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Quantitation of fatty acyl-coenzyme As in mammalian cells by liquid chromatography-electrospray ionization tandem mass spectrometry*

By Christopher A. Haynes, Jeremy C. Allegood, Kacee Sims, Elaine W. Wang, M. Cameron Sullards and Alfred H. Merrill


Fatty acyl-CoAs participate in numerous cellular processes. This article describes a method for the quantitation of subpicomole amounts of long-chain and very-long-chain fatty acyl-CoAs by reverse-phase LC combined with electrospray ionization tandem mass spectrometry in positive ion mode with odd-chain-length fatty acyl-CoAs as internal standards. This method is applicable to a wide range of species [at least myristoyl- (C14:0-) to cerotoyl- (C26:0-) CoA] in modest numbers of cells in culture (∼106–107), with analyses of RAW264.7 cells and MCF7 cells given as examples. Analysis of these cells revealed large differences in fatty acyl-CoA amounts (12 ± 1.0 pmol/106 RAW264.7 cells vs. 80.4 ± 6.1 pmol/106 MCF7 cells) and subspecies distribution. Very-long-chain fatty acyl-CoAs with alkyl chain lengths > C20 constitute <10% of the total fatty acyl-CoAs of RAW264.7 cells versus >50% for MCF7 cells, which somewhat astonishingly contain approximately as much C24:0- and C26:0-CoAs as C16:0- and C18:0-CoAs and essentially equal amounts of C26:1- and C18:1-CoAs. This simple and robust method should facilitate the inclusion of this family of compounds in “lipidomics” and “metabolomics” studies

Topics: Methods
Publisher: American Society for Biochemistry and Molecular Biology
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Provided by: PubMed Central
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