Preparation method of (S)-1-(2, 6-dichlorophenyl) ethanol

Abstract

The invention provides a preparation method of (S)-1-(2, 6-dichlorophenyl) ethanol, which is characterized in that a biological catalytic synthesis method is adopted, a coenzyme circulation system formed by coenzyme, a hydrogen donor and dehydrogenase is used for catalytic reduction of a substrate 2 ', 6'-dichloroacetophenone, and the substrate 2 ', 6'-dichloroacetophenone is converted into a target product. And after the hydrogen transfer step is finished, recovering the coenzyme to the oxidation state, and circularly putting the coenzyme into the oxidation reaction of the hydrogen donor. According to the synthesis route provided by the invention, high-risk hydrogenation operation in a traditional method is avoided, nitrogen protection is not used, and the reaction is carried out under mild and safe conditions. The purity of the product prepared by the method is as high as 99%, and the product has double benefits of high yield and low cost.

Description

Technical Field

[0001] The invention belongs to the technical field of biochemical synthesis, and specifically relates to a method for preparing (S)-1-(2,6-dichlorophenyl)ethanol. Background Art

[0002] Chiral alcohols with 2,6-disubstituted phenyl groups play an important role in the synthesis of many chiral drugs, agrochemicals and bioactive substances. Vishal et al. reported a series of drug molecules that inhibit Gram-negative bacteria, which are of great significance for alleviating the current super-resistant bacteria infection. Their typical structures are shown in Scheme I below. (S)-1-(2,6-dichlorophenyl)ethanol is a key structural unit of this type of molecule. In addition, (S)-1-(2,6-dichlorophenyl)ethanol is also an intermediate in the synthesis of c-Met inhibitors and can be used as a nicotinic acid receptor antagonist.

[0003]

[0004] Asymmetric reduction of 2',6'-dichloroacetophenone is the most economical solution for the synthesis of (S)-1-(2,6-dichlorophenyl)ethanol, but there are still some challenges in this solution. Chengrong Lu et al. reported a method of using a rare earth metal complex containing Trost ligand to catalyze this reaction, but the selectivity of the catalyst was poor, resulting in the ee value of the product (S)-1-(2,6-dichlorophenyl)ethanol being only 74% (J.Org.Chem.2020,85,10504). Wei Sun et al. also reported a method of asymmetric reduction of large hindered ketones containing 2,6-disubstituted phenyl groups by transfer hydrogenation using a manganese catalyst, which achieved good results in a series of reactions, but for 2',6'-dichloroacetophenone, there was also a problem of poor selectivity, resulting in an ee value of 92% for the product (J.Catal.438,2024,115680). Summary of the invention

[0005] In view of the challenges faced by the preparation of (S)-1-(2,6-dichlorophenyl)ethanol in the prior art, such as insufficient selectivity and purity, the present invention provides a method for preparing (S)-1-(2,6-dichlorophenyl)ethanol, which, under the action of a specific catalyst, combines a coenzyme circulation system composed of a coenzyme, a hydrogen donor and a dehydrogenase to effectively catalyze the reduction of 2',6'-dichloroacetophenone to the desired product. After the hydrogen transfer process is completed, the coenzyme is reoxidized to its active form, so that it can continue to participate in the oxidation cycle of the hydrogen donor and realize the recycling of the coenzyme. The biosynthesis technology adopted in the present invention abandons the dangerous hydrogenation step in the traditional synthesis process, does not require nitrogen protection, and ensures that the entire reaction process is carried out under mild and safe conditions. In addition, the method not only ensures the high reliability of the reaction process, but also the purity of the obtained product can reach 99%, while having the dual advantages of high yield and low cost.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions:

[0007] A method for preparing (S)-1-(2,6-dichlorophenyl)ethanol comprises the following steps: under the conditions of ketoreductase and a coenzyme circulation system consisting of a coenzyme, a hydrogen source and a dehydrogenase, catalytically reducing a substrate 2',6'-dichloroacetophenone to the (S)-1-(2,6-dichlorophenyl)ethanol.

[0008] In some embodiments, the coenzyme is any one or more combinations of NADP salt, NADPH, NADH, and NAD salt.

[0009] In some embodiments, the hydrogen source is any one of isopropanol, n-butanol, glucose, and formic acid.

[0010] In some embodiments, the dehydrogenase is any one of alcohol oxidase, glucose dehydrogenase, and formate dehydrogenase.

[0011] In some embodiments, the ketoreductase is any one of SY-KRED110 to SY-KRED119 or a combination of at least two thereof.

[0012] In some embodiments, the preparation method further comprises: adding a co-solvent for dissolving the substrate into the reaction system; and adding a buffer into the reaction system to adjust the pH value of the reaction system.

[0013] In some embodiments, the co-solvent is any one of dimethyl sulfoxide, isopropanol, n-butanol, methanol, and tetrahydrofuran.

[0014] In some embodiments, the buffer is any one of potassium phosphate buffer, tris(hydroxymethyl)methylamine-hydrochloric acid buffer, and sodium phosphate buffer; and the pH value of the buffer is 5.5-8.0.

[0015] In some embodiments, the reaction temperature of the catalytic reduction is 25-35° C.; the reaction time of the catalytic reduction is 10-24 h.

[0016] Compared with the prior art, the preparation method of (S)-1-(2,6-dichlorophenyl)ethanol provided by the present invention has the following beneficial effects:

[0017] 1. The present invention adopts biological method to synthesize (S)-1-(2,6-dichlorophenyl)ethanol, and efficiently catalyzes the reduction of 2',6'-dichloroacetophenone through a coenzyme circulation system. The method is easy to operate, avoids the traditional dangerous hydrogenation operation, does not require nitrogen protection, and ensures the green, safe and reliable reaction. At the same time, the method also has the advantages of high yield and low cost;

[0018] 2. The preparation method provided by the present invention has excellent chemical selectivity and can prepare products with a purity of more than 99.0%, significantly improving product quality;

[0019] 3. The preparation method provided by the present invention has mild reaction conditions, avoiding the need for low-temperature and high-temperature reactions. Since it does not involve extreme temperature conditions, there is no need to invest in special high-temperature or low-temperature reaction equipment, which not only significantly reduces energy consumption, but also greatly reduces production costs, achieving a dual improvement in economic benefits and environmental protection. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The present invention is further described below in conjunction with the accompanying drawings and embodiments.

[0021] Figure 1 is the HPLC chromatogram of substrate 2',6'-dichloroacetophenone;

[0022] Figure 2 The HPLC chromatogram of (S)-1-(2,6-dichlorophenyl)ethanol provided by the present invention;

[0023] Figure 3 HPLC analysis spectrum of the reaction product in Example 3;

[0024] Figure 4 This is the chiral HPLC analysis spectrum of the reaction product in Example 3;

[0025] Figure 5 This is the HNMR analysis diagram of the S-configuration product;

[0026] Figure 6This is the chiral HPLC analysis spectrum of the reaction product in Example 4. DETAILED DESCRIPTION

[0027] The present invention is further explained in detail below in conjunction with the drawings and descriptions of specific embodiments. However, the following descriptions including the embodiments are only used to enable ordinary technicians in the technical field to which the present invention belongs to more clearly understand the principles and essence of the present invention, and do not mean any form of limitation on the present invention.

[0028] Example 1

[0029] The present invention provides a method for preparing (S)-1-(2,6-dichlorophenyl)ethanol, comprising the following steps:

[0030] Under the conditions of a catalyst and a coenzyme circulation system consisting of a coenzyme, a hydrogen source and a dehydrogenase, the substrate 2',6'-dichloroacetophenone is catalytically reduced to (S)-1-(2,6-dichlorophenyl)ethanol.

[0031] The specific process is briefly described as follows: Under suitable conditions (such as specific pH value and temperature), the substrate, catalyst and coenzyme circulation system are mixed to start the reduction reaction.

[0032] Oxidation of hydrogen source: Dehydrogenase catalyzes the oxidation reaction of hydrogen source, transfers hydrogen atoms in the hydrogen source to the coenzyme, and forms a reduced coenzyme. Under the action of the catalyst, the reduced coenzyme acts as a hydrogen donor, transfers hydrogen atoms to the carbonyl group of 2',6'-dichloroacetophenone, forms a hydroxyl group, and thus reduces 2',6'-dichloroacetophenone to (S)-1-(2,6-dichlorophenyl)ethanol.

[0033] After completing the hydrogen transfer, the coenzyme is oxidized back to its oxidized form, and can then participate in the oxidation reaction of the hydrogen source again and continue to be recycled.

[0034] Compared with the method for preparing (S)-1-(2,6-dichlorophenyl)ethanol by asymmetric hydrogenation reaction with complicated operation commonly used in the prior art, the present invention adopts a coenzyme circulation system for synthesis, avoiding the potential dangerous operation in the traditional hydrogenation process, and also omitting the step of using an inert gas (such as nitrogen) to replace the air in the reactor, thereby greatly simplifying the operation process.

[0035] The coenzyme circulation system provided by the present invention can continuously carry out catalytic reaction by recycling coenzymes, thereby improving the yield of the final product. Moreover, the catalytic reaction is carried out under mild conditions, thereby reducing the generation of harmful byproducts and reducing environmental pollution.

[0036] In some embodiments, the above-mentioned coenzyme is selected from any one or more combinations of NADP salt, NADPH, NADH, and NAD salt.

[0037] Preferably, the coenzyme is NADP salt.

[0038] In some embodiments, the hydrogen source is selected from any one of isopropanol, n-butanol, glucose, and formic acid.

[0039] Preferably, the hydrogen source is glucose.

[0040] In some embodiments, the dehydrogenase is selected from any one of alcohol oxidase, glucose dehydrogenase, and formate dehydrogenase.

[0041] Preferably, the dehydrogenase is selected as glucose dehydrogenase.

[0042] In some embodiments, the ketoreductase is selected from any one or a combination of at least two of SY-KRED110 to SY-KRED119.

[0043] Preferably, the ketoreductase is SY-KRED115.

[0044] In some embodiments, the reaction temperature of the catalytic reduction process is 25-35° C., and the reaction time of the catalytic reduction process is 10-24 h.

[0045] In some embodiments, the preparation method further comprises the following steps:

[0046] (1) Add a co-solvent to the reaction system to dissolve the substrate.

[0047] The cosolvent is any one of dimethyl sulfoxide, isopropanol, n-butanol, methanol and tetrahydrofuran, preferably dimethyl sulfoxide.

[0048] (2) Add a buffer solution to the reaction system to adjust the pH value of the reaction system.

[0049] The buffer is any one of potassium phosphate buffer, tris(hydroxymethyl)methylamine-hydrochloric acid buffer and sodium phosphate buffer, and the pH value of the buffer is 5.5-8.0.

[0050] Some specific examples are provided below to illustrate the preparation method of (S)-1-(2,6-dichlorophenyl)ethanol provided by the present invention:

[0051] Example 2

[0052] This example is a screening example of ketoreductase. The reaction system components are as follows: 0.1M potassium phosphate buffer with pH 7.0, substrate includes: 10g / L 2',6'-dichloroacetophenone and 5% (V / V) isopropanol as a cosolvent, coenzyme is 0.5g / L NADP, ketoreductase is SY-KRED110~SY-KRED119 enzyme powder, each taking 1g / L. Reaction at 30°C for 18h. Among them, the reaction results using various ketoreductases are shown in Table 1 below:

[0053] Table 1 Reaction conversion rate of SY-KRED110 to SY-KRED119 catalysts and chiral purity of products

[0054] Screening Number Ketoreductase Reaction conversion rate Chiral purity (S type) 1-1 SY-KRED110 100.0％ 96.7％ 1-2 SY-KRED111 11.4％ 82.8％ 1-3 SY-KRED112 46.7％ 95.6％ 1-4 SY-KRED113 20.5％ 95.3％ 1-5 SY-KRED114 97.3％ 80.7％ 1-6 SY-KRED115 99.8％ 99.9％ 1-7 SY-KRED116 99.9％ 96.6％ 1-8 SY-KRED117 99.9％ 94.2％ 1-9 SY-KRED118 100.0％ 95.9％ 1-10 SY-KRED119 18.0％ 97.3％

[0055] The results showed that the ketoreductase with the best conversion effect was SY-KRED115 No. 6, and the HPL C conversion rate exceeded 99.8%.

[0056] Example 3

[0057] This example is to analyze the chiral characteristics of the product obtained by ketoreductase catalysis reaction

[0058] The ketoreductase SY-KRED115 enzyme powder with good reaction results was used to prepare (S)-1-(2,6-dichlorophenyl)ethanol, specifically:

[0059] Add 0.05M potassium phosphate buffer with pH 7.0, 1.0g substrate (100g / L 2',6'-dichloroacetophenone), 2.0ml isopropanol as cosolvent, 2mg NADP as coenzyme, 10mg ketoreductase, and water to the 10ml reaction system. React at 30℃ for 24h, and monitor the reaction with HPLC.

[0060] The peak position of the product in HPLC analysis is as follows Figure 2 As shown, the peak position of the substrate in conventional liquid phase analysis is as follows Figure 1 The HPLC analysis spectrum of the reaction product is shown in Figure 3 As shown, the reaction conversion rate is 99.7%, the chiral purity is 99.9%, and the chiral HPLC analysis spectrum of the reaction product is shown in Figure 4 After the product was purified, 0.91 g of the product was obtained, with a yield of 90.2% and a content of 98.0%. The HNMR analysis chart is as follows Figure 5 .

[0061] Example 4

[0062] Based on Example 3, this example is to amplify the reaction catalyzed by ketoreductase to a hundred-gram level:

[0063] Add 0.05M potassium phosphate buffer (pH 7.0), 0.2kg (substrate 100g / L 2',6'-dichloroacetophenone), 400ml isopropanol, 0.8g NADP, 4.0g ketoreductase to a 2L reaction system, and make up the rest with water. React at 30℃ for 24h. The chiral purity of the product is 99.8%, such as Figure 6 After purification, 184.0 g of product was obtained, with a yield of 91.1%.

[0064] Comparative Example 1

[0065] The only difference between this comparative example and Example 3 is that dehydrogenase GDH is added to the reaction system, and the matching hydrogen source is glucose. Because gluconic acid is produced during the reaction, 10% sodium carbonate is also needed to control the pH during the reaction to maintain it at about pH 7.0.

[0066] Add 0.05M potassium phosphate buffer (pH 7.0), 1.0g substrate (100g / L 2',6'-dichloroacetophenone), 2.0ml isopropanol, 2mg NADP, 10mg ketoreductase, 1mg GDH, 2.56g glucose to 10ml reaction system, and make up the rest with water. React at 30℃ for 18h, and monitor the reaction with HPLC.

[0067] Results: The reaction was completed and the chiral purity of the S-type product was 99.8%.

[0068] Conclusion: Although the chirality of the product remains unchanged, the conversion rate remains unchanged, and the chiral purity of the S-type product is also 99.8%, the reaction increases the reaction components and requires monitoring of pH, making the operation relatively complicated.

[0069] Comparative Example 2

[0070] The only difference between this comparative example and Example 3 is that the concentration of the substrate is increased to a certain extent.

[0071] Add 0.05M potassium phosphate buffer (pH 7.0), 1.5g substrate (150g / L 2',6'-dichloroacetophenone), 2.0ml propanol, 2mg NADP, 10mg ketoreductase to 10ml reaction system, and make up the rest with water. React at 30℃ for 24h, and monitor the reaction with HPLC.

[0072] Results: The reaction was not completed, and the conversion rate was about 95%; when the reaction time was extended to 48h, the conversion rate was 97%, and the reaction did not change. The chiral purity of the S-type product was 99.8%.

[0073] Conclusion: With increasing substrate amount, the substrate conversion rate was 95%, and after prolonging the reaction, the conversion rate changed little, to 97%; the chiral purity of the S-type product remained unchanged.

[0074] Comparative Example 3

[0075] The only difference between this comparative example and Example 3 is that the amount of the hydrogen source is increased.

[0076] Add 0.05M potassium phosphate buffer (pH 7.0), 1.5g substrate (150g / L 2',6'-dichloroacetophenone), 2.5ml isopropanol, 2mg NADP, 10mg ketoreductase to 10ml reaction system, and make up the rest with water. React at 30℃ for 24h, and monitor the reaction with HPLC.

[0077] Results: The reaction was not completed, and the conversion rate was about 96%; when the reaction time was extended to 48h, the conversion rate was 97%, and the reaction no longer changed. The chiral purity of the S-type product was 99.8%.

[0078] Conclusion: With increasing substrate amount, the substrate conversion rate was 96%. After prolonging the reaction, the conversion rate changed little, to 97%. The chiral purity of the S-type product remained unchanged.

[0079] Comparative Example 4

[0080] The only difference between this comparative example and Example 3 is that the amount of ketoreductase is reduced.

[0081] Add 0.05M potassium phosphate buffer (pH 7.0), 1.0g substrate (100g / L), 2.0ml isopropanol, 2mg NADP, 8mg ketoreductase to 10ml reaction system, and make up the rest with water. React at 30℃ for 24h, and monitor the reaction with HPLC.

[0082] Results: The reaction was not completed, and the conversion rate was about 94%; the reaction time was extended to 48h, the conversion rate was 96%, and the reaction time was further extended to 72h, the conversion rate was 98%. The chiral purity of the S-type product was 99.8%.

[0083] Results: After reducing the enzyme dosage, the substrate conversion rate was 94%, and after extending the reaction, the conversion rate changed slightly to 98%; the chiral purity of the S-type product remained unchanged.

[0084] Comparative Example 5

[0085] The difference between this comparative example and Example 2 is only that the amount of NADP is reduced.

[0086] Add 0.05M pH 7.0 potassium phosphate buffer, 1.0g substrate (100g / L), 2.0ml propanol, 1.5mg NADP, 10mg ketoreductase to 10ml reaction system, and make up the rest with water. React at 30℃ for 24h, and monitor the reaction status with HPLC.

[0087] Results: The reaction was not completed, and the conversion rate was about 96%; the reaction time was extended to 48h, the conversion rate was 98%, and the reaction time was further extended to 72h, the conversion rate was 99.5%. The chiral purity of the S-type product was 99.8%.

[0088] Conclusion: After reducing the amount of NADP, the substrate conversion rate was 96%, and after extending the reaction for 72 hours, the conversion rate was 99.5%, which was basically completed, but the time was long and the reaction energy consumption increased; the chiral purity of the S-type product remained unchanged.

[0089] In summary, by carefully adjusting the experimental conditions of Examples 2-5, including increasing the substrate concentration, hydrogen source, ketoreductase and coenzyme dosage, it is found that the purity of the obtained product still fails to reach the level shown in Example 3 of the present invention. Even if the reaction time is extended to promote the reaction process, the substrate conversion rate of these comparative examples is still lower than the conversion rate obtained in Example 3. As for Comparative Example 1, although the same substrate conversion rate and S-type product chiral purity as Example 3 can be achieved, other reaction components are additionally introduced during the experimental process of Comparative Example 1, which greatly increases the complexity of the experiment. Therefore, the conditions and parameters provided in Example 3 of the present invention obviously have more significant advantages in terms of product purity, substrate conversion rate and reaction efficiency.

[0090] The ideal embodiment of the present invention is inspiration. Through the above description, relevant personnel can make various changes and modifications without departing from the scope of the technical idea of ​​the present invention.

[0091] The technical scope of this invention is not limited to the contents in the specification, and its technical scope must be determined according to the scope of the claims.

Claims

1. A method for preparing (S)-1-(2,6-dichlorophenyl)ethanol, characterized in that: The steps include: Under the conditions of ketoreductase and a coenzyme circulation system consisting of a coenzyme, a hydrogen source and a dehydrogenase, the substrate 2',6'-dichloroacetophenone is catalytically reduced to the (S)-1-(2,6-dichlorophenyl)ethanol.

2. The preparation method according to claim 1, characterized in that: The coenzyme is any one or more combinations of NADP salt, NADPH, NADH and NAD salt.

3. The preparation method according to claim 1, characterized in that: The hydrogen source is any one of isopropanol, n-butanol, glucose and formic acid.

4. The preparation method according to claim 1, characterized in that: The dehydrogenase is any one of alcohol oxidase, glucose dehydrogenase and formate dehydrogenase.

5. The preparation method according to claim 1, characterized in that: The ketoreductase is any one of SY-KRED110 to SY-KRED119 or a combination of at least two thereof.

6. The preparation method according to any one of claims 1 to 5, characterized in that: Also includes: adding a co-solvent for dissolving the substrate into the reaction system; A buffer solution is added to the reaction system to adjust the pH value of the reaction system.

7. The preparation method according to claim 6, characterized in that: The co-solvent is any one of dimethyl sulfoxide, isopropanol, n-butanol, methanol and tetrahydrofuran.

8. The preparation method according to claim 6, characterized in that: The buffer is any one of potassium phosphate buffer, tris(hydroxymethyl)methylamine-hydrochloric acid buffer, and sodium phosphate buffer; The pH value of the buffer solution is 5.5-8.

0.

9. The preparation method according to any one of claims 1 to 5, characterized in that: The reaction temperature of the catalytic reduction is 25-35°C; The reaction time of the catalytic reduction is 10-24h.

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