Screening Of A High Pullulan-producing Non-pigmented Strain And Optimization Of Fermentation Conditions For Producing Pullulans

Pullulans is homopolysaccharide produced by Aureobasidium pullulans,which comprises of maltotriose by α-(1,6)-glucosidic bonds.Pullulans has lots of great feature,such as impermeable to oxygen,moisturizing,safe and so on,which has been applied in food,pharmaceutical,environmental protection and other companies.With the ever-increasing demands of Pullulans,the study of Pullulans is increased,too.This study includes separation and identification of Aureobasidium pullulans,strain breeding and improvement,which aimed to establish a systemic experiment method of breeding and fermentation of Aureobasidium pullulans.Results includes screening a high Pullulan-producing Non-pigmented strain;establishing fermentation process of pulluans.The main results were described as follows:1 、 One kind of fungus was separated from soil samples from grassland of Poyang lake national wetland park,whose yield of polysaccharide was 11.2 g/L and sugar conversion rate was 22.4%.The colony and cell morphology of fungus was observed,and the sequence of 18 S rDNA of it was analyzed.Then,the fungus was was identified as Aureobasidium pullulans and named as P23.The extracellular polysaccharide of strain P23 was verified as Pullulan by infrared spectrum(FT-IR).2、Strain P23 was compound mutagenized by DES、NTG、UV, then 13 mutant strains were obtained. One of the mutant, whose polysaccharide yield was 25.60g/L,sugar conversion rate was 51.20% up to more than twice rate of strain P23(22.4%)was named as P1012. Compared with strain P23, the cell morphology of strain P1012 was mainly yeast-like; the colony of strain P1012 was white,middle protuberant,and edge radial;the fermentation broth was ivory-white and the absorbance was 0.048(OD654nm).The fermentation stability of P1012 was good,then we chose strain P1012 as candidate strain.3、The fermentation condition and medium component of high yield production were determined by single factor experiments and orthogonal test.The constitute of optimum culture medium was:70 g/Lsucrose,3 g/Lrice bran and 3 g/L vitamin C.Theoptimal fermentation conditions was:fermentation temperature 28℃,initial pH 5.0,4%inoculum,and speed at 200 r/min.After 5-L fermenter culture,the yield of pullulan of strain P1012 was 43.77 g/L,and the sugar conversion rate was 62.53%.After fermentation,pH of the fermentation broth increased from 6.56 to 5.00,and the viscosity was 52.10 mpa.s.