Control Strategies for High-Cell Density Cultivation of Escherichia coli

Abstract

Growth of microorganisms under conditions of substrate saturation in batch processes is often accompanied by substrate inhibition, by-product formation, or enduring adaptation. These problems can be overcome, in principle, by fed-batch or continuous cultivation, where the concentration of one substrate (in general the carbon source) is kept at a limiting level and, furthermore, where the growth rate can be kept at a desired value. Production systems with E. coli in batch culture suffer mainly from the formation of acetate, which is produced in response to oxygen limitation or excess carbon. The prevention of the accumulation of toxic levels of acetic acid is the main task for the achievement of high cell and high product concentrations in the bioreactor. The formation of acetate can be circumvented when cell growth occurs at reduced growth rates, thus preventing carbon overflow metabolism. The critical growth rate above for which acetate formation occurs is influenced by the growth medium, temperature, and, more general, by the overall physiological status of the cells. In addition, it is important, to allow cell growth at a constant growth rate to permit the synthesis of a product (e.g., a recombinant protein) of reproducible and defined quality. In continuous cultivation, a constant and non-critical growth rate with respect to acetate formation can be achieved, but problems with plasmid stability and, therefore, decreasing productivity may occur. So in many research and industrial applications, fed-batch processes for the production of recombinant proteins are employed. With an accurate choice of the process and control parameters, very high cell concentrations in the bioreactor can be achieved. The following will give a concise mathematical description of the related microbial reactions, of the reactor dynamics, and of feasible control strategies, which finally can be called a basic model for high-cell density cultivation (HCDC).