CN116440317B

CN116440317B - Photothermal antibacterial hydrogel and preparation method thereof

Info

Publication number: CN116440317B
Application number: CN202310357917.6A
Authority: CN
Inventors: 程翠; 张媛; 韩霄; 林抒今; 林婷
Original assignee: Fuzhou University
Current assignee: Fuzhou University
Priority date: 2023-04-06
Filing date: 2023-04-06
Publication date: 2024-09-03
Anticipated expiration: 2043-04-06
Also published as: CN116440317A

Abstract

The invention discloses a photothermal antibacterial hydrogel and a preparation method thereof. The invention takes bismuth oxide and epsilon-polylysine modified black phosphorus quantum dots, 3-aminophenylboric acid modified oxidized chondroitin sulfate and polyvinyl alcohol as raw materials, and forms BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel taking dynamic boric acid ester bonds as main materials, hydrogen bonds, pi-pi stacking and dynamic imine bond crosslinking as auxiliary materials in aqueous solution. The hydrogel has good multiple stimulus response, self-healing property, adhesiveness and bleeding stopping property, can realize synergistic antibacterial effect through the photo-thermal chemical effect, and can improve the wound microenvironment. Not only can provide good healing environment for common wounds, but also can accelerate the healing of diabetic infected wounds.

Description

Photothermal antibacterial hydrogel and preparation method thereof

Technical Field

The invention relates to a photothermal antibacterial hydrogel and a preparation method thereof, belonging to the technical field of medical biological materials.

Background

The skin serves as a first barrier for the human body, not only protects the viscera from the external environment, but also prevents the loss of body fluid without restriction. If the skin is improperly treated, bacteria can cause wound infection, which affects tissue repair and delays healing process. Particularly for Diabetic Feet (DF), the formation of drug-resistant bacterial biofilms and deregulation of the wound microenvironment make the wound difficult to heal, and in severe cases, cause infections, ulcers, and result in amputation. Clinically, debridement, systemic antibiotics, negative pressure therapy and maggot therapy are often utilized to combat diabetic foot wound infection caused by bacterial biofilm, but the effect is not ideal. In contrast, topical application of antimicrobial dressings is more beneficial for wound healing. However, traditional antibacterial dressing is easy to adhere to the wound when the wound is dry, and is easy to be polluted by exudates to infect the wound. The antibacterial hydrogel has the advantages of antibacterial property, ventilation, moisture retention, exudate control and absorption, and the like, so that the wound can be helped to heal rapidly.

The multifunctional antibacterial hydrogel dressing is prepared by polymerizing rigid natural polysaccharide functionalized by antibacterial activity quaternary ammonium salt and a flexible polyacrylamide network to form a double network, wherein two-dimensional nanomaterial phenylboronic acid functionalized graphene is taken as a nanofiller in prepolymerized dopamine. The multifunctional hydrogel has excellent adhesion performance and hemostatic performance, but has poor effect in treating diabetes wound infection.

As in the patent application No. CN115317660a, a near infrared corresponding antimicrobial nanocomposite hydrogel dressing is disclosed, which consists of an acrylamide-alginate hydrogel and bismuth sulfide nanocrystal particle-protein complexes loaded in situ in the hydrogel. The antibacterial nano composite hydrogel dressing has photothermal antibacterial property, but can not accelerate the healing of diabetic infected wounds.

Disclosure of Invention

The invention aims to solve the problems, and provides a photothermal antibacterial hydrogel and a preparation method thereof. According to the invention, bi ₂O₃ and epsilon-PL modified BPQDs, APBA modified OCS and PVA are used as raw materials, and BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel taking dynamic borate bonds as main materials and taking various non-covalent adhesion mechanisms as auxiliary materials is formed in an aqueous solution. The hydrogel not only promotes common wound healing, but also can accelerate diabetic infected wound healing by photo-thermal/chemical synergistic antibacterial and improving wound microenvironment.

The technical scheme for solving the problems is as follows:

A photothermal antibacterial hydrogel is prepared by mixing BP/Bi ₂O₃/epsilon-PL nanoparticle aqueous solution, APBA-g-OCS PBS solution and PVA aqueous solution;

The BP/Bi ₂O₃/epsilon-PL nanoparticle is formed by carrying out surface modification on epsilon-polylysine and occupying a defect site of BP by bismuth oxide;

The APBA-g-OCS is oxidized chondroitin sulfate modified by 3-aminophenylboric acid.

As a two-dimensional semiconductor nanomaterial, black Phosphorus (BP) has the advantages of high photo-thermal conversion efficiency, good biocompatibility and no toxicity of metabolites. However, the surface defects of 2D BP make it prone to decomposition under water-oxygen conditions. Bismuth oxide (Bi ₂O₃) can occupy the defect sites of BP, forming heterostructure nanoparticles to improve BP stability. Meanwhile, epsilon-polylysine (epsilon-PL) with excellent chemical antibacterial effect is modified on the surfaces of the nano particles to prepare the photo-thermal conversion agent BP/Bi ₂O₃/epsilon-PL, and the photo-thermal conversion agent BP/Bi ₂O₃/epsilon-PL is loaded into hydrogel, so that the photo-thermal conversion agent can resist bacterial biomembrane in chronic wounds.

The production of excess reactive oxygen species and the over-expression of inflammatory chemokines in diabetic foot wounds results in a deregulation of the wound microenvironment. Negatively charged sulfate groups on extracellular matrix glycosaminoglycans (e.g., chondroitin sulfate CS) can bind to positively charged amino acid residues on inflammatory chemokines through electrostatic interactions, regulating extracellular matrix chemokine transport to improve wound microenvironment and promote DF wound healing.

PVA is a natural polymer material, has good biocompatibility and good stability in aqueous solution, and can be used as a cross-linking agent and a supporting material in hydrogel.

The preparation method of the photothermal antibacterial hydrogel comprises the following steps:

s1, dispersing the ground black phosphorus crystals in methyl pyrrolidone (NMP), carrying out ice bath and ultrasonic centrifugation to obtain supernatant containing Black Phosphorus Quantum Dots (BPQDs) with uniform particle size;

S2, adding epsilon-PL alcohol solution and Bi (NO ₃)₃·5H₂ O alcohol solution) into the BPQDs solution, uniformly stirring, standing, centrifuging, taking precipitate to disperse in water, and preparing BP/Bi ₂O₃/epsilon-PL nanoparticle solution;

S3, adding NaIO ₄ into the Chondroitin Sulfate (CS) solution, uniformly mixing, stirring in a dark place, adding reducing alcohol, dialyzing the obtained solution, and freeze-drying the dialysate to obtain Oxidized Chondroitin Sulfate (OCS);

S4, preparing the OCS into an aqueous solution, adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS), performing a first pH value adjustment and light-proof reaction, adding 3-aminophenylboric acid (APBA) solution, performing a second pH value adjustment and light-proof reaction, dialyzing the obtained reaction solution, and performing freeze drying on the dialyzate to obtain 3-aminophenylboric acid modified oxidized chondroitin sulfate (APBA-g-OCS);

S5, mixing the BP/Bi ₂O₃/epsilon-PL nanoparticle solution, the APBA-g-OCS solution and the PVA aqueous solution to obtain a wet BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel;

The APBA-g-OCS solution is prepared by dissolving the APBA-g-OCS in PBS.

Preferably, in step S1, the grinding of the black phosphorus crystal is performed in N ₂.

Preferably, the supernatant containing BPQDs is further diluted to 0.1 to 0.3 mg/mL.

Preferably, the supernatant containing BPQDs at the concentration of 0.1-0.3 mg/mL is further refrigerated at the temperature of 3-5 ℃ for standby.

In the step S2, preferably, the solvents of the epsilon-PL alcohol solution and the Bi (NO ₃)₃·5H₂ O alcohol solution) are ethylene glycol, the concentration of the epsilon-PL alcohol solution is 2-6 mg/mL, the concentration of the Bi (NO ₃)₃·5H₂ O alcohol solution is 30-34 mg/mL, and the standing time is 4-6 h.

Preferably, in the step S2, the mass ratio of the epsilon-PL to the Bi (NO ₃)₃·5H₂ O to BPQDs) is (0.8-1.2) to (16-24) to (0.8-1.2), and the concentration of the BP/Bi ₂O₃/epsilon-PL nanoparticle solution is 0.01-0.45 mg/mL.

Preferably, in the step S3, the concentration of the CS solution is 8-12 mg/mL; the light-shielding stirring time is 5-7 hours; the reducing alcohol is ethylene glycol; the dialysis time is 2-4 d; the dialysis process comprises the steps of changing water every 6-8 hours; the freeze drying temperature of the dialysate is-70 to-90 ℃; the freeze drying time of the dialysate is 2-4 d; the molar ratio of CS to NaIO ₄ is (0.8-1.2) to (2.4-3.6).

Preferably, in step S4, the concentration of the OCS solution is 3-5 mg/mL; the pH value is adjusted to be 4-5 for the first time, and the light-shielding reaction time is 0.5-1.5 h; the pH value is adjusted to 4-5 for the second time, and the light-shielding reaction time is 10-12 hours; the dialysis condition is that the dialysis is carried out for 1.5-3 d in an aqueous solution with the pH value of 4-5 and for 1.5-3 d in ultrapure water; the freeze drying temperature of the dialysate is-70 to-90 ℃; and the freeze drying time of the dialysate is 2-4 d.

Preferably, in the step S4, the concentration of the APBA solution is 11-13 mg/mL; the solvent of the APBA solution is dimethyl sulfoxide (DMSO); the molar ratio of OCS, EDC, NHS, APBA is (0.8-1.2) to (0.8-1.2).

Preferably, in step S5, the pH of the PBS is 7.2 to 7.4; the concentration of the APBA-g-OCS solution is 6-12 wt%; the concentration of the PVA solution is 2-4wt%; the reaction time is 25-35 s.

Preferably, in the step S5, the mass ratio of BP/Bi ₂O₃/epsilon-PL to APBA-g-OCS to PVA is as follows: (0.01-0.45): (6-12): (2-4).

The invention has the following beneficial effects:

1. the photo-thermal antibacterial hydrogel prepared by the invention has multiple stimulus responsiveness, self-healing property, adhesiveness and hemostatic property, and can effectively close wounds;

2. the photo-thermal antibacterial hydrogel prepared by the invention can realize photo-thermal/chemical synergistic antibacterial effect, and is beneficial to resisting wound infection;

3. The photo-thermal antibacterial hydrogel prepared by the invention has the capabilities of removing active oxygen and adsorbing inflammatory chemokines, and can improve the microenvironment of wounds;

4. the photothermal antibacterial hydrogel prepared by the invention not only can provide a good healing environment for common wounds, but also can accelerate healing of diabetic infected wounds;

5. the raw materials used in the invention are cheap and easy to obtain, the preparation process is simple and pure, the preparation of the hydrogel does not need an organic solvent, the preparation period is extremely short, and the gel can be formed in situ within 30 s, and expensive instruments are not needed.

Drawings

FIG. 1 is a flow chart of the preparation of a photothermal antimicrobial hydrogel;

FIG. 2 is an SEM image of hydrogels with mass ratios of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0:9:3 (Blank gel) and 0.25:9:3 (NPs@gel-2);

FIG. 3 is a graph showing the temperature rise of a hydrogel having a mass ratio of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0:9:3 (Blank gel) and 0.25:9:3 (NPs@gel-2) by NIR irradiation 15min of 808 nm;

FIG. 4 is a self-healing plot of a hydrogel having a mass ratio of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0.25:9:3 (NPs@gel-2);

FIG. 5 shows the adhesion of hydrogel (a) to pig skin with a mass ratio of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0.25:9:3 (NPs@gel-2); (b, d) adhesion to various tissues and different materials; (c) adhesion to human joints;

FIG. 6 shows the degradation rates of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA at a mass ratio of 0.25:9:3 (NPs@gel-2) in different solutions (PBS, 1mM H ₂O₂, 16.6 mM Glucose) immersed in different solutions at different time intervals;

FIG. 7 shows that hydrogels with mass ratios of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0:9:3 (Blank gel) and 0.25:9:3 (NPs@gel-2) scavenge ROS in cells;

FIG. 8 is an adsorption capacity of the hydrogels with a mass ratio of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0:9:3 (Blank gel) and 0.25:9:3 (NPs@gel-2) to inflammatory chemokines IL-8 and MCP-1;

FIG. 9 shows the antibacterial activity of hydrogels with BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA in mass ratios of 0:9:3 (Blank gel) and 0.25:9:3 (NPs@gel-2);

FIG. 10 shows the hemostatic properties of a hydrogel having a mass ratio of BP/Bi ₂O₃/ε -PL, APBA-g-OCS and PVA of 0.25:9:3 (NPs@gel-2).

Detailed Description

The present invention will be described in detail with reference to the accompanying drawings.

Example 1

FIG. 1 is a flow chart showing the preparation of a photothermal antimicrobial hydrogel; comprises the preparation of BP/Bi ₂O₃/epsilon-PL nano particles, the preparation of APBA-g-OCS and the preparation of BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel in sequence.

Preparation of BP/Bi ₂O₃/epsilon-PL nanoparticles

The preparation method of the BP/Bi ₂O₃/epsilon-PL nanoparticle comprises the following steps:

Dispersing black phosphorus crystal ground in N ₂ in NMP, ice-bathing, centrifuging after ultrasonic treatment, quantitatively diluting supernatant containing uniform particle size BPQDs to 0.2 mg/mL, and storing in refrigerator;

Adding 4 mg/mL epsilon-PL alcohol solution and 32 mg/mL Bi (NO ₃)₃·5H₂ O alcohol solution) which are both ethylene glycol into the BPQDs solution, uniformly stirring the obtained mixed solution, standing at room temperature for 5h, centrifuging, dispersing 0mg precipitate in water, and preparing 0 mg/mL BP/Bi ₂O₃/epsilon-PL nanoparticle solution.

In the above method, the mass ratio of each substance is epsilon-PL to Bi (NO ₃)₃·5H₂ O: BPQDs =1:20:1.

Preparation of APBA-g-OCS

The preparation method of the APBA-g-OCS comprises the following steps:

Adding NaIO ₄ into 10 mg/mL CS solution, stirring in dark for 6 h, adding 1 mL ethylene glycol to terminate the reaction, placing the obtained solution in a dialysis bag, dialyzing at room temperature for 3d, and freeze-drying the obtained dialysate at-80 ℃ for 3d to obtain OCS;

EDC and NHS are added into the prepared OCS solution with the concentration of 4 mg/mL, the pH value of the solution is regulated to 4-5, and the reaction is carried out at room temperature and in a dark place for 1 h; adding APBA solution with a solvent of DMSO and a concentration of 12 mg/mL into the solution, adjusting the pH value to 4-5, and carrying out light-shielding reaction at room temperature overnight; the reaction solution is placed in a dialysis bag, dialyzed for 2 days in aqueous solution with pH value of 4-5, dialyzed for 2 days in ultrapure water, and the obtained dialysate is freeze-dried at-80 ℃ for 3d, so that APBA-g-OCS is obtained.

In the method, the molar ratio of the substances is CS to NaIO ₄ =1 to 3; OCS (optical system): EDC: NHS: APBA = 1:1:1:1.

Preparation of BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel

At room temperature: APBA-g-OCS was dissolved in PBS at ph=7.4 to prepare a 9wt% APBA-g-OCS solution; PVA is dissolved in distilled water to prepare a PVA solution with the weight percent of 3; the mass ratio is as follows: BP/Bi ₂O₃/epsilon-PL/APBA-g-OCS/PVA=0:9:3, and reacting the mixed solution with 30: 30 s to obtain the BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel in a wet state; and freeze-drying the obtained wet hydrogel to obtain the dry BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel.

Example 2

Preparation of BP/Bi ₂O₃/epsilon-PL nanoparticles

Adding 4 mg/mL of epsilon-PL alcohol solution and 32 mg/mL of Bi (NO ₃)₃·5H₂ O alcohol solution) which are both ethylene glycol into the BPQDs solution, uniformly stirring the obtained mixed solution, standing at room temperature for 5h, centrifuging, taking precipitate, dispersing in water, and preparing 0.25 mg/mL of BP/Bi ₂O₃/epsilon-PL nanoparticle solution.

In the above method, the mass ratio of each substance is ε -PL to Bi (NO ₃)₃·5H₂ O: BPQDs =1:20:1.

Preparation of APBA-g-OCS

The preparation method of the APBA-g-OCS comprises the following steps:

Adding NaIO ₄ into 10 mg/mLCS solution, stirring in the absence of light for 6 h, adding 1mL ethylene glycol to terminate reaction, placing the obtained solution in a dialysis bag, dialyzing at room temperature for 3d, and freeze-drying the obtained dialysate at-80deg.C for 3d to obtain OCS;

In the method, the molar ratio of the substances is CS to NaIO ₄ =1 to 3; OCS: EDC: NHS: APBA=1:1:1:1.

Preparation of BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel

At room temperature: APBA-g-OCS was dissolved in PBS at ph=7.4 to prepare a 9wt% APBA-g-OCS solution; PVA is dissolved in distilled water to prepare a PVA solution with the weight percent of 3; the mass ratio is as follows: BP/Bi ₂O₃/epsilon-PL/APBA-g-OCS/PVA=0.25:9:3, and reacting the mixed solution with 30: 30 s to obtain the BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel in a wet state; and freeze-drying the obtained wet hydrogel to obtain the dry BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel.

Example 3

Preparation of BP/Bi ₂O₃/epsilon-PL nanoparticles

Adding 4 mg/mL of epsilon-PL alcohol solution and 32 mg/mL of Bi (NO ₃)₃·5H₂ O alcohol solution) which are both ethylene glycol into the BPQDs solution, uniformly stirring the obtained mixed solution, standing at room temperature for 5h, centrifuging, taking precipitate, dispersing in water, and preparing 0.15 mg/mL of BP/Bi ₂O₃/epsilon-PL nanoparticle solution.

Preparation of APBA-g-OCS

The preparation method of the APBA-g-OCS comprises the following steps:

Preparation of BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel

At room temperature: APBA-g-OCS was dissolved in PBS at ph=7.4 to prepare a 7.5wt% APBA-g-OCS solution; PVA is dissolved in distilled water to prepare a PVA solution with the weight percent of 3; the mass ratio is as follows: BP/Bi ₂O₃/epsilon-PL/APBA-g-OCS/PVA=0.15:7.5:3, and reacting the mixed solution with 30: 30 s to obtain wet BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel; and freeze-drying the obtained wet hydrogel to obtain the dry BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel.

Example 4

Preparation of BP/Bi ₂O₃/epsilon-PL nanoparticles

Adding 4 mg/mL of epsilon-PL alcohol solution and 32 mg/mL of Bi (NO ₃)₃·5H₂ O alcohol solution) which are both ethylene glycol into the BPQDs solution, uniformly stirring the obtained mixed solution, standing at room temperature for 5h, centrifuging, taking precipitate, dispersing in water, and preparing 0.35 mg/mL of BP/Bi ₂O₃/epsilon-PL nanoparticle solution.

Preparation of APBA-g-OCS

The preparation method of the APBA-g-OCS comprises the following steps:

Preparation of BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel

At room temperature: APBA-g-OCS was dissolved in PBS at ph=7.4 to prepare a 10.5wt% APBA-g-OCS solution; PVA is dissolved in distilled water to prepare a PVA solution with the weight percent of 3; the mass ratio is as follows: BP/Bi ₂O₃/epsilon-PL/APBA-g-OCS/PVA=0.15:10.5:3, and reacting the mixed solution with 30: 30 s to obtain wet BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel; and freeze-drying the obtained wet hydrogel to obtain the dry BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA hydrogel.

Hydrogel performance detection:

the hydrogels obtained in examples 1 and 2 were designated as Blank gel and NPs@gel-2, respectively.

As can be seen from FIG. 2, both the Blank gel and the NPs@gel-2 hydrogels exhibit a highly porous and interconnected network structure with relatively uniform pores that facilitates the absorption of the tissue excess exudates by the hydrogels as wound dressings.

As can be seen from FIG. 3, after 15min of NIR irradiation of the gel 808 nm, 1W/cm ² with both Blank gel and NPs@gel-2, the Blank gel temperature was raised to 32.5℃and the NPs@gel-2 temperature was raised to 63.8 ℃; it is shown that the NPs@gel-2 hydrogel has good photo-thermal properties, and the photo-thermal properties of the hydrogel are endowed by BP/Bi ₂O₃/epsilon-PL nanoparticles.

As can be seen from fig. 4, the broken hydrogel had no external intervention, and after 300s, the crack disappeared and healed completely, indicating that the nps@gel-2 hydrogel had a rapid, stable, efficient self-healing capacity, which was beneficial to providing a closed, moist environment to the wound.

As can be seen from fig. 5a, in the absence of any external force, the weight 10min of 75 g can be suspended from two pigskin pieces adhered by the hydrogel, and still cannot fall off; as can be seen from fig. 5b and 5d, nps@gel-2 hydrogel can adhere to different visceral surfaces (e.g. kidney, lung, heart, liver) and different material surfaces (e.g. iron, rubber, plastic, glass, paper) of rats and shows good adhesion; as can be seen in FIG. 5c, the NPs@gel-2 hydrogel adheres strongly to the skin surface and accommodates severe frequent and large amounts of finger articulation. These results demonstrate that nps@gel-2 hydrogels are beneficial for wound closure and are not easily destroyed when used as wound dressings, prolonging dressing life.

As can be seen from fig. 6, the degradation rate of nps@gel-2 was slower at ph=7.4 than at ph=5.2, 1 mM H ₂O₂ and 16 mM Glucose. After incubation of 6h, the remaining weight ratios of nps@gel-2 were 51%, 36%, 25% and 19% at ph=7.4, ph=5.2, 1 mM H ₂O₂ and 16 mM Glucose, respectively; these results indicate that hydrogels have pH, hydrogen peroxide and glucose dependent degradation behavior; this facilitates the accelerated degradation of nps@gel-2 hydrogels at DF wounds with high sugar and high active oxygen concentration, releasing more active substances to combat wound infection and regulate wound microenvironment.

As can be seen from FIG. 7, NIH3T3 cells treated with the Blank gel and NPs@gel-2 hydrogels showed significant fluorescence quenching, while control cells remained strongly fluorescent, indicating that Blank gel and NPs@gel-2 hydrogels can effectively scavenge active oxygen in the cells; this is beneficial to the NPs@gel-2 hydrogel to protect wound tissues from active oxygen, improve the microenvironment of the wound and avoid the excessive oxidative stress reaction of the wound.

As can be seen from fig. 8, the control group had almost no adsorption to the inflammatory chemokines MCP-1 and IL-8; the adsorption rates of the Blank gel and the NPs@gel-2 to the MCP-1 are respectively up to 87% and 82%; the adsorption rates of IL-8 by Blank gel and NPs@gel-2 are 81% and 73%, respectively; the NPs@gel-2 of 1 mL can adsorb the MCP-1 of 4.2 ng and the IL-8 of 775 ng, which shows that the hydrogel can effectively remove inflammatory chemokines, is beneficial to the hydrogel to regulate the transportation of the inflammatory chemokines, improves the microenvironment of wounds and accelerates the healing of DF infected wounds.

As can be seen from FIG. 9, the growth conditions of bacteria treated by different groups on an agar plate are observed, and the antibacterial rate of the Blank gel group on three bacteria is 15-18%, because boric acid molecules can be combined with glycol molecules of glycoprotein on the surface of bacterial cells, and a certain antibacterial effect is generated; the antibacterial rate of the NPs@gel-2 group to three bacteria is 84-89%, and the obvious antibacterial capacity mainly derives from the physicochemical effect between the amino group of epsilon-PL and the bacterial membrane, so that the bacterial membrane is broken to kill the bacteria; NPs@gel-2+NIR group shows the strongest antibacterial activity, and the inhibition rate of the NPs@gel-2+NIR group on three bacteria is 100%, which indicates that the combination of photo-thermal and chemical antibacterial is more beneficial to thoroughly killing the bacteria; this is beneficial to the nps@gel-2 hydrogel to resist bacterial infection at DF wounds through photo-thermal-chemical combination antibacterial, reduce inflammatory reaction and promote wound healing.

As can be seen from FIG. 10, the blood loss amounts after 5 min treatments with Control (+), medical gauze [ Control (-) ] and NPs@gel-2 were 253.3mg, 175.3mg and 24.3 mg, respectively; the time required for hemostasis of the Control (+) and Control (-) groups was 4 min, whereas the nps@gel-2 treatment group was immediately hemostatic; this is mainly due to the good adhesion of nps@gel-2, which can effectively close the liver bleeding site and prevent bleeding.

By combining the comparison data and the pictures, the photo-thermal antibacterial hydrogel prepared by the invention has rapid self-healing property, adhesiveness and hemostatic property, is beneficial to closing wounds and provides a good healing environment for the wounds; has photo-thermal/chemical synergistic sterilization effect, and is beneficial to resisting wound infection; the hydrogel has multiple stimulus responsivity such as pH, H ₂O₂, glucose and the like, and is beneficial to regulating and controlling the wound microenvironment; the ROS can be effectively removed, inflammatory chemokines are adsorbed, and the wound microenvironment can be regulated and controlled; the photothermal antibacterial hydrogel prepared by the invention not only can be beneficial to common wound healing, but also can effectively promote the repair of diabetic infected wounds.

Claims

1. The preparation method of the photothermal antibacterial hydrogel is characterized by comprising the following steps of:

s1, dispersing the ground black phosphorus crystals in NMP, carrying out ice bath and ultrasonic treatment, and centrifuging to obtain BPQDs supernatant containing uniform particle size;

S2, adding epsilon-PL alcohol solution and Bi (NO ₃)₃·5H₂ O alcohol solution) into the BPQDs supernatant, uniformly stirring, standing, centrifuging, taking precipitate to disperse in water, and preparing BP/Bi ₂O₃/epsilon-PL nanoparticle solution;

S3, adding NaIO ₄ into the CS solution, uniformly mixing, stirring in a dark place, adding reducing alcohol, dialyzing the obtained solution, and freeze-drying the dialysate to obtain OCS;

S4, preparing the OCS into an aqueous solution, adding EDC and NHS, performing a first pH value adjustment and light-proof reaction, adding an APBA solution, performing a second pH value adjustment and light-proof reaction, dialyzing the obtained reaction solution, and performing freeze drying on the dialyzate to obtain APBA-g-OCS;

S5, mixing the BP/Bi ₂O₃/epsilon-PL nanoparticle solution, the APBA-g-OCS solution and the PVA aqueous solution to obtain a wet BP/Bi ₂O₃/epsilon-PL@APBA-g-OCS/PVA antibacterial hydrogel; the APBA-g-OCS solution is prepared by dissolving the APBA-g-OCS in PBS;

Wherein NMP is N-methylpyrrolidone, BPQDs is black phosphorus quantum dot, epsilon-PL is epsilon-polylysine, bi (NO ₃)₃·5H₂ O is bismuth nitrate pentahydrate, CS is chondroitin sulfate, naIO ₄ is sodium periodate, OCS is oxidized chondroitin sulfate, EDC is 1-ethyl- (3-dimethylaminopropyl) carbodiimide, NHS is N-hydroxysuccinimide, APBA is 3-aminophenylboronic acid, APBA-g-OCS is 3-aminophenylboronic acid modified oxidized chondroitin sulfate, PVA is polyvinyl alcohol, and PBS is phosphate buffer.

2. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S1, the grinding of the black phosphorus crystals is performed in N ₂; the concentration of BPQDs supernatant is 0.1-0.3 mg/mL.

3. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in the step S2, the solvents of the epsilon-PL alcohol solution and the Bi (NO ₃)₃·5H₂ O alcohol solution) are ethylene glycol, the concentration of the epsilon-PL alcohol solution is 2-6 mg/mL, the concentration of the Bi (NO ₃)₃·5H₂ O alcohol solution is 30-34 mg/mL, and the standing time is 4-6 h.

4. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in the step S2, the mass ratio of epsilon-PL to Bi (NO ₃)₃·5H₂ O to BPQDs) is (0.8-1.2) to (16-24) to (0.8-1.2), and the concentration of the BP/Bi ₂O₃/epsilon-PL nanoparticle solution is 0.01-0.45 mg/mL.

5. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S3, the concentration of the CS solution is 8-12 mg/mL; the light-shielding stirring time is 5-7 hours; the reducing alcohol is ethylene glycol; the dialysis time is 2-4 d; the dialysis process comprises the steps of changing water every 6-8 hours; the freeze drying temperature of the dialysate is-70 to-90 ℃; the freeze drying time of the dialysate is 2-4 d; the molar ratio of CS to NaIO ₄ is (0.8-1.2) to (2.4-3.6).

6. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S4, the OCS is prepared to have a concentration of 3-5 mg/mL of aqueous solution; the pH value is adjusted to be 4-5 for the first time, and the light-shielding reaction time is 0.5-1.5 h; the pH value is adjusted to 4-5 for the second time, and the light-shielding reaction time is 10-12 hours; the dialysis condition is that the water solution with the pH value of 4-5 is dialyzed for 1.5-3 d, and the ultrapure water is dialyzed for 1.5-3 d; the freeze drying temperature of the dialysate is-70 to-90 ℃; the freeze drying time of the dialysate is 2-4 d.

7. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S4, the concentration of the APBA solution is 11-13 mg/mL; the solvent of the APBA solution is DMSO; OCS, EDC, NHS, APBA is (0.8-1.2) to (0.8-1.2).

8. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S5, the pH of the PBS is 7.2 to 7.4; the concentration of the APBA-g-OCS solution is 6-12 wt%; the concentration of the PVA aqueous solution is 2-4wt%; the reaction time is 25-35 s.

9. The method for preparing a photothermal antibacterial hydrogel according to claim 1, wherein in step S5, the mass ratio of BP/Bi ₂O₃/epsilon-PL: APBA-g-OCS: PVA is: (0.01-0.45): (6-12): (2-4).

10. A photothermal antibacterial hydrogel, characterized in that the photothermal antibacterial hydrogel is prepared by the preparation method of any one of claims 1 to 9; the photothermal antibacterial hydrogel is formed by mixing BP/Bi ₂O₃/epsilon-PL nanoparticle aqueous solution, APBA-g-OCS PBS solution and PVA aqueous solution;