Microbial high cell density culture system with regenerated high purity oxygen

Abstract

Developing high cell density culture technologies has attracted a lot of attentions as it may guarantee a high product level, which has a significant effect on the product separation processes and the production cost. In large-scale aerobic fermentation processes, however, a high cell density generally could not be achieved due to the oxygen limitation. In this study, high purity oxygen was employed to eliminate the oxygen limitation and to investigate the potential influences of high purity oxygen on fermentation processes. Two important microorganisms, Ralstonia eutropha for producing poly(hydroxyalkanoate) and recombinant E. coli BL21 (DE3) for producing human growth hormone (h-GH), were cultivated in the 5-L, 30-L, and 300-L fermentors with air and high purity oxygen, respectively. It was found that the supply of high purity oxygen could eliminate the oxygen limitation efficiently, and could enhance the cell densities and product levels much in the two model culture systems. No microbial metabolism shift except for the increase in cell density was expected due to the supply of high purity oxygen. Compared to the results found in fed-batch cultures of R. eutropha with air supply, the final dry cell weight (208.2 g/L) and PHB concentration (138.7g/L) obtained with high purity oxygen in the 5-L fermentor increased 2.2 and 2.9 times, respectively. In the 30-L fermentor, they increased 3.8 and 6.2 times, respectively. And only 8.4g/L of PHB was produced in the 300-L fermentor with air supply. This clearly showed the impact of oxygen supply on the high cell density culture, especially in the large-scale fermentation process. It was confirmed that a high $CO_2$ concentration could inhibit the cell growth and PHB synthesis in the culture of R. eutropha, even though it is one of autotrophic microbes and can use $CO_2$ as the main carbon source to produce PHB. We developed a new technique for studying $CO_2$ inhibition on the fermentation, “the natural or autogenou...