A microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials and its application in the treatment of abscess wounds

Abstract

The present invention discloses a microneedle patch loaded with vancomycin and a gold / zinc-cerium composite nanomaterial, and its use in treating abscess wounds. The microneedle patch comprises a backing layer and a needle array disposed on the backing layer. Both the backing layer and the needle array are formed from an aqueous solution of a degradable polymer, and the aqueous solution of the degradable polymer used to form the needle array is loaded with vancomycin and the gold / zinc-cerium composite nanomaterial. The resulting microneedle patch, based on the photothermal synergistic effect of the antibiotic and the gold / zinc-cerium composite nanomaterial, achieves rapid healing of abscess wounds by inhibiting bacterial growth, reducing oxidative stress, and promoting angiogenesis.

Description

Technical Field

[0001] The present invention belongs to the technical field of medical device preparation, and particularly relates to a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials and application thereof in the treatment of abscess wounds. Background Art

[0002] Staphylococcus aureus (MRSA) is a pathogen that can cause skin and soft tissue infections to serious systemic diseases. Abscesses are a disease caused by S. aureus. Clinically, the main treatments are puncture and antibiotics. However, puncture is not sufficient to completely and thoroughly kill the bacteria. Antibiotic therapy is an effective method for killing bacteria and preventing infection. However, the development of antibiotics has been slow, and bacterial resistance to antibiotics has increased dramatically. There is still no consistently effective and efficient antimicrobial strategy to address this resistance problem.

[0003] In recent years, due to the rapid development of nanotechnology, various new strategies based on multifunctional nanomaterials have been applied to synergistically combat drug-resistant bacterial infections. Nanomaterials have the advantages of strong broad-spectrum antibacterial ability, low drug resistance, and low biological toxicity, and have become a new type of antibiotics. At present, some antibacterial therapies, such as photothermal therapy (PTT) of nanomaterials, have become promising therapeutic strategies for the treatment of drug-resistant bacterial infections. "A Vehicle-Free Antimicrobial Polymer Hybrid Gold Nanoparticle as Synergistically Therapeutic Platforms for Staphylococcusaureus Infected Wound Healing" (Advanced Science, Vol. 9, 2022, pp. 2105223) reported a carrier-free antimicrobial polymer polyhexamethylene biguanide (PHMB) mixed with gold nanoparticles (Au NPs) with bactericidal and anti-biofilm functions, which has strong antibacterial ability against resistant Staphylococcus aureus. The excited PHMB@Au NPs improve the photothermal conversion efficiency by increasing the NIR absorbance, thereby producing a thermal killing effect on bacteria in vitro. At the same time, PHMB shows damage to bacterial membranes, which can promote the absorption of PHMB@Au NPs and target the adhesion of Staphylococcus aureus to kill MRSA. However, the current single PTT for the treatment of drug-resistant bacterial infections remains challenging because high-power and long-term laser irradiation may cause damage to surrounding normal cells and tissues. “Thermal-Cascade Multifunctional Therapeutic Systems for Remotely Controlled Synergistic Treatment of Drug-Resistant Bacterial Infections” (Advanced Functional Materials, Vol. 34, 2024, pp. 2311315) reports a hybrid nanocluster loaded with functional antibiotics to achieve targeted therapeutic delivery and synergistic mild thermotherapy-antibiotic treatment. The thermal cascade multifunctional therapeutic system improves the therapeutic bioavailability of bacteria, enhances the permeability of bacterial membranes, promotes the production of reactive oxygen species (ROS), and strongly inhibits drug-resistant bacteria.

[0004] Although the bacteria in the abscess wound are cleared, the inflammation caused by the bacteria and the slow tissue repair process still hinder the healing of the abscess. 4+ and Zn 2+) as a basic substance for basic biochemical reactions, can play an anti-inflammatory role, promote angiogenesis and tissue regeneration. “Ceria Nanoenzyme-Based Hydrogel with Antiglycativeand Antioxidative Performance for Infected Diabetic Wound Healing” (SmallMethods, Vol. 6, 2022, p. 2200949) reported a self-healing and erasable hydrogel loaded with cerium dioxide nanoparticles with unique anti-glycation ability and excellent antioxidant ability. This hydrogel can reshape the wound microenvironment by accelerating hemostasis, inhibiting infection, reducing advanced glycation end products (AGEs) and continuously consuming reactive oxygen species. Eliminating oxidative stress and glycation can synergistically regulate the inflammatory response, promote vascular recanalization and extracellular matrix deposition, thereby accelerating wound repair. In summary, the healing of abscess wounds can be effectively promoted by combining multiple treatment strategies such as antibacterial therapy, anti-inflammatory and angiogenesis. Summary of the Invention

[0005] The present invention provides a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials and its application in the treatment of abscess wounds. The technical problem to be solved is: constructing a transdermal drug delivery system to achieve the photothermal synergistic antibacterial effect of vancomycin and gold / zinc-cerium composite nanomaterials, scavenging reactive oxygen species to regulate the inflammatory microenvironment, promote angiogenesis and tissue repair, and thus promote the healing of abscess wounds.

[0006] In order to solve the technical problem, the present invention adopts the following technical solution:

[0007] The present invention first discloses a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials, which is characterized in that: the microneedle patch comprises a backing layer and a needle array arranged on the backing layer; the backing layer and the needle array are both formed from an aqueous solution of a degradable polymer, and the vancomycin and the gold / zinc-cerium composite nanomaterials are loaded in the aqueous solution of the degradable polymer used to form the needle array.

[0008] Furthermore, the concentration of the degradable polymer in the aqueous solution of the degradable polymer is 3-5 g / 30 mL, the concentration of the gold / zinc-cerium composite nanomaterial in the aqueous solution of the degradable polymer used to form the needle array is 0.1-1 g / 30 mL, and the concentration of vancomycin is 0.01-0.1 g / 30 mL.

[0009] Furthermore, each needle in the needle array is in a pyramid shape with a height of 400 to 1000 μm and a bottom diameter of 150 to 400 μm.

[0010] Furthermore, the degradable polymers used in the backing layer and the needle array are independently selected from at least one of hyaluronic acid, gelatin, polyvinyl pyrrolidone and polyvinyl alcohol.

[0011] The present invention also discloses a method for preparing the microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial, comprising the following steps:

[0012] dissolving a degradable polymer in deionized water to obtain a backing layer solution;

[0013] Dissolve the degradable polymer in deionized water, add vancomycin and gold / zinc-cerium composite nanomaterials, and stir evenly at room temperature in the dark to obtain a needle mixture;

[0014] The needle body mixture is filled into the needle body micropores of the microneedle mold, and the excess part outside the micropores is removed. Then, a backing layer solution is added above the micropores, dried, and demolded to obtain a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials.

[0015] Furthermore, the preparation method of the gold / zinc-cerium composite nanomaterial is:

[0016] Add 340-345 mg of hexadecyltrimethylammonium bromide to 10-20 mL of deionized water, then add 0.2-0.3 mL of 3.5-4.0 mg / mL tetrachloroauric acid solution and 0.6-0.7 mL of 0.35-0.40 mg / mL sodium borohydride solution, stir at room temperature in the dark for 5-10 minutes to obtain a gold seed dispersion, and let it stand for use;

[0017] 1-2 g of hexadecyltrimethylammonium bromide, 1-2 mL of a 3.5-4.0 mg / mL tetrachloroauric acid solution, 0.7-0.8 mL of a 1.5-2.0 mg / mL silver nitrate aqueous solution, and 1.4-1.6 mL of a 10-12 mg / mL hydroquinone solution were added to 20-40 mL of deionized water, and stirred until the solution became clear. 1.2-1.4 mL of the seed gold dispersion was then added, and the mixture was stirred at room temperature in the dark for 10-12 h to obtain a dispersion of gold nanorods.

[0018] Finally, 120-140 mg of hexadecyltrimethylammonium bromide, 20-40 mg of zinc salt, 10-30 mg of hexamethylenetetramine, 2-8 mg of ascorbic acid and 2-4 mL of a dispersion of gold nanorods were weighed and added to 20-40 mL of deionized water. The mixture was stirred at 80-90° C. for 10-12 hours, and then 10-20 mg of cerium salt was added and the stirring was continued for 10-12 hours. The mixture was naturally cooled to room temperature and centrifuged for washing to obtain a gold / zinc-cerium composite nanomaterial.

[0019] Furthermore, the zinc salt is at least one of zinc nitrate hexahydrate, zinc chloride and zinc acetate dihydrate, and the cerium salt is at least one of cerium nitrate and cerium chloride.

[0020] The microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials obtained by the present invention can be used for transdermal drug delivery to treat abscess wounds. The present invention constructs a soluble polymer microneedle drug delivery system loaded with gold / zinc-cerium and vancomycin. Based on the photothermal synergistic effect of the antibiotic and the gold / zinc-cerium composite nanomaterials, it achieves rapid healing of abscess wounds by inhibiting bacterial growth, reducing oxidative stress, and promoting angiogenesis.

[0021] Compared with the prior art, the beneficial effects of the present invention are embodied in:

[0022] 1. The present invention prepares soluble microneedles loaded with vancomycin and gold / zinc-cerium composite nanomaterials for the first time. The preparation method is simple, the reaction conditions are mild, the resulting product has uniform morphology, and the production cost is low, making it suitable for industrial scale-up production.

[0023] 2. The microneedles loaded with vancomycin and gold / zinc-cerium composite nanomaterials prepared by the present invention can achieve rapid dissolution and achieve more precise drug release through transdermal administration in the abscess epidermis, and have better biocompatibility.

[0024] 3. The microneedles prepared in this invention can synergize with vancomycin through mild heat to achieve a synergistic effect in treating drug-resistant bacterial infections. This system not only reduces the development of bacterial resistance, but also effectively inhibits bacterial growth and alleviates infection symptoms.

[0025] 4. The microneedles prepared by the present invention can utilize the anti-inflammatory properties and angiogenesis ability of cerium and zinc ions, thereby significantly accelerating the healing process of abscess wounds. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is a scanning electron microscope (SEM) photograph of the gold / zinc-cerium composite nanomaterial prepared in Example 1.

[0027] Figure 2 This is the ultraviolet absorption curve of the gold / zinc-cerium composite nanomaterial prepared in Example 1.

[0028] Figure 3 This is an optical image of the microneedle prepared in Example 1.

[0029] Figure 4 This is an inverted fluorescence microscope image of the microneedle prepared in Example 1 in the bright field.

[0030] Figure 5 This is a scanning electron microscope (SEM) photograph of the microneedle prepared in Example 1.

[0031] Figure 6 The microneedles prepared in Examples 1-3 were cultured with Staphylococcus aureus in a 37° C. constant temperature shaker, and the bacterial solution was diluted and coated on a plate for counting.

[0032] Figure 7 Fluorescence photos of the microneedles prepared in Example 1-3 after co-culture with fibroblasts (L929).

[0033] Figure 8 This is a bright field photograph of the microneedles prepared in Example 1-3 after co-culture with fibroblasts (L929).

[0034] Figure 9 The diagram shows the therapeutic effect of the microneedles prepared in Examples 1-3 on the abscess area on the back of mice. DETAILED DESCRIPTION

[0035] The following is a detailed description of an embodiment of the present invention. This embodiment is implemented based on the technical solution of the present invention, and provides a detailed implementation method and specific operation process. However, the protection scope of the present invention is not limited to the following embodiment.

[0036] Example 1

[0037] In this example, a hyaluronic acid microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials was prepared in the following steps:

[0038] 1. Add 345 mg of hexadecyltrimethylammonium bromide to 10 mL of deionized water, then add 0.25 mL of 3.9 mg / mL tetrachloroauric acid solution and 0.6 mL of 0.38 mg / mL sodium borohydride solution. Stir at room temperature in the dark for 5 minutes to obtain a gold seed dispersion, and let it stand for use.

[0039] 1 g of hexadecyltrimethylammonium bromide, 1.5 mL of 3.9 mg / mL tetrachloroauric acid solution, 0.75 mL of 1.7 mg / mL silver nitrate aqueous solution, and 1.5 mL of 11 mg / mL hydroquinone solution were added to 30 mL of deionized water and stirred until the solution became clear. 1.3 mL of the seed gold dispersion was then added and the mixture was stirred at room temperature in the dark for 12 h to obtain a dispersion of gold nanorods.

[0040] Finally, 130 mg of hexadecyltrimethylammonium bromide, 30 mg of zinc salt, 15 mg of hexamethylenetetramine, 5 mg of ascorbic acid, and 3 mL of a dispersion of gold nanorods were weighed and added to 30 mL of deionized water. The mixture was stirred at 85°C for 12 h, and then 15 mg of cerium salt was added and stirred for 10 h. The mixture was naturally cooled to room temperature and centrifuged to obtain a gold / zinc-cerium composite nanomaterial.

[0041] 2. Weigh 3.9g of hyaluronic acid, 0.03g of vancomycin, and 0.15g of gold / zinc-cerium composite nanomaterials and add them to 30mL of deionized water. Stir in the dark at room temperature for 24h to form a uniform colloidal needle mixture. Then use a pipette to add the needle mixture dropwise to the microneedle mold, and then place it in a vacuum box for vacuum treatment until the air is completely extracted, so that the colloidal material fills the needle micropores of the microneedle mold. Remove the excess part outside the micropores, then add 1.5g / mL of PVP solution above the needle micropores to form a backing layer. Then place the mold in a 40°C oven to dry in the dark. Finally, gently peel the formed microneedles from the mold to obtain a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials.

[0042] Figure 1 This is a morphology picture of the gold / zinc-cerium composite nanomaterial prepared in this example. From a scanning electron microscope, it can be seen that the gold / zinc-cerium composite nanomaterial is a uniformly dispersed nanosphere structure with a size of about 100 nm.

[0043] Figure 2 This is the ultraviolet absorption spectrum of the gold / zinc-cerium composite nanomaterial absorbing in the near-infrared II region prepared in this example, which shows that the ultraviolet absorption is in the near-infrared II region (1064 nm).

[0044] Figure 3 This is an optical picture of the microneedles prepared in this example. From the picture, it can be seen that the needle body in the microneedle patch is a 12*12 array. The needle body is a pyramid with a height of 1000μm and a bottom diameter of 600μm. The corresponding backing layer is a circle with a side length of 10mm.

[0045] Figure 4 This is a bright field side view of the microneedle prepared in this example under an inverted fluorescence microscope (Olympus CKX41 biological inverted microscope). It can be seen from the figure that the microneedle has a sharp needle tip.

[0046] Figure 5 This is a scanning electron microscope image (ZEISS GeminiSEM 300 field emission scanning electron microscope) of the microneedle prepared in this example. The overall appearance of the microneedle can be clearly seen from the image.

[0047] Example 2

[0048] In this example, a hyaluronic acid microneedle patch containing only vancomycin was prepared:

[0049] 3.9 g of hyaluronic acid and 0.03 g of vancomycin were weighed and added to 30 mL of deionized water. The mixture was stirred in the dark at room temperature for 24 h to form a uniform colloidal needle mixture. The microneedle patch was then prepared using the same method as in Example 1.

[0050] Example 3

[0051] This example prepares a hyaluronic acid microneedle patch containing only gold / zinc-cerium composite nanomaterial microspheres:

[0052] 3.9 g of hyaluronic acid and 0.15 g of the gold / zinc-cerium composite nanomaterial prepared in Example 1 were weighed and added to 30 mL of deionized water. The mixture was stirred in the dark at room temperature for 24 h to form a uniform colloidal needle mixture. The mixture was then prepared into a microneedle patch using the same method as in Example 1.

[0053] To characterize the performance of the microneedle patches prepared in the above examples, the following tests were performed:

[0054] 1. Antibacterial properties

[0055] Take 20 μL of Staphylococcus aureus (1×10 8 CFUmL -1 ) were mixed with different material groups and incubated in a 37°C incubator for 12 h. The total number of viable bacteria was recorded by serial dilution and plate counting. Figure 6 The microneedles prepared in Examples 1-3 were incubated with Staphylococcus aureus in a 37°C incubator for 12 hours. The results of the dilution plate counts show that the vancomycin and gold / zinc-cerium microneedle-photothermal groups exhibited stronger bactericidal activity than either the vancomycin microneedle group or the gold / zinc-cerium microneedle-photothermal group. This also demonstrates that the synergistic antibacterial effect of the photothermal effect and antibiotics results in stronger bactericidal properties.

[0056] 2. Anti-inflammatory properties

[0057] Fibroblasts (L929) (2×105 cells mL -1 ) After incubation with different material groups, the intracellular fluorescence signal was monitored using a laser confocal microscope to evaluate the level of ROS. Figure 7 Images obtained using a fluorescence microscope (Olympus CKX41 inverted biological microscope) after co-culturing the microneedles prepared in Examples 1-3 with fibroblasts (L929) for 24 hours show that the fluorescent probe DCFH-DA can be converted to DCF in the presence of ROS, which can be excited by a 488nm laser to produce green fluorescence. As can be seen from the figure, in the absence of added ROS (H2O2), the control group cells did not produce green fluorescence. Compared with the ROS (H2O2) group, the green fluorescence in the vancomycin and gold / zinc-cerium microneedle combined with photothermal therapy faded, indicating optimal ROS scavenging effectiveness.

[0058] 3. Promote angiogenesis

[0059] Matrigel was evenly added to each well and fixed in a 37°C incubator for 30 min. HUVECs were then plated in DMEM medium at a rate of 3 × 104 Cells were seeded at a density of 100 μg / mL onto Matrigel, and different materials were added to each well for co-culture. After 4-6 h of culture, the vascular structure in each well was observed. Figure 8 After the microneedles prepared in Examples 1-3 were co-cultured with fibroblasts (L929) for 4 hours, images were obtained using a fluorescence microscope (Olympus CKX41 biological inverted microscope) in the bright field. It can be seen from the figure that the cells in the control group were dispersed, while the cells in the vancomycin and gold / zinc-cerium microneedle combined with photothermal group were ring-shaped, indicating that the microneedles have the ability to promote angiogenesis.

[0060] 4. Treatment effect on abscess wounds

[0061] Mice were randomly divided into three groups: a control group, a vancomycin microneedle group, a gold / zinc-cerium microneedle combined with photothermal therapy group, and a vancomycin and gold / zinc-cerium microneedle combined with photothermal therapy group, with five mice per group. The abscess model was established by subcutaneously injecting methicillin-resistant Staphylococcus aureus into the mice. Treatment consisted of applying a microneedle patch to the abscess and removing it after 5 minutes. The day of successful modeling was designated Day 0, and treatments were administered on Days 1, 3, and 5, for a total of three times. Photos were taken on Days 0, 3, 7, 11, and 14 to document the therapeutic effects.

[0062] Control group: no treatment.

[0063] Vancomycin microneedle group: The hyaluronic acid microneedles containing only vancomycin described in Example 2 were administered to the modeled mice.

[0064] Gold / zinc-cerium microneedle combined with photothermal group: The mice were treated with hyaluronic acid microneedles containing only gold / zinc-cerium composite nanomaterials as described in Example 3, and the 1064 nm laser (1 W cm -2 , 5min) irradiation treatment.

[0065] Vancomycin and gold / zinc cerium microneedle combined with photothermal therapy: The model mice were administered with hyaluronic acid microneedles loaded with vancomycin and gold / zinc cerium composite nanomaterials as described in Example 1, and the 1064 nm laser (1 W cm -2 , 5min) irradiation treatment.

[0066] Figure 9 The images show the therapeutic effects of the microneedles prepared in various examples on abscesses on the back of mice. As can be seen from the images, the vancomycin and gold / zinc-cerium microneedle combined with photothermal therapy group showed the fastest healing rate compared to the other groups, demonstrating its ability to promote the healing of abscess wounds.

[0067] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials, characterized by: The microneedle patch includes a backing layer and a needle array disposed on the backing layer; the backing layer and the needle array are both formed from an aqueous solution of a degradable polymer, and the aqueous solution of the degradable polymer used to form the needle array is loaded with vancomycin and a gold / zinc-cerium composite nanomaterial; the degradable polymer is hyaluronic acid; and the preparation method of the gold / zinc-cerium composite nanomaterial is as follows: Add 340-345 mg of hexadecyltrimethylammonium bromide to 10-20 mL of deionized water, then add 0.2-0.3 mL of 3.5-4.0 mg / mL tetrachloroauric acid solution and 0.6-0.7 mL of 0.35-0.40 mg / mL sodium borohydride solution. Stir at room temperature in the dark for 5-10 minutes to obtain a gold seed dispersion, and let it stand for use. Add 1-2 g of hexadecyltrimethylammonium bromide, 1-2 mL of 3.5-4.0 mg / mL tetrachloroauric acid solution, 0.7-0.8 mL of 1.5-2.0 mg / mL silver nitrate aqueous solution, and 1.4-1.6 mL of 10-12 mg / mL hydroquinone solution to 20-40 mL of deionized water, stir until the solution is clear, then add 1.2-1.4 mL of the seed gold dispersion, stir and react at room temperature in the dark for 10-12 h to obtain a dispersion of gold nanorods; Finally, 120-140 mg of hexadecyltrimethylammonium bromide, 20-40 mg of zinc salt, 10-30 mg of hexamethylenetetramine, 2-8 mg of ascorbic acid, and 2-4 mL of a dispersion of gold nanorods were weighed and added to 20-40 mL of deionized water. The mixture was stirred at 80-90°C for 10-12 h, and then 10-20 mg of cerium salt was added and stirred for another 10-12 h. The mixture was naturally cooled to room temperature and centrifuged for washing to obtain a gold / zinc-cerium composite nanomaterial.

2. The microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to claim 1, characterized in that: The concentration of the degradable polymer in the aqueous solution of the degradable polymer is 3-5 g / 30 mL.

3. The microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to claim 1 or 2, characterized in that: The concentration of the gold / zinc-cerium composite nanomaterial in the aqueous solution of the degradable polymer used to form the needle array is 0.1-1 g / 30 mL, and the concentration of vancomycin is 0.01-0.1 g / 30 mL.

4. The microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to claim 1, characterized in that: Each needle in the needle array is in a pyramid shape with a height of 400 to 1000 μm and a bottom diameter of 150 to 400 μm.

5. The microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to claim 1, characterized in that: The zinc salt is at least one of zinc nitrate hexahydrate, zinc chloride and zinc acetate dihydrate, and the cerium salt is at least one of cerium nitrate and cerium chloride.

6. A method for preparing a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to any one of claims 1 to 5, characterized in that: The steps include: dissolving a degradable polymer in deionized water to obtain a backing layer solution; Dissolve the degradable polymer in deionized water, add vancomycin and gold / zinc-cerium composite nanomaterials, and stir evenly at room temperature in the dark to obtain a needle mixture; The needle body mixture is filled into the needle body micropores of the microneedle mold, the excess part outside the micropores is removed, and then a backing layer solution is added above the micropores. After drying and demolding, a microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterials is obtained.

7. Use of the microneedle patch loaded with vancomycin and gold / zinc-cerium composite nanomaterial according to any one of claims 1 to 5 in the preparation of a medicine for healing abscess wounds.

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