Biotransformation of musk xylene in trout haemoglobin: Dose-response and toxicokinetics of musk xylene metabolites haemoglobin adducts by gas chromatography-mass spectrometry

Musk xylene (MX) is frequently used as a fragrance in commercial toiletries. Biotransformation of MX into 4-amino-MX (4-AMX) and 2-amino-MX (2-AMX) metabolites in rainbow trout haemoglobin (Hb) has been described. The dose-response relationship and toxicokinetics of the metabolites as adducts in the Hb were determined by gas chromatography (GC)-electron capture negative chemical ionization (NCI)-mass spectrometry (MS), and GC-electron ionization (EI)-MS/MS, using selected ion monitoring (SIM). The trout were subjected to a single exposure of 0.010, 0.030, 0.10, and/or 0.30 mg MX/g of fish. Hb samples were collected from exposed and control fish, and analysed subsequent to exposure at intervals of 24, 72, and 168 h. Alkaline hydrolysis released 4-AMX and 2-AMX metabolites from the Hb, and the solutes were extracted into n -hexane. The extracts were preconcentrated and analysed. The presence of the metabolites in the Hb extracts was confirmed based on agreement of similar mass spectral features from NCI/MS and EI-MS/MS spectra, and retention times of the metabolites with standards. The NCI/MS results were used for dose-response and toxicokinetics measurements. For dose-response, the concentrations of adducts of the metabolites increased with dosage, and a maximum adduct formation was observed at 0.10 mg g -1 , beyond which it decreased. The average concentrations of 4-AMX and 2-AMX at a dosage of 0.10 mg g -1 were 700 and 7.4 ng g -1 , respectively. For toxicokinetics, the concentration of the metabolites in the Hb reached a maximum in the 3 day sample after administration of MX. Further elimination of the metabolites exhibited kinetics with a half-life estimated to be 1-2 days, assuming first-order kinetics. Quantitations were made based on an internal standard and a calibration plot. In control samples, non-hydrolysed Hb, and reagent blank extracts, the metabolites were not detected. The limits of detection for 4-AMX and 2-AMX in the Hb were approximately 1.7 and 1.4 μg L -1 , respectively, based on a signal-to-noise ratio of 3 with NCI/MS.