BENZO[A]PYRENE METABOLISM AND XENOBIOTIC-STIMULATED REACTIVE OXYGEN SPECIES GENERATION BY SUBCELLULAR FRACTION OF LARVAE OF TURBOT (SCOPHTHALMUS-MAXIMUS L)

NADPH-dependent H-3-benzo[a]pyrene (BaP) metabolism and basal and xeno biotic-stimulated NAD(P)H-dependent reactive oxygen species (ROS) prod uction were investigated in 11,600 g supernatants of 4-day-old (yolk s ac) larvae of turbot (Scophthalmus maximus L.). BaP metabolites were r esolved by HPLC and detected radiometrically. ROS were quantified by t he iron-EDTA mediated production of hydroxyl radical (OH) that was det ected by its oxidation of 2 keto-4-methiolbutyric acid (KMBA) to yield ethylene. BaP metabolism produced phenols, dihydrodiols and diones (q uinones) (respectively, 54, 32, and 14% of free metabolites) and putat ive protein adducts. Metabolites identified by retention time included the 7,8-dihydrodiol, the 1,6-, 3,6- and 6,12-diones and the 3- and 9- phenols. Pre-exposure of turbot larvae to 5 ppb BaP for 24 hr caused a n approximate 2-fold increase in both BaP metabolism and 7-ethoxyresor ufin O-deethylase activity, indicative of the induction of cytochrome P4501A and its involvement in BaP metabolism. Basal KMBA oxidation rat es were similar for NADH and NADPH. inhibition studies indicate that O H was formed via the production of superoxide anion radical and hydrog en peroxide. Basal ROS production was stimulated up to 3-fold by a ran ge of redox cycling aromatic hydrocarbon quinones and indicated to be stimulated by other xenobiotics, including nitroaromatics. The results indicate biotransformation and ROS production as potential mechanisms of toxicity in larval fish.