Benzotriazole-mediated synthesis of aza-peptides: En route to an aza-leuenkephalin analogue

Article abstract of DOI:10.1021/jo302251e

Novel N-(N-Pg-azadipeptidoyl)benzotriazoles 20a-e couple efficiently with α-amino acids 21a-e, dipeptides 22a-c, aminoxyacetic acid 23a, depsidipeptide 23b, and α-hydroxy-β-phenylpropionic acid 27 yielding, respectively, azatripeptides 24a-g, azatetrapeptides 25a,b, a hybrid azatripeptide with an oxyamide bond 26a, a hybrid azatetrapeptide with an ester bond 26b, and a hybrid azatripeptide with an ester bond 28. A new protocol for the synthesis of N-Pg-azatripeptides 33a,b and 35a,b, each containing a natural amino acid at the N-terminus, avoids the low coupling rates of the aza-amino acid residue and enables the solution-phase synthesis of an azaphenylalanine analogue of Leu-enkephalin 40.

Full text of DOI:10.1021/jo302251e

Article
pubs.acs.org/joc
Benzotriazole-Mediated Synthesis of Aza-peptides: En Route to an
Aza-Leuenkephalin Analogue
Nader E. Abo-Dya,‡,§,# Suvendu Biswas,‡,# Akash Basak,‡ Ilker Avan,‡,∥ Khalid A. Alamry,⊥
and Alan R. Katritzky*,‡,⊥
‡Center for Heterocyclic Compounds, Department of Chemistry, University of Florida, Gainesville, Florida 32611-7200, United
States
§Department of Pharmaceutical Organic Chemistry, Faculty of Pharmacy, Zagazig University, Zagazig 44519, Egypt
∥Department of Chemistry, Anadolu University, 26470, Eskise
̧ hir, Turkey
⊥Department of Chemistry, King Abdulaziz University, Jeddah 21589, Saudi Arabia
S
* Supporting Information
ABSTRACT: Novel N-(N-Pg-azadipeptidoyl)benzotriazoles 20a−e couple efficiently with α-amino acids 21a−e, dipeptides
22a−c, aminoxyacetic acid 23a, depsidipeptide 23b, and α-hydroxy-β-phenylpropionic acid 27 yielding, respectively,
azatripeptides 24a−g, azatetrapeptides 25a,b, a hybrid azatripeptide with an oxyamide bond 26a, a hybrid azatetrapeptide
with an ester bond 26b, and a hybrid azatripeptide with an ester bond 28. A new protocol for the synthesis of N-Pg-azatripeptides
33a,b and 35a,b, each containing a natural amino acid at the N-terminus, avoids the low coupling rates of the aza-amino acid
residue and enables the solution-phase synthesis of an azaphenylalanine analogue of Leu-enkephalin 40.
geometry of an aza-amino acid is similar to that of a β-turn in
changing the chemical and biological properties of the parent
peptide. Azapeptides may exhibit better interactions with
protein receptors and enhanced stability to enzymatic and
chemical degradation.5,6 Azapeptides are therefore leads for the
generation of receptor ligands, enzyme inhibitors, and clinically
approved drugs.7 Azapeptides possessing electrophilic moieties
also act as inhibitors of cysteine proteases.8−10
INTRODUCTION
■
Azapeptides are peptidomimetics in which the α-CH groups of
one or more amino acid residues are replaced by a nitrogen
atom (Figure 1A, B). This decreases the electrophilicity of the
Azapeptides can be synthesized both in solution and on solid
phase by combining hydrazine chemistry and conventional
peptide synthesis.11 The nitrogen atoms of hydrazine need to
be differentiated to produce N′-alkyl-N-Pg-hydrazines 1;
phosgene (or an equivalent) introduces the carbonyl group
into the skeleton by reacting with either the hydrazine
derivative 1 giving 2 which then reacts with dipeptide ester 3
(Scheme 1A). Alternatively, the phosgene reacts with the
peptide N-terminus of dipeptide 3 (Scheme 1B).12−14 Route B
Figure 1. Comparison of azadipeptide and native dipeptide.
−(NR)CO− carbonyl group and changes the geometry at the α
positions from tetrahedral to trigonal-planar, hence eliminating
chirality. Relative to the natural peptides, azapeptides occupy a
limited conformational space with dihedral angles (φ = 90°
30° or −90° 30° and ψ = 0° 30° or 180° 30°) close to
those in the polyproline type II helix (φ = −78°, ψ = 149°) and
other types of β-turns (Figure 1C).1−4 The effect of the
Received: October 23, 2012
Published: February 1, 2013
© 2013 American Chemical Society
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Scheme 1. Construction of an Azapeptide by (A) a Hydrazine Pathway and (B) a Peptide N-Terminus Pathway
can produce a hydantoin byproduct 7 by intramolecular
nucleophilic attack of the secondary amide nitrogen of the
preceding amino acid residue on the activated isocyanate 6.
Lubell et al. coupled N-Boc-azadi- and tripeptide segments to
the amine terminus of a growing peptide chain to circumvent
formation of the hydantoin byproduct.13
N-Acylbenzotriazoles are advantageous for N-, O-, C-, and S-
acylation, especially when the corresponding acid chlorides are
unstable or difficult to prepare, and we have demonstrated the
use of benzotriazolides for the synthesis of peptidomimetics
such as aminoxypeptides and depsipeptides.15−17 In the present
work, stable, crystalline, and easy to handle N-(N-Pg-α-
azadipeptidoyl)benzotriazoles 20a−e were prepared, and their
synthetic utility was demonstrated by the synthesis of N-Pg-
azatripeptides 24a−h, N-Pg-azatetrapeptides 25a,b, and hybrid
azapeptides containing oxyamide 26a or depsi bonds 26b and
28. In addition, we developed a new route toward N-Pg-
azatripeptides containing a natural amino acid at their N-
terminus 33a,b and 35a,b. The novel pathway enabled the
solution phase synthesis of aza-Leu-enkephalin analogue 40.
fluorenyl-9-ylmethyl carbazates 14b as their HCl salts in 70−
73% overall yield (Scheme 3). Reaction of Cbz-NHNH2 13c
and methyl 2-bromoacetate in dry DMF in the presence of
DIPEA gave benzyl 2-(2-methoxy-2-oxoethyl)hydrazine-1-
carboxylate 14c (Scheme 3).
Construction of Protected Azadipeptides. The activa-
tion of N′-alkyl tert-butyl carbazates into N-(Boc)aza-amino
acid building blocks using p-nitrophenyl chloroformate,18
bis(2,4-dinitrophenyl) carbonate, and carbonyldiimidazole
(CDI)9 gave nitrophenylcarbazates and imidazolides which
required long reaction times and high temperatures to provide
aza-peptides in poor yields. Phosgene and triphosgene activate
N′-alkyl tert-butyl carbazates and N′-substituted fluorenylmeth-
yl carbazates efficiently, but these reagents are extremely toxic.
Although several literature papers describe synthetic protocols,
a general, high-yielding, and efficient synthetic method has yet
to be reported. After examination of the aza-peptide synthetic
literature,5 we chose to synthesize azadipeptides via activation
of the amino acid ester hydrochloride salts 15a−e by
carbonyldiimidazole (CDI) in the presence of 2.5 equiv of
DIPEA (Hunig’s base) in dry DCM to afford the active
̈
RESULTS AND DISCUSSION
■
carbamates 16a−e. Stirring 16a−e with N′-alkyl-N-Pg-
hydrazines 10a−c or 14a,b at 20 °C for 16 h in dry THF
containing 1.0 equiv of DIPEA provided protected azadipep-
tides 18a−h (Scheme 4). DIPEA establishes an equilibrium
between the active carbamates 16 and the isocyanate
intermediates 17 which react quickly with N′-alkyl-N-Pg-
hydrazines 10a−c or 14a,b to afford 18a−h. The method
tolerated N′-alkyl-N-Fmoc-hydrazines 10c and 14b without any
sign of N-Fmoc deprotection. A simple extractive workup using
2 N HCl gave rise to N-Pg-azadipeptide esters 18a−h (Table
Preparation of N′-Alkyl-N-(Pg)-hydrazines. Reaction of
Boc-NHNH2 8a and Fmoc-NHNH2 8b with the appropriate
aldehyde or ketone followed by reduction with NaCNBH3
furnished N′-alkyl tert-butyl carbazates 10a,b and N′-alkyl
fluorenyl-9-ylmethyl carbazates 10c (Scheme 2). Selective
protection of N-methyl hydrazine with (Boc)2O 11 afforded
N′-Boc-N′-methyl hydrazine 12 which was coupled with benzyl
chloroformate and fluorenyl-9-ylmethyl chloroformate to give
N′-Boc-N′-methyl-N-Cbz-hydrazine 13a and N′-Boc-N′-meth-
yl-N-Fmoc-hydrazine 13b. Subsequent Boc deprotection of
13a,b afforded N′-methyl benzyl carbazates 14a and N′-methyl
1
1) displaying satisfactory H NMR and 13C NMR and were
used in the next step without full characterization. In an
attempt to show general applicability of the protocol, we chose
to synthesize an azadipeptide containing a polar side chain.
Compound 14c was subjected to react with 16b, which gave
the protected azadipeptide 18i with the polar side chain (Cbz-
AzaAsp-Val-OtBu). It proves the methodology is general, and
by choosing appropriate coupling partners and protection/
deprotection methods when reacting with 16a−e, other
azapeptides with the polar side chains can be achieved.
Scheme 2. Preparation of N′-Alkyl-N-(Pg)-hydrazines 10a−c
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Scheme 3. Preparation of N′-Methyl Benzyl Carbazates 14a, N′-Methyl Fluorenyl-9-ylmethyl Carbazates 14b, and Benzyl 2-(2-
Methoxy-2-oxoethyl)hydrazine-1-carboxylate 14c
Scheme 4. Preparation of Protected Azadipeptides 18a−i
Table 1. Preparation of Protected Azadipeptides 18a−i
Scheme 5. Preparation of N-(N-Pg-
azadipeptidoyl)benzotriazoles 20a−e
18,
a
yield
entry
Pg
R2
R
R1
CH3
(%)
a
b
c
d
e
f
Cbz
Cbz
Boc
CH3
CH3
CH2Ph
18a, 90
CH(CH3)2
C(CH3)3 18b, 82
CH(CH3)2
CH(CH3)2
CH2Ph
CH2CH(CH3)2
CH2CH2SCH3
CH2CH(CH3)2
CH2Ph
CH3
CH3
CH3
CH3
18c, 89
18d, 90
18e, 95
18f, 80
Boc
Boc
Boc
H
g
h
i
Fmoc
Fmoc
Cbz
CH3
CH(CH3)2
C(CH3)3 18g, 85
C(CH3)3 18h, 78
C(CH3)3 18i, 92
CH(CH3)2
CH2CO2CH3
CH2CH(CH3)2
CH(CH3)2
a
Table 2. Preparation of N-Pg-azadipeptide-OH 19a−h and
N-(N-Pg-azadipeptidoyl)benzotriazoles 20a−e
Isolated yield over two steps.
a
a
Preparation of N-(N-Pg-azadipeptidoyl)benzotri-
azoles 20a−e. N-Pg-Azadipeptides 19a−h were prepared
either by hydrolysis of N-Pg-azadipeptides methyl esters 18a,c−
f using lithium hydroxide in methanol/water mixture (10:1, v/
v) or cleavage of the tert-butyl group of 18b,g,h with
trifluoroacetic acid in DCM (1:1 v/v). N-(N-Pg-
azadipeptidoyl)benzotriazoles 20a−e were synthesized in 81−
92% yields by treatment of N-Pg-azadipeptides 19a−d,h with
3.0 equiv of 1H-benzotriazole, 1.0 equiv of thionyl chloride, and
2.0 equiv of DIPEA in DCM at −30 °C (Scheme 5, Table 2).
The presence of DIPEA neutralizes the HCl liberated from the
reaction of benzotriazole and thionyl chloride, thus preventing
N-Boc-deprotection. N-(N-Pg-azadipeptidoyl)benzotriazoles
20a−e were characterized by 1H NMR, 13C NMR, and
elemental analysis.
N-Pg-azadipeptide- yield
N-(N-Pg-azadipeptidoyl)
benzotriazoles, 20a−e
yield
(%)
entry
A
OH, 19a−h
(%)
Cbz-AzaAla-Phe
90
83
79
86
89
76
69
80
Cbz-AzaAla-Phe-Bt 20a
Cbz-AzaAla-Val-Bt 20b
Boc-AzaVal-Leu-Bt 20c
Boc-AzaVal-Met-Bt 20d
(not attempted)
87
81
90
85
-OH 19a
B
Cbz-AzaAla-Val
-OH 19b
C
D
E
Boc-AzaVal-Leu
-OH 19c
Boc-AzaVal-Met
-OH 19d
Boc-AzaPhe-Leu
-OH 19e
F
Boc-AzaGly-Phe
-OH 19f
(not attempted)
G
H
Fmoc-AzaAla-Val
-OH 19g
(not attempted)
Fmoc-AzaVal-Leu
-OH 19h
Fmoc-AzaVal-Leu-Bt 20e
92
Coupling of N-(N-Pg-azadipeptidoyl)benzotriazoles
20a−e with Amino Acids 21a−e, Dipeptides 22a,b,
Aminoxyacetic Acid 23a, Depsidipeptide 23b, and α-
a
Isolated yield.
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Scheme 6. Coupling Reactions of N-(N-Pg-azadipeptidoyl)benzotriazoles 20a−e
Table 3. Preparation of Azapeptides 24a−g, 25a,b, 26a,b, and 28
a
entry
20
nucleophiles, 21a−e, 22a,b, 23a,b, and 27
products 24a−g, 25a,b, 26a,b, and 28
yield (%)
1
2
20a
20b
20b
20b
20c
20d
20c
20e
20a
20c
20a
20d
20b
H-Cys-OH 21a
Cbz-AzaAla-Phe-Cys-OH 24a
Cbz-AzaAla-Val-Ser-OH 24b
80
80
83
85
89
83
90
86
77
87
90
85
87
H-Ser-OH 21b
3
H-Cys-OH 21a
Cbz-AzaAla-Val-Cys-OH 24c
3′
4
(D,L) H-Cys-OH 21a + 21a′
H-Trp-OH 21c
Cbz-AzaAla-Val-(D,L)Cys-OH 24c + 24c′
Boc-AzaVal-Leu-Trp-OH 24d
Boc-AzaVal-Met-Ser-OH 24e
5
H-Ser-OH 21b
6
H-Asp-(OBn)-OH 21d
H-Pro-OH 21e
Boc-AzaVal-Leu-Asp-(OBn)-OH 24f
Fmoc-AzaVal-Leu-Pro-OH 24g
Cbz-AzaAla-Phe-Gly-Gly-OH 25a
Boc-AzaVal-Leu-Gly-Phe-OH 25b
Cbz-AzaAla-Phe-AOGly-OH 26a
Boc-AzaVal-Met-Gly-OPhe-OH 26b
Cbz-AzaAla-Val-OPhe-OH 28
7
8
H-Gly-Gly-OH 22a
H-Gly-Phe-OH 22b
H-AOGly-OH 23a
H-Gly-OPhe 23b
HO-Phe-OH 27
9
10
11
12
a
Isolated yield.
Hydroxy-β-phenylpropionic Acid 27. Dipeptides are useful
building blocks for longer peptide analogues; however, the
functions and applications of dipeptides have been previously
neglected, probably because of lack of an efficient protocol for
the synthesis of dipeptides.19 However, we now find that N-(N-
Pg-azadipeptidoyl)benzotriazoles 20a−e may be used to attain
longer azapeptide sequences by coupling with (i) α-amino acids
21a−e, (ii) dipeptides 22a,b, (iii) aminoxyacetic acid 23a, and
(iv) depsidipeptide 23b in aqueous acetonitrile−triethylamine
(TEA) at 20 °C for 0.5−2 h. These coupling reactions afford,
respectively, (i) N-Pg-azatripeptides 24a−g, (ii) N-Pg-azate-
trapeptides 25a,b, (iii) hybrid N-Pg-azatripeptides containing
the oxyamide bond 26a, and (iv) hybrid azatetrapeptide with an
ester bond 26b in 77−90% yields (Scheme 6, Table 3). The
coupling of α-hydroxy-β-phenylpropionic acid 27 with 20b in
dry THF containing 2.0 equiv of 4-dimethylaminopyridine
(DMAP) gave hybrid peptide 28. The target compounds were
all characterized by 1H NMR, 13C NMR, and elemental
analysis.
To show that no racemization occurs in the synthetic
protocol used in the paper, we conducted reactions between
the active benzotriazole dipeptide species 20b and amino acid
cysteine (both L and DL forms). The absence of racemization in
1
the azapeptide (24c + 24c′) was deduced from the H NMR,
where the −SH proton signal showed two separated triplets
split in the D,L-cysteine moiety at 1.63 ppm (J = 9.0 Hz) and
1.49 ppm (J = 9.0 Hz), while compound 24c showed a clear
triplet at 1.61 ppm (J = 9.0 Hz). Retention of the chirality in
the products was further confirmed by chiral HPLC analysis
using a (S,S) Welk-O1 column (MeCN/H2O 50:50, flow rate
0.25 mL/min, detection at 210 nm). The diastereomer 24c
showed a single retention-time peak in chiral HPLC at 10.2
min, while its corresponding diastereomeric mixture (24c +
24c′) showed two peaks at 9.3 and 10.1 min. In previous
studies on benzotriazole based peptide coupling we also
demonstrated chirality of the reaction is maintained on N-
acylation with N-acylbenzotriazoles for peptides17a depsipep-
tides16 and aminoxy hybrid-peptides.17b
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Scheme 7. Preparation of Free Azapeptides 29a−c and Coupling with N-(N-Pg-α-aminoacyl)benzotriazoles To Prepare 31a,b
Scheme 8. Preparation of Azatripeptide Starting from N-(N-Pg-α-aminoacyl)benzotriazoles 30
Preparation of Free-Amino Azadipeptides 29a−c and
Their Coupling with N-(N-Pg-α-aminoacyl)benzo-
triazoles. The addition of a new unit to a peptide (or
peptidomimetics) chain is accomplished chemically by
coupling. However, chain extension of a peptidomimetic by
N-acylation of an aza-amino acid residue is hampered by the
low reactivity of the semicarbazide nitrogen. Such coupling
difficulties have previously been addressed by prolonged
coupling time20 mixed coupling reagents and double
coupling.7,13 In the present study, we have investigated the
ability of N-acylbenzotriazoles to acylate hydrochloride or p-
TsOH salts of free-amino azadipeptides 29a−c. Salts 29a−c
were prepared in 90−100% yields according to Scheme 7.
Azadipeptide 29c was conveniently acylated with N-(N-Pg-α-
aminoacyl)benzotriazoles 30a,b in acetonitrile/water (3:1 v/v)
in the presence of 2.0 equiv of Et3N to give azatripeptides
31a,b. However, azapeptides 29a,b were recovered unreacted
upon treatment with 30a,b under the same reaction conditions,
and hydrolysis of 30a was the only reaction observed when the
coupling between 29a and 30a was attempted under microwave
irradiation (1 h, 50 W, 70 °C) (Scheme 7).
Alternative Facile Route to the Synthesis of N-Pg-
azatripeptides 33a,b and 35a,b. The low reactivity of 29a,b
is due to greater stabilization of the amide electron delocalized
contributing structure by the N-alkyl groups thus inductively
reduces the electron density at the terminal amino group and
makes it less nucleophilic. Also steric hindrance arising from
substitution on adjacent nitrogen could play a part. To bypass
the low coupling rates of the aza-amino acid residue, we
developed a novel route to azatripeptides: first we converted N-
(N-Pg-α-aminoacyl)benzotriazoles 30a,c−e to the correspond-
ing hydrazides 32a−d. Coupling active carbamates 16b,d with
hydrazides 32a,b furnished azatripeptides 33a,b containing an
azaglycine residue. Alternatively, hydrazides 32c,d were
benzylated by reductive amination to give N′-benzyl-N-(N-
Pg-glycyl)hydrazines 34a,b which upon coupling with 16b,d
gave azatripeptides 35a,b (Scheme 8).
Solution-Phase Synthesis of Hybrid Azapeptide 40:
Analogue of Leu-enkephalin. Leu-enkephalin is an
endogenous opioid peptide neurotransmitter found naturally
in the brains of many animals, including humans. The amino
acid sequence of Leu-enkephalin is Tyr-Gly-Gly-Phe-Leu.
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Scheme 9. Solution-Phase Synthesis of Azapeptide Analogue of Leu-enkephalin 40
despsipeptide H-Gly-OPhe.HCl 23b was prepared according to the
literature method.16
Azapeptide mimetics of Leu-enkephalin were synthesized, and
their binding affinity was examined in the context of
monoclonal antibody 3-E7 known to strongly bind the
[Leu5] enkephalin sequence.21 Our new route to the synthesis
of azatripeptides (Scheme 8) inserted the aza-amino acid in the
middle of azatripeptides, thus enabling the synthesis of the yet
unknown AzaLeu-enkephalin analogue 40 by segment con-
densation of free-amino azatripeptide fragment 36 and the
benzotriazolide 39 (Scheme 9).
Fmoc-NH-NH2 (8b). Compound 8b was prepared according to the
literature method:11 white microcrystals (8.756 g, 89%); mp 170.0−
172.0 °C (lit.11 mp 172.0−173.0 °C); H NMR (300 MHz, DMSO-
1
d6) δ 8.36 (br s, 1H), 7.90 (d, J = 7.8 Hz, 1H), 7.70 (d, J = 7.2 Hz,
2H), 7.42 (t, J = 6.9 Hz, 1H), 7.33 (t, J = 6.9 Hz, 1H), 4.36−4.16 (m,
3H), 4.09 (br s, 2H); 13C NMR (75 MHz, DMSO-d6) δ 158.2, 143.8,
140.7, 127.6, 127.0, 125.2, 120.1, 65.6, 46.7.
Boc-NH-NCMe2 (9a). Compound 9a was prepared according to
the literature method:13 white microcrystals (1.156 g, 89%); mp
102.0−104.0 °C (lit.22 mp 103.0−104.0 °C); H NMR (300 MHz,
1
CONCLUSIONS
■
CDCl3) δ 7.43 (br s, 1H), 2.05−1.98 (m, 3H), 1.83−1.76 (m, 3H),
1.52−1.45 (m, 9H); 13C NMR (75 MHz, CDCl3) δ 152.8, 149.7, 80.8,
28.2, 25.3, 15.9.
In conclusion, a mild and efficient method for the preparation
of N-Pg-azatripeptides, N-Pg-azatetrapeptides, hybrid N-Pg-
azatripeptides, and hybrid N-Pg-azatetrapeptide has been
developed by reacting N-(N-Pg-azadipeptidoyl)benzotriazoles
with α-aminoacids, α-dipeptides, α-aminoxyacetic acid, a
depsidipeptide, and α-hydroxy-β-phenylpropionic acid. These
azatripeptides and tetrapeptides could be valuable building
blocks for the synthesis of longer or cyclic azapeptides. In
addition, N-(α-aminoacyl)benzotriazoles were easily converted
to N-Pg-α-amino acid hydrazides which were used either
directly or after alkylation to construct N-Pg-azatripeptides,
thus inserting an aza-amino acid in the middle of the
azatripeptide unit. An aza-leu-enkephalin analogue was
synthesized by adopting this novel protocol. The new method
avoids the low coupling rates of aza amino acids and provides
an excellent alternative to the construction of azapeptides
starting from N-Boc- and N-Fmoc hydrazides.
Boc-NH-NCHPh (9b). Compound 9b was prepared according to
the literature method:13 white microcrystals (2.980 g, 89%); mp 199−
202 °C (lit.23 mp 203 °C); 1H NMR (300 MHz, DMSO-d6) δ 8.00 (br
s, 1H), 7.69−7.49 (m, 2H), 7.48−7.28 (m, 3H), 1.47 (s, 9H); 13C
NMR (75 MHz, DMSO-d6) δ 152.3, 143.0, 134.6, 129.3, 128.7, 126.4,
79.4, 28.1.
Fmoc-NH-NCMe2 (9c). To a solution of benzaldehyde (10.0
mmol, 1.0 equiv) in diethyl ether (50 mL) were added (9H-fluoren-9-
yl)methyl hydrazinecarboxylate 8b (10.0 mmol, 1.0 equiv) and two
drops of glacial acetic acid. The reaction mixture was heated under
reflux for 2−3 h and then cooled to room temperature. The white
solid precipitate was collected by filtration, washed with cold diethyl
ether, and dried under vacuum to yield 9c. Compound 9c was
characterized by 1H and 13C NMR and taken to the next step without
further purification: white microcrystals (2.796 g, 95%); mp 147.0−
1
149.0 °C; H NMR (300 MHz, CDCl3) δ 7.76 (d, J = 7.7 Hz, 2H),
7.63 (d, J = 7.1 Hz, 2H), 7.39 (t, J = 7.5 Hz, 2H), 7.30 (t, J = 7.5 Hz,
2H), 4.52 (d, J = 7.2 Hz, 2H), 4.30 (t, J = 7.1 Hz, 1H), 2.07 (d, J = 2.1
Hz, 3H), 1.84 (d, J = 3.0 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ
151.1, 148.8, 143.6, 141.2, 127.7, 127.0, 125.1, 119.9, 67.2, 47.0, 25.4,
EXPERIMENTAL SECTION
■
Melting points were determined on a capillary melting point apparatus
equipped with a digital thermometer and are uncorrected. NMR
spectra were recorded in acetone-d6, CDCl3, and DMSO-d6 with TMS
for 1H (300 MHz) and 13C (75 MHz) as an internal reference. DCM
was dried and distilled over CaH2, whereas tetrahydrofuran (THF)
was used after distillation over Na-benzophenone. Carbonyldiimida-
zole (CDI), Boc-hydrazide 8a, (Boc)2O 11, L-amino methyl/tert-butyl
ester hydrochloride 15a−e, amino acids 21a−e, free dipeptides H-Gly-
Gly-OH 22a, H-Gly-Phe-OH 22b, α-aminoxyacetic acid 23a, and α-
hydroxy-β-phenylpropionic acid 27 were purchased from chemical
supply companies and used without further purification. Free
1
16.2. Compound 9c was partially characterized by H and 13C NMR
and taken to the next step without further purification.
tert-Butyl 2-Isopropylhydrazinecarboxylate (10a). Compound
10a was prepared according to the literature method:13 white
microcrystals (0.364 g, 70%); mp 45.0−47.0 °C (lit.13 mp 47.0−
1
49.0 °C); H NMR (300 MHz, CDCl3) δ 6.42 (br s, 1H), 3.19−2.97
(m, 1H), 1.43 (s, 9H), 1.00 (d, J = 6.6 Hz, 3H), 0.98 (d, J = 6.3 Hz,
3H); 13C NMR (75 MHz, CDCl3) δ 156.8, 80.2, 50.7, 28.3, 20.5.
tert-Butyl 2-Benzylhydrazinecarboxylate (10b). Compound
10b was prepared according to the literature method:13 colorless oil
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(0.310 g, 75%); lit.13 reported as low melting solid; H NMR (300
MHz, CDCl3) δ 7.41−7.21 (m, 5H), 6.13 (br s, 1H), 4.21 (br s, 1H),
3.99 (s, 2H), 1.46 (s, 9H); 13C NMR (75 MHz, CDCl3) δ 156.6,
137.6, 128.9, 128.4, 127.4, 80.5, 55.8, 28.3.
1.0, CH3OH); H NMR (300 MHz, CDCl3) δ 8.09 (d, J = 1.1 Hz,
1H), 7.34 (s, 1H), 7.04 − 6.97 (m, 1H), 6.99 − 6.89 (m, 1H), 4.58 −
4.43 (m, 1H), 1.76 − 1.52 (m, 4H), 1.45 (s, 12H), 0.94 (d, J = 4.2 Hz,
4H), 0.91 (d, J = 4.3 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ 170.7,
148.7, 136.0, 130.5, 115.8, 83.0, 59.0, 53.4, 31.4, 28.0, 18.8, 17.8.
Compound 16e was partially characterized by 1H, 13C NMR and taken
to the next step without further purification.
1
1
(9H-Fluoren-9-yl)methyl 2-Isopropylhydrazinecarboxylate
(10c). Compound 10c was prepared according to the literature
method:11 white microcrystals (0.786 g, 68%); mp 161.0−163.0 °C
(lit.11 mp 163.0−164.0 °C); H NMR (300 MHz, DMSO-d6) δ 8.63
1
General Methods for the Preparation of N-Pg-azadipeptide
18a−h. The residue 16a−e (1.0 equiv) was dissolved in dry DCM (20
mL) and reacted with N′-alkyl-N-Pg-hydrazines 10a−c or 14a,b (1.0
equiv; Pg = Boc-, Fmoc-, Cbz-) in the presence of DIPEA (1.0 equiv)
at 20 °C overnight. The reaction mixture was poured into a separatory
funnel, washed with water (2 × 20 mL), 2 N HCl (3 × 20 mL), brine
solution (2 × 20 mL), dried over MgSO4, and evaporated under
vacuum to give both −NH2 and −CO2H side protected azadipeptide
18a−h, which were used in the next step without further purification.
Cbz-AzaAla-Phe-OMe (18a). Compound 18a was prepared
according to the given general procedure for 18a−h: white
microcrystals (1.734 g, 90%); mp 51.0−53.0 °C (lit.28 mp not
reported); [α]20D −25 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3)
δ 7.31 (s, 5H), 7.27−7.11 (m, 3H), 7.02 (dd, J = 7.2, 2.4 Hz, 2H), 6.86
(s, 1H), 5.77 (d, J = 8.1 Hz, 1H), 5.13 (s, 2H), 4.72−4.64 (m, 1H),
3.62 (s, 3H), 3.13−2.94 (m, 5H); 13C NMR (75 MHz, CDCl3) δ
172.6, 157.0, 155.2, 135.9, 135.3, 129.2, 128.6, 128.5, 128.4, 128.1,
126.9, 67.9, 54.2, 52.1, 38.1, 35.8. Anal. Calcd for C20H23N3O5: C,
62.33; H, 6.01; N, 10.90. Found: C, 62.42; H, 6.17; N, 10.84.
Cbz-AzaAla-Val-OtBu (18b). Compound 18b was prepared
according to the given procedure for 18a: low melting solid (1.556
g, 82%); [α]20D −22.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3)
δ 7.47−7.18 (m, 5H), 7.07 (s, 1H), 5.81 (d, J = 8.8 Hz, 1H), 5.18 (s,
2H), 4.28 (dd, J = 8.8, 4.5 Hz, 1H), 3.10 (s, 3H), 2.20−1.96 (m, 1H),
1.43 (s, 9H), 0.87 (d, J = 6.0 Hz, 3H), 0. 78 (d, J = 6.6 Hz, 3H); 13C
NMR (75 MHz, CDCl3) δ 171.8, 163.2, 157.4, 135.3, 128.5, 128.4,
128.2, 81.7, 67.9, 58.4, 35.9, 31.5, 28.0, 18.8, 17.3. Compound 18b was
(br s, 1H), 7.84 (d, J = 7.4 Hz, 2H), 7.65 (d, J = 7.5 Hz, 2H), 7.36 (t, J
= 7.5 Hz, 2H), 7.27 (t, J = 7.4 Hz, 2H), 4.42−4.21 (m, 3H), 4.18 (d, J
= 6.5 Hz, 1H), 3.05−2.82 (m, 1H), 0.86 (d, J = 5.6 Hz, 6H); 13C
NMR (75 MHz, DMSO-d6) δ 156.8, 143.8, 140.7, 127.6, 127.0, 125.2,
120.1, 65.4, 49.5, 46.7, 20.6.
Benzyl 2-Methylhydrazinecarboxylate Hydrochloride (14a).
Compound 14a was prepared according to the literature method:24
white microcrystals (3.00 g, 70%); mp 174.0−177.0 °C (lit.24 mp
1
175.0−177.0 °C); H NMR (300 MHz, DMSO-d6) δ 10.76 (s, 1H),
7.55−7.05 (m, 5H), 5.13 (s, 2H), 2.67 (s, 3H); 13C NMR (75 MHz,
DMSO-d6) δ 155.4, 136.3, 129.1, 129.0, 128.7, 67.9, 36.4.
(9H-Fluoren-9-yl)methyl 2-methylhydrazinecarboxylate Hy-
drochloride (14b). Compound 14b was prepared according to the
literature method:25 white microcrystals (0.850 g, 73%); mp 160.0−
161.0 °C (lit.25 mp 160.0 °C); H NMR (300 MHz, DMSO-d6) δ
1
10.79 (br s, 1H), 7.90 (d, J = 7.4 Hz, 2H), 7.72 (d, J = 7.4 Hz, 2H),
7.43 (t, J = 7.5 Hz, 2H), 7.34 (t, J = 7.4 Hz, 2H), 4.52 (d, J = 6.5 Hz,
2H), 4.29 (t, J = 6.6 Hz, 1H), 2.70 (s, 3H); 13C NMR (75 MHz,
DMSO-d6) δ 155.5, 143.9, 141.4, 128.4, 127.8, 125.8, 120.9, 67.7, 47.0,
36.3.
General Methods for the Preparation of 16a−e. To a
suspension of L-amino methyl/tert-butyl ester hydrochloride 15a−e
(10.0 mmol, 1.0 equiv) in DCM (20 mL) at 20 °C were added 2.5
equiv of DIPEA and CDI (carbonyldiimidazole, 1.1 equiv). The
reaction mixture was stirred for 3 h at rt, and the organic layer was
washed with water (2 × 20 mL), NaHCO3 (3 × 20 mL) and brine
solution (2 × 20 mL). The organic layer was dried over MgSO4 and
evaporated under vacuum to give oily mono substituted imidazole
derivative 16a−e.
1
partially characterized by H, 13C NMR and taken to the next step
without further purification.
Boc-AzaVal-Leu-OMe (18c). Compound 18c was prepared
Im-Phe-OMe (16a). Compound 16a was prepared according to the
given general procedure for 16a−e: colorless oil26 (2.596 g, 95%);
[α]20D −20.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ 8.02 (s,
1H), 7.34−7.17 (m, 3H), 7.15−7.03 (m, 2H), 7.00 (d, J = 1.7 Hz,
1H), 6.71 (d, J = 8.1 Hz, 1H), 4.90−4.80 (m, 1H), 3.75 (s, 3H), 3.30−
3.11 (m, 2H); 13C NMR (75 MHz, CDCl3) δ 171.8, 148.7, 136.3,
130.4, 129.3, 129.0, 128.6, 127.6, 116.3, 55.0, 53.0, 37.6.
according to the given procedure for 18a: white microcrystals (1.537
1
g, 89%); mp 60.0−62.0 °C; [α]20 −27.0 (c 1.0, CH3OH); H NMR
D
(300 MHz, CDCl3) δ 6.65 (br s, 1H), 5.57 (d, J = 9.1 Hz, 1H), 4.66−
4.53 (m, 1H), 4.47 (q, J = 7.7 Hz, 1H), 3.68 (s, 3H), 1.76−1.48 (m,
3H), 1.44 (s, 9H), 1.09 (d, J = 6.6 Hz, 3H), 1.04 (d, J = 5.7 Hz, 3H),
0.95−0.83 (m, 6H); 13C NMR (75 MHz, CDCl3) δ 174.9, 157.0,
156.2, 82.0, 52.4, 51.9, 48.1, 41.8, 28.2, 24.9, 23.2, 22.0, 19.9, 19.5.
Anal. Calcd for C16H31N3O5: C, 55.63; H, 9.05, N, 12.16. Found: C,
55.99; H, 9.18; N, 12.09.
Im-Val-OtBu (16b). Compound 16b was prepared according to the
given procedure for 16a: oil (2.406 g, 90%); [α]20 −28.0 (c 1.0,
D
1
CH3OH); H NMR (300 MHz, CDCl3) δ 8.15 (s, 1H), 7.38 (d, J =
Boc-AzaVal-Met-OMe (18d). Compound 18d was prepared
according to the given procedure for 18a: colorless oil (1.636 g,
90%); [α]20D −35.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ
6.26 (br s, 1H), 5.90 (br s, 1H), 4.75−4.42 (m, 2H), 3.74 (s, 3H), 2.54
(t, J = 7.5 Hz, 2H), 2.29−1.90 (m, 5H), 1.49 (s, 9H), 1.11 (d, J = 5.7
Hz, 3H), 1.08 (d, J = 7.8 Hz, 3H). Compound 18d was partially
1.5 Hz, 1H), 7.06 (d, J = 1.8 Hz, 1H), 6.69 (d, J = 8.1 Hz, 1H), 4.45
(dd, J = 8.1, 4.8 Hz, 1H), 2.30−2.18 (m, 1H), 1.48 (s, 9H), 0.99 (d, J =
6.9 Hz, 3H), 0.96 (d, J = 6.9 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ
170.7, 148.7, 136.0, 130.5, 115.8, 83.0, 59.0, 31.4, 28.0, 18.8, 17.8.
Compound 16b was partially characterized by 1H, 13C NMR and taken
to the next step without further purification.
1
characterized by H NMR and taken to the next step without further
Im-Met-OMe (16c). Compound 16c was prepared according to the
given procedure for 16a; colorless oil27 (2.419 g, 94%); [α]20D −19 (c
1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ 8.19 (s, 1H), 7.73 (d, J
= 7.8 Hz, 1H), 7.44 (d, J = 1.2 Hz, 1H), 7.04 (d, J = 1.5 Hz, 1H),
4.79−4.71 (m, 1H), 3.77 (s, 3H), 2.62−2.55 (m, 2H), 2.31−2.01 (m,
5H); 13C NMR (75 MHz, CDCl3) δ 171.9, 148.7, 136.1, 130.1, 116.2,
60.4, 53.1, 30.4, 30.1, 15.4.
purification.
Boc-AzaPhe-Leu-OMe (18e). Compound 18e was prepared
according to the given procedure for 18a: low melting solid (1.869
g, 95%); lit.6 mp not reported; [α]D −25.0 (c 1.0, CH3OH); H
NMR (300 MHz, CDCl3) δ 7.47−7.18 (m, 6H), 6.20 (s, 1H), 5.82 (d,
J = 8.6 Hz, 1H), 4.60−4.51 (m, 1H), 3.75−3.65 (m, 5H), 1.70−1.51
(m, 3H), 1.44 (s, 9H), 0.96 (d, J = 6.0 Hz, 3H), 0.94 (d, J = 6.3 Hz,
3H); 13C NMR (75 MHz, CDCl3) δ 174.1, 157.1, 154.2, 135.9, 128.8,
128.6, 127.7, 82.2, 51.8, 41.8, 27.9, 24.7, 22.9, 21.8.
20
1
Im-Leu-OMe (16d). Compound 16d was prepared according to the
given procedure for 16a: white microcrystals (2.297 g, 96%); mp
80.0−82.0 °C (lit.27 mp 80.0−81.0 °C); [α]20 −47.0 (c 1.0,
Boc-AzaGly-Phe-OMe (18f). Compound 18f was prepared
according to the given procedure for 18a: colorless oil (1.350 g,
80%); [α]20D −16.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ
7.29−7.17 (m, 3H), 7.13−7.05 (m, 2H), 6.71 (s, 1H), 5.91 (d, J = 8.1
Hz, 1H), 5.50 (d, J = 8.2 Hz, 1H), 4.80−4.71 (m, 1H), 3.65 (d, J = 7.2
Hz, 3H), 3.07 (d, J = 5.7 Hz, 1H), 3.01 (d, J = 6.0 Hz, 1H), 1.44 (s,
9H); 13C NMR (75 MHz, CDCl3) δ 172.9, 158.3, 156.3, 136.3, 129.5,
128.6, 127.0, 81.8, 54.1, 52.4, 38.4, 28.3. Compound 18f was partially
D
1
CH3OH); H NMR (300 MHz, CDCl3) δ 8.15 (s, 1H), 8.00 (d, J
= 6.0 Hz, 1H), 7.44 (d, J = 3.0 Hz, 1H), 6.95 (d, J = 3.0 Hz, 1H),
4.69−4.39 (m, 1H), 3.69 (s, 3H), 1.82−1.41 (m, 3H), 0.89 (d, J = 6.0
Hz, 3H), 0.86 (d, J = 6.0 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ
173.3, 148.9, 136.1, 129.7, 116.4, 52.5, 52.3, 40.4, 24.8, 22.8, 21.4.
Im-Leu-OtBu (16e). Compound 16e was prepared according to the
given procedure for 16a: colorless oil (2.589 g, 92%); [α]20D −23.0 (c
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1
characterized by H, 13C NMR and taken to the next step without
128.6, 128.4, 68.1, 58.7, 36.2, 31.7, 19.1, 17.6. Compound 19b was
1
partially characterized by H, 13C NMR and taken to the next step
further purification.
Fmoc-AzaAla-Val-OtBu (18g). Compound 18g was prepared
according to the given procedure for 18a: low melting solid (1.987
g, 85%); [α]20D −10.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3)
δ 7.75 (d, J = 7.5 Hz, 2H), 7.58 (t, J = 6.1 Hz, 2H), 7.39 (t, J = 7.5 Hz,
2H), 7.31 (t, J = 7.5 Hz, 2H), 7.13 (s, 1H), 5.85 (d, J = 7.8 Hz, 1H),
4.62−4.44 (m, 2H), 4.30 (dd, J = 8.1, 3.8 Hz, 1H), 4.23 (t, J = 6.9 Hz,
1H), 3.09 (s, 3H), 2.20−2.00 (m, 1H), 1.42 (s, 9H), 0.91 (d, J = 6.9
Hz, 3H), 0.84 (d, J = 6.6 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ
171.9, 157.4, 155.5, 143.2, 141.2, 127.8, 127.1, 124.9, 120.0, 81.8, 67.9,
58.5, 46.9, 35.9, 31.5, 28.0, 18.9, 17.5. Compound 18g was partially
characterized by 1H and 13C NMR and taken to the next step without
further purification.
without further purification.
Boc-AzaVal-Leu-OH (19c). Compound 19c was prepared from 18c
according to the given method for 19a: white microcrystals (1.047 g,
79%); mp 56.0−58.0 °C; [α]20 −25.0 (c 1.0, CH3OH); H NMR
1
D
(300 MHz, DMSO-d6) δ 8.81 (s, 0.6H), 8.39 (s, 0.3H), 6.05−5.90 (m,
1H), 4.50−4.25 (m, 1H), 4.24−3.97 (m, 1H), 1.80−1.46 (m, 3H),
1.41 (s, 9H), 0.97 (d, J = 6.4 Hz, 6H), 0.88−0.83 (m, 6H); 13C NMR
(75 MHz, DMSO-d6) δ 174.9, 156.6, 155.4, 79.4, 51.3, 47.2, 27.9, 24.0,
23.0, 22.1, 21.5, 20.1, 19.4; Anal. Calcd for C15H29N3O5: C, 54.36; H,
8.82, N, 12.68. Found: C, 54.02; H, 9.15; N, 12.30.
Boc-AzaVal-Met-OH (19d). Compound 19d was prepared from
18d according to the given method for 19a: low melting point solid
Fmoc-AzaVal-Leu-OtBu (18h). Compound 18h was prepared
according to the given procedure for 18a: low melting solid (1.988
g, 78%); [α]20D −4.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ
7.77 (d, J = 7.5 Hz, 2H), 7.60 (t, J = 6.6 Hz, 2H), 7.41 (t, J = 7.4 Hz,
2H), 7.32 (t, J = 7.4 Hz, 2H), 6.81−6.48 (m, 0.5H), 5.67−5.42 (m,
0.5H), 4.63−4.36 (m, 3H), 4.31−4.07 (m, 1H), 1.78−1.50 (m, 3H),
1.45 (s, 9H), 1.07 (d, J = 6.7 Hz, 6H), 1.00−0.84 (m, 6H); 13C NMR
(75 MHz, CDCl3) δ 173.7, 164.9, 157.0, 143.6, 141.6, 128.1, 127.4,
125.3, 120.3, 81.9, 52.7, 48.6, 47.3, 42.5, 28.2, 25.1, 23.0, 22.4, 19.6.
Anal. Calcd for C29H39N3O5: C, 68.34; H, 7.71, N, 8.24. Found: C,
68.04; H, 8.14; N, 8.47.
(1.202 g, 86%); [α]20 −20.0 (c 1.0, CH3OH); H NMR (300 MHz,
1
D
DMSO-d6) δ 8.84 (br s, 1H), 8.43 (br s, 1H), 6.60−6.12 (m, 1H),
4.52−3.98 (m, 2H), 2.44 (d, J = 7.4 Hz, 2H), 2.11−1.86 (m, 5H), 1.42
(s, 9H), 0.99 (d, J = 6.3 Hz, 6H). Compound 19d was partially
1
characterized by H NMR and taken to the next step without further
purification.
Boc-AzaPhe-Leu-OH (19e). Compound 19e was prepared from 18e
according to the given method for 19a: white microcrystals (1.351 g,
89%); mp 60.0−62.0 °C; [α]20 −22.0 (c 1.0, CH3OH); H NMR
1
D
(300 MHz, CDCl3) δ 7.46−7.11 (m, 5H), 5.94 (d, J = 8.0 Hz, 1H),
4.60−4.38 (m, 1H), 1.78−1.49 (m, 3H), 1.40 (s, 9H), 0.96 (d, J = 6.6
Hz, 4H), 0.94 (d, J = 6.6 Hz, 3H); 13C NMR (75 MHz, acetone-d6) δ
174.1, 158.0, 154.4, 137.7, 128.9, 128.4, 127.4, 80.5, 52.0, 41.4, 27.6,
24.6, 22.8, 21.3. Compound 19e was partially characterized by 1H and
13C NMR.
Cbz-AzaAsp(OMe)-Val-OtBu (18i). Compound 18i was prepared
according to the given procedure for 18a: low melting solid (2.012 g,
92%); [α]20D −21.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ
7.38−7.18 (m, 5H), 5.98 (d, J = 8.7 Hz, 1H), 5.13 (s, 2H), 4.22 (dd, J
= 8.7, 4.5 Hz, 1H), 3.67−3.57 (m, 5H), 2.27 −1.71 (m, 1H), 1.38 (s,
9H), 0.83 (d, J = 6.8 Hz, 3H), 0.77 (d, J = 7.1 Hz, 3H); 13C NMR (75
MHz, CDCl3) δ 171.5, 170.2, 156.9, 155.4, 135.4, 128.8, 128.7, 128.4,
82.0, 68.3, 58.8, 52.4, 52.2, 31.9, 28.2, 19.0, 17.7; HRMS (ESI) calcd
for C21H31N3O7Na [M + Na]+ 460.2054, found 460.2075.
Boc-AzaGly-Phe-OH (19f). Compound 19f was prepared from 18f
according to the given method for 19a: white microcrystals (0.983 g,
76%); mp 159.0−161.0 °C (lit.13 mp 163.0− 164.0 °C); [α]20D −69.0
(c 1.0, CH3OH); 1H NMR (300 MHz, DMSO-d6) δ 12.77 (br s, 1H),
8.55 (s, 1H), 7.84 (s, 1H), 7.42−7.06 (m, 5H), 6.26 (d, J = 7.6 Hz,
1H), 4.45−4.32 (m, 1H), 3.08−2.86 (m, 2H), 1.39 (s, 9H); 13C NMR
(75 MHz, DMSO-d6) δ 173.3, 157.5, 155.9, 137.2, 129.3, 128.2, 126.5,
79.0, 53.7, 37.4, 28.1. Anal. Calcd for C15H20LiN3O5: C, 54.71; H,
6.12, N, 12.76. Found: C, 54.70; H, 6.41; N, 12.30.
General Methods for the −OMe and −OtBu Group
Deprotection. Method 1. −OMe deprotection: N-Pg-AzaAA1-AA2-
OMe (4.0 mmol, 1.0 equiv) was dissolved in a solution of MeOH and
water (10 mL, 9:1 v/v). LiOH (8.0 mmol, 2.0 equiv) was added, and
the mixture was stirred for 2 h at room temperature. After completion
of the reaction (followed by TLC), MeOH was evaporated, water (5.0
mL) was added, and the aqueous layer was washed with ether (2 × 20
mL) and then acidified with 2 N HCl. The aqueous layer was extracted
with EtOAc (2 × 10 mL) solution, and the organic layer was dried
over anhydrous MgSO4 and evaporated to give N-Pg-azadipeptide.
Method 2. −OtBu deprotection: N-Pg-AzaAA1-AA2-OtBu (4.0
mmol) was dissolved in a solution of dry DCM (10 mL) containing
trifluoroacetic acid (5.0 mL) and stirred for 2 h at room temperature.
After completion of the reaction [followed by TLC] and TFA were
evaporated, 5 mL of water was added, the aqueous layer was extracted
with EtOAc (10 mL × 2) solution, and the organic layer was dried
over anhydrous MgSO4 and evaporated to give white solid.
Compounds 19b,d were characterized by 1H and 13C NMR and
taken to the next step without further purification.
Cbz-AzaAla-Phe-OH (19a). Compound 19a was prepared from 18a
according to the given method 1 for −OMe deprotection. White
microcrystals (1.337 g, 90%); mp 58.0− 60.0 °C; [α]D20 −30.0 (c 1.0,
CH3OH); 1H NMR (300 MHz, DMSO-d6) δ 12.73 (br s, 1H), 9.50−
9.39 (m, 1H), 7.47−7.29 (m, 5H), 7.27−7.15 (m, 5H), 6.67−6.50 (m,
1H), 5.13 (s, 2H), 4.36−4.28 (m, 1H), 3.00 (d, J = 6.5 Hz, 2H), 2.93
(s, 3H); 13C NMR (75 MHz, DMSO-d6) δ 173.5, 157.2, 155.5, 137.6,
136.3, 129.3, 128.5, 128.3, 128.1, 127.9, 126.4, 66.4, 54.5, 39.9, 36.9;
Anal. Calcd for C19H21N3O5: C, 61.45; H, 5.70, N, 11.31. Found: C,
61.13; H, 5.90; N, 11.18.
Fmoc-AzaAla-Val-OH (19g). Compound 19g was prepared from
18g according to the given method for 19b: white microcrystals (1.136
g, 69%); mp 178.0−180.0 °C; [α]20D −7.0 (c 1.0, CH3OH); 1H NMR
(300 MHz, CDCl3) δ 7.71 (d, J = 7.5 Hz, 2H), 7.53 (d, J = 7.1 Hz,
2H), 7.35 (t, J = 7.0 Hz, 2H), 7.29−7.18 (m, 2H), 4.57−4.30 (m, 1H),
4.18 (t, J = 6.8 Hz, 2H), 3.03 (s, 3H), 2.23−2.00 (m, 1H), 0.89 (d, J =
6.3 Hz, 3H), 0.82 (d, J = 6.6 Hz, 3H); 13C NMR (75 MHz, DMSO-d6)
δ 174.1, 158.0, 156.1, 144.1, 141.4, 128.3, 127.7, 125.8, 120.8, 58.9,
47.2, 36.1, 31.3, 30.9, 19.7, 18.6. Compound 19g was partially
1
characterized by H and 13C NMR.
Fmoc-AzaVal-Leu-OH (19h). Compound 19h was prepared from
18h according to the given method for 19b: white microcrystals (1.451
g, 80%); mp 157.0−159.0 °C; [α]20 −16.0 (c 1.0, CH3OH); H
1
D
NMR (300 MHz, DMSO-d6) δ 9.25 (d, J = 10.9 Hz, 1H), 7.90 (d, J =
7.2 Hz, 2H), 7.76 (d, J = 7.3 Hz, 2H), 7.42 (t, J = 7.2 Hz, 2H), 7.33 (t,
J = 7.1 Hz, 2H), 6.40−6.20 (m, 1H), 6.29 (dd, J = 33.4, 8.1 Hz, H),
4.65−3.93 (m, 5H), 1.80−1.12 (m, 3H), 0.97 (dd, J = 13.2, 5.7 Hz,
6H), 0.82 (d, J = 6.7 Hz, 3H); 13C NMR (75 MHz, DMSO-d6) δ
174.9, 156.9, 156.5, 140.7, 127.7, 127.1, 125.3, 120.1, 66.1, 64.9, 51.4,
47.6, 46.6, 24.2, 22.9, 21.5, 19.8, 19.4. Anal. Calcd for C25H31N3O5: C,
66.21; H, 6.89, N, 9.26. Found: C, 65.90; H, 6.98; N, 9.21.
General Method for the Preparation of N-(N-Pg-
azadipeptidoyl)benzotriazoles 20a−e. Benzotriazole (6.0 mmol,
3.0 equiv) was dissolved in dry DCM (50 mL). SOCl2 (2.2 mmol) was
added by syringe, and the mixture was stirred for 15 min at rt under
argon. The solution temperature was lowered to −30 to −40 °C (dry
ice + acetone), and 2.0 equiv of TEA was added. After 5 min of
stirring, N-Pg-AzaAA1-AA2-OH (2.0 mmol) was added and the
mixture stirred for 2 h keeping the temperature at −30 to −40 °C.
After completion of the reaction, ice-cold water (20 mL) was added,
and the organic layer was washed with water (20 mL × 2), NaHCO3
Cbz-AzaAla-Val-OH (19b). Compound 19b was prepared from 18b
according to the given method 2 for −OtBu deprotection: white
microcrystals (1.074 g, 83%); mp 65.0−67.0 °C; [α]20 −15.0 (c 1.0,
D
CH3OH); 1H NMR (300 MHz, CDCl3) δ 7.63−7.41 (m, 1H), 7.34 (s,
5H), 5.99−5.90 (m, 1H), 5.19 (s, 2H), 4.30−4.20 (m, 1H), 3.09 (s,
3H), 2.18−2.07 (m, 1H), 0.89 (d, J = 7.5 Hz, 3H), 0.80 (d, J = 6.6 Hz,
3H); 13C NMR (75 MHz, CDCl3) δ 172.1, 163.4, 157.7, 135.6, 128.8,
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(20 mL × 4), and then brine (20 mL × 2). The organic layer was dried
over MgSO4 and evaporated to yield a white solid 20a−e.
solution (4 × 10 mL), water (10 mL), and brine (10 mL). The solvent
was dried over MgSO4 and evaporated to give various N-Pg-aza-tri-
and -tetrapeptides.
Cbz-AzaAla-Phe-Bt (20a). Compound 20a was prepared according
to the general method for the preparation of N-(N-Pg-azadipeptidoyl)-
benzotriazoles 20a−e: white microcrystals (0.822 g, 87%); mp 70.0−
72.0 °C; [α]20D −15.0 (c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3)
δ 8.04 (d, J = 8.4 Hz, 1H), 8.01 (d, J = 8.1 Hz, 1H), 7.51 (t, J = 7.8 Hz,
1H), 7.40 (t, J = 7.7 Hz, 1H), 7.30−7.15 (m, 5H), 7.09−6.92 (m, 5H),
6.14 (d, J = 7.8 Hz, 1H), 6.04−5.98 (m, 1H), 5.01 (s, 2H), 3.30 (dd, J
= 14.0, 5.2 Hz, 1H), 3.12 (dd, J = 14.0, 7.5 Hz, 1H), 2.98 (s, 3H); 13C
NMR (75 MHz, CDCl3) δ 171.5, 157.2, 155.3, 145.7, 135.2, 130.8,
130.5, 129.0, 128.5, 128.3, 127.9, 127.1, 126.3, 120.1, 114.1, 67.7, 55.1,
38.2, 35.7. Anal. Calcd for C25H24N6O4: C, 63.55; H, 5.12, N, 17.79.
Found: C, 63.23; H, 5.14; N, 18.00.
Cbz-AzaAla-Phe-Cys-OH (24a). Compound 24a was prepared
according to the general procedure for the coupling of N-(N-Pg-
azadipeptidoyl)benzotriazoles 20a−e with various nucleophiles: white
microcrystals (0.379 g, 80%); mp 89.0−92.0 °C; [α]20 −25.0 (c 1.0,
D
CH3OH); 1H NMR (300 MHz, DMSO-d6) δ 9.47 (br s, 1H), 8.25 (br
s, 1H), 7.48−7.33 (m, 5H), 7.28−7.08 (m, 6H), 6.62 (br s, 1H), 5.13
(s, 2H), 4.70−4.30 (m, 2H), 3.06−2.78 (m, 7H); 13C NMR (75 MHz,
DMSO-d6) δ 171.5, 171.4, 157.1, 155.6, 137.7, 136.2, 129.4, 129.2,
128.5, 128.1, 127.9, 126.2, 66.4, 54.6, 54.4, 37.6, 35.5, 25.7. Anal. Calcd
for C22H26N4O6S: C, 55.68; H, 5.52; N, 11.81. Found: C, 55.56; H,
5.64; N, 11.78.
Cbz-AzaAla-Val-Ser-OH (24b). Compound 24b was prepared
Cbz-AzaAla-Val-Bt (20b). Compound 20b was prepared according
according to the given method for 24a: white microcrystals (0.328
to the given method for 20a: white microcrystals (0.688 g, 81%); mp
1
g, 80%); mp 50.0−52.0 °C; [α]20 −5.0 (c 1.0, CH3OH); H NMR
1
57.0−59.0 °C; [α]20 −12.0 (c 1.0, CH3OH); H NMR (300 MHz,
D
D
(300 MHz, CDCl3) δ 8.07 (br s, 1H), 7.95−7.70 (m, 1H), 7.35−7.25
(m, 6H), 5.24−5.07 (m, 2H), 4.67−4.54 (m, 1H), 4.33−4.16 (m, 1H),
4.06−3.81 (m, 2H), 3.03 (s, 3H), 2.20−1.92 (m, 1H), 0.85 (d, J = 6.3
Hz, 3H), 0.77 (d, J = 5.4 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ
175.5, 173.1, 172.4, 158.4, 138.5, 135.3, 128.5, 128.0, 67.9, 62.5, 59.8,
54.7, 36.1, 31.2, 19.1, 17.9; HRMS (ESI) calcd for C18H25N4O7 [M −
H]− 409.1729, found 409.1726.
CDCl3) δ 8.09 (d, J = 8.2 Hz, 1H), 7.98 (d, J = 8.2 Hz, 1H), 7.51 (t, J
= 7.7 Hz, 1H), 7.38 (t, J = 7.4 Hz, 1H), 7.30−7.13 (m, 5H), 6.16 (d, J
= 7.7 Hz, 1H), 5.69 (dd, J = 8.9, 5.4 Hz, 1H), 5.07 (s, 2H), 3.04 (s,
3H), 2.40−2.26 (m, 1H), 0.91 (d, J = 6.9 Hz, 3H), 0.77 (d, J = 6.6 Hz,
3H); 13C NMR (75 MHz, CDCl3) δ 172.0, 157.6, 155.4, 145.7, 135.2,
130.8, 130.4, 128.4, 128.2, 128.0, 126.2, 120.0, 114.1, 67.7, 58.8, 35.8,
31.4, 19.5, 17.2; HRMS (ESI) calcd for C21H24N6O4Na [M + Na]+
447.1751, found 447.1771.
Cbz-AzaAla-Val-Cys-OH (24c). Compound 24c was prepared
according to the given method for 24a: white microcrystals (0.354
Boc-AzaVal-Leu-Bt (20c). Compound 20c was prepared according
1
g, 83%); mp 113.0−115.0 °C; [α]20 −17.0 (c 1.0, CH3OH); H
to the given method for 20a: white microcrystals (0.779 g, 90%); mp
D
1
92.0−95.0 °C; [α]20 −34.0 (c 1.0, CH3OH); H NMR (300 MHz,
NMR (300 MHz, CDCl3) δ 7.75−7.50 (m, 2H), 7.28−7.22 (m, 5H),
5.10 (s, 2H), 4.73−4.63 (m, 1H), 4.16 (t, J = 8.1 Hz, 1H), 3.00 (s,
3H), 2.96−2.78 (m, 1H), 2.07−1.81 (m, 1H), 1.61 (t, J = 9.0 Hz, 1H),
0.81 (d, J = 6.0 Hz, 3H), 0.74 (d, J = 6.6 Hz, 3H); 13C NMR (75 MHz,
CDCl3) δ 179.5, 173.0, 172.0, 158.2, 135.2, 128.5, 128.4, 128.0, 67.9,
54.2, 53.4, 36.3, 30.6, 26.3, 19.1, 18.2; HRMS (ESI) calcd for
C18H25N4O6S [M − H]− 425.1500, found 425.1521.
D
CDCl3) δ 8.21 (d, J = 8.2 Hz, 1H), 8.09 (d, J = 8.1 Hz, 1H), 7.62 (t, J
= 7.7 Hz, 1H), 7.48 (t, J = 7.7 Hz, 1H), 6.52−6.25 (m, 1H), 5.94−5.78
(m, 2H), 4.70−4.56 (m, 1H), 1.92−1.65 (m, 3H), 1.49 (s, 9H), 1.18−
1.0 (m, 9H), 0.94 (d, J = 4.3 Hz, 2H); 13C NMR (75 MHz, CDCl3) δ
173.6, 157.0, 146.1, 131.4, 130.7, 126.5, 120.4, 114.6, 82.2, 53.1, 48.4,
41.9, 28.3, 25.5, 23.5, 21.7, 19.8, 19.5; HRMS (ESI) calcd for
C21H32N6O4Na [M + Na]+ 455.2377, found 455.2399.
Cbz-AzaAla-Val-(D,L)Cys-OH (24c + 24c′). Compound 24c + 24c′
was prepared according to the given method for 24a: white
microcrystals (0.363 g, 85%); mp 90.0−92.0 °C; 1H NMR (300
MHz, CDCl3) δ 7.71 (d, J = 7.1 Hz, 1H), 7.39 −7.27 (m, 5H), 5.18 (s,
2H), 4.89−4.56 (m, 1H), 4.46−4.13 (m, 1H), 3.09 (s, 3H), 3.04−2.79
(m, 2H), 2.10−1.94 (m, 1H), 1.63 (t, J = 9.0 Hz, 0.5H), 1.49 (t, J = 8.9
Hz, 0.5H), 0.90 (d, J = 6.6 Hz, 3H), 0.83 (d, J = 4.9 Hz, 3H); 13C
NMR was similar to the compound 24c. Anal. Calcd for
C18H26N4O6S: C, 50.69; H, 6.15; N, 13.14. Found: C, 50.48; H,
6.21; N, 13.08.
Boc-AzaVal-Leu-Trp-OH (24d). Compound 24d was prepared
according to the given method for 24a: white microcrystals (0.461
g, 89%); mp 110.0−113.0 °C; [α]20D −4.0 (c 1.0, CH3OH); 1H NMR
(300 MHz, DMSO-d6) δ 10.66 (s, 1H), 8.58 (br s, 1H), 7.94 (d, J =
6.2 Hz, 1H), 7.52 (d, J = 7.7 Hz, 1H), 7.32 (d, J = 7.8 Hz, 1H), 7.12 (s,
1H), 7.05 (t, J = 7.5 Hz, 1H), 6.97 (t, J = 7.4 Hz, 1H), 5.89 (br s, 1H),
4.53−4.50 (m, 1H), 4.37 (qt, J = 6.6 Hz, 1H), 4.28−4.13 (m, 1H),
3.18 (dd, J = 14.7, 5.9 Hz, 1H), 3.05 (dd, J = 14.7, 7.4 Hz, 1H), 1.66−
1.55 (m, 1H), 1.50−1.38 (m, 10H), 1.00 (d, J = 6.6 Hz, 6H), 0.85 (d, J
= 6.3 Hz, 6H); 13C NMR (75 MHz, DMSO-d6) δ 173.1, 172.6, 156.2,
155.3, 136.0, 127.2, 123.6, 120.9, 118.3, 118.2, 111.4, 109.7, 79.6, 52.9,
51.7, 47.1, 42.0, 27.9, 27.1, 23.7, 23.4, 22.0, 19.8, 19.3. Anal. Calcd for
C26H39N5O6: C, 60.33; H, 7.59, N, 13.53. Found: C, 60.48; H, 8.07; N,
13.17.
Boc-AzaVal-Met-Bt (20d). Compound 20d was prepared according
to the given method for 20a: white microcrystals (0.766 g, 85%); mp
1
80.0−82.0 °C; [α]20 −22.0 (c 1.0, CH3OH); H NMR (300 MHz,
D
CDCl3) δ 8.23 (d, J = 7.7 Hz, 1H), 8.11 (d, J = 8.3 Hz, 1H), 7.64 (t, J
= 8.1 Hz, 1H), 7.50 (t, J = 8.1 Hz, 1H), 6.54−6.24 (m, 1H), 4.67−4.59
(m, 1H), 2.68 (q, J = 7.2, 6.1 Hz, 2H), 2.21−2.10 (m, 3H), 2.07−2.05
(m, 3H), 1.50 (s, 9H), 1.13−1.06 (m, 6H); 13C NMR (75 MHz,
CDCl3) δ 172.0, 165.2, 156.7, 145.9, 131.1, 130.6, 126.4, 120.2, 114.3,
83.1, 53.4, 48.2, 32.1, 30.1, 28.1, 19.6, 19.2, 15.4. Anal. Calcd for
C20H30N6O4S: C, 53.32; H, 6.71, N, 18.65. Found: C, 53.04; H, 6.78;
N, 18.33.
Fmoc-AzaVal-Leu-Bt (20e). Compound 20e was prepared
according to the given method for 20a: white microcrystals (1.021
g, 92%); mp 100.0−102.0 °C; [α]20 −19.0 (c 1.0, CH3OH); 1H
D
NMR (300 MHz, DMSO-d6) δ 9.26 (d, J = 4.3 Hz, 1H), 8.28 (d, J =
8.1 Hz, 1H), 8.22 (d, J = 8.1 Hz, 1H), 7.90 (d, J = 7.4 Hz, 2H), 7.85−
7.70 (m, 3H), 7.63 (t, J = 7.7 Hz, 2H), 7.44−7.39 (m, 2H), 7.33 (t, J =
7.5 Hz, 2H), 7.23−6.90 (m, 1H), 5.64−5.56 (m, 1H), 4.56−4.47 (m,
1H), 4.44−3.98 (m, 3H), 1.99−1.41 (m, 3H), 1.07−0.89 (m, 9H),
0.86−0.76 (m, 3H); 13C NMR (75 MHz, DMSO-d6) δ 173.3, 157.2,
156.4, 145.3, 143.7, 143.5, 140.7, 131.0, 130.6, 127.7, 127.1, 126.6,
125.4, 125.2, 120.1, 120.1, 113.9, 65.9, 52.5, 48.0, 46.7, 24.6, 22.9, 21.0,
19.8, 19.2; HRMS (ESI) calcd for C31H34N6O4Na [M + Na]+
577.2534, found 577.2526.
Boc-AzaVal-Met-Ser-OH (24e). Compound 24e was prepared
according to the given method for 24a: white microcrystals (0.362
General Procedure for the Coupling of N-(N-Pg-
azadipeptidoyl)benzotriazoles 20a−e To Prepare 24a−g,
25a,b, and 26a,b. Different amino acids 21a−e or dipeptides
22a,b, 23a,b (1.2 mmol, 1.2 equiv), and TEA (1.2 mmol, 1.2 equiv)
were dissolved in a minimum amount of cold water (5.0 mL).
Acetonitrile (10 mL) was added, and the solutions were cooled to 10
°C. A solution of N-Pg-azadipeptidoyl-Bt (1.0 mmol, 1.0 equiv) in
acetonitrile (5.0 mL) was added and the mixture stirred for 2 h at 20
°C. The reaction mixture was monitored by TLC [EtOAc−hexanes
(1:2)]. After completion of reaction, the solvent was evaporated. The
residue was dissolved in DCM (30 mL) and washed with 2 N HCl
1
g, 83%); mp 48.0−50.0 °C; [α]20 −21.0 (c 1.0, CH3OH); H NMR
D
(300 MHz, Acetone−d6) δ 8.17−8.10 (m, 1H), 7.83−7.73 (m, 1H),
7.56 (t, J = 8.2 Hz, 1H), 7.39−7.31 (m, 1H), 6.41−6.22 (m, 1H), 4.42
(dt, J = 14.8, 7.0 Hz, 3H), 3.84 (d, J = 11.7 Hz, 1H), 3.74 (d, J = 11.1
Hz, 1H), 2.43 (q, J = 7.4 Hz, 2H), 2.16−1.68 (m, 5H), 1.34 (s, 9H),
0.97 (d, J = 7.5 Hz, 6H); 13C NMR (75 MHz, CDCl3) δ 172.9, 172.7,
157.9, 157.7, 62.5, 55.1, 53.9, 48.9, 31.2, 30.3, 28.4, 19.7, 15.5; HRMS
(ESI) calcd for C17H31N4O7S [M − H]− 435.1919, found 435.1935.
Boc-AzaVal-Leu-Asp(OBn)-OH (24f). Compound 24f was prepared
according to the given method for 24a: white microcrystals (0.483 g,
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90%); mp 108.0−110.0; [α]20D −12.0 (c 1.0, CH3OH); 1H NMR (300
MHz, DMSO-d6) δ 8.91 (s, 1H), 8.65−8.10 (m, 1H), 7.40−7.30 (m,
5H), 6.13− 5.86 (m, 1H), 5.10 (s, 2H), 4.62−4.56 (m, 1H), 4.40−4.35
(m, 1H), 4.26−4.21 (m, 1H), 2.87 (dd, J = 17.1, 6.9 Hz, 1H), 2.73
(dd, J = 16.5, 7.2 Hz, 1H), 1.89−1.45 (m, 3H), 1.41 (s, 9H), 0.97 (d, J
= 6.6 Hz, 6H), 0.86 (d, J = 7.1 Hz, 6H); 13C NMR (75 MHz, DMSO-
d6) δ 172.7, 172.0, 170.0, 156.4, 155.4, 136.0, 128.4, 128.0, 127.9, 79.6,
65.8, 51.7, 48.6, 42.2, 35.9, 27.9, 23.2, 21.9, 19.7, 19.3; HRMS (ESI)
calcd for C26H39N4O8 [M − H]− 535.2773, found 535.2782.
The residue was dissolved in DCM, washed with 2 N hydrochloric
acid solution (3 × 5 mL) and brine (5 mL) and dried over MgSO4.
The solvent was evaporated to yield the hybrid peptides: colorless oil
1
(0.410 g, 87%); [α]20 −13.0 (c 1.0, CH3OH); H NMR (300 MHz,
D
CDCl3) δ 7.50 (br s, 1H), 7.23−7.05 (m, 10H), 6.05−5.80 (m, 1H),
5.10 (t, J = 5.4 Hz, 1H), 5.04 (s, 2H), 4.34−4.25 (m, 1H), 3.03−2.79
(m, 5H), 2.01 (s, 1H), 0.71 (d, J = 6.9 Hz, 3H), 0.64 (d, J = 5.7 Hz,
3H); 13C NMR (75 MHz, CDCl3) δ 176.5, 175.5, 171.8, 157.9, 136.3,
135.3, 129.3, 129.1, 128.3, 128.1, 126.9, 126.6, 73.2, 70.9, 67.9, 39.9,
36.8, 30.9, 18.6, 17.0; HRMS (ESI) calcd for C24H28N3O7 [M − H]−
470.1933, found 470.1942.
Fmoc-AzaVal-Leu-Pro-OH (24g). Compound 24g was prepared
according to the given method for 24a: white microcrystals (0.474 g,
1
86%); mp 210.0−212.0 °C; [α]20 −37.0 (c 1.0, CH3OH); H NMR
H-AzaAla-Phe-OH (29a). Cbz-AzaAla-Phe-OH 19a (0.75 g, 2.0
mmol) was dissolved in THF (20 mL), treated with a suspension of 10
mol % of Pd on carbon (20 wt %) in THF, placed under hydrogen gas
at pressure, and stirred at room temperature overnight. The reaction
mixture was filtered through Celite. The filtrate was evaporated on a
rotary evaporator to give H-AzaAla-Phe-OH (29a): white micro-
crystals (0.427 g, 90%); mp 63.0−65.0 °C (lit.29 mp not reported);
D
(300 MHz,DMSO-d6) δ 0.75 (d, 6 Hz, 3H), 1.30−1.48 (m, 2H),
(1.50−2.0, m, 4H), 2.0−2.2 (m, 1H), 3.7 (s, 1H), 4.1−4.4 (m, 4H),
4.44 (s, 2H), 6.10−6.40 (rotamars, 1H), 7.33 (t, 10 Hz, 7.42 (t, 7.5
Hz), 7.75 (d, 7.2 Hz, 2 H), 7.89 (d, 7.2 Hz, 2H), 9.30 (d, 13.2 Hz,
1H); 13C NMR (75 MHz, DMSO-d6) δ 172.6, 171.2, 156.8, 156.5,
143.7, 143.6, 143.5, 140.8, 129.0, 127.8, 127.4, 127.2, 125.4, 125.3,
121.5, 120.2, 120.1, 66.2, 52.3, 48.1, 47.7, 46.8, 40.8, 24.1, 23.3, 21.8,
19.8, 19.4, 18.4. Anal. Calcd for C30H38N4O6: C, 65.44; H, 6.96, N,
10.17. Found: C, 65.65; H, 7.36; N, 10.01.
1
1
[α]20 −3.0 (c 1.0, CH3OH); H NMR (300 MHz, CDCl3) δ H
D
NMR (300 MHz, DMSO-d6) δ 7.32−7.08 (m, 5H), 7.07−6.87 (m,
1H), 4.44−4.07 (m, 1H), 3.03 − 2.73 (m, 5H); 13C NMR (75 MHz,
DMSO-d6) δ 174.2, 159.0, 138.2, 129.8, 128.9, 127.1, 55.0, 38.0, 37.8;
HRMS (ESI) calcd for C11H15N3O3Na [M + Na]+ 260.1006, found.
260.1010.
Cbz-AzaAla-Phe-Gly-Gly-OH (25a). Compound 25a was prepared
according to the given method for 24a: white microcrystals (0.374 g,
1
77%); mp 72.0−74.0 °C; [α]20 = −22.0 (c 1.0, CH3OH); H NMR
D
(300 MHz, DMSO-d6) δ 9.45 (br s, 1H), 8.30−8.10 (m, 2H), 7.44−
7.30 (m, 5H), 7.25−7.10 (m, 5H), 6.70 (br s, 1H), 5.12 (s, 2H), 4.45−
4.35 (m, 1H), 3.89−3.59 (m, 4H), 3.00 (d, J = 6.3 Hz, 1H), 2.94 (d, J
= 6.3 Hz, 1H), 2.91 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ 171.9,
171.1, 169.1, 157.3, 155.6, 137.9, 136.2, 129.3, 128.5, 128.1, 128.0,
127.9, 126.2, 66.5, 55.2, 41.9, 40.7, 37.6, 35.5; HRMS (ESI) calcd for
C46H53N10O14 [2M − H]− 969.3748, found 969.3740.
H-AzaPhe-Leu-OH·HCl (29b). Compound 19e (1.0 mmol) was
dissolved in a solution of dry 4 N HCl/dioxane (10 mL) solution and
stirred for 2 h at room temperature. After completion of the reaction
(followed by TLC) ether (5 mL) was added and the white solid was
filtered off to give H-AzaPhe-Leu-OH·HCl (29b): white microcrystals
(0.294 g, 93%); mp 170.0−172.0 °C; [α]20 −31.0 (c 1.0, CH3OH);
D
1H NMR (300 MHz, DMSO-d6) δ 7.75 (d, J = 7.9 Hz, 1H), 7.47−7.16
(m, 5H), 4.92 (s, 2H), 4.17−4.07 (m, 1H), 1.82−1.40 (m, 3H), 0.87
(d, J = 6.6 Hz, 3H), 0.82 (d, J = 6.1 Hz, 3H); 13C NMR (75 MHz,
DMSO-d6) δ 174.3, 157.0, 135.2, 128.5, 128.3, 127.9, 66.4, 52.1,
51.9,24.3, 23.1, 21.0; HRMS (ESI) calcd for C14H20N3O3 [M − H]−
278.1583, found 278.1564.
Boc-AzaVal-Leu-Gly-Phe-OH (25b). Compound 25b was prepared
according to the given method for 24a: white microcrystals (0.466 g,
1
87%); mp 82.0−85.0; [α]20 −6.0 (c 1.0, CH3OH); H NMR (300
D
MHz, DMSO-d6) δ 8.85 (d, J = 11.1 Hz, 0.7H), 8.37 (s, 0.3H), 8.24−
7.94 (m, 2H), 7.35−7.15 (m, 5H), 6.30 (d, J = 8.1 Hz, 0.5H), 5.99 (d,
J = 8.1 Hz, 0.5H), 4.45−4.35 (m, 2H), 4.25−4.16 (m, 1H), 3.77 (dd, J
= 16.9, 6.2 Hz, 1H), 3.59 (dd, J = 17.0, 5.2 Hz, 1H), 3.04 (dd, J = 13.7,
4.8 Hz, 1H), 2.87 (dd, J = 13.7, 8.7 Hz, 1H), 1.73−1.43 (m, 3H), 1.42
(s, 9H), 0.97 (d, J = 6.6 Hz, 3H), 0.94 (d, J = 6.3 Hz, 3H), 0.88−0.77
(m, 6H); 13C NMR (75 MHz, DMSO-d6) δ 173.1, 172.7, 168.6, 156.5,
155.3, 137.4, 129.1, 128.2, 126.4, 79.5, 53.5, 52.1, 47.1, 41.6, 36.9, 27.9,
23.9, 23.2, 21.8, 20.0, 19.3. Anal. Calcd for C26H41N5O7: C, 58.30; H,
7.72, N, 13.07. Found: C, 58.00; H, 7.98; N, 12.87.
H-AzaGly-Phe-OH·TsOH (29c). Compound 19f (3.0 mmol, 1.0
equiv) was dissolved in a solution of dry DCM (10 mL) containing p-
TsOH (1.0 equiv) and stirred for 12 h at room temperature. After
completion of the reaction [followed by TLC] ether (5 mL) was
added and the white solid was filtered off to give H-AzaGly-Phe-
OH·TsOH (29c): white microcrystals (1.186 g, 100%); mp 159.0−
160.0 °C; [α]20 −36.0 (c 1.0, CH3OH); 1H NMR (300 MHz,
D
DMSO-d6) δ 9.88 (s, 2H), 8.72 (s, 1H), 7.51 (d, J = 8.0 Hz, 2H), 7.36
(d, J = 8.1 Hz, 1H), 7.32−7.18 (m, 5H), 7.14 (d, J = 8.3 Hz, 2H),
4.42−4.32 (m, 1H), 3.10 (dd, J = 13.9, 4.5 Hz, 1H), 2.90 (dd, J = 13.9,
9.4 Hz, 1H), 2.30 (s, 3H); 13C (75 MHz, DMSO-d6) δ 173.0, 156.7,
145.4, 137.8, 137.4, 129.2, 128.3, 128.1, 126.5, 125.5, 54.3, 36.8, 20.9.
Anal. Calcd for C17H21N3O6S: C, 51.64; H, 5.35, N, 10.63. Found: C,
51.85; H, 5.25; N, 10.52.
Cbz-AzaAla-Phe-AOGly-OH (26a). Compound 26a was prepared
according to the given method for 24a: white microcrystals (0.400 g,
1
90%); mp 57.0−59.0 °C; [α]20 = −21.0 (c 1.0, CH3OH); H NMR
D
(300 MHz, DMSO-d6) δ 11.45 (br s, 1H), 9.40 (s, 1H), 7.92 (s, 1H),
7.49−7.38 (m, 5H), 7.25−7.15 (m, 5H), 6.62 (s, 1H), 5.13 (s, 2H),
4.40−4.20 (m, 3H), 3.42−2.75 (m, 5H); 13C NMR (75 MHz, DMSO-
d6) δ 170.7, 169.4, 157.6, 156.1, 137.9, 136.9, 129.9, 129.1, 128.7,
128.4, 127.0, 72.3, 67.0, 53.5, 38.6, 36.1; HRMS (ESI) calcd for
C42H47N8O14 [2M − H]− 887.3217, found 887.3214.
Cbz-Gly-AzaGly-Phe-OH (31a). Tosylate salt of free aza-dipeptide
29c (1.0 mmol, 1.0 equiv) and TEA (1.0 mmol, 1.0 equiv) were
dissolved in minimum amount of cold water (5.0 mL). Acetonitrile
(10 mL) was added to this solution and cooled to 10 °C. A solution of
Cbz-Gly-Bt 30a (1.0 mmol, 1.0 equiv) in acetonitrile (5.0 mL) was
added and stirred for 2 h at 20 °C. The reaction mixture was
monitored by TLC [EtOAc−hexanes (1:2)]. After completion of
reaction, the solvent was evaporated. The residue was dissolved in
DCM (30 mL) and washed with 2 N HCl solution (4 × 10 mL), water
(10 mL) and brine (10 mL). The solvent was dried over MgSO4 and
evaporated to give Cbz-Gly-AzaGly-Phe-OH (31a): white micro-
Boc-AzaVal-Met-Gly-OPhe-OH (26b). Compound 26b was pre-
pared according to the given method for 24a: white microcrystals
(0.471 g, 85%); mp 44.0−46.0 °C; [α]20D −15.0 (c 1.0, CH3OH); 1H
NMR (300 MHz, CDCl3) δ 7.74−7.34 (m, 1H), 7.35−6.91 (m, 5H),
6.80−6.23 (m, 1H), 5.23 (dd, J = 8.6, 4.1 Hz, 1H), 4.75−4.30 (m,
2H), 4.21−3.63 (m, 2H), 3.04 (dd, J = 18.6, 4.1 Hz, 2H), 2.65−2.34
(m, 2H), 2.20−1.80 (m, 5H), 1.47 (s, 9H), 1.19−1.11 (m, 6H); 13C
NMR (75 MHz, acetone-d6) δ 174.8, 171.5, 170.2, 169.4, 157.4, 136.7,
129.8, 128.8, 127.2, 73.6, 73.2, 53.5, 48.3, 42.9, 41.0, 37.3, 32.9, 28.0,
19.7, 19.4, 14.8; HRMS (ESI) calcd for C25H37N4O8S [M − H]−
553.2338, found 553.2350.
crystals (0.340 g, 82%); mp 205.0−207.0 °C (lit.30 mp not reported);
1
[α]20 −12.0 (c 1.0, CH3OH); H NMR (300 MHz, DMSO-d6) δ
D
12.70 (s, 1H), 9.59 (d, J = 2.1 Hz, 1H), 7.96 (s, 1H), 7.46 (t, J = 6.1
Hz, 1H), 7.32 - 7.11 (m, 10H), 6.38 (d, J = 8.0 Hz, 1H), 4.98 (s, 2H),
4.56 - 4.16 (m, 1H), 3.60 (d, J = 6.0 Hz, 2H), 3.29 (s, 3H), 2.97 (dd, J
= 13.7, 5.2 Hz, 1H), 2.86 (dd, J = 13.7, 7.4 Hz, 1H). 13C NMR (75
MHz, DMSO-d6) δ 173.3, 168.9, 157.2, 156.5, 137.3, 137.0, 129.3,
128.4, 128.2, 127.8, 127.7, 126.5, 65.5, 53.9, 42.1, 37.3. Anal. Calcd for
Cbz-AzaAla-Val-OPhe-OH (28). 4-(Dimethylamino)pyridine (1.2
mmol) was added to a stirred solution of Cbz-AzaAla-Val-Bt 20b (1.0
mmol) and α-hydroxycarboxylic acid 27 (1.0 mmol) in dry THF (5.0
mL) at 4 °C. The reaction mixture was stirred for 4 h at room
temperature until the reaction was complete by TLC [EtOAc−hexanes
(1:2)]. After completion of the reaction, the solvent was evaporated.
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C20H22N4O6: C, 57.97; H, 5.35, N, 13.52. Found: C, 57.73; H, 5.17; N,
13.49.
Cbz-Met-Ala-AzaGly-Phe-OH (31b). Compound 31b was prepared
according to the method for preparation of Cbz-GlyAza-Gly-Phe-OH
brine solution (2 × 20 mL), dried over MgSO4, and evaporated under
vacuum to give aza-tripeptide 35a,b.
Boc-Gly-AzaPhe-Leu-OMe (35a). Compound 35a was prepared
according to the general method given for the preparation of 35a,b:
white microcrystals (0.405 g, 90%); mp 167.0−170.0 °C (lit.32 mp not
(31a): white microcrystals (0.476 g, 85%); mp 185.0−187.0 °C;
reported); [α]20 −9.0 (c 1.0, CH3OH); 1H NMR (300 MHz,
1
[α]20 = −40.0 (c 1.0, CH3OH); H NMR (300 MHz, DMSO-d6) δ
D
D
acetone-d6) δ 9.41 (s, 1H), 7.50−7.17 (m, 5H), 6.47 (d, J = 7.2 Hz,
2H), 4.54−4.29 (m, 1H), 3.69 (d, J = 5.5 Hz, 2H), 3.64 (s, 3H), 3.30
(s, 2H), 1.88−1.49 (m, 3H), 1.40 (s, 9H), 0.92 (d, J = 6.6 Hz, 3H),
0.90 (d, J = 6.6 Hz, 3H); 174.8, 168.9, 157.4, 156.8, 136.7, 128.9,
128.8, 127.9, 81.1, 52.5, 52.4, 51.3, 44.0, 41.2, 28.5, 24.9, 23.1, 21.9;
HRMS (ESI) calcd for C22H34N4O6Na [M + Na]+ 473.2371, found
473.2395.
9.70 (d, J = 11.0 Hz, 1H), 8.20−7.96 (m, 2H), 7.59−7.49 (m, 1H),
7.40 − 7.16 (m, 10H), 6.38 (dd, J = 12.5, 7.9 Hz, 1H), 5.03 (s, 2H),
4.42 − 4.20 (m, 2H), 4.18 − 4.05 (m, 1H), 3.10−2.90 (m, 2H), 2.56 −
2.47 (m, 2H), 2.02 (s, 3H), 1.96 − 1.73 (m, 2H), 1.23 (d, J = 6.5 Hz,
3H); HRMS (ESI) calcd for C26H32N5O7S [M − H]− 558.2028, found
558.2029.
General Method for the Preparation of 33a,b. Hydrazine hydrate
(1.0 mmol, 1.0 equiv) and DIPEA (1.0 mmol, 1.0 equiv) were
dissolved in ether (5.0 mL). A solution of Cbz-Phe-Bt 30c or Fmoc-
Cys(S-trt)-Bt 30d (1.0 mmol, 1.0 equiv) in ether (5.0 mL) was added
and the mixture stirred for 10 min at 20 °C. The white precipitate
32a,b was formed and collected by filtration. Compounds 32a,b were
used for the next step without further purification.31 The residue was
dissolved in dry DCM (20 mL) and reacted with 16b,d (1.0 equiv) in
the presence of DIPEA (1.0 equiv) at 20 °C overnight. The reaction
mixture was poured into a separatory funnel, washed with water (2 ×
20 mL), NaHCO3 (3 × 20 mL), and brine solution (2 × 20 mL), dried
over MgSO4, and evaporated under vacuum to give both −NH2 and
−CO2H side protected aza-tripeptides 33a,b.
Cbz-Gly-AzaPhe-Val-OtBu (35b). Compound 35b was prepared
according to the method for preparation of 35a: white microcrystals
(0.410 g, 80%); mp 48.0−49.0 °C; [α]20D −12.0 (c 1.0, CH3OH); 1H
NMR (300 MHz, CDCl3) δ 8.24 (s, 0.5H), 7.37−7.14 (m, 11H), 5.86
(d, J = 8.6 Hz, 1H), 5.61 (s, 0.5H), 5.15−4.94 (m, 2H), 4.96−4.77 (m,
0.5H), 4.49 (d, J = 14.6 Hz, 0.5H), 4.30 (d, J = 8.7 Hz, 1H), 4.28 (d, J
= 8.7 Hz, 1H), 3.69 (d, J = 5.5 Hz, 2H), 2.19−1.98 (m, 1H), 1.40 (s,
9H), 0.90 (d, J = 6.9 Hz, 3H), 0.83 (d, J = 6.9 Hz, 3H); 13C NMR (75
MHz, CDCl3) δ 172.4, 169.0, 157.3, 136.7, 136.1, 129.9, 128.9, 128.8,
128.7, 128.5, 128.3, 127.9, 82.1, 67.6, 59.2, 51.5, 43.9, 31.7, 28.3, 19.2,
18.0; HRMS (ESI) calcd for C27H36N4O6Na [M + Na]+ 535.2527,
found 535.2560.
H-Gly-AzaPhe-Leu-OMe·TsOH (36). Compound 36 was prepared
Cbz-Phe-AzaGly-Leu-OMe (33a). Compound 33a was prepared
according to the general method given for the preparation of 33a,b:
white microcrystals (0.417 g, 86%); mp 179.0−181.0 °C; [α]20D −20.0
(c 1.0, CH3OH); 1H NMR (300 MHz, CDCl3) δ 9.35 (s, 0.5H), 7.97
(s, 0.5H), 7.34−7.02 (m, 11H), 6.40 (d, J = 7.9 Hz, 0.5H), 6.05 (d, J =
7.0 Hz, 0.5H), 4.99 (d, J = 12.6 Hz, 1H), 4.85 (d, J = 12.3 Hz, 1H),
4.63−4.27 (m, 2H), 3.59 (s, 3H), 3.13 (dd, J = 13.9, 5.4 Hz, 1H), 2.92
(dd, J = 14.0, 9.0 Hz, 1H), 1.77−1.42 (m, 3H), 0.85 (d, J = 6.2 Hz,
6H); 13C NMR (75 MHz, CDCl3) δ 174.9, 170.9, 157.8, 156.6, 136.5,
136.2, 129.5, 128.7, 128.6, 128.2, 128.0, 127.0, 67.2, 55.1, 52.5, 51.8,
41.4, 38.1, 24.9, 23.0, 22.0; HRMS (ESI) calcd for C25H32N4O6Na [M
+ Na]+ 507.2214, found 507.2232.
according to the method for preparation of 29c: white microcrystals
(0.372 g, 95%); mp 155.0−157.0 °C; [α]20 −26.0 (c 1.0, CH3OH);
D
1H NMR (300 MHz, DMSO-d6) δ 8.01 (s, 1H), 7.43 (d, J = 8.1 Hz,
2H), 7.34−7.14 (m, 5H), 7.07 (d, J = 7.8 Hz, 2H), 4.25−4.04 (m,
1H), 3.74−3.52 (m, 7H), 2.24 (s, 3H), 1.70−1.37 (m, 3H), 0.82 (d, J
= 6.3 Hz, 3H), 0.79 (d, J = 6.3 Hz, 3H); 13C NMR (75 MHz, DMSO-
d6) δ 174.2, 166.7, 157.4, 146.2, 138.3, 138.2, 128.8, 128.7, 128.6,
126.1, 119.9, 79.0, 52.4, 51.7, 43.0, 28.8, 24.7, 23.4, 22.0, 21.5.
1
Compound 36 was partially characterized by H and 13C NMR and
used for the next step without further purification
Cbz-Tyr-(OBn)-Bt (38). Benzotriazole (6.0 mmol, 3.0 equiv) was
dissolved in dry DCM (50 mL). SOCl2 (2.2 mmol) was added by
syringe, and the mixture was stirred for 15 min at rt under argon. The
solution temp was lowered to −30 to −40 °C (dry ice + acetone), and
2.0 equiv of TEA was added. After 5 min of stirring, Cbz-Tyr-(OBn)-
OH 37 (2.0 mmol, 1.0 equiv) was added and the mixture stirred for 2
h keeping the temperature at −30 to −40 °C. After completion of the
reaction, ice-cold water (20 mL) was added and the organic layer was
washed with water (20 mL × 2), NaHCO3 (20 mL × 4), and then
brine (20 mL × 2). The organic layer was dried over MgSO4 and
Fmoc-Cys(S-trt)-AzaGly-Val-OtBu (33b). Compound 33b was
prepared according to the method for preparation of 33a: white
microcrystals (0.599 g, 75%); mp 79.0−81.0 °C; [α]20D −23.0 (c 1.0,
1
CH3OH); H NMR (300 MHz, CDCl3) δ 7.71 (dd, J = 7.4, 3.9 Hz,
2H), 7.59−7.46 (m, 2H), 7.45−7.30 (m, 8H), 7.30−7.08 (m, 12H),
6.74 (s, 1H), 6.23−6.02 (m, 1H), 4.45−4.24 (m, 3H), 4.14−4.10 (m,
2H), 2.80−2.55 (m, 2H), 1.50−1.38 (m, 9H), 1.29−1.23 (m, 1H),
0.93 (d, J = 6.6 Hz, 3H), 0.87 (d, J = 6.9 Hz, 3H). 13C NMR (75 MHz,
CDCl3) δ 171.1, 170.1, 166.4, 158.2, 144.2, 143.6, 141.2, 129.5, 128.1,
127.7, 126.9, 125.0, 119.9, 82.0, 67.1, 65.8, 58.1, 47.0, 31.6, 31.3, 28.0,
19.0, 17.5; HRMS (ESI) calcd for C47H50N4O6SNa [M + Na]+
821.3343, found 821.3358.
evaporated to yield a white solid: white microcrystals (0.841 g, 83%);
1
mp 87.0−89.0 °C; [α]20 −20.0 (c 1.0, CH3OH); H NMR (300
D
MHz, CDCl3) δ 8.20 (d, J = 8.2 Hz, 1H), 8.11 (d, J = 8.6 Hz, 1H),
7.63 (t, J = 7.6 Hz, 1H), 7.50 (t, J = 7.5 Hz, 1H), 7.41 − 7.23 (m,
10H), 7.03 (d, J = 8.2 Hz, 3H), 6.82 (dd, J = 8.8, 3.1 Hz, 2H), 6.15 −
5.88 (m, 1H), 5.68 (d, J = 8.1 Hz, 1H), 5.07 (d, J = 3.5 Hz, 2H), 4.96
(d, J = 2.8 Hz, 2H), 3.40 (dd, J = 13.9, 5.2 Hz, 1H), 3.14 (dd, J = 14.1,
7.6 Hz, 1H); 13C NMR (75 MHz, CDCl3) δ 170.8, 158.0, 155.7, 145.9,
136.7, 135.9, 130.9, 130.6, 130.2, 128.4, 128.0, 127.8, 127.3, 127.1,
126.4, 120.2, 114.9, 114.2, 69.8, 67.1, 55.7, 37.8. Compound 38 was
partially characterized by 1H and 13C NMR and taken to the next step
without further purification.
General Method for the Preparation of 35a,b. To a solution of
benzaldehyde (1.0 equiv) in diethyl ether (10 mL) were added 32c,d
(prepared by using literature method)31 (1.0 equiv) and two drops of
glacial acetic acid. The reaction mixture was heated under reflux for 2−
3 h and then cooled to room temperature. The white solid precipitate
was collected by filtration, washed with cold diethyl ether and dried
under vacuum to yield the desired hydrazone. The hydrazone
intermediate (1.0 equiv) was treated with sodium cyanoborohydride
(NaBH3CN) (1.1 equiv) in absolute methanol (20 mL). HCl (1 N) in
MeOH was added dropwise over 1 h to maintain the reaction pH in
between 3.5−5.0. After being stirred at room temperature overnight,
the solvent was removed and the residue was partitioned between
ether (30 mL) and brine. The organic phase was washed with
saturated NaHCO3 (2 × 20 mL) and brine (2 × 10 mL), dried over
MgSO4, and evaporated to give the 34a,b (yield 70−76%).
Compounds 34a,b were taken to the next step without further
purification. Each residue was dissolved in dry DCM (20 mL) and
reacted with 16b,d (1.0 equiv) in the presence of TEA (1.0 equiv.) at
20 °C overnight. The reaction mixture was poured into a separatory
funnel, washed with water (2 × 20 mL), NaHCO3 (3 × 20 mL), and
Cbz-Tyr-(OBn)-Gly-Bt (39). Gly (1.2 mmol, 1.2 equiv) and TEA
(1.2 mmol, 1.2 equiv) were dissolved in a minimum amount of cold
water (5.0 mL). Acetonitrile (10 mL) was added, and the solution was
cooled to 10 °C. A solution of Cbz-Tyr-(OBn)-Bt 38 (1.0 mmol, 1.0
equiv) in acetonitrile (5.0 mL) was added and the mixture stirred for 2
h at 20 °C. The reaction mixture was monitored by TLC [EtOAc−
hexanes (1:2)]. After completion of reaction, the solvent was
evaporated. The residue was dissolved in DCM (30 mL) and washed
with 2 N HCl solution (4 × 10 mL), water (10 mL), and brine (10
mL). The solvent was dried over MgSO4 and evaporated to give Cbz-
Tyr-(OBn)-Gly-OH, which was directly taken to the next step without
further purification. Benzotriazole (3.0 mmol, 3.0 equiv) was dissolved
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in dry DCM (50 mL). SOCl2 (1.1 mmol) was added by syringe, and
the mixture was stirred for 15 min at rt. under argon. The solution
temperature was lowered to −30 to −40 °C (dry ice + acetone), and
2.0 equiv of TEA was added. After 5 min of stirring, Cbz-Tyr-(OBn)-
Gly-OH was added, and the mixture was stirred for 2 h keeping the
temperature at −30 to −40 °C. After completion of the reaction, ice-
cold water (20 mL) was added, and the organic layer was washed with
water (20 mL × 2); NaHCO3 (20 mL × 4) and then brine (20 mL ×
2). The organic layer was dried over MgSO4 and evaporated to yield
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Cbz-Tyr-(OBn)-Gly-Bt (39): white microcrystals (0.451 g, 80%); mp
1
132−133 °C; [α]20 −19.0 (c 1.0, CH3OH); H NMR (300 MHz,
D
CDCl3) δ 8.10 (d, J = 9.0 Hz, 1H), 8.06 (d, J = 8.1 Hz, 1H), 7.58 (t, J
= 7.1 Hz, 1H), 7.46 (t, J = 7.1 Hz, 1H), 7.36−7.29 (m, 5H), 7.28−7.22
(m, 6H), 7.19−7.00 (m, 3H), 6.91−6.75 (m, 2H), 5.74 (d, J = 7.1 Hz,
1H), 5.18−4.99 (m, 4H), 4.93 (s, 2H), 4.75−4.44 (m, 1H), 3.24−2.96
(m, 2H); 13C NMR (75 MHz, CDCl3) δ 172.6, 167.7, 158.1, 156.6,
146.1, 139.0, 137.1, 136.1, 131.0, 130.6, 128.7, 128.3, 128.1, 127.6,
126.7, 126.2, 120.5, 115.2, 114.2, 70.1, 67.5, 56.6, 43.5, 37.9.
Compound 39 was partially characterized by 1H, 13C NMR and
taken to the next step without further purification.
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Leu-enkephalin-aza Analogue (40). The tosylate salt of free aza-
tripeptide 36 (0.5 mmol, 1.0 equiv) and DIPEA (1.0 mmol, 2.0 equiv)
were dissolved in dry THF. A solution of Cbz-Tyr-(OBn)-Gly-Bt 39
(0.5 mmol, 1.0 equiv) in THF (5.0 mL) was added and the mixture
stirred for 12 h at 20 °C. The reaction mixture was monitored by TLC
[EtOAc−hexanes (1:2)]. After completion of the reaction, the solvent
was evaporated. The residue was dissolved in DCM (30 mL) and
washed with 2 N HCl solution (4 × 10 mL), water (10 mL), and brine
(10 mL). The solvent was dried over MgSO4 and evaporated to give
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ARKIVOC 2011, 212−220.
(16) Avan, I.; Tala, S. R.; Steel, P. J.; Katritzky, A. R. J. Org. Chem.
2011, 76, 4884−4893.
Leu-enkephalin-aza analogue (40): white microcrystals (0.278 g,
1
70%); mp 89.0−91.0 °C; [α]20 −13.0 (c 1.0, CH3OH); H NMR
D
(17) (a) Katritzky, A. R; Singh, A.; Haase, D. N.; Yoshioka, M.
ARKIVOC 2009, 47−56. (b) Katritzky, A. R.; Avan, I.; Tala, S. R. J.
Org. Chem. 2009, 74, 8690−8694.
(300 MHz, DMSO-d6) δ 9.84 (s, 1H), 8.17−7.91 (m, 2H), 7.42−7.25
(m, 20H), 6.90 (dd, J = 5.8, 2.7 Hz, 2H), 6.46 (d, J = 7.9 Hz, 1H), 5.06
(s, 2H), 5.03−4.82 (m, 3H), 4.77−4.37 (m, 2H), 4.32−4.16 (m, 2H),
3.93−3.68 (m, 4H), 3.62 (s, 3H), 3.01 (dd, J = 13.9, 4.2 Hz, 2H),
2.94−2.65 (m, 2H), 1.77−1.41 (m, 3H), 0.91−0.84 (m, 6H); 13C
NMR (75 MHz, DMSO-d6) δ 174.2, 172.8, 172.5, 170.3, 168.9, 157.4,
156.5, 138.4, 137.8, 137.5, 131.1, 130.9, 130.8, 129.0, 128.8, 128.7,
128.5, 128.4, 128.2, 128.1, 128.0, 127.9, 127.6, 127.5, 127.3, 114.9,
70.5, 69.7, 67.0, 65.9, 57.0, 52.3, 42.7, 42.0, 37.1, 31.8, 24.6, 23.5, 21.9;
HRMS (ESI) calcd for C43H50N6O9Na [M + Na]+ 817.3531, found
817.3551.
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ASSOCIATED CONTENT
■
(23) Loser, R.; Schilling, K.; Dimmig, E.; Gutschow, M. J. Med. Chem.
̈
̈
S
* Supporting Information
1H and/or 13C spectra of all compounds listed in the
Experimental Section. This material is available free of charge
via the Internet at http://pubs.acs.org.
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10701−10708.
AUTHOR INFORMATION
Corresponding Author
*E-mail: katritzky@chem.ufl.edu.
■
(26) Groutas, W. C.; Brubaker, M. J.; Zandler, M. E.; Mazo-Gray, V.;
Rude, S. A.; Crowley, J. P.; Castrisos, J. C.; Dunshee, D. A.; Giri, P. K.
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Author Contributions
(27) Groutas, W. C.; Abrams, W. R.; Theodorakis, M. C.; Kasper, A.
M.; Rude, S. A.; Badger, R. C.; Ocain, T. D.; Miller, K. E.; Moi, M. K. J.
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#N.E.A-D. and S.B. contributed equally.
Notes
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517−522.
The authors declare no competing financial interest.
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316, 741−749.
ACKNOWLEDGMENTS
■
This paper was funded by King Abdulaziz University under
Grant No. D- 006/431. The authors, therefore, acknowledge
technical and financial support of KAU. We thank Dr. C. D.
Hall and Mr. Z. Wang for helpful discussions.
(30) Morley, J. S. Eur. Pat. Appl. 1983, 22.
(31) Ghazvini Zadeh, E. H.; El-Gendy, B. E.-D. M.; Pop, A. G.;
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