Evaluating the acute toxicity of binary and ternary mixtures of Cu, Zn, As and Se to Daphnia magna: a component-based approach using isobologram analyses and ternary diagrams

The acute toxicity of binary and ternary combinations of Cu, Zn, As and Se to Daphnia magna was investigated. The aim was to provide a detailed characterization of the interactive effects of these elements in relation to aquatic toxicity. The binary and ternary combinations were evaluated using isobolograms, the concentration addition (CA) model, the independent action (IA) model, and the Hewlett model. They were also implemented using the MixModel package in R. This approach facilitated the identification of potential synergism, additivity, and antagonism, in addition to the characterization of binary and ternary interactions within a mixture composition space. For the binary combinations, Cu-Zn and Zn-Se had the most robust evidence for synergism with EC₅₀ estimates <1 toxicity unit (TU), as low as 0.27 TU for Cu-Zn, and 0.30 TU for Zn-Se. Combinations of As-Cu, As-Se, Cu-Se, and As-Zn were mostly additive, with EC₅₀ estimates ∼1 TU for all evaluated ratios. In all of the tested ternary combinations, Cu was identified the most toxic component. Emergent effects, which were defined as toxicity responses arising from combinations of toxicity which were not dependent on binary combinations, were observed in some ternary combinations. Synergism was observed in the As-Cu-Se and As-Cu-Zn mixtures, suggesting that binary combinations alone are insufficient for predicting interactions involving more than two components. Ternary diagrams also showed potential areas of high toxicity (TU>3), mostly in the Cu-dominated mixtures. These observations suggest that the toxicity assessment of trace metals and metalloids commonly found in mining and metallurgical effluents (in this case Cu, Zn, As, and Se) must take into account the effect of mixtures and not rely on substance-by-substance assessments.