Determination of theophylline and its metabolites in human urine by high-performance liquid chromatography

초록

High-performance liquid chromatographic method with UV detection was developed for the determination of theophylline and its metabolites in human urine using β-hydroxyethyl theophylline (β-HET) as an internal standard. For extraction of urine sample, quality control sample and xanthine-free blank urine were mixed with decylamine (ion-paring reagent) and β-HET. After saturation with ammonium sulfate, the mixture was then extracted with organic solvent at pH values of 4.0-4.5. All separations were performed with ion-pair chromatography using decylamine as an ion-pairing reagent and 3 mM sodium acetate buffered mobile phase (pH 4.0) containing 1% (v/v) acetonitrile and 0.75 mM decylamine. The detection limits of theophylline, 1,3-DMU, 1-MU, 3-MX and 1-MX in human urine were 0.17, 0.17, 0.39, 0.19 and 0.19 μg/ml, based on a signal-to-noise ratios of 3.0. The mean intraday coefficients of variation (C.V.s) of each compound on nine replicates were lower than 2.0%, while mean interday C.V.s on three days were lower than 1.6%. All separations were finished within 40 minutes.

키워드