Study On Enzymatic Preparation Of Heparin Sodium Critical Technolorigy

The purpose of the research was to provide data for improving extracting rate inheparin sodium production.In this experiment,took the porcine small intestine mucousmembrane as raw materials, effects of protease,adsorption and purification wereevaluated on their performances of hydrolyzates potency,adsorption quantity, heparinpotency, extraction rate and recovery.2709alkaline protease,papain,trypsin to the intestinal mucosa for enzymesolution, Respectively, single factor experiment were used to study the effoct ofenzyme substrate ratio,enzyme hydrosis temperature,enzyme hydrosis time,saltconcentration,pH and ratio of material to solution on the yield of small intestinemucous membrane, and the response surface methodology was used to optimize thetechnology parameters. Different types of ion exchange resin for enzyme hydrolysisadsorption,the effections of salt concentration,temperature,time,pH and mass fractionof resin to resin adsorption were researched with single factor experiment, andOrthogonal experimental design was used to optimize the technology conditions.Useof ethanol precipitation on the crude heparin sodium refining,signle factor experimentwere carried out to study the effect of temparature,time,salt concentration and alcohol,and orthogonal experimental design was used to optimize the technologyparameters.The results are as follows:1.Technology parameters of the intestine mucosa membrane hydrolysis（1）2709alkaline protease is the most suitable for hydrolysis conditions:enzyme substrate ratio0.80%,temperature55℃,time3h,pH8.8, salt concentration0.3mol/L, ratio of material1:7.（2）Papain is the most suitable for hydrolysis parameters: enzyme substrateratio0.80%,temperature60℃,time3h,pH8.8, salt concentration0.3mol/L, Materialliquid ratio1:7.（3）Tyrpsin is the most suitable for hydrolysis conditions: enzyme substrateratio0.70%,temperature60℃,time3h,pH8.5, salt concentration0.3mol/L, Materialliquid ratio1:7.2709alkaline protease,papain,trypain,respectively,in optimal enzymatic conditions of mucous,potency of enzyme hydrolysis is9.55U/mg,9.12U/mg,9.72U/mg.2. Technology conditions of the ion exchange resin adsorbed mucous hydrolysisThe most suitable resin exchange adsorption conditions: resin type FPA98cL, saltconcentration0.3mol/L, temperature50℃, pH9.0, time5h, resin concentration3.2%.the FPA98cL exchange resin adsorbed mucous enzyme hydrolysis,heparinsodium adsorption capacity was218.47mg/g,potency to90U/mg.3. Ethanol precipitation refining crude heparin sodium technology parametersCrude heparin refined the most suitable conditions: temperature40℃, altconcentration0.35mol/L, time35min, Alcohol35°.The use of ethanol precipitation to crude heparin in the optimum conditions,thepotency of crude heparin in80U/mg refined to120U/mg,the recovery rate of98.7%.