Evaluation and optimization of analytical procedure and sample preparation for polar Streptomyces albus J1074...

Metabolomics is an essential discipline in omics technology that promotes research on the biology of microbial systems. Streptomyces albus J1074 is a model organism used in fundamental research and industrial microbiology. Nevertheless, a comprehensive and standardized method for analyzing the metabolome of S. albus J1074 is yet to be developed. Thus, we comprehensively evaluated and optimized the analytical procedure and sample preparation for profiling polar metabolites using hydrophilic interaction liquid chromatography (HILIC) coupled with high-resolution mass spectrometry (HRMS). We systematically examined the HILIC columns, quenching solutions, sample-to-quenching ratios, and extraction methods. Then, the optimal protocol was used to investigate the dynamic intracellular polar metabolite profile of the engineered S. albus J1074 strains during spinosad (spinosyn A and spinosyn D) fermentation. A total of 3648 compounds were detected, and 83 metabolites were matched to the standards. The intracellular metabolomic profiles of engineered S. albus J1074 strains (ADE-AP and OE3) were detected; furthermore, their metabolomes in different stages were analyzed to reveal the reasons for their differences in their spinosad production, as well as the current metabolic limitation of heterologous spinosad production in S. albus J1074. The HILIC-HRMS method is a valuable tool for investigating polar metabolomes, and provides a reference methodology to study other Streptomyces metabolomes.