Enteric-coated sustained-release preparation of artesunate

Abstract

The present invention provides an enteric-coated sustained-release preparation of artesunate, the preparation comprising artesunate, L-arginine and a solid dispersion carrier, wherein the solid dispersion carrier is PEG4000. The present invention improves the water solubility and stability of artesunate through the synergistic effect of the alkalizing agent L-arginine and the solid dispersion carrier PEG400, improves the bioavailability of artesunate, reduces the number of administrations of artesunate drugs, and improves patient compliance.

Description

Technical Field

[0001] The invention relates to the technical field of pharmaceutical preparations, and in particular to an enteric-coated sustained-release preparation of artesunate. Background Art

[0002] Artesunate (AS) is one of the artemisinin derivatives with a sesquiterpene structure independently developed in my country. In addition to its significant effects on malaria and some blood protozoan diseases, it also has anti-tumor, anti-inflammatory, immunomodulatory, anti-fibrosis and other effects, and is not easy to produce drug tolerance, so it is very popular among people.

[0003] Artesunate is an odorless white crystalline powder belonging to Class 2 of the biopharmaceutics classification system (BCS). The chemical name of artesunate is dihydroartemisinin-1,2α-succinic acid monoester, and its molecular formula is C 19 H 28 O8, with a relative molecular weight of 384, is very soluble in ethanol, acetone, and dichloromethane, easily soluble in ethanol or ethyl acetate, and almost insoluble in water. It can always maintain a high biomembrane permeability under acid and alkaline conditions in vivo and in vitro. Artesunate is sensitive to acid, alkali, heat, oxygen and other environments, especially in acidic and alkaline environments. Its chemical structure is easily destroyed. In addition, artesunate has a short half-life in the body (about 30 minutes) and low bioavailability, which greatly limits its application.

[0004] Therefore, how to improve the water solubility and stability of artesunate, improve its bioavailability, reduce the number of dosing times, and improve patient compliance is one of the hot topics that technicians in this field continue to pay attention to. Summary of the invention

[0005] The invention provides an enteric-coated sustained-release preparation of artesunate, which is used for improving the water solubility and stability of artesunate, improving the bioavailability thereof, reducing the number of administrations and improving the compliance of patients.

[0006] The first aspect of the present invention provides an enteric-coated sustained-release preparation of artesunate, the preparation comprising a solid dispersion, the solid dispersion comprising artesunate, L-arginine and a solid dispersion carrier, and the solid dispersion carrier is PEG4000.

[0007] In a specific embodiment, the mass ratio of artesunate, L-arginine and PEG4000 is 1:1:1.

[0008] In a specific embodiment, the solid dispersion is prepared by the following preparation method:

[0009] The PEG4000 and L-arginine are dissolved in a solvent, artesunate is added and stirred evenly, and the solid dispersion is obtained after removing the solvent.

[0010] In a specific embodiment, the stirring is carried out at 30° C. and the stirring time is 1 h.

[0011] In a specific embodiment, the preparation further comprises a sustained-release material, and the sustained-release material is HPMC.

[0012] In a specific embodiment, based on 50 mg of artesunate, the mass of HPMC is 146 mg.

[0013] In a specific embodiment, the preparation further comprises a filler, and the filler is alpha lactose monohydrate.

[0014] In a specific embodiment, the preparation further comprises a lubricant, and the lubricant is talc.

[0015] In a specific embodiment, the preparation further comprises an enteric coating, and the enteric coating comprises Eudragit L100-55, triethyl citrate and talc.

[0016] In a specific embodiment, the weight gain percentage of the enteric coating is not less than 5%.

[0017] The present invention improves the water solubility and stability of artesunate through the synergistic effect of the alkalizing agent L-arginine and the solid dispersion carrier PEG400, improves the bioavailability of artesunate, reduces the number of times the artesunate drug is administered, and improves the compliance of patients. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 The saturated solubility test results of the complex and artesunate provided in one embodiment of the present invention in purified water at different pH values;

[0019] Figure 2 The test results of the cumulative dissolution percentage of artesunate in different solid dispersions provided in one embodiment of the present invention;

[0020] Figure 3 The test results of the cumulative dissolution percentage of artesunate in different solid dispersions provided in one embodiment of the present invention;

[0021] Figure 4 The test results of the cumulative dissolution percentage of artesunate in different solid dispersions provided in one embodiment of the present invention;

[0022] Figure 5 The cumulative dissolution percentage test results of artesunate at different temperatures for the solid dispersion provided by one embodiment of the present invention;

[0023] Figure 6 The solid dispersion provided in one embodiment of the present invention is a test result of the cumulative dissolution percentage of artesunate at different stirring times;

[0024] Figure 7 The test results of the cumulative dissolution percentage of artesunate in the solid dispersion provided by an embodiment of the present invention and the original drug;

[0025] Figure 8 The test results of the cumulative dissolution percentage of artesunate at different times for different sustained-release materials provided in one embodiment of the present invention;

[0026] Fig. 9 The test results of the cumulative dissolution percentage of artesunate at different times for different auxiliary materials provided in one embodiment of the present invention;

[0027] Fig.10 A three-dimensional effect diagram and a contour diagram of two factors and response values ​​of HPMC and α-lactose monohydrate provided in one embodiment of the present invention;

[0028] Fig.11 It is an enlarged contour diagram of two factors and response values ​​of HPMC and α-lactose monohydrate provided in one embodiment of the present invention;

[0029] Fig.12 The test results of the cumulative dissolution percentage of artesunate in the sustained-release tablets and the original drug provided in one embodiment of the present invention;

[0030] Fig.13 The cumulative dissolution percentage test results of artesunate when the enteric coating weight gain is 3%, 5%, and 7% provided in one embodiment of the present invention;

[0031] Fig.14 The test results of the cumulative dissolution percentage of artesunate in different dissolution media for the sustained-release tablets provided in one embodiment of the present invention;

[0032] Fig.15 This is a pharmacokinetic blood concentration diagram of an enteric-coated sustained-release preparation of artesunate provided in one embodiment of the present invention and a commercially available preparation. DETAILED DESCRIPTION

[0033] In order to make the purpose, technical solutions and advantages of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0034] Example 1

[0035] Artesunate and L-arginine were mixed in a mass ratio of 2:1 to obtain complex 1, and in a mass ratio of 1:1 to obtain complex 2. The saturated solubility of the complex and artesunate in purified water at different pH values ​​was determined by the saturated solution method. The test results are shown in FIG. Figure 1 shown.

[0036] according to Figure 1 It can be seen that L-arginine helps to improve the saturated solubility of artesunate under different pH environments. At the same time, the mass ratio of L-arginine to artesunate has little effect on the improvement of the saturated solubility of artesunate.

[0037] Example 2

[0038] Artesunate was mixed with polyvinyl alcohol 188, HPMC, poloxamer 188, and PEG4000 at a mass ratio of artesunate: carrier material = 1:3 to obtain a solid dispersion. The prepared solid dispersion was subjected to a dissolution test according to the following method, and the cumulative dissolution percentage of artesunate at different times was calculated. The test results are shown in FIG. Figure 2 shown.

[0039] The dissolution conditions under the artesunate tablets in Part II of the 2020 edition of the Pharmacopoeia of the People's Republic of China were used as a reference. 900 mL of phosphate buffer with a pH of 6.8 was used as the dissolution medium, the medium temperature was 37±0.5°C, the rotation speed was 75r / min, 3 mL of samples were taken at 10, 20, 30, 40, and 50 min, and 3 mL of phosphate buffer with a pH of 6.8 at the same temperature was added to the operating container in time, filtered through a 0.45μm microporous membrane, the initial filtrate was discarded, and an appropriate amount of the subsequent filtrate was taken for HPLC detection, and the cumulative dissolution percentage of artesunate at different times was calculated, and the cumulative time-dissolution percentage curve was drawn.

[0040] Chromatographic conditions: Chromatographic column: Diamonsil C18 (2) (5 μm, 4.6 mm × 200, Dikma Technologies); Detector: SPD-20A UV / visible detector; Mobile phase: acetonitrile-phosphate buffer = 50:50 (take 1.36 mg of potassium dihydrogen phosphate, add 900 mL of water to dissolve, adjust the pH to 3.0 with 10% phosphoric acid, and add water to 1000 mL); Column temperature: room temperature (25 ° C); Injection volume: 20 μL; Detection wavelength: 210 nm; Flow rate: 1.2 mL / min.

[0041] according to Figure 2It can be seen that the dissolution effect of artesunate is different in the solid dispersions prepared with different carrier materials. The carrier materials are ranked according to the dissolution effect of artesunate, and it can be seen that PEG4000>polyvinyl alcohol 188>poloxamer 188>HPMC. Within 50 minutes of the solid dispersion with PEG4000 as the carrier, the cumulative dissolution rate of artesunate is about 90%, indicating that adding a solid dispersion carrier helps to improve the solubility of artesunate, and PEG4000 has the best dissolution effect.

[0042] Example 3

[0043] Artesunate, L-arginine, polyvinyl alcohol 188, HPMC, poloxamer 188, and PEG4000 were mixed at a mass ratio of artesunate: L-arginine: carrier material = 1:1:3 to obtain a solid dispersion. A dissolution test was performed on the prepared solid dispersion in the same manner as in Example 2, and the cumulative dissolution percentage of artesunate at different times was calculated. The test results are shown in FIG. Figure 3 shown.

[0044] according to Figure 3 It can be seen that the dissolution effect of artesunate is improved to varying degrees after the addition of L-arginine, and the solid dispersion with PEG4000 as the carrier has the best dissolution effect within 50 minutes, indicating that L-arginine and PEG4000 have a synergistic effect on improving the solubility of artesunate.

[0045] Example 4

[0046] According to the experimental results of Example 3, PEG4000 was selected as the carrier, and the carrier: artesunate: L-arginine were mixed at a mass ratio of 2:2:1, 4:2:1, 6:2:1, and 8:2:1 to prepare a solid dispersion. The same method as in Example 2 was used to perform a dissolution test on the prepared artesunate solid dispersion. The experimental results are as follows: Figure 4 shown.

[0047] according to Figure 4 It can be seen that in the solid dispersion, the larger the proportion of carrier PEG4000, the more conducive it is to the dissolution of artesunate, but excessive use of the carrier leads to an increase in the cost of the preparation and is not conducive to the preparation of the artesunate dosage form in the later stage. Therefore, considering various factors, the next experiment was carried out with a mass ratio of PEG4000, artesunate and L-arginine of 2:2:1.

[0048] Example 5

[0049] The mass ratio of carrier: artesunate: L-arginine was 2:2:1, and artesunate solid dispersions were prepared at 25, 35, 45, and 55° C., respectively. The dissolution test was performed on the prepared solid dispersions in the same manner as in Example 2. The experimental results are shown in FIG. Figure 5 shown.

[0050] according to Figure 5 It can be seen that as the temperature increases, the dissolution effect becomes worse. This may be due to the physical and chemical properties of artesunate itself. In a hot and humid environment, its chemical structure is prone to destructive changes and is very unstable. Under a higher preparation environment, the drug has been degraded, so 25°C was selected as the preparation temperature for the next investigation.

[0051] Example 6

[0052] The solid dispersion was prepared with a mass ratio of carrier: artesunate: L-arginine of 2:2:1 and a preparation temperature of 25°C. The solid dispersions were stirred for 1 h, 2 h, 3 h, and 4 h, respectively. The dissolution test was performed on the prepared solid dispersions in the same manner as in Example 2. The experimental results are shown in FIG. Figure 6 shown.

[0053] according to Figure 6 It can be seen that as the stirring time increases, the cumulative dissolution becomes relatively smaller. This may be due to the degradation of part of the drug in a hot and humid environment for a long time, so 1h was selected as the stirring time for the next investigation.

[0054] Example 7

[0055] The basic prescription process was determined according to Examples 1, 2, 3, 4, 5, and 6: 30 mg of L-arginine and 60 mg of PEG4000 were weighed and added to 30 mL of purified water to fully dissolve, and then 60 mg of artesunate was added. The mixture was stirred at 500 rpm for 1 h at 25 ° C, pre-frozen at -20 ° C for 6 h, then transferred to -80 ° C for freezing overnight, and then transferred to a freeze dryer for freeze drying for 48 h to remove the solvent. The mixture was sealed with an EP tube to block light and refrigerated.

[0056] According to the orthogonal experimental design table L9(3 4 ) was used to conduct experiments, with the cumulative dissolution percentage in 50 min as the evaluation index, and the best prescription was selected. The factor level table is shown in Table 1, the experimental design and results are shown in Table 2, and the variance analysis table is shown in Table 3.

[0057] Table 1 Factor level table

[0058]

[0059]

[0060] Table 2L9(34) Experimental design and results

[0061]

[0062] Table 3 Analysis of variance table

[0063]

[0064] According to Tables 1, 2 and 3, the order of influence of the factors on the preparation of artesunate solid dispersion is A>C>B>D, and the best combination is A3B1C2D3, that is, the ratio of L-arginine:drug is 2:1, the ratio of carrier:drug is 1:1, the stirring time is 1h, and the stirring temperature is 30℃.

[0065] The optimal formulation solid dispersion uses a large amount of L-arginine, and the prepared solid dispersion is very easy to absorb water and liquefy, which is not conducive to the development of later preparations of artesunate. At the same time, considering factors such as preparation cost and tablet weight, the ratio of L-arginine:drug was adjusted to 1:1.

[0066] Example 8

[0067] Determine the optimal prescription process according to Example 7: weigh 60 mg L-arginine and 60 mg PEG4000, add to 30 mL purified water, fully dissolve, then add 60 mg ART, stir magnetically at 500 rpm for 1 hour at 30°C, pre-freeze at -20°C for 6 hours, then transfer to -80°C for freezing overnight, then transfer to a freeze dryer for freeze drying for 48 hours to remove the solvent, seal with an EP tube to block light and refrigerate, and you're done. Prepare 3 batches in parallel. The saturated solubility of artesunate in release media with different pH values ​​was investigated at different times compared with the original artesunate tablets (Guilin Nanyao Co., Ltd., specification: 50 mg). The results are shown in Table 5, and the cumulative dissolution percentage is shown in Table 5. Figure 7 .

[0068] Table 5

[0069]

[0070] Example 9

[0071] 150 mg of the solid dispersion prepared in Example 8 was used to select HPMC, EC, and three different ratios of sustained-release materials for investigation. The investigation method is as follows. The prescription ratio is shown in Table 6. The dissolution results are shown in Figure 8 .

[0072] The dissolution and release rate determination method (General Rule 0931 Method 2) of Part IV of the 2020 edition of the Pharmacopoeia of the People's Republic of China was used as the dissolution medium. 900 mL of phosphate buffer with a pH of 6.8 was used as the dissolution medium, the medium temperature was 37±0.5°C, the rotation speed was 75 r / min, 3 mL of samples were taken at 2, 4, 6, 8, 10, and 12 h, and 3 mL of phosphate buffer with a pH of 6.8 at the same temperature was added to the operating container in time, filtered through a 0.45 μm microporous filter membrane, the initial filtrate was discarded, and an appropriate amount of the subsequent filtrate was taken for HPLC detection, and the cumulative dissolution percentage of artesunate at different times was calculated, and the cumulative time-dissolution percentage curve was drawn.

[0073] Chromatographic conditions: Chromatographic column: Diamonsil C18 (2) (5 μm, 4.6 mm × 200, Dikma Technologies); Detector: SPD-20A UV / visible detector; Mobile phase: acetonitrile-phosphate buffer = 50:50 (take 1.36 mg of potassium dihydrogen phosphate, add 900 mL of water to dissolve, adjust the pH to 3.0 with 10% phosphoric acid, and add water to 1000 mL); Column temperature: room temperature (25 ° C); Injection volume: 20 μL; Detection wavelength: 210 nm; Flow rate: 1.2 mL / min.

[0074] Table 6 Composition of sustained-release tablets with different sustained-release materials and dosages

[0075]

[0076] According to Table 6, the sustained-release tablets prepared by the six prescriptions all have good finished product appearance, hardness between 9-12kgf, and little weight difference, which meets the requirements. The specific properties of the preparations are determined by referring to the dissolution conditions. Figure 8 Comparing the dissolution curves shown, the sustained-release effect of the sustained-release material HPMC is better than that of EC, and the sustained-release effect is better as the amount of HPMC increases. Considering the later tablet weight and preparation cost, the HPMC dosage of 125 mg was finally selected for the next step of optimization.

[0077] Example 8

[0078] According to Example 7, the amount of solid dispersion and sustained-release material in the sustained-release tablet was determined, and three fillers with different proportions of soluble starch, α-lactose monohydrate, and microcrystalline cellulose were selected for investigation. The formulation composition is shown in Table 7, and the dissolution results are shown in Fig. 9 .

[0079] Table 7 Composition of sustained-release tablets with different fillers and dosages

[0080]

[0081] According to Table 7, soluble starch has poor compressibility and is easy to break into pieces; microcrystalline cellulose itself can be used as a disintegrant, which will accelerate the disintegration of tablets; therefore, α-lactose monohydrate has the best sustained release effect as a filler. Combined with the results of the dissolution curve, α-lactose monohydrate was used as the final filler.

[0082] The prescription was optimized by using a central point design experiment. HPMC was selected as the sustained-release material, α-lactose monohydrate as the filler, and talcum powder as the lubricant. The dosage of HPMC (X1) and α-lactose monohydrate (X2) were used as two factors, and 5 levels of central point experiments were set up to investigate the cumulative release of 2h (R1), 6h (R2), and 12h (R3). The total score (Y) = (1 / R1+1 / R2)*1900 was used as the evaluation index. The experimental results were processed using Design-Expert 8.0.6 software, and the response surface method was used to optimize the optimal preparation process. The factor level table and experimental design are shown in Tables 8 and 9.

[0083] Table 8 Factor level table

[0084]

[0085] Table 9 Experimental design and results

[0086]

[0087] The data in the table were subjected to a secondary multiple regression fitting analysis using Design-Expert 8.0.6 software, and the regression equation was obtained as Y = 92.32 + 7.50*A-5.04*B-13.43*A2-11.28*B2 (R2 = 0.9718, R 2adj =0.9517) shows that the model has good fitting degree and small experimental error, and this model can be used for analysis and prediction. The significance test of quadratic polynomial fitting and variance analysis are shown in Table 10.

[0088] Table 10 Analysis of variance table

[0089]

[0090]

[0091] *P<0.05, significant difference; **P<0.01, highly significant difference; ***P<0.0001, extremely significant difference.

[0092] According to the fitted quadratic polynomial model, the three-dimensional effect diagram and contour diagram of HPMC (A) and α-lactose monohydrate (B) and the response value were drawn using Design-Expert 8.0.6 software to screen the optimal prescription level. The influence of each factor on the response value is shown in Fig.10, 11 .from Fig.10 From the three-dimensional effect diagram, we can see that the change from the blue area at the edge to the red area in the center indicates the change from less to more extraction quality. The faster the change, the greater the slope, that is, the more significant the test result. Fig.10 The two-dimensional contour map shown is similar to Fig.11 The enlarged contour map shown is elliptical, indicating that the interaction between the factors is significant.

[0093] The three-dimensional effect surface plot shows the areas of maximum and minimum response. Fig.10 It can be seen that within the test range, the comprehensive score increases first and then decreases with the increase of HPMC (A) and α-lactose monohydrate (B), indicating that both factors have optimal values.

[0094] In summary, Design-Expert 8.0.6 software was used to set the response value to the maximum value to obtain the best prescription conditions. The best conditions after optimization are shown in Table 11.

[0095] Table 11. Composition and dosage of sustained-release tablet core

[0096]

[0097] Example 9

[0098] According to the optimal formulation process of sustained-release tablets predicted by the star-point design in Example 8, its dissolution was verified. The results are shown in Fig.12 .

[0099] according to Fig.12 It can be seen that the true value is basically consistent with the predicted value, and the reproducibility is good, indicating that the prescription conditions of artesunate solid dispersion tablets optimized by central composite design-response surface methodology are reasonable and reliable, and the preparation process adopted is feasible.

[0100] Example 10

[0101] Enteric coating prescription: Dissolve 6.2g of Eudragit L100-55 in 50g of anhydrous ethanol and add 0.6g of triethyl citrate. Also weigh 1.5g of talcum powder that has passed 80 mesh and add it to 50g of anhydrous ethanol. Use a high shear dispersing emulsifier to fully homogenize for about 10 minutes. Slowly pour the homogenized talcum powder suspension into the Eudragit L100-55 solution, stir thoroughly and coat. Investigate the effect of enteric coating weight gain of 3%, 5%, and 7% on the cumulative dissolution of the drug. Results are shown in Fig.13 The formula for calculating enteric coating weight gain is as follows:

[0102] Enteric coating weight gain percentage = (average tablet weight after coating - average tablet weight of plain tablets) / average tablet weight of plain tablets

[0103] according to Fig.13 The drug release curve shows that when the enteric coating weight increases by 3%, the drug is released in the acidic medium in the first 2 hours. As the thickness of the enteric coating increases, when the enteric coating weight increases by 5% and 7%, the drug release in the first 2 hours is basically 0. Therefore, the weight gain of the enteric coating needs to be above 5% to ensure that the drug is not released in the stomach. This paper finally chooses 5% as the weight gain of the enteric coating.

[0104] The final enteric coating formulation is shown in Table 12.

[0105] Table 12 Enteric coating prescription

[0106]

[0107] Embodiment 11

[0108] Through the above examples, the optimal process formula of ART solid dispersion enteric-coated sustained-release tablets was finally determined, as shown in Table 13. Three batches of ART enteric-coated sustained-release tablets were prepared, and the reproducibility of the preparation process was investigated. The results are shown in Fig.14 .

[0109] Table 13 Optimal process formula for artesunate solid dispersion enteric-coated sustained-release tablets

[0110]

[0111] Example 12

[0112] The enteric-coated sustained-release artesunate preparation prepared in Example 11 was used for pharmacokinetic test:

[0113] Twelve 200-250g SD male rats were selected and fed adaptively for one week. They were fasted for 12 hours before the experiment but not water. After fasting for 24 hours, they were randomly divided into a control group and an experimental group, with 6 rats in each group, and they were allowed to drink water freely.

[0114] The mice in the control group were orally administered with a commercially available drug (Guilin Pharmaceutical Co., Ltd.), and the mice in the experimental group were orally administered with the enteric-coated sustained-release preparation (PO) of artesunate prepared in Example 7. Blood was collected from the eye sockets at different time points after administration, and the serum was separated and processed with plasma. The content of artesunate was analyzed by liquid chromatography-mass spectrometry. The analysis results are as follows: Fig.15 As shown in Table 14, the sustained-release tablets provided by the present invention can increase the half-life t1 / 2 of the drug, and at the same time also improve the MRT, prolonging the retention time of the drug to achieve a sustained-release effect, and compared with commercially available preparations, it is increased by 232.76%, which can significantly improve the bioavailability of the drug.

[0115] Table 14 Pharmacokinetic parameters (n=6)

[0116]

[0117] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. An enteric-coated sustained-release preparation of artesunate, characterized in that: The preparation comprises a solid dispersion, HPMC, alpha lactose monohydrate and talc, the solid dispersion comprises artesunate, L-arginine and a solid dispersion carrier, the solid dispersion carrier is PEG4000, and the mass ratio of artesunate, L-arginine and PEG4000 is 1:1:

1.

2. The preparation according to claim 1, characterized in that The solid dispersion is prepared by the following preparation method: The PEG4000 and L-arginine are dissolved in a solvent, artesunate is added and stirred evenly, and the solid dispersion is obtained after removing the solvent.

3. The preparation according to claim 2, characterized in that The stirring was carried out at 30° C. for 1 h.

4. The preparation according to claim 1, characterized in that Based on every 50 mg of artesunate, the mass of the HPMC is 146 mg.

5. The preparation according to claim 1, characterized in that The preparation further comprises an enteric coating, which comprises Eudragit L100-55, triethyl citrate and talc.

6. The preparation according to claim 5, characterized in that The weight gain percentage of the enteric coating is not less than 5%.

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