High Pressure Liquid Chromatographic Determination of Hydrochlorothiazide (HCT) in Pharmaceutical Preparations and Human Serum After Solid Phase Extraction

Abstract

A rapid, accurate, and sensitive method has been developed for the quantitative determination of hydrochlorothiazide (HCT), an antihypertensive-diuretic agent. A Nucleosil C₁₈ 100×4.6 mm, 5 μm analytical column was used with a mixture of CH₃CN-1% acetic acid, at a volume ratio 20:80. Detection was performed with a variable wavelength UV-visible detector at 270 nm, resulting in detection limits of 0.2 ng per 20 μL injection.

For the quantitative determination, hydroflumethazide (HFM) was used as internal standard at a concentration of 4.08 ng/μL. A rectilinear relationship was observed up to 20 ng/μL. Analysis time was less than 6 min.

The statistical evaluation of the method was examined, performing intra-day (n=8) and inter-day calibration (n=8) and was found to be satisfactory with high accuracy and precision results.

The method was applied to the direct determination of HCT in pharmaceutical preparations (tablets) and biological fluids (human serum). Solid phase extraction was used for sample clean-up and analyte retention using Bakerbond C₁₈ cartridges. Recovery of HCT in spiked samples was 100.3 ± 0.92% over the range of 0.5–5 ng/μL. No interferences were observed in the assay from other drugs which might be concurrently present during hydrochlorothiazide therapy, such as captopril. Endogenous compounds of human serum did not interfere.