CN104151572A - Method for preparing medical cross-linking sodium hyaluronate gel - Google Patents
Method for preparing medical cross-linking sodium hyaluronate gel Download PDFInfo
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Abstract
The invention discloses a method for preparing medical cross-linking sodium hyaluronate gel. The method comprises the following steps: mixing and reacting sodium hyaluronate with glycidyl ether under the alkali condition, namely, reacting for 1-4 hours at 40-60 DEG C firstly, further reacting for more than 48 hours at 15-30 DEG C to form the cross-linking sodium hyaluronate gel, washing, performing gelation and crushing the gel into grains of certain sizes, and subsequently sterilizing, thereby obtaining the medical cross-linking sodium hyaluronate gel. By adopting the method, the residual amount of a cross-linking agent can be reduced without a purification step by prolonging the cross-linking reaction time at low temperature, the difficulty that the residual cross-linking agent is hard to remove by using a conventional method is solved, and the prepared gel is uniform in grain and can be used in fields such as the cosmetic industry and the medicinal industry.
Description
Technical field
The present invention relates to a kind of medical cross-linking sodium hyaluronate gel and preparation method thereof, this cross-linking sodium hyaluronate gel is for medical science or beauty treatment fields.
Background technology
Hyaluronic acid (Hyalouronic Acid, hereinafter to be referred as HA) or its sodium salt be the straight-chain polysaccharide being connected to form again by D-glucuronic acid and N-acetyl-glucosamine disaccharide unit weight, be the important component of humans and animals skin, vitreum, joint lubrication liquid and cartilaginous tissue.Because thering is viscoelasticity, plasticity-and the biocompatibility of height, be widely used in medical treatment, beauty industry.But water-soluble extremely strong, easily diffusion and the degraded in tissue of natural HA, in body, retention time is shorter, so be restricted in application.In order to make up these defects, conventionally adopt and modify and crosslinked method, hyaluronan molecule is changed to structure, change its some attribute, obtain injectable, retention time is long, toxicity is little, the product of good biocompatibility.To be hyaluronic acid modify through linking agent the high-molecular gel obtaining to cross-linking hyaluronic acid sodium, can make up the features such as hyaluronic acid retention time is short, do not change its biocompatibility simultaneously.
The conventional preparation method of cross-linking sodium hyaluronate gel is uniformly mixed hyaluronate sodium and linking agent in basic solution, by linking agent, makes the incompatible preparation of chemical bond between hyaluronic acid macromolecular chain.But linking agent used remains in easily initiation inflammatory reaction in organism.If reduce the consumption of linking agent, gained cross-linking sodium hyaluronate gel visco-elasticity is low, the ability of the interior enzymolysis of opposed body is low, not soft, and the residence time is short in vivo.If dosage of crosslinking agent is high, corresponding prolongation of the residence time in vivo, but hardness increases, and syringeability reduces, and meanwhile, can make excessive linking agent residue in product, and very difficult purifying cleans.And the linking agent in removal jel product is also a large problem at present, the method of the most frequently used removal or reduction linking agent residual quantity comprises dialysis method or cleans with water or buffered soln, but content of crosslinking agent cannot effectively be removed or reduce to these class methods all, and the cycle is long, be difficult for realizing industrial production.Separately have a kind ofly, if prepare crosslinked hyaluronic acid solids, the conventional method of removing linking agent is the precipitator method, clean and make cross-linking sodium hyaluronate gel precipitation, but this method still cannot be removed residual free linking agent completely with organic solvent.Domestic a lot of patent (CN101759881, CN101502677, CN1590444, CN102558600 etc.) is all to use hyaluronate sodium and linking agent 1,4-butanediol diglycidyl ether (hereinafter to be referred as BDDE) reacts the gel making in basic solution, its Patent CN101759881 discloses the crosslinked HA of BDDE and has prepared medical cross-linking sodium hyaluronate gel derivative product, although retention time is long in the gelinite of preparation, Bc good, remove long, the very difficult residual quantity of removing or reduce linking agent completely of cycle of linking agent.
When the present invention utilizes prolongation low-temp reaction, chien shih crosslinking reaction is more abundant, then utilizes buffered soln and water washing, gelation, makes gel pass through the sub-sieve of certain order number under the adequately expanded state of gel, forms the uniform gel particle of particle diameter.
Summary of the invention
The invention provides a kind of method of manufacturing cross-linking sodium hyaluronate gel.Under alkaline environment, hyaluronate sodium first reacts 1～4 hour with linking agent at the temperature of 40 ℃ to 60 ℃, then at 15 ℃～30 ℃, react 48 hours above cross-linking hyaluronic acid sodiums that form, then through washing, the broken particle that forms a certain size, then sterilizing makes.
The object of the invention is to: a kind of simple method of preparing cross-linking sodium hyaluronate gel is provided, be applicable to scale operation, the method according to this invention need to be via purification step, the cross-linking sodium hyaluronate gel that can make that content of crosslinking agent is low in product, gel particle is even, impurity is few, do not increase heavy metal content, biocompatibility is high, to overcome in routine techniques the long and residual high shortcoming of linking agent of purifying cycle, be more suitable in medical cosmetology field.
For this reason, by the following technical solutions, it comprises the following steps in the present invention:
(1) take highly purified hyaluronate sodium as raw material, in being mixed with the basic solution of linking agent, stir, prior to 40 ℃～60 ℃ reactions 1～4 hour, then at 48 hours above cross-linking hyaluronic acid sodiums that form of 15 ℃～30 ℃ insulations of low temperature.
(2) by (1) to gel to be soaked in pH be in 7.0 PBS damping fluid 2～5 hours, then through deionized water wash, gelation.
(3) collect the aqueous gel that step (2) obtains, make it pass through 50～200 object screen clothes, be broken into a certain size gel particle particle.
(4) collect the aqueous gel particle that step (3) obtains, sterilizing, packing makes product.
The invention is characterized in the pyroreaction of first using the short period of time, then use long low-temp reaction condition to carry out crosslinking reaction.Pyroreaction temperature of the present invention refers to 40 ℃～60 ℃, and the best is 45 ℃～50 ℃; Low-temp reaction temperature of the present invention refers to 15 ℃～30 ℃, and the best is 20 ℃～25 ℃.Cross-linking sodium hyaluronate gel prepared by the present invention has the advantages such as linking agent residual quantity is low, the interior retention time of body is long.
The said alkaline environment of the present invention is to be provided by mineral alkali, comprises sodium hydroxide, potassium hydroxide, and sodium carbonate or salt of wormwood, wherein alkali concn is 0.05M～1.5M, wherein preferred 0.2M～0.5M.
Preparation method of the present invention, the molecular weight of hyaluronate sodium is 50～3,000,000 dalton, is preferably 100～1,750,000 dalton, and the mass percent in reaction mixture is 10%～30%, is preferably 12%～20%.
The present invention's linking agent used is BDDE or diglycidylether, and wherein the concentration of linking agent in mixture is 0.1w/v%～2.0w/v%.After the damping fluid that is 7.0 with pH in preparation method's step (2) soaks, use deionized water soaking and washing gel 6～48 hours.
The method that the aqueous gel of collecting described in step of the present invention (3) adopts is screen filtration or centrifugal.Comprising in addition dilution step, is to be required concentration and physiologically acceptable osmotic pressure by the cross-linked-hyaluronic acid dilution of gained in crosslinking reaction; Finally by sub-sieve, collecting median size is gel particle, the sterilizing of 100～300 μ m, obtains needed product.
Embodiment
For better understanding the present invention, below in conjunction with embodiment, the present invention is described further, but the present invention is not limited to the following example.
Embodiment 1
Get aqueous sodium hydroxide solution and 50 μ L linking agents 1 that 7.0ml concentration is 0.25M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,700,000 hyaluronate sodium 1.0g, stir and within 5 minutes, be placed in constant water bath box after 40 ℃ of reaction 3h, at 25 ℃, react 3 days again, after reaction finishes, to adding 100mL pH in products therefrom, it is 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 100 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Embodiment 2
Hyaluronic acid na concn=14w/v%.The BDDE of alkali concn=0.5M, high reaction temperature=50 ℃, low reaction temperatures=25 ℃, crosslinker concentration=1.0v/v%
Get aqueous sodium hydroxide solution and 70 μ L linking agents 1 that 7.0ml concentration is 0.5M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,700,000 hyaluronate sodium 1.0g, stir and within 5 minutes, be placed in constant water bath box after 50 ℃ of reaction 1h, at 25 ℃, react 3 days again, after reaction finishes, in gained reactant, add 100mL pH and be 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 100 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Embodiment 3
Get aqueous sodium hydroxide solution and 50 μ L linking agents 1 that 5.0ml concentration is 0.5M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,700,000 hyaluronate sodium 0.9g, stir in the constant water bath box be placed on temperature controllable for 5 minutes after 50 ℃ of reaction 1h, at 20 ℃, react 3 days again, after reaction finishes, in gained reactant, add 100mL pH and be 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 150 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Embodiment 4
Get aqueous sodium hydroxide solution and 50 μ L linking agents 1 that 5.0ml concentration is 0.25M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,000,000 hyaluronate sodium 0.9g, stir and within 5 minutes, be placed in constant water bath box after 40 ℃ of reaction 3h, at 25 ℃, react 3 days again, after reaction finishes, in gained reactant, add 100mL pH and be 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 150 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Embodiment 5
Hyaluronic acid na concn=18w/v%.The BDDE of alkali concn=0.3M, high reaction temperature=50 ℃, low reaction temperatures=20 ℃, crosslinker concentration=0.7v/v%
Get aqueous sodium hydroxide solution and 50 μ L linking agents 1 that 7.0ml concentration is 0.3M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,000,000 hyaluronate sodium 1.26g, stir and within 5 minutes, be placed in constant water bath box after 50 ℃ of reaction 1h, at 20 ℃, react 3 days again, after reaction finishes, in gained reactant, add 100mL pH and be 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 100 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Embodiment 6
Get aqueous sodium hydroxide solution and 50 μ L linking agents 1 that 7.0ml concentration is 0.25M, 4-butanediol diglycidyl ether, both mix, under magnetic agitation, adding molecular-weight average is 1,000,000 hyaluronate sodium 1.0g, stir and within 5 minutes, be placed in constant water bath box after 40 ℃ of reaction 3h, at 20 ℃, react 3 days again, after reaction finishes, in gained reactant, add 100mL pH and be 7.0 0.1M phosphate buffer soln soaking and washing repeatedly, so that obtain pH < 7.5 and physiologically acceptable osmotic pressure value, drain away the water, collect gel, by external force, make it pass through 150 object screen clothes, then sterilizing, can obtain cross-linking sodium hyaluronate gel product.
Above-described embodiment is only for the present invention is further elaborated, and is not to self-limit of the present invention.Thereby the modification that researcher in this field makes above-described embodiment is not to have exceeded the present invention.
Claims (10)
1. a preparation method for Medical sodium hyaluronate gel, is characterized in that, comprises the following steps:
(1) take high purity hyaluronate sodium as raw material, in being mixed with the basic solution of linking agent, stir, prior to 40 ℃～60 ℃ reactions 1～4 hour, then at 48 hours above cross-linking hyaluronic acid sodiums that form of 15 ℃～30 ℃ insulations.
(2) it is in 7.0 PBS damping fluid 2～5 hours that the gel (1) being obtained is soaked in pH, then through deionized water wash, gelation.
(3) collect the aqueous gel that step (2) obtains, make it pass through 50～200 object screen clothes, be broken into a certain size gel particle.
(4) collect the aqueous gel particle that step (3) obtains, sterilizing, packing makes product.
2. preparation method according to claim 1, wherein said alkaline environment is to be provided by mineral alkali, wherein alkali concn is 0.05M～1.5M.Wherein said mineral alkali is sodium hydroxide, potassium hydroxide, sodium carbonate, or salt of wormwood.
3. preparation method according to claim 1, the molecular weight of hyaluronate sodium used is 50～3,000,000 dalton, the mass percent of hyaluronate sodium in described mixture is 10%～30%.
4. gel process for preparing according to claim 1, the described linking agent of step (1) is BDDE or diglycidylether.The concentration of linking agent in mixture is 0.1w/v%～2.0w/v%.
5. preparation method according to claim 1, wherein crosslinking reaction is carried out 2～5 days.
6. preparation method according to claim 1, wherein the temperature in pyroreaction is 40 ℃～60 ℃.
7. preparation method according to claim 1, wherein the temperature of low temperature insulation reaction is 15 ℃～30 ℃.
8. preparation method according to claim 1, is characterised in that the method for collecting aqueous gel described in step (3) is to filter or centrifugal.The median size of collected gel particle is 100 μ m～300 μ m.
9. preparation method according to claim 1, comprises dilution step in addition, and it is to be required concentration and physiologically acceptable osmotic pressure by the cross-linked hyaluronic acid gel dilution of gained in crosslinking reaction.
10. the cross-linking sodium hyaluronate gel that preparation method according to claim 1 makes is for beauty treatment or medical use.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105497975A (en) * | 2015-12-17 | 2016-04-20 | 中国科学院长春应用化学研究所 | Preparation method and application of medical composite hydrogel dressing |
| CN106167549A (en) * | 2016-08-10 | 2016-11-30 | 浙江美华鼎昌医药科技有限公司 | A kind of preparation method of slow degraded cross-linked hyaluronic acid gel |
| CN107635656A (en) * | 2015-05-29 | 2018-01-26 | 花王株式会社 | The manufacture method of hydrogel particle |
| CN107793577A (en) * | 2017-11-29 | 2018-03-13 | 桂林华诺威生物科技有限公司 | A kind of preparation method of Medical sodium hyaluronate gel |
| WO2019033596A1 (en) * | 2017-08-16 | 2019-02-21 | 杭州协合医疗用品有限公司 | Method for preparing single-phase modified sodium hyaluronate gel |
| CN112812330A (en) * | 2021-01-08 | 2021-05-18 | 青岛琛蓝海洋生物工程有限公司 | Compound polysaccharide sodium hyaluronate gel and preparation method thereof |
| CN113214511A (en) * | 2020-01-21 | 2021-08-06 | 华熙生物科技股份有限公司 | Preparation method of colored cross-linked sodium hyaluronate gel and gel particles |
| CN114573841A (en) * | 2022-03-02 | 2022-06-03 | 杭州科腾生物制品有限公司 | Non-granular sodium hyaluronate linear cross-linking modification process method |
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2013
- 2013-05-16 CN CN201310180379.4A patent/CN104151572A/en active Pending
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107635656A (en) * | 2015-05-29 | 2018-01-26 | 花王株式会社 | The manufacture method of hydrogel particle |
| CN107635656B (en) * | 2015-05-29 | 2020-12-04 | 花王株式会社 | Method for producing hydrogel particles |
| CN105497975A (en) * | 2015-12-17 | 2016-04-20 | 中国科学院长春应用化学研究所 | Preparation method and application of medical composite hydrogel dressing |
| CN105497975B (en) * | 2015-12-17 | 2018-10-16 | 中国科学院长春应用化学研究所 | A kind of preparation method and applications of medical composite hydrogel dressing |
| CN106167549A (en) * | 2016-08-10 | 2016-11-30 | 浙江美华鼎昌医药科技有限公司 | A kind of preparation method of slow degraded cross-linked hyaluronic acid gel |
| WO2019033596A1 (en) * | 2017-08-16 | 2019-02-21 | 杭州协合医疗用品有限公司 | Method for preparing single-phase modified sodium hyaluronate gel |
| CN107793577A (en) * | 2017-11-29 | 2018-03-13 | 桂林华诺威生物科技有限公司 | A kind of preparation method of Medical sodium hyaluronate gel |
| CN113214511A (en) * | 2020-01-21 | 2021-08-06 | 华熙生物科技股份有限公司 | Preparation method of colored cross-linked sodium hyaluronate gel and gel particles |
| CN112812330A (en) * | 2021-01-08 | 2021-05-18 | 青岛琛蓝海洋生物工程有限公司 | Compound polysaccharide sodium hyaluronate gel and preparation method thereof |
| CN114573841A (en) * | 2022-03-02 | 2022-06-03 | 杭州科腾生物制品有限公司 | Non-granular sodium hyaluronate linear cross-linking modification process method |
| CN114573841B (en) * | 2022-03-02 | 2022-11-01 | 杭州科腾生物制品有限公司 | Non-granular sodium hyaluronate linear cross-linking modification process method |
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Application publication date: 20141119 |