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Keywords:

  • Polycyclic aromatic hydrocarbon;
  • Polychaetes;
  • Pyrene hydroxylase;
  • Glucuronosyl transferase;
  • Sulfotransferase

Abstract

In vivo and in vitro biotransformation of the polycyclic aromatic hydrocarbon (PAH) pyrene was investigated in the marine polychaete Nereis virens. Assays were designed to characterize phase I and II enzymes isolated from gut tissue. High-pressure liquid chromatography measurement of 1-hydroxypyrene, pyrene-1-glucuronide, pyrene-1-sulfate, and pyrene-1-glucoside appeared to be a sensitive method for estimating the activity of pyrene hydroxylase, glucuronosyl transferase, and sulfotransferase. Total pyrene in gut tissue after a 5-d exposure to 10 μg/g dry weight pyrene constituted 65% pyrene-1-glucuronide, 12% pyrene-1-sulfate, 2% pyrene-1-glucoside, 4% 1-hydroxypyrene, and 17% pyrene, indicating that glucuronidation is the prominent phase II pathway in this organism. Only pyrene hydroxylase activity was induced significantly after pre-exposure to sediment-associated pyrene (10 μg/g dry wt). Apparent kinetic parameters were determined for all enzymatic reactions. Glucuronidation was confirmed as being the prominent phase II reaction, based on its high apparent maximum velocity (Vmax(a)). Sulfation had the lowest apparent Michaelis constant (Km(a)), indicating high specificity. Apparent kinetic parameters for pyrene hydroxylase activity were changed after induction with pyrene. Induced worms showed increased Vmax(a) and decreased Km(a) compared to noninduced worms, indicating that the relative amount of the cytochrome P450 enzyme(s) responsible for pyrene hydroxylation is increased. Our findings show that N. virens has a high biotransformation capacity for PAHs, with inducible pyrene hydroxylase activity, and that the most prominent phase II pathway in this organism is glucuronidation.