Preparation and determination method of furazolidone single crystal with double bond cis-trans configuration

Abstract

The present application relates to the technical field of medicine preparation, more particularly, to a preparation and determination method of furazolidone single crystal with double bond cis-trans configuration. Furazolidone is added into a solvent, and is fully dissolved by vortexing, and is filtered to obtain a furazolidone saturated solution; the saturated solution is taken into a sample vial, and is sealed with a sealing film, a plurality of small holes are punched on the sealing film, and is left to stand, so that acetonitrile is slowly volatilized, and a crystal is grown to obtain a furazolidone single crystal with double bond cis-trans configuration. The solid state configuration of medicinal furazolidone is first confirmed: the furazolidone single crystal is successfully prepared by a solvent evaporation method, and high-precision single crystal X-ray diffraction technology (MoKα radiation, HyPix-Arc 150 detector) is combined with SHELXT program analysis, and it is first confirmed from the crystallographic level that the medicinal furazolidone is in a trans configuration in a solid state, which provides direct and irrefutable experimental evidence for the stereoscopic structure identification of the medicine.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical preparation technology, and more specifically, to a method for preparing and determining a single crystal of furazolidone with a double bond cis-trans configuration. Background Technology

[0002] Furazolidone, chemically known as 3-[[(5-nitro-2-furanyl)methylene]amino]-2-oxazolidinone, molecular formula: C8H7N3O 5， Molecular weight: 225.16, Structural formula: Furazolidone is a broad-spectrum antibacterial drug belonging to the nitrofuran class. It exhibits good antibacterial activity against a variety of Gram-positive and Gram-negative bacteria, particularly showing a high eradication rate against Helicobacter pylori infection. It remains an important drug in the treatment of clinically difficult-to-eradicate Helicobacter pylori infections. Furazolidone-containing compound preparations have been approved for marketing in my country since the 1970s, primarily used for various gastrointestinal infectious diseases caused by bacteria and protozoa. It has a long history of clinical application and a wide coverage.

[0003] However, furazolidone contains a C=N double bond (connecting the 5-nitrofuran ring and the 2-oxazolidinone ring), theoretically resulting in two geometric isomers: cis (Z) and trans (E). Drug molecules with different configurations often exhibit significant differences in biological activity, metabolic pathways, toxicity, and stability. For furazolidone, the exact configuration in which its active pharmaceutical ingredient is stably present, and whether different configurations interconvert under specific conditions (such as light, temperature, and solvents), has long lacked clear crystallographic data.

[0004] Currently, domestic and international drug standards (such as the Chinese Pharmacopoeia and the United States Pharmacopeia) mainly focus on content determination, related substance testing, and dissolution rate for the quality control of furazolidone raw materials and their preparations. There are no specific regulations for the identification or content limits of furazolidone geometric isomers. This is mainly due to the following unresolved technical bottlenecks: 1. Unclear solid-state configuration: For a long time, there have been no systematic reports on the single-crystal cultivation and structural analysis of furazolidone, and direct evidence for its precise geometric configuration in the solid state is lacking, resulting in a lack of data on crystal form research and stereochemistry. 2. Difficulty in assigning the structure of degradation impurities: In light stability studies, furazolidone easily produces a degradation product with the same molecular weight, but the chemical structure of this impurity has long been unconfirmed, making it impossible to determine whether it belongs to cis-trans isomers or to determine the relationship between its formation pathway and configuration transformation. 3. Lack of targeted analytical methods: Due to the unclear configuration and lack of isomer reference standards, existing analytical methods are unable to effectively distinguish furazolidone from its cis isomers, hindering the improvement of drug quality standards and the optimization of formulation processes.

[0005] In summary, there is an urgent need for a technical solution that can confirm the solid configuration of pharmaceutical furazolidone and elucidate its photoisomerization rules, in order to fill the gap in crystallographic data, clarify the structural attribution of degradation impurities, and provide a scientific basis for establishing a dedicated quality control method for isomers, thereby ensuring the clinical safety and efficacy consistency of furazolidone-containing preparations. Summary of the Invention

[0006] The present invention aims to at least solve one of the technical problems existing in the prior art. To this end, one aspect of the present invention is to provide a method for preparing a single crystal of furazolidone with a double bond cis-trans configuration, the specific steps of which are as follows: S1. Add furazolidone to a solvent, vortex to dissolve completely, filter, and obtain a saturated furazolidone solution; S2. Take a saturated solution of furazolidone and place it in a sample vial. Seal the vial with sealing film, make several small holes in the sealing film, let it stand, and allow the acetonitrile to evaporate slowly. This will grow crystals and obtain single crystals of furazolidone with double bond cis-trans configuration.

[0007] Preferably, the mass fraction ratio of furazolidone to solvent in S1 is 0.3-0.5 mg / mL.

[0008] Preferably, the solvent in S1 is acetonitrile.

[0009] Preferably, the vortex in S1 is a magnetic stirrer, and the vortex time is 1-1.5 hours.

[0010] Preferably, the acetonitrile in S2 slowly evaporates over a period of 3-5 days.

[0011] Another objective of this invention is to provide a method for determining the cis-trans configuration of furazolidone single crystals, the specific steps of which are: S1. Single crystal diffraction data collection: A single crystal sample of furazolidone with double bond cis-trans configuration was taken and data was collected at 100K on a 'ROD, Synergy Custom DW system' single crystal diffractometer, MoKα radiation; S2. Single Crystal Structure Analysis: Analysis of the color, size, space group, cell parameters, cell volume, number of molecules within the cell, and number of molecules contained in an independent region of the unit lattice for the double bond cis-trans configuration of furazolidone single crystal.

[0012] Preferably, in S1, the distance between the double-bond cis-trans configuration furazolidone single crystal and the 'HyPix-Arc 150' detector is 60 mm, the instrument operating voltage is 45 kV, the current is 66 mA, the maximum 2θ angle is 60.634°, the scanning step size is 0.5°, the scanning time is 0.2 s / °, each frame is scanned once, a total of 536 images are captured, the total number of diffraction points is 8683, and the number of independent diffraction points is 1344 [R] int =0.0252], the data integrity is 99.8% when the angle θ reaches 25.242°.

[0013] Preferably, the single crystal of furazolidone with double bond cis-trans configuration in S2 is orange-yellow in shape, with a crystal size of 0.24×0.2×0.12mm, belonging to the orthorhombic crystal system, space group Pnma, and cell parameters: a=22.5308(8)Å, b=6.0608(3)Å, c=6.5249(3)Å, α=90°, β=90°, γ=90°, and cell volume V=891.01(7)Å. 3 The number of molecules in the unit cell is Z=4, and each independent region of the unit lattice contains 0.5 molecules.

[0014] Preferably, in S2, the space group Pnma is determined using the software Olex2 on a computer by calling the program SHELXT to analyze the crystal structure. All non-hydrogen atom positions are obtained from the E diagram. The least squares method (program SHELXL) is used to correct the structural parameters and identify the atom types. Geometric calculations and the Fourier difference method are used to obtain the positions of all hydrogen atoms. The final reliability factors are R1=0.0322[I≧2σ(I)], wR2=0.0908 (all data), S=1.085, determining the molecular configuration to be trans, the minimum stoichiometric formula to be C8H7N3O5, the corresponding molecular weight to be 225.17, and the calculated crystal density to be 1.679 g / cm³. 3 .

[0015] The beneficial effects of this invention are as follows: For the first time, the solid-state configuration of pharmaceutical furazolidone has been confirmed: single crystals of furazolidone were successfully prepared by solvent evaporation method, and high-precision single-crystal X-ray diffraction technology (MoKα radiation, HyPix-Arc 150 detector) combined with SHELXT program analysis was used to clearly confirm for the first time from the crystallographic level that pharmaceutical furazolidone has a trans configuration in the solid state, providing direct and irrefutable experimental evidence for the stereostructure identification of this drug.

[0016] Revealing the structural nature of photodegradation impurities: Combined with liquid chromatography-mass spectrometry analysis, it was found that the photodegradation product has the same molecular weight as furazolidone. Based on this, it was inferred that the impurity is a cis-trans isomer of its double bond. This discovery not only reasonably explains the intrinsic reason for the drug's photoinstability, but also indirectly confirms that the trans configuration is absolutely dominant in the active pharmaceutical ingredient, providing key data support for tracing the source and structural attribution of photodegradation impurities.

[0017] Improving drug quality control: The superior configuration and photoisomerization pathway of furazolidone have been clarified, providing a scientific basis for establishing a dedicated detection method for cis impurities. This will help to formulate stricter quality standards for active pharmaceutical ingredients and formulations, ensuring the safety and effectiveness of clinical drug use.

[0018] Additional aspects and advantages of the invention will become apparent from the description which follows, or may be learned by practice of the invention. Attached Figure Description

[0019] The above and / or additional aspects and advantages of the present invention will become apparent and readily understood from the description of the embodiments taken in conjunction with the following drawings, in which: Figure 1 This is a stereoscopic ellipsoidal diagram of the single crystal molecule of furazolidone with double bond cis-trans configuration according to an embodiment of the present invention; Figure 2 This is a molecular packing diagram of a single crystal unit cell of furazolidone with double bond cis-trans configuration according to an embodiment of the present invention; Figure 3 This is a 3D ball-and-stick model of a single crystal of furazolidone with double bond cis-trans configuration according to an embodiment of the present invention; Figure 4 This is a liquid chromatogram of a single-crystal sample of furazolidone with double bond cis-trans configuration before photodegradation according to an embodiment of the present invention; Figure 5 This is a liquid chromatography-mass spectrum of a single crystal of furazolidone with double bond cis-trans configuration as described in an embodiment of the present invention. Figure 6 This is a liquid chromatography-mass spectrum of a single crystal of furazolidone with double bond cis-trans configuration as described in an embodiment of the present invention. Figure 7 This is a liquid chromatography-mass spectrum of a single crystal of furazolidone with double bond cis-trans configuration obtained according to an embodiment of the present invention. Detailed Implementation

[0020] To better understand the above-mentioned objectives, features, and advantages of the present invention, the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments. It should be noted that, unless otherwise specified, the embodiments and features described in these embodiments can be combined with each other.

[0021] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be implemented in other ways different from those described herein. Therefore, the scope of protection of the invention is not limited to the specific embodiments disclosed below.

[0022] Example Single crystal growth process Weigh 200 mg of furazolidone and add it to a 10 ml reaction tube. Add a magnetic stir bar and then add 4 ml of acetonitrile. Stir on a magnetic stirrer for 1 hour to fully dissolve the sample. Filter to obtain a saturated solution of furazolidone. Take 1.5 ml of the saturated solution and place it in a 2 ml vial. Seal the vial with sealing film and poke several small holes in the sealing film with a steel needle. Let it stand to allow the acetonitrile to evaporate slowly. Crystals will grow after 5 days.

[0023] Single crystal diffraction data collection The above sample was used to collect data at 100K on a 'ROD, Synergy Custom DW system' single crystal diffractometer with MoKα radiation. The distance between the crystal and the 'HyPix-Arc 150' detector was 60mm. The instrument operating voltage was 45kV, the current was 66mA, the maximum 2θ angle was 60.634°, the scan step size was 0.5°, the scan time was 0.2s / °, and each frame was scanned once, for a total of 536 images, with a total of 8683 diffraction points and 1344 independent diffraction points. int =0.0252], the data integrity is 99.8% when the angle θ reaches 25.242°.

[0024] Single crystal structure analysis The crystals are orange-yellow in shape. The crystal size used for diffraction analysis is 0.24×0.2×0.12mm. It belongs to the orthorhombic crystal system with space group Pnma. The unit cell parameters are: a=22.5308(8)Å, b=6.0608(3)Å, c=6.5249(3)Å, α=90°, β=90°, γ=90°, and the unit cell volume V=891.01(7)Å. 3 The number of molecules in the unit cell is Z=4, and each independent region of the unit lattice contains 0.5 molecules.

[0025] The molecular arrangement in the crystalline state belongs to space group Pnma. The crystal structure was analyzed using the software Olex2 on a computer by calling the program SHELXT. All non-hydrogen atom positions were obtained from the E diagram. The least squares method (program SHELXL) was used to correct the structural parameters and identify the atom types. The positions of all hydrogen atoms were obtained using geometric calculations and the Fourier difference method. The final reliability factors were R1 = 0.0322 [I≧2σ(I)], wR2 = 0.0908 (all data), and S = 1.085. The molecular configuration was determined to be trans, with the minimum stoichiometry C8H7N3O5, corresponding to a molecular weight of 225.17. The calculated crystal density was 1.679 g / cm³. 3 .

[0026] Single crystal structure expression The structure of the molecule is , Figure 1 This is an ellipsoidal diagram of the three-dimensional structure of the molecule. Figure 2 Table 1 shows the molecular packing diagram of the unit cell, Table 2 shows the coordinates and average thermal vibration parameters of non-hydrogen atoms, Table 3 shows the thermal vibration parameters of non-hydrogen atoms, Table 4 shows the bond lengths of non-hydrogen atoms, Table 5 shows the bond angles of non-hydrogen atoms, and Table 6 shows the twist angles of non-hydrogen atoms.

[0027] Table 1. Crystal data and structure optimization

[0028] Table 2. Fractional atomic coordinates (×10⁴) and equivalent isotropic displacement parameters (Ų×10³)

[0029] Table 3. Anisotropic displacement parameters (Ų×10³)

[0030] Table 4. Bond Lengths

[0031] Table 5. Bond angles

[0032] Table 6. Torsion Angle

[0033] Simulated furazolidone ball-and-stick model A 3D ball-and-stick model of furazolidone was simulated using Chem3D 20.0 in ChemOffice 2020. Figure 3The image shows a 3D ball-and-stick model of furazolidone. As can be seen from the image, the parts enclosed in red (the nitrogen and carbon-nitrogen double bonds of the oxazole ring, the hydrogen on the carbon of the double bond, the furan ring, and the nitro group) are on the same plane, which can form cis-trans isomerism of the carbon-nitrogen double bond.

[0034] Sample preparation process and liquid chromatography-mass spectrometry results for photodegradation 1. Preparation process of light-induced damage to test samples Place 20.40 mg in a 20 ml volumetric flask, add about 10 ml of acetonitrile, sonicate to dissolve, add water to the mark, shake well, pour into a transparent vial, and place in an expandable test chamber (BXS-250s, Shanghai Boxun Medical Biological Instrument Co., Ltd.), and irradiate with light and ultraviolet light for 4 hours.

[0035] 2. Liquid chromatography-mass spectrometry (LC-MS) test results Figure 4 This is a liquid chromatogram of the sample before photodegradation of furazolidone. Figures 5 to 7 The liquid chromatography-mass spectrum of the sample after photodegradation of furazolidone. As shown in the above liquid chromatography and mass spectrometry chromatograms, the purity of furazolidone before photodegradation is 100%. After photodegradation, there is a 5.95% photodegradation impurity at 19.50 min. The ES+ signal is the same as that of the main peak of furazolidone at 20.10 min, which is very likely due to the cis-trans isomerism of the C=N double bond in the structure of furazolidone. This further confirms that furazolidone is a stable trans configuration and that a small amount of cis configuration impurity was generated after light irradiation.

[0036] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the invention by those skilled in the art. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the invention should be included within the scope of protection of the invention.

Claims

1. A method for preparing a single crystal of furazolidone with double bond cis-trans configuration, characterized in that: The specific steps of the preparation method are as follows: S1. Add furazolidone to a solvent, vortex to dissolve completely, filter, and obtain a saturated furazolidone solution; S2. Take a saturated solution of furazolidone and place it in a sample vial. Seal the vial with sealing film, make several small holes in the sealing film, let it stand, and allow the acetonitrile to evaporate slowly. This will grow crystals and obtain single crystals of furazolidone with double bond cis-trans configuration.

2. The method for preparing furazolidone single crystals according to claim 1, characterized in that: The mass fraction ratio of furazolidone to solvent in S1 is 0.3-0.5 mg / mL.

3. The method for preparing furazolidone single crystals according to claim 1, characterized in that: The solvent in S1 is acetonitrile.

4. The method for preparing furazolidone single crystals according to claim 1, characterized in that: The vortex in S1 uses a magnetic stirrer, and the vortex time is 1-1.5 hours.

5. The method for preparing furazolidone single crystals according to claim 1, characterized in that: The acetonitrile in S2 evaporates slowly over a period of 3-5 days.

6. A method for determining the cis-trans configuration of a furazolidone single crystal, characterized in that: The specific steps of the determination method are as follows: S1. Single crystal diffraction data collection: A single crystal sample of furazolidone with double bond cis-trans configuration was taken and data was collected at 100K on a 'ROD, SynergyCustom DW system' single crystal diffractometer, MoKα radiation; S2. Single Crystal Structure Analysis: Analysis of the color, size, space group, cell parameters, cell volume, number of molecules within the cell, and number of molecules contained in an independent region of the unit lattice for the double bond cis-trans configuration of furazolidone single crystal.

7. The method for determining the double bond cis-trans configuration of furazolidone single crystal according to claim 6, characterized in that: The distance between the double-bond cis-trans configuration furazolidone single crystal in S1 and the 'HyPix-Arc 150' detector is 60 mm. The instrument operating voltage is 45 kV, the current is 66 mA, the maximum 2θ angle is 60.634°, the scanning step size is 0.5°, the scanning time is 0.2 s / °, and each frame is scanned once, for a total of 536 images, with a total of 8683 diffraction points and 1344 independent diffraction points. int =0.0252], the data integrity is 99.8% when the angle θ reaches 25.242°.

8. The method for determining the double bond cis-trans configuration of furazolidone single crystal according to claim 6, characterized in that: The S2 single crystal of furazolidone with double bond cis-trans configuration is orange-yellow in shape, with a crystal size of 0.24×0.2×0.12mm. It belongs to the orthorhombic crystal system, space group Pnma, and has the following cell parameters: a=22.5308(8)Å, b=6.0608(3)Å, c=6.5249(3)Å, α=90°, β=90°, γ=90°, and cell volume V=891.01(7)Å. 3 The number of molecules in the unit cell is Z=4, and each independent region of the unit lattice contains 0.5 molecules.

9. The method for determining the double bond cis-trans configuration of furazolidone single crystal according to claim 8, characterized in that: The space group Pnma in S2 was analyzed using the software Olex2 on a computer by calling the program SHELXT. All non-hydrogen atom positions were obtained from the E diagram. The least squares method was used to correct the structural parameters and identify the atom types. Geometric calculations and the Fourier difference method were used to obtain the positions of all hydrogen atoms. The final reliability factors were R1 = 0.0322 [I≧2σ(I)], wR2 = 0.0908 (all data), and S = 1.

085. The molecular configuration was determined to be trans configuration, with the minimum stoichiometry being C8H7N3O5, corresponding to a molecular weight of 225.17, and a crystal density of 1.679 g / cm³. 3 .

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