Abstract
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a useful tool to characterize the behavior of natural lipids within biological matrices. We report a LC-MS/MS method developed specifically to analyze CYP products of the arachidonoyl ethanolamide (anandamide, AEA), the epoxyeicosatrienoic acid ethanolamides (EET-EAs) and their hydrolyzed metabolites, and the dihydroxyeicosatrienoic acid ethanolamides (DHET-EAs). This method was used to measure EET-EA biotransformation to DHET-EAs by two human epoxide hydrolases: the soluble EH (sEH) and the microsomal EH (mEH). In general, sEH and mEH substrate preference was similar, based on kcat/KM. The 14,15-EET-EA and 11,12-EET-EA were the most efficiently hydrolyzed, followed by 8,9-EET-EA and 5,6-EET-EA. The method was also used to detect endogenous levels of these lipids in mouse tissues, although levels were below the instrumental detection limit (0.1–3.4 nM). Because both AEA and EETs are biologically active, the method described herein will be invaluable in revealing the role(s) of EET-EAs in vivo.
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Acknowledgments
This work received support in part from the National Institute of Environmental Health Sciences R01 ES002710 and NIEHS Superfund Program P42 ES004699. A. Rand acknowledges support from the OSCB training grant NIH/NIEHS T32 CA108459 and from the 2016 AACR Judah Folkman Fellowship for Angiogenesis Research, Grant Number 16-40-18-RAND.
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Rand, A.A., Helmer, P.O., Inceoglu, B., Hammock, B.D., Morisseau, C. (2018). LC-MS/MS Analysis of the Epoxides and Diols Derived from the Endocannabinoid Arachidonoyl Ethanolamide. In: Giera, M. (eds) Clinical Metabolomics. Methods in Molecular Biology, vol 1730. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7592-1_10
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DOI: https://doi.org/10.1007/978-1-4939-7592-1_10
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