Effects of various pretreatments of the anaerobic sludge on hydrogen production were investigated for enhancing hydrogen production via dark anaerobic fermentation of organic substrates. The pretreatment methods included heat treatment, acid or base treatment, acid/heat treatment and base/heat treatment. After the pretreatment of the anaerobic sludge, productivity of H-2 by the pretreated sludge were evaluated when the culture were grown on glucose-based medium at ambient conditions. The detailed experimental investigation showed highest production of H-2 by the heat-treated culture followed by the heat and acid-treated culture and the heat and base-treated culture. (C) 2012 Published by Elsevier Ltd.