The application of thermophilic bacteria to the production of hydrogen (H-2) from a cellulosic biomass has captured attention of engineering researchers. Thermotoga neupolitana is considered as a potential hydrogen producer because of its ability to be used for the direct H-2 fermentation from raw cellulose, but its H-2 yield is not high. In this study, three chemical pretreatment methods with an ionic liquid, acid and alkali, respectively, were tested on cellulose to enhance the cellulose conversion of this strain into H-2. Batch cultivations were carried out to investigate the influence of the chemical pretreatment methods on H-2 production under non-sterile conditions. The highest H-2 concentration in the head-space obtained from the ionic liquid-pretreated cellulose was 36.1% (v/v) with ethanol as an anti-solvent. This was clearly higher than that from the acid- and alkali-pretreated celluloses (22-24%). Cultivation with cellulose pretreated with the ionic liquid at a concentration of 10% combining with N-2 sparging showed the maximum cumulative H-2 yield of 1280 mL H-2/L culture. This value is approximately 10-fold higher than that of raw cellulose (122 mL H-2/L culture). The H-2 yield obtained from fermentation with ionic liquid- pretreated cellulose was 2.2 mol H-2/mol glucose equivalents. The results showed that the pretreatment of cellulose using an ionic liquid has considerable potential for improving the direct conversion of cellulosic substrates into H-2 by bacteria. (C) 2008 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved,