A normal phase detection method for imine reaction in-process control

Abstract

The application belongs to the technical field of detection and analysis, and discloses a normal phase detection method for imine reaction control. The method is detected by normal phase liquid chromatography, and the detection conditions include: a polysaccharide derivative normal phase coated type chiral chromatographic column is used, and a mixed solution of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid is used as the mobile phase, wherein the volume ratio of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid is (75-85):(15-25):(0.05-0.15):(0.05-0.15). The method can realize effective separation, detect the content of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]azepine in the reaction solution, and has the advantages of simplicity, accuracy, rapidness and reliability.

Description

Technical Field

[0001] This invention belongs to the field of detection and analysis technology, and specifically relates to a normal-phase detection method for the control of imine reaction. Background Technology

[0002] 1,2,3,4,10,14b-hexahydro-2-methyldibenzo[c,f]pyrazino[1,2-a]azapyridine hydrochloride (mianserin hydrochloride), CAS number 21535-47-7, is an antidepressant. 1,2,3,4,10,14B-hexahydro-2-methyldibenzo[c,f]pyrazino[1,2-a]azapyridine hydrochloride, CAS number 71936-92-0, is an important intermediate in the synthesis of 1,2,3,4,10,14b-hexahydro-2-methyldibenzo[c,f]pyrazino[1,2-a]azapyridine hydrochloride. The completeness of its reaction directly affects the yield of subsequent reactions and the quality of the final product; therefore, its content in the reaction solution needs to be monitored during production. Because imine compounds are generated during this reaction, the system must be free of water, and reversed-phase liquid chromatography cannot be used to monitor the reaction progress.

[0003] Currently, no literature or reports have been found regarding the monitoring of the reaction process of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza. In order to strengthen the quality control of mianserin hydrochloride, it is particularly important to provide a normal-phase detection method for the mid-control of the imine reaction. Summary of the Invention

[0004] The present invention aims to solve at least one of the technical problems existing in the prior art. To this end, the present invention proposes a normal-phase detection method for the control of imine reaction. This method can effectively separate and detect the content of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazine[1,2-A]aza in the reaction solution, and has the advantages of being simple, accurate, rapid and reliable.

[0005] In a first aspect, the present invention provides a normal-phase detection method for controlling an imine reaction, which uses normal-phase liquid chromatography to detect the content of components in the reaction solution during the reaction that generates imine compounds;

[0006] The components in the reaction solution include 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza and its racemic mixture;

[0007] The detection conditions of the normal phase liquid chromatography method include: using a polysaccharide derivative normal phase coated chiral column, and using a mobile phase of a mixed solution of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid, wherein the volume ratio of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid is (75-85):(15-25):(0.05-0.15):(0.05-0.15).

[0008] In some embodiments of the present invention, the volume ratio of hexane, ethanol, ethylenediamine and trifluoroacetic acid is 80:20:0.1:0.1.

[0009] In some embodiments of the present invention, the normal-phase coated chiral chromatographic column of the polysaccharide derivative is a Chiralcel OD-H, 4.6*250mm, 5μm.

[0010] In some embodiments of the present invention, the detection conditions of the normal phase liquid chromatography method further include:

[0011] Injection volume: 5-15 μL;

[0012] Flow rate: 0.8-1.2 mL / min;

[0013] Column temperature: room temperature;

[0014] Detection wavelength: 240-300nm;

[0015] Detector: Ultraviolet detector.

[0016] In some embodiments of the present invention, the reaction solution further includes compound MIA08 and its racemic mixture, wherein the structural formula of compound MIA08 is as follows:

[0017]

[0018] In some embodiments of the present invention, the detection using normal-phase liquid chromatography includes the following steps:

[0019] Preparation of test solution: Take the test sample, add diluent to dissolve it, and obtain the test solution;

[0020] Preparation of positioning solution: Take 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza standard, dissolve it in diluent to obtain positioning solution;

[0021] The test solution and the positioning solution were tested under the aforementioned detection conditions, and the chromatograms were recorded respectively. If the chromatogram of the test solution contained a chromatographic peak corresponding to the retention time of the racemic substance in the chromatogram of the positioning solution, the area was normalized.

[0022] In some embodiments of the present invention, the diluent has the same composition as the mobile phase.

[0023] In some embodiments of the present invention, each 1 mL of the test solution contains 0.4-0.6 mg of the test sample.

[0024] In some embodiments of the present invention, each 1 mL of the positioning solution contains 0.4-0.6 mg of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza-standard.

[0025] A second aspect of the present invention provides the application of the normal-phase detection method described in the first aspect of the present invention in the preparation of mianserin hydrochloride.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0027] 1. This invention employs normal-phase liquid chromatography for detection, and uses specific mobile phase components and ratios to improve separation efficiency and target peak response. This enables the effective separation of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza and its racemic mixture, as well as compound MIA08 and its racemic mixture, in the reaction solution. Detection is performed using a UV detector, with retention time for qualitative analysis and peak area for quantitative analysis, allowing for accurate and reliable monitoring of the reaction process.

[0028] 2. The normal phase detection method of the present invention has the advantages of being simple, accurate, fast and reliable, thereby effectively controlling the quality of mianserin hydrochloride. Attached Figure Description

[0029] Figure 1 This is a liquid chromatogram of the positioning solution of Example 1 of the present invention;

[0030] Figure 2 This is a liquid chromatogram of the test solution of Example 1 of the present invention;

[0031] Figure 3 This is a liquid chromatogram of the test solution in Example 2 of the present invention;

[0032] Figure 4 This is a liquid chromatogram of the test solution in Example 3 of the present invention;

[0033] Figure 5 This is a superimposed liquid chromatogram of the positioning solution and the test solution of Comparative Example 1 of the present invention;

[0034] Figure 6 This is a superimposed liquid chromatogram of the positioning solution and the test solution of Comparative Example 2 of the present invention;

[0035] Figure 7This is a superimposed liquid chromatogram of the positioning solution and the test solution of Comparative Example 3 of the present invention. Detailed Implementation

[0036] To enable those skilled in the art to more clearly understand the technical solutions described in this invention, the following embodiments are provided for illustration. It should be noted that the following embodiments do not constitute a limitation on the scope of protection claimed by this invention.

[0037] Unless otherwise specified, the raw materials, reagents or devices used in the following examples are available from conventional commercial sources or can be obtained by existing known methods.

[0038] Three batches of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza (denoted as MIA07) reaction solutions reacted for 2 hours at different times were taken as samples 1-3. They were detected according to the detection methods of Examples 1-3, and quantification was performed using the area normalization method.

[0039] Example 1

[0040] A normal-phase detection method for the control of an imine reaction, employing normal-phase liquid chromatography, includes the following steps:

[0041] 1) Preparation of test solution: Take an appropriate amount of sample 1, weigh it accurately, and dilute it with diluent to prepare a solution containing 0.5 mg of sample 1 per 1 mL, which is used as the test solution;

[0042] 2) Preparation of MIA07 positioning solution: Take an appropriate amount of MIA07 standard, accurately weigh it, and dilute it with diluent to prepare a solution containing 0.5 mg of MIA07 standard per 1 mL, which is used as the MIA07 positioning solution;

[0043] 3) Accurately measure 10 μL of the positioning solution, inject it into the liquid chromatograph, and record the chromatogram;

[0044] 4) Accurately measure 10 μL of the test solution and inject it into the liquid chromatograph, and record the chromatogram; if there is a chromatographic peak in the chromatogram of the test solution that corresponds to the retention time of the racemic mixture in the positioning solution, calculate the area using the normalization method.

[0045] The detection conditions for the liquid chromatograph in this embodiment are set as follows:

[0046] Chromatographic column: Chiralcel OD-H (4.6*250mm, 5μm);

[0047] Injection volume: 10 μL;

[0048] Flow rate: 1.0 mL / min;

[0049] Column temperature: room temperature;

[0050] Detection wavelength: 257nm;

[0051] Mobile phase (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 80:20:0.1:0.1;

[0052] Diluent (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 80:20:0.1:0.1;

[0053] Detector: Ultraviolet detection.

[0054] The localization spectrum of MIA07 in this embodiment is as follows: Figure 1 As shown, the retention time of 6.325 min is MIA07, and the retention time of 9.292 min is the racemic form of MIA07; the spectrum of sample 1 is shown below. Figure 2 As shown, the retention time of 6.342 min is MIA07, the retention time of 9.334 min is the racemic form of MIA07, the retention time of 14.251 min is MIA08, and the retention time of 19.901 min is the racemic form of MIA08.

[0055] Example 2

[0056] A normal-phase detection method for the control of an imine reaction, employing normal-phase liquid chromatography, includes the following steps:

[0057] 1) Preparation of test solution: Take an appropriate amount of sample 2, weigh it accurately, and dilute it with diluent to prepare a solution containing 0.5 mg of sample 1 per 1 mL, which is used as the test solution;

[0058] 2) Preparation of MIA07 positioning solution: Take an appropriate amount of MIA07 standard, accurately weigh it, and dilute it with diluent to prepare a solution containing 0.5 mg of MIA07 standard per 1 mL, which is used as the MIA07 positioning solution;

[0059] 3) Accurately measure 10 μL of the positioning solution, inject it into the liquid chromatograph, and record the chromatogram;

[0060] 4) Accurately measure 10 μL of the test solution and inject it into the liquid chromatograph, and record the chromatogram; if there is a chromatographic peak in the chromatogram of the test solution that corresponds to the retention time of the racemic mixture in the positioning solution, calculate the area using the normalization method.

[0061] The detection conditions for the liquid chromatograph in this embodiment are set as follows:

[0062] Chromatographic column: Chiralcel OD-H (4.6*250mm, 5μm);

[0063] Injection volume: 10 μL;

[0064] Flow rate: 1.0 mL / min;

[0065] Column temperature: room temperature;

[0066] Detection wavelength: 257nm;

[0067] Mobile phase (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 75:25:0.05:0.15;

[0068] Diluent (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 75:25:0.05:0.15;

[0069] Detector: Ultraviolet detection.

[0070] The spectrum of sample 2 in this embodiment is as follows: Figure 3 As shown, the retention time of 6.317 min is MIA07, the retention time of 9.284 min is the racemic form of MIA07, the retention time of 14.101 min is MIA08, and the retention time of 19.734 min is the racemic form of MIA08.

[0071] Example 3

[0072] A normal-phase detection method for the control of an imine reaction, employing normal-phase liquid chromatography, includes the following steps:

[0073] 1) Preparation of test solution: Take an appropriate amount of sample 3, weigh it accurately, and dilute it with diluent to prepare a solution containing 0.5 mg of sample 1 per 1 mL, which is used as the test solution;

[0074] 2) Preparation of MIA07 positioning solution: Take an appropriate amount of MIA07 standard, accurately weigh it, and dilute it with diluent to prepare a solution containing 0.5 mg of MIA07 standard per 1 mL, which is used as the MIA07 positioning solution;

[0075] 3) Accurately measure 10 μL of the positioning solution, inject it into the liquid chromatograph, and record the chromatogram;

[0076] 4) Accurately measure 10 μL of the test solution and inject it into the liquid chromatograph, and record the chromatogram; if there is a chromatographic peak in the chromatogram of the test solution that corresponds to the retention time of the racemic mixture in the positioning solution, calculate the area using the normalization method.

[0077] The detection conditions for the liquid chromatograph in this embodiment are set as follows:

[0078] Chromatographic column: Chiralcel OD-H (4.6*250mm, 5μm);

[0079] Injection volume: 10 μL;

[0080] Flow rate: 1.0 mL / min;

[0081] Column temperature: room temperature;

[0082] Detection wavelength: 257nm;

[0083] Mobile phase (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 85:15:0.15:0.05;

[0084] Diluent (volume ratio): n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 85:15:0.15:0.05;

[0085] Detector: Ultraviolet detection.

[0086] The spectrum of sample 3 in this embodiment is as follows: Figure 4 As shown, the retention time of 6.326 min is MIA07, the retention time of 9.292 min is the racemic form of MIA07, the retention time of 14.067 min is MIA08, and the retention time of 19.701 min is the racemic form of MIA08.

[0087] The results of the area normalization method used in Examples 1-3 are shown in Table 1.

[0088] Table 1

[0089] batch number MIA07 MIA07 racemic mixture MIA08 MIA08 racemic mixture Sample 1 41.88％ 41.32％ 8.03％ 6.65％ Sample 2 37.49％ 36.84％ 12.02％ 10.64％ Sample 3 40.21％ 39.56％ 9.62％ 7.70％

[0090] from Figure 2-4 As shown in Table 1, the detection methods of Examples 1-3 can effectively separate 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza, its racemic mixture, and the imine compound MIA08 generated during the reaction process, its racemic mixture.

[0091] Comparative Example 1

[0092] The only difference between Comparative Example 1 and Example 1 is that the mobile phase used in Comparative Example 1 is a solution of n-hexane and ethanol mixed in a volume ratio of 70:30, and the diluent and mobile phase are the same.

[0093] The spectrum of the localization solution (black and white lines) of Comparative Example 1 and the spectrum of the test sample solution are superimposed as follows: Figure 5 As shown, only one peak appears for MIA07, and the separation from MIA08 is poor. MIA07, MIA08 and their racemic mixtures cannot be separated.

[0094] Comparative Example 2

[0095] The only difference between Comparative Example 2 and Example 1 is that the mobile phase used in Comparative Example 2 is a solution of n-hexane and ethanol mixed in a volume ratio of 80:20, and the diluent and mobile phase are the same.

[0096] The spectrum of the localization solution (black and white lines) of Comparative Example 2 and the spectrum of the test sample solution are superimposed as follows: Figure 6 As shown, the separation of MIA07 and MIA08 meets the requirements, but MIA07, MIA08 and their racemates cannot be separated, and only two main peaks are shown in the spectrum.

[0097] Comparative Example 3

[0098] The only difference between Comparative Example 3 and Example 1 is that the mobile phase used in Comparative Example 3 is a solution of n-hexane, ethanol and ethylenediamine mixed in a volume ratio of 80:20:0.1, and the diluent used is a solution of n-hexane and ethanol mixed in a volume ratio of 80:20.

[0099] The spectrum of the localization solution (black and white lines) of Comparative Example 3 and the spectrum of the test sample solution are superimposed as follows: Figure 7 As shown, the separation of MIA07 and MIA08 and their corresponding racemates meets the requirements, but the response of MIA07 and its racemate is low; for the same concentration of test sample, the peak height of MIA07 and its racemate does not exceed 25, which is about half of the peak height under the conditions of Example 1 (n-hexane: ethanol: ethylenediamine: trifluoroacetic acid = 80:20:0.1:0.1).

[0100] The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.

Claims

1. A normal-phase detection method for the intermediate control of an imine reaction, characterized in that, The content of components in the reaction solution during the reaction process that generates imine compounds was determined by normal phase liquid chromatography. The components in the reaction solution include 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza and its racemic mixture; The detection conditions of the normal phase liquid chromatography method include: using a polysaccharide derivative normal phase coated chiral column, and using a mobile phase of a mixed solution of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid, wherein the volume ratio of n-hexane, ethanol, ethylenediamine and trifluoroacetic acid is (75-85):(15-25):(0.05-0.15):(0.05-0.15).

2. The positive phase detection method according to claim 1, characterized in that, The volume ratio of hexane, ethanol, ethylenediamine, and trifluoroacetic acid is 80:20:0.1:0.

1.

3. The positive phase detection method according to claim 1, characterized in that, The normal-phase coated chiral chromatographic column for the polysaccharide derivative was a Chiralcel OD-H, 4.6*250mm, 5μm.

4. The positive phase detection method according to claim 1, characterized in that, The detection conditions for the normal phase liquid chromatography method also include: Injection volume: 5-15 μL; Flow rate: 0.8-1.2 mL / min; Column temperature: room temperature; Detection wavelength: 240-300nm; Detector: Ultraviolet detector.

5. The positive phase detection method according to claim 1, characterized in that, The reaction solution also includes compound MIA08 and its racemic mixture. The structural formula of compound MIA08 is as follows:

6. The positive phase detection method according to claim 1, characterized in that, Includes the following steps: Preparation of test solution: Take the test sample, add diluent to dissolve it, and obtain the test solution; Preparation of positioning solution: Take 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza standard, dissolve it in diluent to obtain positioning solution; The test solution and the positioning solution were tested under the aforementioned detection conditions, and the chromatograms were recorded respectively. If the chromatogram of the test solution contained a chromatographic peak corresponding to the retention time of the racemic substance in the chromatogram of the positioning solution, the area was normalized.

7. The positive phase detection method according to claim 6, characterized in that, The diluent has the same composition as the mobile phase.

8. The positive phase detection method according to claim 6, characterized in that, Each 1 mL of the test solution contains 0.4-0.6 mg of the test sample.

9. The positive phase detection method according to claim 6, characterized in that, Each 1 mL of the positioning solution contains 0.4-0.6 mg of 1,2,3,4,10,14B-hexahydrodibenzo[C,F]pyrazino[1,2-A]aza-standard.

10. The application of the normal-phase detection method according to any one of claims 1-9 in the preparation of mianserin hydrochloride.

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