响应面法优化棘孢木霉产厚垣孢子发酵工艺

Optimization of Fermentation Process for Chlamydospores of Trichoderma asperellum by Response Surface Methodology

采用响应面法（Box—Benhnken Design）对影响棘孢木霉（Trichoderma asperellum）ZJSX5003菌株产厚垣孢子的4个主要因素（黄豆饼粉．玉米粉用量，甘油添加量，装液量，初始pH）进行了研究。通过对试验数据进行方差分析考察了各因素及其交互作用对厚垣孢子产量的影响；进行回归分析并建立基于统计学的回归方程，计算获得最佳发酵条件：大豆饼粉．玉米粉用量33．25g／L、甘油添加量8．86ml／L、装液量99．35ml／500ml、初始pH值3．26，此时预测的厚垣孢子产量为9．56×107个／ml。在此条件下，实际测得厚垣孢子产量为9．84×107个／ml，达到理论预测值的97．07％，且比优化前提高了69．07％，进一步采用10L发酵罐进行适用性验证，厚垣孢子产量达到1．75X108个／ml，比摇瓶发酵的产量高，为木霉菌厚垣孢子菌剂开发提供了理论依据。

The response surface methodology （Box-Benhnken Design） was applied to investigate the four major factors （the concentration of soybean cake powder and maize powder used in medium, the volume of glycerol addition added to the medium, loading volume of liquid medium, initial pH value of medium） that have importent effect on the production of chlamydospores of Trichoderma asperellum ZJSXS003. By variance analysis on the experiment results, the influences of the the four factors and their interactions on the production of chlamydospores was investigated. By performing regression analysis and establishing regression equation based on statistics, the optimal fermentation condition was obtained, that as follow：the concentration of soybean cake powder and maize powder used in medium 33.25g/L, glycerol 8.86ml/L, each 500ml bottles contains 99.35ml liquid medium, initial pH 3.26. Under the optimal conditions, the yield of chlamydospore is expected to be 9.56 x 107 spores/ml, theoretically. The practical yield of chlamydospores approached 9.84 x 10v spores/ml, reached 97.07% of the expected value, increased by 69.07% compared with pre-optimization. In order to test its applicability, used a IOL fermentation tank under the optimal condition, the yield of chlamydospore reached up to 1.75 x lOSspores/ml, which is better than shake flask fermentation. Further researches of developing microbial inoculum of Trichoderma ehlamydospore could be conducted base on this research.