Detection method for determining impurity pyridine-3-sulfonic acid in Vonoprazan fumarate tablets
The method of separating and detecting pyridine-3-sulfonic acid, an impurity in vonoprazan fumarate tablets, by high performance liquid chromatography (HPLC) solves the problem of inaccurate separation and quantification in existing technologies, and achieves detection results with high sensitivity and high repeatability.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- HAINAN ASIA PHARM CO LTD
- Filing Date
- 2026-03-19
- Publication Date
- 2026-05-05
AI Technical Summary
Existing technologies are insufficient for effectively separating and sensitively detecting the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets, and conventional methods are easily affected by matrix interference, leading to inaccurate quantification and potential safety hazards.
High-performance liquid chromatography (HPLC) was employed, using octadecylsilane-bonded silica gel as the stationary phase. Mobile phase A consisted of ammonium acetate solution-sodium octanesulfonate-methanol, and mobile phase B consisted of methanol-water solution. A gradient elution program was used, with a detection wavelength of 260-265 nm, a column temperature of 32-35 °C, and a flow rate of 0.8-1.2 mL/min. This method enabled the efficient separation and quantification of the impurity pyridine-3-sulfonic acid.
It achieves high specificity, high sensitivity, and good repeatability for the impurity pyridine-3-sulfonic acid, effectively eliminates matrix interference, achieves accurate quantification, and improves the level of product quality control.
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Figure CN121978251A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of pharmaceutical analysis and detection technology, and in particular to a method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets. Background Technology
[0002] Vonoprazan fumarate (TAK-438) is a novel oral potassium-competitive acid blocker (P-CAB), chemically named 1-[5-(2-fluorophenyl)-1-(pyridin-3-sulfonyl)-1H-pyrrolo-3-yl]-N-methylmethylamine monofumarate. It effectively inhibits the formation of upper gastrointestinal mucosal damage and is used to treat reflux esophagitis and, in combination with appropriate antibiotics, to eradicate Helicobacter pylori. The key starting material for the synthesis of vonoprazan fumarate, pyridine-3-sulfonyl chloride, is a strong oxidizing agent with high chemical reactivity. It readily hydrolyzes to pyridine-3-sulfonic acid. Furthermore, vonoprazan fumarate itself can degrade to pyridine-3-sulfonic acid. This impurity of pyridine-3-sulfonic acid can be introduced into the finished product, leading to a lack of therapeutic effect, affecting drug stability and efficacy, and even exhibiting genotoxicity, directly causing DNA damage, leading to DNA mutations, and potentially causing cancer, posing a safety risk. Because the impurity pyridine-3-sulfonic acid is highly polar and has no or very weak UV absorption in the weakly basic drug matrix of vonoprazan fumarate, conventional HPLC-UV methods often fail to achieve effective separation and sensitive quantitative detection, resulting in poor repeatability. Furthermore, the complex formulation of vonoprazan fumarate tablets means that excipients and main components can easily interfere with the elution of pyridine-3-sulfonic acid, leading to inaccurate quantification.
[0003] Therefore, there is an urgent need to develop a detection method that is highly specific, highly sensitive, has poor anti-interference ability, and is easy to operate. Summary of the Invention
[0004] Therefore, this invention proposes a method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets.
[0005] The technical solution of this invention is implemented as follows: A method for determining pyridine-3-sulfonic acid, an impurity in vonoprazan fumarate tablets, is disclosed. The method employs high-performance liquid chromatography (HPLC) with the following chromatographic conditions: the stationary phase is octadecylsilane-bonded silica gel; mobile phase A is ammonium acetate solution-sodium octanesulfonate-methanol; mobile phase B is methanol-water solution; and elution is performed using a gradient elution program.
[0006] Furthermore, in the mobile phase A, the volume ratio of ammonium acetate solution - sodium octane sulfonate - methanol is 92-94:1-3:5; in the mobile phase B, the volume concentration of the methanol aqueous solution is 50%-70%.
[0007] Furthermore, in the mobile phase A, the volume ratio of ammonium acetate solution - sodium octane sulfonate - methanol is 93:2:5; in the mobile phase B, the volume concentration of the methanol aqueous solution is 60%.
[0008] Furthermore, the concentration of the ammonium acetate solution is 25-30 mmol / L.
[0009] Furthermore, the pH of the mobile phase A is 6.0-6.2.
[0010] Furthermore, the gradient elution procedure is as follows:
[0011] Furthermore, the chromatographic column used in the high-performance liquid chromatography determination is an Incrtsil ODS-3 column with dimensions of 4.6 mm × 250 mm and a diameter of 5 μm.
[0012] Furthermore, the detection wavelength in the high-performance liquid chromatography determination process is 260-265 nm, the column temperature is 32-35 °C, and the flow rate is 0.8-1.2 mL / min.
[0013] Furthermore, the injection volume in the high-performance liquid chromatography determination process is 10 μL.
[0014] Compared with the prior art, the beneficial effects of the present invention are: 1. This invention establishes a method for detecting pyridine-3-sulfonic acid, an impurity in vonoprazan fumarate tablets, using high performance liquid chromatography (mobile phase A is ammonium acetate solution-sodium octanesulfonate-methanol, mobile phase B is methanol aqueous solution, gradient elution). The method is stable and feasible, and can effectively control the quality of this product. Furthermore, the results of methodological studies on linearity, precision, repeatability, recovery rate, and robustness show that the method is reasonably designed, highly operable, and precise and reliable.
[0015] 2. The detection method for pyridine-3-sulfonic acid, an impurity in vonoprazan fumarate tablets of the present invention has good specificity and high detection sensitivity. The impurity peak can be completely separated from the main peak, effectively eliminating matrix interference and achieving accurate quantification of the target impurity. It has the characteristics of high sensitivity, good repeatability and simple operation, which greatly improves the product quality control level. Attached Figure Description
[0016] Figure 1 The chromatogram of the diluent for the specificity test in Example 3 is shown.
[0017] Figure 2 The chromatogram of the test sample for the specificity test in Example 3 is shown.
[0018] Figure 3 The chromatogram of the vonoprazan fumarate control solution is shown in Example 3 for the specificity test.
[0019] Figure 4 This is the chromatogram of the reference solution for the specificity test in Example 3.
[0020] Figure 5 The chromatogram of the spiked test solution for the specificity test in Example 3 is shown. Detailed Implementation
[0021] To better understand the technical content of this invention, specific embodiments are provided below to further illustrate the invention.
[0022] Unless otherwise specified, the experimental methods used in the embodiments of this invention are all conventional methods.
[0023] Unless otherwise specified, all materials and reagents used in the embodiments of this invention are commercially available.
[0024] The impurity of this invention, pyridine-3-sulfonic acid, has the following CAS number: 636-73-7; molecular formula: C5H5NO3S; molecular weight: 159.16; and structural formula: .
[0025] The impurity pyridine-3-sulfonic acid reference standard of this invention is obtained from CATO of Guangzhou Jiatu Technology Co., Ltd., batch number 0712-RC-0047.
[0026] The vonoprazan fumarate tablets of this invention are from Hainan Asia Pharmaceutical Co., Ltd., batch number 03240801.
[0027] The vonoprazan fumarate raw material of this invention is sourced from Shandong Langnuo Pharmaceutical Co., Ltd., batch number 231200330603.
[0028] Example 1 Diluent (blank solution): 30 mmol / L ammonium acetate solution - methanol (volume ratio 95:5).
[0029] Test solution: Take vonoprazan fumarate tablets, place them in a volumetric flask, add an appropriate amount of diluent, shake for about 30 minutes to allow the tablets to completely disintegrate, then dilute to the mark with diluent, shake well, centrifuge at 10,000 rpm for 10 minutes, and take the supernatant as the test solution.
[0030] Stock solution of impurity pyridine-3-sulfonic acid reference standard: Accurately weigh the impurity pyridine-3-sulfonic acid reference standard, place it in a volumetric flask, dilute to the mark with diluent, shake well, and prepare a solution containing approximately 100 μg per ml.
[0031] Impurity pyridine-3-sulfonic acid reference solution: Accurately measure the stock solution of impurity pyridine-3-sulfonic acid reference solution, place it in a volumetric flask, dilute to the mark with diluent, shake well, and prepare a solution containing approximately 2 μg of impurity pyridine-3-sulfonic acid per 1 ml.
[0032] System suitability solution: Same as "Impurity pyridine-3-sulfonic acid reference solution".
[0033] Spiked test solution: Take vonoprazan fumarate tablets, place them in a volumetric flask, add an appropriate amount of diluent, shake for about 30 minutes to allow the tablets to completely disintegrate, accurately measure the impurity pyridine-3-sulfonic acid reference standard stock solution, place it in a volumetric flask, dilute to the mark with diluent, shake well, centrifuge at 10000 rpm for 10 minutes, and take the supernatant, which is the spiked test solution.
[0034] Vonoprazan fumarate control solution: Accurately weigh the vonoprazan fumarate raw material, place it in a volumetric flask, add diluent to dilute to the mark, and shake well.
[0035] Chromatographic conditions:
[0036] The gradient elution procedure is as follows:
[0037] Example 2 System suitability test method: Take blank solution and system suitability solution for analysis, and record the chromatograms. Investigate the theoretical plate number and tailing factor of the pyridine-3-sulfonic acid peak in the system suitability solution.
[0038] The experimental results are shown in Table 1.
[0039] Table 1
[0040] As shown in Table 1, the theoretical plate number of the reference solution was ≥16041 for five consecutive determinations, and the tailing factor was between 1.0 and 1.1, which met the requirements.
[0041] Example 3 Specificity test method: Take diluent, vonoprazan fumarate control solution, test solution and spiked test sample, inject and determine, record the chromatogram, and investigate the retention time of chromatographic peaks in each solution.
[0042] The experimental results are shown in Table 2.
[0043] Table 2
[0044] As shown in Table 2, no pyridine-3-sulfonic acid peak was detected in the diluent, test solution, and vonoprazan fumarate reference solution, indicating that it did not interfere with the detection. The retention time of pyridine-3-sulfonic acid in the spiked test solution was basically consistent with that in the system suitability solution.
[0045] Example 4 Linearity test method: Based on the 100% concentration limit level of impurity pyridine-3-sulfonic acid, the stock solution of impurity pyridine-3-sulfonic acid reference standard in Example 1 was diluted to prepare a series of linear concentrations of LOQ, 50%, 80%, 100%, 120%, and 150%, respectively, as linearity determination solutions. Injection and detection were performed, and chromatograms were recorded. Linear regression was performed with the impurity pyridine-3-sulfonic acid concentration as the x-axis and peak area as the y-axis to obtain the regression equation and correlation coefficient. Specific methods for preparing linear solutions are shown in Table 3. Table 3
[0046] The experimental results are shown in Table 4.
[0047] Table 4
[0048] As can be seen from Table 4, the correlation coefficient of the regression line is R=0.9995, the Y-intercept is 1.3% of the 100% response value, and the data points in the regression residual and residual distribution plot are randomly distributed, all of which meet the requirements.
[0049] Example 5 Test methods for limits of quantitation and detection: The limits of quantitation (LOQ) and detection (LOD) of impurity pyridine-3-sulfonic acid were calculated using the slope (S) and residual standard deviation (σ) of the linear equation for impurity pyridine-3-sulfonic acid under Example 4.
[0050] Take the stock solution of impurity pyridine-3-sulfonic acid, dilute it stepwise, inject it for analysis, and confirm that the signal-to-noise ratio (S / N) of the pyridine-3-sulfonic acid peak is ≥10, which is the LOQ concentration solution of this component. Prepare 6 limit of quantitation solutions of this concentration using the same method or inject 6 samples consecutively, analyze them, and calculate the RSD value of the peak area of the 6 solutions.
[0051] Dilute the solution according to the above method for preparing the limit of quantitation solution to the LOD concentration, and confirm that its S / N>3, which is the limit of detection concentration solution.
[0052] The experimental results are shown in Table 5.
[0053] Table 5
[0054] As shown in Table 5, the signal-to-noise ratio of the pyridine-3-sulfonic acid peak in the six limit-of-quantitation solutions was >28.40, and the RSD of the peak area was 3%; the signal-to-noise ratio of the pyridine-3-sulfonic acid peak in the limit-of-detection solutions was 10.31, both of which met the requirements.
[0055] Example 6 Accuracy test method: Using the limit of impurity pyridine-3-sulfonic acid (0.20%) as a 100% concentration baseline, accuracy solutions containing impurity pyridine-3-sulfonic acid equivalent to LOQ, 25%, 50%, and 120% were prepared. Each concentration was prepared in triplicate, and the chromatograms were recorded. The recovery rate and RSD values were calculated. Specific accuracy solution preparations are shown in Table 6. Table 6
[0056] Preparation of recovery solution: Take this product, place it in a volumetric flask, add diluent and shake for 30 minutes to disintegrate the tablets, add impurity pyridine-3-sulfonic acid reference standard stock solution, and then dilute to the mark with diluent. Shake well to obtain the solution.
[0057] After centrifuging the above solution at 10,000 rpm for 10 min, the supernatant was collected and injected into the liquid chromatograph along with the test solution and the impurity pyridine-3-sulfonic acid reference solution.
[0058] The experimental results are shown in Table 7.
[0059] Table 7
[0060] As shown in Table 7, the recovery rate at each concentration level ranged from 95.9% to 108.0%, and the RSD (n=3) ranged from 0.6% to 4.9%. The average recovery rate of the accuracy solution at all concentration levels was 98.8%, and the RSD (n=12) for all concentration levels was 3%, all of which met the requirements.
[0061] Example 7 Precision 1. Repetitiveness Test method: Repeatability - Test solution: Prepared according to the spiked test solution in Example 1, with 6 parallel preparations.
[0062] Take 6 test solutions and pyridine-3-sulfonic acid reference solution, inject them for analysis, and record the chromatograms. Calculate the content and RSD value of pyridine-3-sulfonic acid in the 6 test solutions.
[0063] The results are shown in Table 8.
[0064] Table 8
[0065] As can be seen from Table 8, the RSD of the impurity pyridine-3-sulfonic acid content in the 6 spiked test sample solutions was 0.1%.
[0066] 2. Intermediate precision Test method: Different analysts, different dates, and different instruments prepared relevant solutions under the "Repeatability" section, injected and detected, and recorded chromatograms. Calculate the RSD values of the impurity pyridine-3-sulfonic acid content in 6 test sample solutions, and combine them with the results of 6 tests under the "Repeatability" section to calculate the RSD values of the impurity pyridine-3-sulfonic acid content in 12 test samples.
[0067] The results are shown in Table 9.
[0068] Table 9
[0069] As can be seen from Table 9, the RSD of the impurity pyridine-3-sulfonic acid content in the 6 spiked test sample solutions was 0.1%; the combined RSD of the 12 samples from the two individuals was 0.1%, which meets the requirements.
[0070] Example 8 Solution stability test method: Take the reference solution and the spiked test solution under the repeatability section of "Example 7", place them under the proposed conditions, and inject them at time points of 1 day, 2 days and 3 days to investigate the change in the peak area of the impurity pyridine-3-sulfonic acid.
[0071] The proposed conditions for studying solution stability are: room temperature, no illuminance control, colorless container, and the test samples are pyridine-3-sulfonic acid reference solution and spiked test solution. The study period is 1, 2, and 3 days.
[0072] The results are shown in Table 10.
[0073] Table 10
[0074] As shown in Table 10, the ratio of the main peak area to the 0h peak area in the reference solution after being placed at room temperature for 45.7 hours was 99.4%-99.7%; the ratio of the main peak area to the 0h peak area in the spiked test sample after being placed at room temperature for 39 hours was 100.2%-100.9%, which meets the requirements.
[0075] Example 9 Durability test method: Take diluent and system suitability solution, inject them into the liquid chromatograph under different conditions for durability testing, record the chromatograms, and examine the system suitability results.
[0076] The specific durability test conditions are shown in Table 11.
[0077] Table 11
[0078] The results are shown in Table 12.
[0079] Table 12
[0080] As shown in Table 12, under various conditions of durability variation, the theoretical plate number of the pyridine-3-sulfonic acid peak in the reference solution was >6134, and the tailing factor was between 1.0 and 1.1, all of which met the requirements, indicating that the method has good durability.
[0081] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.
Claims
1. A method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets, characterized in that, The determination was performed using high performance liquid chromatography (HPLC). The chromatographic conditions were as follows: the stationary phase was octadecylsilane-bonded silica gel; mobile phase A was ammonium acetate solution-sodium octanesulfonate-methanol; mobile phase B was methanol-water solution; and a gradient elution program was used for elution.
2. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 1, characterized in that, In the mobile phase A, the volume ratio of ammonium acetate solution - sodium octane sulfonate - methanol is 92-94:1-3:5; in the mobile phase B, the volume concentration of the methanol aqueous solution is 50%-70%.
3. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 2, characterized in that, In the mobile phase A, the volume ratio of ammonium acetate solution - sodium octane sulfonate - methanol is 93:2:5; in the mobile phase B, the volume concentration of the methanol aqueous solution is 60%.
4. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 3, characterized in that, The concentration of the ammonium acetate solution is 25-30 mmol / L.
5. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 3, characterized in that, The pH of the mobile phase A is 6.0-6.
2.
6. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 1, characterized in that, The gradient elution procedure is as follows:
7. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 1, characterized in that, The high-performance liquid chromatography (HPLC) used an Incrtsil ODS-3 column with dimensions of 4.6 mm × 250 mm and a diameter of 5 μm.
8. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 1, characterized in that, The detection wavelength in the high-performance liquid chromatography (HPLC) determination process is 260-265 nm, the column temperature is 32-35 °C, and the flow rate is 0.8-1.2 mL / min.
9. The method for determining the impurity pyridine-3-sulfonic acid in vonoprazan fumarate tablets as described in claim 1, characterized in that, The injection volume during the high-performance liquid chromatography determination process is 10 μL.