Biological Degradation Of Soybean Allergens

Based on the purpose of degradation of α-subunit of β-conglycinin (Gly m Bd60K),casein plate selecting, soybean isolate protein re-screening and SDS-PAGE verification wasused for screening certain microorganism. And the production of protease by themicroorganism, the effect of its degradation and the functional properties of soybean isolateprotein (SPI) hydrolysate were further investigated. Moreover, strain identification andfermentation optimization were also studied. Main results were described as following:(1) A strain which can remove α-subunit and β-subunit of β-conglycinin was isolatedfrom soybean slurry, and then the selected strain was identified as Bacillus sp. BBE201108.The strains’ preservation number is CCTCC M2012165. The screening strategy includedcasein plate selecting, soybean isolate protein re-screening and SDS-PAGE verification.(2) Some enzymology character of protease produced by Bacillus sp. BBE201108anddesensitization conditions of soybean protein isolate (SPI) were investigated. And it wasfound that the optimal temperature is50oC, the optimal pH is8.0, the optimal proportion is7.5U Bacillus sp. BBE201108protease per milliliter of1%SPI solution, the optimal enzymeactivity assay method is Folin-determination using casein as substrate. Then, Polyclonalantibody in Rabbit resisted to SPI was prepared. The result showed that the polyclonalantibody can identify Gly m Bd60K and β-subunit of β-conglycinin. Moreover, the antibodyhas higher sensitivity in Gly m Bd60K, which can be identified even in500pg. Then,SDS-PAGE verification and Western blot verification of desensitized SPI treated with optimaldesensitization conditions were investigated. And it was found that both Gly m Bd60K andβ-subunit of β-conglycinin lose their allergenicity due to the degradation by Bacillus sp.BBE201108protease.(3) Partial functional properties of desensitized SPI were investigated. The result showedthat as compared to SPI, emulsifyingactivity and foaming capacity of desensitized SPI weredecreased, while emulsifyingactivity stability, foaming stability and gelation of desensitizedSPI were increased.(4) The wild bacteria Bacillus sp. BBE201108got a shake flask fermentationoptimization. Based on fermentation optimization of carbon resources, nitrogen resources andorthogonal experiment, the result showed that the optimum fermentation medium consisted2%glucose,2%soybean peptone,1%yeast extract,0.4%Na2HPO4·12H2O and0.06%KH2PO4. Moreover, the effects of primary pH, inoculation ages, inoculation volume, pouredsolution volume and temperature on protease production were also studied. Under the optimalconditions, protease activity reached the highest level at36h, up to a maximum of292.74U/mL.