Determination of baclofen and α-baclofen in rat liver homogenate and human urine using solid-phase extraction, o-phthalaldehyde- tert.-butyl thiol derivatization and high-performance liquid chromatography with amperometric detection

Abstract

Methods were developed for the determination of the zwitterionic compounds baclofen and α-baclofen in complex biological samples (rat liver homogenates and human urine) in concentration ranges that would be suitable for pharmacokinetic studies of these compounds. In the procedure, the biological samples along with an internal standard were selectively concentrated using C 18 solid-phase extraction cartridges, evaporated, derivatized with o-phthalaldehyde and tert.-butyl thiol at room temperature for 2 min, then subjected to high-performance liquid chromatographic analysis. The reversed-phase (C 18) chromatographic analysis with amperometric detection (glassy carbon electrode in oxidative mode, +0.6 V vs. Ag/AgCl) was found to be useful for the measurement of both baclofen and α-baclofen from 10 ng/ml to 10 μg/ml in these complex biological samples. The primary advantage of the method was that the derivatives formed using tert.-butyl thiol were markedly more stable than the previously reported derivatives prepared using mercaptoethanol.