高抑菌活性物质产生菌的选育、优化及其特性表征

Screening of Bacteriocin Producing Bacillus and Study on Fermentation Conditions and Its Antibacteriocin’s Properties

以发酵液对金黄色葡萄球菌的抑菌能力作为指标,从实验室保藏菌株中筛选出一株产抑菌活性物质水平较高的菌株JN-814C,经鉴定其为地衣芽孢杆菌。进一步研究该菌株产生与分泌细菌素的特性,确定其最适培养基为糊精2.5 g/L,酵母粉7.5 g/L,胰蛋白胨10 g/L,氯化钠3 g/L,最佳培养条件为培养时间22 h,培养温度37℃,初始pH 7.0,装液量50 mL,接种量2%。通过硫酸铵分级沉淀与DEAE-Sepharose-FF阴离子交换层析可初步纯化得到抑菌活性物质样品。纯化后的抑菌物质对革兰氏阳性菌有较好的拮抗性,对样品进行蛋白酶处理与氨基酸分析,发现其对碱性蛋白酶敏感,组成中谷氨酸含量较高,对温度、酸碱的敏感性较低,因此可基本被确定为一种抑菌肽。

In this study,bacterial strain preserved in laboratory was chosen with the inhibitory activity against Staphylococcus aureus,it was identified as Bacillus licheniformis JN-814C.The optimal conditions for Bacillus licheniformis JN-814C to produce bacteriocin were as follow:2.5 g/L dextrin,7.5 g/L yeast extract,10 g/L peptone,3.0 g/L NaCl,at pH 7.0 and at 37℃for 22 h,with an inoculum density of 2%and inoculum age of 22 h,with fluid volume 50 mL/250 mL.The bacteriocin was purified through salting-out 40-80%and DEAE-Sepharose-FF anion exchange chromatography respectively.This purified bacteriocin had the inhibitory activity on gram-positive bacterium.It is sensitive to alkaline protease and is not sensitive to temperature or acid or alkali and it have high glutamate content.Therefore,it can be basically determined as an antibacterial peptide.