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A previously developed biotic ligand model (BLM) was validated for its capacity to predict acute 48-h EC50 values of copper to Daphnia magna in 25 reconstituted media with different pH values and concentrations of artificial dissolved organic carbon, Ca, Mg and Na. Before the BLM validation, fitting of measured (with a copper ion-selective electrode) and calculated (with the BLM) Cu²⁺-activity was performed by adjusting the wham model V (i.e. the metal–organic speciation part of the BLM) copper–proton exchange constant to pK_MHA=1.9. Using this value, the 48-h EC50 values observed agreed very well with BLM-predicted EC50 values for tests performed at pH<8, but not at all for tests performed at pH>8. Additional experiments demonstrated that this was due to toxicity of the CuCO₃ complex, which is the most abundant inorganic copper species at pH>8. This was incorporated into the initial BLM by allowing …