This paper reports investigations carried out to determine the optimum culture conditions for the production of hydrogen with a recently isolated strain Clostridium butyricum CWBI1009. The production rates and yields were investigated at 30 degrees C in a 2.3 L bioreactor operated in batch and sequenced-batch mode using glucose and starch as substrates. In order to study the precise effect of a stable pH on hydrogen production, and the metabolite pathway involved, cultures were conducted with pH controlled at different levels ranging from 4.7 to 7.3 (maximum range of 0.15 pH unit around the pH level). For glucose the maximum yield (1.7 mol H(2) mol(-1) glucose) was measured when the pH was maintained at 5.2. The acetate and butyrate yields were 0.35 mol acetate mol(-1) glucose and 0.6 mol butyrate mol(-1) glucose. For starch a maximum yield of 2.0 mol H(2) mol(-1) hexose, and a maximum production rate of 15 mol H(2) mol(-1) hexose h(-1) were obtained at pH 5.6 when the acetate and butyrate yields were 0.47 mol acetate mol(-1) hexose and 0.67 mol butyrate mol(-1) hexose. (C) 2010 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved.