Screening And Identification Of Bacillus Sp With Elastase Activity And Optimization Of Its Fermentation Condition

Elastase is a kind of protease that can specially degrade elastin, Which is draw fromanimal pancreas mainly. Because of short of animal pancreas, elastase has demand exceedssupply in medication, food, chemical industry and environmental protection et al areas.Sreening the strais that producing elalaste is become thecrosscut to settle the demand andsupply.In this study, start with screenedand breed the elastase-producing strains, improved.the strains which producing elastase activity by mutation, optimized fermentationparameter, and researched on the application characteristic. This paper main results asfollows:(1) Strains capable of producing extracellular elastase were isolated from the soil andwastewater samples that were near the slaughterhouse, milk factory and beer factory. Ofthem 209 strains produced elastase. Hold the 83 strains which has a higher ratio of hydrolyzeloop and the colony diameter(HC)on the elementary screening flat, And swaying vasescreened again, strain NO.SCEL01 has the highest elastase enzyme,it was up to 33U/mL.Combine individual shape on the flat, biochemical feature and the homologous sequence by16SrDNA, the strain was identified as Bacillus subtilis. This strain SCEL01 irradiated by UV,mutated by DES, single out one strains BEM01 has higher elastase activity and stability onproducing-elastase. BEM 01 elastase activity growth 36%,it was up to 45U/mL.(2) The best carbon and nitrogen of the Bacillus subilitis strain,the result wass glucoseand casein, and toween can stimulate the strain to produce elastase and develop. Optimizedthe fermentation culture medium proportion of glucose, casein and toween by theL₉(3³)orthogonall experiment, Glucose has themainly influence the strain producing-elastase,then was casein, and the last was toween, meanwhile, the results was glucose2%, casin2%,toween 0.01%, K₂HPO₄ 0.2%, MgSO₄ 0.01%were the best proportion of culture medium.With the fermentation parameter as follows: the culture medium initial pH was 7.5, 37℃,20mL/300mLbottle, inoculation age was 24h, inoculation was 3%, cultivate 36h. Through the above condition,the strains has highest elastase, it was up to 59U/mL, and growth 31%than have not optimized the fermentation parameter.(3) With the same activity, pancreaselastase and Bacillus subtilis elastase have higherhydrolyze activity than papain on casein, gelatin and albumin, also on pork, beef, hogskin,oxhide, the same result with the culture medium flats to inspect. Elastase can hydrolyzeelastin strongly, and papain can not hydrolyze elastin.