The effects of C/N and C/P ratio on hydrogen yield and specific production rate were investigated in dark fermentation of wheat powder solution (WPS) by external addition of N and P since the wheat powder was N and P deficient. A Box-Wilson statistical experiment design approach was used. C/N ratio was varied between 20 and 200 while C/P ratio was between 50 and 1000. A quadratic response function was used to correlate the objective functions with the independent variables and the response function coefficients were determined by regression analysis. Hydrogen yield increased with increasing C/N and C/P ratios. Low nitrogen concentrations required low phosphorous contents for high hydrogen yields. A C/N ratio of 200 and C/P ratio of 1000 yielded the highest H-2 yield of 281. ml H-2 g(-1) starch at STP. SHPR also increased with increasing C/N and C/P ratios. Again the highest SHPR (98 ml H-2 g(-1) biomass h(-1) at STP) was obtained at C/N and C/P ratios of 200 and 1000, respectively. The C/N/P ratio maximizing the yield and formation rate of hydrogen was 100/0.5/0.1 (w w(-1) w(-1)). (C) 2008 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.