Hydrogen gas production from acid hydrolyzed waste wheat starch by combined dark and photo-fermentation was investigated in continuous mode with periodic feeding and effluent removal. A mixture of heat treated anaerobic sludge and Rhodobacter sphaeroides (NRRL-B 1727) were used as the seed culture for dark and light fermentations, respectively with biomass ratio of Rhodobacter/sludge = 3. Hydraulic residence time (HRT) was changed between 1 and 8 days by adjusting the feeding periods. Ground waste wheat was acid hydrolyzed at pH = 3 and 121 degrees C for 30 min using an autoclave and the resulting sugar solution was used as the substrate for combined fermentation after pH adjustment and nutrient addition. The highest daily hydrogen gas production (41 ml d(-1)), hydrogen yield (470 ml g(-1) total sugar = 3.4 mol H(2) mol(-1)glucose), volumetric and specific hydrogen production rates were obtained at the HRT of 8 days. The highest biomass and the lowest total volatile fatty acids (TVFA) concentrations were also realized at HRT = 8 days indicating VFA fermentation by Rhodobacter sp. at high HRTs. The lowest total sugar loading rate of 0.625 g L(-1) d(-1) resulted in the highest hydrogen yield and formation rate. Hydrogen gas production by combined fermentation with periodic feeding was proven to be an effective method resulting in high hydrogen yields at long HRTs. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.