CN110042139A - A kind of preparation method and applications of animal blood activity complex peptides - Google Patents
A kind of preparation method and applications of animal blood activity complex peptides Download PDFInfo
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- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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- A23K20/00—Accessory food factors for animal feeding-stuffs
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- A61K8/00—Cosmetics or similar toiletry preparations
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- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
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- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/005—Preparations for sensitive skin
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- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
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- C07K1/34—Extraction; Separation; Purification by filtration, ultrafiltration or reverse osmosis
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- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
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- C12Y—ENZYMES
- C12Y115/00—Oxidoreductases acting on superoxide as acceptor (1.15)
- C12Y115/01—Oxidoreductases acting on superoxide as acceptor (1.15) with NAD or NADP as acceptor (1.15.1)
- C12Y115/01001—Superoxide dismutase (1.15.1.1)
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Abstract
A kind of preparation method of animal blood activity complex peptides, this method combine membrane separation technique and bio-orientation enzyme incision technology, can prepare the animal blood active peptide of high activity with effective protection animal blood serum albumen and polypeptide active.The method of the present invention eliminates chemistry in conventional animal blood polypeptide extraction process or high-temperature denaturation and spray drying, by the way of grading extraction, it not only takes the lead in proposing the active peptide A(blood plasma ultrafiltration peptide inherently contained in blood plasma with 5000Da ultrafiltration membrane in same process flow), and using the blood plasma after being concentrated by ultrafiltration as raw material direct enzyme cutting, ultrafiltration, reverse osmosis concentration has extracted plasma protein peptide B(plasma protein digestion peptide), the active polypeptide complexes type formed after the two is compound is more abundant comprehensive, and two class active peptides mixing after can complement each other, play bio-cooperation, compared with traditional handicraft extract relatively single animal blood polypeptide in medical treatment, beauty, the fields such as health care can play stronger effect.
Description
Technical field
The invention patent belongs to biomedicine field, and in particular to a kind of preparation method of animal blood activity complex peptides and its
Using.
Background technique
Biologically active peptide is made of amino acid with the different arrangement modes formed, from dipeptides to complicated linear, ring
The general name of the different peptides of shape structure.These peptides can be activated by phosphorylation, glycosylation acylation living and play physiology work
With.Peptide is the basic material for constituting all cells of human body, and in body, various active peptides are generated in respective gland cell
And secrete, amount is seldom, but peptide is related to each of update, metabolism, growth and the reparation of human hormone, nerve, reproduction and cell
A field.Peptide is the basis of life, and it is exactly because they can produce peptide in vivo why life, which exists,.Lack polypeptide cell just
Meeting aging loses activity, is even dead, it may be said that without peptide just without life.
Scientist has confirmed: generation, development, treatment, the rehabilitation of all diseases are all related with peptide.Biologically active peptide can
The ability of various diseases is resisted in activating immune system, enhancing, and active peptide can make one the tissue of the vitals such as body-centered, liver, lung, kidney
Cell is repaired and is regenerated, and the physiological function gradually degenerated is made to be restored to health status；Active peptide, which has hormone and adjusts, to swash
The effect of element has the function of adjusting and inhibition to the intracorporal enzyme of biology, can enhancing development；Active peptide can increase vascular wall
Elasticity, has antithrombotic, anti-hypertension, and norcholesterol inhibits bacterium, virus, antitumaous effect；Active peptide has anti-oxidant and clear
Except Free Radical, element absorption and minerals transport can be improved, the elasticity of skin is can increase, restore the pale, fine and smooth of skin
And gloss；Active peptide has facilitation to deep sleep, has good effect to chronic fatigue syndrome；Active peptide also has simultaneously
There is the ability for promoting wound healing.
Contain a large amount of active peptide in animal blood, they, all over whole body, are more than 250 kinds of human body with blood circulation of human body system fortune
Different cells provide cytotrophy agent, while wide participation and having adjusted the proliferation of human body cell, growth, metabolism, aging, death
Etc. vital movements.Clinically for calf serum de-protein injection by application many years, main composition is exactly serum peptide, it
It is widely used in treating the diseases such as all kinds of burn and scalds, bedsore, diabetes, this not only demonstrates the important function of serum active peptide
Effect effect, while the precedent of different animals serum peptide mutually is also further started, it is established for the safety of serum active clinical application
Solid foundation is determined.
People have been able to isolate from animal blood, plant and microorganism a variety of using the means digested in vitro at present
The biologically active peptide of multiplicity.There is the listing of a large amount of animal blood (deer, camel, ox, sheep blood) polypeptide powder, both at home and abroad with raw material shape
Formula is widely used food or field of health care products, for improving the abilities such as immunity of organisms, antifatigue, anti-ageing, but current animal
For the traditional extraction technique of serum polypeptide mostly using haemocyanin as raw material, crushing heat aging through albumen, proteolytic cleavage, ultrafiltration are dense
The techniques such as contracting, spray drying are prepared, this not only allows in serum the active peptide loss of activity contained originally, greatly reduces blood
The physiological activity of clear active peptide, while the various polypeptides of serum and proteolytic cleavage peptide also lose after high-temperature maturing and spray drying treatment
The physiological activity of peptide is lost.
Summary of the invention
The purpose of the present invention be exactly directed to it is above-mentioned there are problems, and on the basis of existing technology, a kind of animal of proposition
The preparation method of blood activity complex peptides can will utilize film under the premise of effective protection animal blood serum albumen and polypeptide active
Isolation technics and bio-orientation enzyme incision technology combine, and invent a kind of technique side that can prepare high activity animal blood active peptide
Method.
The purpose of the present invention is achieved through the following technical solutions:
(1) the healthy animal blood qualified through quarantine is acquired, tube centrifuge separation, collection blood plasma is spare, and blood cell can be used for making
Standby animal protein feed or the SOD of extraction, or the directly qualified fresh animal blood plasma of purchase quarantine；
(2) the tubular type filter filtration sterilization by blood plasma through 2 ~ 3 concatenated 0.22 micron pore sizes；
(3) by blood plasma after filtration sterilization through the ultrafiltration apparatus ultrafiltration that molecular weight is 5000Da of shutting off, permeate is collected respectively and is cut
Flow liquid is spare；
(4) by the permeate in step (3) through shut off molecular weight be 100-200Da reverse osmosis membrane be concentrated by ultrafiltration to permeate original
The 1/8-1/10 of volume collects shut off liquid and permeate respectively, and the liquid that shuts off is animal blood active peptides A(blood plasma ultrafiltration peptide)
(content of peptides >=8mg/ml, content of peptides can be used UV-absorptivity method in " Chinese Pharmacopoeia " to be detected), permeate can be made
It is recycled for industrial water (as being used as equipment water for cleaning)；
(5) liquid that shuts off in step (3) is put into reaction kettle, compound protease is added, each protease additional amount is 200-
600U/mL(is in terms of trapped fluid volume)), i.e., 4 hours or more are kept the temperature under the conditions of pH7.0-8.5,35-50 DEG C of temperature, obtained
Animal blood albumen enzymolysis liquid；
(6) by the enzymolysis liquid in step (5), the ultrafiltration apparatus ultrafiltration through the molecular weight 3000Da that shuts off, respectively collect shut off liquid and
Permeate continues to digest, recycle wherein the liquid that shuts off can be added in lower batch of material liquid step (5)；
(7) permeate in step (6) is concentrated by ultrafiltration through the reverse osmosis membrane for the molecular weight 100-200Da that shuts off to permeate substance
Long-pending 1/3-1/5, collection is shut off respectively and permeate, the liquid that shuts off are animal blood polypeptide B(plasma protein digestion peptide) (>=
20mg/ml), permeate can be used as industrial water (as equipment water for cleaning) and recycle；
(8) it will be mixed with the animal blood polypeptide in the animal blood polypeptide A and step (7) in batch step (4), as animal blood is living
Property complex peptides.
Compound protease described in step (5) can be trypsase, papain, alkali protease and neutral proteinase
In it is any two or more.
Extraction process involved by the present invention can also be used for its hetero-organization (such as spleen, pancreas, placenta) of animal be material into
The extraction process of row active peptides.
The animal blood refers to the blood of the mammals such as pig blood, deer blood, sheep blood, ox blood, camel blood.
The animal blood activity complex peptides prepared through the invention can be applied to many aspects such as medical treatment, beauty, health care.It is curing
Prescription face can be used for can be used in terms of the Regeneration and Repair of tissue, beauty the reparation of allergy skin quality, and body can be improved in health care
Immunity.
Compared with prior art, innovative point of the invention essentially consists in:
1. present invention process makes full use of UF membrane extractive technique and bio-orientation enzyme incision technology, conventional animal blood polypeptide is eliminated
Chemistry or high-temperature denaturation and spray drying in extraction process, entire process flow, which is established, is ensureing plasma protein and more
On the basis of the bioactivity of peptide, high activity animal blood active peptide truly has been extracted, has been existed for animal blood active peptide
Punching point, which plays a role, in body has established Basic of Biology.
2. active complex peptides extraction process involved by the present invention, by the way of grading extraction, in same process flow
It not only takes the lead in proposing the active peptide A inherently contained in blood plasma with 5000Da ultrafiltration membrane, but also with the blood after being concentrated by ultrafiltration
It starches and has gone out plasma protein peptide (fibrin former peptide, ovalbumin peptide, globulin peptide) for raw material direct enzyme cutting, ultrafiltration, concentration extraction
Isoreactivity peptide B, the active polypeptide complexes formed after the two is compound, not only active peptide type is more abundant comprehensive, but also two classes
It can complement each other after active peptides mixing, play bio-cooperation, the relatively single animal blood extracted compared with traditional handicraft
Polypeptide can play stronger effect in fields such as medical treatment, beauty, health cares.
3. complex peptides active peptide A(blood plasma ultrafiltration peptide of the present invention) and active peptide B(plasma protein digestion peptide) extraction process
In be respectively adopted shut off molecular weight 5000Da and the ultrafiltration membrane of molecular weight 3000Da of shutting off, this key is shut off molecular weight
Selection and setting, realize shutting off for bacterium in composite reactive peptide, virus and bacterial endotoxin well, are animal blood active peptide
Biological safety provide technology ensuring.
4. using the molecular weight that shuts off in the extraction process of active peptide A of the present invention and active peptide B as the reverse osmosis of 100-200Da
Permeable membrane is concentrated, and the effective concentration of animal blood active peptide is not only increased, and reduces the water content of active peptide, for activity
The production of peptide freeze-dried powder preparation is provided convenience.
5. " waste liquid " of upper batch in animal blood active peptide preparation process of the present invention, the raw material that can become lower batch carry out
It is recycled, realizes the ecological extraction operation of closed loop, overall process does not generate any waste liquid, waste residue, not only realizes animal
The high value added utilization of blood, and accomplished truly environmentally protective.
6. carrying out ultrafiltration to enzymolysis liquid using the ultrafiltration membrane for the molecular weight 3000Da that shuts off in the present invention, 3000Da points are extracted
Son measures active peptide B below, and entire enzymolysis process does not need to carry out enzymolysis reaction by high temperature enzyme deactivation or chemical reagent, this
Sample not only saves the dosage of reagent, and makes extraction process more environmentally-friendly.
7. production process of the present invention uses modern biotechnology integrated technology, robotic degree is high, easy to operate, at low cost,
Convenient for large-scale production, there is huge economic benefit and social benefit.
Specific embodiment
Following example is only to further illustrate the present invention, and the invention is not limited in any way.
Embodiment 1
(1) the health deer blood qualified through quarantine is acquired, tube centrifuge separation, collection blood plasma is spare, and blood cell can be used for preparing
Animal protein feed or the SOD of extraction；
(2) the tubular type filter filtration sterilization by blood plasma through 3 concatenated 0.22 micron pore sizes；
(3) by blood plasma after filtration sterilization through the ultrafiltration apparatus ultrafiltration that molecular weight is 5000Da of shutting off, permeate is collected respectively and is cut
Flow liquid is spare；
(4) by the permeate in step (3) through shut off molecular weight be 200Da reverse osmosis membrane be concentrated by ultrafiltration to step 3 permeate
Original volume 1/8 collects shut off liquid and permeate respectively, and the liquid that shuts off is deer blood active peptides A(content of peptides >=8mg/ml), thoroughly
It crosses liquid and can be used as equipment water for cleaning and recycle；
(5) liquid that shuts off in step (3) is put into reaction kettle, compound protease (trypsase 600U/mL, pawpaw egg is added
White enzyme 200U/mL(is in terms of trapped fluid volume)), i.e., 6 hours are kept the temperature under the conditions of pH8.5,40 DEG C of temperature, obtains deer hemalbumin
Enzymolysis liquid；
(6) by the enzymolysis liquid in step (5), the ultrafiltration apparatus ultrafiltration through the molecular weight 3000Da that shuts off, respectively collect shut off liquid and
Permeate continues to digest, recycle wherein the liquid that shuts off can be added in lower batch of material liquid step (5)；
(7) permeate in step (6) is concentrated by ultrafiltration through the reverse osmosis membrane for the molecular weight 200Da that shuts off and is penetrated into step (6)
The 1/3 of liquid original volume, collection is shut off respectively and permeate, the liquid that shuts off are deer blood polypeptide B (>=20mg/ml), and permeate can be made
It is recycled for equipment water for cleaning；
(8) it will be mixed with the deer blood polypeptide B in the deer blood polypeptide A and step (7) in batch step (4), as deer blood activity is multiple
Close peptide.
Embodiment 2
(1) it is spare that anti-coagulants is added in the qualified fresh pig blood plasma of purchase quarantine；
(2) the tubular type filter filtration sterilization by blood plasma through 2 concatenated 0.22 micron pore sizes；
(3) by blood plasma after filtration sterilization through the ultrafiltration apparatus ultrafiltration that molecular weight is 5000Da of shutting off, permeate is collected respectively and is cut
Flow liquid is spare；
(4) by the permeate in step (3) through shut off molecular weight be 100Da reverse osmosis membrane be concentrated by ultrafiltration to the 1/ of original volume
10, shut off liquid and permeate are collected respectively, and the liquid that shuts off is pig blood active peptides A(content of peptides >=10mg/ml), permeate can
It is recycled as equipment water for cleaning；
(5) liquid that shuts off in step (3) is put into reaction kettle, compound protease (trypsase 300U/mL, neutral egg is added
White enzyme 500U/mL, alkali protease 200U/mL(are in terms of trapped fluid volume)), i.e., 4 are kept the temperature under the conditions of pH7.5, temperature 45 C
Hour, obtain porcine haemoglobin enzymolysis liquid
(6) by the enzymolysis liquid in step (5), the ultrafiltration apparatus ultrafiltration through the molecular weight 3000Da that shuts off, respectively collect shut off liquid and
Permeate continues to digest, recycle wherein the liquid that shuts off can be added in lower batch of material liquid step (5)；
(7) permeate in step (6) is concentrated by ultrafiltration through the reverse osmosis membrane for the molecular weight 100Da that shuts off to the 1/5 of original volume, point
It Shou Ji not shut off and permeate, the liquid that shuts off are peptides from swine blood B (>=33mg/ml), permeate can be used as equipment water for cleaning and follow
Ring utilizes；
(8) the peptides from swine blood B in the peptides from swine blood A and step (7) in same batch step 4 is mixed, as pig blood activity is compound
Peptide.
Claims (8)
1. a kind of preparation method of animal blood activity complex peptides, which comprises the following steps:
(1) the healthy animal blood qualified through quarantine is acquired, centrifuge separation, collection blood plasma is spare, or directly purchase quarantine is qualified
Fresh animal blood plasma；
(2) the tubular type filter filtration sterilization by blood plasma through 2 ~ 3 concatenated 0.22 micron pore sizes；
(3) by blood plasma after filtration sterilization through the ultrafiltration apparatus ultrafiltration that molecular weight is 5000Da of shutting off, permeate is collected respectively and is cut
Flow liquid is spare；
(4) by the permeate in step (3) through shut off molecular weight be 100-200Da reverse osmosis membrane be concentrated by ultrafiltration to permeate original
The 1/8-1/10 of volume collects shut off liquid and permeate respectively, and the liquid that shuts off is animal blood active peptides A i.e. blood plasma ultrafiltration peptide,
Permeate can be used as industrial water and recycle；
(5) liquid that shuts off in step (3) is put into reaction kettle, compound protease is added, each protease additional amount is with trapped fluid
Volume is calculated as 200-600U/mL, i.e., keeps the temperature 4 hours or more under the conditions of pH7.0-8.5,35-50 DEG C of temperature, obtain animal blood
Protein enzymatic hydrolyzate；
(6) by the enzymolysis liquid in step (5), the ultrafiltration apparatus ultrafiltration through the molecular weight 3000Da that shuts off, respectively collect shut off liquid and
Permeate continues to digest, recycle wherein the liquid that shuts off can be added in lower batch of material liquid step (5)；
(7) permeate in step (6) is concentrated by ultrafiltration through the reverse osmosis membrane for the molecular weight 100-200Da that shuts off to permeate substance
Long-pending 1/3-1/5, collection is shut off respectively and permeate, the liquid that shuts off are animal blood polypeptide B i.e. plasma protein digestion peptide, permeate
It can be used as industrial water to recycle；
(8) it will be mixed with the animal blood polypeptide B in the animal blood polypeptide A and step (7) in batch step (4), as animal blood
Active complex peptides.
2. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that divide in step (1)
From when seperator used be channel separator.
3. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that divide in step (1)
Blood cell from after can be used for preparing the SOD of animal protein feed or extraction.
4. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that the compound protein
Enzyme can in trypsase, papain, alkali protease and neutral proteinase it is any two or more, additional amount
For 200-600U/mL.
5. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that step (4), (7)
Described in refer to equipment water for cleaning as industrial water.
6. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that the animal blood is
Refer to pig blood, deer blood, sheep blood, ox blood, camel blood mammal blood.
7. a kind of preparation method of animal blood activity complex peptides as described in claim 1, which is characterized in that involved extraction process
It can also be used for the extraction process that active peptides are carried out using its hetero-organization of animal such as spleen, pancreas, placenta as material.
8. a kind of application such as the animal blood activity complex peptides of claim 1 method preparation in terms of medical treatment, beauty, health care,
Medical aspect can be used for can be used in terms of the Regeneration and Repair of tissue, beauty the reparation of allergy skin quality, and machine can be improved in health care
Body immunity.
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Application publication date: 20190723 |