边进行综合利用，本文选择由中性蛋白酶、胰蛋白酶和复合蛋白酶3种酶组合成3对混合酶对扇贝裙边进行酶水解实验，以水解液中游离氨基酸态氮为考察指标，采用正交实验设计研究了加酶量、酶解时间、酶解温度及酶的种类对酶解效果的影响，从而优化了扇贝裙边的酶法水解工艺条件。以最佳水解工艺条件获得的酶解液经喷雾干燥制成的氨基酸营养粉，经检测，其粗蛋白含量为82.89%，粗脂肪含量为1.55%，总糖含量为6.82%。该氨基酸营养粉中氨基酸种类齐全，每100g粗蛋白中氨基酸总量达79.06%，其中游离氨基酸总量达51.53%。