Hydrogel microsphere with core-shell structure as well as preparation method and application of hydrogel microsphere

Abstract

The invention discloses a hydrogel microsphere with a core-shell structure, the hydrogel microsphere comprises a sodium hyaluronate hydrogel core and a sodium alginate shell, and the sodium hyaluronate hydrogel core also comprises a slow release factor and divalent metal ions. The hydrogel microspheres with the sodium hyaluronate hydrogel as the inner core and the alginic acid as the shell can realize accurate targeted blasting in the intestinal tract, and can be adhered to the local part of the intestinal wall to induce benign intestinal flora to treat the inflammatory bowel disease, so that the phenomenon that when the medicine arrives at the intestinal tract, the medicine cannot be damaged is avoided; due to peristalsis of the intestinal tract, the medicine can not be accurately released to a specific position. The mass ratio of sodium hyaluronate to sodium alginate is controlled to be (4-20): 1, and the explosion speed of the microspheres is controlled, so that the release speed and the position accuracy of the medicine are controlled, the problem that the blood concentration of the medicine cannot be effectively controlled due to the fact that the release speed of the medicine is too high or too low is solved, the curative effect of the medicine is improved, and side effects are reduced.

Description

Technical Field

[0001] The present invention relates to the field of medical preparations, and in particular to a hydrogel microsphere with a core-shell structure and a preparation method and application thereof. Background Art

[0002] Inflammatory bowel disease is a lifelong intestinal inflammation caused by damage to epithelial cells and related ileum and colorectal mucosa. Existing drugs have certain efficacy in the short term, but long-term use has certain side effects and creates a metabolic burden on the human body. Quercetin has certain efficacy for inflammatory bowel disease, but it will be metabolized in large quantities in the intestine. Therefore, developing a system that can deliver quercetin in a targeted manner has excellent therapeutic potential for the treatment of inflammatory bowel disease.

[0003] Chinese invention patent CN118236352A discloses a gel microsphere containing drug-loaded microspheres in the inner core and a preparation method thereof. The gel microspheres include drug-loaded microspheres and a gel coating layer coated on the outside of the drug-loaded microspheres, thereby forming a ball-in-ball structure. It solves the problems of low drug loading and low encapsulation rate, fast drug release rate and poor sustained release effect of ordinary drug-loaded microspheres, but it adopts injection administration and cannot be used for long-term oral treatment. Chinese invention patent CN113041232B discloses an oral colon-targeted hydrogel with targeted intestinal explosion and its preparation method and application. It uses thiolated hyaluronic acid and silver ions to cross-link, and uses airflow control technology to prepare HA-SH-Ag hydrogel microspheres containing calcium ions. After oral administration, it can accurately "explode" and release hydrogel in the colon. It can regulate the polarization of intestinal macrophages and induce benign intestinal flora composition by adhering to the intestinal wall to treat IBD, but the preparation process is cumbersome and the cost is high. Summary of the invention

[0004] Sodium alginate is stable in the gastrointestinal tract, but is biodegradable in the colonic microenvironment under the action of colonic flora and enzymes, and can interact with divalent cations to form a gel structure. Sodium alginate gel is less sensitive to the acidic pH of the stomach (pH 1-2) and can be used to reduce drug release and degradation in the gastrointestinal tract. As the pH in the intestine increases, the sodium alginate gel swells and causes drug release to improve IBD treatment.

[0005] In the intestine, under the action of intestinal fluid, the outer shell of sodium alginate will gradually dissolve, causing the intestinal bursting microspheres with a core-shell structure to rupture, thereby releasing the sodium hyaluronate core containing quercetin and divalent metal ions. Since the sodium hyaluronate core has good biocompatibility and degradability in the intestine, it can effectively reduce the side effects of drugs. Through this method, the precise controlled release of drugs in the intestine can be achieved, the absorption efficiency and efficacy of drugs can be improved, and the side effects of drugs can be reduced.

[0006] In order to develop a system for targeted delivery of quercetin, the first aspect of the present invention provides a core-shell structured hydrogel microsphere comprising a sodium hyaluronate hydrogel core and a sodium alginate shell, wherein the sodium hyaluronate hydrogel core also includes a sustained release factor and a divalent metal ion.

[0007] As a preferred embodiment, the sodium hyaluronate hydrogel core is prepared by emulsifying the aqueous phase and the oil phase and adding a cross-linking agent to undergo a cross-linking reaction, wherein the aqueous phase includes a sustained-release factor, sodium hyaluronate and an alkaline aqueous solution, and the oil phase includes a surfactant and liquid paraffin.

[0008] As a preferred embodiment, the volume ratio of the water phase to the oil phase is 1:(6-14).

[0009] As a preferred embodiment, the volume ratio of the water phase to the oil phase is 1:10.

[0010] The inventors found during the experiment that when the water-oil ratio of the prepared sodium hyaluronate hydrogel was 1:10, the hydrogel formed had a rounded morphology, uniform particle size, and the obtained hydrogel microspheres had consistent efficacy, which was suitable for actual production. The reason may be that when the water-oil ratio was 1:10, the microspheres formed under stirring had a good emulsification effect, the internal cross-linked structure of the microspheres was clear, no flocs were produced, the structure was transparent and the morphology was good. If the proportion of the oil phase increased, the volume of the hydrogel would be small, and in the subsequent washing process, the loss would be large and the yield would be low.

[0011] As a preferred embodiment, the average molecular weight of the sodium hyaluronate is 100,000-3,000,000 Da.

[0012] As a preferred embodiment, the average molecular weight of the sodium hyaluronate is 200,000 Da.

[0013] As a preferred embodiment, the mass fraction of the surfactant in the liquid paraffin is 1-5%.

[0014] As a preferred embodiment, the mass fraction of the surfactant in the liquid paraffin is 3%.

[0015] As a preferred embodiment, the surfactant is a nonionic surfactant, and the nonionic surfactant is selected from one of Span and Tween.

[0016] As a preferred embodiment, the crosslinking agent is selected from at least one of 1,4-butanediol diglycidyl ether, divinyl sulfone, pentaerythritol tetrakis-(3-mercaptopropionate), and trimethylolpropane triacrylate.

[0017] As a preferred embodiment, the cross-linking agent is 1,4-butanediol diglycidyl ether.

[0018] As a preferred embodiment, the mass ratio of the cross-linking agent to sodium hyaluronate is (0.5-2):1.

[0019] As a preferred embodiment, the mass ratio of the cross-linking agent to sodium hyaluronate is 0.5:1.

[0020] As a preferred embodiment, the temperature of the cross-linking reaction is 35-70°C.

[0021] As a preferred embodiment, the temperature of the cross-linking reaction is 40-70°C.

[0022] As a preferred embodiment, the temperature of the cross-linking reaction is 50°C.

[0023] As a preferred embodiment, the mass concentration of the alkaline aqueous solution is 1%-12%.

[0024] As a preferred implementation, the mass concentration of the alkaline aqueous solution is 1%.

[0025] The second aspect of the present invention provides a method for preparing hydrogel microspheres with a core-shell structure, comprising the following steps:

[0026] S1: adding a sustained-release factor to an alkaline aqueous solution containing sodium hyaluronate to obtain an aqueous phase, dissolving a surfactant in liquid paraffin to obtain an oil phase, dropping the aqueous phase into the oil phase under stirring for emulsification, adding a cross-linking agent for cross-linking reaction, and washing to obtain a sodium hyaluronate hydrogel;

[0027] S2: immersing the sodium hyaluronate hydrogel in a divalent metal ion aqueous solution to obtain a sodium hyaluronate hydrogel core;

[0028] S3 prepares sodium alginate aqueous solution;

[0029] S4 adds the sodium hyaluronate hydrogel core dropwise into the sodium alginate aqueous solution to form a sodium alginate shell, thereby obtaining hydrogel microspheres with a core-shell structure.

[0030] As a preferred embodiment, the divalent metal ion is selected from at least one of calcium ion, iron ion, magnesium ion, barium ion, zinc ion and copper ion.

[0031] As a preferred embodiment, the divalent metal ion is a divalent calcium ion or a divalent magnesium ion.

[0032] As a preferred embodiment, the mass concentration of the divalent calcium ion aqueous solution is 5-15%; preferably, the mass concentration of the divalent calcium ion aqueous solution is 10%.

[0033] As a preferred embodiment, the mass fraction of the sodium alginate aqueous solution is 0.1%-1%.

[0034] As a preferred embodiment, the mass fraction of the sodium alginate aqueous solution is 0.1%-0.5%.

[0035] As a preferred embodiment, the mass ratio of sodium hyaluronate to sodium alginate is (4-20):1.

[0036] As a preferred embodiment, the mass ratio of sodium hyaluronate to sodium alginate is (5-10):1.

[0037] As a preferred embodiment, the mass ratio of sodium hyaluronate to sodium alginate is 6.7:1.

[0038] The inventors found during the experiment that the prepared hydrogel microspheres with sodium hyaluronate hydrogel as the core and alginate as the shell can achieve accurate targeted explosion in the intestine, and treat inflammatory bowel disease by adhering to the local intestinal wall to induce benign intestinal flora. The possible reason is speculated to be: using sodium hyaluronate and sodium alginate with a mass fraction of 0.1%-0.5% at a weight ratio of (6-7): 1, the drug can be accurately released in the intestine. Under the action of intestinal fluid in the intestine, the sodium alginate shell will gradually dissolve, causing the core-shell structure to explode in the intestine, releasing quercetin and the core of the sodium hyaluronate hydrogel. If the proportion of sodium hyaluronate is too large, the hydrogel microspheres cannot be released at the lesion site, and the drug release rate is too fast, resulting in a sharp increase in the blood concentration of the drug. When the mass proportion of sodium alginate is too large, the drug release rate will be too slow. At the same time, too much sodium alginate will also affect the morphology of the hydrogel microspheres, and a microsphere structure with uniform morphology and sufficient quercetin cannot be obtained.

[0039] The third aspect of the present invention provides an application of hydrogel microspheres with a core-shell structure for preparing drugs for inflammatory bowel disease.

[0040] Compared with the prior art, the present invention has the following beneficial effects:

[0041] (1) The core-shell hydrogel microspheres of the present invention, which are prepared with sodium hyaluronate hydrogel as the core and alginate as the shell, can achieve accurate targeted explosion in the intestine, and can treat inflammatory bowel disease by adhering to the local intestinal wall to induce benign intestinal flora, thereby avoiding the problem that the drug cannot be accurately released to a specific location due to intestinal peristalsis when it reaches the intestine.

[0042] (2) The core-shell structure hydrogel microspheres of the present invention, when the water-to-oil ratio of the prepared sodium hyaluronate hydrogel is 1:10, the hydrogel formed has a rounded morphology and uniform particle size. The obtained hydrogel microspheres have consistent efficacy and are suitable for actual production.

[0043] (3) The core-shell hydrogel microspheres of the present invention control the mass ratio of sodium hyaluronate to sodium alginate to (4-20):1, thereby controlling the blasting speed of the microspheres and thus controlling the release speed and position accuracy of the drug, thereby avoiding the problem of the drug release speed being too fast or too slow and being unable to effectively control the blood drug concentration, thereby improving the efficacy of the drug and reducing side effects.

[0044] (4) The core-shell structured hydrogel microspheres of the present invention include a sodium hyaluronate hydrogel core and a sodium alginate shell. The sodium hyaluronate hydrogel core includes quercetin, which can protect the drug from degradation and prolong the circulation time of the drug in the body, thereby improving the absorption efficiency and efficacy of the drug.

[0045] (5) The core-shell hydrogel microspheres of the present invention are made of materials with good biocompatibility, are non-toxic and harmless to the human body, and have good biosafety. At the same time, the preparation method is simple and easy to industrialize, which is conducive to popularization and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 This is a morphology picture of the inner core of the sodium hyaluronate hydrogel prepared in Example 11;

[0047] Figure 2 A schematic diagram of the core-shell structure of the core-shell hydrogel microspheres prepared in Example 13;

[0048] In the figure: 1. Sodium hyaluronate hydrogel core; 2. Sodium alginate shell.

[0049] Figure 3 This is a morphology picture of the core-shell structured hydrogel microspheres prepared in Example 12;

[0050] Figure 4 This is a morphology picture of the core-shell structured hydrogel microspheres prepared in Example 13;

[0051] Figure 5 This is a morphology image of the core-shell structured hydrogel microspheres prepared in Example 14;

[0052] Figure 6 This is the in vitro release curve of the core-shell hydrogel microspheres prepared in Example 13.

[0053] In the figure: A: ultrapure water; B: simulated gastric fluid; C: simulated intestinal fluid. DETAILED DESCRIPTION

[0054] Example A

[0055] Example A provides a method for preparing a sodium hyaluronate hydrogel core, comprising the following steps:

[0056] 0.6 g of sustained-release factor was added to an alkaline aqueous solution containing 4 g of sodium hyaluronate to obtain an aqueous phase, 3 wt % of a surfactant was dissolved in 200 mL of liquid paraffin to obtain an oil phase, and the aqueous phase was added dropwise to the oil phase under stirring for emulsification, 2 g of a cross-linking agent was added to react and cross-link for 4 hours, and the sodium hyaluronate hydrogel was obtained after washing with clean water.

[0057] The sustained-release factor is quercetin; the average molecular weight of the sodium hyaluronate solution is 2.6 million Da, the surfactant is Span 80; the cross-linking agent is 1,4-butanediol diglycidyl ether; and the cross-linking temperature is 35°C.

[0058] The alkaline aqueous solution includes 2 g of sodium hydroxide and 40 g of ultrapure water. Sodium hyaluronate is added to the alkaline aqueous solution with continuous stirring.

[0059] Results: Sodium hyaluronate was not completely transformed into gel microspheres, and some floccules appeared.

[0060] Example B

[0061] Example 1

[0062] A method for preparing a sodium hyaluronate hydrogel core, the specific implementation method is the same as that of Example A, except that the temperature of the cross-linking reaction is 45°C.

[0063] Example 2

[0064] A method for preparing a sodium hyaluronate hydrogel core, the specific implementation method is the same as that of Example A, except that the temperature of the cross-linking reaction is 50°C.

[0065] Example 3

[0066] A method for preparing a sodium hyaluronate hydrogel core, the specific implementation method is the same as that of Example A, except that the temperature of the cross-linking reaction is 70°C.

[0067] Results: The inner core of the sodium hyaluronate hydrogel of Example 1 was not completely spherical and had flocs; the inner core of the sodium hyaluronate hydrogel of Example 2 had round microspheres and flocs; the inner core microspheres of the sodium hyaluronate hydrogel of Example 3 were not round and had flocs.

[0068] Conclusion: At different cross-linking temperatures, the viscoelasticity of the prepared microspheres is different, and the morphology and water absorption of the microspheres are quite different. When the cross-linking temperature is 50℃, the morphology of the prepared microspheres is round.

[0069] Example C

[0070] Example 4

[0071] A method for preparing a sodium hyaluronate hydrogel core, the specific implementation method is the same as Example 2, except that the mass ratio of the cross-linking agent to sodium hyaluronate is 1:1; and the water-to-oil ratio is 1:5.

[0072] Example 5

[0073] A method for preparing a sodium hyaluronate hydrogel inner core, the specific implementation method is the same as Example 4, except that the water-to-oil ratio is 1:15.

[0074] Example 6

[0075] A method for preparing a sodium hyaluronate hydrogel inner core, the specific implementation method is the same as Example 4, except that the water-to-oil ratio is 1:10.

[0076] Results: The sodium hyaluronate hydrogel inner core microspheres of Example 4 were round and had flocs; the sodium hyaluronate hydrogel inner core microspheres of Example 5 had good morphology and no flocs; the sodium hyaluronate hydrogel inner core microspheres of Example 6 had good morphology and no flocs.

[0077] Conclusion: Increasing the proportion of oil phase can significantly improve the morphology of microspheres. When the water-oil volume ratio is 1:10, the prepared microspheres have round morphology and uniform particle size. However, when the volume of oil phase is further increased, the particle size of microspheres is small, and more losses occur during the washing process, resulting in low yield.

[0078] Example D

[0079] Example 7

[0080] A method for preparing a sodium hyaluronate hydrogel inner core, with specific parameters as shown in Table 1, and other parameters are the same as in Example 2.

[0081] Table 1

[0082]

[0083]

[0084] Conclusion: When the molecular weight of sodium hyaluronate is 200,000, the mass ratio of crosslinker to sodium hyaluronate is 1:2, the mass percentage of sodium hydroxide is 1%, the water-oil ratio is 1:10, and the reaction temperature is 50°C, the prepared microspheres have a rounded morphology and uniform particle size, and the drug loading rate of quercetin is 15%. (Drug loading rate of quercetin = quercetin mass / sodium hyaluronate mass × 100%)

[0085] Example E

[0086] A method for preparing hydrogel microspheres with a core-shell structure comprises the following steps:

[0087] S1: 0.3 g of sustained-release factor is added to an alkaline aqueous solution containing 2 g of sodium hyaluronate to obtain an aqueous phase, 3 wt % of a surfactant is dissolved in 200 mL of liquid paraffin to obtain an oil phase, the aqueous phase is added dropwise to the oil phase under stirring for emulsification, 1 g of a cross-linking agent is added, cross-linked at 50° C. for 4 h, and the sodium hyaluronate hydrogel is obtained after washing;

[0088] S2: Soaking the sodium hyaluronate hydrogel in 20 mL of a divalent metal ion aqueous solution to obtain a sodium hyaluronate hydrogel core;

[0089] S3 prepares sodium alginate aqueous solution;

[0090] S4 adds the sodium hyaluronate hydrogel inner core dropwise into 100 mL of sodium alginate aqueous solution. The calcium ions in the microspheres gradually diffuse to the surface of the microspheres and cross-link the sodium alginate to form a calcium alginate hydrogel outer shell, thereby obtaining hydrogel microspheres with a core-shell structure.

[0091] The sustained-release factor is quercetin; the average molecular weight of the sodium hyaluronate solution is 200,000 Da; the surfactant is Span 80; and the cross-linking agent is 1,4-butanediol diglycidyl ether.

[0092] The alkaline aqueous solution includes 0.2 g of sodium hydroxide and 20 g of ultrapure water. Sodium hyaluronate is added to the alkaline aqueous solution with continuous stirring.

[0093] The divalent metal ion aqueous solution is a divalent calcium ion aqueous solution with a mass concentration of 10%.

[0094] The mass fraction of the sodium alginate aqueous solution is 1%.

[0095] Results: The morphology of the microspheres was observed. Due to the high concentration of sodium alginate, the microspheres aggregated in the sodium alginate aqueous solution.

[0096] Example F

[0097] Example 12

[0098] A method for preparing hydrogel microspheres with a core-shell structure, the specific implementation method is the same as that of Example E, except that the mass fraction of the sodium alginate aqueous solution is 0.1%, and the mass ratio of sodium hyaluronate to sodium alginate is 20:1.

[0099] Embodiment 13

[0100] A method for preparing hydrogel microspheres with a core-shell structure, the specific implementation method is the same as that of Example E, except that the mass fraction of the sodium alginate aqueous solution is 0.3%, and the mass ratio of sodium hyaluronate to sodium alginate is 6.7:1.

[0101] Embodiment 14

[0102] A method for preparing hydrogel microspheres with a core-shell structure, the specific implementation method is the same as that of Example E, except that the mass fraction of the sodium alginate aqueous solution is 0.5%, and the mass ratio of sodium hyaluronate to sodium alginate is 4:1.

[0103] Results: The shell of the core-shell hydrogel microspheres of Example 12 was thin; the core-shell structure of the core-shell hydrogel microspheres of Example 13 was uniform; and the microspheres of the core-shell hydrogel microspheres of Example 14 were agglomerated.

[0104] Conclusion: When the concentration of sodium alginate is higher than 0.5%, the core-shell microspheres are prone to agglomeration. When the concentration of sodium alginate is lower than 0.1%, the shell wall of the core-shell microspheres is thinner.

[0105] Performance Testing

[0106] 1. Morphology test: The morphology of the sodium hyaluronate hydrogel core and the core-shell structure hydrogel microspheres prepared in each example was observed using a WD300-48LT biological microscope. The morphology of Example 11 is shown in Figure 1 , the morphology of Examples 12-14 is shown in Figure 3-5 The structural diagram of Example 13 is shown in Figure 2 .

[0107] 2. Drug loading rate test: The test results were measured by UV-visible spectrophotometer at a wavelength of 357 nm. Drug loading rate = (the measured drug mass in the microspheres / the total mass of the microspheres) × 100%.

[0108] 3. In vitro release test: Take 0.5 g of the dried microspheres of Example 13, add 7.5 mL of ultrapure water, simulated gastric fluid, and simulated colon fluid respectively, and place them in a shaker, take the supernatant at 0.5 h, 1 h, 2 h, 3 h, 4 h, 5 h, 20 h, and 22 h, and test the drug concentration by UV.

[0109] The cumulative dissolution rate after each time point (%) = [(C1 + C2 + ... Cn) × V] / L × 100%, where Cn is the drug concentration after each time point, V is the fixed sampling volume at each time point (because all the drugs are taken out and then supplemented with the same volume of medium, the sampling volume is equivalent to the dissolution medium volume), and L is the total drug mass input. The test results are shown in Table 2.

[0110] Table 2

[0111]

Claims

1. A core-shell hydrogel microsphere, characterized in that: It comprises a sodium hyaluronate hydrogel core and a sodium alginate shell, wherein the sodium hyaluronate hydrogel core also comprises a sustained-release factor and a divalent metal ion.

2. The core-shell hydrogel microsphere according to claim 1, characterized in that: The sodium hyaluronate hydrogel core is prepared by emulsifying the water phase and the oil phase and adding a cross-linking agent to undergo a cross-linking reaction. The water phase includes a sustained-release factor, sodium hyaluronate and an alkaline aqueous solution, and the oil phase includes a surfactant and liquid paraffin.

3. The core-shell hydrogel microspheres according to claim 2, characterized in that: The volume ratio of the water phase to the oil phase is 1:(6-14).

4. The core-shell hydrogel microspheres according to claim 2, characterized in that: The crosslinking agent is selected from at least one of 1,4-butanediol diglycidyl ether, divinyl sulfone, pentaerythritol tetrakis-(3-mercaptopropionate), and trimethylolpropane triacrylate.

5. The core-shell hydrogel microsphere according to claim 2, characterized in that: The mass ratio of the cross-linking agent to sodium hyaluronate is (0.5-2):

1.

6. The core-shell hydrogel microspheres according to claim 2, characterized in that: The temperature of the cross-linking reaction is 35-70°C.

7. The core-shell hydrogel microspheres according to claim 2, characterized in that: The mass concentration of the alkaline aqueous solution is 1%-12%.

8. A method for preparing the core-shell hydrogel microspheres according to any one of claims 2 to 7, characterized in that: The following steps are involved: S1: adding a sustained-release factor to an alkaline aqueous solution containing sodium hyaluronate to obtain an aqueous phase, dissolving a surfactant in liquid paraffin to obtain an oil phase, dropping the aqueous phase into the oil phase under stirring for emulsification, adding a cross-linking agent for cross-linking reaction, and washing to obtain a sodium hyaluronate hydrogel; S2: immersing the sodium hyaluronate hydrogel in a divalent metal ion aqueous solution to obtain a sodium hyaluronate hydrogel core; S3 prepares sodium alginate aqueous solution; S4 adds the sodium hyaluronate hydrogel core dropwise into the sodium alginate aqueous solution to form a sodium alginate shell, thereby obtaining hydrogel microspheres with a core-shell structure.

9. The method for preparing the core-shell structured hydrogel microspheres according to claim 8, characterized in that: The mass ratio of sodium hyaluronate to sodium alginate is (4-20):

1.

10. An application of the core-shell structured hydrogel microspheres according to any one of claims 2 to 7, characterized in that: The invention is used in preparing medicines for inflammatory bowel disease.

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