Sodium alginate-based composite hydrogel as well as preparation method and application thereof

Abstract

The invention provides sodium alginate-based composite hydrogel, a preparation method thereof and application of the sodium alginate-based composite hydrogel in wound dressing. The preparation method provided by the invention comprises the following steps: preparing nano-silver by adopting a chemical reduction method, preparing chitosan-nano-silver gel beads, and further embedding gallic acid and the chitosan-nano-silver gel beads in sodium alginate / gelatin hydrogel together to obtain the sodium alginate / gelatin / gallic acid / chitosan-nano-silver gel bead hydrogel. The mixture is the sodium alginate-based composite hydrogel disclosed by the invention. The chitosan-nano-silver gel beads and gallic acid play a synergistic antibacterial role, so that escherichia coli and staphylococcus aureus can be effectively inhibited; meanwhile, gallic acid has remarkable antioxidant activity and can remove excessive active oxygen accumulated on the wound so as to promote wound healing. The composite hydrogel is simple to prepare and high in biocompatibility, and can be widely applied to the field of biological medicines.

Description

Technical Field

[0001] This invention belongs to the fields of materials synthesis and biomedicine, specifically relating to a sodium alginate-based composite hydrogel with highly efficient antibacterial and antioxidant properties, its preparation method, and its application. Background Technology

[0002] Traditional dressings such as gauze and cotton have poor antibacterial and antioxidant activity, cannot provide a moist environment for the wound, and have low absorption capacity, leading to frequent and cumbersome dressing changes. In addition, these dressings tend to stick to the wound, often causing pain and discomfort to the patient when removed.

[0003] Unlike traditional dressings, hydrogel dressings provide a moist environment for the wound, accelerating granulation tissue formation and promoting wound healing. They are easy to remove from the wound, reducing pain during dressing changes. More importantly, antibacterial and antioxidant agents can be easily loaded or encapsulated within the hydrogel, thus endowing the dressing with excellent antibacterial and antioxidant activity.

[0004] Sodium alginate, derived from brown algae, is a highly biocompatible polysaccharide widely used in the biomedical field. Because its carboxyl groups can undergo cross-linking in the presence of divalent cations (such as calcium ions), calcium ions are often used as cross-linking agents to prepare sodium alginate hydrogels. However, sodium alginate hydrogels formed by calcium ion cross-linking are typically quite rigid, making them unsuitable for flexible hydrogel dressings.

[0005] Gelatin, as a hydrolyzed product of collagen, is an important material in the biomedical field. However, its poor mechanical strength limits its application in forming hydrogels on its own. Studies have shown that compounding gelatin with other polymers can significantly improve its mechanical properties. Therefore, introducing gelatin into sodium alginate hydrogel systems can improve both the flexibility of sodium alginate hydrogels and the mechanical properties of gelatin, making composite hydrogels an ideal wound dressing material.

[0006] It is essential to provide wound dressing materials with highly effective antibacterial and antioxidant properties based on sodium alginate / gelatin hydrogel. Summary of the Invention

[0007] The purpose of this invention is to provide a sodium alginate-based composite hydrogel, its preparation method, and its application.

[0008] This invention provides a sodium alginate-based composite hydrogel comprising the following raw material components in parts by weight: nano silver, chitosan, sodium alginate, gelatin, and gallic acid.

[0009] The present invention also provides a method for preparing the above-mentioned sodium alginate-based composite hydrogel, the method comprising: a. Prepare a nano-silver solution; b. Chitosan is dissolved in an aqueous acetic acid solution and mixed evenly at room temperature to obtain a chitosan-acetic acid aqueous solution; a nano silver solution is added to the chitosan-acetic acid aqueous solution and mixed evenly to obtain a chitosan-nano silver mixed solution; then the chitosan-nano silver mixed solution is dropped into a sodium hydroxide solution, and the resulting product is immersed in the sodium hydroxide solution for incubation to perform gelation treatment, and then washed to obtain chitosan-nano silver gel beads; c. Dissolve sodium alginate, gelatin, and gallic acid together in deionized water, heat in a water bath and mix evenly to obtain a sodium alginate / gelatin / gallic acid mixed solution; mix the chitosan-silver nanogel beads with the sodium alginate / gelatin / gallic acid mixed solution, and then add calcium chloride solution for immersion to perform calcium ion crosslinking; wash the obtained product after crosslinking to obtain sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel, that is, the sodium alginate-based composite hydrogel.

[0010] Furthermore, the preparation method specifically includes the following steps: a. Disperse silver nitrate in deionized water, stir until dissolved, and transfer to a flask. Place the flask in an oil bath and heat while maintaining magnetic stirring. After the silver nitrate solution boils, add trisodium citrate solution dropwise to the stirred silver nitrate solution. After it has been completely added, continue heating until a distinct pale yellow color appears. Stop heating and allow it to cool to obtain a nano silver solution. b. Prepare an aqueous acetic acid solution using acetic acid and deionized water. Dissolve chitosan in the aqueous acetic acid solution and magnetically stir for 12 hours at room temperature to obtain a chitosan-acetic acid aqueous solution. Add nano-silver solution dropwise to the chitosan-acetic acid aqueous solution and stir to mix evenly to obtain a chitosan-nano-silver mixed solution. Then, dropwise add the chitosan-nano-silver mixed solution into a sodium hydroxide solution and immerse the resulting product in the sodium hydroxide solution for 12 hours to complete the gelation process. After gelation, wash with deionized water 10-15 times to remove residual sodium hydroxide solution from the surface. After washing, chitosan-nano-silver gel beads are obtained. c. Dissolve sodium alginate, gelatin, and gallic acid together in deionized water and heat in a water bath while magnetically stirring during heating. After the solution is stirred evenly, a sodium alginate / gelatin / gallic acid mixed solution is obtained. Add the chitosan-silver nanogel beads and the sodium alginate / gelatin / gallic acid mixed solution together into a mold, add calcium chloride solution, and immerse for 3 hours to perform calcium ion crosslinking. After crosslinking, remove the hydrogel from the mold and wash it with deionized water to remove excess calcium chloride solution. After rinsing, the sodium alginate-based composite hydrogel is obtained.

[0011] Further, step a is carried out according to the following dosage ratios and operating parameters: the mass of silver nitrate is 0.0325~0.0525g; the volume of deionized water is 100~150mL; the concentration of trisodium citrate solution is 0.025~0.045M, and the volume is 3~7mL; the stirring speed is 800~1200rpm; the temperature of the oil bath is 110~130℃; and the concentration of the nano-silver solution is 0.00186~0.00197M.

[0012] Further, step b is carried out according to the following dosage ratios and operating parameters: the volume of acetic acid is 0.1~0.3mL, the volume of deionized water is 10~17mL, the mass of chitosan is 0.5~0.9g, the stirring speed is 1000~1400rpm, the concentration of the nano silver solution is 0.00186~0.00197M and the volume is 2~4mL, and the concentration of the sodium hydroxide solution is 1~3M and the volume is 40~60mL.

[0013] Further, step c is carried out according to the following dosage ratios and operating parameters: sodium alginate is 0.1~0.3g, gelatin is 0.5~1.0g, gallic acid is 0.05~0.2g, deionized water is 6~10mL, stirring speed is 900~1100rpm, water bath temperature is 60~80℃, chitosan-silver nanogel beads are 3~7g, sodium alginate / gelatin / gallic acid mixed solution is 6~9mL, calcium chloride solution concentration is 0.12~0.24M, and volume is 3~7mL.

[0014] The present invention also provides the application of the above-mentioned sodium alginate-based composite hydrogel in the field of wound dressings.

[0015] Furthermore, the sodium alginate-based composite hydrogel described above has antioxidant capacity, in vitro antibacterial effect, and can be used as an excipient for in vivo wound healing.

[0016] Specifically, this invention provides a method for preparing sodium alginate-based composite hydrogels and testing their high-efficiency antibacterial and antioxidant properties, comprising the following steps: a. Preparation of nano-silver solution: Disperse a certain amount of silver nitrate in deionized water, stir until dissolved, and then transfer to a flask. Place the flask in an oil bath and heat while maintaining magnetic stirring. After the silver nitrate solution boils, add a certain amount of trisodium citrate solution dropwise to the stirred silver nitrate solution. After it has been completely added, continue heating until a distinct pale yellow color appears, then stop heating and allow it to cool to obtain the nano-silver solution. b. Preparation of chitosan-silver nanogel beads: Prepare a certain amount of acetic acid aqueous solution, weigh a certain amount of chitosan and dissolve it in the acetic acid aqueous solution, and magnetically stir the mixed solution at room temperature for 12 hours; take a certain amount of nanosilver solution and add it dropwise to the chitosan-acetic acid aqueous solution, and stir to mix it evenly; then add the chitosan-silver nanogel solution dropwise to a certain concentration of sodium hydroxide solution to form chitosan-silver nanogel beads; immerse the gel beads in sodium hydroxide solution and incubate for 12 hours to complete the gelation process; after gelation, wash the chitosan-silver nanogel beads with deionized water 10-15 times to remove the residual sodium hydroxide solution on the surface, and obtain chitosan-silver nanogel beads after washing; c. Preparation of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel: A certain amount of sodium alginate, a certain amount of gelatin, and a certain amount of gallic acid are dissolved together in a certain amount of deionized water and heated in a water bath while being magnetically stirred during the heating process; after the solution is stirred evenly, a certain amount of chitosan-nano silver gel beads and the sodium alginate / gelatin / gallic acid mixed solution are added to a mold, and a certain amount of calcium chloride solution is added, and the mixture is immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel is removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel is obtained. d. Free radical scavenging and antioxidant capacity test of sodium alginate / gelatin / gallic acid / chitosan-silver nanogel beads hydrogel: To evaluate the antioxidant capacity of the material, the antioxidant capacity of 1,1-diphenyl-2-trinitrophenylhydrazine free radical and 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radical was tested. For the determination of 1,1-diphenyl-2-trinitrophenylhydrazine free radical, 3 mL of 0.1 mM 1,1-diphenyl-2-trinitrophenylhydrazine ethanol solution was taken, 50 mg of freeze-dried hydrogel was added, and after standing in the dark at room temperature for 2 hours, the absorption peak intensity of each group of 1,1-diphenyl-2-trinitrophenylhydrazine free radical at 517 nm was measured using a UV-Vis spectrophotometer. For 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt, the antioxidant capacity of sodium alginate / gelatin / gallic acid / chitosan-silver nanogel beads hydrogel was tested. The determination of 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radicals involved mixing 7.0 mM 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt with 2.5 mM potassium persulfate at a 1:1 volume ratio and reacting in the dark for 24 hours. The mixture was then diluted 20 times to obtain a 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radical solution. 3 mL of this solution was added to 50 mg of freeze-dried hydrogel. After standing in the dark at room temperature for 0.5 hours, the absorption peak intensity of each group of 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radicals at 734 nm was measured using a UV-Vis spectrophotometer. e. Application of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel hydrogel in in vitro antibacterial activity testing of different bacterial species: To evaluate the in vitro antibacterial effect of the material, Escherichia coli grown in the logarithmic phase was diluted with sterile water to 1×10⁻⁶. 8 CFU / mL, add 1 mg of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel hydrogel to a 5 mL centrifuge tube and heat to 37°C. o C, then add 1 mL of E. coli bacterial suspension and mix well; then dilute 100 μL of bacterial suspension to 1 × 10⁻⁶. 5 CFU / mL was plated on agar plates, and colony counts were performed after 24 hours; the same antimicrobial evaluation was performed against Staphylococcus aureus. f. In vivo wound healing experiment of sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel beads hydrogel in rats: Six-week-old male rats, pre-cultured for 7 days, were used to study wound healing in vivo. The rats were anesthetized with 10% chloral hydrate, and a circular wound with a diameter of 1.0 cm was constructed on the back of the rats to complete the rat wound model. The rats were randomly divided into two groups: control group and sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel beads hydrogel group. Photos were taken at 0, 3, 5, 7, 10, and 14 days after surgery to observe the growth and calculate the wound area. Compared with the prior art, the present invention has at least the following beneficial effects: The sodium alginate-based composite hydrogel provided by this invention can achieve highly efficient sterilization through the synergistic antibacterial effect of gallic acid and chitosan-silver nanogel beads. Simultaneously, the pyrogallol groups of gallic acid have excellent antioxidant effects, which can scavenge free radicals in wounds to promote wound healing. This hydrogel is simple to prepare, highly biocompatible, and can be widely used in the biomedical field. Attached Figure Description

[0017] One or more embodiments are illustrated by way of example with reference to the accompanying drawings. These illustrations do not constitute a limitation on the embodiments, and unless otherwise stated, the figures in the drawings are not to be limited by scale.

[0018] Figure 1 Scanning electron microscope (SEM) images of sodium alginate / gelatin / gallic acid / chitosan-silver nanogel beads hydrogel in Example 1 and sodium alginate / gelatin / gallic acid hydrogel in Comparative Example 1. Figure 2 Infrared spectra of gelatin, sodium alginate, chitosan-silver nanogel beads, gallic acid, and sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel in Example 1; Figure 3 The bar chart shows the scavenging rates of 1,1-diphenyl-2-trinitrophenylhydrazine free radical and 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radicals of the sodium alginate / gelatin / gallic acid / chitosan-nano silver gel beads hydrogel in Example 1 and the sodium alginate / gelatin hydrogel in Comparative Example 2. Figure 4 The images show the antibacterial effects of sodium alginate / gelatin / gallic acid / chitosan-silver nanogel beads hydrogel in Example 1, chitosan-silver nanogel beads in Comparative Example 3, and sodium alginate / gelatin / gallic acid hydrogel in Comparative Example 4 on Escherichia coli and Staphylococcus aureus. Figure 5 The image shows the in vivo wound healing experiment in rats using the sodium alginate / gelatin / gallic acid / chitosan-nano silver gel hydrogel in Example 1 and the blank control group. Detailed Implementation

[0019] Gallic acid is a highly bioactive natural polyphenol that can be extracted from natural plants such as gallnut, oak bark, and witch hazel. It also possesses antibacterial, antioxidant, and anti-inflammatory properties. As a highly effective free radical scavenger, gallic acid has been incorporated into hydrogel dressings to eliminate reactive oxygen species and promote wound healing.

[0020] The inventors discovered that while nanosilver is widely used as an antibacterial agent due to its broad-spectrum antibacterial activity, excessive amounts can lead to adverse reactions such as gastrointestinal disturbances and spasms. Fortunately, combining it with chitosan can significantly reduce the potential toxicity of nanosilver. Furthermore, chitosan, as a cationic natural polysaccharide, possesses excellent biocompatibility, biodegradability, and antibacterial activity. Under neutral / acidic conditions, the amino groups of chitosan can undergo protonation, interacting with negatively charged molecules on the cell membrane, thereby leading to microbial death.

[0021] Based on this, the present invention embeds gallic acid and chitosan-silver nanogel beads into sodium alginate / gelatin hydrogel to prepare a sodium alginate-based natural polysaccharide hydrogel for promoting wound healing. The carboxyl groups on the sodium alginate molecular chain undergo ionic cross-linking with calcium ions to form a hydrogel, while gallic acid and chitosan-silver nanogel beads are embedded in the hydrogel dressing. Due to the synergistic antibacterial effect of gallic acid and chitosan-silver nanogel beads, the dressing possesses significant antibacterial activity; simultaneously, the presence of gallic acid gives it good antioxidant activity. Therefore, a sodium alginate-based composite hydrogel is provided that can be used to promote wound healing.

[0022] The present invention will now be described in detail with reference to specific embodiments.

[0023] Example 1: A method for preparing a sodium alginate-based composite hydrogel includes the following steps: (1) Preparation of nano-silver solution: Weigh 0.0425g of silver nitrate and disperse it in 125mL of deionized water. Stir until dissolved and transfer to a flask. Place the flask in an oil bath at 120℃ and heat it while maintaining magnetic stirring at 1000rpm. After the silver nitrate solution boils, use a dropper to add 5mL of 0.034M trisodium citrate solution dropwise to the stirred silver nitrate solution. After it is completely added, continue heating until a distinct pale yellow color appears. Stop heating and cool to obtain a nano-silver solution with a concentration of 0.00192M.

[0024] (2) Preparation of chitosan-silver nanogel beads: Use a dropper to add 0.2 mL of acetic acid to a beaker containing 15 mL of deionized water. Weigh 0.7 g of chitosan and dissolve it in the acetic acid aqueous solution. Stir the mixed solution magnetically at 1200 rpm for 12 hours at room temperature. Add 3 mL of 0.00192 M silver nanosol to the chitosan-acetic acid aqueous solution dropwise and stir to mix evenly. Then add the chitosan-silver nanosol mixed solution to 50 mL of 2 M sodium hydroxide solution to form chitosan-silver nanogel beads. Immerse the gel beads in the sodium hydroxide solution and incubate for 12 hours to complete the gelation process. After gelation, wash the chitosan-silver nanogel beads with deionized water 10-15 times to remove the residual sodium hydroxide solution on the surface. After washing, chitosan-silver nanogel beads are obtained.

[0025] (3) Preparation of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel: 0.2g sodium alginate, 0.8g gelatin and 0.1g gallic acid were dissolved together in 9mL deionized water and heated in a water bath at 70℃ while magnetically stirring at 1000rpm during the heating process; after the solution was stirred evenly, 5g chitosan-nano silver gel beads and 7mL sodium alginate / gelatin / gallic acid mixed solution were added to the mold, and 5mL of 0.18M calcium chloride solution was added and immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel was obtained.

[0026] (4) For the determination of 1,1-diphenyl-2-trinitrophenylhydrazine radical, 3 mL of 0.1 mM ethanol solution of 1,1-diphenyl-2-trinitrophenylhydrazine was taken, and 50 mg of freeze-dried hydrogel was added. After standing in the dark at room temperature for 2 hours, the absorption peak intensity of each group of 1,1-diphenyl-2-trinitrophenylhydrazine radicals at 517 nm was measured using a UV-Vis spectrophotometer. For the determination of 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt radical, 7.0 mM 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt was added to a solution of 50 mg of 0.1 mM ethanol solution of 1,1-diphenyl-2-trinitrophenylhydrazine was measured. Potassium persulfate of 2.5 mM was mixed at a 1:1 volume ratio and reacted in the dark for 24 hours. The mixture was then diluted 20 times to obtain a 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radical solution. 3 mL of the 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radical solution was taken, and 50 mg of freeze-dried hydrogel was added. After standing in the dark at room temperature for 0.5 hours, the absorption peak intensity of each group of 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt free radicals at 734 nm was measured using a UV-Vis spectrophotometer.

[0027] (5) To evaluate the in vitro antibacterial effect of the material, Escherichia coli grown in the logarithmic phase was diluted with sterile water to 1×10⁻⁶. 8 CFU / mL, add 1 mg of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel hydrogel to a 5 mL centrifuge tube and heat to 37°C. o C, then add 1 mL of E. coli bacterial suspension and mix well; then dilute 100 μL of bacterial suspension to 1 × 10⁻⁶. 5 CFU / mL was plated on an agar plate, and the colony count was performed after 24 hours; the same antimicrobial evaluation was performed on Staphylococcus aureus.

[0028] (6) To evaluate the effect of the material on wound healing in rats, sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel beads hydrogel were applied to rats for in vivo healing experiments. Six-week-old male rats (200-220g) that had been pre-fed for 7 days were used for in vivo wound healing studies. The rats were anesthetized with 10% chloral hydrate, and a circular wound with a diameter of 1.0cm was constructed on the back of the rats to complete the rat wound model. The rats were randomly divided into two groups: a control group and a sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel beads hydrogel group. Photos were taken on days 0, 3, 5, 7, 10, and 14 after surgery to observe their growth and calculate the wound area.

[0029] Reference Figure 1The field emission scanning electron microscope (FESEM) image of the sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel prepared in Example 1 is as follows: Figure 1 A. As can be clearly seen from the figure, chitosan-silver nanogel beads were successfully embedded in sodium alginate / gelatin / gallic acid hydrogel. At the same time, it can be visually seen that the hydrogel has a porous three-dimensional network structure.

[0030] The infrared spectra of the gelatin, sodium alginate, chitosan-silver nanogel beads, gallic acid, and sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogels prepared in Example 1 are shown below. Figure 2 As shown, the gelatin is at 3435cm. –1 The peak is attributed to the stretching vibration of N–H, while the 1546 cm peak is attributed to the stretching vibration of N–H. –1 The peak is attributed to the bending vibration of N–H in the amide II band. Sodium alginate at 3408 cm⁻¹ –1 The peak belongs to the O–H stretching vibration, 1623 cm⁻¹. –1 The peak and 1417cm –1 The peaks are respectively attributed to –COO – Asymmetric stretching vibration and –COO – Symmetric stretching vibrations. For chitosan-silver nanogel beads, 3446 cm⁻¹ –1 The characteristic peak at 2929 cm⁻¹ is attributed to the stretching vibrations of N–H and O–H. –1 and 2857cm –1 The characteristic peaks at 1642 cm⁻¹ belong to the asymmetric and symmetric stretching vibrations of C–H, respectively. –1 The peak belongs to the bending vibration of N–H. Gallic acid at 3499 cm⁻¹ –1 and 3283cm –1 The characteristic peak at 1028 cm⁻¹ belongs to the stretching vibration of O–H. –1 The peak belongs to the C–H bending vibration. For sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel, 3446 cm⁻¹ –1 Belonging to the N–H stretching vibration and O–H stretching vibration, 2929 cm –1 and 2857cm –1 Asymmetric and symmetric stretching vibrations of C–H from chitosan-silver nanogel beads, 1664 cm⁻¹ –1 and 1542cm –1 The characteristic peaks correspond to the –COO of sodium alginate, respectively. – The asymmetric stretching vibrations and the bending vibrations of N–H in the amide II band of gelatin, 1028 cm⁻¹ –1The characteristic peaks correspond to the C–H bending vibrations of gallic acid. These results demonstrate the successful synthesis of the target product, sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel.

[0031] The free radical scavenging and antioxidant capacity of the sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel prepared in Example 1 is as follows: Figure 3 As shown, the antioxidant capacity of the sodium alginate / gelatin / gallic acid / chitosan-silver nanogel hydrogel was tested using 1,1-diphenyl-2-trinitrophenylhydrazine radical and 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt radical. The scavenging rates of 1,1-diphenyl-2-trinitrophenylhydrazine radical and 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt radical reached 91.81% and 99.43%, respectively, indicating its highly efficient free radical scavenging ability. This is because the pyrogallol group in the gallic acid molecule effectively scavenge free radicals.

[0032] The sodium alginate / gelatin / gallic acid / chitosan-silver nanogel hydrogel prepared in Example 1 exhibits the following antibacterial activity against Escherichia coli and Staphylococcus aureus: Figure 4 As shown, the sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel exhibited inhibition rates of 99.93% and 99.99% against *Escherichia coli* and *Staphylococcus aureus*, respectively. The results demonstrate that this hydrogel effectively inhibits bacteria. This is attributed to the synergistic antibacterial effect of gallic acid and chitosan-silver nanogel beads, which endows the hydrogel with highly efficient antibacterial properties. The silver nanoparticles in the chitosan-silver nanogel beads can penetrate the interior of bacteria, disrupting their internal structure and biomolecules. Simultaneously, gallic acid has an affinity for the lipophilic membrane layer of bacterial cell membranes. This affinity alters the cell's electrochemical potential, leading to a decrease in cell membrane integrity and promoting bacterial death. It is precisely this synergistic antibacterial effect of gallic acid and chitosan-silver nanogel beads that gives this hydrogel its powerful antibacterial efficacy.

[0033] The in vivo wound healing experiment in rats using the sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel prepared in Example 1 is as follows: Figure 5 As shown. Figure 5 Photographs A show the wound repair effects on rats in the control group and the sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel bead hydrogel group. As can be seen from the figures, at the same time point, the wound contraction in the sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel bead hydrogel group was significantly greater than that in the control group, indicating that the sodium alginate / gelatin / gallic acid / chitosan-nanosilver gel bead hydrogel can effectively promote wound healing. Furthermore, compared to the control group, almost complete wound healing was achieved after two weeks of using this hydrogel as a dressing. Figure 5 B is a graph showing the percentage of unhealed wound area. As can be seen from the graph, the area of ​​unhealed wound in the sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel group is much smaller than that in the blank control group, further confirming the therapeutic potential of this hydrogel in the field of wound repair.

[0034] Example 2: A method for preparing a sodium alginate-based composite hydrogel includes the following steps: (1) Preparation of nano-silver solution: Weigh 0.0325g of silver nitrate and disperse it in 100mL of deionized water. Stir until dissolved and transfer to a flask. Place the flask in an oil bath at 110℃ and heat it while maintaining magnetic stirring at 800rpm. After the silver nitrate solution boils, use a dropper to add 3mL of 0.025M trisodium citrate solution dropwise to the stirred silver nitrate solution. After it is completely added, continue heating until a distinct pale yellow color appears. Stop heating and cool to obtain a nano-silver solution with a concentration of 0.00186M.

[0035] (2) Preparation of chitosan-silver nanogel beads: Use a dropper to add 0.1 mL of acetic acid to a beaker containing 10 mL of deionized water. Weigh 0.5 g of chitosan and dissolve it in the acetic acid aqueous solution. Stir the mixed solution magnetically at 1000 rpm for 12 hours at room temperature. Add 2 mL of 0.00186 M silver nanosol dropwise to the chitosan-acetic acid aqueous solution and stir to mix evenly. Then add the chitosan-silver nanosol mixed solution to 40 mL of 1 M sodium hydroxide solution to form chitosan-silver nanogel beads. Immerse the gel beads in the sodium hydroxide solution and incubate for 12 hours to complete the gelation process. After gelation, wash the chitosan-silver nanogel beads with deionized water 10-15 times to remove the residual sodium hydroxide solution on the surface. After washing, chitosan-silver nanogel beads are obtained.

[0036] (3) Preparation of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel: 0.1g sodium alginate, 0.5g gelatin and 0.05g gallic acid were dissolved together in 6mL deionized water and heated in a water bath at 60℃ while magnetically stirring at 900rpm during the heating process; after the solution was stirred evenly, 3g chitosan-nano silver gel beads and 6mL sodium alginate / gelatin / gallic acid mixed solution were added to the mold, and 3mL of 0.12M calcium chloride solution was added and immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel was obtained.

[0037] Example 3: A method for preparing a sodium alginate-based composite hydrogel includes the following steps: (1) Preparation of nano-silver solution: Weigh 0.0525g of silver nitrate and disperse it in 150mL of deionized water. Stir until dissolved and transfer to a flask. Place the flask in an oil bath at 130℃ and heat it while maintaining magnetic stirring at 1200rpm. After the silver nitrate solution boils, use a dropper to add 7mL of 0.045M trisodium citrate solution dropwise to the stirred silver nitrate solution. After it is completely added, continue heating until a distinct pale yellow color appears. Stop heating and cool to obtain a nano-silver solution with a concentration of 0.00197M.

[0038] (2) Preparation of chitosan-silver nanogel beads: 0.3 mL of acetic acid was dropped into a beaker containing 17 mL of deionized water using a dropper. 0.9 g of chitosan was weighed and dissolved in the acetic acid aqueous solution. The mixed solution was magnetically stirred at 1400 rpm for 12 hours at room temperature. 4 mL of 0.00197 M silver nanosol was added dropwise to the chitosan-acetic acid aqueous solution and stirred to mix evenly. Then, the chitosan-silver nanosol mixed solution was dropped into 60 mL of 3 M sodium hydroxide solution to form chitosan-silver nanogel beads. The gel beads were immersed in the sodium hydroxide solution and incubated for 12 hours to complete the gelation process. After gelation, the chitosan-silver nanogel beads were washed with deionized water 10-15 times to remove the residual sodium hydroxide solution on the surface. After washing, chitosan-silver nanogel beads were obtained.

[0039] (3) Preparation of sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel: 0.3g sodium alginate, 1.0g gelatin and 0.2g gallic acid were dissolved together in 10mL deionized water and heated in a water bath at 80℃ while magnetically stirring at 1100rpm during the heating process; after the solution was stirred evenly, 7g chitosan-nano silver gel beads and 9mL sodium alginate / gelatin / gallic acid mixed solution were added to the mold, and 7mL of 0.24M calcium chloride solution was added and immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid / chitosan-nano silver gel bead hydrogel was obtained.

[0040] Comparative Example 1: (1) Preparation of sodium alginate / gelatin / gallic acid hydrogel: 0.2g sodium alginate, 0.8g gelatin and 0.1g gallic acid were dissolved together in 9mL of deionized water and heated in a water bath at 70℃ while magnetically stirring at 1000rpm during the heating process; after the solution was stirred evenly, 7mL of sodium alginate / gelatin / gallic acid mixed solution was added to the mold, and 5mL of 0.18M calcium chloride solution was added and immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid hydrogel was obtained.

[0041] Continue to refer to Figure 1 The field emission scanning electron microscope image of the sodium alginate / gelatin / gallic acid hydrogel prepared in Comparative Example 1 is as follows: Figure 1 B. As can be seen from the figure, the sodium alginate / gelatin / gallic acid hydrogel exhibits a porous three-dimensional network structure.

[0042] Comparative Example 2: (1) Preparation of sodium alginate / gelatin hydrogel: 0.2 g of sodium alginate and 0.8 g of gelatin were dissolved together in 9 mL of deionized water and heated in a water bath at 70 °C while magnetically stirring at 1000 rpm. After the solution was stirred evenly, 7 mL of sodium alginate / gelatin mixed solution was added to a mold, and 5 mL of 0.18 M calcium chloride solution was added. The mixture was immersed for 3 hours to allow calcium ion crosslinking. After crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin hydrogel was obtained.

[0043] The sodium alginate / gelatin hydrogel prepared in Comparative Example 2 exhibited the following antioxidant capacity for scavenging free radicals: Figure 3 As shown, the sodium alginate / gelatin hydrogel exhibited scavenging rates of 12.07% and 14.86% for 1,1-diphenyl-2-trinitrophenylhydrazine radical and 2,2'-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt radical, respectively, and its antioxidant activity was related to the hydrophobic amino acids in gelatin.

[0044] Comparative Example 3: (1) Preparation of nano-silver solution: Weigh 0.0425g of silver nitrate and disperse it in 125mL of deionized water. Stir until dissolved and transfer to a flask. Place the flask in an oil bath at 120℃ and heat it while maintaining magnetic stirring at 1000rpm. After the silver nitrate solution boils, use a dropper to add 5mL of 0.034M trisodium citrate solution dropwise to the stirred silver nitrate solution. After it is completely added, continue heating until a distinct pale yellow color appears. Stop heating and cool to obtain a nano-silver solution with a concentration of 0.00192M. (2) Preparation of chitosan-silver nanogel beads: Use a dropper to add 0.2 mL of acetic acid to a beaker containing 15 mL of deionized water. Weigh 0.7 g of chitosan and dissolve it in the acetic acid aqueous solution. Stir the mixed solution magnetically at 1200 rpm for 12 hours at room temperature. Add 3 mL of 0.00192 M silver nanosol to the chitosan-acetic acid aqueous solution dropwise and stir to mix evenly. Then add the chitosan-silver nanosol mixed solution to 50 mL of 2 M sodium hydroxide solution to form chitosan-silver nanogel beads. Immerse the gel beads in the sodium hydroxide solution and incubate for 12 hours to complete the gelation process. After gelation, wash the chitosan-silver nanogel beads with deionized water 10-15 times to remove the residual sodium hydroxide solution on the surface. After washing, chitosan-silver nanogel beads are obtained.

[0045] The antibacterial activity of chitosan-silver nanogel beads against Escherichia coli and Staphylococcus aureus in Comparative Example 3 is as follows: Figure 4 As shown. From Figure 4 It can be seen that the chitosan-silver nanogel beads exhibited inhibition rates of 99.10% and 99.07% against Escherichia coli and Staphylococcus aureus, respectively. This is because silver nanoparticles adhere to the cell membrane and affect microbial activity by altering the cell membrane structure and permeability. When silver nanoparticles penetrate into the microbial cell, they interact with the cell structure and biomolecules, thus exerting an inhibitory effect on the microorganisms.

[0046] Comparative Example 4: (1) Preparation of sodium alginate / gelatin / gallic acid hydrogel: 0.2g sodium alginate, 0.8g gelatin and 0.1g gallic acid were dissolved together in 9mL of deionized water and heated in a water bath at 70℃ while magnetically stirring at 1000rpm during the heating process; after the solution was stirred evenly, 7mL of sodium alginate / gelatin / gallic acid mixed solution was added to the mold, and 5mL of 0.18M calcium chloride solution was added and immersed for 3 hours to carry out calcium ion crosslinking; after crosslinking, the hydrogel was removed from the mold and washed with deionized water to remove excess calcium chloride solution. After rinsing, sodium alginate / gelatin / gallic acid hydrogel was obtained.

[0047] The antibacterial activity of sodium alginate / gelatin / gallic acid hydrogel against Escherichia coli and Staphylococcus aureus in Comparative Example 4 is as follows: Figure 4 As shown. From Figure 4 It can be seen that the sodium alginate / gelatin / gallic acid hydrogel exhibited inhibition rates of 98.02% and 92.28% against *Escherichia coli* and *Staphylococcus aureus*, respectively. This is because gallic acid has an affinity for the lipophilic membrane layer of bacterial cell membranes. This affinity causes changes in the cell's electrochemical potential, leading to a decrease in cell membrane integrity and thus promoting bacterial death. Furthermore, compared with Example 1 and Comparative Example 3, the synergistic effect of gallic acid and chitosan-silver nanogel beads demonstrates that Example 1 possesses excellent antibacterial properties and can effectively inhibit bacterial activity.

[0048] Those skilled in the art will understand that the above embodiments are specific examples of implementing the present invention, and in practical applications, various changes in form and detail can be made without departing from the spirit and scope of the present invention. Any person skilled in the art can make their own modifications and alterations without departing from the spirit and scope of the present invention; therefore, the scope of protection of the present invention should be determined by the scope defined in the claims.

Claims

1. A sodium alginate-based composite hydrogel, characterized in that, It includes the following raw material components: nano silver, chitosan, sodium alginate, gelatin, and gallic acid.

2. A method for preparing a sodium alginate-based composite hydrogel according to claim 1, characterized in that, The preparation method includes: a. Prepare a nano-silver solution; b. Chitosan is dissolved in an aqueous acetic acid solution and mixed evenly at room temperature to obtain a chitosan-acetic acid aqueous solution; a nano silver solution is added to the chitosan-acetic acid aqueous solution and mixed evenly to obtain a chitosan-nano silver mixed solution; then the chitosan-nano silver mixed solution is dropped into a sodium hydroxide solution, and the resulting product is immersed in the sodium hydroxide solution for incubation to perform gelation treatment, and then washed to obtain chitosan-nano silver gel beads; c. Dissolve sodium alginate, gelatin, and gallic acid together in deionized water, heat in a water bath and mix evenly to obtain a sodium alginate / gelatin / gallic acid mixed solution; mix the chitosan-silver nanogel beads with the sodium alginate / gelatin / gallic acid mixed solution, and then add calcium chloride solution for immersion to perform calcium ion crosslinking; wash the obtained product after crosslinking to obtain sodium alginate / gelatin / gallic acid / chitosan-silver nanogel bead hydrogel, that is, the sodium alginate-based composite hydrogel.

3. The method for preparing sodium alginate-based composite hydrogel according to claim 2, characterized in that, The preparation method specifically includes the following steps: a. Disperse silver nitrate in deionized water, stir until dissolved, and transfer to a flask. Place the flask in an oil bath and heat while maintaining magnetic stirring. After the silver nitrate solution boils, add trisodium citrate solution dropwise to the stirred silver nitrate solution. After it has been completely added, continue heating until a distinct pale yellow color appears. Stop heating and allow it to cool to obtain a nano silver solution. b. Prepare an aqueous acetic acid solution using acetic acid and deionized water. Dissolve chitosan in the aqueous acetic acid solution and magnetically stir for 12 hours at room temperature to obtain a chitosan-acetic acid aqueous solution. Add nano-silver solution dropwise to the chitosan-acetic acid aqueous solution and stir to mix evenly to obtain a chitosan-nano-silver mixed solution. Then, dropwise add the chitosan-nano-silver mixed solution into a sodium hydroxide solution and immerse the resulting product in the sodium hydroxide solution for 12 hours to complete the gelation process. After gelation, wash with deionized water 10-15 times to remove residual sodium hydroxide solution from the surface. After washing, chitosan-nano-silver gel beads are obtained. c. Dissolve sodium alginate, gelatin, and gallic acid together in deionized water and heat in a water bath while magnetically stirring during heating. After the solution is stirred evenly, a sodium alginate / gelatin / gallic acid mixed solution is obtained. Add the chitosan-silver nanogel beads and the sodium alginate / gelatin / gallic acid mixed solution together into a mold, add calcium chloride solution, and immerse for 3 hours to perform calcium ion crosslinking. After crosslinking, remove the hydrogel from the mold and wash it with deionized water to remove excess calcium chloride solution. After rinsing, the sodium alginate-based composite hydrogel is obtained.

4. The method for preparing sodium alginate-based composite hydrogel according to claim 1, characterized in that, Step a is carried out according to the following dosage ratios and operating parameters: the mass of silver nitrate is 0.0325~0.0525g; the volume of deionized water is 100~150mL; the concentration of trisodium citrate solution is 0.025~0.045M, and the volume is 3~7mL; the stirring speed is 800~1200rpm; the temperature of the oil bath is 110~130℃; and the concentration of the nano-silver solution is 0.00186~0.00197M.

5. The method for preparing sodium alginate-based composite hydrogel according to claim 1, characterized in that, Step b is performed according to the following proportions and operating parameters: the volume of acetic acid is 0.1~0.3mL, the volume of deionized water is 10~17mL, the mass of chitosan is 0.5~0.9g, the stirring speed is 1000~1400rpm, the concentration of the nano silver solution is 0.00186~0.00197M, and the volume is 2~4mL; the concentration of the sodium hydroxide solution is 1~3M, and the volume is 40~60mL.

6. The method for preparing sodium alginate-based composite hydrogel according to claim 1, characterized in that, Step c is performed according to the following dosage ratios and operating parameters: sodium alginate is 0.1~0.3g, gelatin is 0.5~1.0g, gallic acid is 0.05~0.2g, deionized water is 6~10mL, stirring speed is 900~1100rpm, water bath temperature is 60~80℃, chitosan-silver nanogel beads are 3~7g, sodium alginate / gelatin / gallic acid mixed solution is 6~9mL, calcium chloride solution concentration is 0.12~0.24M, and volume is 3~7mL.

7. The application of a sodium alginate-based composite hydrogel according to claim 1 in the field of wound dressings.

8. The application according to claim 7, characterized in that, The sodium alginate-based composite hydrogel described above has antioxidant capabilities.

9. The application according to claim 7, characterized in that, The sodium alginate-based composite hydrogel described above has an in vitro antibacterial effect.

10. The application according to claim 7, characterized in that, The sodium alginate-based composite hydrogel is used as an excipient for in vivo wound healing.

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