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A high-performance liquid chromatographic method with electrochemical detection is described for the simultaneous determination of the loop diuretic 1-isopropil-3-[4-(3-methylphenylamino)-3-pyridinesulphonyl]urea (torasemide) and its major metabolite M5 in human urine. The assay is simple, fast, and easy. It requires a sample cleanup consisting of a solid-phase extraction under acidic conditions followed by chromatographic separation with a C18 microBondapack column. The use of a water-acetonitrile mobile phase (80:20, v/v, pH 3) ensures total separation from urine-interfering substances, and both compounds can be quantitated amperometrically at a glassy carbon electrode set to +1300 mV versus Ag-AgCl. The method demonstrates linearity for both the parent drug and the metabolite over a wide concentration range (up to 7 microg/mL) and reproducibility with relative standard deviation lower than 2% in intraday and 5% in interday assays. The mean extraction recoveries are 78% for M5 and 60% for torasemide, and the limits of quantitation are 1 and 8 ng/mL, respectively. The method developed is applied to the analysis of healthy volunteers' urine samples collected at different time intervals after the oral ingestion of a single dose of 10 mg torasemide, and the results obtained are in agreement with the pharmacokinetic profile of torasemide.

journal of chromatographic science

Simultaneous Determination of Torasemide and Its Major Metabolite M5 in Human Urine by High-Performance Liquid Chromatography-Electrochemical Detection