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KMID : 0369819910210010033
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Jorunal of Korean Pharmaceutical Sciences
1991 Volume.21 No. 1 p.33 ~ p.41
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HPLC Assay and Renal Excretion Characteristics of Theophylline and Its Metabolites in Rat
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구효정/Kuh HJ
심창구/이민화/김신근/Shim CK/Lee MH/Kim SK
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Abstract
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A high-performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of theophylline(TP) and its metabolites, 1-methyluric acid (1-MU) and 1,3-dimethyluric acid (1,3-DMU), in rat plasma and urine. An 100μl aliquot of a plasma or urine sample was mixed with 250μl of acetonitrite and vortexed. After centrifugation, 200μl (plasma) or 20μl (urine) aliquot of the supernatant was dried by N2 stream and redissolved in 100μl (plasma) or 200μl (urine) of the mobile phase. A 20μl of the mobile phase solution was injected onto a C18 reversed-phase column. The column was maintained at 45?C by the aid of electric heating jacket. The mobile phase was a 3%(v/v) methanol solution in deionized water which contains sodium acetate (100 mM) and tetrabutyl ammonium hydroxide (4 mM). pH of the mobile phase was adjusted 4.5 by the addition of acetic acid. Detection limits for TP, 1-MU, and 1,3-DMU in plasma were 0.2, 0.1 and 0.1μ/ml, respectively and the corresponding values in urine were all 5μg/ml. Inter- and intra-day variability of the assay for all compounds in the plasma samples was less than 5.5 and 3.8%, respectively. The retention times for 1-MU, 1,3-DMU, and TP were approximately 7, 8.5 and 18 min, respectively. Sample preparation procedure used in this method was simple, rapid and reproducible. Renal clearance of TP and its metabolites in rats showed plasma concentration dependency indicating renal tubular secretion and reabsorption of them.
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KEYWORD
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HPLC, theophylline, 1-methyluric acid (1-MU), 1,3-dimethyluric acid (1,3-DMU), nonlinear renal clearance, renal tubular secretion
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