WO2024164432A1 - Tacrolimus sustained-release preparation and preparation method therefor - Google Patents

Abstract

The present invention relates to the technical field of pharmaceutical preparations. Specifically, disclosed in the present invention are a tacrolimus sustained-release preparation and a preparation method therefor. In the tacrolimus sustained-release preparation provided in the present invention, sorbitol is selected as a filler, hydroxypropyl methyl cellulose is used as a solubilizer, ethyl cellulose is used as a sustained-release framework, calcium stearate and/or stearic acid are used as a lubricant, and cross-linked sodium carboxymethyl cellulose is used as a disintegrant. The present invention also achieves the aim of sustained release of a primary drug while increasing the solubility of tacrolimus, and achieves effective control over the release rate of tacrolimus, can more stably exert the efficacy of tacrolimus, reduce the frequency of administration, and improve the compliance of a patient. In addition, the present invention also solves the problem of ethanol dumping which easily occurs in the tacrolimus sustained-release preparation, effectively improves the effectiveness and safety of the medicine therein, has a simple preparation process and is suitable for industrial production.

Description

A tacrolimus sustained-release preparation and preparation method thereof Technical Field

The present invention relates to the technical field of pharmaceutical preparations, and in particular to a tacrolimus sustained-release preparation and a preparation method thereof.

Background Art

Tacrolimus was first extracted from the fermentation broth of Streptococcus Tsukuba by Fujisawa Pharmaceutical Co., Ltd. in Japan in 1984. It is one of the three pillar products for anti-transplant immune rejection at home and abroad, along with cyclosporine and mycophenolate mofetil. Tacrolimus was approved by the FDA for marketing in the United States in 1995 and entered the Chinese market in 1998. Both tacrolimus and cyclosporine A are calcineurin inhibitors, with similar but more extensive immunosuppressive effects as cyclosporine A. They mainly inhibit the release of interleukin-2 (L-2) to achieve a comprehensive inhibition of T lymphocytes. The efficacy is 10-100 times higher than that of cyclosporine A, and the adverse reactions are fewer. At present, tacrolimus is widely used in the transplantation of solid organs such as liver, pancreas, kidney, heart, and lung. It has become a first-line drug for organ transplant immunosuppressive therapy. At the same time, it also plays an active role in the treatment of autoimmune diseases such as atopic dermatitis (AD), systemic lupus erythematosus (SLE), and autoimmune eye diseases, and has a high clinical application value.

However, the therapeutic index (NTI) of tacrolimus is narrow, and its blood concentration must be maintained within a certain level. Too high a concentration will increase the risk of toxic reactions, while insufficient concentration will increase rejection reactions. Tacrolimus sustained-release preparations can stabilize the blood concentration of tacrolimus, reduce the CV of blood concentrations, and reduce the number of medications, which has a high clinical advantage. However, the formulation and production process of tacrolimus sustained-release preparations are difficult. The difficulties in its formulation and process are as follows: (1) Tacrolimus has very poor solubility in water (about 1.58 μM in water at 25°C), and the selected excipients need to have a sustained-release function while solubilizing tacrolimus; (2) Ethanol dumping is prone to occur in the preparation product; (3) The development of solubilizing and sustained-release excipients is difficult. The material and the main drug can be fully combined, and the process suitable for industrialization is difficult. Therefore, it is of great significance to develop a sustained-release preparation suitable for industrial production, which can slowly release tacrolimus and does not cause ethanol dumping.

Summary of the invention

In view of the above problems existing in the existing tacrolimus sustained-release preparations, the present invention provides a tacrolimus sustained-release preparation and a preparation method thereof.

In order to solve the above technical problems, the technical solution provided by the present invention is:

A tacrolimus sustained-release preparation comprises tacrolimus or a pharmaceutically acceptable salt thereof, sorbitol, hydroxypropyl methylcellulose, ethyl cellulose, calcium stearate and/or stearic acid, and cross-linked sodium carboxymethyl cellulose.

Compared with the prior art, the tacrolimus sustained-release preparation provided by the present invention selects sorbitol as a filler, hydroxypropyl methylcellulose as a solubilizer, ethyl cellulose as a sustained-release matrix, calcium stearate and/or stearic acid as a lubricant, and cross-linked sodium carboxymethyl cellulose as a disintegrant, thereby increasing the solubility of tacrolimus while achieving the purpose of sustained-release of the main drug, realizing effective control of the release rate of tacrolimus, being able to more stably exert the efficacy of tacrolimus, reducing the number of dosing times, and improving patient compliance; in addition, the problem of ethanol dumping that is prone to occur in tacrolimus sustained-release preparations is solved, and the drug efficiency and safety thereof are effectively improved.

Preferably, the tacrolimus sustained-release preparation comprises the following components in weight percentage: 1.02% tacrolimus or a pharmaceutically acceptable salt thereof, 85%-90% sorbitol, 1%-5% hydroxypropyl methylcellulose, 4%-10% ethyl cellulose, 0.5%-1.0% calcium stearate and/or stearic acid, and 0.5-2.0% cross-linked sodium carboxymethyl cellulose.

The preferred components are compounded in a specific ratio, which can effectively improve the solubility of tacrolimus and better control the sustained-release effect of the drug, especially improve the sustained-release effect of the tacrolimus sustained-release preparation in an ethanol environment, so that the blood drug concentration of the tacrolimus sustained-release preparation in an ethanol environment can also be maintained in a relatively stable and lasting effective range, thereby improving the safety of the drug.

Preferably, the sustained-release preparation is a sustained-release capsule.

The present invention also provides a method for preparing a tacrolimus sustained-release preparation, comprising the following steps:

Step a, preparing tacrolimus raw material;

Step b, crushing sorbitol, and sieving to obtain sorbitol fine powder;

Step c, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in an organic solvent, then sequentially adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder, mixing evenly, drying, and pulverizing to obtain a solid dispersion;

Step d, uniformly mixing cross-linked sodium carboxymethyl cellulose, calcium stearate and/or stearic acid with the solid dispersion, and filling to obtain the tacrolimus sustained-release preparation.

The preparation method of the tacrolimus sustained-release preparation provided by the present invention has simple process, good stability, no need for special equipment, low production cost, and low organic solvent residue in the prepared pentoxifylline sustained-release tablets, further improving the safety of the drug and being suitable for industrial production.

Preferably, the method for preparing the tacrolimus raw material comprises the following steps:

Step S1, adding a short-chain alcohol solvent to soak the mycelium obtained by separating the tacrolimus fermentation liquid to obtain an alcohol soaking liquid; adding an inorganic salt solution to the alcohol soaking liquid, continuing to soak, and separating the solid and the liquid to obtain a salt soaking liquid;

Step S2, concentrating the salt soaking solution to a potency of 100,000 μg/mL-150,000 μg/mL using a nanofiltration concentrator with a cut-off molecular weight of 200Da-300Da to obtain a nanofiltrate;

Step S3, adding sodium dodecylbenzene sulfonate to the nanofiltrate, mixing evenly, adding purified water, crystallizing, solid-liquid separation, washing, and drying to obtain tacrolimus crude powder;

Step S4, dissolving the tacrolimus crude powder with a polar solvent, adding purified water, mixing evenly, adding to a dynamic axial compression column, eluting with a mixed solution of polar solvent and purified water as a chromatography liquid, collecting target components, and obtaining a layer washing liquid; wherein the filler of the dynamic axial compression column is UniSil C8 UItra Plus, and the polar solvent is at least one of ethanol, methanol or acetonitrile;

Step S5, concentrating the chromatographic liquid to a non-polar solvent to obtain a concentrated liquid; adding an acetate solvent to the concentrated liquid for extraction, concentrating the extract to dryness, adding an ethanol solution to dissolve, cooling and crystallizing, solid-liquid separation, washing, and drying to obtain a tacrolimus raw material.

The preparation method of tacrolimus raw material provided by the present invention comprises the following steps: soaking mycelium obtained by separating tacrolimus fermentation broth in a short-chain alcohol solvent and an inorganic salt solution to preliminarily remove pigments and other protein impurities contained in the mycelium, and then concentrating and removing ions and small molecular impurities with a nanofiltration concentrator with a cut-off molecular weight of 200Da-300Da, and then adding sodium dodecylbenzene sulfonate to the concentrated solution to change the distribution ratio of tacrolimus and impurities in the mother liquor and crystals, and increase the solubility of impurities in the mother liquor, so that tacrolimus can be fully crystallized and precipitated, and further remove secondary metabolites produced in the fermentation process, thereby improving the quality of the target product, and finally purifying and separating with a dynamic axial compression column filled with UniSil 10-120 C8 UItra Plus to fully remove impurities such as ascomycin and dihydrotacrolimus and other analogues, thereby greatly improving the purity of tacrolimus, and finally obtaining a high-purity tacrolimus raw material with a content of more than 99.98%, which is beneficial to improving the clinical drug safety of tacrolimus preparation products.

Preferably, in step S1, the short-chain alcohol solvent is at least one of methanol, ethanol or isopropanol.

Preferably, in step S1, the inorganic salt solution is at least one of an aluminum sulfate solution, a ferrous sulfate solution or a zinc sulfate solution with a mass concentration of 20%-25%.

Preferably, in step S1, the content of the short-chain alcohol solvent in the alcohol soaking solution is 70%-75%.

Preferably, in step S1, the amount of the inorganic salt solution added is 4%-6% of the volume of the alcohol soaking solution.

Preferably, in step S1, the soaking time is 2h-3h.

The preferred short-chain alcohol solvent, inorganic salt and added amount can fully remove the pigment and other protein impurities contained in the mycelium.

Preferably, in step S3, the mass volume ratio of the sodium dodecylbenzene sulfonate to the nanofiltrate is 0.02-0.05:1, wherein the mass unit is g and the volume unit is mL.

Preferably, in step S3, the volume ratio of the purified water to the nanofiltrate is 0.75-1.0:1.

Preferably, in step S3, the purified water is added slowly, and the adding time is controlled to be 1h-2h.

Preferably, in step S3, the crystallization temperature is 15°C-35°C, and the crystallization time is 12h-24h.

The preferred addition amounts of sodium dodecylbenzene sulfonate and purified water and the crystallization conditions can fully crystallize tacrolimus and reduce the content of impurities in the crystals, thereby improving the separation effect of tacrolimus and impurities, and further facilitating the improvement of the purification effect and efficiency of subsequent high pressure chromatography.

Furthermore, in step S3, the filter cake is washed with 45wt%-50wt% ethanol solution.

Preferably, in step S4, the loading amount of the tacrolimus crude powder is 0.8%-1.0% of the column volume, wherein the unit of the loading amount is g and the unit of the column volume is mL.

Preferably, in step S4, the volume-to-mass ratio of the polar solvent to the tacrolimus crude powder is 8-12:1, wherein the volume unit is mL and the mass unit is g.

Preferably, in step S4, the amount of purified water added is 20%-25% of the volume of the dissolved solution.

Preferably, in step S4, the content of the polar solvent in the chromatography fluid is 55wt%-65wt%.

Preferably, in step S4, the chromatography flow rate is 5 BV/h-6 BV/h.

Preferably, in step S4, the diameter of the dynamic axial compression column is 80 mm, the particle size of the filler is 10 μm, and the pore size of the filler is The height of the packing is 250 mm.

The preferred high-pressure chromatography conditions are conducive to effectively separating tacrolimus from impurities, especially ascomycin and dihydrotacrolimus and other analogs, achieving better separation effects in a shorter time and improving separation efficiency.

It should be noted that in step S4, the filling steps of the dynamic axial compression column are as follows: 900 g of the chromatographic filler is homogenized with 2.5 L of isopropanol to obtain a homogenate, the homogenate is added to the dynamic axial compression column, the isopropanol is discharged and the chromatographic filler is compacted to complete the filling of the dynamic axial compression column, and then the dynamic axial compression column is balanced with the chromatographic fluid.

It should be noted that in step S4, the chromatography process is monitored by high performance liquid chromatography, and the chromatographic solution containing ascomycin <0.1% and dihydrotacrolimus <0.1% is collected.

Preferably, in step S5, the acetate solvent is one or both of ethyl acetate and butyl acetate.

Preferably, in step S5, the volume ratio of the acetate solvent to the concentrated solution is 0.8-1.0:1.

Preferably, in step S5, the concentration of the ethanol solution is 45wt%-50wt%, and the potency of tacrolimus in the system after dissolution is 15μg/mL-300,000μg/mL.

Preferably, in step S5, the crystallization temperature is 2°C-8°C, and the crystallization time is 4h-6h.

Exemplarily, in step S5, the filter cake is washed with 45 wt%-50 wt% ethanol solution.

Optimal extraction solvent and crystallization conditions can further improve the purity of tacrolimus.

It should be noted that, in step S5, an alcohol meter is used to detect whether the concentrated liquid contains a polar solvent.

Preferably, in step b, the sorbitol is crushed and then passed through a 100-200 mesh sieve.

Preferably, in step c, the organic solvent is at least one of ethanol, propylene glycol or ethyl acetate.

Preferably, in step c, the amount of the organic solvent added is 35-60 times the mass of the solid formulation.

Furthermore, in step c, the content of particles smaller than 40 μm is greater than 90%.

Furthermore, in step c, the drying temperature is 40-60° C., and the drying time is 1.5 h-6 h.

The tacrolimus sustained-release preparation provided by the present invention has a simple formula, a simple and easy preparation process, an in vitro dissolution curve that is highly consistent with the original preparation, high drug stability, and can better exert the efficacy of tacrolimus. In an ethanol environment, the solubility of the tacrolimus sustained-release preparation prepared by the present invention does not change significantly, and the dissolution property is stable, which effectively improves the stability of the tacrolimus sustained-release preparation, thereby facilitating the improvement of the safety of its clinical application, and can achieve effective substitution of the original product, thus having broad application prospects.

DETAILED DESCRIPTION

In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

In order to better illustrate the present invention, further examples are given below.

Example 1

This embodiment provides a method for preparing a tacrolimus raw material, comprising the following steps:

Step a, taking 50L of tacrolimus fermentation broth with a fermentation unit of 1024 μg/mL, vacuum filtering the tacrolimus fermentation broth, retaining mycelium, adding the mycelium to a soaking tank, adding 40L of anhydrous ethanol to the soaking tank, detecting the ethanol content in the soaking liquid with an alcohol meter to 72%, soaking at room temperature for 2 hours, then adding 2L of aluminum sulfate solution (450g of aluminum sulfate dissolved in 2L of deionized water) to the fermentation tank, soaking at room temperature for 2 hours, filtering with a plate and frame filter, and retaining 40L of the soaking liquid;

Step b, adding the soaking liquid to a D2540 nanofiltration concentrator for pre-concentration, and then transferring to a D1812 nanofiltration concentrator for concentration to a potency of 138450 μg/mL, with a concentrated liquid volume of 300 mL;

Step c, add 6g of sodium dodecylbenzene sulfonate to the concentrated solution, stir, slowly add 225mL of purified water, the addition time is controlled to be 1h, after the addition of water is completed, crystallize at room temperature for 14h, filter, rinse the filter cake with 50% ethanol solution, and dry in a vacuum drying oven at 40°C to obtain 41.95g of crude powder;

Step d, 10 g of each batch of crude powder was taken, dissolved in 100 mL of anhydrous ethanol, and then 25 mL of purified water was added, and then loaded into a 1.25 L dynamic axial compression column, and chromatographed with 60% ethanol aqueous solution at a chromatography flow rate of 5 VB/h, and the peak platform part of the chromatographic solution was collected, and all the chromatographic solutions were combined to a volume of 8200 mL with a titer of 4095 μg/mL;

Step e, concentrating the chromatographic liquid until there is no ethanol, adding 700 mL of ethyl acetate for extraction, concentrating the extract to dryness, adding 170 mL of 50% ethanol aqueous solution to dissolve while hot, cooling to 6° C., crystallizing for 5 hours, filtering with suction, rinsing the filter cake with 50% ethanol solution, and drying in a vacuum drying oven at 40° C. to obtain 23.15 g of tacrolimus product with HPLC purity of 99.99%, 0.01% of dihydrotacrolimus, and a total yield of 45.12%.

The preparation method of the dynamic axial compression column in step d is as follows: 900 g of UniSil 10-120 C8 UItra Plus silica gel filler with a particle size of 10 μm is homogenized with 2.5 L of isopropanol to obtain a homogenate, the homogenate is added to a dynamic axial compression column (height 650 mm, diameter 80 mm), the isopropanol is discharged and the chromatographic filler is compacted to a filling height of 250 mm, the dynamic axial compression column is filled, and then the dynamic axial compression column is balanced with a chromatography fluid.

By adopting other process conditions defined in the present invention to prepare tacrolimus raw materials, as long as each parameter is within the range defined in the present invention, the technical effect equivalent to that of Example 1 can be achieved.

Example 2

This embodiment provides a tacrolimus sustained-release capsule, and the prescription dosage is shown in the following table:

The preparation method of the above-mentioned tacrolimus sustained-release capsule is as follows:

Step a, crushing sorbitol, sieving, and obtaining sorbitol fine powder;

Step b, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in anhydrous ethanol, then adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder in sequence, stirring evenly, drying in a drying oven at a drying temperature of 40° C. for a drying time of 6 hours, and crushing the dried material until the content of particles less than 40 μm is greater than 90%, to obtain a solid dispersion;

Step c, mixing the weighed cross-linked sodium carboxymethyl cellulose and calcium stearate with the solid dispersion evenly, and filling to obtain the tacrolimus sustained-release capsules.

Example 3

This embodiment provides a tacrolimus sustained-release capsule, and the prescription dosage is shown in the following table:

The preparation method of the above-mentioned tacrolimus sustained-release capsule is as follows:

Step a, crushing sorbitol, sieving, and obtaining sorbitol fine powder;

Step b, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in anhydrous ethanol, then adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder in sequence, stirring evenly, drying in a drying oven at a drying temperature of 60° C. for a drying time of 1.5 h, and crushing the dried material until the content of particles less than 40 μm is greater than 90%, to obtain a solid dispersion;

Step c, mixing the weighed cross-linked sodium carboxymethyl cellulose and calcium stearate with the solid dispersion evenly, and filling to obtain the tacrolimus sustained-release capsules.

Example 4

This embodiment provides a tacrolimus sustained-release capsule, and the prescription dosage is shown in the following table:

The preparation method of the above-mentioned tacrolimus sustained-release capsule is as follows:

Step a, crushing sorbitol, sieving, and obtaining sorbitol fine powder;

Step b, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in anhydrous ethanol, then adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder in sequence, stirring evenly, drying in a drying oven at a drying temperature of 50° C. for a drying time of 3 hours, and crushing the dried material until the content of particles less than 40 μm is greater than 90%, to obtain a solid dispersion;

Step c, mixing the weighed cross-linked sodium carboxymethyl cellulose and calcium stearate with the solid dispersion evenly, and filling to obtain the tacrolimus sustained-release capsules.

Example 5

This embodiment provides a tacrolimus sustained-release capsule, and the prescription dosage is shown in the following table:

The preparation method of the above-mentioned tacrolimus sustained-release capsule is as follows:

Step a, crushing sorbitol, sieving, and obtaining sorbitol fine powder;

Step b, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in anhydrous ethanol, then adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder in sequence, stirring evenly, and drying in a drying oven at a drying temperature of 55° C. for a drying time of 4 hours, and crushing the dried material until the content of particles less than 40 μm is greater than 90%, to obtain a solid dispersion;

Step c, mixing the weighed cross-linked sodium carboxymethyl cellulose and calcium stearate with the solid dispersion evenly, and filling to obtain the tacrolimus sustained-release capsules.

Comparative Example 1

This comparative example provides a tacrolimus sustained-release capsule, the prescription and preparation method of which are exactly the same as those of Example 1, except that sorbitol is replaced by an equal amount of mannitol.

The above-mentioned Examples 2-5 and Comparative Example 1 all use the high-purity tacrolimus raw material prepared in Example 1.

Residual solvent determination

According to the second method of residual solvent determination in Part IV 0861 of the Chinese Pharmacopoeia 2020: gas chromatography, the detection conditions are: (5%) diphenyl-(95%) dimethylsiloxane as the stationary liquid, the injection temperature is 200°C, the detector temperature is 250°C, the headspace bottle temperature is 80°C, and the equilibrium time is 60 minutes. The residual solvent standard is not higher than 5‰.

The residual solvents in the solid dispersion in step b in Example 2-5 were tested to be 2.4‰, 2.2‰, 2.3‰ and 2.6‰, respectively, which met the standard requirements.

Dissolution curve determination

In order to investigate the dissolution of the tacrolimus sustained-release capsules prepared in the examples of the present invention, the dissolution curves of Examples 2-5 and the original formulation (Astellas Pharmaceuticals (China) Co., Ltd., batch number 1R3034A) were measured with reference to the dissolution and release test method (Chinese Pharmacopoeia 2015 edition, Part IV, General Rules, 0931, Method 2). The test results are shown in Tables 1 and 2.

Test solution:

pH 4.5 dissolution medium: pH 4.5 phosphoric acid solution (containing 0.1% sodium lauryl sulfate and 0.005% hydroxypropyl cellulose) 900 mL.

pH 6.8 dissolution medium: pH 6.8 potassium dihydrogen phosphate-disodium hydrogen phosphate buffer (containing 0.005% hydroxypropyl cellulose) 900 mL.

Dissolution temperature: 37.0±0.5℃; Rotation speed: 50rpm;

Chromatographic column: Merck-RP18 (4.0*55mm, 3μm);

Detector: UV spectrophotometer (measurement wavelength: 205nm);

Injection volume: 100mL;

Flow rate: 1.0mL/min;

Column temperature: 30°C;

Mobile phase: Measure 530 ml of water, 360 ml of acetonitrile, add 1 ml of phosphoric acid solution (precisely measure 3 ml of phosphoric acid, place in a 50 ml volumetric flask, and dilute to the scale with water), and mix well.

Table 1 Dissolution curve data of pH 4.5 dissolution medium

Table 2 Dissolution curve data of pH 6.8 dissolution medium

From the above data, it can be seen that under the dissolution medium conditions of pH 4.5 and pH 6.8, the dissolution curve of the tacrolimus sustained-release capsules prepared in the embodiment of the present invention is similar to that of the original product, which can achieve the same quality as the original product and can achieve a better replacement for the original product.

Ethanol pour test

Dissolution medium:

pH 1.2 hydrochloric acid solution + 0.005% HPC (5% concentration): according to the volume ratio, 95% pH 1.2 hydrochloric acid solution + 5% anhydrous ethanol, stir evenly.

pH 1.2 hydrochloric acid solution + 0.005% HPC (20% concentration): according to the volume ratio, 80% pH 1.2 hydrochloric acid solution + 20% anhydrous ethanol, stir evenly.

pH 1.2 hydrochloric acid solution + 0.005% HPC (40% concentration): according to the volume ratio, 60% pH 1.2 hydrochloric acid solution + 40% anhydrous ethanol, stir evenly.

The dissolution test method at each time point is the same as above. The results are shown in Tables 3 to 5.

Table 3 Ethanol pouring test in pH 1.2 medium (ethanol concentration 5%)

Table 4 Ethanol pouring test in pH 1.2 medium (ethanol concentration 20%)

Table 5 Ethanol pouring test in pH 1.2 medium (ethanol concentration 40%)

It can be seen from the above table that, compared with the comparative example, no burst release was found in the tacrolimus sustained-release capsules prepared in the examples of the present invention in different concentrations of ethanol, while the burst release of the tacrolimus sustained-release capsules prepared in the comparative example became more obvious with the increase of ethanol concentration.

Stability test

Examples 2-5 and Comparative Example 1, the original research (Stellar Pharmaceuticals (China) Co., Ltd., batch number 1R3034A) were subjected to long-term and accelerated tests. The long-term and accelerated test conditions are shown in Table 6, and the test results are shown in Tables 7-10.

Table 6 Long-term and accelerated test conditions

Table 7 Stability data of content

Table 8 Release stability data

Table 9 Moisture stability data

Table 10 Impurity stability data

The results of the stability test of the samples show that the tacrolimus sustained-release capsules prepared by the present invention have low residual solvent content, low moisture content, good dissolution, small changes in content and impurities after long-term and accelerated testing, good stability, and high safety for clinical use.

In summary, the tacrolimus sustained-release capsules prepared in the embodiments of the present invention have a high degree of similarity with the original product, good quality stability, and can replace the original product; the preparation production process is simple, and the process energy consumption is low, and it is easy to realize large-scale industrial production.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modification, equivalent substitution or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims (10)

A tacrolimus sustained-release preparation, characterized in that it comprises tacrolimus or a pharmaceutically acceptable salt thereof, sorbitol, hydroxypropyl methylcellulose, ethyl cellulose, calcium stearate and/or stearic acid, and cross-linked sodium carboxymethyl cellulose. The tacrolimus sustained-release preparation according to claim 1, characterized in that it comprises the following components in weight percentage: 1.02% tacrolimus or a pharmaceutically acceptable salt thereof, 85%-90% sorbitol, 1%-5% hydroxypropyl methylcellulose, 4%-10% ethyl cellulose, 0.5%-1.0% calcium stearate and/or stearic acid, and 0.5-2.0% cross-linked sodium carboxymethyl cellulose. The tacrolimus sustained-release preparation according to claim 1 or 2, characterized in that the sustained-release preparation is a sustained-release capsule. A method for preparing the tacrolimus sustained-release preparation according to any one of claims 1 to 3, characterized in that it comprises the following steps: Step a, preparing tacrolimus raw material; Step b, crushing sorbitol, and sieving to obtain sorbitol fine powder; Step c, weighing each component according to the designed ratio, then dissolving the weighed tacrolimus raw material in an organic solvent, then sequentially adding the weighed ethyl cellulose, hydroxypropyl methyl cellulose and sorbitol fine powder, mixing evenly, drying, and pulverizing to obtain a solid dispersion; Step d, uniformly mixing cross-linked sodium carboxymethyl cellulose, calcium stearate and/or stearic acid with the solid dispersion, and filling to obtain the tacrolimus sustained-release preparation. The method for preparing a tacrolimus sustained-release preparation according to claim 4, characterized in that in step a, the method for preparing the tacrolimus raw material comprises the following steps: Step S1, adding a short-chain alcohol solvent to soak the mycelium obtained by separating the tacrolimus fermentation liquid to obtain an alcohol soaking liquid; adding an inorganic salt solution to the alcohol soaking liquid, continuing to soak, and separating the solid and the liquid to obtain a salt soaking liquid; Step S2, concentrating the salt soaking solution to a potency of 100,000 μg/mL-150,000 μg/mL using a nanofiltration concentrator with a cut-off molecular weight of 200Da-300Da to obtain a nanofiltrate; Step S3, adding sodium dodecylbenzene sulfonate to the nanofiltrate, mixing evenly, adding purified water, crystallizing, solid-liquid separation, washing, and drying to obtain tacrolimus crude powder; Step S4, dissolving the tacrolimus crude powder with a polar solvent, adding purified water, mixing evenly, adding to a dynamic axial compression column, eluting with a mixed solution of polar solvent and purified water as a chromatography liquid, collecting target components, and obtaining a layer washing liquid; wherein the filler of the dynamic axial compression column is UniSil C8 UItra Plus, and the polar solvent is at least one of ethanol, methanol or acetonitrile; Step S5, concentrating the chromatographic liquid to a non-polar solvent to obtain a concentrated liquid; adding an acetate solvent to the concentrated liquid for extraction, concentrating the extract to dryness, adding an ethanol solution to dissolve, cooling and crystallizing, solid-liquid separation, washing, and drying to obtain a tacrolimus raw material. The method for preparing a tacrolimus sustained-release preparation according to claim 5, characterized in that in step S1, the short-chain alcohol solvent is at least one of methanol, ethanol or isopropanol; and/or In step S1, the inorganic salt solution is at least one of an aluminum sulfate solution, a ferrous salt solution or a zinc sulfate solution with a mass concentration of 20%-25%; and/or In step S1, the content of the short-chain alcohol solvent in the alcohol soaking solution is 70%-75%; and/or In step S1, the amount of the inorganic salt solution added is 4%-6% of the volume of the alcohol soaking solution; and/or In step S1, the soaking time is 2h-3h. The method for preparing a tacrolimus sustained-release preparation according to claim 5, characterized in that, in step S3, the mass volume ratio of the sodium dodecylbenzene sulfonate to the nanofiltrate is 0.02-0.05:1, wherein the mass unit is g and the volume unit is mL; and/or In step S3, the volume ratio of the purified water to the nanofiltrate is 0.75-1.0:1; and/or In step S3, the purified water is added slowly, and the adding time is controlled to be 1h-2h; and/or In step S3, the crystallization temperature is 15°C-35°C, and the crystallization time is 12h-24h. The method for preparing a tacrolimus sustained-release preparation according to claim 5, characterized in that, in step S4, the loading amount of the tacrolimus crude powder is 0.8%-1.0% of the column volume, wherein the unit of the loading amount is g and the unit of the column volume is mL; and/or In step S4, the volume-to-mass ratio of the polar solvent to the tacrolimus crude powder is 8-12:1, wherein the volume unit is mL and the mass unit is g; and/or In step S4, the amount of purified water added is 20%-25% of the volume of the dissolved solution; and/or In step S4, the content of the polar solvent in the chromatography liquid is 55wt%-65wt%; and/or In step S4, the chromatography flow rate is 5 BV/h-6 BV/h; In step S4, the diameter of the dynamic axial compression column is 80 mm, the particle size of the filler is 10 μm, and the pore size of the filler is The height of the packing is 250 mm. The method for preparing a tacrolimus sustained-release preparation according to claim 5, characterized in that in step S5, the acetate solvent is one or both of ethyl acetate or butyl acetate; and/or In step S5, the volume ratio of the acetate solvent to the concentrated solution is 0.8-1.0:1; and/or In step S5, the concentration of the ethanol solution is 45wt%-50wt%, and the potency of tacrolimus in the system after dissolution is 15μg/mL-300,000μg/mL; and/or In step S5, the temperature of the cooling crystallization is 2°C-8°C, and the crystallization time is 4h-6h. The method for preparing a tacrolimus sustained-release preparation according to claim 4, characterized in that, in step b, the sorbitol is crushed and passed through a 100-200 mesh sieve; and/or In step c, the organic solvent is at least one of ethanol, propylene glycol or ethyl acetate; and/or In step c, the amount of the organic solvent added is 35-60 times the mass of the solid formulation.