Novel and facile solution-phase synthesis of 2,5-diketopiperazines and O-glycosylated analogs

Article abstract of DOI:10.1016/j.tet.2009.04.069

This work describes the synthesis in solution of a series of related diketopiperazines with potential biological activities: cyclo(l-Pro-l-Ser), cyclo(l-Phe-l-Ser), cyclo(d-Phe-l-Ser) and the corresponding glycosylated analogs of the latter, cyclo[d-Phe-l

Full text of DOI:10.1016/j.tet.2009.04.069

Tetrahedron 65 (2009) 5343–5349
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Novel and facile solution-phase synthesis of 2,5-diketopiperazines
and O-glycosylated analogs
*
Vanessa L. Campo, Maristela B. Martins, Carlos H.T.P. da Silva, Ivone Carvalho
Faculdade de Cieˆncias Farmaceˆuticas de Ribeir˜ao Preto, Universidade de S˜ao Paulo, Av. do Cafe´ s/n, Ribeir˜ao Preto 14040-930, Brazil
a r t i c l e i n f o
a b s t r a c t
Article history:
This work describes the synthesis in solution of a series of related diketopiperazines with potential
Received 5 February 2009
Received in revised form 13 April 2009
Accepted 20 April 2009
biological activities: cyclo(
glycosylated analogs of the latter, cyclo[
synthetic approach involved coupling reactions of –OH or O-glycosylated serine benzyl esters with
NFmoc-protected amino acids (Pro or Phe), followed by one-pot deprotection–cyclization reaction in the
presence of 20% piperidine in DMF.
L
-Pro-
L
-Ser), cyclo(
L
-Phe-
L
-Ser), cyclo(
D
-Phe-
L
-Ser) and the corresponding
D-Phe-
L-Ser(
a
GlcNAc)] and cyclo[
D
-Phe- -Ser( GlcNAc)]. The
L
b
Available online 3 May 2009
Keywords:
Diketopiperazine
Ó 2009 Elsevier Ltd. All rights reserved.
Dipeptide cyclization
Glycosylated amino acid
Solution-phase synthesis
1. Introduction
chains as well as nearly any other type of substituents of interest can
be incorporated into a DKP scaffold, comprising a versatile strategy to
2,5-Diketopiperazines (DKPs) are peculiar heterocyclic systems
presenting spatially defined substituents around a proteolysis-re-
sistant peptide-mimicking scaffold, characteristically derived from
the cross-link between two amino acid residues via a pair of
complementary amide bonds. Diketopiperazine derivatives are
known by properties such as antitumor, antimicrobial, and antiviral
activities, and modulation of enzymes, receptors, and biochemical
mediators. These naturally occurring cyclic dipeptides are consid-
ered privileged structures for medicinal chemistry, since the DKP
backbone is straightforwardly obtained and provides a stable and
rigid core that can be conveniently functionalized at different po-
sitions and configurations.1–5
Besides, being small and fairly constrained structures with mod-
erately polar groups like amide, particularly when functionalized
with balanced hydrophilic and lipophilic residues, DKP derivatives
give rise to valuable physicochemical properties for bioavailable
compounds, which may include carrier pro-drugs and blood–brain
barrier shuttles.1,6 Thus, DKPs constitute a rich source of new phar-
macologically active compounds that can thrust drug discovery.
Frequent positive hits in biological screens raised the interest for
DKP motifs, which demands the development of efficient synthetic
methods leading to diversely substituted diketopiperazines.1 In ad-
ditiontoordinaryaminoacidfunctionalities, unusualormodifiedside
display pharmacophoric groups in a controlled fashion and increase
both selectivity and binding affinity toward biological targets.7,8 De-
spite numerous derivatives from natural origin, further structural
variety can be achieved by combinatorial chemistry through a num-
ber of conventional procedures that lead to head-to-tail cyclization of
selected dipeptides, both in solution and solid-phase.5,9
Although many reports describe support-based general methods
for synthesizing DKP combinatorial libraries,10 problems with
scaling up are important shortcomings9 and the advantages of solid-
phase reactions concerning to peptide chemistry do not completely
apply to diketopiperazines, as the attachment of the growing chain
and its cleavage from the resin introduces worthless steps for the in
situ generation of only one or two peptide bonds intended for the
product. Synthetic approaches in solution leading to DKPs are
scarcer, except for novel strategies involving Ugi multicomponent
reactions11,12 and DKP enolates alkylation,13,14 which might, in turn,
provide poor stereocontrol because generally do not employ linear
dipeptides with predefined configuration as precursors. In parallel,
microwave-promoted cyclization reactions have been increasingly
used with promising results.3,9,15 Notwithstanding, comprehensive
procedures in satisfactory yields for conveniently substituted DKP
by homogeneous reactions are still lacking.9
Hence, this study exploited the synthesis in solution of a series
of related diketopiperazines cyclo(
Ser) (2), cyclo( -Phe- -Ser) (3), and the corresponding O-glycosylated
analogs of the latter, cyclo[ -Phe- -Ser( GlcNAc)] (4) and cyclo
-Phe- -Ser( GlcNAc)] (5) (Fig. 1).
L-Pro-L-Ser) (1), cyclo(L-Phe-L-
D
L
D
L
a
* Corresponding author. Tel.: þ55 16 36024709; fax: þ55 16 36024879.
E-mail address: carronal@usp.br (I. Carvalho).
[D
L
b
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.04.069

5344
V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
decoration with several types of substituents of interest.1,9 Fur-
thermore, the interest for these amino acids is also justified by their
expressive presence in biologically active diketopiperazines, either
isolated from natural source or synthetically accessed.2,19
Diketopiperazines are occasionally formed as byproducts in
solid-phase peptide synthesis (SPPS) after removal of the amino
protecting group under basic cleavage.20 Thus, our approach
exploited this condition to extend the well-known SPPS protocols to
solution-phase synthesis, employing the stable OBn group for C-
terminal orthogonal protection instead of the solid support.
Considering the importance of the nucleophilic strength of the N-
terminal nitrogen for efficient cyclization,3 some commonly used N-
protecting groups, such as Boc and Alloc, require further treatment
with organic bases to induce cyclization after their acid cleavage.20
Differently, Fmoc group is advantageous due to its cleavage under
basic conditions that allow performing dipeptide deprotection and
cyclization in a single step.
2.1. Synthesis of diketopiperazines cyclo(
L-Pro-L-Ser) 1,
cyclo( -Phe- -Ser) 2, and cyclo( -Phe- -Ser) 3
L
L
D
L
The synthesis of diketopiperazines 1, 2, and 3 was developed in
three steps (Scheme 1), by the treatment of Fmoc– -serine benzyl
ester (6) with 20% piperidine in DMF, followed by coupling of the
resultant -Ser–OBn (7) (94%) with the amino acids Fmoc– -proline
(8), Fmoc– -phenylalanine (9), or Fmoc- -phenylalanine (10), re-
spectively, in the presence of the coupling reagents PyBOP, HOBt,
and DIEA in acetonitrile.20 The dipeptides Fmoc–
-Pro- -Ser–OBn
(11), Fmoc– -Phe- -Ser–OBn (12), and Fmoc– -Phe- -Ser–OBn (13)
L
L
L
L
D
Figure 1. Diketopiperazines and O-glycosylated analogs prepared by solution-phase
synthesis.
L
L
2. Results and discussion
L
L
D
L
were obtained in 80, 89, and 85% yields, respectively, and their
Selection of amino acid residues was carefully based on chemical
reactivity and equilibrium between hydrophilic and hydrophobic
properties, characteristic of most drugs, and desired features in
eventual hits. Residues such as proline, phenylalanine, and serine,
brought together in DKPs, suitably combine affinity for aqueous and
organic phases, besides belonging to representative classes of
amino acids, according to side chain chemical properties. In a sense,
structural variety was narrowed by the preference for residues
bearing neutral character side chains under all reaction conditions
of the synthetic route.
The aliphatic amino acid proline is often emphasized by the
peculiarity of its secondary amino group, which induces amide
bonds to adopt a cis conformation favoring intramolecular cycliza-
tion into bicyclic diketopiperazine structures.9,10,16 Aromatic resi-
dues, including the simplest phenylalanine, are known to establish
non-covalent contacts that assist molecular folding, self-assembly,
and ligand–receptor interaction.17,18 Reactive side chain functions,
such as the hydroxyl group in serine, comprise versatile sites for
structures were confirmed by 1H NMR spectroscopy.
The dipeptides 11,12, and 13 were easily cyclized in the presence
of 20% piperidine in DMF. Considering that this step involves the
well known NFmoc deprotection followed by intramolecular cy-
clization, it demands longer reaction times (15–18 h) compared to
the single cleavage of Fmoc group. It is suggested that, favored by
the basic reaction condition, the dipeptide reacts intramolecularly
by nucleophilic attack of the free amino group toward the terminal
carbonyl with displacement of benzyloxy group, giving a stable six-
membered ring. Nevertheless, some N-deprotected dipeptides may
not undergo cyclization, remaining as linear byproducts.
The formed diketopiperazines 1, 2, and 3 were thus obtained in
respective yields of 68, 57, and 53%. The absence of aromatic and
alkylic hydrogens of both NFmoc and particularly OBn protecting
groups in the 1H NMR spectra of 1, 2, and 3 indicated the occurrence
of cyclization. Subsequently, their structures were confirmed by
ESI-MS, which displayed the characteristic adducts [MþNa]þ
207.07 for 1 and [MþH]þ 235.10 for 2 and 3.
Scheme 1. Synthesis of diketopiperazines 1, 2, and 3. Reagents and conditions: (i) 20% piperidine/DMF, rt; (ii) PyBOP, HOBt, DIEA/MeCN, rt.

V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
5345
Figure 2. Structures of diketopiperazine 2 in the two main low-energy conformations. In (A), the folded conformer solvated (one layer) with DMSO, and the overall system ge-
ometry-optimized at AM1 level of calculation. After the simulation, short distance of 2.69 Å was obtained for the CH/ contact, here represented by a dashed line. Hydrogen atoms
of the DMSO molecules were omitted for the sake of the clarity. In (B), the extended conformer geometry-optimized in gas phase at B3LYP/6-31G level of calculation is shown.
p
*
The aromatic containing diketopiperazines 2 and 3 showed re-
markable differences between their serine methylenic signals, due
to the conformational restriction imposed by ring closure, fixing
the amino acid side chains in defined cis and trans configurations,
fixed compound 2 and subsequently relaxing all the solute–solvent
system. From this resulting optimized system and for the sake of
calculation limits, only the first layer of DMSO was energy mini-
mized together with compound 2 using a more robust algorithm,
the semi-empirical AM1 model. On the other hand, the extended
respectively. CH2 Ser chemical shifts were
comparable to both linear dipeptide precursors 12 (
and 13 ( 3.83 and 3.65), whilst the corresponding diastereoisomer
2 showed 3.30 and 2.83 values. This considerable shielding effect
d
3.59 and 3.42 for 3,
d
3.81 and 3.69)
conformer was fully optimized in gas phase at B3LYP/6-31G
of calculation.22 A short distance of 2.69 Å was obtained for the CH/
contact, thus evidenced with the simulation in explicit solvation,
*
level
d
d
p
observed for compound 2 indicated that, after cyclization, serine
and whose result agrees with the expected and reported values
methylenic hydrogens are influenced by the paramagnetic current
obtained for this kind of interaction.21
field of phenylalanine benzene ring, through a CH/
p
interaction.21
The folded conformation of 2 led to spatial proximity of meth-
ylenic serine hydrogens toward the shielding region of the aromatic
ring field of phenylalanine residue, thus explaining the observed
effect.9,14 Results of the simulations for the two low-energy con-
formers of diketopiperazine 2 are shown in Figure 2.
For better understanding this different behavior at 1H NMR spectra,
we underwent some conformational analysis of compound 2.
2.2. Cyclization-derived conformational effects in 2
In order to investigate the energetic profiles of diketopiperazine
2 in both solvated and non-solvated conditions, this compound was
initially submitted to a conformational search in gas phase. Fol-
lowing this approach, the MMFF molecular mechanics model
implemented in the Spartan ’06 software22 is successful in assigning
low-energy conformers and in providing quantitative estimates of
conformational energy differences, and it has been the method of
choice for large scale conformational surveys.23,24 Thus, two low-
energy conformations, one extended and another folded, have been
preferentially obtained for compound 2. As a second step of the
simulation, the folded conformer was immersed in a boundary
condition containing five layers of DMSO molecules surrounding
the diketopiperazine. The overall system was then energy mini-
mized using the CVFF force field thus implemented in the DIS-
COVER3 module of the Insight II package,25 initially maintaining
2.3. Synthesis of O-glycosylated diketopiperazines cyclo-
[D
-Phe-L-Ser(aGlcNAc)] 4 and cyclo[D-Phe-L-Ser(bGlcNAc)] 5
Glycosylated amino acids such as O-GlcNAc-Ser/Thr are of utmost
importance considering their involvement in cellular recognition,
signaling, and regulation processes.26,27 The will to investigate their
display on DKP core prompted us to use the glycosylation method
previously reported by our group28,29 to prepare the building blocks
Fmoc–L-Ser(aGlcNAc)–OBn (14) and Fmoc–L-Ser(bGlcNAc)–OBn
(15). As a replacement for serine amino acid 6, these compounds
could be readily employed for the synthesis of glycosylated DKPs.
The glycosylated amino acids 14 and 15 were prepared by gly-
cosylation reaction using aGlcNAcCl (16) and the amino acid 6, in
the presence of HgBr2 as catalyst28,29 (Scheme 2), being obtained in
20 and 40% yields, respectively, after the separation of the anomeric
Scheme 2. Synthesis of the building blocks 14 and 15 by glycosylation reaction. Reagents and conditions: (i) HgBr2, DCE, reflux 9 h.

5346
V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
Scheme 3. Synthesis of diketopiperazines 4 and 5. Reagents and conditions: (i) 20% piperidine/DMF, rt; (ii) Fmoc-D-Phe-OH (10), PyBOP, HOBt, DIEA/MeCN, rt.
Table 1
mixture by chromatographic column. The assignment of the gly-
cosidic bond configuration of 14 and 15 was based on the coupling
constants of the anomeric hydrogens in the 1H NMR spectra, re-
Cyclization reactions under conventional and microwave conditions
Cyclized product
Conventional
Time (h)
Microwave
Time (min)
spectively, at
The NFmoc deprotection of the glycosylated amino acids 14 and
15 with 20% piperidine in DMF gave the products -Ser( GlcNAc)–
OBn (17) (74%) and -Ser( GlcNAc)–OBn (18) (85%), which were
subsequently treated with the amino acid 10 under coupling
conditions20 (Scheme 3), affording the glycodipeptides Fmoc–
Phe- -Ser( GlcNAc)–OBn (19) and Fmoc– -Phe- -Ser( GlcNAc)–
OBn (20) in 75 and 68%, respectively.
The structures of the obtained glycodipeptides were confirmed
by 1H NMR spectra, being observed characteristic doublets at
4.70
(J1,2 3.7) and 4.62 (J1,2 7.9) relative to H-1 of 19 and 20, respectively,
d 4.70 (J1,2 3.6 Hz) and d 4.67 (J1,2 8.4 Hz).
Yield (%)
Yield (%)
1
2
3
4
5
15
18
18
41
68
68
57
53
35
43
4
4
4
4
4
82
78
70
da
da
L
a
L
b
D
-
a
Product not obtained.
L
a
D
L
b
The cyclization reactions of the dipeptide precursors in the
presence of 20% piperidine in DMF were successfully carried out in
open vessels under microwave irradiation (200 W, 163 ꢀC), which
led to the total consumption of the starting materials 11, 12, and 13
after 4 min and to the generation of the corresponding cyclized
products 1, 2, and 3 as major products in high and satisfactory
yields of 82, 78, and 70%, respectively (Table 1).
The use of reactions under microwave irradiation for obtaining
the glycosylated diketopiperazines 4 and 5 was also investigated
with the aim to get them in shorter periods of time and in higher
yields, as described for the non-glycosylated DKPs. Nevertheless,
the employment of the same microwave conditions utilized for
cyclization of dipeptides 11–13 led to the degradation of the re-
action mixtures containing the glycodipeptides 19 or 20 in 20%
piperidine/DMF. This may be explained by the lability of the gly-
cosidic bond under harsh conditions, meaning that milder micro-
wave reaction settings have to be further investigated for the
synthesis of glycosylated diketopiperazines.
d
d
signals of aromatic hydrogens (Fmoc, OBn and the side chain of
phenylalanine) and signals of methylenic hydrogens of phenylala-
nine at
d 3.11–2.94 (19), and at d 3.15 and d 2.95 (20).
Finally, the NFmoc deprotection of the glycodipeptides 19 and 20
by treatment with 20% piperidine in DMF, as described for com-
pounds 1–3, led to the formation of the diketopiperazinic ring by
intramolecular cyclization (Scheme 3). However, the cyclizationwas
not so straightforward, as judged by TLC, requiring longer reaction
times, 41 h for the
a isomer and 68 h for the b isomer. The final
glycosylated diketopiperazines 4 and 5 were obtained in reasonable
yields, 35 and 43%, respectively. Presumably, owing to the influence
of the bulky glycosidic unit in the cyclization reaction, the yields of 4
and 5 were slightly lower compared to the non-glycosylated analog
2 (53%). Although direct glycosylation of 3 could be an alternative
approach to get higher yields, diketopiperazine peptide bonds could
not resist to the harsh glycosylation conditions.
These final compounds were suitably characterized by means of
1H NMR and ESI-MS analysis, which demonstrated, respectively,
the lack of the aromatic and aliphatic hydrogens of NFmoc and OBn,
and the presence of the characteristic adduct [MþNa]þ 586.20.
3. Conclusion
In summary, we developed an alternative strategy for the
synthesis of diketopiperazines by exploiting one-pot deprotection–
cyclization reactions in solution-phase. Such straightforward
approach proved to be suitable for both –OH and O-glycosylated
amino acids, suggesting that this method can be effective for
substituents of distinct nature. Furthermore, varying the amino
acid units with properly protected side chain functionalities may
provide DKPs containing sites for additional modification by
conventional orthogonal chemistry.
2.4. Microwave-assisted cyclization reactions
An innovative approach employing microwave-assisted heating
in peptide chemistry provided intermolecular coupling resulting in
dimerization to symmetric diketopiperazines in higher yields
(about 90%) and shorter reaction times than conventional thermal
heating.7 Other studies about the use of reactions under microwave
irradiation for the synthesis of diketopiperazines derived from di-
peptide esters also verified higher yields (97%) compared to clas-
sical thermal heating.9 Therefore, based on the described
advantages of reactions under microwave irradiation for synthesis
of diketopiperazines, we investigated the cyclization of the corre-
sponding protected dipeptides into DKPs 1, 2, and 3 using this
approach.7
4. Experimental
4.1. General
All chemicals were purchased as reagent grade and used with-
out further purification. Solvents were dried according to standard

V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
5347
methods.30 Column chromatography was performed on silica gel
60 (0.040–0.063 mm).
Microwave reactions were performed in an open vessel labo-
ratorial oven (CEM DISCOVER). Nuclear magnetic resonance spectra
were recorded on Bruker Advance DRX 300 (300 MHz), DPX 400
(400 MHz) or DPX 500 (500 MHz) spectrometers. Chemical shifts
29.3 (
b
-CH2 Pro), 24.5 (
g
-CH2 Pro). ESI-HRMS: calculated for
C30H30N2O6Na [MþNa]þ 537.1996, found 537.2039.
4.2.3.2. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-L-phenylalanyl-L-ser-
ine benzyl ester (12). Following the procedure described in Section
4.2.1, the coupling reaction of compound 7 (60 mg, 0.30 mmol),
PyBOP (203 mg, 0.39 mmol), HOBt (53 mg, 0.39 mmol), and N-
(d) are given in parts per million downfield from tetramethylsilane.
Assignments were made with the aid of HMQC and COSY experi-
ments. Optical rotations were measured at ambient temperature on
a Jasco DIP-370 digital polarimeter using a sodium lamp. Accurate
mass electrospray ionization mass spectra (ESI-HRMS) were
obtained using positive ionization mode on a Bruker Daltonics
UltrOTOF-Q-ESI-TOF mass spectrometer.
(fluoren-9-ylmethoxycarbonyl)-
mmol) for 15 h afforded the product 12 as an amorphous solid
(156 mg, 0.27 mmol, 89%) after purification. Rf 0.64 [EtOAc]. [a]
D
þ1.45 (c 1.0, CHCl3). dH (CDCl3, 400 MHz) 7.63 (2H, d, J 7.6 Hz, CH
Fmoc arom.), 7.38 (2H, t, J 6.8 Hz, CH Fmoc arom.), 7.26–7.04 (14H, m,
CH Bn arom., CH Phe arom., CH Fmoc arom.), 6.90 (1H, d, NH Fmoc),
6.20 (1H, d, NH Ser), 5.05 (2H, AB, JAB 12.2 Hz, CH2 Bn), 4.50 (1H, m,
L-phenylalanine 9 (151 mg, 0.39
25
4.2. Synthesis of diketopiperazines 1, 2, and 3
a-CH Ser), 4.31 (1H, m, a-CH Phe), 4.23–4.09 (2H, m, CH2 Fmoc), 4.0
(1H, t, J 6.8 Hz, CH Fmoc), 3.81 (1H, dd, J1 3.8 Hz, J2 11.6 Hz, CH2a Ser),
3.69 (1H, m, CH2b Ser), 2.98 (1H, dd, J1 5.8 Hz, J2 13.5 Hz, CH2a Phe),
2.80 (1H, dd, J1 8.4 Hz, J2 14 Hz, CH2b Phe). dC (CDCl3, 400 MHz) 172.1,
169.3 (CO Ser, CO Phe), 156.1 (CO Fmoc), 143.9, 141.3 (Cquat. Fmoc),
135.2 (Cquat. Bn), 129.1–127.5 (CH Fmoc arom., CH Phe arom., CH Bn
arom.),124.9 (CH Fmoc arom.),119.7 (CH Fmoc arom.), 67 (CH2 Fmoc,
4.2.1. General procedure for NFmoc deprotection and coupling
reactions
The C-terminal amino acid N-(9-fluorenylmethoxycarbonyl)–
serine benzyl ester 6 (prepared from commercially available amino
L-
acid Fmoc–L-serine by treatment with cesium carbonate and benzyl
bromide in DMF)31 was treated with 20% piperidine/DMF (6 equiv)
and stirred for 20 min at room temperature. After concentration in
vacuo, the residue was purified by column chromatography (EtOAc/
CH2 Bn), 62.1 (CH2 Ser), 55.9 (a-CH Phe), 54.8 (a-CH Ser), 46.8 (CH
Fmoc), 38.1 (CH2 Phe). ESI-HRMS: calculated for C34H32N2O6Na
[MþNa]þ 587.2153, found 587.2205.
hexane 7:3 v/v; MeOH/DCM 1:9 v/v). To the obtained product L-
serine benzyl ester 7, PyBOP (1.25 equiv), HOBt (1.25 equiv), and the
Fmoc-protected N-terminal amino acid (1.25 equiv) were added
and dissolved in acetonitrile. The reaction started with the addition
of DIEA (2.5 equiv) was allowed to stir at room temperature for
periods varying from 15 to 18 h. The resulting mixture was dis-
solved in EtOAc, washed with dilute HCl solution, saturated aque-
ous Na2CO3 solution and brine, dried over MgSO4, and concentrated
in vacuo. Purification was carried out by column chromatography
(EtOAc/hexane 7:3, v/v).
4.2.3.3. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-D-phenylalanyl-L-serine
benzyl ester (13). Following the procedure described in Section 4.2.1,
compound 7 (70 mg, 0.35 mmol), PyBOP (224 mg, 0.43 mmol), HOBt
(58 mg, 0.43 mmol), and N-(fluoren-9-ylmethoxycarbonyl)-D-phe-
nylalanine 10 (170 mg, 0.43 mmol) reacted for 15 h, being isolated the
product 13 as an amorphous solid (172 mg, 0.30 mmol, 85%). Rf 0.64
25
[EtOAc]. [
a
]
þ14.7 (c 1.0, CHCl3). dH (CDCl3, 400 MHz) 7.75 (2H, d, J
D
7.6 Hz, CH Fmoc arom.), 7.51 (2H, t, J 6.8 Hz, CH Fmoc arom.), 7.40–7.21
(14H, m, CH Bn arom., CH Phe arom., CH Fmoc arom.), 6.80 (1H, d, NH
Phe), 5.56 (1H, d, NH Ser), 5.13 (2H, s, CH2 Bn), 4.61–4.27 (4H, m,
a-CH
4.2.2. General procedure for cyclization
Ser, -CH Phe, CH2 Fmoc), 4.15 (1H, t, J 6.8 Hz, CH Fmoc), 3.83 (1H, dd, J1
a
Method A. The protected dipeptide was treated with 20% pi-
peridine/DMF (6 equiv) and allowed to stir at room temperature for
periods varying from 15 to 18 h. After concentration in vacuo, the
residue was purified by column chromatography (EtOAc/hexane
1:1 v/v, MeOH/DCM 1:9 v/v).
3.8 Hz, J2 11.6 Hz, CH2a Ser), 3.65 (1H, m, CH2b Ser), 3.07 (2H, m, CH2
Phe). dC (CDCl3, 100 MHz) 172.1, 169.3 (CO Ser, CO Phe), 156.1 (CO
Fmoc), 143.9, 141.3 (Cquat. Fmoc), 135.2 (Cquat. Bn), 129.1–127.5 (CH
Fmoc arom., CH Phe arom., CH Bn arom.),124.6 (CH Fmoc arom.), 119.7
(CH Fmoc arom.), 67.5 (CH2 Bn), 67.2 (CH2 Fmoc), 62.4 (CH2 Ser), 56.4
Method B. The solution of the protected dipeptide in 20% pi-
peridine/DMF (6 equiv) was reacted for 4 min in an open vessel
microwave oven under 200 W and at 163 ꢀC as set temperature. The
product was purified as described in Method A.
(a-CH Phe), 54.6 (a-CH Ser), 47.2 (CH Fmoc), 38.8 (CH2 Phe). ESI-HRMS:
calculated for C34H32N2O6Na [MþNa]þ 587.2153, found 587.2211.
4.2.4. c(L-Prolyl-L-seryl) (1)
Method A. Compound 11 (55 mg, 0.11 mmol) was reacted for
15 h as described in the general procedure (Section 4.2.2). The pure
product 1 was obtained in 68% yield (13 mg, 0.072 mmol) as an
amorphous solid.
4.2.3. Specific procedures
4.2.3.1. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-L-prolyl-L-serine ben-
zyl ester (11). Following the procedure described in Section 4.2.1, the
coupling reaction of compound 7 (44 mg, 0.23 mmol), PyBOP (146
mg, 0.28 mmol), HOBt (38 mg, 0.28 mmol), and N-(fluoren-9-
Method B. Compound 11 (30 mg, 0.058 mol) was reacted as de-
scribed in the general procedure (Section 4.2.2). The product 1 was
obtained in 82% yield (8.7 mg, 0.047 mmol) after purification.
25
ylmethoxycarbonyl)-L-proline 8 (95 mg, 0.28 mol) was carried out
Rf 0.04 [EtOAc]. [
a
]
ꢁ84.5 (c 0.94, MeOH). dH (CD3OD,
-CH Pro), 4.08–4.05 (1H, m, -CH Ser),
3.86–3.76 (2H, m, CH2 Ser), 3.55–3.46 (1H, m, -CH2 Pro), 3.43–3.35
(1H, m, -CH2 Pro), 2.25–2.19 (1H, m, -CH2a Pro), 1.96–1.78 (3H, m,
-CH2b Pro, -CH2 Pro). dC (CD3OD, 100 MHz) 170.5, 165 (CO Ser, CO
Pro), 60.3 (CH2 Ser), 58.8 ( -CH Pro), 56.9 ( -CH Ser), 44.8 ( -CH2
D
for 18 h. The pure product 11 was obtained as an amorphous solid
300 MHz) 4.17–4.11 (1H, m,
a
a
25
(93 mg, 0.18 mmol, 80%). Rf 0.40 [EtOAc]. [
a
]
ꢁ29.2 (c 1.0, CHCl3).
d
D
dH (CDCl3, 400 MHz) 7.65 (2H, d, J 7.3 Hz, CH Fmoc arom.), 7.49 (2H,
dd, J 7.9 Hz, CH Fmoc arom.), 7.32–7.20 (9H, m, CH Bn arom., CH
Fmoc arom.), 7.10 (1H, d, J 7.3 Hz, NH Ser), 5.09 (2H, s, CH2Bn), 4.56
d
b
b
g
a
a
d
(1H, m,
CH Pro, CH Fmoc), 3.91 (1H, dd, J1 3.4 Hz, J2 11.5 Hz, CH2a Ser), 3.85
(1H, dd, J1 3.4 Hz, J2 11.3 Hz, CH2b Ser), 3.53–3.37 (2H, m, -CH2 Pro),
2.06–1.79 (4H, m, -CH2 Pro, -CH2 Pro). dC (CDCl3, 100 MHz) 172.1,
a
-CH Ser), 4.31–4.21 (2H, m, CH2 Fmoc), 4.19–4.12 (2H, m,
a
-
Pro), 28.2 (b-CH2 Pro), 21.8 (g-CH2 Pro). ESI-HRMS: calculated for
C8H12N2O3Na$[MþNa]þ 207.0740, found 207.0746.
d
b
g
4.2.5. c(L-Phenylalanyl-L-seryl) (2)
170.1 (CO Ser, CO Pro), 156.1 (CO Fmoc), 143.9, 141.3 (Cquat. Fmoc),
135.2 (Cquat. Bn), 128.5, 128.1, 127.7, 127.0, 125.1, 119.9 (CH Fmoc
arom., CH Bn arom.), 67.8 (CH2 Fmoc), 67.4 (CH2 Bn), 62.4 (CH2 Ser),
Method A. Compound 12 (50 mg, 0.080 mmol) was reacted for
18 h as described in the general procedure (Section 4.2.2). The pure
product 2 was obtained in 57% yield (12 mg, 0.050 mmol) as an
amorphous solid.
60.7 (a-CH Pro), 55.2 (a-CH Ser), 47.2 (CH Fmoc), 47.1 (d-CH2 Pro),

5348
V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
Method B. Compound 12 (25 mg, 0.044 mmol) was reacted as
described in the general procedure (Section 4.2.2). The product 2
was obtained in 78% yield (8.0 mg, 0.034 mmol) after purification.
7.64 (2H, d, J 7.8 Hz, CH Fmoc arom.), 7.40–7.26 (9H, m, CH Fmoc
arom., CH Bn arom.), 5.81 (1H, d, J 8.7 Hz, NH Ser), 5.35 (1H, d, J
8.4 Hz, NHAc), 5.25 (1H, t, J3,4 9.6 Hz, H-3), 5.20 (2H, AB, JAB 12.2 Hz,
CH2Bn), 5.02 (1H, t, J3,4 10 Hz, H-4), 4.67 (1H, d, J1,2 8.4 Hz, H-1),
4.55–4.38 (3H, m, CH2 Fmoc, CH Ser), 4.28–4.19 (3H, m, H-6a, CH
Fmoc, CH2a Ser), 4.08 (1H, dd, J5,6b 2.2 Hz, J6a,6b 12.3 Hz, H-6b), 3.85
(1H, dd, J 2.7,10.8 Hz, CH2b Ser), 3.70 (1H, apparent t, J1,2, J2,3 10.0 Hz,
H-2), 3.64–3.59 (1H, m, H-5), 2.04, 2.03, 2.02, 1.82 (12H, 4s, COCH3).
dC (CDCl3, 100 MHz) 170.8, 170.7, 169.5, 169.4 (COCH3, COCH2Bn),
156.1 (CO Fmoc),143.7; 143.6; 141.3 (Cquat. Fmoc),135.3 (Cquat. Bn),
128.5, 128.4, 128.2, 127.7, 127.1, 125.1, 119.9 (CH Fmoc arom., CH Bn
arom.), 100.6 (C-1), 71.9, 71.7, (C-5, C-3), 68.9; 68.3 (C-4, CH2 Ser),
67.3 (CH2Bn), 66.7 (CH2 Fmoc), 61.8 (C-6), 54.6; 54.1 (C-2, CH Ser),
47.0 (CH Fmoc), 22.8–20.7 (COCH3). ESI-HRMS: calculated for
C39H42N2O13NH4 [MþNH4]þ 764.3031, found 764.3025.
25
Rf 0.15 [MeOH/DCM: 1:9 v/v]. [
a]
ꢁ124.6 (c 0.36, MeOH). dH
D
(DMSO, 400 MHz) 7.30–7.15 (5H, m, CH arom.), 5.00 (1H, t, J 5.7 Hz,
OH), 4.05 (1H, m, CH Phe), 3.66 (1H, m, CH Ser), 3.31–3.26 (1H, m,
CH2a Ser), 3.10 (1H, dd, J1 6.0 Hz, J2 13.5 Hz CH2a Phe), 2.98 (1H, dd, J1
4.6 Hz, J2 13.5 Hz CH2b Phe), 2.83 (1H, m, dd, J17.3 Hz, J2 11.4 Hz CH2b
Ser). dC (DMSO, 100 MHz) 129.8–126.3 (CH arom.), 62.9 (CH2 Ser),
56.9 (a-CH Ser), 55.2 (a-CH Phe), 39.6 (CH2 Phe). ESI-HRMS: cal-
culated for C12H15N2O3$[MþH]þ 235.1004, found 235.1083.
4.2.6. c(D-Phenylalanyl-L-seryl) (3)
Method A. Compound 13 (41 mg, 0.070 mmol) was reacted for
18 h as described in the general procedure (Section 4.2.2). The pure
product 3 was obtained in 53% yield (9.1 mg, 0.040 mmol) as an
amorphous solid.
Method B. Compound 13 (60 mg, 0.10 mol) was reacted as de-
scribed in the general procedure (Section 4.2.2). The product 3 was
4.3.2. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-
O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-
-serine benzyl ester (19)
Compound 14 (61 mg, 0.082 mmol) was treated with 20% pi-
D-phenylalanyl-
a-D-glucopyranosyl)-
L
obtained in 70% yield (17 mg, 0.074 mmol) after purification.
25
Rf 0.15 [MeOH/DCM 1:9 v/v]. [
a
]
D
ꢁ117.4 (c 0.35, MeOH). dH
peridine/DMF (0.5 mL) for removal of the NFmoc group. The re-
action mixture was stirred for 1 h at room temperature and then
purified by column chromatography (EtOAc/hexane, 7:3 v/v;
MeOH/DCM 1:9 v/v). The product (2-acetamido-2-deoxy-3,4,6-tri-
(DMSO, 400 MHz) 7.24–7.06 (5H, m, CH arom.), 5.00 (1H, sl, OH),
4.15 (1H, m, CH Phe), 3.59 (1H, dd, J1 3.0 Hz, J2 11.6 Hz, CH2a Ser),
3.42 (1H, dd, J1 2.8 Hz, J2 11.6 Hz, CH2b Ser), 3.15 (1H, dd, J1 3.8 Hz, J2
13.6 Hz, CH2a Phe), 3.06 (1H, m, CH Ser), 2.87 (1H, dd, J1 4.8 Hz, J2
13.6 Hz, CH2b Phe). dC (DMSO, 100 MHz) 129.9–126.4 (CH arom.),
O-acetyl-b-D-glucopyranosyl)-L-serine benzyl ester 17 was
obtained as a pale oil (32 mg, 0.061 mmol, 75%). PyBOP (37 mg,
0.071 mmol), HOBt (9.6 mg, 0.071 mmol), and compound 10
(27 mg, 0.070 mmol) were then added to 14 (30 mg, 0.057 mmol)
and dissolved in DMF (0.8 mL), being the reaction initiated with
the addition of DIEA (0.05 mL, 0.28 mmol). The mixture was
allowed to stir for 20 h at room temperature and purification by
column chromatography (EtOAc/hexane, 7:3 v/v) afforded the
62.7 (CH2 Ser), 56.3 (a-CH Ser), 54.9 (a-CH Phe), 37.6 (CH2 Phe). ESI-
HRMS: calculated for C12H15N2O3$[MþH]þ 235.1004, found
235.1083.
4.3. Synthesis of glycosylated diketopiperazines 4 and 5
4.3.1. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-O-(2-acetamido-
product 19 as an amorphous solid (38 mg, 0.043 mmol, 75%). Rf
25
3,4,6-tri-O-acetyl-2-deoxy-
ester (14) and N-[(9H-fluoren-9-ylmethoxy)carbonyl]-O-
(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy- -glucopyranosyl)-
-serine benzyl ester (15)29
A mixture of 2-acetamido-2-deoxy-3,4,6-tri-O-acetyl-
copyranosyl chloride 16 (300 mg, 0.82 mmol) and N-(9-fluo-
renylmethoxycarbonyl)- -serine benzyl ester 6 (171 mg, 0.41 mmol)
a-D
-glucopyranosyl)-
L
-serine benzyl
0.36 [EtOAc/hexane, 7:3 v/v]. [
a
]
ꢁ5.0 (c 0.36, CHCl3). dH (CDCl3,
D
300 MHz) 7.68 (2H, d, J 7.7 Hz, CH Fmoc arom.), 7.45–7.41 (2H, m,
CH Fmoc arom.), 7.35–7.21 (14H, m, CH Fmoc arom., CH Phe arom.,
CH Bn arom.,), 5.84 (1H, d, J 8.8 Hz, NH Ser), 5.33 (1H, d, J 7.4 Hz,
NH Phe), 5.07 (2H, s, CH2 Bn), 5.01–4.96 (2H, m, H-3, H-4), 4.70
b-D
L
a-D-glu-
(1H, d, J 3.7 Hz, H-1), 4.44–4.34 (3H, m, H-6a, H-6b,
a-CH Phe),
L
4.20–3.94 (5H, m, H-5, -CH Ser, CH Fmoc, CH2 Fmoc), 3.84 (1H, dd,
a
in 1,2-dichloroethane (4.3 mL) was refluxed with mercuric bromide
(400 mg, 0.82 mmol) for 9 h and followed by TLC (EtOAc/hexane 7:3,
v/v). The resulting amber mixture was concentrated in vacuo and
the residue was purified by a silica gel column chromatography
(EtOAc/hexane 7:3, v/v). The products 14 (61 mg, 0.081 mmol, 20%)
and 15 (122 mg, 0.16 mmol, 40%) were obtained as amorphous
J1,2 3.3 Hz, J2,3 10.2 Hz, H-2), 3.74–3.66 (1H, m, CH2a Ser), 3.64–3.55
(1H, m, CH2b Ser), 3.11–2.94 (2H, m, CH2 Phe), 1.98, 1.97, 1.93, 1.92
(12H, 4s, COCH3). dC (CDCl3, 100 MHz) 129.2–127.1 (CH Fmoc arom.,
CH Phe arom., CH Bn arom.), 125.0, 119.8 (CH Fmoc arom.), 98.78
(C-1), 70.73 (C-3), 69.56 (C-2), 68.2 (CH2 Ser), 67.9 (C-4), 67.74 (CH2
Bn), 67.23 (C-6), 67.12 (C-5), 61.5 (CH2 Fmoc), 56.30 (a-CH Phe),
solids.
51.8 (a-CH Ser), 47.0 (CH Fmoc), 38.37 (CH2 Phe), 23.03–20.54
25
a-Glycoside 14. Rf 0.31 (EtOAc/hexane, 7:3 v/v). [
a
]
þ14.8 (c 1.0,
(COCH3). ESI-HRMS: calculated for C48H52N3O14 [MþH]þ 894.3443,
D
CHCl3). dH (CDCl3, 400 MHz) 7.66 (2H, d, J 7.5 Hz, CH Fmoc arom.),
7.62 (2H, d, J 6.0 Hz, CH Fmoc arom.), 7.45–7.26 (9H, m, CH Fmoc
arom., CH Bn arom.), 5.95 (1H, d, J 9.3 Hz, NHAc), 5.85 (1H, d, J
9.5 Hz, NH Ser), 5.24–5.16 (3H, m, AB, JAB 12.2 Hz, CH2Bn, H-3), 5.08
(1H, t, J3,4 9.4 Hz, H-4), 4.70 (1H, d, J1,2 3.6 Hz, H-1), 4.60 (1H, m, CH
Ser), 4.45 (2H, m, CH2 Fmoc), 4.30 (1H, dt, J1,2 3.6 Hz, J2,3 9.4 Hz, H-
2), 4.25 (1H, t, J 7.3 Hz, CH Fmoc), 4.20–4.06 (3H, m, CH2a Ser, H-6a,
H-6b), 3.88 (2H, m, CH2bSer), 3.84 (1H, m, H-5), 2.06, 2.05, 2.03, 2.02
(12H, 4s, COCH3). dC (CDCl3, 100 MHz) 171.5, 170.9, 170.8, 170.6
(COCH3), 169.5 (COCH2Bn), 156.7 (CO Fmoc), 143.6, 141.3 (Cquat.
Fmoc), 134.5 (Cquat. Bn), 129.2, 129.1, 128.8, 128.0, 127.3, 125.3,
120.2 (CH Fmoc arom., CH Bn arom.), 99.5 (C-1), 71.4 (C-4), 68.5,
68.4 (C-3, C-5), 68.0 (CH2Bn), 67.9 (CH2b Ser), 67.7 (CH2 Fmoc), 62.3
(CH2a Ser), 62.2 (C-6), 55.0 (CH Ser), 52.5 (C-2), 47.3 (CH Fmoc),
23.3–20.8 (COCH3). ESI-HRMS: calculated for C39H42N2O13Na
found 894.3406.
4.3.3. N-[(9H-Fluoren-9-ylmethoxy)carbonyl]-
O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-
-serine benzyl ester (20)
The NFmoc deprotection of the building block 15 was first re-
D-phenylalanyl-
b-D-glucopyranosyl)-
L
alized by treatment with 20% piperidine/DMF (0.3 mL). Starting
from 50 mg (0.067 mmol) of 15, the product (2-acetamido-2-de-
oxy-3,4,6-tri-O-acetyl-b-D-glucopyranosyl)-L-serine benzyl ester 18
was obtained in 85% yield (29.7 mg, 0.057 mmol) as an oil. Then,
following the procedure described for compound 17, from 28 mg
(0.053 mmol) of 18, PyBOP (37.2 mg, 0.071 mmol), HOBt (9.6 mg,
0.071 mmol), compound 10 (27.6 mg, 0.071 mmol), and DIEA
(0.05 mL, 0.28 mmol), it was possible to obtain 32.5 mg
(0.036 mmol, 68%) of the product 20. Rf 0.37 [EtOAc/hexane, 7:3 v/
25
[MþNa]þ 764.2579, found 764.2593.
v]. [
a]
ꢁ13.0 (c 0.4, CHCl3). dH (CDCl3, 300 MHz) 7.69 (2H, d, J
D
25
b-Glycoside 15. Rf 0.25 (EtOAc/hexane, 7:3 v/v). [
a
]
ꢁ4.6 (c 1.0,
7.3 Hz, CH Fmoc arom.), 7.45–7.14 (16H, m, CH Fmoc arom., CH Bn
arom., CH Phe arom.), 5.44 (1H, d, J 7.7 Hz, NH Phe), 5.22 (1H, t, J3,4
D
CHCl3). dH (CDCl3, 400 MHz) 7.77 (2H, d, J 7.8 Hz, CH Fmoc arom.),

V.L. Campo et al. / Tetrahedron 65 (2009) 5343–5349
5349
9.9 Hz, H-3), 5.10 (2H, AB, JAB 12.3 Hz, CH2 Bn), 4.91 (1H, t, J3,4
9.6 Hz, H-4), 4.62 (1H, d, J1,2 7.9 Hz, H-1), 4,40 (2H, m, CH Fmoc,
CH Phe), 4.18–3.98 (6H, m, H-5, H-6a, H-6b, -CH Ser, CH2 Fmoc),
3.77 (1H, dd, J 9.4, 1.5 Hz, CH2a Ser), 3.53 (1H, dd, J 9.4, 1.5 Hz, CH2b
Ser), 3.39 (1H, dd, J1,2 8.0 Hz, J2,3 10.2 Hz, H-2), 3,15 (1H, dd, J 13.1,
5.3 Hz, CH2a Phe), 2.95 (1H, dd, J 13.1, 7.6 Hz, CH2b Phe), 1.98, 1.97,
1.92, 1.81 (12H, 4s, COCH3). dC (CDCl3, 100 MHz) 129.5–127.9 (CH
Fmoc arom., CH Phe arom., CH Bn arom.), 125.1, 120.3 (CH Fmoc
arom.),100.9 (C-1), 71.9 (CH2 Ser), 71.93 (C-3), 68.53 (C-5), 68.46 (C-
4), 67.74 (CH2 Bn), 67.35 (CH Fmoc), 62.08 (CH2 Fmoc), 61.86 (C-6),
conformer, which was immersed in five layers of DMSO molecules.
The overall system was then energy minimized using the CVFF
force field thus implemented in the DISCOVER3 module of the In-
sight II package, initially maintaining fixed compound 2 and sub-
sequently relaxing all the solute–solvent system. From this
resulting optimized system and for the sake of calculation limits,
only the first layer of DMSO was energy minimized together with
compound 2 using the semi-empirical AM1 model. The molecule
a
-
a
was then fully optimized in gas phase at B3LYP/6-31G level of
*
calculation. All the quantum chemical methods here used are
56.71 (
a
-CH Phe), 55.44 (C-2), 47.22 (
a
-CH Ser), 38.37 (CH2 Phe),
implemented in the Spartan ’06 1.1.2 software.
23.73–20.5 (COCH3). ESI-HRMS: calculated for C48H51N3O14Na
[MþNa]þ 916.3263, found 916.3339.
Acknowledgements
4.3.4. c(
O-acetyl-
D
-Phenylalanyl-O-(2-acetamido-2-deoxy-3,4,6-tri-
`
Authors are thankful to Fundaça˜o de Amparo a Pesquisa do
a
-
D
-glucopyranosyl)- -seryl) (4)
L
Estado de Sa˜o Paulo (FAPESP) for financial support and fellowships.
Compound 19 (30 mg, 0.034 mmol) was treated with 20% pi-
peridine/DMF (0.5 mL) and the reaction mixture was stirred for
41 h, being followed by TLC (EtOAc/hexane, 7:3 v/v). The mixture
was then concentrated in vacuo and the residue was purified by
column chromatography (EtOAc/hexane 7:3 v/v; MeOH/CH2Cl2 1:9
v/v). The product 4 was obtained as an amorphous solid (6.7 mg,
0.012 mmol, 35%). Rf 0.15 [EtOAc/hexane 7:3 v/v]. [
0.4, MeOH). dH (CDCl3, 300 MHz) 7.33–7.22 (5H, m, CH Phe arom.),
6.68 (1H, d, J 9.3 Hz, NH Phe), 5.15–5.01 (2H, m, H-3, H-4), 4.77 (1H,
Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2009.04.069.
25
a
]
ꢁ186.9 (c
D
References and notes
1. Gellerman, G.; Hazan, E.; Brider, T.; Traube, T.; Albeck, A.; Shatzmiler, S. Int. J.
Pept. Res. Ther. 2008, 14, 183.
2. Martins, M. B.; Carvalho, I. Tetrahedron 2007, 63, 9923.
3. Tullberg, M.; Grøtli, M.; Luthman, K. J. Org. Chem. 2007, 72, 195.
4. Rodionov, I. L.; Rodionova, L. N.; Baidakova, L. K.; Romashko, A. M.; Balashova,
T. A.; Ivanov, V. T. Tetrahedron 2002, 58, 8515.
5. Horton, D. A.; Bourne, G. T.; Smythe, M. L. Mol. Diversity 2000, 5, 289.
6. Teixido´, M.; Zurita, E.; Malakoutikhah, M.; Tarrago´ , T.; Giralt, E. J. Am. Chem. Soc.
2007, 129, 11802.
7. Santagada, V.; Fiorino, F.; Perissutti, E.; Severino, B.; Terraciano, S.; Cirino, G.;
Caliendo, G. Tetrahedron Lett. 2003, 44, 1145.
d, J 3.7 Hz, H-1), 4.32–4.27 (2H, m, H-2,
a-CH Phe), 4.18–3.95 (3H, m,
H-6a, H-6b, -CH Ser), 3.85–3.79 (2H, m, CH2a Ser), 3.58–3.50 (2H,
a
m, H-5, CH2b Ser), 3.24 (1H, dd, J 13.7, 3.7 Hz, CH2a Phe), 3.02 (1H,
dd, J 13.7, 7.5 Hz, CH2b Phe), 2.03, 1.96, 1.93, 1.84 (12H, 4s, COCH3). dC
(CDCl3, 75 MHz) 170.9, 169.3, 166.6 (CO Ser, CO Phe, COCH3), 129.9–
128.0 (CH arom.), 98 (C-1), 70.7 (C-3), 68.4 (C-5), 68.3 (
68.15 (CH2 Ser), 68 (C-4), 61.7 (C-6), 56.1 ( -CH Phe), 51.5 (C-2), 40.1
a-CH Ser),
a
(b-CH2 Phe), 22.9–20.5 (COCH3). ESI-HRMS: calculated for
C26H33N3O11Na [MþNaþ] 586.2013, found 586.2028.
8. Giraud, M.; Bernad, N.; Martinez, J.; Cavelier, F. Tetrahedron Lett. 2001,
42, 1895.
9. Tullberg, M.; Grøtli, M.; Luthman, K. Tetrahedron 2006, 62, 7484.
10. Fischer, P. M. J. Pept. Sci. 2003, 9, 9.
11. Santra, S.; Andreana, P. Org. Lett. 2008, 9, 5035.
12. Marcaccini, S.; Torroba, T. Nat. Protocols 2007, 2, 632.
13. Pichowicz, M.; Simpkins, N. S.; Blake, A. J.; Wilson, C. Tetrahedron 2008,
64, 3713.
14. Bull, S. D.; Davies, S. G.; Garner, A. C.; Parkers, A. L.; Roberts, P. M.; Sellers, T. G.
R.; Smith, A. D.; Tamayo, J. A.; Thomson, J. E.; Vickers, R. J. New J. Chem. 2007, 31,
486.
15. O’Neill, J. C.; Blackwell, H. E. Comb. Chem. High Throughput Screening
2007, 10, 857.
16. Santos, C.; Mateus, M. L.; Santos, A. P.; Moreira, R.; Oliveira, E.; Gomes, P. Bioorg.
Med. Chem. Lett. 2005, 15, 1595.
17. Joshi, K. B.; Verma, S. Tetrahedron Lett. 2008, 49, 4231.
18. Kilian, G.; Jamie, H.; Brauns, S. C. A.; Dyason, K.; Milne, P. J. Pharmazie
2005, 60, 305.
19. Isaka, M.; Palasarn, S.; Rachtawee, P.; Vimuttipong, S.; Kongsaeree, P. Org. Lett.
2005, 7, 2257.
20. Chan, W. C.; White, P. D. Fmoc Solid Phase Peptide Synthesis: A Practical Approach;
Oxford University Press: New York, NY, 2000.
4.3.5. c(
O-acetyl-
D
-Phenylalanyl-O-(2-acetamido-2-deoxy-3,4,6-tri-
b
-D
-glucopyranosyl)- -seryl) (5)
L
According to the procedure described for the synthesis of the
product 4, compound 20 (30 mg, 0.033 mmol) was treated with
20% piperidine/DMF (0.5 mL) and the reaction mixture was stirred
for 68 h, being followed by TLC analysis (EtOAc/hexane, 7:3 v/v).
The mixture was then concentrated in vacuo and the residue was
purified by column chromatography (EtOAc/hexane 7:3 v/v;
MeOH/CH2Cl2 1:9 v/v). The product 5 was obtained as an amor-
phous solid (8.1 mg, 0.014 mmol, 43%). Rf 0.13 [EtOAc:hexane 7:3
25
v/v]. [
a
]
ꢁ192.1 (c 0.4, MeOH). dH (CDCl3, 300 MHz) 7.27–7.20 (5H,
D
m, CH Phe arom.), 6.64 (1H, d, NH Phe), 6.41 (1H, d, J 9.0 Hz, NHAc),
5.09–4.94 (2H, m, H-3, H-4), 4.49 (1H, d, J 8.5 Hz, H-1), 4.30–4.27
(1H, m, a-CH Phe), 4.22–3.95 (4H, m, H-2, H-6a, H-6b, a-CH Ser),
3.67–3.56 (3H, m, H-5, CH2 Ser), 3.25 (1H, dd, J 13.8, 3.8 Hz, CH2a
Phe), 2.91 (1H, dd, J 13.8, 8.0 Hz, CH2b Phe), 2.01, 1.98, 1.96, 1.93
(12H, 4s, COCH3). dC (CDCl3, 75 MHz) 170.9, 169.3, 166.6 (CO Ser, CO
Phe, COCH3), 129.9–127.5 (CH arom.), 101.4 (C-1), 72.5 (C-3), 71.9
21. Umezawa, Y.; Tsuboyama, S.; Takahashi, H.; Uzawa, J.; Nishio, M. Tetrahedron
1999, 55, 10047.
22. Spartan ’06 full v.1.1.2; Tutorial and User’s Guide; Wavefunction: Irvine, CA, USA,
2006.
23. Hehre, W. J. A Guide to Molecular Mechanics and Quantum Chemical Calculations;
Wavefunction: Irvine, CA, 2003.
24. Leach, A. R. Molecular Modelling: Principles and Applications, 2nd ed.; Pearson:
England, 2001.
(C-5), 70.9 (a-CH Ser), 68.3 (C-4), 61.9 (C-6), 55.5 (CH2 Ser), 55.0 (a-
CH Phe), 53.8 (C-2), 40.9 (b-CH2 Phe), 23.03–20.54 (COCH3). ESI-
HRMS: calculated for C26H33N3O11Na [MþNa]þ 586.2013, found
586.2038.
25. Insight II; Accelrys: San Diego, CA, 2005.
26. Campo, V. L.; Carvalho, I. Quim. Nova 2008, 31, 1027.
27. Zachara, N. E.; Hart, G. W. Biochim. Biophys. Acta 2006, 1761, 599.
28. Campo, V. L.; Carvalho, I.; Allman, S.; Davis, B. G.; Field, R. A. Org. Biomol. Chem.
2007, 5, 2645.
4.4. Molecular modeling procedures
29. Carvalho, I.;Scheuerl,S.L.;Kartha, K.P. R.;Field,R.A.Carbohydr.Res.2003, 338,1039.
30. Perrin, D. D.; Armarego, W. L. F.; Perrin, D. R. Purification of Laboratory Chemicals,
2nd ed.; Pergamon: New York, NY, 1980; EUA.
31. Wang, S. S.; Gisin, B. F.; Winter, D. P.; Makofske, R.; Kulesha, I. D.; Tzougraki, C.;
Meienhofer, J. J. Org. Chem. 1977, 42, 1286.
Conformational search was performed on diketopiperazine 2
structure using the MONTE CARLO method with the MMFF mo-
lecular mechanics model, thus implemented in the Spartan ’06 1.1.2
software. Boundary conditions were applied to the folded