CN116676358A - Preparation method of 2-fluoro-2-deoxyuridine - Google Patents
Preparation method of 2-fluoro-2-deoxyuridine Download PDFInfo
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- CN116676358A CN116676358A CN202310899885.2A CN202310899885A CN116676358A CN 116676358 A CN116676358 A CN 116676358A CN 202310899885 A CN202310899885 A CN 202310899885A CN 116676358 A CN116676358 A CN 116676358A
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- 238000002360 preparation method Methods 0.000 title claims abstract description 16
- 238000006243 chemical reaction Methods 0.000 claims abstract description 39
- 102000004190 Enzymes Human genes 0.000 claims abstract description 33
- 108090000790 Enzymes Proteins 0.000 claims abstract description 33
- 238000006555 catalytic reaction Methods 0.000 claims abstract description 27
- 239000002798 polar solvent Substances 0.000 claims abstract description 13
- 238000002156 mixing Methods 0.000 claims abstract description 4
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 12
- 238000000034 method Methods 0.000 claims description 11
- 238000001953 recrystallisation Methods 0.000 claims description 11
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 9
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 9
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 8
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 8
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 8
- 239000000243 solution Substances 0.000 claims description 8
- 229930027917 kanamycin Natural products 0.000 claims description 7
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 claims description 7
- 229960000318 kanamycin Drugs 0.000 claims description 7
- 229930182823 kanamycin A Natural products 0.000 claims description 7
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 6
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 claims description 6
- 239000007853 buffer solution Substances 0.000 claims description 6
- 239000007795 chemical reaction product Substances 0.000 claims description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- 240000006439 Aspergillus oryzae Species 0.000 claims description 4
- 235000002247 Aspergillus oryzae Nutrition 0.000 claims description 4
- 238000000746 purification Methods 0.000 claims description 4
- 108010006654 Bleomycin Proteins 0.000 claims description 3
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 3
- JLTDJTHDQAWBAV-UHFFFAOYSA-N N,N-dimethylaniline Chemical compound CN(C)C1=CC=CC=C1 JLTDJTHDQAWBAV-UHFFFAOYSA-N 0.000 claims description 3
- SECXISVLQFMRJM-UHFFFAOYSA-N N-Methylpyrrolidone Chemical compound CN1CCCC1=O SECXISVLQFMRJM-UHFFFAOYSA-N 0.000 claims description 3
- 239000007983 Tris buffer Substances 0.000 claims description 3
- 229960001561 bleomycin Drugs 0.000 claims description 3
- OYVAGSVQBOHSSS-UAPAGMARSA-O bleomycin A2 Chemical compound N([C@H](C(=O)N[C@H](C)[C@@H](O)[C@H](C)C(=O)N[C@@H]([C@H](O)C)C(=O)NCCC=1SC=C(N=1)C=1SC=C(N=1)C(=O)NCCC[S+](C)C)[C@@H](O[C@H]1[C@H]([C@@H](O)[C@H](O)[C@H](CO)O1)O[C@@H]1[C@H]([C@@H](OC(N)=O)[C@H](O)[C@@H](CO)O1)O)C=1N=CNC=1)C(=O)C1=NC([C@H](CC(N)=O)NC[C@H](N)C(N)=O)=NC(N)=C1C OYVAGSVQBOHSSS-UAPAGMARSA-O 0.000 claims description 3
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 claims description 3
- 229910000403 monosodium phosphate Inorganic materials 0.000 claims description 3
- 235000019799 monosodium phosphate Nutrition 0.000 claims description 3
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 claims description 3
- 229910000162 sodium phosphate Inorganic materials 0.000 claims description 3
- 239000001488 sodium phosphate Substances 0.000 claims description 3
- 235000011008 sodium phosphates Nutrition 0.000 claims description 3
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 claims description 3
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 claims description 3
- JLEXUIVKURIPFI-UHFFFAOYSA-N tris phosphate Chemical compound OP(O)(O)=O.OCC(N)(CO)CO JLEXUIVKURIPFI-UHFFFAOYSA-N 0.000 claims description 2
- KRHYYFGTRYWZRS-UHFFFAOYSA-N Fluorane Chemical compound F KRHYYFGTRYWZRS-UHFFFAOYSA-N 0.000 abstract description 8
- 239000002253 acid Substances 0.000 abstract description 6
- 239000002351 wastewater Substances 0.000 abstract description 6
- 239000003153 chemical reaction reagent Substances 0.000 abstract description 4
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 abstract description 2
- 230000003197 catalytic effect Effects 0.000 abstract description 2
- 238000003912 environmental pollution Methods 0.000 abstract description 2
- 229910052731 fluorine Inorganic materials 0.000 abstract description 2
- 239000011737 fluorine Substances 0.000 abstract description 2
- 231100000086 high toxicity Toxicity 0.000 abstract description 2
- 238000003786 synthesis reaction Methods 0.000 abstract description 2
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 24
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 24
- 239000007864 aqueous solution Substances 0.000 description 15
- 239000002904 solvent Substances 0.000 description 9
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical class O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 description 7
- 238000001914 filtration Methods 0.000 description 7
- 238000004128 high performance liquid chromatography Methods 0.000 description 7
- 239000007787 solid Substances 0.000 description 6
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 6
- 239000012065 filter cake Substances 0.000 description 5
- 230000002035 prolonged effect Effects 0.000 description 5
- 230000035484 reaction time Effects 0.000 description 5
- 239000000706 filtrate Substances 0.000 description 4
- 239000011259 mixed solution Substances 0.000 description 4
- 230000001105 regulatory effect Effects 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 239000000337 buffer salt Substances 0.000 description 3
- 238000002425 crystallisation Methods 0.000 description 3
- 230000008025 crystallization Effects 0.000 description 3
- 229960002063 sofosbuvir Drugs 0.000 description 3
- TTZHDVOVKQGIBA-IQWMDFIBSA-N sofosbuvir Chemical compound N1([C@@H]2O[C@@H]([C@H]([C@]2(F)C)O)CO[P@@](=O)(N[C@@H](C)C(=O)OC(C)C)OC=2C=CC=CC=2)C=CC(=O)NC1=O TTZHDVOVKQGIBA-IQWMDFIBSA-N 0.000 description 3
- 208000005176 Hepatitis C Diseases 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- -1 hydroxyl compound Chemical class 0.000 description 2
- 229910017053 inorganic salt Inorganic materials 0.000 description 2
- NROKBHXJSPEDAR-UHFFFAOYSA-M potassium fluoride Chemical compound [F-].[K+] NROKBHXJSPEDAR-UHFFFAOYSA-M 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000008213 purified water Substances 0.000 description 2
- 238000007142 ring opening reaction Methods 0.000 description 2
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 description 2
- 229940045145 uridine Drugs 0.000 description 2
- 208000006154 Chronic hepatitis C Diseases 0.000 description 1
- 102000014150 Interferons Human genes 0.000 description 1
- 108010050904 Interferons Proteins 0.000 description 1
- 241001460073 Trichoderma asperellum Species 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000010511 deprotection reaction Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 238000003682 fluorination reaction Methods 0.000 description 1
- 238000011010 flushing procedure Methods 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 208000010710 hepatitis C virus infection Diseases 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 229940079322 interferon Drugs 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 235000003270 potassium fluoride Nutrition 0.000 description 1
- 239000011698 potassium fluoride Substances 0.000 description 1
- GRJJQCWNZGRKAU-UHFFFAOYSA-N pyridin-1-ium;fluoride Chemical compound F.C1=CC=NC=C1 GRJJQCWNZGRKAU-UHFFFAOYSA-N 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000010025 steaming Methods 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- NTJBWZHVSJNKAD-UHFFFAOYSA-N triethylazanium;fluoride Chemical compound [F-].CC[NH+](CC)CC NTJBWZHVSJNKAD-UHFFFAOYSA-N 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/26—Preparation of nitrogen-containing carbohydrates
- C12P19/28—N-glycosides
- C12P19/38—Nucleosides
- C12P19/385—Pyrimidine nucleosides
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biotechnology (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Saccharide Compounds (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The application relates to the technical field of organic synthesis, in particular to a preparation method of 2-fluoro-2-deoxyuridine. The application provides a preparation method of 2-fluoro-2 deoxyuridine, which comprises the following steps: mixing 2-fluoro-2-deoxycytidine, biological enzyme and polar solvent for enzyme catalytic reaction to obtain 2-fluoro-2-deoxyuridine. The preparation method provided by the application does not need to use high-toxicity acid reagents such as hydrofluoric acid and the like, and prepares the 2-fluoro-2-deoxyuridine through enzyme catalysis, so that the generation of fluorine-containing wastewater or acid wastewater is avoided, the safety of the preparation process is ensured, and the problem of environmental pollution is avoided. In addition, the preparation method provided by the application has high biological enzyme catalytic conversion rate and high yield.
Description
Technical Field
The application relates to the technical field of organic synthesis, in particular to a preparation method of 2-fluoro-2-deoxyuridine.
Background
2-fluoro-2-deoxyuridine is a key intermediate for preparing chronic hepatitis C drug sofosbuvir. Sofosbuvir is the first drug to safely and effectively treat certain types of hepatitis C without combining interferon, and has the following structural formula:
the 2-fluoro-2-deoxyuridine is used as a key intermediate of sofosbuvir, the intermediate can be used for preparing 2,2 '-dehydrated uridine from uridine, and then the 2,2' -dehydrated uridine is subjected to ring opening by using reagents such as hydrofluoric acid, pyridine hydrofluoric acid complex, triethylamine hydrofluoric acid complex, potassium fluoride and the like to prepare the 2-fluoro-2-deoxyuridine, wherein the specific synthetic route is shown in a formula A.
The 2-fluoro-2-deoxyuridine can also be prepared into 2,2' -dehydrated uridine by taking uridine as a raw material, then sequentially carrying out hydroxyl protection reaction and ring-opening reaction at the second position to prepare a hydroxyl compound, and then carrying out fluorination deprotection on the hydroxyl compound to obtain the target compound 2-fluoro-2-deoxyuridine, wherein the specific synthetic route is shown in a formula B.
However, both the preparation methods need to be carried out in an acidic environment, the reaction conditions are harsh, and a large amount of acidic wastewater which is difficult to treat is produced.
Disclosure of Invention
In view of this, the present application provides a method for preparing 2-fluoro-2-deoxyuridine. The application only uses 2-fluoro-2-deoxycytidine to carry out enzyme catalytic reaction in the presence of biological enzyme and polar solvent, thus obtaining 2-fluoro-2-deoxyuridine without using high toxic acid reagents such as hydrofluoric acid and the like and generating acid wastewater.
In order to achieve the above object, the present application provides the following technical solutions:
a method for preparing 2-fluoro-2 deoxyuridine, comprising the following steps:
mixing 2-fluoro-2-deoxycytidine, biological enzyme and polar solvent for enzyme catalytic reaction to obtain 2-fluoro-2-deoxyuridine.
Preferably, the polar solvent comprises purified water, N dimethylaniline, N-dimethylformamide, dimethylsulfoxide, N-methylpyrrolidone, acetonitrile, tetrahydrofuran or acetone.
Preferably, the biological enzyme comprises one or more of kanamycin deaminase, aspergillus oryzae deaminase and bleomycin deaminase.
Preferably, the mass ratio of the 2-fluoro-2-deoxycytidine to the biological enzyme is 1:0.001-0.01.
Preferably, the mass-volume ratio of the 2-fluoro-2-deoxycytidine to the polar solvent is 1:3-30.
Preferably, the reaction temperature of the enzyme catalytic reaction is 10-40 ℃ and the time is 20-48 h.
Preferably, the pH value of the reaction solution of the enzyme catalytic reaction is 7.2-8.0.
Preferably, the pH value of the reaction solution of the enzyme-catalyzed reaction is adjusted by a buffer solution.
Preferably, the solute in the buffer solution comprises one or more of potassium dihydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate, disodium hydrogen phosphate, tris and sodium phosphate.
Preferably, after the enzyme-catalyzed reaction, the method further comprises the step of carrying out recrystallization and purification treatment on the enzyme-catalyzed reaction product.
The application provides a preparation method of 2-fluoro-2 deoxyuridine, which comprises the following steps: 2-fluoro-2-deoxycytidine, biological enzyme and polar solvent are mixed for enzyme catalytic reaction to prepare the 2-fluoro-2-deoxyuridine. The preparation method provided by the application does not need to use high-toxicity acid reagents such as hydrofluoric acid and the like, and prepares the 2-fluoro-2-deoxyuridine through enzyme catalysis, so that the generation of fluorine-containing wastewater or acid wastewater is avoided, the safety of the preparation process is ensured, and the problem of environmental pollution is avoided. In addition, the preparation method provided by the application has high biological enzyme catalytic conversion rate and high yield.
Detailed Description
The application provides a preparation method of 2-fluoro-2 deoxyuridine, which comprises the following steps:
mixing 2-fluoro-2-deoxycytidine, biological enzyme and polar solvent for enzyme catalytic reaction to obtain 2-fluoro-2-deoxyuridine.
In the present application, the desired materials are commercially available products well known to those skilled in the art unless specified otherwise.
In the present application, the polar solvent preferably includes water, N-dimethylaniline, N-dimethylformamide, dimethylsulfoxide, N-methylpyrrolidone, acetonitrile, tetrahydrofuran or acetone, and more preferably water.
In the present application, the biological enzyme preferably includes one or more of kanamycin deaminase, aspergillus oryzae deaminase and bleomycin deaminase, more preferably kanamycin deaminase.
In the present application, the mass ratio of the 2-fluoro-2-deoxycytidine to the biological enzyme is preferably 1:0.001-0.01, and more preferably 1:0.005; the mass-volume ratio of the 2-fluoro-2-deoxycytidine to the polar solvent is preferably 1:3-30, and more preferably 1:15.
In the present application, the reaction temperature of the enzyme-catalyzed reaction is preferably 10 to 40 ℃, more preferably 30 ℃, and the time is preferably 20 to 48 hours, more preferably 25 hours. In the present application, the pH of the reaction solution of the enzyme-catalyzed reaction is preferably 7.2 to 8.0, more preferably 7.5, and the pH of the reaction solution of the enzyme-catalyzed reaction is preferably adjusted by a buffer solution, and the solute in the buffer solution preferably comprises one or more of potassium dihydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate, disodium hydrogen phosphate, tris (hydroxymethyl) aminomethane and sodium phosphate, more preferably a mixed buffer salt of dipotassium hydrogen phosphate and tris (hydroxymethyl) aminomethane. In the specific embodiment of the application, preferably, the 2-fluoro-2-deoxycytidine, the polar solvent and the buffer salt are mixed to obtain a reaction system, the pH value of the reaction system is regulated to 7.2-8.0 by the buffer salt, and then biological enzyme is added for enzyme catalytic reaction to obtain the 2-fluoro-2-deoxyuridine.
In the present application, the enzymatic reaction has the following reaction formula:
after the enzyme catalytic reaction, the application preferably further comprises the step of carrying out recrystallization and purification treatment on the enzyme catalytic reaction product; in a specific embodiment of the present application, the recrystallization purification treatment preferably includes the steps of: and (3) steaming the enzyme catalytic reaction product in a rotary way, dissolving the obtained residue in a first recrystallization solvent, filtering to obtain filtrate, adding a second recrystallization solvent into the filtrate under a heating state, cooling, crystallizing and filtering in sequence, flushing a filter cake with a third recrystallization solvent, and drying the flushed filter cake to obtain the 2-fluoro-2-deoxyuridine. In the present application, the temperature in the heated state is preferably 40 to 50 ℃, and more preferably 45 ℃; the temperature of the cooling crystallization is preferably 0 to 10 ℃, and more preferably 0 ℃.
In the present application, the first, second and third recrystallization solvents independently preferably include at least one or more of purified water, methanol, ethanol, methylene chloride and acetone, the first recrystallization solvent is more preferably methanol, the second recrystallization solvent is more preferably methylene chloride, the third recrystallization solvent is more preferably a mixed solution of methanol and methylene chloride, and the volume ratio of methanol and methylene chloride in the mixed solution is preferably 1:1 to 3, more preferably 1:2.
The following description of the technical solutions in the embodiments of the present application will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present application, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the application without making any inventive effort, are intended to be within the scope of the application.
Example 1
10g of 2-fluoro-2-deoxyuridine, 10mL of 0.1mol/L of tris aqueous solution and 5mL of 0.1mol/L of dipotassium hydrogen phosphate aqueous solution are added into a 250mL three-necked flask to obtain a reaction system, the pH value=7.5 of the reaction system is regulated by using the 0.1mol/L of potassium dihydrogen phosphate aqueous solution, 50mg of kanamycin deaminase is added, the reaction is carried out for 25 hours at 30 ℃, the conversion of 2-fluoro-2-deoxyuridine is monitored by HPLC, the enzyme catalytic reaction product is distilled to remove the solvent, the residue is dissolved in 100mL of methanol, the filtration is carried out, insoluble inorganic salt and insoluble enzyme protein after wall breaking are filtered, the filtrate is heated to 45 ℃, 200mL of dichloromethane is dripped, the temperature is reduced to 0-10 ℃ for crystallization, the filtration is carried out, the filter cake is dried by using cold methanol/dichloromethane mixed solution (volume ratio is 1:2), 9.63 g of white solid is obtained, the white solid is 2-fluoro-2-deoxyuridine, and the yield of 2-fluoro-2-deoxyuridine is 96%.
Example 2
10g of 2-fluoro-2-deoxyuridine, 10mL of 0.1mol/L of tris (hydroxymethyl) aminomethane aqueous solution and 5mL of 0.1mol/L of dipotassium hydrogen phosphate aqueous solution are added into a 250mL three-port bottle to obtain a reaction system, the pH=7.5 of the reaction system is regulated by 0.1mol/L of potassium dihydrogen phosphate aqueous solution, 50mg of Aspergillus oryzae deaminase is added, the reaction is carried out at 30 ℃ for 25 hours, the conversion of 2-fluoro-2-deoxyuridine is monitored by HPLC, the reaction time is prolonged to 48 hours, the conversion is monitored by HPLC to about 92%, the reaction time is prolonged to 72 hours, the conversion rate is not changed obviously, the enzyme catalytic reaction product is distilled to remove a solvent, the residue is dissolved in 100mL of methanol, the filtration is carried out, insoluble inorganic salt and insoluble enzyme protein after filtration, the filtrate is heated to 45 ℃, 200mL of dichloromethane is dripped, the temperature is reduced to 0-10 ℃, the wall breaking crystallization is carried out, the filtration is carried out, the filter cake is mixed with cold methanol/dichloromethane mixed solution (volume ratio is 1:2), the filter cake is leached into a mixed solvent (volume ratio is 1:2) and the white solid (2:2) is dehydrated into the white solid, the yield is about 12-2-fluorouridine, the white solid is obtained, and the white solid is dried.
Example 3
10g of 2-fluoro-2-deoxyuridine, 10mL of 0.1mol/L of an aqueous solution of tris (hydroxymethyl) aminomethane and 5mL of 0.1mol/L of an aqueous solution of dipotassium hydrogenphosphate were added to a 250mL three-necked flask to obtain a reaction system, the pH=7.5 of the reaction system was adjusted with 0.1mol/L of an aqueous solution of potassium dihydrogenphosphate, 50mg of kanamycin deaminase was added, the reaction was carried out at 30℃for 25 hours, the HPLC was monitored to convert about 80%, and the reaction time was prolonged to 72 hours without significant change in conversion.
Comparative example 1
10g of 2-fluoro-2-deoxyuridine, 10mL of 0.1mol/L of tris (hydroxymethyl) aminomethane aqueous solution and 5mL of 0.1mol/L of dipotassium hydrogen phosphate aqueous solution are added into a 250mL three-necked flask to obtain a reaction system, the pH=7.5 of the reaction system is regulated by using the 0.1mol/L of potassium dihydrogen phosphate aqueous solution, 50mg of trichoderma asperellum deaminase is added, the temperature is kept at 30 ℃ for reaction for 25 hours, HPLC (high performance liquid chromatography) monitoring is carried out for about 5 percent conversion, the reaction time is prolonged to 72 hours, and the conversion rate is not changed obviously.
Comparative example 2
10g of 2-fluoro-2-deoxyuridine, 10mL of 0.1mol/L of an aqueous solution of tris (hydroxymethyl) aminomethane and 5mL of 0.1mol/L of an aqueous solution of dipotassium hydrogenphosphate were added to a 250mL three-necked flask to obtain a reaction system, the pH=9.0 of the reaction system was adjusted by using the 0.1mol/L aqueous solution of potassium dihydrogenphosphate, 50mg of kanamycin deaminase was added, the reaction was carried out at 30℃for 25 hours, the HPLC was monitored to convert about 3%, the product was extremely small, the reaction time was prolonged to 72 hours, and the conversion rate was not significantly changed.
The foregoing is merely a preferred embodiment of the present application and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present application, which are intended to be comprehended within the scope of the present application.
Claims (10)
1. The preparation method of the 2-fluoro-2 deoxyuridine is characterized by comprising the following steps:
mixing 2-fluoro-2-deoxycytidine, biological enzyme and polar solvent for enzyme catalytic reaction to obtain the 2-fluoro-2-deoxyuridine.
2. The method according to claim 1, wherein the polar solvent comprises water, N dimethylaniline, N-dimethylformamide, dimethylsulfoxide, N-methylpyrrolidone, acetonitrile, tetrahydrofuran or acetone.
3. The method of claim 1, wherein the biological enzyme comprises one or more of kanamycin deaminase, aspergillus oryzae deaminase, and bleomycin deaminase.
4. The method according to claim 1 or 3, wherein the mass ratio of the 2-fluoro-2-deoxycytidine to the biological enzyme is 1:0.001-0.01.
5. The preparation method according to claim 1 or 2, wherein the mass-volume ratio of the 2-fluoro-2-deoxycytidine to the polar solvent is 1:3-30.
6. The method according to claim 1, wherein the reaction temperature of the enzyme-catalyzed reaction is 10 to 40 ℃ and the time is 20 to 48 hours.
7. The method according to claim 1, wherein the pH of the reaction solution of the enzyme-catalyzed reaction is 7.2 to 8.0.
8. The method according to claim 1 or 7, wherein the pH of the reaction solution of the enzyme-catalyzed reaction is adjusted by a buffer solution.
9. The method of claim 8, wherein the solute in the buffer solution comprises one or more of potassium dihydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate, disodium hydrogen phosphate, tris and sodium phosphate.
10. The method according to claim 1, further comprising subjecting the enzyme-catalyzed reaction product to a recrystallization purification treatment after the enzyme-catalyzed reaction.
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