title: Determination of rofecoxib (MK-0966), a cyclooxygenase-2 inhibitor, in human plasma by high-performance liquid chromatography with tandem mass spectrometric detection.
abstract: A method for the determination of 4-(4-methanesulfonylphenyl)-3-phenyl-5H-furan-2-one (Rofecoxib, Vioxx, MK-0966, I) a cyclooxygenase-2 inhibitor, in human plasma has been developed. The method is based on high-performance liquid chromatography (HPLC) with atmospheric pressure chemical ionization tandem mass spectrometric (APCI-MS-MS) detection in negative ionization mode using a heated nebulizer interface. Drug and internal standard (II) were isolated from basified plasma using liquid-liquid extraction. The organic extracts were dried, reconstituted in mobile phase and injected into the HPLC-MS-MS system. Compounds I and II were chromatographed on a narrow bore (100 mm x 3.0 mm) C18 analytical column, with mobile phase consisting of acetonitrile:water (1:1, v/v) at a flow-rate of 0.4 ml/min. The MS-MS detection was performed on a PE-Sciex API III Plus tandem mass spectrometer operated in selected reaction monitoring mode. The parent-->product ion combinations of m/z 313-->257 and 327-->271 were used to quantify I and II, respectively, after chromatographic separation of the analytes. The assay was validated in the concentration range of 0.1 to 100 ng/ml of plasma. The precision of the assay (expressed as coefficient of variation) was less than 10% at all concentrations within the standard curve range, with adequate assay accuracy. The effect of HPLC mobile phase components on the ionization efficiency and sensitivity of detection in the positive and negative ionization modes, and the detailed description of all necessary steps involved in the assay for I in plasma are presented.
authors.affiliation: Merck Research Laboratories, West Point, PA 19486, USA.
authors.forename: C M
authors.initials: CM
authors.lastname: Chavez-Eng
authors.forename: M L
authors.initials: ML
authors.lastname: Constanzer
authors.forename: B K
authors.initials: BK
authors.lastname: Matuszewski
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D016861:Cyclooxygenase Inhibitors / Q000097:blood*; D006801:Humans; D007783:Lactones / Q000097:blood*; D013058:Mass Spectrometry / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013450:Sulfones
chemical_list: D016861:Cyclooxygenase Inhibitors; D007783:Lactones; D013450:Sulfones; C116926:rofecoxib