凌宏志，葛菁萍，平文祥，许修宏

1 东北农业大学资源与环境学院，哈尔滨 150080

2 黑龙江大学生命科学学院 微生物黑龙江省高校重点实验室，哈尔滨 150080

响应面法优化黑曲霉HDF05产β-葡萄糖苷酶过程参数

凌宏志1,2，葛菁萍2，平文祥2，许修宏1

1 东北农业大学资源与环境学院，哈尔滨 150080

2 黑龙江大学生命科学学院 微生物黑龙江省高校重点实验室，哈尔滨 150080

为获得黑曲霉 Aspergillus niger HDF05菌株较高的 β-葡萄糖苷酶酶活，对其发酵条件进行了优化。采用Plackett-Burman实验设计考察关键发酵操作参数对产酶的影响。继而采用最陡爬坡路径逼近最大响应区域，并结合中心组合实验和响应面对4个显著性因素进行分析。Plackett-Burman实验结果表明，发酵温度、装液量、麦麸和 (NH4)2SO4浓度对 β-葡萄糖苷酶合成影响显著。通过响应面分析得到一元二阶方程，对方程求解得到优化的发酵过程参数：发酵温度为28 ℃，装液量为71.4 mL/250 mL，麸皮浓度为36 g/L，(NH4)2SO4浓度为5.5 g/L。采用该优化的过程参数，菌株的最大产β-葡萄糖苷酶活力可达60.06 U/mL，较优化前提高了23.9%。将黑曲霉HDF05产生的β-葡萄糖苷酶用于酸解玉米芯纤维残渣的酶解实验中，可明显降低纤维二糖的积累，48 h内可使玉米芯纤维素残渣酶解得率达到80.4%。

黑曲霉，β-葡萄糖苷酶，响应面，过程参数优化

Abstract:In order to obtain high β-glucosidase productivity, we optimized the fermentation parameters for β-glucosidase production by Aspergillus niger HDF05. First, we screened the important parameters by Plackeet-burman design. Second, we usedthe path of steepest ascent to approach to the biggest response region of parameters affecting β-glucosidase production. Then, we obtained the optimal parameters by central composite design and response surface analysis. We developed a quadratic polynomial equation for predicting β-glucosidase production level. The results showed that the important parameters were temperature, packing volume, concentrations of wheat bran and (NH4)2SO4. The optimal fermentation parameters were as follows: temperature 28 °C,packing volume 71.4 mL/250 mL, wheat bran 36 g/L and (NH4)2SO45.5 g/L. Under the optimal conditions, we obtained the maximum enzyme activity of 60.06 U/mL, with an increase of 23.9% compared to the original fermentation parameters. During enzymatic hydrolysis of acid-pretreated corncob, addition of β-glucosidase from Aspergillus niger HDF05 greatly reduced the inhibition caused by cellobiose, and the hydrolysis yield was improved from 66.7% to 80.4%.

Keywords:Aspergillus niger, β-glucosidase, response surface methodology, parameters optimization

我国纤维质资源非常丰富，其中工业纤维质废渣每年有数千万吨。如能利用生物技术手段将其中一部分转化为酒精，无疑是解决现行酒精生产工艺中“与人争粮”的重要途径之一。而利用纤维质为原料生产酒精，纤维素酶的作用在其中起到至关重要的作用。……