High-performance liquid chromatographic analysis of triamterene and p-hydroxytriamterene in plasma

A rapid and sensitive high-performance liquid chromatographic assay was developed for triamterene and 6-p-hydroxytriamterene in plasma. Plasma (0.5 ml), after addition of the internal standard, was extracted with 10 ml of ether-isopropanol (95:5). After thorough mixing and separation of phases, the organic layer was evaporated to dryness under nitrogen. The residue was reconstituted with 500 microliters of mobile phase [acetonitrile-water-acetic acid (60:39.5:0.5)], and 100 microliters was injected into the chromatograph. Chromatographic separation was carried out on a C18 muBondapak column at a flow rate of 1 ml/min. Detection of compounds in the column eluent was by UV absorption at 365 nm. The retention times for 6-p-hydroxytriamterene, triamterene, and the internal standard were 7.5, 9.0, and 12.0 min, respectively. The lower limit of detection for each compound was 20 ng/ml. Recoveries of triamterene and 6-p-hydroxytriamterene were 91--99 and 828--95%, respectively, over a 40--240-ng/ml range.