JPH0249799A - Production of peptide - Google Patents

Abstract

PURPOSE:To obtain peptides using a protecting group capable of removal under so mild conditions as to be stable against acids, bases and reduction by protecting a imidazole group of histidine with 4-methoxy-2,3,6-trimethylbenzenesulfonyl group and synthesizing a peptide. CONSTITUTION:In production of a peptide containing histidines, imidazole groups of the histidines are reacted with 4-methoxy-2,3,6-trimethylbenzenesulfonyl chloride to form histidine derivatives represented by the formula (R is H or protective group of-amino group; 4-Methoxy-2,3,6-trimethylbenzenesulfonyl group is bonded to N at 1 or 3 position of A ring.) where each imidazole group is protected with a 4-methoxy-2,3,6-trimethylbenzenesulfonyl group and then peptide condensation is carried out. The protecting groups are subsequently removed using an acid or 1-hydroxybenzotriazole to produce the objective peptide.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for producing a peptide by protecting the imidazole group of histidine or a salt thereof.

When synthesizing peptides containing histidine, the imidazole group dissociates near neutrality and exhibits weak basicity, so it is difficult to solve various technical problems, such as unbinding the imidazole group during synthesis. ification.

This often causes difficulties in purification after synthesis and ease of racemization, so a useful protecting group is desired.

The protecting group is preferably one that can selectively cleave the α-amino protecting group, is stable to 1-lifluoroacetic acid treatment or catalytic reduction, and can be removed by a mild method.

As conventional protecting groups, benzyl groups, carbobenzoxy groups, etc. are known, but since these are removed by catalytic reduction, it is therefore not possible to use a carbobenzoxy group as the α-amino (ν5 group). Valatoluenesulfonyl group has been known as a protecting group that is stable against both catalytic reduction and trifluoroacetic acid treatment, but surprisingly this protecting group is extremely unstable under basic conditions. It is stable, and a considerable amount of the protecting group is often removed during neutralization after acid treatment (for example, when neutralizing with triethylamine).Protecting groups that are improved from this type of protecting group Recently, the 4-methoxybenzenesulfonyl group has been known as a protective group, but even in the case of this protecting group, the stability under basic conditions is insufficient. As a result of examining the protective groups for It was discovered that it could be removed through treatment, and as a result of further research, the present invention was completed.

That is, in the production of a peptide having histidine, the present invention protects the imidazole group of histidine with a 4-methoxy-2,3,6-trimethylbenzenesulfonyl group, performs peptide condensation, and then converts the 1y-protecting group with an acid. Alternatively, the method for producing a peptide is characterized in that the peptide is eliminated using l-hydroxybensito J:r sol.

In the present invention, - maximum [wherein R is hydrogen or α-amino group HS l+'
; represents that the 4-methoxy-2,3,6-1-limethylbenzenesulfonyl group is bonded to either the 1st or 3rd position of the ring (A)]; and its salts are used.

The 4-methoxy-2,3,6-trimethylbenzenesulfonyl group used in the present invention is a novel ("a NX Jf
It is usually used in the form of a halide, and is produced, for example, by the method of Reference Example 1.

In the present invention, histidine having an imidazole group protected with a 4-methoxy-2,3,6-toJ methylbenzenesulfonyl group can be produced, for example, by the following conventional method. In other words, it can be produced by reacting 4-methoxy-2,3,6-1-limethylbenzenesulfonyl halide, preferably chloride, with histidine in which the α-ami7 group is protected. This reaction is, for example, about -1
Solvents (e.g., water, aqueous dihydrofuran, aqueous dioxane) at a suitable temperature ranging from 0° to +50°C.

It may also be carried out using aqueous acetone, aqueous acetonitrile, aqueous dimethylformamide). As a base,
Various bases can be used, and among them, sodium hydrogen carbonate, sodium carbonate, triethylamine, etc. are preferred. As the protecting group for α-ami7 group, known protecting groups such as carbobenzoxy group, p-nitrobenzyloxycarbonyl group, p-methoxybenzyloxycarbonyl group, L-butoxycarbonyl group, L-amyloxycarbonyl group, etc. group, 9-fluorenylmethoxycarbonyl group, innicotinyloxycarbonyl group.

0-nitrophenylsulfenyl L2-(p-biphenyl)isopropyloxycarbonyl group, etc. can be used,
Among these, those in which the α-amino group of histidine is protected with a carbobenzoxy group or an L-butquine carbonyl group are advantageously used, such as dichlorohexylamine and cyclohexylamine.

It is used in peptide condensation as a salt such as sodium.

The thus obtained 4-methoxy-2,3゜6-drimethylbenzenesulfonyl-promoted histidine having an imidazole group is subjected to a peptide condensation reaction with the amino acid residue by a conventional method. Advantageously (
Can be used for cars. As a common method for this, for example, M1 mouth od
Peptide Synthesis, by M.
sis).

1ater 5science, New York,
1966; F. M. Finn and I.
lormann'if The Proteins
Proteins), 2nd Q, Il, Neura
th, R, I7. Edited by l1iee, Acade
++ic Press Inc, New York,
1976: Methods described in "Peptide Synthesis" by Nobuo Izumiya et al., Marukubi Co., Ltd., 1975, such as the azide method, chloride method, acid anhydride method, mixed acid anhydride method, and DCC method.

Active ester method, method using Woodward reagent I (
Carbonimidazole method, redox method, DCC/1
Examples include the 1ON13 method.

By the way, in order to perform peptide condensation, α-・r-mino (1, kQN whose word group is L-butoxycarbonyl group) needs to be acid-treated with trifluoroacetic acid or the like, and then neutralized with a base such as triethylamine. However, the para-toluenesulfonyl group that has been commonly used in the past is extremely unstable under the basic conditions described in 1, and even the para-methoxybenzenesulfonyl group is insufficiently stable. In comparison, 4-methoxy-2,3,6-1-
Limethylbenzenesulfonyl group is extremely stable, as shown in the examples below, and its usefulness is extremely high.

After peptide condensation, the N5L of the invention is then removed by acid or l-hydroxybenzotriazole.

As the desorption method using an acid, known acid treatment methods such as anhydrous hydrogen fluoride, methanesulfonic acid, trifluoromethanesulfonic acid, etc. can be applied. Furthermore, in the case of the method of the present invention,
As a new acid treatment method, trifluoroacetic acid or trifluoroacetic acid containing a small amount of methanesulfonic acid can be advantageously used, and the elimination reaction proceeds favorably, especially when carried out under rjIL of thioanisole, anisole or dimethyl sulfide. .

As described below, the protecting group of the present invention can be used for catalytic reduction,
It has characteristics not found in the conventional imidazole group (J, NY 4N), such as being stable against acids such as trifluoroacetic acid and stable against bases such as triethylamine.

Next, the present invention will be described in more detail with reference to reference examples and examples. In this specification, regarding amino acids, peptides, protecting groups, active groups, etc., ItlPAC-1u
11commission on 13io1og
It may be indicated by an abbreviation based on icalNomenclaLure or by an abbreviation commonly used in the field. Examples of them are shown below. Trp: hributophane; Ly
s: Rigi7; His+histidine; Δrg: arginine; Ser: serine; Gly nigericin; Ala: alanine; Pro nibroline; Thr: threonine; Gin:
Glutamine; Val: valine; Leu: oisine; li
e: Isoleucine; Met: Methionine; Tyr: Tyrosine (Hereinafter, unless otherwise specified, amino acids refer to the L-form. (excluding jj L G I y <);
Z: carbobenzoxy; Boc: L-butoxycarbonyl; OBu': t-butyl ester・110 N B
and ONB: N-noloxy-5-fulbornene-2,3-dicarboxinimide and its ester; 1
(OBt:N-'\idroquine benztriazole,
1) CC: N, N'-one/chlorohexyl/carbodiimide; DCU: N.

N'-dicycloheki/luurea; Mbs: 4-methoxy7
Benzenesulfonyl; Pme: pentamethylbenzenesulfonyl; Mtr: 4-methoxy-2,3,6-drimethylbenzenesulfonyl; C11A: cyclohexylamine; DC; HΔnidicyclohexylamine +D
MF Nidimethylformamide; 'rEA Nitriethylamine; Also, the developing solvent for thin layer chromatography shown in this specification is R1'': Chloroform-methanol-acetic acid (9:1:0.5); Rr' Ethyl acetate-pyridine -acetic acid-water (60:206: lO); R
r3: Chloroform-methanol water (7: 3: 0
．． 5); Rr4 di-n-butanol-hybrid.

Lysine-acetic acid-water (30:20:6:24).

Reference Example 1 (1) Synthesis of 2,3,5-trimethylanisole 2.
3.5-t.limethylphenol 10g, methyl iodide I
O,/Iau to dimethyl sulfoxide 7 100 rr&
The mixture was stirred and cooled with water, and 5.6 g of 60% oily hydrogenated thorium was added thereto, followed by stirring for 10 hours. After adding water to this, it was extracted with ether, and the ether layer was washed with water and dried over anhydrous sodium sulfate. The solvent was distilled off to obtain a sheet. Yield: 12.9 g (quantitative).

(2) Synthesis of 4-methoxy-2,3,6-1-limethylbenzenesulfonyl chloride 2.3.5-1 Limethylanisole (4.5 g) was dissolved in 500 methylene chloride, -5° to -10° After cooling to 0.degree. C., 400 g of a methylene chloride solution containing 6 chlorsulfonic acid and Ohfl was added dropwise. Thereafter, the temperature was returned to room temperature and poured onto water containing 5% sodium bicarbonate/water. The methylene chloride layer was washed with water and then dried over anhydrous magnesium sulfate. After the solvent was distilled off, crystals were collected by filtration from n-hexane. Yield 11t5.0g (67,
0%) Melting point 56-58℃ Elemental analysis C, oII, 303S Calculated value as CQ: C48,29, H5,21: S 12.89; C
& 14.26 Experimental value C4B, 42: II5.21: S
12.61: Cf214.25 Reference Example 2 Production of 110c11is(Mbs)-011-DCIIA 130G-11is-0111, 50g, water 101n
1. It was dissolved in a mixed solution of 10 d of acetone and cooled with water. After adding TEAL, 65- to this, while stirring vigorously, a solution prepared by dissolving 21 g of 1Abs-CQl in acetone was added dropwise, and the mixture was stirred at room temperature for 1 hour. After adding citric acid, the acetone was distilled off, extracted with ethyl acetate, and dried using anhydrous sodium sulfate.

After adding D CHA 1 and 061tIl and distilling off the solvent, the residual oil crystallized when left overnight in a refrigerator, so ether was added and this was collected by filtration. Yield: 2.65g (74.0%) Melting point: 146-
147°C Elemental bone Fr C30114807N Old calculation value as 4S: C59,38; H7,64; N 9.23
:S 5.29 Experimental value: C59,16: [17,70・N8.9
53 5.28 Reference Example 3 Production of 130cm11is(MLr)-011'DCII^ BOC-11is-Off 5.1 g was mixed with 30+J of water, 301r of acetone, 1. and cooled with water. After adding '1゛IζΔ 5.60bfl to this, while stirring vigorously, Mtr-CQ
30 J of a solution prepared by dissolving 4.97 g in acetone was added dropwise, and the mixture was stirred for 2 hours.

After distilling off the acetone, the mixture was acidified with citric acid, extracted with ethyl acetate, and dried over anhydrous sodium sulfate. D
After adding CIIA 3.60d, the solvent was distilled off,
When the residual oil was left in the refrigerator overnight, it crystallized, so ether was added to recrystallize it, and this was collected by filtration. Yield ff17.80g (60°1%) Melting point 136-137
°C R [” 0.63 Elemental analysis C,, I2.0.N3S -C,, H13N
Calculated value as: C61,08;'118.08;
N 8.64; 3 4.94 Experimental value: C61,19; H8,05; N 8
．． 89;3 4.13 Reference Example 4 Production of Z-11is(MLr)-011・DCllA 7
, -11is-0112, 90 g was dissolved in a mixture of 15 parts of water and 15 parts of acetone and cooled with water. Add to this TEA2.
After adding 80ml, stir vigorously and add Mtr-
15 J of a solution prepared by dissolving 22.49 g of CI in acetone was added dropwise, and the mixture was stirred for about 1 hour. After distilling off the acetone, acidify with citric acid and extract with ethyl acetate.
It was dried using anhydrous sodium sulfate. DCl-I
After adding A 2 rrdl, the solvent was distilled off, ether was added, and the crystals were collected by filtration.

Revenue! it 4. lOg (60,0%), melting point 149
150°C Rr 0, 6 2 Elemental analysis Cz-14I707N 3S' C1zl
Calculated value as l t, N・C63,32; I+ 7
．． 38; N 8.21+3 4.70 Experimental value: C62,94; H7,11; N 8.
11;3 4.80 Test Example 4-MeI-xy-2,3,6-)limethylbenzenesulfonyl J! (MLr), 4-methoxybenzenesulfonyl group (Mbs), valatoluenesulfonyl group (
Comparison of stability under basic conditions of BOC-11
is(Mtr)-011・DCllA, BOC-11
is(Mbs)-011・DCi! A.

Take 100 mg each of 130C-11is(Tos)-011/DCIIA, dissolve it in 50% water-containing methanol containing Q,5N-triethylamine, and leave it at 26°C for a certain period of time. 1 knee 1 fixed, fiL. As shown in the attached table, the MLr group is much more stable than Ike's protective groups (Mbs, Tos), confirming its usefulness.

Reference Example 5 Production of It-11is (MLr)-011 Z-11is
After 3.4 g of (MLr)-011'DCII^ was centrifuged to 100% ethyl acetate, IN-sulfuric acid was added thereto, water was further added, the mixture was stirred, and then dried over anhydrous sodium sulfate. After distilling off the solvent, methanol 10(b
When dissolved in J solution, catalytically reduced, and distilled off the solvent, crystals will precipitate, so add ether and remove all of them. Collection ff
i 1.80 g (98.3%) Melting point 162-164
°C (α] −271 (C = 1.01.acetic acid), Rr”
0.35 Elemental analysis C,,11,. 05N 3S -1/31-
1, Calculated value as O: C51,60,1-15,55
: N 11.28°S 8.61 Experimental value: C51,76; II 5.63; N
11.08°S 8.63 Example 1, lJG RI) Production of (Gastrin Relieving Peptide) (1) Production BO of 130cmLeu-MetNII
C-MetNIIt (10,3g) to 4N-)4C
After adding 40 d of C/acetic acid and shaking, crystals precipitated, so ether was added, and the crystals were collected by filtration and dried.

Dissolve this in 200ml of DMr, cool on ice, and serve 7 people for 12 people.
0 Jll was added to neutralize. To this, BOC-Leu-O
N[(BOC-Leu-OR8,0g, HONB6
．． 85E, prepared from 7.83 g of BOC),
- Stirred at night. After the solvent was distilled off, the residue was dissolved in ethyl acetate, washed with 11W water and citric acid water, and then dried using anhydrous sulfuric acid (IJum). When the solvent was distilled off, crystals were precipitated, so ether was added thereto, the crystals were collected by filtration, and recrystallized from methanol-ether. Storage lid 10.8g (
86.3%) Melting point 152-154t:, (α]2b"
-34-2°(c-1,03,I)JF'),
Rflo, 65 elemental analysis Calculated value as C engineering 6H3104N3S: C53,15,1 (8,64 pieces NIL63
iS8.87'# Numerical value: C53-56i [8,72;
N 11. 4'r; 38.92 (21BOCR18 (Mtr) OK-DCHAod construction B
OC-f (isoH5, 11 g was dissolved in a mixture of 30 d of water and 30 g of acetone, 1 O?11, and cooled on ice. After adding 5.6 d of TE to this, Mtr-C14, 97
30ml/g of acetone solution was added and stirred for 2 hours.

After acetone was distilled off under reduced pressure, the residue was acidified with citric acid, extracted with ethyl acetate, washed with water, and then dried using anhydrous sulfuric acid. After evaporating the solvent, it was dissolved in a small amount of ethyl acetate, D CEi & 3.6d was added, the solvent was evaporated and the mixture was left overnight in a refrigerator. Ether was added to the precipitated crystals, which were collected by filtration. Income *7. gog(
6o, 1%) Melting point 136-137℃, [α]
18.8° (c-1,01, methanol), Rfl
o, 63 elemental analysis C33I (calculated value as 520'7"43: C61,08; ■8.08i N 8.
64i34.94 Experimental value: C61,19; Hs, 05; N 8-
89iS4.73 (31BOC-Hls(Mtr)-Lau-Met12
Production of BOC-Leu-%AetNH210, Og,
4 N-11C1/acetic acid 3011 was added, and the mixture was allowed to stand at room temperature for 20 minutes, and then ether was added to form a precipitate, which was collected by filtration and dried.

On the other hand, 130C41s (lJtr) Oli-DCHA
16.Og was heated to ethyl acetate, branched by adding 27 ml of IN-Me acid, and dried using anhydrous sulfuric acid.

After distilling off the solvent, dissolve in 100 ml of acetonitrile, add 5.0 g of HONB, cool on ice, and add 5.8 g of DCC.
was added and reacted for one night, and then the precipitated DCU was filtered off. The previously prepared amine component was dissolved in DMF'lOOgZ, neutralized with 4.6 liters of TEA, added with active ester, stirred for 6 hours, distilled off the dissolved μ, and then dissolved in ethyl acetate. ,′! It was washed with backwater, citric acid water, and water in this order, and dried with anhydrous sulfuric acid. After distilling off the solution, ether was added and the powder was collected by filtration.

Yield 15.1g (8'6.0%) Melting point 129-131℃, Ca)v-13,so(c
-0,87,DIJF), Rfl O,64 elemental analysis C32H5008”632 as total Xff1:
C54,06i) 17.09; N 11.82;
39.02 Experimental value: C54,69; H7,39; N 11.
97iS8.39 (4) Production of Z-Val-Gly-OBut Z-G
ly-OBut12. After catalytic reduction of Og in methanol, the solvent was distilled off, and the residue was dissolved in DMF' and cooled on ice. To this, Z-Mal-OH8, 80g, HONB7.2
0g and 8.24g of DCC were added and stirred overnight. After filtering off the precipitated DCU, the residue was dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and then washed with anhydrous acid.)
It was dried with IJum. The solvent was distilled off, petroleum benzine was added to the residue, the crystals were collected by filtration, and then recrystallized from ethyl acetate-petroleum benzine. Accommodation f1111.1g (87
,0%) Melting point 141℃, (α)'b3-2.1°(c
-1,16°DMF'), Rflo, 80 Elemental analysis Calculated value as C19H2805N2: C6
2,62i H7,74i N 7.69 Experiment f1:
C62,49i H7,60i N 7.72(5)
UaZ-V of Z-Ala-Mal-Gly-OBut
After catalytic reduction of al-Gly-OButl 0 and Og in methanol, it was dissolved in DIJ Floogl, and 5.7 g of ZAla-OH and 4.1 g of HONB were added.
Ice cold. 6.2 g of DCC was added to this and stirred overnight. When the precipitated DCU was filtered off and the solvent was distilled off, crystals were precipitated, so ethyl acetate was added, the crystals were collected by filtration, and thoroughly washed.

Revenue! i 10.5g (96.4%) Melting point 184-185℃, [α)%3 7. B°(c-
1, 06, DMF'), Rfl Q. 67 elemental analysis C
Total X value as 22H3306N3: C60,67i
H7,64i N 9.65 East experimental value: C60,93i
H7,86i N 9.77(6) Z-Trp(
Production Z of Mtr)-Ala-Mal-Gly-OBut
-Ala-4al-Gly-OBut5, Og was catalytically reduced in methanol, the solvent was distilled off, and DM
F' It was dissolved in 100 ml. In addition, Z-Trp(M
tr) 16.34 g of OF and 86 g of HOBTl were added, cooled on ice, and stirred overnight with 2.85 g of DCC.

The precipitated DCU was filtered and the solvent was distilled off, then extracted with ethyl acetate containing a small amount of n-butanol, washed with aqueous sodium bicarbonate, and dried over IJum. The solvent was distilled off, ether was added, and the precipitate was collected by filtration, crystallized from methanol-ethyl acetate-ether, and filtered.

Yield s, sog (9s, 6%) Melting point 154-155℃, [α] -22.7° (c=
0,88. DMF'), Rflo, 68 elemental analysis Calculated value as C43H5601°N5S: C
61,85+ H6,76; N 8.39iS3.8
4 Experiment @i: C62, 06i H7, Oli N 8
．． 58i33.58 (7) BOC-1(1s(Mtr)-Trp(ut
r) Production of -Ala-val-Gly-OBut Z-Trp(Mtr)-Ala-Mal-Gly 0B
After catalytic reduction of ut8 and Og in DMF'-methanol, methanol was distilled off. In addition, BOC-L
lia(Mtr)0)1(BOC-Hls(lJtr)
OH-DCHA 6. prepared from 22g), HONB
After adding 90 g of BOC and cooling on ice, 2.18 g of BOC was added and stirred overnight.

After the precipitated DCU was filtered off, the solvent was distilled off, ether was added, and the precipitate was collected by filtration. This is methanol,
Washed with a mixture of ethyl acetate/I/ and ether. Yield 1
o, 2g (92,5%) melting point 202-204℃,
[α] Xi-16.1° (C-1, 18, DIF”)
, Rflo, 68 elemental analysis C56"77014N
Total X rank as 832: C58,46; H6,75i
N 9.74iS5.57 Experimental value: C58,32i H6,54i N 9.5
8i35.51 f8) BOC-8er-Hls(Mtr)-Trp
Production of (Mtr)-Ala-Val-01yOH BOC-Hla (Mtr)-Trp(Mtr)-
Ala-”/al-GlyOBut4.OgKTF'A
Add 30gj, shake for 50 minutes at room temperature, then add T.
I'A was distilled off, ether was added, and the precipitate was collected by filtration and dried. On the other hand, BOC-8er-OH0, 76-E
, HON B 0 , 72 g in acetonitrile 20
11, cooled on ice, added 83 g of DCCo, and stirred for 4 hours. The previously prepared amine component was DM
F' was dissolved in 50 ml and neutralized by adding 0ml of TEAl, followed by adding the active ester and stirring overnight.

After distilling off the nitrogen, add a small amount of acetic acid, then add water and filter it as a precipitate. Mr. Denshi reprecipitates this from DMF'-water. Yield i 3.60g (85.0%) Melting point 1
48-152°C, [α)26'-t6.4° (0 pieces 1.
09. DMF') Elemental analysis Calculated value as C55H74016N9S2・2H20: C54,26i H6,46i N 10
．． 36iS5.27 Experiment (ili: C54,58; H6,33i
N 10.82iS5.51 (91Production of Z-Arg(Pme)-Gly-OBut 13 g of Z-Gly-OBut was mixed with 300 g of methanol/
After catalytic reduction, the solvent was distilled off and the residue was purified by D! ,
I combed it to IF'200d. To this, Z-Arg(Pm
s)OH(Z-Arg(Pme)OH・CHA 2
(prepared from 0 g), cooled on ice, and added 5.4 g of HOBT,
8.2g of BOC was added and stirred overnight. Precipitated DC
After U was filtered off, the solvent was distilled off and the residue was dissolved in ethyl acetate. After washing this with a sodium bicarbonate solution and a citric acid solution, it was dried with anhydrous acid (IJum). After distilling off the solvent, petroleum benzine was added and the powder was collected by filtration. Yield 19. sg (9s, o%) Melting point 55-60℃, [α
] Kanju〇, 2° (c -0,88, DMF')
, Rflo, 62 Elemental analysis Calculated value as C31H4507N5S: C
58,93i H7,18i N 11.09iS5.
08 Experimental values: C58,96i H7, Oli N 10.
67i(10) Production of Z-Tyr-Pro-OBut Z-Pro○But 15. Og to Methano-A/30
After catalytic reduction, the solvent was distilled off to 0MF.
I combed it to 400sl. To this, Z-Tyr-OH (Z-
Tyr-OH-DCHA 20, from Og M),
Add 6.75 g of HoBT, cool on ice, and add DCCIo.
.

Added 4g and stirred overnight. After the precipitated DCU was filtered off and the solvent was distilled off, it was dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and dried over anhydrous sodium sulfate. After distilling off the solvent, a silica gel cuff (7.5x
9c+++) and eluted with 1% MeOH/chloroform. Fractions of the target product were collected and concentrated. Petroleum benzine was added thereto and the powder was collected by filtration. Yield 15.4g (82
,2%)(α)2D3-39.9°(Q-0,83,D
MF'), Rflo, 62 Elemental analysis C26H3206N2 ・Calculated value as 'AH20: C65,39i El 6.97; N 5
．． 87 experimental value: C65,7oi H6,93i N
5.66 (11) Z-Engineering 1e-Tyr-Pro-O
ButZ-Tyr-Pro-OButl 5, 2 g
After dissolving in methanol 300ze and catalytic reduction,
The solvent was distilled off and the residue was dissolved in 200 ml of DMF'. To this, Z-11eOH8, Og, HONB6.5g
After adding and cooling on ice, 7.4 g of BOC was added and stirred overnight. After adding a small amount of N,N-dimethylpropanthamine, the DCU was filtered off and the solvent was distilled off under reduced pressure.

The residue was dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and dried over anhydrous sodium sulfate. When the solvent was distilled off, crystals precipitated, so petroleum benzine was added and the crystals were collected by filtration, and then recrystallized with methanol and ether/L/1 petroleum benzine. Yield 10.9g (62,5
%) Melting point 177-178℃, [α] also 3-3g, 3°
(c-1, ll, DMF'), Rflo-
62 element analysis Calculated value as C32H4307N3:
C66,07i H7,45i N 7.22 Experimental value:
C66, 07i H7, 74i N 7.19 (12
) Production of Z-IIs-Tyr-Pro -OH Z-
IIs-Tyr-Pro OBu 6, T to Og
Add 60ml of F' and stir at room temperature for 1 hour.
The residue was dissolved in ethyl acetate, washed with water, and dried over anhydrous sulfuric acid. After distilling off the solvent, ether was added to form a powder, which was collected by filtration.

Yield 5.10g (94.2%) Melting point 72-74°C, [α] also 3-25.2° (c -
1,01, DMr), Rflo, 44 Elemental analysis Calculated value as 028H3507N3: C6
3,98i H6,71i N 7.99 trial [1:
C63,75i H6,67; tJ 7.84 (13
) Z-11e-Tyr-Pro-Arg(Pme)
-Production of GlyOBut Z-Arg(Pme)-Gly-OBut7, 58
Dissolve g in methanol 300g/IN-hydrochloric acid 12
After adding d, catalytic reduction was carried out. After distilling off the solvent, the residue was dissolved in DIJrlOOgl, cooled on ice, and TEA
2.00M1 was added to neutralize. In addition, Z-11e-
Tyr-Pro-OH5,78q, HOBT2.
23g and 3.40g of DCC were added and stirred overnight.

After precipitated DCU was filtered off, the solvent was distilled off and the residue was dissolved in ethyl acetate. This was washed with aqueous sodium bicarbonate and citric acid, and dried with anhydrous sulfuric acid (IJ worm). The solvent was distilled off and the residue was poured into a silica gel cuff (5.5×10C
11) and eluted with 2% Meal (/chloroform). Fractions of the target product were collected and separated into 1 layer, then ether was added and the powder was collected by filtration.

Yield 7.2ag (62.2%) Melting point 110-112℃, [α] -28.3° (c
-1,19,DMF'), Rflo, 59 elemental analysis Calculated value as C51''72011NBS-H2O:
C59,85; H7,29i N 10.93iS
3.13 Experimental values: C60,16i H7,56i N
10.95iS2.98 (14) Z-I, ys(lJtr)-Eng 1e-Ty
Production of r-Pro-Arg(Pms)-Gly0But Z-11e-Tyr-Pro-Arg(Pme)-G
ly-OBut7, Og was dissolved in methanol 350111, and 7 ml of IN-hydrochloric acid was added for catalytic reduction. After evaporating the solvent, the residue was dissolved in DIJF'200m, cooled on ice, and neutralized by adding TEAl, 0m/. To this, Z-Lys(Mtr)-OH(Z-Lys(lJ
tr)OH-DCHA4.

prepared from 74g), HonTt2og, Dccl, 80
Add g and mix overnight. The precipitated DCU was filtered off and the residue was dissolved in ethyl acetate. This was washed with M stored water and citric acid water, and then dried over anhydrous sodium sulfate.

The solvent was distilled off, ether was added to form a powder, and the powder was reprecipitated twice from ethyl acetate-ether. Revenue: 118.40
g (87,5%) Melting point 116-118°C, [α] temporary -1969° (c'-1,05, DMF'), Rflo
, 59 elemental analysis Calculated as C66H96015N10S2 [: C59,44; H7,26i N 10.
50iS4.81 Experimental value: C59,10i H7,44; N 10.
46; (15) Z-Thr-Lys(Mtr)-1
1e-Tyr-Pro-Arg(Pme)-Gly-
Production of OBut Z-Ly8(Mtr)-(1a-Tyr
-Pro-Arg(Pme)-GlyOBut7,5
After catalytic reduction of 0 g in 350 ml of methanol,
The solvent was distilled off, and the residue was dissolved in 100 g/dMF'.

To this, Z-Thr-OHL49g, HONBl, 51
After cooling on ice, 73 g of DCCl was added and stirred overnight. After processing a small amount of N,N-dimethylpropanediamine and filtering off DCU, the solvent was distilled off and the residue was dissolved in ethyl acetate containing a small amount of n-butanol.

This was washed with aqueous sodium bicarbonate, then water, and dried over anhydrous sodium sulfate. The solvent was distilled off, and ether was added to the residue to form a powder, which was then collected by filtration. Yield 8. Og
(99,2%) Melting point 122-124℃, [α] also 3-2s, s° (c
-0,82,DuF'), Rflo, 59 elemental analysis Calculated value as C'70H103017"1.LS2: C58,60i H7,24; N 10.74i
S4.47 Experimental value: C58,61i fl 7.29i N 1
0.47i(16) BOC-Lau-Thr-Ly
s(Mtr)-IIs-Tyr-Pro-Arg(P
Production of Z-Thr-Lya(me)-Gly 0But
Mtr)-11e-Tyr-Pro-Arg(Pma)
-G1yOBut7, 5 g methanol 350g1
! After adding 0.9914-puff toluenesulfonic acid and catalytic reduction, the solvent was distilled off. D this
The mixture was dissolved in Md. 100d, cooled with water, and neutralized by adding 74mt of TEAo. This Ic, BOC-Leu-OH1
37 g of HONBl, 41 g of DCCl, and 62 g of DCCl were added and stirred overnight. After filtering off DCU and distilling off the solvent, it was dissolved in ethyl acetate and diluted with aqueous sodium bicarbonate.

It was washed with citric acid water and dried with anhydrous sulfuric acid (IJum). After distilling off the spies, ether was added and the mixture was filtered as a powder. Yield ff17.55g (95.4%) Melting point 1
36-138°C, [α)2b3-26. (3°(o-
x, oO, DMr), Rfl O, 59 elemental analysis C'
Calculated value as 73F1116018N12S2・H2O: C57,23i H7,76; N 10.97i
S4.19 Experimental value: C57-53+ H8,12i N 10.
65i33.90 (17) BOC-Ala-Leu-Thr-Lye
(Mtr)-IIs-Tyr-Pro-human rg(Pm
a) -Gly-OR H'5hBOC-Leu-Thr
-Lya (Mtr)-11e-Tyr-Pro-
Arg(Pme)-Gly-OBut4, 20 g
40d containing KTF' was added, and after shaking at room temperature for 60 minutes, the solvent was distilled off, ether was added, the powder was collected by filtration, and the mixture was dried. This was dissolved in DMF' 50 #l, cooled with water, and TIAO-80gt was added. to this,
BOC-Ala-ONB (BOC-Ala-OH0,5
2g, HONBo.

54 g (prepared from 62 g of DCCO) were added and stirred overnight. After adding a small amount of N,N-dimethylpropanediamine and distilling off the solvent, aqueous acetic acid was added to form a precipitate, which was filtered. This was reprecipitated with aqueous methanol. Yield 3.60g (sg, t%) RA point 128-130
℃, Ca3%” -32,9°(c-0,98,DM
F'), Rfl(L40 elemental analysis C'72"113
Calculated value as 019N13S2/2H20: C55,
26i H7,s4; N 11.64; S4.10 Experimental value: C55,02i L(6,89i N 1
1.38iS3.77 (18) Z-3er-Pro 0But manufacturing Z-
Pro-OButl l, Og with methanol 300at
After catalytic reduction, the solvent was distilled off and DMF'
I combed it to 200tZ. Z-3er-OH7,2 to this
E, add 4.90g of HOBT and 7.50g of BOC,
- Stirred at night. The precipitated DCU was filtered off, the solvent was distilled off, and the residue was dissolved in ethyl acetate. Add this to baking soda water,
After washing with citric acid water, it was dried with anhydrous sulfuric acid ram. The solvent was distilled off, petroleum benzine was added, the crystals were collected by filtration, and then recrystallized from ethyl acetate-ether.

Yield fi 9.50g (67.2%) Melting point 126-127℃, [a]′2b”-so,
9° (c-0, 95, DMF'), Rfl 0
．． 65 element analysis Calculated value as C20H2806N2:
C61,21; H7,19; H7,14 experimental value:
061.45i 1(7,16i N 7.31(1
9) Manufacture of Z-Gly-8er-Pro-OBut Z-3er-Pro-OButl Q, Q g was dissolved in 300 g of methanol for catalytic reduction, the solvent was distilled off, and it was dissolved in DIJ'F300 m/. . This is K
After adding 5.06 g of Z-Gly-OH and 5.13 g of HONB and cooling on ice, 5.89 g of BOC was added and stirred overnight. After adding a small amount of N,N-dimethylpropanediamine, DCU was filtered off and f8B was distilled off. The residue was dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and dried over anhydrous sulfuric acid. Distill the solvent,
Petroleum benzine was added to the crystals, which were collected by filtration and then recrystallized from ether. Yield 7.70g (72,1%) Melting point 96-98℃, (a) Xi-53,4° (c-
x.

05, DMF'), Rflo, 61 Elemental analysis Calculated value as C22H3□07N3: C5
8,78i H6,95; N 9.35 real M[: C
5B, 86+ H7,04i N (146(20)
Production of Z-Gly-Gly-3er-Pro-OBut Z-Gly-8er-Pro-OBut7,
After Og was dissolved in 200 g/methanol and subjected to catalytic reduction, the solvent was distilled off and the residue was dissolved in 100 g/DMF. This is KZ-Gly-OR3.0g, UONB3.1
After adding 0 g and cooling on ice, 3.60 g of DCC was added and stirred overnight. After adding a small amount of N,N-dimethylpropanediamine, the DCU was filtered off and the solvent was distilled off.

The residue was dissolved in ethyl acetate, washed with saturated brine, and then dried over anhydrous sodium sulfate. The solvent was distilled off and ether was added to form crystals, which were then collected and recrystallized from ethyl acetate. Yield 5.7og (7g, 5%) Melting point 129-130℃, [α] Temporary -47.00 (c=
0.81, DMF'), Rflo, 47 elemental analysis
Total X[:C56,90i as C24H3408N4
1 (6,77i N 11.06 trial [: C56,7
5; H6,68; N 10.90 (21) Z-
Manufacturing of Gln-Pro-OBut Z-Pro-OBut
16. Dissolve 2 g in methanol/L/350ml,
After catalytic reduction, the solvent was distilled off and dissolved in DMr2001/. To this, Z-Gin-OH12゜4g, ) (OB
T7. 16 g was added, cooled with water, 9 g of DCCIo was added, and the mixture was stirred overnight. The precipitated DCU was filtered and the solvent was distilled off, then dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and dried over anhydrous sulfuric acid.

After distilling off the solvent, petroleum benzine was added to form crystals, which were collected by filtration and recrystallized from ethyl acetate-petroleum benzine.

Yield 15. sg (s2.5%) Melting point 106-107℃, [α] -51.2° (c-
=-1,06, DMF'), Rf" 0.62 Elemental analysis Calculated as C22H3106N3 ill: C60,
95; H7, 21i N 9.69 East D*: C6
0,951H7-36; N 9.41(22) Z
-Production of Leu-Gln-Pro-OBut Z-Gln
-Pro-OBu 8. 3.51 g of para-toluenesulfonic acid was dissolved in methanol #300 for catalytic reduction, and then the solvent was distilled off. DMF'2 the residue
After thawing to 0.00ml and cooling on ice, T'EA 2.6g? was added to neutralize it. To this, Z-LeuOH (Z-Leu-
OH-DCHA 8, 24 g) JJW),
HoNB4, OOg, and DCC4.60g were added and stirred overnight. After adding a small amount of N,N-dimethylpropanediamine, DCU was filtered off and the solvent was distilled off.

The residue was dissolved in ethyl acetate, washed with aqueous sodium bicarbonate and citric acid, and dried over anhydrous chloride. After the solvent was distilled off to precipitate crystals, petroleum benzine was added and the crystals were collected by filtration and recrystallized from ethyl acetate-petroleum benzine. Yield 7.7og (76.3%) Melting point 62-64℃, [α
] Kan-51.7° (c-t.

11, DMF'), Rfl O,61 elemental analysis Calculated value as C28''4207N4: C61,5
2i H7,75; N 10.25%! Experiment [: C6
1,19; H7,75; N 10.11 (23)
BOC-Pro-Leu-Gin-Pro-OBut
Production of Z-Leu-Gin-Pro-OBu 7,
After dissolving Og in methanol/I/300m/ and performing catalytic reduction, the solvent was distilled off and the residue was dissolved in DMF'100m.
I made it to 1.

To this, BOC-Pro O [(2,42g, HON
After adding 76g of B2 and cooling on ice, add 3.17g of DCC.
Added and stirred overnight. After adding a small amount of N,N-dimethylpropanediamine, the DCU was filtered off, the solvent was evaporated and dissolved in ethyl acetate. This is l1tV
After washing with water and citric acid solution, it was dried with anhydrous acid solution. After distilling off the solvent, the powder was filtered from ether-petroleum benzine. Yield 6-5og (s3
．． 3%) Melting point 74-76℃, C(1)2D3-76, 3°(
c-1゜19, DMF'), RflO-60 elemental analysis Calculated value as C30H5108''5: C59,o
si H8,43; N 11.49 Experiment 411:
C5B, 899 H8,i2; N 11.08 (2
4) BOC-Ala-Pro-Leu-Gln-P
Production of ro-OH BOC-Pro-Leu-Gin-P
60 g of TF'A was added to ro-OBut6 and Og, and after stirring at room temperature for 60 minutes, the solvent was distilled off, and ether was added to the residue to form a powder, which was collected by filtration and dried. Dissolve this in DMF'1QQd, cool on ice, and use TEA2.80.
After adding d, BOC-Ala-○HE (BOC-
Ala-OH1, 95g, I (ONB2.07g, BO
C2 (prepared from 38 g) was added and stirred overnight. After distilling off the solvent, add a small amount of acetic acid and then add ether/l.
/ was added and filtered as a powder. Dissolve this in chloroform and use a silica gel column (!ls, 5x8c+++
) and eluted with 5% methanol/chloroform to take a fraction of the desired product, concentrate it, and then add ether and collect it by filtration as a powder. Yield fi 4.50g (73-2%) Melting point 124-128℃, [α] also 3-82.10 (c
-1,04,DMF'), RflO, 18 elemental analysis Calculated as C29H4809N6 tit: C55,
75; H7,74i N 13.45 Experimental value: C5
5,38i H7,65i N 13.21 (2
5) BOC-Ala-Pro-Leu-Gln-P
Production of ro-Gly-Gly-8or-Pro-OBut Z-Gly-Gly-3er-Pro-OBut2.2
3 g was dissolved in 100 jl of methanol for catalytic reduction, the solvent was distilled off, and the residue was dissolved in DMF' 50 at
I grinned.

To this, BOC-Aha-Pr□-Leu-Gln-Pr
50 g of o-OH2 and 90 g of HONBo were added and cooled on ice, and further 3 g of DCCl was added and stirred overnight.

After the precipitated DCU was filtered off, the solvent was distilled off, and this was dissolved in ethyl acetate containing Major's n-BuOH, washed with saturated brine, and dried over anhydrous sodium sulfate.

After distilling off the solvent, ether was added and the powder was collected by filtration. Yield 3.85g (98.3%) Melting point 100-1
05℃, [α] Suga-74.0' (c-0, 92, DII
F), Rfl 0.19 elemental analysis C45H74
Calculated as 014NIO”'20ff: C54,2
oi H7,68i N 14.05 experiment if: C
54,01$ H7,45B N 13.44 (26)
BOC-Ala-Pro-Leu-Gln-Pro
-Production of Gly-Gly-3et-Pro-OH noc-Human 1a-Pro-Leu-01n-Pro-G
ly-Gay-6ar-Pro-OButl, Og
After adding 10 ml of fcT F' and stirring at room temperature for 1 hour, the solvent was distilled off, ether was added, and a powder was collected by filtration. Dissolve this in 10ml of DMF, cool on ice, and add TEAo, 46z? elJD, t, next KBOC-0) 1
0.27 g was added and stirred for 4 hours. After distilling off the solvent, 1.5 al of acetic acid was added, ether was added, and a powder was collected by filtration.

This was reprecipitated from methanol-ether. Yield 0
．． 87g (92.4%) Melting point 141-145℃, [α]2♂-72.9° (c
-0,98,DMr), Rf30.29 elemental analysis Calculated as C41H66014NIO”3H20 f
fL: C50-40i f(7,43+ N 14-
34 experimental value: C50,16i)! 6.76; N
14.11(27) BOC-3ar-[(1s(
utr)-Trp(Mtr)-Ala-Val-Gl
Production of y-Hla(Mtr)Leu-MetNl2BOC-H1s()Atr)-Leu-MetNH23
, Tl'''A 40 d was added to 97 g, and the mixture was stirred at room temperature for 10 minutes, then 4.1 al of 1.3N hydrochloric acid was added, the solvent was distilled off, ether was added, the powder was collected by filtration, and dried. .Dissolve this in DMF'40ml, cool with water, and
Add Bko, 86ml, then BOC-8ep-1 (
18(Mtr)-Trp(Mtr)-Ala-'V
al-Gly-○H5,50g, HONBl, Og, D
15 g of CCl was added and stirred overnight. Precipitated DCU
After filtering and distilling off the solvent, water was added and filtered to make a powder. This was then washed with aqueous ethal.

Yield ffk 7.35g (88.9%) Melting point 192-193℃, [α] Sweet 12.5° (c-
1,05,DMF'), RflO,50 elemental analysis Calculated as C82H114021"15"4 [: C
55,51i H6,48i N 11.84i37.
23 Experiment ill: C55,56i 1 (6,74i N
11.83iS6.55 (28) BOC-8er-Hls-Trp(hlt
r)-Ala-4al-G17-flIB-Leu
Production of Mat Nl2 BOC-8or-H1s (Mtr
)-Trp(Mtr)-Ala-Val-Gly-H
la(Mtr)-Leu-Met-Nl23, O
g was dissolved in 15 liters of DMF', 2.30 g of HOBT was added, and the mixture was treated for 30 minutes.The solvent was distilled off, and ether was added to obtain a powder by filtration. Yield 2.15g (92,
5%) Melting point 191-193℃, [α] Suga -18,8° (C stripe 0, 94.DMF'), Rf30.64 Elemental analysis Total as C62H90015N15S2'''20; Value: C54,45〼H6,78HN 15.36;S4
．． 69 Token Town: C53,92i H6,43i N 15.
5oiS4.34 (29) BOC-Ala-Leu-Thr-Lye
(Mtr)-11e -Tyr-Pro-Arg(Pm
a)-Gly-3er-f(ia-Trp(Mtr
)-Ala-Van-Gly-Hl 5-Leu-IJ
Production of et-Nl2 BOC-3er-Hla-Trp(Mtr)-Ala
-'/al-Gly-H1s-Leu-MetNH22
, OgKT F'A (20 ml) was added thereto, and the mixture was treated at room temperature for 15 minutes and then distilled off. Ko-Te/J was added thereto, the powder was collected by filtration, and the mixture was dried. This was dissolved in DMF5ffil, 24 gt of TEAl was added thereto, and the mixture was stirred well. Ether was added to settle the mixture, and the mixture was filtered as a powder. Combining this to Aete/'20 stl, BOC-Al
a-Leu-Thr-Lya(Mtr)-Eng 1e-Ty
r-Pro-Arg(Pma)-Gly-01(1,7
After adding 8g, L (ONBo, 41g), cool on ice, DC
47 g of Co was added and stirred overnight. After distilling off the solvent, ethanol-ethyl acetate was added and the powder was filtered and washed with hot ethanol.

Yield 3.30g (9 (17%) 1 point 222-223℃ (decomposition), [α is '1)'' -
22.8°cc=t, 02. DMI'), Rf30.6
7 element analysis C129H193031”2834・8H
Calculated value as 20: C53,34i f'(7,25
; N 13.50; S4.42 Experiment ffl: C5135; l(6,98; N
12.91; 34.58 (30) BOC-Ala-Pro-Leu-01n
, -Pro-Gly-Gly-Bor-Pro-Ala
-Leu-Thr-Ly8(Mtr)-11e-Ty
r-Pro-Arg(Pme)-Gly-3er-H
l 80Jtr)-Trp(Mtr)-Ala-'/a
Production of l-Gly-H1s-Leu-Met-Nl2 BOC-Ala-Leu-Thr-Lya (jJtr)
-Xle-Tyr-Pro-λrg(Pme)-Gly
-3er-Hls-Trp(Mtr)-Ala-Van
-Gly-Hls-Leu-Mat-Nl2500 q
T F A 5 at was added to the mixture, stirred, and then distilled off. Ether was added and the mixture was filtered as a powder. DI this
The mixture was dissolved in JFld, 2 ml of TEAo was added thereto, and after stirring well, ether was added and the mixture was filtered as a powder.

This was dissolved in DMF'5m, and BOC-Ala-Pr
o-Leu-Gin-Pro-Gly-Gly-3er
-Pro-OH217 da and EOBT50q were added, and then cooled on ice.7 and DCC16 (1') were added and stirred overnight.

After the precipitated DCU was filtered off, the solvent was distilled off, and ethyl acetate was added to the residue, which was filtered as a powder. DM this
After dissolving it in P''-methanol and heating for about 30 minutes, the methanol was distilled off, and ethyl acetate was added to the residue, which was filtered as a powder.Then, this was washed with methanol-water. Yield: 1480M (72°) 2%) ; Melting point 203-208°C (decomposition), [α] Sweet 35.
2゜(c-0,93,DIJP), Rf30.66 elemental analysis C165H24q042N3asc6H20tL
Calculation fit: C53, 95i H7, iN 1
4.49iS3.49 Experimentffl: C54,08i H6,87i N
14.19i33.43 (31) H-λ1a-Pro-Leu-Gln −
Pro-Gly-]1y-8er-Pro-A
ha-Lau-Thr-Lya-Ile-Tyr-Pr
.

-Arg-Gly-3er-Hls-Trp-Ala-
Mal-Gly-Hla-Leu-Met-Nl2 (
, k G RP ) production BOC-Aha-Pro-
Leu-Gln-Pro-Gly-Gly-8ar-P
ro-Ala-Lau-Thr-Lye(Mtr)-English-Tyr-Pro-Arg(Pme)-Gly-
8ar-Hla-Trp(Mtr)-Person 1a-1/a
l-Gly-H1s-Leu-Mat-Nl2 1 0
Add 0.3M methane-nulphonic acid/TF'A-naoaniso-/I/(9:t)sg,g to 0q, stir at room temperature for 2 hours, then add 1ooq of acetic acid ammonia,
TF'A was distilled off, ether was added, and the powder was collected by filtration. A small amount of IN vinegar was dissolved in reverse water, and this was applied to a Sephadex G-25 column (2.2 x 120 cm). Elute with IN-acetic acid 170m? Ambs
After passing through a rlite RA410 (1xlOcm) column, a carboxymethylcellulose column (2,
2 x 17 cm). Water (400g/) - 0.4M
Elution was performed using a linear gradient method using aqueous acetic acid ammonia (40011t), and fractions from 335 to 365 d were collected and lyophilized.

Revenue: 28 Da (35%).

(α)%2-102.2° (Q-0,32,1% acetic acid)
.

Rf' O-39 Amino acid analysis 1t (4% thioglycolic acid/6N hydrochloric acid hydrolysis): Lys 1.00(1)i Hla 1
．． 73(2); Arg 1-04(1); Trp
0.67(1), Thrl, 06(1)+Sar
1.77(2)iGlu 1.11(1)iPro
4.20(4); Gly 4.23(4); Ala
3.19(3)i val 1.06(1)i Me
t 1-02(1)+l1eO,96(1)+Le
u 3.04 (3) + Tyr 1.00 (1) (average recovery rate 73.7%)

Claims (3)

[Claims] (1) When producing a peptide containing histidine, the imidazole group of histidine was replaced with 4-methoxy-2,3,
A method for producing a peptide, which comprises protecting the peptide with a 6-trimethylbenzenesulfonyl group, condensing the peptide, and then removing the protecting group with an acid or 1-hydroxybenzotriazole. (2) The manufacturing method according to claim (1), using trifluoroacetic acid as the acid. (3) The production method according to claim (1) or (2), wherein the protecting group is removed in the presence of thioanisole or dimethyl sulfide.