Method for preparing milobalin by chemical-enzyme method

Abstract

The invention discloses a method for preparing miloabalin by a chemical-enzyme method. According to the method disclosed by the invention, a compound II is taken as a starting material, the starting material is converted into a compound IV through Horner-Wadsword-Emmons and addition reaction, then the compound IV is converted into a five-membered lactam ring compound V, and finally the compound V is converted into the milobalin under the action of amide hydrolase. The method disclosed by the invention is simple to operate, high in yield, low in cost and more suitable for industrial production, and the use of expensive resolving agents and material waste are avoided.

Description

Technical field:

[0001] The invention belongs to the technical field of biocatalysis, and in particular relates to a method for preparing milobarlin by a chemical-enzymatic method. Background technology:

[0002] Mirogabalin was developed by Daiichi Sankyo Co., Ltd. and launched in Japan on January 8, 2019 under the trade name Tarlige. It is mainly used to treat peripheral neuropathic pain (PNP), including diabetic PNP (DPNP) and postherpetic neuralgia (PHN). Its chemical structure is as follows:

[0003]

[0004] Daiichi Sankyo Co., Ltd. disclosed a method for preparing milobarline in patent CN101878193B, and the route is shown in Scheme 1. The method uses compound II as the starting material, and obtains compound 1 through Horner-Wadsworth-Emmons reaction, and then undergoes Michael addition with nitromethane to obtain compound 2. Compound 2 is separated by column chromatography to obtain intermediate 3, and finally milobarline is obtained through nitro reduction, de-tert-butylation and other reactions. Chromatographic separation is used in this route, which is cumbersome to operate, has low yield, high cost, and is not suitable for industrial production. Daiichi Sankyo Co., Ltd. improved the above route in patent CN102356061B, that is, compound 2 is first reduced to nitro to generate compound 5, and then separated with D-mandelic acid to obtain compound 6. The optimized route avoids the complicated column chromatography, but still cannot avoid the loss of nearly 50% of the product.

[0005]

[0006] Scheme 1

[0007] Patent CN111116345A discloses a new route, as shown in Scheme 2. This method uses compound 7 as the starting material, and sequentially reacts with cyanoacetate, sulfuric acid, urea, hypohalite, etc. to obtain racemic milobarlin hydrochloride 11, and finally obtains milobarlin by splitting. This route still involves the use of a splitting agent, and the reaction conditions from compound 9 to compound 11 are harsh, with a reaction temperature as high as 155°C, which poses certain safety risks.

[0008]

[0009] Scheme 2

[0010] In the prior art, the preparation methods of milobarlin have low yields and require splitting, which is not suitable for industrialization. Therefore, it is necessary to develop a preparation method that is simple to operate, high in yield, low in cost and suitable for industrialization. Summary of the invention:

[0011] The object of the present invention is to provide a method for preparing milopalline by an efficient chemical-enzymatic method in view of the deficiencies in the prior art.

[0012] The technical solution adopted by the present invention is as follows:

[0013]

[0014] The present invention provides a method for preparing milopalline, which specifically comprises the following steps:

[0015] Step 1: Compound II undergoes Horner-Wadsworth-Emmons reaction with dimethoxyphosphorylethyl tert-butyl ester to generate compound III;

[0016] Step 2: Under the action of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), compound III reacts with nitromethane to obtain compound IV;

[0017] Step 3: Under acidic conditions, compound IV is reduced and amine esterified under the action of iron powder to generate compound V;

[0018] Step 4: The racemic compound V undergoes hydrolysis reaction under the action of amidohydrolase to obtain milobarlin.

[0019] Furthermore, the acid in step 3 is acetic acid or hydrochloric acid, preferably acetic acid.

[0020] Furthermore, in step 3, the equivalent ratio of compound IV to acid is 1:1.5-1:3, preferably 1:2.

[0021] Furthermore, in step 3, the reaction temperature is 20-70°C, preferably 50-55°C.

[0022] Further, the solvent used in step 3 includes but is not limited to methanol, ethanol, propanol, isopropanol, tetrahydrofuran and combinations thereof, preferably ethanol.

[0023] Furthermore, the amide hydrolase in step 4 is derived from Acetohalobium arabaticum, Streptomyces griseofuscus, or Streptomyces anthocyanicus.

[0024] Furthermore, in step 4, the amide hydrolase participates in the catalytic reaction in the form of amide hydrolase powder, amide hydrolase solution, amide hydrolase homogenate solution, cells containing amide hydrolase, etc., preferably amide hydrolase homogenate solution.

[0025] Furthermore, the concentration of the amidohydrolase homogenate solution is 10 to 100 g / L.

[0026] Furthermore, the amide hydrolase is selected from the amide hydrolase ES-AMD-101 to ES-AMD-119 produced by Shangke Biopharmaceutical (Shanghai) Co., Ltd.

[0027] The beneficial effect of the present invention is that the present invention provides a new method for preparing milopalline, the new route disclosed by the present invention uses an amidohydrolase to construct a chiral center, avoids the use of expensive resolving agents and waste of materials, the total yield is as high as more than 70%, and the chiral purity is ≥99%. The route is simple to operate, has high atom economy, and can be used for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] none DETAILED DESCRIPTION

[0029] The technical content of the present invention is further described below in conjunction with specific embodiments, the purpose of which is to provide a better understanding of the content of the present invention, but the protection scope of the present invention is not limited thereto.

[0030] Example 1 Preparation of Compound III

[0031] Under nitrogen protection, compound II (130g, 0.955mol, 1.0eq), triethyl phosphoacetate (256.9g, 1.15mol, 1.2eq), and 1300mL of THF were added to a 250mL three-necked flask. After the addition was complete, the temperature was lowered to 0°C, and NaH (57.3g, 1.43mol, 1.5eq, 60%) was added to the reaction system in batches. After the addition was complete, the temperature was raised to room temperature and stirred for 1h to terminate the reaction. Water was added to the mixture to quench the reaction, the system was separated, the aqueous phase was washed with ethyl acetate 3 times, the organic phases were combined, and concentrated under reduced pressure to obtain 187.1g of compound III, the product was a brown oily liquid, and the reaction yield was 95.1%.

[0032] Example 2 Preparation of Compound IV

[0033] Under nitrogen protection, compound III (140 g, 0.68 mol, 1.0 eq) was dissolved in nitromethane (430 mL), and then 1,8-diazabicyclo[5.4.0]undec-7-ene (310 g, 2.03 mol, 3.0 eq) was slowly added dropwise. After the addition was complete, the temperature was raised to 50-55 °C and stirred for 3 h to complete the reaction. 200 mL of saturated KH2 PO 4 The solution was used to quench the reaction, and the aqueous phase was extracted three times with methyl tert-butyl ether. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 170.54 g of compound IV. The product was an oily liquid, and the reaction yield was 94%.

[0034] Example 3 Preparation of Compound V

[0035] Compound IV (25 g, 0.094 mol, 1.0 eq), acetic acid (11.2 g, 0.187 mol, 2.0 eq), iron powder (31.5 g, 0.56 mol, 6.0 eq), and anhydrous ethanol (250 mL) were added to a 250 mL three-necked flask, and the temperature was raised to 55 ° C. The reaction was stirred for 5 h and then cooled to room temperature. The solid in the system was filtered out and the organic phase was dried to obtain a crude compound V. The crude product was further dissolved in 100 mL of dichloromethane. After filtering out the insoluble matter, the filtrate was dried to obtain 15.8 g of compound V with a yield of 88% and a purity of 97%.

[0036] Example 4 Preparation of Compound V

[0037] Compound IV (25 g, 0.094 mol, 1.0 eq), 8.3 g of hydrochloric acid, iron powder (31.5 g, 0.56 mol, 6.0 eq), and anhydrous ethanol (250 mL) were added to a 250 mL three-necked flask, and the temperature was raised to 50 ° C. The reaction was stirred for 5 h and then cooled to room temperature. The solid in the system was filtered out and the organic phase was dried to obtain a crude compound V. The crude product was further dissolved in 100 mL of dichloromethane. After filtering out the insoluble matter, the filtrate was dried to obtain 14.4 g of compound V with a yield of 80% and a purity of 95%.

[0038] Example 5 Preparation of Compound I

[0039] Compound V (21 g) and amide hydrolase homogenate (1.0 v / w) and water (15 v / w) were added to a 500 mL three-necked flask, and the temperature was raised to 40° C. with stirring. During the reaction, a 10% sodium carbonate aqueous solution was used to adjust the pH to 7.9-8.0. After TLC monitoring showed that the substrate disappeared, the temperature was lowered to room temperature, the reaction solution was centrifuged, the supernatant obtained was filtered, and the solution was washed twice with 20 mL of pure water. The filtrate was concentrated to 100 mL under reduced pressure, and the pH was adjusted to 5 with 2N hydrochloric acid. A large amount of solid was precipitated, filtered, and the filter cake was washed twice with 20 mL of pure water. The obtained solid was dried under vacuum at 50° C. Finally, 21.38 g of compound I was obtained, and the reaction yield was 93.0%, the purity was >99.2%, and the chiral purity was >99.1%.

[0040] Example 6 Preparation of Compound I

[0041] 400g of 0.05M phosphate buffer (pH=7.5) was added to a 1000mL reaction bottle, 21g of compound V was added under stirring, the system temperature was controlled to 35°C, stirred evenly, and 1g of amidohydrolase powder (ES-AMD-101) was added at one time under stirring. The stirring reaction was started, and the pH was adjusted to 7.9-8.0 with 10% sodium carbonate aqueous solution during the reaction. The HPLC test was performed, and the conversion rate was more than 98%, and the reaction was completed. 300mL of ethyl acetate was added to the system, the reaction solution was filtered, hydrochloric acid was added to the filtrate to adjust the pH to 5, a large amount of solid was precipitated, filtered, the filter cake was washed twice with 20mL of pure water, and the obtained solid was vacuum dried at 50°C to finally obtain 20.68g of compound I, the reaction yield was 90.1%, the purity was>99.0%, and the chiral purity was>99.3%.

Claims

1. A method for preparing milopalline by chemical-enzymatic method, characterized in that: Compound II is subjected to Horner-Wadsworth-Emmons and Michael addition reactions to obtain compound IV, and then compound IV is reduced and esterified by amines under the action of a reducing agent to obtain compound V. Finally, compound V is subjected to the action of an amidohydrolase to obtain milobarlin. The reaction formula is shown below:

2. The method according to claim 1, characterized in that The reducing agent is selected from iron powder.

3. The method according to claim 1, characterized in that The equivalent ratio of the compound IV to the acid is 1:1.5 to 1:

3.

4. The method according to claim 1, characterized in that The amide hydrolase is selected from the amide hydrolase ES-AMD-101 to ES-AMD-119 produced by Shangke Biopharmaceuticals (Shanghai) Co., Ltd.

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