Nimodipine impurity E as well as preparation method and application thereof

Abstract

The invention provides a nimodipine analogue E with a brand new structure or a pharmaceutically acceptable salt thereof. And a pharmaceutical composition comprising nimodipine and the nimodipine analogue E; the invention also provides a preparation method of the nimodipine analogue E. The nimodipine analogue E is prepared by carrying out formylation reaction on a nimodipine impurity F and formic acid in an organic solvent. The application of the nimodipine analogue E in the field of medicine includes but not limited to impurity detection and quality control of nimodipine raw materials or preparations thereof, the impurity detection method can separate and detect eight impurities at the same time, and the nimodipine analogue E can be applied to the field of medicine for the nimodipine raw materials or the preparations thereof, especially injections and oral solutions. And the method has extremely important quality control guiding significance.

Description

Technical Field

[0001] The invention belongs to the field of nimodipine impurities, and particularly relates to a nimodipine impurity E and a preparation method and application thereof. Background Art

[0002] Nimodipine, chemically known as isopropyl-2-methoxyethyl-1,4-dihydro-2,6-dimethyl-4(3-nitrophenyl)-3,5-pyridinedicarboxylate, is a second-generation 1,4-dihydropyridine calcium channel antagonist. It is clinically used to prevent and treat ischemic nerve damage caused by cerebral vasospasm after subarachnoid hemorrhage, as well as senile brain function damage, migraine, sudden deafness, etc. It is the first choice for the treatment of cerebrovascular diseases.

[0003] Nimodipine is a dihydropyridine derivative, which is characterized by light instability. In order to control the quality of nimodipine API and related preparations, pharmacopoeias of various countries have made corresponding regulations on nimodipine impurities. The Chinese Pharmacopoeia discloses nimodipine impurity I and stipulates the impurity limit: impurity I≤0.5%, other single impurities≤1.0%, and total impurities≤2.0%. The European Pharmacopoeia discloses impurity A (corresponding to impurity I in the Chinese Pharmacopoeia), impurity B and impurity C, and stipulates the impurity limit: impurity C≤0.2%, other single impurities≤0.10%, and total impurities≤0.5%.

[0004]

[0005] It can be seen that in addition to impurities A, B, and C, there are many unknown impurities in nimodipine raw materials and related preparations. Luo Jing et al. [1] The impurities of nimodipine were studied, and their structures were determined by analytical methods such as HPLC-MS and NMR. A new impurity 2 was discovered for the first time. Riekes MK et al. [2] showed that nimodipine produces a variety of degradation products under acid degradation, alkaline degradation, and photodegradation conditions, but the specific structures of the degradation products were not studied.

[0006]

[0007] The inventors of the present application discovered p-hydroxybenzoic acid, methylparaben, impurity F, and impurities E and G of a completely new structure during the research of nimodipine API and related preparations, and impurities E, F, and G increased significantly during the stability period. Therefore, it is necessary to provide a nimodipine impurity E and a preparation method thereof for the quality control of nimodipine API and related preparations.

[0008]

[0009] References:

[0010] [1] Luo Jing, Liu Haijing, He Langchong, et al. Determination of related substances in nimodipine and its preparations by HPLC [J]. Journal of Pharmaceutical Analysis, 2007, 27(7): 1039-1042.

[0011] [2]Riekes MK, Rauber GS, Gislaine K, et al. Determination of Nimodipinein the Presence of its Degradation Products and Overall Kinetics through aStability-Indicating LC Method[J]. Journal of Chromatographic Science, 2013(6):511-516. Summary of the invention

[0012] The technical problem to be solved by the present invention is to provide a nimodipine impurity E with a completely new structure, a pharmaceutical composition comprising nimodipine and impurity E, a preparation method of impurity E, and application of impurity E in the medical field.

[0013] The above technical problems to be solved are achieved through the following technical solutions:

[0014] The present invention provides a nimodipine analogue, which is a compound represented by formula E or a pharmaceutically acceptable salt thereof:

[0015]

[0016] The present invention provides a pharmaceutical composition comprising nimodipine and nimodipine analog E.

[0017] Preferably, the content of nimodipine analogue E in the pharmaceutical composition is ≤0.5%.

[0018] More preferably, the content of nimodipine analogue E in the pharmaceutical composition is ≤0.3%.

[0019] Preferably, the pharmaceutical composition further comprises a solvent, and the solvent includes but is not limited to one or more of water and ethanol.

[0020] Preferably, the pharmaceutical composition further comprises a solubilizer, which includes but is not limited to one or more of glycerol, propylene glycol, and polyethylene glycol.

[0021] Preferably, the pharmaceutical composition further comprises an antibacterial agent, which includes but is not limited to one or more of methyl parahydroxybenzoate, ethyl parahydroxybenzoate, propyl parahydroxybenzoate, and butyl parahydroxybenzoate.

[0022] Preferably, the pharmaceutical composition further comprises a pH adjuster, which includes but is not limited to one or more of citric acid, sodium citrate, sodium monohydrogen phosphate, and sodium dihydrogen phosphate.

[0023] More preferably, the pharmaceutical composition further comprises the following components: ethanol, glycerol, methyl paraben and polyethylene glycol.

[0024] More preferably, the pharmaceutical composition further comprises the following components: ethanol, glycerol, methyl parahydroxybenzoate and polyethylene glycol, sodium monohydrogen phosphate, sodium dihydrogen phosphate and water.

[0025] More preferably, the pharmaceutical composition further comprises the following components: ethanol, polyethylene glycol, sodium citrate, citric acid and water.

[0026] The present invention also provides a method for preparing nimodipine analogue E, and the reaction scheme is as follows:

[0027]

[0028] The reaction steps include: impurity F and formic acid undergo formylation reaction in an organic solvent to obtain impurity E.

[0029] Preferably, the organic solvent includes, but is not limited to, one or more of dichloromethane, toluene, xylene, and tetrahydrofuran.

[0030] More preferably, the organic solvent is dichloromethane.

[0031] Preferably, the formylation reaction further comprises the use of an activating agent.

[0032] More preferably, the activating agent is carbonyl imidazole.

[0033] Furthermore, the preparation method of nimodipine analogue E provided by the present invention comprises the following steps:

[0034] 1) impurity F, formic acid and an activator are subjected to a formylation reaction in an organic solvent;

[0035] 2) Extract, wash and purify on a silica gel column to obtain impurity E.

[0036] Preferably, in step 1), the usage ratio of impurity F, formic acid and activator is 1:0.4-10:0-2 (w / w).

[0037] More preferably, in step 1), the usage ratio of impurity F, formic acid and activator is 1:0.4:1.4 (w / w).

[0038] Preferably, in step 1), the formylation reaction temperature is 25° C. to 120° C., and the reaction time is 4 h to 16 h.

[0039] More preferably, in step 1), the formylation reaction temperature is 50° C. and the reaction time is 5 h.

[0040] Most preferably, step 1) is to take 0.2 g of formic acid, 0.7 g of carbonyl imidazole, and 20 mL of dichloromethane, stir evenly, add 0.5 g of impurity F, and react at 50° C. for 5 h.

[0041] Preferably, the extraction and washing solvent in step 2) includes, but is not limited to, one or more of water, saturated sodium bicarbonate, and dilute ammonia water.

[0042] More preferably, the extraction and washing solvent in step 2) is water.

[0043] Preferably, the eluent for silica gel column purification in step 2) includes, but is not limited to, one or more of ethyl acetate, petroleum ether, dichloromethane, and methanol.

[0044] More preferably, the eluent for silica gel column purification in step 2) is ethyl acetate:petroleum ether=1:2.

[0045] Most preferably, step 2) is to extract with water and dry with anhydrous sodium sulfate; concentrate the organic phase and purify it with a silica gel column, the eluent is ethyl acetate: petroleum ether = 1:2, collect the eluate and carry out the flow with a small amount of methyl tert-butyl ether to obtain an off-white solid which is impurity E.

[0046] The present invention also provides the application of nimodipine analogue E in the medical field, and its uses include but are not limited to impurity detection and quality control of nimodipine raw materials or preparations thereof.

[0047] A nimodipine analogue E can be used as nimodipine impurity E for detecting impurities in nimodipine raw materials or preparations.

[0048] Preferably, the impurity detection method is to detect nimodipine impurities by high performance liquid chromatography. The nimodipine impurities referred to herein include but are not limited to known impurities that have been disclosed (e.g., p-hydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity F, etc.), and unknown impurities E and impurity G discovered by the applicant.

[0049] More preferably, when high performance liquid chromatography is used to detect nimodipine impurities, the chromatographic column is selected to use octadecylsilane bonded silica gel as a filler.

[0050] More preferably, when nimodipine impurities are detected by high performance liquid chromatography, the mobile phase is: potassium dihydrogen phosphate as mobile phase A, methanol-acetonitrile as mobile phase B, and linear gradient elution is performed as shown in the following table:

[0051]

[0052] Most preferably, linear gradient elution is performed as shown below:

[0053]

[0054] Most preferably, linear gradient elution is performed as shown below:

[0055]

[0056] More preferably, the concentration of potassium dihydrogen phosphate in the mobile phase is 20 mmol / L.

[0057] More preferably, the pH range of potassium dihydrogen phosphate in the mobile phase is 3.0 to 3.2.

[0058] More preferably, the chromatographic column has a specification of 4.6 mm×250 mm.

[0059] The most preferred chromatographic columns are Thermo Syncronis C18 (4.6×250 mm, 5 μm), AcchromTech

[0060] Unitary C18 100A (4.6×250mm, 5μm), Agilent Zorbax sb-C18 (4.6×250mm, 5μm), or a column with equivalent performance.

[0061] More preferably, the detection wavelength is 230 to 240 nm.

[0062] Most preferably, the detection wavelength is 235 nm.

[0063] More preferably, the column temperature is 28-32°C.

[0064] Most preferably, the column temperature is 30°C.

[0065] More preferably, the flow rate is 0.8 to 1.2 ml / min.

[0066] Most preferably, the flow rate is 1.0 ml / min.

[0067] A nimodipine analogue E can be used as a standard substance or reference substance for nimodipine impurity E, and can be used for quality control of nimodipine raw materials or preparations thereof, or for establishing quality standards.

[0068] Preferably, the nimodipine preparation is selected from a solid preparation, a semisolid preparation and a liquid preparation.

[0069] More preferably, the nimodipine preparation is selected from liquid preparations.

[0070] Most preferably, the nimodipine formulation is an injection or an oral solution.

[0071] Compared with the prior art, the beneficial effects of the nimodipine injection of the present invention are:

[0072] 1) The present application provides a nimodipine impurity E which is not disclosed in the prior art and has a completely new structure. The structure of the impurity E is confirmed by liquid chromatography-mass spectrometry, carbon nuclear magnetic resonance spectroscopy, and hydrogen nuclear magnetic resonance spectroscopy, and is a degradation impurity of nimodipine.

[0073] 2) The present application also provides a method for preparing impurity E, which can be prepared by formylation reaction of nimodipine impurity F with formic acid, and purified by silica gel column to obtain pure impurity E, which can be used as a standard or reference substance for nimodipine degradation impurity E.

[0074] 3) The present application also provides a method for detecting impurity E or simultaneously detecting impurity E and other nimodipine impurities. By optimizing the mobile phase and elution gradient of high performance liquid chromatography, the technical effect of simultaneously separating and detecting 8 impurities (parahydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity E, impurity F and impurity G) is achieved, so as to control the quality of nimodipine raw materials or their preparations and establish corresponding quality standards; in particular, it has extremely important quality control guiding significance for the currently marketed nimodipine injection and oral solution. BRIEF DESCRIPTION OF THE DRAWINGS

[0075] Figure 1 is the mass spectrum of nimodipine impurity E in Example 2;

[0076] Figure 2 is the carbon NMR spectrum of the nimodipine impurity E in Example 2;

[0077] Figure 3 is the hydrogen nuclear magnetic resonance spectrum of the nimodipine impurity E in Example 2;

[0078] Figure 4 The results of the nimodipine related substance test in Example 3 are as follows;

[0079] Figure 5 This is the detection result of nimodipine related substances in comparative example 1. DETAILED DESCRIPTION

[0080] The following are specific embodiments of the present invention, which further describe the technical solution of the present invention, but the protection scope of the present invention is not limited to these embodiments. Any changes or equivalent substitutions that do not deviate from the concept of the present invention are protected within the protection scope of the present invention.

[0081] The reagents and instruments used in the examples are all conventional reagents and instruments in the art and can be purchased through conventional channels. The following is only for illustration.

[0082] Reagents:

[0083] Nimodipine oral solution (trade name: Nymalize, specification: 60mg / 20ml, manufacturer: Arbor Pharmaceuticals)

[0084] p-Hydroxybenzoic acid (purity: 100.0%, manufacturer: Sigma)

[0085] Methylparaben (purity: 100.0%, manufacturer: Sigma)

[0086] Nimodipine impurity A (purity: 87.0%, manufacturer: China National Institute for Food and Drug Control)

[0087] Nimodipine impurity B (purity: 99.37%, manufacturer: GP)

[0088] Nimodipine impurity C (purity: 98.12%, manufacturer: GP)

[0089] instrument:

[0090] High performance liquid chromatograph (model: U3000, manufacturer: Thermo Fisher Scientific, USA)

[0091] Liquid chromatography-mass spectrometry (Model: Thermo U3000 (Double Ternary) + Q-EXACTIVE, Manufacturer: Thermo Fisher Scientific, USA)

[0092] Nuclear magnetic resonance spectrometer (model: Bruker BioSpin GmbH 400MHz, manufacturer: Bruker, USA).

[0093] Example 1 Preparation of Impurity E

[0094] Take 0.2 g of formic acid, 0.7 g of carbonyl imidazole, and 20 mL of dichloromethane, stir evenly, then add 0.5 g of impurity F to react, and monitor the reaction by TLC; after the reaction is completed, wash with water twice, and then dry with anhydrous sodium sulfate; concentrate the organic phase and purify it with a silica gel column, the eluent is ethyl acetate: petroleum ether = 1:2, collect the eluate and carry out the flow with a small amount of methyl tert-butyl ether to obtain an off-white solid, which is impurity E (purity: 98.71%, yield: 95.0%).

[0095]

[0096] Example 2 Characterization of Impurity E

[0097] The molecular weight and structure of the nimodipine impurity E obtained in Example 1 were characterized by liquid chromatography-mass spectrometry, carbon nuclear magnetic resonance spectroscopy and hydrogen nuclear magnetic resonance spectroscopy.

[0098] The mass spectrum of impurity E is as follows Figure 1 As shown, the mass-to-charge ratio (m / z) of impurity E [M+H] + It is 417.0.

[0099] The carbon NMR spectrum of impurity E is as follows Figure 2 As shown, 13 C-NMR(DMSO-d6,100MHz)δ166.87(C13), 166.40(C9), 159.26(C24), 148.87(C2) , 145.71(C6), 145.03(C17), 137.80(C21), 128.11(C19), 122.87(C18), 118.42( C22), 116.83(C20), 102.09(C3), 101.35(C5), 69.93(C14), 66.08(C11), 62.35( C10), 58.09(C12), 40.14(C4), 21.84(C15), 21.56(C16), 18.23(C8), 18.15(C7).

[0100] The H NMR spectrum of impurity E is as follows Figure 3 As shown, 1 H-NMR(DMSO-d6,400MHz)δ10.05(s,1H,C24–CH), 8.21,8.81(s,2H,C1–CH,C23 –CH), 6.88-7.40(m,4H,C18–CH,C19–CH,C20–CH,C22–CH), 4.78-4.84(m,2H,C4 –CH,C14–CH), 3.48-4.07(m,4H,C10–CH2,C11–CH2), 3.25(s,3H,C12–CH3), 2. 26(s,6H,C7–CH3,C8–CH3), 1.05-1.06,1.17-1.19(dd,6H,C15–CH3,C16–CH3).

[0101] Therefore, the molecular weight and structure of impurity E are as follows:

[0102]

[0103] Example 3

[0104] A method for detecting nimodipine-related substances comprises the following steps:

[0105] 1) Sample treatment: Take appropriate amounts of nimodipine, p-hydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity E, impurity F and impurity G reference substances, dissolve them in methanol-acetonitrile-water (35:38:27) solvent and quantitatively dilute them to prepare a mixed solution containing approximately 300 μg nimodipine, 6 μg p-hydroxybenzoic acid, 200 μg methylparaben, 1.5 μg impurity A, 0.6 μg impurity B, 0.6 μg impurity C, 0.6 μg impurity E, 0.6 μg impurity F and 0.6 μg impurity G per 1 ml as the system suitability solution.

[0106] 2) High performance liquid chromatography detection: Take the sample solution in step 1) and inject it. The detection conditions are as follows:

[0107] Chromatographic column: Thermo Syncronis C18 (4.6×250mm, 5μm) or chromatographic column with equivalent performance;

[0108] Mobile phase: 20 mmol / L (Example 3) potassium dihydrogen phosphate (adjusted to pH 3.0-3.2 with phosphoric acid) as mobile phase A, methanol-acetonitrile (50:50) as mobile phase B, linear gradient elution according to the following table:

[0109]

[0110] Detection wavelength: 235nm;

[0111] Column temperature: 30°C;

[0112] Flow rate: 1.0ml / min;

[0113] Injection volume: 20 μl.

[0114] The results of the investigation are shown in Table 1:

[0115] Table 1. Detection method of nimodipine related substances - Investigation results of Example 3

[0116]

[0117] The results show that by adding potassium dihydrogen phosphate to the mobile phase, preferably at a concentration of 20 mmol / L, under the gradient elution conditions shown in Example 3, the separation degree of impurity E and impurity F can be made greater than 1.5, and the simultaneous separation and detection of eight related substances of nimodipine can be achieved: p-hydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity E, impurity F and impurity G.

[0118] Example 4

[0119] A method for detecting nimodipine-related substances comprises the following steps:

[0120] 1) Sample treatment: Take appropriate amounts of nimodipine, p-hydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity E, impurity F and impurity G reference substances, dissolve them in methanol-acetonitrile-water (35:38:27) solvent and quantitatively dilute them to prepare a mixed solution containing approximately 300 μg nimodipine, 6 μg p-hydroxybenzoic acid, 200 μg methylparaben, 1.5 μg impurity A, 0.6 μg impurity B, 0.6 μg impurity C, 0.6 μg impurity E, 0.6 μg impurity F and 0.6 μg impurity G per 1 ml as the system suitability solution.

[0121] 2) High performance liquid chromatography detection: Take the sample solution in step 1) and inject it. The detection conditions are as follows:

[0122] Chromatographic column: Thermo Syncronis C18 (4.6×250mm, 5μm) or chromatographic column with equivalent performance;

[0123] Mobile phase: 20 mmol / L (Example 4) potassium dihydrogen phosphate (pH adjusted to 3.0-3.2 with phosphoric acid) as mobile phase A, methanol-acetonitrile (50:50) as mobile phase B, linear gradient elution according to the following table:

[0124]

[0125] Detection wavelength: 235nm;

[0126] Column temperature: 30°C;

[0127] Flow rate: 1.0ml / min;

[0128] Injection volume: 20 μl.

[0129] The results of the investigation are shown in Table 2:

[0130] Table 2. Elution gradient of nimodipine related substance detection method - investigation results of Example 4

[0131]

[0132] The results show that when the elution gradient shown in Implementation 4 is selected, the separation degree of impurities E and F can be made greater than 1.5, and the simultaneous separation and detection of eight related substances of nimodipine can be achieved: p-hydroxybenzoic acid, methylparaben, impurity A, impurity B, impurity C, impurity E, impurity F and impurity G.

[0133] Comparative Example 1: Refer to the relevant substance detection method of Chinese Pharmacopoeia

[0134] A method for detecting nimodipine-related substances comprises the following steps:

[0135] 1) Sample treatment: Take appropriate amounts of nimodipine, methylparaben, impurity A, impurity B, impurity C, impurity D, impurity E, impurity F and impurity G reference substances, dissolve them in methanol-acetonitrile-water (35:38:27) solvent and quantitatively dilute them to prepare a mixed solution containing approximately 300 μg nimodipine, 200 μg methylparaben, 1.5 μg impurity A, 0.6 μg impurity B, 0.6 μg impurity C, 0.6 μg impurity E, 0.6 μg impurity F and 0.6 μg impurity G per 1 ml as the system suitability solution.

[0136] 2) High performance liquid chromatography detection: Take the sample solution in step 1) and inject it. The detection conditions are as follows:

[0137] Chromatographic column: Thermo Syncronis C18 (4.6×250mm, 5μm) or chromatographic column with equivalent performance;

[0138] Mobile phase: methanol-acetonitrile-water (35:38:27)

[0139] Detection wavelength: 235nm;

[0140] Column temperature: 30°C;

[0141] Flow rate: 1.0ml / min;

[0142] Injection volume: 20 μl.

[0143] Test results such as Figure 5 As shown in Table 3:

[0144] Table 3. Results of testing nimodipine related substances with reference to the Chinese Pharmacopoeia method

[0145]

[0146] The results showed that the separation degree of impurity E and impurity F was only 1.28<1.5, which could not be effectively separated and detected. Therefore, it was necessary to optimize the detection method based on the relevant substances in the Chinese Pharmacopoeia.

Claims

1. A nimodipine analogue, which is a compound represented by formula E or a pharmaceutically acceptable salt thereof:

2. A pharmaceutical composition comprising nimodipine and the nimodipine analog E according to claim 1.

3. The pharmaceutical composition according to claim 2, characterized in that The content of the nimodipine analogue E is ≤0.5%.

4. A method for preparing the nimodipine analogue E according to claim 1, wherein the reaction scheme is as follows: The reaction steps include: Impurity F and formic acid undergo formylation reaction in an organic solvent to obtain impurity E.

5. The method for preparing nimodipine analogue E according to claim 4, characterized in that: The organic solvent includes, but is not limited to, one or more of dichloromethane, toluene, xylene, and tetrahydrofuran.

6. The method for preparing nimodipine analogue E according to claim 4, characterized in that: The formylation reaction further comprises using an activator; preferably, the activator is carbonyl imidazole.

7. Use of the nimodipine analogue E according to claim 1 in the medical field, including but not limited to use for impurity detection, quality control of nimodipine raw materials or their preparations.

8. Use of the nimodipine analogue E as claimed in claim 7 in the medical field, including as nimodipine impurity E for detecting impurities in nimodipine raw materials or preparations thereof.

9. The use of the nimodipine analogue E in the medical field as claimed in claim 8, wherein the impurity detection method is to detect nimodipine impurities by high performance liquid chromatography.

10. The use of the nimodipine analogue E in the medical field as claimed in claim 9, wherein the chromatographic conditions for detecting nimodipine impurities by high performance liquid chromatography are as follows: chromatographic column selection: octadecylsilane bonded silica gel is used as filler. Further preferably, the mobile phase is selected as follows: potassium dihydrogen phosphate as mobile phase A, methanol-acetonitrile as mobile phase B, and linear gradient elution is performed as shown in the following table:

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