title: In-line capillary electrophoretic evaluation of the enantioselective metabolism of verapamil by cytochrome P3A4.
abstract: In this paper a methodology for the in-line evaluation of enantioselective metabolism by capillary electrophoresis has been developed and applied to the study of verapamil metabolism by cytochrome P3A4. The developed methodology comprises an in-capillary reaction step carried out by electrophoretically mediated microanalysis and a separation step in which highly sulfated β-cyclodextrin with partial filling technique has been employed as chiral selector for verapamil and norverapamil enantiomers resolution, joining the advantages of both methodologies in a unique assay. Kinetic parameters of the enzymatic reaction (Km and Vmax) have been evaluated for both verapamil enantiomers by non-linear fitting of experimental data obtained under intermediate precision conditions to Michaelis-Menten equation. Km and Vmax estimated values were 51±9 μM and 22±2 pmol min(-1) (pmol CYP)(-1) for S-VER and 47±9 μM and 21±2 pmol min(-1) (pmol CYP)(-1) for R-VER. Consequently, slight enantioselectivity was found for the CYP3A4 metabolism of verapamil. However, since confidence intervals of estimates overlap, we cannot assure a significant enantioselectivity. Intrinsic clearance values were also estimated from Km and Vmax for both enantiomers.
authors.affiliation: Departamento de Química Analítica, Facultad de Farmacia, Universidad de Valencia, Burjassot, Valencia, Spain.
authors.forename: L
authors.initials: L
authors.lastname: Asensi-Bernardi
authors.forename: Y
authors.initials: Y
authors.lastname: Martín-Biosca
authors.forename: L
authors.initials: L
authors.lastname: Escuder-Gilabert
authors.forename: S
authors.initials: S
authors.lastname: Sagrado
authors.forename: M J
authors.initials: MJ
authors.lastname: Medina-Hernández
mesh_terms: D019075:Electrophoresis, Capillary / Q000379:methods*; D015394:Molecular Structure; D012984:Software; D013237:Stereoisomerism; D014700:Verapamil / Q000737:chemistry*
chemical_list: D014700:Verapamil