title: LC-MS/MS analysis of epoxyalcohols and epoxides of arachidonic acid and their oxygenation by recombinant CYP4F8 and CYP4F22.
abstract: CYP4F22 and CYP4F8 are expressed in epidermis, and mutations of CYP4F22 are associated with lamellar ichthyosis. Epoxyalcohols (HEETs) and epoxides (EETs) of 20:4n-6 appear to be important for the water permeability barrier of skin. Our aim was to study the MS/MS spectra and fragmentation of these compounds and to determine whether they were oxidized by CYP4F22 or CYP4F8 expressed in yeast. HEETs were prepared from 15-hydroperoxyeicosatetraenoic acid (15-HPETE), 12-HPETE, and their [(2)H(8)]labeled isotopomers, and separated by normal phase-HPLC with MS/MS analysis. CYP4F22 oxygenated 20:4n-6 at C-18, whereas metabolites of HEETs could not be identified. CYP4F8 formed omega3 hydroxy metabolites of HEETs derived from 12R-HPETE with 11,12-epoxy-10-hydroxy configuration, but not HEETs derived from 15S-HPETE. 8,9-EET and 11,12-EET were also subject to omega3 hydroxylation by CYP4F8. We conclude that CYP4F8 and CYP4F22 oxidize 20:4n-6 and that CYP4F8 selectively oxidizes 8,9-EET, 11,12-EET, and 10,11R,12R-HEET at the omega3 position.
authors.affiliation: Division of Biochemical Pharmacology, Department of Pharmaceutical Biosciences, Biomedicum, Uppsala University, Uppsala, Sweden.
authors.forename: T
authors.initials: T
authors.lastname: Nilsson
authors.forename: I V
authors.initials: IV
authors.lastname: Ivanov
authors.forename: E H
authors.initials: EH
authors.lastname: Oliw
mesh_terms: D000438:Alcohols / Q000032:analysis* / Q000378:metabolism; D016718:Arachidonic Acid / Q000737:chemistry* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003315:Cornea / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism; D004852:Epoxy Compounds / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D010100:Oxygen / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000438:Alcohols; D004852:Epoxy Compounds; D011994:Recombinant Proteins; D016718:Arachidonic Acid; D003577:Cytochrome P-450 Enzyme System; C559989:CYP4F22 protein, human; D010100:Oxygen