A kind of acellular matrix hydrogel microneedle with biological activity and a preparation method thereof

Abstract

The application provides a kind of extracellular matrix hydrogel microneedle with biological activity and a preparation method thereof, and belongs to the technical field of medicine.The preparation method of the extracellular matrix hydrogel microneedle with biological activity of the application comprises the following steps: preparing a hydrogel from methacrylated porcine small intestinal submucosa (SISMA) after ultrasonic treatment, adding a photoinitiator to the prepared hydrogel, injecting the hydrogel into a microneedle mold, concentrating and solidifying, and obtaining the microneedle.The hydrogel microneedle has good biological activity and better swelling properties, and has good prospects in biomedicine.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a bioactive decellularized matrix hydrogel microneedle and its preparation method. Background Technology

[0002] Microneedles are an array of needle tips with a length of 0.2mm-2.0mm that can penetrate the stratum corneum of the epidermis with minimal stimulation of pain nerves, providing a transdermal drug delivery protocol for non-lipid-soluble, non-small molecule drugs.

[0003] Microneedle drug delivery technologies are diverse, primarily including surface coating, hollow microneedles, soluble microneedles, and hydrogel microneedles. Among these, hydrogel microneedles are a novel type, utilizing highly biocompatible hydrogels. They leverage the mechanical strength of the gel scaffold in its dry state to penetrate the stratum corneum, and utilize the hydrophilic properties of the hydrogel to absorb tissue fluid, swelling subcutaneously and releasing the drug in a sustained manner. Compared to other microneedle technologies, hydrogel microneedles offer advantages such as higher drug loading capacity, better biocompatibility, and more controllable drug release, making them a current hot topic in microneedle technology.

[0004] Currently, most materials used in the preparation of hydrogel microneedles are polyvinyl alcohol (PVA), polyethylene glycol (PEG), methacrylic anhydride gelatin (GelMA), and methacryloyl hyaluronic acid (HAMA). These materials have a single composition and lack bioactive substances, serving only as drug carriers and not promoting tissue repair themselves. Furthermore, existing hydrogel microneedle materials are mostly compounds with relatively small molecular weights, limiting their swelling properties (including swelling rate and swelling ratio).

[0005] The fabrication of hydrogel microneedles typically requires the use of high-concentration, high-flowability hydrogel materials to ensure the mechanical properties and surface morphology of the microneedles. Current techniques for preparing microneedles using bioactive materials such as extracellular matrix (ECM) usually only improve the hydrophilicity of ECM through enzymatic digestion and chemical modification. The resulting ECM materials still have limited hydrophilicity and often cannot form high-concentration, high-flowability solutions, thus failing to meet the requirements for microneedle fabrication. Furthermore, most ECM materials obtained by existing fabrication techniques are difficult to dissolve due to their high collagen content and have poor flowability, making them unsuitable for microneedle preparation.

[0006] Therefore, in order to obtain microneedles with bioactivity and better swelling properties, it is necessary to explore a method for preparing modified decellularized matrix materials (SISMA) with excellent solubility and flowability. Summary of the Invention

[0007] The purpose of this invention is to provide an extracellular matrix for the preparation of microneedles.

[0008] A bioactive decellularized matrix hydrogel microneedle is made from methacrylamide porcine small intestinal submucosa material;

[0009] The methacrylamide porcine small intestinal submucosa material is a methacrylic anhydride-grafted small intestinal submucosa with a grafting rate of 40-90%.

[0010] Preferably, the methacrylic anhydride grafting rate is 40-90%.

[0011] Preferably, the methacrylic anhydride grafting rate is 80%.

[0012] Preferably, it is prepared by reacting the submucosa of the small intestine with methacrylic anhydride in an alkaline aqueous solution.

[0013] Preferably, the molar ratio of free amino groups to methacrylic anhydride in the submucosa of the small intestine is 1:(5-100).

[0014] Preferably, the molar ratio of free amino groups to methacrylic anhydride in the submucosa of the small intestine is 1:20.

[0015] Preferably, the alkaline aqueous solution is an aqueous solution with a pH of 8 to 10.

[0016] Preferably, the reaction conditions are: 0-50°C for 1-24 hours.

[0017] Preferably, the reaction conditions are: 0-20℃ for 20-24 hours, 20-35℃ for 10-20 hours, and 35-50℃ for 1-10 hours.

[0018] Preferably, the amino content of the submucosa of the small intestine is 400–800 μM.

[0019] Preferably, the submucosal layer of the small intestine is a submucosal layer of the small intestine that has undergone defatting, decellularization, and digestion treatment.

[0020] Preferably, it is prepared by the following method:

[0021] (1) The methacrylamide porcine small intestinal submucosa material is mixed with a photoinitiator to obtain a mixture. The mixture is injected into a microneedle mold, defoamed, and concentrated in an oven.

[0022] The photoinitiator in the mixture has a mass-volume fraction of 0.05%-1%;

[0023] (2) The concentrated SISMA in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0024] Preferably, the photoinitiator is LAP or I2959.

[0025] Preferably, in step (1), the oven concentration temperature is 20℃~50℃; the oven concentration time is 12h~72h; and the photocuring time is 15s~1min.

[0026] The present invention also provides a method for preparing the above-mentioned hydrogel microneedles, comprising the following steps:

[0027] (1) The methacrylamide porcine small intestinal submucosa material is mixed with a photoinitiator to obtain a mixture. The mixture is injected into a microneedle mold, defoamed, and concentrated in an oven.

[0028] The photoinitiator in the mixture has a mass-volume fraction of 0.05%-1%;

[0029] (2) The concentrated SISMA in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0030] The present invention also provides the use of the above-described hydrogel microneedles in the preparation of drug delivery carriers.

[0031] This invention provides an ECM hydrogel microneedle and its preparation method, enabling the microneedle to act as a drug delivery carrier while possessing activities such as promoting angiogenesis, regulating immune stress, and promoting collagen deposition. Simultaneously, the rich collagen network structure of the ECM material improves the swelling properties of the microneedle, thereby enhancing its drug delivery efficiency.

[0032] Obviously, based on the above description of the present invention, and according to common technical knowledge and conventional methods in the field, various other modifications, substitutions or alterations can be made without departing from the basic technical concept of the present invention.

[0033] The following detailed embodiments further illustrate the above-described content of the present invention. However, this should not be construed as limiting the scope of the present invention to the following examples. All technologies implemented based on the above-described content of the present invention fall within the scope of the present invention. Attached Figure Description

[0034] Figure 1 The full view of SISMA microneedles under low magnification (1mm);

[0035] Figure 2 The side view of a SISMA microneedle under high magnification (200 μm);

[0036] Figure 3 Images of angiogenesis in different hydrogel microneedles;

[0037] Figure 4Histograms were plotted to show the number of vascular intersections, meshes, nodes, and total length of the main vascular trunk in HUVEC cells after treatment with different hydrogel microneedles. ns: P>0.05; *: P<0.05; **: P<0.01; ***: P<0.001; ****: P<0.0001.

[0038] Figure 5 The expression levels of inflammatory genes in RAW264.7 cells after treatment with different hydrogel microneedles; ns: P>0.05; **: P<0.01; ***: P<0.001; ****: P<0.0001

[0039] Figure 6 The results of experiments on the number of NIH-3T3 cells migrating after different hydrogel microneedle treatments;

[0040] Figure 7 The results of NIH-3T3 cell proliferation assay after different hydrogel microneedle treatments;

[0041] Figure 8 Results of experiments on the swelling properties of SISMA hydrogel microneedles;

[0042] Figure 9 This relates the ratio of raw materials to the grafting rate of the modified small intestinal submucosa in Experiment Example 1. Detailed Implementation

[0043] LAP phenyl(2,4,6-trimethylbenzoyl) lithium phosphate;

[0044] I2959: 2-Hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone;

[0045] The raw materials and equipment used in this invention are all known products, obtained by purchasing commercially available products.

[0046] The porcine small intestinal submucosa (SIS), the raw material of this invention, can be prepared by the following method:

[0047] 1) Take fresh pig small intestine, rinse away the contents with water, rub with salt, and rinse repeatedly with water 3 times. Use a scalpel to cut open the small intestine and cut it into 10-20cm long segments;

[0048] 2) Use a tongue depressor to scrape off the muscular and serosa layers of the small intestine, and store it in physiological saline at 4°C;

[0049] 3) After rinsing with deionized water and filtering, immerse it in a mixture of chloroform and methanol (chloroform:methanol = 1:1) and leave it at room temperature overnight.

[0050] 4) After repeated rinsing with deionized water, place the solution in a 0.25% trypsin solution and incubate overnight at 4°C.

[0051] 5) Rinse 10 times with deionized water and then treat with 0.5% SDS for at least 4 hours;

[0052] 6) After cleaning, freeze-dry at -70℃ for 24 hours to obtain the SIS membrane.

[0053] 7) The SIS membrane was pulverized at low temperature using a ball mill to obtain SIS powder.

[0054] 8) Digest the SIS powder with pepsin solution and freeze-dry it again under vacuum for later use.

[0055] Example 1: Preparation of methacrylamide porcine submucosal (SISMA) hydrogel microneedles

[0056] 1. Preparation of methacrylamide porcine small intestinal submucosal layer (SISMA) material

[0057] (1) Dissolve SIS in ultrapure water at a ratio of 1%, stir continuously, and after it is fully dissolved, add NaOH to adjust the pH to 9.

[0058] (2) After adding methacrylic anhydride (MA) dropwise in batches, NaOH was added to adjust the pH to the range of 8-9. The molar ratio of free amino groups (from SIS) to MA was 1:20. The mixture was obtained by ultrasonic treatment (ultrasonic power 100W, 4℃, 3h). Dialyzed and lyophilized for later use. In this embodiment, the amount of MA was calculated as follows: the content of free amino groups after SIS digestion was first determined by the OPA method, and the actual amount of MA was calculated based on the ratio of free amino group to MA of 1:20.

[0059] 2. Preparation of SISMA hydrogel microneedles using microneedle molds

[0060] (1) Inject SISMA mixed with photoinitiator LAP into a silicone resin (PDMS) microneedle mold. The concentration of LAP in the mixture is 0.25% (g / ml). Defoaming is performed under negative pressure, and the mixture is concentrated in an oven at 20°C for 12 hours.

[0061] (2) The concentrated SISMA (LAP-405nm, irradiated for 55s) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0062] Example 2: Preparation of methacrylamide porcine small intestinal submucosal (SISMA) hydrogel microneedles

[0063] The preparation method in this embodiment is the same as that in Example 1, except that the preparation method of SISMA is different. The SISMA in this embodiment is prepared according to the following method:

[0064] Dissolve SIS at a concentration of 0.25% in an ice-water bath for 4–6 hours. After the SIS is dissolved, add free amino groups and methacrylic anhydride (MA) at a molar ratio of 1:20, adjust the pH to 9–10, and react in an ice-water bath for 24 hours. Dialyze and freeze-dry for later use.

[0065] Example 3: Preparation of methacrylamide porcine small intestinal submucosal (SISMA) hydrogel microneedles

[0066] The preparation method in this embodiment is the same as that in Example 1, except that the preparation method of SISMA is different. The SISMA in this embodiment is prepared according to the following method:

[0067] Dissolve SIS at a concentration of 0.5% at room temperature. Add free amino groups and methacrylic anhydride (MA) at a molar ratio of 1:20. Adjust the pH to 9-10 and react at 25°C for 12 hours. Dialyze and lyophilize for later use.

[0068] Example 4: Preparation of methacrylamide porcine small intestinal submucosal (SISMA) hydrogel microneedles

[0069] The preparation method in this embodiment is the same as in Example 1, except that the method for preparing SISMA hydrogel microneedles using a microneedle mold is different, specifically:

[0070] (1) Inject SISMA mixed with photoinitiator LAP into a silicone resin (PDMS) microneedle mold. The concentration of LAP in the mixture is 0.25% (g / ml). Defoaming is performed under negative pressure, and the mixture is concentrated in an oven at 50°C for 24 hours.

[0071] (2) The concentrated SISMA (LAP-405nm, irradiated for 1 min) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0072] Example 5: Preparation of methacrylamide porcine small intestinal submucosal (SISMA) hydrogel microneedles

[0073] The preparation method in this embodiment is the same as in Example 1, except that the method for preparing SISMA hydrogel microneedles using a microneedle mold is different, specifically:

[0074] (1) SISMA mixed with photoinitiator LAP was injected into a silicone resin (PDMS) microneedle mold. The concentration of LAP in the mixture was 0.25% (g / ml). The mixture was defoamed under negative pressure and concentrated in an oven at 30°C for 48 hours.

[0075] (2) The concentrated SISMA (LAP-405nm, irradiated for 30s) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0076] (3) Scanning electron microscopy to examine the surface morphology of microneedles

[0077] SEM analysis of the microneedle surface morphology, such as Figure 1 , Figure 2 .

[0078] Example 6: Preparation of methacrylamide porcine small intestinal submucosal (SISMA) hydrogel microneedles

[0079] The preparation method in this embodiment is the same as in Example 1, except that the method for preparing SISMA hydrogel microneedles using a microneedle mold is different, specifically:

[0080] (1) SISMA mixed with photoinitiator I2959 was injected into a silicone resin (PDMS) microneedle mold. The concentration of LAP in the mixture was 0.25% (g / ml). The mixture was defoamed under negative pressure and concentrated in an oven at 20°C for 72 hours.

[0081] (2) The concentrated SISMA (I2959-365 nm, irradiated for 25s) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0082] Example 7: Preparation of methacrylamide porcine submucosal (SISMA) hydrogel microneedles

[0083] The preparation method in this embodiment is the same as in Example 1, except that the method for preparing SISMA hydrogel microneedles using a microneedle mold is different, specifically:

[0084] (1) SISMA mixed with photoinitiator I2959 was injected into a silicone resin (PDMS) microneedle mold. The concentration of LAP in the mixture was 0.25% (g / ml). The mixture was defoamed under negative pressure and concentrated in an oven at 50°C for 24 hours.

[0085] (2) The concentrated SISMA (I2959-365 nm, irradiated for 1 min) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0086] Example 8: Preparation of methacrylamide porcine small intestinal submucosa (SISMA) hydrogel microneedles

[0087] The preparation method in this embodiment is the same as in Example 1, except that the method for preparing SISMA hydrogel microneedles using a microneedle mold is different, specifically:

[0088] (1) Inject SISMA mixed with photoinitiator LAP or I2959 into a silicone resin (PDMS) microneedle mold. The concentration of LAP or I2959 in the mixture is 0.25% (g / ml). Defoaming is performed under negative pressure, and the mixture is concentrated in an oven at 30°C for 48 hours.

[0089] (2) The concentrated SISMA (I2959-365 nm, irradiated for 30s) in the mold was photocured and demolded to obtain SISMA hydrogel microneedles.

[0090] The following experimental examples demonstrate the beneficial effects of the hydrogel microneedles prepared in this invention.

[0091] Experimental Example 1: Performance testing of the SISMA hydrogel microneedles of the present invention:

[0092] 1. Experimental Methods

[0093] (1) Sample preparation

[0094] Referring to Example 5, SISMA hydrogel microneedles and GelMA hydrogel microneedles were prepared. The preparation method of GelMA hydrogel microneedles was the same as that of SISMA hydrogel microneedles, except that SISMA was replaced with GelMA in the raw materials. The preparation method of GelMA was as follows: gelatin was dissolved in a warm water bath at a ratio of 0.5% for 1-4 hours. After complete dissolution, free amino groups and methacrylic anhydride (MA) were added at a molar ratio of 1:20, the pH was adjusted to 9-10, and the reaction was carried out in an ice-water bath for 24 hours. The mixture was then dialyzed and lyophilized for later use. SISMA hydrogel microneedles and GelMA hydrogel microneedle extracts were prepared according to the national standard GB / T 16886.12-2017.

[0095] (1) Determination of the angiogenesis-promoting ability of SISMA hydrogel microneedles

[0096] To verify whether SISMA hydrogel microneedles have bioactivity, we verified the angiogenesis-promoting ability of SISMA hydrogel microneedles and compared it with GelMA hydrogel microneedles.

[0097] The following steps were followed for testing: 1) Matrigel deposition: A 24-well plate was placed on an ice plate, and 30 μL of Matrigel was added to each well, ensuring the bottom of the well was evenly covered. After equilibration on the ice plate for 5 min, the plate was incubated at 37°C for 15 min to form a gel. 2) Cell seeding and treatment: Human umbilical vein endothelial cells (HUVECs) in the logarithmic growth phase were digested and resuspended in SISMA hydrogel microneedles and GelMA hydrogel microneedle extract to prepare a cell extract. HUVEC cells were also resuspended in complete culture medium as a control group. 100,000 cells were added to each well of the Matrigel-deposited 24-well plate and incubated at 37°C for 8 h. 3) Observation: After 8 h of culture, the cells were observed under an optical microscope and images were acquired. ImageJ software was used for image analysis.

[0098] (2) SISMA hydrogel microneedle immunomodulation assay

[0099] The anti-inflammatory properties of SISMA hydrogel microneedles were verified using mouse mononuclear macrophage leukemia cells (RAW264.7) and compared with those of GelMA hydrogel microneedles.

[0100] RAW264.7 medium: 90% α-MEM medium + 10% fetal bovine serum (FBS) + 100 U / mL penicillin (P) and 100 μg / mL streptomycin (S).

[0101] Log-phase RAW264.7 cells were used to prepare cell suspensions, and 100,000 cells / well were inoculated into 6-well plates. After overnight adhesion, the cells were treated with 100 ng / mL lipopolysaccharide (LPS) for 4 hours to simulate an inflammatory microenvironment. Subsequently, SISMA hydrogel microneedles and GelMA hydrogel microneedles extracts were added, and a control group without LPS stimulation was set up. After 48 hours of culture, RNA was extracted using Promega's RNA kit (LS1040) and reverse transcribed using Promega's reverse transcription kit (A2791). Finally, qPCR experiments were performed using a two-step method with TaKaRa's TB Green Premix Ex Taq∥ reagent (RR820A) to determine the expression of inflammatory genes Il-6, Cd86, and Il-1β.

[0102] (3) Assay of the ability of SISMA hydrogel to promote fibroblast migration

[0103] The ability of SISMA hydrogel microneedles to promote fibroblast migration was measured and compared with that of GelMA hydrogel microneedles.

[0104] Mouse embryonic fibroblasts (NIH-3T3) were used as a cell model. The culture medium consisted of 90% high-glucose DMEM, 10% FBS, 100 U / mL P, and 100 μg / mL S. Logarithmic-phase NIH-3T3 cells were prepared into an FBS-free cell suspension. 200 μL of the FBS-free cell suspension was seeded into the upper chamber of a Transwell microarray at 50,000 cells / well. 700 μL of SISMA hydrogel microneedle and GelMA hydrogel microneedle extract was added to the lower chamber. After 24 h of culture, the culture medium was removed, and the cells were washed three times with PBS. The cells were fixed with 10% neutral formaldehyde for 30 min, washed three times with PBS, and stained with crystal violet for 1 h. The crystal violet was washed away with PBS, and the cells in the upper chamber were gently wiped with a cotton swab. Subsequently, the cells were observed and images were acquired using an optical microscope.

[0105] (4) Assay on the ability of SISMA hydrogel microneedles to promote fibroblast proliferation

[0106] The ability of SISMA hydrogel microneedles to promote fibroblast proliferation was measured and compared with that of GelMA hydrogel microneedles.

[0107] NIH-3T3 was used as a cell model, with the same culture medium as above. The proliferation capacity of NIH-3T3 cells was verified by CCK-8 assay.

[0108] Log-phase NIH-3T3 cells were prepared into a cell suspension, and 3,000 NIH-3T3 cells were seeded into each well of a 96-well plate at three time points. After cell attachment, the SISMA hydrogel microneedle and GelMA hydrogel microneedle extraction medium were replaced at 2, 4, and 6 days. At 1, 3, and 7 days, the conditioned medium was removed, and 110 μL of high-glucose DMEM medium containing 10% CCK-8 was added to each well. The plates were incubated at 37°C in the dark for 1.5 h. Then, 100 μL of the incubation medium was added to a new 96-well plate, and the absorbance was measured at 450 nm using a microplate reader.

[0109] (5) Determination of swelling properties of SISMA hydrogel microneedles

[0110] The swelling properties of SISMA hydrogel microneedles were determined.

[0111] Take SISMA hydrogel microneedles and GelMA hydrogel microneedles, weigh and record the initial weight. After soaking thoroughly in 5 mL of PBS, wipe off excess water with filter paper, weigh and record the weight. Swelling rate calculation: Let M0 be the initial mass and M1 be the mass after thorough soaking, then the swelling rate formula is:

[0112]

[0113] 2. Experimental Results

[0114] Results of angiogenesis capacity assay as follows Figure 3 and Figure 4 As shown, SISMA hydrogel microneedles exhibit better angiogenesis ability compared to the control group and the GelMA hydrogel microneedle group.

[0115] Immunomodulatory assay results as follows Figure 5 As shown, compared with the LPS and GelMA hydrogel microneedle groups, SISMA hydrogel microneedles were able to downregulate the expression of inflammatory factors Il-6 and CD86. Compared with the LPS group, SISMA microneedles were able to reduce the expression of inflammatory factor Il-1β.

[0116] Results of fibroblast migration ability assay as follows Figure 6As shown, compared with GelMA hydrogel microneedles, SISMA hydrogel microneedles have a stronger ability to promote NIH-3T3 migration.

[0117] Results of fibroblast proliferation assay as follows Figure 7 As shown, on day 5, compared with the control group, the OD value of SISMA hydrogel microneedles was significantly larger (P<0.05), and compared with GelMA hydrogel microneedles, the OD value of SISMA hydrogel microneedles was also significantly larger. The increase in OD value over time for SISMA hydrogel microneedles indicates that SISMA hydrogel microneedles themselves have the ability to promote NIH-3T3 proliferation.

[0118] The results of the swelling property determination experiment are as follows: Figure 6 As shown, compared with GelMA hydrogel microneedles, SISMA hydrogel microneedles absorb PBS liquid faster and have a higher swelling rate in the same time.

[0119] The experimental data above show that the SISMA hydrogel microneedles provided by this invention have excellent angiogenesis-promoting ability, immunomodulatory function, ability to promote fibroblast migration and proliferation, and good swelling properties.

[0120] Example 2: Optimal ratio of submucosal layer of small intestine to methacrylic anhydride

[0121] (1) Experimental methods

[0122] The grafting rate of SISMA is closely related to the mechanical properties of the hydrogel. In this experimental example, the grafting rate was used as an indicator to optimize the ratio of small intestinal submucosa to methacrylic anhydride during the preparation of SISMA.

[0123] The preparation method of SISMA in this experiment is the same as that in Example 1, except that the molar ratio of free amino groups to MA is set to 1:5, 1:20, and 1:100, respectively. The grafting rate of the prepared SISMA is determined by the o-phthalaldehyde (OPA) method.

[0124] (2) Experimental Results

[0125] The results are as follows Figure 9 As shown, when the molar ratio of free amino to MA is 1:20, the prepared SISMA has the highest grafting rate, with an average of 70%-90%.

[0126] In summary, the hydrogel microneedles made from modified decellularized matrix material (SISMA) provided by this invention have good bioactivity and better swelling properties, and have promising prospects in biomedicine.

Claims

1. A bioactive decellularized matrix hydrogel microneedle, characterized in that: It is prepared by the following method: (1) Mix methacrylamide porcine small intestinal submucosa material SISMA with a photoinitiator to obtain a mixture, inject the mixture into a microneedle mold, remove bubbles, and concentrate in an oven; (2) The concentrated SISMA in the mold was photocured and demolded to obtain SISMA hydrogel microneedles; The methacrylamide porcine small intestinal submucosa material SISMA is prepared by reacting small intestinal submucosa with methacrylic anhydride in an alkaline aqueous solution. The methacrylic anhydride grafting rate is 40-90%, and the small intestinal submucosa is a small intestinal submucosa that has undergone defatting, decellularization, and digestion treatment.

2. The hydrogel microneedles as described in claim 1, characterized in that, The methacrylic anhydride grafting rate is 80%.

3. The hydrogel microneedles as described in claim 1, characterized in that, The molar ratio of free amino groups to methacrylic anhydride in the submucosa of the small intestine is 1:(5~100).

4. The hydrogel microneedles as described in claim 3, characterized in that, The molar ratio of free amino groups to methacrylic anhydride in the submucosa of the small intestine is 1:

20.

5. The hydrogel microneedles as described in claim 1, characterized in that, The alkaline aqueous solution is an aqueous solution with a pH of 8 to 10.

6. The hydrogel microneedles as described in claim 1, characterized in that, The reaction conditions are: 0~50℃ for 1~24h.

7. The hydrogel microneedles as described in claim 6, characterized in that, The reaction conditions are selected from one of the following: 0~20℃ for 20~24 hours, 20~35℃ for 10~20 hours, or 35~50℃ for 1~10 hours.

8. The hydrogel microneedles as described in claim 1, characterized in that, The amino content of the submucosa of the small intestine is 400~800 μM.

9. The hydrogel microneedles according to claim 1, characterized in that: The photoinitiator has a mass-volume fraction of 0.05%-1% in the mixture.

10. The hydrogel microneedles according to claim 9, characterized in that: The photoinitiator is LAP or I2959.

11. The hydrogel microneedles according to claim 9, characterized in that: The oven concentration temperature is 20℃~50℃; the oven concentration time is 12h~72h; and the photocuring time is 5s~1min.

12. The method for preparing the hydrogel microneedles according to any one of claims 1-11, characterized in that, Includes the following steps: (1) The methacrylamide porcine small intestinal submucosa material SISMA was mixed with a photoinitiator to obtain a mixture. The mixture was injected into a microneedle mold, defoamed, and concentrated in an oven. The photoinitiator in the mixture has a mass-volume fraction of 0.05%. (2) The concentrated SISMA in the mold is photocured and demolded to obtain SISMA hydrogel microneedles.

13. Use of the hydrogel microneedles according to any one of claims 1-11 in the preparation of drug delivery carriers.

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