响应面法和变温培养优化海洋黑曲霉产纤维素酶发酵条件

Optimization of Fermentation Conditions of Cellulase Produced by a Marine Aspergillus Niger with Response Surface Methodology and Variable Temperature Culture

选用响应面法和变温培养相结合的方法,优化了一种海洋黑曲霉生产耐盐性纤维素酶的液体发酵条件。首先通过单因素实验初步确定碳源、氮源、温度、表面活性剂、接种量、装液量以及初始pH;随后选择Plackett-Burman(PB)设计挑选出影响滤纸酶活(FPA)的主要成分:麸皮和吐温-80(Tween-80);再通过爬坡实验,使麸皮和吐温-80浓度接近最大酶活区域;接着选择响应面分析快速有效地求得该菌株产耐盐性纤维素酶的最佳培养基配方;最后进行了变温培养实验确定适宜的稳定期培养温度。得到的优化发酵条件为:麸皮5.53%(w),玉米浆0.5%(w),磷酸二氢钾0.2%(w),Tween-80 0.197%(w),初始pH 5.0,接种量8%(mol),装液量40 m L,180 r/min,37℃培养2 d后继续28℃培养2 d。优化后的滤纸酶活为0.566 U/m L,与未优化时的酶活(0.283 U/m L)相比增加了94.5%。

The response surface analysis and variable temperature culture were used to optimize the liquid fermentation conditions for the production of salt-tolerant cellulose by a marine Aspergillus niger. Firstly, the single factor tests were carried out to determine the carbon source, nitrogen source, temperature, surfactant, inoculum amount, liquid volume and initial pH. Then the Plackett-Burman（PB） design was used and the wheat bran and Tween-80 were identified as the two major components that affect the filter paper activity（FPA）. The concentrations of these two components were ascertained to be close to the maximum enzyme activity area by steepest ascent path. Finally, the response surface analysis was used to obtain the optimal fermentation medium formulation of salt-tolerance cellulose and the variable temperature experiment was carried out to determine the optimum culture temperature. The optimal formulation was determined as 40 m L of medium consisting of 5.53 % wheat bran, 0.5 % corn steep liquor, 0.2 % KH_2PO4, 0.197 % Tween-80 in a 150 m L flask, and the optimal fermentation conditions were initial pH of 5, inoculum concentration of 8%, liquid volume of 40 m L at 180 r/min, culture temperature at 37 ℃ for 2 days followed at 28 ℃ for another 2 days. The maximum FPA under the optimized conditions was 0.566 U/m L, which was increased by 94.5 % compared with the 0.283 U/m L obtained before optimization.