Toxicity of metal mixtures to a tropical freshwater alga (Chlorella sp.): The effect of interactions between copper, cadmium, and zinc on metal cell binding and uptake

The individual and combined effects of copper, cadmium, and zinc on the cell division rate of the tropical freshwater alga Chlorella sp. were determined over 48 to 72 h. Metal mixtures were prepared based on multiples of their single‐metal median effective concentration (EC50) values, i.e., toxic units (TU) using a triangular mixture design with five toxicant levels (0, 0.75, 1.0, 1.25, and 1.5 TU). Single‐metal EC50 values after a 72‐h exposure were 0.11, 0.85, and 1.4 μM for copper, cadmium, and zinc, respectively. Significant interactions were observed for all metal combinations after 48 and 72 h. An equitoxic mixture of Cu 1 Cd was more than concentration additive (synergistic) to the growth of Chlorella sp., while combinations of Cu + Zn, Cd + Zn, and Cu + Cd + Zn were all less than concentration additive or were antagonistic. To determine the effect of each metal on the uptake of the other, extracellular (membrane‐bound) and intracellular metal concentrations, both alone and in mixtures, were compared. The increased growth inhibition observed for mixtures of Cu + Cd was due to higher concentrations of cell‐bound and intracellular copper in the presence of cadmium compared with copper alone (i.e., cadmium‐enhanced copper uptake). In contrast, both extra‐ and intracellular cadmium concentrations were reduced in the presence of copper. In mixtures of Cu + Zn, copper also inhibited the binding and cellular uptake of zinc, which resulted in decreased toxicity. Zinc had no appreciable effect on the uptake of copper by Chlorella sp. Our results suggest that all three metals share some common uptake and transport sites on Chlorella cells and that copper out competes both cadmium and zinc for cell binding. Determination of metal cell distribution coefficients (Kd) confirmed that Kd values for cadmium and zinc in single‐metal exposures decreased in the presence of copper.