一株将甜菊苷转化为甜茶甙的细菌鉴定及转化特性

摘要

[Objective]The purpose of our study was to separate and identify a bacillus that could convert stevioside specifically.Then we identified the conversion product and studied the conversion capability of the bacillus.We also studied the enzyme with conversion capability and the conversion characteristic of the enzyme.[Methods]The bacillus was identified on the basis of morphology features and 16S rDNA sequence analysis.Phylogenetic tree was constructed to determine its taxonomic status.The product was detected and identified by high performance liquid chromatography and liquid chromatography-mass spectrometry methods.We use bacteria media directly to studied the conversion capability of the bacillus, and use resting cells, extracellular fluid and intracellular fluid to convert stevioside to determine the enzyme and made further study to learn its conversion characteristic.[Results]The 16S rDNA sequence of the strain had 99％ similarity with Chryseobacterium sp., which was ultimately identified as Chryseobacterium sp.JH.The product of biotransformation was rubusoside and the enzyme that converts stevioside into rubusoside was intracellular enzyme.The conversion rate could reach 100％ , obtained 5.7 g/L rubusodide solution after 48 h by bacteria media when the concentration of stevia glycosides was 10 g/L, including 7.2 g/L stevioside.[Conclusion]The isolated strain JH was identified as Chryseobacterium sp..It was a novel strain with high, specific ability to convert stevioside into rubusoside which had potential applications.%[目的]筛选一株对甜菊苷具有特异转化性能的细菌,并对该菌及转化产物进行鉴定,探讨转化酶及酶对甜菊苷的转化特性.[方法]通过16S rDNA序列分析,构建该菌系统进化树,结合菌体形态及菌落特征,确立该菌系统发育学地位.通过高效液相色谱(HPLC)及液质联用(LC-MS)法检测并鉴定转化产物.用菌液直接对甜菊苷进行转化以研究菌的转化能力.用静息细胞、胞外液和胞内液对甜菊糖分别转化法,确定转化酶与菌体的关系,并用该酶液进行转化特性研究.[结果]该菌株与黄杆菌属的16S rDNA序列相似性为99%,结合菌体及菌落的形态特征,鉴定该菌为黄杆菌属(Chryseobacterium sp.)黄杆菌.转化产物经高效液相色谱及液质联用法鉴定为甜茶甙.用菌液直接转化,48 h可将10 g/L甜菊糖中甜菊苷(7.2 g/L)完全转化,转化率达100%,得到5.7 g/L甜茶甙溶液.[结论]确定该菌为黄杆菌(Chryseobacterium sp.),能将甜菊苷高效、特异性地转化为甜茶甙.