Screening Strain And Optimization Of Fermentation And Extraction Technology For Clavulanic Acid

Based on the biosynthesis pathway and metabolic mechanism of Streptomycesclavuligerus producing clavulanic acid, rational selection was performed to obtain aclavulanic acid high-producing strain. Zr2002bs-r2029was the variant strain ofzr2002bs-001obtained through ultra-violet mutation combined with high concertration of10%glycerol selection, and60Co mutation combined with1.0%sodium sulbactainselection. The productivity of this strain reached4698mg/L, which was increased by38.24%compared with the parent strain zr2002bs-001.With the mutant strain zr2002bs-r2029as original strain, the seed culture technique inshake flask was optimized, including initial pH, temperature and seed age. Based on theabove research, the optimal initial pH of medium was7.0, the seed age was54h, and theoptimal temperature was25℃. By the research of fermentation medium, soybean mealwas the best nitrogen source for clavulanic acid fermentation, and the bean oil could beused for clavulanic acid fermentation instead of triolein. During the period of clavulanicacid fermentation, by feeding glycerol as precursor after60h of incubation, theproductivity of zr2002bs-r2029could be improved.Based on the culture technique in shake flask, the fermentation medium inpilot-plant-scale was optimized, and the better quantity of soybean meal, bean oil andglycerol was4.2%,1.0%and1.0%. The concentration of glycerol should be maintained1~3g/L after22h of incubation for clavulanic acid biosynthesis. Druing the fermentation,the temperature should be controlled28±2℃, the air flow rate was15±10%, the agitationrate was500rpm, pH was6.90~7.10, and the PMV was25~30%. During the wholefermentation process, when the dissolved oxygen was above20%, the clavulanic acidproductivity reached5800mg/L.The extraction process was optimized. The potassium clavulanate preparation couldbe carried out in water phase, the yield of each step was above80%, and the productquality was qualification. For improving the safety of extraction process, microcrystallinecellulose was added to co-crystalling and co-drying with potassium clavulanate, themixture was well mixed, and the particle size was small.