pH and base counterion affect succinate production in dual-phase Escherichia coli fermentations
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Succinate production was studied in Escherichia coli AFP111, which contains mutations in pyruvate formate lyase (pfl), lactate dehydrogenase (ldhA) and the phosphotransferase system glucosephosphotransferase enzyme II (ptsG). Two-phase fermentations using a defined medium at several controlled levels of pH were conducted in which an aerobic cell growth phase was followed by an anaerobic succinate production phase using 100% (v/v) CO2. A pH of 6.4 yielded the highest specific succinate productivity. A metabolic flux analysis at a pH of 6.4 using 13C-labeled glucose showed that 61% of the PEP partitioned to oxaloacetate and 39% partitioned to pyruvate, while 93% of the succinate was formed via the reductive arm of the TCA cycle. The flux distribution at a pH of 6.8 was also analyzed and was not significantly different compared to that at a pH of 6.4. Ca(OH)2 was superior to NaOH or KOH as the base for controlling the pH. By maintaining the pH at 6.4 using 25% (w/v) Ca(OH)2, the process achieved an average succinate productivity of 1.42 g/l h with a yield of 0.61 g/g.