An investigation of biological hydrogen production from glucose by Clostridium beijerinckii was conducted in a synthetic wastewater solution. A study examining the effect of initial pH (range 5.7-6.5) and substrate loading (range 1-3 g COD/L) on the specific conversion and hydrogen production rate has shown interaction behaviour between the two independent variables. Highest conversion of 10.3 mL H(2)/(g COD/L) was achieved at pH of 6.1 and glucose concentration of 3 g COD/L, whereas the highest production rate of 71 mL H(2)/(h L) was measured at pH 6.3 and substrate loading of 2.5 g COD/L. In general, there appears to be a strong trend of increasing hydrogen production rate with an increase in both substrate concentration and pH. Butyrate (14-63%), formate (10-45%) and ethanol (16-40%) were the main soluble products with other volatile fatty acids and alcohols present in smaller quantities. (C) 2009 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.