CN107126579A - A kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder and its preparation method and application - Google Patents
A kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder and its preparation method and application Download PDFInfo
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- CN107126579A CN107126579A CN201710337394.3A CN201710337394A CN107126579A CN 107126579 A CN107126579 A CN 107126579A CN 201710337394 A CN201710337394 A CN 201710337394A CN 107126579 A CN107126579 A CN 107126579A
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- yak
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- 235000011852 gelatine desserts Nutrition 0.000 title claims abstract description 73
- 230000000025 haemostatic effect Effects 0.000 title claims abstract description 48
- 229940030225 antihemorrhagics Drugs 0.000 title claims abstract description 46
- 239000000843 powder Substances 0.000 title claims abstract description 45
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 43
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 43
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 238000004108 freeze drying Methods 0.000 claims abstract description 12
- 230000000740 bleeding effect Effects 0.000 claims abstract description 11
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- 238000001035 drying Methods 0.000 claims abstract description 5
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- 238000002682 general surgery Methods 0.000 claims abstract description 4
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- 239000000243 solution Substances 0.000 claims description 31
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- 239000007788 liquid Substances 0.000 claims description 18
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- PMMYEEVYMWASQN-DMTCNVIQSA-N Hydroxyproline Chemical compound O[C@H]1CN[C@H](C(O)=O)C1 PMMYEEVYMWASQN-DMTCNVIQSA-N 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 1
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- PMMYEEVYMWASQN-UHFFFAOYSA-N dl-hydroxyproline Natural products OC1C[NH2+]C(C([O-])=O)C1 PMMYEEVYMWASQN-UHFFFAOYSA-N 0.000 description 1
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- FGMPLJWBKKVCDB-UHFFFAOYSA-N trans-L-hydroxy-proline Natural products ON1CCCC1C(O)=O FGMPLJWBKKVCDB-UHFFFAOYSA-N 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/04—Surgical adhesives or cements; Adhesives for colostomy devices containing macromolecular materials
- A61L24/10—Polypeptides; Proteins
- A61L24/102—Collagen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/001—Use of materials characterised by their function or physical properties
- A61L24/0015—Medicaments; Biocides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/001—Use of materials characterised by their function or physical properties
- A61L24/0036—Porous materials, e.g. foams or sponges
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/001—Use of materials characterised by their function or physical properties
- A61L24/0042—Materials resorbable by the body
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/04—Surgical adhesives or cements; Adhesives for colostomy devices containing macromolecular materials
- A61L24/10—Polypeptides; Proteins
- A61L24/104—Gelatin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
- A61L2300/252—Polypeptides, proteins, e.g. glycoproteins, lipoproteins, cytokines
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/412—Tissue-regenerating or healing or proliferative agents
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/418—Agents promoting blood coagulation, blood-clotting agents, embolising agents
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/60—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a special physical form
- A61L2300/602—Type of release, e.g. controlled, sustained, slow
- A61L2300/604—Biodegradation
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- A—HUMAN NECESSITIES
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- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2400/00—Materials characterised by their function or physical properties
- A61L2400/04—Materials for stopping bleeding
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- Veterinary Medicine (AREA)
- Chemical & Material Sciences (AREA)
- Materials Engineering (AREA)
- Engineering & Computer Science (AREA)
- Dispersion Chemistry (AREA)
- Materials For Medical Uses (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention discloses former protein sponge of the Yak-skin Gelatin of a kind of quick-acting haemostatic powder and its preparation method and application.The collagen protein sponge of the quick-acting haemostatic powder is to extract I-type collagen and gelatin by yak skin, and mixed solution freeze-drying or convection drying obtain the support of three-dimensional porous structure.The collagen protein sponge of quick-acting haemostatic powder of the present invention can accelerate the anthemorrhagic speed of wound, promote wound cell propagation, prevent adhesion, accelerating wound healing serves as filler.With good compatibility, and energy is uniform, be closely adhere on the surface of a wound；Wound healing, there is certain mechanical strength, soft, is not deformed, good biocompatibility.The collagen protein sponge of quick-acting haemostatic powder of the present invention is widely used, available for the hemostasis of exocuticle bleeding, the hemostat application that viscera tissue is bled profusely, and for neurosurgery, orthopaedics, gynaecology, department of general surgery, the clinical medicine other field such as operating room.
Description
Technical field
The invention belongs to field of biomedical materials, and in particular to a kind of former egg of Yak-skin Gelatin acted on quick-acting haemostatic powder
Bai Haimian and its preparation method and application.
Background technology
Collagen is the main component of extracellular matrix, is distributed in each organ of body, be biological in-vivo content most
Many, most abundant protein, accounts for the 25%~30% of vivo protein total amount.Collagen contains a large amount of polar groups, tool
There are surface-active and good compatibility.The unique triple helix structure of collagen, assigns its good biocompatibility, life
The characteristics such as Biodegradable, low antigenicity.In addition, tropocollagen molecule can activate intrinsic coagulation pathway, and the hydroxyl dried meat contained
Propylhomoserin is the amino acid of a kind of repair again, after collagen and Platelet, can quickly cause follow-up poly- with blood
A series of progress of the associated processes of collection, so that rapid blood coagulation, reaches haemostatic effect；Meanwhile, collagen can promote into fibre
Propagation and the differentiation of cell are tieed up, induction karyoblast, lymphocyte, the aggregation of macrophage promote angiogenesis, and then
Accelerate regeneration and wound healing.Therefore, collagen shows greatly latent in terms of medicine, bioengineered tissue
Power.
Gelatin is the gentle fracture catabolite of collagen, is molecular weight distribution very wide peptide molecule mixture, molecular weight
It is general tens of thousands of to ten tens of thousands of.Gelatin is hydrophilic colloid, is a kind of ampholytes.There is substantial amounts of hydroxyl in gelatin molecule structure
Base, in addition also many carboxyls and amino；Gelatin contains the needed by human body such as interior propylhomoserin, glycine, proline and hydroxyproline
18 kinds of amino acid, but gelatin belongs to nutrition incomplete protein, lacks some essential amino acids, especially tryptophan.Gelatin is easy
Warm water is dissolved in, its solubility differs very little with setting temperature, is easily influenceed and gone bad by moisture content, temperature, humidity.Just because of gelatin
The multifrequency nature such as gelation, solid water, cohesive and dissolubility, its pharmaceuticals industry is had been widely used.
It is that raw material extract after collagen or gelatin that existing hemostasis collagen protein sponge, which uses ox heel string, with reference to it
His polymeric biomaterial, high molecular polymer or technology of preparing carry out the preparation of collagen protein sponge, single varieties, therefore open
The various hemostasis collagen protein sponge for sending out better has great importance.
The content of the invention
In order to solve problems of the prior art, primary and foremost purpose of the invention is to provide a kind of with quick-acting haemostatic powder
The former protein sponge of the Yak-skin Gelatin of effect, the collagen protein sponge can accelerate the anthemorrhagic speed of bleeding part, and be cured in tissue
Gradually degraded while conjunction, catabolite plays a part of promoting wound repair by promoting the effects such as cell propagation.
Another object of the present invention is to provide the system of the above-mentioned former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder
Preparation Method.
It is still another object of the present invention to provide answering for the above-mentioned former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder
With.
In order to realize the purpose of the present invention, the technical solution adopted by the present invention is as follows：
A kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder, it is characterised in that described that there is quick-acting haemostatic powder
The former protein sponge of the Yak-skin Gelatin of effect is made up of the former albumen of Yak-skin Gelatin and yak skin gelatin；
Further, it is described with the former protein sponge of Yak-skin Gelatin that quick-acting haemostatic powder is acted on be by mixed solution freeze-drying or
The support with three-dimensional porous structure that convection drying is obtained；
Further, it is described with the former protein sponge of Yak-skin Gelatin that quick-acting haemostatic powder is acted on be by mixed solution freeze-drying or
The support with three-dimensional porous structure that convection drying is obtained, the support possesses following characteristics：Quality >=5mg/cm3, pulling force
≥65g/cm2, water absorption is not less than 17 times of own wt before water suction, 65% (m/m) of total protein content >=gross weight, thickness
For 1~6 millimeter.
A kind of preparation method of the former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder, it is characterised in that including with
Lower step：
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, then rejects yak hair, be cut into length
5~8cm, wide 5~8cm fritters, and remove top layer and hypodermis layer, obtained yak skin skin corium fritter；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 1~3mm, wide by 1~
3mm.0.5molL-1 acetums, solid-liquid ratio 1:10~30, yak skin:Pepsin weight compares 100:1.0~2.5.Low temperature
16~36h is extracted in stirring, is filtered, saltouts, is purified, freeze-dried back；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, obtains extract solution, this is carried
Take liquid to utilize after the filtering with microporous membrane with a certain size aperture, obtain the extract solution of levels different molecular weight ranges, will
The upper strata extract solution concentration, freeze-dried back；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water low temperature molten after 1~4 mixing
Solution, concentration solid-liquid ratio to 1:0.6~0.2；
(5) grinding tool will be poured into after the concentrate solution vacuum obtained by step (4) or centrifugation degassing, mould is freezed 4 in -80 DEG C
It is freeze-dried after~8h, obtains the former protein sponge of described quick-acting haemostatic powder Yak-skin Gelatin.
It is preferred that, described in step (2) plus NaCl is saltoutd, NaCl ultimate densities to 3~5mol/L, afterwards repeatedly
Saltout and saltoutd step by step with 2.5~0.9mol/L of gradient；
It is preferred that, the 10~180kDa of molecular weight for the gelatin that step (3) the yak skin is extracted；
It is preferred that, the condition of step (3) described water extraction refers to that solid-liquid ratio is 1：10~20, temperature is 65~100 DEG C, is extracted
Number of times 1~4 time, every time 3~8 hours；
It is preferred that, the miillpore filter described in step (3) refers to that filtering filter membrane to the aperture that molecular weight is 10kDa apertures is
0.1 μm of filter membrane；
It is preferred that, the condition of the concentration described in step (3) (4) is low-temperature atmosphere-pressure concentration, and 30~40 DEG C of temperature or pressure are
0.09MPa, vacuum concentrates for 500~600mmHg pressurization；
It is preferred that, the freeze-drying described in step (2) (3) (5) be -10~-80 DEG C freeze 4~8h, then vacuum,
16~20h is freeze-dried at -60 DEG C of condenser temperature；
It is preferred that, the freeze-drying described in step (3) is to freeze 5~7h at -20~-80 DEG C, then in vacuum, cold-trap temperature
16~20h is freeze-dried at -60 DEG C of degree.
A kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder is in the hemostasis of exocuticle bleeding, internal organ group
Knit hemostasis and wound repair application in the hemostasis bled profusely, neurosurgery, orthopaedics, gynaecology and general surgery.
Compared with prior art, the present invention has advantages below and beneficial effect：
1. the present invention is using the skin of yak in Qinghai-tibet as raw material, using Yak Growth in extremely frigid zones, yak skin without dirt
Metachromia, and its extracted collagen/gelatine is compared to possible cause --- the glue of the denatured temperature (referring to table 1) of other source gelatin
The particularity of former protein amino acid sequence, develops the raw material that prepare of new collagen protein sponge, improves the added value of yak skin.
2. Extraction solvent used is pure water in extracting method of the present invention, later stage only cryogenic freezing and freeze-drying extract system
Preparation Method cost is low.
3. quick-acting haemostatic powder collagen protein sponge of the present invention uses easy to operate, film-strength height；Pliability is good.Can be according to using
It is required that difference, select proper strength timbering material.
4. quick-acting haemostatic powder collagen protein sponge of the present invention is widely used, can quick-acting haemostatic powder, protection wound, promote healing,
And for neurosurgery, orthopaedics, gynaecology, department of general surgery, the clinical medicine other field such as operating room.
5. the yak skin gelatin of HMW is intercepted in raw material of the present invention, because different molecular weight gelatin is to platelet aggregation
Facilitation it is inconsistent, the gelatin haemostatic effect of high molecular weight range becomes apparent from.
6. the present invention can accelerate the anthemorrhagic speed of bleeding part with the former protein sponge of Yak-skin Gelatin that quick-acting haemostatic powder is acted on,
And gradually degraded while organization healing, catabolite plays promotion wound repair by promoting the effects such as cell propagation
Effect.
The different enzymolysis time Yak-skin Gelatin samples of table 1 DSC values (N=3)
Different subscripts, which is represented, statistical discrepancy, P<0.05.
Embodiment
Below in conjunction with the embodiment of the present invention, the technical scheme in the embodiment of the present invention is clearly and completely described,
Obviously, described embodiment is only a part of embodiment of the invention, rather than whole embodiments.Based in the present invention
Embodiment, the every other embodiment that those of ordinary skill in the art are obtained under the premise of creative work is not made, all
Belong to the scope of protection of the invention.
Embodiment 1
1. prepare the former protein sponge of Yak-skin Gelatin
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, and 48h is soaked with 5% NaOH solution
After reject yak hair, 3 times are cleaned with distilled water to neutral, long 5~7cm, wide 5~7cm fritters is cut into, and remove epidermis and subcutaneous
Organized layer, obtains skin corium；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 1~2mm, wide by 1~
2mm.0.5mol/L acetums, solid-liquid ratio 1:15, yak skin:Pepsin weight compares 100:1.0.16h is extracted in low temperature stirring,
NaCl is added after filtering, in filtrate to be saltoutd, NaCl ultimate densities to 4mol/L stir 10h, and low-temperature centrifugation will be precipitated molten
In 0.5mol/L glacial acetic acid, then saltoutd repeatedly with the NaCl of 2.5,1.8 and 0.9mol/L step by step, acid it is molten, finally obtain collagen
Crude extract；In the bag filter that crude extract is fitted into 100kDa apertures, dialysed 16h with 0.04mol/L acetum, then with distilling
Water is dialysed 16h, and 4h rocks bag filter once, obtain after purified, is freeze-dried standby；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, and solid-liquid ratio is 1:15, temperature
Spend for 80 DEG C, extraction time 3 times, 5 hours every time, No. three extract solution mixing, will after the extract solution is using 0.1 μm of membrane filtration
Upper strata extract solution pressure be 0.09MPa, vacuum be 550mmHg under conditions of be concentrated into 1:0.2, it is freeze-dried standby；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water dissolution in low temperature after 1 mixing, it is dense
Contracting solid-liquid ratio is to 1:0.2；
(5) grinding tool will be poured into after the concentrate solution vacuum outgas of gained, mould is freezed after 6h in -20 DEG C in vacuum, cold
16h is freeze-dried at -60 DEG C of hydrazine temperature, that is, obtains the former protein sponge of quick-acting haemostatic powder Yak-skin Gelatin.
Prepared quick-acting haemostatic powder Yak-skin Gelatin olynthus thickness is about 3~5 millimeters, standby after co-60 radiation sterilizing.
2. Yak-skin Gelatin original protein sponge quick-acting haemostatic powder and the effect experiment for repairing wound：Tail bleeding is cut as mould using mouse
Type, by a diameter of 0.4 millimeter, the former protein sponge of thick 3~5 millimeters former sheet Yak-skin Gelatin attaches to the mouse for just cutting tail bleeding
At tail, rat-tail stops the bleeding time less than 10s, and the time for stopping bleeding than blank rat-tail shortens 25s or so.Use collagen egg
The mouse tail 1 week or so of Bai Haimian hemostasis, the mouse tail stopped blooding compared to unused collagen protein sponge, wound has similar new
The generation of saccus setigerus, epithelial tissue has growth tendency, and the surface of a wound and surrounding skin are close, and surface of a wound paler colour is more smooth, surface quilt
Cuticular cellulose is covered.
Embodiment 2
1. prepare the former protein sponge of Yak-skin Gelatin
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, and 12h is soaked with 10% NaOH solution
After reject yak hair, 5 times are cleaned with distilled water to neutral, long 6~8cm, wide 6~8cm fritters is cut into, and remove epidermis and subcutaneous
Organized layer, obtains skin corium；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 2~3mm, wide by 2~
3mm.0.5molL-1 acetums, solid-liquid ratio 1:20, yak skin:Pepsin weight compares 100:1.5.Low temperature stirring is extracted
24h, adds NaCl after filtering, in filtrate and is saltoutd, NaCl ultimate densities to 5mol/L, stirs 12h, low-temperature centrifugation will be heavy
Shallow lake is dissolved in 0.5mol/L glacial acetic acid, then saltoutd repeatedly with the NaCl of 2.1,1.5 and 1.0mol/L step by step, acid it is molten, finally obtain
Collagen crude extract；In the bag filter that crude extract is fitted into 100kDa apertures, dialysed 24h with 0.08mol/L acetum, then
Rocked bag filter with distilled water dialysis 24h, 6h once, obtained after purified, is freeze-dried standby；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, and solid-liquid ratio is 1:20, temperature
Spend for 90 DEG C, extraction time 3 times, 6 hours every time, No. three extract solution mixing, will after the extract solution is using 0.1 μm of membrane filtration
Upper strata extract solution pressure be 0.09MPa, vacuum be 600mmHg under conditions of be concentrated into 1:0.4, it is freeze-dried standby；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water dissolution in low temperature after 3 mixing, it is dense
Contracting solid-liquid ratio is to 1:0.3；
(5) grinding tool will be poured into after the concentrate solution vacuum outgas of gained, mould is freezed after 5h in -80 DEG C in vacuum, cold
- 60 DEG C of freeze-drying 14h of hydrazine temperature, that is, obtain quick-acting haemostatic powder Yak-skin Gelatin olynthus.
Prepared quick-acting haemostatic powder Yak-skin Gelatin olynthus thickness is about 2~4 millimeters, standby after co-60 radiation sterilizing.
2. Yak-skin Gelatin original protein sponge quick-acting haemostatic powder and the effect experiment for repairing wound：Tail bleeding is cut as mould using mouse
Type, by a diameter of 0.4 millimeter, the former protein sponge of thick 2~4 millimeters former sheet Yak-skin Gelatin attaches to rabbit ear central artery
The surface of a wound, the bleeding stopping period of the rabbit ear surface of a wound shortens 140s or so between 80~92s than the blank physical pressing haemostatic time.2 weeks
Afterwards, the rabbit ear wound face of collagen protein sponge hemostasis is more smooth, paler colour, close to surrounding skin.
Embodiment 3
1. prepare the former protein sponge of Yak-skin Gelatin
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, and 36h is soaked with 8% NaOH solution
After reject yak hair, 4 times are cleaned with distilled water to neutral, long 5~6cm, wide 5~6cm fritters is cut into, and remove epidermis and subcutaneous
Organized layer, obtains skin corium；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 1~2mm, wide by 1~
2mm.0.5mol/L acetums, solid-liquid ratio 1:30, yak skin:Pepsin weight compares 100:2.24h, mistake are extracted in low temperature stirring
NaCl is added after filter, in filtrate to be saltoutd, NaCl ultimate densities to 3mol/L stir 10h, and precipitation is dissolved in by low-temperature centrifugation
In 0.5mol/L glacial acetic acid, then saltoutd repeatedly with the NaCl of 2.0,1.4 and 0.9mol/L step by step, acid it is molten, finally obtain collagen thick
Extract；In the bag filter that crude extract is fitted into 100kDa apertures, dialysed 24h with 0.1mol/L acetum, then use distilled water
Dialyse 24h, and 4h rocks bag filter once, obtained after purified, is freeze-dried standby；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, and solid-liquid ratio is 1:20, temperature
Spend for 90 DEG C, extraction time 2 times, 8 hours every time, extract solution is mixed twice, the extract solution utilizes the membrane filtration in 15kDa apertures
Afterwards, by upper strata extract solution pressure be 0.09MPa, vacuum be 600mmHg under conditions of be concentrated into 1:0.2, it is freeze-dried standby
With；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water dissolution in low temperature after 1 mixing, it is dense
Contracting solid-liquid ratio is to 1:0.4；
(5) grinding tool will be poured into after the concentrate solution vacuum outgas of gained, mould is freezed after 6h in -80 DEG C in vacuum, cold
- 60 DEG C of freeze-drying 16h of hydrazine temperature, that is, obtain quick-acting haemostatic powder Yak-skin Gelatin olynthus.
Prepared quick-acting haemostatic powder Yak-skin Gelatin olynthus thickness is about 1~3 millimeter, standby after co-60 radiation sterilizing.
2. Yak-skin Gelatin original protein sponge is used as the effect experiment of tissue engineering material：Collagen protein sponge and application on human skin table
Skin fibroblast co-incubation, fibroblast can be with normal growth, and cellular morphology is normal, and over time retired, carefully
Born of the same parents attenuate with the part collagen in collagen sponge coculture, and cell quantity increase, the space of sponge is less and less.
Embodiment 4
1. prepare the former protein sponge of Yak-skin Gelatin
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, and 24h is soaked with 10% NaOH solution
After reject yak hair, 5 times are cleaned with distilled water to neutral, long 7~8cm, wide 7~8cm fritters is cut into, and remove epidermis and subcutaneous
Organized layer, obtains skin corium；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 2~3mm, wide by 2~
3mm.0.5mol/L acetums, solid-liquid ratio 1:25, yak skin:Pepsin weight compares 100:2.5.36h is extracted in low temperature stirring,
NaCl is added after filtering, in filtrate to be saltoutd, NaCl ultimate densities to 3.5mol/L stir 12h, and low-temperature centrifugation will be precipitated
Be dissolved in 0.5mol/L glacial acetic acid, then saltoutd repeatedly with the NaCl of 2.5,1.6 and 1.1mol/L step by step, acid it is molten, finally obtain glue
Former crude extract；In the bag filter that crude extract is fitted into 100kDa apertures, dialysed 36h with 0.08mol/L acetum, then with steaming
Distilled water is dialysed 36h, and 6h rocks bag filter once, obtain after purified, is freeze-dried standby；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, and solid-liquid ratio is 1:30, temperature
Spend for 85 DEG C, extraction time 4 times, 5 hours every time, No. four extract solution mixing, the extract solution utilizes the membrane filtration in 15kDa apertures
Afterwards, by upper strata extract solution pressure be 0.09MPa, vacuum be 580mmHg under conditions of be concentrated into 1:0.5, it is freeze-dried standby
With；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water dissolution in low temperature after 4 mixing, it is dense
Contracting solid-liquid ratio is to 1:0.4；
(5) grinding tool will be poured into after the concentrate solution vacuum outgas of gained, mould is freezed after 8h in -50 DEG C in vacuum, cold
18h is freeze-dried at -60 DEG C of trap temperature, that is, obtains quick-acting haemostatic powder Yak-skin Gelatin olynthus.
Prepared quick-acting haemostatic powder Yak-skin Gelatin olynthus thickness is about 2~4 millimeters, standby after co-60 radiation sterilizing.
2. Yak-skin Gelatin original protein sponge is used as the effect experiment of tissue engineering material：Collagen protein sponge and application on human skin table
Skin fibroblast co-incubation, fibroblast can be with normal growth, and over time, cell quantity increase is extra large
Continuous space is less and less.After being detected with AO/EB fluorescence colours, when finding culture 24h, the metabolic activity of cell not by
Have a strong impact on, and basic apoptotic cell-free.
Although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
A variety of changes, modification can be carried out to these embodiments, replace without departing from the principles and spirit of the present invention by understanding
And modification, the scope of the present invention is defined by the appended.
Claims (10)
1. a kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder, it is characterised in that described that there is quick-acting haemostatic powder to make
Yak-skin Gelatin original protein sponge is made up of the former albumen of Yak-skin Gelatin and yak skin gelatin.
2. a kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 1, it is characterised in that
It is described to be obtained with the former protein sponge of Yak-skin Gelatin that quick-acting haemostatic powder is acted on by mixed solution freeze-drying or convection drying
Support with three-dimensional porous structure.
3. a kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 1, it is characterised in that
It is described to be obtained with the former protein sponge of Yak-skin Gelatin that quick-acting haemostatic powder is acted on by mixed solution freeze-drying or convection drying
Support with three-dimensional porous structure, the support possesses following characteristics：Mass volume ratio (density) >=5mg/cm3, pulling force >=
65g/cm2, water absorption is not less than 17 times of own wt before water suction, 65% (m/m) of total protein content >=gross weight, and thickness is
1~6 millimeter.
4. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 1,
It is characterised in that it includes following steps：
(1) fresh yak in Qinghai-tibet Wild Yak yak skin is cleaned up, then rejects yak hair, be cut into it is long by 5~
8cm, wide 5~8cm fritters, and remove top layer and hypodermis layer, obtained yak skin skin corium fritter；
(2) prepared by type i collagen：The yak skin skin corium fritter that step (1) is obtained further is shredded, long 1~3mm, wide 1~3mm.
0.5molL-1 acetums, solid-liquid ratio 1:10~30, yak skin:Pepsin weight compares 100:1.0~2.5.Low temperature is stirred
16~36h is extracted, filters, saltout, purify, freeze-dried back；
(3) prepared by gelatin：The yak skin skin corium fritter that step (1) is obtained directly carries out water extraction, obtains extract solution, the extract solution
After the filtering with microporous membrane with a certain size aperture, the extract solution of levels different molecular weight ranges is obtained, will be described
Upper strata extract solution concentration, freeze-dried back；
(4) by the sample obtained by step (2) and step (3) according to mass ratio 1:Add distilled water dissolution in low temperature after 1~4 mixing, it is dense
Contracting solid-liquid ratio is to 1:0.6~0.2；
(5) grinding tool will be poured into after the concentrate solution vacuum obtained by step (4) or centrifugation degassing, mould is freezed into 4~8h in -80 DEG C
After be freeze-dried, obtain the former protein sponge of described quick-acting haemostatic powder Yak-skin Gelatin.
5. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 4,
Characterized in that, described in step (2) plus NaCl saltoutd, NaCl ultimate densities to 3~5mol/L, saltouing repeatedly afterwards
Saltoutd step by step with 2.5~0.9mol/L of gradient.
6. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 4,
Characterized in that, 10~the 180kDa of molecular weight for the gelatin that step (3) the yak skin is extracted, the condition of the water extraction refers to material
Liquor ratio is 1：10~20, temperature is 65~100 DEG C, extraction time 1~4 time, and 3~8 hours every time, described miillpore filter referred to
It is that filter membrane to the aperture in 10kDa apertures is 0.1 μm of filter membrane to filter molecular weight.
7. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 4,
Characterized in that, the condition of the concentration described in step (3) (4) is low-temperature atmosphere-pressure concentration, 30~40 DEG C of temperature or pressure are
0.09MPa, vacuum concentrates for 500~600mmHg pressurization.
8. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 4,
Characterized in that, the freeze-drying described in step (2) (3) (5) is to freeze 4~8h at -10~-80 DEG C, then in vacuum, cold-trap
16~20h is freeze-dried at -60 DEG C of temperature.
9. a kind of preparation method of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder according to claim 4,
Characterized in that, the freeze-drying described in step (3) be -20~-80 DEG C freeze 5~7h, then vacuum, condenser temperature -
16~20h is freeze-dried at 60 DEG C.
10. a kind of former protein sponge of Yak-skin Gelatin acted on quick-acting haemostatic powder is in exocuticle bleeding according to claim 1
Hemostasis, the hemostasis that viscera tissue is bled profusely, neurosurgery, orthopaedics, hemostasis and wound repair in gynaecology and general surgery
Using.
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