Hydrogel with photo-thermal multiple antibacterial and hemostatic functions as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to hydrogel with photo-thermal multiple antibacterial and hemostatic functions. The multifunctional hydrogel is prepared from epsilon-polylysine (EPL), gallic acid (GA), tannic acid (TA) and ferric trichloride hexahydrate. The polyphenols are obtained through multiple mechanisms such as reaction between the polyphenols and amino groups, metal chelation, pi-pi interaction and electrostatic attraction (as shown in Figure 1). The metal ion and multiple dynamic cross-linking system hydrogel is constructed, the hydrogel with positive charges enriched on the surface can act with blood platelets with negative charges, red blood cell aggregation, blood platelet adhesion and vasoconstriction effects are promoted, and the blood coagulation efficiency is improved. The hydrogel shows excellent antibacterial performance and photo-thermal performance, can prevent infection in the inflammation stage of wound healing, can effectively accelerate wound healing, and has good biological safety.

Description

Technical Field

[0001] The present invention relates to a hydrogel with photothermal multiple antibacterial and hemostatic functions, belonging to the field of biomedical technology. By constructing a dynamic crosslinking network system of metal-polyphenol / cationic polymer, the present invention realizes the synergistic improvement of hemostatic efficacy, antibacterial activity and tissue regeneration ability, accelerates the wound hemostasis and healing process and prevents infection, providing a breakthrough solution for the comprehensive management of complex wounds. Background Art

[0002] In contemporary clinical medical practice, wound management, especially the treatment of chronic refractory wounds and postoperative infectious wounds, has always been a highly challenging clinical problem. Traditional wound dressings such as gauze, bandage, and cotton are prone to adhesion to newly formed tissues during the treatment process, resulting in secondary trauma and unable to meet the needs of moist wound healing. Approximately 6.79 million people worldwide are troubled by chronic wounds, and the demand for new medical dressings is becoming increasingly urgent. Hydrogel materials are reticular crosslinked polymers with water as the dispersion medium and containing both hydrophilic groups and hydrophobic residues. Due to their excellent water absorption and storage capacity, biocompatibility, environmental responsiveness, and ease of processing and functional modification, they are widely used in the medical and nursing fields, including wound dressings, tissue repair, drug delivery, and biological lubrication. Compared with traditional wound dressings, new hydrogel dressings can absorb wound secretions, maintain a moist environment, promote wound healing and reduce the risk of infection. In addition, the adhesion and mechanical properties of hydrogels enable them to achieve good adhesion effects and reduce irritation during dressing removal. Although the application of hydrogels as new dressings has made significant progress, many challenges still remain. These mainly include the following three problems: (1) The functional module has an obvious single feature and is difficult to meet the requirements of complex wound microenvironment regulation; (2) The traditional hemostatic mechanism overly relies on physical adsorption of material pore size and lacks the ability to activate the coagulation cascade at the molecular level, resulting in low hemostatic efficiency; (3) The cytotoxicity caused by residual initiators on the material surface requires urgent improvement in biosafety. These problems have hindered the further popularization and application of hydrogel dressings and are difficult to meet more complex medical needs.

[0003] The present invention first functionalizes ε-polylysine (EPL) with gallic acid (GA) to obtain an EPL-GA complex. Subsequently, EPL-GA and Fe 3+The coordination effect forms a metal-polyphenol network. On this basis, a multiple dynamic cross-linking system is further constructed by loading tannic acid (TA). During the preparation process, multiple mechanisms such as the reaction between polyphenols and amino groups, metal chelation, π-π interaction, and electrostatic attraction work together to form an intelligent responsive hydrogel with a three-dimensional network structure. As a natural cationic antibacterial polypeptide, the molecular properties of ε-polylysine endow it with dual biological functions. The lysine residues densely distributed on its molecular chain are one of the 8 essential amino acids for the human body. It can replace the hydrated cations on the surface of moist tissues through the cation replacement mechanism, prompting the polyphenol groups to form a stable bond with the wound tissue, achieving persistent adhesion in a high-humidity environment. The positively charged amino groups in the ε-polylysine molecule can produce electrostatic interaction with the negatively charged region on the platelet surface, inducing red blood cell aggregation and platelet adhesion by activating coagulation factors, thereby enhancing the hemostatic efficacy. As a natural polyphenol compound, gallic acid has three mechanisms to achieve antibacterial synergism: penetrating the bacterial cell wall, blocking the quorum sensing system, and inhibiting protein biosynthesis. It enhances vascular smooth muscle contraction by activating signal pathways and reduces blood loss. After the chelation of gallic acid with metal ions, not only the molecular stability is improved, but also the antibacterial efficacy is enhanced by increasing the cell membrane permeability. Tannic acid exhibits antibacterial properties by disrupting the bacterial membrane structure through hydrogen bonds, chelating essential metal ions with carboxyl groups to form chelates, and scavenging free radicals with phenolic structures to produce antioxidant properties. Thus, it helps to inhibit the growth of bacteria in wound dressings and reduce the risk of infection. When Fe in ferric chloride hexahydrate 3+ forms a coordination structure with catechol groups, the ligand-to-metal charge transfer mechanism triggers a significant enhancement effect of light absorption. Under the excitation of near-infrared light (NIR, 808 nm), the π→π* electronic transition of the polyphenol ligand and the d-d orbital transition of Fe 3+ produce a synergistic effect, enabling the composite to possess broadband absorption characteristics, a high molar extinction coefficient, and excellent photothermal conversion efficiency. This intelligent responsive system maintains the temperature of the treatment area through a temperature control feedback mechanism, which can not only effectively inactivate pathogenic bacteria but also avoid thermal damage to normal tissues.

[0004] In summary, it is of great significance to develop a hydrogel dressing with photothermal multiple antibacterial and hemostatic functions. This dressing is based on ε-polylysine, gallic acid, and tannic acid, and forms a composite system by loading Fe 3+ Under the irradiation of an 808 nm laser, this hydrogel can intelligently adjust its properties to achieve a photothermal response, thereby providing multiple antibacterial protection and enhancing the hemostatic effect. This innovative dressing not only improves the success rate of wound treatment but also significantly improves the quality of life of patients. Summary of the Invention

[0005] The present invention relates to a hydrogel dressing with photothermal multiple antibacterial and hemostatic functions. This dressing is based on loading Fe 3+Based on ε-polylysine / gallic acid / tannic acid, it aims to provide a new method for wound treatment to effectively prevent and treat wound infections. It mainly introduces metal-polyphenol complexes through gallic acid-functionalized ε-polylysine to construct a hydrogel system. Mainly utilize gallic acid-functionalized ε-polylysine to introduce metal-polyphenol complexes into the hydrogel system. The components of this hydrogel dressing, ε-polylysine, gallic acid, and tannic acid, have significant broad-spectrum antibacterial activity, excellent wound-healing promotion ability, and good biocompatibility. In addition, when Fe in ferric chloride hexahydrate 3+ forms a coordination structure with catechol groups, it greatly enhances its light absorption ability in the near-infrared light (NIR) region and endows the hydrogel with superior photothermal conversion performance. Under light illumination, this hydrogel can effectively inhibit various pathogens through the photothermal effect, thus providing a good aseptic healing environment for the wound.

[0006] The object of the present invention is achieved through the following scheme: First, ε-polylysine is modified by gallic acid functionalization to obtain an EPL-GA complex, and then coordinated with Fe3+ to form a metal-polyphenol network, and a multiple dynamic cross-linking system is constructed by loading tannic acid. During the preparation process, through the synergistic action of multiple mechanisms such as polyphenol-amino reaction, metal chelation, π-π interaction, and electrostatic attraction, a hydrogel with a three-dimensional network structure is formed. The design of the hydrogel takes into account hemostasis and multiple antibacterial properties. ε-polylysine, gallic acid, and tannic acid have antibacterial effects. ε-polylysine has strong antibacterial, anti-inflammatory, and antioxidant abilities, excellent wound-healing promotion ability, and good biocompatibility. Gallic acid and tannic acid produce antibacterial effects by forming hydrogen bonds with hydroxyl groups in the hydrogel network. The preparation of a hydrogel for photothermal multiple antibacterial hemostasis function includes the following steps:

[0007] 1. Preparation process of the cross-linking agent: Dissolve 4-formylbenzoic acid (4-FA), 4-dimethylaminopyridine (DMAP), and dicyclohexylcarbodiimide (DCC) in 50 mL of tetrahydrofuran (THF). The molar ratio of 4-formylbenzoic acid: 4-dimethylaminopyridine: dicyclohexylcarbodiimide is 1:4:6. Activate 4-formylbenzoic acid in a nitrogen atmosphere, and drop polyethylene glycol (PEG) into the reaction system, and react for 24 - 48 h. After the reaction is completed, precipitate and purify with ice ether, and the product is end-diphenylaldehyde polyethylene glycol (PEGDA).

[0008] 2. Preparation process of polymer ligand: Dissolve ε-polylysine (EPL), gallic acid (GA), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl), and N-hydroxysuccinimide (NHS) in 50 mL of 2-morpholinoethanesulfonic acid (MES) buffer solution. The molar ratio of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride:N-hydroxysuccinimide:gallic acid is 1:1:0.8 - 0.9. The molar ratio of ε-polylysine:gallic acid is 1:100. Activate gallic acid in a nitrogen atmosphere and react for 2 - 4 h. After the reaction, perform dialysis purification, dialyze with 0.01 - 0.05 mol / L hydrochloric acid (HCl) solution for 24 - 48 h, and then dialyze with deionized water for 48 - 60 h, followed by freeze-drying. The product is gallic acid-modified ε-polylysine polymer (EPL-GA).

[0009] 3. Preferably, the relative molecular mass of the polyethylene glycol described in step 1 is 2000.

[0010] 4. Preferably, the molar ratio of polyethylene glycol, p-formylbenzoic acid, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide described in step 1 can be 1:4:4.4:6.

[0011] 5. Preferably, the relative molecular mass of the ε-polylysine described in step 2 is 3800 - 4200.

[0012] 6. Preferably, the molar ratio of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride:N-hydroxysuccinimide:gallic acid described in step 2 is 1:1:0.8 - 0.9.

[0013] 7. Preferably, the molar ratio of the ε-polylysine and gallic acid described in step 2 can be 1:100.

[0014] 8. Preferably, the concentration of the MES buffer solution described in step 2 is 0.05 mol / L and pH = 5.5.

[0015] 9. Preferably, weigh the gallic acid-modified ε-polylysine polymer, terminal dibenzaldehyde polyethylene glycol, ferric chloride hexahydrate, and tannic acid, and ultrasonically dissolve them respectively to prepare solutions. Add them to the mold in sequence, stir evenly with a vortex stirrer, and adjust the pH to 8.5 - 9 with sodium hydroxide solution. React for 30 s to obtain EPL-GA-TA@Fe hydrogel.

[0016] 10. Preferably, the mass ratio of the gallic acid-modified ε-polylysine polymer:terminal dibenzaldehyde polyethylene glycol:tannic acid:ferric chloride hexahydrate described in step 9 is 1:(4.5 - 5):(0 - 0.2):(0 - 0.05).

[0017] 11. Preferably, the concentration of the sodium hydroxide solution described in step 9 is 0.1 mol / L.

[0018] 12. Use of the hydrogel having photothermal multiple antibacterial hemostatic functions in the preparation of wound dressings.

[0019] The present invention has the following advantages and effects compared with the prior art:

[0020] 1. The present invention has a "chemical-physical" dual hemostatic mechanism: the present invention modifies ε-polylysine by grafting gallic acid, so that the modified ε-polylysine and Fe 3+ The complex is formed by complexation and blended with tannic acid. Among them, the cationic properties of ε-polylysine enable it to interact with the negative charge on the platelet surface, thereby inducing red blood cell aggregation and platelet adhesion. At the same time, gallic acid has a vasoconstrictive effect, which helps to reduce bleeding. The introduction of the "physical-chemical" dual hemostatic mechanism effectively improves the hemostatic efficiency and breaks through the limitations of single physical hemostasis.

[0021] 2. The present invention has a light-responsive antibacterial effect: through Fe 3+ When forming a coordination structure with the catechol group, the charge transfer mechanism from the ligand to the metal triggers an enhanced light absorption effect. Under near-infrared light, the hydrogel can quickly rise from 20°C to 50°C within 3 minutes and tend to be stable; after removing the light source, the gel can return to its initial temperature in a very short time. It has fast and reversible photothermal properties, which helps to avoid the invasion of external bacteria and reduce the risk of post-injury infection.

[0022] 3. The present invention exhibits excellent biocompatibility, which is mainly attributed to the dual biological functional characteristics of the ε-polylysine molecule. The densely distributed lysine residues in its molecular structure, as one of the eight essential amino acids for the human body, achieve a high affinity with the organism through the bionic mussel design. This innovative design effectively overcomes the drug resistance bottleneck of traditional materials and significantly improves the chemical stability of the active ingredients, thereby raising biosafety to a new standard. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 It is a representation of the principle of the present invention, forming a process and a functional demonstration diagram.

[0024] Figure 2 This is the synthesis route of the cross-linking agent (PEGDA) and the synthesis route of the polymer ligand (EPL-GA) in Example 1.

[0025] Figure 3 The H NMR spectra of the polymer ligand (EPL-GA) and ε-polylysine grafted gallic acid in Example 1, the solvent is D 2O。

[0026] Figure 4 Scanning electron microscope images of the hydrogels of Examples 2, 3, and 4.

[0027] Figure 5 Photothermal imaging diagrams of the temperature change over time of the hydrogels containing different masses of ferric chloride hexahydrate in Examples 2, 3, and 4, graphs of the temperature change over time, and the photothermal cycling curve of the hydrogel of Example 4.

[0028] Figure 6 Blood compatibility diagrams of the hydrogels of Examples 2, 3, and 4.

[0029] Figure 7 Pictures of the hemostasis process of the hydrogels of Examples 2, 3, and 4 with different materials applied to the mouse tail and liver bleeding models and pictures of the filter paper after hemostasis; the blood loss after hemostasis with different materials (*p < 0.05, **p < 0.01, ***p < 0.001).

[0030] Figure 8 Macrographs of the antibacterial properties of the hydrogels of Examples 2, 3, and 4.

[0031] Figure 9 Wound healing effect diagrams of the hydrogel of Example 4 and the blank group. Detailed implementation manners

[0032] The following examples are only applicable to further illustrate the present invention. It should be noted that all technologies and scientific terms used in the present invention have the same meaning as those in the technical field to which the present invention belongs unless otherwise specified. The experimental methods without specific conditions mentioned in the following examples are all conventional techniques in this technical field or are carried out according to the conditions recommended by the manufacturers; the reagents or instruments without indicating the manufacturers are all conventional products that can be obtained through commercial purchase.

[0033] The technical solutions of the present invention are further described below through specific examples. The following examples are further explanations of the present invention and do not limit the scope of the present invention.

[0034] Example 1

[0035] (1) 0.601 g of p-formylbenzoic acid, 0.538 g of 4-dimethylaminopyridine, and 1.238 g of dicyclohexylcarbodiimide were placed together in a two-necked flask with a magnetic stirring rotor, 50 mL of tetrahydrofuran was added to completely dissolve the drugs, and the mixed solution was reacted for 2 h in a nitrogen atmosphere. Subsequently, 2 g of polyethylene glycol (average molecular weight 2000) was dissolved in 10 mL of tetrahydrofuran and added to the two-necked flask, and the reaction was continued for 24 - 48 h in a nitrogen atmosphere. After the reaction was completed, it was precipitated and purified with ice ether, and the product was end-diphenylaldehyde polyethylene glycol (PEGDA).

[0036] (2) Place 2 g of ε-polylysine, 9.016 g of gallic acid, 11.176 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, and 6.709 g of N-hydroxysuccinimide together in a three-necked flask equipped with a magnetic stirring rotor. Add them to 50 mL of MES buffer (0.05 mol / L, pH = 5.5) to completely dissolve the drugs. The mixed solution reacts for 2 h in a nitrogen atmosphere. After the reaction, dialysis purification is carried out. Dialyze with 0.01 mol / L hydrochloric acid solution for 24 h, and then dialyze and purify with excess deionized water for 48 h. Freeze-dry, and the product is gallic acid-modified ε-polylysine polymer (EPL-GA).

[0037] Example 2

[0038] (1) Add 0.1 g of gallic acid-modified ε-polylysine polymer (EPL-GA) to 2.4 ml of deionized water and dissolve it by ultrasonic treatment to form a transparent solution.

[0039] (2) Add 0.435 g of end-dibenzaldehyde polyethylene glycol crosslinker to 2.4 ml of deionized water and dissolve it by ultrasonic treatment to form a solution.

[0040] (3) Add the solution in step (2) to the solution in step (1), and adjust the pH of the mixed solution to 8.5 - 9 with 0.1 mol / L sodium hydroxide solution. Stir evenly with a vortex stirrer to form the hydrogel EPL-GA.

[0041] Example 3

[0042] (1) Add 0.1 g of gallic acid-modified ε-polylysine polymer (EPL-GA) to 2 ml of deionized water and dissolve it by ultrasonic treatment to form a transparent solution.

[0043] (2) Add 0.435 g of end-dibenzaldehyde polyethylene glycol crosslinker to 2.4 ml of deionized water and dissolve it by ultrasonic treatment to form a solution.

[0044] (3) Add 0.02 g of tannic acid to 0.4 ml of deionized water and dissolve it by ultrasonic treatment to form a solution.

[0045] (4) Add the solution in step (2) and the solution in step (3) to the solution in step (1), and adjust the pH of the mixed solution to 8.5 - 9 with 0.1 mol / L sodium hydroxide solution. Stir evenly with a vortex stirrer to form the hydrogel EPL-GA-TA.

[0046] Example 4

[0047] (1) Add 0.1 g of gallic acid-modified ε-polylysine polymer (EPL-GA) to 2 ml of deionized water and dissolve it by ultrasonic treatment to form a transparent solution.

[0048] (2) Add 0.435 g of terminal dibenzaldehyde polyethylene glycol crosslinker and 4.5 mg of ferric chloride hexahydrate to 2.4 ml of deionized water and dissolve it by ultrasonic treatment to form a solution.

[0049] (3) Add 0.02 g of tannic acid to 0.4 ml of deionized water and dissolve it by ultrasonic treatment to form a solution.

[0050] (4) Add the solution of step (2) and the solution of step (3) to the solution of step (1), and adjust the pH of the mixed solution to 8.5 - 9 with 0.1 mol / L sodium hydroxide solution. Stir evenly with a vortex stirrer to form the hydrogel EPL-GA-TA@Fe.

[0051] The preparation steps of the hydrogel in this example are as follows:

[0052] In the present invention, a series of performance tests were carried out on the hydrogel dressings prepared in the examples and comparative examples to evaluate their applicability as photothermal hemostatic and antibacterial dressings in the treatment of wounds. The following are the detailed steps and purposes of the tests:

[0053] 1. Nuclear magnetic resonance spectrum:

[0054] Determine the molecular structure by nuclear magnetic resonance hydrogen spectrum. Dissolve the polymer in deuterated water and configure it to a concentration in the range of 0.5 - 1 mol / L to ensure the signal intensity. Then place the solution in an NMR sample tube for subsequent experiments.

[0055] The test results show that: Figure 3 a shows that the peaks with δ values between 3.49 and 3.55 ppm come from the two protons (-CH 2 -O-CH 2 -) of the ether methylene monomer unit, and the chemical shift peaks with δ values of 3.78 and 4.38 ppm are attributed to the protons of H 4 and H 3 respectively. The peak with δ value between 7.87 and 8.08 ppm corresponds to the aromatic protons of H 2 , and the aldehyde proton peak corresponding to H 1 is at δ value of 9.90 ppm, indicating the successful synthesis of the terminal dibenzaldehyde polyethylene glycol crosslinker. Figure 3 b shows that the position of specific H on EPL is between 1.2 - 3.8 ppm, while GA has a very obvious peak at 6.9 ppm. After the amidation reaction is completed, the ratio peak of EPL still exists in the product EPL-GA, proving the successful synthesis of the polymer.

[0056] 2. Scanning electron microscope:

[0057] The microscopic morphology of the hydrogel was observed by a scanning electron microscope. Before the scanning electron microscope observation, the hydrogel sample was frozen and crushed in liquid nitrogen, and then dried in a freeze-drying vacuum oven for 48 hours to maintain its structural integrity.

[0058] The test results showed that the hydrogel sample was freeze-dried and its internal structure was observed by scanning electron microscopy. The results Figure 4 showed that the hydrogel exhibited an obvious porous structure, and these three-dimensional interconnected porous structures were beneficial to the mechanical properties and response properties of the hydrogel. The morphologies of EPL-GA and EPL-GA-TA hydrogels at different magnifications showed that the pores were evenly distributed in the spatial structure and had similar sizes and shapes. However, the EPL-GA-TA@Fe hydrogel showed more microfibril structures than other hydrogels.

[0059] 3. Photothermal effect:

[0060] Near-infrared light with a wavelength of 808 nm (1 W·cm -2 ) was irradiated above each group of hydrogels, and the irradiation time for each group of hydrogels was set to 3 min. A thermal imaging camera was used to take photothermal images and record the temperature at specific times. The photothermal images of EPL-GA, EPL-GA-TA, and EPL-GA-TA@Fe hydrogels with different Fe 3+ contents under 808 nm near-infrared light irradiation were as shown in Figure 5 a, Figure 5 and the heating curves and photothermal curves of EPL-GA, EPL-GA-TA, and EPL-GA-TA@Fe hydrogels with different Fe 3+ contents under 808 nm near-infrared light irradiation were shown in b and 5c, respectively.

[0061] The test results showed that: as can be seen from Figure 5 a and 5b, due to the introduction of Fe 3+ , the photothermal effect of the EPL-GA-TA@Fe hydrogel increased significantly. As time went by, the photothermal effects of EPL-GA-TA@Fe hydrogels with different Fe 3+ contents were different. The hydrogel with a higher Fe 3 + concentration had a better photothermal effect and a higher temperature rise. After 3 min of irradiation, the maximum temperatures of EPL-GA-TA@Fe hydrogels with different Fe 3+ contents increased to 35.1 °C, 40 °C, and 52.4 °C, respectively, which further indicated that the main source of the photothermal effect was the chelation of Fe 3+ with the carboxyl groups in polyphenols to form Fe 3+ chelates. Figure 5c is the photothermal cycling curve of the hydrogel after turning on and off the near-infrared light source, indicating that the EPL-GA-TA@Fe hydrogel has excellent photothermal cycling effect.

[0062] 4. Hemolysis assay:

[0063] Preparation of platelet-rich plasma: The traditional two-step centrifugation method was used. 5 mL of fresh rat blood was drawn and stored in an anticoagulant tube containing sodium citrate and then centrifuged. The supernatant, the middle layer, and 1-2 mm below the erythrocyte layer were aspirated, and after centrifugation again, platelet deposition was visible at the bottom layer. The upper liquid was discarded, and the remaining serum and platelets were mixed evenly to obtain platelet-rich plasma. The obtained erythrocytes were washed 3 times with PBS and diluted to a final concentration of 5% (v / v). 0.2 g of EPL-GA, EPL-GA-TA, and EPL-GA-TA@Fe hydrogels were placed in a 48-well plate at 37 °C, and 1 mL of mouse erythrocyte suspension was added to each well. After the 48-well plate was incubated at 37 °C for 1 h, the suspension was centrifuged, and the supernatant was transferred to a 48-well plate. The absorbance of the supernatant was read at 540 nm by a microplate reader (Molecular Devices). 0.1% Triton X-100 (polyethylene glycol octylphenyl ether) was used as the positive control, and PBS was used as the negative control.

[0064] The test results show that: with the increase of the material, the hemolysis rate of the hydrogel sample decreased significantly, showing a hemolysis rate of less than 5%, Figure 6 indicating that the hydrogel has good blood compatibility.

[0065] 5. Antibacterial test:

[0066] The antibacterial activity of each group of hydrogels against Gram-negative bacterium Escherichia coli (E. coli) was determined by the plate counting method. The specific method was as follows: First, the hydrogel and Gram-negative Escherichia coli were cultured in NB culture medium. After irradiating vertically above the hydrogel with 808 nm near-infrared light for 15 min, it was cultured at 37 °C for 12 h to obtain a sample solution of bacteria co-incubated. The sample solution of bacteria co-incubated was diluted with NB culture medium by the ten-fold dilution method. 200 μL of the diluted sample solution of bacteria co-incubated was evenly spread on the agar medium. Finally, the agar culture dish was placed at 37 °C for culture, and the agar medium was photographed. The colony numbers of the treated sample group and the control group were calculated. Each group had 3 parallel samples, and the average value was taken. The colony numbers were counted in CFU / mL.

[0067] The test results show that: with the increase of the antibacterial material, the antibacterial performance of the hydrogel sample increased significantly, Figure 7 showing an obvious difference in the number of colonies. The prepared EPL-GA-TA@Fe showed good multiple antibacterial effects under photothermal conditions.

[0068] 6. Animal hemostasis test:

[0069] Mouse tail and liver defect bleeding models were used to evaluate the hemostatic effect of the adhesive hydrogel. Using surgical scissors, 50% of the tail length was cut off. After cutting, the mouse tail was placed in the air for 3 s to ensure normal blood loss. Immediately after applying the hydrogel to the bleeding site for 30 s, the weight of the filter paper containing the absorbed blood was measured and compared with the blank control group without hemostatic treatment. The liver of the mouse was exposed through an abdominal incision, and part of the large liver lobe was cut off with tissue scissors to construct a liver bleeding model. Immediately after applying the hydrogel to the bleeding site for 30 s, the weight of the filter paper containing the absorbed blood was measured and compared with the blank control group without hemostatic treatment. The bleeding condition of the wound was observed and photographed. All of the above operations needed to be performed on the mouse under a respiratory anesthesia machine.

[0070] The test results showed that: Figure 8 It was shown that the hydrogel of the present invention had a hemostatic effect, and the hemostasis rate reached 70.0% (P < 0.001).

[0071] 7. Animal full-thickness skin wound infection model:

[0072] The full-thickness skin wound infection model was used to further prove its effect on the wound. One day before the experiment, the hair on the back skin surface of the mouse was shaved off. During the experiment, the anesthetized rat was fixed in the prone position. The skin at the experimental site was disinfected with 70% alcohol. The epidermis and dermis on the left side of the midline of the rat's back were removed with a skin puncher and surgical forceps to form a circular incision with a diameter of 8 mm. Infection group: After treating the wound surface with the hydrogel of the present invention (the control group was not treated), 1×10 10 Staphylococcus aureus was added to the wound surface. Then the wound was covered with a medical surgical sterile dressing. Thereafter, the wound surface was wiped with sterile normal saline, and the dressing was changed every 3 days. The healing condition of the wound surface was observed and photographed at fixed time points. Two weeks after modeling, the back skin samples of the rats were taken and fixed with 4% paraformaldehyde. Histological analysis was performed on paraffin-embedded tissue sections (40 μm) by hematoxylin and eosin staining. All sections were observed and scanned using a microscope.

[0073] The test results showed that: The hydrogel was placed in a full-thickness infected wound of 8 mm×8 mm on the back of the mouse. Figure 9 As shown, the wound site gradually closed over time and was almost completely closed by the 14th day. The wound surface of the Staphylococcus aureus infection group healed more slowly. In contrast, the addition of the hydrogel of the present invention significantly reduced the inflammatory reaction caused by bacteria and accelerated the wound healing rate.

[0074] The above results indicated that the EPL-GA-TA@Fe hydrogel prepared by the present invention had excellent near-infrared assisted antibacterial ability, which was beneficial for it to become a potential biomedical material.

[0075] In summary, the present invention successfully prepares a photothermal antibacterial modified responsive hydrogel, which can aggregate blood cells and complete the hemostasis process in a short time. At the same time, it also has excellent antioxidant performance, can effectively eliminate oxygen free radicals, provide a suitable environment for the rapid healing of wounds, and has good photothermal response and high sterilization ability, providing a good sterile healing environment for the wound surface and promoting local tissue regeneration and wound surface healing.

[0076] Obviously, the above embodiments are merely examples given for clear illustration and are not limitations on the embodiments. For those of ordinary skill in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to enumerate all embodiments here. And the obvious changes or modifications derived therefrom are still within the protection scope of the present invention.

Claims

1. A photothermal multi-antibacterial hemostatic hydrogel, characterized in that: It includes the following steps: (1) First, p-formylbenzoic acid (4-FA), 4-dimethylaminopyridine (DMAP), and dicyclohexylcarbodiimide (DCC) are dissolved in 50 mL of tetrahydrofuran (THF), and the molar ratio of p-formylbenzoic acid: 4-dimethylaminopyridine: dicyclohexylcarbodiimide is 1:4:6; p-formylbenzoic acid is activated in a nitrogen atmosphere, and polyethylene glycol (PEG) is added dropwise to the reaction system for 24-48 hours; after the reaction is completed, the product is purified by precipitation with glacial ether, and the product is dibenzaldehyde-terminated polyethylene glycol (PEGDA); (2) Secondly, ε-polylysine (EPL), gallic acid (GA), 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC·HCl) and N-hydroxysuccinimide (NHS) were dissolved in 50 mL 2-morpholineethanesulfonic acid (MES) buffer, the molar ratio of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride: N-hydroxysuccinimide: gallic acid was 1:1:0.8-0.9; the molar ratio of ε-polylysine: gallic acid was 1:100; the gallic acid was activated in a nitrogen atmosphere and reacted for 2-4 hours; after the reaction, it was dialyzed and purified, dialyzed against 0.01-0.05 mol / L hydrochloric acid (HCl) solution for 24-48 hours, dialyzed against deionized water for 48-60 hours, and freeze-dried, and the product was gallic acid-modified ε-polylysine polymer (EPL-GA); (3) Weigh gallic acid-modified ε-polylysine polymer, dibenzaldehyde-terminated polyethylene glycol, ferric chloride hexahydrate, and tannic acid and dissolve them by ultrasonication to prepare solutions; add them into the mold in turn, stir them evenly with a vortex stirrer, and adjust the pH to 8.5-9 with sodium hydroxide solution; react for 30 seconds to obtain EPL-GA-TA@Fe hydrogel.

2. The photothermal multi-antibacterial hemostatic hydrogel according to claim 1, characterized in that: The relative molecular mass of the polyethylene glycol described in step (1) is 2000.

3. The photothermal multi-antibacterial hemostatic hydrogel according to claim 1, characterized in that: The relative molecular mass of the ε-polylysine described in step (2) is 3800-4200.

4. The photothermal multi-antibacterial hemostatic hydrogel according to claim 1, characterized in that: The molar ratio of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride:N-hydroxysuccinimide:gallic acid in step (2) is 1:1:0.8-0.

9.

5. The photothermal multi-antibacterial hemostatic hydrogel according to claim 1, characterized in that: The MES buffer concentration in step (2) is 0.05 mol / L, and the pH is 5.

5.

6. The photothermal multi-antibacterial hemostatic hydrogel according to claim 1, characterized in that: Gallic acid modified ε-polylysine polymer described in step (3): dibenzaldehyde-terminated polyethylene glycol: tannic acid: The mass ratio of ferric chloride hexahydrate is 1:(4.5-5):(0-0.2):(0-0.05).

7. The medical material according to any one of claims 1 to 6 comprises a drug sustained-release carrier and a wound dressing.

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