Combined Mutagenizing And Optimizing Fermenting Conditions Of Aureobasidium Pullulans

Pullulan is an unbranched homopolysaccharide which consists of maltotriose units withα-(1,6)-glucosidic bonds. There is extensive applying prospect for Pullulan in the fields, such as environment polluted disposal and food industry. We discussed combined mutagenizing to Aureobasidium Pullulans and screening for mutant strain with low pigment content and high Pullulan yield, and also investigated the conditons of the fermentation and the growth curve of the excellent mutant strain.By using Aureobasidium Pullulans N3.837 as the original strain, the strain was cultured in liquid basal medium for 16-20 hours, then washed by physiological saline twice and suspended. Suspensions, radiated by microwave, were diluted and spread on solid basal medium plates and cultured. After 48-72h, we counted the colonies, compared with no microwave mutagenize colonies, calculated the lethal rate and drew the lethal rate curve. Then, 16 strains which were alive while the lethal rate reached or surpassed 95% were selected to ferment for screening. After measureing the concentration of total sugar and extropolysaccharide, mutant KL15, of which the extropolysaccharide yield was 17.45%, was obtained. The Pullulan yield of KL15 is nearly one times more than that of Aureobasidium Pullulans N3.837(8.87%).By adding DES and EMS severally to the suspensions of mutant KL15 as chemistry mutation, 32 strains were taken out from culture dishes where the lethal rate reached or surpassed 90% and deepfreezed at 4℃. After screening by fermenting these strains, we knowed that mutant KL22, of which the extropolysaccharide yield reaches 23.88%, has the lowest pigment content. Repeated fermentative experiments proved the character of KL22 is stabile.Then, single factor experiment was used to study the optimum fermentation condition by fermenting mutant KL22. Initial pH of fermentative medium was selected to be the single factor. The result of experiment showed that 6.6, uder which the extropolysaccharide yield of KL22 reaches 25.41%, is the most suitable initial pH. What's more, we know that a small deviation of initial pH will affect the Pullulan productivy of KL22 observably. At last, we fermented mutant KL22 for 75 hours to determine the growth curve. The extropolysaccharide yield, the color and pH of fermentative culture, the variation of the cells'morphological forms and density were surveyed and recorded every 5 hours. By analyzing the relationships between these variable factors, we submitted optimizational suggestions for the fermentative production of Pullulan in future. Meanwhile, it has been proved that when KL22 is cultivated nearly 65 hours, the highest extropolysaccharide yield appears(26.63%) . This fact shows that the Pullulan producting cycle of mutant KL22 is shower than that of Aureobasidium Pullulans N3.837, and will bring many benefits for the industrial production of Pullulan by shortening the fermentative cycle of Aureobasidium Pullulans.