A method for separating and purifying erythromycin-6,9-imino ether

Abstract

The invention discloses a separation and purification method of erythromycin-6,9-imidoether, and erythromycin-6,9-imidoether is separated and purified by three-stage countercurrent centrifugal extraction technology and composite solvent crystallization technology. Erythromycin oxime salt is freed with a hydrophobic solvent, and extraction and separation are performed by three-stage countercurrent centrifugal extraction technology, that is, the free extraction time is shortened, and erythromycin oxime is prevented from being too long under alkaline conditions, and degradation occurs and affects product quality. Composite solvent crystallization technology is used for purification, that is, the residue of hydrophobic solvent dichloromethane in the reaction solution can be effectively removed, and erythromycin-6,9-imidoether can be directly purified and extracted, while replacing the refining process of ethanol and acetone in the traditional process, so as to achieve the purpose of reducing production costs and improving product yield.

Description

Technical Field

[0001] The invention belongs to the technical field of biopharmaceuticals, and in particular relates to a method for separating and purifying erythromycin-6,9-imidoether. Background Art

[0002] Azithromycin is a 15-membered macrolide antibiotic with a similar antibacterial mechanism to erythromycin, but with a broader antibacterial spectrum. It has strong antibacterial activity against both Gram-positive bacteria and Gram-negative bacteria (such as Haemophilus influenzae, Salmonella, Escherichia coli, Shigella, etc.). It is also stable to acid and has good tolerance. Related literature reports that the preparation method of azithromycin is usually as follows: erythromycin thiocyanate reacts with hydroxylamine hydrochloride to convert into erythromycin oxime salt; erythromycin oxime salt is first freed to erythromycin oxime (conversion rate is about 85%), and then converted into erythromycin-6,9-imido ether by Beckmann rearrangement; erythromycin-6,9-imido ether is hydrogenated and reduced to azithromycin amine, and then further methylated and hydrolyzed to obtain azithromycin.

[0003] Erythromycin 6,9-Imino Ether, also known as erythromycin 6,9-imino ether, azithromycin impurity 17, is easily soluble in water and poorly soluble in hydrophobic solvents (such as ethyl acetate, dichloromethane, chloroform, dichloroethane, toluene, petroleum ether, etc.). The US Chemical Abstracts Registry Number is CAS: 99290-97-8, and its structural formula is:

[0004]

[0005] Erythromycin-6,9-imidoether is an important precursor for the synthesis of azithromycin. Its synthesis route is as follows:

[0006]

[0007] At present, the HPLC purity of erythromycin-6,9-iminoether prepared by ordinary process is about 92%, and the purity of azithromycin synthesized by it is generally about 96%. It is necessary to increase the refining process to make its purity reach more than 98% (USP43 quality standard). Although the refining can also meet the quality standard of azithromycin USP43, it undoubtedly increases manpower, material resources and production time, resulting in a series of problems such as increased product cost and decreased yield. In addition, since the free erythromycin oxime salt is mainly carried out under alkaline conditions, the conventional extraction method takes too long to cause the erythromycin oxime to degrade, affecting the product yield and its quality. Summary of the invention

[0008] The object of the present invention is to provide a method for separating and purifying erythromycin-6,9-imino ether, which optimizes the azithromycin rearrangement reaction process through a three-stage countercurrent centrifugal extraction combined with a composite solvent extraction technology, effectively improves the purity and yield of azithromycin, and reduces production costs. In order to achieve the above object, the technical solution adopted by the present invention is:

[0009] A method for separating and purifying erythromycin-6,9-imino ether, comprising the following specific steps:

[0010] (1) dissolving erythromycin oxime salt in deionized water, adding a hydrophobic solvent, and performing free extraction using a three-stage countercurrent centrifugal extraction technique under alkaline conditions to obtain an erythromycin oxime organic phase;

[0011] (2) adding deionized water, an acid binding agent and a sulfonylating agent to the organic phase of step (1) to carry out a Beckmann rearrangement reaction to obtain an erythromycin-6,9-imidoether mixed phase liquid;

[0012] (3) subjecting the erythromycin-6,9-iminoether mixed phase solution obtained in step (2) to back extraction under acidic conditions to obtain an erythromycin-6,9-iminoether aqueous phase solution;

[0013] (4) adding the aqueous phase solution of erythromycin-6,9-imino ether prepared in step (3) into a composite solvent for dropwise alkali crystallization, and centrifuging to obtain erythromycin-6,9-imino ether.

[0014] In some embodiments of the present invention, the hydrophobic solvent in step (1) is one of ethyl acetate, dichloromethane, chloroform, dichloroethane, toluene, and petroleum ether.

[0015] In some embodiments of the present invention, the alkaline condition in step (1) is 10%-15% liquid alkali for alkali extraction, and the pH value is 10.5-12.5.

[0016] In some embodiments of the present invention, the temperature of the three-stage countercurrent centrifugal extraction in step (1) is 20-25°C.

[0017] The three-stage countercurrent centrifugal extraction technology route is as follows:

[0018]

[0019] In some embodiments of the present invention, the volume ratio of the hydrophobic solvent in step (1): deionized water is 1:2-3.

[0020] Preferably, in step (1), the volume ratio of the hydrophobic solvent to deionized water is 1:2.5.

[0021] In some embodiments of the present invention, the sulfonylating agent in step (2) is p-toluenesulfonyl chloride or benzenesulfonyl chloride.

[0022] In some embodiments of the present invention, the acid binding agent in step (2) is sodium bicarbonate.

[0023] In some embodiments of the present invention, the amount of the acid binding agent (calculated by molar ratio) is 1-2 times of the amount of erythromycin A (the starting reactant for azithromycin synthesis), and the amount of the sulfonylating agent (calculated by molar ratio) is 1-2.5 times of the amount of erythromycin A (the starting reactant for azithromycin synthesis). The amount of the acid binding agent is preferably 1.5 times, and the amount of the sulfonylating agent is preferably 2 times.

[0024] In some embodiments of the present invention, the Beckmann rearrangement reaction temperature in step (2) is 0-5°C, and the reaction time is 2.5-3h, preferably 2.5h.

[0025] In some embodiments of the present invention, the acidic condition in step (3) is 30%-40% glacial acetic acid or formic acid for acid adjustment and stripping, and the pH value is 4.8-6.0.

[0026] In some embodiments of the present invention, the composite solvent in step (4) is one or both of ethyl acetate and toluene.

[0027] Preferably, the composite solvent in step (4) is composed of ethyl acetate-toluene in a volume ratio of 1:2-5.

[0028] In some embodiments of the present invention, the extraction volume ratio of the composite solvent to the aqueous phase liquid in step (4) is 1:2-3 (V:V).

[0029] In some embodiments of the present invention, the concentration of the liquid alkali solution in step (4) is 10%-15% (W / W).

[0030] The technical solution of the present invention has the following beneficial technical effects:

[0031] 1. The three-stage countercurrent centrifugal extraction technology adopted in the present invention not only shortens the extraction cycle, but also prevents erythromycin oxime from being degraded under alkaline conditions for too long, thereby affecting product quality, and significantly improves production efficiency.

[0032] 2. The composite solvent crystallization technology adopted in the present invention can effectively remove the residual hydrophobic solvent dichloromethane in the reaction solution, and can directly purify erythromycin-6,9-imido ether, thereby avoiding the problems of solvent loss and yield reduction caused by the single purification of erythromycin-6,9-imido ether.

[0033] 3. The present invention adopts three-stage countercurrent centrifugal extraction technology and composite solvent crystallization technology to separate and purify erythromycin-6,9-imido ether. The purity of HPLC can reach more than 97%, and the purity of azithromycin synthesized by the method can reach more than 98%. At the same time, it replaces the refining steps of ethanol and acetone in the traditional process, significantly improving the purity and yield of the azithromycin product.

[0034] 4. Based on the above advantages, the present invention makes the process more suitable for industrial production and can effectively achieve the goals of improving quality and reducing costs. DETAILED DESCRIPTION

[0035] In the following embodiment, the preparation process of erythromycin oxime salt is as follows: in a 500ml reaction bottle, 100g methanol, 40g hydroxylamine hydrochloride, 45g sodium acetate, 100g erythromycin thiocyanate are added, the temperature is raised to 40-70°C, the reaction is carried out for 24-72h, and the erythromycin content is less than 0.5% by HPLC monitoring. After the reaction is completed, 100ml deionized water is added dropwise, stirred, and centrifuged to obtain erythromycin oxime salt.

[0036] In the above process, erythromycin oxime salt is prepared by using the preparation process recorded in the current public document (patent CN103319551A).

[0037] Example 1 Three-stage countercurrent extraction technology + single solvent (ethyl acetate) drop alkali crystallization

[0038] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust the pH to 10.5-12.5, and extract the dichloromethane organic phase by three-stage countercurrent centrifugation. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Adjust the pH to 4.8-6.0 with 35% glacial acetic acid, let stand and separate, add 400ml of single solvent ethyl acetate to the water layer, adjust the pH to 10.5-12.5 with 10% sodium hydroxide solution, and centrifuge to obtain 68g of erythromycin-6,9-imido ether (yield is about 80%), with an HPLC purity of 97.1%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 98.2%.

[0039] Example 2 Three-stage countercurrent extraction technology + single solvent (toluene) drop alkali crystallization

[0040] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust the pH to 10.5-12.5, and extract the dichloromethane organic phase by three-stage countercurrent centrifugation. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Adjust the pH to 4.8-6.0 with 35% glacial acetic acid, let stand and separate, add 400ml of single solvent toluene to the water layer, adjust the pH to 10.5-12.5 with 10% sodium hydroxide solution, and centrifuge to obtain 68g of erythromycin-6,9-imido ether (yield is about 80%), with an HPLC purity of 97.3%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 98.5%.

[0041] Example 3 Three-stage countercurrent extraction technology + composite solvent (ethyl acetate-toluene) drop alkali crystallization

[0042] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust pH=10.5-12.5, and extract dichloromethane organic phase by three-stage countercurrent centrifugation. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Adjust pH=4.8-6.0 with 35% glacial acetic acid, let stand and separate, add 400ml of composite solvent ethyl acetate-toluene (80ml of ethyl acetate + 320ml of toluene) to the water layer, adjust pH=10.5-12.5 with 10% sodium hydroxide solution, and centrifuge to obtain 68g of erythromycin-6,9-imido ether (yield is about 80%), with HPLC purity of 97.5%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 98.8%.

[0043] Comparative Example 1 Conventional extraction technology + water phase alkali drop crystallization

[0044] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust pH=10.5-12.5, extract conventionally, stand for stratification, and discard the water layer to obtain the dichloromethane organic phase. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Use 35% glacial acetic acid to adjust pH=4.8-6.0, stand for stratification, add 10% sodium hydroxide solution to the water layer to adjust pH=10.5-12.5, filter and dry to obtain 63g of erythromycin-6,9-imido ether (yield is about 74%), and the HPLC purity is 92.5%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 96.2%.

[0045] Comparative Example 2: Three-stage countercurrent extraction technology + water phase alkali drop crystallization

[0046] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust the pH to 10.5-12.5, and extract the dichloromethane organic phase by three-stage countercurrent centrifugation. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Use 35% glacial acetic acid to adjust the pH to 4.8-6.0, let stand and separate, add 10% sodium hydroxide solution to the water layer to adjust the pH to 10.5-12.5, filter and dry to obtain 68g of erythromycin-6,9-imido ether (yield is about 80%), and the HPLC purity is 94.8%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 96.3%.

[0047] Comparative Example 3 Conventional extraction technology + composite solvent (ethyl acetate-toluene) drop alkali crystallization

[0048] Take 100g of erythromycin oxime salt (the conversion rate of erythromycin oxime is 85%), add 280ml of deionized water, 700ml of dichloromethane, add 10% sodium hydroxide solution, adjust pH=10.5-12.5, extract conventionally, stand for stratification, and discard the water layer to obtain the dichloromethane organic phase. Add 1000ml of deionized water and 14g of sodium bicarbonate to dichloromethane, stir and cool to 0-5°C, add 42g of p-toluenesulfonyl chloride and react for 2.5h. Use 35% glacial acetic acid to adjust pH=4.8-6.0, stand for stratification, add 400ml of composite solvent ethyl acetate-toluene (ethyl acetate 80ml + toluene 320ml) to the water layer, adjust pH=10.5-12.5 with 10% sodium hydroxide solution, and centrifuge to obtain 63g of erythromycin-6,9-imido ether (yield is about 74%), with HPLC purity of 96.5%. The prepared erythromycin-6,9-imino ether is further subjected to potassium borohydride reduction and methylation reaction to obtain azithromycin. The purity of azithromycin detected by USP43 quality standard is 97.8%.

[0049] The above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, ordinary technicians in the field should understand that the specific implementation methods of the present invention can still be modified or some technical features can be replaced by equivalents without departing from the spirit of the technical solution of the present invention, which should be included in the scope of the technical solution for protection of the present invention.

Claims

1. A method for separating and purifying erythromycin-6,9-imino ether, comprising the following steps: (1) dissolving erythromycin oxime salt in deionized water, adding a hydrophobic solvent, and performing free extraction using a three-stage countercurrent centrifugal extraction technique under alkaline conditions to obtain an erythromycin oxime organic phase; (2) adding deionized water, an acid binding agent and a sulfonylating agent to the organic phase of step (1) to carry out a Beckmann rearrangement reaction to obtain an erythromycin-6,9-imidoether mixed phase liquid; (3) subjecting the mixed phase liquid of erythromycin-6,9-imino ether in step (2) to back extraction under acidic conditions to obtain an aqueous phase liquid of erythromycin-6,9-imino ether, wherein: The acidic condition is 30%-40% glacial acetic acid or formic acid for acid adjustment and stripping, and the pH value is 4.8-6.0; (4) adding a solvent to the aqueous phase solution of erythromycin-6,9-imidoether prepared in step (3) to perform alkali dropwise crystallization, and centrifuging to obtain erythromycin-6,9-imidoether, wherein the solvent is one or both of ethyl acetate and toluene.

2. The separation and purification method as claimed in claim 1, characterized in that The hydrophobic solvent in step (1) is one of ethyl acetate, dichloromethane, chloroform, dichloroethane, toluene and petroleum ether.

3. The separation and purification method as claimed in claim 1, characterized in that The alkaline condition in step (1) is 10%-15% liquid alkali for alkali extraction, and the pH value is 10.5-12.

5.

4. The separation and purification method as claimed in claim 1, characterized in that The volume ratio of the hydrophobic solvent in step (1): deionized water is 1:2-3.

5. The separation and purification method as claimed in claim 1, characterized in that The sulfonylating agent in step (2) is p-toluenesulfonyl chloride or benzenesulfonyl chloride.

6. The separation and purification method as claimed in claim 1, characterized in that The acid binding agent in step (2) is sodium bicarbonate.

7. The separation and purification method as claimed in claim 1, characterized in that The solvent in step (4) is composed of ethyl acetate-toluene in a volume ratio of 1:2-5.

8. The separation and purification method as claimed in claim 1, characterized in that In step (4), the extraction volume ratio of the solvent to the aqueous phase liquid is 1:2-3 (V:V).

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