一株快速转化甜菊苷的细菌鉴定、产酶及转化特性  

Identification,culture optimization and biotransformation of a stevioside-degrading bacterium

作　　者：刘虎 陈育如[1,2] 姜中玉[1] 

机构地区：[1]南京师范大学生命科学学院,江苏省微生物资源工程技术研究中心,南京210046 [2]南京师范大学泰州学院生物技术系,泰州225300

出　　处：《微生物学报》2010年第7期885-890,共6页Acta Microbiologica Sinica

基　　金：林业公益性行业科研专项基金(200904055)~~

摘　　要：【目的】本研究鉴定了一株筛选的甜菊苷特异降解菌、优化了该菌产β-葡萄糖苷酶的条件以及研究了该菌对甜菊苷的转化特性。【方法】经16SrRNA基因序列测序和系统发育学分析,结合形态学特征确定该菌株的系统发育地位。用单因素及多因素分析探讨了其对甜菊苷的降解,通过液质联用检测了降解产物。【结果】菌株-J2与巨大芽孢杆菌的16SrRNA基因序列相似性达到100%,结合形态学特征,鉴定该菌为巨大芽孢杆菌(Bacillus megaterium)。在玉米淀粉4%、豆粕粉1%、硫酸镁0.04%、pH7.0、37℃、220r/min、接种量10%、培养36h的条件下,该菌产β-葡萄糖苷酶活力为779.68U/mL。甜菊苷转化的结果表明:3d可将10mg/mL甜菊糖溶液中甜菊苷转化74%,使莱鲍迪甙A和甜菊苷的比例(RA/SS)由转化前的0.38上升至0.99,RA的相对量增加160.5%,5d时转化完全。转化产物经液质联用鉴定为甜菊双糖甙。【结论】确定菌株-J2为巨大芽孢杆菌(Bacillus megaterium),该菌对甜菊苷具有高效、特异的转化能力,为首次报道的新型、安全菌株。[Objective]Our study aimed at screening and identifying a specific bacterium capable of degrading stevioside.We also studied the conditions of enzyme production and stevioside conversion.[Methods]Taxonomic group of the strain was confirmed by physical characterization and phylogenetic analysis by 16S rRNA gene sequence analysis and phylogenetic tree construction of the strain.The optimum conditions of enzyme producing and stevioside degrading were studied by single factor and multi-factor statistical analysis.Degradation product was detected and identified via liquid chromatography-mass spectrometry.[Results]Based on the result of 16S rRNA gene sequence analysis,the strain named J2 shares 100% sequence identity with the sequence of the Bacillus megaterium.The activity of beta-Glucosidase produced by this Bacillus megaterium strain was up to 779.68 U /ml with 4% maize starch,1% defatted soybean,0.04% MgSO4 and 0.2% stevioside as culture medium when fermented under the condition of pH 7.0,37℃,220 r /min and 10% inoculum for 36 h.The results of conversion showed that 10 mg /ml stevioside can be converted to steviolbioside by 74% after 3 days which has been identified by LC-MS.The ratio of rebaudioside A and stevioside was increased to 0.99 compared to original solution 0.38,which lead to 160.5% increasement of rebaudioside A in the relative amount.Stevioside can be converted completely after 5 days.[Conclusion] The isolated strain J2 was identified as Bacillus megaterium.It was a novel and safe strain with high,specific conversion stevioside to steviolbioside ability.

关 键 词：甜菊苷 生物转化 莱鲍迪甙A Β-葡萄糖苷酶 巨大芽孢杆菌 

分 类 号：TQ920.1[轻工技术与工程—发酵工程]