WO2016070569A1 - Sofosbuvir crystal and preparation method thereof - Google Patents

Abstract

Disclosed are four novel sofosbuvir crystals, namely Crystal H1, Crystal H2, Crystal H3 and Crystal H4, and method for preparing the four crystals. The structures and unit cell parameters of the four novel sofosbuvir crystals are determined by employing a single crystal X-ray diffraction analysis. A diffraction angle 2θ of the Crystal H1 has characteristic peaks at 4.9°, 6.6°, 7.1°, 8.2°, 9.6°, 16.5°, 19.0°, 19.2°, 19.9° and 24.9°; a diffraction angle 2θ of the Crystal H2 has characteristic peaks at 6.7°, 7.0°, 9.7°, 19.1°, 19.3°, 19.8°, 20.3°, 24.9°, 25.1° and 25.6°; a diffraction angle 2θ of the Crystal H3 has characteristic peaks at 5.1°, 6.7°, 7.1°, 9.6°, 15.8°, 17.2°, 19.3°, 19.9°, 20.7° and 24.9°; and a diffraction angle 2θ of the Crystal H4 has characteristic peaks at 5.5°, 5.6°, 10.0°, 10.9°, 10.9°, 16.6°, 19.8°, 20.0°, 20.4° and 25.0°. The four novel sofosbuvir crystals respectively have good physicochemical properties, good stability and good solubility, and are easy to prepare and handle.

Description

Crystal form of flubuvir and preparation method thereof Technical field

The present invention relates to sofosbuvir (isopropyl(2S)-2-[[[(2R,3R,4R,5R)-5-(2,4-dioxapyrimidin-1-yl)-4-fluoro-) 3-hydroxy-4-methyl-tetrahydrofuran-2-yl]-methoxy-phenoxy-phosphoryl]amino]propionate, four novel crystals of the formula: C ₂₂ H ₂₉ FN ₃ O ₉ P) Type and its preparation method.

Background technique

Sofosbuvir, chemical name isopropyl (2S)-2-[[[(2R,3R,4R,5R)-5-(2,4-dioxapyrimidin-1-yl)-4 -Fluoro-3-hydroxy-4-methyl-tetrahydrofuran-2-yl]-methoxy-phenoxy-phosphoryl]amino]propionate, CAS No. 1190307-88-0, the chemical structural formula is as follows:

Soofibide is an NS5B polymerase inhibitor developed by Glead Sciences that blocks the specificity required for hepatitis C virus replication when used alone or in combination with other drugs. Protein, which is used in the treatment of hepatitis C. Soofibide was approved by the US Food and Drug Administration in December 2013 under the trade name Sovaldi. Soofibrevir is the first drug approved for total oral therapy for hepatitis C. It can eliminate the need for traditional injectable drug interferon (IFN) when used in the treatment of specific genotypes of chronic hepatitis C. With global sales exceeding US$10 billion, it has broad market prospects.

U.S. Patent No. 1,201,051,152 describes six crystal forms of sofosbuvir and teaches a process for the preparation of Form 6 therein. The six crystal forms described in this document are as follows:

The crystalline form Form 1 has powder X-ray diffraction characteristic peaks of 5.2°, 7.5°, 9.6°, 16.7°, 18.3°, and 22.2°. The patent US20110251152 does not teach a specific preparation method of the crystal form 1, but indicates that the crystal form 1 can be converted from the crystal forms 2 to 5. The crystal form 1 itself is highly hygroscopic, and is easily converted into a gel-like substance or converted into a crystal form 6 in an open container.

Form 2 is a solvate crystal of dichloromethane having powder X-ray diffraction characteristic peaks of 4.9, 6.9, 9.8, 19.8, 20.6, 24.7 and 26.1. Form 2 is crystallized from dichloromethane and converted to Form 1 upon drying.

The crystal form 3 is a chloroform solvate crystal having a powder X-ray diffraction characteristic peak of 6.9, 9.8, 19.7, 20.6 and 24.6. Form 3 is crystallized from chloroform and converted to Form 1 upon drying.

Form 4 has a powder X-ray diffraction characteristic peak of 5.0°, 6.8°, 19.9°, 20.6°, and 24.6°. The crystal form 4 was crystallized from acetonitrile, and the properties were unstable, and it was converted into the crystal form 1 when it was separated by filtration.

The crystalline form 5 has powder X-ray diffraction characteristic peaks of 5.2°, 6.6°, 7.1°, 15.7°, 19.1°, and 25.0°. The crystal form 5 is crystallized from anisole and has an unstable property, which is converted into a crystal form 1 when separated by filtration.

The crystalline form 6 has powder X-ray diffraction characteristic peaks of 6.1, 8.2, 10.4, 12.7, 17.2, 17.7, 18.0, 18.8, 19.4, 19.8, 20.1, and 20.8. The patent US20110251152 discloses the following two methods for preparing the crystalline form 6, which are obtained by conversion from the crystalline form 1:

1) The crystal form 1 powder was placed in a room temperature for several days to form a gel, and the gel was ground into a powder (the powder X-ray diffraction was the same as that of form 1). After the powder was placed in an open container for 6-10 weeks, the sample slowly became crystalline form 6.

2) The crystal form 1 is added to water of 5 to 50 mg/ml at normal temperature, and stirred at normal temperature or 50 ° C, and the crystal form 1 is converted into the crystal form 6 in a few hours.

The existing crystal forms of the six kinds of sofosbuvir have disadvantages such as low stability, low solubility and poor reproducibility of the production process, which directly affects the production and storage of the solfifil drug substance. The effect of its pharmaceutical preparations in the treatment of hepatitis C.

It is well known that the crystalline form of a drug can greatly affect the properties of the drug, such as chemical stability, melting point, solubility, dissolution rate, and the like. These characteristics can directly affect the production, storage and use of APIs and preparations. Therefore, controlling the crystal form of sofosbuvir is essential for obtaining a better formulation of sofosbuvir, and it is expected to develop new novels with higher stability, higher solubility and easier preparation of sofosbuvir. Crystal form.

Summary of the invention

It is an object of the present invention to provide new crystalline forms of sofosbuvir having superior physicochemical properties, as well as methods for preparing these crystalline forms.

The inventors of the present invention conducted intensive studies to solve the above problems, and as a result, four new crystalline forms H1, H2, H3, and H4 of sofosbuvir were surprisingly found. Among them, the crystal form H1 is a hydrate crystal containing 1:1 water molecules, the crystal form H2 is a methanol crystal containing 1:1 methanol molecules, and the crystal form H3 is an ethanolate crystal containing 1:1 ethanol molecules, crystal The type H4 is a crystal of a diphenyl ether compound containing a 1:1 diphenyl ether molecule. Further, the inventors have also found a method of selectively obtaining a crystal form H1, a crystal form H2, a crystal form H3, and a crystal form H4, respectively. These four new crystalline forms have the advantages of excellent physical and chemical properties, good stability, good solubility, simple preparation and operation, and have superiority in industrial production and medical applications.

The diffraction angle 2θ of the crystal form H1 disclosed in the present invention in the powder X-ray diffraction pattern using Cu-Kα ray is 4.9°, 6.6°, 7.1°, 8.2, 9.6°, 16.5°, 19.0°, 19.2°, 19.9°. There are characteristic peaks at 24.9° and the like.

After single crystal X-ray diffraction experiments and crystal structure calculations, it was confirmed that the crystal form H1 belongs to the monoclinic system, the space group P21, and the unit cell parameters:

α=90.00°, β=108.05(3)°, γ=90.00°; unit cell volume

The unit cell contains an asymmetric unit number Z=2, and the stoichiometric formula in one asymmetric unit is C ₂₂ H ₃₁ FN ₃ O ₁₀ P, the molecular weight is 547.47, and the crystal density d=1.251 g/cm ³ . Single crystal X-ray diffraction structure analysis confirmed that the crystal form H1 lattice contained 1:1 water molecules.

The crystal form H1 of the present invention has a small endothermic peak at 65 to 80 ° C and a large endothermic at 95 to 110 ° C in the range of 50-200 ° C in the DSC chart (differential scanning calorimetry). Peak (its peak temperature is 100.6 ° C).

The invention provides a preparation method of the crystal form H1, wherein the soffluzil and water are added to a mixed solvent of anisole and dibutyl ether or a mixed solvent of anisole and diisopropyl ether, and heated to 70 to 95 ° C. Then, the temperature was lowered to 0 to 35 ° C to carry out crystallization, and the crystallization time was 1 to 10 days, whereby a colorless needle-like crystal form H1 was obtained.

In the method for producing the crystalline form H1 of the present invention, the volume ratio of anisole to dibutyl ether or isopropyl ether is from 3:1 to 1:1, preferably 1:1; the concentration of sofosbuvir (g The ratio of the mixed solvent volume (mL) is 1:5 to 1:20, more preferably 1:10; the weight ratio of water to sofosbuvir is 1:100 to 1:20, more preferably 3:100; The heating temperature is preferably 80 to 85 ° C; preferably, the temperature is lowered to 20 to 25 ° C; and the crystallization time is preferably 3 days.

The diffraction angle 2θ of the crystal form H2 of the present invention in a powder X-ray diffraction pattern using Cu-Kα ray is 6.7°, 7.0°, 9.7°, 19.1°, 19.3°, 19.8°, 20.3°, 24.9°, 25.1°. There are characteristic peaks at 25.6° and the like.

After single crystal X-ray diffraction experiments and crystal structure calculations, it was confirmed that the crystal form H2 belongs to the monoclinic system, the space group P21, and the unit cell parameters: α=90.00°, β=108.86(3)°, γ=90.00°; unit cell volume

The unit cell contains an asymmetric unit number Z=2, and the stoichiometric formula in an asymmetric unit is C ₂₃ H ₃₃ FN ₃ O ₁₀ P, the molecular weight is 561.49, and the crystal density d=1.313 g/cm ³ . Single crystal X-ray diffraction structural analysis confirmed that the crystalline H2 lattice contained 1:1 methanol solvent molecules.

The crystal form H2 of the present invention has an endothermic peak at 84 to 100 ° C in the range of 50 to 200 ° C in the DSC chart, and the peak temperature is 89.1 ° C.

The invention provides a preparation method of the crystal form H2, wherein the Sophorabine and methanol are added to a mixed solvent of an isophthalic ether solvent, isophthalic ether and dibutyl ether, or m-xylylene ether and diisopropyl ether. In the mixed solvent, the mixture is heated to 70 to 95 ° C, and then cooled to 0 to 35 ° C to carry out crystallization, and the crystallization time is 1 to 10 days, thereby obtaining a colorless needle-like crystal form H2.

In the method for producing the crystalline form H2 of the present invention, the volume ratio of the m-xylylene ether to the dibutyl ether or isopropyl ether is from 3:1 to 1:1, more preferably 1:1; sofosbuvir The ratio of the weight (g) to the solvent volume (mL) is 1:5 to 1:20, preferably 1:8; the weight ratio of methanol to sofosbuvir is 1:20 to 3:20, more preferably 1:10. The heating temperature is preferably 80 to 85 ° C; preferably, the temperature is lowered to 20 to 25 ° C; and the crystallization time is preferably 3 days.

The diffraction angle 2θ of the crystal form H3 of the present invention in a powder X-ray diffraction pattern using Cu-Kα ray is 5.1°, 6.7°, 7.1°, 9.6°, 15.8°, 17.2°, 19.3°, 19.9°, 20.7°. There are characteristic peaks at 24.9° and the like.

After single crystal X-ray diffraction experiments and crystal structure calculations, it was confirmed that the crystal form H3 belongs to the monoclinic system, the space group P21, and the unit cell parameters:

α=90.00°, β=108.78(3)°, γ=90.00°; unit cell volume

The unit cell contains an asymmetric unit number Z=2, and the stoichiometric formula in one asymmetric unit is C ₂₄ H ₃₅ FN ₃ O ₁₀ P, the molecular weight is 575.52, and the crystal density d is 1.342 g/cm ³ . Single crystal X-ray diffraction structure analysis confirmed that the crystalline H3 lattice contains 1:1 ethanol solvent molecules

The crystal form H3 of the present invention has an endothermic peak at 70 to 88 ° C in the range of 50 to 200 ° C in the DSC chart, and the peak temperature is 74.4 ° C. The crystalline form H3 of the present invention is a sophorabate ethanol solvate containing one molecule of ethanol.

The invention provides a preparation method of the crystal form H3, wherein the Soofibide and ethanol are added to the solvent of dibenzyl ether, heated to 70-95 ° C, and then cooled to 0-35 ° C for crystallization, and the crystallization time is 1 ～. After 10 days, a colorless flake-form crystal form H3 was obtained.

In the method for preparing the crystalline form H3 of the present invention, the ratio of the weight (g) of sofosbuvir to the volume of the solvent (mL) is from 1:10 to 1:20, preferably 1:10; ethanol and soffloxacin The weight ratio of wei is 1:50 to 1:10, preferably 1:20; the heating temperature is preferably 85 to 90 ° C; preferably, the temperature is lowered to 20 to 25 ° C; and the crystallization time is preferably 3 days.

The diffraction angle 2θ of the crystal form H4 of the present invention in a powder X-ray diffraction pattern using Cu-Kα ray is 5.5°, 5.6°, 10.0°, 10.9°, 10.9°, 16.6°, 19.8°, 20.0°, 20.4°. There are characteristic peaks at 25.0° and the like.

After single crystal X-ray experiments and crystal structure calculations, it was confirmed that the crystal form H4 belongs to the monoclinic system, the space group P21, and the unit cell parameters:

α=90.00°, β=116.64(3)°, γ=90.00°; unit cell volume

The unit cell contains an asymmetric unit number Z=2, and the stoichiometric formula in one asymmetric unit is C ₃₄ H ₃₉ FN ₃ O ₁₀ P, the molecular weight is 699.65, and the crystal density d=1.313 g/cm ³ . Single crystal X-ray diffraction structural analysis confirmed that the crystalline H4 crystal lattice contained 1:1 diphenyl ether solvent molecules.

The crystal form H4 of the present invention has an endothermic peak at 64 to 76 ° C in the range of 50 to 200 ° C in the DSC chart, and the peak temperature is 68.1 ° C.

The invention provides a preparation method of the crystalline form H4, wherein the Soofibide is added to a solvent of diphenyl ether, heated to 70-95 ° C, and then cooled to 0-35 ° C for crystallization, and the crystallization time is 1 to 10 days. Thus, a colorless needle-like crystal form H4 was obtained.

In the method for producing the crystalline form H4 of the present invention, the ratio of the weight (g) of sofosbuvir to the volume of the solvent (mL) is from 1:3 to 1:20, preferably 1:10; the heating temperature is preferably 85. ～90° C.; preferably, the temperature is lowered to 20 to 25° C.; the crystallization time is preferably 3 days.

DRAWINGS

1 is an X-ray single crystal molecular structure ellipsoid of Soofibvir crystal form H1 according to Example 1 of the present invention;

2 is an X-ray powder diffraction pattern of Soofibvir crystal form H1 according to Example 1 of the present invention;

Figure 3 is a DSC-TGA chart of the Soofibvir crystal form H1 according to Example 1 of the present invention;

4 is an X-ray single crystal molecular structure ellipsoid of Soofibvir crystal form H2 according to Example 2 of the present invention;

Figure 5 is an X-ray powder diffraction pattern of sofosbuvir crystal form H2 according to Example 2 of the present invention;

Figure 6 is a DSC-TGA chart of Soofibvir crystal form H2 according to Example 2 of the present invention;

7 is an X-ray single crystal molecular structure ellipsoid of Soofibvir crystal form H3 according to Example 3 of the present invention;

Figure 8 is an X-ray powder diffraction pattern of sofosbuvir crystal form H3 according to Example 3 of the present invention;

Figure 9 is a DSC-TGA chart of Soofibvir crystal form H3 according to Example 3 of the present invention;

Figure 10 is an X-ray single crystal molecular structure ellipsoid of Soofibvir crystal form H4 according to Example 4 of the present invention;

Figure 11 is an X-ray powder diffraction pattern of sofosbuvir crystal form H4 according to Example 4 of the present invention;

Figure 12 is a DSC-TGA diagram of sofosbuvir crystal form H4 according to Example 4 of the present invention.

detailed description

The invention is described in more detail below with reference to the examples, but the invention is not intended to be limited thereby. The experimental methods in the following examples, which do not specify the specific conditions, were carried out in accordance with conventional methods and conditions.

The room temperature referred to in the present invention means 10 ° C to 30 ° C.

In the preparation method of the four crystal forms disclosed in the present invention, sofosbuvir is prepared using the method reported in the prior literature (J. Med. Chem. 2010, 53: 7202-7228); Both solvents and reagents are commercially available as chemically pure or analytically pure products.

The four novel crystal forms disclosed in the present invention were all tested and characterized by single crystal X-ray diffraction structure analysis. The single crystal X-ray diffraction structure analysis can directly obtain the unit cell parameters, space group, number of molecules in the unit cell, solvent in the unit cell and the three-dimensional structure information of the molecule. It is the most direct, accurate and effective crystal in the current crystal research method. Type analysis method.

The crystal structure measuring instrument used in the examples of the present invention was Enraf Noius & Enraf Noius's CAD4/PC single crystal X-ray diffractometer, and the test temperature was 293 (2) K. Monochromating MoKα ray with graphite

Structural analysis and correction were performed using SHELXS-97 (Sheldrick, 1990) and SHELXL-97 (Sheldrick, 1997). A single crystal molecular structure ellipsoid was obtained using the program ORTEP (Fig. 1, Fig. 4, Fig. 7, Fig. 10).

The crystal cell parameters obtained by crystal structure determination have the following meanings:

a, b, c are the side lengths of the unit cell;

α, β, γ are the angles of the crystal plane of the unit cell;

V is the unit cell volume;

Z is the number of asymmetric units contained in the unit cell.

The X-ray powder diffraction apparatus used in the examples of the present invention is a X'pert PRO type X-ray powder diffractometer manufactured by PANalytical Co., Ltd. The Cu-Kα ray was used, and the test power was 40 kV×250 mA, the scanning speed was 5°/min, and the scanning range was 4 to 80° (2θ).

In the X-ray powder diffraction pattern (Fig. 2, Fig. 5, Fig. 8, and Fig. 11) obtained in the examples of the present invention, the horizontal axis represents the diffraction peak 2θ position (unit: degree); and the vertical axis represents the diffraction peak intensity.

The differential scanning calorimetry-thermogravimetric (DSC-TGA) analytical instrument used in the examples of the present invention is a TGA/DSC type 1 synchronous thermal analyzer from Mettler Toledo. The measurement range is 25-350 ° C, the heating rate is 10 ° C / min, and nitrogen protection is used.

In the differential scanning calorimetry-thermal weight (DSC-TGA) diagram (Fig. 3, Fig. 6, Fig. 9, Fig. 12) obtained in the embodiment of the present invention, the thermogravimetric (TGA) diagram and the calorimetric (DSC) diagram are upper and lower. Side by side. In the thermogravimetric diagram, the horizontal axis is temperature (°C), and the vertical axis is mass (mg). In the calorimetric diagram, the horizontal axis is temperature (unit: °C), and the vertical axis is power (unit: milliwatt).

Example 1: Preparation of a Form H1 single crystal.

0.5 g of sofosbuvir and 15 mg of water were added to a mixed solvent of 2.5 ml of anisole and 2.5 ml of dibutyl ether, and the mixture was heated to 80 to 85 ° C and stirred to completely dissolve the solid, followed by 1 to 3 ° C / min. The rate was slowly cooled to room temperature and allowed to stand at room temperature for 72 hours, and the precipitated colorless needle-shaped crystal form H1 single crystal was taken out by filtration.

Alternatively, the mixed solvent in the above production method may be replaced with a mixed solvent of 2.5 ml of m-xylylene ether and 2.5 ml of isopropyl ether.

Measurement of the X-ray structure of the crystal form H1 single crystal in Example 1.

The Sooflovon crystal form H1 single crystal having a size of 0.30 mm×0.20 mm×0.10 mm in the crystal obtained in Example 1 was selected and placed on a single crystal X-ray diffractometer, and collected in the range of 1.20° ≤ 2θ ≤ 25.42°. 5532 diffraction data, of which 5,310 were independent diffraction points. The collected data was corrected by Lp factor and empirical absorption. Using the direct method, and after multiple rounds of difference Fourier synthesis, find all hydrogen atoms. All non-hydrogen atom coordinates and their equivalent anisotropic displacement parameters are all refined using the full matrix least squares method.

After single crystal X-ray diffraction test and crystal structure calculation, it was confirmed that the crystalline form H1 of soflubluff belongs to monoclinic system, space group P21, unit cell parameters:

α=90.00°, β=108.05(3)°, γ=90.00°;

Z = 2, the stoichiometric formula in an asymmetric unit is C ₂₂ H ₃₁ FN ₃ O ₁₀ P, the molecular weight is 547.47, and the crystal density d is 1.251 g/cm ³ . The single crystal molecular structure ellipsoid of the Sophobvir crystal form H1 was determined as shown in Fig. 1, and the non-hydrogen atom coordinate data is shown in Table 1.

Table 1 Non-hydrogen atom coordinate parameters of crystal form H1

Powder X-ray diffraction analysis of the crystalline H1 single crystal of Example 1 :

The crystal form H1 single crystal obtained in Example 1 was ground into a powder, and subjected to powder X-ray diffraction analysis, and the results are shown in Fig. 2 . The X-ray diffraction data of the crystal form H1 corresponding to Fig. 2 is shown in Table 2.

Table 2 Powder X-ray Diffraction Analysis of Form H1

As a result of X-ray diffraction, the position of the characteristic diffraction peak 2θ of the crystal form H1 was 4.9°, 6.6°, 7.1°, 8.2°, 9.6°, 16.5°, 19.0°, 19.2°, 19.9°, 24.9°, and the like.

The powder sample was subjected to differential scanning calorimetry-thermogravimetric analysis, and the results are shown in Fig. 3.

As can be seen from Fig. 3, the crystal form H1 has a small endothermic peak at 65 to 80 ° C and a large endothermic peak at 95 to 110 ° C in the thermogravimetric diagram in the range of 50 to 200 °C. The difference thermogram shows that the weight loss rate is 2.3% in the range of room temperature to 150 ° C, and the error range is consistent with the structure obtained by single crystal X-ray diffraction analysis.

Example 2: Preparation of crystalline H2 single crystal.

Add 0.5 g of sofosbuvir and 0.05 g of methanol to 5.0 ml of m-xylylene ether solvent, heat to 85-90 ° C to completely dissolve the solid, then slowly cool to room temperature at room temperature of 1-3 ° C / min and at room temperature After standing for 24 to 72 hours, the colorless needle-like crystal form H2 was taken out by filtration.

Alternatively, the 5.0 ml of the m-xylylene ether solvent in the above preparation method may be replaced with a mixed solvent of 5.0 ml of m-xylylene ether and 5.0 ml of isopropyl ether.

Determination of the crystal structure of the crystalline form H2 in Example 2.

The Sooflovon crystal form H2 single crystal having a size of 0.20 mm×0.10 mm×0.10 mm in the crystal obtained in Example 2 was selected and placed on a single crystal X-ray diffractometer, and collected in the range of 1.21°≤2θ≤25.40°. 5446 diffraction data, of which 5216 were independent diffraction points. The collected data was corrected by Lp factor and empirical absorption. Using the direct method, and after multiple rounds of difference Fourier synthesis, find all hydrogen atoms. All non-hydrogen atom coordinates and their equivalent anisotropic displacement parameters are all refined using the full matrix least squares method.

After single crystal X-ray diffraction test and crystal structure calculation, it was confirmed that the crystalline form H2 of soflubluff belongs to monoclinic system, space group P21, unit cell parameters:

α=90.00°, β=108.86(3)°, γ=90.00°;

Z = 2, the stoichiometric formula in an asymmetric unit is C ₂₃ H ₃₃ FN ₃ O ₁₀ P, the molecular weight is 561.49, and the crystal density d = 1.313 g/cm ³ .

The single crystal molecular structure ellipsoid obtained by measuring the crystalline form H2 of sofosbuvir is shown in Fig. 4, and the non-hydrogen atom coordinate data is shown in Table 3.

Table 3 Non-hydrogen atom coordinate parameters of crystal form H2

Atomic number (eg

X coordinate

Y coordinate

Z coordinate

U eq (temperature equivalence factor

Figure 4)				child)
					C1	0.8703 (5)	‐0.0224(6)	0.8922(4)	0.0478(6)
C2	0.9650 (4)	‐0.1377(4)	0.8947 (3)	0.0409(5)
					C3	1.0421(5)	‐0.3253(6)	0.8132(3)	0.0394 (6)
C4	1.1213(4)	‐0.1659(5)	0.8652 (3)	0.0384 (6)
					C5	1.0560(5)	0.0049 (6)	0.8909 (3)	0.0377(6)
C6	1.1991(6)	‐0.0781(6)	0.8323(4)	0.062 (2)
					C7	0.9731 (5)	‐0.1294(6)	0.6856(3)	0.0462(6)
C8	0.9529 (5)	‐0.1148(6)	0.6095 (4)	0.0483 (6)
					C9	0.9750 (5)	‐0.2902(6)	0.5645(4)	0.0477(6)
C10	1.0505 (5)	‐0.4857(6)	0.6897(4)	0.0466(6)
					C11	0.6957 (6)	0.4796(6)	0.6733 (4)	0.059(2)
C12	0.6753 (6)	0.7220 (6)	0.6432(5)	0.094(3)
					C13	0.7411(6)	0.3577 (6)	0.6199(4)	0.0548(6)
C14	0.6971 (6)	0.183 (2)	0.4917(5)	0.109 (4)
					C15	0.7361(6)	0.334(3)	0.4502 (6)	0.158(5)
C16	0.5987(5)	0.062 (3)	0.4420 (5)	0.163 (4)
					C17	0.5580(5)	0.2608(6)	0.7866(4)	0.0498(6)
C18	0.5414(5)	0.4768 (5)	0.8131(5)	0.077 (3)
					C19	0.4402 (5)	0.5304(5)	0.8009 (4)	0.093(3)
C20	0.3614(5)	0.3978(5)	0.7702(4)	0.086(3)
					C21	0.3806(5)	0.190 (2)	0.7483 (6)	0.101(3)
C22	0.4795(6)	0.1290(6)	0.7552(5)	0.075 (3)
					C23	0.6038(5)	0.739 (3)	0.9962(5)	0.192
F	1.1711(3)	‐0.2860(6)	0.93449 (7)	0.0525(7)
					N1	1.0225(4)	‐0.3090(6)	0.7291(3)	0.0390(5)
N2	1.0278(4)	‐0.4584(4)	0.6110(3)	0.0472(4)
					N3	0.7579(4)	0.4846(5)	0.7541(3)	0.0492(5)
O1	0.8375(3)	0.1267(5)	0.8238(2)	0.0490(5)
					O2	0.9474(3)	‐0.2805(4)	0.8269(2)	0.0437(5)
O3	1.1057 (3)	0.1207(4)	0.9599 (2)	0.0481(4)
					O4	0.9534 (4)	‐0.2994(4)	0.4923(2)	0.0673 (5)
O5	1.0947 (4)	‐0.6461(5)	0.7265(3)	0.0584(5)
					O6	0.6702(4)	0.3082(6)	0.5524(3)	0.0950(6)
O7	0.8293 (4)	0.3076(5)	0.6382 (3)	0.0814(6)
					O8	0.7812(4)	0.4183 (5)	0.9016(2)	0.0536(5)
O9	0.6550 (3)	0.1850(5)	0.7932 (3)	0.0538 (5)
					O10	0.5485(5)	0.966 (2)	0.9894(4)	0.248

P

0.76017 (7)

0.3141(3)

0.82483 (5)

0.0433(4)

Powder X-ray diffraction analysis of the crystalline H2 single crystal of Example 2:

The crystal form H2 single crystal obtained in Example 2 was ground into a powder, and subjected to powder X-ray diffraction analysis, and the results are shown in Fig. 5 . The X-ray diffraction data of this crystal form H2 is shown in Table 4.

Table 4 X-ray diffraction analysis of crystalline H2 powder

As a result of X-ray diffraction, the position of the characteristic diffraction peak 2θ of the crystal form H2 was 6.7°, 7.0°, 9.7°, 19.1°, 19.3°, 19.8°, 20.3°, 24.9°, 25.1°, 25.6°, and the like.

The powder sample was subjected to differential scanning calorimetry-thermogravimetric analysis, and the results are shown in Fig. 6.

As can be seen from Fig. 6, the crystal form H2 has an endothermic peak at 84-100 ° C in the thermogravimetric diagram in the range of 50-200 °C. The difference thermogram shows that the weight loss rate is 5.5% from room temperature to 180 °C, and the error range is consistent with the structure obtained by single crystal X-ray diffraction analysis.

Example 3: Preparation of crystalline H3 single crystal

Mix 0.5 g of sofosbuvir with 25 mg of ethanol and 5 ml of dibenzyl ether, heat the mixture to 85-90 ° C and stir to dissolve the solid completely, then slowly cool to room temperature at 1-3 ° C / min and After standing at room temperature for 72 hours, the precipitated colorless needle-shaped crystal form H3 single crystal was taken out by filtration.

Determination of crystal structure of crystal form H3 of Example 3

The Sooflovon crystal form H3 single crystal having a size of 0.30 mm×0.20 mm×0.10 mm in the crystal obtained in Example 3 was selected and placed on a single crystal X-ray diffractometer, and collected in the range of 1.20° ≤ 2θ ≤ 25.38°. 5460 diffraction data, of which 5230 were independent diffraction points. The collected data was corrected by Lp factor and empirical absorption. Using the direct method, and after multiple rounds of difference Fourier synthesis, find all hydrogen atoms. All non-hydrogen atom coordinates and their equivalent anisotropic displacement parameters are all refined using the full matrix least squares method.

After single crystal X-ray diffraction test and crystal structure calculation, it was confirmed that the crystalline form H3 of the sofosbuvir belongs to the monoclinic system, the space group P21, and the unit cell parameters:

α=90.00°, β=108.78(3)°, γ=90.00°;

Z = 2, the stoichiometric formula in an asymmetric unit is C ₂₄ H ₃₅ FN ₃ O ₁₀ P, the molecular weight is 575.52, and the crystal density d is 1.342 g/cm ³ .

The single crystal molecular structure ellipsoid obtained by measuring the crystal form H3 of sofosbuvir is shown in Fig. 7, and the non-hydrogen atom coordinate data is shown in Table 5.

Table 5 Non-hydrogen atom coordinate parameters of crystal form H3

Powder X-ray diffraction analysis of the crystalline H3 single crystal of Example 3:

The crystal form H3 single crystal obtained in Example 3 was ground into a powder, and subjected to powder X-ray diffraction analysis, and the results are shown in Fig. 8 . Corresponding to Fig. 8, the X-ray diffraction data of the crystal form H3 is shown in Table 6.

Table 6 Powder X-ray Diffraction Analysis of Form H3

From the results of X-ray diffraction, the positions of the diffraction peaks 2θ of the crystal form H3 were 5.1, 6.7, 7.1, 9.6, 15.8, 17.2, 19.3, 19.9, 20.7, and 24.9.

The powder sample was subjected to differential scanning calorimetry-thermogravimetric analysis, and the results are shown in Fig. 9.

According to the thermogravimetric diagram of the crystal form H3 in Fig. 9, the crystal form H3 has an endothermic peak at 70 to 88 ° C in the range of 50 to 200 °C. The difference thermogram shows that the weight loss rate is 5.3% from room temperature to 180 °C, and the error range is consistent with the structure obtained by single crystal X-ray diffraction analysis.

Example 4: Preparation of crystalline H4 single crystal

Mix 0.5 g of sofobuvir with 5 ml of diphenyl ether solvent, heat to 85-90 ° C to form an oily mixture, then slowly cool to room temperature at 1-3 ° C / min and let stand at room temperature for 72 hours. The colorless needle-shaped crystal form H4 single crystal was taken out by filtration.

Determination of Crystal Structure of Form H4 of Example 4

The Sooflovon crystal form H4 single crystal having a size of 0.20 mm×0.10 mm×0.10 mm in the crystal obtained in Example 4 was selected and placed on a single crystal X-ray diffractometer, and collected in the range of 1.28° ≤ 2θ ≤ 25.39°. There are 6737 diffraction data, of which 6514 are independent diffraction points. The collected data was corrected by Lp factor and empirical absorption. Using the direct method, and after multiple rounds of difference Fourier synthesis, find all hydrogen atoms. All non-hydrogen atom coordinates and their equivalent anisotropic displacement parameters are all refined using the full matrix least squares method.

After single crystal X-ray diffraction test and crystal structure calculation, it was confirmed that the Sooflovon crystal form H4 belongs to the monoclinic system, the space group P21, and the unit cell parameters:

α=90.00°, β=116.64(3)°, γ=90.00°;

Z = 2, the stoichiometric formula in an asymmetric unit is C ₃₄ H ₃₉ FN ₃ O ₁₀ P, the molecular weight is 699.65, and the crystal density d = 1.313 g/cm ³ .

The single crystal molecular structure ellipsoid obtained by measuring the crystalline form H4 of sofosbuvir is shown in Fig. 10, and the non-hydrogen atom coordinate data is shown in Table 7.

Table 7 Non-hydrogen atom coordinate parameters of crystal form H4

Powder X-ray diffraction analysis of the crystalline H4 single crystal of Example 4:

The crystal form H4 single crystal obtained in Example 4 was ground into a powder, and subjected to powder X-ray diffraction analysis, and the results are shown in Fig. 11 . Corresponding to Fig. 11, the X-ray diffraction data of the crystal form H4 is shown in Table 8.

Table 8 X-ray diffraction analysis of crystalline H4 powder

From the results of X-ray diffraction, the positions of the diffraction peaks 2θ of the crystal form H4 were 5.5°, 5.6°, 10.0°, 10.9°, 10.9°, 16.6°, 19.8°, 20.0°, 20.4°, 25.0°, and the like.

The powder sample was subjected to differential scanning calorimetry-thermogravimetric analysis, and the results are shown in Fig. 12.

According to the thermogravimetric diagram of the crystal form H4 in Fig. 12, the crystal form H4 has an endothermic peak at 64 to 76 ° C in the range of 50 to 200 °C. The difference thermogram shows that the weight loss rate is 17.4% in the range of room temperature to 200 ° C, and the error range is consistent with the structure obtained by single crystal X-ray diffraction analysis.

Test example 1

The high-humidity stability measurement of the crystal form H1, the crystal form H2, the crystal form H3, and the form H4 obtained according to Example 1-4 was carried out, and was compared with the crystal forms of Form 1 and Form 6 of the known Soofibide.

Samples of various crystal forms of Soofibide are evenly spread in an open container, the sample thickness is ≤ 5 mm, placed in a constant temperature and humidity incubator at a temperature of 25 ± 2 ° C and a relative humidity of 60 ± 5%. Observations were made after a fixed time interval t and compared to the starting crystal form. The results are shown in Table 9.

Table 9 Stability of sofbuvir crystal form under humidity conditions (25 ° C / 60% relative humidity)

Crystal form	t=2 hours	t=4 hours	t=24 hours	t=72 hours
					Crystal form H1	No moisture solution	No moisture solution	No moisture solution	Partial deliquescence
Crystal form H2	No moisture solution	No moisture solution	No moisture solution	No moisture solution
					Crystal form H3	No moisture solution	No moisture solution	No moisture solution	No moisture solution
Form H4	No moisture solution	No moisture solution	No moisture solution	No moisture solution
Crystal form 1	No moisture solution	No moisture solution	Partial deliquescence	deliquescence
					Crystal form 6	No moisture solution	No moisture solution	No moisture solution	Partial deliquescence

The temperature and humidity conditions were changed, and the same sample was placed in a constant temperature and humidity incubator at a temperature of 40 ± 2 ° C and a relative humidity of 75 ± 5%, and observed at a fixed time t and compared with the starting sample crystal form. The results are shown in Table 10.

Table 10 Stability of sofosbuvir crystal form under humidity conditions (40 ° C / 75% relative humidity)

Crystal form	t=2 hours	t=4 hours	t=24 hours	t=72 hours
					Crystal form H1	No moisture solution	No moisture solution	Partial deliquescence	deliquescence
Crystal form H2	No moisture solution	No moisture solution	No moisture solution	No moisture solution
					Crystal form H3	No moisture solution	No moisture solution	No moisture solution	No moisture solution
Form H4	No moisture solution	No moisture solution	No moisture solution	No moisture solution
Crystal form 1	Partial deliquescence	deliquescence	deliquescence	deliquescence
					Crystal form 6	No moisture solution	No moisture solution	Partial deliquescence	deliquescence

The test results show that the crystalline form H2, the crystalline form H3 and the crystalline form H4 exhibit less hygroscopicity under different temperature and humidity conditions, have good stability, and effectively solve the existing crystal form of sofofubuvir. The problem of deliquescence.

Test example 2

The crystal form H1, the crystal form H2, the crystal form H3, and the form H4 obtained according to Examples 1-4 were subjected to water solubility measurement, and were compared with the crystal forms of Form 1 and Form 6 of the known Soofibide. The test method was carried out in accordance with the method specified in the Chinese Standards of the Chinese Pharmacopoeia 2010. The specific method is as follows: accurately weigh the appropriate amount of different crystal form of sofosbuvir, slowly add a certain amount of water as a solvent, shake vigorously for 30 seconds every 5 minutes, observe the dissolution within 30 minutes. The test results are shown in Table 11.

Table 11 Solubility of different crystal forms of sofosbuvir in water

The test results show that the solubility of crystal form H1, crystal form H2 and form H3 in water is higher than that of the known crystal form of sofosbuvir, which effectively solves the problem of poor solubility of the existing sofosbuvir crystal form. .

Claims (16)

1. A crystal form H1 of sofosbuvir characterized in that, using Cu-Kα ray, the diffraction angle 2θ in the powder X-ray diffraction pattern of the crystal form H1 is 4.9°, 6.6°, 7.1°, 8.2°, 9.6° Characteristic peaks at 16.5°, 19.0°, 19.2°, 19.9°, and 24.9°.
2. The crystal form H1 according to claim 1, wherein the differential scanning calorimetry chart of the crystal form H1 has an endothermic peak at 65 to 80 ° C and 95 to 110 ° C.
3. A crystalline form H2 of sofosbuvir characterized in that, using Cu-Kα ray, the diffraction angle 2θ in the powder X-ray diffraction pattern of the crystalline form H2 is at 6.7°, 7.0°, 9.7°, 19.1°, 19.3°. Characteristic peaks at 19.8°, 20.3°, 24.9°, 25.1°, and 25.6°.
4. The crystal form H2 according to claim 3, wherein the differential scanning calorimetry chart of the crystal form H2 has an endothermic peak at 84 to 100 °C.
5. A crystalline form H3 of sofosbuvir characterized in that, using Cu-Kα ray, the diffraction angle 2θ in the powder X-ray diffraction pattern of the crystalline form H3 is at 5.1°, 6.7°, 7.1°, 9.6°, 15.8°. Characteristic peaks at 17.2°, 19.3°, 19.9°, 20.7°, and 24.9°.
6. The crystal form H3 according to claim 5, wherein the differential scanning calorimetry chart of the crystal form H3 has an endothermic peak at 70 to 88 °C.
7. A crystalline form H4 of sofosbuvir characterized by using Cu-Kα ray, the diffraction angle 2θ of the powder X-ray diffraction pattern of the crystalline form H4 is 5.5°, 5.6°, 10.0°, 10.9°, 10.9° Characteristic peaks at 16.6°, 19.8°, 20.0°, 20.4°, and 25.0°.
8. The crystal form H4 according to claim 7, wherein the differential scanning calorimetry chart of the crystal form H4 has an endothermic peak at 64 to 76 °C.
9. A method for producing a crystalline form H1 of sofosbuvir according to claim 1 or 2, characterized in that sofobuvir and water are added to a mixed solvent of anisole and dibutyl ether or benzate In a mixed solvent of ether and isopropyl ether, the mixture is heated to 70 to 95 ° C, and then cooled to 0 to 35 ° C to carry out crystallization, and the crystallization time is 1 to 10 days, thereby obtaining a crystal form H1.
10. The preparation method according to claim 9, wherein the volume ratio of anisole to dibutyl ether or isopropyl ether is from 3:1 to 1:1; the weight of the sofosbuvir in grams and the The ratio of the mixed solvent to the volume in milliliters is 1:5 to 1:20; the weight ratio of water to sofosbuvir is 1:100 to 1:20.
11. Process for the preparation of crystalline form H2 of sofosbuvir according to claim 3 or 4, characterized in that the sofosbuvir and methanol are added to the m-xylylene ether solvent or m-xylylene ether a mixed solvent of dibutyl ether or a mixed solvent of m-xylylene ether and isopropyl ether, heated to 70-95 ° C, and then cooled to 0-35 ° C for crystallization, crystallization time of 1 to 10 days Thereby, the crystal form H2 is obtained.
12. The preparation method according to claim 11, wherein the volume ratio of m-xylylene ether to dibutyl ether or diisopropyl ether is from 3:1 to 1:1; and the weight of the sofosbuvir is in terms of grams. The ratio of the solvent to the volume in milliliters is 1:5 to 1:20; the weight ratio of methanol to sofosbuvir is 1:20 to 3:20.
13. A method for producing the crystalline form H3 of the sofosbuvir according to claim 5 or 6, wherein the sofosbuvir and ethanol are added to the solvent of dibenzyl ether and heated to 70 to 95 ° C. Then, the temperature was lowered to 0 to 35 ° C to carry out crystallization, and the crystallization time was 1 to 10 days, thereby obtaining the crystal form H3.
14. The preparation method according to claim 13, wherein the ratio of the weight of the sofosbuvir to the volume of the solvent in the milliliter is 1:10 to 1:20; the weight ratio of the ethanol to the sofosbuvir is 1:50~1:10.
15. A process for the preparation of the crystalline form H4 of the sofosbuvir according to claim 7 or 8, characterized in that the sofosbuvir is added to a solvent of diphenyl ether, heated to 70 to 95 ° C, and then cooled. The crystallization was carried out at 0 to 35 ° C, and the crystallization time was 1 to 10 days, whereby the crystal form H4 was obtained.
16. The method according to claim 15, wherein the ratio of the weight of the sofosbuvir to the volume of the solvent in terms of milliliters is from 1:3 to 1:20.

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