产漆酶细菌筛选鉴定及固体发酵条件研究

Screening and Identification of Laccase-producing Bacteria and Study on Solid-state Fermentation Conditions

为了获得能降解木质素的细菌,采用愈创木酚法和BR亮蓝平板法从枯枝落叶自然堆积物中分离漆酶活力较高的菌株,通过形态学观察、生理生化试验,16S rRNA序列分析及系统发育树的构建对菌株进行鉴定。以芦苇秸秆粉为主要原料,采用单因素试验和正交试验研究并优化了菌株适宜的发酵培养基和发酵条件。结果表明,菌株H-1鉴定为蜡状芽孢杆菌H-1(Bacillus cereus H-1);适宜的发酵培养基和发酵条件为芦苇粉100%,(NH4)2SO 42.5%,酵母抽提粉0.3%,KH2PO40.15%,MgSO4·7H2O 0.15%,NaCl 0.5%,CaCl 20.1%,发酵温度35℃,固水比1.0∶1.1(g∶mL),初始pH 7.2~7.4,接种量3%,装量50 g,发酵时间144 h。在此条件下,漆酶活力达0.992 U/g。

In order to obtain lignin-degrading bacteria,the strains with high laccase activity were isolated from natural litter by guaiacol and BR brilliant blue plate method which were identified by morphological observation,physiological and biochemical experiments,16S rRNA sequence analysis and phylogenetic tree construction.The suitable fermentation medium and fermentation conditions of the strain were optimized by single factor and orthogonal experiments with reed straw powder as the main raw material.The results showed that strain H-1 was identified as Bacillus cereus H-1.The suitable fermentation medium and conditions were as follows:reed powder 100%,(NH4)2SO 42.5%,yeast extract powder 0.3%,KH2PO40.15%,MgSO4·7H2O 0.15%,NaCl 0.5%,CaCl 20.1%,fermentation temperature 35℃,solid-water ratio 1.0∶1.1(g∶mL),initial pH 7.2-7.4,inoculation volume 3%,loading capacity 50 g,fermentation time 144 h.Under those conditions,laccase activity reached 0.992 U/g.