产几丁质酶菌株GXUN-20的筛选、鉴定及其产酶条件优化

Screening and Identification of Chitinase-producing Strain GXUN-20 and Optimization of Its Enzyme Producing Conditions

为了提高虾蟹等海产品废弃物利用率及其附加值,从广西合浦县大神木海鸭场附近滩涂沉积物(虾蟹及贝壳废弃物)中分离、纯化、筛选出一株产几丁质酶菌株GXUN-20,经过生理生化鉴定以及16S rDNA序列构建系统发育树,初步判断该菌株为厚壁溶杆菌(Lysobacter firmicutimachus)。以几丁质酶活力为指标,对GXUN-20菌株进行发酵条件单因素优化,在单因素优化结果较显著的4因素(氮源浓度、发酵液初始pH、几丁质粉末浓度、接种量)基础上进行4水平正交试验及正交实验方差分析。结果显示:氮源浓度对产酶影响显著(P=0.013<0.05),在氮源为1 g/L黄豆饼粉,几丁质粉末为20 g/L,接种量为3%,发酵液初始pH8.4,30℃、200 r/min发酵6 d的条件下,发酵液中几丁质酶活力最大,达到0.965 U/mL,是优化前酶活力的6.89倍。本研究结果为GXUN-20菌株产几丁质酶进一步开发与应用提供了基础数据和实验方法。

In order to improve that utilization rate and added value of seafood wastes such as shrimp and crab. The chitinase-producing strain GXUN-20 was isolated, purified and screened from the mudflat sediments(shrimp, crab and shell wastes) near Damenhai Duck Farm in Hepu county, Guangxi Province. After physiological and biochemical identification and the construction of phylogenetic tree based on 16 S rDNA sequence, the strain was preliminarily identified as Lysobacter firmicutimachus. Taking chitinase activity as index, the single factors optimization of fermentation conditions were conducted for strain GXUN-20. On the basis of four factors(nitrogen source concentration, initial pH of fermentation broth, chitin powder concentration and inoculation amount) that had significant results in the single factor optimization, four levels of orthogonal experiment and variance analysis of orthogonal experiment were carried out. The results showed that the concentration of nitrogen source had a significant effect on enzyme production(P=0.013<0.05). When the nitrogen source was 1 g/L soybean cake powder, chitin powder was 20 g/L, the inoculation amount was 3%, the initial pH of the fermentation broth was 8.4, and the fermentation temperature was 30 ℃and 200 r/min for 6 days, the chitinase activity in the fermentation broth reached the maximum value of 0.965 U/mL, which was 6.89 times of the enzyme activity before optimization. The results of this study would provide the basic data and experimental methods for the further development and application of the chitinase produced by GXUN-20 strain.