CN114540455A - Process for the preparation of chiral beta-amino acid derivatives - Google Patents
Process for the preparation of chiral beta-amino acid derivatives Download PDFInfo
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- CN114540455A CN114540455A CN202011331904.4A CN202011331904A CN114540455A CN 114540455 A CN114540455 A CN 114540455A CN 202011331904 A CN202011331904 A CN 202011331904A CN 114540455 A CN114540455 A CN 114540455A
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- amino acid
- chiral
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- acid derivative
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- 150000001576 beta-amino acids Chemical class 0.000 title claims abstract description 61
- 238000000034 method Methods 0.000 title claims abstract description 38
- 230000008569 process Effects 0.000 title claims abstract description 29
- 238000002360 preparation method Methods 0.000 title claims abstract description 20
- 239000000758 substrate Substances 0.000 claims abstract description 117
- 239000007787 solid Substances 0.000 claims abstract description 85
- 239000002904 solvent Substances 0.000 claims abstract description 55
- 239000002245 particle Substances 0.000 claims abstract description 52
- 238000006911 enzymatic reaction Methods 0.000 claims abstract description 42
- 239000002994 raw material Substances 0.000 claims abstract description 33
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims abstract description 25
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 23
- 102000004190 Enzymes Human genes 0.000 claims abstract description 22
- 108090000790 Enzymes Proteins 0.000 claims abstract description 22
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 20
- 239000003054 catalyst Substances 0.000 claims abstract description 13
- 239000012046 mixed solvent Substances 0.000 claims abstract description 7
- 125000003158 alcohol group Chemical group 0.000 claims abstract description 5
- 238000002156 mixing Methods 0.000 claims abstract description 4
- 238000006243 chemical reaction Methods 0.000 claims description 79
- 239000000243 solution Substances 0.000 claims description 68
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 42
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 36
- JJWLVOIRVHMVIS-UHFFFAOYSA-N isopropylamine Chemical group CC(C)N JJWLVOIRVHMVIS-UHFFFAOYSA-N 0.000 claims description 27
- NGVDGCNFYWLIFO-UHFFFAOYSA-N pyridoxal 5'-phosphate Chemical compound CC1=NC=C(COP(O)(O)=O)C(C=O)=C1O NGVDGCNFYWLIFO-UHFFFAOYSA-N 0.000 claims description 26
- 125000000217 alkyl group Chemical group 0.000 claims description 17
- 235000007682 pyridoxal 5'-phosphate Nutrition 0.000 claims description 13
- 239000011589 pyridoxal 5'-phosphate Substances 0.000 claims description 13
- 229960001327 pyridoxal phosphate Drugs 0.000 claims description 13
- 102000003929 Transaminases Human genes 0.000 claims description 12
- 108090000340 Transaminases Proteins 0.000 claims description 12
- HQABUPZFAYXKJW-UHFFFAOYSA-N butan-1-amine Chemical compound CCCCN HQABUPZFAYXKJW-UHFFFAOYSA-N 0.000 claims description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 9
- 125000003118 aryl group Chemical group 0.000 claims description 9
- 125000001153 fluoro group Chemical group F* 0.000 claims description 9
- 239000007788 liquid Substances 0.000 claims description 9
- 230000035484 reaction time Effects 0.000 claims description 9
- 125000001424 substituent group Chemical group 0.000 claims description 9
- 239000000337 buffer salt Substances 0.000 claims description 7
- 235000019000 fluorine Nutrition 0.000 claims description 7
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 claims description 6
- -1 C1~8Alkyl Chemical group 0.000 claims description 6
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 claims description 6
- 150000003973 alkyl amines Chemical class 0.000 claims description 6
- 150000003863 ammonium salts Chemical class 0.000 claims description 6
- 125000005842 heteroatom Chemical group 0.000 claims description 6
- 125000000623 heterocyclic group Chemical group 0.000 claims description 6
- 229910052739 hydrogen Inorganic materials 0.000 claims description 6
- 239000001257 hydrogen Substances 0.000 claims description 6
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 6
- 229910052757 nitrogen Inorganic materials 0.000 claims description 6
- 125000004433 nitrogen atom Chemical group N* 0.000 claims description 6
- 229910052760 oxygen Inorganic materials 0.000 claims description 6
- DPBLXKKOBLCELK-UHFFFAOYSA-N pentan-1-amine Chemical compound CCCCCN DPBLXKKOBLCELK-UHFFFAOYSA-N 0.000 claims description 6
- 229920006395 saturated elastomer Polymers 0.000 claims description 6
- 229910052717 sulfur Inorganic materials 0.000 claims description 6
- 125000002023 trifluoromethyl group Chemical group FC(F)(F)* 0.000 claims description 6
- 102000004625 Aspartate Aminotransferases Human genes 0.000 claims description 5
- 108010003415 Aspartate Aminotransferases Proteins 0.000 claims description 5
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 5
- 239000011737 fluorine Substances 0.000 claims description 5
- GSEJCLTVZPLZKY-UHFFFAOYSA-N Triethanolamine Chemical compound OCCN(CCO)CCO GSEJCLTVZPLZKY-UHFFFAOYSA-N 0.000 claims description 4
- 239000007983 Tris buffer Substances 0.000 claims description 4
- 125000003545 alkoxy group Chemical group 0.000 claims description 4
- 229910052731 fluorine Inorganic materials 0.000 claims description 4
- ZMJGSOSNSPKHNH-UHFFFAOYSA-N pyridoxamine 5'-phosphate Chemical compound CC1=NC=C(COP(O)(O)=O)C(CN)=C1O ZMJGSOSNSPKHNH-UHFFFAOYSA-N 0.000 claims description 4
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 claims description 4
- 125000003341 7 membered heterocyclic group Chemical group 0.000 claims description 3
- 239000004471 Glycine Substances 0.000 claims description 3
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 3
- 235000019270 ammonium chloride Nutrition 0.000 claims description 3
- VZTDIZULWFCMLS-UHFFFAOYSA-N ammonium formate Chemical group [NH4+].[O-]C=O VZTDIZULWFCMLS-UHFFFAOYSA-N 0.000 claims description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 claims description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 claims description 3
- 235000011130 ammonium sulphate Nutrition 0.000 claims description 3
- 125000006615 aromatic heterocyclic group Chemical group 0.000 claims description 3
- 125000004122 cyclic group Chemical group 0.000 claims description 3
- 229910052736 halogen Inorganic materials 0.000 claims description 3
- 150000002367 halogens Chemical class 0.000 claims description 3
- 125000001072 heteroaryl group Chemical group 0.000 claims description 3
- 150000002431 hydrogen Chemical group 0.000 claims description 3
- 125000004356 hydroxy functional group Chemical group O* 0.000 claims description 3
- 238000004519 manufacturing process Methods 0.000 claims description 3
- 125000004043 oxo group Chemical group O=* 0.000 claims description 3
- 229940100684 pentylamine Drugs 0.000 claims description 3
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 3
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 claims description 3
- WHOMFKWHIQZTHY-UHFFFAOYSA-L pyridoxine 5'-phosphate(2-) Chemical compound CC1=NC=C(COP([O-])([O-])=O)C(CO)=C1O WHOMFKWHIQZTHY-UHFFFAOYSA-L 0.000 claims description 3
- 125000000876 trifluoromethoxy group Chemical group FC(F)(F)O* 0.000 claims description 3
- 230000001476 alcoholic effect Effects 0.000 claims description 2
- 125000004849 alkoxymethyl group Chemical group 0.000 claims description 2
- HRYZPBSOAMTFEL-UHFFFAOYSA-N 1,3-diazinane-2,4,6-trione;sodium Chemical compound [Na].O=C1CC(=O)NC(=O)N1 HRYZPBSOAMTFEL-UHFFFAOYSA-N 0.000 claims 1
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical group [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 claims 1
- 229940111685 dibasic potassium phosphate Drugs 0.000 claims 1
- 229940111688 monobasic potassium phosphate Drugs 0.000 claims 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims 1
- 239000003960 organic solvent Substances 0.000 abstract description 12
- 230000007547 defect Effects 0.000 abstract description 7
- 238000009776 industrial production Methods 0.000 abstract description 4
- 239000002244 precipitate Substances 0.000 abstract description 2
- MFFMDFFZMYYVKS-SECBINFHSA-N sitagliptin Chemical compound C([C@H](CC(=O)N1CC=2N(C(=NN=2)C(F)(F)F)CC1)N)C1=CC(F)=C(F)C=C1F MFFMDFFZMYYVKS-SECBINFHSA-N 0.000 description 59
- 229960004034 sitagliptin Drugs 0.000 description 59
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 36
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 27
- 239000007864 aqueous solution Substances 0.000 description 11
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 9
- 238000001035 drying Methods 0.000 description 9
- 238000001914 filtration Methods 0.000 description 9
- 238000004537 pulping Methods 0.000 description 9
- 238000001514 detection method Methods 0.000 description 8
- 238000004128 high performance liquid chromatography Methods 0.000 description 8
- 238000001704 evaporation Methods 0.000 description 7
- 238000003756 stirring Methods 0.000 description 7
- 230000000052 comparative effect Effects 0.000 description 6
- 238000009792 diffusion process Methods 0.000 description 6
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 5
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 5
- MFFMDFFZMYYVKS-VIFPVBQESA-N (3s)-3-amino-1-[3-(trifluoromethyl)-6,8-dihydro-5h-[1,2,4]triazolo[4,3-a]pyrazin-7-yl]-4-(2,4,5-trifluorophenyl)butan-1-one Chemical compound C([C@@H](CC(=O)N1CC=2N(C(=NN=2)C(F)(F)F)CC1)N)C1=CC(F)=C(F)C=C1F MFFMDFFZMYYVKS-VIFPVBQESA-N 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 3
- 238000005054 agglomeration Methods 0.000 description 3
- 230000002776 aggregation Effects 0.000 description 3
- 239000007853 buffer solution Substances 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 230000003287 optical effect Effects 0.000 description 3
- 230000010355 oscillation Effects 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000001228 spectrum Methods 0.000 description 3
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 229940125717 barbiturate Drugs 0.000 description 2
- HNYOPLTXPVRDBG-UHFFFAOYSA-N barbituric acid Chemical compound O=C1CC(=O)NC(=O)N1 HNYOPLTXPVRDBG-UHFFFAOYSA-N 0.000 description 2
- 238000006555 catalytic reaction Methods 0.000 description 2
- 230000002349 favourable effect Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 238000001308 synthesis method Methods 0.000 description 2
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 238000005576 amination reaction Methods 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000007810 chemical reaction solvent Substances 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- 125000004093 cyano group Chemical group *C#N 0.000 description 1
- 230000032798 delamination Effects 0.000 description 1
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000011031 large-scale manufacturing process Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 150000002894 organic compounds Chemical class 0.000 description 1
- 230000001376 precipitating effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 239000013558 reference substance Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
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- 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
- C12P41/00—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture
- C12P41/006—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture by reactions involving C-N bonds, e.g. nitriles, amides, hydantoins, carbamates, lactames, transamination reactions, or keto group formation from racemic mixtures
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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
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/18—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing at least two hetero rings condensed among themselves or condensed with a common carbocyclic ring system, e.g. rifamycin
- C12P17/182—Heterocyclic compounds containing nitrogen atoms as the only ring heteroatoms in the condensed system
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Abstract
The invention provides a preparation method of a chiral beta-amino acid derivative. The preparation method also comprises the following steps: s1, mixing the first solvent, the biological enzyme catalyst and the amino donor to form a mixed raw material solution; the first solvent is an alcohol solvent or a mixed solvent of water and the alcohol solvent; s2, adding a solid particle substrate A into the mixed raw material solution in a continuous solid stream mode through a continuous feeding device to carry out enzymatic reaction, and obtaining the chiral beta-amino acid derivative. By applying the technical scheme of the invention, the defect that the substrate is dissolved in the organic solvent and then is dripped is effectively overcome, the problem that the substrate is easy to agglomerate and precipitate in the enzymatic reaction process is solved, and the yield of the chiral beta-amino acid derivative is improved. Moreover, the preparation method is simpler and more convenient, the raw materials are more easily obtained, the repeatability is better, and the industrial production prospect is better.
Description
Technical Field
The invention relates to the technical field of synthesis of organic compounds, in particular to a preparation method of a chiral beta-amino acid derivative.
Background
Chiral beta-amino acid derivatives are of interest to many scientists as important drugs. The synthesis methods of chiral β amino acid derivatives are many, and the synthesis method using an enzyme as a catalyst is particularly concerned by those skilled in the art due to the characteristics of mild conditions, little pollution and the like. Chinese patent CN108586346B and chinese patent CN102405281B are two typical patents for preparing chiral β amino acid derivatives using enzyme catalysis.
Both CN108586346B and CN102405281B report that the substrate is dissolved by an organic solvent and then fed to a reaction system comprising an amino donor and an enzyme catalyst in the form of a solution to react, wherein the concentration of the substrate is 100g/L-250 g/L. In the field of synthesizing small molecular organic matters, the reaction substrate is generally regarded as being more beneficial to the stability of a reaction system in a form of dropwise adding of solution. However, according to the embodiments in the above patent, a plurality of similar experiments were carried out in combination with practical circumstances, and it was found that when a chiral β -amino acid derivative is prepared, precipitation of a substrate in a reaction system is caused by dropping a substrate solution, and then agglomeration is generated to cause difficulty in complete reaction of an enzymatic reaction, thereby reducing the yield of the chiral β -amino acid derivative.
Disclosure of Invention
The invention mainly aims to provide a preparation method of a chiral beta-amino acid derivative, which aims to solve the problems that in the prior art, when the chiral beta-amino acid derivative is prepared, a substrate is easy to precipitate in a reaction system in a manner of dropwise adding a substrate solution, and then, enzymatic reaction is difficult to completely react due to caking, so that the yield of the beta-amino acid derivative is reduced.
In order to achieve the above objects, according to one aspect of the present invention, there is provided a method for preparing a chiral β -amino acid derivative. The chiral beta-amino acid derivative has a structure shown in the following general formula I:
in formula I, the stereogenic center marked with x has the (R) -or (S) -configuration; an enantiomeric excess of at least 70% compared to the corresponding enantiomer; z is OR2、SR2Or NR2R3(ii) a R is C1～8Alkyl, aryl, heteroaryl, aryl-C1～2Alkyl or heteroaryl-C1～2An alkyl group; r2And R3Each independently is hydrogen, C1～8Alkyl, aryl or aryl-C1～2An alkyl group; or R2And R3Together with the nitrogen atom to which they are attached form a ring optionally containing a nitrogen atom selected from O, S, NH and NC1～4A 4-to 7-membered heterocyclic ring system of other heteroatoms of the alkyl radical, the heterocyclic ring being unsubstituted or independently selected from oxo, hydroxy, halogen, C1～4Alkoxy and C1～4Alkyl substituted with 1-3 substituents, wherein alkyl and alkoxy are unsubstituted or substituted with 1-5 fluorines; and the heterocyclic ring system is optionally combined with a 5-to 6-membered saturated or aromatic carbocyclic ring system or contains a ring selected from O, S and NC0～4Fused to 5-to 6-membered saturated or aromatic heterocyclic ring systems of 1-2 heteroatoms of the alkyl radical, the fused ring systems being unsubstituted or selected from hydroxy, amino, fluoro, C1～4Alkyl radical, C1～4Alkoxy and trifluoromethyl substituted with 1-2 substituents; wherein, the preparation method also comprises the following steps: s1, mixing the first solvent, the biological enzyme catalyst and the amino donorMixing to form a mixed raw material solution; the first solvent is an alcohol solvent or a mixed solvent of water and the alcohol solvent; s2, adding a solid particle substrate A in the form of continuous solid stream into the mixed raw material solution through a continuous feeding device for enzymatic reaction, wherein the structural formula of the solid particle substrate A is shown in the specification
R and Z have the same definitions as before, to give chiral β -amino acid derivatives.
Further, the adding speed of the solid particle substrate A is 0.5-12.0 g/s; preferably, during the process of adding the solid particle substrate A, the continuous solid stream is controlled to be a plurality of strands, and the strands are independent and do not interfere with each other; more preferably, the plurality of continuous solid streams are controlled to oscillate relative to the liquid surface as they enter the mixed feedstock solution during the addition of the solid particulate substrate a.
Further, the adding time of the solid particle substrate A is 4-12 h; preferably, the adding time of the solid particle substrate A is 6-10 h.
Further, the addition amount of the solid particle substrate A is 80-500 g per liter of the first solvent.
Furthermore, the molar ratio of the solid particle substrate A to the amino donor is 1 (3-10).
Further, the mixed raw material solution also comprises a buffer salt system.
Further, the buffer salt system is one or more of potassium dihydrogen phosphate/dipotassium hydrogen phosphate, Tris/hydrochloric acid, triethanolamine, barbiturate sodium/hydrochloric acid, glycine/sodium hydroxide or amino donor/hydrochloric acid.
Further, the alcohol solvent is selected from one or more of methanol, ethanol and propanol.
Further, the first solvent is a mixed solvent of an alcohol solvent and water, wherein the mass fraction of the alcohol solvent is 20-40%.
Further, the amino donor is selected from C1～6One or more of alkylamine and inorganic ammonium salt; preferably, C1～6The alkylamine is isopropylamine, butylamine orA pentylamine; preferably, the inorganic ammonium salt is ammonium formate, ammonium chloride or ammonium sulfate.
Further, the mixed raw material solution also comprises an amino transfersome, and the amino transfersome is selected from one or more of pyridoxal phosphate, pyridoxamine phosphate or pyridoxine phosphate.
Further, the bio-enzyme catalyst is transaminase; the transaminase is selected from one or more of aspartate aminotransferase (AspATs), aromatic aminotransferase (AroATs), omega-transaminase (omega-ATs), branched-chain transaminase (BCATs), and D-transaminase (DATs).
Further, in the enzymatic reaction process, the reaction temperature is 20-60 ℃, the reaction pH value is 7.0-11, and the reaction time is 18-36 h; preferably, the reaction temperature is 35-55 ℃, and the reaction pH value is 8.0-10.5.
Further, R is aryl-C1～2Alkyl, wherein aryl-C1～2The phenyl group of the alkyl group is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of fluorine, trifluoromethyl and trifluoromethoxy; z is NR2R3；NR2R3Has the structural formula
R4Is hydrogen or C which is unsubstituted or substituted by 1 to 5 fluorine1～4An alkyl group.
The technical scheme of the invention effectively improves the defect that the substrate is dissolved in the organic solvent and then is dripped, overcomes the technical prejudice that the dripping of the substrate into the solution is more favorable for reaction stability, utilizes the special reaction solvent-alcohol solvent to be matched with a special feeding mode, more effectively improves the defect that the substrate is dissolved in the organic solvent and then is dripped, solves the problem that the substrate is easy to be separated out in a conglobation manner in the enzymatic reaction process, and promotes the yield of the chiral beta-amino acid derivative to be higher. Moreover, the preparation method is simpler and more convenient, the raw materials are more easily obtained, the repeatability is better, and the industrial production prospect is better. The main object of the present invention is to provide a solution to the problems of the prior art.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this application, illustrate embodiments of the invention and, together with the description, serve to explain the invention and not to limit the invention. In the drawings:
FIG. 1 shows a spectrum of a reference substance for detecting the purity of sitagliptin prepared in example 1 of the invention; and
fig. 2 shows a spectrum of a sitagliptin optical isomer detection control prepared in example 1 of the present invention.
Detailed Description
It should be noted that the embodiments and features of the embodiments in the present application may be combined with each other without conflict. The present invention will be described in detail below with reference to the embodiments with reference to the attached drawings.
As described in the background section, in the prior art, when chiral beta-amino acid derivatives are prepared, a substrate solution is dripped to cause the substrate to be separated out in a reaction system, and then agglomeration is generated to cause the enzymatic reaction to be difficult to completely react, so that the yield of the beta-amino acid derivatives is reduced. In order to solve the problem, the invention provides a preparation method of a chiral beta-amino acid derivative, wherein the chiral beta-amino acid derivative has a structure shown in the following general formula I:
in formula I, the stereogenic center marked with x has the (R) -or (S) -configuration; an enantiomeric excess of at least 70% compared to the corresponding enantiomer; z is OR2、SR2Or NR2R3(ii) a R is C1～8Alkyl, aryl, heteroaryl, aryl-C1～2Alkyl or heteroaryl-C1～2An alkyl group; promote more solid particle substrate A to be converted into chiral beta amino acid derivative R2And R3Each independently is hydrogen, C1～8Alkyl, aryl or aryl-C1～2An alkyl group; or R2And R3Together with the nitrogen atom to which they are attached form a ring optionally containing a nitrogen atom selected from O, S, NH and NC1～4Of alkyl groupsA 4-to 7-membered heterocyclic ring system of further hetero atoms, the heterocyclic ring being unsubstituted or independently selected from oxo, hydroxy, halogen, C1～4Alkoxy and C1～4Alkyl substituted with 1-3 substituents, wherein alkyl and alkoxy are unsubstituted or substituted with 1-5 fluorines; and the heterocyclic ring system is optionally combined with a 5-to 6-membered saturated or aromatic carbocyclic ring system or contains a ring selected from O, S and NC0～4Condensed with 5-to 6-membered saturated or aromatic heterocyclic ring systems of 1-2 heteroatoms of the alkyl radical, the condensed ring systems being unsubstituted or selected from hydroxy, amino, fluoro, C1～4Alkyl radical, C1～41-2 substituent substitutions of alkoxy and trifluoromethyl; wherein, the preparation method also comprises the following steps: s1, mixing a first solvent, a biological enzyme catalyst and an amino donor to form a mixed raw material solution, wherein the first solvent is an alcohol solvent or a mixed solvent of water and the alcohol solvent; s2, adding a solid particle substrate A in the form of continuous solid stream into the mixed raw material solution through a continuous feeding device for enzymatic reaction, wherein the structural formula of the solid particle substrate A is shown in the specification
R and Z have the same definitions as before, giving chiral beta-amino acid derivatives.
Specifically, the enzymatic reaction is as follows:
according to the invention, the first solvent, the biological enzyme catalyst and the amino donor are mixed to form a mixed raw material solution, so that a more stable and uniform reaction environment is provided for the subsequent solid particle substrate A, and the subsequent enzymatic reaction is promoted to be more stable. And then adding the solid particle substrate A into the mixed raw material solution in a continuous solid stream mode through a continuous feeding device to carry out enzymatic reaction. On one hand, the alcohol solvent is used as the reaction solvent of the chiral beta-amino acid derivative, and compared with the condition that the solid particle substrate A is prepared into a solution and is dripped into the mixed raw material solution of the alcohol solvent, the solid particle substrate A is added in a continuous solid stream mode, so that the diffusion speed of the solid particle substrate A after entering the alcohol solvent can be better controlled, and the enzymatic reaction can be gradually completed in the gradual diffusion process. Moreover, due to the specific surface tension of the solid particle substrate A in the reaction system of the alcohol solvent, the solid particle substrate A can be relatively stably kept at the position on the upper part of the reaction system, and the obtained enzymatic product chiral beta-amino acid derivative can gradually approach the lower part of the reaction system of the alcohol solvent along with the gradual diffusion and reaction of molecules of the solid particle substrate A, so that the time difference and the space difference of the gradual diffusion and reaction of the solid particle substrate A and the gradual diffusion of the product are formed, the enzymatic reaction of the invention is more stable in the process, is not easy to agglomerate, and has higher reaction yield.
In conclusion, the preparation method of the chiral beta-amino acid derivative overcomes the technical prejudice that the dropwise addition of the substrate into the solution is more favorable for the reaction stability, utilizes the special reaction solvent-alcohol solvent to be matched with a special charging mode, more effectively overcomes the defect that the substrate is dissolved into the organic solvent and then is dropwise added, overcomes the defect that the substrate is easy to agglomerate and separate out in the enzymatic reaction process, and promotes the higher yield of the chiral beta-amino acid derivative. Moreover, the preparation method is simpler and more convenient, the raw materials are more easily obtained, the repeatability is better, and the industrial production prospect is better.
Preferably, the adding speed of the solid particle substrate A is 0.5-12.0 g/s; in the process of adding the solid particle substrate A, controlling the continuous solid stream to be a plurality of strands, wherein the strands are independent and do not interfere with each other; more preferably, the plurality of continuous solid streams are controlled to be in a rotating state relative to the liquid surface when entering the mixed raw material solution during the process of adding the solid particle substrate A. Therefore, the dispersion state of the solid particle substrate A in the mixed raw material solution can be greatly increased, the solid particle substrate A is fully positioned in each direction of the upper position of the alcohol solvent, the solid particle substrate A is promoted to be better dispersed in the mixed raw material solution, the solid particle substrate A in each position is promoted to participate in the enzymatic reaction, better distribution is formed in the space and the diffusion speed of a target product, on one hand, the enzymatic reaction can be continuously maintained to be forward, the enzymatic reaction is more complete, the yield of the chiral beta-amino acid derivative is higher, on the other hand, a more proper distance is kept between reaction products, and the caking problem is better solved.
In order to further improve the yield of the chiral beta-amino acid derivative, the addition time of the solid particle substrate A is 4-12 h. More preferably, the adding time of the solid particle substrate A is 6-10 h.
Preferably, the solid particulate substrate A is added in an amount of 80 to 500g per litre of first solvent. Within the range, the solid particle substrate A has better solubility in the first solvent, and the better solubility can more effectively avoid the solid particle substrate A from precipitating and agglomerating in the mixed raw material solution, so that the solid particle substrate A can be more fully contacted with the catalyst and can react, the enzymatic reaction is more complete, more solid particle substrate A is converted into chiral beta-amino acid derivatives, and the yield of the chiral beta-amino acid derivatives is higher.
In order to further improve the yield of the chiral beta-amino acid derivative, the molar ratio of the solid particle substrate A to the amino donor is 1 (3-10). Within this range, the solid particulate substrate a and the amino donor react more completely, more of the solid particulate substrate a is converted to the chiral β -amino acid derivative, and the yield of the chiral β -amino acid derivative is higher.
In a preferred embodiment, the mixed raw material solution further comprises a buffer salt system; the buffer salt system is one or more of potassium dihydrogen phosphate/dipotassium hydrogen phosphate, Tris/hydrochloric acid, triethanolamine, barbiturate sodium/hydrochloric acid, glycine/sodium hydroxide or amino donor/hydrochloric acid. The buffer system can keep the pH value of the reaction solution system in a range more suitable for the enzymatic reaction, and promote the enzyme catalyst to have better catalytic activity, better stability and higher stereoselectivity. Simultaneously, the enzymatic reaction is more complete, and the yield of the chiral beta-amino acid derivative is higher.
Preferably, the alcoholic solvent is one or more of methanol, ethanol, propanol. For the purpose of promoting better solubility of the solid particle substrate A in the first solvent, in a preferred embodiment, the first solvent is a mixed solvent of water and an alcohol solvent, and the mass fraction of the alcohol solvent is 20-40%.
In a preferred embodiment, the amino donor is preferably selected from C1～6One or more of alkylamine and inorganic ammonium salt can have good amination effect under the condition of low addition amount, so that more solid particle substrates A are promoted to be converted into chiral beta amino acid derivatives, and the yield of the chiral beta amino acid derivatives is higher. At the same time, the carbonyl by-product formed by the enzymatic reaction is more easily removed based on the above-mentioned amino donor. More preferably, C1～6The alkylamine is isopropylamine, butylamine or pentylamine, and the inorganic ammonium salt is ammonium formate, ammonium chloride or ammonium sulfate. The amino donor is lower in cost and has the possibility of large-scale production.
For the purpose of improving the effect of the amino donor on amino transfer, it is preferable that the mixed raw material solution further includes an amino mediator selected from one or more of pyridoxal phosphate, pyridoxamine phosphate, and pyridoxine phosphate.
In a preferred embodiment, the bio-enzyme catalyst is a transaminase; the transaminase is selected from one or more of aspartate aminotransferase (AspATs), aromatic aminotransferase (AroATs), omega-transaminase (omega-ATs), branched-chain transaminase (BCATs), and D-transaminase (DATs). The enzyme catalysis activity is better, and the stability is better.
Preferably, in the enzymatic reaction process, the reaction temperature is 20-60 ℃, the reaction pH value is 7.0-11, and the reaction time is 18-36 hours; preferably, the reaction temperature is 35-55 ℃, and the reaction pH value is 8.0-10.5. Under the reaction condition, the enzymatic reaction is more stable and complete, and the yield of the chiral beta amino acid derivative is higher.
In a preferred embodiment, R is aryl-C1～2Alkyl, wherein aryl-C1～2The phenyl group of the alkyl group is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of fluorine, trifluoromethyl and trifluoromethoxy; z is NR2R3；NR2R3Has the structural formula
R4Is hydrogen or unsubstituted or substituted by 1 to 5Fluorine substituted C1～4An alkyl group. For example, the chiral beta-amino acid derivative is sitagliptin with the structural formula
The present application is described in further detail below with reference to specific examples, which should not be construed as limiting the scope of the invention as claimed.
And (3) performance characterization:
the purity detection and central control method comprises the following steps:
the instrument comprises the following steps: high performance liquid chromatograph
A detector: an ultraviolet detector; and (3) chromatographic column: discovery Cyano, 150X 4.6mm, 5 μm; detection wavelength: 205 nm; column temperature: 30 ℃; flow rate: 1.0 ml/min; sample introduction volume: 20 mu L of the solution; operating time: 25 min; sample introduction concentration: 0.1 mg/ml;
buffer salt solution: potassium dihydrogen phosphate (1.36 g) was dissolved in 1000ml of water, and the solution was adjusted to pH 2.0 with phosphoric acid.
Mobile phase: buffer solution: acetonitrile 85:15
Fig. 1 shows a sitagliptin purity detection profile prepared in example 1 of the present invention.
(II) an optical isomer detection method:
the instrument comprises the following steps: high performance liquid chromatograph
A chromatographic column: chiralpak AD 4.6mm × 25cm, 5 μm or equivalent column; sample introduction volume: 10 mu l of the mixture; flow rate: 0.8 ml/min; column temperature: 35 ℃; detection wavelength: 268 nm; operating time: and 20 min.
Mobile phase: anhydrous ethanol: n-heptane: diethylamine: water 600: 400: 1: 1
Fig. 2 shows a detection spectrum of sitagliptin optical isomers prepared in example 1 of the present invention.
Example 1
Adding a first solvent (150L methanol and 210L water), 60kg isopropylamine, 100g potassium dihydrogen phosphate and 500g dipotassium hydrogen phosphate into a reaction kettle respectively, adjusting the pH of the solution to 9.0 by using hydrochloric acid, and adding 90L (5.0 g/L) (R) -omega-ATs-117 enzyme solution and 1kg pyridoxal phosphate;
the reaction kettle was warmed to 45 ℃ and stirred while the sitagliptin substrate was added as a continuous solid stream in a state of oscillation relative to the liquid surface. Wherein the adding speed of the sitagliptin substrate is 12g/s, the total feeding time is 10h, the adding amount of the sitagliptin substrate is 88g for each liter of the first solvent, the pH value in the enzymatic reaction process is maintained at 9.0 by using 4mol/L of isopropylamine aqueous solution, and the reaction time is 20 h. The reaction was monitored by HPLC with a medium control, and after completion of the reaction, the sitagliptin content was 87%, corresponding to which the (S) -configurational isomer content was 1.5%. Extracting with 500L ethyl acetate, evaporating to remove organic solvent, adding 200kg toluene, pulping, filtering, and drying to obtain 35.1kg sitagliptin with yield of 82%.
Wherein the molar ratio of sitagliptin substrate to isopropylamine is 1: 10.
example 2
Adding a first solvent (25L methanol and 16L water), 4kg isopropylamine, 16L Tris/hydrochloric acid buffer solution with pH of 8.0 into a reaction kettle respectively, adjusting the pH of the solution to 8.0 by using hydrochloric acid, and adding 9L (2.5 g/L) (R) -omega-ATs-117 enzyme solution and 1kg pyridoxal phosphate;
the reaction kettle is placed in an oil bath at 30 ℃ for stirring, and meanwhile, the sitagliptin substrate is added in a continuous solid stream mode, wherein the adding speed of the sitagliptin substrate is 2g/s, the total adding time of the streams is 6h, the adding amount of the sitagliptin substrate is 83g for each liter of first solvent, 4mol/L of isopropylamine aqueous solution is used for maintaining the pH value of the enzymatic reaction process at 8.0, and the reaction time is 24. The reaction was monitored by HPLC with a central control, and after completion of the reaction, the sitagliptin content was 85% and the corresponding (S) -configurational isomer content was 1.0%. Extracting with 50L ethyl acetate, evaporating to remove organic solvent, adding 20kg toluene, pulping, filtering, and drying to obtain 3.5kg sitagliptin with yield of 80%.
Wherein the molar ratio of the sitagliptin substrate to the isopropylamine is 1: 6.
Example 3
Adding a first solvent (50ml of methanol and 20ml of water) and 60g of butylamine into a reaction flask respectively, adjusting the pH of the solution to 8.5 by using hydrochloric acid, and adding 90ml of 5.0g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxal phosphate;
and (3) placing the reaction bottle in a 45 ℃ oil bath for stirring, and simultaneously adding the sitagliptin substrate in a continuous solid stream mode, wherein the adding speed of the sitagliptin substrate is 0.5g/s, the total adding time of the sitagliptin substrate is 6h, the adding amount of the sitagliptin substrate is 504g for each liter of the first solvent, 4mol/L of isopropylamine aqueous solution is used for maintaining the pH value of the enzymatic reaction process at 8.5, and the reaction time is 30 h. The reaction was monitored by HPLC with a central control, and after completion of the reaction, the sitagliptin content was 80% and the corresponding (S) -configurational isomer content was 2.1%. Extracting with 500mL ethyl acetate, evaporating to remove organic solvent, adding 1kg toluene, pulping, filtering, and drying to obtain 861.8g sitagliptin with yield of 80%.
Wherein the molar ratio of the sitagliptin substrate to the butylamine is 1: 3.3.
Example 4
Adding a first solvent (150L methanol and 210L water), 60kg isopropylamine, 100g potassium dihydrogen phosphate and 500g dipotassium hydrogen phosphate into a reaction kettle respectively, adjusting the pH of the solution to 9.0 by using hydrochloric acid, and adding 90L (5.0 g/L) (R) -omega-ATs-117 enzyme solution and 1kg pyridoxal phosphate;
the reaction kettle was warmed to 45 ℃ and stirred while adding the sitagliptin substrate as a continuous solid stream in an oscillating state relative to the liquid surface. Wherein the adding speed of the sitagliptin substrate is 12g/s, the total feeding time is 12h, the adding amount of the sitagliptin substrate is 103g for each liter of the first solvent, the pH value is maintained at 9.0 in the enzymatic reaction process by using 4mol/L isopropylamine aqueous solution, and the reaction time is 20 h. The reaction was monitored by HPLC with a medium control, and after completion of the reaction, the sitagliptin content was 88%, corresponding to which the (S) -configurational isomer content was 1.6%. Extracting with 500L ethyl acetate, evaporating to remove organic solvent, adding 200kg toluene, pulping, filtering, and drying to obtain 42.9kg sitagliptin with yield of 83%.
Wherein the molar ratio of sitagliptin substrate to isopropylamine is 1: 10.
example 5
Adding a first solvent (150L methanol and 210L water), 60kg isopropylamine into a reaction kettle, adjusting the pH of the solution to 9.0 by using hydrochloric acid, and adding 90L 10.0g/L (S) -omega-ATs-Vf enzyme solution and 1kg pyridoxal phosphate;
the reaction kettle was warmed to 50 ℃ and stirred while the sitagliptin substrate was added as a continuous solid stream in a state of oscillation relative to the liquid surface. Wherein the adding speed of the sitagliptin substrate is 12g/s, the total feeding time is 12h, the adding amount of the sitagliptin substrate is 103g for each liter of the first solvent, the pH value in the enzymatic reaction process is maintained at 9.0 by using 4mol/L of isopropylamine aqueous solution, and the reaction time is 24 h. The reaction was monitored by HPLC with a medium control, and after completion of the reaction, the content of (S) -sitagliptin was 71%, and the content of the corresponding (R) -configurational isomer was 3.4%. Extracting with 500L ethyl acetate, evaporating to remove organic solvent, adding 100kg toluene, pulping, filtering, and drying to obtain 16.5kg (S) -sitagliptin with yield of 32%.
Example 6
Adding a first solvent (1L methanol and 2L water), 600g isopropylamine into a reaction flask, adjusting the pH of the solution to 8.0 by using hydrochloric acid, and adding 500mL of 10.0g/L (S) -omega-ATs-113 enzyme solution and 1g pyridoxal phosphate;
the reaction flask was warmed to 45 ℃ and stirred while the sitagliptin substrate was added as a continuous solid stream in a state of oscillation relative to the liquid surface. Wherein the adding speed of the sitagliptin substrate is 2g/s, the total feeding time is 1h, the adding amount of the sitagliptin substrate is 180g for each liter of the first solvent, the pH value in the enzymatic reaction process is maintained at 9.0 by using 4mol/L of isopropylamine aqueous solution, and the reaction time is 30 h. The reaction was monitored by HPLC with a medium control, and after completion of the reaction, the content of (S) -sitagliptin was 70%, and the content of the corresponding (R) -configurational isomer was 4.6%. Extracting with 5L ethyl acetate, evaporating to remove organic solvent, adding 1kg toluene, pulping, filtering, and drying to obtain 275.9g of (S) -sitagliptin with a yield of 38%.
Comparative example 1
Adding 50ml of methanol, 360ml of water and 60g of isopropylamine into a reaction flask respectively, adjusting the pH of the solution to 8.5 by using hydrochloric acid, and adding 90ml of 5g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxal phosphate;
and (3) placing the reaction bottle in an oil bath at 45 ℃ for stirring, and simultaneously slowly adding sitagliptin substrate solution, wherein the total flow adding time is 8h, the sitagliptin substrate solution is 100ml of methanol for dissolving 200.0g of sitagliptin substrate, and 4mol/L of isopropylamine aqueous solution is used for maintaining the pH value to be 8.5 in the enzymatic reaction process.
After 6 hours of reaction, the solid is separated out from the reaction flask, and the solid is detected by HPLC, and the separated solid is confirmed to be a substrate and to have an agglomeration phenomenon. After completion of the reaction, the sitagliptin content was 26%, corresponding to which the (S) -configurational isomer content was 96%. Due to the low sitagliptin content, sitagliptin cannot be obtained by post-treatment.
Comparative example 2
Adding 360ml of water and 60g of isopropylamine into a reaction flask respectively, adjusting the pH of the solution to 8.5 by using hydrochloric acid, and adding 90ml of 5g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxal phosphate;
and (3) placing the reaction bottle in an oil bath at 45 ℃ for stirring, and simultaneously slowly adding a sitagliptin substrate solution for 8 hours in total, wherein the sitagliptin substrate solution is prepared by dissolving 200.0g of sitagliptin substrate in 50ml of methanol, and 4mol/L of isopropylamine aqueous solution is used for maintaining the pH value of the enzymatic reaction process at 8.5.
After 4 hours of reaction, a solid was separated out from the reaction flask, and the solid was confirmed to be a substrate by HPLC detection and to be agglomerated. After completion of the reaction, the sitagliptin content was 12%, and the corresponding (S) -configurational isomer content was 96%. Due to the low sitagliptin content, sitagliptin cannot be obtained by post-treatment.
Comparative example 3
100ml of DMSO, 300ml of water, 60g of isopropylamine and 10g of triethanolamine are added into a 1L reaction bottle, the pH is adjusted to 10.0 by using hydrochloric acid, then 90ml of 2.5g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxal phosphate are added into an oil bath at 55 ℃, stirring is carried out, 30.0g of sitagliptin substrate is slowly added, the total flow adding time is 4h, the pH is maintained at about 10.0 by using 4mol/L of isopropylamine aqueous solution in the flow adding process, the reaction condition is monitored by central control, the content of the sitagliptin is 83%, and the content of the corresponding (S) -configurational isomer is 97%. And after the reaction is finished, adding 1.5L of ethyl acetate for extraction, evaporating to remove the ethyl acetate after layering, adding 1L of toluene for pulping and crystallizing, filtering and drying to obtain 12.87g of sitagliptin with the yield of 43%.
Comparative example 4
Adding 50ml of DMSO, 360ml of water and 60g of isopropylamine into a 1L reaction bottle, adjusting the pH to 8.5 by using hydrochloric acid, adding 90ml of 5g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxamine phosphate, putting the mixture into a 45 ℃ oil bath, stirring, slowly adding 200.0g of sitagliptin substrate, adding the mixture for 8 hours in total, maintaining the pH at about 8.5 by using 4mol/L of isopropylamine aqueous solution in the process of adding the mixture, monitoring the reaction condition by a central control system, and after the reaction is finished, keeping the content of the sitagliptin 86% and the content of the corresponding (S) -configurational isomer at 96%. After the reaction is finished, 1.5L of ethyl acetate is added for extraction, ethyl acetate is evaporated after delamination, 1.5L of toluene is added for pulping and crystallization, and 83.79g of sitagliptin is obtained after filtration and drying, with the yield of 42%.
Comparative example 5
Adding 360ml of water and 60g of isopropylamine into a reaction flask respectively, adjusting the pH of the solution to 8.5 by using hydrochloric acid, and adding 90ml of 5g/L (R) -omega-ATs-117 enzyme solution and 1g of pyridoxal phosphate;
placing the reaction bottle in an oil bath at 45 ℃ for stirring, and simultaneously slowly adding a sitagliptin substrate solution, wherein the total feeding time is 8h, the sitagliptin substrate solution is 50ml DMSO for dissolving 200.0g of sitagliptin substrate, 4mol/L of isopropylamine aqueous solution is used for maintaining the pH value at 8.5 in the enzymatic reaction process, the reaction condition is monitored by a central control, after the reaction is completed, the content of sitagliptin is 84%, and the content of the corresponding (S) -configurational isomer is 97%. And after the reaction is finished, adding 1L of ethyl acetate for extraction, distilling to remove the ethyl acetate after layering, adding 1L of toluene for pulping and crystallizing, filtering and drying to obtain 77.81g of sitagliptin with the yield of 39%.
From the above description, it can be seen that the above-described embodiments of the present invention achieve the following technical effects:
as can be seen from the data of examples 1 to 4 and comparative examples 1 to 5, the first solvent, the bio-enzyme catalyst and the amino donor are mixed to form a mixed raw material solution, so as to provide a more stable and uniform reaction environment for the subsequent solid particle substrate A and promote a more stable subsequent enzymatic reaction. And then adding the solid particle substrate A into the mixed raw material solution in a continuous solid stream mode through a continuous feeding device to carry out enzymatic reaction. On the one hand, the content of the solid particle substrate A in the mixed raw material solution is controlled more effectively, the content change of the solid particle substrate A in the unit volume of the mixed raw material solution is enabled to be more stable, the content of the solid particle substrate A in the unit volume of the mixed raw material solution can be better kept to be more soluble, the solubility of the solid particle substrate A in the mixed raw material solution is enabled to be better, the solid particle substrate A is prevented from being separated out and agglomerated in the mixed raw material solution more effectively due to good solubility, enzymatic reaction is enabled to be more complete, more solid particle substrate A is converted into chiral beta-amino acid derivatives, and the yield of the chiral beta-amino acid derivatives is higher. On the other hand, the contact between the solid particle substrate A and the mixed raw material solution is greatly increased, so that more solid particle substrates A are promoted to participate in the enzymatic reaction as much as possible, the enzymatic reaction can continuously keep going forward, the enzymatic reaction is more complete, more solid particle substrates A are converted into chiral beta-amino acid derivatives, and the yield of the chiral beta-amino acid derivatives is higher.
In conclusion, the preparation method of the chiral beta-amino acid derivative effectively overcomes the defect that a substrate is dissolved in an organic solvent and then is dripped, overcomes the defect that the substrate is easy to agglomerate and precipitate in the enzymatic reaction process, and promotes higher yield of the chiral beta-amino acid derivative. Moreover, the preparation method is simpler and more convenient, the raw materials are more easily obtained, the repeatability is better, and the industrial production prospect is better.
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (14)
1. A method for preparing a chiral beta-amino acid derivative, wherein the chiral beta-amino acid derivative has a structure represented by the following general formula I:
in the general formula I, the stereogenic center marked by has (R) -or (S) -configuration; an enantiomeric excess of at least 70% compared to the corresponding enantiomer;
z is OR2、SR2Or NR2R3；
R is C1～8Alkyl, aryl, heteroaryl, aryl-C1～2Alkyl or heteroaryl-C1～2An alkyl group;
R2and R3Each independently is hydrogen, C1～8Alkyl, aryl or aryl-C1～2An alkyl group; or R2And R3Together with the nitrogen atom to which they are attached form a ring optionally containing a nitrogen atom selected from O, S, NH and NC1～44-to 7-membered heterocyclic ring system of further heteroatoms of alkyl, said heterocyclic ring being unsubstituted or being independently selected from oxo, hydroxy, halogen, C1～4Alkoxy and C1～4Alkyl substituted with 1-3 substituents, wherein alkyl and alkoxy are unsubstituted or substituted with 1-5 fluorines; and said heterocyclic ring system optionally being associated with a 5-to 6-membered saturated or aromatic carbocyclic ring system or containing a ring selected from O, S and NC0～4Fused to 5-to 6-membered saturated or aromatic heterocyclic ring systems of 1-2 heteroatoms of the alkyl radical, the fused ring systems being unsubstituted or selected from hydroxy, amino, fluoro, C1～4Alkyl radical, C1～4Alkoxy and trifluoromethyl substituted with 1-2 substituents;
wherein, the preparation method also comprises the following steps:
s1, mixing the first solvent, the biological enzyme catalyst and the amino donor to form a mixed raw material solution; the first solvent is an alcohol solvent or a mixed solvent of water and the alcohol solvent;
s2, adding a solid particle substrate A in the form of continuous solid stream into the mixed raw material solution through a continuous feeding device for enzymatic reaction, wherein the structural formula of the solid particle substrate A is shown in the specification
R and Z have the same definitions as described hereinbefore, giving the chiral β -amino acid derivative.
2. The method for preparing a chiral β -amino acid derivative according to claim 1, wherein the solid particulate substrate a is added at a rate of 0.5 to 12.0 g/s; preferably, the continuous solid stream is controlled to be a plurality of strands in the process of adding the solid particle substrate A, and the plurality of strands are independent and do not interfere with each other; more preferably, the plurality of continuous solid streams are controlled to oscillate relative to the liquid level as they enter the mixed feedstock solution during the addition of the solid particulate substrate a.
3. The method for preparing chiral beta-amino acid derivatives according to claim 2, wherein the solid particulate substrate A is added for 4-12 h; preferably, the adding time of the solid particle substrate A is 6-10 h.
4. The process for the preparation of a chiral β -amino acid derivative according to claim 1, wherein the solid particulate substrate a is added in an amount of 80 to 500g per liter of the first solvent.
5. The method for preparing a chiral beta-amino acid derivative according to any one of claims 1 to 4, wherein the molar ratio of the solid particulate substrate A to the amino donor is 1 (3-10).
6. The method for producing a chiral β -amino acid derivative according to any one of claims 1 to 4, wherein the mixed raw material solution further comprises a buffer salt system.
7. The method of claim 6, wherein the buffer salt system is one or more of monobasic potassium phosphate/dibasic potassium phosphate, Tris/hydrochloric acid, triethanolamine, barbituric sodium/hydrochloric acid, glycine/sodium hydroxide, or amino donor/hydrochloric acid.
8. The process for the preparation of a chiral β -amino acid derivative according to any one of claims 1 to 7, wherein the alcoholic solvent is selected from one or more of methanol, ethanol, propanol.
9. The method for preparing a chiral β -amino acid derivative according to claim 8, wherein the first solvent is a mixed solvent of the alcohol solvent and water, and the mass fraction of the alcohol solvent is 20 to 40%.
10. Process for the preparation of chiral β -amino acid derivatives according to any of claims 1 to 4, characterized in that the amino donor is selected from C1～6One or more of alkylamine and inorganic ammonium salt;
preferably, said C1～6The alkylamine is isopropylamine, butylamine or pentylamine;
preferably, the inorganic ammonium salt is ammonium formate, ammonium chloride or ammonium sulfate.
11. The method for producing a chiral β -amino acid derivative according to any one of claims 1 to 6, wherein an amino mediator selected from one or more of pyridoxal phosphate, pyridoxamine phosphate, and pyridoxine phosphate is further included in the mixed raw material solution.
12. The process for the preparation of a chiral β -amino acid derivative according to any one of claims 1 to 6, wherein the biological enzyme catalyst is a transaminase; the transaminase is selected from one or more of aspartate transaminase, aromatic transaminase, omega-transaminase, branched-chain transaminase, and D-transaminase.
13. The method for preparing the chiral beta-amino acid derivative according to any one of claims 1 to 6, wherein in the enzymatic reaction process, the reaction temperature is 20-60 ℃, the reaction pH value is 7.0-11, and the reaction time is 18-36 h;
preferably, the reaction temperature is 35-55 ℃, and the reaction pH value is 8.0-10.5.
14. The process for the preparation of chiral β -amino acid derivatives of claim 1, wherein R is said aryl-C1～2Alkyl, wherein said aryl-C1～2The phenyl group of the alkyl group is unsubstituted or substituted with 1 to 3 substituents selected from the group consisting of fluorine, trifluoromethyl and trifluoromethoxy; z is NR2R3；NR2R3Has the structural formula
R4Is hydrogen or C unsubstituted or substituted by 1 to 5 fluorine atoms1～4An alkyl group.
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| CN102405281A (en) * | 2009-02-26 | 2012-04-04 | 科德克希思公司 | Transaminase biocatalysts |
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