Formation of nitro musk adducts of rainbow trout hemoglobin for potential use as biomarkers of exposure

The high use of nitro musk xylene (MX) and musk ketone (MK) as fragrances, and their persistence and bioaccumulation potential make them ubiquitous environmental contaminants. The 4-amino-MX (AMX) and 2-amino-MK (AMK) metabolites have been detected in trout fish hemoglobin (Hb) samples by gas chromatography-ion trap-mass spectrometry (GC-MS). Twelve Hb samples prepared from rainbow trout that were exposed to MX and MK, over a period of 24 and 72h, were analyzed. Amino metabolites were liberated by basic hydrolysis and extracted from the fish Hb into n-hexane. The extract was concentrated, analyzed, and spiked with a standard solution (80pg/μl) of AMX or AMK and reanalyzed. Concentrations of AMX from 10 to 25ng/g were detected in Hb from fish taken 24 and 72h after MX exposure. At 24 and 72h after MK exposure, the concentration of AMK was found to be 25-51ng/g and 9.5-25ng/g, respectively. Concentrations of AMK in Hb from two of the three trout were substantially lower after 72h compared with 24h exposure. The AMX and AMK metabolites were not detected in four control samples. Average recoveries exceeding 89 and 86% could be achieved for AMX and AMK, respectively, with a coefficient of variation (CV) around 5%.