Utility of liquid chromatography/fast atom bombardment mass spectrometry and liquid chromatography/thermospray mass spectrometry for structure identification of metabolites of a fluorinated herbicide

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Abstract
This study demonstrates a useful application of on‐line microbore high‐performance liquid chromatography (HPLC) fast atom bombardment (FAB) and thermospray (TSP) mass Spectrometry techniques for identification of metabolites from the in vitro metabolism of an experimental Monsanto herbicide: 2‐chloro‐N‐(ethoxymethyl‐N‐[2‐methyl‐6‐(trifluoromethyl) phenyl]acetamide), ‘chloroacetanilide’. The microbore HPLC FAB technique on a high‐resolution sector mass spectrometer accelerated identification of polar metabolites from the in vitro metabolism study of the herbicide. It provided good chromatographic resolution and excellent FAB sensitivity with strong protonated molecular ions. Scanning high‐resolution LC/FAB mass spectrometry also provided molecular formulae for structural elucidation of unknown metabolites. Sample purification and concentration were minimized. Identification of less polar metabolites was carried out using LC/TSP mass spectrometry with a quadrupole mass spectrometer. LC/TSP mass spectrometry provided useful structural information for both polar and less polar metabolites because their spectra showed more fragmentation than FAB spectra. Glutathione conjugation was the major reaction observed during in vitro incubation of the herbicide. Oxidation of the chloroacetanilide by rat liver enzymes was also a significant metabolic reaction. Seven metabolites were identified, of which four were glutathione conjugates.