枯草芽孢杆菌对环境耐受性和产中性蛋白酶能力研究  

Study on environmental tolerance and neutral protease production capability of Bacillus subtilis

作　　者：李振 崔艳红 刘长忠 袁梦 LI Zhen;CUI Yanhong;LIU Changzhong;YUAN Meng(College of Animal Science and Technology/Xinxiang Key Laboratory of Animal Feed Nutrition,Henan University of Science and Technology,Xinxiang 453000,China)

机构地区：[1]河南科技学院动物科技学院新乡市动物饲料营养重点实验室,河南新乡453000

出　　处：《黑龙江畜牧兽医》2024年第4期81-87,92,共8页Heilongjiang Animal Science And veterinary Medicine

基　　金：河南省科技攻关项目(202102110092,212102110170)。

摘　　要：为了探究枯草芽孢杆菌对环境耐受性及产中性蛋白酶的能力,试验模拟饲料加工温度、胃肠道酸性环境、胆盐对枯草芽孢杆菌进行环境耐受性评定,在单因素(pH值、时间、温度、接种量、碳源、碳源添加量和金属离子)试验基础上采用正交试验以中性蛋白酶活力为评定指标,对枯草芽孢杆菌培养条件进行优化。结果表明:枯草芽孢杆菌对温度、胃肠道酸性环境和胆盐均具有一定耐受性;单因素试验中最佳pH值为5,培养时间为24 h,培养温度为30℃,接种量为2%,碳源为淀粉,淀粉添加量为3%;金属离子Mg^(2+)、Fe^(2+)、Mn^(2+)、Cu^(2+)和Zn^(2+)对产酶均具有抑制作用,不在培养基中添加金属离子。正交试验优化后枯草芽孢杆菌产中性蛋白酶活力最佳培养条件为pH值6,淀粉添加量3%,培养温度35℃,培养时间24 h,枯草芽孢杆菌种子菌液接种量3%,在最优条件下,酶活力可达40.05 U/mL。说明枯草芽孢杆菌可耐受动物胃肠道环境且培养条件优化后中性蛋白酶酶活力显著提高。In order to explore the environmental tolerance and the ability of neutral protease production of Bacillus subtilis,feed processing temperature,gastrointestinal acid environment,and bile salt were simulated,and the environmental tolerance of Bacillus subtilis to bile salts was assessed.On the basis of single factor tests(pH value,time,temperature,inoculation amount,carbon source,carbon source addition amount and metal ion),the neutral protease activity was used as the evaluation index to optimize the culture conditions of Bacillus subtilis by orthogonal test.The results showed that Bacillus subtilis had a certain tolerance to temperature,gastrointestinal pH value and bile salt.In the single factor test,the optimal pH value was 5;the culture time was 24 h;the culture temperature was 30℃;the inoculation amount was 2%;the carbon source was starch;the addition amount of starch was 3%,and metal ions Mg^(2+),Fe^(2+),Mn^(2+),Cu^(2+)and Zn^(2+)all had inhibitory effects on the enzyme production,and didn't add metal ions to the medium.After the optimization of orthogonal experiment,the optimal culture conditions for the activity of neutral protease production from Bacillus subtilis were pH value 6,starch addition amount 3%,temperature 35℃,culture time 24 h and 3%inoculum size of Bacillus subtilis seed.Under the optimal conditions,the enzyme activity reached 40.05 U/mL.The results indicated that Bacillus subtilis could tolerate the environment of animal gastrointestinal tract and the enzyme activity of dispase was significantly improved after the optimization of culture conditions.

关 键 词：枯草芽孢杆菌 胃肠道环境 耐受性 中性蛋白酶 产酶条件 

分 类 号：S816.79[农业科学—饲料科学]