title: Validated HPLC/MS/MS assay for CI-1011 in rat plasma and a comparison with an HPLC/UV assay.
abstract: A liquid chromatographic/mass spectrometric (LC/MS/MS) method to quantitate CI-1011 in rat plasma has been validated and compared to an LC/UV assay. The analyte and internal standard were isolated from the plasma matrix by using liquid/liquid extraction with diethyl ether. The ether layer was evaporated to dryness and the residue reconstituted in acetonitrile-water (70:30, v/v). A 2.1 x 150 mm x 5 microns 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 ml 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 presence of a precursor-product ion pair for both the analyte and the internal standard. Samples were introduced into the mass spectrometer using electrospray ionization. Retention times of CI-1011 and of the internal standard (IS), [13C6]CI-1011, were approximately 4.2 min. No peaks interfering with the quantitation of CI-1011 were observed throughout 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 precision for CI-1011, based on the percent relative standard deviation of replicate quality controls, was less than or equal to 5.60% with an accuracy 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 degrees C. This assay has been proven suitable for routine quantitation of CI-1011 in rat plasma at concentrations from 0.500 (100 pg on-column) to 500 ng ml-1. The applicability of this method to determine CI-1011 concentrations in rat plasma is reported in this manuscript. CI-1011 concentrations, in plasma samples from cholesterol- and chow-fed rats administered single daily oral doses of CI-1011 in a CMC/Tween suspension, obtained using a validated LC/UV assay were compared to concentrations obtained using the reported LC/MS/MS assay over the concentration range 0.0806-12.3 micrograms ml-1. The concordance correlation coefficient determined for this comparison was 0.9977, suggesting that the CI-1011 concentrations obtained by the two assays are in excellent agreement.
authors.affiliation: Parke-Davis Pharmaceutical Research Division, Warner-Lambert Company, Ann Arbor, MI 48105, USA. BullenW@aa.wl.com
authors.forename: W W
authors.initials: WW
authors.lastname: Bullen
authors.forename: C D
authors.initials: CD
authors.lastname: Lathia
authors.forename: R B
authors.initials: RB
authors.lastname: Abel
authors.forename: R N
authors.initials: RN
authors.lastname: Hayes
mesh_terms: D000081:Acetamides; D000085:Acetates; D000284:Administration, Oral; D000818:Animals; D000924:Anticholesteremic Agents / Q000008:administration & dosage / Q000097:blood*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D008297:Male; D013058:Mass Spectrometry; D051381:Rats; D015203:Reproducibility of Results; D013449:Sulfonamides; D013451:Sulfonic Acids / Q000008:administration & dosage / Q000097:blood*
chemical_list: D000081:Acetamides; D000085:Acetates; D000924:Anticholesteremic Agents; D013449:Sulfonamides; D013451:Sulfonic Acids; C423185:avasimibe