Metabolic Flux Analysis of Simultaneous Production of Vitamin B12 and Propionic Acid in a Coupled Fermentation Process by Propionibacterium freudenreichii

Abstract

The metabolic processes involved in simultaneous production of vitamin B₁₂ and propionic acid by Propionibacterium freudenreichii are very complicated. To further investigate the regulatory mechanism of this metabolism, a simplified metabolic network was established. The effects of glucose feeding, propionic acid removal, and 5,6-dimethylbenzimidazole (DMB) addition on the metabolic flux distribution were investigated. The results showed that synthesis of propionic acid can be increased by increasing the metabolic flux through the oxaloacetate and methylmalonyl-CoA branches in the early and middle stages of the coupled fermentation. After DMB addition, the synthesis of vitamin B₁₂ was significantly enhanced via increased metabolic flux through the δ-aminolevulinate branch, which promoted the synthesis of uroporphyrinogen III, a precursor of vitamin B₁₂. Therefore, the analysis of metabolic flux at key nodes can provide theoretical guidance for the optimization of P. freudenreichii fermentation processes. In an experimental coupled fermentation process, the concentrations of vitamin B₁₂ and propionic acid reached 21.6 and 50.12 g/L respectively, increased by 105.71% and 73.91% compared with batch fermentation, which provides a new strategy for industrial production.

Keywords: Membrane separation coupled fermentation; Metabolic flux analysis; Propionibacterium freudenreichii; Propionic acid; Vitamin B12.