title: Assay of Endocannabinoid Oxidation by Cyclooxygenase-2.
abstract: The endocannabinoids, 2-arachidonoylglycerol (2-AG) and arachidonylethanolamide (AEA), are endogenous ligands for the cannabinoid receptors (CB1 and CB2) and are implicated in a wide array of physiological processes. These neutral arachidonic acid (AA) derivatives have been identified as efficient substrates for the second isoform of the cyclooxygenase enzyme (COX-2). A diverse family of prostaglandin glycerol esters (PG-Gs) and prostaglandin ethanolamides (PG-EAs) is generated by the action of COX-2 (and downstream prostaglandin synthases) on 2-AG and AEA. As the biological importance of the endocannabinoid system becomes more apparent, there is a tremendous need for robust, sensitive, and efficient analytical methodology for the endocannabinoids and their metabolites. In this chapter, we describe methodology suitable for carrying out oxygenation of endocannabinoids by COX-2, and analysis of products of endocannabinoid oxygenation by COX-2 and of endocannabinoids themselves from in vitro and cell assays.
authors.affiliation: A.B. Hancock Jr. Memorial Laboratory for Cancer Research, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.|Department of Biochemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.
authors.forename: Shalley N
authors.initials: SN
authors.lastname: Kudalkar
authors.affiliation: A.B. Hancock Jr. Memorial Laboratory for Cancer Research, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.|Department of Biochemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.|Department of Chemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA.
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Kingsley
authors.affiliation: A.B. Hancock Jr. Memorial Laboratory for Cancer Research, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA. larry.marnett@vanderbilt.edu.|Department of Biochemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA. larry.marnett@vanderbilt.edu.|Department of Chemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA. larry.marnett@vanderbilt.edu.|Department of Pharmacology, Vanderbilt Institute of Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA. larry.marnett@vanderbilt.edu.
authors.forename: Lawrence J
authors.initials: LJ
authors.lastname: Marnett
mesh_terms: D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001095:Arachidonic Acids / Q000378:metabolism; D001681:Biological Assay; D002460:Cell Line; D002853:Chromatography, Liquid; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D063388:Endocannabinoids / Q000737:chemistry / Q000378:metabolism*; D005989:Glycerides / Q000378:metabolism; D066298:In Vitro Techniques; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D010084:Oxidation-Reduction; D053284:Polyunsaturated Alkamides / Q000378:metabolism; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry
chemical_list: D001095:Arachidonic Acids; D063388:Endocannabinoids; D005989:Glycerides; D053284:Polyunsaturated Alkamides; D016718:Arachidonic Acid; C094503:glyceryl 2-arachidonate; D051546:Cyclooxygenase 2; C078814:anandamide
keywords: Cell assay; Cyclooxygenase-2; Endocannabinoids; In vitro assay; LC-MS/MS; PG-EAs; PG-Gs