Triphenylacetic acid rafenasin crystal form and preparation method thereof

Abstract

The invention belongs to the technical field of medicinal chemistry, and particularly relates to a crystal form of triphenylacetic acid rafenasin and a preparation method thereof. The crystal form has good chemical stability and crystal form purity, is easy for large-scale preparation, can be better suitable for preparation of pharmaceutical preparations and large-scale production, and has a wide application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical chemistry, and specifically relates to a refenacin triphenylacetate crystal form and a preparation method thereof. The crystal form has good chemical stability and crystal purity, is easy to prepare on a large scale, can be better applied to the preparation of pharmaceutical preparations and large-scale production, and has broad application prospects. Background Art

[0002] Revenacin is a long-acting muscarinic antagonist with a structural formula as shown in Formula 1. It is mainly administered by inhalation solution spray to improve lung function, reduce clinical symptoms of COPD, and prevent further deterioration of the lesion. It is used for the maintenance treatment of COPD patients. In 2018, the FDA approved Revenacin for the maintenance treatment of patients with chronic obstructive pulmonary disease (COPD). COPD is a progressive, irreversible, persistent airway obstructive lung disease with no single cause. It is also a general term for chronic bronchitis and emphysema. COPD is believed to affect approximately 328 million people worldwide. It is already the third leading cause of death in the United States. By 2020, it will become the third leading cause of death worldwide. There is a serious unmet need for COPD treatment drugs.

[0003]

[0004] The original research company has two crystal patents authorized in China, namely CN101163677B and CN102470130B. Among them, CN101163677B discloses three salt forms of diphosphate, monosulfate and hydrogen oxalate, and protects diphosphate. CN102470130B discloses four crystal forms of free base crystal form I, crystal form II, crystal form III and crystal form IV. The DSC peak temperatures of crystal form I and crystal form II are 102.7℃ and 98.64℃ respectively, and the DSC peak temperatures of crystal form III and crystal form IV are 123.1℃ and 118.8℃ respectively. The patent protects crystal form III, which is an anhydrous form with an acceptable degree of hygroscopicity and a high melting point. The patent will expire in China on July 13, 2030.

[0005] Salt formation can improve the physicochemical properties of drug molecules, significantly increase solubility and dissolution rate, and is one of the effective means to improve its drugability. It can also improve drug compliance, improve drug stability, and reduce its adverse reactions. When there are problems with the stability of free bases or free acids, salt forms are the preferred option. For ionizable molecules, salt forms can improve the physicochemical properties of compounds, such as solubility, crystallinity, hygroscopicity, melting point, and stability, which can not only improve bioavailability, but also improve solid-state stability.

[0006] The diphosphate, monosulfate, and hydrogen oxalate disclosed in the original research patent CN101163677B are all obtained by unconventional means such as freeze drying, and mass production cannot be achieved. The subsequent original research patents are all free forms, and there is no literature report on other salt forms, indicating that it is difficult to form a salt of refenacin. The inventor of this application screened 39 acids, and only triphenylacetic acid can form a salt with refenacin, but the yield is low. The diphenylacetic acid, which is very similar in structure, cannot form a salt with refenacin. After the deep optimization of the conditions for the formation of salts of triphenylacetic acid, refenacin triphenylacetic acid can be stably prepared. The solid state of refenacin triphenylacetic acid finally prepared is good, and TG, DSC, PXRD, mass spectrum, hydrogen spectrum, and carbon spectrum all show that it has formed a salt, and there is currently no relevant literature or patent report on this salt form.

[0007] The drug has been disclosed in various salt forms, but there is no report on the triphenylacetic acid crystal form of the drug. The inventors of the present application obtained the triphenylacetic acid salt form of the drug in the process of preparing revenacin. This crystal form has a high melting point, good chemical stability and crystal purity, is easy to prepare on a large scale, can be better applied to the preparation of pharmaceutical preparations and large-scale production, and has broad application prospects. Summary of the invention

[0008] The invention belongs to the technical field of pharmaceutical chemistry, and specifically relates to a refenacin triphenylacetate crystal form and a preparation method thereof. The crystal form has good chemical stability and crystal purity, is easy to prepare on a large scale, can be better applied to the preparation of pharmaceutical preparations and large-scale production, and has broad application prospects.

[0009] The object of the present invention is to provide a new salt form (triphenylacetate form) of revenacin compound and a preparation method thereof. The specific technical scheme of the present invention is as follows:

[0010] The structural formula of refenacin triphenylacetate is as follows:

[0011]

[0012] The X-ray powder diffraction pattern of refenacin triphenylacetate has characteristic peaks at 2θ diffraction angles of 6.07±0.2°, 12.05±0.2°, 18.58±0.2°, 20.79±0.2°, and 21.46±0.2°.

[0013] Furthermore, the X-ray powder diffraction pattern of the crystal form has characteristic peaks in 2θ diffraction angles at 3.27±0.2°, 6.94±0.2°, 10.19±0.2°, 13.85±0.2°, 14.87±0.2°, 15.83±0.2°, 22.61±0.2°, 24.24±0.2°, and 31.55±0.2°.

[0014] The refenacin triphenylacetate crystal form has the following characteristics: Figure 1 The X-ray powder diffraction peak pattern is shown.

[0015] The DSC analysis of the crystal form has a peak value at 172.9±3.0°C thermal absorption peak, and further, has the following Figure 2 DSC spectrum shown.

[0016] The TG analysis of the crystal form shows thermal weight loss of 7.98±2% and 53.99±2%, respectively, and further has the following characteristics: Figure 3 DSC spectrum shown.

[0017] The infrared spectrum of the triphenylacetate refenacin is 3401±2cm -1 、3290±2cm -1 、3149±2cm -1 、1717±2cm -1 、1673±2cm -1 、1624±2cm -1 There is infrared absorption at. Further, it has the infrared spectrum characteristics as shown in the figure.

[0018] The preparation method of the refenacin triphenylacetate crystal form of the present invention comprises: (1) adding refenacin to an acetone solution, heating and stirring until dissolved; (2) adding triphenylacetic acid, cooling and crystallizing, separating the crystals, and drying.

[0019] Preferably, the ratio of acetone to refenacin is 1-30 ml:1 g, more preferably 5-10 ml:1 g.

[0020] Preferably, the equivalent of the added triphenylacetic acid is 1 to 3, more preferably 1.2 to 1.5.

[0021] Preferably, the crystallization temperature is 0 to 30°C, more preferably 20 to 25°C.

[0022] Advantages of the present invention:

[0023] The invention relates to a refenacin triphenylacetate crystal form, which has a high DSC, is very stable under conditions of high temperature, high humidity, light, etc., has better chemical stability and crystal purity, has simple preparation process steps, is easy to produce and operate, has high yield and purity, is suitable for industrial large-scale production, and has high industrial production value. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 X-ray powder diffraction pattern of revenacin triphenylacetate.

[0025] Figure 2Differential scanning calorimetry (DSC) curve of revenacin triphenylacetate.

[0026] Figure 3 Thermogravimetric analysis (TG) curve of revenacin triphenylacetate.

[0027] Figure 4 Infrared test image of revenacin triphenylacetate.

[0028] Figure 5 Mass spectrum of refenacin triphenylacetate.

[0029] Figure 6 H NMR spectrum of refenacin triphenylacetate.

[0030] Figure 7 NMR carbon spectrum of refenacin triphenylacetate. DETAILED DESCRIPTION

[0031] The present invention is described in detail below in conjunction with specific implementation methods and exemplary embodiments, but these descriptions cannot be understood as any limitation of the present invention. Those skilled in the art understand that, without departing from the spirit and scope of the present invention, a variety of equivalent substitutions, modifications or improvements can be made to the technical solutions and implementation methods of the present invention, all of which fall within the scope of the present invention. The scope of protection of the present invention shall be subject to the attached claims. Specific embodiments are described as follows:

[0032] Example 1 Preparation of refenacin free base (refer to Chinese patent CN101163677B)

[0033] Add isopiperoidinecarboxamide (59.9 g, 400 mmol), acetic acid (25.7 ml), sodium sulfate (64.4 g) and isopropanol (4000 ml) to a 20 L reactor. Cool to 0-10 °C and slowly add a solution of 1-{2-[(4-formylbenzoyl)methylamino]ethyl}piperidin-4-ylbiphenyl-2-ylcarbamate (110 g, 227 mmol) in isopropanol (3000 ml). React at room temperature for about 2 h and cool to 0-10 °C. Add sodium triacetoxyborohydride (151.6 g, 685 mmol) in portions and react at room temperature for about 16 h. Concentrate the reaction solution to a volume of about 500 ml and adjust the pH to about 3 with 1N HCl (about 2000 ml). Dichloromethane (3

[0034] × 2500ml). Cool to 0-5°C, adjust pH to about 10 with 50% NaOH aqueous solution. Extract with isopropyl acetate (3×3000ml), wash the organic layer with water (1000ml) and saturated sodium chloride aqueous solution (2×500ml), dry with anhydrous sodium sulfate, filter and concentrate to obtain refenacin free base.

[0035] Example 2 Preparation of revenacin diphosphate (refer to Chinese patent CN101163677B)

[0036] Dissolve 5.0 g of refenacin free base in 50 ml of water and 15 ml of 1M phosphoric acid. Adjust the pH to about 5.3 with 2.5 ml of 1M phosphoric acid. Filter and freeze-dry to obtain an amorphous diphosphate. Dissolve the amorphous diphosphate in 20 ml of isopropanol: acetonitrile (1:1). Add 1 ml of water, heat to 60°C, cool to 20-25°C, continue stirring for about 12 hours, filter, and vacuum dry to obtain refenacin diphosphate.

[0037] Example 3 Preparation of revenacin crystal form III (refer to Chinese patent CN102470130B)

[0038] Dissolve 3.07 g of refenacin free base in 15 ml of acetonitrile and stir at room temperature for about 80 min. Place the mixture in an oscillator block for thermal cycling (0°C to 40°C in a 1 h block) for 48 h. Add 15 ml of acetonitrile and then place the mixture back in the oscillator block and keep for 2 h. Filter and vacuum dry to obtain refenacin Form III.

[0039] Example 4 Preparation of rafenacin monosulfate (refer to Chinese patent CN101163677B)

[0040] Dissolve 44.2 g of refenacin free base in 500 ml of H2O:ACN (1:1), slowly add about 145 ml of 1N sulfuric acid, adjust the pH to about 3.3, filter through a 0.2 micron filter, and freeze-dry to obtain the monosulfate salt.

[0041] Dissolve 3.0 g of monosulfate in 165 ml of IPA:ACN (10:1), add to a 60°C water bath and stir for 30 min, heat to 70°C and stir for 30 min, cool to 60°C and stir for 1 h. Cool to room temperature, leave at room temperature for about 13 days, filter, and vacuum dry to obtain refenacin monosulfate.

[0042] Example 5 Preparation of revenacin monooxalate (refer to Chinese patent CN101163677B)

[0043] 51.0 g of revenacin free base was dissolved in 500 ml of H2O:ACN (1:1), and about 170 ml of 1M aqueous oxalic acid was slowly added to adjust the pH to about 3.0. The mixture was filtered through a 0.2 micron filter and freeze-dried to obtain the hydrogen oxalate salt.

[0044] Dissolve 3.2 g of hydrogen oxalate in 276 ml of 94% IPA / 6% H2O, add to a 60°C water bath and stir for 2.5 hours. Cool to room temperature, place at room temperature for about 30 days, filter, and vacuum dry to obtain revenacin monooxalate.

[0045] Example 6 Study on the preparation of revenacin triphenylacetate

[0046] 10.0 g of refenacin free base was added to a round-bottom flask, followed by the addition of different reaction solvents in amounts shown in Table 1, and the addition of triphenylacetic acid in amounts shown in Table 1, and stirring was started, and the temperature was raised to dissolve. The temperature was then lowered for crystallization, filtration was performed, and vacuum drying was performed to obtain each group of triphenylacetic acid refenacin. The experimental results are shown in Table 1.

[0047] Table 1 Preparation of refenacin triphenylacetate

[0048]

[0049]

[0050] *: The negligible limit is 0.02%, after deducting the influence of corresponding acid radicals.

[0051] PXRD,DSC,TG,IR, 1 H NMR, 13 H NMR and related substances detection methods are as follows:

[0052] X-ray powder diffraction (PXRD): model Empyrean, step width 0.026°, step time 50 s.

[0053] Differential scanning calorimeter (DSC): Model DSC214Polyma, nitrogen purge gas 40ml / min, protective gas 60ml / min. Heating rate 10K / min. Temperature range 25~200℃.

[0054] Thermogravimetric analyzer (TG): Model: TG 209F3. Nitrogen purge gas 20 ml / min, protective gas 20 ml / min. Heating rate 10 K / min. Temperature range 25~400℃.

[0055] Infrared spectrometry test: Model: Nicolet iS5 infrared spectrophotometer, and KBr pellet detection was used.

[0056] High-resolution mass spectrometer (MS): Model: Waters H-Class / G2-XS QTOF LC-MS

[0057] Nuclear Magnetic Resonance Spectrometer ( 1 H NMR, 13 C NMR): Model: Bruker AVANCEⅢ-600, Solvent: DMSO-d6

[0058] High performance liquid chromatography (HPLC): model waters e2695, mobile phase: 0.01 mol / L ammonium dihydrogen phosphate buffer (pH adjusted to 2.9 with phosphoric acid)-acetonitrile (95:5) as mobile phase A, acetonitrile as mobile phase B, column temperature: 40°C, detection wavelength: 220 nm, chromatographic column: YMC Triart C18 column (4.6 mm×150 mm, 3 μm).

[0059] The X-ray powder diffraction pattern of refenacin triphenylacetate is as follows Figure 1 As shown, the differential scanning calorimetry spectrum is as Figure 2 As shown in the thermogravimetric analysis (TG) curve Figure 3 As shown, the infrared spectrum is Figure 4 As shown in the high-resolution mass spectrum Figure 5 As shown, the H NMR spectrum is Figure 6 As shown, the NMR carbon spectrum is as follows Figure 7 shown.

[0060] Example 7 Study on organic / inorganic acid salt formation

[0061] This example screened 39 acids, and ultimately only triphenylacetic acid could form a salt with revenacin. The preparation process during screening was the same as that of Example 6, No. 2. The screening results are shown in Table 2.

[0062] Table 2 Results of screening of refenacin salts

[0063]

[0064]

[0065] Example 8 Study on the solubility of revenacin triphenylacetate and other salt forms and crystal forms of the present invention

[0066] Weigh 100 mg of the test sample, place it in 2 ml of purified water at 25 ± 2 ° C, shake vigorously for 10 seconds every 1 minute, and observe the dissolution within 3 minutes. If there are no visible solute particles, it is considered to be completely dissolved; if there are visible solute particles, add 2 ml of water and repeat the above operation until it is completely dissolved. Record the total amount of water used and the total time. Refenacin Form III was prepared with reference to Chinese patent document CN102470130A. The results are shown in Table 3.

[0067] Table 3 Solubility study of refenacin triphenylacetate and other crystal forms

[0068]

[0069] As shown in Table 3, the revenacin triphenylacetate of the present invention has better solubility and faster dissolution rate than other salt forms and crystal forms, and can be better used in formulation development.

[0070] Example 9 Stability Study of Refenacin Triphenylacetate and Other Salt Forms of the Present Invention

[0071] Stability test conditions include:

[0072] 1. Thermal degradation: Take about 200 mg of refenacin triphenylacetate and place it in a drying oven at 60°C;

[0073] 2. Photodegradation: Take about 200 mg of refenacin triphenylacetate and place it in an environment with an illumination of 4500±500lx;

[0074] 3. High humidity degradation: Take about 200 mg of revenacin triphenylacetate and place it in a desiccator containing a saturated KNO3 solution at room temperature.

[0075] The test methods for related substances of revenacin are as follows:

[0076] Method basis: High performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition Part IV General Rules 0512)

[0077] Chromatographic conditions:

[0078] High performance liquid chromatography (HPLC): model waters e2695, mobile phase: 0.01 mol / L ammonium dihydrogen phosphate buffer (pH adjusted to 2.9 with phosphoric acid)-acetonitrile (95:5) as mobile phase A, acetonitrile as mobile phase B, column temperature: 40°C, detection wavelength: 220 nm, chromatographic column: YMC Triart C18 column (4.6 mm×150 mm, 3 μm).

[0079] Gradient elution:

[0080] Time (min) Mobile phase A% Mobile phase B% 0 85 15 3 85 15 30 35 65 35 35 65 37 85 15 45 85 15

[0081] Solution preparation:

[0082] (1) Diluent: mobile phase A.

[0083] (2) Test solution: Take an appropriate amount of the product, weigh it accurately, add an appropriate amount of acetonitrile to dissolve it, and quantitatively dilute it with mobile phase A to make a solution containing about 1 mg per 1 ml.

[0084] (3) Control solution: Accurately measure an appropriate amount of the test sample solution and quantitatively dilute it with mobile phase A to make a solution containing approximately 1 μg per 1 ml.

[0085] The results of the stability study are shown in Table 4.

[0086] Table 4 Stability study of revenacin triphenylacetate, revenacin diphosphate, revenacin monosulfate, revenacin monooxalate and revenacin crystal form III

[0087]

[0088] Note: The negligible limit is 0.02%, after deducting the influence of corresponding acid radicals.

[0089] The DSC peak of refenacin triphenylacetate is 172.9℃, which is much higher than 154.5℃ of diphosphate, about 57℃ and 73.2℃ of monosulfate, 69.2℃ and 122.8℃ of monooxalate and 123.1℃ of crystal form III. Theoretically, it should have good chemical stability. As shown in Table 4, diphosphate, monosulfate, monooxalate and crystal form III have a tendency to deteriorate under conditions of light, high temperature and high humidity, especially under high temperature. In comparison, refenacin triphenylacetate has better stability, no significant increase in impurities, and almost no decrease in purity, indicating that refenacin triphenylacetate has better stability than other salts of refenacin triphenylacetate and free base crystal form III.

[0090] Example 10 Preparation of revenacin triphenylacetate inhalation powder

[0091] 1) Weigh 0.087g of magnesium stearate and 2.446g of refenacin triphenylacetate, stir and mix for 20min, transfer to a jet mill and grind to powder at 5 bar pressure. 10 :0.6μm; D 50 :2.7μm; D 90 : 5.0 μm, and the refenacin complex was obtained.

[0092] 2) Weigh the remaining amount of magnesium stearate (0.313 g) and the total amount of lactose (327.154 g) and place them in a TRV high shear mixer, mix them at a mixing speed of 500.0 r / min for 6 min, and discharge the mixture to obtain a lactose premix.

[0093] 3) Add about 1 / 2 of the lactose premix, all of the refenacin complex and the remaining about 1 / 2 of the lactose premix to the TRV mixing device in sequence, set the mixing speed to 1350 r / min, mix for 3 min, discharge, and obtain the refenacin total mixture.

[0094] 4) Take the revenacin total mixture powder and fill it into capsules according to the filling amount of 25 mg±1 mg to prepare revenacin inhalation powder capsules that can be delivered to the lungs for administration through a dry powder inhalation device.

[0095] 5) Double-aluminum packaging is performed on the above-mentioned drug-loaded capsules to obtain the product.

[0096] Table 5 Prescription list of revenacin inhalation powder (revenacin triphenylacetate)

[0097] Element Prescription dosage effect lactose 327.154g Carrier Magnesium Stearate 0.40g Dispersing aids Refenacin (triphenylacetate) 2.446g Active ingredients

[0098] Example 11 Preparation of revenacin diphosphate, revenacin monosulfate, revenacin monooxalate and revenacin crystal form III inhalation powder

[0099] Weigh 0.087g of magnesium stearate and 2.191g of revenacin diphosphate and stir for 20min, then transfer to a jet mill and grind to a fine powder of D at a pressure of 5 bar. 10 :0.6μm; D 50 :2.5μm; D 90 : 4.9 μm, and the revenacin complex was obtained. The revenacin diphosphate inhalation powder was prepared by referring to the method of Example 10.

[0100] Weigh 0.087g of magnesium stearate and 1.650g of refenacin crystal form III raw materials, stir and mix for 20 minutes, transfer to a jet mill and grind at 5 bar pressure to co-fine powder to D 10 :0.5μm; D 50 :2.3μm; D 90 : 4.5 μm, and the revenacin complex was obtained. The revenacin crystal form III inhalation powder was prepared according to the method of Example 10.

[0101] Weigh 0.087g of magnesium stearate and 1.921g of refenacin monosulfate, stir and mix for 20min, transfer to a jet mill and grind to powder D at 5 bar pressure. 10 :0.7μm; D 50 :2.7μm; D 90 : 5.5 μm, and the revenacin complex was obtained. The revenacin monosulfate inhalation powder was prepared by referring to the method of Example 8.

[0102] Weigh 0.087g of magnesium stearate and 1.899g of revenacin monooxalate, stir and mix for 20min, transfer to a jet mill and grind to powder D at 5 bar pressure. 10 :0.6μm; D 50 :2.4μm; D 90 : 4.8 μm, and the revenacin complex was obtained. Referring to the method of Example 8, the revenacin monooxalate inhalation powder was prepared.

[0103] Example 12 Study on the stability of different revenacin inhalation powder preparations

[0104] The different revenacin inhalation powders prepared in Examples 10 and 11 were placed in an environment of 40°C ± 2°C and RH 75% ± 5%, and the changes in the fine particle dosage, fine particle fraction, content and related substances of the products were measured at regular intervals. The results are shown in Table 5.

[0105] The testing methods for the relevant substances are as follows:

[0106] Method basis: High performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0107] Chromatographic conditions:

[0108] High performance liquid chromatography (HPLC): model waters e2695, mobile phase: 0.01 mol / L ammonium dihydrogen phosphate buffer (pH adjusted to 2.9 with phosphoric acid)-acetonitrile (95:5) as mobile phase A, acetonitrile as mobile phase B, column temperature: 40°C, detection wavelength: 220 nm, chromatographic column: YMC Triart C18 column (4.6 mm×150 mm, 3 μm).

[0109] Gradient elution:

[0110] Time (min) Mobile phase A% Mobile phase B% 0 85 15 3 85 15 30 35 65 35 35 65 37 85 15 45 85 15

[0111] Solution preparation:

[0112] (1) Diluent: mobile phase A.

[0113] (2) Test solution: Take 10 capsules of this product and transfer the contents to a 25 ml volumetric flask. Wash the capsule shells with 10 ml of mobile phase A in batches. Add the washing solution to the volumetric flask and shake to dissolve. Dilute to the mark with mobile phase A and shake well.

[0114] (3) Control solution: Accurately measure 1 ml of the test solution and place it in a 100 ml volumetric flask. Add diluent to dilute to the mark.

[0115] Table 6 Stability test conditions

[0116] Inspection conditions specific conditions accelerate Temperature: 40℃±2℃, relative humidity 75%±5%

[0117] Table 7 Stability test results

[0118]

[0119] Table 7: Stability test results

[0120]

[0121] The fine particle dose and fine particle fraction of the inhalation powder prepared with refenacin triphenylacetate were significantly higher than those of the inhalation powder prepared with refenacin diphosphate and crystalline form III. In the stability study, the fine particle dose and fine particle fraction of refenacin triphenylacetate inhalation powder remained stable within 3 months of acceleration, while the fine particle dose and fine particle fraction of refenacin diphosphate and crystalline form III inhalation powder showed a downward trend within 3 months of acceleration. The related substances of refenacin triphenylacetate inhalation powder increased slowly within 3 months of acceleration, while the related substances of refenacin diphosphate and crystalline form III inhalation powder increased significantly within 3 months of acceleration. The inhalation powder prepared with refenacin monosulfate and refenacin oxalate had the worst fine particle dose and fine particle fraction, and the related substances of both increased most significantly in the stability study.

[0122] In summary, the inhalation powder prepared using refenacin triphenylacetate has the best performance in the stability study. Example 13 Pharmacokinetic performance of different refenacin inhalation powders in rats

[0123] Male SD rats (SPF grade) were used in this embodiment, aged 9 months and weighing 180-220g. During the whole experiment, the rats were free to eat and drink. The SD rats for the experiment were grouped by randomized block design and divided into three groups: refenacin triphenylacetate crystalline inhalation powder group, refenacin diphosphate inhalation powder group and refenacin crystalline III inhalation powder group. The drug was administered by oral and nasal inhalation in rats, and the dosage was set to 375μg / kg (in terms of refenacin).

[0124] 0.5 ml of venous blood was collected from the fundus venous plexus at 0.1 h, 0.2 h, 0.5 h, 2 h, 6 h, 12 h, 24 h, 48 h, 60 h, 72 h, 96 h and 120 h after administration and placed in a pre-labeled EDTA (4 mM) anticoagulated EP tube. After whole blood collection, it was placed on ice and then centrifuged at 4 ° C, 8000 rpm, 5 min to collect plasma, transferred to a 96-well plate, and stored at -20 ° C until LC-MS / MS detection. The drug concentration in EDTA (4 mM) anticoagulated SD rat plasma was determined using the LC / MS / MS (Agilent 6460) method. WinNonlin 5.2 software was used to calculate the relevant pharmacokinetic parameters of the concentration at each time point after administration of each group according to the statistical distance theory. See Table 8 for details.

[0125] Table 8 Pharmacokinetic performance of different crystalline forms of revenacin inhalation powder in rats

[0126]

[0127] The pharmacokinetic results in rats showed that revenacin triphenylacetate inhalation powder had the lowest peak blood concentration C max and AUC 0-∞ , indicating that the safety of revifenacin triphenylacetate inhalation powder is significantly better than that of revifenacin diphosphate inhalation powder and revifenacin form III inhalation powder.

[0128] Example 14 Tissue distribution of different crystalline forms of revenacin inhalation powder in rats

[0129] Preparation selection: revifenacin triphenylacetate inhalation powder prepared in Examples 10 and 11, revifenacin diphosphate inhalation powder, revifenacin crystal form III inhalation powder

[0130] Dosing frequency: Single dose

[0131] Route of administration: Inhalation through the mouth and nose

[0132] Experimental animals: SD rats

[0133] Grouping: A total of 108 SD rats were divided into 375 μg / kg (calculated as refenacin) dose group of refenacin triphenylacetate inhalation powder, 375 μg / kg (calculated as refenacin) dose group of refenacin diphosphate inhalation powder and 375 μg / kg (calculated as refenacin) dose group of refenacin crystal form III inhalation powder, 36 rats in each group. The drug concentrations of animal bronchoalveolar lavage fluid, lung tissue, main trachea and bronchus, bladder, heart, liver and kidney were collected at 0.5 h after administration.

[0134] Table 9 Distribution of revenacin in major organs and tissues in rats after inhalation of two preparations for 0.5 h

[0135]

[0136] Revenacin is a long-acting anticholinergic drug that has similar affinity to the subtypes of sarcosine receptors M1-M5. In the airway, it exhibits pharmacological effects by inhibiting smooth muscle M3 receptors to cause bronchial dilation. Smooth muscle M3 receptors are mainly located in the trachea and bronchi and are the main pharmacodynamic site of Revenacin. The above-mentioned animal tissue distribution test shows that 0.5h after inhalation administration, the concentration of refenacin triphenylacetate inhalation powder in the lung tissue, main trachea and bronchus at the drug effect site is significantly higher than that of refenacin diphosphate inhalation powder and refenacin crystal form III inhalation powder, while in the bladder, heart and other places where clinical side effects (such as urinary retention) may occur, the concentration of refenacin triphenylacetate inhalation powder is significantly lower than that of the other two crystal forms of refenacin inhalation powder, indicating that refenacin triphenylacetate inhalation powder may produce a higher bronchodilator effect than refenacin diphosphate inhalation powder and refenacin crystal form III inhalation powder and produce lower clinical side effects.

Claims

1. A crystalline form of refenacin triphenylacetate, wherein the crystalline form of refenacin triphenylacetate has the following structural formula: Features: The X-ray powder diffraction spectrum of the crystal form has characteristic peaks at 2θ diffraction angles of 6.07±0.2°, 12.05±0.2°, 18.58±0.2°, 20.79±0.2°, and 21.46±0.2°.

2. The refenacin triphenylacetate crystal form according to claim 1, characterized in that: The X-ray powder diffraction pattern of the crystal form further has characteristic peaks at 2θ diffraction angles of 3.27±0.2°, 6.94±0.2°, 10.19±0.2°, 13.85±0.2°, 14.87±0.2°, 15.83±0.2°, 22.61±0.2°, 24.24±0.2°, and 31.55±0.2°.

3. The refenacin triphenylacetate crystal form according to claim 2, characterized in that: It has an X-ray powder diffraction peak pattern as shown in FIG1 .

4. The refenacin triphenylacetate crystal form according to any one of claims 1 to 3, characterized in that: The DSC analysis of the crystal form has a thermal absorption peak at a peak value of 172.9±3.0°C.

5. The refenacin triphenylacetate crystal form according to claim 4, characterized in that: It has a DSC spectrum as shown in Figure 2.

6. The refenacin triphenylacetate crystal form according to any one of claims 1 to 3, characterized in that: The TG analysis of the crystal form showed thermal weight losses of 7.98±2% and 53.99±2%, respectively.

7. The refenacin triphenylacetate crystal form according to claim 6, characterized in that: It has a TG spectrum as shown in FIG3 .

8. The refenacin triphenylacetate crystal form according to any one of claims 1 to 3, characterized in that: The infrared spectrum of the crystal form is at 3401±2cm -1 、3290±2cm -1 、3149±2cm -1 、1717±2cm -1 、1673±2cm -1 、1624±2cm -1 There is infrared absorption.

9. The refenacin triphenylacetate crystal form according to claim 8, characterized in that It has the infrared spectrum characteristics shown in FIG6 .

10. The method for preparing the revenacin triphenylacetate crystal form according to any one of claims 1 to 9, characterized in that: The steps include: a) adding revenacin and acetone into a reactor, heating and stirring until dissolved; b) adding triphenylacetic acid, cooling and crystallizing, separating the crystals and drying them.

11. The preparation method according to claim 10, characterized in that: The ratio of acetone to refenacin in step a) is 5-30 ml: 1 g; the equivalent of triphenylacetic acid in step b) is 1-3; and the crystallization temperature in step b) is 0-30°C.

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