A microsphere-containing wound infection monitoring and repair gel dressing

Abstract

The present application relates to the field of biological medicine, disclose a kind of containing microsphere's wound infection monitoring and repair gel dressing.Preparation method includes the following steps: molybdenum disulfide solution is dropped into sodium alginate solution, fully stir;Molybdenum disulfide sodium alginate solution is shot to calcium chloride solution, crosslinking is obtained by stationary, molybdenum disulfide gel microspheres;Sodium alginate and acrylamide are dissolved in water after stirring, add methylene bisacrylamide, polyacrylamide, acid-base indicator and ammonium persulfate, fully mixed and dissolved, obtain mixed solution;Molybdenum disulfide gel microspheres are added in mixed solution, tetramethyl ethylenediamine and calcium sulfate dihydrate are sequentially added under continuous stirring, and stirring is uniformly;After stationary coagulation, removing impurities, obtain containing microsphere's wound infection monitoring and repair gel dressing.The gel dressing of the present application has excellent tensile elastic modulus, real-time wound infection state monitoring, near-infrared light-controlled antibacterial treatment and the ability to promote wound healing.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biological medicine, and more particularly to a microsphere-containing wound infection monitoring and repairing gel dressing. BACKGROUND

[0002] Severe wound injuries, such as acute wounds of trauma, burns, etc., or chronic wounds of diabetes, pressure sores, etc., are often accompanied by complications such as bacterial infection, which seriously hinders the effective healing of the wound, and even causes severe sepsis leading to death. At present, the use of antibacterial drugs is still a common choice for local disinfection of wounds. In clinical treatment, various antibacterial preparations, including antibiotics (such as tetracyclines, penicillins and streptomycin), metal compounds (such as silver nitrate, bismuth sulfide and copper nicotinate), quaternary ammonium salts, peptides, etc., have been widely used to combat bacterial infections. However, the antibacterial process of these antibacterial preparations is usually difficult to accurately control, and in order to quickly suppress infection, the overuse of antibacterial agents often occurs in actual treatment. In most cases, the misuse or overuse of antibacterial drugs has been generally considered to have an important mutual relationship with bacterial drug-resistant infections and adverse reactions caused by treatment. In view of the above problems, new strategies for preventing and treating wound infections are being continuously explored. Among them, researchers have found that the combination of existing antibacterial preparations with hydrogel wound dressings is expected to improve the therapeutic effect of wound infection prevention and treatment. Because the hydrogel dressing not only provides a hydrophilic temporary physical barrier to resist external bacterial infection, but also maintains a moist healing environment to promote rapid healing of the wound. Some hydrogel dressings even have a loose porous structure, which can effectively absorb the tissue exudate of the wound, improve the oxygen exchange capacity of the wound, and guide the rapid reconstruction of the wound tissue.

[0003] In terms of literature reports, in 2021, Professor Wu Xiaoyang of the University of Chicago and Professor Zhu Linyong of East China University of Technology invented a light-induced imine cross-linked hydrogel wound dressing that can release transforming growth factor-β (TGF-β) inhibitors within a certain time after the wound occurs, and successfully achieved scar-free wound healing in mouse and large animal preclinical models. In 2022, Professor Gu Baolin of Xi'an Jiaotong University successfully developed a self-repairing hydrogel with good conductivity and antioxidant activity based on quaternized chitosan, oxidized dextran, tobramycin, and polydopamine-coated polypyrrole nanowires. In 2023, the team of Professor Du Yongzhong of Zhejiang University developed a new type of extracellular matrix hydrogel. By incorporating fat-derived mesenchymal stem cell-derived exosomes into the extracellular matrix hydrogel, the exosomes can be continuously released from the hydrogel, significantly increasing the concentration of exosomes at the wound site and promoting the rapid recovery of the wound. Related research results show that this hydrogel can effectively reduce inflammation, accelerate cell proliferation and migration, promote the formation of new blood vessels, improve collagen deposition at the wound site, and accelerate wound repair. Although there have been some breakthroughs in the use of hydrogel dressings to treat wound infections, existing research techniques still cannot achieve the real-time monitoring and precise on-demand treatment technology strategy proposed in this patent.

[0004] In the existing patent (A skin damage repair hydrogel dressing and a preparation method thereof, CN117357693B), the inventors synthesized a skin damage repair hydrogel using polypeptides, cordycepin, and oxidized hyaluronic acid, which can remove active oxygen in damaged tissues and produce nitric oxide, promote the migration function of vascular endothelial cells after radiation damage, significantly reduce DNA damage in cells, enhance the stability of cordycepin in tissues, reduce vascular endothelial cell aging, and promote skin damage repair. The invention patent (An antibacterial hydrogel material and a preparation method thereof, CN117582537A) prepared a hydrogel with high-efficiency antibacterial function, which can effectively maintain a microenvironment suitable for wound healing, promote the proliferation and differentiation of fibroblasts, accelerate the formation of new blood vessels and the reconstruction process of wound granulation tissue, and promote the healing process of the wound. The invention patent (Protein hybrid gel dressing with composite drug-loaded fibers and preparation method and application thereof, CN116942891A) discloses a protein hybrid gel dressing with composite drug-loaded fibers, which is mainly composed of a hybrid hydrogel and hydrophobic antibiotics loaded in the gel, where the hydrophobic antibiotics can be released from the gel dressing for long-term antibacterial effect.

[0005] However, most of the hydrogel dressings currently applied to the prevention and treatment of wound infection are based on the concentration-dependent drug diffusion release mechanism, which releases the loaded antibacterial drugs to the wound site for antibacterial purpose. The release process is usually difficult to control accurately, resulting in poor treatment effect. The main technical defects include the following two aspects: 1) The infection of the wound cannot be accurately monitored. Most hydrogel dressings lack the feedback ability of the wound infection. It cannot indicate whether the wound is infected or cannot monitor the process change of the wound from non-infection to infection in real time, resulting in the failure to discover the infection in the early stage of the wound and treat it in time, thus missing the best treatment period. 2) The conventional hydrogel wound infection treatment dressing has drug controlled release ability, but cannot realize accurate controlled release of the drug, resulting in mismatch between drug release and antibacterial treatment demand of the wound, such as too fast or too slow drug release, or excessive drug release, which cannot realize accurate on-demand treatment of the wound infection. SUMMARY

[0006] To overcome the defects of the prior art that the infection of the wound cannot be accurately monitored and the drug cannot be accurately controlled, the present application provides a microsphere-containing wound infection monitoring and repairing gel dressing.

[0007] Another object of the present application is to provide a preparation method of the microsphere-containing wound infection monitoring and repairing gel dressing.

[0008] Another object of the present application is to provide an application of the microsphere-containing wound infection monitoring and repairing gel dressing.

[0009] To solve the above technical problems, the technical solution of the present application is as follows:

[0010] A preparation method of a microsphere-containing wound infection monitoring and repairing gel dressing, comprising the following steps:

[0011] S1, adding a molybdenum disulfide solution to a sodium alginate solution, and fully stirring to obtain a molybdenum disulfide sodium alginate solution;

[0012] S2, injecting the molybdenum disulfide sodium alginate solution into a calcium chloride solution, and standing for crosslinking to obtain molybdenum disulfide gel microspheres;

[0013] S3, dissolving sodium alginate and acrylamide in water after stirring, adding methylene bisacrylamide, polyacrylamide, an acid-base indicator and ammonium persulfate, and fully mixing and dissolving to obtain a mixed solution;

[0014] S4, adding the molybdenum disulfide gel microspheres to the mixed solution, and sequentially adding tetramethyl ethylenediamine and calcium sulfate dihydrate under continuous stirring, and stirring uniformly;

[0015] S5, after standing, coagulation, and removal of impurities, a wound infection monitoring and repairing gel dressing containing microspheres is obtained.

[0016] Preferably, the molybdenum disulfide nanoparticles and sodium alginate are dispersed in deionized water respectively to obtain a molybdenum disulfide solution and a sodium alginate solution.

[0017] Preferably, when the molybdenum disulfide nanoparticles are dispersed in deionized water, the stirring speed is 100-1000 r / min and the stirring time is 5-20 min; when the sodium alginate is dispersed in deionized water, the stirring speed is 50-200 r / min and the stirring time is 1-3 h.

[0018] Preferably, when the molybdenum disulfide nanoparticles are dispersed in water, the stirring speed is 800 r / min and the stirring time is 10 min; when the sodium alginate is dispersed in water, the stirring speed is 100 r / min and the stirring time is 2 h.

[0019] Further, the concentration of the molybdenum disulfide solution in S1 is 0.01-1 g / mL, and the concentration of the obtained sodium alginate solution is 0.025-0.6 g / mL.

[0020] Preferably, the concentration of the obtained molybdenum disulfide solution in S1 is 0.1 g / mL, and the concentration of the obtained sodium alginate solution is 0.15 g / mL.

[0021] Preferably, in S1, the molybdenum disulfide solution is added to the sodium alginate solution.

[0022] Further, in S1, the volume ratio of the added molybdenum disulfide solution to the sodium alginate solution is 0.1:5-20.

[0023] Preferably, in S1, the speed of the sufficient stirring is 200-1000 r / min and the time is 5-20 min.

[0024] Preferably, in S1, the speed of the sufficient stirring is 800 r / min and the time is 10 min.

[0025] Further, in S2, the molybdenum disulfide sodium alginate solution is extruded into the calcium chloride solution at a flow rate of 0.02-1.0 mL / min by external force; the speed generated by the external force is 10.5-15.5 m / s.

[0026] Preferably, in S2, the molybdenum disulfide sodium alginate solution is extruded and blown into the calcium chloride solution by wind at a flow rate of 0.02-1.0 mL / min; the wind speed of the wind is 10.5-15.5 m / s.

[0027] Preferably, the flow rate is 0.1 mL / min and the wind speed of the wind is 12.5 m / s.

[0028] Preferably, the blowing is performed using a micro-injection pump.

[0029] Further, the mass percentage of the calcium chloride solution in S2 is 0.5% to 2%.

[0030] Preferably, the mass percentage of the calcium chloride solution in S2 is 1.5%.

[0031] Preferably, the standing time in S2 is 10 to 50 min.

[0032] Preferably, the standing time in S2 is 30 min.

[0033] Preferably, after the standing, the molybdenum disulfide gel microspheres are washed to be redispersed in deionized water.

[0034] Further, the concentration of sodium alginate in the mixed solution in S3 is 0.01 to 0.04 g / mL, the concentration of acrylamide is 0.05 to 0.25 g / mL, the concentration of methylene bis-acrylamide is 0.05 to 0.15 mg / mL, the concentration of polyacrylamide is 0.5 to 3 mg / mL, the concentration of acid-base indicator is 0.05 to 0.5 mg / mL, and the concentration of ammonium persulfate is 0.5 to 3 mg / mL.

[0035] Preferably, the concentration of sodium alginate in the mixed solution in S3 is 0.025 g / mL, the concentration of acrylamide is 0.135 g / mL, the concentration of methylene bis-acrylamide is 0.085 mg / mL, the concentration of polyacrylamide is 1 mg / mL, the concentration of acid-base indicator is 0.2 mg / mL, and the concentration of ammonium persulfate is 1.5 mg / mL.

[0036] Preferably, the acid-base indicator is phenol red.

[0037] Preferably, the stirring time of sodium alginate and acrylamide in S3 is 3 h.

[0038] Preferably, when the mixing and dissolving are sufficient, the stirring speed is 200 to 1000 r / min, and the stirring time is 5 to 50 min.

[0039] Preferably, when the mixing and dissolving are sufficient, the stirring speed is 500 r / min, and the stirring time is 30 min.

[0040] Further, after the stirring in S4 is uniform, the concentration of the molybdenum disulfide gel microspheres is 0.2 to 0.8 g / mL, the concentration of tetramethyl ethylenediamine is 0.3 to 2 μL / mL, and the concentration of calcium sulfate dihydrate is 0.001 to 0.01 g / mL.

[0041] Preferably, the concentration of the molybdenum disulfide gel microspheres is 0.4 g / mL, the concentration of the tetramethylethylenediamine is 1.5 μL / mL, and the concentration of the calcium sulfate dihydrate is 0.004 g / mL after the stirring in S4.

[0042] Preferably, the mixed solution is defoamed in S4.

[0043] Preferably, the tetramethylethylenediamine is added and stirred for 2-10 min in S4, and then the calcium sulfate dihydrate powder is slowly added through a sieve while stirring.

[0044] Preferably, the sieve is 100 mesh.

[0045] Preferably, the standing time in S5 is 1-3 days.

[0046] Preferably, the standing is at 4°C for 2 days.

[0047] A wound infection monitoring and repairing gel dressing containing microspheres is prepared by the preparation method.

[0048] Further, the microspheres are molybdenum disulfide gel microspheres with a diameter of 300-1000 μm.

[0049] The application of the wound infection monitoring and repairing gel dressing containing microspheres, and the application of the wound infection monitoring and repairing gel dressing containing microspheres in preparing a wound treatment consumable. Preferably, the near-infrared light wavelength is 800-1650 nm, the irradiation time is 5-20 min, and the concentration of hydrogen peroxide is 10-100 mM.

[0050] Preferably, the hydrogen peroxide is added dropwise to the wound, and the wound infection monitoring and repairing gel dressing containing microspheres is covered, and then irradiated with near-infrared light.

[0051] The change of pH value is crucial for monitoring the state of the wound, because the change of pH value indirectly reflects the infection of the wound and the remodeling process of the wound tissue. Generally, acidic keratin sebum and sweat gland secretions usually form a slightly acidic keratin layer on the surface of healthy skin, and the pH value is mainly distributed between 4.0 and 6.0. However, in infected wounds, due to the action of microorganisms and the presence of bacterial enzymes, the pH value will gradually deviate to alkaline, and the value is mainly distributed between 7.0 and 9.0.

[0052] Based on this, the polyacrylamide-alginate double network gel is used as the main component of the dressing, on the one hand, the pH value change of the wound surface is observed by using the acid-base indicator to realize the real-time infection monitoring of the wound surface; on the other hand, the molybdenum disulfide gel microspheres in the gel dressing can rapidly decompose low-concentration hydrogen peroxide under the irradiation of near-infrared laser to generate active free radicals to kill pathogenic microorganisms on the wound surface, so that the real-time infection monitoring and precise on-demand antibacterial treatment of the wound surface are realized. At the same time, when the wound is not infected, the over-release of antibacterial drugs will not occur, and the precise on-demand treatment can be carried out according to the infection of the wound.

[0053] Compared with the prior art, the beneficial effects of the technical scheme of the present application are:

[0054] 1. Real-time monitoring of wound infection: the color change of the gel dressing in different pH environments can sensitively reflect the change of the acidity and alkalinity of the wound, which can help to quickly judge whether the wound is infected. When the wound is infected with bacteria, the pH value rises, and the gel color changes from yellow to red.

[0055] 2. Precise control of on-demand antibacterial treatment: the sodium alginate hydrogel microspheres containing molybdenum disulfide nanomaterials can decompose hydrogen peroxide to generate active oxygen (·OH) under near-infrared light irradiation, thereby killing pathogenic microorganisms. The antibacterial experiment results show that only under the condition that low-concentration hydrogen peroxide is combined with the gel dressing and receives near-infrared laser irradiation, the antibacterial ability can be significantly improved. For example, the bacteriostatic rate of Escherichia coli is 94.44%, that of Staphylococcus aureus is 97.80%, that of Pseudomonas aeruginosa is 92.68%, and that of Acinetobacter baumannii is 85.68%.

[0056] 3. Excellent mechanical properties and adaptability: the gel dressing exhibits good tensile deformation ability and can be stretched to more than 3 times the original length without breaking or releasing the wrapped microspheres. This feature makes it particularly suitable for body parts that need frequent movement or deformation (such as elbows, knees, etc.). The loose and porous internal structure of the gel dressing helps to improve the air permeability and the ability to absorb wound exudates, thereby providing a better environment for wound healing.

[0057] 4. Promote wound healing: after using the gel dressing of the present application and cooperating with near-infrared laser irradiation, the healing speed of the wound is significantly accelerated, the infection is effectively controlled, the production of pus is reduced, and the rapid regeneration of the wound tissue is promoted. BRIEF DESCRIPTION OF DRAWINGS

[0058] Figure 1 Take photos for body microscope to observe the particle size difference of molybdenum disulfide gel microspheres prepared by using different wind power;

[0059] Figure 2Observation of the color change of the gel dressing after being placed in physiological saline with different pH values for 10 minutes;

[0060] Figure 3 Scanning electron microscope photos of the surface and fracture surface of the polyacrylamide-alginate double network gel prepared in the present application compared with the polyacrylamide gel;

[0061] Figure 4 Observation of the color change of the gel dressing after being placed in physiological saline with different pH values for 10 minutes;

[0062] Figure 5 Detection of the absorbance of methylene blue oxidized by hydrogen peroxide decomposed by the gel dressing promoted by near-infrared laser;

[0063] Figure 6 Plate colony counting method for testing the efficacy of the gel dressing in inhibiting bacterial growth by decomposing hydrogen peroxide promoted by near-infrared laser;

[0064] Figure 7 Bacterial live and dead fluorescent labeling for observing the bacterial activity (green fluorescence) and death (red fluorescence) of the gel dressing prepared under the condition of low-concentration hydrogen peroxide solution without and with near-infrared laser irradiation;

[0065] Figure 8 Observation of the effect of the gel dressing on the infection of mouse wounds treated under different treatment regimens for different time periods;

[0066] Figure 9 Photographing of mouse wounds for different time periods to observe the effect of the gel dressing in inhibiting wound infection and promoting wound healing by decomposing hydrogen peroxide promoted by near-infrared laser. DETAILED DESCRIPTION

[0067] The present application will be further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present application in any form. Unless otherwise specified, the reagents, methods and devices used in the present application are conventional reagents, methods and devices in the technical field.

[0068] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0069] Example 1

[0070] Preparation of a microsphere-containing wound infection monitoring and repair gel dressing

[0071] Step 1: At room temperature, 0.4 g of molybdenum disulfide nanoparticles was added to 4 mL of deionized water, and a magnetic stirrer was used to stir at a stirring speed of 800 revolutions per minute for 10 minutes. The molybdenum disulfide nanoparticles were fully dispersed in the aqueous solution.

[0072] Second step: 1.5 g of sodium alginate powder was sieved into 10 mL of deionized water and stirred at a stirring rate of 100 rpm for 2 hours. After the sodium alginate was fully dissolved, 100 μL of the molybdenum disulfide solution obtained in the first step was added dropwise to the sodium alginate solution, and then stirred at a stirring rate of 800 rpm for 10 minutes to fully mix the molybdenum disulfide solution and the sodium alginate solution.

[0073] Third step: The molybdenum disulfide sodium alginate solution obtained in the second step was sucked into a syringe, extruded by a micro-injection pump at a flow rate of 0.1 mL / min, and blown by wind at a wind speed of 12.5 m / s into a 1.5% (mass percentage) calcium chloride solution, and left to stand in the solution for 30 minutes. The sodium alginate hydrogel microspheres doped with molybdenum disulfide nanomaterials were obtained by cross-linking with calcium ions in the calcium chloride solution, and then the gel balls were washed with deionized water three times and redispersed in deionized water for use.

[0074] Fourth step: 2.5 g of sodium alginate powder and 13.5 g of acrylamide were dissolved in 100 mL of deionized water after magnetic stirring for 3 hours, and then 8.5 mg of methylene bisacrylamide, 0.1 g of polyacrylamide, 0.002 g of phenol red powder, and 0.15 g of ammonium persulfate powder were added, and the sample was fully mixed and dissolved by stirring at a stirring rate of 500 rpm for 30 minutes using a magnetic stirrer.

[0075] Fifth step: After the mixture obtained in the fourth step was defoamed using an ultrasonic defoaming instrument, 4 g of the sodium alginate hydrogel microspheres doped with molybdenum disulfide nanomaterials prepared in the third step were added to every 10 mL of the mixture. After 15 μL of tetramethyl ethylenediamine was added under magnetic stirring for 5 minutes, 0.04 g of calcium sulfate dihydrate powder was added while stirring through a 100-mesh sieve. After the mixture was stirred uniformly, it was poured into a glass mold with a length of 100 mm, a width of 83 mm, and a height of 1.5 mm, and placed in a 4°C refrigerator for 2 days to allow the mixture to fully cross-link, polymerize, and solidify into a gel. Then the gel was taken out of the glass mold, soaked in deionized water, and washed in a horizontal dehydration shaker for 3 times (10 minutes each time) to remove residual monomers that did not undergo cross-linking. Finally, a wound infection monitoring and treatment gel dressing containing microspheres was obtained.

[0076] Performance detection.

[0077] Example 2

[0078] Preparation of a wound infection monitoring and repair gel dressing containing microspheres

[0079] First step: 0.1 g of molybdenum disulfide nanoparticles was added into 10 mL of deionized water at room temperature, and stirred for 5 minutes at a stirring rate of 100 revolutions per minute using a magnetic stirrer. The molybdenum disulfide nanoparticles were fully dispersed in the aqueous solution.

[0080] Second step: 0.5 g of sodium alginate powder was sieved and added into 20 mL of deionized water, and stirred for 1 hour at a stirring rate of 50 revolutions per minute. After the sodium alginate was fully dissolved, 100 μL of the molybdenum disulfide solution obtained in the first step was added dropwise into the sodium alginate solution, and then stirred for 5 minutes at a stirring rate of 200 revolutions per minute to fully mix the molybdenum disulfide solution and the sodium alginate solution.

[0081] Third step: The molybdenum disulfide sodium alginate solution obtained in the second step was sucked into a syringe, and extruded at a flow rate of 0.02 mL / min through a microsyringe pump and blown by air at a wind speed of 15.5 m / s into a 0.5% (mass percentage) calcium chloride solution, and left to stand in the solution for 50 minutes. The molybdenum disulfide nanomaterial-doped sodium alginate hydrogel microspheres were obtained by cross-linking of calcium ions in the calcium chloride solution, and then the gel spheres were washed three times with deionized water and redispersed in deionized water for use.

[0082] Fourth step: 1 g of sodium alginate powder and 25 g of acrylamide were dissolved in 100 mL of deionized water after magnetic stirring for 3 hours, and then 15 mg of methylene bisacrylamide, 0.05 g of polyacrylamide, 0.05 g of phenol red powder, and 0.3 g of ammonium persulfate powder were further added, and the sample was fully mixed and dissolved by stirring for 5 minutes at a stirring rate of 200 revolutions per minute using a magnetic stirrer.

[0083] Fifth step: After the mixture obtained in the fourth step was defoamed using an ultrasonic defoaming instrument, 8 g of the molybdenum disulfide nanomaterial-doped sodium alginate hydrogel microspheres prepared in the third step was added into every 10 mL of the mixture, 3 μL of tetramethyl ethylenediamine was added under magnetic stirring for 2 minutes, and then 0.1 g of calcium sulfate dihydrate powder was added through a 100-mesh sieve while stirring, and the mixture was poured into a glass mold with a length of 100 mm, a width of 83 mm, and a height of 1.5 mm, and placed in a 4-degree Celsius refrigerator for 3 days to allow the mixture to fully cross-link, polymerize, and solidify into a gel. Then the gel was taken out of the glass mold, soaked in deionized water, and washed in a horizontal dehydration shaker for 3 times (10 minutes each time) to remove the residual monomers that did not undergo cross-linking. Finally, a wound infection monitoring and treatment gel dressing containing microspheres was obtained.

[0084] Performance detection.

[0085] Example 3

[0086] Preparation of microsphere-containing wound infection monitoring and repair gel dressing

[0087] First step: 10 g of molybdenum disulfide nanoparticles were added to 10 mL of deionized water at room temperature, and stirring was performed using a magnetic stirrer at a stirring rate of 1000 revolutions per minute for 20 minutes. The molybdenum disulfide nanoparticles were fully dispersed in the aqueous solution.

[0088] Second step: 12 g of sodium alginate powder was sieved and added to 20 mL of deionized water, and stirring was performed at a stirring rate of 200 revolutions per minute for 3 hours. After the sodium alginate was fully dissolved, 100 μL of the molybdenum disulfide solution obtained in the first step was taken and added dropwise to the sodium alginate solution, and then stirring was performed at a stirring rate of 1000 revolutions per minute for 20 minutes to fully mix the molybdenum disulfide solution and the sodium alginate solution.

[0089] Third step: the molybdenum disulfide sodium alginate solution obtained in the second step was sucked into a syringe, and was extruded through a microsyringe pump at a flow rate of 1 mL / min and blown by air at a wind speed of 12.5 m / s into a 2% (mass percentage) calcium chloride solution, and was left to stand in the solution for 10 minutes. The molybdenum disulfide nanomaterial-doped sodium alginate hydrogel microspheres were obtained by cross-linking of calcium ions in the calcium chloride solution, and then the gel balls were washed three times with deionized water and were redispersed in deionized water for use.

[0090] Fourth step: 4 g of sodium alginate powder and 5 g of acrylamide were dissolved in 100 mL of deionized water after magnetic stirring for 3 hours, and then 5 mg of methylene bisacrylamide, 0.3 g of polyacrylamide, 0.005 g of phenol red powder, and 0.05 g of ammonium persulfate powder were further added, and the sample was fully mixed and dissolved by stirring at a stirring rate of 1000 revolutions per minute using a magnetic stirrer for 50 minutes.

[0091] Fifth step: after the mixture obtained in the fourth step was defoamed using an ultrasonic defoaming instrument, 2 g of the molybdenum disulfide nanomaterial-doped sodium alginate hydrogel microspheres prepared in the third step were added to every 10 mL of the mixture, 20 μL of tetramethyl ethylenediamine was added under magnetic stirring for 2 minutes, and then 0.01 g of calcium sulfate dihydrate powder was slowly added through a 100-mesh sieve while stirring, and after uniform stirring, the mixture was poured into a glass mold with a length of 100 mm, a width of 83 mm, and a height of 1.5 mm, and was left to stand in a 4-degree Celsius refrigerator for 1 day, so that the mixture was fully cross-linked, polymerized, and solidified into a gel. Then the gel was taken out of the glass mold, was soaked in deionized water, and was washed three times (10 minutes each time) in a horizontal shaking bed to remove residual monomers that did not undergo cross-linking. After the washing was completed, a microsphere-containing wound infection monitoring and treatment gel dressing was finally obtained.

[0092] Performance detection.

[0093] Example 4

[0094] Preparation of molybdenum disulfide gel microspheres

[0095] 0.5 g of molybdenum disulfide was dispersed in 4 mL of deionized water, and continuously stirred at 1000 rpm for 10 minutes using a magnetic stirrer. After that, 100 μL of the obtained molybdenum disulfide solution was added dropwise to 5 mL of a 1.5% (mass / volume ratio) sodium alginate solution under stirring at a speed of 800 rpm, and stirring was continued for 10 minutes to obtain a uniform solution. The mixture was then jetted into 3 liters of a 1.5% (mass / volume ratio) calcium chloride solution to crosslink under the wind force of 13.5 m / s at an extrusion flow rate of 0.1 mL / min using a microsyringe pump extrusion device to obtain gel microspheres containing molybdenum disulfide nanomaterials of different particle sizes.

[0096] Examples 5-8

[0097] The technical solutions of Examples 5-8 are similar to those of Example 4, except that the wind forces of Examples 5-6 are 14.5 m / s, 15.5 m / L, 10.5 m / s and 11.5 m / L, respectively.

[0098] Comparative Example 1

[0099] A conventional polyacrylamide gel was prepared as follows: 4.1 mL of deionized water was added to a 30 mL beaker, followed by sequentially adding 30% Acr-Bis (29:1) 3.3 mL, lower gel buffer (4x) 2.5 mL, 10% gel polymerization catalyst 0.1 mL, all purchased from the Biyun Tian Biological Technology Company, and stirring uniformly, then quickly pouring into a 1.5 mm thick electrophoresis gel preparation glass plate sandwich, carefully sucking out the air bubbles, and then injecting deionized water until the top of the sandwich glass plate. After standing for 1 h, a 10% concentration SDS-PAGE gel was obtained.

[0100] Detection method

[0101] 1. Particle size observation

[0102] The particle sizes of the molybdenum disulfide gel microspheres prepared using different wind forces were observed by taking pictures with a stereomicroscope.

[0103] 2. Tensile deformation performance detection of the gel dressing

[0104] The gel dressing prepared in Example 1 was cut into a long strip with a length of 130 mm, a width of 23 mm and a thickness of 1.5 mm, and the two ends of the gel were stretched by hand.

[0105] 3. Observation of the micro-morphology structure of the gel dressing

[0106] The surface and fracture surface of the gels of Example 2 and Comparative Example 1 were observed by scanning electron microscopy.

[0107] 4. Observation of the indicating performance of the gel dressing to pH change

[0108] The gel dressing prepared in Example 3 was placed in phosphate buffer with different pH values for 10 minutes and then removed. The gel showed a clear color change.

[0109] 5. Detection of active radicals

[0110] The production of active radicals OH in the solution was detected using methylene blue reagent. The absorbance of methylene blue oxidized by hydrogen peroxide promoted by near-infrared laser was detected.

[0111] 6. Test of antibacterial efficacy

[0112] E. coli (gram-negative bacteria), P. aeruginosa (gram-negative bacteria), A. baumannii (gram-negative bacteria), and S. aureas (gram-positive bacteria) were used as representative model bacteria to test the antibacterial performance of the prepared gel dressing. First, single colonies of the above four model bacteria were incubated in 5 mL sterile bacterial broth medium (purchased from Aladdin) respectively. The shaking speed of the bacterial culture shaker was 250 revolutions per minute, and the bacteria were kept shaking at 37°C overnight to obtain logarithmic phase bacterial suspension. Subsequently, the bacterial suspension was transferred into a new test tube, and the sterile bacterial broth medium was used for dilution at a dilution ratio of 10 times, and the bacterial concentration was diluted to 1 x 10 6 CFU / mL. Then, the four bacterial suspensions were aliquoted into centrifuge tubes, centrifuged at 4000 revolutions per minute for 5 minutes, and the bacterial culture medium was removed, and the bacteria were resuspended in sterile phosphate buffer to a bacterial concentration of 1 x 10 6CFU / mL. Subsequently, 1 mL of the bacterial suspension was added to each well of a 24-well cell culture plate and further manipulated to form the following six groups: (I) bacteria alone, (II) bacteria + H2O2 (20 mM H2O2), (III) bacteria + H2O2 + near-infrared laser irradiation (20 mM H2O2, 1.5 W / cm2at 808 nm for 10 min), (IV) bacteria + gel dressing, (V) bacteria + H2O2 + gel dressing, and (VI) bacteria + H2O2 + gel dressing + near-infrared laser irradiation. After incubation for 10 min or near-infrared laser irradiation for 10 min, the above groups were diluted twice with sterile phosphate-buffered saline at a dilution factor of 10, and 100 μL of the diluted bacterial suspension was spread on the surface of a sterile bacterial agar plate. The plates were incubated at 37°C for 24 h in a bacterial incubator, removed, and photographed. Colony counts were then performed and analyzed statistically.

[0113] 7. Fluorescent staining observation

[0114] Commercially available bacterial live / dead fluorescent staining kits (purchased from Invitrogen) were used to stain and observe bacteria treated with low concentrations of hydrogen peroxide solution (20 mM) with or without the gel dressing of Example 1 and with or without near-infrared laser irradiation (1.5 W / cm2for 10 min).

[0115] 8. Monitoring ability for wound infection

[0116] Full-thickness skin defect models and S. aureus-infected full-thickness skin defect models were constructed in mice. C57 male mice (8 weeks old) were anesthetized by continuous inhalation of 1.5% isoflurane using an animal anesthetizing machine. The back hair of the mice was then removed using a hair clipper, and the back was depilated using commercially available depilatory cream (Veet, Reckitt Benckiser). Before surgery, the skin surface was disinfected with iodophor solution and 75% alcohol, respectively. Subsequently, a circular full-thickness skin with a diameter of 10 ± 2 mm was excised from the back of each mouse using a scalpel and surgical scissors, and the wound was rinsed with sterile saline to remove exuded blood. The mice were then grouped, and sterile phosphate-buffered saline was added to the wound site of the mice in the non-infected group (negative control group), and 20 μL of a sterile phosphate-buffered saline-diluted S. aureus suspension (10 6 CFU / mL) was added to the wound site of the mice in the infected wound group. The wounds of the mice in the two groups were covered with the gel dressing prepared in Example 1, and the wounds were sealed with a 3M polyacrylamide film. The color change of the gel dressing was observed after 1 day, 2 days, and 3 days, respectively.

[0117] 9. Inhibition of wound infection

[0118] The wounds of mice were photographed at different time periods to observe how near-infrared laser promoted the gel dressing to decompose hydrogen peroxide, inhibiting wound infection and promoting wound healing.

[0119] Test results

[0120] 1. Particle size observation

[0121] The particle size of the molybdenum disulfide gel microspheres in Examples 4 to 8 is as follows: Figure 1 As shown. When the wind speed at the needle tip is 10.5m / s (Example 7), the average particle size of the gel microspheres is 951.4 microns. When the wind speed at the needle tip is 11.5m / s (Example 8), the average particle size of the gel microspheres is reduced to 846.97 microns. Within a certain range, the particle size of the prepared gel gradually becomes smaller as the wind speed increases. When the wind speed reaches 13.5m / s (Example 4), the average particle size of the gel microspheres is reduced to 546.30 microns; when the wind speed reaches 14.5m / s (Example 5), the average particle size of the gel microspheres is reduced to 389.25 microns. When the wind speed reaches 15.5m / s (Example 6), the size of the gel ball particle size no longer shows a linear relationship with the increase in wind force. If the particle size is too small, the molybdenum disulfide content in the microspheres is less, which is not conducive to the subsequent rapid decomposition of hydrogen peroxide to kill bacteria under near-infrared light irradiation. If the particle size is too large, it will affect the visual observation of the color change of the gel and affect the cross-linking preparation of the gel. The concentration of alginate, the extrusion rate of the microinjection pump, and the wind force at the needle tip of the present invention are matched within appropriate intervals to obtain gel microspheres with smaller particle sizes, which are more conducive to the performance of the gel dressing.

[0122] 2. Testing of tensile deformation properties of gel dressing

[0123] like Figure 2 As shown in the figure, the tensile deformation capacity of the gel dressing prepared in Example 1 can reach more than 3 times of its original length, and compared with conventional wound gel dressings, it has excellent deformation and stretching capacity. In addition, the black molybdenum disulfide gel microspheres wrapped in the gel also show excellent deformation capacity as the gel is stretched. During the stretching process, the molybdenum disulfide gel microspheres and the gel dressing show good adaptability, and do not separate from the gel due to the significant deformation of the gel dressing. This shows that the prepared gel dressing exhibits excellent tensile deformation capacity, making it well suitable for the bending and straightening deformation of the wound, such as the large-scale bending deformation requirements of the elbow and knee joints, so that the gel dressing can adapt well to the deformation and displacement of the wound.

[0124] 3. Observation of the microstructure of gel dressing

[0125] likeFigure 3 As shown, the polyacrylamide-alginate double-network hydrogel dressing containing molybdenum disulfide sodium alginate gel balls prepared in Example 2 has obvious differences from the conventional polyacrylamide gel of Comparative Example 1. The surface structure of the polyacrylamide gel of Comparative Example 1 is relatively dense, and the pore structure is relatively small. In contrast, the surface of the gel dressing prepared in Example 2 presents a loose gel pore structure, and the number of pores is also significantly improved. When the polyacrylamide gel of Comparative Example 1 and the prepared gel dressing are broken and the fracture cross-section of the gel is observed, it can be found that the gel cross-linking inside the fracture cross-section of the polyacrylamide gel of Comparative Example 1 is relatively dense, and the number of pores in the gel is relatively small, while the fracture cross-section of the gel dressing of Example 2 is significantly different. The gel fracture surface presents a clear honeycomb sponge structure, and not only on the surface of the gel, but also in the interior of the gel, it presents an extremely high porosity. Therefore, with the help of a loose porous structure, the gel dressing prepared by the present invention will have a more excellent deformation and stretching ability, better air permeability and the ability to absorb and drain wound exudate.

[0126] 4. Observation of the gel dressing's performance in indicating pH changes

[0127] like Figure 4 As shown, as the pH value of the solution gradually increases from 4.5 to 9.0, the color of the gel prepared in Example 3 gradually changes from yellow to brown. When the solution becomes alkaline, the gel gradually changes from brown to red, and finally reaches deep red. This demonstrates that the gel dressing prepared by the present invention can sensitively respond to changes in pH, and the obvious color change of the gel reflects the acidity or alkalinity of the pH environment in which the gel is located.

[0128] 5. Detection of active free radicals

[0129] Methylene blue is a commonly used hydroxyl radical scavenger. Its working principle is that methylene blue is oxidized by OH to generate colorless methylene white, which causes the color of the methylene blue solution to gradually fade from blue to colorless. Figure 5The UV absorbance curve of the solution sample shown in 400-800 nm indicates that the 20 mM hydrogen peroxide solution without methylene blue cannot detect obvious absorbance signal. The solution of methylene blue prepared by the gel dressing of Example 1 shows the highest absorbance value at the same wavelength, indicating that the solution of methylene blue has no obvious active free radical ·OH, and the blue methylene blue solution does not fade obviously. When the gel dressing of Example 1 is irradiated by 808 nm near-infrared laser with a power of 0.5 W / cm2for 10 minutes, the absorbance value of the methylene blue solution decreases, indicating that the production of active free radical ·OH in the solution increases, and the methylene blue is oxidized and faded by ·OH. Under the same conditions, when the power of the irradiated laser is increased from 0.5 W / cm2to 1.0 W / cm2, 1.5 W / cm2, and 2.0 W / cm2, respectively, it can be found that the stronger the laser irradiation intensity, the stronger the ability of the prepared gel dressing to decompose hydrogen peroxide to produce ·OH. It is proved that the use of near-infrared laser irradiation can accurately control and improve the ability of the prepared gel dressing to decompose hydrogen peroxide to produce ·OH.

[0130] 6. Anti-bacterial efficacy test

[0131] As shown in Figure 6 , the anti-bacterial ability of the diluted hydrogen peroxide solution (20 mM) is greatly reduced. After counting the bacterial colonies of the bacterial culture dish and comparing with the blank control group, it is found that more than 71.93% of Escherichia coli, more than 76.80% of Staphylococcus aureus, more than 85.37% of Pseudomonas aeruginosa, and more than 78.78% of Acinetobacter baumannii survive. On this basis, the addition of near-infrared laser irradiation cannot significantly enhance the anti-bacterial efficacy of low-concentration hydrogen peroxide solution. Similarly, when only the gel dressing prepared in Example 1 is used, or the gel dressing is used in combination with low-concentration hydrogen peroxide, no obvious bacteriostatic effect can be observed. Only when low-concentration hydrogen peroxide is added and the gel dressing is used in combination with 808 nm near-infrared laser irradiation, a significant bacteriostatic effect can be observed. Among them, the inhibition rate of Escherichia coli reaches 94.44%, that of Staphylococcus aureus is 97.80%, that of Pseudomonas aeruginosa is 92.68%, and that of Acinetobacter baumannii is 85.68%, showing good anti-bacterial efficacy, indicating that the use of near-infrared laser irradiation can significantly improve the anti-bacterial ability of the prepared gel dressing.

[0132] 7. Fluorescent staining observation

[0133] Under the confocal fluorescence microscope, SYTO-9 green fluorescent nucleic acid dye in the bacterial live / dead fluorescent staining kit can successfully label the bacteria and present bright green fluorescence. At the same time, another iodinated propyl red fluorescent nucleic acid dye (PI) is also present in the kit. Since PI can only penetrate the damaged bacterial membrane, and at the same time, the penetration of PI and the insertion staining of bacterial nucleic acid will reduce the green fluorescence signal of SYTO-9 dye, therefore, according to this principle, the bacteria with intact membrane structure and good growth state present bright green fluorescence under the labeling of the kit, while the bacteria with damaged membrane present red fluorescence. As shown in Figure 7 , under the condition of low concentration hydrogen peroxide solution (20mM), only using gel dressing cannot damage the bacteria, and almost all the bacteria present bright green fluorescence in the four tested bacteria. On the contrary, after using gel dressing combined with near-infrared laser irradiation, except for a few green fluorescent points observed in Acinetobacter baumannii, the remaining three bacteria all present obvious red fluorescence, indicating that most of the bacteria are significantly damaged.

[0134] 8. Monitoring ability of wound infection

[0135] As shown in Figure 8 , after placing the gel dressing on the wound of mice for 1 day, the non-infected group (normal saline group) without the addition of Staphylococcus aureus presents yellow-brown color, and the remaining gel dressing groups with the addition of Staphylococcus aureus, H2O2+gel dressing group, H2O2+gel dressing+near-infrared laser group present pink color. By observing the color change of the gel, it is found that the pH value of the non-infected normal saline group wound is distributed in the range of 6.0-7.0, and the wound of mice infected with Staphylococcus aureus generally presents alkaline, and the pH value is distributed in the range of 7.5-8.5. This indicates that by observing the color change of the wound gel, it can be effectively distinguished whether the wound is infected.

[0136] 9. Inhibition of wound infection

[0137] As shown in Figure 9As shown, the dressing changed from the initial bright yellow to magenta after 1 day of S. aureus infection, indicating that the wound of the mouse gradually turned to alkaline under the continuous infection of bacteria. Meanwhile, under the same conditions of applying the gel, there was a significant difference in the healing of the wound of the mouse without near-infrared laser irradiation and the wound of the mouse with near-infrared laser irradiation. The wound of the mouse without near-infrared laser irradiation showed obvious signs of wound infection on the third day after S. aureus infection, and the wound showed obvious yellow pus, indicating that the wound had been infected and inflamed. Moreover, due to the failure to inhibit the infection in time, the inflammation of the wound gradually increased with time, which significantly hindered the healing and repair of the wound. In contrast, the inflammation of the wound of the mouse with near-infrared laser irradiation was significantly inhibited on the second day after treatment, and the healing rate of the wound was significantly better than that of the mouse without near-infrared laser irradiation under the same healing time, indicating that near-infrared laser can promote the decomposition of hydrogen peroxide in the gel dressing to inhibit wound infection and promote wound healing.

[0138] Obviously, the above embodiments of the present application are only examples for clearly illustrating the present application, and are not intended to limit the embodiments of the present application. Based on the above description, other different forms of changes or variations can be made by those skilled in the art. Here, it is not necessary and impossible to exhaust all the embodiments. Any modification, equivalent replacement and improvement made within the spirit and principle of the present application shall be included in the protection scope of the claims of the present application.

Claims

1. A method of preparing a microsphere-containing wound infection monitoring and repair gel dressing, characterized by, The method comprises the following steps: S1, adding a molybdenum disulfide solution to a sodium alginate solution, and stirring to obtain a molybdenum disulfide sodium alginate solution; S2, adding the molybdenum disulfide sodium alginate solution to a calcium chloride solution, and standing to crosslink to obtain molybdenum disulfide gel microspheres; S3, dissolving sodium alginate and acrylamide in water by stirring, and adding methylene bisacrylamide, polyacrylamide, an acid-base indicator, and ammonium persulfate, and fully mixing and dissolving to obtain a mixed solution; S4, adding the molybdenum disulfide gel microspheres to the mixed solution, and adding tetramethylethylenediamine and calcium sulfate dihydrate in sequence under continuous stirring, and stirring uniformly; S5, standing to solidify, removing impurities, and obtaining a wound infection monitoring and repairing gel dressing containing the microspheres. In S2, the molybdenum disulfide sodium alginate solution is injected into the calcium chloride solution at a flow rate of 0.02-1.0 mL / min under external force; the speed generated by the external force is 10.5-15.5 m / s.

2. The method of claim 1, wherein the microsphere-containing wound infection monitoring and repair gel dressing is prepared by the steps of: The concentration of the molybdenum disulfide solution in S1 is 0.01-1 g / mL, and the concentration of the obtained sodium alginate solution is 0.025-0.6 g / mL.

3. The method of claim 1, wherein the microsphere-containing wound infection monitoring and repair gel dressing is prepared by the steps of: The volume ratio of the molybdenum disulfide solution to the sodium alginate solution added in S1 is 0.1:5-20.

4. The method of claim 1, wherein the microsphere-containing wound infection monitoring and repair gel dressing is prepared by the steps of: The mass percentage of the calcium chloride solution in S2 is 0.5%-2.0%.

5. The method of claim 1, wherein the microsphere-containing wound infection monitoring and repair gel dressing is prepared by the steps of: In the mixed solution of S3, the concentration of sodium alginate is 0.01-0.04 g / mL, the concentration of acrylamide is 0.05-0.25 g / mL, the concentration of methylene bisacrylamide is 0.05-0.15 mg / mL, the concentration of polyacrylamide is 0.5-3 mg / mL, the concentration of the acid-base indicator is 0.05-0.5 mg / mL, and the concentration of ammonium persulfate is 0.5-3 mg / mL.

6. The method of claim 1, wherein the microsphere-containing wound infection monitoring and repair gel dressing is prepared by the steps of: After uniform stirring in S4, the concentration of the molybdenum disulfide gel microspheres is 0.2-0.8 g / mL, the concentration of tetramethylethylenediamine is 0.3-2 μL / mL, and the concentration of calcium sulfate dihydrate is 0.001-0.01 g / mL.

7. A microsphere-containing wound infection monitoring and repair gel dressing, characterized in that, The method is prepared by the method of any one of claims 1-6.

8. The microsphere-containing wound infection monitoring and repair gel dressing of claim 7, wherein, The microspheres are molybdenum disulfide gel microspheres, and the diameter is 300-1000 μm.

9. Use of a microsphere-containing wound infection monitoring and repair gel dressing according to claim 8, characterised in that, The wound infection monitoring and repairing gel dressing containing the microspheres is applied to the preparation of a wound treatment consumable.

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