For over 40 years, anurans have been used as a study model to assess the adverse effects of benzo(α)pyrene (BαP), which include genotoxic, hepatotoxic, and immunotoxic effects. In these studies, BαP is administered cutaneously or by injection, with no comparison between two or more routes. The purpose of this study is to assess whether the effect of BαP is influenced by its route of administration, using the response of hepatic biomarkers of Physalaemus nattereri. Specimens (n = 108) were collected and divided into three experimental treatments (cutaneous, injection, and oral) and three experimental times (one, three, and seven days). Specimens received 0.02 ml of pure mineral oil (control) or mineral oil containing 2 mg/kg of BαP. The BαP causes changes in morphological (melanin, hemosiderin, lipofuscin, and mast cells) and biochemical (superoxide dismutase and glutathione S-transferase) hepatic biomarkers. Compared to biochemical, morphological biomarkers underwent a greater number of significant changes due to the treatment with BαP. The route of exposure alters the effects of BαP, mainly seen in morphological biomarkers, especially the pigments melanin, hemosiderin, and lipofuscin. In these pigments, the effect of the exposure pathway changes according to the analyzed biomarker, and the exposure time modulates the exposure pathway effect. These results are unprecedented for anurans and contribute to the field of herpetology and ecotoxicology.
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