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In vivo  rat kidney tissue metabolites of an anticancer drug, cisplatin (cis-diamminedichloroplatinum [II]) (CP) which is used for the treatment of testicular, ovarian, bladder, cervical, esophageal, small cell lung, head and neck cancers, have been identified and characterized by using liquid chromatography positive ion electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) in combination with on line hydrogen/deuterium exchange (HDX) experiments. To identify  in vivo  metabolites, kidney tissues were collected after intravenous administration of CP to adult male Sprague-Dawley rats (n = 3 per group). The tissue samples were homogenized and extracted using newly optimized metabolite extraction procedure which involves liquid extraction with phosphate buffer containing ethyl acetate and protein precipitation with mixed solvents of methanol-water-chloroform followed by solid-phase clean-up procedure on Oasis HLB 3cc cartridges and then subjected to LC/ESI-HRMS analysis. A total of thirty one unknown  in vivo  metabolites have been identified and the structures of metabolites were elucidated using LC-MS/MS experiments combined with accurate mass measurements. Online HDX experiments have been used to further support the structural characterization of metabolites. The results showed that CP undergoes a series of ligand exchange biotransformation reactions with water and other nucleophiles like thio groups of methionine, cysteine, acetylcysteine, glutathione and thioether. This is the first research approach focused on the structure elucidation of biotransformation products of CP in rats, and the identification of metabolites provides essential information for further pharmacological and clinical studies of CP, and may also be useful to develop various effective new anticancer agents.

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Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC/ESI-MS/MS) Study for the Identification and Characterization of In Vivo Metabolites of Cisplatin in Rat Kidney Cancer Tissues: Online Hydrogen/Deuterium (H/D) Exchange Study