CN116987138A - Preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester - Google Patents
Preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 26
- 238000000034 method Methods 0.000 claims abstract description 26
- DGABKXLVXPYZII-SIBKNCMHSA-N hyodeoxycholic acid Chemical compound C([C@H]1[C@@H](O)C2)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)CC1 DGABKXLVXPYZII-SIBKNCMHSA-N 0.000 claims abstract description 10
- DGABKXLVXPYZII-UHFFFAOYSA-N Hyodeoxycholic acid Natural products C1C(O)C2CC(O)CCC2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)CC2 DGABKXLVXPYZII-UHFFFAOYSA-N 0.000 claims abstract description 9
- 239000002994 raw material Substances 0.000 claims abstract description 8
- 238000005886 esterification reaction Methods 0.000 claims abstract description 6
- 230000032050 esterification Effects 0.000 claims abstract description 3
- 238000006243 chemical reaction Methods 0.000 claims description 39
- 239000004367 Lipase Substances 0.000 claims description 33
- 102000004882 Lipase Human genes 0.000 claims description 33
- 108090001060 Lipase Proteins 0.000 claims description 33
- 235000019421 lipase Nutrition 0.000 claims description 33
- 150000001875 compounds Chemical class 0.000 claims description 31
- 229940126062 Compound A Drugs 0.000 claims description 21
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 21
- NLDMNSXOCDLTTB-UHFFFAOYSA-N Heterophylliin A Natural products O1C2COC(=O)C3=CC(O)=C(O)C(O)=C3C3=C(O)C(O)=C(O)C=C3C(=O)OC2C(OC(=O)C=2C=C(O)C(O)=C(O)C=2)C(O)C1OC(=O)C1=CC(O)=C(O)C(O)=C1 NLDMNSXOCDLTTB-UHFFFAOYSA-N 0.000 claims description 21
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 18
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 18
- ZAFNJMIOTHYJRJ-UHFFFAOYSA-N Diisopropyl ether Chemical compound CC(C)OC(C)C ZAFNJMIOTHYJRJ-UHFFFAOYSA-N 0.000 claims description 17
- 238000001914 filtration Methods 0.000 claims description 16
- 239000007800 oxidant agent Substances 0.000 claims description 15
- 238000001035 drying Methods 0.000 claims description 14
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical class [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 12
- 125000000218 acetic acid group Chemical group C(C)(=O)* 0.000 claims description 11
- 239000012074 organic phase Substances 0.000 claims description 10
- 230000001590 oxidative effect Effects 0.000 claims description 10
- HETCEOQFVDFGSY-UHFFFAOYSA-N Isopropenyl acetate Chemical group CC(=C)OC(C)=O HETCEOQFVDFGSY-UHFFFAOYSA-N 0.000 claims description 9
- 239000005708 Sodium hypochlorite Substances 0.000 claims description 9
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical group [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 claims description 9
- PCLIMKBDDGJMGD-UHFFFAOYSA-N N-bromosuccinimide Chemical compound BrN1C(=O)CCC1=O PCLIMKBDDGJMGD-UHFFFAOYSA-N 0.000 claims description 8
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 8
- QYTDEUPAUMOIOP-UHFFFAOYSA-N TEMPO Chemical class CC1(C)CCCC(C)(C)N1[O] QYTDEUPAUMOIOP-UHFFFAOYSA-N 0.000 claims description 8
- 229940099352 cholate Drugs 0.000 claims description 8
- 239000000706 filtrate Substances 0.000 claims description 8
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 8
- 239000003960 organic solvent Substances 0.000 claims description 8
- KJIFKLIQANRMOU-UHFFFAOYSA-N oxidanium;4-methylbenzenesulfonate Chemical compound O.CC1=CC=C(S(O)(=O)=O)C=C1 KJIFKLIQANRMOU-UHFFFAOYSA-N 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 238000005406 washing Methods 0.000 claims description 7
- 239000002904 solvent Substances 0.000 claims description 6
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 4
- XTXRWKRVRITETP-UHFFFAOYSA-N Vinyl acetate Chemical compound CC(=O)OC=C XTXRWKRVRITETP-UHFFFAOYSA-N 0.000 claims description 4
- 239000003377 acid catalyst Substances 0.000 claims description 4
- 239000012043 crude product Substances 0.000 claims description 4
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 3
- 239000003054 catalyst Substances 0.000 claims description 3
- HOSGXJWQVBHGLT-UHFFFAOYSA-N 6-hydroxy-3,4-dihydro-1h-quinolin-2-one Chemical group N1C(=O)CCC2=CC(O)=CC=C21 HOSGXJWQVBHGLT-UHFFFAOYSA-N 0.000 claims description 2
- DWAQJAXMDSEUJJ-UHFFFAOYSA-M Sodium bisulfite Chemical compound [Na+].OS([O-])=O DWAQJAXMDSEUJJ-UHFFFAOYSA-M 0.000 claims description 2
- 230000000397 acetylating effect Effects 0.000 claims description 2
- 239000012295 chemical reaction liquid Substances 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims description 2
- 235000010267 sodium hydrogen sulphite Nutrition 0.000 claims description 2
- 239000004289 sodium hydrogen sulphite Substances 0.000 claims description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 2
- 238000003786 synthesis reaction Methods 0.000 abstract description 10
- 230000015572 biosynthetic process Effects 0.000 abstract description 9
- 102000004190 Enzymes Human genes 0.000 abstract description 2
- 108090000790 Enzymes Proteins 0.000 abstract description 2
- 239000003814 drug Substances 0.000 abstract description 2
- 230000021736 acetylation Effects 0.000 abstract 1
- 238000006640 acetylation reaction Methods 0.000 abstract 1
- 125000000217 alkyl group Chemical group 0.000 abstract 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 abstract 1
- 238000005832 oxidative carbonylation reaction Methods 0.000 abstract 1
- QWXYZCJEXYQNEI-OSZHWHEXSA-N intermediate I Chemical compound COC(=O)[C@@]1(C=O)[C@H]2CC=[N+](C\C2=C\C)CCc2c1[nH]c1ccccc21 QWXYZCJEXYQNEI-OSZHWHEXSA-N 0.000 description 29
- 239000000243 solution Substances 0.000 description 14
- 239000007787 solid Substances 0.000 description 13
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 8
- 239000002244 precipitate Substances 0.000 description 8
- 238000004321 preservation Methods 0.000 description 5
- 238000012512 characterization method Methods 0.000 description 4
- 230000035484 reaction time Effects 0.000 description 4
- 239000011734 sodium Substances 0.000 description 4
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 4
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- BHQCQFFYRZLCQQ-UHFFFAOYSA-N (3alpha,5alpha,7alpha,12alpha)-3,7,12-trihydroxy-cholan-24-oic acid Natural products OC1CC2CC(O)CCC2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)C(O)C2 BHQCQFFYRZLCQQ-UHFFFAOYSA-N 0.000 description 2
- RUDATBOHQWOJDD-UHFFFAOYSA-N (3beta,5beta,7alpha)-3,7-Dihydroxycholan-24-oic acid Natural products OC1CC2CC(O)CCC2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)CC2 RUDATBOHQWOJDD-UHFFFAOYSA-N 0.000 description 2
- 239000004380 Cholic acid Substances 0.000 description 2
- 238000007239 Wittig reaction Methods 0.000 description 2
- BHQCQFFYRZLCQQ-OELDTZBJSA-N cholic acid Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)[C@@H](O)C1 BHQCQFFYRZLCQQ-OELDTZBJSA-N 0.000 description 2
- 229960002471 cholic acid Drugs 0.000 description 2
- 235000019416 cholic acid Nutrition 0.000 description 2
- KXGVEGMKQFWNSR-UHFFFAOYSA-N deoxycholic acid Natural products C1CC2CC(O)CCC2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)C(O)C2 KXGVEGMKQFWNSR-UHFFFAOYSA-N 0.000 description 2
- 230000008030 elimination Effects 0.000 description 2
- 238000003379 elimination reaction Methods 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 229960001601 obeticholic acid Drugs 0.000 description 2
- ZXERDUOLZKYMJM-ZWECCWDJSA-N obeticholic acid Chemical compound C([C@@]12C)C[C@@H](O)C[C@H]1[C@@H](CC)[C@@H](O)[C@@H]1[C@@H]2CC[C@]2(C)[C@@H]([C@H](C)CCC(O)=O)CC[C@H]21 ZXERDUOLZKYMJM-ZWECCWDJSA-N 0.000 description 2
- 238000001953 recrystallisation Methods 0.000 description 2
- GEHJYWRUCIMESM-UHFFFAOYSA-L sodium sulfite Chemical compound [Na+].[Na+].[O-]S([O-])=O GEHJYWRUCIMESM-UHFFFAOYSA-L 0.000 description 2
- 150000003431 steroids Chemical class 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000000967 suction filtration Methods 0.000 description 2
- SMEROWZSTRWXGI-UHFFFAOYSA-N Lithocholsaeure Natural products C1CC2CC(O)CCC2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)CC2 SMEROWZSTRWXGI-UHFFFAOYSA-N 0.000 description 1
- 238000006882 Shapiro reaction Methods 0.000 description 1
- NBVHDOZEOGAKLK-UHFFFAOYSA-N [N]=O.CC1C(N(CCC1)C)(C)C Chemical compound [N]=O.CC1C(N(CCC1)C)(C)C NBVHDOZEOGAKLK-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229960003328 benzoyl peroxide Drugs 0.000 description 1
- 238000005893 bromination reaction Methods 0.000 description 1
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229960001091 chenodeoxycholic acid Drugs 0.000 description 1
- RUDATBOHQWOJDD-BSWAIDMHSA-N chenodeoxycholic acid Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)CC1 RUDATBOHQWOJDD-BSWAIDMHSA-N 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- KPUWHANPEXNPJT-UHFFFAOYSA-N disiloxane Chemical compound [SiH3]O[SiH3] KPUWHANPEXNPJT-UHFFFAOYSA-N 0.000 description 1
- 150000002085 enols Chemical class 0.000 description 1
- 238000006735 epoxidation reaction Methods 0.000 description 1
- 150000007857 hydrazones Chemical class 0.000 description 1
- 238000009413 insulation Methods 0.000 description 1
- SMEROWZSTRWXGI-HVATVPOCSA-N lithocholic acid Chemical compound C([C@H]1CC2)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)CC1 SMEROWZSTRWXGI-HVATVPOCSA-N 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
- 238000006722 reduction reaction Methods 0.000 description 1
- 238000007142 ring opening reaction Methods 0.000 description 1
- 235000010265 sodium sulphite Nutrition 0.000 description 1
- 238000009987 spinning Methods 0.000 description 1
- 238000001308 synthesis method Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- RUDATBOHQWOJDD-UZVSRGJWSA-N ursodeoxycholic acid Chemical compound C([C@H]1C[C@@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)CC1 RUDATBOHQWOJDD-UZVSRGJWSA-N 0.000 description 1
- 229960001661 ursodiol Drugs 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07J—STEROIDS
- C07J9/00—Normal steroids containing carbon, hydrogen, halogen or oxygen substituted in position 17 beta by a chain of more than two carbon atoms, e.g. cholane, cholestane, coprostane
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses a preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester, relating to the technical field of medicine synthesis; the invention uses hyodeoxycholic acid as raw material, and is obtained by alkyl esterification, selective acetylation of 3 rd position by biological enzyme method, and oxidative carbonylation of 6 th hydroxyl. The 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester prepared by the synthetic route has wide raw material sources and high product yield and purity, is suitable for mass preparation, and can be subjected to subsequent industrialized development.
Description
Technical Field
The invention relates to the technical field of medicine synthesis, in particular to a preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester.
Background
3 alpha-acetoxy-6-keto-5 beta-cholanic acid methyl ester (hereinafter referred to as intermediate I) is an important intermediate, and various high-value cholic acid substances can be synthesized by using the intermediate in various reported documents and patents.
In patent publication nos. CN108264532a and CN108264533a, zhang Jie and the like use intermediate I to perform the synthesis of obeticholic acid by Wittig reaction (Wittig reaction). The specific reaction process is as follows:
in patent publication No. CN 10823913a, gao Heyong and the like attack carbonyl group by an ethyl-formatted reagent using intermediate I, and then undergo steps such as elimination and hydrolysis to obtain obeticholic acid. The specific reaction process is as follows:
furthermore, ursodeoxycholic acid (Dou Qian et al, chemical report, 2018, 81 (08): 720-726.) was prepared from intermediate I by both Shapiro reaction and enol silyl ether oxidation. The specific reaction process is as follows:
in patent publication No. CN 109134576a, han Ying et al synthesized lithocholic acid by hydrazone reduction using intermediate I. The specific reaction process is as follows:
in the applicant's earlier patent CN 109369765a, also intermediate I was used to obtain β -allo-murine cholic acid by steps such as α -bromination, hydrolysis, etc. The specific reaction process is as follows:
in addition, chenodeoxycholic acid can be synthesized by steps of reduction, elimination, epoxidation ring opening, hydrolysis, and the like using intermediate I (Steroids, 2020, 157:108594). The specific reaction process is as follows:
in summary, 3 alpha-acetoxyl group-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) has wide application prospect and industrial value. The existing preparation method of the intermediate I is mainly obtained by a chemical synthesis method, but the preparation process is complex, column chromatography purification is needed, the yield is low, and the intermediate I with high purity is not obtained by a better method and cannot be produced in a large scale. Thus, it is highly desirable to find a simple, efficient, economical and inexpensive method for mass-synthesizing the intermediate I to solve the source problem.
In view of this, the present invention has been made.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide a preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester, which uses hyodeoxycholic acid (3 alpha, 6 alpha-dihydroxyl-5 beta-cholanic acid) as a raw material, uses biological enzyme as a catalyst to selectively acetylate 3-hydroxyl of a compound A, and solves the problem of poor selectivity in the prior synthesis method; the synthesis route provided by the invention has the advantages of mild condition, good selectivity, high yield and less pollution emission, is suitable for mass preparation, and solves the technical problems of complex synthesis process, poor selectivity and lower yield of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) in the prior art.
In order to solve the technical problems, the invention provides the following technical scheme:
the invention provides a preparation method of 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester, which comprises the following steps:
step a: the method comprises the steps of (1) performing side chain esterification on hyodeoxycholic acid serving as a raw material to obtain a compound A;
step b: dissolving a compound A, lipase and an acetyl donor in a first organic solvent, and selectively acetylating 3-hydroxyl in the compound A by taking lipase as a catalyst to obtain a compound B; wherein the lipase is novelin lipase 435, novelin lipase 40086, lipase CAL B or lipase CAL B-2;
step c: dissolving a compound B in a second organic solvent, adding an oxidant, and oxidizing the 6-hydroxyl in the compound B to obtain 3 alpha-acetoxy-6-ketone-5 beta-cholanic acid methyl ester;
the reaction route of the preparation method is as follows:
the 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester intermediate I is prepared by the synthetic route, and has the advantages of wide raw material sources, sufficient supply, simple and convenient reaction process and good selectivity; in addition, the 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) obtained through the steps has high yield which can reach 94.1 percent, is suitable for mass preparation, and can provide support for subsequent related research and industrialization.
The hyodeoxycholic acid is 3 alpha, 6 alpha-dihydroxyl-5 beta-cholanic acid.
Further, the lipase is preferably novelin lipase 435.
Further, step a comprises: under the action of an acid catalyst, performing esterification reaction on hyodeoxycholic acid and alcohol at 60-70 ℃ to obtain a compound A.
In step a, it is preferable to carry out the esterification reaction using hyodeoxycholic acid and methanol under the above conditions.
Preferably, the acid catalyst is one or more of concentrated hydrochloric acid, concentrated sulfuric acid, and p-toluenesulfonic acid monohydrate, and more preferably p-toluenesulfonic acid monohydrate.
Further, in the step b, the first organic solvent is one or more of isopropyl ether, ethyl acetate and n-hexane; isopropyl ether is preferred.
Further, in the step b, the reaction temperature is 35-45 ℃; step b may adjust the reaction time according to the reaction progress, preferably the reaction time is 6 to 18 hours, to complete the reaction.
The preferable scheme of the step b is as follows: the compound A and isopropyl ether are mixed in advance, after fully and uniformly mixing, the Norwestine lipase is added, after fully and uniformly mixing again, the mixture is heated, and then the acetyl donor is added for thermal insulation reaction. Wherein the reaction temperature is 35-45 ℃.
Further, in step b, the acetyl donor is isopropenyl acetate, vinyl acetate or ethyl acetate; preferably isopropenyl acetate.
Further, in the step b, the mass ratio of the compound A to the lipase is 1:0.01-0.5; the molar ratio of the compound A to the acetyl donor is 1:0.7-1.5.
Preferably, the mass ratio of the compound A to the lipase is 1:0.01-0.5, i.e. in various embodiments of the invention, the mass ratio of compound a to novelin lipase may be 1:0.05, 1:0.1, 1:0.3, 1:0.4, 1:0.5, etc. The preferred mass ratio is 1:0.2.
preferably, the molar ratio of compound a to acetyl donor is 1:0.7 to 1.5, i.e. in various embodiments of the invention, the molar ratio of compound a to acetyl donor may be 1:0.7, 1:0.8, 1:1.0, 1:1.2, 1:1.5, etc. The preferred molar ratio is 1:1.2.
further, the step b further comprises the steps of purifying: after the reaction is finished, filtering the reaction liquid, collecting filtrate, concentrating to obtain a crude product of the sample B, dissolving the crude product with dichloromethane, washing with saturated sodium bicarbonate solution and saturated saline water in sequence, collecting an organic phase, drying, and removing the solvent to obtain the compound B.
Further, the reaction time in step c can be adjusted according to the reaction progress, preferably the reaction time is 2-3h, so that the reaction is complete.
Further, in step c, the reaction temperature is 0 to 20 ℃.
Further, in the step c, the second organic solvent is one or more of dichloromethane and ethyl acetate.
Further, in step c, the step of,the oxidant is sodium hypochlorite, tetramethyl piperidine nitrogen oxide, mnO 2 、CrO 3 、H 2 O 2 One or more of N-bromosuccinimide;
preferably one or more of sodium hypochlorite and tetramethyl piperidine nitroxide;
more preferably, the oxidizing agent is a combination of sodium hypochlorite/tetramethylpiperidine nitroxide, and the molar ratio of sodium hypochlorite to tetramethylpiperidine nitroxide in the oxidizing agent is 1:0.001-0.1; the molar ratio of the compound B to the oxidant is 1:1.2-3.0
That is, in various embodiments of the invention, the molar ratio of sodium hypochlorite to tetramethylpiperidine nitroxide may be 1:0.001, 1:0.003, 1:0.005, 1:0.007, 1:0.009, 1:0.01, 1:0.03, 1:0.05, 1:0.08, 1:0.1, etc.;
the molar ratio of compound B to oxidant may be 1:1.2, 1:1.5, 1:1.8, 1:2.0, 1:2.5, 1:2.7, 1:2.9, 1:3.0, etc.
Further, the step c further comprises the steps of purifying: after the reaction, the reaction was quenched with 10% sodium bisulphite solution, the organic phase was washed with saturated sodium bicarbonate solution and then with saturated saline solution, dried, the solvent was removed, and methanol was recrystallized to obtain methyl 3 a-acetoxy-6-keto-5 β -cholate.
The reaction conditions of the steps are relatively mild, the process is simple, the product is obtained through 3 steps of reaction, the control is easy, and the target compound can be obtained without complex equipment.
Compared with the prior art, the invention has the beneficial effects that:
(1) The invention develops a method for synthesizing 3 alpha-acetoxyl-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) by taking hyodeoxycholic acid as a raw material, which has wide raw material sources and low cost.
(2) In the synthetic route, the method has the advantages of mild conditions, easiness in control, good selectivity and high yield, is suitable for mass preparation, and solves the technical problems of complex synthesis process, low yield and poor selectivity of the intermediate I in the prior art.
(3) According to the invention, the reaction conditions of each step in the synthetic route are further optimized, and the yield of the intermediate I is improved.
Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention.
Drawings
The accompanying drawings, which are included to provide a further understanding of the invention and constitute a part of this specification, are incorporated in and constitute a part of this specification and do not limit the invention in any way, and in which:
FIG. 1 is a synthetic route for 3 alpha-acetoxy-6-keto-5 beta-cholanic acid methyl ester provided by the example of the present invention.
Detailed Description
For a more complete understanding of the present invention, reference should be made to the following descriptions and illustrations of the present invention in conjunction with the accompanying drawings and the detailed description thereof; it should be noted that the positional or positional relationship indicated by the terms "upper", "lower", "front", "rear", "inner", "outer", etc. are not given to the positional or positional relationship shown in the drawings, and are merely for convenience of description of the present usage confidence and simplification of the description, and are not indicative or implying that the apparatus or element in question must have a specific orientation, be constructed and operated in a specific orientation, and therefore should not be construed as limiting the present invention. Furthermore, the terms "first," "second," and the like, herein, are used for distinguishing between different elements and not necessarily for describing a sequential or chronological order, and not for limiting that "first" and "second" are of different types.
The following description of the embodiments of the present invention will be made clearly and fully with reference to the accompanying drawings, in which it is evident that the embodiments described are only some, but not all embodiments of the invention; all other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to fall within the scope of the invention.
Example 1
The preparation method of the 3 alpha-acetoxy-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) in the embodiment refers to a synthetic route chart shown in fig. 1, and comprises the following steps:
(a) Synthesis of 3 alpha, 6 alpha-dihydroxy-5 beta-cholanic acid methyl ester (Compound A)
Reference has been made to the preparation of Steroids,2020,157:108594, in 98.3% yield.
The structural characterization data for compound a are as follows:
HRMS:Calcd for C 25 H 42 O 4 [M+Na] + 429.2981，Found 429.2985。
(b) Synthesis of 3 alpha-acetoxy-6 alpha-hydroxy-5 beta-cholanic acid methyl ester (Compound B) Compound A (5.00 g,12.3 mmol), isopropyl ether 50mL and NoveXin lipase 435 (1.00 g) were stirred well, isopropenyl acetate (4.0 mL,36.6 mmol) was added at 40℃and the reaction was stirred at constant temperature for 8h. After the reaction, insoluble matter was removed by filtration, and the precipitate was washed with 20mL of isopropyl ether; the filtrate was dried by spinning, dissolved in 40mL of methylene chloride, washed successively with a saturated sodium hydrogencarbonate solution (20 mL. Times.3) and a saturated brine (20 mL. Times.3), and the organic phase was dried over anhydrous magnesium sulfate to remove water, filtered and concentrated to give 5.42g of a white solid as Compound B in 98.3% yield.
The structural characterization data for compound B are as follows:
1 H NMR(400MHz,CDCl 3 )δ4.68(m,1H),4.03(m,1H),3.63(s,3H),2.32(m,1H),2.18(m,1H),1.99(s,3H),0.91–0.85(m,6H),0.61(s,3H)。
13 C NMR(101MHz,CDCl 3 )δ174.7,170.5,74.2,67.5,56.1,55.9,51.4,48.3,42.8,39.9,39.8,35.9,35.3,35.2,34.7,34.7,31.0,30.9,28.1,26.5,25.3,24.1,23.4,21.4,20.7,18.2,12.0。
HRMS:Calcd for C 27 H 44 O 5 [M+Na] + 471.3086，Found 471.3092。
(c) Synthesis of 3 alpha-acetoxy-6-keto-5 beta-cholanic acid methyl ester (intermediate I)
Compound B (5.00 g,11.15 mmol) and 50mL of methylene chloride were added to the reaction vessel, dissolved by stirring, tetramethylpiperidine nitroxide (15.0 mg,0.1 mmol) was added at about 5℃and stirred uniformly, dropwise addition of sodium hypochlorite solution was started, and after completion of the dropwise addition, a total of 12.0mL (about 20.3 mmol) was added, and the reaction was continued for 2 hours. After the reaction, 30mL of 10% sodium sulfite solution was slowly added to quench the reaction, followed by washing with saturated sodium bicarbonate solution (25 mL. Times.2) and saturated brine (25 mL. Times.3), drying over anhydrous magnesium sulfate to remove water, removing solids by suction filtration, and concentrating to remove solvent to obtain pale yellow solid. 20mL of methanol was added for recrystallization, and filtration and drying gave 4.83g of a white powdery solid, methyl 3 a-acetoxy-6-keto-5 β -cholate (intermediate I) in 97.1% yield.
Structural characterization data for 3 α -acetoxy-6-keto-5 β -cholanic acid methyl ester (intermediate I) are as follows:
1 H NMR(400MHz,CDCl 3 )δ4.62(m,1H),3.60(s,3H),1.96(s,3H),0.86(d,J＝6.5Hz,3H),0.78(s,3H),0.59(s,3H)。
13 C NMR(101MHz,CDCl 3 )δ212.7,174.6,170.2,72.4,59.1,56.8,55.8,51.5,43.1,42.8,39.9,39.6,37.9,37.1,35.2,34.1,31.0,31.0,30.9,28.0,26.2,23.9,23.1,21.3,20.8,18.2,11.9。
HRMS:Calcd for C 27 H 42 O 5 [M+Na] + 469.2930，Found 469.2939。
according to the results, the preparation method provided by the invention has the advantages of mild conditions of each step, easiness in control, good selectivity and high yield.
Example 2
This example refers to the preparation method of example 1, differing only in: in step (b), the acetyl donor is vinyl acetate.
The method comprises the following specific steps: compound A (1.00 g,2.5 mmol), isopropyl ether 10mL and Norwechat lipase 435 (0.20 g) were stirred well, vinyl acetate (0.8 mL,8.6 mmol) was added at 40℃and stirred for 8h with heat preservation, the precipitate was removed by filtration and washed with isopropyl ether; spin-drying the filtrate with CH 2 Cl 2 Dissolving, washing sequentially with saturated sodium bicarbonate solution (10 mL×3) and saturated saline (10 mL×3), drying the organic phase with anhydrous magnesium sulfate for removing water, filtering, concentrating to obtain white solid 0.53g as compoundB, yield 48.0%.
From the above results, it is found that the choice of acetyl donor in the present invention affects the yield of compound B and thus the final yield of 3 α -acetoxy-6-keto-5 β -cholanic acid methyl ester (intermediate I).
Example 3
This example refers to the preparation method of example 1, differing only in: in step (b), the lipase is novelin lipase 40086.
The method comprises the following specific steps: compound A (1.00 g,2.5 mmol), isopropyl ether 10mL and Norwechat lipase 40086 (0.20 g) were stirred well, isopropenyl acetate (0.8 mL,7.3 mmol) was added at 40℃and stirred for 8h with heat preservation, the precipitate was removed by filtration, and the precipitate was washed with isopropyl ether; spin-drying the filtrate with CH 2 Cl 2 Dissolving, washing with saturated sodium bicarbonate solution (10 mL. Times.3) and saturated saline (10 mL. Times.3) in sequence, drying the organic phase with anhydrous magnesium sulfate to remove water, filtering and concentrating to obtain white solid 0.55g, which is compound B, with a yield of 50.0%.
From the above results, it is found that the selection of lipase in the present invention affects the yield of compound B, and thus the final yield of 3 α -acetoxy-6-keto-5 β -cholanic acid methyl ester (intermediate I).
Example 4
This example refers to the preparation method of example 1, differing only in: in step (B), the lipase is lipase CAL B.
The method comprises the following specific steps: compound A (1.00 g,2.5 mmol), isopropyl ether 10mL and NoveXin lipase CAL B (0.20 g) were stirred well, isopropenyl acetate (0.8 mL,7.3 mmol) was added at 40℃and stirred for 8h with heat preservation, the precipitate was removed by filtration, and the precipitate was washed with isopropyl ether; spin-drying the filtrate with CH 2 Cl 2 Dissolving, washing with saturated sodium bicarbonate solution (10 mL. Times.3) and saturated saline (10 mL. Times.3) in sequence, drying the organic phase with anhydrous magnesium sulfate to remove water, filtering and concentrating to obtain 0.39g of white solid, which is compound B, with a yield of 35.3%.
From the above results, it is found that the selection of lipase in the present invention affects the yield of compound B, and thus the final yield of 3 α -acetoxy-6-keto-5 β -cholanic acid methyl ester (intermediate I).
Example 5
This example refers to the preparation method of example 1, differing only in: in step (b), the reaction temperature was 35 ℃.
The method comprises the following specific steps: compound A (1.00 g,2.5 mmol), isopropyl ether 10mL and NoveXin lipase 435 (0.20 g) were stirred well, isopropenyl acetate (0.8 mL,7.3 mmol) was added at 35℃and stirred for 8h with heat preservation, the precipitate was removed by filtration and washed with isopropyl ether; spin-drying the filtrate with CH 2 Cl 2 Dissolving, washing with saturated sodium bicarbonate solution (10 mL. Times.3) and saturated saline (10 mL. Times.3) in sequence, drying the organic phase with anhydrous magnesium sulfate to remove water, filtering and concentrating to obtain 1.04g of white solid, which is compound B, with a yield of 94.3%.
The structural characterization data for compound B are as follows:
HRMS:Calcd for C 27 H 44 O 5 [M+Na] + 471.3086，Found 471.3094。
example 6
This example refers to the preparation method of example 1, differing only in: in step (b), the reaction temperature was 45 ℃.
The method comprises the following specific steps: compound A (1.00 g,2.5 mmol), isopropyl ether 10mL and NoveXin lipase 435 (0.20 g) were stirred well, isopropenyl acetate (0.8 mL,7.31 mmol) was added at 45℃and stirred for 8h with heat preservation, the precipitate was removed by filtration and washed with isopropyl ether; spin-drying the filtrate with CH 2 Cl 2 The mixture was dissolved, washed with a saturated sodium hydrogencarbonate solution (10 mL. Times.3) and a saturated brine (10 mL. Times.3) in this order, and the organic phase was dried over anhydrous magnesium sulfate and dehydrated, and after filtration, it was concentrated to give 1.02g of a white solid as Compound B in 92.4% yield.
According to the test results of example 1, example 5 and example 6, the reaction temperature of step B is between 35 and 45 ℃, and the compound B has higher yield, so that the high yield of 3 alpha-acetoxy-6-ketone-5 beta-cholanic acid methyl ester (intermediate I) can be ensured.
Example 7
This example refers to the preparation method of example 1, differing only in: in step (c), the oxidizing agent is N-bromosuccinimide.
The method comprises the following specific steps: adding compound B (1.00 g,2.23 mmol) and dichloromethane (15 mL) into a reaction vessel, stirring for dissolving, slowly adding MnO in batches at about 15 DEG C 2 The solid (0.34 g,3.91 mmol) was stirred for 4h. After the completion of the reaction, the solid was removed by filtration, and then washed with a saturated sodium hydrogencarbonate solution (10 mL. Times.2) and a saturated brine (10 mL. Times.3), dried over anhydrous magnesium sulfate, removed by suction filtration, and the solvent was concentrated to give a pale yellow solid. 5mL of methanol was added for recrystallization, and filtration and drying gave 0.76g of a white powdery solid, methyl 3 a-acetoxy-6-keto-5 β -cholate (intermediate I) in a yield of 76.0%.
As a result, the selection of the oxidizing agent in the present invention affects the yield of methyl 3 a-acetoxy-6-keto-5 β -cholate (intermediate I), and example 7 uses N-bromosuccinimide as the oxidizing agent, and the yield of methyl 3 a-acetoxy-6-keto-5 β -cholate (intermediate I) is lower than that of example 1, but the yield is also higher, indicating that the oxidizing agent used in the present invention can increase the yield of methyl 3 a-acetoxy-6-keto-5 β -cholate (intermediate I).
The foregoing has described in detail the technical solutions provided by the embodiments of the present invention, and specific examples have been applied to illustrate the principles and implementations of the embodiments of the present invention, where the above description of the embodiments is only suitable for helping to understand the principles of the embodiments of the present invention; meanwhile, as for those skilled in the art, according to the embodiments of the present invention, there are variations in the specific embodiments and the application scope, and the present description should not be construed as limiting the present invention.
Claims (10)
- A process for the preparation of methyl 3 α -acetoxy-6-keto-5 β -cholate, the process comprising the steps of:step a: the method comprises the steps of (1) performing side chain esterification on hyodeoxycholic acid serving as a raw material to obtain a compound A;step b: dissolving a compound A, lipase and an acetyl donor in a first organic solvent, and selectively acetylating 3-hydroxyl in the compound A by taking lipase as a catalyst to obtain a compound B; wherein the lipase is novelin lipase 435, novelin lipase 40086, lipase CAL B or lipase CAL B-2;step c: dissolving a compound B in a second organic solvent, adding an oxidant, and oxidizing the 6-hydroxyl in the compound B to obtain 3 alpha-acetoxy-6-ketone-5 beta-cholanic acid methyl ester;the reaction route of the preparation method is as follows:
- 2. the method of claim 1, wherein step a comprises: under the action of an acid catalyst, performing esterification reaction on hyodeoxycholic acid and alcohol at 60-70 ℃ to obtain a compound A;preferably, the acid catalyst is one or more of concentrated hydrochloric acid, concentrated sulfuric acid, and p-toluenesulfonic acid monohydrate, more preferably p-toluenesulfonic acid monohydrate;the alcohol is methanol.
- 3. The process according to claim 1, wherein in step b, the reaction temperature is from 35 to 45 ℃.
- 4. The method of claim 1, wherein in step b, the acetyl donor is isopropenyl acetate, vinyl acetate or ethyl acetate; preferably isopropenyl acetate.
- 5. The preparation method according to claim 1, wherein in the step b, the mass ratio of the compound a to the lipase is 1:0.01-0.5; the molar ratio of the compound A to the acetyl donor is 1:0.7-1.5.
- 6. The preparation method according to claim 1, wherein in the step b, the first organic solvent is one or more of isopropyl ether, ethyl acetate and n-hexane; isopropyl ether is preferred.
- 7. The method according to any one of claims 1 to 6, wherein the purification in step b further comprises: after the reaction is finished, filtering the reaction liquid, collecting filtrate, concentrating to obtain a crude product of the sample B, dissolving the crude product with dichloromethane, washing with saturated sodium bicarbonate solution and saturated saline water in sequence, collecting an organic phase, drying, and removing the solvent to obtain the compound B.
- 8. The method according to claim 1, wherein in the step c, the oxidizing agent is sodium hypochlorite, tetramethylpiperidine nitroxide, mnO 2 、CrO 3 、H 2 O 2 One or more of N-bromosuccinimide;preferably, the oxidant is a combination of sodium hypochlorite/tetramethylpiperidine nitroxide, and the molar ratio of sodium hypochlorite to tetramethylpiperidine nitroxide in the oxidant is 1:0.001-0.1; the molar ratio of the compound B to the oxidant is 1:1.2-3.0.
- 9. The process according to claim 1, wherein in step c, the reaction temperature is 0-20 ℃; the second organic solvent is one or more of dichloromethane and ethyl acetate.
- 10. The method according to any one of claims 1, 8 to 9, wherein step c further comprises purifying: after the reaction, the reaction was quenched with 10% sodium bisulphite solution, the organic phase was washed with saturated sodium bicarbonate solution and then with saturated saline solution, dried, the solvent was removed, and methanol was recrystallized to obtain methyl 3 a-acetoxy-6-keto-5 β -cholate.
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