Font Size: a A A

Study On Screening Of Higy-yield Neutral Proteiase Producing Aspergillus Oryzae And Fermentation Conditions

Posted on:2014-10-12Degree:MasterType:Thesis
Country:ChinaCandidate:Y L YuanFull Text:PDF
GTID:2250330401475032Subject:Microbiology
Abstract/Summary:PDF Full Text Request
The neutral protease is a protease whose optimum catalytic pH is6.5~7.5.It can catalyze thecleavage of peptide bonds in protein and is widely used in pharmacy, leather and food.A series of mutations and screenings for higher enzyme producers were performed using the ultraviolet (UV) and diethyl sulphate (DES) from parent strain Aspergillus oryzae H0. One high-yield neutralprotease-producing strain named H2-5was selected and enzyme activity reached2158U/g. This strain hada good genetic stability by inoculating10times.The mutation conditions: the distance is25cm,the time of irradiation is120s with15W ultraviolet lamp, the concentration of diethyl sulphate was2%and the time of irradiation is15min.The strainof H2-5was screened with high neutral protease. The neutral protease activity has improved108.5%thanthe parent strain.To determine the effect of the fermentation conditions and medium ingredinents on fermentationproduction, flask solid-state fermentation experiments were carried out. It was found that carbon source,nitrogen source, fermentation time, fermentation temperature, water ratio of the medium and pH havegreater effect and activator, inorganic salt also have effect. The optimal medium composition andfermentation conditions on fermentation were determined as: wheat bran as carbon source, soybean mealas nitrogen source, compounding ratio of the solid state and water5:4, pH7.5, tween-800.2%,as a result,high neutral protease activity was obtained after84h culture at30~32℃.With reference of single factor, we optimized fermentation conditions of Aspergillus oryzae byresponse surface methodology (RSM). The optimum concentrations obtained from RSM were13.42g ofwheat bran,3.45g of soybean meal and1.29(m/m) in ratio of material to water, respectively, resultingin maximum neutral protease activity of5561.76U/g, which was confirmed the validity of the establishedmodel.An experiment was conducted to study the effects of soybean fermentation using the neutralprotease on the optimal condition. We found that the concentration of free amino acids was more than30%.
Keywords/Search Tags:Aspergillus oryzae, neutral protease, mutation breeding, fermentation optimization
PDF Full Text Request
Related items