IN-VIVO FORMATION OF MUTAGENS BY INTRAPERITONEAL ADMINISTRATION OF POLYCYCLIC AROMATIC-HYDROCARBONS IN ANIMALS DURING EXPOSURE TO NITROGEN-DIOXIDE

Consumption of fossil fuels has increased indoor and outdoor concentra tions of polycyclic aromatic hydrocarbons (PAHs) and nitrogen dioxide (NO2). To study the combined effect of PAH administration and NO2 expo sure on mutagenicity of urine from animals we injected 400 mg/kg body wt i.p. one of five kinds of PAH (pyrene, fluoranthene, fluorene, anth racene and chrysene) into ICR mice, Wistar rats, Syrian golden hamster s or Hartley guinea pigs after exposure to 20 p.p.m. NO2 gas for 24 h and then exposed the animals to NO2 gas for an additional 24 h, During the latter 24 h we collected the urine and assayed its mutagenicity w ith the Ames Salmonella strains after treatment with P-glucuronidase a nd arylsulfatase and extraction with dichloromethane. The urine from m ice treated with both PAH and NO? showed high mutagenicity for Salmone lla typhimurium strains TA98 and TA100, whereas the urine from mice tr eated with PAH and air showed almost no mutagenic activity, The mutage nicity was decreased in nitroreductase- and acetyltransferase-deficien t strains TA98NR and TA98/1,8-DNP6 respectively, Treatment with a mixt ure of 20 % of each of the five kinds of PAH and NO2 augmented the uri nary mutagenicity of mice 1.5-fold, The urine from hamsters treated wi th pyrene or fluoranthene and NO2 was also highly mutagenic, but that from rats or guinea pigs was not very mutagenic, The mutagenicity was also decreased in strains TA98NR and TA98/1,8-DNP6. These results sugg est that the urine contains nitro compounds and that the nitration of PAHs occurs in the body of animals under exposure to NO2 gas. Actually , the nitrated metabolites of pyrene, 1-nitro-6/8-hydroxypyrene and 1- nitro-3-hydroxypyrene, were detected in the urine from mice treated wi th pyrene under exposure to NO2 gas, To elucidate the mechanism of in vivo nitration, NO2 (20 p.p.m.) was bubbled through 50 mM Tris-HCl buf fer (pH 7.4) or dichloromethane solution containing pyrene or l-hydrox ypyrene (10 mu g/ml). Pyrene was not nitrated by NO2 in either aqueous or organic solutions, However, l-hydroxypyrene was changed to nitrohy droxy-pyrenes by NO2 in the Tris-HCl buffer, but not in the organic so lution, Ascorbic acid, alpha-tocopherol, glutathione, oleic acid and h emoglobin were found to inhibit the nitration of 1-hydroxypyrene in aq ueous solution, The urinary mutagenicity of mice treated with both pyr ene and NO2 was also decreased by oral administration of ascorbic acid and alpha-tocopherol, These results suggest that 1-hydroxypyrene is n itrated by an ionic reaction in the animal body after hydroxylation of pyrene in the liver.