title: Fully automated electrophoretically mediated microanalysis for CYP1A1 activity monitoring optimized by multivariate approach.
abstract: In this study, a fully automated incapillary system was developed to monitor the activity of CYP1A1 (Cytochrome P450, family 1, subfamily A, polypeptide 1) in physiological conditions. Ethoxycoumarin, the selected substrate, undergoes an inline bioreaction in the presence of CYP1A1 supersomes and Nicotinamide adenine dinucleotide phosphate reduced as cofactor, giving rise to hydroxycoumarin, the product that was assayed. The optimization of the experimental conditions was supported by the application of a design of experiment, providing a better understanding of electrophoretic mixing parameters that influence the metabolic reactions. The results obtained in optimal conditions were compared not only to those achieved after offline metabolization but also with liver microsomes. Finally, inhibition studies were conducted showing an important decrease of hydroxycoumarin formation using apigenin as CYP1A1 potent inhibitor. This study demonstrates the usefulness of our inline system for the fully automated in vitro metabolism studies and the screening of new CYP1A1 inhibitors.
authors.affiliation: Laboratory for the Analysis of Medicines (LAM), Department of Pharmacy, CIRM, University of Liège, Liège, Belgium.
authors.forename: Elena
authors.initials: E
authors.lastname: Farcaş
authors.affiliation: Laboratory for the Analysis of Medicines (LAM), Department of Pharmacy, CIRM, University of Liège, Liège, Belgium.
authors.forename: Anne-Catherine
authors.initials: AC
authors.lastname: Servais
authors.affiliation: Laboratory for the Analysis of Medicines (LAM), Department of Pharmacy, CIRM, University of Liège, Liège, Belgium.
authors.forename: Caroline
authors.initials: C
authors.lastname: Lamalle
authors.affiliation: Advanced Technology Corporation (ATC), University Hospital Centre of Liège, Liège, Belgium.
authors.forename: Patrice
authors.initials: P
authors.lastname: Chiap
authors.affiliation: Namur Medicine & Drug Innovation Center (NAMEDIC-NARILIS), University of Namur, Namur, Belgium.
authors.forename: Lionel
authors.initials: L
authors.lastname: Pochet
authors.affiliation: Laboratory for the Analysis of Medicines (LAM), Department of Pharmacy, CIRM, University of Liège, Liège, Belgium.
authors.forename: Marianne
authors.initials: M
authors.lastname: Fillet
mesh_terms: D000818:Animals; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D019363:Cytochrome P-450 CYP1A1 / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006801:Humans; D008862:Microsomes, Liver / Q000378:metabolism; D051381:Rats
chemical_list: D019363:Cytochrome P-450 CYP1A1
keywords: CYP1A1 activity monitoring; Electrophoretically mediated microanalysis; Ethoxycoumarin; Metabolization inhibition