CN108187129A - A kind of absorbable gelatin styptic powder and preparation method thereof - Google Patents
A kind of absorbable gelatin styptic powder and preparation method thereof Download PDFInfo
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- CN108187129A CN108187129A CN201810124860.4A CN201810124860A CN108187129A CN 108187129 A CN108187129 A CN 108187129A CN 201810124860 A CN201810124860 A CN 201810124860A CN 108187129 A CN108187129 A CN 108187129A
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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
- A61L24/104—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
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/02—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques
- C08J3/03—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media
- C08J3/07—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media from polymer solutions
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/12—Powdering or granulating
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J9/00—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof
- C08J9/28—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof by elimination of a liquid phase from a macromolecular composition or article, e.g. drying of coagulum
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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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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2201/00—Foams characterised by the foaming process
- C08J2201/04—Foams characterised by the foaming process characterised by the elimination of a liquid or solid component, e.g. precipitation, leaching out, evaporation
- C08J2201/048—Elimination of a frozen liquid phase
- C08J2201/0484—Elimination of a frozen liquid phase the liquid phase being aqueous
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2207/00—Foams characterised by their intended use
- C08J2207/10—Medical applications, e.g. biocompatible scaffolds
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2389/00—Characterised by the use of proteins; Derivatives thereof
Abstract
The present invention provides a kind of absorbable gelatin styptic powder and preparation method thereof, the BET specific surface area of the gelatin styptic powder is 30m2/g~70m2The step of/g, preparation process includes carrying out crosslinking Treatment, water absorption and swelling, mechanical crushing, freeze-drying and separating twice to gelatin.The styptic powder can be attached on the surface of a wound well, and the quick moisture absorbed in blood realizes the effect well stopped blooding.And preparation method simple process, it is further advanced by using the crosslinking agent for being not involved in reaction, and mechanical crushing, ultrasonic centrifuge cycle cleaning or ethanol water cleaning operation are carried out after gelatin crosslinking and fully swelling, effectively remove crosslinking agent, cause crosslinking agent noresidue in styptic powder, avoiding presence irritation caused by organism of crosslinking agent influences, and improves the safety of product, has good promotion and application value.
Description
Technical field
The present invention relates to hemostatic material technical fields, relate more specifically to a kind of absorbable gelatin styptic powder and its preparation
Method.
Background technology
Surgical hemostasis is one of important step of clinical operation, good hemostatic technique and hemostatic material be ensure operation into
The key of work(.Currently used hemostatic material includes the variforms such as styptic sponge, hemostatic gauze, hemostasis gel, styptic powder,
Middle styptic powder is not influenced by surface of a wound size and position, cannot be only used for surgical wound surface, can be also used for the minimally invasive hand such as endoscope
Art has the advantages that uniqueness.
In the prior art, the hemostatic material of powdery is mainly the micropore polysaccharide product using starch to prepare raw material, is passed through
The technologies such as supercritical ultrasonics technology, wet heating processing, microwave method, Mechanical Method or enzyme perforation realize more microporous of starch particle surface,
The specific surface area and hydrophilicity of material are promoted, molecular sieve is played the role of by the micropore of microsphere surface, is adsorbed in blood
Moisture promotes the concentration of coagulation factor, accelerates the generation of clotting mechanism, so as to fulfill anastalsis.The material of this kind of product is
Natural polysaccharide, nonhazardous and biocompatibility is preferable, but there are preparation process it is complicated, easily cause environmental pollution, water absorbing properties
The problems such as not good enough, with surface of a wound poor adhesion.
Gelatin is as a kind of natural water-soluble Biodegradable polymer material, structure and bio-tissue structure
It is similar, there is good biocompatibility, while can realize that biodegradable absorb does not generate inflammatory reaction, be widely used in curing
In medicine field.Wherein, in terms of hemostatic material, absorbable gelatin sponge, injectable gelatin hemostasia products is common are, but are met
There is not been reported for the gelatin class styptic powder of Clinical practice requirement.
In addition, during gelatin class hemostasia products are prepared, in order to meet the requirement of mechanical strength and/or degradability, usually
It needs to carry out crosslinking Treatment to it.Last crosslinking agent can be present in final products, and in the subsequent degradation process of gelatin materials
It gradually discharges, bringing tissue irritation, possibly even there are carcinogenicities, and the safety of product is affected.
Therefore, a kind of gelatin class absorbable hemostatic powder with good haemostatic effect is researched and developed, is further carried on this basis
The security performance of high styptic powder, has great importance.
Invention content
It is of the invention in view of the deficiencies of the prior art, it is an object of the present invention to provide a kind of absorbable gelatin styptic powder
Gelatin styptic powder has very high specific surface area, can be attached on the surface of a wound well, and the quick moisture absorbed in blood is realized
Good haemostatic effect.
Another object of the present invention is to provide a kind of preparation method of absorbable gelatin styptic powder, the preparation method work
Skill is easy, and styptic powder obtained has good performance and haemostatic effect, and can significantly reduce crosslinking agent in styptic powder
Content promotes the safety of product, has good promotion and application value.
In order to solve the above technical problems, the technical solution adopted in the present invention is：
A kind of absorbable gelatin styptic powder, the BET specific surface area of the styptic powder is 30 m2/g ~70m2/ g, prepared
The step of journey includes carrying out crosslinking Treatment, water absorption and swelling, mechanical crushing, freeze-drying and separating twice to gelatin.
The saturated water absorption of the gelatin styptic powder is 50% ~ 200%, and volume sweell(ing) rate is 250% ~ 550%.
A kind of preparation method of absorbable gelatin styptic powder, includes the following steps：
S1. aqueous gelatin solution is prepared；
S2. crosslinking agent is added in institute's gelatin water solution, carries out crosslinking Treatment；
S3. the gelatin after crosslinking Treatment is placed in water and is fully swollen；
S4. the gelatin after being swollen in step S3 is divided into fritter, is then ground into micron-sized gelatin particle；
S5. the gelatin particle is subjected to cryogenic freezing, is freeze-dried after complete freezing, obtains dry gelatin；
S6. the gelatin after freeze-drying is subjected to separating twice, obtains absorbable gelatin styptic powder.
The present invention is adequately water-swellable by the way that the gelatin after crosslinking Treatment is carried out, and then carries out mechanical crushing, freezing is done
Dry and separating twice obtains absorbable hemostatic powder.The styptic powder obtained in this way can not only be attached on the surface of a wound well, had
In addition good water absorbing properties and haemostatic effect can realize that degradation absorbs in vivo completely.
Preferably, in step S1, the mass concentration of institute's gelatin water solution is 5% ~ 50%.
Preferably, in step S2, the dosage of crosslinking agent is the 4% ~ 20% of gelatin quality.It is further preferred that the crosslinking
Agent adds in during whisking gelatin solution.
Preferably, in step S4, the crushing is using water phase materials crusher.
Preferably, in step S6, the crushing speed of the separating twice is 10000r/min ~ 40000r/min, during crushing
Between be 10s ~ 5min.
Preferably, the preparation method, which further includes, packs obtained gelatin styptic powder and irradiation sterilization, you can
To the hemostasia products that can be applied on the surface of a wound.
Preferably, in order to reduce the residual quantity of crosslinking agent in the gelatin styptic powder, the safety of gelatin styptic powder is improved,
The present invention is using the crosslinking agent water-soluble carbodiimide for being not involved in reaction（EDC）Or glutamine transaminage（TG enzymes）, and
Crosslinking agent is removed by particular step in preparation method, the styptic power product crosslinking agent residual quantity finally obtained can be in 100ppm
Even 10ppm is non-stimulated to organizing hereinafter, safety is good.It is obtained especially by above-mentioned preparation method combination following steps：
（1）Crosslinking agent described in step S2 is water-soluble carbodiimide（EDC）Or TG enzymes, preferably EDC, the quality of crosslinking agent are
The 4% ~ 20% of gelatin quality, crosslinking agent add in whisking gelatin solution processes, and the mixing speed of gelatin solution is 100r/
Min ~ 5000r/min, preferably 1000r/min, the addition speed of crosslinking agent is 0.1mL/s ~ 10mL/s, preferably 2mL/s；
（2）Between step S4 and step S5, the gelatin particle is placed in pure water carry out the processing of ultrasound-centrifuge cycle or
It is cleaned repeatedly using ethanol water, then removes ethyl alcohol and being replaced in pure water and be fully swollen, will be crosslinked
Agent removes.
Preferably, the rotating speed of the centrifugation is 10000r/min ~ 20000r/min, time of single spin for 5min ~
10min, supersonic frequency are 25KHz ~ 130KHz, and the time of single sonication is 5min ~ 30min.
Preferably, the ethanol water is using ethyl alcohol:The volume ratio 4 of water:1 is prepared to obtain.
Gelatin crosslinking is to adjust a major way of gelatin degradation speed and anthemorrhagic performance, but usually adopt in the prior art
It is gelatin cross-linker with glutaraldehyde, and the rear crosslinking agents of crosslinking can not remove well, the irritation in later stage and potential can be brought
Safety issue.This method is operated by carrying out ultrasound-centrifuge cycle cleaning and separating twice after cross-linked gelatin swelling is crushed,
Gelatin cross-linker can be removed well, reached good hemostatic material purification effect, effectively avoided the residual of pungent
It stays and toxicity.
The crosslinking agent that the present invention selects is the water-soluble carbodiimide for being not involved in reaction（EDC）Or glutamine transaminage
（TG enzymes）, and be crosslinked rear crosslinking agents and can be removed from final system, crosslinking agent is not present in final product, is avoided
The presence of impurity irritation caused by organism influences and potential safety issue.
In addition, crosslinking agent causes, cross-linking reaction is very rapid, and disposable add in is easy to cause cross-linking reaction degree in system
Unevenness, being added dropwise can make system crosslinking uniform while stirring as possible.In cross-linking process, amino and carboxyl in gelatin lead to
The effect for crossing crosslinking agent forms amido bond, and the increase of system lactam bond can extend the degradation process of material, degradation time meeting
Extend with the increase of system internal crosslinker, so by the way that cross-linking reaction is controlled to can be very good the swelling ratio of modulating hemostasis powder
And degradation cycle.
Compared with prior art, the beneficial effects of the invention are as follows：
The present invention provides a kind of absorbable gelatin styptic powder and preparation method thereof, the gelatin styptic powder has very high ratio
Surface area can be attached on the surface of a wound well, and the quick moisture absorbed in blood realizes the effect well stopped blooding.And it makes
Preparation Method simple process, be further advanced by using be not involved in reaction crosslinking agent, and gelatin crosslinking and fully it is water-swellable after
Mechanical crushing, ultrasound-centrifuge cycle cleaning or ethanol water cleaning operation are carried out, effectively removes crosslinking agent so that hand over
Join agent noresidue in styptic powder, avoiding presence irritation caused by organism of crosslinking agent influences, and improves product
Safety has good promotion and application value.
Figure of description
Fig. 1 is the scanning electron microscope (SEM) photograph of the absorbable gelatin styptic powder of the present invention.
Fig. 2 is result of the leaching liquor culture cell of the absorbable gelatin styptic powder of embodiment 1 after 24 hours.
Fig. 3 is result of the leaching liquor culture cell of the absorbable gelatin styptic powder of embodiment 1 after 72 hours.
Specific embodiment
The present invention is further illustrated With reference to embodiment, but embodiment the present invention is not done it is any
The restriction of form.
Embodiment 1
A kind of preparation method of absorbable styptic powder, includes the following steps：
S1. existing gelatin 40g is taken, 60g water is added in, is placed in 80 DEG C of baking ovens and treats that gelatin is completely dissolved, being prepared into mass concentration is
40% aqueous gelatin solution；
S2. water-soluble carbodiimide（EDC）1.6g is dissolved in 10ml purified waters（EDC mass is the 4% of gelatin quality）, institute
Crosslinking agent is added in gelatin water solution while stirring, carries out crosslinking Treatment, wherein churned mechanically rotating speed is 1000r/min,
The addition speed 2mL/s of crosslinking agent；
S3. the gelatin after crosslinking Treatment is placed in water and is fully swollen；
S4. the gelatin after being swollen in step S3 is divided into fritter, micron is then ground into using water phase materials crusher
The gelatin particle of grade；
S5. gelatin particle is placed in pure water by way of centrifugal deposition and carries out ultrasound-centrifuge cycle processing, centrifugal rotational speed
15000r/min, single spin time 5min, ultrasonic frequency are 100KHz, and the time of single sonication is 10min, is recycled
Then the clean rear tiling of gelatin particle filtering elution is placed in low temperature refrigerator and freezes, treats to freeze completely laggard by number 3 times
Row freeze-drying, obtains dry gelatin；
S6. the gelatin after freeze-drying is subjected to separating twice, crushing speed is 30000r/min, and grinding time 1min is obtained
Absorbable gelatin styptic powder, filling in auxiliary instrument, irradiation is spare.
Fig. 1 is the surface texture scanning electron microscope (SEM) photograph of gelatin styptic powder made from embodiment 1, it can be seen that it is with microcosmic
Porous structure.
Embodiment 2
A kind of preparation method of absorbable styptic powder, includes the following steps：
S1. existing gelatin 20g is taken, 80g water is added in, is placed in 50 DEG C of baking ovens and treats that gelatin is completely dissolved, being prepared into mass concentration is
20% aqueous gelatin solution；
S2. water-soluble carbodiimide（EDC）2g is dissolved in 10ml purified waters（EDC mass is the 10% of gelatin quality）, institute
Crosslinking agent is added in gelatin water solution while stirring, carries out crosslinking Treatment, the churned mechanically rotating speed is 2000r/min,
The addition speed 5mL/s of crosslinking agent；
S3. the gelatin after crosslinking Treatment is placed in water and is fully swollen；
S4. the gelatin after being swollen in step S3 is divided into fritter, micron is then ground into using water phase materials crusher
The gelatin particle of grade；
S5. gelatin particle is placed in pure water by way of centrifugal deposition and carries out ultrasound-centrifuge cycle processing, centrifugal rotational speed
10000r/min, single spin time 5min, ultrasonic frequency are 25KHz, and the time of single sonication is 5min, and cycle is secondary
Number is 3 times, and then tiling after gelatin particle filtering elution totally is placed in low temperature refrigerator and is freezed, treats to freeze completely laggard
Row freeze-drying, obtains dry gelatin；
S6. the gelatin after freeze-drying is subjected to separating twice, crushing speed is 25000r/min, and grinding time 2min is obtained
Absorbable gelatin styptic powder, filling in auxiliary instrument, irradiation is spare.
Embodiment 3
A kind of preparation method of absorbable styptic powder, includes the following steps：
S1. existing gelatin 5g is taken, 95g water is added in, is placed in 50 DEG C of baking ovens and treats that gelatin is completely dissolved, being prepared into mass concentration is
5% aqueous gelatin solution；
S2. water-soluble carbodiimide（EDC）0.5g is dissolved in 10ml purified waters（EDC mass is the 10% of gelatin quality）,
Crosslinking agent is added in institute's gelatin water solution while stirring, crosslinking Treatment is carried out, wherein churned mechanically turn to gelatin solution
Speed is 2000r/min, the addition speed 5mL/s of crosslinking agent；
S3. the gelatin after crosslinking Treatment is placed in water and is fully swollen；
S4. the gelatin after being swollen in step S3 is divided into fritter, micron is then ground into using water phase materials crusher
The gelatin particle of grade；
S5. the gelatin particle after crushing is placed in pure water and ethanol solution to be stirred repeatedly, and the volume of wherein ethyl alcohol is the volume of water
Four times, after cleaning up repeatedly, completely remove ethyl alcohol phase, be dispersed in again water phase saturation swelling after, tiling be placed in Low-temperature Ice
It is freezed in case, completely after freezing, is placed in freeze-drying in freeze dryer and obtains dry gelatin；
S6. the gelatin after freeze-drying is subjected to separating twice, crushing speed is 40000r/min, and grinding time 3min is obtained
Absorbable gelatin styptic powder, filling in auxiliary instrument, irradiation is spare.
Performance detection
1st, crosslinking agent residues detection
The detection of crosslinking agent residual quantity is detected using LC-MS/MS, and LC-MS/MS uses tandem mass spectrum, can obtain molecular ion
Peak, and have fragment ion peak, thus can be used for carrying out the qualitative and quantitative analysis of material.Crosslinking agent residues detection result is such as
Shown in table 1, its content of crosslinking agent of gelatin styptic powder obtained is in 10ppm hereinafter, showing that the preparation method of the present invention can be notable
Content of crosslinking agent is reduced, greatly promotes the safety of product.
1 content of crosslinking agent testing result of table
| Serial number | Crosslinking agent | By inspection sample size/g | Detection method | Testing result/mg/kg |
| Embodiment 1 | EDC | 0.1157 | LC-MS/MS | 5.3 |
| Embodiment 2 | EDC | 1.9830 | LC-MS/MS | 9.8 |
| Embodiment 3 | EDC | 2.3627 | LC-MS/MS | 6.7 |
2nd, specific surface area test and saturated water absorption and the test of volume sweell(ing) rate
The test method of specific surface area：Product to be measured is taken to be put into the sample cell of analytical instrument, wherein, analytical instrument is quick complete
Automatic specific surface area and Porosimetry, model U.S. health tower NOVA 4200e.In low temperature（Liquid nitrogen bath）Under the conditions of, to sample
A certain amount of Adsorbate Gas is passed through in pipe（N2）, determine sample to absorption according to the variation of gas volume before and after absorption
Matter molecule（N2）Adsorbance；Solid matter is measured with reference to standard GB/T/T24533-2009-gas absorption BET principles
Specific surface area.
The calculation of specific surface area is：It is put into the sample in gaseous environment, material surface（Extra-granular and inside
The surface area in gap）Physical absorption will occur at low temperature.When absorption reaches balance, the adsorbed gas of equilibrium adsorptive pressure is measured
The scale of construction calculates sample mono layer adsorption amount, so as to calculate the specific surface area of sample according to BET equations.Wherein, BET
Equation is：
In formula：
P --- adsorbate divides, unit Pa；
P0--- adsorbent saturated vapor pressure, unit Pa；
V --- the practical adsorbance of sample, unit cm3；
Vm--- individual layer saturated extent of adsorption, unit cm3；
C --- with the relevant constant of sample adsorption capacity.
It is tested with gelatin styptic powder made from embodiment 1-3, test result is as shown in table 2.
The test method of saturated water absorption：By the sample of certain mass（m1）It is placed in culture dish, weighs up sample and add culture
The quality m of ware2, then add in and be preheated to（37±1）DEG C 0.9% physiological saline, the quality of physiological saline is material to be tested quality
20 times.Culture dish is moved into drying box,（37±1）60min is kept at DEG C.Culture dish is taken out, its careful inclination is fallen
Go out unabsorbed excessive moisture, until then adding culture dish with electronic balance precise sample under anhydrous drippage in 30s
Quality m3, parallel determination 3 times.By the saturated water absorption that can be calculated styptic powder（X）, wherein saturated water absorption（X）Meter
Calculating formula is：X=(m3-m2)/m1×100%。
The test method of volume sweell(ing) rate：The volume V of sample is measured by solvent completion method1.Then by sample（V1）It is placed in
It is equipped with certain volume, be preheated to（37±1）DEG C physiological saline graduated cylinder in, graduated cylinder is placed in（37±1）It is protected under DEG C environment
60min is held, the height that liquid level is caused to rise after then being added according to sample obtains the sample volume after the water suction of styptic powder saturation
V2, parallel determination 3 times.The volume sweell(ing) rate of styptic powder is obtained by calculation（Y）, volume sweell(ing) rate（Y）Calculation formula be：Y
=(V2-V1)/V1×100%。。
The specific surface area and swelling ratio test result of styptic powder are as shown in table 2.
2 styptic powder specific surface area of table and swelling ratio test result
| Serial number | BET specific surface area/m2/g | Volume sweell(ing) rate/% | Quality swelling ratio/% |
| Embodiment 1 | 30 | 296 | 53 |
| Embodiment 2 | 40 | 396 | 126 |
| Embodiment 3 | 70 | 512 | 180 |
3rd, hemostasis validity experiment
（1）Experimental design
Healthy new zealand rabbit 18 is selected, is divided into six groups, wherein three groups are experimental group, respectively experimental group 1, experimental group 2 and real
Test group 3, the gelatin hemostasis powder material that experimental group is prepared using embodiment 1 to embodiment 3.Other three groups are control group, respectively
Gauze control group, gelfoam control group and other brand styptic powder control groups, other described brand styptic powders stop for chitosan
Blood meal, producer are Nanning Bo Enkang bio tech ltd.
（2）Experimental method
Specific experiment method is：
After 1. rabbit is anesthetized, back fixation, abdomen upward, opens abdominal cavity, liver is fully exposed, with scalpel in liver table
Upper work about 1cm × 1cm wounds, cause liver trauma Hemorrhage Model；
2. after model foundation, take gauze of having weighed（w0）, the rapid Cover surface of a wound slightly firmly presses, starts simultaneously at timing, after 10s
Gauze is removed, weighs weight（w1）, w1-w0For bleeding numerical quantity；
3. covering hemostatic material rapidly, and gauze is covered on hemostatic material after removing gauze, gauze is firmly pressed, presses the phase
Between every 5s remove gauze observation surface of a wound bleeding, when loseing oozing of blood, stop timing, record bleeding stopping period.
（3）Experimental result
Bleeding stopping period is then judged to hemostasis failure more than 5min in experimentation, and every group carries out five experiments respectively, calculates every group and puts down
Mean value, bleeding stopping period and the haemostatic effect table of comparisons are as shown in table 3.
The hemostasis laboratory test results of table 3
| Group | Average bleeding | Average bleeding stopping period |
| Experimental group 1 | 2.1g | 1min55s |
| Experimental group 2 | 1.9g | 1min20s |
| Experimental group 3 | 3.0g | 1min40s |
| Control group 1 | 2.0g | ＞ 5min |
| Control group 2 | 2.5g | 1min40s |
| Control group 3 | 2.2g | 2min00s |
4th, cellulotoxic experiment
（1）Experimental design
Experimental group：Using complete medium：+ 10% fetal calf serum of DMEM culture mediums（FBS）+ 1% is dual anti-（Penicillin and streptomysin
Mixed liquor）, leaching liquor is added, adds in cell suspension, carries out cell culture.
Control group：Using complete medium：+ 10% fetal calf serum of DMEM culture mediums（FBS）+ 1% is dual anti-（Penicillin and strepto-
The mixed liquor of element）, cell suspension is added in, carries out cell culture.Leaching liquor is not added in control group, remaining cell culture condition with
Experimental group is identical.
Blank group：Using complete medium：+ 10% fetal calf serum of DMEM culture mediums（FBS）+ 1% is dual anti-（Penicillin and chain
The mixed liquor of mycin）, leaching liquor and cell suspension are not added in blank group, are positioned over and experimental group and control group in same time
In identical environment, reference during measuring absorbance value as experimental group and control group.
（2）Experimental method
Culture cell is contacted by leaching liquor, with L929For experimental cell, cell proliferation observation is evaluated prepared by embodiment 1
Hemostatic material is to the toxic effect of cell in vitro.
The detailed process of experiment is as follows：
1. use complete medium（+ 10% fetal calf serum of DMEM culture mediums（FBS）+ 1% is dual anti-（The mixing of penicillin and streptomysin
Liquid））According to the gelatin styptic powder of the ratio extraction embodiment 1 of 0.1g/mL, 100% leaching liquor is obtained, is carried out with 100% leaching liquor
Dilution, obtains 75% leaching liquor and 25% leaching liquor.
2. carrying out the preparation of leaching liquor using gelatin styptic powder made from embodiment 1, wherein extraction temperature is (37 ± 1)
DEG C, extraction time is (24 ± 2) h.Derect seething obtains being 100% leaching liquor, is diluted with 100% leaching liquor, can obtain
75% leaching liquor and 25% leaching liquor.Then L929 cells are resuspended using complete medium, are made a concentration of 4 × 104A cell/mL
Cell suspension.Using 96 hole plates as culture plate, wherein experimental group and control group are that above-mentioned cell suspension is added in culture plate,
100 μ L are added in per hole, complete medium of the blank group to add in 100 μ L per hole in culture plate is added without cell suspension.
3. the culture plate of experimental group, control group and blank group is placed in preculture 24 hours in incubator（At 37 DEG C, 5%
CO2Under the conditions of）.Then the leaching liquor of above-mentioned concentration gradient is added in into experimental group, 100 μ L, control group and blank are added in per hole
Group is not added with leaching liquor, each culture plate is placed in incubator cultivates 24 hours, 72 hours later（At 37 DEG C, 5%CO2Under the conditions of）.
Then the culture solution in hole is carefully sucked out, then the CCK-8 mixed liquors of 100 μ L are added in into every hole, mixed liquor is complete by 90 μ L's
The CCK-8 of+10 μ L of culture medium is mixed, then culture plate is placed in incubator and is incubated 2 hours.Then it is measured with microplate reader
Absorbance at 450nm.
（3）Experimental result
Fig. 2, Fig. 3 are in cellulotoxic experiment, and the absorbance value after blank group is individually subtracted in experimental group, control group（OD values）.From figure
Upper can be seen that is approached with the cell proliferation rate after leaching liquor culture with control group, and therefore, hemostatic material of the invention has good
Good safety, has no toxic side effect to cell growth.
Claims (10)
1. a kind of absorbable gelatin styptic powder, which is characterized in that the BET specific surface area of the styptic powder is 30 m2/g ~
70m2/ g, preparation process include carrying out crosslinking Treatment, water absorption and swelling, mechanical crushing, freeze-drying and separating twice to gelatin
The step of.
2. absorbable gelatin styptic powder according to claim 1, which is characterized in that the saturation water suction of the gelatin styptic powder
Rate is 50% ~ 200%, and volume sweell(ing) rate is 250% ~ 550%.
3. a kind of preparation method of absorbable gelatin styptic powder, which is characterized in that include the following steps：
S1. aqueous gelatin solution is prepared；
S2. crosslinking agent is added in institute's gelatin water solution, carries out crosslinking Treatment；
S3. the gelatin after crosslinking Treatment is placed in water and is fully swollen；
S4. the gelatin after being swollen in step S3 is divided into fritter, is then ground into micron-sized gelatin particle；
S5. the gelatin particle is subjected to cryogenic freezing, is freeze-dried after complete freezing, obtains dry gelatin；
S6. the gelatin after freeze-drying is subjected to separating twice, obtains absorbable gelatin styptic powder.
4. the preparation method of absorbable gelatin styptic powder according to claim 3, which is characterized in that in step S2, crosslinking
The dosage of agent is the 4% ~ 20% of gelatin quality.
5. the preparation method of absorbable gelatin styptic powder according to claim 3, which is characterized in that described in step S4
It crushes using water phase materials crusher.
6. the preparation method of absorbable gelatin styptic powder according to claim 3, which is characterized in that described in step S6
The crushing speed of separating twice is the r/min of 10000 r/min ~ 40000, and grinding time is 10s ~ 5min.
7. the preparation method of absorbable gelatin styptic powder according to claim 3, which is characterized in that further include：（1）Step
Crosslinking agent described in S2 is EDC or TG enzymes, and the quality of crosslinking agent is the 4% ~ 20% of gelatin quality, and crosslinking agent is molten in whisking gelatin
It is added in during liquid, the mixing speed of gelatin solution is 100 r/min ~ 5000r/min, and the addition speed of crosslinking agent is
0.1mL/s ~10mL/s；
（2）Between step S4 and step S5, the gelatin particle is placed in pure water carry out the processing of ultrasound-centrifuge cycle or
It is cleaned repeatedly using ethanol water, then removes ethyl alcohol and being replaced in pure water and be fully swollen, will be crosslinked
Agent removes.
8. the preparation method of absorbable gelatin styptic powder according to claim 7, which is characterized in that the rotating speed of the centrifugation
For 10000r/min ~ 20000r/min, the time of single spin is 5min ~ 10min；The frequency of ultrasound is 25KHz ~ 130KHz,
The time of single sonication is 5min ~ 30min.
9. the preparation method of absorbable gelatin styptic powder according to claim 7, which is characterized in that the ethanol water
It is using ethyl alcohol:Water is according to volume ratio 4:1 is prepared to obtain.
10. according to the preparation method of any one of claim 3 to the 9 absorbable gelatin styptic powder, which is characterized in that described
Preparation method, which further includes, packs obtained gelatin styptic powder and irradiation sterilization step.
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