Studies On The Extraction And Separation Of Neutral Protease From Bacillus Subtilis

In this thesis,the extraction and separation technology of the neutral protease fermentation broth produced by Jinan Nornoon Biological Engineering Co.,Ltd.were studied,and the study showed that food grade solid neutral protease can be industrialized.Firstly,the neutral protease acid-alkali stability and thermal stability were tested.Fermentation broth of neutral protease is stable at the pH4.80-5.20,with acetate-sodium acetate adjusted.Adding 0.2% potassium sorbate can inhibit the growth of stray bacteria in a certain period.And the adding 0.1% anhydrous calcium chloride can enhance the vitality and stability.With the company's existing fermentation apacity,the fermentation broth treatment selects the diaphragm filter press plate frame filter,with the filtration area of 150 ?,the filtration pressure 0.6MPa,3% corn starch as the filter aid,0.3% polyacrylamide as the flocculant filter cake,and the amount of top water into the cake keeping 10% of the volume of fermentation broth.The next treatment of filtrate of neutral protease chooses tubular ultrafiltration machine,the whole process ensure that the temperature is below 25?,and the interception molecular weight of pan film is 10000 Da,with pressure about 0.4MPa,the liquid neutral protease activity was about 6.7 times the initial enzyme activity after ultrafiltration,and the rate of yield was higher than 95%?In the process of salting out,ammonium sulfate was added to 25% of the quality of neutral protease concentrate,the cake enzyme activity is about 760 000u/g,salting out yield is higher than 95%;The method of adding water ultrafiltration is used to wash the desalination of ammonium sulfate in the filter cake,the liquid enzyme activity is about 230 000u/g after ammonium sulfate,and the solid content is 18.2%.The pressure spray drying tower is selected,the inlet temperature is 138 ?,the wind temperature is 76?,swirl slice 3.9,spray hole 1.5,feed speed is 200L/h.The obtained powder neutral protease 60 target quasi sieve passes rate 90%,the total yield is about 85%,enzyme activity 1 100 000u/g,and the enzyme activity is 73 times of fermentation broth.Target product hygiene indicators and physicochemical indicators are in line with food-grade enzyme preparation requirements.