WO2022110842A1 - Crystal form a of levorotatory fluoropyrrolidone and preparation method therefor - Google Patents

Abstract

Disclosed are a crystal form A of levorotatory fluoropyrrolidone and a preparation method therefor. In a powder X-ray diffraction pattern of the crystal form A, the crystal form A has a characteristic peak at a diffraction angle 2θ, wherein the diffraction angle 2θ has a value of 5.52°±0.2°, 11.06°±0.2°, 14.84°±0.2°, 15.78°±0.2°, 16.64°±0.2°, 17.28°±0.2°, 18.34°±0.2°, 19.19°±0.2°, 20.12°±0.2°, 20.67°±0.2°, 22.57°±0.2°, 23.24°±0.2°, 23.80°±0.2°, 25.24°±0.2°, 25.86°±0.2°, 27.08°±0.2°, 27.82°±0.2°, 28.60°±0.2°, 30.16°±0.2°, 30.94°±0.2°, 32.26°±0.2°, 33.66°±0.2°, 34.14°±0.2°, 35.32°±0.2°, 36.21°±0.2°, 37.16°±0.2° and 38.06°±0.2°. The crystal form A has the advantages of good chemical stability, no hygroscopicity, thermodynamic stability and the like, and can be used for subsequent crystal form research and development.

Description

Crystal form A of L-fluoropyrrolidone and preparation method thereof technical field

The invention relates to a crystal form A of levoflopyrrolidone and a preparation method thereof, belonging to the field of pharmaceutical crystal forms.

Background technique

L-fluoropyrrolidone, its chemical formula is C ₁₈ H ₁₆ FNO ₂ , its Chinese name is (S)-5-benzyl-4-((4-fluorobenzyl)oxo)-1H-pyrrole-2(5H )-ketone. L-fluoropyrrolidone is an inhibitor of Rac1 (Ras-related C3 botulinum toxin substrate 1, Ras-related C3 botulinum toxin substrate 1) activity, which may prevent the complexation of RhoGDI, Coronin1A and Pak-1 by binding to F-actin in the cytoplasm The formation of the body, thereby inhibiting the activation of the juxtamembrane domain Rac1, and thus rescue the memory deficit. A number of pharmacodynamic trials have shown that its efficacy is superior to donepezil.

Alzheimer's disease (AD) is a common cause of dementia and one of the leading causes of complications and death in an aging population. Neuropathological hallmarks of AD include amyloid plaques formed by the deposition of extracellular Aβ protein and neurofibrillary tangles formed by the aggregation of intracellular phosphorylated Tau protein. In addition, the inflammatory response in the patient's brain increased, the metabolic rate decreased, and neuronal death and brain atrophy occurred. Severe brain damage results in cognitive impairment and motor deficits, and dramatically reduces life expectancy. Due to the loss of cognitive function and mobility, patients cannot take care of themselves and can only receive professional care, which brings a great financial burden to the family and society.

The prevalence of AD increases with age. Generally speaking, after the age of 60, the incidence of dementia doubles for every 10-year increase in age, and the burden of treatment gradually increases. With the intensification of aging in China, AD patients are increasing year by year, and there are currently more than 6 million patients in China. The team of Professor Jia Jianping from Xuanwu Hospital of Capital Medical University conducted a survey on the per capita annual expenditure of AD patients in China in 2015 and found that the total expenditure of AD patients in my country has reached 167.74 billion US dollars, accounting for 1% of China's GDP, and this number may increase by 2030. to $507.49 billion.

Since the first AD patient was reported more than 100 years ago, countless scientists have conducted drug research and development for the disease, and marketed two types of therapeutic drugs, namely cholinesterase inhibitors and glutamate receptor antagonists. However, these clinical drugs The curative effect is relatively weak. In addition, scientists' antibodies against the toxic proteins Aβ and tau proteins have mostly failed, and most of the drugs against inflammation have been eliminated. Recently, the Chinese drug regulatory authority has conditionally approved GV-971, a new anti-AD drug that reduces brain inflammation through intestinal flora, but it may still be required to be supplemented after listing due to its weak improvement in patients' cognition. In efficacy studies, AD patients still face the situation that no specific drugs are available.

Previous studies have found that the Rac1 protein is involved in the physiological regulation of active forgetting. The higher the activity of this protein, the faster the memory forgetting speed, and the lower the activity, the better the memory is maintained. In addition, in clinical studies, it was found that the activity of Rac1 protein in the brain tissue (hippocampus brain region) of AD patients was significantly abnormally increased, and it was directly related to memory impairment, suggesting that the increased activity of Rac1 in patients caused accelerated forgetting of memory, which in turn led to memory impairment. In related basic research, it was found that the increase of Rac1 activity led to spatial memory deficits in AD flies and AD mice, and inhibition of Rac1 activity could improve cognitive impairment in AD animal models. Based on this, two AD models of Drosophila and double transgenic mice were used to evaluate the pharmacodynamics of a large number of active compounds, and it was found that JKF-012 inhibited the activity of the amnestic molecule Rac1, and also had excellent curative effect and huge effect in animal models. in-depth development value. Based on the structural modification of JFK-012, 200 derivatives were synthesized and evaluated using two AD models of Drosophila and double transgenic mice, and several new chemical entities with the best efficacy were found. After six of them were evaluated for druggability, pharmacokinetics, and in vitro and in vivo toxicology, levoflopyrrolone stood out due to its excellent pharmacokinetic properties, blood-brain barrier permeability, and low toxicity. L-fluropyrrolone has a positive effect in rescue of memory deficits in Drosophila and mouse models. Since L-fluropyrrolidone belongs to a new chemical entity, it is still uncertain which crystal form it exists in and how stable the different crystal forms are. Therefore, it is necessary to study its crystal form.

SUMMARY OF THE INVENTION

The object of the present invention is to provide a crystal form A of L-fluoropyrrolidone, the crystal form A is a bulk crystal with a particle size of about 50 μm, and has good physical and chemical stability.

The crystal form A of L-fluoropyrrolidone provided by the present invention has characteristic peaks at the diffraction angle 2θ in the powder X-ray diffraction diagram, wherein the values of the diffraction angle 2θ are 5.52°±0.2°, 11.06°±0.2° , 14.84°±0.2°, 15.78°±0.2°, 16.64°±0.2°, 17.28°±0.2°, 18.34°±0.2°, 19.19°±0.2°, 20.12°±0.2°, 20.67°±0.2°, 22.57 °±0.2°, 23.24°±0.2°, 23.80°±0.2°, 25.24°±0.2°, 25.86°±0.2°, 27.08°±0.2°, 27.82°±0.2°, 28.60°±0.2°, 30.16°± 0.2°, 30.94°±0.2°, 32.26°±0.2°, 33.66°±0.2°, 34.14°±0.2°, 35.32°±0.2°, 36.21°±0.2°, 37.16°±0.2°, 38.06°±0.2° ;

The differential scanning calorimetry (DSC) spectrum of the crystal form A has an endothermic peak between 117°C and 119°C;

When the crystal form A is heated to 110° C., the weight loss is only 1.4%, so the crystal form A is an anhydrous crystal form.

The equilibrium solubility of the crystal form A in H ₂ O is about 0.077 mg/mL, and the crystal form has not changed; DVS test results show that the crystal form A sample has no hygroscopicity; in the PLM test, all The crystal form A is a bulk crystal with a particle size of about 50 μm, and there is agglomeration phenomenon at the same time; the stability test shows that the crystal form A has good physical and chemical stability.

The present invention also provides the preparation method of the crystal form A of L-fluoropyrrolidone, which is specifically the following:

The first method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolving L-fluoropyrrolidone in a good solvent to obtain a clear solution, then adding an anti-solvent dropwise to the clear solution, crystallizing under stirring conditions, and filtering and drying to obtain the crystal of the L-fluoropyrrolidone. Type A;

If there is no solid precipitation after adding about 10.0 mL of the anti-solvent, the test is stopped;

The good solvent and the anti-solvent are any of the following 1)-4):

1) the good solvent is any one in methanol, 4-methyl-2-pentanone, ethyl acetate, 1,4-dioxane and dichloromethane; the anti-solvent is n-heptane;

2) the good solvent is any one in isopropanol, acetonitrile, dimethyl sulfoxide, N-methylpyrrolidone, methanol and N,N-dimethylacetamide; the anti-solvent is water;

3) the good solvent is any one in chloroform, isopropyl acetate and 2-methyltetrahydrofuran, and the anti-solvent is n-hexane;

4) the good solvent is any one in toluene, acetone and tetrahydrofuran, and the anti-solvent is cyclohexane;

The dosage of the good solvent is 0.1-0.4 mL/15 mg of L-fluoropyrrolidone.

The second method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolving L-fluoropyrrolidone in a good solvent to obtain a clear solution, then adding the clear solution dropwise to an anti-solvent, crystallizing under stirring conditions, filtering and drying to obtain the L-fluoropyrrolidone solution. Form A;

The good solvent and the anti-solvent are any of the following 1)-3):

1) the good solvent is any one in chloroform and toluene, and the anti-solvent is n-heptane;

2) the good solvent is any one of methanol, 1,4-dioxane and acetone; the anti-solvent is water;

3) the good solvent is any one in ethylene dichloride and isopropyl acetate, and the anti-solvent is cyclohexane;

The dosage of the good solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone;

The volume ratio of the good solvent to the anti-solvent is 0.2-0.4:5.

The third method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolve L-fluoropyrrolidone in an organic solvent, take the supernatant after sonicating or shaking; place the supernatant in a container and seal it with a film, the film is provided with 3 to 5 through holes, and then place at room temperature Slowly volatilize and crystallize, and obtain the crystal form A of the L-fluoropyrrolidone after filtration and drying;

The organic solvent is methanol, ethanol, isopropanol, acetonitrile, acetone, ethyl acetate, toluene, tetrahydrofuran, 2-methyltetrahydrofuran, isopropyl acetate, methyl tert-butyl ether, dichloromethane and chloroform any of them;

The room temperature refers to a temperature of 20 to 25°C;

The dosage of the organic solvent is 0.1-0.6 mL/15 mg of L-fluoropyrrolidone.

The fourth method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolve L-fluoropyrrolidone in an organic solvent, stir and take the supernatant; cool the supernatant from 50°C to 5°C at a speed of 0.1°C/min and keep the temperature at 5°C until a solid is precipitated, after filtration and drying Obtain the crystal form A of described L-fluoropyrrolidone;

Described organic solvent is methyl tertiary butyl ether, ethanol and n-heptane volume ratio of 1:9 mixed solution, acetone and cyclohexane volume ratio of 1:9 mixed solution, ethyl acetate and cyclohexane volume At least one of a mixed solution with a ratio of 1:9 and a mixed solution with a volume ratio of acetonitrile to water of 1:4;

The dosage of the organic solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone, preferably 0.3 mL/15 mg of L-fluoropyrrolidone.

The fifth method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolving L-fluoropyrrolidone in organic solvent I or organic solvent II, stirring at room temperature for 9-11 days or at 50°C for 6-11 days, and collecting the solid by centrifugation is the crystal form A of the L-fluoropyrrolidone;

The organic solvent I is a mixture of n-heptane, n-hexane, cyclohexane, water, methyl tert-butyl ether, methanol and water in a volume ratio of 1:9, and a mixture of ethanol and water in a volume ratio of 1:9 liquid, a mixed solution with a volume ratio of acetone and water of 1:9, a mixed solution with a volume ratio of acetonitrile and water of 1:9, a mixed solution of isopropanol and n-heptane with a volume ratio of 1:9, isopropyl acetate and The mixed solution of n-heptane volume ratio of 1:9, the mixed solution of toluene and n-heptane volume ratio of 1:9, the mixed solution of methyl tert-butyl ether and cyclohexane volume ratio of 1:9, toluene and Any one of the mixed solution whose volume ratio of cyclohexane is 1:9 and the mixed solution whose volume ratio of 2-methyltetrahydrofuran and cyclohexane is 1:9;

When using the organic solvent I, stirring is carried out at room temperature;

The organic solvent II is a mixed solution of n-heptane, n-hexane, cyclohexane, water, isopropanol and n-heptane in a volume ratio of 1:9, and a mixed solution of ethanol and n-heptane in a volume ratio of 1:9 , a mixed solution with a volume ratio of acetonitrile and n-heptane of 1:9, and a mixed solution with a volume ratio of ethyl acetate and cyclohexane of 1:9;

When using the organic solvent II, stirring is carried out under the condition of 50 °C;

The dosage of the organic solvent I or the organic solvent II is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone, preferably 0.3 mL/15 mg of L-fluoropyrrolidone.

The sixth method for preparing crystal form A provided by the present invention comprises the following steps:

The L-fluoropyrrolidone is placed in an open container I, then the container I is placed in a container II filled with an organic solvent, the container II is sealed and left standing at room temperature for 11 to 18 days, filtered and dried. Then obtain the crystal form A of described L-fluoropyrrolidone;

The organic solvent is any one of water, acetone, methyl tert-butyl ether, ethyl acetate, isopropanol, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, acetonitrile and dichloromethane .

The seventh method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolve L-fluoropyrrolidone solvent in a good solvent, stir and take the supernatant; then place the vial in a container filled with anti-solvent, seal the container and let stand at room temperature for 11 to 18 days, filter and drying to obtain the crystal form A of the L-fluoropyrrolidone;

The good solvent and the anti-solvent are any of the following 1)-3):

1) the good solvent is any one in isopropanol, methyl isobutyl ketone, 1,4-dioxane and isopropyl acetate, and the anti-solvent is n-heptane;

2) the good solvent is any one in acetone, ethanol, acetonitrile and methanol, and the anti-solvent is water;

3) the good solvent is 2-methyltetrahydrofuran or chloroform, and the anti-solvent is cyclohexane;

The dosage of the good solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone;

The volume ratio of the good solvent to the anti-solvent is 0.2-0.4:4.

The eighth method for preparing crystal form A provided by the present invention comprises the following steps:

Dissolving L-fluoropyrrolidone in a good solvent to obtain a clear solution, then adding a polymer to the clear solution, placing it at room temperature to slowly volatilize and crystallize, filtering and drying to obtain the crystal form A of the L-fluoropyrrolidone ;

The good solvent and the polymer are any of the following 1)-2):

1) Described good solvent is any in acetone, chloroform, isopropanol and acetonitrile; Described polymer is polyvinylpyrrolidone, polyvinyl alcohol, polyvinyl acetate, polyvinyl chloride, polyvinyl acetate or hydroxyl Mixture of propyl methylcellulose and methylcellulose (mass ratio can be 1:1);

2) described good solvent is any in methanol, isopropyl acetate, methyl tert-butyl ether and toluene; described polymer is polycaprolactone, polyethylene glycol, polymethyl methacrylate or A mixture of sodium alginate and hydroxyethyl cellulose (mass ratio can be 1:1).

Description of drawings

Fig. 1 is the powder diffraction XRPD spectrum of the L-fluoropyrrolidone sample used in the embodiment of the present invention.

Fig. 2 is the DSC spectrogram of the L-fluoropyrrolidone sample adopted in the embodiment of the present invention.

Fig. 3 is the TGA spectrum of the L-fluoropyrrolidone sample adopted in the embodiment of the present invention.

Figure 4 is a XRPD comparison diagram of Form A and Form B.

FIG. 5 is an XRPD pattern of Form A. FIG.

FIG. 6 is a TGA/DSC chart of Form A. FIG.

FIG. 7 is an XRPD pattern of Form B. FIG.

Figure 8 is the XRPD pattern of Form B after drying at 5°C.

FIG. 9 is a XRPD comparison diagram of Form B before and after drying.

Figure 10 is the TGA/DSC chart of the crystal form B after drying at 5°C.

Figure 11 is an XRPD overlay of an anhydrous A/B suspension competing for separation solids.

Figure 12 is an XRPD overlay of Form A before and after stable placement.

FIG. 13 is a DVS diagram of Form A. FIG.

Figure 14 is a comparison diagram of the crystal form before and after the DVS test of the crystal form A.

FIG. 15 is a PLM image of Form A. FIG.

Figure 16 is a XRPD comparison chart of Form A after equilibrating in pure water for 24 hours.

Figure 17 is a XRPD comparison chart of Form B after equilibrating in pure water for 24 hours.

Detailed ways

The experimental methods used in the following examples are conventional methods unless otherwise specified.

The materials, reagents, etc. used in the following examples can be obtained from commercial sources unless otherwise specified.

In the following examples, the XRPD pattern was collected on the X-ray powder diffraction analyzer of PANalytical, and the XRPD test parameters are shown in Table 1.

Table 1 XRPD test parameters

In the following examples, TGA and DSC spectra were collected on a TAQ500/5000 thermogravimetric analyzer and a TAQ200/2000 differential scanning calorimeter, respectively, and the test parameters are shown in Table 2.

Table 2 DSC and TGA test parameters

In the following examples, polarized light microscopy (PLM) polarized light microscopy data were collected at room temperature with an Axio Lab.A1 upright microscope.

In the following examples, Dynamic Moisture Sorption (DVS) curves were collected on DVS Intrinsic of SMS (Surface Measurement Systems). The relative humidity at 25°C was corrected for the deliquescence points of LiCl, Mg( _NO3 ) ₂ and KCl. DVS test parameters are shown in Table 3.

Table 3 DVS test parameters

In the following examples, high performance liquid chromatography (HPLC) high performance liquid chromatography was collected on Agilent 1260 HPLC, and the instrument and test parameters were shown in Table 4.

Table 4 HPLC parameters for purity and solubility tests

In the following examples, the L-fluoropyrrolidone sample adopted is prepared according to the following method:

1) Preparation of Intermediate 1

Dissolve L-phenylalanine (20 g, 121.07 mmol) in 500 mL of a mixed solution of tetrahydrofuran and water (v/v 1:1), stir under ice bath for 10 minutes, add sodium hydroxide (10.7 g, 266.4 mmol) and di-tert-butyl dicarbonate (29.1 g, 133.2 mmol) were added and stirred at room temperature for 10 hours. TLC monitoring showed that the reaction was complete. The tetrahydrofuran was evaporated under reduced pressure, 500 mL of dichloromethane was added, 2N aqueous hydrochloric acid solution was added dropwise with stirring until the pH value of the aqueous layer was about 5, the organic layer was separated and washed once with 200 mL of saturated brine, dried over anhydrous magnesium sulfate, filtered and reduced in size. The solvent was evaporated to dryness under pressure to obtain a crude product with a yield of 99%, and the next reaction was carried out without further purification.

2) Preparation of Intermediate 2

Intermediate 1 (25 g, 94.2 mmol) was dissolved in 500 mL of anhydrous dichloromethane, followed by adding Michaelis acid (14.9 g, 103.7 mmol), 4-dimethylaminopyridine (17.3 g, 141.3 mmol), under ice bath After stirring for 10 minutes, a solution of dicyclohexylcarbodiimide (21.4 g, 103.7 mmol) in dichloromethane (100 mL) was added dropwise. After the dropwise addition, the reaction solution was continued to be stirred under an ice bath for 10 hours, and the completion of the reaction was monitored by TLC. Filter, wash the filtrate with 5% potassium hydrogen sulfate aqueous solution 6 times, each 200mL, wash with saturated brine once, dry over anhydrous sodium sulfate, evaporate the solvent under reduced pressure to obtain a light yellow solid, add 500mL petroleum ether and stir and filter to obtain the target The product was a white solid (30 g, 77% yield), which was carried to the next step without further purification.

3) Preparation of Intermediate 3

Intermediate 2 (25 g, 63.9 mmol) was dissolved in 400 mL of ethyl acetate and reacted under reflux for 5 hours. It was cooled to room temperature, and the ethyl acetate was evaporated to dryness under reduced pressure to obtain the target product (17.6 g, 95%) as a white solid, which did not require further purification.

4) Preparation of intermediate 15

Intermediate 3 (289 mg, 1.0 mmol) was dissolved in 5 mL of acetonitrile, K ₂ CO ₃ (207 mg, 1.5 mmol) and 4-fluorobenzyl bromide (227 mg, 1.2 mmol) were sequentially added at room temperature, and the reaction was refluxed for 5 hours. After cooling to room temperature, the solid was removed by filtration, the filtrate was evaporated to dryness, and the intermediate 15 was purified by column chromatography to obtain intermediate 15 as a colorless oil (158 mg, yield 40%). Molecular weight: 397.45. ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.34 (dd, J=8.2, 5.4 Hz, 2H), 7.19 (m, 3H), 7.10 (t, J=8.5 Hz, 2H), 6.98 ( m, 2H), 4.91(s, 1H), 4.84(m, 2H), 4.71(dd, J=5.3, 3.0Hz, 1H), 3.43(dd, J=14.0, 5.2Hz, 1H), 3.14(dd , J=14.0, 2.8Hz, 1H), 1.59(s, 9H). ¹³ C NMR(100MHz, CDCl ₃ )δ174.79, 168.75, 163.06(d, J=248.3Hz), 149.47, 134.24, 130.21(d, J =8.4Hz), 130.05(d, J=3.3Hz), 129.59, 128.35, 127.11, 115.93(d, J=21.7Hz), 96.18, 82.78, 72.65, 60.38, 35.55, 28.30. ¹⁹ F NMR(376MHz, CDCl ₃ ) δ-112.20.

5) Preparation of L-fluoropyrrolidone

Intermediate 15 (397 mg, 1.0 mmol) was dissolved in 10 mL of dichloromethane, trifluoroacetic acid (223 μL, 3.0 mmol) was added under ice bath, and the reaction solution was stirred at room temperature for 5 hours. After the completion of the reaction, 90 mL of dichloromethane was added to dilute the reaction solution, washed with 10% NaHCO ₃ aqueous solution once, washed with saturated brine once, dried over anhydrous sodium sulfate, filtered, evaporated to dryness, and separated and purified by column chromatography to obtain formula 1: white Solid 360 mg, 82% yield. Molecular weight: 297.33. ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.37 (dd, J=8.4, 5.3 Hz, 2H), 7.27 (m, 3H), 7.18 (m, 2H), 7.11 (t, J=8.5 Hz, 2H), 5.77(s, 1H), 5.06(s, 1H), 4.95(m, 2H), 4.27(dd, J=9.2, 3.6Hz, 1H), 3.21(dd, J=13.7, 3.7Hz) , 1H), 2.67 (dd, J=13.7, 9.0 Hz, 1H). ¹³ C NMR (100 MHz, CDCl ₃ ) δ 175.94, 173.55, 163.04 (d, J=247.8 Hz), 136.55, 130.64 (d, J=3.5 Hz), 130.06(d, J=8.4Hz), 129.27, 128.79, 127.17, 115.91(d, J=21.7Hz), 95.23, 72.60, 58.86, 38.77. ¹⁹ F NMR(376MHz, CDCl ₃ )δ-112.58.

The powder diffraction XRPD, DSC and TGA spectra of L-fluoropyrrolidone samples are shown in Figures 1 to 3, respectively.

Example 1. Preparation of crystal form

The present invention adopts the methods of anti-solvent addition, anti-anti-solvent addition, slow volatilization, slow cooling, suspension stirring, gas-solid infiltration and gas-liquid infiltration to set up 96 polymorph screening tests. The specific test methods and results are shown in Table 5. , a total of 2 crystal forms were found, named as crystal form A and crystal form B.

Table 5 Polymorph screening test methods and results

1. Anti-solvent addition test

A total of 17 anti-solvent addition experiments were set up. Weigh about 15 mg of each L-fluoropyrrolidone sample into a 20 mL vial, dissolve it with 0.1-0.4 mL of good solvent, add the anti-solvent in Table 6 dropwise to the clear solution, and stir while adding dropwise to the solution. A solid precipitated out. If there is no solid precipitation after adding about 10.0 mL of anti-solvent, the test is stopped, and the solid is collected and tested by XRPD. The test results are shown in Table 6, and the crystal form A was obtained by the anti-solvent addition test.

Table 6 Anti-solvent addition test method and results

2. Anti-anti-solvent addition test

A total of 8 anti-antisolvent addition experiments were set up. Weigh about 15 mg of each L-fluoropyrrolidone sample into a 3 mL vial, dissolve it with 0.2 to 0.4 mL of a good solvent, and quickly add the above solution to a 20 mL glass bottle containing 5 mL of the anti-solvent in Table 7. Add dropwise with stirring until solids are precipitated. The solution without precipitation of solids was transferred to stand at 5°C, and if there was still no solid precipitation, it was transferred to stand at -20°C. The test results are shown in Table 7, and the crystal form A/B was obtained by the anti-anti-solvent addition test.

Table 7 Anti-anti-solvent addition test method and results

*: The solution was clear, no solid was precipitated after transferring to 5°C and stirring, and then it was left open to volatilize and precipitated solid at room temperature.

A total of 13 slow evaporation experiments were set up. Weigh about 15 mg of each L-fluoropyrrolidone sample into a 3 mL vial, add 0.1 to 0.6 mL of solvent respectively, ultrasonically or vibrate to dissolve the sample as completely as possible, and take the clear liquid after filtration. The vial was sealed with parafilm and 3-5 pinholes were pricked on it, and it was slowly evaporated at room temperature, and the obtained solid was collected and tested by XRPD. The test results are shown in Table 8, and the crystal form A was obtained in the slow volatilization crystallization test.

Table 8 Slow volatilization test method and results

4. Slow down the temperature

A total of 6 slow cooling experiments were set up. Weigh about 15 mg of each sample of L-fluoropyrrolidone into a 1.5 mL vial, add 0.3 mL of the solvent in Table 9, stir at 50 °C for about 2 hours, and filter the supernatant. The temperature was lowered from 50°C to 5°C/min at a constant temperature of 5°C, the precipitated solids were collected and tested by XRPD, and the samples without precipitation of solids were transferred to -20°C and stirred. The test results are shown in Table 9, and the crystal form A was obtained by the slow cooling test.

Table 9 Slow cooling test method and results

*: No solid was precipitated after stirring at -20°C for 4 days, transferred to room temperature and volatilized for 3 days. A4 and A6 precipitated solid, and A3 turned into oil. 0.3 mL of the solvents listed in Table 10 were added to HPLC vials, respectively, and the resulting suspension was stirred at room temperature for about 9 to 11 days, and then the solid was collected by centrifugation and tested by XRPD. The test results are shown in Table 10, and the crystal form A was obtained by the suspension stirring test.

Table 10 Room temperature suspension test method and results

6. Suspension stirring at 50°C

A total of 9 suspension stirring tests at 50°C were set up. About 15 mg of each sample of L-fluoropyrrolidone was weighed into an HPLC vial, 0.3 mL of the solvent listed in Table 11 was added, and the resulting suspension was stirred at 50 °C for 6 to 11 days, and the solid was collected by centrifugation and tested by XRPD . The test results are shown in Table 11, and the crystal form A was obtained in the suspension stirring test.

Table 11 50℃ suspension stirring test method and results

^# : Compared with the crystal form A, the XRPD pattern of this sample has one more diffraction peak at ~6.3°. The mL vials were placed open in 20 mL vials containing different solvents, the 20 mL vials were sealed and allowed to stand at room temperature for 11 to 18 days. The solids were collected and tested by XRPD. The test results are shown in Table 12, and the crystal form A was obtained in the gas-solid permeation test.

Table 12 Gas-solid penetration test method and results

*: No visible solid, the solution was left at room temperature to evaporate and crystallize.

^# : Compared with the crystal form A, the XRPD pattern of this sample has one more diffraction peak at ~6.3°.

In 0.2 mL of good solvent, filter the supernatant into a 3 mL vial, take another 20 mL vial and add about 4 mL of anti-solvent to it, place the above 3 mL vial open in a 20 mL vial, seal the 20 mL vial and store it at room temperature. Let stand. When solid precipitation is observed, take out the solid for XRPD measurement. If there is no solid precipitation, take out and volatilize at room temperature until the solvent evaporates to dryness, collect the solid and perform XRPD measurement. The test results are shown in Table 13, and the crystal form A was obtained by the gas-liquid diffusion test.

Table 13 Gas-liquid diffusion test method and results

*: No visible solid, the solution was left at room temperature to evaporate and crystallize.

^# : Compared with the crystal form A, the XRPD pattern of this sample has one more diffraction peak at ~6.3°.

If as shown in Table 14, Form A was obtained in the polymer-induced crystallization test.

Table 14 High polymer induction test method and results

It can be seen that there are absorption peaks at 2θ in the XRPD spectrum of Form A: 5.522, 11.059, 14.842, 15.799, 16.640, 17.279, 18.339, 19.180, 20.119, 20.660, 22.560, 23.239, 23.800, 25.240, 25.860 , 27.079, 27.819, 28.598, 30.160, 30.940, 32.261, 33.661, 34.141, 35.320, 36.205, 37.160, 38.060.

The TGA/DSC results are shown in Figure 6. It can be seen that the weight loss is 1.4% when heated to 110°C, and there is an endothermic peak at 117.8°C (initial temperature). According to the small weight loss of TGA before heating to decomposition, Form A is presumed to be an anhydrous form.

2. Form B

The XRPD results of Form B (814901-08-A1) are shown in Figure 7 . After the sample of crystal form B was dried at 5°C for 1 hour, it began to transform to crystal form A. The XRPD results are shown in Figure 8. After standing at room temperature overnight, it partially transformed into crystal form A to form a mixture of crystal form A and crystal form B, as shown in FIG. 9 . The TGA/DSC results of the sample after drying at 5°C for 1 hour (Figure 10) showed a weight loss of 1.3% when heated to 110°C; the sample had a broad exothermic peak at 86.8°C (peak) and was heated to 118.0°C (initial temperature), there is a sharp endothermic peak. Combined with the XRPD pattern and the DSC results of crystal form A, it is speculated that this exothermic peak is the signal of the transformation of crystal form B to crystal form A. According to the small weight loss of TGA before heating to decomposition, it is presumed that Form B is an anhydrous form.

Example 3. Thermodynamic stability study

Suspension competition tests at room temperature and 50°C were carried out for the crystal forms A+B.

Weigh approximately 4.5 mg of Form B sample into an HPLC vial containing a saturated Form A sample suspension, and separate after approximately 4 hours at room temperature (25°C ± 3°C) and 50°C with magnetic stirring (~1000 rpm). Solid and tested XRPD. The test results are shown in Table 15, and the XRPD characterization results (FIG. 11) show that the crystal forms A+B are all converted into the crystal form A after being stirred in the H ₂ O and MTBE systems. The test results show that, in the range from room temperature to 50 °C, compared with the crystal form B, the crystal form A is a more thermodynamically stable anhydrous crystal form.

Table 15 Suspension competition stirring test and results between anhydrous crystals A+B

Example 4. Research on the physicochemical properties of crystal form A

The physicochemical stability, hygroscopicity and particle morphology of Form A were determined, and the 24-hour equilibrium solubility of Form A and Form B in pure water was determined.

1. Physical and chemical stability of Form A

The solid samples of Form A were placed at 80°C/24 hours, 25°C/60%RH/1 week and 40°C/75%RH/1 week, respectively, and their chemical purity and solidity were evaluated by HPLC area purity and XRPD characterization results The change of crystal form, the results are shown in Table 16.

The purity/initial purity (area)% of the samples of crystal form A placed at 80°C for 1 day and at 25°C/60%RH and 40°C/75%RH for 1 week were both 100.0%. The three conditions The lower solid crystal form did not change, indicating that the crystal form A has better physical and chemical stability. The XRPD overlay of the solid is shown in Figure 12.

Table 16 Physical and chemical stability results of crystal form A

2. The hygroscopicity of Form A

The hygroscopicity of Form A was evaluated by Dynamic Moisture Sorption (DVS) testing.

At a constant temperature of 25 °C, the DVS test results are shown in Figure 13. Figure 13 shows that when the humidity increases from 0% to 80% RH, the sample of crystal form A has a hygroscopic weight gain of 0.03%, indicating that crystal form A has no hygroscopicity. The XRPD results (Figure 14) show that the XRPD of the solids before and after DVS is basically the same.

3. Characterization of particle morphology and size of crystal form A

The crystal form A was characterized and tested by polarized light microscopy (PLM), as shown in Figure 15, the batch of crystal form A samples were bulk crystals accompanied by agglomeration, and the particle size was about 50 μm.

4. Equilibrium solubility of crystal-free A and B in pure water

At room temperature, the two crystal forms (crystal form A and crystal form B) were equilibrated with magnetic stirring in pure water for 24 hours, and then the supernatant liquid and the lower solid were separated by centrifugation. The obtained supernatant was filtered through a 0.22 μm filter to test the solubility, and the isolated solid was tested by XRPD to test whether the crystal form changed. The results are shown in Table 17, and the XRPD characterization results are shown in Figures 16 and 17. It can be seen that after the two crystal forms are stirred and equilibrated in pure water for 24 hours, the crystal forms are both crystal form A.

Table 17 24-hour equilibrium solubility of anhydrous crystals in pure water

It can be seen from the above test results that the crystal form A prepared by the present invention has good stability, which is beneficial to the storage of the raw drug and the development of the preparation; the crystal form with good stability can effectively avoid the phenomenon of crystal transformation and Bioavailability effects due to transformation.

Industrial application

The crystal form A of L-fluoropyrrolidone provided by the present invention has the advantages of good chemical stability, no hygroscopicity, thermodynamic stability and the like, and can be used for subsequent crystal form research and development.

Claims (17)

1. The crystal form A of L-fluoropyrrolidone represented by formula I has characteristic peaks at the diffraction angle 2θ in the powder X-ray diffraction pattern, wherein the values of the diffraction angle 2θ are 5.52°±0.2°, 11.06°±0.2°, 14.84°±0.2°, 15.78°±0.2°, 16.64°±0.2°, 17.28°±0.2°, 18.34°±0.2°, 19.19°±0.2°, 20.12°±0.2°, 20.67°±0.2°, 22.57° ±0.2°, 23.24°±0.2°, 23.80°±0.2°, 25.24°±0.2°, 25.86°±0.2°, 27.08°±0.2°, 27.82°±0.2°, 28.60°±0.2°, 30.16°±0.2 °, 30.94°±0.2°, 32.26°±0.2°, 33.66°±0.2°, 34.14°±0.2°, 35.32°±0.2°, 36.21°±0.2°, 37.16°±0.2°, 38.06°±0.2°;

   
2. The crystal form A according to claim 1, wherein the differential scanning calorimetry analysis pattern of the crystal form A has an endothermic peak between 117°C and 119°C.
3. The application of the crystal form A of claim 1 or 2 in the preparation of a pharmaceutical crystal form.
4. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

   Dissolve L-fluoropyrrolidone in a good solvent to obtain a clear solution, then add anti-solvent dropwise to the clear solution, perform crystallization under stirring conditions, and obtain crystals of the L-fluoropyrrolidone after filtration and drying. Type A;

   The good solvent and the anti-solvent are any of the following 1)-4):

   1) the good solvent is any one in methanol, 4-methyl-2-pentanone, ethyl acetate, 1,4-dioxane and dichloromethane; the anti-solvent is n-heptane;

   2) the good solvent is any one in isopropanol, acetonitrile, dimethyl sulfoxide, N-methylpyrrolidone, methanol and N,N-dimethylacetamide; the anti-solvent is water;

   3) the good solvent is any one in chloroform, isopropyl acetate and 2-methyltetrahydrofuran, and the anti-solvent is n-hexane;

   4) The good solvent is any one of toluene, acetone and tetrahydrofuran, and the anti-solvent is cyclohexane.
5. The preparation method according to claim 4, wherein the amount of the good solvent is 0.1-0.4 mL/15 mg of L-fluropyrrolidone.
6. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

   Dissolving L-fluoropyrrolidone in a good solvent to obtain a clear solution, then adding the clear solution dropwise to an anti-solvent, crystallizing under stirring conditions, and filtering and drying to obtain the L-fluoropyrrolidone solution. Form A;

   The good solvent and the anti-solvent are any of the following 1)-3):

   1) the good solvent is any one in chloroform and toluene, and the anti-solvent is n-heptane;

   2) the good solvent is any one of methanol, 1,4-dioxane and acetone; the anti-solvent is water;

   3) The good solvent is any one of dichloroethane and isopropyl acetate, and the anti-solvent is cyclohexane.
7. The preparation method according to claim 6, wherein the amount of the good solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone;

   The volume ratio of the good solvent to the anti-solvent is 0.2-0.4:5.
8. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

   Dissolve L-fluoropyrrolidone in an organic solvent, take the supernatant after ultrasonication or vibration; place the supernatant in a container and seal it with a film, the film is provided with through holes, and then slowly volatilize and crystallize at room temperature , after filtration and drying, the crystal form A of the L-fluoropyrrolidone is obtained;

   The organic solvent is methanol, ethanol, isopropanol, acetonitrile, acetone, ethyl acetate, toluene, tetrahydrofuran, 2-methyltetrahydrofuran, isopropyl acetate, methyl tert-butyl ether, dichloromethane and chloroform any of them.
9. The preparation method according to claim 8, wherein the amount of the organic solvent is 0.1-0.6 mL/15 mg of L-fluoropyrrolidone.
10. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

    Dissolve L-fluoropyrrolidone in an organic solvent, stir and take the supernatant; cool the supernatant from 50°C to 5°C at a speed of 0.1°C/min and keep the temperature at 5°C until a solid is precipitated, after filtration and drying Obtain the crystal form A of described L-fluoropyrrolidone;

    Described organic solvent is methyl tertiary butyl ether, the mixed solution of ethanol and n-heptane volume ratio is 1:9, the mixed solution of toluene and n-heptane volume ratio is 1:9, the volume ratio of acetone and cyclohexane is At least one of a 1:9 mixture, a 1:9 mixture of ethyl acetate and cyclohexane, and a 1:4 mixture of acetonitrile and water.
11. The preparation method according to claim 10, wherein the amount of the organic solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone.
12. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

    Dissolving L-fluoropyrrolidone in organic solvent I or organic solvent II, stirring at room temperature for 9-11 days or at 50°C for 6-11 days, and collecting the solid by centrifugation is the crystal form A of the L-fluoropyrrolidone;

    The organic solvent I is a mixture of n-heptane, n-hexane, cyclohexane, water, methyl tert-butyl ether, methanol and water in a volume ratio of 1:9, and a mixture of ethanol and water in a volume ratio of 1:9 liquid, a mixed solution with a volume ratio of acetone and water of 1:9, a mixed solution with a volume ratio of acetonitrile and water of 1:9, a mixed solution of isopropanol and n-heptane with a volume ratio of 1:9, isopropyl acetate and n-heptane volume ratio of 1:9 mixed solution, methyl tert-butyl ether and cyclohexane volume ratio of 1:9 mixed solution, toluene and cyclohexane volume ratio of 1:9 mixed solution and 2- Methyltetrahydrofuran and cyclohexane volume ratio is any in the mixed solution of 1:9;

    The organic solvent II is a mixed solution of n-heptane, n-hexane, cyclohexane, water, isopropanol and n-heptane in a volume ratio of 1:9, and a mixed solution of ethanol and n-heptane in a volume ratio of 1:9 , a mixed solution with a volume ratio of acetonitrile and n-heptane of 1:9, and a mixed solution with a volume ratio of ethyl acetate and cyclohexane of 1:9.
13. The preparation method according to claim 12, wherein the amount of the organic solvent I or the organic solvent II is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone.
14. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

    The L-fluoropyrrolidone is placed in an open container I, then the container I is placed in a container II filled with an organic solvent, the container II is sealed and left standing at room temperature for 11 to 18 days, filtered and dried. Then obtain the crystal form A of described L-fluoropyrrolidone;

    The organic solvent is any one of water, acetone, methyl tert-butyl ether, ethyl acetate, isopropanol, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, acetonitrile and dichloromethane .
15. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

    Dissolve L-fluoropyrrolidone solvent in a good solvent, stir and take the supernatant; then place the vial in a container filled with anti-solvent, seal the container and let stand at room temperature for 11 to 18 days, filter and drying to obtain the crystal form A of the L-fluoropyrrolidone;

    The good solvent and the anti-solvent are any of the following 1)-3):

    1) the good solvent is any one in isopropanol, methyl isobutyl ketone, 1,4-dioxane and isopropyl acetate, and the anti-solvent is n-heptane;

    2) the good solvent is any one in acetone, ethanol, acetonitrile and methanol, and the anti-solvent is water;

    3) the good solvent is 2-methyltetrahydrofuran or chloroform, and the anti-solvent is cyclohexane.
16. The preparation method according to claim 15, wherein the amount of the good solvent is 0.2-0.4 mL/15 mg of L-fluoropyrrolidone;

    The volume ratio of the good solvent to the anti-solvent is 0.2-0.4:4.
17. The preparation method of the described crystal form A of claim 1 or 2, comprises the steps:

    Dissolving L-fluoropyrrolidone in a good solvent to obtain a clear solution, then adding a polymer to the clear solution, placing it at room temperature to slowly volatilize and crystallize, filtering and drying to obtain the crystal form A of the L-fluoropyrrolidone ;

    The good solvent and the polymer are any of the following 1)-2):

    1) Described good solvent is any in acetone, chloroform, isopropanol and acetonitrile; Described polymer is polyvinylpyrrolidone, polyvinyl alcohol, polyvinyl acetate, polyvinyl chloride, polyvinyl acetate or hydroxyl Mixtures of propyl methylcellulose and methylcellulose;

    2) described good solvent is any in methanol, isopropyl acetate, methyl tert-butyl ether and toluene; described polymer is polycaprolactone, polyethylene glycol, polymethyl methacrylate or A mixture of sodium alginate and hydroxyethyl cellulose.

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