Method for detecting trans-isomers in cis-perhydroisoindole and salts thereof

Abstract

The invention discloses a method for detecting trans-isomers in cis-perhydroisoindole and salts thereof, and belongs to the technical field of medicine and chemical detection. The method for detecting trans-isomers in cis-perhydroisoindole and salts thereof comprises the following steps: placing a sample to be detected in a vessel, adding a sodium hydroxide solution and a chloroform solution for extraction, and taking a chloroform layer to obtain a solution to be detected; measuring the solution to be measured by adopting a gas chromatograph; wherein the chromatographic conditions of the gas chromatograph are as follows: a chromatographic column is a capillary column taking 5% diphenyl-95% dimethyl polysiloxane as a stationary phase. The method has the advantages of strong specificity, rapidness, sensitivity, high accuracy and the like, and can reliably perform qualitative and quantitative analysis on trans-perhydroisoindole which is an impurity in cis-perhydroisoindole and salts thereof.

Description

Technical Field

[0001] The invention relates to a method for detecting trans isomers in cis-perhydroisoindole and its salt, belonging to the technical field of medicine and chemical detection. Background Art

[0002] Cis-perhydroisoindole, also known as cis-octahydroisoindole in Chinese; REL-(3AR,7AS)-octahydro-1H-isoindole; CAS number is 1470-99-1; the structural formula of cis-perhydroisoindole and its salts is as follows:

[0003] and its salts

[0004] Mitiglinide calcium was applied for use in controlling postprandial blood sugar in patients with type 2 diabetes in December 2002 and was launched on the market in April 2004. Mitiglinide calcium works by shutting down the ATP-dependent K + channels, causing Ca 2+ Influx of intracellular Ca 2+ The increase in concentration causes the extracellular insulin-containing vesicles to degranulate, thereby stimulating the secretion of insulin. Mitiglinide calcium is clinically used to treat type 2 diabetes, but it has a faster onset and a shorter half-life. It is beneficial for lowering the postprandial blood sugar of diabetic patients and can avoid hypoglycemia caused by continuous blood sugar reduction. It has obvious advantages over other traditional blood sugar-lowering drugs and can be used as a first-line diabetes treatment drug with broad market prospects.

[0005] Cis-perhydroisoindole is a key intermediate of mitiglinide calcium, and its trans isomer content directly affects the quality of mitiglinide calcium. Therefore, a detection method for trans-perhydroisoindole in cis-perhydroisoindole should be developed to control the quality of cis-perhydroisoindole; the structural formula of trans-perhydroisoindole is as follows:

[0006] and its enantiomers.

[0007] The prior art JP1998287648A discloses a method for detecting the trans isomer in cis-perhydroisoindole. However, in this method, the trans isomer and cis-perhydroisoindole have poor separation and cannot be effectively separated, so it is difficult to quantitatively detect. Summary of the invention

[0008] [Technical issues]

[0009] The prior art is aimed at detecting the trans isomer in cis-perhydroisoindole. However, the separation degree between the trans isomer and cis-perhydroisoindole is poor and cannot be effectively separated, so it is difficult to perform quantitative detection.

[0010] [Technical solution]

[0011] In view of the defects of the prior art, the purpose of the present invention is to provide a method for detecting trans isomers in cis-perhydroisoindole and its salts. The method uses gas chromatography to qualitatively and quantitatively analyze the trans isomers in cis-perhydroisoindole and its salts, and the methodological verification is carried out. The results show that the method has the advantages of strong specificity, rapidity, sensitivity, accuracy, etc., and can reliably perform qualitative and quantitative analysis on the impurity in cis-perhydroisoindole and its salts, namely, trans-perhydroisoindole.

[0012] In order to achieve the above purpose, the technical solutions provided are as follows:

[0013] The present invention provides a method for detecting trans isomers in cis-perhydroisoindole and its salts, the method comprising the following steps:

[0014] (1) Preparation of test solution

[0015] Place the sample to be tested in a container, add sodium hydroxide solution and chloroform solution for extraction, and take the chloroform layer to obtain the solution to be tested;

[0016] (2) Determination of trans isomers in cis-perhydroisoindole and its salts in the test solution

[0017] The solution to be tested obtained in step (1) is measured by a gas chromatograph, wherein the chromatographic column of the gas chromatograph is a capillary column with 5% diphenyl-95% dimethylpolysiloxane as the stationary phase.

[0018] In one embodiment, the cis-perhydroisoindole salt includes any one or more of cis-perhydroisoindole hydrochloride, cis-perhydroisoindole phosphate, cis-perhydroisoindole sulfate, and cis-perhydroisoindole methanesulfonate.

[0019] In one embodiment, the concentration of the sodium hydroxide in step (1) is 2 to 10 mol / L, preferably 6 mol / L.

[0020] In one embodiment, the ratio of the sample to be tested to the sodium hydroxide and chloroform solution in step (1) is: 80-120:1-10:10; mg:g:mL.

[0021] In one embodiment, the capillary column with 5% diphenyl-95% dimethylpolysiloxane as the stationary phase in step (2) has a specification of: 15m×0.25mm, 0.5μm.

[0022] In one embodiment, the chromatographic conditions of the gas chromatograph in step (2) are as follows: temperature program: starting temperature is 60-85°C, maintained for 5-15 minutes, heated to 100-120°C at a rate of 2-8°C per minute, heated to 200-270°C at a rate of 10-30°C per minute, and maintained for 10-30 minutes.

[0023] In one embodiment, the chromatographic conditions of the gas chromatograph in step (2) are as follows: temperature program: starting temperature is 75°C, maintained for 10 minutes, heated to 110°C at a rate of 5°C per minute, heated to 270°C at a rate of 20°C per minute, and maintained for 20 minutes.

[0024] In one embodiment, the gas chromatograph detection conditions in step (2) are: injection port temperature 200-350° C., FID detector temperature 200-350° C., carrier gas is nitrogen, and injection volume is 0.1-10 μl.

[0025] In one embodiment, the gas chromatograph detection conditions in step (2) are: injection port temperature: 300° C., FID detector temperature: 300° C., carrier gas: nitrogen, injection volume: 1 μl.

[0026] In one embodiment, when the test solution obtained in step (1) is measured by gas chromatograph in step (2), the separation degree between the cis-isomer and the trans-isomer in cis-perhydroisoindole and its salts is greater than 1.5, which can be used for the qualitative detection of the trans-isomer in cis-perhydroisoindole and its salts.

[0027] In one embodiment, in step (2) quantitative detection of the trans isomer of cis-perhydroisoindole in the test solution: the peak area of ​​the trans isomer of cis-perhydroisoindole measured in the gas chromatograph is substituted into the linear relationship model to obtain the content of the trans isomer of cis-perhydroisoindole in the test sample.

[0028] In one embodiment, the linear relationship model is established by preparing trans-perhydroisoindole and its salt into a series of standard solutions, then adding sodium hydroxide solution and chloroform solution for extraction, taking the chloroform layer to obtain the standard solution to be tested; using a gas chromatograph to measure the standard solution, with the concentration of different standard solutions as the horizontal coordinate and the corresponding peak area as the vertical coordinate, to construct a linear relationship model.

[0029] In one embodiment, the linear relationship model is: y=1046.9508x-2142.7641; R 2 =0.9957.

[0030] The present invention also provides an application of the method described above in monitoring the production quality of cis-perhydroisoindole and its salt.

[0031] Beneficial effects:

[0032] The method for detecting the trans isomer in cis-perhydroisoindole and its salt provided by the present invention can better realize the separation of cis-perhydroisoindole and trans-perhydroisoindole, making the separation degree greater than 1.5; and improves the theoretical plate number of the chromatographic peak of trans-perhydroisoindole, and can effectively detect the content of trans-perhydroisoindole, further ensuring the safety, effectiveness and controllability of clinical drug use. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 is the chromatogram of Example 1;

[0034] Figure 2 It is the chromatogram of Comparative Example 1;

[0035] Figure 3 This is a linear relationship model data diagram of Example 2;

[0036] Figure 4 The chromatogram of the chromatographic column SH-Rtx-624 30m×0.32mm×0.25μm of Example 3;

[0037] Figure 5 The chromatogram of the chromatographic column SH-1 30m×0.32mm×1.5μm of Example 3;

[0038] Figure 6 This is the chromatogram of the chromatographic column DB-WAX 30m×0.32mm×0.25μm of Example 3. DETAILED DESCRIPTION

[0039] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention. The following specific implementation methods further describe the present invention.

[0040] The raw material sources involved in the present invention are:

[0041] Table 1. Raw material information

[0042] name factory purity Cis-perhydroisoindole hydrochloride Shandong Boyuan Pharmaceutical Co., Ltd. 99.95％ Trans-perhydroisoindole hydrochloride Shenzhen Hengfeng Wanda Pharmaceutical Technology Co., Ltd. 99.54％

[0043] Preparation of trans-perhydroisoindole hydrochloride stock solution:

[0044] Accurately weigh 15 mg of trans-perhydroisoindole hydrochloride, place it in a separatory funnel, add 5 ml of 6 mol / L sodium hydroxide solution, and then add 100 ml of chloroform solution for extraction. Take the chloroform layer to obtain the stock solution of trans-perhydroisoindole hydrochloride.

[0045] Example 1

[0046] A qualitative detection method for trans isomers in cis-perhydroisoindole comprises the following steps:

[0047] (1) Preparation of test solution

[0048] Accurately weigh 100 mg of cis-perhydroisoindole hydrochloride and 1 ml of trans-perhydroisoindole hydrochloride stock solution, place in a separatory funnel, add 5 ml of 6 mol / L sodium hydroxide solution, and then add 10 ml of chloroform solution for extraction. Take the chloroform layer to obtain the test solution;

[0049] (2) Determination of the trans isomer of cis-perhydroisoindole in the test solution

[0050] The liquid to be tested is measured by gas chromatograph, wherein the chromatographic conditions of the gas chromatograph are:

[0051] Chromatographic column: capillary column with 5% diphenyl-95% dimethylpolysiloxane as stationary phase (Rtx-5 Amine 15m×0.25mm, 0.5μm);

[0052] Heating program: starting temperature is 75°C, maintained for 10 min, then increased to 110°C at a rate of 5°C per minute, then increased to 270°C at a rate of 20°C per minute, and maintained for 20 min;

[0053] Inlet temperature: 300°C; FID detector temperature: 300°C; carrier gas: nitrogen; linear velocity: 25 cm / s; split ratio: 10; injection volume: 1 μl.

[0054] The test results are shown in Table 2 and Figure 1 As shown:

[0055] Table 2. Separation of cis-perhydroisoindole and trans-isomers

[0056]

[0057] From Table 2 and Figure 1 and Figure 2 The results show that the separation degree of Example 1 is 5.973, which is significantly greater than 1.5, indicating that cis-perhydroisoindole and trans-perhydroisoindole can be completely separated and can be used for the qualitative detection of trans-isomers in cis-perhydroisoindole.

[0058] Comparative Example 1

[0059] A qualitative detection method for trans isomers in cis-perhydroisoindole comprises the following steps:

[0060] (1) Preparation of test solution

[0061] Accurately weigh 100 mg of cis-perhydroisoindole hydrochloride and 1 ml of trans-perhydroisoindole hydrochloride stock solution, place in a separatory funnel, add 5 ml of 6 mol / L sodium hydroxide solution, and then add 10 ml of chloroform solution for extraction. Take the chloroform layer to obtain the test solution;

[0062] (2) Determination of the trans isomer of cis-perhydroisoindole in the test solution

[0063] The liquid to be tested is measured by gas chromatograph, wherein the chromatographic conditions of the gas chromatograph are:

[0064] Chromatographic column: GL Science NEUTRAB OND-5 (0.25 mm × 60 m × 1.5 μm);

[0065] Heating program: starting temperature 100°C, maintain for 1 min, increase to 160°C at a rate of 10°C, maintain for 8 min, and then increase to 220°C at a rate of 10°C.

[0066] Inlet temperature: 150°C; FID detector: 300°C; carrier gas: helium; flow rate: 3 ml / min; split ratio: 33:1; injection volume: 1 μl.

[0067] Example 2

[0068] A method for quantitatively detecting trans isomers in cis-perhydroisoindole comprises the following steps:

[0069] (1) Preparation of standard test solution

[0070] Linearity - 200%: Take 7.5 mg of trans-perhydroisoindole hydrochloride, weigh accurately, place in a separatory funnel, accurately add 12.5 ml of 6 mol / L sodium hydroxide solution, then add 25 ml of chloroform solution for extraction, take 5 ml of the chloroform layer and place in a 50 ml volumetric flask, dilute to the mark with chloroform, and shake well;

[0071] Linearity -120%: Take 6 ml of linearity -200% solution and place it in a 10 ml volumetric flask, dilute to the mark with chloroform, and shake well;

[0072] Linearity - 100%: Take 5 ml of linearity - 200% solution and place it in a 10 ml volumetric flask, dilute to the mark with chloroform, and shake well;

[0073] Linearity -80%: Take 4 ml of linearity -200% solution and place it in a 10 ml volumetric flask, dilute to the mark with chloroform, and shake well;

[0074] Linearity -50%: Take 5 ml of linearity -200% solution and place it in a 20 ml volumetric flask, dilute to the mark with chloroform, and shake well;

[0075] (2) Determination of the trans isomer of cis-perhydroisoindole in the standard test solution

[0076] The liquid to be tested is measured by gas chromatograph, wherein the chromatographic conditions of the gas chromatograph are:

[0077] Chromatographic column: capillary column with 5% diphenyl-95% dimethylpolysiloxane as stationary phase (Rtx-5 Amine 15m×0.25mm, 0.5μm);

[0078] The test conditions were as follows: heating program: starting temperature at 75°C, maintained for 10 min, heated to 110°C at a rate of 5°C per minute, heated to 270°C at a rate of 20°C per minute, maintained for 20 min;

[0079] Inlet temperature: 300°C; FID detector temperature: 300°C; carrier gas: nitrogen; linear velocity: 25 cm / s; split ratio: 10; injection volume: 1 μl.

[0080] The quantitative relationship model was constructed with the concentration of trans-perhydroisoindole in the standard solution as the horizontal axis and the corresponding peak area as the vertical axis. 2 =0.9957; the results are shown in Table 3 and Figure 3 As shown:

[0081] Table 3. Trans-perhydroisoindole hydrochloride linear results

[0082]

[0083] (3) Determination of the trans isomer of cis-perhydroisoindole in the sample to be tested

[0084] The liquid to be tested is measured by gas chromatography, and the content of the trans isomer of the cis-perhydroisoindole in the liquid to be tested is calculated according to the quantitative relationship model constructed in step (2);

[0085] Among them, the samples to be tested are:

[0086] Linearity -200%: Take 7.5 mg of trans-perhydroisoindole hydrochloride, accurately weigh it, put it in a separatory funnel, accurately add 12.5 ml of 6 mol / L sodium hydroxide solution, and then add 25 ml of chloroform solution for extraction. Take 10 ml of the chloroform layer and put it in a 100 ml volumetric flask, dilute it to the scale with chloroform, and shake well.

[0087] Control solution: Take 5 ml of linear -200% solution and place it in a 10 ml volumetric flask, dilute to the mark with chloroform, and shake well.

[0088] Test solution: Take 100 mg of cis-perhydroisoindole hydrochloride, accurately weigh it, put it in a separatory funnel, accurately add 5 ml of 6 mol / L sodium hydroxide solution, then add 10 ml of chloroform solution for extraction, and take the chloroform layer to obtain.

[0089] For test +0.075% trans-perhydroisoindole hydrochloride solution: Take 100 mg of cis-perhydroisoindole hydrochloride, accurately weigh, place in a separatory funnel, accurately add 5 ml of 6 mol / L sodium hydroxide solution and 2.5 ml of linear -200% solution, then add 7.5 ml of chloroform solution for extraction, take the chloroform layer, and you are done.

[0090] For test +0.15% trans-perhydroisoindole hydrochloride solution: Take 100 mg of cis-perhydroisoindole hydrochloride, accurately weigh, place in a separatory funnel, accurately add 5 ml of 6 mol / L sodium hydroxide solution and 5 ml of linear -200% solution, then add 5 ml of chloroform solution for extraction, take the chloroform layer, and you are done.

[0091] For test +0.225% trans-perhydroisoindole hydrochloride solution: Take 100 mg of cis-perhydroisoindole hydrochloride, accurately weigh, place in a separatory funnel, accurately add 5 ml of 6 mol / L sodium hydroxide solution and 7.5 ml of linear -200% solution, then add 2.5 ml of chloroform solution for extraction, take the chloroform layer, and you are done.

[0092] The results are shown in Table 4:

[0093] Table 4. Recovery data

[0094]

[0095] Note: A 杂 represents the peak area of ​​trans-perhydroisoindole hydrochloride in the test+trans-perhydroisoindole hydrochloride solution;

[0096] A 对 represents the peak area of ​​trans-perhydroisoindole hydrochloride in the control solution;

[0097] m represents the sample amount of cis-perhydroisoindole hydrochloride in the test+trans-perhydroisoindole hydrochloride solution;

[0098] m 对 It represents the sample amount of trans-perhydroisoindole hydrochloride in the control solution.

[0099] Example 3

[0100] A qualitative detection method for trans isomers in cis-perhydroisoindole comprises the following steps:

[0101] (1) Preparation of test solution

[0102] Accurately weigh 100 mg of cis-perhydroisoindole hydrochloride and 1 ml of trans-perhydroisoindole hydrochloride stock solution, place in a separatory funnel, add 5 ml of 6 mol / L sodium hydroxide solution, and then add 10 ml of chloroform solution for extraction. Take the chloroform layer to obtain the test solution;

[0103] (2) Determination of the trans isomer of cis-perhydroisoindole in the test solution

[0104] The liquid to be tested is measured by gas chromatograph, wherein the chromatographic conditions of the gas chromatograph are:

[0105] Chromatographic column: as shown in Table 5:

[0106] Heating program: starting temperature is 75°C, maintained for 10 min, then increased to 110°C at a rate of 5°C per minute, then increased to 270°C at a rate of 20°C per minute, and maintained for 20 min;

[0107] Inlet temperature: 300°C; FID detector temperature: 300°C; carrier gas: nitrogen; linear velocity: 25 cm / s; split ratio: 10; injection volume: 1 μl.

[0108] The results are shown in Table 5 and Figure 4 , 5 , as shown in Figure 6.

[0109] Table 5. Column types

[0110]

[0111] Example 4

[0112] A method for detecting trans isomers in cis-perhydroisoindole comprises the following steps:

[0113] (1) Preparation of test solution

[0114] Accurately weigh 100 mg of cis-perhydroisoindole hydrochloride and 1 ml of trans-perhydroisoindole hydrochloride stock solution, place in a separatory funnel, add 5 ml of 6 mol / L sodium hydroxide solution, and then add 10 ml of chloroform solution for extraction. Take the chloroform layer to obtain the test solution;

[0115] (2) Determination of the trans isomer of cis-perhydroisoindole in the test solution

[0116] The liquid to be tested is measured by gas chromatograph, wherein the chromatographic conditions of the gas chromatograph are:

[0117] Chromatographic column: capillary column with 5% diphenyl-95% dimethylpolysiloxane as stationary phase (Rtx-5 Amine 15m×0.25mm, 0.5μm);

[0118] Heating program: Different heating programs are shown in Table 6;

[0119] Inlet temperature: 300°C; FID detector temperature: 300°C; carrier gas: nitrogen; linear velocity: 25 cm / s; split ratio: 10; injection volume: 1 μl.

[0120] The results are shown in the table below:

[0121] Table 6. Temperature program and resolution

[0122]

[0123] The embodiments provided above are not intended to limit the scope of the present invention, and the steps described are not intended to limit the execution order thereof. Those skilled in the art may make obvious improvements to the present invention in combination with existing common knowledge, which also fall within the scope of protection defined by the claims of the present invention.

Claims

1. A method for detecting trans isomers in cis-perhydroisoindole and its salts, characterized in that: The method comprises the following steps: (1) Preparation of test solution Place the sample to be tested in a container, add sodium hydroxide solution and chloroform solution for extraction, and take the chloroform layer to obtain the solution to be tested; (2) Determination of trans isomers in cis-perhydroisoindole and its salts in the test solution The solution to be tested obtained in step (1) is measured by a gas chromatograph, wherein the chromatographic column of the gas chromatograph is a capillary column with 5% diphenyl-95% dimethylpolysiloxane as the stationary phase.

2. The method according to claim 1, characterized in that The cis-perhydroisoindole salt includes one or more of cis-perhydroisoindole hydrochloride, cis-perhydroisoindole phosphate, cis-perhydroisoindole sulfate, and cis-perhydroisoindole methanesulfonate.

3. The method according to claim 1, characterized in that The ratio of the sample to be tested to the sodium hydroxide and chloroform solution in step (1) is: 80-120:1-10:10; mg:g:mL.

4. The method according to claim 1, characterized in that: The chromatographic conditions of the gas chromatograph in step (2) are as follows: heating program: starting temperature is 60-85°C, maintained for 5-15 minutes, heated to 100-120°C at a rate of 2-8°C per minute, heated to 200-270°C at a rate of 10-30°C per minute, and maintained for 10-30 minutes.

5. The method according to claim 1, characterized in that The chromatographic conditions of the gas chromatograph in step (2) are as follows: heating program: starting temperature is 75°C, maintained for 10 min, heated to 110°C at a rate of 5°C per minute, heated to 270°C at a rate of 20°C per minute, and maintained for 20 min.

6. The method according to claim 1, characterized in that The gas chromatograph detection conditions in step (2) are: injection port temperature 200-350° C., FID detector temperature 200-350° C., carrier gas nitrogen, and injection volume 0.1-10 μl.

7. The method according to claim 1, characterized in that The gas chromatograph detection conditions in step (2) are: injection port temperature: 300° C., FID detector temperature: 300° C., carrier gas: nitrogen; injection volume: 1 μl.

8. The method according to claim 1, characterized in that In step (2), in the qualitative detection of the trans isomers in the cis-perhydroisoindole and its salts in the test solution, the separation degree between the cis-structure and the trans isomers in the cis-perhydroisoindole and its salts in the test solution is greater than 1.

5.

9. The method according to claim 1, characterized in that: In step (2), quantitative detection of the trans isomer of cis-perhydroisoindole in the test solution: the peak area of ​​the trans isomer of cis-perhydroisoindole measured in the gas chromatograph is brought into the linear relationship model to obtain the content of the trans isomer of cis-perhydroisoindole in the test sample.

10. Use of the method according to any one of claims 1 to 9 in monitoring the production quality of cis-perhydroisoindole and its salts.

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