响应面法优化革耳Panus rudis FG-35菌株产漆酶培养基

Optimization of Components for Laccase Production by Panus rudis FG-35 Using Response Surface Methodology

采用Plackett-Burman设计和响应面分析相结合的方法,对革耳Panus rudis FG-35菌株产漆酶的液体培养基配方进行优化。单因素试验结果显示,发酵培养基中的最优碳源为可溶性淀粉,最优氮源为蛋白胨;Plackett-Burman设计筛选出影响漆酶产量的3个重要因素为可溶性淀粉、金属Ca2+离子和吐温-40,在此基础上运用最陡爬坡试验逼近最大响应值区域,最后利用Box-Behnken试验设计及响应面分析法进行回归分析,获得最佳培养基配方为:可溶性淀粉10.040 4 g/L、蛋白胨0.2 g/L、K2HPO41.00g/L、ZnSO4·7H2O 0.008 g/L、MgSO4·7H2O 0.5 g/L、CuSO4·7H2O 0.007 g/L、FeSO4·7H2O 0.005 g/L、MnSO40.035 g/L、CaCl20.0816 g/L、VB10.1 g/L、吐温-40 0.428%。在优化后的条件下摇瓶发酵产漆酶酶活力为263.31 U/mL,与模型预测值接近,发酵产酶量比优化前提高1.07倍,同时优化后的发酵液对木质素降解进行试验发现,优化后漆酶对木质素降解率提高了14.34%。

We optimized the liquid medium components to produce the laccase activity of Panus rudis FG-35 by using Plackett-Burman design and response surface methodology. The results of single-factor test showed that soluble starch and peptone were the best sources of carbon and nitrogen. Then the Plackett-Burman design was applied to determine the specific medium components affecting laccase activity and found that soluble starch, Ca2＋and Tween-40 were the key factors. Based on these results, steepest ascent experiments were applied to find central points of laccase activity. These significant parameters were further optimized using Box-Behnken design, response surface methodology and regression analysis. Finally, the optimal medium was：soluble starch 10.040 4 g/L, peptone 0.2 g/L, K2HPO4 1.00 g/L, ZnSO4·7H2O 0.008 g/L, MgSO4·7H2O 0.5 g/L, CuSO4·7H2O 0.007 g/L, FeSO4·7H2O 0.005 g/L, MnSO4 0.035 g/L, CaCl2 0.081 6 g/L, VB1 0.1 g/L, Tween-40 0.428%. Under these optimal conditions, the activity of laccase increased from 263.31 U/mL to 127.194 U/mL（1.0-fold increase in total yield）, similar to the predictions. And the results of lignin degradation experiments indicated that the optimal medium made contribution to the degradation rate of lignin which about 14.34%improvement over before.