Screening Of Cellulase-producing Bacteria In Liquor Pit Mud And Its Application In Liquor Fermentation

Liquor is mainly produced from corn and sorghum.In order to keep the water content and oxygen content of fermented grains,we need to add moderate rice husk and other accessories to loose fermented grains and expand fermentation interface.The cellulose contained in the raw material can not be used by yeast in directly.The cellulose is degraded into glucose by the bacteria which produced cellulase,then alcohol can be obtained from glucose by the metabolism of the yeast.This will not only save costs,improve utilization of raw materials,but also to reduce mash viscosity,shorten the fermentation cycle,improve liquor liquor yield.Therefore,the bacteria which produced cellulase has great potential in liquor production.In addition,the powerful aroma of liquor is mainly from the esterification flavored period,and the bacteria multiply rapidly in this period.So screening a kind of cellulase-producing bacteria which can not only produce cellulase but also has the role of liquor flavoring from the mud of the cellar have practical significance.In this research,cellar mud samples of fuyu old cellar spirit Co.,Ltd Heilongjiang Province were gathered by ecology methods.We used CMC-Na as a selective medium,the pure culture of cellulase-producing bacteria was obtained from the sample by classical microorganism separation and purification technology,the diversity of cellulase-producing bacteria from cellar mud were statistical comparative analysised by morphology,physiological and biochemical indexes.High yielding cellulose bacteria were screened by transparent circle and measuring enzyme activity.And then the functional strains were indentified to genus by molecule appraisal.Based on comprehensive analysis of other enzyme activity and aroma function,we obtained the desired strain and it was used in liquid liquor fermentation.The culture conditions were optimized in order to improve the production of cellulase activity,and the strain will initially be applied in liquid liquor fermentation.Through the analysis of the bacterial distribution that produced cellulase in the pit mud,we can know that bacteria and fungi are in the majority.Bacteria include Bacillus,Paenibacillus and Breuibacillus.Fungi include Aspergillus,Penicillium and Absidia.By molecular identification of fungal ITS sequences,AZ8 is Penicillium chrysogenum strain,AZ10 is Aspergillus sydowii.Based on the characteristic that bacteria can reproduce rapidly,short productive cycle and easy to culture,totally four strains with high cellulase yields were isolated.After 16SrDNA molecular identification,NA21 and NB29 are Bacillus subtilis,NA48 is Bacillus amyloliquefaciens,NC40 is Geobacillus stearothermophilus.Comprehensive strains producing the enzyme and production of other fragrant functional analysis to determine the purpose of the strain NB29.In this paper,NB29 is selected as the best strain after analysis of the enzyme activity and aroma function.The best culture condition was established by single factor analysis.The conditions were as follows:18 h of incubation time,140r/min of shaking spreed,37 C of culture temperature,4%inoculation.Through the response surface method,three main influence factors were determined,and the optimum concentration of these factors was:3.621g/100 mL glucose,0.183g/100 mL K2HPO4,0.062g/100 mL CaCl2.The cell concentration could reach up to 5.067×1010cfu/mL.The cellulase activity was doubled.NB29 was fermentated liquid in the sweet mashes of our lab.Liquor yield increased 6.13%and raw material utilization ratio increased 9.58%.After distilled into 40%(vol)liquor,the total acid amount was 0.876 g/L,the total ester content was 1.132g/L,and ethyl acetate content was 0.721 g/L.Liquor with flavored fragrance and pure mellow taste meted the secondary standard specifications of the state Fen-flavor liquor.