CN103444979B - Method for lowering albumen allergenicity by immobilized enzyme hydrolysis - Google Patents
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Abstract
The invention relates to a method for lowering albumen allergenicity by immobilized enzyme hydrolysis, which comprises the following steps: immobilizing crosslinking proteinase with magnetic beads, hydrolyzing albumen with the immobilized proteinase, removing fishy smell from the hydrolysate by using activated carbon, carrying out vacuum concentration at 60-70 DEG C, and carrying out freeze-drying to obtain the albumen powder of which the water content is less than 5%. The albumen powder has low allergenicity, contains abundant peptides and amino acids required by the human body, can be easily absorbed by the human body, and does not have bad flavor; and the method has favorable popularization and application prospects, can be widely used in the fields of food, drugs and the like, and is especially suitable to be used as a food raw material for allergic patients.
Description
Technical field
The invention belongs to egg-products processing technique field, relate to the preparation method of low irritated albumen.
Background technology
Food anaphylaxis is a kind of untoward reaction that body produces some food, has now become the ubiquitous food-safety problem in the whole world.In food anaphylaxis reaction, egg be FAO/WHO assert cause one of 8 large group foods of mankind's allergy, occupy food anaphylaxis ranking list second.According to statistics, there are 0.5%～2.5% children in the whole world to egg allergy, shows as clinically the symptoms such as urticaria, asthma, stomachache and diarrhoea, is having a strong impact on the healthy of irritated infant and is growing up.
Up to now, egg allergy there is no specific short, and strictly avoiding the edible food containing egg is allergy patient's optimal selection.But egg is often added in numerous food, avoid edible egg very difficult completely; And concerning the low patient of egg degree of allergic reaction, neither optimum method.At present, mainly reduce egg allergen by working methods such as hydrolysis method, thermal treatment, glycosylation and fermentations, to reduce the generation of egg allergy.When Choice and process method reduces the allergenicity of allergenic foods, meanwhile, also should consider organoleptic quality and the nutritive value of food.In these working methods, hydrolysis is considered to reduce the effective means of allergenic foods sensitization.The fragment that anaphylactogen has an epi-position (linear epitope and conformational epitope) by proteolytic enzyme catalytic hydrolysis after, its sensitization can significantly reduce; Anaphylactogen hydrolysis also may produce the biologically active peptides of functions such as having immunomodulatory, anti-oxidant, hypotensive, and its processing characteristics can improve.But at present, still there are many key issues urgently to be resolved hurrily in enzymic hydrolysis anaphylactogen: in (1) traditional resolvase protolysate process, enzyme can produce from hydrolysis fragment, disturbs downstream analysis to detect; Meanwhile, hydrolysis reaction is difficult to control, and after hydrolysis reaction finishes, enzyme Separation and Recovery exists certain difficulty, and repeating utilization factor is low; (2) exposure of a large amount of hydrophobic groupings in hydrolytic process, makes hydrolyzate with bitter off-flavors.These problems are perplexing and are being hydrolyzed and are affecting its in the widespread use reducing aspect food anaphylaxis originality.
In the last few years, magnetic microsphere is applied to enzyme immobilization field as carrier becomes study hotspot, enzyme is fixed on magnetic microsphere, both improved the stability of enzyme, also have simultaneously easily separated, reuse and improve the advantages such as immobilized enzyme catalysis efficiency continuously, be conducive to realize serialization and the industrialization of production technique.In view of the problem existing in the harm of egg allergy and current enzymic hydrolysis anaphylactogen technique.The invention provides a kind of method that immobilized enzyme hydrolysis reduces albumen sensitization, described method comprises: first use magnetic bead immobilization crosslinking protein enzyme, then utilize immobilization proteinase hydrolysis albumen, hydrolyzed solution carries out vacuum concentration after the de-raw meat of charcoal absorption at 60-70 DEG C, and recycling lyophilize obtains the albumen powder that water content is less than 5%.Low according to the inventive method gained albumen powder sensitization, and contain a large amount of peptide classes and the amino acid of needed by human body, be very easily absorbed by the body, without bad flavor; There is good promotion and application prospect, can be used as widely allergy patient's food raw material.
Summary of the invention
Main purpose of the present invention is to provide a kind of immobilized enzyme hydrolysis to reduce the method for albumen sensitization.Hydrolysis egg-white powder of the present invention nutrition, good palatability, safe, sensitization is low.
For achieving the above object, the technical solution used in the present invention is as follows.
(1) preparation of biotinylation labelled protein enzyme: take a certain amount of vitamin H, dissolve with appropriate 0.02M PBS damping fluid (pH5.5), be that 1:3:3 ratio is mixed in vitamin H with 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide (EDC), N-hydroxy-succinamide (NHS) mol ratio, under room temperature, react 1.5 h; Be dissolved in and in phosphate buffered saline buffer, prepare protein enzyme solution with any one enzyme in papoid, trypsinase, Flavourzyme flavor protease, Alcalase Sumizyme MP, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, be adjusted to enzyme optimum pH, after at 4 DEG C, reaction is spent the night, remove unreacted vitamin H with the dialysis of PBS damping fluid, dialysis product adds glycerine to save backup at-20 DEG C.
(2) Streptavidin (SA) coupling magnetic bead: get a certain amount of carboxylated magnetic bead, successively with after dehydrated alcohol, NaOH, HCl and the washing of PBS damping fluid, with the resuspended magnetic bead of PBS damping fluid.Be that 1:3:3 adds respectively EDC and Sulfo-NHS by magnetic bead surfaces carboxyl and EDC, N-hydroxy thiosuccinimide (Sulfo-NHS) mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls.Magnetic bead reclaims by magnet stand, and with after PBS damping fluid washing three times, is resuspended in PBS damping fluid.Magnetic bead after activation adds containing 37 DEG C of reaction 2 h in the PBS damping fluid (pH8.0) of SA, and final reaction product adds thanomin room temperature to seal 2 h.Magnet stand reclaims magnetic bead, and PBS damping fluid washs after three times, is resuspended in containing in the PBS damping fluid (pH8.0) of 0.05% sodium azide, and 4 DEG C save backup.
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:1-1:4, reaction oscillation incubation 20 min at 37 DEG C.Carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme.
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, control temperature is 37-60 DEG C, pH value 6.0-8.0, then living with the ratio of protein substrate in enzyme is that 5000-30000 U/g protein adds immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value 6.0-8.0 of controlled enzymatic hydrolysis system, hydrolysis time is 2.0-4.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution.
(5) in the albumen enzymolysis solution of collecting in step (4), add 0.5-3.0% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min.
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%.
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
Advantage of the present invention is: 1, the present invention can significantly reduce the sensitization of albumen; 2, albumen powder provided by the invention, has absorption easy to digest, is of high nutritive value, and without bitter off-flavors and other bad flavor, has good promotion and application prospect.
Embodiment
The present invention will be described further by following specific examples.
Embodiment 1.
(1) preparation of biotinylation labelled protein enzyme: taking a certain amount of vitamin H, with appropriate 0.02M PBS dissolving, is that 1:3:3 ratio is mixed in vitamin H and EDC, NHS mol ratio, reacts 1.5 h under room temperature; Be dissolved in phosphate buffered saline buffer and prepare protein enzyme solution with papoid, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, adjust pH value to 6.0, after reaction is spent the night at 4 DEG C, dialyse and remove unreacted vitamin H with PBS damping fluid.
(2) Streptavidin (SA) coupling magnetic bead: get a certain amount of carboxylated magnetic bead, use successively after dehydrated alcohol, NaOH, HCl and PBS solution washing, with the resuspended magnetic bead of PBS.Be that 1:3:3 adds respectively EDC and Sulfo-NHS by magnetic bead surfaces carboxyl and EDC, Sulfo-NHS mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls.Magnetic bead reclaims by magnet stand, and with after PBS washing three times, is resuspended in PBS.Magnetic bead after activation adds containing 37 DEG C of reaction 2 h in the PBS damping fluid (pH8.0) of SA, and final reaction product adds thanomin room temperature to seal 2 h.Magnet stand reclaims magnetic bead, and PBS washs after three times, is resuspended in the PBS(pH8.0 containing 0.05% sodium azide) in.
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:1, reaction oscillation incubation 20 min at 37 DEG C.Carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme.
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, controlling temperature is 50 DEG C, pH value 6.0, then live with the ratio of protein substrate in enzyme and be that 5000U/g protein adds immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value of controlled enzymatic hydrolysis system, hydrolysis time is 4.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution.
(5) in the albumen enzymolysis solution of collecting in step (4), add 0.5% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min.
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%.
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
Embodiment 2.
(1) preparation of biotinylation labelled protein enzyme: take a certain amount of vitamin H, with appropriate 0.02M PBS(pH5.5) dissolve, be that 1:3:3 ratio is mixed in vitamin H and EDC, NHS mol ratio, under room temperature, react 1.5 h; Be dissolved in phosphate buffered saline buffer and prepare protein enzyme solution with trypsinase, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, adjust pH value to 8.0, after reaction is spent the night at 4 DEG C, dialyse and remove unreacted vitamin H with PBS damping fluid.
(2) Streptavidin (SA) coupling magnetic bead: get a certain amount of carboxylated magnetic bead, use successively after dehydrated alcohol, NaOH, HCl and PBS solution washing, with the resuspended magnetic bead of PBS.Be that 1:3:3 adds respectively EDC and Sulfo-NHS by magnetic bead surfaces carboxyl and EDC, Sulfo-NHS mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls.Magnetic bead reclaims by magnet stand, and with after PBS washing three times, is resuspended in PBS.Magnetic bead after activation adds containing 37 DEG C of reaction 2 h in the PBS damping fluid (pH8.0) of SA, and final reaction product adds thanomin room temperature to seal 2 h.Magnet stand reclaims magnetic bead, and PBS washs after three times, is resuspended in the PBS(pH8.0 containing 0.05% sodium azide) in, 4 DEG C save backup.
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:2, reaction oscillation incubation 20 min at 37 DEG C.Carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme.
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, controlling temperature is 37 DEG C, pH value 8.0, then live with the ratio of protein substrate in enzyme and be that 10000 U/g protein add immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value of controlled enzymatic hydrolysis system, hydrolysis time is 4.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution.
(5) in the albumen enzymolysis solution of collecting in step (4), add 3.0% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min.
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%.
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
Embodiment 3.
(1) preparation of biotinylation labelled protein enzyme: take a certain amount of vitamin H, with appropriate 0.02M PBS(pH5.5) dissolve, be that 1:3:3 ratio is mixed in vitamin H and EDC, NHS mol ratio, under room temperature, react 1.5 h; Be dissolved in and in phosphate buffered saline buffer, prepare protein enzyme solution with Flavourzyme flavor protease, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, adjust pH value to 7.5, after reaction is spent the night at 4 DEG C, dialyse and remove unreacted vitamin H with PBS damping fluid.
(2) Streptavidin (SA) coupling magnetic bead: get a certain amount of carboxylated magnetic bead, use successively after dehydrated alcohol, NaOH, HCl and PBS solution washing, with the resuspended magnetic bead of PBS.Be that 1:3:3 adds respectively EDC and Sulfo-NHS by magnetic bead surfaces carboxyl and EDC, Sulfo-NHS mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls.Magnetic bead reclaims by magnet stand, and with after PBS washing three times, is resuspended in PBS.Magnetic bead after activation adds containing 37 DEG C of reaction 2 h in the PBS damping fluid (pH8.0) of SA, and final reaction product adds thanomin room temperature to seal 2 h.Magnet stand reclaims magnetic bead, and PBS washs after three times, is resuspended in the PBS(pH8.0 containing 0.05% sodium azide) in, 4 DEG C save backup.
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:3, reaction oscillation incubation 20 min at 37 DEG C.Carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme.
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, controlling temperature is 50 DEG C, pH value 7.5, then live with the ratio of protein substrate in enzyme and be that 20000 U/g protein add immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value of controlled enzymatic hydrolysis system, hydrolysis time is 3.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution.
(5) in the albumen enzymolysis solution of collecting in step (4), add 2.0% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min.
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%.
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
Embodiment 4.
(1) preparation of biotinylation labelled protein enzyme: take a certain amount of vitamin H, with appropriate 0.02M PBS(pH5.5) dissolve, be that 1:3:3 ratio is mixed in vitamin H and EDC, NHS mol ratio, under room temperature, react 1.5 h; Be dissolved in and in phosphate buffered saline buffer, prepare protein enzyme solution with Alcalase Sumizyme MP, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, adjust pH value to 8.0, after reaction is spent the night at 4 DEG C, dialyse and remove unreacted vitamin H with PBS damping fluid.
(2) Streptavidin (SA) coupling magnetic bead: get a certain amount of carboxylated magnetic bead, use successively after dehydrated alcohol, NaOH, HCl and PBS solution washing, with the resuspended magnetic bead of PBS.Be that 1:3:3 adds respectively EDC and Sulfo-NHS by magnetic bead surfaces carboxyl and EDC, Sulfo-NHS mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls.Magnetic bead reclaims by magnet stand, and with after PBS washing three times, is resuspended in PBS.Magnetic bead after activation adds containing 37 DEG C of reaction 2 h in the PBS damping fluid (pH8.0) of SA, and final reaction product adds thanomin room temperature to seal 2 h.Magnet stand reclaims magnetic bead, and PBS washs after three times, is resuspended in the PBS(pH8.0 containing 0.05% sodium azide) in.
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:4, reaction oscillation incubation 20 min at 37 DEG C.Carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme.
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, controlling temperature is 60 DEG C, pH value 8.0, then live with the ratio of protein substrate in enzyme and be that 30000 U/g protein add immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value of controlled enzymatic hydrolysis system, hydrolysis time is 2.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution.
(5) in the albumen enzymolysis solution of collecting in step (4), add 1.0% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min.
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%.
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
Claims (1)
1. immobilized enzyme hydrolysis reduces a method for albumen sensitization, it is characterized in that according to the following steps:
(1) preparation of biotinylation labelled protein enzyme: take a certain amount of vitamin H, with appropriate pH5.5, the dissolving of 0.02M PBS damping fluid, be that 1:3:3 ratio is mixed in vitamin H with 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide, N-hydroxy-succinamide mol ratio, under room temperature, react 1.5 h; Be dissolved in and in phosphate buffered saline buffer, prepare protein enzyme solution with any one enzyme in papoid, trypsinase, Flavourzyme flavor protease, Alcalase Sumizyme MP, the vitamin H of activation is mixed with proteolytic enzyme mol ratio 3:1 ratio, be adjusted to enzyme optimum pH, after at 4 DEG C, reaction is spent the night, remove unreacted vitamin H with the dialysis of PBS damping fluid, dialysis product adds glycerine to save backup at-20 DEG C;
(2) Streptavidin coupling magnetic bead: get a certain amount of carboxylated magnetic bead, successively with after dehydrated alcohol, NaOH, HCl and the washing of PBS damping fluid, with the resuspended magnetic bead of PBS damping fluid, be that 1:3:3 adds respectively 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide and N-hydroxy thiosuccinimide by magnetic bead surfaces carboxyl and 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide, N-hydroxy thiosuccinimide mol ratio, 37 DEG C of oscillatory reaction 2 h activation magnetic bead surfaces carboxyls; Magnetic bead reclaims by magnet stand, and with after PBS damping fluid washing three times, is resuspended in PBS damping fluid; Magnetic bead after activation adds 37 DEG C of reaction 2 h of PBS damping fluid containing Streptavidin pH8.0, and final reaction product adds thanomin room temperature to seal 2 h; Magnet stand reclaims magnetic bead, and PBS damping fluid washs after three times, is resuspended in the PBS damping fluid of the pH8.0 that contains 0.05% sodium azide, and 4 DEG C save backup;
(3) preparation of immobilized enzyme: immobilized enzyme utilizes Streptavidin MagneSphere and the preparation of biotinylated protein enzyme association reaction, in reaction system, the mol ratio of Streptavidin MagneSphere and biotinylated protein enzyme is 1:1-1:4, reaction oscillation incubation 20 min at 37 DEG C, carry out magnetic separation through magnetic frame, remove supernatant liquor, reclaim magnetic bead, obtain immobilized enzyme;
(4) albumen solution is mixed by a certain percentage with buffered soln, be transferred to afterwards hydrolytic decomposition pot, regulate the temperature of hydrolyzed solution, control temperature is 37-60 DEG C, pH value 6.0-8.0, then living with the ratio of protein substrate in enzyme is that 5000-30000 U/g protein adds immobilized enzyme, after mixing, in thermostat water bath, be hydrolyzed, constantly stir during this time, and the pH value 6.0-8.0 of controlled enzymatic hydrolysis system, hydrolysis time is 2.0-4.0 h, after reaction finishes, carries out magnetic separation through magnetic frame, collects albumen enzymolysis solution;
(5) in the albumen enzymolysis solution of collecting in step (4), add 0.5-3.0% food grade powdered carbon, hydrolyzed solution is placed in the water-bath of 50 DEG C and processes 60 min, then through the centrifugal acquisition supernatant liquor of 5000 r/min;
(6) be that 0.08 MPa, temperature are 50 DEG C and carry out vacuum concentration by the handled enzymolysis solution of step (5) in vacuum tightness, be concentrated into solid substance more than 60%;
(7) by step (6) gained egg white hydrolyzed peptide enriched material at-40 DEG C of pre-freeze 12 h, condenser temperature is-65 DEG C, vacuum tightness 100 Pa, 50 DEG C of temperature of heating plate, vacuum lyophilization 24 h, moisture content is below 5%, and dry rear gained powder is low sensitization albumen powder.
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| CN105018569B (en) * | 2015-07-16 | 2019-05-31 | 北京中科紫鑫科技有限责任公司 | The biotin labeling method of ATP sulfurylase |
| CN106978429B (en) * | 2017-03-16 | 2019-11-15 | 中国人民解放军第四军医大学 | A kind of bovine enterokinase light chain load magnetic bead and its preparation method and application |
| CN110498830A (en) * | 2018-05-16 | 2019-11-26 | 康码（上海）生物科技有限公司 | A kind of preparation method of the magnetic bead for separation and purification of protein |
| CN110144374B (en) * | 2019-05-30 | 2022-04-29 | 吉林大学 | Egg white peptide capable of promoting skin wound healing and preparation method thereof |
| CN110463925B (en) * | 2019-08-22 | 2022-11-04 | 华中农业大学 | Method for preparing high-foaming egg white liquid by immobilized protease |
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| EP0577162A1 (en) * | 1992-04-29 | 1994-01-05 | Genencor International, Inc. | Immobilized enzyme on a carrier of cross-linked gelatin and active carbon |
| CN1596676A (en) * | 2004-08-30 | 2005-03-23 | 江南大学 | Method of modifying lactoalbumin by enzymatic method and its application |
| CN101724675A (en) * | 2008-10-28 | 2010-06-09 | 陈栋梁 | Process for preparing albumin polypeptide |
| CN102429149A (en) * | 2011-11-17 | 2012-05-02 | 吉林大学 | Salted egg white protein polypeptide containing enteral nutrition and preparation method thereof |
| CN102994489A (en) * | 2012-10-09 | 2013-03-27 | 山西农业大学 | Biotin-avidin system immobilized glucoamylase and preparation method thereof |
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|---|---|---|---|---|
| EP0577162A1 (en) * | 1992-04-29 | 1994-01-05 | Genencor International, Inc. | Immobilized enzyme on a carrier of cross-linked gelatin and active carbon |
| CN1596676A (en) * | 2004-08-30 | 2005-03-23 | 江南大学 | Method of modifying lactoalbumin by enzymatic method and its application |
| CN101724675A (en) * | 2008-10-28 | 2010-06-09 | 陈栋梁 | Process for preparing albumin polypeptide |
| CN102429149A (en) * | 2011-11-17 | 2012-05-02 | 吉林大学 | Salted egg white protein polypeptide containing enteral nutrition and preparation method thereof |
| CN102994489A (en) * | 2012-10-09 | 2013-03-27 | 山西农业大学 | Biotin-avidin system immobilized glucoamylase and preparation method thereof |
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