Method for catalytically synthesizing (S)-3-chloro-1-phenyl-1-propanol by infrared light-driven WO3-based photo-enzyme catalyst

Abstract

The invention relates to the technical field of NADPH regeneration and the technical field of antidepressant drug synthesis, and discloses a method for catalytically synthesizing (S)-3-chloro-1-phenyl-1-propanol by driving a WO3-based photoenzyme catalyst through infrared light, and the method comprises the following steps: step S1, preparing an oxygen-deficient WO3 nanosheet; s2, preparing a NaCBR carbonyl reductase (NaCBR carbonyl reductase); s3, mixing the oxygen-deficient WO3 nanosheet and NaCBR carbonyl reductase according to the mass ratio of (1: 0.5)-(1: 2), and stirring for 3-6 hours to obtain the photo-enzyme catalyst (WO3 / NaCBR). According to the method, artificial photosynthesis, namely a method for driving NADPH regeneration, is utilized, other materials are not additionally used as hydrogen transfer and electron transfer media, and the (S)-3-chloro-1-phenylpropanol is catalytically synthesized by utilizing infrared light to drive a photo-enzyme catalyst (WO3 / NaCBR). The method can greatly reduce the cost, is green and environment-friendly, has high catalytic efficiency and mild reaction conditions, is simple to operate, is easy to recover and recycle, and is suitable for large-scale industrial production and application.

Description

Technical Field

[0001] The invention relates to the technical field of NADPH regeneration and the technical field of antidepressant drug synthesis, and in particular to a method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by catalytic synthesis of a WO3-based photoenzyme catalyst driven by infrared light. Background Art

[0002] (S)-3-Chloro-1-phenyl-1-propanol ((S)-CPPO) is a key chiral intermediate for the synthesis of chiral side chains of fluoxetine and tosooxetine. Fluoxetine is a class of drugs known as selective serotonin reuptake inhibitors and is a prescription drug for the treatment of depression. According to Newport Premium, the global market for fluoxetine hydrochloride was approximately US$7 billion in 2017, with global consumption of approximately 84 tons. (S)-Fluoxetine or (R)-fluoxetine is an isomer of fluoxetine and can be prepared from the chiral alcohol intermediate (S)-CPPO, which can be produced by enzymatic reduction of carbonyl ketones in the substrate 3-chloro-1-phenyl-1-propanone (3-CPP). Among them, optically active secondary alcohols are important components of many biologically active compounds in the pharmaceutical industry. In industrial production, asymmetric hydrogenation (AH) of prochiral ketones is a practical and simple method to obtain enantiomerically pure secondary alcohols. Although the method of preparing optical alcohols by AH has attracted widespread interest in academic and industrial research, due to the complexity of catalysts or harsh conditions, the chemical catalytic asymmetric synthesis of chiral alcohols usually has the problems of harsh conditions, expensive catalysts, and low enantiomeric excess of chiral alcohols. Most research is still in the stage of small-scale production or difficult large-scale production.

[0003] In recent years, the sudden rise of enzyme catalysis has made biocatalysis an important part that cannot be ignored. Enzyme-catalyzed reactions are simple, easy to control, highly efficient, and have mild reaction conditions, making them suitable for industrial applications. Moreover, the products are easy to separate and purify, and have high stereo and chemical selectivity, which has led to exponential growth in recent decades. Previous studies have found that carbonyl reductase (NaCBR) from Novosphingobium aromaticivorans can be used to reduce 3-CPP to synthesize (S)-CPPO due to its reliable chemical and stereo selectivity.

[0004] However, NAD(P)H is required as a cofactor in the process of catalyzing the carbonyl reductase reaction. NAD(P)H is not only expensive but also indispensable as a cofactor, and the amount used is often large, which is basically equivalent to the substrate ketone, which shows the economic disadvantage of this biocatalytic process. Enzyme-catalyzed regeneration of NAD(P)H (such as alcohol dehydrogenase and formate dehydrogenase (FDH)) is already established in industrial biocatalysis, but this process usually requires the regeneration of both the enzyme and the corresponding substrate, which is limited by the limited stability of the enzyme and the high separation cost. In addition, current research on NADPH regeneration basically requires the use of such as [Cp*Rh(bpy)(H2O)] 2+ The purpose of NADPH regeneration can only be achieved by using precious metal ligands such as hydrogen transfer and electron transfer media, but this will inevitably increase the high cost and greatly limit its industrial application.

[0005] Therefore, developing a simple, efficient and low-cost scheme for NAD(P)H regeneration and (S)-CPPO synthesis has become an issue that needs to be explored urgently by cutting-edge scholars in the industry. Summary of the invention

[0006] 1. Technical issues to be resolved In view of the deficiencies in the prior art, the present invention provides a method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by catalytically using a WO3-based photoenzyme catalyst driven by infrared light. Artificial photosynthesis is used to drive NADPH regeneration, and no other materials are used as hydrogen transfer and electron transfer media. Infrared light is used to drive the photoenzyme catalyst (WO3 / NaCBR) to catalyze the synthesis of (S)-3-chloro-1-phenyl-1-propanol. The present invention has the advantages of simple synthesis method, green environmental protection, high catalytic efficiency, mild reaction conditions, simple operation, easy recovery and recycling, etc., and can greatly reduce costs, and is suitable for large-scale industrial production applications.

[0007] (II) Technical solution In order to achieve the above-mentioned synthesis method is simple, green and environmentally friendly, has high catalytic efficiency, mild reaction conditions, simple operation, easy recovery and recycling, and can significantly reduce costs, suitable for large-scale industrial production applications and the like, the present invention provides the following technical scheme: a method for catalytically synthesizing (S)-3-chloro-1-phenyl-1-propanol by an infrared light-driven WO3-based photoenzyme catalyst, comprising the following steps: Step S1: preparing oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to a certain amount of ethanol in a certain proportion, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 80-120°C for 20-26 h, cooled naturally to room temperature to collect the product, washed with distilled water and anhydrous ethanol several times, and vacuum dried at 50-80°C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 to obtain oxygen-deficient WO3 nanosheets; Step S2: preparing NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 30-37°C until the OD 600 =0.6-0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 0.1-1.0 mM, induce protein expression at 18-25°C for 15-20 hours, centrifuge at 5000-10000 rpm for 5-15 minutes to obtain bacteria of carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 5000-10000 rpm for 10-15 minutes to obtain free enzyme; Step S3: preparing WO3 / NaCBR photoenzyme catalyst; the specific method is: The oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed in a mass ratio of 1:0.5 to 1:2, and the mixture was stirred at 10°C for 3 to 6 hours to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying.

[0008] Step S4: WO3 / NaCBR photoenzyme catalyst catalyzes the synthesis of (S)-3-chloro-1-phenyl-1-propanol; the specific method is: A reaction solvent, a WO3 / NaCBR photoenzyme catalyst and 3-chloro-1-phenyl-1-propanone are added to a reactor, and the reaction is carried out under xenon light illumination for 0.25 to 2.0 hours; the supernatant of the reaction solution is taken, extracted with anhydrous n-hexane, and then heated and decompressed to dryness to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0009] Preferably, the molar ratio of WCl6 to oxalic acid in step S1 is between 1:20 and 1:50.

[0010] Preferably, the molar ratio of NaBH4 and WO3 in step S1 is between 1:1 and 1:4.

[0011] Preferably, the kanamycin content in the LB culture medium in step S2 is 80-120 μg / mL.

[0012] Preferably, the concentration of isopropyl-β-D-thiogalactoside (IPTG) added during protein expression induction in step S2 is 0.1-1.0 mg / mL.

[0013] Preferably, the mass ratio of the oxygen-deficient WO3 nanosheets and the NaCBR carbonyl reductase in step S3 is between 1:0.5 and 1:2.

[0014] Preferably, the reaction solvent described in step S4 is one of isopropanol, methanol, n-hexane, PBS buffer, sodium hydroxide solution, and hydrochloric acid solution.

[0015] Preferably, the illumination reaction time in step S4 is 0.25 to 2.0 h.

[0016] Preferably, the wavelength λ of the xenon lamp in step S4 is greater than 800 nm, but is not limited to this wavelength range.

[0017] Preferably, the concentration of the photoenzyme catalyst WO3 / NaCBR added to the reaction system described in step S4 is 0.5-2.0 mg / mL; the concentration of 3-chloro-1-phenyl-1-propanone is 30-60 mmol / L.

[0018] (III) Beneficial effects Compared with the prior art, the present invention provides a method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst, which has the following beneficial effects: 1. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by catalytic synthesis of (S)-3-chloro-1-phenyl-1-propanol by infrared light-driven WO3-based photoenzyme catalyst is to use artificial photosynthesis, that is, to regenerate NADPH by synthesized WO3 / NaCBR photoenzyme catalyst for catalytic synthesis of (S)-3-chloro-1-phenyl-1-propanol. It has the advantages of simple synthesis method, green environmental protection, high catalytic efficiency, mild reaction conditions, simple operation, easy recovery and recycling, etc., and can greatly reduce costs, and is suitable for large-scale industrial production applications.

[0019] 2. The infrared light-driven WO3-based photoenzyme catalyst catalyzes the synthesis of (S)-3-chloro-1-phenyl-1-propanol. For the first time, the WO3 / NaCBR artificial photoenzyme system realizes the directional conversion of 3-CPP to (S)-CPPO without the assistance of expensive coenzyme factors and precious metal ligands, and the enantiomeric excess (ee) is over 99.9%.

[0020] 3. The WO3 / NaCBR artificial photoenzyme system uses infrared light to catalyze water decomposition to directly provide active hydrogen atoms, and uses the composite structure of the WO3 / NaCBR photoenzyme catalyst to achieve direct transfer of active hydrogen, further improving the efficiency of sunlight utilization without using other materials as hydrogen transfer and electron transfer media.

[0021] 4. The photoenzyme catalyst (WO3 / NaCBR) synthesized by this method is expected to use artificial photosynthesis to drive other reactions, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a transmission electron microscope (TEM) microscopic morphology of the oxygen-deficient WO3 nanosheets prepared in the present invention; Figure 2 The X-ray diffraction diagram of the oxygen-deficient WO3 nanosheets and photoenzyme catalyst (WO3 / NaCBR) prepared in the present invention; Figure 3 This is a scanning electron microscope (SEM) microscopic morphology image of the photoenzyme catalyst (WO3 / NaCBR) prepared in the present invention; Figure 4 The HPLC analysis spectrum of (S)-3-chloro-1-phenyl-1-propanol in the artificial photoenzyme system after the catalysis of the embodiment of the present invention is completed; DETAILED DESCRIPTION

[0023] As mentioned above, in view of the deficiencies of the prior art, the inventors of this case proposed the technical solution of the present invention after long-term research and extensive practice, which is mainly based on at least: the method of the present invention using artificial photosynthesis to regenerate NADPH for photoenzyme-catalyzed synthesis of (S)-3-chloro-1-phenyl-1-propanol, so as to reduce steps, reduce environmental pollution, reduce production costs, improve production safety, and provide a competitive route for industrialization; the method of the present invention using (near) infrared light to drive oxygen-deficient tungsten trioxide (WO3) to decompose water to provide hydrogen and electrons, and does not use other materials as hydrogen transfer and electron transfer media, assisted by efficient and mild enzyme catalytic reactions; the reaction conditions are mild, the operation and separation are simple, easy to recover and recycle, less pollution to the environment, greatly reducing production costs, and suitable for large-scale industrial production applications.

[0024] In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention is further described in detail below in conjunction with the accompanying drawings and examples. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention. In the following examples, unless otherwise specified, the raw materials and equipment used can be purchased from the market or are commonly used in the art, and the methods involved, if not otherwise specified, are all conventional methods in the art.

[0025] Example 1: Preparation of oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to 40 mL of ethanol at a molar ratio of 1:40, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 100 °C for 24 h, and naturally cooled to room temperature to collect the product, which was washed with distilled water and anhydrous ethanol several times and dried in vacuo at 60 °C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 at a molar ratio of NaBH4:WO3=1:1 to obtain oxygen-deficient WO3 nanosheets; Example 2: Preparation of oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to 40 mL of ethanol at a molar ratio of 1:40, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 120 °C for 20 h, cooled naturally to room temperature to collect the product, washed with distilled water and anhydrous ethanol several times, and vacuum dried at 60 °C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 at a molar ratio of NaBH4:WO3=3:4 to obtain oxygen-deficient WO3 nanosheets; Example 3: Preparation of oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to 40 mL of ethanol at a molar ratio of 1:30, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 110 °C for 22 h, cooled naturally to room temperature to collect the product, washed with distilled water and anhydrous ethanol several times, and vacuum dried at 60 °C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 at a molar ratio of NaBH4:WO3=1:2 to obtain oxygen-deficient WO3 nanosheets; Example 4: Preparation of oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to 40 mL of ethanol at a molar ratio of 1:30, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 80 °C for 26 h, cooled naturally to room temperature to collect the product, washed with distilled water and anhydrous ethanol several times, and vacuum dried at 60 °C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 at a molar ratio of NaBH4:WO3=1:4 to obtain oxygen-deficient WO3 nanosheets; Example 5: Preparation of NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 37°C until the OD 600=0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 0.5 mM, induce protein expression at 18°C ​​for 20 hours, centrifuge at 8000 rpm for 10 minutes to obtain bacteria of carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 8000 rpm for 15 minutes to obtain free enzyme; Example 6: Preparation of NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 37°C until the OD 600 =0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 0.5 mM, induce protein expression at 20°C for 20 hours, centrifuge at 8000 rpm for 10 minutes to obtain bacteria of carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 8000 rpm for 15 minutes to obtain free enzyme; Example 7: Preparation of NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 37°C until the OD 600 =0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 1.0 mM, induce protein expression at 23 °C for 18 hours, centrifuge at 8000 rpm for 10 minutes to obtain bacteria with carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 8000 rpm for 15 minutes to obtain free enzyme; Example 8: Preparation of NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 37°C until the OD 600 =0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 1.0 mM, induce protein expression at 25°C for 18 hours, centrifuge at 8000 rpm for 10 minutes to obtain bacteria of carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 8000 rpm for 15 minutes to obtain free enzyme; The WO3 / NaCBR photoenzyme catalyst is prepared by using the oxygen-deficient WO3 nanosheets prepared by any method of Examples 1 to 4 and the NaCBR carbonyl reductase prepared by any method of Examples 5 to 8.

[0026] Example 9: Preparation of WO3 / NaCBR photoenzyme catalyst; the specific method is: Oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed at a mass ratio of 1:0.5, and the mixture was stirred at 10 °C for 3 hours to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying; Example 10: Preparation of WO3 / NaCBR photoenzyme catalyst; the specific method is: The oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed in a mass ratio of 1:1, and the mixture was stirred at 10 °C for 3 hours to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying; Example 11: Preparation of WO3 / NaCBR photoenzyme catalyst; the specific method is: Oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed at a mass ratio of 1:1.5, and the mixture was stirred at 10 °C for 6 hours to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying; Example 12: Preparation of WO3 / NaCBR photoenzyme catalyst; the specific method is: Oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed in a mass ratio of 1:2, and the mixture was stirred at 10 °C for 6 hours to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying; (S)-3-chloro-1-phenyl-1-propanol was synthesized by catalyzing the WO3 / NaCBR photoenzyme catalyst prepared by any method of Examples 9 to 12.

[0027] Example 13: Synthesis of (S)-3-chloro-1-phenyl-1-propanol by WO3 / NaCBR photoenzyme catalyst; the specific method is: Isopropanol solvent, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 0.5 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 30 mmol / L. The reaction was carried out under xenon light for 0.25 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0028] Example 14: Synthesis of (S)-3-chloro-1-phenyl-1-propanol catalyzed by WO3 / NaCBR photoenzyme catalyst; the specific method is: Methanol solvent, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 0.8 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 35 mmol / L. The reaction was carried out under xenon light for 0.5 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0029] Example 15: Synthesis of (S)-3-chloro-1-phenyl-1-propanol by WO3 / NaCBR photoenzyme catalyst; the specific method is: n-Hexane solvent, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 1.0 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 40 mmol / L. The reaction was carried out under xenon light for 1.0 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0030] Example 16: Synthesis of (S)-3-chloro-1-phenyl-1-propanol by WO3 / NaCBR photoenzyme catalyst; the specific method is: PBS buffer, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 1.5 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 50 mmol / L. The reaction was carried out under xenon light for 1.0 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0031] Example 17: Synthesis of (S)-3-chloro-1-phenyl-1-propanol catalyzed by WO3 / NaCBR photoenzyme catalyst; the specific method is: Sodium hydroxide solution, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 1.2 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 45 mmol / L. The reaction was carried out under xenon light illumination for 1.5 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0032] Example 18: Synthesis of (S)-3-chloro-1-phenyl-1-propanol catalyzed by WO3 / NaCBR photoenzyme catalyst; the specific method is: Hydrochloric acid solution, WO3 / NaCBR photoenzyme catalyst, and 3-chloro-1-phenyl-1-propanone were added to the reactor. The concentration of WO3 / NaCBR photoenzyme catalyst in the reaction system was 2.0 mg / mL, and the concentration of 3-chloro-1-phenyl-1-propanone was 60 mmol / L. The reaction was carried out under xenon light for 2.0 h. The supernatant of the reaction solution was taken, extracted with anhydrous n-hexane, heated and decompressed, and dried by rotation to obtain (S)-3-chloro-1-phenyl-1-propanol.

[0033] Embodiment 19: After the reaction, the photoenzyme catalyst (WO3 / NaCBR) was filtered and washed with deionized water and dried for recycling. The cycle lasted for 0.5 h. After 5 cycles, the conversion rate of (S)-3-chloro-1-phenyl-1-propanol detected by HPLC remained basically unchanged.

[0034] Embodiment 20: After the reaction, the photoenzyme catalyst (WO3 / NaCBR) was filtered and washed with deionized water and dried for recycling. The cycle lasted for 1.0 h. After 5 cycles, the conversion rate of (S)-3-chloro-1-phenyl-1-propanol detected by HPLC remained basically unchanged.

[0035] In the case implementation, the morphology of oxygen-deficient tungsten trioxide (WO3) was first studied by transmission electron microscopy (TEM). Figure 1 This is the TEM image of oxygen-deficient WO3 nanosheets. It can be clearly seen that the sample has a good nanosheet structure.

[0036] In the case implementation, the phase and structure information of carbonyl reductase (NaCBR) and photoenzyme catalyst (WO3 / NaCBR) were obtained through X-ray diffraction (XRD) testing. Figure 2As shown. Comparing the peak position and diffraction peak intensity of carbonyl reductase (NaCBR), the peak position of the photoenzyme catalyst (WO3 / NaCBR) sample did not change, indicating that the photoenzyme catalyst (WO3 / NaCBR) formed by the composite of oxygen-deficient WO3 nanosheets and carbonyl reductase (NaCBR) did not change the structure of carbonyl reductase (NaCBR). Moreover, the XRD diffraction peak intensity of the photoenzyme catalyst (WO3 / NaCBR) increased significantly, which was mainly caused by the crystallinity of oxygen-deficient WO3. At the same time, it also proved the successful preparation of the photoenzyme catalyst (WO3 / NaCBR) from the side.

[0037] In the case implementation, the successful preparation of the photoenzyme catalyst (WO3 / NaCBR) was further confirmed by scanning electron microscopy (SEM). Figure 3 As shown, the results show the state of oxygen-deficient WO3 and NaCBR complexed together, and the WO3 / NaCBR complex exhibits an irregular structure, which is consistent with the structure of the precursor carbonyl reductase NaCBR.

[0038] In the case implementation, the content of (S)-3-chloro-1-phenyl-1-propanol in the system was detected by high performance liquid chromatography after the reaction was completed. High performance liquid chromatography conditions: C18 column, column temperature of 25°C, detector of 210nm, mobile phase of methanol: water = 75:25 (v / v), flow rate of 0.4mL / min. (S)-3-chloro-1-phenyl-1-propanol purchased from TCI (Tokyo Chemical Industry Co., Ltd.) was used as a standard, and HPLC analysis was performed with the product prepared by the method of the present invention. The retention times of the two were consistent, and the product structure was determined. The results are as follows. Figure 4 As shown. The conversion rate is calculated by the standard curve of the HPLC peak area of ​​the standard sample (S)-3-chloro-1-phenyl-1-propanol, and the standard curve of the standard sample is y=527.73x+34.29, wherein x represents the product concentration in the reaction solution, in mg / mL, and y represents the HPLC peak area of ​​the product. The highest conversion rate of (S)-3-chloro-1-phenyl-1-propanol detected by HPLC is 32.58%.

[0039] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Although the present invention has been disclosed as a preferred embodiment as above, it is not used to limit the present invention. Any technician familiar with this profession can make some changes or modifications to equivalent embodiments of equivalent changes using the technical contents disclosed above without departing from the scope of the technical solution of the present invention. However, any simple modification, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. A method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst, characterized in that: The following steps are involved: Step S1: preparing oxygen-deficient WO3 nanosheets; the specific method is: Oxalic acid and WCl6 were added to a certain amount of ethanol in a certain proportion, stirred evenly at room temperature, transferred to a Teflon-lined autoclave, sealed and reacted at 80-120°C for 20-26 h, cooled naturally to room temperature to collect the product, washed with distilled water and anhydrous ethanol several times, and vacuum dried at 50-80°C overnight to obtain WO3 nanosheets, and WO3 was reduced with NaBH4 to obtain oxygen-deficient WO3 nanosheets; Step S2: preparing NaCBR carbonyl reductase; the specific method is: The NaCBR carbonyl reductase mutant cells were inoculated into LB medium containing kanamycin and cultured in a shaking incubator at 30-37°C until the OD 600 =0.6-0.8; add isopropyl-β-D-thiogalactoside (IPTG) to a final concentration of 0.1-1.0 mM, induce protein expression at 18-25°C for 15-20 hours, centrifuge at 5000-10000 rpm for 5-15 minutes to obtain bacteria of carbonyl reductase modified with non-natural amino acids; resuspend the bacterial precipitate of aldehyde-keto reductase modified with non-natural amino acids in PBS buffer, treat with ultrasound to lyse the bacteria; centrifuge at 5000-10000 rpm for 10-15 minutes to obtain free enzyme; Step S3: preparing WO3 / NaCBR photoenzyme catalyst; the specific method is: The oxygen-deficient WO3 nanosheets and NaCBR carbonyl reductase were mixed in a mass ratio of 1:0.5 to 1:2, and the mixture was stirred at 10 °C for 3 to 6 h to obtain a mixed material. After centrifugation, the photoenzyme catalyst (WO3 / NaCBR) was collected by freeze drying; Step S4: WO3 / NaCBR photoenzyme catalyst catalyzes the synthesis of (S)-3-chloro-1-phenyl-1-propanol; the specific method is: A reaction solvent, a WO3 / NaCBR photoenzyme catalyst and 3-chloro-1-phenyl-1-propanone are added to a reactor, and the reaction is carried out under xenon light illumination for 0.25 to 2.0 hours; the supernatant of the reaction solution is taken, extracted with anhydrous n-hexane, and then heated and decompressed to dryness to obtain (S)-3-chloro-1-phenyl-1-propanol.

2. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The molar ratio of WCl6 to oxalic acid in step S1 is between 1:20 and 1:

50.

3. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The molar ratio of NaBH4 and WO3 in step S1 is between 1:1 and 1:

4.

4. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The kanamycin content in the LB culture medium described in step S2 is 80-120 μg / mL.

5. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The concentration of isopropyl-β-D-thiogalactoside (IPTG) added during protein expression induction in step S2 is 0.1 to 1.0 mg / mL.

6. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The mass ratio of the oxygen-deficient WO3 nanosheets and the NaCBR carbonyl reductase described in step S3 is 1:5 to 1:

2.

7. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The reaction solvent described in step S4 is one of isopropanol, methanol, n-hexane, PBS buffer, sodium hydroxide solution, and hydrochloric acid solution.

8. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The illumination reaction time in step S4 is 0.25 to 2.0 h.

9. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The wavelength λ of the xenon lamp in step S4 is greater than 800 nm, but is not limited to this wavelength range.

10. The method for synthesizing (S)-3-chloro-1-phenyl-1-propanol by using an infrared light-driven WO3-based photoenzyme catalyst according to claim 1, characterized in that: The concentration of the photoenzyme catalyst WO3 / NaCBR added to the reaction system described in step S4 is 0.5-2.0 mg / mL; the concentration of 3-chloro-1-phenyl-1-propanone is 30-60 mmol / L.

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