title: Liquid chromatography-tandem mass spectrometric assay for pravastatin and two isomeric metabolites in mouse plasma and tissue homogenates.
abstract: A bioanalytical assay for pravastatin and two isomeric metabolites, 3'α-isopravastatin and 6'-epipravastatin, was developed and validated. Mouse plasma and tissue homogenates from liver, kidney, brain and heart were pre-treated using protein precipitation with acetonitrile containing deuterated internal standards of the analytes. The extract was diluted with water and injected into the chromatographic system. This system consisted of a polar embedded octadecyl silica column using isocratic elution with formic acid in a water-acetonitrile mixture. The eluate was transferred to an electrospray interface using negative ionization and the analytes were detected and quantified with the selected reaction monitoring mode of a triple quadrupole mass spectrometer. The assay was successfully validated in a 3.4-7100ng/ml concentration range for pravastatin, 1.3-2200ng/ml for 3'α-isopravastatin and 0.5-215ng/ml for 6'-epipravastatin using only plasma for calibration. For plasma samples, subjected to full validation, within and between day precisions were 1-7% (9-18% at the LLQ level) and accuracies were between 91% and 103%. For tissue homogenates, subjected to partial validation, within and between day precisions were 2-12% (6-19% at the LLQ level) and accuracies were between 87% and 113% (81 and 113% at the LLQ level). Drug and metabolites were shown to be chemically stable under most relevant analytical conditions. Finally, the assay was successfully applied for a pilot study in mice. After intravenous administration of the drug, all isomeric compounds were found in plasma; however, in liver and kidney homogenate only the parent drug showed levels exceeding the LLQ.
authors.affiliation: Universiteit Utrecht, Faculty of Science, Department of Pharmaceutical Sciences, Section of Biomedical Analysis, Division of Drug Toxicology, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands. R.W.Sparidans@uu.nl
authors.forename: Rolf W
authors.initials: RW
authors.lastname: Sparidans
authors.forename: Dilek
authors.initials: D
authors.lastname: Iusuf
authors.forename: Alfred H
authors.initials: AH
authors.lastname: Schinkel
authors.forename: Jan H M
authors.initials: JH
authors.lastname: Schellens
authors.forename: Jos H
authors.initials: JH
authors.lastname: Beijnen
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D005260:Female; D007668:Kidney / Q000737:chemistry; D008099:Liver / Q000737:chemistry; D008297:Male; D051379:Mice; D009206:Myocardium / Q000737:chemistry; D017035:Pravastatin / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014018:Tissue Distribution
chemical_list: D017035:Pravastatin