Hydrogen production from the anaerobic fermentation of feedlot cattle manure was examined in batch cultures over a temperature range from 36 to 60 degrees C at a pH of 5.2. The amount of hydrogen produced increased with temperature to a maximum of 65 L H-2 kg TS-1 at 52 degrees C. At temperatures > 52 degrees C, acetate was the main volatile fatty acid (VFA) accumulated, while at <52 degrees C butyrate accumulated the most. Formate was detected in the 56 and 60 degrees C treatments but was absent in all others. Thermophilic conditions resulted in the highest hydrogen production rates, with maximum hydrogen production occurring 52 degrees C. Changing incubation temperature by small (4 degrees C) increments up or down from 52 degrees C resulted in changes in the metabolic flux (conversion of substrate to VFA and gaseous products) of the anaerobic digestion system. These findings indicate that the hydrogen production potential of anaerobic systems utilizing heat treated cattle manure as inoculum is affected greatly by incubation temperature. (C) 2008 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.