Separation and quantitation of hydralazine metabolites by high-performance liquid chromatography

A selective high-performance liquid chromatographic assay for the separation and quantitation of the proposed hepatic microsomal metabolites of hydralazine (HP), phthalazine, phthalazinone, s-triazolo[3,4-a]phthalazine, 3-methyl-s-triazolo[3,4-a]phthalazine and 3-hydroxymethyl-s-triazolo [3,4-a]phthalazine, is described. An extraction technique was developed for the removal of HP from hepatic microsomal samples in order to minimize the measurement of products resulting from the chemical degradation of HP. The effects of pH and composition of the mobile phase on the retention times and resolution of the five compounds were examined. The methods presented are accurate and reliable, permitting the baseline separation of five HP metabolites with reasonable analysis time and sensitivity.