Preparation method of ritonavir degradation impurity hydantoin amino alcohol

Abstract

The invention relates to a method for preparing impurity hydantoin amino alcohol by high-temperature degradation of ritonavir, which comprises the following steps of: heating and damaging a ritonavir solid, and separating and purifying by preparative chromatography to obtain the impurity hydantoin amino alcohol. The method is simple to operate, efficient and low in cost, and the prepared hydantoamino alcohol is high in yield and purity and can be used as a standard substance for quantitative research of impurities, so that the safety and effectiveness of a ritonavir preparation are ensured.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and in particular relates to a method for preparing ritonavir high-temperature degradation impurity hydantoin. Background Art

[0002] In December 2021, the U.S. Food and Drug Administration approved Pfizer's emergency use authorization application for Paxlovid, a new oral antiviral drug for the treatment of COVID-19. In February 2022, the National Medical Products Administration conditionally approved the import registration of Paxlovid for use in adult patients with mild to moderate COVID-19 who are at high risk of progression to severe illness.

[0003] The simplified chemical structure of ritonavir is: thiazol-5-ylmethyl[(1S,2R,4S)-1-benzyl-2-hydroxy-4-[[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol-4-yl]methyl]carbamoyl]amino]butyryl]amino]-5-phenylpentyl]carbamate, see Formula 1.

[0004]

[0005] Formula 1 Ritonavir structure In the antiviral process, it slows down the CYP3A-mediated metabolism of namatevir and increases its therapeutic concentration. It is an antiviral enhancer and has been approved for use in combination with namatevir to achieve an antiviral effect.

[0006] Hydantoin amino alcohol is included in USP2022 as a known impurity of ritonavir API and ritonavir tablets. Its production mechanism is that the amide bond of ritonavir breaks under high temperature environment and forms a ring with the active hydrogen on the amino group. The reaction formula is shown in Formula 2.

[0007]

[0008] Formula 2: Hydantoin amino alcohol reaction formula In order to better control the quality of ritonavir tablets, improve the qualitative analysis and quantitative research of products, to ensure drug safety, it is necessary to study the impurity hyaluronic acid in ritonavir tablets, and control its content in the final product, therefore, it is necessary to obtain high-purity hyaluronic acid impurity standards, to improve the quality control level of ritonavir products. Up to now, only one document has been reported on the impurity at home and abroad, and the document is to destroy the impurity by refluxing the ritonavir aqueous solution in a water bath, not only complicated operation, but also low recovery rate, only 1%, and in this patent, only the impurity can be obtained by heating the ritonavir solid, and the recovery rate can reach 35%. Using this patented method to prepare hyaluronic acid, not only simple to operate, convenient, and high impurity recovery rate, high purity, can be used as a standard substance for quantitative research of impurities. Summary of the invention

[0009] The technical problem solved by the present invention is to provide a preparation method of ritonavir high temperature degradation impurity hyaluronic acid amino alcohol, and provide an impurity reference substance that meets the requirements for the quality control of ritonavir. The method is simple to operate, efficient, low in cost, and the prepared hyaluronic acid amino alcohol has high yield and high purity.

[0010] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions.

[0011] In one embodiment, the method for preparing ritonavir degradation impurity hydantoin amino alcohol of the present invention is characterized in that: the ritonavir solid is destroyed by heating and then separated and purified by preparative chromatography to obtain it.

[0012] In the preparation method of the present invention, the heating temperature is 100-200°C and the reaction time is 5-20h; preferably, the heating temperature is 150°C and the reaction time is 10-17h.

[0013] Furthermore, in the preparation method of the present invention, water and an organic solvent are added to dissolve and destroy the sample before the chromatographic separation and purification; Wherein, the organic solvent is methanol, acetonitrile or ethanol, preferably acetonitrile.

[0014] The preparation method of the present invention further comprises collecting the target fraction of the preparation chromatography, removing the organic phase under nitrogen, and freeze-drying, namely the impurity hydantoin.

[0015] In the preparation method of the present invention, the preparative chromatography column is an octadecylsilane bonded silica gel column, the mobile phase A is purified water, the mobile phase B is 90% organic phase, the mobile phase flow rate is 27ml / min~33ml / min, and the detection wavelength is 200~250nm.

[0016] Preferably, in the preparation method of the present invention, the chromatographic column model is Agilent SB-C18, 250mm×21.2mm, 7μm; the organic phase is selected from methanol, acetonitrile or a mixture of the two, preferably acetonitrile; the mobile phase flow rate is 30ml / min, and the detection wavelength is 240nm.

[0017] Beneficial technical effects of the present invention: (1) Simple operation and high reaction efficiency. The high-temperature degradation impurity of ritonavir, hydantoin, can be obtained by simply heating the sample to destroy and degrade it.

[0018] (2) Easy purification and high purity. The sample solution can reach 100% purity after chromatographic preparation, freeze-drying and other operations. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is a purity detection chromatogram of hydantoin prepared in Example 3 of the present application; Figure 2 This is the mass spectrum of hydantoin amino alcohol prepared in Example 4 of the present application; Figure 3 is the hydrogen spectrum of hydantoin amino alcohol prepared in Example 4 of the present application; Figure 4 This is the carbon spectrum of hydantoin prepared in Example 4 of the present application. Implementation

[0020] The following examples are used to further illustrate and understand the spirit of the present invention, but are not intended to limit the scope of protection of the present invention in any way.

[0021] Example 1 Preparation of hydantoin amino alcohol Take an appropriate amount of ritonavir, place it in a muffle furnace, and heat it at 150° C. for 17 h to thermally destroy the ritonavir sample. Run it according to the chromatographic conditions in Table 1 to monitor the reaction.

[0022] Table 1 Reaction monitoring chromatographic conditions

[0023] After heat destruction, the content of hydantoin can reach 35%. The sample can be taken for preparative separation.

[0024] Comparative Example 1 Preparation of hydantoin Take an appropriate amount of ritonavir and dissolve it completely in 0.01M ammonium acetate-acetonitrile (1:1) solution according to the conditions in the literature. Then place it in a water bath reflux device and reflux it in a water bath at 70°C for 5 hours to hydrolyze the ritonavir and destroy the sample. Run it according to the chromatographic conditions in Table 1 and monitor the reaction.

[0025] After heat destruction, the content of hydantoin is only 0.9%.

[0026] Example 2 Preparation of hydantoin amino alcohol Take an appropriate amount of ritonavir, place it in a muffle furnace, and heat it at 150° C. for 10 h to thermally destroy the ritonavir sample. Run it according to the chromatographic conditions in Table 1 to monitor the reaction.

[0027] After thermal destruction, the content of hydantoin can reach 22.1%. The sample can be taken for preparative separation.

[0028] Comparative Example 2 Preparation of hydantoin Take an appropriate amount of ritonavir and dissolve it completely in 0.01M ammonium acetate-acetonitrile (1:1) solution according to the conditions in the literature. Then place it in a water bath reflux device and reflux it in a water bath at 70°C for 10 hours to hydrolyze the ritonavir and destroy the sample. Run it according to the chromatographic conditions in Table 1 and monitor the reaction.

[0029] After heat destruction, the content of hydantoin is only 2.7%.

[0030] Example 3 Separation and purification of hydantoin Take an appropriate amount of ritonavir heat-destroyed sample, dissolve the sample with an appropriate amount of acetonitrile water, filter it through a 0.45 μm filter membrane, run it according to the chromatographic conditions in Table 2 below, and receive the target fraction.

[0031] Table 2 Preparative chromatographic conditions

[0032] After the target fraction is collected, the purity of the target fraction is tested using the ritonavir tablets related substance analysis method specified in USP2022. Figure 1 , the purity reaches 100%.

[0033] All the fractions were mixed, the organic phase was removed under nitrogen, and the above solution was freeze-dried in a freeze dryer to obtain the impurity. The mass spectrum is shown in FIG. Figure 2 , mass-to-charge ratio is 551.3, molecular formula is C 29 H 34 N4O5S.

[0034] Example 4 Separation and purification of hydantoin Take an appropriate amount of ritonavir heat-destroyed sample, dissolve the sample with an appropriate amount of acetonitrile water, filter it through a 0.45 μm filter membrane, run it according to the chromatographic conditions in the table below, and receive the target fraction.

[0035] Table 3 Preparative chromatographic conditions

[0036] After the target fraction is collected, the purity of the target fraction is tested using the ritonavir tablet related substance analysis method specified in USP2022.

[0037] All the fractions were mixed, the organic phase was removed under nitrogen, and the above solution was freeze-dried in a freeze dryer to obtain a chromatographic purity of 96.8%. 1H-NMR showed Figure 3 , 13C-NMR see Figure 4 .

Claims

1. A method for preparing ritonavir-degraded impurity hydantoin, characterized in that: The ritonavir solid is destroyed by heating and separated and purified by preparative chromatography to obtain the product.

2. According to the preparation method according to claim 1, the heating temperature is 100-200°C and the reaction time is 5-20h.

3. The preparation method according to claim 2, wherein the heating temperature is 150°C and the reaction time is 10 to 17 hours.

4. The preparation method according to claim 1, wherein water and an organic solvent are added to dissolve and destroy the sample before separation and purification by preparative chromatography.

5. The preparation method according to claim 4, wherein the organic solvent is methanol, acetonitrile or ethanol, preferably acetonitrile.

6. The preparation method according to claim 1 to 5, further comprising collecting the target fraction of the preparative chromatography, removing the organic phase under nitrogen, and freeze-drying to obtain the impurity hydantoin.

7. The preparation method according to claim 1, wherein the preparative chromatography, the chromatographic column is an octadecylsilane bonded silica chromatographic column, the mobile phase A is purified water, the mobile phase B is 90% organic phase, the mobile phase flow rate is 27ml / min~33ml / min, and the detection wavelength is 200~250nm.

8. The preparation method according to claim 7, wherein the chromatographic column model is Agilent SB-C18, 250 mm×21.2 mm, 7 μm.

9. The preparation method according to claim 7, wherein the mobile phase flow rate is 30 ml / min and the detection wavelength is 240 nm.

10. The preparation method according to claim 7, wherein the organic phase is selected from methanol, acetonitrile or a mixture of the two, preferably acetonitrile.

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