期刊
ANALYTICAL CHEMISTRY
卷 83, 期 6, 页码 2152-2161出版社
AMER CHEMICAL SOC
DOI: 10.1021/ac102981k
关键词
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资金
- California Institute of Regenerative Medicine [TR1-01219]
- National Institutes of Health [R24 EY017540-04, P30 MH062261-10, P01 DA026146-02]
- NIH/NIA [L30 AG0 038036]
- Department of Energy [FG02-07ER64325, DE-AC0205CH11231]
Mass spectrometry-based metabolomics is the comprehensive study of naturally occurring small molecules collectively known as the metabolome. Given the vast structural diversity and chemical properties of endogenous metabolites, biological extraction and chromatography methods bias the number, property, and concentration of metabolites detected by mass spectrometry and creates a challenge for global untargeted studies. In this work, we used Escherichia coli bacterial cells to explore the influence of solvent polarity, temperature, and pH in extracting polar and nonpolar metabolites simultaneously. In addition, we explored chromatographic conditions involving different stationary and mobile phases that optimize the separation and ionization of endogenous metabolite extracts as well as a mixture of synthetic standards. Our results reveal that hot polar solvents are the most efficient in extracting both hydrophilic and hydrophobic metabolites simultaneously. In addition, ammonium fluoride in the mobile phase substantially improved ionization efficiency in negative electrospray ionization mode by an average increase in signal intensity of 5.7 and over a 2-fold increase in the total number of features detected. The improvement in sensitivity with ammonium fluoride resulted in 3.5 times as many metabolite hits in databases compared to ammonium acetate or formic acid enriched mobile phases and allowed for the identification of unique metabolites involved in fundamental cellular pathways.
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