Synthesis method of fusidic acid impurity B

Abstract

The invention relates to the field of organic synthesis, and discloses a synthesis method of an impurity B of fusidic acid. The method comprises the following steps: 1, dissolving fusidic acid in a first organic solvent, adding an epoxidation agent, reacting for a first time at a first temperature, and extracting to obtain a compound I; 2, dissolving the compound I in a second organic solvent, adding a catalyst, reacting for a second time at a second temperature, and pulping and washing by using a third organic solvent to obtain a compound II; in the embodiment of the invention, reagents used in the synthesis route of the fusidic acid impurity B are easy to obtain and low in cost, the whole route only needs two steps, the total yield can reach more than 50%, and the technical problem that the cost is too high when a large amount of fusidic acid impurity B is synthesized in the prior art is solved.

Description

Technical Field

[0001] The present invention relates to the field of organic synthesis, and particularly to a method for synthesizing impurity B of fusidic acid. Background Art

[0002] Fusidic acid, with the English name Fusidic acid and the chemical name (Z)-2-((3R,4S,5S,8S,9S,10S,11R,13R,14S,16S)-16-acetoxy-3,11-dihydroxy-4,8,10,14-tetramethyldodecahydro-1H-cyclopenta[a]phenanthren-17(2H,10H,14H)-ylidene)-6-methylhept-5-enoic acid, contains a steroid nucleus in its molecular structure and has a high similarity to aspergillic acid.

[0003] Fusidic acid is an important antibiotic. Research shows that it is highly sensitive to various Gram-positive cocci related to skin infections, especially to staphylococci, is also effective against drug-resistant Staphylococcus aureus, has a certain antibacterial effect on some Gram-negative bacteria, and there is no cross-resistance with other antibiotics. These characteristics have enabled it to be widely used in the antibacterial field. The common salt form of fusidic acid is its sodium salt, and the commercial dosage forms mainly include ointments, injections, and dry suspensions, etc. These different dosage forms are for treating infection diseases caused by various sensitive bacteria. For example, fusidic acid injection can be used for infections of the limbs and joints, bacterial sepsis, endocarditis, cystic fibrosis, osteomyelitis, skin tissue infections, pneumonia, and some traumatic infections, etc.; the ointment can be used for treating infections caused by staphylococci, streptococci, Corynebacterium minutissimum, and other sensitive bacteria, etc.

[0004] Fusidic acid belongs to steroid compounds, but its three-dimensional structure is different from that of traditional steroid compounds. Due to the complexity of its structure, the chemical synthesis steps are cumbersome and lengthy, and the yield is low, so the cost is extremely high. Currently, almost all fusidic acid on the market is isolated from the secondary metabolites of fermentation of Fusidium coccineum. Inevitably, other substances are mixed in during the fermentation and extraction processes, or partial deterioration occurs during storage. These factors are important sources of impurities in fusidic acid raw materials. In practice, the research and control of impurities in raw materials are related to the clinical safety of drugs, so it has become one of the key links in the quality control of raw materials. The adverse reactions that occur during the clinical use of drugs are often related to some impurities in the drugs in addition to being caused by the pharmacological characteristics of the drugs themselves. Therefore, it is very necessary to conduct in-depth research and strict control on the impurities in drugs.

[0005]

[0006] The impurity shown in formula (II) is included in EP 9.5, with the code name B, and is an isomer of fusidic acid EP impurities C, D, and E. The Journal of China Pharmaceutical University (2018, 49(3): 322) reported the generation conditions of this impurity. Experiments showed that fusidic acid can generate impurity B under both oxidative degradation and photodegradation conditions, but the HPLC content is less than 0.1%. Except for the above literature, there is no other reported synthetic method. However, the cost of synthesizing this impurity B in large quantities by this method is too high, and a new synthetic technology is needed to solve the current problem. Summary of the Invention

[0007] The main object of the present invention is to solve the technical problem that the cost of synthesizing impurity B of fusidic acid in large quantities in the prior art is too high.

[0008] The first aspect of the present invention provides a synthetic method for impurity B of fusidic acid, and the synthetic method for impurity B of fusidic acid includes:

[0009] Step 1: Dissolve fusidic acid in a first organic solvent, add an epoxidizing agent, react at a first temperature for a first duration, and extract to obtain compound I;

[0010] Step 2: Dissolve compound I in a second organic solvent, add a catalyst, react at a second temperature for a second duration, and repulp and wash with a third organic solvent to obtain compound II. The reaction equations for steps 1-2 are as follows:

[0011]

[0012] Optionally, in the first implementation manner of the first aspect of the present invention, the first organic solvent includes: one or a mixture of two or more of dichloromethane, chloroform, 1,2-dichloroethane, acetonitrile, and ethyl acetate.

[0013] Optionally, in the second implementation manner of the first aspect of the present invention, the epoxidizing reagent includes: any one of 3-chloroperoxybenzoic acid, peroxybenzoic acid, peracetic acid, and carboxylic acid derivatives.

[0014] Optionally, in the third implementation manner of the first aspect of the present invention, the molar ratio of fusidic acid to epoxidizing agent is = 1:(1.5-3).

[0015] Optionally, in the fourth implementation manner of the first aspect of the present invention, the first temperature includes: -10°C to 10°C, and the first duration includes: 0.5 h to 1.5 h.

[0016] Optionally, in the fifth implementation manner of the first aspect of the present invention, the second organic solvent includes: one or a mixture of two or more of dichloromethane, chloroform, and 1,2-dichloroethane.

[0017] Optionally, in the sixth implementation manner of the first aspect of the present invention, the catalyst includes one or a mixture of two or more of sulfuric acid, benzenesulfonic acid, 4-methylbenzenesulfonic acid, (R)- or (S)-camphorsulfonic acid, methanesulfonic acid, and trifluoroacetic acid.

[0018] Optionally, in the seventh implementation manner of the first aspect of the present invention, the molar ratio of fusidic acid to the catalyst is 1:(0.05 - 0.2).

[0019] Optionally, in the eighth implementation manner of the first aspect of the present invention, the second temperature includes -10°C to 10°C, and the second duration includes 3h to 5h.

[0020] Optionally, in the ninth implementation manner of the first aspect of the present invention, the third organic solvent includes one or a mixture of two or more of dichloromethane, 1,2-dichloroethane, acetonitrile, or chloroform.

[0021] In the embodiments of the present invention, a synthesis method is proposed. The entire route only requires two steps and the total yield can reach more than 50%. Compared with the yield of the generation route of fusidic acid impurity B reported in the Journal of China Pharmaceutical University (2018, 49(3): 322), there is a great improvement. In the synthesis route of fusidic acid impurity B provided by the present invention, the reagents used are easily obtained and the cost is low. In addition, the method provided by the present invention has the advantages of mild reaction conditions and simple operation. Column chromatography is not required for both steps of the reaction, and pure products can be obtained only by repeated slurrying. The HPLC purity is more than 90%. The synthesized product can be used as a reference substance for the quality research of fusidic acid, meeting the requirements of fusidic acid-related preparations in aspects such as research, production, storage, and clinical applications, and solving the technical problem of the too high cost of synthesizing impurity B of fusidic acid in the prior art. Description of the Drawings

[0022] Figure 1 It is the synthesis route diagram of the synthesis method of fusidic acid EP impurity B in the embodiments of the present invention;

[0023] Figure 2 It is the liquid chromatogram of fusidic acid EP impurity B in the embodiments of the present invention;

[0024] Figure 3 It is the mass spectrum of fusidic acid EP impurity B in the embodiments of the present invention;

[0025] Figure 4 It is the nuclear magnetic resonance hydrogen spectrum of fusidic acid EP impurity B in the embodiments of the present invention. Detailed Embodiments

[0026] An embodiment of the present invention provides a method for synthesizing impurity B of fusidic acid.

[0027] Embodiments of the present invention disclosed will be described in more detail below with reference to the accompanying drawings. Although some embodiments of the present invention are shown in the drawings, it should be understood that the present invention can be implemented in various forms and should not be construed as limited to the embodiments set forth herein. Instead, these embodiments are provided to more thoroughly and completely understand the present disclosure. It should be understood that the drawings and embodiments of the present invention are only for exemplary purposes and are not intended to limit the scope of protection of the present invention.

[0028] In the description of the embodiments of the present invention disclosed, the term "including" and its similar terms should be understood as open inclusion, that is, "including but not limited to". The term "based on" should be understood as "at least partially based on". The term "an embodiment" or "the embodiment" should be understood as "at least one embodiment". The terms "first", "second", etc. may refer to different or the same objects. There may also be other explicit and implicit definitions hereinafter.

[0029] For ease of understanding, the specific process of the embodiment of the present invention will be described below. Please refer to Figure 1 the synthetic route diagram of the method for synthesizing impurity B of fusidic acid in the embodiment of the present invention, which includes the steps:

[0030] Step 1: Fusidic acid is dissolved in a first organic solvent, an epoxidizing agent is added, and the reaction is carried out at a first temperature for a first duration, and then extracted to obtain compound I;

[0031] Step 2: Compound I is dissolved in a second organic solvent, a catalyst is added, and the reaction is carried out at a second temperature for a second duration, and then repulped and washed with a third organic solvent to obtain compound II. The reaction equations of steps 1-2 are as follows:

[0032]

[0033] In the embodiments of the present invention, various raw materials, reagents, and solvents used are commercially available without special instructions.

[0034] In the embodiments of the present invention, high performance liquid chromatography is used for purity detection. Chromatographic column model: Waters Sunfire C18 (4.6×150mm, 3.5um); detection wavelength: 235nm; column oven temperature: 30°C; flow rate: 1.0 mL / min; injection volume: 10.0 μL;

[0035] Mobile phase A: MeOH-ACN-5g / L H3PO4 = 20:40:40;

[0036] Mobile phase B: 5 g / L H3PO4 - MeOH - ACN = 10:20:70

[0037] The changes of mobile phase A and mobile phase B with time for purity detection by high - performance liquid chromatography are shown in Table 1, the table of the change of the proportion of mobile phase with time.

[0038] Table 1, the table of the change of the proportion of mobile phase with time

[0039] Time / min A％ B％ 0 100.0 0.0 3 100.0 0.0 28 0.0 100.0 33 0.0 100.0 34 100.0 0.0 40 100.0 0.0

[0040] In the example of Step 1, fusidic acid is dissolved in the first organic solvent, an epoxidizing agent is added, and the reaction is carried out at the first temperature for the first duration, and compound I is obtained by extraction. The reaction equation is as follows:

[0041]

[0042] Example 1

[0043] The molar ratio of fusidic acid to the epoxidizing agent is = 1:3.

[0044] 5.0 g of fusidic acid was added to the reaction flask, 80 mL of dichloromethane was added, and the reaction flask was placed in an ice - water bath. 5.8 g of 3 - chloroperoxybenzoic acid (abbreviation: m - CPBA, content 85%) was added at one time, and the reaction was kept at 0 °C. According to the reaction progress, after 0.5 h, it was detected that the fusidic acid raw material was completely consumed. A saturated sodium thiosulfate solution was added to quench the reaction, and it was extracted three times with dichloromethane. The organic phases were combined and dried with anhydrous sodium sulfate, then filtered. The filtrate was concentrated under vacuum conditions to obtain the crude intermediate shown in compound I, which was used as the raw material for the next step without further purification immediately.

[0045] Example 2

[0046] The molar ratio of fusidic acid to the epoxidizing agent is = 1:3.

[0047] 5.0 g of fusidic acid was added to the reaction flask, 70 mL of chloroform was added, and the reaction solution was cooled to - 10 °C. 5.8 g of 3 - chloroperoxybenzoic acid (abbreviation: m - CPBA, content 85%) was added at one time, and the reaction was kept at - 10 °C. According to the reaction progress, after 1.5 h, it was detected that the fusidic acid raw material was completely consumed. A saturated sodium thiosulfate solution was added to quench the reaction, and it was extracted three times with chloroform. The organic phases were combined and dried with anhydrous sodium sulfate, then filtered. The filtrate was concentrated under vacuum conditions to obtain the crude intermediate shown in compound I, which was used as the raw material for the next step without further purification immediately.

[0048] Example 3

[0049] The molar ratio of fusidic acid to the epoxidizing agent is = 1:3.

[0050] Add 5.0 g of fusidic acid into a reaction flask, add 70 mL of dichloromethane, cool this reaction solution to -5 °C, and add 5.8 g of 3-chloroperoxybenzoic acid (abbreviation: m-CPBA, content 85%) all at once. Keep the reaction proceeding at -5 °C. According to the reaction progress, after 1 h, it is detected that the fusidic acid raw material is completely consumed. Add a saturated sodium thiosulfate solution to quench the reaction, extract with dichloromethane three times, combine the organic phases and dry with anhydrous sodium sulfate, then filter. Concentrate the filtrate under vacuum conditions to obtain the crude intermediate shown in Compound I, which is used as the raw material for the next reaction immediately without further purification.

[0051] Example 4

[0052] The molar ratio of fusidic acid to the epoxidizing agent is = 1:1.5.

[0053] Add 5.0 g of fusidic acid into a reaction flask, add 80 mL of dichloromethane, place this reaction flask in an ice-water bath, and slowly dropwise add 1.3 mL (1.06 g / mL) of peracetic acid. Keep the reaction proceeding at 10 °C. According to the reaction progress, after 0.5 h, it is detected that the fusidic acid raw material is completely consumed. Keep the temperature at 0 °C and slowly add a saturated sodium thiosulfate solution to quench the reaction, extract with dichloromethane three times, combine the organic phases and dry with anhydrous sodium sulfate, then filter. Concentrate the filtrate under vacuum conditions to obtain the crude intermediate shown in Compound I, which is used as the raw material for the next reaction immediately without further purification.

[0054] In the example of Step 2, dissolve Compound I in a second organic solvent, add a catalyst, and react at a second temperature for a second duration. Then, repulp and wash with a third organic solvent to obtain Compound II. The reaction equation is as follows:

[0055]

[0056] Example 1

[0057] The molar ratio of fusidic acid to the catalyst is = 1:0.1.

[0058] Place the crude intermediate obtained in Example 1 of Step 1 in a reaction flask, add 80 mL of dichloromethane, and cool the reaction flask to -5 °C. Add 225 mg of camphorsulfonic acid (abbreviation CSA). Keep the reaction temperature not exceeding -10 °C, and continue the reaction for 3.5 h. Then, the reaction mixture is slowly restored to room temperature, stirred vigorously for 20 minutes, filtered, the filter cake is washed with an appropriate amount of dichloromethane, drained, the filter cake is collected, dichloromethane is added again, and the pulping operation is repeated three times in total. The obtained white insoluble matter is collected in a container and dried under vacuum to obtain 2.63 g of pure fusidic acid EP impurity B, with a total yield of 51% and a measured purity of 91.3%.

[0059] Example 2

[0060] Fusidic acid: The molar ratio of the catalyst is = 1:0.2.

[0061] Place the crude intermediate obtained in Example 1 of Step 1 in a reaction flask, add 80 mL of chloroform, and place the reaction flask in normal temperature water (the measured temperature is 10 °C). Add 449 mg of camphorsulfonic acid (abbreviation CSA). Keep the reaction temperature at 10 °C, and continue the reaction for 3 h. Then, the reaction mixture is slowly restored to room temperature, stirred vigorously for 20 minutes, filtered, the filter cake is washed with an appropriate amount of chloroform, drained, the filter cake is collected, chloroform is added again, and the pulping operation is repeated three times in total. The obtained white insoluble matter is collected in a container and dried under vacuum to obtain 2.53 g of pure fusidic acid EP impurity B, with a total yield of 49% and a measured purity of 91.0%.

[0062] Example 3

[0063] Fusidic acid: The molar ratio of the catalyst is = 1:0.05.

[0064] Place the crude intermediate obtained in Example 1 of Step 1 in a reaction flask, add 80 mL of 1,2-dichloroethane, and cool the reaction flask to -10 °C. Add 167 mg of 4-methylbenzenesulfonic acid. Keep the reaction temperature at -10 °C, and continue the reaction for 5 h. Then, the reaction mixture is slowly restored to room temperature, stirred vigorously for 20 minutes, filtered, the filter cake is washed with an appropriate amount of 1,2-dichloroethane, drained, the filter cake is collected, 1,2-dichloroethane is added again, and the pulping operation is repeated three times in total. The obtained white insoluble matter is collected in a container and dried under vacuum to obtain 2.49 g of pure fusidic acid EP impurity B, with a total yield of 48% and a measured purity of 90.6%.

[0065] Example 4

[0066] Fusidic acid: The molar ratio of the catalyst is = 1:0.05.

[0067] The crude intermediate obtained in Example 1 of Step 1 was placed in a reaction flask, 80 mL of dichloromethane was added, and the reaction flask was placed in an ice-water bath (the measured temperature was 0 °C). 112 mg of camphorsulfonic acid (abbreviation CSA) was added. The reaction temperature was maintained at 0 °C, and the reaction continued for 4.5 h. Then the reaction mixture was slowly restored to room temperature, stirred vigorously for 20 minutes, filtered, the filter cake was washed with an appropriate amount of dichloromethane, drained, the filter cake was collected, dichloromethane was added again, and the pulping operation was repeated three times in total. The obtained white insoluble matter was collected in a container and dried under vacuum to obtain 2.46 g of pure fusidic acid EP impurity B, with an overall yield of 48% and a measured purity of 90.9%.

[0068] Please refer to Figure 2 , Figure 2 which is the liquid chromatogram of fusidic acid EP impurity B in the embodiment of the present invention. In the positive ion mass spectrum of the fusidic acid EP impurity B sample, there is a strong ion peak at 550.3, which is [M+NH4] of the sample + , which is consistent with the calculated value corresponding to C 31 H 52 NO7 + . The specific spectrum is shown in Figure 3 , Figure 3 which is the mass spectrum of fusidic acid EP impurity B in the embodiment of the present invention;

[0069] Figure 4 which is the 1H nuclear magnetic resonance spectrum of fusidic acid EP impurity B in the embodiment of the present invention. The analysis data is: 1 1H NMR (400 MHz, DMSO-d6).

[0070] In the embodiment of the present invention, a synthesis method is proposed. The entire route only requires two steps and the overall yield can reach more than 50%. Compared with the yield of the generation route of fusidic acid impurity B reported in the Journal of China Pharmaceutical University (2018, 49(3): 322), there is a great improvement. In the synthesis route of fusidic acid impurity B provided by the present invention, the reagents used are easily obtained and have low cost. In addition, the method provided by the present invention has the advantages of mild reaction conditions and simple operation. Column chromatography is not required for both steps of the reaction, and pure products can be obtained only by pulping multiple times, with an HPLC purity of more than 90%. The synthesized product can be used as a reference substance for the quality research of fusidic acid, meeting the requirements of fusidic acid-related preparations in aspects such as research, production, storage, and clinical application, and solving the technical problem of the too high cost of synthesizing a large amount of impurity B of fusidic acid in the prior art.

[0071] In addition, although the operations are depicted in a particular order, this should be understood as requiring that the operations be performed in the particular order shown or in sequential order, or that all of the illustrated operations be performed to achieve the desired result. In certain circumstances, multitasking and parallel processing may be advantageous. Similarly, although several specific implementation details are included in the above discussion, these should not be construed as limiting the scope of the present disclosure. Certain features described in the context of separate embodiments may also be implemented in combination in a single implementation. Conversely, the various features described in the context of a single implementation may also be implemented separately or in any suitable sub-combination in multiple implementations.

[0072] Although the subject matter has been described in language specific to structural features and / or methodological logical acts, it should be understood that the subject matter defined in the appended claims is not necessarily limited to the specific features or acts described above. Rather, the specific features and acts described above are merely example forms of implementing the claims.

Claims

1. A method for synthesizing impurity B of fusidic acid, characterized in that, Comprising the steps: Step 1: Fusidic acid is dissolved in a first organic solvent, an epoxidizing agent is added, and the reaction is carried out at a first temperature for a first duration, and compound I is obtained by extraction; Step 2: Compound I is dissolved in a second organic solvent, a catalyst is added, and the reaction is carried out at a second temperature for a second duration, and compound II is obtained by repulping and washing with a third organic solvent. The reaction equations for Steps 1-2 are as follows:

2. The synthetic method of impurity B of fusidic acid according to claim 1, characterized in that, The first organic solvent includes: one or a mixture of two or more of dichloromethane, chloroform, 1,2-dichloroethane, acetonitrile, and ethyl acetate.

3. The synthesis method of impurity B of fusidic acid according to claim 1, characterized in that, The epoxidizing reagent includes: any one of 3-chloroperoxybenzoic acid, peroxybenzoic acid, peracetic acid, and carboxylic acid derivatives.

4. The synthetic method of impurity B of fusidic acid according to claim 1, characterized in that, The molar ratio of fusidic acid to epoxidizing agent is = 1:(1.5-3).

5. The synthesis method of impurity B of fusidic acid according to claim 1, characterized in that, The first temperature includes: -10°C to 10°C, and the first duration includes: 0.5 h to 1.5 h.

6. The synthetic method of impurity B of fusidic acid according to claim 1, characterized in that, The second organic solvent includes: one or a mixture of two or more of dichloromethane, chloroform, and 1,2-dichloroethane.

7. The method for synthesizing impurity B of fusidic acid according to claim 1, characterized in that, The catalyst includes: one or a mixture of two or more of sulfuric acid, benzenesulfonic acid, 4-methylbenzenesulfonic acid, (R)- or (S)-camphorsulfonic acid, methanesulfonic acid, and trifluoroacetic acid.

8. The synthetic method of impurity B of fusidic acid according to claim 1, characterized in that, The molar ratio of fusidic acid to catalyst is = 1:(0.05-0.2).

9. The synthetic method of impurity B of fusidic acid according to claim 1, characterized in that, The second temperature includes: -10°C to 10°C, and the second duration includes: 3 h to 5 h.

10. The method for synthesizing impurity B of fusidic acid according to claim 1, characterized in that, The third organic solvent includes: one or a mixture of two or more of dichloromethane, 1,2-dichloroethane, acetonitrile, or chloroform.

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