CN106434811A - Preparation method and application of cyclina sinensis immune-enhancing peptide - Google Patents
Preparation method and application of cyclina sinensis immune-enhancing peptide Download PDFInfo
- Publication number
- CN106434811A CN106434811A CN201611020866.4A CN201611020866A CN106434811A CN 106434811 A CN106434811 A CN 106434811A CN 201611020866 A CN201611020866 A CN 201611020866A CN 106434811 A CN106434811 A CN 106434811A
- Authority
- CN
- China
- Prior art keywords
- meretricis seu
- seu cyclinae
- concha meretricis
- weight portion
- preparation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/01—Hydrolysed proteins; Derivatives thereof
- A61K38/012—Hydrolysed proteins; Derivatives thereof from animals
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/30—Extraction; Separation; Purification by precipitation
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/34—Extraction; Separation; Purification by filtration, ultrafiltration or reverse osmosis
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/36—Extraction; Separation; Purification by a combination of two or more processes of different types
Abstract
The invention provides a preparation method and application of cyclina sinensis immune-enhancing peptide. The preparation method includes: preprocessing, to be more specific, removing the shells of cyclina sinensis, soaking cyclina sinensis meat into deionized water, homogenizing, performing ultrasonic treatment, and storing at low temperature; performing enzymolysis, to be more specific, adding a sodium hydroxide solution to regulate pH, adding compound enzymes with the weight being 2-10% of the weight of the cyclina sinensis meat to perform the enzymolysis, deactivating the enzymes, centrifuging, and taking supernate; performing membrane separation, to be more specific, adding a precipitating agent with the weight being 1-2% of the weight of the cyclina sinensis meat into the supernate, performing the membrane separation, and performing vacuum concentration, wherein the precipitating agent comprises, by weight, 3-5 parts of ammonium sulfate, 1-3 parts of sodium chloride, 0.5-1 part of potassium nitrate, 0.2-0.7 part of potassium dihydrogen phosphate and 0.1-0.3 part of resveratrol; performing freeze drying on concentrated liquor, and preserving to obtain the cyclina sinensis immune-enhancing peptide. The cyclina sinensis immune-enhancing peptide extracted by the method is high in concentration and extraction rate, high in activity, capable of being absorbed by human bodies favorably, and applicable to antitumor medicine and immune-enhancing medicine.
Description
Technical field
The invention belongs to technical field of biological extraction, and in particular to a kind of preparation method of Concha Meretricis Seu Cyclinae mucin peptide and should
With.
Background technology
Immunity is the defense function that body is attacked to pathogenic microorganism and the stable removing aging of environment in maintaining, dead
Die, injured cell and supervision, the function of mutant cell removed, when the low or obstacle of body's immunity, it will serious symptom occurs
Infection, tumor and human body senilism etc..With transformation of the people to disease treatment idea, the emphasis for the treatment of via direct killing outside
Borne causal agent turns to adjustment living organism itself function, causes extensive pass so as to application of the immunostimulant in medical science
Note, the research of immunostimulant has become as one of most active research field of medical science, finds natural immunomodulator, strengthens
Immunity of organism is the important and effective approach of current oncotherapy.
Concha Meretricis Seu Cyclinae is the common shellfish of China coast, belongs to high-quality protein original.Concha Meretricis Seu Cyclinae extract has non-oxidizability, antitumor
Function with enhance immunity.
Prior art provides the method that multiple biologies prepare mucin peptide, for example, Chinese invention granted patent document one
Plant the preparation method Authorization Notice No. CN 102952838 of immune-enhancing mussel enzymatic hydrolytic polypeptide and preparation method of corresponding tablet thereof
A kind of immunostimulant mussel enzymolysis polypeptide of B and its preparation method of respective tablets, step is：Using mussel dry powder as raw material, with
Buffer mixes according to 1: 20～1: 25 w/v, homogenate, then adjusts pH value to 9.5～10, obtains
Mixed liquor；Mixeding liquid temperature is risen to 50～55 DEG C preheat 10～15 minutes, according to 1.0%～1.5% point of raw material
Not Jia Ru alkaline protease and neutral protease, hydrolysis temperature be 50～55 DEG C, 2～4h of enzymolysis time；Enzyme by gained
Solution product obtains enzymolysis solution first through destroy the enzyme treatment, and the enzymolysis solution obtains immunostimulant enzyme successively through desalination, concentration and spray drying
Solution polypeptide；The invention also discloses the immunity enhancement type mussel polypeptide piece of correlation.The preparation method of the invention is simple, and obtained exempts from
Epidemic disease strengthens peptide and its tablet has higher activity, while raw material resources are abundant, with stronger industrial implementation.
The preparation method of prior art is simple, and the immunostimulant peptide activity for obtaining is higher, but only by alkaline protease and
Neutral protease is digested, and hydrolysis result is general, is had much room for improvement on the extraction ratio for extracting mucin peptide.
Content of the invention
The present invention provides one kind for above-mentioned technical problem and adopts ultrasonic assistant pretreatment, and hydrolysis result is excellent, has
A kind of preparation method and application of the high Concha Meretricis Seu Cyclinae mucin peptide of active immunostimulant peptides extraction rate.
The present invention by solving the scheme taken of above-mentioned technical problem is：A kind of preparation side of Concha Meretricis Seu Cyclinae mucin peptide
Method, preparation process is as follows：
1）Pretreatment：Concha Meretricis Seu Cyclinae is shelled take meat and be put in deionized water and soak, homogenate, ultrasonic Treatment, low-temp storage；By leaching
Bubble by impurity dissolving in Concha Meretricis Seu Cyclinae meat and kills antibacterial, harmless to Concha Meretricis Seu Cyclinae meat, does not affect to extract the activity of mucin peptide, with even
Pulp grinder by further for Concha Meretricis Seu Cyclinae meat tissue micronization processes, can lift the effect of follow-up enzymolysis using ultrasonic Treatment after being homogenized,
Low-temp storage ultrasonic Treatment rear slurry.
2）Enzymolysis：Add sodium hydroxide solution that pH value is adjusted, add the compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat to carry out enzyme
Solution, enzyme denaturing, centrifugation, take supernatant；Add sodium hydroxide solution quick regulation pH value to specified range, reach the pH needed for enzymolysis
Value, adds compound enzyme to be digested, and whole enzymolysis process is controlled, and the immunostimulant peptide content in enzymolysis solution is significantly improved, and
Prevent mucin peptide from losing activity, lift the mucin peptide effect for obtaining.Again through enzyme denaturing, supernatant is centrifuged to obtain.
3）Membrance separation：The precipitation agent of 1 ~ 2% weight portion of Concha Meretricis Seu Cyclinae meat being added in supernatant, carries out membrane separation, reduces pressure dense
Contracting；Add precipitation agent that presence is not digested sufficient protein and mucin peptide is separated out, to improve mucin peptide in supernatant
Content, improve Concha Meretricis Seu Cyclinae mucin peptide extraction ratio, through membrane separation obtain molecular weight for 3000 ~ 10000Da Concha Meretricis Seu Cyclinae
Mucin peptide liquid, the Concha Meretricis Seu Cyclinae mucin peptide closeness height of this scope, with higher activity, anti-tumor function and enhancing are exempted from
Epidemic disease power excellent effect, beneficial to absorption of human body.
4）Lyophilizing：Concentrated solution lyophilizing is preserved, using vacuum freeze drier, concentrated solution lyophilizing is preserved, obtain Concha Meretricis Seu Cyclinae immunity
Strengthen peptide.Concha Meretricis Seu Cyclinae mucin peptide effectively can be preserved for a long time, and Concha Meretricis Seu Cyclinae mucin peptide never degenerates, it is ensured that activity, loss amount
Few.
Precipitation agent is made up of following component and weight portion：3 ~ 5 parts of ammonium sulfate, 1 ~ 3 part of Sodium Chloride, 0.5 ~ 1 part of potassium nitrate, phosphorus
0.2 ~ 0.7 part of acid dihydride potassium and 0.1 ~ 0.3 part of resveratrol.Using compound enzyme can digest protein in Concha Meretricis Seu Cyclinae meat, enzymolysis
Excellent effect, there may be the protein not digested in enzymolysis process, for preventing such case, add precipitation agent to reach egg
White matter is sufficiently separated out the effect of Concha Meretricis Seu Cyclinae mucin peptide, lifts in supernatant the content of mucin peptide and intensive with this
Degree, is improved the extraction ratio of Concha Meretricis Seu Cyclinae mucin peptide.
In step 1, deionized water volume is 3 ~ 5 times of Concha Meretricis Seu Cyclinae meat, processes Concha Meretricis Seu Cyclinae meat slurry, ultrasonic power using ultrasonoscope
For 90 ~ 290W, ultrasonic time is to deposit 10 ~ 50min under the conditions of 2 ~ 6 DEG C.Refiner adopts super after being homogenized
Sonicated by further for Concha Meretricis Seu Cyclinae meat tissue micronization processes, can lift the effect of follow-up enzymolysis.
In step 1, deionized water is added with：3 ~ 7 weight portion of sodium citrate, 5 ~ 9 weight portion of Sodium Chloride, 8 ~ 12 weight of magnesium sulfate
Amount part, 2 ~ 4 weight portion of para-amino benzoic acid, 1 ~ 5 weight portion of 4-AA, 3 ~ 7 weight portion of disodium hydrogen phosphate, di(2-ethylhexyl)phosphate
1 ~ 5 weight portion of hydrogen potassium, 1 ~ 2 weight portion of sodium carboxymethyl cellulose and 0.3 ~ 0.7 weight portion of monoethyl amine.
The sodium hydroxide solution of 3.0 ~ 10.0mol/L is added to adjust pH value under the conditions of 35 ~ 45 DEG C to 6 ~ 8 in step 2.Reach
To the effect of quick regulation pH value, time-consuming, improve preparation efficiency.
In step 2, sodium hydroxide solution is added with 0.7 ~ 0.9 weight portion of phthalandione butyl benzyl ester, coordinates sodium hydroxide to adjust pH
Value, lifts pH value governing speed, saves the time, enzymolysis process is no affected, does not damage Concha Meretricis Seu Cyclinae protein.
The compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat is added to digest 1 ~ 3h under the conditions of 30 ~ 50 DEG C in step 2.Controlled enzymatic hydrolysis
Process, makes compound enzyme fully digest Concha Meretricis Seu Cyclinae protein, lifts in the enzymolysis solution that obtains Concha Meretricis Seu Cyclinae immunostimulant peptide content and intensive
Degree.
By enzymolysis solution boiling water bath enzyme denaturing in step 2, under the conditions of 90 ~ 95 DEG C, it is incubated 10 ~ 30min.Compound enzyme is avoided to continue
Enzymolysis and the activity to the follow-up mucin peptide for adding precipitation agent and acquisition, extraction ratio affects.
Compound enzyme is made up of following component and weight portion：10 ~ 15 parts of papain, 6 ~ 12 parts of neutral protease, alkalescence
2 ~ 4 parts of protease, 1 ~ 3 part of adenosinetriphosphataes, 2 ~ 3 parts of alteplase, 0.7 ~ 1.5 part of Cobra venom endonuclease.
It is centrifuged using centrifuge in step 2,8000 ~ 10000r/min of centrifugal rotational speed, 10 ~ 20min of centrifugation time, is gone
Except insoluble impurity.
In step 3, the film of membrane separation divides combined closure system for 3000 ~ 10000Da, obtains 3000 ~ 10000Da's of molecular weight
Concha Meretricis Seu Cyclinae mucin peptide liquid, in the Concha Meretricis Seu Cyclinae mucin peptide closeness height of this scope, with higher activity, anti-tumor function and
Enhancing immunity excellent effect, beneficial to absorption of human body.
The mucin peptide for extracting from Concha Meretricis Seu Cyclinae is applied in antitumor drug and medicament for immunity enhancement.
Beneficial effect：The present invention is by soaking impurity dissolving in Concha Meretricis Seu Cyclinae meat and killing antibacterial, harmless to Concha Meretricis Seu Cyclinae meat, no
The activity of mucin peptide is extracted in impact, can will be further for Concha Meretricis Seu Cyclinae meat tissue using ultrasonic Treatment after being homogenized with refiner
Micronization processes, the effect of the follow-up enzymolysis of lifting.Sodium hydroxide solution quick regulation pH value is added to specified range, time-consuming,
Compound enzyme is added to be digested, whole enzymolysis process is controlled, and the immunostimulant peptide content in enzymolysis solution is significantly improved, and is prevented
Mucin peptide loses activity, and lifts the mucin peptide effect for obtaining.Add precipitation agent that presence is not digested sufficient albumen
Matter separates out mucin peptide, to improve the content of mucin peptide in supernatant, improves the extraction ratio of Concha Meretricis Seu Cyclinae mucin peptide.This
Concha Meretricis Seu Cyclinae mucin peptide closeness and extraction ratio height that invention is extracted, with higher activity, anti-tumor function and enhancing immunity
Power excellent effect, beneficial to absorption of human body.
Specific embodiment
It is described in further detail below in conjunction with specific embodiment：
Embodiment 1：
A kind of preparation method of Concha Meretricis Seu Cyclinae mucin peptide, preparation process is as follows：
1）Pretreatment：Concha Meretricis Seu Cyclinae is shelled take meat and be put in deionized water and 10 ~ 12h is soaked, homogenate, ultrasonic Treatment, low-temp storage；
By soaking impurity dissolving in Concha Meretricis Seu Cyclinae meat and killing antibacterial, harmless to Concha Meretricis Seu Cyclinae meat, do not affect to extract the work of mucin peptide
Property, after being homogenized with refiner, further for Concha Meretricis Seu Cyclinae meat tissue micronization processes can be lifted by follow-up enzymolysis using ultrasonic Treatment
Effect, the activity of the stable ultrasonic Treatment rear slurry of low-temp storage.
2）Enzymolysis：Add sodium hydroxide solution that pH value is adjusted, add the compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat to carry out enzyme
Solution, enzyme denaturing, centrifugation, take supernatant；Add sodium hydroxide solution quick regulation pH value to specified range, reach the pH needed for enzymolysis
Value, adds compound enzyme to be digested, and whole enzymolysis process is controlled, and the immunostimulant peptide content in enzymolysis solution is significantly improved, and
Prevent mucin peptide from losing activity, lift the mucin peptide effect for obtaining.Again through enzyme denaturing, supernatant is centrifuged to obtain.
3）Membrance separation：The precipitation agent of 1 ~ 2% weight portion of Concha Meretricis Seu Cyclinae meat being added in supernatant, carries out membrane separation, reduces pressure dense
Contracting；Add precipitation agent that presence is not digested sufficient protein and mucin peptide is separated out, to improve mucin peptide in supernatant
Content, improve Concha Meretricis Seu Cyclinae mucin peptide extraction ratio, through membrane separation obtain molecular weight for 3000 ~ 10000Da Concha Meretricis Seu Cyclinae
Mucin peptide liquid, the Concha Meretricis Seu Cyclinae mucin peptide closeness height of this scope, with higher activity, anti-tumor function and enhancing are exempted from
Epidemic disease power excellent effect, beneficial to absorption of human body.
4）Lyophilizing：Concentrated solution lyophilizing is preserved, using vacuum freeze drier, concentrated solution lyophilizing is preserved, obtain Concha Meretricis Seu Cyclinae immunity
Strengthen peptide.Concha Meretricis Seu Cyclinae mucin peptide effectively can be preserved for a long time, and Concha Meretricis Seu Cyclinae mucin peptide never degenerates, it is ensured that activity, loss amount
Few.
Precipitation agent is made up of following component and weight portion：3 ~ 5 parts of ammonium sulfate, 1 ~ 3 part of Sodium Chloride, 0.5 ~ 1 part of potassium nitrate, phosphorus
0.2 ~ 0.7 part of acid dihydride potassium and 0.1 ~ 0.3 part of resveratrol.Using compound enzyme can digest protein in Concha Meretricis Seu Cyclinae meat, enzymolysis
Excellent effect, there may be the protein not digested in enzymolysis process, for preventing such case, add precipitation agent to reach egg
White matter is sufficiently separated out the effect of Concha Meretricis Seu Cyclinae mucin peptide, lifts in supernatant the content of mucin peptide and intensive with this
Degree, makes the extraction ratio of Concha Meretricis Seu Cyclinae mucin peptide be greatly improved.
In step 1, deionized water volume is 3 ~ 5 times of Concha Meretricis Seu Cyclinae meat, processes Concha Meretricis Seu Cyclinae meat slurry, ultrasonic power using ultrasonoscope
For 90 ~ 290W, ultrasonic time is to deposit 10 ~ 50min under the conditions of 2 ~ 6 DEG C.Refiner adopts super after being homogenized
Sonicated by further for Concha Meretricis Seu Cyclinae meat tissue micronization processes, can lift the effect of follow-up enzymolysis.
In step 1, deionized water is added with：3 ~ 7 weight portion of sodium citrate, 5 ~ 9 weight portion of Sodium Chloride, 8 ~ 12 weight of magnesium sulfate
Amount part, 2 ~ 4 weight portion of para-amino benzoic acid, 1 ~ 5 weight portion of 4-AA, 3 ~ 7 weight portion of disodium hydrogen phosphate, di(2-ethylhexyl)phosphate
1 ~ 5 weight portion of hydrogen potassium, 1 ~ 2 weight portion of sodium carboxymethyl cellulose and 0.3 ~ 0.7 weight portion of monoethyl amine.
The sodium hydroxide solution of 3.0 ~ 10.0mol/L is added to adjust pH value under the conditions of 35 ~ 45 DEG C to 6 ~ 8 in step 2.Reach
To the effect of quick regulation pH value, time-consuming, improve preparation efficiency.
In step 2, sodium hydroxide solution is added with 0.7 ~ 0.9 weight portion of phthalandione butyl benzyl ester, coordinates sodium hydroxide to adjust pH
Value, lifts pH value governing speed, saves the time, enzymolysis process is no affected, does not damage Concha Meretricis Seu Cyclinae protein.
The compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat is added to digest 1 ~ 3h under the conditions of 30 ~ 50 DEG C in step 2.Controlled enzymatic hydrolysis
Process, makes compound enzyme fully digest Concha Meretricis Seu Cyclinae protein, lifts in the enzymolysis solution that obtains Concha Meretricis Seu Cyclinae immunostimulant peptide content and intensive
Degree.
By enzymolysis solution boiling water bath enzyme denaturing in step 2, under the conditions of 90 ~ 95 DEG C, it is incubated 10 ~ 30min.Compound enzyme is avoided to continue
Enzymolysis and the activity to the follow-up mucin peptide for adding precipitation agent and acquisition, extraction ratio affects.
Compound enzyme is made up of following component and weight portion：10 ~ 15 parts of papain, 6 ~ 12 parts of neutral protease, alkalescence
2 ~ 4 parts of protease, 1 ~ 3 part of adenosinetriphosphataes, 2 ~ 3 parts of alteplase, 0.7 ~ 1.5 part of Cobra venom endonuclease.
It is centrifuged using centrifuge in step 2,8000 ~ 10000r/min of centrifugal rotational speed, 10 ~ 20min of centrifugation time, is gone
Except insoluble impurity.
In step 3, the film of membrane separation divides combined closure system for 3000 ~ 10000Da, obtains 3000 ~ 10000Da's of molecular weight
Concha Meretricis Seu Cyclinae mucin peptide liquid, in the Concha Meretricis Seu Cyclinae mucin peptide closeness height of this scope, with higher activity, anti-tumor function and
Enhancing immunity excellent effect, beneficial to absorption of human body.
The mucin peptide for extracting from Concha Meretricis Seu Cyclinae is applied in antitumor drug and medicament for immunity enhancement.
Embodiment 2：
A kind of preparation method of Concha Meretricis Seu Cyclinae mucin peptide, preferably preparation process is as follows：
1）Pretreatment：Concha Meretricis Seu Cyclinae is shelled take meat and be put in deionized water and preferably soak after 10h, deionized water volume is that Concha Meretricis Seu Cyclinae meat is excellent
5 times of choosing, are homogenized with refiner and are processed Concha Meretricis Seu Cyclinae meat slurry with ultrasonoscope again, and ultrasonic power is preferably 270W, when ultrasonic
Between be preferably 15min, deposit 30min preferably under the conditions of 4 DEG C.By soaking impurity dissolving in Concha Meretricis Seu Cyclinae meat and killing antibacterial,
Harmless to Concha Meretricis Seu Cyclinae meat, do not affect to extract the activity of mucin peptide, can using ultrasonic Treatment after being homogenized with refiner
By further for Concha Meretricis Seu Cyclinae meat tissue micronization processes, the effect of the follow-up enzymolysis of lifting, the stable ultrasonic Treatment rear slurry of low-temp storage
Activity.
2）Enzymolysis：The sodium hydroxide solution of preferred 8mol/L is added preferably to adjust pH value under the conditions of preferably 40 DEG C to 8,
Add the compound enzyme of preferred 7% weight portion of Concha Meretricis Seu Cyclinae meat 2h preferably to be digested under the conditions of preferably 45 DEG C, enzymolysis solution boiling water bath is gone out
Enzyme, is preferably incubated 20min, it is to avoid compound enzyme is persistently digested, and is centrifuged using centrifuge, preferably under the conditions of preferred 92 DEG C
Centrifugal rotational speed 9000r/min, preferably centrifugation time 15min, remove insoluble impurity, take supernatant；Add sodium hydroxide solution
Quick regulation pH value reaches the pH value needed for enzymolysis to designated value, adds compound enzyme to be digested, and whole enzymolysis process is controlled,
Immunostimulant peptide content in enzymolysis solution is significantly improved, and prevents mucin peptide from losing activity, the immunity increasing that lifting is obtained
Strong peptide effect.
3）Membrance separation：The precipitation agent of preferred 1.2% weight portion of Concha Meretricis Seu Cyclinae meat is added in supernatant, carries out membrane separation,
The film of membrane separation divides combined closure system for preferred 3000Da, obtains Concha Meretricis Seu Cyclinae mucin peptide liquid, concentrating under reduced pressure；Add precipitation agent
Sufficient protein precipitation mucin peptide is not digested existing, so that the content of mucin peptide in supernatant is improved, improve blue or green
The extraction ratio of clam mucin peptide, obtains Concha Meretricis Seu Cyclinae mucin peptide liquid through membrane separation, and closeness height, with higher work
Property, anti-tumor function and enhancing immunity excellent effect, beneficial to absorption of human body.
4）Lyophilizing：Concentrated solution lyophilizing is preserved, using vacuum freeze drier, concentrated solution lyophilizing is preserved, obtain Concha Meretricis Seu Cyclinae immunity
Strengthen peptide.Concha Meretricis Seu Cyclinae mucin peptide effectively can be preserved for a long time, and Concha Meretricis Seu Cyclinae mucin peptide never degenerates, it is ensured that activity, loss amount
Few.
Precipitation agent is made up of following component and weight portion：4 parts of preferably sulfuric acid ammonium, 2 parts of preferred Sodium Chloride, preferred potassium nitrate
0.7 part, 0.5 part of preferably phosphoric acid potassium dihydrogen and preferred 0.2 part of resveratrol.Using compound enzyme can digest the albumen in Concha Meretricis Seu Cyclinae meat
Matter, hydrolysis result is excellent, there may be the protein not digested in enzymolysis process, for preventing such case, adds precipitation agent
The effect of Concha Meretricis Seu Cyclinae mucin peptide is sufficiently separated out to reach protein, and the content of mucin peptide in supernatant is lifted with this
And closeness, it is improved the extraction ratio of Concha Meretricis Seu Cyclinae mucin peptide.
Deionized water is added with：5 weight portion of preferably citric acid sodium, 6 weight portion of preferred Sodium Chloride, 10 weight of preferably sulfuric acid magnesium
Part, 3 weight portion of preferred para-amino benzoic acid, 3 weight portion of preferred 4-AA, 4 weight portion of preferably phosphoric acid disodium hydrogen, excellent
Select 2 weight portion of potassium dihydrogen phosphate, 1.5 weight portion of preferably carboxymethyl cellulose sodium and 0.4 weight portion of preferred monoethyl amine.
Sodium hydroxide solution is added with preferred 0.75 weight portion of phthalandione butyl benzyl ester, coordinates sodium hydroxide to adjust pH value,
Lifting pH value governing speed, saves the time, enzymolysis process is no affected, and active immunostimulant peptides extraction rate is high, does not damage
Hinder Concha Meretricis Seu Cyclinae protein.
Compound enzyme is made up of following component and weight portion：It is preferred that 13 parts of papain, 8 parts of preferred neutral protease, excellent
Select 3 parts of alkaline protease, 2 parts of preferred adenosinetriphosphataes, 2 parts of preferred alteplase, 1.2 parts of preferred nucleic acid endonuclease.
The mucin peptide for extracting from Concha Meretricis Seu Cyclinae is applied in antitumor drug and medicament for immunity enhancement.
Above-described embodiment is the present invention preferably embodiment, but embodiments of the present invention are not by above-described embodiment
Limit, other any spirit without departing from the present invention and the change that is made under principle, modification, replacement, combine, simplify,
Equivalent substitute mode is all should be, is included in protection scope of the present invention.
Claims (9)
1. a kind of preparation method of Concha Meretricis Seu Cyclinae mucin peptide, it is characterised in that following preparation process：
1）Pretreatment：Concha Meretricis Seu Cyclinae is shelled take meat and be put in deionized water and soak, homogenate, ultrasonic Treatment, low-temp storage；
2）Enzymolysis：Add sodium hydroxide solution that pH value is adjusted, add the compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat to be digested, go out
Enzyme, centrifugation, take supernatant；
3）Membrance separation：In supernatant, the precipitation agent of 1 ~ 2% weight portion of addition Concha Meretricis Seu Cyclinae meat, membrane separation, concentrating under reduced pressure, obtain blue or green
Clam mucin peptide concentrated solution；
4）Lyophilizing：Concentrated solution lyophilizing is preserved, obtains Concha Meretricis Seu Cyclinae mucin peptide.
2. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：Described precipitation agent
It is made up of following component and weight portion：3 ~ 5 parts of ammonium sulfate, 1 ~ 3 part of Sodium Chloride, 0.5 ~ 1 part of potassium nitrate, potassium dihydrogen phosphate 0.2 ~
0.7 part and 0.1 ~ 0.3 part of resveratrol.
3. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：In the step 1
Deionized water volume is 3 ~ 5 times of Concha Meretricis Seu Cyclinae meat, processes Concha Meretricis Seu Cyclinae meat slurry using ultrasonoscope, and ultrasonic power is 90 ~ 290W, when ultrasonic
Between be 10 ~ 30min, deposit 10 ~ 50min after ultrasonic Treatment under the conditions of 2 ~ 6 DEG C.
4. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：In the step 1
Deionized water is added with：3 ~ 7 weight portion of sodium citrate, 5 ~ 9 weight portion of Sodium Chloride, 8 ~ 12 weight portion of magnesium sulfate, p-aminophenyl first
Sour 2 ~ 4 weight portions, 1 ~ 5 weight portion of 4-AA, 3 ~ 7 weight portion of disodium hydrogen phosphate, 1 ~ 5 weight portion of potassium dihydrogen phosphate,
1 ~ 2 weight portion of sodium carboxymethyl cellulose and 0.3 ~ 0.7 weight portion of monoethyl amine.
5. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：In the step 2
The sodium hydroxide solution of 3.0 ~ 10.0mol/L is added to adjust pH value under the conditions of 35 ~ 45 DEG C to 6 ~ 8；Hydroxide in the step 2
Sodium solution is added with 0.7 ~ 0.9 weight portion of phthalandione butyl benzyl ester.
6. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：In the step 2
The compound enzyme of 2 ~ 10% weight portion of Concha Meretricis Seu Cyclinae meat is added to digest 1 ~ 3h under the conditions of 30 ~ 50 DEG C；By enzymolysis solution boiling water in the step 2
Bath enzyme denaturing, is incubated 10 ~ 30min under the conditions of 90 ~ 95 DEG C.
7. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：Described compound enzyme
It is made up of following component and weight portion：10 ~ 15 parts of papain, 6 ~ 12 parts of neutral protease, 2 ~ 4 parts of alkaline protease, three
1 ~ 3 part of adenosine phosphate enzyme, 2 ~ 3 parts of alteplase, 0.7 ~ 1.5 part of Cobra venom endonuclease.
8. the preparation method of a kind of Concha Meretricis Seu Cyclinae mucin peptide according to claim 1, it is characterised in that：In the step 2
It is centrifuged using centrifuge, 8000 ~ 10000r/min of centrifugal rotational speed, 10 ~ 20min of centrifugation time；Membrance separation in the step 3
The film of process divides combined closure system for 3000 ~ 10000Da.
9. Concha Meretricis Seu Cyclinae mucin peptide according to claim 1 is applied in antitumor drug and medicament for immunity enhancement.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611020866.4A CN106434811A (en) | 2016-11-21 | 2016-11-21 | Preparation method and application of cyclina sinensis immune-enhancing peptide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611020866.4A CN106434811A (en) | 2016-11-21 | 2016-11-21 | Preparation method and application of cyclina sinensis immune-enhancing peptide |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106434811A true CN106434811A (en) | 2017-02-22 |
Family
ID=58221075
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201611020866.4A Pending CN106434811A (en) | 2016-11-21 | 2016-11-21 | Preparation method and application of cyclina sinensis immune-enhancing peptide |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106434811A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107007831A (en) * | 2017-06-16 | 2017-08-04 | 南京佰泰克生物技术有限公司 | The immunologic adjuvant of hepatitis B DNA vaccine |
| CN108339109A (en) * | 2018-01-17 | 2018-07-31 | 浙江海洋大学 | A kind of clam immunoloregulation polypeptide tablet and preparation method |
| CN108396046A (en) * | 2018-01-17 | 2018-08-14 | 浙江海洋大学 | A kind of preparation method of clam immunoloregulation polypeptide |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101736064A (en) * | 2010-01-20 | 2010-06-16 | 山东大学 | Process for preparing immune reinforcement peptide for enzymolysis on ocean shell and application thereof |
| CN104789626A (en) * | 2015-03-06 | 2015-07-22 | 浙江海洋学院 | Preparation method and application of Cyclina sinensis enzymolysis polypeptide |
| CN105821103A (en) * | 2015-12-30 | 2016-08-03 | 浙江海洋学院 | Method for preparing angiotensin-converting enzyme inhibitory peptide from clams |
-
2016
- 2016-11-21 CN CN201611020866.4A patent/CN106434811A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101736064A (en) * | 2010-01-20 | 2010-06-16 | 山东大学 | Process for preparing immune reinforcement peptide for enzymolysis on ocean shell and application thereof |
| CN104789626A (en) * | 2015-03-06 | 2015-07-22 | 浙江海洋学院 | Preparation method and application of Cyclina sinensis enzymolysis polypeptide |
| CN105821103A (en) * | 2015-12-30 | 2016-08-03 | 浙江海洋学院 | Method for preparing angiotensin-converting enzyme inhibitory peptide from clams |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107007831A (en) * | 2017-06-16 | 2017-08-04 | 南京佰泰克生物技术有限公司 | The immunologic adjuvant of hepatitis B DNA vaccine |
| CN108339109A (en) * | 2018-01-17 | 2018-07-31 | 浙江海洋大学 | A kind of clam immunoloregulation polypeptide tablet and preparation method |
| CN108396046A (en) * | 2018-01-17 | 2018-08-14 | 浙江海洋大学 | A kind of preparation method of clam immunoloregulation polypeptide |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104004813B (en) | A kind of preparation of mushroom biologically active peptide | |
| CN106434811A (en) | Preparation method and application of cyclina sinensis immune-enhancing peptide | |
| CN107868805B (en) | Longan polysaccharide degraded by lactobacillus fermentation and preparation method thereof | |
| CN110801012A (en) | Active polysaccharide compound nutrient for enhancing immunity and resisting fatigue and preparation method thereof | |
| CN103271951B (en) | Method for preparing cordyceps preparations with high adenosine contents | |
| CN106491662B (en) | A method of polysaccharide and flavones are extracted simultaneously from Phellinus cultivation waste material | |
| CN112999127B (en) | Gentiana scabra bunge compound enzyme and preparation method and application thereof | |
| CN106359839A (en) | Extraction method of oyster peptides | |
| CN105146045B (en) | Sea-fish protein prepares the method for combining blood pressure lowering peptide with fresh water fish protein | |
| CN108497118B (en) | Alcohol-dispelling health tea and preparation method thereof | |
| CN104762356B (en) | A kind of pumpkin leaf polyose and polypeptide and preparation method thereof | |
| CN101057659B (en) | Preparation method for black fungus blood fat-reducing products | |
| CN108208848A (en) | The preparation method of red algae protein polypeptide compound | |
| CN111662946A (en) | Method for preparing oligopeptide by enzymolysis of ganoderma lucidum water-soluble protein | |
| CN106834399A (en) | A kind of Antihypertensive Peptides from Trachyostracous mussel closed shell flesh | |
| CN106188329A (en) | The extracting method of a kind of scallop polysaccharide and goods | |
| CN114766678B (en) | Extraction method of roxburgh rose flavone, spirulina instant powder and preparation method thereof | |
| CN110835379A (en) | Process for extracting sparassis crispa polysaccharide by ultrahigh pressure auxiliary method | |
| CN106282281A (en) | A kind of method extracting small-molecular peptides from periplaneta americana | |
| CN106668867A (en) | Mumps vaccine freeze-drying protective additive free of gelatin and human albumin | |
| CN109907304A (en) | A kind of preparation method of Polysaccharide in Pleurotus eryngii | |
| CN102078335A (en) | Preparation method of latic acid hirudo liquid | |
| CN1261120C (en) | Lucid ganoderma extract for relieving or neutralizing alcohol effect and its extracting method and uses | |
| CN107793491A (en) | A kind of preparation method of squid ink polysaccharide oligosaccharides | |
| CN112375799A (en) | Method for improving oxidation resistance of abalone biological product |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20170222 |