Problem statement: Some component of fermentation medium showed to reduce the bacterial production of hydrogen. Approach: Reinforced clostridium medium is a selected medium for Clostridium species. Reformulation this medium regarding hydrogen production may focus on such medium composition that enhance or reduce the bacterial productivity. The optimum pH and temperature for hydrogen production were at initial pH of 7.0 and 30ºC. Results: The results show that both nitrogen source and its concentration affected biomass growth as well as H 2 yield. Yeast extract at concentration of 13 gL −1 was the best organic nitrogen source and resulted in hydrogen yield (Y P/S) of 308 mL g −1 glucose utilized with biomass concentration of 1.1 gL −1 , hydrogen yield per biomass (Y P/X) of 280 mL g −1 L −1 , biomass per substrate utilized (Y X/S) of 0.22 and produced hydrogen in gram per gram of glucose utilized (Y H2/S) of 0.0275. C/N of 70 enhanced the Y P/S from 308 mL g −1 to 350 mL g −1 glucose utilized with biomass concentration of 1.22 gL −1 , Y P/X of 287 mL g −1 L −1 , Y X/S of 0.244 and (Y H2/S) of 0.03125. In the absence of sodium chloride and sodium acetate further enhanced Y P/S from 350 mL g −1 glucose utilized to 391 mL g −1 glucose utilized with maximum hydrogen productivity of 77.5 mL L −1 h −1 , whereas RCM medium gave the highest hydrogen productivity of 63.5 mL L −1 h −1. Results also show that Sodium Chloride and Sodium Acetate in the medium adversely affect growth. Removal of both components from the medium enhanced the biomass concentration from 1.22-1.34 gL −1 , Y P/X of 254 mL g −1 L −1 , Y X/S of 0.268 and (Y H2/S) of 0.0349. Conclusion: The medium an improved containing (glucose 5 gL −1 , Yeast extract gL −1 , L-Cystine. HCl 1 gL −1 and Bacteriological agar 0.5 gL −1), was able to enhance the hydrogen productivity.
