Risedronate sodium-bletilla striata polysaccharide soluble microneedle as well as preparation method and application thereof

Abstract

The invention discloses a risedronate sodium-bletilla striata polysaccharide soluble microneedle as well as a preparation method and application thereof. The risedronate sodium and the rhizoma bletillae polysaccharide are combined to prepare the soluble microneedle, so that the treatment effect after percutaneous delivery of the microneedle is equivalent to the oral administration effect; according to the soluble microneedle disclosed by the invention, the discomfort of gastrointestinal tracts of postmenopausal women after oral administration is reduced, the compliance of the postmenopausal women is also greatly improved, and a new alternative way is provided for the administration mode of osteoporosis of the postmenopausal women.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and particularly relates to a risedronate sodium-Bletilla striata polysaccharide soluble microneedle and a preparation method and application thereof. Background Art

[0002] Osteoporosis (OP) patients have a significant decrease in bone mass and damage to the microstructure of trabeculae, which greatly reduces bone strength and increases the risk of fractures. With the aging of the population, the incidence of primary osteoporosis is increasing, and postmenopausal osteoporosis (PMOP) is becoming more and more common.

[0003] Bisphosphonate anti-osteoporosis drugs are an important drug for the treatment of osteoporosis. Risedronate sodium is a representative drug among them, which mainly acts on bone metabolism by inhibiting osteoclast activity. Risedronate sodium can effectively improve the bone density of postmenopausal osteoporosis patients, effectively avoid the complications of fractures, and is beneficial to the quality of life of patients. Its administration method is mainly oral administration, but studies have shown that risedronate sodium is mucosal irritant, and oral administration can cause gastric mucosal damage, causing patients to have stomach pain, lack of appetite, indigestion, nausea and vomiting. In order to prevent these adverse reactions, osteoporosis patients should sit up or walk for at least 30 minutes after oral administration. Since osteoporosis patients mainly show fatigue, back pain or limb pain in clinical practice, many osteoporosis patients have difficulty taking risedronate sodium orally, and their compliance and quality of life are poor.

[0004] As a new transdermal drug delivery technology, microneedles are micron-sized needle structures with a height of 10-2000μm and a diameter of 10-50μm. They can penetrate the epidermis to form micropores, allowing drugs to pass through the stratum corneum and be delivered to the dermis. Compared with traditional patches, they improve the transdermal efficiency of drugs. Moreover, due to the length under the skin, they do not touch the nerve endings, just like the feeling of mosquito bites. Compared with subcutaneous injections, they have better patient compliance. Compared with oral administration, they avoid a series of gastrointestinal adverse reactions caused by oral administration, becoming an efficient and convenient transdermal drug delivery method. Bletilla striata polysaccharide (BSP) is the dominant active ingredient of Bletilla striata, with good hydrophilicity and stability, abundant availability, good bioadhesion and low toxicity. Bletilla striata polysaccharide helps to restore micro-trauma caused by microneedle insertion, such as preventing bacterial infection and reducing inflammation. Therefore, a Bletilla striata polysaccharide soluble microneedle technology is developed to deliver risedronate sodium percutaneously. The drug is delivered into the blood and into the systemic circulatory system by only piercing the stratum corneum barrier on the skin surface. It has the advantages of being painless, efficient, easy to operate and can be self-administered. Summary of the invention

[0005] In view of the above problems, the object of the present invention is to provide a risedronate sodium-Bletilla striata polysaccharide soluble microneedle and a preparation method thereof.

[0006] To achieve the above purpose, the technical solution adopted by the present invention is:

[0007] Application of risedronate sodium-Bletilla striata polysaccharide soluble microneedles in the preparation of a drug for treating postmenopausal osteoporosis.

[0008] The risedronate sodium-Bletilla striata polysaccharide soluble microneedle comprises the following components: risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide.

[0009] Preferably, the mass ratio of risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide is 3-5:100-300:50-200.

[0010] Particularly preferably, the mass ratio of risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide is 3:200:100.

[0011] Preferably, the polyvinyl pyrrolidone is replaced by one of polyvinyl alcohol or carboxymethyl chitosan.

[0012] The preparation method of the risedronate sodium-Bletilla striata polysaccharide soluble microneedle comprises the following steps:

[0013] (1) Preparation of matrix solution

[0014] 1) adding risedronate sodium into ultrapure water and dissolving it to obtain a risedronate sodium drug solution with a concentration of 3 to 5 mg / mL;

[0015] 2) Weighing polyvinyl pyrrolidone and Bletilla striata polysaccharide and placing them in a centrifuge tube, adding the risedronate sodium drug solution, stirring evenly, placing in a water bath to fully swell, and obtaining a risedronate sodium-Bletilla striata polysaccharide matrix liquid;

[0016] (2) Preparation of soluble microneedles

[0017] The risedronate sodium-Bletilla striata polysaccharide matrix liquid is placed in a microneedle concave mold, centrifuged, dried, and demolded to obtain the microneedle.

[0018] Preferably, the temperature of the water bath in step (1) is 20°C to 90°C; the centrifugal speed in step (2) is 3500 to 5500 rpm, and the centrifugal time is 10 to 60 min; the drying method is vacuum drying or room temperature drying, and the drying time is 6 to 48 hours.

[0019] Preferably, the temperature of the water bath in step (1) is 30°C to 60°C; the centrifugal speed in step (2) is 4000 to 4500 rpm, the centrifugal time is 20 to 40 min; and the drying time is 10 to 15 hours.

[0020] Preferably, the demoulding further includes drying at room temperature, and the drying time at room temperature is 3 to 7 days.

[0021] In the present invention, unless otherwise specified, % refers to mass percentage.

[0022] The present invention has the following beneficial effects:

[0023] The present invention combines risedronate sodium and Bletilla striata polysaccharide to prepare soluble microneedles, which can ensure that the therapeutic effect of the microneedles after transdermal delivery is equivalent to that of oral administration; therefore, the soluble microneedles in the present invention can reduce the discomfort in the gastrointestinal tract of postmenopausal women after oral administration, and also greatly improve their compliance, providing an alternative new way of administration for osteoporosis in postmenopausal women. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The appearance of soluble microneedles with different mixed matrices;

[0025] Figure 2 Parafilm insertion experiments for soluble microneedles with different mixed matrices;

[0026] Figure 3 This is a physical picture of risedronate sodium-Bletilla striata polysaccharide microneedle;

[0027] Figure 4 Optical microscope and scanning electron microscope images of risedronate sodium-Bletilla striata polysaccharide microneedles;

[0028] Figure 5 The mechanical properties of risedronate sodium-Bletilla striata polysaccharide microneedles;

[0029] Figure 6 The in vitro transdermal absorption of risedronate sodium-Bletilla striata polysaccharide microneedles;

[0030] Figure 7 To determine the bone metabolism related indexes of risedronate sodium-Bletilla striata polysaccharide microneedle;

[0031] Figure 8 Micro-CT of risedronate sodium-Bletilla striata polysaccharide microneedle;

[0032] Fig. 9 HE staining of gastric tissue of risedronate sodium-Bletilla striata polysaccharide. DETAILED DESCRIPTION

[0033] The following is further described in conjunction with the specific embodiments of the present invention. It should be noted that the description of these embodiments is used to help understand the present invention, but does not constitute a limitation of the present invention. In addition, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0034] Example 1

[0035] The method for making soluble microneedles with different mixed matrices includes the following steps: 10% Bletilla striata polysaccharide is mixed with matrix materials of different concentrations (5% / 10% / 20% carboxymethyl chitosan, 10% / 20% / 30% polyvinyl alcohol, 10% / 20% / 30% polyvinyl pyrrolidone), mixed and swollen in a 50°C water bath, and placed in a 4°C refrigerator overnight. Scrape an appropriate amount of the mixture onto the PDMS mold, put it into a centrifuge tube, centrifuge at 4500rpm for 30min to allow the solution to fill the micropores in the mold, put it into a room temperature dryer, and carefully demold after completely drying for 10 to 15 hours to obtain a composite microneedle. Results are shown in Figure 1 . The appearance of the microneedles prepared by mixing Bletilla striata polysaccharide (BSP) with different proportions of polyvinyl alcohol (PVA) is relatively flat. As the proportion of PVA increases, the hardness of the needle tip gradually increases, but the drying time takes longer than other matrices. The microneedles prepared by mixing different proportions of carboxymethyl chitosan CMC with BSP have good molding integrity. When CMC and BSP are mixed, the severe bending of the CMC backing is improved. The higher the proportion of CMC, the higher the hardness of the microneedles, but the backing is too hard and lacks the flexibility required for microneedles. The microneedles prepared by reducing the proportion of CMC are overall thin and lack the hardness of microneedles. The microneedles prepared by the mixed matrix of polyvinyl pyrrolidone (PVP) and BSP, when the proportion of PVP is 10%, the microneedles are well formed, but the hardness is slightly lower. When it is increased to 20%, the microneedles are well formed and have corresponding flexibility, but when it is increased to 30%, the flexibility of the microneedles decreases and they are easily brittle. And due to the particularity of the microneedle dosage form, the needle tip must have sufficient mechanical force to pierce the skin, and the backing needs to be flexible enough to fit the skin. Therefore, during the preparation process, it was found that a mixed matrix of PVP:BSP = 2:1 is more suitable.

[0036] Example 2

[0037] In order to more intuitively observe the depth of microneedles with different mixed matrices inserted into the skin, 9 Parafilm films were stacked together (the thickness of one Parafilm film was about 126 μm) and placed on the foam. The microneedles were inserted into the Parafilm film and pressed for about 1 minute. The microneedles were removed and the number of holes in each layer of the film and the number of layers of the film pierced by the microneedles were recorded to screen out the best mixed matrix. Parafilm film is widely used in human skin models because it has an elastic modulus and insertion resistance similar to that of the skin. Figure 2As shown in the figure, the percentage of micropores produced by microneedles penetrating the parafilm membrane at different depths accounts for the total number of microneedles in the array. It can be seen that basically the microneedles prepared by the mixed matrix investigated all penetrated the first layer. Except for CMC:BSP=1:2 and PVA:BSP=1:1, basically all the microneedles made of matrix materials can penetrate the second layer of parafilm membrane (252μm). Except for PVA:BSP=1:1 microneedles, the rest of the microneedles can achieve a pore production rate of more than 80% in the third layer. The percentage of micropores produced by CMC:BSP=2:1, PVP:BSP=2:1 and PVP:BSP=3:1 microneedles on the fourth layer of parafilm membrane is the highest among all types of mixed microneedles. The hardness of PVA:BSP=3:1 is slightly lower than these three groups, but its flexibility is the best. CMC:BSP=2:1 and PVP:BSP=3:1 have poor backing flexibility and are easily broken during pressing, which may affect the transdermal effect. Therefore, PVP:BSP=2:1 was finally selected for subsequent experiments. The penetration depth of the blank microneedle into the simulated skin was approximately in the range of 504-630μm, so the PVP:BSP=2:1 composite matrix is ​​the best.

[0038] Example 3

[0039] A method for preparing risedronate sodium-Bletilla striata polysaccharide soluble microneedles comprises the following process steps: preparing matrix liquid, putting the matrix liquid into a concave mold, centrifuging, drying and forming, demoulding and cutting.

[0040] 1. Configuration of matrix solution:

[0041] 1) Add risedronate sodium into a centrifuge tube, add ultrapure water, and dissolve by ultrasonication to obtain a risedronate sodium drug solution with a concentration of 3 mg / mL.

[0042] 2) Weigh 200 mg of polyvinyl pyrrolidone and 100 mg of Bletilla striata polysaccharide and place them in a centrifuge tube, then add 1 mL of the above-mentioned risedronate sodium drug solution to the mixture, stir evenly, and place in a water bath to allow it to fully swell, thereby obtaining a risedronate sodium-Bletilla striata polysaccharide matrix solution.

[0043] 2. Preparation of soluble microneedles

[0044] Scrape a small amount of matrix liquid and place it in the microneedle concave mold, centrifuge at 4500rpm for 30min, place the microneedle mold containing the matrix liquid in a room temperature dryer and let it stand for 10 to 15 hours, demould, peel the microneedles from the grooves of the microneedle mold along the edges of the microneedle mold to obtain a semi-finished patch, and then cut the excess parts around the patch to finally obtain risedronate sodium-Bletilla striata polysaccharide soluble microneedles, which are dried and stored at room temperature.

[0045] Example 4

[0046] Characterization of risedronate sodium-Bletilla striata polysaccharide soluble microneedle patch

[0047] The risedronate sodium-Bletilla striata polysaccharide soluble microneedles prepared in Example 3 are as follows Figure 3 and Figure 4 It is clearly visible that the morphology is a cone array. The prepared microneedles contain 100 needle tips arranged in a 10×10 array. The needle height is 800μm and the bottom diameter is 400μm. The microneedles prepared by this method have uniform shape and size, the microneedle array is complete and beautiful, the needle body surface is smooth, and the microscopic morphology is good.

[0048] Example 5

[0049] Study on the mechanical properties of risedronate sodium-Bletilla striata polysaccharide soluble microneedle patch

[0050] Take the risedronate sodium-Bletilla striata polysaccharide soluble microneedles prepared in Example 3 and test their mechanical properties through a texture analyzer to measure the relationship between the stress on the microneedle tip during the downward pressure of the probe and the displacement of the probe. Place the microneedle tip upward on the metal plate of the texture analyzer, start the instrument, and start the test. The metal probe of the instrument runs downward, the trigger force is set to 0.049N, the probe compresses the microneedle at a speed of 0.1mm / s, the compression distance is 0.8mm, and the force-displacement curve is recorded. According to the test results, the data is converted into the compressive elastic modulus of a single needle. Existing studies have shown that the maximum pressure that a single microneedle can withstand exceeds 0.048N to penetrate the epidermis of the skin. Figure 5 As shown, when the drug-loaded microneedle is compressed to 0.2mm, the pressure that a microneedle can withstand is 0.32N. Therefore, the mechanical strength of the drug-loaded microneedle is sufficient to pierce the skin, which provides a theoretical basis for the next step of skin experiments.

[0051] Example 6

[0052] In vitro transdermal absorption experiment of risedronate sodium-Bletilla striata polysaccharide soluble microneedles

[0053] After the rats were raised normally for 7 days, Veet depilatory cream was applied to the back skin of the rats for about 3 minutes, the depilatory cream and hair were wiped clean with clean water, and the rats were put back into the cage for normal feeding for 48 hours, and the back skin was observed to be in good condition. The mice were anesthetized with 10% urethane and killed, and their back skin was immediately cut off, the excess fascia of the rat back skin was cut off, and it was rinsed with saline and placed in saline, and placed in the freezer at -20°C for storage.

[0054] Prepare PBS solution as receiving solution, remove bubbles by ultrasonication for 30 minutes, pour the receiving solution into the diffusion cell, fill the diffusion cell with PBS solution, cover the diffusion cell with rat skin, with the stratum corneum facing the supply cell, cover the release cell on the rat skin, and fix the whole device with clips in the order of receiving cell, ex vivo skin, and release cell. Place the transdermal device in a 37°C constant temperature water bath, balance for 30 minutes, remove bubbles, and after the receiving solution temperature in the receiving cell is maintained at a constant temperature, contact or pierce the skin with free risedronate sodium drug solution, drug-loaded microneedles, and drug-loaded patches. Take samples of 1.0 mL at 0.5h, 1h, 2h, 4h, 8h, 12h, 24h, 36h, and 48h, respectively, and immediately add an equal amount of 37°C PBS solution after each sampling.

[0055] The sample was centrifuged at 5000 rpm for 10 min, and the supernatant was taken and measured by high performance liquid chromatography, and the peak area was recorded. The cumulative permeability per unit area of ​​risedronate sodium Q (μg·cm 2 ).

[0056]

[0057] Q is the cumulative permeation amount, Cn is the drug concentration measured at the nth sampling time point, Ci is the drug concentration measured at the ith sampling time point, V is the volume of the receiving solution, Vi is the sampling volume at the ith time point, and A is the transdermal diffusion area (1x1cm 2 )

[0058] Example 7

[0059] Pharmacodynamics study of risedronate sodium-Bletilla striata polysaccharide soluble microneedles

[0060] 1. Establishment of osteoporosis model in postmenopausal rats

[0061] Experimental animals: SD rats (SPF grade, female, 3 months old, provided by Beijing Huafukang Biotechnology Co., Ltd., license number: SCXK (Beijing) 2019-0008, approved by the Experimental Animal Ethics Committee of Guizhou Medical University, approval number: No. 2402936).

[0062] Feeding conditions: Raised in an ordinary animal room, ambient temperature: 17-25°C, relative humidity: 50-70%, 6 rats per frame, drinking water replaced daily, fed with standard feed, litter replaced every three days, adaptive feeding for 7 days.

[0063] The rat osteoporosis model was established by removing the ovaries of SD female rats. SD female rats were fasted for 12 hours, anesthetized by intraperitoneal injection of 10% urethane, and fixed on the surgical board in prone position. The excess hair was shaved with a razor, disinfected with alcohol, and a longitudinal incision was made 1 to 1.5 cm below the rear edge of the ribs and 1 to 1.5 cm beside the spine. The skin was pulled up with forceps, and the skin and muscles were peeled off layer by layer to fully expose the skin. A large mass of white fat was visible. A long strip of fallopian tube with horizontal texture could be found along the fat. Then, the mulberry-shaped ovary with light pink or dark red color could be found by exploring the fallopian tube to the distal end. The fallopian tube was clamped 0.5 cm below the ovary with curved forceps, the fallopian tube was ligated below the curved forceps, and the ovary was completely removed with ophthalmic scissors close to the curved forceps above the curved forceps. After loosening the curved forceps, carefully observe whether the hemostasis is complete, put it into the incision, and then operate on the other side, suture the muscles and skin respectively, and disinfect the wound with iodine. After the model is established, it is raised for three months, and finally the body weight, uterine index and bone tissue HE staining are observed to verify the success of the model.

[0064] 2. To investigate the therapeutic effect of risedronate sodium-Bletilla striata polysaccharide soluble microneedling on castrated osteoporotic rats

[0065] The osteoporotic rats after modeling were randomly divided into 5 groups, 5 rats in each group, and treated for a total of 28 days, including normal group, model group, RIS oral group, blank microneedle group and drug-loaded microneedle group;

[0066] Normal group: gavage with the same volume of 0.9% saline once a day during the treatment period;

[0067] Model group: During the treatment period, the same volume of 0.9% saline was administered once a day;

[0068] RIS oral group: 0.45 mg / kg risedronate sodium RIS (prepared with 0.9% saline) was administered intragastrically once a day during the treatment period;

[0069] Blank microneedle group: blank microneedles were given to the back during the treatment, one piece per day;

[0070] Drug-loaded microneedle group: drug-loaded microneedles were administered to the back during treatment, one tablet per day;

[0071] After 28 days of drug treatment, blood was collected from the femoral artery of the rats. After the blood samples were placed at room temperature for 1 to 2 hours, they were centrifuged (4°C, 3500 rpm, 10 min) and serum was separated. The serum of the rats was divided into EP tubes and stored at -20°C. Analysis of ELISA method was used to detect inflammatory factors related to rat bone metabolism. The tibia of the rats was scanned by Micro-CT, and image analysis of the selected area and calculation of related bone metabolism indicators were performed. Since the decline of estrogen will lead to the decline of the gastric mucosal system and the decrease of gastrointestinal function, oral treatment with sodium risedronate, which is irritating to the mucosa, may aggravate gastrointestinal damage. Therefore, the rats after blood collection were cut open along the midline of the abdomen of the rats, the stomach was taken out, and the stomach was cut open along the greater curvature of the stomach. The stomach contents were washed with saline and the stomach tissue was collected in a plastic bag. The collected stomach tissue was taken out and the pathological condition of the stomach tissue was evaluated by HE staining.

[0072] 3. Experimental Results

[0073] Figure 6 The in vitro permeation curves of RIS solution, drug-loaded patch RIS-BM (without needle tip) and drug-loaded microneedles (131.5±10.73μg) on ​​rat skin within 48 hours. The results showed that after 48 hours of drug-loaded microneedles, 103.50% of RIS was released from drug-loaded microneedles, 30.05% of RIS was released from free RIS drug solution, and 36.74% of RIS was released from drug-loaded patches. Therefore, the results show that the microneedle array loaded with RIS can significantly improve the transdermal delivery of RIS.

[0074] Three months after modeling, the rats were given oral administration of risedronate sodium RIS, blank microneedles, and drug-loaded microneedles for 28 days, and serum was collected to detect serum biochemical indicators using ELISA. Figure 7As shown in the results, compared with the normal group, the levels of serum osteocalcin (BGP), interleukin-6 (IL-6), tartrate-resistant acid phosphatase (TRAP) and cross-linked carboxyl-terminal peptide of type I collagen (CTX-I) in the model group were significantly increased (P < 0.001, P < 0.001, P < 0.001, P < 0.001), and the levels of osteoprotegerin (OPG) and amino-terminal propeptide of type I procollagen (PINP) were significantly decreased (P < 0.001, P < 0.001). 01), compared with the model group, after treatment, the levels of BGP, IL-6, TRAP and CTX-I in the RIS oral group and the drug-loaded microneedle group decreased significantly (P < 0.05, P < 0.001, P < 0.001, P < 0.001), and the levels of OPG and PINP increased significantly (P < 0.001, P < 0.001) (P < 0.01, P < 0.01), and there was no statistical difference between the RIS oral group and the drug-loaded microneedle group. Correspondingly, the blank microneedle group without drug loading did not show any therapeutic effect. The RIS oral group and the drug-loaded microneedle group showed similar anti-osteoporosis effects, indicating that the RIS oral group and the drug-loaded microneedle group had equivalent efficacy.

[0075] After treatment, the rat tibiae were removed for Micro-CT scanning. Figure 8 As shown in the figure, the trabeculae in the normal group were normal, dense, evenly distributed, crisscrossed into a network, and the bone microstructure was intact. Obviously, the trabeculae in the model group and the blank microneedle group were significantly sparse and broken, the number of trabeculae decreased, the separation was relatively wide, the bone microstructure was destroyed, and the structure was loosely arranged. After oral administration of RIS and drug-loaded microneedle treatment, it can be seen that the trabeculae and bone density were relatively improved, and the trabeculae and overall bone density increased significantly.

[0076] Image analysis was performed on the Micro-CT scans of rat tibiae and relevant bone metabolism indices were calculated, such as Figure 8As shown in the figure, compared with the normal group, the bone tissue fraction (BV / TV), trabecular number (Th.N) and total bone density (BMD) of the rats in the model group were significantly decreased (P<0.001, P<0.001, P<0.001), and the trabecular space (Th.Sp) was significantly increased (P<0.001), suggesting that the osteoporosis model of rats was successfully established; compared with the model group, the bone tissue fraction (BV / TV), trabecular number (Th.N) and total bone density (BMD) of the rats in the RIS oral group and the drug-loaded microneedle group were significantly decreased (P<0.001, P<0.001, P<0.001), and the trabecular space (Th.Sp) was significantly increased (P<0.001). .N) and total bone density (BMD) were significantly increased (P < 0.001, P < 0.001, P < 0.05) (P < 0.001, P < 0.001, P < 0.05, P < 0.01), and the trabecular space (Th.Sp) was significantly increased (P < 0.05), indicating that both oral administration of RIS and transdermal absorption of drug-loaded microneedles can delay bone loss and have anti-osteoporosis effects, and there is no significant statistical difference between the two groups, indicating that the two have equivalent efficacy. There is no significant difference in the trabecular thickness (Th.Th) between the groups, and it is speculated that osteoporosis after castration has no effect on trabecular thickness.

[0077] In summary, the therapeutic effect of risedronate sodium-Bletilla striata polysaccharide soluble microneedle was comprehensively evaluated by detecting Micro-CT and bone metabolic factors, and it was finally concluded that the therapeutic effects of oral administration of risedronate sodium and transdermal absorption of risedronate sodium-Bletilla striata polysaccharide soluble microneedle were equivalent.

[0078] Depend on Fig. 9 It can be seen that the normal group and gastric tissue structure are relatively normal, the mucosal layer, submucosa, muscular layer and adventitia structure are complete and clearly layered, blood vessels are scattered in the loose connective tissue of the submucosa, and the muscular layer and adventitia are tightly connected; in the model group, blank microneedle group, and drug-loaded microneedle group, slight necrosis of the surface layer of the local mucosal layer of the gastric tissue can be observed, a small amount of gastric glands can be seen falling off in the necrotic area, the structure is blurred, and a small amount of inflammatory cells are scattered, mainly lymphocytes with round nuclei and dark staining; while the RIS oral group showed large-area necrosis of the mucosal layer of the gastric tissue, cell disintegration in the necrotic area, blurred or disappeared gastric gland structure, accompanied by inflammatory cell infiltration, mainly lymphocytes with round nuclei and dark staining.

[0079] In summary, compared with the normal group, the model group showed slight pathological changes in gastric tissue. Compared with the model group, there was no significant change in gastric tissue lesions in the blank microneedle group and the drug-loaded microneedle group; the overall lesion severity in the RIS oral group was slightly aggravated, and the gastric tissue necrosis area was more extensive. Therefore, after changing the administration method of risedronate sodium, the damage of risedronate sodium to gastric tissue was avoided.

[0080] Example 8

[0081] By conducting a study on the pharmacokinetic of risedronate sodium RIS, the functional relationship between the blood concentration of the drug in vivo and time can be established, which is helpful to describe the quantitative change law of the drug in vivo and provide a certain reference for the in vivo application research of RIS-BMNs. The analysis was performed by a TQS triple quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source on the Acquity-Class UPLC system.

[0082] Ultra-high performance liquid chromatography conditions are as follows: Column: Waters BEH C 18 Column (2.1 mm × 50 mm, 1.7 μm), guard column: Waters van Guard BEH C 18 , flow rate: 0.35mL / min, mobile phase: 0.2% formic acid water (A)-0.2% formic acid acetonitrile (B), injection volume: 1μL; column temperature: 40℃, gradient elution conditions:

[0083] Table 1 Risedronate sodium gradient elution conditions

[0084] Time (min) Run parameters 0-0.4 A:98%; B:2% 0.4-0.6 A: 85%; B: 15% 0.6-0.8 A: 30%; B: 70% 0.8-1.0 A: 10%; B: 90% 1.0-1.5 A: 10%; B: 90% 1.5-2.0 A:98%; B:2% 2.0-3.0 A:98%; B:2%

[0085] The mass spectrometry conditions were as follows: electrospray ionization source (ESI), capillary ionization voltage: 2.0 KV, ion source temperature: 150°C, spray gas and backflush gas: N2, desolvation gas temperature: 400°C, desolvation gas flow rate: 1000 L / hr, collision gas flow rate: 0.15 mL / min.

[0086] Table 2 Risedronate sodium mass spectrometry conditions

[0087] Compound Precursor ion (m / z) Product ion (m / z) Cone voltage (V) Impact voltage(V) Puerarin (internal standard) 417 267 40 20 Risedronate 304.1 286 35 12

[0088] The above is a detailed description of the embodiments of the present invention, but the present invention is not limited to the described embodiments. For those skilled in the art, various changes, modifications, substitutions and variations of these embodiments are made without departing from the principles and spirit of the present invention, and still fall within the scope of protection of the present invention.

Claims

1. Application of risedronate sodium-Bletilla striata polysaccharide soluble microneedle in the preparation of a drug for treating postmenopausal osteoporosis.

2. The use according to claim 1, characterized in that: The risedronate sodium-Bletilla striata polysaccharide soluble microneedle comprises the following components: risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide.

3. The use according to claim 2, characterized in that: The mass ratio of risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide is 3-5:100-300:50-200.

4. The use according to claim 3, characterized in that: The mass ratio of risedronate sodium, polyvinyl pyrrolidone and Bletilla striata polysaccharide is 3:200:

100.

5. The use according to claim 2 or 3, characterized in that: The polyvinyl pyrrolidone is replaced by one of polyvinyl alcohol and carboxymethyl chitosan.

6. The use according to claim 1, characterized in that: The preparation method of the risedronate sodium-Bletilla striata polysaccharide soluble microneedle comprises the following steps: (1) Preparation of matrix solution 1) adding risedronate sodium into ultrapure water and dissolving the solution to obtain a risedronate sodium drug solution; 2) Weighing polyvinyl pyrrolidone and Bletilla striata polysaccharide and placing them in a centrifuge tube, adding the risedronate sodium drug solution, stirring evenly, placing in a water bath to fully swell, and obtaining a risedronate sodium-Bletilla striata polysaccharide matrix liquid; (2) Preparation of soluble microneedles The risedronate sodium-Bletilla striata polysaccharide matrix liquid is placed in a microneedle concave mold, centrifuged, dried, and demolded to obtain the microneedle.

7. The use according to claim 6, characterized in that: The temperature of the water bath in step (1) is 20°C to 90°C; the centrifugal speed in step (2) is 3500 to 5500 rpm, and the centrifugal time is 10 to 60 min; the drying method is vacuum drying or room temperature drying, and the drying time is 6 to 48 hours.

8. The use according to claim 7, characterized in that: In the step (1), the temperature of the water bath is 30° C. to 60° C.; in the step (2), the centrifugal speed is 4000 to 4500 rpm, the centrifugal time is 20 to 40 minutes; and the drying time is 10 to 15 hours.

9. The use according to claim 6, characterized in that: The process further comprises drying at room temperature after demoulding, and the drying time at room temperature is 3 to 7 days.

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