Quality control method of fenerenone

Abstract

The invention discloses a quality control method of fenerenone, which is characterized in that specific fenerenone impurities are subjected to gradient elution by adopting a high performance liquid chromatography, and the quality condition of the fenerenone can be accurately and effectively reflected. The method is simple and convenient to operate, meets the quality control requirement, and can be used for the whole process of finerenone preparation and quality control of raw material medicines and preparation products thereof.

Description

Technical Field

[0001] The present invention relates to a quality control method for finerenone, in particular to a quality control method for finerenone and related impurities thereof. Background Art

[0002] Type 2 diabetes mellitus (T2DM) is a major risk factor for chronic kidney disease (CKD) and warrants significant attention. CKD is a clinical syndrome characterized by persistent abnormalities in kidney structure and function. When T2DM and CKD coexist, the risk of cardiovascular disease increases. This is because persistently high blood sugar levels can cause systemic vascular damage, which in turn adversely affects the heart, eyes, kidneys, and nerves, leading to various complications. Diabetic nephropathy (DKD) is one of the common complications of diabetes.

[0003] For patients with T2DM who have CKD, proteinuria symptoms, and are at high risk of cardiovascular events or CKD progression, the ADA Guidelines recommend the use of nonsteroidal mineralocorticoid receptor antagonists (MRAs) to delay the progression of CKD and reduce the risk of cardiovascular events.

[0004] Finerenone (C 21 H 22 N4O3, (4S)-4-(4-cyano-2-methoxyphenyl)-5-ethoxy-1,4-dihydro-2,8-dimethyl-1,6-naphthyridine-3-carboxamide) is a non-steroidal MRA and a target organ protective drug. In the treatment of patients with combined CKD and DKD, it can not only effectively reduce proteinuria, but also protect the cardiovascular and renal functions of patients with T2DM-related CKD.

[0005] The finerenone preparation process, as described below, generates a range of impurities that can be transferred to the finerenone API and related finished products during subsequent production, directly impacting the quality, safety, and efficacy of the API and finished products. However, there is currently no effective HPLC method for quality control of finerenone impurities generated during this preparation process.

[0006]

[0007] Summary of the Invention

[0008] Purpose of the invention: The present invention aims to provide an effective and reliable method for quality control of finerenone.

[0009] Technical solution: The quality control method of finerenone of the present invention comprises the following steps:

[0010] (1) Prepare sample solution;

[0011] (2) The sample solution prepared in step (1) is detected by high performance liquid chromatography, wherein the mobile phase A is selected from trifluoroacetic acid buffer, formic acid buffer, phosphate buffer, and phosphate buffer, and the mobile phase B is selected from acetonitrile and methanol; based on the volume fraction of mobile phase A, gradient elution is performed according to the following procedure:

[0012] 0-15 minutes: from 100% to 87%-77%, 15-25 minutes: from 87%-77% to 81%-71%, 25-55 minutes: from 81%-71% to 35%-25%, 55-60 minutes: maintained at 35%-25%;

[0013] (3) Analyze the content of each quality control component detected in step (2).

[0014] Preferably, the quality control components described in step (3) are as follows:

[0015]

[0016] Further preferably, in step (2), gradient elution is performed according to the following procedure based on the volume fraction of mobile phase A:

[0017] 0-15 minutes: from 100% to 84%-80%, 15-25 minutes: from 84%-80% to 78%-74%, 25-55 minutes: from 78%-74% to 32%-28%, 55-60 minutes: maintained at 32%-28%.

[0018] Preferably, in step (2), the gradient elution procedure further comprises the following procedure, based on the volume fraction of mobile phase A:

[0019] 60-60.1 minutes: from 35%-25% to 100%, 60.1-70 minutes: maintained at 100%.

[0020] Further preferably, in step (2), the gradient elution procedure further comprises the following procedure, based on the volume fraction of mobile phase A:

[0021] 60-60.1 minutes: changed from 32%-28% to 100%, 60.1-70 minutes: maintained at 100%.

[0022] More preferably, the procedure of the gradient elution is as follows:

[0023]

[0024]

[0025] Preferably, the mobile phase A in step (2) is selected from phosphate buffer, and the mobile phase B is selected from acetonitrile.

[0026] More preferably, the phosphate buffer is dipotassium hydrogen phosphate or potassium dihydrogen phosphate buffer.

[0027] More preferably, the mobile phase A is a 10 mM potassium dihydrogen phosphate buffer with a pH of 6.9 to 7.1 (pH adjusted with phosphoric acid).

[0028] Still more preferably, the mobile phase A is a 10 mM potassium dihydrogen phosphate buffer with a pH of 7.0 (pH adjusted with phosphoric acid).

[0029] Preferably, the high performance liquid chromatography method described in step (2) uses an alkylsilane bonded silica gel phase chromatography column, the detection wavelength is 210 to 310 nm, the mobile phase flow rate is 0.8 to 1.2 ml / min, and the chromatography column temperature is 20 to 40°C.

[0030] More preferably, the detection wavelength is 248-252 nm, the mobile phase flow rate is 0.9-1.1 ml / min, and the chromatographic column temperature is 23-27°C.

[0031] More preferably, the detection wavelength is 250 nm, the flow rate of the mobile phase is 1.0 ml / min, and the column temperature is 25°C.

[0032] More preferably, the injector temperature used in the HPLC method is 2-8°C.

[0033] More preferably, the injector temperature used in the HPLC method is 5°C.

[0034] Preferably, the chromatographic column used in the high performance liquid chromatography method is selected from C18 chromatographic column, C8 chromatographic column, phenyl column, cyano column, amino column, column length is 150-250 mm, column diameter is 2.0-4.6 mm, and column packing particle size is 3.0-5.0 μm.

[0035] More preferably, the chromatographic column is a C18 chromatographic column with a column length of 250 mm, a column diameter of 4.6 mm, and a column packing particle size of 5.0 μm.

[0036] More preferably, the chromatographic column is Thermo Hypersil GOLD TM The column or an equivalent chromatographic column has a column length of 250 mm, a column diameter of 4.6 mm, and a column packing particle size of 5.0 μm.

[0037] Preferably, the linear range of detection of each quality control component described in step (3) is as follows:

[0038]

[0039]

[0040] Further preferably, the detection linear equations of the quality control components are as follows:

[0041]

[0042] Preferably, the sample solution prepared in step (1) includes a reference solution, a test solution, and a separation solution.

[0043] More preferably, the preparation method of the reference solution is as follows:

[0044] The finerenone reference substance is added to an acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), dissolved and prepared into a solution with a concentration of 0.1 to 1.0 mg / ml, and more preferably, prepared into a reference substance solution with a concentration of 1.0 mg / ml.

[0045] Further preferably, the preparation method of the test solution is as follows:

[0046] Add the finerenone test sample to acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), dissolve it and prepare a test solution with a concentration of 0.1-1.0 mg / ml.

[0047] More preferably, the separation solution is prepared as follows:

[0048] Impurity A to impurity F reference substances were added to acetonitrile solution (the volume ratio of acetonitrile to water was 7:3), dissolved and prepared into a stock solution of each impurity with a concentration of 0.25 mg / ml.

[0049] Dissolve the finerenone reference substance in acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), add the stock solution of each impurity, and then add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3) to dissolve and prepare a separation solution containing 5 μg of each impurity and 1.0 mg of finerenone per 1 ml.

[0050] Preferably, the injection volume of the sample solution in step (2) is 5 μl.

[0051] More preferably, the injection volume of the test solution in the sample solution is 5 μl.

[0052] Preferably, the content of each quality control component in step (3) is analyzed using the area normalization method.

[0053] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0054] The present invention establishes a detection method for specific finerenone impurities that can accurately and effectively reflect the quality status of finerenone. The method is simple to operate, has excellent specificity (the separation between each impurity and the main component is not less than 1.5), sensitivity (the sensitivity of the limit of quantification is not greater than 0.03%), accuracy (linearity and range: the linear correlation coefficients of finerenone and each impurity are both greater than 0.999, and the percentage of the Y-axis intercept to the 100% concentration level response value is within 25%; recovery: the recovery rate of each impurity is between 90% and 110%), stability (the system suitability solution is stable within 61 hours), and durability, meeting quality control requirements. It is widely applicable and can be used for the entire finerenone preparation process, the quality control of raw materials and their preparations. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] Figure 1 This is the HPLC chromatogram of Test 1 in Example 1;

[0056] Figure 2 This is the HPLC chromatogram of Test 2 in Example 1;

[0057] Figure 3 This is the HPLC chromatogram of Test 3 in Example 1;

[0058] Figure 4 This is the HPLC chromatogram of Test 1 in Example 2;

[0059] Figure 5 This is the HPLC detection chromatogram of Test 2 in Example 2;

[0060] Figure 6 This is the HPLC chromatogram of Test 3 in Example 2;

[0061] Figure 7 This is the HPLC chromatogram of Test 1 in Example 3;

[0062] Figure 8 This is the HPLC chromatogram of Test 2 in Example 3;

[0063] Figure 9 The HPLC chromatogram of the blank solution in Example 4 is shown;

[0064] Figure 10 The HPLC chromatogram of the separation solution in Example 4 is shown;

[0065] Figure 11 The HPLC chromatogram of the quantitative limit solution in Example 5 is shown;

[0066] Figure 12 The HPLC detection chromatogram of the detection limit solution in Example 5 is shown. DETAILED DESCRIPTION

[0067] The technical solution of the present invention will be further described below in conjunction with embodiments.

[0068] Example 1: Mobile phase screening test

[0069] (1) Test 1

[0070] Mobile phase A1: 0.1% phosphoric acid solution;

[0071] Mobile phase B1: acetonitrile.

[0072] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0073] Chromatographic conditions: Column: Thermo Hypersil GOLD TM , 250mm×4.6mm, 5μm; gradient elution was performed with 0.1% phosphoric acid solution-acetonitrile as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25℃; detection wavelength: 250nm; flow rate: 1.0ml / min.

[0074]

[0075] Test results: See Figure 1 As shown in Table 1-1, impurity A was not retained, impurity C was not separated from the main peak, impurity D was not separated from impurity E, and the separation degree was less than 1.5, which did not meet the quality control requirements.

[0076] Table 1-1 Test results of test 1 in embodiment 1

[0077] Impurity name Retention time (min) Separation Impurity A 3.570 / Impurity B 4.187 4.89 Impurity C 9.423 / Impurity D 14.830 20.23 Impurity E 15.027 0.73 Impurity F 27.327 32.16

[0078] (2) Test 2

[0079] Mobile phase A2: 10 mmol / L potassium dihydrogen phosphate buffer;

[0080] Mobile phase B2: acetonitrile.

[0081] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0082] Chromatographic conditions: Column: Thermo Hypersil GOLD TM, 250mm×4.6mm, 5μm; gradient elution was performed with 10mmol / L potassium dihydrogen phosphate buffer-acetonitrile as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25℃; detection wavelength: 250nm; flow rate: 1.0ml / min.

[0083]

[0084] Test results: See Figure 2 As shown in Table 1-2, the chromatographic peak of impurity F has an obvious front extension, which does not meet the quality control requirements.

[0085] Table 1-2 Test results of test 2 in Example 1

[0086] Impurity name Retention time (min) Separation Symmetry Factor Impurity A 14.977 / 0.97 Non-specific impurities 20.473 24.41 1.01 Finerenone 25.080 15.89 1.00 Impurity B 30.783 3.21 0.95 Impurity C 28.640 9.98 0.94 Impurity D 29.647 2.73 0.96 Impurity E 34.470 11.62 0.94 Impurity F 43.990 5.74 0.57

[0087] (3) Test 3

[0088] Mobile phase A3: 10 mM potassium dihydrogen phosphate buffer, pH = 7.0 (pH adjusted with phosphoric acid);

[0089] Mobile phase B3: acetonitrile.

[0090] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0091] Chromatographic conditions: Column: Thermo Hypersil GOLD TM , 250mm×4.6mm, 5μm; gradient elution was performed with potassium dihydrogen phosphate buffer-acetonitrile as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25℃; detection wavelength: 250nm; flow rate: 1.0ml / min.

[0092]

[0093] Test results: See Figure 3 As shown in Table 1-3, the separation degree between each impurity is not less than 1.5, which meets the quality control requirements.

[0094] Table 1-3 Test results of test 3 in Example 1

[0095] Impurity name Retention time (min) Separation Symmetry Factor Impurity A 15.000 / 1.05 Impurity B 19.523 23.21 1.04 Non-specific impurities 24.703 21.88 1.05 Finerenone 25.503 2.95 1.04 Impurity C 29.030 11.77 1.02 Impurity D 29.977 3.05 1.02 Impurity E 34.720 17.41 1.02 Impurity F 45.137 41.05 0.93

[0096] Example 2: Chromatographic column screening test

[0097] (1) Test 1

[0098] Column 1: Kromasil 100-5C8, 4.6 mm × 250 mm, 5 μm.

[0099] Gradient elution was performed using 10 mM potassium dihydrogen phosphate buffer-acetonitrile at pH 7.0 (pH adjusted with phosphoric acid) as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25°C; detection wavelength: 250 nm; flow rate: 1.0 ml / min.

[0100]

[0101] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0102] Test results: See Figure 4 As shown in Table 2-1, impurity C and impurity D were not separated, which did not meet the quality control requirements.

[0103] Table 2-1 Test results of test 1 in Example 2

[0104] Impurity name Retention time (min) Separation Impurity A 16.993 / Impurity B 22.643 20.30 Non-specific impurities 27.830 15.41 Finerenone 29.763 5.22 Impurity C+Impurity D 33.597 9.72 Impurity D 33.597 Not separated from impurity C Impurity E 38.010 12.61 Impurity F 48.187 30.73

[0105] (2) Test 2

[0106] Column 2: Agilent Eclipse XDB-Phenyl, 4.6 mm × 250 mm, 5 μm.

[0107] Gradient elution was performed using 10 mM potassium dihydrogen phosphate buffer-acetonitrile at pH 7.0 (pH adjusted with phosphoric acid) as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25°C; detection wavelength: 250 nm; flow rate: 1.0 ml / min.

[0108]

[0109] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0110] Test results: See Figure 5 As shown in Table 2-2, the chromatographic peaks of impurity A, impurity B, impurity C, and impurity F have poor symmetry and do not meet the quality control requirements.

[0111] Table 2-2 Test results of test 2 in embodiment 2

[0112] Impurity name Retention time (min) Symmetry Factor Impurity A 15.650 1.38 Impurity B 18.837 0.85 Non-specific impurities 25.113 1.26 Finerenone 26.750 1.09 Impurity C 29.157 0.87 Impurity D 30.960 1.02 Impurity E 36.060 1.06 Impurity F 43.583 0.74

[0113] (3) Test 3

[0114] Column 3: Thermo Hypersil GOLD TM 4.6mm×250mm, 5μm.

[0115] The remaining chromatographic conditions were the same as those for mobile phase A3 / B3 in Example 1.

[0116] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0117] Test results: See Figure 6 As shown in Table 2-3, the separation between finerenone and adjacent impurities and between each impurity is not less than 1.5, and the impurity peak shapes are significantly improved, meeting the quality control requirements.

[0118] Table 2-3 Test results of test 3 in embodiment 2

[0119] Impurity name Retention time (min) Separation Symmetry Factor Impurity A 15.000 / 1.05 Impurity B 19.523 23.21 1.04 Non-specific impurities 24.703 21.88 1.05 Finerenone 25.503 2.95 1.04 Impurity C 29.030 11.77 1.02 Impurity D 29.977 3.05 1.02 Impurity E 34.720 17.41 1.02 Impurity F 45.137 41.05 0.93

[0120] Example 3: Gradient elution screening

[0121] (1) Test 1

[0122] Column 3: Thermo Hypersil GOLD TM 4.6mm×250mm, 5μm.

[0123] Gradient elution was performed using 10 mM potassium dihydrogen phosphate buffer-acetonitrile at pH 7.0 (pH adjusted with phosphoric acid) as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25°C; detection wavelength: 250 nm; flow rate: 1.0 ml / min.

[0124]

[0125] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0126] Test results: See Figure 7 As shown in Table 3-1, finerenone was not separated from adjacent impurities (non-specific impurities, RT: 23.333 min), which did not meet the quality control requirements.

[0127] Table 3-1 Test results of test 1 in Example 3

[0128]

[0129]

[0130] (2) Test 2

[0131] Column 3: Thermo Hypersil GOLD TM 4.6mm×250mm, 5μm.

[0132] The remaining chromatographic conditions were the same as those for mobile phase A3 / B3 in Example 1.

[0133] Test solution: Weigh about 20 mg of finerenone into a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), sonicate to dissolve and dilute to the mark, and shake well.

[0134] Test results: See Figure 8 As shown in Table 3-2, the separation between finerenone and adjacent impurities and between each impurity is not less than 1.5, and the impurity peak shapes are symmetrical, meeting the quality control requirements.

[0135] Table 3-2 Test results of test 2 in Example 3

[0136] Impurity name Retention time (min) Separation Symmetry Factor Impurity A 15.000 / 1.05 Impurity B 19.523 23.21 1.04 Non-specific impurities 24.703 21.88 1.05 Finerenone 25.503 2.95 1.04 Impurity C 29.030 11.77 1.02 Impurity D 29.977 3.05 1.02 Impurity E 34.720 17.41 1.02 Impurity F 45.137 41.05 0.93

[0137] Example 4: Specificity test

[0138] Specificity testing requires verification of the blank solution's absence of interference at the retention time of the main peak in the test and reference solutions, as well as the resolution between the impurities and the main component. The following are the preparation methods for each impurity and main component:

[0139] Impurity A localization solution: Accurately weigh 12.5 mg of the impurity A reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use it as the impurity A stock solution; accurately pipette the impurity A stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity A localization solution.

[0140] Impurity B localization solution: Accurately weigh 12.5 mg of the impurity B reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use as the impurity B stock solution; accurately pipette the impurity B stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity B localization solution.

[0141] Finerenone localization solution: Accurately weigh 12.5 mg of finerenone reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile:water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use as the localization solution.

[0142] Impurity C localization solution: Accurately weigh 12.5 mg of the impurity C reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use as the impurity C stock solution; accurately pipette the impurity C stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity C localization solution.

[0143] Impurity D localization solution: Accurately weigh 12.5 mg of the impurity D reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use as the impurity D stock solution; accurately pipette the impurity D stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity D localization solution.

[0144] Impurity E localization solution: Accurately weigh 12.5 mg of the impurity E reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use it as the impurity E stock solution; accurately pipette the impurity E stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity E localization solution.

[0145] Impurity F localization solution: Accurately weigh 12.5 mg of the impurity F reference substance, place it in a 50 ml volumetric flask, add appropriate amount of acetonitrile: water (7:3 V / V), sonicate to dissolve and dilute to the scale, shake well, and use as the impurity F stock solution; accurately pipette the impurity F stock solution and quantitatively dilute it to 5 μg / ml, which is used as the impurity F localization solution.

[0146] Blank solution: acetonitrile:water (7:3 V / V).

[0147] Resolving power solution: Accurately weigh 20 mg of finerenone reference substance, place it in a 20 ml volumetric flask, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), and dissolve it by ultrasonication. Accurately pipette the stock solution of the above impurities, add acetonitrile solution (the volume ratio of acetonitrile to water is 7:3), dissolve and dilute to make a solution containing 5 μg of each related substance and 1.0 mg of finerenone per 11 ml, which is used as the resolving power solution.

[0148] Test solution: Weigh about 20 mg of phenelline into a 20 ml volumetric flask, add acetonitrile:water (7:3 V / V), sonicate to dissolve and dilute to the mark, and shake well.

[0149] Reference solution: Prepare in the same manner as the test solution.

[0150] Reference solution of each impurity: Accurately pipette the stock solution of each impurity, place it in a volumetric flask, add diluent (acetonitrile solution, the volume ratio of acetonitrile to water is 7:3), dilute to the scale, shake well, and prepare a 5μg / ml reference solution of each impurity.

[0151] Chromatographic conditions: Column: Thermo Hypersil GOLD TM, 250mm×4.6mm, 5μm; gradient elution was performed with 10mM potassium dihydrogen phosphate buffer-acetonitrile at pH=7.0 (pH adjusted with phosphoric acid) as the mobile phase; the elution gradient was as follows based on the volume ratio of mobile phase A; column temperature: 25°C; detection wavelength: 250nm; flow rate: 1.0ml / min.

[0152]

[0153] Accurately measure 5 μl of each of the above solutions, inject them into the liquid chromatograph, and record the chromatogram. The results are shown in Table 4 and Figures 9 and 10 .

[0154] Table 4 Results of specificity investigation

[0155]

[0156] From Table 4 and Figures 9 and 10 It can be seen that the blank solution has no interference at the retention time of the main peak in the test and reference solutions respectively; the separation degree between the impurity and the main component is ≥1.5, indicating that the specificity of the detection method of the present invention meets the quality control requirements.

[0157] Example 5: Sensitivity test

[0158] The impurity A stock solution, impurity B stock solution, impurity C stock solution, impurity D stock solution, impurity E stock solution, impurity F stock solution, and reference solution prepared in Example 3 were diluted with solvent to prepare solutions of appropriate concentrations, and diluted stepwise to appropriate multiples. The solution with a signal-to-noise ratio ≥10:1 was used as the quantification limit solution; the solution with a signal-to-noise ratio ≥3:1 was used as the detection limit solution.

[0159] 5 μl of the quantitative limit solution and the detection limit solution were accurately measured and injected into the liquid chromatograph. The chromatographic conditions were the same as those in Example 3. One injection was made for each of the quantitative limit solution and the detection limit solution. The results are shown in Tables 5-1 to 5-2 and Figures 11 and 12 .

[0160] Table 5-1 Results of Quantitation Limit Investigation

[0161] name Concentration (μg / ml) S / N Sensitivity (%) Impurity A 0.2945 39.7 0.03 Impurity B 0.2972 38.1 0.03 Finerenone 0.2989 28.9 0.03 Impurity C 0.2974 45.8 0.03 Impurity D 0.2988 22.4 0.03 Impurity E 0.2991 30.3 0.03 Impurity F 0.2953 25.5 0.03

[0162] Table 5-2 Detection limit investigation results

[0163] name Concentration (μg / ml) S / N Sensitivity (%) Impurity A 0.0883 18.6 0.01 Impurity B 0.0892 18.7 0.01 Finerenone 0.0897 13.0 0.01 Impurity C 0.0892 22.4 0.01 Impurity D 0.0896 11.6 0.01 Impurity E 0.0897 15.6 0.01 Impurity F 0.0886 11.7 0.01

[0164] From Table 5-1 to Table 5-2 and Figures 11 and 12 It can be seen that the sensitivity of the quantitative limit is no more than 0.03%, and the sensitivity of the detection limit is no more than 0.01%, indicating that the sensitivity of the detection method of the present invention meets the quality control requirements.

[0165] Example 6: Linearity and range test

[0166] Reference solution: Prepare the reference solution in the same manner as in Example 3.

[0167] Impurity A to F stock solutions: prepared in the same manner as the impurity A to F stock solutions in Example 3.

[0168] Linear stock solution: Take appropriate amount of finerenone reference solution and impurity A~F stock solution, add acetonitrile:water (7:3V / V) to dilute to the scale, and shake well (the concentration of finerenone and impurity A~F is 10μg / ml).

[0169] Accurately measure the corresponding volume of linear stock solution into a volumetric flask of the corresponding volume, add acetonitrile: water (7:3 V / V) to dilute to the scale, shake well, and use as the linear solution (the concentration of each linear solution is shown in Table 5-1 to Table 5-7). Accurately measure 10 μl of each linear solution and inject it into the liquid chromatograph. The chromatographic conditions are the same as those in Example 4. The results are shown in Table 6-1 to Table 6-7.

[0170] Table 6-1 Results of linearity and range investigation of finerenone

[0171]

[0172] Table 6-2 Linearity and range investigation results of impurity A

[0173]

[0174]

[0175] Table 6-3 Linearity and range investigation results of impurity B

[0176]

[0177] Table 6-4 Linearity and range investigation results of impurity C

[0178]

[0179] Table 6-5 Linearity and range investigation results of impurity D

[0180]

[0181] Table 6-6 Linearity and range investigation results of impurity E

[0182]

[0183]

[0184] Table 6-7 Linearity and range investigation results of impurity F

[0185]

[0186] As can be seen from Tables 6-1 to 6-7, the linear correlation coefficients of finerenone and each impurity are all >0.999, and the percentage of the Y-axis intercept and the 100% concentration level response value is within 25%, indicating that the linearity and range of the detection method of the present invention meet the quality control requirements.

[0187] Example 7: Methodology Validation Test

[0188] (1) Methodological validation test method

[0189]

[0190] (2) Methodology validation test results

[0191]

[0192]

Claims

1. A quality control method for finerenone, characterized in that: The following steps are involved: (1) Prepare sample solution; (2) The sample solution prepared in step (1) is detected by high performance liquid chromatography, wherein the mobile phase A is selected from trifluoroacetic acid buffer, formic acid buffer, phosphate buffer, and phosphate buffer, and the mobile phase B is selected from acetonitrile and methanol; based on the volume fraction of mobile phase A, gradient elution is performed according to the following procedure: 0-15 minutes: from 100% to 87%-77%, 15-25 minutes: from 87%-77% to 81%-71%, 25-55 minutes: from 81%-71% to 35%-25%, 55-60 minutes: maintained at 35%-25%; (3) Analyze the content of each quality control component detected in step (2).

2. The quality control method according to claim 1, characterized in that The quality control components described in step (3) are as follows:

3. The quality control method according to claim 1 or 2, characterized in that: In step (2), based on the volume fraction of mobile phase A, the gradient elution procedure further comprises the following procedure: 60-60.1 minutes: from 35%-25% to 100%, 60.1-70 minutes: maintained at 100%.

4. The quality control method according to claim 3, characterized in that: The procedure of gradient elution in step (2) is as follows:

5. The quality control method according to claim 1 or 2, characterized in that: The mobile phase A described in step (2) is selected from phosphate buffer, and the mobile phase B is selected from acetonitrile.

6. The quality control method according to claim 1 or 2, characterized in that: The high performance liquid chromatography method described in step (2) adopts an alkylsilane bonded silica gel phase chromatography column, the detection wavelength is 210-310 nm, the mobile phase flow rate is 0.8-1.2 ml / min, and the chromatography column temperature is 20-40°C.

7. The quality control method according to claim 6, characterized in that: The chromatographic column is selected from C18 chromatographic column, C8 chromatographic column, phenyl column, cyano column and amino column, with a column length of 150-250 mm, a column diameter of 2.0-4.6 mm and a column filler particle size of 3.0-5.0 μm.

8. The quality control method according to claim 7, characterized in that: The chromatographic column is a C18 chromatographic column with a column length of 250 mm, a column diameter of 4.6 mm, and a column filler particle size of 5.0 μm.

9. The quality control method according to claim 6, characterized in that: The detection wavelength is 248-252 nm, the flow rate of the mobile phase is 0.9-1.1 ml / min, and the column temperature of the chromatographic column is 23-27° C.

10. The quality control method according to claim 1 or 2, characterized in that: The linear range of each quality control component described in step (3) is as follows:

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