title: A simple method for simultaneous determination of N-arachidonoylethanolamine, N-oleoylethanolamine, N-palmitoylethanolamine and 2-arachidonoylglycerol in human cells.
abstract: The endocannabinoid system has been considered as a target for pharmacological intervention. Accordingly, inhibition of fatty acid amide hydrolase (FAAH), a degrading enzyme of the endocannabinoids N-arachidonoylethanolamine (anandamide; AEA) and 2-arachidonoylglycerol (2-AG) as well as of the endocannabinoid-like substances N-oleoylethanolamine (OEA) and N-palmitoylethanolamine (PEA), can cause augmented endogenous cannabinoid tone. Using liquid chromatography coupled with positive electrospray ionisation mass spectrometry, we herein describe a method to simultaneously quantify levels of AEA, OEA, PEA and 2-AG in cultured cells. The procedure was developed according to the FDA guidelines for bioanalytical methods validation. The limits of quantification (LOQs) were 0.05 pmol for AEA, 0.09 pmol for OEA, 0.10 pmol for PEA and 0.80 pmol for 2-AG when molecular ion monitoring was used. In H460 human lung carcinoma cells, basal levels of all four analytes ranged between 2 and 17 pmol mg(-1) protein with PEA showing the lowest and OEA the highest concentrations. Endocannabinoid levels observed in mesenchymal stem cells were of the same order of magnitude when compared to those in H460 human lung carcinoma cells.
authors.affiliation: Institute of Toxicology and Pharmacology, University of Rostock, Schillingallee 70, 18057, Rostock, Germany, igor.ivanov@med.uni-rostock.de.
authors.forename: Igor
authors.initials: I
authors.lastname: Ivanov
authors.forename: Philipp
authors.initials: P
authors.lastname: Borchert
authors.forename: Burkhard
authors.initials: B
authors.lastname: Hinz
mesh_terms: D000577:Amides; D001095:Arachidonic Acids / Q000032:analysis*; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid / Q000379:methods; D063388:Endocannabinoids / Q000032:analysis*; D004983:Ethanolamines / Q000032:analysis*; D005989:Glycerides / Q000032:analysis*; D006801:Humans; D057230:Limit of Detection; D008175:Lung Neoplasms / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods; D059630:Mesenchymal Stem Cells / Q000737:chemistry; D009829:Oleic Acids / Q000032:analysis*; D010169:Palmitic Acids / Q000032:analysis*; D053284:Polyunsaturated Alkamides; D015203:Reproducibility of Results
chemical_list: D000577:Amides; D001095:Arachidonic Acids; D063388:Endocannabinoids; D004983:Ethanolamines; D005989:Glycerides; D009829:Oleic Acids; D010169:Palmitic Acids; D053284:Polyunsaturated Alkamides; C033595:N-oleoylethanolamine; C005958:palmidrol; C094503:glyceryl 2-arachidonate; C078814:anandamide