Method for synthesizing glufosinate-ammonium from L-ethylglycine

Abstract

The present invention relates to a method for synthesizing glufosinate-ammonium by using L-vinylglycine. Under the protection of nitrogen or helium, diethyl methylphosphite is added dropwise to an acetic acid solution of L-vinylglycine for reaction. After the reaction is completed, the reaction is refluxed in a hydrochloric acid solution, and finally refluxed in an ammonia water solution for 2-8 h. The obtained product is subjected to vacuum distillation and recrystallized from methanol to obtain glufosinate-ammonium crystals. By adding L-vinylglycine to ethyl methylphosphite under acetic acid conditions, followed by acidolysis, ammonia neutralization, and recrystallization from methanol, glufosinate-ammonium with an optical purity greater than 99% and a yield greater than 95% can be obtained, which has broad application prospects.

Description

Technical Field

[0001] The present invention belongs to the fields of fine chemical engineering, genetic engineering and enzyme engineering, and relates to the application of L-vinylglycine in the synthesis of glufosinate-ammonium. Background Art

[0002] The structural formula of L-vinylglycine is shown below. It is a natural, non-protein amino acid, a pyridoxal phosphate (PLP) coenzyme inhibitor, and can also be used as a raw material for the synthesis of the novel herbicide glufosinate-ammonium (L-phosphinothricin). L-vinylglycine derivatives can also be used as preservatives to delay the loss of fruit quality and bioactive components during cold storage and shelf life. Therefore, the low-cost and high-efficiency synthesis of L-vinylglycine is crucial.

[0003] 。

[0004] Currently, the synthesis of L-vinylglycine is mainly by chemical methods, and enzymatic synthesis has not been reported yet. Chemical synthesis has been reported more by foreign scholars. Its synthesis raw materials mainly include methionine and homocysteine. After protecting the amino and carboxyl groups, it is prepared by dehydration, and the total yield is less than 50%. The main process is as follows:

[0005]

[0006] Due to the long synthesis route, low yield, high temperature and hazardous chemicals involved in the chemical synthesis of L-vinylglycine, high requirements for equipment, and difficulty in industrialization. Therefore, the present invention uses a biocatalyst to first prepare its pyruvic acid compound using L-amino acid oxidase or D-amino acid oxidase, then uses dehydratase to prepare the corresponding vinylpyruvic acid, and finally ammoniates and reduces it using L-amino acid dehydrogenase and coenzyme NADH or NADPH to prepare L-vinylglycine. Considering that the final product will have a certain toxicity to amino acid oxidase and dehydratase, a multi-enzyme cascade reaction system will be used in the process. The research group of the present invention has applied for a corresponding patent (Gong Dachun, Wang Delin, Zhang Shuyin, etc., Multi-enzyme cascade reaction separation coupling system and method, CN 113088443 A). Therefore, the method for efficiently synthesizing L-vinylglycine using a multi-enzyme cascade reaction has not been reported yet and has broad application prospects. Summary of the Invention

[0007] To solve the technical problems such as low yield and harsh conditions of the above chemical synthesis, the present invention provides a method for preparing glufosinate-ammonium from L-vinylglycine.

[0008] The method for synthesizing glufosinate-ammonium using L-vinylglycine includes the following steps:

[0009] Under the protection of nitrogen or helium, diethyl methylphosphite was added dropwise to an acetic acid solution of L-vinylglycine for reaction. After the reaction was completed, the mixture was refluxed in a hydrochloric acid solution and finally refluxed in an ammonia water solution. The obtained product was subjected to vacuum distillation and recrystallization from methanol to obtain the glufosinate-ammonium crystals.

[0010] In the acetic acid solution, the reaction was carried out at a temperature of 10 - 30 °C for 2 - 10 h.

[0011] The mass ratio of L-vinylglycine to diethyl methylphosphite was 1:1 - 3.0.

[0012] The concentration of the hydrochloric acid solution was 1% - 30%. After adjusting the pH of the system to 1 - 6, the mixture was stirred and reacted at a temperature of 0 - 80 °C for 2 - 10 h.

[0013] The concentration of the ammonia water solution was 1% - 30%. After adjusting the pH of the system to 6.8 - 7.8, at a temperature of

[0014] 0 - 80 °C, the mixture was stirred and reacted for 2 - 10 h.

[0015] The synthetic route of L-vinylglycine described in the present invention is as follows:

[0016]

[0017] The main method of the present invention is as shown above. Using threonine as the raw material, biological catalysts such as L-amino acid oxidase and D-amino acid oxidase were selectively added according to the configuration of the substrate. At 20 - 35 °C and a reaction pH of 5.5 - 7.5, 3-substituted γ-substituted hydroxy (mercapto) or β-substituted hydroxy pyruvic acid was prepared, and the substrate could be added in batches or all at once; then under the action of lactate dehydrogenase or phenyl lactate dehydrogenase, at a reaction temperature of 15 - 35 °C and a reaction pH of 4.5 - 7, vinylpyruvic acid was prepared; then using L-glutamate dehydrogenase and glucose dehydrogenase, at a reaction temperature of 25 - 55 °C and a reaction pH of 6.5 - 8.5, L-vinylglycine was prepared (the preparation method of L-vinylglycine comes from another patent applied on the same day: A method for preparing L-vinylglycine by multi-enzyme cascade), and the substrate could be added in batches or all at once. The whole reaction was carried out by continuous production in a tubular reactor or batch preparation in a batch reactor. Through adsorption and elution by a hydrogen-type 001x7 cation exchange resin, high-purity L-vinylglycine was obtained, and then it was reacted with diethyl methylphosphite under the condition of 10 - 30 °C for the preparation of glufosinate-ammonium.

[0018] The present invention reacts L-vinylglycine with ethyl methylphosphonate under acetic acid conditions, and after acidolysis, ammonia neutralization, and methanol recrystallization, the optically pure glufosinate-ammonium with an optical purity greater than 99% and a yield greater than 95% can be obtained, having broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 is the HNMR spectrum of L-vinylglycine 1 HNMR spectrum.

[0020] Figure 2 is the 1 HNMR spectrum of L-glufosinate-ammonium. DETAILED DESCRIPTION OF THE INVENTION

[0021] The present invention will be further described below in conjunction with the embodiments, but the scope of protection required by the present invention is not limited to the scope described in the embodiments.

[0022] Example 1 Preparation of Glufosinate-ammonium (L-Glufosinate-ammonium)

[0023] In a 500 L batch reactor, under the condition of 15 °C and protected by nitrogen or helium, an acetic acid solution of L-vinylglycine was added, and diethyl methylphosphonate was added dropwise. The mass ratio of L-vinylglycine to diethyl methylphosphonate was 1.01:1.36, and the mass concentration of the acetic acid solution of L-vinylglycine was 40%. The reaction was carried out for 6 hours, then the pH was adjusted to 3.5 with a 10% hydrochloric acid solution, heated to 50 °C, and refluxed for 5 hours. Finally, a 20% ammonia aqueous solution was added to adjust the pH to 6.8, and refluxed at 50 °C for 6 hours. After vacuum distillation, it was recrystallized with methanol to obtain white glufosinate-ammonium crystals, 188.1 kg was obtained, the yield was 95%, and the optical purity of the product was 99.6%. After NMR detection, it was 1 HNMR (300 MHz, D2O) δ: 1.12 (d, J = 13.50 Hz, 3H); 1.86 - 1.94 (m, 2H); 1.34 - 1.58 (m, 2H); 3.64 (t, J = 5.9 Hz, 1H) (see attachment Figure 2 ). After liquid phase detection, the retention time of L-glufosinate-ammonium was: 7.010 min (mobile phase; 50 mM ammonium acetate buffer solution (Ph8.0): methanol = 9:1 flow rate; 0.8 mL / min, chromatographic column; Acclaim TM 120 C18 5um 120A 4.6×150mm, detection wavelength; 350 nm column temperature: 35 °C, high performance liquid chromatograph model; ThermoUItiMate3000).

[0024] Example 2 Preparation of Glufosinate-ammonium (L-Glufosinate-ammonium)

[0025] In a 500 L batch reactor, under the condition of 15 °C and protected by nitrogen or helium, an acetic acid solution of L-vinylglycine was added, and diethyl methylphosphite was added dropwise. The mass ratio of L-vinylglycine to diethyl methylphosphite was 1.01:2.72, and the mass concentration of the acetic acid solution of L-vinylglycine was 35%. The reaction was carried out for 6 hours, then the pH was adjusted to 3.2 with a hydrochloric acid solution with a mass concentration of 10%, heated to 50 °C, and refluxed for 5 hours. Finally, a 20% ammonia water solution was added to adjust the pH to 7.2, and refluxed at 50 °C for 6 hours. After vacuum distillation, recrystallization was carried out with methanol to obtain white glufosinate-ammonium crystals, with a yield of 95.6% and an optical purity of the product of 99.6%. After NMR detection, it was 1H NMR (300 MHz, D2O) δ: 1.12 (d, J = 13.50 Hz, 3H); 1.86 - 1.94 (m, 2H); 1.34 - 1.58 (m, 2H); 3.64 (t, J = 5.9 Hz, 1H). mp 208.5 - 211 °C, [α] D 19.5 = + 16.02 °C (c = 0.70, H2O), ee > 99.6%.

[0026] The above examples only represent the preferred embodiments of the present invention, and the description is relatively specific and detailed, but it should not be construed as a limitation on the scope of the present invention patent. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications, improvements and substitutions can be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the present invention patent shall be subject to the appended claims.

Claims

1. A method for synthesizing glufosinate-ammonium from L-ethylglycine, characterized in that, It includes the following steps: In a 500 L batch reactor, under the condition of 15 °C and protected by nitrogen or helium, an acetic acid solution of L-vinylglycine is added, and diethyl methylphosphonate is added dropwise. The mass ratio of L-vinylglycine to diethyl methylphosphonate is 1.01:1.36, and the mass concentration of the acetic acid solution of L-vinylglycine is 40%. The reaction is carried out for 6 hours, and then the pH is adjusted to 3.5 with a 10% hydrochloric acid solution, heated to 50 °C, and refluxed for 5 hours. Finally, a 20% ammonia water solution is added to adjust the pH to 6.8, and refluxed at 50 °C for 6 hours. After vacuum distillation, recrystallization is carried out with methanol to obtain white glufosinate-ammonium crystals, obtaining 188.1 kg, with a yield of 95% and an optical purity of the product of 99.6%.

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