VALIDATED HPLC MS/MS ASSAY FOR CI-1011 IN RAT PLASMA AND A COMPARISONWITH AN HPLC/UV ASSAY/

A liquid chromatographic/mass spectrometric (LC/MS/MS) method to quant itate CI-1011 in rat plasma has been validated and compared to an LC/U V assay. The analyte and internal standard were isolated from the plas ma matrix by using liquid/liquid extraction with diethyl ether. The et her layer was evaporated to dryness and the residue reconstituted in a cetonitrile-water (70:30, v/v). A 2.1 x 150 mm x 5 mu m Zorbax RX-C18 column with a mobile phase of acetonitrile-ammonium acetate (pH 8.0: 5 mM)-triethylamine (70.30:0.03, v/v/v) delivered at a flow rate of 0.2 mi min(-1) was used for chromatography. Analyte and internal standard ion chromatograms were obtained by operating the mass spectrometer in the negative ion multiple reaction monitoring mode to detect the pres ence of a precursor-product ion pair for both the analyte and the inte rnal standard. Samples were introduced into the mass spectrometer usin g electrospray ionization. Retention times of CI-1011 and of the inter nal standard (IS), [C-13(6)]CI-1011, were approximately 4.2 min. No pe aks interfering with the quantitation of CI-1011 were observed through out the validation process. Mean recoveries of CI-1011 from rat plasma ranged from 98.2 to 105%. The recovery of the IS was 100%. Assay prec ision for CI-1011, based on the percent relative standard deviation of replicate quality controls, was less than or equal to 5.60% with an a ccuracy of +/- 8.80%. The lower limit of quantitation for CI-1011 was 0.500 ng ml(-1) for a 0.2-ml sample aliquot. CI-1011 is stable in rat plasma for 24 h at room temperature and for at least 34 days at -20 de grees C. This assay has been proven suitable for routine quantitation of CI-1011 in rat plasma at concentrations from 0.500 (100 pg on-colum n) to 500 ng ml(-1). The applicability of this method to determine CI- 1011 concentrations in rat plasma is reported in this manuscript. CI-1 011 concentrations, in plasma samples from cholesterol- and chow-fed r ats administered single daily oral doses of CI-1011 in a CMC/Tween sus pension, obtained using a validated LC/UV assay were compared to conce ntrations obtained using the reported LC/MS/MS assay over the concentr ation range 0.0806-12.3 mu g ml(-1). The concordance correlation coeff icient determined for this comparison was 0.9977, suggesting that the CI-1011 concentrations obtained by the two assays are in excellent agr eement. (C) 1998 Elsevier Science B.V. All rights reserved.