Analysis method of azilsartan raw material and 14 related substances in preparation of azilsartan raw material

Abstract

The invention discloses a method for analyzing 14 related substances in an azilsartan raw material and a preparation of the azilsartan raw material, belongs to the technical field of pharmaceutical analysis, and is characterized in that impurities in the azilsartan raw material and the preparation of the azilsartan raw material are detected by adopting a high performance liquid chromatography, and the method aims at solving the problem of detection of the azilsartan related substances. The invention provides a convenient, efficient and accurate detection method, the method can be used for simultaneously analyzing 14 related impurities in azilsartan raw materials and preparations thereof, the content of known impurities is effectively controlled by a self-contrast method with correction factors, the separation degree between each impurity and a main component is greater than 1.5, and the detection result is accurate. The method is high in sensitivity, simple and rapid, strong in specificity, good in accuracy, high in precision, good in stability and good in linear relation, and can be used for quality control of azilsartan raw materials and preparations thereof.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical analysis, and particularly to an analytical method for 14 related substances in azilsartan raw materials and its preparations. Background Art

[0002] The chemical name of azilsartan is 2-ethoxy-1-[[2'-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)biphenyl-4-yl]methyl]-1H-benzimidazole-7-carboxylic acid (CAS No.: 147403-03-0), and its chemical structural formula is as follows.

[0003] Azilsartan is an angiotensin II receptor blocker (ARBs) developed by Takeda Pharmaceutical Company of Japan, which exerts its antihypertensive effect by blocking the vasopressor hormone angiotensin II. Compared with angiotensin-converting enzyme inhibitor (ACEI) antihypertensive drugs, this product has the advantages of stable antihypertensive effect and no dry cough. Research shows that as a new generation of dual-function ARBs, azilsartan can not only reduce blood pressure stably, but also reduce the risks of cardiovascular diseases and diabetes.

[0004] In order to ensure the safety and effectiveness of drugs, it is necessary to study, detect and monitor the related substances in drug raw materials and their preparations. Patents CN103743826B and CN106841415B both disclose a high-performance liquid chromatography analysis method for azilsartan, which can effectively detect azilsartan and its related substances.

[0005] The inventor of the present invention studied azilsartan and found that the azilsartan raw materials and their preparations may contain the following impurities: Among them, impurity 1 is the starting material, and impurities 2 and 3 are reaction intermediates. Impurity 1, impurity 2, impurity 3, impurity 4, impurity 5, impurity 7, impurity 8, and impurity 10 are the same as impurities D, E, J, A, C, B, F, and G in patent CN103743826B respectively. The remaining 6 impurities are different, and the remaining 6 impurities are also different from the impurities in patent CN106841415B.

[0006] Due to different synthesis processes of drugs and different prescriptions of preparations, the impurity profiles of drugs will also change. The analytical methods provided by patents CN103743826B and CN106841415B are not applicable to the detection of related substances in all azilsartan raw materials and their preparations. Therefore, it is necessary to establish a suitable analytical method according to different synthesis processes and preparation prescriptions to achieve accurate and effective detection of related substances in azilsartan raw materials and their preparations. SUMMARY OF THE INVENTION

[0007] In order to solve the problems in the prior art, the present invention provides a method for analyzing 14 related substances in azilsartan raw materials and their preparations. The analysis method of the present application has good specificity, high sensitivity, and can quickly, effectively and accurately perform quantitative analysis on 14 related substances in azilsartan raw materials and their preparations.

[0008] The method for analyzing 14 related substances in azilsartan raw materials and their preparations provided by the present invention adopts the following technical solutions: A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is detected by high performance liquid chromatography. The chromatographic conditions include: mobile phase A is potassium dihydrogen phosphate buffer solution, and mobile phase B is acetonitrile; The gradient elution program is: 25%-65% B from 0 to 30 min, 65% B from 30 to 40 min, 65%-25% B from 40 to 41 min, and 25% B from 41 to 50 min; The 14 related substances involved in the azilsartan raw materials and preparations are: Among them, impurity 1 is the starting material, impurities 2 and 3 are reaction intermediates, impurities 4, 5, 6, 8, 9 are process impurities, impurities 12, 13, 14 are degradation impurities, and impurities 7, 10, 11 are both process impurities and degradation impurities.

[0009] Preferably, the high performance liquid chromatography conditions further include that the chromatographic column is a chromatographic column filled with octadecylsilane-bonded silica gel, and more preferably Kromasil 100-5-C18 4.6 mm×250 mm, 5 μm.

[0010] Preferably, the high performance liquid chromatography conditions further include that the flow rate is 0.9-1.1 mL / min, the injection volume is 10 μL, and the column temperature is 28-32 °C.

[0011] Preferably, the detection wavelength in the high performance liquid chromatography conditions is 220 nm.

[0012] Preferably, the pH value of the potassium dihydrogen phosphate buffer solution of mobile phase A is 2.8-3.2.

[0013] Preferably, it further includes the preparation of the test solution. Weigh the test sample, dissolve it with a solvent and dilute it with a diluent to a solution containing 1 mg of azilsartan per 1 mL.

[0014] Preferably, the solvent is composed of a 0.02 mol / L dipotassium hydrogen phosphate solution and acetonitrile in a volume ratio of 50:50, and the diluent is composed of a 0.272 wt% potassium dihydrogen phosphate solution and acetonitrile in a volume ratio of 75:25.

[0015] Preferably, the pH value of the dipotassium hydrogen phosphate solution in the solvent is 8.0, and the pH value of the potassium dihydrogen phosphate solution in the diluent is 3.0.

[0016] In summary, the present invention has the following beneficial effects: The analytical method of the present application has high sensitivity, strong specificity, good accuracy, high precision, high stability, and good linear relationship, and can effectively and accurately perform quantitative analysis on 14 related substances in azilsartan raw materials and preparations. Description of the Drawings

[0017] Figure 1 It is the chromatogram of the system suitability solution in Example 1.1 of the present application.

[0018] Figure 2 It is the chromatogram of the test sample of azilsartan raw material in Example 2.1 of the present application.

[0019] Figure 3 It is the chromatogram of the test sample of azilsartan tablets in Example 2.2 of the present application. Detailed Embodiments

[0020] The present invention will be further described in detail below with reference to the drawings. All reagents not indicating the manufacturer can be obtained as conventional reagent products through commercial purchase.

[0021] Example 1.1 An analytical method for 14 related substances in azilsartan raw materials and their preparations includes the following steps: 1.1 Instruments and Detection Conditions High performance liquid chromatograph Column model: The packing material is octadecylsilane-bonded silica gel (Kromasil 100-5-C18 4.6×250 mm, 5 μm); Mobile phase A: Potassium dihydrogen phosphate buffer solution (take 2.72 g of potassium dihydrogen phosphate, dissolve it in water and dilute it to 1000 mL, adjust the pH value to 3.0 with phosphoric acid); Mobile phase B: Acetonitrile; Gradient elution is used for elution, and the elution program is shown in the following table: The flow rate is 1.0 mL / min, the injection volume is 10 μL, and the column temperature is 30 °C.

[0022] 1.2 Solution preparation requires operation under light protection. On the premise that the target concentration and weighing accuracy of all solutions meet the requirements, their volumes can be enlarged or reduced to meet the reagent needs.

[0023] 1.2.1 Preparation of system suitability solution Take appropriate amounts of reference substances of impurity 1, impurity 2, impurity 3, impurity 4, impurity 6, impurity 10, and impurity 11, weigh accurately, dissolve in acetonitrile - water (90:10) and dilute to a solution containing about 10 μg of impurity 1, impurity 2, impurity 3, impurity 4, impurity 6, impurity 10, and impurity 11 per 1 mL as impurity stock solution 1; Take appropriate amounts of reference substances of impurity 5, impurity 7, impurity 8, impurity 9, impurity 12, impurity 13, and impurity 14, weigh accurately, dissolve in an appropriate amount of N,N - dimethylformamide, and dissolve and dilute with N,N - dimethylformamide - water (9:1) to make a solution containing about 10 μg of impurity 5, impurity 7, impurity 8, impurity 9, impurity 12, impurity 13, and impurity 14 per 1 mL as impurity stock solution 2; Take 20 mg of azilsartan reference substance, weigh accurately, place it in a 20 mL brown volumetric flask, add 10 mL of solvent (the solvent is composed of 0.02 mol / L dipotassium hydrogen phosphate solution (pH 8.0) and acetonitrile in a volume ratio of 50:50) to dissolve, accurately add 2 mL of each of impurity stock solution 1 and impurity stock solution 2, dilute to the mark with diluent (composed of 0.272 wt% potassium dihydrogen phosphate solution (pH 3.0) and acetonitrile in a volume ratio of 75:25), shake well to make a solution containing about 1 mg of azilsartan and about 1 μg of each impurity per 1 mL as the system suitability solution.

[0024] Experimental operation: Inject 10 μL of the system suitability solution, record the typical chromatogram. The typical chromatogram Figure 1 As shown, it can be seen from Figure 1 that the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0025] Example 1.2 A method for analyzing 14 related substances in azilsartan raw material and its preparations, which is different from Example 1.1 in that the flow rate is 0.9 mL / min, and the others are the same as in Example 1.1. The typical chromatogram obtained from the experimental operation is the same as the typical chromatogram in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0026] Example 1.3 A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is different from Example 1.1 in that the flow rate is 1.1 mL / min, and the others are the same as in Example 1.1. The typical chromatogram obtained by the experimental operation is the same as that in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0027] Example 1.4 A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is different from Example 1.1 in that the column temperature is 28 °C, and the others are the same as in Example 1.1. The typical chromatogram obtained by the experimental operation is the same as that in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0028] Example 1.5 A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is different from Example 1.1 in that the column temperature is 32 °C, and the others are the same as in Example 1.1. The typical chromatogram obtained by the experimental operation is the same as that in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0029] Example 1.6 A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is different from Example 1.1 in that the pH of the potassium dihydrogen phosphate solution in mobile phase A is 2.8, and the others are the same as in Example 1.1. The typical chromatogram obtained by the experimental operation is the same as that in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0030] Example 1.7 A method for analyzing 14 related substances in azilsartan raw materials and their preparations, which is different from Example 1.1 in that the pH of the potassium dihydrogen phosphate solution in mobile phase A is 3.2, and the others are the same as in Example 1.1. The typical chromatogram obtained by the experimental operation is the same as that in Example 1.1, and the resolution between each impurity peak, between the azilsartan main peak and its adjacent impurity peaks is greater than 1.5.

[0031] Example 2.1 A method for analyzing 14 related substances in azilsartan raw materials and their preparations includes the following steps: Sample preparation Test solution: Weigh accurately about 20 mg of azilsartan raw material test sample, place it in a 20 mL brown volumetric flask, add 10 mL of solvent (the solvent is composed of 0.02 mol / L dipotassium hydrogen phosphate solution (pH 8.0) and acetonitrile in a volume ratio of 50:50) to dissolve it, and then dilute it to the mark with diluent (composed of 0.272 wt% potassium dihydrogen phosphate solution (pH 3.0) and acetonitrile in a volume ratio of 75:25), shake well to obtain the test solution.

[0032] Experimental operation: Inject 10 mL of the test solution into the high performance liquid chromatograph. The instrument parameters are the same as those in Example 1.1, record the chromatogram, as Figure 2 shown.

[0033] Example 2.2 An analytical method for 14 related substances in azilsartan raw material and its preparations, comprising the following steps: Sample preparation Test solution: Take 10 tablets of azilsartan tablets with a specification of 20 mg, grind them finely, take an appropriate amount of the fine powder (equivalent to about 20 mg of azilsartan), weigh accurately, place it in a 20 mL brown volumetric flask, add 10 mL of solvent (the solvent is composed of 0.02 mol / L dipotassium hydrogen phosphate solution (pH 8.0) and acetonitrile in a volume ratio of 50:50) to dissolve it, and then dilute it to the mark with diluent (composed of 0.272 wt% potassium dihydrogen phosphate solution (pH 3.0) and acetonitrile in a volume ratio of 75:25), shake well, filter, and take the filtrate as the test solution.

[0034] Experimental operation: Inject 10 mL of the test solution into the high performance liquid chromatograph. The instrument parameters are the same as those in Example 1.1, record the chromatogram, as Figure 3 shown.

[0035] Example 3 Verify the analytical method of Example 1.1 to prove that the established method is suitable for the corresponding detection requirements. The verification process is as follows: 3.1 Specificity verification The inventor of the present invention screened and optimized the mobile phase with appropriate components and ratios for chromatographic analysis of azilsartan and the above 14 impurities, determined the analytical method of the present invention, and verified the specificity of the present invention through peak location tests of the starting material (Impurity 1), reaction intermediates (Impurities 2 and 3), azilsartan, and degradation experiments of azilsartan. The test results are shown in Table 1 and Figure 1 shown.

[0036] Table 1 Specificity verification results Name Retention Time (min) Resolution Theoretical Plate Number Impurity 14 8.894 / 11624 Impurity 5 9.740 2.844 21625 Impurity 12 11.315 6.278 36486 Impurity 7 14.746 11.759 28963 Impurity 2 16.397 5.766 84244 Impurity 8 17.147 3.090 69707 Impurity 9 19.185 8.150 101890 Impurity 6 21.830 11.047 134175 Azilsartan 22.407 2.148 89707 Impurity 4 24.083 6.026 140678 Impurity 13 27.143 11.663 164183 Impurity 11 27.852 2.681 182041 Impurity 3 29.788 7.317 197548 Impurity 10 32.561 10.187 222205 Impurity 1 37.235 15.670 216273 As can be seen from the above table, the resolution between each impurity peak, between the main peak of azilsartan and its adjacent impurity peaks is greater than 1.5, and the theoretical plate number of the main peak of azilsartan is not less than 3000. The impurities and the main peak can be effectively separated, indicating that the specificity of the analysis method of this application is good.

[0037] The inventor of the present invention also studied the commonly used pharmaceutical excipients for preparing azilsartan pharmaceuticals on the market and found that the commonly used pharmaceutical excipients have no interference with this application.

[0038] 3.2 Detection Limit and Quantitation Limit The inventor of the present invention detected the detection limit, quantitation limit, linearity, and correction factor of azilsartan and 14 impurities, and carried out sensitivity and quantitative analysis verification. The results are shown in Table 2.

[0039] Table 2 Quantitative Analysis Verification Results of Azilsartan and Each Impurity As can be seen from the above table, the sensitivity of the detection of azilsartan and each impurity in this application is relatively high, and the linear relationship of each impurity is good in the extremely low concentration range. The correlation coefficient R is greater than 0.990. The correction factors of impurities 1-13 are all in the range of 0.8-1.2, and the correction factor is calculated as 1.0. The correction factor of impurity 14 is calculated as 1.5. The analysis method of this application can accurately control the content of each impurity.

[0040] In addition, the detection limit and quantitation limit of this application are relatively small, indicating that the sensitivity of this application is good. This application uses the self-control method of the main component with correction factor to quantitatively analyze the above 14 impurities, which improves the accuracy of the related substance detection of this application. It also shows that the analysis method of this application can effectively and accurately quantitatively analyze the related impurities in azilsartan raw materials and preparations.

[0041] 3.3 Stability Test The solution stability of azilsartan and each impurity was studied. A mixed solution was prepared from azilsartan and 14 impurities. The concentration of azilsartan and each impurity in the mixed solution was 1 μg / ml. The detection results are shown in Table 3.

[0042] Table 3 Determination Results of the Stability Test of the Mixed Solution of Azilsartan and Each Impurity As can be seen from the above table, the solutions of azilsartan and each impurity are stable within 63 h, indicating that the solution stability of this application is relatively good and can stably and effectively quantitatively analyze the related substances in azilsartan raw materials and their preparations.

[0043] The embodiments of this specific implementation manner are all preferred embodiments of the present invention, and do not limit the protection scope of the present invention accordingly. Therefore, all equivalent changes made according to the structure, shape, and principle of the present invention shall be covered within the protection scope of the present invention.

Claims

1. An analytical method for 14 related substances in azilsartan raw materials and its preparations, characterized in that: Detection was carried out by high performance liquid chromatography. The chromatographic conditions included: mobile phase A was potassium dihydrogen phosphate buffer solution, and mobile phase B was acetonitrile; The gradient elution program was: 25%-65% B from 0 to 30 min, 65% B from 30 to 40 min, 65%-25% B from 40 to 41 min, and 25% B from 41 to 50 min; The 14 related substances involved in the azilsartan raw material and its preparations were:

2. The analytical method for 14 related substances in azilsartan raw materials and its preparations according to claim 1, wherein: The high performance liquid chromatography conditions also included a chromatographic column with octadecylsilane chemically bonded silica gel as the packing material.

3. The analytical method for 14 related substances in an azilsartan raw material and its preparation according to claim 1, characterized in that: The high performance liquid chromatography conditions also included a flow rate of 0.9 - 1.1 mL / min, an injection volume of 10 μL, and a column temperature of 28 - 32 °C.

4. The analytical method for 14 related substances in an azilsartan raw material and its preparation according to claim 1, characterized in that: The detection wavelength in the high performance liquid chromatography conditions was 220 nm.

5. The analytical method for 14 related substances in azilsartan raw materials and its preparations according to claim 1, characterized in that: The pH value of the potassium dihydrogen phosphate buffer solution of mobile phase A was 2.8 - 3.

2.

6. The analytical method for 14 related substances in an azilsartan raw material and its preparation according to claim 1, characterized in that: It also included the preparation of the test solution. Weigh the test sample, dissolve it with a solvent and dilute it with a diluent to a solution containing 1 mg of azilsartan per 1 mL.

7. The analytical method for 14 related substances in an azilsartan raw material and its preparation according to claim 6, characterized in that: The solvent was composed of 0.02 mol / L potassium hydrogen phosphate solution and acetonitrile in a volume ratio of 50:50, and the diluent was composed of 0.272 wt% potassium dihydrogen phosphate solution and acetonitrile in a volume ratio of 75:

25.

8. The analytical method for 14 related substances in an azilsartan raw material and its preparation according to claim 7, characterized in that: The pH value of the potassium hydrogen phosphate solution in the solvent was 8.0, and the pH value of the potassium dihydrogen phosphate solution in the diluent was 3.0.

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