Metabolite analysis, isolation and purity assessment using various liquid chromatographic techniques combined with radioactivity detection

During the discovery of metabolic routes of a drug candidate, radioactively labeled substances are administered. This study reports the multidimensional application of overpressured layer chromatography (OPLC) and high-performance liquid chromatography (HPLC) coupled with online or off-line nondestructive radioactivity detection methods in metabolism studies. Among these methods, digital autoradiography and flow-cell radioactivity detectors (RD) using solid scintillators are used. In this study, the hyphenation of OPLC with RD is reported. The application of the OPLC-RD technique is demonstrated on a metabolism study as well as the multidimensional chromatographic selectivity using normal-phase OPLC for the separation in the first dimension, followed by reversed-phase HPLC-RD, which provides additional selectivity to the separation. Information regarding the identity of radiolabeled metabolites and data obtained from spectroscopic methods could be advantageously used during structure elucidation.