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In the environment, chemical substances appear as complex mixtures and consequently organisms are exposed to a variety of chemicals from different sources (e.g. wastewater treatment plants, agriculture runoffs). When studying chemical mixtures, there are two conceptual models usually used to predict toxicity: the Independent Action (IA) and Concentration Addition (CA) models. However, deviations from these reference models can occur as synergism or antagonism, dose ratio or dose level dependency. The aim of the present study was to investigate the effects of triclosan and carbendazim, and their binary mixture to Daphnia magna. With this purpose, immobilisation, feeding inhibition, and reproduction were assessed as main ecotoxicity endpoints. In addition, in vivo genotoxicity of both chemicals was investigated using the comet assay. In the single exposure, carbendazim was more toxic to D. magna than triclosan. When daphnids were exposed to both single compounds, DNA damage was observed. Concerning mixture exposures, different endpoints followed different patterns of response, from additivity: IA model (feeding inhibition and reproduction data), to deviations that indicate interaction between chemicals inside the organism: dose level dependency (immobilisation data) and dose ratio dependency (DNA damage). This study showed that additivity does not rule the dose-effect relation in chemical mixtures of carbendazim and triclosan and interactions between both chemicals might induce generally higher toxicity than predicted based on single chemical exposures.

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Carbendazim is a fungicide largely used in agriculture as a plant protection product. As a result of agricultural runoffs, drainage, and leaching, it reaches surface waters at concentrations possibly hazardous to aquatic communities. Because of potential and continuous release of carbendazim to aquatic systems, long-term exposure to aquatic organisms should be addressed. To fill the knowledge gap, the present study evaluated the responses of multiple generations of Daphnia magna (clone K6) to an environmentally relevant concentration of carbendazim (5 μg/L). Twelve successive generations were evaluated, and the effects in these offspring were compared with those from a control population. Neonates' fitness was assessed through immobilization, reproduction, and feeding activity tests, along with the comet assay for in vivo DNA damage evaluation. Recovery from long-term exposure was also assessed. In the F5 generation, the results revealed that when daphnids were re-exposed to carbendazim, DNA damage was higher in daphnids continuously exposed to carbendazim than those from clean medium. After daphnids were moved to a clean medium, a low recovery potential was observed for DNA damage. Daphnids exposed continuously for 6 generations (F6) to carbendazim displayed an increase in feeding rates when re-exposed to carbendazim compared with F6 daphnids reared in clean medium. Continuous exposure of daphnids to carbendazim induced a significant increase in DNA damage from the F0 to the F12 generation. Deleterious effects of the multigenerational exposure to carbendazim were more prominent at a subcellular level (DNA damage) compared with the individual level. Environ Toxicol Chem 2017;36:383-394. © 2016 SETAC.