An antifungal microneedle patch, a preparation method and application thereof

Abstract

The present application relates to an antifungal microneedle patch and its preparation method and application, the antifungal microneedle patch includes backing layer and the needle head arranged on the surface of the backing layer;The needle head constitutes n×n array on the surface of the backing layer;The surface of the needle head includes fatty acid coating containing antibacterial drug;The inside of the needle head includes photosensitizer;The n is selected from integer.The antibacterial drug is lipophilic triazole antifungal drug.The fatty acid is selected from saturated fatty acid containing 6-12 carbon atoms.The present application innovatively combines antifungal drug itraconazole and photo-thermal agent / photosensitizer indocyanine green, loads in microneedle, can carry out painless minimally invasive local administration, through the antibacterial action, photo-thermal effect and photodynamic effect of antifungal drug released under the excitation of near-infrared light, realize the effective killing to fungi, and will not cause irreversible damage to normal tissue.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to an antifungal microneedle patch, its preparation method, and its application. Background Technology

[0002] Globally, approximately 20-25% of the population suffers from fungal skin infections. If left untreated, these infections can develop into invasive fungal infections and threaten life. Currently, most treatments for fungal skin infections use topical preparations such as lotions, creams, and gels containing antifungal drugs. However, these topical preparations are only effective against superficial fungal skin infections. Because the drugs have difficulty penetrating the stratum corneum, topical preparations are less effective against deep infections caused by fungi that can migrate from the epidermis and dermis to the subcutaneous tissue. Treatment for deep fungal infections usually involves oral antifungal medications, but this systemic administration method lacks targeting and can cause significant toxic side effects. Therefore, there is an urgent need to develop effective, safe, and easy-to-use methods to combat skin infections.

[0003] Microneedles are patches composed of an array of tiny needles ranging from 100 to 1000 μm in length. They can penetrate the stratum corneum to deliver medication, and because the puncture depth does not reach the nerve endings in the dermis, it is a painless or minimally invasive local drug delivery method. Most microneedles are made of polymers for encapsulating and controlling the release of drugs. Current research is exploring the use of microneedles for the detection and treatment of various diseases, including skin cancer, skin infections, diabetes, and eye diseases.

[0004] Several studies have reported the use of microneedles for treating fungal skin infections, such as microneedle patches made using chitosan-polyethyleneimine copolymer as an antibacterial polymer and encapsulating amphotericin B to treat deep fungal skin infections. There are also hyaluronic acid microneedle patches encapsulated with itraconazole nanocrystals for treating Candida albicans skin infections. However, in recent years, the number of multidrug-resistant fungi discovered clinically has been increasing, and antibiotic-based antifungal drugs alone are insufficient to eradicate them.

[0005] Photothermal therapy refers to the physical destruction of fungal structures by raising the temperature of a photothermal agent under light excitation, thus achieving a bactericidal effect. This physical destruction mechanism does not lead to fungal resistance. However, photothermal sterilization alone usually requires high temperatures and long durations, which inevitably damages surrounding normal cells and tissues. Photodynamic therapy is a treatment method that uses light to activate photosensitizers to generate cytotoxic reactive oxygen species, causing oxidative damage to cellular components (lipids, proteins, and nucleic acids), thereby inactivating fungi. Photodynamic therapy exhibits broad-spectrum antibacterial properties and avoids the development of fungal resistance. However, the low skin penetration efficiency, poor water solubility, and instability of free photosensitizers limit the further clinical application of photodynamic therapy.

[0006] Therefore, it is urgent to provide new technical solutions based on existing technologies to solve the problems existing in the existing technologies. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention innovatively combines the antifungal drug itraconazole with the photothermal / photosensitizer indocyanine green, loading them into microneedles for painless, minimally invasive local drug delivery. By combining the antifungal effects, photothermal effects, and photodynamic effects of the antifungal drug released under near-infrared light excitation, it effectively kills fungi without causing irreversible damage to normal tissues.

[0008] One object of the present invention is to provide an antifungal microneedle patch, the antifungal microneedle patch comprising a backing layer and needles disposed on the surface of the backing layer;

[0009] The needles form an n×n array on the surface of the backing layer;

[0010] The needle surface includes a fatty acid coating containing an antibacterial agent;

[0011] The needle contains a photosensitizer;

[0012] The n is selected from integers.

[0013] Furthermore, the antibacterial drug is a lipophilic triazole antifungal drug.

[0014] Furthermore, the antibiotic is itraconazole.

[0015] Furthermore, the fatty acid is selected from saturated fatty acids containing 6-12 carbon atoms.

[0016] Furthermore, the fatty acid is preferably lauric acid.

[0017] Furthermore, the needle is made of chitosan, and the backing layer is made of hyaluronic acid.

[0018] Furthermore, the photosensitizer is indocyanine green.

[0019] Itraconazole (ITZ, CAS Registry No.: 84625-61-6) is a lipophilic triazole antifungal drug used for a wide range of fungal infections. Its mechanism of action involves highly selectively binding to fungal cytochrome P450 isoenzymes, inhibiting ergosterol synthesis, leading to fungal cell membrane damage and ultimately cell death. Its binding to the mammalian P450 system is weak, significantly reducing its toxicity to humans. Clinically, itraconazole is primarily used for systemic infections caused by deep fungi, such as chromoblastomycosis, histoplasmosis, coccidioidomycosis, sporotrichosis, coccidioidomycosis, candidiasis, and aspergillosis. Its structural formula is as follows:

[0020]

[0021] Indocyanine green (ICG, CAS Registry No.: 3599-32-4) is a commonly used contrast agent in the clinical diagnosis of cardiovascular diseases in my country, used for imaging choroidal and retinal vessels. Indocyanine green exhibits strong absorption of 808nm near-infrared laser light. The absorbed energy dissipates in three ways: producing fluorescence, generating heat, and generating reactive oxygen species. The heat and reactive oxygen species produced can damage cells; therefore, indocyanine green has been used in studies to treat tumors and bacterial infections. Its structural formula is as follows:

[0022]

[0023] Chitosan (CS, CAS Registry No.: 9012-76-4) is a product of the natural polysaccharide chitin, which has undergone partial acetylation. It possesses various physiological functions, including biodegradability, biocompatibility, antibacterial properties, and anti-inflammatory effects, and is widely used in numerous fields such as food additives, cosmetics and health products, antibacterial agents, medical fibers, medical dressings, and drug sustained-release materials. Its structural formula is as follows:

[0024]

[0025] Sodium hyaluronate (HA, CAS Registry No.: 9004-61-9), also known as sodium hyaluronate, is a naturally occurring biological macromolecule in the human body, constituting the dermis layer of the skin, with a molecular weight ranging from hundreds of thousands to millions. Due to its excellent biocompatibility, moisturizing, lubricating, and anti-wrinkle properties, it has been widely used in drug carriers, skincare products, and medical aesthetic products. Its structural formula is as follows:

[0026]

[0027] Lauric acid (LA, CAS Registry No.: 143-07-7) is a medium-chain saturated fatty acid containing 12 carbon atoms, mainly found in coconut oil and palm seed oil. It has numerous applications in food additives, flavorings, pharmaceuticals, and surfactants. Lauric acid is a phase change material with a phase change temperature of 44.4℃; when the temperature exceeds 44.4℃, lauric acid will change from a solid phase to a liquid phase. Its structural formula is as follows:

[0028]

[0029] Another object of the present invention is to provide a method for preparing the above-mentioned antifungal microneedle patch, comprising the following steps:

[0030] S1. Add chitosan solution to mold, centrifuge, treat with alkaline solution, dry, add photosensitizer solution, centrifuge, dry, add hyaluronic acid salt solution 1, centrifuge, add hyaluronic acid salt solution 2, then centrifuge, dry and fix to obtain intermediate product;

[0031] S2. Add the fatty acid solution of the antibacterial drug to a blank mold, centrifuge, and obtain a mold containing the antibacterial drug solution;

[0032] S3. The intermediate product is placed in the mold containing the antibacterial drug solution, then dried, and this process is repeated several times to obtain the antifungal microneedle patch.

[0033] Further, in step S1, the concentration of the chitosan solution is 3-5%; and the concentration of the photosensitizer solution is 0.01-0.2%.

[0034] Further, in step S1, the concentration of hyaluronic acid solution 1 is 40-60%, and the concentration of hyaluronic acid solution 2 is 10-30%.

[0035] Furthermore, in step S2, the content of the antibacterial drug in the fatty acid solution of the antibacterial drug is 10-20%.

[0036] Another object of the present invention is to provide the application of the above-mentioned antifungal microneedle patch in antifungal drugs, medical devices and disinfection products.

[0037] The present invention has the following beneficial effects:

[0038] The antifungal microneedle patch of this invention incorporates indocyanine green and itraconazole into the microneedles. Through the synergistic effects of photothermal, photodynamic, and antibiotic action, it achieves a significant enhancement in antifungal efficacy, reducing the dosage of itraconazole and lowering the risk of fungal resistance. Furthermore, this invention is a detachable microneedle that can remain embedded in the skin, eliminating the need for continuous application and effectively reducing discomfort. In addition, the use of lauric acid, a phase change material, to encapsulate itraconazole enables on-demand photothermal drug release, demonstrating promising application prospects. Attached Figure Description

[0039] Figure 1 The antifungal microneedle patch prepared in Example 1 of the present invention is shown;

[0040] in,

[0041] Figure 1 a is a schematic diagram of the preparation method of antifungal microneedle patches;

[0042] Figure 1 b is a photo of the antifungal microneedle patch;

[0043] Figure 1 c and d are bright-field micrographs of the antifungal microneedle patch;

[0044] Figure 1 e is a fluorescence micrograph after using Nile Red as a model drug;

[0045] Figure 1 f is a scanning electron microscope image; the left side shows the image without drug loading, and the right side shows the image with drug loading.

[0046] Figure 1 g is a microscopic image of a skin tissue section after insertion into pig skin (scale bar: 10 μm).

[0047] Figure 2 The near-infrared light response behavior of CS(ICG)@LA(ITZ)MNs is shown;

[0048] in,

[0049] Figure 2 a represents CS(ICG)@LA(ITZ)MNs containing different amounts of ICG under an 808nm laser (1W / cm²). 2 Temperature rise curve under irradiation;

[0050] Figure 2 b represents the temperature rise curves of CS(ICG)@LA(ITZ)MNs under irradiation with 808nm lasers of different powers;

[0051] Figure 2 c represents CS(ICG)@LA(ITZ)MNs under three 808nm laser beams (1W / cm²). 2 Temperature rise / fall curves under on / off cycles;

[0052] Figure 2 d represents the drug release curve of the CS(ICG)@LA needle under on / off 808nm laser irradiation (n=3);

[0053] Figure 2 e represents the reactive oxygen species probe DPBF solution and different samples at 808 nm (1W / cm). 2 DPBF balance (%) - time curve after laser irradiation;

[0054] Figure 2 f represents the reactive oxygen species probe SOSG solution and different samples at 808 nm (1W / cm). 2 Fluorescence spectrum after laser irradiation.

[0055] Figure 3 The results of in vitro antifungal activity tests are shown;

[0056] in,

[0057] Figure 3 a is a colony count diagram of the standard strain of Candida albicans (ATCC 64548) after different sample treatments;

[0058] Figure 3 b is a photograph of the growth of Candida albicans standard strain (ATCC 64548) colonies on an agar plate after different sample treatments;

[0059] Figure 3 c is a confocal micrograph of the standard strain of Candida albicans (ATCC 64548) after different sample treatments and subsequent live / dead staining (scale bar: 10 μm);

[0060] Figure 3 d represents the colony count for each of the five batches of Candida albicans standard strain (ATCC64548) treated with the same CS(ICG)@LA(ITZ)MNs.

[0061] Figure 3 e represents the colony count of Candida albicans itraconazole-resistant strain (ATCC 64550) after different sample treatments;

[0062] Figure 3 f shows the growth photos of colonies of Candida albicans itraconazole-resistant strain (ATCC 64550) on agar plates after different sample treatments;

[0063] Figure 3 g represents the colony count of chromogenic budding bacteria (cbs269.37, SUMS0310) after different sample treatments;

[0064] Figure 3 h is a photograph of the growth of colonies of chromogenic blastobacteria (cbs269.37, SUMS0310) on an agar plate after different sample treatments.

[0065] Figure 4 The results of in vitro anti-biofilm activity tests are shown;

[0066] in,

[0067] Figure 4 a is a 3D image of a biofilm after staining for live / dead (SYTO9 / PI);

[0068] Figure 4 b is a photograph of the biological membrane after crystal violet staining;

[0069] Figure 4 c represents the absorbance (OD) of the biological membrane after crystal violet staining. 595 ).

[0070] Figure 5 The results of the antibacterial mechanism study are shown;

[0071] in,

[0072] Figure 5 a represents the colony count in the antibacterial test (n=3) with / without the addition of vitamin C (Vc) peroxide quencher;

[0073] Figure 5 b is a bright-field / fluorescence field confocal micrograph of intracellular peroxides in Candida albicans ATCC 64548 after co-incubation with DCFH-DA (scale bar: 10 μm).

[0074] Figure 6 The results of in vivo antifungal activity studies are shown;

[0075] in,

[0076] Figure 6 a is a schematic diagram of the in vivo infection and treatment process;

[0077] Figure 6 b is an in vivo thermal imaging image;

[0078] Figure 6 c represents the colony count of the in vivo standard strain of Candida albicans ATCC 64548 after different sample treatments.

[0079] Figure 6 Image d shows the growth of the in vivo Candida albicans standard strain ATCC 64548 on an agar plate after different sample treatments. Detailed Implementation

[0080] To more clearly illustrate the technical solution of the present invention, the following embodiments are provided. Unless otherwise stated, the raw materials, reactions, and post-processing methods appearing in the embodiments are all common raw materials in the same field both domestically and internationally, as well as technical methods well known to those skilled in the art.

[0081] The terms "preferred," "more preferably," "more suitable," etc., used in this invention refer to embodiments of the invention that provide certain beneficial effects under certain circumstances. However, other embodiments may also be preferred in the same or other circumstances. Furthermore, the description of one or more preferred embodiments does not imply that other embodiments are unavailable, nor is it intended to exclude other embodiments from the scope of the invention. It should be understood that, except in any operational instance or otherwise indicated, all figures representing the amounts of ingredients used, for example, in the specification and claims, should be understood to be modified by the term "about" in all cases. Therefore, unless indicated to the contrary, the numerical parameters set forth in the following specification and appended claims are approximate values ​​varying with the desired performance to be obtained according to the invention.

[0082] The itraconazole (ITZ), chitosan (CS, degree of deacetylation ≥95%, viscosity 100-200 apm.s) and vitamin C used in the embodiments of this invention were purchased from Shanghai Maclean Biochemical Technology Co., Ltd.

[0083] Indocyanine green (ICG) and sodium hyaluronate (HA, 10kDa / 150-250kDa) were purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0084] Methanol and dichloromethane were purchased from Guangzhou Chemical Reagent Factory;

[0085] Sabouraud broth medium (SDB) was purchased from Beijing Solarbio Science & Technology Co., Ltd.

[0086] 1640 culture medium was purchased from Gibco, Inc., USA.

[0087] SOSG was purchased from Dalian Meilun Biotechnology Co., Ltd.

[0088] LIVE / The BacLight™ Bacterial Viability Kit was purchased from Invitrogen Life Sciences, Inc., USA.

[0089] DCFH-DA was purchased from MCE (MedChemExpress).

[0090] The Candida albicans standard strain (ATCC 64548), the Candida albicans itraconazole-resistant strain (ATCC 64550), and the Chromogenic Blastodiformis standard strain (cbs269.37, SUMS0310) in the test examples of this invention were all kindly provided by Professor Lu Sha of Sun Yat-sen Memorial Medical School, Sun Yat-sen University.

[0091] Example 1

[0092] An antifungal microneedle patch includes a backing layer and needles disposed on the surface of the backing layer;

[0093] The needles are arranged in a 10×10 array on the surface of the backing layer;

[0094] The needle surface includes an LA coating containing ITZ;

[0095] The needle contains the photosensitizer ICG.

[0096] The needle is made of CS material, and the backing layer is made of HA material.

[0097] The preparation method of the above-mentioned antifungal microneedle patch includes the following steps:

[0098] S1. Add 200 μL of 4% CS solution (prepared with 1% glacial acetic acid) to the PDMS microneedle mold, centrifuge for 30 min (4500 rpm, 20℃) using a basket centrifuge, remove excess CS solution after centrifugation, and centrifuge again for 5 min (4500 rpm, 20℃).

[0099] Then add 200 μL of 10% NaOH solution, centrifuge for 5 min (4500 rpm, 20℃), then soak the mold in 10% NaOH solution for 1 h, take it out and rinse with pure water, and dry it in an oven at 37℃ for 15 min;

[0100] Then add 200 μL of 1 mg / mL ICG (methanol solution), centrifuge for 20 min (4500 rpm, 20 °C) to remove excess ICG (methanol solution), and dry at 37 °C for 5 min;

[0101] Then add 150 μL of 50% HA solution (10 kDa), centrifuge for 5 min (3500 rpm, 20℃) to remove excess 50% HA solution (10 kDa); finally add 250 μL of 20% HA solution (150 kDa), centrifuge for 30 min (3500 rpm, 20℃), and dry in a 37℃ oven to obtain the intermediate product, namely CS(ICG)MNs;

[0102] S2. Prepare an ITZ solution containing 15% ITZ and 10% LA (solvents are 70% methanol and 30% dichloromethane); add 200 μL of the ITZ solution to a blank mold, centrifuge for 1 min (3500 rpm, 20 °C) using a basket centrifuge to remove excess solution and obtain a mold containing the antibacterial solution;

[0103] S3. Press the intermediate product into the mold containing the antibacterial drug solution, dry it in an oven at 37°C, and repeat the above drug loading step twice to obtain the antifungal microneedle patch, namely CS(ICG)@LA(ITZ)MNs.

[0104] Figure 1 The antifungal microneedle patch prepared in Example 1 of the present invention is shown;

[0105] in,

[0106] Figure 1 a is a schematic diagram of the preparation method of antifungal microneedle patches;

[0107] Figure 1 b is a photo of the antifungal microneedle patch;

[0108] Figure 1 c and d are bright-field micrographs of the antifungal microneedle patch;

[0109] Figure 1 e is a fluorescence micrograph after using Nile Red as a model drug;

[0110] Figure 1 f is a scanning electron microscope image; the left side shows the image without drug loading, and the right side shows the image with drug loading.

[0111] Figure 1 g is a microscopic image of a skin tissue section after insertion into pig skin (scale bar: 10 μm).

[0112] according to Figure 1 It can be concluded that the CS(ICG)@LA(ITZ)MNs prepared in this invention consists of a 10×10 array with an area of ​​13×13 mm. 2 ( Figure 1 b). Stereoscopic images show that the needle tips are uniform, each needle is a prismatic shape with a pointed tip, approximately 1200 μm in height, approximately 350 μm in base side length, and approximately 1000 μm in tip-to-tip distance. The needles exhibit a distinct double-layered structure. Figure 1 c and Figure 1 d). To characterize whether the needle tip surface was coated with a lauric acid coating containing itraconazole, Nile red was used as a model drug. Under the fluorescence field of a stereomicroscope, a uniform red color was observed on the needle tip surface, indicating that Nile red could be uniformly coated on the needle tip surface. Figure 1 e). Meanwhile, scanning electron microscopy images showed that the surface of the needle tip without lauric acid coating was smoother, while the surface of the needle tip with lauric acid coating was rougher, which also indicates that the needle tip surface was successfully coated with lauric acid. Figure 1 f).

[0113] One of the characteristics of microneedles is their ability to penetrate the stratum corneum for drug delivery. To characterize whether CS(ICG)@LA(ITZ)MNs have sufficient mechanical strength for skin insertion, we conducted the following test: Flat pig skin was taken, its fat removed, and cut into uniformly thick pieces. CS(ICG)@LA(ITZ)MNs were pressed vertically onto the skin for 8 minutes, then left on for another 20 minutes before removing the backing layer. The pig skin containing the needles was embedded in an embedding medium (OCT) and frozen at -80°C. The pig skin was then sectioned into 10 μm sections using a frozen section microtome. The sections were observed using a laser confocal microscope.

[0114] A tear-like indentation can be seen on the skin at the microneedle insertion site, with the needle tip remaining in the indentation. This indicates that the mechanical strength of CS(ICG)@LA(ITZ)MNs is sufficient to pierce the skin, and as a detachable microneedle, it also allows the needle tip to remain inside the skin after insertion. Figure 1 g).

[0115] Test Example 1

[0116] Determination of drug loading in CS(ICG)@LA(ITZ)MNs

[0117] Test method:

[0118] Indocyanine green (ICG): First, an ICG standard curve was plotted: ICG (DMSO) solutions with concentrations of 10, 5, 2.5, 1.25, 0.625, 0.3125, and 0.15625 μg / mL were prepared, and their absorption peaks were measured using a UV spectrophotometer. The absorbance at 794 nm was recorded, and a concentration-absorbance standard curve was plotted. In Example 1, excess indocyanine green was scraped from the mold and dissolved in DMSO. The absorbance of the solution at 794 nm was measured using a UV spectrophotometer. The ICG concentration was calculated by substituting the absorbance into the standard curve. The ICG loading per microneedle was finally calculated by subtracting the remaining ICG from the initial total amount added.

[0119] Itraconazole (ITZ): First, an ITZ standard curve was plotted: ITZ (methanol) solutions with concentrations of 4.496, 11.24, 17.984, 24.728, 31.472, and 3.216 μg / mL were prepared. The absorption peaks were measured using a UV spectrophotometer, and the absorbance at 266 nm was recorded. A concentration-absorbance standard curve was plotted. CS(ICG)@LA(ITZ)MNs were then immersed in 500 μL of methanol. The absorbance of the solution at 266 nm was measured using a UV spectrophotometer. The ITZ concentration was calculated by substituting the absorbance into the standard curve, and finally, the ITZ loading per microneedle was calculated.

[0120] The test results are shown in Table 1.

[0121] Table 1. Drug loading of CS(ICG)@LA(ITZ)MNs

[0122]

[0123]

[0124] Quantitative studies using a UV spectrophotometer revealed that each CS(ICG)@LA(ITZ)MNs patch contained 96.38±19.65 μg of indocyanine green (ICG) and 15.19±0.62 μg of itraconazole (ITZ).

[0125] Test Example 2

[0126] Near-infrared light response behavior of CS(ICG)@LA(ITZ)MNs

[0127] Test method:

[0128] (1) Photothermal behavior of CS(ICG)@LA(ITZ)MNs

[0129] Photothermal behavior dependent on ICG concentration: CS(ICG)@LA(ITZ)MNs with ICG concentrations of 0, 25, 50, 100, and 200 μg were prepared, and the photothermal behavior was investigated using 808 nm (1 W / cm²). 2 The laser was used to irradiate the microneedles, and a thermal imager was used to detect the temperature of the microneedles. The temperature was recorded every 5 seconds, and a temperature rise curve was plotted.

[0130] Dependence of photothermal behavior on laser power: Several CS(ICG)@LA(ITZ)MNs with an ICG content of 100 μg were prepared, and laser power was applied at 0, 300, 500, 700, and 1000 mW / cm². 2 The microneedles were irradiated with an 808nm laser, and the temperature of the microneedles was detected using a thermal imager. The temperature was recorded every 10 seconds, and a temperature rise curve was plotted.

[0131] (2) Photothermal drug release behavior of CS(ICG)@LA(ITZ)MNs

[0132] Press the microneedle patch onto a slightly damp paper towel for 2 minutes, then leave it on for another 5 minutes. Remove the backing; the microneedle tips will remain in the paper towel. Use a small scraper to remove the tips (this method will be used to remove the tips from the microneedle patch in subsequent examples). Place the patch in a 96-well plate (one needle per well), and add 250 μL of 1640 culture medium to each well. Perform three laser on / off cycles, irradiating with an 808 nm laser at minutes 53, 113, and 173 for 7 minutes each time, maintaining the temperature at 50 ± 2 °C. The control group received no laser irradiation. At minutes 30, 60, 90, 120, and 180, remove 200 μL of the solution from each well into centrifuge tubes, and then add another 200 μL of 1640 culture medium to each well. Add 300 μL of dichloromethane to a centrifuge tube, shake thoroughly, and allow to stand for extraction. Measure the absorbance of the dichloromethane layer at 268 nm using a UV spectrophotometer. Substitute the absorbance into the itraconazole standard curve to calculate the amount of itraconazole released.

[0133] (3) Photodynamic properties of CS(ICG)MNs

[0134] Remove the needle containing 200 μg ICG in CS(ICG)MNs and place it in a 96-well plate for later use.

[0135] DPBF peroxide determination: Four experimental groups were set up: ① H2O + NIR (near-infrared light); ② Free ICG (total content 200 μg) + NIR; ③ CS(ICG)MNs without NIR; ④ CS(ICG)MNs + NIR. 475 μL of pure water and 25 μL of 2 mg / mL DPBF (DMSO) solution were added to each well. Groups ①, ②, and ④ were measured using 808 nm (1 W / cm²). 2 After laser irradiation for 30 minutes, the absorption peak of the solution was measured every 5 minutes using an ultraviolet spectrophotometer, and the absorbance at 420 nm was recorded.

[0136] SOSG determination of peroxides: Four experimental groups were set up: ① H₂O + NIR; ② Free ICG (total content 200 μg) + NIR; ③ CS(ICG)MNs without NIR; ④ CS(ICG)MNs + NIR. Each group was treated with 500 μL of pure water and 1 μL of SOSG (10 μM) methanol solution. Groups ①, ②, and ④ were measured using an 808 nm (1 W / cm) wavelength. 2 After 15 minutes of laser irradiation, the absorption peaks of each group of solutions at 500-600 nm (EX = 488 nm) were recorded using a fluorescence spectrophotometer.

[0137] Figure 2 The near-infrared light response behavior of CS(ICG)@LA(ITZ)MNs is shown;

[0138] in,

[0139] Figure 2 a represents CS(ICG)@LA(ITZ)MNs containing different amounts of ICG under an 808nm laser (1W / cm²). 2 Temperature rise curve under irradiation;

[0140] Figure 2 b represents the temperature rise curves of CS(ICG)@LA(ITZ)MNs under irradiation with 808nm lasers of different powers;

[0141] Figure 2 c represents CS(ICG)@LA(ITZ)MNs under three 808nm laser beams (1W / cm²). 2 Temperature rise / fall curves under on / off cycles;

[0142] Figure 2 d represents the drug release curve of the CS(ICG)@LA needle under on / off 808nm laser irradiation (n=3);

[0143] Figure 2 e represents the reactive oxygen species probe DPBF solution and different samples at 808 nm (1W / cm). 2 DPBF balance (%) - time curve after laser irradiation;

[0144] Figure 2 f represents the reactive oxygen species probe SOSG solution and different samples at 808 nm (1W / cm). 2 Fluorescence spectrum after laser irradiation.

[0145] according to Figure 2 We can conclude that:

[0146] Due to the inclusion of the near-infrared fluorescent dye indocyanine green, CS(ICG)@LA(ITZ)MNs exhibit certain near-infrared photoresponsiveness. Under laser irradiation at a wavelength of 808 nm, CS(ICG)@LA(ITZ)MNs can rapidly heat up, and this heating behavior is ICG concentration-dependent. Figure 2 a) and laser power dependence ( Figure 2 b) This demonstrates that CS(ICG)@LA(ITZ)MNs possesses excellent photothermal properties. CS(ICG)@LA(ITZ)MNs were subjected to three 808nm laser pulses (1W / cm²). 2 Three similar heating / cooling curves were obtained by cyclic irradiation (on / off), indicating that CS(ICG)@LA(ITZ)MNs exhibit similar heating behavior in three repeated laser irradiations, demonstrating good photothermal stability. Figure 2 c).

[0147] In this invention, lauric acid is used to encapsulate itraconazole. Lauric acid is a phase change material; when the temperature exceeds 44°C, lauric acid changes from a solid phase to a liquid phase, releasing itraconazole, thereby achieving photothermal controlled release. The release curves show that, over a certain time period, the laser-irradiated group released more drug than the non-laser-irradiated group. Furthermore, within the laser-irradiated group, the drug release rate was higher during the laser-irradiated phase than during the non-laser-irradiated phase. Figure 2 d). This indicates that CS(ICG)@LA(ITZ)MNs has a certain photothermal controlled release effect, which can realize on-demand drug release.

[0148] To investigate the photodynamic effects of CS(ICG)MNs, two reactive oxygen species (ROS) detection probes, DPBF and SOSG, were selected. DPBF can recognize various ROS; once bound to an ROS, it undergoes irreversible oxidation, resulting in a rapid decrease in UV-vis (420 nm) absorption intensity. SOSG highly selectively binds to singlet oxygen; in the presence of singlet oxygen, SOSG produces green fluorescence. CS(ICG)MNs were co-incubated with DPBF and then subjected to an 808 nm laser (1 W / cm²). 2 When irradiated with laser light, the absorbance of the solution at 420 nm decreased significantly, indicating the formation of peroxides. Without laser irradiation, almost no peroxides were produced. Figure 2e). It is worth noting that free ICG did not produce peroxides after laser irradiation. This may be because high concentrations of ICG molecules undergo aggregation and quenching, preventing peroxide formation. The porous structure of the chitosan needle allows for better dispersion of ICG, limiting aggregation and quenching, thus enhancing its photodynamic effect. Similar results were obtained when using SOSG as a probe: CS(ICG)MNs combined with 808nm laser irradiation produced peroxides, while CS(ICG)MNs without laser irradiation or free ICG under laser irradiation produced almost no peroxides. Figure 2 f). In summary, this microneedle can generate a large amount of reactive oxygen species that are harmful to fungi through photodynamic therapy, which can be used to achieve the synergistic antifungal effect of the microneedle.

[0149] Test Example 3

[0150] In vitro antifungal activity

[0151] Test method:

[0152] The needles from CS@LA(ITZ)MNs (microneedles containing only itraconazole), CS(ICG)MNs (microneedles containing only indocyanine green), and CS(ICG)@LA(ITZ)MNs patches were removed and placed in 96-well plates (one needle from one patch was placed in one well). Four experimental groups were set up: ①CS@LA(ITZ)MNs; ②CS(ICG)MNs; ③CS(ICG)@LA(ITZ)MNs; ④blank control. 100 μL of SDB and 100 μL of bacterial suspension (Candida albicans standard strain ATCC 645481 × 10⁻⁶) were added to all wells. 8 CFU / mL, Candida albicans itraconazole-resistant strain ATCC 64550 1×10 8 CFU / mL, chromogenic blastomyces standard strain cbs269.37 1×10 7 CFU / mL). Each group has 6 wells, 3 of which are treated with an 808nm laser (1W / cm). 2 Irradiate for 90 seconds, leaving 3 wells un-irradiated. After irradiation, incubate at 37°C with shaking for 1 hour. Then, dilute the bacterial suspension appropriately and spread it evenly on SDB agar plates. Incubate at 37°C for 24 hours, and then count the colonies.

[0153] Using LIVE / The BacLight™ Bacterial Viability Kit was used to detect the viability of three fungi—ATCC 64548, ATCC 64550, and a chromogenic blastomyces standard strain—after different sample treatments. After treating the fungi using the same method as described above, all wells from the same group of fungi were collected in a centrifuge tube, centrifuged (4500g, 4℃, 10min), and the supernatant was removed. Then, 100μL of dye (1mL PBS + 1.5μL PI + 1.5μL SYTO9) was added, and the mixture was incubated in a metal bath for 15min (37℃, 200rpm). 10μL of the bacterial culture was then added to a glass slide, covered with a coverslip, and inverted under a laser confocal microscope for observation using a 63x objective lens.

[0154] To investigate the antibacterial effect of CS(ICG)@LA(ITZ)MNs after multiple uses, the needle tip of CS(ICG)@LA(ITZ)MNs was placed in a 96-well plate, and 200 μL of Candida albicans suspension (SDB, 1×10⁻⁶) was added. 8 CFU / mL), 808nm (1W / cm) 2 Irradiate with laser for 2 minutes. After irradiation, aspirate the bacterial culture and place it in another 96-well plate. Add 200 μL of Candida albicans (SDB, 1×10⁻⁶) to the plate again. 8 CFU / mL), 808nm (1W / cm) 2 Irradiate for 2 minutes, and repeat the above steps five times. After five irradiations, place the 96-well plates containing the bacterial suspensions treated with CS(ICG)@LA(ITZ)MNs five times and the untreated bacterial suspensions in a 37°C incubator and incubate with shaking for 1 hour. Then, dilute the bacterial suspensions appropriately and spread them evenly on SDB agar plates. Incubate at 37°C for 24 hours, and then count the colonies.

[0155] Figure 3 The results of in vitro antifungal activity tests are shown;

[0156] in,

[0157] Figure 3 a is a colony count diagram of the standard strain of Candida albicans (ATCC 64548) after different sample treatments;

[0158] Figure 3 b is a photograph of the growth of Candida albicans standard strain (ATCC 64548) colonies on an agar plate after different sample treatments;

[0159] Figure 3 c is a confocal micrograph of the standard strain of Candida albicans (ATCC 64548) after different sample treatments and subsequent live / dead staining (scale bar: 10 μm);

[0160] Figure 3 d represents the colony count for each of the five batches of Candida albicans standard strain (ATCC64548) treated with the same CS(ICG)@LA(ITZ)MNs.

[0161] Figure 3 e represents the colony count of Candida albicans itraconazole-resistant strain (ATCC 64550) after different sample treatments;

[0162] Figure 3 f shows the growth photos of colonies of Candida albicans itraconazole-resistant strain (ATCC 64550) on agar plates after different sample treatments;

[0163] Figure 3 g represents the colony count of chromogenic budding bacteria (cbs269.37, SUMS0310) after different sample treatments;

[0164] Figure 3 h is a photograph of the growth of colonies of chromogenic blastobacteria (cbs269.37, SUMS0310) on an agar plate after different sample treatments.

[0165] according to Figure 3 It can be concluded that without 808nm laser irradiation, all groups showed no significant antibacterial effect against the standard strain of Candida albicans (ATCC 64548); with 808nm laser irradiation (1W / cm²), the antibacterial effect was significantly reduced. 2 After irradiation, CS@LA(ITZ)MNs still showed no significant antibacterial effect, while CS(ICG)MNs exhibited some antibacterial activity, inhibiting at least 90% of colonies. CS(ICG)@LA(ITZ)MNs showed the best antibacterial effect, inhibiting over 99.9% of colonies. Figure 3 a and Figure 3 b).

[0166] After processing ATCC 64548 with different samples, it was then used with LIVE / Staining was performed using the BacLight™ kit with a mixture of SYTO 9 and PI in appropriate proportions. Bacteria with intact membrane structures and high activity exhibited green fluorescence, while bacteria with damaged membrane structures and low activity exhibited red fluorescence. Observation was performed using a laser confocal microscope after staining. Results showed that fungi treated with CS(ICG)@LA(ITZ)MNs+NIR exhibited the strongest red fluorescence and the weakest activity. Some fungi treated with CS(ICG)MNs+NIR showed red fluorescence, while the remaining groups showed weaker red fluorescence. Figure 3 c). The experimental results are basically consistent with the colony count results, and also indicate that the antibacterial mechanism of CS(ICG)@LA(ITZ)MNs+NIR is related to the disruption of fungal cell membranes.

[0167] Because the microneedles designed in this invention can leave the needle tip inside the skin, it is desirable to be able to perform multiple laser irradiations to achieve repeated treatments. To explore the feasibility of multiple treatments, the same CS(ICG)@LA(ITZ)MNs was repeatedly used with an 808nm laser to treat fungi five times. The colony count results show that even after five treatments, the antibacterial rate of CS(ICG)@LA(ITZ)MNs remained as high as 99.9%. Figure 3 d) indicates that it can exert its antibacterial effect multiple times, thus achieving multiple treatments.

[0168] The antifungal effects of the micro-target on two types of fungi—itraconazole-resistant Candida albicans (ATCC 64550) and standard strains of Chromogenic Blastomonas (Fonsecaeamonophora (cbs269.37, SUMS0310))—showed that the antifungal performance of the micro-target against drug-resistant Candida albicans was similar to that against standard Candida albicans. Figure 3 e and 3f). For *Cycloblastomyces*, without 808 nm laser irradiation, both the CS@LA(ITZ)MNs and CS(ICG)@LA(ITZ)MNs groups containing itraconazole showed some antibacterial activity, inhibiting approximately 60% of colonies. This may be because *Cycloblastomyces* is more sensitive to itraconazole than *Candida albicans*. After laser irradiation, both the CS(ICG)MNs and CS(ICG)@LA(ITZ)MNs groups also showed antibacterial activity, with the CS(ICG)@LA(ITZ)MNs group exhibiting the best antibacterial effect, inhibiting over 95% of colonies. Figure 3 (g and 3h). The above antibacterial experimental results show that CS(ICG)@LA(ITZ)MNs combined with 808nm laser irradiation can produce a very good antifungal effect, which is speculated to be the synergistic effect of itraconazole, photothermal and peroxide.

[0169] Test Example 4

[0170] In vitro anti-biofilm effect

[0171] Test method:

[0172] Live / dead staining: 1 mL of Candida albicans ATCC 64548 (SDB, 1×10⁻⁶) was used for staining. 5 CFU / mL was added to a confocal dish and incubated at 37°C for 48 h. Four experimental groups were set up: ① blank control; ② CS@LA(ITZ)MNs; ③ CS(ICG)MNs; ④ CS(ICG)@LA(ITZ)MNs. After biofilm formation, the culture medium was aspirated, microneedles were inserted into the biofilm, and 100 μL SDB was added at 808 nm (1 W / cm²) per group. 2Irradiate with laser for 2 min, controlling the temperature at 50±2℃. Remove the microneedle patch, add 1 mL of dye (1 mL PBS + 1.5 μL PI + 1.5 μL SYTO9) and incubate at 37℃ for 20 min. Then use a laser confocal microscope to scan and obtain a 3D image of the biofilm.

[0173] Biomass loss: 1 mL of CA (SDB, 1×10⁻⁶) was added. 5 CFU / mL was added to 12-well plates and incubated at 37°C for 48 h. Four experimental groups were set up: ① blank control; ② CS@LA(ITZ)MNs; ③ CS(ICG)MNs; ④ CS(ICG)@LA(ITZ)MNs. After biofilm formation, the culture medium was aspirated, microneedles were inserted into the biofilm, and 100 μL of SDB was added. Each group was incubated at 808 nm (1 W / cm²). 2 Irradiate with laser for 2 min, maintaining the temperature at 50±2℃. Remove the microneedle patch, add 1 mL of SDB, and incubate in a shaking incubator for 1 h (30℃, 190 rpm). Aspirate the SDB, open the plate, and dry it in a clean bench for 10 min. Then add 600 μL of 1% crystal violet (PBS) solution and incubate at room temperature for 1 h. Aspirate excess crystal violet, add anhydrous ethanol, and shake at 30℃ for 30 min. Aspirate 200 μL of ethanol into a 96-well plate, dilute 10-fold, and measure the absorbance at 595 nm using a microplate reader.

[0174] Among them, CS@LA(ITZ)MNs are microneedles containing only itraconazole, CS(ICG)MNs are microneedles containing only indocyanine green, and CS(ICG)@LA(ITZ)MNs are microneedles containing both itraconazole and indocyanine green.

[0175] Figure 4 The results of in vitro anti-biofilm activity tests are shown;

[0176] in,

[0177] Figure 4 a is a 3D image of a biofilm after staining for live / dead (SYTO9 / PI);

[0178] Figure 4 b is a photograph of the biological membrane after crystal violet staining;

[0179] Figure 4 c represents the absorbance (OD) of the biological membrane after crystal violet staining. 595 ).

[0180] according to Figure 4It can be concluded that the biofilms in the blank control group and the CS@LA(ITZ)MNs group exhibited widespread green fluorescence, indicating that the number of dead bacteria was extremely small; the CS(ICG)MNs+NIR group showed partial red fluorescence, indicating that some bacteria in the biofilm had died; the CS(ICG)@LA(ITZ)MNs+NIR group exhibited widespread red fluorescence, indicating that a significant number of bacteria in the biofilm had died. Figure 4 a). Crystal violet can stain the extracellular matrix of biofilms. Results showed that biofilms treated with the blank control group and the CS@LA(ITZ)MNs group retained most of their activity and structural integrity. In contrast, the CS(ICG)MNs+NIR and CS(ICG)@LA(ITZ)MNs+NIR groups disrupted the compact biofilm and led to significant biomass loss. Figure 4 (b, c) The above experiments all demonstrate that CS(ICG)@LA(ITZ)MNs, after near-infrared light irradiation, exhibit a stronger antifungal biofilm effect compared to microneedles containing only drugs or photosensitizers.

[0181] Test Example 5

[0182] Research on antibacterial mechanisms

[0183] To investigate the effect of peroxides on antibacterial efficacy, vitamin C was used to quench peroxides. The needle was removed and placed in a 96-well plate, with five experimental groups: ① blank control group; ② CS@LA(ITZ)MNs; ③ free ICG; ④ CS(ICG)MNs; ⑤ CS(ICG)@LA(ITZ)MNs. Each group was further divided into a vitamin C-containing group (to quench peroxides) and a vitamin C-free group. The Candida albicans in the vitamin C-containing group had been co-incubated with 10 mM vitamin C for 24 hours. Groups ①②④⑤ were each added with 100 μL of SDB and 100 μL of Candida albicans bacterial suspension (SDB, ATCC 64548, 1×10⁻⁶). 8 Group ③ received 50 μL of ICG (2 mg / mL, DMSO), 50 μL of SDB, and 100 μL of Candida albicans suspension (SDB, ATCC 64548, 1×10⁻⁶ CFU / mL). 8 CFU / mL). All groups were treated at 808nm (1W / cm). 2 Irradiate with laser for 2 minutes. After irradiation, incubate with shaking in a 37°C incubator for 50 minutes. Then, dilute the bacterial solution to an appropriate ratio and spread it evenly on an SDB agar plate. Incubate at 37°C for 24 hours, and then count the colonies.

[0184] To detect intracellular peroxides in fungi, a CA bacterial suspension (SDB, ATCC 64548, 10) was prepared. 8CFU / mL and 40 μmol / mL DCFH-DA were co-incubated at 37℃ for 1 h. 100 μL of this bacterial culture was then added to 96-well plates containing the following samples: ① blank culture medium; ② CS@LA(ITZ)MNs; ③ free ICG; ④ CS(ICG)MNs; ⑤ CS(ICG)@LA(ITZ)MNs; ⑥ CS(ICG)@LA(ITZ)MNs without NIR. Except for group ⑥, groups ①-⑤ were treated with an 808 nm laser (1 W / cm²). 2 Irradiate for 2 minutes, then incubate at 37°C with shaking for 15 minutes. Add 10 μL of bacterial culture to a glass slide, cover with a coverslip, invert the slide, and observe under a laser confocal microscope with a 63x objective lens.

[0185] Figure 5 The results of the antibacterial mechanism study are shown;

[0186] in,

[0187] Figure 5 a represents the colony count in the antibacterial test (n=3) with / without the addition of vitamin C (Vc) peroxide quencher;

[0188] Figure 5 b is a bright-field / fluorescence field confocal micrograph of intracellular peroxides in Candida albicans ATCC 64548 after co-incubation with DCFH-DA (scale bar: 10 μm).

[0189] according to Figure 5 It can be concluded from the colony count results that the antibacterial effect is CS(ICG)@LA(ITZ)MNs > CS(ICG)MNs > free ICG > CS@LA(ITZ)MNs. Free ICG has a weak antibacterial effect due to its photothermal effect. The antibacterial effect of CS(ICG)MNs is better than that of free ICG. Based on the previous photodynamic experiments, it is speculated that this is because CS(ICG)MNs can produce peroxides, while free ICG cannot. Combined with the results of Vc quenching peroxides, it can be seen that quenching peroxides will reduce the antibacterial effect of CS(ICG)MNs but has no effect on free ICG. This further proves that the bactericidal effect of CS(ICG)MNs is the combined effect of peroxides produced by photothermal and photodynamic effects. The antibacterial effect of CS(ICG)@LA(ITZ)MNs was significantly better than that of CS(ICG)MNs, indicating that the itraconazole released by photothermal activity played a role. After quenching with vitamin C, the antibacterial effect decreased significantly, and the decrease was greater than that of CS(ICG)MNs, indicating that peroxides participated in the antibacterial activity and had a synergistic effect with photothermal activity and itraconazole. Figure 5 a).

[0190] To further demonstrate the peroxides generated by ICG photodynamic therapy, the intracellular reactive oxygen species probe DCFH-DA was co-incubated with ATCC 64548. DCFH-DA, after being oxidized by the peroxides, becomes the fluorescent compound DCF. Confocal fluorescence microscopy revealed that some bacteria in the CS(ICG)MNs+NIR group produced intracellular peroxides, and almost all bacteria in the CS(ICG)@LA(ITZ)MNs+NIR group produced intracellular peroxides. However, the blank control group, the CS@LA(ITZ)MNs group, the free ICG group, and the CS(ICG)@LA(ITZ)MNs group without 808nm laser irradiation produced almost no intracellular peroxides. Figure 5 b).

[0191] In summary, when irradiated with an 808nm laser, CS(ICG)@LA(ITZ)MNs generate heat and peroxides. The heat generated can promote the release of itraconazole. Ultimately, the antifungal drug itraconazole, photothermal effect, and photodynamic effect work synergistically to achieve a significant antifungal effect.

[0192] Test Example 6

[0193] In vivo antifungal activity

[0194] Test method:

[0195] Female Balb-c mice aged 6-8 weeks were randomly divided into four groups (① no treatment; ② CS@LA(ITZ)MNs; ③ CS(ICG)MNs; ④ CS(ICG)@LA(ITZ)MNs), with four mice in each group. All hair on the back of the mice was removed. 24 hours after hair removal, 25 μL of 1×10⁻⁶ mice was injected intradermally into the back of the mice. 8 CFU / mL Candida albicans (ATCC 64548) PBS suspension was used. Twelve hours after infection, microneedle patches were inserted into the infection site, followed by 30 minutes of 808nm laser irradiation. Temperature changes were monitored using a thermal imaging camera. Twelve hours after treatment, mice were euthanized by spinal dislocation, and tissue from the infection site was collected, homogenized, and evenly spread on SDB agar plates. After incubation at 37°C for 24 hours, colonies were counted.

[0196] Figure 6 The results of in vivo antifungal activity studies are presented;

[0197] in,

[0198] Figure 6 a is a schematic diagram of the in vivo infection and treatment process;

[0199] Figure 6 b is an in vivo thermal imaging image;

[0200] Figure 6 c represents the colony count of the standard strain of Candida albicans ATCC 64548 after different sample treatments.

[0201] Figure 6 Image d shows the growth of the in vivo Candida albicans standard strain ATCC 64548 on an agar plate after different sample treatments.

[0202] according to Figure 6 It can be concluded that red bumps were observed on the backs of mice 12 hours after infection, indicating that the intradermal Candida albicans infection model was successfully established. Thermal imaging revealed that CS(ICG)MNs and CS(ICG)@LA(ITZ)MNs also exhibited good photothermal activity in vivo, rapidly heating to 53℃ under laser irradiation. Figure 6 b). Infected tissue was collected 12 hours after treatment, ground, and plated. Colony counting results showed that CS(ICG)@LA(ITZ)MNs could kill more than 90% of Candida albicans in the body, while CS@LA(ITZ)MNs and CS(ICG)MNs had slightly weaker antibacterial effects. Figure 6 (c, d). The experimental results indicate that, thanks to the synergistic effect of itraconazole with photothermal and photodynamic effects, CS(ICG)@LA(ITZ)MNs combined with 808nm laser irradiation also have a good antifungal effect in vivo.

[0203] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.

[0204] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

[0205] References:

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[0207] [2]Chandra J,Kuhn DM,Mukherjee PK,Hoyer LL,McCormick T,GhannoumMA.Biofilm formation by the fungal pathogen Candida albicans:development,architecture,and drug resistance.J Bacteriol.2001Sep;183(18):5385-94.

[0208] [3]Verma S,Utreja P.Vesicular nanocarrier based treatment of skinfungal infections:Potential and emerging trends in nanoscalepharmacotherapy.Asian J Pharm Sci.2019Mar;14(2):117-129.

[0209] [4]Kyle AA,Dahl MV.Topical therapy for fungal infections.Am J ClinDermatol.2004;5(6):443-51.

[0210] [5]Spernovasilis N,Kofteridis DP.Pre-Existing Liver Disease andToxicity of Antifungals.JFungi(Basel).2018Dec 10;4(4):133.

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Claims

1. An antifungal microneedle patch, characterized in that, The antifungal microneedle patch includes a backing layer and needles disposed on the surface of the backing layer; The needles form an n×n array on the surface of the backing layer; The needle surface includes a lauric acid coating containing an antibacterial agent; The needle contains a photosensitizer; The n is selected from integers; The preparation method of the antifungal microneedle patch includes the following steps: S1. Add the chitosan solution to the mold, centrifuge, treat with alkaline solution, dry, add photosensitizer solution, centrifuge, dry, add hyaluronic acid salt solution 1, centrifuge, add hyaluronic acid salt solution 2, then centrifuge, dry and fix to obtain intermediate product; S2. Add the lauric acid solution of the antibacterial drug to a blank mold, centrifuge, and obtain a mold containing the antibacterial drug solution; S3. The intermediate product is placed in the mold containing the antibacterial drug solution, then dried, and this process is repeated several times to obtain the antifungal microneedle patch.

2. The antifungal microneedle patch according to claim 1, characterized in that, The antibacterial drug is a lipophilic triazole antifungal drug.

3. The antifungal microneedle patch according to claim 2, characterized in that, The antibiotic in question is itraconazole.

4. The antifungal microneedle patch according to claim 1, characterized in that, The needle is made of chitosan, and the backing layer is made of hyaluronic acid salt.

5. The antifungal microneedle patch according to claim 1, characterized in that, The photosensitizer is indocyanine green.

6. The antifungal microneedle patch according to claim 1, characterized in that, In step S1, the concentration of the chitosan solution is 3-5%; the concentration of the photosensitizer solution is 0.01-0.2%; the concentration of hyaluronic acid salt solution 1 is 40-60%; and the concentration of hyaluronic acid salt solution 2 is 10-30%.

7. The antifungal microneedle patch according to claim 1, characterized in that, In step S2, the content of the antibacterial drug in the lauric acid solution is 10-20%.

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