title: Rapid determination of five probe drugs and their metabolites in human plasma and urine by liquid chromatography/tandem mass spectrometry: application to cytochrome P450 phenotyping studies.
abstract: A liquid chromatography/mass spectrometry method, for rapid determination of five cytochrome P450 (CYP) probe drugs and their relevant metabolites in human plasma and urine, is described. The five specific probe substrates/metabolites, caffeine/paraxanthine (CYP1A2), tolbutamide/4-hydroxytolbutamide/carboxytolbutamide (CYP2C9), omeprazole/5-hydroxyomeprazole (CYP2C19), debrisoquine/5-hydroxydebrisoquine (CYP2D6) and midazolam/1'-hydroxymidazolam (CYP3A), together with the internal standards (phenacetin and paracetamol), in plasma and urine, were extracted using solid-phase extraction. The chromatography was performed using a C18 column with an isocratic mobile phase consisting of acetonitrile and 0.1% formic acid in water (70:30). The triple-quadrupole mass spectrometer was operated in both positive and negative modes, and multiple reaction monitoring was used for quantification. The method was validated over the concentration ranges 0.05-5 microg/mL for caffeine and paraxanthine, 0.02-2 microg/mL for tolbutamide, 0.1-20 microg/mL for 4-hydroxytolbutamide, carboxytolbutamide, debrisoquine and 5-hydroxydebrisoquine, 5-2500 ng/mL for omeprazole and 5-hydroxyomeprazole, and 1-100 ng/mL for midazolam and 1'-hydroxymidazolam. The intra- and inter-day precision were 0.3-13.7% and 1.9-14.3%, respectively, and the accuracy ranged from 93.5-107.2%. The lower limit of quantification varied between 1 and 100 ng/mL. The present method provides a robust, fast and sensitive analytical tool for the five-probe drug cocktail, and has been successfully applied to a clinical phenotyping study in 16 subjects.
authors.affiliation: School of Pharmacy, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong. qpyin@cuhk.edu.uk
authors.forename: Ophelia Q P
authors.initials: OQ
authors.lastname: Yin
authors.forename: Sherry S L
authors.initials: SS
authors.lastname: Lam
authors.forename: Cindy M Y
authors.initials: CM
authors.lastname: Lo
authors.forename: Moses S S
authors.initials: MS
authors.lastname: Chow
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003577:Cytochrome P-450 Enzyme System / Q000145:classification / Q000235:genetics / Q000378:metabolism; D004338:Drug Combinations; D006801:Humans; D004364:Pharmaceutical Preparations / Q000097:blood* / Q000652:urine*; D010641:Phenotype; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D004338:Drug Combinations; D004364:Pharmaceutical Preparations; D003577:Cytochrome P-450 Enzyme System