Screening Of Cellulose Degradation Bacteria In Inner Mongolia Grasslands And It's Ethanol Fermentation From Corn Stalk

Utilization of cellulose without pollution is the main strategy to solve the shortage of energy.The biotransformation of cellulose has absorbed the attention of scientist in the field of energy,environment and chemical industries,and using cellulase to degrade cellulose efficiently and environmentally friendly to generate all kinds of energy required is the main way of cellulose biotransformation.So,how to obtain efficient cellulase is the key of the utilization of cellulose.This experiment uses CMC-Na selection medium to screen bacteria strains that could secrete cellulase from soil samples collected from Inner Mongolia.Then,the selected strains were stained with Congo red and cultured at different temperatures for enzyme activity determination.Among them,the strain S5 was screened out for it could produce cellulase at low temperatures.The strain was identified as Brevibacterium frigoritolerans.through morphological observation,physiological and biochemical verification and 16S rDNA gene sequence homology analysis.In order to optimize the enzyme production conditions of S5 strain,single factor optimization experiments were carried out on the carbon source,nitrogen source,inorganic salt,inoculum amount,temperature and pH of the enzyme production medium.The results showed that S5 produced cellulase under various conditions.At the same time,response surface method was used to optimize factors mentioned above.The final results showed that the enzyme amount was highest when the composition of fermentation was 3%corn straw,0.5%tryptone,0.5%tryptone,0.51%ferrous sulfate,2%inoculation amount.Fermentation pH was 6,and temperature was 30.59?.Under this condition,the cellulase activity of S5 was 24.78 U-mL-1,near to the expected value from response surface,and which is 179%of the enzyme activity of crude enzyme solution,which was 13.8 U·mL-1.In order to find out the optimum and stable temperature and pH of cellulase produced by S5 strain,the enzymatic activity was assayed under 20?,30?,40?,50?,60?,70?,80?and pH 3.0,4.0,5.0,6.0,7.0,8.0 respectively.The results showed that cellulase had the highest enzymatic activity under the reaction conditions of 50? and pH 4.0.At 45? and pH 4.0,cellulase had the best enzyme stability.This was in accordance with the result of optimum culture components and fermentation conditions in enzyme production.Finally,agricultural waste corn stalk powder was used to produce bioethanol.At first,the corn stalks were degraded into reducing sugars by S5 under the fermentation conditions optimized above.Then,Saccharomyces cerevisiae BY4741 was used to convert sugar into bioethanol,and the maximum concentration of ethanol was 0.47 g-L-1.So,it could be concluded that,in this study,a bacterium strain with the ability to degrade cellulose was found in the soil of Inner Mongolia,it was Brevibacterium frigoritolerans S5,which showed higher cellulase activity at lower temperature,and the enzyme activity of its cellulase reached to 24.78 U·mL-1 after the optimization of formation condition.Corn stalks could be converted into bioethanol by S5,together with Saccharomyces cerevisiae BY4741.The strain could be used to degrade cellulose and this study was benefit to solve the urgent problem of energy shortage.