Fulltext

Evaluation of an in-capillary approach for performing quantitative cytochrome P450 activity studies

Curcio, R. ; Nicoli, R. ; Rudaz, S. ; Veuthey, J.-L

In: Analytical and Bioanalytical Chemistry, 2010, vol. 398, no. 5, p. 2163-2171

Ajouter à la liste personnelle
    Titre
    • Evaluation of an in-capillary approach for performing quantitative cytochrome P450 activity studies
    Auteur
    • Curcio, R.. Laboratory of Pharmaceutical Analytical Chemistry, School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Boulevard d'Yvoy 20, 1211, Geneva 4, Switzerland
    • Nicoli, R.. Laboratory of Pharmaceutical Analytical Chemistry, School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Boulevard d'Yvoy 20, 1211, Geneva 4, Switzerland
    • Rudaz, S.. Laboratory of Pharmaceutical Analytical Chemistry, School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Boulevard d'Yvoy 20, 1211, Geneva 4, Switzerland
    • Veuthey, J.-L. Laboratory of Pharmaceutical Analytical Chemistry, School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Boulevard d'Yvoy 20, 1211, Geneva 4, Switzerland
    Type de document
    • Postprint
    Langue
    • Anglais
    Publié dans
    • Analytical and Bioanalytical Chemistry, 2010, vol. 398, no. 5, p. 2163-2171. Springer-Verlag
    Autre version électronique
    Identifiant OAI-PMH
    • oai:doc.rero.ch:316222
    Summary
    An automated in-capillary assay requiring very small quantities of reagents was developed for performing in vitro cytochrome P450 (CYP450) drug metabolism studies. The approach is based on the following: (i) hydrodynamic introduction of nanoliter volumes of substrate and enzyme solutions in the sandwich mode, within a capillary; (ii) mixing the reagents by diffusion across the interfaces between the injected solutions; (iii) collection of the capillary content at the end of the in-capillary assay; and (iv) off-line analysis of the incubation mixture by ultrahigh pressure liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). After optimizing the injection sequence of the reagents, the in-capillary approach was applied to the quantitative determination of the kinetics of drug metabolism reactions catalyzed by three CYP450 isozymes involved in human drug metabolism: CYP1A2, CYP2D6, and CYP3A4. It was demonstrated that this in-capillary method was able to provide similar kinetic parameters for CYP450 activity (e.g., Michaelis constants and turnover values) as the classical in vitro method, with a drastic reduction of reagent consumption. Injection setups used for in-capillary CYP450 assays