γ-Glutamyl-dipeptides: Easy tools to rapidly probe the stereoelectronic properties of the ionotropic glutamate receptor binding pocket

Article abstract of DOI:10.1016/j.tet.2016.11.025

γ-Glutamyl-dipeptides, built by condensing the distal carboxylate of L-Glu (or D-Glu) onto a series of differently functionalized amino acids, were prepared and used as tools for rapidly probing the stereo-electronic properties of iGluRs, searching for subtype-selective ligands.

Full text of DOI:10.1016/j.tet.2016.11.025

Tetrahedron 72 (2016) 8486e8492
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g
-Glutamyl-dipeptides: Easy tools to rapidly probe the
stereoelectronic properties of the ionotropic glutamate receptor
binding pocket
Lucia Tamborini a, Veronica Nicosia a, Paola Conti a, Federica Dall'Oglio a,
Carlo De Micheli a, Birgitte Nielsen b, Anders A. Jensen b, Darryl S. Pickering b,
,
*
Andrea Pinto a
a Department of Pharmaceutical Sciences (DISFARM), University of Milan, Via Mangiagalli 25, 20133 Milan, Italy
b Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen
OE, Denmark
a r t i c l e i n f o
a b s t r a c t
Article history:
g
-Glutamyl-dipeptides, built by condensing the distal carboxylate of L-Glu (or D-Glu) onto a series of
Received 1 September 2016
Received in revised form
21 October 2016
Accepted 7 November 2016
Available online 9 November 2016
differently functionalized amino acids, were prepared and used as tools for rapidly probing the stereo-
electronic properties of iGluRs, searching for subtype-selective ligands.
© 2016 Elsevier Ltd. All rights reserved.
Keywords:
Glutamic acid
Dipeptides
Ionotropic glutamate receptors
Flow-chemistry
Polymer-supported reagents
1. Introduction
Starting from the structure of the endogenous ligand L-Glu,
selectivity for iGluRs vs mGluRs can be achieved through confor-
L-Glutamic acid (
L-Glu, Fig. 1) is the main excitatory neuro-
mational constraint, locking the ligand into a folded or extended
transmitter in the central nervous system (CNS), where it is
involved in the modulation of many physiological processes such as
learning, memory, and synaptic plasticity.1 Once released into the
conformation, respectively (e.g., compound L-CCG-IV vs L-CCG-I,
Fig. 1).6 In contrast, when it comes to the design of ligands selective
for a specific iGluR or, ideally, for an iGluR subtype, common stra-
tegies often rely on the increase of molecular complexity.7 This
strategy aims at exploiting the subtle differences among iGluRs in
terms of stereo-electronic properties of the ligand binding domain
(LBD) of the different receptor subunits (GluN1, GluN2A-C, GluN3;
GluA1-4 and GluK1-5).
synaptic cleft,
-Glu receptors (iGluRs and mGluRs, respectively). The iGluRs
are divided into three classes based on sequence homology and
ligand selectivity: N-methyl- -aspartic acid (NMDA) receptors,
L-Glu acts through both ionotropic and metabotropic
L
D
(RS)-2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid
(AMPA) receptors, and kainic acid (KA) receptors. These classes of
receptors are further subdivided into subtypes.1e5 The action of
For instance, GluK1 selective agonists, such as the 5-tert-butyl
derivative of AMPA, 2-amino-3-(5-tert-butyl-3-hydroxy-4-isoxazolyl)
propionic acid (ATPA) and 5-iodowillardine (Fig. 1), are sterically
hindered by the side chain of Asp690 when docked into the GluK2
pocket and this is relieved by the presence of the smaller Ser706 at the
equivalent position in GluK1.7 Steric obstruction may also explain the
reduced selectivity of ATPA for AMPARs. Steric occlusion partly ex-
plains also the higher selectivity of the agonist (2S,4R)-4-
methylglutamate (SYM 2081, Fig. 1) for kainate receptors over AMPA
L-Glu is then silenced by a specific reuptake transport system, the
excitatory amino acid transporters (EAATs), which are localized
both at the synaptic nerve terminals and at glial cells.
* Corresponding author.
E-mail address: andrea.pinto@unimi.it (A. Pinto).
http://dx.doi.org/10.1016/j.tet.2016.11.025
0040-4020/© 2016 Elsevier Ltd. All rights reserved.

L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
8487
establishment of additional interaction/steric occlusion with the
binding pocket.
2. Results and discussion
2.1. Chemistry
L-Glu (or D-Glu) was used as the starting material for the syn-
thesis of the suitably protected amino acid 23 (Scheme 1), obtained
in good yield and high purity, following a literature procedure.11 On
the other hand, each amino acid used in the coupling reaction was
protected at the carboxylic function using thionyl chloride in
MeOH, yielding the corresponding methyl ester in quantitative
yield as a hydrochloride salt. The amino acids Lys, Glu and Ser were
further protected at the side chain functional groups.12 The
coupling reaction was performed exploiting the use of a flow
chemistry reactor in combination with an immobilized coupling
reagent (polymer supported (PS)-HOBt) and scavengers.13 PS-HOBt
was packed into an Omnifit® glass column, then, after a washing/
swelling step with DMF, a solution of the carboxylic acid 23, DIPEA
and the phoshponium coupling reagent bromo-tris-pyrrolidino
phosphonium hexafluorophosphate (PyBroP) in DMF was fluxed
Fig. 1. Structure of model compounds.
receptors.8 In addition, homologation of the amino acidic chain is a
strategy often pursued to turn agonists into antagonists, because it
prevents the closure of the clamshell like LBD, thus leaving the
channel pore closed.9,10 Finally, it has to be noticed that, whereas
AMPA and KA receptor ligands, analogously to the endogenous
through it at 100 mL/min (Scheme 1, step A). The column was then
washed with DMF, leaving an activated amino ester covalently
attached to the polymer support. The column was then connected
with two prewashed columns of Amberlyst A-21 and Amberlyst A-
15 linked in series (Scheme 1, step B). A solution of the second
amino acid methyl ester as hydrochloride salt in DMF was fluxed
through the column containing A-21 to liberate the free amine,
which was then fluxed into the column containing the activated
ester, where the coupling took place. The exiting flow stream was
finally directed into the column filled with A-15 to perform an in-
line purification, by scavenging any unreacted amine.
agonist L-Glu, usually are characterized by an S-configuration at the a-
amino acid stereogenic center, NMDA receptors often exhibit a pref-
erence for R-configured amino acids, as in the case of the prototypical
agonist NMDA.
On this ground, we envisaged that
g-glutamyl-dipeptides, built
by condensing the distal carboxylate of
L
-Glu (or -Glu) to a series of
D
differently functionalized amino acids, could represent an easy
strategy to synthesize ligands, featuring as ideal tools for rapidly
probing the stereo-electronic properties of iGluRs, searching for
The resulting solution was then evaporated to obtain the pro-
tected dipeptides 24e45 in high yields (75e90%), in high purities
(>95%) without any aqueous work up or column chromatography
and with a significant reduction of time compared to traditional
subtype-selective antagonists (Fig. 2). Indeed,
g-glutamyl-di-
peptides are acidic amino acids characterized by a 6-atom spacer
connecting the proximal and the distal acidic functions and by
different side chains useful to confer an increasing degree of mo-
lecular complexity to the molecules, making possible the
Fig. 2. Structure of target compounds.
Scheme 1. In flow coupling reaction.

8488
L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
Table 1
batch synthesis (20 min vs 2 h). The final compounds 1e22 were
obtained after deprotection of intermediates 24e45 (Scheme 2).
These steps were easily carried out in batch, through the hydrolysis
of the esters with 1 N NaOH in MeOH and removing the Boc pro-
tecting groups in acidic conditions using a 30% trifluoroacetic acid
solution in CH2Cl2 (Scheme 2). These acidic conditions allowed also
the removal of the trityl protecting group in intermediates 29, 30,
40 and 41. In the case of compounds 21 and 22, which possess the
distal amino function of Lys, the acidic ion exchange resin Amber-
lite IR-120 was used to liberate the distal amine from the tri-
fluoroacetate salt.
Receptor binding affinities at native rat iGluRs.a
Compd
[3H]AMPA Ki (
mM)
[3H]KAIN Ki (
m
M)
[3H]CGP39653 Ki (
mM)
1
2
3
4
5
6
7
8
>100
>100
>100
>100
>100
>100
>100
8.4 [5.08 0.07]
>100
>100
4.1 [5.39 0.05]
10 [5.00 0.03]
>100
15 [4.81 0.02]
>100
4.0 [5.40 0.04]
42 [4.38 0.07]
0.74 [6.15 0.10]
24 [4.62 0.03]
9.8 [5.06 0.14]
>100
5.4 [5.29 0.07]
40 [4.42 0.10]
>100
44 [4.39 0.11]
>100
5.8 [5.27 0.10]
21 [4.73 0.10 ]
0.92 [6.04 0.03]
9.6 [5.04 0.08]
22 [4.73 0.14]
>100
7.2 [5.15 0.05]
21 [4.67 0.03]
>100
9
10
11
12
13
14
15
16
17
18
19
20
21
22
>100
>100
>100
3. Pharmacology
>100
>100
>100
>100
All dipeptides were preliminarily submitted to binding assays at
native iGluRs, using rat brain synaptic membranes from male
SpragueeDawley rats (Table 1).
>100
>100
ꢁ100
>100
>100
>100
>100
>100
6.8 [5.18 0.06]
33 [4.51 0.09]
3.3 [5.48 0.01]
85 [4.07 0.01]
72 [4.14 0.02]
>100
ꢁ100
25 [4.60 0.03]
>100
These binding data show that several compounds characterized
>100
by the presence of the
L-Glu moiety displayed moderate affinity
>100
>100
(IC50 ꢀ 10 M) for native KARs (i.e., compounds 1, 2, 6, 8 and 10).
m
>100
>100
>100
>100
Unfortunately, the compounds lack KAR selectivity because they
also exhibited affinity towards NMDA receptors. Indeed, the
highest-affinity hit 8 and the compounds 1 and 6 displayed high
a
Data are given as mean [mean pKi SEM] of three to four independent ex-
periments each conducted in triplicate.
affinity also for NMDARs (Ki ¼ 0.9, 5.4 and 5.8 M, respectively), The
m
ability of the endogenous dipeptide -Glu-L-Glu and few other
dipeptides or tripeptides (such as glutathione) to bind with
g
-
L
4. Conclusion
micromolar affinity to ionotropic glutamate receptors was previ-
ously observed.14 Interestingly, several derivatives within the
D-Glu
In the present work, we have developed an easy route to rapidly
generate a small library of ligands able to interact, with different
binding affinities and degrees of selectivity, with KA and NMDA
receptors. The length of the amino acidic skeleton obtained by
series (i.e., compounds 12, 13, 17 and 19) behaved as selective
NMDA receptor ligands exhibiting low micromolar binding affin-
ities (Ki ¼ 7.2, 21, 6.8 and 3.3
mM, respectively) at these receptors.
coupling the distal carboxylate of D- or L-Glu, to an a-amino acid
In parallel, since it is known that an increase in molecular
complexity of the aspartate/glutamate skeleton may generate
molecules that act as blockers of the EAATs,15 we have also tested
the new derivatives 1e11 as potential inhibitors of the EAAT sub-
types 1, 2 and 3 (EAAT1-3). In detail, the inhibition of the com-
pounds at human EAAT1-3 stably expressed in HEK293 cells was
seems to be appropriate to fit the iGluR binding site, whereas
additional natural and non-natural amino acids remain to be
investigated, in order to maximize the receptor affinity and possibly
increase the selectivity profile.
determined in a [3H]-
viously described using
D
-Aspartate uptake assay performed as pre-
5. Experimental
L-Glu and
D,L
-TBOA as reference ligands.16
Compound 6 displayed weak inhibitory activity at all the three
transporters at high micromolar concentrations, while the other
5.1. Materials and methods
derivatives proved to be inactive up to concentrations of 300
1000 M.
mM or
All reagents were purchased from Sigma. 1H NMR and 13C NMR
spectra were recorded with a Varian Mercury 300 (300 MHz)
m
spectrometer. Chemical shifts (d) are expressed in ppm downfield
from tetramethylsilane, and coupling constants (J) are expressed in
Hz. Optical rotation determinations were carried out using a Jasco
P-1010 spectropolarimeter, coupled with a Haake N3-B thermostat.
TLC analyses were performed on commercial silica gel 60 F254
aluminum sheets; spots were further evidenced by spraying with a
dilute alkaline potassium permanganate solution or ninhydrin. MS
analyses were performed on a Varian 320-MS triple quadrupole
mass spectrometer with ESI source. Microanalyses (C, H, N) of new
compounds were within
0.4% of theoretical values. The
continuous-flow peptide synthesis was performed using a R2þ/R4
flow reactor, commercially available from Vapourtec equipped with
Omnifit glass column.
5.2. Binding assays
Affinities for native NMDA, AMPA, and KA receptors in rat
cortical synaptosomes were determined using 2 nM [3H]CGP39653,
5 nM [3H]AMPA, and 5 nM [3H]KA, respectively, with minor mod-
ifications as previously described.17
Scheme 2. Reagents and conditions: a) 1 N NaOH, MeOH, b) 30% TFA, CH2Cl2.

L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
8489
5.3. General procedures
5.6. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-1-
oxo-3-phenylpropan-2-ylamino)-5-oxopentanoate (26)
5.3.1. General procedure for peptide coupling
0.5 g of PS-HOBt was packed (loading 1.0 mmoL/g) into a glass
column (Ø: 6.6 mm; h: 100 mm) and DMF was fluxed for 15 min
Yield: 88%; yellow oil; Rf (cyclohexane/EtOAc 6/4): 0.22;
[a
]
20: ꢄ20.3 (c: 1.00 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t,
D
(flow rate: 200
(261 L; 1.5 mmol) were added to a 0.5 M solution of 23 (275 mg;
1.0 mmol in 2.0 mL of DMF) and the resulting solution was fluxed
into the flow reactor at 100 L/min 0.5 g of Amberlyst A-21 was
packed into a glass column (Ø: 6.6 mm; h: 100) and DMF was fluxed
for 10 min (flow rate: 200 L/min). 0.5 g of Amberlyst A-15 were
packed into a third glass column (Ø: 6.6 mm; h: 100 mm) and DMF
was fluxed for 10 min (flow rate: 200 L/min). The column con-
m
L/min). PyBroP (559 mg, 1.2 mmol) and DIPEA
J ¼ 7.0, 3H); 1.45 (s, 9H); 1.80e1.90 (m, 1H); 2.10e2.30 (m, 3H); 3.06
(dd, J ¼ 6.8, 14.1, 1H); 3.17 (dd, J ¼ 5.4, 14.1, 1H); 3.75 (s, 3H); 4.20 (q,
J ¼ 7.0, 2H); 4.35e4.45 (m, 1H); 4.80e4.90 (m, 1H); 5.30 (bd, J ¼ 6.8,
m
m
1H); 6.75 (bd, J ¼ 5.9,1H); 7.10e7.20 (m, 2H); 7.20e7.30 (m, 3H); 13
C
NMR (75 MHz, CDCl3): 14.4, 28.5, 29.7, 32.6, 38.0, 52.6, 53.1, 53.8,
61.8, 80.4, 127.3, 128.8, 129.5, 136.3, 156.2, 172.1, 172.4, 172.5. Anal.
calcd for C22H32N2O7: C, 60.54; H, 7.39; N, 6.42; found: C, 60.30; H,
7.28; N, 6.55.
m
m
taining A-21 was connected in series with the column packed with
PS-HOBt and with the column containing A-15. A 100 psi back-
pressure regulator was connected. A 0.2 M solution of the amino
acid methyl ester hydrochloride (0.5 mmol in 2.5 mL of DMF) was
injected into the reactor. The flow rate was set in order to obtain a
residence time of 20 min.
5.7. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-4-
methyl-1-oxopentan-2-ylamino)-5-oxopentanoate (27)
Yield: 82%; yellow oil; Rf (cyclohexane/EtOAc 6/4): 0.30;
[a
]
D
20: þ5.5 (c: 1.10 in CHCl3); 1H NMR (300 MHz, CDCl3): 0.94 (d,
J ¼ 6.3, 6H); 1.25 (t, J ¼ 7.1, 3H); 1.45 (s, 9H); 1.50e1.70 (m, 3H);
1.80e1.90 (m, 1H); 2.10e2.25 (m, 1H); 2.28e2.36 (m, 2H); 3.75 (s,
3H); 4.20 (q, J ¼ 7.1, 2H); 4.20e4.30 (m, 1H); 4.55e4.65 (m, 1H);
5.25 (bd, J ¼ 7.7, 1H); 6.35 (bd, J ¼ 6.6, 1H); 13C NMR (75 MHz,
CDCl3): 14.4, 22.1, 23.1, 25.1, 28.5, 29.3, 32.6, 41.7, 51.0, 52.5, 53.2,
61.8, 80.3, 156.1, 171.9, 172.5, 173.8. Anal. calcd for C19H34N2O7: C,
56.70; H, 8.51; N, 6.96; found: C, 57.01; H, 8.65; N, 7.17.
5.3.2. General procedure for dipeptide deprotection
1) The protected dipeptide (0.5 mmol) was dissolved in MeOH
(1.5 mL) and 1 N NaOH (1.5 mL, 1.5 mmol) was added. The mixture
was stirred at rt for 3 h and the disappearance of the starting ma-
terial was monitored by TLC (CH2Cl2/MeOH 9/1 þ 1% AcOH). After
evaporation of MeOH, the residue was diluted with distilled H2O
(5 mL) and washed with CH2Cl2 (2 ꢂ 5 mL). After that, the aqueous
phase was made acidic (pH ¼ 2) with 2 N aqueous HCl and then
extracted with EtOAc (3 ꢂ 5 mL). The organic phase was dried over
anhydrous Na2SO4 and, after evaporation of the solvent, the inter-
mediate was obtained as a white solid.
2) The intermediate obtained from the previous step (0.5 mmol)
was treated with a 30% solution of TFA in CH2Cl2 (1.25 mL) at 0 ꢃC.
The solution was stirred at rt for 2 h and the reaction was followed
by TLC (CH2Cl2/MeOH 9/1 þ 1% AcOH). The volatiles were removed
under reduced pressure. The obtained solid was dissolved in water
(1 mL) and lyophilized to give the final dipeptide.
5.8. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-4-
methyl-1-oxopentan-2-ylamino)-5-oxopentanoate (28)
Yield: 75%; yellow oil; Rf (cyclohexane/EtOAc 6/4): 0.20;
[a
]
D
20: þ12.7 (c: 1.55 in CHCl3); 1H NMR (300 MHz, CDCl3): 0.94 (d,
J ¼ 6.0, 6H); 1.25 (t, J ¼ 7.0, 3H); 1.45 (s, 9H); 1.50e1.70 (m, 3H);
1.80e1.90 (m, 1H); 2.10e2.25 (m, 1H); 2.28e2.36 (m, 2H); 3.75 (s,
3H); 4.20 (q, J ¼ 7.0, 2H); 4.35e4.45 (m, 1H); 4.50e4.60 (m, 1H);
5.32 (bd, J ¼ 7.1, 1H); 6.90 (bd, J ¼ 6.8, 1H); 13C NMR (75 MHz,
CDCl3): 14.4, 22.1, 23.1, 25.1, 28.5, 29.9, 32.7, 41.4, 51.3, 52.5, 53.1,
61.8, 80.5, 156.3, 172.3, 172.6, 173.8. Anal. calcd for C19H34N2O7: C,
56.70; H, 8.51; N, 6.96; found: C, 56.96; H, 8.67; N, 7.12.
5.4. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-(2-methoxy-2-
oxoethylamino)-5-oxopentanoate (24)
5.9. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-1-
oxo-3-(trityloxy)propan-2-ylamino)-5-oxopentanoate (29)
Yield: 80%; purple oil; Rf (cyclohexane/EtOAc 1/1): 0.41;
[
a
]
20: þ11.6 (c: 1.00 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t,
D
Yield: 75%; Solid state: pale yellow oil; Rf (cyclohexane/EtOAc 7/
20: ꢄ21.0 (c: 1.00 in CHCl3); 1H NMR (300 MHz, CDCl3):
J ¼ 7.0, 3H); 1.45 (s, 9H); 1.80e2.00 (m, 1H); 2.10e2.25 (m, 1H);
2.30e2.38 (m, 2H); 3.75 (s, 3H); 4.05 (dd, J ¼ 5.5, 9.4, 2H); 4.20 (q,
J ¼ 7.0, 2H); 4.25e4.40 (m, 1H); 5.35 (bd, J ¼ 7.1, 1H); 6.70 (bd,
J ¼ 5.5, 1H); 13C NMR (75 MHz, CDCl3): 14.4, 28.5, 29.4, 32.5, 41.5,
52.6, 53.1, 61.8, 80.3, 156.2, 170.7, 172.5, 172.6. Anal. calcd for
3): 0.21; [
a
]
D
1.25 (t, J ¼ 7.0, 3H); 1.45 (s, 9H); 1.85e2.00 (m, 1H); 2.10e2.20 (m,
1H); 2.30 (m, 2H); 3.35 (dd, J ¼ 3.3, 9.4, 1H); 3.60 (dd, J ¼ 3.3, 9.4,
1H); 3.75 (s, 3H); 4.15 (q, J ¼ 7.0, 2H); 4.20e4.30 (m, 1H); 4.70 (ddd,
J ¼ 3.3, 3.3, 8.0, 1H); 5.24 (bd, J ¼ 8.0, 1H); 6.55 (bd, J ¼ 8.3, 1H);
7.20e7.40 (m, 15H); 13C NMR (75 MHz, CDCl3): 14.4, 28.5, 28.9, 32.6,
52.6, 52.9, 60.6, 61.7, 63.8, 80.7, 86.9, 127.5, 128.1, 128.7, 143.6, 155.8,
171.2, 171.8, 172.5. Anal. calcd for C35H42N2O8: C, 67.94; H, 6.84; N,
4.53; found: C, 67.70; H, 6.67; N, 4.68.
C
15H26N2O7: C, 52.01; H, 7.57; N, 8.09; found: C, 52.30; H, 7.68; N,
8.17.
5.5. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-1-
oxo-3-phenylpropan-2-ylamino)-5-oxopentanoate (25)
5.10. (S)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-1-
Yield: 76%; yellow oil; Rf (cyclohexane/EtOAc 6/4): 0.3;
oxo-3-(trityloxy)propan-2-ylamino)-5-oxopentanoate (30)
[
a]
20: þ37.5 (c: 1.00 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t,
D
J ¼ 7.0, 3H); 1.45 (s, 9H); 1.85e2.00 (m, 1H); 2.10e2.20 (m, 1H);
2.20e2.30 (m, 2H); 3.10 (dd, J ¼ 6.8, 14.1, 1H); 3.15 (dd, J ¼ 5.4, 14.1,
1H); 3.75 (s, 3H); 4.20 (q, J ¼ 7.0, 2H); 4.35e4.45 (m, 1H); 4.80e4.90
(m, 1H); 5.20 (bd, J ¼ 6.9, 1H); 6.35 (bd, J ¼ 5.8, 1H); 7.10e7.15 (m,
2H); 7.20e7.30 (m, 3H); 13C NMR (75 MHz, CDCl3): 14.4, 28.5, 29.7,
32.6, 38.0, 52.6, 53.1, 53.8, 61.8, 80.4,127.3, 128.8, 129.5,136.3,156.2,
172.1, 172.4, 172.5. Anal. calcd for C22H32N2O7: C, 60.54; H, 7.39; N,
6.42; found: C, 60.21; H, 7.22; N, 6.54.
Yield: 77%; Solid state: yellow oil; Rf (cyclohexane/EtOAc 1/1):
0.63; [
a
]
20: ꢄ1.0 (c: 1.00 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25
D
(t, J ¼ 7.0, 3H); 1.45 (s, 9H); 1.80e2.00 (m, 1H); 2.10e2.40 (m, 3H);
3.35 (dd, J ¼ 3.2, 9.4,1H); 3.60 (dd, J ¼ 3.2, 9.4,1H); 3.75 (s, 3H); 4.20
(q, J ¼ 7.0, 2H); 4.35e4.45 (m, 1H); 4.72 (ddd, J ¼ 3.2, 3.2, 7.9, 1H);
5.35 (bd, J ¼ 7.9, 1H); 6.86 (bd, J ¼ 7.9, 1H); 7.20e7.40 (m, 15H); 13
C
NMR (75 MHz, CDCl3): 14.4, 28.5, 29.0, 32.5, 52.9, 53.1, 59.4, 61.8,
63.1, 80.6, 86.9, 127.5, 128.1, 128.9, 143.6, 156.1, 171.2, 171.2, 172.5.

8490
L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
Anal. calcd for C35H42N2O8: C, 67.94; H, 6.84; N, 4.53; found: C,
68.24; H, 7.00; N, 4.40.
5.16. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-1-
oxo-3-phenylpropan-2-ylamino)-5-oxopentanoate (36)
[
a
]
20: þ19.7 (c: 1.00 in CHCl3). Anal. calcd for C22H32N2O7: C,
D
5.11. (S)-Dimethyl 2-((S)-4-(tert-butoxycarbonylamino)-5-ethoxy-
5-oxopentanamido)pentanedioate (31)
60.54; H, 7.39; N, 6.42; found: C, 60.28; H, 7.23; N, 6.60.
Yield: 83%; crystallized from diisopropyl ether as white crystals;
5.17. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-1-
oxo-3-phenylpropan-2-ylamino)-5-oxopentanoate (37)
mp: 95e97 ꢃC; Rf (cyclohexane/EtOAc 4/6): 0.37; [
a]
20: þ21.4 (c:
D
1.00 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t, J ¼ 7.1, 3H); 1.45
(s, 9H); 1.90e2.10 (m, 2H); 2.15e2.25 (m, 2H); 2.30e2.40 (m, 2H);
2.40e2.50 (m, 2H); 3.65 (s, 3H); 3.75 (s, 3H); 4.20 (q, J ¼ 7.1, 2H);
4.20e4.30 (m, 1H); 4.60 (ddd, J ¼ 5.0, 7.6, 7.6, 1H); 5.25 (bd, J ¼ 7.6,
1H); 6.60 (bd, J ¼ 7.6, 1H). Anal. calcd for C19H32N2O9 C, 52.77; H,
7.46; N, 6.48; found: C, 52.96; H, 7.70; N, 6.25.
[
a
]
20: ꢄ37.0 (c: 1.00 in CHCl3). Anal. calcd for C22H32N2O7: C,
D
60.54; H, 7.39; N, 6.42; found: C, 60.30; H, 7.20; N, 6.61.
5.18. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-4-
methyl-1-oxopentan-2-ylamino)-5-oxopentanoate (38)
5.12. (R)-Dimethyl 2-((S)-4-(tert-butoxycarbonylamino)-5-ethoxy-
5-oxopentanamido)pentanedioate (32)
[
a
]
20: ꢄ13.1 (c: 1.10 in CHCl3). Anal. calcd for C19H34N2O7: C,
D
56.70; H, 8.51; N, 6.96; found: C, 56.93; H, 8.68; N, 7.15.
Yield: 73%; crystallized from diisopropyl ether as white crystals;
mp: 75e77 ꢃC; Rf (cyclohexane/EtOAc 1/1): 0.19; [
a]
D
20: ꢄ6.5 (c: 1.00
5.19. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-4-
methyl-1-oxopentan-2-ylamino)-5-oxopentanoate (39)
in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t, J ¼ 7.0, 3H); 1.45 (s,
9H); 1.85e2.00 (m, 1H); 2.00e2.10 (m, 1H); 2.15e2.25 (m, 2H);
2.30e2.40 (m, 2H); 2.40e2.50 (m, 2H); 3.65 (s, 3H); 3.75 (s, 3H);
4.20 (q, J ¼ 7.0, 2H); 4.35e4.45 (m, 1H); 4.60 (ddd, J ¼ 5.0, 7.6, 7.6,
1H); 5.31 (bd, J ¼ 7.6, 1H); 6.75 (bd, J ¼ 7.6, 1H). Anal. calcd for
[
a
]
20: ꢄ5.3 (c: 1.00 in CHCl3). Anal. calcd for C19H34N2O7: C,
D
56.70; H, 8.51; N, 6.96; found: C, 57.00; H, 8.71; N, 7.20.
C
19H32N2O9 C, 52.77; H, 7.46; N, 6.48; found: C, 53.05; H, 7.70; N,
5.20. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((S)-1-methoxy-1-
oxo-3-(trityloxy)propan-2-ylamino)-5-oxopentanoate (40)
6.30.
20: þ1.3 (c: 1.00 in CHCl3). Anal. calcd for C35H42N2O8: C,
5.13. (S)-Methyl 6-(tert-butoxycarbonylamino)-2-((S)-4-(tert-
butoxycarbonylamino)-5-ethoxy-5-oxopentanamido)hexanoate
(33)
[
a
]
D
67.94; H, 6.84; N, 4.53; found: C, 68.15; H, 7.04; N, 4.38.
5.21. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-((R)-1-methoxy-1-
oxo-3-(trityloxy)propan-2-ylamino)-5-oxopentanoate (41)
Yield: 79%; yellow oil; Rf (cyclohexane/EtOAc 1/1): 0.33;
[
a]
20: þ8.0 (c: 0.50 in CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t,
D
J ¼ 7.0, 3H); 1.45 (s, 18H); 1.30e1.50 (m, 1H); 1.65e1.95 (m, 6H);
2.10e2.25 (m, 1H); 2.30e2.40 (m, 2H); 3.05e3.15 (m, 2H); 3.75 (s,
3H); 4.20 (q, J ¼ 7.0, 2H); 4.20e4.30 (m, 1H); 4.55 (ddd, J ¼ 5.0, 7.7,
7.7, 1H); 4.65 (bs., 1 H); 5.25 (bd, J ¼ 7.7, 1H); 6.50 (bd, J ¼ 7.7, 1H);
13C NMR (75 MHz, CDCl3): 14.4, 22.7, 28.5, 28.6, 29.6, 29.8, 31.9,
32.6, 40.3, 52.4, 52.5, 53.1, 61.8, 79.3, 80.4, 156.2, 156.3, 172.3, 172.6,
173.1. Anal. calcd for C24H43N3O9 C, 55.69; H, 8.37; N, 8.12; found: C,
55.91; H, 8.56; N, 7.90.
[
a
]
20: þ20.6 (c: 1.00 in CHCl3). Anal. calcd for C35H42N2O8: C,
D
67.94; H, 6.84; N, 4.53; found: C, 68.20; H, 6.98; N, 4.32.
5.22. (S)-Dimethyl 2-((R)-4-(tert-butoxycarbonylamino)-5-ethoxy-
5-oxopentanamido)pentanedioate (42)
[
a]
20: þ6.9 (c: 1.00 in CHCl3). Anal. calcd for C19H32N2O9 C,
D
52.77; H, 7.46; N, 6.48; found: C, 53.00; H, 7.70; N, 6.32.
5.14. (R)-Methyl 6-(tert-butoxycarbonylamino)-2-((S)-4-(tert-
butoxycarbonylamino)-5-ethoxy-5-oxopentanamido)hexanoate
(34)
5.23. (R)-Dimethyl 2-((R)-4-(tert-butoxycarbonylamino)-5-ethoxy-
5-oxopentanamido)pentanedioate (43)
[a]
20: e 21.0 (c: 0.95 in CHCl3). Anal. calcd for C19H32N2O9 C,
D
52.77; H, 7.46; N, 6.48; found: C, 52.94; H, 7.65; N, 6.34.
Yield: 87%; crystallized from diisopropyl ether as white crystals;
mp: 75 ꢃC; Rf (cyclohexane/EtOAc 1/1): 0.40; [
a]
20: þ5.3 (c: 0.50 in
D
CHCl3); 1H NMR (300 MHz, CDCl3): 1.25 (t, J ¼ 7.1, 3H); 1.45 (s, 18H);
1.35e1.50 (m, 1H); 1.65e1.95 (m, 6H); 2.10e2.25 (m, 1H); 2.30e2.40
(m, 2H); 3.05e3.15 (m, 2H); 3.75 (s, 3H); 4.20 (q, J ¼ 7.1, 2H);
4.35e4.45 (m, 1H); 4.55 (ddd, J ¼ 5.0, 7.4, 7.4, 1H); 4.65 (bs., 1 H);
5.30 (bd., J ¼ 6.6, 1H); 6.90 (bd., J ¼ 6.6, 1H); 13C NMR (75 MHz,
CDCl3): 14.4, 22.7, 28.5, 28.6, 29.5, 29.8, 31.9, 32.6, 40.3, 52.4, 52.5,
53.1, 61.8, 79.3, 80.4, 156.2, 156.3, 172.3, 172.5, 173.1. Anal. calcd for
5.24. (S)-Methyl 6-(tert-butoxycarbonylamino)-2-((R)-4-(tert-
butoxycarbonylamino)-5-ethoxy-5-oxopentanamido)heptanoate
(44)
[a]
20: e 5.0 (c: 0.50 in CHCl3). Anal. calcd for C24H43N3O9 C,
D
55.69; H, 8.37; N, 8.12; found: C, 55.84; H, 8.49; N, 8.00.
C
24H43N3O9 C, 55.69; H, 8.37; N, 8.12; found: C, 55.99; H, 8.50; N,
7.95.
5.25. (R)-Methyl 6-(tert-butoxycarbonylamino)-2-((R)-4-(tert-
butoxycarbonylamino)-5-ethoxy-5-oxopentanamido)heptanoate
(45)
5.15. (R)-Ethyl 2-(tert-butoxycarbonylamino)-5-(2-methoxy-2-
oxoethylamino)-5-oxopentanoate (35)
[
a
]
20: ꢄ11.3 (c: 1.00 in CHCl3). Anal. calcd for C15H26N2O7: C,
[
a
]
20: ꢄ8.0 (c: 0.50 in CHCl3). Anal. calcd for C24H43N3O9 C,
D
D
52.01; H, 7.57; N, 8.09; found: C, 52.27; H, 7.70; N, 8.22.
55.69; H, 8.37; N, 8.12; found: C, 55.90; H, 8.52; N, 7.95.

L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
8491
5.26. (S)-2-Amino-5-(carboxymethylamino)-5-oxopentanoic acid
(1)
3H); 4.38 (dd, J ¼ 4.1, 4.1, 1H); 13C NMR (75 MHz, D2O): 26.5, 31.9,
54.5, 57.3, 62.1, 174.3, 174.6, 176.4; [MþH]þ: 235.0. [
a
]
20: ꢄ13 (c:
D
0.35 in H2O). Anal. calcd for C8H14N2O6 C, 41.03; H, 6.03; N, 11.96;
found: C, 41.30; H, 6.22; N, 11.78.
Yield: 31%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.18; mp: dec.
T > 159 ꢃC; [
a
]
20: þ10.0 (c: 0.30 in H2O); lit.18
[
a
]
20: þ11.1 (c: 1.00 in
D
D
H2O); 1H NMR (300 MHz, D2O): 2.00e2.08 (m, 2H); 2.35 (ddd,
J ¼ 1.4, 8.5, 8.5, 2H); 3.70 (dd, J ¼ 6.3, 6.3, 1H); 3.80 (s, 2H); 13C NMR
(75 MHz, D2O): 26.1, 31.2, 41.5, 53.6, 173.3, 174.0, 175.2; [MþH]þ:
205.0. Anal. calcd for C7H12N2O5 C, 41.18; H, 5.92; N, 13.72; found: C,
41.00; H, 5.80; N, 13.89.
5.32. (S)-2-Amino-5-((R)-1-carboxy-2-hydroxyethylamino)-5-
oxopentanoic acid (7)
Yield: 40%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.26; mp: dec.
T > 156 ꢃC; [
a]
D
20: þ12.3 (c: 0.40 in H2O); 1H NMR (300 MHz, D2O):
2.00e2.10 (m, 2H); 2.40 (ddd, J ¼ 3.5, 6.2, 6.2, 2H); 3.70e3.85 (m,
5.27. (S)-2-Amino-5-((S)-1-carboxy-2-phenylethylamino)-5-
oxopentanoic acid (2)
3H); 4.38 (dd, J ¼ 4.1, 4.1, 1H); 13C NMR (75 MHz, D2O): 25.9, 31.0,
53.2, 55.1, 61.2, 172.9, 173.8, 174.6; [MþH]þ: 235.0. [
a
]
20: ꢄ13 (c:
D
0.35 in H2O). Anal. calcd for C8H14N2O6 C, 41.03; H, 6.03; N, 11.96;
found: C, 41.28; H, 6.20; N, 11.80.
Yield: 80%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.49; [
a
]
20: þ18.2
D
(c: 0.25 in H2O); mp: T > 186e189 ꢃC; 1H NMR (300 MHz, D2O):
1.80e1.90 (m, 2H); 2.25 (m, 2H); 2.85 (dd, J ¼ 9.3,14.0,1H); 3.10 (dd,
J ¼ 5.3, 14.0, 1H); 3.60 (dd, J ¼ 6.5, 6.5, 1H); 4.55 (dd, J ¼ 5.3, 9.3 1H);
7.10e7.30 (m, 5H); 13C NMR (75 MHz, D2O): 26.1, 31.3, 36.8, 53.2,
54.3, 127.3, 128.8, 129.3, 136.8, 172.7, 174.2, 175.3; [MþH]þ: 295.0.
Anal. calcd for C14H18N2O5 C, 57.13; H, 6.16; N, 9.52; found: C, 56.90;
H, 6.28; N, 9.40.
5.33. (S)-2-((S)-4-Amino-4-carboxybutanamido)pentanedioic acid
(8)
Yield: 59%, Rf (n-butanol/H2O/AcOH 4:2:1): 0.29; [
a
]
]
20: ꢄ2.5 (c:
D
0.50 in H2O); [
a]
20: þ3.2 (c: 0.50 in 0.5 N HCl); lit.20
[
a
20: þ3.4 (c:
D
D
1.00 in 0.5 N HCl); mp: 188e190 ꢃC; 1H NMR (300 MHz, D2O):
1.80e1.95 (m, 1H); 2.00e2.15 (m, 3H); 2.30e2.45 (m, 4H); 3.80 (dd,
J ¼ 6.3, 6.3, 1H); 4.25 (dd, J ¼ 5.0, 9.1, 1H); 13C NMR (75 MHz, D2O):
25.9, 26.0, 30.3, 31.3, 52.5, 53.5, 173.1, 174.7, 175.5, 177.3; [MþH]þ:
277.0. Anal. calcd for C10H16N2O7 C, 43.48; H, 5.84; N, 10.14; found:
C, 43.70; H, 6.00; N, 9.98.
5.28. (S)-2-Amino-5-((R)-1-carboxy-2-phenylethylamino)-5-
oxopentanoic acid (3)
Yield: 89%; Rf (CH2Cl2/MeOH 9:1 þ 1% AcOH): 0.62; [
a
]
20: þ3.7
D
(c: 0.30 in H2O); mp: 150 ꢃC; 1H NMR (300 MHz, D2O): 1.80e2.00
(m, 2H); 2.28 (ddd, J ¼ 3.0, 8.2, 8.2, 2H); 2.82 (dd, J ¼ 9.7, 14.1, 1H);
3.12 (dd, J ¼ 5.3, 14.1, 1H); 3.60 (dd, J ¼ 3.0, 6.5, 1H); 4.55 (dd, J ¼ 5.3,
9.7,1H); 7.10e7.25 (m, 5H); 13C NMR (75 MHz, D2O): 25.7, 30.4, 36.8,
52.2, 54.0, 127.2, 128.8, 129.4, 136.8, 171.8, 174.0, 175.1; [MþH]þ:
295.0. Anal. calcd for C14H18N2O5 C, 57.13; H, 6.16; N, 9.52; found: C,
57.32; H, 5.98; N, 9.78.
5.34. (S)-2-((R)-4-Amino-4-carboxybutanamido)pentanedioic acid
(9)
Yield: 44%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.29; [
a
]
20: þ15.2
D
(c: 0.25 in H2O); mp: 103e105 ꢃC; 1H NMR (300 MHz, D2O):
1.80e1.95 (m, 1H); 2.00e2.10 (m, 3H); 2.30e2.40 (m, 4H); 3.80 (dd,
J ¼ 6.3, 6.3, 1H); 4.25 (dd, J ¼ 5.0, 9.1, 1H); 13C NMR (75 MHz, D2O):
25.9, 26.0, 30.2, 31.0, 52.4, 53.3, 172.9, 174.6, 175.5, 177.3; [MþH]þ:
277.0. Anal. calcd for C10H16N2O7 C, 43.48; H, 5.84; N, 10.14; found:
C, 43.75; H, 6.06; N, 9.90.
5.29. (S)-2-Amino-5-((S)-1-carboxy-3-methylbutylamino)-5-
oxopentanoic acid (4)
Yield: 100%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.56; [
a
]
20: ꢄ15.0
D
(c: 0.25 in H2O); lit.19
[
a]
D
20: ꢄ13.5 (c: 1.00 in H2O); mp: 187 ꢃC; 1H
5.35. (S)-6-Amino-2-((S)-4-amino-4-carboxybutanamido)
hexanoic acid (10)
NMR (300 MHz, D2O): 0.70 (d, J ¼ 5.8, 3H); 0.80 (d, J ¼ 5.8, 3H);
1.45e1.55 (m, 3H); 1.95e2.05 (m, 2H); 2.30e2.40 (m, 2H); 3.72 (dd,
J ¼ 6.2, 6.2, 1H); 4.20 (dd, J ¼ 6.4, 6.4, 1H); 13C NMR (75 MHz, D2O):
20.7, 22.3, 24.6, 26.2, 31.3, 39.5, 51.9, 53.5, 173.1, 174.7, 177.0;
[MþH]þ: 261.0. Anal. calcd for C11H20N2O5 C, 50.76; H, 7.74; N,
10.76; found: C, 50.45; H, 7.60; N, 10.50.
The residue was dissolved in a few drops of bidistilled water and
purified on a column packed with Amberlite IR120. The resin was
washed with bidistilled H2O until the pH was neutral and then the
product was released from the resin with aqueous 1 N NH3. The
fractions containing the dipeptide were collected and lyophilized to
give the desired product.
5.30. (S)-2-Amino-5-((R)-1-carboxy-3-methylbutylamino)-5-
oxopentanoic acid (5)
Yield: 79%, Rf (n-butanol/H2O/AcOH 4:2:1): 0.14, [
a
]
20: þ4.0 (c:
D
0.20 in H2O); lit.21
[
a]
20: þ3.5 (c: 1.00 in H2O); mp: dec. T > 215 ꢃC;
D
Yield: 86%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.56; [
a
]
20: þ35.0
1H NMR (300 MHz, D2O): 1.25e1.35 (m, 2H); 1.50e1.60 (m, 3H);
1.60e1.70 (m, 1H); 1.95e2.05 (m, 2H); 2.30e2.40 (m, 2H); 2.86
(ddd, J ¼ 7.6, 7.6, 7.6, 2H); 3.62 (dd, J ¼ 6.5, 6.5, 1H); 4.00 (dd, J ¼ 5.3,
8.5, 1H); 13C NMR (75 MHz, D2O): 22.3, 26.5, 26.5, 31.0, 31.9, 39.4,
54.4, 55.2, 174.2, 174.4, 179.2; [MþH]þ: 276.0. Anal. calcd for
D
(c: 0.25 in H2O); mp: 100e102 ꢃC; 1H NMR (300 MHz, D2O): 0.70 (d,
J ¼ 6.2, 3H); 0.80 (d, J ¼ 6.2, 3H); 1.45e1.55 (m, 3H); 1.95e2.10 (m,
2H); 2.35 (ddd, J ¼ 1.8, 8.8, 8.8, 2H); 3.80 (dd, J ¼ 6.4, 6.4, 1H); 4.22
(dd, J ¼ 7.0, 7.0, 1H); 13C NMR (75 MHz, D2O): 20.6, 22.3, 24.5, 25.9,
30.9, 39.4, 51.7, 53.0,172.5,174.5, 176.9; [MþH]þ: 261. Anal. calcd for
C
11H21N3O5 C, 47.99; H, 7.69; N, 15.26; found: C, 47.70; H, 7.48; N,
C
11H20N2O5 C, 50.76; H, 7.74; N, 10.76; found: C, 50.98; H, 7.90; N,
15.46.
10.53.
5.36. (R)-6-Amino-2-((S)-4-amino-4-carboxybutanamido)
5.31. (S)-2-Amino-5-((S)-1-carboxy-2-hydroxyethylamino)-5-
hexanoic acid (11)
oxopentanoic acid (6)
Yield: 65%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.14; [
a
]
20: ꢄ4.5 (c:
D
Yield: 40%; Rf (n-butanol/H2O/AcOH 4:2:1): 0.24; mp: dec.
0.35 in H2O); mp: dec. T > 205 ꢃC; 1H NMR (300 MHz, D2O):
1.20e1.40 (m, 2H); 1.50e1.60 (m, 2H); 1.60e1.75 (m, 2H); 1.95e2.05
(m, 2H); 2.25e2.35 (m, 2H); 2.86 (ddd, J ¼ 7.3, 7.3, 7.3, 2H); 3.66 (dd,
T > 164 ꢃC; [
a
]
D
20: þ11.6 (c: 0.35 in H2O); 1H NMR (300 MHz, D2O):
2.00e2.10 (m, 2H); 2.40 (ddd, J ¼ 3.5, 6.2, 6.2, 2H); 3.70e3.85 (m,

8492
L. Tamborini et al. / Tetrahedron 72 (2016) 8486e8492
J ¼ 5.9, 5.9, 1H); 4.02 (dd, J ¼ 5.0, 8.2, 1H); 13C NMR (75 MHz, D2O):
22.3, 26.4, 26.5, 31.0, 31.2, 39.4, 54.3, 55.2, 174.2, 174.9, 179.2;
[MþH]þ: 276.0. Anal. calcd for C11H21N3O5 C, 47.99; H, 7.69; N,
15.26; found: C, 47.75; H, 7.50; N, 15.50.
5.46. (S)-6-Amino-2-((R)-4-amino-4-carboxybutanamido)
hexanoic acid (21)
The residue was dissolved in a few drops of bidistilled water and
purified on a column packed with Amberlite IR120. The resin was
washed with bidistilled H2O until the pH was neutral and then the
product was released from the resin with aqueous 1 N NH3. The
fractions containing the dipeptide were collected and lyophilized to
give the desired product.
5.37. (R)-2-Amino-5-(carboxymethylamino)-5-oxopentanoic acid
(12)
[
a
]
20: ꢄ10.0 (c: 0.30 in H2O). Anal. calcd for C7H12N2O5 C, 41.18;
D
[
a
]
20: þ4.0 (c: 0.35 in H2O). Anal. calcd for C11H21N3O5 C, 47.99;
D
H, 5.92; N, 13.72; found: C, 40.98; H, 5.81; N, 13.93.
H, 7.69; N, 15.26; found: C, 48.23; H, 7.90; N, 15.00.
5.38. (R)-2-Amino-5-((S)-1-carboxy-2-phenylethylamino)-5-
oxopentanoic acid (13)
5.47. (R)-6-Amino-2-((R)-4-amino-4-carboxybutanamido)
hexanoic acid (22)
[
a
]
20: ꢄ5.0 (c: 0.25 in H2O). Anal. calcd for C14H18N2O5 C, 57.13;
D
H, 6.16; N, 9.52; found: C, 57.38; H, 6.30; N, 9.40.
[
a]
20: ꢄ4.0 (c: 0.20 in H2O). Anal. calcd for C11H21N3O5 C, 47.99;
D
H, 7.69; N, 15.26; found: C, 48.18; H, 7.93; N, 14.98.
5.39. (R)-2-Amino-5-((R)-1-carboxy-2-phenylethylamino)-5-
oxopentanoic acid (14)
References
[
a
]
20: ꢄ17.1 (c: 0.30 in H2O). Anal. calcd for C14H18N2O5 C, 57.13;
D
H, 6.16; N, 9.52; found: C, 57.30; H, 6.00; N, 9.80.
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2. Brauner-Osborne H, Egebjerg J, Nielsen EØ, Madsen U, Krogsgaard-Larsen P.
5.40. (R)-2-Amino-5-((S)-1-carboxy-3-methylbutylamino)-5-
oxopentanoic acid (15)
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ꢀ
[a
]
D
20: ꢄ33.0 (c: 0.25 in H2O). Anal. calcd for C11H20N2O5 C, 50.76;
H, 7.74; N, 10.76; found: C, 50.93; H, 7.90; N, 10.50.
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10. Dolman NP, More JC, Alt A, et al. J Med.Chem. 2006;49:2579.
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5.41. (R)-2-Amino-5-((R)-1-carboxy-3-methylbutylamino)-5-
oxopentanoic acid (16)
[
a
]
20: þ15.6 (c: 0.25 in H2O). Anal. calcd for C11H20N2O5 C, 50.76;
D
H, 7.74; N, 10.76; found: C, 50.50; H, 7.58; N, 10.87.
5.42. (R)-2-Amino-5-((S)-1-carboxy-2-hydroxyethylamino)-5-
oxopentanoic acid (17)
[
a
]
20: ꢄ12.3 (c: 0.40 in H2O). Anal. calcd for C8H14N2O6 C, 41.03;
D
H, 6.03; N, 11.96; found: C, 40.80; H, 5.80; N, 12.10.
5.43. (R)-2-Amino-5-((R)-1-carboxy-2-hydroxyethylamino)-5-
oxopentanoic acid (18)
iii) Rajca A, Wiessler M. Carbohydr Res. 1995;274:123.
[
a
]
20: ꢄ13.0 (c: 0.35 in H2O). Anal. calcd for C8H14N2O6 C, 41.03;
D
13. Baxendale IR, Ley SV, Smith CD, Tranmer GK. Chem Commun. 2006;46:4835.
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15. i) Shimamoto K, Lebrun B, Yasuda-Kamatani Y, et al. Mol Pharmacol. 1998;53:
195;
H, 6.03; N, 11.96; found: C, 40.85; H, 5.82; N, 12.16.
5.44. (R)-2-((S)-4-Amino-4-carboxybutanamido)pentanedioic acid
(19)
ii) Colleoni S, Jensen AA, Landucci E, et al. J Pharmacol Exp Ther. 2008;326:646;
iii) Pinto A, Conti P, De Amici M, et al. Tetrahedron Asymmetry. 2008;19:867;
iv) Tamborini L, Conti P, Pinto A, Colleoni S, Gobbi M, De Micheli C. Tethrahe-
dron. 2009;65:6083;
[
a
]
20: ꢄ16.0 (c: 0.25 in H2O). Anal. calcd for C10H16N2O7 C, 43.48;
D
H, 5.84; N, 10.14; found: C, 43.69; H, 5.98; N, 9.97.
v) Tsukada S, Iino M, Takayasu Y, Shimamoto K, Ozawa S. Neuropharmacology.
2005;48:479.
5.45. (R)-2-((R)-4-amino-4-carboxybutanamido)pentanedioic acid
(20)
€
16. Jensen AA, Brauner-Osborne H. Biochem Pharmacol. 2004;67:2115.
_
17. Assaf Z, Larsen AP, Venskutonyte R, et al. J Med Chem. 2013;56:1614.
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19. Rowlands DA, Young GT. J Chem Soc. 1950:3937.
20. Waley SG. J Chem Soc. 1955:517.
[
a
]
20: þ3.0 (c: 0.50 in H2O). Anal. calcd for C10H16N2O7 C, 43.48;
D
H, 5.84; N, 10.14; found: C, 43.80; H, 6.00; N, 9.93.
21. Chaturvedi NC, Khosla MC, Anand N. Ind J Chem. 1965;3:554.