Optimization of recombinant bacterial fermentations for pharmaceutical production
Baheri, Hamid Reza
Two computer programs were developed and used to determine the optimum operating parameters of a fedbatch and a continuous two-stage process for fermentation of recombinant bacteria. The study was conducted in three phases: (a) developing two computer programs for simulation and optimization of the above processes, (b) conducting batch culture fermentations to verify the performance of the biokinetic model, and (c) conducting fedbatch and two-stage continuous fermentation experiments to closely examine the simulation and optimization results. The Miao and Kompala (1992) biokinetic model was used for simulation of the bacterial growth and cloned gene expression. The Pattern-Search method, developed by Hooke and Jeeves (1962), was incorporated in the programs to determine the optimum values of the parameters. Extensive studies of the optimization results showed 30-40% higher productivities for the two stage continuous process over the fedbatch process when using the same media in both processes. In addition, increasing the number of stages in the continuous two-stage process resulted in very limited improvement in the productivity of the process (10-12%). The information from the process optimization was then used to design batch, fedbatch nd two stage continuous experiments. Recombinant E. coli (strain BL21DE3) with an inducible gene (sensitive to IPTG, isopropyl-â-D-thiogalactopyranoside) was used throughout the experiments. The experimental results from the fedbatch and two stage continuous processes clearly showed good agreement with the simulation and optimization results $(\cong$15% deviation). The experiments also revealed that the maintenance of plasmid harboring cells over the long-term operation could be an important barrier in achieving the predicted high productivity in the two stage continuous process. Finally, in addition to computer programs for optimization of genetically modified microorganisms, a new computer program with a generic algorithm for optimization of multiple CFSTR fermentation with any kind of biokinetic model was developed. The program was used to optimize multiple CFSTRs with the cybernetic biokinetic model for the first time. Besides finding the optimum residence times for multiple CFSTRs operation, the effect of inaccuracies in different cybernetic model parameters on the overall productivity of the process was investigated. The simulation results illustrated that, a single CFSTR was more sensitive in its operation to inaccuracies in the biokinetic constants as compared to optimized CFSTRs in series (2-8 times more sensitive).
DegreeDoctor of Philosophy (Ph.D.)
SupervisorHill, Gordon A.
Copyright DateApril 1998
pattern search optimization
Continuous Flow Stirred Tank Reactor
recombinant bacterial fermentation
genetic engineering - computer simulation
two stage CFSTR optimization