Screening Of Moderate Molecular Weight Hyaluronic Acid Producing Strain And Optimization Of Fermentation Process

Hyaluronic acid(hyaluronic acid,HA)is a kind of straight chain polymer alternately connected together by GlcA and GlcNAC by ?-1,3 and ?-1,4 glycosidic bond which has special water retaining effect.Hyaluronic acid has been widely used in food,cosmetics,medicine and other fields and has a huge market potential.It is found that the different molecular weight hyaluronic acid application performance are quite different: high molecular weight hyaluronic acid application in osteoarthritis treatment,operation defensive adhesion,ophthalmic surgery viscoelastic agent and drop eye liquid because of its good viscoelasticity and moisture,moderate molecular weight hyaluronic acid has advantages in compact skin and moisturizing properties,low molecular weight hyaluronic acid has better effect on anti-tumor,wound healing,angiogenesis,immune regulation.In this paper,the selection of fermentation strains,fermentation process optimization and separation and extraction process conditions to obtain the medium molecular weight HA.The main research contents and results are as follows:(1)Streptococcus zooepidemicus SL-2 which screened and preserved by the early stage was used as a starting strain,a good strain of medium molecular weight HA was obtained By ARTP system.it was identified as Streptococcus zooepidemicus XKW-104.(2)According to the single factor experiment and orthogonal experiment,the composition of the culture medium was determined:glucose 80 g/L,peptone 20 g/L,yeast powder 5 g/L,MgSO4·7H2O 1.5 g/L,K2HPO4·3H2O 2 g/L;culture condition for initial pH 7.0 medium volume 100 mL,inoculation amount of 10%(v/v),shaker speed 200 r/min,training temperature of 37 ?,the fermentation period is 24 h.(3)According to process optimization experiment,in 10 L automatic fermentor on the fermentation characteristics of strain XKW-104.Through control the amount of added glucose 80 g/L by initial glucose concentration of 40 g/L,6 h of fermentation one-time feeding glucose,to maintain the concentration of 40 g/L can make transparent qualitative acid yield increased by 28.5%(7.63±0.52 g/L),the accumulation of lactic acid decreased 7.89%,average molecular weight reached 1.56 x 106±0.45 u.The maximum yield of hyaluronic acid reached 9.66 + 0.58 g/L in 10 m3 fermentation tank,the rate of fermentation was 0.483±0.42 g/(L·h),the fermentation period was reduced to 20 h.(4)The method of extracting HA from two times alcohol precipitation was adopted.A fermentation liquor to precipitate crude best wine precision was 58%(v/v),optimal standing time of 10 h;solution decolorization activated carbon optimum adding amount of 0.8%(m/v);secondary precipitation best alcohol was 52%(v/v),the best standing time is 8 h;drying conditions for 50 ? drying under 5 h.