High cell density culture of Anabaena variabilis using repeated injections of carbon dioxide for the production of hydrogen

Cyanobacteria have the unique characteristic of using CO₂ in the air as a carbon source and solar energy as an energy source. Reducing equivalents from the fermentation of carbohydrates are used as the primary electron donors in cyanobacteria for the hydrogen producing enzymes. The cells take up CO₂ first to produce cellular substances, which are subsequently used for H₂ production. Since the optimal operating conditions for the CO₂ uptake and H₂ production are different, a two-stage system can be effectively employed to separate these two phases. In this study, for the efficient production of H₂ in the second stage, the conditions for the effective CO₂ uptake and cell growth in the first stage were characterized, and high cell density culture was carried out using repeated injections of CO₂. The specific growth rate and growth yield based on CO₂ decreased with an increase in light intensity or CO₂ concentration. However, the effect of CO₂ concentration on the growth yield was much smaller than that of a light intensity. A CO₂ uptake rate per unit cell decreased linearly with the initial CO₂ concentration in the gas phase. With repeated injections of CO₂, the CO₂ was continuously consumed and the cell concentration reached 3.7 g dry cell/l in 20 days, which is 6.7 times higher than that in a batch culture without further supply of CO₂. The CO₂ injection in the cell growth phase increased not only the cell concentration but also the hydrogen production per gram cell.