Method for preparing 1, 3, 4-oxadiazine compound through catalysis of oxidase

Abstract

The invention provides a method for preparing a 1, 3, 4-oxadiazine compound through catalysis of oxidase, and relates to the technical field of organic synthesis. 4-aminophenol and a hydrazone compound are subjected to an intermolecular cycloaddition reaction in the presence of a buffer solution, oxidase and a peroxide catalyst, and the 1, 3, 4-oxadiazine compound is obtained. 4-aminophenol and a hydrazone compound which are low in price and easy to obtain are used as raw materials, a buffer solution is used as a reaction solvent, oxidase is used as a biocatalyst, peroxide is cooperated, and intermolecular cycloaddition reaction of the 4-aminophenol and the hydrazone compound is catalyzed to synthesize the 1, 3, 4-oxadiazine compound. The method uses a biological catalysis method, does not need to react under harsh conditions such as strong acid, strong alkali or a strong reducing agent, is mild in reaction condition, few in generated by-products, high in product yield, high in atom economy, simple to separate and purify, low in production cost, simple and convenient to operate and pollution-free, meets the requirements of green chemical development, and is suitable for industrial production.

Description

Technical Field

[0001] The invention relates to the technical field of organic synthesis, and in particular to a method for preparing 1,3,4-oxadiazine compounds by oxidase catalysis. Background Art

[0002] 1,3,4-oxadiazines are a class of oxadiazines with a unique six-membered ring skeleton structure consisting of two N atoms and one O atom. They are widely found in medicines, agricultural chemicals and many biologically active molecules, such as the Bulgarian chemotherapy drug Oxadin (trade name) with antiviral activity against herpes simplex virus (HSV), and Indoxacarb, an oxadiazine pesticide developed by DuPont that can act on the larvae of lepidopteran insects.

[0003]

[0004] At present, there are mainly the following methods for synthesizing 1,3,4-oxadiazine structures: (1) β-hydroxylamine is reacted with sodium nitrite and hydrochloric acid to obtain nitrosyl, and then LiAlH4 and imidazolone are used in turn to obtain 1,3,4-oxadiazine compounds through cycloaddition reaction; (2) p-hydroxyquinone and azomethine imine are used as raw materials to prepare 1,3,4-oxadiazine through [3+3] cycloaddition reaction under the condition of potassium tert-butoxide strong base reagent; (3) Azomethine imine and cumulative diene are used as raw materials to obtain the target product 1,3,4-oxadiazine through cycloaddition reaction under the conditions of sodium nitrite / nitric acid / oxygen and 4-dimethylaminopyridine. Although the above synthetic methods can successfully prepare 1,3,4-oxadiazine compounds, they need to be completed under harsh conditions such as strong acid, strong base or strong reducing agent, and the reaction is accompanied by the production of a large amount of by-products, and the product yield is low. Summary of the invention

[0005] In view of this, the object of the present invention is to provide a method for preparing 1,3,4-oxadiazines by oxidase catalysis. The method provided by the present invention uses oxidase as a biocatalyst to prepare 1,3,4-oxadiazines, with mild preparation conditions, high atom economy and high product yield.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The invention provides a method for preparing 1,3,4-oxadiazine compounds by oxidase catalysis, comprising the following steps: mixing 4-aminophenol, a hydrazone compound, a catalyst and a buffer solution, and performing an intermolecular cycloaddition reaction to obtain the 1,3,4-oxadiazine compounds; the catalyst comprises an oxidase and a peroxide.

[0008] Preferably, the mass ratio of 4-aminophenol to oxidase is 0.05-0.5:1.

[0009] Preferably, the oxidase comprises one or more of horseradish peroxidase, peroxidase and haloperoxidase.

[0010] Preferably, the peroxidase comprises a peroxidase derived from fungi.

[0011] Preferably, the oxidase is used in the form of one or more of whole fungal cells, crude enzyme powder, crude enzyme solution, pure enzyme and immobilized enzyme.

[0012] Preferably, the peroxide comprises an inorganic peroxide and / or an organic peroxide;

[0013] Preferably, the peroxide concentration is 100-300 mM; when 1 mL of buffer solution is added, the addition rate of the peroxide solution is 10-30 μL / h; and the peroxide solution is continuously added during the intermolecular cycloaddition reaction.

[0014] Preferably, the hydrazone compound includes hydrazone or halogenated hydrazone; the molar ratio of 4-aminophenol to the hydrazone compound is 1:1.2-2;

[0015] The volume ratio of the amount of 4-aminophenol to the buffer solution is 1 mmol: 100-200 mL.

[0016] Preferably, the buffer solution includes a citrate buffer solution or a phosphate buffer solution; the pH value of the buffer solution is 4-9.

[0017] Preferably, the 4-aminophenol is used in the form of a 4-aminophenol solution, and the hydrazone compound is used in the form of a hydrazone compound solution; the solvents in the 4-aminophenol solution and the hydrazone compound solution independently include one or more of dimethyl sulfoxide, ethyl acetate, acetone and isopropanol.

[0018] Preferably, the temperature of the intermolecular cycloaddition reaction is 25 to 40° C., and the time is 5 to 24 hours.

[0019] The present invention provides a method for preparing 1,3,4-oxadiazine compounds by oxidase catalysis, comprising the following steps: mixing 4-aminophenol, hydrazone compounds, catalysts and buffer solutions, and performing intermolecular cycloaddition reactions to obtain 1,3,4-oxadiazine compounds; the catalysts include oxidase and peroxide. The present invention uses cheap and readily available 4-aminophenol and hydrazone compounds as raw materials, a buffer solution as a reaction solvent, an oxidase as a biocatalyst, and peroxide as an auxiliary catalyst to catalyze the intermolecular cycloaddition reactions of 4-aminophenol and hydrazone compounds to synthesize 1,3,4-oxadiazine compounds. The biocatalytic method used in the present invention does not need to react under harsh conditions such as strong acids, strong bases or strong reducing agents, has mild reaction conditions, produces few by-products, has a high product yield, has high atom economy, is simple to separate and purify, has low production costs, is easy to operate, is pollution-free, meets the requirements of green chemical development, and is suitable for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The hydrogen spectrum of 1,3,4-oxadiazine prepared in Example 1;

[0021] Figure 2 This is a high performance liquid chromatogram of 1,3,4-oxadiazine prepared in Example 1;

[0022] Figure 3 This is the hydrogen spectrum of 3-fluoro-1,3,4-oxadiazine prepared in Example 2;

[0023] Figure 4 This is the HPLC chromatogram of 3-fluoro-1,3,4-oxadiazine prepared in Example 2;

[0024] Figure 5 This is the hydrogen spectrum of 4-chloro-1,3,4-oxadiazine prepared in Example 3;

[0025] Figure 6 This is the HPLC chromatogram of 4-chloro-1,3,4-oxadiazine prepared in Example 3;

[0026] Figure 7 This is the hydrogen spectrum of 3-bromo-1,3,4-oxadiazine prepared in Example 4;

[0027] Figure 8 This is the high performance liquid chromatogram of 3-bromo-1,3,4-oxadiazine prepared in Example 4. DETAILED DESCRIPTION

[0028] The invention provides a method for preparing 1,3,4-oxadiazine compounds by oxidase catalysis, comprising the following steps: mixing 4-aminophenol, a hydrazone compound, a catalyst and a buffer solution, and performing an intermolecular cycloaddition reaction to obtain the 1,3,4-oxadiazine compounds; the catalyst comprises an oxidase and a peroxide.

[0029] Unless otherwise specified, the materials and equipment used in the present invention are all commercially available products in the art.

[0030] In the present invention, the 4-aminophenol is preferably used in the form of a 4-aminophenol solution, the concentration of the 4-aminophenol solution is preferably 5 to 50 mM (mmol / L), more preferably 5 to 20 mM, and in a specific embodiment can be 5 mM, 10 mM, 15 mM, 20 mM, 25 mM, 30 mM, 35 mM, 40 mM, 45 mM or 50 mM; the solvent in the 4-aminophenol solution preferably includes one or more of dimethyl sulfoxide, ethyl acetate, acetone and isopropanol.

[0031] In the present invention, the hydrazone compound preferably includes a hydrazone or a halogenated hydrazone; the halogenated hydrazone preferably includes one or more of a fluorohydrazone, a chlorohydrazone and a bromohydrazone, and more preferably includes one or more of a 3-fluorohydrazone, a 4-chlorohydrazone and a 3-bromohydrazone. In the present invention, the hydrazone compound is preferably used in the form of a hydrazone compound solution; the concentration of the hydrazone compound solution is preferably 10 to 100 mM, more preferably 30 to 80 mM, and further preferably 50 mM; the solvent in the hydrazone compound solution preferably includes one or more of dimethyl sulfoxide, ethyl acetate, acetone and isopropanol.

[0032]

[0033] In the present invention, when the hydrazone compound is a hydrazone, the prepared 1,3,4-oxadiazine compound is 1,3,4-oxadiazine; when the hydrazone compound is a halogenated hydrazone, the prepared 1,3,4-oxadiazine compound is a halogenated 1,3,4-oxadiazine; the halogenated 1,3,4-oxadiazine includes one or more of fluoro-1,3,4-oxadiazine, chloro-1,3,4-oxadiazine and brominated 1,3,4-oxadiazine, more preferably includes one or more of 3-fluoro-1,3,4-oxadiazine, 4-chloro-1,3,4-oxadiazine and 3-bromo-1,3,4-oxadiazine, and the structural formula is as follows:

[0034]

[0035] In the present invention, the molar ratio of 4-aminophenol to hydrazone compound is preferably 1:1.2-2, more preferably 1:1.2-1.5, and further preferably 1:1.2-1.3.

[0036] In the present invention, the oxidase preferably includes one or more of horseradish peroxidase (GenBank: AAA72223.1), peroxidase and haloperoxidase; the peroxidase preferably includes peroxidase derived from fungi; the fungi preferably include one or more of Marasmius rotula, Coprinopsis cinerea, Myceliophthorathermophila and Coprinellus radians; the oxidase specifically preferably includes horseradish peroxidase (HRP, GenBank: AAA72223.1) derived from horseradish (Armoracia rusticana), peroxidase derived from Marasmius rotula peroxidase (MroUPO, GenBank: JQ654436.1), Coprinellusradians peroxidase (CraUPO, GenBank: FM872459.1) and Caldariomycesfumago haloperoxidase (CfCPO, GenBank: GCA001660795.1) one or more. In the present invention, the oxidase is preferably used in the form of one or more of fungal whole cells, crude enzyme powder, crude enzyme liquid, pure enzyme and immobilized enzyme; the fungi in the fungal whole cells preferably include one or more of Marasmius rotula, Coprinopsis cinerea, Myceliophthorathermophila and Coprinellus radians. The present invention has no particular limitation on the preparation method of the fungal whole cell, crude enzyme powder, crude enzyme solution, pure enzyme and immobilized enzyme, and the preparation method of the fungal whole cell, crude enzyme powder, crude enzyme solution, pure enzyme and immobilized enzyme containing oxidase known to those skilled in the art can be used. The oxidase used in the present invention has the activity of oxidizing phenolic substrates, and can catalyze the intermolecular cycloaddition reaction of 4-aminophenol and hydrazone compounds under mild conditions to generate 1,3,4-oxadiazine compounds.

[0037] In the present invention, the concentration of the oxidase is preferably 1 to 2 mg / mL, more preferably 1 to 1.5 mg / mL, and even more preferably 1 to 1.2 mg / mL.

[0038] In the present invention, the mass ratio of 4-aminophenol to oxidase is preferably 0.05 to 0.5:1, more preferably 0.1 to 0.4:1, and further preferably 0.2 to 0.3:1.

[0039] In the present invention, the peroxide preferably includes inorganic peroxide and / or organic peroxide; the inorganic peroxide preferably includes hydrogen peroxide; the organic peroxide preferably includes one or more of tert-butyl hydroperoxide, benzoyl peroxide, methyl ethyl ketone peroxide, urea hydrogen peroxide, triacetone triperoxide, ethylene glycol dimethyl ether peroxide, benzoic acid peroxide, peroxyacetyl nitrate, ether peroxide, peracetic acid and cumene peroxide.

[0040] In the present invention, the hydrogen peroxide is preferably used in the form of a peroxide solution, and the peroxide solution includes an inorganic peroxide solution and / or an organic peroxide solution; the concentration of the peroxide solution is preferably 100-300mM, more preferably 150-250mM, and in a specific embodiment, it can be 100mM, 110mM, 120mM, 130mM, 140mM, 150mM, 160mM, 170mM, 180mM, 190mM, 200mM, 210mM, 220mM, 230mM, 240mM, 250mM, 260mM, 270mM, 280mM, 290mM or 3000mM. In the present invention, the solvent in the inorganic peroxide solution is preferably water; the solvent in the organic peroxide solution preferably includes one or more of ethyl acetate, acetone and isopropanol. In the present invention, the peroxide solution is preferably added to the reaction system via a syringe, and the peroxide is continuously added during the intermolecular cycloaddition reaction.

[0041] In the present invention, when 1 mL of buffer solution is added, the addition rate of the peroxide solution is preferably 10 to 30 μL / h, more preferably 20 μL / h, and in a specific embodiment can be 10 μL / h, 15 μL / h, 20 μL / h, 25 μL / h or 30 μL / h; the peroxide solution is continuously added during the intermolecular cycloaddition reaction, that is, the addition time of the peroxide solution is the same as the time of the intermolecular cycloaddition reaction.

[0042] The invention can add peroxide to cooperate with oxidase to catalyze the intermolecular cycloaddition reaction between 4-aminophenol and hydrazone compounds to generate 1,3,4-oxadiazine compounds.

[0043] In the present invention, the buffer solution preferably includes a citrate buffer solution or a phosphate buffer solution; the pH value of the buffer solution is preferably 4 to 9, more preferably 5 to 8, and specifically preferably 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5 or 9. In the present invention, the citrate buffer solution preferably includes citric acid, sodium citrate and water, and the sodium citrate preferably includes trisodium citrate and / or disodium citrate; the concentration of sodium citrate in the citrate buffer solution is preferably 50 to 100 mM, more preferably 80 to 100 mM. In the present invention, the phosphate buffer solution preferably includes disodium hydrogen phosphate and / or sodium dihydrogen phosphate; the concentration of sodium dihydrogen phosphate in the phosphate buffer solution is preferably 50 to 100 mM, more preferably 80 to 100 mM.

[0044] In the present invention, the ratio of the amount of 4-aminophenol to the volume of the buffer solution is preferably 1 mmol: 100-200 mL, more preferably 1 mmol: 100-150 mL.

[0045] In the present invention, the temperature of the intermolecular cycloaddition reaction is preferably 25 to 40°C, more preferably 30 to 35°C; the intermolecular cycloaddition reaction time is preferably 5 to 24h, more preferably 10 to 24h, and further preferably 15 to 24h. In the present invention, taking the synthesis of 1,3,4-oxadiazine from 4-aminophenol and hydrazone as an example, the reactions occurring during the intermolecular cycloaddition reaction are as follows:

[0046]

[0047] After the intermolecular cycloaddition reaction is completed, the present invention preferably further comprises: extracting the reaction solution obtained from the intermolecular cycloaddition reaction, and sequentially drying, concentrating and purifying the obtained organic phase with a desiccant to obtain 1,3,4-oxadiazine compounds.

[0048] In the present invention, the extractant used in the extraction preferably includes one or more of dichloromethane, ethyl acetate and methyl tert-butyl ether.

[0049] In the present invention, the desiccant preferably includes anhydrous sodium sulfate and / or anhydrous magnesium sulfate.

[0050] The present invention has no particular limitation on the concentration, and the solvent can be removed by a concentration method well known to those skilled in the art, such as reduced pressure distillation.

[0051] In the present invention, the purification preferably includes thin layer chromatography purification, and the developing agent used in the thin layer chromatography purification preferably includes a mixed solvent including a polar solvent and petroleum ether, and the polar solvent preferably includes one or more of ethyl acetate, dichloromethane and methanol; the volume ratio of the polar solvent to petroleum ether is preferably 1 to 3:5, more preferably 1.5 to 2.5:5, and further preferably 2:5.

[0052] To further illustrate the present invention, the method for preparing 1,3,4-oxadiazine compounds catalyzed by oxidase provided by the present invention is described in detail below in conjunction with examples, but they should not be construed as limiting the scope of protection of the present invention.

[0053] Example 1

[0054] Preparation of 1,3,4-oxadiazine

[0055] Add 50mL phosphate buffer solution (pH=8.0, NaH2PO4 concentration of 100mM), 50μL 4-aminophenol dimethyl sulfoxide solution with a concentration of 5mM, 60μL hydrazone dimethyl sulfoxide solution with a concentration of 5mM, and then add 2mL HRP enzyme (enzyme concentration of 1mg / mL). Use a syringe pump to add a 200mM H2O2 aqueous solution at a rate of 500μL / h to mix, react in a 30℃ constant temperature vibration metal bath for 24h, add ethyl acetate to the resulting reaction solution for extraction, dry the resulting organic phase with anhydrous sodium sulfate, distill under reduced pressure to remove the organic solvent, and then separate by thin layer chromatography (developing solvent is ethyl acetate and petroleum ether with a volume ratio of 1:3), and obtain 1,3,4-oxadiazine (yield of 87%, HPLC purity of 94%). Among them, the volume ratio of the reaction solution to ethyl acetate is 1:2, and the addition time of the H2O2 aqueous solution is the same as the reaction time, both of which are 24h.

[0056] Figure 1 is the hydrogen spectrum of 1,3,4-oxadiazine (the solvent is deuterated chloroform), Figure 2 1,3,4-oxadiazine is a high performance liquid chromatogram. It can be seen that the target product with high purity is prepared in this embodiment.

[0057] Example 2

[0058] Preparation of 3-fluoro-1,3,4-oxadiazine:

[0059] Add 50mL phosphate buffer solution (pH=8.0, NaH2PO4 concentration of 100mM), 50μL 4-aminophenol dimethyl sulfoxide solution with a concentration of 50M, 60μL fluorohydrazone dimethyl sulfoxide solution with a concentration of 5mM, and then add 2mL HRP enzyme (enzyme concentration of 1mg / mL). During the reaction, add 200mM H2O2 aqueous solution at a rate of 500μL / h using a syringe pump to mix, react in a 30℃ constant temperature vibration metal bath for 24h, add ethyl acetate to the obtained reaction solution for extraction, dry the obtained organic phase with anhydrous sodium sulfate, remove the organic solvent by vacuum distillation, and then separate by thin layer chromatography (developing solvent is ethyl acetate and petroleum ether with a volume ratio of 1:3), and analyze by high performance liquid chromatography to obtain 3-fluoro 1,3,4-oxadiazine (yield is 80%, HPLC purity is 93%). The volume ratio of the reaction solution to ethyl acetate was 1:2, and the addition time of the H2O2 aqueous solution was the same as the reaction time, both of which were 24 h.

[0060] Figure 3 is the hydrogen spectrum of 3-fluoro-1,3,4-oxadiazine (the solvent is deuterated chloroform), Figure 4 3-fluoro-1,3,4-oxadiazine is a high performance liquid chromatogram. It can be seen that the target product with high purity is prepared in this example.

[0061] Example 3

[0062] Preparation of 4-chloro-1,3,4-oxadiazine:

[0063] Add 50mL phosphate buffer solution (pH=8.0, NaH2PO4 concentration of 100mM), 50μL 4-aminophenol dimethyl sulfoxide solution with a concentration of 5mM, 60μL chlorohydrazone dimethyl sulfoxide solution with a concentration of 5mM, and then add 2mL HRP enzyme (enzyme concentration is 1mg / mL). Use a syringe pump to add a 200mM H2O2 aqueous solution at a rate of 500μL / h to mix, react in a 30℃ constant temperature vibration metal bath for 24h, add ethyl acetate to the obtained reaction solution for extraction, dry the obtained organic phase with anhydrous sodium sulfate, remove the organic solvent by reduced pressure distillation, and then separate by thin layer chromatography (developing solvent is ethyl acetate and petroleum ether with a volume ratio of 1:3), and analyze by high performance liquid chromatography to obtain 4-chloro-1,3,4-oxadiazine (yield is 80%, HPLC purity is 91%). The volume ratio of the reaction solution to ethyl acetate was 1:2, and the addition time of the H2O2 aqueous solution was the same as the reaction time, both of which were 24 h.

[0064] Figure 5 is the hydrogen spectrum of 4-chloro-1,3,4-oxadiazine (the solvent is deuterated chloroform), Figure 64-chloro-1,3,4-oxadiazine. It can be seen that the target product with high purity is prepared in this example.

[0065] Example 4

[0066] Preparation of 3-bromo-1,3,4-oxadiazine:

[0067] Add 50mL phosphate buffer solution (pH=8.0, NaH2PO4 concentration of 100mM), 50μL 50mM 4-aminophenol dimethyl sulfoxide solution, 60μL 50mM bromohydrazone dimethyl sulfoxide solution, and then add 2mL HRP enzyme (enzyme concentration of 1mg / mL). Use a syringe pump to add a 200mM H2O2 aqueous solution at a rate of 500μL / h to mix, react in a 30℃ constant temperature vibration metal bath for 24h, add ethyl acetate to the obtained reaction solution for extraction, dry the obtained organic phase with anhydrous sodium sulfate, remove the organic solvent by reduced pressure distillation, and then separate by thin layer chromatography (developing solvent is ethyl acetate and petroleum ether with a volume ratio of 1:3), and analyze by high performance liquid chromatography to obtain 3-bromo-1,3,4-oxadiazine (yield is 83%, HPLC purity is 92%). The volume ratio of the reaction solution to ethyl acetate was 1:2, and the addition time of the H2O2 aqueous solution was the same as the reaction time, both of which were 24 h.

[0068] Figure 7 is the hydrogen spectrum of 3-bromo-1,3,4-oxadiazine (the solvent is deuterated chloroform), Figure 8 3-bromo-1,3,4-oxadiazine is shown in the HPLC chromatogram. It can be seen that the target product with high purity is prepared in this example.

[0069] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for preparing 1,3,4-oxadiazine compounds by oxidase catalysis, characterized in that: The following steps are involved: 4-aminophenol, hydrazone compounds, catalysts and buffer solutions are mixed to carry out intermolecular cycloaddition reaction to obtain 1,3,4-oxadiazine compounds; the catalysts include oxidase and peroxide.

2. The method according to claim 1, characterized in that The mass ratio of the 4-aminophenol to the oxidase is 0.05-0.5:

1.

3. The method according to claim 1 or 2, characterized in that: The oxidase includes one or more of horseradish peroxidase, peroxidase and haloperoxidase.

4. The method according to claim 3, characterized in that The peroxidase includes peroxidase derived from fungi.

5. The method according to claim 3, characterized in that: The oxidase is used in the form of one or more of fungal whole cells, crude enzyme powder, crude enzyme liquid, pure enzyme and immobilized enzyme.

6. The method according to claim 1, characterized in that The peroxide includes inorganic peroxide and / or organic peroxide; The peroxide is used in the form of a peroxide solution, the concentration of which is 100-300 mM; when 1 mL of buffer solution is added, the addition rate of the peroxide solution is 10-30 μL / h; and the peroxide solution is continuously added during the intermolecular cycloaddition reaction.

7. The method according to claim 1, characterized in that The hydrazone compound includes hydrazone or halogenated hydrazone; the molar ratio of 4-aminophenol to the hydrazone compound is 1:1.2-2; The volume ratio of the amount of 4-aminophenol to the buffer solution is 1 mmol: 100-200 mL.

8. The method according to claim 1, characterized in that The buffer solution includes a citrate buffer solution or a phosphate buffer solution; the pH value of the buffer solution is 4-9.

9. The method according to claim 1, 2, 6 or 7, characterized in that: The 4-aminophenol is used in the form of a 4-aminophenol solution, and the hydrazone compound is used in the form of a hydrazone compound solution; the solvents in the 4-aminophenol solution and the hydrazone compound solution independently include one or more of dimethyl sulfoxide, ethyl acetate, acetone and isopropanol.

10. The method according to any one of claims 1 to 2 and 4 to 8, characterized in that The temperature of the intermolecular cycloaddition reaction is 25-40° C. and the time is 5-24 hours.

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