大丽轮枝菌拮抗细菌枯草芽孢杆菌12-34菌株发酵产抗菌蛋白的条件优化

采用单因素试验及正交试验结合的方法，优化大丽轮枝菌拮抗细菌枯草芽孢杆菌（Bacillus subtilis）12-34菌株产抗菌蛋白的发酵条件。通过单因素试验和正交试验，确定了B. subtilis 12-34菌株产抗菌蛋白的适宜碳源、氮源和无机离子，优化了培养基的组成；并对B. subtilis 12-34菌株产抗菌蛋白的发酵时间、装瓶量、接种量、培养基初始pH、摇床转速和发酵温度进行了优化。结果显示，B. subtilis 12-34菌株产抗菌蛋白的最佳培养基组成为玉米粉5%，牛肉浸膏3%，CaCl 2 0.10%，KCl 0.05%；B. subtilis 12-34菌株产抗菌蛋白的最佳发酵条件为培养基初始pH9.0，种子液按10%接种量接种至50 mL/250 mL三角瓶中，200 r/min，37 ℃培养48 h。优化后表征抑菌蛋白产量的抑菌圈直径增大了115.5%。

Optimization of the fermentation conditions of the antagonistic strain 12-34 of Bacillus subtilis against Verticillium dahliae

The strain 12-34 of Bacillus subtilis is an antagonistic strain with high antagonistic activity against Verticillium dahliae Kleb. The protein from a fractional precipitation of the fermentation broth showed antagonistic activity. The filtered fermentation broth after 121 ℃ heat treatment showed no antifungal activity. The extract of the fermentation broth using chloroform showed no antifungal activity. So it is concluded that the protein had the antifungal activity. Further studies on composition of media and fermentation conditions of antifungal protein of the strain were carried out. Through single factor experiments, the suitable carbon source, nitrogen source and inorganic salt were chosen. The optimal composition of media was determined by orthogonal experiments. The optimal medium of strain B. subtilis 12-34 contained 5% maizena, 3% beef extraction, 0.10% CaCl 2 and 0.05% KCl. The optimization of four kinds of fermentation factors, such as initial pH, inoculum size, media volume and fermentation time, was made by orthogonal experiments. Other fermentation factors, such as rotation speed and fermentation temperature, were also optimized by single factor experiments. The optimal fermentation conditions were pH9.0 and 10% inoculum size, 37 ℃, 200 r/min and 48 h. The results showed that calcium salts and beef extraction played an important role in the fermentation process. And the volume was the most important factor in fermentation. A distinguished elevation of antifungal protein yield by about 115.5%was observed.