CN114002336A - Method for analyzing and detecting thiophene ethylamine and related substances thereof - Google Patents

Method for analyzing and detecting thiophene ethylamine and related substances thereof Download PDF

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CN114002336A
CN114002336A CN202010736471.4A CN202010736471A CN114002336A CN 114002336 A CN114002336 A CN 114002336A CN 202010736471 A CN202010736471 A CN 202010736471A CN 114002336 A CN114002336 A CN 114002336A
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CN114002336B (en
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赵涛涛
徐煜涵
王成
李天晴
洪健
张琦
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WUHAN WUYAO PHARMACEUTICAL CO Ltd
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Abstract

The invention relates to the technical field of chemical analysis, in particular to a method for analyzing and detecting thiophene ethylamine and related substances thereof. The invention provides a method for analyzing and detecting thiophene ethylamine and related substances thereof, which effectively separates the thiophene ethylamine and the related substances thereof by using a high performance liquid chromatography analysis method, and further analyzes the purity of the thiophene ethylamine, thereby realizing the quality control of clopidogrel bulk drugs and preparations thereof.

Description

Method for analyzing and detecting thiophene ethylamine and related substances thereof
Technical Field
The invention relates to the technical field of chemical analysis, in particular to a method for analyzing and detecting thiophene ethylamine and related substances thereof.
Background
The thiophene ethylamine is a key raw material for producing 4,5,6, 7-tetrahydrothieno [3,2-c ] pyridine hydrochloride, and the 4,5,6, 7-tetrahydrothieno [3,2-c ] pyridine hydrochloride is an important medical intermediate and is mainly used for synthesizing platelet coagulation inhibitors such as clopidogrel drugs. In order to improve the quality of clopidogrel drug production, the quality of thiopheneethylamine needs to be effectively analyzed and controlled according to drug production specifications and product standards, such as those specified in the United States Pharmacopeia (USP). Analysis of the thiophene ethylamine process route, several impurity components that may be present are:
Figure BDA0002605158600000011
at present, a method for analyzing and detecting thiophene ethylamine and related substances thereof simply, quickly and at low cost is not reported for a long time.
Therefore, a method for simply, quickly and conveniently analyzing and detecting the thiophene ethylamine and related substances thereof is needed to be developed, so that the quality control of the clopidogrel bulk drug and the preparation thereof is realized.
Disclosure of Invention
The present invention is directed to solving, at least to some extent, one of the technical problems in the related art. Therefore, an object of the present invention is to provide a method for analyzing and detecting thiophene ethylamine and related substances thereof, wherein the thiophene ethylamine and related substances thereof are effectively separated by a high performance liquid chromatography analysis method, and the purity of the thiophene ethylamine is further analyzed, so as to realize quality control of clopidogrel bulk drug and preparations thereof.
Therefore, the invention provides a method for analyzing and detecting the thiophene ethylamine and related substances thereof in a first aspect. According to the embodiment of the invention, the method utilizes high performance liquid chromatography to detect a sample to be detected so as to obtain a chromatogram map; and
obtaining the content of thiophene ethylamine in the sample to be detected based on the chromatogram;
wherein the chromatographic conditions of the high performance liquid chromatography are as follows:
the chromatographic column adopts an octadecylsilane chemically bonded silica gel column;
the Ghost peak catching column is Welch Ghost-Buster;
taking buffer solution as mobile phase A and organic solvent as mobile phase B to carry out gradient elution,
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
The mobile phase A is an ammonium salt solution, preferably an ammonium chloride aqueous solution, and the mobile phase B is acetonitrile.
The analysis and detection method can separate the thiophene ethylamine from the impurity A (2- (thiophene-3-yl) ethylamine), the impurity C (thiophene-2-formaldehyde) and the impurity D (thiophene) existing in the production process of the thiophene ethylamine, and further analyze and detect the purity of the thiophene ethylamine so as to realize the quality control of the thiophene ethylamine and further realize the quality control of the clopidogrel bulk drug and the preparation thereof.
The method for analyzing and detecting the thiophene ethylamine and related substances thereof according to the embodiment of the invention can also have at least one of the following additional technical characteristics:
according to an embodiment of the invention, the concentration of mobile phase a is 10 mM.
According to an embodiment of the invention, the pH of the mobile phase a is 9.10 ± 0.05.
Preferably, the mobile phase a is an aqueous ammonium chloride solution, and is prepared by the following method: dissolving a certain amount of ammonium chloride solid in water, dissolving by ultrasonic, and adjusting the pH value to 9.10 +/-0.05 by using ammonia water.
The pH value of the mobile phase A can influence the separation result, and the main component peak can generate tailing phenomenon, poor peak shape and interference with the separation effect of other impurity peaks when the pH value is too high or too low.
According to the embodiment of the invention, the chromatographic column in the high performance liquid chromatography is Waters Xbridge C18.
According to a preferred embodiment of the present invention, the high performance liquid chromatography column is Waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm), and the Ghost capturing column is Welch Ghost-Buster (50 mm. times.2.1 mm).
In chromatographic separation, particularly in a gradient elution process, famous and wonderful chromatographic peaks, commonly called ghost peaks, are easily generated and mainly come from a mobile phase and a pipeline, and once ghost peaks appear in method development, much time and energy are needed for an analyst to eliminate the ghost peaks. In order to effectively trap the generated ghost peaks and eliminate the interference of the ghost peaks on material analysis, the invention is provided with the ghost peak trapping column which can remove the impurities in the solvent including the organic solvent and can effectively trap the impurities in the pipeline and the mixer; impurities are adsorbed, so that the peaks are more gradually seen, and the interference of impurity peaks and solvent peaks thereof is reduced.
According to the embodiment of the invention, the temperature of the chromatographic column is 38-42 ℃, and is preferably 40 ℃.
According to an embodiment of the invention, the flow rate of the mobile phase is 0.9-1.1 ml/min, preferably 0.9-1.0 ml/min.
The column temperature of the chromatographic column and the flow rate of the mobile phase affect the separation effect, and if the column temperature of the chromatographic column or the flow rate of the mobile phase is not within the above range, the separation effect is poor, the separation degree is less than 1.5, the main peak and the impurity peak cannot be well separated, and the peak shape is also poor.
According to the embodiment of the invention, the detection wavelength in the high performance liquid chromatography is 208-212 nm, preferably 210 nm.
According to an embodiment of the invention, the method further comprises the steps of:
(1) dissolving a sample to be detected containing thiophene ethylamine by using acetonitrile to prepare a sample solution with the concentration of 2 mg/ml;
(2) and (2) injecting 5 mu l of the sample solution obtained in the step (1) into a high performance liquid chromatograph, and recording a chromatogram to finish the separation and detection of the sample containing the thiophene ethylamine.
The second aspect of the invention provides the application of the method of the first aspect in the analytical detection of clopidogrel or the salt thereof by using thiophene ethylamine as a raw material.
The method for analyzing and detecting the thiophene ethylamine and related substances thereof can be realized according to the following method:
chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 0.9-1.1 ml/min;
column temperature of the chromatographic column: 38-42 ℃;
detection wavelength: 208 to 212nm
Sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample by using mobile phase B acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mul of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and completing the analysis and separation of the sample containing the thiophene ethylamine.
When each component dissolved in the mobile phase passes through the stationary phase, the components flow out of the stationary phase successively due to different retention times in the stationary phase due to different sizes and strengths of actions (adsorption, distribution, ion attraction, exclusion and affinity) with the stationary phase. High performance liquid chromatography is based on this principle to achieve separation of substances.
The inventor creatively discovers that the method for analyzing and detecting the thiophene ethylamine and the related substances thereof can analyze the purity of the thiophene ethylamine, thereby realizing the quality control of the thiophene ethylamine and further realizing the quality control of clopidogrel bulk drugs and preparations thereof, and if the content of the thiophene ethylamine in a sample to be detected does not reach the standard, the sample to be detected cannot be used as a raw material for producing clopidogrel for producing the clopidogrel.
Additional aspects and advantages of the invention will be set forth in part in the description which follows and, in part, will be obvious from the description, or may be learned by practice of the invention.
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The above and/or additional aspects and advantages of the present invention will become apparent and readily appreciated from the following description of the embodiments, taken in conjunction with the accompanying drawings of which:
FIG. 1 shows a high performance liquid chromatogram of a sample obtained according to example 1 of the present invention;
FIG. 2 shows a high performance liquid chromatogram of a sample obtained according to example 2 of the present invention;
FIG. 3 shows a high performance liquid chromatogram of a sample obtained according to example 3 of the present invention;
FIG. 4 shows a high performance liquid chromatogram of a sample obtained according to example 4 of the present invention;
FIG. 5 shows a high performance liquid chromatogram of a sample obtained according to example 5 of the present invention;
FIG. 6 shows a high performance liquid chromatogram of a sample obtained according to example 6 of the present invention;
FIG. 7 shows a high performance liquid chromatogram of a sample obtained according to example 7 of the present invention;
FIG. 8 shows high performance liquid chromatograms (enlarged chromatogram pool around main peak) of the samples of example 8 of the present invention and comparative examples 1 to 7.
Detailed Description
The following describes embodiments of the present invention in detail. The following examples are illustrative only and are not to be construed as limiting the invention. The examples, where specific techniques or conditions are not indicated, are to be construed according to the techniques or conditions described in the literature in the art or according to the product specifications. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.
In the embodiment of the invention, the sample containing thiophene ethylamine is purchased from Michelin, a high performance liquid chromatograph: shimadzu LC-20A series high performance liquid chromatograph.
Example 1
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 1.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 1, the peak 1 is impurity A, the peak 2 is thiophene ethylamine, the peak 4 is impurity C, and the peak 11 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 8.560 min.
Example 2
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 2.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 208 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 2, the peak 1 is impurity A, the peak 2 is thiophene ethylamine, the peak 4 is impurity C, and the peak 12 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 8.560 min.
Example 3
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 3.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 212 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 3, the peak 1 is impurity A, the peak 2 is thienylethylamine, the peak 4 is impurity C, and the peak 11 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 8.560 min.
Example 4
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 4.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 0.9 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 4, the peak 1 is impurity A, the peak 2 is thienylethylamine, the peak 5 is impurity C, and the peak 11 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 9.309 min.
Example 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 5.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.1 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 5, the peak 1 is impurity A, the peak 2 is thienylethylamine, the peak 4 is impurity C, and the peak 10 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 7.944 min.
Example 6
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 6.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 38 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 6, the peak 1 is impurity A, the peak 3 is thienylethylamine, the peak 5 is impurity C, and the peak 12 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 8.197 min.
Example 7
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mu l of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and finishing the analysis and separation of the sample containing the thiophene ethylamine as shown in figure 7.
Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM ammonium chloride buffer (pH 9.10 ± 0.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 42 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
In FIG. 7, the peak 1 is impurity A, the peak 2 is thienylethylamine, the peak 4 is impurity C, and the peak 11 is impurity D. Under the condition, the main peak of the thiophene ethylamine can be well separated from impurities, the separation degree of the main peak and the adjacent peak is more than 1.5, and the main peak of the thiophene ethylamine is about 8.035 min.
Example 8
Compared with example 1, the difference is only that the temperature of the chromatographic column is changed from 40 ℃ to 41 ℃ in the chromatographic conditions, and the rest are the same, and the chromatogram is shown in FIG. 8 a. In fig. 8a the main component peaks are well symmetrical, the peak pattern is better and the degree of separation from adjacent peaks is better.
Comparative example 1
Compared with example 8, the difference is that the pH of the mobile phase A ammonium chloride buffer solution in the chromatographic condition is 8.07, the rest is the same, the obtained spectrum is shown in figure 8b, the main peak in figure 8b is triangular, the symmetry is poor, the peak emergence time is advanced, the main component peak has a tailing phenomenon, and whether the main component peak is separated from other peaks or not cannot be judged.
Comparative example 2
Compared with example 8, the difference is only that the pH of the mobile phase A ammonium chloride buffer solution in the chromatographic condition is 7.77, and the rest is the same, and the obtained spectrum is shown as the attached figure 8 c. In fig. 8c, the main peak is triangular, the peak-off time is advanced, the symmetry is poor, the response is low, the main component peak has some trailing phenomena, and whether the main component peak is separated from other peaks or not cannot be judged.
Comparative example 3
1. Chromatographic conditions are as follows:
a chromatographic column: CSH-C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM NH4FA buffer (pH 8.32); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mul of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and completing the analysis and separation of the sample containing the thiophene ethylamine.
The obtained separation result is shown in figure 8d, and a small peak appears before the main peak, which indicates that the separation effect of the main component peak and other impurity peaks is not good.
Comparative example 4
Comparative example 4 differs from comparative example 3 only in the chromatographic column used: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm) obtained the separation as shown in FIG. 8e, with some tailing of the main component peaks.
Comparative example 5
1. Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mM NH4HCO3Buffer (pH 9.56); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mul of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and completing the analysis and separation of the sample containing the thiophene ethylamine.
The separation results obtained are shown in fig. 8f, with some tailing of the main component peaks.
Comparative example 6
1. Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 10mMK2HPO4Buffer (pH 9.05); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mul of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and completing the analysis and separation of the sample containing the thiophene ethylamine.
The obtained separation result is shown in fig. 8g, where the main component has a triangular peak shape, an asymmetric and sharp peak shape, and a poor peak shape.
Comparative example 7
1. Chromatographic conditions are as follows:
a chromatographic column: waters Xbridge C18(150 mm. times.4.6 mm, 3.5 μm);
ghost peak trapping column: welch Ghost-Buster 50mm × 2.1 mm;
mobile phase: phase A: 0.01% NH3.H2O buffer (pH 9.17); phase B: acetonitrile
Flow rate of mobile phase: 1.0 ml/min;
column temperature of the chromatographic column: 40 ℃;
detection wavelength: 210 nm;
sample introduction amount: 5 mu L of the solution;
the gradient elution conditions were:
time (min) A(％) B(％)
0 95 5
20 20 80
25 20 80
26 95 5
40 95 5
1) Taking a sample containing thiophene ethylamine, dissolving the sample with acetonitrile, and preparing a sample solution with the concentration of 2 mg/ml;
2) injecting 5 mul of the sample solution obtained in the step 1) into a high performance liquid chromatograph, recording a chromatogram, and completing the analysis and separation of the sample containing the thiophene ethylamine.
The obtained separation result is shown in fig. 8h, and the peak shape of the main component is triangular, the peak shape is asymmetric and sharp, and the peak shape is poor.
In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.

Claims (9)

1. A method for analyzing and detecting thiophene ethylamine and related substances thereof is characterized in that a sample to be detected is detected by using a high performance liquid chromatography so as to obtain a chromatogram map; and
obtaining the content of thiophene ethylamine in the sample to be detected based on the chromatogram;
wherein the chromatographic conditions of the high performance liquid chromatography are as follows:
the chromatographic column adopts an octadecylsilane chemically bonded silica gel column;
the Ghost peak catching column is Welch Ghost-Buster;
taking buffer solution as mobile phase A and organic solvent as mobile phase B to carry out gradient elution,
the gradient elution conditions were:
time (min) A(％) B(％) 0 95 5 20 20 80 25 20 80 26 95 5 40 95 5
The mobile phase A is an ammonium salt solution, preferably an ammonium chloride aqueous solution, and the mobile phase B is acetonitrile.
2. The method according to claim 1, wherein the concentration of mobile phase a is 10 mM.
3. The method according to claim 1, wherein the mobile phase a has a pH of 9.10 ± 0.05.
4. The method of claim 1, wherein the high performance liquid chromatography column is Waters Xbridge C18.
5. The method according to claim 4, wherein the column temperature of the chromatographic column is 38 to 42 ℃, preferably 40 ℃.
6. The method according to claim 1, wherein the flow rate of the mobile phase is 0.9 to 1.1ml/min, preferably 0.9 to 1.0 ml/min.
7. The method according to claim 1, wherein the detection wavelength in the high performance liquid chromatography is 208-212 nm, preferably 210 nm.
8. A method according to any one of claims 1 to 7, further comprising the steps of:
(1) dissolving a sample to be detected containing thiophene ethylamine by using acetonitrile to prepare a sample solution with the concentration of 2 mg/ml;
(2) and (2) injecting 5 mu l of the sample solution obtained in the step (1) into a high performance liquid chromatograph, and recording a chromatogram to finish the separation and detection of the sample containing the thiophene ethylamine.
9. Use of the method of any one of claims 1-8 in analytical testing of clopidogrel or its salts produced from thienylethylamine.
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