Effect of process parameters on succinic acid production in Escherichia coli W3110 and enzymes involved in the reductive tricarboxylic acid cycle

The effect of process optimization on succinic acid production by Escherichia coli W3110 and on enzymes involved in the reverse tricarboxylic acid cycle was studied. Approximately, 7.02 g L–1 of succinic acid was produced in 60 h at pH 7.0 in 500 mL anaerobic bottles containing 300 mL of the medium, wherein the sucrose concentration was 2.5%, the ratio of tryptone to ammonium hydrogen phosphate was 1:1, and the concentration of magnesium carbon ate was 1.5%. When these optimized fermentation conditions were employed in a 10 L bioreactor, 11.2 g L–1 of succinic acid was produced in 48 h. This is a 10-fold increase in succinic acid production from the initial titer of 0.94 g L–1. This clearly indicates the importance of process optimization, where by manipulating the media composition and production conditions, a remarkable increase in the production of the desired biomolecule can be obtained. The production of succinic acid is a multi-step reaction through the reverse tricarboxylic acid cycle. A linear relationship was observed between succinic acid production and the enzyme activities. The enzyme activities were found to increase in the order phospho-enol-pyruvate carboxylase < malate dehydrogenase < fumarase < fumarate reductase. The activity of phospho-enol-pyruvate carboxykinase was also estimated. Results indicate that this enzyme was not a very active participant in the production of succinic acid, since it catalyzes the phosphorylation of oxaloacetic acid to yield phospho-enol-pyruvate.Key words: anaerobic production, succinic acid, Escherichia coli, process optimization, reverse tricarboxylic acid cycle enzymes, fermentation.