title: Analysis of Cell Metabolism Using LC-MS and Isotope Tracers.
abstract: 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.
authors.affiliation: Cancer Research UK Beatson Institute, Glasgow, UK. Electronic address: g.mackay@beatson.gla.ac.uk.
authors.forename: Gillian M
authors.initials: GM
authors.lastname: Mackay
authors.affiliation: Cancer Research UK Beatson Institute, Glasgow, UK.
authors.forename: Liang
authors.initials: L
authors.lastname: Zheng
authors.affiliation: Cancer Research UK Beatson Institute, Glasgow, UK.
authors.forename: Niels J F
authors.initials: NJ
authors.lastname: van den Broek
authors.affiliation: Cancer Research UK Beatson Institute, Glasgow, UK. Electronic address: e.gottlieb@beatson.gla.ac.uk.
authors.forename: Eyal
authors.initials: E
authors.lastname: Gottlieb
mesh_terms: D000818:Animals; D002244:Carbon / Q000378:metabolism*; D002247:Carbon Isotopes; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*
chemical_list: D002247:Carbon Isotopes; D002244:Carbon
keywords: Exactive and Q-Exactive mass spectrometry; Fumarate hydratase; HILIC chromatography; LC-MS; Metabolite extraction; Metabolite quantification; Stable isotope tracers