Acute Toxicity of Electronic Cigarette Refill Liquids in Aquatic Vertebrates: Comparative Responses of Fish Embryos, Fish Juveniles and Amphibian Larvae

The increasing use of electronic cigarettes has raised concerns regarding the potential environmental impact of e-cigarette refill liquids and their release into aquatic ecosystems. Despite their growing consumption worldwide, information concerning their ecotoxicological effects on aquatic vertebrates remains limited. In the present study, the acute toxicity of an electronic cigarette refill liquid was evaluated using different aquatic vertebrate models, including fish eggs (Danio rerio), juvenile fish (Jenynsia multidentata), and amphibian larvae (Rhinella arenarum). Acute toxicity assays were performed using a stock solution of e-cigarette liquid (1 mL？L^？1), from which different dilutions were prepared. Juvenile fish and amphibian larvae were exposed to concentrations ranging from 5% to 50% of the stock solution for 96 h, while fish eggs were exposed to concentrations between 3% and 50% for 72 h. Mortality was recorded at regular intervals throughout the exposure period. No mortality was observed in J. multidentata juveniles or R. arenarum larvae at any of the concentrations tested, indicating low acute toxicity under the experimental conditions employed. In contrast, D. rerio eggs exhibited concentration-dependent mortality, with the highest effects observed at 50% of the stock solution, where mortality ranged between 66% and 100% in duplicate assays. Lower concentrations produced variable responses, although embryonic stages consistently showed greater sensitivity than juvenile organisms. The results demonstrate that the toxicological response to e-cigarette refill liquids is strongly influenced by developmental stage. While juvenile fish and amphibian larvae appeared relatively resistant to exposure, fish embryos showed increased susceptibility, suggesting that early developmental stages may represent sensitive indicators for assessing the environmental risk of emerging contaminants associated with electronic cigarettes. These findings contribute to the understanding of the ecotoxicological effects of vaping-related products and highlight the importance of including multiple biological models and life stages in environmental risk assessments.