A quality detection method for suvorexan intermediates

Abstract

The present invention relates to the field of drug analysis and quality control technology, and is specifically related to a quality detection method for a suvorexan intermediate. The quality detection method of the present invention comprises the following steps: step 1, dissolving the test sample of (R)-1-benzyl-5-methyl-1,4-diazacyclic with a diluent, and preparing a test solution; step 2, using high performance liquid chromatography to detect (R)-1-benzyl-5-methyl-1,4-diazacyclic and / or (S)-1-benzyl-5-methyl-1,4-diazacyclic in the test solution, wherein the chromatographic conditions are: the stationary phase adopts a polysaccharide derivative covalently bonded chiral chromatographic column, and the mobile phase adopts a mixed solution of water, acetonitrile and diethylamine. The present invention realizes the separation of IN5 and its enantiomeric impurity C, accurately determines the impurity C content, and the measurement result is stable and reliable, with strong specificity and good application prospects.

Description

Technical Field

[0001] The present invention relates to the technical field of drug analysis and quality control, and specifically to a quality detection method for a suvorexant intermediate. Background Art

[0002] Suvorexant is a new drug for the treatment of sleep difficulties approved by the US FDA in 2014. Its original manufacturer is Merck Pharmaceuticals. Suvorexant is an orexin receptor antagonist that can affect the signal transduction of the orexin pathway; orexin is a small molecule neuropeptide that is related to the regulation of the sleep-wake cycle. Suvorexant can be used clinically to treat chronic insomnia.

[0003] The chemical name of Suvorexant is 5-chloro-2-[(5R)-5-methyl-4-[5-methyl-2-(2H-1,2,3-triazol-2-yl)benzoyl]-1,4-diazepan-1-yl]-1,3-benzoxazole, and its molecular formula is C 23 H 23 ClN6O2, its structural formula is as follows:

[0004]

[0005] IN5 is a key intermediate in the synthesis of Suvorexan API. The chemical name of IN5 is (R)-1-benzyl-5-methyl-1,4-diazoheterocycle. The chemical structure of IN5 is as follows:

[0006]

[0007] In order to ensure the product quality of suvorexan API or raw materials, the content of IN5 in suvorexan API or raw materials is required to be strictly controlled during drug development and drug registration. In addition, IN5 contains a certain number of enantiomers, the chemical name of which is (S)-1-benzyl-5-methyl-1,4-diazacyclopentane (also referred to as enantiomeric impurity, enantiomeric impurity C or impurity C in this article).

[0008] The basic information of impurity C is shown in the table below:

[0009]

[0010]

[0011] The synthetic process route of svoraxant shows that impurity C and IN5 participate in subsequent reactions. IN5, as the main reaction material, will eventually produce svoraxant API. Impurity C in IN5 will also participate in the reaction, thereby deriving a series of impurities with structures similar to impurity C. These impurities can cause the isomers of svoraxant API to exceed the limit requirements for raw materials, making the quality of svoraxant API unqualified and unable to be registered as a raw material.

[0012] Therefore, it is urgent to find a quality control method that can accurately determine impurity C so as to control the content and limit of impurity C in advance in IN5. The improvement of IN5 purity also effectively reduces the purification process of suvorexan API, thereby reducing the direct production cost of suvorexan API or API. At the same time, the effective control of impurity C also indirectly controls the product quality of suvorexan API or API, making the product quality of suvorexan API more stable, measurable and controllable.

[0013] However, the prior art does not show that IN5 and its enantiomeric impurity C have a significant impact on the quality of suvorexan API or raw materials. At the same time, since impurity C is very similar to IN5 in structure, it is difficult to separate and detect impurity C in IN5. There is no detection method for suvorexan intermediate IN5 and its enantiomeric impurity C in the currently publicly reported technology. Summary of the invention

[0014] In view of the problems of the prior art, the present invention provides a quality detection method for a suvorexan intermediate, which aims to realize a high performance liquid chromatography analysis method for a suvorexan intermediate IN5 and an enantiomeric impurity C thereof, so as to effectively control the quality of the suvorexan intermediate IN5, and further effectively control the quality of the suvorexan API or raw material drug; meanwhile, the present invention also has certain guiding significance for the research on the synthesis process of the suvorexan API or raw material drug.

[0015] A quality detection method for a suvorexan intermediate comprises the following steps:

[0016] Step 1, dissolving a test sample of (R)-1-benzyl-5-methyl-1,4-diazacyclic ring in a diluent to prepare a test sample solution;

[0017] Step 2, using high performance liquid chromatography to detect (R)-1-benzyl-5-methyl-1,4-diazacyclopentane and / or (S)-1-benzyl-5-methyl-1,4-diazacyclopentane in the test solution,

[0018] The chromatographic conditions are as follows: a polysaccharide derivative covalently bonded chiral chromatographic column is used as the stationary phase, a mixed solution of water, acetonitrile and diethylamine is used as the mobile phase, and the volume ratio of water, acetonitrile and diethylamine in the mobile phase is 900-700:100-300:0.1-2.

[0019] Preferably, the diluent is selected from 0.5-2 mg / ml 4-chloro-7-nitrobenzofuran-acetonitrile solution.

[0020] Preferably, the diluent is selected from 1.0 mg / ml 4-chloro-7-nitrobenzofuran-acetonitrile solution;

[0021] Preferably, in the test solution, the dosage ratio of the test sample to the diluent is 1.0-5.0 mg:1 ml.

[0022] Preferably, the volume ratio of water, acetonitrile and diethylamine in the mobile phase is 900:100:1.

[0023] Preferably, the filler of the polysaccharide derivative covalently bonded chiral chromatographic column is: silica gel with amylose-tris(3,5-xylylcarbamate) covalently bonded on the surface.

[0024] Preferably, the chromatographic conditions further include: the elution procedure is isocratic elution for 5 to 30 minutes.

[0025] Preferably, the chromatographic conditions also include: detection wavelength of 190 to 800 nm; flow rate of 0.5 to 1.0 ml / min; column temperature of 25 to 35° C.; injection volume of 10 to 30 μl.

[0026] Preferably, step 1 further comprises: taking reference substances of (R)-1-benzyl-5-methyl-1,4-diazacyclopentane and (S)-1-benzyl-5-methyl-1,4-diazacyclopentane, dissolving them in a diluent to prepare a system suitability solution; the concentration of (R)-1-benzyl-5-methyl-1,4-diazacyclopentane in the system suitability solution is 1.0 to 5.0 mg / ml, and the concentration of (S)-1-benzyl-5-methyl-1,4-diazacyclopentane is 5 to 50 μg / ml;

[0027] Step 2 also includes: using high performance liquid chromatography to detect the system suitability solution, comparing the chromatograms of the test solution and the system suitability solution, and determining whether the test solution contains (S)-1-benzyl-5-methyl-1,4-diazacyclic.

[0028] Preferably, the method further comprises the following steps:

[0029] Step 3, calculate the content of (S)-1-benzyl-5-methyl-1,4-diazacyclic in the test sample, and the calculation formula is as follows:

[0030] Xi%=Ai / A×100%

[0031] Wherein, Xi% is the content of (S)-1-benzyl-5-methyl-1,4-diazacyclopentane in the test solution, Ai is the peak area of ​​(S)-1-benzyl-5-methyl-1,4-diazacyclopentane in the chromatogram of the test solution, and A is the sum of all peak areas in the chromatogram of the test solution.

[0032] The present invention proposes for the first time a quality control method for the intermediate IN5 of suvorexan. Through optimized high-performance liquid chromatography conditions, the separation of IN5 and its enantiomer impurity C is achieved, and the content of impurity C is accurately determined. The measurement results are stable and reliable with strong specificity. The methodological validation results show that the present invention meets the requirements of the 2020 edition of the "Chinese Pharmacopoeia" for analytical methodology, thereby effectively controlling the quality of suvorexan intermediates and improving the quality of suvorexan raw materials. At the same time, the synthesis process of suvorexan intermediates and the synthesis process of suvorexan raw materials can be effectively monitored.

[0033] Obviously, according to the above contents of the present invention, in accordance with common technical knowledge and customary means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0034] The above contents of the present invention are further described in detail below through specific implementation methods in the form of embodiments. However, this should not be understood as the scope of the above subject matter of the present invention being limited to the following examples. All technologies realized based on the above contents of the present invention belong to the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 It is the HPLC chart of the test solution in Example 1.

[0036] Figure 2 It is the HPLC chart of the test solution in Example 2.

[0037] Figure 3 It is a typical HPLC profile of the comparative system suitability solution.

[0038] Figure 4 is a typical HPLC profile of a system suitability solution. DETAILED DESCRIPTION

[0039] The reagents and materials used in the following examples and experimental examples are all commercially available unless otherwise specified.

[0040] (1) Materials

[0041] IN5 (provided by Chengdu Weibang Pharmaceutical Co., Ltd., batch number 221201, 221202);

[0042] Impurity C reference substance (provided by Chengdu Weibang Pharmaceutical Co., Ltd., batch number MS203423-221001).

[0043] (2) Main instruments

[0044] High performance liquid chromatograph: Shimadzu LC-20AT, chromatographic column: CHIRALCEL IA, 4.6×250 mm, 5 μm.

[0045] Example 1: Determination of the content of impurity C in the intermediate IN5 of Suvorexan

[0046] (1) Preparation of test solution: Accurately weigh IN5 (batch number 221201) and dissolve it in 1.0 mg / ml 4-chloro-7-nitrobenzofuran (NBD-Cl)-acetonitrile solution to prepare a 2.0 mg / ml test solution.

[0047] (2) Preparation of system suitability solution: Accurately weigh the impurity C reference substance and dissolve it in 1.0 mg / ml 4-chloro-7-nitrobenzofuran (NBD-Cl)-acetonitrile solution to prepare a solution with a concentration of 1.0 mg / ml as the impurity C reference substance stock solution. Accurately weigh 20.00 mg of IN5 and accurately measure 0.1 mL of the impurity C stock solution, place them in a 10 mL volumetric flask, dissolve it with a diluent, shake well, and make up to volume to prepare the system suitability solution.

[0048] (3) Chromatographic condition setting

[0049] Chromatographic column: CHIRALCEL IA, specification 4.6×250mm, 5μm;

[0050] Detection wavelength: 484nm;

[0051] Flow rate: 1.0 ml / min;

[0052] Column temperature: 30°C;

[0053] Injection volume: 20 μl;

[0054] Mobile phase: water-acetonitrile-diethylamine = (80%:20%:0.1%, v / v / v);

[0055] Gradient elution program: isocratic elution for 15 min.

[0056] (4) Determination

[0057] Pipette 10 μl of the test solution and system suitability solution into the chromatograph, record the chromatogram, and record the peak area A of the impurity C in the test sample. i And the sum of all peak areas A in the test solution, the content of impurity C in the test sample is calculated by area normalization method. The calculation formula is as follows:

[0058] Xi%=Ai / A×100%

[0059] Among them, X i %: the content of impurity C in the test sample,

[0060] A i : Peak area of ​​impurity C in the test sample,

[0061] A: The sum of all peak areas in the test solution. The HPLC spectrum of the test solution is shown in Figure 1 .

[0062] Calculation results: The impurity C content of IN5 sample batch number 221001 is: impurity C = 0.14%.

[0063] Example 2: Determination of the content of impurity C in the intermediate IN5 of Suvorexan

[0064] (1) Preparation of test solution: Accurately weigh IN5 (batch number 221202) and dissolve it in 1.0 mg / ml 4-chloro-7-nitrobenzofuran (NBD-Cl)-acetonitrile solution to prepare a 2.0 mg / ml test solution.

[0065] (2) Preparation of system suitability solution: Accurately weigh the impurity C reference substance and dissolve it in 1.0 mg / ml 4-chloro-7-nitrobenzofuran (NBD-Cl)-acetonitrile solution to prepare a solution with a concentration of 1.0 mg / ml as the impurity C reference substance stock solution. Accurately weigh 20.00 mg of IN5 and accurately measure 0.1 mL of the impurity C stock solution, place them in a 10 mL volumetric flask, dissolve it with a diluent, shake well, and make up to volume to prepare the system suitability solution.

[0066] (3) Chromatographic condition setting

[0067] Chromatographic column: CHIRALCEL IA, specification 4.6×250mm, 5μm;

[0068] Detection wavelength: 484nm;

[0069] Flow rate: 1.0 ml / min;

[0070] Column temperature: 30°C;

[0071] Injection volume: 20 μl;

[0072] Mobile phase: water-acetonitrile-diethylamine = (80%:20%:0.1%, v / v / v);

[0073] Gradient elution program: isocratic elution for 15 min.

[0074] (4) Determination

[0075] Pipette 10 μl of the test solution and system suitability solution into the chromatograph, record the chromatogram, and record the peak area A of the impurity C in the test sample. i And the sum of all peak areas A in the test solution, the content of impurity C in the test sample is calculated by area normalization method. The calculation formula is as follows:

[0076] Xi%=Ai / A×100%

[0077] Among them, X i %: the content of impurity C in the test sample,

[0078] A i : Peak area of ​​impurity C in the test sample,

[0079] A: The sum of all peak areas in the test solution. The HPLC spectrum of the test solution is shown in Figure 2 .

[0080] Calculation results: The impurity C content of IN5 sample batch number 221002 is: Impurity C = ND.

[0081] The technical solution of the present invention is further described below through experiments. In the following experimental examples, the HPLC detection steps or chromatographic conditions not specifically described are performed according to the description of Example 1.

[0082] Experimental Example 1: Chromatographic Condition Comparison Experiment

[0083] This experimental example provides a comparative experiment of different chromatographic conditions. The specific conditions and experimental results are as follows:

[0084] (1) Preparation of system suitability solution: Accurately weigh 20.00 mg of IN5 and 20.00 mg of impurity C reference substance, place in a 10 ml volumetric flask, dissolve with diluent, shake well, and make up to volume to prepare the system suitability solution.

[0085] (2) Chromatographic condition setting

[0086] Chromatographic column: CHIRALCEL IA, specification 4.6×250mm, 5μm;

[0087] Detection wavelength: 200nm;

[0088] Flow rate: 1.0 ml / min;

[0089] Column temperature: 30°C;

[0090] Injection volume: 5 μl;

[0091] Mobile phase: n-hexane-ethanol-diethylamine = (1000ml:10ml:1ml, v / v / v);

[0092] Gradient elution program: isocratic elution for 19 min.

[0093] (3) Determination

[0094] Pipette 5 μl of the system suitability solution and inject it into the chromatograph, record the chromatogram, and record the peak area A of the impurity C in the test sample. i And the sum of all peak areas A in the test solution, the content of impurity C in the test sample is calculated by area normalization method. The calculation formula is as follows:

[0095] Xi%=Ai / A×100%

[0096] Among them, X i %: Content of impurity C in the test sample.

[0097] A i : Peak area of ​​impurity C in the test sample.

[0098] A: The sum of all peak areas in the test solution. The HPLC spectrum of the test solution is shown in Figure 2 .

[0099] Results: The separation degree between IN5 and impurity C in the system suitability solution was 0.77, but the tailing factor was 3.05 in order to achieve baseline separation.

[0100] Conclusion: The separation and peak shape are insufficient under this mobile phase. The compound has a large polarity. The peak shape becomes worse as the ethanol ratio is subsequently reduced. There is insufficient room for optimization of the normal phase chromatography method.

[0101] Experimental Example 2: System Applicability and Specificity

[0102] Instrument: High performance liquid chromatograph: Shimadzu LC-20AT, chromatographic column: CHIRALCEL IA chromatographic column, 4.6×250 mm, 5 μm, flow rate: 1.0 ml / min, chromatographic column temperature: 30°C, injection volume 20 ul, detector wavelength: 484 nm, mobile phase is water-acetonitrile-diethylamine = (800 ml:200 ml:1 ml, v / v / v), isocratic elution for 15 min.

[0103] (1) Blank solution (the following dilution): 1.0 mg / ml 4-chloro-7-nitrobenzofuran (NBD-Cl)-acetonitrile solution;

[0104] (2) Impurity C stock solution: Accurately weigh 10 mg of impurity C into a 10 ml volumetric flask, dilute the solution and make up to volume (impurity C: 1.0 mg / ml);

[0105] (3) System suitability solution: Accurately weigh 20 mg of IN5 into a 10 ml volumetric flask, accurately add 0.1 ml of impurity C stock solution, and dilute to volume with diluent (IN5 2.0 mg / ml, impurity C 10 μg / ml);

[0106] (4) Impurity C localization solution: Weigh 0.1 mg of impurity C and place it in a 10 ml volumetric flask, then add diluent to dissolve and make up the volume (impurity C: 10 μg / ml).

[0107] (5) Test solution: Accurately weigh 20 mg of IN5 into a 10 ml volumetric flask and add diluent solution to make up to volume.

[0108] Inject blank solution, system suitability solution, and test solution according to the chromatographic conditions, record the chromatographic process, and the results are shown in Table 1. The HPLC spectrum of the system suitability solution is shown in Figure 4 .

[0109] Table 1 System applicability and specificity

[0110]

[0111] Conclusion: The separation degree of this method is 3.55, the separation degree is good, and the impurity C peak has no interference, which shows that the system applicability and specificity of the present invention meet the analysis requirements.

[0112] Experimental Example 3: Limit of Detection and Limit of Quantitation

[0113] The experimental conditions, liquid chromatography method and solution preparation were the same as those in Experimental Example 1.

[0114] For known impurity C, the detection limit (LOD) and quantification limit (LOQ) are determined based on the signal-to-noise ratio. The impurity C stock solution with known concentration is diluted to a low concentration for injection. The detection signal-to-noise ratio S / N ≥ 8 is determined as the quantification limit, and S / N = 2 to 8 is determined as the detection limit. The test results are shown in Table 2.

[0115] Table 2 Limit of quantitation and detection limit

[0116]

[0117] Results: The quantitative limit of impurity C was 0.25 μg / ml, equivalent to 0.0125% of the sample concentration, and the detection limit was 10 μg / ml, equivalent to 0.50% of the sample concentration, which fully demonstrated that the method of the present invention had good sensitivity and met the requirements for drug-related analysis.

[0118] Experimental Example 4: Solution Stability

[0119] The experimental conditions, liquid chromatography method and solution preparation were the same as those in Experimental Example 1.

[0120] The system suitability solution was placed at room temperature (25°C) for 0, 2, 6, 12, 18, and 24 hours, and then injected in sequence. The solution stability results are shown in Table 3.

[0121] Table 3 Stability

[0122] time Impurity C% 0 0.461 2 0.469 6 0.469 12 0.464 18 0.471 24 0.471 RSD 0.87％

[0123] Conclusion: The peak area %RSD of impurity C in the solution at room temperature (25°C) within 24 hours is 0.87%, indicating that the solution of the present invention has good stability and meets the requirements of drug-related analysis.

[0124] The results of Experimental Examples 1-3 show that the method for determining IN5 impurity C provided by the present invention has good specificity, sensitivity and durability (good solution stability within 24 hours at room temperature of 25°C), and can be applied to the quality control of intermediate impurity C in the synthesis of bulk drugs. The quality analysis of suvorexan intermediates is summarized by the method of the present invention. In order to ensure the quality of suvorexan API prepared with suvorexan intermediates as raw materials, the content of impurity C in suvorexan intermediates should be controlled at 0.5%.

[0125] In summary, the present invention meets the requirements of the 2020 edition of the "Chinese Pharmacopoeia" for analytical methodology, and effectively controls the quality of the svoraxant intermediate IN5, thereby improving the quality of the svoraxant raw material. At the same time, it can effectively monitor the synthesis process of the intermediates of svoraxant and the synthesis process of the svoraxant raw material, which has a guiding role in the drug research and development of svoraxant.

Claims

1. A quality detection method for a suvorexan intermediate, characterized in that: The steps include: Step 1, dissolving a test sample containing (R)-1-benzyl-5-methyl-1,4-diazacyclic ring in a diluent to prepare a test sample solution; Step 2, using high performance liquid chromatography to detect (R)-1-benzyl-5-methyl-1,4-diazacyclopentane and (S)-1-benzyl-5-methyl-1,4-diazacyclopentane in the test solution, The chromatographic conditions are as follows: a polysaccharide derivative covalently bonded chiral chromatographic column is used as the stationary phase; the filler of the polysaccharide derivative covalently bonded chiral chromatographic column is silica gel with amylose-tris(3,5-xylylcarbamate) covalently bonded on the surface; a mixed solution of water, acetonitrile and diethylamine is used as the mobile phase; and the volume ratio of water, acetonitrile and diethylamine in the mobile phase is 900-700:100-300:0.1-2.

2. The quality inspection method according to claim 1, characterized in that: The diluent is a 0.5-2 mg / ml 4-chloro-7-nitrobenzofuran-acetonitrile solution.

3. The quality inspection method according to claim 2, characterized in that: The diluent is a 1.0 mg / ml 4-chloro-7-nitrobenzofuran-acetonitrile solution.

4. The quality inspection method according to claim 1, characterized in that: In the test solution, the ratio of the test sample to the diluent is 1.0-5.0 mg:1 ml.

5. The quality inspection method according to claim 1, characterized in that: The volume ratio of water, acetonitrile and diethylamine in the mobile phase is 900:100:

1.

6. The quality inspection method according to claim 1, characterized in that: The chromatographic conditions also include: the elution procedure is isocratic elution for 5 to 30 minutes.

7. The quality inspection method according to claim 1, characterized in that: The chromatographic conditions also include: a detection wavelength of 190 to 800 nm; a flow rate of 0.5 to 1.0 ml / min; a column temperature of 25 to 35° C.; and an injection volume of 10 to 30 μl.

8. The quality inspection method according to claim 1, characterized in that: Step 1 also includes: taking reference substances of (R)-1-benzyl-5-methyl-1,4-diazacyclopentane and (S)-1-benzyl-5-methyl-1,4-diazacyclopentane, dissolving them in a diluent to prepare a system suitability solution; the concentration of (R)-1-benzyl-5-methyl-1,4-diazacyclopentane in the system suitability solution is 1.0 to 5.0 mg / ml, and the concentration of (S)-1-benzyl-5-methyl-1,4-diazacyclopentane is 5 to 50 μg / ml; Step 2 also includes: using high performance liquid chromatography to detect the system suitability solution, comparing the chromatograms of the test solution and the system suitability solution, and determining whether the test solution contains (S)-1-benzyl-5-methyl-1,4-diazacyclic.

9. The quality inspection method according to claim 1, characterized in that: The following steps are also included: Step 3, calculate the content of (S)-1-benzyl-5-methyl-1,4-diazacyclic in the test sample, and the calculation formula is as follows: Xi %= Ai / A ×100% in, Xi % is the content of (S)-1-benzyl-5-methyl-1,4-diazacyclic in the test solution, Ai is the peak area of ​​(S)-1-benzyl-5-methyl-1,4-diazacyclopentane in the chromatogram of the test solution, A It is the sum of all peak areas in the chromatogram of the test solution.

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