Studies concerning environmental impact of construction materials are very rarely based on experiments performed with real mixed samples and test organisms (ecotoxicological bioassays). In our study, ecotoxicity of two concrete samples from different producers and one concrete-based construction waste in leachate form were assessed. Leachates were treated by various designs, i.e. i) without any treatment, ii) original pH + nutrients addition, iii) pH adjustment to 7.0, and iv) pH adjustment to 7.0 + addition of inorganic nutrients. Ecotoxicological bioassays with freshwater algae Desmodesmus subspicatus, freshwater plant Lemna minor and freshwater invertebrate Daphnia magna were performed. The metal content was determined both in solid samples and leachates. Results showed differences in toxicity level among the concrete sources when original leachates without any treatment were tested. On the contrary, the test design recommended by Czech legislation, i.e. lowering of the pH and addition of nutrients usually significantly decreased the sample toxicity and the potential differences among concrete samples. We suggest that the toxicity of concrete leachates may result not only from the highly alkaline pH but also from the potential persistence of high pH values both within dilution process and time. The higher toxicity was in accordance with higher level of leachate conductivity. Therefore, for the purposes of aquatic ecotoxicity assessment, we recommend either making no pH adjustment or performing the bioassays both with treated and untreated leachates.