WO2022063312A1 - 透明质酸水凝胶和透明质酸膜及其制备方法和应用 - Google Patents

透明质酸水凝胶和透明质酸膜及其制备方法和应用 Download PDF

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WO2022063312A1
WO2022063312A1 PCT/CN2021/121184 CN2021121184W WO2022063312A1 WO 2022063312 A1 WO2022063312 A1 WO 2022063312A1 CN 2021121184 W CN2021121184 W CN 2021121184W WO 2022063312 A1 WO2022063312 A1 WO 2022063312A1
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hyaluronic acid
substituted
unsubstituted
acid derivative
preparation
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French (fr)
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董群
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吾奇生物医疗科技(江苏)有限公司
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Priority claimed from CN202011043357.XA external-priority patent/CN112159534B/zh
Priority claimed from CN202011043309.0A external-priority patent/CN112156234B/zh
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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
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/14—Macromolecular materials
    • A61L27/20—Polysaccharides
    • 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
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • 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
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/52—Hydrogels or hydrocolloids
    • 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
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/56—Porous materials, e.g. foams or sponges
    • 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
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/04—Macromolecular materials
    • 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
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/14—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
    • C08B37/0024—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid beta-D-Glucans; (beta-1,3)-D-Glucans, e.g. paramylon, coriolan, sclerotan, pachyman, callose, scleroglucan, schizophyllan, laminaran, lentinan or curdlan; (beta-1,6)-D-Glucans, e.g. pustulan; (beta-1,4)-D-Glucans; (beta-1,3)(beta-1,4)-D-Glucans, e.g. lichenan; Derivatives thereof
    • C08B37/0027—2-Acetamido-2-deoxy-beta-glucans; Derivatives thereof
    • C08B37/003—Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
    • 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
    • C08J3/075—Macromolecular gels
    • 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
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L5/00—Compositions of polysaccharides or of their derivatives not provided for in groups C08L1/00 or C08L3/00
    • C08L5/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof

Definitions

  • the invention belongs to the field of hyaluronic acid, and in particular relates to a hyaluronic acid hydrogel, a preparation method and application thereof, and a hyaluronic acid film and a preparation method and application thereof.
  • Hyaluronic acid is a natural transparent polysaccharide. It is a straight-chain polymer polysaccharide composed of glucuronic acid-N-acetic acid glucosamine as a disaccharide molecular unit. It has good biocompatibility. , biodegradability, bioactivity and rheological properties, and contain free carboxyl and hydroxyl groups, can be modified under mild conditions, has been favored.
  • Hydrogel is a kind of extremely hydrophilic three-dimensional network structure gel, which swells rapidly in water and can keep a large volume of water in this swollen state without dissolving.
  • the existing hyaluronic acid hydrogel is a bulk gel formed by random cross-linking of hyaluronic acid macromolecules, which has disadvantages such as low mechanical properties, inability to control release of biologically active molecules, or the use of toxic cross-linking agents. , which limits the application of hyaluronic acid hydrogels.
  • CN102942699A discloses a self-enhancing double-crosslinked hyaluronic acid hydrogel and a preparation method thereof.
  • the preparation method is divided into the following steps: preparation of double bond-activated hyaluronic acid, preparation of hyaluronic acid microspheres, preparation of double bond-activated hyaluronic acid microspheres, and preparation of self-enhancing double-crosslinked hyaluronic acid hydrogel.
  • the self-enhancing double-crosslinked hyaluronic acid hydrogel prepared by this method is prepared by reacting double-shoulder-activated hyaluronic acid microspheres as reinforcing particles with double-bond-activated hyaluronic acid molecules, and has a double-crosslinked network structure.
  • the diameter of the double bond-activated hyaluronic acid microspheres is 1 ⁇ m-10 ⁇ m
  • the pore size of the double cross-linked hyaluronic acid hydrogel is 10 ⁇ m-70 ⁇ m
  • the double bond substitution degree is 2.8%-65%.
  • the gel has good elastic properties and can prolong the sustained controlled release time of bovine serum albumin.
  • CN103724455A discloses a hyaluronic acid derivative and a preparation method thereof.
  • hyaluronic acid or its salt and acid anhydride are mixed and reacted in an anhydrous solvent, and the product is precipitated, separated and purified to obtain a cross-linkable hyaluronic acid.
  • Hyaluronic acid derivatives The preparation process of the method is simple, the reaction degree can be realized by controlling the reaction parameters, and the obtained product has stable performance and can be stored for a long time.
  • the obtained hyaluronic acid derivatives with cross-linkability can prepare hyaluronic acid hydrogels under mild conditions, have good biocompatibility, and can be used as bioactive substance carrier materials, tissue engineering scaffold materials and soft tissue filling materials .
  • CN108341976A discloses a derivative of a methacrylated matrix material based on copper-free catalytic click chemistry and its synthesis method and application.
  • the invention provides a click chemistry-based methacrylated base material derivative, two derivatives obtained by introducing an alkynyl group or an azide group into the methacrylated base material respectively.
  • These two materials can not only be covalently cross-linked under the action of photo-initiated radical polymerization to form hydrogels, but also the aqueous solutions of the two materials can be simply blended in a certain proportion to form hydrogels.
  • the two materials were prepared into hydrogels by photo-initiated free radical polymerization, and click chemistry can also occur through the alkynyl and azide groups on the surface of the hydrogel. , enabling the two hydrogels to bond on the surface.
  • CN111253591A discloses a double cross-linked hyaluronic acid hydrogel, its preparation method and application.
  • the preparation method includes: modifying hyaluronic acid with a modifier to obtain double-bonded hyaluronic acid; in the presence of a cross-linking agent and a photoinitiator, subjecting the double-bonded hyaluronic acid to physical treatment.
  • Cross-linking and photochemical cross-linking reactions were carried out to obtain double-cross-linked hyaluronic acid hydrogels.
  • the double-crosslinked hyaluronic acid hydrogel in the invention has good mechanical properties, and the hyaluronic acid derivative is self-crosslinked in the whole scheme, which effectively avoids the introduction of other macromolecular chains and solves the problem of hyaluronic acid hydrogel. Due to its low strength, it needs to be compounded with other macromolecular chain materials, which leads to the problem that its practical application is limited; and the invention uses a photoinitiator with better water solubility and less cytotoxicity, which has fast gel formation speed and double cross-linking. The mechanical properties of the obtained hydrogel have been greatly improved, and the maximum compressive stress can reach more than 2.5MPa.
  • the biomaterial film dosage form has macroscopic size, stable three-dimensional network structure and good mechanical strength, and is suitable for cosmetic applications, wound dressings, drug delivery preparations, various patches, anti-adhesion films, corneas, tissue engineering scaffolds, artificial organs, etc. .
  • the existing hyaluronic acid film has the disadvantage of low mechanical properties, or uses a toxic crosslinking agent, which greatly limits the application of the hyaluronic acid film.
  • CN105113054A discloses a preparation method of hyaluronic acid derivative cross-linked fiber.
  • the hyaluronic acid is grafted and modified through a ring-opening reaction to obtain a hyaluronic acid derivative with photopolymerization activity, and then a photoinitiator is added to mix After dissolving, freezing and crystallizing at low temperature, carrying out photocrosslinking polymerization at low temperature, and then removing the solvent by freeze drying to obtain a hyaluronic acid derivative crosslinked fiber material.
  • the hyaluronic acid derivative cross-linked fiber material with good mechanical properties and mechanical strength is obtained by this method, and no reagents that change the properties of hyaluronic acid are added in the preparation process, and some excellent properties of hyaluronic acid as a natural polymer will not be changed.
  • the cross-linked hyaluronic acid derivative fiber material can be widely used in drug release system, wound healing material, sewage treatment, heavy metal recovery, membrane separation, daily chemical industry and other fields. However, this method only performs photocrosslinking once, and the mechanical properties of the obtained hyaluronic acid derivative crosslinked fibers need to be further improved.
  • the purpose of the present invention is to provide a hyaluronic acid hydrogel and a preparation method and application thereof.
  • the hyaluronic acid hydrogel of the present invention has good elasticity, mechanical properties and fatigue resistance, and can be deformed under large deformation. It can withstand more than 10,000 cycles of compression without damage and fragmentation, effectively expanding the application range of hyaluronic acid hydrogels.
  • One of the objects of the present invention is to provide a kind of preparation method of hyaluronic acid hydrogel, in order to achieve this purpose, the present invention adopts the following technical scheme:
  • a preparation method of hyaluronic acid hydrogel comprising the steps:
  • step 2) adding a photoinitiator to the hyaluronic acid hydrogel combination obtained in step 1), and then performing a photocrosslinking reaction under illumination to obtain the hyaluronic acid hydrogel.
  • the hyaluronic acid hydrogel of the present invention adopts the cycloaddition reaction of the hyaluronic acid derivative M and the hyaluronic acid derivative F, and introduces a ring structure into the hyaluronic acid without adding other macromolecular chains.
  • the photocrosslinking reaction occurs when the light is illuminated, so that the crosslinked hydrogel has good elasticity, mechanical properties and fatigue resistance, and can withstand more than 10,000 cycles of compression under large deformation without Damaged and fragmented, effectively expanding the application range of hyaluronic acid hydrogels.
  • the cycloaddition reaction of the hyaluronic acid derivative M and the hyaluronic acid derivative F in the dark is the first cross-linking, and the hyaluronic acid hydrogel is cross-linked for the second time after adding the photoinitiator to light, and the obtained
  • the elasticity, mechanical properties and fatigue resistance of the hyaluronic acid hydrogel were further improved.
  • the anti-fatigue properties are based on the use of the conjugated diene structure on the hyaluronic acid derivative F as an electron donor due to electron delocalization. Under certain conditions, it can form Diels with the alkenyl group on the hyaluronic acid derivative M. - Alder reaction to form a six-membered heterocyclic ring, and the newly generated unsaturated double bond on the heterocyclic ring can undergo radical polymerization reaction under the condition of photoinitiator and specific wavelength of light, thereby obtaining double-crosslinked hydrogel.
  • the ring structure formed by the double cross-linked hyaluronic acid hydrogel at the connection site when the hydrogel is stimulated by external force, can absorb part of the energy through the rotation of the single bond to play the role of internal energy dissipation , and due to the steric hindrance of the ring, the mechanical movement inside the hydrogel is hindered to a certain extent, which enhances the resistance of the hydrogel to external force stimulation, so that it has good anti-fatigue performance and can withstand large deformations. Withstands multiple compressions without shattering.
  • step 1) the hyaluronic acid hydrogel conjugate includes modified hyaluronic acid whose structural formula is shown in formula 1;
  • R 1 is the residue part of the hyaluronic acid polymer that removes the hydroxyl group on C 6 ;
  • R 2 , R 3 , R 4 and R 6 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or any one of unsubstituted hydroxyl group, substituted or unsubstituted carboxyl group, substituted or unsubstituted amino group and substituted or unsubstituted sulfonic acid group;
  • R 5 , R 7 and R 9 are independently selected from oxygen, sulfur, substituted or unsubstituted Any one of the substituted alkylene groups;
  • R 8 is the residue part of the hyaluronic acid polymer that removes the carboxyl group.
  • step 1) the structural formula of the hyaluronic acid derivative M is shown in formula 2, and the structural formula of the hyaluronic acid derivative F is shown in formula 3.
  • R 1 is the residue part of hyaluronic acid polymer that removes the hydroxyl group on C 6 ;
  • R 2 , R 3 , R 4 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted Any of a hydroxyl group, a substituted or unsubstituted carboxyl group, a substituted or unsubstituted amino group, and a substituted or unsubstituted sulfonic acid group.
  • R 5 is the residue part of the hyaluronic acid polymer that removes the carboxyl group
  • R 6 , R 10 and R 11 are independently selected from any one of oxygen, sulfur, substituted or unsubstituted alkylene
  • R 7 , R 8 , R 9 , R 12 , R 13 , R 1 4 , R 15 and R 16 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted hydroxy, substituted or unsubstituted carboxyl, Any of a substituted or unsubstituted amino group and a substituted or unsubstituted sulfonic acid group.
  • step 1) the volume ratio of the hyaluronic acid derivative M and the hyaluronic acid derivative F is (0.5-1.5):(0.5-1.5); for example, the hyaluronic acid derivative M and the hyaluronic acid derivative
  • the volume ratio of the hyaluronic acid derivative F is 1:0.5, 1:0.7, 1:0.9, 1:1.1, 1:1.3 or 1:1.5, 0.5:1, 0.7:1, 0.9:1, 1.1:1, 1.3:1 or 1.5:1 etc.
  • the final mass concentrations of the hyaluronic acid derivative M and the hyaluronic acid derivative F in the reaction system are both 0.01-0.05 g/mL; for example, the hyaluronic acid derivative M and the hyaluronic acid derivative
  • the final concentration of F in the reaction system can all be 0.01 g/mL, 0.02 g/mL, 0.03 g/mL, 0.04 g/mL or 0.05 g/mL.
  • the time of the cycloaddition reaction is 6 to 24h, for example, the time of the cycloaddition reaction is 6h, 7h, 8h, 9h, 10h, 11h, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h or 24h, etc.;
  • the temperature of the cycloaddition reaction is 20 to 70°C, for example, the temperature of the cycloaddition reaction is 20°C, 30°C, 40°C, 50°C, 60°C or 70°C, etc.
  • the function of the photoinitiator is to act when the prepared hyaluronic acid hydrogel combination is prepared into a hyaluronic acid hydrogel.
  • the photoinitiator-added hyaluronic acid hydrogel combination is protected from light prior to the hyaluronic acid hydrogel.
  • the addition of the photoinitiator can be added before the preparation of the hyaluronic acid hydrogel combination, or after the preparation of the hyaluronic acid hydrogel combination, which belongs to the protection scope of the present invention.
  • step 2) the volume ratio of the hyaluronic acid derivative M, the hyaluronic acid derivative F and the photoinitiator is (8.5-9.5):(8.5-9.5):(1.5-2.5).
  • the mass concentration of the photoinitiator is 0.01-0.10g/mL, for example, the mass concentration of the photoinitiator is 0.01g/mL, 0.02g/mL, 0.03g/mL, 0.04g/mL, 0.05g/mL mL, 0.06g/mL, 0.07g/mL, 0.08g/mL, 0.09g/mL or 0.1g/mL, etc.
  • the photoinitiator is lithium phenyl(2,4,6-trimethylbenzoyl)phosphate, 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylbenzene Any one or a mixture of at least two of acetone and 1-hydroxycyclohexyl phenyl ketone.
  • a typical but non-limiting combination of the mixture is lithium phenyl(2,4,6-trimethylbenzoyl)phosphate, 2-hydroxy-4'-(2-hydroxyethoxy)-2-methyl Mixture of Propiophenone, Lithium Phenyl(2,4,6-Trimethylbenzoyl) Phosphate, Mixture of 1-Hydroxycyclohexyl Phenone, 2-Hydroxy-4'-(2-Hydroxyethoxy )-2-methylpropiophenone and 1-hydroxycyclohexyl phenyl ketone mixture, phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt, 2-hydroxy-4'-(2- A mixture of hydroxyethoxy)-2-methylpropiophenone and 1-hydroxycyclohexyl phenyl ketone.
  • the wavelength of the light source used in the photocrosslinking reaction is 360 nm to 450 nm, for example, the wavelength of the light source is 360 nm, 370 nm, 380 nm, 390 nm, 400 nm, 410 nm, 420 nm, 430 nm, 440 nm or 450 nm.
  • the illumination time of the illumination photocrosslinking reaction is 30s ⁇ 30min, for example, the illumination time is 30s, 40s, 50s, 60s, 1min, 5min, 10min, 15min, 20min, 25min or 30min, etc.; the illumination distance is 1-30cm, for example, the distance of light is 1cm, 2cm, 3cm, 4cm, 5cm, 6cm, 7cm, 8cm, 9cm, 10cm, 15cm, 20cm, 25cm or 30cm, etc.
  • step 1) the hyaluronic acid derivative M is obtained by modifying hyaluronic acid with 2-methacrylic anhydride.
  • the modification method is as follows: using 2-methacrylic anhydride solution and hyaluronic acid aqueous solution to react for 8 to 48 hours at 0 to 8° C. to obtain the hyaluronic acid derivative M; for example, the reaction temperature 0°C, 1°C, 2°C, 3°C, 4°C, 5°C, 6°C, 7°C or 8°C, etc., the reaction time is 8h, 10h, 12h, 15h, 16h, 18h, 20h, 24h, 26h, 28h, 30h, 32h, 35h, 36h, 38h, 40h, 42h, 46h or 48h etc.
  • the 2-methacrylic anhydride solution is a solution prepared by dissolving 2-methacrylic anhydride in acetone; the volume ratio of the 2-methacrylic anhydride to the acetone is 1:(1 ⁇ 5 ), for example, the volume ratio of the 2-methacrylic anhydride to the acetone is 1:1, 1:2, 1:3, 1:4 or 1:5, etc.
  • the hyaluronic acid aqueous solution is prepared by mixing hyaluronic acid and water; the mass concentration of the hyaluronic acid aqueous solution is 0.001-0.010 g/mL, for example, the mass concentration of the hyaluronic acid aqueous solution is 0.001 g/mL, 0.002g/mL, 0.003g/mL, 0.004g/mL, 0.005g/mL, 0.006g/mL, 0.007g/mL, 0.008g/mL, 0.009g/mL or 0.01g/mL, etc.
  • the molar ratio of the repeating units of 2-methacrylic anhydride in the 2-methacrylic anhydride solution to the hyaluronic acid in the hyaluronic acid aqueous solution is (0.54-5.42): 1, for example 2 -The molar ratio of repeating units of methacrylic anhydride to hyaluronic acid is 0.54:1, 0.55:1, 0.6:1, 0.8:1, 0.9:1, 1:1, 1.5:1, 2:1, 2.5 :1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.1:1, 5.2:1, 5.3:1, 5.4:1 or 5.42 etc.
  • the hyaluronic acid derivative M is further subjected to dialysis treatment and freeze-drying treatment.
  • the dialysis treatment is: using deionized water or 0.1M sodium chloride solution to dialysis the hyaluronic acid derivative M for 3-7 days, and the dialysis bag used in the dialysis treatment has a molecular weight cut-off of 3500 Da.
  • step 1) the hyaluronic acid derivative F is obtained by modifying hyaluronic acid with 2-furan methylamine.
  • the modification method is as follows: using a mixture of 2-furan methylamine and a carboxyl activator to react with a hyaluronic acid buffer solution at 15-40° C. for 8-72 hours to obtain the hyaluronic acid derivative F; for example, the reaction temperature is 15°C, 16°C, 18°C, 20°C, 25°C, 30°C, 35°C or 40°C, etc., and the reaction time is 8h, 10h, 12h, 15h, 18h, 20h, 24h, 25h , 28h, 30h, 32h, 36h, 38h, 40h, 42h, 46h, 48h, 50h, 52h, 56h, 60h, 64h, 66h, 68h, 70h or 72h, etc.
  • the hyaluronic acid buffer solution is prepared by adding hyaluronic acid to 80-120 mM 2-(N-morpholine)ethanesulfonic acid buffer solution, and the mass concentration of the hyaluronic acid buffer solution is 0.001 ⁇ 0.01g/mL, for example, the mass concentration of the hyaluronic acid buffer solution is 0.001g/mL, 0.002g/mL, 0.003g/mL, 0.004g/mL, 0.005g/mL, 0.006g/mL, 0.007g /mL, 0.008g/mL, 0.009g/mL or 0.01g/mL, etc.
  • the 2-(N-morpholine)ethanesulfonic acid buffer solution is prepared by dissolving 2-(N-morpholine)ethanesulfonic acid in water and adjusting the pH to 5.3-5.8, for example, the pH is 5.3, 5.4, 5.5, 5.6, 5.7 or 5.8 etc.
  • the molar ratio of the 2-furan methylamine to the repeating units of hyaluronic acid in the hyaluronic acid buffer solution is (1-6): 1, for example, the 2-furan methylamine and the transparent
  • the molar ratio of the repeating units of hyaluronic acid in the uronic acid buffer solution is 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5: 1, 5.5:1 or 6:1 etc.
  • the molar ratio of the carboxyl activator to the repeating units of hyaluronic acid in the hyaluronic acid buffer solution is (1-10):1, for example, the carboxyl activator and the hyaluronic acid buffer solution
  • the molar ratio of repeating units of hyaluronic acid is 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5: 1, 6:1, 7:1, 8:1, 9:1 or 10:1 etc.
  • the carboxyl activator is 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride.
  • the hyaluronic acid derivative F is further subjected to dialysis treatment and freeze-drying treatment.
  • the dialysis treatment is: using deionized water or 0.1M sodium chloride solution to dialysis the hyaluronic acid derivative F for 3-7 days, and the dialysis bag used in the dialysis treatment has a molecular weight cut-off of 3500 Da.
  • the second purpose of the present invention is to provide a hyaluronic acid hydrogel, which is prepared by the preparation method of the hyaluronic acid hydrogel described in the first purpose.
  • the hyaluronic acid hydrogel comprises a modified hyaluronic acid whose structural formula of the repeating unit is shown in formula 4.
  • R 5 , R 14 , R 18 and R 31 are the residues of the hyaluronic acid polymer by removing the hydroxyl group on C 6 .
  • R 1 , R 12 , R 21 and R 28 are the residue parts of the hyaluronic acid macromolecule with the carboxyl group removed.
  • R 2 , R 3 , R 8 , R 11 , R 13 , R 20 , R 23 , R 26 , R 27 , R 29 , R 32 , R 33 , R 34 and R 35 are independently selected from oxygen, Any of sulfur, substituted or unsubstituted alkylene.
  • R 4 , R 6 , R 7 , R 9 , R 10 , R 15 , R 16 , R 17 , R 19 , R 22 , R 24 , R 25 and R 30 are independently selected from hydrogen, substituted or unsubstituted Any of a substituted alkyl group, a substituted or unsubstituted hydroxyl group, a substituted or unsubstituted carboxyl group, a substituted or unsubstituted amino group, a substituted or unsubstituted amino derivative, and a sulfonic acid group.
  • the third object of the present invention is the application of the hyaluronic acid hydrogel described in the second object, and the hyaluronic acid hydrogel is used for the preparation of biomedical materials or tissue engineering materials.
  • the beneficial effects of the present invention include:
  • the hyaluronic acid hydrogel of the present invention adopts the cycloaddition reaction of the hyaluronic acid derivative M and the hyaluronic acid derivative F, and introduces a ring structure into the hyaluronic acid without adding other macromolecular chains. After adding the photoinitiator, the photocrosslinking reaction occurs when the light is illuminated, so that the crosslinked hydrogel has good elasticity, mechanical properties and fatigue resistance.
  • the compressive stress of the prepared hyaluronic acid hydrogel is 100kPa ⁇ 1.1MPa
  • the elastic modulus is 1 ⁇ 30kPa, under large deformation, it can withstand 10,000 cycles of cyclic compression without damage and fragmentation, effectively expanding the application range of hyaluronic acid hydrogel.
  • the purpose of the present invention is also to provide a hyaluronic acid film and a preparation method and application thereof.
  • the hyaluronic acid film of the present invention has better mechanical properties, and the obtained freeze-dried film has suitable microscopic pores and can be stretched at the maximum.
  • the stress is 4.5MPa, and the Young's modulus is 39.4MPa; the maximum tensile stress of the air-dried film is 145MPa, and the Young's modulus is 3.8GPa, which effectively expands the application range of the hyaluronic acid film.
  • the fourth object of the present invention is to provide a preparation method of a hyaluronic acid film, and for this purpose, the present invention adopts the following technical solutions:
  • a preparation method of hyaluronic acid film comprising the steps:
  • step 1) In the mixed system of the hyaluronic acid derivative M and the hyaluronic acid derivative F, a cycloaddition reaction occurs to obtain a hyaluronic acid pretreatment combination; 2)
  • the hyaluronic acid pretreatment compound obtained in step 1) After adding a photoinitiator to the treatment combination, stirring uniformly and removing air bubbles, it is cast in a mold to form a film, photocrosslinking reaction occurs under illumination, and then freeze-drying or air-drying to obtain the hyaluronic acid film.
  • the hyaluronic acid film of the present invention adopts the cycloaddition reaction between the hyaluronic acid derivative M and the hyaluronic acid derivative F, and in the case of avoiding the addition of other macromolecular chains, a ring structure is introduced into the hyaluronic acid, and light is added to the hyaluronic acid.
  • the photo-crosslinking reaction occurs after irradiation, and after freeze-drying/air-drying, the obtained hyaluronic acid film has good tensile mechanical properties and different microscopic pores, which effectively expands the application range of the hyaluronic acid film.
  • the tensile properties are based on the use of the conjugated diene structure on the hyaluronic acid derivative F, which can form Diels with the alkenyl group on the hyaluronic acid derivative M under certain conditions due to electron delocalization as an electron donor.
  • - Alder reaction to form a six-membered heterocyclic ring, and the newly generated unsaturated double bond on the heterocyclic ring can undergo radical polymerization reaction under the condition of photoinitiator and specific wavelength of light, thereby obtaining double cross-linked hyaluronic acid material.
  • the ring structure formed by the double cross-linked hyaluronic acid material at the connection site can absorb part of the energy through the rotation of the single bond when stimulated by an external force, thereby playing the role of internal energy dissipation.
  • the steric hindrance enhances the resistance of the membrane material to external force stimulation, making it have good tensile properties.
  • the hyaluronic acid pretreatment combination described in step 1) includes the modified hyaluronic acid whose structural formula is shown in formula 1;
  • R 1 is the residue part of the hyaluronic acid polymer that removes the hydroxyl group on C 6 ;
  • R 2 , R 3 , R 4 and R 6 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or any one of unsubstituted hydroxyl group, substituted or unsubstituted carboxyl group, substituted or unsubstituted amino group and substituted or unsubstituted sulfonic acid group;
  • R 5 , R 7 and R 9 are independently selected from oxygen, sulfur, substituted or unsubstituted Any one of the substituted alkylene groups;
  • R 8 is the residue part of the hyaluronic acid polymer that removes the carboxyl group.
  • step 1) the structural formula of the hyaluronic acid derivative M is shown in formula 2, and the structural formula of the hyaluronic acid derivative F is shown in formula 3.
  • R 1 is the residue part of hyaluronic acid polymer that removes the hydroxyl group on C 6 ;
  • R 2 , R 3 , R 4 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted Any of a hydroxyl group, a substituted or unsubstituted carboxyl group, a substituted or unsubstituted amino group, and a substituted or unsubstituted sulfonic acid group.
  • R 5 is the residue part of the hyaluronic acid polymer that removes the carboxyl group
  • R 6 , R 10 and R 11 are independently selected from any one of oxygen, sulfur, substituted or unsubstituted alkylene
  • R 7 , R 8 , R 9 , R 12 , R 13 , R 1 4 , R 15 and R 16 are independently selected from hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted hydroxy, substituted or unsubstituted carboxyl, Any of a substituted or unsubstituted amino group and a substituted or unsubstituted sulfonic acid group.
  • step 1) the volume ratio of the hyaluronic acid derivative M and the hyaluronic acid derivative F is (0.5-1.5):(0.5-1.5); for example, 1:0.5, 1:0.7, 1:0.9, 1:1.1, 1:1.3 or 1:1.5, 0.5:1, 0.7:1, 0.9:1, 1.1:1, 1.3:1 or 1.5:1 etc.
  • the final mass concentrations of the hyaluronic acid derivative M and the hyaluronic acid derivative F in the reaction system are both 0.01-0.05 g/mL; 0.03g/mL, 0.04g/mL or 0.05g/mL.
  • the time of the cycloaddition reaction is 6-24h, such as 6h, 7h, 8h, 9h, 10h, 11h, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h or 24h, etc.; the temperature of the cycloaddition reaction is 20-70°C, such as 20°C, 30°C, 40°C, 50°C, 60°C or 70°C, etc.
  • the function of the photoinitiator lies in preparing the prepared hyaluronic acid pretreatment combination into a hyaluronic acid film. Before carrying out the hyaluronic acid film, the hyaluronic acid pretreatment combination with added photoinitiator can be protected from light. It should be noted that the addition of the photoinitiator can be before the preparation of the hyaluronic acid pre-treatment combination, or after the preparation of the hyaluronic acid pre-treatment combination, no matter when it is added, it belongs to the protection scope of the present invention.
  • step 2) the volume ratio of the hyaluronic acid derivative M, the hyaluronic acid derivative F and the photoinitiator is (8.5-9.5):(8.5-9.5):(1.5-2.5).
  • the mass concentration of the photoinitiator is 0.01-0.10g/mL, such as 0.01g/mL, 0.02g/mL, 0.03g/mL, 0.04g/mL, 0.05g/mL, 0.06g/mL, 0.07g/mL, 0.08g/mL, 0.09g/mL or 0.1g/mL etc.
  • the photoinitiator is lithium phenyl(2,4,6-trimethylbenzoyl)phosphate, 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylbenzene Any one or a mixture of at least two of acetone and 1-hydroxycyclohexyl phenyl ketone.
  • a typical but non-limiting combination of the mixture is lithium phenyl(2,4,6-trimethylbenzoyl)phosphate, 2-hydroxy-4'-(2-hydroxyethoxy)-2-methyl Mixture of Propiophenone, Lithium Phenyl(2,4,6-Trimethylbenzoyl) Phosphate, Mixture of 1-Hydroxycyclohexyl Phenone, 2-Hydroxy-4'-(2-Hydroxyethoxy )-2-methylpropiophenone and 1-hydroxycyclohexyl phenyl ketone mixture, phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt, 2-hydroxy-4'-(2- A mixture of hydroxyethoxy)-2-methylpropiophenone and 1-hydroxycyclohexyl phenyl ketone.
  • the wavelength of the light source used in the photo-crosslinking reaction is 360 nm to 450 nm, for example, 360 nm, 370 nm, 380 nm, 390 nm, 400 nm, 410 nm, 420 nm, 430 nm, 440 nm or 450 nm.
  • the illumination time of the illumination photocrosslinking reaction is 30s ⁇ 30min, such as 30s, 40s, 50s, 60s, 1min, 5min, 10min, 15min, 20min, 25min or 30min, etc.; the illumination distance is 1 ⁇ 30cm, For example, 1cm, 2cm, 3cm, 4cm, 5cm, 6cm, 7cm, 8cm, 9cm, 10cm, 15cm, 20cm, 25cm or 30cm, etc.
  • step 1) the hyaluronic acid derivative M is obtained by modifying hyaluronic acid with 2-methacrylic anhydride.
  • the modification method is as follows: using 2-methacrylic anhydride solution and hyaluronic acid aqueous solution to react for 8 to 48 hours at 0 to 8° C. to obtain the hyaluronic acid derivative M; for example, the reaction temperature 0°C, 1°C, 2°C, 3°C, 4°C, 5°C, 6°C, 7°C or 8°C, etc., the reaction time is 8h, 10h, 12h, 15h, 16h, 18h, 20h, 24h, 26h, 28h, 30h, 32h, 35h, 36h, 38h, 40h, 42h, 46h or 48h etc.
  • the 2-methacrylic anhydride solution is a solution prepared by dissolving 2-methacrylic anhydride in acetone; the volume ratio of the 2-methacrylic anhydride to the acetone is 1:(1 ⁇ 5 ), for example, the volume ratio of the 2-methacrylic anhydride to the acetone is 1:1, 1:2, 1:3, 1:4 or 1:5, etc.
  • the hyaluronic acid aqueous solution is prepared by mixing hyaluronic acid and water; the mass concentration of the hyaluronic acid aqueous solution is 0.001-0.010 g/mL, such as 0.001 g/mL, 0.002 g/mL, 0.003 g/mL mL, 0.004g/mL, 0.005g/mL, 0.006g/mL, 0.007g/mL, 0.008g/mL, 0.009g/mL or 0.01g/mL, etc.
  • the molar ratio of the repeating units of 2-methacrylic anhydride in the 2-methacrylic anhydride solution to the hyaluronic acid in the hyaluronic acid aqueous solution is (0.54-5.42): 1, for example 0.54 :1, 0.55:1, 0.6:1, 0.8:1, 0.9:1, 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1 , 5:1, 5.1:1, 5.2:1, 5.3:1, 5.4:1 or 5.42 etc.
  • the hyaluronic acid derivative M is further subjected to dialysis treatment and freeze-drying treatment.
  • the dialysis treatment is: using deionized water or 0.1M sodium chloride solution to dialysis the hyaluronic acid derivative M for 3-7 days, and the dialysis bag used in the dialysis treatment has a molecular weight cut-off of 3500 Da.
  • the hyaluronic acid derivative F is obtained by modifying hyaluronic acid with 2-furan methylamine.
  • the modification method is as follows: using a mixture of 2-furan methylamine and a carboxyl activator to react with a hyaluronic acid buffer solution at 15-40° C.
  • the reaction temperature is 15°C, 16°C, 18°C, 20°C, 25°C, 30°C, 35°C or 40°C, etc.
  • the reaction time is 8h, 10h, 12h, 15h, 18h, 20h, 24h, 25h , 28h, 30h, 32h, 36h, 38h, 40h, 42h, 46h, 48h, 50h, 52h, 56h, 60h, 64h, 66h, 68h, 70h or 72h, etc.
  • the hyaluronic acid buffer solution is prepared by adding hyaluronic acid to 80-120 mM 2-(N-morpholine)ethanesulfonic acid buffer solution, and the mass concentration of the hyaluronic acid buffer solution is 0.001 ⁇ 0.01g/mL, e.g. 0.001g/mL, 0.002g/mL, 0.003g/mL, 0.004g/mL, 0.005g/mL, 0.006g/mL, 0.007g/mL, 0.008g/mL, 0.009g/ mL or 0.01g/mL, etc.
  • the 2-(N-morpholine)ethanesulfonic acid buffer solution is prepared by dissolving 2-(N-morpholine)ethanesulfonic acid in water and adjusting the pH to 5.3-5.8, for example, the pH is 5.3, 5.4, 5.5, 5.6, 5.7 or 5.8 etc.
  • the molar ratio of the 2-furanmethylamine to the repeating units of hyaluronic acid in the hyaluronic acid buffer solution is (1-6):1, such as 1:1, 1.5:1, 2:1 , 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5:1 or 6:1, etc.
  • the molar ratio of the carboxyl activator to the repeating units of hyaluronic acid in the hyaluronic acid buffer solution is (1-10):1, such as 1:1, 1.5:1, 2:1, 2.5 :1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5:1, 6:1, 7:1, 8:1, 9:1 or 10:1 etc.
  • the carboxyl activator is 4-(4,6-dimethoxytriazin-2-yl)-4-methylmorpholine hydrochloride.
  • the hyaluronic acid derivative F is further subjected to dialysis treatment and freeze-drying treatment.
  • the dialysis treatment is: using deionized water or 0.1M sodium chloride solution to dialysis the hyaluronic acid derivative F for 3-7 days, and the dialysis bag used in the dialysis treatment has a molecular weight cut-off of 3500 Da.
  • the temperature of the freeze-drying is -50 to -20°C, such as -50°C, -45°C, -40°C, -35°C, -30°C, -25°C or -20°C, etc.
  • the air-drying temperature is 20-80°C, such as 20°C, 30°C, 40°C, 50°C, 60°C, 70°C or 80°C, and the like.
  • the fifth object of the present invention is to provide a hyaluronic acid film, which is prepared by the preparation method of the hyaluronic acid film described in the fourth object.
  • the sixth object of the present invention is the application of the hyaluronic acid film described in the fifth object, and the hyaluronic acid film is used for the preparation of biomedical materials or tissue engineering materials.
  • the beneficial effects of the present invention also include: the hyaluronic acid film of the present invention adopts the cycloaddition reaction of the hyaluronic acid derivative M and the hyaluronic acid derivative F to avoid adding other macromolecular chains.
  • hyaluronic acid a ring structure is introduced into hyaluronic acid, photo-crosslinking reaction occurs after adding a photoinitiator, and after freeze-drying/air-drying, the obtained hyaluronic acid film has good tensile mechanical properties.
  • the obtained hyaluronic acid film freeze-dried film has uniform, loose and porous microscopic pores, the maximum tensile stress is 4.5MPa, the Young's modulus is 39.4MPa; the surface is smooth and flat, without obvious microscopic pores, and the maximum tensile stress is 145MPa , Young's modulus is 3.8GPa, which effectively expands the application range of hyaluronic acid film.
  • Fig. 1 is the actual picture of the hyaluronic acid hydrogel prepared by the embodiment 1a of the present invention
  • Fig. 2 is the hyaluronic acid hydrogel mechanical property test result graph of verification example 1 of the present invention
  • Fig. 3 is the hyaluronic acid hydrogel anti-fatigue test result diagram of verification example 2 of the present invention.
  • Fig. 4 is the hyaluronic acid hydrogel mechanical property test result graph of comparative example 5a of the present invention.
  • Fig. 5 is the SEM image of the hyaluronic acid film freeze-dried film prepared in Example 1b of the present invention.
  • Fig. 6 is the SEM image of the hyaluronic acid film air-dried film prepared in Example 2b of the present invention.
  • Example 7 is a graph showing the results of testing the mechanical properties of the hyaluronic acid film freeze-dried film prepared in Example 1b of the present invention.
  • Example 8 is a graph showing the results of testing the mechanical properties of the air-dried hyaluronic acid film prepared in Example 2b of the present invention.
  • Fig. 9 is the Young's modulus test result diagram of the single cross-linked freeze-dried film prepared by comparative example 5b of the present invention.
  • Fig. 10 is a graph showing the results of Young's modulus test of the single cross-linked air-dried film prepared in Comparative Example 5b of the present invention.
  • Hyaluronic acid aqueous solution
  • a hyaluronic acid aqueous solution with a mass concentration of 0.005 g/mL was prepared by mixing hyaluronic acid and deionized water.
  • the 2-methacrylic anhydride solution prepared above was mixed with the hyaluronic acid aqueous solution, and then the pH value was adjusted to 10 with NaOH solution, and the reaction was carried out at 4 °C for 20 hours. After the reaction, the reaction system was successively dialyzed and freeze-dried. , the hyaluronic acid derivative M is obtained, and the structural formula of the hyaluronic acid derivative M is as follows:
  • the dialysis was dialysis with deionized water for 5 days, and the molecular weight cut-off of the dialysis bag used for the dialysis was 3500 Da.
  • Hyaluronic acid was added to 100mM 2-(N-morpholine)ethanesulfonic acid buffer solution to prepare hyaluronic acid buffer solution, and the mass concentration of hyaluronic acid buffer solution was 0.005g/mL.
  • the dialysis was dialysis with deionized water for 5 days, and the molecular weight cut-off of the dialysis bag used for the dialysis was 3500 Da.
  • the hyaluronic acid derivative M prepared in step (1), the hyaluronic acid derivative F prepared in step (2), and phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) ) is mixed according to the volume ratio of 9:9:2; wherein the concentration of the hyaluronic acid derivative M is 0.03g/mL, the concentration of the hyaluronic acid derivative F is 0.03g/mL, and the concentration of the photoinitiator is 0.005g /mL;
  • the preparation method of the hyaluronic acid hydrogel in this embodiment is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, and the difference is as follows:
  • Hyaluronic acid derivative M, hyaluronic acid derivative F and phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) were mixed in a volume ratio of 9:9:2 ;
  • the concentration of the hyaluronic acid derivative M is 0.02g/mL
  • the concentration of the hyaluronic acid derivative F is 0.02/mL
  • the concentration of the photoinitiator is 0.03g/mL.
  • the preparation method of the hyaluronic acid hydrogel in this embodiment is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, and the difference is as follows:
  • Hyaluronic acid derivative M, hyaluronic acid derivative F and phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) were mixed in a volume ratio of 5:5:2 ;
  • the concentration of the hyaluronic acid derivative M is 0.05g/mL
  • the concentration of the hyaluronic acid derivative F is 0.05g/mL
  • the concentration of the photoinitiator is 0.008g/mL.
  • Sample 1 adopts the preparation method provided in Example 3a
  • Sample 2 adopts the preparation method provided in Example 1a
  • Sample 3 adopts the preparation method provided in Example 2a.
  • Test after preparation use a 2kN sensor, place it on the sensor compression substrate, set the test parameters according to the size of the sample, and stop the compression when the test curve changes abruptly.
  • the test parameters are: diameter of 10mm, height of 5mm, and compression rate of 3mm/min.
  • the test results are shown in Figure 2.
  • the maximum compressive stress of sample 1 is 1085kPa
  • the Young's modulus is about 25kPa
  • the maximum compressive stress of sample 2 is 530kPa
  • the Young's modulus is about 11kPa
  • the maximum compressive stress of sample 3 is 158kPa
  • the Young's modulus is about 3kPa .
  • the prepared samples were subjected to 10,000-cycle compression tests with an instron mechanical testing machine, and the set frequency was about 0.93 Hz, and the test curve as shown in Figure 3 was obtained. It can be seen from Figure 3 that the gel can withstand large deformation, And after 10,000 times of compression under 60% deformation, there is no breakage and fragmentation. After 1000 times of cyclic compression, the modulus and maximum compressive stress of the gel do not change greatly, and the maximum compressive stress remains the same. It can reach 75.2% of the original, while the modulus can reach 82.6% of the original, and the deformation recovery rate after 10,000 cycles of compression is also above 71%, which proves that the hydrogel has good anti-fatigue performance and can be under a certain external force. Withstand multiple squeezing without breaking.
  • the preparation method of the hyaluronic acid hydrogel of this comparative example is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, the difference is as follows:
  • the preparation method of the hyaluronic acid hydrogel of this comparative example is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, the difference is as follows:
  • the preparation method of the hyaluronic acid hydrogel of this comparative example is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, the difference is as follows:
  • the preparation method of the hyaluronic acid hydrogel of this comparative example is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, the difference is as follows:
  • the second cross-linking of the hyaluronic acid hydrogel is omitted, that is, the step of performing the light reaction is omitted, and only one cross-linking is performed without forming a gel.
  • the preparation method of the hyaluronic acid hydrogel of this comparative example is roughly the same as the preparation method provided in Example 1a, the difference lies in the preparation method of the hyaluronic acid hydrogel, the difference is as follows:
  • the two hydrogel samples were subjected to a compression test with an instron mechanical testing machine.
  • the test method and parameters were the same as those in Verification Example 1.
  • the results are shown in Figure 4.
  • the maximum compressive stress of sample 4 is 81 kPa, and the Young's modulus is about 4 kPa, the maximum compressive stress of sample 5 is 158 kPa, and the Young's modulus is about 2 kPa.
  • the present invention introduces a ring structure into hyaluronic acid, and after adding a photoinitiator, a photocrosslinking reaction occurs when the light is illuminated, so that the crosslinked hydrogel has better elasticity and mechanics. Its performance and fatigue resistance can withstand more than 10,000 cyclic compressions under large deformation without breakage and fragmentation, effectively expanding the application range of hyaluronic acid hydrogels.
  • Hyaluronic acid aqueous solution a hyaluronic acid aqueous solution with a mass concentration of 0.005 g/mL is prepared by mixing hyaluronic acid and deionized water.
  • 2-Methacrylic anhydride solution Dissolve each 500 ⁇ L of 2-methacrylic anhydride in 2mL of acetone solution.
  • Hyaluronic acid derivative M Mix the 2-methacrylic anhydride solution prepared above with the hyaluronic acid aqueous solution, then adjust the pH value to 10 with NaOH solution, and react at 4°C for 20 hours. After the reaction, the reaction The system is successively dialyzed and freeze-dried to obtain hyaluronic acid derivative M, and the structural formula of hyaluronic acid derivative M is as follows:
  • the dialysis was dialysis with deionized water for 5 days, and the molecular weight cut-off of the dialysis bag used was 3500 Da.
  • Hyaluronic acid buffer solution dissolve 2-(N-morpholine)ethanesulfonic acid particles in deionized water, adjust the pH to 5.5 with NaOH and/or hydrochloric acid, and prepare 2-(N-morpholine) Ethanesulfonic acid buffer solution, after preparation, refrigerate at 4°C for 24 hours before use; add hyaluronic acid to 100mM 2-(N-morpholine)ethanesulfonic acid buffer solution to prepare hyaluronic acid buffer solution, hyaluronic acid buffer solution The mass concentration of the solution was 0.005 g/mL.
  • the dialysis was dialysis with deionized water for 5 days, and the molecular weight cut-off of the dialysis bag used was 3500 Da.
  • the hyaluronic acid derivative M prepared in step (1), the hyaluronic acid derivative F and the phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) in step (2) ) is mixed according to the volume ratio of 9:9:2; wherein the concentration of the hyaluronic acid derivative M is 0.03g/mL, the concentration of the hyaluronic acid derivative F is 0.03g/mL, and the concentration of the photoinitiator is 0.005g /mL;
  • the preparation method of the hyaluronic acid film in this example is roughly the same as the preparation method provided in Example 1b, the difference lies in the preparation method of the hyaluronic acid film, which is an air-drying film. 10 hours.
  • the hyaluronic acid film was prepared, and its SEM image was shown in FIG. 6 . It can be seen from FIG. 6 that the surface of the prepared hyaluronic acid film is smooth and flat, without obvious microscopic pores.
  • the preparation method of the hyaluronic acid film in this example is roughly the same as the preparation method provided in Example 1b, the difference lies in the preparation method of the hyaluronic acid film, the difference is: the hyaluronic acid derivative M, the hyaluronic acid Derivative F and phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) are mixed according to the volume ratio of 9:9:2; wherein the concentration of hyaluronic acid derivative M is 0.02 g/mL, the concentration of the hyaluronic acid derivative F was 0.02/mL, and the concentration of the photoinitiator was 0.03 g/mL.
  • the preparation method of the hyaluronic acid film in this example is roughly the same as the preparation method provided in Example 1b, the difference lies in the preparation method of the hyaluronic acid film, the difference is: the hyaluronic acid derivative M, the hyaluronic acid Derivative F and phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt (photoinitiator) are mixed according to the volume ratio of 5:5:2; wherein the concentration of hyaluronic acid derivative M is 0.05 g/mL, the concentration of the hyaluronic acid derivative F was 0.05 g/mL, and the concentration of the photoinitiator was 0.008 g/mL.
  • the freeze-dried film and air-dried film samples were subjected to tensile testing with an instron mechanical testing machine.
  • the test conditions were as follows: after the samples were prepared according to the standard tensile testing method, the samples were tested using a 10N sensor, and the tensile samples were fixed on the clamps at the upper and lower ends. , to keep it in a normal stretching state, set the test parameters according to the size of the sample, and stop stretching when the test curve changes abruptly. Test parameters: length 15mm, width 5mm, height 1mm, tensile rate 0.5mm/min. The results show that the mechanical properties of the hyaluronic acid film are enhanced as the material concentration increases, reaching the highest at the concentrations used in Examples 1b and 2b, and then the mechanical strength decreases with increasing concentrations.
  • Figure 7 shows the test results of the mechanical properties of the hyaluronic acid freeze-dried film prepared in Example 1b. It can be seen that the obtained hyaluronic acid freeze-dried film has a maximum tensile stress of 4.5 MPa and a Young's modulus of 39.4 MPa.
  • Figure 8 shows the test results of the mechanical properties of the hyaluronic acid film air-dried film prepared in Example 2b. It can be seen that the maximum tensile stress of the hyaluronic acid air-dried film is 145MPa, and the Young's modulus is 3.8GPa.
  • the preparation method of the hyaluronic acid film of this comparative example is roughly the same as the preparation method provided in Example 1b, except that only one hyaluronic acid derivative M is used in this comparative example, no light is applied, and no gel is formed.
  • the preparation method of the hyaluronic acid film of this comparative example is roughly the same as the preparation method provided in Example 1b, except that only one hyaluronic acid derivative F is used in this comparative example, no light is applied, and no gel is formed.
  • the preparation method of the hyaluronic acid film of this comparative example is roughly the same as the preparation method provided in Example 1b, the difference is that only one hyaluronic acid derivative F is used in this comparative example, and no gel is formed after illumination.
  • the preparation method of the hyaluronic acid film in this comparative example is roughly the same as the preparation method provided in Example 1b, except that the second cross-linking of the hyaluronic acid film is omitted in this comparative example, that is, the light reaction is omitted. In one step, only one cross-linking is performed, and no gel is formed.
  • the preparation method of the hyaluronic acid film of this comparative example is roughly the same as the preparation method provided in Example 1b, except that only one hyaluronic acid derivative M is used in the preparation of the hyaluronic acid freeze-dried film and the air-dried film sample, light into glue;
  • the tensile test of the two film samples was carried out with an instron mechanical testing machine.
  • the test method and parameters were the same as those in Example 1b.
  • the test results of the single cross-linked freeze-dried film were shown in Figure 9, and its Young's modulus was 2.5MPa. ;
  • the test results of the single cross-linked air-dried film are shown in Figure 10, and its Young's modulus is 129MPa.
  • the present invention illustrates the detailed process equipment and process flow of the present invention through the above-mentioned embodiments, but the present invention is not limited to the above-mentioned detailed process equipment and process flow, that is, it does not mean that the present invention must rely on the above-mentioned detailed process equipment and process flow to be implemented. .
  • the preferred embodiments of the present invention are described above in detail, however, the present invention is not limited to the specific details of the above embodiments, and various simple modifications can be made to the technical solutions of the present invention within the scope of the technical concept of the present invention.
  • the specific technical features described in the above-mentioned specific embodiments can be combined in any suitable manner under the condition of no contradiction. In order to avoid unnecessary repetition, the present invention has The combination method will not be specified otherwise.

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Abstract

本发明公开了透明质酸水凝胶和透明质酸膜及制备方法和应用。本发明采用透明质酸衍生物M与透明质酸衍生物F发生环加成反应，在避免加入其他大分子链的情况下，在透明质酸中引入环结构，加入光引发剂后光照发生交联反应，由此制得的透明质酸水凝胶和透明质酸膜具有较好的弹性、力学性能和抗疲劳性，可承受万次循环压缩而不损坏，有效扩展了透明质酸水凝胶和透明质酸膜的应用范围。

Description

透明质酸水凝胶和透明质酸膜及其制备方法和应用 技术领域
本发明属于透明质酸领域，具体涉及一种透明质酸水凝胶及其制备方法和应用，以及一种透明质酸膜及其制备方法和应用。
背景技术
透明质酸(Hyaluronic acid，简称HA)是一种天然透明的多醣体，是由葡萄醛酸-N-乙酸氨基葡萄糖为双糖分子单位组成的直链高分子多醣，具有良好的生物相容性、生物可降解性、生物活性以及流变学特性，并且含有自由羧基和羟基，可在温和条件下进行改性，一直以来备受青睐。
水凝胶(Hydrogel)是一类极为亲水的三维网络结构凝胶，它在水中迅速溶胀并在此溶胀状态可以保持大量体积的水而不溶解。但是，现有的透明质酸水凝胶是由透明质酸大分子无规交联形成的本体胶，存在力学性能低、不能对生物活性分子进行控制释放等缺点，或采用有毒的交联剂，使透明质酸水凝胶的应用受到了很大的限制。
CN102942699A公开了一种自增强双交联透明质酸水凝胶及其制备方法。制备方法分为以下步骤：双键活化的透明质酸制备、透明质酸微球制备、双键活化的透明质酸微球的制备、自增强双交联透明质酸水凝胶的制备。通过该方法制备得到的自增强双交联透明质酸水凝胶由双肩活化的透明质酸微球作为增强颗粒与双键活化的透明质酸分子反应制得，具有双交联网络结构。其中双键活化的透明质酸微球的直径大小为1μm～10μm，双交联透明质酸水凝胶的孔径大小为10μm～70μm；双键取代度为2.8％～65％。与一次性交联透明质酸水凝胶相比，其胶弹性能良好，而且较好地延长对牛血清白蛋白的持续控制释放时间。
CN103724455A公开了一种透明质酸衍生物及其制备方法，所述方法将透明质酸或其盐与酸酐在无水溶剂中混合反应，产物经沉淀、分离、纯化后获得具有可交联性的透明质酸衍生物。该方法制备过程简单且反应程度可通过控制反应参数实现，获得的产物性能稳定，可长期保存。所得的具有可交联性的透明质酸衍生物可在温和条件下制备透明质酸水凝胶，具有良好的生物相容性，可用于生物活性物质载体材料、组织工程支架材料和软组织填充材料。
CN108341976A公开了一种基于无铜催化点击化学的甲基丙烯化基质材料的衍生物及其合成方法和应用。该发明提供了一种基于点击化学的甲基丙烯化基质材料的衍生物，将炔基或叠氮基团分别引入到甲基丙烯酰化的基质材料上 得到的两种衍生物。这两种材料不仅能够分别在光引发的自由基聚合作用下共价交联形成水凝胶，还可以将这两种材料的水溶液按一定比例简单共混形成水凝胶。并且，与自愈合水凝胶类似，利用光引发自由基聚合的方法分别将这两种材料制备成水凝胶，还可以通过水凝胶表面的炔基和叠氮基团发生点击化学反应，使这两种水凝胶能够在表面粘合。
CN111253591A公开了一种双交联透明质酸水凝胶、其制备方法与应用。所述制备方法包括：以改性剂对透明质酸进行改性处理，获得双键化透明质酸；在交联剂和光引发剂存在的条件下，使所述双键化透明质酸进行物理交联和光化学交联反应，从而获得双交联透明质酸水凝胶。该发明中的双交联透明质酸水凝胶具有良好的力学性能，且整个方案中透明质酸衍生物自身交联，有效避免了其他大分子链的引入，解决了透明质酸水凝胶因强度过低需与其他大分子链材料复合导致其在实际应用中受限的难题；并且，该发明使用水溶性更好、细胞毒性更小的光引发剂，成胶速度快，且双交联使所获水凝胶的力学性能得到了大大提升，最大压缩应力可达2.5MPa以上。
上述制得的透明质酸水凝胶的力学性能及抗疲劳性能有待进一步加强。
生物材料膜剂型具有宏观尺寸、稳定的三维网络结构和良好的力学强度，适用于美容、创伤敷料、给药制剂、各种补片、防粘连膜、角膜以及组织工程支架、人造器官等方面应用。但是，现有的透明质酸膜存在力学性能低的缺点，或采用有毒的交联剂，使透明质酸膜的应用受到了很大的限制。
CN105113054A公开了一种透明质酸衍生物交联纤维的制备方法，通过开环反应对透明质酸进行接枝改性，得到具有光聚合反应活性的透明质酸衍生物，然后加入光引发剂混溶后低温冷冻结晶，并在低温下进行光照交联聚合，再通过冷冻干燥除去溶剂，得到透明质酸衍生物交联纤维材料。通过该方法得到良好力学性能和机械强度的透明质酸衍生物交联纤维材料，制备过程中没有添加任何改变透明质酸特性的试剂，不会改变透明质酸作为天然高分子原有的一些优良性能，交联的透明质酸衍生物纤维材料可广泛应用于药物释放体系、伤口愈合材料、污水处理、重金属回收、膜分离、日用化工等领域。但是该方法只进行一次光交联，制得的透明质酸衍生物交联纤维的力学性能有待进一步提高。
因此，开发一种安全无毒害、力学性能优异的透明质酸膜很有必要。
发明内容
本发明的目的在于提供一种透明质酸水凝胶及其制备方法和应用，本发明的透明质酸水凝胶，具有较好的弹性、力学性能和抗疲劳性，能够在较大的形 变下承受一万次以上的循环压缩而不破损、碎裂，有效扩展了透明质酸水凝胶的应用范围。
本发明的目的之一在于提供一种透明质酸水凝胶的制备方法，为达此目的，本发明采用以下技术方案：
一种透明质酸水凝胶的制备方法，包括如下步骤：
1)在透明质酸衍生物M和透明质酸衍生物F的混合体系中，发生环加成反应，得到透明质酸水凝胶结合物；
2)在步骤1)得到的所述透明质酸水凝胶结合物中加入光引发剂，再经光照发生光交联反应，得到所述透明质酸水凝胶。
本发明的透明质酸水凝胶，采用透明质酸衍生物M与透明质酸衍生物F发生环加成反应，在避免加入其他大分子链的情况下，在透明质酸中引入环结构，加入光引发剂后光照发生光交联反应，使交联后的水凝胶具有较好的弹性、力学性能和抗疲劳性，能够在较大的形变下承受一万次以上的循环压缩而不破损、碎裂，有效扩展了透明质酸水凝胶的应用范围。其中，透明质酸衍生物M和透明质酸衍生物F避光发生环加成反应为第一次交联，加入光引发剂光照后透明质酸水凝胶进行了二次交联，制得的透明质酸水凝胶的弹性、力学性能和抗疲劳性进一步得到提升。
需要说明的是，抗疲劳特性是利用透明质酸衍生物F上的共轭双烯结构因电子离域作为给电子体一定条件下可与透明质酸衍生物M上的烯基发生狄尔斯-阿尔德反应，形成六元杂环，杂环上新生成的不饱和双键在光引发剂和特定波长光照条件下可发生自由基聚合反应，从而得到双交联水凝胶。本发明中，双交联透明质酸水凝胶在连接位点处形成的环结构，在水凝胶受到外力刺激下，可通过单键的旋转吸收部分能量从而起到内部能量耗散的作用，并由于环的空间位阻作用，使水凝胶内部的机械运动收到一定阻碍，增强了水凝胶对外力刺激的抗性，使其具有良好的抗疲劳性能，能够在较大形变下承受多次的压缩而不碎裂。
步骤1)中，所述透明质酸水凝胶结合物，包括结构式如式1所示的改性透明质酸；
式1：
Figure PCTCN2021121184-appb-000001
其中，式1中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4和R 6均独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任意一种；R 5、R 7和R 9均独立地选自氧、硫、取代或未取代亚烷基中的任意一种；R 8为透明质酸高分子去除羧基的残基部分。
步骤1)中，所述透明质酸衍生物M的结构式如式2所示，所述透明质酸衍生物F的结构式如式3所示。
其中，式2为
Figure PCTCN2021121184-appb-000002
式3为
Figure PCTCN2021121184-appb-000003
优选地，式2中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种。优选地，式3中，R 5为透明质酸高分子去除羧基的残基部分；R 6、R 10和R 11独立地选自氧、硫、取代或未取代亚烷基中的任一种；R 7、R 8、R 9、R 12、R 13、R 14、R 15和R 16独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种。
步骤1)中，所述透明质酸衍生物M与所述透明质酸衍生物F的体积比为(0.5～1.5):(0.5～1.5)；例如所述透明质酸衍生物M与所述透明质酸衍生物F的体积比为1:0.5、1:0.7、1:0.9、1:1.1、1:1.3或1:1.5、0.5:1、0.7:1、0.9:1、1.1:1、 1.3:1或1.5:1等。
优选地，所述透明质酸衍生物M与所述透明质酸衍生物F在反应体系中的终质量浓度均为0.01～0.05g/mL；例如透明质酸衍生物M与透明质酸衍生物F在反应体系中的终浓度均可以为0.01g/mL、0.02g/mL、0.03g/mL、0.04g/mL或0.05g/mL。
优选地，所述环加成反应的时间为6～24h，例如环加成反应的时间为6h、7h、8h、9h、10h、11h、12h、13h、14h、15h、16h、17h、18h、19h、20h、21h、22h、23h或24h等；所述环加成反应的温度为20～70℃，例如环加成反应的温度为20℃、30℃、40℃、50℃、60℃或70℃等。
需要说明的是，光引发剂的作用在于将制备得到的透明质酸水凝胶结合物制备为透明质酸水凝胶时发挥作用。在进行透明质酸水凝胶之前，对加入了光引发剂的透明质酸水凝胶结合物进行避光处理。需要说明的是，光引发剂的加入可以在制备透明质酸水凝胶结合物之前，也可以在制备透明质酸水凝胶结合物后，无论何时加入，均属于本发明的保护范围。
步骤2)中，所述透明质酸衍生物M、所述透明质酸衍生物F与所述光引发剂的体积比为(8.5～9.5):(8.5～9.5):(1.5～2.5)。
优选地，所述光引发剂的质量浓度为0.01～0.10g/mL，例如光引发剂的质量浓度为0.01g/mL、0.02g/mL、0.03g/mL、0.04g/mL、0.05g/mL、0.06g/mL、0.07g/mL、0.08g/mL、0.09g/mL或0.1g/mL等。
优选地，所述光引发剂为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮中的任意一种或至少两种的混合物。所述混合物典型但非限制的组合为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮的混合物，苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、1-羟基环己基苯基酮的混合物，2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮的混合物，苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮的混合物。
优选地，所述光照光交联反应所用的光源波长为360nm～450nm，例如光源波长为360nm、370nm、380nm、390nm、400nm、410nm、420nm、430nm、440nm或450nm等。
优选地，所述光照光交联反应的光照时间为30s～30min，例如光照的时间为30s、40s、50s、60s、1min、5min、10min、15min、20min、25min或30min等；光照的距离为1～30cm，例如光照的距离为1cm、2cm、3cm、4cm、5cm、6cm、 7cm、8cm、9cm、10cm、15cm、20cm、25cm或30cm等。
其中，步骤1)中，所述透明质酸衍生物M是采用2-甲基丙烯酸酐对透明质酸进行改性得到的。
优选地，所述改性的方法为：采用2-甲基丙烯酸酐溶液与透明质酸水溶液在0～8℃的条件下反应8～48h，得到所述透明质酸衍生物M；例如反应温度为0℃、1℃、2℃、3℃、4℃、5℃、6℃、7℃或8℃等，反应时间为8h、10h、12h、15h、16h、18h、20h、24h、26h、28h、30h、32h、35h、36h、38h、40h、42h、46h或48h等。
优选地，所述2-甲基丙烯酸酐溶液是2-甲基丙烯酸酐溶解在丙酮中配成的溶液；所述2-甲基丙烯酸酐与所述丙酮的体积比为1:(1～5)，例如所述2-甲基丙烯酸酐与所述丙酮的体积比为1:1、1:2、1:3、1:4或1:5等。
优选地，所述透明质酸水溶液由透明质酸与水混合配成；所述透明质酸水溶液的质量浓度为0.001～0.010g/mL，例如透明质酸水溶液的质量浓度为0.001g/mL、0.002g/mL、0.003g/mL、0.004g/mL、0.005g/mL、0.006g/mL、0.007g/mL、0.008g/mL、0.009g/mL或0.01g/mL等。
优选地，所述2-甲基丙烯酸酐溶液中的2-甲基丙烯酸酐与所述透明质酸水溶液中的透明质酸的重复单元的摩尔数比为(0.54～5.42):1，例如2-甲基丙烯酸酐与透明质酸的重复单元的摩尔数比为0.54:1、0.55:1、0.6:1、0.8:1、0.9:1、1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.1:1、5.2:1、5.3:1、5.4:1或5.42等。
优选地，所述透明质酸衍生物M还经过透析处理和冷冻干燥处理。
优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物M透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
其中，步骤1)中，所述透明质酸衍生物F是采用2-呋喃甲胺对透明质酸进行改性得到的。
优选地，所述改性的方法为：采用2-呋喃甲胺和羧基活化剂的混合物与透明质酸缓冲溶液在15～40℃的条件下反应8～72h，得到所述透明质酸衍生物F；例如反应的温度为15℃、16℃、18℃、20℃、25℃、30℃、35℃或40℃等，反应时间为8h、10h、12h、15h、18h、20h、24h、25h、28h、30h、32h、36h、38h、40h、42h、46h、48h、50h、52h、56h、60h、64h、66h、68h、70h或72h等。
优选地，所述透明质酸缓冲溶液是将透明质酸加入80～120mM的2-(N-吗啉)乙磺酸缓冲溶液中制得的，所述透明质酸缓冲溶液的质量浓度为 0.001～0.01g/mL，例如所述透明质酸缓冲溶液的质量浓度为0.001g/mL、0.002g/mL、0.003g/mL、0.004g/mL、0.005g/mL、0.006g/mL、0.007g/mL、0.008g/mL、0.009g/mL或0.01g/mL等。
优选地，所述2-(N-吗啉)乙磺酸缓冲溶液是将2-(N-吗啉)乙磺酸溶于水中，pH值调至5.3～5.8制得的，例如pH值为5.3、5.4、5.5、5.6、5.7或5.8等。
优选地，所述2-呋喃甲胺与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～6):1，例如所述2-呋喃甲胺与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.5:1或6:1等。
优选地，所述羧基活化剂与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～10):1，例如所述羧基活化剂与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.5:1、6:1、7:1、8:1、9:1或10:1等。
优选地，所述羧基活化剂为4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐。
优选地，所述透明质酸衍生物F还经过透析处理和冷冻干燥处理。
优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物F透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
本发明的目的之二在于提供一种透明质酸水凝胶，由目的之一所述的透明质酸水凝胶的制备方法制备得到。
优选地，所述透明质酸水凝胶包括重复单元的结构式如式4所示的改性透明质酸。
式4：
Figure PCTCN2021121184-appb-000004
其中，式4中，R 5、R 14、R 18和R 31为透明质酸高分子去除C 6上羟基的残基部分。
优选地，R 1、R 12、R 21和R 28为透明质酸高分子去除羧基的残基部分。
优选地，R 2、R 3、R 8、R 11、R 13、R 20、R 23、R 26、R 27、R 29、R 32、R 33、R 34和R 35独立地选自氧、硫、取代或未取代亚烷基中的任一种。
优选地，R 4、R 6、R 7、R 9、R 10、R 15、R 16、R 17、R 19、R 22、R 24、R 25和R 30独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基、取代或未取代氨基衍生物和磺酸基中的任一种。
本发明的目的之三在于目的之二所述的透明质酸水凝胶的应用，将所述透明质酸水凝胶用于生物医疗材料或组织工程材料的制备。
与现有技术相比，本发明的有益效果包括：
本发明的透明质酸水凝胶，采用透明质酸衍生物M与透明质酸衍生物F发生环加成反应，在避免加入其他大分子链的情况下，在透明质酸中引入环结构，加入光引发剂后光照发生光交联反应，使交联后的水凝胶具有较好的弹性、力学性能和抗疲劳性，具体的，制得的透明质酸水凝胶的压缩应力为100kPa～1.1MPa，弹性模量为1～30kPa，在较大形变下，能承受循环压缩一万次而不破损、碎裂，有效扩展了透明质酸水凝胶的应用范围。
本发明的目的还在于提供一种透明质酸膜及其制备方法和应用，本发明的透明质酸膜，具有较好的力学性能，制得的冻干膜具有适宜的微观孔隙，最大拉伸应力为4.5MPa，杨氏模量为39.4MPa；制得的风干膜最大拉伸应力为145MPa，杨氏模量为3.8GPa，有效扩展了透明质酸膜的应用范围。
本发明的目的之四在于提供一种透明质酸膜的制备方法，为达此目的，本发明采用以下技术方案：
一种透明质酸膜的制备方法，包括如下步骤：
1)在透明质酸衍生物M和透明质酸衍生物F的混合体系中，发生环加成反应，得到透明质酸前处理结合物；2)在步骤1)得到的所述透明质酸前处理结合物中加入光引发剂，搅拌均匀并去除气泡后，在模具中流延成膜，经光照发生光交联反应，再经冻干或风干，得到所述透明质酸膜。
本发明的透明质酸膜，采用透明质酸衍生物M与透明质酸衍生物F发生环加成反应，在避免加入其他大分子链的情况下，在透明质酸中引入环结构，加入光引发剂后光照发生光交联反应，再经冻干/风干后，获得的透明质酸膜具有较好的拉伸力学性能并具有不同的微观孔隙，有效扩展透明质酸膜的应用范围。
需要说明的是，拉伸性能是利用透明质酸衍生物F上的共轭双烯结构因电子离域作为给电子体一定条件下可与透明质酸衍生物M上的烯基发生狄尔斯-阿尔德反应，形成六元杂环，杂环上新生成的不饱和双键在光引发剂和特定波长光照条件下可发生自由基聚合反应，从而得到双交联透明质酸材料。本发明中，双交联透明质酸材料在连接位点处形成的环结构，在受到外力刺激下，可通过单键的旋转吸收部分能量从而起到内部能量耗散的作用，并由于环的空间位阻作用，增强了膜材料对外力刺激的抗性，使其具有良好的拉伸性能。
步骤1)中所述透明质酸前处理结合物包括结构式如式1所示改性透明质酸；
式1：
Figure PCTCN2021121184-appb-000005
其中，式1中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4和R 6均独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任意一种；R 5、R 7和R 9均独立地选自氧、硫、取代或未取代亚烷基中的任意一种；R 8为透明质酸高分子去除羧基的残基部分。
步骤1)中，所述透明质酸衍生物M的结构式如式2所示，所述透明质酸衍生物F的结构式如式3所示。
其中，式2为
Figure PCTCN2021121184-appb-000006
式3为
Figure PCTCN2021121184-appb-000007
优选地，式2中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、 取代或未取代氨基和取代或未取代磺酸基中的任一种。
优选地，式3中，R 5为透明质酸高分子去除羧基的残基部分；R 6、R 10和R 11独立地选自氧、硫、取代或未取代亚烷基中的任一种；R 7、R 8、R 9、R 12、R 13、R 14、R 15和R 16独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种。
步骤1)中透明质酸衍生物M与透明质酸衍生物F的体积比为(0.5～1.5):(0.5～1.5)；例如1:0.5、1:0.7、1:0.9、1:1.1、1:1.3或1:1.5、0.5:1、0.7:1、0.9:1、1.1:1、1.3:1或1.5:1等。
优选地，所述透明质酸衍生物M与所述透明质酸衍生物F在反应体系中的终质量浓度均为0.01～0.05g/mL；例如可以为0.01g/mL、0.02g/mL、0.03g/mL、0.04g/mL或0.05g/mL。优选地，所述环加成反应的时间为6～24h，例如为6h、7h、8h、9h、10h、11h、12h、13h、14h、15h、16h、17h、18h、19h、20h、21h、22h、23h或24h等；所述环加成反应的温度为20～70℃，例如20℃、30℃、40℃、50℃、60℃或70℃等。
需要说明的是，光引发剂的作用在于将制备得到的透明质酸前处理结合物制备为透明质酸膜时发挥作用。在进行透明质酸膜之前，对加入了光引发剂的透明质酸前处理结合物可进行避光处理。需要说明的是，光引发剂的加入可以在制备透明质酸前处理结合物之前，也可以在制备透明质酸前处理结合物后，无论何时加入，均属于本发明的保护范围。
步骤2)中，透明质酸衍生物M、透明质酸衍生物F与光引发剂的体积比为(8.5～9.5):(8.5～9.5):(1.5～2.5)。
优选地，所述光引发剂的质量浓度为0.01～0.10g/mL，例如0.01g/mL、0.02g/mL、0.03g/mL、0.04g/mL、0.05g/mL、0.06g/mL、0.07g/mL、0.08g/mL、0.09g/mL或0.1g/mL等。优选地，所述光引发剂为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮中的任意一种或至少两种的混合物。所述混合物典型但非限制的组合为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮的混合物，苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、1-羟基环己基苯基酮的混合物，2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮的混合物，苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮的混合物。优选地，所述光照光交联反应所用的光源波长为360nm～450nm，例如360nm、370nm、380nm、390nm、400nm、410nm、420nm、430nm、440nm 或450nm等。优选地，所述光照光交联反应的光照时间为30s～30min，例如30s、40s、50s、60s、1min、5min、10min、15min、20min、25min或30min等；光照的距离为1～30cm，例如1cm、2cm、3cm、4cm、5cm、6cm、7cm、8cm、9cm、10cm、15cm、20cm、25cm或30cm等。
其中，步骤1)中，所述透明质酸衍生物M是采用2-甲基丙烯酸酐对透明质酸进行改性得到的。
优选地，所述改性的方法为：采用2-甲基丙烯酸酐溶液与透明质酸水溶液在0～8℃的条件下反应8～48h，得到所述透明质酸衍生物M；例如反应温度为0℃、1℃、2℃、3℃、4℃、5℃、6℃、7℃或8℃等，反应时间为8h、10h、12h、15h、16h、18h、20h、24h、26h、28h、30h、32h、35h、36h、38h、40h、42h、46h或48h等。优选地，所述2-甲基丙烯酸酐溶液是2-甲基丙烯酸酐溶解在丙酮中配成的溶液；所述2-甲基丙烯酸酐与所述丙酮的体积比为1:(1～5)，例如所述2-甲基丙烯酸酐与所述丙酮的体积比为1:1、1:2、1:3、1:4或1:5等。优选地，所述透明质酸水溶液由透明质酸与水混合配成；所述透明质酸水溶液的质量浓度为0.001～0.010g/mL，例如0.001g/mL、0.002g/mL、0.003g/mL、0.004g/mL、0.005g/mL、0.006g/mL、0.007g/mL、0.008g/mL、0.009g/mL或0.01g/mL等。优选地，所述2-甲基丙烯酸酐溶液中的2-甲基丙烯酸酐与所述透明质酸水溶液中的透明质酸的重复单元的摩尔数比为(0.54～5.42):1，例如0.54:1、0.55:1、0.6:1、0.8:1、0.9:1、1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.1:1、5.2:1、5.3:1、5.4:1或5.42等。
优选地，所述透明质酸衍生物M还经过透析处理和冷冻干燥处理。优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物M透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
其中，步骤1)中，所述透明质酸衍生物F是采用2-呋喃甲胺对透明质酸进行改性得到的。优选地，所述改性的方法为：采用2-呋喃甲胺和羧基活化剂的混合物与透明质酸缓冲溶液在15～40℃的条件下反应8～72h，得到所述透明质酸衍生物F；例如反应的温度为15℃、16℃、18℃、20℃、25℃、30℃、35℃或40℃等，反应时间为8h、10h、12h、15h、18h、20h、24h、25h、28h、30h、32h、36h、38h、40h、42h、46h、48h、50h、52h、56h、60h、64h、66h、68h、70h或72h等。优选地，所述透明质酸缓冲溶液是将透明质酸加入80～120mM的2-(N-吗啉)乙磺酸缓冲溶液中制得的，所述透明质酸缓冲溶液的质量浓度为0.001～0.01g/mL，例如0.001g/mL、0.002g/mL、0.003g/mL、0.004g/mL、0.005g/mL、 0.006g/mL、0.007g/mL、0.008g/mL、0.009g/mL或0.01g/mL等。优选地，所述2-(N-吗啉)乙磺酸缓冲溶液是将2-(N-吗啉)乙磺酸溶于水中，pH值调至5.3～5.8制得的，例如pH值为5.3、5.4、5.5、5.6、5.7或5.8等。优选地，所述2-呋喃甲胺与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～6):1，例如1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.5:1或6:1等。优选地，所述羧基活化剂与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～10):1，例如1:1、1.5:1、2:1、2.5:1、3:1、3.5:1、4:1、4.5:1、5:1、5.5:1、6:1、7:1、8:1、9:1或10:1等。
优选地，所述羧基活化剂为4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐。
优选地，所述透明质酸衍生物F还经过透析处理和冷冻干燥处理。
优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物F透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
步骤2)中，所述冻干的温度为-50～-20℃，例如-50℃、-45℃、-40℃、-35℃、-30℃、-25℃或-20℃等。优选地，所述风干的温度为20～80℃，例如20℃、30℃、40℃、50℃、60℃、70℃或80℃等。
本发明的目的之五在于提供一种透明质酸膜，由目的之四所述的透明质酸膜的制备方法制备得到。
本发明的目的之六在于目的之五所述的透明质酸膜的应用，将所述透明质酸膜用于生物医疗材料或组织工程材料的制备。
与现有技术相比，本发明的有益效果还包括：本发明的透明质酸膜，采用透明质酸衍生物M与透明质酸衍生物F发生环加成反应，在避免加入其他大分子链的情况下，在透明质酸中引入环结构，加入光引发剂后光照发生光交联反应，再经冻干/风干后，获得的透明质酸膜具有较好的拉伸力学性能，具体的，制得的透明质酸膜冻干膜具有均匀的、疏松多孔的微观孔隙，最大拉伸应力4.5MPa，杨氏模量39.4MPa；表面光滑平整，无明显的微观孔隙，最大拉伸应力145MPa，杨氏模量3.8GPa，有效扩展了透明质酸膜的应用范围。
附图说明
图1为本发明的实施例1a制备的透明质酸水凝胶实物图；
图2为本发明的验证例1的透明质酸水凝胶力学性能测试结果图；
图3为本发明的验证例2的透明质酸水凝胶抗疲劳测试结果图；
图4为本发明的对比例5a的透明质酸水凝胶力学性能测试结果图；
图5为本发明实施例1b制备的透明质酸膜冻干膜的SEM图；
图6为本发明实施例2b制备的透明质酸膜风干膜的SEM图；
图7为本发明的实施例1b制备的透明质酸膜冻干膜力学性能测试结果图；
图8为本发明实施例2b制备的透明质酸膜风干膜的力学性能测试结果图；
图9为本发明对比例5b制备的单交联冻干膜的杨氏模量测试结果图；
图10为本发明对比例5b制备的单交联风干膜的杨氏模量测试结果图。
具体实施方式
下面结合附图，并通过具体实施方式来进一步说明本发明的技术方案。
如无具体说明，本发明的实施例中未注明具体条件者，按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者，均为可以通过市售购买获得的常规产品。
实施例1a
本实施例的透明质酸水凝胶的制备方法，其包括以下步骤：
(1)透明质酸衍生物M的制备：
透明质酸水溶液：
由透明质酸与去离子水混合配成质量浓度为0.005g/mL的透明质酸水溶液。
2-甲基丙烯酸酐溶液：
每500μL2-甲基丙烯酸酐溶于2mL的丙酮溶液中配成。
透明质酸衍生物M：
将上述配制得到的2-甲基丙烯酸酐溶液与透明质酸水溶液混合，然后采用NaOH溶液调整pH值至10，在4℃的条件下反应20h，反应结束后，反应体系依次经透析和冷冻干燥，得到透明质酸衍生物M，透明质酸衍生物M的结构式如下：
Figure PCTCN2021121184-appb-000008
其中，透析为采用去离子水透析5天，透析所用的透析袋的截留分子量为3500Da。
(2)透明质酸衍生物F的制备：
透明质酸缓冲溶液：
将2-(N-吗啉)乙磺酸颗粒溶于去离子水中，再用NaOH和/或盐酸调pH值至5.5，定容配制成2-(N-吗啉)乙磺酸缓冲溶液，配制后冷藏于4℃的条件下24h后使用；
将透明质酸加入100mM2-(N-吗啉)乙磺酸缓冲溶液中制得透明质酸缓冲溶液，透明质酸缓冲溶液的质量浓度为0.005g/mL。
透明质酸衍生物F：
在磁力搅拌的条件下，在透明质酸缓冲溶液中，加入用量为透明质酸重复单元数5倍的4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐，搅拌20min后，加入用量为透明质酸重复单元数2倍的2-呋喃甲胺，在20℃的条件下反应40h，反应结束后，依次经透析和冷冻干燥，得到透明质酸衍生物F，透明质酸衍生物F的结构式如下：
Figure PCTCN2021121184-appb-000009
其中，透析为采用去离子水透析5天，透析所用的透析袋的截留分子量为3500Da。
(3)透明质酸水凝胶的制备：
将步骤(1)制备的透明质酸衍生物M、步骤(2)制备的透明质酸衍生物F以及苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为9:9:2进行混合；其中透明质酸衍生物M的浓度为0.03g/mL，透明质酸衍生物F的浓度为0.03g/mL，光引发剂的浓度为0.005g/mL；
混合后，避光，除去气泡，然后置于37℃的水浴锅中，避光过夜反应，得到一步交联后的水凝胶结合物，再经光照，光照波长为360nm，光照时间为8min，光照距离为5cm，得到双交联的透明质酸水凝胶，如图1所示。
实施例2a
本实施例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
将透明质酸衍生物M、透明质酸衍生物F以及苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为9:9:2进行混合；其中透明质酸衍生物M的 浓度为0.02g/mL，透明质酸衍生物F的浓度为0.02/mL，光引发剂的浓度为0.03g/mL。
实施例3a
本实施例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
将透明质酸衍生物M、透明质酸衍生物F以及苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为5:5:2进行混合；其中透明质酸衍生物M的浓度为0.05g/mL，透明质酸衍生物F的浓度为0.05g/mL，光引发剂的浓度为0.008g/mL。
验证例1
通过用instron力学测试机对三种水凝胶样品进行压缩测试。
样品1采用实施例3a提供的制备方法，样品2采用实施例1a提供的制备方法，样品3采用实施例2a提供的制备方法。
制备好后进行测试，使用2kN传感器，置于传感器压缩基底上，根据样品的大小设置好测试参数，测试曲线出现突变后，停止压缩。
测试参数为：直径为10mm，高5mm，压缩速率3mm/min。测试结果如图2所示，随着材料浓度的提高，水凝胶的力学性能也随之增强。样品1的最大压缩应力为1085kPa，杨氏模量约为25kPa，样品2的最大压缩应力为530kPa，杨氏模量约为11kPa，样品3的最大压缩应力为158kPa，杨氏模量约为3kPa。
验证例2
通过用instron力学测试机对制备的样品进行万次循环压缩测试，设置的频率约为0.93Hz，得到如图3的测试曲线，从图3中可以看到，凝胶能够承受较大的形变，并且在60％形变下承受一万次的压缩之后不出现破损和碎裂的情况，在循环压缩一千次之后，凝胶的模量和最大压缩应力没有发生较大的改变，最大压缩应力仍然可以达到原来的75.2％，而模量则可以达到原来的82.6％，万次循环压缩之后的形变恢复率也在71％以上，证明水凝胶具有良好的抗疲劳性能，能在一定外力作用下承受多次的挤压而不至破碎。
对比例1a
本对比例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
本对比例中，仅使用一种透明质酸衍生物M，不光照，未成胶。
对比例2a
本对比例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
本对比例中，仅使用一种透明质酸衍生物F，不光照，未成胶。
对比例3a
本对比例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
本对比例中，仅使用一种透明质酸衍生物F，光照后，未成胶。
对比例4a
本对比例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
本对比例中，省略了将透明质酸水凝胶的第二次交联，即省略了进行光照反应的一步，只进行了一次交联，未成胶。
对比例5a
本对比例的透明质酸水凝胶的制备方法，其与实施例1a提供的制备方法大致相同，区别在于透明质酸水凝胶的制备方法的不同，区别如下：
水凝胶样品4制备时，仅使用一种透明质酸衍生物M，光照成胶；
水凝胶样品5制备时，两种透明质酸衍生物混合并除气泡后不经第一步交联，立刻光照成胶；
通过用instron力学测试机对两种水凝胶样品进行压缩测试，测试方法及参数与验证例1中相同，结果如图4所示，样品4的最大压缩应力为81kPa，杨氏模量约为4kPa，样品5的最大压缩应力为158kPa，杨氏模量约为2kPa。
由验证例1a及对比例5a中结果可知，本发明在透明质酸中引入环结构，加入光引发剂后光照发生光交联反应，使交联后的水凝胶具有较好的弹性、力学性能和抗疲劳性，能够在较大的形变下承受一万次以上的循环压缩而不破损、碎裂，有效扩展了透明质酸水凝胶的应用范围。
实施例1b
本实施例的透明质酸膜的制备方法，其包括以下步骤：
(1)透明质酸衍生物M的制备：
透明质酸水溶液：由透明质酸与去离子水混合配成质量浓度为0.005g/mL的透明质酸水溶液。
2-甲基丙烯酸酐溶液：每500μL2-甲基丙烯酸酐溶于2mL的丙酮溶液中配成。
透明质酸衍生物M：将上述配制得到的2-甲基丙烯酸酐溶液与透明质酸水溶液混合，然后采用NaOH溶液调整pH值至10，在4℃的条件下反应20h，反应结束后，反应体系依次经透析和冷冻干燥，得到透明质酸衍生物M，透明质酸衍生物M的结构式如下：
Figure PCTCN2021121184-appb-000010
其中，透析为采用去离子水透析5天，所用透析袋的截留分子量为3500Da。
(2)透明质酸衍生物F的制备：
透明质酸缓冲溶液：将2-(N-吗啉)乙磺酸颗粒溶于去离子水中，再用NaOH和/或盐酸调pH值至5.5，定容配制成2-(N-吗啉)乙磺酸缓冲溶液，配制后冷藏于4℃的条件下24h后使用；将透明质酸加入100mM2-(N-吗啉)乙磺酸缓冲溶液中制得透明质酸缓冲溶液，透明质酸缓冲溶液的质量浓度为0.005g/mL。
透明质酸衍生物F：
在磁力搅拌的条件下，在透明质酸缓冲溶液中，加入用量为透明质酸重复单元数5倍的4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐，搅拌20min后，加入用量为透明质酸重复单元数2倍的2-呋喃甲胺，在20℃的条件下反应40h，反应结束后，依次经透析和冷冻干燥，得到透明质酸衍生物F，透明质酸衍生物F的结构式如下：
Figure PCTCN2021121184-appb-000011
其中，透析为采用去离子水透析5天，所用透析袋的截留分子量为3500Da。
(3)透明质酸膜的制备：
将步骤(1)制备的透明质酸衍生物M、透明质酸衍生物F以及步骤(2)中的苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为9:9:2进行 混合；其中透明质酸衍生物M的浓度为0.03g/mL，透明质酸衍生物F的浓度为0.03g/mL，光引发剂的浓度为0.005g/mL；
混合后，避光，除去气泡，然后置于37℃的水浴锅中，避光过夜反应，得到一步交联后的水凝胶结合物，在模具中流延成膜，再经光照，光照波长为360nm，光照时间为8min，光照距离为5cm，再经-20℃冻干，冻干时间为16小时，制备得到透明质酸膜，其SEM图如图5所示。由图5可看出，制备得到的透明质酸膜具有均匀的、疏松多孔的微观形貌。
实施例2b
本实施例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别在于透明质酸膜的制备方法的不同，为风干膜，风干条件为：温度20℃，风干时间10小时。制备得到透明质酸膜，其SEM图如图6所示。由图6可看出，制备得到的透明质酸膜表面光滑平整，无明显的微观孔隙。
实施例3b
本实施例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别在于透明质酸膜的制备方法的不同，区别为：将透明质酸衍生物M、透明质酸衍生物F以及苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为9:9:2进行混合；其中透明质酸衍生物M的浓度为0.02g/mL，透明质酸衍生物F的浓度为0.02/mL，光引发剂的浓度为0.03g/mL。
实施例4b
本实施例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别在于透明质酸膜的制备方法的不同，区别为：将透明质酸衍生物M、透明质酸衍生物F以及苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐(光引发剂)按照体积比为5:5:2进行混合；其中透明质酸衍生物M的浓度为0.05g/mL，透明质酸衍生物F的浓度为0.05g/mL，光引发剂的浓度为0.008g/mL。
用instron力学测试机对冻干膜和风干膜样品进行拉伸测试，测试条件如下：按标准拉伸测试方法制备好样品后进行测试，使用10N传感器，将拉伸样品固定在上下两端的夹具上，使其保持正常伸展状态，根据样品的大小设置好测试参数，测试曲线出现突变后停止拉伸。测试参数：长15mm，宽5mm，高1mm，拉伸速率0.5mm/min。结果显示，随着材料浓度提高，透明质酸膜的力学性能增强，在实施例1b和2b所用浓度时达到最高，随后力学强度随浓度增加而降低。
实施例1b制备的透明质酸冻干膜力学性能测试结果图如图7所示，可以看出，制得的透明质酸冻干膜最大拉伸应力为4.5MPa，杨氏模量为39.4MPa。实 施例2b制备的透明质酸膜风干膜的力学性能测试结果图如图8所示，可以看出，透明质酸风干膜最大拉伸应力为145MPa，杨氏模量为3.8GPa。
对比例1b
本对比例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别为：本对比例中仅使用一种透明质酸衍生物M，不光照，未成胶。
对比例2b
本对比例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别为：本对比例中仅使用一种透明质酸衍生物F，不光照，未成胶。
对比例3b
本对比例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别为：本对比例中仅使用一种透明质酸衍生物F，光照后，未成胶。
对比例4b
本对比例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别为：本对比例中省略了将透明质酸膜的第二次交联，即省略光照反应的一步，只进行一次交联，未成胶。
对比例5b
本对比例的透明质酸膜的制备方法，其与实施例1b提供的制备方法大致相同，区别为：透明质酸冻干膜和风干膜样品制备时仅使用一种透明质酸衍生物M，光照成胶；
通过用instron力学测试机对两种膜样品进行拉伸测试，测试方法及参数与实施例1b中相同，单交联冻干膜的测试结果如图9所示，其杨氏模量为2.5MPa；单交联风干膜的测试结果如图10所示，其杨氏模量为129MPa。
本发明通过上述实施例来说明本发明的详细工艺设备和工艺流程，但本发明并不局限于上述详细工艺设备和工艺流程，即不意味着本发明必须依赖上述详细工艺设备和工艺流程才能实施。以上详细描述了本发明的优选实施方式，但是，本发明并不限于上述实施方式中的具体细节，在本发明的技术构思范围内，可以对本发明的技术方案进行多种简单变型。另外需要说明的是，在上述具体实施方式中所描述的各个具体技术特征，在不矛盾的情况下，可以通过任何合适的方式进行组合，为了避免不必要的重复，本发明对各种可能的组合方式不再另行说明。

Claims (20)

  1. 一种透明质酸水凝胶的制备方法，其特征在于，所述制备方法包括如下步骤：
    1)在透明质酸衍生物M和透明质酸衍生物F的混合体系中，发生环加成反应，得到透明质酸水凝胶结合物；
    2)在步骤1)得到的所述透明质酸水凝胶结合物中加入光引发剂，再经光照发生光交联反应，得到所述透明质酸水凝胶。
  2. 根据权利要求1所述的制备方法，其特征在于，步骤1)中，所述透明质酸结合物包括结构式如式1所示的改性透明质酸；
    式1：
    Figure PCTCN2021121184-appb-100001
    其中，式1中，R 1为透明质酸高分子去除C 6上羟基的残基部分；
    R 2、R 3、R 4和R 6均独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任意一种；
    R 5、R 7和R 9均独立地选自氧、硫、取代或未取代亚烷基中的任意一种；
    R 8为透明质酸高分子去除羧基的残基部分；
    优选地，步骤1)中，所述透明质酸衍生物M的结构式如式2所示，所述透明质酸衍生物F的结构式如式3所示；
    式2为
    Figure PCTCN2021121184-appb-100002
    式3为
    Figure PCTCN2021121184-appb-100003
    和/或
    Figure PCTCN2021121184-appb-100004
    优选地，式2中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代 羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种；
    优选地，式3中，R 5为透明质酸高分子去除羧基的残基部分；R 6、R 10和R 11独立地选自氧、硫、取代或未取代亚烷基中的任一种；R 7、R 8、R 9、R 12、R 13、R 14、R 15和R 16独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种。
  3. 根据权利要求1或2所述的制备方法，其特征在于，步骤1)中，所述透明质酸衍生物M与所述透明质酸衍生物F的体积比为(0.5～1.5):(0.5～1.5)；
    优选地，所述透明质酸衍生物M与所述透明质酸衍生物F在反应体系中的终质量浓度均为0.01～0.05g/mL；
    优选地，所述环加成反应的时间为6～24h，所述环加成反应的温度为20～70℃。
  4. 根据权利要求1-3之一所述的制备方法，其特征在于，步骤2)中，所述透明质酸衍生物M、所述透明质酸衍生物F与所述光引发剂的体积比为(8.5～9.5):(8.5～9.5):(1.5～2.5)；
    优选地，所述光引发剂的质量浓度为0.01～0.10g/mL；
    优选地，所述光引发剂为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮中的任意一种或至少两种的混合物。
  5. 根据权利要求1-4之一所述的制备方法，其特征在于，步骤2)中，所述光照的光源波长为360nm～450nm，所述光照的时间为30s～30min，所述光照的距离为1～30cm。
  6. 根据权利要求1-5之一所述的制备方法，其特征在于，步骤1)中，所述透明质酸衍生物M是采用2-甲基丙烯酸酐对透明质酸进行改性得到的；
    优选地，所述改性的方法为：采用2-甲基丙烯酸酐溶液与透明质酸水溶液在0～8℃的条件下反应8～48h，得到所述透明质酸衍生物M；
    优选地，所述2-甲基丙烯酸酐溶液是2-甲基丙烯酸酐溶解在丙酮中配成的溶液；所述2-甲基丙烯酸酐与所述丙酮的体积比为1:(1～5)；
    优选地，所述透明质酸水溶液由透明质酸与水混合配成；所述透明质酸水溶液的质量浓度为0.001～0.010g/mL；
    优选地，所述2-甲基丙烯酸酐溶液中的2-甲基丙烯酸酐与所述透明质酸水溶液中的透明质酸的重复单元的摩尔数比为(0.54～5.42):1；
    优选地，所述透明质酸衍生物M还经过透析处理和冷冻干燥处理；
    优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物M透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
  7. 根据权利要求1-6之一所述的制备方法，其特征在于，所述透明质酸衍生物F是采用2-呋喃甲胺对透明质酸进行改性得到的；
    优选地，所述改性的方法为：采用2-呋喃甲胺和羧基活化剂的混合物与透明质酸缓冲溶液在15～40℃的条件下反应8～72h，得到所述透明质酸衍生物F；
    优选地，所述透明质酸缓冲溶液是将透明质酸加入80～120mM的2-(N-吗啉)乙磺酸缓冲溶液中制得的，所述透明质酸缓冲溶液的质量浓度为0.001～0.01g/mL；
    优选地，所述2-(N-吗啉)乙磺酸缓冲溶液是将2-(N-吗啉)乙磺酸溶于水中，pH值调至5.0～6.0制得的；
    优选地，所述2-呋喃甲胺与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～6):1；
    优选地，所述羧基活化剂与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～10):1；
    优选地，所述羧基活化剂为4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐；
    优选地，所述透明质酸衍生物F还经过透析处理和冷冻干燥处理；
    优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物F透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
  8. 一种如权利要求1-7之一所述的制备方法得到的透明质酸水凝胶。
  9. 根据权利要求8所述的透明质酸水凝胶，其特征在于，所述透明质酸水凝胶包括重复单元的结构式如式4所示的改性透明质酸；
    式4：
    Figure PCTCN2021121184-appb-100005
    其中，式4中，R 5、R 14、R 18和R 31为透明质酸高分子去除C 6上羟基的残基部分；
    优选地，R 1、R 12、R 21和R 28为透明质酸高分子去除羧基的残基部分；
    优选地，R 2、R 3、R 8、R 11、R 13、R 20、R 23、R 26、R 27、R 29、R 32、R 33、R 34和R 35独立地选自氧、硫、取代或未取代亚烷基中的任一种；
    优选地，R 4、R 5、R 6、R 7、R 9、R 10、R 12、R 15、R 16、R 17、R 19、R 22、R 24、R 25和R 30独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基、取代或未取代氨基衍生物和磺酸基中的任一种。
  10. 一种如权利要求8或9所述的透明质酸水凝胶的应用，其特征在于，将所述透明质酸水凝胶用于生物医疗材料或组织工程材料的制备。
  11. 一种透明质酸膜的制备方法，其包括如下步骤：
    1)在透明质酸衍生物M和透明质酸衍生物F的混合体系中，发生环加成反应，得到透明质酸前处理结合物；
    2)在步骤1)得到的所述透明质酸前处理结合物中加入光引发剂，在模具中流延成膜，经光照发生光交联反应，再经冻干或风干，得到所述透明质酸膜。
  12. 根据权利要求11所述的制备方法，其特征在于，步骤1)中，所述透明质酸前处理结合物包括结构式如式1所示的改性透明质酸；
    式1：
    Figure PCTCN2021121184-appb-100006
    其中，式1中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4和R 6均独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任意一种；R 5、R 7和R 9均独立地选自氧、硫、取代或未取代亚烷基中的任意一种；R 8为透明质酸高分子去除羧基的残基部分；
    优选地，步骤1)中，所述透明质酸衍生物M的结构式如式2所示，所述透明质酸衍生物F的结构式如式3所示；
    式2为
    Figure PCTCN2021121184-appb-100007
    式3为
    Figure PCTCN2021121184-appb-100008
    和/或
    Figure PCTCN2021121184-appb-100009
    优选地，式2中，R 1为透明质酸高分子去除C 6上羟基的残基部分；R 2、R 3、R 4独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种；
    优选地，式3中，R 5为透明质酸高分子去除羧基的残基部分；R 6、R 10和R 11独立地选自氧、硫、取代或未取代亚烷基中的任一种；R 7、R 8、R 9、R 12、R 13、R 14、R 15和R 16独立地选自氢、取代或未取代烷基、取代或未取代羟基、取代或未取代羧基、取代或未取代氨基和取代或未取代磺酸基中的任一种。
  13. 根据权利要求11或12所述的制备方法，其特征在于，步骤1)中，所述透明质酸衍生物M与所述透明质酸衍生物F的体积比为(0.5～1.5):(0.5～1.5)；
    优选地，所述透明质酸衍生物M与所述透明质酸衍生物F在反应体系中的 终质量浓度均为0.01～0.05g/mL；
    优选地，所述环加成反应的时间为6～24h，所述环加成反应的温度为20～70℃。
  14. 根据权利要求11-13之一所述的制备方法，其特征在于，步骤2)中，所述透明质酸衍生物M、所述透明质酸衍生物F与所述光引发剂的体积比为(8.5～9.5):(8.5～9.5):(1.5～2.5)；
    优选地，所述光引发剂的质量浓度为0.01～0.10g/mL；
    优选地，所述光引发剂为苯基(2,4,6-三甲基苯甲酰基)磷酸锂盐、2-羟基-4’-(2-羟乙氧基)-2-甲基苯丙酮和1-羟基环己基苯基酮中的任意一种或至少两种的混合物。
  15. 根据权利要求11-14之一所述的制备方法，其特征在于，步骤2)中，所述光照的光源波长为360nm～450nm，所述光照的时间为30s～30min，所述光照的距离为1～30cm。
  16. 根据权利要求11-15之一所述的制备方法，其特征在于，步骤1)中，所述透明质酸衍生物M是采用2-甲基丙烯酸酐对透明质酸进行改性得到的；
    优选地，所述改性的方法为：采用2-甲基丙烯酸酐溶液与透明质酸水溶液在0～8℃的条件下反应8～48h，得到所述透明质酸衍生物M；
    优选地，所述2-甲基丙烯酸酐溶液是2-甲基丙烯酸酐溶解在丙酮中配成的溶液；所述2-甲基丙烯酸酐与所述丙酮的体积比为1:(1～5)；
    优选地，所述透明质酸水溶液由透明质酸与水混合配成；所述透明质酸水溶液的质量浓度为0.001～0.010g/mL；
    优选地，所述2-甲基丙烯酸酐溶液中的2-甲基丙烯酸酐与所述透明质酸水溶液中的透明质酸的重复单元的摩尔数比为(0.54～5.42):1；
    优选地，所述透明质酸衍生物M还经过透析处理和冷冻干燥处理；
    优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物M透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
  17. 根据权利要求11-16之一所述的制备方法，其特征在于，所述透明质酸衍生物F是采用2-呋喃甲胺对透明质酸进行改性得到的；
    优选地，所述改性的方法为：采用2-呋喃甲胺和羧基活化剂的混合物与透明质酸缓冲溶液在15～40℃的条件下反应8～72h，得到所述透明质酸衍生物F；
    优选地，所述透明质酸缓冲溶液是将透明质酸加入80～120mM的2-(N-吗啉)乙磺酸缓冲溶液中制得的，所述透明质酸缓冲溶液的质量浓度为 0.001～0.01g/mL；
    优选地，所述2-(N-吗啉)乙磺酸缓冲溶液是将2-(N-吗啉)乙磺酸溶于水中，pH值调至5.0～6.0制得的；
    优选地，所述2-呋喃甲胺与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～6):1；
    优选地，所述羧基活化剂与所述透明质酸缓冲溶液中透明质酸的重复单元的摩尔数比为(1～10):1；
    优选地，所述羧基活化剂为4-(4,6-二甲氧基三嗪-2-基)-4-甲基吗啉盐酸盐；
    优选地，所述透明质酸衍生物F还经过透析处理和冷冻干燥处理；
    优选地，所述透析处理为：采用去离子水或0.1M氯化钠溶液对透明质酸衍生物F透析3～7天，所述透析处理所用的透析袋的截留分子量为3500Da。
  18. 根据权利要求11-17之一所述的制备方法，其特征在于，步骤2)中，所述冻干的温度为-50～-20℃；优选地，所述风干的温度为20-80℃。
  19. 一种如权利要求11-18之一所述的制备方法得到的透明质酸膜。
  20. 一种如权利要求19所述的透明质酸膜的应用，其特征在于，将所述透明质酸膜用于生物医疗材料或组织工程材料的制备。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116942890A (zh) * 2023-06-09 2023-10-27 西南交通大学 具有促伤口闭合的抗菌抗炎温敏性水凝胶及其制备方法

Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005023906A1 (ja) * 2003-09-08 2005-03-17 Chugai Seiyaku Kabushiki Kaisha ヒアルロン酸修飾物、及びそれを用いた薬物担体
US20130231474A1 (en) * 2010-11-18 2013-09-05 Universite Joseph Fourier - Grenoble 1 Polysaccharide derivatives including an alkene unit and thiol-click chemical coupling reaction
CN103724455A (zh) * 2013-12-11 2014-04-16 四川大学 一种透明质酸衍生物及其水凝胶的制备方法
US20150291706A1 (en) * 2012-11-27 2015-10-15 Contipro Biotech S.R.O. Photoreactive Derivative of Hyaluronic Acid, Method of Preparation Thereof, 3D.Crosslinked Derivative of Hyaluronic Acid, Method of Preparation and Use Thereof
US20160312385A1 (en) * 2013-11-21 2016-10-27 Contipro Biotech S R.O. Nanofibers Containing Photocurable Ester Derivative of Hyaluronic Acid or Its Salt, Photocured Nanofibers, Method of Synthesis Thereof, Preparation Containing Photocured Nanofibers and Use Thereof
CN107976472A (zh) * 2017-11-22 2018-05-01 江南大学 一种传感器电极表面抗生物污染涂层的制备方法
CN108341976A (zh) * 2017-01-25 2018-07-31 四川大学 基于点击化学的甲基丙烯化基质材料的衍生物及合成方法
CN112159534A (zh) * 2020-09-28 2021-01-01 吾奇生物医疗科技(镇江)有限公司 一种透明质酸水凝胶及其制备方法和应用
CN112156234A (zh) * 2020-09-28 2021-01-01 吾奇生物医疗科技(镇江)有限公司 一种透明质酸膜及其制备方法和应用
CN112192797A (zh) * 2020-09-28 2021-01-08 吾奇生物医疗科技(镇江)有限公司 一种制膜装置及制膜方法
CN213797685U (zh) * 2020-09-28 2021-07-27 吾奇生物医疗科技(江苏)有限公司 一种制膜装置

Patent Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005023906A1 (ja) * 2003-09-08 2005-03-17 Chugai Seiyaku Kabushiki Kaisha ヒアルロン酸修飾物、及びそれを用いた薬物担体
US20130231474A1 (en) * 2010-11-18 2013-09-05 Universite Joseph Fourier - Grenoble 1 Polysaccharide derivatives including an alkene unit and thiol-click chemical coupling reaction
US20150291706A1 (en) * 2012-11-27 2015-10-15 Contipro Biotech S.R.O. Photoreactive Derivative of Hyaluronic Acid, Method of Preparation Thereof, 3D.Crosslinked Derivative of Hyaluronic Acid, Method of Preparation and Use Thereof
US20160312385A1 (en) * 2013-11-21 2016-10-27 Contipro Biotech S R.O. Nanofibers Containing Photocurable Ester Derivative of Hyaluronic Acid or Its Salt, Photocured Nanofibers, Method of Synthesis Thereof, Preparation Containing Photocured Nanofibers and Use Thereof
CN103724455A (zh) * 2013-12-11 2014-04-16 四川大学 一种透明质酸衍生物及其水凝胶的制备方法
CN108341976A (zh) * 2017-01-25 2018-07-31 四川大学 基于点击化学的甲基丙烯化基质材料的衍生物及合成方法
CN107976472A (zh) * 2017-11-22 2018-05-01 江南大学 一种传感器电极表面抗生物污染涂层的制备方法
CN112159534A (zh) * 2020-09-28 2021-01-01 吾奇生物医疗科技(镇江)有限公司 一种透明质酸水凝胶及其制备方法和应用
CN112156234A (zh) * 2020-09-28 2021-01-01 吾奇生物医疗科技(镇江)有限公司 一种透明质酸膜及其制备方法和应用
CN112192797A (zh) * 2020-09-28 2021-01-08 吾奇生物医疗科技(镇江)有限公司 一种制膜装置及制膜方法
CN213797685U (zh) * 2020-09-28 2021-07-27 吾奇生物医疗科技(江苏)有限公司 一种制膜装置

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116942890A (zh) * 2023-06-09 2023-10-27 西南交通大学 具有促伤口闭合的抗菌抗炎温敏性水凝胶及其制备方法
CN116942890B (zh) * 2023-06-09 2024-04-02 西南交通大学 具有促伤口闭合的抗菌抗炎温敏性水凝胶及其制备方法

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