Curcumin-loaded claspable composite microneedle and preparation method thereof

Abstract

The application belongs to the field of biomedical materials, targeted drug delivery technology and ulcerative colitis treatment, and discloses a curcumin-loaded claspable composite microneedle and a preparation method thereof. The composite microneedle comprises a curcumin-loaded gelatin hemisphere base and a curcumin-loaded chitosan-polyvinyl alcohol microneedle, and the microneedle is located on the circular end face of the hemisphere base. The curcumin in the base and the microneedle is loaded in the form of curcumin-cyclodextrin inclusion compound. The claspable composite microneedle is formed by designing a hemisphere base + microneedle array, and the hemisphere base + microneedle part of the composite microneedle is loaded with curcumin, which can relieve and treat intestinal wall and intestinal mucosa damage caused by ulcerative colitis, indirectly inhibit intestinal flora disorder and abnormal activation of the inflammatory immune system, so as to solve the technical problem of low cure rate of ulcerative colitis, and provide a more safe and efficient treatment strategy for targeted intestinal diseases.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of biomedical materials, targeted drug delivery technology and ulcerative colitis (UC) treatment, and more particularly relates to a compound microneedle loaded with curcumin and capable of being clamped and a preparation method thereof. BACKGROUND

[0002] Currently, the existing treatment scheme for ulcerative colitis in clinical practice mainly involves internal medicine drug treatment, and various drugs such as amino salicylic acid preparations (e.g., mesalazine, sulfasalazine), glucocorticoids (e.g., prednisone, budesonide), immunosuppressants (e.g., azathioprine, cyclosporine), biological agents (e.g., infliximab, vedolizumab) can be selected according to the severity of the disease; in addition, fecal flora transplantation and other means can also achieve therapeutic effects. However, the existing treatment methods face many challenges: most of the conventional drugs have systemic adverse reactions, such as long-term use of steroid hormones leading to osteoporosis and increased risk of infection (drug side effects); some drugs (e.g., immunosuppressants, biological agents, etc.) are expensive, and combined with the chronic and recurrent nature of ulcerative colitis, long-term medication is required, further increasing the economic burden on patients (heavy social and economic burden); fecal flora transplantation can regulate intestinal flora balance and relieve ulceration, but the efficacy and durability are limited, and there are problems such as "donor-recipient pairing intolerance". For the problems of systemic administration, the clinic currently uses suppositories and retention enemas for intestinal local administration to make up for the deficiencies, but it is particularly worth noting that the lesion is characterized by "continuous distribution from the rectum upwards", which poses three bottlenecks for traditional local administration: ① enema and suppository are difficult to cover the deep lesions of the right half of the colon; ② the short drug retention time caused by intestinal fluid flushing; ③ diagnosis and treatment are separated, and separate operation is required for drug administration, increasing the discomfort and inconvenience of patients (low drug target efficiency, complicated operation). Therefore, it is urgent to explore a new treatment method and corresponding preparation to achieve targeted drug delivery, integrated diagnosis and treatment, and cost reduction and efficiency improvement.

[0003] Curcumin, as a "food and medicine" and GRAS (Generally Recognized as Safe) substance, contains natural polyphenolic compounds, which are widely available and easy to obtain. Curcumin can inhibit the levels of inflammatory factors such as IL-6 and TNF-α through multiple signaling pathways such as NF-κB and JAK / STAT, can regulate the immune of target organs, and has obvious antioxidant effect, and has certain ability to relieve ulcerative colitis, and has small side effects and adverse reactions. However, the core defect of its extremely low oral bioavailability (affected by the first-pass effect and "low drug concentration-target tissue response concentration") has long restricted its clinical application. In order to overcome the defects of oral administration of curcumin, researchers at home and abroad have explored various new drug delivery strategies. Microneedle technology, as a popular minimally invasive local drug delivery method, can effectively bypass the first-pass effect, improve local drug concentration, and reduce systemic exposure and side effects by creating micron-sized channels to deliver drugs directly to subcutaneous or mucosal tissues. However, traditional microneedle technology is mainly used for transdermal drug delivery, and its application range is relatively limited. In addition, there is a lack of a drug delivery device in the prior art that can be used in conjunction with standard endoscopic procedures (such as colonoscopy), can be reliably held by endoscopic clamps, can be accurately delivered and implanted into specific lesion sites in the intestinal tract, and can then be firmly adhered to the lesion site and controlled drug release. Therefore, the development of a new type of microneedle system suitable for intraluminal drug delivery is of great significance for improving the therapeutic effect of curcumin and other drugs for UC and promoting the development of diagnosis and treatment integration. SUMMARY

[0004] In view of the above defects or improvement needs of the prior art, the purpose of the present application is to provide a curcumin-loaded clippable composite microneedle and a preparation method thereof, wherein a clippable composite microneedle is formed by designing a hemispherical base + microneedle array, and the hemispherical base + microneedle part of the composite microneedle is loaded with curcumin, which can relieve and treat intestinal wall and intestinal mucosa damage caused by ulcerative colitis, indirectly inhibit intestinal flora disorder and abnormal activation of the inflammatory immune system, in order to solve the technical problem of low cure rate of ulcerative colitis, and provide a more safe and efficient targeted treatment strategy for intestinal diseases.

[0005] To achieve the above-mentioned purpose, according to one aspect of the present application, a curcumin-loaded clippable composite microneedle is provided, characterized in that it comprises a gelatin hemispherical base loaded with curcumin and a chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin, the microneedle being located on the circular end face of the hemispherical base, the height of the microneedle being 1.90-2.00 mm, and the height direction of the microneedle being perpendicular to the circular end face.

[0006] The curcumin in the base and the microneedle is loaded in the form of curcumin-cyclodextrin inclusion complex.

[0007] As a further preferred embodiment of the present application, the gelatin hemispheroid substrate loaded with curcumin is prepared by mixing gelatin powder with deionized water, hot injection molding after adding curcumin-cyclodextrin inclusion compound, and obtaining the hemispheroid after cooling and drying.

[0008] The curcumin-loaded chitosan (CS) -polyvinyl alcohol (PVA) microneedle is prepared by mixing the CS solution and the PVA solution, adding the curcumin-cyclodextrin inclusion compound to configure a mixed injection solution, repeatedly injecting into a microneedle mold, and obtaining the microneedle part after drying and demolding.

[0009] As a further preferred embodiment of the present application, the gelatin hemispheroid substrate loaded with curcumin and the curcumin-loaded chitosan (CS) -polyvinyl alcohol (PVA) microneedle are bonded by hot-melt gelatin and then cooled and solidified to form a composite microneedle.

[0010] As a further preferred embodiment of the present application, the curcumin-cyclodextrin inclusion compound is prepared by mixing a β-cyclodextrin solution and a curcumin solution under heating, and then cooling, precipitating, suction filtering, and drying; wherein the β-cyclodextrin solution is obtained by dissolving β-cyclodextrin in deionized water; and the curcumin solution is obtained by dissolving curcumin powder in ethanol.

[0011] As a further preferred embodiment of the present application, the microneedle is a quadrangular pyramid with a bottom side length of 1.0 mm-1.4 mm.

[0012] As a further preferred embodiment of the present application, the hemispheroid substrate has a spherical diameter of 10 mm-14 mm.

[0013] According to another aspect of the present application, the present application provides a preparation method of the above-mentioned curcumin-loaded claspable composite microneedle, characterized in that it comprises the following steps:

[0014] S1. Synthesis of the gelatin hemispheroid substrate loaded with curcumin: mixing gelatin powder with deionized water, hot injection molding after adding curcumin-cyclodextrin inclusion compound, and obtaining the gelatin hemispheroid substrate loaded with curcumin after cooling and drying;

[0015] S2. Synthesis of the curcumin-loaded chitosan (CS) -polyvinyl alcohol (PVA) microneedle: mixing the CS solution and the PVA solution, adding the curcumin-cyclodextrin inclusion compound to configure a mixed injection solution, repeatedly injecting into a microneedle mold, and obtaining the curcumin-loaded chitosan (CS) -polyvinyl alcohol (PVA) microneedle after drying and demolding;

[0016] S3. Synthesis of the composite microneedle: the curcumin-loaded gelatin hemisphere base is bonded with the curcumin-loaded chitosan (CS)-polyvinyl alcohol (PVA) microneedle through hot-melt gelatin, and then solidified after cooling to form the composite microneedle.

[0017] According to still another aspect of the present application, the present application provides use of the above-mentioned curcumin-loaded grippable composite microneedle in the preparation of a drug delivery platform for relieving ulcerative colitis.

[0018] As a further preferred embodiment of the present application, the drug delivery platform is a sustained-release implant-type drug delivery platform.

[0019] As a further preferred embodiment of the present application, the drug delivery platform can reduce the risk of intestinal obstruction.

[0020] By the above technical solutions conceived by the present application, compared with the prior art, the present application designs a hemisphere base + microneedle array to form a grippable composite microneedle, which can be gripped to the lesion site by a colonoscope with gripping forceps and fixed, and a curcumin-loaded grippable composite microneedle is constructed by using hot-melt gelatin, beta-cyclodextrin-curcumin powder (i.e., curcumin-cyclodextrin inclusion compound), and CS-PVA composite material to realize colon administration, which can be used for ulcerative colitis, can be fixed to the intestinal lesion site by a colonoscope, realizes local sustained release of curcumin, and can be used for relieving ulcerative colitis. The composite microneedle can stably adhere in the dynamic peristalsis and liquid environment of the intestinal tract, while reducing the risk of intestinal obstruction.

[0021] In the design of the height of the microneedle, the present application takes into account that the grippable composite microneedle needs to be stably implanted into the intestinal wall and not to fall off in a long period of time, improves the design of the conventional microneedle height <=1 mm, and adopts a longer microneedle design; at the same time, in order to avoid damage to the intestinal wall by the microneedle which is too long, the final microneedle height is designed to be 1.90-2.00 mm (e.g., 1.95 mm). The implantable depth of the microneedle can cover the intestinal mucosa layer (0.8-1.2 mm) to the submucosa layer (0.4-0.7 mm), avoiding damage to the intestinal tissue by being too deep. Moreover, the base of the composite microneedle is designed to be a hemisphere shape, the main body of the grippable hemisphere part is composed of gelatin material, which can maintain a solid state in vitro and be slowly degraded in vivo, and this design makes the curcumin-loaded grippable composite microneedle have the characteristics of being grippable and degradable. In addition, the smooth appearance of the hemisphere-shaped base will not scratch the intestinal tract.

[0022] The obtained curcumin-loaded claspable composite microneedle can be targeted for relieving and treating ulcerative colitis, the curcumin is uniformly wrapped in the interspace of the porous gelatin material and the CS-PVA composite material, and is attached to the surface of a colonic ulcer focus through the microneedle structure, so that the colonic ulcer focus can be protected, the abnormal activation of an immune inflammatory response can be inhibited, and the intestinal flora can be improved, thereby delaying the further development of ulcerative colitis. The present application can bypass the first-pass effect of oral administration, improve the local drug concentration of the lesion, relieve the symptoms of ulcerative colitis through the mechanisms of anti-inflammatory, antioxidant and regulation of intestinal flora, improve the mucosal healing rate, reduce the disease activity index, and provide an efficient auxiliary diagnosis and treatment integrated strategy for the precise treatment of ulcerative colitis.

[0023] The present application forms a claspable composite microneedle by designing a hemispherical base + microneedle array, the hemispherical base is the upper half of the curcumin-loaded claspable composite microneedle, which is tough and homogeneous, and functions to increase the drug loading capacity of the curcumin-loaded claspable composite microneedle; the microneedle is the lower half of the curcumin-loaded claspable composite microneedle, which has hardness and toughness, and functions to increase the intestinal wall adhesion of the curcumin-loaded claspable composite microneedle. In subsequent use, the hemispherical base part of the curcumin-loaded claspable composite microneedle can be fixed with a medical clamping forceps, slowly sent into the colon cavity from the rectum under the assistance of a colonoscope, and placed at the ulcer focus. Based on the composite microneedle obtained by the present application, the drug administration scene can be, for example, mild to moderate active ulcerative colitis (MAYO endoscopic score 1-2 points, UCEIS score 1-6 points), and the drug administration mode can be: after the lesion position is determined by colonoscopy, the claspable composite microneedle preparation is clamped with a colonoscope with a clamping forceps, and is accurately sent to the intestinal ulcer and fixed by the clamping arm, so that the microneedle part penetrates into the submucosa, and the gelatin hemispherical base is attached to the surface of the lesion. The curcumin-loaded claspable composite microneedle can be used in cooperation with colonoscopy, and the implantation and administration of the composite microneedle preparation are completed immediately after the lesion needs clinical intervention is determined by colonoscopy, without the need for additional surgical operation, reducing the pain and treatment cycle of the patient; and since the target of curcumin drug action is clear, the curcumin-loaded claspable composite microneedle of the present application can release curcumin, thereby persistently playing the role of inhibiting macrophage activation, regulating neutrophil infiltration, and regulating Treg cell function, thereby reducing intestinal inflammatory response and improving intestinal mucosal barrier function, avoiding the first-pass effect of oral administration and the damage of the intestinal environment to the drug.

[0024] The present application can uniformly wrap the beta-cyclodextrin-curcumin powder in the pores of the porous material to form a slow-release system by using gelatin material and CS-PVA composite material with natural biocompatibility and biodegradability; the curcumin-loaded claspable composite microneedle is fixed on the surface of the ulcer focus of the intestinal wall through the microneedle structure, which maximizes the possibility of contact between the beta-cyclodextrin-curcumin powder and the necrotic tissue, and the beta-cyclodextrin-curcumin powder wrapped therein is slowly released as the gelatin material and the CS-PVA composite material decompose, which not only targets the colon for slow drug release and persistent drug efficacy, but also forms a film on the surface of the ulcer focus, which isolates the ulcer focus from harmful substances in the intestinal cavity and delays the further development of ulcerative colitis; the curcumin-loaded claspable composite microneedle has pharmacological and physical therapeutic effects, has good prospects for treating ulcerative colitis and improving intestinal function, and has the advantages of high targeting and high safety.

[0025] Meanwhile, the main materials used in the present application are only gelatin powder, curcumin powder, PVA powder, CS powder, beta-cyclodextrin powder, etc., and the raw material cost is low; the preparation process is only simple operations such as mixing, drying, hot infusion, cooling and precipitation, and the preparation cost is low. This to some extent makes up for the shortcomings of some clinically common expensive drugs.

[0026] The present application first uses beta-cyclodextrin to embed curcumin to obtain beta-cyclodextrin-curcumin powder (i.e., curcumin-cyclodextrin inclusion compound), which has high biocompatibility, biodegradability, low toxicity and other characteristics, and improves the problems of low water solubility and low biocompatibility of curcumin powder; in combination with gelatin material and CS-PVA composite material (which have good biodegradability, tissue compatibility and porous structure), conditions are provided for slow release of the embedded curcumin drug. The curcumin-loaded claspable composite microneedle prepared in the present application has a strong targeted ulcerative colitis ulcer focus drug slow-release delivery capacity, and the beta-cyclodextrin-curcumin powder wrapped therein is slowly released as the gelatin material and the CS-PVA composite material decompose and absorb, which lasts for a long time and realizes precise and efficient treatment of ulcerative colitis.

[0027] Taking the following embodiments as examples, the composite microneedles obtained based on the present invention consist of gelatin hemispheres and chitosan (CS)-polyvinyl alcohol (PVA) microneedles, and are loaded with β-cyclodextrin-curcumin powder (curcumin loading reaches 86.5 mg / composite microneedle). In the following embodiments, the mass ratio of β-cyclodextrin powder to curcumin powder is 3:1 to satisfy the molecular weight ratio between the two is 1:1, so as to obtain 1:1 embedded β-cyclodextrin-curcumin powder; the mass ratio of CS to PVA is 3:10 to ensure that the microneedles have mechanical strength and are not easily broken, so that the product can be fixed on the intestinal surface and not easily fall off (these ratios are all obtained after a large number of experiments). Taking the following embodiments as examples, the mechanical properties of the composite microneedle formulation obtained based on the present invention meet the following requirements: maximum tensile force ≈ 0.4 N, theoretical propulsion force ≈ 2.17 × 10 -6 N (calculated as in the examples below) means that the maximum tensile force is greater than the theoretical propulsion force; during simulated colonoscopy drug delivery, it can be stably embedded in the intestinal tissue surface with a low risk of detachment or displacement.

[0028] Specifically, the present invention can achieve the following beneficial effects:

[0029] (1) The curcumin-loaded clampable composite microneedles obtained in this invention can be clamped by clamping forceps and placed at the ulcer site with the assistance of colonoscopy. They can be fixed at the ulcer site under the action of colonic peristalsis and gravity, and gradually soften under the action of body temperature and colonic fluid, adhering to the surface of the ulcer site, providing a physical barrier for the ulcer site area, preventing the ulcer site from contacting harmful substances in the intestinal lumen, and providing a favorable environment for the healing of the ulcer site.

[0030] (2) The porous CS-PVA composite material and gelatin material encapsulate β-cyclodextrin-curcumin powder in their pores and adhere it to the surface of ulcerative colitis lesions. On the one hand, it can increase the adhesion of β-cyclodextrin-curcumin powder to the tissue surface, so that it can play a long-lasting and uniform role. On the other hand, the clampable composite microneedles loaded with curcumin prepared by the present invention have a strong ability to deliver sustained-release drugs to ulcerative colitis lesions. As the CS-PVA composite material and gelatin material decompose and absorb, the β-cyclodextrin-curcumin powder encapsulated therein is slowly released, which effectively prolongs the circulation and retention time of β-cyclodextrin-curcumin powder at the ulcer site, and the drug effect is more lasting.

[0031] This invention not only provides a treatment strategy for ulcerative colitis with higher therapeutic efficacy and lower toxicity, but also provides a drug sustained-release delivery platform based on CS-PVA composite materials and gelatin materials. Attached Figure Description

[0032] Figure 1This is a photograph of the mold used in Example 1 to prepare the clampable composite microneedles loaded with curcumin.

[0033] Figure 2 This is a mold drawing for the microneedle portion of the clampable composite microneedle loaded with curcumin, prepared in Example 1 (the unit of the dimensions marked in the figure is mm).

[0034] Figure 3 This is a photograph of the mold used in Example 1 to prepare the hemispherical part of the clampable composite microneedle loaded with curcumin.

[0035] Figure 4 This is a flowchart illustrating the preparation process of the clampable composite microneedles loaded with curcumin in this invention.

[0036] Figure 5 This is a flowchart of the method for applying curcumin-loaded clampable composite microneedles to ulcerative colitis in this invention.

[0037] Figure 6 These are magnified images of the curcumin-loaded clampable composite microneedles prepared in Example 1, and a macro lens image of the tip structure of the curcumin-loaded clampable composite microneedles; wherein, Figure 6 (a) in the image corresponds to an enlarged photograph of the actual object. Figure 6 (b) in the diagram corresponds to the needle tip structure under a macro lens.

[0038] Figure 7 This describes the dissolution of the clampable composite microneedles loaded with curcumin prepared in Example 2.

[0039] Figure 8 This is the dissolution curve of the clampable composite microneedles loaded with curcumin prepared in Example 3.

[0040] Figure 9 This is a stability test of the clampable composite microneedles loaded with curcumin prepared in Example 4.

[0041] Figure 10 This is a further test of the stability of the clampable composite microneedles loaded with curcumin prepared in Example 4.

[0042] Figure 11 This is a section image obtained in Example 5, showing the perforation of intestinal wall tissue by a clampable composite microneedle loaded with curcumin under a high-power microscope.

[0043] Figure 12 This is a schematic diagram of the clampable composite microneedles loaded with curcumin prepared in Example 6, which are implanted into the intestinal lumen and fixed to the intestinal wall tissue. Detailed Implementation

[0044] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention. Furthermore, the technical features involved in the various embodiments of this invention described below can be combined with each other as long as they do not conflict with each other.

[0045] In summary, the preparation process of the curcumin-loaded clampable composite microneedles for treating ulcerative colitis in this invention includes the following steps:

[0046] Step 1: Preparation of β-cyclodextrin-curcumin inclusion complex powder (i.e., β-cyclodextrin-curcumin powder), including the following sub-steps:

[0047] Preparation of β-cyclodextrin solution: Prepare a 1.2% (w / v) β-cyclodextrin solution from β-cyclodextrin powder using deionized water. For example, take 6 g of β-cyclodextrin (98% purity) and dissolve it in 500 mL of deionized water at 60 °C.

[0048] Preparation of curcumin solution: Prepare a 0.4% (w / v) curcumin solution from curcumin powder using anhydrous ethanol. For example, take 2 g of curcumin powder (98% purity) and dissolve it in 500 mL of anhydrous ethanol.

[0049] Synthesis of the target β-cyclodextrin-curcumin inclusion complex powder: β-cyclodextrin solution and curcumin solution were mixed in a 1:1 ratio (volume ratio), stirred at 37 ℃ for 1 h, heating was stopped, and the mixture was placed in a refrigerator at 2-8 ℃ for 40 min to cool. The mixture was then filtered, and the precipitate was dried to obtain the β-cyclodextrin-curcumin inclusion complex powder. The powder was weighed and sealed for later use.

[0050] Step 2: Preparation of the microneedle portion, including the following sub-steps:

[0051] Preparation of chitosan solution: Prepare a 2% (w / v) chitosan solution from chitosan powder using a 1% acetic acid solution. For example, dissolve 1 mL of glacial acetic acid in 99 mL of water to obtain a 1% acetic acid solution, and then dissolve 0.6 g of chitosan powder (degree of deacetylation 80.0-95.0) in 30 mL of the above acetic acid solution.

[0052] Preparation of PVA solution: Prepare a 10% (w / v) PVA solution from PVA powder using deionized water. For example, 1 g of PVA powder can be dissolved in 10 mL of deionized water.

[0053] Synthesis of the target microneedle portion: Chitosan solution and PVA solution were mixed in a 3:1 volume ratio. Then, a corresponding proportion of β-cyclodextrin-curcumin inclusion complex powder was added to the mixture (i.e., 26 g of β-cyclodextrin-curcumin inclusion complex powder was added to every 100 mL of the mixture). The mixture was stirred at 37°C for 1 h to obtain a final solution. This final solution was then added to a PDMS microneedle mold (the mold's chamber for containing the final solution was a 7×7 array of square pyramids, where each pyramid had a height of 1.95 mm, a base length of 1.2 mm, and a spacing of 0.5 mm between pyramids). Figure 1 , Figure 2 As shown in the figure, the mold was degassed under vacuum and placed in a vacuum drying oven at 37 ℃ for 2 h. After 2 h, the mold was removed and the mixed solution was added to the mold (considering water evaporation). The mold was then placed in a vacuum drying oven again for 2 h, and the mixed solution was added to the mold again. This process of drying and adding the mixed solution was repeated 2 to 3 times. The mold was then dried at room temperature (23 ± 2 ℃) for more than 12 h. After thorough drying and solidification, the mold was demolded to obtain the microneedle portion.

[0054] Step 3: Preparation of the hemispherical portion: Gelatin powder (250 g / L) and β-cyclodextrin-curcumin inclusion complex powder are uniformly mixed at a mass ratio of 1325:163. The resulting mixture is then mixed with an equal volume of deionized water to obtain a solution. Finally, the solution is injected into a silicone mold (the mold is a hemispherical silicone mold, meaning the cavity in the mold used to contain the solution is hemispherical, and the diameter of the sphere is 14 mm, see...). Figure 3 The mold was heated in a 37°C water bath for 1 hour, cooled for 1 hour, and then demolded to obtain the hemispherical part.

[0055] After the above steps are completed, the microneedle part is bonded to the hemispherical part with molten gelatin, and after cooling, a clampable composite microneedle loaded with curcumin is obtained.

[0056] The following are specific embodiments (unless otherwise specified, all embodiments in the following text were carried out at room temperature of 23 ± 2 ℃):

[0057] Example 1:

[0058] like Figure 4 As shown, the preparation process of the curcumin-loaded clampable composite microneedles for the treatment of ulcerative colitis provided in this embodiment of the invention includes the following steps:

[0059] Step 1: Preparation of β-cyclodextrin-curcumin powder: Take 6 g of β-cyclodextrin (98% purity) and dissolve it in 500 mL of deionized water at 60 ℃ to obtain 500 mL of β-cyclodextrin solution; then take 2 g of curcumin powder (98% purity) and dissolve it in 500 mL of anhydrous ethanol to obtain 500 mL of curcumin solution. Pour the obtained β-cyclodextrin solution and curcumin solution into the same container and mix thoroughly. Stir at 37 ℃ for 1 h, then stop heating and place in a refrigerator at 2-8 ℃ to cool for 40 min. Filter, dry the precipitate, and obtain β-cyclodextrin-curcumin inclusion complex powder. Weigh and seal for later use.

[0060] Step 2: Preparation of the microneedle portion: Take 60 mL of 2% (w / v) chitosan solution and 20 mL of 10% (w / v) PVA solution, mix them thoroughly, and then add 20.8 g of β-cyclodextrin-curcumin inclusion complex powder to the mixture. Stir at 37℃ for 1 h to obtain a mixed solution. Add the mixed solution to a PDMS microneedle mold, remove bubbles under vacuum, and place in a vacuum drying oven at 37℃ for 2 h. After 2 h, remove the mold and add more mixed solution (considering water evaporation); dry at 37℃ again for 2 h, and repeat this process 3-4 times. Finally, place the mold in a dark environment and air dry at 23±2℃ for at least 12 h. After thorough air drying, demold to obtain the microneedle portion.

[0061] Step 3: Take 2.65 g of gelatin powder (250 g / L) into a hemispherical silicone mold, then mix in 326 mg of β-cyclodextrin-curcumin inclusion complex powder. Mix the two thoroughly in a small beaker, add an equal volume of deionized water, mix thoroughly, and pour the mixture into the silicone mold. Place the mold in a 37 ℃ water bath and heat for 1 h. After cooling for 1 h, demold to obtain the hemispherical part.

[0062] After the above steps are completed, the microneedle part is joined to the hemispherical part with molten gelatin (i.e., hot melt gelatin), and after cooling, a clampable composite microneedle loaded with curcumin is obtained.

[0063] In the clampable composite microneedles loaded with curcumin obtained in this embodiment, the β-cyclodextrin-curcumin powder loading in the hemisphere is approximately 326 mg, and the curcumin drug loading is approximately 81.5 mg; the β-cyclodextrin-curcumin powder loading in the microneedles is approximately 20 mg, and the curcumin drug loading is approximately 5 mg; in the entire clampable composite microneedles loaded with curcumin, the β-cyclodextrin-curcumin powder loading is approximately 346 mg, and the curcumin drug loading is approximately 86.5 mg.

[0064] In the curcumin-loaded clampable composite microneedles obtained in this embodiment, the bottom side length of the microneedle is 1.2 mm and the length of the tip is 1.95 mm. It can be clamped to the lesion and fixed by a colonoscope with clamping forceps. The implantation depth of the microneedle can cover the intestinal mucosa layer (0.8-1.2 mm) to the submucosa layer (0.4-0.7 mm), avoiding damage to intestinal tissue due to excessive depth.

[0065] Example 2:

[0066] This invention provides a method for determining the solubility of clampable composite microneedles loaded with curcumin, including:

[0067] (1) Prepare clampable composite microneedles loaded with curcumin according to the method provided in Example 1.

[0068] (2) Cut a 3 cm × 3 cm piece of fresh pig large intestine and place it in a petri dish. Add an appropriate amount of artificial colon fluid (commercially available, product number: PH1842, Feijing Biotechnology). The amount added should be such that the liquid level covers the pig large intestine. Fix the curcumin-loaded clampable composite microneedles prepared in (1) onto the above-mentioned pig large intestine-artificial colon fluid composite using clamping forceps.

[0069] (3) The porcine colon-artificial colon fluid complex with curcumin-loaded clampable composite microneedles fixed in (2) was placed in a flask and then fixed on the clamp of a water bath constant temperature shaker tray. The parameters were set as follows: shaker speed 100 rpm; shaking amplitude 30 mm; motion mode reciprocating; temperature 37.5 ℃. The above parameter settings are all based on simulating the normal physiological characteristics of the human colon.

[0070] (4) The dissolution of the curcumin-loaded clampable composite microneedles was observed at 0.5 h, 1 h, 1.5 h, 2 h, 4 h, 6 h, 8 h, 10 h, and 12 h. The dissolution at 0 h, 4 h, 8 h, and 12 h is shown below. Figure 7 As shown, the clampable composite microneedles loaded with curcumin began to soften after 4 hours, no longer exhibiting a spherical shape. Between 8 and 12 hours, the curcumin-loaded clampable composite microneedles gradually softened and adhered to the surface of the pig large intestine in a layered, muddy manner. Given that the gastric emptying time for solid food in humans is 4-5 hours, the risk of intestinal obstruction is relatively low even if the patient eats immediately after the insertion of the curcumin-loaded clampable composite microneedles.

[0071] Example 3:

[0072] This invention further provides a sustained-release assay of clampable composite microneedles loaded with curcumin, including:

[0073] (1) Five clampable composite microneedles loaded with curcumin were prepared according to the method provided in Example 1. The weight of each clampable composite microneedle loaded with curcumin was weighed and recorded.

[0074] (2) Each clampable composite microneedle loaded with curcumin was operated according to (2) to (3) in Example 2.

[0075] (3) For any composite microneedle, at 0.5 h, 1 h, 1.5 h, 2 h, 4 h, 6 h, 8 h, 10 h and 12 h respectively, the incompletely dissolved curcumin-loaded clampable composite microneedle was picked up with tweezers and the timing was paused. The surface moisture was absorbed with absorbent paper and then weighed. The weight of the incompletely dissolved curcumin-loaded clampable composite microneedle at each time was recorded (after weighing, the composite microneedle was put back into the shaker to continue the experiment, and the timing was continued at the same time), and the mass change rate at each time was calculated.

[0076] (4) Using Origin plotting software, the SWeibull1 model was used to fit the dissolution curve of the clampable composite microneedles loaded with curcumin, where the dissolution rate was approximated by the rate of mass change. The resulting curve is shown in Figure 1. Figure 8 As shown, R 2 The value was 0.99747, indicating that the model could fit the dissolution curve of the curcumin-loaded clampable composite microneedles well, and further demonstrating that the curcumin-loaded clampable composite microneedles conform to conventional drug dissolution kinetics. In five repeated experiments, the average dissolution rate over 12 hours was 86.2%. Figure 8 As shown, the composite microneedles exhibit specific dissolution and degradation properties in a simulated in vivo intestinal environment (artificial colonic fluid): they are hard at 0 h, begin to soften after 4 h, and remain in an adhesive state from 8 to 12 h. The dissolution trend of curcumin is "fast at first and then slow": 17.15% at 0.5 h, 51.11% at 2 h, 66.80% at 4 h, and 86.20% at 12 h. This indicates that the clampable composite microneedles loaded with curcumin can stably release the loaded curcumin within 12 h, exhibiting good sustained-release properties, and can achieve slow release and long-lasting effect of curcumin at intestinal lesions.

[0077] Example 4:

[0078] This invention provides the stability of the binding of curcumin-loaded clampable composite microneedles to the colon, including:

[0079] (1) Prepare clampable composite microneedles loaded with curcumin according to the method provided in Example 1.

[0080] (2) Cut a 3 cm × 3 cm piece of fresh pig large intestine and place it in a petri dish. Add an appropriate amount of artificial colon fluid, with the amount added enough to cover the pig large intestine. Fix the curcumin-loaded clampable composite microneedles prepared in (1) onto the above-mentioned pig large intestine-artificial colon fluid composite using clamping forceps.

[0081] (3) Use tweezers to pick up the pig large intestine and shake it left and right with appropriate force. Observe whether the clampable composite microneedles loaded with curcumin fall off. The results are as follows: Figure 9 As shown, even when the pig large intestine is vibrated left and right with force, the clampable composite microneedles loaded with curcumin remain stably attached to the pig large intestine.

[0082] (4) Place the pig large intestine with the curcumin-loaded clampable composite microneedles fixed on the experimental table. Use a thin, circular thread to connect the hemisphere and the microneedle array. Pull the other end of the thread with a spring balance, gradually increasing the tension on the spring balance and observing the change in the spring balance reading. When the curcumin-loaded clampable composite microneedles loosen, record the spring balance reading. Figure 10 As shown, when the clampable composite microneedle loaded with curcumin loosens, the spring tension gauge reading is recorded as 0.4 N.

[0083] (5) Calculate the propulsion force of intestinal peristalsis on the composite microneedles that can be clamped by curcumin: The effect of intestinal peristalsis on an object is approximated as the effect of fluid on an object, and the resistance formula in fluid mechanics (as shown below) is used for estimation.

[0084]

[0085] Where F is the resistance generated by the fluid on the object (in this example, the propulsive force generated by intestinal peristalsis on the clampable composite microneedles loaded with curcumin is equal in magnitude but opposite in direction to this resistance). It is the drag coefficient, which is related to the shape of the object. The drag coefficient of the clampable composite microneedle loaded with curcumin is approximately equal to that of the hemisphere, which is 0.47 (this is an empirical value obtained through experimental and theoretical research). The density of colonic contents is approximately 1.2 g / cm³. 3 ; Let v be the velocity of the fluid relative to the object, specifically the velocity of the intestinal contents relative to the curcumin-loaded clampable composite microneedles. Since intestinal peristalsis is generally slow, we assume v = 0.01 m / s. Let A be the projected area of ​​the object perpendicular to the fluid flow direction. The projection of the curcumin-loaded clampable composite microneedles perpendicular to the fluid flow direction is a semicircle. Based on the formula for the area of ​​a circle, A is approximately 7.7 × 10⁻⁶ m / s. -5 m 2 Substituting the above data into the formula yields:

[0086] F = 2.17 × 10 -6 N

[0087] The propulsive force of intestinal peristalsis on the curcumin-loaded clampable composite microneedles is much less than the resistance (0.4 N). Therefore, the curcumin-loaded clampable composite microneedles have good stability in the colon and will not easily slip off.

[0088] Example 5:

[0089] This invention provides embodiments of the microscopic binding of curcumin-loaded clampable composite microneedles to the colon, including:

[0090] (1) Prepare clampable composite microneedles loaded with curcumin according to the method provided in Example 1.

[0091] (2) Cut a 3 cm × 3 cm piece of fresh pig large intestine and place it in a petri dish. Add an appropriate amount of artificial colon fluid, with the amount added enough to cover the pig large intestine. Fix the curcumin-loaded clampable composite microneedles prepared in (1) onto the above-mentioned pig large intestine-artificial colon fluid composite using clamping forceps.

[0092] (3) Remove the curcumin-loaded clampable composite microneedles with forceps, cut out the colon tissue bound to the microneedles, place it in a cryostat, and add a small amount of OCT embedding agent. After the tissue is completely frozen, section it. Then fix the tissue sections in 95% ethanol for 1-2 minutes, wash with water, and perform H&E staining. After staining, add neutral resin to the sections, cover with a coverslip, gently flatten, remove air bubbles, and observe under a microscope. Figure 11 As shown, although the tissue cavities created by the partially curcumin-loaded clampable composite microneedles did not reach the muscle layer due to factors such as the slicing angle, there were still complete tissue cavities that penetrated the mucosa and submucosa, which is consistent with the histological characteristics of ulcerative colitis that only involves the mucosa and submucosa.

[0093] Example 6:

[0094] This invention provides the in vitro feasibility of using curcumin-loaded clampable composite microneedles to target colonic ulcer lesions via colonoscopy, including:

[0095] (1) Prepare clampable composite microneedles loaded with curcumin according to the method provided in Example 1.

[0096] (2) Place a 1 m long fresh pig large intestine segment flat on the experimental table, and inflate one end with an air pump at an appropriate speed to keep the pig large intestine full. Use clamping forceps attached to the colonoscope to hold the clampable composite microneedle loaded with curcumin and slowly insert it into the pig large intestine at the other end. Observe the internal condition of the pig large intestine through the monitor connected to the colonoscope. Then place the clampable composite microneedle loaded with curcumin in the designated position and press it with the clamping forceps to make it firmly attached to the colon tissue.

[0097] (3) Slowly move the pig's large intestine left and right, and up and down to simulate the peristalsis of the large intestine in vivo, and observe whether the clampable composite microneedles loaded with curcumin are firmly attached to the pig's large intestine through colonoscopy. The results are as follows: Figure 12 As shown, even when simulating colonic peristalsis, the clampable composite microneedles loaded with curcumin remain firmly attached to the designated location in the colon.

[0098] Advantages of this invention:

[0099] To make the objectives, technical solutions, and advantages of this invention clearer, the following description, demonstration, and discussion are based on a comparison of this invention with other inventions using curcumin in the field of colon treatment in terms of targeting, clinical ease of use, safety, and medication cost.

[0100] (1) Targeting:

[0101] Currently, most common colon-targeted drug delivery systems are based on oral administration, including pH-controlled release systems, time-controlled release systems, and enzyme-triggered controlled release systems (microbial-triggered controlled release systems). The principles of these systems all relate to the specific dissolution of drug coatings in the colonic environment, releasing the drug. However, these oral-based targeted drug delivery systems are not precise: firstly, they can only achieve drug release near the colonic segment, not guaranteeing release near the colonic lesion; secondly, they cannot address the differences in colonic environments across species and populations, leading to a significant reduction in targeting accuracy.

[0102] For example, a Chinese patent (application number 201511027771.0) provides a curcumin colon-targeted drug formulation and its preparation method. This patent involves preparing an inclusion complex of curcumin and cyclodextrin in a specific ratio, combining them with various natural polysaccharide materials to form insoluble colon-targeted microparticles, and then coating them with an enteric coating solution to create the colon-targeted drug formulation. Although this patent emphasizes that its invention achieves a certain degree of colon-targeting effect, it does not conduct in vivo or in vitro animal experiments to verify this. It only demonstrates a high drug release rate in artificial colonic fluid and rat colonic contents release media through in vitro release tests, thus failing to fully prove that its invention has significant targeting properties.

[0103] This invention achieves targeted drug delivery by using a colonoscope to hold a clampable composite microneedle loaded with curcumin and fix it at the intestinal lesion site, thus achieving local sustained release of curcumin. This invention, therefore, utilizes a colonoscope to precisely locate the drug at the lesion site, avoiding the relatively low targeting accuracy and high uncertainty of oral-based targeted drug delivery systems. It also bypasses the first-pass effect of oral curcumin administration, increasing the local drug concentration at the lesion site, thereby alleviating the symptoms of ulcerative colitis through anti-inflammatory, antioxidant, and intestinal flora regulation mechanisms.

[0104] (2) Clinical usability:

[0105] While common oral-administered colon-targeted drug delivery systems offer portability, many patents are limited to conventional drug delivery approaches. Their core focus remains on improving the efficiency of oral administration, without considering the clinical diagnosis and treatment of colonic diseases such as ulcerative colitis. For example, Chinese patent application (application number: 202010775144.X) proposes a solubilization method and agent for curcumin drugs to achieve targeted drug delivery to colonic tissue, and its application in the preparation of drugs for treating colitis. This patent aims to provide a curcumin-containing agent that uses curcumin as the active molecule and forms a covalent complex with hydrophilic excipients to increase the curcumin loading and improve its oral bioavailability. The patent demonstrates the high drug release rate of its invention through in vitro release tests, but it does not reflect specific clinical usability: on the one hand, the patent does not clearly propose clinical application scenarios and specific usage methods; on the other hand, it only increases the efficiency of oral administration by increasing the drug loading of curcumin, without demonstrating a significant therapeutic advantage compared with other existing oral drugs. Therefore, it may not have an application advantage in clinical practice and has poor clinical usability.

[0106] This invention, considering the clinical context of ulcerative colitis and leveraging the need for colonoscopy in ulcerative colitis patients, proposes a method to implant curcumin-loaded, clampable composite microneedles into the colonic lumen using clamping forceps during the anesthesia interval of colonoscopy. This allows for precise treatment without increasing the patient's treatment burden, demonstrating innovation in both treatment method and approach. Furthermore, this invention is a non-oral medication, which can be used in combination with oral medications, making it highly clinically usable.

[0107] (3) Safety:

[0108] Currently, most common oral-administered colon-targeted drug delivery systems focus their safety concerns primarily on the safety of the drug's materials. To meet the basic requirements of being non-toxic, harmless, highly biocompatible, and biodegradable, oral medications often use substances approved by the U.S. Food and Drug Administration (FDA) as raw materials, such as cyclodextrin, alginate, pectin, and rice protein hydrolysate. These substances are mostly extracted from natural crops and are widely used in the manufacture of various clinical drugs and medical devices. However, many patents only describe safety in terms of the materials themselves, without designing experiments to verify the safety of the product, which does not meet the requirements for clinical application. This is mainly reflected in whether the product's shape, size, and degradation time meet the needs of human safety.

[0109] For example, Chinese patent application (application number: 202310823602.6) discloses a drug-loaded and stabilized curcumin composite nanoparticle and its preparation method. This invention uses rice protein hydrolysate as a carrier to load curcumin, and uses chondroitin sulfate to stabilize the rice protein hydrolysate loaded with curcumin, thus creating a colon-targeted drug formulation. The main materials used in this invention include rice protein hydrolysate, curcumin, and chondroitin sulfate, all of which have been certified by the U.S. Food and Drug Administration (FDA) and are recognized as non-toxic and harmless substances. However, this invention has not undergone in vitro or in vivo experiments to verify its safety. Therefore, the possibility of a series of safety issues arising after the composite nanoparticles are ingested by the human body, such as intestinal obstruction, intestinal irritability, and intestinal spasm, cannot be ruled out. It does not meet the standards for clinical drug use and may even pose significant safety risks.

[0110] This invention has verified its safety from multiple perspectives. In terms of materials, the invention primarily uses chitosan, gelatin, curcumin, and polyvinyl alcohol, all of which are certified by the U.S. Food and Drug Administration (FDA). Furthermore, we used Origin plotting software (version 9.1) and the SWeibull1 model to fit the dissolution curve of the curcumin-loaded clampable composite microneedles, simulating the product's dissolution in animals and humans. The curcumin-loaded clampable composite microneedles began to soften after 4 hours, no longer exhibiting a spherical shape. Between 8 and 12 hours, the curcumin-loaded clampable composite microneedles gradually softened and adhered to the surface of the pig large intestine in a layered, muddy manner. Given that the gastric emptying time for solid food in humans is 4-5 hours, the risk of intestinal obstruction is relatively low even if the patient eats immediately after the insertion of the curcumin-loaded clampable composite microneedles. We have verified the product's safety from multiple angles and aspects through various experiments.

[0111] (4) Medication costs:

[0112] Due to the low efficiency of oral administration, current common oral-based colon-targeted drug delivery systems are often designed to increase drug loading or targeting. This often increases the overall cost of the drug or device. For example, the manufacturing processes of commonly used nanoparticle-loaded, enzyme-loaded, and microparticle-loaded drugs are all relatively complex, costly, and expensive. If applied clinically, they may not be acceptable to patients.

[0113] The process of this invention is simple, involving only simple operations such as mixing, drying, hot perfusion, and cooling precipitation, resulting in low production costs. Furthermore, the raw materials used in this invention are inexpensive, have high drug utilization efficiency, and minimize raw material loss, further reducing production costs.

[0114] Those skilled in the art will readily understand that the above description is merely a preferred embodiment of the present invention and is 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 within the scope of protection of the present invention.

Claims

1. Use of curcumin loaded grippable composite microneedle in the preparation of a sustained release implantable drug delivery platform for the relief of ulcerative colitis, characterized in that, The curable compound microneedle loaded with curcumin comprises a gelatin hemispherical base loaded with curcumin and a chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin, which is located on the circular end face of the hemispherical base, the height of the microneedle is 1.90-2.00 mm, and the height direction of the microneedle is perpendicular to the circular end face; The curcumin in the base and the microneedle is loaded in the form of curcumin-cyclodextrin inclusion compound; The gelatin hemispherical base loaded with curcumin is prepared by mixing gelatin powder and deionized water, adding curcumin-cyclodextrin inclusion compound and then hot pouring and forming, and then cooling and drying to obtain a hemispherical body; The chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin is prepared by mixing CS solution and PVA solution, adding curcumin-cyclodextrin inclusion compound to configure a mixed pouring solution, repeatedly pouring into a microneedle mold, and then drying and demolding to obtain a microneedle part; The curable compound microneedle loaded with curcumin is used to be clamped to intestinal lesions by a colonoscope with clamping forceps and fixed, which can reduce the risk of intestinal obstruction, wherein the microneedle can be stably implanted into the intestinal wall, and the implantation depth can cover the intestinal mucosa layer to the submucosa layer; and the compound microneedle including the gelatin hemispherical base loaded with curcumin and the chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin can form a film on the surface of the ulcer, isolate the ulcer from harmful substances in the intestinal cavity, and delay the further development of ulcerative colitis.

2. The use according to claim 1, characterized in that, The gelatin hemispherical base loaded with curcumin and the chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin are bonded by hot-melt gelatin and then cooled and solidified to form a compound microneedle.

3. The use according to claim 1, characterized in that, The curcumin-cyclodextrin inclusion compound is prepared by mixing β-cyclodextrin solution and curcumin solution under heating, and then cooling, precipitating, filtering and drying; wherein the β-cyclodextrin solution is obtained by dissolving β-cyclodextrin in deionized water; and the curcumin solution is obtained by dissolving curcumin powder in ethanol.

4. The use according to claim 1, characterized in that, The microneedle is a quadrangular pyramid, and the bottom side length is 1.0-1.4 mm.

5. The use according to claim 1, characterized in that, The spherical diameter of the hemispherical base is 10-14 mm.

6. The use according to claim 1, characterized in that, The curable compound microneedle loaded with curcumin is prepared by the following preparation method comprising the following steps: S1. Synthesis of the gelatin hemispherical base loaded with curcumin: mixing gelatin powder and deionized water, adding curcumin-cyclodextrin inclusion compound and then hot pouring and forming, and then cooling and drying to obtain the gelatin hemispherical base loaded with curcumin; S2. Synthesis of the chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin: mixing CS solution and PVA solution, adding curcumin-cyclodextrin inclusion compound to configure a mixed pouring solution, repeatedly pouring into a microneedle mold, and then drying and demolding to obtain the chitosan (CS)-polyvinyl alcohol (PVA) microneedle loaded with curcumin; S3. Synthesis of the composite microneedle: The gelatin hemispheroid base loaded with curcumin was adhered to the curcumin-loaded chitosan (CS)-polyvinyl alcohol (PVA) microneedle by hot-melt gelatin and solidified by cooling to form the composite microneedle.

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