基于HPLC-MS/MS快速筛选阿卡波糖及其类似物的菌株

Rapid Screening of Acarbose and Its Analogs in Fermentation Broth Based on HPLC-MS/MS

依据阿卡波糖及其类似物的质谱特征碎片,建立了高效液相色谱-串联质谱(HPLC-MS/MS)法从20种不同来源的发酵液中快速筛选阿卡波糖及其类似物产生菌。发酵液经阳离子型固相萃取柱纯化后,采用HPLC-MS/MS筛选分析。通过归纳阿卡波糖及其类似物的特征离子碎片,总结裂解途径,得出具有C7N单元化合物的共有特征碎片,并以质谱能量分辨曲线对实验结果加以判断。该分析方法结合超高效液相色谱分离与质谱多反应监测扫描模式,可实现阿卡波糖及其类似物产生菌的筛选,对其选育提供了重要的实验思路。

High performance liquid chromatography tandem mass spectrometry(HPLC-MS/MS)is a comprehensive analytical technology developed in recent years for structural identification,qualitative and quantitative detection of compound components.Acarbose is currently one of the most widely used drugs in the treatment of typeⅡdiabetes,and it has important medical significance.In this experiment,based on the fragments mass spectrometric of acarbose and its analogs,HPLC-MS/MS method was established for rapid screening of acarbose and its analog-producing bacteria from dozens of fermentation broths with different sources.After the fermentation broth was purified by a cationic solid-phase extraction column,simple dilution and centrifugation,separation was performed with a Waters CORTECS UPLC C18+chromatography column.Methanol-water was selected as the mobile phase with the speed of 0.2 mL/min under gradient conditions.At positive ion mode,twenty strains of fermentation broth from different sources were screened experimentally.They were monitored by multiple reaction monitoring,parent ion scanning mode to detect acarbose,the analogs,and unknown compounds with a core structure of C7 N.By summarizing the characteristic product ion fragments of acarbose and its analogs,the cleavage pathways of acarbose and its analogs were summarized,and the common characteristic fragments of compounds with C7 N units were obtained,which was m/z 304.The two isomers of acarbose and acarbose impurity A were determined by mass spectrum energy resolution curve.The analysis method can realize the screening of acarbose and its analogue-producing bacteria,and provides the breeding of acarbose and important experimental ideas of its analogue-producing bacteria.