基于GC-MS的阿维链霉菌代谢物组学研究方法的建立

Optimized Sample Preparation for Metabolome Studies on Streptomyces avermitilis by GC-MS

基于GC-MS分析平台,建立了阿维链霉菌代谢物组样品制备过程中的菌体分离、细胞清洗、细胞淬灭及代谢物提取条件,并对阿维菌素高产菌株和野生菌株胞内代谢物组进行了初步分析。采用"冷冻离心-细胞清洗(3次)-液氮淬灭(5min)-珠磨破壁(3个循环,每个循环48 s)-提取(氯仿∶乙醇∶水=2∶2∶1)"的方法,可以定性和定量检测阿维链霉菌胞内包括氨基酸、有机酸、脂肪酸、糖类等59种差异化合物。建立的方法能够有效地用于阿维链霉菌胞内代谢物的分析。

We established a method for preparing metabolome sample of Streptomyces avermitilis, including cell separation, washing, quenching, disruption and extraction conditions based on GC-MS analysis system, and applied it in the analysis of the metabolome of industrial strain 9-39 and wild type strain. The optimal conditions for sample preparation were as follows： separating cells by centrifugation, washing 3 times, quenching 5 minutes with liquid nitrogen, cell disruption with bead mill （ 3 cycles and 48 seconds per each cycle ） , and extraction of metabolites （ chloroform ： ethanol ： water = 2 ： 2 ： 1 ） . By using this optimized protocol, 59 differential compounds were identified and quantified, including amino acids, organic acids, fatty acids and saccharides. Our established method can be used to effectively analyze the Streptomyces avermitilis metabolites.