CN108384817B

CN108384817B - Synthesis method of (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid

Info

Publication number: CN108384817B
Application number: CN201810099344.0A
Authority: CN
Inventors: 徐红岩; 陆广; 汪海明
Original assignee: Kanghua Shanghai New Drug R & D Co ltd
Current assignee: Kanghua Shanghai New Drug R & D Co ltd
Priority date: 2018-02-01
Filing date: 2018-02-01
Publication date: 2021-07-06
Anticipated expiration: 2038-02-01
Also published as: CN108384817A

Abstract

The invention relates to a synthesis method of (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid. The invention solves the problems that the chiral reagent is expensive and most synthetic routes reported in literatures are suitable for gram-grade production and cannot be used for kilogram-grade production, and the like, and the cost is high due to the synthesis by a chiral reagent method. The synthesis method comprises the following steps: (1) condensing L-alanine ethyl ester hydrochloride and benzyl carbonyl. (2) The alkyl group is introduced with 5-bromo-1-pentene. (3) And (3) a hydrochloric acid deprotection agent. (4) And (4) enzymolysis. (5) Fmoc protection. In the whole synthesis process, the intermediate and the target product do not need to be separated by a chromatographic column, the raw materials are cheap, and the purification is simple. The method is suitable for synthesizing (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid with low cost and high efficiency.

Description

Synthesis method of (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid

Technical Field

The invention relates to synthesis of (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid (CAS: 288617-73-2).

Background

The Stapled Peptide (Stapled Peptide) is an important active polypeptide structure modification mode, and the whole carbon skeleton of the Stapled Peptide forms alpha helical Peptide with stable side chain ring closing structure. The key building blocks for the synthesis of stapled peptides are generally unnatural chiral amino acids with alpha olefin side chains. (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid is widely applied to medical intermediates as an important raw material for synthesizing the staple peptide.

The asymmetric synthesis method of chiral amino acid is adopted at home and abroad. Wherein route 1: in the literature reported at present, the synthetic route of "(5S, 6R) -4-tert-butoxycarbonyl-5, 6-diphenyl morpholine-2-hydroxy amino acid" disclosed in chemical synthesis (2003.80.18), chiral auxiliary compound 6 is not produced in kilogram scale, and the intermediate material has large mass, which is extremely unfavorable for scale-up production.

Route 2: the chiral auxiliary material in the synthetic route of tetrahedron (56 (2000) (2577) (2582) 'large-scale asymmetric synthesis of enantiomer trans-cinnamyl glycine and-alpha-alanine)' is a coordination compound (compound 8) of metallic nickel, the synthetic difficulty is large, and the separation of free amino acid needs to use DOWEX ion exchange resin. Produces a large amount of waste liquid in industrial production and is not environment-friendly.

Synthetic scheme 1 reaction scheme:

；

synthetic scheme 2 reaction scheme:

。

disclosure of Invention

The invention aims to provide a synthetic method of (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid, which mainly solves the problem that the existing synthetic method can not realize kilogram-level production and adopts Lipase enzyme to split to obtain an intermediate with high chiral purity.

The technical scheme of the invention is as follows: step one, adding benzaldehyde into dichloromethane and triethylamine to obtain a compound 1, wherein the product is directly used for the next reaction without purification; secondly, adding 5-bromo-1-pentene into the compound 1 and potassium tert-butoxide in tetrahydrofuran for reaction, and stirring for reaction to obtain a compound 2; step three, reacting the compound 2 with a hydrochloric acid solution at room temperature to obtain a compound 3; fourthly, adding Lipase (Lipase) into the compound 3 and pure water, and carrying out enzymolysis to obtain a compound 4; and fifthly, reacting the compound 4 in acetone and sodium hydroxide, adding Fmoc-OSu, reacting at room temperature, and acidifying by hydrochloric acid to obtain a target compound 5.

The synthesis route is as follows:

in the reaction, the reaction temperature in the step 2 is 20-30 ℃, and the preferable reaction temperature is 25 ℃; the reaction time of the step 2 is 12 to 24 hours, and the preferable reaction time is 16 hours; step 4, the reaction temperature is 35-40 ℃, and the preferable reaction temperature is 37 ℃; step 4 has a pH of 7.0 to 9.0, preferably a pH of 7.8.

The invention has the beneficial effects that: the synthetic route adopted by the invention has the advantages that asymmetric synthetic chiral synthesis target compounds reported in documents are abandoned, racemate intermediates are synthesized by adopting a conventional method, and researches show that industrial production can be formed through enzymolysis, and finally the enzymolysis effect of the Lipase is optimal through screening of a large number of various enzymes. The route solves the problem of kilogram-level production of the product and provides an effective solution for the synthesis of similar compounds. The reagent is cheap, the reaction condition is simple, and the target product and the intermediate do not need to be purified by a chromatographic column.

Detailed Description

Example 1: the synthetic route is as follows:

step 1:

to a three-necked flask was added L-alanine ethyl ester hydrochloride (1.0 kg, 6.47 mol), dichloromethane (6L); triethylamine (1.63 kg, 16.1 mol) was added in an ice bath. The reaction solution was stirred at room temperature for 20 minutes. Benzaldehyde (0.82 kg, 7.76 mol) was added to the ice bath and stirred at room temperature for 24 hours. Saturated brine (1L x 3) was added, and the organic phase was dried over sodium sulfate and filtered. The filtrate was spin-dried to give a colorless liquid, compound 1 (1.33 kg, 6.47 mol, 100%) which was used directly in the next reaction;

step 2:

into a three-necked flask, Compound 1 (1.3 kg, 6.34 mol), tetrahydrofuran (6L) and potassium tert-butoxide (1.77 kg, 15.85 mol) were added; 5-bromo-1-pentene is added dropwise in an ice bath, and the temperature is naturally raised to 25 ℃ after the addition for reaction for 16 hours. Water (1.5L) was added and extracted with ethyl acetate (2L x 3), the organic phases combined and water (100 mL x 2) was used to spin dry to give compound 2 (1.77 kg, 6.5 mol, 100%) as a yellow liquid which was used directly in the next reaction;

and step 3:

compound 2 (1.77 kg, 6.5 mol) and methylene chloride (10L) were added to a three-necked flask followed by 6N HCl (10L), and stirred at room temperature for 5 hours. The reaction mixture was adjusted to pH =7-8 with saturated sodium bicarbonate, the layers were separated, the aqueous phase was extracted with dichloromethane (2L x 3), the dichloromethane phases were combined, washed with saturated brine (1.0L), dried over sodium sulfate and filtered. The filtrate was spun dry to give compound 3 (562 g, 3.03 mol, 46%) as a yellow liquid. Directly used for the next reaction;

and 4, step 4:

to a three-necked flask was added compound 3 (560 g, 3.02 mol), purified water (15.1L) was adjusted to pH 7.8 with 1N NaOH, and Lipase (50 g) (purchased from Sigma Aldrich trade Co., Ltd., Sigmaaldrich) was added, and the reaction mixture was stirred at 37 ℃ for 24 hours. The reaction was cooled to room temperature, filtered, the aqueous phase extracted with dichloromethane (2L x 3), the aqueous phase was concentrated to 2L and a solid precipitated. Filtration and drying afforded compound 4 as a white solid (161 g, 1.02 mol, 68%, ee: 99%).¹H NMR (400 MHz, D₂O) 1.21, (m, 1 H), 1.35, (m, 1 H), 1.35, (s,3 H) ,1.61, (m, 1 H), 1.65, (m, 1 H),2.0 (dd, 2 H), 4.95 (q, 2 H), 5.74 (m, 1 H) ppm；

And 5:

compound 4 (160 g, 1.02 mol), acetone (1L), water (1L) were added to a three-necked flask, followed by sodium bicarbonate (214.2 g, 2.55 mol) and Fmoc-OSu (343.7 g, 1.02 mol). The reaction solution was stirred at room temperature for 12 hours. Petroleum ether extraction (200 mL x 3); the aqueous phase was acidified to pH 3 with 1N hydrochloric acid, extracted with ethyl acetate (200 mL x 3), the organic phases combined, washed with saturated brine (150 mL), dried over sodium sulfate and filtered. The filtrate was spin-dried to give title compound 5 (304 g, 0.8 mol, 78%, Pu:99%, ee: 99%) as a white solid.¹H NMR (400 MHz, D₂O) 1.32, (m, 5 H), 1.64, (m, 1 H), 1.66 (m,1 H) , 2.0 (dd, 2 H), 4.32 (m, 3H), 4.96(q, 2 H), 5.75(m, 1 H) 7.33(m, 2 H), 7.40(m, 3 H) ,7.71(m, 2 H) ,7.79(m, 2 H), 12.39(s, 1 H) ppm。

Example 2, step 2 reaction temperature was 20 ℃; step 2, the reaction time is 24 hours; step 4, the reaction temperature is 35 ℃; the pH was 7.0 as in example 1.

Example 3, step 2 reaction temperature was 30 ℃; step 2, the reaction time is 12 hours; step 4, the reaction temperature is 40 ℃; the pH was 9.0 as in example 1.

Claims

1. A method for synthesizing (2S) -2-N-fluorenylmethoxycarbonylamino-2-methyl-6-heptenoic acid is characterized by comprising the following steps: comprises the following steps: step one, adding benzaldehyde into dichloromethane and triethylamine to obtain a compound 1, wherein the product is directly used for the next reaction without purification; secondly, adding 5-bromo-1-pentene sodium methoxide into the compound 1 and potassium tert-butoxide in tetrahydrofuran for reaction, and stirring at 20-30 ℃ for reaction for 12-24 hours to obtain a compound 2; step three, reacting the compound 2 with a hydrochloric acid solution at room temperature to obtain a compound 3; fourthly, adding lipase into the compound 3 and pure water, controlling the pH value to be 7.0-9.0, controlling the reaction temperature to be 35-40 ℃, and carrying out enzymolysis to obtain a compound 4; fifthly, reacting the compound 4 in acetone and sodium hydroxide, adding Fmoc-OSu, reacting at room temperature, and acidifying by hydrochloric acid to obtain a target compound 5, wherein the synthetic route is as follows:

。