红球菌利用小龙虾壳粉产几丁质脱乙酰酶的培养条件优化

Optimization of Cultivation Conditions of Rhodococcus for Producing Chitin Deacetylase Using Crayfish Shell Powder

为了利用小龙虾壳和降低几丁质脱乙酰酶的生产成本,以小龙虾壳粉作为培养基主要成分,接种红球菌11-3菌株发酵生产几丁质脱乙酰酶。首先对培养基进行蛋白酶水解预处理;然后,通过单因素试验、Plackett-Burman设计、最陡爬坡试验及Box-Behnken试验,对补充营养成分和环境条件进行优化。结果表明:19%虾壳粉起始培养基,用5 U/mg添加量的碱性蛋白酶处理4 h,补充1%蔗糖、0.25%硫酸铵和0.75%玉米浆,以3%的接种量接入红球菌,200 r/min振荡培养2.5 d,产酶活力高达2 723 U/mL,比起始培养基提高约53倍。以虾壳粉为主要成分的培养基可产生高酶活力的几丁质脱乙酰酶,为小龙虾壳的综合利用提供了一条新途径,也为未来使用生物法从虾壳中直接提取壳聚糖提供基础数据。

In order to comprehensively utilize the crayfish shell and reduce the production cost of chitin deacetylase,in this study we used crayfish shell powder as the main component of the medium,and inoculated with Rhodococcus sp.strain 11-3 to produce chitin deacetylase.The crayfish shell powder was firstly proteolytically hydrolyzed;then,supplemental nutrients and environmental conditions were optimized by single factor test,Plackett-Burman design,steepest climbing test and Box-Behnken test.The results showed that 19%crayfish shell powder medium was treated with alkaline protease at 5 U/mg for 4 h,supplemented with 1%sucrose,0.25%ammonium sulphate and 0.75%corn syrup.Rhodococcus was introduced into the medium at a seeding rate of 3%,and cultured at 200 r/min for 2.5 days.The enzyme production activity reached 2723 U/mL,which was about 53 times higher than the initial crayfish shell powder medium.Therefore,Rhodococcus in the obtained medium having crayfish shell powder as a main component can produce chitin deacetylase having high enzyme activity.It provides a new way for the comprehensive utilization of crayfish shells,and also provides a research basis for the future direct extraction of chitosan from crayfish shells using biological methods.