书虱伴生菌中纤维素酶产生菌的筛选、鉴定及最佳产酶发酵条件的优化

从以啃食树皮为生的书虱伴生菌中筛选得到5株产纤维素酶的菌株,编号分别为S2、S6、N10、N11和N12。结合菌株的形态学及16S rDNA序列分析等结果将这些菌株分别鉴定为Bacillus methylotrophicus, Streptomyces sp., Pseudomans fluorescens, Bacillus sp.及Pseudomans sp.。分别使用单因素分析法和响应面分析法对纤维素酶活最高的S2菌株的产酶发酵条件进行了优化。单因素实验结果显示,B. methylotrophicus S2的最适产酶发酵条件为:0.01 g/mL 的CMC碳源、10 g/L蛋白胨的氮源、48 h的发酵时间、28℃、初始pH为7.0,此时总纤维素酶活达到204.37 u/g。然后选取发酵时间、温度和初始pH作为3个因素,通过BBD实验,用响应面法对S2的发酵条件进行优化分析,最后得到一个拟合度良好的二次多项方程模型(R2=0.9948) 。方差分析结果显示,发酵温度与培养基初始pH之间的交互作用极显著。响应面分析优化后的反应体系为: 温度24℃、初始pH为7.7、发酵59 h,测得酶活力为303.18 u/g,比优化前单因素最佳纤维素酶活力提高45%。

Screening,identification cellulose-degrading bacteria associated with booklice and optimization of fermentation conditions

Five bacteria numbered S2, S6, N10, N11, N12 were isolated from booklice with bark for food. The bacterial strains were classified to be individually Bacillus methylotrophicus, Streptomyces sp., Pseudomans fluorescens, Bacillus sp. and Pseudomans sp. by phenotypic and genotypic characteristics.We aimed to study the influencing factors and to investigate the combined interactions between them by applying conventional one-factor-at-a-time (OFAT) method and response surface methodology (RSM) for cellulase activates using S2 as targeted strain. Through the OFAT method, the optimum conditions for fermentation were 0.01 g/mL CMC as carbon source, 10 g/L peptone as nitrogen source, 48 h, 28℃,pH 7.0. Under these conditions, the total cellulase activity reached to 204.37 u/g. Moreover, in order to optimize the reaction fermentation conditions, the fermentation time, temperature and pH were selected as three factors for systematical analysis. Response surface analysis was conducted through BBD and a reduced cubic model with a good fit of regression equation (R2=0.9948) was generated. The optimum conditions through RSM were found to be 59 h, 24℃,initial pH 7.7, for the maximum activity 303.18 u/g, which increased by 45%.