Synthetic process for preparing rifampin using rifamycin O as the starting material

Abstract

The application relates to the technical field of organic synthesis, and relates to a synthetic process of rifampicin by taking rifamycin O as a starting material, which comprises the following steps: after rifamycin O is hydrolyzed, a cyclization reaction is carried out on the hydrolyzed rifamycin O and dimethylol tertiary butylamine, and crystallization is carried out, so that rifamycin oxazine is obtained; a condensation reaction is carried out on the rifamycin oxazine and 1-methyl-4-aminopiperazine, and crystallization is carried out, so that rifampicin is obtained. Through the above technical scheme, the problems of low raw material output, insufficient purity and complex preparation in the production of rifampicin in the prior art are solved.

Description

Technical Field

[0001] This invention relates to the field of rifampicin synthesis technology, specifically to a synthesis process for preparing rifampicin using rifamycin O as a starting material. Background Technology

[0002] Rifampin, chemically known as 3-[[(4-methyl-1-piperazinyl)imino]methyl]-rifamycin, is an organic compound and a member of the rifamycin family of antibiotics. It is a low-toxicity, highly effective, broad-spectrum antibiotic with strong antibacterial activity against Mycobacterium tuberculosis, and is also effective against Gram-positive and Gram-negative bacteria and viruses. It is primarily used to treat tuberculosis, meningitis, and Staphylococcus aureus infections, and can be used topically to treat trachoma. Currently, industrial production uses rifamycin S sodium salt or rifamycin S as raw materials and employs an oxazine process to manufacture rifampin. Rifamycin S sodium salt is produced through processes such as fermentation, filtration, oxidation, extraction, washing, and crystallization; rifamycin S is produced by acidifying, purifying, and drying rifamycin S sodium salt.

[0003] The preparation process of rifamycin S sodium salt or rifamycin S is essentially a long-chain production process of "microbial fermentation - multi-step extraction and purification," with each step having technical barriers and efficiency losses. Specifically, the fermentation process is difficult due to the dependence on and controllability of the strain: it requires a specific "Mediterranean amylopectin bacterium rifamycin SV strain" as the starting strain. The fermentation process of this strain has strict requirements on the environment (temperature, pH, dissolved oxygen) and culture medium composition (carbon and nitrogen source ratio, trace elements). Once the fermentation conditions fluctuate, it can easily lead to a decrease in the yield of rifamycin SV or an increase in impurities, directly affecting the output and purity of subsequent raw materials.

[0004] The extraction and purification processes involve multiple losses: extracting rifamycin S sodium salt from fermentation broth requires multiple steps, including plate and frame filtration, oxidation, extraction, washing, and crystallization, each of which results in material loss. Furthermore, rifamycin S requires further "acidification-purification" from rifamycin S sodium salt, increasing the consumption of acid and alkali reagents, the investment in purification equipment, and the processing time. The acidification process may also introduce new impurities, necessitating additional purification steps and further increasing the complexity of raw material preparation. Therefore, a novel synthetic process for rifampicin is needed. Summary of the Invention

[0005] This invention proposes a synthetic process for preparing rifampin using rifamycin O as the starting material, which solves the problems of low raw material yield, insufficient purity, and complex preparation in related technologies.

[0006] The technical solution of the present invention is as follows: This invention proposes a synthetic process for preparing rifampin using rifamycin O as a starting material, comprising the following steps: S1. After hydrolyzing rifamycin O, it undergoes a cyclization reaction with dimethylol tert-butylamine to obtain a cyclized liquid, which is then crystallized to obtain rifamycin oxazine. S2. The rifamycin oxazine is subjected to a condensation reaction with 1-methyl-4-aminopiperazine to obtain a condensation solution, which is then crystallized to obtain rifampin.

[0007] As a further technical solution, the mass-to-volume ratio of rifamycin O and dimethylol tert-butylamine is 20g:5~10mL; Preferably, the mass-to-volume ratio of rifamycin O and dimethylol tert-butylamine is 20 g: 7.14~8 mL.

[0008] As a further technical solution, the mass-to-volume ratio of rifamycin oxazine and 1-methyl-4-aminopiperazine is 50~53g:5mL; Preferably, the mass-to-volume ratio of rifamycin oxazine and 1-methyl-4-aminopiperazine is 51.34~52.57 g: 5 mL.

[0009] As a further technical solution, the hydrolysis is carried out in an acidic solution.

[0010] As a further technical solution, the acidic solution comprises concentrated hydrochloric acid and a solvent.

[0011] As a further technical solution, the solvent includes dimethylformamide.

[0012] As a further technical solution, the hydrolysis reaction temperature is ≥20℃; Preferably, the hydrolysis reaction temperature is 25~30℃ and the time is 6~8h.

[0013] As a further technical solution, the mass-to-volume ratio of rifamycin O, solvent, and concentrated hydrochloric acid is 20g:24~40mL:4~4.5mL.

[0014] As a further technical solution, the cyclization reaction is carried out at a temperature of 50~55℃ for a time of 145~155 min.

[0015] As a further technical solution, ethyl acetate and glacial acetic acid were also added during the cyclization reaction.

[0016] As a further technical solution, the volume ratio of ethyl acetate, glacial acetic acid, and dimethylol tert-butylamine is 8:3.5:5~10.

[0017] As a further technical solution, in step S1, the crystallization is water crystallization.

[0018] As a further technical solution, the water crystallization process includes the following steps: adding the cyclized liquid to water, stirring, letting stand, and filtering to obtain rifamycin oxazine.

[0019] As a further technical solution, during the water crystallization process, the stirring time is 5-10 minutes and the rotation speed is 290 r / min; The settling time is 30-40 minutes.

[0020] As a further technical solution, the temperature of the condensation reaction is 60~65℃ and the time is 70~75min.

[0021] As a further technical solution, the condensation reaction includes the following steps: under stirring, the rifamycin oxazine and reaction aid are added to the solvent, heated to 68~72°C for the first reaction, cooled to 65°C, and then 1-methyl-4-aminopiperazine is added for the second reaction to obtain the condensation solution.

[0022] As a further technical solution, the solvent in the condensation reaction is n-butanol; The temperature of the solvent is 25~30℃; The reaction aids are sodium carbonate, vitamin C, and urea; The mass-to-volume ratio of rifamycin oxazine, sodium carbonate, vitamin C, urea, n-butanol, and 1-methyl-4-aminopiperazine is 51.34~52.57 g:1 g:0.6 g:6 g:54 mL:5 mL; The duration of the first reaction is 90-95 minutes; The second reaction takes 70-75 minutes.

[0023] As a further technical solution, in step S2, the crystallization is cooling crystallization.

[0024] As a further technical solution, the cooling crystallization includes the following steps: mixing water and glacial acetic acid, adding the filtered condensation solution, reacting at 32~40℃, cooling to below 30℃, refrigerating, filtration, and rinsing.

[0025] As a further technical solution, during the cooling crystallization, the volume ratio of water to glacial acetic acid is 100:6; The mixing temperature is 22~30℃; The reaction time is 1~1.5h; The refrigeration temperature was 5°C, and the time was 14 hours.

[0026] As a further technical solution, step S2 includes a refining step after crystallization.

[0027] As a further technical solution, the refining process includes the following steps: Vitamin C and the crystalline product were added to a solvent, dissolved, crystallized, filtered, and dried to obtain rifampin.

[0028] As a further technical solution, during the purification process, the mass-to-volume ratio of the crystallized product, vitamin C, and solvent is 18.53~18.82g:0.2g:60mL; The temperature of the solvent is 70~76℃; The stirring speed is 15~25 r / min; The crystallization temperature is ≤8℃ and the time is 2~6h; The drying temperature is 70~80℃, and the time is 5 hours.

[0029] The working principle and beneficial effects of this invention are as follows: In this invention, rifamycin O has mature industrial production technology and higher production capacity; when used as a starting material, the raw materials are readily available and inexpensive, and large-scale procurement can further reduce raw material costs and enhance price competitiveness, giving it a low-cost advantage; moreover, it does not rely on special scarce raw materials, reducing the risk of raw material supply disruptions during production and ensuring process continuity. Replacing rifamycin S sodium salt / rifamycin S with rifamycin O can effectively reduce the number of production process steps without requiring changes to existing equipment; In the chemical structure of rifamycin O, the functional groups such as phenolic hydroxyl and quinone groups are more stable than some homologues, which makes the physicochemical properties stable and reduces the generation of impurities during storage, transportation and reaction, indirectly improving the purity of the final product rifampin and reducing the difficulty of quality control. Because rifamycin O material is stable and has fewer production steps, the batch-to-batch content fluctuations will be greatly reduced, significantly improving production consistency and meeting drug quality standards. Attached Figure Description

[0030] The present invention will now be described in further detail with reference to the accompanying drawings and specific embodiments.

[0031] Figure 1 This is a physical image of the rifampicin product from Embodiment 1 of the present invention; Figure 2 This is a high-performance liquid chromatogram of the rifampicin product from Example 1 of the present invention. Detailed Implementation

[0032] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0033] In the following examples and comparative examples, rifamycin O was manufactured by Hisun Pharmaceutical (Hangzhou) Co., Ltd.; dimethylol tert-butylamine was manufactured by Hebei Xingang Pharmaceutical Co., Ltd.; concentrated hydrochloric acid, dimethylformamide, ethyl acetate, glacial acetic acid, and n-butanol were all of analytical grade.

[0034] Example 1 The synthetic process for preparing rifampin using rifamycin O as the starting material includes the following steps: Add 30 mL of dimethylformamide and 4.0 mL of concentrated hydrochloric acid to a 500 mL three-necked flask, heat to 30 °C, then add 20 g of rifamycin O, stir at 264 r / min for 6 h, and test the reaction endpoint by thin-layer chromatography to obtain solution A; Add 8 mL of ethyl acetate, 3.5 mL of glacial acetic acid, and 8 mL of dimethylol tert-butylamine to solution A, maintain the temperature at 53 °C, react for 150 min, take a sample, and test the reaction endpoint by thin-layer chromatography to obtain the cyclized solution; Add 300 mL of drinking water to a 500 mL beaker, start stirring, pour in the cyclized solution, stir at 290 r / min for 8 min, let stand for 35 min, filter with a Buchner funnel, and obtain 51.55 g of rifamycin oxazine. Add 54 mL of n-butanol to a 500 mL three-necked flask, stir at 215 r / min and heat to 25 °C, add the rifamycin oxazine obtained above, 1 g of sodium carbonate, 0.6 g of vitamin C and 6 g of urea, heat to 70 °C and keep the reaction at this temperature for 93 min. Use thin-layer chromatography to determine the reaction endpoint; cool to 65 °C, add 5 mL of 1-methyl-4-aminopiperazine, keep the reaction at this temperature for 70 min, use thin-layer chromatography to determine the reaction endpoint, and obtain the condensate; Add 100 mL of purified water and 6 mL of glacial acetic acid to a 500 mL three-necked flask, stir at 264 r / min and heat to 28 °C, add the filtered condensate, continue to heat to 38 °C, stir at 264 r / min for 1 h, then cool to 30 °C and refrigerate at 5 °C for 14 h, filter with a Buchner funnel and wash with purified water and n-butanol to obtain 18.82 g of crude rifampicin wet product; Add 60 mL of n-butanol to a 500 mL three-necked flask, stir at 20 r / min and heat to 73 °C. Quickly add 0.2 g of vitamin C and the above-mentioned crude rifampicin wet product, stir until completely dissolved, stop heating, cool to below 8 °C, and allow crystals to grow for 6 h. Filter using a Buchner funnel. Dry the obtained crude rifampicin wet product under vacuum at 80 °C for 5 h to obtain 14.1 g of rifampicin. Yield = rifampicin yield / mass of added rifamycin O × 100% = 14.1 / 20 × 100% = 70.5%.

[0035] Example 2 The synthetic process for preparing rifampin using rifamycin O as the starting material includes the following steps: Add 40 mL of dimethylformamide and 4.5 mL of concentrated hydrochloric acid to a 500 mL three-necked flask, heat to 25 °C, add 20 g of rifamycin O, stir and react for 8 h, and use thin-layer chromatography to check the reaction endpoint to obtain solution A; Add 8 mL of ethyl acetate, 3.5 mL of glacial acetic acid, and 8 mL of dimethylol tert-butylamine to solution A, maintain the temperature at 50 °C, react for 155 min, take a sample, and use thin-layer chromatography to test the reaction endpoint to obtain the cyclized solution; Add 300 mL of drinking water to a 500 mL beaker, start stirring, pour in the cyclized solution, stir at 290 r / min for 5 min, let stand for 30 min, filter with a Buchner funnel, and obtain 52.57 g of rifamycin oxazine. Add 54 mL of n-butanol to a 500 mL three-necked flask, stir at 215 r / min and heat to 30 °C, add the rifamycin oxazine obtained above, 1 g of sodium carbonate, 0.6 g of vitamin C and 6 g of urea, heat to 68 °C and keep the reaction at this temperature for 95 min. Use thin-layer chromatography to determine the reaction endpoint; cool to 65 °C, add 5 mL of 1-methyl-4-aminopiperazine, keep the reaction at this temperature for 70 min, use thin-layer chromatography to determine the reaction endpoint, and obtain the condensate; Add 100 mL of purified water and 6 mL of glacial acetic acid to a 500 mL three-necked flask, stir at 264 r / min and heat to 22 °C, add the filtered condensate, continue to heat to 32 °C, stir at 264 r / min for 1.5 h, then cool to 30 °C and refrigerate at 5 °C for 14 h, filter with a Buchner funnel and wash with purified water and n-butanol to obtain 18.53 g of crude rifampicin wet product; Add 60 mL of n-butanol to a 500 mL three-necked flask, stir at 15 r / min and heat to 70 °C. Quickly add 0.2 g of vitamin C and the above-mentioned wet crude rifampin. After stirring until completely dissolved, stop heating, cool to below 8 °C, and allow crystals to grow for 2 hours. Filter using a Buchner funnel. Dry the obtained wet crude rifampin at 70 °C under vacuum for 5 hours to obtain 13.6 g of rifampin. Yield = rifampin yield / mass of added rifamycin O × 100% = 13.6 / 20 × 100% = 68%.

[0036] Example 3 The synthetic process for preparing rifampin using rifamycin O as the starting material includes the following steps: Add 24 mL of dimethylformamide and 4.0 mL of concentrated hydrochloric acid to a 500 mL three-necked flask, heat to 30 °C, then add 20 g of rifamycin O, stir and react for 6 h, and use thin-layer chromatography to check the reaction endpoint to obtain solution A; Add 8 mL of ethyl acetate, 3.5 mL of glacial acetic acid, and 7.14 mL of dimethylol tert-butylamine to solution A, maintain the temperature at 55 °C, react for 145 min, take a sample, and use thin-layer chromatography to test the reaction endpoint to obtain the cyclized solution; Add 300 mL of drinking water to a 500 mL beaker, start stirring, pour in the cyclized solution, stir at 290 r / min for 10 min, let stand for 40 min, filter with a Buchner funnel, and obtain 51.34 g of rifamycin oxazine. Add 54 mL of n-butanol to a 500 mL three-necked flask, stir at 215 r / min and heat to 25 °C, add the rifamycin oxazine obtained above, 1 g of sodium carbonate, 0.6 g of vitamin C and 6 g of urea, heat to 72 °C and keep the reaction at this temperature for 90 min. Use thin-layer chromatography to determine the reaction endpoint; cool to 60 °C, add 5 mL of 1-methyl-4-aminopiperazine, keep the reaction at this temperature for 75 min, use thin-layer chromatography to determine the reaction endpoint, and obtain the condensate; Add 100 mL of purified water and 6 mL of glacial acetic acid to a 500 mL three-necked flask, stir at 264 r / min and heat to 30 °C, add the filtered condensate, continue to heat to 40 °C, stir at 264 r / min for 1.3 h, then cool to 29 °C and refrigerate at 5 °C for 14 h, filter with a Buchner funnel and wash with purified water and n-butanol to obtain 18.61 g of wet crude rifampin; Add 60 mL of n-butanol to a 500 mL three-necked flask, stir at 25 r / min and heat to 76 °C. Quickly add 0.2 g of vitamin C and the above-mentioned wet crude rifampin. After stirring until completely dissolved, stop heating, cool to below 8 °C, and allow crystals to grow for 4 hours. Filter using a Buchner funnel. The obtained wet purified rifampin is vacuum dried at 75 °C for 5 hours to obtain 13.85 g of rifampin. Yield = rifampin yield / mass of added rifamycin O × 100% = 13.8 / 20 × 100% = 69%.

[0037] Example 4 The synthetic process for preparing rifampin using rifamycin O as the starting material includes the following steps: Add 30 mL of dimethylformamide and 4.0 mL of concentrated hydrochloric acid to a 500 mL three-necked flask, control the temperature at 15 °C, then add 20 g of rifamycin O, stir the mixture at 264 r / min for 6 h, and test the reaction endpoint by thin-layer chromatography to obtain solution A; Add 8 mL of ethyl acetate, 3.5 mL of glacial acetic acid, and 8 mL of dimethylol tert-butylamine to solution A, maintain the temperature at 53 °C, react for 150 min, take a sample, and test the reaction endpoint by thin-layer chromatography to obtain the cyclized solution; Add 300 mL of drinking water to a 500 mL beaker, start stirring, pour in the cyclized solution, stir at 290 r / min for 8 min, let stand for 35 min, filter with a Buchner funnel, and obtain 46.55 g of rifamycin oxazine. Add 54 mL of n-butanol to a 500 mL three-necked flask, stir at 215 r / min and heat to 25 °C, add the rifamycin oxazine obtained above, 1 g of sodium carbonate, 0.6 g of vitamin C and 6 g of urea, heat to 70 °C and keep the reaction at this temperature for 93 min. Use thin-layer chromatography to determine the reaction endpoint; cool to 65 °C, add 5 mL of 1-methyl-4-aminopiperazine, keep the reaction at this temperature for 70 min, use thin-layer chromatography to determine the reaction endpoint, and obtain the condensate; Add 100 mL of purified water and 6 mL of glacial acetic acid to a 500 mL three-necked flask, stir at 264 r / min and heat to 28 °C, add the filtered condensate, continue to heat to 38 °C, stir at 264 r / min for 1 h, then cool to 30 °C and refrigerate at 5 °C for 14 h, filter with a Buchner funnel and wash with purified water and n-butanol to obtain 13.79 g of crude rifampicin wet product; Add 60 mL of n-butanol to a 500 mL three-necked flask, stir at 20 r / min and heat to 75 °C. Quickly add 0.2 g of vitamin C and the above-mentioned wet crude rifampin. After stirring until completely dissolved, stop heating, cool to below 8 °C, and allow crystals to grow for 6 hours. Filter using a Buchner funnel. Dry the obtained wet crude rifampin at 80 °C under vacuum to obtain 10.24 g of rifampin. Yield = Rifampin yield / Mass of added rifamycin O × 100% = 10.24 / 20 × 100% = 51.2%.

[0038] Experimental Example The rifampicin prepared in Examples 1-3 was tested according to the following method; The content and related substances of rifampicin were characterized and detected in accordance with the Chinese Pharmacopoeia; the content was determined by external standard method and high performance liquid chromatography (General Rule 0512). High performance liquid chromatography (HPLC) conditions: octylsilane-bonded silica gel was used as the stationary phase; the mobile phase consisted of methanol, acetonitrile, potassium dihydrogen phosphate solution (0.075 mol / L) and citric acid solution (1.0 mol / L) in a volume ratio of 30:30:36:4; the injection volume was 10 μL; and the detection wavelength was 254 nm. If impurity peaks are present in the chromatogram of the test solution, the peak areas of quinone rifampin, N-oxyrifampin, and 3-formylrifamycin SV, calculated using the external standard method, shall not exceed 1.5%, 0.5%, and 0.5%, respectively; the area of ​​any other single impurity peak shall not exceed the area of ​​the main peak of the control solution (1.0%), and the sum of the areas of all other impurity peaks shall not exceed three times the area of ​​the main peak of the control solution (3.0%). The test results are shown in Table 1 below: Table 1. High-performance liquid chromatography (HPLC) test results of rifampicin prepared in Examples 1-3

[0039] The test results showed that the content of quinone rifampin was less than 1.5%, the content of N-oxyrifampin was less than 0.5%, the content of 3-formylrifamycin SV was less than 0.5%, and the peak area of ​​other individual impurities was not greater than the main peak area of ​​the control solution (1.0%), which meets the standards of the Chinese Pharmacopoeia. Therefore, this production method is feasible.

[0040] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A synthetic process for preparing rifampin using rifamycin O as a starting material, characterized in that, Includes the following steps: S1. After hydrolyzing rifamycin O, it undergoes a cyclization reaction with dimethylol tert-butylamine to obtain a cyclized liquid, which is then crystallized to obtain rifamycin oxazine. S2. The rifamycin oxazine is subjected to a condensation reaction with 1-methyl-4-aminopiperazine to obtain a condensation solution, which is then crystallized to obtain rifampin.

2. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The mass-to-volume ratio of rifamycin O and dimethylol tert-butylamine is 20 g: 5-10 mL.

3. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The mass-to-volume ratio of rifamycin oxazine and 1-methyl-4-aminopiperazine is 50-53 g: 5 mL.

4. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The hydrolysis is carried out in an acidic solution; The hydrolysis reaction temperature is ≥20℃; Preferably, the hydrolysis reaction temperature is 25~30℃.

5. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The cyclization reaction is carried out at a temperature of 50-55°C for a time of 145-155 min. Ethyl acetate and glacial acetic acid were also added during the cyclization reaction.

6. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, In step S1, the crystallization is water precipitation crystallization.

7. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The condensation reaction is carried out at a temperature of 60-65°C for 70-75 minutes.

8. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, The condensation reaction includes the following steps: under stirring, the rifamycin oxazine and reaction aid are added to the solvent, heated to 68~72°C for the first reaction, cooled to 65°C, and then 1-methyl-4-aminopiperazine is added for the second reaction to obtain the condensation solution.

9. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, In step S2, the crystallization is performed by cooling crystallization.

10. The synthetic process for preparing rifampin using rifamycin O as a starting material according to claim 1, characterized in that, In step S2, the crystallization process further includes a refining step.

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