High-performance liquid chromatographic determination of tracazolate and its major metabolite in plasma

A high-performance liquid chromatographic (HPLC) method is reported that provides for either separate or simultaneous plasma determination of tracazolate and its major metabolite, the free carboxylic acid. Tracazolate is extracted from plasma with hexane and is quantitated by reverse-phase HPLC using an internal standard. The metabolite is extracted in an additional step, methylated, and either quantitated using the same HPLC conditions in a separate determination or combined with the hexane extract for the simultaneous HPLC determination of drug and metabolite. The plasma concentrations of metabolite were as much as 60 times greater than free drug in some animal species, therefore requiring the use of separate determinations for the two compounds. The analyses have limits of reproducible quantitation of 20 ng/mL for tracazolate and 50 ng/mL for the metabolite in plasma. The simultaneous analysis for tracazolate and the metabolite was used to evaluate plasma levels of both compounds versus time and dose. Data generated in conjunction with a toxicology study are reported to demonstrate the applicability of the procedure.