Silver-loaded nano-cluster-adriamycin composite hydrogel and preparation method thereof

Abstract

The invention discloses a preparation method of silver-loaded nano-cluster-adriamycin composite hydrogel. The method comprises the following steps: preparing a silver nano-cluster from an AgNO3 solution and a gallic acid solution; preparing a prepolymer from chitosan, hyaluronic acid, an acetic acid solution and methacrylic anhydride; mixing silver nanoclusters, a doxorubicin hydrochloride solution and the prepolymer, and adding a photoinitiator to obtain preliminarily formed hydrogel; and immersing the primarily formed hydrogel into a sodium citrate solution to obtain the silver-loaded nano-cluster-adriamycin composite hydrogel. The composite hydrogel prepared by the invention has a pH-responsive drug release characteristic, the drug release is accelerated in an acidic environment, and the drug release is slow in a neutral environment; the antibacterial agent has an inhibiting effect on bacteria such as escherichia coli and staphylococcus aureus, and shows good antibacterial performance; the toxicity to cancer cells is obviously higher than that of normal cells, and certain tumor targeting is achieved; and the polymer has good mechanical strength and good swelling performance in different pH environments, and has a wide application prospect.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a silver-loaded nanocluster-doxacin composite hydrogel and its preparation method. Background Technology

[0002] In the biomedical field, with the increasing demand for disease treatment and tissue repair, the development of novel materials with multifunctional properties has become a research hotspot. Hydrogels, as polymeric materials with a three-dimensional network structure capable of absorbing large amounts of water, exhibit enormous application potential in drug delivery, wound healing, and tissue engineering due to their similarity to biological tissues.

[0003] In drug delivery, traditional drug delivery systems often struggle to achieve precise drug release. The microenvironment of tumor tissue differs from that of normal tissue; for example, tumor tissue is typically acidic. Traditional drug delivery systems cannot effectively utilize this difference for targeted drug release, leading to the release of large amounts of drug into normal tissue, increasing side effects, and simultaneously reducing drug concentration at the tumor site, thus impacting treatment efficacy. Therefore, developing pH-responsive drug delivery systems that enable specific drug release within the tumor microenvironment is crucial for improving cancer treatment outcomes.

[0004] Infection is a significant factor affecting wound healing. Currently, commonly used antibacterial materials in clinical practice suffer from narrow antibacterial spectra and are prone to drug resistance. Therefore, developing novel materials with highly effective antibacterial properties and low resistance to drug development is of great importance for preventing and treating wound infections and promoting wound healing.

[0005] In the field of tissue engineering, ideal tissue engineering scaffold materials need to possess good mechanical properties and biocompatibility. Traditional scaffold materials often lack sufficient mechanical properties, making them unable to withstand external forces in the physiological environment, and their cytotoxicity may also affect the effectiveness of tissue repair and regeneration. Furthermore, the swelling properties of the material can also affect its stability in vivo and drug release behavior. Therefore, developing tissue engineering scaffold materials with good mechanical properties, low cytotoxicity, and suitable swelling properties is a current research focus.

[0006] In recent years, nanomaterials have been widely used in the biomedical field due to their unique physicochemical properties. For example, silver nanoclusters (AgNCs) possess excellent optical properties and antibacterial activity, while natural polymers such as chitosan and hyaluronic acid exhibit good biocompatibility and biodegradability. Combining these materials holds promise for preparing multifunctional composite hydrogels to meet diverse needs in the biomedical field.

[0007] Therefore, there is a need for a silver-loaded nanocluster-doxacin composite hydrogel and its preparation method. Summary of the Invention

[0008] The technical problem to be solved by this invention is to address the shortcomings of the prior art by providing a method for preparing a silver-loaded nanocluster-doxorubicin composite hydrogel. This method, through the rational design and preparation of the composite hydrogel, comprehensively utilizes the advantages of each component, endowing it with multiple functions such as pH-responsive drug release, antibacterial properties, low cytotoxicity, good mechanical properties, and swelling properties. This provides a novel, high-performance, multifunctional material for fields such as tumor treatment, wound healing, and tissue engineering.

[0009] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is: a method for preparing silver-loaded nanoclusters-doxacin composite hydrogel, characterized in that the method includes the following steps: Step 1: Stir AgNO3 solution and gallic acid solution evenly, then add sodium hydroxide solution dropwise until the pH is 8-9, and then treat with water bath to obtain silver nanocluster dispersion; Step 2: Mix chitosan, hyaluronic acid, acetic acid solution and methacrylic anhydride to react and obtain photosensitive prepolymer; Step 3: Mix the silver nanocluster dispersion obtained in Step 1, the doxorubicin hydrochloride solution, and the photosensitive prepolymer obtained in Step 2 evenly, and then add phenyl-2,4,6-trimethylbenzoyl lithium phosphine as a photoinitiator to carry out a photocrosslinking reaction to obtain a preliminary hydrogel. Step 4: Immerse the preliminarily formed hydrogel obtained in Step 3 into sodium citrate solution to carry out ionic cross-linking reaction, and obtain silver-loaded nanoclusters-doxacin composite hydrogel.

[0010] This invention involves a redox reaction between AgNO3 and gallic acid under alkaline conditions via a water bath, where gallic acid acts as a reducing agent to react with AgNO3. +The process involves reducing the substance to Ag, while simultaneously encapsulating it as a stabilizer to form silver nanoclusters (AgNCs), preventing aggregation and creating a stable dispersion. Chitosan, hyaluronic acid, and acetic acid solutions are mixed; the acetic acid provides an acidic environment to promote chitosan dissolution, and the chitosan and hyaluronic acid swell and dissolve in the acetic acid solution. Subsequently, they undergo esterification with methacrylic anhydride, introducing double bonds into the polysaccharide molecules, endowing them with photoresponsiveness, and forming a photocrosslinkable photosensitive prepolymer. The silver nanocluster dispersion, doxorubicin hydrochloride (DOX) solution, and the photosensitive prepolymer are then mixed... The mixture is homogenized, and then lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) is added as a photoinitiator. In the photocrosslinking reaction under light irradiation, the photoinitiator initiates the free radical polymerization of the double bonds in the photosensitive prepolymer to form a three-dimensional crosslinked network, achieving the initial formation of the hydrogel. At the same time, silver nanoclusters and doxorubicin are loaded. By immersing the initially formed hydrogel in sodium citrate solution, the citrate ions coordinate with the amino groups of chitosan to carry out ionic crosslinking, further enhancing the network structure, improving mechanical strength and stability, and finally forming a composite hydrogel.

[0011] This invention constructs a stable network through a dual crosslinking mechanism of photocrosslinking and ionic crosslinking, balancing molding efficiency and structural strength; it steps to encapsulate silver nanoclusters and doxorubicin to achieve synergistic antibacterial and antitumor functions, with controllable conditions at each step, making it suitable for large-scale preparation.

[0012] The preparation method of the above-mentioned silver nanocluster-doxacin composite hydrogel is characterized in that the concentration of the AgNO3 solution in step one is 0.08 mM to 0.12 mM, the concentration of the gallic acid solution is 1.8 mM to 2.2 mM, and the volume ratio of the AgNO3 solution to the gallic acid solution is 1:2 to 4. This invention ensures complete reduction of Ag⁺ by controlling the concentration and volume ratio of the AgNO3 solution and the gallic acid solution, while providing sufficient stabilizer to prevent the aggregation of silver nanoclusters, ensuring their dispersibility and antibacterial activity, and controlling their characteristic peak to 420 nm to 430 nm to maintain stability.

[0013] The preparation method of the above-mentioned silver nanocluster-doxorubicin composite hydrogel is characterized in that the water bath treatment in step one is carried out at a temperature of 50℃~60℃ for 1.5h~2.5h. This invention, by controlling the parameters of the water bath, forms uniform and stable silver nanoclusters, preventing incomplete reaction due to excessively low temperature and insufficient AgNCs aggregation due to excessively high temperature. It also prevents insufficient reaction due to excessively short time and excessively long time, which may result in excessively large nanocluster sizes.

[0014] The preparation method of the above-mentioned silver nanocluster-doxorubicin composite hydrogel is characterized in that the characteristic peak of the silver nanocluster dispersion in step one is 420 nm to 430 nm. This invention controls the characteristic peak of the silver nanocluster dispersion because the characteristic peak of 420 nm to 430 nm is a marker of stable AgNCs.

[0015] The preparation method of the above-mentioned silver-loaded nanoclusters-doxacin composite hydrogel is characterized in that, in step two, the mass ratio of chitosan, hyaluronic acid, acetic acid, and methacrylic anhydride is 1.8~2.5:0.8~1.2:100:4.5~6, the degree of deacetylation of the chitosan is 83%~87%, and the concentration of the acetic acid solution is 0.08M~0.12M. This invention controls the degree of deacetylation of chitosan to balance solubility and reactivity; controls the amount of chitosan added to ensure the strength of the network framework; controls the amount of acetic acid added to ensure uniform dissolution of the polysaccharide, preventing degradation due to excessive concentration and insufficient dissolution due to insufficient concentration; controls the amount of hyaluronic acid added to enhance biocompatibility and prevent insufficient mechanical strength due to excessive addition; and controls the amount of methacrylic anhydride added to introduce an appropriate amount of photosensitive groups to ensure photocrosslinking efficiency, preventing reduced biocompatibility due to excessive addition and insufficient crosslinking due to insufficient addition.

[0016] The preparation method of the above-mentioned silver-loaded nanoclusters-doxacin composite hydrogel is characterized in that the mixing reaction temperature in step two is 35℃~40℃, and the time is 5h~7h. This invention prevents the problems of methacrylic anhydride volatilization or polysaccharide degradation due to excessively high temperatures and slow reaction rates due to excessively low temperatures by controlling the mixing reaction temperature. Simultaneously, by controlling the time, it ensures sufficient esterification reaction and guarantees the amount of photosensitive groups introduced.

[0017] The preparation method of the above-mentioned silver nanocluster-doxorubicin composite hydrogel is characterized in that, in step three, the volume ratio of the silver nanocluster dispersion, doxorubicin hydrochloride solution, and photosensitive prepolymer is 0.8~1.2:0.8~1.2:3~5; the concentration of the doxorubicin hydrochloride solution is 0.8 mg / mL~1.2 mg / mL; and the added mass of lithium phenyl-2,4,6-trimethylbenzoylphosphine is 0.4%~0.6% of the total mass of the silver nanocluster dispersion, doxorubicin hydrochloride solution, and photosensitive prepolymer. This invention ensures uniform loading of the drug and AgNCs by controlling the volume ratio of the silver nanocluster dispersion, doxorubicin hydrochloride solution, and photosensitive prepolymer, as well as the concentration of the doxorubicin hydrochloride solution. The concentration balances efficacy and hydrogel stability. By controlling the amount of LAP added, the crosslinking efficiency and biosafety are balanced, preventing incomplete crosslinking due to insufficient addition and increased toxicity due to residual initiator caused by excessive addition.

[0018] The preparation method of the above-mentioned silver-loaded nanoclusters-doxacin composite hydrogel is characterized in that the wavelength of the light source used in the photocrosslinking reaction in step three is 400nm~410nm, and the irradiation time of the light source in the photocrosslinking reaction is 25s~35s. This invention controls the wavelength of the light source to match the photoresponse characteristics of LAP, and controls the time to ensure complete crosslinking and avoid excessive irradiation that could damage the drug or nanoclusters.

[0019] The preparation method of the above-mentioned silver-loaded nanoclusters-doxacin composite hydrogel is characterized in that the mass fraction of the sodium citrate solution in step four is 1.5%~2.5%, and the ionic crosslinking reaction time is 10h~15h. This invention prevents problems such as excessively dense networks affecting swelling and drug release due to excessively high concentrations and insufficient ionic crosslinking due to excessively low concentrations by controlling the mass fraction of the sodium citrate solution. By controlling the ionic crosslinking reaction time, sufficient crosslinking is ensured, forming a dual network in synergy with photocrosslinking, thereby improving mechanical properties.

[0020] In addition, this application also discloses a silver-loaded nanocluster-doxacin composite hydrogel, which is prepared by the above method.

[0021] Compared with the prior art, the present invention has the following advantages: 1. The composite hydrogel prepared by this invention has many excellent properties. It has pH-responsive drug release characteristics, with faster drug release in acidic environments and slower release in neutral environments; it has inhibitory effects on bacteria such as Escherichia coli and Staphylococcus aureus, exhibiting good antibacterial properties; its toxicity to cancer cells is significantly higher than that to normal cells, showing certain tumor targeting; it has good mechanical strength, elasticity, and good swelling properties under different pH environments. These properties make this composite hydrogel of great significance.

[0022] 2. The composite hydrogel prepared in this invention has the advantages of pH-responsive drug release and selective killing of cancer cells in tumor treatment, which helps to achieve precise treatment, improve efficacy, and reduce damage to normal tissues. Its good antibacterial properties make it very useful in wound dressings and medical devices, effectively preventing and treating infections and promoting wound healing. Its low cytotoxicity ensures its safety in biomedical applications and broadens its application scope. Its excellent mechanical and swelling properties provide a solid foundation for its use as a tissue engineering scaffold and drug delivery system, and have broad application prospects.

[0023] 3. The drug release test results of the composite hydrogel prepared in this invention show that the composite hydrogel has obvious pH-responsive drug release characteristics. In an environment with pH 7.4, drug release is relatively slow, with a release rate of only 8.2%±1.3% in 2 hours and a cumulative release of only 21.5%±2.8% in 72 hours. However, in acidic environments with pH 6.5 and 5.5, the drug release rate is significantly accelerated, with a cumulative release of 82.4%±3.5% and 97.1%±1.8% in 72 hours, respectively. This pH-responsive drug release characteristic makes the composite hydrogel potentially valuable in fields such as tumor treatment, because the microenvironment of tumor tissue is usually acidic. The composite hydrogel can release drugs more effectively at the tumor site, improve the therapeutic effect, and at the same time reduce drug exposure to normal tissues and reduce side effects.

[0024] 4. The antibacterial performance test results of the composite hydrogel prepared in this invention show that it has an inhibitory effect on both Escherichia coli and Staphylococcus aureus. The inhibition zone diameter for Escherichia coli is 15.2 mm ± 0.8 mm, and the minimum inhibitory concentration (MIC) is 16 μg / mL; the inhibition zone diameter for Staphylococcus aureus is 13.7 mm ± 1.1 mm, and the MIC is 32 μg / mL. This indicates that the composite hydrogel can effectively inhibit bacterial growth and has application potential in wound dressings, medical devices, and other fields. It can prevent and treat infections and promote wound healing.

[0025] 5. The cytotoxicity test results of the composite hydrogel prepared in this invention show that there are significant differences in toxicity to normal cells and cancer cells. The half-maximal inhibitory concentration (IC50) for MCF-7 cancer cells is 0.38 μg / mL ± 0.05 μg / mL, while the IC50 for L929 normal cells is 12.6 μg / mL ± 1.2 μg / mL. This means that the composite hydrogel can inhibit cancer cells at a relatively low concentration, while having relatively low toxicity to normal cells. It has a certain degree of tumor targeting and can reduce damage to normal cells in cancer treatment, thereby improving the safety and effectiveness of treatment.

[0026] 6. The mechanical property test results of the composite hydrogel prepared by this invention show that the compressive modulus of the composite hydrogel is 28.5 kPa ± 2.3 kPa, which is 42% higher than that of the single crosslinked system. This indicates that the composite hydrogel has good mechanical strength and elasticity, and can withstand certain external forces without easily deforming or being damaged. At the same time, its swelling rate is 358.66% ± 16.32% at pH 7.4 and 621.79% ± 20.24% at pH 5.5, indicating that the composite hydrogel has good swelling performance under different pH environments and can absorb and retain a certain amount of water. This is of great significance for its application in the biomedical field, such as tissue engineering scaffolds and drug delivery systems.

[0027] 7. This invention lays the foundation for the application of silver-loaded nanoclusters-doxacin composite hydrogels in the biomedical field by controlling the preparation parameters of the composite hydrogels and optimizing their performance. This is based on performance optimization and takes into account the structural stability, functional effectiveness (pH-responsive release, antibacterial, tumor targeting) and biosafety of the composite hydrogels.

[0028] The technical solution of the present invention will be further described in detail below through embodiments. Detailed Implementation

[0029] Example 1 This embodiment includes the following steps: Step 1: Mix 0.1 mM AgNO3 solution and 2 mM gallic acid solution in a clean beaker at a volume ratio of 1:3. Stir thoroughly with a magnetic stirrer until the mixture is homogeneous. Then, slowly add sodium hydroxide solution dropwise while monitoring the pH value of the solution in real time with a pH meter. Adjust the pH of the solution to 8.5, then transfer it to a constant temperature water bath at 55°C and react for 2 hours. During the reaction, the solution gradually turns wine-red. After the reaction is complete, a wine-red AgNCs dispersion is obtained. The obtained AgNCs dispersion is detected using a UV-Vis spectrophotometer, and its characteristic peak is determined to be at 425 nm. Step 2: Dissolve chitosan and hyaluronic acid with a degree of deacetylation of 85% in a 0.1M acetic acid solution. Stir thoroughly on a magnetic stirrer until completely dissolved to form a homogeneous solution. Then add methacrylic anhydride and continue stirring until homogeneous. Transfer the solution to a 37°C constant temperature water bath and react for 6 hours to obtain the photosensitive prepolymer. The mass ratio of chitosan, hyaluronic acid, acetic acid and methacrylic anhydride is 2:1:100:5. Step 3: Mix the AgNCs dispersion obtained in Step 1 (volume ratio 1:1:4), the DOX solution (concentration 1 mg / mL), and the photosensitive prepolymer obtained in Step 2 in a clean container. Stir thoroughly with a magnetic stirrer until the three are evenly mixed. Then add 0.5% LAP of the total mass of AgNCs dispersion, DOX solution, and photosensitive prepolymer and stir evenly. Pour the mixture into a mold and irradiate with an LED light source with a wavelength of 405 nm for 30 seconds to carry out the photocrosslinking reaction and achieve preliminary molding, resulting in a preliminary hydrogel. Step 4: Remove the pre-formed hydrogel obtained in Step 3 from the mold and immerse it in a 2% sodium citrate solution for 12 hours to carry out an ionic crosslinking reaction, finally obtaining a composite hydrogel.

[0030] The composite hydrogel prepared in this embodiment was subjected to in vitro drug release tests, antibacterial performance tests (ASTM E2149 standard), cytotoxicity tests (CCK-8 method), and mechanical property tests. The results are as follows: The results of the in vitro drug release test are shown in Table 1; Table 1

[0031] The results of the antimicrobial performance test (ASTM E2149 standard) are shown in Table 2; Table 2

[0032] The results of the cytotoxicity assay (CCK-8 assay) are shown in Table 3; Table 3

[0033] The results of the mechanical performance test are as follows: Compression modulus: 28.5 kPa ± 2.3 kPa (42% higher than the single crosslinked system); Swelling rate: 358.66%±16.32% (pH 7.4), 621.79%±20.24% (pH 5.5).

[0034] Example 2 This embodiment includes the following steps: Step 1: Mix 0.1 mM AgNO3 solution and 2 mM gallic acid solution in a clean beaker at a volume ratio of 1:2. Stir thoroughly with a magnetic stirrer until the mixture is homogeneous. Then, slowly add sodium hydroxide solution dropwise while monitoring the pH value of the solution in real time with a pH meter. Adjust the pH of the solution to 9.0, then transfer it to a constant temperature water bath at 60°C and react for 1.5 hours. During the reaction, the solution gradually turns wine-red. After the reaction is complete, a wine-red AgNCs dispersion is obtained. The obtained AgNCs dispersion is detected using a UV-Vis spectrophotometer, and its characteristic peak is determined to be 430 nm. Step 2: Dissolve chitosan and hyaluronic acid with a degree of deacetylation of 83% in a 0.1M acetic acid solution. Stir thoroughly on a magnetic stirrer until completely dissolved to form a homogeneous solution. Then add methacrylic anhydride and continue stirring until homogeneous. Transfer the solution to a 40°C constant temperature water bath and react for 5 hours to obtain the photosensitive prepolymer. The mass ratio of chitosan, hyaluronic acid, acetic acid and methacrylic anhydride is 1.5:0.8:100:4. Step 3: Mix the AgNCs dispersion obtained in Step 1 (volume ratio 0.8:0.8:3), the DOX solution (concentration 0.8 mg / mL), and the photosensitive prepolymer obtained in Step 2 in a clean container. Stir thoroughly with a magnetic stirrer until the three are evenly mixed. Then add 0.4% LAP (total mass of AgNCs dispersion, DOX solution, and photosensitive prepolymer) and stir evenly. Pour the mixture into a mold and irradiate with an LED light source at a wavelength of 405 nm for 25 seconds to carry out a photocrosslinking reaction, achieving preliminary molding and obtaining a preliminary hydrogel. Step 4: Remove the pre-formed hydrogel obtained in Step 3 from the mold and immerse it in a 1.5% sodium citrate solution for 15 hours to carry out an ionic crosslinking reaction, and finally obtain the composite hydrogel.

[0035] Example 3 Step 1: Mix 0.08 mM AgNO3 solution and 1.8 mM gallic acid solution in a clean beaker at a volume ratio of 1:4. Stir thoroughly with a magnetic stirrer until the mixture is homogeneous. Then, slowly add sodium hydroxide solution dropwise while monitoring the pH value of the solution in real time with a pH meter. Adjust the pH of the solution to 8.0, then transfer it to a constant temperature water bath at 50°C and react for 2.5 hours. During the reaction, the solution gradually turns wine-red. After the reaction is complete, a wine-red AgNCs dispersion is obtained. The obtained AgNCs dispersion is detected using a UV-Vis spectrophotometer, and its characteristic peak is determined to be at 420 nm. Step 2: Dissolve chitosan and hyaluronic acid with a degree of deacetylation of 87% in a 0.12M acetic acid solution. Stir thoroughly on a magnetic stirrer until completely dissolved to form a homogeneous solution. Then add methacrylic anhydride and continue stirring until homogeneous. Transfer the solution to a 35°C constant temperature water bath and react for 7 hours to obtain the photosensitive prepolymer. The mass ratio of chitosan, hyaluronic acid, acetic acid and methacrylic anhydride is 2.5:1.2:100:6. Step 3: Mix the AgNCs dispersion obtained in Step 1 (volume ratio 1.2:1.2:5), the DOX solution (concentration 1.2 mg / mL), and the photosensitive prepolymer obtained in Step 2 in a clean container. Stir thoroughly with a magnetic stirrer until the three are evenly mixed. Then add the AgNCs dispersion, DOX solution, and 0.6% LAP (total mass of the photosensitive prepolymer) and stir evenly. Pour the mixture into a mold and irradiate with an LED light source with a wavelength of 400 nm for 35 seconds to carry out the photocrosslinking reaction and achieve preliminary molding, resulting in a preliminary hydrogel. Step 4: Remove the pre-formed hydrogel obtained in Step 3 from the mold and immerse it in a 2.5% sodium citrate solution to carry out an ionic crosslinking reaction for 10 hours, finally obtaining a composite hydrogel.

[0036] Example 4 Step 1: Mix 0.12 mM AgNO3 solution and 2.2 mM gallic acid solution in a clean beaker at a volume ratio of 1:2.5. Stir thoroughly with a magnetic stirrer until the mixture is homogeneous. Then, slowly add sodium hydroxide solution dropwise while monitoring the pH value of the solution in real time with a pH meter. Adjust the pH of the solution to 8.2, then transfer it to a constant temperature water bath at 52°C and react for 2.2 hours. During the reaction, the solution gradually turns wine-red. After the reaction is complete, a wine-red AgNCs dispersion is obtained. The AgNCs dispersion is detected using a UV-Vis spectrophotometer, and its characteristic peak is determined to be at 422 nm. Step 2: Dissolve chitosan and hyaluronic acid with a degree of deacetylation of 85% in a 0.08M acetic acid solution. Stir thoroughly on a magnetic stirrer until completely dissolved to form a homogeneous solution. Then add methacrylic anhydride and continue stirring until homogeneous. Transfer the solution to a 36°C constant temperature water bath and react for 6 hours to obtain the photosensitive prepolymer. The mass ratio of chitosan, hyaluronic acid, acetic acid and methacrylic anhydride is 1.8:0.9:100:4.5. Step 3: Mix the AgNCs dispersion obtained in Step 1 (volume ratio 1:1:3.5), the DOX solution (concentration 0.9 mg / mL), and the photosensitive prepolymer obtained in Step 2 in a clean container. Stir thoroughly with a magnetic stirrer until the three are evenly mixed. Then add 0.45% LAP of the total mass of AgNCs dispersion, DOX solution, and photosensitive prepolymer and stir evenly. Pour the mixture into a mold and irradiate with an LED light source with a wavelength of 400 nm for 28 seconds to carry out the photocrosslinking reaction and achieve preliminary molding, obtaining a preliminary hydrogel. Step 4: Remove the pre-formed hydrogel obtained in Step 3 from the mold and immerse it in a 1.8% sodium citrate solution to carry out an ionic crosslinking reaction for 13 hours, finally obtaining a composite hydrogel.

[0037] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention in any way. Any simple modifications, alterations, and equivalent changes made to the above embodiments based on the technical essence of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A method for preparing a silver-loaded nanocluster-doxacin composite hydrogel, characterized in that, The method includes the following steps: Step 1: Stir AgNO3 solution and gallic acid solution evenly, then add sodium hydroxide solution dropwise until the pH is 8-9, and then treat with water bath to obtain silver nanocluster dispersion; Step 2: Mix chitosan, hyaluronic acid, acetic acid solution and methacrylic anhydride to react and obtain photosensitive prepolymer; Step 3: Mix the silver nanocluster dispersion obtained in Step 1, the doxorubicin hydrochloride solution, and the photosensitive prepolymer obtained in Step 2 evenly, and then add phenyl-2,4,6-trimethylbenzoyl lithium phosphine as a photoinitiator to carry out a photocrosslinking reaction to obtain a preliminary hydrogel. Step 4: Immerse the preliminarily formed hydrogel obtained in Step 3 into sodium citrate solution to carry out ionic cross-linking reaction, and obtain silver-loaded nanoclusters-doxacin composite hydrogel.

2. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, In step one, the concentration of the AgNO3 solution is 0.08 mM to 0.12 mM, the concentration of the gallic acid solution is 1.8 mM to 2.2 mM, and the volume ratio of the AgNO3 solution to the gallic acid solution is 1:2 to 4.

3. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, The water bath treatment in step one is carried out at a temperature of 50℃~60℃ for 1.5h~2.5h.

4. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, The characteristic peak of the silver nanocluster dispersion in step one is 420 nm to 430 nm.

5. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, In step two, the mass ratio of chitosan, hyaluronic acid, acetic acid, and methacrylic anhydride is 1.8~2.5:0.8~1.2:100:4.5~6, the degree of deacetylation of the chitosan is 83%~87%, and the concentration of the acetic acid solution is 0.08M~0.12M.

6. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, The mixing reaction in step two is carried out at a temperature of 35℃~40℃ for 5h~7h.

7. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, In step three, the volume ratio of the silver nanocluster dispersion, doxorubicin hydrochloride solution, and photosensitive prepolymer is 0.8~1.2:0.8~1.2:3~5; the concentration of the doxorubicin hydrochloride solution is 0.8 mg / mL~1.2 mg / mL; and the added mass of lithium phenyl-2,4,6-trimethylbenzoylphosphinic acid is 0.4%~0.6% of the total mass of the silver nanocluster dispersion, doxorubicin hydrochloride solution, and photosensitive prepolymer.

8. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, The wavelength of the light source used in the photocrosslinking reaction in step three is 400nm~410nm, and the irradiation time of the light source in the photocrosslinking reaction is 25s~35s.

9. The method for preparing a silver-loaded nanocluster-doxacin composite hydrogel according to claim 1, characterized in that, The sodium citrate solution in step four has a mass fraction of 1.5% to 2.5%, and the ion cross-linking reaction takes 10 to 15 hours.

10. A silver-loaded nanocluster-doxacin composite hydrogel, characterized in that, The composite hydrogel is prepared by the method described in any one of claims 1 to 9.

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