The copper extraction yield from thermophilic bioleaching of chalcopyrite depends on temperature, pH, and the oxidation-reduction potential (ORP), as well as on the activity of the thermophile used. The copper extraction yields obtained with three thermophiles under various pH and temperature conditions and with different initial amounts of Fe3+ were studied. The results indicated that because of the low ability of Acidianus brierleyi to leach iron as Fe3+, high biomass concentrations were reflected by ORP close to a critical value (450 mV, Ag-0/AgCI reference), at which copper extractions were highest. By contrast, because of the higher ability of Sulfolobus metallicus and Metallosphaera sedula to leach iron as Fe3+, high biomass concentrations were reflected by high ORP which in combination with the precipitation of Fe3+ as jarosite attenuated the leaching rates. Therefore, optimum temperatures for the growth of thermophiles did not always mean high copper extraction yields. Generally, highest copper extractions were obtained at initial pH 1.5. However, higher copper extractions were observed at initial pH 2.5 than at pH 2.0. suggesting that at high pH the bioleaching of chalcopyrite is controlled by the ORP rather than by the pH or temperature. The bioleaching capacity of A. brierleyi was reduced or suppressed when insufficient initial Fe3+ was provided to trigger the leaching reaction, whereas S. metallicus and M. sedula were less sensitive to the initial availability of Fe3+. This result confirmed that a direct enzymatic attack on the mineral surface could initiate the leaching reaction, but later ORP governed the leaching rate of chalcopyrite. (C) 2008 Elsevier B.V. All rights reserved.