High Performance Liquid Chromatography-Tandem Mass Spectrometry Method for Ex vivo Metabolic Studies of a Rhenium-Labeled Radiopharmaceutical for Liver Cancer

The radio-isotope rhenium-labeled N-[2-(triphenylmethyl)thioethyl]-3-aza-19-ethyloxycarbonyl-3-[2-(triphenylmethyl)thioethyl] octadecanoate) ligand (188Re-MN-16ET) is a novel therapeutic agent under preclinical evaluation for hepatoma. A reversed-phase high performance liquid chromatography coupled with a tandem mass spectrometric analysis method and diode array detector (DAD) involving a T type splitter was developed to characterize this pharmaceutical in rat liver tissue solution and determine its biotransformation rate. The separation was accomplished on a C18 column (chromolith silica, 4.6 mm × 100 mm) using an acetonitrile–ammonium acetate buffer gradient as the mobile phase. The detection was achieved by DAD set at 250 nm and tandem mass spectrometry using electrospray ionization in the positive ion mode. Re-MN-16ET displayed a retention time of 23.2 min and a transition ion pair corresponding to m/z 677 → 631 for multiple reaction monitoring. Its biotransformation reaction in rat liver homogenate proceeded for 90 min in a 37°C water bath. The characterization was conducted using aliquots that were extracted and concentrated from the reaction mixture for various incubation times. Re-MN-16ET exhibited a biotransformation half-life ( t1/2) of 8–9 min in liver tissue solution and was almost completely exhausted after 90 min. Two of its metabolites, consisting of the Re-labeled carboxylic acid derivative, predominately, and its corresponding demetallized disulfide ligand were found in the liver homogenate, providing a metabolism pathway for the radiopharmaceutical.