Method for detecting genetic toxic impurity D of varenicline tartrate

Abstract

The invention provides the method for detecting the genetic toxic impurity D of varenicline tartrate, and the method has the advantages of strong instrument universality, strong specificity, no interference of blank solvent on detection, high accuracy, good durability and more convenient operation, can more effectively realize quality control on the links of the product production process, improves the quality of the final product, and is suitable for industrial production. The safety and the effectiveness of clinical medication are ensured.

Description

1. Technical field

[0001] The invention belongs to the technical field of drug analysis, and particularly relates to an HPLC method for detecting the genetic toxic impurity D of varenicline tartrate. 2. Background technology

[0002] Varenicline tartrate, a nicotinic acetylcholine receptor alpha 4 β 2 Subtype partial stimulant oral smoking cessation drug, mainly used for adult smoking cessation. Varenicline tartrate tablets were developed by Pfizer, USA. On May 10, 2006, Pfizer's Varenicline tartrate tablets were approved by the US FDA under the trade name Chantix. On September 26 of the same year, they were approved by the European Union under the trade name Champix. At the end of July 2008, Varenicline tartrate tablets were approved for import in my country.

[0003] Varenicline tartrate, CAS number: 341031-54-7, chemical name: 7,8,9,10-tetrahydro-6,10-methylene-6H-pyrazino[2,3-h][3]benzazepine -(2R,3R)-2,3-dihydroxysuccinate (1:1), the structural formula is as follows:

[0004]

[0005] Varenicline tartrate impurity D, with the structure shown below, is an intermediate in the synthesis of varenicline tartrate raw materials. It contains aromatic amine warning structures and is a genotoxic impurity.

[0006]

[0007] No toxicological information related to impurity D was found through searching. Its limit was calculated according to the ICH M7 guidelines. The maximum daily dose of varenicline tartrate is 3.4 mg (calculated as varenicline tartrate). According to the toxicological concern threshold TTC of 1.5 μg / day, the impurity D limit = 1.5 μg / 3.4 mg × 100% = 0.04%.

[0008] At present, the pharmacopoeias of various countries do not include the standard of tartrate varenicline, and there is no relevant statutory standard for reference. Reference (Song Fan, Song Xiaoni, etc. LC-MS determination of 6 aniline-like genotoxic impurities in tartrate varenicline. Drug Analysis Impurities, 2018.) discloses a method for detecting genotoxic impurities of tartrate varenicline. The analysis method is a high-performance liquid chromatography-mass spectrometry, which requires a mass spectrometer. The instrument is expensive and has poor universality. Patent CN114689778 A discloses an analysis method for determining tartrate varenicline-related substances by high-performance liquid chromatography. In this patent, impurity D is only controlled as a common impurity, and the detection sensitivity of the method cannot be guaranteed when it is controlled as a genotoxic impurity. Therefore, it is necessary to develop a high-performance liquid chromatography method that can detect genotoxic impurity D in tartrate varenicline. 3. Summary of the invention

[0009] The invention develops a high performance liquid chromatography method, which can detect the genotoxic impurity D in varenicline tartrate, has strong instrument universality, strong method specificity, blank solvent does not interfere with detection, has high accuracy, good durability, and is more convenient to operate. It can more effectively achieve quality control of product production process links, improve the quality of final products, and ensure the safety and effectiveness of clinical drug use.

[0010] The present invention provides a method for detecting the genetic toxic impurity D of varenicline tartrate, comprising the following steps: respectively sampling a sensitivity solution, a reference solution and a test solution, detecting according to high performance liquid chromatography conditions, and recording a chromatogram. The chromatographic conditions are: chromatographic column: a chromatographic column with octylsilane bonded silica gel as a filler; mobile phase: mobile phase A is a mixed solution of a buffer solution and an organic solvent, mobile phase B is a mixed solution of a buffer solution and an organic solvent, and gradient elution is performed according to the following table:

[0011]

[0012] The chromatographic conditions also include:

[0013] Detector: UV detector;

[0014] Flow rate: 1.0ml / min;

[0015] Column temperature: 30-40°C, preferably 35°C;

[0016] Detection wavelength: 238nm;

[0017] Injection volume: 30 μl;

[0018] Run time: at least 1 minute after the main peak of the sensitivity and reference solutions is collected; 45 minutes for the test solution.

[0019] The chromatographic column is selected from Agilent ZORBAX RX-C8, 250 mm × 4.6 mm, 5 μm, Agilent ZORBAXSB-C 8 , 4.6mm×250mm, 5μm, Agela Venusil XBP C 8 , 4.6mm×250mm, 5μm, preferably Agilent ZORBAX SB-C8, 4.6mm×250mm, 5μm.

[0020] The mobile phase A, the buffer solution can be selected from one or more of sodium perchlorate solution, potassium dihydrogen phosphate solution, sodium dihydrogen phosphate solution, preferably sodium perchlorate solution. Wherein, the sodium perchlorate solution is adjusted to pH 2.0-3.0, preferably 2.5, with an acidic reagent, and its concentration is 30-70 mmol / L, preferably 50 mmol / L. The acidic reagent is selected from one or more of glacial acetic acid, phosphoric acid, and formic acid, preferably phosphoric acid. The organic solvent is selected from one or more of acetonitrile, methanol, and tetrahydrofuran, preferably acetonitrile. The ratio of the sodium perchlorate solution to acetonitrile is 78-82:22-18, preferably 80:20.

[0021] The mobile phase B, the buffer solution can be selected from one or more of sodium perchlorate solution, potassium dihydrogen phosphate solution, sodium dihydrogen phosphate solution, preferably sodium perchlorate solution. Wherein, the sodium perchlorate solution is adjusted to pH 2.0-3.0, preferably 2.5, with an acidic reagent, and its concentration is 30-70 mmol / L, preferably 50 mmol / L. The acidic reagent is selected from one or more of glacial acetic acid, phosphoric acid, and formic acid, preferably phosphoric acid. The organic solvent is selected from one or more of acetonitrile, methanol, and tetrahydrofuran, preferably acetonitrile. The ratio of the sodium perchlorate solution to acetonitrile is 50:50.

[0022] The preparation process of the reference solution is as follows: take an appropriate amount of impurity D reference substance, add solvent to dissolve, dilute and shake well.

[0023] The preparation process of the sensitivity solution is as follows: accurately measure 5 ml of impurity D reference solution, dilute to the scale with solvent, and shake well.

[0024] The preparation process of the test solution is as follows: take about 50 mg of varenicline tartrate, accurately weigh it, dilute it to the scale with a solvent, and shake it well.

[0025] The solvent is mobile phase A.

[0026] The present invention provides a method for detecting the genetic toxic impurity D of varenicline tartrate, comprising the following steps: respectively taking a sensitive solution, a reference solution and a test solution for sampling, detecting according to the conditions of high performance liquid chromatography, and recording a chromatogram. The chromatographic conditions are: chromatographic column: Agilent ZORBAX SB-C8, 4.6mm×250mm, 5μm; mobile phase: mobile phase A: 50mmol / L sodium perchlorate solution (adjusting the pH value to 2.5 with phosphoric acid): acetonitrile = 80:20; mobile phase B: 50mmol / L sodium perchlorate solution (adjusting the pH value to 2.5 with phosphoric acid): acetonitrile = 50:50, and gradient elution is performed according to the following table:

[0027]

[0028] The chromatographic conditions also include:

[0029] Detector: UV detector;

[0030] Flow rate: 1.0ml / min;

[0031] Column temperature: 35 °C;

[0032] Detection wavelength: 238nm;

[0033] Injection volume: 30 μl;

[0034] Run time: at least 1 minute after the main peak of the sensitivity and reference solutions is collected; 45 minutes for the test solution.

[0035] The preparation process of the reference solution is as follows: take an appropriate amount of impurity D reference substance, accurately weigh it, dissolve it with mobile phase A and quantitatively dilute it to make a solution containing about 0.24 μg per 1 ml.

[0036] The preparation process of the sensitivity solution is as follows: accurately measure 5 ml of impurity D reference solution, place it in a 10 ml volumetric flask, dilute to the scale with mobile phase A, and shake well.

[0037] The preparation process of the test solution is as follows: take about 50 mg of varenicline tartrate (equivalent to 30 mg of varenicline), accurately weigh, place in a 50 ml volumetric flask, add mobile phase A to dissolve and dilute to the scale, and shake well.

[0038] The beneficial technical effects of the present invention are as follows: the present invention develops a reverse phase HPLC method, which can detect the genotoxic impurity D in varenicline tartrate. The detection method used has strong instrument universality, strong method specificity, blank solvent does not interfere with the detection, has high accuracy, good durability, and is more convenient to operate. It can more effectively achieve quality control of the product production process, improve the quality of the final product, and ensure the safety and effectiveness of clinical drug use. 4. Description of the drawings

[0039] Figure 1 The overlay graph was selected for the Example 1 Test Condition 1 method.

[0040] Figure 2 The overlay graph was selected for the Test Condition 2 method of Example 1.

[0041] Figure 3 This is the blank solvent spectrum of test condition 2 in Example 1.

[0042] Figure 4 This is an overlapping diagram of the accuracy test of Example 2.

[0043] Figure 5 This is an overlay diagram of the reference solution in the solution stability test of Example 3.

[0044] Figure 6 This is an overlapping diagram of the test sample solution in the solution stability test of Example 3.

[0045] Figure 7 This is the linear test diagram of Example 4.

[0046] Figure 8 This is an overlap diagram of the quantitative limit solution of the range test in Example 6.

[0047] Fig. 9 This is the overlap graph of the 150% solution for the range test limit of Example 6.

[0048] Fig.10 This is the precision overlay diagram of the precision test system in Example 7.

[0049] Fig.11 This is an overlapping diagram of reference solutions of different chromatographic columns in the durability test of Example 8.

[0050] Fig.12 This is an overlapping diagram of the test sample solutions of different chromatographic columns in the durability test of Example 8.

[0051] Fig.13 This is an overlay diagram of reference solutions with different mobile phase pH values ​​in the durability test of Example 8.

[0052] Fig.14 This is an overlapping diagram of the test solution at different mobile phase pH values ​​in the durability test of Example 8.

[0053] Fig.15 This is an overlapping diagram of the reference substance solutions at different column temperatures in the durability test of Example 8.

[0054] Fig.16 This is an overlapping diagram of the test sample solutions at different column temperatures in the durability test of Example 8.

[0055] Fig.17This is an overlay diagram of the reference solution with different acetonitrile ratios of mobile phase A in the durability test of Example 8.

[0056] Fig.18 This is an overlap diagram of the test sample solutions with different acetonitrile ratios of mobile phase A in the durability test of Example 8. 5. Specific implementation methods

[0057] Specific embodiments of the present invention are described in detail below. It should be noted that the embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0058] Instrument and chromatographic conditions:

[0059] Instrument: High performance liquid chromatography, UV detector;

[0060] Chromatographic column: a chromatographic column with octylsilane bonded silica as filler, preferably Agilent ZORBAX SB-C8, 4.6mm×250mm, 5μm or a chromatographic column with equivalent performance;

[0061] Flow rate: 1.0ml / min;

[0062] Run time: at least 1 minute after the main peak of the sensitivity and reference solution is collected; 45 minutes for the test solution;

[0063] Detection wavelength: 238nm.

[0064] Reagent preparation: Keep away from light and prepare freshly before use.

[0065] Preparation of test solution: Take about 50 mg of varenicline tartrate, accurately weigh it, place it in a 50 ml volumetric flask, add mobile phase A to dissolve and dilute to the scale, and shake well.

[0066] Preparation of reference solution: Take about 4.8 mg of impurity D reference substance, accurately weigh it, place it in a 200 ml volumetric flask, add mobile phase A and sonicate to dissolve it, dilute it to the scale with mobile phase A, shake well, accurately measure 1 ml, place it in a 100 ml volumetric flask, add mobile phase A to dilute it to the scale, shake well, and prepare a solution containing about 0.24 μg per 1 ml.

[0067] Sensitivity solution: Accurately measure 5 ml of the reference solution, place it in a 10 ml volumetric flask, dilute to the mark with mobile phase A, and shake well.

[0068] Example 1 Selection of chromatographic conditions

[0069] Based on the above instruments and chromatographic conditions, continue to screen the most suitable chromatographic conditions.

[0070] Test condition 1:

[0071] Mobile Phase:

[0072] Mobile phase A: 50 mmol / L sodium perchlorate solution (pH adjusted to 2.5 with phosphoric acid);

[0073] Mobile phase B: acetonitrile;

[0074] Gradient elution program:

[0075]

[0076]

[0077] Column temperature: 20°C;

[0078] Injection volume: 30 μl;

[0079] Solvent: Mobile phase A-acetonitrile (80:20).

[0080] Preparation of impurity D localization solution: Take an appropriate amount of impurity D reference substance, weigh accurately, dissolve in solvent and dilute to make a solution containing approximately 0.24 μg per 1 ml.

[0081] Preparation of impurity interference solution: Take varenicline tartrate and other common impurities, add solvent to dissolve and dilute to make a mixed solution containing about 0.1 mg of varenicline tartrate and 1 μg of each impurity per 1 ml.

[0082] Accurately measure 30 μl of impurity D positioning solution and impurity interference solution, respectively, inject them into the chromatograph, and record the chromatogram.

[0083] The results show that the impurity interference solution has a chromatographic peak at the peak of impurity D. The interference impurity D is checked. Figure 1 .

[0084] Test condition 2:

[0085] Mobile Phase:

[0086] Mobile phase A: 50 mmol / L sodium perchlorate solution (pH adjusted to 2.5 with phosphoric acid)-acetonitrile (80:20);

[0087] Mobile phase B: 50 mmol / L sodium perchlorate solution (pH adjusted to 2.5 with phosphoric acid)-acetonitrile (50:50);

[0088] Gradient elution program:

[0089]

[0090] Column temperature: 35 °C;

[0091] Solvent: Mobile phase A.

[0092] The preparation of impurity D localization solution, interfering solution and other chromatographic conditions were the same as those in test condition 1.

[0093] Accurately measure 30 μl each of the impurity D positioning solution, impurity interference solution, and blank solvent, inject them into the chromatograph respectively, and record the chromatogram.

[0094] The results show that: the impurity interference solution has no chromatographic peak at the peak of impurity D, and the impurities in the impurity interference solution do not interfere with the impurity D inspection. Figure 2 The blank solvent does not produce a peak at the position of impurity D and does not interfere with the inspection of impurity D. Figure 3 .

[0095] The method in test condition 2 was determined as the inspection method for impurity D in varenicline tartrate.

[0096] Example 2 Accuracy Test

[0097] Stock solution of reference substance: Take about 6 mg of impurity D reference substance, accurately weigh it, place it in a 25 ml volumetric flask, add solvent to dissolve and dilute to the scale, shake well, accurately measure 1 ml, place it in a 100 ml volumetric flask, dilute to the scale with solvent, shake well, and obtain.

[0098] Quantitative limit stock solution: Take about 6 mg of impurity D reference substance, accurately weigh, place in a 25 ml volumetric flask, add solvent to dissolve and dilute to the scale, shake well, accurately measure 0.3 ml, place in a 100 ml volumetric flask, dilute to the scale with solvent, shake well, and obtain.

[0099] Recovery solution: Take three portions of 2.5ml, 5ml and 7.5ml of the reference stock solution respectively, place them in 9 50ml volumetric flasks, add 10ml of the blank test sample stock solution to each, dilute to the scale with solvent, shake well, and use them as 50% recovery, 100% recovery and 150% recovery test solutions respectively.

[0100] Quantitative limit recovery solution: Take three 5ml portions of the quantitation limit stock solution and place them in three 50ml volumetric flasks. Add 10ml of the blank test sample stock solution, dilute to the scale with solvent, shake well, and the solution is ready.

[0101] Accurately measure 30 μl of the test solution, quantitative limit recovery solution, and recovery solution, respectively, and inject them into the liquid chromatograph to record the chromatogram; calculate the detection amount according to the external standard method, and correct it with the blank solution, calculate the recovery rate, and calculate the relative standard deviation of the results.

[0102] The results showed that the average recovery rate was in the range of 80-120%, and the relative standard deviation was less than 10%, which met the requirements and the method had good accuracy. The results are shown in Table 1 and Figure 4 .

[0103] The accuracy recovery rate was calculated by measured amount / added amount × 100%.

[0104] Table 1 Impurity D recovery test results

[0105]

[0106] Example 3 Solution stability test

[0107] Stability of reference solution

[0108] The stability of the reference solution was determined by examining the peak area change of the reference solution at different times. The reference solution was placed at room temperature and protected from light, and then injected at 0, 2, 3, 4, and 8 hours. The results are shown in Table 2. Figure 5 .

[0109] The results showed that when the reference solution was placed in a dark environment for 40 minutes, the change in the peak area of ​​impurity D was -4.86%, which was less than 5%; when placed in natural light for 20 minutes, the change in the peak area of ​​impurity D was -5.06%, which was greater than 5%, indicating that within 8 hours, the peak area of ​​impurity D in the reference solution gradually decreased, and the maximum change rate was 47.84%, which was greater than 5%. The reference solution was unstable within 8 hours, and the stability of the reference solution was poor, and it had to be prepared fresh before use.

[0110] Table 2 Results of stability test of reference solution

[0111] Time (hours) 0 2 3 4 8 Maximum change rate (%) Impurity D peak area 13212 10593 9467 8872 6892 47.84

[0112] Stability of test solution

[0113] The stability of the test solution is achieved by examining the change in impurity content of the test solution at different times.

[0114] Take an appropriate amount of the test solution, place it at room temperature and avoid light, and inject samples at 0, 2, 5, 8, 16, and 24 hours. The results show that impurity D was not detected within 24 hours. Figure 6 The test solution is stable within 24 hours. Since the stability of impurity D is poor, in order to prevent the degradation of impurity D during the stability study, the test solution still needs to be freshly prepared before use.

[0115] Table 3 Test results of the stability of the test solution

[0116]

[0117] Example 4 Linearity Test

[0118] Impurity D stock solution: Take 15.53 mg of impurity D reference substance, accurately weigh it, put it in a 25 ml volumetric flask, add solvent to dissolve and dilute to the scale, shake well, and obtain.

[0119] Linear stock solution: Accurately measure 0.4 ml of impurity D stock solution, place it in a 100 ml volumetric flask, dilute to the scale with solvent, shake well, and obtain.

[0120] Quantitation limit solution: Take an appropriate amount of impurity D stock solution, add solvent to dilute and gradually dilute to a linear solution with a measured main peak signal-to-noise ratio ≥ 10.

[0121] Linear solution: Accurately measure 1 ml, 2.5 ml, 5 ml, 7.5 ml and 10 ml of the linear stock solution, place them into 5 25 ml volumetric flasks, dilute to the scale with solvent, shake well, and use them as solutions with concentrations of 40%, 100%, 200%, 300% and 400% respectively.

[0122] Accurately measure 30 μl of the quantitative limit solution and the linear 40% to linear 400% solution, respectively, and inject them into the liquid chromatograph, record the chromatogram, plot the peak area against the concentration, and calculate the regression equation and correlation coefficient by the least squares method. The results are shown in Table 4 and Figure 7 The results showed that the impurity D had a good linear relationship within the concentration range of 0.072 to 0.980 μg / ml with r ≥ 0.990 and intercept ≤ 25%.

[0123] Table 4 Impurity D linearity test results

[0124] Sample name Concentration range (μg / ml) Linear equations Correlation coefficient intercept Impurity D 0.072～0.980 y=50561x+341.92 r＝0.9997 2.74

[0125] Example 5 Quantitative Limit and Detection Limit Test

[0126] Preparation of quantitative limit solution: see Example 4 linearity test.

[0127] Preparation of detection limit solution: dilute the quantitation limit solution 5 times.

[0128] Accurately measure 30 μl of the quantitative limit solution and the detection limit solution, inject them into the liquid chromatograph, and record the chromatogram. The results are shown in Table 5. The results show that the quantitative limit concentration is less than 50% of the limit concentration, and the method has high sensitivity.

[0129] Table 5 Results of quantitative limit and detection limit of impurity D

[0130]

[0131] Example 6 Range Test

[0132] Preparation of reference solution with a limit concentration of 150%: Take about 6 mg of impurity D reference substance, place it in a 25 ml volumetric flask, add solvent to dissolve and dilute to the scale, shake well, and use it as the impurity stock solution; measure 0.15 ml of the impurity stock solution, place it in a 100 ml volumetric flask, dilute to the scale with solvent, shake well, and obtain.

[0133] Quantitation limit solution: see Example 4 linearity test.

[0134] The quantitative limit solution and the reference solution with a concentration of 150% of the limit were accurately measured and injected 6 times continuously. The relative standard deviation of the peak area of ​​impurity D and the main component was calculated. The results are shown in Table 6, and the quantitative limit concentration overlap diagram is shown in Figure 8 , limit 150% concentration overlap diagram see Fig. 9 The results showed that in the chromatogram of the quantitative limit solution, the RSD of the impurity D peak area was ≤20%, and in the chromatogram of the limit 150% concentration solution, the RSD of the impurity D peak area was ≤5%, which met the requirements.

[0135] Table 6 Verification results of the inspection range of impurity D

[0136]

[0137] Example 7 Precision Test

[0138] The precision test is carried out through three items: system precision, method repeatability and intermediate precision.

[0139] System precision

[0140] Take the solution with a linear concentration of 100% under the linear test item of Example 4, inject 6 injections continuously, record the chromatogram, and calculate the relative standard deviation of the peak area of ​​impurity D. The results are shown in Table 6 and Fig.10 The results showed that the RSD of the peak area of ​​impurity D was less than 10%, which met the acceptance criteria.

[0141] Table 7 Impurity D system precision test results

[0142]

[0143] Method reproducibility

[0144] Personnel A prepared the same batch of test solution 6 times, calculated the content of impurity D by external standard method, and calculated the relative standard deviation of the 6 determination results. The results are shown in Table 8. The results show that Personnel A measured impurity D 6 times in parallel, and no impurity D was detected, with good repeatability.

[0145] Intermediate precision

[0146] Personnel B repeated the "method repeatability" verification on different dates and with different instruments, and calculated the relative deviation of the repeatability and intermediate precision results, and the results are shown in Table 8. The results show that personnel A and B respectively measured impurity D 6 times on different dates, using different instruments and chromatographic systems, and no impurity D was detected, and the intermediate precision of the method was good.

[0147] Table 8 Impurity D repeatability and intermediate precision test results

[0148]

[0149] Example 8 Durability Test

[0150] The durability is mainly investigated through the durability of the chromatographic conditions.

[0151] Based on the established chromatographic conditions, three chromatographic columns with the same filler but different brands, mobile phases with different pH values, different column temperatures, and different acetonitrile ratios in mobile phase A were used to determine the impurity D in varenicline tartrate to investigate the robustness of the method. Only one parameter was changed in each test, and the results were compared with those obtained without changing the parameters.

[0152] Using 3 different models and brands of C8 chromatographic columns, the pH value of the buffer salt varied within the range of 2.5±0.5, the column temperature varied within the range of 35℃±5℃, and the proportion of acetonitrile in the mobile phase A varied within the range of 20%±2% (parameters are detailed in Table 9). All of them met the system suitability test requirements, and impurity D was not detected, indicating that the method has good durability under these conditions. The results are shown in Table 10 and Figures 11 to 18 .

[0153] Table 9 Impurity D Check the robustness of the HPLC method

[0154]

[0155] Table 10 Impurity D inspection durability test results

[0156]

[0157] Note: * indicates standard conditions.

Claims

1. A method for detecting the genotoxic impurity D in varenicline tartrate, It is characterized in that The method comprises the following steps: respectively sampling a sensitivity solution, a reference solution and a test solution, testing them according to high performance liquid chromatography conditions, and recording a chromatogram; the chromatographic conditions are: a chromatographic column: a chromatographic column with octylsilane bonded silica gel as a filler; Mobile phase: Mobile phase A is a mixed solution of buffer solution and organic solvent, and mobile phase B is a mixed solution of buffer solution and organic solvent. Gradient elution is performed according to the following table: The structure of the impurity D is shown below:

2. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 1, It is characterized in that The chromatographic conditions also include: Detector: UV detector; Flow rate: 1.0ml / min; Column temperature: 30-40°C; Detection wavelength: 238nm; Injection volume: 30 μl.

3. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 1, It is characterized in that The chromatographic column is selected from Agilent ZORBAX RX-C8, 250 mm × 4.6 mm, 5 μm, Agilent ZORBAX SB-C 8 , 4.6mm×250mm, 5μm, Agela Venusil XBP C 8 , 4.6mm×250mm, 5μm.

4. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 1, It is characterized in that The buffer solution is selected from one or more of sodium perchlorate solution, potassium dihydrogen phosphate solution, and sodium dihydrogen phosphate solution, and the organic solvent is selected from one or more of acetonitrile, methanol, and tetrahydrofuran.

5. The method for detecting the genotoxic impurity D of varenicline tartrate according to claim 4, It is characterized in that The pH value of the sodium perchlorate buffer is adjusted to 2.0-3.0 with an acidic reagent, and the concentration thereof is 30-70 mmol / L.

6. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 5, It is characterized in that The acidic reagent is selected from one or more of glacial acetic acid, phosphoric acid and formic acid.

7. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 1, It is characterized in that The ratio of the buffer solution to the organic solvent in the mobile phase A is 78-82:22-18.

8. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 1, It is characterized in that The ratio of the buffer solution to the organic solvent in the mobile phase B is 50:

50.

9. The method for detecting genotoxic impurity D of varenicline tartrate according to claim 2, It is characterized in that The column temperature was 35°C.

10. Use of the method for detecting genotoxic impurity D in varenicline tartrate according to claim 1 in quality control of varenicline tartrate.

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