Preparation method of triamcinolone acetonide particles and application of drug-containing microneedle delivery system

Abstract

The invention relates to the technical field of medical materials, in particular to a preparation method of triamcinolone acetonide particles and application of a drug-containing microneedle delivery system, local delivery, slow release and better transdermal absorption of drugs are achieved by developing a new triamcinolone acetonide administration mode, the action time of the drugs is prolonged, and granulomatous mastitis is effectively treated. In addition, the microneedle array has the advantages of being good in biocompatibility, controllable in drug release, convenient to use and the like, and is suitable for local treatment of granulomatous mastitis.

Description

Technical Field

[0001] This application relates to the technical field of medical materials, and particularly to a preparation method of triamcinolone acetonide granules for treating granulomatous mastitis and the application of a drug-loaded microneedle delivery system. Background Art

[0002] Triamcinolone acetonide (TA), as a potent glucocorticoid drug, is widely used in the treatment of skin inflammation, arthritis, ophthalmic diseases, chronic pain and other indications due to its significant anti-inflammatory, immunosuppressive and anti-proliferative effects. However, its clinical application has long been limited by the defects of the physical and chemical properties of the drug itself: (1) extremely low solubility - the hydrophobic groups in the triamcinolone acetonide molecular structure account for a high proportion (such as acetylated side chains and fluorine atom substitution), resulting in an equilibrium solubility of less than 0.03 mg / mL (25 °C) in aqueous media, and it is difficult for conventional preparations to form a stable drug dispersion system; (2) low diffusion efficiency - after local administration, the drug is prone to accumulate on the stratum corneum or mucosal surface due to its high hydrophobicity, and it is difficult to penetrate the tissue barrier through passive diffusion, and the bioavailability is significantly limited (experimental data show that only about 1% - 3% of the drug in traditional creams can penetrate into the dermis layer); (3) weak transmembrane transport ability - although there is an affinity between the hydrophobic core of the cell membrane phospholipid bilayer and the triamcinolone acetonide molecule, its large molecular weight (434.5 Da) and rigid cyclic structure make it difficult to cross the membrane through free diffusion or carrier-mediated active transport, and it is difficult for the drug concentration in target cells (such as macrophages at the inflammation site) to reach the treatment threshold.

[0003] To improve the above defects, the existing technologies mainly adopt the following strategies:

[0004] The first category is the improvement of traditional preparations. For example, the apparent solubility is increased by adding surfactants (such as polysorbate 80) or co-solvents (such as propylene glycol). However, such methods have significant limitations: (a) the solubilization effect depends on high-concentration excipients, which may cause local irritation or allergic reactions; (b) the drug still exists in the form of crystals or micelle-wrapped, and cannot fundamentally change its diffusion kinetic characteristics, and the excipients may interfere with the binding efficiency of the drug to the target.

[0005] The second category is nanocarrier technology, including liposomes, polymeric nanoparticles, solid lipid nanoparticles, etc. For example, a kind of triamcinolone acetonide encapsulated by poly(lactic-co-glycolic acid) (PLGA) can improve its dispersibility through nanosizing. However, this kind of technology still faces the following challenges: (a) When the surface modification of nanoparticles is insufficient, drug burst release or carrier aggregation is likely to occur due to hydrophobic interaction; (b) There is a contradiction between the particle size control of nanoparticles (usually <200 nm) and the drug loading amount, and the process is complex and costly; (c) It is difficult to match the degradation rate of carrier materials (such as PLGA) with the drug release kinetics, which may lead to local pH changes or aggravated inflammatory reactions.

[0006] Meanwhile, as a new transdermal drug delivery tool, microneedle technology has attracted attention due to its minimally invasive property, high drug loading efficiency and controlled release characteristics. The microneedle array can form micron-sized pores through mechanical puncture, bypass the stratum corneum barrier and directly deliver drugs to the dermis layer, significantly improving drug utilization and reducing systemic toxicity. However, there are still technical bottlenecks in the combination of existing microneedle systems and triamcinolone acetonide nanoformulations: (1) Traditional silicon-based or metal microneedles have high mechanical strength but limited drug loading amount, and secondary surgery is required for removal, resulting in poor patient compliance; (2) Although soluble microneedles (such as hyaluronic acid matrix) can achieve non-invasive drug delivery, there is a contradiction between their rapid dissolution characteristics and the sustained release requirements of triamcinolone acetonide; (3) Existing microneedle preparation processes are difficult to simultaneously take into account the stability of nanoparticles, drug loading uniformity and needle body mechanical properties, resulting in obstacles to clinical translation.

[0007] In summary, how to further achieve precise drug control release, long-term effect and patient-friendly drug delivery through the "nanocarrier-microneedle delivery" synergistic technology on the basis of improving the solubility and transmembrane efficiency of triamcinolone acetonide, while avoiding the safety risks of traditional therapies, is a technical problem that needs to be solved urgently by those skilled in the art. Summary of the Invention

[0008] The purpose of this application aims to solve at least one of the above technical defects. The technical solutions adopted in this application are as follows:

[0009] In the first aspect, an embodiment of this application provides a preparation method of triamcinolone acetonide nanoparticles, which is characterized by including the following steps:

[0010] Step S1: Dissolve triamcinolone acetonide in propanol to form a first drug solution;

[0011] Step S2: Add the first drug solution described in Step S1 into an aqueous chitosan solution for mixing operation to obtain a second drug solution;

[0012] Step S3: Add a crosslinking agent solution to the second drug solution in Step S2 and react. During the reaction, use an alkaline solution to adjust the pH of the second drug solution to be maintained between 3.5 and 6.5 to obtain a first product;

[0013] Step S4: Centrifuge the first product obtained in Step S3 and take the lower layer precipitate to obtain a second product. Further, the preparation method of the crosslinking agent solution in Step S3 is: dissolve sodium tripolyphosphate and sodium dodecyl sulfate in pure water.

[0014] Further, the reaction in Step S3 is to continuously react at a stirring speed of 300 to 1000 revolutions per minute for 30 - 60 minutes.

[0015] Further, the alkaline solution in Step S3 is a 0.1 - 1.0 mol / L NaOH solution, and the pH value of the reaction system is between 5.5 ± 0.2.

[0016] Further, in Step S4, it also includes washing the second product with a sodium acetate buffer solution, then centrifuging again to take the lower layer precipitate to obtain a third product, and resuspending the third product in pure water, followed by ultrasonic dispersion and freeze - drying to obtain a fourth product.

[0017] In a second aspect, an embodiment of the present application provides a method for preparing a microneedle array, characterized in that,

[0018] Step S1: Dissolve polyvinyl alcohol in water to obtain a polyvinyl alcohol solution;

[0019] Step S2: Apply the polyvinyl alcohol solution obtained in Step S1 to the surface of the microneedle array mold and centrifuge;

[0020] Step S3: Thoroughly mix the triamcinolone acetonide nanoparticles obtained according to Claims 1 - 5 with the polyvinyl alcohol solution, and continue to apply it to the microneedle mold and centrifuge;

[0021] Step S4: Place the centrifuged microneedle mold in an oven for heating;

[0022] Step S5: Repeat Step S2, Step S3, and Step S4 until the water is completely evaporated;

[0023] Step S6: Add pullulan to the microneedle mold obtained in Step S5 to seal the bottom layer and then dry;

[0024] Step S7: Demold the microneedle array after drying is completed.

[0025] Further, in Step S1, during the process of dissolving polyvinyl alcohol in water, heat the solution for dissolution and centrifuge to remove air bubbles.

[0026] Further, the polyvinyl alcohol solution is a 15% polyvinyl alcohol solution.

[0027] Further, in step S2, before the polyvinyl alcohol solution is applied to the surface of the microneedle array mold, the polyvinyl alcohol solution is filtered using a 0.45 μm filter.

[0028] In a third aspect, an embodiment of the present application provides a triamcinolone acetonide nanoparticle, which is prepared by using the preparation method of the above-mentioned triamcinolone acetonide nanoparticle.

[0029] In a fourth aspect, an embodiment of the present application provides a microneedle array, including:

[0030] A polyvinyl alcohol (PVA) microneedle matrix;

[0031] Triamcinolone acetonide, as an active pharmaceutical ingredient;

[0032] Chitosan, as a drug loading and sustained release material, is used to control the modification and release of triamcinolone acetonide. In a fifth aspect, an embodiment of the present application provides an application of the microneedle array in the treatment of granulomatous mastitis.

[0033] The beneficial effects of the present application are as follows:

[0034] By constructing triamcinolone acetonide nanoparticles encapsulated with chitosan, and utilizing the electrostatic interaction between the cationic properties of chitosan and drug molecules, the in vivo solubility of triamcinolone acetonide is effectively improved, and with the high specific surface area and surface hydrophilic modification of the nanoparticles, the diffusion rate of the drug between tissues and the ability to penetrate cell membranes are enhanced; through microneedle sustained release and chitosan encapsulation, drug sustained release, better transdermal absorption and extended drug action time are achieved, and the treatment dosage required is significantly reduced. In addition, the microneedle array loads triamcinolone acetonide through chitosan, realizing local sustained release and targeted delivery of the drug, and can effectively treat granulomatous mastitis. The microneedle array has the advantages of good biocompatibility, controllable drug release, convenient use, etc., and is suitable for local treatment of granulomatous mastitis; and the present application realizes controllable sustained release drug delivery of triamcinolone acetonide in a minimally invasive manner, with high drug utilization; the minimally invasive drug delivery mode avoids the tissue damage and infection risks caused by traditional injection, and there is no needle body residue (using soluble microneedles), reducing the treatment pain of patients, and the acceptance of patients is high and it is suitable for popularization and use in clinical practice. Description of the Drawings

[0035] In order to more clearly illustrate the technical solutions in the embodiments of the present application, the following will briefly introduce the drawings required to be used in the description of the embodiments of the present application.

[0036] Figure 1 Schematic diagram of chitosan-loaded triamcinolone acetonide nanoparticles provided for the embodiment;

[0037] Figure 2 Schematic diagram of the microneedle array provided for the embodiment;

[0038] Figure 3a Partially enlarged schematic diagram of the microneedle array provided for the embodiment;

[0039] Figure 3b Partially enlarged schematic diagram of the microneedle array provided for the embodiment;

[0040] Figure 4a Comparison diagram of the rat inflammation model provided for the embodiment;

[0041] Figure 4b Schematic diagram after microneedle treatment of the rat inflammation model provided for the embodiment;

[0042] Figure 5 Comparison diagram before injection treatment and microneedle treatment and after injection treatment and microneedle treatment provided for the embodiment;

[0043] Figure 6 Schematic diagram of the changes in lymphocytes and macrophages in the injection group and the microneedle group provided for the embodiment;

[0044] Figure 7 Microneedle sustained-release time release curve provided for the embodiment;

[0045] Figure 8 Microneedle penetration ability test curve provided for the embodiment. Detailed implementation manners

[0046] The embodiments of the present application will be described in detail below. Examples of each embodiment are shown in the accompanying drawings, where the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions from beginning to end. The embodiments described below by referring to the accompanying drawings are exemplary and are only used to explain the present application, and cannot be construed as a limitation to the present application.

[0047] Those skilled in the art of the present technology can understand that unless specifically stated otherwise, the singular forms "a", "an" and "the" used herein may also include the plural forms. It should be further understood that the term "including" used in the specification of the present application means the presence of features, integers, steps, operations, elements and / or components, but does not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components and / or their groups. The term "and / or" used herein includes all or any unit and all combinations of one or more of the associated listed items.

[0048] To make the purpose, technical solutions and advantages of the present application clearer, the specific implementation manners of the present application will be further described in detail below with reference to the accompanying drawings.

[0049] Embodiment 1

[0050] This embodiment relates to a method for preparing triamcinolone acetonide nanoparticles. Figure 1 The following is a schematic diagram of chitosan-loaded triamcinolone acetonide nanoparticles in this embodiment. The preparation method includes the following steps:

[0051] Step S1: Dissolve triamcinolone acetonide in propanol to form a first drug solution.

[0052] Step S2: Add the first drug solution obtained in Step S1 to an aqueous chitosan solution and mix well to obtain a second drug solution.

[0053] In this embodiment, preferably, dissolve 4 mg of triamcinolone acetonide in propanol to obtain a first drug solution, add the first drug solution to a 10 ml polymer solution containing 0.1% chitosan (0.01 g), and ultrasonicate for 10 s × 3 times to obtain a second drug solution.

[0054] Step S3: Add a crosslinking agent solution to the second drug solution obtained in Step S2 for reaction. During the reaction, use an alkaline solution to adjust the pH of the system to be maintained between 3.5 and 6.5 to obtain a first product.

[0055] In this embodiment, preferably, the preparation method of the crosslinking agent solution in Step S3 is: dissolve sodium tripolyphosphate and sodium dodecyl sulfate in pure water; more preferably, the crosslinking agent solution is prepared by dissolving 3 mg of sodium tripolyphosphate and 1.5 mg of sodium dodecyl sulfate in 2 ml of pure water.

[0056] In this embodiment, preferably, the reaction in Step S3 is carried out at a stirring speed of 300 - 1000 revolutions per minute for 30 - 60 minutes; more preferably, the reaction in Step S3 is carried out at a stirring speed of 500 revolutions per minute for 45 minutes.

[0057] In this embodiment, preferably, the alkaline solution is a 0.1 - 1.0 mol / L NaOH solution, and the pH of the second drug solution is adjusted to be between 5.5 ± 0.2; more preferably, the pH value is 5.5.

[0058] Step S4: Centrifuge the first product obtained in Step S3 and take the lower layer precipitate to obtain a second product.

[0059] In this embodiment, preferably, centrifuge the first product obtained in Step S3 at 8000 r / 5 min.

[0060] In this embodiment, preferably, step S4 also includes washing the second product with sodium acetate buffer and then centrifuging again to remove the lower layer of precipitate to obtain a third product, and resuspending the third product in pure water, ultrasonically dispersing and freeze-drying to obtain a fourth product; more preferably, resuspending the third product in 1 ml of pure water, ultrasonically dispersing the particles for 5 seconds, and freeze-drying to obtain the fourth product.

[0061] Embodiment 2

[0062] This embodiment relates to a method for preparing a microneedle array, and the preparation method comprises the following steps:

[0063] Step S1: dissolving polyvinyl alcohol in water to obtain a polyvinyl alcohol solution;

[0064] In this embodiment, preferably, in the process of dissolving polyvinyl alcohol in water, the solution is heated to dissolve, and centrifuged to remove bubbles; more preferably, 200 mg of polyvinyl alcohol is dissolved in 1300 ul of pure water, heated to dissolve, and centrifuged to remove bubbles.

[0065] Step S2: Apply the polyvinyl alcohol solution obtained in step S1 to the microneedle array mold ( Figure 2 , Figure 3a , Figure 3b Centrifugation on the surface shown;

[0066] In this embodiment, preferably, the polyvinyl alcohol solution is a 1% to 20% polyvinyl alcohol solution; more preferably, the polyvinyl alcohol solution is a 15% polyvinyl alcohol solution.

[0067] In this embodiment, preferably, the 15% polyvinyl alcohol solution obtained in step S1 is applied to the surface of the microneedle array mold, and is sterilized by a filter, centrifuged, and bubbles are removed; more preferably, the filter is a 0.45 um filter.

[0068] Step S3: The triamcinolone acetonide nanoparticles prepared in Example 1 are fully mixed with the polyvinyl alcohol solution, and then applied to the microneedle mold and centrifuged.

[0069] In this embodiment, preferably, the triamcinolone acetonide nanoparticles are ground and then ultrasonicated for 5 minutes to be fully mixed with the polyvinyl alcohol solution and then applied to the microneedle mold.

[0070] Step S4: placing the centrifuged microneedle mold in a drying oven for heating;

[0071] In this embodiment, preferably, the microneedle mold is centrifuged at 4000 r / min for 20 min and then placed in a 60° C. drying oven and heated for 10 min to allow bubbles to float out.

[0072] Step S5: Repeat Step S2, Step S3, and Step S4 until the water is completely evaporated;

[0073] Step S6: Add the microneedle mold obtained in Step S5 to the pullulan polysaccharide sealing base layer and then dry it;

[0074] In this embodiment, preferably, add the microneedle mold obtained in Step S5 to the 10% pullulan polysaccharide sealing base layer and place it in an oven to dry at 37 °C for 24 h.

[0075] Step S7: Demold the microneedle array after drying is completed.

[0076] Example Three

[0077] This embodiment provides a triamcinolone acetonide nanoparticle, which is prepared by using the preparation method of a triamcinolone acetonide nanoparticle described in Example One above.

[0078] Example Four

[0079] This embodiment provides a microneedle array, and the microneedle array is prepared by using the preparation method of a microneedle array described in Example Two above.

[0080] The preferred solution in this embodiment is:

[0081] 1. The concentration of the PVA microneedle matrix is 15% (w / v), the concentration of chitosan is 0.1% (w / v), and the loading amount of triamcinolone acetonide is 0.6 mg per microneedle matrix.

[0082] 2. The length of the microneedles is 700 ± 20 μm, the diameter is from 230 μm to 190 μm, and the area of the microneedle array is 1.26 cm 2 *1.26 cm 2 .

[0083] Example Five

[0084] This embodiment provides an application of a microneedle array in the treatment of granulomatous mastitis. The application frequency of the microneedle array is once a week, and it is specifically adjusted according to the patient's condition and the drug release curve.

[0085] Animal experiment:

[0086] Construct a rat granulomatous mastitis model, and analyze the efficacy and safety of microneedle sustained-release treatment from phenotypes, imaging, histopathology, and humoral immune mechanisms after respectively administering triamcinolone acetonide by microneedle and by injection.

[0087] Experimental purpose:

[0088] 1. Evaluate whether the transdermal administration of TA microneedles can achieve a treatment effect equivalent to that of TA injection treatment

[0089] 2. Observe the safety and feasibility of microneedle drug delivery

[0090] Experimental grouping:

[0091] Control group; microneedle treatment group;

[0092] Experimental procedures:

[0093] 1. Animal model establishment: Homogenize human granulomatous lobular mastitis tissue, mix it with complete Freund's adjuvant at a volume ratio of 1:1 to make an oil-in-water emulsion suspension, and inject it into the mammary gland of female SD rats. Metoclopramide is subcutaneously injected to induce hyperprolactinemia. Pathological sections and imaging diagnosis are granulomatous mastitis.

[0094] 2. Apply the microneedle array to the mammary gland lesion site, treat once a week for 1 consecutive week.

[0095] 3. After treatment, the inflammatory symptoms of the diseased rats were significantly improved, and no obvious side effects were observed.

[0096] Experimental results:

[0097] The experimental comparison results of the rat inflammation model are shown in Figure 4a , Figure 4b ; among them, Figure 4a is the control group, Figure 4b is the microneedle treatment group; it can be seen that the inflammatory mass significantly regressed after microneedle treatment.

[0098] Figure 5 is the experimental result comparison chart before injection treatment and microneedle treatment and after injection treatment and microneedle treatment; it can be seen that in both the injection treatment group and the microneedle treatment group, it was observed that the originally large mass gradually shrank and even disappeared after treatment.

[0099] Figure 6 As shown in

[0100] In both the injection group and the microneedle group, it was observed that the infiltration of lymphocytes and macrophages was significantly reduced, the granuloma structure gradually regressed, and the inflammatory response was controlled. Figure 7 The drug release performance test of the microneedle array is as

[0101] 1. Place the microneedle array in phosphate buffer (PBS, pH 7.4) and conduct a drug release experiment at 37°C.

[0102] 2. Sampling at different time points, and determine the release amount of triamcinolone acetonide by high performance liquid chromatography (HPLC).

[0103] 3. The results showed that the microneedle array released approximately 30% of the drug within 24 hours and approximately 80% of the drug within 5 days, indicating its good sustained-release performance. This microneedle system conforms to the non-Fickian release mechanism dominated by initial diffusion and synergistic diffusion-erosion in the later stage, and is applicable to the treatment of granulomatous mastitis diseases that require rapid onset and continuous drug administration.

[0104] Safety: Liver and kidney function tests

[0105]

[0106] Table 1

[0107]

[0108] Table 2

[0109] Con: Blank control group; C+M: Microneedle alone group; C+SC: Injection alone group

[0110] Note: One case of abnormal liver function occurred in the injection group, and obvious liver and kidney toxicity occurred in the microneedle treatment group.

[0111] Transdermal release in breast tissue: Human breast tissue was taken and microneedles were inserted. On the 1st day, 3rd day, and 6th day, the content of triamcinolone acetonide in the adipose layer and glandular layer was detected to reflect the percutaneous penetration and diffusion ability. The results showed that triamcinolone acetonide was detected in both the adipose layer and glandular layer. The content release curve is shown in Figure 8 as shown.

[0112] Conclusion:

[0113] 1. In the treatment of granulomatous mastitis, animal experiments showed that microneedle administration could achieve the same therapeutic effect as injection administration. In clinical application, compared with traditional drug administration methods, microneedle treatment is not only simple to operate but also reduces the pain of patients, which is more conducive to clinical promotion and application.

[0114] 2. No obvious toxic and side effects were observed in the microneedle treatment group. Compared with injection, microneedle controlled-release drug administration improves the safety of hormone use.

[0115] The above content is only an example and explanation of the structure of this application. Those skilled in the art of this technology can make various modifications or supplements to the specific embodiments described or use similar methods to replace them. As long as they do not deviate from the structure of the application or exceed the scope defined by this claim book, they should fall within the protection scope of this application.

Claims

1. A preparation method of triamcinolone acetonide nanoparticles, characterized in that, The following steps are involved: Step S1: dissolving triamcinolone acetonide in propanol to form a first drug solution; Step S2: adding the first drug solution in step S1 to the chitosan sodium acetate solution and mixing them to obtain a second drug solution; Step S3: adding a crosslinking agent solution to the second drug solution of step S2 for reaction, during which an alkaline solution is used to adjust the pH of the second drug solution to be maintained between 3.5 and 6.5, to obtain a first product; Step S4: After centrifuging the first product obtained in step S3, the lower precipitate is removed to obtain the second product.

2. The preparation method according to claim 1, wherein, The preparation method of the cross-linking agent solution in step S3 is: dissolving sodium tripolyphosphate and sodium dodecyl sulfate in pure water.

3. The preparation method according to claim 1, characterized in that, The reaction in step S3 is continued for 30 to 60 minutes at a stirring speed of 300 to 1000 rpm.

4. The preparation method according to claim 1, wherein, In step S3, the alkaline solution is a 0.1-1.0 mol / L NaOH solution, and the pH value of the second drug solution is adjusted to be between 5.5±0.

2.

5. The preparation method according to claim 1, characterized in that, Step S4 also includes washing the second product with sodium acetate buffer and centrifuging again to remove the lower precipitate to obtain a third product, and resuspending the third product in pure water, dispersing by ultrasonication, and freeze-drying to obtain a fourth product.

6. A method for preparing a microneedle array, characterized in that: Step S1: dissolving polyvinyl alcohol in water to obtain a polyvinyl alcohol solution; Step S2: applying the polyvinyl alcohol solution obtained in step S1 to the surface of the microneedle array mold and centrifuging; Step S3: fully mixing the triamcinolone acetonide nanoparticles obtained according to claims 1-5 with the polyvinyl alcohol solution, and then applying the mixture to the microneedle mold and centrifuging the mixture; Step S4: placing the centrifuged microneedle mold in a drying oven for heating; Step S5: repeating steps S2, S3, and S4 until the water is completely evaporated; Step S6: adding pullulan to the microneedle mold obtained in step S5 to seal the base layer and then drying; Step S7: demoulding the microneedle array after drying is completed.

7. The preparation method according to claim 6, characterized in that, In step S1, during the process of dissolving polyvinyl alcohol in water, the solution is heated to dissolve and centrifuged to remove bubbles.

8. The preparation method according to claim 6, characterized in that, The polyvinyl alcohol solution is a 15% polyvinyl alcohol solution.

9. The preparation method according to claim 6, characterized in that, In step S2, before applying the polyvinyl alcohol solution to the surface of the microneedle array mold, the polyvinyl alcohol solution is filtered using a 0.45 um filter.

10. A triamcinolone acetonide nanoparticle, characterized in that, The method is prepared by the method as described in claims 1-5.

11. A microneedle array, characterized in that, include: Polyvinyl alcohol (PVA) microneedle matrix; triamcinolone acetonide, as the active pharmaceutical ingredient; Chitosan, as a drug loading and sustained-release material, was used to control the modification and release of triamcinolone acetonide.

12. Use of the microneedle array according to claim 11 in treating granulomatous mastitis.

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