Analysis of Cell Metabolism Using LC-MS and Isotope Tracers

Here we discuss our methods to analyze small polar compounds involved in central carbon metabolism using LC-MS. Methods described include sample extraction procedures for cells and medium, as well as for plasma/serum, urine, CSF, and tissue samples. Different extraction solvents are assessed. Our me...

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Bibliographic Details
Published inMethods in enzymology Vol. 561; p. 171
Main Authors Mackay, Gillian M, Zheng, Liang, van den Broek, Niels J F, Gottlieb, Eyal
Format Journal Article
LanguageEnglish
Published United States 01.01.2015
Subjects
Animals
Carbon - metabolism
Carbon Isotopes
Chromatography, Liquid - methods
Humans
Isotope Labeling - methods
Mass Spectrometry - methods
Fumarate hydratase
LC-MS
Stable isotope tracers
HILIC chromatography
Exactive and Q-Exactive mass spectrometry
Metabolite extraction
Metabolite quantification
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Summary:Here we discuss our methods to analyze small polar compounds involved in central carbon metabolism using LC-MS. Methods described include sample extraction procedures for cells and medium, as well as for plasma/serum, urine, CSF, and tissue samples. Different extraction solvents are assessed. Our methods for using (13)C stable isotope tracers to examine the kinetics and distributions of mass isotopologues of many metabolites are discussed. Quantification methods are described for (13)C stable isotope tracer experiments as well as for unlabeled experiments. These methods were applied in a fumarate hydratase deficient cell model to show how isotope tracing can demonstrate shifts in metabolic pathways and, together with metabolite exchange rates, can be used to gain insights into changes in cell metabolism.
ISSN:1557-7988
DOI:10.1016/bs.mie.2015.05.016