Method for determining content of dibromohydantoin in anastrozole starting material by ion chromatography

Abstract

The invention relates to the field of analytical chemistry, in particular to a method for determining the content of dibromohydantoin in an anastrozole starting material through ion chromatography. The invention provides a method capable of precisely and accurately determining the content of dibromohydantoin in the anastrozole starting material, and the method is simple, convenient, sensitive and good in specificity, and is used for quality control of the anastrozole starting material penta-methyl-1, 3-diethyl cyanobenzene, so that the quality of an anastrozole raw material medicine is ensured.

Description

Technical Field

[0001] The invention relates to the field of analytical chemistry, and in particular to a method for determining the content of dibromohydantoin in anastrozole starting materials by using an ion chromatography method. Background Art

[0002] According to data from the World Health Organization's International Cancer Research Center, breast cancer is a common clinical malignancy, and its incidence rate has shown an upward trend, seriously endangering human life and health. At present, the main treatments for breast cancer include surgery, chemotherapy, targeted drug therapy, and endocrine drug therapy. Compared with other patients, for patients with advanced breast cancer who are receiving chemotherapy, if endocrine therapy can be used to specifically inhibit hormone receptors, the proliferation of breast cancer cells can be significantly inhibited.

[0003] Among the drugs used to treat breast cancer, tamoxifen was the first to be used, but tamoxifen can cause serious liver and kidney damage and lipid metabolism disorders, and increase the incidence of cardiovascular complications. Anastrozole is a selective non-steroidal aromatase inhibitor, with a chemical name of α,α,α',α-tetramethyl-5-(1H-1,2,4-triazol-1-yl-methyl)-1,3-diacetonitrilebenzene, which can inhibit hormone synthesis and significantly reduce hormone metabolism disorders. Anastrozole was patented in 1987 and launched in 1995. The original manufacturer of anastrozole tablets is AstraZeneca. It is currently the most commonly used breast cancer treatment drug in clinical practice. It is mainly suitable for the treatment of advanced breast cancer in postmenopausal women, for the adjuvant treatment of hormone receptor-positive early breast cancer in postmenopausal women, and for the adjuvant treatment of hormone receptor-positive early breast cancer in postmenopausal women who have received 2 to 3 years of tamoxifen adjuvant treatment.

[0004]

[0005] The synthesis process of anastrozole raw material is to use penta-methyl-1,3-diethylcyanobenzene as the starting material and synthesize intermediates and final products in sequence according to the route.

[0006]

[0007] In the synthesis process of penta-methyl-1,3-diethylcyanobenzene, dibromohydantoin is used as a brominating agent to participate in the reaction:

[0008]

[0009] Whether the impurities in the production process of Anastrozole can be fully and accurately controlled will directly affect the safety, controllability and quality stability of the drug. Therefore, the study of various impurities in the starting materials of Anastrozole is a key factor in the quality assurance of raw materials. It is to ensure the smooth, orderly and efficient connection of each step of the reaction and provide high-quality products. According to the synthesis route map of the starting material of Anastrozole, penta-methyl-1,3-diethylcyanobenzene, dibromohydantoin participates in the reaction as a brominating agent during the synthesis process of penta-methyl-1,3-diethylcyanobenzene. Therefore, it is necessary to control the residual amount of dibromohydantoin in the starting material and formulate reasonable limits according to the ICH Q3 guidelines to ensure the quality of Anastrozole.

[0010] At present, there are few reports on the detection method of dibromohydantoin in medicines. "Study on the Residual and Digestion of Bromide Ions Produced by Dibromohydantoin Acting on Tilapia" discloses a method for detecting bromide ions produced by dibromohydantoin acting on Gifu Tilapia, but this method is not suitable for the detection of dibromohydantoin in medicines. CN114216990A discloses a method for detecting dibromohydantoin in brominated trityl tetrazolyl biphenyl. This method uses a headspace gas chromatograph equipped with an ECD detector to detect the content of dibromohydantoin in brominated trityl tetrazolyl biphenyl. However, the boiling points of penta-methyl-1,3-diethylcyanobenzene and dibromohydantoin, the starting materials of anastrozole, are close, and the above method cannot separate the two well. At present, there is no literature reporting the relevant detection method of dibromohydantoin in penta-methyl-1,3-diethylcyanobenzene. Summary of the invention

[0011] Based on the above technical status, the purpose of the present invention is to provide a method for precisely and accurately determining the content of dibromohydantoin in anastrozole starting materials. The method is simple, sensitive and has good specificity, and is used for quality control of anastrozole starting material penta-methyl-1,3-diethylcyano, thereby ensuring the quality of anastrozole raw materials.

[0012] In order to achieve the above technical purpose, the technical solution adopted by the present invention is as follows:

[0013] A method for determining the content of dibromohydantoin in anastrozole starting materials by ion chromatography comprises the following steps:

[0014] The solution obtained by fully reacting sodium triacetoxyborohydride and dibromohydantoin is used as the reference solution;

[0015] The solution obtained by fully reacting sodium triacetoxyborohydride and anastrozole starting materials is used as the test solution;

[0016] The reference solution and the test solution are detected by an ion chromatograph, and the content of dibromohydantoin in the anastrozole starting material is calculated by an external standard method; the chromatographic column is an anion exchange column, and the eluent is an alkaline aqueous solution.

[0017] Dibromohydantoin can decompose into hypobromous acid in water, releasing bromide ions, and is more easily decomposed in strong acids and alkalis. Therefore, in the prior art, dilute sulfuric acid is added to promote its decomposition when determining dibromohydantoin. The present invention has found through experiments that the addition of a reducing agent can make dibromohydantoin decompose more thoroughly, thereby making the test results more accurate. However, different reducing agents have different reduction effects in specific drugs. The present invention selects sodium triacetoxyborohydride as a reducing agent, which can fully reduce dibromohydantoin in the presence of penta-methyl-1,3-diethylcyanobenzene. The generated bromide ions are adsorbed by an anion exchange column and then eluted with an alkaline aqueous solution. The bromide ion content can be accurately determined, and then the content of dibromohydantoin can be converted.

[0018] In some embodiments, the concentration of dibromohydantoin in the reference solution is 1-10 μg / ml, and the mass ratio of sodium triacetoxyborohydride to dibromohydantoin is 50-100: 1. The concentration of sodium acetoxyborohydride in the test solution is 0.08-1 mg / ml, and the mass ratio of anastrozole starting material to sodium triacetoxyborohydride is 10-15: 1. Under the above conditions, the reduction reaction can be fully carried out, and excess sodium triacetoxyborohydride will not have a negative impact on the detection.

[0019] In some embodiments, the solvent of the reference solution and the test solution is a mixed solution of an organic solvent and water, wherein the volume content of the organic solvent does not exceed 8%; and the order of adding the solvent is: first dissolve the solid material with an organic solvent and then add water to make up the volume. Since penta-methyl-1,3-diethylcyanobenzene is almost insoluble in water, an organic solvent needs to be added to dissolve it, and the organic solvent is easy to damage the ion chromatography suppressor, so the present invention controls the content of the organic solvent to be no more than 8vt%. The addition method of sodium triacetoxyborohydride also has a certain influence on the test results. Adding it in a solid form can make the reaction more complete, while first preparing it into an aqueous solution and then adding it will not cause the reduction reaction to be sufficient, and the test results are inaccurate. Sodium triacetoxyborohydride can react with water, and the generated borane is also reducing, but it does not have a positive effect on the reduction of dibromohydantoin.

[0020] In some embodiments, the organic solvent is methanol, acetonitrile, toluene, ethyl acetate or dichloromethane, preferably acetonitrile. Acetonitrile has good solubility for penta-methyl-1,3-diethylcyanobenzene, dibromohydantoin and sodium triacetoxyborohydride, and can remain dissolved after dilution with water, ensuring the accuracy of the test results.

[0021] In some embodiments, the reduction reaction time is 0.1-24 hours. In order to fully carry out the reaction, ultrasound or oscillation can be used to shorten the reaction time. There is no special requirement for the reaction temperature, which can be at natural temperature or heated to a temperature not higher than the boiling point of the solution.

[0022] In some embodiments, the alkaline aqueous solution is a sodium hydroxide aqueous solution or a potassium hydroxide aqueous solution, and the concentration of the eluent is 8-11 mmol / L. At this concentration, the eluent can fully and efficiently elute the bromide ions to ensure the accuracy of the test results.

[0023] In some embodiments, the elution time of the eluent is 13-15 minutes; after the elution is completed, the system is flushed with an alkaline aqueous solution with a concentration of 35-40 mmol / L. Specifically, the mobile phase can be divided into liquid A and liquid B. Liquid A is an alkaline aqueous solution with a concentration of 8-11 mmol / L, which is used as an eluent to elute bromide ions, and liquid B is an alkaline aqueous solution with a concentration of 35-40 mmol / L, which is used as a system flushing liquid. Specifically, the following elution procedure can be used:

[0024] Time (min) A(％) B(％) 0 100 0 14.0 100 0 14.5 0 100 24.0 0 100 24.5 100 0 30.0 100 0

[0025] In some embodiments, the chromatographic column is Dionex TM IonPac TM AS19.

[0026] In some embodiments, a guard column is further provided before the chromatographic column.

[0027] In some embodiments, the guard column is Dionex IonPac™ AG 19.

[0028] In some embodiments, the detector of the ion chromatograph is a suppressed conductivity detector, with a column temperature of 25-35° C. and a flow rate of 1-1.5 ml / min.

[0029] Chromatographic conditions are important factors that affect the test results. The present invention studies the chromatographic column, column temperature, flow rate, etc., and ultimately determines the chromatographic conditions that, while ensuring separation, precision, accuracy, etc., also shorten the test time as much as possible. The test of a sample can be completed within 30 minutes.

[0030] In some embodiments, the content of dibromohydantoin in the test sample is calculated according to the external standard method, and the calculation formula is as follows:

[0031]

[0032] W std is the sample weight of dibromohydantoin reference substance, mg;

[0033] P is the content of dibromohydantoin reference substance;

[0034] A spl is the main peak area in the test solution;

[0035] V spl is the dilution volume of the test sample, ml;

[0036] A std is the main peak area in the reference solution;

[0037] V std is the total dilution volume of the reference substance, ml;

[0038] W spl is the sample weight of the test product, mg.

[0039] In summary, the method of the present invention can fully release the bromide ions in dibromohydantoin, and then accurately and precisely determine the bromide ion content, and further convert the content of dibromohydantoin according to the bromide ion content. The method is simple, sensitive, and has good specificity, which provides a guarantee for the quality control of anastrozole starting materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 It is the chromatogram of the blank solution in the specificity test;

[0041] Figure 2 It is the chromatogram of the potassium bromide positioning solution in the specificity test;

[0042] Figure 3 It is the chromatogram of the reference solution in the specificity test;

[0043] Figure 4 It is the chromatogram of the 10% level spike solution in the accuracy test;

[0044] Figure 5 It is the chromatogram of the 100% level loading solution in the accuracy test;

[0045] Figure 6 It is the chromatogram of the 150% level loading solution in the accuracy test;

[0046] Figure 7 is a chromatogram of a batch of samples in Example 2;

[0047] Figure 8 is a chromatogram of the reference solution in Comparative Example 1 without sodium triacetoxyborohydride reduction;

[0048] Fig. 9 This is the chromatogram of the reference solution after adding dilute sulfuric acid treatment in Comparative Example 1;

[0049] Fig.10 is a chromatogram of the reference solution reduced with sodium nitrite in Comparative Example 2;

[0050] Fig.11 is a chromatogram of the reference solution reduced with sodium sulfite in Comparative Example 2;

[0051] Fig.12It is the chromatogram of the blank solution in Comparative Example 5;

[0052] Fig.13 It is the chromatogram of the reference substance solution in Comparative Example 5;

[0053] Fig.14 is the chromatogram of the reference substance solution in Comparative Example 6;

[0054] Fig.15 is the chromatogram of the reference solution in Comparative Example 7;

[0055] Fig.16 It is the chromatogram of the reference substance solution in Comparative Example 8.

[0056] In the above chromatogram, the peak with "Br" after the peak number is the bromide ion peak. DETAILED DESCRIPTION

[0057] Example 1 Methodological Experiment

[0058] 1. Test instruments, material information and test methods

[0059] 1.1 Test equipment

[0060] Ion chromatograph: (Thermo Fisher Scientific Inc. ICS-6000), chromatographic column Dionex IonPac TM AS19 (4×250 mm), guard column: Dionex IonPac TM AG 19 (4×50 mm), the detector is a conductivity detector, and the suppressor is Dionex TM ASRS TM 300 4mm; electronic balance (XPE105 from Mettler, Switzerland); water bath constant temperature oscillator (SHA-B from Tianjin Guangfeng Technology Co., Ltd.); CNC ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd., Model KQ-500DE).

[0061] 1.2 Test reagents

[0062] Acetonitrile (HPLC, Merck); sodium triacetoxyborohydride (purity: 97%, Maya reagent); potassium bromide (spectral grade, Merck); sodium hydroxide solution (purity: 50w / w%, Alfa Aesar); dibromohydantoin (purity: 98.3%, Shandong Xiya Chemical Co., Ltd.); three batches of penta-methyl-1,3-diethylcyanobenzene (Changzhou Liren Pharmaceutical Technology Co., Ltd., batch numbers LR-TMB-200723, LR-TMB-221208, LR-TMB-221224) of anastrozole starting material

[0063] 1.3 Chromatographic conditions

[0064] Chromatography Column Dionex IonPac TM AS19 (4×250 mm), guard column: Dionex IonPac TM AG 19 (4×50mm), eluent A is 9.5mmol / L sodium hydroxide solution (precisely measure 1ml of 50% sodium hydroxide solution and put it into 2000ml purified water, shake well); eluent B is 38mmol / L sodium hydroxide solution (precisely measure 4ml of 50% sodium hydroxide solution and put it into 2000ml purified water, shake well). The elution procedure is shown in Table 1; the detector is a conductivity detector, and the suppressor is Dionex TM ASRS TM 3004mm, column temperature 30℃, flow rate 1.2ml / min.

[0065] Table 1 Elution program

[0066] Time (min) A(％) B(％) 0 100 0 14.0 100 0 14.5 0 100 24.0 0 100 24.5 100 0 30.0 100 0

[0067] 1.4 Solution preparation

[0068] Blank solution: Weigh 20.24 mg of sodium triacetoxyborohydride reagent into a 100 ml volumetric flask, accurately measure 5 ml of acetonitrile, dilute to the scale with water, and shake well.

[0069] Potassium bromide positioning solution: Weigh 10.77 mg of potassium bromide reagent into a 100 ml volumetric flask, dilute to the scale with water, and shake well.

[0070] Stock solution of reference substance: Weigh 12.56 mg of dibromohydantoin reference substance accurately, place in a 50 ml volumetric flask, dissolve and dilute to the mark with water, and shake well.

[0071] Reference substance solution: Weigh 19.99 mg of sodium triacetoxyborohydride reagent and place it in a 100 ml volumetric flask. Accurately measure 5 ml of acetonitrile and accurately measure 1 ml of reference substance stock solution and place it in the same volumetric flask. Add 20 ml of water, shake for 1 hour, dilute to the scale with water, and shake well.

[0072] Test solution: Take about 250 mg of penta-methyl-1,3-diethylcyanobenzene, the starting material of anastrozole, accurately weigh it, and place it in a 100 ml volumetric flask. Then weigh about 20 mg of sodium triacetoxyborohydride reagent and place it in the same volumetric flask. Accurately measure 5 ml of acetonitrile to dissolve the anastrozole starting material, add 20 ml of water, shake for 1 hour, dilute with water to the scale, shake well, and filter through a 0.45 μm nylon membrane to obtain.

[0073] 1.5 Test methods

[0074] Inject the reference solution and the test solution into the ion chromatograph respectively, and measure according to the chromatographic conditions under 1.3. Calculate the content of dibromohydantoin in the test sample according to the external standard method:

[0075]

[0076] W std is the sample weight of dibromohydantoin reference substance, mg;

[0077] P is the content of dibromohydantoin reference substance;

[0078] A spl is the main peak area in the test solution;

[0079] V spl is the dilution volume of the test sample, ml;

[0080] A std is the main peak area in the reference solution;

[0081] V std is the total dilution volume of the reference substance, ml;

[0082] W spl is the sample weight of the test product, mg.

[0083] 2 Methodological validation

[0084] 2.1 Exclusivity

[0085] Accurately measure 100 μl of blank solution, potassium bromide solution and reference solution, and inject them into the ion chromatograph according to the chromatographic parameters under 1.3. The chromatogram is shown in Figure 1-3 Results: The blank solution had no interference with the determination of bromide ions, and the retention time of the main peak of the dibromohydantoin reference solution was consistent with that of the potassium bromide positioning solution, indicating that the method had good specificity.

[0086] 2.2 Limit of detection and limit of quantification

[0087] Weigh 20.37 mg of sodium triacetoxyborohydride reagent and place it in a 100 ml volumetric flask. Accurately measure 5 ml of acetonitrile, and then accurately measure an appropriate amount of diluted reference stock solution (quantitative limit solution: the signal-to-noise ratio of dibromohydantoin peak is not less than 10; detection limit solution: the signal-to-noise ratio of dibromohydantoin peak is not less than 3) and place it in the same volumetric flask, add 20 ml of water, shake for 1 hour, dilute with water to the scale, and shake well. Accurately measure 100 μl of quantitative limit solution and record the chromatogram. Results: The concentration of the quantitative limit solution of dibromohydantoin is 0.2422 μg / ml, equivalent to 10% of the limit, and the concentration of the detection limit solution is 0.0727 μg / ml, equivalent to 3% of the limit, which meets the determination requirements.

[0088] 2.3 Linearity and range of dibromohydantoin

[0089] Weigh 20.61 mg of sodium triacetoxyborohydride reagent and place it in a 100 ml volumetric flask. Accurately measure 5 ml of acetonitrile, and then accurately measure a series of concentrations of dibromohydantoin reference solution and place it in the same volumetric flask. Add 20 ml of water, shake for 1 hour, dilute to the scale with water, and shake well. Accurately measure 100 μl of linear solution and record the chromatogram. Plot the measured dibromohydantoin peak area against the concentration, and perform linear regression using the least squares method. The linear equation is y=0.0628x-0.0055, R 2 The linear relationship of dibromohydantoin was good in the concentration range of 0.2422 to 4.8422 μg / ml.

[0090] 2.4 Method precision and repeatability

[0091] Six test solutions were prepared in parallel according to the test solution preparation method. According to the chromatographic parameters under 3.1, 100 μl of the test solution was accurately measured and the chromatogram was recorded. As a result, no dibromohydantoin was detected in the six test solutions, indicating that the method has good precision. Six test solutions were prepared in parallel according to the test solution preparation method by different experimenters on different dates. The chromatographic parameters under 3.1 were measured. As a result, no dibromohydantoin was detected in the six test solutions measured by different experimenters. The results show that the method has good reproducibility.

[0092] 2.5 Method accuracy

[0093] Take about 250 mg of penta-methyl-1,3-diethylcyanobenzene, the starting material of anastrozole, accurately weigh it, and place it in a 100 ml volumetric flask. Then weigh about 20 mg of sodium triacetoxyborohydride reagent and place it in the same volumetric flask. Weigh 9 portions in parallel, accurately measure 5 ml of acetonitrile respectively to dissolve the starting material of anastrozole, add 0.1 ml, 1 ml, and 1.5 ml of the reference substance stock solution, and then add 20 ml of water respectively, shake for 1 hour, dilute with water to the scale, shake well, and filter to obtain 10% (LOQ level), 100% and 150% level sample solutions (three solutions for each of the three concentration levels of low, medium and high). According to the chromatographic parameters under 3.1, accurately measure 100 μl of each solution, determine according to the law, and investigate the recovery rate of dibromohydantoin. See the chromatogram. Figure 4-Figure 6 The results were calculated by external standard method. The recovery of dibromohydantoin at each concentration level was in the range of 94% to 106%, with an average recovery of 102% and an RSD of 3.3%. The results showed that this method can accurately quantify the content of dibromohydantoin in the starting material of anastrozole.

[0094] 2.6 Method Durability

[0095] The chromatographic parameters were adjusted as follows: column temperature ±5°C, flow rate ±0.1ml / min, eluent A sodium hydroxide solution ±0.1ml. Other parameters remained unchanged. Chromatograms of blank solution, reference solution and test solution were collected. The results showed that the separation between the main peak and the adjacent peaks of the reference solution was greater than 1.5 under all changing conditions. No dibromohydantoin was detected in the test solution. The results showed that this method was durable.

[0096] Example 2 Sample determination

[0097] Weigh 3 different batches of anastrozole starting material samples, the sample weights were 250.50 mg, 249.99 mg and 250.17 mg respectively, and prepare according to the preparation method of the test solution. The amount of sodium triacetoxyborohydride added was 20.08 mg, 20.50 mg and 20.56 mg respectively. According to the chromatographic conditions under 1.3 and the test method under 1.5, the content of dibromohydantoin in the test sample was detected. The results showed that the three batches of samples did not have a peak at the corresponding retention time of the bromide ion peak (about 11.9 min). Figure 7 This is the chromatogram of one of the batches of samples. No dibromohydantoin was detected in any of the three batches of samples.

[0098] Comparative Example 1

[0099] No sodium triacetoxyborohydride was added to the reference solution. The reference stock solution was diluted with water and the same chromatographic conditions as in Example 1 were used for determination. The chromatogram is shown in FIG. Figure 8 The reference solution was not added with sodium triacetoxyborohydride, but 1 ml of concentrated sulfuric acid was added to promote the decomposition of dibromohydantoin. The same chromatographic conditions as in Example 1 were used for determination. The chromatogram is shown in FIG. Fig. 9 . Figure 3 , Figure 8 and Fig. 9 In the experiment, the peak areas of bromide ion peaks are 0.0688, 0.0312, and 0.1434, respectively. It can be seen that the addition of sodium triacetoxyborohydride for reduction can make the release of bromide ions more complete.

[0100] Comparative Example 2

[0101] The sodium triacetoxyborohydride in the reference solution was replaced by sodium nitrite and sodium sulfite, respectively, and the determination was carried out under the same chromatographic conditions as in Example 1. The chromatogram is shown in FIG. Figure 10-11 Compared with sodium triacetoxyborohydride as a reducing agent, the bromide ion peak and the adjacent solvent peak did not reach baseline separation when sodium nitrite was used for reduction. When sodium sulfite was used for reduction, the resolution of the unknown ion peak (peak 4) on the right side and the bromide ion peak was low (1.7), which interfered with the determination of bromide ion content. However, the resolution of the bromide ion peak and the adjacent peaks when sodium triacetoxyborohydride was used for reduction was 6.8.

[0102] Comparative Example 3

[0103] The sodium triacetoxyborohydride under the item "2.5 Method Accuracy" was replaced by sodium nitrite and sodium sulfite, respectively, and the 10% sample solution was measured according to the same method. Each reducing agent was measured in parallel three times, and the recovery rate of dibromohydantoin was calculated. The results are shown in Table 2.

[0104] Table 2 Effect of different reducing agents on accuracy

[0105]

[0106] It can be seen that when sodium sulfite and sodium nitrite are used as reducing agents, the recovery rate of the sample is low, and this method is not suitable for the detection of dibromohydantoin in the starting material of anastrozole.

[0107] Comparative Example 4

[0108] Prepare the stock solution of dibromohydantoin reference substance: take about 12.5 mg of dibromohydantoin reference substance, accurately weigh it, put it in a 50 ml volumetric flask, add appropriate amount of water to dissolve it, dilute it to the scale with water, and shake well.

[0109] Prepare reference solution:

[0110] Join Method 1:

[0111] Weigh about 1500 mg of sodium triacetoxyborohydride into a 100 ml volumetric flask, dissolve it with water and dilute to the mark, shake well. Accurately measure 20 ml of the above solution and 1 ml of dibromohydantoin reference stock solution into the same 100 ml volumetric flask, dilute to the mark with water, and sonicate for 2 hours. (Sodium triacetoxyborohydride: 3 mg / ml; dibromohydantoin: 2.5 μg / ml).

[0112] Join Method 2:

[0113] Weigh about 300 mg of sodium triacetoxyborohydride into a 100 ml volumetric flask, then accurately measure 1 ml of dibromohydantoin reference stock solution into the same volumetric flask, dissolve with water and dilute to scale, and sonicate for 2 hours. (Sodium triacetoxyborohydride: 3 mg / ml; dibromohydantoin: 2.5 μg / ml).

[0114] According to the preparation method of addition method 1 and 2, the addition of other concentrations of sodium triacetoxyborohydride was compared to investigate the effect of the addition method on the main peak area of ​​the reference solution.

[0115] Results: The main peak area of ​​dibromohydantoin reference substance with the same concentration of sodium triacetoxyborohydride and different addition methods is as follows: the main peak area of ​​sodium triacetoxyborohydride added to the reference substance solution in solution form is 0.0722, 0.0833, 0.1006, 0.1107, 0.1143 when sodium triacetoxyborohydride concentration is 0.2mg / ml, 1mg / ml, 1.5mg / ml, 2mg / ml, 3mg / ml, and the main peak area is 0.1497, 0.1392, 0.1420, 0.1427, 0.1449 when sodium triacetoxyborohydride is added to the reference substance solution in solid form. The results show that sodium triacetoxyborohydride reacts more completely when added in solid form.

[0116] Comparative Example 5

[0117] The eluent A was replaced with a sodium hydroxide solution with a concentration of 6.3 mmol / L. The preparation method was as follows: 1 ml of 50% (w / w) sodium hydroxide solution was added to 3000 ml of water and mixed. The other conditions were the same as those in Example 1. The blank solution and the reference solution were tested respectively. The results are shown in Table 1. Fig.12 and Fig.13 It can be seen that the blank solution has an interference peak (15.653min) at the position of the bromide ion peak, which affects the accuracy of the test results.

[0118] Comparative Example 6

[0119] The eluent A was replaced with a sodium hydroxide solution with a concentration of 12.3 mmol / L. The preparation method was as follows: 1.3 ml of 50% (w / w) sodium hydroxide solution was added to 2000 ml of water and mixed. The other conditions were the same as those in Example 1. The reference solution was tested. The results are shown in Table 1. Fig.14 It can be seen that when the eluent of this concentration is used, the baseline of the chromatogram of the reference solution fluctuates greatly, the chromatographic system is unstable, and this method is not suitable for the detection of dibromohydantoin.

[0120] Comparative Example 7

[0121] The eluent A was replaced with a sodium hydroxide solution with a concentration of 15.2 mmol / L. The preparation method was as follows: 2 ml of 50% (w / w) sodium hydroxide solution was added to 2500 ml of water and mixed. Other conditions were the same as those in Example 1. The reference solution was tested. The results are shown in Table 1. Fig.15 It can be seen that when the eluent of this concentration is used, the baseline of the chromatogram of the reference solution fluctuates greatly, the chromatographic system is unstable, and this method is not suitable for the detection of dibromohydantoin.

[0122] Comparative Example 8

[0123] The chromatographic column in Example 1 was replaced with Dionex IonPac TMAS11 (4×250 mm), guard column is Dionex IonPac TM AG 11 (4×50mm), the reference solution was determined using the same method, the chromatogram is shown in Fig.16 It can be seen that when the above chromatographic column is used, the bromide ion peak emerges earlier, fails to achieve baseline separation with the solvent peak, has a poor peak shape, and has a low column efficiency.

[0124] Example 2

[0125] In this embodiment, the eluent is a 9.5 mmol / L potassium hydroxide aqueous solution, and the rest is the same as in Example 1. The detection effect on dibromohydantoin is equivalent to that in Example 1.

[0126] Example 3

[0127] The concentration of sodium triacetoxyborohydride in the test solution of this embodiment is 0.8 mg / ml, and the rest is the same as that of embodiment 1. The detection effect on dibromohydantoin is equivalent to that of embodiment 1.

[0128] Other condition parameters within the protection scope of the method of the present invention can achieve the same detection effect as that of Example 1, and will not be described in detail here.

[0129] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modification, equivalent substitution or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for determining the content of dibromohydantoin in anastrozole starting material by ion chromatography, wherein the starting material is penta-methyl-1,3-diethylcyanobenzene, characterized in that: The steps include: The solution obtained by fully reacting sodium triacetoxyborohydride and dibromohydantoin is used as the reference solution; The solution obtained by fully reacting sodium triacetoxyborohydride and anastrozole starting materials is used as the test solution; The reference solution and the test solution are detected by an ion chromatograph, and the content of dibromohydantoin in the anastrozole starting material is calculated by an external standard method; the chromatographic column is an anion exchange column, and the eluent is an alkaline aqueous solution.

2. The method according to claim 1, characterized in that The concentration of dibromohydantoin in the reference solution is 1-10 μg / ml, and the mass ratio of sodium triacetoxyborohydride to dibromohydantoin is 50-100:

1.

3. The method according to claim 1, characterized in that The concentration of sodium acetoxyborohydride in the test solution is 0.08-1 mg / ml, and the mass ratio of anastrozole starting material to sodium triacetoxyborohydride is 10-15:

1.

4. The method according to claim 1, characterized in that: The solvents of the reference solution and the test solution are mixed solutions of organic solvent and water, wherein the volume content of the organic solvent does not exceed 8%; and the order of adding the solvents is: first dissolving the solid material with the organic solvent and then adding water to make up the volume.

5. The method according to claim 4, characterized in that The organic solvent is methanol, acetonitrile, toluene, ethyl acetate or dichloromethane.

6. The method according to any one of claims 1 to 5, characterized in that: The reaction time for preparing the reference solution and the test solution is 0.1-24h.

7. The method according to any one of claims 1 to 5, characterized in that: The alkaline aqueous solution is a sodium hydroxide aqueous solution or a potassium hydroxide aqueous solution, and the concentration of the eluent is 8-11 mmol / L.

8. The method according to claim 7, characterized in that The elution time of the eluent is 13-15 min; and / or: After elution, flush the system with an alkaline aqueous solution with a concentration of 35-40mmol / L for 13-15 minutes.

9. The method according to any one of claims 1 to 5, characterized in that: The chromatographic column is AS19.

10. The method according to any one of claims 1 to 5, characterized in that: A guard column is also arranged before the chromatographic column.

11. The method according to any one of claims 1 to 5, characterized in that: The detector of the ion chromatograph is a suppressed conductivity detector, with a column temperature of 25-35° C. and a flow rate of 1-1.5 ml / min.

12. The method according to any one of claims 1 to 5, characterized in that: The content of dibromohydantoin in the anastrozole starting material was calculated using the external standard method according to the following formula: W std is the sample weight of dibromohydantoin reference substance, mg; P is the content of dibromohydantoin reference substance; A spl is the main peak area in the test solution; V spl is the dilution volume of the test sample, ml; A std is the main peak area in the reference solution; V std is the total dilution volume of the reference substance, ml; W spl is the sample weight of the test product, mg.

Images