Method for producing peptide compounds

Abstract

The present invention addresses the problem of providing a method for efficiently producing a peptide and provides a method for producing a peptide, said method comprising: step (1) for mixing an N-protected amino acid or an N-protected peptide with a carboxylic acid halide represented by formula (I) (wherein: X represents a halogen atom; and R1, R2 and R3 independently represent an optionally substituted aliphatic hydrocarbon group, provided that the total number of carbon atoms in R1, R2 and R3 is 3 to 40); and step (2) for mixing the product obtained in step (1) with a C-protected amino acid or a C-protected peptide.

Description

[Technical field]

[0001] The present invention relates to a method for producing a peptide compound. [Background technology]

[0002] Condensation reagents for forming amide bonds are widely used in the production of biologically active substances such as medicines and agricultural chemicals. In particular, in the production of peptide compounds, it is necessary to form multiple amide bonds, and therefore there is a strong demand for condensation reagents that can give the target product in high yield (Non-Patent Document 1). In the production of peptide compounds, (1-cyano-2-ethoxy-2-oxoethylideneaminooxy)dimethylamino-morpholino-carbenium hexafluorophosphate (COMU), isobutyl chloroformate, pivaloyl chloride, and the like are known as condensation reagents that can be industrially used (Patent Documents 1 and 2, Non-Patent Documents 2 and 3). On the other hand, COMU and isobutyl chloroformate are known to be exothermic due to autolysis (Non-Patent Document 4). [Prior art documents] [Patent documents]

[0003] [Patent Document 1] International Publication No. 2017 / 129796 [Patent Document 2] International Publication No. 2012 / 004554

[0004] [Non-Patent Document 1] Chemical Reviews 2011, Vol. 111, pp. 6557-6602 [Non-Patent Document 2] Reports of the Bulgarian Academy of Sciences 2004, volume 57, pages 53-58. [Non-Patent Document 3] Russian Journal of Bioorganic Chemistry 2009, Vol. 35, pp. 150-156 [Non-Patent Document 4] Organic Process Research and Development 2018, Vol. 22, pp. 1262-1275 Summary of the Invention [Problem to be solved by the invention]

[0005] The present inventors have confirmed that when pivaloyl chloride is used as a condensation reagent to react the N-terminus of a C-protected amino acid or a C-protected peptide with the C-terminus of an N-protected amino acid or an N-protected peptide, a by-product is generated in which a pivaloyl group derived from the condensation reagent is introduced into the N-terminus of the C-protected amino acid or the C-protected peptide, and the yield of the target peptide compound decreases. In the production of a peptide compound that repeatedly forms amide bonds, the by-product accumulates as an impurity. Therefore, the above method is expected to have problems in terms of efficiency and economy as an industrial production method.

[0006] The present invention provides a method for producing a peptide compound in high yield using a safe condensation reagent that does not generate heat due to autolysis. [Means for solving the problem]

[0007] As a result of extensive investigations, the present inventors have found that the above-mentioned problems can be solved by using a carboxylic acid halide having a specific structure, and have thus completed the present invention. That is, the present invention is characterized as follows. [1] The following steps (1) and (2): (1) and an N-protected amino acid or an N-protected peptide of formula (I) [ka] (wherein X represents a halogen atom, R 1 , R 2 and R 3 each independently represents an aliphatic hydrocarbon group which may have a substituent; R 1 , R 2and R 3 wherein the total number of carbon atoms is 3 to 40; and (2) mixing the product obtained in step (1) with a C-protected amino acid or a C-protected peptide; A method for producing a peptide, comprising: [2] Further, the following steps (3) to (5): (3) a step of removing the protecting group at the N-terminus of the peptide obtained in step (2) or (5); (4) mixing an N-protected amino acid or an N-protected peptide with a carboxylic acid halide of formula (I); and (5) mixing the product obtained in step (4) with the product obtained in step (3); A method for producing the peptide according to [1], which contains one or more repeats of the above. [3] R 1 , R 2 and R 3 The method for producing a peptide according to [1] or [2], wherein two or more of the above are independently an aliphatic hydrocarbon group containing one or more tertiary or quaternary carbon atoms. [4] R 1 is a methyl group, and R 2 and R 3 are each independently an aliphatic hydrocarbon group containing one or more tertiary or quaternary carbon atoms. [5] R 2 and R 3 each independently represents one or more tertiary or quaternary carbon atoms; 3-10 The method for producing the peptide according to [4], wherein the peptide is an alkyl group. [6] The carboxylic acid halide is represented by the formula (II) [ka] (wherein X represents a halogen atom). [7] The method for producing a peptide according to any one of [1] to [6], wherein X is a chlorine atom or a bromine atom. [8] The method for producing the peptide according to any one of [1] to [6], wherein X is a chlorine atom. [9] The method for producing a peptide according to any one of [1] to [8], wherein the N-protected amino acid is an α-amino acid other than glycine.

[10] The method for producing a peptide according to [9], wherein the amino acid in the N-protected amino acid is an α-amino acid other than glycine, and the reactive functional group of the amino acid side chain is protected.

[11] The method for producing a peptide according to [9] or

[10] , wherein the α-amino acid other than glycine is valine, phenylalanine, threonine, leucine, tryptophan, serine, cysteine, aspartic acid or tyrosine.

[12] The method for producing a peptide according to any one of [1] to

[11] , wherein the amino acid in the C-protected amino acid or the amino acid at the N-terminal residue of the C-protected peptide is an α-amino acid other than an N-substituted amino acid.

[13] The method for producing a peptide according to any one of [1] to

[12] , wherein step (2) is a step of mixing the product obtained in step (1) with a C-protected peptide.

[14] The method for producing a peptide according to any one of [1] to

[13] , wherein the N-terminal protecting group of the N-protected amino acid or the N-protected peptide is a carbamate protecting group.

[15] The method for producing a peptide according to

[14] , wherein the carbamate protecting group is a 9-fluorenylmethyloxycarbonyl group or a benzyloxycarbonyl group. Effect of the Invention

[0008] According to the present invention, it is possible to obtain the target peptide in high yield using industrially applicable reagents. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0009] The present invention will be described in detail below. In this specification, "n-" means normal, "s-" means secondary, "t-" and "tert-" mean tertiary, "Me" means methyl, "Et" means ethyl, "Bu" means butyl, "Bn" means benzyl, "Boc" means t-butoxycarbonyl, "Cbz" means benzyloxycarbonyl, "Fmoc" means 9-fluorenylmethoxycarbonyl, "Trt" means trityl, and "NMP" means N-methylpyrrolidone.

[0010] The term "halogen atom" means a fluorine atom, chlorine atom, bromine atom or iodine atom.

[0011] "Alkyl group" means a linear or branched, saturated aliphatic hydrocarbon monovalent group. 1-6 The term "alkyl group" means a straight-chain or branched-chain alkyl group having 1 to 6 carbon atoms, and specific examples include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, a s-butyl group, a t-butyl group, an n-pentyl group, a 3-pentyl group, a 2-methylbutyl group, a 3-methylbutyl group, a 1,1-dimethylpropyl group, a 1,2-dimethylpropyl group, a 2,2-dimethylpropyl group, a 1-ethylpropyl group, an n-hexyl group, and a 3,3-dimethylbutan-2-yl group.

[0012] "C 1-40The term "alkyl group" means a straight-chain or branched-chain alkyl group having 1 to 40 carbon atoms, and specific examples include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, a s-butyl group, a t-butyl group, an n-pentyl group, an n-hexyl group, an octyl group, a decyl group, a dodecyl group, a hexadecyl group, an octadecyl group, a docosyl group, a triacontyl group, a tetracontyl group, and a 3,7,11,15-tetramethylhexadecyl group (hereinafter sometimes referred to as a 2,3-dihydrophytyl group).

[0013] "C 3-10 The term "alkyl group" means a straight-chain or branched-chain alkyl group having 3 to 10 carbon atoms, and specific examples include an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, a s-butyl group, a t-butyl group, an n-pentyl group, an n-hexyl group, an octyl group, a decyl group, a 2,3-dimethylbutyl group, a 2,2-dimethylbutyl group, a 2,2,4-trimethylpentyl group, a 2,2,4-trimethylhexyl group, a 2,2,3,4-tetramethylhexyl group, and a 4-ethyl-2,2-dimethylhexyl group.

[0014] "Cycloalkyl group" means a monovalent group of a cyclic saturated aliphatic hydrocarbon. 3-6 The term "cycloalkyl group" means a cycloalkyl group having 3 to 6 carbon atoms, and specific examples include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, and a cyclohexyl group.

[0015] The term "alkenyl group" refers to a monovalent linear or branched unsaturated aliphatic hydrocarbon group having a double bond. 2-6 The term "alkenyl group" means an alkenyl group having 2 to 6 carbon atoms, and specific examples include a vinyl group, a 1-propenyl group, an allyl group, an isopropenyl group, a 2-butenyl group, and a 3-butenyl group.

[0016] The term "alkynyl group" refers to a monovalent linear or branched unsaturated aliphatic hydrocarbon group having a triple bond. 2-6The term "alkynyl group" means an alkynyl group having 2 to 6 carbon atoms, and specific examples include an ethynyl group, a 1-propynyl group, and the like.

[0017] The term "aralkyl group" refers to an alkyl group having an aromatic hydrocarbon as a substituent. 7-14 The term "aralkyl group" means an aralkyl group having 7 to 14 carbon atoms, and specific examples include a benzyl group, a 1-phenylethyl group, a 2-phenylethyl group, a 1-phenylpropyl group, a naphthylmethyl group, a 1-naphthylethyl group, and a 1-naphthylpropyl group.

[0018] "C 6-14 The term "aryl group" means an aromatic hydrocarbon group having 6 to 14 carbon atoms, and specific examples thereof include a phenyl group, a 1-naphthyl group, a 2-naphthyl group, a 1-anthryl group, a 2-anthryl group, a 9-anthryl group, and a biphenyl group.

[0019] "C 6-14 The term "haloaryl group" means an aromatic hydrocarbon group having 6 to 14 carbon atoms substituted with one or more halogen atoms, and specific examples thereof include a 4-chlorophenyl group, a 2,4-dichlorophenyl group, a 5-fluoro-1-naphthyl group, a 6-bromo-2-naphthyl group, a 6,7-diiodo-1-anthryl group, a 10-bromo-9-anthryl group, and a 4'-chloro-(1,1'-biphenyl)-2-yl group.

[0020] "C 6-14 The term "aryloxy group" means an aryloxy group having 6 to 14 carbon atoms, and specific examples include a phenoxy group, a 1-naphthyloxy group, a 2-naphthyloxy group, a 1-anthryloxy group, a 2-anthryoxy group, a 9-anthryoxy group, and a biphenyloxy group.

[0021] The term "5- to 10-membered heterocyclic group" refers to a monocyclic or condensed ring heterocyclic group having 5 to 10 ring-constituting atoms and containing, among the ring-constituting atoms, 1 to 4 heteroatoms independently selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms. This heterocyclic group may be saturated, partially unsaturated, or unsaturated. Specific examples include a pyrrolidinyl group, a tetrahydrofuryl group, a tetrahydrothienyl group, a piperidyl group, a tetrahydropyranyl group, a tetrahydrothiopyranyl group, a pyrrolyl group, a furyl group, a thienyl group, a pyridyl group, a pyrimidinyl group, a pyridazinyl group, an azepanyl group, an oxepanyl group, a thiepanyl group, an azepinyl group, an oxepinyl group, a thiepinyl group, an imidazolyl group, a pyrazolyl group, an oxazolyl group, a thiazolyl group, an imidazolinyl group, a pyrazinyl group, a morpholinyl group, a thiazinyl group, an indolyl group, an isoindolyl group, a benzimidazolyl group, a purinyl group, a quinolyl group, an isoquinolyl group, a quinoxalinyl group, a cinnolinyl group, a pteridinyl group, a chromenyl group, and an isochromenyl group.

[0022] "C 1-6 The term "alkoxy group" means a straight-chain or branched-chain alkoxy group having 1 to 6 carbon atoms, and specific examples include a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an isobutoxy group, a t-butoxy group, an n-pentyloxy group, and an n-hexyloxy group.

[0023] "C 1-40 The term "alkoxy group" means a straight-chain or branched-chain alkoxy group having 1 to 40 carbon atoms, and specific examples include a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an isobutoxy group, a t-butoxy group, an n-pentyloxy group, an n-hexyloxy group, an octyloxy group, a decyloxy group, a dodecyloxy group, a hexadecyloxy group, an octadecyloxy group, a docosyloxy group, a triacontyloxy group, a tetracontyloxy group, and a 3,7,11,15-tetramethylhexadecyloxy group (hereinafter sometimes referred to as a 2,3-dihydrophytyloxy group).

[0024] "C 3-6 The term "cycloalkoxy group" means a cycloalkyloxy group having 3 to 6 carbon atoms, and specific examples include a cyclopropoxy group, a cyclobutoxy group, a cyclopentyloxy group, and a cyclohexyloxy group.

[0025] "Mono C 1-6 The term "alkylamino group" refers to one of the above "C 1-6 "Alkyl group" means a group in which an "alkyl group" is bonded to an amino group, and specific examples include a monomethylamino group, a monoethylamino group, a mono-n-propylamino group, a monoisopropylamino group, a mono-n-butylamino group, a monoisobutylamino group, a mono-t-butylamino group, a mono-n-pentylamino group, and a mono-n-hexylamino group.

[0026] "JiC 1-6 The term "alkylamino group" refers to a group having two identical or different "C 1-6 Specific examples of the alkyl group include dimethylamino, diethylamino, di-n-propylamino, diisopropylamino, di-n-butylamino, diisobutylamino, di-t-butylamino, di-n-pentylamino, di-n-hexylamino, N-ethyl-N-methylamino, N-methyl-Nn-propylamino, N-isopropyl-N-methylamino, Nn-butyl-N-methylamino, Examples of such amino groups include N-isobutyl-N-methylamino group, Nt-butyl-N-methylamino group, N-methyl-Nn-pentylamino group, Nn-hexyl-N-methylamino group, N-ethyl-Nn-propylamino group, N-ethyl-N-isopropylamino group, Nn-butyl-N-ethylamino group, N-ethyl-N-isobutylamino group, Nt-butyl-N-ethylamino group, N-ethyl-Nn-pentylamino group, and N-ethyl-Nn-hexylamino group.

[0027] "C 1-6The term "alkoxycarbonyl group" means a straight-chain or branched-chain alkoxycarbonyl group having 1 to 6 carbon atoms, and specific examples include a methoxycarbonyl group, an ethoxycarbonyl group, an n-propoxycarbonyl group, an isopropoxycarbonyl group, an n-butoxycarbonyl group, an isobutoxycarbonyl group, a t-butoxycarbonyl group, an n-pentyloxycarbonyl group, and an n-hexyloxycarbonyl group.

[0028] "Tori C 1-6 The term "alkylsilyl group" refers to a group having three identical or different "C 1-6 "Alkyl group" means a group in which the alkyl group is bonded to a silyl group, and specific examples include a trimethylsilyl (TMS) group, a triethylsilyl group, a triisopropylsilyl group, a t-butyldimethylsilyl group, and a di-t-butylisobutylsilyl group.

[0029] "Tori C 1-6 The term "alkylsilyloxy group" refers to a group having three identical or different "C 1-6 Specific examples of the silyloxy group include a trimethylsilyloxy group, a triethylsilyloxy group, a triisopropylsilyloxy group, a t-butyldimethylsilyloxy group, and a di-t-butylisobutylsilyloxy group.

[0030] The term "bicycloalkyl group" refers to a monovalent group of a saturated aliphatic hydrocarbon containing two bridgehead carbons and having two rings. Specific examples include an octahydroinden-3-yl group, an octahydronaphthalene-4-yl group, a bicyclo[2.2.1]heptan-1-yl group, and a bicyclo[2.2.1]heptan-2-yl group. 5-10 A bicycloalkyl group has 5 to 10 carbon atoms and is defined as "C 5-7 The term "bicycloalkyl group" refers to a bicycloalkyl group having from 5 to 7 carbon atoms.

[0031] The term "tricycloalkyl group" refers to a monovalent group of a saturated aliphatic hydrocarbon having at least three bridgehead carbons and three rings. Specific examples include tricyclo[3.3.1.1 3,7 ]decan-1-yl (adamantan-1-yl) group or tricyclo[3.3.1.1 3,7 ]decan-2-yl (adamantan-2-yl) group. 5-15 A "tricycloalkyl group" is a group having 5 to 15 carbon atoms. 7-15 The term "tricycloalkyl group" refers to a tricycloalkyl group having from 7 to 15 carbon atoms.

[0032] The term "aliphatic hydrocarbon group" refers to a linear, branched or cyclic, saturated or unsaturated aliphatic hydrocarbon group, and examples thereof include an alkyl group, a cycloalkyl group, an alkenyl group, an alkynyl group, and an aralkyl group. Specific examples thereof include C 1-40 Alkyl group, C 3-6 Cycloalkyl groups, C 2-6 Alkenyl group, C 2-6 Alkynyl group, C 7-14 Examples include aralkyl groups.

[0033] The term "aliphatic hydrocarbon group containing one or more tertiary or quaternary carbon atoms" refers to a group in which one or more of the carbon atoms constituting the aliphatic hydrocarbon group are tertiary or quaternary carbon atoms. Specific examples include a 2-methylbutan-2-yl group, a 3-methylbutan-2-yl group, a 3,3-dimethylbutan-2-yl group, a t-butyl group, a 3-pentyl group, a 2,2,4-trimethylpentan-3-yl group, a 2,4,4-trimethylpentyl group, a 2,4-dimethylpentan-3-yl group, a 4-ethyl-2,2-dimethylhexane-3-yl group, a 3-heptyl group, a 2,2,4,8,10,10-hexamethylundecane-5-yl group, a 3-methylcyclobutyl group, a 2-methylcyclopentyl group, and a 4-isopropylcyclohexyl group.

[0034] "C containing one or more tertiary or quaternary carbon atoms 3-10 "Alkyl group" means C 3-10It means a group in which one or more of the carbon atoms constituting the alkyl group are tertiary or quaternary carbon atoms, and specific examples include an isobutyl group, a t-butyl group, an isopentyl group, a neopentyl group, a 2,2-dimethylbutyl group, a 2,3-dimethylbutyl group, a 2,2,4-trimethylpentyl group, a 2,4,4-trimethylpentyl group, a 2,2,4-trimethylhexyl group, a 2,2,3,4-tetramethylhexyl group, a 2,2,4,4-tetramethylpentyl group, and a 4-ethyl-2,2-dimethylhexyl group.

[0035] "Reactive functional group of amino acid side chain" means a group present in the amino acid side chain that can react with other groups to form a covalent bond, specific examples of which include hydroxyl group, carboxyl group, amino group, amide group, thiol group, etc. Amino acids containing these functional groups in their side chains include arginine, asparagine, aspartic acid, lysine, glutamine, cysteine, serine, threonine, tyrosine, tryptophan, histidine, etc. Furthermore, "the reactive functional group of the amino acid side chain is protected" means that a protecting group that is generally used in peptide chemistry and the like is introduced into the reactive functional group.

[0036] The term "optionally substituted" means that the group is unsubstituted or substituted with any number of substituents that are chemically permissible.

[0037] The term "having a substituent" means substituted with any number of optional substituents which are chemically permissible.

[0038] The above-mentioned "optional substituent" is not particularly limited in type, so long as it is a substituent that does not adversely affect the reaction that is the subject of the present invention.

[0039] Examples of the "substituent" in the "optionally substituted aliphatic hydrocarbon group" include C 6-14 Aryl group, C 6-14 Aryloxy group, 5-10 membered heterocyclic group, hydroxy group, C 1-40 Alkoxy group, C 3-6Cycloalkoxy group, acetoxy group, benzoyloxy group, amino group, mono C 1-6 Alkylamino group, N-acetylamino group, diC 1-6 Alkylamino group, halogen atom, C 1-6 Alkoxycarbonyl group, phenoxycarbonyl group, N-methylcarbamoyl group, N-phenylcarbamoyl group, triC 1-6 Alkylsilyl group, triC 1-6 Examples of the alkyl group include an alkylsilyloxy group, a cyano group, a nitro group, and a carboxy group.

[0040] The terms "N-protected amino acid" and "N-protected peptide" refer to an amino acid or peptide in which the amino group present in the amino acid backbone or the amino group at the N-terminus of the peptide is protected and the carboxy group present in the amino acid backbone or the carboxy group at the C-terminus of the peptide is unprotected.

[0041] The terms "C-protected amino acid" and "C-protected peptide" refer to an amino acid or peptide in which the carboxy group present in the amino acid backbone or the carboxy group at the C-terminus of the peptide is protected and the amino group present in the amino acid backbone or the amino group at the N-terminus of the peptide is unprotected.

[0042] The amino acids used in the present invention also include N-substituted amino acids.

[0043] The term "N-substituted amino acid" refers to an amino acid in which one substituent has been introduced into the amino group present in the amino acid backbone. A specific example is NC 1-6 Alkylamino acids, NC 2-6 Alkenyl amino acid, NC 2-6 Alkynyl amino acids, NC 6-14 Aryl amino acid, NC 1-6 Alkoxyamino acid (C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 6-14 Aryl and C 1-6The alkoxy may have a substituent, and examples of such substituents are the same as the examples of the "substituent" in the above "aliphatic hydrocarbon group which may have a substituent".

[0044] Above NC 1-6 The alkylamino acid is preferably C 6-14 Aryl group, C 1-6 Alkoxy group, C 1-6 Alkoxycarbonyl group, diC 1-6 Alkylamino group, 5-10 membered heterocyclic group or triC 1-6 C which may have an alkylsilyl group 1-6 An amino acid having one alkyl group introduced therein, preferably C 6-14 Aryl group, C 1-6 Alkoxy group or C 1-6 C which may have an alkoxycarbonyl group 1-6 It is an amino acid having one alkyl group introduced, and more preferably an amino acid having one methyl group, ethyl group, n-propyl group, n-butyl group or benzyl group introduced.

[0045] The term "group derived from an amino acid" as used herein means a divalent group in which a hydrogen atom has been removed from the nitrogen atom of a primary or secondary amino group present in the main chain of an amino acid and a hydroxyl group has been removed from a carboxy group present in the main chain.

[0046] The term "peptide-derived group" as used herein means a divalent group in which a hydrogen atom has been removed from the nitrogen atom of the primary or secondary amino group of the amino acid constituting the N-terminus and a hydroxyl group has been removed from the carboxy group of the amino acid constituting the C-terminus.

[0047] The stereostructure of the α-amino acid is not particularly limited, but is preferably the L-form.

[0048] All technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are described below. All publications and patents mentioned herein are incorporated by reference for the purpose of describing and disclosing, for example, the constructs and methodologies described in the publications that can be used in connection with the described invention.

[0049] (Specific Description of the Method for Producing the Peptide of the Present Invention) The method for producing a peptide of the present invention can be carried out by performing all or a suitable combination of the unit steps described below as steps (1) to (5). This specific description will be given based on the following. (a) R in the description of steps (1) to (5) 1 , R 2 and R 3 is synonymous with the above. (b) The specific conditions for the reactions are not particularly limited as long as the production of the peptide of the present invention is achieved. Preferred conditions for each reaction will be described in detail as appropriate. (c) Each reaction can be carried out in a solvent as necessary, and is preferably carried out in a solvent. The solvents described in each reaction can be used alone or in combination of two or more kinds.

[0050] Process (1) In this step, an N-protected amino acid or an N-protected peptide is mixed with a carboxylic acid halide. In this step, the C-terminus of the N-protected amino acid or the N-protected peptide is activated with a carboxylic acid halide. In one embodiment of the present invention, a compound represented by formula (III)P 1 -A 1 -OH(in the formula, P 1 is an N-terminal protecting group, and A 1 represents a group derived from an amino acid or a group derived from a peptide.

[0051] The carboxylic acid halide is represented by the following formula (I).

[0052] [ka] (In the formula, X represents a halogen atom, R 1 , R 2 and R 3 each independently represents an aliphatic hydrocarbon group which may have a substituent; R 1 , R 2 and R 3 The total number of carbon atoms in the

[0053] The carboxylic acid halide of formula (I) is preferably 1 , R 2 and R 3 Among these, two or more are independently an aliphatic hydrocarbon group containing one or more tertiary or quaternary carbon atoms, and more preferably, R 1 is a methyl group, and R 2 and R 3 are each independently an aliphatic hydrocarbon group containing one or more tertiary or quaternary carbon atoms, and more preferably 1 is a methyl group, and R 2 and R 3 each independently represents one or more tertiary or quaternary carbon atoms; 3-10 The alkyl group is a carboxylic acid halide, and the compound shown in the following formula (II) is particularly preferred. [ka]

[0054] The halogen atom represented by X in formula (I) and formula (II) is not particularly limited as long as it is a halogen atom, but is preferably a chlorine atom or a bromine atom, and more preferably a chlorine atom.

[0055] The amount of the carboxylic acid halide used is preferably 0.2 to 50 equivalents, more preferably 0.5 to 20 equivalents, and even more preferably 0.8 to 5 equivalents, relative to the N-protected amino acid or N-protected peptide.

[0056] P in formula (III) 1 The N-terminal protecting group represented by the formula (I) is not particularly limited, and specific examples thereof include carbamate protecting groups (9-fluorenylmethoxycarbonyl group, t-butoxycarbonyl group, benzyloxycarbonyl group, allyloxycarbonyl group, 2,2,2-trichloroethoxycarbonyl group, 2-(p-biphenyl)isopropyloxycarbonyl group, etc.), amide protecting groups (acetyl group, trifluoroacetyl group, etc.), imide protecting groups (phthaloyl group, etc.), sulfonamide protecting groups (p-toluenesulfonyl group, 2-nitrobenzenesulfonyl group, etc.), benzyl group, etc., preferably 9-fluorenylmethoxycarbonyl group, t-butoxycarbonyl group, or benzyloxycarbonyl group, more preferably 9-fluorenylmethoxycarbonyl group, or benzyloxycarbonyl group.

[0057] The amino acid in the N-protected amino acid represented by formula (III) and the amino acid in the C-terminal residue of the N-protected peptide are not particularly limited, but are preferably α-amino acids, β-amino acids, γ-amino acids or δ-amino acids, more preferably α-amino acids or β-amino acids, even more preferably α-amino acids, still more preferably α-amino acids other than glycine, and particularly preferably valine, phenylalanine, threonine, leucine, tryptophan, serine, cysteine, aspartic acid or tyrosine. If these amino acids have two or more amino groups (e.g., arginine, lysine, etc.), two or more carboxy groups (e.g., glutamic acid, aspartic acid, etc.), or reactive functional groups (e.g., cysteine, serine, etc.), the reactive functional groups present in the side chains of amino acids not involved in the formation of the peptide may be protected.

[0058] In formula (III), A1 When is a group derived from a peptide, the number of amino acid residues contained in the group is not particularly limited, but is preferably 2 to 40, and more preferably 2 to 20.

[0059] This step can be carried out by adding a base, if necessary.

[0060] The base used in this step is not particularly limited, and examples thereof include aliphatic amines (e.g., triethylamine, N,N-diisopropylethylamine, N-methylmorpholine), aromatic amines (e.g., pyridine, imidazole, N,N-dimethyl-4-aminopyridine), amidines (e.g., diazabicycloundecene), alkali metal salts (e.g., sodium hydrogen carbonate, potassium carbonate), etc. Preferred are aliphatic amines, and more preferred are N,N-diisopropylethylamine, triethylamine, or N-methylmorpholine.

[0061] The amount of the base used in this step is preferably 0.2 to 50 equivalents, more preferably 0.5 to 20 equivalents, and further preferably 0.8 to 5 equivalents, relative to the carboxylic acid halide.

[0062] The solvent used in this step is not particularly limited as long as it does not interfere with the activation reaction, and examples thereof include halogen-containing hydrocarbon solvents (e.g., dichloromethane, chloroform), aromatic hydrocarbon solvents (e.g., toluene, xylene), ether solvents (e.g., tetrahydrofuran, 1,4-dioxane, cyclopentyl methyl ether, methyl-t-butyl ether), amide solvents (e.g., N,N-dimethylformamide, N,N-dimethylacetamide), nitrile solvents (e.g., acetonitrile), ketone solvents (e.g., acetone, methyl ethyl ketone), aliphatic hydrocarbon solvents (e.g., hexane, heptane, cyclohexane), ester solvents (e.g., ethyl acetate), etc. Preferred are nitrile solvents, amide solvents, or ether solvents, and more preferred are acetonitrile, tetrahydrofuran, or N,N-dimethylacetamide.

[0063] The amount of the solvent used is preferably 100 times by mass or less, more preferably 1 to 50 times by mass, and further preferably 3 to 20 times by mass, relative to the amount of the carboxylic acid halide.

[0064] In this step, the temperature may be controlled using an oil bath or a cooling bath as necessary. The temperature is not particularly limited, but is preferably from -40°C to the reflux temperature of the mixture, more preferably from -20°C to 50°C, and even more preferably from -10°C to 30°C.

[0065] This step forms an N-protected amino acid or an N-protected peptide with an activated C-terminus. Therefore, the product obtained by this step means an N-protected amino acid or an N-protected peptide with an activated C-terminus, or a mixture containing either of them. The N-protected amino acid or the N-protected peptide with an activated C-terminus thus obtained may be mixed with the C-protected peptide as it is in the reaction solution or isolated as a (crude) purified product without going through a purification step.

[0066] Process (2) This step is a step of mixing the product obtained in step (1) with a C-protected amino acid or a C-protected peptide. This step is a step of mixing the N-protected amino acid or the N-protected peptide whose C-terminus is activated obtained in step (1) with a C-protected amino acid or a C-protected peptide. In one embodiment of the present invention, the N-protected amino acid or the N-protected peptide whose C-terminus is activated obtained in step (1) with a compound represented by formula (IV) HA 2 -OP 2 (A 2 represents a group derived from an amino acid or a group derived from a peptide, P 2 is a C-terminal protecting group.

[0067] The amino acid in the C-protected amino acid represented by formula (IV) and the amino acid in the N-terminal residue of the C-protected peptide are not particularly limited, but are preferably α-amino acids, β-amino acids, γ-amino acids, or δ-amino acids, more preferably α-amino acids or β-amino acids, even more preferably α-amino acids, and even more preferably phenylalanine, glycine, valine, proline, leucine, or ornithine. If these amino acids have two or more amino groups (e.g., arginine, lysine, etc.), two or more carboxy groups (e.g., glutamic acid, aspartic acid, etc.), or reactive functional groups (e.g., cysteine, serine, etc.), the reactive functional groups of the amino acid side chains that are not involved in the formation of the peptide may be protected.

[0068] In formula (IV), A 2 When is a group derived from a peptide, the number of amino acid residues contained in the group is not particularly limited, but is preferably 2 to 40, and more preferably 2 to 20.

[0069] P in formula (IV) 2 The C-terminal protecting group represented by the formula (I) is not particularly limited as long as it is a protecting group commonly used in the synthesis of amino acids or peptides, and examples thereof include a methyl group, an ethyl group, a t-butyl group, a benzyl group, an allyl group, and a silyl group.

[0070] In formula (IV), P 2may be a group derived from a solid phase carrier, and the embodiment is not particularly limited, and is bound directly or via a linker. The solid phase carrier is not particularly limited, and examples thereof include nitrocellulose, agarose beads, modified cellulose fibers, polypropylene, polyethylene glycol, polystyrene, polyacrylamide, and other synthetic resins. The linker is not particularly limited, and examples thereof include 2-chlorotrityl (2-ClTrt), 4-(hydroxymethyl)benzoic acid, 3,4-dihydro-2H-pyran-2-ylmethanol, 4-(hydroxymethyl)phenoxyacetic acid, 3-hydroxy-xanthen-9-one, N-methoxy-3-aminopropionic acid, 3-methoxy-2-nitropyridine, and the like.

[0071] The solvent used in this step is not particularly limited, but examples thereof include halogen-containing hydrocarbon solvents (e.g., dichloromethane, chloroform), aromatic hydrocarbon solvents (e.g., toluene, xylene), ether solvents (e.g., tetrahydrofuran, 1,4-dioxane, cyclopentyl methyl ether, methyl-t-butyl ether), amide solvents (e.g., N,N-dimethylformamide), nitrile solvents (e.g., acetonitrile), etc. Nitrile solvents, amide solvents, or ether solvents are preferred, and acetonitrile, tetrahydrofuran, or N,N-dimethylacetamide are more preferred.

[0072] The amount of the solvent used is preferably 100 times or less by mass, more preferably 1 to 50 times by mass, and even more preferably 3 to 20 times by mass, relative to the amount of the C-protected amino acid or C-protected peptide.

[0073] The temperature of the obtained mixture is controlled using an oil bath or a cooling bath as necessary. The temperature of the mixture is not particularly limited, but is preferably from -40°C to the reflux temperature of the reaction mixture, more preferably from -20°C to 50°C, and further preferably from -10°C to 30°C.

[0074] When the C-protected amino acid or C-protected peptide used in this step is in the form of a salt, it can be converted into the free form by adding an organic amine.

[0075] The organic amine used when converting the salt of the C-protected amino acid or C-protected peptide into the free form is not particularly limited, and examples thereof include aliphatic amines (e.g., dicyclohexylamine, piperidine, triethylamine, N,N-diisopropylethylamine, N-methylmorpholine), aromatic amines (e.g., pyridine, imidazole, N,N-dimethyl-4-aminopyridine), etc. Preferred are aliphatic amines, and more preferred are triethylamine or N,N-diisopropylethylamine.

[0076] The amount of the organic amine used in converting a salt of a C-protected amino acid or a C-protected peptide into a free form is preferably 0.01 to 50 equivalents, more preferably 0.1 to 20 equivalents, and even more preferably 0.2 to 5 equivalents, relative to the C-protected amino acid or C-protected peptide.

[0077] In the method for producing a peptide of the present invention, the peptide chain can be further elongated by repeating the following steps (3) to (5) for the peptide obtained in step (2) a desired number of times. (3) A step of removing the protecting group at the N-terminus of the peptide obtained in step (2) or (5). (4) mixing an N-protected amino acid or an N-protected peptide with a carboxylic acid halide of formula (I); (5) A step of mixing the product obtained in step (4) with the product obtained in step (3). Steps (4) and (5) can be carried out by the same procedures as in steps (1) and (2) above, or by a general peptide synthesis reaction.

[0078] The "N-protected amino acid or N-protected peptide" used in step (4) may be the same as or different from that used in step (1). Similarly, the "carboxylic acid halide represented by formula (I)" used in step (4) may be the same as or different from that used in step (1).

[0079] In the method for producing a peptide of the present invention, purification of the peptide obtained in steps (1) to (5) can be omitted as appropriate, as long as it does not affect the reaction in the subsequent step.

[0080] Process (3) This step is a step of removing the N-terminal protecting group from the peptide obtained in step (2) or step (5).

[0081] The deprotection conditions used in this step are appropriately selected depending on the type of N-terminal protecting group. For example, in the case of a 9-fluorenylmethoxycarbonyl group, deprotection is carried out by treatment with a secondary or tertiary amine (e.g., pyrrolidine, piperidine, morpholine, triethylamine); in the case of a t-butoxycarbonyl group, deprotection is carried out by treatment with an acid (e.g., trifluoroacetic acid, hydrochloric acid, Lewis acid); and in the case of a benzyloxycarbonyl group or an allyloxycarbonyl group, deprotection is carried out in a neutral environment, for example, by hydrogenation in the presence of a metal catalyst.

[0082] In each reaction, when the reaction substrate has a hydroxy group, a mercapto group, an amino group, a carboxy group, or a carbonyl group (particularly when the amino acid or peptide has a functional group in the side chain), a protecting group generally used in peptide chemistry or the like may be introduced into these groups, and the target compound can be obtained by removing the protecting group as necessary after the reaction.

[0083] The protection and deprotection can be carried out by carrying out a protection-deprotection reaction using a commonly known protecting group (see, for example, Protective Groups in Organic Synthesis, Fourth edition, TW Greene, John Wiley & Sons Inc. (2006)). EXAMPLES

[0084] The present invention will be described in more detail below with reference to Reference Examples, Comparative Examples and Synthesis Examples as Examples, but the present invention is not limited to these Examples.

[0085] In this specification, when amino acids and the like are represented by abbreviations, each representation is based on the abbreviations defined by the IUPAC-IUB Commission on Biochemical Nomenclature or the commonly used abbreviations in the relevant field.

[0086] In this specification, a 2-(4,4-dimethylpentan-2-yl)-5,7,7-trimethyloctanoyl group may be abbreviated as ISTA, and a pivaloyl group may be abbreviated as Piv.

[0087] In the synthesis examples, "(v / v)" means (volume / volume) and "M" means mol / L.

[0088] The safety evaluation of ISTA-Cl and ISTA-Br was performed using a differential scanning calorimeter manufactured by METTLER TOLED under the measurement conditions shown below. <Measurement conditions> Temperature range: 30-300℃ Heating rate: 10℃ / min Pan used: Swiss Institute AuPan M20

[0089] Unless otherwise specified, high performance liquid chromatography / mass spectrometry was measured using either Waters ACQUITY UPLC H-Class / QDa, Waters ACQUITY UPLC H-Class / SQD2, or Shimadzu LC-20AD / Triple Tof5600.

[0090] In the description of HPLC / mass spectrometry, ESI+ is electrospray ionization positive mode, and M+H means protonated.

[0091] Unless otherwise specified below, the quantitative yield of each product and by-product was calculated by analysis using high performance liquid chromatography <Analysis Condition 1>. <Analysis conditions 1> High performance liquid chromatography: SHIMADZU LC-20 Column: Agilent Poroshell 120 EC-C18 (2.7 μm, 3.0 x 100 mm) Column oven temperature: 40℃ Eluent: 0.025 vol% trifluoroacetic acid, acetonitrile: 0.025 vol% trifluoroacetic acid aqueous solution 95:5 (0-12 minutes), 95:5 (12 minutes-18 minutes), 10:90 (18.1 minutes-23 minutes) (v / v) Eluent rate: 0.7mL / min Detection wavelength: 210nm

[0092] For purification by silica gel column chromatography, unless otherwise specified, either a Yamazen Hi-Flash column, a Biotage SNAP Ultra Silica Cartridge, Merck Silica Gel 60, or Fuji Silysia Chemical PSQ60B was used.

[0093] In the following Reference Examples and Examples, the yield or quantitative yield may exceed 100%. These are all due to measurement errors, the influence of the purity of the raw materials or products, or other factors based on common technical knowledge. In the following Examples, the causes of the yield exceeding 100% are not mentioned individually, but a person skilled in the art would be able to fully understand the scientific validity of these Examples.

[0094] In the following Reference Examples and Synthesis Examples, quantitative yields of the target product and by-products may be shown, but these are the results of the reaction. In addition, when a quantitative yield is shown, unless otherwise specified regarding the yield of the by-product, it means that no by-products were produced.

[0095] Reference Example 1: Safety evaluation of ISTA-Cl by differential scanning calorimetry ISTA-Cl (16.8 mg) was placed in a gold pan, sealed, and calorimetry was performed. No exothermic peak was observed. This revealed that ISTA-Cl does not have exothermic properties due to self-decomposition. The calorific values ​​of COMU and isobutyl chloroformate are 773.17 J / g and 467.31 J / g, respectively (Non-Patent Document 4).

[0096] Reference Example 2: Safety evaluation of ISTA-Br by differential scanning calorimetry ISTA-Br (14.9 mg) was placed in a gold pan, sealed, and calorimetry was performed. No exothermic peak was observed. This demonstrated that ISTA-Br does not generate heat due to self-decomposition.

[0097] Reference Example 3: Synthesis of ISTA-Br [ka] 2-(4,4-Dimethylpentan-2-yl)-5,7,7-trimethyloctanoic acid (4.0g, 14.06mmol), N,N-dimethylformamide (0.031g, 0.422mmol) were mixed with xylene (4.0mL), thionyl bromide (4.8g, 21.15mmol) was added at 0℃, the mixture was heated to room temperature, and stirred for 1 hour. The reaction solution was distilled under reduced pressure to obtain 2-(4,4-dimethylpentan-2-yl)-5,7,7-trimethyloctanoyl bromide (4.31g, 88% yield) as a transparent liquid.

[0098] Reference Example 4: Synthesis of Fmoc-Val-Phe-OEt [ka]

[0099] Fmoc-Val-OH (0.177g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) were mixed with acetonitrile (1.8mL), and pivaloyl chloride (0.058g, 0.479mmol) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Val-Phe-OEt (target product) in the obtained organic layer was 79%, and the quantitative yield of Piv-Phe-OEt (by-product) was 21%. Piv-Phe-OEt MASS(ESI+)m / z;278.0(M+H)+

[0100] Reference Example 5: Synthesis of Cbz-Val-Phe-OEt [ka]

[0101] Cbz-Val-OH (0.131g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) and acetonitrile (1.3mL) were mixed, and pivaloyl chloride (0.058g, 0.479mmol) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with ethyl acetate (3.0g), and then saturated ammonium chloride aqueous solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Cbz-Val-Phe-OEt (target product) in the obtained organic layer was 77%, and the quantitative yield of Piv-Phe-OEt (by-product) was 22%.

[0102] Reference Example 6: Synthesis of Boc-Val-Phe-OEt [ka]

[0103] Boc-Val-OH (0.114g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) were mixed with acetonitrile (1.1mL), and pivaloyl chloride (0.058g, 0.479mmol) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with ethyl acetate (3.0g), and then saturated ammonium chloride aqueous solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Boc-Val-Phe-OEt (target product) in the obtained organic layer was 79%, and the quantitative yield of Piv-Phe-OEt (by-product) was 18%.

[0104] Reference Example 7: Synthesis of Fmoc-Trp(Boc)-Ala-OBn [ka]

[0105] Fmoc-Trp(Boc)-OH (0.293g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) were mixed with acetonitrile (2.9mL), and pivaloyl chloride (0.061g, 0.509mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with ethyl acetate (3.0g), and then saturated ammonium chloride aqueous solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Trp(Boc)-Ala-OBn (target product) in the obtained organic layer was 84%, and the quantitative yield of Piv-Ala-OBn (by-product) was 10%. Piv-Ala-OBn MASS(ESI+)m / z;264.9(M+H)+

[0106] Reference Example 8: Synthesis of Fmoc-Val-Phe-Phe-OEt [ka]

[0107] Fmoc-Val-OH (0.118g, 0.348mmol), N,N-diisopropylethylamine (0.048g, 0.377mmol) were mixed with acetonitrile (1.2mL), and pivaloyl chloride (0.039g, 0.32mmol) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Phe-Phe-OEt (0.1g, 0.29mmol), N,N-diisopropylethylamine (0.045g, 0.348mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Val-Phe-Phe-OEt (target product) in the obtained organic layer was 85%, and the quantitative yield of Piv-Phe-Phe-OEt (by-product) was 18%. Piv-Phe-Phe-OEt MASS(ESI+)m / z;425.2(M+H)+

[0108] Reference Example 9: Synthesis of Fmoc-Trp(Boc)-Ala-Phe-OEt [ka]

[0109] Fmoc-Trp(Boc)-OH (0.209g, 0.40mmol), N,N-diisopropylethylamine (0.055g, 0.43mmol) were mixed with acetonitrile (2.1mL), and pivaloyl chloride (0.044g, 0.36mmol) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Ala-Phe-OEt (0.1g, 0.33mmol), N,N-diisopropylethylamine (0.051g, 0.40mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The obtained reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Trp(Boc)-Ala-Phe-OEt (target product) in the obtained organic layer was 69%, and the quantitative yield of Piv-Ala-Phe-OEt (by-product) was 26%. Piv-Ala-Phe-OEt MASS(ESI+)m / z;349.1(M+H)+

[0110] Synthesis Example 1: Synthesis of Fmoc-Val-Phe-OEt [ka]

[0111] Fmoc-Val-OH (0.177g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) and acetonitrile (1.8mL) were mixed, and 50% by mass toluene solution (0.29g, 0.479mmol) of 2-(4,4-dimethylpentan-2-yl)-5,7,7-trimethyloctanoyl chloride (ISTA-Cl) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol) and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The reaction mixture was diluted with tetrahydrofuran (3.0 g) and ethyl acetate (3.0 g), and then saturated aqueous ammonium chloride solution (1.0 g) and water (1.0 g) were added to separate the mixture. The aqueous layer was separated again with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Val-Phe-OEt (target product) in the obtained organic layer was 95%, and the quantitative yield of ISTA-Phe-OEt (by-product) was 1%. Fmoc-Val-Phe-OEt MASS(ESI+)m / z;515.7(M+H)+ ISTA-Phe-OEt MASS(ESI+)m / z;461.1(M+H)+

[0112] Synthesis Example 2: Synthesis of Fmoc-Val-Phe-OEt [ka]

[0113] Fmoc-Val-OH (0.207g, 0.610mmol), N,N-diisopropylethylamine (0.084g, 0.650mmol) were mixed with acetonitrile (1mL), and a solution of 2-(4,4-dimethylpentan-2-yl)-5,7,7-trimethyloctanoyl bromide (ISTA-Br) (0.197g, 0.566) and toluene (0.197g) was added at 0℃ and stirred for 1 hour. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.526mmol), and acetonitrile (1.0mL) was added to the reaction solution and stirred for 1 hour. The resulting reaction solution was diluted with tetrahydrofuran (3.0 g) and ethyl acetate (6.0 g), and then 10% by mass potassium hydrogen sulfate aqueous solution (1.0 g) and water (2.0 g) were added and separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Val-Phe-OEt (target product) in the obtained organic layer was 94%, and the quantitative yield of ISTA-Phe-OEt (by-product) was 1%. Fmoc-Val-Phe-OEt MASS(ESI+)m / z;515.7(M+H)+ ISTA-Phe-OEt MASS(ESI+)m / z;461.1(M+H)+

[0114] Synthesis Example 3: Synthesis of Cbz-Val-Phe-OEt [ka]

[0115] Cbz-Val-OH (0.131g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) and acetonitrile (1.3mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.29g, 0.479mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Cbz-Val-Phe-OEt (target product) in the obtained organic layer was 95%, and the quantitative yield of ISTA-Phe-OEt (by-product) was 1%. Cbz-Val-Phe-OEt MASS(ESI+)m / z;427.6(M+H)+

[0116] Synthesis Example 4: Synthesis of Boc-Val-Phe-OEt [ka]

[0117] Boc-Val-OH (0.114g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) and acetonitrile (1.1mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.29g, 0.479mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Boc-Val-Phe-OEt (target product) in the obtained organic layer was 96%, and the quantitative yield of ISTA-Phe-OEt (by-product) was 1%. Boc-Val-Phe-OEt MASS(ESI+)m / z;393.5(M+H)+

[0118] Synthesis Example 5: Synthesis of Fmoc-Phe-Phe-OEt [ka]

[0119] Fmoc-Phe-OH (0.202 g, 0.522 mmol), N,N-diisopropylethylamine (0.073 g, 0.566 mmol) were mixed with acetonitrile (2.0 mL), and a 50% by mass solution of ISTA-Cl in toluene (0.29 g, 0.479 mmol) was added at 0°C and stirred for 2 hours. A solution of HCl·H-Phe-OEt (0.1 g, 0.435 mmol), N,N-diisopropylethylamine (0.068 g, 0.522 mmol), and acetonitrile (1.0 mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with tetrahydrofuran (3.0 g) and ethyl acetate (3.0 g), and then saturated aqueous ammonium chloride solution (1.0 g) and water (1.0 g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Phe-Phe-OEt (target compound) in the obtained organic layer was 101%. Fmoc-Phe-Phe-OEt MASS(ESI+)m / z;563.6(M+H)+

[0120] Synthesis Example 6: Synthesis of Cbz-Phe-Phe-OEt [ka]

[0121] Cbz-Phe-OH (0.156g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) and acetonitrile (1.6mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.29g, 0.479mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Cbz-Phe-Phe-OEt (target compound) in the obtained organic layer was 99%. Cbz-Phe-Phe-OEt MASS(ESI+)m / z;563.6(M+H)+

[0122] Synthesis Example 7: Synthesis of Fmoc-Phe-MePhe-OMe [ka]

[0123] Fmoc-Phe-OH (0.202 g, 0.522 mmol), N,N-diisopropylethylamine (0.073 g, 0.566 mmol) were mixed with acetonitrile (2.0 mL), and a 50% by mass solution of ISTA-Cl in toluene (0.29 g, 0.479 mmol) was added at 0°C and stirred for 2 hours. A solution of HCl·H-MePhe-OMe (0.1 g, 0.435 mmol), N,N-diisopropylethylamine (0.068 g, 0.522 mmol), and acetonitrile (1.0 mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0 g), and then saturated aqueous ammonium chloride solution (1.0 g) and water (1.0 g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Phe-MePhe-OMe (target product) in the obtained organic layer was 91%. Fmoc-Phe-MePhe-OMe MASS(ESI+)m / z;563.7(M+H)+

[0124] Synthesis Example 8: Synthesis of Fmoc-Leu-Gly-OBn [ka]

[0125] Fmoc-Leu-OH (0.210g, 0.60mmol), N,N-diisopropylethylamine (0.083g, 0.65mmol) were mixed with acetonitrile (2.1mL), and a 50% by mass solution of ISTA-Cl in toluene (0.330g, 0.55mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Gly-OBn (0.1g, 0.50mmol), N,N-diisopropylethylamine (0.077g, 0.60mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Leu-Gly-OBn (target product) in the obtained organic layer was 98%, and the quantitative yield of Piv-Gly-OBn (by-product) was 1%. Fmoc-Leu-Gly-OBn MASS(ESI+)m / z;501.6(M+H)+ ISTA-Gly-OBn MASS(ESI+)m / z;432.7(M+H)+

[0126] Synthesis Example 9: Synthesis of Fmoc-Trp(Boc)-Gly-OBn [ka]

[0127] Fmoc-Trp(Boc)-OH (0.313g, 0.60mmol), N,N-diisopropylethylamine (0.083g, 0.65mmol) were mixed with acetonitrile (3.1mL), and a 50% by mass solution of ISTA-Cl in toluene (0.330g, 0.55mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Gly-OBn (0.1g, 0.50mmol), N,N-diisopropylethylamine (0.077g, 0.60mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Trp(Boc)-Gly-OBn (target product) in the obtained organic layer was 93%, and the quantitative yield of ISTA-Gly-OBn (by-product) was 2%. Fmoc-Trp-Gly-OBn MASS(ESI+)m / z; 574.3(M+H)+ (detected as Boc-free product)

[0128] Synthesis Example 10: Synthesis of Fmoc-Leu-Ala-OBn [ka]

[0129] Fmoc-Leu-OH (0.197g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) and acetonitrile (2.0mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.309g, 0.509mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Leu-Ala-OBn (target product) in the obtained organic layer was 98%. Fmoc-Leu-Ala-OBn MASS(ESI+)m / z;515.7(M+H)+

[0130] Synthesis Example 11: Synthesis of Fmoc-Trp(Boc)-Ala-OBn [ka]

[0131] Fmoc-Trp(Boc)-OH (0.293g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) were mixed with acetonitrile (2.9mL), and a 50% by mass solution of ISTA-Cl in toluene (0.309g, 0.509mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Trp(Boc)-Ala-OBn (target product) in the obtained organic layer was 96%, and the quantitative yield of ISTA-Ala-OBn (by-product) was 1%. Fmoc-Trp(Boc)-Ala-OBn MASS(ESI+)m / z;688.4(M+H)+ ISTA-Ala-OBn MASS(ESI+)m / z;446.3(M+H)+

[0132] Synthesis Example 12: Synthesis of Fmoc-Ser(tBu)-Ala-OBn [ka]

[0133] Fmoc-Ser(tBu)-OH (0.213g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) were mixed with acetonitrile (2.1mL), and a 50% by mass solution of ISTA-Cl in toluene (0.309g, 0.509mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Ser(tBu)-Ala-OBn (target compound) in the obtained organic layer was 97%. Fmoc-Ser(tBu)-Ala-OBn MASS(ESI+)m / z;545.3(M+H)+

[0134] Synthesis Example 13: Synthesis of Fmoc-Cys(Trt)-Ala-OBn [ka]

[0135] Fmoc-Cys(Trt)-OH (0.326g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) were mixed with acetonitrile (3.3mL), and a 50% by mass solution of ISTA-Cl in toluene (0.309g, 0.509mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Cys(Trt)-Ala-OBn (target compound) in the obtained organic layer was 96%.

[0136] Synthesis Example 14: Synthesis of Fmoc-Thr(tBu)-Ala-OBn [ka]

[0137] Fmoc-Thr(tBu)-OH (0.221g, 0.556mmol), N,N-diisopropylethylamine (0.078g, 0.602mmol) and acetonitrile (2.2mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.309g, 0.509mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Ala-OBn (0.1g, 0.463mmol), N,N-diisopropylethylamine (0.072g, 0.556mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Thr(tBu)-Ala-OBn (target compound) in the obtained organic layer was 92%. Fmoc-Thr(tBu)-Ala-OBn MASS(ESI+)m / z;559.4(M+H)+

[0138] Synthesis Example 15: Synthesis of Fmoc-Asp(tBu)-Phe-OEt [ka]

[0139] Fmoc-Asp(tBu)-OH (0.215g, 0.522mmol), N,N-diisopropylethylamine (0.073g, 0.566mmol) were mixed with acetonitrile (2.2mL), and a 50% by mass solution of ISTA-Cl in toluene (0.29g, 0.479mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Phe-OEt (0.1g, 0.435mmol), N,N-diisopropylethylamine (0.068g, 0.522mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Fmoc-Asp(tBu)-Phe-OEt (target product) in the obtained organic layer was 99%, and the quantitative yield of ISTA-Phe-OEt (by-product) was 1%. Fmoc-Asp(tBu)-Phe-OEt MASS(ESI+)m / z;587.7(M+H)+

[0140] Synthesis Example 16: Synthesis of Boc-Gly-Pro-OBn [ka]

[0141] Boc-Gly-OH (0.086g, 0.50mmol), N,N-diisopropylethylamine (0.070g, 0.538mmol) and acetonitrile (0.9mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.27g, 0.456mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Pro-OBn (0.1g, 0.414mmol), N,N-diisopropylethylamine (0.053g, 0.50mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Boc-Gly-Pro-OBn (target compound) in the obtained organic layer was 91%. Boc-Gly-Pro-OBn MASS(ESI+)m / z;363.1(M+H)+

[0142] Synthesis Example 17: Synthesis of Boc-Asp(tBu)-Leu-OMe [ka]

[0143] Boc-Asp(tBu)-OH (0.191g, 0.66mmol), N,N-diisopropylethylamine (0.092g, 0.715mmol) were mixed with acetonitrile (2.0mL), and a 50% by mass solution of ISTA-Cl in toluene (0.367g, 0.61mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Leu-OMe (0.1g, 0.55mmol), N,N-diisopropylethylamine (0.085g, 0.66mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Boc-Asp(tBu)-Leu-OMe (target compound) in the obtained organic layer was 99%. Boc-Asp(tBu)-Leu-OMe MASS(ESI+)m / z;417.1(M+H)+

[0144] Synthesis Example 18: Synthesis of Boc-Tyr(tBu)-Orn(Boc)-OMe [ka]

[0145] Boc-Tyr(tBu)-OH (0.142g, 0.42mmol), N,N-diisopropylethylamine (0.059g, 0.46mmol) were mixed with acetonitrile (1.4mL), and a 50% by mass solution of ISTA-Cl in toluene (0.233g, 0.39mmol) was added at 0℃ and stirred for 2 hours. A solution of HCl·H-Orn(Boc)-OMe (0.1g, 0.35mmol), N,N-diisopropylethylamine (0.054g, 0.42mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added to separate the liquids. The aqueous layer was again separated with ethyl acetate (2.0 g), and the quantitative yield of Boc-Tyr(tBu)-Orn(Boc)-OMe (target product) in the obtained organic layer was 99%. Boc-Tyr(tBu)-Orn(Boc)-OMe MASS(ESI+)m / z;566.4(M+H)+

[0146] Synthesis Example 19: Synthesis of Fmoc-Val-Phe-Phe-OEt [ka]

[0147] Fmoc-Val-OH (0.118g, 0.348mmol), N,N-diisopropylethylamine (0.048g, 0.377mmol) were mixed with acetonitrile (1.2mL), and a 50% by mass solution of ISTA-Cl in toluene (0.193g, 0.32mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Phe-Phe-OEt (0.1g, 0.29mmol), N,N-diisopropylethylamine (0.045g, 0.348mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Val-Phe-Phe-OEt (target product) in the obtained organic layer was 96%, and the yield of ISTA-Phe-Phe-OEt (by-product) was 1%. Fmoc-Val-Phe-Phe-OEt MASS(ESI+)m / z;662.4(M+H)+ ISTA-Phe-Phe-OEt MASS(ESI+)m / z;607.4(M+H)+

[0148] Synthesis Example 20: Synthesis of Fmoc-Asp(tBu)-Phe-Phe-OEt [ka]

[0149] Fmoc-Asp(tBu)-OH (0.143g, 0.348mmol), N,N-diisopropylethylamine (0.048g, 0.377mmol) were mixed with acetonitrile (1.4mL), and a 50% by mass solution of ISTA-Cl in toluene (0.193g, 0.32mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Phe-Phe-OEt (0.1g, 0.29mmol), N,N-diisopropylethylamine (0.045g, 0.348mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Asp(tBu)-Phe-Phe-OEt (target compound) in the obtained organic layer was 94%. Fmoc-Asp(tBu)-Phe-Phe-OEt MASS(ESI+)m / z;734.4(M+H)+

[0150] Synthesis Example 21: Synthesis of Fmoc-Trp(Boc)-Ala-Phe-OEt [ka]

[0151] Fmoc-Trp(Boc)-OH (0.209g, 0.40mmol), N,N-diisopropylethylamine (0.055g, 0.43mmol) were mixed with acetonitrile (2.1mL), and a 50% by mass solution of ISTA-Cl in toluene (0.220g, 0.36mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Ala-Phe-OEt (0.1g, 0.33mmol), N,N-diisopropylethylamine (0.051g, 0.40mmol), and acetonitrile (1.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with tetrahydrofuran (3.0g) and ethyl acetate (3.0g), and then saturated aqueous ammonium chloride solution (1.0g) and water (1.0g) were added and the liquids were separated. The aqueous layer was again separated with ethyl acetate (3.0 g), and the quantitative yield of Fmoc-Trp(Boc)-Ala-Phe-OEt (target product) in the obtained organic layer was 97%, and the quantitative yield of ISTA-Ala-Phe-OEt (by-product) was 1%. Fmoc-Trp(Boc)-Ala-Phe-OEt MASS(ESI+)m / z;773.4(M+H)+ ISTA-Ala-Phe-OEt MASS(ESI+)m / z;531.3(M+H)+

[0152] Synthesis Example 22: Synthesis of Boc-Val-Phe-OEt [ka]

[0153] Boc-Val-OH (0.340g, 1.56mmol), N,N-diisopropylethylamine (0.219g, 1.69mmol) and acetonitrile (3.4mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.870g, 1.43mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H-Phe-OEt (0.3g, 1.3mmol), N,N-diisopropylethylamine (0.203g, 1.32mmol), and acetonitrile (3.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with ethyl acetate (6.0g), and then saturated aqueous ammonium chloride solution (3.0g) and water (2.0g) were added to separate the liquids. The aqueous layer was separated again with ethyl acetate (3.0 g), and the resulting organic layer was concentrated and purified by silica gel column chromatography to obtain Boc-Val-Phe-OEt (0.49 g, yield 95%) as a white solid. Boc-Val-Phe-OEt MASS(ESI+)m / z;393.2(M+H)+

[0154] Synthesis Example 23: Synthesis of Cbz-Val-Val-Phe-OEt [ka]

[0155] Boc-Val-Phe-OEt (0.20 g, 0.51 mmol) was mixed with 4M-HCl / 1,4-dioxane (1.0 mL) and stirred for 2 hours at 25° C. The reaction solution was concentrated, and the resulting white solid HCl·H-Val-Phe-OEt was used in the next step. Cbz-Val-OH (0.154g, 0.612mmol), N,N-diisopropylethylamine (0.086g, 0.663mmol) and acetonitrile (3.0mL) were mixed, and a 50% by mass solution of ISTA-Cl in toluene (0.340g, 0.561mmol) was added at 0℃ and stirred for 3 hours. A solution of HCl·H―Val-Phe-OEt (0.51mmol), N,N-diisopropylethylamine (0.079g, 0.612mmol), and acetonitrile (2.0mL) was added to this solution and stirred for 1 hour. The resulting reaction solution was diluted with acetonitrile (4.0g) and ethyl acetate (8.0g), and then saturated aqueous ammonium chloride solution (2.0g) and water (2.0g) were added and the liquids were separated. The aqueous layer was separated again with ethyl acetate (5.0 g), dried over sodium sulfate, and the organic layer was concentrated to give a white solid. Ethyl acetate (4.0 g) was added to the white solid, and the precipitated solid was collected by filtration to give Cbz-Val-Val-Phe-OEt (0.26 g, yield 97%) as a white solid. Cbz-Val-Val-Phe-OEt MASS(ESI+)m / z;526.7(M+H)+

[0156] Synthesis Example 24: Synthesis of Fmoc-Val-Val-Val-Phe-OEt [ka]

[0157] Cbz-Val-Val-Phe-OEt (0.10 g, 0.19 mmol) was dissolved in 2,2,2-trifluoroethanol (4 mL), and 10% by mass Pd-C (20 mg) was added, followed by stirring for 1 hour under a hydrogen gas atmosphere at 25° C. The reaction solution was filtered and concentrated, and the resulting H-Val-Val-Phe-OEt was used in the next step. Fmoc-Val-OH (0.077 g, 0.228 mmol), N,N-diisopropylethylamine (0.032 g, 0.247 mmol) were mixed with acetonitrile (0.77 mL), and a 50% by mass solution of ISTA-Cl in toluene (0.127 g, 0.21 mmol) was added at 0°C and stirred for 3 hours. A solution obtained in the previous step, H-Val-Val-Phe-OEt (0.19 mmol), acetonitrile (1.0 mL), and NMP (0.2 mL) were added to this solution and stirred for 1 hour. Diisopropyl ether (10 mL) was added to the obtained reaction solution, and the precipitated solid was filtered to obtain Fmoc-Val-Val-Val-Phe-OEt (0.14 g, 103% yield) as a white solid. Fmoc-Val-Val-Val-Phe-OEt MASS(ESI+)m / z;713.4(M+H)+

[0158] Synthesis Example 25: Synthesis of Fmoc-Phe-Phe-OH (1) NMP (1.0 mL) was added to H-Phe-2-ClTrt resin (109.9 mg, 0.1 mmol, Merck, resin is polystyrene), and the mixture was stirred for 15 minutes, after which the solvent was removed. (2) Fmoc-Phe-OH (155.0 mg, 0.4 mmol) and N,N-diisopropylethylamine (0.07 mL, 0.4 mmol) were dissolved in NMP (0.8 mL), and then a 50% by mass solution of ISTA-Cl in toluene (0.24 mL, 0.4 mmol) was added at room temperature and stirred for 3 hours. (3) The solid obtained in (1) was added to the solution obtained in (2) above and stirred for 1 hour. After removing the reaction solvent, the solid obtained was washed with NMP and methanol in that order and then dried to obtain Fmoc-Phe-Phe-2-ClTrt resin. (4) Acetic acid / 2,2,2-trifluoroethanol / dichloromethane (volume ratio 1 / 2 / 7) (2 mL) was added to the total amount of Fmoc-Phe-Phe-2-ClTrt resin obtained in (3) above, and the mixture was stirred for 2 hours. After removing the desorbed resin by filtration, the reaction solution was concentrated under reduced pressure, diisopropyl ether was added to the residue, and the precipitated solid was collected by filtration to obtain Fmoc-Phe-Phe-OH (51.4 mg, yield 96.1%) as a white solid. Fmoc-Phe-Phe-OH MASS(ESI+)m / z;535.3(M+H)+ [Industrial Applicability]

[0159] The present invention provides a highly efficient method for producing peptides.

Claims

1. The following steps (1) and (2): （１） an N-protected amino acid or an N-protected peptide and a compound represented by formula (II) 【Chemistry 1】 (wherein X represents a halogen atom) and a carboxylic acid halide represented by the formula: （２） mixing the product obtained in step (1) with a C-protected amino acid or a C-protected peptide; A method for producing a peptide, comprising:

2. Further, the following steps (3) to (5): （３） A step of removing the protecting group at the N-terminus of the peptide obtained in step (2) or (5); （４） mixing an N-protected amino acid or an N-protected peptide with a carboxylic acid halide of formula (II); and （５） mixing the product obtained in step (4) with the product obtained in step (3); The method for producing the peptide according to claim 1, comprising one or more repeats of the formula:

3. The method for producing the peptide according to claim 1 or 2, wherein X is a chlorine atom or a bromine atom.

4. The method for producing the peptide according to any one of claims 1 to 3, wherein X is a chlorine atom.

5. The method for producing a peptide according to any one of claims 1 to 4, wherein the N-protected amino acid is an α-amino acid other than glycine.

6. The method for producing a peptide according to claim 5, wherein the amino acid in the N-protected amino acid is an α-amino acid other than glycine, and the reactive functional group of the amino acid side chain is protected.

7. 7. The method for producing a peptide according to claim 5 or 6, wherein the α-amino acid other than glycine is valine, phenylalanine, threonine, leucine, tryptophan, serine, cysteine, aspartic acid or tyrosine.

8. The method for producing a peptide according to any one of claims 1 to 7, wherein the amino acid in the C-protected amino acid or the amino acid at the N-terminal residue of the C-protected peptide is an α-amino acid other than an N-substituted amino acid.

9. The method for producing a peptide according to any one of claims 1 to 8, wherein the step (2) is a step of mixing the product obtained in the step (1) with a C-protected peptide.

10. 10. The method for producing a peptide according to claim 1, wherein the N-terminal protecting group of the N-protected amino acid or the N-protected peptide is a carbamate protecting group.

11. The method for producing a peptide according to claim 10, wherein the carbamate-based protecting group is a 9-fluorenylmethyloxycarbonyl group or a benzyloxycarbonyl group.

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