It may be hypothesised that as the bioavailable background concentration of an essential metal increases (within natural limits), the natural tolerance (to the metal) of the acclimated/adapted organisms and communities will increase. In this study the influence of acclimation to different copper concentrations on the sensitivity of the freshwater cladoceran Daphnia magna Straus was investigated. D. magna was acclimated over three generations to environmentally relevant copper concentrations ranging from 0.5 to 100 microg Cu/l (copper activity: 7.18 x 10(-15) to 3700 x 10(-12) M Cu2+). A modified standard test medium was used as culture and test medium. Medium modifications were: reduced hardness (lowered to 180 mg CaCO3/l) and addition of Aldrich humic acid at a concentration of 5 mg DOC/l (instead of EDTA). The effects of acclimation on these organisms were monitored using acute mortality assays and long-term assays in which life table parameters, copper body concentrations and energy reserves were used as test endpoints. Our results showed a two-fold increase in acute copper tolerance with increasing acclimation concentration for second and third generation organisms. Copper acclimation concentrations up to 35 microg Cu/l (80 pM Cu2+) did not affect the net reproduction and the intrinsic growth rate. The energy reserves of the acclimated daphnids revealed an Optimal Concentration range (OCEE) and concentrations between 5 and 12 microg Cu/l (0.5-4.1 pM Cu2+) and 1 and 35 microg Cu/l (0.023-80 pM Cu2+) seemed to be optimal for first and third generation daphnids, respectively. Lower and higher copper concentrations resulted in deficiency and toxicity responses. It was also demonstrated that up to 35 microg Cu/l, third generation daphnids were able to regulate their total copper body concentration. These results clearly indicate that bioavailable background copper concentrations present in culture media have to be considered in the evaluation of toxicity test results, especially when the toxicity data are used for water quality guideline derivation and/or ecological risk assessment for metals.