CN105567771A - Pigskin small peptide chelate and preparation method and application thereof - Google Patents
Pigskin small peptide chelate and preparation method and application thereof Download PDFInfo
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- CN105567771A CN105567771A CN201610021722.4A CN201610021722A CN105567771A CN 105567771 A CN105567771 A CN 105567771A CN 201610021722 A CN201610021722 A CN 201610021722A CN 105567771 A CN105567771 A CN 105567771A
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- 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
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/36—Skin; Hair; Nails; Sebaceous glands; Cerumen; Epidermis; Epithelial cells; Keratinocytes; Langerhans cells; Ectodermal cells
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/39—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
- A61K8/65—Collagen; Gelatin; Keratin; Derivatives or degradation products thereof
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
- A61K8/981—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin of mammals or bird
- A61K8/985—Skin or skin outgrowth, e.g. hair, nails
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/4841—Filling excipients; Inactive ingredients
- A61K9/4858—Organic compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/4841—Filling excipients; Inactive ingredients
- A61K9/4866—Organic macromolecular compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
Abstract
The invention discloses a pigskin small peptide chelate and a preparation method and application thereof and belongs to the field of biotechnology. The pigskin small peptide chelate and the preparation method are characterized in that pigskin serves as a starting material, enzymolysis, ultrafiltration, resin column purification, a chelating reaction and drying are conducted, the pigskin small peptide chelate is obtained, and preparation auxiliaries can be added, and the pigskin small peptide chelate can be prepared into corresponding preparations according to a conventional method. By the adoption of the pigskin small peptide chelate, wound healing can be remarkably promoted, the beautifying and anti-wrinkle functions are achieved, and the pigskin small peptide chelate has the advantages of being free of toxic or side effect, free of drug injury and low in cost.
Description
Technical field
The present invention relates to biological technical field, be specifically related to small-peptide chelated thing of a kind of pigskin and its preparation method and application.
Background technology
Pigskin begins to be loaded in Materia Medica of Decoction, and from ancient times to the present, pigskin is applied to the treatment of various relative disease always, and modern clinic is used for the treatment of surgical skin damage.Protein content in pigskin is up to 33%, and wherein collagen content is 87.8%.Pigskin and human body skin structural similitude, containing I abundant type and III Collagen Type VI, and and homology between people comparatively ox, mouse are good, potential disease source is few, in addition China Shi Chan pig big country, have the pigskin resource of abundant super quality and competitive price, people are more and more deep to the research of extracting collagen from pigskin, and the existing research with pigskin being collagen prepared by raw material gets more and more.
Pigskin is the thing of normal food, but as medicine, early on the books.The medicinal pig skin by name of pigskin, taste is sweet cool in nature, the effect have promoting blood circulation and hemostasis, enrich and benefit essence and blood, Glycerin, gloss hair, reducing wrinkle, delay senility, because containing a large amount of collagen proteins in pigskin, the body cell that can slow down is aging, especially to yin asthenia generating intrinsic heat, occur having sore throat, the edible for patients of the disease such as low-heat is better.Pig skin can be processed to decoct and be smelt gelatin, and nowadays commercially available " new donkey-hide gelatin " is exactly replace donkey hide to endure " donkey-hide gelatin " of glue with pigskin.China is that the Eastern Han Dynasty cures holy Zhang Zhongjing with pig skin beautifying with what cure the disease the earliest, he has formulated " pig skin soup " in treatise on Febrile Diseases, control SHAO YIN syndrome, lower profit, pharyngalgia, fullness sensation in chest, vexed: 1 jin, pig skin, a pipeful of with water, boil and get five liters, go dregs, with whitish honey one liter, white powder pentahapto, endure fragrant and close, temperature point six clothes; And point out, pig skin function " and blood vessels, profit skin ", and the effect having yin-nourishing, promote the production of body fluid, quench the thirst.
Pharmacological research shows, the new donkey-hide gelatin produced by pigskin has the effect of hemopoietic, separately has report, new donkey-hide gelatin pair
60co irradiates caused hemopoietic function infringement mouse, make content of hemachrome in its peripheral blood, and leukocyte count and bonemarrow nucleated cells number all show higher than control group, and its curative effect is better than donkey-hide gelatin; The collagen produced by fresh porcine skin, in mouse burnt degree partial smearing immediately, can promote that skin mucosa injury heals, Wound healing rate is apparently higher than control group, light microscopy checking, the hyperplasia phenomenon of fibrous tissue under new epithelize, inoblast is all obviously better than control group to the trend of fibrocyte differentiation and collagenzation, heals have obvious promoter action to wound.
The little peptide of pigskin is the small-molecule peptide using biotechnology and zymolysis technique to extract from pigskin, its relative molecular mass at below 3KD, easy Transdermal absorption, activity is stronger, solvability and digestive utilization ratio high, safety has no side effect, no dependence.
Trace element is closely bound up with HUMAN HEALTH, micro-Science Research Association president professor Chen Xiangyou of China points out: " people is by elementary composition; people sick, old, dead invariably relevant with element balance; the breakthrough of the difficult and complicated cases of world today's prestige association human health longevity, awaits the development of element balance medical science ".Trace element has been successfully applied to the treatment of disease, research shows, iron deficiency can cause anaemia, scarce copper causes anaemia, osteoporosis, coronary heart disease, scarce cobalt can cause macrocytic anemia, leukemia, cataract, stomatocace etc., scarce chromium can cause diabetes, coronary heart disease, and scarce molybdenum there will be cardiovascular disorder, as myocardial necrosis coronary heart diseases and angina pectoris etc.
Zinc is one of trace element of needed by human, plays an important role in the important physiological processs such as growth in humans's growth, reproduction heredity, immunity, internal secretion, by the laudatory title of people titled with " flowers of life ", " the harmonious element of marriage ".Zinc is present in numerous enzyme systems, as in carbonic anhydrase, respiratory enzyme, LDH, superoxide-dismutase, alkaline phosphatase, the poly-middle enzyme of DNA and RNA etc., nucleic acid, protein, the synthesis of carbohydrate and the required material of vitamin A utilization, there is enhancing development, improve the effect of the sense of taste.Zinc is mainly present in animal food, is main mainly with plant food in Chinese family, and the absorption of the severe jamming zinc such as oxalic acid, phytic acid, Mierocrystalline cellulose in vegetable food.Modern science research proves, scarce zinc can cause various illness: such as, and zinc is to promoter action of having grown, by participating in the formation of enzyme, zinc promotes that nucleic acid protein synthesizes, affect growth and the division of cell, so the serious children lacking zinc will show as hypoevolutism, cretinism, malnutritive, thin and weak, anaemia, hair is yellow.The serious pregnant woman lacking zinc is out of shape due to karyomit(e), can cause the increase of abortion rate, cause habitual abortion; Zinc can strengthen wound tissue's regenerative power, scarce zinc will cause collagen quantity in wound epidermis and callus to reduce, wound not easily heals, so, zinc can assisting therapy stomatocace, stomach ulcer, burn, wound and adolescent acne, lower limb fester, the illness such as bedsore; Zinc maintains the normal growth of brain and brain function grows indispensable trace element, lacks the formation that zinc affects brain DNA, RND, protein, cerebellum development will be caused slow, behavior departs from, mental retardation, so, scarce zinc is the main reason causing brain paralysis, and this point is even more important to pregnant woman; Zinc still promotes the important element of the secretion of male testical ketone and spermiogenesis tail, so scarce zinc is also the major cause of male sterility, sexual dysfunction.Allly above show, important to human body of zinc element, should cause the attention of each crowd.
Copper is a kind of metallic chemical element, is also the necessary a kind of trace element of human body.Human body lacks copper can cause anaemia, paratrichosis, and bone and artery exception, so that cerebral disorders.Copper and HUMAN HEALTH in close relations.Human body all will take in various trace element every day, and copper is one of human body metallic element that can not lack.In adult human body, in 1 kg body weight, copper content is approximately 1.4mg ~ 2.1mg; In blood, the content of copper is about 1.0mg ~ 1.5mg.Though this quantity is little, it is healthy indispensable with normal operation that is organ for maintenance.This is because copper has special effect in body runs.Copper is the important component part of body internal protein and enzyme, the participation of many important enzyme require trace coppers and activation.Such as, copper can the synthesis of catalysis oxyphorase.Research shows, scarce copper can cause plasma cholesterol to raise, and increases atherosclerotic danger, because of but cause the important factor of coronary heart disease.Scientist also finds, it is relevant that the generation of the diseases such as nutritional anemia, vitiligo, osteoporosis, cancer of the stomach and esophagus cancer also all lacks copper with human body.Serious scarce copper and long-term marginality lack copper, also can cause dysplasia of children and some endemy.Copper is the indispensable micro-nutrients of HUMAN HEALTH, and for blood, nervus centralis and immunity system, growth and the function of hair, skin and bone tissue and brain and the internal organ such as liver, the heart have material impact.Copper is mainly taken in from diet.The World Health Organization advises, in order to remain healthy, adult's per kilogram of body weight should take in 0.03 milligram of copper every day.Pregnant woman and infant should double.Scarce copper can give rise to diseases, and can take cupric tonic and pill is supplemented.Copper is very light to the genotoxic potential of human body, only has when intake greatly exceed normal value, and side can cause the untoward reactions such as gastrointestinal dysfunction.Result of study shows, when adult Men's and Women's intake every day is respectively more than 12mg and 10mg, can produce slight impact to human biochemistry's process.In view of the genotoxic potential of copper is lower, it is but the indispensable element of HUMAN HEALTH on the contrary, and the expert group of the World Health Organization draws a conclusion, and the harm lacking copper is much bigger more than the toxicity of copper.Except the heredity that some is rare, what people mainly prevented is scarce copper.
Some metallics is that collage synthesis must obtain, and as copper, when the content of copper in skin cells raises, collagen protein synthesis increases.But, copper is a kind of poisonous metal, and copper peptide is not only without toxicity, and can promote collage synthesis, wound healing, effective smooth out wrinkles.Copper peptide (GHK-Cu), have another name called palmitoyl tripeptide 3, blue copper victory peptide, the complex compound of glycyl-L-histidyl--1B and copper, copper peptide was separated in 1973 at first and obtains from the blood plasma of people, and found that it had wound repair function in 1985, investigator in 1999 thinks that copper peptide and copper composition thereof can as the activator of tissue remodeling, it is also a signal peptide, promote that the outside a large amount of collagen protein of scar gathers the degraded of thing, the synthesis of the normal collagen protein of skin, elastin, the generation of proteoglycan and grape amine glycan, the speed of growth of different cell type and migration, anti-inflammatory, Antioxidation reaction.Johnson & Johnson's skin research centre research in University of Pennsylvania dermatology professor and this school shows, copper peptide is used in skin care and makeup helpful to improving skin ageing phenomenon, product containing copper peptide can improve skin problem fast, comprises and reduces microgroove, wrinkle, coarse, jaundice, cutis laxa and pigementation (foxiness).
The small-peptide chelated thing of pigskin further can not only improve effect of little peptide, the specific absorption of the trace element such as zinc, copper can also be improved, and human body can also be made to take in two class materials (little peptide and trace element), respective and collaborative performance promotes the effect of wound healing and beautifying face, preserving moisture.
Summary of the invention
The first object of the present invention is to provide a kind of pigskin small-peptide chelated thing, and the second object of the present invention is to provide the small-peptide chelated thing of this pigskin for the preparation of the application in the food of short wound healing and beautifying wrinkle-removing, protective foods, medicine and makeup.
The object of the invention first object is achieved in that
The small-peptide chelated thing of a kind of pigskin, is characterized in that adopting following processing step to prepare:
(1) preparation of the little peptide of pigskin: by fresh pig skin stripping and slicing, add 8-12 times amount 1-3% soaking in sodium carbonate solution 6 hours, discard alkali lye, rinse to weakly acidic pH with water, add 8-12 times of water gaging homogenate, add water boil, wait to be chilled to about 35 ~ 45 DEG C, adjust ph 1.0 ~ 3.0, add 0.5 ~ 2% stomach en-, be incubated 1 ~ 3 hour, adjust ph 7.0 ~ 9.0, add 0.5 ~ 2% trypsinase, be incubated 3 ~ 5 hours, boil 10 ~ 15 minutes, centrifugal, supernatant liquor is the ultra-filtration membrane ultrafiltration of 3KD by relative molecular mass, permeate is added on DA201-C macroporous adsorptive resins, with water elution, discard water lotion, again with 60 ~ 90% ethanol elutions, collect ethanol eluate, decompression recycling ethanol also continues to be concentrated into the clear cream that 60 DEG C of relative densities are 1.10 ~ 1.15, (inlet temperature is 180 ~ 200 DEG C to spraying dry, air outlet temperature is 90 ~ 100 DEG C), obtain the little peptide extract of pigskin.Containing Gln-Pro-Gly-Gly-Gly-Lys-Ala-Ser-Gly-Ala-Gly-Asn-Asp-Ala-Leu-Ser-Gly-Ile-Trp in the little peptide extract of pigskin, Val-Ser-Gly-Gly-Leu-Pro-Gly-Leu-Pro-Gly-Ala-Asp-Ala-Leu-Ser-Pro-Gly-Pro-Ala-Leu-Gly-Ala-Gly, Gln-Pro-Gly-Ala-Val-Ser-Gly-Ser-Ala-Ser-Thr-Pro-Gly, Lys-Gly-Gly-Pro-Gly-Ala-Gly-Ser-Ala-Gly-Gly-Thr-Pro-Gly-Ala-Pro-Gln, Arg-Thr-Leu-Pro-Gly-Ser-Pro-Ala-Val-Leu-Leu-Gly-Gly-Gly-Pro-Val-Ala, Pro-Gly-Gly-Ala-Gly-Val-Pro-Ala-Pro-Gly-Ala-Pro-Gln-Gly-Glu six kinds of little peptides, total content is 50% ~ 90%.
(2) huge legendary turtle closes reaction: by zinc chloride (or Zinc Gluconate, or zinc lactate, or Cuprocitrol, or copper gluconate, or copper sulfate) being dissolved in distilled water, to be made into concentration be 10 ~ 15mg/ml salts solution, with dilute hydrochloric acid adjust ph 2 ~ 4, the little peptide extract of pigskin is added in salts solution and pigskin little peptide extract ratio 8 ~ 10:1 ratio, stirring and dissolving, under temperature is 50 ~ 60 DEG C of conditions, chelatropic reaction 60 ~ 90 minutes, with nanofiltration membrane (retaining relative molecular mass is 150 ~ 300Da) nanofiltration desalination, obtains the small-peptide chelated thing solution of pigskin.
(3) spraying dry obtains inner complex: it is 1.10 ~ 1.15 that small-peptide chelated for pigskin thing solution decompression is concentrated into 60 DEG C of relative densities, and spraying dry (inlet temperature is 180 ~ 200 DEG C, and air outlet temperature is 90 ~ 100 DEG C), obtains the small-peptide chelated thing of pigskin.
Through the small-peptide chelated thing of pigskin prepared by aforesaid method, for the preparation of the following preparation of one-tenth::
(1) get pure water, add carbomer-980NF, moisten the 12h that rises, add triethylamine and adjust pH7.5, slaking 12h, add the small-peptide chelated thing of pigskin, add ethyl p-hydroxybenzoate, tween-80, glycerine, stir evenly, and is prepared into gelifying agent;
(2) by the small-peptide chelated thing of pigskin: Semen Maydis oil: beeswax: lecithin is 10:36:3:1 mixture, through colloidal mill grinding, soft capsule is prepared into;
(3) in the small-peptide chelated thing of pigskin: poly(oxyethylene glycol) 400: propylene glycol ratio is 10:38:2 mixture, through colloidal mill grinding, soft capsule is prepared into;
(4) the small-peptide chelated thing of pigskin, adds customary adjuvant, is prepared into oral liquid, capsule, tablet, granule;
(5) by following prescription proportioning, newborn cream cosmetic is prepared into:
The small-peptide chelated thing of pigskin 10 parts,
Shea butter 2 parts,
Jojoba oil 2 parts,
305 emulsifying agents (France's match Bick) 2 parts,
Glycerine 10 parts,
Carbomer 20200.1 parts,
Trolamine 0.1 part,
D-panthenol 1 part,
EDTA-2Na0.1 part,
Water-solubleazone 1 part,
0.05 part, Fragrant Lily essence,
Pure water 71.65 parts.
But the invention is not restricted to above-mentioned formulation.Moulding process conventional dose auxiliary material used can replace, as assistant formation pharmaceutical excipients such as Icing Sugar, honey, Xylo-Mucines with other kin auxiliary material; Also can share with other medicinal substances, jointly reach the effect of short wound healing and beautifying wrinkle-removing.
The second object of the present invention is to provide the purposes of the small-peptide chelated thing of above-mentioned pigskin, and it has the effect of short wound healing and beautifying wrinkle-removing, and it can apply in for the preparation of the food of short wound healing and beautifying wrinkle-removing, protective foods, medicine and makeup.
The invention has the beneficial effects as follows: the small-peptide chelated thing of pigskin of the present invention, prepare easy, cost is low; The little peptide of pigskin and trace element work in coordination with the effect playing short wound healing and beautifying wrinkle-removing, have simultaneously toxic side effect little, have no drug resistance, the not easily advantage such as residual, reach the requirement of green food pure natural, can good economic benefit be obtained.
The test of pesticide effectiveness
For the effect of the short wound healing and beautifying wrinkle-removing of verifying the small-peptide chelated thing of pigskin of the present invention further, inventors performed the test of pesticide effectiveness and investigate, test method and result as follows:
1. on the impact of rat skin mechanical trauma healing
(1) test medicine
The little peptide copper chelate of pigskin, self-control, containing the little peptide of pigskin (with total phthaleinometer) 71.8%, cupric 1.37%.CMC-Na solution is adopted to be made into respective concentration liquid before use.
The little peptide chelates of zinc of pigskin, self-control, containing the little peptide of pigskin (with total phthaleinometer) 72.4%, containing zinc 1.40%.Sterile purified water is adopted to be made into respective concentration liquid before use.
The little peptide of pigskin, self-control, containing the little peptide of pigskin (with total phthaleinometer) 73.8%.Sterile purified water is adopted to be made into respective concentration liquid before use.
Zinc Gluconate, food grade, Zn content is 14.3%.Sterile purified water is adopted to be made into respective concentration liquid for gavage before use.
Copper gluconate, food grade, copper content is 14.1%.Sterile purified water is adopted to be made into respective concentration liquid for gavage before use.
(2) animal and raising
1. animal species, source
SD rat, male, body weight 180 ~ 220g, provides purchased from by Shandong University's experimental animal center, animal conformity certification number: SCXK(Shandong) 20130004.
2. rearing conditions
Rat feeding in independent air-feeding cage (IVC), air purity 10000 grades, laboratory temperature 24 ± 2 DEG C; Relative humidity 60% ~ 80%; Air inerchange number of times per hour: 10-15 time/hour; Periodicity of illumination: 12(day)/12(night) hour.Male and female are separately raised, and every cage is no more than 5.
Feed: mouse full-valence pellet feed, works in coordination with medical bioengineering limited liability company purchased from Jiangsu Province, and its quality meets GB14924.1-2001 " laboratory animal mixed feed general-quality criteria ".
Bedding and padding: sterilizing particle bedding and padding, work in coordination with medical bioengineering limited liability company purchased from Jiangsu Province.
Drinking-water: drink purified water, freely drink after acidifying.
(3) test method and result
Rat 60 is got in animal model and grouping, by the anesthesia of 0.4ml/100g abdominal injection 1% Nembutal sodium solution, and local cropping preserved skin, ethanol for disinfection is sterilized, and is the circular incision of 2cm, cuts off skin holostrome in rat back operating scissors clip diameter, to subcutaneous fascia, cause mechanical trauma model.
Rat model is divided into 6 groups at random, only often organize lO, i.e. pigskin little peptide copper chelate group (100mg/kg), pigskin little peptide chelates of zinc group (100mg/kg), the little peptide group (100mg/kg) of pigskin, Cuprocitrol group (10mg/kg), Zinc Gluconate group (10mg/kg), blank group (physiological saline).
Administering mode is next day after model is made, and starts to give medicine: each test group respectively at wound surface smearing (or coated with) relative medicine, blank group in wound surface smearing physiological saline, every day 2 times, continuous 10d.
Every day is observed wound healing situation in the evaluation of wound healing situation, (surface of a wound is traced on Semitransparent Paper in lO days record surface of a wound areas, again as template, homogeneous hard paper is cut into pattern size, with hard paper weight secondary indication surface of a wound size, calculate with following formula: Wound healing rate (%)=(wound area-do not heal surface of a wound area in beginning)/start wound area) and put to death rat, cut part healing surface of a wound place tissue, fix with formalin solution and carry out histological examination.
Experimental result measurement data represents with mean+SD (± S), compares with one-way analysis of variance between two groups of sample averages; Numeration data is with chi square test.The Wound healing rate of each experimental group is in table 1.
The impact (± S) that table 1 heals on rat skin mechanical trauma
Note: compare with blank group, * P<0.05, * * P<0.01.
Above test-results shows: pigskin little peptide copper chelate group, pigskin little peptide chelates of zinc group, obviously can promote wound healing, have significant (P<0.01) with blank group comparing difference; Obviously can extend the swimming time of mouse in 10 DEG C of cold water, significant (P<0.05) is had with blank group comparing difference, the little peptide group of pigskin, Cuprocitrol group, Zinc Gluconate group compare can promote wound healing with blank groups, and effect pigskin little peptide group > Cuprocitrol group > Zinc Gluconate group, but difference is not significantly (P > 0.05).Illustrate that the small-peptide chelated thing effect of silkworm pigskin obviously strengthens.
Embodiment
Enumerate embodiment below, further illustrate the present invention, each embodiment, only for illustration of the present invention, does not limit the present invention:
Embodiment 1
By fresh pig skin stripping and slicing, add 10 times amount 2% soaking in sodium carbonate solution 6 hours, discard alkali lye, rinse to weakly acidic pH with water, add 10 times of water gaging homogenate, heated and boiled, wait to be chilled to about 40 DEG C, adjust ph 2.0, add 1% stomach en-, be incubated 1 hour, adjust ph 8.0, add 1% trypsinase, be incubated 3 hours, boil 15 minutes, centrifugal, supernatant liquor is the ultra-filtration membrane ultrafiltration of 3KD by relative molecular mass, permeate is added on DA201-C macroporous adsorptive resins, with water elution, discard water lotion, again with 90% ethanol elution, collect ethanol eluate, decompression recycling ethanol also continues to be concentrated into the clear cream that 60 DEG C of relative densities are 1.10 ~ 1.15, (inlet temperature is 180 ~ 200 DEG C to spraying dry, air outlet temperature is 90 ~ 100 DEG C), obtain the little peptide extract of pigskin.
Cuprocitrol being dissolved in distilled water, to be made into concentration be 15mg/ml salts solution, with dilute hydrochloric acid adjust ph 3, the little peptide extract of pigskin is added in salts solution and pigskin little peptide extract ratio 8:1 ratio, stirring and dissolving, under temperature is 60 DEG C of conditions, chelatropic reaction 90 minutes, with the nanofiltration membrane nanofiltration desalination of relative molecular mass 3KD, obtain the small-peptide chelated thing solution of pigskin, being evaporated to 60 DEG C of relative densities is 1.10 ~ 1.15, spraying dry (inlet temperature is 180 ~ 200 DEG C, and air outlet temperature is 90 ~ 100 DEG C), obtains the small-peptide chelated thing of pigskin.
Embodiment 2
By fresh pig skin stripping and slicing, add 12 times amount 3% soaking in sodium carbonate solution 6 hours, discard alkali lye, rinse to weakly acidic pH with water, add 12 times of water gaging homogenate, heated and boiled, wait to be chilled to about 45 DEG C, adjust ph 1.5, add 2% stomach en-, be incubated 2 hours, adjust ph 9.0, add 2% trypsinase, be incubated 4 hours, boil 15 minutes, centrifugal, supernatant liquor is the ultra-filtration membrane ultrafiltration of 3KD by relative molecular mass, permeate is added on DA201-C macroporous adsorptive resins, with water elution, discard water lotion, again with 75% ethanol elution, collect ethanol eluate, decompression recycling ethanol also continues to be concentrated into the clear cream that 60 DEG C of relative densities are 1.10 ~ 1.15, (inlet temperature is 180 ~ 200 DEG C to spraying dry, air outlet temperature is 90 ~ 100 DEG C), obtain the little peptide extract of pigskin.
Zinc Gluconate being dissolved in distilled water, to be made into concentration be 10mg/ml salts solution, with dilute hydrochloric acid adjust ph 4, the little peptide extract of pigskin is added in salts solution and pigskin little peptide extract ratio 10:1 ratio, stirring and dissolving, under temperature is 60 DEG C of conditions, chelatropic reaction 60 minutes, with the nanofiltration membrane nanofiltration desalination of relative molecular mass 3KD, obtain the small-peptide chelated thing solution of pigskin, being evaporated to 60 DEG C of relative densities is 1.10 ~ 1.15, spraying dry (inlet temperature is 180 ~ 200 DEG C, and air outlet temperature is 90 ~ 100 DEG C), obtains the small-peptide chelated thing of pigskin.
Embodiment 3
The small-peptide chelated thing 100g of pigskin of Example 1 method gained, by the small-peptide chelated thing of pigskin: Semen Maydis oil: beeswax: lecithin is 10:36:3:1 mixture, 30 minutes are ground through colloidal mill, with gelatin, glycerine, purified water, iron oxide black makes soft capsule hide glue gelatin in 100:40:100:1 ratio, control glue box temperature 55 ± 5 DEG C, heat insulation tank 40 ± 5 DEG C, sprinkler body temperature 36 ± 5 DEG C, with pellet press compacting soft capsule, the tolerant heavy 0.50g of every intragranular, 18 DEG C ~ 26 DEG C cold air drying 4h(relative humidity are 30% ~ 40%) sizing, wash ball, dry, select ball, packaging, be prepared into soft capsule finished product.
Embodiment 4
The small-peptide chelated thing 100g of pigskin of Example 1 method gained, in the small-peptide chelated thing of pigskin: poly(oxyethylene glycol) 400: propylene glycol ratio is 10:38:2 mixture, grind through colloidal mill, with gelatin, glycerine, purified water, iron oxide black makes soft capsule hide glue gelatin in 100:40:100:1 ratio, control glue box temperature 55 ± 5 DEG C, heat insulation tank 40 ± 5 DEG C, sprinkler body temperature 36 ± 5 DEG C, with pellet press compacting soft capsule, the tolerant heavy 0.50g of every intragranular, 18 DEG C ~ 26 DEG C cold air drying 4h(relative humidity are 30% ~ 40%) sizing, wash ball, dry, select ball, packaging, be prepared into soft capsule finished product.
Embodiment 5
The small-peptide chelated thing 200g of pigskin of Example 2 method gained, adds starch to total amount 250g, and granulation, dry, whole grain, load capsule, packaging, is prepared into capsule finished product.
Embodiment 6
The small-peptide chelated thing of pigskin of Example 1 method gained, by following prescription proportioning, is prepared into newborn cream cosmetic:
The small-peptide chelated thing of pigskin 10 parts,
Shea butter 2 parts,
Jojoba oil 2 parts,
305 emulsifying agents (France's match Bick) 2 parts,
Glycerine 10 parts,
Carbomer 20200.1 parts,
Trolamine 0.1 part,
D-panthenol 1 part,
EDTA-2Na0.1 part,
Water-solubleazone 1 part,
0.05 part, Fragrant Lily essence,
Pure water 71.65 parts.
Embodiment 7
Example 6 gained pigskin small-peptide chelated thing breast frost, carry out controlled trial on probation, include 60 examples altogether in, this product with contrast pigskin little peptide breast frost and organize each 30 routine, result shows: in pigskin small-peptide chelated thing breast frost group 30 examples, skin exquisiteness is soft and smooth, glossy, has slight sense of compacting, full flexible, canthus microgroove, corners of the mouth line, decree line, crow's feet shoal.In white 30 examples of control group pigskin little peptide breast, skin is fine and smooth, and have slight sense of compacting, canthus microgroove, corners of the mouth line, decree line, crow's feet shoal not obvious.Two groups are compared, pigskin small-peptide chelated thing breast frost is obviously better than pigskin little peptide breast frost, the small-peptide chelated thing of pigskin can be attacked directly from source and go deep into maintenance damage at the bottom of flesh, and strengthen skin maintenance ability, promote skin oneself nascent capacity, make the more smooth children of skin tender, Q bullet jade-like stone profit, can skin nutrition, unfold wrinkle, strengthen skin elasticity.
Claims (9)
1. the small-peptide chelated thing of pigskin, is characterized in that adopting following processing step to prepare:
(1) preparation of the little peptide of pigskin: by fresh pig skin stripping and slicing, add 8-12 times amount 1-3% soaking in sodium carbonate solution 6 hours, discard alkali lye, rinse to weakly acidic pH with water, add 8-12 times of water gaging homogenate, heated and boiled, wait to be chilled to about 35 ~ 45 DEG C, adjust ph 1.0 ~ 3.0, add 0.5 ~ 2% stomach en-, be incubated 1 ~ 3 hour, adjust ph 7.0 ~ 9.0, add 0.5 ~ 2% trypsinase, be incubated 3 ~ 5 hours, boil 10 ~ 15 minutes, centrifugal, supernatant liquor is the ultra-filtration membrane ultrafiltration of 3KD by relative molecular mass, permeate is added on DA201-C macroporous adsorptive resins, with water elution, discard water lotion, again with 60 ~ 90% ethanol elutions, collect ethanol eluate, decompression recycling ethanol also continues to be concentrated into the clear cream that 60 DEG C of relative densities are 1.10 ~ 1.15, spraying dry, obtain the little peptide extract of pigskin,
(2) huge legendary turtle closes reaction: zinc chloride is dissolved in distilled water and is made into salts solution, with dilute hydrochloric acid adjust ph 2 ~ 4, add the little peptide extract of above-mentioned pigskin, stirring and dissolving, under temperature is 50 ~ 60 DEG C of conditions, chelatropic reaction 60 ~ 90 minutes, with nanofiltration membrane nanofiltration desalination, obtains the small-peptide chelated thing solution of pigskin;
(3) spraying dry obtains inner complex: it is 1.10 ~ 1.15 that small-peptide chelated for pigskin thing solution decompression is concentrated into 60 DEG C of relative densities, and spraying dry obtains the small-peptide chelated thing of pigskin.
2. the small-peptide chelated thing of pigskin according to claim 1, it is characterized in that in the little peptide extract of step (1) described pigskin containing Gln-Pro-Gly-Gly-Gly-Lys-Ala-Ser-Gly-Ala-Gly-Asn-Asp-Ala-Leu-Ser-Gly-Ile-Trp, Val-Ser-Gly-Gly-Leu-Pro-Gly-Leu-Pro-Gly-Ala-Asp-Ala-Leu-Ser-Pro-Gly-Pro-Ala-Leu-Gly-Ala-Gly, Lys-Gly-Gly-Pro-Gly-Ala-Gly-Ser-Ala-Gly-Gly-Thr-Pro-Gly-Ala-Pro-Gln, Arg-Thr-Leu-Pro-Gly-Ser-Pro-Ala-Val-Leu-Leu-Gly-Gly-Gly-Pro-Val-Ala, Pro-Gly-Gly-Ala-Gly-Val-Pro-Ala-Pro-Gly-Ala-Pro-Gln-Gly-Glu, Gln-Pro-Gly-Ala-Val-Ser-Gly-Ser-Ala-Ser-Thr-Pro-Gly six kinds of little peptides, total content is 50% ~ 90%.
3. the small-peptide chelated thing of pigskin according to claim 1, is characterized in that step (2) described zinc chloride can use Zinc Gluconate, zinc lactate, Cuprocitrol, copper gluconate, copper sulfate to replace.
4. the small-peptide chelated thing of pigskin according to claim 1, is characterized in that the concentration of step (2) described salts solution is 10 ~ 15mg/ml.
5. the small-peptide chelated thing of pigskin according to claim 1, is characterized in that step (2) described salts solution and the little peptide extract ratio of pigskin are 8 ~ 10:1.
6. the small-peptide chelated thing of pigskin according to claim 1, is characterized in that the relative molecular mass that retains of step (2) described nanofiltration membrane is 150 ~ 300Da.
7. the small-peptide chelated thing of pigskin according to claim 1, it is characterized in that step (1) and (3) described spraying dry inlet temperature are 180 ~ 200 DEG C, air outlet temperature is 90 ~ 100 DEG C.
8. the small-peptide chelated thing of pigskin according to claim 1, is characterized in that being prepared into following preparation:
(1) get pure water, add carbomer-980NF, moisten the 12h that rises, add triethylamine and adjust pH7.5, slaking 12h, add the small-peptide chelated thing of pigskin, add ethyl p-hydroxybenzoate, tween-80, glycerine, stir evenly, and is prepared into gelifying agent;
(2) by the small-peptide chelated thing of pigskin: Semen Maydis oil: beeswax: lecithin is 10:36:3:1 mixture, through colloidal mill grinding, soft capsule is prepared into;
(3) in the small-peptide chelated thing of pigskin: poly(oxyethylene glycol) 400: propylene glycol ratio is 10:38:2 mixture, through colloidal mill grinding, soft capsule is prepared into;
(4) the small-peptide chelated thing of pigskin, adds customary adjuvant, is prepared into oral liquid, capsule, tablet, granule;
(5) by following prescription proportioning, newborn cream cosmetic is prepared into:
The small-peptide chelated thing of pigskin 10 parts,
Shea butter 2 parts,
Jojoba oil 2 parts,
305 emulsifying agents (France's match Bick) 2 parts,
Glycerine 10 parts,
Carbomer 20200.1 parts,
Trolamine 0.1 part,
D-panthenol 1 part,
EDTA-2Na0.1 part,
Water-solubleazone 1 part,
0.05 part, Fragrant Lily essence,
Pure water 71.65 parts.
9. the small-peptide chelated thing of pigskin according to claim 1 is used for short wound healing and beautifying wrinkle-removing.
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| CN105925652A (en) * | 2016-07-15 | 2016-09-07 | 安徽精准医疗产业研究院有限公司 | Method for producing small peptides through bionic enzymatic hydrolysis |
| CN105962106A (en) * | 2016-07-15 | 2016-09-28 | 安徽精准医疗产业研究院有限公司 | Method for producing donkey-hide gelatin small peptide by utilizing bionic enzymatic hydrolysis technology and application thereof |
| CN106975068A (en) * | 2017-04-28 | 2017-07-25 | 安徽生物肽产业研究院有限公司 | A kind of pigskin collagen small peptide active component composition |
| CN107126555A (en) * | 2017-04-28 | 2017-09-05 | 安徽生物肽产业研究院有限公司 | A kind of pigskin collagen small peptide active component composition of wound healing |
| CN107279458A (en) * | 2017-07-13 | 2017-10-24 | 安徽生物肽产业研究院有限公司 | It is a kind of that there is zinc-rich pea small peptide of enhancing immunity of organisms and preparation method thereof |
| CN112813123A (en) * | 2020-12-04 | 2021-05-18 | 苏州艾博迈尔新材料有限公司 | Preparation method of copper-containing complex |
| CN113980122A (en) * | 2021-10-28 | 2022-01-28 | 滨州医学院 | Preparation method and application of pigskin collagen peptide-ferrous chelate |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105925652A (en) * | 2016-07-15 | 2016-09-07 | 安徽精准医疗产业研究院有限公司 | Method for producing small peptides through bionic enzymatic hydrolysis |
| CN105962106A (en) * | 2016-07-15 | 2016-09-28 | 安徽精准医疗产业研究院有限公司 | Method for producing donkey-hide gelatin small peptide by utilizing bionic enzymatic hydrolysis technology and application thereof |
| CN106975068A (en) * | 2017-04-28 | 2017-07-25 | 安徽生物肽产业研究院有限公司 | A kind of pigskin collagen small peptide active component composition |
| CN107126555A (en) * | 2017-04-28 | 2017-09-05 | 安徽生物肽产业研究院有限公司 | A kind of pigskin collagen small peptide active component composition of wound healing |
| CN107279458A (en) * | 2017-07-13 | 2017-10-24 | 安徽生物肽产业研究院有限公司 | It is a kind of that there is zinc-rich pea small peptide of enhancing immunity of organisms and preparation method thereof |
| CN112813123A (en) * | 2020-12-04 | 2021-05-18 | 苏州艾博迈尔新材料有限公司 | Preparation method of copper-containing complex |
| CN113980122A (en) * | 2021-10-28 | 2022-01-28 | 滨州医学院 | Preparation method and application of pigskin collagen peptide-ferrous chelate |
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