This study determined hydrogen production, volatile fatty acids (VFAs) generation and cellulose solubilisation from anaerobic dark fermentation of wheat stalk and showed the effect of different mixed microflora. The cumulative hydrogen yields of anaerobic digested activated sludge (AS)-inoculated and anaerobic digested dairy manure (DM)-inoculated system were 23.3 and 37.0 mL/g VS at 204 h, respectively. A modified Gompertz equation was able to adequately describe the production of hydrogen from the batch fermentation by both mixed microflora. During the process, acetate and butyrate accounted for more than 76.1% of total VFAs for both fermentations. The extent of cellulose solubilisation approached 46.6% and 75.2% for AS- and DM-inoculated fermentation, respectively. The X-ray diffraction (XRD) showed that the crystallinities of both fermented stalks were partly disrupted by the mixed microflora, and DM-inoculated fermentation had more disruption than AS-inoculated one. (C) 2010 Elsevier Ltd. All rights reserved.