An integrated wound dressing with bacteriostatic and nursing functions and a preparation method thereof

Abstract

The application provides an integrated wound dressing with bacteriostasis and nursing functions and a preparation method thereof, and relates to the technical field of medical wound nursing materials. The integrated wound dressing with bacteriostasis and nursing functions comprises, from outside to inside, a medical adhesive base layer, a ROS sterilization layer, a sample directional flow guiding layer, an integrated detection layer and a transparent protective observation layer. The medical adhesive base layer is a composite structure of a low-sensitivity medical pressure-sensitive adhesive coating and a micro-permeable hole array. The thickness of the pressure-sensitive adhesive coating is 25-50 microns. The diameter of the micro-permeable hole is 50-150 microns, the hole spacing is 200-400 microns, and the opening rate is 15%-25%. Through the synergistic structure design of the five functional layers, the active oxygen photodynamic sterilization, the directional micro-fluidic flow guiding and the multi-index fluorescence immunoassay are integrated in the same dressing, the integrated operation of local bacteriostasis treatment of the wound and in-situ visual evaluation of the infection state is realized, and the technical prejudice that the active oxygen sterilization component interferes with the immunoassay is overcome.

Description

Technical Field

[0001] This invention relates to the field of medical wound care materials, specifically to an integrated wound dressing that combines antibacterial and nursing functions, and its preparation method. Background Technology

[0002] Chronic, non-healing wounds, including diabetic foot ulcers, pressure injuries, and venous ulcers, are characterized by prolonged course, difficulty in healing, and frequent recurrent infections, posing a significant challenge to clinical wound management. Existing wound care products exhibit the following prominent problems in practical application.

[0003] Firstly, existing wound dressings have limited functionality, with sterilization and detection being disconnected. Traditional moist wound dressings only have the basic functions of absorbing exudate and maintaining a moist environment, without any active bactericidal components. When a wound becomes infected, the dressing needs to be changed frequently and combined with systemic antibiotic treatment, which increases patient suffering and easily leads to the growth of drug-resistant bacteria. On the other hand, separate wound detection reagents or biosensors require the collection of wound exudate samples for testing, which cannot achieve immediate assessment at the bedside. Furthermore, the sampling process requires removing the dressing, causing mechanical damage to newly formed granulation tissue and introducing the risk of exogenous contamination.

[0004] Secondly, the release of active ingredients in existing bactericidal dressings lacks targeting and controllability. Broad-spectrum antibacterial products such as silver ion dressings and iodine-based dressings, while killing bacteria, also exhibit cytotoxicity to normal wound tissue cells, and the accumulation of silver ions in the wound may lead to silver deposition in the skin. Antibiotic-containing dressings face the risk of selection by drug-resistant bacteria, and their antibacterial effect diminishes with prolonged use. While photodynamic sterilization technology can avoid the problem of drug resistance, existing photosensitizers are mostly in a free state, easily diffusing and being lost in wound exudate, and the reactive oxygen species they generate may quench the signal of nearby immunoassay reagents. Those skilled in the art generally believe that integrating photodynamic sterilization materials with oxidative activity into the same dressing would lead to reagent inactivation due to reactive oxygen species diffusion; therefore, there is no technological inspiration for integrating the two.

[0005] Third, existing methods for assessing wound infection rely on laboratory testing, which is not timely. Clinically, wound infection is mainly assessed based on physical signs such as redness, swelling, heat, and pain, as well as serological indicators such as complete blood count, C-reactive protein, and procalcitonin. These indicators reflect the systemic inflammatory state rather than the local microenvironment of the wound, and the testing cycle is long and expensive, making continuous dynamic monitoring impossible. Although the local pH value of the wound can reflect the infection status, current pH testing methods mostly use colorimetric strips, which require removing the dressing and directly contacting the wound, making the operation inconvenient and lacking in quantitative accuracy.

[0006] Fourth, existing multilayer composite dressings suffer from low interlayer liquid transfer efficiency and poor detection consistency. Ordinary absorbent pad dressings passively absorb exudate through fiber capillary action, resulting in uneven liquid distribution between layers. This leads to insufficient sample volume or over-wetting in the detection area, resulting in poor repeatability of test results. At the same time, the bactericidal layer and the detection layer are not physically isolated, and the bactericidal active ingredients may interfere with the antigen-antibody binding reaction of the immunoassay reagent, causing false negatives or signal quenching. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention provides an integrated wound dressing that combines antibacterial and nursing functions, as well as its preparation method. This solves the problems of existing wound dressings, such as the separation of bactericidal and detection functions, uncontrollable release of active ingredients, poor timeliness of infection assessment, and uneven interlayer transport interfering with detection.

[0008] To achieve the above objectives, the present invention provides the following technical solution: an integrated wound dressing that combines antibacterial and nursing functions, comprising, sequentially arranged from the wound contact side outwards, a medical adhesive base layer, a ROS sterilization layer, a sample guiding layer, an integrated detection layer, and a transparent protective observation layer: The medical adhesive substrate is a composite structure of a low-sensitivity medical pressure-sensitive adhesive coating and a microporous array. The thickness of the pressure-sensitive adhesive coating is 25-50 micrometers, the diameter of the microporous pores is 50-150 micrometers, the pore spacing is 200-400 micrometers, and the porosity is 15%-25%. The ROS sterilization layer is a mesoporous nanoparticle composite hydrogel membrane loaded with photosensitive active ingredients, with a thickness of 100-300 micrometers, a particle size of 50-100 nanometers and a pore size of 2-5 nanometers, and a photosensitive active ingredient loading of 5%-15% of the dry weight of the ROS sterilization layer; The sample directional flow layer is a hydrophilic polymer film with built-in tree-like branched microchannels. The main channel width is 200-300 micrometers, the branch channel width is 100-150 micrometers, the channel depth is 50-100 micrometers, and the channel spacing is 200-500 micrometers. The integrated detection layer includes three independent PCT fluorescence immunoassay regions, IL-6 fluorescence immunoassay regions, and pH fluorescence detection regions, which are arranged side by side on the same nitrocellulose substrate. The transparent protective observation layer is a waterproof and breathable PET film with a thickness of 20-50 micrometers and a water vapor permeability of not less than 2000 grams per square meter per 24 hours.

[0009] Preferably, the mesoporous nanoparticles in the ROS sterilization layer are mesoporous silica nanoparticles, the photosensitive active ingredient is methylene blue, and the methylene blue is loaded on the inner and outer surfaces of the pores of the mesoporous silica through a dual mechanism of physical adsorption and Schiff base covalent bonding. The composite hydrogel matrix is ​​a crosslinking system of sodium alginate and gelatin with a mass ratio of 1:1 to 3:1, and the crosslinking agent is a calcium chloride solution with a mass fraction of 0.5% to 2%. The methylene blue is excited to generate singlet oxygen under 660 nm red light irradiation. The pore confinement effect of the mesoporous nanoparticles makes the singlet oxygen concentration in the diffusion-sample directional flow layer less than 1 μmol per liter.

[0010] Preferably, the tree-like branched microchannels of the sample directional flow layer adopt a hierarchical structure in which a primary main channel branches into four secondary branch channels, and each secondary branch channel further branches into two tertiary terminal channels. The inlet of the main channel is directly opposite the liquid outlet area of ​​the ROS sterilization layer, and the tertiary terminal channels are respectively connected to the three detection area inlets of the integrated detection layer. The inner wall of the microchannel is treated with oxygen plasma hydrophilic treatment, and the contact angle is less than 30 degrees. The tree-shaped branched microchannels adopt an equal flow resistance design, and the ratio of the cross-sectional area of ​​the main channel to the total cross-sectional area of ​​the branch channels is 1:1.02 to 1:1.08.

[0011] Preferably, the nitrocellulose membrane of the PCT fluorescent immunoassay region is coated with anti-human PCT polyclonal antibody as a detection line, and the upstream binding pad of the detection line is sprayed with europium ion chelate-labeled anti-human PCT monoclonal antibody. The nitrocellulose membrane of the IL-6 fluorescent immunoassay area is coated with anti-human IL-6 polyclonal antibody as a detection line, and the upstream binding pad of the detection line is sprayed with europium ion chelate-labeled anti-human IL-6 monoclonal antibody. The pH fluorescence detection area is a cellulose membrane with a composite indicator of fluorescein isothiocyanate and bromothymol blue immobilized. The europium ion chelate has an excitation wavelength of 340–365 nm and an emission wavelength of 610–620 nm.

[0012] Preferably, the semi-quantitative determination threshold of the PCT fluorescence immunoassay detection area is as follows: the fluorescence signal intensity corresponds to a PCT concentration of 0.5 ng / mL as the threshold for mild infection, 2 ng / mL as the threshold for moderate infection, and 10 ng / mL as the threshold for severe infection. The semi-quantitative determination thresholds for the IL-6 fluorescent immunoassay detection area are as follows: 50 picograms per milliliter of fluorescence signal intensity corresponds to a critical IL-6 concentration of mild infection, 200 picograms per milliliter corresponds to a critical IL-6 concentration of moderate infection, and 1000 picograms per milliliter corresponds to a critical IL-6 concentration of severe infection. The semi-quantitative determination thresholds for the pH fluorescence detection area are: pH less than 7.2 indicates a normal healing environment, pH between 7.2 and 7.5 indicates a mild infection, and pH greater than 7.5 indicates a moderate to severe infection.

[0013] Preferably, the micropores of the medical adhesive substrate are prepared by laser drilling, with a drilling power of 6-10 watts, a pulse width of 0.1-0.3 milliseconds, and a repetition frequency of 15-25 kHz. The transparent protective observation layer is fabricated with a conical microstructure array on its surface using a nanoimprinting process. The conical microstructure has a bottom diameter of 1–3 micrometers, a height of 4–6 micrometers, and a spacing of 3–5 micrometers. The hydrostatic pressure of the film is greater than 30 kPa.

[0014] Preferably, the photosensitive active ingredient in the ROS sterilization layer is curcumin, the mesoporous nanoparticles are mesoporous carbon nanoparticles, the composite hydrogel matrix is ​​a chitosan and polyvinyl alcohol crosslinking system, and the fluorescent marker in the integrated detection layer is CdSe / ZnS quantum dots with an emission wavelength of 600-630 nm.

[0015] A method for preparing an integrated wound dressing that combines antibacterial and nursing functions includes the following steps: Step S1, Preparation of medical adhesive base layer: A low-sensitivity acrylic pressure-sensitive adhesive with a thickness of 25-50 micrometers is coated on a polyurethane elastic pad. A microporous array with a diameter of 50-150 micrometers, a spacing of 200-400 micrometers, and an open area of ​​15%-25% is prepared on the pressure-sensitive adhesive surface using a laser drilling process. After drilling, the surface is ultrasonically cleaned with 75% ethanol for 3-8 minutes. Step S2, Preparation of ROS sterilization layer: Synthesize mesoporous nanoparticles and load them with photosensitive active ingredients. Disperse the loaded mesoporous nanoparticles in a hydrogel matrix solution, and obtain a ROS sterilization layer with a thickness of 100-300 micrometers by cross-linking curing and freeze-drying. Step S3, Preparation of sample directional flow layer: Dendritic branched microchannels are prepared on the surface of polymer film by photolithography and etching process, and oxygen plasma hydrophilization treatment is performed after resist removal; Step S4, Preparation of integrated detection layer: Three independent detection areas are separated on the nitrocellulose membrane to prepare PCT fluorescence immunoassay area, IL-6 fluorescence immunoassay area and pH value fluorescence detection area respectively; Step S5, Composite Assembly and Sterilization: Align and stack the five layers in sequence, apply medical-grade pressure-sensitive adhesive to the non-circulation areas between the layers, heat-press the composite, then cut, sterilize, and seal the packaging.

[0016] Preferably, step S2 includes the following sub-steps: Step S2-1, Synthesis of mesoporous nanoparticles: Using hexadecyltrimethylammonium bromide as a template agent and tetraethyl orthosilicate as a silicon source, the reaction was carried out in a water-ethanol mixture catalyzed by ammonia at 55-65 degrees Celsius for 1.5-2.5 hours with stirring. After centrifugation and washing, the template agent was removed by calcination at 500-600 degrees Celsius for 4-6 hours to obtain mesoporous silica nanoparticles with a particle size of 50-100 nanometers and a pore size of 2-5 nanometers. Step S2-2, loading of photosensitive active ingredient: Mesoporous silica nanoparticles are dispersed in a methylene blue phosphate buffer solution with a mass fraction of 0.5–2 mg / mL. The mixture is shaken and adsorbed at 20–30°C for 10–14 hours under light-protected conditions. After centrifugation and washing, the mixture is activated with 3-aminopropyltriethoxysilane, so that methylene blue is loaded through a dual mechanism of physical adsorption and covalent bonding with Schiff base. The loading amount is 5%–15% of the mass of the mesoporous nanoparticles. Step S2-3, hydrogel composite molding: Mesoporous nanoparticles loaded with methylene blue are dispersed in a mixed solution of sodium alginate and gelatin at a mass fraction of 2% to 4%, with a mass ratio of sodium alginate to gelatin of 1:1 to 3:1. After casting, the mixture is immersed in a calcium chloride solution at a mass fraction of 0.5% to 2% for crosslinking for 10 to 20 minutes, rinsed, and then freeze-dried.

[0017] Preferably, step S4 includes the following sub-steps: Step S4-1, detection area separation: Separate the PCT detection area, IL-6 detection area and pH detection area by printing spacer lines on the nitrocellulose membrane with hydrophobic wax lines. Step S4-2, preparation of the immunodetection zone: Anti-human PCT polyclonal antibody and anti-human IL-6 polyclonal antibody were diluted with phosphate buffer to 0.8-1.2 mg / mL, respectively, and streaked onto the corresponding nitrocellulose membrane as detection lines. After drying at 37°C for 1-3 hours, the membrane was blocked with 1% bovine serum albumin. Europium ion chelate-labeled monoclonal antibodies were then sprayed onto the conjugate pads and vacuum dried at 37°C for 3-5 hours. Step S4-3, pH detection zone preparation: Dissolve fluorescein isothiocyanate and bromothymol blue in a mass ratio of 1:1 to 1:3 in a borate buffer solution with a pH of 7.5 to 8.5, with a total concentration of 0.3 to 0.8 mg per milliliter. Immerse the cellulose membrane in the solution for 3 to 8 minutes, then remove it, dry it at 37 degrees Celsius in the dark, and cut and fix it in the pH detection zone.

[0018] This invention provides an integrated wound dressing that combines antibacterial and nursing functions, as well as its preparation method. It has the following beneficial effects: 1. This invention achieves an integrated nursing effect by layering and combining a ROS photodynamic sterilization layer, a sample directional flow layer, and a three-index integrated detection layer into an integrated dressing structure, and combining it with a transparent protective observation layer for non-removable visualization design, thereby realizing in-situ antibacterial and real-time assessment of infection status in chronic and difficult-to-heal wounds.

[0019] 2. This invention achieves a safe antibacterial effect by physically adsorbing and covalently binding photosensitive active ingredients with mesoporous nanoparticles and co-loading them with Schiff bases, combined with the confined and sustained-release effect of the mesoporous channels, thus achieving targeted immobilization and controlled release of bactericidal ingredients while avoiding the risks of drug resistance and toxicity to normal tissue cells.

[0020] 3. This invention achieves rapid, visual assessment of the local microenvironment of the wound at the bedside and without the need to remove dressings for sampling by using a parallel detection design of PCT and IL-6 dual-index fluorescence immunochromatography and pH fluorescence colorimetry, combined with semi-quantitative threshold grading colorimetric determination.

[0021] 4. This invention utilizes a tree-like branched microchannel design with equal flow resistance for graded and directional delivery, combined with the physical barrier isolation of the polymer substrate of the flow guide layer, to accurately and equally distribute the sterilization exudate to each independent detection area, while preventing the diffusion of sterilization active ingredients from interfering with the stable detection effect of the immunoassay reagent. Attached Figure Description

[0022] Figure 1 This is an exploded view of the layered structure of the wound dressing of the present invention; Figure 2 This is a schematic diagram of the tree-like branched microchannel structure of the sample directional flow layer of the present invention; Figure 3 This is a schematic diagram of the arrangement of the three detection zones in the integrated detection layer of the present invention; Figure 4 This is a flowchart of the integrated wound dressing preparation process of the present invention; Figure 5 This is a comparison chart of the antibacterial rates of embodiments and comparative examples of the present invention; Figure 6 This is a standard curve of PCT and IL-6 fluorescence detection according to the present invention.

[0023] The components include: 1. Medical adhesive base layer; 2. ROS sterilization layer; 3. Sample directional flow layer; 4. Integrated detection layer; and 5. Transparent protective observation layer. Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] Example: like Figure 1-6 As shown, this embodiment of the invention provides an integrated wound dressing that combines antibacterial and nursing functions. It includes a medical adhesive base layer 1, a ROS sterilization layer 2, a sample guiding layer 3, an integrated detection layer 4, and a transparent protective observation layer 5, arranged sequentially from the wound contact side outwards. The medical adhesive base layer 1 is a composite structure of a low-sensitivity medical pressure-sensitive adhesive coating and a microporous array. The pressure-sensitive adhesive coating has a thickness of 25–50 micrometers, the microporous pores have a diameter of 50–150 micrometers, a pore spacing of 200–400 micrometers, and an open porosity of 15%–25%. The microporous pores of the medical adhesive base layer 1 are prepared using a laser drilling process with a drilling power of 6–10 watts, a pulse width of 0.1–0.3 milliseconds, and a repetition frequency of 15–25 kilohertz. ROS sterilization layer 2 is a mesoporous nanoparticle composite hydrogel membrane loaded with photosensitive active ingredients, with a thickness of 100–300 micrometers. The mesoporous nanoparticles have a particle size of 50–100 nanometers and a pore size of 2–5 nanometers. The loading of photosensitive active ingredients is 5%–15% of the dry weight of ROS sterilization layer 2. The mesoporous nanoparticles in ROS sterilization layer 2 are mesoporous silica nanoparticles, and the photosensitive active ingredient is methylene blue. Methylene blue is loaded on the inner and outer surfaces of the pores of the mesoporous silica through a dual mechanism of physical adsorption and Schiff base covalent bonding. The composite hydrogel matrix has a mass ratio of 1:1–3: The sodium alginate and gelatin crosslinking system of layer 1 uses calcium chloride solution with a mass fraction of 0.5% to 2% as the crosslinking agent. Methylene blue is excited to generate singlet oxygen under 660 nm red light irradiation. The pore confinement effect of the mesoporous nanoparticles makes the singlet oxygen concentration in the diffusion-sample directional flow layer 3 less than 1 μmol / L. The photosensitive active ingredient in the ROS sterilization layer 2 is curcumin, and the mesoporous nanoparticles are mesoporous carbon nanoparticles. The composite hydrogel matrix is ​​a crosslinking system of chitosan and polyvinyl alcohol. The fluorescent label of the integrated detection layer 4 is CdSe / ZnS quantum dots with an emission wavelength of 600 to 630 nm. The sample directional flow guide layer 3 is a hydrophilic polymer film with built-in tree-like branched microchannels. The main channel width is 200-300 micrometers, the branch channel width is 100-150 micrometers, the channel depth is 50-100 micrometers, and the channel spacing is 200-500 micrometers. The tree-like branched microchannels of the sample directional flow guide layer 3 adopt a hierarchical structure in which the primary main channel branches into four secondary branch channels, and each secondary branch channel further branches into two tertiary terminal channels. The main channel inlet is directly opposite the liquid outlet area of ​​the ROS sterilization layer 2, and the tertiary terminal channels are connected to the three detection area inlets of the integrated detection layer 4 one by one. The inner wall of the microchannel is treated with oxygen plasma hydrophilization, and the contact angle is less than 30 degrees. The tree-like branched microchannels adopt an equal flow resistance design, and the ratio of the cross-sectional area of ​​the main channel to the total cross-sectional area of ​​the branch channels is 1:1.02 to 1:1.08. The integrated detection layer 4 includes three independent detection zones: a PCT fluorescence immunoassay zone, an IL-6 fluorescence immunoassay zone, and a pH fluorescence detection zone. These three zones are arranged side-by-side on the same nitrocellulose substrate. The PCT fluorescence immunoassay zone has an anti-human PCT polyclonal antibody coated on its nitrocellulose membrane as a detection line. An anti-human PCT monoclonal antibody labeled with europium ion chelates is sprayed onto the upstream conjugate pad of the detection line. Similarly, the IL-6 fluorescence immunoassay zone has an anti-human IL-6 polyclonal antibody coated on its nitrocellulose membrane as a detection line. An anti-human IL-6 monoclonal antibody labeled with europium ion chelates is sprayed onto the upstream conjugate pad of the detection line. The pH fluorescence detection zone is a cellulose membrane immobilized with a fluorescein isothiocyanate and bromothymol blue composite indicator. The excitation wavelength of the europium ion chelate is [wavelength value missing]. The emission wavelength is 610–620 nm. The semi-quantitative thresholds for the PCT fluorescence immunoassay area are: 0.5 ng / mL for mild infection, 2 ng / mL for moderate infection, and 10 ng / mL for severe infection. The semi-quantitative thresholds for the IL-6 fluorescence immunoassay area are: 50 picograms / mL for mild infection, 200 picograms / mL for moderate infection, and 1000 picograms / mL for severe infection. The semi-quantitative thresholds for the pH fluorescence immunoassay area are: pH less than 7.2 indicates a normal healing environment, pH between 7.2 and 7.5 indicates mild infection, and pH greater than 7.5 indicates moderate to severe infection. The transparent protective observation layer 5 is a waterproof and breathable PET film with a thickness of 20-50 micrometers and a water vapor permeability of not less than 2000 grams per square meter per 24 hours. The surface of the transparent protective observation layer 5 is prepared with a conical microstructure array by nanoimprinting process. The bottom diameter of the conical microstructure is 1-3 micrometers, the height is 4-6 micrometers, the spacing is 3-5 micrometers, and the hydrostatic pressure of the film is greater than 30 kPa.

[0026] A method for preparing an integrated wound dressing that combines antibacterial and nursing functions includes the following steps: Step S1, Preparation of medical adhesive base layer: A low-sensitivity acrylic pressure-sensitive adhesive with a thickness of 25-50 micrometers is coated on a polyurethane elastic pad. A microporous array with a diameter of 50-150 micrometers, a spacing of 200-400 micrometers, and an open area of ​​15%-25% is prepared on the pressure-sensitive adhesive surface using a laser drilling process. After drilling, the surface is ultrasonically cleaned with 75% ethanol for 3-8 minutes. Step S2, Preparation of ROS sterilization layer: Synthesize mesoporous nanoparticles and load them with photosensitive active ingredients. Disperse the loaded mesoporous nanoparticles in a hydrogel matrix solution, and obtain a ROS sterilization layer 2 with a thickness of 100-300 micrometers by cross-linking curing and freeze drying. Step S2-1, Synthesis of mesoporous nanoparticles: Using hexadecyltrimethylammonium bromide as a template agent and tetraethyl orthosilicate as a silicon source, the reaction was carried out in a water-ethanol mixture catalyzed by ammonia at 55-65 degrees Celsius for 1.5-2.5 hours with stirring. After centrifugation and washing, the template agent was removed by calcination at 500-600 degrees Celsius for 4-6 hours to obtain mesoporous silica nanoparticles with a particle size of 50-100 nanometers and a pore size of 2-5 nanometers. Step S2-2, loading of photosensitive active ingredient: Mesoporous silica nanoparticles are dispersed in a methylene blue phosphate buffer solution with a mass fraction of 0.5–2 mg / mL. The mixture is shaken and adsorbed at 20–30°C for 10–14 hours under light-protected conditions. After centrifugation and washing, the mixture is activated with 3-aminopropyltriethoxysilane, so that methylene blue is loaded through a dual mechanism of physical adsorption and covalent bonding with Schiff base. The loading amount is 5%–15% of the mass of the mesoporous nanoparticles. Step S2-3, hydrogel composite molding: Mesoporous nanoparticles loaded with methylene blue are dispersed in a mixed solution of sodium alginate and gelatin at a mass fraction of 2% to 4%, with a mass ratio of sodium alginate to gelatin of 1:1 to 3:1. After casting and molding, the mixture is immersed in a calcium chloride solution at a mass fraction of 0.5% to 2% for crosslinking for 10 to 20 minutes, rinsed, and then freeze-dried. Step S3, Preparation of sample directional flow layer: Dendritic branched microchannels are prepared on the surface of polymer film by photolithography and etching process, and oxygen plasma hydrophilization treatment is performed after resist removal; Step S4, Preparation of integrated detection layer: Three independent detection areas are separated on the nitrocellulose membrane to prepare PCT fluorescence immunoassay area, IL-6 fluorescence immunoassay area and pH value fluorescence detection area respectively; Step S4-1, detection area separation: Separate the PCT detection area, IL-6 detection area and pH detection area by printing spacer lines on the nitrocellulose membrane with hydrophobic wax lines. Step S4-2, preparation of the immunodetection zone: Anti-human PCT polyclonal antibody and anti-human IL-6 polyclonal antibody were diluted with phosphate buffer to 0.8-1.2 mg / mL, respectively, and streaked onto the corresponding nitrocellulose membrane as detection lines. After drying at 37°C for 1-3 hours, the membrane was blocked with 1% bovine serum albumin. Europium ion chelate-labeled monoclonal antibodies were then sprayed onto the conjugate pads and vacuum dried at 37°C for 3-5 hours. Step S4-3, pH detection zone preparation: Dissolve fluorescein isothiocyanate and bromothymol blue in a mass ratio of 1:1 to 1:3 in a borate buffer solution with a pH of 7.5 to 8.5, with a total concentration of 0.3 to 0.8 mg per milliliter. Immerse the cellulose membrane in the solution for 3 to 8 minutes, then remove it, dry it at 37 degrees Celsius in the dark, and cut and fix it in the pH detection zone. Step S5, Composite Assembly and Sterilization: Align and stack the five layers in sequence, apply medical-grade pressure-sensitive adhesive to the non-circulation areas between the layers, heat-press the composite, then cut, sterilize, and seal the packaging.

[0027] Example 1 This embodiment provides an integrated wound dressing that combines antibacterial and nursing functions. The overall dimensions are 10 cm in length and 8 cm in width. From bottom to top, it is composed of a medical adhesive base layer 1, a ROS sterilization layer 2, a sample guiding layer 3, an integrated detection layer 4, and a transparent protective observation layer 5, which are hot-pressed together. The layers are bonded point-to-point with medical-grade polyurethane acrylate pressure-sensitive adhesive, and the bonding area avoids the liquid flow path.

[0028] The medical adhesive base layer 1 is made of a 38-micron thick low-sensitivity acrylic pressure-sensitive adhesive coated on a polyurethane elastic pad. Cylindrical micro-permeable pores are evenly distributed on the pressure-sensitive adhesive surface. The pore diameter is 100 microns, the pore spacing is 300 microns, and the porosity is 20%. The micro-permeable pores are prepared by laser drilling process with a drilling power of 8 watts, a pulse width of 0.2 milliseconds, and a repetition frequency of 20 kHz. After drilling, the molten residue is removed by ultrasonic cleaning with 75% ethanol for 5 minutes.

[0029] The preparation process of ROS sterilization layer 2 is as follows: First, mesoporous silica nanoparticles are synthesized using the sol-gel method. Hexadecyltrimethylammonium bromide is used as a template agent, and tetraethyl orthosilicate is used as the silicon source. The reaction is carried out in a water-ethanol mixture catalyzed by ammonia at 60°C for 2 hours. After centrifugation, washing, and calcination at 550°C for 5 hours to remove the template agent, mesoporous silica nanoparticles with a particle size of approximately 80 nm, a pore size of approximately 3.5 nm, and a specific surface area of ​​approximately 850 m² / g are obtained. The mesoporous silica nanoparticles are then dispersed in a methylene blue phosphate buffer solution with a mass fraction of 1 mg / mL. Adsorption is carried out under light-protected conditions at 25°C with shaking for 12 hours. After centrifugation, the adsorption is... Unadsorbed free methylene blue was removed by washing with phosphate buffer. After activation with 3-aminopropyltriethoxysilane, it formed a Schiff base covalently bonded to the phenolic hydroxyl groups of methylene blue, resulting in a final methylene blue loading of 10% of the mass of mesoporous silica. The methylene blue-loaded mesoporous silica nanoparticles were dispersed at 3% by mass in a mixed solution of 2% by mass sodium alginate and 2% by mass gelatin (sodium alginate to gelatin mass ratio of 2:1). After stirring evenly, the mixture was poured into a polytetrafluoroethylene mold and immersed in a 1% by mass calcium chloride solution for crosslinking for 15 minutes. After removal, it was rinsed with deionized water and freeze-dried to obtain a ROS sterilization layer 2 with a thickness of approximately 200 micrometers. After being irradiated with 660 nm red light for 5 minutes at a power density of 50 mW / cm², the singlet oxygen generation of the ROS sterilization layer 2 reached a plateau value. Due to the confinement effect of the mesoporous channels, the singlet oxygen concentration diffused to the outer surface of the ROS sterilization layer 2 was less than 0.8 μmol / L.

[0030] The sample directional flow layer 3 uses a 150-micron-thick polyethylene terephthalate (PET) film as the substrate. Dendritic branched microchannels are fabricated using UV lithography and reactive ion etching (RIE) processes: First, a 5-micron-thick positive photoresist is spin-coated onto the PET film surface, pre-baked at 90 degrees Celsius for 30 seconds, then exposed to UV light through a mask for 12 seconds and developed for 30 seconds to obtain the microchannel pattern. Next, reactive oxygen species (ROS) etching is performed for 120 seconds to an etching depth of 80 microns. After removing the photoresist, the microchannel structure is obtained. The microchannels branch from a primary main channel. A hierarchical structure consisting of four secondary branch channels, each further branching into two tertiary terminal channels. The main channel width is 250 micrometers, the secondary branch channel width is 150 micrometers, and the tertiary terminal channel width is 100 micrometers. The channel depth is 80 micrometers, and the channel spacing is 300 micrometers. After etching, the sample was placed in an oxygen plasma treatment instrument with a directional flow layer 3 and treated for 60 seconds at a power of 100 watts and an oxygen flow rate of 50 standard milliliters per minute, reducing the contact angle of the microchannel inner wall to below 25 degrees. This tree-like branching structure employs an equal flow resistance design, with the ratio of the cross-sectional area of ​​the main channel to the total cross-sectional area of ​​the branch channels being 1:1.05.

[0031] The integrated detection layer 4 uses a nitrocellulose membrane with a pore size of 5 micrometers as a substrate. Three independent detection zones are arranged horizontally side by side on the substrate: a PCT fluorescence immunoassay zone, an IL-6 fluorescence immunoassay zone, and a pH fluorescence detection zone. Each zone is separated by a 1 mm wide hydrophobic wax line. Preparation of the PCT detection zone: Anti-human PCT polyclonal antibody was diluted to 1 mg / mL with phosphate buffer and applied as a detection line on the nitrocellulose membrane using a three-dimensional spotting apparatus at a rate of 1 μL / cm. After drying at 37°C for 2 hours, it was blocked with 1% bovine serum albumin for 1 hour. The binding pad is made of glass cellulose membrane, coated with europium ion chelate-labeled anti-human PCT monoclonal antibody at a concentration of 0.5 mg / mL and a coating amount of 2 μL / cm². It was then vacuum dried at 37°C for 4 hours. The preparation method of the IL-6 detection area is the same as that of the PCT detection area, except that the antibody is replaced with anti-human IL-6 polyclonal antibody and europium-labeled anti-human IL-6 monoclonal antibody. Preparation of the pH fluorescence detection area: Fluorescein isothiocyanate and bromothymol blue are dissolved in borate buffer at a mass ratio of 1:2 to 8.0, with a total concentration of 0.5 mg / mL. Quantitative filter paper is immersed in the solution for 5 minutes, then removed and dried at 37°C in the dark. It is then cut into 5 mm x 5 mm membranes and fixed to the detection area.

[0032] The transparent protective observation layer 5 uses a 30-micron thick transparent PET film. The surface is prepared with a conical microstructure array through nanoimprinting to achieve waterproof and breathable properties. The bottom diameter of the conical microstructure is 2 microns, the height is 5 microns, and the spacing is 4 microns. The water vapor permeability of the film is 2800 grams per square meter per 24 hours, and the hydrostatic pressure is greater than 30 kPa.

[0033] The five layers are aligned and stacked in sequence. Medical-grade polyurethane acrylate pressure-sensitive adhesive is applied to the non-circulating areas between the layers. The layers are then hot-pressed for 30 seconds at a temperature of 60 degrees Celsius and a pressure of 0.3 MPa. The mixture is then cut to the specified size and sterilized with ethylene oxide at a temperature of 37 degrees Celsius, a relative humidity of 60%, an EO concentration of 600 mg / L, and a sterilization time of 4 hours. After 7 days of analysis, the mixture is sealed and packaged.

[0034] The dressing in this embodiment is used as follows: the medical adhesive base layer 1 is applied to the intact skin around the wound, so that the microporous area covers the wound. The wound exudate permeates upward through the microporous area into the ROS sterilization layer 2. During use, the ROS sterilization layer 2 area is irradiated with 660 nm red light through the transparent protective observation layer 5 for 5 minutes to activate methylene blue and generate singlet oxygen to complete local antibacterial action. The sterilized exudate is transported to three detection areas through the tree-like microchannels of the sample directional flow layer 3. After 15 minutes, the fluorescence image is observed with the naked eye through the transparent protective observation layer 5 or captured by a mobile phone under a 365 nm ultraviolet light source. The concentrations of PCT and IL-6 and the pH value are semi-quantitatively determined by comparing with the colorimetric card, thereby assessing the degree of wound infection and healing status.

[0035] Example 2 The difference between this embodiment and Embodiment 1 is as follows: the photosensitive active ingredient in the ROS sterilization layer is replaced by curcumin instead of methylene blue, the mesoporous nanoparticles are replaced by mesoporous carbon nanoparticles instead of mesoporous silica, the curcumin loading is 8%, the composite hydrogel matrix is ​​a chitosan and polyvinyl alcohol crosslinking system, and the crosslinking agent is a 2% (w / w) mixed solution of sodium hydroxide and anhydrous ethanol; the sample directional flow layer uses a polydimethylsiloxane casting process instead of a PET etching process, using a silicon wafer etching master plate as a template, the PDMS prepolymer and curing agent are mixed at a mass ratio of 10:1 and cast onto the master plate, cured at 80 degrees Celsius for 2 hours and then peeled off, the microchannel depth is 100 micrometers; the remaining structure and parameters are the same as in Embodiment 1.

[0036] Example 3 The difference between this embodiment and Example 1 is as follows: the fluorescent label of the integrated detection layer uses CdSe / ZnS quantum dots with an emission wavelength of 620 nm, replacing europium ion chelates; the antibody coating concentration of the detection lines in the PCT and IL-6 detection areas is adjusted to 1.5 mg / mL; the quantum dot-labeled antibody is prepared by activating carboxyl quantum dots with carbodiimide hydrochloride and then coupling them with the amino group of the antibody; the pH detection area uses a phenol red and Nile blue composite indicator instead of fluorescein isothiocyanate and bromothymol blue; the remaining structures and parameters are the same as in Example 1.

[0037] Comparative Example 1 This comparative example uses a commercially available, ordinary moist wound dressing, specifically a calcium alginate fiber dressing. It does not include the ROS sterilization layer 2, sample-directing layer 3, or integrated detection layer 4; it consists only of a calcium alginate fiber absorbent layer and a non-woven fabric backing layer. The calcium alginate fiber layer is made of calcium alginate fibers through a hydroentangling process, with a fiber diameter of 15 to 25 micrometers, a fiber weight of 120 grams per square meter, and a thickness of 3 millimeters. The non-woven fabric backing layer is made of polypropylene spunbond non-woven fabric with a weight of 30 grams per square meter. The liquid absorption mechanism of this dressing involves ion exchange between the calcium alginate fibers and sodium ions in the wound exudate, transforming it into sodium alginate gel to absorb and lock in the exudate. The saturated absorbency is approximately 18 times its own dry weight. This dressing does not contain any active bactericidal ingredients and does not have infection indicator detection capabilities; it only maintains a moist wound environment. Under the same test conditions as in Example 1, the dressing showed inhibition rates of 12.3%, 8.7%, and 6.5% against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, respectively, and could not provide any assessment information on the wound infection status.

[0038] Comparative Example 2 This comparative example uses a commercially available silver ion antibacterial dressing, specifically a nano-silver-loaded sodium carboxymethyl cellulose dressing. It comprises a silver ion bactericidal layer and a waterproof and breathable backing layer, but excludes the sample-directing flow layer 3 and the integrated detection layer 4. The bactericidal layer is prepared by dissolving sodium carboxymethyl cellulose in deionized water to a 3% (w / w) solution, adding nano-silver sol (20-50 nm particle size, 1000 μg / mL silver content) to achieve a final silver loading of 0.5% of the dressing's dry weight. After thorough mixing, the mixture is poured and freeze-dried to obtain a 2 mm thick silver ion bactericidal layer. The backing layer is a 25 μm thick polyurethane breathable film. This dressing achieves broad-spectrum antibacterial activity through the sustained release of silver ions, but the silver ions act indiscriminately on both bacteria and normal tissue cells, inhibiting fibroblasts in the wound while simultaneously killing bacteria. Under the same test conditions as in Example 1, the dressing showed inhibition rates of 98.2%, 97.5%, and 95.8% against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, respectively. However, the cytotoxicity test showed that the cell survival rate of its extract was only 71.3%, the toxicity grade was 2, and it did not have infection detection function.

[0039] Comparative Example 3 This comparative example removes the sample-directing flow layer from Example 1, allowing the ROS sterilization layer 2 to directly contact the integrated detection layer 4. The structure, materials, parameters, and preparation methods of all other layers are identical to those in Example 1. Due to the lack of the physical barrier and microchannel-based directional delivery function of the sample-directing flow layer 3, wound exudate, after being sterilized by the ROS sterilization layer 2, diffuses freely on the nitrocellulose membrane surface of the integrated detection layer. Liquid distribution is uncontrolled, resulting in significant differences in the influent time and volume among the three detection zones. Simultaneously, trace amounts of free methylene blue unloaded by mesoporous nanoparticles in the ROS sterilization layer 2 can directly diffuse into the detection zone, non-specifically binding to antibodies on the detection line and generating background fluorescence interference within the fluorescence detection wavelength range. Under the same test conditions as in Example 1, the antibacterial rate of this comparative example was comparable to that of Example 1, with antibacterial rates of 99.88%, 99.60%, and 99.22% against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, respectively. However, the clinical concordance rate of PCT detection dropped to 76.7%, the concordance rate of IL-6 detection dropped to 73.3%, the deviation of liquid volume in the three detection areas was greater than 35%, and the time for liquid to reach the detection area fluctuated randomly between 60 and 120 seconds.

[0040] Comparative Example 4 This comparative example, based on Example 1, replaces the methylene blue in ROS sterilization layer 2 with an equal mass of free methylene blue. That is, instead of using mesoporous silica nanoparticles for loading, methylene blue powder is directly dissolved and dispersed at a mass fraction of 0.3% in a mixed solution of sodium alginate and gelatin. ROS sterilization layer 2 is prepared using the same calcium chloride crosslinking and freeze-drying process. The structure, material, parameters, and preparation methods of the remaining layers are completely identical to those in Example 1. Because the methylene blue is not immobilized by mesoporous nanoparticles, it is only physically entangled and retained in the hydrogel network. Upon contact with wound exudate, a burst release effect occurs, with a large amount of methylene blue diffusing and being lost with the exudate within the first hour. After 24 hours, the residual amount of methylene blue in ROS sterilization layer 2 is less than 15% of the initial amount. Simultaneously, the free methylene blue lacks the confined protection of mesoporous channels, and the singlet oxygen it generates can freely diffuse to the downstream integrated detection layer 4, causing oxidative damage to the immunoassay reagents. Under the same test conditions as in Example 1, the 24-hour inhibition rates of Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa in this comparative example decreased to 85.6%, 78.3%, and 71.2%, respectively, and the PCT detection signal intensity decreased by 22% compared to Example 1, verifying the sustained-release immobilization and pore confinement effect of the dual loading of mesoporous nanoparticles on the active ingredients.

[0041] Comparative Example 5 This comparative example differs from Example 1 by removing the ROS sterilization layer 2 and directly contacting the medical adhesive base layer 1 with the sample directional flow layer 3. The structure, materials, parameters, and preparation methods of all other layers are identical to those in Example 1. This comparative example retains the directional flow and three-index fluorescence detection functions but lacks active sterilization capability; wound exudate enters the detection area directly without antibacterial treatment. Under the same test conditions as Example 1, the antibacterial rates against the three pathogens in this comparative example are comparable to those in Example 1, at 10.8%, 7.2%, and 5.9%, respectively. Due to the large number of viable bacteria and their metabolites in the exudate entering the detection area, the antibody activity on the nitrocellulose membrane is affected, resulting in a PCT detection concordance rate of 83.3% and an IL-6 detection concordance rate of 80.0%, both lower than in Example 1. This comparative example is used to verify the necessity of the ROS sterilization layer in integrated dressings, demonstrating that sterilization is not only an independent functional requirement but also improves the accuracy of downstream detection by reducing interference from viable bacteria and metabolites in the exudate.

[0042] Comparative Example 6 This comparative example replaces the tree-like branched microchannels of the sample directional flow layer 3 with a parallel straight channel structure, based on Example 1. Specifically, it uses eight parallel straight channels, each 150 micrometers wide, 80 micrometers deep, and spaced 300 micrometers apart. The channel inlets are evenly distributed below the liquid outlet area of ​​the ROS sterilization layer 2, and the outlets are evenly distributed above the integrated detection layer 4. The structure, materials, parameters, and preparation methods of the remaining layers are completely identical to those in Example 1. The parallel straight channel structure lacks the equal flow resistance distribution characteristic of the tree-like branched structure. Due to processing errors and differences in inlet position, the flow resistance of each channel is inconsistent. Furthermore, the straight channel outlets cannot precisely correspond to the inlets of the three detection zones, with some channel outlets falling on the hydrophobic gaps between the detection zones, resulting in liquid waste and insufficient liquid intake in the detection zones. Under the same test conditions as Example 1, the liquid intake deviation of the three detection zones in this comparative example was 28%, the liquid arrival time deviation was 25 seconds, the intra-batch coefficient of variation for PCT detection increased to 9.6%, and the intra-batch coefficient of variation for IL-6 detection increased to 11.2%, verifying the crucial role of the equal flow resistance design of the tree-like branched microchannels in ensuring detection consistency.

[0043] Performance test and verification results I. Antibacterial Performance Test Referring to GB / T20944.3-2008 "Evaluation of Antimicrobial Properties of Textiles - Part 3: Shaking Method", the antibacterial rates of Staphylococcus aureus (ATCC6538), Escherichia coli (ATCC25922), and Pseudomonas aeruginosa (ATCC27853) were tested in Examples 1 to 3 and Comparative Examples 1 to 6. The contact time between the samples and the bacterial solution was 24 hours, and the test temperature was 37 degrees Celsius. During the contact period, the Example groups and Comparative Examples 3, 4, 5, and 6 were all irradiated with 660 nm red light for 5 minutes (except for Comparative Examples 1 and 2). The results are as follows: Example 1 showed an inhibition rate of 99.92% against Staphylococcus aureus, 99.68% against Escherichia coli, and 99.35% against Pseudomonas aeruginosa; Example 2 showed inhibition rates of 99.87%, 99.52%, and 99.18% against the same three bacteria; Example 3 showed inhibition rates of 99.90%, 99.61%, and 99.27% ​​against the same three bacteria; Comparative Example 1 showed inhibition rates of 12.3%, 8.7%, and 6.5% against the same three bacteria; Comparative Example 2 showed inhibition rates of 98.2%, 97.5%, and 95.8% against the same three bacteria; Comparative Example 3 showed inhibition rates of 99.88%, 99.60%, and 99.22% against the same three bacteria; Comparative Example 4 showed inhibition rates of 85.6%, 78.3%, and 71.2% against the same three bacteria; Comparative Example 5 showed inhibition rates of 10.8%, 7.2%, and 5.9% against the same three bacteria; and Comparative Example 6 showed inhibition rates of 99.85%, 99.55%, and 99.10% against the same three bacteria.

[0044] The above results show that the antibacterial rate of the embodiments of the present invention is significantly better than that of ordinary dressings, free methylene blue dressings, and dressings without a sterilizing layer, and is comparable to or even better than that of silver ion dressings, while avoiding the risks of cytotoxicity and drug resistance of silver ions; the antibacterial rate of Comparative Example 4 decreased significantly after 24 hours due to the sudden release and loss of free methylene blue, which verifies the sustained release and immobilization effect of mesoporous nanoparticle loading on active ingredients; the antibacterial rate of Comparative Example 6 is close to that of Example 1, indicating that the structure of the directional flow layer 3 of the sample does not affect the sterilization effect, and the difference is mainly reflected in the detection performance.

[0045] II. Detection Performance Verification Standard solutions with PCT concentrations of 0.1, 0.5, 1, 2, 5, 10, and 20 ng / mL, standard solutions with IL-6 concentrations of 10, 50, 100, 200, 500, 1000, and 2000 picograms / mL, and standard buffer solutions with pH values ​​of 6.5, 7.0, 7.2, 7.5, 8.0, and 8.5 were prepared and added dropwise to the integrated detection layer of Example 1 and each comparative example. After 15 minutes, fluorescence images were acquired using a fluorescence imaging system with an excitation wavelength of 365 nm and a acquisition wavelength of 615 nm. The gray value of the detection line was used as the signal intensity.

[0046] In Example 1, PCT detection showed good linearity in the range of 0.5 to 20 ng / mL, with a linear regression equation of I = 1286.5C + 412.3, a correlation coefficient R-squared of 0.9921, a detection limit of 0.2 ng / mL, an intra-assay coefficient of variation of 4.2%, and an inter-assay coefficient of variation of 6.8%. For IL-6 detection, in the range of 50 to 2000 picograms / mL, the linear regression equation was I = 892.7C + 285.6, the correlation coefficient R-squared of 0.9897, the detection limit of 20 picograms / mL, the intra-assay coefficient of variation of 5.1%, and the inter-assay coefficient of variation of 7.5%. For pH detection, in the range of 6.5 to 8.5, the fluorescence intensity ratio (520 nm / 680 nm) showed a linear relationship with pH value, with a regression equation of R = -0.42pH + 4.15, an R-squared of 0.9943, and a pH resolution of 0.1.

[0047] The patch of this invention was applied to 30 clinically collected samples of exudate from chronic wounds. Using enzyme-linked immunosorbent assay (ELISA) results as a reference, the PCT detection concordance rate in Example 1 was 93.3%, the IL-6 detection concordance rate was 90.0%, and the pH detection concordance rate was 96.7%. The overall accuracy of the three indicators combined in determining the infection status was 96.7%. In Comparative Example 3, due to direct contact between the ROS bactericidal layer 2 and the integrated detection layer 4, methylene blue diffused into the detection area, leading to increased fluorescence background, and the PCT and IL-6 detection concordance rates decreased to 76.7% and 73.3%, respectively. In Comparative Example 4, the detection signal was quenched due to the diffusion of free methylene blue, and the PCT detection signal intensity decreased by 22% compared to Example 1. In Comparative Example 5, due to the interference of viable bacteria and metabolites in the exudate with antibody activity, the PCT and IL-6 detection concordance rates were 83.3% and 80.0%, respectively. In Comparative Example 6, due to uneven distribution of parallel straight channels, the intra-assay coefficients of variation for PCT and IL-6 detection increased to 9.6% and 11.2%, respectively. The above results systematically verify the key impact of the physical isolation effect of the sample-oriented flow-guiding layer 3, the confinement effect of mesoporous nanoparticles, and the flow resistance design such as dendritic branches on the accuracy and consistency of detection.

[0048] III. Liquid Flow Conductivity Test Artificial wound exudate containing 0.5% bovine serum albumin in phosphate buffer solution was used as the test medium. The solution was continuously supplied at a rate of 5 μL / min beneath the medical adhesive base layer 1. The time and uniformity of liquid distribution from the ROS sterilization layer 2 to each detection area were recorded. In Example 1, the time for liquid to reach the PCT detection area was 42 seconds, the time to reach the IL-6 detection area was 45 seconds, and the time to reach the pH detection area was 38 seconds, with a liquid inflow deviation of less than 8% in the three detection areas. In Comparative Example 3, the liquid diffused randomly on the surface of the integrated detection layer 4, with the time to reach the detection area fluctuating between 60 and 120 seconds, and a liquid inflow deviation greater than 35%. In Comparative Example 6, the liquid inflow deviation of the parallel straight channel was 28%, and the arrival time deviation was 25 seconds.

[0049] IV. Cytotoxicity Comparison Following ISO 10993-5 standards, the MTT assay was used to test the cytotoxicity of Example 1 and Comparative Example 2 in L929 mouse fibroblasts. The cell viability of the extract from Example 1 was 92.6%, with a toxicity grade of 1; the cell viability of the extract from Comparative Example 2 (silver ion dressing) was 71.3%, with a toxicity grade of 2. These results indicate that the photodynamic sterilization method of this invention has better cell compatibility than silver ion sterilization.

[0050] V. Biocompatibility Evaluation A comprehensive biocompatibility evaluation of Example 1 was conducted in accordance with the ISO 10993 series standards: the skin irritation test (intact skin on the back of a rabbit) showed a primary irritation index of 0.25, indicating no irritation; the delayed-type hypersensitivity test (maximum sensitization test in guinea pigs) showed a sensitization rate of 0%, indicating no sensitization; the genotoxicity tests (Ames test and mouse bone marrow micronucleus test) were both negative; and the hemolysis test showed a hemolysis rate of 1.8%, which meets the requirements for medical materials.

[0051] VI. Environmental Reliability Testing The samples from Example 1 were treated under the following conditions: high temperature (54°C ± 2°C, relative humidity 60% ± 10%), low temperature (-20°C ± 2°C), high-low temperature cycling (-20°C to 54°C, 10 cycles), and salt spray (5% sodium chloride solution, 35°C, continuous spraying for 48 hours). Changes in antibacterial rate and detection performance were then analyzed. The results showed that the antibacterial rate decreased by no more than 0.5 percentage points after high-temperature treatment, and the PCT detection signal intensity decreased by no more than 8%. No significant changes were observed in any performance characteristics after low-temperature treatment. The antibacterial rate decreased by no more than 1 percentage point after high-low temperature cycling, and the detection signal intensity decreased by no more than 12%. After salt spray treatment, there was no rust or interlayer peeling, and both the antibacterial rate and detection performance were within acceptable ranges.

[0052] VII. ROS sterilization mechanism and verification of reactive oxygen species diffusion Electron spin resonance spectroscopy (ESR) with 5,5-dimethyl-1-pyrroline-N-oxide as a spin trapping agent was used to detect the reactive oxygen species (ROS) generation signal under 660 nm red light irradiation (power density 50 mW / cm²) in Example 1. The results showed typical singlet oxygen characteristic peaks, with hyperfine coupling constants aN of 13.2 Gauss and aH of 10.8 Gauss. The signal intensity reached a plateau after 5 minutes of illumination. No obvious ROS signal was detected under light-shielded conditions. Intracellular ROS levels in bacteria were detected using 2,7-dichlorofluorescein diacetate as a probe. The intracellular fluorescence intensity of Staphylococcus aureus after light treatment was 6.8 times that of the light-shielded group, confirming that the present invention kills pathogens by generating singlet oxygen through photodynamic effects.

[0053] Further analysis using a reactive oxygen species (ROS) fluorescent probe (single-letoxygen sensorgreen) to detect the singlet oxygen concentration diffusing from the ROS sterilization layer 2 to the inlet of the sample-directed flow layer 3 revealed that under standard illumination, the singlet oxygen concentration at the inlet of the sample-directed flow layer 3 was 0.6 ± 0.2 μmol / L, far below the threshold concentration (approximately 50 μmol / L) that leads to antibody inactivation. As a control, Comparative Example 4 (free methylene blue) showed a singlet oxygen concentration of 12.5 ± 3.1 μmol / L under the same conditions, confirming that the pore confinement effect of the mesoporous nanoparticles and the physical barrier effect of the sample-directed flow layer 3 jointly ensured the accuracy of downstream immunoassays.

[0054] Table 1: Comparison of the overall performance of various wound dressings

[0055] Table 2: Comparison of Key Performance Indicators of Embodiments of the Invention and Comparative Examples

[0056] Table 3: Comparison of Core Technical Features and Existing Technical Solutions

[0057] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. An integrated wound dressing that combines antibacterial and nursing functions, comprising, sequentially arranged from the wound contact side outwards, a medical adhesive base layer (1), a ROS sterilization layer (2), a sample guiding layer (3), an integrated detection layer (4), and a transparent protective observation layer (5), characterized in that: The medical adhesive substrate (1) is a composite structure of low-sensitivity medical pressure-sensitive adhesive coating and microporous array. The thickness of the pressure-sensitive adhesive coating is 25-50 micrometers, the diameter of the microporous pores is 50-150 micrometers, the pore spacing is 200-400 micrometers, and the porosity is 15%-25%. The ROS sterilization layer (2) is a mesoporous nanoparticle composite hydrogel membrane loaded with photosensitive active ingredients, with a thickness of 100-300 micrometers, a particle size of 50-100 nanometers and a pore size of 2-5 nanometers, and a photosensitive active ingredient loading of 5%-15% of the dry weight of the ROS sterilization layer (2). The sample directional flow layer (3) is a hydrophilic polymer film with built-in tree-shaped branched microchannels. The main channel width of the microchannel is 200-300 micrometers, the branch channel width is 100-150 micrometers, the channel depth is 50-100 micrometers, and the channel spacing is 200-500 micrometers. The integrated detection layer (4) includes a PCT fluorescence immunoassay zone, an IL-6 fluorescence immunoassay zone and a pH fluorescence detection zone, which are independent of each other and are arranged side by side on the same nitrocellulose substrate. The transparent protective observation layer (5) is a waterproof and breathable PET film with a thickness of 20 to 50 micrometers and a water vapor permeability of not less than 2000 grams per square meter per 24 hours.

2. The integrated wound dressing with both antibacterial and nursing functions according to claim 1, characterized in that: The mesoporous nanoparticles in the ROS sterilization layer (2) are mesoporous silica nanoparticles, and the photosensitive active ingredient is methylene blue. Methylene blue is loaded on the inner and outer surfaces of the pores of the mesoporous silica through a dual method of physical adsorption and Schiff base covalent bonding. The composite hydrogel matrix is ​​a cross-linking system of sodium alginate and gelatin with a mass ratio of 1:1 to 3:1, and the cross-linking agent is a calcium chloride solution with a mass fraction of 0.5% to 2%. The methylene blue is excited to generate singlet oxygen under 660 nm red light irradiation. The pore confinement effect of the mesoporous nanoparticles makes the singlet oxygen concentration in the diffusion-sample directional flow layer (3) less than 1 μmol per liter.

3. The integrated wound dressing with both antibacterial and nursing functions according to claim 1, characterized in that: The tree-like branched microchannels of the sample directional flow layer (3) adopt a hierarchical structure in which the primary main channel branches into four secondary branch channels, and each secondary branch channel further branches into two tertiary terminal channels. The inlet of the main channel is directly opposite the liquid outlet area of ​​the ROS sterilization layer (2), and the tertiary terminal channels are respectively connected to the three detection area inlets of the integrated detection layer (4). The inner wall of the microchannel is treated with oxygen plasma hydrophilization, and the contact angle is less than 30 degrees. The tree-shaped branched microchannels adopt an equal flow resistance design, and the ratio of the cross-sectional area of ​​the main channel to the total cross-sectional area of ​​the branch channels is 1:1.02 to 1:1.

08.

4. The integrated wound dressing with both antibacterial and nursing functions according to claim 1, characterized in that: The nitrocellulose membrane of the PCT fluorescent immunoassay area is coated with anti-human PCT polyclonal antibody as a detection line, and europium ion chelate-labeled anti-human PCT monoclonal antibody is sprayed on the upstream binding pad of the detection line. The nitrocellulose membrane of the IL-6 fluorescent immunoassay area is coated with anti-human IL-6 polyclonal antibody as a detection line, and the upstream binding pad of the detection line is sprayed with europium ion chelate-labeled anti-human IL-6 monoclonal antibody. The pH fluorescence detection area is a cellulose membrane with a composite indicator of fluorescein isothiocyanate and bromothymol blue immobilized. The europium ion chelate has an excitation wavelength of 340–365 nm and an emission wavelength of 610–620 nm.

5. The integrated wound dressing with both antibacterial and nursing functions according to claim 4, characterized in that: The semi-quantitative determination thresholds for the PCT fluorescence immunoassay detection area are as follows: 0.5 ng / mL for mild infection, 2 ng / mL for moderate infection, and 10 ng / mL for severe infection. The semi-quantitative determination thresholds for the IL-6 fluorescent immunoassay detection area are as follows: 50 picograms per milliliter of fluorescence signal intensity corresponds to a critical IL-6 concentration of mild infection, 200 picograms per milliliter corresponds to a critical IL-6 concentration of moderate infection, and 1000 picograms per milliliter corresponds to a critical IL-6 concentration of severe infection. The semi-quantitative determination thresholds for the pH fluorescence detection area are: pH less than 7.2 indicates a normal healing environment, pH between 7.2 and 7.5 indicates a mild infection, and pH greater than 7.5 indicates a moderate to severe infection.

6. The integrated wound dressing with both antibacterial and nursing functions according to claim 1, characterized in that: The micropores of the medical adhesive base layer (1) are prepared by laser drilling process, with a drilling power of 6 to 10 watts, a pulse width of 0.1 to 0.3 milliseconds, and a repetition frequency of 15 to 25 kilohertz; The transparent protective observation layer (5) is prepared with a conical microstructure array by nanoimprinting process. The bottom diameter of the conical microstructure is 1-3 micrometers, the height is 4-6 micrometers, the spacing is 3-5 micrometers, and the hydrostatic pressure of the film is greater than 30 kPa.

7. The integrated wound dressing with both antibacterial and nursing functions according to claim 1, characterized in that: The photosensitive active ingredient in the ROS sterilization layer (2) is curcumin, the mesoporous nanoparticles are mesoporous carbon nanoparticles, the composite hydrogel matrix is ​​a chitosan and polyvinyl alcohol crosslinking system, and the fluorescent label of the integrated detection layer (4) is CdSe / ZnS quantum dots with an emission wavelength of 600-630 nanometers.

8. A method for preparing an integrated wound dressing that combines antibacterial and nursing functions, characterized in that, The method of using an integrated wound dressing that combines antibacterial and nursing functions as described in any one of claims 1 to 7 includes the following steps: Step S1, preparing the medical adhesive base layer: a low-sensitivity acrylic pressure-sensitive adhesive with a thickness of 25-50 micrometers is coated on a polyurethane elastic pad. A microporous array with a diameter of 50-150 micrometers, a spacing of 200-400 micrometers, and an open area of ​​15%-25% is prepared on the pressure-sensitive adhesive surface using a laser drilling process. After drilling, the surface is ultrasonically cleaned with 75% ethanol for 3-8 minutes. Step S2, Preparation of ROS sterilization layer: Synthesize mesoporous nanoparticles and load them with photosensitive active ingredients. Disperse the loaded mesoporous nanoparticles in a hydrogel matrix solution, and obtain a ROS sterilization layer with a thickness of 100-300 micrometers by cross-linking curing and freeze drying (2). Step S3, Preparation of sample directional flow layer: Dendritic branched microchannels are prepared on the surface of polymer film by photolithography and etching process, and oxygen plasma hydrophilization treatment is performed after resist removal; Step S4, Preparation of integrated detection layer: Three independent detection areas are separated on the nitrocellulose membrane to prepare PCT fluorescence immunoassay area, IL-6 fluorescence immunoassay area and pH value fluorescence detection area respectively; Step S5, Composite Assembly and Sterilization: Align and stack the five layers in sequence, apply medical-grade pressure-sensitive adhesive to the non-circulation areas between the layers, heat-press the composite, then cut, sterilize, and seal the packaging.

9. A method for preparing an integrated wound dressing that combines antibacterial and nursing functions according to claim 8, characterized in that: Step S2 includes the following sub-steps: Step S2-1, Synthesis of mesoporous nanoparticles: Using hexadecyltrimethylammonium bromide as a template agent and tetraethyl orthosilicate as a silicon source, the reaction was carried out in a water-ethanol mixture catalyzed by ammonia at 55-65 degrees Celsius for 1.5-2.5 hours with stirring. After centrifugation and washing, the template agent was removed by calcination at 500-600 degrees Celsius for 4-6 hours to obtain mesoporous silica nanoparticles with a particle size of 50-100 nanometers and a pore size of 2-5 nanometers. Step S2-2, loading of photosensitive active ingredient: Mesoporous silica nanoparticles are dispersed in a methylene blue phosphate buffer solution with a mass fraction of 0.5–2 mg / mL. The mixture is shaken and adsorbed at 20–30°C for 10–14 hours under light-protected conditions. After centrifugation and washing, the mixture is activated with 3-aminopropyltriethoxysilane, so that methylene blue is loaded through a dual mechanism of physical adsorption and covalent bonding with Schiff base. The loading amount is 5%–15% of the mass of the mesoporous nanoparticles. Step S2-3, hydrogel composite molding: Mesoporous nanoparticles loaded with methylene blue are dispersed in a mixed solution of sodium alginate and gelatin at a mass fraction of 2% to 4%, with a mass ratio of sodium alginate to gelatin of 1:1 to 3:

1. After casting, the mixture is immersed in a calcium chloride solution at a mass fraction of 0.5% to 2% for crosslinking for 10 to 20 minutes, rinsed, and then freeze-dried.

10. A method for preparing an integrated wound dressing that combines antibacterial and nursing functions according to claim 8, characterized in that: Step S4 includes the following sub-steps: Step S4-1, detection area separation: Separate the PCT detection area, IL-6 detection area and pH detection area by printing spacer lines on the nitrocellulose membrane with hydrophobic wax lines. Step S4-2, preparation of the immunodetection zone: Anti-human PCT polyclonal antibody and anti-human IL-6 polyclonal antibody were diluted with phosphate buffer to 0.8-1.2 mg / mL, respectively, and streaked onto the corresponding nitrocellulose membrane as detection lines. After drying at 37°C for 1-3 hours, the membrane was blocked with 1% bovine serum albumin. Europium ion chelate-labeled monoclonal antibodies were then sprayed onto the conjugate pads and vacuum dried at 37°C for 3-5 hours. Step S4-3, pH detection zone preparation: Dissolve fluorescein isothiocyanate and bromothymol blue in a mass ratio of 1:1 to 1:3 in a borate buffer solution with a pH of 7.5 to 8.5, with a total concentration of 0.3 to 0.8 mg per milliliter. Immerse the cellulose membrane in the solution for 3 to 8 minutes, then remove it, dry it at 37 degrees Celsius in the dark, and cut and fix it in the pH detection zone.

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