Research And Hydrolyzed Soy Protein Hydrolyzate Low Protease HDW7

Low-temperature protease HDW7is produced by a psychrotroph namedPseudomonas sp.W7.The protease,whose optimum temperature is37℃. Basesd ofthis,author’s laboratory has carried the experiments that apply low-temperature proteaseHDW7to hydrolyze animal proteins such as milk proteins and bovine proteins and soon.These experiments has acpuired some research findings better than mesophilicproteases.So,the study aims at hydrolyzing soyabean protein with low-temperatureprotease HDW7,to expand its applied fields further.In this paper,it was researched to hydrolyze soyabean protein with low-temperatureprotease HDW7.Firstly,we chose amount of enzyme,time,temperature,pH value andconcentration of substrate for analysis of single factor.Then,we optimized latter threefactors with response surface method(RSM),and finally achieved optimumtechnological condition,which hydrolyzes soyabean protein with low-temperatureprotease HDW7,which is amount of enzyme6000U/g,time3h,temperature37℃,pHvalue6.98and concentration of substrate4.09%.The hydrolyzing degree after optimalreaches to13.95%,slightly lower than two mesophilic proteases.But HDW7has distinctadvantage in saving energy and some applied fields strict with temperture,due to its lowwork temperture lower20℃than mesophilic proteases.On the base of achieving optimum technological condition,we researched thereaction kinetics. Firstly,we got Km=1.303g/L，Vmax=6.489mg/(L min) with themethod of Lineweaver-Burk,then got the Michaelis-Menten equation of hydrolyzingsoyabean protein with low-temperature protease HDW7under the optimumconditions,which is V=(6.489[S])/(1.303+[S]).Then we obtained the typical reactionkinetics equation under the same condition through mathematical deduction andnumerical fitting,which is DH=1/b·ln(1+abt)=6.061ln[1+(6.239e0/s0-0.011t)],and thismodel is quite consistent with the actual with quite low maximum deviation of7.42%.The enzymatic reaction rate constant k2is equal to37.81min-1,and enzymeinactivation constants K4is equal to6.714min-1,which can be used to describe andknow the properties of protease.In addition,we deduced and calculated the model equation of the main constituents level in hydrolysis system,which can describehydrolysis process in detail and be used as application of protease.We measured theactivation energy Ea=29.4kJ/mol of hydrolysis reaction,which indicates HDW7hasdistinct advantage in saving energy and catalyze hydrolysis reaction moreeasily,compared with mesophilic proteases.Our work researched the hydrolysis product further. We made sure that themolecular weights of hydrolysis product were less than10kDa by SDS-PAGE gelelectrophoresis. Then we separated and purificated the hydrolysis product bySephadex-50chromatography,and got two small molecular components SP1andSP2,whose proportions are17.86%and14.29%separately.The DPPH free radicalscavenging ability of SP2reaches to85.73%higher than SP1,which is obviously betterof mesophilic proteases.So,SP2deserves additional studies.