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GODOY, A.A. Assessment of the single and mixture ecotoxicity of pharmaceuticals of environmental concern using aquatic test organisms. 2019. 386 f. Tese (Doutorado) – Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, São Paulo, 2019. Pharmaceuticals are contaminants of emerging concern which have been a target of increasing attention by the scientific community. Pharmaceuticals presenting high consumption, incomplete metabolism and incomplete removal at wastewater treatment plants have been frequently detected in aquatic ecosystems worldwide. This is the case of the pharmaceuticals metformin (MET), bisoprolol (BIS), sotalol (SOT) and ranitidine (RAN). However, ecotoxicity data for these contaminants are scarce, especially regarding behavior effects and chronic toxicity. In addition, the knowledge regarding the joint toxicity of these pharmaceuticals on non-target organisms is still incipient, which makes their environment risk assessment uncertain. This study aimed to fill these knowledge gaps for these four pharmaceuticals, by carrying out toxicity tests using five test organisms from three trophic levels. Different endpoints were assessed in tests with Raphidocelis subcapitata (algae), Lemna minor (macrophyte), Daphnia similis (crustacean), Hydra attenuate (cnidarian) and Danio rerio (fish). The binary and quaternary mixture acute toxicity for these pharmaceuticals were assessed on D. similis and D. rerio embryo tests, respectively. This study also aimed to evaluate the predictive accuracy of the Concentration addition (CA) and the Independent action (IA) classic models. In addition, the nature of the possible toxicological interactions between the pharmaceuticals in binary mixtures were also evaluated, using the Combination Indexisobologram (CI) method. The modelling of the concentration-response curves and the associated statistical analyses were performed using the automated spreadsheet ToxCalcMix v.1.0 and the software OriginPro 2015. The software CompuSyn was used for performing the mixture analyses with the CI method. The experimental planning of the binary mixture tests was performed using the fractioned factorial design, in order to cover several possible ratio and level-dependent effects with a reduced number of test organisms. The results obtained in this study are shown in four articles. In article 1, we provided a critical review and discussed the misunderstandings, deficiencies and data gaps on the ecotoxicity data of pharmaceuticals and personal care products mixtures published in the literature. In the following articles, the results obtained from the single and mixture toxicity tests performed in this study were presented and discussed. The pharmaceuticals MET (article 2) and BIS (article 3) were classified as hazardous to the aquatic environment, in the acute toxicity category. However, an ecological risk is not expected for the pelagic freshwater species exposed to these two pharmaceuticals, based on the chronic data obtained. The results obtained from the mixture toxicity tests (article 4) showed that most of the observed toxicity effects from the binary mixtures were in the zone between the predicted effects by the CA and IA models. The CI model showed to be an useful tool to describe the possible toxicological interactions occurring between the pharmaceuticals in joint action. Even statistically significant non-effect concentrations of the pharmaceuticals added up to induce significant adverse effects in mixtures (something from nothing). It was concluded that ecological risk assessment based on single toxic effects can underestimate the real impact of environmental contaminants on aquatic ecosystems.