title: Improved procedure for the the determination of rofecoxib in human plasma involving 96-well solid-phase extraction and fluorescence detection.
abstract: An improved assay for the determination of rofecoxib in human plasma samples is described. The analyte and an internal standard were extracted from the plasma matrix using solid-phase extraction in the 96-well format with an Empore C8-SD extraction plate. The analytes are chromatographed on a Waters Symmetry C18 analytical column (3.5 microm, 50x4.6 mm) with a mobile phase consisting of acetonitrile-water (35:65, v/v). Analyte detection was via fluorescence following post-column photochemical derivatization. Eight point calibration curves over the concentration range of 0.5-80 ng/ml yielded a linear response when a 1/y weighted linear regression model was employed. Based on the replicate analyses (n=5) of spiked standards, the within-day assay precision was better than 8% RSD at all points on the calibration curve, within-day accuracy was within 6% of nominal at all standard concentrations. The between-run precision and accuracy of the assay, as calculated from the results of the analysis of quality control samples, was better than 7% RSD and within 5% of nominal. Assay throughput was improved by a factor of three as compared to previously described methods. The method was partially automated using a combination of a Packard Multi-Probe liquid handling system and a TomTec Quadra 96 workstation.
authors: Department of Drug Metabolism, Merck Research Laboratories, West Point, PA 19486, USA.
authors: C Z
authors: CZ
authors: Matthews
authors: E J
authors: EJ
authors: Woolf
authors: B K
authors: BK
authors: Matuszewski
mesh_terms: D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D016861:Cyclooxygenase Inhibitors / Q000097:blood*; D006801:Humans; D007783:Lactones / Q000097:blood*; D012015:Reference Standards; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013450:Sulfones
chemical_list: D016861:Cyclooxygenase Inhibitors; D007783:Lactones; D013450:Sulfones; C116926:rofecoxib