CN116870263B - Cornea bandage mirror with bionic layer structure and preparation method thereof - Google Patents

Cornea bandage mirror with bionic layer structure and preparation method thereof Download PDF

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Publication number
CN116870263B
CN116870263B CN202311151277.XA CN202311151277A CN116870263B CN 116870263 B CN116870263 B CN 116870263B CN 202311151277 A CN202311151277 A CN 202311151277A CN 116870263 B CN116870263 B CN 116870263B
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cornea
mirror
bandage mirror
cornea bandage
solution
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CN116870263A (en
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陈江
张泽真
曲超
罗越
张诗萌
肖文哲
王彬鉴
赵弋豪
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Sichuan Peoples Hospital of Sichuan Academy of Medical Sciences
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Sichuan Peoples Hospital of Sichuan Academy of Medical Sciences
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L31/00Materials 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
    • A61L31/08Materials for coatings
    • A61L31/10Macromolecular materials
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L31/00Materials 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
    • A61L31/08Materials for coatings
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L31/00Materials 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
    • A61L31/08Materials for coatings
    • A61L31/082Inorganic materials
    • A61L31/088Other specific inorganic materials not covered by A61L31/084 or A61L31/086
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L31/00Materials 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
    • A61L31/14Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L31/00Materials 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
    • A61L31/14Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L31/16Biologically active materials, e.g. therapeutic substances
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L2300/00Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/216Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials with other specific functional groups, e.g. aldehydes, ketones, phenols, quaternary phosphonium groups
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS 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
    • A61L2300/00Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/40Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
    • A61L2300/404Biocides, antimicrobial agents, antiseptic agents
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2400/00Materials characterised by their function or physical properties
    • A61L2400/18Modification of implant surfaces in order to improve biocompatibility, cell growth, fixation of biomolecules, e.g. plasma treatment

Abstract

The invention discloses a cornea bandage mirror with a bionic layer structure and a preparation method thereof, belonging to the technical field of cornea bandage mirrors, wherein the preparation method comprises the following steps: immersing cornea bandage mirror in water solution A containing polyphenol compound, water solution B containing silver ion, water solution C containing polyphenol compound and polyamine group compound, water solution D containing polyphenol compound and water solution E containing sulfhydryl hyaluronic acid in turn, and finally washing with deionized water to obtain the cornea bandage mirror; wherein, the cornea bandage mirror is soaked in the solution B and the solution E and simultaneously irradiated by ultraviolet light. The surface of the cornea bandage mirror prepared by the method forms a bionic layer structure, so that the problem that the existing cornea bandage mirror is easy to cause microbial keratitis can be solved, the using effect of the cornea bandage mirror is greatly improved, and the purpose of preventing and treating eye diseases is improved.

Description

Cornea bandage mirror with bionic layer structure and preparation method thereof
Technical Field
The invention relates to the technical field of cornea bandage mirrors, in particular to a cornea bandage mirror with a bionic layer structure and a preparation method thereof.
Background
The human eyes are easily affected by various external adverse factors. Such as an impaired ocular surface barrier, is susceptible to infection by pathogenic microorganisms. Keratitis is one of the most common ocular disorders, such as early treatment failure, which can further exacerbate the corneal ulceration and even perforation, ultimately leading to corneal blindness. 85% of corneal blindness is due to infectious keratopathy. Infectious keratitis is mainly caused by pathogenic microorganisms such as bacteria (e.g., staphylococcus aureus, pseudomonas aeruginosa) or fungi (e.g., aspergillus fumigatus, fusarium). The disease is urgent, the disease is serious, if the correct treatment is not carried out in time, serious complications such as corneal ulcer, perforation, eye content removal, endophthalmitis and the like can be caused, the infection can be controlled only by cornea transplantation, but the cornea donation is limited, and many patients can finally carry out eyeball excision only.
The cornea bandage mirror can be used as a bandage and a medicine warehouse to help treat intractable cornea defect and cornea wound after repair operation and provide required medicine, so that the medicine is released continuously, the residence time of the medicine on the ocular surface is greatly prolonged, the ocular bioavailability of the medicine is improved from 1% -5% of eye drops to 50%, and better curative effect and less side effect are generated. However, conventional keratobandaged lenses often cause various types of microbial keratitis during wear, and are still considered contraindicated in infectious keratitis. Therefore, how to surface modify traditional cornea bandages, develop cornea bandages with effective antibacterial performance, and establish an effective drug delivery system, which is helpful for wide use of cornea bandages and more effective prevention and control of eye infection.
Disclosure of Invention
In order to solve the defects in the prior art, the invention aims to provide a cornea bandage mirror with a bionic layer structure and a preparation method thereof, so as to solve the problem that microbial keratitis is easy to cause in the use process of the traditional cornea bandage mirror and improve the prevention effect and the treatment effect of the cornea bandage mirror.
The technical scheme for solving the technical problems is as follows:
a preparation method of a cornea bandage mirror with a bionic layer structure comprises the following steps:
immersing cornea bandage mirror in water solution A containing polyphenol compound, water solution B containing silver ion, water solution C containing polyphenol compound and polyamine group compound, water solution D containing polyphenol compound and cross-linking agent and water solution E containing sulfhydrylation hyaluronic acid in turn, and finally washing with deionized water to obtain the cornea bandage mirror;
wherein, the cornea bandage mirror is soaked in the solution B and the solution E and simultaneously irradiated by ultraviolet light.
Further, the polyphenol compound in the aqueous solution a includes at least one of dopamine, tea polyphenol, tannic acid, anthocyanin, catechol, gallic acid and epicatechin, and the mass concentration of the polyphenol compound in the aqueous solution a is 0.5-4%.
Further, the pH value of the aqueous solution A is 7.0-9.0, the soaking temperature of the cornea bandage mirror in the aqueous solution A is 20-40 ℃, and the soaking time is 1-8h.
Further, silver ions are provided by silver nitrate, the mass concentration of the silver ions in the aqueous solution B is 0.5-4%, the soaking temperature of the cornea bandage mirror in the aqueous solution B is 20-40 ℃, and the soaking time is 1-30min.
Further, the polyphenol compound in the aqueous solution C comprises at least one of dopamine, tea polyphenol, tannic acid, anthocyanin, catechol, gallic acid and epicatechin; the polyamine group compound includes at least one of hexamethylenediamine and polyethylene polyamine; the mass concentration of the polyphenol compound in the aqueous solution C is 0.5-2%, and the mass concentration of the polyamine group compound in the aqueous solution C is 0.5-5%.
Further, the soaking temperature of the cornea bandage mirror in the aqueous solution C is 25-40 ℃ and the soaking time is 12-48h.
Further, the solvent of the solution D is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, wherein the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 70-73%, the mass fraction of the N-hydroxysuccinimide in the solvent is 22-25%, and the mass fraction of the water in the solvent is 2-8%;
the mass concentration of the polyphenol compound in the solution D is 1-10%, and the polyphenol compound is at least one of gallic acid and baicalin.
Further, the soaking temperature of the cornea bandage mirror in the solution D is 25-40 ℃ and the soaking time is 0.5-2h.
Further, the mass concentration of the sulfhydryl hyaluronic acid in the aqueous solution E is 1-5%, the soaking temperature of the cornea bandage mirror in the aqueous solution E is 25-40 ℃, and the soaking time is 0.1-2h.
A cornea bandage mirror with a bionic layer structure is prepared by adopting the method.
The invention has the following beneficial effects:
when the cornea bandage mirror is soaked in the aqueous solution A, the polyphenol compound is fixed on the cornea bandage mirror through the action modes of hydrogen bond, van der Waals force, coordination bond, pi-pi accumulation and the like, so that an active site is provided for subsequent materials; when the film is soaked in the aqueous solution B, the phenolic hydroxyl groups on the polyphenol compound fixed on the surface of the cornea bandage mirror and silver ions are subjected to complexation, so that the purpose of fixing the silver ions on the surface of the polyphenol compound is realized; when the coating is soaked in the aqueous solution C, the polyphenol compound and the polyamine group compound form covalent bonds through Michael addition reaction of oxidized derivative quinone of catechol on the polyphenol compound, and more amino groups are introduced to the coating of the cornea bandage mirror; when the hydrophilic adhesive is soaked in the aqueous solution D, the polyphenol compound is grafted onto polyamine through covalent bonds formed by the reaction of amino groups and carboxyl amides, the fixation of the polyphenol compound is further realized, when the hydrophilic adhesive is soaked in the aqueous solution E, the thiol-modified hyaluronic acid is fixed on the polyphenol compound through Michael addition reaction of thiol groups and catechol derivative quinone on the polyphenol, and after the operation, a bionic ocular surface glycocalyx structure is formed on the surface of the cornea bandage mirror. The ocular surface glycocalyx is an epithelial cell epitaxial barrier formed by mucin, and plays a plurality of roles of resisting pollution and invasion of microorganisms, maintaining stable ocular surface tear layer, keeping lubrication, preserving antibacterial biomolecules, regulating and controlling ocular surface microenvironment steady state and the like. The occurrence of keratitis is often accompanied by the destruction of the eye surface glycocalyx, and the bionic eye surface glycocalyx structure formed on the surface of the cornea bandage mirror can play roles in promoting the moisture retention of eyes, promoting the repair of cornea epithelium, preventing infection and enhancing anti-pollution, thereby prolonging the wearing time of the cornea bandage mirror and realizing targeted regulation of the change of pathological microenvironment caused by the eye surface inflammation and oxidative stress.
Drawings
FIG. 1 is a schematic illustration of the preparation flow of a keratology procedure in example 1;
FIG. 2 is a graph showing the results of water contact angles before and after modification of a keratoscope;
FIG. 3 is a graph of silver ion release from a modified keratoscope;
FIG. 4 is a graph of oxidation resistance results of a modified keratology;
FIG. 5 is a fluorescence micrograph of Staphylococcus aureus and Pseudomonas aeruginosa adhesion on the surface of the corneal bandage mirror prior to modification;
FIG. 6 is a high-magnification fluorescence micrograph of modified corneal bandaging lens staphylococcus aureus and pseudomonas aeruginosa adhesion;
FIG. 7 is a fluorescence micrograph of macrophage adhesion on the surface of a corneal bandage mirror prior to modification;
FIG. 8 is a fluorescence micrograph of macrophage adhesion on the surface of the modified cornea bandages;
FIG. 9 is a fluorescence microscope image of macrophage adhesion after hyaluronidase action before and after corneal bandage mirror modification;
FIG. 10 is a fluorescence micrograph of epithelial cell adhesion on the surface of the corneal bandages before and after modification.
Detailed Description
The examples given below are only intended to illustrate the invention and are not intended to limit the scope thereof. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing the cornea bandage mirror into 2wt% dopamine aqueous solution with pH value of 8.5, and soaking for 4h at 30deg.C;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing dopamine and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then putting the cornea bandage mirror into a solution containing baicalin, soaking the cornea bandage mirror in the solution for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of baicalin in the solution is 2wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 2
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing cornea bandage mirror into tannic acid water solution with mass concentration of 2wt%, wherein pH value of water solution is 8.5, and soaking for 4h at 30deg.C;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing dopamine and polyethylene polyamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then placing the cornea bandage mirror into a solution containing gallic acid, and soaking for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of gallic acid in the solution is 4wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 4wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 3
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing the cornea bandage mirror into a dopamine aqueous solution with the mass concentration of 1wt%, wherein the pH value of the aqueous solution is 7, and soaking for 8 hours at the temperature of 30 ℃;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 5min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing tannic acid and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then putting the cornea bandage mirror into a solution containing baicalin, soaking the cornea bandage mirror in the solution for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of baicalin in the solution is 2wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 4
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing cornea bandage mirror into 2wt% tea polyphenol water solution with pH value of 8.5, soaking for 4 hr at 30deg.C;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing tea polyphenol and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of tea polyphenol in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then putting the cornea bandage mirror into a solution containing baicalin, soaking the cornea bandage mirror in the solution for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of baicalin in the solution is 2wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 5
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing the cornea bandage mirror into 2wt% dopamine aqueous solution with pH value of 7.8, and soaking for 4h at 30deg.C;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 365nm at the temperature of 30 ℃;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing dopamine and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 2wt% and the mass concentration of hexamethylenediamine is 5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then placing the cornea bandage mirror into a solution containing gallic acid, and soaking for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of gallic acid in the solution is 1wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 1wt%, soaking the cornea bandage mirror for 1h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 6
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing cornea bandage mirror into tannic acid water solution with mass concentration of 2wt%, wherein pH value of water solution is 7.8, and soaking for 4h at 30deg.C;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing catechol and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then putting the cornea bandage mirror into a solution containing baicalin, soaking the cornea bandage mirror in the solution for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of baicalin in the solution is 1wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 7
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing the cornea bandage mirror into catechol water solution with the mass concentration of 2wt%, wherein the pH value of the water solution is 8.5, and soaking for 4 hours at the temperature of 30 ℃;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 2wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing catechol and hexamethylenediamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 2wt% and the mass concentration of hexamethylenediamine is 5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), cleaning with deionized water, then placing the cornea bandage mirror into a solution containing gallic acid, and soaking the cornea bandage mirror in the solution for 0.5h at the temperature of 40 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of gallic acid in the solution is 2wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Example 8
The preparation method of the cornea bandage mirror with the bionic layer structure comprises the following steps:
(1) Placing the cornea bandage mirror into a 4wt% dopamine aqueous solution, wherein the pH value of the aqueous solution is 8.5, and soaking for 2 hours at 30 ℃;
(2) Taking out the cornea bandage mirror treated in the step (1), putting the cornea bandage mirror into a silver nitrate aqueous solution with the mass concentration of 4wt%, and soaking the cornea bandage mirror for 1min under the ultraviolet irradiation condition of 25 ℃ and 365 nm;
(3) Taking out the cornea bandage mirror treated in the step (2), cleaning with deionized water, then putting into an aqueous solution containing dopamine and polyethylene polyamine, and soaking for 24 hours at 25 ℃; the mass concentration of dopamine in the aqueous solution is 1wt%, and the mass concentration of hexamethylenediamine is 2.5wt%;
(4) Taking out the cornea bandage mirror treated in the step (3), washing with deionized water, then placing the cornea bandage mirror into a solution containing gallic acid, and soaking for 1h at the temperature of 30 ℃, wherein the solvent in the solution is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water, the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 71%, the mass fraction of the N-hydroxysuccinimide in the solvent is 24%, and the mass fraction of the water in the solvent is 5%; the mass concentration of gallic acid in the solution is 4wt%;
(5) Taking out the cornea bandage mirror treated in the step (4), cleaning with deionized water, then placing the cornea bandage mirror in a thiolated hyaluronic acid aqueous solution with the mass concentration of 2wt%, soaking the cornea bandage mirror for 0.5h at the temperature of 25 ℃ and then irradiating the cornea bandage mirror with ultraviolet light for 0.1h, and then cleaning the cornea bandage mirror with deionized water to obtain the cornea bandage mirror with the bionic layer structure.
Experimental example
FIG. 1 is a schematic view of the preparation process of a cornea bandage mirror with a bionic layer structure in example 1.
Taking the cornea bandage mirror before and after modification in the example 1 as an example, the performance change before and after modification is detected, and the specific detection process is as follows:
1. water contact angle test
The change in hydrophilicity of each sample before and after surface modification was examined using a droplet shape analysis system (DSA 100, kruss, germany), and the specific examination results are shown in fig. 2.
As can be seen from fig. 2, the water contact angle of the surface of the modified cornea bandage mirror is 33 °, and the water contact angle of the cornea bandage mirror before modification is 92 °, and the water contact angle of the cornea bandage mirror before modification is far greater than that of the cornea bandage mirror after modification, which proves that the cornea bandage mirror after modification has better hydrophilicity and wettability. The modified cornea bandage mirror can improve the comfort level of eyes in the use process, and is convenient for eyes to recover.
2. Silver ion release experiment
(1) Soaking the modified cornea bandage mirror in 4mL of PBS solution at 37 ℃, collecting the soaked PBS solution at 1, 3, 5, 7, 10, 15 and 30 days respectively, and adding 4mL of new PBS solution for continuous soaking;
(2) And (3) detecting (1) the silver particle content in the collected PBS solution by adopting an atomic absorption spectrum (ASS, japen-Hitachi-Z-2300), and calculating the total silver particle content in the coating. The specific results are shown in FIG. 3.
As can be seen from fig. 3, as the soaking time is prolonged, the content of silver ions in the PBS solution gradually increases, which indicates that the modified coating can release silver ions for a long time, and the silver ions can be sterilized in a broad spectrum, thereby realizing a long-term antibacterial effect.
3. Antioxidant experiment
(1) Taking an unmodified cornea bandage mirror as a control group, and placing the unmodified cornea bandage mirror and cornea bandage mirror samples at different modification stages in a hydrogen peroxide aqueous solution with the concentration of 0.01M;
(2) The contact angles of water on the surface of the cornea bandage mirror before soaking, at 6h, 12h and 24h are respectively tested. The hydrophilicity change is used to characterize the oxidation resistance of all samples before and after modification. The specific results are shown in FIG. 4.
As can be seen from fig. 4, the surface structure of the modified sample is destroyed by oxidizing the surface of the modified sample with hydrogen peroxide, and the hydrophilicity is reduced, so that the water contact angle is changed. Baicalin has a large amount of polyphenols, has a certain reducibility, and can resist oxidation. Before baicalin grafting modification, the water contact angle increases with the increase of the hydrogen peroxide soaking time. However, after baicalin is grafted, the change of the water contact angle is obviously reduced, which proves that the modified coating has good oxidation resistance.
4. Bacterial adhesion experiments
(1) The cornea bandage mirror before and after modification is placed at a concentration of 1X 10 6 Incubating CFU/ml in PBS solution of pseudomonas aeruginosa or staphylococcus aureus for 1h at 37 ℃;
(2) Removing the treated keratoscope of (1) from the PBS solution and rinsing 5 times with PBS solution;
(3) Treating the cornea bandage mirror treated in the step (2) with 2.5% glutaraldehyde solution at 4 ℃ for 24 hours;
(4) Taking out the cornea bandage mirror treated in the step (3), cleaning with normal saline, and then dyeing for 15min by using rhodamine normal saline solution;
(5) Taking out the cornea bandage mirror treated in the step (4), washing 3 times by using normal saline, and drying and using a fluorescence microscope to adhere bacteria on the surface of the cornea bandage mirror. The specific results are shown in FIGS. 5-6.
As can be seen from FIG. 5, after the corneal bandage mirror before modification is treated with Pseudomonas aeruginosa and Staphylococcus aureus, the surface of the corneal bandage mirror contains a large amount of Pseudomonas aeruginosa or Staphylococcus aureus.
As can be seen from fig. 6, after the modified cornea bandage mirror is treated by pseudomonas aeruginosa and staphylococcus aureus, the number of the pseudomonas aeruginosa and the staphylococcus aureus on the surface of the cornea bandage mirror is obviously reduced, which proves that silver ions added in the cornea bandage mirror in the modification process have stronger bactericidal effect, can effectively kill the pseudomonas aeruginosa or the staphylococcus aureus, and reduce the pollution to eyes of patients.
5. Inflammatory cell culture experiments
(1) Cutting the cornea bandage mirrors before and after modification into wafers with diameters of 8mm, and placing the wafers in a 48-hole plate;
(2) Adding 3X 10 to the well plate in which the sample was placed in (1) 4 Mouse macrophage cell/mL, and placing in a cell incubator for culturing for 1 and 3 days;
(3) Taking out the cornea bandage mirror after incubation in the step (2), lightly flushing for 3 times by using normal saline, sucking out, and then adding glutaraldehyde normal saline solution with the volume concentration of 2.5% for fixation for 24 hours;
(4) The cornea bandage mirror treated in the step (3) is taken out and stained with rhodamine physiological saline solution for 15min.
(5) Taking out the cornea bandage mirror treated in the step (4), washing 3 times by using normal saline, and drying and using a fluorescence microscope to adhere macrophages on the cornea bandage mirror surface. The specific results are shown in FIGS. 7-8.
As can be seen from fig. 7, the surface of the cornea bandage mirror before modification contains a large number of macrophages, which proves that the surface of the cornea bandage mirror generates inflammation;
as can be seen from fig. 8, the macrophages on the surface of the modified cornea bandage mirror are obviously reduced, and the modified cornea bandage mirror has a certain anti-inflammatory effect.
6. Anti-hyaluronidase assay
Omitting the operation in step (4) on the basis of the production method in example 1 to produce a keratome, and conducting an experiment using the keratome and the keratome produced in example 1 as samples; the method comprises the following steps:
(1) 1mL of hyaluronidase PBS solution with the concentration of 100mg/L is added to the surface of the sample, the cornea bandage mirror sample is collected after incubation for 3 h at 37 ℃, and the cornea bandage mirror sample is rinsed by deionized water;
(2) Cutting the cornea bandage mirror collected in the step (1) into wafers with diameters of 8mm, and placing the wafers in a 48-well plate;
(3) Adding 3X 10 to the well plate in which the sample was placed in (2) 4 Mouse macrophages at each mL and placed in a cell incubator for 1 day;
(4) Taking out the cornea bandage mirror after incubation in the step (3), lightly flushing for 3 times by using normal saline, sucking out, and then adding glutaraldehyde normal saline solution with the volume concentration of 2.5% for fixation for 24 hours;
(5) The cornea bandage mirror treated in the step (4) is taken out and stained with rhodamine physiological saline solution for 15min.
(6) Taking out the cornea bandage mirror treated in the step (5), washing the cornea bandage mirror with normal saline for 3 times, and detecting adhesion of macrophages on the surface of the cornea bandage mirror by using a fluorescence microscope after the cornea bandage mirror is dried. The specific results are shown in FIG. 9.
As can be seen from fig. 9, the anti-inflammatory properties of the hyaluronic acid layer without baicalin grafted thereto were significantly reduced after hyaluronidase action, because part of the hyaluronic acid was decomposed and the insulation effect was reduced. The difference between the front and the back of the hyaluronic acid layer grafted with the baicalin is not great, because the baicalin has the function of inhibiting the enzyme activity, and the grafting of the baicalin can reduce the decomposition of the hyaluronic acid by the hyaluronidase.
7. Corneal epithelial cell culture experiments
(1) Add 3X 10 to 24 well plate 4 individual/mL human corneal epithelial cells and placed in a cell incubator for 1 day;
(2) Placing the cornea bandage mirrors before and after modification in a 24-hole plate in which the human cornea epithelial cells are cultured in the step (1), and placing the cornea bandage mirrors in a cell incubator for culturing for 2 days;
(3) Taking out the cornea bandage mirror after incubation in the step (2), lightly flushing a 24-pore plate with physiological saline for 3 times and sucking out, and then adding glutaraldehyde physiological saline solution with the volume concentration of 2.5% into the 24-pore plate for fixation for 24 hours;
(4) Staining the corneal epithelial cells in the well plate with rhodamine physiological saline solution for 15min;
(5) The 24-well plate treated in the above step (4) was washed 3 times with physiological saline, and after drying, adhesion of corneal epithelial cells inside the well was observed by using a fluorescence microscope. The specific results are shown in FIG. 10.
As can be seen from fig. 10, the number of corneal epithelial cells on the surface of the modified sample is increased, which indicates that the constructed bionic glycocalyx structure has good biocompatibility and can effectively promote cornea repair.
The foregoing description of the preferred embodiments of the invention is not intended to limit the invention to the precise form disclosed, and any such modifications, equivalents, and alternatives falling within the spirit and scope of the invention are intended to be included within the scope of the invention.

Claims (10)

1. The preparation method of the cornea bandage mirror with the bionic layer structure is characterized by comprising the following steps of:
immersing cornea bandage mirror in water solution A containing polyphenol compound, water solution B containing silver ion, water solution C containing polyphenol compound and polyamine group compound, water solution D containing polyphenol compound and water solution E containing sulfhydryl hyaluronic acid in turn, and finally washing with deionized water to obtain the cornea bandage mirror;
the polyphenol compound in the aqueous solution A and the aqueous solution C comprises at least one of dopamine, tea polyphenol, tannic acid, anthocyanin, catechol, gallic acid and epicatechin; the polyphenol compound in the solution D is at least one of gallic acid and baicalin;
wherein, the cornea bandage mirror is soaked in the solution B and the solution E and simultaneously irradiated by ultraviolet light.
2. The method for producing a cornea bandage mirror having a bionic layer structure according to claim 1, wherein the mass concentration of the polyphenol compound in the aqueous solution a is 0.5 to 4%.
3. The method for preparing a cornea bandage mirror with a bionic layer structure according to claim 2, wherein the pH value of the aqueous solution a is 7.0-9.0, the soaking temperature of the cornea bandage mirror in the aqueous solution a is 20-40 ℃ and the soaking time is 1-8h.
4. The method for preparing a cornea bandage mirror with a bionic layer structure according to claim 1, wherein silver ions are provided by silver nitrate, the mass concentration of the silver ions in the aqueous solution B is 0.5-4%, the soaking temperature of the cornea bandage mirror in the aqueous solution B is 20-40 ℃, and the soaking time is 1-30min.
5. The method for producing a cornea bandage mirror having a bionic layer structure according to claim 1, wherein the polyamine group compound in the aqueous solution C comprises at least one of hexamethylenediamine and polyethylene polyamine; the mass concentration of the polyphenol compound in the aqueous solution C is 0.5-2%, and the mass concentration of the polyamine group compound in the aqueous solution C is 0.5-5%.
6. The method for producing a cornea bandage mirror with a bionic layer structure according to claim 5, wherein the immersing temperature of the cornea bandage mirror in the aqueous solution C is 25-40 ℃ and the immersing time is 12-48h.
7. The method for preparing a cornea bandage mirror with a bionic layer structure according to claim 1, wherein the solvent in the solution D is a mixture of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and water; wherein the mass fraction of the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in the solvent is 70-73%, the mass fraction of the N-hydroxysuccinimide in the solvent is 22-25%, and the mass fraction of the water in the solvent is 2-8%;
the mass concentration of the polyphenol compound in the solution D is 1-10%.
8. The method for preparing a cornea bandage mirror with a bionic layer structure according to claim 1 or 7, wherein the soaking temperature of the cornea bandage mirror in the solution D is 25-40 ℃ and the soaking time is 0.5-2h.
9. The method for preparing the cornea bandage mirror with the bionic layer structure according to claim 1, wherein the mass concentration of the sulfhydryl hyaluronic acid in the aqueous solution E is 1-5%, the soaking temperature of the cornea bandage mirror in the aqueous solution E is 25-40 ℃, and the soaking time is 0.1-2h.
10. A keratology with biomimetic layered structure, characterized in that it is produced by the method of any one of claims 1-9.
CN202311151277.XA 2023-09-07 2023-09-07 Cornea bandage mirror with bionic layer structure and preparation method thereof Active CN116870263B (en)

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