CN109200329A - A kind of preparation method and application of cloth-like hemostatic material - Google Patents
A kind of preparation method and application of cloth-like hemostatic material Download PDFInfo
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- CN109200329A CN109200329A CN201811251659.9A CN201811251659A CN109200329A CN 109200329 A CN109200329 A CN 109200329A CN 201811251659 A CN201811251659 A CN 201811251659A CN 109200329 A CN109200329 A CN 109200329A
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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
- A61L24/0005—Ingredients of undetermined constitution or reaction products thereof
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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
- A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
- A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
- A61L15/40—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons containing ingredients of undetermined constitution or reaction products thereof, e.g. plant or animal extracts
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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
- A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
- A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
- A61L15/42—Use of materials characterised by their function or physical properties
- A61L15/44—Medicaments
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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
- A61L24/0015—Medicaments; Biocides
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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
- A61L2400/00—Materials characterised by their function or physical properties
- A61L2400/04—Materials for stopping bleeding
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Abstract
The invention belongs to medical bio medical material fields, and in particular to a kind of preparation method and application of the cloth-like hemostatic material of animal origin, the material have many advantages, such as good haemostatic effect, it is easy to carry, have preferable safety and biocompatibility.A kind of preparation method of cloth-like hemostatic material, using silk cocoon as raw material, silk cocoon pull into cloth-like be placed in oven bake, it bakes to the quality of best haemostatic effect, after baking, cloth-like coke silk cocoon is trimmed to specific shape, sterilization packaging, it is final to obtain the cloth-like bleeding-stopping dressing with stronger anastalsis.Coke silk cocoon hemostatic material of the invention is easy to get, and cost is relatively low, and cost performance is high, and carrying portable, not with the tissue easy debridement of adhesion, can satisfy clinical demand.
Description
Technical field
The invention belongs to medical bio medical material fields, and in particular to a kind of system of the cloth-like hemostatic material of animal origin
Preparation Method and application, the material with good haemostatic effect, it is easy to carry, have preferable safety and biocompatibility etc. excellent
Point.
Background technique
Clinically surgical operation can all form bleeding, and the patient for clinically dying of excessive blood loss accounts for very big ratio, hemostasis
It is one of the core link in hospital surgical, good hemostatic technique is the key that guarantee successful surgery.In schedule life
Relatively mostly occur skin surface local hemorrhage.The development and improvement of hemostatic material have become the hot spot of current field of medicaments research.
With being constantly progressive for medical technology, new medical material and the relevant technologies have obtained good development.
Currently used effective Hemostasis is using the preferable resorbable material hemostasis of biocompatibility, preferably only
The key property of blood material is general are as follows: safety is good, insensitive, not allergy；It is bonded with wound closely, and there is good suction
Moisture performance；It does not need often to replace；Cost performance is high；Adherency and easy debridement etc. are not easy with wound.As people have hemostatic material
Excellent biomaterial in nature is found in the raising of effect property and security requirement, develop haemostatic effect more preferably, nontoxic secondary make
It is imperative with, hemostatic material nonirritant, easily processed into type.
Currently, hemostatic material in the market is mainly include the following types: Fibrin Glue class, collagen class, oxidized fibre
Plain class hemostatic gauze, chitosan class styptic sponge, starch-polysaccharides class or chitosan class styptic powder.Currently, these materials are in animal
All achieve good results in experiment and clinical application, but also all come with some shortcomings, show as follows: gelatin, which absorbs, generally to be needed
4 weeks or more are wanted, the infection risk of wound, and gelatin bad mechanical property itself are increased；Collagenous source is xenogenesis in animal tissue
Albumen has potential sensitization；Cellulose material degradation time is long, may bring the side effects such as infection, and structure pine
It dissipates, poor adhesion may cause bleeding again；Starch is easy to microorganism microbiological contamination, and the Arista of Medafor company of U.S. production can inhale
The property received starch hemostatic microsphere preparation process is more complex, expensive, can not popularize use；Degradation of chitosan is poor, and there are safety
Risk, pure chitosan material anastalsis is limited, the wound big for amount of bleeding, and adhesion is poor and hemostasis is slower, for
Extensive bleeding surface of a wound haemostatic effect is not highly desirable, is only used for the hemostasis of exterior trauma bleeding, internal without that can be used for internal organ etc.
The hemostasis of bleeding.Simultaneously the maximum bottleneck of domestic one for influencing the industry development of chitosan hemostatic material is chitosan material
Biocompatibility, state food pharmaceuticals administration general bureau (China Food and Drug Administration, CFDA)
Also the supervision to such product is increased, the chitosan product of II class is changed into Group III by force after 2014 and is supervised.The U.S.
The QuikClot zeolite base styptic powder (Z-Medica company) of FDA approval is used for the first aid of severe haemorrhage wound, however when use
More than 100 DEG C high temperature cause the damage of wound tissue.Existing hemostatic material is confined to simple hemostasis more, is promoting healing side
The research in face is also fewer.Develop it is a kind of with quick-acting haemostatic powder, promote the novel hemostatic materials of functions such as healing to have become hemostasis
The certainty of Materials.
This application provides the cloth-like coke silk cocoons after silk cocoon baking to have quick-acting haemostatic powder effect, and zoopery is found to artery
The advantages that bleeding also has preferable haemostatic effect, and has higher-security, carrying portable, inexpensive and nontoxic is ideal
Hemostatic material, but cloth-like coke silk cocoon did not did the hemostasis research and Mechanism Study of system, and this patent is using cloth-like coke silk cocoon as base
The silk cocoon dressing of plinth preparation, can be directly used for the big bleeding choke material of deep and big wound, or patch is made for skin
Surface local hemorrhage, or for the hemostasis of other Bleeding patterns, develops that easy to use, effectiveness is significant, take effect odd fast, safe nothing
Malicious and cheap and good-quality ideal bleeding-stopping dressing.
Summary of the invention
The present invention for the number of drawbacks of hemostatic material in the market, prepared it is a kind of it is easy to carry, can quick-acting haemostatic powder,
It is highly-safe, biocompatibility is good, cost performance is high, and will not with tissue the easy debridement of adhesion novel cloth-like coke silk cocoon hemostasis apply
Material, this method preparation process is simple, is expected to be widely used for the big bleeding choke material of deep and big wound, clinical operation hemostasis and skin
Skin surface local hemorrhage, or the hemostasis for other Bleeding patterns.
A kind of preparation method of cloth-like hemostatic material, using silk cocoon as raw material, silk cocoon pull into cloth-like be placed in oven bake, bake
To the quality of best haemostatic effect, after baking, cloth-like coke silk cocoon is trimmed to specific shape, sterilization packaging, it is final obtain have compared with
The cloth-like bleeding-stopping dressing of strong anastalsis.
Silk cocoon after baking is khaki or isabelline, water content 3-5%.
The baking condition is that 200 DEG C of bakings baking in 2 hours or 230 DEG C is baked 2 hours for 30 minutes or 230 DEG C.
Silk cocoon is pulled into cloth-like silk, cloth-like silk is placed in oven, which can be electric oven or solar energy oven, most
It is well transparent glass door, roasted hemostasis auxiliary material quality can be observed in real time.
Sterilization method is that irradiation sterilization Co-60 irradiates 10-30h or ethylene oxide sterilization processes, vacuum are protected from light aluminium foil bag packet
Dress, sticks mark.
The specific shape is the hemostasis auxiliary material shape for being suitble to operation or patch to use.
The application is additionally provided with a kind of hemostasis cloth and its preparation method and application.
Stop blooding cloth the preparation method is as follows:
Using silk cocoon as raw material, silk cocoon is pulled into cloth-like it is placed in oven and bake, bake to the quality of best haemostatic effect, baking
Afterwards, cloth-like coke silk cocoon is trimmed to specific shape, and sterilization packaging is final to obtain the cloth-like bleeding-stopping dressing with stronger anastalsis.
Silk cocoon after baking is khaki or isabelline, water content 3-5%.
The baking condition is that 200 DEG C of bakings baking in 2 hours or 230 DEG C is baked 2 hours for 30 minutes or 230 DEG C.
Cloth stop blooding for clinically surgical operation bleeding or skin surface local hemorrhage.The clinically group of a variety of surgical operations
It knits or organ hemorrhage.
The application method of hemostasis cloth, including following several: similar hemostatic gauze is directly clogged, especially for the big bleeding of wound
Big and deep wound；Or it is made into patch, for controlling serious skin surface local hemorrhage under contingency condition；Or for tearing
The skin surface of the bleedings such as lacerated wound, small incision and damaged wound is locally disposed, or for the hemostasis of other Bleeding patterns.
Realization process of the invention is as follows:
A kind of preparation method of animal origin cloth-like coke silk cocoon hemostasis auxiliary material, steps are as follows: being original with the cloth-like silk of drawing
Material, is baked several hours in a few Baidu's high temperature roasters, after being baked the quality with preferable haemostatic effect (such as khaki or brown
Yellow, about 4%), cloth-like coke silk cocoon is trimmed to specific shape to water content, and using Co-60 radiation sterilization, a kind of tool can be obtained
There is the sterile cloth-like coke silk cocoon bleeding-stopping dressing of higher styptic activity.
Specific implementation process of the present invention is as follows:
(1) silk cocoon removes silkworm chrysalis, to remove the silk cocoon of silkworm chrysalis as raw material；
(2) the silk cocoon drawing of removal silkworm chrysalis is made into cloth-like silk, the way of cloth-like silk can be placed on bow with the silk cocoon of softening, be drawn
It is made into cloth-like, cleans and dries；
(3) cloth-like silk raw material is baked several hours in a few Baidu's high temperature roasters, for example, 230 DEG C baking 2-3 hours, with not
Subject to black wood charcoal or preservation styptic activity (such as khaki or isabelline, water content is about 4%)；
(4) through roasted cloth-like silk through Co-60 radiation sterilization 10-30h, or other sterilization methods are used.
Using cloth-like silk as raw material, several hours are baked in a few Baidu's high temperature roasters, which can be electric oven or too
Sun can energize oven, preferably transparent glass door, can observe baking state in real time.Cloth-like silk raw material shelf is stacked, and avoids stacking
It is overstocked to cause baking temperature inhomogenous.
Unless otherwise specified, the reagent consumptive material in the present invention can be obtained by commercially available purchase, be set used in the present invention
The standby conventional equipment that can be used in fields is carried out referring to the prior art of fields.
The invention has the following advantages that
1, burnt silk cocoon hemostatic material biocompatibility provided by the invention is high, has significant anastalsis, may be used as
The big bleeding choke material of deep and big wound, the hemostatic material of the multiple uses such as clinical operation hemostasis and skin surface local hemorrhage
Material has quickly water suction hemostasis, the quickly hemostasis of agglutination red blood cell and highly-safe effect；The most key, the present invention is burnt
Silk cocoon hemostatic material is easy to get, and cost is relatively low, and cost performance is high, and carrying portable, not with the tissue easy debridement of adhesion, can satisfy and faces
Bed demand.
2, the preparation section of hemostatic material of the present invention is few, simple process, can be significantly reduced in large-scale production artificial
Cost and time cost.
Detailed description of the invention
Fig. 1 is the cell toxicity test of more than a kind of cloth coke silk cocoon leaching liquor of embodiment；
Fig. 2 is to measure in the external clotting time (CT) of more than 2 kinds of cloth coke silk cocoons of embodiment；
Fig. 3 is that the thrombelastogram (TEG) of 3 cloth coke silk cocoon of embodiment is tested；
Fig. 4 is that the blood platelet flow cytometer detection of 4 cloth coke silk cocoon of embodiment is tested；
Fig. 5 is the hemostasis that more than 5 kinds of cloth coke silk cocoons of embodiment are used for small Rat common carotid Rupture Model；
Fig. 6 is that hemostasis of the 6 cloth coke silk cocoon of embodiment for pig femoral artery Slicing Model for Foreign is tested；
Fig. 7 is the hemostasis blood routine that 6 cloth coke silk cocoon of embodiment is used for pig femoral artery Slicing Model for Foreign；
Fig. 8 is hemostasis blood coagulation four that 6 cloth coke silk cocoon of embodiment is used for pig femoral artery Slicing Model for Foreign
Fig. 9 is that TEG of the 6 cloth coke silk cocoon of embodiment for pig femoral artery cutting bleeding model is tested
Figure 10 is the hemostasis blood pressure that 6 cloth coke silk cocoon of embodiment is used for pig femoral artery Slicing Model for Foreign
Figure 11 is the hemostasis blood pressure that 6 gauze of embodiment is used for pig femoral artery Slicing Model for Foreign
Specific embodiment
In order to illustrate more clearly of the present invention, the present invention is done further below with reference to preferred embodiments and drawings
It is bright.Below with reference to preferred embodiment, the present invention is described further.It will be appreciated by those skilled in the art that institute is specific below
The content of description is illustrative and be not restrictive, and should not be limited the scope of the invention with this, further includes each specific reality
Any combination between mode is applied, those of ordinary skill in the art adopt technical solution of the present invention by reading description of the invention
Any equivalent transformation taken, all are covered by the claims of the invention.
Experimental method used in following embodiments is conventional method unless otherwise specified；Material used, reagent
Deng being commercially available unless otherwise specified.
The preparation of more than a kind of cloth coke silk cocoon of embodiment and its cell toxicity test of leaching liquor
The external clotting time (CT) of more than 2 kinds of cloth coke silk cocoons of embodiment measures
The thrombelastogram (TEG) of 3 cloth coke silk cocoon of embodiment is tested
The blood platelet flow cytometer detection of 4 cloth coke silk cocoon of embodiment is tested
More than 5 kinds of cloth coke silk cocoons of embodiment are used for the hemostasis of small Rat common carotid Rupture Model
Hemostasis of the 6 cloth coke silk cocoon of embodiment for pig femoral artery Slicing Model for Foreign is tested
The preparation and its leaching liquor cell toxicity test of more than a kind of cloth coke silk cocoon of embodiment
The preparation of more than a kind of cloth coke silk cocoon
1) silk cocoon removes silkworm chrysalis, to remove the silk cocoon of silkworm chrysalis as raw material；
2) the silk cocoon drawing of removal silkworm chrysalis is made into cloth-like silk.The way of cloth-like silk is exemplified below: drawing is knitted in advance that bamboo chip is complete
It at arciform, and is fixed on one piece of bottom plate, in this, as the small bow for drawing woven fabric-like silk；When drawing is knitted, it is torn by hand silk cocoon, removes cocoon
The sundries such as pupa, clean up, and soften silk cocoon, open cocoon mouth, are placed on bow, silk is pulled to the bottom plate of bow, be allowed to form cloth-like silk,
Drawing is knitted repeatedly, after overworking small bow, is removed pouch-shaped silk floss from bamboo bow, is cleaned and dry；
3) cloth-like silk raw material is baked several hours in a few Baidu's high temperature roasters, for example, 230 DEG C baking 2-3 hours, with not
Subject to black wood charcoal or preservation styptic activity (such as khaki or isabelline, water content is about 4%)；
4) the cloth-like coke silk cocoon after being baked is cut into specific shape, is prepared into suitable clinical operation use or local skin goes out
The form now used through Co-60 radiation sterilization 10-30h, or uses other sterilization methods.
The cell toxicity test of more than 2 kinds of cloth coke silk cocoon leaching liquors
The preparation method of leaching liquor: all operations of this experiment are completed in Biohazard Safety Equipment.Extract liquid proportional reference
The preparation of the 12nd sample segment of national standard BiologicalEvaluationofMedicalDevice is executed with the requirement in reference sample, and extraction should use
Asptic technique carries out in sterile, chemically inert closed container.By the cloth-like coke silk cocoon to sterilize with the ratio of 0.1g/mL
It immerses in serum-free RPMI 1640 culture medium, is extracted for 24 hours with ParafilmTM and being placed under 37 DEG C of constant temperatures, take out sample
It is filtered after product with 0.2 μm of sterile filters, to remove the particulate matter in leaching liquor.
Cell culture and detection: by the L929 cell after passage after trypsin digestion, fresh medium is added, instead
Multiple piping and druming is evenly distributed on cell in culture solution, is counted under inverted microscope using blood cell counting plate, cell is suspended
Liquid is diluted to 104~105A cell/mL, is seeded in respectively on 96 orifice plates, each 100 μ L of hole, and one group of control group is arranged.Then
Inoculated 96 orifice plate is placed in 37 DEG C of CO2It is cultivated in incubator for 24 hours, after cell is adherent, original culture solution is sucked out, is added
Enter sample leaching liquor, wherein the corresponding comparison liquid of 100 μ L is added in negative control group, and the complete of 0.3% phenol is added in positive controls
Culture medium places into biochemical cultivation case and continues to cultivate 48h.Culture plate is taken out, the 5mg/mL MTT solution of 10 μ L is added in every hole,
Continue to cultivate 4h at 37 DEG C, discard stoste, 150 hole μ L/ dimethyl sulfoxides are added, shake 10min, is surveyed using microplate reader
Absorption value when 570nm is calculate by the following formula opposite proliferation rate (relative growth rate, RGR): RGR (%)=A1/
A2 × 100%
In formula: A1: experimental group mean light absorbency A2: negative control group mean light absorbency
It is no cytotoxicity that experimental result, which is 0 grade, and it is qualification that 1 grade, which is slight cytotoxic,.Experimental result is 2 grades anti-
It answers, is moderate cytotoxicity, answers combination cell morphological analysis, overall merit.Experimental result is that 3-5 order reaction is severe cell
Toxicity, it is unqualified.
MTT colorimetric method is a kind of method for detecting cell survival and proliferation, and testing principle is in living cells mitochondria
Succinate dehydrogenase makes exogenous MTT be reduced to the bluish violet crystallization first a ceremonial jade-ladle, used in libation of water-insoluble and is deposited in cell, and dead cell
Without this function, first a ceremonial jade-ladle, used in libation in dmso solution cell, with extinction degree at ELISA instrument detection 570nm wavelength
Value, the numerical value can reflect living cells quantity indirectly.Within the scope of certain cell number, MTT crystallization content is directly proportional to cell number, the method
High sensitivity and economy.
The influence that cloth-like coke silk cocoon bleeding-stopping dressing leaching liquor grows cell is shown in Fig. 1, as can be seen from the figure
Opposite proliferation degree (RGR) of cloth-like coke silk cocoon powder bleeding-stopping dressing is 0 grade (no cytotoxicity RGR >=100) or 1 order reaction is (slight
Cytotoxicity RGR 75-99), Cytotoxic evaluation is qualification, and the Cytotoxic evaluation of 0.3% phenol of positive control is 3 grades
(RGR 25-49) or 4 order reactions (RGR 1-24) are reacted, are severe cytotoxicity.Therefore, our conclusion is cloth-like coke silk cocoon
Powder bleeding-stopping dressing is highly-safe, good biocompatibility, meets the requirement of bio-medical material.
The external clotting time (CT) of more than 2 kinds of cloth coke silk cocoons of embodiment measures
1) 25mg cloth-like coke silk cocoon hemostatic material is accurately weighed respectively, send the test tube bottom for being 2cm to diameter；
2) people's whole blood 3ml is acquired, autoblood enters syringe and starts timing, and blood is slowly injected into test tube bottom along tube wall
Portion, each test tube 1ml, mixes gently 3s, and then test tube is put into 37 DEG C of electric heating constant-temperature water-bath tanks；
3) a test tube is tilted every 5s, is not flowed until test tube is inverted blood, records whole blood clotting time；
4) under kindred circumstances, measurement is not added the negative control of material and adds Celox chitosan positive control hemostatic material
The external clotting time, and record；Every group of sample tests three parallel samples, assembly average respectively；
CT refers to blood from the time in vitro with foreign matter face contact to needed for solidifying.After blood is in vitro, contact negatively charged
Surface when, the XII factor is activated, and a series of coagulation factors activate in succession thereafter, finally fibrinogen is made to be changed into fiber
Albumen and blood coagulation.CT experiment is the first experiment that test medicine must carry out clotting mechanism whether there is or not influence.
Fig. 2's the result shows that, compared with negative control group (CT about 15min) and positive controls (CT about 10min), cloth-like
The clotting time of burnt silk cocoon (black mulberry, yellowish mulberry and black toothed oak) bleeding-stopping dressing substantially reduces (about 5min).
The thrombelastogram (TEG) of 3 cloth coke silk cocoon of embodiment is tested
1) fresh whole blood is added to sodium citrate anticoagulant tube, gently inverts upside down sodium citrate pipe 5 times to mix well, then
It is vertically disposed on table and waits test.It waits at least for 15min just to start to test and it is necessary to be tested (citric acid in 2h
Sodium sufficiently works the required time).
2) it before testing, is first moved on in the test tube of 25mg experimental material with the sodium citrate anticoagulation that pipettor draws 1ml, on
Lower inverted tubes 5 times to mix well.
3) positive control has 2 groups, first is that the standard kaolin activator sodium citrate anticoagulation reaction group of instrument configuration, two
It is the sodium citrate anticoagulation reaction group of the experimental material of Celox containing 25mg；
4) 20 μ l calcium chloride CaCl are added2Then solution is drawn 340 μ l blood from reaction tube again and is moved on into cup
It in cup, selects sample type " CK ", instrument starts to detect.
5) instrument output TEG figure and major parameter.
The principle of TEG thrombelastogram experiment: the final result based on coagulation process is to form blood clot, and blood clot
Physical characteristic decides whether there is normal coagulation.TEG is using Cytological Basis as mode, in a short time with a small amount of whole blood
Intracorporal coagulation process and fibrinolytic process are simulated, so that the participations such as overall monitor coagulation factor, blood platelet, fibrinogen is solidifying
Blood process overall picture.
Fig. 3, A and B are Celox positive control and standard kaolin activator, and C and D are black toothed oak coke silk cocoon and Hei Sangjiao silkworm
The TEG of cocoon schemes.
R (clotting time): it is the coagulation factor reaction time, main to reflect coagulation factor function, when the smaller expression blood coagulation of numerical value
Between it is shorter.It is to react from blood coagulation system starting until one section of incubation period between fibrin clot formation.The R value for scheming C and D is aobvious
Show cloth-like coke silk cocoon powder (JCJ) about 5min, considerably less than Celox control group group and standard kaolin activator control group
(10.3min and 7.5min).CI (blood coagulation composite index): reflecting overall coagulation, and the smaller reflection of value is low solidifying, is worth bigger reflection
Hypercoagulative state, the CI value of figure C and D shows that cloth-like coke silk cocoon (JCJ) is in normal range value, and the CI of Cleox is -4.7, is exceeded
Normal range (NR) (- 3 to 3 range).K value reflects the function and level of fibrinogen, when assessment blood clotting bulk strength reaches certain 20mm
Time, the smaller expression fibrinogen function of numerical value is stronger.Compared to Cleox, the fibrinogen function of cloth-like JCJ increases
By force.
The blood platelet flow cytometer detection of 4 cloth coke silk cocoon of embodiment is tested
CD61, CD62P- selectin and Intracellular Ca2+ concentration are all the indexs for reflecting platelet activation.CD61 is expressed in blood
On platelet, work in platelet activation and aggregation.CD62P- selectin is present on Platelet alpha granule film, mainly mediates grain
The adherency of cell and monocyte and blood platelet.Fluo-3-Am is high degree of specificity Ca2+ fluorescence indicator, can be delicately anti-
Reflect the variation of free intracellular calcium concentration.
Thrombocyte CD61 measurement
1) it takes people's health whole blood and sodium citrate anticoagulant with the mixing of 9:1 ratio, prepares anticoagulated whole blood.
2) whole blood is centrifuged 15min with 200g, takes 100 μ l of supernatant platelet-rich blood plasma in EP pipe, 10mg material is added
Material, 37 DEG C of incubation 30min.
3) 1ml PBS is added in every pipe, removes from EP pipe after sample is rinsed, and remaining liquid is centrifuged 15min with 1500g,
Remove supernatant.
4) the CD61 antibody of 50 μ l FITC fluorescent markers after dilution is added, is protected from light effect 20min for 4 DEG C after mixing.
5) PBS is then added to mix, is measured with BD Accuri C6 flow cytometer.
The measurement of blood platelet CD62P- selectin
Experimental procedure is measured with Thrombocyte CD61, in addition to above-mentioned steps 4) use the CD62P- selectin of FITC fluorescent marker instead
Antibody.
Intracellular calcium ion measurement
1) 5mM Fluo-3-Am ester mother liquor is prepared with high quality, anhydrous DMSO.
2) when testing, DMSO mother liquor or packing liquid is dissolved, room temperature is returned back to.Prepare 20 μM of working solutions (such as Hanks and
Hepes buffer).
3) it takes people's health whole blood and sodium citrate anticoagulant to mix with the ratio of 9:1, prepares anticoagulated whole blood.
4) whole blood is centrifuged 15min with 200g, takes 100 μ l of supernatant platelet-rich blood plasma in EP pipe, 10mg material is added
Material.
5) isometric dyestuff working solution is added into the EP pipe containing material, 10 μM of final concentration.
6) 37 DEG C of incubation 1h of the room temperature of plate containing dyestuff, therebetween slight oscillatory 3 times are added without calcium buffer 1ml rinse material, from
The heart removes extra dyestuff, then is resuspended with no calcium buffer, with BD Accuri C6 flow cytomery.
Fig. 4, the blood platelet flow cytometer detection of cloth coke silk cocoon the experimental results showed that, cloth-like coke silk cocoon all makes blood platelet
CD61 (Fig. 4 A), CD62P- selectin (Fig. 4 B) and intracellular calcium concentration (Fig. 4 C) increase, and show that blood platelet activates.
The small Rat common carotid of more than 5 kinds of cloth coke silk cocoons of embodiment ruptures hemostasis
1) amobarbital mouse is injected intraperitoneally, is anaesthetized；
2) anesthetized mice is faced upward placement, and four limbs are fixed, cut with surgical instrument (microforceps and microscissors) along neck median line
1cm is opened, exposes arteria carotis communis through separation below nutator, dissociate about 0.5cm, arteria carotis communis is separated, with microscissors cross
To cutting arteria carotis communis；
3) arteria carotis communis, bloodletting 10s are broken, a variety of cloth coke silk cocoon bleeding-stopping dressings are placed at bleeding；
4) when the non-oozing of blood of hemostatic material, record hemostasis is time-consuming.
Fig. 5, the small Rat common carotid brake test of a variety of cloth coke silk cocoons is the result shows that Hei Sangjiao silk cocoon and black toothed oak coke silkworm
The femoral artery bleeding hemostasis time-consuming (1.5min) of cocoon is approximately the half (Fig. 5 A) of Celox (2.5min), hemostasis and hemostasis knot
Beam is shown in that Fig. 5 B and Fig. 5 C, cloth coke silk cocoon, easily by debridement, can completely be taken out after quick-acting haemostatic powder respectively.
The hemostasis of the pig femoral artery Slicing Model for Foreign of 6 cloth coke silk cocoon of embodiment is tested
Experimental method
1) experimental animal
Experiment Ba-Ma mini pig, 35 ± 5kg of weight (same to gender).
2) experimental group
Cloth-like hemostatic material and blank gauze negative control group.
3) instrument and equipment
Animal blood pressure instrument, Medlab, Nanjing Calvin Biotechnology Co., Ltd；
ECG monitor (PM-9000Vet)；
Anesthetic gas machine (Matrx, MODEL 3000)；
Conventional surgical instrument, constant operation table.
4) anesthesia
After being anaesthetized in advance using Su Mian Xin II 0.1ml/kg, promoting the circulation of qi cannula, isoflurane breathing are anaesthetized, root in experimentation
According to General reactions (heart rate, blood pressure, blink reflection, reaction etc. of wagging the tail) adjustment anesthesia soakage of animal.
5) experimental procedure
Dorsal position is fixed on 37 DEG C of constant operation tables, neck and iliac region preserved skin after Animal Anesthesia；
A bilateral common carotid artery is separated, is intubated, connection MP150 leads physiological monitor, is mainly used for dynamic monitoring experimentation more
The mean arterial pressure of middle animal；
It pops one's head in through tongue and connects ECG monitor, be mainly used for monitor heart rate, adjust anaesthesia dosage in due course；
Side femoral artery is separated, is found at femoral artery downstream branches, above the branch point at proximal part about 0.5cm, along blood
Pipe cross-sectional direction vertically cuts 1/2 perimeter vascular wall (releasing vasopasm in advance with 2% lidocaine), sees femoral artery in spray
Blood shape is projected, freely sprays blood 30s, amount of bleeding during collecting and record with blank gauze；
Be prepared in advance hemostatic material product, freely sprays after blood by specification operation immediately for tested products with certain
Pressure pressing acts at femoral artery injury bleeding, and primary pressing continues 3 minutes；
Withdraw from pressure after 3 minutes, whether observation wound still has active hemorrhage, if any then continuing to press a cycle with method
3 minutes.
Evaluation index
1) hemostatic healing efficacy evaluation index
A) amount of bleeding
The amount of bleeding of all generations during dressing stops blooding is collected with blank gauze.
Amount of bleeding=gauze collects weight-blank gauze weight after blood.
B) bleeding stopping period
Pressing period N (N≤3) needed for record product is finally reached haemostatic effect, and calculate last hemostasis and taken
Between.
Bleeding stopping period=3 minute × N
2) Safety Evaluation Index
A) blood routine
Whether the ingredients such as red blood cell, blood platelet occur significant change in the animal blood sample of evaluation hemostasis front and back.
B) blood coagulation four
Whether the coagulation indexes such as evaluation hemostasis front and back PT, APTT, FIB, TT occur significant change.
C) thrombelastogram (TEG) is tested
Whether the coagulation indexes such as evaluation hemostasis front and back thrombelastogram (TEG) occur significant change.
D) blood pressure
Evaluate the situation of change of blood pressure in hemostasis.
Cloth coke silk cocoon shows that the stock of cloth-like coke silk cocoon is dynamic for the hemostasis experiment effect (Fig. 6) of pig femoral artery Slicing Model for Foreign
Arteries and veins amount of bleeding is 51.57g, and the femoral artery amount of bleeding of gauze group is 134g, and cloth coke silk cocoon is in the hemostasis non-oozing of blood of 9min, easily quilt
Debridement can completely be taken out, and gauze group still oozing of blood.
Fig. 7-9 is blood routine, blood coagulation four and the thrombus elastic force that cloth coke silk cocoon is used for pig femoral artery Slicing Model for Foreign respectively
Figure (TEG) is as a result, show the ingredients such as red blood cell, blood platelet in the animal blood sample of hemostasis front and back, hemostasis front and back PT, APTT, FIB, TT
Significant change does not all occur for the coagulation indexes such as equal blood coagulations four indices and hemostasis front and back thrombelastogram (TEG).
Figure 10 and Figure 11 is the blood pressure of cloth and gauze coke silk cocoon for the hemostasis of pig femoral artery Slicing Model for Foreign respectively
Variation, the results showed that significant change does not occur for hemostasis front and back blood pressure.
Claims (10)
1. a kind of preparation method of cloth-like hemostatic material, it is characterised in that: using silk cocoon as raw material, silk cocoon pull into cloth-like be placed in it is roasting
It bakes, bakes to the quality of best haemostatic effect, after baking, cloth-like coke silk cocoon is trimmed to specific shape, and sterilization packaging finally obtains in case
There must be the cloth-like bleeding-stopping dressing of stronger anastalsis.
2. preparation method according to claim 1, it is characterised in that: the silk cocoon after baking is khaki or isabelline, is contained
Water is 3-5%.
3. preparation method according to claim 1, it is characterised in that: the baking condition be 200 DEG C be baked 2 hours or
230 DEG C are baked 2 hours for baking 30 minutes or 230 DEG C.
4. preparation method according to claim 1, it is characterised in that: silk cocoon is pulled into cloth-like silk.
5. the preparation method according to claim 4, it is characterised in that: cloth-like silk is placed in oven, which can be electricity
Oven or solar energy oven, preferably transparent glass door can observe roasted hemostasis auxiliary material quality in real time.
6. preparation method according to claim 1, it is characterised in that: sterilization method is that irradiation sterilization Co-60 irradiates 10-
30h or ethylene oxide sterilization processes, vacuum are protected from light packaging of aluminium foil bag, stick mark.
7. preparation method according to claim 1, it is characterised in that: the specific shape is that operation or patch is suitble to use
Hemostasis auxiliary material shape.
8. using any the method for claim 1-7 cloth obtained that stops blooding.
9. hemostasis cloth is for clinically surgical operation bleeding or skin surface local hemorrhage described in claim 8.
10. the application method for the cloth that stops blooding described in claim 8, including following several: similar hemostatic gauze is directly clogged, for hurting
The especially big and deep wound of the big bleeding of mouth；Or it is made into patch, for controlling serious skin surface part under contingency condition
Bleeding；Or the skin surface for bleedings such as lacerated wound, small incision and damaged wounds is locally disposed, or is used for other hemorrhagics
The hemostasis of type.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1407992A (en) * | 1999-12-09 | 2003-04-02 | 独立行政法人农业生物资源研究所 | Process for producing functional silk fibroin and utilization of the same |
| CN106317446A (en) * | 2016-09-14 | 2017-01-11 | 西南大学 | Preparation method of silk fiber fibroin hemostasis material |
-
2018
- 2018-10-25 CN CN201811251659.9A patent/CN109200329A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1407992A (en) * | 1999-12-09 | 2003-04-02 | 独立行政法人农业生物资源研究所 | Process for producing functional silk fibroin and utilization of the same |
| CN106317446A (en) * | 2016-09-14 | 2017-01-11 | 西南大学 | Preparation method of silk fiber fibroin hemostasis material |
Non-Patent Citations (2)
| Title |
|---|
| 中国药学会上海分会、上海市药材公司: "《药材资料汇编 下集》", 31 August 1959, 上海科技卫生出版社 * |
| 冉懋雄等: "《现代中药炮制手册》", 31 January 2002, 中国中医药出版社 * |
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Application publication date: 20190115 |