Abstract
Aerobic, hydrogen oxidizing bacteria are capable of efficient, non-phototrophic CO(2) assimilation, using H(2) as a reducing agent. The presence of explosive gas mixtures requires strict safety measures for bioreactor and process design. Here, we report a simplified, reproducible, and safe cultivation method to produce Cupriavidus necator H16 on a gram scale. Conditions for long-term strain maintenance and mineral media composition were optimized. Cultivations on the gaseous substrates H(2), O(2), and CO(2) were accomplished in an explosion-proof bioreactor situated in a strong, grounded fume hood. Cells grew under O(2) control and H(2) and CO(2) excess. The starting gas mixture was H(2):CO(2):O(2) in a ratio of 85:10:2 (partial pressure of O(2) 0.02 atm). Dissolved oxygen was measured online and was kept below 1.6 mg/L by a stepwise increase of the O(2) supply. Use of gas compositions within the explosion limits of oxyhydrogen facilitated production of 13.1 ± 0.4 g/L total biomass (gram cell dry mass) with a content of 79 ± 2% poly-(R)-3-hydroxybutyrate in a simple cultivation set-up with dissolved oxygen as the single controlled parameter. Approximately 98% of the obtained PHB was formed from CO(2).