Preparation method of ubenimex gamma crystal form

文档序号：163010 发布日期：2021-10-29 浏览：61次中文

阅读说明：本技术 一种乌苯美司γ晶型的制备方法 (Preparation method of ubenimex gamma crystal form ) 是由卢铁刚邵波李晓迅刘时奎王颖于 2018-07-09 设计创作，主要内容包括：本发明涉及一种乌苯美司γ晶型制备,通过该制备方法过程中的各关键参数控制,本申请制备方法能够显著提高γ晶型的收率,并得到单一晶型,工艺重现性好,使得实现工业化生产条件可控,有利于大规模产业化。(The invention relates to preparation of ubenimex gamma crystal form, and the preparation method can obviously improve the yield of the gamma crystal form and obtain single crystal form through controlling each key parameter in the preparation process, has good process reproducibility, realizes controllable industrial production conditions and is beneficial to large-scale industrialization.)

1. The preparation method of ubenimex gamma crystal form is characterized by comprising the following steps:

(1) stirring and pulping ubenimex in purified water, wherein the dosage ratio of ubenimex to purified water is 1 g: (5 ml-30 ml), stirring and beating temperature is 40-70 ℃, stirring and beating time is 2-16 h, and stirring speed is 200 r/min-500 r/min;

(2) cooling the reaction liquid obtained in the step (1) to 0-20 ℃, stirring and crystallizing, wherein the crystallization time is 0.5-3 h;

(3) carrying out suction filtration on the solution obtained in the step (2);

(4) putting the filter cake obtained in the step (3) into a vacuum oven at 50-80 ℃ for drying for 4-24 h;

wherein the gamma crystal has a diffraction angle as follows: 3.238 + -0.2, 6.479 + -0.2, 12.999 + -0.2, 15.960 + -0.2, 17.441 + -0.2, 18.560 + -0.2, 19.560 + -0.2, 20.181 + -0.2, 23.602 + -0.2 and 26.122 + -0.2.

2. The preparation method according to claim 1, wherein the ratio of ubenimex to purified water in step (1) is 1 g: (10 ml-15 ml), the stirring and beating temperature is 50-60 ℃, the stirring and beating time is preferably 2-4 h, and the stirring speed is 300 r/min-400 r/min.

3. The preparation method according to claim 1, wherein the stirring crystallization temperature in the step (2) is 0-10 ℃ and the crystallization time is 1-2 h.

4. The preparation method according to claim 1, wherein the drying time in the step (4) is 8 to 16 hours, and the temperature is 70 to 80 ℃.

5. The method of claim 1, comprising the steps of:

(1) dispersing 19.40g of ubenimex in 194ml of purified water, stirring and pulping at the temperature of 45 ℃, stirring and pulping for 3 hours, wherein the stirring speed is 360 r/min;

(2) cooling the reaction liquid obtained in the step (1) to 5 ℃, and stirring for crystallization for 1 h;

(3) carrying out negative pressure suction filtration on the reaction liquid obtained in the step (2);

(4) and (4) putting the filter cake obtained in the step (3) into a vacuum oven at 70 ℃ for drying for 8 hours to obtain 17.90g of white solid powder with the yield of 92.3%.

6. The method of claim 1, comprising the steps of:

dispersing 10.0g ubenimex in 150ml purified water, stirring and pulping at 40 ℃ for 16h, stirring at the speed of 500r/min, cooling to 10 ℃ in an ice bath, and stirring and crystallizing for 1 h; and (3) performing suction filtration under negative pressure, and drying the filter cake in a vacuum oven at 50 ℃ for 16h to obtain 9.07g of white solid powder with the yield of 90.07%. HPLC purity 99.94%, max mono-heterozygote 0.03%, total heterozygote 0.06%.

Technical Field

The application relates to the field of pharmaceutical crystal forms, in particular to an ubenimex gamma crystal form and a preparation method thereof.

Background

Ubenimex (Ubenimex), its chemical name: n- [ (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyryl]-L-leucine; the molecular formula is as follows: c₁₆H₂₄N₂O₄(ii) a Molecular weight: 308.37, respectively; the structural formula is as follows:

ubenimex is a dipeptide compound separated from the culture solution of streptomyces olivorum by merzuschin who was a japanese scholarler in 1976, and can competitively inhibit aminopeptidase B, leucine peptidase, caspase, etc., induce apoptosis of tumor cells and promote host immune function. Can be used in combination with chemotherapy and radiotherapy for treating leukemia, multiple myeloma, myelodysplastic syndrome, hematopoietic stem cell transplantation, and other solid tumor patients.

The preparation method of ubenimex provided by the US patent US4281180 comprises the following steps: dissolving ubenimex with 1N hydrochloric acid, decolorizing with activated carbon, separating insoluble substances, adjusting pH of the separated substance to 5-6 with dilute ammonia water, filtering, collecting precipitated solid, washing with acetone, and vacuum drying to obtain ubenimex beta crystal form.

US4786754 discloses two processes for preparing ubenimex gamma crystalline form and indicates that the beta crystalline form will become the alpha crystalline form of dihydrate by absorbing moisture when placed in air. Firstly, heating alpha and beta crystal forms at 148-200 ℃ to convert the alpha and beta crystal forms into gamma crystal forms, but the method has high crystal conversion temperature, so that related substances of products are increased; secondly, the alpha crystal form and the beta crystal form are dispersed in acetone, methyl ethyl ketone, diethyl ketone, tetrahydrofuran, ethyl acetate, dioxane, isopropanol and methanol water solution and stirred to obtain the gamma crystal form, and the solvents have high toxicity and serious environmental pollution.

Chinese patent CN101891647A discloses a process of using glacial acetic acid as a solvent, hydrogenating a compound (12) with palladium on carbon to obtain a crude ubenimex product, then dissolving the crude ubenimex product in hydrochloric acid, adjusting the pH to 5-6 with ammonia water, filtering, soaking a wet ubenimex product in acetone, filtering, and vacuum drying to obtain a finished ubenimex product, which is substantially the same as the process disclosed in US4786754, and soaking in an organic solvent acetone to obtain a ubenimex gamma crystal form.

Chinese patent CN103910648A provides a preparation method of ubenimex hydrochloride crystal form compound: adding ubenimex hydrochloride into a mixed solvent of isopropyl acetate and dimethylformamide for slightly heating and dissolving, stirring for 1-5h, and standing to volatilize the solvent. The method has long volatilization time, is only suitable for preparing small-dose crystal forms, and cannot meet the requirement of preparing large-batch crystal forms.

Therefore, there is still a need in the art to provide gamma crystals with a simple preparation process and a better product quality, and the present invention meets such a need.

Disclosure of Invention

In the continuous research process, a novel preparation method of the gamma crystal form of ubenimex is unexpectedly obtained through experiments.

The application discloses a preparation method of ubenimex gamma crystal form, which specifically comprises the following steps:

(2) cooling the reaction liquid obtained in the step (1) to 0-20 ℃, stirring and crystallizing, wherein the crystallization time is 0.5-3 h;

(3) carrying out suction filtration on the solution obtained in the step (2);

(4) and (4) putting the filter cake obtained in the step (3) into a vacuum oven at the temperature of 50-80 ℃ for drying for 4-24 h.

In the step (1), the dosage ratio of ubenimex to purified water is preferably 1 g: (10 ml-15 ml), the stirring and beating temperature is preferably 50-60 ℃, the stirring and beating time is preferably 2 h-4 h, and the stirring speed is 300 r/min-400 r/min.

In the step (2), the stirring crystallization temperature is preferably 0-10 ℃, and the crystallization time is preferably 1-2 h.

In the step (4), the drying time is 8-16 h, and the temperature is 70-80 ℃.

The ubenimex gamma crystal form prepared by the method has the following advantages:

1. in the prior art, the preparation of the gamma crystal form is that the alpha and beta crystal forms of ubenimex are prepared firstly, and then dried at the temperature of 148 plus 200 ℃ for crystal transformation, and the temperature is higher, so that related substances are obviously increased (the maximum single impurity before crystal transformation is 0.06%, the total impurity is 0.19%, the maximum single impurity after crystal transformation is 1.13%, and the total impurity is 2.04%), which is obviously higher than the limit of the single impurity and the total impurity in the raw material medicine in the national medicine registration process, and does not meet the product quality requirement. The other preparation method is to stir and convert alpha and beta crystal forms in an organic solvent, wherein the used solvent comprises methyl ethyl ketone, diethyl ketone, ethyl acetate, dioxane, tetrahydrofuran and the like, and the solvents have large dosage (1 g: 10ml), high generation cost, large toxicity and serious environmental pollution. The preparation process of the crystal form has mild reaction conditions and simple operation, and the used solvent is only purified water, so that the preparation process is beneficial to environmental protection, reduces the cost and energy consumption of reagents and has a certain refining effect on products. The obtained product has high quality and purity and low content of related substances, is obviously superior to the prior art, and meets the registration requirements of related raw material medicines.

2. Through the control of each key parameter in the preparation process of the gamma crystal form, the preparation method can obviously improve the yield of the gamma crystal form, obtain a single crystal form, has good process reproducibility, realizes controllable industrial production conditions, and is beneficial to large-scale industrialization.

Drawings

FIG. 1 HPLC chromatogram of ubenimex gamma crystal form of example 1

FIG. 2 XRPD pattern of ubenimex gamma crystalline form example 1

FIG. 3 Infrared spectrum of ubenimex gamma crystal form of example 1

FIG. 4 XRPD pattern of ubenimex beta crystalline form of comparative example

Detailed Description

The present application will be described in further detail with reference to the following examples, which are only for illustrating the technical solutions of the present application and are not intended to limit the spirit and scope of the present application.

The structure of the compound is determined by nuclear magnetic resonance¹HNMR) assayAnd (4) determining. Nuclear magnetic resonance (¹H NMR) shifts (δ) are given in parts per million (ppm). The instrument comprises the following steps: a Bruker AVII-600MHz nuclear magnetic resonance spectrometer; the detection basis is as follows: 0441 NMR spectroscopy in the pharmacopoeia of the people's republic of China 2015 edition, four general rules; solvent: DMSO, internal standard TMS.

The X-ray powder diffraction (XRPD) measurement described in this application is collected by Liaoning Danhao Yuan DX-2700X powder diffractometer, the specific parameters are as follows:

the HPLC spectra were determined using an Agilent model 1260DAD, Agilent, Ach.

Instrument model of infrared determination: fourier transform infrared spectrometer Spectrum Two

In the present application, ubenimex HPLC purity was performed as follows:

(1) a chromatographic column (5 μm particle diameter, column temperature 25 ℃) using octadecylsilane chemically bonded silica as a filler;

(2) mobile phase A: diluted 0.1mol/L potassium dihydrogen phosphate solution (13 → 20) and acetonitrile mixture (V/V ═ 17/3);

mobile phase B: a mixture of acetonitrile and diluted 0.1mol/L potassium dihydrogen phosphate solution (13 → 20) (V/V. 2/1);

elution gradients for mobile phases a and B are shown in the table below:

time (min)	Mobile phase A (%, V/V)	Mobile phase B (%, V/V)
			0-20	100	0
20-60	100→0	0→100
			60-70	0	100
70-71	0→100	100→0
			71-80	100	0

(3) A detector: ultraviolet absorption photometer (wavelength: 220 nm);

(4) flow rate: adjusting the flow rate to ensure that the retention time of the ubenimex is about 14 minutes;

(5) operating time: starting timing from the solvent peak, the retention time of ubenimex is about 5 times;

(6) test solution: taking 30mg of the product, precisely weighing, adding diluent to dissolve and dilute into a solution containing about 3mg of the product in each 1ml, and using the solution as a test solution;

(7) sample introduction amount: and (3) measuring the sample solution by an automatic integration method, and calculating the purity of the ubenimex in the sample according to the peak area.

The specific implementation mode is as follows:

example 1: preparation of ubenimex gamma crystal form

Dispersing 19.40g ubenimex in 195ml of purified water, stirring and pulping at 45 ℃ for 3h, stirring at a speed of 360r/min, cooling to 5 ℃ in an ice bath, and stirring and crystallizing for 1 h; and (3) performing suction filtration under negative pressure, and drying the filter cake in a vacuum oven at 70 ℃ for 8 hours to obtain 17.90g of white solid powder with the yield of 92.3%. HPLC purity 99.96%, max monohetero 0.02%. X-ray powder measurements were performed using Cu-ka radiation, the spectra having diffraction angles and interplanar spacings as shown in the following table:

the nuclear magnetic results were as follows:

¹H NMR(600MHz，DMSO)：δ8.08(d,1H，J＝7.8Hz)，7.32(m,5H),3.96(q,1H，J＝7.2Hz),3.87(d,1H，J＝3.0Hz),3.54(m,1H),2.96(m,2H),1.61(d,2H，J＝4.2Hz),0.86(m,6H)

¹³C NMR(600MHz,DMSO)：δ175.52(s,1C),172.09(s,1C),137.91(s,1C),129.92(s,2C),128.84(s,2C),126.94(s,1C),69.20(s,1C),55.76(s,1C),53.41(s,1C),40.70(s,1C),36.12(s,1C),25.18(s,1C)，23.72(s,1C),21.84(s,1C)

the infrared spectrum is shown in figure 3.

Example 2: preparation of ubenimex gamma crystal form

Dispersing 10.0g ubenimex in 150ml purified water, stirring and pulping at 40 ℃ for 16h, stirring at the speed of 500r/min, cooling to 10 ℃ in an ice bath, and stirring and crystallizing for 1 h; and (3) performing suction filtration under negative pressure, and drying the filter cake in a vacuum oven at 50 ℃ for 16h to obtain 9.07g of white solid powder with the yield of 90.07%. HPLC purity 99.94%, max mono-heterozygote 0.03%, total heterozygote 0.06%.