School of Life Sciences, Liaocheng University, Liaocheng 252059, People's Republic of China.
World J Microbiol Biotechnol. 2012 Mar;28(3):1163-74. doi: 10.1007/s11274-011-0919-8. Epub 2011 Oct 18.
Solid-state fermentation conditions for cellulases production by a newly isolated Penicillium chrysogenum QML-2 were investigated using statistical methods. At first, significant variables for cellulases production including (NH(4))(2)SO(4), initial pH and inoculum size were screened by using Plackett-Burman Design. Then the optimal regions of the significant variables were investigated by using the method of steepest ascent. Finally, central composite design and response surface analysis were adopted to determine the optimal values of the significant variables and investigate the combined effects of each variable's pair on cellulases production. The results showed that the optimal ranges of (NH(4))(2)SO(4) concentration, initial pH and inoculum size for three types of cellulases activities were 1.97-2.15 g, pH 4.32-4.41 and 13.3-13.7% (v/w), respectively. Using the mixture of corn stover powder and wheat bran (CSP/WB, 1/1) as carbon source, the optimization resulted in 370.15, 101.76 and 321.56 U/g for maximal endoglucanase activity, filter paper activity and β-glucosidase activity, respectively. Compared with maximum values of cellulases activities (endoglucanase activity 85.21 U/g, filter paper activity 16.62 U/g and β-glucosidase activity 67.68 U/g) obtained under unoptimized conditions, the optimization resulted in 3.34, 5.12 and 3.75 folds improvement for endoglucanase activity, filter paper activity and β-glucosidase activity, respectively. For chitosan hydrolysis, the crude cellulases had the optimal temperature of 55°C, pH of 4.4 and exhibited Michaelis constant (K (m)) value of 8.34 mg/ml and maximum velocity (V (max)) of 2.21 μmol glucosamine/min by 1 ml of the crude cellulases.
采用统计方法研究了新分离的产黄青霉 QML-2 固态发酵生产纤维素酶的条件。首先，利用 Plackett-Burman 设计筛选出纤维素酶生产的显著变量，包括(NH4)2SO4、初始 pH 和接种量。然后，采用最陡爬坡法考察显著变量的最优区域。最后，采用中心复合设计和响应面分析确定显著变量的最优值，并考察各变量对纤维素酶生产的组合效应。结果表明，三种纤维素酶活性的(NH4)2SO4浓度、初始 pH 和接种量的最佳范围分别为 1.97-2.15 g、pH 4.32-4.41 和 13.3-13.7%（v/w）。采用玉米秸秆粉和麦麸（CSP/WB，1/1）混合物作为碳源，优化后内切葡聚糖酶活性、滤纸酶活性和β-葡萄糖苷酶活性分别达到 370.15、101.76 和 321.56 U/g。与未优化条件下纤维素酶活性的最大值（内切葡聚糖酶活性 85.21 U/g、滤纸酶活性 16.62 U/g 和β-葡萄糖苷酶活性 67.68 U/g）相比，优化后内切葡聚糖酶活性、滤纸酶活性和β-葡萄糖苷酶活性分别提高了 3.34、5.12 和 3.75 倍。壳聚糖水解试验表明，粗酶的最适温度为 55°C，最适 pH 为 4.4，米氏常数（K(m)）值为 8.34 mg/ml，最大反应速度（V(max)）为 2.21 μmol 氨基葡萄糖/min，每毫升粗酶用量为 1 ml。
