Toxicity effects of copper (II) on the marine dinoflagellateAmphidinium carterae: Influence of metal speciation

Abstract

The influence of copper(II) concentration and speciation on the growth of Amphidinium carterae in batch culture in a modified ESAW medium was evaluated by two different experimental designs: (i) copper(II) was added to the cultures 2 h after cell inoculation and (ii) copper(II) had a 24 h pre-equilibration period in the medium before cell inoculation. To complement the biological data, the following information on the chemical/biological system was obtained: (a) computational speciation of the culture medium under chemical equilibrium conditions: (b) total and electrochemically labile copper concentrations in the culture medium during the toxicity experiments; and (c) cellular levels of copper. The labile fraction of copper was measured by potentiometric stripping analysis (PSA) at a deposition potential (E _d) of — 0·3 V. At this E _d the dissociation of Cu-EDTA complexes was negligible. In both kinds of experiments the effects of copper were more evident after 4 days than after 7 or 10 days of exposure. When compared with situation (ii) pronounced toxic effects were observed under situation (i): (1) A. carterae growth rate was affected at lower copper concentration and in a more acute way: (2) the cells were exposed to significantly higher levels of labile copper (2- to 3-fold more than in situation (ii) for 13·4 µm total copper) and (3) after 2 h of exposure to the metal the copper sorbed by the cells was about 2-fold higher. These results provide evidence for the importance of kinetic parameters in toxicity studies. Experimental copper lability together with speciation calculations provided additional evidence that the bioavailable copper and its cellular toxicity are directly related to the labile metal concentration. This is higher than but directly proportional to the initial free metal concentration in the medium, as was experimentally verified. PSA-derived copper lability provided a reasonable indicator of metal bioavailability to A. carterae in this synthetic seawater medium. A first response of A. carterae to copper toxicity is the release of the metal due to re-equilibration between copper bound to the cell surface and to the ligands in the medium and/or efflux. A different mechanism must subsequently be activated, allowing growth despite the relatively high cellular levels of copper observed.