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  • Title: A stereospecific high-performance liquid chromatographic assay for the determination of ketoconazole enantiomers in rat plasma.
    Author: Hamdy DA, Brocks DR.
    Journal: Biomed Chromatogr; 2008 May; 22(5):542-7. PubMed ID: 18205135.
    Abstract:
    A stereospecific high-performance liquid chromatographic assay was developed for the quantitation of ketoconazole enantiomers (KTZ) in rat plasma. After protein precipitation of 100 microL plasma using acetonitrile, a wash step was performed using hexane. The supernatant was removed and KTZ enantiomers and amiodarone, the internal standard, were extracted using liquid-liquid extraction with tert-butyl methyl ether. After transfer and evaporation of the organic layer, the residue was reconstituted in mobile phase and injected into the HPLC through a chiral column. The mobile phase consisted of hexane:ethanol:2-propanol with diethyl amine, pumped at 1.5 mL/min. All components eluted within 18 min. KTZ enantiomers were baseline resolved and peaks were symmetrical in appearance with no interferences. Calibration curves were linear over the range 62.5-5000 ng/mL of enantiomer. The intraday and interday CV% assessments were <or=19 and <13%, respectively, and mean error was <4% for both enantiomers. The lower limit of quantitation was 62.5 ng/mL for each enantiomer based on 100 microL rat plasma. In rats, plasma concentrations of (+)-KTZ were higher than those of antipode after single oral doses. The assay was shown to be sensitive and appropriate for use in pharmacokinetics study of KTZ in rat.
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