Preparation method and application of antibacterial medical gel coating

Abstract

The invention belongs to the technical field of biological materials, and discloses a preparation method and application of an antibacterial medical gel coating, and the method comprises the following steps: preparing a natural polymer solution; adding the fritillaria alkaloid into a natural polymer solution, and fully mixing to form a gel coating raw material; adding a cross-linking agent into the gel coating raw material, rapidly stirring, and dipping or spraying on the surface of the biomedical material to form the antibacterial medical gel coating; the antibacterial rate of the prepared antibacterial medical gel coating is greater than 90%, and the antibacterial rate of the antibacterial medical gel coating is still greater than 70% after the antibacterial medical gel coating is soaked in normal saline for 7 days; the raw materials are safe and natural, the biocompatibility is good, and the residual risk is avoided; the preparation process is simple, efficient and high in applicability; through a two-step core process of solution preparation and spraying / dipping, the operation is simple and convenient, complicated treatment conditions such as high temperature and high pressure are not needed, and the energy consumption is low; the preparation process is mild, and the activity of the fritillary alkaloid can be maintained.

Description

Technical Field

[0001] This invention belongs to the field of biomaterials technology, specifically relating to a method for preparing and applying an antibacterial medical gel coating. Background Technology

[0002] In the field of modern biomedical materials and tissue engineering, infection control remains one of the core issues affecting clinical treatment outcomes. Whether in surgical implants, wound repair materials, or long-term indwelling catheters, artificial joints, and dental implants, bacterial colonization and biofilm formation significantly increase the risk of infection. Once formed, biofilms not only exhibit extremely high resistance to conventional antibiotics but are also very difficult to clear through the body's immune system or drug treatment, easily leading to chronic, recurrent, and even life-threatening infections.

[0003] To address this challenge, researchers have proposed various antibacterial surface modification strategies, including metal ion or nanomaterial loading (such as silver, copper, and zinc), synthetic polymer antibacterial agent coatings, and functionalized surfaces such as photocatalysis and photothermal sterilization. However, these methods have certain limitations in clinical translation: metal ion materials may accumulate during long-term release, causing cytotoxicity and tissue damage, and even creating screening pressure for drug-resistant strains; some synthetic antibacterial agents (such as quaternary ammonium salts and polyguanidines) have strong bactericidal effects, but their biocompatibility is limited, and the safety of their degradation products in vivo is controversial; photocatalysis or photothermal technology relies on external excitation sources, limiting its application scenarios.

[0004] In recent years, natural medicinal active ingredients have attracted attention due to their wide availability, multi-target mechanisms of action, good biocompatibility, and low incidence of drug resistance. Fritillaria spp., a traditional Chinese medicine, is rich in various alkaloids. These alkaloids not only possess broad-spectrum antibacterial activity but also exhibit anti-inflammatory and antitussive activities. Compared with single-target synthetic antibacterial agents, the multi-target mechanism of Fritillaria alkaloids can effectively reduce the risk of drug resistance and has good inhibitory effects on both Gram-positive and Gram-negative bacteria. Therefore, directly introducing Fritillaria alkaloids into medical coating systems is a promising approach that combines safety and broad-spectrum antibacterial activity.

[0005] On the other hand, natural biodegradable polymers (such as chitosan, gelatin, and alginate) are widely used in wound dressings and drug delivery systems due to their excellent biocompatibility, biodegradability, and film-forming and gel-forming properties. If fritillaria alkaloids can be efficiently combined with such polymer matrices and a stable three-dimensional gel network can be constructed through appropriate cross-linking reactions, the dual effects of sustained release and long-lasting antibacterial action can be achieved while maintaining the stability of the active ingredients. Furthermore, the softness and good adhesion of the gel coating can effectively cover wounds or implanted surfaces, reducing bacterial adhesion and promoting tissue repair.

[0006] While there are reports of introducing plant extracts into medical materials, most of them have the following shortcomings: ① The active ingredients are unstable during processing and are easily deactivated; ② The binding efficiency with the polymer matrix is ​​low, resulting in insufficient antibacterial durability; ③ The coating molding method is complex or the conditions are harsh, which limits industrial application. Summary of the Invention

[0007] One object of the present invention is to solve at least the above-mentioned problems and / or defects, and to provide at least the advantages described below.

[0008] To achieve these objectives and other advantages of the present invention, a method for preparing an antibacterial medical gel coating is provided, comprising the following steps: Step 1: Add the natural polymer to the solvent to prepare a natural polymer solution; Step 2: Add fritillaria alkaloids to the natural polymer solution and mix thoroughly to form a gel coating material; Step 3: Add a crosslinking agent to the gel coating material, stir rapidly, then immerse the biomedical material in the coating material to achieve adhesion, and then remove it to allow it to complete the gel reaction and form an antibacterial medical gel coating, or spray the gel coating material onto the surface of the biomedical material to form an antibacterial medical gel coating on the surface of the biomedical material.

[0009] Preferably, in step one, the concentration of the natural polymer solution is 0.01wt% to 20wt%, the natural polymer is one or more of chitosan, dextran, gelatin, collagen, hyaluronic acid, chitin, cellulose, and sodium alginate, and the solvent is one of water and acetic acid solution, with the concentration of the acetic acid solution being 1 to 2wt%.

[0010] Preferably, in step two, the fritillaria alkaloids are one or more of fritillaria alkaloids, fritillaria alkaloid A, fritillaria alkaloid B, fritillaria alkaloid sine ...ketone alkaloid.

[0011] Preferably, in step two, the amount of fritillaria alkaloids added is 0.05wt% to 3wt% of the total mass of the gel coating raw material.

[0012] Preferably, in step two, the fritillaria alkaloids are added to the natural polymer solution in any one of the following forms: aqueous solution, acetic acid extract, ethanol extract, ethyl acetate extract, suspension, or powder.

[0013] Preferably, in step three, the amount of crosslinking agent added is 0.1wt% to 0.5wt% of the gel coating raw material, and the crosslinking agent is either glutaraldehyde or calcium chloride.

[0014] Preferably, before step one, the natural polymer is modified by means of: S11. Dissolve the natural polymer in an organic solvent, add succinic anhydride, adjust the pH with triethylamine, stir and react for a period of time, and then adjust the pH to neutral with hydrochloric acid to obtain a pretreated natural polymer solution. S12. Dissolve the antimicrobial peptide in phosphate buffer to prepare an antimicrobial peptide solution. Then add the antimicrobial peptide solution to the natural polymer solution and stir at low speed in the dark. After the reaction is complete, dialyze with phosphate buffer solution and finally freeze dry to obtain the modified natural polymer.

[0015] Preferably, in S11, the organic solvent is dimethyl sulfoxide, the ratio of natural polymer to organic solvent is 10g:100~150mL, the mass ratio of natural polymer to succinic anhydride is 10:0.2~0.3, the pH is adjusted to 5~6 with triethylamine, and the reaction is stirred at room temperature for 5~6h.

[0016] Preferably, in step S12, the ratio of antimicrobial peptide to phosphate buffer solution is 1-3 mg:1 mL, the concentration of phosphate buffer solution is 0.01 M, and when adding the antimicrobial peptide solution, it is added according to the mass ratio of natural polymer to antimicrobial peptide of 1:0.11-0.12. The stirring speed is 100-200 rpm, and the stirring reaction is carried out for 8-12 hours. During dialysis, a dialysis bag with a molecular weight cutoff of 3500-4000 Da is used, the dialysate is 0.01 M phosphate buffer solution, and dialysis is carried out for 24-32 hours, with the dialysate being replaced every 8 hours.

[0017] Preferably, in step three, the antibacterial medical gel coating formed on the surface of the biomedical material is a dry film or a wet coating.

[0018] The present invention also provides an application of an antibacterial medical gel coating, wherein the antibacterial medical gel coating is attached to the surface of a biomedical material to inhibit the growth and reproduction of fungi and bacteria.

[0019] The present invention has the following beneficial effects: 1. Excellent broad-spectrum antibacterial properties: This invention utilizes fritillaria alkaloids as the core active antibacterial component, which, after being embedded in a natural polymer gel network, can significantly inhibit the growth and reproduction of a variety of key pathogenic fungi and putrefactive bacteria. The resulting antibacterial medical gel coating has an inhibition rate of more than 90%, and the antibacterial medical gel coating still has an inhibition rate of more than 70% after being soaked in physiological saline for 7 days. 2. The raw materials are safe and natural, with good biocompatibility and no risk of residue: All the main components of the coating prepared by this invention are derived from natural sources, are safe to eat or have a long history of medicinal and edible use (such as some fritillaria bulbs being used as ingredients in health products), and do not contain any chemically synthesized preservatives or bactericides. 3. The preparation process is simple, efficient, and highly applicable: The method of this invention uses two core processes, solution preparation and spraying / immersion, which are simple and convenient to operate, do not require complex treatment conditions such as high temperature and high pressure, and have low energy consumption; the preparation process is mild, which is conducive to maintaining the activity of fritillaria alkaloids. Attached Figure Description

[0020] Figure 1 The antibacterial medical gel coatings prepared in Examples 1-3, the silver nanoparticle coating in Comparative Example 2, and the antibacterial rate of untreated Comparative Example 1 are shown. Figure 2 The antibacterial rates of the antibacterial medical gel coatings prepared in Examples 1-3, the silver nanoparticle coating in Comparative Example 2, and Comparative Example 1 after being soaked in physiological saline for 7 days and then subjected to antibacterial experiments are shown. Figure 3 Photographs of cells cultured for 3 days on the antibacterial medical gel coatings prepared in Examples 1-3; Figure 4 The absorbance of cells after 3 and 5 days of co-culturing in well plates, with the antibacterial medical gel coating prepared in Examples 1-3, and with the silver nanoparticle coating of Comparative Example 2. Figure 5 The antibacterial rates of Examples 1 and 4, and the antibacterial rate after soaking in physiological saline for 7 days before conducting the antibacterial experiment; Figure 6 The absorbance of cells after 3 and 5 days of co-culturing in well plates with the antibacterial medical gel coatings prepared in Examples 1 and 4. Detailed Implementation

[0021] The present invention will now be described in further detail with reference to the embodiments and accompanying drawings, so that those skilled in the art can implement it based on the description.

[0022] Example 1 A method for preparing an antibacterial medical gel coating includes the following steps: Step 1: Add 2g of chitosan (degree of deacetylation > 90%) to 98mL of 1wt% acetic acid solution, and then make up to 100mL to prepare a 2.0wt% chitosan-acetic acid aqueous solution. Step 2: Concentrate the water extract of Fritillaria cirrhosa (containing total alkaloids such as Fritillaria cirrhosa and Fritillaria cirrhosa) into an aqueous solution with a solid content of 15%. Then, add 2 mL of the water extract of Fritillaria cirrhosa to 98 mL of chitosan acetic acid aqueous solution (so that the final amount of Fritillaria cirrhosa alkaloids added is 0.3 wt% of the total amount of gel coating). Stir magnetically for 1 hour to form the gel coating material. Step 3: Add 0.1g of glutaraldehyde as a crosslinking agent to the above 100mL gel coating material, stir evenly, and then spray the gel coating material onto the surface of the biomedical material to form an antibacterial medical gel coating on the surface of the biomedical material.

[0023] Example 2 A method for preparing an antibacterial medical gel coating includes the following steps: Step 1: Dissolve 1.5g of sodium alginate in 98mL of water, and bring the volume up to 100mL to prepare a 1.5wt% sodium alginate aqueous solution. Step 2: Dissolve 1g of fritillary alkaloid A standard (purity >98%) in a small amount of edible ethanol, then add pure water to prepare a 1.0wt% aqueous solution. Take 199mL of sodium alginate aqueous solution and add 1mL of fritillary alkaloid A solution (so that the final amount of fritillary alkaloid added is 0.5wt% of the total amount of gel coating). Sonicate for 10 minutes to ensure uniform dispersion and form the gel coating material. Step 3: Add 0.2g of calcium chloride to the above 200mL gel coating material, then immerse the biomedical material in the gel coating material, and then remove it to allow it to complete the gel reaction and form an antibacterial medical gel coating.

[0024] Example 3 A method for preparing an antibacterial medical gel coating includes the following steps: Step 1: Add 2g of gelatin and 0.5g of dextran to 98mL of water, heat to 60℃ to dissolve, and then bring the volume up to 100mL to obtain a composite aqueous solution of gelatin (2.0wt%) and dextran (0.5wt%). Step 2: Take 100 mL of gelatin and dextran composite aqueous solution, cool it to 40℃, add 0.25 g of fritillaria alkaloid powder (purity >95%, micron particle size <50 μm), and disperse it in a high-speed homogenizer at 8000 rpm for 5 min to form a stable suspension, thus obtaining the gel coating raw material (the final amount of fritillaria alkaloid added is 0.25 wt% of the total amount of gel coating). Step 3: Add 0.1g of glutaraldehyde as a crosslinking agent to the above 100mL gel coating material, stir evenly, and then spray the gel coating material onto the surface of the biomedical material to form an antibacterial medical gel coating on the surface of the biomedical material.

[0025] Example 4 A method for preparing an antibacterial medical gel coating includes the following steps: Step 1 includes: S11. Dissolve 10g of chitosan in 100mL of dimethyl sulfoxide, add 0.2g of succinic anhydride, adjust the pH to 6 with triethylamine, stir and react for 5h, then adjust the pH to neutral with hydrochloric acid to obtain a pretreated chitosan solution. S12. Dissolve 1 mg of antimicrobial peptide in 1 mL of 0.01 M phosphate buffer to prepare an antimicrobial peptide solution of 1 mg / mL. Then add 110 mL of the antimicrobial peptide solution to 100 mL of chitosan solution and stir at low speed of 150 rpm for 8 h in the dark. After the reaction is complete, dialyze with 0.01 M phosphate buffer solution using a dialysis bag with a molecular weight cutoff of 3500 Da for 24 h. Change the dialysate every 8 h. Finally, freeze dry to obtain modified chitosan. S13. Add 2g of modified chitosan to 98mL of 1wt% acetic acid solution, and then adjust the volume to 100mL to prepare a 2.0wt% modified chitosan acetic acid aqueous solution. Step 2: Concentrate the water extract of Fritillaria cirrhosa (containing total alkaloids such as Fritillaria cirrhosa and Fritillaria cirrhosa) into an aqueous solution with a solid content of 15%. Then, add 2 mL of the water extract of Fritillaria cirrhosa to 98 mL of modified chitosan acetic acid aqueous solution (so that the final amount of alkaloids added is 0.3 wt% of the total amount of coating). Stir magnetically for 1 hour to form a gel coating material. Step 3: Add 0.1g of glutaraldehyde as a crosslinking agent to the above 100mL gel coating material, stir evenly, and then spray the gel coating material onto the surface of the biomedical material to form an antibacterial medical gel coating on the surface of the biomedical material.

[0026] Comparative Example 1 The biomedical materials used in this comparative example have no coating on their surface.

[0027] Comparative Example 2 This comparative example uses common silver nanoparticles as an antibacterial coating on the surface of biomedical materials.

[0028] The antibacterial medical gel coatings prepared in Examples 1-4 and Comparative Example 2 were subjected to antibacterial tests. The antibacterial medical gel coatings were co-cultured with bacteria, and the bacteria were counted by absorbance. Sterility was negative (0), and 100% bacteria was positive (100). The antibacterial inhibition rate of the antibacterial medical gel coatings against bacteria was calculated. Figure 1 The antibacterial medical gel coatings prepared in Examples 1-3, the silver nanoparticle coating in Comparative Example 2, and the antibacterial rate of untreated Comparative Example 1 show that the antibacterial medical gel coatings activated by Fritillaria alkaloids have good inhibitory ability against bacteria (all exceeding 90%). Figure 2 The antibacterial rates of the antibacterial medical gel coatings prepared in Examples 1-3, the silver nanoparticle coating in Comparative Example 2, and Comparative Example 1 after soaking in physiological saline for 7 days were compared. It can be seen that the gelatin / dextran-based, chitosan-based, and sodium alginate-based coatings still exhibited excellent antibacterial properties (>70%). Figure 3The images show cells cultured on the antibacterial medical gel coatings prepared in Examples 1-3 for 3 days (a corresponds to Example 1, b corresponds to Example 2, and c corresponds to Example 3). It can be seen that the cells are still adhered to the material (cells can only adhere to the material when they are alive), indicating that the antibacterial medical gel coatings prepared in Examples 1-3 have good biocompatibility. Figure 4 The absorbance values ​​are measured after cells are co-cultured in a well plate, the antibacterial medical gel coating prepared in Examples 1-3, and the silver nanoparticle coating in Comparative Example 2 for 3 and 5 days. Higher absorbance indicates a larger number of cells and excellent biocompatibility of the well plate, hence its highest absorbance. Notably, the absorbance of the antibacterial medical gel coating prepared in Examples 1-3 is close to that of the well plate, indicating that the cell compatibility of the antibacterial medical gel coating is similar to that of the well plate, both being very good. In contrast, the absorbance of the silver nanoparticle coating in Comparative Example 2 is low, indicating poor biocompatibility.

[0029] Furthermore, Example 4 of this invention modifies the natural polymer by grafting antimicrobial peptides onto it. The antibacterial rate of Example 4 and its antibacterial effect after soaking in physiological saline for 7 days are shown in the figure. Figure 5 As shown, the antibacterial medical gel coating grafted with antimicrobial peptides effectively improves the antibacterial ability; the absorbance of the antibacterial medical gel coating prepared in Example 4 is as follows: Figure 6 As shown, the absorbance of Example 4 is very close to that of the well plate, indicating that it has good biocompatibility.

[0030] Although embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, the present invention is not limited to the specific details and illustrations shown and described herein.

Claims

1. A method of preparing an antimicrobial medical gel coating, characterized by, The method comprises the following steps: Step one, adding natural polymer into solvent to prepare natural polymer solution; Step two, adding fritillaria alkaloids into natural polymer solution, mixing well to form gel coating raw material; Step three, adding crosslinking agent into gel coating raw material, stirring rapidly, then dipping biomaterial into coating raw material to realize adhesion, then taking out to complete gel reaction and form antibacterial medical gel coating, or spraying gel coating raw material on biomaterial surface to form antibacterial medical gel coating.

2. The method for preparing the antibacterial medical gel coating as described in claim 1, characterized in that, In step one, the concentration of natural polymer solution is 0.01wt%-20wt%, the natural polymer is one or more of chitosan, dextran, gelatin, collagen, hyaluronic acid, chitin, cellulose and sodium alginate, and the solvent is one of water and acetic acid solution with a concentration of 1-2wt%.

3. The method for preparing the antibacterial medical gel coating as described in claim 1, characterized in that, In step two, the fritillaria alkaloids are one or more of fritillaria sinica alkaloids, fritillarin A, fritillarin B, veratrum alkaloids, isosachalinine, chuanbeium and chuanbeium ketone.

4. The method for preparing the antibacterial medical gel coating as described in claim 1, characterized in that, In step two, the addition amount of fritillaria alkaloids is 0.05wt%-3wt% of the total mass of gel coating raw material.

5. The method of making an antimicrobial medical gel coating of claim 1, wherein, In step two, the fritillaria alkaloids are added into natural polymer solution in the form of water solution, acetic acid extract, ethanol extract, ethyl acetate extract, suspension or powder.

6. The method of making an antimicrobial medical gel coating of claim 1, wherein, In step three, the addition amount of crosslinking agent is 0.1wt%-0.5wt% of gel coating raw material, and the crosslinking agent is one of glutaraldehyde and calcium chloride.

7. The method of making an antimicrobial medical gel coating of claim 1, wherein, Before step one, the natural polymer is modified by the following method: S11, dissolving natural polymer in organic solvent, adding succinic anhydride, adjusting pH with triethylamine, stirring for a period of time, then adjusting pH to neutral with hydrochloric acid to obtain pretreated natural polymer solution; S12, dissolving antibacterial peptide in phosphate buffer solution to prepare antibacterial peptide solution, then adding antibacterial peptide solution into natural polymer solution, stirring at low speed in dark, dialyzing with phosphate buffer solution after reaction, and finally freeze-drying to obtain modified natural polymer.

8. The method of claim 7, wherein the antimicrobial medical gel coating is prepared by the steps of: In S11, the organic solvent is dimethyl sulfoxide, the dosage ratio of natural polymer to organic solvent is 10g:100-150mL, the mass ratio of natural polymer to succinic anhydride is 10:0.2-0.3, and the pH is adjusted to 5-6 with triethylamine, and the stirring reaction is carried out at room temperature for 5-6h; In S12, the dosage ratio of antibacterial peptide to phosphate buffer solution is 1-3mg:1mL, the concentration of phosphate buffer solution is 0.01M, the addition of antibacterial peptide solution is carried out according to the mass ratio of natural polymer to antibacterial peptide of 1:0.11-0.12, the stirring speed is 100-200rpm, the stirring reaction is carried out for 8-12h, the dialysis bag with a molecular weight cut-off of 3500-4000Da is used for dialysis, the dialysis liquid is 0.01M phosphate buffer solution, and the dialysis is carried out for 24-32h, and the dialysis liquid is replaced every 8h.

9. The method of making an antimicrobial medical gel coating of claim 1, wherein, In step three, the antibacterial medical gel coating formed on the surface of biomaterial is dry film or wet coating.

10. Use of an antimicrobial medical gel coating prepared according to the method of any one of claims 1 to 9, characterized in that, The antimicrobial medical gel coating is attached to the surface of a biomaterial to inhibit the growth and proliferation of fungi and bacteria.

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