A liquid chromatography/tandem mass spectrometry method for measuring the in vivo incorporation of plasma free fatty acids into intramyocellular ceramides in humans
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RATIONALE: Sphingolipids are important components of cell membranes that serve as cell signaling molecules; ceramide plays a central role in sphingolipid metabolism. De novo ceramide biosynthesis depends on fatty acid availability, but whether muscle uses circulating free fatty acids or pre‐existing intracellular stores is unknown. Our goal was to develop a method to detect the incorporation of intravenously infused [U‐ 13 C]palmitate into intramyocellular ceramides. METHODS: We used liquid chromatography/tandem mass spectrometry (LC/MS/MS) to measure the concentrations of different sphingolipid species and 13 C‐isotopic enrichment of 16:0‐ceramide. Chromatographic separation was performed using ultra‐performance liquid chromatography. The analysis was performed on a triple quadrupole mass spectrometer using a positive ion electrospray ionization source with selected reaction monitoring (SRM). RESULTS: The sphingolipids ions, except enriched ceramide, were monitored as [M+2+H] + . The [ 13 C 16 ]16:0‐ceramide was monitored as [M+16+H] + . By monitoring two different transitions of the [ 13 C 16 ]16:0‐ceramide (554/536 and 554/264) we could indirectly measure enrichment of the palmitate that is not a part of the sphingoid base. Concentration and enrichment could be measured using 20 mg of muscle obtained from volunteers receiving a low dose [U‐ 13 C]palmitate infusion. CONCLUSIONS: LC/MS/MS can be used to detect the incorporation of plasma palmitate into muscle ceramides in humans, in vivo . Copyright © 2012 John Wiley & Sons, Ltd.
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Different approaches for the measurement of ceramide synthesis in vivo have been proposed, e.g. infusion of U13C palmitate as ceramide precursors in humans to measure de novo synthesis in skeletal muscle or labelled palmitate to measure hepatic ceramide kinetics in rat models.
“Different approaches for the measurement of ceramide synthesis in vivo have been proposed, e.g. the infusion of U13C palmitate as ceramide precursors, used in humans to measure de novo synthesis of ceramides in skeletal muscle[119], [120], [121] or labelled palmitate used to measure hepatic ceramide kinetics in rat models.122,123”
From Lipid metabolism in MASLD and MASH: From mechanism to the clinic (2024) · cites this paper as [119]
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- Authors
- Blachnio‐Zabielska, A. U., Persson, X. T., Koutsari, C., Zabielski, P., & Jensen, M. D.
- Affiliations
- Endocrine Research Unit Mayo Clinic Rochester MN 55905 USA
- Mayo Clinic (US)
- Venue
- Rapid Communications in Mass Spectrometry
- Publisher
- Wiley
- Published
- 2012-03-22
- Volume
- 26
- Issue
- 9
- Pages
- 1134-1140
- ISSN
- 0951-4198 (ISSN-L)
- 1097-0231
- PMC
- PMC3370409
- OpenAlex
- W1608393181
- Cited by
- 102
- References
- 24
- Citations by year
- 2026: 2 · 2025: 7 · 2024: 7 · 2023: 8 · 2022: 5 · 2021: 8 · 2020: 4 · 2019: 11 · 2018: 10 · 2017: 9 · 2016: 9 · 2015: 6 · 2014: 6 · 2013: 7 · 2012: 3
- FWCI
- 2.83
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Rapid Communications in Mass Spectrometry (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- Funders
- American Diabetes Association
- National Institutes of Health (UL1TR000135)
- U.S. Public Health Service
- National Institute of Diabetes and Digestive and Kidney Diseases (DK 40484, P30DK050456, DK 45343, DK50456, R37DK040484, R01DK045343, R01DK040484)
- National Center for Research Resources (RR-00585, M01RR000585, UL1RR024150)
- National Center for Advancing Translational Sciences
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Sphingolipid Metabolism and Signaling
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolomics and Mass Spectrometry Studies
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Lipid metabolism and biosynthesis
- Keywords
- ceramide, sphingolipid metabolism, palmitate incorporation, isotope enrichment, muscle, plasma free fatty acids, ceramide biosynthesis
- MeSH terms
- Carbon Isotopes, Ceramides, Chromatography, High Pressure Liquid, Humans, Palmitates, Linear Models, Muscle, Skeletal, Tandem Mass Spectrometry
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