Minocycline hydrochloride-melatonin compound colon targeting capsule and preparation method thereof

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a minocycline hydrochloride-melatonin compound colon targeting capsule and a preparation method thereof. The capsule comprises a pellet and a capsule shell, wherein the pellet comprises a medicine core and a colon-targeted sustained-release layer wrapping the medicine core. The medicine core comprises active ingredients of minocycline hydrochloride and melatonin; the colon-targeted sustained-release layer comprises a special pH-enzyme colon-targeted response agent and a self-made plasticizer. The pH-enzyme responsive agent is based on a hydroxypropyl methylcellulose grafted vanillin derivative, and can be degraded in response to pH change and azo reductase at the colon part; the plasticizer has good compatibility with a coating material, and can effectively reduce leakage of drugs at the front ends of the stomach and intestines. The capsule prepared by the invention can obviously improve gastrointestinal stimulation of minocycline hydrochloride, realizes targeted release of the medicine at the colon part, and shows an excellent treatment effect on ulcerative colitis while reducing the dosage of antibiotics by utilizing the synergistic effect of melatonin.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of pharmaceutical preparations, and particularly relates to a minocycline hydrochloride-melatonin compound colon-targeting capsule and a preparation method thereof. BACKGROUND

[0002] Minocycline hydrochloride is a semisynthetic tetracycline antibiotic with a broad spectrum of activity. Its hydrochloride salt is a yellow crystalline powder with a bitter taste, easy to absorb moisture and degrade under light. It is easily soluble in alkali hydroxide or carbonate solution, slightly soluble in ethanol and water. The antibacterial activity of the product is the highest in the tetracycline family, and it still has a killing effect on tetracycline-resistant Staphylococcus aureus, Streptococcus and Escherichia coli, and the relative low drug resistance mutation selection pressure.

[0003] Minocycline hydrochloride is rapidly and completely absorbed after oral administration, has strong tissue penetration, and can be widely distributed in various organs and body fluids of the body, especially easily penetrates the blood-brain barrier and reaches a high concentration in the cerebrospinal fluid. Long-term clinical application has confirmed that the main adverse reactions include gastrointestinal irritation symptoms (such as nausea, vomiting, abdominal pain, diarrhea) and esophageal ulcers. Among the above adverse reactions, gastrointestinal discomfort is mainly caused by direct irritation of the drug to the upper gastrointestinal mucosa and imbalance of the small intestinal flora, and esophageal ulcers are directly related to the local disintegration of the capsule in the esophagus.

[0004] Melatonin can synchronize circadian rhythms, shorten sleep latency, and improve shift work, jet lag, and primary insomnia. It has antioxidant and immunomodulatory effects, but needs to be used in low doses and for a short period of time, and long-term and large amounts of endocrine suppression. Patent CN117643569A discloses that melatonin and minocycline hydrochloride synergistically improve the success rate of periodontitis treatment. However, the effect of the synergistic action of melatonin and minocycline hydrochloride on colitis has not been further studied.

[0005] Therefore, there is an urgent need for a minocycline hydrochloride-melatonin compound colon-targeting capsule that can be targeted to the colon for controlled release, overcome the irritation of minocycline hydrochloride to the stomach, and enhance its colon targeting to exert anti-inflammatory effects. SUMMARY

[0006] In order to overcome the shortcomings of the prior art, the purpose of the present application is to provide a minocycline hydrochloride-melatonin compound colon-targeting capsule and a preparation method thereof.

[0007] The purpose of the present application is achieved by the following technical solutions: A minocycline hydrochloride-melatonin compound colon-targeting capsule, comprising pellets and a capsule shell, wherein the pellets comprise a drug core and a colon-targeting sustained-release layer wrapped outside the drug core, and the mass ratio of the two is 1: (0.1-0.15). The drug core comprises the following raw materials in parts by weight: minocycline hydrochloride 20-40 parts, melatonin 10-20 parts, filler 20-40 parts, adhesive I 4-11 parts; The colon-targeted sustained-release layer comprises the following raw materials in parts by weight: adhesive II 5-15 parts, pH-enzyme colon-targeted response agent 10-20 parts, plasticizer 2-4 parts, light shielding agent 3-5 parts, sustained-release agent 5-20 parts, pore-forming agent 2-10 parts.

[0008] Preferably, the pH-enzyme colon-targeted response agent is prepared by the following preparation process: (1) vanillic acid and potassium carbonate are added to acetonitrile, and 4-bromo-2-methoxy aniline is added, and the reaction is carried out under heating conditions, and the intermediate 1 is obtained by purification treatment; the structure of the intermediate 1 is shown in formula I: ; (2) 4-aminophenyl acrylate ester and the intermediate 1 are added to sodium hypochlorite solution, and hydrochloric acid solution is added to carry out the reaction, and the vanillic acid derivative is obtained by purification treatment; the structure of the vanillic acid derivative is shown in formula II: ; (3) hydroxypropyl methyl cellulose is dissolved in water, and potassium sulfate is added after the temperature is raised to 50-60°C under nitrogen atmosphere to initiate, and then the vanillic acid derivative is added to continue the reaction, and the pH-enzyme colon-targeted response agent is obtained by dialysis and freeze-drying after cooling.

[0009] Preferably, in step (1), the molar ratio of the vanillic acid, potassium carbonate and 4-bromo-2-methoxy aniline is 1:(2.2-2.8):(1.1-1.5); the temperature of the heating is 60-80°C, and the reaction time is 12-18 h.

[0010] Preferably, in step (2), the amount ratio of the intermediate 1, 4-aminophenyl acrylate ester, sodium hypochlorite solution and hydrochloric acid solution is 1 mmol:(1-1.5) mmol:(3.7-4.5) mL:(0.4-0.8) mL; the concentration of the sodium hypochlorite solution is 10 wt%, and the concentration of the hydrochloric acid solution is 2.5 mol / L; the reaction time is 3-5 h.

[0011] Preferably, in step (3), the mass ratio of the hydroxypropyl methyl cellulose, potassium sulfate and vanillic acid derivative is 1:(0.012-0.016):(3.2-4); the initiation time is 12-15 min; the reaction time is 0.5-1 h; and the molecular weight cut-off of the dialysis is 10 kDa.

[0012] Preferably, the plasticizer is prepared by the following preparation process: The phthalic acid, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 4-dimethylaminopyridine and 4-hydroxy-2-pyrrolidinone are added into acetonitrile, reacted under heating condition, and the plasticizer is obtained after purification treatment; the structure of the plasticizer is shown in formula III: .

[0013] Preferably, the molar ratio of the phthalic acid, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 4-dimethylaminopyridine and 4-hydroxy-2-pyrrolidinone is 1: (2.5-2.8): (5-5.6): (2-2.5); the temperature of the heating is 40-60 ℃, and the reaction time is 0.5-1.5 h.

[0014] Preferably, the filler is any one selected from lactose and microcrystalline cellulose; the binder I and the binder II are both povidone K30; the light shielding agent is titanium dioxide; the sustained release agent is any one selected from polyacrylic acid ester and ethyl cellulose; and the pore-forming agent is any one selected from polysorbate and polyethylene glycol 6000.

[0015] A preparation method of the above-mentioned minocycline hydrochloride-melatonin compound colon-targeting capsule, comprising the following steps: (a) preparing a drug core: according to the weight fractions of the raw materials in the drug core, the minocycline hydrochloride, the melatonin, the filler and part of the binder I are mixed to obtain a mixed drug powder; the remaining binder I is dissolved in water to form a binder solution and added to the mixed drug powder, and after stirring, extruding and drying, the drug core is obtained; (b) preparing a pellet: according to the weight fractions of the raw materials in the colon-targeting sustained release layer, the binder II, the pH-enzyme colon-targeting response agent, the plasticizer, the light shielding agent, the sustained release agent and the pore-forming agent are added into an ethanol-water solution to obtain a mixed solution; the mixed solution is wrapped onto the surface of the drug core obtained in step (a) through a fluidized bed bottom spraying process to form a colon-targeting sustained release layer, and after drying, the pellet is obtained; (c) preparing a capsule: the pellet is loaded into a capsule shell to obtain the minocycline hydrochloride-melatonin compound colon-targeting capsule.

[0016] Preferably, the content of the minocycline hydrochloride in each minocycline hydrochloride-melatonin compound colon-targeting capsule is 100-120 mg.

[0017] The present application has the following effects relative to the prior art: 1. The invention is a compound colon-targeted preparation of minocycline hydrochloride and melatonin, which improves the gastrointestinal irritation of minocycline hydrochloride by adding a pH-enzyme colon-targeting response agent and a self-made plasticizer, realizes the targeted release of the drug in the colon, reduces the dosage of antibiotics while reducing the risk of drug resistance, and enhances the therapeutic effect on colitis by utilizing the anti-inflammatory, antioxidant and circadian rhythm regulation effects of melatonin.

[0018] 2. The pH-enzyme colon-targeting response agent of the invention is a graft copolymer obtained by grafting carboxylated vanillic acid derivative side chains to hydroxypropyl methyl cellulose (HPMC) as the main chain. This material has excellent film-forming properties and shows significant swelling differences in different pH environments in the gastrointestinal tract: in the stomach acid environment, the side chain carboxyl group is protonated, the hydrophilicity of the material decreases, and the swelling degree is limited, effectively inhibiting drug release; when reaching the terminal ileum to the ascending colon, the carboxyl group deprotonates to produce electrostatic repulsion, and the copolymer rapidly swells to form a continuous diffusion channel, realizing drug release. Further, the diazo groups introduced into the side chain by azoization can be specifically broken by azo reductase secreted by anaerobic bacteria in the colon, realizing pH-enzyme dual triggering and enhancing colon targeting. In addition, the ester groups of the graft side chains are similar to the ester groups in the plasticizer, which can reduce drug leakage in the acid stage through dipole-dipole interaction; the benzyl ether group has a hydrophobic aromatic ether structure that synergizes with the protonated carboxyl group to inhibit swelling in the stomach acid, and its rigid aromatic ring enhances the mechanical properties of the film and is compatible with ester plasticizers, reducing the risk of acid leakage.

[0019] 3. The invention further comprises plasticizers, light shielding agents, pore-forming agents and other excipients in the colon-targeted sustained-release layer, wherein the plasticizer is esterified from phthalic acid and 4-hydroxy-2-pyrrolidone, and its five-membered lactam structure is similar to the pyrrolidone unit of povidone as the binder, with good compatibility, which can significantly improve the flexibility and ductility of the coating, avoid cracks when the tablet core swells, and reduce drug leakage in the acid stage; at the same time, it does not affect the swelling jump of the copolymer at the colon pH, ensuring the targeted release of the colon-targeted sustained-release layer. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 FT-IR spectrum of hydroxypropyl methyl cellulose and pH-enzyme colon-targeting response agent in Preparation Example 1 of the invention; Figure 2 Expression of IL-4 in the serum and colon tissue of rats in each group in Test Example 2 of the invention; Figure 3 Expression of TNF-α in the serum and colon tissue of rats in each group in Test Example 2 of the invention; Figure 4 Expression of MPO in the serum and colon tissue of rats in each group in Test Example 2 of the invention. DETAILED DESCRIPTION

[0021] The technical solutions of the present application are further described below in combination with specific embodiments. However, those skilled in the art should understand that the following examples are only used to illustrate the present application, and should not be regarded as limiting the present application. The specific conditions not mentioned in the examples are carried out according to the conventional conditions or the conditions recommended by the manufacturer. The reagents or instruments used, if not specifically mentioned, are conventional products obtained through commercial channels. In the following examples, the polyacrylic acid ester is selected as Eudragit RS 100; the polysorbate is selected as Tween 80, and the binder I and the binder II are both selected as povidone K30.

[0022] Preparation Example 1 Preparation Example 1 provides a pH-enzyme colon-targeting response agent, which is prepared by the following preparation process: (1) Vanillic acid (1.68 g, 10 mmol) and potassium carbonate (3.60 g, 26 mmol) were added to 100 mL of acetonitrile, and then 4-bromo-2-methoxyaniline (2.62 g, 13 mmol) was slowly added after stirring. The temperature was raised to 70°C and reacted for 15 h. The obtained mixture was extracted with ethyl acetate and distilled water, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by silica gel column chromatography (ethyl acetate: dichloromethane = 1:1, v / v) to obtain intermediate 1 (weight 2.41 g, yield 83.38%); 1 H NMR (C 15 H 15 NO5, 400 MHz, DMSO): δ 12.62 (s, 1H), 7.75-7.69 (m,2H), 7.45 (d, 1H), 6.78 (d, 1H), 6.49-6.45 (m, 2H), 4.92 (s, 2H), 3.88 (s,3H), 3.82 (s, 3H); HRMS(ESI+): [M+H] + Calculated 290.10, found 290.10.

[0023] (2) Intermediate 1 (2.89 g, 10 mmol), 4-aminophenyl acrylate (1.96 g, 12 mmol), 2.5 M hydrochloric acid solution (6 mL), and 10 wt% sodium hypochlorite solution (41 mL, 55 mmol) were added to a reaction bottle, and reacted at room temperature for 4 h. Ethyl acetate was added to the reaction mixture for quenching, the water layer was extracted with ethyl acetate, the combined organic layer was dried over anhydrous sodium sulfate and concentrated, and purified by flash column chromatography (n-heptane: ethyl acetate = 80:20, v / v) to obtain vanillic acid derivative (3.53 g, yield 78.80%).1 H NMR (C 24 H 20 N2O7, 400 MHz, DMSO): δ 12.62 (s, 1H),7.88 (d, 3H), 7.75-7.69 (m, 2H), 7.45 (q, 3H), 6.84 (dd, 2H), 6.26 (dd, 1H),6.12 (dd, 1H), 5.76 (dd, 1H), 4.05 (s, 3H), 3.82 (s, 3H); HRMS(ESI+): [M+H] + Calculated 449.13, found 449.10.

[0024] (3) 2.5 g of hypromellose was weighed into 50 g of water, and the reaction solution was heated to 55°C under a nitrogen atmosphere, then 0.03 g of potassium sulfate (KPS) was added to initiate the reaction for 14 min, then 9 g of the vanillic acid derivative was added to the reaction solution, and after the dropwise addition was completed, the reaction was continued at 55°C for 1 h, and after cooling to room temperature, the dialysis bag (10 kDa) was used for interception, and the intercepted part was freeze-dried to obtain the pH-enzyme colon-targeting responsive agent.

[0025] The FT-IR spectra of hypromellose and the pH-enzyme colon-targeting responsive agent are shown in Figure 1 From Figure 1 it can be seen that the characteristic peak of the pH-enzyme colon-targeting responsive agent at 1430 cm -1 corresponds to the N=N stretching vibration, the characteristic peak at 1250 cm -1 corresponds to the asymmetric stretching of Ar-O-C, the characteristic peak at 1100 cm -1 corresponds to the symmetric stretching of Ar-O-C, and the characteristic peak at 3200 cm -1 corresponds to the stretching vibration of O-H in the carboxyl group, and the characteristic peak at 1680 cm -1 corresponds to the stretching vibration of the carbon-oxygen double bond in the carboxyl group; the above results confirm the successful preparation of the pH-enzyme colon-targeting responsive agent.

[0026] Preparation Example 2 Preparation Example 2 provides a pH-enzyme colon-targeting responsive agent, which is prepared by the following preparation process: (1) Add 1.68 g of vanillic acid (10 mmol) and 3.04 g of potassium carbonate (22 mmol) to 100 mL of acetonitrile, stir uniformly, and then slowly add 2.22 g of 4-bromo-2-methoxyaniline (11 mmol), and react at 60°C for 18 h; the obtained mixture is extracted with ethyl acetate and distilled water, the organic phase is washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by silica gel column chromatography (ethyl acetate: dichloromethane = 1:1, v / v) to obtain 2.10 g of intermediate 1 (yield 72.66%); the structure of intermediate 1 is confirmed by H NMR and HRMS. 1 H NMR and HRMS results are the same as those of Preparation Example 1.

[0027] (2) Add 2.89 g of intermediate 1 (10 mmol), 4-aminophenyl acrylate (1.63 g, 10 mmol), 2.5 M hydrochloric acid solution (4 mL), and 10 wt% sodium hypochlorite solution (37 mL, 50 mmol) to a reaction bottle, and react at room temperature for 3 h; add ethyl acetate to the reaction mixture for quenching, extract the aqueous layer with ethyl acetate, combine the organic layers, dry over anhydrous sodium sulfate, and concentrate, and purify by flash column chromatography (n-heptane: ethyl acetate = 80:20, v / v) to obtain a vanillic acid derivative; the structure of the vanillic acid derivative is confirmed by H NMR and HRMS. 1 H NMR and HRMS results are the same as those of Preparation Example 1.

[0028] (3) Weigh 2.5 g of hypromellose and dissolve it in 50 g of water, and then add 0.03 g of potassium sulfate (KPS) to initiate the reaction under a nitrogen atmosphere at 50°C for 15 min, and then add 8 g of the vanillic acid derivative to the reaction solution, and continue to react at 50°C for 1 h after the dropwise addition is completed, cool to room temperature, and then cut with a dialysis bag (10 kDa), and freeze-dry the cut part to obtain the pH-enzyme colon-targeting responsive agent.

[0029] Preparation Example 3 Preparation Example 3 provides a pH-enzyme colon-targeting responsive agent, which is prepared by the following preparation process: (1) Add 1.68 g of vanillic acid (10 mmol) and 3.88 g of potassium carbonate (28 mmol) to 100 mL of acetonitrile, stir uniformly, and then slowly add 3.03 g of 4-bromo-2-methoxyaniline (15 mmol), and react at 80°C for 12 h; the obtained mixture is extracted with ethyl acetate and distilled water, the organic phase is washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by silica gel column chromatography (ethyl acetate / dichloromethane = 1 / 1, v / v) to obtain 2.48 g of intermediate 1 (yield 85.80%); the structure of intermediate 1 is confirmed by H NMR and HRMS. 1H NMR and HRMS results are the same as Preparation Example 1.

[0030] (2) Intermediate 1 (2.89 g, 10 mmol), 4-aminophenyl acrylate (2.45 g, 15 mmol), 2.5 M hydrochloric acid solution (8 mL), and 10 wt% sodium hypochlorite solution (45 mL, 60 mmol) were added to a reaction bottle and reacted at room temperature for 5 h; ethyl acetate was added to the reaction mixture for quenching, the water layer was extracted with ethyl acetate, the combined organic layer was dried over anhydrous sodium sulfate and concentrated, and purified by flash column chromatography (n-heptane / ethyl acetate = 80 / 20, v / v) to obtain the vanillic acid derivative (3.59 g, yield 80.14%); the vanillic acid derivative was analyzed by H NMR and HRMS. 1 H NMR and HRMS results are the same as Preparation Example 1.

[0031] (3) 2.5 g of hypromellose was dissolved in 50 g of water, the reaction solution was heated to 60°C under a nitrogen atmosphere, 0.04 g of potassium sulfate (KPS) was added to initiate reaction for 12 min, then 10 g of the vanillic acid derivative was added to the reaction solution, after the dropwise addition was completed, the reaction was continued at 60°C for 0.5 h, after cooling to room temperature, it was intercepted with a dialysis bag (10 kDa), and the intercepted part was freeze-dried to obtain the pH-enzyme colon-targeting responsive agent.

[0032] Preparation Example 4 Preparation Example 4 provides a plasticizer prepared by the following preparation process: Phthalic acid (1.66 g, 10 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI, 5 g, 26 mmol), 4-dimethylaminopyridine (DMAP, 6.48 g, 53 mmol), and 4-hydroxy-2-pyrrolidone (2.32 g, 23 mmol) were added to 20 mL of acetonitrile, stirred uniformly, heated to 50°C, and reacted for 1 h; the acetonitrile was removed under reduced pressure, the residue was dissolved in ethyl acetate, the ethyl acetate phase was sequentially washed with 1M hydrochloric acid, saturated sodium bicarbonate, and saturated brine, dried over anhydrous magnesium sulfate and filtered, concentrated, and purified by column chromatography (n-heptane / ethyl acetate = 5 / 1, v / v) to obtain the plasticizer (2.75 g, yield: 82.82%); 1 H NMR (C 16 H 16N2O6, 400 MHz, DMSO): δ 8.07-8.04 (m, 2H), 7.73-7.71 (m, 2H), 5.30-5.27 (m, 4H), 3.96 (q, 2H), 3.71 (q, 2H), 2.76 (q, 2H),2.54 (q, 2H); HRMS(ESI+): [M+H] + Calculated 333.10, found 333.10.

[0033] Preparation Example 5 Preparation Example 5 provides a plasticizer, which is prepared by the following preparation process: Phthalic acid (1.66 g, 10 mmol), EDCI (4.79 g, 25 mmol), DMAP (6.15 g, 50 mmol) and 4-hydroxy-2-pyrrolidinone (2.02 g, 20 mmol) were added to 20 mL of acetonitrile, stirred uniformly and heated to 40°C for 1.5 h; acetonitrile was removed under reduced pressure, the residue was dissolved in ethyl acetate, the ethyl acetate phase was washed with 1M hydrochloric acid, saturated sodium bicarbonate and saturated brine in turn, dried over anhydrous magnesium sulfate and filtered, concentrated and purified by column chromatography (n-heptane / ethyl acetate = 5 / 1, v / v) to obtain the plasticizer (2.48 g, yield 74.69%).

[0034] Preparation Example 6 Preparation Example 6 provides a plasticizer, which is prepared by the following preparation process: Phthalic acid (1.66 g, 10 mmol), EDCI (4.79 g, 25 mmol), DMAP (6.15 g, 50 mmol) and 4-hydroxy-2-pyrrolidinone (2.02 g, 20 mmol) were added to 20 mL of acetonitrile, stirred uniformly and heated to 40°C for 1.5 h; acetonitrile was removed under reduced pressure, the residue was dissolved in ethyl acetate, the ethyl acetate phase was washed with 1M hydrochloric acid, saturated sodium bicarbonate and saturated brine in turn, dried over anhydrous magnesium sulfate and filtered, concentrated and purified by column chromatography (n-heptane / ethyl acetate = 5 / 1, v / v) to obtain the plasticizer (2.48 g, yield 74.69%).

[0035] Example 1 Example 1 provides a minocycline hydrochloride-melatonin compound colon-targeting capsule, which comprises pellets and a capsule shell, wherein: The pellets comprise a drug core and a colon-targeting sustained-release layer wrapped outside the drug core, and the mass ratio of the two is 1:0.12; The drug core comprises the following raw materials in parts by weight: minocycline hydrochloride 30 parts, melatonin 15 parts, lactose 30 parts, povidone K30 (adhesive I) 7 parts; The colon-targeting sustained-release layer comprises the following raw materials in parts by weight: povidone K30 (adhesive II) 10 parts, the pH-enzyme colon-targeting response agent of Preparation Example 1 15 parts, the plasticizer of Preparation Example 4 3 parts, titanium dioxide 4 parts, polyacrylic acid ester 12 parts, and Tween 80 6 parts.

[0036] Example 1 also provides a preparation method of the above-mentioned minocycline hydrochloride-melatonin compound colon-targeting capsule, comprising the following steps: (a) Preparation of the drug core: according to the weight parts of each raw material in the above-mentioned drug core, minocycline hydrochloride, melatonin, lactose and part of the adhesive I (5 parts) are mixed uniformly after being passed through a 100-mesh sieve to obtain a mixed drug powder; the remaining adhesive I (2 parts) is dissolved in water to form a binding liquid and is added to the mixed drug powder, which is stirred for 3-5 min under the condition of a rotation speed of 100 rpm and a shear speed of 1200 rpm to prepare a soft material; the soft material is placed into an out-roller compactor (extrusion frequency 25 HZ, shear frequency 15 HZ / min) to prepare wet pellets, which are dried at 60-65℃ until the moisture content is <2.5-3% to prepare the drug core; (b) Preparation of the pellets: according to the weight parts of each raw material in the above-mentioned colon-targeting sustained-release layer, the adhesive II, the pH-enzyme colon-targeting response agent, the plasticizer, titanium dioxide, polyacrylic acid ester and polysorbate are dissolved in a mixed solution of 160 parts of pure water and 500 parts of ethanol to obtain a mixed liquid; the drug powder prepared in step (a) is placed in a fluidized bed (inlet air temperature 45℃, preheating for 10 min), and the mixed liquid is sprayed at a spraying pressure of 0.1 MPa, a flow rate of 2 ml / min and an increase in weight of 12%; after the spraying is completed, drying is continued for 20 min, and the 16-24-mesh pellets are screened to obtain the pellets; (c) The above-mentioned pellets are loaded into a No. 0 hard capsule shell (the material of the capsule shell is hydroxypropyl methyl cellulose) to obtain the minocycline hydrochloride-melatonin compound colon-targeting capsule, and the content of minocycline hydrochloride in each capsule is 100 mg.

[0037] Example 2 Example 2 provides a minocycline hydrochloride-melatonin compound colon-targeting capsule, which comprises pellets and a capsule shell, wherein: The pellets comprise a drug core and a colon-targeting sustained-release layer wrapped outside the drug core, and the mass ratio of the two is 1:0.1; The drug core comprises the following raw materials in parts by weight: minocycline hydrochloride 20 parts, melatonin 10 parts, microcrystalline cellulose 20 parts, and povidone K30 (adhesive I) 4 parts; The colon-targeting sustained-release layer comprises the following raw materials in parts by weight: povidone K30 (binder II) 5 parts, the pH-enzyme colon-targeting response agent of Preparation Example 2 10 parts, the plasticizer of Preparation Example 5 2 parts, titanium dioxide 3 parts, ethyl cellulose 5 parts, and polyethylene glycol 6000 2 parts.

[0038] Example 2 also provides a preparation method of the above-mentioned minocycline hydrochloride-melatonin compound colon-targeting capsule, comprising the following steps: (a) preparing a drug core: according to the parts by weight of each raw material in the above-mentioned drug core, minocycline hydrochloride, melatonin, microcrystalline cellulose and part of binder I (3 parts) are mixed uniformly after being passed through a 100-mesh sieve to obtain a mixed drug powder; the remaining binder I (1 part) is dissolved in water to form a binding liquid and is added to the mixed drug powder, which is stirred under the condition of a rotation speed of 100 rpm and a shear speed of 1200 rpm for 3-5 min to prepare a soft material; the soft material is placed into an out-roller compactor (extrusion frequency 20 HZ, shear frequency 20 HZ / min) to prepare wet pellets, which are dried at 60-65°C until the moisture content is <2.5-3% to prepare the drug core; (b) preparing pellets: according to the parts by weight of each raw material in the above-mentioned colon-targeting sustained-release layer, the binder II, the pH-enzyme colon-targeting response agent, the plasticizer, titanium dioxide, ethyl cellulose and polyethylene glycol 6000 are dissolved in a mixed solution of 160 parts of pure water and 500 parts of ethanol to obtain a mixed liquid; the drug powder prepared in step (a) is placed in a fluidized bed (inlet air temperature 45°C, preheating for 10 min), and the mixed liquid is sprayed at a spraying pressure of 0.1 MPa, a flow rate of 2 ml / min and an increase of 10% to carry out spraying, and after the spraying is completed, drying is continued for 20 min, and the 16-24-mesh pellets are screened to obtain the pellets; (c) the above-mentioned pellets are loaded into a No. 0 hard capsule shell (the material of the capsule shell is hydroxypropyl methyl cellulose) to obtain the minocycline hydrochloride-melatonin compound colon-targeting capsule, and the content of minocycline hydrochloride in each capsule is 120 mg.

[0039] Example 3 Example 3 provides a minocycline hydrochloride-melatonin compound colon-targeting capsule, which comprises pellets and a capsule shell, wherein: the pellets comprise a drug core and a colon-targeting sustained-release layer wrapped outside the drug core, and the mass ratio of the two is 1:0.15; the drug core comprises the following raw materials in parts by weight: minocycline hydrochloride 40 parts, melatonin 20 parts, lactose 40 parts, and povidone K30 (binder I) 11 parts; the colon-targeting sustained-release layer comprises the following raw materials in parts by weight: povidone K30 (binder II) 15 parts, the pH-enzyme colon-targeting response agent of Preparation Example 3 20 parts, the plasticizer of Preparation Example 6 4 parts, titanium dioxide 5 parts, polyacrylic acid resin 20 parts, and polysorbate 10 parts.

[0040] Example 3 also provides a preparation method of the above minocycline hydrochloride-melatonin compound colon-targeting capsule, comprising the following steps: (a) Preparation of the drug core: according to the weight fraction of each raw material in the above drug core, minocycline hydrochloride, melatonin, lactose and part of the binder I (8 parts) are mixed uniformly after being sieved through a 100-mesh sieve to obtain a mixed drug powder; the remaining binder I (3 parts) is dissolved in water to form a binding liquid and added to the mixed drug powder, which is stirred at a rotation speed of 100 rpm and a shear speed of 1200 rpm for 3-5 min to prepare a soft material; the soft material is placed into an out-roller compactor (extrusion frequency 30 HZ, shear frequency 20 HZ / min) to prepare wet pellets, which are dried at 60-65°C until the moisture content is <2.5-3% to prepare the drug core; (b) Preparation of the pellets: according to the weight fraction of each raw material in the above colon-targeting sustained-release layer, the binder II, the pH-enzyme colon-targeting responsive agent, the plasticizer, titanium dioxide, polyacrylic acid resin, and polysorbate are dissolved in a mixed solution of 160 parts of pure water and 500 parts of ethanol to obtain a mixed solution; the drug powder prepared in step (a) is placed in a fluidized bed (inlet air temperature 45°C, preheating for 10 min), and the mixed solution is sprayed at a spraying pressure of 0.1 MPa, a flow rate of 2 ml / min, and an increase of 15%, and after spraying, it is dried for 20 min, and the 16-24-mesh pellets are screened to obtain the pellets; (c) The above pellets are loaded into a No. 0 hard capsule shell (the material of the capsule shell is hydroxypropyl methyl cellulose) to obtain the minocycline hydrochloride-melatonin compound colon-targeting capsule, and the content of minocycline hydrochloride in each capsule is 110 mg.

[0041] Comparative Example 1 Comparative Example 1 is basically the same as Example 1, except that the plasticizer is replaced by dimethyl phthalate.

[0042] Comparative Example 2 Comparative Example 2 is basically the same as Example 1, except that the pH-enzyme colon-targeting responsive agent is replaced by hydroxypropyl methyl cellulose.

[0043] Test Example 1 According to the Paddle Method of Dissolution and Release Determination Method in Chinese Pharmacopoeia 2020 Edition Part IV 0931 Method II, the drug release characteristics of the capsules obtained in Examples 1-3 and Comparative Examples 1-2 are evaluated.

[0044] ① Capsules of Examples 1-3 and Comparative Examples 1-2 were weighed respectively, 50 mg each was placed in a dissolution cup, the volume of dissolution medium was 750 mL, the rotation speed was 100 r / min, and the temperature was (37±0.5) °C. First, release in artificial gastric juice with pH=1.2 for 2 h, and then in artificial small intestinal fluid with pH=6.8 for 4 h, the volume of dissolution medium met the sink condition. 1 mL was sampled at 2 h and 6 h respectively, and then 1 mL of the same volume of medium at the same temperature was immediately added. The collected sample was filtered through a 0.45 μm filter, and the content of minocycline hydrochloride and melatonin was determined by LC-MS / MS respectively, and the cumulative drug release rate was calculated. The results are shown in Table 1.

[0045] ② Capsules of Examples 1-3 and Comparative Examples 1-2 were weighed respectively, 50 mg each was placed in a dissolution cup, the volume of dissolution medium was 750 mL, the rotation speed was 100 r / min, and the temperature was (37±0.5) °C. Release in pH=7.4 phosphate buffer containing 4% rat cecal contents for 10 h, the volume of dissolution medium met the sink condition. 1 mL was sampled at 1 h, 2 h, 4 h, 6 h, 8 h, 10 h, and 12 h respectively, and then 1 mL of the same volume of medium at the same temperature was immediately added. The collected sample was filtered through a 0.45 μm filter, and the content of minocycline hydrochloride and melatonin was determined by LC-MS / MS respectively, and the cumulative drug release rate was calculated. The results are shown in Table 2.

[0046] Table 1 Table 2 As shown in Table 1, the capsules obtained in Examples 1-3 were basically not released in artificial gastric juice with pH=1.2, and only a small amount was released in artificial small intestinal fluid with pH=6.8, and the cumulative release amount within 6 h was less than 10%, while the cumulative release amount of Comparative Example 1 and Comparative Example 2 within 6 h was more than 10%.

[0047] As shown in Table 2, the capsules obtained in Examples 1-3 released rapidly in the medium containing rat cecal contents and could be basically released completely within 12 h. Combined with the results in Table 1, it can be proved that the minocycline hydrochloride-melatonin compound capsules prepared in the application have obvious colon targeting, and the in vitro release behavior meets the requirements of colon targeting. Comparative Example 2 replaces the pH-enzyme colon targeting response material with hydroxypropyl methyl cellulose which does not have enzyme responsiveness, and the cumulative release rate within 12 h is only about 22%, indicating that in the absence of pH-enzyme response material, the drug cannot be effectively released in the colon environment.

[0048] The pH-enzyme colon-targeting responsive agent of this invention is a graft copolymer obtained by free radical grafting of carboxylated vanillic acid derivative side chains onto hydroxypropyl methylcellulose (HPMC) as the main chain. This material exhibits excellent film-forming properties and shows significant swelling differences in different pH environments of the gastrointestinal tract: in the acidic environment of the stomach (pH=1.2), the carboxyl groups of the side chains are protonated, reducing the hydrophilicity of the material and limiting the swelling degree, effectively inhibiting drug release (see Table 1, Example 1 and Comparative Example 2); when it reaches the terminal ileum to the ascending colon (pH=7.4), the carboxyl groups are deprotonated, generating electrostatic repulsion, and the copolymer rapidly swells to form continuous diffusion channels, achieving drug release (see Table 2). Furthermore, the diazo groups introduced by azotization of the side chains can be specifically cleaved by azoreductase secreted by anaerobic bacteria in the colon, thereby achieving dual pH-enzyme triggering and enhancing colon-targeting.

[0049] The plasticizer used in the colon-targeted sustained-release layer of the present invention is formed by esterification of phthalic acid and 4-hydroxy-2-pyrrolidone. Its five-membered lactam structure is similar to the pyrrolidone unit of povidone as a binder, and has good compatibility. It can significantly improve the flexibility and extensibility of the coating, avoid cracking when the tablet core absorbs water and expands, and reduce drug leakage in the acid stage (see Table 1 Example 1 and Comparative Example 1).

[0050] Experimental Example 2 1. Laboratory animals and grouping Eighty healthy male SPF-grade SD rats (weighing 170±10 g) were randomly divided into eight groups of ten each: blank group, model group, positive control group, Examples 1-3 groups, and Comparative Examples 1-2 groups. The rats were acclimatized for three days before the experiment.

[0051] 2. Modeling A 5% sodium dextran solution was prepared using distilled water as a modeling agent. Except for the control group, which drank distilled water, all other groups drank the 5% sodium dextran solution for 7 consecutive days to establish a rat model of ulcerative colitis.

[0052] 3. Administration method Starting from day 8, the administration was changed, and the modeling agent sodium dextran solution was replaced with distilled water for the rats' daily drinking water. The blank control group and the model group were administered physiological saline by gavage; the positive control group was administered sulfasalazine suspension (0.1 g / mL, prepared by adding sulfasalazine to 4% hydroxypropyl methylcellulose solution) by gavage at a dose of 100 mg / kg; and groups 1-3 of Examples and groups 1-2 of Comparative Examples were administered the corresponding capsules by gavage at a dose of 100 mg crude drug / kg. The administration was repeated once daily for 6 consecutive days.

[0053] 4. Detection Indicators After the administration was finished on the 13th day, the rats were normally fed for 24 h, and 1 mL of blood was taken from the retro-orbital plexus on the 14th day and the rats were killed, and the serum was taken after the blood was centrifuged at 3000 r / min for 10 min at room temperature for 2 h, and the expression levels of IL-4, TNF-α and MPO in the serum were detected according to the operation instruction of the kit; a certain mass of colon tissue was taken, and physiological saline was used to prepare 10% tissue homogenate, and then the homogenate was diluted with physiological saline to 5%, and the expression levels of IL-4, TNF-α and MPO in the tissue homogenate were detected according to the operation instruction of the kit; the expression levels of IL-4 in the serum and the tissue homogenate are shown in Figure 2 , the expression levels of TNF-α in the serum and the tissue homogenate are shown in Figure 3 , and the expression levels of MPO in the serum and the tissue homogenate are shown in Figure 4 .

[0054] 5. Experimental results As shown in Figure 2 , 3 and 4, compared with the blank group, the contents of TNF-α and MPO in the serum and the colon tissue of the model group were significantly increased, and the content of IL-4 was significantly reduced. Compared with the model group, the contents of TNF-α and MPO in the serum and the colon tissue of the positive control group, examples 1-3 were decreased, and the content of IL-4 was increased, and the effect of comparative examples 1-2 was obviously not as good as example 1. The above results show that the capsule preparation prepared by the application can target the colon lesion site and effectively inhibit the inflammatory response.

[0055] Finally, it should be pointed out that the above examples are only used to illustrate the technical solutions of the application, but not to limit it. The basic principles and main features of the application have been described above with specific embodiments, and some modifications or replacements can be made on the basis of the application, but these modifications or replacements do not make the corresponding technical solutions deviate from the scope of the application claimed.

Claims

1. A compound colon-targeted capsule of minocycline hydrochloride-melatonin, comprising pellets and a capsule shell, characterized in that: the pellets comprise a drug core and a colon-targeted sustained-release layer wrapped outside the drug core, and the mass ratio of the two is 1: (0.1-0.15) ; the drug core comprises the following raw materials in parts by weight: minocycline hydrochloride 20-40 parts, melatonin 10-20 parts, filler 20-40 parts, and adhesive I 4-11 parts; the colon-targeted sustained-release layer comprises the following raw materials in parts by weight: adhesive II 5-15 parts, pH-enzyme colon-targeted response agent 10-20 parts, plasticizer 2-4 parts, light shielding agent 3-5 parts, sustained-release agent 5-20 parts, and pore-forming agent 2-10 parts; the pH-enzyme colon-targeted response agent is prepared by the following process: (1) adding vanillic acid and potassium carbonate into acetonitrile, then adding 4-bromo-2-methoxyaniline, and reacting under heating conditions to obtain intermediate 1, the structure of which is shown in formula I: (2) adding 4-aminophenyl acrylate and the intermediate 1 into a sodium hypochlorite solution, then adding a hydrochloric acid solution to react, and purifying to obtain a vanillic acid derivative, the structure of which is shown in formula II: (3) dissolving hydroxypropyl methyl cellulose in water, adding potassium sulfate to initiate under a nitrogen atmosphere after heating to 50-60℃, then adding the vanillic acid derivative to continue the reaction, and cooling, dialyzing, and freeze-drying to obtain the pH-enzyme colon-targeted response agent; in step (1), the molar ratio of the vanillic acid, potassium carbonate, and 4-bromo-2-methoxyaniline is 1: (2.2-2.8) : (1.1-1.5), the heating temperature is 60-80℃, and the reaction time is 12-18 h; in step (2), the amount ratio of the intermediate 1, 4-aminophenyl acrylate, sodium hypochlorite solution, and hydrochloric acid solution is 1 mmol: (1-1.5) mmol: (3.7-4.5) mL: (0.4-0.8) mL, the concentration of the sodium hypochlorite solution is 10 wt%, the concentration of the hydrochloric acid solution is 2.5 mol / L, and the reaction time is 3-5 h; in step (3), the mass ratio of the hydroxypropyl methyl cellulose, potassium sulfate, and vanillic acid derivative is 1: (0.012-0.016) : (3.2-4), the initiation time is 12-15 min, the reaction time is 0.5-1 h, and the molecular weight cut-off of the dialysis is 10 kDa; the plasticizer is prepared by the following process: adding o-phthalic acid, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, 4-dimethylaminopyridine, and 4-hydroxy-2-pyrrolidone into acetonitrile, and reacting under heating conditions to obtain the plasticizer after purification, the structure of which is shown in formula III: ​ ​ ​ 2. The minocycline-hydrochloride-melatonin compound colon-targeted capsule according to claim 1, characterized in that, ​ ​ ； ​ ； ​ 3. The minocycline-melatonin compound colon-targeted capsule according to claim 2, characterized in that, ​ 4. The minocycline-melatonin compound colon-targeted capsule of claim 2, characterized in that, ​ 5. The minocycline-melatonin compound colon-targeted capsule according to claim 2, characterized in that, ​ 6. The minocycline-hydrochloride-melatonin compound colon-targeted capsule according to claim 1, characterized in that, ​ ​ 。 7. The minocycline-melatonin compound colon-targeted capsule according to claim 6, characterized in that, The molar ratio of the phthalic acid, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, 4-dimethylaminopyridine and 4-hydroxy-2-pyrrolidinone is 1:(2.5-2.8):(5-5.6):(2-2.5); the heating temperature is 40-60 DEG C, and the reaction time is 0.5-1.5 h.

8. The minocycline-melatonin compound colon-targeted capsule according to claim 1, characterized in that, The filler is selected from any one of lactose and microcrystalline cellulose; the binder I and the binder II are both povidone K30; the light shielding agent is titanium dioxide; the sustained release agent is selected from any one of polyacrylic acid ester and ethyl cellulose; and the pore forming agent is selected from any one of polysorbate and polyethylene glycol 6000.

9. A process for preparing the minocycline hydrochloride-melatonin combination colon-targeted capsule according to any one of claims 1 to 8, characterized by, The method comprises the following steps: (a) preparing a drug core: mixing minocycline hydrochloride, melatonin, a filler and part of a binder I according to the weight fractions of the raw materials in the drug core to obtain a mixed drug powder; dissolving the remaining binder I in water to form a binder solution and adding the binder solution to the mixed drug powder, stirring, extruding and drying to obtain the drug core; (b) preparing pellets: adding a binder II, a pH-enzyme colon-targeting responsive agent, a plasticizer, a light shielding agent, a sustained release agent and a pore forming agent to an ethanol-water solution to obtain a mixed solution; wrapping the mixed solution onto the surface of the drug core obtained in step (a) by a fluidized bed bottom spraying process to form a colon-targeting sustained release layer, and drying to obtain the pellets; (c) preparing capsules: loading the pellets into a capsule shell to obtain the minocycline hydrochloride-melatonin compound colon-targeting capsules.

10. The method of claim 9, wherein the minocycline-melatonin compound colon-targeted capsule is prepared by, The content of minocycline hydrochloride in each minocycline hydrochloride-melatonin compound colon-targeting capsule is 100-120 mg.

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