In this study, the effects of alginate and chitosan as entrapped materials in the biofilm formation of microbial attachment on activated carbon was determined for biohydrogen production. Five different batch fermentations, consisting of mixed concentration alginate (Alg), were carried out in a bioreactor at temperature of 60 °C and pH 6.0, using granular activated carbon (GAC) as a primer for cell attachment and colonisation. It was found that the highest hydrogen production rate (HPR) of the GAC–Alg beads was 2.47 ± 0.47 mmol H2/l.h, and the H2 yield of 2.09 ± 0.22 mol H2/mol sugar was obtained at the ratio of 2 g/L of Alg concentration. Next, the effect of chitosan (C) as an external polymer layer of the GAC–Alg beads was investigated as an alternative approach to protecting the microbial population in the biofilm in a robust environment. The formation of GAC with Alg and chitosan (GAC–AlgC) beads gave the highest HPR of 0.93 ± 0.05 mmol H2/l.h, and H2 yield of 1.11 ± 0.35 mol H2/mol sugar was found at 2 g/L of C concentration. Hydrogen production using GAC-attached biofilm seems promising to achieve consistent HPRs at higher temperatures, using Alg as immobilised bead material, which has indicated a positive response in promoting the growth of hydrogen-producing bacteria and providing excellent conditions for microorganisms to grow and colonise high bacterial loads in a bioreactor.

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