LC-MS/MS analysis of epoxyalcohols and epoxides of arachidonic acid and their oxygenation by recombinant CYP4F8 and CYP4F22

• Cytochrome P450 4F22 (CYP4F22) and CYP4F8 are expressed in epidermis, and mutations of CYP4F22 are associated with lamellar ichthyosis. Epoxyalcohols (HEETs) and epoxides (EETs) derived from 20:4n-6 appear to be important for the water permeability barrier of normal skin. Our aim was to systematically 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 [2H8]labeled isotopomers, and separated by normal phase-HPLC with on-line MS/MS analysis. CYP4F22 oxygenated 20:4n-6 at C-18, whereas metabolites of HEETs could not be identified. CYP4F8 formed w3-hydroxy metabolites of HEETs derived from 12R-HPETE with 11,12-epoxy-10-hydroxy configuration, but not its 8-hydroxy isomer, or HEETs from 15S-HPETE. 8,9-EET and 11,12-EET were also subject to ω3 hydroxylation by CYP4F8, whereas 14,15-EET was not a substrate. 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 ω3 position.

Subject headings

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Farmakologi och toxikologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Pharmacology and Toxicology (hsv//eng)

Keyword

Epoxyeicosatrienoic acids

electrospray ionization

hepoxilins

ichthyosis

Pharmaceutical pharmacology

Farmaceutisk farmakologi

Biochemical Pharmacology

biokemisk farmakologi

Publication and Content Type

ref (subject category)

ovr (subject category)