Lysozyme functionalized tissue-adhesive hemostatic patch

Abstract

This invention relates to the field of medical hemostatic materials, specifically a lysozyme-functionalized tissue-adhesive hemostatic patch, comprising a superhydrophobic conductive layer, a nanofiber barrier layer, an interpenetrating network adhesion layer, a hydrophobic and breathable structural layer, and a release layer. The upper surface of the superhydrophobic conductive layer has a fluorosilane-modified titanium dioxide nanocoating; the nanofiber barrier layer is a polycaprolactone-based lysozyme composite nanofiber layer; the interpenetrating network adhesion layer is an interpenetrating network hydrogel layer formed by lysozyme, oxidized dextran, and gelatin; the hydrophobic and breathable structural layer includes a hydrophobic and breathable microporous membrane and a nonwoven fabric support layer. Through the synergistic effect of the multi-layered structure, it achieves rapid hemostasis, strong wet tissue adhesion, prevention of functional component leakage, and continuous active antibacterial effects, making it particularly suitable for treating superficial wounds with oozing bleeding.

Description

Technical Field

[0001] This invention relates to the field of medical hemostatic materials technology, specifically to a lysozyme-functionalized tissue adhesive hemostatic patch. Background Technology

[0002] In daily life, superficial wounds such as cuts, abrasions, and punctures are extremely common, often accompanied by persistent bleeding. If not treated promptly, they can not only affect wound healing but also easily lead to bacterial infection. Currently available adhesive bandages typically consist of non-woven adhesive tape and absorbent pads, relying primarily on physical pressure and absorption of exudate to achieve passive hemostasis. However, these products have low hemostatic efficiency and struggle to quickly form stable blood clots in wounds with even moderate bleeding. Furthermore, once the pressure-sensitive adhesive layer becomes soaked with blood or sweat, its adhesiveness decreases drastically, causing the dressing to shift or even fall off, exposing the wound and increasing the risk of infection. In addition, ordinary adhesive bandages lack active antibacterial properties and are ineffective at inhibiting bacterial growth on the wound surface. In recent years, some functional hemostatic dressings containing chitosan, oxidized cellulose, and other ingredients have been developed. These dressings accelerate hemostasis by activating clotting factors or concentrating blood. However, most of these products have insufficient tissue adhesion in a wet state and easily detach from the wound surface after contact with water or tissue exudate, making it difficult to maintain reliable fixation during daily activities. Furthermore, the structural design of existing hemostatic dressings often fails to consider the leakage of functional ingredients at the edges after wetting, leading to the easy loss of hemostatic and antibacterial active ingredients with liquid, shortening the effective action time and potentially staining clothing. Therefore, developing a patch-type hemostatic product that has fast hemostasis, strong tissue adhesion, prevents leakage of functional components, and has continuous antibacterial effects is of great practical significance. Summary of the Invention

[0003] The purpose of this invention is to provide a lysozyme-functionalized tissue adhesive hemostatic patch to solve the problems of insufficient wet adhesion, easy leakage of functional components, and lack of sustained antibacterial ability in existing hemostatic dressings. To achieve the above objectives, the present invention provides the following technical solution: a lysozyme-functionalized tissue adhesive hemostatic patch, comprising: A superhydrophobic flow-guiding layer, the lower surface of which is divided into a central region and a peripheral region surrounding the central region; A nanofiber barrier layer is composited in the central region; An interpenetrating network adhesion layer is composited on the lower surface of the nanofiber barrier layer; A hydrophobic and breathable structural layer is composited in the peripheral region and surrounds the nanofiber barrier layer and the interpenetrating network adhesion layer. The lower surface of the hydrophobic and breathable structural layer is an adhesive region. Release layer, which peels off the hydrophobic and breathable structural layer and the interpenetrating network adhesion layer. Lower surface; The superhydrophobic guiding layer is a thermoplastic material with a fluorosilane-modified titanium dioxide nanocoating on its upper surface. A polyurethane-based membrane; the nanofiber barrier layer is a polycaprolactone-based lysozyme composite nanofiber layer. The interpenetrating network adhesion layer is an interpenetrating network hydrogel formed by lysozyme, oxidized dextran, and gelatin. The hydrophobic and breathable structural layer includes at least one hydrophobic and breathable microporous membrane and one nonwoven fabric support layer. Furthermore, the lysozyme in the nanofiber barrier layer is distributed in an island structure inside the polycaprolactone nanofibers, and the pore size of the nanofiber barrier layer is 0.5–5 μm. Furthermore, in the interpenetrating network adhesion layer, lysozyme forms a Schiff base through dynamic covalent cross-linking with the aldehyde groups of oxidized dextran via its free amino groups, thus constituting the first network of the interpenetrating network; gelatin forms a second network through physical cross-linking. Furthermore, the lower surface of the interpenetrating network adhesion layer is flush with the lower surface of the hydrophobic and breathable structural layer. Furthermore, the adhesive area on the lower surface of the hydrophobic and breathable structural layer is made of medical pressure-sensitive adhesive. Furthermore, the mass of lysozyme in the nanofiber barrier layer accounts for 5% to 15% of the mass of polycaprolactone. Furthermore, the mass ratio of lysozyme to oxidized dextran in the interpenetrating network adhesion layer is 1:0.5-2, and the oxidation degree of oxidized dextran is 30%-60%. A method for preparing a lysozyme-functionalized tissue adhesive hemostatic patch includes the following steps: S1. Using a thermoplastic polyurethane film as the base film, a fluorosilane-modified titanium dioxide nanoparticle dispersion is sprayed onto its upper surface. After curing, the superhydrophobic guiding layer is formed, and the central and peripheral areas of the lower surface of the superhydrophobic guiding layer are defined. S2. The mixed solution of polycaprolactone and lysozyme is directly spun into the nanofiber barrier layer in the central region by electrospinning. S3. A mixed solution containing gelatin, lysozyme and oxidized dextran is coated on the lower surface of the nanofiber barrier layer, and an interpenetrating network hydrogel is formed by physical cross-linking and Schiff base cross-linking to obtain the interpenetrating network adhesion layer. S4. Prepare a hydrophobic and breathable structural layer comprising a hydrophobic and breathable microporous membrane and a nonwoven fabric, and attach the hydrophobic and breathable structural layer to the peripheral area, and coat the lower surface of the hydrophobic and breathable structural layer with medical pressure-sensitive adhesive to form an adhesive area. S5. The release layer is peelably adhered to the adhesive area on the lower surface of the hydrophobic and breathable structural layer to obtain a hemostatic patch. Further, in step S2, the parameters of the electrospinning process are: voltage 15-20kV, receiving distance 15cm, temperature 25℃, and humidity 40%; in step S3, the gelatin solution has a mass-volume fraction of 10%-15%, lysozyme concentration of 5-20mg / mL, oxidized dextran oxidation degree of 30%-60%, and the mass ratio of lysozyme to oxidized dextran is 1:0.5-2. Further, in step S4, when preparing the hydrophobic and breathable structural layer, a polytetrafluoroethylene (PTFE) hydrophobic and breathable microporous membrane with a thickness of 20–30 μm and a pore size range of 0.1–0.5 μm and a polyester nonwoven fabric with a thickness of 80–150 μm are selected first; then, the PTFE hydrophobic and breathable microporous membrane and the polyester nonwoven fabric are stacked and hot-pressed together for 60–120 seconds at a temperature of 120–150 °C and a pressure of 0.3–0.6 MPa to form a hydrophobic and breathable substrate with a total thickness of 100–180 μm; then, a die-cutting process is used to cut the hydrophobic and breathable substrate into rings according to the outer contour size and shape of the peripheral area. Compared with the prior art, the beneficial effects of the present invention are: The present invention relates to a lysozyme-functionalized tissue adhesive hemostatic patch that uses an interpenetrating network adhesive layer to directly contact the wound surface. The gelatin and oxidized dextran in the patch can quickly absorb wound exudate and form a physical gel barrier. At the same time, the Schiff base dynamic covalent cross-linking network gives it excellent wet tissue adhesion, which can firmly adhere to moist tissue and is not easy to fall off. This invention relates to a lysozyme-functionalized tissue adhesive hemostatic patch that forms a unidirectional synergistic mechanism of flow and barrier by incorporating a superhydrophobic conductive layer and a nanofiber barrier layer. The superhydrophobic conductive layer prevents external moisture from penetrating, while the nanofiber barrier layer (containing lysozyme) effectively prevents the diffusion of active ingredients (such as lysozyme and oxidized dextran) in the adhesive layer to the outside of the patch, ensuring that the functional ingredients are concentrated in the wound area, prolonging the action time and avoiding staining clothing. This invention relates to a lysozyme-functionalized tissue adhesive hemostatic patch in which both the nanofiber barrier layer and the interpenetrating network adhesion layer are loaded with lysozyme. Lysozyme, as a natural antibacterial protein, can disrupt bacterial cell walls, achieving a long-lasting and safe antibacterial effect, and is less likely to induce drug resistance. Attached Figure Description Figure 1 This is an exploded structural diagram of the entire invention. In the diagram: 1-Superhydrophobic flow layer; 1.1-Central region; 1.2-Outer region; 2-Nanofiber barrier layer; 3-Interpenetrating network adhesion layer; 4-Hydrophobic and breathable structural layer; 5-Release layer. Detailed Implementation 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. Example 1: Please refer to Figure 1 This invention provides a technical solution: a lysozyme-functionalized tissue adhesive hemostatic patch, comprising: The superhydrophobic guiding layer 1 has its lower surface divided into a central region 1.1 and a peripheral region 1.2 surrounding the central region 1.1; Nanofiber barrier layer 2 is composited in the central region 1.1; An interpenetrating network adhesion layer 3 is composited on the lower surface of the nanofiber barrier layer 2; A hydrophobic and breathable structural layer 4 is composited in the peripheral region 1.2 and surrounds the nanofiber barrier layer 2 and the interpenetrating network adhesion layer 3. The lower surface of the hydrophobic and breathable structural layer 4 is an adhesive region. Release layer 5 is peelable and covers the lower surface of the hydrophobic and breathable structural layer 4 and the interpenetrating network adhesive layer 3. Specifically, the superhydrophobic guiding layer 1 uses a thermoplastic polyurethane (TPU) base film with a thickness of 50 μm, and a dispersion of fluorosilane-modified titanium dioxide nanoparticles is sprayed onto its upper surface. After curing at 80°C for 30 minutes, a superhydrophobic coating is formed. The nanofiber barrier layer 2 was prepared by electrospinning: polycaprolactone (PCL, molecular weight 80 kDa) and lysozyme (mass ratio PCL:lysozyme = 10:1) were co-dissolved in hexafluoroisopropanol to prepare a 10% (w / v) spinning solution. Under the conditions of 18 kV voltage, 15 cm receiving distance, 25 °C temperature, and 40% humidity, direct spinning was performed in the central region 1.1 for 30 min to obtain a nanofiber layer with a pore size of 1-3 μm and a thickness of about 50 μm, in which the lysozyme was distributed in an island-like pattern inside the PCL fibers. Preparation of interpenetrating network adhesion layer 3: A 10% (w / v) gelatin aqueous solution was prepared, and lysozyme was added to a final concentration of 10 mg / mL, and stirred until homogeneous. Separately, an aqueous solution of oxidized dextran (oxidation degree 45%) was prepared with a concentration of 10 mg / mL. The gelatin / lysozyme solution and the oxidized dextran solution were mixed at a volume ratio of 1:1 and immediately coated onto the lower surface of the nanofiber barrier layer 2, with a thickness controlled at 200 μm. The mixture was allowed to stand at 25 °C for 30 min for cross-linking to form an interpenetrating network hydrogel. In this structure, the free amino groups of lysozyme and the aldehyde groups of oxidized dextran form Schiff base dynamic covalent cross-links (first network), while the gelatin segments form physical cross-links through hydrogen bonds and chain entanglement (second network). Preparation of the hydrophobic and breathable structural layer 4: A polytetrafluoroethylene (PTFE) microporous membrane with a thickness of 25 μm and a pore size of 0.2 μm was selected, along with a polyester nonwoven fabric with a thickness of 100 μm. The two were then laminated and hot-pressed at 130℃ and 0.5 MPa for 90 seconds to obtain a composite substrate with a total thickness of 125 μm. Subsequently, a die-cutting process was used to cut the substrate to a ring-shaped dimension of 1.2 mm in the outer region. Medical-grade acrylic pressure-sensitive adhesive was coated onto the lower surface of this annular hydrophobic and breathable structural layer 4 to form the adhesive area. The upper surface of the hydrophobic and breathable structural layer 4 is bonded to the outer periphery 1.2 of the superhydrophobic flow guiding layer 1, so that the lower surface of the interpenetrating network adhesion layer 3 is flush with the lower surface of the hydrophobic and breathable structural layer 4. Finally, a release layer 5 (silicone release paper) is applied to obtain the finished product. Example 2: Essentially the same as Example 1, except that the mass of lysozyme in the nanofiber barrier layer 2 accounts for 15% of the mass of polycaprolactone (i.e., PCL:lysozyme = 6.7:1), the electrospinning voltage is 15kV, and the fiber pore size is 0.5–2μm. In the interpenetrating network adhesion layer 3, the mass ratio of lysozyme to oxidized dextran is 1:2, and the oxidation degree of oxidized dextran is 30%. Example 3: Essentially the same as Example 1, except that the mass of lysozyme in the nanofiber barrier layer 2 accounts for 5% of the mass of polycaprolactone (i.e., PCL:lysozyme = 20:1), the electrospinning voltage is 20 kV, and the fiber pore size is 3–5 μm. In the interpenetrating network adhesion layer 3, the mass ratio of lysozyme to oxidized dextran is 1:0.5, and the oxidation degree of oxidized dextran is 60%. Comparative Example 1: Ordinary adhesive bandages (non-woven tape + absorbent cotton pad) were used. Comparative Example 2: Prepared according to the method of Example 1, but omitting the nanofiber barrier layer 2, and directly composited the interpenetrating network adhesion layer 3 onto the central region 1.1 of the superhydrophobic conductive layer 1. Performance testing: Wet tissue adhesion test: Take fresh pigskin and moisten the surface. Attach each sample (2cm×2cm) to the pigskin, apply 500g pressure for 30 seconds, and then perform a 180° peel test using a universal testing machine (tensile speed 50mm / min). The results showed that the peel strength of Example 1 was 4.8 N / cm, Example 2 was 4.2 N / cm, and Example 3 was 4.5 N / cm, while Comparative Example 1 was only 0.6 N / cm (after peeling off), and Comparative Example 2 was 1.8 N / cm. This indicates that the interpenetrating network adhesive layer 3 of the present invention has excellent wet adhesion performance. Functional component leakage test: Each sample was attached to an agar gel wound model containing simulated blood (PBS + hemoglobin) and incubated at 37°C for 6 hours. Exudate from the outer edge of the patch was collected, and the lysozyme content was detected by BCA method. The results showed that the lysozyme concentration in the edge exudate of Example 1 was less than 0.05 μg / mL, while the lysozyme concentration in the edge exudate of Comparative Example 2 was as high as 12.3 μg / mL. This indicates that the nanofiber barrier layer 2 of the present invention can effectively prevent the leakage of functional components. 3. Antibacterial Performance Test: The inhibition zone method was used, with Staphylococcus aureus as the indicator bacterium. Each sample was affixed to an agar plate coated with bacterial suspension and incubated at 37°C for 24 hours. Examples 1, 2, and 3 all formed distinct inhibition zones (diameters of 18 mm, 16 mm, and 17 mm, respectively), while Comparative Example 1 showed no inhibition zone, and Comparative Example 2 had a smaller inhibition zone (8 mm), which significantly decreased in size after 6 hours. This indicates that the present invention possesses sustained and effective antibacterial activity. It should be noted that in this article, relational terms such as first and second are only used to refer to... Distinguishing one entity or operation from another does not necessarily require or imply any such actual relationship or order between those entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus. 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. A lysozyme-functionalized tissue adhesive hemostatic patch, characterized in that, include: The superhydrophobic flow-guiding layer (1) has a lower surface divided into a central region (1.1) and a peripheral region (1.2) surrounding the central region (1.1). A nanofiber barrier layer (2) is composited in the central region (1.1). An interpenetrating network adhesion layer (3) is composited on the lower surface of the nanofiber barrier layer (2); A hydrophobic and breathable structural layer (4) is composited in the peripheral region (1.2) and surrounds the nanofiber barrier layer (2) and the interpenetrating network adhesion layer (3). The lower surface of the hydrophobic and breathable structural layer (4) is the adhesive region. Release layer (5) is peelably applied to the lower surface of the hydrophobic and breathable structural layer (4) and the interpenetrating network adhesive layer (3); The superhydrophobic guiding layer (1) is a thermoplastic polyurethane base film with a fluorosilane-modified titanium dioxide nanocoating on its upper surface; the nanofiber barrier layer (2) is a polycaprolactone-based lysozyme composite nanofiber layer; the interpenetrating network adhesion layer (3) is an interpenetrating network hydrogel layer formed by lysozyme, oxidized dextran and gelatin; and the hydrophobic and breathable structural layer (4) includes at least one hydrophobic and breathable microporous membrane and one nonwoven fabric support layer.

2. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 1, characterized in that: The lysozyme in the nanofiber barrier layer (2) is distributed in an island structure inside the polycaprolactone nanofibers, and the pore size of the nanofiber barrier layer (2) is 0.5 to 5 μm.

3. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 1, characterized in that: In the interpenetrating network adhesion layer (3), lysozyme forms a Schiff base dynamic covalent cross-link with the aldehyde group of oxidized dextran through its free amino group, thus forming the first network of the interpenetrating network; gelatin forms the second network through physical cross-linking.

4. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 1, characterized in that: The lower surface of the interpenetrating network adhesion layer (3) is flush with the lower surface of the hydrophobic and breathable structural layer (4).

5. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 1, characterized in that: The adhesive area on the lower surface of the hydrophobic and breathable structural layer (4) is made of medical pressure-sensitive adhesive.

6. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 2, characterized in that: The mass of lysozyme in the nanofiber barrier layer (2) accounts for 5% to 15% of the mass of polycaprolactone.

7. The lysozyme-functionalized tissue adhesive hemostatic patch according to claim 3, characterized in that: The mass ratio of lysozyme to oxidized dextran in the interpenetrating network adhesion layer (3) is 1:0.5-2, and the oxidation degree of oxidized dextran is 30%-60%.

8. A method for preparing a lysozyme-functionalized tissue adhesive hemostatic patch as described in claim 1, characterized in that, Includes the following steps: S1. Using a thermoplastic polyurethane film as the base film, a dispersion of fluorosilane-modified titanium dioxide nanoparticles is sprayed onto its upper surface. After curing, the superhydrophobic guiding layer (1) is formed, and the central region (1.1) and the peripheral region (1.2) of the lower surface of the superhydrophobic guiding layer (1) are defined. S2. In the central region (1.1), a mixed solution of polycaprolactone and lysozyme is directly spun into the nanofiber barrier layer (2) by electrospinning. S3. A mixed solution containing gelatin, lysozyme and oxidized dextran is coated on the lower surface of the nanofiber barrier layer (2), and an interpenetrating network hydrogel is formed by physical cross-linking and Schiff base cross-linking to obtain the interpenetrating network adhesion layer (3). S4. Prepare a hydrophobic and breathable structural layer (4) comprising a hydrophobic and breathable microporous membrane and a nonwoven fabric, and attach the hydrophobic and breathable structural layer (4) to the peripheral area (1.2), and coat the lower surface of the hydrophobic and breathable structural layer (4) with medical pressure-sensitive adhesive to form an adhesive area. S5. The release layer (5) is peelably attached to the adhesive area on the lower surface of the hydrophobic and breathable structural layer (4) to obtain a hemostatic patch.

9. The method for preparing a lysozyme-functionalized tissue adhesive hemostatic patch according to claim 8, characterized in that: The parameters for the electrospinning process in step S2 are: voltage 15-20kV, receiving distance 15cm, temperature 25℃, and humidity 40%. In step S3, the gelatin solution has a mass-volume fraction of 10%–15%, the lysozyme concentration is 5–20 mg / mL, the oxidation degree of oxidized dextran is 30%–60%, and the mass ratio of lysozyme to oxidized dextran is 1:0.5–2.

10. The method for preparing a lysozyme-functionalized tissue adhesive hemostatic patch according to claim 8, characterized in that: In step S4, when preparing the hydrophobic and breathable structural layer (4), a polytetrafluoroethylene hydrophobic and breathable microporous membrane with a thickness of 20-30 μm and a pore size range of 0.1-0.5 μm and a polyester nonwoven fabric with a thickness of 80-150 μm are selected. Then, the polytetrafluoroethylene hydrophobic and breathable microporous membrane and the polyester nonwoven fabric are stacked and hot-pressed together for 60-120 seconds at a temperature of 120-150℃ and a pressure of 0.3-0.6 MPa to form a hydrophobic and breathable substrate with a total thickness of 100-180 μm. Then, a die-cutting process is used to cut the hydrophobic and breathable substrate in a ring according to the outer contour size and shape of the outer periphery (1.2).

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