解淀粉芽孢杆菌抑菌物质粗提取的优化及分析

为探究解淀粉芽孢杆菌（Bacillus amyloliquefaciens）A13抑菌物质适宜提取方法，分别采用硫酸铵盐析法、酸沉淀法、甲醇抽提法、乙酸乙酯萃取法进行抑菌粗提液的制备，并通过SuperdexTM75凝胶层析柱分离纯化。结果表明，以80%饱和硫酸铵法制备粗提液抑菌效果最佳，抑菌圈直径为23.67 mm，蛋白质量浓度最高，为582.50 μg/mL，其回收率为17.8%；以甲醇抽提法制备粗提液的抑菌圈直径为25.17 mm，其回收率最高，为32.1%，蛋白质量浓度为505.00 μg/mL；以酸沉淀法制备粗提液的抑菌圈直径为22.67 mm，其回收率与蛋白质量浓度分别为13.7%与273.75 μg/mL；以乙酸乙酯法制备粗提液时，乙酸乙酯与发酵液为3∶1（V/V）的抑菌效果最佳，抑菌圈直径为25.5 mm，并且其回收率和蛋白质量浓度分别为23.6%和255.20 μg/mL。因此，硫酸铵盐析法为粗提解淀粉芽孢杆菌A13抑菌物质的适宜方法。

Optimization and analysis of crude extraction of bacteriostatic substances from Bacillus amyloliquefaciens

In order to explore the suitable extraction method of antibacterial substances from Bacillus amyloliquefaciens A13, antibacterial crude extract was prepared by ammonium sulfate precipitation method, acid precipitation method, methanol extraction method and ethyl acetate extraction method, and the crude extract was separated and purified by SuperdexTM75 gel chromatography column. The results showed that the crude extract prepared by 80% saturation ammonium sulfate had the optimal bacteriostatic effect, the diameter of bacteriostatic circle was the largest of 23.67 mm, the protein concentration was the highest of 582.50 μg/ml, and the recovery rate was 17.8%. The diameter of inhibition zone of crude extract prepared by methanol extraction was 25.17 mm with the highest recovery rate of 32.1% and protein concentration of 505.00 μg/ml. The inhibition zone diameter of crude extract prepared by acid precipitation method was 22.67 mm, and the recovery rate and protein concentration were 13.7% and 273.75 μg/ml, respectively. When the crude extract was prepared by ethyl acetate method, the antibacterial effect was the optimal when the ratio of ethyl acetate to fermentation broth was 3∶1 (V/V). The diameter of inhibition zone was 25.5 mm, and the recovery rate and protein concentration were 23.6% and 255.20 μg/ml, respectively. Therefore, ammonium sulfate salting-out method was the appropriate method for crude extraction of antimicrobial substances from B. amyloliquefaciens A13.