Propylthiouracil liposome injection and preparation method thereof

Abstract

The invention provides a propylthiouracil liposome injection and a preparation method thereof. The propylthiouracil liposome injection comprises the following components: propylthiouracil, hydrogenated soybean lecithin, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000, alpha-tocopherol, a phosphate buffer solution and cane sugar. The preparation process comprises the steps of lipid film preparation, hydrated active drug loading, high-pressure homogenization, concentration and purification, sterilization and filtration, subpackaging and freeze-drying. The prepared propylthiouracil injection has the advantages of being fast in redissolution, good in stability, low in toxic and side effect, fast in clinical use effect and the like, is suitable for industrial large-scale production, can be used by patients who have hyperthyroidism emergency, coma, dysphagia and the like and cannot take oral liquid as the injection, and fills the blank in the current clinical treatment field.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of pharmaceutical preparations, and particularly relates to a liposome injection of propylthiouracil and a preparation method thereof. BACKGROUND

[0002] Propylthiouracil was discovered in the middle of the 20th century and has been widely used in the field of hyperthyroidism treatment. Compared with similar drugs, it has significant advantages: (1) high safety in pregnancy, which can effectively reduce the risk of teratogenicity; (2) can quickly inhibit the conversion of T4 to T3 in peripheral tissues, and rapidly relieve the symptoms of hyperthyroidism.

[0003] At present, propylthiouracil tablets have been widely used in clinical practice, but the large side effects and narrow therapeutic window of the existing oral dosage forms have been a major problem in clinical practice. Patent CN1602874A discloses a propylthiouracil sustained-release preparation and a preparation method thereof, and patent CN102764239A discloses a propylthiouracil sustained-release pellet. The above two patents improve the release of the drug and reduce the side effects by preparing propylthiouracil into a sustained-release preparation. However, due to the slow release of the sustained-release preparation, it often cannot quickly reach the blood drug concentration required for treatment in the face of hyperthyroidism crisis, and oral preparations cannot be used for patients who cannot take orally, such as coma and difficulty swallowing. Therefore, it is urgent to develop a propylthiouracil injection with rapid onset and small side effects to meet the clinical use requirements of the drug in specific situations.

[0004] Propylthiouracil has poor solubility in water, and it is difficult to prepare a uniform and stable solution. In addition, in the solution state, it is also susceptible to factors such as light, temperature and pH value, and it can be oxidized and degraded, resulting in a decrease in drug activity and an increase in impurities, which greatly hinders the development of propylthiouracil injection forms. SUMMARY

[0005] The purpose of the present application is to develop a propylthiouracil injection with rapid onset, small side effects and suitable for patients who cannot take orally, such as coma and difficulty swallowing.

[0006] To solve the above problems, the inventors have found through a large number of research and experimental attempts that a specific ratio of propylthiouracil, hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000, alpha-tocopherol, phosphate buffer and sucrose can be prepared into a propylthiouracil liposome injection with good solubility and stable quality. The preparation process uses high-pressure homogenization technology and tangential flow filtration technology to further reduce the particle size of the liposome, increase the uniformity of the particle size and the encapsulation efficiency. The preparation of the present application has the characteristics of rapid onset, low side effects and stable quality. The retention time of the active ingredient in the body circulation is significantly prolonged by the liposome technology, the drug has good biocompatibility, high bioavailability and enhanced efficacy.

[0007] The specific technical solutions are as follows:

[0008] 1. A propylthiouracil liposome injection, the components of which comprise propylthiouracil 100 parts, hydrogenated soybean lecithin 240-360 parts, cholesterol 80-120 parts, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 30-70 parts, alpha-tocopherol 1-3 parts, sucrose 100-300 parts, and a suitable amount of a buffer solution.

[0009] 2. The propylthiouracil liposome injection, characterized in that the dosage form is 50 mg or 100 mg.

[0010] 3. The propylthiouracil liposome injection, characterized in that the weight ratio of hydrogenated soybean lecithin to cholesterol is 3:1.

[0011] 4. The propylthiouracil liposome injection, characterized in that the buffer solution used is preferably a phosphate buffer solution (pH 7.4).

[0012] 5. A preparation method of a propylthiouracil liposome injection, characterized by comprising the following steps:

[0013] (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000, and alpha-tocopherol are added to a phosphate buffer solution with a pH value of 7.4 in a proportion, and stirred and dissolved at 55-70°C to form a uniform lipid solution.

[0014] (2) Drug-loaded hydration: propylthiouracil is added to the phosphate buffer solution at 55-70°C, stirred and dissolved, and then added to the lipid solution of step (1), and stirred for 2-4 hours to form multilayer liposomes.

[0015] (3) High-pressure homogenization: the crude liposome suspension of step (2) is homogenized by a high-pressure homogenizer to form single-chamber liposomes, and the average particle size of the liposomes is controlled to be 80-160 nm.

[0016] (4) Concentration and purification: the drug solution of step (3) is filtered by a tangential flow filtration system, and the cycle filtration is continued until the free drug content is less than 5%, and the prepared 8% sucrose phosphate buffer solution is continuously added during the cycle filtration process.

[0017] (5) Sterilization filtration and sub-packaging: the drug solution of step (4) is sequentially passed through a 1.2 μm pre-filter membrane and a 0.22 μm sterile filter cartridge, and then filled into a Schott bottle.

[0018] (6) Freeze-drying: first stage: cooling to -50℃ to -30℃, holding for 2 to 4 hours; second stage: warming to -25℃ to -10℃, holding for 8 to 12 hours, controlling the vacuum degree to be 10 to 40 Pa; third stage: warming to 20℃ to 60℃, holding for 6 to 8 hours, controlling the vacuum degree to be 10 to 30 Pa; controlling the final water content to be ≤2%.

[0019] 6. The preparation method of the propylthiouracil liposome injection, characterized in that the stirring and dissolving temperature in step (1) is preferably 60℃.

[0020] 7. The preparation method of the propylthiouracil liposome injection, characterized in that the average particle size of the liposome in step (3) is preferably controlled in the range of 110 to 130 nm.

[0021] Beneficial effects

[0022] The present application effectively solves the problems of poor solubility and stability of propylthiouracil in water by specific component proportioning composition and preparation process, develops propylthiouracil into an injection form, solves the problems that oral administration cannot rapidly reach a therapeutic blood drug concentration in hyperthyroidism emergency and oral preparations cannot be used for patients in coma or with difficulty in swallowing, and fills the clinical use blank of the drug. The preparation method is simple and suitable for industrialized mass production. DETAILED DESCRIPTION

[0023] The present application will be further described below in combination with specific examples, so that those skilled in the art can better understand the present application and implement it, but the examples are not used as a limitation to the present application.

[0024] Example 1: Propylthiouracil liposome injection and its preparation method

[0025] Propylthiouracil 100 parts, hydrogenated soybean lecithin 300 parts, cholesterol 100 parts, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, and phosphate buffer (pH 7.4) in an appropriate amount.

[0026] Preparation method:

[0027] (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 and α-tocopherol are added to a phosphate buffer solution (pH 7.4) in proportion, stirred and dissolved at 60℃ to form a uniform lipid solution.

[0028] (2) Drug-loaded hydration: propylthiouracil is added to the 60℃ phosphate buffer solution after stirring and dissolving, and then added to the lipid solution of step (1), stirred and hydrated for 2 hours to form a multilayer liposome.

[0029] (3) High pressure homogenization: the crude liposome suspension of step (2) is homogenized by a high pressure homogenizer to form single chamber liposomes.

[0030] (4) Concentration and purification: the drug solution of step (3) is subjected to tangential flow filtration system, and is filtered repeatedly until the free drug content is less than 5%, and the prepared 8% sucrose phosphate buffer is added constantly during the process.

[0031] (5) Bacterial removal filtration and sub-packaging: the drug solution of step (4) is subjected to 1.2 μm pre-filter membrane and 0.22 μm sterile filter core in sequence, and is filled into a vial.

[0032] (6) Freeze-drying: first stage: cooling to -40 °C, and keeping for 4 hours; second stage: warming to -15 °C, and keeping for 10 hours, with the vacuum degree controlled at 20 Pa; third stage: warming to 40 °C, and keeping for 6 hours, with the vacuum degree controlled at 10 Pa; the final water content is controlled to be less than or equal to 2%.

[0033] Example 2 Propylthiouracil liposome injection and its preparation method

[0034] Propylthiouracil 100 parts, hydrogenated soybean lecithin 240 parts, cholesterol 80 parts, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, phosphate buffer (pH 7.4) in proper amount.

[0035] Preparation method: same as example 1

[0036] Example 3 Propylthiouracil liposome injection and its preparation method

[0037] Propylthiouracil 100 parts, hydrogenated soybean lecithin 360 parts, cholesterol 120 parts, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, phosphate buffer (pH 7.4) in proper amount.

[0038] Preparation method: same as example 1

[0039] Comparative Example 1 Propylthiouracil liposome injection and its preparation method

[0040] Propylthiouracil 100 parts, hydrogenated soybean lecithin 300 parts, cholesterol 50 parts, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, phosphate buffer (pH 7.4) in proper amount.

[0041] Preparation method: same as example 1

[0042] Comparative Example 2 Propylthiouracil liposome injection and its preparation method

[0043] Propylthiouracil 100 parts, hydrogenated soybean lecithin 300 parts, cholesterol 100 parts, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, phosphate buffer (pH 7.4) in proper amount.

[0044] Preparation method:

[0045] (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000, α-tocopherol were added to the phosphate buffer solution (pH 7.4) in proportion, stirred and dissolved at 50°C to form a uniform lipid solution.

[0046] (2) Drug-loaded hydration: after propylthiouracil was added to the phosphate buffer solution stirred and dissolved at 50°C, it was added to the lipid solution of step (1), stirred and hydrated for 2 hours to form multilayer liposomes.

[0047] (3) High pressure homogenization: the crude liposome suspension of step (2) was homogenized by high pressure homogenizer to form single chamber liposomes.

[0048] (4) Concentration and purification: the drug solution of step (3) was filtered by tangential flow filtration system, and the cycle filtration was continued until the free drug content was less than 5%, and the prepared 8% sucrose phosphate buffer was continuously added during the cycle filtration process.

[0049] (5) Sterile filtration and dispensing: the drug solution of step (4) was sequentially passed through 1.2 μm prefilter membrane and 0.22 μm sterile filter cartridge, and was filled into a test tube.

[0050] (6) Freeze-drying: first stage: cooling to -40°C, holding for 4 hours; second stage: warming to -15°C, holding for 10 hours, controlling the vacuum degree to be 20 Pa; third stage: warming to 40°C, holding for 6 hours, controlling the vacuum degree to be 10 Pa; controlling the final water content to be ≤2%.

[0051] Propylthiouracil liposome injection and its preparation method

[0052] Propylthiouracil 100 parts, hydrogenated soybean lecithin 300 parts, cholesterol 100 parts, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, phosphate buffer (pH 7.4) in proper amount.

[0053] Preparation method:

[0054] (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000, α-tocopherol were added to the phosphate buffer solution (pH 7.4) in proportion, stirred and dissolved at 60°C to form a uniform lipid solution.

[0055] (2) Drug loading: Propylthiouracil was added to 60°C phosphate buffer solution and stirred to dissolve, then added to the lipid solution of step (1) and stirred for 2 hours to form multilayer liposomes. The prepared 8% sucrose phosphate buffer solution was added.

[0056] (3) Sterile filtration and dispensing: The drug solution of step (4) was passed through a 1.2 μm prefilter and a 0.22 μm sterile filter in sequence, and then filled into a vial.

[0057] (4) Freeze-drying: First stage: cooling to -40°C and holding for 4 hours; second stage: warming to -15°C and holding for 10 hours, with a vacuum degree of 20 Pa; third stage: warming to 40°C and holding for 6 hours, with a vacuum degree of 10 Pa; and the final water content was controlled to be ≤2%.

[0058] Propylthiouracil liposome injection and a preparation method thereof

[0059] Propylthiouracil 100 parts, hydrogenated soybean lecithin 300 parts, cholesterol 100 parts, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000 50 parts, α-tocopherol 1 part, sucrose 200 parts, and phosphate buffer solution (pH 7.4) in an appropriate amount.

[0060] Preparation method:

[0061] (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000, and α-tocopherol were added to a phosphate buffer solution (pH 7.4) in a proportion, stirred to dissolve at 60°C, and a uniform lipid solution was formed.

[0062] (2) Drug loading: propylthiouracil was added to 60°C phosphate buffer solution and stirred to dissolve, then added to the lipid solution of step (1) and stirred for 2 hours to form multilayer liposomes.

[0063] (3) High-pressure homogenization: the crude liposome suspension of step (2) was homogenized by a high-pressure homogenizer to form single-chamber liposomes.

[0064] (4) Concentration and purification: the drug solution of step (3) was subjected to tangential flow filtration to recycle filter until the free drug content was less than 5%, and the prepared 8% sucrose phosphate buffer solution was continuously added during the recycling filtration process.

[0065] (5) Sterile filtration and dispensing: the drug solution of step (4) was passed through a 1.2 μm prefilter and a 0.22 μm sterile filter in sequence, and then filled into a vial.

[0066] Test Example 1: content, encapsulation efficiency, and particle size detection

[0067] The propylthiouracil liposome injection prepared in Examples 1-3 and Comparative Examples 1-4 was detected for content, encapsulation rate and particle size, and the detection method was as follows:

[0068] The detection results are shown in the following table: Table 1 Detection results

[0069] From the above detection results, it can be seen that:

[0070] The propylthiouracil liposome injection prepared in Examples 1-3 has a content of more than 99%, a high encapsulation rate (>95%), an average particle size of 110-130 nm, and a span of <1.5, and the particle size distribution is uniform;

[0071] In Comparative Example 1, the weight ratio of hydrogenated soybean lecithin to cholesterol is 6:1, the proportion of cholesterol is not enough, the stability of the liposome membrane is poor, the drug loading rate is low, the particle size distribution is wide, and the content is low;

[0072] In Comparative Example 2, the temperature of the system during the preparation of the liposome is only 50°C, which is lower than the phase transition temperature, the phospholipid dispersion is not sufficient, the drug loading rate is low, the particle size distribution range is wide, and the content is low;

[0073] In Comparative Example 3, the homogenization process and tangential flow filtration process are not used, the average particle size of the liposome is very large, the encapsulation rate is low, and the particle size distribution is very wide.

[0074] Therefore, when the amount of ingredients is outside the amount of ingredients defined in the present application, the preparation process is not within the scope defined in the present application or the specified process steps are not used, it is impossible to obtain a propylthiouracil liposome injection with qualified average particle size and encapsulation rate.

[0075] Test Example 2 Leakage rate test

[0076] The samples prepared in Examples 1-3 and Comparative Examples 1-4 were periodically inspected at room temperature at 0 days, 30 days, 60 days, 90 days and 180 days, and the encapsulation rate was measured and compared with the drug amount encapsulated at 0 days to calculate the leakage rate, and the results are shown in the following table: Table 2 Detection results

[0077] From the above detection results, it can be seen that:

[0078] The propylthiouracil liposome injection prepared in Examples 1-3 has low leakage rate and little change in encapsulation rate within 180 days. In Comparative Example 1, the amount of cholesterol is insufficient, the fluidity of the lipid membrane is increased, the diffusion resistance of the drug is decreased, the membrane structure is easily collapsed, and the leakage rate is obviously increased. In Comparative Example 3, homogenization and tangential flow filtration are not used in the preparation process, the drug is not tightly embedded, and free drug is not removed, thus continuously inducing leakage. In Comparative Example 4, the product is not lyophilized, and the phospholipid is hydrolyzed and oxidized during liquid storage, the integrity of the lipid membrane is gradually destroyed, and the leakage rate is continuously increased.

[0079] Therefore, when the amount of the components is outside the range defined in the present application, and the preparation process produces a product that is not within the range defined in the present application or does not use the specified process steps, the encapsulation rate of the propylthiouracil liposome injection during storage cannot be effectively guaranteed.

[0080] Test Example 3: Stability Investigation

[0081] The samples prepared in Examples 1-3 and Comparative Examples 1-4 were subjected to accelerated (40°C) stability investigation, and the product properties, content, and related substances were detected. The results are shown in the following table: Table 3: Results of accelerated (40°C) condition investigation test

[0082] From the above detection results, it can be seen that:

[0083] The properties, content, and related substances of the propylthiouracil liposome injection prepared in Examples 1-3 do not change significantly under accelerated (40°C) conditions. However, the properties, content, and related substances of the propylthiouracil liposome injection prepared in Comparative Examples 1-4 change and increase during the accelerated storage period, because the amount of the components and the preparation process do not meet the requirements of the present application.

[0084] Test Example 4: Vessel Irritation Test

[0085] The propylthiouracil liposome injection prepared in Example 1 was prepared into a 2 mg / ml solution using 0.9% sodium chloride injection as the solvent, and 0.9% sodium chloride injection was used as the control. Twelve New Zealand white rabbits were randomly divided into two groups, and were injected with the drug solution (1 ml / kg) once a day for three consecutive days using the alternate ear vein injection method. The local tissue reaction was observed and scored as follows: 0 points: no redness and induration, 1 point: mild redness (diameter <5 mm), 2 points: moderate redness (diameter 5-10 mm), and 3 points: severe redness / induration / necrosis. The test results are as follows: Table 4: Results of propylthiouracil liposome injection irritation test

[0086] From the results of the above table, the propylthiouracil liposome injection of the present application and the negative control are not found obvious local reaction and pathological changes, and no blood vessel irritation.

[0087] Hemolysis test of test example 5

[0088] Take one experimental rabbit, take blood from the central ear artery, prepare 2% red blood cell suspension, and prepare 2mg / ml propylthiouracil solution with the propylthiouracil liposome injection prepared in Example 1 as solvent. Take 7 test tubes, number 1-7, 1-5 are sample tubes of Example 1, 6 is a negative control tube, and 7 is a positive control tube. According to the proportioning amount in Table 5, add 2% red blood cell suspension and 0.9% sodium chloride injection, pure water into the tubes in turn, mix well, and then immediately place in a 37℃ water bath box for 30min. Then add different amounts of 2mg / ml propylthiouracil solution, place in a 37℃ water bath box and observe for 3h, observe every 15min within 1h, observe every 1h within 2-3h, and after 3h, centrifuge the solution in each tube and test the absorbance value of each tube. The results are shown in Table 6 and Table 7. Table 5: In vitro red blood cell hemolysis test of experimental rabbits Table 6: Results of macroscopic observation of hemolysis test Table 7: Results of spectrophotometry

[0089] From the results of the above table, in the drug hemolysis test, no hemolysis and coagulation phenomenon occurred under different drug concentrations, and the hemolysis rate was less than 5%, indicating that the propylthiouracil liposome injection of the present application has no hemolytic and red blood cell aggregation reaction.

[0090] From the results of the above examples and test examples, the propylthiouracil liposome injection of the present application has good appearance, small particles, uniform particle size, high encapsulation efficiency, high stability, low leakage rate, high bioavailability, and the key quality attributes of the product can meet the requirements. During the stability sample (at 40℃) period, the product quality meets the standard, the product clinical use safety is higher, and it is suitable for industrial production.

[0091] The above content is a further detailed description of the present application in combination with specific embodiments, which cannot be considered as limiting the specific implementation of the present application to these descriptions. For ordinary skilled persons in the technical field to which the present application belongs, without departing from the concept of the present application, a number of simple deductions or substitutions can be made.

Claims

1. A liposomal injection of propylthiouracil, characterized by: The composition comprises the following ingredients by weight: propylthiouracil 100 parts, hydrogenated soybean lecithin 240-360 parts, cholesterol 80-120 parts, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000 30-70 parts, alpha-tocopherol 1-3 parts, sucrose 100-300 parts, and a buffer solution in an appropriate amount.

2. The liposomal injection of propylthiouracil according to claim 1, characterized by The dosage form is 50 mg or 100 mg.

3. The liposomal injection of propylthiouracil according to claim 1, characterized by The weight ratio of hydrogenated soybean lecithin to cholesterol is 3:

1.

4. The liposomal injection of propylthiouracil according to claim 1, characterized by The buffer solution used is preferably a phosphate buffer (pH 7.4).

5. A process for the preparation of the liposomal injection of propylthiouracil according to claim 1, characterized in that The method comprises the following steps: (1) Lipid film preparation: hydrogenated soybean lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol 2000, and alpha-tocopherol are added to a phosphate buffer solution with a pH of 7.4 in the appropriate proportions, and stirred and dissolved at 55-70°C to form a uniform lipid solution. (2) Drug-loaded hydration: propylthiouracil is added to the phosphate buffer solution at 55-70°C, stirred and dissolved, and then added to the lipid solution of step (1), and stirred for 2-4 hours to form multilayer liposomes. (3) High-pressure homogenization: the crude liposome suspension of step (2) is homogenized by a high-pressure homogenizer to form single-chamber liposomes, and the average particle size of the liposomes is controlled to be 80-160 nm. (4) Concentration and purification: the drug solution of step (3) is filtered by a tangential flow filtration system, and the cycle is filtered until the free drug content is less than 5%, and 8% sucrose phosphate buffer solution is continuously added during the cycle filtration process. (5) Sterilization filtration and dispensing: the drug solution of step (4) is passed through a 1.2 μm prefilter and a 0.22 μm sterile filter in sequence, and then filled into a Schott bottle. (6) Freeze-drying: first stage: cooling to -50°C to -30°C, and holding for 2-4 hours; second stage: warming to -25°C to -10°C, and holding for 8-12 hours, with a vacuum degree of 10-40 Pa; third stage: warming to 20°C to 60°C, and holding for 6-8 hours, with a vacuum degree of 10-30 Pa; and the final water content is controlled to be ≤2%.

6. The method of claim 5, wherein the liposomal injection of propylthiouracil is prepared by the steps of: The stirring and dissolving temperature of step (1) is preferably 60°C.

7. The method of claim 5, wherein the liposomal injection of propylthiouracil is prepared by the steps of: The average particle size of the liposomes in step (3) is preferably controlled to be in the range of 110-130 nm.

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