CN107596431A - A kind of natural glycan Quito functional microsphere and Preparation method and use - Google Patents
A kind of natural glycan Quito functional microsphere and Preparation method and use Download PDFInfo
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Abstract
The invention discloses a kind of natural glycan Quito functional microsphere and Preparation method and use, its preparation method to be：Natural glycan is made into aqueous dispersions, adds carboxymethyl carbohydrate, the material with wound healing promoting effect, stirs, obtains aqueous phase；Emulsifying agent is dissolved in fat-soluble solvent, obtains oil phase, and aqueous phase is added dropwise in oil phase, stirring；Crosslinking agent, stirring is added dropwise；Stand, cleaned using the first polar solvent, remove oil phase, water layer is cleaned using second of polar solvent, is settled out microballoon, water layer is extracted, and microballoon is dried, and obtains a kind of natural glycan Quito functional microsphere.Microballoon of the present invention can effectively block bleeding site, promote hemostasis, without any chemical cross-linking agent or other auxiliary agents, biological safety is high, without haemolysis, can degrade absorption in a short time, product of the present invention also has the effect of preventing the growth of bacterium, promoting the quick healing of wound while surface of a wound hemostasis is completed.
Description
Technical field
The invention belongs to hemostatic material technical field, is related to biological medical polymer material and compatibility modifier field, tool
Body is related to the natural glycan Quito functional microsphere and Preparation method and use of a kind of degradable absorption.
Background technology
Uncontrollable bleeding is to cause the major reason of war, traffic accident and other Accidents deaths, at the same be also
The problem often to be faced in emergency care of trauma and surgical operation.How surface of a wound hemostasis is effectively completed, and this is for saving patient's life
Life has vital meaning, and therefore, fast and effectively hemostatic material turns into worldwide study hotspot for exploitation.
Powdery hemostatic material obtains extensively because its is easy to operate, surface of a wound fitting effect is good in surgery surgical hemostasis field
Application.At present, powdery hemostatic material is broadly divided into Aluminosilicates, chitosan class and biological products class, such as
Quickclot, Celox, HemCon, TachoComb H etc..However, Aluminosilicates hemostasia products are with preferably hemostatic
Can, it is a kind of not biodegradable product, but later stage application finds that such product easily largely generates heat, and causes tissue to be burnt,
Secondary injury is brought to patient.Poly- sugar products with sharpest edges be such product with good biocompatibility and
Degradability, the surface of the Arista microballoons of Medafor companies of U.S. production have fissured structure, enhance it for water
Adsorption capacity, chemical cross-linking agent epoxychloropropane, sodium tripolyphosphate have been used in preparation process yet with it, cause its
Due to the residual of crosslinking agent in degradation process, and certain toxicity is produced, so as to limit its application.Complete hemostatic function
Afterwards, microballoon easily causes bacteria breed and the infection of the surface of a wound, further delays the healing of wound because having stronger hydrophily.
Some products are anti-to realize by adding antipathogenic composition, such as silver ion, antibiotic, quaternary ammonium salt, chitosan antiseptic wherein
The purpose of bacterium.But silver ion has stronger cytotoxicity, the abuse of antibiotics also results in bad antibacterial effect,
Although the cationic antibacterial agents such as quaternary ammonium salt, chitosan have certain biocidal property, but do not possess broad-spectrum antiseptic function.And dopamine,
Catechol, tannic acid are all from naturally extracting product, have stronger bacteriostasis property, and may act as styptic, at present, there has been no
Make the report that crosslinking agent prepares natural glycan Quito functional microsphere with above-mentioned several natural extract matter.
The content of the invention
The purpose of the present invention is overcome the deficiencies in the prior art, there is provided a kind of anthemorrhagic performance is good, without any chemical crosslinking
Agent or other auxiliary agents, biological safety be high, without haemolysis, the absorption that can degrade in a short time, wound healing can be promoted, prevent surface of a wound bacterium
Natural glycan Quito functional microsphere of infection.
Second object of the present invention is to provide a kind of preparation method of natural glycan Quito functional microsphere.
Third object of the present invention is to provide a kind of purposes of natural glycan Quito functional microsphere.
Technical solution of the present invention is summarized as follows：
A kind of preparation method of natural glycan Quito functional microsphere, comprises the following steps：Natural glycan is dispersed in 70-90
DEG C ultra-pure water in, be made into mass fraction be 5%-30% dispersion liquid, then add equivalent to natural glycan quality 1%-6%
Carboxymethyl carbohydrate, add the material with wound healing promoting effect equivalent to natural glycan quality 0%-2%, stirring is equal
It is even, obtain aqueous phase；Emulsifying agent is dissolved in 45-75 DEG C of fat-soluble solvent, is made into the solution that mass fraction is 0.03%-1%
It is by volume 1 under the conditions of 30-40 DEG C, 500-1000rpm as oil phase:15-3 ratio, aqueous phase is added dropwise to oil phase
In, it is added dropwise, continues to stir 30min-120min；Mixing speed is down to 400-600rpm, is added dropwise equivalent to natural glycan
Quality 5%-30% crosslinking agent, is added dropwise, and mixing speed is adjusted into 500-1000rpm continues to stir 2-5h；Stand 10-
30min, cleaned 3-5 times using the first polar solvent, remove oil phase, water layer is cleaned 3-5 times using second of polar solvent, sunk
Formed sediment microballoon, and water layer is extracted, and microballoon is dried, and obtains a kind of natural glycan Quito functional microsphere.
Preferably, natural glycan is at least one in hydroxyethyl cellulose, chitosan, starch, sodium alginate and hyaluronic acid
Kind；
Preferably, starch is cornstarch, farina, tapioca or starch from sweet potato；The viscosity of the chitosan
For 200-400mPa.s, deacetylation 50%-80%；The sodium alginate viscosity is 200 ± 20mPa.s；The ethoxy
Cellulose viscosity is 1000-5000mPa.s；The molecular weight of the hyaluronic acid is 4 × 105-2.2×106Dalton.
Preferably, carboxymethyl carbohydrate is that viscosity is 200-6000mPa.s, and substitution value is 80%-95% carboxymethyl chitosan
Sugar and viscosity are 600-4500mPa.s, and substitution value is at least one of 60%-95% carboxymethyl cellulose.
Preferably, the material with wound healing promoting effect is molecular weight 1 × 105-3×105The hyaluronic acid of dalton
With molecular weight 6 × 103At least one of dalton epithelical cell growth factor.
Preferably, emulsifying agent Tween-60, Tween-80, Span-60, Span-80, Span-85 and Triton X-
At least one of 100；The fat-soluble solvent is at least one of atoleine, vegetable oil, hexamethylene and n-hexane.
Preferably, crosslinking agent is at least one of catechol, tannic acid and dopamine.
Preferably, the first polar solvent is hexamethylene, n-hexane or petroleum ether.Second of polar solvent be water,
At least one of methanol and ethanol；The drying condition is 30-60 DEG C of vacuum drying or freeze-drying.
A kind of natural glycan Quito functional microsphere prepared by the above method.
A kind of purposes of the above-mentioned natural glycan Quito functional microsphere in styptic powder is prepared.
Advantages of the present invention：
Natural glycan Quito functional microsphere prepared by the present invention can effectively block bleeding site, promote hemostasis, anthemorrhagic performance
Good, without any chemical cross-linking agent or other auxiliary agents, biological safety is high, and no haemolysis, can degrade absorption in a short time, present invention production
Product can promote the wound healing of wound surface by loading hyaluronic acid, EGF, more so as to assign hemostatic microsphere
Can property.Product of the present invention also has the growth for preventing bacterium, promotes the quick healing of wound while surface of a wound hemostasis is completed
Effect.
Brief description of the drawings
Fig. 1 is the SEM shape appearance figure of natural glycan Quito functional microsphere.
Fig. 2 is that natural glycan Quito functional microsphere is used for SD injury of liver bleeding haemostatic effect figures.
Fig. 3 is the wound healing assay design sketch that natural glycan Quito functional microsphere is used for SD rat skins.
Fig. 4 is the 3rd day design sketch of wound healing that natural glycan Quito functional microsphere is used for SD rat skins.
Fig. 5 is the wound healing assay result figure that natural glycan Quito functional microsphere is used for SD rat skins.
Embodiment
With reference to specific embodiment, the invention will be further described, and the embodiment of the present invention is the skill in order that this area
Art personnel better understood when the present invention, but not carry out any restrictions to the present invention.
Embodiment 1
A kind of preparation method of natural glycan Quito functional microsphere, comprises the following steps：Natural glycan is dispersed in 70 DEG C
Ultra-pure water in, be made into mass fraction be 5% dispersion liquid, then add equivalent to natural glycan quality 1% carboxymethyl sugar
Class, stir, obtain aqueous phase；Emulsifying agent is dissolved in 45 DEG C of fat-soluble solvent, be made into mass fraction be 0.03% it is molten
Liquid is by volume 1 under the conditions of 30 DEG C, 500rpm as oil phase:15 ratio, aqueous phase is added dropwise in oil phase, is dripped
Finish, continue to stir 120min；Mixing speed is down to 400rpm, the crosslinking agent equivalent to natural glycan quality 5% is added dropwise, is added dropwise
Finish, mixing speed is adjusted into 500rpm continues to stir 5h；30min is stood, is cleaned 5 times, removed using the first polar solvent
Oil phase, water layer are cleaned 5 times using second of polar solvent, are settled out microballoon, water layer is extracted, and 30 DEG C of vacuum drying of microballoon, are obtained
To a kind of natural glycan Quito functional microsphere (Fig. 1), its particle diameter is 15-50um.
Natural glycan is that mass ratio is 1:1:1 hydroxyethyl cellulose, chitosan and cornstarch；Hydroxyethyl cellulose glues
Spend for 1000mPa.s；The viscosity of chitosan is 200mPa.s, deacetylation 50%；
Carboxymethyl carbohydrate is that viscosity is 200mPa.s, and substitution value is 80% carboxymethyl chitosan.
Emulsifying agent is that mass ratio is 1:1:5 Tween-60, Tween-80, Span-85；
Fat-soluble solvent is atoleine.
Crosslinking agent is that mass ratio is 1:2 catechol and tannic acid.
The first polar solvent is that volume ratio is 1:2 hexamethylene and n-hexane, second of polar solvent are ethanol；
Embodiment 2
A kind of preparation method of natural glycan Quito functional microsphere, comprises the following steps：Natural glycan is dispersed in 90 DEG C
Ultra-pure water in, be made into mass fraction be 30% dispersion liquid, then add equivalent to natural glycan quality 6% carboxymethyl sugar
Class, the material with wound healing promoting effect equivalent to natural glycan quality 2% is added, stirs, obtains aqueous phase；Emulsification
Agent is dissolved in 75 DEG C of fat-soluble solvent, be made into mass fraction be 1% solution as oil phase, in 40 DEG C, 1000rpm conditions
Under, it is by volume 1:3 ratio, aqueous phase is added dropwise in oil phase, is added dropwise, and continues to stir 30min；Mixing speed is dropped
To 600rpm, the crosslinking agent equivalent to natural glycan quality 30% is added dropwise, is added dropwise, by mixing speed be adjusted to 1000rpm after
Continuous stirring 2h；10min is stood, is cleaned 3 times using the first polar solvent, removes oil phase, water layer uses second of polar solvent
Cleaning 3 times, is settled out microballoon, water layer is extracted, microballoon freeze-drying, obtains a kind of natural glycan Quito functional microsphere.Particle diameter
For 20-60um.
Natural glycan is that mass ratio is 1:1:1 farina, sodium alginate, hyaluronic acid, the sodium alginate glue
Spend for 200 ± 20mPa.s；The molecular weight of hyaluronic acid is 4 × 105Dalton.
Carboxymethyl carbohydrate is that mass ratio is 1:1 viscosity is 6000mPa.s, carboxymethyl chitosan that substitution value is 95% and
Viscosity is 600mPa.s, and substitution value is 60% carboxymethyl cellulose.
Material with wound healing promoting effect is molecular weight 1 × 105The hyaluronic acid of dalton.
Emulsifying agent is that mass ratio is 4:1 Tween-80 and Span-60；Fat-soluble solvent is liquid soy oil.
Crosslinking agent is that mass ratio is 1:1 tannic acid and dopamine.
The first polar solvent is petroleum ether.Second of polar solvent is methanol.
Embodiment 3
A kind of preparation method of natural glycan Quito functional microsphere, comprises the following steps：Natural glycan is dispersed in 80 DEG C
Ultra-pure water in, be made into mass fraction be 15% dispersion liquid, then add equivalent to natural glycan quality 3% carboxymethyl sugar
Class, the material with wound healing promoting effect equivalent to natural glycan quality 1% is added, stirs, obtains aqueous phase；Emulsification
Agent is dissolved in 60 DEG C of fat-soluble solvent, be made into mass fraction be 0.5% solution as oil phase, in 35 DEG C, 800rpm bars
It is by volume 1 under part:5 ratio, aqueous phase is added dropwise in oil phase, is added dropwise, and continues to stir 60min；By mixing speed
Be down to 500rpm, the crosslinking agent equivalent to natural glycan quality 12% be added dropwise, is added dropwise, by mixing speed be adjusted to 700rpm after
Continuous stirring 3h；20min is stood, is cleaned 4 times using the first polar solvent, removes oil phase, water layer uses second of polar solvent
Cleaning 4 times, is settled out microballoon, water layer is extracted, and 60 DEG C of vacuum drying of microballoon, obtains a kind of natural glycan Quito functional microsphere.
Natural glycan is that mass ratio is 1:1:5 chitosan, tapioca (can also use starch from sweet potato to substitute para arrowroot
Powder) and hydroxyethyl cellulose；The viscosity of chitosan is 400mPa.s, deacetylation 80%；Hydroxyethylcellulose viscosity is
5000mPa.s。
Carboxymethyl carbohydrate is that viscosity is that mass ratio is 2:1 viscosity 5000mPa.s, substitution value are 90% carboxymethyl chitosan
Sugar and viscosity are 4000mPa.s, and substitution value is 90% carboxymethyl cellulose.
Material with wound healing promoting effect is that mass ratio is 5：1 molecular weight is 3 × 105Dalton hyaluronic acid and
Molecular weight is 6 × 103The epithelical cell growth factor of dalton.
Emulsifying agent is that mass ratio is 1:1 Span-80 and Triton X-100；
Fat-soluble solvent is that volume ratio is 1:2 hexamethylene and n-hexane.
Crosslinking agent is dopamine.
The first polar solvent is petroleum ether.Second of polar solvent is the ethanol solution that mass fraction is 80%.
Embodiment 4
A kind of preparation method of natural glycan Quito functional microsphere, comprises the following steps：Natural glycan is dispersed in 78 DEG C
Ultra-pure water in, be made into mass fraction be 10% dispersion liquid, then add equivalent to natural glycan quality 4% carboxymethyl sugar
Class, the material with wound healing promoting effect equivalent to natural glycan quality 0.5% is added, stirs, obtains aqueous phase；Breast
Agent is dissolved in 60 DEG C of fat-soluble solvent, be made into mass fraction be 0.6% solution as oil phase, in 40 DEG C, 800rpm
Under the conditions of, it is by volume 1:10 ratio, aqueous phase is added dropwise in oil phase, is added dropwise, and continues to stir 50min；Will stirring
Speed is down to 600rpm, and the crosslinking agent equivalent to natural glycan quality 5% is added dropwise, is added dropwise, and is 600rpm by mixing speed
Continue to stir 3h；20min is stood, is cleaned 5 times using the first polar solvent, removes oil phase, water layer is molten using second of polarity
Agent is cleaned 3 times, is settled out microballoon, water layer is extracted, and 50 DEG C of vacuum drying of microballoon, it is micro- to obtain a kind of natural glycan Quito function
Ball.Its particle diameter is 15-25um.
Natural glycan is that mass ratio is 3:1 sodium alginate and hyaluronic acid；
Sodium alginate viscosity is 200 ± 20mPa.s；The molecular weight of the hyaluronic acid is 2.2 × 106Dalton.
Carboxymethyl carbohydrate is that viscosity is that mass ratio is 2:1 5000mPa.s, substitution value be 90% carboxymethyl chitosan and
Viscosity is 4500mPa.s, and substitution value is 95% carboxymethyl cellulose.
Material with wound healing promoting effect is that quality is 4:1 molecular weight is 3 × 105Dalton hyaluronic acid and point
Son amount 6 × 103Dalton epithelical cell growth factor.
Emulsifying agent is Span-80；
Fat-soluble solvent is atoleine.
Crosslinking agent is tannic acid.
The first polar solvent is hexamethylene.Second of polar solvent is 70% methanol solution.
Experimental example 1
A kind of natural glycan Quito functional microsphere obtained to embodiment 1-4 carries out performance detection, and specific detection method is such as
Under：
Hemolysis rate：It is measured according to method as defined in GB/T 16175-1996.
After testing, the natural glycan Quito functional microsphere of the present invention has a good biocompatibility, the day in each embodiment
The hemolysis rate of right glycan Quito functional microsphere is respectively less than 5% (table 1), meets national standard.
Table 1 is unlike material hemolysis rate statistical result：
| Sample ID | Average value (X ± s) | Hemolysis rate (%) |
| Positive controls | 0.642±0.0387 | 100 |
| Negative control group | 0.048±0.0008 | 1.03 |
| Embodiment 1 | 0.048±0.0025 | 1.12 |
| Embodiment 2 | 0.048±0.0032 | 1.25 |
| Embodiment 3 | 0.048±0.0035 | 1.30 |
| Embodiment 4 | 0.048±0.0005 | 1.20 |
Note：1 positives control group of table is distilled water；Negative control group is physiological saline；Experimental group is that each embodiment is a kind of
The physiological saline leaching liquor of natural glycan Quito functional microsphere；X is absorbance under 545nm wavelength；S is that absorbance measurement is inclined
Difference.
Experimental example 2
Natural glycan Quito functional microsphere a kind of to each embodiment carries out external hemostatic function detection, specific detection method
For：SD rats are subjected to anaesthetic treatment, it is fixed, about 0.5cm × 1.0cm shearing wound is manufactured with operating scissors in its left lateral lobe of liver
Mouthful, after bleeding, spilt in bleeding part and apply the natural glycan Quito functional microspheres of 0.01g (blank control is without any processing).
Natural glycan Quito functional microsphere hemostatic microsphere prepared by embodiment 3 stops blooding (see Fig. 2), and can promote wound healing
(see Fig. 3).And there is certain adhesiveness, the effect that protective layer is formed at wound can be attached at.
After testing, the natural glycan Quito functional microsphere of the present invention effectively can stop blooding (table 2) in 36-76s.
A kind of natural glycan Quito functional microsphere that table 2 is embodiment 1-4 counts knot in the bleeding stopping period of SD rat livers
Fruit.
| Sample number into spectrum | Blank control | Embodiment 1 | Embodiment 2 | Embodiment 3 | Embodiment 4 |
| Bleeding stopping period (s) | 207±10 | 56±20 | 50±5 | 60±10 | 55±5 |
Embodiment described above is a kind of preferable scheme of the present invention, not the present invention is made any formal
Limitation, there are other variants and remodeling on the premise of without departing from the technical scheme described in claim.
It is good that a kind of new type natural glycan Quito functional microsphere combination property of gained is prepared according to this technology, it is easy to operation
And preservation, and preparation technology is simple, is adapted to industrialized production.
Experimental example 3
The wound healing promoting that skin is carried out to the multifunctional hemostatic microballoon provided in embodiment 2 and embodiment 4 is tested, specifically
Experimental method is：SD rats are anaesthetized, it is fixed, make a call to a depth about 2mm using a diameter of 5mm card punch at its back
Skin wounds (see Fig. 5 first row), then by the full-filling of 0.02g hemostatic microspheres at its wound (see the of Fig. 5 secondary series
2nd, the third line), rat dorsum skin wound healing situation was observed at the 3rd day, the 7th day and the 14th day respectively, blank control group is not
Do any processing, the results showed that：Compared with control group, hemostatic microsphere provided by the invention can promote skin to a certain extent
Speed of wound healing, wherein, dotted line frame position represents situation about adding after hemostatic microsphere.Hemostatic microsphere provided by the invention, no
Only there is preferable haemostatic effect, and there is certain adhesiveness, the effect (Fig. 4) that protective layer is formed at wound can be attached at.
Claims (10)
1. a kind of preparation method of natural glycan Quito functional microsphere, its feature comprise the following steps：Natural glycan is dispersed in
In 70-90 DEG C of ultra-pure water, the dispersion liquid that mass fraction is 5%-30% is made into, is then added equivalent to natural glycan quality
1%-6% carboxymethyl carbohydrate, the material with wound healing promoting effect equivalent to natural glycan quality 0%-2% is added,
Stir, obtain aqueous phase；Emulsifying agent is dissolved in 45-75 DEG C of fat-soluble solvent, and it is 0.03%-1% to be made into mass fraction
Solution as oil phase, be by volume 1 under the conditions of 30-40 DEG C, 500-1000rpm:15-3 ratio, aqueous phase is added dropwise
Into oil phase, it is added dropwise, continues to stir 30min-120min；Mixing speed is down to 400-600rpm, is added dropwise equivalent to day
Right glycan quality 5%-30% crosslinking agent, is added dropwise, and mixing speed is adjusted into 500-1000rpm continues to stir 2-5h；It is quiet
10-30min is put, is cleaned 3-5 times using the first polar solvent, removes oil phase, water layer cleans 3-5 using second of polar solvent
It is secondary, microballoon is settled out, water layer is extracted, microballoon is dried, and obtains a kind of natural glycan Quito functional microsphere.
2. according to the method for claim 1, it is characterised in that the natural glycan is hydroxyethyl cellulose, chitosan, shallow lake
At least one of powder, sodium alginate, hyaluronic acid；
3. according to the method for claim 2, it is characterised in that the starch is cornstarch, farina, para arrowroot
Powder or starch from sweet potato；The viscosity of the chitosan is 200-400mPa.s, deacetylation 50%-80%；The sodium alginate
Viscosity is 200 ± 20mPa.s；The hydroxyethylcellulose viscosity is 1000-5000mPa.s；The molecular weight of the hyaluronic acid
4 × 105-2.2×106Dalton.
4. according to the method for claim 1, it is characterised in that the carboxymethyl carbohydrate is that viscosity is 200-6000mPa.s,
The carboxymethyl chitosan and viscosity that substitution value is 80%-95% are 600-4500mPa.s, and substitution value is 60%-95% carboxylic first
At least one of base cellulose.
5. according to the method for claim 1, it is characterised in that the material with wound healing promoting effect is molecular weight
1 × 105-3×105The hyaluronic acid and molecular weight of dalton are 6 × 103In dalton epithelical cell growth factor at least
It is a kind of.
6. according to the method for claim 1, it is characterised in that the emulsifying agent is Tween-60, Tween-80, Span-
60th, at least one of Span-80, Span-85 and Triton X-100；The fat-soluble solvent be atoleine, vegetable oil,
At least one of hexamethylene and n-hexane.
7. according to the method for claim 1, it is characterised in that the crosslinking agent be catechol, tannic acid and dopamine in extremely
Few one kind.
8. according to the method for claim 1, it is characterised in that the first described polar solvent is hexamethylene, n-hexane or stone
Oily ether.Second of polar solvent is at least one of water, methanol and ethanol；The drying condition is 30-60 DEG C of vacuum
Dry or be freeze-dried.
A kind of 9. natural glycan Quito functional microsphere prepared by one of claim 1-8 method.
A kind of 10. purposes of the natural glycan Quito functional microsphere of claim 9 in styptic powder is prepared.
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