响应面法对多黏菌素E发酵工艺的优化

Optimization of fermentation process for polymyxin E by response surface methodology

为了优化多黏菌素E的发酵工艺,以多黏菌素含量为考核指标,在单因素实验确定因素和水平的基础上,利用响应面法对多黏类芽孢杆菌产多黏菌素E的发酵培养基和工艺进行了初步优化。对7个因素进行Plackett-Burman(P-B)实验,得到3个显著因子。Plackett-Burman实验结果表明,蛋白胨、温度和硫酸铵是影响多黏菌素E发酵产量的显著因子。然后进行最陡爬坡实验逼近最佳响应面区域,最后通过Central Composite Design,利用Design-Expert V8.06软件进行回归分析,得到最佳条件,并进行响应面的预测值和实测值的验证。多黏菌素E的优化发酵条件结果为:葡萄糖20g/L、蛋白胨21.05g/L、硫酸铵16.25g/L、种龄21h、接种量10%(V/V)、温度33.3℃、装液量1L/1.8L。在该优化的发酵条件下,多黏类芽孢杆菌产多黏菌素E的最大效价达到28577.2U/m L,较优化前的多黏菌素E效价(25103.4U/m L)提高了13.8%。优化的发酵工艺合理、稳定、可行,为进一步工业化生产优化奠定了基础。

In order to optimize the fermentation process ofpolymyxin E, polymyxin content as evaluation index, the single factor experiment, on the basis of determining factors and levels, the response surface method was used to optimize polymyxin E fermentation process. Three significant factors were obtained from seven influencing factors of Plackett-Burman experiments. Based on Plackett-Burman, the peptone, temperature, （NH4）2SO4 were chosen as influencing factors. The path of steepest ascent was used to approach the optimal region of the fermentation conditions subsequently. In the third step, the concentrations of those three main factors were further optimized by using central composite design and response surface analysis by solving the quadratic regression model equation by Design-expert, the predicted value and measured value were also contrasted. The optimal fermentation conditions were determined as follows： glucose 20g/L, peptone 21.05g/L, （NH4）2SO4 16.25g/L, seed age 21h, inoculum concentration 10%（V/V）, temperature 33.3℃, broth content 1L/1.8L, respectively. In optimization of fermentation conditions, the titer of polymyxin E is 28577.2U/mL, which was increased by 13.8% in comparison with that of the before optimization of fermentation. The optimized fermentation condition is reasonable and scientific, and it could provide scientific theoretical basis for the future industrialized production.