Study On Optimization Of Fermentation Conditions, Strain Selection And Middle Substrates By Aureobasidium Pullulans

Pullulan, kind of natural and water solubility exopolysaccharides. It can be used in many fields because of unique physical properties. So a large number of pullulan are needed.This article started from the angle of the optimization of fermentation conditions to improve the production of pullulan. Response surface methodology(RSM) was been used to analyze the six factors. Firstly, the optimal range of temperature, culture time,agitation speed, initial p H, loaded liquid and inoculated volume. Then Plackett-Burmen experiment showed three significant factors, they were culture time, agitation speed and initial p H. The value of other three factors chose their the maximum value if they were positive effects, otherwise chose the minimum value. Box-Behnken experiments were do for the three significant factors.The optimum fermentation technological conditions were as follows: the culture temperature was 25 ℃; culture fermentation time was 5.5days; culture agitation speed was 240 r/min; initial p H was 6.6; loaded liquid was 30 m L and inoculum size was 2%(V/V). Under the fermentation technological conditions,the predicted maximal pullulan yield was 26.31 mg/m L which closed to the predicted value 26.68 mg/m L, it had increased by about 63.1 % compared with the previous condition 16.13 mg/m L.The second parts of experiment was started from strain induced. Mutation mode can be divided into single mutagenesis and composite mutagenesis, composite mutagenesis also can be divided into using of the same mutagen repeatedly and using two or more mutagen at the same time or staggered to use. In this experiment, UV and Li Cl has been used as mutagen respectively and commonly. UV induced can improve the produce of pullulan, and Li Cl induced can reduce melanin production. We chose composite mutagenesis to induced, and chose the FAS-01 strain as the final purpose because of the experiment purpose, it can improve the produce of pullulan and the fermented liquid was yellow.In the last part of this experiment, HPLC was used to study the middle substrateof fermentation process. The mobile phase is 70% acetonitrile, the chromatographic column temperature is 35 ℃, the flow rate is 1 m L/min, sample quantity is 20 μL.Glucose, fructose, sucrose can be separated very well under the condition of the chromatographic analysis. Then contents of three saccharides obtained from fermentation broth fermented different time are be studyed by HPLC. It shows that only two kinds of hexose glucose and fructose can be detected in the fermentation broth, and by this way the more accurate content of pullulan can be known.