Development of Isotope Labeling Liquid Chromatography–Mass Spectrometry for Metabolic Profiling of Bacterial Cells and Its Application for Bacterial Differentiation

Quantitative and comprehensive profiling of cellular metabolites is currently a challenging task in bacterial metabolomics. In this work, a simple and robust method for profiling the amine- and phenol-containing metabolome of bacterial cells is described. The overall workflow consists of methanol-based cell lysis and metabolite extraction with ultrasonication, differential isotope dansylation labeling of cellular metabolites, and analysis of the labeled metabolites by liquid chromatography–mass spectrometry (LC–MS). Over a thousand peak pairs or putative metabolites can be detected from bacterial cells in a 25 min LC–MS run and near 2500 putative metabolites can be found in one bacterium from combined results of multiple analyses. After careful examination and optimization of the sample preparation process, this method is shown to be effective for both Gram-positive and Gram-negative bacteria. An idea of applying LC–ultraviolet (UV) detection to quantify the total amount of labeled metabolites is shown to...