CN115109073A

CN115109073A - Crystalline tylorravone

Info

Publication number: CN115109073A
Application number: CN202110294050.5A
Authority: CN
Inventors: 苏国强; 苏柏朗; 杨加宾
Original assignee: Nanjing Zhongrui Pharmaceutical Co ltd
Current assignee: Nanjing Zhongrui Pharmaceutical Co ltd
Priority date: 2021-03-19
Filing date: 2021-03-19
Publication date: 2022-09-27

Abstract

The invention discloses a tylorravone crystal A, wherein the 2 theta angle of a characteristic peak of which the relative intensity of an X-ray powder diffraction pattern is 100% is 8.35 +/-0.2 degrees. The tylorravone crystal A has stable property, is convenient to store and use, has relatively low requirement on dissolution condition, has higher dissolution speed, and has the solubility meeting the requirement of a preparation. The invention also discloses a pharmaceutical composition which contains the tylorravone crystal A with the pharmacologically effective dose and pharmaceutically acceptable auxiliary materials. The invention also discloses an improved synthesis process of the tyraravone, which comprises the following steps: and (3) taking a saturated hydrogen chloride ethanol solution as a reaction solvent, and condensing and cyclizing the 2-cyanobenzene hydrazine and the methyl acetoacetate in the saturated hydrogen chloride ethanol solution to generate the tylorravone. The method adopts a one-pot method to prepare the tylorravone, has mild reaction conditions, can be carried out at room temperature, avoids high-temperature by-products, has high reaction yield at least up to 75 percent, and is easy to purify the product.

Description

Crystalline tylorravone

Technical Field

The invention belongs to the field of pharmaceutical chemistry, and relates to a tylravone crystal, a preparation method and application thereof.

Background

Tyrularine, chemical name: the structure of the 2-methyl-5-imino-benzo [ d ] [1,3] oxazine [5-b ] pyrazole is shown as the formula I. The tyraravone is a brand new compound (CN101508696A) designed and synthesized by a molecular skeleton migration drug design method based on a lead compound edaravone structure. In vitro experiments show that the tylorravone has the effect of remarkably eliminating hydroxyl oxygen and superoxide radical. In vivo experiments show that the tylorula can obviously improve the nerve defect symptoms of cerebral ischemia reperfusion animals in a dose-dependent manner, reduce the cerebral infarction area, reduce the brain injury degree, relieve cerebral edema and inhibit lipid peroxidation of damaged brain tissues.

Clinical research results show that the tylorrazone can obviously improve cerebral apoplexy symptoms, and has good safety and tolerance without serious adverse events. However, the tylorravone has rigid structure, stable chemical property, relatively low solubility in water or common organic solvents, is basically insoluble in water and has poor pharmacy property. The inventor Chinese patent CN 101966146A has carried out various solubilization tests, but the tylorravone is dissolved in the arginine water solution under stirring and under the condition of introducing nitrogen gas at 60-85 ℃.

Disclosure of Invention

The invention aims to provide a tylravone crystal with medicinal value.

The object of the invention can be achieved by the following measures:

a crystalline Talaravone A having an X-ray powder diffraction pattern with a 2 theta angle of 8.35 + -0.2 DEG relative to a characteristic peak having a relative intensity of 100%.

Preferably, the tylorravone crystal A has an X-ray powder diffraction pattern with characteristic peaks at 2 theta values of 8.35 +/-0.2, 10.55 +/-0.2, 13.80 +/-0.2, 14.60 +/-0.2, 15.90 +/-0.2, 17.95 +/-0.2, 19.20 +/-0.2, 21.65 +/-0.2 and 27.33 +/-0.2 degrees.

Further preferably, the X-ray powder diffraction pattern of the tylorravone crystal a is as shown in fig. 2, 3 or 4.

A preparation method of tylorravone crystal A comprises the following steps: at room temperature, adding the tylorravone into an organic solvent, adding concentrated ammonia water to dissolve the tylorravone, adding organic acid, and stirring to separate out a solid; filtering, washing with absolute ethyl alcohol, purified water and absolute ethyl alcohol in sequence, and drying at 60 ℃ to obtain the tylorravone crystal A.

The organic solvent is an organic solvent which can be mutually soluble with ammonia water, and is specifically selected from one or more of methanol, ethanol or dimethylformamide.

The organic acid is formic acid, acetic acid and the like.

The mass ratio of the organic solvent to the tylaravone is 8-10: 1, the mass ratio of the organic solvent to the ammonia water is 2-2.5: 1, and the mass ratio of the organic solvent to the organic acid is 1.5-2: 1.

As a further preferable technical scheme of the preparation method of the tylorravone crystal A, the preparation method comprises the following steps: at room temperature, adding the tylorravone into an organic solvent, adding concentrated ammonia water to dissolve the tylorravone, adding activated carbon, stirring, filtering, adding an organic acid into the filtrate, and stirring to separate out a solid; filtering, washing with absolute ethyl alcohol, purified water and absolute ethyl alcohol in sequence, and drying at 60 ℃ to obtain the tylorravone crystal A.

Another preparation method of the tylravone crystal a comprises the following steps: heating and dissolving the tylorravone in dimethylformamide, cooling to room temperature, and separating out a solid; filtering, washing a filter cake with dimethylformamide and ethanol in sequence, then soaking with purified water and ethanol in sequence, draining, and drying at 60 ℃ to obtain the tylorravone crystal A.

The mass-volume ratio of the tylorravone to the dimethylformamide is 1: 8-10.

As a further preferable technical scheme of the preparation method of the tylorravone crystal A, the preparation method comprises the following steps: heating and dissolving the tyloruon in dimethylformamide, adding activated carbon, filtering while hot, cooling to room temperature, and separating out a solid; filtering, washing a filter cake with dimethylformamide and ethanol in sequence, then soaking in purified water and ethanol in sequence, draining, and drying at 60 ℃ to obtain the tylorravone crystal A.

The solubility of the tylorravone in the dimethylformamide is obviously increased along with the increase of the temperature, and the tylorravone can be used as a recrystallization solvent. But the dimethyl formamide can be decomposed at high temperature to generate formaldehyde, and the formaldehyde and the two-molecule tylaravone are condensed to generate the dipolymer tylaravone (formula II). It is considered that the use of dimethylformamide as a recrystallization solvent for tylaravone produces a small amount of polymerized tylaravone impurity. Thus, dimethylformamide is not a preferred recrystallization solvent for tylaravone crystals a.

A crystalline Talaravone B having a characteristic peak with an X-ray powder diffraction pattern relative intensity of 100% at a 2 [ theta ] angle of 12.88 + -0.2 deg.

Preferably, the tylorravone crystal B has an X-ray powder diffraction pattern having characteristic peaks at 2 theta values of 7.22 + -0.2, 10.64 + -0.2, 12.88 + -0.2, 14.42 + -0.2, 17.79 + -0.2, 21.42 + -0.2, 25.54 + -0.2, 26.62 + -0.2, 27.06 + -0.2, 31.46 + -0.2 and 34.26 + -0.2 deg.

Further preferably, an X-ray powder diffraction pattern of the tylorravone crystal B is shown in fig. 6.

The preparation method of the tylravone crystal B comprises the following steps: heating and dissolving the tylravone in a recrystallization solvent by using micromolecular carboxylic acid or micromolecular carboxylic acid aqueous solution as a recrystallization solvent, cooling to room temperature, stirring for crystallization, filtering, washing with absolute ethyl alcohol, purified water and absolute ethyl alcohol in sequence, and drying at 60 ℃ to obtain a tylravone crystal B; wherein the small molecular carboxylic acid is water-soluble small molecular carboxylic acid such as formic acid and acetic acid; the concentration of the small molecular carboxylic acid in the small molecular carboxylic acid aqueous solution is 50% (v/v); the volume-mass ratio of the recrystallization solvent to the tylorravone is more than or equal to 50:1mL/g or L/kg.

As a further preferable technical scheme of the preparation method of the tylorravone crystal B, the preparation method comprises the following steps: heating and dissolving the terepratuone in a recrystallization solvent, adding activated carbon, filtering while hot, cooling to room temperature, stirring for crystallization, filtering, washing with absolute ethyl alcohol, purified water and absolute ethyl alcohol in sequence, and drying at 60 ℃ to obtain terepratuone crystal B.

In small molecular carboxylic acid (such as formic acid, acetic acid and the like) or small molecular carboxylic acid aqueous solution, the solubility of the tylravone does not increase remarkably along with the change of temperature, and a large amount of recrystallization solvent (such as 50% acetic acid aqueous solution and the tylravone volume-mass ratio of more than or equal to 50:1mL/g or L/kg) is required to be adopted to dissolve the tylravone by heating. The method for preparing the tylravone crystal B has low production efficiency, so that the tylravone crystal B is not a preferred medicinal crystal form of the tylravone.

A crystalline Talaravone C having an absorption peak with 100% relative intensity in X-ray powder diffraction pattern at a 2-theta angle of 10.61 + -0.2 deg.

Preferably, the tylravone crystal C has an X-ray powder diffraction pattern having characteristic peaks at 2 theta values of 7.16 + -0.2, 10.61 + -0.2, 12.81 + -0.2, 14.37 + -0.2, 17.77 + -0.2, 18.66 + -0.2, 21.35 + -0.2, 25.52 + -0.2, 26.53 + -0.2, 27.09 + -0.2 and 32.22 + -0.2 degrees.

Further preferably, an X-ray powder diffraction pattern of the tylorravone crystal C is shown in fig. 8.

The preparation method of the tylravone crystal C comprises the following steps: the method comprises the steps of taking a mixed solvent of ethanol, concentrated hydrochloric acid and water as a recrystallization solvent, heating and refluxing to dissolve the tylorhoduone, cooling and crystallizing, filtering, washing a filter cake with the recrystallization solvent and water in sequence, dispersing in water, neutralizing with a 40% sodium hydroxide solution to reach the pH of 7-8, filtering, washing the filter cake with water and absolute ethyl alcohol in sequence, and drying at about 60 ℃ to obtain the tylorhoduone crystal C. Wherein the mass ratio of the ethanol to the concentrated hydrochloric acid to the water is 8-10: 1-1.5: 1; the mass ratio of the recrystallization solvent to the tylaravone is 15-20: 1.

The imine structure of the terepratuone has acidity, so that the terepratuone is dissolved in ammonia water, and the organic acid is used for neutralizing the ammonia water to obtain the terepratuone crystal. The tylorravone crystal A prepared by the method has stable properties, is convenient to store and use, has relatively low requirement on dissolution conditions, has higher dissolution speed, and has solubility meeting the requirement of a preparation. Therefore, the terepratvone crystal A is a preferable medicinal crystal form of the terepratvone.

TABLE 1 solubility of crystalline tylorravone A in aqueous arginine solution (25 ℃ C.)

TABLE 2 dissolution of tylorravone crystals in 5% arginine in water (pH9)

Another object of the present invention is to provide a pharmaceutical composition containing a pharmacologically effective amount of crystalline a, crystalline B or crystalline C of tylaravone, and a pharmaceutically acceptable adjuvant.

Preferably, the pharmaceutical composition is a pharmaceutical preparation for injection.

The invention also aims to provide the application of the pharmaceutical composition in preparing medicaments for treating or preventing cardiovascular and cerebrovascular diseases.

The inventor discloses a synthetic method of tyraravone in Chinese patent CN 101508696A: under the alkaline condition, 2-cyano phenylhydrazine and methyl acetoacetate are condensed at the high temperature of 40-80 ℃ to generate 2- (3-methyl-5-oxo-4, 5-dihydro-1H-pyrazol-1-yl) benzonitrile, and then the tyrravone is generated through catalytic cyclization by hydrogen chloride. The reaction condition has high temperature, more byproducts and difficult product purification, and the yield is about 40 percent.

Another object of the present invention is to provide an improved synthesis process of tylravone, which comprises: and (3) taking a saturated hydrogen chloride ethanol solution as a reaction solvent, and condensing and cyclizing the 2-cyanobenzene hydrazine and the methyl acetoacetate in the saturated hydrogen chloride ethanol solution to generate the tylorravone.

The molar ratio of the 2-cyanophenylhydrazine to the methyl acetoacetate is 0.8-1: 1.

The reaction temperature was room temperature.

After the reaction is finished, cooling the reaction solution to about 5 ℃, adding ice water with the mass 2 times that of the reaction solvent, stirring to separate out a product, filtering, and washing by adopting 33% ethanol and absolute ethanol in sequence to obtain a pure Tairui ravone product.

The method adopts a one-pot method to prepare the tylaravone, has mild reaction conditions, can be carried out at room temperature, avoids high-temperature by-products, has high reaction yield at least up to 75 percent, and is easy to purify the product. The synthesis process of the teriravone is more suitable for industrial production.

Drawings

FIG. 1 is a photomicrograph of crystalline Talaravone A (example 2, aqueous ammonia/acetic acid solution).

FIG. 2 is an X-ray powder diffraction pattern of crystalline Talaravone A (example 2, aqueous ammonia/acetic acid solution, 0 months).

FIG. 3 is an X-ray powder diffraction pattern of crystalline Talaravone A (example 2, aqueous ammonia/acetic acid solution, examined at 40 ℃ for 6 months).

FIG. 4 is an X-ray powder diffraction pattern of crystalline Talaravone A (example 3, DMF solution).

Fig. 5 is a micrograph of tylravone crystal B.

FIG. 6 is an X-ray powder diffraction pattern of crystalline Talaravone B (example 4, 50% acetic acid solution).

Fig. 7 is a photomicrograph of crystalline tylravone C.

FIG. 8 is an X-ray powder diffraction pattern of crystalline Talaravone C (example 5, aqueous ethanolic hydrochloric acid).

Detailed Description

The technical solution of the present invention is further described below with reference to specific embodiments.

The concentration of the concentrated ammonia water (sold on the market) used in the invention is 25-28%, and the concentration of the concentrated hydrochloric acid (sold on the market) is 36-38%.

Example 1

Preparation of Tairuiravone

Adding 10.5kg of saturated hydrogen chloride ethanol solution, 1.5kg of 2-cyanophenylhydrazine and 1.57kg of methyl acetoacetate into a 50L reaction kettle under stirring, and stirring at room temperature for reacting overnight; and cooling the reaction liquid to about 5 ℃, adding 21kg of ice water, stirring for 1-1.5 hours at 5 ℃, carrying out suction filtration, washing the filter cake for 2-3 times by using a 33% ethanol solution, washing the filter cake for 1 time by using absolute ethanol, drying at about 60 ℃ after suction drying, and obtaining 1.91kg of the tylorravone.

Example 2

Adding 1.97kg of Tairui ravone (prepared according to the method of example 1) and 17.57kg of absolute ethyl alcohol into a 50L reaction kettle, stirring at room temperature, adding 7.64kg of concentrated ammonia water, adding 0.1kg of activated carbon after the Tairui ravone is completely dissolved, stirring for 30 minutes at 25-35 ℃, carrying out suction filtration, transferring filtrate into a clean reaction kettle, adding 8.83kg of glacial acetic acid under stirring, naturally cooling to room temperature, stirring, and crystallizing overnight; filtering, washing with absolute ethyl alcohol, soaking and washing with pure water for 4 times, finally washing with absolute ethyl alcohol, fully draining, and drying at 60 ℃ to obtain 1.58kg of the tylravone crystal.

A micrograph of crystalline Talaravone A is shown in FIG. 1.

The X-ray powder diffraction pattern (0 month) of the tylorravone crystal A is shown in figure 2, the stability of the tylorravone crystal A is examined (40 ℃ accelerated test for 6 months), the X-ray powder diffraction pattern is shown in figure 3, and the tylorravone crystal A is stable in property and convenient to store and use.

TABLE 3X-ray powder diffraction data for crystalline Talaravone A

Example 3

Adding 43g of Tairuiravone (prepared according to the method of example 1) and 400mL of Dimethylformamide (DMF) into a 1000mL reaction bottle, heating to slightly boil, completely dissolving, slightly cooling, adding 1g of activated carbon, and heating for 20 min; hot filtering, cooling to room temperature for crystallization, and filtering; and washing a filter cake by using 50mL of dimethylformamide and 50mL of ethanol in sequence, then soaking for 20min by using 50mL of purified water, finally soaking for 20min by using 50mL of ethanol, fully draining, and drying at 60 ℃ to obtain a tylaravone crystal A36 g.

An X-ray powder diffraction pattern of crystalline Talaravone A is shown in FIG. 4.

TABLE 4X-ray powder diffraction data for crystalline Talaravone A

2θ	d value	Relative intensity/%)
			8.388	10.532	100
10.431	8.474	3.1
			13.772	6.425	43.4
14.767	5.994	15.0
			16.049	5.518	21.6
17.907	4.949	4.5
			19.236	4.611	6.7
21.651	4.101	30.6
			27.284	3.266	43.7

Example 4

Adding 10g of Tairuiravone, 250mL of acetic acid and 250mL of water into a 1000mL reaction bottle, heating and refluxing for dissolution, after all the components are dissolved, slightly cooling, adding 0.5g of activated carbon, heating to reflux, filtering while hot, naturally cooling the filtrate to room temperature, and stirring for crystallization overnight. Filtering, washing with anhydrous ethanol, soaking with purified water for 4 times, washing with anhydrous ethanol, and completely draining. Drying at 60 ℃ to obtain 6.58g of crystalline Tararravone B.

A micrograph of crystalline Talaravone B is shown in FIG. 5.

An X-ray powder diffraction pattern of crystalline form B of tirilavone is shown in figure 6.

TABLE 5X-ray powder diffraction data for crystalline tylorrazone A

2θ	d value	Relative intensity/%)
			7.228	12.220	1.5
10.639	8.309	18.6
			12.876	6.870	100.0
14.416	6.139	12.5
			17.797	4.980	3.7
21.422	4.145	8.4
			25.549	3.484	2.8
26.621	3.346	4.6
			27.057	3.293	9.2
31.469	2.841	2.5
			34.261	2.615	3.5

Example 5

Adding 1.91kg of Tairuiravone (prepared according to the method of the embodiment 1), 26kg of purified water, 3.91kg of concentrated hydrochloric acid and 2.6kg of absolute ethyl alcohol into a 50L reaction kettle, heating to reflux, keeping refluxing for 20-30 minutes to dissolve the materials, stopping heating, reducing the temperature to 85-90 ℃, adding activated carbon, and continuing heating and refluxing for 30-40 minutes; filtering while hot, and cooling the filtrate to room temperature for crystallization; and (2) carrying out suction filtration, sequentially washing a filter cake with a recrystallization solvent (prepared from purified water, concentrated hydrochloric acid and absolute ethyl alcohol according to the mass ratio of 26:3.91:2.6 kg) and water, adding the filter cake into a reaction kettle, adding 16kg of water, dropwise adding a 40% sodium hydroxide solution under stirring at room temperature to neutralize the solution until the pH value is 7-8, carrying out suction filtration, washing the filter cake with water for 3-4 times, washing with absolute ethyl alcohol for 1 time, and drying at about 60 ℃ to obtain 0.86kg of the Taylovone crystal C.

A micrograph of crystalline tylravone C is shown in fig. 7.

An X-ray powder diffraction pattern of crystalline form C of tirilavone is shown in figure 8.

TABLE 6X-ray powder diffraction data of crystalline Talaravone C

2θ	d value	Relative intensity/%)
			7.165	12.327	2.5
10.607	8.334	100.0
			12.810	6.905	69.2
14.371	6.158	27.1
			17.775	4.986	9.2
18.662	4.751	8.2
			21.355	4.157	10.8
25.529	3.486	22.1
			26.534	3.357	14.9
27.090	3.289	15.5
			32.219	2.776	5.6

Claims

1. A crystalline Talaravone A characterized in that: the 2 theta angle of a characteristic peak of the tylorravone crystal A with 100% of relative intensity in an X-ray powder diffraction pattern is 8.35 +/-0.2 degrees.

2. A method for preparing the crystalline tylravone a of claim 1, which is characterized in that: the method comprises the following steps: at room temperature, adding the tylorravone into an organic solvent, adding concentrated ammonia water to dissolve the tylorravone, adding organic acid, and stirring to separate out a solid; filtering, washing with absolute ethyl alcohol, purified water and absolute ethyl alcohol in sequence, and drying to obtain a tylorravone crystal A; wherein the organic solvent is an organic solvent which can be mutually dissolved with ammonia water; the mass ratio of the organic solvent to the tyraravone is 8-10: 1, the mass ratio of the organic solvent to the ammonia water is 2-2.5: 1, and the mass ratio of the organic solvent to the organic acid is 1.5-2: 1.

3. The preparation method of tylravone crystal a according to claim 2, characterized in that: the organic solvent is selected from one or more of methanol, ethanol or dimethylformamide; the organic acid is formic acid or acetic acid.

4. A method for preparing the crystalline tylravone a of claim 1, which is characterized in that: the method comprises the following steps: heating and dissolving the tylorravone in dimethylformamide, cooling to room temperature, and separating out a solid; filtering, washing a filter cake with dimethylformamide and ethanol in sequence, then soaking with purified water and ethanol in sequence, draining, and drying to obtain a tylorravone crystal A; the mass-volume ratio of the tylorravone to the dimethylformamide is 1: 8-10.

5. A crystalline Talaravone B characterized in that: the 2 theta angle of a characteristic peak of the tyloruone crystal B with 100% relative intensity in an X-ray powder diffraction pattern is 12.88 +/-0.2 degrees.

6. A crystalline Talaravone C characterized in that: the 2 theta angle of an absorption peak of the tyloruone crystal C with 100% relative intensity of an X-ray powder diffraction pattern is 10.61 +/-0.2 degrees.

7. A pharmaceutical composition characterized by: the pharmaceutical composition contains a pharmacologically effective amount of the terepratvone crystal A of claim 1, the terepratvone crystal B of claim 5, or the terepratvone crystal C of claim 6, and a pharmaceutically acceptable excipient.

8. An improved synthesis process of tylorravone, which is characterized in that: the method comprises the following steps: taking a saturated hydrogen chloride ethanol solution as a reaction solvent, and condensing and cyclizing 2-cyanobenzene hydrazine and methyl acetoacetate in the saturated hydrogen chloride ethanol solution to generate the tylorravone; wherein the molar ratio of the 2-cyanophenylhydrazine to the methyl acetoacetate is 0.8-1: 1.

9. The synthesis process of teriravone according to claim 8, wherein: the reaction temperature was room temperature.

10. The synthesis process of teriravone according to claim 8, wherein: after the reaction is finished, cooling the reaction liquid to about 5 ℃, adding ice water, stirring to separate out a product, filtering, and washing by adopting 33% ethanol and absolute ethanol in sequence to obtain a pure Tairui ravone product.