In this study, biohydrogen production from glucose by two fermentative bacteria (Clostridium butyricum, a typical strictly anaerobic bacterium, and Klebsiella pneumoniae, a well-studied facultative anaerobic and nitrogen-fixing bacterium) are stiochiometrically analyzed according to energy (ATP), reducing equivalent and mass balances. The theoretical analysis reveals that the maximum yield of hydrogen on glucose by Clostridium butyricum is 3.26 mol/mol when all acetyl-CoA entering into the acetate pathway (alpha= 1), which is higher than that by Klebsiella pneumoniae under strictly anaerobic conditions. In the latter case, the maximum yield by Klebsiella pneumoniae is 2.86 mol hydrogen per mol glucose when five sevenths of acetyl-CoA is transformed to acetate. However, under microaerobic condition the maximum yield of hydrogen on glucose by Klebsiella pneumoniae could reach 6.68 mol/mol if all acetyl-CoA entered into tricarboxylic acid (TCA) cycle (gamma = 1) and a quantity of 53% of the reducing equivalents generated in the metabolism were completely oxidized by molecular oxygen. On the other hand, the relationship between hydrogen production and biomass formation is distinct by Clostridium butyricum from that by Klebsiella pneumoniae. The former yield of hydrogen on glucose increases as biomass. In contrast, the latter one decreases as biomass in a certain range of molar fraction of acetate in total acetyl-CoA metabolism (5/7 >= beta >= 0). Microacrobic condition is favorable for high hydrogen production with low biomass formation by Klebsiella pneumoniae in a certain range of the molar fraction of all reducing equivalents oxidized completely by molecular oxygen (0.53 <= 6 <= 0.83). (c) 2005 International Association for Hydrogen Energy. Published by Elsevier Ltd. All fights reserved.