Biopharmaceutical studies of thiazide diuretics. II. High-performance liquid chromatographic method for determination of hydrochlorothiazide in plasma, urine, blood cells and bile.

Abstract

A rapid, specific and sensitive high-performance liquid chromatographic method for the determination of hydrochlorothiazide in small volumes of plasma, urine, blood cell and bile was developed. The method has been applied in a drug level study in patients given a single oral dose of 100 mg of hydrochlorothiazide. The drug was extracted into ethyl acetate from biological fluid, and then the endogenous substances were eliminated with ion exchange cellulose and silica gel. A methanol solution of the drug extract, containing hydroflumethiazide as an internal standard, was chromatographed on a silica gel column with a mobile phase containing n-hexane and ethanol. Eluted components were detected by ultraviolet absorption measurement at 270 nm. Based on 0.3ml of plasma, the limit of detection of the method was 5 ng/ml, and the detector response was linear in the range of 20-1600-ng/ml with an overall recovery of 96.8±1.5%. Similarly, detector responses were linear over the concentration ranges of 5 to 200 μg/ml for urine, 20 to 400 ng/ml for hemolysate, and 20 to 600 ng/ml for bile. No interference was observed in this assay from the following drugs which may be administered concurrently with hydrochlorothiazide therapy : prednisolone, spironolactone, deslanoside, nifedipine, penicillins and cephalosporins.