基于复合诱导剂诱导枯草工程菌产饲用蛋白酶的响应面优化研究

Response surface optimization study on production of feed proteinase by Bacillus subtilis with composite inducers

试验为建立一种采用乳糖和异丙基-β-D-硫代半乳糖苷(IPTG)复合诱导剂代替IPTG作为诱导剂进行枯草芽孢杆菌产饲用中性蛋白酶发酵工艺。通过初步筛选获得最佳诱导剂配比为0.025‰IPTG和11.25 g/L乳糖,在原有工艺下发酵得到饲用蛋白酶活性为1066.09 U/mL;经过单因素试验和响应面优化发酵工艺,获得最佳发酵工艺条件为初始p H值7.5、培养温度36.5℃、诱导剂添加时间3.2 h、发酵时间45 h,中性蛋白酶活性为1398.54 U/mL,提高至优化前的1.31倍。研究表明,复合剂诱导条件比IPTG单独诱导更复杂,在蛋白酶的规模化生产中具有潜在的工业应用价值,为蛋白酶制剂在饲料添加剂的应用提供了基础。

The experiment aimed to establish a fermentation process for the production of neutral protease for feed using a composite inducer of lactose and isopropyl-β-D-thiogalactopyranoside(IPTG)as a substitute for IPTG in Bacillus subtilis.Through preliminary screening,the optimal inducer ratio was determined to be 0.025‰IPTG and 11.25 g/L lactose,resulting in a feed protease activity of 1066.09 U/mL under the original process.After optimization of the fermentation process through single-factor experiments and response surface methodology,the best fermentation conditions were found to be an initial pH of 7.5,cultivation temperature of 36.5℃,inducer addition time of 3.2 hours,and fermentation time of 45 hours,yielding a neutral protease activity of 1398.54 U/mL,which was 1.31 times higher than before optimization.The study indicates that the induction conditions with the composite inducer are more complex than those with IPTG alone,and it has potential industrial application value in the large-scale production of protease.It provides a foundation for the application of protease preparations as feed additives.