CN105372376B - Detection method and application of parecoxib sodium genotoxic impurity - Google Patents
Detection method and application of parecoxib sodium genotoxic impurity Download PDFInfo
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Abstract
The invention provides a detection method and application of parecoxib sodium genotoxic impurities. The method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows: the chromatographic columns comprise a C18 column, a C8 column, a phenyl column and a Hilic column; the mobile phase composition is water-acetonitrile, diluted phosphoric acid-acetonitrile, phosphate-acetonitrile; the mobile phase ratio is 90: 10-10: 90 of water phase-organic phase; the column temperature is 25-40 ℃; the flow rate of the mobile phase is 0.2-2 ml/min; the detection wavelength is 205-290 nm; the detector is an ultraviolet detector or a diode array (PDA) detector. The sample amount is 0.1-40 μ l. The detection method of parecoxib sodium genotoxic impurities provided by the invention realizes the separation of parecoxib sodium and 3 genotoxic impurities in a short time, has high sensitivity and specificity, is simple and convenient to operate, achieves the separation degree of main components, genotoxic impurities and genotoxic impurities which is more than 1.5, can be used for quality control of parecoxib sodium, and has practical value.
Description
Technical Field
The invention belongs to the technical field of pharmaceutical analysis, and particularly relates to a detection method of parecoxib sodium genotoxic impurities, and more particularly relates to an analytical detection method of parecoxib sodium genotoxic impurities by reversed phase liquid chromatography.
Background
Nonsteroidal anti-inflammatory drugs (NSAIDs), which are analgesics commonly used for acute and chronic pain, act by inhibiting synthesis and release of prostaglandins through inhibition of Cyclooxygenase (COX), resulting in analgesic effect. The traditional non-steroidal anti-inflammatory drugs (such as naproxen, indomethacin, diclofenac, nimesulide and the like) are clinically applied for decades, but the application of the traditional non-steroidal anti-inflammatory drugs is limited in recent years, because the traditional non-steroidal anti-inflammatory drugs have obvious side effects after long-term use, gastrointestinal side effects such as dyspepsia and abdominal pain can be generated, and diseases such as gastric perforation and duodenal ulcer can be caused under severe conditions.
Parecoxib sodium is a novel COX-2 specific inhibitor. COX-2 is an inducible enzyme that is normally not or only slightly expressed in most tissues of the body, but is rapidly produced when stimulated. Studies have shown that specific COX-2 inhibitors are more therapeutically effective than non-selective cyclooxygenase inhibitors, with fewer side effects. In the treatment concentration, the parecoxib sodium can selectively inhibit COX-2 and Prostaglandin (PG) synthesis, so that the effects of analgesia and anti-inflammation are achieved, the COX-1 inhibition effect is not obvious, and the parecoxib sodium is widely used for clinically preventing and treating moderate and severe pain and preoperative analgesia.
Parecoxib Sodium (English name: Parecoxib Sodium) with molecular formula C19H18N2O4Na, molecular weight 392.40, CAS accession no: 198470-85-8. The pure parecoxib sodium product is white crystalline powder, and the melting point of the parecoxib sodium product is 273-275 ℃. The structure is as follows.
The research on impurities is a key content in the quality research of raw material medicaments. At present, the Chinese pharmacopoeia and the foreign pharmacopoeia do not contain a detection method for related substances or genotoxic impurities of parecoxib sodium raw materials and preparations. In the published patent, CN104749288A discloses a liquid chromatography analysis method for parecoxib sodium related substances, which controls 6 impurities such as 4- (5-methyl-3-phenyl-4-isoxazole) sodium benzenesulfonate, and does not search any patent about detection methods for parecoxib sodium genotoxic impurities.
In recent years, the quality control of raw material medicines increasingly pays more attention to the research on genotoxic (or called genotoxic) impurities. When the concentration of the genotoxic impurities is very low, the genetic materials of the human body can be damaged, so that gene mutation is caused, and tumors can be promoted. The refining step of the parecoxib sodium production process may generate impurities containing genotoxicity warning structures, namely sulfonates (see table 1), and requires special attention for quality control.
Since parecoxib sodium is commonly used as a postoperative analgesic, the treatment period is less than 1 month, and the acceptable intake of genotoxic impurities is 120 mug/day as regulated by the limit control of genotoxic impurities. The maximum daily dose of parecoxib sodium for injection is 80mg, and the control limit of genotoxic impurities is converted into single impurities (or total impurities) which do not exceed 1500 ppm.
TABLE 1 parecoxib sodium genotoxic impurities
Disclosure of Invention
The invention aims to provide a method for detecting 3 genotoxic impurities of parecoxib sodium.
The invention further aims to provide an application of the parecoxib sodium 3 genotoxic impurity detection method.
The purpose of the invention is mainly realized by the following technical scheme: the detection method adopts reversed phase liquid chromatography, and detection objects, control limits, sample preparation, chromatographic columns, mobile phase compositions, column temperature, flow rate, detection wavelength, sample introduction amount and the like are described as follows.
The detection objects are 4- (5-methyl-3-phenyl-4-isoxazole) benzene sulfonic acid methyl ester, 4- (5-methyl-3-phenyl-4-isoxazole) benzene sulfonic acid ethyl ester and 4- (5-methyl-3-phenyl-4-isoxazole) benzene sulfonic acid isopropyl ester.
The control limit of the invention is that each genotoxic impurity is controlled to be 100-2000 ppm, preferably 1000 ppm.
The sample is prepared by dissolving a sample with water, acetonitrile, methanol, water and acetonitrile (50:50), preferably methanol; the sample preparation concentration is 0.5-4 mg/ml, and preferably 1 mg/ml.
The chromatographic columns of the invention are C18 columns, C8 columns, phenyl columns and Hilic columns, and preferably C18 columns.
The mobile phase composition of the invention is water-acetonitrile, dilute phosphoric acid-acetonitrile, 0.01mol/L disodium hydrogen phosphate-acetonitrile, 0.01mol/L potassium dihydrogen phosphate-acetonitrile, 0.01mol/L ammonium dihydrogen phosphate-acetonitrile, preferably 0.01mol/L ammonium dihydrogen phosphate-acetonitrile; the mobile phase ratio is 90: 10-10: 90 of water phase-organic phase.
The column temperature of the invention is 25-40 ℃, preferably 35 ℃.
The flow rate of the mobile phase is 0.2-2.0ml/min, preferably 1.0 ml/min.
The detector is an ultraviolet detector or a diode array (PDA) detector;
the detection wavelength of the invention is 205-290 nm, preferably 230 nm.
The sample amount of the invention is 0.1-40 μ l, preferably 20 μ l.
The detection method of parecoxib sodium genotoxic impurities provided by the invention realizes the separation of parecoxib sodium and 3 genotoxic impurities in a short time, has high sensitivity and specificity, is simple and convenient to operate, achieves the separation degree of main components, genotoxic impurities and genotoxic impurities which is more than 1.5, can be used for quality control of parecoxib sodium, and has practical value.
Drawings
FIG. 1 is a liquid chromatogram of example 7;
FIG. 2 is a liquid chromatogram of the first injection in example 11;
FIG. 3 is a liquid chromatogram of a control sample mixture of genotoxic impurities from example 12;
FIG. 4 is a liquid chromatogram of the sample measurement in example 12;
Detailed Description
The present invention is further described in detail below with reference to specific examples, which are given only for the purpose of illustrating the present invention and are not intended to limit the scope of the present invention.
Example 1
The invention relates to a preparation method of a parecoxib sodium genotoxicity impurity detection solution.
In the test, water, acetonitrile, methanol and water acetonitrile (1:1) are selected to dissolve a sample, the sample is slightly soluble in water, slightly soluble in acetonitrile and easily soluble in methanol and water acetonitrile (1: 1). 3 genotoxic impurities are required to be added into a parecoxib sodium sample during chromatographic condition examination, the structures of the impurity A, the impurity B and the impurity C are ester substances, the ester substances are easy to hydrolyze, and the solvent is required to be not suitable for containing water, so methanol is preferably used as the solvent for dissolving the sample.
In the test, the parecoxib sodium sample is prepared into a solution of 0.5-4 mg/ml for investigation, and the result shows that the 0.5mg/ml impurity response is small, the asymmetry of the 4mg/ml main peak is serious, the detection of the impurity is influenced, and 1mg/ml is preferably used as the concentration of the test sample.
Example 2
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-uv detector;
a chromatographic column: c8 column (4.6X 150mm,5 μm);
mobile phase: water acetonitrile (50: 50);
column temperature: 25 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 10 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 40mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The results showed that the separation degree between each impurity and the main component was good, and the main component peak was split.
Example 3
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-uv detector;
a chromatographic column: c8 column (4.6X 150mm,5 μm);
mobile phase: pH3.0 Dilute phosphoric acid acetonitrile (70: 30);
column temperature: 30 ℃; flow rate: 2 mL/min;
detection wavelength: 290 nm; sample introduction amount: 40 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The results showed that the degree of separation between the components was good and the peak shape of each component was good.
Example 4
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-uv detector;
a chromatographic column: c18 column (4.6X 250mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (10: 90);
column temperature: 35 ℃; flow rate: 0.8 mL/min;
detection wavelength: 230 nm; sample introduction amount: 20 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 20mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The result shows that the base line of the spectrogram is stable, all the impurities are subjected to peak emergence within 5min, and the components are not separated from each other by the base line.
Example 5
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters UPLC-PDA detector;
a chromatographic column: phenyl column (2.1X 50mm,1.7 μm);
mobile phase: 0.01mol/L sodium dihydrogen phosphate (pH 3.0 adjusted to phosphoric acid) acetonitrile (40: 60);
column temperature: 35 ℃; flow rate: 0.2 mL/min;
detection wavelength: 205 nm; sample introduction amount: 0.1. mu.l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.1mg/ml solution as impurity stock solution; accurately weighing 5mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The result shows that the base line of the spectrogram is stable, all the impurities are peaked within 5min, the separation degree among the components is good, the peak shape of each component is good, and the separation degree between the known impurities and the unknown impurities is 1.35 (less than 1.5).
Example 6
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters UPLC-PDA detector;
chromatography column Hilic column (2.1X 50mm,1.7 μm);
mobile phase: 0.01mol/L sodium dihydrogen phosphate (pH 3.0 adjusted to phosphoric acid) acetonitrile (90: 10);
column temperature: 40 ℃; flow rate: 0.3 mL/min;
detection wavelength: 230 nm; sample introduction amount: 0.5. mu.l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.1mg/ml solution as impurity stock solution; accurately weighing 5mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The result shows that the spectrogram base line is stable, all the impurities are subjected to peak extraction within 10min, the separation degree among the components is good, and the peak shape symmetry is poor.
Example 7
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-uv detector;
a chromatographic column: c18 column (4.6X 250mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (45: 55);
column temperature: 35 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 20 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and examining separation degree of main component and each impurity (see figure 1 in detail). The result shows that the base line of the spectrogram is stable, all the impurities are subjected to peak emergence within 10min, the separation degree among the components is good, and the peak shape of each component is good.
Example 8
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-uv detector;
column C18 (4.6X 150mm,5 μm);
mobile phase: 0.01mol/L potassium dihydrogen phosphate (adjusted to pH3.0 with phosphoric acid) acetonitrile (45: 55);
column temperature: 40 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 10 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The result shows that the base line of the spectrogram is stable, all the impurities are subjected to peak emergence within 7min, the separation degree among the components is good, and the peak shape of each component is good.
Example 9
The invention discloses a detection method of parecoxib sodium genotoxic impurities
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-PDA detector;
column C18 (4.6X 150mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (45: 55);
column temperature: 35 ℃; flow rate: 0.8 mL/min;
detection wavelength: 230 nm; sample introduction amount: 10 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 0.5mg/ml solution as impurity stock solution; accurately weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, accurately weighing 1ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a chromatographic condition screening solution.
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, running for 20min, recording chromatogram, and inspecting the separation degree of main components and each impurity. The results show that the base line of the spectrogram is stable, the impurity C generates a peak after 10min, the peak shape is wide, the separation degree among the components is good, and the peak shape of each component is good.
Examples 2 to 7 focus on the examination of the influence of chromatographic conditions on the peak shape and the degree of separation of the main component and the genotoxic impurities, and each example basically realizes the specificity examination of 3 genotoxic impurities. Through comparison of system applicability parameters, the genetic toxicity impurities of the parecoxib sodium are preferably inspected by using a C18 chromatographic column, 0.01mol/L ammonium dihydrogen phosphate (phosphoric acid is adjusted to pH3.0) and acetonitrile (45:55) as a mobile phase under chromatographic conditions of column temperature of 35 ℃, flow rate of 1.0ml/min and detection wavelength of 230 nm.
Under the optimized conditions (see figure 1), the main components and the impurity attributes of each isomer are shown in table 2.
TABLE 2 Peak assignment of parecoxib sodium genotoxic impurities
| Serial number | Retention time (RT, min) | Relative Retention Time (RRT) | Attribution |
| 1 | 2.951 | 1.00 | Parecoxib sodium |
| 2 | 4.663 | 1.58 | Impurity A |
| 3 | 5.785 | 1.96 | Impurity B |
| 4 | 7.217 | 2.45 | Impurity C |
Example 10
System applicability test of parecoxib sodium genotoxicity impurity detection method
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-PDA detector;
column C18 (4.6X 150mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (45: 55);
column temperature: 35 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 20 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 10 μ g/ml solution as impurity stock solution; the method comprises the steps of precisely weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, precisely weighing 0.5ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a system applicability solution 1 (each genotoxic impurity contains 500 ppm).
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, continuously injecting sample for 5 needles, operating for 15min each time, recording chromatogram (figure 2 is chromatogram for first injection), and inspecting main component and each impurity, separation degree between each impurity and peak area repeatability of each impurity (see table 3). The result shows that the base line of the spectrogram is stable, the peak appearance of each component is finished within 8min, the separation degree among the components is good, and the repeatability of the peak area of each impurity is good.
TABLE 3 precision test results of System suitability solution 1
| Number of samples taken | Area of impurity A peak | Area of impurity B peak | Peak surface of impurity CProduct of large quantities |
| 1 | 24776 | 19480 | 14201 |
| 2 | 25780 | 18733 | 14557 |
| 3 | 25991 | 19093 | 14366 |
| 4 | 25164 | 19958 | 14557 |
| 5 | 24948 | 19648 | 13867 |
| Mean value of | 25332 | 19382 | 14310 |
| RSD％ | 2.09 | 2.47 | 2.02 |
Example 11
System applicability test of parecoxib sodium genotoxicity impurity detection method
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-PDA detector;
column C18 (4.6X 250mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (45: 55);
column temperature: 35 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 20 μ l.
2) Solution preparation
Precisely weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into 10 μ g/ml solution as impurity stock solution; the method comprises the steps of precisely weighing 10mg to 10ml of parecoxib sodium in a volumetric flask, precisely weighing 0.5ml of each impurity stock solution in the volumetric flask, adding methanol to dissolve and dilute the impurity stock solution to a scale, and using the impurity stock solution as a system applicability solution 2 (each genotoxic impurity contains 1000 ppm).
3) Measurement method and results
Injecting sample according to 1) chromatographic condition, continuously injecting sample for 5 needles, operating for 15min each time, recording chromatogram, and inspecting main components and each impurity, separation degree among impurities and peak area repeatability of each impurity (see table 4). The result shows that the base line of the spectrogram is stable, the peak appearance of each component is finished within 8min, the separation degree among the components is good, and the repeatability of the peak area of each impurity is good.
TABLE 4 precision test results of System suitability solution 2
| Number of samples taken | Area of impurity A peak | Area of impurity B peak | Area of impurity C peak |
| 1 | 42257 | 39337 | 27905 |
| 2 | 43042 | 39091 | 29566 |
| 3 | 42570 | 38737 | 28860 |
| 4 | 42741 | 38147 | 28034 |
| 5 | 42930 | 38905 | 29278 |
| Mean value of | 42708 | 38842 | 28729 |
| RSD％ | 0.73 | 1.15 | 2.57 |
Examples 10 and 11 focus on the establishment of limits for genotoxic impurities, and each impurity was added at 500ppm and 1000ppm, and the amount of the sample was increased to 20. mu.l to increase the amount of the sample to be detected, and the system applicability was examined, preferably 1000 ppm.
Example 12
The invention relates to sample detection of a parecoxib sodium genotoxicity impurity detection method.
1) Instrumentation and test conditions
The instrument comprises the following steps: waters E2695 high performance liquid chromatograph-PDA detector;
column C18 (4.6X 250mm,5 μm);
mobile phase: 0.01mol/L ammonium dihydrogen phosphate (pH 3.0 adjusted by phosphoric acid) acetonitrile (45: 55);
column temperature: 35 ℃; flow rate: 1.0 mL/min;
detection wavelength: 230 nm; sample introduction amount: 20 μ l.
2) Solution preparation
Accurately weighing a proper amount of parecoxib sodium sample, adding methanol to dissolve and dilute the parecoxib sodium sample into a solution of 1mg/ml, and taking the solution as a test solution; accurately weighing appropriate amount of reference substances of impurity A, impurity B and impurity C, adding methanol to dissolve and dilute into mixed solution of 1 μ g/ml as reference substance solution of impurity.
3) Measurement method and results
Sequentially sampling the test solution and impurity reference solution under 1) chromatographic conditions, running for 15min each time, and recording chromatogram. If the test solution contains genotoxic impurities, the content of each impurity is calculated by an external standard method. The result shows that the impurity A is detected in the parecoxib sodium sample, the content is 300ppm, and the impurity B and the impurity C are not detected. The liquid chromatogram of the reference substance mixture of genotoxic impurities is shown in figure 3; the liquid chromatogram of sample measurement is shown in figure 4.
Claims (6)
1. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C8 column;
the mobile phase composition is water-acetonitrile 50: 50;
the column temperature was 25;
the flow rate of the mobile phase is 1.0 ml/min;
the detection wavelength is 230 nm;
the detector is an ultraviolet detector;
the sample amount is 10 mul;
the sample solution is dissolved by methanol, and the concentration is 4 mg/ml.
2. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C8 column;
the mobile phase composition is diluted phosphoric acid with pH3.0-acetonitrile 70: 30;
the column temperature is 30 ℃;
the flow rate of the mobile phase is 2.0 ml/min;
the detection wavelength is 290 nm;
the detector is an ultraviolet detector;
the sample amount is 40 mul;
the sample solution is dissolved by methanol with the concentration of 1 mg/ml.
3. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C18 column;
mobile phase composition 0.01mol/L ammonium dihydrogen phosphate-acetonitrile ph3.0 ═ 10: 90;
the column temperature was 35 ℃;
the flow rate of the mobile phase is 0.8 ml/min;
the detection wavelength is 230 nm;
the detector is an ultraviolet detector;
the sample amount is 20 mul;
the sample solution is dissolved by methanol with the concentration of 2 mg/ml.
4. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C18 column;
mobile phase composition 0.01mol/L ammonium dihydrogen phosphate-acetonitrile ph3.0 ═ 45: 55;
the column temperature was 35 ℃;
the flow rate of the mobile phase is 1.0 ml/min;
the detection wavelength is 230 nm;
the detector is an ultraviolet detector;
the sample amount is 20 mul;
the sample solution is dissolved by methanol with the concentration of 1 mg/ml.
5. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C18 column;
mobile phase composition 0.01mol/L ph3.0 potassium dihydrogen phosphate-acetonitrile 45: 55;
the column temperature was 40 ℃;
the flow rate of the mobile phase is 1.0 ml/min;
the detection wavelength is 230 nm;
the detector is an ultraviolet detector;
the sample amount is 10 mul;
the sample solution is dissolved by methanol with the concentration of 1 mg/ml.
6. The detection method of parecoxib sodium genotoxic impurities is characterized in that the parecoxib sodium genotoxic impurities comprise 4- (5-methyl-3-phenyl-4-isoxazole) methyl benzenesulfonate, 4- (5-methyl-3-phenyl-4-isoxazole) ethyl benzenesulfonate and 4- (5-methyl-3-phenyl-4-isoxazole) isopropyl benzenesulfonate; the detection method adopts reversed phase liquid chromatography, and the chromatographic conditions are as follows:
the chromatographic column is a C18 column;
mobile phase composition 0.01mol/L ammonium dihydrogen phosphate-acetonitrile ph3.0 ═ 45: 55;
the column temperature was 35 ℃;
the flow rate of the mobile phase is 0.8 ml/min;
the detection wavelength is 230 nm;
the detector is an ultraviolet detector;
the sample amount is 10 mul;
the sample solution is dissolved by methanol with the concentration of 1 mg/ml.
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| CN107037153B (en) * | 2017-04-21 | 2019-05-14 | 常州佳德医药科技有限公司 | The method that high performance liquid chromatography detects genotoxicity impurity in AL58805 bulk pharmaceutical chemicals or pharmaceutical preparation |
| CN107976500B (en) * | 2017-11-29 | 2021-09-28 | 浙江震元制药有限公司 | Diaryl-substituted isoxazole compound and preparation method and application thereof |
| CN107782832B (en) * | 2017-12-11 | 2019-12-06 | 重庆华邦制药有限公司 | Method for separating and determining bepotastine besilate and potential genotoxic impurities thereof by HPLC (high performance liquid chromatography) |
| CN108828127A (en) * | 2018-06-21 | 2018-11-16 | 上药东英（江苏）药业有限公司 | Liquid-phase chromatography method in relation to substance in a kind of detection Parecoxib Sodium and synthetic intermediate |
| CN110642801A (en) * | 2019-09-20 | 2020-01-03 | 深圳市祥根生物科技有限公司 | Preparation method of parecoxib meta-isomer impurity |
| CN111307958A (en) * | 2019-11-25 | 2020-06-19 | 澳美制药(苏州)有限公司 | Method for detecting genotoxic impurities in bepotastine besilate |
| CN110938043A (en) * | 2019-11-27 | 2020-03-31 | 上海秀新臣邦医药科技有限公司 | Parecoxib sodium genotoxic impurity and preparation method thereof |
| CN111089931A (en) * | 2019-11-28 | 2020-05-01 | 上海秀新臣邦医药科技有限公司 | Detection method of parecoxib sodium gene genotoxicity impurity |
| CN114441666B (en) * | 2020-11-05 | 2024-02-27 | 成都百裕制药股份有限公司 | Method for detecting impurities in 4- (5-methyl-3-phenyl-4-isoxazole) benzenesulfonyl chloride |
| CN114460203B (en) * | 2022-02-15 | 2023-12-22 | 浙江海正药业股份有限公司 | Method for simultaneously detecting methyl p-toluenesulfonate, ethyl p-toluenesulfonate and isopropyl p-toluenesulfonate in medicine |
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