title: Identification of phase I and phase II metabolites of benfluron and dimefluron in rat urine using high-performance liquid chromatography/tandem mass spectrometry.
abstract: Biotransformation products of two potential antineoplastic agents, benfluron and dimefluron, are characterized using our integrated approach based on the combination of high-performance liquid chromatography (HPLC) separation of phase I and phase II metabolites followed by photodiode-array UV detection and electrospray ionization tandem mass spectrometry (MS/MS). High mass accuracy measurement allows confirmation of an elemental composition and metabolic reactions according to exact mass defects. The combination of different HPLC/MS/MS scans, such as reconstructed ion current chromatograms, constant neutral loss chromatograms or exact mass filtration, helps the unambiguous detection of low abundance metabolites. The arene oxidation, N-oxidation, N-demethylation, O-demethylation, carbonyl reduction, glucuronidation and sulfation are typical mechanisms of the metabolite formation. The interpretation of their tandem mass spectra enables the distinction of demethylation position (N- vs. O-) as well as to differentiate N-oxidation from arene oxidation for both phase I and phase II metabolites. Two metabolic pathways are rather unusual for rat samples, i.e., glucosylation and double glucuronidation. The formation of metabolites that lead to a significant change in the chromophoric system of studied compounds, such as the reduction of carbonyl group in 7H-benzo[c]fluorene-7-one chromophore, is reflected in their UV spectra, which provides valuable complementary information to MS/MS data.
authors.affiliation: Department of Analytical Chemistry, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 53210 Pardubice, Czech Republic.
authors.forename: Robert
authors.initials: R
authors.lastname: Jirásko
authors.forename: Michal
authors.initials: M
authors.lastname: Holčapek
authors.forename: Milan
authors.initials: M
authors.lastname: Nobilis
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism / Q000652:urine; D001669:Biochemical Phenomena; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005449:Fluorenes / Q000378:metabolism / Q000652:urine*; D005960:Glucosides / Q000378:metabolism / Q000652:urine; D020719:Glucuronides / Q000378:metabolism / Q000652:urine; D008297:Male; D053858:Metabolic Networks and Pathways; D051381:Rats; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000970:Antineoplastic Agents; D005449:Fluorenes; D005960:Glucosides; D020719:Glucuronides; C507439:dimefluron; C034234:VUFB 13468