Identification of phase I and phase II metabolites of benfluron and dimefluron in rat urine using high-performance liquid chromatography/tandem mass spectrometry

• Published in: Rapid communications in mass spectrometry 2011-08, Vol.25 (15), p.2153-2162
• Main Authors: Jirásko, Robert, Holčapek, Michal, Nobilis, Milan
• Format: Article
• Language: English
• Subjects: Animals; Antineoplastic Agents - metabolism; Antineoplastic Agents - urine; Biochemical Phenomena; Chromatography, High Pressure Liquid - methods; Fluorenes - metabolism; Fluorenes - urine; Glucosides - metabolism; Glucosides - urine; Glucuronides - metabolism; Glucuronides - urine; Male; Metabolic Networks and Pathways; Rats; Tandem Mass Spectrometry - methods
• ISSN: 0951-4198; 1097-0231
• DOI: 10.1002/rcm.5097

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. Copyright © 2011 John Wiley & Sons, Ltd.