Quantification of cortisol and 6 beta-hydroxycortisol in human urine by LC-MS/MS, and gender-specific evaluation of the metabolic ratio as biomarker of CYP3A activity.
Drug-drug and food-drug interactions are often due to an inhibition or induction of drug-metabolizing cytochrome P450 (CYP) enzymes and may result in non-response or adverse reactions. Hence, phenotypic biomarkers of CYP activity appear as useful tools for individualized pharmacotherapy. The metabolic ratio (MR) of the concentration of 6beta-hydroxycortisol (6beta-OHC) to cortisol (MR 6beta-OHC/cortisol) in human urine had been proposed as an endogenous marker for CYP3A activity. Here, we report on the improvement of published LC-MS/MS methods for the simultaneous quantification of cortisol and 6beta-OHC, using on-line sample cleanup by column switching and isotope-labeled analogues as internal standards. [(2)H(2)]6beta-OHC was prepared by incubation of human recombinant CYP3A4 with commercially available [(2)H(2)]cortisol. Analytical sensitivity could be increased about 10-fold. The first morning urine of 69 female and 27 male healthy volunteers was analyzed for cortisol and 6beta-OHC. Concentrations ranged from 1.0 to 142 and 24 to 670ng/mL, respectively. Individual MR 6beta-OHC/cortisol varied more than 20-fold and we were able to show for the first time for a Caucasian population significantly higher MR values in females as compared to males. This non-invasive biomarker for CYP3A activity lends itself for the study of genetic differences as well as enzyme induction or inhibition in the clinical setting without the need of using a probe drug.
Lutz
Ursula
U
Department of Toxicology, University of Würzburg, 9 Versbacher St., D-97078 Würzburg, Germany.
Bittner
Nataly
N
Ufer
Mike
M
Lutz
Werner K
WK
D000328:Adult; D015415:Biomarkers / Q000378:metabolism / Q000652:urine*; D002853:Chromatography, Liquid / Q000379:methods*; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D005260:Female; D006801:Humans; D006854:Hydrocortisone / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000652:urine*; D008297:Male; D008875:Middle Aged; D010641:Phenotype; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012737:Sex Factors; D018709:Statistics, Nonparametric; D053719:Tandem Mass Spectrometry / Q000379:methods*
D015415:Biomarkers; C001380:6 beta-hydroxycortisol; D003577:Cytochrome P-450 Enzyme System; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; D006854:Hydrocortisone