CN114277080B - Process for preparing L-glufosinate-ammonium by enzyme resolution method - Google Patents
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- 238000000034 method Methods 0.000 title claims abstract description 37
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 35
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 35
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 19
- CRZQGDNQQAALAY-UHFFFAOYSA-N Methyl benzeneacetate Chemical compound COC(=O)CC1=CC=CC=C1 CRZQGDNQQAALAY-UHFFFAOYSA-N 0.000 claims abstract description 40
- 239000005561 Glufosinate Substances 0.000 claims abstract description 33
- 239000002253 acid Substances 0.000 claims abstract description 29
- IAJOBQBIJHVGMQ-BYPYZUCNSA-N glufosinate-P Chemical compound CP(O)(=O)CC[C@H](N)C(O)=O IAJOBQBIJHVGMQ-BYPYZUCNSA-N 0.000 claims abstract description 28
- 238000002425 crystallisation Methods 0.000 claims abstract description 15
- 230000008025 crystallization Effects 0.000 claims abstract description 15
- IAJOBQBIJHVGMQ-SCSAIBSYSA-N (2R)-glufosinate Chemical compound C[P@@](O)(=O)CC[C@@H](N)C(O)=O IAJOBQBIJHVGMQ-SCSAIBSYSA-N 0.000 claims abstract description 10
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 63
- 238000006243 chemical reaction Methods 0.000 claims description 48
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 claims description 42
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 41
- 239000000243 solution Substances 0.000 claims description 35
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 26
- 238000001914 filtration Methods 0.000 claims description 25
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 24
- 108010073038 Penicillin Amidase Proteins 0.000 claims description 22
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 21
- 239000003960 organic solvent Substances 0.000 claims description 15
- WLJVXDMOQOGPHL-UHFFFAOYSA-N phenylacetic acid Chemical compound OC(=O)CC1=CC=CC=C1 WLJVXDMOQOGPHL-UHFFFAOYSA-N 0.000 claims description 14
- 238000000605 extraction Methods 0.000 claims description 11
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 9
- 230000001105 regulatory effect Effects 0.000 claims description 9
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 8
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 8
- 230000035484 reaction time Effects 0.000 claims description 8
- 238000010992 reflux Methods 0.000 claims description 8
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 7
- 235000011114 ammonium hydroxide Nutrition 0.000 claims description 7
- 238000001704 evaporation Methods 0.000 claims description 7
- 238000010438 heat treatment Methods 0.000 claims description 7
- SJWFXCIHNDVPSH-UHFFFAOYSA-N octan-2-ol Chemical compound CCCCCCC(C)O SJWFXCIHNDVPSH-UHFFFAOYSA-N 0.000 claims description 7
- 229960003424 phenylacetic acid Drugs 0.000 claims description 7
- 239000003279 phenylacetic acid Substances 0.000 claims description 7
- 239000000725 suspension Substances 0.000 claims description 7
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical compound CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 claims description 6
- 238000011033 desalting Methods 0.000 claims description 6
- ZXEKIIBDNHEJCQ-UHFFFAOYSA-N isobutanol Chemical compound CC(C)CO ZXEKIIBDNHEJCQ-UHFFFAOYSA-N 0.000 claims description 6
- 239000012046 mixed solvent Substances 0.000 claims description 6
- BWDBEAQIHAEVLV-UHFFFAOYSA-N 6-methylheptan-1-ol Chemical compound CC(C)CCCCCO BWDBEAQIHAEVLV-UHFFFAOYSA-N 0.000 claims description 4
- 230000006340 racemization Effects 0.000 claims description 4
- 239000002904 solvent Substances 0.000 claims description 4
- 239000008096 xylene Substances 0.000 claims description 4
- 239000012452 mother liquor Substances 0.000 claims description 3
- 238000010790 dilution Methods 0.000 claims description 2
- 239000012895 dilution Substances 0.000 claims description 2
- 239000011259 mixed solution Substances 0.000 claims description 2
- 238000001953 recrystallisation Methods 0.000 claims description 2
- DAJSVUQLFFJUSX-UHFFFAOYSA-M sodium;dodecane-1-sulfonate Chemical compound [Na+].CCCCCCCCCCCCS([O-])(=O)=O DAJSVUQLFFJUSX-UHFFFAOYSA-M 0.000 claims 2
- IAJOBQBIJHVGMQ-UHFFFAOYSA-N 2-amino-4-[hydroxy(methyl)phosphoryl]butanoic acid Chemical class CP(O)(=O)CCC(N)C(O)=O IAJOBQBIJHVGMQ-UHFFFAOYSA-N 0.000 abstract description 9
- 239000003513 alkali Substances 0.000 abstract description 8
- 239000002351 wastewater Substances 0.000 abstract description 6
- 230000003287 optical effect Effects 0.000 abstract description 5
- -1 phenylacetyl glufosinate-ammonium Chemical compound 0.000 abstract description 5
- 230000009615 deamination Effects 0.000 abstract description 4
- 238000006481 deamination reaction Methods 0.000 abstract description 4
- 239000000575 pesticide Substances 0.000 abstract description 4
- 238000002360 preparation method Methods 0.000 abstract description 4
- 238000009776 industrial production Methods 0.000 abstract description 3
- 239000000463 material Substances 0.000 abstract description 3
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- 238000005903 acid hydrolysis reaction Methods 0.000 abstract 1
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 21
- 239000000706 filtrate Substances 0.000 description 15
- 238000001035 drying Methods 0.000 description 8
- 238000003756 stirring Methods 0.000 description 8
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 7
- 239000007787 solid Substances 0.000 description 6
- LOUPRKONTZGTKE-WZBLMQSHSA-N Quinine Chemical compound C([C@H]([C@H](C1)C=C)C2)C[N@@]1[C@@H]2[C@H](O)C1=CC=NC2=CC=C(OC)C=C21 LOUPRKONTZGTKE-WZBLMQSHSA-N 0.000 description 4
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- 230000000052 comparative effect Effects 0.000 description 3
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- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 235000001258 Cinchona calisaya Nutrition 0.000 description 2
- LOUPRKONTZGTKE-UHFFFAOYSA-N cinchonine Natural products C1C(C(C2)C=C)CCN2C1C(O)C1=CC=NC2=CC=C(OC)C=C21 LOUPRKONTZGTKE-UHFFFAOYSA-N 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000004821 distillation Methods 0.000 description 2
- GVGUFUZHNYFZLC-UHFFFAOYSA-N dodecyl benzenesulfonate;sodium Chemical compound [Na].CCCCCCCCCCCCOS(=O)(=O)C1=CC=CC=C1 GVGUFUZHNYFZLC-UHFFFAOYSA-N 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 230000002363 herbicidal effect Effects 0.000 description 2
- 239000010413 mother solution Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 229960000948 quinine Drugs 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 229940080264 sodium dodecylbenzenesulfonate Drugs 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- IHFRMUGEILMHNU-UHFFFAOYSA-N 2-hydroxy-5-nitrobenzaldehyde Chemical compound OC1=CC=C([N+]([O-])=O)C=C1C=O IHFRMUGEILMHNU-UHFFFAOYSA-N 0.000 description 1
- VMZCDNSFRSVYKQ-UHFFFAOYSA-N 2-phenylacetyl chloride Chemical compound ClC(=O)CC1=CC=CC=C1 VMZCDNSFRSVYKQ-UHFFFAOYSA-N 0.000 description 1
- 150000001413 amino acids Chemical group 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 238000005349 anion exchange Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
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- 239000012153 distilled water Substances 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 1
- 102000005396 glutamine synthetase Human genes 0.000 description 1
- 108020002326 glutamine synthetase Proteins 0.000 description 1
- 150000002309 glutamines Chemical class 0.000 description 1
- 239000004009 herbicide Substances 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
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- 229910021645 metal ion Inorganic materials 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
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- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
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- 239000002699 waste material Substances 0.000 description 1
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention relates to a process for preparing L-glufosinate by an enzyme splitting method, which belongs to the technical field of pesticide preparation and comprises three steps of preparing N-phenylacetyl-L-glufosinate by using DL-glufosinate and methyl phenylacetate, obtaining L-glufosinate solution by acid hydrolysis of the N-phenylacetyl-L-glufosinate and preparing the L-glufosinate by crystallization. The invention solves the problems of long process route, repeated acid and alkali adjustment, complex post treatment and large wastewater amount in the enzyme resolution method, uses the glufosinate-ammonium salt to directly enzyme synthesize the phenylacetyl glufosinate-ammonium, omits the deamination step required by the chemical synthesis of the phenylacetyl glufosinate-ammonium in the original method, and avoids the generation of deamination wastewater; the invention has high yield, the single resolution yield reaches more than 85 percent, the optical purity reaches more than 99 percent, and the invention is suitable for industrial production; the D-glufosinate which is separated out is racemized and then continuously used, so that the material is not wasted, the environment is not polluted, and the environment is protected.
Description
Technical Field
The invention relates to a process for preparing L-glufosinate-ammonium by an enzyme resolution method, and belongs to the technical field of pesticide preparation.
Background
Glufosinate is a glutamine analogue which binds to glutamine synthetase in plants and inhibits enzymatic activity, resulting in hindered glutamine synthesis in plants and death of the plants. Glufosinate has the characteristics of high efficiency, broad spectrum and low toxicity, and has grown as the second largest herbicide in the world. The glufosinate has an amino acid structure and has D, L two configurations, wherein only the L configuration has the herbicidal activity, so that the development of the optically pure L-glufosinate can effectively reduce the pesticide usage amount and has important significance for pesticide usage reduction and synergy.
The existing process of the mixed glufosinate is mature, the yield is high, and the separation of the mixed glufosinate into the L-glufosinate is an important research direction for synthesizing the L-glufosinate, and comprises a chemical separation method and an enzyme separation method.
The chemical resolution of DL-glufosinate is reported in US patent 5767309: and (3) salifying quinine and L-glufosinate, crystallizing and separating out, and neutralizing with ammonia to obtain the L-glufosinate. This method requires the use of expensive quinine and the resulting product has low optical purity. DL-glufosinate and chiral prosthetic groups and metal ions in the CN201811235641 are complexed to form coordination compounds, so that the configuration of D-glufosinate is reversed to generate L-glufosinate, and the L-glufosinate is obtained by hydrolysis.
U.S. Pat. Nos. 3,379, 4922013 report that the derivative is phenylacetylated with DL-glufosinate and then selectively hydrolyzed with penicillin G acylase to give L-glufosinate with high optical rotation, and the enzymatic resolution is continued after racemization of the N-glufosinate. In the method, glufosinate is deaminated and then reacts with phenylacetyl chloride to obtain phenylacetyl derivatives, acid is added until the pH value is 1-2, crystallization is carried out to obtain N-phenylacetyl-DL-glufosinate, N-phenylacetyl-DL-glufosinate is dissolved by alkali, penicillin G acylase is added for selectively hydrolyzing N-phenylacetyl-L-glufosinate, acid is added until the pH value is 1-2, crystallization is carried out to remove unreacted N-phenylacetyl-D-glufosinate, the pH value of the obtained L-glufosinate solution is regulated by alkali, and L-glufosinate is obtained after purification, and further reaction is carried out to obtain L-glufosinate. The process yield reaches more than 80%, the optical purity of the product is more than 99%, but the process is long, two times of alkali adjustment and two times of acid adjustment are needed, a large amount of acid and alkali are used, and a large amount of strong acid wastewater is generated. The specific synthetic route is shown in FIG. 2.
Disclosure of Invention
The invention aims to provide a process for preparing L-glufosinate-ammonium by an enzyme resolution method, which has the advantages of short process route, simple and convenient operation, high resolution efficiency and less three wastes, and is suitable for industrial production.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
a process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
adding DL-glufosinate and methyl phenylacetate into a mixed solvent, adding sodium dodecyl sulfate, adding acid to adjust the pH to 5.5-6.5, adding penicillin acylase to react, controlling the pH in the reaction process to be 5.5-6.5, adding an organic solvent to extract unreacted methyl phenylacetate after the reaction is finished, adding acid to adjust the pH to 1.5-2.5, crystallizing to obtain N-phenylacetyl-L-glufosinate, wherein a mother solution is D-glufosinate solution, and the D-glufosinate solution is used after racemization at high temperature;
step two, adding the N-phenylacetyl-L-glufosinate obtained in the step one into water for suspension, adding acid, heating for reflux, adding an organic solvent for extraction reaction to generate phenylacetic acid after the reaction is completed, and obtaining an L-glufosinate solution;
and thirdly, adding ammonia water into the L-glufosinate-ammonium solution obtained in the second step, regulating the pH value to 7-8, decompressing and dehydrating until the water content is 30-40%, adding methanol for dilution until the water content is 10-20% for crystallization, filtering and desalting to obtain an L-glufosinate-ammonium methanol solution, evaporating to dryness, and adding methanol for recrystallization to obtain the L-glufosinate-ammonium.
The technical scheme of the invention is further improved as follows: the adding amount of the sodium dodecyl sulfate in the first step is 0.5-1% of the mass of the DL-glufosinate.
The technical scheme of the invention is further improved as follows: the penicillin acylase in the first step is penicillin acylase solution or immobilized penicillin acylase, the adding amount of the penicillin acylase is 2-5% of the mass of DL-glufosinate, the reaction temperature is 15-20 ℃, and the reaction time is 10-20h.
The technical scheme of the invention is further improved as follows: the molar ratio of DL-glufosinate to methyl phenylacetate in the first step is 1:0.6-1.0.
The technical scheme of the invention is further improved as follows: the mixed solvent in the first step is a mixed solution of water and alcohol, the amount of the solvent is 3-4 times of the mass of DL-glufosinate-ammonium, the amount of the alcohol is 5-20% of the mass of water, and the alcohol is one of isopropanol, isobutanol, sec-octanol and isooctanol.
The technical scheme of the invention is further improved as follows: the adding amount of water in the second step is 3-5 times of the mass of the N-phenylacetyl-L-glufosinate-ammonium, and the adding amount of acid is 0.5-2 times of the molar amount of the N-phenylacetyl-L-glufosinate-ammonium.
The technical scheme of the invention is further improved as follows: the organic solvent in the first step and the second step is one of dichloromethane, chloroform, toluene and xylene, the adding mass of the organic solvent in the first step is 1-3 times of the mass of DL-glufosinate-ammonium, and the adding mass of the organic solvent in the second step is 2-3 times of the mass of N-phenylacetyl-L-glufosinate-acid.
The technical scheme of the invention is further improved as follows: the acid in the first step and the second step is hydrochloric acid or sulfuric acid.
By adopting the technical scheme, the invention has the following technical effects:
the invention solves the problems of long process route, repeated acid and alkali adjustment, complex post treatment and large wastewater amount in the enzyme resolution method, uses the glufosinate-ammonium salt to directly enzyme synthesize the phenylacetyl glufosinate-ammonium, omits the deamination step required by the chemical synthesis of the phenylacetyl glufosinate-ammonium in the original method, and avoids the generation of deamination wastewater. The invention realizes the production of L-glufosinate-ammonium by acid regulation for 1 time and alkali regulation for 1 time, and compared with the prior art, the method has the advantages of greatly shortening the operation flow and reducing the wastewater quantity by acid regulation and alkali regulation for two times.
The invention has high yield, the single resolution yield reaches more than 85 percent, the optical purity reaches more than 99 percent, and the invention is suitable for industrial production; the D-glufosinate which is separated out is racemized and then continuously used, so that the material is not wasted, the environment is not polluted, and the environment is protected.
Drawings
FIG. 1 is a process route diagram of the present invention;
fig. 2 is a prior art process roadmap.
Detailed Description
The invention is described in further detail below in connection with specific examples:
the process for preparing the L-glufosinate-ammonium by the enzyme splitting method is shown in figure 1, and comprises the following steps:
step one, adding DL-glufosinate and methyl phenylacetate into a solvent, wherein the molar ratio of the DL-glufosinate to the methyl phenylacetate is 1:0.6-1.0, preferably 1:0.6-0.75; the solvent is a mixed solvent of water and alcohol, the using amount of the alcohol is 5-20% of the mass of the water, the alcohol is one of isopropanol, isobutanol, sec-octanol and isooctanol, preferably sec-octanol, and the using amount of the mixed solvent is 3-4 times of the mass of DL-glufosinate; adding sodium dodecyl sulfate with the addition of 0.5-1% of DL-glufosinate, adding hydrochloric acid or sulfuric acid to regulate the pH to 5.5-6.5, adding penicillin acylase for reaction, wherein the penicillin acylase is penicillin acylase solution or immobilized penicillin acylase, preferably immobilized penicillin acylase, the addition of the enzyme is 2-5% of DL-glufosinate mass (the enzyme activity is 200-500 IU/g), the reaction temperature is 15-20 ℃, the pH is controlled to be 5.5-6.5 in the reaction process, the reaction time is 10-20h, the enzyme is filtered out after the reaction is finished, the filtrate is added with an organic solvent to extract unreacted methyl phenylacetate, the amount of the organic solvent is 1-3 times of the DL-glufosinate mass, the organic solvent is one of dichloromethane, chloroform, toluene and xylene, the obtained water phase is added with hydrochloric acid or sulfuric acid to regulate the pH to 1.5-2.5, and the obtained water phase is crystallized to obtain N-phenylacetyl-L-glufosinate acid, the mother solution is D-glufosinate, and D-glufosinate is crystallized at high temperature;
adding the N-phenylacetyl-L-glufosinate obtained in the step one into water with the mass of 3-5 times for suspension, adding hydrochloric acid or sulfuric acid, wherein the addition amount is 0.5-2 times of the molar amount of the N-phenylacetyl-L-glufosinate, heating for reflux, and adding an organic solvent for extraction reaction to generate phenylacetic acid after the reaction is finished, wherein the organic solvent is one of dichloromethane, chloroform, toluene and xylene, and the addition amount is 2-3 times of the mass of the N-phenylacetyl-L-glufosinate, so as to obtain an L-glufosinate solution;
and thirdly, adding ammonia water into the L-glufosinate-ammonium solution obtained in the second step, regulating the pH value to 7-8, decompressing and dehydrating until the water content is 30-40%, adding methanol to dilute until the water content is 10-20% for crystallization, filtering and desalting to obtain an L-glufosinate-ammonium methanol solution, evaporating to dryness, and adding the rest 2 times of methanol for crystallization to obtain the L-glufosinate-ammonium.
Example 1
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 45g (0.3 mol) of methyl phenylacetate are taken and added into a 1L four-mouth bottle, 300g of water, 15g of sec-octanol and 0.5g of sodium dodecyl benzene sulfonate are added, stirring is carried out, the pH is regulated to 5.5 by sulfuric acid, 5.2g (200 IU/g) of immobilized penicillin acylase is added for reaction at 15 ℃, the reaction time is 10h, and 2% sodium hydroxide solution is dropwise added in the reaction process to control the pH to be 5.5. Filtering out enzyme after the reaction is finished, adding 104g of toluene into the filtrate for extraction, adding sulfuric acid into the filtrate for regulating the pH value to 1.5, stirring for crystallization, filtering and drying to obtain 65.8g of N-phenylacetyl-L-glufosinate with the content of 99.1 percent and the yield of 87.1 percent. The mother liquor is D-glufosinate-ammonium solution. The toluene phase contains unreacted methyl phenylacetate, and toluene and methyl phenylacetate are recovered by distillation.
And step two, adding 200g of N-phenylacetyl-L-glufosinate in the step one into water for suspension, adding 11g (98%) of sulfuric acid, heating and refluxing for 25h, and adding 130g of toluene after the reaction is completed to extract phenylacetic acid generated by the reaction, thus obtaining the L-glufosinate solution.
And thirdly, adding ammonia water into the L-glufosinate-ammonium solution to adjust the pH value to 7-8, decompressing and dehydrating until the solid content is 70%, adding methanol to adjust the water content to 20%, crystallizing for 30min, filtering and desalting, decompressing and evaporating filtrate to dryness, adding 100g of methanol for crystallizing, filtering and drying to obtain 44.1g of L-glufosinate, wherein the content is 95.5%, the ee value is 99.7%, and the yield is 85% based on the L-glufosinate in the raw material DL-glufosinate. The content and ee value detection method is high performance liquid chromatography, chromatographic column 250 mm ×4.6 mm (i.d.) strong anion exchange stainless steel column, filled with SAX,5 μm filler; mobile phase: weighing 6.80g of potassium dihydrogen phosphate, dissolving with 970mL of redistilled distilled water, adding 30mL of methanol, filtering with a filter membrane, and degassing, c (KH 2 PO 4 ) =0.05 mol/L. Detection wavelength: 195 nm. And (5) measuring the content of L-glufosinate and D-glufosinate, and calculating the ee value.
Example 2
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 75g (0.5 mol) of methyl phenylacetate are taken and added into a 1L four-mouth bottle, 350g of water, 70g of sec-octanol and 1g of sodium dodecyl sulfate are added, the mixture is stirred, the pH is adjusted to 6.5 by hydrochloric acid, 2g (200 IU/g) of immobilized penicillin acylase is added, the reaction is carried out at the temperature of 20 ℃, the reaction time is 20 hours, and 2% sodium hydroxide solution is dropwise added in the reaction process to control the reaction solution to maintain the pH to 6.5. Filtering out enzyme after the reaction is finished, adding 312g of toluene into the filtrate for extraction, adding hydrochloric acid to adjust the pH to 2.5 for stirring crystallization, filtering, and drying to obtain 68.0g of N-phenylacetyl-L-glufosinate with the content of 99 percent and the yield of 90.0 percent. The toluene phase contains unreacted methyl phenylacetate, and toluene and methyl phenylacetate are recovered by distillation.
Step two, adding 340g of N-phenylacetyl-L-glufosinate acid in the step one into water for suspension, adding 45g (98%) of sulfuric acid, heating and refluxing for 15h, and adding 200g of toluene for extraction reaction to generate phenylacetic acid after the reaction is completed, thus obtaining the L-glufosinate acid solution.
Adding ammonia water into the L-glufosinate-ammonium solution to adjust the pH value to 7-8, decompressing and dehydrating until the solid content is 70%, adding methanol to adjust the water content to 20%, crystallizing for 30min, filtering and desalting, decompressing and evaporating filtrate to dryness, adding 100g of methanol for crystallizing, filtering and drying to obtain 45.0g of L-glufosinate, wherein the content is 95.7%, the ee value is 99.8%, and the yield is 87% of that of the L-glufosinate in the raw material DL-glufosinate.
Example 3
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
step one, 104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 56.2g (0.375 mol) of methyl phenylacetate are added into a four-necked flask of 1Lml, 300g of water, 50g of isooctanol and 0.7g of sodium dodecyl sulfate are added, the mixture is stirred, the pH is adjusted to 6.5 by hydrochloric acid, 3g (200 IU/g) of immobilized penicillin acylase is added for reaction at the temperature of 20 ℃, the reaction time is 15h, and 2% of sodium hydroxide solution is dropwise added in the reaction process to control the pH=6.0. Filtering out enzyme after the reaction is finished, adding 200g of dichloromethane into the filtrate for extraction, adding hydrochloric acid into the filtrate for regulating the pH value to 2.0, stirring for crystallization, and filtering to obtain 90g of wet solid N-phenylacetyl-L-glufosinate with water content of about 25%.
Step two, adding 250g of N-phenylacetyl-L-glufosinate acid in the step one into water for suspension, adding 50g (38%) of hydrochloric acid, heating and refluxing for 20h, and adding 200g of dichloromethane to extract phenylacetic acid generated by the reaction after the reaction is completed, so as to obtain the L-glufosinate acid solution.
Step three, decompressing, dehydrating and concentrating the L-glufosinate-ammonium acid solution, adding ammonia water to pH 7-8, decompressing, dehydrating to solid content 70%, adding methanol to regulate the water content to 15% by mass, crystallizing for 30min, filtering to remove salt, decompressing, evaporating filtrate to dryness, adding 100g of methanol for crystallization, filtering and drying to obtain 45.7g of L-glufosinate-ammonium, wherein the content is 95.4%, the ee value is 99.7%, and the yield is 88% of that of the L-glufosinate-ammonium in the raw material DL-glufosinate-ammonium.
Example 4
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
step one, 104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 56.2g (0.375 mol) of methyl phenylacetate are taken and added into a four-necked flask of 1Lml, 300g of water, 50g of isopropanol and 0.5g of sodium dodecyl sulfate are added, the mixture is stirred, the pH is adjusted to 6.5 by hydrochloric acid, 5g (200 IU/g) of immobilized penicillin acylase is added for reaction at the temperature of 20 ℃, the reaction time is 10 hours, and 2% of sodium hydroxide solution is dropwise added in the reaction process to control the pH=6.0. Filtering out enzyme after the reaction is finished, adding 200g of dichloromethane into the filtrate for extraction, adding hydrochloric acid into the filtrate for regulating the pH value to 2.0, stirring for crystallization, and filtering to obtain 87g of wet solid N-phenylacetyl-L-glufosinate with water content of about 25%.
And step two, adding 250g of N-phenylacetyl-L-glufosinate in the step one into water for suspension, adding 11g (98%) of sulfuric acid, heating and refluxing for 25h, and adding 200g of dimethylbenzene for extraction reaction to generate phenylacetic acid after the reaction is completed, so as to obtain the L-glufosinate solution.
And thirdly, adding ammonia water into the L-glufosinate-ammonium solution to adjust the pH value to 7-8, decompressing and dehydrating until the solid content is 70%, adding methanol to adjust the water content to 20%, crystallizing for 30min, filtering and desalting, decompressing and evaporating filtrate to dryness, adding 100g of methanol for crystallizing, filtering and drying to obtain 44.8g of L-glufosinate, wherein the content is 95.1%, the ee value is 99.3%, and the yield is 86% of that of the L-glufosinate in the raw material DL-glufosinate.
EXAMPLE 5D racemization of glufosinate
In example 2, the mother liquor obtained by filtering the crystallized N-phenylacetyl-L-glufosinate acid in the first step is D-glufosinate acid solution, 2g of 5-nitrosalicylaldehyde is added, the mixture is heated to reflux reaction for 20 hours to obtain racemic DL-glufosinate, part of water is distilled out, methanol is added to the water until the water content is 20%, salt is filtered, the filtrate is diluted by methanol until the water content is 10-15%, and the DL-glufosinate acid is obtained by crystallization at room temperature, and enzyme resolution is continued.
Comparative example 1
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
preparation of N-phenylacetyl-L-glufosinate: 104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 45g (0.3 mol) of methyl phenylacetate are taken, a 1L four-mouth bottle is added, 300g of water, 60g of methanol and 0.5g of sodium dodecyl benzene sulfonate are added, stirring is carried out, the pH is regulated to 5.5 by sulfuric acid, 5.2g (200 IU/g) of immobilized penicillin acylase is added, the reaction is carried out at 15 ℃ for 10 hours, and 2% sodium hydroxide solution is dropwise added in the reaction process to control the reaction solution to maintain the pH to 5.5. Filtering out enzyme after the reaction is finished, adding 200g of toluene into the filtrate for extraction, adding sulfuric acid to adjust the pH to 1.5, stirring for crystallization, filtering and drying to obtain 34.1g of N-phenylacetyl-L-glufosinate with the content of 99.1 percent and the yield of 45.2 percent.
Comparative example 2
A process for preparing L-glufosinate-ammonium by an enzyme resolution method comprises the following steps:
preparation of N-phenylacetyl-L-glufosinate: 104g (95%, 0.5 mol) of DL-glufosinate-ammonium and 75g (0.5 mol) of methyl phenylacetate are taken and added into a 1L four-mouth bottle, 350g of water, 70g of sec-octanol and 1g of sodium dodecyl sulfate are added and stirred, the pH is adjusted to 8 by sodium hydroxide, 2g (200 IU/g) of immobilized penicillin acylase is added for reaction at 20 ℃, the reaction time is 20h, and 2% sodium hydroxide solution is dropwise added in the reaction process to control the reaction solution to maintain the pH8. Filtering out enzyme after the reaction is finished, adding 312g of toluene into the filtrate for extraction, adding hydrochloric acid to adjust the pH to 2.5 for stirring crystallization, filtering, and drying to obtain 4.5g of N-phenylacetyl-L-glufosinate with the content of 99.0 percent and the yield of 6 percent.
The reaction materials and enzymes used in the examples and comparative examples were all of the same batch.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, or alternatives falling within the spirit and principles of the invention.
Claims (5)
1. The process for preparing the L-glufosinate-ammonium by the enzyme splitting method is characterized by comprising the following steps of:
step one, adding DL-glufosinate and methyl phenylacetate into a mixed solvent, wherein the molar ratio of the DL-glufosinate to the methyl phenylacetate is 1:0.6-1.0, adding sodium dodecyl sulfonate, wherein the adding amount of the sodium dodecyl sulfonate is 0.5-1% of the mass of DL-glufosinate, adding acid to adjust the pH value to 5.5-6.5, adding penicillin acylase to react, wherein the adding amount of enzyme is 2-5% of the mass of DL-glufosinate, the reaction temperature is 15-20 ℃, the reaction time is 10-20h, the pH value is controlled to be kept at 5.5-6.5 in the reaction process, adding an organic solvent to extract unreacted methyl phenylacetate after the reaction is completed, adding water to adjust the pH value to 1.5-2.5, crystallizing to obtain N-phenylacetyl-L-glufosinate, and applying the D-glufosinate solution as mother liquor after high-temperature racemization; the mixed solvent is a mixed solution of water and alcohol, the amount of the solvent is 3-4 times of the mass of DL-glufosinate-ammonium, the amount of the alcohol is 5-20% of the mass of water, and the alcohol is one of isopropanol, isobutanol, sec-octanol and isooctanol;
step two, adding the N-phenylacetyl-L-glufosinate obtained in the step one into water for suspension, adding acid, heating for reflux, adding an organic solvent for extraction reaction to generate phenylacetic acid after the reaction is completed, and obtaining an L-glufosinate solution;
and thirdly, adding ammonia water into the L-glufosinate-ammonium solution obtained in the second step, regulating the pH value to 7-8, decompressing and dehydrating until the water content is 30-40%, adding methanol for dilution until the water content is 10-20% for crystallization, filtering and desalting to obtain an L-glufosinate-ammonium methanol solution, evaporating to dryness, and adding methanol for recrystallization to obtain the L-glufosinate-ammonium.
2. The process for preparing L-glufosinate-ammonium by an enzyme resolution method according to claim 1, wherein the process comprises the following steps: in the first step, the penicillin acylase is penicillin acylase solution or immobilized penicillin acylase.
3. The process for preparing L-glufosinate-ammonium by an enzyme resolution method according to claim 1, wherein the process comprises the following steps: the adding amount of water in the second step is 3-5 times of the mass of the N-phenylacetyl-L-glufosinate-ammonium, and the adding amount of acid is 0.5-2 times of the molar amount of the N-phenylacetyl-L-glufosinate-ammonium.
4. The process for preparing L-glufosinate-ammonium by an enzyme resolution method according to claim 1, wherein the process comprises the following steps: the organic solvent in the first step and the second step is one of dichloromethane, chloroform, toluene and xylene, the adding mass of the organic solvent in the first step is 1-3 times of the mass of DL-glufosinate-ammonium, and the adding mass of the organic solvent in the second step is 2-3 times of the mass of N-phenylacetyl-L-glufosinate-acid.
5. The process for preparing L-glufosinate-ammonium by an enzyme resolution method according to claim 1, wherein the process comprises the following steps: the acid in the first step and the second step is hydrochloric acid or sulfuric acid.
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