高效溶栓菌株的筛选鉴定及发酵条件优化  

Screening and Determination of a Bacterium Strain with High Fibrinolytic Activities and the Optimization of Its Fermentation Conditions

作　　者：王丽[1] 郝友进[1] 何正波[1] 和七一[1] 陈治霖[1] 陈斌[1] 

机构地区：[1]重庆师范大学活性物质生物技术教育部工程研究中心重庆高校生物活性物质工程研究中心昆虫与分子生物学研究所,重庆401331

出　　处：《重庆师范大学学报（自然科学版）》2013年第3期121-126,共6页Journal of Chongqing Normal University:Natural Science

基　　金：重庆市科技攻关重点项目(No.CSTC2012GG-YYJSB80002);重庆市自然科学基金重点项目(No.CSTC2008BA5030)

摘　　要：采用酪蛋白平板法,从纳豆激酶胶囊、纳豆及多种发酵食品中筛选出了146株产溶栓酶菌株;然后利用纤维蛋白平板法比较发酵后溶栓酶活力,并从初筛菌株中筛选出1株溶栓酶活达1 637.6IU/mL的高产菌株HT8;依据菌种形态和生理生化特征以及16SrRNA基因序列分析,鉴定为枯草芽孢杆菌(Bacillus subtilis)。在此基础上,对此高效溶栓菌株的液体培养条件进行了单因素初步实验,方差分析后设计正交试验L18(37),通过软件SPSS分析确定该菌最佳发酵条件为初始pH值8.0、温度40℃、培养体积90mL(容量为500mL的三角瓶装90mL的培养液)、0.5%牛肉膏和2%大豆蛋白胨;优化后的相对酶活力达1 804.08IU/mL。研究结果为进一步开发纳豆等保健食品提供理论基础。In recent years, natto and natto bacteria have become a research hotpoint in many countries because its metabolite nattoki- nase has high fibrinolytic activity. In the present study, 146 fibrinolytic enzyme producing bacterial strains were isolated from nat- tokinase capsules, natto, and a variety of fermented foods using casein-plate method. The fibrinolytic activities of the fermentation broth of these isolated bacteris were compared with the fibrin plate method, and one strain （named as HT8） with the highest fi- brinolytic activity （1 637.6 IU/mL） was identified. It was classified as Bacillus subtilis based on the clony morphological, physio- logical and biochemical characteristics and 16S rRNA gene sequences. Based on this result, the fermentation conditions were deter mined, then orthogonal experiments using L18 （37） array were performed and statistically analyzed by ANOVA in SPSS. The result showed that the optimal fermentation condition was medium＇s initial pH value 8.0, fermentation temperature 40 ℃, medium volume 90/500 mL, 0.5% beef extract and 2% soybean peptone. The relative enzyme activity was 1 804.08 IU/mL under the optimal fer- mentation condition, which provided a theoretical basis for the further development of natto and of healthy food.

关 键 词：溶栓菌 菌种筛选 枯草芽孢杆菌 发酵条件 纳豆激酶 

分 类 号：Q503[生物学—生物化学] Q556