The world has been using fossil fuels to generate energy for centuries and has had adverse effects on the environment; hence renewable energy needs to be discovered and developed. Biohydrogen production is renewable energy since it emits no greenhouse gases and may provide clean energy. Therefore, this study aimed to investigate the optimum headspace ratio and biohydrogen production for suspended and immobilized cells using Palm Oil Mill Effluent (POME) as the fermentation substrate, while its anaerobic sludge acted as the inoculum. Five different ratios were investigated, which are 0.2, 0.3, 0.4, 0.5, and 0.6. These are equivalent to working volume (WV) of 80 mL, 70 mL, 60 mL, 50 mL, and 40 mL, respectively. The solution contained 10 % of inoculum and 90 % (v/v) of the feedstock. For immobilized cells, additional of glass beads as carrier material was added into the solution, using the ratio of 1:1 for anaerobic sludge (mL) to support carrier (g). The kinetic study was investigated using a modified Gompertz equation whereby for suspended cells, the best ratio was 0.3, with the highest biohydrogen concentration of 357.6 ppm. Meanwhile, the optimum ratio for the immobilized cell was 0.2, with the highest biohydrogen concentration of 479.3 ppm. Based on the kinetic studies, the kinetic parameters for suspended cells were: Hm = 89.8 mL, Rm = 6.8 mL/h, and λ = 0.1 hr. Meanwhile for immobilized cell, the kinetic parameters were: Hm = 73.6 mL, Rm = 6.9 mL/h and X λ 0 hr. In conclusion, selecting the suitable headspace ratio could affect the biohydrogen quality and improve the effectiveness of the production rate.
