title: A simple validated RP-HPLC bioanalytical method for the quantitative determination of a novel valproic acid arylamide derivative in rat hepatic microsomes.
abstract: A simple and specific bioanalytical method based on reversed-phase high-performance liquid chromatography (RP-HPLC) coupled with ultraviolet detection was developed and validated for the determination of a novel valproic acid arylamide, N-(2-hydroxyphenyl)-2-propylpentanamide (HO-AAVPA) in rat hepatic microsomes (a subcellular fraction containing phase I enzymes, especially cytochrome P450). The chromatographic separation was achieved using a reversed-phase Zorbax SB-C18 column and a mobile phase of acetic acid in water (0.2% v/v) and acetonitrile (40:60 v/v) with a flow rate of 0.5 mL/min. The calibration curve was linear over the range of 882-7060 ng/mL (r(2)  = 0.9987), and the lower limit of quantification and the lower limit of determination were found to be 882 and 127.99 ng/mL, respectively. The method was validated with excellent sensitivity, and intra-day accuracy and precision varied from 93.79 to 93.12%, and from 2.12 to 4.36%, respectively. The inter-day accuracy and precision ranged from 93.29 to 97.30% and from 0.68 to 3.60%, respectively. The recovery of HO-AAVPA was measured between 91.36 and 97.98%. The assay was successfully applied to the analysis of kinetic metabolism and pharmacokinetic parameters in vitro by a substrate depletion approach.
authors.affiliation: Laboratorio de Biofísica y Biocatálisis y Laboratorio de Modelado Molecular y Diseño de Fármacos de la Escuela Superior de Medicina, Instituto Politécnico Nacional, Plan de San Luis y Díaz Mirón s/n, Casco de Santo Tomás, Distrito Federal, 11340, Mexico.
authors.forename: Arianna
authors.initials: A
authors.lastname: Silva-Trujillo
authors.forename: José
authors.initials: J
authors.lastname: Correa-Basurto
authors.forename: Aurelio
authors.initials: A
authors.lastname: Romero-Castro
authors.forename: Arnulfo
authors.initials: A
authors.lastname: Albores
authors.forename: Jessica Elena
authors.initials: JE
authors.lastname: Mendieta-Wejebe
mesh_terms: D000818:Animals; D000927:Anticonvulsants / Q000032:analysis* / Q000493:pharmacokinetics; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D008297:Male; D008862:Microsomes, Liver / Q000737:chemistry*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014635:Valproic Acid / Q000032:analysis* / Q000493:pharmacokinetics
chemical_list: D000927:Anticonvulsants; D014635:Valproic Acid
keywords: CYP; RP-HPLC; metabolic stability; microsomes; validation