Separation And Application Of The Enzymes In The Solid State Fermented Matrix Of Pummelo Peel By Aspergillus Niger

As major catalysts in industry,enzymes were used in many area widely.Microorganisms could produce a variety of enzymes,so they are main resouce of enzymes.However,there are short of the technology to analyse fermented matrix,and because of complex culture components and microbial metabolic production,industry usually separates and purifies out one or two enzymes alone,while others are wasted.In this paper,the solid state fermented matrix of pummelo peel by Aspergillus niger was studied as the object.Proteomics technology was used to analyze the enzymes.The main enzymes in solid state media were separated thoroughly and identified by proteomics technology.Finally,enzymology properties and application properties of the major enzymes were studied,the main results were as follows:(1)Aspergillus niger 41034 as the strain of fermentation,submerged Czapek-Dox Medium with naringin and Czapek-Dox Medium as the control cultures,were identified 656 proteins totally by LTQ technology,such as glucosidase,glucanase,glucoamylase and mannosidase,etc.,these proteins were classified by function,and the result showed that A.niger produced carbohydrase,cell metabolic enzymes,lipid metabolic enzymes and oxidordeuctase.Narigin-added-Czapek-Dox Medium produced carbohydrase,cell metabolic enzymes and few oxidordeuctase while standard Czapek-Dox Medium produced only carbohydrase and cell metabolic enzymes.Most proteins were acidic protein,molecular weight between 20-135 kDa.(2)Aspergillus niger extracellular enzymes of solid state fermentation were separated by combined procedure consisting of DEAE-Sepharose F.F and HiTrap Blue HP,the major enzyme in four parts identified by proteomics technology,they wereα-L-rhamnosidase,β-glucosidase,α-L-arabinosidase,andβ-galactosidase.After separated,the enzyme activity were 1.2,737.2,146.9 and 15.5 U/mL,respectively.Specific activity were 4.7,5542.9,1514.3,234.4 U/mg respectively.(3)Enzymology properties and application properties ofα-L-rhamnosidase,β-glucosidase,α-L-arabinosidase,andβ-galactosidase were studied.The results showed that the optimal pH and temperature ofα-L-rhamnosidase was 4.0-5.0 and 50 ~oC respectively.α-L-rhamnosidase could hydrolyze naringin in citrus juices efficiently;β-glucosidase,optimal pH was 3.0,optimal temperature was 70 ~oC,this enzyme could hydrolyzed isoflavone to daidzein obviously;α-L-arabinosidase,optimal pH was 4.0 and temperature was 60 ~oC,the hydrolysis efficiency of this enzyme was equal to coorperated with xylanase;β-galactosidase,optimal pH was 3.0 and temperature at 70 ~oC,this enzyme could hydrolyze lactose to galactose efficiently.To sum up,this study set up a method to separate and applicate enzymes in solid state fermentation of pummelo peel by Aspergillus niger based proteomic technology.It might offer a reference to utilize the matrix enzymes in industry.