A gram negative hydrogen producing facultative anaerobe was isolated and characterized as Enterobacter cloacae IIT-BT 08. Hydrogen yields by using this microorganism varied from substrate to substrate (2.2 mol/mol glucose, 6 mol/mol of sucrose and 5.4 mol/mol cellobiose considering 1% w/v substrate in MY medium). The maximum rate of hydrogen production achieved was at 36 degrees C and initial pH 6.0. The maximum rate was 29.63 mmol/(g dry cell per h). The pH profiles of the fermentation broth under aerobic and anaerobic conditions were monitored and found to differ from each other particularly beyond the pH of 4.8. About 28% of substrate energy were recovered in the form of hydrogen using sucrose as a substrate. (C) 2000 Elsevier Science Ltd. All rights reserved.