Hydrogen Production by Immobilized Cells of Clostridium intestinale Strain URNW Using Alginate Beads

Biological hydrogen (H₂) is a promising candidate for production of renewable hydrogen. Using entrapped cells rather than conventional suspended cell cultures for the production of H₂ offers several advantages, such as improved production yields related to higher cell density, and enhanced resistance to substrate and end-product inhibition. In this study, H₂ production by a novel isolate of Clostridium intestinale (strain URNW) was evaluated using cells entrapped within 2% calcium-alginate beads under strictly anaerobic conditions. Both immobilized cells and suspended cultures were studied in sequential batch-mode anaerobic fermentation over 192 h. The production of H₂ in the headspace was examined for four different initial cellobiose concentrations (5, 10, 20, and 40 mM). Although a lag period for initiation of the fermentation process was observed for bacteria entrapped within hydrogel beads, the immobilized cells achieved both higher volumetric production rates (mmol H₂/(L culture h)) and molar yields (mol H₂/mol glucose equivalent) of H₂ compared with suspended cultures. In the current study, the maximum cellobiose consumption rate of 0.40 mM/h, corresponding to 133.3 mg/(L h), was achieved after 72 h of fermentation by immobilized cells, generating a high hydrogen yield of 3.57 mol H₂/mol cellobiose, whereas suspended cultures only yielded 1.77 mol H₂/mol cellobiose. The results suggest that cells remain viable within the hydrogels and proliferated with a slow rate over the course of fermentation. The stable productivity of immobilized cells over 8 days with four changes of medium depicted that the immobilized cells of the isolated strain can successfully yield higher hydrogen and lower soluble metabolites than suspended cells suggesting a feasible process for future applications for bioH₂ production.