JPH0328418B2 - - Google Patents

Description

[Detailed description of the invention] The present invention is useful as a converting enzyme inhibitor and as an antihypertensive agent.
Carboxyalkyl dipeptides useful as pressure agents
This invention relates to aza analogues of derivatives. Main departure
The general structural formula of light compounds is: [In the ceremony: R and R₃are independently hydrogen, lower alkyl, ara
is alkyl; R₁is aaralkyl, heteroaralkyl,
Substituted aralkyl, substituted heteroaralkyl (aryl)
The substituents of aryl and heteroaryl are independently
halo, amino lower alkyl, hydroxy, lower
alkoxy, lower alkyl and 1-8 carbon atoms
alkyl, and amino, acylamino, hetero
Arylamino, aryloxy, heteroary
ruoxy, arylthio, heteroarylthio,
1-8 carbon atoms substituted by hydroxyl
is an alkyl); R₂is hydrogen, lower alkyl, amino lower alkyl
ru, araalkyl; A is hydrogen, cycloalkyl having 5-7 carbon atoms
aryl, heteroaryl; B is hydrogen, lower alkyl; or A and B are N-CHCO₂R₃and structural formula:
[Formula] and [Formula] (In the formula: R₃is defined above; Q grouped together₁and Q₂is CH₂CH₂,CH₂
-S,R_Fouris aryl or hydroxyl, amino
Lower alkyl, amino, halo, alkoxy
CHR is an aryl substituted with_FourS, R_Fiveis hydrogen,
lower alkyl, aryl, aralkyl,
CONR₆R₇(R₆and R₇are independently hydrogen, lower alkyl
CH is₂-CH-
OR_Fiveselected from; W is a bond or CH₂And; Y is a bond, CH₂,CH₂CH₂) with
[can be combined to form a structure], and its pharmaceutical
Indicated by acceptable salts. Preferred structural formulas include: R and R₃are independently hydrogen, lower alkyl, ara
is alkyl; R₁is an ara in which the alkyl group contains 1 to 4 carbon atoms.
alkyl, heteroaralkyl; alkyl group is carbon
1-4 substituents including halo, amino, amino
Lower alkyl, hydroxy, substituted alkyl (carbon
1 to 6 substituents, amino, arylamino,
Aryloxy, alkylthio, arylthio,
heteroarylthio, hydroxy)
Substituted aralkyl, substituted heteroaralkyl
the law of nature; R₂is hydrogen, lower alkyl, amino lower alkyl
is le; A is cycloalkyl having 5-7 carbon atoms.
the law of nature; B is hydrogen; or A and B are NCHCO₂R₃and structural formula: (In the formula: R₃is defined above; Q grouped together₁and Q₂is CH₂CH₂，
CH₂S, R_Fourby aryl or hydroxyl
CHR is a substituted aryl_FourS, R_Fiveis a lower grade
CH that is alkyl, araalkyl₂CH-OR_FiveAnd
Compounds that can be bonded by forming a ring structure with
It is. More preferred structural formulas are: R and R₃are independently hydrogen, lower alkyl, ara
is alkyl; R₁is an ara in which the alkyl group contains 1 to 4 carbon atoms.
alkyl, heteroaralkyl; alkyl group is carbon
1-4 substituents including halo, hydroxy,
Substituted alkyl (1-6 carbon atoms, substituent is aryl)
oxy or arylthio)
laalkyl, substituted heteroaralkyl; R₂is hydrogen, lower alkyl, amino lower alkyl
is le; A and B are NCH-CO₂R₃and structural formula: (In the formula: R₃is defined above; Q grouped together₁and Q₂is CH₂CH₂，
CH₂S, R_Fourby aryl or hydroxyl
CHR is a substituted aryl_FourS, R_Fiveis a lower grade
CH that is Lukiru₂CH-OR_Five) A compound that is bonded to form a ring structure with
be. The most preferred structural formula is: R is hydrogen or lower alkyl; R₁is an araa in which the alkyl group contains 1 to 3 carbon atoms.
alkyl, heteroaralkyl; alkyl group is carbon
1-3 substituents including halo, hydroxy, substituted
Substituted alkyl (1-6 carbon atoms and aryl substituent)
oxy or arylthio)
Raalkyl; R₂is hydrogen, methyl, 4-aminobutyl
the law of nature; R₃is hydrogen; A and B are NCHCO₂R₃and structural formula: (In the formula, R₃is defined above)
bond by forming a ring structure with
Phosphorus, thiaproline, 4-carboxy-2-(2
-Hydroxyphenyl)thiazolidine
It is a thing. Preferred, more preferred, most preferred compounds are also
Contains pharmaceutically acceptable salts of. Unless otherwise noted, the values shown are subject to change.
The lower alkyl group is a straight chain or branched chain with 1 to 6 carbon atoms.
chains, saturated and unsaturated hydrocarbon groups: e.g.
thyl, ethyl, propyl, isopropyl, buty
isobutyl, t-butyl, pentyl, isopeyl
ethyl, hexyl, vinyl, allyl, butenyl
What is it? Subject to change unless otherwise noted.
The above aralkyl group shown in
have 1-6 carbon atoms, such as benzyl,
Contains netyl, cinnamoyl, etc. halo is black
It means bro, bromo, iodine, and fluoro. especially
Any of the aryl groups shown except as noted
Represented by phenyl, naphthyl, biphenyl
Ru. The heteroaryl groups shown are, for example, pyridyl,
Thienyl, frill, indolyl, benzthienyl
Contains imidazolyl, imidazolyl, and thiazolyl. Acyl group
indicates lower alkanoyl and aroyl groups. Structural formula () and preferred subgroup compounds are as follows:
Reaction route diagram (R, R unless otherwise specified)₁,R₂，
R₃, A, B, W, Y are defined above.
one or more methods and submethods shown in
It can be manufactured by As shown in the reaction route diagram, the aza of the present invention
Analog compounds are generally imino acids (A≠H).
t-Butyl ester and phosgene in methylene chloride
By reacting in the presence of triethylamine, N
- Manufactured by obtaining chlorocarbonyl derivatives
be done. This intermediate was dissolved in triethyl chloride in methylene chloride.
For example, isopropyl or N-
Isopropyl-N'-carbobenzyloxyhydride
Razin [C.J.Gray et al.
Tetrahedron, 33 volumes, 837-40
(1977) according to known methods.
for example, isolation, filtration,
Condensation yields a protected intermediate. ethanol
hydrogen in the presence of 10% palladium-activated carbon catalyst in a
to obtain a deprotected intermediate derivative.
Ru. Alternatively, triethyl in methylene chloride
React with substituted hydrazine in the presence of amine to obtain
Ru. t-Butyl ester of amino acid (A=H)
When using phosgene, carbonyl diimide
acylimidazole intermediate by replacing sol with
This was reacted as described above to give N-chlorocarboxylic acid.
obtain the nil derivative. Tetrahydrof derivative
α-keto ester or
α-Ketoester-carbazone by reaction with acid
obtained, cyanoborohydride in ethanol
α-aza-t-butyl ester peptide
A tide derivative is obtained. Trifuses peptide derivatives
T-butyl ester is reacted with fluoroacetic acid (TFA).
The tel group is removed to obtain a. When R is other than hydrogen
R₃X (X = bromo, iodine) and cesium hydroxide
It is introduced using a base such as Protecting group is present
If so, removing it will yield a compound with the structural formula. of
Bis-esters are available from Fisher.
Stellification, e.g., a compound with structural formula at room temperature
Obtained by treatment with dry HCl in alcohol
I can do it. Alternatively, R₂If =H, the compound with the structural formula
can be obtained as shown in the reaction diagram.
can. This method uses α-hydrazinoic acid or
Synthesis of tels X, then in N-chlorocarbonyl
This includes condensation with intermediate bodies. if you need it
for example, by subsequent esterification and deprotection under normal conditions.
Structural formula (R₂=H) is obtained. obvious to those skilled in this field.
As shown in the example below, and as shown in the examples below,
Amino, carboxy, hydroxy
Reactive residues such as syl are
Protected and reversed by the usual methods in petido chemistry.
The desired compound is obtained by subsequent deprotection.
be able to. The protection of reactive residues used during condensation reactions is an analogy.
N-formyl, N-t-butoxycarbonyl,
known methods such as N-carbobenzyloxy
By including and then removing them we get ()
Ru. Furthermore, R and R₃The residues are benzyl, ethyl
removable ester residues such as
can include. The starting materials required for the process of the invention may be known in the literature.
or prepared by a known method from known starting materials.
I can do it. The above syntheses can be performed using racemic or paired compounds as starting materials.
Can use palm body. diastema depending on the synthesis method.
If a rheomeric product is obtained, the diastereomeric
The product of the mer can be obtained by chromatography or by fractional crystallization.
It can be separated by method. intermediate or final
If optical resolution of the final product is required, an optically active acid
or by forming a salt with a base and crystallizing it.
can be achieved. Many of the compounds of this invention are available in a variety of inorganic and organic salts.
forming salts with groups, which are also within the scope of the invention.
It will be done. The cationic salts include sodium salt, potassium salt,
Alkali metal salts like salt, calcium salts, mag
Alkaline earth metal salts such as nesium salts,
lohexylamine salt, N-methyl-D-glucami
salts with organic bases such as arginine, lysine
including salts with amino acids such as R₁If
is R₂The product containing a basic functional group in the male
Organic and inorganic acids and salts such as inic acid, hydrochloric acid, etc.
form. Non-toxic and pharmacologically acceptable salts
Although suitable, other salts may also be used to isolate the product, e.g.
Or it is useful as in purification. Salts are prepared using conventional methods in solvents or media in which the salts are insoluble.
Remove in a vacuum or under reduced pressure or by freeze-drying.
or the cation of the salt present on a suitable ion exchange resin.
A solution such as water that can exchange cations for other cations.
1 equivalent of the free acid or free base of the product in the medium.
React with one or more amounts of a suitable base or acid.
It can be formed by making it correspond. The compounds of the present invention are angiotensin converting enzymes.
Inhibits decapeptide, angiotensin
It interferes with the conversion of angiotensin to angiotensin. Ann
Geotensin is a powerful blood pressure enhancer. blood
The decrease in pressure is associated with angiotensile stress, especially in animals and humans.
Inhibition of biosynthesis in high blood pressure caused by
It depends. Additionally, converting enzymes can lower vascular blood pressure.
Decomposes bradykinin, which is a sub-substance. Therefore,
bradykini is an angiotensin-converting enzyme inhibitor
It also lowers blood pressure by maintaining the effectiveness of drugs.
The relative importance of these and other possible mechanisms has been established.
However, angiotensin-converting enzyme
The natural inhibitor is effective in various animal models.
It is an antihypertensive agent, and is used to treat malignant and essential diseases such as renal vascular disease.
clinically effective in many human patients with hypertensive hypertension.
be. For example, D.D.
(D.W.Cushman) et al., Biochemistry E.
(Biochemistry), vol. 16, p. 5484 (1977).
Yo. Evaluation of converting enzyme inhibitors involves in vitro enzyme inhibition tests.
Guided by established law. As an example, a valid method
Y. Piquilloud, E.
- A. Reinharz and M. B
M.Roth, Biochem, Biophys.Acta, 206
volume, page 136 (1970).
Lebobenzyloxyphenylalanyl histidine
This test measures the hydrolysis of leleucine. Living
For in-body evaluation, for example, J. R. Ui
J.R. Weeks, G.A.
J.A.Jones, Proc.Soc.Exp.Biol, Med, 104
Volume, 646 pages (1960)
In normotensive rats stimulated with
vinegar. S. Koletsky et al., Proc. Soc.
Technology of Exp.Biol.Med., vol. 125, p. 96 (1967)
by high renin rat models such as . Compounds of the invention may be used in hypertensive mammals, including humans.
Useful as an antihypertensive agent in the treatment of orally administered
Compositions like tablets, capsules and elixirs for
into a sterile solution or suspension for parenteral administration.
It can be used to achieve lower blood pressure by
Can be used. Compounds of the present invention require such treatment.
patients (animals and humans) with at least a pharmaceutical
An amount that provides a beneficial effect can be administered. Dosage depends on the severity of the disease, the patient's weight, and the field in question.
Varies depending on other factors recognized by the skilled person
However, the dosage range is generally 1 kg of body weight per day.
The amount is approximately 0.5 to 50 mg. This dosage range can be adjusted with unit ingredients if necessary.
It can be divided into daily doses. Current
However, the dosage depends on the severity of the illness, concurrently administered drugs, and the
may vary depending on other factors recognized by those skilled in the field.
become The compounds of the invention may also be used as diuretics or other
Can be administered in combination with antihypertensive agents. Typically 1
In the case of single administration, the daily dosage of individual drugs is
Can be recommended from 1/5 of the minimum recommended dose
range up to the maximum dose. These combinations
To explain, in the range of 5-500 milligrams per day.
One of the clinically effective antihypertensive agents of the present invention is as follows:
Antihypertensive drugs and diuretics listed above and 1-500 mg per day.
Can be effectively combined within the regram range.
Wear. The combined drugs and their daily doses are shown below: Hydrochlorothiazide (10-200mg), Timorrow
(5-60mg), methyldopa (65-2000mg),
Childopa pivaloyloxyethyl ester
(30−1000mg), indacrinone and various ratios of
Its enantiomer (25-150mg), (-)-4-{3-{[2
-(1-hydroxycyclohexyl)-ethyl]-
4-oxo-2-thiazolidinylpropyl
Zozoic acid (10-100 mg). Additionally, hydrochlorothiazide (15-200 mg)
amiloride (5-20 mg) and the converting enzyme inhibitor of the present invention.
Harmful agent (1-500mg) or hydrochlorothiazide
(15-200mg) and timolol (5-50mg) and
Three types of Ming converting enzyme inhibitors (1-500mg)
The combination is also effective in controlling blood pressure in hypertensive patients.
It's a combination. The above dosage ranges are based on unit ingredients if necessary.
It can be divided into daily doses. Ma
In addition, the dosage depends on the severity of the disease, the patient's weight, and the field of study.
may vary depending on other factors recognized by those skilled in
Ru. Typically, the compounds of the invention are as shown below.
can be formulated into pharmaceutical compositions. Structural formula or a pharmacologically acceptable salt thereof
Approximately 5 to 500 mg of the compound or compounds are permitted.
pharmacologically acceptable excipients necessary for pharmaceutical science
Excipients, carriers, binders, preservatives, stabilizers, flavorings, etc.
and mix into unit dosage form. These compositions
The amount of active substance in the formulation or combination should be adjusted within the indicated range.
It is designed so that a suitable dosage can be obtained. Examples of adjuvants added to tablets, capsules, etc. are as follows:
As shown in: Gum tragacanth, Arabia
Bonds like gum, corn starch, and gelatin
agent; excipients such as microcrystalline cellulose; corn
Rokoshi starch, pre-gelatinized starch,
Disintegrants such as lugic acid, stearic acid mag
Lubricants like nesium, sugar, and saccharin
Una sweetener, peppermint, wintergreen oil (cherry)
flavoring agent such as oil), unit dosage form is capsule
In addition to the above types of substances, fatty acids such as
A liquid carrier may be included. various other things
quality as a coating or in the physical form of the dosage unit.
It can be included if it is changed. For example, tablet
The agent may be coated with cilantro, sugar, or both.
I can do it. Syrup or elixir is an active substance
sugar as a sweetener, methyl and preservatives
Propylparaben, color, cherry or orange flavor
Contains flavoring agents such as. Sterile compositions for injection are active according to conventional pharmaceutical sciences
Substances for injection with carriers like water, sesame oil, coconut
Natural botanicals like tree oil, peanut oil, cottonseed oil, etc.
Synthetic oils, such as ethyl oleate, etc.
Can be formulated dissolved or suspended in a fatty carrier
can. Buffers, preservatives, antioxidants as required
etc. can be added. The following examples illustrate the invention:
It consists of particularly preferred embodiments. these
Preferred diastereomers of the examples are R₁as well as
C.O.₂R₃Natural L- at the two carbons to which is bonded
Although it has the configuration of amino acids, the column
Isolated by chromatography or fractional crystallization
It will be done. In most cases, the preferred absolute placement of these is
It is also indicated by (S)-. In the examples below, the NMR chemical shift is
from the internal standard tetramethylsilane to the low magnetic field side.
It is expressed in ppm. Optical rotation is measured in methanol solution
did. Example 1 α-Azaalanyl-(L)-proline t-butyl
ester(2) Proline t-butyl ester (10.0g; 0.058
mol) and triethylamine (8.56 ml; 6.22 g;
0.0614 mol) of dehydrated THF solution (60 ml)
(12.5%; 0.070 mol) in a cold solution of benzene (61
ml, 0°C) over 1 hour. Further dehydration
Add 50 ml of THF and continue stirring for 2 hours. reaction mixture
After filtration, concentration under reduced pressure yields a dark red oily substance.
lorocarbonyl-(L)-proline t-butyl ester
Tell (1) (13.35g) was obtained. crude carbamoyl chloride1(9.35g) dehydrated
Add THF solution (40 ml) to methylhydrazine (3.46
g; 4.00ml; 0.075 mol) and triethylamine
16.33 g; 8.73 ml; 0.063 mol) of water-cooled dehydrated
The mixture was added to a THF solution (40 ml). Stir at room temperature for 18 hours.
Stirred. When the reaction mixture is filtered and concentrated under reduced pressure, it turns pale yellow.
A colored solid was obtained. Reconstituted from hexane-chloroform
When crystallization is carried out, white solid 2 (6.62 g; 0.0272 mole
68%). Melting point 89-92℃, TLC (silica
Gel; CH₂Cl₂−CH₃OH, 10:1)R_f=0.45.Element
Analysis: C₁₁H_{twenty one}N₃O₃Calculated value; C54.30; H8.70;
N, 17.27. Actual value: C54.45; H8.96; N, 17.29.
NMR (CDCl)₃): δ1.42 (9H, s); 1.7−2.3 (4H,
s); 3.00 (3H, s); 3.4-3.8 (2H, m); 3.8
(2H, bs); 4.4 (1H, m). IR (CHCl₃): 3500−
3300, 1730, 1630cm^-1・[α]_D−55.7°±0.5°. Example 2 α-Azaglycinyl-(L)-proline t-butylene
Le Esther (3) carbamoyl chloride1(1.00g; above)
Hydrazine dehydrated from water THF solution (3.5ml)
(0.269g; 0.310ml; 8.40mmol) and triethyl
Dehydration of amine (0.708; 0.975 ml; 7.00 mmol)
Added to ice-cold THF solution (4 ml) over 10 minutes.
Stir overnight at room temperature. When the mixture is filtered and concentrated
A pale yellow solid was obtained. From hexane-ethyl acetate
Recrystallization produces pure carbazide 3 as a white solid.
(0.533 g; 2.33 mmol; 52%) was obtained. melting point 83
-84℃. TLC (silica gel; CH₂Cl₂−CH₃Oh,
10:1)R_f=0.25~0.35.MSm/e 173(M⁺−
56). Elemental analysis: C_TenH₁₉N₃O₃Calculated value: C, 52.39;
H, 8.35; N, 18.33. Actual value: C, 52.08; H,
8.58; N, 18.01; NMR (CDCl₃) δ1.43 (9H,
s); 2.0-2.3 (4H, m); 3.1-3.4 (2H, m); 3.8
(2H, bs); 4.3−4.6 (1H, m); 6.2 (1H, bs);
IR (CHCl₃): 3500−3300, 1730, 1660, 1640cm
^-1.〔α〕_D−46.5±0.5°. Example 3 N-carbobenzyloxy-N'-isopropyl
Ruhydrazine (4) Carbobenzyloxyhydrazine (1.00g;
6.02 mmol) in anhydrous ethanol solution (6 ml)
Acetone (0.377g; 0.477ml; 6.50 mmol)
added. After 1 hour, the solution was concentrated to dryness and diluted with ethanol.
(5ml) and sodium cyanoborohydride
(0.19 g; 3.0 mmol) was added. Mixture at 6pm
After stirring for a while, it was concentrated to a thick syrup. hydrochloric acid
(0.50N; 5ml) and ether (50ml),
The layer mixture was stirred for 1 hour. solid water layer
NaHCO₃Neutralize by adding ether (2 x 25 ml)
Extracted with. The collected ether part is mixed with water and saturated salt.
Washed with water and dried (MgSO_Four)did. distill off the solvent
The solid obtained is recrystallized from hexane-ether.
and pure hydrazide 4 (1.09 g; 5.25 mm
87%). Melting point 61-62℃. TLC (Silica
Gel; CH₂Cl₂−CH₃OH, 20:1)R_f=0.5.MS
m/e 208 (M⁺). Elemental analysis: C₁₁H₁₆N₂O₂total
Calculated value: C, 63.44; H, 7.44; N, 13.45. Actual value:
C, 63.22; H, 7.67; N, 13.71; NMR
(CDCl₃); δ0.97 (6H, d, J=6); 3.15 (1H,
Septet, J = 6); 3.8-4.6 (1H, br); 5.13 (2H,
s); 6.2−7.0 (1H, br); 7.33 (5H, s); IR
(CHCl₃): 3500, 1720cm^-1, Example 4 α-Azabarinyl-(L)-proline t-butyl
ester(5) Carbamoyl chloride 1 (4 ml) prepared above
carbazide 4 (3.50 mmol, crude product)
0.728g) and triethylamine (0.557ml;
4.00 mmol) in dehydrated THF solution (2 ml).
Ta. The mixture was heated under reflux for 7 hours, filtered and concentrated.
It turned out to be a thick oil. Dissolve this in benzene (10ml)
and 1 atm with 10% Pd(C) (0.35 g) as a catalyst at 6 o'clock.
Hydrogenated for a while. When the catalyst is separated and concentrated, it turns pale yellow.
An oily substance was obtained, and further silica gel (CH₂Cl₂−
CH₃OH, 50:1) flash chromatograph
Purified by phytochemistry. Pure as a pale yellow solid
carbazide 5 (0.622g; 2.30 mmol; 66%)
I got it. When recrystallized from hexane, melting point 87−91
Samples were obtained at ℃. TLC (silica gel; CH₂Cl₂−
CH₃OH, 20:1)R_f=0.4.MSm/e 271
(M⁺). Elemental analysis: C₁₃H_{twenty five}N₃O₃Calculated value; C,
57.54; H, 9.29; N, 15.48. Actual value; C, 57.67;
H, 9.46; N, 15.44.NMR (CDCl₃): δ1.08 (3H,
d, J=6); 1.13 (3H, d, J=6); 1.43 (9H,
s); 1.7-2.2 (4H, m); 2.4-2.7 (2H, m); 3.3
(2H, bs); 4.07 (1H, septet, J=6); 4.3−
4.5 (1H, m). IR (CHCl₃): 3600−3300, 1730,
1630cm^-1.〔α〕_D15.9゜±0.5゜. Example 5 N-t-BOC-2-pyrrolidinol (6)
Di-t-butyl dicarbonate (2.3ml; 10ml
mole) to 4-aminobutyraldehyde diethyla
Setal (1.73ml: 10mmol) in chloroform
Add dropwise to the solution (20ml). Temperature rise of solution and gas
occurrence was observed. After 10 minutes, TLC (silicage)
LE；CH₃Cl₂−CH₃starting material by OH, 20:1)
of amine is consumed and R_f= 0.8 of a single product
Got a spot. The solvent was distilled off to obtain a colorless oil.
Ta. The crude product was mixed with THF (30 ml) and hydrochloric acid (0.50N, 30
ml) mixture. Stirring was continued for 2 hours.
The mixture was then diluted with saturated saline (50 ml) and
(3 x 25 ml). The collected ether part
Wash and dry with water, saturated sodium bicarbonate solution, and saturated salt solution
(MgSO_Four)did. The colorless oil obtained by distilling off the solvent is
Silicage using hexane-ether (2:1)
Baker (2 x 18 cm) chromatography
When purified, pure 6 (0.35g:
1.87 mmol; 19%). TLC (Shirikage
R; hexane-ether, 2:1) R_f=0.5.MS
m/e 170 (M⁺-OH). NMR (CDCl)₃): δ1.48
(9H, s); 1.8-2.1 (4H, m); 3.2-3.7 (2H,
m); 4.2−4.4 (1H, br); 5.4−5.6 (1H, br), IR
(CHCl₃): 3700−3400, 3000, 2920, 1675cm^-1. Example 6 N-(t-BOC-4-amino-1-butyl)-
N′-CBZ-hydrazine (8) N-t-BOC-2-pyrrolidinol (0.187
g; 1.00 mmol) and carbobenzyl oxyhydride
Radin (0.166 g; 1.00 mmol) was dehydrated.
Mix with THF (2 ml) and heat the resulting solution for 2 hours.
It refluxed. TLC (silica gel; hexane-
EtOAc, 2:1) is two carbazone isomers7a，
7b, R _f =0.2, 0.3. The mixture was concentrated to dryness and the residue was mixed with ethanol (3 ml) and sodium cyanoborohydride (63 mg; 1.00 mmol). After stirring the solution for 3 hours, the solvent was concentrated and exchanged with hydrochloric acid (0.50N, 3ml) and ether (15ml).
After stirring the two-layer mixture for 30 minutes, remove the aqueous layer from the solid.
Neutralized with _NaHCO3 and extracted with ethyl acetate (3x10ml). The combined solvent layer was dried (MgSO ₄ ) and the solvent was distilled off to give a pale yellow oil. Pure carbazide 8 (0.155 g; 0.46 mmol; 46
%) was obtained and recrystallized from hexane-EtOAc.
Melting point 73-74℃, TLC (silica gel; hexane-
EtOAc, 2:1) R _f =0.25.MSm/e 281(M ⁺
-56). Elemental analysis: Calculated value of C ₁₇ H ₂₇ N ₃ O ₄ : C,
60.51; H, 8.06; N, 12.45. Actual value: C, 60.18;
H, 8.11; N, 12.26, NMR ( _CDCl3 ): δ1.42
(9H, s); 2.8-3.3 (4H, m); 3.9-4.1 (1H,
br); 4.8−5.0 (1H, br); 5.17 (2H, s); 6.7−
6.9 (1H, br); 7.37 (5H, s); IR (CHCl ₃ ):
3500, 2980, 1720 cm ^-1 . Example 7 N-t-BOC-α-azalidyl-(L)-proline t-butyl ester ( 10 ) Carbazide 8 (0.941 g; 2.79 mmol) and carbamoyl chloride 1 (4 mmol; A solution of the crude product in dehydrated THF (7 ml) containing triethylamine (0.56 ml; 4.00 mmol) was refluxed for 8 hours.
TLC (silica gel; hexane-EtOAc, 1:1,
Two developments) showed a single product (9), R _f =0.5. The residue obtained by distilling off the solvent was dissolved in a mixture of ethyl acetate (10 ml) and acetic acid (3 ml), and the solution was diluted with 10%
Hydrogenation was carried out using Pd(C) (0.20 g) at 1 atm for 4 hours.
The crude product was flash chromatographed on silica gel (E. Merck No. 9385, 1.5 x 22 cm) with hexane-EtOAc (2:1) to give pure 10 (0.579 g; 1.45 mmol; 52%) as a colorless oil. ) was obtained. TLC (silica gel, EtOAc) R _f =0.3.MSm/
e 400 (M ⁺ ). NMR (CDCl ₃ ): δ1.43 (18H,
s); 1.8-2.1 (8H, m); 3.0-3.8 (8H, m); 4.3
−4.7 (1H, m); 4.8−5.1 (1H, br). IR
( _CHCl3 ): 3600−3300, 1730 (sh), 1720, 1630cm
^-1 . Example 8 α-Azaalanyl-(L)-phenylalanine t
-Butyl ester ( 11 ) Triethylamine (0.28 ml; 2 mmol) was added to a suspension of (L)-phenylalanine t-butyl ester hydrochloride (0.50 g; 1.94 mmol) in methylene chloride (4 ml). Carbonyldiimidazole (0.325 g; 2 mmol) was added and stirred for 30 minutes.
Then methylhydrazine (0.11 ml; 2 mmol)
was added over 5 minutes and stirred for 18 hours. The solvent was distilled off and the resulting solid residue was partitioned between ether and water. The ether layer was washed twice with water and once with saturated brine, and dried (MgSO ₄ ). The solvent was distilled off to obtain crude carbazide 11 as a colorless oil. Silica gel (E. Merck No. 9385; 2 x 12 cm) flash chromatography with methylene chloride:methanol (100:1) gave pure 11 (0.480 g; 1.63 mmol; 84%). TLC ( _{CH2Cl2 : CH3OH} ,
20:1) R _f =0.3.MSm/e 294(M ⁺ ). Elemental analysis: Calculated value of C ₁₅ H ₂₄ N ₃ O ₃ : C, 61.20; H,
8.22; N, 14.27. Actual value: C, 61.75; H, 8.28;
N, 14.00.NMR (CDCl ₃ ): δ1.33 (9H, s); 3.00
(3H, s); 3.05 (2H, d, J=6); 3.6 (2H,
bs); 4.13 (1H, dt, J=8.6); 6.7−7.0 (1H,
m); 7.25 (5H, s); IR (CHCl ₃ ): 3450, 3000,
1730, 1650cm ^-1 . Example 9 α-Azaphenylalanyl-(L)-proline t
-Butyl ester ( 12 ) Carbamoyl chloride 1 was prepared from (L)-proline t-butyl ester (10 g) as described above and obtained as a pale red oil (15.7 g). this
An ice-cold mixture of benzylhydrazine dihydrochloride (0.029 mol; 5.7 g), triethylamine (0.088 mol; 12.2 ml) and THF (40 ml) was added as a THF solution (50 ml). The mixture was stirred at room temperature for 18 hours. After filtration and distillation of the solvent, a crude product was obtained as a thick syrup. This is CH ₂ Cl ₂ −CH ₃ OH
(100:1) silica gel (E.MercNo.9385;
(3 x 30 cm) was subjected to flash chromatography. Collect fractions containing pure 12, 4.06g
(0.0127 mol; 44%) was obtained. An earlier fraction contained a small amount of the isomeric carbazide 13, but this was not fully confirmed. TLC _{( CH2Cl2} :
CH ₃ OH = 20:1) R _f = 0.5.MSm/e 319
(M ⁺ ). NMR (CDCl ₃ ): δ1.42 (9H, s); 1.8−
2.2 (4H, m); 3.4-3.5 (2H, bs); 3.5-3.8 (2H,
m); 4.3-4.6 (1H, m); 4.62 (2H, s); 7.3-
7.4 (5H, s). IR ( _CHCl3 ): 3000, 1740, 1630cm
^-1 . Example 10 N-(1-carbobenzyloxy-3-phenylpropyl)-d-azaalanyl-(L)-proline t-butyl esters-( 14R , 14S ). Carbazide 2 (0.243 g; 1.00 mmol) and benzyl 4-phenyl-2-oxobutanoate (0.268 g; 1.00 mmol) were dissolved in THF (2 ml) and 2
The resulting solution was mixed with a drop of acetic acid and heated to 60° for 1 hour. The mixture was stirred at room temperature overnight and then concentrated to give a thick, yellow syrup. TLC (silica gel; CH ₂ Cl ₂ -CH ₃ OH, 20:1) showed a single product spot at R _f =0.6. To a solution of the crude product in absolute ethanol (3 ml) was added sodium cyanoborohydride (63 mg; 1.00 mmol). 3
After an hour, the ethanol was distilled off and the residue was mixed with dilute hydrochloric acid (0.5N, 3ml) and ethyl acetate (15ml). The biphasic mixture was stirred vigorously for 1 hour, and the aqueous layer was neutralized by the addition of solid potassium carbonate and extracted three times with ethyl acetate. The collected solvent layers were washed with water and saturated brine, and dried (MgSO ₄ ). The solvent was distilled off to give a thick colorless oil. TLC (silica gel; hexane-ethyl acetate 2:1, developed twice) showed two diastereomers, R _f =0.25, 0.30. Separation of the two was achieved using medium pressure chromatography on silica gel (E. Merck 60B; 6.5 ml/min) with hexane-ethyl acetate (4:1). The following were obtained: 1 14S; the more mobile diastereomer (158
mg). MSm/e495 (M ⁺ ). NMR ( _CDCl3 ):
δ1.42 (9H, s): 1.8-2.3 (6H, m); 2.5-2.9
(2H, m); 2.90 (3H, s); 3.1−3.6 (3H,
m); 4.3-4.6 (1H, m); 5.0 (1H, br); 5.10
(2H, s); 7.10 (5H, s); 7.28 (5H, s). IR
(CHCl ₃ ): 3000, 1740, 1640 cm ^-1 .2 Mixed fraction 53.5 mg 3 14 R; less mobile diastereomer (170 mg). Melting point 76-79°C (hexane-EtOAc). MSm/e495 (M ⁺ ). NMR (CDCl ₃ ): δ1.42 (9H,
s); 1.7-2.2 (6H, m); 2.5-2.9 (2H, m);
2.92 (3H, s); 3.1-3.6 (3H, m); 4.2-4.5 (1H,
m); 5.07 (2H, s); 5.1-5.3 (1H, br); 7.10
(5H, s); 7.27 (5H, s). IR ( _CHCl3 ): 3000,
1740, 1640 cm ^-1 . (total amount 382 mg; 0.771 mmol; 77
%). Elemental analysis: Calculated value of C ₂₈ H ₃₇ N ₃ O ₅ : C,
67.86; H, 7.52; N, 8.48. Actual value: C, 67.74;
H, 7.39, N, 8.29. Example 11 N-(1-carbobenzyloxy-3-phenylpropyl)-α-azaalanyl-(L)-prolines ( 15 R, 15 S) More mobile aza Peptide ester 14S
(158 mg) was left in a solution of trifluoroacetic acid (10 ml) for 3 hours. The solution was concentrated to dryness, and the residue in ether was washed several times with water and dried (MgSO ₄ ). The concentrated residue obtained by distilling off the solvent was purified using a LH-20 (methanol) column to obtain 118 mg of azapeptide 15S as a colorless oil. TLC (silica gel; ethyl acetate-pyridine-acetic acid-water (EPAW) 20:
5:1:1) R _f = 0.6. NMR (CDCl ₃ ): δ1.7-2.3 (6H, m); 2.5-2.9
(2H, m); 2.95 (3H, s); 3.3−4.2 (3H, m);
4.4-4.8 (1H, m); 5.3 (2H, s); 7.1-7.4 (5H,
m); 7.30 (5H, s). The less mobile azapeptide diastereomer 14 R (170 mg) was similarly treated and the resulting product was purified by chromatography on LH-20 (methanol) to yield the pure azapeptide 15 R92.
I got mg. TLC (silica gel; EPAW20:5:
1:1) R _f = 0.6. NMR (CDCl ₃ ): δ1.6-2.2 (6H, m); 2.5-2.9
(2H, m); 3.08 (3H, s); 3.3−3.6 (2H, m);
3.8-4.1 (1H, m); 4.7-4.9 (1H, m); 5.02 (2H,
s); 5.8-6.0 (1H, br); 7.1-7.3 (5H, m);
7.18 (5H, s). Example 12 N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl-(L)-proline t
-Butyl esters ( 16R , 16S ) Using the method described in Example 11 above, carbazide 2 (0.486 g; 2.00 mmol) and 4-phenyl-
Azapeptide 15 (R,S) was prepared using 2-oxobutanoate (0.412 g; 2.00 mmol),
This was flushed with silica gel (E.Merck No. 9385; 2 x 25 cm) using CH ₂ Cl ₂ -CH ₃ OH (100:1).
Purified by chromatography. Product (0.753
g: 1.74 mmol; 87%) was a colorless oil. Separation of diastereomers was performed by pipetting E.
This was accomplished on a column (3 x 18 cm) of Merck silica gel H (following the method of Stahl). more mobile isomer (116 mg), less mobile isomer (75.4
mg), a mixture of the two (300 mg) was obtained. The more mobile isomer 16S : TLC (silica gel; hexane-
EtOAc, 2:1, 2 times) R _f =0.30, MSm/
e433(M ⁺ ). NMR (CDCl ₃ ): δ1.25 (3H, τ, J
=7); 1.43 (9H, s); 1.7-2.3 (6H, m); 2.6-
3.0 (2H, m); 2.95 (3H, s); 3.4−3.7 (3H,
m); 4.12 (2H, q, J = 7); 4.45 (1H, wide q, J = 6); 4.8-5.0 (1H, br); 7.2-7.4 (5H,
bs); IR (CHCl ₃ ); 1730, 1640 cm ^-1 . Less mobile isomer (16R): TLC, R _f =
0.25.MSm/e433(M ⁺ ). NMR ( _CDCl3 ): δ1.23
(3H, t, J=7); 1.42 (9H, s); 1.8−2.4
(6H, m); 2.6−3.0 (2H, m); 2.95 (3H, s);
3.4−3.7 (3H, m); 4.12 (2H, q, J=7); 4.3
−4.5 (1H, m); 5.0−5.3 (1H, br); 7.2−7.4
(5H, bs), IR (CHCl ₃ ): 1730, 1640 cm ^-1 . Example 13 N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl-(L)-prolines (17R, 17S ) The respective diastereomers of azapeptide 16 R, 16 S were treated with trifluoroacetic acid as described in Example 11 above with the following results. Azapeptide 16S (0.116g; 0.238 mmol) was purified by LH-20 (methanol) and azapeptide 17S
(0.0448 g; 0.119 mmol; 50%).
TLC (silica gel; EPAW, 20:5:1:1) R _f
=0.65MSm/e449 (M ⁺ , monotrimethylsilyl), NMR (CDCl ₃ ): δ1.28 (3H, t, J = 7);
1.8-2.4 (6H, m); 2.5-2.9 (2H, m); 3.07 (3H,
S); 3.5−4.0 (3H, m); 4.17 (2H, q, J=
7); 4.3-4.7 (1H, m); 7.23 (5H, bs); IR
(CHCl ₃ ): 3500−2900, 1740, 1670 cm ⁻¹ .Azapeptide 16R (0.0754 g; 0.174 mmol) is LH−20
After purification with azapeptide 17R (0.0398g; 0.106
mmol; 61%). TLC, R _f =0.65.MS
m/e449 (M ⁺ , monotrimethylsilyl). NMR
(CDCl ₃ ): δ1.28 (3H, t, J=7); 1.8−2.4
(6H, m); 2.7−3.0 (2H, m); 3.10 (3H, S);
3.5−4.0 (3H, m); 4.17 (2H, q, J=7); 4.5
−4.9 (1H, m); 7.15 (5H, m); IR (CHCl ₃ ):
3500−2900, 1740, 1670 cm ^-1 . Example 14 N-(1-carboxy-3-phenylpropyl)
-α-azaalanyl-(L)-proline t-butyl ester ( 18 ) carband 2 (0.243 g; 1.00 mmol) and 4
-Phenyl-2-oxobutanoic acid (0.534g;
An aqueous solution (1 ml) of 3.00 mmol) was brought to pH=7 by addition of aqueous sodium hydroxide (2N). Next, sodium cyanoborohydride (0.0628g;
An aqueous solution (1 ml) of 1.00 mmol) was added over 4 hours. Washed DOWEX ^R 50M-×4 resin (3
g) was added and stirred for 90 minutes. Mixture water _CH3OH
(1:1; 5 ml) and transferred to a column of the same resin (5 g). Elute with _H2O - _CH3OH (10:1)
This was followed by H ₂ O-pyridine (50:1). The latter solvent mixture was lyophilized to yield pure azapeptide ester 18 (0.300 g; 0.742 mmol; 74%) as a white foam. TLC (silica gel;
EPAW, 20:5:1:1) R _f =0.6.MSm/e477
(M ⁺ , monotrimethylsilyl). NMR ( _CDCl3 );
δ1.43 (9H, S); 1.7-2.3 (6H, m); 2.6-3.0
(2H, m); 3.00 (3H, S); 3.4−3.7 (2H, m);
4.3−4.6 (1H, m); 7.20 (5H, s); 7.25 (2H,
bs) Example 15 N-(1-carboxy-3-phenylpropyl)
An EtOAc solution (2 ml) of -α-azaalanyl-(L)-proline ( 19 ) azapeptide ester 18 (10.301 g; 0.742 mmol) was added via syringe to an ice-cold EtOAc solution of anhydrous hydrogen chloride (5 ml; 5.16 N). Stirring was performed at room temperature for 2 hours, and the solution was concentrated to give a white solid. Purification is done using DOWEX ^R 50W-×4
Resin (7 g), H ₂ O-CH ₃ OH (1:1, then
10:1) and _H2O -pyridine (50:1). The latter solvent mixture gave azapeptide 19 (0.100 g; 0.287 mmol; 39%) as a white foam. Further purification by LH-20 (methanol) column chromatography yielded pure azapeptide 19 (71 mg) as a white foam. TLC (silica gel; EPAW, 10:5:1:3) R _f =0.5. Example 16 N-(1-carboxyethyl)-α-azaalanyl-(L)-proline t-butyl ester ( 20 ) Aqueous solution of carbazide 2 (0.486 g; 2.00 mmol) and sodium pyruvate (1.10 g; 10 mmol) (30ml) was stirred for 15 minutes. An aqueous solution (2 ml) of sodium cyanoborohydride (0.251 g; 4.00 mmol) was added over 4 hours and stirred for 18 hours.
Then 5 g of DOWEX ^R 50W-x4 resin (washed with water) was added and the mixture was stirred for 4 hours. The mixture was added to a column of the same resin (5 g) and diluted with _H2O - _CH3OH (10:
1), eluted with H ₂ O-pyridine (50:1). The latter solvent was pure azapeptide 20 (0.504 g; 1.6
mmol; 80%), and when this was lyophilized, it became a white foam. TLC (silica gel; EPAW, 20:5:1:1) R _f =0.7, NMR
(CDCl ₃ ): δ1.0 (3H, d, J=7); 1.45 (9H,
s); 1.8-2.3 (4H, m); 3.03 (3H, s) 3.5-3.9
(3H, m); 4.4-4.7 (1H, m); 8.2-8.5 (1H,
br). Example 17 A solution of N-(1-carboxyethyl)-α-azaalanyl-(L)-proline ( 21 ) azapeptide 20 (0.504 g; 1.60 mmol) in ethyl acetate (2.5 ml) was dissolved in ice-cold ethyl acetate with hydrogen chloride. solution (7.5ml; 5.2N). The resulting solution was stirred at room temperature for 4 hours. Distillation of the solvent gave a white solid residue. This solid is DOWEX ^R
Purification with _H2O -pyridine (50:1) as eluent at 50W-4 (7.5g) yields pure azapeptide 21 (0.290g), which becomes a white foam upon lyophilization.
g; 1.12 mmol; 70%). This substance (50
When purified with LH-20 (methanol) column, azapeptide 21 (20.0
mg) was given. TLC (silica gel; EPAW, 10:
5:1:3) R _f =0.26, 0.30 (2 diastereomers). Example 18 N-(1-carboxy-4-3-methylbutyl)
-α-Azaalanyl-(L)-proline t-butyl ester ( 22 ) Carbazide 2 (0.468 g; 2.00 mmol) and 4-
An aqueous solution (4 ml) of methyl-2-oxopentanoic acid (1.52 g; 10.0 mmol) was added to an aqueous solution (2 ml) of sodium cyanoborohydride (0.251 g; 4.00 mmol) over a period of 7 hours. Stirring was continued for 18 hours. Washed DOWEX ^R 50W-×4 resin (7.5
g) was added to the mixture (PH=2) and stirred vigorously for 2 hours. H ₂ O-CH ₃ OH (1:1) was used to dissolve any remaining insoluble material in the flask and the entire mixture was transferred to a column of DOWEX ^R resin (5 g).
The column was heated with _H2O - _CH3OH (1:1, then 10:1),
It was then eluted with _H2O -pyridine (50:1).
This final solvent makes the azapeptide 22 pure.
(0.596 g; 1.67 mmol; 84%) was eluted and became a white foam upon lyophilization. TLC (silica gel; EPAW, 20:5:1:1) R _f =0.80, 0.85
(2 diastereomers) MSm/e429 (M ⁺ , monotrimethylsilyl), NMR (CDCl ₃ ): δ0.95 (6H,
d, J=6): 1.38 (9H, s); 1.8−2.2 (7E,
m); 3.03 (3H, s); 3.3-3.6 (3H, m); 4.2-
4.5 (1H, m); 7.6-7.8 (2H, br), IR ( _CHCl3 ):
3600−2500, 1740, 1650cm ^-1 . Example 19 N-(1-carboxybutyl)-α-azaalanyl-(L)-proline ( 23 ) A trifluoroacetic acid solution (2 ml) of azapeptide 22 (74.4 mg; 0.208 mmol) was allowed to stand for 2 hours.
The solvent was concentrated and the residue was purified on DOWEX ^R 50W-x4 resin (4 g). _H2O -pyridine (50:1)
Azapeptide 23 (56.9
mg; 0.189 mmol; 91%). TLC (silica gel; EPAN, 10:5:1:3) R _f =0.5, 0.6
(2 diastereomers), MSm/e445 (M ⁺ , bistrimethylsilyl) NMR (CDCl ₃ ): δ0.98 (6H,
d, J=6); 1.4-1.8 (2H, m); 1.8-2.3 (5H,
m); 3.10 (3H, s); 3.4-3.8 (3H, m); 4.3-
4.7 (1H, m); 9.0−9.7 (3H, br), IR ( _CHCl3 ):
3600−2500, 1720, 1670, ^{1610cm -1} . Example 20 N-(1-carbobenzyloxy-5-phthalimidopentyl)-α-azaalanyl-(L)-
Proline t-butyl ester ( 24 ) carbazide 2 (0.665 g; 2.74 mmol) and benzyl 6-phthalimido-2-oxohexanoate (1.00 g; 2.74 mmol) in THF (5 ml) containing glacial acetic acid (3 drops). The mixture was heated under reflux for 3 hours.
TLC (silica gel; _{CH2Cl2 -CH3OH ,} 20:1)
showed a single product spot, R _f =0.5.
The crude product was treated with sodium cyanoborohydride (172 mg; 2.74 mmol) as in 14 (Example 10) and then purified with silica gel (hexane-EtOAc, 1:1).
Pure carbazide 24 (1.18 g; 2.00 mmol; 73%) was obtained by flash chromatography. TLC (silica gel; hexane-EtOAc,
1:1) is a mixture of diastereomers (1:1),
It showed R _f =0.25, 0.30. MSm/e592 (M ⁺ ), elemental analysis: Calculated value of C ₃₂ H ₄₀ N ₄ O ₇ : C, 64.85: H,
6.80; N, 9.45. Actual value: C, 64.24; H, 6.98;
N, 8.95.NMR ( _CDCl3 ): δ1.40 (9H, s); 1.5-
2.0 (8H, m); 2.88, 2.90 (3H, 2×s); 3.2−
3.7 (6H, m); 4.2-4.4 (1H, m); 4.8-5.1 (1H,
m); 5.12, 5.15 (2H, 2×s); 7.27 (5H, s);
7.6−7.9 (4H, m): IR (CHCl ₃ ): 2980, 1770,
1730, 1720, 1640 cm ^-1 Example 21 N-(1-carboxy-5-phthalimidopentyl)-α-azaalanyl-(L)-proline t
-butyl ester ( 25 ) A solution of azapeptide 24 (0.402 g; 0.679 mmol) in ethyl acetate (5 ml) and acetic acid (0.2 ml) was added to
Hydrogenation was performed using 110 mg of %Pd/C for 18 hours. The catalyst was separated and the solvent was distilled off to obtain azapeptide 25 (0.330 g) in the form of a white foam. TLC (silica gel;
EPAW, 20:5:1:1) R _f =0.7, MSm/
e574 (M ⁺ , monotrimethylsilyl). NMR
(CDCl ₃ ): δ1.42 (9H, s); 1.5−2.1 (10H, m);
3.00 (3H, s); 3.4-3.8 (5H, m); 4.3-4.6 (1H,
m); 7.6-7.8 (4H, m); 7.8-8.0 (1H, br). Example 22 N-(5-amino-1-carboxypentyl)-
α-Azaalanyl-(L) -proline- ( 27 ) Anhydrous hydrazine (1.70 mmol;
0.054g). After 3 hours, the mixture was filtered to remove a white precipitate and concentrated to give a white foam. Purification on DOWEX ^R 50W-×4 (7.5 g) using H ₂ O-pyridine (50:1) as eluent yielded a white foamy N-(5-amino-1-carboxypentyl)-α-azaalanyl- (L)-proline t-butyl ester 26 (0.264 g) was obtained. TLC
(Silica gel; EPAW, 10:5:1:3) R _f =
0.25.MSm/e516 (M ⁺ , bistrimethylsilyl). A trifluoroacetic acid solution (5 ml) of azapeptide ester 26 (0.264 g) was allowed to stand for 2 hours. Remove the solvent and use _DOWEX ^R 50W-×4 (10
Chromatography was performed according to g). 0.166 g of white foamy azapeptide 27 was obtained. TLC (silica gel; n-butanol, H ₂ O, acetic acid,
EtOAc1:1:1:1) R _f =0.1−0.2.MSm/
e517 (M ⁺ ; tristrimethylsilyl-15).
NMR (D ₂ O): δ1.2−2.2 (10H, m); 2.85 (3H,
s); 3.1-3.7 (5H, m); 4.0-4.4 (1H, m). Example 23 N-(1-carboethoxy-3-phenylpropyl)-α-azaglycinyl-(L)-proline t-butyl ester ( 28 ) Similarly to using the method described in Example 10 above for 14, Azapeptide using carbazide 3 (0.458 g; 2.00 mmol) and ethyl 4-phenyl-2-butanoate (0.412 g; 2.00 mmol)
28 was prepared and purified by flash chromatography on silica gel (0.633 g;
1.51 mmol; 76%). TLC (silica gel;
CH ₂ Cl ₂ −CH ₃ OH, 10:1) R _f =0.75.MSm/
e419(M ⁺ ). NMR (CDCl ₃ ): δ1.22 (3H, t, J
=7); 1.37 (9H, s); 1.8-2.3 (6H, m); 2.6-
3.0 (2H, m); 3.2-3.8 (3H, m); 4.0-4.4 (1H,
m); 4.08 (1H, q, J = 7); 4.6-4.8 (1H,
br); 6.6 (1H, bs); 7.17 (5H, bs). IR
(CHCl ₃ ): 3500-3300, 1730, 1660 cm ^-1 . Example 24 N-(1-carboethoxy-3-phenylpropyl)-α-azaglycinyl-(L)-proline (29) Azapeptide 28 (0.284 g; 0.677 mmol) with trifluoroacetic acid (as described in Example 11) and purified by DOWEX ^R 50W-×4 (8 g) using H ₂ O-pyridine (50:1) as eluent. Then, pure azapeptide 29 (0.196 g; 0.541 mmol; 80%) was obtained as a white foam by lyophilization. TLC (silica gel; EPAW, 20:5:1:
1) R _f =0.5.MSm/e507 (M ⁺ , disilyl).
NMR (CDCl ₃ ): δ1.22 (3H, t, J=7); 1.7−
2.4 (6H, m); 2.5-3.0 (2H, m); 3.1-3.9 (3H,
m); 4.0−4.6 (1H, m); 4.08 (2H, q, J=
7); 7.18 (5H, bs); 8.4-9.0 (2H, br). IR
( _CHCl3 ): 3600−2500, 1780 (sh), 1730, 1670,
1610cm ^-1 . Example 25 N-(1-carboxy-3-phenylpropyl)
-α-Azaglycinyl-(L)-proline ( 30 ) To a solution of azapeptide 29 (0.132 g; 0.362 mmol) in ethanol (1.5 ml) was added a sodium hydroxide solution (0.80 ml; 0.80 mmol). mixture
The mixture was stirred for 24 hours and applied to a column of DOWEX ^R 50W-x4 (6 g). Azapeptide 30 is eluted with _H2O -pyridine (50:1) and lyophilized as a white powder.
(0.080 g; 0.239 mmol; 66%), TLC
(Silica gel; EPAW, 10:5:1:3) R _f =
0.45.MSm/e551 (M ⁺ , trisilyl). NMR
(CDCl ₃ ): δ1.7−2.4 (6H, m); 2.7−3.1 (2H,
m); 3.3-4.2 (4H, m); 7.1-7.3 (5H, bs); 8.4
−9.4 (2H, br). IR ( _CHCl3 ): 3600−2500, 1780
(sh), 1730, 1670 cm ^-1 . Example 26 Ethyl-4-phenyl-2-oxobutanoate (α-azaglycinyl-(L)-proline t-
Butyl ester) Carbazone ( 31 ) A THF solution (3 ml) of carbazide 3 (0.458 g; 2.00 mmol) and ethyl 4-phenyl-2-oxobutanoate (0.412 g; 2.00 mmol) was stirred overnight. The obtained carbazone was treated with CH ₂ Cl ₂ −
Purification by flash chromatography on silica gel with CH ₃ OH (100:1) gave 31,0.515 g (1.23 mmol; 62%) of a colorless oil.
TLC (silica gel; _{CH2Cl2 -CH3OH ,} 20:1)
R _f =0.2−0.4.MSm/e417(M ⁺ ). IR ( _CHCl3 ):
1740, 1720, 1660, 1610cm ^-1 , elemental analysis:
Calculated value of C ₂₂ H ₃₁ N ₃ O ₅ : C, 63.29; H, 7.48; N,
10.06, Actual value: C, 63.69; H, 7.66; N, 10.02.
NMR (CDCl ₃ ): δ1.25 (3H, t, J=7); 1.37
(9H, s); 1.7-2.1 (4H, m); 2.7-2.9 (4H,
bs); 3.4−3.7 (2H, m); 4.13 (2H, q, J=
7); 4.5-4.7 (1H, m); 7.2-7.4 (5H, bs). Example 27 Ethyl-4-phenyl-α-oxobutanoate (α-azaglycinyl-(L)-proline)carbazone ( 32 ) A trifluoroacetic acid solution (3 ml) of carbazone 31 (80.0 mg; 0.192 mmol) was added for 2 hours. I left it alone. The light brown solid obtained by distilling off the solvent was recrystallized from hexane-ethanol (2:1) to yield pure carbazone 32 (31 mg; 0.085 mmol; 44%), mp 126-
Obtained 127°C. TLC (silica gel; EPAW, 20:
5:1:1) R _f = 0.5, elemental analysis: Calculated value of C ₁₈ H ₂₃ N ₃ O ₅ : C, 59.82; H, 6.41; N, 11.63, actual value: C, 59.66; H, 6.44; N ,11.48,MSm/
e361 (M ⁺ ), NMR (CDCl ₃ ): δ1.32 (3H, t, J
=7); 1.9−2.4 (4H, m); 2.80 (4H, bs); 3.4−
3.8 (2H, m); 4.23 (2H, q, J=7); 4.5−4.8
(1H, m); 7.13 (5H, s), IR (CHCl ₃ ): 3600−
2500, 1760, 1740 (sh), 1670, 1630, 1580 cm ^-1 , Example 28 N-(1-carboethoxy-3-phenylpropyl)-α-azaphenylalanyl-(L)-proline t- A solution of butyl ester ( 34 ) carbazide 12 (0.568 g; 1.78 mmol) and ethyl 4-phenyl-2-oxobutanoate (0.367 g; 1.78 mmol) in absolute ethanol (2 ml) containing glacial acetic acid (3 drops) was added to The mixture was heated to reflux for an hour. The obtained carbazone 33 was diluted with CH ₂ Cl ₂ −CH ₃ OH.
Purification by flash chromatography on silica gel (100:1) gave the pure material (0.387 g; 0.763 mmol; 43%) as a colorless oil. TLC (silica gel; _{CH2Cl2 - CH3OH} , 20:
1) R _f =0.7, MSm/e507(M ⁺ ), NMR
(CDCl ₃ ): δ1.32 (3H, t, J=7); 1.40 (9H,
s); 1.7-2.4 (4H, m); 2.68 (4H, s); 3.4-
3.8 (2H, m); 4.22 (2H, q, J=7); 4.5−4.8
(1H, m); 4.93 (2H, s); 6.8−7.3 (5H, m);
7.13 (5H, s), IR (CHCl ₃ ): 3000, 1730, 1660,
At 1600 cm ^-1 , carbazone 33 (0.387 g; 0.763 mmol) was treated with sodium cyanoborohydride (0.032 g; 0.509 mmol) in absolute ethanol (3 ml) for 24 hours. Work-up as described in Example 10 provided azapeptide 34 as a colorless oil. _{CH2Cl2− _}
Flash chromatography on silica gel with CH ₃ OH (100:1) gave the pure azapeptide (0.293 g; 0.575 mmol; 75%).
TLC (silica gel; _{CH2Cl2 -CH3OH ,} 20:1)
R _f =0.60, 0.65 (two diastereomers, 1:
1), IR (CHCl ₃ ): 1740, 1630 cm ^-1 , MSm/
e509 (M ⁺ ), NMR (CDCl ₃ ): δ1.22 (3H, t, J
=7); 1.42 (9H, s); 1.7-2.2 (6H, m); 2.4-
2.8 (2H, m); 3.4−3.8 (2H, m); 4.03 (2H, q,
J=7); 4.4-4.7 (3H, m); 7.22 (5H, bs);
7.32 (5H, bs), Example 29 N-(1-carboethoxy-3-phenylpropyl)-α-azaphenylalanyl-(L)-proline ( 35 ) Azapeptide 34 (0.293 g; 0.575 mmol A trifluoroacetic acid solution (6 ml) of ) was allowed to stand for 2 hours. The solution was concentrated to dryness and the light brown residue was dissolved in ether (25ml). The solution was washed several times with saturated saline, dried (MgSO ₄ ), and concentrated to give a white foamy 35
(0.193 g; 0.427 mmol; 74%) was obtained. TLC
(Silica gel; EPAW, 20:5:1:1) R _f =
0.75, MSm/e525 (M ⁺ , monotrimethylsilyl), NMR (CDCl ₃ ): δ1.22 (3H, t, J = 7);
1.7-2.3 (6H, m); 2.4-2.7 (1H, m); 3.4-3.8
(3H, m); 4.05 (2H, q, J=7); 4.1−4.4
(1H, m); 4.52 (2H, bs); 7.0−7.6 (10H, m),
IR (CHCl ₃ ): 3600-2500, 1740, 1680 cm ^-1 , Example 30 N-(1-carboxy-3-phenylpropyl)
-α-Azaphenylalanyl-(L)-proline ( 36 ) Aqueous sodium hydroxide solution (0.70 ml; 0.70 mmol) was added to a solution of azapeptide 35 (0.140 g; 0.309 mmol) in absolute ethanol (1.5 ml). ,twenty four
Stir for hours. DOWEX ^R 50W−× reaction mixture
4 (6 g). Elution with _H2O followed by _H2O -pyridine (50:1) and lyophilization afforded pure azapeptide 36 (0.0626 g; 0.147 mmol; 48%) as a white powder. TLC (silica gel;
EPAW, 10:5:1:3) R _f =0.80, 0.85 (two diastereomers), MSm/e569 (M ⁺ ) bistrimethylsilyl), NMR (CDCl ₃ ): δ1.6−2.2
(6H, m); 2.4-2.8 (2H, m); 3.5-3.9 (3H,
m); 4.3-4.7 (3H, m); 7.1-7.5 (10H, m), IR
(CHCl ₃ ): 3500-2900, 1730, 1650 cm ^-1 , Example 31 N-(1-carbobenzyloxy-3-phenylpropyl)-α-azabalinyl-(L)-proline t-butyl ester ( 37 ) A THF solution (3 ml) containing carbazide 36 (0.271 g; 1.00 mmol) and benzyl 4-phenyl-2-oxobutanoate (0.295 g; 1.10 mmol) in glacial acetic acid (2 drops) was heated under reflux for 8 hours.
TLC (silica gel; hexane-EtOAc, 2:1)
showed the product carbazone at R _f =0.50, 0.55. MSm/e521 (M ⁺ ), the residue after concentration was dissolved in ethanol and treated with sodium cyanoborohydride (63 mg; 1.00 mmol). 4 hours later,
The product was worked up as described in Example 10 and purified by chromatography on silica gel (Baker, 1.5 x 20 cm) with hexane-EtOAc (10:1). Pure azapeptide (0.307
g; 0.587 mmol; 59%) on silica gel (hexane-EtOAc, 5:1) gave R _f
It showed two diastereomers of =0.30, 0.35 (1:1 mixture). MSm/e523(M ⁺ ), NMR
(CDCl ₃ ): 1.28, 1.30 (6H, paired triplet,
J=6); 1.42 (9H, s); 1.8-2.4 (6H, m); 2.6
−3.0 (2H, m); 3.4−3.7 (4H, m); 4.3−4.7
(1H, m); 5.1-5.3 (2H, m); 7.17 (5H, bs);
7.33 (5H, s), IR (CHCl ₃ ): 3000, 1740, 1650
cm ^-1 , Example 32 1-(carboxy-3-phenylpropyl)-α
-Azabarinyl-(L)-proline ( 39 ) A trifluoroacetic acid solution (4 ml) of azapeptide diester 37 (0.105 g; 0.201 mmol) was allowed to stand for 2 hours. The solvent was concentrated and the residue in ethyl acetate (25 ml) was washed several times with H ₂ O and dried (MgSO ₄ ).
did. The residue obtained after concentration (0.100 g) showed a single spot with R _f =0.7 on TLC (silica gel; EPAW, 20:5:1:1). MSm/e539 (M ⁺ , moritrimethylsilyl) and crude azapeptide (38) were dissolved in ethyl acetate (3 ml) and glacial acetic acid (1 ml),
Hydrogenation was performed with 10% Pd/C (100 mg) for 2 hours. After removing the catalyst and concentrating the solvent, a white foam was obtained which was purified by chromatography on a column of LH-20 (CH ₃ OH). Obtained azapeptide 39
(colorless oil) (52 mg; 0.138 mmol; 69%)
TLC (silica gel; EPAW, 20:5:1:1) showed a single spot. MSm/e521 (M ⁺ , bistrimethylsilyl), NMR (CDCl ₃ ): δ1.0−1.4
(6H, m); 1.8-2.4 (6H, m); 2.6-3.0 (2H,
m); 3.2-3.8 (4H, m); 4.2-4.6 (1H, m); 7.1
−7.3 (5H, bs), IR (CHCl ₃ ): 3600−2500,
1730, 1640 cm ^-1 , Example 33 N〓-(1-carbobenzyloxy-3-phenylpropyl)-N〓-t-BOC-α-azalidyl-(L)-proline t-butyl ester ( 41 ) Carbazide 10 (0.308 g; 0.779 mmol) and benzyl 4-phenyl-2-oxobutanoate (0.230 g; 0.866 mmol) were mixed in dry tetrahydrofuran (2.5 ml) and the solution heated to reflux for 4 hours. The product was purified by silica gel (Baker, 1.5 x 20
cm), and this step allows excess α-
The ketoester was eluted first to allow separation. The product obtained was a mixture of 40 carbazone isomers with R _f =0.3 and 0.5 by TLC (silica gel; hexane-EtOAc, 2:1). This was treated with sodium cyanoborohydride (27 mg; 0.52 mmol) dissolved in ethanol (2 ml). After 4 hours, the reaction mixture was worked up as described in Example 10 to give the crude product as a colorless oil. Hexane-EtOAc
Silica gel ((Baker, 1.5×20
cm) to give azapeptide 41 (0.181 g; 0.279 mmol; 36%) as a 1:1 mixture of diastereomers. TLC (silica gel; hexane-EtOAc, 2:1, developed twice) R _f =0.25, 0.30,
MSm/e652 (M ⁺ ), NMR (CDCl ₃ ): δ1.42 (18H,
s); 1.7-2.8 (12H, m); 3.0-3.5 (6H, m);
4.2-4.6 (2H, m); 4.8-5.0 (1H, br); 5.08,
5.12 (2H, bs); 7.1−7.2 (5H, bs); 7.25 (5H,
s), Example 34 N-(1-carbobenzyloxy-3-phenylpropyl)-α-azalidyl-(L)-proline ( 42 ) azapeptide ester 41 (0.181 g; 0.279 mmol) in trifluoroacetic acid The solution (4ml) was left at room temperature for 4 hours. The mixture was concentrated to dryness, and the resulting light brown residue was mixed with DOWEX ^R 50W-×4 resin (10 g).
It was purified by chromatography. H ₂ O−
Elution with pyridine (50:1) resulted in pure azapeptide 42 (84.3 mg; 0.170 mmol; 61
%) was given. TLC (silica gel; EPAW, 10:
5:1:3) R _f = 0.5, NMR (DMSO−d ₆ ; 300M
Hz): δ1.4-2.0 (10H, m); 2.5-2.9 (2H, m);
3.1-3.7 (7H, m); 4.0-4.1 (1H, m); 5.09,
5.11 (2H, 2×s); 7.1−7.3 (5H, m); 7.40
(5H, s), IR (CHCl ₃ ): 3600−2500, 1730,
1630 cm ^-1 , Example 35 N〓-(1-carboxy-3-phenylpropyl)-α-azalidyl-(L)-proline ( 43 ) Azapeptide 42 (34.3 mg; 0.069 mmol) was dissolved in ethyl acetate (1.5 ml). ) and acetic acid (0.5ml),
The resulting solution was exposed to 1 atmosphere of hydrogen using 10% Pd/C (50 mg) as a catalyst. After 4 hours, the catalyst was removed by filtration and the resulting product (14.5 mg)
It was purified by chromatography using an LH-20 (methanol) column. Product 43 (9.4
mg; 0.023 mmol; 33%) TLC (silica gel;
EPAW, 10:5:1:3) shows a single spot with R _f =0.35. NMR (DMSO− _d6 ): δ1.6−2.0
(10H, m); 2.4-2.6 (2H, m); 2.8 (2H, bs);
3.0−3.5 (6H, m); 4.4 (1H, bs); 7.1−7.3 (5H,
m), Example 36 N〓-(1-carbobenzyloxy-5-phthalimidopentyl)-N〓-t-BOC-α-azalidyl-(L)-proline t-butyl ester ( 44 ) Performed on azapeptide 14 Using the method described in Example 10, carbazide 10 (0.142 g; 0.356 mmol)
and benzyl 6-phthalimido-2-hexanoate (0.130 g; 0.356 mmol) after flash chromatography on silica gel (1.5 x 22 cm; hexane-EtOAc, 1:1) with a yield of 50% (0.133 mg; Azapeptide 44 was prepared in mmol). TLC (silica gel; hexane-
EtOAc, 1:1, 2 times expansion) is R _f = 0.45, 0.50
A mixture of two diastereomers (1:1) was shown. MSm/e749 (M ⁺ ), IR (CHCl ₃ ): 1775,
1730 (sh), 1710, 1645 cm ^-1 , NMR (CDCl ₃ ):
1.42 (18H, s); 1.3-2.2 (14H, m); 3.0-3.8
(9H, m); 4.3-4.5 (2H, br); 5.10, 5.13 (2H,
2×s); 7.27 (5H, s); 7.7-7.9 (4H, m), Example 37 N〓-(1-carboxyl-5-phthalimidopentyl)-N〓-t-BOC-α-azalidyl-
A solution of (L)-proline t-butyl ester ( 45 ) azapeptide 44 in ethyl acetate (3 ml) containing 3 drops of glacial acetic acid was heated with 10% Pd/C (60 mg) at 1 atm, 18
Hydrogenated for hours. Obtained azapeptide 45
(89.2 mg; 0.135 mmol; 94%) was purified by TLC (silica gel; EPAW, 20:5:1:1) R _f =0.75. MSm/e803 (M ⁺ , bistrimethylsilyl), NMR (CDCl ₃ ): δ1.43 (18H, m); 1.4−
2.0 (14H, m); 3.0-3.8 (9H, m); 4.3-4.5 (1H,
m); 5.0-5.3 (1H, br); 7.2-7.4 (1H, br); 7.7
-7.9 (4H, m), Example 38 N〓-(1-carboxy-5-phthalimidopentyl)-α-azalidyl-(L)-proline ( 46 ) Trifluoro of azapeptide 45 (89.2 mg; 0.135 mmol) The acetic acid solution (3ml) was allowed to stand for 2 hours. Distill the solvent and remove the light brown residue by DOWEX ^R.
A 50W-×4 (7.5 g) chromatograph was performed.
Elution with H ₂ O-pyridine (50:1) gave pure azapeptide 46 (61.6 mg; 0.122 mmol; 91%) as a white foam. TLC (silica gel;
EPAW, 10:5:1:3) R _f =0.25, NMR
(CD ₃ OD): 1.3-2.2 (14H, m); 2.9-3.1 (2H,
m); 3.4-3.8 (7H, m); 4.3-4.6 (1H, m);
7.80 (4H, s), Example 39 N〓-(5-amino-1-carboxypentyl)
-α-Azaridyl-(L) -proline ( 47 ) Anhydrous hydrazine (12
mg, 0.378 mmol) was added. The solution was stirred for 18 hours and the white precipitate was removed by filtration. After concentration, the residue was chromatographed on DOWEX ^R 50W-x4 (8 g) using H ₂ O-pyridine (50:1) as eluent. Product ( 47 ) (35.9mg;
0.0923 mmol; 84%) was a white powder.
TLC (silica gel; _H2O -BuOH-AcOH, 1:
1:1) R _f =0.2, 0.4, MSm/e661 (tetratrimethylsilyl), NMR (DMSO-d ₆ ): δ1.3-1.8
(12H, m); 2.6-2.8 (2H, m); 3.0-3.6 (4H,
m), Example 40 N-(α-azaalanyl)-(L)-1,2,3,
4-Tetrahydro-3-isoquinolinecarboxylic acid t-butyl ester ( 48 ) (L)-1,2,3,4-tetrahydro-3-
Isoquinolinecarboxylic acid [prepared according to the method of Yamada and Kunieda; Chem.Pharm.Bull., 15 , 490-8
(1967)] to the corresponding t-butyl ester using the method of Taschner et al. (Liebigs. Ann.
Chem., 646 , 134-6 (1961)].
A solution of the ester and 1 equivalent of triethylamine in methylene chloride was added to a slight excess of phosgene (12.5% in benzene). Filter the resulting mixture;
The residue obtained after concentration was dissolved in methylene chloride, cooled on ice, and added to a methylene chloride solution of methylhydrazine (1 equivalent) and triethylamine (1 equivalent).
The mixture was stirred for several hours, filtered and concentrated. The resulting crude product was purified by silica gel chromatography using a methylene chloride-methanol mixture as the eluent. Example 41 N-[N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl]-(L)-1,
2,3,4-Tetrahydro-3-isoquinolinecarboxylic acid t-butyl ester ( 49 ) Azapeptide (49) was prepared using carbazide 48 and ethyl 4-phenyl-2-oxobutanoate using the method described in Example 10. 49 ) was prepared and purified by silica gel chromatography using a methylene chloride-methanol mixture as the eluent. Example 42 N-[N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl]-(L)-1,
A trifluoroacetic acid solution of 2,3,4-tetrahydro-3-isoquinolinecarboxylic acid ( 50 ) azapeptide ( 49 ) was left at room temperature for 4 hours. Distill the solvent and remove the residue.
H ₂ O-pyridine (50:1) was used as the eluent.
Chromatography on DOWEX ^R 50W-x4 resin gave a product which became a white foam upon lyophilization. Example 43 N-[N-(1-carboxy-3-phenylpropyl)-α-azaalanyl]-(L)-1,2,
3,4-tetrahydro-3-isoquinolinecarboxylic acid ( 51 ) azapeptide ( 50 ) _{H2O -CH3OH} (1:1)
The solution was treated with sodium hydroxide (2.5 eq.) for 24 hours. The mixture was applied to a DOWEX ^R 50W-x4 column. Elution with H ₂ O-pyridine (50:1) gave the product which lyophilized to a white foam. Example 44 N〓-(1-carboethoxy-3-phenylpropyl)-N〓-t-BOC-α-azalidyl-(L)
-Proline t-butyl esters ( 52R , 52S ) Dehydrated THF solution (7.5
ml) was heated under reflux for 16 hours. The solvent was distilled off, and absolute ethanol (5 ml) and sodium cyanoborohydride (93 mg; 1.48 mmol) were added. 4 of the solution
After stirring for an hour and working up as described in Example 10, the crude product was obtained as a colorless oil. TLC (silica gel; hexane-EtOAc, 1:1) showed a mixture of diastereomers (1:1), R _f =0.25, 0.30. Silica gel (hexane-EtOAc, 1:1)
column chromatography (yield = 0.755 g;
1.28 mmol; 87%) and then careful separation of the diastereomers on silica gel (hexane-EtOAc, 2:1) yielded: 1 The more mobile diastereomer (184 mg) of 52S, MS: m/e 591 (M ⁺ +1), Elemental analysis: Calculated value of C ₃₁ H ₅₀ N ₄ O ₇ : C, 63.02; H,
8.53; N, 9.49, actual value: C, 63.17; H, 8.60;
N, 9.12, NMR (CDCl ₃ ): 1.28 (3H, t, J=
6); 1.43 (9H, s); 1.48 (9H, s); 1.6-2.1
(10H, m); 2.2-2.3 (2H, m); 2.67 (2H, t,
J=8); 3.0−3.5 (7H, m); 4.17 (2H, q, J
= 7); 4.22 (1H, t, J = 7); 4.94 (1H, bs);
5.05 (1H, bs); 7.1-7.3 (5H, m), 2 Mixed fraction (70 mg) 3 Diastereomer less mobile than 52R (327 mg), MS: m/e591 (M ⁺ +1), element Analysis: Actual value: C, 62.59; H, 8.53; N, 9.21,
NMR (CDCl ₃ ): δ1.28 (3H, t, J=7); 1.46
(18H, s); 1.6-2.1 (10H, m); 2.2-2.3
(2H, m); 2.6-2.8 (2H, m); 3.1-3.3 (4H,
m); 3.4−3.5 (3H, m); 4.17 (2H, q, J=
7); 4.40 (1H, t, J = 7); 4.9-5.0 (1H,
br); 7.2-7.4 (5H, m), Example 45 N-(1-carboethoxy-3-phenylpropyl)-α-azalidyl-(L)-proline dihydrochloride ( 53R , 53S ) 52S (more Mobile diastereomer; 108mg)
An ice-cooled concentrated hydrochloric acid solution (6 ml) was stirred at 5°C for 18 hours. The solution was diluted with _H2O (25ml) and concentrated to give 53S (89mg) as a white solid. TLC (silica gel;
_CHCl3 : _CH3OH : _H2O :AcOH, 70:30:6:
4) R _f = 0.5, NMR (D ₂ O): δ1.18 (3H, t, J =
7); 1.5-2.0 (10H, m); 2.1-2.3 (2H, m);
2.5-2.7 (2H, m); 2.9-3.1 (2H, m); 3.1-3.6
(5H, m); 4.09 (2H, q, J=7); 4.3−4.4
(1H, m); 7.1−7.3 (5H, m), 52R (less mobile diastereomer;
Starting from 111 mg), a white solid was prepared using the same method.
53R (90 mg) was obtained. TLC (silica gel;
_CHCl3 : _CH3OH : _H2O :AcOH, 70:30:6:
4) R _f = 0.5, NMR (D ₂ O): δ1.24 (3H, t, J =
7); 1.5-2.1 (10H, m); 2.2-2.4 (2H, m);
2.6-2.8 (2H, m); 2.9-3.1 (2H, m); 3.2-3.7
(5H, m); 4.12 (2H, q, J=7); 4.6−4.7
(1H, m); 7.2-7.4 (5H, m), Example 46 N-(1-carboxy-3-phenylpropyl)
-α-Azaridyl-(L)-proline dihydrochloride ( 54R , 54S ) An aqueous sodium hydroxide solution (0.25N; 3ml) of 53S (89mg) was stirred for 18 hours and then treated with hydrochloric acid (0.50N).
Neutralized by addition of The product was lyophilized and then purified by ion exchange on a DOWEX ^R 50W-x4. Elute with H ₂ O-pyridine (25:1),
54S (free base) (65
mg; 0.16 mmol; 87%). This was further mixed with silica gel (CHCl ₃ :CH ₃ OH:H ₂ O:AcOH,
70:30:6:4) and reconcentration from hydrochloric acid (6N; 5 ml) gave 54S (5 mg) as the dihydrochloride. TLC (silica gel; CMWA, 70:30:6:4) R _f =0.35, elemental analysis: Calculated value of C ₂₀ H ₂₈ N ₄ O ₅・2HCl・7/2H ₂ O: C, 44.44; H, 6.90; N, 10.37, actual value:
C, 44.44; H, 6.78; N, 10.15, NMR ( _D2O ):
δ1.5−2.1 (10H, m); 2.2−2.4 (2H, m); 2.6−
2.8 (2H, m); 2.9-3.1 (2H, m); 3.3-3.6 (5H,
m); 4.3-4.45 (1H, m); 7.2-7.4 (5H, m); Starting from 53R (90 mg) and using a similar method, the dihydrochloride 54R (54 mg) was obtained. NMR
(D ₂ O): δ1.5−2.1 (10H, m); 2.1−2.3 (2H,
m); 2.6-2.8 (1H, m); 3.2-3.7 (5H, m); 4.4
−4.45 (1H, m); 7.2−7.4 (5H, m), TLC (silica gel; CMWA, 70:30:6:4) R _f =0.35 Example 47 N-(1-carboethoxy-3-phenyl propyl)-α-azaalanyl-(L)-proline ethyl ester hydrochloride ( 55 ) N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl-(L)-proline t-
An ice-cold anhydrous ethanol solution of butyl ester (16S) was saturated with hydrogen chloride. After 1 hour, the solution was concentrated to dryness and the product was purified by chromatography on LH-20 (methanol). Example 48 Ethyl 4-phenyl-2-hydrazibutanoate trifluoroacetate ( 56 ) Ethyl 4-phenyl-2-oxobutanoate (2.06 g; 10 mmol) and t-butylcarbazate (1.32 g) ;10 mmol) in THF solution (50 ml)
The mixture was heated under reflux for 8 hours. The solvent was distilled off and the residue was dissolved in absolute ethanol (25 ml). Add sodium cyanoborohydride (0.31 g; 5 mmol),
The solution was stirred for 4 hours. After concentration, the residue was dissolved in EtOAc.
(50ml) and hydrochloric acid (0.5N; 20ml) for 30 minutes.
The aqueous layer was neutralized by addition of _K2CO3 and EtOAc (3 x 25 ml) _.
Extracted with. The collected solvent layers were washed with H ₂ O and saturated brine, and dried (MgSO ₄ ). After concentration, the residue was purified by chromatography on silica gel to obtain N-(1-carboethoxy-3-phenylpropyl)-N'-BOC-hydrazine. The ice-cold trifluoroacetic acid solution of hydrazine was allowed to stand for 1 hour. The solvent was distilled off to obtain trifluoroacetate of hydrazine ( 56 ). Example 49 (4R)-4-carboethoxy-2-(2-hydroxyphenyl)thiazolidine ( 57 ) M. Schubert's method (J. Biol. Chem., 114 , 341 (1936)) 57 was prepared starting from L-cysteine ethyl ester and salicylaldehyde using the method of . Purification by silica gel chromatography gave 57 as a thick oil. TLC (silica gel; cyclohexane-
EtOAc3:1) showed a mixture of two isomers with R _f =0.4, 0.5. IR (thin film): 3300 (sharp), 3000,
1740, 1430 cm ^-1 , NMR (CCl ₄ ): 1.28 (3H, t,
J=7); 2.9-3.5 (2H, m); 3.8-4.4 (3H,
m); 5.50, 5.80 (1H, 2 x bs); 6.5-7.3 (4H,
m), Example 50 N-[N-(1-carboxy-3-phenylpropyl)-α-azaglycinyl]-(4R)-carboethoxy-2-(2-hydroxyphenyl)thiazolidine ( 58 ) Phosgene ( An ice-cold solution of 1.1 M toluene solution; 0.91 ml) was diluted with CH ₂ Cl ₂ (3 ml) and then 56
(0.34 g; 1.0 mmol) and triethylamine (0.28 ml; 2.0 mmol) in CH ₂ Cl ₂ solution (3 ml) was slowly added under cooling (ice bath). then 57
(0.25 g; 1.0 mmol) and triethylamine (0.14 ml; 1.0 mmol) in CH ₂ Cl ₂ (3 ml) were added and stirred at room temperature for 6 hours. reaction mixture
It was diluted with EtOAc (25 ml), extracted with H ₂ O (2 x 10 ml) and saturated brine, and dried (MgSO ₄ ). After concentration,
The residue was chromatographed on silica gel and 58 was isolated. Example 51 N-[(1-carboxy-3-phenylpropyl)-α-azaglycyl]-(4R)-carboxy-2-(2-hydroxyphenyl)thiazolidine ( 59 ) The above diester (Example 50) It was dissolved in aqueous sodium hydroxide (2N; 5 ml) and stirred for 1 hour.
The resulting solution was made acidic with dilute hydrochloric acid and the product was
Isolated by ion exchange chromatography on DOWEX ^R 50W-x4 resin. Example 52 Improved method of N-t-BOC-4-aminobutyraldehyde carbobenzoxyhydrazone ( 7 ) Chloroform solution of 4-aminobutyraldehyde diethyl acetal (9.33 g) and di-t-butyl dicarbonate (13.3 g) (120ml) was stirred for 2 hours. Concentrate the mixture and add THF (110ml) to the residue.
and hydrochloric acid (0.5N; 120ml) were added. The mixture was stirred vigorously for 1 hour, then cooled in an ice bath and diluted with ether (100ml). The aqueous layer is slightly basic (PH
= 8) Sodium hydroxide solution (1N)
added. Separate the two layers and separate the aqueous layer with ether (3×
100ml). The collected ether layer was mixed with H ₂ O,
It was washed with saturated saline and dried (K ₂ CO ₃ ). When concentrated, N-t-BOC-2-pyrrolidinol ( 6 )
A colorless oil (20 g) was obtained. This material was combined with carbobenzoxyhydrazine (6.64 g) and THF (60 g).
ml). The resulting solution was heated to reflux for 3 hours and stirred at room temperature for 20 hours. When the solvent was distilled off and the resulting solid was recrystallized from hexane-EtOAc, 7
(8.4 g; 43%), melting point 99-104°C. TLC
(Silica gel; EtOAc) showed a mixture of isomers (1:1) with R _f =0.50, 0.55. Elemental analysis:
Calculated value of C ₁₇ H ₂₆ N ₃ O ₄ : C, 60.70; H, 7.79; N,
12.49, Actual value: C, 61.09; H, 7.69; N,
12.45, NMR (CDCl ₃ ): δ1.43, 1.47 (9H, 2×
s); 1.8-2.1 (2H, m); 2.2-2.5 (1H, m); 3.0
-3.5 (3H, m); 4.8-5.0 (1H, m); 5.18, 5.27
(2H, 2×s); 7.3 (5H, s), Example 53 N-(t-BOC-4-aminobutyl)-N'-
CBZ-Hydrazine (8) Sodium cyanoborohydride (1.1 g) was added to a solution of Carbazone 7 (6.0 g) in THF (35 ml) and ethanol (20 ml). Cool the mixture (ice bath)
An ethanol solution of anhydrous HCl (a saturated solution diluted 4 times with ethanol) was slowly added. After adding 6 ml, the remaining carbazone is removed by TLC.
It disappeared at the top. Concentrate the mixture with EtOAc (125 ml)
and diluted with aqueous hydrochloric acid (0.5N; 30ml). The mixture was stirred vigorously for 15 minutes and the aqueous layer was neutralized by addition of _K2CO3 and extracted with EtOAc (3 x 50ml) _. The collected solvent layer was washed with H ₂ O, saturated brine, and dried (MgSO ₄ ).
did. After concentration, the residue was purified by chromatography on silica gel and recrystallized from hexane-EtOAc to give 8 (2.7 g; 8.2 mmol; 46%), mp.
73-74°C was given. Example 54 N-t-BOC-α-azalidyl-(L)-proline t-butyl ester ( 10 ) Carbazide 8 (1.50 g; 4.45 mmol) and carbamoyl chloride (prepared as described in Example 1; 6.7 mmol) A dehydrated THF solution (7.5 ml) containing triethylamine (0.66 ml, crude product) (4.50 mmol) was heated under reflux for 8 hours. The residue after concentration
It was suspended in EtOAc. Purification by column chromatography using silica gel (hexane-EtOAc) produces a colorless oil N〓-CBZ-N〓-t
-BOC-α-azalidyl-(L)-proline t-
Butyl ester (2.29g; 4.29mmol; 96%)
I got it. TLC (silica gel; hexane-EtOAc,
1:1) R _f = 0.35, elemental analysis: Calculated value of C ₂₇ H ₄₂ N ₂ O ₇ : C, 60.66; H, 7.92; N, 10.48, actual value: C, 61.15; H, 8.33; N, 10.04 , NMR
(CDCl ₃ ): δ1.43 (18H, s); 1.5−1.7 (4H, m);
1.75-2.3 (4H, m); 3.1-3.4 (3H, m); 3.4-3.6
(4H, m); 4.3 (1H, br); 4.8 (1H, br); 5.22
(2H, bs); 7.4 (5H, bs), a solution of this substance in absolute ethanol (20 ml) was dissolved in 10% Pd/C (100 mg).
Hydrogenation was carried out for an hour (1 atm). The catalyst was separated and the solvent was distilled off to obtain carbazide 10 as a colorless oil. TLC (silica gel; CH ₂ Cl ₂ :CH ₃ OH, 10:
1) R _f = 0.7, a portion (0.10 g) of this substance was subjected to silica gel chromatography (ether:methanol, 10:1)
It was purified by Elemental analysis: Calculated value _{of C19H36N4O5 :} C, 56.97; H, 9.06; _N , 13.99, actual _value :
C, 57.00; H, 9.29; N, 14.06, Example 55 Improvement method of N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl-(L)-proline ( 17S ) Azapeptide 16S (0.180 g; 0.416 mmol) with concentrated hydrochloric acid (5 ml) and the resulting solution was stirred at 5°C overnight. After dilution with H ₂ O (25 ml), the solvent was evaporated to give 17S as a pale yellow oil. This was mixed with silica gel (CHCl ₃ :CH ₃ OH:H ₂ O:AcOH,
85:30:5:1) and reconcentrated from dilute hydrochloric acid to give the product as an oil (as the hydrochloride salt; 0.141 g; 0.341 mmol; 82%).
I got it. TLC (silica gel: EPAW, 20:5:1:1) R _f
=0.65 Example 57 N-(1-carboxy-3-phenylpropyl)
-α-Azaalanyl-(L)-proline ( 19S ) A solution of the azapeptide 16S (more mobile diastereomer; 0.335 g; 0.774 mmol) in trifluoroacetic acid (5 ml) was allowed to stand for 3 hours. After concentration, the residue was mixed with sodium hydroxide (1N; 3ml) and THF (0.5
ml) and the resulting solution was stirred for 18 hours under a nitrogen stream. After neutralizing with dilute hydrochloric acid, the solution was transferred to DOWEX ^R 50W.
A column of -×4 ion exchange resin (5 g) was applied. The product was eluted with _H2O -pyridine (20:1) and lyophilized to give a white foam (0.215 g; mmol; 80%). Part of this substance (0.052
g) into silica gel (CHCl ₃ :CH ₃ OH:H ₂ O:
Purification by chromatography with AcOH (85:30:5:1) gave the pure product (0.031 g). Disodium salt precipitated from CH ₂ Cl ₂ ; melting point
220−225℃ (decomposition). TLC (silica gel; EPAW,
10:5:1:3) R _f =0.45, elemental analysis:
Calculated value of C ₁₇ H ₂₃ N ₃ O ₅・2Na: C, 51.65; H,
5.82; N, 10.63, actual value: C, 51.54; H,
5.79; N, 10.28, MS (as dicarboxylic acid):
m/e403 (monotrimethylsilyl of M ⁺ −H ₂ O),
NMR ( _CDCl3 ): δ1.7-2.3 (6H, m); 2.6-2.8
(2H, m); 3.00 (3H, s); 3.5−3.6 (3H, m);
4.3-4.5 (1H, m); 7.1-7.3 (5H, m); 7.4-8.0
(2H,br), Example 57 Additional Compounds of Structural Formula Using the methods and intermediates shown in the examples above, additional compounds of the structural formula shown in the table can be synthesized. These syntheses are based on the structural formula HN− from the table.
It can be made using CHCO ₂ H amino acid t-butyl esters and the known keto acids or esters listed in the table. [Table] [Table] [Table]

Claims (1)

[Claims] 1 Structural formula: [In the formula: R and R ₃ are independently hydrogen, lower alkyl, aralkyl; R ₁ is aralkyl, heteroaralkyl, substituted aralkyl, substituted heteroaralkyl (substituents of aryl and heteroaryl are independently amino, Halo, amino lower alkyl, hydroxy, lower alkoxy, lower alkyl and carbon number 1-8
alkyl and amino, acylamino, heteroarylamino, aryloxy, heteroaryloxy, arylthio, heteroarylthio,
alkyl having 1-8 carbon atoms substituted by hydroxyl; R ₂ is hydrogen, lower alkyl, amino-lower alkyl, aralkyl; A is cycloalkyl having 5-7 carbon atoms, aryl , heteroaryl; B is hydrogen, lower alkyl; or A and B contain N- _CHCO2R3 and have _the structural formula:
[Formula] and [Formula] (wherein; R ₃ is defined above; Q ₁ and Q ₂ taken together are CH ₂ CH ₂ , CH ₂
-S, R ₄ is aryl or aryl substituted by hydroxyl, amino lower alkyl, amino, halo, alkoxy CHR ₄ S, R ₅ is hydrogen,
lower alkyl, aryl, aralkyl,
CONR ₆ R ₇ (R ₆ and R ₇ are independently hydrogen, lower alkyl, aralkyl) CH ₂ −CH−
OR ₅ ; W is a bond or CH ₂ ; Y is a bond, CH ₂ , CH ₂ CH ₂ ) and its pharmaceutically acceptable salt. 2. The compound according to claim 1, wherein the asymmetric carbon has the configuration of an L-amino acid. 3 N-(1-carboxy-3-phenylpropyl)-α-azaalanyl-L-proline; N-(1
-(S)-carboxy-3-phenylipropyl]-
α-Azaalanyl-L-proline; N-(1-ethoxycarbonyl-3-phenylpropyl)-α
-Azaalanyl-L-proline; N-[1(S)-
Carboethoxy-3-phenylpropyl]-α-
Azaalanyl-L-proline; N-(1-carboethoxy-3-phenylpropyl)-α-azaalanyl-L-proline ethyl ester; N-[1
(S)-carboethoxy-3-phenylpropyl]
-α-Azaalanyl-L-proline ethyl ester; N-[1-carboethoxy-3-phenylpropyl)-α-azaglycyl]-4(R)-carboxy-2-phenylthiadiazolidine; N〓-( 1-
N〓-[1(S)-carboxy-3-phenylpropyl]-α-azalidyl-
3. The compound according to claim 2, which is a member of the group consisting of L-proline. 4 Structural formula: [In the formula: R and R ₃ are independently hydrogen, lower alkyl, aralkyl; R ₁ is aralkyl, heteroaralkyl, substituted aralkyl, substituted heteroaralkyl (substituents of aryl and heteroaryl are independently amino, Halo, amino lower alkyl, hydroxy, lower alkoxy, lower alkyl and carbon number 1-8
alkyl and amino, acylamino, heteroarylamino, aryloxy, heteroaryloxy, arylthio, heteroarylthio,
alkyl having 1-8 carbon atoms substituted by hydroxyl; R ₂ is hydrogen, lower alkyl, amino-lower alkyl, aralkyl; A is cycloalkyl having 5-7 carbon atoms, aryl , heteroaryl; B is hydrogen, lower alkyl; or A and B contain N- _CHCO2R3 and have _the structural formula:
[Formula] and [Formula] (wherein; R ₃ is defined above; Q ₁ and Q ₂ taken together are CH ₂ CH ₂ , CH ₂
-S, R ₄ is aryl or aryl substituted by hydroxyl, amino lower alkyl, amino, halo, alkoxy, CHR ₄ S, R ₅ is hydrogen,
lower alkyl, aryl, aralkyl,
CONR ₆ R ₇ (R ₆ and R ₇ are independently hydrogen, lower alkyl, aralkyl) CH ₂ −CH−
OR ₅ ; W is a bond or CH ₂ ; Y is a bond, CH ₂ , CH ₂ CH ₂ ). Structural formula: (wherein R, R ₃ , A, B are as defined above) and a protected semicarbazide derivative having the structural formula: R ₁ COCO ₂ R (R and R ₁ are as defined above) by reductive condensation with a keto acid or ketoester having the structure and, after removal of the protecting group, to form compounds of the structural formula, if necessary by chromatography or crystallization, to obtain a biologically more active diastereomer. A method comprising separating the mer and, if necessary, preparing a pharmaceutically acceptable salt thereof according to conventional methods.