A method for synthesizing cyclopentapeptide-4

Abstract

This invention discloses a method for synthesizing cyclic pentapeptide-4. Specifically, this invention provides a method for synthesizing compound I-5, which includes the following steps: in an organic solvent, in the presence of a fluorinated organic acid and an organic tertiary amine base, compound I-4 undergoes an intramolecular cyclization reaction to obtain compound I-5. The synthesis method provided by this invention eliminates the need for multi-step reactions and purification; direct cyclization yields an intermediate with high yield and high purity. Further reaction of the intermediate does not require further purification to obtain cyclic pentapeptide-4 with a purity higher than 99%, thus achieving high-efficiency liquid-phase synthesis of cyclic pentapeptide-4.

Description

Technical Field

[0001] This invention belongs to the field of polypeptide synthesis technology, and more specifically relates to a method for synthesizing cyclic pentapeptide-4. Background Technology

[0002] Pentapeptide-4 is an important fragment of type I collagen. Pentapeptide-4 and its derivatives can stimulate the production of collagen, elastin, and hyaluronic acid, increasing skin hydration and moisture retention, improving skin thickness, and reducing fine lines. Cyclic pentapeptide-4 is more stable than pentapeptide-4 and can enhance absorption.

[0003] Cyclic peptides are gaining increasing attention as a type of marine drug. Cyclization modification can enhance the spatial complexity of peptides, increase their specific surface area, and provide more opportunities for target contact, aiming to achieve more diverse efficacy, higher utilization of peptide raw materials, and expand the application prospects of these anti-wrinkle peptides in the field of beauty and skincare. Cyclic peptides possess a larger surface area, high affinity for targets, recognition specificity, and excellent stability. They are not easily degraded and have very strong skin permeability. Compared with the original linear structure, they are more stable and have greater affinity, showing stronger application potential in beauty, skincare, and anti-aging medical aesthetics. Cyclic peptides can bind to targets with higher affinity than linear analogs, and cyclic peptides are generally superior to linear peptide members. Peptides cyclized through their terminal amino acids are protected from exopeptidases that break down peptides from the terminal end.

[0004] There are very few literature reports on the synthesis of cyclic pentapeptide-4, and there are no published invention patents. Solid-phase synthesis uses a lot of solvents, produces a lot of waste liquid, has expensive raw materials, low yield, and high cost. Conventional liquid-phase synthesis requires protection and deprotection of each amino acid, which is a long process with many side reactions and impurities. Summary of the Invention

[0005] The technical problem to be solved by the present invention is that the existing methods for synthesizing polypeptide compounds have many steps, long cycles, and many side reactions and impurities. The present invention provides a method for synthesizing cyclic pentapeptide-4.

[0006] The present invention solves the above-mentioned technical problems through the following technical solutions.

[0007] This invention provides a method for synthesizing compounds of formula I-5, comprising the following steps: In an organic solvent, in the presence of a fluorinated organic acid and an organic tertiary amine base, compound I-4 undergoes an intramolecular cyclization reaction to yield compound I-5. ; The molar ratio of the fluorinated organic acid to the organic tertiary amine base is 1:(1.1-1.8).

[0008] In some embodiments, the molar ratio of the fluorinated organic acid and the organic tertiary amine base in the intramolecular cyclization reaction is 1:(1.2-1.5), for example, 1:1.3, 1:1.4 or 1:1.5.

[0009] In some embodiments, in the intramolecular cyclization reaction, the organic solvent is a conventional organic solvent in the art, preferably a haloalkane solvent and / or an ether solvent, more preferably a haloalkane solvent. The haloalkane solvent may be a chloromethane solvent, such as dichloromethane. The ether solvent may be a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, such as tetrahydrofuran and / or 1,4-dioxane.

[0010] In some embodiments, in the intramolecular cyclization reaction, the molar volume ratio of the compound of formula I-4 to the organic solvent is (0.1-1.0) mol / L, preferably (0.1-0.5) mol / L, for example (0.15-0.2) mol / L.

[0011] In some embodiments, the fluorinated organic acid in the intramolecular cyclization reaction is a fluorinated carboxylic acid, such as trifluoroacetic acid.

[0012] In some embodiments, in the intramolecular cyclization reaction, the molar ratio of the compound of formula I-4 to the fluorinated organic acid is 1:(1-5), preferably 1:(2.5-3.5), for example 1:3.0.

[0013] In some embodiments, in the intramolecular cyclization reaction, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine (DIEA).

[0014] In some embodiments, the organic tertiary amine base is added dropwise during the intramolecular cyclization reaction at a rate of (1.2-3.0) mol / h, preferably (1.4-2.7) mol / h.

[0015] In some embodiments, the fluorinated organic acid and the organic tertiary amine base in the intramolecular cyclization reaction are trifluoroacetic acid and DIEA.

[0016] In some embodiments, the progress of the intramolecular cyclization reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC, or NMR), and the reaction endpoint is generally defined as the disappearance of the starting material or the cessation of the reaction; the reaction time of the intramolecular cyclization reaction is 2h-5h, preferably 4.5h-5h.

[0017] In some embodiments, the reactants for the intramolecular cyclization reaction consist of the following substances: the compound of formula I-4, the organic solvent, the fluorinated organic acid, and the organic tertiary amine base.

[0018] In some embodiments, the method for synthesizing the compound of formula I-5 includes the following steps: At 30-40°C, the organic solvent is added to the mixture of the compound of formula I-4 and the fluorinated organic acid, the temperature is lowered, and the organic tertiary amine base is added to carry out an intramolecular cyclization reaction to obtain compound I-5.

[0019] In some embodiments, the cooling is from 30-40°C to -5°C to -10°C, preferably from 30-40°C to 0°C to -5°C.

[0020] In some embodiments, the method for synthesizing the compound of formula I-5 further includes a method for synthesizing the compound of formula I-4, which includes the following steps: In an organic solvent, in the presence of an activator, a condensing agent and a tertiary amine organic base, compounds I-2 and I-3 undergo a condensation reaction to obtain compound I-4. .

[0021] In some embodiments, the organic solvent is a conventional organic solvent in the art, preferably an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent. The halogenated hydrocarbon solvent may be a chloromethane solvent, such as dichloromethane. The ether solvent may be a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, such as tetrahydrofuran and / or 1,4-dioxane.

[0022] In some embodiments, the molar volume ratio of the compound of formula I-2 to the organic solvent is (0.2-1.5) mol / L, preferably (0.2-0.7) mol / L, for example 0.4 mol / L, 0.6 mol / L or 0.3 mol / L.

[0023] In some embodiments, the molar ratio of the compound of formula I-2 to the compound of formula I-3 is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1.

[0024] In some embodiments, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine (DIEA).

[0025] In some embodiments, the molar ratio of the compound of formula I-2 to the organic tertiary amine base is 1:(1.0-2.0), preferably 1:(1.0-1.5), for example 1:1.4.

[0026] In some embodiments, the activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole (HOAT) or 1-hydroxybenzotriazole (HOBT), for example 1-hydroxybenzotriazole (HOBT).

[0027] In some embodiments, the molar ratio of the compound of formula I-2 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0028] In some embodiments, the condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide (DIC).

[0029] In some embodiments, the molar ratio of the compound of formula I-2 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0030] In some embodiments, the reaction temperature of the condensation reaction is 10-50°C, preferably 20-40°C, for example 20-30°C.

[0031] In some embodiments, the progress of the condensation reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC, or NMR), generally with the disappearance of the starting material or the cessation of the reaction as the endpoint. The reaction time of the condensation reaction is preferably 1-8 hours, more preferably 2-5 hours, for example, 3 hours.

[0032] In some embodiments, the reactants for the condensation reaction consist of the following substances: the organic solvent, the activator, the condensing agent, the tertiary amine organic base, the compound of formula I-2, and the compound of formula I-3.

[0033] In some embodiments, the method for synthesizing the compound of formula I-5 further includes a method for synthesizing the compound of formula I-3, which includes the following steps: In an organic solvent, in the presence of an activator, a condensing agent, a tertiary amine organic base, and an ether solution of an inorganic acid, compound I-1 undergoes a condensation reaction with serine methyl ester to obtain compound I-3. .

[0034] In some embodiments, the organic solvent is a conventional organic solvent in the art, preferably an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent. The halogenated hydrocarbon solvent may be a chloromethane solvent, such as dichloromethane. The ether solvent may be a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, such as tetrahydrofuran and / or 1,4-dioxane.

[0035] In some embodiments, the molar volume ratio of the compound of formula I-1 to the organic solvent is (0.2-1.5) mol / L, preferably (0.5-0.8) mol / L, for example 0.5 mol / L, 0.7 mol / L or 0.8 mol / L.

[0036] In some embodiments, the molar ratio of the compound of formula I-1 to the serine methyl ester is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1.

[0037] In some embodiments, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine (DIEA).

[0038] In some embodiments, the molar ratio of the compound of formula I-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4.

[0039] In some embodiments, the activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole (HOAT) or 1-hydroxybenzotriazole (HOBT), for example 1-hydroxybenzotriazole (HOBT).

[0040] In some embodiments, the molar ratio of the compound of formula I-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0041] In some embodiments, the condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide (DIC).

[0042] In some embodiments, the molar ratio of the compound of formula I-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0043] In some embodiments, the ether solution of the inorganic acid is a saturated ether solution of the inorganic acid.

[0044] In some embodiments, the inorganic acid in the ether solution is selected from one or more of hydrochloric acid, hydrofluoric acid, hydrobromic acid, and phosphoric acid, preferably hydrochloric acid.

[0045] In some embodiments, the ether solution of the inorganic acid is a cyclic ether solution of the inorganic acid, preferably a 1,4-dioxane solution of the inorganic acid and / or a tetrahydrofuran solution of the inorganic acid.

[0046] In some embodiments, the ether solution of the inorganic acid is a 1,4-dioxane solution of saturated hydrochloric acid and / or a tetrahydrofuran solution of saturated hydrochloric acid.

[0047] In some embodiments, the molar mass ratio of the compound of formula I-1 to the ether solution of the inorganic acid is (0.3-1.5) mol / kg, preferably (0.5-1.0) mol / kg, for example (0.6-0.7) mol / kg.

[0048] In some embodiments, the reaction temperature of the condensation reaction is 10-60°C, preferably 20-50°C.

[0049] In some embodiments, the progress of the condensation reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC, or NMR), generally with the disappearance of the starting material or the cessation of the reaction as the endpoint. The reaction time of the condensation reaction is preferably 2-10 hours, more preferably 3-7 hours, for example 5 hours.

[0050] In some embodiments, the reactants for the condensation reaction consist of the following substances: the organic solvent, the activator, the condensing agent, the tertiary amine organic base, the ether solution of the inorganic acid, the compound of formula I-1, and the serine methyl ester.

[0051] In some embodiments, the method for synthesizing the compound of formula I-3 includes the following steps: At 10-60℃, a tertiary amine organic base is added to a mixture of an organic solvent, compound of formula I-1, serine methyl ester, activator and catalyst, followed by the addition of an ether solution of an inorganic acid, to carry out a condensation reaction and obtain compound of formula I-3.

[0052] In some embodiments, the method for synthesizing the compound of formula I-5 further includes a method for synthesizing the compound of formula I-2, which includes the following steps: In organic solvents and water, in the presence of an activator, a condensing agent, a tertiary amine organic base and an inorganic base, compound I-2-1 undergoes a condensation reaction with threonine methyl ester to give compound I-2. .

[0053] In some embodiments, the organic solvent is a conventional organic solvent in the art, preferably an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent. The halogenated hydrocarbon solvent may be a chloromethane solvent, such as dichloromethane. The ether solvent may be a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, such as tetrahydrofuran and / or 1,4-dioxane.

[0054] In some embodiments, the molar volume ratio of the compound of formula I-2-1 to the organic solvent is (0.1-1.5) mol / L, preferably (0.3-0.7) mol / L, for example 0.4 mol / L, 0.5 mol / L or 0.6 mol / L.

[0055] In some embodiments, the mass ratio of the organic solvent to the water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1.

[0056] In some embodiments, the molar ratio of the compound of formula I-2-1 to the threonine methyl ester is 1:(0.5-2.0), preferably 1:(0.8-1.2), for example 1:1.

[0057] In some embodiments, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine (DIEA).

[0058] In some embodiments, the molar ratio of the compound of formula I-2-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4.

[0059] In some embodiments, the activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole (HOAT) or 1-hydroxybenzotriazole (HOBT), for example 1-hydroxybenzotriazole (HOBT).

[0060] In some embodiments, the molar ratio of the compound of formula I-2-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0061] In some embodiments, the condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide (DIC).

[0062] In some embodiments, the molar ratio of the compound of formula I-2-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0063] In some embodiments, the inorganic base is a metal hydroxide base, preferably selected from one or more of sodium hydroxide, potassium hydroxide, calcium hydroxide and lithium hydroxide, and more preferably lithium hydroxide, such as lithium hydroxide monohydrate.

[0064] In some embodiments, the molar ratio of the compound of formula I-2-1 to the inorganic base is 1:(1-3), preferably 1:(1.5-2.5), for example 1:2.

[0065] In some embodiments, the molar volume ratio of the compound of formula I-2-1 to the water is (0.1-1.5) mol / L, preferably (0.3-0.8) mol / L, for example 0.4 mol / L, 0.5 mol / L or 0.7 mol / L.

[0066] In some embodiments, the reaction temperature of the condensation reaction is 10-60°C, preferably 20-50°C.

[0067] In some embodiments, the progress of the condensation reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC, or NMR), generally with the disappearance of the starting material or the cessation of the reaction as the endpoint. The reaction time of the condensation reaction is preferably 2-10 hours, more preferably 3-7 hours, for example, 6 hours.

[0068] In some embodiments, the reaction raw materials for the condensation reaction consist of the following substances: the organic solvent, water, the activator, the condensing agent, the tertiary amine organic base, the inorganic base, the compound of formula I-2-1, and the threonine methyl ester.

[0069] In some embodiments, the method for synthesizing the compound of formula I-2 includes the following steps: An organic tertiary amine base is added to a mixture of an organic solvent, a compound of formula I-2-1, threonine methyl ester, an activator, and a condensing agent. Then, water and an inorganic base are added to carry out a condensation reaction to obtain a compound of formula I-2.

[0070] In some embodiments, the method for synthesizing the compound of formula I-5 further includes a method for synthesizing the compound of formula I-2-1, which includes the following steps: In an organic solvent and water, in the presence of an activator, a condensing agent, a tertiary amine organic base and an inorganic base, the compound of formula I-1 undergoes a condensation reaction with threonine methyl ester to give the compound of formula I-2-1. .

[0071] In some embodiments, the organic solvent is a conventional organic solvent in the art, preferably an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent. The halogenated hydrocarbon solvent may be a chloromethane solvent, such as dichloromethane. The ether solvent may be a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, such as tetrahydrofuran and / or 1,4-dioxane.

[0072] In some embodiments, the molar volume ratio of the compound of formula I-1 to the organic solvent is (0.1-1.5) mol / L, preferably (0.4-0.9) mol / L, for example 0.8 mol / L, 0.5 mol / L or 0.7 mol / L.

[0073] In some embodiments, the mass ratio of the organic solvent to the water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1.

[0074] In some embodiments, the molar ratio of the compound of formula I-1 to the threonine methyl ester is 1:(0.5-2.0), preferably 1:(0.8-1.2), for example 1:1.

[0075] In some embodiments, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine (DIEA).

[0076] In some embodiments, the molar ratio of the compound of formula I-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4.

[0077] In some embodiments, the activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole (HOAT) or 1-hydroxybenzotriazole (HOBT), for example 1-hydroxybenzotriazole (HOBT).

[0078] In some embodiments, the molar ratio of the compound of formula I-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0079] In some embodiments, the condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide (DIC).

[0080] In some embodiments, the molar ratio of the compound of formula I-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1.

[0081] In some embodiments, the inorganic base is a metal hydroxide base, preferably selected from one or more of sodium hydroxide, potassium hydroxide, calcium hydroxide and lithium hydroxide, and more preferably lithium hydroxide, such as lithium hydroxide monohydrate.

[0082] In some embodiments, the molar ratio of the compound of formula I-1 to the inorganic base is 1:(1-3), preferably 1:(1.5-2.5), for example 1:1.9.

[0083] In some embodiments, the molar volume ratio of the compound of formula I-1 to the water is (0.1-1.5) mol / L, preferably (0.4-1.0) mol / L, for example 0.9 mol / L, 0.5 mol / L or 0.7 mol / L.

[0084] In some embodiments, the reaction temperature of the condensation reaction is 10-60°C, preferably 20-50°C.

[0085] In some embodiments, the progress of the condensation reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC, or NMR), generally with the disappearance of the starting material or the cessation of the reaction as the endpoint. The reaction time of the condensation reaction is preferably 2-10 hours, more preferably 3-7 hours, for example, 6 hours.

[0086] In some embodiments, the reaction raw materials for the synthesis method of Formula I-2-1 consist of the following substances: the organic solvent, the water, the activator, the condensing agent, the tertiary amine organic base, the inorganic base, the compound of Formula I-1, and the threonine methyl ester.

[0087] In some embodiments, the synthesis method of formula I-2-1 includes the following steps: An organic tertiary amine base is added to a mixture of an organic solvent, a compound of formula I-1, serine methyl ester, an activator, and a condensing agent, followed by the addition of water and an inorganic base, to carry out a condensation reaction and obtain a compound of formula I-2-1.

[0088] The present invention also provides a method for synthesizing a compound of formula I, comprising the following steps: (1) Compound I-5 was prepared by the method described above; (2) In a mixed solvent of ether solvent and water, in the presence of a catalyst and a hydrogen source, compounds of formula I-5 undergo a reduction reaction to obtain compound I; .

[0089] In some embodiments, the ether solvent in the reduction reaction is a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as 1,4-dioxane and / or tetrahydrofuran.

[0090] In some embodiments, in the reduction reaction, the mass ratio of the ether solvent to water in the mixed solvent of the ether solvent and water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1.

[0091] In some embodiments, in the reduction reaction, the molar volume ratio of the compound of formula I-5 to the ether solvent is (0.1-1.0) mol / L, preferably (0.3-0.6) mol / L, for example 0.5 mol / L, 0.35 mol / L or 0.54 mol / L.

[0092] In some embodiments, the catalyst in the reduction reaction is a metal catalyst, preferably a palladium catalyst, more preferably a zero-valent palladium catalyst, for example a 10% wt palladium on carbon catalyst.

[0093] In some embodiments, the molar ratio of the compound of formula I-5 to the catalyst in the reduction reaction is 1:(0.3-2.0), preferably 1:(0.4-1.0), for example 1:0.77, 1:0.58 or 1:0.4.

[0094] In some embodiments, the hydrogen source in the reduction reaction is hydrogen gas.

[0095] In some embodiments, during the reduction reaction, when the hydrogen source is hydrogen gas, the pressure of the hydrogen gas is 0.3-1.5 MPa, preferably 0.5-1.0 MPa, for example 0.6-0.7 MPa.

[0096] In some embodiments, the reduction reaction is carried out at a temperature of 20-60°C, preferably 30-50°C, for example 40-50°C.

[0097] In some embodiments, the progress of the reduction reaction can be detected using conventional monitoring methods in the art (e.g., TLC, HPLC or NMR), and the reaction endpoint is generally defined as the disappearance of the starting material or the cessation of the reaction; the reaction time of the reduction reaction is 1-8 hours, preferably 2-5 hours, for example 3 hours.

[0098] In some embodiments, the reaction feedstock for the reduction reaction consists of the following substances: the ether solvent and the water mixture, the catalyst, the compound of formula I-5, and the hydrogen source.

[0099] In some embodiments, the synthesis method of the compound of formula I further includes crystallization, wherein the crystallization solution is a nitrile solvent or a ketone solvent. The nitrile solvent may be acetonitrile. The ketone solvent may be acetone.

[0100] The present invention also provides a method for synthesizing compounds of formula I-4, which includes the following steps: In an organic solvent, in the presence of an activator, a condensing agent and a tertiary amine organic base, compounds I-2 and I-3 undergo a condensation reaction to obtain compound I-4. .

[0101] In some embodiments, the reaction conditions for synthesizing the compound of formula I-4 are as described above.

[0102] The present invention also provides a compound of formula I-4: .

[0103] The present invention also provides a compound of formula I-5: .

[0104] Based on common knowledge in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0105] The reagents and raw materials used in this invention are all commercially available.

[0106] The positive and progressive effects of this invention are as follows: the synthesis method provided by this invention does not require multiple reaction steps and purification. It can directly cyclize to obtain intermediates with high yield and high purity. After further reaction of the intermediates, cyclic pentapeptide-4 with a purity of more than 99% can be obtained without further purification, thus realizing the high-efficiency liquid phase synthesis of cyclic pentapeptide-4. Detailed Implementation

[0107] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0108] Example 1

[0109] Step 1: Synthesis of Cbz-Lys(Boc)-Thr-OH

[0110] In a 5L three-necked flask, 1 kg (1.12 L) of tetrahydrofuran, 200 g (0.53 mol) of Cbz-Lys(Boc)-OH, 70 g (0.53 mol) of threonine methyl ester, and 106.55 g (0.79 mol) of 1-hydroxybenzotriazole were added. Under ice bath conditions, 99.52 g (0.79 mol) of DIC (N,N'-diisopropylcarbodiimide) was added, followed by dropwise addition of 27.18 g (0.21 mol) of DIEA. After the addition was complete, the reaction temperature was raised to 25±2℃ and reacted for 3 hours. Then, 1 kg of water and 44.12 g (1 mol) of lithium hydroxide monohydrate were added, and the reaction was continued at 45±2℃ for 3 hours. The pH was then adjusted to neutral, and the mixture was concentrated at 45℃ for 1 hour. The pH of the residue was adjusted to 3.8-4.0, and crystals were grown for 2 hours. The mixture was then filtered, and the filter cake was dried under vacuum at 45±2℃ to obtain the product. Yield: 95.0%. Step 2: Synthesis of Cbz-Lys(Boc)-Thr-Thr-OH

[0111] 1202.5 g (1.35 L) of tetrahydrofuran, 240.5 g (0.5 mol) of Cbz-Lys(Boc)-Thr-OH, 66.5 g (0.5 mol) of threonine methyl ester, and 101.23 g (0.75 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 94.54 g (0.75 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 25.82 g (0.2 mol) of DIEA. After reacting at 25 ± 2 °C for 3 h, 1202.5 g of water and 41.91 g (1 mol) of lithium hydroxide monohydrate were added. The reaction was continued at 45 ± 2 °C for 3 h. The pH was then adjusted to neutral, and the mixture was concentrated. The pH of the residue was adjusted to 3.8-4.0, and crystals were allowed to grow for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product. Yield: 95.8%. Step 3: Synthesis of H-Lys(Cbz)-Ser-Ome

[0112] 1 kg (1.12 L) of tetrahydrofuran, 200 g (0.53 mol) of Boc-Lys(Cbz)-OH, 62.62 g (0.53 mol) of serine methyl ester, and 106.55 g (0.79 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 99.52 g (0.79 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 27.18 g (0.21 mol) of DIEA. After reacting at 25 ± 2 °C for 2 h, 800 g of a saturated solution of tetrahydrofuran hydrochloride was added, and the reaction was continued at 45 ± 2 °C for 3 h. Then, 3 kg of methyl tert-butyl ether was added, and crystallization was allowed to continue for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 95.3%. Step 4: Synthesize Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-Ome

[0113] 1.116 kg (1.25 L) of tetrahydrofuran, 279 g (0.48 mol) of Cbz-Lys(Boc)-Thr-Thr-OH, 182.65 g (0.48 mol) of H-Lys(Cbz)-Ser-OMe, and 97.06 g (0.72 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 90.65 g (0.72 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 86.65 g (0.67 mol) of DIEA. After reacting at 25 ± 2 °C for 3 h, 2.232 kg of water was added, and crystallization was allowed to continue for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 94.0%. Step 5: Synthesize Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser)

[0114] 282.79 g (0.30 mol) of Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe and 102.64 g (0.9 mol) of trifluoroacetic acid were added to a 5 L three-necked flask. After reacting at an internal temperature of 35 ± 2 °C for 3 h, 2554.8 g (1.93 L) of dichloromethane was added, and the internal temperature was lowered to 0-5 °C. 174.5 g of DIEA (1.35 mol) was added dropwise over a period of 30-40 min at an internal temperature of 0-5 °C for 1 h. The dichloromethane was concentrated, and 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C for 24 h. The dried solid was dissolved in 1277.4 g of tetrahydrofuran, and then 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C. The product was obtained with a yield of 86.0%. Step Six: Synthesis of Cyclic Pentapeptide-4

[0115] 315 g (0.39 mol) Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser), 945 g (1.1 L) tetrahydrofuran, 945 g water, and 31.5 g (0.3 mol) 10% palladium on carbon catalyst were added to a 10 L hydrogenation reactor. The reaction was carried out at an internal temperature of 45±2℃ and a hydrogen pressure of 0.6-0.7 MPa for 3 h. The solution was filtered and separated into layers. The lower aqueous phase was concentrated to 730 g, and 1890 g acetonitrile was added. Crystallization was carried out for 2 h. The solution was then filtered and the filter cake was dried under vacuum at 45±2℃ to obtain the product with a yield of 71.0% and a purity of 99.60%.

[0116] Example 2

[0117] Step 1: Synthesis of Cbz-Lys(Boc)-Thr-OH

[0118] In a 5L three-necked flask, 800g (0.78L) of 1,4-dioxane, 200g (0.53mol) of Cbz-Lys(Boc)-OH, 70g (0.53mol) of threonine methyl ester, and 92.35g (0.68mol) of 1-hydroxybenzotriazole were added. Under ice bath conditions, 86.25g (0.68mol) of DIC was added, followed by dropwise addition of 27.18g (0.21mol) of DIEA. After the addition was complete, the reaction temperature was raised to 25±2℃ and reacted for 3 hours. Then, 800g of water and 44.12g (1mol) of lithium hydroxide monohydrate were added, and the reaction was continued at 45±2℃ for 3 hours. The pH was then adjusted to neutral, and the mixture was concentrated at 45℃ for 1 hour. The pH of the residue was adjusted to 3.8-4.0, and crystals were grown for 2 hours. The mixture was then filtered, and the filter cake was dried under vacuum at 45±2℃ to obtain the product. The yield was 94.8%. Step 2: Synthesis of Cbz-Lys(Boc)-Thr-Thr-OH Add 960 g (0.93 L) of 1,4-dioxane, 240.0 g (0.5 mol) of Cbz-Lys(Boc)-Thr-OH, 66.36 g (0.5 mol) of threonine methyl ester, and 87.55 g (0.65 mol) of 1-hydroxybenzotriazole to a 5 L three-necked flask. Add 81.77 g (0.65 mol) of DIC under ice bath conditions, followed by dropwise addition of 25.77 g (0.2 mol) of DIEA. After reacting at an internal temperature of 25 ± 2 °C for 3 h, add 960 g of water and 41.83 g (1 mol) of lithium hydroxide monohydrate. React at 45 ± 2 °C for 3 h. Adjust the pH to neutral, then concentrate the product. Adjust the pH of the residue to 3.8-4.0, allow it to crystallize for 2 h, filter, and dry the filter cake under vacuum at 45 ± 2 °C to obtain the product. Yield: 96.0%. Step 3: Synthesis of H-Lys(Cbz)-Ser-OMe 800 g (0.77 L) of 1,4-dioxane, 200 g (0.53 mol) of Boc-Lys(Cbz)-OH, 62.62 g (0.53 mol) of serine methyl ester, and 92.35 g (0.68 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 86.25 g (0.68 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 27.18 g (0.21 mol) of DIEA. After reacting at an internal temperature of 25 ± 2 °C for 2 h, 800 g of a saturated solution of 1,4-dioxane hydrogen chloride was added, and the reaction was continued at 45 ± 2 °C for 3 h. Then, 2.4 kg of methyl tert-butyl ether was added, and crystallization was allowed to continue for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 94.9%. Step 4: Synthesize Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe 836.4 g (0.8 L) of 1,4-dioxane, 278.8 g (0.48 mol) of Cbz-Lys(Boc)-Thr-Thr-OH, 182.51 g (0.48 mol) of H-Lys(Cbz)-Ser-OMe, and 84.06 g (0.62 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 78.5 g (0.62 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 86.58 g (0.67 mol) of DIEA. After reacting at 25 ± 2 °C for 3 h, 1.673 kg of water was added, and crystallization was allowed to occur for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 93.8%. Step 5: Synthesize Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser) 246.87 g (0.26 mol) of Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe and 89.6 g (0.79 mol) of trifluoroacetic acid were added to a 5 L three-necked flask. After reacting at an internal temperature of 35 ± 2 °C for 3 h, 2124 g (1.6 L) of dichloromethane was added, and the internal temperature was lowered to 0-5 °C. 132.03 g (1.02 mol) of DIEA was added dropwise, and the reaction was carried out for 30-40 min at an internal temperature of 0-5 °C for 1 h. The dichloromethane was concentrated, and 2336.4 g of water was added, and the mixture was allowed to crystallize for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C for 24 h. The dried solid was dissolved in 1062 g of tetrahydrofuran, and then 2124 g of water was added, and the mixture was allowed to crystallize for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 85.5%. Step Six: Synthesis of Cyclic Pentapeptide-4 312.5 g (0.38 mol) Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser), 781.25 g (0.76 L) 1,4-dioxane, 781.25 g water, and 23.4 g (0.22 mol) 10% palladium on carbon catalyst were added to a 10 L hydrogenation reactor. The reaction was carried out at an internal temperature of 45±2℃ and a hydrogen pressure of 0.6-0.7 MPa for 3 h. The solution was filtered, separated into layers, and the lower aqueous phase was concentrated to 625 g. 1562.5 g acetonitrile was added, and crystals were grown for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45±2℃ to obtain the product with a yield of 70.6% and a purity of 99.14%.

[0119] Example 3

[0120] Step 1: Synthesis of Cbz-Lys(Boc)-Thr-OH

[0121] In a 5L three-necked flask, 600g (0.67L) of tetrahydrofuran, 200g (0.53mol) of Cbz-Lys(Boc)-OH, 70g (0.53mol) of threonine methyl ester, and 78.14g (0.58mol) of 1-hydroxybenzotriazole were added. Under ice bath conditions, 72.98g (0.58mol) of DIC was added, followed by dropwise addition of 27.18g (0.21mol) of DIEA. After the addition was complete, the reaction temperature was raised to 25±2℃ and reacted for 3 hours. Then, 600g of water and 44.12g (1mol) of lithium hydroxide monohydrate were added, and the reaction was continued at 45±2℃ for 3 hours. The pH was then adjusted to neutral, and the mixture was concentrated at 45℃ for 1 hour. The pH of the residue was adjusted to 3.8-4.0, and crystals were grown for 2 hours. The mixture was then filtered, and the filter cake was dried under vacuum at 45±2℃ to obtain the product. The yield was 94.7%. Step 2: Synthesis of Cbz-Lys(Boc)-Thr-Thr-OH 719.4 g (0.8 L) tetrahydrofuran, 239.8 g (0.5 mol) Cbz-Lys(Boc)-Thr-OH, 66.3 g (0.5 mol) threonine methyl ester, and 74.02 g (0.55 mol) 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 69.13 g (0.55 mol) DIC was added under ice bath conditions, followed by dropwise addition of 25.74 g (0.2 mol) DIEA. After reacting at 25 ± 2 °C for 3 h, 719.4 g of water and 41.79 g (1 mol) lithium hydroxide monohydrate were added, and the reaction was continued at 45 ± 2 °C for 3 h. The pH was then adjusted to neutral, and the mixture was concentrated. The pH of the residue was adjusted to 3.8-4.0, and crystals were grown for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product. Yield: 95.8%. Step 3: Synthesis of H-Lys(Cbz)-Ser-OMe 600 g (0.67 L) of tetrahydrofuran, 200 g (0.53 mol) of Boc-Lys(Cbz)-OH, 62.62 g (0.53 mol) of serine methyl ester, and 78.14 g (0.58 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 72.98 g (0.58 mol) of DIC was added under ice bath conditions, followed by the dropwise addition of 27.18 g (0.21 mol) of DIEA. After reacting at an internal temperature of 25 ± 2 °C for 2 h, 800 g of a saturated solution of tetrahydrofuran hydrochloride was added, and the reaction was continued at 45 ± 2 °C for 3 h. Then, 1.8 kg of methyl tert-butyl ether was added, and crystallization was allowed to continue for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 95.1%. Step 4: Synthesize Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe 1390 g (1.6 L) of tetrahydrofuran, 278.1 g (0.48 mol) of Cbz-Lys(Boc)-Thr-Thr-OH, 182.06 g (0.48 mol) of H-Lys(Cbz)-Ser-OMe, and 70.95 g (0.53 mol) of 1-hydroxybenzotriazole were added to a 5 L three-necked flask. 66.26 g (0.53 mol) of DIC was added under ice bath conditions, followed by dropwise addition of 86.37 g (0.67 mol) of DIEA. After reacting at 25 ± 2 °C for 3 h, 2.78 kg of water was added, and crystallization was allowed to occur for 2 h. The mixture was then filtered, and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product, with a yield of 94.0%. Step 5: Synthesize Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser) 211.41 g (0.22 mol) of Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe and 76.73 g (0.67 mol) of trifluoroacetic acid were added to a 5 L three-necked flask. After reacting at an internal temperature of 35 ± 2 °C for 3 h, 1697.6 g (1.28 L) of dichloromethane was added, and the internal temperature was lowered to 0-5 °C. 121.76 g (0.94 mol) of DIEA was added dropwise over a period of 30-40 min at an internal temperature of 0-5 °C for 1 h. The dichloromethane was concentrated, and 2122 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C for 24 h. The dried solid was dissolved in 848.8 g of tetrahydrofuran, and then 1697.6 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C. The product was obtained with a yield of 85.0%. Step Six: Synthesis of Cyclic Pentapeptide-4 310.5 g (0.38 mol) Cyclo (Cbz-Lys-Thr-Thr-Lys(Cbz)-Ser), 621 g (0.7 L) tetrahydrofuran, 621 g water, and 15.6 g (0.15 mol) 10% palladium on carbon catalyst were added to a 10 L hydrogenation reactor. The reaction was carried out at an internal temperature of 45 ± 2 °C and a hydrogen pressure of 0.6-0.7 MPa for 3 h. The solution was filtered and separated into layers. The lower aqueous phase was concentrated to 621 g, and 1242 g acetone was added. Crystallization was carried out for 2 h. The solution was then filtered and the filter cake was dried under vacuum at 45 ± 2 °C to obtain the product with a yield of 72.2% and a purity of 99.17%.

[0122] Comparative Example 1

[0123] The preparation method is the same as in Example 1, except that in the preparation process of this comparative example, step three follows the conventional synthesis method, as follows: Step 3: Synthesis of H-Lys(Cbz)-Ser-OMe Add 1 kg tetrahydrofuran, 220 g Boc-Lys(Cbz)-OH, 68.88 g serine methyl ester, and 117.2 g 1-hydroxybenzotriazole to a 5 L three-necked flask. Add 109.5 g DIC under ice bath conditions, then add 29.9 g of [a specific ingredient, likely DIC] dropwise. DIEA; after reacting at an internal temperature of 25±2℃ for 2 hours, tetrahydrofuran was concentrated at 45±2℃; then added to 1.1 kg ethyl acetate and 1.1 kg water, pH adjusted to 9.5, stirred for 20 min, allowed to stand for 20 min, and separated; the aqueous phase was extracted once with 600 g ethyl acetate; the organic phases were combined and washed twice with 1.1 kg water at pH=9.5; the organic phase was washed three times with 1.1 kg water at pH=2.0, washed once with 1.1 kg saturated brine, dried with 60 g anhydrous magnesium sulfate for 30 min, filtered, and the filtrate was concentrated at 45℃ to obtain a white solid; then 220 g trifluoroacetic acid was added, reacted at 45±2℃ for 3 hours, added to 3.3 kg methyl tert-butyl ether, crystallized for 2 hours, filtered, and the filter cake was dried under vacuum at 45±2℃ to obtain the product, with a yield of 82.0%.

[0124] Comparative Example 2

[0125] The preparation method is the same as in Example 1, except that in step three of the preparation process of this comparative example, the saturated solution of tetrahydrofuran hydrogen chloride is replaced with a saturated solution of ethyl acetate hydrogen chloride to obtain the product, with a yield of 85.8%.

[0126] Comparative Example 3

[0127] The preparation method is the same as in Example 1, except that the addition ratio of trifluoroacetic acid to DIEA is different in step five, and the addition method is also different. Specifically, DIEA is added rapidly in step five, with a trifluoroacetic acid:DIEA ratio of 1:1, resulting in a low cyclization yield. Details are as follows: 282.8 g of Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe and 102.64 g of trifluoroacetic acid were added to a 5 L three-necked flask. After reacting at an internal temperature of 35 ± 2 °C for 3 h, 2554.8 g of dichloromethane was added, and the internal temperature was lowered to 0-5 °C. 116.34 g of DIEA was added dropwise over 10 min, and the reaction was carried out at 0-5 °C for 1 h. The dichloromethane was concentrated, and 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C for 24 h. The dried solid was dissolved in 1277.4 g of tetrahydrofuran, and then 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C. The product was obtained with a yield of 56.5%.

[0128] Comparative Example 4

[0129] The preparation method is the same as in Example 1, except that the addition ratio of trifluoroacetic acid to DIEA is different in step five of the preparation process of this comparative example, and the addition method is also different. Specifically, DIEA is added slowly in step five, with a trifluoroacetic acid:DIEA ratio of 1:2. The cyclization yield is low, as detailed below: 282.8 g of Cbz-Lys(Boc)-Thr-Thr-Lys(Cbz)-Ser-OMe and 102.64 g of trifluoroacetic acid were added to a 5 L three-necked flask. After reacting at an internal temperature of 35 ± 2 °C for 3 h, 2554.8 g of dichloromethane was added, and the internal temperature was lowered to 0-5 °C. 232.63 g of DIEA was added dropwise over a period of 60 min at an internal temperature of 0-5 °C for 1 h. The dichloromethane was concentrated, and 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C for 24 h. The dried solid was dissolved in 1277.4 g of tetrahydrofuran, and then 2554.8 g of water was added. Crystallization was carried out for 2 h. The mixture was filtered, and the filter cake was dried under vacuum at 45 ± 2 °C. The product was obtained with a yield of 65.8%.

[0130] Table 1: Comparison of product indicators in different embodiments and comparative examples

[0131] The experimental data analysis is as follows: (1) Compared with Comparative Example 1, the difference lies in the fact that step three uses a specific hydrochloric acid gas / organic solvent saturated solution to remove BOC, obtaining a qualified intermediate in one step without purification; wherein, the yield of step three of the present invention is as high as 95.3%, while that of Comparative Example 1 is only 82.0% according to conventional synthesis methods. (2) Compared with Example 1, the difference in Comparative Example 2 is that in step three, the saturated solution of tetrahydrofuran hydrogen chloride is replaced with the saturated solution of ethyl acetate hydrogen chloride. Although Comparative Example 2 uses hydrochloric acid gas / organic solvent saturated solution to remove BOC, the yield of Comparative Example 2 is not substantially improved, only 85.8%, which is not much different from the 82.0% of the conventional synthesis method of Comparative Example 1. (3) Compared with Example 1, the difference in Comparative Example 3 is that the addition ratio of trifluoroacetic acid and DIEA is different in step five of Comparative Example 3. The ratio of trifluoroacetic acid to DIEA is 1:1, and the cyclization yield is low. The comparison revealed that although Comparative Example 3 used the direct cyclization method of the present invention, the yield of the intermediate in step five of Comparative Example 3 was only 56.5% due to the different addition ratio of trifluoroacetic acid and DIEA, which was far lower than the 86.0% of the present invention. (4) The difference between Comparative Example 4 and Example 1 is that in step five of Comparative Example 4, the ratio of trifluoroacetic acid to DIEA is different, with trifluoroacetic acid:DIEA = 1:2, resulting in a lower cyclization yield. The comparison revealed that although Comparative Example 4 used the direct cyclization method of the present invention, the yield of the intermediate in step five of Comparative Example 4 was only 65.8% due to the different addition ratio of trifluoroacetic acid and DIEA, which was much lower than the 86.0% of the present invention. A comparison between Example 1 and Comparative Examples 3-4 shows that: The amount of DIEA added affected the yield of the intermediate in step five of Comparative Examples 3-4, and the amount of DIEA added was related to the amount of trifluoroacetic acid added. Therefore, it can be seen that the amount of DIEA added cannot be adjusted through a limited number of experiments. This invention creatively achieves direct cyclization by adding an excess of trifluoroacetic acid and DIEA in a specific ratio, and effectively avoids hydrolysis or ring-opening that may be caused by local high concentrations of DIEA, thereby greatly avoiding the generation of impurities.

[0132] In summary: This method prepares cyclic pentapeptide-4 via liquid-phase synthesis. After recrystallization once, it is directly hydrogenated and crystallized, resulting in a simple, rapid synthesis that yields products with high purity and yield. This method innovatively uses specific hydrochloric acid gas / organic solvent solutions to remove BOC, especially hydrochloric acid gas / tetrahydrofuran or hydrochloric acid gas / 1,4-dioxane, to obtain qualified intermediates in one step without purification, unexpectedly avoiding the generation of impurities and greatly improving the yield. This method innovatively incorporates a limited ratio of trifluoroacetic acid and DIEA, achieving direct cyclization and significantly improving the yield.

Claims

1. A method for synthesizing a compound of formula I-5, comprising the following steps: In an organic solvent, in the presence of a fluorinated organic acid and an organic tertiary amine base, compound I-4 undergoes an intramolecular cyclization reaction to yield compound I-5. ； The molar ratio of the fluorinated organic acid to the organic tertiary amine base is 1:(1.1-1.8).

2. The synthesis method according to claim 1, characterized in that, It meets one or more of the following conditions: (1) In the intramolecular cyclization reaction, the molar ratio of the fluorinated organic acid and the organic tertiary amine base is 1:(1.2-1.5), for example, 1:1.3, 1:1.4 or 1:1.5; (2) In the intramolecular cyclization reaction, the organic solvent is a halohydrocarbon solvent and / or an ether solvent, more preferably a halohydrocarbon solvent; The halocarbon solvent may be a chloromethane solvent, such as dichloromethane; The ether solvent may be a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as tetrahydrofuran and / or 1,4-dioxane; (3) In the intramolecular cyclization reaction, the molar volume ratio of the compound of formula I-4 to the organic solvent is (0.1-1.0) mol / L, preferably (0.1-0.5) mol / L, for example (0.15-0.2) mol / L; (4) In the intramolecular cyclization reaction, the fluorinated organic acid is a fluorinated carboxylic acid, such as trifluoroacetic acid; (5) In the intramolecular cyclization reaction, the molar ratio of the compound of formula I-4 to the fluorinated organic acid is 1:(1-5), preferably 1:(2.5-3.5), for example 1:3.0; (6) In the intramolecular cyclization reaction, the organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine; (7) In the intramolecular cyclization reaction, the organic tertiary amine base is added dropwise at a rate of (1.2-3.0) mol / h, preferably (1.4-2.7) mol / h; (8) The reaction time of the intramolecular cyclization reaction is 2h-5h, preferably 4.5h-5h; (9) The reactants for the intramolecular cyclization reaction consist of the following substances: the compound of formula I-4, the organic solvent, the fluorinated organic acid, and the organic tertiary amine base; (10) The method for synthesizing the compound of formula I-5 includes the following steps: at 30-40°C, the organic solvent is added to the mixture of the compound of formula I-4 and the fluorinated organic acid, the temperature is lowered, the organic tertiary amine base is added, and an intramolecular cyclization reaction is carried out to obtain the compound of formula I-5; Preferably, the temperature drop is from 30-40℃ to -5℃-10℃, and more preferably from 30-40℃ to 0℃-5℃.

3. The synthesis method as described in claim 1, characterized in that, It includes a method for synthesizing compounds of formula I-4, the method comprising the following steps: In an organic solvent, in the presence of an activator, a condensing agent and a tertiary amine organic base, compounds I-2 and I-3 undergo a condensation reaction to obtain compound I-4. ； Preferably, the method for synthesizing the compound of formula I-4 satisfies one or more of the following conditions: (1) The organic solvent is an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent; The halocarbon solvent may be a chloromethane solvent, such as dichloromethane; The ether solvent may be a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as tetrahydrofuran and / or 1,4-dioxane; (2) The molar volume ratio of the compound of formula I-2 to the organic solvent is (0.2-1.5) mol / L, preferably (0.2-0.7) mol / L, for example 0.4 mol / L, 0.6 mol / L or 0.3 mol / L; (3) The molar ratio of the compound of formula I-2 to the compound of formula I-3 is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1; (4) The organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine; (5) The molar ratio of the compound of formula I-2 to the organic tertiary amine base is 1:(1.0-2.0), preferably 1:(1.0-1.5), for example 1:1.4; (6) The activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole or 1-hydroxybenzotriazole, for example 1-hydroxybenzotriazole; (7) The molar ratio of the compound of formula I-2 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (8) The condensing agent is a carbodiamine condensing agent, for example, N,N'-diisopropylcarbodiimide; (9) The molar ratio of the compound of formula I-2 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (10) The reaction temperature of the condensation reaction is 10-50℃, preferably 20-40℃, for example 20-30℃; (11) The reaction time of the condensation reaction is 1-8h, more preferably 2-5h, for example 3h; (12) The reaction raw materials for the condensation reaction consist of the following substances: the organic solvent, the activator, the condensing agent, the tertiary amine organic base, the compound of formula I-2 and the compound of formula I-3.

4. The synthesis method according to claim 1, characterized in that, It includes a method for synthesizing compounds of formula I-3, the method comprising the following steps: In an organic solvent, in the presence of an activator, a condensing agent, a tertiary amine organic base, and an ether solution of an inorganic acid, compound I-1 undergoes a condensation reaction with serine methyl ester to obtain compound I-3. ； Preferably, the method for synthesizing the compound of formula I-3 satisfies one or more of the following conditions: (1) The organic solvent is an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent; The halocarbon solvent may be a chloromethane solvent, such as dichloromethane; The ether solvent may be a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as tetrahydrofuran and / or 1,4-dioxane; (2) The molar volume ratio of the compound of formula I-1 to the organic solvent is (0.2-1.5) mol / L, preferably (0.5-0.8) mol / L, for example 0.5 mol / L, 0.7 mol / L or 0.8 mol / L; (3) The molar ratio of the compound of formula I-1 to the serine methyl ester is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1; (4) The organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine; (5) The molar ratio of the compound of formula I-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4; (6) The activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole or 1-hydroxybenzotriazole, for example 1-hydroxybenzotriazole; (7) The molar ratio of the compound of formula I-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (8) The condensing agent is a carbodiamine condensing agent, for example, N,N'-diisopropylcarbodiimide; (9) The molar ratio of the compound of formula I-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (10) The ether solution of the inorganic acid is a saturated ether solution of the inorganic acid; (11) In the ether solution of the inorganic acid, the inorganic acid is selected from one or more of hydrochloric acid, hydrofluoric acid, hydrobromic acid and phosphoric acid, preferably hydrochloric acid; (12) The ether solution of the inorganic acid is a cyclic ether solution of the inorganic acid, preferably a 1,4-dioxane solution of the inorganic acid and / or a tetrahydrofuran solution of the inorganic acid; (13) The molar mass ratio of the compound of formula I-1 to the ether solution of the inorganic acid is (0.3-1.5) mol / kg, preferably (0.5-1.0) mol / kg, for example (0.6-0.7) mol / kg; (14) The reaction temperature of the condensation reaction is 10-60℃, preferably 20-50℃; (15) The reaction time of the condensation reaction is 2-10h, more preferably 3-7h, for example 5h; (16) The reaction raw materials for the condensation reaction consist of the following substances: the organic solvent, the activator, the condensing agent, the tertiary amine organic base, the ether solution of the inorganic acid, the compound of formula I-1, and the serine methyl ester; (17) The method for synthesizing the compound of formula I-3 includes the following steps: at 10-60°C, a tertiary amine organic base is added to a mixture of organic solvent, compound of formula I-1, serine methyl ester, activator and catalyst, and then an ether solution of inorganic acid is added to carry out a condensation reaction to obtain compound of formula I-3. More preferably, the method for synthesizing the compound of formula I-3 satisfies the following conditions: The ether solution of the inorganic acid is a 1,4-dioxane solution of saturated hydrochloric acid and / or a tetrahydrofuran solution of saturated hydrochloric acid.

5. The synthesis method according to claim 1, characterized in that, It includes a method for synthesizing a compound of formula I-2, the method comprising the following steps: In organic solvents and water, in the presence of an activator, a condensing agent, a tertiary amine organic base and an inorganic base, compound I-2-1 undergoes a condensation reaction with threonine methyl ester to give compound I-2. ； Preferably, the method for synthesizing the compound of formula I-2 satisfies one or more of the following conditions: (1) The organic solvent is an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent; The halocarbon solvent may be a chloromethane solvent, such as dichloromethane; The ether solvent may be a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as tetrahydrofuran and / or 1,4-dioxane; (2) The molar volume ratio of the compound of formula I-2-1 to the organic solvent is (0.1-1.5) mol / L, preferably (0.3-0.7) mol / L, for example 0.4 mol / L, 0.5 mol / L or 0.6 mol / L; (3) The mass ratio of the organic solvent to the water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1; (4) The molar ratio of the compound of formula I-2-1 to the threonine methyl ester is 1:(0.5-2.0), preferably 1:(0.8-1.2), for example 1:1; (5) The organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine; (6) The molar ratio of the compound of formula I-2-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4; (7) The activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole or 1-hydroxybenzotriazole, for example 1-hydroxybenzotriazole; (8) The molar ratio of the compound of formula I-2-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (9) The condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide; (10) The molar ratio of the compound of formula I-2-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (11) The inorganic base is a metal hydroxide base, preferably selected from one or more of sodium hydroxide, potassium hydroxide, calcium hydroxide and lithium hydroxide, and more preferably lithium hydroxide, such as lithium hydroxide monohydrate; (12) The molar ratio of the compound of formula I-2-1 to the inorganic base is 1:(1-3), preferably 1:(1.5-2.5), for example 1:2; (13) The molar volume ratio of the compound of formula I-2-1 to the water is (0.1-1.5) mol / L, preferably (0.3-0.8) mol / L, for example 0.4 mol / L, 0.5 mol / L or 0.7 mol / L; (14) The reaction temperature of the condensation reaction is 10-60℃, preferably 20-50℃; (15) The reaction time of the condensation reaction is 2-10h, more preferably 3-7h, for example 6h; (16) The reaction raw materials for the condensation reaction consist of the following substances: the organic solvent, water, the activator, the condensing agent, the tertiary amine organic base, the inorganic base, the compound of formula I-2-1, and the threonine methyl ester; (17) The method for synthesizing the compound of formula I-2 includes the following steps: adding an organic tertiary amine base to a mixture of an organic solvent, the compound of formula I-2-1, threonine methyl ester, an activator and a condensing agent, then adding water and an inorganic base to carry out a condensation reaction to obtain the compound of formula I-2.

6. The synthesis method according to claim 1, characterized in that, It includes a method for synthesizing a compound of formula I-2-1, the method comprising the following steps: In an organic solvent and water, in the presence of an activator, a condensing agent, a tertiary amine organic base and an inorganic base, the compound of formula I-1 undergoes a condensation reaction with threonine methyl ester to give the compound of formula I-2-1. ； Preferably, the method for synthesizing the compound of formula I-2-1 satisfies one or more of the following conditions: (1) The organic solvent is an ether solvent and / or a halogenated hydrocarbon solvent, more preferably an ether solvent; The halocarbon solvent may be a chloromethane solvent, such as dichloromethane; The ether solvent may be a cyclic ether solvent, preferably a five-membered cyclic ether solvent and / or a six-membered cyclic ether solvent, such as tetrahydrofuran and / or 1,4-dioxane; (2) The molar volume ratio of the compound of formula I-1 to the organic solvent is (0.1-1.5) mol / L, preferably (0.4-0.9) mol / L, for example 0.8 mol / L, 0.5 mol / L or 0.7 mol / L; (3) The mass ratio of the organic solvent to the water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1; (4) The molar ratio of the compound of formula I-1 to the threonine methyl ester is 1:(0.5-2.0), preferably 1:(0.8-1.2), for example 1:1; (5) The organic tertiary amine base is an aliphatic tertiary amine base, preferably a non-nucleophilic aliphatic tertiary amine base, such as N,N-diisopropylethylamine; (6) The molar ratio of the compound of formula I-1 to the organic tertiary amine base is 1:(0.1-1.0), preferably 1:(0.3-0.5), for example 1:0.4; (7) The activator is a triazole activator, preferably a hydroxybenzotriazole activator, and more preferably 1-hydroxy-7-azabenzotriazole or 1-hydroxybenzotriazole, for example 1-hydroxybenzotriazole; (8) The molar ratio of the compound of formula I-1 to the activator is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (9) The condensing agent is a carbodiamine condensing agent, such as N,N'-diisopropylcarbodiimide; (10) The molar ratio of the compound of formula I-1 to the condensing agent is 1:(0.5-2.0), preferably 1:(1.0-1.5), for example 1:1.5, 1:1.3 or 1:1.1; (11) The inorganic base is a metal hydroxide base, preferably selected from one or more of sodium hydroxide, potassium hydroxide, calcium hydroxide and lithium hydroxide, and more preferably lithium hydroxide, such as lithium hydroxide monohydrate; (12) The molar ratio of the compound of formula I-1 to the inorganic base is 1:(1-3), preferably 1:(1.5-2.5), for example 1:1.9; (13) The molar volume ratio of the compound of formula I-1 to the water is (0.1-1.5) mol / L, preferably (0.4-1.0) mol / L, for example 0.9 mol / L, 0.5 mol / L or 0.7 mol / L; (14) The reaction temperature of the condensation reaction is 10-60℃, preferably 20-50℃; (15) The reaction time of the condensation reaction is 2-10h, more preferably 3-7h, for example 6h; (16) The reaction raw materials for the condensation reaction consist of the following substances: the organic solvent, the water, the activator, the condensing agent, the tertiary amine organic base, the inorganic base, the compound of formula I-1, and the threonine methyl ester; (17) The method for synthesizing the compound of formula I-2-1 includes the following steps: adding an organic tertiary amine base to a mixture of an organic solvent, the compound of formula I-1, serine methyl ester, an activator and a condensing agent, then adding water and an inorganic base to carry out a condensation reaction to obtain the compound of formula I-2-1.

7. A method for synthesizing a compound of formula I, comprising the following steps: (1) Compounds of formula I-5 are prepared by the synthetic method according to any one of claims 1-6; (2) In a mixed solvent of ether solvent and water, in the presence of a catalyst and a hydrogen source, compounds of formula I-5 undergo a reduction reaction to obtain compound I; ； Preferably, it is characterized by satisfying one or more of the following conditions: (1) The ether solvent is a cyclic ether solvent, preferably a five-membered ring ether solvent and / or a six-membered ring ether solvent, for example, 1,4-dioxane and / or tetrahydrofuran; (2) In the mixed solvent of ether solvent and water, the mass ratio of ether solvent to water is 1:(0.5-1.5), preferably 1:(0.8-1.2), for example 1:1; (3) The molar volume ratio of the compound of formula I-5 to the ether solvent is (0.1-1.0) mol / L, preferably (0.3-0.6) mol / L, for example 0.5 mol / L, 0.35 mol / L or 0.54 mol / L; (4) The catalyst is a metal catalyst, preferably a palladium catalyst, and more preferably a zero-valent palladium catalyst, for example, a 10%wt palladium on carbon catalyst; (5) The molar ratio of the compound of formula I-5 to the catalyst is 1:(0.3-2.0), preferably 1:(0.4-1.0), for example 1:0.77, 1:0.58 or 1:0.4; (6) The hydrogen source is hydrogen gas; preferably, the pressure of the hydrogen gas is 0.3-1.5 MPa, more preferably 0.5-1.0 MPa, for example 0.6-0.7 MPa; (7) The reaction temperature of the reduction reaction is 20-60℃, preferably 30-50℃, for example 40-50℃; (8) The reaction time of the reduction reaction is 1-8 hours, preferably 2-5 hours, for example 3 hours; (9) The reaction raw materials for the reduction reaction consist of the following substances: the mixed solvent of the ether solvent and water, the catalyst, the compound of formula I-5, and the hydrogen source; (10) The method for synthesizing the compound of formula I further includes crystallization, wherein the crystallization solution is a nitrile solvent or a ketone solvent; the nitrile solvent may be acetonitrile; the ketone solvent may be acetone.

8. A method for synthesizing a compound of formula I-4, comprising the following steps: In an organic solvent, in the presence of an activator, a condensing agent and a tertiary amine organic base, compounds I-2 and I-3 undergo a condensation reaction to obtain compound I-4. ； Preferably, the reaction conditions for synthesizing the compound of formula I-4 are as described in claim 3.

9. A compound of formula I-4: 。 10. A compound of formula I-5: 。

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