Preparation method and application of drug delivery system based on hollow mesoporous polydopamine nanoparticles

Abstract

The invention belongs to the field of drug preparation, and particularly relates to a preparation method and application of a drug delivery system based on hollow mesoporous polydopamine nanoparticles. The drug delivery system comprises hollow mesoporous polydopamine nanoparticles, a drug loaded inside and / or outside the hollow mesoporous polydopamine nanoparticles, and pectin wrapping the hollow mesoporous polydopamine nanoparticles, the medicine is a medicine with a nerve protection effect and an acetylcholin esterase inhibitor. The drug delivery system can reduce proinflammatory cytokines, regulate functions of target immune cells, synergistically reduce necrosis of neurons in intestinal tracts and induce neural crest stem cells in the intestinal tracts to be differentiated into neurons.

Description

Technical Field

[0001] The invention belongs to the field of drug preparation, and in particular relates to a preparation method and application of a drug delivery system based on hollow mesoporous polydopamine nanoparticles. Background Art

[0002] Inflammatory bowel diseases (IBDs) mainly include Crohn's disease and ulcerative colitis. IBD has a high incidence worldwide, with the highest incidence of Crohn's disease in North America. Statistics show that an estimated 129,000 people in Canada suffer from the disease. Although the onset of the disease is usually in adulthood, an increasing number of children are diagnosed with IBD. Although the mortality rate of the disease is low, it is still incurable and requires lifelong treatment. In addition, IBD patients have a much higher risk of depression and neurodegenerative diseases than ordinary people. Due to the prolonged and recurrent course of the disease, it seriously affects the quality of life of patients and also places a huge burden on the medical system.

[0003] IBD is usually treated with drugs that can relieve symptoms and reduce inflammation of the colon lining, such as anti-inflammatory drugs including 5-aminosalicylic acid, and immunosuppressants such as azathioprine, mercaptopurine, and methotrexate. These therapies are very effective in reducing the level of inflammation, but they also have many side effects. For example, taking these drug compounds can cause fluid retention, insomnia, weight gain, drowsiness, high blood pressure, constipation, and vomiting.

[0004] At present, the Western medicine treatment of IBD has also shifted from simple anti-inflammatory to correcting disordered intestinal immune function. More and more Western medicine treatment plans focus on biological treatment of immune-related factors such as TNF-α, IL-12, and IL-23. However, more than 1 / 3 of IBD patients do not respond to biological therapy against TNF-α. Among those patients who respond to infliximab or adalimumab treatment, there is still a 13% or 20% risk of losing responsiveness to these two biological agents each year, leading to treatment failure. At the same time, biological agents are expensive, and long-term use brings a heavy economic burden to patients, and can cause patients to suffer from autoimmune diseases, skin diseases (such as eczema, psoriasis), and even induce malignant tumors. Therefore, immunosuppressants are still the main drugs for Western medicine to maintain the treatment of IBD. Summary of the invention

[0005] In order to solve the problems in the prior art, such as strong drug side effects, expensive biological preparations, and poor drug delivery efficiency, the present invention mainly provides a drug delivery system based on hollow mesoporous polydopamine nanoparticles that can deliver drugs to IBDs diseased intestinal tissues:

[0006] A drug delivery system based on hollow mesoporous polydopamine nanoparticles comprises a drug-loaded precursor formed by hollow mesoporous polydopamine nanoparticles and a drug loaded inside and / or outside the hollow mesoporous polydopamine nanoparticles, and an enteric material wrapped outside the drug-loaded precursor; the drug comprises a drug with a neuroprotective effect and an acetylcholinesterase inhibitor.

[0007] Furthermore, the drug with neuroprotective effect is piracetam; the acetylcholinesterase inhibitor is galantamine hydrobromide; and the enteric material includes one of pectin, shellac, polyvinyl alcohol acetate phthalate, acrylic resin, cellulose or chitosan.

[0008] Furthermore, the drug loading of the neuroprotective drug is 35-45%; and the drug loading of the acetylcholinesterase inhibitor is 15-25%.

[0009] Furthermore, the average particle size of the drug-loaded precursor is 300 to 1000 nm.

[0010] A method for preparing the above-mentioned hollow mesoporous polydopamine nanoparticle-based drug delivery system comprises the following steps:

[0011] a. Adding polyoxyethylene-polyoxypropylene-polyoxyethylene to a mixed solution of water and ethanol under stirring and ultrasonic conditions, dispersing the mixture uniformly to obtain a polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, adding 3,3',5,5'-tetramethylbenzidine to the dispersion, mixing for 1 to 3 hours to obtain a first mixed system; adding tris(hydroxymethyl)aminomethane hydrochloride) solution and dopamine hydrochloride solution to the first mixed system, continuing to stir and react, to obtain a mixed solution; after centrifugation and washing, collecting and obtaining hollow mesoporous polydopamine nanoparticles;

[0012] b, the hollow mesoporous polydopamine nanoparticles are fully dispersed in the saturated solution of the drug, and fully mixed to obtain a mixed solution; the solid is separated, collected and freeze-dried to obtain a drug-loaded precursor;

[0013] c. Prepare a drug-loaded precursor dispersion, then add the enteric material solution into the drug-loaded precursor dispersion, and mix thoroughly to obtain the product.

[0014] Furthermore, in step a, the concentration of the polyoxyethylene-polyoxypropylene-polyoxyethylene in the mixed solution of water and ethanol is 2 to 10 mg / mL; the volume ratio of water to ethanol solution in the mixed solution of water and ethanol is 1:0.25 to 4; the volume ratio of the polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion to 3,3',5,5'-tetramethylbenzidine is 80 to 120:1; the concentration of the tris(hydroxymethyl)aminomethane hydrochloride) solution is 40 to 50 mg / mL; the concentration of the dopamine hydrochloride solution is 5 to 10 mg / mL; the volume ratio of the tris(hydroxymethyl)aminomethane hydrochloride solution to the dopamine hydrochloride solution is 1:1; and the volume ratio of the first mixed system to the dopamine hydrochloride solution is 12 to 15:1.

[0015] Furthermore, the stirring speed of step a is 400-500 rpm, the ultrasonic frequency is 30-40 kHz, and the ultrasonic power is 200-300 W; the stirring reaction time of step a is 18-24 h; the washing liquid of the centrifugal washing in step a is a mixture of acetone and ethanol in a volume ratio of 1:2; the centrifugal conditions are 13000 rpm, 10 min, and the hollow mesoporous polydopamine nanoparticles are collected by centrifugal washing twice.

[0016] Furthermore, the stirring speed of the mixing in step b is 300-800 rpm, and the mixing time is 20-28 h; the centrifugal condition of the separation in step b is 13000 rpm, 10 min; the separation in step b is centrifuged once to collect the precipitate; the freeze-drying condition in step b is -80

[0017] ℃, freeze-dried at 2Pa.

[0018] Furthermore, the concentration of the drug-loaded precursor aqueous dispersion in step c is 2-20 mg / mL; the concentration of the enteric material in step c is 2-20 mg / mL; and the volume ratio of the drug-loaded precursor dispersion to the enteric material solution is 1:0.25-4.

[0019] An application of the above-mentioned hollow mesoporous polydopamine nanoparticle-based drug delivery system in the preparation of a drug for treating inflammatory bowel disease is characterized in that the nerve abundance of damaged intestinal tissue is maintained and the neuroimmune function in the diseased intestinal tissue is restored.

[0020] By adopting the above scheme, the method of the present invention has the following advantages:

[0021] 1. The drug delivery system of the present invention creatively uses piracetam, a neurotrophic drug that is not used to treat gastrointestinal diseases, and galantamine hydrobromide, an anticholinesterase. The two drugs work together to restore the neural network structure in the intestine of IBD patients, reduce proinflammatory cytokines through the CAP pathway, regulate the function of target immune cells, synergistically reduce the necrosis of neurons in the intestine, and induce the differentiation of neural crest stem cells in the intestine into neurons.

[0022] 2. The present invention loads drugs with neuroprotective effects and acetylcholinesterase inhibitors into hollow porous polydopamine nanoparticles, which can synergistically exert anti-inflammatory effects while repairing the intestinal barrier system and rebuilding intestinal homeostasis. Enteric materials are selected to modify the co-drug delivery system to achieve the function of targeted immune cell drug delivery, prevent drug leakage and premature release after oral administration of the drug delivery system, and achieve the purpose of targeted intestinal immune cell delivery.

[0023] 3. The drug delivery system of the present invention is based on hollow mesoporous polydopamine nanoparticles, which can greatly increase the drug-loading capacity of the nano-platform and facilitate the co-delivery of multiple therapeutic drugs. The construction of the nano-sized drug delivery system can make full use of the leakage characteristics of the IBD intestine to achieve passive targeting and further improve the drug delivery efficiency. No hazardous reagents are used in the preparation process, making the construction safer and non-toxic.

[0024] 4. Neuron-macrophage crosstalk is one of the key units for the vagus nerve-cholinergic anti-inflammatory pathway (CAIP). Stimulating the vagus nerve (VNS) to obtain significant anti-inflammatory effects has been used in clinical treatment, but VNS therapy has its limitations, that is, it is subject to the structural and functional integrity of the nervous tissue. The present invention is based on repairing the structure and function of damaged intestinal nervous tissue, which can correct the intestinal neuroimmune function of IBD disorders and provide support for improving the therapeutic effect of IBD.

[0025] 5. The drug delivery system of the present invention can significantly increase the proportion of M2 macrophages in tissues. Compared with the pro-inflammatory phenotype M1 macrophages, M2 macrophages have anti-inflammatory, lymphangiogenic and wound healing effects, and can also increase the barrier function of intestinal epithelial cells.

[0026] 6. The hollow mesoporous polydopamine nanoparticles used in the drug delivery system of the present invention also have good SOD-like enzyme activity and can effectively remove ROS in tissues. Removing excess ROS in tissues helps neural stem cells to differentiate into neurons and helps macrophages to differentiate into M2 type. These effects of hollow mesoporous polydopamine are very beneficial for controlling the inflammatory state of the intestine and promoting the repair of intestinal neuroimmune function.

[0027] 7. The drug delivery system of the present invention uses an enteric material with intestinal targeting capability to wrap the drug delivery system to prevent the drug from being released prematurely or destroyed in the gastric acid environment. Pectin can be prevented from being degraded prematurely by gastric acid or digestive enzymes, and the intestinal targeting property enables the drug to act more directly on the intestine. This property helps to prolong the release time of the drug, thereby improving the bioavailability of the drug.

[0028] 8. The present invention directly mixes pectin with the drug-loaded hollow mesoporous polydopamine to form a coating, which fully reduces drug leakage from the drug-loaded hollow mesoporous polydopamine and works together with the hollow mesoporous polydopamine to achieve uniform sustained release of the drug. The coating step is simple and does not introduce impurities.

[0029] 9. The pectin in the drug delivery system of the present invention can promote the growth of beneficial bacteria in the intestines, help maintain a healthy microbial community, thereby improving digestion and preventing constipation. The natural antioxidants contained in the pectin can reduce the damage of free radicals in the body and reduce the risk of chronic diseases. Pectin can increase the protection of the intestinal mucosa and improve the absorption of nutrients. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 It is a comparison diagram of the particle size and potential of PDA and Drugs@PDA obtained in step 1 and step 2 of Example 1, respectively.

[0031] Figure 2 These are transmission electron micrographs of PDA (a) and Drugs@PDA (b) obtained in step 1 and step 2 of Example 1, respectively.

[0032] Figure 3 It is a comparison chart of the ratios of M2 macrophages and TNF-α, IL-1β, and IL-6 in comparative example 1 and comparative example 3 in the anti-inflammatory test.

[0033] Figure 4 It is a comparison chart of the weight changes of mice after administration of Example 1 and Comparative Examples 1 to 3.

[0034] Figure 5 It is a comparison chart of the colon length of mice after administration of Example 1 and Comparative Examples 1 to 3.

[0035] Figure 6 It is a stacked column chart of the intestinal flora of mice after administration of Example 1 and Comparative Examples 1 to 3.

[0036] Figure 7 It is a Venn diagram of the intestinal flora of mice after administration of Example 1 and Comparative Examples 1 to 3.

[0037] Figure 8 It is a comparison chart of the abundance of enteric neurons in mice after administration of Example 1 and Comparative Examples 1 to 3. DETAILED DESCRIPTION

[0038] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0039] Embodiment 1:

[0040] (1) under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene was added to a mixed solution of water and ethanol in a volume ratio of 1:1, and dispersed evenly to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine was added to the dispersion in a volume ratio of 100:1, and mixed for 2 hours to obtain a first mixed system; 45 mg / mL tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL dopamine hydrochloride aqueous solution were added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture was stirred and reacted for 20 hours to obtain a mixed solution; the mixture was centrifuged at 13000 rpm for 10 minutes, and washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and centrifuged and washed twice to collect hollow mesoporous polydopamine nanoparticles;

[0041] (2) the hollow mesoporous polydopamine nanoparticles are fully dispersed in a saturated aqueous solution of the drug, and the mixture is stirred at a speed of 400 to 500 rpm for 24 hours to obtain a mixed solution; the solid is collected by centrifugation and freeze-dried at -80°C and 2 Pa to obtain a drug-loaded precursor;

[0042] (3) Prepare a drug-loaded precursor aqueous dispersion, then add a 10 mg / mL pectin solution to the drug-loaded precursor aqueous dispersion at a volume ratio of 1:1, and mix thoroughly to obtain (i.e., Drugs@PDA+Pectin).

[0043] Depend on Figure 1 It can be seen that the average particle size of the hollow mesoporous polydopamine nanoparticles (PDA) obtained in step 1 is 200-400 nm, and the zeta potential is -12 mV; the average particle size of the drug-loaded precursor (i.e., Drugs@PDA) obtained in step 2 is 600 nm, and the zeta potential is -19 mV.

[0044] Depend on Figure 2 From the transmission electron microscopy photos, it can be seen that the PDA obtained in step 1 of Figure a has a hollow structure observable under the microscope, and the Drugs@PDA obtained in step 2 of Figure b is observed to be spherical after drug loading, with no clearly visible hollow structure, indicating that the drug has been successfully loaded.

[0045] Embodiment 2:

[0046] (1) under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene was added to a mixed solution of water and ethanol in a volume ratio of 1:1, and dispersed evenly to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine was added to the dispersion in a volume ratio of 100:1, and mixed for 2 hours to obtain a first mixed system; 45 mg / mL tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL dopamine hydrochloride aqueous solution were added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture was stirred and reacted for 20 hours to obtain a mixed solution; the mixture was centrifuged at 13000 rpm for 10 minutes, and washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and centrifuged and washed twice to collect hollow mesoporous polydopamine nanoparticles;

[0047] (2) the hollow mesoporous polydopamine nanoparticles are fully dispersed in a saturated aqueous solution of the drug, and the mixture is stirred at a speed of 400 to 500 rpm for 24 hours to obtain a mixed solution; the solid is collected by centrifugation and freeze-dried at -80°C and 2 Pa to obtain a drug-loaded precursor;

[0048] (3) Prepare a drug-loaded precursor aqueous dispersion, then add a 10 mg / mL pectin solution to the drug-loaded precursor aqueous dispersion at a volume ratio of 1:0.5, and mix thoroughly to obtain the product.

[0049] Embodiment 3:

[0050] (1) under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene was added to a mixed solution of water and ethanol in a volume ratio of 1:1, and dispersed evenly to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine was added to the dispersion in a volume ratio of 100:1, and mixed for 2 hours to obtain a first mixed system; 45 mg / mL tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL dopamine hydrochloride aqueous solution were added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture was stirred and reacted for 20 hours to obtain a mixed solution; the mixture was centrifuged at 13000 rpm for 10 minutes, and washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and centrifuged and washed twice to collect hollow mesoporous polydopamine nanoparticles;

[0051] (2) the hollow mesoporous polydopamine nanoparticles are fully dispersed in a saturated aqueous solution of the drug, and the mixture is stirred at a speed of 400 to 500 rpm for 24 hours to obtain a mixed solution; the solid is collected by centrifugation and freeze-dried at -80°C and 2 Pa to obtain a drug-loaded precursor;

[0052] (3) Prepare a drug-loaded precursor aqueous dispersion, then add 10 mg / mL pectin solution to the drug-loaded precursor aqueous dispersion at a volume ratio of 1:1.5, and mix thoroughly to obtain. Comparative Example 1:

[0053] (1) under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene was added to a mixed solution of water and ethanol in a volume ratio of 1:1, and dispersed evenly to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine was added to the dispersion in a volume ratio of 100:1, and mixed for 2 hours to obtain a first mixed system; 45 mg / mL tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL dopamine hydrochloride aqueous solution were added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture was stirred and reacted for 20 hours to obtain a mixed solution; the mixture was centrifuged at 13000 rpm for 10 minutes, and washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and centrifuged and washed twice to collect hollow mesoporous polydopamine nanoparticles;

[0054] (2) The hollow mesoporous polydopamine nanoparticles are fully dispersed in a saturated aqueous solution of the drug, and the mixture is stirred at a speed of 400 to 500 rpm for 24 hours to obtain a mixed solution; the solid is collected by centrifugation and freeze-dried at -80°C and 2Pa to obtain (i.e., Drugs@PDA).

[0055] Comparative Example 2:

[0056] (1) under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene was added to a mixed solution of water and ethanol in a volume ratio of 1:1, and dispersed evenly to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine was added to the dispersion in a volume ratio of 100:1, and mixed for 2 hours to obtain a first mixed system; 45 mg / mL tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL dopamine hydrochloride aqueous solution were added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture was stirred and reacted for 20 hours to obtain a mixed solution; the mixture was centrifuged at 13000 rpm for 10 minutes, and washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and centrifuged and washed twice to collect hollow mesoporous polydopamine nanoparticles;

[0057] (2) The hollow mesoporous polydopamine nanoparticles are fully dispersed in a saturated aqueous solution of the drug, and stirred and mixed at a speed of 400 to 500 rpm for 24 hours to obtain (i.e., PDA+Drugs Mixture).

[0058] Comparative Example 3:

[0059] (1) Under the conditions of stirring at a speed of 400 to 500 rpm and ultrasound at a frequency of 30 to 40 kHz and a power of 200 to 300 W, polyoxyethylene-polyoxypropylene-polyoxyethylene is added to a mixed solution of water and ethanol in a volume ratio of 1:1, and the mixture is evenly dispersed to obtain a 6 mg / mL polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, 3,3',5,5'-tetramethylbenzidine is added to the dispersion in a volume ratio of 100:1, and the mixture is mixed for 2 hours to obtain a first mixed system; 45 mg / mL of tris(hydroxymethyl)aminomethane hydrochloride aqueous solution and 7.5 mg / mL of dopamine hydrochloride aqueous solution are added to the first mixed system in a volume ratio of 27.25:1:1, and the mixture is stirred and reacted for 20 hours to obtain a mixed solution; the mixture is centrifuged at 13000 rpm for 10 minutes, and the mixture is washed with a mixed solution of acetone and ethanol in a volume ratio of 1:2, and the mixture is centrifuged and washed twice to obtain hollow mesoporous polydopamine nanoparticles (i.e., PDA).

[0060] Example sample test:

[0061] Drug loading analysis: The prepared Drugs@PDA of Comparative Example 1 was freeze-dried, three portions of a certain amount of Drugs@PDA nanoparticle freeze-dried powder were accurately weighed and placed in centrifuge tubes, a certain amount of deionized water was added, and the solution was fully dissolved in an ultrasonic washer, and then the solution was transferred to a 10mL volumetric flask to make up the volume, 1mL was taken out, and a 0.22μm organic nylon filter was passed, and the subsequent filtrate was taken as a test sample. The drug content in the subsequent filtrate was detected by HPLC, and the drug loading of the Drugs@PDA nanoparticle preparation was calculated. The concentration of piracetam was quantitatively analyzed using a DIONEX Ultimate 3000 high-performance liquid chromatography system (ThermoFisherScientific, Carlsbad, USA) equipped with a C18 reverse phase chromatographic column (4.6mm×250mm, 5μm). The mobile phase was a mixture of water and methanol (90:10, v / v), the flow rate was 0.8 mL / min, the detection temperature was 40°C, the injection volume was 20 μL, and the detection wavelength of piracetam was 210 nm. The concentration of galanthamine hydrobromide was quantitatively analyzed using a DIONEX Ultimate 3000 HPLC system (Thermo Fisher Scientific, Carlsbad, USA) equipped with a C18 reversed-phase column (4.6 mm×250 mm, 5 μm). The mobile phase was a mixture of triethylamine phosphate buffer and methanol (75:25, v / v) (triethylamine phosphate buffer: take 7 mL of triethylamine, add 900 mL of deionized water, adjust the pH to 6.0 with 0.5 mol / L phosphoric acid solution, and add deionized water to 1000 mL), the flow rate was 1.0 mL / min, the detection temperature was 35°C, the injection volume was 20 μL, and the detection wavelength of galanthamine hydrobromide was 228 nm.

[0062] The results show that the drug loading of piracetam is between 35-45%, and the drug loading of galantamine hydrobromide is between 15-25%.

[0063] Anti-inflammatory effect test: Flow cytometry was used to detect the phenotype of the drug-intervention cell line BMDM. The culture medium was DMEM basic culture medium containing 10% fetal bovine serum and 1% penicillin / streptomycin. Set up blank group Control group, LPS group, Drugs@PDA group and PDA group. The cells were cultured in a sterile six-well plate, and lipopolysaccharide (LPS) (5μg / ml) was added for 12h before adding drugs, and incubated together for 24h at 37℃, 5% CO2 environment. The supernatant was collected and divided into -80℃ refrigerator for storage for subsequent ELISA detection (TNF-α, IL-1β, IL-6). The cells were washed three times with PBS and then digested with trypsin, collected by centrifugation, and antibodies (CD86-PE and CD206-APC) were added. After centrifugation and washing, PBS was added to resuspend and test on the machine.

[0064] The results are as follows Figure 3 As shown in a, the proportion of M2 in the Drugs@PDA group and the PDANPs group was significantly higher than that in the non-PDA Blankcontrol and LPS groups, and there was a significant difference. Figure 3 As shown in Figures b to 3d, compared with the blank control group, the various indicators (TNF-α, IL-1β, IL-6) of the LPS group were significantly upregulated, while the groups added with Drugs@PDA and PDA showed different degrees of downregulation, among which the Drugs@PDA group showed the most obvious downregulation and there was a significant difference between the other groups. It can be seen that the Drugs@PDA and PDA of the present invention both have anti-inflammatory effects, and the combination of drugs and PDA can produce a synergistic effect.

[0065] Oral efficacy test of dextran sulfate sodium salt (DSS) induced colitis in mice: 6-8 week old Balb / c female mice were taken and an inflammatory bowel disease model was constructed by letting them drink 3.5% DSS (MW36-50KD) on their own. A total of six groups were divided: blank control group, DSS control group, DSS+PDA group, DSS+PDA+Drugs Mixture group, DSS+Drugs@PDA group and DSS+Drugs@PDA+Pectin group. Daily gavage began on the eighth day of mouse modeling, and the experimental period was 14 days. During the experiment, the weight of the mice was tested and recorded every day. The types and numbers of bacterial flora in the mouse intestine were calculated. The abundance of neurons in the mouse intestine was observed.

[0066] The results are as follows Figure 4As shown in the figure, compared with the control group, the body weight of the DSS control group decreased significantly, while the weight loss trend of the other drug-treated groups slowed down, among which the decline trend of the Drugs@PDA+Pectin group was the slowest. By analyzing and comparing the colon length between each group, the results are as follows Figure 5 As shown, compared with the Blank control group, the colon length of the DSScontrol group was significantly shortened, while the other drug-administered groups were relieved by drug intervention, among which the therapeutic effect of the Drugs@PDA+Pectin group was the most obvious, and there was a significant difference with other treatment groups. The above results show that the piracetam and galanthamine hydrobromide of the present invention, combined with hollow mesoporous polydopamine, have therapeutic effects on DSS-induced colitis in mice. Among them, the weight of the DSS+Drugs@PDA group and the DSS+Drugs@PDA+Pectin group, which are loaded with drugs and hollow mesoporous polydopamine, showed an early decline in the degree of body weight that was only slightly higher than that of the DSS+PDA+Drugs Mixture group, but a significant upward trend was generated in the later period, and the final weight increase result was significantly better than that of the DSS+PDA+Drugs Mixture group, which was simply mixed and had no upward trend, indicating that the inclusion treatment of piracetam and galanthamine hydrobromide did not significantly affect the onset time of the drug, and could effectively treat the colitis of mice while achieving sustained release. Combination Figure 4 and Figure 5 It can be seen that the weight gain of the DSS+Drugs@PDA+Pectin group was much higher than that of the other groups except the blank group, and the colon length was even longer than that of the blank group, indicating that the combination of Drugs@PDA and pectin can make the drugs have a targeted effect, which can better exert the therapeutic effect of DSS-induced colitis in mice and help repair the colon of mice.

[0067] Figure 6 The results of the microbial study showed that the number of Bacteroides in the intestines of DSS-modeled IBD mice was significantly increased compared with the normal group, while the number of Firmicutes was significantly reduced. The empty PDA group and the empty PDA and drug physical mixture group can significantly increase the number of Firmicutes and reduce the number of Bacteroides, and the double-drug-loaded PDA group also has a better improvement effect. Among them, the treatment effect of the pectin-coated DSS+Drugs@PDA+Pectin group was the most significant, and the number of Bacteroides and Firmicutes in the intestines of mice returned to the level of microbial distribution in the intestines of normal mice. Figure 7 The Venn diagram results of mouse intestinal flora showed that the types of flora in the intestines of pectin-coated DSS+Drugs@PDA+Pectin mice were the most abundant, significantly higher than those in the IBD mouse group modeled by DSS, and also significantly higher than those in the normal group of mice.

[0068] Effects of different treatments on the abundance of enteric neurons in IBD mice Figure 8 As shown in the figure, the neuron abundance in the DSS+Drugs@PDA+Pectin group is much higher than that in the other groups, indicating that the method of the present invention using hollow mesoporous polydopamine and fructose to continue treating the drugs can synergistically improve the drug's ability to repair mouse intestinal neurons.

[0069] For those skilled in the art, various other corresponding changes and deformations can be made according to the technical solutions and concepts described above, and all of these changes and deformations should fall within the protection scope of the claims of the present invention.

Claims

1. A drug delivery system based on hollow mesoporous polydopamine nanoparticles, characterized in that: It comprises a drug-loaded precursor formed by hollow mesoporous polydopamine nanoparticles and drugs loaded inside and / or outside the hollow mesoporous polydopamine nanoparticles, and an enteric material wrapped outside the drug-loaded precursor; the drugs include drugs with neuroprotective effects and acetylcholinesterase inhibitors.

2. The drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 1, characterized in that: The drug with neuroprotective effect is piracetam; the acetylcholinesterase inhibitor is galantamine hydrobromide; and the enteric material includes one of pectin, shellac, polyvinyl alcohol acetate phthalate, acrylic resin, cellulose or chitosan.

3. The drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 1, characterized in that: The drug loading of the drug with neuroprotective effect is 35-45%; the drug loading of the acetylcholinesterase inhibitor is 15-25%.

4. The drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 1, characterized in that: The average particle size of the drug-loaded precursor is 300-1000 nm.

5. A method for preparing a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 1, characterized in that: The following steps are involved: a. Adding polyoxyethylene-polyoxypropylene-polyoxyethylene to a mixed solution of water and ethanol under stirring and ultrasonic conditions, dispersing the mixture uniformly to obtain a polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion; then, adding 3,3',5,5'-tetramethylbenzidine to the dispersion, mixing for 1 to 3 hours to obtain a first mixed system; adding tris(hydroxymethyl)aminomethane hydrochloride) solution and dopamine hydrochloride solution to the first mixed system, continuing to stir and react, to obtain a mixed solution; after centrifugation and washing, collecting and obtaining hollow mesoporous polydopamine nanoparticles; b, the hollow mesoporous polydopamine nanoparticles are fully dispersed in the saturated solution of the drug, and fully mixed to obtain a mixed solution; the solid is separated, collected and freeze-dried to obtain a drug-loaded precursor; c. Prepare a drug-loaded precursor dispersion, then add the enteric material solution into the drug-loaded precursor dispersion, and mix thoroughly to obtain the product.

6. The method for preparing a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 5, characterized in that: Step a: The concentration of the polyoxyethylene-polyoxypropylene-polyoxyethylene in the mixed solution of water and ethanol is 2-10 mg / mL; the volume ratio of water to ethanol solution in the mixed solution of water and ethanol is 1:0.25-4; the volume ratio of the polyoxyethylene-polyoxypropylene-polyoxyethylene dispersion to 3,3',5,5'-tetramethylbenzidine is 80-120:1; the concentration of the tris(hydroxymethyl)aminomethane hydrochloride) solution is 40-50 mg / mL; the concentration of the dopamine hydrochloride solution is 5-10 mg / mL; the volume ratio of the tris(hydroxymethyl)aminomethane hydrochloride solution to the dopamine hydrochloride solution is 1:1; and the volume ratio of the first mixed system to the dopamine hydrochloride solution is 12-15:

1.

7. The method for preparing a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 5, characterized in that: The stirring speed of step a is 400-500 rpm, the ultrasonic frequency is 30-40 kHz, and the ultrasonic power is 200-300 W; the stirring reaction time of step a is 18-24 h; the washing liquid of the centrifugal washing in step a is a mixture of acetone and ethanol in a volume ratio of 1:2; the centrifugal condition is 13000 rpm, 10 min, and the hollow mesoporous polydopamine nanoparticles are collected after two centrifugal washings.

8. The method for preparing a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 5, characterized in that: The stirring speed of the mixing in step b is 300-800 rpm, and the mixing time is 20-28 h; the centrifugal conditions for the separation in step b are 13000 rpm, 10 min; the separation in step b is centrifugation once to collect the precipitate; the freeze-drying conditions in step b are freeze-drying at -80°C and 2 Pa.

9. The method for preparing a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to claim 5, characterized in that: The concentration of the drug-loaded precursor aqueous dispersion in step c is 2-20 mg / mL; the concentration of the enteric material in step c is 2-20 mg / mL; the volume ratio of the drug-loaded precursor dispersion to the enteric material solution is 1:0.25-4.

10. Use of a drug delivery system based on hollow mesoporous polydopamine nanoparticles according to any one of claims 1 to 4 in the preparation of a drug for treating inflammatory bowel disease, characterized in that: Maintain the neural abundance of damaged intestinal tissue and restore the neuroimmune function within the diseased intestinal tissue.

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