【作者】 殷金莲；

【导师】 徐怀德；

【作者基本信息】 西北农林科技大学 ， 食品科学， 2007， 硕士

【摘要】 甲鱼具有很高的营养价值和药用价值,是开发高级保健食品的良好原料。近年来由于甲鱼产量的迅速增加,为甲鱼的深加工提供了良好的机遇。为了科学认识甲鱼的营养保健功能,本试验以甲鱼为原料,利用三种蛋白酶酶解甲鱼蛋白,研究了酶解产物的水解特性、抗氧化性及体外ACE抑制功能,并对木瓜蛋白酶酶解产物进行了分离;尝试性地用枯草芽孢杆菌发酵生产甲鱼多肽,并开发制作了甲鱼多肽酒。主要研究结果如下:随着酶解时间的延长,三种酶解产物的氨基态氮含量、水解度、多肽含量、溶解度都呈增大的趋势,肽分子平均链长不断减小,三种酶解产物的自由基清除能力和还原能力整体也呈增大的趋势,比较得到木瓜蛋白酶为酶解甲鱼蛋白的最佳蛋白酶。以体外ACE抑制率、·OH、O^2-·和DPPH·清除率为指标,通过正交试验确定了木瓜蛋白酶酶解甲鱼蛋白制备ACE抑制肽的最佳酶解条件为:温度60℃、pH7.0、酶解时间3h、加酶量1.0%;在此酶解条件下酶解产物的ACE抑制率为:99.96%,利用SephadexG-25进行分离,获得了ACE抑制活性较高的组分F4和F1,其ACE抑制率分别为:67.27%和52.73%,制备抗氧化肽的最佳酶解条件为:温度60℃、pH5.0、时间4h、加酶量0.6%,得到了抗氧化活性最高的组分F2 ,该组分的·OH、O^2-·和DPPH·清除率分别为:10.39%、33.00%和17.68%,其分子量范围集中在307左右。以甲鱼肉为原料,接种枯草芽孢杆菌,在37℃、120r/min摇瓶条件下,发酵72h,将甲鱼蛋白水解为甲鱼多肽,随着发酵时间的延长,甲鱼多肽溶液的氨基态氮含量、水解度、溶解度、多肽含量都在增大;肽分子平均链长在不断缩短;发酵液表现出一定的OH·、O^2-·清除率和还原能力。发酵72h,发酵液的水解度达到7.96%;氨基态氮含量增大到0.0827g/100mL;多肽平均链长由41.36缩短到12.09;溶解度增大到11.90%;多肽含量达到3.34 mg/mL;·OH清除率增大到33.53%;O^2-·清除率增大到49.92%;总还原能力达到0.416。添加酵母于木瓜蛋白酶酶解液中进行发酵,制成的甲鱼多肽酒为淡黄色,酒体丰厚,清亮透明,无悬浮物或沉淀产生,具有淡淡的醇香和甲鱼特有的香味。甲鱼多肽酒中多肽含量高达9.85 mg/mL,同时具有较强的抗氧化性,·OH、O^2-·、DPPH·清除率最高分别达到54.36%、87.27%、90.16%,总还原能力最大达到1.138。通过t-检验,蔗糖的添加量对·OH、O^2-·、DPPH·清除率和总还原能力没有显著影响（P>0.05）。更多还原

【Abstract】 There are quite high nutrition and medicine values in Soft-shelled Turtle and it is the favorable material of health food. In recent years, turtle’s production was quickly increasing, bringing favorable opportunities to turtle’s deep-processing. To know turtle’s nutrition and health care function sciencly, using Turtle as material ,Turtle protein was enzyme hydrolyzed by papain,trypsin and subtilisin,mainly researching on some hydrolytic properties,the antioxidant functional properties and ACE inhibitory ratio in vitro of three hydrolysates.. papain hydrolysates were separated,The feasibility of Bacillus subtilis fermenting and producing Turtle peptides was test,And Turtle peptide wine was prepared. The main results are as follows:For three hydrolysates, with time’s extension, the content of amino nitrogen, the degree of solubility, the degree of hydrolysis, and peptides content were all increased, the average length of peptides chains were decreased gradually, the scavenging ability of free radical and the ability of deoxidization were also increased. By comparing, papain protease was the optimum protease.We did orthogonal trial with the ACE inhibitory ratio in vitro、·OH、O^2-·and DPPH·scavenging ratio being index. The hydrolysate with the ACE inhibitory ratio being 99.96%, was gained under the condition of temperature 60℃, pH 7.0, enzymatic hydrolysis time 3 h, enzyme concentration 1.0%. Trough separating by SephadexG-25, the fractions F4 and F1 with the higher activity was collected, with the ACE inhibitory ratio being 67.27% and 52.73%. The optimal conditions of preparing antioxidant was temperature 60℃, pH5.0, enzymatic hydrolysis time 4 h, and enzyme concentration 0.6%. The fraction F2 with the highest activity was collected, with the·OH、O^2-·and DPPH·scavenging ratio being 10.39%、33.00% and 17.68%. Molecular weight distribution was located 307 around.Using Soft-shelled Turtle protein as raw materials and Bacillus subtilis as starter, Turtle protein was hydrolyzed into polypeptide with 120r/min in shake flask at 37℃for 72h. With time’s extension, amino nitrogen, degree of hydrolysis, peptides content, and degree of solubility all increased the average length of peptides chain decreased gradually. Turtle polypeptide solutions put up scavenging capacities of·OH and O^2-·, deoxidization ability. When fermenting 72h, the degree hydrolysis of fermented solution reached 7.96%; amino nitrogen content was reached to 0.0827g/100mL, the average length of peptides chains was shorted from 41.36 to12.09, the degree of solubility was increased to 11.90%, the peptides content reached 3.34 mg/mL, the·OH scavenging ratio was increased to 33.53%, the O^2-·scavenging ratio was reached 49.92%, the ability of deoxidization reached 0.416.Adding yeast into papain hydrolysis fluid, Turtle peptide wine was prepared. With the color being light yellow, wine body rich and generous, limpidity, having no suspended substance and deposit. It had light wine aroma and Soft-shelled Turtle‘s special fragrance. After fermentation, peptides content reached 9.85 mg/mL at highest. Turtle peptide wine had·OH O^2-·, DPPH·scavenging ability, respectly reaching 54.36%,87.27%, 90.16% at highest, The highest deoxidization ability reached 1.138.Trough t-detection, the recruitment of sugar had no marked affection on·OH ,O^2-·, DPPH·scavenging ratio and deoxidization ability（P>0.05）.更多还原