Hydrogen gas production from cheese whey powder (CWP) solution by thermophilic dark fermentation was investigated at 55 degrees C. Experiments were performed at different initial total sugar concentrations varying between 5.2 and 28.5 g L-1 with a constant initial bacteria concentration of 1 g L-1. The highest cumulative hydrogen evolution (257 mL) was obtained with 20 g L-1 total sugar (substrate) concentration within 360 h while the highest H-2 formation rate (2.55 mL h(-1)) and yield (1.03 mol H-2 mol(-1) glucose) were obtained at 5.2 and 9.5 g L-1 substrate concentrations, respectively. The specific H-2 production rate (SHPR = 4.5 mL h(-1) reached the highest level at 20 g L-1 total sugar concentration. Total volatile fatty acid (TVFA) concentration increased with increasing initial total sugar content and reached the highest level (14.15 g L-1) at 28.5 g L-1 initial substrate concentration. The experimental data was correlated with the Gompertz equation and the constants were determined. The optimum initial total sugar concentration was 20 g L-1 above which substrate and product (VFA) inhibitions were observed. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.