Compound mutation breeding of Streptococcus equisimilis ZC-95 for producing high molecular weight hyaluronic acid and optimization of culture conditions

A strain of mutant S. equisimilis ZC-95, which can produce high molecular weight (HMW) hyaluronic acid (HA), was obtained from compound mutation breeding of S. equisimilis ZC-0 using 5-Bromouracil, UV rays and nitrosoguanidine as mutagenic agents. After inoculation for 20 generations, S. equisimilis ZC-95 showed inheritance stability, but not hemolysis. The culture conditions for producing HMW-HA were optimized in a 5-L fermentor through single-factor and multi-factor orthogonal tests. Results showed that cheap corn steep liquor was an appropriate nitrogen source for S. equisimilis ZC-95, and the optimum ratio of fermentation medium comprised 50 g/L sucrose, 80 g/L corn steep liquor, 10 g/L glucose, 2 g/L MgSO4, 2 g/L NaHCO3, 2 g/L Na2HPO4, and 0.1 g/L UTP. Under these fermentation conditions, the molecular weight of the produced HA was 2.76 × 106Da, and HA yield reached 4.12 g/L.