Continuous cell recycle succinic acid fermentation by Escherichia coli KJ 122

Abstract

The effectiveness of the genetically modified E. coli strain KJ 122 with regard to succinic acid (SA) production was evaluated under high cell density fermentation conditions in a continuous cell recycle reactor equipped with a hollow fibre filter. Batch fermentations were performed in a standard 1,5 L bioreactor for the purposes of comparison and to investigate the productivity, yield and titre that could be obtained. Continuous cell recycle fermentation led to a significant increase in volumetric productivity when compared with batch fermentations, albeit at a lower SA titre. The highest continuous volumetric productivity of 3 g/L/h was achieved at the highest dilution rate (0.15 h-1), at an SA titre of 19 g/L, which was five times higher than the overall batch productivity. The batch fermentations did, however, reach a final SA titre of 56 g/L. Unfortunately, severe product inhibition, at SA concentrations above 25 g/L, makes continuous production at high titres unfeasible and limits the cellular concentration in the fermenter due to cell death and subsequent cell lysis. Therefore, although temporary high dry cell weight was achieved, the biomass died off until an equilibrium was established between the cell growth, cell death and cells removed through the bleed stream. The SA yields obtained during batch fermentation (0.85 g/g glucose) were, however, superior to those obtained during continuous cell recycle fermentations (0.69 to 0.77 g/g). This was due mainly to the utilisation of pyruvate and formate at high SA titres during the latter part of the batch fermentations. The SA yield did, however, increase as the dilution rate increased, with the maximum yield (0.77 g/g) being obtained at a dilution rate of 0.15 h-1. Based on the metabolic flux analysis, this is believed to be due to an increase in pyruvate dehydrogenase activity at higher dilution rates. This increase led to a decrease in pyruvate and formate concentrations, and an increase in the flux through the reductive branch of the tricarboxylic acid cycle (due to the additional nicotinamide adenine dinucleotide produced). Low titres would increase the downstream processing requirements. To evaluate the economic feasibility of high cell density fermentation, the low titres and high productivities of continuous cell recycle fermentation would therefore have to be evaluated against the lower productivities and higher titres of batch fermentations.