Gas chromatography detection method for related substances of 2, 5-dimethoxytetrahydrofuran

Abstract

The invention solves the problems in the prior art, and provides a gas chromatography detection method for related substances of 2, 5-dimethoxytetrahydrofuran, which is simple and convenient to operate, good in specificity, linearity and precision, high in sensitivity and good in durability, can be used for quality control of a compound (Va), namely 2, 5-dimethoxytetrahydrofuran, and meets process requirements.

Description

Technical Field

[0001] The invention belongs to the technical field of drug analysis, and in particular relates to a gas chromatography method for separating and detecting 2,5-dimethoxytetrahydrofuran-related substances. Background Art

[0002] The chemical name of ilaprazole is 2-[(RS)-[(4-methoxy-3-methylpyridin-2-yl)methyl]sulfinyl]-5-(1H-pyrrol-1-yl)-1H-benzimidazole (CAS: 172152-36-2). Ilaprazole is a potent proton pump inhibitor that works by interfering with the proton pump that secretes gastric acid on gastric parietal cells. Its effect in inhibiting gastric acid is the strongest among similar drugs. It is mainly used for adult duodenal ulcers, gastric ulcers and reflux esophagitis.

[0003] Compound (Va), 2,5-dimethoxytetrahydrofuran (CAS No.: 696-59-3), is a key starting material for the preparation of ilaprazole API, and its structural formula is as follows:

[0004]

[0005] Compound (Va) is usually obtained by catalytic hydrogenation of 2,5-dimethoxydihydrofuran (CAS No.: 332-77-4) and is composed of a mixture of cis and trans isomers. According to the chemical structure and preparation process of compound (Va), it may contain process impurities and degradation impurities, which will directly participate in the reaction during the synthesis process to produce process by-product impurities corresponding to ilaprazole. It is difficult to remove them during the synthesis process, which may affect the quality of the final product ilaprazole. Compound (Va) and its related impurities are low-boiling substances without ultraviolet absorption. Therefore, it is very necessary to establish a gas phase detection method for related substances of compound (Va).

[0006] Currently, the enterprise standard of Laohekou Guanglian Technology Co., Ltd. (standard number: Q / 420682GL02-2023) is retrieved: 2,5-dimethoxytetrahydrofuran quality standard, in which the method for determining the content of 2,5-dimethoxytetrahydrofuran is gas chromatography, and the area normalization method is used to determine the content of its main component. This method has a short retention time for the main peak and impurity peaks, and can only roughly examine the content of the main component of 2,5-dimethoxytetrahydrofuran. Other impurities merge after the main peak and have poor separation from the main peak, making it impossible to achieve quality control of ilaprazole. Summary of the invention

[0007] The present invention solves the problems existing in the prior art and provides a gas chromatography detection method for 2,5-dimethoxytetrahydrofuran-related substances. The method is simple to operate, has good specificity, linearity and precision, high sensitivity and good method durability, and can be used for quality control of ilaprazole starting material compound (Va), namely 2,5-dimethoxytetrahydrofuran, to meet process requirements.

[0008] The invention provides a gas chromatography detection method for 2,5-dimethoxytetrahydrofuran related substances, comprising the following steps: respectively sampling a separation solution and a test solution, performing detection according to gas chromatography conditions, recording a chromatogram, and calculating the detection amount of a single impurity and total impurities according to a peak area normalization method;

[0009] The structures of the related substances are shown below:

[0010] ;

[0011] Among them, impurity Va-1 is the starting material of compound (Va), impurity Va-2, impurity Va-3, and impurity Va-4 are process impurities of compound (Va), and butanedial and methanol are degradation impurities of compound (Va).

[0012] The chromatographic conditions are:

[0013] Instrument: Agilent gas chromatograph, flame ionization detector (FID);

[0014] Chromatographic column: A capillary column with 6% cyanopropylphenyl-94% dimethylpolysiloxane as the stationary liquid;

[0015] Column temperature: starting temperature is 35-70°C, maintained for 0-5 minutes, then heated to 200-240°C at a rate of 10-20°C / min, maintained for 10-20 minutes; preferably 55-65°C, maintained for 2 minutes, then heated to 240°C at a rate of 12-18°C / min, maintained for 12 minutes;

[0016] Inlet temperature: 150-250°C, preferably 200°C;

[0017] Carrier gas: nitrogen;

[0018] Carrier gas flow rate: 0.9~1.1ml / min;

[0019] Detector temperature: 200-300°C, preferably 250°C;

[0020] Split ratio: 50:1-150:1, preferably 100:1;

[0021] Injection volume: 1μl.

[0022] The chromatographic column is DB-624, with an inner diameter of 0.32 mm, a length of 60 m, and a film thickness of 1.8 μm.

[0023] The test solution is prepared as follows: take an appropriate amount of compound (Va), dissolve it in a solvent and dilute it to make a solution containing 0.1 g per 1 ml.

[0024] The preparation method of the separation solution is as follows: take appropriate amounts of methanol, impurity Va-2 reference substance, succinaldehyde reference substance, impurity Va-3 reference substance, impurity Va-1 reference substance and impurity Va-4 reference substance, place them in the same volumetric bottle, dilute to the mark with solvent, shake well, and use the solution as the impurity stock solution. Take appropriate amount of compound (Va) and place it in a volumetric bottle, add appropriate amount of impurity stock solution, dilute to the mark with solvent, and shake well.

[0025] The solvent is selected from one or more of acetonitrile, ethanol, acetone and dichloromethane, preferably acetonitrile.

[0026] The invention provides a gas chromatography detection method for 2,5-dimethoxytetrahydrofuran related substances, comprising the following steps: respectively sampling a separation solution and a test solution, performing detection according to gas chromatography conditions, recording a chromatogram, and calculating the detection amount of a single impurity and total impurities according to a peak area normalization method;

[0027] The structures of the related substances are shown below:

[0028] ;

[0029] The chromatographic conditions are:

[0030] Instrument: Agilent gas chromatograph, flame ionization detector (FID);

[0031] Chromatographic column: A capillary column with 6% cyanopropylphenyl-94% dimethylpolysiloxane as the stationary liquid;

[0032] Column temperature: start at 60°C, maintain for 2 minutes, increase to 240°C at a rate of 15°C per minute, and maintain for 12 minutes;

[0033] Inlet temperature: 200°C;

[0034] Carrier gas: nitrogen;

[0035] Carrier gas flow rate: 1.0 ml / min;

[0036] Detector temperature: 250°C;

[0037] Split ratio: 100:1;

[0038] Injection volume: 1μl.

[0039] The chromatographic column is DB-624, with an inner diameter of 0.32 mm, a length of 60 m, and a film thickness of 1.8 μm.

[0040] The test solution was prepared as follows: 1.0 g of compound (Va) was placed in a 10 ml volumetric flask, acetonitrile was added to dissolve to the mark, and shaken well.

[0041] The preparation method of the separation solution is as follows: 25 mg each of methanol, impurity Va-2 reference substance, succinaldehyde reference substance, impurity Va-3 reference substance, impurity Va-1 reference substance and impurity Va-4 reference substance are placed in the same 5 ml volumetric bottle, diluted to the mark with acetonitrile, and shaken to obtain the impurity stock solution. 0.2 g of compound (Va) is placed in a 2 ml volumetric bottle, 0.2 ml of the impurity stock solution is added, diluted to the mark with acetonitrile, and shaken to obtain the solution.

[0042] The present invention further provides a method for detecting 2,5-dimethoxytetrahydrofuran-related substances by gas chromatography and its use in the quality control of ilaprazole.

[0043] The beneficial technical effects of the present invention are:

[0044] (1) The present invention provides a method for detecting related substances of compound (Va) i.e. 2,5-dimethoxytetrahydrofuran, a starting material of ilaprazole. The method can effectively separate the two components of compound (Va) i.e. 2,5-dimethoxytetrahydrofuran (a mixture of cis and trans isomers) and various impurities.

[0045] (2) The detection method of the present invention is easy to operate, has good specificity, linearity, precision, high sensitivity, and good method durability. It can be used for quality control of the starting material compound (Va) of ilaprazole, namely 2,5-dimethoxytetrahydrofuran, and meets the process requirements. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 This is the chromatogram of condition ① of Example 1.

[0047] Figure 2 This is the chromatogram of condition ② of Example 1.

[0048] Figure 3 This is the chromatogram of condition ③ of Example 1.

[0049] Figure 4 This is the separation solution chromatogram of Example 2.

[0050] Figure 5 This is the separation solution chromatogram of Example 2.

[0051] Figure 6 This is the linear test diagram of Example 3.

[0052] Figure 7 This is the chromatogram of the quantitative limit of compound (Va) in Example 4.

[0053] Figure 8 This is the quantification limit chromatogram of impurity Va-3 in Example 4.

[0054] Fig. 9 The chromatograms of the quantitative limits of other impurities in Example 4 are shown.

[0055] Fig.10 This is the detection limit chromatogram of compound (Va) in Example 4.

[0056] Fig.11 This is the detection limit diagram of impurity Va-3 in Example 4.

[0057] Fig.12 This is the detection limit diagram of other impurities in Example 4.

[0058] Fig.13 This is the precision test spectrum of Example 5.

[0059] Fig.14 This is the stability test spectrum of Example 6.

[0060] Fig.15 The chromatograms of the chromatographic columns with different durability of chromatographic conditions in Example 7 are shown.

[0061] Fig.16 The chromatograms of different carrier gas flow rates for the durability of the chromatographic conditions in Example 7 are shown.

[0062] Fig.17 The chromatograms of different column temperatures for the durability of the chromatographic conditions in Example 7 are shown. DETAILED DESCRIPTION

[0063] 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.

[0064] Solution preparation:

[0065] Preparation of test solution: Take an appropriate amount of compound (Va), dissolve it in acetonitrile and dilute it to make a solution containing 0.1 g per 1 ml.

[0066] Preparation of separation solution: Take 25 mg each of methanol, impurity Va-2 reference substance, succinaldehyde reference substance, impurity Va-3 reference substance, impurity Va-1 reference substance and impurity Va-4 reference substance, place them in the same 5 ml volumetric flask, dilute to scale with acetonitrile, shake well, and use it as impurity stock solution. Take 0.2 g of compound (Va), place it in a 2 ml volumetric flask, add 0.2 ml of impurity stock solution, dilute to scale with acetonitrile, shake well, and obtain.

[0067] Example 1 Selection of Chromatographic Conditions

[0068] ① Refer to the corporate standard of Laohekou Guanglian Technology Co., Ltd. (standard number: Q / 420682GL02-2023): Chromatographic conditions in the quality standard of 2,5-dimethoxytetrahydrofuran:

[0069] Instrument: Agilent gas chromatograph, flame ionization detector (FID);

[0070] Chromatographic column: A capillary column with 5% phenyl-95% dimethylpolysiloxane as the stationary phase was used as the chromatographic column (HP-5, 0.32 mm × 60 m, 1.0 μm);

[0071] Column temperature: 150°C, maintained for 30 min;

[0072] Carrier gas: nitrogen;

[0073] Carrier gas flow rate: 1.0 ml / min;

[0074] Inlet temperature: 200°C;

[0075] Detector temperature: 250°C;

[0076] Split ratio: 100:1;

[0077] Injection volume: 1µl.

[0078] Preparation of separation solution: Take 25 mg of impurity Va-1 reference substance and impurity Va-4 reference substance, place them in the same 5 ml volumetric flask, dilute to scale with acetonitrile, shake well, and use as impurity stock solution. Take 1.0 g of compound (Va), place it in a 10 ml volumetric flask, add 1 ml of impurity stock solution, dilute to scale with acetonitrile, shake well, and obtain.

[0079] Accurately measure 1 μl of the separation solution, inject it into the gas chromatograph, and record the chromatogram.

[0080] Results: The retention time of the main peak in the chromatogram of the separated solution was 1.990 minutes. The number of impurity peaks was small and inconsistent with the number of impurities added. It should be that impurities Va-1, Va-4 and unknown peaks merged into one peak. The resolution between the main peak and the adjacent impurity peak was 1.38, which was less than 1.5. The resolution was poor. Figure 1 shown.

[0081] ②Change the temperature program to: start at 50℃, maintain for 5 minutes, increase the temperature to 240℃ at a rate of 15℃ per minute, and maintain for 8 minutes. The other conditions are the same as condition ①.

[0082] Accurately measure 1 μl of the separation solution, inject it into the gas chromatograph, and record the chromatogram.

[0083] Results: In the chromatogram of the separated solution, the resolution between the impurity Va-1 and impurity Va-4 peaks was 1.42, less than 1.5, and the resolution was poor. The tailing factor of the main peak of compound (Va) was 0.65, less than 1.2, and the peak shape was poor. Figure 2 shown.

[0084] ③Chromatographic column: a capillary column with 6% cyanopropylphenyl-94% dimethylpolysiloxane as the stationary liquid (model DB-624, inner diameter 0.32mm, length 60m, film thickness 1.8μm);

[0085] Column temperature: start at 60°C, maintain for 2 minutes, increase to 240°C at a rate of 15°C per minute, and maintain for 12 minutes;

[0086] The rest of the conditions are the same as condition ①.

[0087] Accurately measure 1 μl of the separation solution, inject it into the gas chromatograph, and record the chromatogram.

[0088] Results: There are two main peaks in the chromatogram of the test solution (cis-trans isomers, with two main peaks), the minimum theoretical plate number of the main peak is 818204, greater than 5000, the peak shape is good, the minimum resolution between the main peak and the adjacent peak is 4.58, greater than 1.5, and the separation is good. Figure 3 shown.

[0089] Example 2 Separation Test

[0090] The separation test was continued using the chromatographic conditions screened in Condition ③ of Example 1.

[0091] Preparation of impurity locating solution: Take 20 mg each of methanol, impurity Va-2 reference substance, succinaldehyde reference substance, impurity Va-3 reference substance, impurity Va-1 reference substance and impurity Va-4 reference substance, place them in 6 10 ml volumetric flasks, add acetonitrile to dissolve and dilute to the scale, shake well, and use them as the locating solutions for each impurity.

[0092] Accurately measure 1.0 µl of the test solution, positioning solution and separation solution, respectively, and inject them into the gas chromatograph to record the chromatogram. The results are shown in Table 1. Figure 4 , Figure 5 .

[0093] The results showed that: in the separation solution chromatogram, butanedialdehyde and impurity Va-3 co-existed, and the two impurities were combined and controlled in this method; the minimum separation between the two main peaks and the adjacent impurity peaks was 4.56, and the minimum separation between the impurity peaks was 5.10, both greater than 1.5, indicating good separation; in the chromatogram of the test solution, the tailing factor of the main peak was 1.04, less than 2.0, indicating that the method had good specificity.

[0094] Table 1 Positioning and separation test results

[0095]

[0096] Example 3 Linearity test

[0097] Preparation of linear solution: take 1.2 g of compound (Va), accurately weigh it, place it in a 10 ml volumetric flask, add acetonitrile to dissolve and dilute to the scale, shake well, and use it as linear solution 1; take another 1.0 g of compound (Va), accurately weigh it, place it in a 10 ml volumetric flask, add acetonitrile to dissolve and dilute to the scale, shake well, accurately measure 5 ml, 1 ml, and 0.1 ml of the solution, place them in three 10 ml volumetric flasks respectively, dilute to the scale with acetonitrile, shake well, and use them as linear solution 2, linear solution 3, and linear solution 4 respectively; accurately measure 0.1 ml of linear solution 3, place it in a 10 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and use them as linear solution 5.

[0098] Preparation of quantitative limit solution: Take 1.0019g of compound (Va), accurately weigh, place in a 10ml volumetric flask, add acetonitrile to dissolve and dilute to the scale, shake well, accurately measure 0.1ml, place in a 10ml volumetric flask, dilute to the scale with acetonitrile, shake well, measure 0.3ml of the sensitivity stock solution of compound (Va), place in a 10ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0099] Accurately measure 1 μl of the linear solution and the quantitative limit solution, respectively, and inject them into the gas chromatograph. Record the chromatogram, plot the peak area against the concentration, and calculate the regression equation and correlation coefficient using the least squares method. The results are shown in Table 2. Figure 6 .

[0100] Table 2 Results of linearity test for related substances of compound (Va)

[0101]

[0102] Example 4 Limit of Quantitation and Limit of Detection

[0103] Preparation of sensitivity stock solution of compound (Va): Take 1.0019 g of compound (Va), accurately weigh it, place it in a 10 ml volumetric flask, add acetonitrile to dissolve and dilute to the scale, shake well, accurately measure 0.1 ml, place it in a 10 ml volumetric flask, dilute to the scale with acetonitrile, and shake well.

[0104] Preparation of the quantitative limit solution of compound (Va): Measure 0.3 ml of the sensitive stock solution of compound (Va), place it in a 10 ml volumetric flask, dilute to the scale with acetonitrile, shake well, and obtain.

[0105] Preparation of the detection limit solution of compound (Va): Measure 0.1 ml of the sensitivity stock solution of compound (Va), place it in a 10 ml volumetric flask, dilute to the scale with acetonitrile, and shake well to obtain the solution.

[0106] Preparation of impurity stock solution: Take 41.8 mg of methanol, 38.58 mg of impurity Va-2 reference substance, 101.63 mg of succinaldehyde reference substance, 29.63 mg of impurity Va-3 reference substance, 39.98 mg of impurity Va-1 reference substance and 40.97 mg of impurity Va-4 reference substance, place them in the same 5 ml volumetric flask, add acetonitrile solvent and dilute to the scale, shake well, accurately measure 1 ml, place it in a 10 ml volumetric flask, dilute to the scale with acetonitrile, and shake well.

[0107] The impurity stock solution was diluted to different concentrations with acetonitrile step by step as the quantitative limit and detection limit solutions of each impurity. 1µl was accurately measured and injected into the gas chromatograph to record the chromatogram. The measured response value was greater than or equal to 10 times and 3 times the noise signal, which was the quantitative limit and detection limit of the corresponding compound. The results are shown in Table 3, Table 4 and Figure 7 to Figure 12 .

[0108] Table 3 Quantitative limit results of related substances of compound (Va)

[0109]

[0110] Table 4 Detection limit results of related substances of compound (Va)

[0111]

[0112] Example 5 Precision Test

[0113] Prepare 6 parallel portions of the test solution. Accurately measure 1 μl of each, inject into the gas chromatograph, record the chromatogram, calculate by area normalization method, analyze and compare the detection amount of the largest single impurity and total impurities. The results are shown in Table 5. Fig.13 The maximum single impurity and total impurities detected were consistent, and the precision test results of the related substances of compound (Va) were good.

[0114] Table 5 Results of precision test of related substances of compound (Va)

[0115]

[0116] Example 6 Stability Test

[0117] Place the test solution at room temperature under natural light. Accurately measure 1 μl at 0, 2, 4, 6, 8, and 24 hours, inject it into the gas chromatograph, record the chromatogram, analyze and compare the detection amount of the largest single impurity and the total impurities. The results are shown in Table 6. Fig.14 The maximum change rate of the largest single impurity detected was 5.26%, which was less than 20%. The total impurity detected was consistent, and no new impurities were generated at each time point, indicating that the test solution was stable within 24 hours.

[0118] Table 6 Results of stability test of solution of related substances of compound (Va)

[0119]

[0120] Example 7 Durability Test

[0121] On the basis of standard chromatographic conditions, different chromatographic columns, different carrier gas flow rates, and different column temperature conditions are used to determine the related substances of this product according to Table 7. Accurately measure 1 μl of the test solution, inject it into the gas chromatograph, record the chromatogram, and compare the determination results.

[0122] Accurately measure 1 μl of the test solution, inject it into the gas chromatograph, and record the chromatogram. The results are shown in Table 8. Figures 15 to 17 .

[0123] The results showed that when using chromatographic columns of the same model but different numbers, the carrier gas flow rate was in the range of 0.9~1.1ml / min, the starting temperature was in the range of 55~65℃ (heating rate was in the range of 12~18℃ / min), and the system suitability results were good; compared with standard conditions, the maximum change in the detection amount of the maximum single impurity and the total impurities was 0.01%, indicating that the detection method for related substances of compound (Va) has good durability.

[0124] Table 7 Durability of the method for detecting related substances of compound (Va)

[0125]

[0126] Table 8 Results of durability test for related substances of compound (Va)

[0127]

[0128] Note: ① The test product has cis and trans isomers and has two main peaks. The theoretical plate number of the main peak is calculated based on the minimum value, and the tailing factor of the main peak is calculated based on the maximum value; ②* is the standard condition.

Claims

1. A gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran, characterized in that: The method comprises the following steps: sampling the separation solution and the test solution respectively, testing according to gas chromatography conditions, recording the chromatogram and calculating the detected amount of impurities; The chromatographic conditions are as follows: chromatographic column: a capillary column with 6% cyanopropylphenyl-94% dimethylpolysiloxane as a stationary liquid; a heating program: a starting temperature of 35-70°C, maintained for 0-5 minutes, and heated to 200-240°C at a rate of 10-20°C / min, and maintained for 10-20 minutes; an injection port temperature of 150-250°C; a detector temperature of 200-300°C; a split ratio of 50:1-150:1; The structures of the related substances are shown below: 。 2. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The chromatographic conditions also include: Detector: Hydrogen flame ionization detector; Carrier gas: nitrogen; Carrier gas flow rate: 1.0 ml / min; Injection volume: 1μl.

3. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The chromatographic column is DB-624, 0.32 mm×60 m, 1.8 μm.

4. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The heating program is as follows: the starting temperature is 55-65°C, maintained for 2 minutes, and then heated to 240°C at a rate of 12-18°C / min, and maintained for 12 minutes.

5. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The injection port temperature is 200°C.

6. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The detector temperature was 250°C.

7. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The split ratio is 100:

1.

8. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The preparation method of the test solution is as follows: take an appropriate amount of 2,5-dimethoxytetrahydrofuran, add a solvent to dissolve and dilute to prepare a solution containing 0.1 g per 1 ml; the preparation method of the separation solution is as follows: take an appropriate amount of methanol, impurity Va-2 reference substance, succinaldehyde reference substance, impurity Va-3 reference substance, impurity Va-1 reference substance and impurity Va-4 reference substance, place them in the same volumetric bottle, dilute to the scale with a solvent, shake well, and use it as an impurity stock solution; take an appropriate amount of 2,5-dimethoxytetrahydrofuran and place it in a volumetric bottle, add an appropriate amount of impurity stock solution, dilute to the scale with a solvent, and shake well to obtain.

9. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 8, characterized in that: The agent is selected from one or more of acetonitrile, ethanol, acetone and dichloromethane.

10. The gas chromatography detection method for related substances of 2,5-dimethoxytetrahydrofuran according to claim 1, characterized in that: The impurity detection amount is calculated according to the peak area normalization method.

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