| Propionic acid is widely used in food and dairy industries. As a result of its antimicrobial activity, propionic acid and its salts are widely used as preservatives in foods and grains. Currently, the market of propionic acid is mainly supplied by production via petrochemical routes. Fermentation by propionibacteria produces mainly propionic and acetic acids from sugars; however, the fermentation suffers from low propionic acid production due to by-product formation and strong propionic acid inhibition on cell growth and the fermentation. The high demand of propionic acid for use as a natural preservative in foods and grains has stimulated developments of new fermentation processes to achieve improved propionic acid production from low-cost biomass and food processing wastes. In this research, novel approaches, at process engineering, metabolic engineering, and genetic engineering levels, were developed for enhanced propionic acid production by Propionibacterium acidipropionici.; Fed-batch fermentation of glucose by P. acidipropionici immobilized in a fibrous-bed bioreactor (FBB) with a high cell density (>45 g/L) produced a high final propionic acid concentration of 72 g/L and a high propionate yield of up to 0.65 g/g. A mutant with improved propionate tolerance was obtained by adaptation in the FBB, which resulted in significant physiological and morphological changes. The mutant cells were less sensitive to propionate inhibition and had a higher saturated fatty acid content in the cell membrane and a slimmer shape with an increased specific surface area.; Metabolic stoichiometric analysis was applied to quantitatively describe the global cellular mechanism in propionic acid fermentation. By feeding carbon sources with different oxidation states, different fermentation end-product compositions were obtained, indicating different controlling mechanisms involving various acid-forming enzymes with significant changes in their activities and overall protein expression pattern. In general, the metabolic pathway shifted toward more propionate formation with a more-reduced substrate.; Gene inactivation via gene disruption and integrational mutagenesis was used to knock out the acetate kinase (ack) gene with the goal of eliminating acetate formation and further enhancing propionic acid production by P. acidipropionici. Mutants were obtained by transforming the cells with a partial ack gene fragment, which was introduced either as a linear DNA fragment with a tetracycline resistance cassette within the partial ack gene or in a non-replicative integrational plasmid containing the tetracycline resistance cassette. The ack inactivation in the mutants showed a profound impact on cell growth rate. Compared to the wild type, the ack-deleted mutants achieved ∼10% increase in propionate yield and ∼10% decrease in acetate yield. (Abstract shortened by UMI.)... |
