Optimization Of Pullulan Polysaccharide Fermentation By Aureobasidium Pullulans With Glucose As Carbon Source

Pullulan polysaccharide is an extracellular polysaccharide produced by Aureobasidium pullulan.As a new biopolymer,pullulan polysaccharide has many excellent physical and chemical properties and is widely used in food,environmental protection and other fields.At present,the industrial production of pullulan polysaccharide mostly uses sucrose as substrate for fermentation.However,organic sucrose is expensive as a raw material in industrial production.Therefore,it is of great significance to develop the pullulan fermentation process of non sucrose carbon source and provide organic production process for industrial production.In this paper,using Aureobasidium pullulans（CGMCCNo.7055）as the starting strain,the optimization of medium composition and culture conditions for pullulan production by Aureobasidium pullulans with glucose as carbon source were studied.In this paper,the optimal medium composition and culture conditions for high yield of pullulan polysaccharide were optimized by single factor and response surface methodology,and then the intracellular metabolites and transcription of pullulan polysaccharide produced by Aureobasidium pullulans under different ventilation conditions were analyzed by transcriptomics and metabolomics.The aim is to explore the biological mechanism of pullulan polysaccharide production through omics technology.The results are as follows:（1）The components of seed medium and culture conditions were optimized.The optimized medium components were（g/L）:glucose 75.0,yeast extract 2.0,（NH₄）₂SO₄0.5,K₂HPO₄3.0,Mg SO₄·7H₂O 0.8,Na Cl 3.0,Fe SO₄·7H₂O 0.06.The final culture conditions were:initial p H 7.0.（2）Through single factor and response surface experiments,the optimum medium composition and culture conditions for pullulan polysaccharide synthesis were as follows:glucose 132.0 g/L,yeast extract 7.2 g/L,Mg SO₄·7H₂O 1.4 g/L,K₂HPO₄6.9 g/L,Na Cl 7.0g/L and initial p H raw,seed age 24 h,inoculation amount 3%（V/V）.The yield of pullulan polysaccharide after optimization reached 77.97±1.56 g/l,which was 70.50%higher than that before optimization.（3）The fermentation conditions of 5 L fermentor were determined as follows:the initial p H was original,the p H was not regulated in the fermentation process,the ventilation ratio was 1:1.4（V/V）,and the stirring speed was 400 r/min.（4）Based on GC-MS technology,the effects of different ventilation on the changes of intracellular metabolites of pullulan polysaccharide synthesized by Aureobasidium pullulans during fermentation were studied.A total of 45 kinds of intracellular small molecule metabolites were detected,mainly including amino acids,organic acids,sugars,alcohols and some other metabolites.The metabolic pathway of pullulan combined with KEGG analysis mainly involves pentose glucuronic acid conversion pathway,fructose mannose metabolic pathway,citric acid cycle pathway and galactose metabolic pathway.（5）In this paper,the effects of different ventilation conditions on gene expression in the process of pullulan polysaccharide synthesis were studied by transcriptome technology.It was found that 693 segments of up-regulated genes and 598 segments of down-regulated genes were screened out at 36 h of fermentation,with a total of 1291 segments.The gene was up-regulated in segment 669 and down regulated in segment 811.The related differentially expressed genes had certain effects on glycolysis,citric acid metabolism,galactose metabolism,fructose mannose metabolism,inositol phosphate metabolism and propionic acid metabolism.