Inhibitory effect of carbon dioxide on the fed‐batch culture of Ralstonia eutropha: Evaluation by CO₂ pulse injection and autogenous CO₂ methods

Abstract

In order to see the effect of CO(2) inhibition resulting from the use of pure oxygen, we carried out a comparative fed-batch culture study of polyhydroxybutyric acid (PHB) production by Ralstonia eutropha using air and pure oxygen in 5-L, 30-L, and 300-L fermentors. The final PHB concentrations obtained with pure O(2) were 138.7 g/L in the 5-L fermentor and 131.3 g/L in the 30-L fermentor, which increased 2.9 and 6.2 times, respectively, as compared to those obtained with air. In the 300-L fermentor, the fed-batch culture with air yielded only 8.4 g/L PHB. However, the maximal CO(2) concentrations in the 5-L fermentor increased significantly from 4.1% (air) to 15.0% (pure O(2)), while it was only 1.6% in the 30-L fermentor with air, but reached 14.2% in the case of pure O(2). We used two different experimental methods for evaluating CO(2) inhibition: CO(2) pulse injection and autogenous CO(2) methods. A 10 or 22% (v/v) CO(2) pulse with a duration of 3 or 6 h was introduced in a pure-oxygen culture of R. eutropha to investigate how CO(2) affects the synthesis of biomass and PHB. CO(2) inhibited the cell growth and PHB synthesis significantly. The inhibitory effect became stronger with the increase of the CO(2) concentration and pulse duration. The new proposed autogenous CO(2) method makes it possible to place microbial cells under different CO(2) level environments by varying the gas flow rate. Introduction of O(2) gas at a low flow rate of 0.42 vvm resulted in an increase of CO(2) concentration to 30.2% in the exit gas. The final PHB of 97.2 g/L was obtained, which corresponded to 70% of the PHB production at 1.0 vvm O(2) flow rate. This new method measures the inhibitory effect of CO(2) produced autogenously by cells through the entire fermentation process and can avoid the overestimation of CO(2) inhibition without introducing artificial CO(2) into the fermentor.

Ho Nam Chang Special Issue: Life of a great biochemical engineer and his life-time contribution to high cell density culture

“…Each HCDC unit is equipped with cell recycling device of upflow packed-bed biofilter that might capture 95% of the cells with minimum pressure drop. Pure oxygen is generated from pressure swing adsorption method and regenerated by removing CO 2 from fermentation off-gas [17][18][19].…”

Low-cost noninvasive optical CO2 sensing system for fermentation and cell culture

“…Monitoring of carbon dioxide in bioprocesses is of importance because it affects microbial growth in various ways (Dixon and Kell, 1989;Kato and Tanaka, 1998;Shang et al, 2003;Sparringa et al, 2002). It is a product of cell respiration and can pass through cell membranes to influence the pH inside cells.…”

Production of biomass and recombinant human-like collagen in Escherichia coli processes with different CO2 pulses

“…Meanwhile, the utilization of pure O 2 and pressurized culture may dramatically increase CO 2 concentration in the exit gas, thus increasing dissolved CO 2 concentration (Lee 1996;Shang et al 2003;Castan et al 2002). However, the reported CO 2 effects depended on CO 2 concentration and the organism employed (Shang et al 2003); therefore, the effects of CO 2 should be examined in each specific fermentation system.…”

“…CO 2 inhibition or stimulation is likely to be most apparent in large fermentors (i.e., industrial-scale fermentors) in which increased hydrostatic pressure near the base of vessels results in increased dissolved CO 2 concentrations (Onken and Liefke 1989). Meanwhile, the utilization of pure O 2 and pressurized culture may dramatically increase CO 2 concentration in the exit gas, thus increasing dissolved CO 2 concentration (Lee 1996;Shang et al 2003;Castan et al 2002). However, the reported CO 2 effects depended on CO 2 concentration and the organism employed (Shang et al 2003); therefore, the effects of CO 2 should be examined in each specific fermentation system.…”

“…In recent studies, one or more pulses of CO 2 -enriched gas, rather than the continuous CO 2 -enriched gassing of batch cultures from time zero, had been used in batch cultures to avoid an overestimation of CO 2 effects (Shang et al 2003;McIntyre and McNeil 1997). In fact, the effects of CO 2 on fermentation processes may differ with cell growth phase, due to different cell growth and metabolic characteristics.…”