One of the challenges in the development of integrated dark and photofermentative biological hydrogen production systems is the presence of ammonium ions in dark fermentation effluent (DFE). Ammonium strongly inhibits the sequential photofermentation process, and so its removal is required for successful process integration. In this study, the removal of ammonium ions from molasses DFE using a natural zeolite (clinoptilolite) was investigated. The samples were treated with batch suspensions of Na-form clinoptilolite. The ammonium ion concentration could be reduced from 7.60 mM to 1.60 mM and from 12.30 mM to 2.40 mM for two different samples. Photofermentative hydrogen production on treated and untreated molasses DFE samples were investigated in batch photo-bioreactors by an uptake hydrogenase deleted (hup(-)) mutant strain of Rhodobacter capsulotus. Maximum hydrogen productivities of 1.11 mmol H-2/L-c.h and 1.16 mmol H-2/L-c.h and molar yields of 79% and 90% were attained in the treated DFE samples, while the untreated samples resulted in no hydrogen production. The results showed that ammonium ions in molasses DFE could be effectively removed using clinoptilolite by applying a cost-effective, simple batch process. Copyright (C) 2012, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.