Active and passive synergistic antibacterial and healing-promoting dressings, their preparation methods and applications

Abstract

This invention discloses a synergistic active and passive antibacterial and healing-promoting dressing, its preparation method, and its application, belonging to the field of burn dressing technology. The technical solution is as follows: the dressing is a dopamine-sodium alginate-zinc ion composite functional dressing based on short-cycle, cyclic cross-linking coating technology. The preparation method includes the following steps: 1) compounding dopamine with polyurethane sponge; 2) short-cycle, cyclic cross-linking of sodium alginate and zinc ions to form a gelled coating; 3) drying the cross-linked dressing to obtain the synergistic active and passive antibacterial and healing-promoting dressing. This invention obtains a synergistic active and passive antibacterial dressing through short-time, multiple-cycle cross-linking technology, achieving a highly efficient antibacterial effect that integrates active and passive antibacterial principles. It also achieves multiple functions, including absorbing exudate and promoting healing, and is suitable for the production and application of dressings for open wounds such as burns and scalds.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of open wound dressings for burns and scalds, and particularly relates to a synergistic antibacterial and healing-promoting dressing, and a preparation method and application thereof. BACKGROUND

[0002] Burns and scalds refer to skin tissue damage caused by heat, electric current, chemicals, lasers, and radiation, and are common accidental injuries in life and production. Improper treatment of the wound at the burn site can be life-threatening. Currently, wound infection is one of the main causes of death in burn patients. In clinical work, wound dressing change is one of the most effective ways to control infection. The traditional method is to change dressing with gauze combined with various antibacterial drugs. As the most common medical dressing, gauze has the advantages of protecting the wound, good absorbability, wide source, simple preparation, and low price. However, the disadvantages are also prominent: it cannot keep the wound moist, hinders tissue cell epithelialization, delays wound healing, affects healing, causes pain during dressing change, and makes it difficult to fully drain the exudate from the infected burn wound, which can easily lead to the accumulation of exudate and aggravate local infection.

[0003] Infection is the most common complication after burns. Skin is the main barrier against pathogens in the environment, and burns destroy the integrity of the skin, allowing the invasion of infection sources (mainly bacteria, but also yeast, fungi, and viruses) to cause skin tissue and systemic infection, and in severe cases, sepsis, which is the main cause of death in burn patients. Due to the diversity of infections and the changing drug resistance of pathogenic bacteria, antibiotic therapy is facing challenges such as limited efficacy and misuse. Therefore, there is an urgent need for new anti-infection wound dressings with great potential.

[0004] In recent years, composite materials combining active and passive antibacterial strategies have gradually become a research hotspot and have been reported in the field of environmental science, but there is no report on active and passive synergistic antibacterial dressings for skin wounds. The existing antibacterial wound dressing technology is mainly divided into two categories: active sterilization strategy (such as silver ions, antibiotics, etc.) and passive bacteria prevention strategy (such as polymer surface modification to form physical bacteria prevention); metal ions can provide good antibacterial effect, and the commonly used antibacterial ion is silver ion, but it has the disadvantages of high cytotoxicity, high cost, and limited clinical application. Chinese invention patent CN109010898A discloses a chitosan-based gauze dressing loaded with metal ions, the metal ions loaded on the gauze dressing can be added one or more as needed, and the gauze dressing can be directly applied to the wound surface or applied after being stacked layer by layer. The concentration of the loaded metal ions can be adjusted by changing the concentration of the prepared metal ion solution; Chinese invention patent CN114456437A discloses an antibacterial polyurethane dressing modified by a polydopamine coating layer, a preparation method and application thereof, and the preparation method comprises the following steps: adding dopamine, sulfobetaine methacrylate, H2O2 and CuSO4 into a Tris solution in sequence, mixing uniformly to obtain a co-deposition solution; immersing a sponge substrate in the co-deposition solution and performing oscillation treatment, taking out and drying to constant weight to obtain the antibacterial polyurethane dressing modified by the polydopamine coating layer. The co-deposition reaction of dopamine and sulfobetaine methacrylate is initiated by CuSO4 / H2O2, and the polyurethane sponge is co-deposited and modified, so that the prepared polyurethane dressing exhibits broad-spectrum, long-acting and stable antibacterial performance. However, the dressing with a single antibacterial strategy has certain limitations in actual application, such as excessive cytotoxicity, limited antibacterial effect, and short-lasting curative effect. Therefore, how to develop a wound dressing with active and passive antibacterial properties is a problem to be solved by those skilled in the art. SUMMARY

[0005] The present application provides a kind of active and passive synergistic antibacterial and promote healing dressing and its preparation method and application, active and passive synergistic antibacterial strategy composite function dressings, can effectively prevent and control burn and scald wound infection and promote healing.The dressing is constructed by dopamine (DOP), sodium alginate (SA) and zinc ion (Zn 2+ )It realizes multiple functions such as efficient antibacterial, absorption of exudate and promotion of healing by active and passive synergistic antibacterial mechanism and short-time crosslinking technology, and is suitable for dressing production for burn and scald and other open wound related.

[0006] The technical scheme of the present application is as follows:

[0007] In a first aspect, an active and passive synergistic antibacterial and healing-promoting dressing is disclosed, which is a dopamine-sodium alginate-zinc ion composite functional dressing. The dressing is a modified hydrogel coating polyurethane dressing formed by modifying dopamine with sodium alginate and zinc ions.

[0008] Preferably, the dressing has excellent passive adhesion and antibacterial performance and zinc ion release active antibacterial performance; the passive antibacterial efficiency can reach 80±1.2%, and the antibacterial rate of the active and passive synergistic antibacterial wound dressing can reach more than 99%.

[0009] Preferably, the dressing has good mechanical properties and liquid management ability, the liquid absorption speed of the dressing is more than twice that of traditional polyurethane sponge PU, and has excellent water absorption and water retention performance, which can effectively control wound exudate.

[0010] Preferably, the dressing uses polyurethane sponge as the substrate, and through in-situ polymerization of dopamine and polyurethane sponge, a passive antibacterial layer with hydrophilicity and good water absorption is formed; then short-period and cyclic cross-linking treatment is performed on the sodium alginate and zinc ion solution to form a zinc ion-containing hydrogel coating, thereby obtaining a modified polyurethane dressing.

[0011] In a second aspect, a preparation method of the active and passive synergistic antibacterial and healing-promoting dressing is disclosed, which comprises the following steps:

[0012] 1) Complexing dopamine with polyurethane sponge: mixing polyurethane sponge with dopamine solution and performing self-polymerization in an alkaline environment to form a polydopamine coating;

[0013] 2) Sodium alginate-zinc ion gelation: soaking the dopamine-modified polyurethane sponge in a sodium alginate and zinc ion solution for short-time and multiple cross-linking to form a zinc-containing sodium alginate hydrogel coating;

[0014] 3) Freeze-drying the cross-linked dressing to obtain the active and passive synergistic antibacterial and healing-promoting dressing.

[0015] Preferably, the concentration of the dopamine solution in step 1) is 2±0.5 mg / mL, and the alkaline environment in step 1) is a tris(hydroxymethyl)aminomethane hydrochloride (Tris-HCl) buffer.

[0016] Preferably, the concentration of the sodium alginate in step 2) is 0.01-2 g / mL, and the concentration of the zinc ion is 8-40 mg / mL.

[0017] Preferably, the short-time and multiple cross-linking time in step 2) is 0.5-3 min, and the cross-linking frequency is 2-6 times.

[0018] In a third aspect, the application discloses an application of the active and passive synergistic antibacterial and healing-promoting dressing in the preparation of an open wound-related dressing.

[0019] Sodium alginate, as a natural polymer, has a negative charge characteristic, and the carboxyl group (-COO -) can form ionic bonds with metal ions, thereby constructing a network structure and promoting the formation of a gel. In this cross-linking process, zinc ions (Zn 2+ ) play a crucial role. Not only do they interact with the carboxyl groups of alginate to form stable complexes, but their selective action also significantly enhances the mechanical properties and stability of the gel.

[0020] Instant multiple cross-linking technology: Unlike traditional methods that solidify into gels after a single cross-linking reaction, instant multiple cross-linking involves introducing zinc ions using an instant multiple technique during gel formation. This aims to increase the cross-linking density of the gel network, thereby constructing a more stable three-dimensional network structure. Precise control can be achieved by flexibly adjusting the concentration of zinc ions or the reaction time.

[0021] Instant multiple cross-linking has the following advantages:

[0022] Improved gel strength: Instant multiple cross-linking can enhance the structural stability of the gel, improve its mechanical strength, and enable it to withstand more stress, adapting to different application requirements.

[0023] Controllability: By adjusting the number of cross-linking times and reaction conditions, the structure and properties of the gel, such as swelling, mechanical strength, and slow-release ability, can be precisely controlled.

[0024] Improved metal ion release rate: Due to the increased cross-linking density, the pore structure of the gel is more uniform, and the release rate of zinc ions can be effectively controlled, thereby improving the bioavailability and therapeutic effect of the antibacterial factor.

[0025] Improved stability: Multiple cross-linking enhances the network structure of the gel and maintains the integrity of the sponge pores after freeze-drying. This reduces the degradation rate of the gel during use and prolongs its use time in vivo and in vitro.

[0026] Active and passive synergistic antibacterial mechanism: Dopamine forms a polydopamine (PDA) coating through self-polymerization, creating a physical barrier on the wound surface and adhering bacteria to the sponge, preventing bacterial invasion and playing a passive antibacterial role. Zinc ions, on the other hand, exert an antibacterial effect through an active release mechanism, effectively killing bacteria. The synergistic effect of the two significantly enhances the antibacterial ability of the dressing.

[0027] Promoting wound healing: The dressing has strong antibacterial properties and effective absorption of wound exudate, combined with pro-healing factors to improve the rate of wound healing. In addition, the dressing has advantages such as promoting collagen deposition and reducing inflammatory infiltration.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] 1.The dressing prepared by the present application has high-efficiency antibacterial performance, and through the modification of dopamine and the combination with sodium alginate and zinc ions, a dressing with a synergistic antibacterial mechanism of active and passive is successfully constructed. The polydopamine coating formed by dopamine can form a physical barrier on the wound surface, effectively adhere and prevent bacterial invasion, and the active release of zinc ions further enhances the antibacterial effect. Experiments prove that the dressing has a bacteriostatic rate of more than 99% on Staphylococcus aureus (S.aureus) and Escherichia coli (E.coli), and can continuously kill bacteria for 11 days and 9 days, significantly improving the anti-infection ability, and has an 80% ± 1.2% antibacterial ability, realizing the protection ability of short-term blocking and long-term sterilization.

[0030] 2.The dressing of the present application adopts polyurethane sponge as the substrate, and through in-situ polymerization of dopamine and polyurethane sponge, a passive antibacterial layer with hydrophilicity and good water absorption is formed. The preparation method of the present application adopts the instant multiple cross-linking treatment of sodium alginate and zinc ions to form a zinc ion-containing hydrogel coating, and the dressing has high porosity, good mechanical properties and excellent elasticity, and is suitable for long-term use in the treatment of burn and scald wounds.

[0031] 3.The dressing of the present application has excellent liquid management ability, the liquid absorption speed is twice that of traditional polyurethane (PU), and has good water retention performance, which can effectively control wound exudate and provide a drier and cleaner healing environment for the wound.

[0032] 4.The dressing of the present application also has a significant effect of promoting wound healing. In a rat scald model, the dopamine-zinc alginate dressing can accelerate the wound healing process and improve the wound healing rate by 27.5%, and performs excellently in promoting microvascular generation, collagen deposition and inflammation control. Its strong antibacterial ability can reduce wound infection and provide strong guarantee for wound healing. At the same time, the dressing can effectively absorb wound exudate, reduce the burden of the wound, promote collagen deposition and reduce the degree of inflammatory infiltration, thereby accelerating the wound healing process.

[0033] 5.The preparation method of the dressing of the present application is simple and easy to operate, and the raw materials required are widely available and low in cost. Through the steps of compounding dopamine with polyurethane sponge, gelation of sodium alginate-zinc ions and freeze-drying treatment, a dressing with excellent performance of synergistic antibacterial and healing promotion can be prepared, which provides strong support for the wide application and industrialization of the dressing.

[0034] 6.The active and passive synergistic antibacterial and healing promotion dressing provided by the present application is suitable for burn and scald wounds and other open wounds, which not only can efficiently prevent and control wound infection, but also can promote wound healing and improve the life quality of patients. At the same time, the preparation method is simple and easy to popularize, and has high practical application value and broad market prospect. BRIEF DESCRIPTION OF DRAWINGS

[0035] Figure 1 Figure 1 is a surface morphology chart of the polyurethane (PU) and dopamine-zinc alginate blended sponge (APBM-L) prepared in accordance with the present application and Comparative Example 3, and the dopamine-zinc alginate dressing (APBM) prepared in accordance with Example 1.

[0036] Figure 2 Figure 2 is a Fourier transform infrared spectroscopy (FTIR) test chart of the active and passive synergistic antibacterial and healing-promoting dressing (APBM) prepared in accordance with Example 1 of the present application, the polyurethane (PU), the DPU prepared in accordance with Comparative Example 1, and the ZnPU prepared in accordance with Comparative Example 2.

[0037] Figure 3 Figure 3 is a water absorption rate result chart of the polyurethane sponge PU and the APBM dressing prepared in accordance with Example 1 of the present application.

[0038] Figure 4 Figure 4 is a water retention rate determination result chart of the polyurethane sponge PU, the APBM prepared in accordance with Example 1, and the APBM-L prepared in accordance with Comparative Example 3 of the present application.

[0039] Figure 5 Figure 5 is a permeability and porosity determination result chart of the active and passive synergistic antibacterial and healing-promoting dressing (APBM) prepared in accordance with Example 1 of the present application and the PU.

[0040] Figure 6 Figure 6 is a stress-strain test chart of the dopamine-zinc alginate dressing APBM prepared in accordance with Example 1 of the present application and the PU.

[0041] Figure 7 Figure 7 is a result chart of the in vitro antibacterial performance test of the PU, the DPU prepared in accordance with Comparative Example 1, the ZnPU prepared in accordance with Comparative Example 2, and the APBM prepared in accordance with Example 1 of the present application.

[0042] Figure 8 Figure 8 is a bacterial ring experiment result chart of the dressing PU, the DPU prepared in accordance with Comparative Example 1, the ZnPU prepared in accordance with Comparative Example 2, and the APBM prepared in accordance with Example 1 of the present application.

[0043] Figure 9 Figure 9 is a bacteria blocking test result chart of the PU, the DPU prepared in accordance with Comparative Example 1, the ZnPU prepared in accordance with Comparative Example 2, and the APBM prepared in accordance with Example 1 of the present application.

[0044] Figure 10 Figure 10 is a biological safety detection (cytotoxicity, hemolyticity) result chart of the dressing prepared in accordance with Example 1 of the present application.

[0045] Figure 11 Figure 11 is a wound healing effect comparison chart of the APBM prepared in accordance with Example 1 of the present application, the DPU prepared in accordance with Comparative Example 1, and the PU in a rat burn and scald model.

[0046] Figure 12 Figure 1 is a histogram showing the histological index analysis of the APBM prepared in Example 1 of the present application, the DPU prepared in Comparative Example 1, and the PU dressing in a rat burn and scald model.

[0047] Figure 13 Figure 2 is a surface morphology diagram of the dressing prepared in Comparative Example 4 of the present application.

[0048] Figure 14 Figure 3 is a surface morphology diagram of the dressing prepared in Comparative Example 5 of the present application. DETAILED DESCRIPTION

[0049] In order to enable persons skilled in the art to better understand the technical solutions in the present application, the technical solutions in the embodiments of the present application will be clearly and completely described below. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments.

[0050] Example 1

[0051] The preparation method of the active-passive synergistic antibacterial and healing-promoting dressing comprises the following steps:

[0052] 1) Complexing of dopamine and polyurethane sponge: The medical polyurethane sponge PU is cut into a cylinder with a diameter of 10 mm and a height of 5 mm, washed and dried, and subjected to ultraviolet sterilization. At the same time, Tris-HCl buffer solution (10 mmol / L, pH = 8.5) is prepared. 2 mg of dopamine hydrochloride (Dop-HCl) powder is added to the Tris-HCl buffer solution to maintain a dopamine hydrochloride concentration of 2 mg / mL. After dissolution, the dried sponge is placed in a wide-mouth bottle and placed in a 37℃ shaking bed for reaction for 12 hours. Then, the sponge is washed with deionized water and anhydrous ethanol alternately for 3 times, and then freeze-dried to obtain polyurethane sponge (DPU) with a polydopamine (PDA) coating.

[0053] 2) Sodium alginate-zinc ion gelation:

[0054] A. The sponge with a polydopamine coating (DPU) is immersed in a sodium alginate solution (0.01 g / mL, viscosity of 80 mPa·s) for 5 minutes, and the residual sodium alginate on the surface of the sponge is squeezed off gently;

[0055] B. After being immersed in a zinc chloride solution (12 mg / mL) for 10 seconds, the sponge is again immersed in the prepared sodium alginate solution (0.01 g / mL, viscosity of 80 mPa·s) for 10 seconds;

[0056] C. Repeat step B) three times, then put into zinc chloride solution (12 mg / mL) and put into 37℃ shaker for 2 hours, freeze-dried to get dopamine-zinc alginate dressing, i.e. the active and passive synergistic antibacterial and healing-promoting dressing, hereinafter referred to as (APBM).

[0057] This embodiment carries out short-time multiple cross-linking to form a sodium alginate hydrogel coating containing zinc, which not only provides active antibacterial effect, but also optimizes the water absorption performance and flexibility of the dressing.

[0058] Example 2

[0059] The preparation method of the active and passive synergistic antibacterial and healing-promoting dressing comprises the following steps:

[0060] 1) Complexing of dopamine and polyurethane sponge: Cut medical polyurethane sponge PU into a cylinder with a diameter of 10 mm and a height of 5 mm, wash and dry, and perform ultraviolet sterilization. At the same time, prepare Tris-HCl buffer solution (10 mmol / L, pH = 8.5). Add 1.5 mg of dopamine hydrochloride (Dop-HCl) powder to the Tris-HCl buffer solution to maintain a dopamine hydrochloride concentration of 1.5 mg / mL. After dissolving, place the dry sponge in a wide-mouth bottle and react in a 37℃ shaker for 12 hours. Then wash with deionized water and anhydrous ethanol alternately for 3 times, and freeze-dry to obtain a sponge with polydopamine coating (DPU).

[0061] 2) Sodium alginate-zinc ion gelation:

[0062] A. Immerse the sponge with polydopamine coating (DPU) in sodium alginate solution (0.01 g / mL, viscosity 80 mPa·s) for 5 minutes, and gently squeeze the residual sodium alginate on the surface of the sponge;

[0063] B. Immerse in zinc chloride solution (20 mg / mL) for 10 seconds, then immerse in prepared sodium alginate solution (0.01 g / mL, viscosity 80 mPa·s) for 10 seconds;

[0064] C. Repeat step B) three times, then put into zinc chloride solution (12 mg / mL) and put into 37℃ shaker for 2 hours, freeze-dried to get dopamine-zinc alginate sponge, i.e. the active and passive synergistic antibacterial and healing-promoting dressing.

[0065] Comparative Example 1

[0066] The preparation method of passive antibacterial dressing polydopamine coated sponge (DPU) is as follows: the medical polyurethane sponge PU is cut into a cylinder with a diameter of 10 mm and a height of 5 mm, washed and dried, and subjected to ultraviolet sterilization. Meanwhile, a Tris-HCl buffer solution (10 mmol / L, pH = 8.5) is prepared. 2 mg of dopamine hydrochloride (Dop-HCl) powder is added to the Tris-HCl buffer solution to maintain a dopamine hydrochloride concentration of 2 mg / mL. After dissolution, the dried sponge is placed in a jar and reacted in a shaking bed at 37°C for 12 hours, then washed with deionized water and anhydrous ethanol alternately for 3 times, and freeze-dried to obtain the sponge with a polydopamine coating (DPU).

[0067] Comparative Example 2

[0068] The preparation method of active antibacterial dressing zinc polyurethane sponge (ZnPU) is as follows: the medical polyurethane sponge PU is cut into a cylinder with a diameter of 10 mm and a height of 5 mm, washed and dried, and subjected to ultraviolet sterilization, then immersed in a zinc chloride solution (12 mg / mL) and placed in a shaking bed at 37°C for 3 hours, washed with deionized water for 3 times, and freeze-dried to obtain the zinc polyurethane sponge ZnPU.

[0069] Comparative Example 3

[0070] The dopamine-zinc alginate blended sponge (APBM-L) is prepared by a blending method. The DPU prepared in Comparative Example 1 is first immersed in a zinc ion solution (12 mg / mL) for 30 seconds, then directly added to a sodium alginate solution (0.01 g / mL), and placed in a shaking bed at 37°C for 2 hours, and freeze-dried to obtain the APBM-L.

[0071] Comparative Example 4

[0072] Different from Example 1, in step 2), the B step is immersed in a zinc chloride solution (12 mg / mL) for 20 seconds, and then immersed in the prepared sodium alginate solution again and placed for 20 seconds; the rest is the same as Example 1. The surface morphology of the prepared dressing is shown in Figure 13 , and the sponge inside the opening structure appears a gelatinized wrinkle state with unclear levels.

[0073] Comparative Example 5

[0074] Different from Example 1, in step 2), the C step of “repeat step B) three times” is changed to “repeat step B) once”; the rest is the same as Example 1. The surface morphology of the prepared dressing is shown in Figure 14 , and the morphology and structural integrity of the sponge are not affected. And it shows similar opening structure and soft foam characteristics as the base sponge.

[0075] The physicochemical properties and biological properties of the active and passive synergistic antibacterial and healing-promoting dressing prepared in Example 1, as well as the dressing prepared in the comparative example, were characterized and tested, as follows:

[0076] The surface morphology of polyurethane (PU), the dopamine-zinc alginate blended sponge (APBM-L) prepared in Comparative Example 3, and the dopamine-zinc alginate dressing (APBM) prepared in Example 1 were observed using scanning electron microscopy (SEM) at three different magnifications. Changes in the pore structure of the materials were observed using SEM (JSM-6390LV, NEC Japan) to confirm coating uniformity and porous structure integrity. Surface and cross-sectional morphology were observed using SEM. The sponge samples were observed at 15 kV after being coated with gold for 160 seconds. The results are as follows. Figure 1 As shown, the PU sponge cross-section surface is very smooth, without impurities or deposits. Deposition of self-polymerized polydopamine (PDA) particles was observed on the surface of the dopamine-zinc alginate blended sponge (APBM-L), indicating successful PDA adhesion to the PU surface. In the SEM image of the dopamine-zinc alginate dressing (APBM), a gelled wrinkled state can be seen on the sponge skeleton, forming a rough surface structure, but the morphology and structural integrity are completely unaffected, exhibiting good open-cell structure and soft foam characteristics, with a relatively concentrated pore size distribution. The porous structure is not destroyed in the dopamine-zinc alginate dressing (APBM). However, the dopamine-zinc alginate blended sponge (APBM-L) synthesized through long-term blending exhibits severe gelation, with the internal open-cell structure broken, becoming loose and lacking clear layers.

[0077] The active-passive synergistic antibacterial and healing-promoting dressing (APBM) prepared in Example 1, polyurethane (PU), DPU prepared in Comparative Example 1, and ZnPU prepared in Comparative Example 2 were tested by Fourier transform infrared spectroscopy (FTIR). The results are as follows: Figure 2 As shown, the successful introduction of zinc ions and sodium alginate is verified. The spectrum of DPU is similar to that of PU, at 3314 cm⁻¹. -1 The increased intensity of one peak indicates the stretching vibration of the polydopamine-OH groups, proving that PDA was successfully coated onto the PU surface. For zinc alginate, the peak intensity is between 3000-3600 cm⁻¹. -1 A broad hydroxyl absorption peak is observed, with peak values ​​for both asymmetric and symmetric stretching vibrations at approximately 1618 cm⁻¹. -1 and 1419cm -1 The display is due to its carboxylate ionic state. This is achieved through the interaction of sodium alginate (SA) and Zn. 2+ Cross-linking to prepare hydrogels. Hydrogels are prepared by cross-linking sodium alginate (SA) molecular chains with Zn. 2+ The bonds combine to form hydrogels via ionic bonds. Therefore, their active hydrogen absorption peaks, such as those of hydroxyl, amino, and carboxyl groups, are located at 3000-3500 cm⁻¹. -1tends to be flat. In addition, due to the strong interaction between the carboxylate anion and Zn 2+ the stretching vibration absorption peak of the carbonyl group (COO ) shifts from 1618 cm -1 and 1419 cm -1 to 1624 cm -1 and 1502 cm -1 .

[0078] The water absorption and water retention capacity of the wound dressing is an important characteristic for evaluating the cleaning effect and maintaining the moist effect of wound exudate. The ideal wound dressing should maintain the wetness of the wound while effectively preventing excessive exudate from accumulating on the wound surface.

[0079] The water absorption rate of the polyurethane sponge PU and the APBM dressing prepared in Example 1 and the water retention rate were measured, which can be evaluated according to the method of YY / T0471.1-2004. The dry samples for measurement were weighed in triplicate, then immersed in phosphate buffered saline (PBS, pH = 7.4), and finally placed in an oven at 37°C for 10 minutes. After standing for 30 seconds, the sample was taken out with tweezers, weighed until the liquid droplet formation stopped, and the water absorption capacity of the sponge was calculated according to the formula.

[0080]

[0081] wherein m1 is the initial mass of the sponge, g; m2 is the mass of the sponge after water absorption, g; the test results are shown in Table 1. Figure 3

[0082] The water retention rate of the polyurethane sponge PU, the APBM prepared in Example 1 and the APBM-L prepared in Comparative Example 3 was measured, and the specific test method was as follows: the dressings were respectively placed in pure water, and after the water absorption reached the saturated state, the samples were taken out at different time points, the excess liquid on the surface was wiped off and weighed. The calculation formula is as follows:

[0083]

[0084] wherein W2 is the mass of the sponge after the water absorption reaches the saturated state, g; W1 is the mass of the sponge after being placed for different time, g; the test results are shown in Table 2.

[0085] Figure 4

[0086] Due to the hydrophilic modification, the dopamine-zinc alginate dressing APBM and APBM-L have improved water absorption rate and water retention capacity. Among them, compared with the polyurethane sponge PU, the dopamine-zinc alginate dressing APBM can absorb liquid up to 1704% of high water absorption within 10 minutes and the water retention rate is 54%. It is sufficient to make the wound dressing absorb the wound exudate while keeping the moist wound surface. ​​​

[0087] An ideal wound dressing should maintain the wound moist while effectively preventing excessive exudate from accumulating on the wound surface. A certain degree of moisture and gas permeability through the wound dressing is important to prevent excessive dehydration and exudate accumulation in the wound. Therefore, the appropriate water vapor transmission rate (WVTR) is also particularly important. The ideal WVTR is 2000-2500 g·m -2 / day. The permeability of the active and passive synergistic antibacterial and healing-promoting dressing (APBM) prepared in Example 1 and polyurethane sponge PU was measured, and the specific test method was as follows: the water vapor transmission rate (WVTR) of the dressing was measured according to YY / T0471.2-2004. Briefly, a certain amount of pure water was first added to a centrifuge tube, the sponge dressing APBM prepared in Example 1 was cut into a uniform size and placed in the centrifuge tube, and then the device was placed in a 37°C incubator. After different time periods, the volume change was measured, and the specific formula is as follows:

[0088]

[0089] In the formula: m0- the initial mass of the centrifuge tube device with sponge and water, g;

[0090] m t - the mass of the entire device after time t, g;

[0091] S- the area of the centrifuge tube opening, mm;

[0092] t- the number of days the entire device was placed in the incubator, d; the test results are shown in Figure 5 .

[0093] The dopamine-zinc alginate dressing can reduce 18.17% compared with the polyurethane sponge PU dressing (2450.9 g·m -2 ·day -1 ). Compared with the blank group (16417.9 g·m -2 ·day -1 ), the moisture loss can be reduced by 82.35%, which has an ideal permeability value. This not only helps to create a moist environment, but also prevents excessive dehydration.

[0094] Porosity is an important parameter for medical sponge applications. High porosity structure not only has high specific surface area, but also helps to absorb exudates. The porosity of the active and passive bacteria-mitigating and healing-promoting dressing (APBM) prepared in Example 1 and polyurethane sponge PU was determined by liquid volume displacement method. The specific test method was as follows: the sample was weighed and the volume was measured and calculated (cylindrical, radius r was about 1.1 cm, thickness h was about 0.5 cm), the mass was weighed, recorded as W1, and placed in anhydrous ethanol, soaked for 5 min, so as to fully absorb anhydrous ethanol. Then the sponge sample was taken out from the anhydrous ethanol, the excess anhydrous ethanol was absorbed and wiped dry, and immediately weighed and recorded as W2. The porosity of the sample was calculated using the following formula:

[0095]

[0096] wherein W1 and W2 are the initial weight and the final weight of the sponge sample, respectively. The porosity was calculated based on three parallel sample tests, expressed as the arithmetic mean and standard deviation (M±SD, n=3).

[0097] Compared with the porosity of PU (81%), the porosity of dopamine-zinc alginate dressing sponge can reach 86.7%. After modification by dopamine DOP and gelation, the porosity of dopamine-zinc alginate dressing composite dressing is increased. Combined with the SEM image, it shows a regular and homogeneous high-porosity structure. This makes the composite sponge dopamine-zinc alginate dressing have potential application prospect in wound dressings. Figure 1

[0098] The stress-strain test was performed on the dopamine-zinc alginate dressing APBM prepared in Example 1 and the polyurethane sponge PU to test the stiffness of the dressing. The specific test method was as follows: the mechanical properties of the dressing were evaluated by a universal testing machine (WDW-05L, China) according to GB / T6344-2008 and GB / T33609-2017. The standard rectangular sample (10 cm x 1.5 cm x 0.5 cm under dry conditions) was stretched at a rate of 100 mm / min. The compression test was performed at a constant speed of 5 mm / min. The experiment was carried out at room temperature. Each specimen was tested at least 3 times. The test results were as follows: Figure 6 ​The tensile strength of polyurethane sponge PU and dopamine-zinc alginate dressing APBM were 137.3 MPa and 238.67 MPa, respectively, and the corresponding breaking strain was 479.99% and 669.01%, respectively. After introducing polydopamine PDA oxidation polymerization and zinc alginate multilayer coating, the mechanical properties of PU sponge were significantly improved. Compared with PU, the tensile strength of dopamine-zinc alginate dressing increased by 101.37 kPa, and the elongation at break increased by 189.02%. The compression test results show that the compressive strength of dopamine-zinc alginate dressing is 28.4 kPa, which is 11.1 kPa higher than that of PU. The mechanical properties are improved and the flexibility of PU is enhanced due to the addition of sodium alginate gel.

[0099] The in vitro antibacterial performance of PU, DPU prepared in Comparative Example 1, ZnPU prepared in Comparative Example 2, and APBM prepared in Example 1 was tested. The specific test method was: colony counting method, using only the culture medium with Staphylococcus aureus, Escherichia coli and Candida albicans added as the control group, and the antibacterial activity of the dressing on Escherichia coli, Staphylococcus aureus and fungi was evaluated respectively. The sponge was cut into a cylinder with a height of 5 mm and a diameter of 20 mm, and ultraviolet sterilized for 30 min. 4 mL of phosphate buffer solution (PBS) was added to the sterilized sponge for soaking, and then 200 uL of three kinds of bacterial solution (10 6 CFU / mL) was added respectively, and incubated at 37°C for 2 h. After incubation, the bacterial solution was diluted and 100 uL of bacterial solution was spread on agar plates. After incubation at 37°C for 24 h (fungal medium was placed at 30°C), the colonies on the agar plates were observed and photographed. The test results are shown in Figure 7

[0100] Compared with the control group, zinc ions can destroy the structure of bacteria by inhibiting the formation of bacterial biofilm and interfering with amino acid metabolism and enzyme systems, thereby inducing bacterial death. 99% of Staphylococcus aureus and Escherichia coli were killed after being in contact with dopamine-zinc alginate dressing APBM for only two hours. Among them, the inhibition of DPU and ZnPU on Staphylococcus aureus was 60% and 75%, respectively, and the inhibition on Escherichia coli was 50% and 65%, respectively. PU has no chemical antibacterial property, and the inhibition rate is only 8.69%. The above results show that the antibacterial effect of dopamine-zinc alginate dressing APBM is better than that of ZnPU (active sterilization) and DPU (passive bacteria prevention) dressing.

[0101] ​To test the in vitro bacterial barrier performance of APBM, the antibacterial effects of DPU, ZnPU, and APBM dressings were evaluated using an agar diffusion assay. PU served as the control group. Specifically, PU, ​​DPU, ZnPU, and APBM samples were cut into cylindrical sections (10mm × 10mm × 5mm) and sterilized under UV light for 30 minutes. Bacterial suspensions (Escherichia coli and Staphylococcus aureus, 1×10⁻⁶) were prepared. 6 CFU / L, 1 mL) was evenly dropped onto one side of the dressing (i.e., the top layer). The sample with the bacterial solution dropped onto the surface was placed on an LB agar plate (with the surface in contact with the bacteria facing upwards) and placed in a bacterial incubator at 37°C. The plate was removed daily to observe changes in bacterial penetration and growth area. Four groups of samples were tested according to the above test procedure. The test results are as follows: Figure 8 As shown, in the PU group, both Staphylococcus aureus and Escherichia coli grew on the plate through the dressing and covered the largest area, indicating that the PU dressing had no antibacterial effect. In the APBM group, no infection appeared on the dressing surface after a week of continuous incubation. However, the control dressing was covered with bacterial colonies, which turned yellow, indicating that the entire dressing was infected with bacteria. Furthermore, it can be seen that the bacterial infection area of ​​PU, DPU (passive antibacterial), and ZnPU (active antibacterial) increased over time. Compared with the PU group, the bacterial growth area in the DPU group decreased by 20% and 40%, respectively. This is mainly due to the self-polymerization of dopamine on the sponge surface to form a firm, dark brown PDA coating. Besides its antibacterial effect, the rough surface facilitates bacterial adhesion to the material surface, preventing some bacteria from growing on the plate through the sponge. Calculating the bacterial infection area on day 7 of incubation, the infection area of ​​Staphylococcus aureus (SA) in the PU group reached as high as 12 cm². 2 The area infected with E. coli (EC) was as high as 16 cm. 2 No bacterial infection occurred at the APBM, indicating that the APBM provides a good barrier for surface bacteria, making it difficult for external bacteria to invade.

[0102] To investigate the independent antibacterial properties of APBM, the antibacterial properties of PU, DPU, and APBM dressings were tested separately. A bacterial capture experiment was conducted, specifically using a soluble hyaluronic acid filter membrane (HAFM) to quantitatively verify the antibacterial ability of APBM. The test method was as follows: each dressing was cut into a cylinder 5 mm high and 10 mm in diameter. After UV sterilization for 30 min, the cylinder was placed inside an acrylic cylinder with a diameter of 10 mm and a height of 10 mm. A transparent air filter membrane (HAFM) with a diameter of 12 mm was placed on a clean transparent plate, with the bottom of the cylinder in contact with the filter membrane (but not with the sponge). An aerosol bottle (containing Staphylococcus aureus bacterial suspension, 1×10⁻⁶) was then used. 4CFU / L, 5 mL) was sprayed over the top of the PMMA cylinder, aligned with the sponge, and timed for 30 s. After the end of spraying, the HAFM was left to stand for 1 min, removed and placed in an LB Petri dish for 30 s, and then the membrane was left to dissolve before plating. After 24 h of incubation at 37 °C, the colonies on the agar plate were observed and photographed. It should be noted that the HAFM is a porous structure filter membrane (porous filter membrane for efficient capture and instant quantitative detection of air pathogens prepared in Example 1 of patent 202410540252.7) constructed by an ethanol-induced phase separation strategy, which can capture and collect pathogens in the air with high performance. Therefore, it is extremely effective to collect pathogens in the closed environment formed between the PMMA cylinder and the aerosol bottle by the HAFM. The test results after spraying and plating are shown in Figure 9 The results show that the APBM group can achieve a bacteria blocking effect of 80%, and the bacteria blocking rate of the DPU group is greater than 55%, indicating that the passive design of the dopamine-modified layer can play a role in blocking and adhering bacteria, and the APBM as a whole has a more superior effect.

[0103] Biocompatibility is a basic prerequisite for the clinical transformation of wound dressings. To test the biocompatibility of dopamine-zinc alginate dressing, we conducted cytotoxicity tests and hemolysis tests.

[0104] The specific method of the cytotoxicity test is as follows: the CCK-8 method was used to evaluate the cytotoxicity of the sponge. L929 cells were cultured in DMEM medium containing 10% fetal bovine serum and double antibodies. The control group was a group with only the addition of culture medium. The dopamine-zinc alginate dressing sponge APBM prepared in Example 1 was soaked in DMEM high-sugar medium for 24 h, and a 0.22 μm filter membrane (Sartorius, Germany) was used for filtration to obtain a sponge extract. Then it was diluted to 0.25, 0.5, 0.75 and 1 mg / mL. The concentration of L929 cells was adjusted to 5 × 10 4 cells / mL, 100 μL per well was inoculated in a 96-well plate, and after the cells adhered, different concentrations of sponge extract were replaced, and then 100 μL of sponge extract was used to replace the culture medium for further culture for 24 h and 72 h. After the corresponding culture time, the original culture medium was aspirated, 100 μL of CCK-8 detection solution (CCK-8:DMEM medium = 1:9) was added to each well, and it was incubated at 37 °C for 2 h. The OD value was measured at 450 nm by using an enzyme-labeled instrument. The calculation formula of cell proliferation rate is as follows:

[0105]

[0106] wherein OD1 and OD0 represent the absorbance values of the experimental object and the negative control, respectively.

[0107] The test results are shown in Figure 10(a) shows that the results show that each concentration group has no effect on cell activity, with no statistical difference compared with the control group.

[0108] Hemolysis experiment: New Zealand white rabbits' anticoagulant whole blood was used to evaluate the hemolysis of PU and dopamine-zinc alginate dressing APBM prepared in Example 1. Red blood cells were obtained by centrifuging whole blood at 1500 rpm for 15 minutes, then washed in PBS until the supernatant was colorless. 1 mL of red blood cells was added to 9 mL of sterile PBS to prepare a red blood cell suspension. Then, 1 mg of sample was added to 1 mL of diluted blood as the experimental group, and red blood cells were added to deionized water and PBS as positive and negative controls, respectively. After incubation at 37°C for 1 hour, all groups were centrifuged at 2000 rpm for 5 minutes. The absorbance of the supernatant at 540 nm was measured by UV-visible spectrophotometer. The hemolysis ratio (HR) of the sponge was calculated according to the following formula (5).

[0109]

[0110] where OD S represents the absorbance of the dressing, OD (+) represents the absorbance of the positive sample, and OD (-) represents the absorbance of the negative sample.

[0111] The test results are shown in Figure 10 (b), the hemolysis rate of the wound dressing is less than 2%, which is the minimum standard for the clinical application of biomedical materials. After evaluation, the hemolysis activity test of dopamine-zinc alginate dressing APBM showed that its hemolysis rate was less than 1.5% compared with the positive group. The above results show that dopamine-zinc alginate dressing has good biocompatibility, providing a reliable basis for wound treatment.

[0112] The main difficulty in the treatment of burns and scalds is microbial infection and inflammation, which ultimately leads to delayed wound healing. To explore the efficacy of dopamine-zinc alginate dressing in promoting healing and anti-inflammatory effects, the wound healing effect of rat scalded infected wounds was verified, and the specific experimental method was as follows:

[0113] In order to evaluate the bacterial barrier performance and healing-promoting performance of dopamine-zinc alginate dressing as a wound dressing, a rat bacterial infected scald model was designed. Normal SD rats were randomly divided into control group, PU group, DPU group and dopamine-zinc alginate dressing group. The back of the anesthetized rats was shaved and depilated with depilatory cream, and the exposed skin was sterilized with 1% povidone-iodine solution and washed with normal saline. Two deep second-degree burns with a diameter of 1.5 cm were made on the back of the rats using a burn instrument, and thirty minutes later the scab part of the wound was removed, and then dopamine-zinc alginate dressing and PU dressing were fixed on the surface of the wound. Subsequently, 1 mL of Staphylococcus aureus suspension (1 x 10 6 CFU / mL-1 ) Sprayed on the wound and the surface of the dressing, the control group was sprayed with bacterial liquid on the surface of the wound. The rats were labeled and fed separately, and the wound was photographed, while the dressing was changed every 4-5 days and the recovery of the wound was observed. ImageJ was used to analyze the wound closure area. The wound healing rate was calculated by the following formula.

[0114]

[0115] Where S0 and S represent the initial wound area and the wound area on the day of examination, respectively.

[0116] It should be noted that all experiments were carried out in accordance with the guidelines for the care and use of laboratory animals, and every effort was made to minimize suffering. This study was approved by the Animal Ethics Committee of Qingdao University, China (No. QDU-AEC-2024708). The experimental protocol was performed in accordance with the ARRIVE guidelines. Sprague Dawley rats (male, 250-280 grams, production license number: SCXK(Lu)2022-0006) were purchased from Jinan Pengyue Experimental Animal Co., Ltd.

[0117] The results are shown in Figure 11 , with the blank treatment as a control, to further evaluate the effect of APBM on the healing ability of scald infected wounds. The wound area was quantitatively analyzed on days 0, 5, 8, 11, and 15 after wounding. The wound after scalding was porcelain white with mild swelling. The skin in the wound area began to discolor and scab on the fifth day. The color of the skin changed from porcelain white to light yellow, brown or light brown, and finally red black. By day 8, there was a clear demarcation line between the scalded area and normal tissue. The wound showed tissue hyperemia with local brown and red borders. Compared with the other three groups (blank control group, PU, DAPU), APBM sponges significantly promoted wound closure, showing a faster healing rate than the other groups. In particular, during this period from day 2 to day 10. By day 15, the average wound closure rates for the blank control group, PU, DPU, and APBM were 60.27%, 64.32%, 73.49%, and 87.39%, respectively; the wound treated with APBM healed significantly faster than the other three groups during the 15-day healing period.

[0118] To fully assess the therapeutic effect of APBM, histopathological evaluation of tissue samples was performed using hematoxylin and eosin (H&E) staining and Masson's trichrome staining to analyze the degree of inflammatory infiltration, re-epithelialization, angiogenesis, and collagen content. Post-surgery, soft tissue surrounding the wound was collected at day 11 and day 15 for histopathological analysis. Briefly, tissues were dehydrated in ethanol, cleared in xylene, and then embedded in paraffin. H&E staining, Masson's staining, and immunofluorescence staining were used to evaluate the wound bed for collagen deposition, epithelialization, angiogenesis, and inflammatory infiltration, and the blank control group was the untreated group, which was only scalded without drug administration. The test results are shown in Figure 12 H&E staining showed that in the blank control group and the PU group, immune cells infiltrated into the interstitial region and muscle tissue around the implant, and there was a significant accumulation of inflammatory cells and inflammatory necrotic foci on the surface of the tissue. The APBM group significantly inhibited neutrophil infiltration. During wound healing, epithelial cells can act as a skin barrier to avoid pathogen infection, so evaluating re-epithelialization is crucial to wound healing. Histological results showed that compared with the blank control group and the PU group, re-epithelialization occurred in the wounds treated with DPU and APBM sponges, especially in the central region of the defect. In addition, through HE staining at day 11 and day 15, it was observed that the new epidermis formed well in the APBM group. By day 15, in the APBM group, the new epidermis was well integrated with the dermis, and a relatively complete epidermis and dermis were observed. This highlights the role of APBM in the early regenerative phase of inflammation, proliferation, and healing. Collagen can repair defects, restore the organizational structure and function of tissues, and improve the extracellular matrix framework and lay the foundation for the formation of wound cell matrix. Collagen deposition is necessary in the remodeling phase of blood vessels and skin healing to enhance the tensile strength of the tissue and support wound healing. This study observed the formation of collagen through Masson's staining, and the results showed that at day 11, only a small amount of loose and disordered collagen fibers were present in the wounds of the blank control group and the PU-treated group, in contrast, at day 11 and day 15 post-surgery, the rat skin tissue in the APBM group exhibited denser collagen fibers and more collagen deposition. Meanwhile, the histological results at day 11 and day 15 showed that the APBM group had more than 2.5 times the amount of collagen fiber deposition compared with the blank control group and the PU group. It was also better than the collagen deposition results of the DPU group. The collagen was denser and more mature, with a highly ordered fibrous collagen structure similar to the histopathological structure of normal skin tissue, which was beneficial for maintaining the mechanical integrity of the skin and reducing scar formation. This also indicates that the incorporation of zinc ions and the design of the antibacterial strategy significantly enhance collagen fiber deposition.

[0119] In the wound healing process, neovascularization is essential to ensure sufficient nutrients and oxygen to the wound tissue to support fibroblast proliferation, collagen synthesis and re-epithelialization. Immunofluorescence staining of CD31, a-smooth muscle actin (a-SMA) was performed to study the vascularization and recovery of wound tissue. Angiogenesis is essential for tissue regeneration and repair. Impaired vascularization delays wound closure and increases the risk of infection. CD31 is a transmembrane protein expressed in angiogenesis, indicating neovascularization. The effect of APBM on CD31 expression is shown in the figure. On day 11, the expression level of CD31 in the APBM group was significantly higher than that in the blank control group and the PU group (P < 0.01). On day 15, the expression trend of CD31 in the APBM group was consistent with that on day 11 (P < 0.001). These studies show that APBM dressing can promote angiogenesis during wound healing and further accelerate wound healing. a-SMA is a cytoplasmic protein expressed in late angiogenesis, indicating the maturation of vascular smooth muscle cells. Immunofluorescence experiments showed that compared with the control group blank control group, PU and DPU group, the expression of a-SMA in the APBM treatment group was significantly higher than the other three groups on days 11 and 15, and the fluorescence expression intensity was more than 2 times. This indicates that APBM dressing can more effectively stimulate wound angiogenesis.

[0120] In summary, the APBM group accelerated the healing of rat scald wounds by reducing inflammatory infiltration in the wound, promoting re-epithelialization in damaged skin lesions, collagen deposition, and vascular regeneration. The above results strongly support the better application prospects and potential of APBM dressing in open wounds and infected wounds.

[0121] Although the present application has been described in detail through the preferred embodiments, the present application is not limited thereto. Those skilled in the art can make various equivalent modifications or replacements to the embodiments of the present application without departing from the spirit and essence of the present application, and these modifications or replacements shall be within the scope of the present application. Therefore, the protection scope of the present application shall be subject to the protection scope of the claims.

Claims

1. A method of preparing a passive-active synergistic antibacterial and healing-promoting dressing, characterized in that, The method comprises the following steps: 1) Complexing dopamine with polyurethane sponge: mixing polyurethane sponge with dopamine solution, and performing self-polymerization in an alkaline environment to form a polydopamine coating; 2) sodium alginate Zinc ion gelation: dopamine-modified polyurethane sponge was immersed in a solution of sodium alginate and zinc ions, and short-time multiple cycles of crosslinking were performed to form a zinc-containing sodium alginate hydrogel coating; 3) Freeze-drying the crosslinked dressing to obtain the active and passive synergistic antibacterial and healing-promoting dressing; The concentration of the dopamine solution in step 1) is 2 mg ± 0.5 mg / mL, and the alkaline environment in step 1) is a tris-hydroxymethyl aminomethane hydrochloride buffer; The concentration of sodium alginate in step 2) is 0.01 g / mL, and the concentration of zinc ions is 12-20 mg / mL; In step 2), the sponge with a polydopamine coating is immersed in a sodium alginate solution, the material obtained by immersing in the sodium alginate solution is sequentially immersed in a zinc ion solution and a sodium alginate solution for short-time multiple cycle crosslinking, and then immersed in a zinc ion solution to form a zinc-containing sodium alginate hydrogel coating; The short-time multiple cycle crosslinking is 10 s each time in the zinc ion solution and the sodium alginate solution, and the number of crosslinking is 3-6 times; The dressing has passive adhesion and bacteria blocking performance and zinc ion release active antibacterial performance; the passive antibacterial efficiency can reach 80 ± 1.2%, and the antibacterial rate of the active and passive synergistic antibacterial and healing-promoting dressing can reach more than 99%.

2. The application of the dressing prepared by the method of claim 1 in open wounds of burns and scalds.

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