Study On The Fermentation Process Of Coenyzme Q₁₀ With Agrobacterium Tumefaciens

Coenzyme Q₁₀ (CoQ₁₀, ubiquinone), widely presents in animals, plants and microorganisms, is an important electron transient mediator in the respiratory chain. Because of its multiple physiological activities, CoQ₁₀ has been greatly used in medicine, health products and cosmetics, etc. Among all of methods, producing CoQ₁₀ by microbial fermentation is considered as the most potential way in industry. The present study focus on controlling the quality of the strain to overcome the strain regression in previous research, and the fermentation conditions were optimized base on the obtained findings.Above all, study on the strain degeneration and yield reduction resulted from subculture of CoQ₁₀ producing strain illustrates that white colonies would appear in the fifth generation from the original green colonies, and its proportion increased with the subculture. Meanwhile, a variation of cell shape could be recognized in green colonies which hardly happened among white colonies. Besides, the numbers of recession-shaped cells are also different in colonies with various sizes and at various positions.A standard for fast screening of strains with high quality was determined by investigating product yield and strain stability. It was showed that the cells in centre of big and green colonies are the optimal strains. Simultaneously, the most effective method for keeping strain stability was to reduce the subculture times less than five, and perform strain isolation regularly to discard the regressed strains.A two stage fed-batch fermentation process was carried out in shake flask and the optimal conditions were as following: CaCO3 concentration of 5 g/L, glucose concentration between 5 and 15 g/L, adding 10% (v/v) fresh medium at 30, 48 and 66 h, respectively. In the initial 50 hours, temperature and agitating rate were maintained at 30oC and 200 r/min. After that, temperature and agitating rate were controlled at 34oC and 175 r/min. The results showed that a highest biomass of 15.1 g/L, CoQ₁₀ production of 228.2 mg/L and specific CoQ₁₀ content at 15.1 mg/g-DCW, increased by 79.4%, 241.2% and 90.3% respectively compared to batch fermentation, was obtained. The fermentation period was 76 h and productivity was 3.0 mg/(L·h).Subsequently, the yield and stability of strain was verified with the original optimized fermentation conditions in a 15 L fermentor. The highest yield of the strains screened by standard above was stable at 2,100 mg/L or more compared with previous results. Contemporarily, the two stage fed-batch culture system was scaled up. The results showed the fermentation period was 76 h and productivity was 32.9 mg/(L·h). Meanwhile, a highest biomass of 118.9 g/L, CoQ₁₀ production of 2,825.7 mg/L, specific CoQ₁₀ content at 23.8 mg/g-DCW, increased by 17.5%, 22.7% and 4.4% compared to that of the former strategy, were obtained.