Double-layer plasma activated gel patch as well as preparation method and application thereof

Abstract

The invention discloses a double-layer plasma activated gel patch as well as a preparation method and application thereof. Comprising the following steps: dissolving polyvinyl alcohol and polyethylene glycol in a mass ratio of (1: 0.8)-(1: 1.2) in deionized water, stirring and dissolving in a water bath of 90-95 DEG C, and cooling to room temperature to prepare a matrix solution; performing plasma-rich activation on the matrix solution to obtain a lower-layer gel precursor solution, injecting the lower-layer gel precursor solution into a mold, and performing physical freeze-thaw cross-linking treatment for 2-5 times, namely freezing at-20 DEG C to-80 DEG C for 8-12 hours and melting at 4 DEG C to room temperature for 4-8 hours to obtain lower-layer gel; additionally taking a matrix solution, activating the matrix solution through NO-rich plasma to obtain an upper-layer gel precursor solution, pouring the upper-layer gel precursor solution on the surface of the lower-layer gel, and repeating the freeze-thaw cross-linking treatment to form double-layer gel; and immersing the double-layer gel into a solution containing an adhesive polymer for 1-2 hours, and then washing to obtain a finished product. The gel patch prepared by the invention realizes a sequential effect of resisting bacteria first and then promoting healing through a double-layer structure, has no chemical residue in physical freeze-thaw cross-linking, has wound surface fitting property, and is suitable for treating infectious and chronic skin wound surfaces.

Description

Technical Field

[0001] This application relates to the field of pharmaceutical technology, and in particular to a double-layer plasma-activated gel patch, its preparation method, and its application. Background Technology

[0002] With the increasing incidence of chronic wounds (such as diabetic foot ulcers, venous ulcers, and pressure ulcers) and infected skin wounds, these wounds have become a significant treatment challenge in the medical field due to their long healing cycles and susceptibility to recurrent infections. Current clinical treatments mostly rely on antibiotics combined with traditional dressings, or atmospheric pressure cold plasma alone, but both have significant limitations. Long-term use of antibiotics easily leads to drug resistance and can only target the infection stage, failing to meet the sequential needs of wound healing from infection control to tissue regeneration. Although gaseous plasma can generate antibacterial and healing-promoting active particles, when administered via "gas purging," the active particles have short half-lives and limited durations of action, requiring patients to make multiple hospital visits, making it difficult to cover the wound repair cycle of several days to several weeks.

[0003] In existing technologies, hydrogel dressings are widely used because they can carry drugs and maintain a moist environment on the wound. However, there are significant shortcomings in the preparation and functional design of existing hydrogels. On the one hand, most hydrogels are prone to the contradiction of "insufficient antibacterial dosage / excessive healing-promoting dosage" or vice versa, making sequential treatment impossible. On the other hand, some preparation processes rely on chemical cross-linking agents, which are prone to residues that can cause skin irritation. In existing physical cross-linking schemes, an imbalance can occur between the mechanical properties of the gel and the loading capacity of active particles.

[0004] Therefore, how to develop a gel patch preparation method with clear process parameters, no safety residue from physical cross-linking, and a sequential function of "antibacterial first, then healing" to meet the needs of efficient treatment of infected skin wounds is a key problem that urgently needs to be solved in this field. Summary of the Invention

[0005] This application provides a double-layer plasma-activated gel patch, its preparation method, and its application, which solves the problems in the prior art where most hydrogels cannot achieve sequential treatment, are prone to residue causing skin irritation, and have an imbalance between the mechanical properties of the gel and the loading capacity of active particles.

[0006] In a first aspect, embodiments of this application provide a method for preparing a double-layer plasma-activated gel patch, comprising: Polyvinyl alcohol and polyethylene glycol are dissolved in deionized water, stirred and dissolved in deionized water at 90-95℃, and cooled to room temperature to obtain a matrix solution. The mass ratio of polyvinyl alcohol to polyethylene glycol is in the range of 1:0.8-1:1.2. The matrix solution is enriched with... The plasma was used to activate the lower gel precursor solution to obtain the lower gel precursor solution. The lower gel precursor liquid is injected into the mold and subjected to 2-5 physical freeze-thaw crosslinking treatments to obtain the lower gel. The physical freeze-thaw crosslinking treatment is performed by freezing at a temperature of -20℃ to -80℃ for 8-12 hours and thawing at a temperature of 4℃ to room temperature for 4-8 hours. The matrix solution is activated with NO-rich plasma to obtain an upper gel precursor solution; the upper gel precursor solution is poured onto the surface of the lower gel, and after the physical freeze-thaw crosslinking treatment, a bilayer gel is obtained. The bilayer gel is immersed in a solution containing an adhesive polymer for 1-2 hours and then rinsed to obtain a bilayer plasma-activated gel patch.

[0007] In conjunction with the first aspect, in one possible implementation, the matrix solution further includes a biocompatible polymer material, which includes at least one of sodium alginate, hyaluronic acid, sodium hyaluronate, chitosan, carboxylated chitosan, cellulose, cellulose derivatives, pectin, gelatin, and poloxamer.

[0008] In conjunction with the first aspect, in one possible implementation, the working gas for the plasma activation treatment is a mixture of nitrogen and oxygen or air, the flow rate of the working gas is 0-5 SLM, the activation treatment method is irradiation, gas blowing or bubbling, and the activation treatment time is 5-30 min.

[0009] In conjunction with the first aspect, in one possible implementation, the solution containing the adhesive polymer is polydopamine, polyphenols, tannic acid, mussel adhesive protein, or a combination thereof.

[0010] In conjunction with the first aspect, in one possible implementation, the rinsing is performed using deionized water, and the rinsing is performed 2-3 times, with each rinsing lasting 1-2 minutes.

[0011] In conjunction with the first aspect, in one possible implementation, the mold is made of polytetrafluoroethylene or glass, and the mold shape is circular, square, or irregular. The thickness of the double-layer gel is 1-6 mm by controlling the pouring amount of the upper gel precursor liquid and the lower gel precursor liquid.

[0012] In conjunction with the first aspect, in one possible implementation, when the polyvinyl alcohol and polyethylene glycol are dissolved in deionized water at 90-95°C, the stirring speed is 300-500 r / min. After the lower gel precursor liquid is injected into the mold, the mold is sealed before physical freeze-thaw crosslinking treatment. When the bilayer gel is immersed in a solution containing an adhesive polymer, it is completely submerged in the solution.

[0013] In conjunction with the first aspect, in one possible implementation, the plasma generating device is a dielectric barrier discharge device or a sliding arc discharge device.

[0014] Secondly, embodiments of this application provide a double-layer plasma-activated gel patch prepared using the preparation method of the double-layer plasma-activated gel patch described in the first aspect or any possible implementation of the first aspect, comprising: The lower gel and the upper gel are both based on a polyvinyl alcohol-polyethylene glycol blend and are physically cross-linked to form an integral structure. The lower gel is loaded with rich Active particles from plasma conversion; The upper gel is loaded with NO-rich plasma-converted active particles, mainly NO. The bilayer gel, after being treated with a solution containing an adhesive polymer, has the property of adhering to the skin or wound.

[0015] Thirdly, embodiments of this application provide an application of a double-layer plasma-activated gel patch prepared using the preparation method of the double-layer plasma-activated gel patch described in the first aspect or any possible implementation of the first aspect, characterized in that it includes: The double-layer plasma-activated gel patch is used to prepare topical medications for skin wounds; The skin wounds include infected skin wounds and chronic wounds, wherein the chronic wounds are diabetic foot ulcers, venous ulcers, or pressure ulcers.

[0016] One or more technical solutions provided in the embodiments of this application have at least the following technical effects or advantages: In this embodiment, a matrix solution is prepared by dissolving polyvinyl alcohol and polyethylene glycol in deionized water at a mass ratio of 1:0.8-1:1.2, stirring and dissolving in a water bath at 90-95°C, and then cooling to room temperature; the matrix solution is then enriched with... Plasma activation yields a lower gel precursor solution, which is injected into a mold and subjected to physical freeze-thaw crosslinking treatment 2-5 times, involving freezing at -20℃ to -80℃ for 8-12 hours and thawing at 4℃ to room temperature for 4-8 hours, to obtain the lower gel. Separately, a matrix solution is activated with NO-rich plasma to obtain an upper gel precursor solution, which is poured onto the surface of the lower gel and the above freeze-thaw crosslinking treatment is repeated to form a bilayer gel. The bilayer gel is then immersed in a solution containing an adhesive polymer for 1-2 hours and rinsed to obtain the finished product. The gel patch prepared in this application achieves a sequential effect of "antibacterial first, then healing promoted" through its bilayer structure. Physical freeze-thaw crosslinking leaves no chemical residue and provides good wound adhesion, making it suitable for the treatment of infected and chronic skin wounds, thus solving the problems mentioned in the background art. Attached Figure Description

[0017] To more clearly illustrate the technical solutions of the embodiments of this application, the drawings used in the description of the embodiments of this application or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0018] Figure 1 This is a schematic diagram of the design of a plasma-activated gel patch provided in an embodiment of this application; Figure 2 A rich variety of materials provided in the embodiments of this application Fourier transform infrared spectrum of plasma; Figure 3 A Fourier transform infrared spectrum of NO-rich plasma provided for embodiments of this application; Figure 4 A schematic diagram illustrating the antibacterial effect test results of the upper and lower gels provided in the embodiments of this application; Figure 5 The image shows the effect of the double-layer plasma-activated gel patch provided in this application on promoting wound healing in diabetic rats. Detailed Implementation

[0019] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0020] The following description of some technologies involved in the embodiments of this application is provided to aid understanding and should be considered merely exemplary. Therefore, those skilled in the art should recognize that various changes and modifications can be made to the embodiments described herein without departing from the scope and spirit of this application. Similarly, for clarity and brevity, some descriptions of well-known functions and structures are omitted in the following description.

[0021] Example 1: Preparation of a basic double-layer plasma-activated gel patch.

[0022] Figure 1 This is a schematic diagram of a plasma-activated gel patch provided in an embodiment of this application. The double-layer plasma-activated gel patch prepared in this embodiment fully complies with the attached... Figure 1 The functional zoning design shown, consisting of an upper gel layer near the air and a lower gel layer near the wound, follows these steps: Step 1: Precise preparation and performance control of the matrix solution.

[0023] Polyvinyl alcohol (PVA) was selected as the main film-forming material for the gel matrix. The hydroxyl groups on its molecular chains can form hydrogen bonds through physical freeze-thaw cycles, ensuring the mechanical strength of the gel. Polyethylene glycol (PEG) was selected as a plasticizer to improve brittleness and enhance the gel's flexibility. The two were compounded in a 1:1 mass ratio, ensuring gel formability while avoiding excessive swelling due to excessive PEG.

[0024] Polyvinyl alcohol (PVA) and polyethylene glycol (PEG) were added to deionized water in a specific ratio to prepare a mixture with a total solids content of 10 wt%. The mixture was transferred to a double-walled glass reactor equipped with a mechanical stirrer and subjected to a constant temperature water bath at approximately 92°C (temperature fluctuations controlled within ±1°C to prevent localized overheating that could lead to carbonization and yellowing of the PVA). The mixture was stirred continuously at 350 rpm for 2.5 hours, with a small amount of solution observed every 30 minutes using a glass rod, until the solution became homogeneous, transparent, and free of visible undissolved particles. After stirring was stopped, the solution was transferred to a sealed polyethylene container and allowed to cool naturally to room temperature at a constant temperature of 25°C, yielding a final product with a viscosity of [missing value]. The base matrix solution.

[0025] Step 2: Plasma activation and shaping of the lower gel precursor solution.

[0026] The dielectric barrier discharge device and the sliding arc discharge device as defined in the embodiments of this application are used as plasma sources; dry air is selected as the working gas, the gas flow rate through the dielectric barrier discharge device is 5 SLM, the gas flow rate through the sliding arc discharge device is 1 SLM, the discharge power of the two devices is adjusted to 30W, and the preheating device is preheated for 5 minutes to stabilize the discharge state.

[0027] Pour 50 mL of the base matrix solution into a 100 mL gas washing bottle. The gaseous active particles generated by the discharge are enriched by a "bubbling" method. Plasma activation (Fourier transform infrared spectrum of its gaseous active particles, as shown in the figure) Figure 2 As shown, Figure 2 A rich variety of materials provided in the embodiments of this application The Fourier transform infrared spectrum of the plasma confirms that the activation products are... (Mainly). After activation treatment for 15 min, the lower gel precursor solution was obtained. Absorbance (au) is the absorbance. "Absorbance" indicates the degree of light absorption by a substance; "au" is an abbreviation for "arbitrary unit," meaning "any unit." Because the absolute scale of absorbance often varies depending on the detection system, a relative unit is used. Wavenumber ( Wavenumber (unit: centimeters) "Wavenumber" is a physical quantity describing the frequency of electromagnetic waves (such as infrared light), and it is inversely proportional to the wavelength (wavenumber = 1 / wavelength; when the wavelength is in centimeters, the wavenumber is in centimeters). (), is a commonly used abscissa parameter in analyses such as infrared spectroscopy.

[0028] The lower gel precursor solution was slowly injected into the polytetrafluoroethylene mold, and the side wall of the mold was tapped to remove air bubbles. The mold was placed in an ultra-low temperature freezer at -40°C for 10 hours, and then the mold was removed and transferred to a room temperature environment at 25°C for 6 hours to thaw, completing one freeze-thaw cycle. The above freeze-thaw process was repeated 3 times to finally obtain a lower gel with a smooth surface and no cracks.

[0029] Step 3: Plasma activation of the upper gel precursor solution and integral molding of the two layers.

[0030] Take another 50 mL of the base matrix solution and activate it using the sliding arc discharge device provided in the embodiments of this application. The working gas is a mixture of nitrogen and oxygen (volume ratio 9:1, at which ratio NO generation is highest and there is no excessive NO). Impurities were removed, and the flow rate was controlled at 1.5 SLM using a gas mass flow meter. The discharge power was adjusted to 25 W, and the device was preheated for 5 minutes. The matrix solution was poured into a quartz petri dish identical to that in step 2, and activation was performed using a bubbling method. The gas dispersant head was inserted 20 mm into the bottom of the solution to allow the gaseous active particles to fully dissolve in the solution as microbubbles. The activation time was 20 minutes. After activation, the solution was light pink, and the Fourier transform infrared spectrum of its gaseous active particles is shown below. Figure 3 As shown, Figure 3 The Fourier transform infrared spectrum of NO-rich plasma provided in this application embodiment shows no obvious impurity peak interference.

[0031] The upper gel precursor solution was slowly poured onto the surface of the already formed lower gel. During pouring, the solution was poured slowly along the edge of the mold to prevent the precursor solution from impacting the lower gel and causing structural damage. After pouring, the mold was gently shaken to ensure that the upper precursor solution evenly covered the lower gel, ensuring that there were no air bubbles between the two layers. The mold was then placed in a -40°C ultra-low temperature freezer for 10 hours and thawed at 25°C for 6 hours, and this freeze-thaw cycle was repeated 3 times. Through physical freeze-thaw, the polyvinyl alcohol molecular chains of the upper and lower gels interpenetrated to form hydrogen bonds and cross-linked, ultimately resulting in a tightly bonded bilayer gel.

[0032] Step 4: Adhesion post-treatment and finished product preparation.

[0033] The polydopamine provided in the embodiments of this application was selected as the adhesive polymer. Dopamine hydrochloride powder was dissolved in 10 mmol / L Tris-HCl buffer (pH=8.5, at which pH value dopamine is prone to self-polymerization reaction) to prepare a polydopamine solution with a mass concentration of 2wt%. The solution was magnetically stirred for 30 min to completely dissolve and stored in the dark for later use.

[0034] The bilayer gel was completely immersed in a polydopamine solution and left to stand at 25°C in the dark for 1.5 hours (too short an immersion time would result in a thin adhesive layer, while too long an immersion time would cause the gel to absorb too much liquid and swell). After removing the gel, it was rinsed twice with deionized water (1.5 minutes each time) using gentle running water to avoid damaging the adhesive layer on the gel surface. The rinsed gel was then placed in a vacuum drying oven at 4°C for 4 hours to remove excess surface moisture, finally yielding the basic bilayer plasma-activated gel patch. The finished product has a light brown surface and good skin adhesion.

[0035] Example 2: Preparation of a modified bilayer plasma-activated gel patch containing sodium alginate.

[0036] This embodiment, based on Embodiment 1, adds the biocompatible polymer material (sodium alginate) provided in this application embodiment to optimize the gel's permeation performance and biocompatibility. The specific steps are as follows: Step 1: Preparation of modified matrix solution.

[0037] In the basic matrix formulation of Example 1, sodium alginate accounting for 3 wt% of the total solid content was added; polyvinyl alcohol, polyethylene glycol and sodium alginate were added to deionized water in a mass ratio of 1:1:0.03, and the subsequent dissolution process and cooling conditions were the same as in Example 1.

[0038] After dissolution, the viscosity of the modified matrix solution was tested. The viscosity was adjusted by adding a small amount of deionized water. The pH of the solution was measured to be 6.8 using a pH meter, and no additional adjustment was required.

[0039] Steps 2-4: Activation, molding and post-treatment.

[0040] The subsequent activation steps of the lower gel precursor solution (dielectric barrier discharge, air flow rate 5 SLM, sliding arc discharge, air flow rate 1 SLM, treatment for 15 min), upper gel precursor solution activation (sliding arc discharge, nitrogen-oxygen mixture 1.5 SLM, treatment for 20 min), freeze-thaw crosslinking (freezing at -40℃ for 10 h / thawing at 25℃ for 6 h, cycled 3 times) and polydopamine adhesion treatment (2 wt% solution, soaking for 1.5 h) were all completely consistent with those of Example 1. The resulting modified bilayer gel patch had an absorption rate of 450% ± 30%, which was 30% higher than that of the basic patch in Example 1, and a tensile strength of 1.2 MPa, making it more suitable for chronic wounds with a lot of exudate.

[0041] Example 3: Preparation of a highly adhesive bilayer plasma-activated gel patch modified with mussel adhesive protein.

[0042] This embodiment only changes the type of adhesive polymer in step 4 (using the mussel adhesive protein provided in this application embodiment), while the remaining steps are the same as in embodiment 1, aiming to optimize the skin adhesion properties of the gel, as detailed below: Mussel adhesive protein powder was dissolved in 0.01 mol / L phosphate buffer to prepare a 0.5 wt% solution. The solution was magnetically stirred for 1 hour to ensure complete dissolution. Impurities were removed by filtration through a 0.22 μm filter membrane to prevent undissolved particles from affecting the adhesion effect.

[0043] The bilayer gel was completely immersed in mussel adhesive protein solution and allowed to stand for 1 hour at 25°C and 60% humidity. After removing the gel, it was rinsed twice with sterile phosphate-buffered saline (PBS) to remove unbound proteins from the surface. The gel was then dried in a vacuum drying oven at 4°C for 3 hours to obtain a highly adhesive bilayer gel patch.

[0044] Comparative Example 1: Preparation of a single-layer antibacterial plasma-activated gel patch.

[0045] This comparative preparation only loaded with rich The specific steps for creating a monolayer gel patch using plasma-active particles are as follows: Prepare the basic matrix solution according to step 1 of Example 1; Follow step 2 of Example 1 to enrich Plasma activation (dielectric barrier discharge, air flow rate 5 SLM, sliding arc discharge, air flow rate 1 SLM, treatment for 15 min) yielded a monolayer antibacterial precursor solution. The precursor liquid is injected into a polytetrafluoroethylene mold and molded through three freeze-thaw cycles. Following step 4 of Example 1, polydopamine adhesion treatment was performed to finally obtain a single-layer antibacterial gel patch.

[0046] Comparative Example 2: Preparation of a chemically cross-linked bilayer gel patch.

[0047] This comparative example uses a chemical cross-linking agent (glutaraldehyde) to prepare a bilayer gel patch. The specific steps are as follows: Prepare the basic matrix solution according to step 1 of Example 1, add 0.5 wt% glutaraldehyde as a crosslinking agent to the solution, and stir until homogeneous; Subsequent plasma activation (lower layer enrichment) The upper layer is rich in NO), and the double-layer molding steps are the same as in Example 1, but no freeze-thaw cycle is required. Chemical cross-linking is completed by simply standing at 25°C for 24 hours. The polydopamine adhesion treatment was the same as in Example 1, resulting in a chemically cross-linked bilayer gel patch.

[0048] Using Staphylococcus aureus and Escherichia coli as target bacteria, the antibacterial effect was evaluated using the colony forming unit (CFU) method. The specific procedure is as follows: S. aureus and E. coli were inoculated into liquid culture medium and cultured for 12 h in a shaker at 37 °C and 180 r / min. The bacterial culture was then diluted with PBS. spare.

[0049] The gel samples from Example 1, Example 2, and Comparative Example 1 were cut into 1cm × 1cm pieces and placed in a 24-well cell culture plate. 10μL of the above bacterial suspension was added to each well to ensure that the bacterial suspension evenly covered the sample surface.

[0050] Incubate the culture plate in a 37°C incubator for 60 min, then add 1 mL of sterile PBS to each well and gently shake for 10 min to fully elute the surviving bacteria; take the eluent and perform 10-fold serial dilutions (…). ), take 10 μL of each dilution of elution buffer and spread it on LB agar plates. After incubation at 37°C for 24 h, count the number of colonies and calculate the number of viable bacteria and inactivation efficiency; use the sterile PBS treatment group as a negative control.

[0051] like Figure 4 As shown, Figure 4 This is a schematic diagram showing the antibacterial effect test results of the upper and lower gels provided in the embodiments of this application.

[0052] Negative control (PBS): The viable counts of both S. aureus and E. coli were maintained at [value missing]. CFU / mL level, with no obvious antibacterial effect, and Figure 4 The column heights in the "control" group were consistent.

[0053] Example 1: Bilayer Gel: The lower gel showed inactivation efficiencies of more than 6 orders of magnitude against both *S. aureus* and *E. coli* (viable bacterial count < 0.05%). CFU / mL), the upper gel achieved an inactivation efficiency of 2 orders of magnitude against S. aureus (viable bacterial count reduced to 1000 CFU / mL). (CFU / mL), with an inactivation efficiency of over 6 orders of magnitude against E. coli, and... Figure 4 The antibacterial data of the "lower gel" and "upper gel" are completely matched.

[0054] Example 2 Modified bilayer gel: Due to the presence of sodium alginate, the inactivation efficiency of S. aureus and E. coli was improved compared with Example 1, and the number of viable bacteria was further reduced.

[0055] Comparative Example 1: Single-layer antibacterial gel: It has an inactivation effect of 6 orders of magnitude against only two types of bacteria, but has no healing-promoting activity and poor antibacterial persistence.

[0056] Tests to promote wound healing in diabetic rats.

[0057] SPF-grade male SD rats weighing 150±10g were selected and acclimatized for one week. Then, streptozotocin was injected intraperitoneally at a dose of 50mg / kg to induce a diabetes model. One week after the injection, random blood glucose levels at the tail tip of the rats were measured using a blood glucose meter. A blood glucose level ≥16.7mM was considered a successful establishment of a diabetes model.

[0058] After anesthetizing diabetic rats intraperitoneally with 2.5% tribromoethanol solution, the back was shaved and disinfected. A full-thickness skin was removed using a 20mm diameter circular biopsy punch to create a circular wound. Subsequently, 100 μL of L. aureus suspension was dripped onto the wound surface. ,about (CFU / mL), let stand for 30 minutes to allow bacteria to fully colonize, and construct an infected wound model.

[0059] Rats were randomly divided into 3 groups (n=6): "control group" (wound covered with sterile saline gauze, changed every 3 days), "Example 1 double-layer gel group" (wound covered with double-layer gel patch prepared in Example 1, changed every 3 days), and "Comparative Example 2 chemical cross-linking group" (wound covered with chemical cross-linking double-layer gel patch prepared in Comparative Example 2, changed every 3 days).

[0060] On days 3, 6, 9, and 12 of treatment, wound photographs were taken to analyze the wound area and calculate the wound healing rate (healing rate = (initial wound area - remaining wound area) / initial wound area × 100%). At the same time, the appearance of the wound was observed, and the redness, swelling, exudation, and granulation tissue growth were recorded.

[0061] like Figure 5 As shown, Figure 5 The image shows the effect of the double-layer plasma-activated gel patch provided in this application on promoting wound healing in diabetic rats.

[0062] Control group: On day 3, the wound still had significant redness, swelling, and exudation, with a healing rate of only 15%; on day 6, the redness and swelling lessened, but the exudation did not completely disappear, with a healing rate of 30%; on day 9, a small amount of granulation tissue began to appear, with a healing rate of 55%; on day 12, the wound still had some unhealed areas, with a healing rate of 75%. Figure 5 The curve trends were consistent with those of the "control" group.

[0063] Example 1: Bilayer gel assembly: Corresponding Figure 5 The curves of the "double-layer gel" group showed that on the 3rd day, the redness and swelling of the wound significantly subsided, the exudation decreased, and the healing rate reached 35%; on the 6th day, the wound edges began to shrink and fresh granulation tissue appeared, with a healing rate of 60%; on the 9th day, the granulation tissue covered most of the wound, with a healing rate of 85%; and on the 12th day, the wound was basically completely healed, with a healing rate of >95%, and there were no signs of infection recurrence.

[0064] Comparative Example 2 Chemical Cross-linking Group: The healing rates from day 3 to day 12 were 20%, 40%, 65%, and 80%, respectively, which were lower than those in Example 1, and some rats showed slight redness and swelling in their wounds.

[0065] The gel samples from Example 1 and Comparative Example 2 were immersed in DMEM medium to prepare sample extracts. Mouse fibroblasts were seeded into 96-well plates and cultured for 24 hours. Different concentrations of extracts were then added, and the cells were cultured for another 24 hours. A chromogenic reagent was added and incubated for 4 hours. After discarding the supernatant, dimethyl sulfoxide was added to dissolve and crystallize the cells. The absorbance at 490 nm was measured using a microplate reader, and cell viability was calculated. The results showed that the cell viability of all concentration groups of the gel extract from Example 1 was >90%, while the cell viability of the 100% extract group from Comparative Example 2 was only 75%. This demonstrates that the physically freeze-thaw crosslinked gels in this application have no chemical residues and exhibit superior biosafety.

[0066] Three white rabbits were selected, and their backs were shaved to divide the area into three zones. The experimental zone was covered with the gel patch from Example 1, the negative control group was covered with sterile gauze, and the positive control group was covered with gauze soaked in 1% formaldehyde solution. The samples were removed after 24 hours, and skin reactions were observed at 1 hour, 24 hours, 48 ​​hours, and 72 hours. The results showed that no erythema or edema occurred in the experimental zone or the negative control group, while significant erythema and edema appeared in the positive control group, proving that the gel patch from Example 1 is non-irritating to the skin.

[0067] The dual-layer structure of Examples 1 and 2 solves the problem of the "single function" of the single-layer gel in Comparative Example 1 by sequentially releasing "antibacterial in the lower layer + healing-promoting in the upper layer". The dual-layer gel can control infection in the early stage of wound healing by releasing antibacterial active particles in the lower layer, inhibit excessive inflammation in the middle stage by releasing NO in the upper layer, and promote granulation tissue growth in the later stage, covering the entire healing cycle without the need to change dressings.

[0068] In Example 2, the addition of sodium alginate significantly improved the gel's permeability and mechanical properties, making it more suitable for the hyperosmolar environment of chronic wounds. In Example 3, mussel adhesive protein was used for modification, which greatly improved the adhesion performance and solved the problem of easy detachment of traditional gel patches. In contrast, the chemically cross-linked gel in Comparative Example 2 had residual cross-linking agents, which posed risks of cytotoxicity and skin irritation, highlighting the safety of physical freeze-thaw cross-linking.

[0069] The above embodiments are only used to illustrate the technical solutions of this application, and are not intended to limit this application. Although this application has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of this application.

Claims

1. A method for preparing a double-layer plasma-activated gel patch, characterized in that, include: Polyvinyl alcohol and polyethylene glycol are dissolved in deionized water, stirred and dissolved in deionized water at 90-95℃, and cooled to room temperature to obtain a matrix solution. The mass ratio of polyvinyl alcohol to polyethylene glycol is in the range of 1:0.8-1:1.

2. The matrix solution is enriched with... The plasma was used to activate the lower gel precursor solution to obtain the lower gel precursor solution. The lower gel precursor liquid is injected into the mold and subjected to 2-5 physical freeze-thaw crosslinking treatments to obtain the lower gel. The physical freeze-thaw crosslinking treatment is performed by freezing at a temperature of -20℃ to -80℃ for 8-12 hours and thawing at a temperature of 4℃ to room temperature for 4-8 hours. The matrix solution is activated with NO-rich plasma to obtain an upper gel precursor solution; the upper gel precursor solution is poured onto the surface of the lower gel, and after the physical freeze-thaw crosslinking treatment, a bilayer gel is obtained. The bilayer gel is immersed in a solution containing an adhesive polymer for 1-2 hours and then rinsed to obtain a bilayer plasma-activated gel patch.

2. The method according to claim 1, characterized in that, The matrix solution also includes a biocompatible polymer material, which includes at least one of sodium alginate, hyaluronic acid, sodium hyaluronate, chitosan, carboxylated chitosan, cellulose, cellulose derivatives, pectin, gelatin, and poloxamer.

3. The method according to claim 1, characterized in that, The working gas for the plasma activation treatment is a mixture of nitrogen and oxygen or air, the flow rate of the working gas is 0~5 slm, the activation treatment method is irradiation, gas blowing or bubbling, and the activation treatment time is 5-30 min.

4. The method according to claim 1, characterized in that, In the solution containing the adhesive polymer, the adhesive polymer is polydopamine, polyphenols, tannic acid, mussel adhesive protein, or a combination thereof.

5. The method according to claim 1, characterized in that, The rinsing is performed using deionized water, and the rinsing is repeated 2-3 times, with each rinsing lasting 1-2 minutes.

6. The method according to claim 1, characterized in that, The mold is made of polytetrafluoroethylene or glass, and the mold shape is round, square or irregular. The mold controls the pouring amount of the upper gel precursor liquid and the lower gel precursor liquid so that the thickness of the double-layer gel is 1-6mm.

7. The method according to claim 1, characterized in that, When the polyvinyl alcohol and polyethylene glycol are dissolved in deionized water at 90-95°C, the stirring speed is 300-500 r / min. After the lower gel precursor liquid is injected into the mold, the mold is sealed before physical freeze-thaw crosslinking treatment. When the double-layer gel is immersed in a solution containing adhesive polymer, it is completely submerged in the solution.

8. The method according to claim 1, characterized in that, The plasma generating device is a dielectric barrier discharge device or a sliding arc discharge device.

9. A double-layer plasma-activated gel patch prepared using the method described in any one of claims 1-8, characterized in that, include: The lower gel and the upper gel are both based on a polyvinyl alcohol-polyethylene glycol blend and are physically cross-linked to form an integral structure. The lower gel is loaded with rich Active particles from plasma conversion; The upper gel is loaded with NO-rich plasma-converted active particles, mainly NO. The bilayer gel, after being treated with a solution containing an adhesive polymer, has the property of adhering to the skin or wound.

10. An application of a double-layer plasma-activated gel patch prepared using the method described in any one of claims 1-8, characterized in that, include: The double-layer plasma-activated gel patch is used to prepare topical medications for skin wounds; The skin wounds include infected skin wounds and chronic wounds, wherein the chronic wounds are diabetic foot ulcers, venous ulcers, or pressure ulcers.

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