CN111721879A - Method for detecting D-3-acetylmercapto-2-methylpropanoyl chloride in captopril by gas chromatography-mass spectrometry - Google Patents
Method for detecting D-3-acetylmercapto-2-methylpropanoyl chloride in captopril by gas chromatography-mass spectrometry Download PDFInfo
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Abstract
The invention discloses a method for detecting genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril raw material medicine, belonging to the field of medicine analysis. The gas chromatography conditions were: the chromatographic column is HP-5, 30m × 0.25mm × 0.25 μm, UI; the stationary phase is 5% of phenyl-95% of dimethyl polysiloxane, and the carrier gas is helium; temperature programming; the flow rate is 1.4-1.6 mL/min; the split ratio is 10: 1; the amount of sample was 1.0. mu.L. The mass spectrum conditions are as follows: ion source (EI) voltage, ionization energy: 70eV, ion source temperature: 230 ℃, quadrupole temperature: 150 ℃, transmission line temperature: 280 ℃. The method has high accuracy and strong specificity, can qualitatively or quantitatively detect genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril raw material medicines, ensures the quality of captopril and improves the safety of clinical medication.
Description
Technical Field
The invention belongs to the field of pharmaceutical analysis, and particularly relates to a method for detecting genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril.
Background
The genotoxic impurities refer to substances which can directly or indirectly damage DNA, cause genetic material damage of human bodies at low concentration to cause gene mutation or have carcinogenic tendency, and seriously threaten human health in the medication process. In recent years, genotoxic impurities gradually attract wide attention due to important safety risks, and the development of accurate identification and monitoring methods for genotoxic impurities of medicines has a very important significance in order to avoid passive intake of genotoxic impurities and guarantee the quality of medicines and the medication safety of the public. Regulatory agencies in various countries, such as EMEA, FDA, ICH, etc., issue guidelines for genotoxic impurities in succession, and require control and evaluation of such impurities.
Captopril is the earliest found angiotensin converting enzyme inhibitor, has good curative effect, does not produce orthostatic hypotension, improves cardiac function, increases renal blood flow, and is one of the first-choice medicaments for treating hypertension and congestive heart failure. The captopril may have potential genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in the synthesis process, and the measurement method is not reported at present. Because genotoxic impurity detection has special requirements on the aspects of sensitivity, selectivity, stability of an object to be detected, complexity of a matrix and the like, and the analysis method has the characteristic of being different from other impurity detection requirements, the existing analysis technology of genotoxic impurities of drugs is still very incomplete, so that the genotoxic impurities are not completely judged and mastered by enterprises, the research and development cost is high, the problems of 'sending and supplementing' required by drug registration approval analysis and the like are caused, and the research and development speed, the quality of products and the international competitiveness are influenced. Therefore, it is necessary to establish a method for accurately and rapidly quantitatively detecting and analyzing the genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in the medicine captopril.
Disclosure of Invention
The invention aims to solve the technical problem of providing a method for accurately detecting genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril, which has high accuracy, strong specificity and good reproducibility, wherein unstable impurity D-3-acetylmercapto-2-methylpropanoyl chloride is esterified and then detected by adopting a gas chromatography-mass spectrometry combined technology, and the analysis method has the advantages of high analysis speed and high sensitivity, thereby ensuring the quality of captopril raw material medicines and improving the safety of clinical medication.
The invention is realized by the following technical scheme:
a method for detecting genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril bulk drug, which uses a gas chromatography-mass spectrometry method for detection, comprises the following steps:
(1) preparing a test solution: precisely weighing a sample of the nicarduopril, dissolving the sample by using an alcohol solvent, fixing the volume, shaking the sample uniformly to prepare a solution of 0.05-0.20 g/mL, filtering the solution by using a microporous filter membrane, taking a subsequent filtrate as a sample solution, and preparing a blank solution.
(2) Preparing a reference substance solution: precisely weighing a D-3-acetylmercapto-2-methylpropanoyl chloride reference substance, placing the reference substance in a 25mL volumetric flask, dissolving the reference substance by using an alcohol solvent, fixing the volume to a scale, and shaking up to obtain a reference substance solution a; precisely measuring 50 mu L of reference substance solution a, placing in a 2mL sample injection vial, adding 950 mu L of alcohol solvent, and shaking up to obtain reference substance solution b; precisely measuring 50 μ L of the control solution b, placing in a 2mL injection vial, adding 950 μ L of alcohol solvent, and shaking to obtain control solution c. Precisely measuring 100 μ L of the control solution, placing in a 2mL sample injection vial, adding 900 μ L of alcohol solvent, and shaking to obtain the control solution.
Further, the concentration of the test solution is 0.10 g/mL.
Furthermore, the concentration of the reference substance solution is 0.20-0.65 mug/mL.
Further, the diluting solvent used for the test solution and the reference solution is an alcohol solvent such as methanol, ethanol and isopropanol.
(3) Precisely measuring blank solution, reference solution and sample solution 1.0 μ L, respectively, injecting into gas chromatography-mass spectrometer for testing, and recording chromatogram. The gas chromatography-mass spectrometry detection method has the following conditions:
gas chromatography conditions: the chromatographic column is HP-5, 30m × 0.25mm × 0.25 μm, UI; the stationary phase is 5% of phenyl-95% of dimethyl polysiloxane, and the carrier gas is helium; the temperature of a sample inlet is 200-240 ℃; an MS detector; the flow rate is 1.4-1.6 mL/min; the split ratio is 10: 1; the amount of sample was 1.0. mu.L.
Mass spectrum conditions: ion source (EI) voltage, ionization energy: 70eV, ion source temperature: 230 ℃, quadrupole temperature: 150 ℃, transmission line temperature: 280 ℃, scanning mode: selective ion scan, quantification of ions: 144, qualitative ion: 144. 134, 116, 101.
Further, the gas chromatographic column temperature condition is as follows: the initial column temperature is 60-75 ℃, the initial column temperature is maintained for 1min, then the temperature is increased to 180 ℃ at the speed of 20 ℃/min, the temperature is increased to 260 ℃ at the speed of 40 ℃/min, and the initial column temperature is maintained for 10min at 260 ℃.
(4) And (3) calculating: and calculating the content of the D-3-acetylthio-2-methylpropanoyl chloride in the test sample by peak area according to an external standard method.
Has the advantages that:
the method for determining the genotoxic impurity D-3-acetylmercapto-2-methylpropionyl chloride in captopril provided by the invention has the advantages of quick and simple operation, high sensitivity, good repeatability and accurate result, can be used for qualitatively or quantitatively detecting, provides a good reference for controlling the D-3-acetylmercapto-2-methylpropionyl chloride residue in the production process of captopril serving as a raw material medicine, ensures the quality of terbutaline sulfate raw material medicines, and thus improves the safety of clinical medication.
Drawings
FIG. 1A diagram of the detection limit determination by the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 2D-3-acetylmercapto-2-methylpropanoyl chloride method validation quantitation limit determination profile;
FIG. 3D-3-acetylmercapto-2-methylpropanoyl chloride method verifies the profile of the specificity determination control;
FIG. 4 is a diagram of a test sample for specific assay, which is verified by the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 5 is a graph of a control sample for limiting recovery determination using the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 6A-3-acetylmercapto-2-methylpropanoyl chloride method validation limit recovery determination solvent profile;
FIG. 7A diagram of a blank sample for the validation of the limit recovery assay by the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 8 is a 100% plot of a validation limit recovery assay using the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 9 is a graph of the control after the verification of the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 10 is a solvent profile after validation of the D-3-acetylmercapto-2-methylpropanoyl chloride method;
FIG. 11 is a test sample spectrum after the verification of the D-3-acetylmercapto-2-methylpropanoyl chloride method.
Detailed Description
The following examples are provided to further illustrate the technical solutions of the present invention, but not to limit the present invention.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of example embodiments according to the present application. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise.
First, chromatogram/mass spectrum condition and system applicability test
According to the test of gas chromatography (0521 of the general rules of the four ministry of the Chinese pharmacopoeia 2015 edition), an analytical column (HP-5, 30m × 0.25mm × 0.25 μm, UI) with a stationary liquid of 5% phenyl-95% dimethylpolysiloxane is adopted, the initial column temperature is 70 ℃, the initial column temperature is maintained for 1 minute, the temperature is increased to 180 ℃ at the rate of 20 ℃ per minute, the temperature is increased to 260 ℃ at the rate of 40 ℃ per minute, the initial column temperature is maintained for 10 minutes, the injection port temperature is 220 ℃, an MS detector takes helium as a carrier gas, the flow rate is 1.5mL per minute, the split ratio is 10:1, and the sample injection amount is 1.0 μ L.
Mass spectrum conditions: ion source (EI) voltage, ionization energy: 70eV, ion source temperature: 230 ℃, quadrupole temperature: 150 ℃, transmission line temperature: 280 ℃, scanning mode: selective ion scan, quantification of ions: 144, qualitative ion: 144. 134, 116, 101.
The determination method comprises the following steps: taking 1.0g of a captopril sample, precisely weighing, dissolving with methanol, fixing the volume to a scale, shaking up, passing through a microporous filter membrane (the aperture of the filter membrane is 0.45 mu m), taking a subsequent filtrate as a test solution, and simultaneously performing a blank test; taking a proper amount of D-3-acetylmercapto-2-methylpropanoyl chloride reference substance, precisely weighing, dissolving with methanol, and quantitatively diluting to obtain a solution containing about 0.33 μ g of D-3-acetylmercapto-2-methylpropanoyl chloride per 1mL of D-acetylmercapto-2-methylpropanoyl chloride reference substance as a reference substance solution. Precisely measuring blank solution, reference solution and sample solution 1.0 μ L, respectively, injecting into gas chromatography-mass spectrometer, and recording chromatogram. If a chromatographic peak with the same retention time as the main peak of the reference solution exists in the chromatogram of the test solution, the mass-to-charge ratio of the corresponding qualitative ions is consistent with that of the reference solution, the abundance ratio accords with the range specified in table 1 compared with that of the reference solution, and the D-3-acetylthio-2-methylpropanoyl chloride content is not more than 3.33 mug/mL calculated by the external standard method based on the peak area of the quantitative ions.
TABLE 1 maximum permissible deviation of relative ion abundance for qualitative confirmation
| Relative ion abundance (%) | ＞50 | ＞20～50 | ＞10～20 | ≤10 |
| Maximum deviation allowed (%) | ±20 | ±25 | ±30 | ±50 |
Verification of analysis method of di, D-3-acetylmercapto-2-methylpropanoyl chloride
The instrument comprises the following steps: agilent 7890B-5977B gas chromatography-mass spectrometer; a chromatographic column: HP-5(30 m.times.0.25 mm.times.0.25 μm, UI); reagent: methanol (chromatographic purity), D-3-acetylmercapto-2-methylpropanoyl chloride (purity 99.4%), captopril (bulk drug, Waisha Zhejiang pharmaceutical Co., Ltd.).
1. Detection limit and quantitation limit: the detection limit and the quantification limit are determined by a signal-to-noise ratio method, and the detection limit is determined by the corresponding concentration when the signal-to-noise ratio of the quantitative ions is 3:1, so that the result is 0.080 mu g/mL and is 2.402 percent of the limit. The limit of quantitation was determined at the corresponding concentration at a 10:1 signal-to-noise ratio of the quantitative ion, and the result was 0.266. mu.g/mL, which was 7.99% of the limit.
2. The specificity is as follows: taking 26.81mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c. Precisely measuring 100 mu L of reference substance solution c, placing in a 2mL sample injection vial, adding 900 mu L of methanol, and shaking up to obtain reference substance solution; precisely measuring 1.0 μ L of the reference solution, injecting into a gas chromatography-mass spectrometer, and recording chromatogram.
Taking 1.0g of captopril, precisely weighing, placing in a 10mL measuring flask, dissolving with methanol, fixing the volume to a scale, shaking up, filtering with a microporous filter membrane, and taking the subsequent filtrate as a test solution; and simultaneously performing a blank test. Precisely measuring the sample solution and the blank test solution by 1.0 μ L, respectively, injecting into a gas chromatography-mass spectrometer, and recording chromatogram.
And (4) conclusion: the retention time of the D-3-acetylthio-2-methylpropanoyl chloride in the reference solution is 5.16min, no interference peak appears in the blank solvent, the D-3-acetylthio-2-methylpropanoyl chloride in the test solution is not detected, no interference peak exists, and the method has good specificity.
3. Precision of the instrument: a control solution (with a limit concentration of 0.266. mu.g/mL) was sampled 6 times, and the RSD of the area of the quantitative ion peak was calculated.
TABLE 2 precision test of D-3-acetylmercapto-2-methylpropanoyl chloride
The results show that: the precision of the D-3-acetylmercapto-2-methylpropanoyl chloride analysis method is good.
4. Linearity and range: taking 25.45mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c.
The control solutions c 100. mu.L, 130. mu.L, 150. mu.L, 200. mu.L and 250. mu.L were precisely measured, and placed in 2mL injection vials, into which methanol 900. mu.L, 870. mu.L, 850. mu.L, 800. mu.L and 750. mu.L were added, respectively, and shaken well. According to the conditions of the chromatogram/mass spectrum, precisely measuring 1.0 mu L of the solution and injecting the solution into a gas chromatograph-mass spectrometer. Recording chromatogram, measuring peak area, drawing a standard curve by taking linear concentration as abscissa and quantitative ion peak area as ordinate, and calculating regression equation, wherein the result is shown in Table 3.
TABLE 3 determination of the D-3-acetylmercapto-2-methylpropanoyl chloride Linear relationship
| Concentration (x, μ g/mL) | Quantitative ion Peak area (y) |
| 0.2530 | 213 |
| 0.3289 | 268 |
| 0.3795 | 303 |
| 0.5060 | 395 |
| 0.6325 | 502 |
The result shows that the linear relation of the D-3-acetylmercapto-2-methylpropanoyl chloride is good in the range of 0.2530-0.6325 g/mL. The regression equation: Y757.03X +18.261, correlation coefficient: r2 ═ 0.9984.
5. Accuracy of
1) Limit concentration accuracy (recovery) test:
taking 25.45mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c. The control solution c 130. mu.L was precisely measured, placed in a 2mL injection vial, and 870. mu.L of methanol was added thereto, and shaken up to give a control solution.
Taking 1.0g of captopril, precisely weighing, and placing in a 10mL measuring flask to prepare 7 parts; dissolving 1 part of the filtrate with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a test solution, accurately adding the other 6 parts of the filtrate into a control solution c130 muL, dissolving with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a 100% recovery solution, and simultaneously performing a blank test.
Precisely measuring the reference substance solution, the blank solution, the sample solution and the 100% recovery rate solution to be 1.0 μ L respectively, injecting into a gas chromatography-mass spectrometer, and calculating the content, recovery rate and RSD result of D-3-acetylthio-2-methylpropanoyl chloride according to an external standard method.
The results are shown in Table 4.
2) Quantitative limiting concentration accuracy (recovery) test:
taking 23.39mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; precisely measuring 50 μ L of reference substance solution b, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution c;
the control solution c 130. mu.L was precisely measured, placed in a 2mL injection vial, and 870. mu.L of methanol was added thereto, and shaken up to give a control solution.
Taking 1.0g of captopril, precisely weighing, and placing in a 10mL measuring flask to prepare 4 parts; dissolving 1 part of the filtrate with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, collecting the subsequent filtrate as a sample solution, adding the other 3 parts of the filtrate into a control solution c100 μ L, dissolving with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, collecting the subsequent filtrate as a recovery limit solution, and performing a blank test.
Precisely measuring the reference substance solution, the blank solution, the sample solution and the quantitative limit recovery rate solution to be 1.0 μ L respectively, injecting into a gas chromatography-mass spectrometer, and calculating the content, recovery rate and RSD result of D-3-acetylthio-2-methylpropanoyl chloride according to an external standard method. The results are shown in Table 4.
TABLE 4 accuracy (recovery) test of D-3-acetylmercapto-2-methylpropanoyl chloride
The results show that: the accuracy of the D-3-acetylmercapto-2-methylpropanoyl chloride analysis method is good.
6. Repeatability of
(limit concentration accuracy (recovery rate) test under the same accuracy item of preparation method)
Taking 25.45mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c. The control solution c 130. mu.L was precisely measured, placed in a 2mL injection vial, and 870. mu.L of methanol was added thereto, and shaken up to give a control solution.
Taking 1.0g of captopril, precisely weighing, and placing in a 10mL measuring flask to prepare 7 parts; dissolving 1 part of the filtrate with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a test solution, accurately adding the other 6 parts of the filtrate into a control solution c130 muL, dissolving with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a 100% recovery solution, and simultaneously performing a blank test.
Precisely measuring the reference substance solution, the blank solution, the sample solution and the 100% recovery rate solution to be 1.0 μ L respectively, injecting into a gas chromatography-mass spectrometer, and calculating the content, recovery rate and RSD result of D-3-acetylthio-2-methylpropanoyl chloride according to an external standard method. The results are shown in Table 5.
TABLE 5 repeatability test of D-3-acetylmercapto-2-methylpropanoyl chloride
The results show that: the repeatability of the D-3-acetylmercapto-2-methylpropanoyl chloride analysis method is good.
7. Intermediate precision test
Taking 23.39mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c. The control solution c 130. mu.L was precisely measured, placed in a 2ml injection vial, and 870. mu.L of methanol was added thereto, and shaken up to give a control solution.
Taking 1.0g of captopril, precisely weighing, and placing in a 10mL measuring flask to prepare 7 parts; dissolving 1 part of the filtrate with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a test solution, accurately adding the other 6 parts of the filtrate into a control solution c130 muL, dissolving with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, taking the subsequent filtrate as a 100% recovery solution, and simultaneously performing a blank test.
Precisely measuring the reference solution, the blank solution, the sample solution and the 100% recovery rate solution by 1.0 μ L, respectively, injecting into a gas chromatography-mass spectrometer, inspecting different personnel and different dates, measuring the content according to the D-3-acetylthio-2-methylpropanoyl chloride analysis method, and calculating RSD. The results are shown in Table 6.
TABLE 6 intermediate precision test of D-3-acetylmercapto-2-methylpropanoyl chloride
The results show that: the intermediate precision of the D-3-acetylmercapto-2-methylpropanoyl chloride analysis method is good.
8. Stability test of solution
Taking 23.39mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c.
Taking captopril about 1.0g, precisely weighing, placing in a 10mL measuring flask, accurately adding a control solution c130 μ L, dissolving with methanol, fixing the volume to scale, shaking up, filtering with a microporous membrane, and taking the subsequent filtrate as a test solution (100% recovery solution).
Precisely measuring 1.0 μ L of the test solution, injecting into a gas chromatography-mass spectrometer, and recording the chromatogram. The sample injection is carried out continuously for 24 hours, and the RSD of the quantitative ion peak area is calculated. The results are shown in Table 7.
TABLE 7 solution stability test
The results show that: d-3-acetylmercapto-2-methylpropanoyl chloride in the captopril test solution has good stability within 8 hours.
9. Durability test
Taking 24.76mg of D-3-acetylmercapto-2-methylpropanoyl chloride, precisely weighing, placing in a 25mL measuring flask, dissolving with methanol, fixing the volume to a scale, and shaking up to obtain a reference solution a; precisely measuring 50 μ L of reference substance solution a, placing in 2mL sample injection vial, adding 950 μ L of methanol, and shaking to obtain reference substance solution b; the control solution b 50. mu.L was precisely measured, placed in a 2mL injection vial, added with 950. mu.L of methanol, and shaken up to give a control solution c. The control solution c 130. mu.L was precisely measured, placed in a 2mL injection vial, and 870. mu.L of methanol was added thereto, and shaken up to give a control solution.
Taking 1.0g of captopril, precisely weighing, and placing in a 10mL measuring flask to prepare 3 parts; dissolving 1 part of the filtrate with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, collecting the subsequent filtrate as a test solution, adding the rest 2 parts of the filtrate into a control solution c130 μ L, dissolving with methanol, fixing the volume to the scale, shaking up, filtering with a microporous membrane, collecting the subsequent filtrate as a 100% recovery solution, and performing a blank test.
The contents of the above solutions were measured by D-3-acetylmercapto-2-methylpropanoyl chloride analysis by varying the injection port temperature (210 ℃, 220 ℃, 230 ℃), the column flow rate (1.4mL/min, 1.5mL/min, 1.6mL/min) and the initial column temperature (65 ℃, 70 ℃, 75 ℃), respectively, and the results are shown in Table 8.
TABLE 8 durability test of D-3-acetylmercapto-2-methylpropanoyl chloride
The results show that: the durability of the D-3-acetylmercapto-2-methylpropanoyl chloride analysis method is good.
Third, sample detection
9 batches of captopril samples were assayed according to the above-described analytical method for D-3-acetylmercapto-2-methylpropanoyl chloride, and the results are shown in Table 9.
TABLE 99 determination of D-3-acetylmercapto-2-methylpropanoyl chloride in samples
The results show that: 9 batches of captopril samples were determined by D-3-acetylmercapto-2-methylpropanoyl chloride analysis, and none of the results were detected, which met the regulations.
The above description is only a preferred embodiment of the present application and is not intended to limit the present application, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present application shall be included in the protection scope of the present application.
Claims (6)
1. A method for detecting genotoxic impurity D-3-acetylmercapto-2-methylpropanoyl chloride in captopril bulk drug is characterized in that the used detection method is a gas chromatography-mass spectrometry combined method, and the method comprises the following steps:
(1) preparing a test solution: precisely weighing a sample of the nicarduopril, dissolving the sample by using an alcohol solvent, fixing the volume, shaking the sample uniformly to prepare a solution of 0.05-0.20 g/mL, filtering the solution by using a microporous filter membrane, taking a subsequent filtrate as a sample solution, and preparing a blank solution.
(2) Preparing a reference substance solution: precisely weighing a D-3-acetylmercapto-2-methylpropanoyl chloride reference substance, placing the reference substance in a 25mL volumetric flask, dissolving the reference substance by using a solvent, fixing the volume to a scale, and shaking up to obtain a reference substance solution a; precisely measuring 50 mu L of reference substance solution a, placing in a 2mL sample injection vial, adding 950 mu L of alcohol solvent, and shaking up to obtain reference substance solution b; precisely measuring 50 μ L of the control solution b, placing in a 2mL injection vial, adding 950 μ L of alcohol solvent, and shaking to obtain control solution c. Precisely measuring 100 μ L of the control solution, placing in a 2mL sample injection vial, adding 900 μ L of alcohol solvent, and shaking to obtain the control solution.
(3) Precisely measuring blank solution, reference solution and sample solution 1.0 μ L, respectively, injecting into gas chromatography-mass spectrometer for testing, and recording chromatogram.
2. The assay method of claim 1, wherein the concentration of the sample solution of step (1) in claim 1 is 0.10 g/mL.
3. The assay method of claim 1, wherein the concentration of the control solution in step (2) of claim 1 is 0.20-0.65 μ g/mL.
4. The method according to claim 1, wherein the diluting solvent used in step (1) (2) of claim 1 is an alcohol solvent such as methanol, ethanol and isopropanol.
5. The analytical method according to claim 1, wherein the conditions of the GC-MS test in step (3) of claim 1 are as follows:
gas chromatography conditions: the chromatographic column is HP-5, 30m × 0.25mm × 0.25 μm, UI; the stationary phase is 5% of phenyl-95% of dimethyl polysiloxane, and the carrier gas is helium; the temperature of a sample inlet is 200-240 ℃; an MS detector; the flow rate is 1.4-1.6 mL/min; the split ratio is 10: 1; the amount of sample was 1.0. mu.L.
Mass spectrum conditions: ion source (EI) voltage, ionization energy: 70eV, ion source temperature: 230 ℃, quadrupole temperature: 150 ℃, transmission line temperature: 280 ℃, scanning mode: selective ion scan, quantification of ions: 144, qualitative ion: 144. 134, 116, 101.
6. The analytical method according to claim 5, wherein the gas chromatography column temperature conditions are: the initial column temperature is 60-75 ℃, the initial column temperature is maintained for 1min, then the temperature is increased to 180 ℃ at the speed of 20 ℃/min, the temperature is increased to 260 ℃ at the speed of 40 ℃/min, and the initial column temperature is maintained for 10min at 260 ℃.
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| CN112326860A (en) * | 2020-10-30 | 2021-02-05 | 淄博高新技术产业开发区生物医药研究院 | Method for simultaneously detecting genotoxic impurities of 1, 4-dichlorobutane and 1-chloro-4-hydroxybutane in bulk drug or preparation |
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