Method for detecting specific impurities in 2-bromo-4, 5-dimethoxybenzyl bromide

Abstract

The invention discloses a method for detecting specific impurities in 2-bromo-4, 5-dimethoxybenzyl bromide in the field of medical analysis, which is characterized in that octadecylsilane chemically bonded silica is used as a detection chromatographic column, acetonitrile and water are used as mobile phases, and high performance liquid chromatography is carried out. The method has the advantages that the response value is high, the 3, 4-dimethoxybenzyl alcohol and the 2-bromo-4, 5-dimethoxybenzaldehyde impurities in the 2-bromo-4, 5-dimethoxybenzyl bromide can be effectively separated and detected, the detection operation is simple, the product purity and impurity content detection result accuracy is high, the system applicability, specificity, linearity and range all meet the requirements, and the method is suitable for industrial production. The problems of purity detection and impurity control of the substance are solved.

Description

Technical Field

[0001] The invention relates to the field of pharmaceutical analysis technology, and in particular to a method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide, an intermediate of pinaverium bromide. The specific impurities are 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde. Background Art

[0002] 2-Bromo-4,5-dimethoxybenzyl bromide is an intermediate of pinaverium bromide. Its appearance is white to light brown powder. Its CAS number is 53207-00-4. Its structural formula is as follows:

[0003]

[0004] The current process route for 2-bromo-4,5-dimethoxybenzyl bromide is as follows:

[0005]

[0006] The specific impurity structural formula is as follows:

[0007]

[0008] 3,4-Dimethoxybenzyl alcohol is an intermediate in the reaction of 2-bromo-4,5-dimethoxybenzyl bromide, and 2-bromo-4,5-dimethoxybenzaldehyde is a process impurity produced by residual starting materials. The presence of these two specific impurities will affect the quality and yield of the product. Because they are similar in structure to the target product, there is currently no method that can effectively detect these two specific impurities.

[0009] Therefore, in order to accurately, simply and effectively control the quality of 2-bromo-4,5-dimethoxybenzyl bromide, a series of studies were conducted on the detection of specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide. Summary of the invention

[0010] The object of the present invention is to provide a method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide, which uses an exclusive simple and accurate detection method and means to detect and control its specific impurities 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde to determine the product quality.

[0011] The technical solution adopted by the present invention is as follows:

[0012] A method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide, comprising dissolving a test sample and detecting the impurities by liquid chromatography;

[0013] The liquid chromatography method uses an octadecylsilane bonded silica gel chromatography column and uses an ultraviolet detector for detection;

[0014] The mobile phase is acetonitrile-water solution, wherein the volume content of acetonitrile is 70% and the volume content of water is 30%;

[0015] The column temperature of liquid chromatography is 25-40°C and the detection temperature is 25-40°C;

[0016] The flow rate of the mobile phase was 0.5-0.9 ml / min, and the elution mode was isocratic elution;

[0017] The injection volume for liquid chromatography is 10-30 μl.

[0018] The detection steps of the liquid chromatography method are:

[0019] 1) Set the liquid chromatograph according to the chromatographic conditions until the baseline is stable and wait for injection;

[0020] 2) Preparation of impurity solution: Take 10 mg each of 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde, accurately weigh them, place them in the same 100 ml volumetric flask, add acetonitrile to dilute to the scale, and shake well;

[0021] 3) Test solution: Take 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide, accurately weigh it, put it in a 100 ml volumetric flask, add an appropriate amount of acetonitrile to dissolve it, then dilute it to the scale and shake well;

[0022] According to the above chromatographic conditions, the sample was automatically injected, the peak area was recorded, and the content was calculated according to the area normalization method. The calculation formula is as follows:

[0023]

[0024] Where:

[0025] A 杂 It is the impurity peak area after deducting the blank in the test solution;

[0026] A 总 It is the sum of the peak areas of all chromatographic peaks in the test solution after deducting all blank peaks.

[0027] The detection conditions are as follows: the chromatographic column is a YMC-Pack ODS-AQ high performance liquid chromatography column with a specification of 25 cm×4.6 mm×5.0 μm.

[0028] In the present invention, the structural formulas of 2-bromo-4,5-dimethoxybenzyl bromide (1), 3,4-dimethoxybenzyl alcohol (2) and 2-bromo-4,5-dimethoxybenzaldehyde (3) are as follows:

[0029]

[0030] The concentration of the test solution is 0.6-1.0 mg / ml, and the concentration of the impurity localization solution is 0.08-0.12 mg / ml.

[0031] The present invention also includes the determination of the standard curve of 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde, and the determination step includes:

[0032] S01) accurately weighing 40 mg of each of 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde into a 10 ml volumetric flask, dissolving them with acetonitrile to the mark, accurately measuring 1 ml of the above solution into a 100 ml volumetric flask, dissolving them with acetonitrile to the mark, and preparing a single standard solution of 40 μg / ml;

[0033] S02) using a pipette to accurately pipette 1.0 ml, 2.0 ml, 4.0 ml, 5.0 ml, 6.0 ml, 8.0 ml, and 10.0 ml into a 50 ml volumetric flask, and then dilute to the mark with acetonitrile to obtain standard curve solutions of 0.8 μg / ml, 1.6 μg / ml, 3.2 μg / ml, 4.0 μg / ml, 4.8 μg / ml, 6.4 μg / ml, and 8.0 μg / ml;

[0034] S03) taking the standard curve solution for high performance liquid chromatography analysis, the chromatographic conditions are: using a YMC-Pack ODS-AQ column as a HPLC column, its specifications are 25cm×4.6mm×5.0μm, the mobile phase is acetonitrile-water at a volume ratio of 70:30, the flow rate is 0.7ml / min, and the column temperature is 30°C; the detector is a UV detector, and a standard curve is drawn according to the relationship between the concentration mg / ml and the chromatographic peak area.

[0035] Compared with the prior art, the invention has the following beneficial effects:

[0036] 1) Simplicity and accuracy: A simple and accurate HPLC detection method is provided to effectively detect specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide. It not only simplifies the operation process, but also improves the accuracy of detection, which helps to better control product quality.

[0037] 2) Good linear relationship: It shows good linear relationship in different concentration ranges (0.16mg / ml-1.60mg / ml), which means that the test results are more reliable and can obtain accurate quantitative analysis.

[0038] 3) Innovative detection method: In view of the problem that there is currently no accurate method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide, the present invention proposes an innovative solution. Through specific chromatographic conditions and detector settings, the detection method of the present invention can more effectively identify and quantify specific impurities.

[0039] 4) The instrument has low operating cost, simple and convenient detection process, good repeatability, and can be used to evaluate product quality. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 This is the chromatogram of the test solution of 2-bromo-4,5-dimethoxybenzyl bromide.

[0041] Figure 2 This is the chromatogram of the system suitability solution of 2-bromo-4,5-dimethoxybenzyl bromide.

[0042] Figure 3 This is the linear relationship diagram of 2-bromo-4,5-dimethoxybenzyl bromide.

[0043] Figure 4 This is the linear relationship diagram of 3,4-dimethoxybenzyl alcohol.

[0044] Figure 5 This is the linear relationship diagram of 2-bromo-4,5-dimethoxybenzaldehyde. DETAILED DESCRIPTION

[0045] The present invention is further described below in conjunction with embodiments and drawings, but the present invention is not limited to the following embodiments.

[0046] HPLC Detection of Specific Impurities in 2-Bromo-4,5-dimethoxybenzyl Bromide

[0047] The instruments and chemicals used in the examples were: high performance liquid chromatograph (Agilent 1260, Agilent); electronic balance (BS124S, Sartorius); YMC-Pack ODS-AQ chromatographic column (25 cm×4.6 mm×5.0 μm, YMC); acetonitrile was of chromatographic grade.

[0048] Example 1

[0049] HPLC analysis conditions:

[0050] Chromatographic column: YMC-Pack ODS-AQ chromatographic column (25cm×4.6mm×5.0μm, YMC)

[0051] The mobile phase was acetonitrile and aqueous solution;

[0052] The HPLC column temperature was 30°C;

[0053] The flow rate of the mobile phase was 0.7 ml / min, and the elution mode was isocratic;

[0054] The injection volume for HPLC was 20 μl.

[0055] Solution preparation:

[0056] Preparation of mobile phase: acetonitrile, aqueous solution: measure 700 ml of acetonitrile and 300 ml of water and mix well.

[0057] Impurity stock solution: Take 40 mg each of 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde, accurately weigh, place in the same 100 ml volumetric flask, add acetonitrile to dilute to the scale, and shake well.

[0058] System suitability solution: Take 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide into a 100 ml volumetric flask, then transfer 1.0 ml of the impurity stock solution into the same volumetric flask, add acetonitrile to dissolve and dilute to the mark.

[0059] Preparation of test solution: Weigh 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide test sample, accurately weigh it, place it in a 100 ml volumetric flask, add an appropriate amount of acetonitrile to dissolve it, then dilute it to the scale, shake well, and you will get a test solution with a concentration of 0.8 mg / ml.

[0060] Blank solution: acetonitrile.

[0061] Determination:

[0062] HPLC-VWD was used for detection, and the detection conditions were as described above.

[0063] Take blank and test solution and inject them into HPLC, record the chromatogram, the injection sequence is as follows:

[0064] name Number of injection needles blank ≥2, until the two needles completely overlap without interference Test solution 1

[0065] Example 2

[0066] 1. Exclusivity

[0067] 1.1 Solution preparation:

[0068] Blank: Acetonitrile

[0069] Impurity stock solution: Take 10 mg each of 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde, accurately weigh, place in the same 100 ml volumetric flask, add acetonitrile to dilute to the scale, and shake well.

[0070] Sensitivity solution: Take 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide into a 100 ml volumetric flask, then transfer 0.2 ml of the impurity stock solution into the same volumetric flask, add acetonitrile to dissolve and dilute to the scale.

[0071] Test solution: Weigh 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide, accurately weigh it, place it in a 100 ml volumetric flask, add an appropriate amount of acetonitrile to dissolve it, then dilute it to the scale, shake well, and you will get a test solution with a concentration of 0.8 mg / ml.

[0072] 1.2 Determination method

[0073] Inject blank, sensitivity solution and test solution in the following order and record the chromatogram.

[0074] Injection order:

[0075] name Number of injection needles blank ≥2, until the two needles completely overlap without interference Sensitivity solution 1 Test solution 1

[0076] 1.3 Results: No interference in blank; The peak separation of each component in the sensitivity solution is ≥1.5, and the signal-to-noise ratio (S / N) of specific impurities is greater than 10.

[0077]

[0078] 2. Linearity and range

[0079] 2.1 Solution preparation:

[0080] Stock solution: Accurately weigh 40 mg of each of 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde into a 10 ml volumetric flask, dissolve to volume with acetonitrile, accurately measure 1.0 ml of the above solution into a 100 ml volumetric flask, dissolve to volume with acetonitrile to prepare a single standard solution of 40 μg / ml;

[0081] Linear solution ① (0.8 μg / ml, 20%): Accurately measure 1.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0082] Linear solution ② (1.6 μg / ml, 40%): Accurately measure 2.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0083] Linear solution ③ (3.2 μg / ml, 80%): Accurately measure 4.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0084] Linear solution ④ (4.0 μg / ml, 100%): Accurately measure 5.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0085] Linear solution ⑤ (4.8 μg / ml, 120%): Accurately measure 6.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0086] Linear solution ⑥ (6.4 μg / ml, 160%): Accurately measure 8.0 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0087] Linear solution ⑦ (8.0 μg / ml, 200%): Accurately measure 10 ml of the stock solution, place it in a 50 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0088] 2.2 Determination method:

[0089] Take linear solution ① to linear solution ⑦, inject 2 times each, and record the chromatogram. Perform linear regression based on the concentration and peak area of ​​the linear solution to draw a standard curve.

[0090] 2.3 Results:

[0091] Table 1: 2-Bromo-4,5-dimethoxybenzyl bromide linear data table

[0092] Limit concentration ratio / % Concentration / μg / ml Peak area / mAU 20 0.80 16.74 40 1.60 34.31 80 3.20 66.89 100 4.01 81.82 120 4.81 101.94 160 6.41 134.49 200 8.01 167.68

[0093] Table 2: 3,4-Dimethoxybenzyl alcohol linear data table

[0094]

[0095]

[0096] Table 3: 2-Bromo-4,5-dimethoxybenzaldehyde linear data table

[0097] Limit concentration ratio / % Concentration / μg / ml Peak area / mAU 20 0.80 9.16 40 1.60 17.95 80 3.20 35.85 100 4.01 46.93 120 4.81 52.38 160 6.41 72.15 200 8.01 90.94

[0098] Linear regression was performed with concentration (μg / ml) as the horizontal axis and the corresponding peak area as the vertical axis. The linear results and linear equation are shown in Figure 3 , 4 5, the results show that 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde are detected according to the chromatographic conditions described in Example 1, and 2-bromo-4,5-dimethoxybenzyl bromide has a good linear relationship in the concentration range of 0.80 μg / ml to 8.01 μg / ml, and the linear equation is y=22.634x-0.1019, R 2 =0.9996; 3,4-dimethoxybenzyl alcohol has a good linear relationship in the concentration range of 0.80μg / ml to 8.01μg / ml, and the linear equation is y=22.634x-0.1019, R 2=0.9992; 2-bromo-4,5-dimethoxybenzaldehyde has a good linear relationship in the concentration range of 0.80μg / ml to 8.01μg / ml, and the linear equation is y=11.297x-0.055, R 2 =0.9986.

[0099] The above content is a further detailed description of the technical solution provided in combination with the preferred implementation mode of the present invention. It cannot be determined that the specific implementation of the present invention is limited to the above descriptions. For ordinary technicians in the technical field to which the present invention belongs, without departing from the concept of the present invention, some simple deductions or substitutions can be made, which should be considered as falling within the protection scope of the present invention as needed.

Claims

1. A method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide, characterized in that: The test sample was dissolved and tested by liquid chromatography; The liquid chromatography method uses an octadecylsilane bonded silica gel chromatography column and uses an ultraviolet detector for detection; The mobile phase is acetonitrile-water solution, wherein the volume content of acetonitrile is 70% and the volume content of water is 30%; The column temperature of liquid chromatography is 25-40°C and the detection temperature is 25-40°C; The flow rate of the mobile phase was 0.5-0.9 ml / min, and the elution mode was isocratic elution; The injection volume for liquid chromatography is 10-30 μl. The detection steps of the liquid chromatography method are: 1) Set the liquid chromatograph according to the chromatographic conditions until the baseline is stable and wait for injection; 2) Preparation of impurity solution: Take 10 mg each of 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde, accurately weigh them, place them in the same 100 ml volumetric flask, add acetonitrile to dilute to the scale, and shake well; 3) Test solution: Take 80 mg of 2-bromo-4,5-dimethoxybenzyl bromide, accurately weigh it, put it in a 100 ml volumetric flask, add an appropriate amount of acetonitrile to dissolve it, then dilute it to the scale and shake well; According to the above chromatographic conditions, the sample was automatically injected, the peak area was recorded, and the content was calculated according to the area normalization method. The calculation formula is as follows: Where: A 杂 It is the impurity peak area after deducting the blank in the test solution; A 总 It is the sum of the peak areas of all chromatographic peaks in the test solution after deducting all blank peaks.

2. The method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide according to claim 1, characterized in that: The detection conditions are as follows: the chromatographic column is a YMC-Pack ODS-AQ high performance liquid chromatography column with a specification of 25 cm×4.6 mm×5.0 μm.

3. The method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide according to claim 1, characterized in that: The concentration of the test solution is 0.6-1.0 mg / ml, and the concentration of the impurity localization solution is 0.08-0.12 mg / ml.

4. The method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide according to claim 1, characterized in that: It also includes the determination of standard curves for 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol, and 2-bromo-4,5-dimethoxybenzaldehyde.

5. The method for detecting specific impurities in 2-bromo-4,5-dimethoxybenzyl bromide according to claim 4, characterized in that: The steps for determining the standard curve of the 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde include: S01) accurately weighing 40 mg of each of 2-bromo-4,5-dimethoxybenzyl bromide, 3,4-dimethoxybenzyl alcohol and 2-bromo-4,5-dimethoxybenzaldehyde into a 10 ml volumetric flask, dissolving them with acetonitrile to the mark, accurately measuring 1 ml of the above solution into a 100 ml volumetric flask, dissolving them with acetonitrile to the mark, and preparing a single standard solution of 40 μg / ml; S02) using a pipette to accurately pipette 1.0 ml, 2.0 ml, 4.0 ml, 5.0 ml, 6.0 ml, 8.0 ml, and 10.0 ml into a 50 ml volumetric flask, and then dilute to the mark with acetonitrile to obtain standard curve solutions of 0.8 μg / ml, 1.6 μg / ml, 3.2 μg / ml, 4.0 μg / ml, 4.8 μg / ml, 6.4 μg / ml, and 8.0 μg / ml; S03) taking the standard curve solution for high performance liquid chromatography analysis, the chromatographic conditions are: using a YMC-Pack ODS-AQ column as a HPLC column, its specifications are 25cm×4.6mm×5.0μm, the mobile phase is acetonitrile-water at a volume ratio of 70:30, the flow rate is 0.7ml / min, and the column temperature is 30°C; the detector is a UV detector, and a standard curve is drawn according to the relationship between the concentration mg / ml and the chromatographic peak area.

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