Arundo donax hydrolysate (ADH), obtained by steam explosion and enzymatic treatment, was exploited as a substrate for anaerobic digestion aimed to the production of hydrogen. Mixed cultures of hydrogen-forming anaerobic bacteria selected from a primary sludge digester were used as inocula. Methanogens were removed from the wild consortium by the use of thermal treatments (autoclaving, freezing and thawing). Either with glucose or ADH as carbon sources, the soluble anaerobic fermentation products were butyric acid, lactic acid, acetic acid, formic acid and ethanol. The collected biogas consisted mainly of hydrogen (86-97%) and carbon dioxide. Hydrogen yield with ADH was lower than that found with glucose fermentation with the same inoculum. Outgrowth of lactic acid bacteria during ADH fermentation was presumably the cause of the low observed yield. Further optimization of ADH pre-treatment and stabilisation of microbial consortium are necessary in order to improve hydrogen yield on ADH.