响应面法优化产β-内酰胺酶菌Bacillus cereus B03的发酵条件

Optimization of Fermentation Conditions ofβ-lactamase Producing Bacteria Bacillus cereus B03 by Response Surface Methodology

为快速高效地提高菌株Bacillus cereus B03的产酶能力,采用响应面法对Bacillus cereus B03产β-内酰胺酶的发酵培养基进行优化。首先通过单因素实验研究了不同碳源及浓度、不同氮源及浓度、不同金属离子及浓度、氯化钠、磷酸氢二钾以及温度、pH、接种量、装液量对菌株产酶活力的影响,然后设计Plackett-Burman试验筛选出影响产酶的3个显著性因素:温度、pH、接种量。在此基础上,最后设计Box-Behnken中心组合试验确定最优产酶发酵条件。结果表明,最佳发酵培养基成分为葡萄糖20 g/L、酵母浸粉20 g/L、NaCl 2 g/L、MgSO4·7H2O 0.2 g/L,K2HPO4·3H2O 4 g/L,最佳产酶发酵条件为发酵温度37℃,pH为7.3,接种量3%,装液量50 mL/250 mL。在此优化条件下,该菌株产β-内酰胺酶的酶活力为113278.7 U/mL,为优化之前酶活(88792.7 U/mL)的1.28倍。本研究为进一步工业化开发利用性状稳定且高产β-内酰胺酶的菌株提供借鉴。

In order to improve the enzyme production ability of strain Bacillus cereus B03 quickly and efficiently,the fermentation medium of Bacillus cereus B03 producingβ-lactamase was optimized by response surface methodology.Firstly,the single factor experiments were conducted to study the different enzyme sources and concentrations,different nitrogen sources and concentrations,different metal ions and concentrations,sodium chloride,dipotassium hydrogen phosphate,temperature,pH,inoculum and liquid loading.Influence,then design the Plackett-Burman experiment to screen out three significant factors affecting enzyme production:Temperature,pH,inoculum.On this basis,design the Box-Behnken center combination test and response surface analysis method to determine the optimal enzyme fermentation conditions.The results showed that the optimum fermentation medium was 20 g/L glucose,20 g/L yeast extract,NaCl 2 g/L,MgSO4·7 H2O 0.2 g/L,K2HPO4·3 H2O 4 g/L.The optimum enzyme fermentation conditions were fermentation temperature 37℃,pH7.3,inoculum 3%,and liquid volume 50 mL/250 mL.Under this optimized condition,the enzyme activity of the strain producingβ-lactamase was 113278.7 U/mL,which was 1.28 times that of the enzyme activity before optimization(88792.7 U/mL).This study would provide a reference for further industrial development using strains with stable traits and high production ofβ-lactamase.