Selection of a Streptococcus equi mutant and optimization of culture conditions for the production of high molecular weight hyaluronic acid

A high molecular weight hyaluronic acid-producing mutant, designated as Streptococcus equi KFCC 10830, was derived from Streptococcus equi ATCC 6580 by the serial selection programs after NTG treatment. The selected mutant had nonhemolytic, hyaluronidase-negative, kanamycin-resistant, and highly viscous characteristics. Various culture conditions were optimized in a 5-l fermentor for the production of high molecular weight hyaluronic acid. When lysozyme was added to the culture broth during cultivation, the molecular weight of hyaluronic acid increased from 2.9 x 10(6) to 3.8 x 10(6). The optimum temperature and pH were 37 degrees C and 7.0, respectively. The molecular weight of hyaluronic acid at these conditions was 3.8 x 10(6). The effect of impeller type on the molecular weight of hyaluronic acid was also examined. When the intermig-type impeller was applied to the fermentor, the molecular weight of hyaluronic acid was 4.8 x 10(6). The molecular weight of hyaluronic acid increased as high as 5.0 x 10(6) when the agitation speed was increased up to 1.299 rpm. The highest molecular weight of hyaluronic acid, 4.8 x 10(6), was achieved at an aeration rate of 1.0 vvm. In addition, pilot-scale production of hyaluronic acid was successfully performed in a 100-l fermentor. Hyaluronic acid with a molecular weight of 3.2 x 10(6) was accumulated at a concentration of 6-7 g l(-1) in the culture broth. Accordingly, culture conditions, such as the addition of lysozyme, temperature, pH, impeller type, agitation speed, and aeration rate, were found to be major factors affecting the molecular weight of hyaluronic acid as well as its production.