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

Authors

  • Ophelia Q. P. Yin,

    Corresponding author
    1. School of Pharmacy, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
    2. Drug Development Centre, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
    • School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong.
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  • Sherry S. L. Lam,

    1. School of Pharmacy, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
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  • Cindy M. Y. Lo,

    1. School of Pharmacy, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
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  • Moses S. S. Chow

    1. School of Pharmacy, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
    2. Drug Development Centre, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong
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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 μg/mL for caffeine and paraxanthine, 0.02–2 μg/mL for tolbutamide, 0.1–20 μg/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. Copyright © 2004 John Wiley & Sons, Ltd.

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