19 F-NMR and directly coupled 19 F/ 1 H-HPLC-NMR spectroscopic investigations of the metabolism of the model ecotoxin 3-trifluoromethylaniline in the earthworm species Eisenia veneta

1. The metabolic fate of the model ecotoxin 3-trifluoromethylaniline (3-TFMA) in earthworm was studied by 19 F- and directly coupled 19 F/ 1 H-HPLC-NMR spectroscopy. Earthworms of Eisenia veneta spp. were subjected to the ecotoxin during a filter papercontact toxicity test at exposure levels of 1000, 100, 10, 1 and 0.1 µg cm -2. A metabolic profile was obtained previously by 19 F-NMR spectroscopy and metabolites were observed at all the exposure levels. 2. Identification of metabolites in individual worm extracts at the (lethal) exposure levels of 1000 and 100 µg cm -2 could be achieved on-line without sample preparation by 19 F/ 1 H-HPLC-NMR spectroscopy. 19 F-HPLC-NMR spectroscopy was used in the continuous-flow mode, which enabled the HPLC chromatographic retention times (t R) of the metabolites to be established in a single analytical step. 3. In total, three 19 F-NMR signals could be detected, of which one was identified as the parent compound. Two earlier eluting metabolites were identified to be α - and β -glucoside conjugates of 3-TFMA. 4. Metabolites at the lower (sublethal) exposure levels of 10, 1 and 0.1 µg cm -2 escaped identification by 19 F/ 1 H-HPLC-NMR spectroscopy as outlined here and will require concentration prior to analysis.