JPS6259295A - Novel process for producing peptide - Google Patents

Abstract

PURPOSE:To obtain a peptide applicable in the fields of hormone, agricultural chemical, food, etc., efficiently, by reacting a specific alpha-amino acid with a trialkylsilylimidazole and reacting the product with a specific active ester compound in an organic solvent. CONSTITUTION:The objective compound of formula IV [R<1> is H, 1-10C alkyl, 2-6C alkenyl, phenyl, etc.; R<2> is H, amino-protecting group or group of formula V (R<4> is R<2>); R<3> is H or lower alkyl] by reacting (A) an alpha-amino acid of formula I (R<5> is H, 1-10C alkyl, 2-6C alkenyl, phenyl, 7-9C aralkyl, etc.) with (B) a trialkylsilylimidazole and reacting the reaction product with (C) an active ester compound of formula II [R<6> is H, 1-10C alkyl, 2-6C alkenyl, phenyl, 7-9C aralkyl, etc.; R<7> is amino-protecting group or group of formula III (R<8> is amino-protecting group)] (preferably pyridyl thioester) in an organic solvent (preferably amide-type, ether-type or chlorine-containing solvent) preferably at 30-0 deg.C.

Description

Detailed Description of the Invention (Industrial Application Field) The present invention relates to the general formula (1), where R1 is a hydrogen atom, an alkyl group having 1 to 10 carbon atoms, an alkenyl group having 2 to 6 carbon atoms, a phenyl group, Carbon number 7-9
aralkyl group having 1 to 6 carbon atoms substituted with a protected or unprotected hydroxyl group, an aralkyl group having 7 to 9 carbon atoms substituted with a protected or unprotected hydroxyl group, substituted with a lower alkylthio group an alkyl group having 2 to 6 carbon atoms, or a thiol group having 1 carbon number substituted with a protected or unprotected thiol group
~4 alkyl group or indolylmethyl group.

R2 is a hydrogen atom, a protecting group for an amino group, or a group (here, R
1 represents the same meaning as above, R4 represents a hydrogen atom or a protecting group for an amino group), and R3 represents a hydrogen atom or a lower alkyl group] relates to a new method for producing peptides.

More specifically, the present invention is an extremely useful method for synthesizing takuputide without decomposition of active esters in organic solvents or racemization with bases, and is also a breakthrough in that α-amino acids can be used without protective groups. This is a typical method.

The compounds obtained by the method of the present invention include various hormones,
It is useful as an agricultural chemical, a food (for example, sweetening amount), or as an intermediate for synthesizing the final target compound having various uses as described above from the compound.

(Prior Art) Synthesis of Iptite 9 is roughly divided into in-phase synthesis method and liquid phase synthesis method. The in-phase method requires fewer purification steps (b'), and therefore has the advantage of being able to perform rapid synthesis and saving labor by using a synthesis machine. On the other hand, the liquid phase method is generally superior in yield and racemization rate in each step, and both methods have advantages.

In the liquid phase method, there are three basic methods for synthesis. (1) C-terminal activation method (21 condensation method) (3) N-terminal activation method (incyanate method) In method (1), the carboxyl group of one amino acid is activated as an ester, and the other amino acid and a base are present. Alternatively, there is a method of condensation in the absence of the amino acid, and method (2) is a method in which the carboxyl group of one amino acid becomes an active ester in a flask by using a condensing agent, and is condensed with the amino group of the other amino acid. .

These two methods are the most commonly used for each glaze haptide synthesis in the liquid phase method.

However, in both cases, it is usually essential to protect the carboxyl group of the amine component condensed to the active ester by esterification.

Amino acids form dissociated inner salts and are electrically neutralized. Therefore, it is poorly soluble in water and organic solvents.

For this reason, in the liquid phase method, the carboxyl group must be protected and neutralized in order to bring out the basicity (nucleophilicity) of the nitrogen of the amine group.

One way to bring out the basicity of the amine in an amino acid is to add a strong base to form a carboxylate. As this method, Schotten-no-Z-Uman
Baumann method is known (Bodanski (B.
Peptide Synthesis Second Edition (Peptide 5ynthesi)
(s2nd Ea,) p. 85) However, in this method, the active ester group undergoes a competitive reaction between the amino group and the hydrolysis by water, making it unstable (
It is not possible to use active esters with high activity.

(Problems to be Solved by the Invention) The purpose of the present invention is to carry out the reaction in an organic solvent to prevent the decomposition of the active ester by water, and to prevent racemization by the base, under near-neutral reaction conditions. This is to bring out the basicity of the amine.

Another object of the present invention is to provide a method that satisfies the above conditions without protecting α-amino acids in order to further expand the range of applications.

(Unfavorable Means to Solve the Problems) As a method for neutralizing the carboxyl groups of free amino acids, bringing out the basicity of the amine groups, and solubilizing them in organic solvents, the present inventors have investigated a method using silylation.

The silylation was solved by using trimethylsilylimidazole.

α-Amino acid and trimethylsilylimidazole from 0℃
When the reaction is carried out in an organic solvent at 30°C, the reaction proceeds quickly and N
-) trimethylsilylaminotrimethylsilyl ester is obtained.

Taking advantage of this, the general formula (2) (wherein R5 is a hydrogen atom, an alkyl group having 1 to 10 carbon atoms,
Alkenyl group having 2 to 6 carbon atoms, phenyl group, 7 to 6 carbon atoms
9 aralkyl group, an alkyl group with 1 to 6 carbon atoms substituted with a protected or unprotected hydroxyl group, an aralgyl group with 7 to 9 carbon atoms substituted with a hydroxyl group of koji or unprotected, lower alkylthio group An α-amino acid having an alkyl group having 2 to 6 carbon atoms substituted with , an alkyl group having 1 to 4 carbon atoms substituted with a protected thiol group, or an indolylmethyl group is mixed with trimethylsilylimidazole in an organic solvent. When treated, the formula (4) (where A is a hydrogen atom, an alkyl group having 1 to 10 carbon atoms!U, an alkenyl group having 112 to 6 carbon atoms, or a phenyl group) is formed.

Aralkyl group having 7 to 9 carbon atoms, alkyl group having 1 to 6 carbon atoms substituted with a protected hydroxyl group, aralkyl group having 7 to 9 carbon atoms substituted with a protected hydroxyl group, substituted with a lower alkylthio group (representing an alkyl group having 2 to 6 carbon atoms, an alkyl group having 1 to 4 carbon atoms substituted with a protected thiol group, or an indolylmethyl group) is obtained. It will be done.

In this step, the reaction temperature is preferably 06 to 30°C.

The reaction solvent is preferably amide, ether, ester or chlorine. The amount of trimethylsilylimidazole used relative to the α-amino acid is determined by the general formula:

The amount that provides (4) is sufficient.

The advantages of using trimethylsilylimidazole to form N-)limethylsilylaminotrimethylsilyl ester are as follows:
The imidazole that appears in the reaction system is a neutral compound that does not participate in the reaction in any way, and there is no fear of racemization.

This means that the active ester can then be added to synthesize the peptide without isolating the N-)limethylsilylamino trimethylsilyl ester.

Regarding the α-amino acid group represented by general formula (2).

equivalent to amine and carboxyl groups, i.e. 2
Although the current situation is best, the dipeptide formation reaction proceeds even with 1 equivalent by using the following reaction conditions. However, it is inevitable that the yield will decrease when the amount is 1 equivalent.

Next, a condensation reaction between N-)rimestersilylaminotrimethylsilyl ester having the general formula (4) and an active ester compound of an amino acid was investigated.

The α-amino acid of the general formula (2) is reacted with trimethylsilylimidazole to form a compound having the general formula (4), and without isolation, the α-amino acid of the general formula (3) [wherein R6 is a hydrogen atom and has 1 to 10 carbon atoms] an alkyl group, an alkenyl group having 2 to 6 carbon atoms, and a phenyl group.

Aralkyl group having 7 to 9 carbon atoms, alkyl group having 1 to 6 carbon atoms substituted with a protected hydroxyl group, aralkyl group having 7 to 9 carbon atoms substituted with a protected hydroxyl group, lower aralthio group represents an alkyl group having 2 to 6 carbon atoms substituted with , an alkyl group having 1 to 4 carbon atoms substituted with a thiol group, or an indolylmethyl group.

By adding an amino acid active ester compound having an amine-protecting group or a group (wherein R6 represents the same meaning as above and R8 represents an amino group-retaining group), the yield can be improved. General formula (1
a) A compound of the present invention having the formula (wherein A, R' and R7 have the same meanings as above) is obtained.

The active esters used in this reaction are p-nitrophenyl ester and dinitrophenyl ester.

Phenols such as interchlorphenyl ester and trichlorphenyl ester, esters of N-oxysuccinimide, esters of N-oyacyphthalimibi, etc. may be used, but pyridylthioester is preferable.

In the above novel peptide synthesis method, the general formula (
The amino acid of 2) is suspended in an organic solvent, and then trimethylsilylimidazole is added. As the reaction progresses, the insoluble matter dissolves, forming a translucent solution. For example, when dimethylformamide 1 is used as a solvent, a translucent solution is obtained, and the formation of N-)limethylsilylaminotrimethylsilyl ester can be easily confirmed.

If an excess of trimethylsilylimidazole is used, the reaction will not proceed. This is thought to be due to the formation of an N,N-ditrimethylsilylamino compound having the general formula (5) (wherein A has the same meaning as above). This means that citrimethylsilylimidazole must always be used correctly.

When the carboxyl group of the above-mentioned compound (1a) of the present invention is reacted with a general alkylating agent, the formula (1b) (wherein A,
Other compounds of the present invention are obtained in which R6 and R7 have the same meanings as above, and R9 represents a lower alkyl group.

The alkylating agent used here may be a general alkylating agent such as diazomethane.

In addition, by removing the protecting group of the above-mentioned compound (1a) using a method generally used in ipted chemistry, another compound of the present invention (1c) [where RIO is a hydrogen atom and has 1 to 1 carbon atoms] Alkyl &, alkenyl group having 2 to 6 carbon atoms, phenyl group, aralkyl group having 7 to 9 carbon atoms, alkyl group having 1 to 6 carbon atoms substituted with hydroxyl group, 7 to 9 carbon atoms substituted with hydroxyl group It represents an aralkyl group, an alkyl group having 2 to 6 carbon atoms substituted with a lower alkyl group, an alkyl group having 1 to 4 carbon atoms substituted with a thiol group, or an indo9lylmethyl group.

R11 is a hydrogen atom or basis (Here, RIO has the same meaning as above)] The cyptide of the present invention having the following can be obtained.

To summarize the production method of the present invention, first, trimethylsilylimidazole is added to the amino acid to produce N-)limethylsilylamino trimethylsilyl ester of the amino acid.

What is separated here is imidazole, which does not participate in the reaction. Then the active ester (pyridylthioester) is added.

What separates is 2-pyridinethiol.

Under such reaction conditions, there is no need to add a catalyst such as a weak base. There is also no need to isolate silylated amino acids as intermediates.

The compounds present in the reaction system are completely neutral, so there is no need to worry about racemization. The reaction conditions are preferably room temperature or lower, and since the reaction proceeds between room temperature and -20°C, temperature control is not necessary.

After the reaction is complete, diimidazole is removed by washing with water.

2-pyridinethiol can be removed.

Thereby, after straining, it can be easily purified by recrystallization, column chromatography, preparative thin layer chromatography, etc.

It is thought that high yield can be determined quantitatively by considering the type of amino acid, solvent, reaction time, etc.

Using the method of the present invention, it is theoretically possible to extend the takuptedo chain as long as the solvent is compatible.

(Examples) The present invention will be explained in more detail below using Examples and Reference Examples, but it goes without saying that the present invention is not limited to these Examples.

Example I N-(benzyloxycarbonyl)-L-valyl-2-
Trimethylsilylimidazole (amino-4-) was added to a suspension of L-allylglycune ntthenate (230η) in dimethylformamide (5 ml).
560w9) was added at room temperature under a nitrogen stream and stirred for 1 hour.

Next, a dimethylformamide solution (3+t/) of N-penzyloxycarbonylnoline pyridylthioester (550 .mu.) was added at room temperature and stirred for 2 hours.

The reaction solution was poured into water and adjusted to pH 2 with IN hydrochloric acid.

After stirring for 30 minutes, extraction was performed three times with ethyl acetate (200 liters), and the organic layer was washed with saturated brine, dried, and concentrated to obtain 500 w1 of crude crystals (yield: 88%).

Recrystallized from chloroform/hexane to give 467cm (yield: 81
%) of the title compound was obtained.

In Example 1, it was found that trimethylsilyl igudazole had the optimum ratio of 2 to 2 according to the following experimental example.

Further, in order to examine the influence of the solvent, an experiment was conducted using the same method as in Example 1 using the following solvents.

j) N-(indylox7carbonyl)- by dissolving the title compound in ether and adding excess diazomethane.
L-noζly-2-amino-4-pentenoic acid methyl ester.

The physical properties of methyl ester are shown below.

Melting temperature: 116.5-117.0°C [α) D: +12.9° (C-1,01, chloroform)
): 3450° 1740.172 (1,1680 Mass spectrum (m/z): 363 (M+1)", 25
4°NMR spectrum (CD (J3. δ): 0.8
7 (3H, d.

J-7,0H2), 0.93 (3H,d, J=7.0
Hz).

2.06 (IH, m), 2.48 (2H, t, 'J
-7,0Hz).

3.68 (3H, s), 4.00 (IH, drl, J=
6゜8.5 HZ) + 460 (I H, l
ty J-6, 8, 5Hz) #5.04 (2H,
e), 6.46 (1 horse d, J=8.5).

7.28 (5H, s) Example 2 N-(benzyloxycarbonyl)-valyl-L-methionine Trimethylsilylimidazole (176 μl) was added to a dimethylformamide suspension (2 mJ) of L-methionine (901 ng) at room temperature under a nitrogen atmosphere. was added and stirred for 2 hours.

Next, a dimethylformamide solution (2 m/) of N-benzyloxycarbonyl-L-valine pyridylthioester (170111v) was added and stirred for 14 hours.

Add 3 ml of IN hydrochloric acid to stop the reaction, extract ethyl acetate,
The layer was treated in the same manner as in Example 1 to obtain the title compound 168cm (yield: 89cm) as white crystals.

Melting point: 180-182° [α] D knee 22.6° (Cm 1.05. Chloroform) IR spectrum (nujol, cm ): 3350
゜3300.1740.1640 Mass spectrum (m/z): 276.203,183N
MR spectrum (CD30D, δ): 0.96 (3H,
d.

J=7.0Hz), 0.99(3B, (1,J-7,0
Hz).

2.05 (3H,e), 3.96 (IH,d
, J=8.0H2).

4,'58 (IH, aa, J=4.8H2), 5.0
8 (2H.

8), 7.32 (5H, B).

Example 3 Suspension of N-(-<ndyloxycarbonyl)-valyl-L-phenylalanine methyl ester L-phenylalanine (99■) in dimethylformamide (1 m/), ) rimethersilylimidazole (
176μ and N-benzyloxycarbonyl-L-
2-pyridylthioester of valine (1671111>
) was reacted in the same manner as in Example 2 using a dimethylformamide solution (ljIe), and the resulting crude crystals were dissolved in 5++/ml ethyl acetate and an excess of diazomethane in ether was added to give a methyl ester. .

After concentration, the residue was subjected to silica gel column chromatography and eluted with ether/hexane (25 nia 5) to give the title compound 171η (yield: 86 H) as white crystals.

Melting point: 142-143°C [α)D: +36.6° (C-1,1, chloroform]
Mass spectrum (m/z): 412 (M"), 304°R spectrum (chloroform, cm-"): 3
450゜3350.1750, 1740゜1730, 168O NMR shot (CDOA!3. δ): o, 57
(3H.

d, J-6,0H2), 0.93(3H, (1,
J=6.0H2)*2.08 (I H,q, J-
6Hg), 3.10 (2H,d.

J-6Hz), 3.72 (3H,s)*4
．． 00 (I H', d d.

J-6* 9 Hz) + 488 (I H* d
t, J-6r 8 Hzλ5.10 (2H,s
), 5.34 (I H, d, J-9Hz).

6.36 (IH, (1,, J-8H2), 7.36 (5
H, a).

7.0-7.4 (5H,m) Example 4 A suspension of N,-(benzyloxycarbonyl)valylglycyglycine (100 mg) in dimethylformamide (1
me),)limethylsilylimidazole (390 μl)
and N-benzyloxycarbonyl-L-valine pyridylthioester (381yzz) in dimethylformamide 4 solution (1, (1 mJ)) in the same manner as in Example 1, and reacted at room temperature for 16 hours. The oily substance obtained by the same treatment was dissolved in ethyl acetate, and an ether solution of diazomethane was added to give a methyl ester.This methyl ester was subjected to silica gel column chromatography (elution solvent: ether) to obtain the title compound of 3771119 as a white ester. Obtained as crystals (yield: 97 cm).

Melting point: 159.5-161.5°C [α) p knee 13.1° (C-1,16, methanol)
Laboratories (Chloroform, cm): 3450
゜1750.1740,1720゜Mass Haktor (m/z) * 322 (M”
) * 255 +NMR spectrum (CDC13,
δ): 0.94 (3H, (1°J=7Hz), 0
．． 98 (3H, d, J-7H2), 2.05 (
I H, q, J-7H2), 3.72 (3H
, 8) *3.90-4.22 (3H, m), 5.
08 (2H, 8)*5.58 (I H, cl,
J-9H2) + 6.80 (I H.

aa, J-5,6Hz), 7.32(5H,s
) Example 5 N-(benzyloxycarbonyl)-L-paryl-L-
Threonine Methyl Ester H To a suspension of L-threonine (23.81 ng) in dimethylformamide (0.5 ng) was added dropwise trimethylsilylimidazole (88 μl) under a stream of pure atom at room temperature and stirred to obtain a translucent solution. Next, N-benzyloxycarbonyl-L
A dimethylformamide solution of -valine pyridyl thioester (68119) was added dropwise and treated in the same manner as in Example 4. The resulting oil was treated with excess diazomethane ether solution. The crude product was subjected to silica gel column chromatography to obtain the title compound as white crystals. Collection 41
29.5■, yield 41%.

Melting point: 141-142°C [α] D knee 8.3° (Ct, O, chloroform) mass lactate (m/z): 366 (M”), 322IR
Spectrum (react form, cm-”): 345
0°1750.1730,1710°68O NMR spectrum (CDC13, δ): 0.93 (3H
,d.

J-7H2), 0.96 (3H, d, J-7Hz
), 1.16 (3H, d, J-7Hz), 3.70
(3H, s).

4.05 (IH, dd, J-7, 9H2), 4
．． 58 (IH.

ad, J-2,5,8Hz), 5.05(2H,s
), 5.70 (IH, eL, J=8Hz), 7
．． 11 (IH, d, J=9Hz), 7.30 (
5H,e) Example 6 N-(benzyloxycarbonyl)-L-valyl-0-
(1-Butyldimethylsilyl)-L-threonine methyl ester 0-t-butyldimethylsilyl-L-threonine 38■
N-indyloxycarbonyl-L-valine pyridylthioester 56 was added to a clear solution obtained by stirring for 30 minutes using a suspension of 100% in dimethylformamide (0.5 ml) and 49 µm of trimethylsilylimidazole.
1v of dimethylformamide (0.5m/) i solution was added dropwise and reacted at room temperature for 3 hours.

2rxl of 0.5N hydrochloric acid was added, and the mixture was extracted with ethyl acetate and treated in a conventional manner. To the crude product obtained, an excess of diazomethane in ether was added to esterify the mixture. This esterified product was subjected to silica gel column chromatography and eluted with yin-yin/ethyl acetate (3:1) to yield the title compound 55 (
Yield: 7o%) was obtained as an oil.

[α) D: + 4.31° (C-1,16, chloroform) -r x , X-! : Cuttle (m/z): 48
1(M+1)”.

423.315 IR Structur (Film # clIL-”): 34
50°3300.1725,167O NMR speed! ! ri) le (CDC1a e a)
: o, OO(3H2s), 0.06(3H,s),
0.85 (9H, a).

1.00 (3H, d, J-7H2), 1.03
(3H, d.

J-7H2), 1.17 (3H, d, J-7H2), 2
．． 16 (xnt qI LT-7Hz), 3.7
0 (3H, 8).

4.13 (I H, ad, J=6.8Hs),
4.35-4.60 (2 Hlm) -5,12 (2
Ht a ) * 5.50 (I H-d, J-
8Hz), 6.36 (IH, d, J-8Hz).

7.35(5H,s) Example 7 N-(t-nodoxycarbonyl)-2-amino-4-R
Enthenoyl 0-(t-butyldimethylsilyl)-L-
Threonine methyl ester number C00C(CH3)3 0ft-butyldimethylsilylthreonine (953■
4.0 mmol) in dimethylformamide solution (8,
0m! Trimethylsilylimidazole (1,17m
18.0 mmol) was added and stirred for 1 hour at room temperature under nitrogen atmosphere, resulting in a clear solution.

To this solution, a methylene chloride solution (1 (1 m/)) of N-t-phthoxycarbonylallylglycine thioviridyl ester (1,05N, 3.4 mmol) was slowly added dropwise under stirring at room temperature. The mixture was stirred overnight, poured into water, adjusted to pH 3 with a 10% aqueous citric acid solution, and extracted several times with ethyl acetate.The organic layer was dried over anhydrous magnesium sulfate, filtered, and the solvent was distilled off under reduced pressure. A reaction mixture of a yellow oil was obtained.(2,25Ji') A portion (414■) of this reaction mixture was dissolved in ether,
After adding the diazomethane ether solution.

The solution was distilled off, and the resulting colorless oil was subjected to column chromatography to obtain ether/hexane-1/1)N-t.
-Butoxycarbonylallylglycine-〇 -t--
7-Tyldimethylsilylthreonine methyl ester was obtained. (252w9 Yield 95cm) Properties: Colorless oily substance IRx vector (film # “”-”): 34
50 *3335.3080.1?60°1720.1685 mass/%! 'Ktor (m/g): 44s (M+1)''
, 400° NMR spectrum (CDCJ3. δ) knee 0.04 (3H.

8L 0.02 (3H, s), 0.83 (9H, s)
.

1-14 (3H26p J-70Hta), 1
-42 (9Hs s ) s2.52 (2H=
m)s 3,68 (3H,s)s 4.1
~46 (3H9m) +5-0~5-3 (3
H2m)*5180'(I H-ddt, J=
18.0, 10.0, 7.0H2), 6.67
(IH, d,, 10.0 &) Example 8 N-(t-nodoquincarbonyl)-2-amino-4-pentenoyl-0-(t-butyldimethylsilyl)-L-
Trimethylsilylimidazole (0.65 mJ 4.4 mmol) was added to a dimethylformamide solution (5 MI) of threonyl-2-amino-pentenoic acid methyl ester allylgrin (255w 92.2 mmol),
The mixture was stirred at room temperature for 1 hour to obtain a clear solution. Add N to this solution.
-t-putoky7carbonylallylglycine-0-t-,
/ methyldimethylsilylthreonine thiopyridyl ester (9661v 1.85 mmol) in methylene chloride solution (8d) under a nitrogen stream.

The mixture was added dropwise at room temperature, and then stirred overnight. This reaction solution was poured into water, adjusted to pH 3 with 10 citric acid aqueous solution, and then extracted several times with ethyl acetate. The organic layer was dried over anhydrous magnesium sulfate, filtered, and the solvent was distilled off under reduced pressure to obtain a yellow oil.

An ether solution (10 ml) of this product was mixed with a diazomethane ether solution under cooling with water at θ°C, the solvent was distilled off, and the colorless oil obtained was subjected to column chromatography (ether/hexane-1/ 1) to obtain the title compound 875 (if', rate 88).

Properties: Colorless crystal Melting point: 135-136.5°C (recrystallization solvent: hexane)
IH xd (m/z): 542 (M+1
) +, 497° NMR spectrum (CI) C13, δ)
:0.14(θ53HLO, 17(θy3H)
s O,93(9H+θ),1.12(FH,a
, J-6,0H2), 1.45 (9H,s).

2.53 (4H, m), 3.74 (3H, s), 4.1
9 (IH, d, J-6,0Hz), 4.36 (
IH, ad, , T-7,0,3,0H2), 4
．． 41 (I H, brs), 4.65 (I Hp
q * J+=7.0 H2)s 5.00 (
IH,brs) C5,1~5.2 (4H,m)
, 5.6-5.8 (2H, m).

7.02 (] H, d, J-6, 0H2),
7.29 (I H, d.

J-7, aH2).

This compound (Example 8) was hydrolyzed with a 0.5 N aqueous sodium hydroxide solution to give pN-(t-butquincarbonyl)-2-amino-4--! ntenoirou0-(
t-methyldimethylsilyl)-L-threonyl-2-amine gave ntthenic acid.

Properties: Amorphous crystal NMRx vector (100MH2-CD a OD e
δ) = 0.08 (3H, s), 0.10 (θ, 3H
), 0.92 (9H, e), 1.16 (3H, d,, T
-6,0Hys).

1.45 (9H-8) # 2.52 (4He
m) -4.0 to 6.0 (12H, m), 7.
68 (] H, d, J=8.0H2).

7.81 (IH, d, J-8,0Hz).

Example 9 (28,3R)-1'J-(t-butoxycarbonyl)
-2-Amino-3-(tetrahydropyranyl)oxy-4-(t-butyldimethylsilylokinphenyl)-
Pteroyl-0-(t-butyldimethylsilyl)-threonine pyridine thioester 0-(t-butyldimethylsilyl)-threonine 35 #v was suspended in 0.3 xi of anhydrous N,N-dimethylformamide, and 44 μl of trimethylsilylimidazole was suspended in 0.3 xi of anhydrous N,N-dimethylformamide. was added and stirred at room temperature for 1 hour under nitrogen.

A solution of Compound 45.5 of Reference Example 3 in anhydrous dimethylformamide (0.7 ml) was then added to the above silylated product at room temperature, and the mixture was further stirred for 15 hours.
After adjusting the pH to 4 with NtiA acid, the mixture was extracted with ethyl acetate.

The organic layer was dried over anhydrous magnesium sulfate and concentrated to obtain an oil.

This oil was dissolved in 0.511/ml of anhydrous dichloromethane, and 23.8 µm of triphenylphosphine and 16.7 µm of dipyridyl disulfide were added to it at room temperature under a nitrogen stream.
Stir for hours.

The reaction solution was concentrated under reduced pressure, and the resulting oily substance was subjected to silica gel column chromatography using ether/hexane (
1:1) to give the title compound 38911Ig (yield 63
%) was obtained.

Properties of oily substance NMR spectrum (CDC13I δ): 0.01.
0.05 (3H, s each), 0.16 (6H, a), 0.
88 and 0.90 (each 8.9H in total), α95 (9H
, s), 1.14 and 1.20 (each a, aH, J-5H
z), 1.48 (9H.

brs), 6.72 and 673 (each d, 2H, J-
8H2).

7.09 (2H, d, J=8H2), 7.1
6-7.80 (3H, m), 8.59 (IH, a, J-
5H2).

Reference Example I N-(t-nodoxycarbonyl)-2-amino-4-pentenoic acid thiopyridyl ester N-t-butoxycarbonylallylglycine (500
2.3 mmol) of methylene chloride solution (8 lj) was added pyridyl disulfide (51219°2.3 mmol) and then triphenylphosphine (731 mg).
, 2.8 mmol) were added. After stirring overnight at room temperature,
The solvent was distilled off under reduced pressure, and the resulting reaction mixture was subjected to column chromatography using ether/hexane-2/3)N.
-t-Nodoxycarbonylallylglycine thiopyridyl ester was obtained as a pale yellow oil. (650wi, yield 91%) Properties: Pale yellow oil NMR spectrum (100MH2, CDCl3. δ
): 1.44 (9H, s), 2.54 (2H,
m), 4.48 (I H, dt, J-7,0,
7,5H2), 5.0-5.3(3H,m),
5.70 (IH, aat, J-18, 0°9.0.
7.0H2), 7.23(IH, ddd, t
Tx7. o.

5.0. 2.0H2), 7.54 (IH, aa
a, J-8,0°2.0. t, oHz)+7.
70 (IH, ddd, , r-B, 0°7.0. 2.0
H2), 8.56 (I H, aaa, J-5
,0°2.0. 1.0H2).

Reference Example 2 N-(t-nodoxycarbonyl)-2-amino-4-intenoyl-0-(t-butyldimethylsilyl)-L-
Threonine thiopyridyl ester N obtained in Example 7
-t-Butoxycarbonylallylglycine-〇-t-Butyldimethylsilyl-L-threonine (1,3#) in methylene chloride solution (Bat) was added with dipyridyl sulfide (440ml).

Next, triphenylphosphine (629 cm) was added, and the mixture was stirred at room temperature for 4 hours. After distilling off the solvent under reduced pressure, it was subjected to column chromatography to obtain ether/hexane-2/3.
) N-t-nodoxycarbonylallylglycine-〇-t
-Notyldimethylsilylthreonine thiopyridyl ester 842cm (yield: 82cm) was obtained as a pale yellow oil.

Properties: Pale yellow oily substance NMR spectrum (60MHz, CDCl3.δ
) knee-0,04(3H,θ)t O,02(3H,
s), 0.85(9H,s), 1.11
(3H,a, J-7,0H2).

1.40 (9H, e), 2.58 (2H, m), 4.0
-6.0 (7H, m), 6.6~8.6 (5H
, mL Reference Example 3 (28,3R)-4-(4-t-butyldimethylsilyloxyphenyl)-3-(2-te),>human90pyranyl)oxy-2-N-t-nodoxycarbonylamino Butyric acid-2-pyridylthiol ester (2Ss 3R)-
4-(4-t-butyldimethyl7lyloxyphenyl)-
3-(2-tetrahydropyranyl)oxy-2-N-butoxycarbonylaminobutyric acid 1.03# was dissolved in anhydrous dichloromethane 10ffi/triphenylphosphine 954■ and dipyridyl sulfide under a stream at 40°C for 8.5 hours. and stirred at room temperature for 14 hours.

The reaction solution was concentrated and subjected to silica gel column chromatography and eluted with hexane:ether (1:1) to obtain the title compound 846 (yield: 69%).

Properties: Oily substance IR spectrum (film, cWL): 3450° Mass spectrum (m/z): 464 HMR spectrum (CDC13, δ): 0.16 (6H, θ).

o, 96 (9Ht s ) t 1.54 (9H
-8), 6.74 and 706 (each 2H, a, J-g
Hz), 7.24 (IH.

m), 7.44-7.80 (2H, m), 8
．． 52 (IH.

m). (Effects of the invention) Peptides can be synthesized by using the method of the present invention. The special feature of this method is the use of an organic solvent, and it is thought that theoretically any polymeric peptide can be synthesized as long as there is a solvent that can solubilize it. Since the peptide obtained by the method of the present invention is a basic substance that can be applied in a wide range of fields such as hormones, agricultural chemicals, and foods, the method of the present invention, which can efficiently produce peptides, will be of great benefit to this field. Probably.

(5 other people) QQ Chi-

Claims (1)

[Claims] 1. General formula (1) ▲ Numerical formulas, chemical formulas, tables, etc. ▼ (1) [In the formula, R^1 is a hydrogen atom, an alkyl group having 1 to 10 carbon atoms, or a carbon number 2 to 6 alkenyl group, phenyl group, carbon number 7
~9 aralkyl group, a C1-6 alkyl group substituted with a protected or unprotected hydroxyl group, a C7-9 aralkyl group substituted with a protected or unprotected hydroxyl group,
Represents an alkyl group having 2 to 6 carbon atoms substituted with a lower alkylthio group, an alkyl group having 1 to 4 carbon atoms substituted with a protected or unprotected thiol group, or an indolylmethyl group, R^2 is a hydrogen atom, Protecting group or group for amino group ▲ Formula,
There are chemical formulas, tables, etc. ▼ (where R^1 represents the same meaning as above, R^4 represents a hydrogen atom or a protecting group for an amino group), and R^3 represents a hydrogen atom or a lower alkyl group. A method for producing a peptide having the general formula (2) ▲ Numerical formula, chemical formula, table, etc. ▼ (2) (wherein R^5 is a hydrogen atom, an alkyl group having 1 to 10 carbon atoms, Alkenyl group having 2 to 6 carbon atoms, phenyl group, 7 carbon atoms
~9 aralkyl group, a C1-6 alkyl group substituted with a protected or unprotected hydroxyl group, a C7-9 aralkyl group substituted with a protected or unprotected hydroxyl group,
Alkyl group having 2 to 6 carbon atoms substituted with lower alkylthio group, 1 to 4 carbon atoms substituted with protected thiol group
An α-amino acid having an alkyl group or an indolylmethyl group) is reacted with trialkylsilylimidazole, and then the general formula (3) ▲There are mathematical formulas, chemical formulas, tables, etc.▼(3) [In the formula, R^6 is hydrogen atom, alkyl group having 1 to 10 carbon atoms, alkenyl group having 2 to 6 carbon atoms, phenyl group, 7 carbon atoms
-9 aralkyl group, alkyl group having 1 to 6 carbon atoms substituted with a protected hydroxyl group, aralkyl group having 7 to 9 carbon atoms substituted with a protected hydroxyl group, carbon substituted with a lower alkylthio group Represents an alkyl group of 2 to 6 carbon atoms, an alkyl group of 1 to 4 carbon atoms substituted with a protected thiol group, or an indolylmethyl group, R^7 is a protecting group or group for an amino group ▲ Numerical formula, chemical formula, table etc. ▼ (Here, R^6 represents the same meaning as above, and R^8 represents a protecting group for the amino group) An active ester compound having the following is reacted in an organic solvent, or deprotected after the reaction. (2) A method characterized in that the trialkylsilylimidazole is t-butyldimethylsilylimidazole, trimethylsilylimidazole, triethylsilylimidazole, dimethylphenylsilylimidazole, or t-butyldiphenylsilylimidazole. (3) The method according to claim 1, wherein the solvent used in the peptide formation reaction is an amide solvent, an ether solvent, or a chlorine solvent. ) The reaction temperature in the peptide formation method is 30 to 0°C.
The method according to claim 1. (5) The method according to claim 1, wherein the amount of trialkylsilylimidazole used in the reaction is 1 to 2 equivalents relative to the α-amino acid group of formula (2). (6) The method according to claim 1, wherein the formulas (2) and (3) are optically active substances. (7) The method according to claim 1, wherein the formula (1) is an optically active substance.