Extraction Of Immunomodulatory Protein From Fruit Body And The Base Of Stipe Of Flammulina Velutipe

Immunomodulatory protein of Flammulina velutipe is a kind of protein extracted from Flammulina velutipe,which can enhance immunity,promote the organism of produce cytokines,promote cellular biosynthesis nucleic acids and proteins,and inhibit allergic reactions.The aim of this research is to investigate the extraction methord of FIP-fve from Flammulina velutipe,and the grade of purity of different part of Flammulina velutipe.The results of this research can provide method for effective use of Flammulina velutipe in practical application.In this study,we isolated FIP-fve by salting out method and ion exchange chromatography.We used hemagglutination test,SDS-polyacrylamide gel electrophoresis and gel filtration chromatography identify the properties of FIP-fve.We also studied the immunomodulatory protein level of of Flammulina velutipe in fruit body and the base of stipe.The main results were as follows:1)We used cell disruption,salting out method got 1.82g crude proteins from 500g Flammulina velutipe.Hemagglutination test showed the minimal active concentration was 25?g/mL.The extraction rate of crude proteins was 0.364%.2)We get 6 diffierent fractions by anion exchange chromatography which named F1-F6.The result of hemagglutination test of the 6 fractions showed that only F3 was positive and the minimal active concentration was 6.25 ?g/mL.The result indicating that F3 contain the components of FIP-fve.3).The result of SDS-polyacrylamide gel electrophoresis showed only one band of 13KDa.Corresponded with the relative molecular weight of immunomodulatory protein of Flammulina velutipes.4)The result of gel filtration chromatograph showed the molecular weight of F3 was 26KDa.It showed F3 is a kind of pure protein whith constitute by two monomers with the relative molecular weight of 13KDa.Experimental results fit the characteristics of FIP-fve.5)The F3 has effect of promoting red blood cell aggregating,the weight of subunit and native protein were 13KDa and 26KDa.It proved the F3 was FIP-fve.We got 18mg F3 from 500g the fruit body of Flammulina velutipe.The extraction rate of F3 was 0.0036%6)Use the same methods,we got 2.63g crude proteins from 500g the base of stipe of Flammulina velutipe.The extraction rate of crude proteins was 0.526%.Compared with fruit body of Flammulina velutipe increased 44.51%.7)We get 6 diffierent fractions(F1-F6)by anion exchange chromatography from the base of stipe of Flammulina velutipe.The result was the same as the fruit body of Flammulina velutipe.The result of hemagglutination test of the 6 fractions showed that only F3 was positive and the minimal active concentration was 3.13 ?g/mL.We got 29mg F3 from 500g the base of stipe of Flammulina velutipe.The extraction rate of F3 was 0.0058%.Compared with fruit body of Flammulina velutipe increased 61.1%.