Synthesis method of aza-seven-membered ring nitrosamine impurity

Abstract

The invention relates to a synthetic method of an aza-seven-membered ring nitrosamine impurity, which comprises the following steps: S1, condensing a compound I and a compound II in a solvent 1 to obtain a solution A containing a compound III; s2, a reducing agent is added into the solution A containing the compound III obtained in the S1 for a reduction reaction, after the reaction is completed, aftertreatment is conducted, and a compound IV is obtained; s3, hydrolyzing the compound IV obtained in S2 in an aqueous solution of strong acid, filtering after hydrolysis is completed, adjusting the pH value to 6.5-7.2, adding a compound V, heating, refluxing, condensing, dehydrating and cyclizing, and obtaining a solution B containing a compound VI after reaction is completed; and S4, concentrating the solvent in the solution B containing the compound VI obtained in the step S3 under reduced pressure, adding a solvent 2, filtering, adding a nitrosation reagent, and after the reaction is finished, performing column chromatography purification to obtain the target impurity. At present, no report related to synthesis of the nitrosamine impurity exists in the prior art, and the nitrosamine impurity synthesis method lays a foundation for quality control of azelastine.

Description

Technical Field

[0001] The invention belongs to the technical field of medicine, and particularly relates to a method for synthesizing an aza-seven-membered ring nitrosamine impurity. Background Art

[0002] Aza-7-membered ring skeletons are widely present in natural products and have important applications in medicine. For example, galantamine, a natural product extracted from Galanthus amine (Galanthus galanthus), used to treat Alzheimer's disease, contains a seven-membered aza ring structure. Dibenzazepine seven-membered rings are the basic skeleton of psychotropic drugs (antidepressants and antiepileptics), such as imipramine. Azelastine hydrochloride, which contains a seven-membered aza ring structure, exhibits potent antihistamine pharmacological activity and has comprehensive anti-inflammatory properties.

[0003] Following the 2018 incident involving the carcinogenic nitrosamine impurity NDMA (nitrosodimethylamine) in sartan drugs, regulatory agencies such as the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) have consistently emphasized the need for quality control of nitrosamine impurities during the production of both active pharmaceutical ingredients and finished pharmaceuticals. They have repeatedly issued and updated relevant guidelines, instructing pharmaceutical companies and contract development and manufacturing organizations (CDMOs) to strengthen their management of nitrosamine impurities.

[0004] The FDA's regulatory requirements for nitrosamine impurities can be roughly divided into two periods. Initially, driven by the impact of NDMA contamination in drugs like sartans, the FDA focused primarily on the management of simple nitrosamine impurities. During this period, the FDA focused on small nitrosamine impurities resulting from reactions between reagents during drug production, or between reagents and impurities (NDMA contamination in sartans stems from the reaction between nitrite and dimethylamine impurities in the dimethylformamide solvent).

[0005] In the second phase, the FDA and EMA shifted their focus to so-called NDSRIs (nitrosamine-related impurities in the drug matrix). Unlike simple nitrosamine impurities, NDSRIs refer to drugs that participate in the formation of nitrosamine impurities. These drugs typically contain secondary amines (including tertiary amines, which can degrade to secondary amines) that provide amino groups for nitrosamine formation.

[0006] Nitrosamine impurities in drug molecules are receiving increasing attention. Chinese patent CN119569792A discloses a method for synthesizing 3'-N-demethyl-3'-N-nitroso-6-O-methylerythromycin and 3'-N-demethyl-3'-N-nitrosoerythromycin A(E)-9-oxime nitrosamine impurities, and CN118344274A discloses a method for preparing ivocacetyl nitrosamine impurities.

[0007] The nitrogen seven-membered ring structure contains a tertiary amine or secondary amine structure, which has the potential to generate nitrosamine impurities. Its structural formula is as follows: Azelastine (4-(4-chlorobenzyl)-2-(hexahydro-1-methyl-1H-azepine-4-yl)-1-(2H)-phthalazinone) contains a seven-membered nitrogen ring structure and has potent antihistamine pharmacological activity and comprehensive anti-inflammatory mediator effects. However, due to its tertiary amine structure, impurities shown in Formula I may be generated during the production and storage of raw materials and preparations.

[0008] The main production process is as follows: azelastine is oxidatively degraded to remove the methyl group to form demethylazelastine, and then nitrosated to form demethylazelastine nitrosamine impurities.

[0009] The European Medicines Agency (EMA) clearly lists the impurity represented by Formula I as an object that needs to be studied in its guidelines on nitrosamine impurities, and the Center for Drug Evaluation (CDE) of the National Medical Products Administration has the same requirement.

[0010] Currently, there is no literature report on the synthesis method of the nitrosamine impurity in the prior art. Based on the above background, this application is specially proposed. Summary of the Invention

[0011] The technical problem to be solved by the present invention is to provide a method for synthesizing an aza-seven-membered ring nitrosamine impurity.

[0012] In order to solve the above problems, the technical solution adopted by the present invention is: A method for synthesizing nitrogen-containing seven-membered ring nitrosamine impurities, which is synthesized via the following route: S1: Condensing compound I and compound II in solvent 1 to obtain solution A containing compound III; S2: adding a reducing agent to the solution A containing compound III obtained in S1 to carry out a reduction reaction. After the reaction is completed, the solution is post-treated to obtain compound IV; S3: hydrolyze compound IV obtained in S2 in an aqueous solution of a strong acid, filter after completion of the hydrolysis, adjust the pH to 6.5-7.2, add compound V, heat under reflux for condensation and dehydration cyclization, and complete the reaction to obtain solution B containing compound VI; S4: The solvent of solution B containing compound VI obtained in S3 is concentrated under reduced pressure, solvent 2 is added, and after filtration, a nitrosating agent is added. After the reaction is completed, the target impurity is purified by column chromatography to obtain the target impurity.

[0013] As a further improvement of the present invention, the molar ratio of compound I to compound II to compound V is 1:0.95-1.0:0.85-1.0.

[0014] As a further improvement of the present invention, solvent 1 is one of methanol, ethanol, water, a mixed solution of methanol and water, and a mixed solution of ethanol and water; solvent 2 is one of dichloromethane, methanol, ethanol, isopropanol, tetrahydrofuran, and acetonitrile; and the aqueous solution of the strong acid is an aqueous hydrochloric acid solution with a mass fraction of 36%–38%.

[0015] As a further improvement of the present invention, the amount of compound I and solvent 1 is in the following relationship: 0.1 mol: 80-120 ml; The relationship between the amount of compound I and the aqueous solution of strong acid is 0.1 mol: 120-150 ml; The usage ratio of compound I and solvent 2 is 0.1 mol: 160-240 ml.

[0016] As a further improvement of the present invention, the reducing agent in S2 is one of sodium cyanoborohydride, sodium borohydride or potassium borohydride; and the nitrosating agent in S4 is one of tert-butyl nitrite, sodium nitrite or potassium nitrite.

[0017] As a further improvement of the present invention, the reducing agent in S2 is sodium borohydride, and the nitrosating agent in S4 is tert-butyl nitrite.

[0018] As a further improvement of the present invention, the molar ratio of compound I to the reducing agent is 1:0.75-1.0; The molar ratio of compound I to the nitrating agent is 1:1.5-5.

[0019] As a further improvement of the present invention, the reaction temperature in S1 is 20-60°C; the adding temperature of the reducing agent in S2 is 0-10°C, and the reaction temperature is room temperature; and the reaction temperature in S4 is room temperature.

[0020] As a further improvement of the present invention, the reaction temperature in S1 is 30-40°C.

[0021] As a further improvement of the present invention, the pH in S2 is adjusted to 6.8-7.0.

[0022] As a further improvement of the present invention, water is added simultaneously with the addition of the reducing agent in S2. The post-treatment in S2 is to concentrate the reaction solution under reduced pressure until the volume of the solvent is reduced to 1-1.1 times the volume of solvent 1, and to add the same volume of ethyl acetate to the residual solution for extraction, and to concentrate the organic phase under reduced pressure to obtain compound IV. or when the reducing agent is added in S2, water is not added. The post-treatment in S2 is to concentrate the reaction solution under reduced pressure until the volume of the solvent is reduced to an amount equivalent to the volume of solvent 1, add equal volumes of water and ethyl acetate to the residue for extraction, and concentrate the organic phase under reduced pressure to obtain compound IV; The dosage of compound I and water is 0.1 mol: 80-120 ml.

[0023] As a further improvement of the present invention, the purification is carried out by silica gel column chromatography, using n-heptane-ethyl acetate in a volume ratio of 2:1 as the eluent.

[0024] The beneficial effect of adopting the above technical solution is that: this application proposes a method for synthesizing a nitrogen-containing seven-membered ring nitrosamine impurity. Currently, there is no relevant report on the synthesis of this nitrosamine impurity in the existing technology. The proposal of this application lays the foundation for the quality control of azelastine. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 It is a synthetic route diagram of the synthetic impurity of the present invention; Figure 2 is a mass spectrum of the impurity synthesized in Example 1 of the present invention; Figure 3 The impurities synthesized in Example 1 of the present invention 1 H-NMR spectrum; Figure 4 The impurities synthesized in Example 1 of the present invention 13 C-NMR spectrum; Figure 5 1 is an HPLC spectrum of the impurities measured in Example 1 of the present invention and a corresponding peak result table. DETAILED DESCRIPTION

[0026] In order to make the objectives, technical solutions and advantages of the present invention more clear, the invention is clearly and completely described below in conjunction with specific embodiments.

[0027] The examples of this application are synthesized using the following route, and compounds I to VII described in the examples are the corresponding compounds in this route.

[0028] Example 1 S1: Add 21.3 g of compound I and 13.6 g of compound II to 100 mL of methanol, stir and react at 30°C for 6 h. The reaction is complete when detected by TLC to obtain solution A containing compound III.

[0029] S2: Add 100 ml of water to solution A containing compound III obtained in S1, cool to 0°C, add 2.9 g of sodium borohydride in batches, warm to room temperature after addition, and continue stirring to react for 3 hours. TLC detection shows that the reaction is complete. The reaction solution is concentrated under reduced pressure at 40°C to a solvent of about the volume of methanol. 100 ml of ethyl acetate is added to the residual liquid for extraction. The organic phase is concentrated under reduced pressure at 40°C to dryness to obtain compound IV.

[0030] S3: 140 ml of 37% concentrated hydrochloric acid was added to compound IV, and the mixture was refluxed for 4 h. The reaction was completed by TLC. The mixture was filtered, and the pH was adjusted to 7.0 with 50% sodium hydroxide. 24.7 g of compound V was added, and the mixture was refluxed for 7 h. The reaction was completed by TLC to obtain solution B containing compound VI.

[0031] S4: Solution B containing compound VI obtained in S3 was concentrated to dryness under reduced pressure, and then 200 ml of dichloromethane was added. After filtering, 18 ml of tert-butyl nitrite was added and stirred at room temperature for 2 h. The reaction was completed after TLC detection. The solution was purified by silica gel column chromatography using n-heptane / ethyl acetate in a volume ratio of 2:1 as eluent to obtain 8.9 g of the target product. The purity was 98.16% as determined by HPLC.

[0032] The mass spectrum of the compound synthesized in this example is as follows Figure 2 As shown, 1 H-NMR spectrum Figure 3 As shown, 13 C-NMR spectrum Figure 4 As shown, the successful synthesis of the impurity was demonstrated.

[0033] Example 2 S1: Add 5.3 g of compound I and 3.4 g of compound II to 30 mL of ethanol, stir and react at 30°C for 6 h. The reaction is complete when detected by TLC to obtain solution A containing compound III.

[0034] S2: Add 30 ml of water to the solution A containing compound III obtained in S1, cool to 0°C, add 1.1 g of potassium borohydride in batches, warm to room temperature after addition, continue stirring and react for 3 hours, and detect the completion of the reaction by TLC. Concentrate the reaction solution under reduced pressure at 40°C to a solvent about 1.1 times the volume of ethanol, add 30 ml of ethyl acetate to the residual liquid for extraction, and concentrate the organic phase under reduced pressure at 40°C to dryness to obtain compound IV. S3: Add 30 ml of concentrated hydrochloric acid with a mass fraction of 37% to compound IV, reflux for 4 hours, and detect the completion of the reaction by TLC. Filter, adjust the pH to 7.0 with 50% sodium hydroxide, add 6.1 g of compound V, reflux for 7 hours, and detect the completion of the reaction by TLC. S4: Solution B containing compound VI obtained in S3 was concentrated to dryness under reduced pressure, and 50 ml of acetonitrile was added. After filtration, 8.5 g of sodium nitrite and 20 ml of 1N hydrochloric acid were added. The mixture was stirred at room temperature for 4 h. The reaction was detected to be complete by TLC. The solution was purified by silica gel column chromatography using n-heptane / ethyl acetate in a volume ratio of 2:1 as eluent to obtain 1.84 g of the target product. The purity was 97.32% as determined by HPLC.

[0035] Example 3 S1: 4.3 g of compound I and 2.7 g of compound II were added to 20 mL of methanol, stirred at 30°C for 6 h, and the reaction was completed by TLC detection to obtain solution A containing compound III.

[0036] S2: Cool the solution A containing compound III obtained in S1 to 0°C, add 1.0 g of sodium cyanoborohydride in batches, warm to room temperature after addition, and continue stirring to react for 3 hours. TLC detection shows that the reaction is complete. The reaction solution is concentrated under reduced pressure at 40°C to remove methanol, and 20 ml of water and 20 ml of ethyl acetate are added to the residual liquid for extraction, washed with 20 ml of water, and the organic phase is concentrated under reduced pressure at 40°C to dryness to obtain compound IV. S3: 30 ml of 37% concentrated hydrochloric acid was added to compound IV, and the mixture was refluxed for 4 h. The reaction was completed by TLC. The mixture was filtered, and the pH was adjusted to 7.0 with 50% sodium hydroxide. 4.7 g of compound V was added, and the mixture was refluxed for 7 h. The reaction was completed by TLC to obtain solution B containing compound VI.

[0037] S4: The mixture containing compound VI obtained in S3 was concentrated under reduced pressure, and then 40 ml of dichloromethane was added. After filtration, 6.8 g of potassium nitrite and 15 ml of 1N hydrochloric acid were added and stirred at room temperature for 4 h. The reaction was completed after TLC detection. The mixture was purified by silica gel column chromatography using n-heptane / ethyl acetate with a volume ratio of 2:1 as eluent to obtain 1.37 g of the target product. The purity was 96.85% as determined by HPLC. Example 4 S1: Add 0.1 mol of compound I and 0.095 mol of compound II into 80 mL of methanol, stir and react at 20°C. After TLC detection, the reaction is complete to obtain solution A containing compound III.

[0038] S2: Add 80 ml of water to solution A containing compound III obtained in S1, cool to 0°C, add 2.9 g of sodium borohydride in batches, and after addition, warm to room temperature and continue stirring to react for 3 hours. TLC detection shows that the reaction is complete. Concentrate the reaction solution under reduced pressure at 40°C to a solvent with a volume of methanol. Add 80 ml of ethyl acetate to the residual liquid for extraction. Concentrate the organic phase under reduced pressure at 40°C to dryness to obtain compound IV.

[0039] S3: 140 ml of 37% concentrated hydrochloric acid was added to compound IV, and the mixture was refluxed for 4 h. The reaction was detected by TLC to be complete. The mixture was filtered, and the pH was adjusted to 6.5 with 50% sodium hydroxide. 0.09 mol of compound V was added, and the mixture was refluxed. The reaction was detected by TLC to be complete, thereby obtaining solution B containing compound VI.

[0040] S4: Solution B containing compound VI obtained in S3 was concentrated to dryness under reduced pressure, 160 ml of methanol was added, and after filtration, 18 ml of tert-butyl nitrite was added and stirred at room temperature for 2 h. The reaction was detected to be complete by TLC. The product was purified by silica gel column chromatography using n-heptane / ethyl acetate in a volume ratio of 2:1 as eluent to obtain 6.3 g of the target product with a purity of 97.51% as determined by HPLC.

[0041] Example 5 S1: Add 0.1 mol of compound I and 0.1 mol of compound II to 100 mL of water, stir and react at 60°C. After TLC detection, the reaction is complete to obtain solution A containing compound III.

[0042] S2: Add 120 ml of water to solution A containing compound III obtained in S1, cool to 0°C, add 2.9 g of sodium borohydride in batches, and after addition, warm to room temperature and continue stirring to react for 3 hours. TLC detection shows that the reaction is complete. The reaction solution is concentrated under reduced pressure at 40°C to about 100 ml. 120 ml of ethyl acetate is added to the residual liquid for extraction. The organic phase is concentrated under reduced pressure at 40°C to dryness to obtain compound IV.

[0043] S3: 140 ml of 37% concentrated hydrochloric acid was added to compound IV, and the mixture was refluxed for 4 h. The reaction was completed by TLC. The mixture was filtered, and the pH was adjusted to 7.2 with 50% sodium hydroxide. 0.1 mol of compound V was added, and the mixture was refluxed. The reaction was completed by TLC to obtain solution B containing compound VI.

[0044] S4: Solution B containing compound VI obtained in S3 was concentrated to dryness under reduced pressure, and 240 ml of tetrahydrofuran was added. After filtration, 36 ml of tert-butyl nitrite was added and stirred at room temperature. TLC detection showed that the reaction was complete. The product was purified by silica gel column chromatography using n-heptane / ethyl acetate in a volume ratio of 2:1 as eluent to obtain 6.9 g of the target product. The purity was 96.31% as determined by HPLC.

[0045] Comparative Example 1 Based on Example 1, the pH of S3 was adjusted to 5.0, and the rest remained unchanged. TLC detection of S3 showed that a large amount of starting material remained. The subsequent preparation was continued, and after column chromatography purification, almost no target product was obtained.

[0046] Comparative Example 2 Based on Example 1, the pH of S3 was adjusted to 9.0, and the rest remained unchanged. TLC analysis of S3 showed that a large amount of starting material remained, so the preparation was continued. After column chromatography purification, 1.5 g of the target product was obtained, and the purity was 95.06% as determined by HPLC.

[0047] Effect Example 1 Detection method: This example detects the prepared impurities according to General Chapter 0512 (High Performance Liquid Chromatography) of Part IV of the Pharmacopoeia of the People's Republic of China (2020 edition).

[0048] (1) Chromatographic conditions Chromatographic column: Use cyanosilane bonded silica gel as the filler (Agilent ZORBAX SB-CN, 4.6 mm × 250 mm, 5 μm is selected for this application, and other chromatographic columns with equivalent performance can be selected); Detection wavelength: 210nm; Flow rate: 1.5 ml / min; Column temperature: 30°C; Injection volume: 10µl; Mobile phase A: potassium dihydrogen phosphate sodium octane sulfonate solution (dissolve 0.92 g of potassium dihydrogen phosphate and 2.92 g of sodium octane sulfonate in 1000 ml of water and adjust the pH to 4.0 with phosphoric acid)-acetonitrile (90:10, v / v); Mobile phase B: potassium dihydrogen phosphate sodium octane sulfonate solution-acetonitrile (40:60, v / v); The gradient elution program is shown in Table 1: Table 1 (2) Solution preparation Test solution: Accurately weigh about 15 mg of the impurity prepared in Example 1, place it in a 10 ml volumetric flask, add acetonitrile to dissolve and dilute to the scale, and shake well.

[0049] Test results such as Figure 5 As shown, it is proved that the nitrogen seven-membered ring nitrosamine impurity synthesized in the present application has high purity and can be used as a reference substance for azelastine nitrosamine impurities.

[0050] Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art may still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein; and these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for synthesizing nitrogen-containing seven-membered ring nitrosamine impurities, characterized in that: It is synthesized via the following route: ； S1: Condensing compound I and compound II in solvent 1 to obtain solution A containing compound III; S2: adding a reducing agent to the solution A containing compound III obtained in S1 to carry out a reduction reaction. After the reaction is completed, the solution is post-treated to obtain compound IV; S3: hydrolyze compound IV obtained in S2 in an aqueous solution of a strong acid, filter after completion of the hydrolysis, adjust the pH to 6.5-7.2, add compound V, heat under reflux for condensation and dehydration cyclization, and complete the reaction to obtain solution B containing compound VI; S4: The solvent of solution B containing compound VI obtained in S3 is concentrated under reduced pressure, solvent 2 is added, and after filtration, a nitrosating agent is added. After the reaction is completed, the target impurity is purified by column chromatography to obtain the target impurity.

2. The synthesis method according to claim 1, wherein The molar ratio of compound I to compound II to compound V is 1:0.95-1.0:0.85-1.

0.

3. The synthesis method according to claim 1, wherein Solvent 1 is one of methanol, ethanol, water, a mixed solution of methanol and water, and a mixed solution of ethanol and water; solvent 2 is one of dichloromethane, methanol, ethanol, isopropanol, tetrahydrofuran, and acetonitrile; and the aqueous solution of the strong acid is an aqueous hydrochloric acid solution with a mass fraction of 36%–38%.

4. The synthesis method according to claim 1, characterized in that The relationship between the amount of compound Ⅰ and solvent 1 is 0.1 mol: 80-120 ml; The relationship between the amount of compound I and the aqueous solution of strong acid is 0.1 mol: 120-150 ml; The usage ratio of compound I and solvent 2 is 0.1 mol: 160-240 ml.

5. The synthesis method according to claim 1, characterized in that The reducing agent in S2 is one of sodium cyanoborohydride, sodium borohydride or potassium borohydride; the nitrosating agent in S4 is one of tert-butyl nitrite, sodium nitrite or potassium nitrite.

6. The synthesis method according to claim 1, characterized in that The reducing agent in S2 is sodium borohydride, and the nitrosating agent in S4 is tert-butyl nitrite.

7. The synthesis method according to claim 1, characterized in that The molar ratio of compound I to the reducing agent is 1:0.75-1.0; The molar ratio of compound I to the nitrating agent is 1:1.5-5.

8. The synthesis method according to claim 1, characterized in that The reaction temperature in S1 is 20-60°C; the adding temperature of the reducing agent in S2 is 0-10°C, and the reaction temperature is room temperature; the reaction temperature in S4 is room temperature.

9. The synthesis method according to claim 1, characterized in that When the reducing agent is added to S2, water is added simultaneously. The post-treatment in S2 is to concentrate the reaction solution under reduced pressure until the volume of the solvent is reduced to 1-1.1 times the volume of solvent 1, and the same volume of ethyl acetate is added to the residual solution for extraction, and the organic phase is concentrated under reduced pressure to obtain compound IV. or when the reducing agent is added in S2, water is not added. The post-treatment in S2 is to concentrate the reaction solution under reduced pressure until the volume of the solvent is reduced to an amount equivalent to the volume of solvent 1, add equal volumes of water and ethyl acetate to the residue for extraction, and concentrate the organic phase under reduced pressure to obtain compound IV; The dosage of compound I and water is 0.1 mol: 80-120 ml.

10. The synthesis method according to claim 1, characterized in that The column chromatography purification conditions described in S4 are: purification by silica gel column chromatography, using n-heptane-ethyl acetate in a volume ratio of 2:1 as the eluent.

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