Analysis of Fatty Acid Metabolism Using Stable Isotope Tracers and Mass Spectrometry

Methods Enzymol. 2015;561:197-217. doi: 10.1016/bs.mie.2015.05.017. Epub 2015 Aug 17.

Abstract

Cells can synthesize fatty acids by ligating multiple acetyl units from acetyl-CoA. This is followed by desaturation and elongation reactions to produce a variety of fatty acids required for proper cellular functioning. Alternatively, exogenous lipid sources can contribute to cellular fatty acid pools. Here, we present a method based on incorporation of (13)C-carbon from labeled substrates into fatty acids and subsequent mass spectrometry analysis. The resulting labeling patterns can be used to determine (1) (13)C-enrichment of lipogenic acetyl-CoA, (2) the relative contributions of synthesis and uptake, and (3) absolute fatty acid fluxes. We begin by providing a background and general principles regarding the use of stable isotopes to study fatty acid metabolism. We then proceed with detailing procedures for sample preparation and both GC-MS and LC-MS analysis of isotope incorporation. Finally, we discuss the interpretation of the resulting fatty acid-labeling patterns.

Keywords: Acetyl-CoA; De novo lipogenesis; Fatty acid metabolism; Mass spectrometry; Stable isotopes; Tracing.

Publication types

  • Review

MeSH terms

  • Acetyl Coenzyme A / metabolism
  • Animals
  • Carbon Isotopes
  • Fatty Acids / metabolism*
  • Humans
  • Isotope Labeling / methods*
  • Lipid Metabolism
  • Mass Spectrometry / methods*

Substances

  • Carbon Isotopes
  • Fatty Acids
  • Acetyl Coenzyme A