Preparation method of entecavir EP impurity D
The preparation process of entecavir EP impurity D was simplified by using the Mitsunobu reaction and alkaline hydrolysis steps, which solved the problems of numerous steps and complex operations in the existing technology, and obtained high-purity entecavir EP impurity D, which is suitable for quality research of entecavir.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-08-16
- Publication Date
- 2026-03-03
AI Technical Summary
Existing methods for preparing entecavir EP impurity D involve numerous steps, complex operations, high equipment requirements, difficult purification, and low overall yield.
By employing the Mitsunobu reaction and hydrolysis under alkaline conditions, and using specific organic solvents and reagents, the synthetic route was simplified, reaction conditions were optimized, and purity was improved.
A concise synthetic route was achieved, which is simple to operate, has mild reaction conditions, and produces a high-purity product, making it suitable for entecavir quality studies.
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Figure CN121591735A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to a method for preparing entecavir EP impurity D. Background Technology
[0002] Entecavir is a new generation nucleoside analog HBV drug developed by BMS in the 1990s. It is a 2'-pentanedeoxyguanine nucleoside analog and was approved by the US FDA for the treatment of hepatitis B in 2005, the same year it was approved for marketing in China. Entecavir is characterized by its strong antiviral activity and low drug resistance rate, and is currently the most widely used first-line drug in the clinical treatment of hepatitis B.
[0003] Impurity research is a key research area in drug development and commercial production, and it is conducted throughout the entire drug lifecycle. The presence of impurities directly affects the safety, efficacy, and quality control of drugs.
[0004] Entecavir EP impurity D, as a specified impurity in the European Pharmacopoeia, is essential in the process of entecavir quality and impurity research. Therefore, the study of the synthesis of entecavir EP impurity D has very important practical significance.
[0005] Currently, only one paper reports a method for preparing entecavir EP impurity D. The reported route is as follows, consisting of 10 steps. The route is complex, requires sophisticated equipment (using a reaction at -78 degrees Celsius), is difficult to purify, and has a very low overall yield.
[0006] This invention addresses the shortcomings of existing technologies by providing a method for preparing entecavir EP impurity D. The synthetic route of this invention is short, the operation is simple, the reaction conditions are mild, and the product purity is high. Summary of the Invention
[0007] The purpose of this invention is to provide a method for preparing entecavir EP impurity D.
[0008] This invention provides entecavir EP impurity D (compound of formula IV), the structural formula of which is as follows:
[0009] This invention also provides a method for preparing entecavir EP impurity D (compound of formula IV), comprising the following steps: (1) Compound I reacts with compound II in an organic solvent via the Mitsunobu reaction to prepare compound III. (2) The compound of formula III was hydrolyzed under alkaline conditions to prepare entecavir EP impurity D (compound of formula IV).
[0010] The organic solvent in step (1) is selected from anhydrous dichloromethane and anhydrous tetrahydrofuran. The volume of the organic solvent is 5-50 times (mL / g) the mass of the compound of formula I, preferably 20 times.
[0011] In step (1), the Mitsunobu reagent is selected from a combination of diethyl azodicarbonate, diisopropyl azodicarbonate, N,N,N',N'-tetramethylazodicarbonamide, triphenylphosphine, and tributylphosphine. The molar ratio of diethyl azodicarbonate, diisopropyl azodicarbonate, N,N,N',N'-tetramethylazodicarbonamide to the compound of formula I is 2 to 6:1, preferably 4:1. The molar ratio of triphenylphosphine, tributylphosphine to the compound of formula I is 2 to 6:1, preferably 4:1.
[0012] The reaction temperature in step (1) is -10 degrees to 50 degrees, preferably 25 degrees.
[0013] In step (1), the molar ratio of compound II to compound I is 2 to 4:1, preferably 2.5:1.
[0014] The reaction time for step (1) is 2-48 hours, preferably 20 hours.
[0015] The alkali in step (2) is sodium hydroxide, lithium hydroxide, potassium hydroxide, sodium amino acid, or potassium carbonate, preferably sodium hydroxide.
[0016] In step (2), the molar ratio of the base to the compound of formula III is 2 to 10:1, preferably 2:1.
[0017] In step (2), the reaction temperature is 0 degrees to 65 degrees, preferably 25 degrees.
[0018] The reaction time for step (2) is 5-24 hours, preferably 5 hours.
[0019] Due to the application of the above technical solutions, this invention has the following advantages: a short synthetic route, simple operation, mild reaction conditions, and high product purity. The entecavir EP impurity D (compound of formula IV) prepared by this invention can be used as a reference standard for entecavir-related substances, providing a foundation for quality research on entecavir. Attached Figure Description
[0020] Figure 1 This is the high performance liquid chromatogram of entecavir EP impurity D (formula IV compound) in step (2) of Example 1.
[0021] Figure 2This is the mass spectrometry of entecavir EP impurity D (formula IV compound) in step (2) of Example 1.
[0022] Figure 3 The 1H NMR spectrum of entecavir EP impurity D (formula IV compound) in step (2) of Example 1 is shown. Detailed Implementation
[0023] It should be understood that those skilled in the art can make various modifications and improvements to the invention without departing from the spirit and scope of the invention, based on the content disclosed herein. All such modifications and improvements should fall within the patent protection scope defined by the claims of this application. Furthermore, it is understood that the embodiments provided herein are for illustrative purposes only and should not be construed as limiting the invention.
[0024] The present invention will be further described in detail below with reference to specific embodiments.
[0025] Example 1: (1) Preparation of compound III. 5 g of entecavir (compound I), 7.84 g of p-nitrobenzoic acid (compound II), 18.92 g of triphenylphosphine, and 100 ml of tetrahydrofuran were added to a three-necked flask. Under nitrogen protection and ice bath conditions, 11.36 ml of diethyl azodicarbonate (DEAD) was added. After the addition was complete, the reaction was carried out at 25°C for 20 h. The reaction was monitored by TLC until complete. Water was added to quench the reaction mixture, followed by extraction with ethyl acetate. The organic layer was evaporated to dryness, and purified by column chromatography to obtain 5 g of compound III as a white solid. (2) Preparation of entecavir EP impurity D (compound of formula IV): 5 g of compound of formula III was dissolved in 25 ml of methanol and 25 ml of water, and 0.7 g of sodium hydroxide was added. The reaction was carried out at room temperature for 5 h. After the reaction was completed by TLC monitoring, the pH of the reaction solution was adjusted to neutral, the reaction solution was evaporated to dryness, and 1.2 g of compound of formula IV (i.e., entecavir EP impurity D) was obtained by preparative liquid chromatography as a white solid. Characterization data are as follows: Figure 1 High-performance liquid chromatography purity: 98.97%, chromatogram 2 (mass spectrometry) [M+H] + It is 278.1250. Figure 3 1H NMR spectrum (400MHz, DMSO-d6).
[0026] Example 2: (1) Preparation of compound III. 5 g of entecavir (compound I), 7.84 g of p-nitrobenzoic acid (compound II), 18.92 g of triphenylphosphine, and 100 ml of tetrahydrofuran were added to a three-necked flask. Under nitrogen protection and ice bath conditions, 14.15 ml of diisopropyl azodicarbonate (DIAD) was added. After the addition was complete, the reaction was carried out at 25°C for 20 h. The reaction was monitored by TLC until complete. Water was added to quench the reaction mixture, followed by extraction with ethyl acetate. The organic layer was evaporated to dryness, and purified by column chromatography to obtain 4.5 g of compound III as a white solid. (2) Preparation of entecavir EP impurity D (compound of formula IV): 4.5 g of compound of formula III was dissolved in 25 ml of methanol and 25 ml of water, and 4.14 g of sodium carbonate was added. The mixture was heated to 65 degrees Celsius and reacted for 5 h. After the reaction was completed by TLC monitoring, the pH of the reaction solution was adjusted to neutral, the reaction solution was evaporated to dryness, and 0.9 g of compound of formula IV (i.e., entecavir EP impurity D) was obtained by preparative liquid chromatography purification. The solid was white and the HPLC purity was 97.82%.
[0027] Example 3: (1) Preparation of compound III. 5 g of entecavir (compound I), 7.84 g of p-nitrobenzoic acid (compound II), 18.01 ml of tributylphosphine, and 100 ml of tetrahydrofuran were added to a three-necked flask. Under nitrogen protection and ice bath conditions, 12.42 g of N,N,N',N'-tetramethylazodicarbonamide (TMAD) was added. After the addition was complete, the reaction was carried out at 25°C for 20 h. The reaction was monitored by TLC until complete. Water was added to quench the reaction mixture, followed by extraction with ethyl acetate. The organic layer was evaporated to dryness and purified by column chromatography to obtain 4.8 g of compound III as a white solid. (2) Preparation of entecavir EP impurity D (compound of formula IV): 4.8 g of compound of formula III was dissolved in 25 ml of methanol and 25 ml of water, and 1.75 g of lithium hydroxide monohydrate was added. The mixture was heated to 50 degrees Celsius and reacted for 5 h. After the reaction was completed by TLC monitoring, the pH of the reaction solution was adjusted to neutral, the reaction solution was evaporated to dryness, and 1.1 g of compound of formula IV (i.e., entecavir EP impurity D) was obtained by preparative liquid chromatography purification, which was a white solid.
Claims
1. The structural formula of an entecavir EP impurity D (compound of formula IV) is as follows:
2. A method for preparing entecavir EP impurity D (compound of formula IV), characterized in that: The method includes the following steps: (1) Compound I reacts with compound II in an organic solvent via the Mitsunobu reaction to prepare compound III. (2) The compound of formula III was hydrolyzed under alkaline conditions to prepare entecavir EP impurity D (compound of formula IV).
3. The method as described in claim 2, characterized in that: The organic solvent in step (1) is selected from anhydrous dichloromethane and anhydrous tetrahydrofuran. The volume of the organic solvent is 5-50 times (mL / g) the mass of the compound of formula I, preferably 20 times.
4. The method as described in claim 2, characterized in that: The Mitsunobu reagent in step (1) is selected from a combination of diethyl azodicarbonate, diisopropyl azodicarbonate, N,N,N',N'-tetramethylazodicarbonamide, triphenylphosphine, and tributylphosphine. The molar ratio of diethyl azodicarbonate, diisopropyl azodicarbonate, N,N,N',N'-tetramethylazodicarbonamide to the compound of formula I is 2 to 6:1, preferably 4:
1. The molar ratio of triphenylphosphine, tributylphosphine to the compound of formula I is 2 to 6:1, preferably 4:
1.
5. The method as described in claim 2, characterized in that: The reaction temperature in step (1) is -10 degrees to 50 degrees, preferably 25 degrees.
6. The method as described in claim 2, characterized in that: In step (1), the molar ratio of compound II to compound I is 2 to 4:1, preferably 2.5:
1.
7. The method as described in claim 2, characterized in that: The reaction time for step (1) is 2-48 hours, preferably 20 hours.
8. The method as described in claim 2, characterized in that: The alkali in step (2) is sodium hydroxide, lithium hydroxide, potassium hydroxide, sodium amino acid, or potassium carbonate, preferably sodium hydroxide.
9. The method as described in claim 2, characterized in that: In step (2), the molar ratio of the base to the compound of formula III is 2 to 10:1, preferably 2:
1.
10. The method as described in claim 2, characterized in that: The reaction temperature in step (2) is 0 degrees to 65 degrees, preferably 25 degrees.
11. The method as described in claim 2, characterized in that: The reaction time for step (2) is 5-24 hours, preferably 5 hours.