New sialyl Lewisx mimic containing an α-substituted β3-amino acid spacer

Article abstract of DOI:10.1016/j.carres.2007.10.001

A highly convergent and efficient synthesis of a new sialyl Lewis^x (sLe^x) mimic, which was predicted by computational studies to fulfil the spacial requirements for a selectin antagonist, has been developed. With a β^2,3-am

Full text of DOI:10.1016/j.carres.2007.10.001

Available online at www.sciencedirect.com
Carbohydrate Research 343 (2008) 31–38
New sialyl Lewisx mimic containing an a-substituted
b3-amino acid spacer
Silvana Pedatella,a,* Mauro De Nisco,a Beat Ernst,b Annalisa Guaragna,a
Beatrice Wagner,b Robert J. Woodsc and Giovanni Palumboa
a
`
Dipartimento di Chimica Organica e Biochimica, Universita di Napoli Federico II, Via Cynthia, 4 I-80126 Napoli, Italy
bInstitute of Molecular Pharmacy, Pharmacenter, University of Basel, Klingelberstrasse, 50 CH-4056 Basel, Switzerland
cComplex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA
Received 11 June 2007; received in revised form 20 August 2007; accepted 2 October 2007
Available online 7 October 2007
Abstract—A highly convergent and efficient synthesis of a new sialyl Lewisx (sLex) mimic, which was predicted by computational
studies to fulfil the spacial requirements for a selectin antagonist, has been developed. With a b2,3-amino acid residue L-galactose
(bioisostere of the L-fucose moiety present in the natural sLex) and succinate are linked, leading to a mimic of sLex that contains
all the required pharmacophores, namely the 3- and 4-hydroxy group of L-fucose, the 4- and 6-hydroxy group of D-galactose
and the carboxylic acid of N-acetylneuraminic acid. The key step of the synthesis involves a tandem reaction consisting of a N-
deprotection and a suitable O!N intramolecular acyl migration reaction which is promoted by cerium ammonium nitrate
(CAN). Finally, the new sialyl Lewisx mimic was biologically evaluated in a competitive binding assay.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Keywords: sLex; Selectin antagonist; b3-Amino acid; Glycomimetic
1. Introduction
hydrate epitope recognized by E-selectin,4 it became
the lead structure for the design of selectin antagonists.5
The search for novel selectin antagonist with enhanced
adhesion and improved pharmacokinetic properties5 has
led to numerous classes of antagonists. In the initial con-
tributions,6,7 the structure–activity relationship was eluci-
dated, revealing the essential pharmacophores which
are the carboxylic acid function of N-acetylneuraminic
acid, the 3- and 4-hydroxyl group of L-fucose and the
4- and 6-hydroxyl group of D-galactose (highlighted in
Fig. 1).8 In addition, it has been shown that the D-Glc-
NAc moiety is not involved in binding. Its principal func-
tion is that of a rigid spacer to accommodate the
appropriate spacial orientation of L-fucose and D-galact-
ose.5 For the design of simplified sLex antagonists5 a dual
strategy was pursued by (1) eliminating monosaccharide
moieties by non-carbohydrate linkers, and (2) substitut-
ing metabolically labile O-glycosidic bonds.
Selectins1 are involved in the orderly migration of leuco-
cytes from blood vessels to the sites of inflammation.2
Although extravasation of leucocytes represents an
essential defense mechanism against inflammatory stim-
uli, excessive infiltration of leucocytes into the surround-
ing tissue can cause acute or chronic reactions as
observed in reperfusion injuries, stroke, psoriasis, rheu-
matoid arthritis, or respiratory diseases.2 An early step
in this inflammatory cascade is mediated by selectin/carbo-
hydrate interactions. Data from both selectin knock
out mice3a–c and LAD type 2 patients,3d clearly demon-
strate that the selectin–carbohydrate interaction is a pre-
requisite for the inflammatory cascade to take place.
Since the tetrasaccharide sLex (1, Fig. 1) is the carbo-
Considering the glycoaminoacid mimics reported by
Wong,9 which exhibit remarkable pharmacological
*
Corresponding author. Tel.: +39 081 674118; fax: +39 081 674119;
e-mail: pedatell@unina.it
0008-6215/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2007.10.001

32
S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
Figure 1. Identical spacial orientation of the pharmacophores in sLex (1) and the glycoaminoacid mimic 2.
activity, we report herein the synthesis and the biological
evaluation of the scaffold 2, a member of a new family of
antagonists based on a dihydroxylated b-amino acid as a
substitute of galactose. This scaffold is decorated with a
suitably modified L-fucose and a carboxylic acid moiety
mimicking the native N-acetylneuraminic acid. The
corresponding target molecule 2 (Fig. 1) therefore
contains (2S,3S)-2-hydroxy-b3-serine which is coupled
with the free amino group of 6-amino-6-deoxy-L-galact-
ose (L-Fuc replacement) and, at the other end, with a
succinic acid (D-NeuNAc replacement). An important
factor for affinity is the spacial orientation of the phar-
macophores in the bioactive conformation.5d,e This is
expected to be realized with the rigid diamide core.
(3), using CuSO4/H2SO4 in acetone, gave only poor
yields (55%).
Using an improved procedure12 (polystyryl diphenyl
phosphine–iodine complex, prepared in situ, in anhy-
drous acetone, at rt under nitrogen) 3 was obtained in
95% yield. Tosylation of the primary hydroxy group fol-
lowed by substitution with sodium azide and catalytic
hydrogenation (Pd/H2) afforded the free amine 4 in
69% overall yield from L-galactose.
(2S,3S)-2-Hydroxy-b3-serine methyl ester derivative 7,
was obtained diastereoselectively13 from commercially
available O-benzyl-L-serine via enolate formation (by
KHMDS) of the corresponding fully protected b3-serine
5 and the coupling of the enolate with racemic 2-[(4-
methylphenyl)sulfonyl]-3-phenyloxaziridine (6) (Scheme
2).
2. Results and discussion
The hydrolysis of methyl ester 7 afforded 8 in excellent
yield without any traces of racemization.
A conformational analysis of 2 by molecular dynamic
studies indicated that the low energy conformer-2 pre-
sented in Figure 1 exhibits distances between the carbox-
Then, the 2-hydroxy-b3-amino acid 8 was coupled
with the previously prepared 6-amino-L-galactose deriv-
ative 4 by treatment with DCC and HOBT in CH2Cl2,
yielding 9 in good yield (75%) (Scheme 3).
˚
ylate and C-1 and C-2 of L-galactose (9.2 and 9.1 A,
respectively), which are similar to the one found in the
bioactive conformation10 of sLex. Conformer-2 is only
less stable by 5.29 kJ/mol compared to its lowest energy
conformation and is characterized by differing flexibili-
ties of the dihedral angles h, u and x (Fig. 2). Whereas
h and u show considerable flexibility over the 10,000 ps
period of the MD simulation, x is rather stiff, leading to
The last step of the synthesis was the introduction of a
carboxylate side chain at the N-terminus of 9. The first
attempt to remove the 4-methoxybenzyl ether protection
(PMB) with cerium ammonium nitrate (CAN)14 was
accompanied by the formation of several byproducts
due to the cleavage of C-2–C-3 bond of the b-aminoac-
idic moiety. To avoid a possible interference of the free
hydroxyl group with CAN, compound 9 was first acetyl-
ated and then treated with CAN. A single product was
formed, which turned out not to be the desired N-depro-
tected derivative, but the product of a subsequent O!N
intramolecular acetyl migration.15
a
partial preorganization of
2 in the bioactive
conformation.
The synthesis started with the preparation of 6-amino-
6-deoxy-L-galactose derivative 4 from commercially
available L-galactose according to a known procedure11
(Scheme 1). According to this procedure, the acetona-
tion to achieve 1,2:3,4-di-O-isopropyliden-L-galactose
When this O!N intramolecular acyl migration was
applied to ester 10, which was obtained by esterification

S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
33
Figure 2. The plots A, B, and C show the fluctuations of the h, u, and x torsional angles in compound 2 monitored during the 10,000 ps MD
simulation in a box of TIP3P water molecules; A, B: the h and u dihedral angles are highly variable within the MD simulation time frame; C: the x
torsion angle remains practically unchanged during the MD simulation.
HO
O
O
OH
O
O
OH
OH
O
O
i
ii
O
O
O
HO
HO
H2N
ref 11
O
O
3
4
Scheme 1. Synthesis of 6-amino-6-deoxy-L-galactose derivative 4. Reagents and conditions: (i) PDP/I2, anhyd (CH3)2CO, rt, 1 h, 95%; (ii) (a) TsCl,
anhyd py, 0 ꢁC!rt, 45 min, 86%; (b) NaN3, anhyd DMF, 115 ꢁC, 15 h, 94%; (c) H2, 5% Pd/C, MeOH, ultrasound, 90%.
BnO
BnO
H2N
O
O
(PMB)2N
OMe
ref 13
OH
5
O
TolO SN
p
2
Ph
ref 13
6
BnO
BnO
MeOH
1 M aq NaOH
O
O
(PMB)2N
OH
(PMB)2N
OMe
0 ºC
rt, 6.5 h, 92%
OH
OH
7
8
Scheme 2. Synthesis of protected (2S,3S)-2-hydroxy-b3-serine 8.
of 9 with 3-carbomethoxypropionyl chloride, product 11
could be isolated in excellent 90% yield (Scheme 3).
Finally, the removal of the protecting groups by
hydrogenolysis on Pd/C in an ultrasound bath (!12),
followed by alkaline hydrolysis (!13), and acidic
hydrolysis (TFA/H2O) of the isopropylidene groups
(Scheme 3) yielded 2 as an anomeric mixture (a/b = 1:2).
3. Biological evaluation
Various formats of cell-free competitive binding assays
under static conditions have been reported.16–18 We used
microtiter plates coated with human recombinant
E-selectin/IgG or P-selectin/IgG, which were incubated
with 2 and commercially available sLex/biotin-polyacry-

34
S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
O
Cl
(CH2)2COOMe
ii
O
O
BnO
BnO
O
O
O
O
O
i
O
8
O
O
(PMB)2N
N
H
(PMB)2N
O
N
H
O
O
OH
O
9
10
COOMe
iii
O
O
HO
HN
BnO
HN
O
O
O
O
O
O
iv
O
O
N
N
H
H
O
O
OH
OH
O
O
11
12
COOMe
COOMe
v
O
HO
HN
O
O
O
vi
O
2
N
H
O
OH
O
13
COOH
Scheme 3. Synthesis of mimic 2. Reagents and conditions: (i) DCC, HOBT, 4, anhyd CH2Cl2, rt, 15 h, 75%; (ii) anhyd py, CH2Cl2, rt, 4 h, 90%; (iii)
CAN, MeCN/H2O, 0 ꢁC!rt, 19 h, 90%; (iv) H2, Pd/C 5%, MeOH, ultrasound, 40 ꢁC, 22 h, 93%; (v) MeOH, 1 M aq NaOH, 0 ꢁC!rt, 3 h, 91%; (vi)
TFA/H2O, rt, 3 h, 80%.
late. After unbound ligand and polymer were washed
from the plate, streptavidin/horseradish peroxidase con-
jugate was added to enable colorimetric determination
of binding.18b As a reference compound, sialyl Lewisx
with an IC50 value of 0.9 mM for E-selectin and
P3 mM for P-selectin was used. With 2, a moderate
inhibition of the interaction of sLex/E-selectin
(IC50 P 6 mM) could be detected. However, the interac-
tion of P-selectin with the sLex-polyacrylate polymer
could not be inhibited with 2 (see Table 1).
sLex mimic shows only low affinity of E-selectin
(IC50 = 6 mM) and no activity (IC50 >10 mM) for
P-selectin, respectively.
Since the spatially correct arrangement of the phar-
macophores can be realized in 2, the low affinity can also
be due to the considerable conformational flexibility of
the dihedral angles h and u (Fig. 2A and B) leading to
substantial cost of entropy upon binding.
5. Experimental
5.1. General methods
4. Conclusion
The new sLex mimic 2 was designed and synthesized
starting from an L-galactose derivative 4 and orthogo-
nally protected (2S,3S)-2-hydroxy-b3-serine 8 in six
steps. Our synthesis features the use of cerium ammo-
nium nitrate (CAN) to perform a useful O!N intra-
molecular acyl migration reaction on the succinoyl
ester 10. However, biological testing revealed that this
Triphenyl phosphine polymer-bound was purchased
from Fluka Chemical Co. All moisture-sensitive reac-
tions were performed under a nitrogen atmosphere using
oven-dried glassware. Solvents were dried over standard
drying agents and freshly distilled prior to use. Reac-
tions were monitored by TLC (precoated silica gel plate
F254, Merck). Column chromatography: Merck Kiesel-
gel 60 (70–230 mesh); flash chromatography: Merck
Kieselgel 60 (230–400 mesh). Optical rotations were
measured at 25 2 ꢁC in the stated solvent. 1H and
13C NMR spectra were recorded on NMR spectro-
meters operating at 400 or 500 MHz and 50, 100 or
125 MHz, respectively. Wherever necessary, two-dimen-
Table 1. Biological evaluation data
Test compound
E-Selectin (mM)
P-Selectin (mM)
Sialyl Lewisx (1)
0.9
P3
Glycoaminoacid (2)
P6
>10

S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
35
1
sional H–1H COSY experiments were carried out for
complete signal assignments. Combustion analyses were
performed using CHNS analyzer.
5.5. N1-(1,2:3,4-Di-O-isopropylidene-6-deoxy-a-L-galac-
topyranos-6-yl)-(2S,3S)-4-(benzyloxy)-3-[di(4-methoxy-
benzyl)amino]-2-hydroxybutanamide (9)
5.2. 1,2:3,4-Di-O-isopropylidene-a-L-galactopyranose (3)
HOBT (0.35 g, 2.58 mmol) was added to a stirred solu-
tion of b2,3-amino acid 8 (0.60 g, 1.29 mmol) in anhy-
drous CH2Cl2 (10 mL) at rt. After 30 min, to the
resulting mixture a solution of compound 4 (0.30 g,
1.17 mmol) and DCC (0.40 g, 1.42 mmol) in the same
solvent (10 mL) was added dropwise over 5 min at
0 ꢁC. The solution was allowed to warm slowly to rt
and, after 15 h, most of the solvent was evaporated
under reduced pressure and replaced by EtOAc. The
precipitate was filtered and the solution was washed
with saturated NaHCO3 solution, brine until neutral,
then dried (Na2SO4), and concentrated under reduced
pressure. Chromatography of the crude residue on silica
gel (hexane/EtOAc, 1:1) afforded the oily coupling prod-
To a magnetically stirred suspension of dry polystyryl
diphenyl phosphine (1.12 g, ꢀ3.34 phosphine units) in
anhydrous acetone (10 mL) at rt, a solution of I2
(0.85 g, 3.34 mmol) in the same solvent (30 mL) was
added dropwise in the dark and under dry nitrogen
atmosphere. After 15 min, solid L-galactopyranose
(0.33 g, 1.67 mmol) was added in one portion to the sus-
pension. TLC monitoring (CHCl3/CH3OH, 9:1) showed
that the starting sugar was completely consumed within
30 min. The reaction mixture was then filtered through a
glass sinter funnel and washed with acetone. The solvent
was removed under reduced pressure and the solid resi-
due recrystallized from CHCl3/hexane (1:2) to give the
25
1
uct 9 (0.62 g, 75%). ½aꢁD +8.5 (c 0.9, CHCl3); H NMR
25
final product 3 (0.41 g, 95% yield); ½aꢁD ꢂ59.5 (c 1.5,
(500 MHz, CDCl3) d 1.31, 1.32, 1.43, 1.45 (4s, 12H,
25
1
CHCl3), lit.19 ½aꢁD ꢂ55.0 (c 3.6, CHCl3). H and 13C
4 · CH3), 3.24 (ddd, 1H, J6 a;6 b 13:9; J6 a;5 8:1;
0
0
0
0
NMR spectral data matched that reported.12
J6 a;NH 4:4Hz, Ha-60), 3.36–3.43 (m, 1H, H-3), 3.51
0
(ddd, 1H, J6 b;6 a 13:9; J6 b;5 6:8; J6 b;NH 4:9 Hz, Hb-60),
3.67 (d, 2H, Ja,b 13.2 Hz, 2 · CHaPMB), 3.73 (d, 2H,
Jb,a 13.2 Hz, 2 · CHbPMB), 3.79 (s, 6H, 2 · OCH3),
3.80–3.85 (m, 1H, H-50), 3.92 (dd, 1H, J4a,4b 10.7, J4a,3
4.4 Hz, Ha-4), 3.95–4.08 (m, 3H, H-40, H-2, Hb-4),
0
0
0
0
0
5.3. 6-Amino-6-deoxy-1,2:3,4-di-O-isopropylidene-a-L-
galactopyranose (4)
The title compound was prepared following May’s pro-
cedure11 starting from 3.
4.27 (dd, 1H, J2 ;1 4:9; J2 ;3 1:9 Hz, H-20), 4.50 (dd,
0
0
0
0
1H, J3 ;4 7:8; J3 ;2 1:9 Hz, H-30), 4.55 (d, 1H, Ja,b
11.7 Hz, CHaPh), 4.58 (d, 1H, Jb,a 11.7 Hz, CHbPh),
0
0
0
0
5.4. (2S,3S)-4-(Benzyloxy)-3-[di(4-methoxybenzyl)-
amino]-2-hydroxybutanoic acid (8)
5.48 (d, 1H, J1 ;2 4:9 Hz, H-10), 6.84 (d, 4H,
Jortho 8.3 Hz, ArH), 7.18 (d, 4H, Jortho 8.3 Hz, ArH),
7.28–7.40 (m, 5H, PhH), 7.50–7.58 (m, 1H, NH); 13C
NMR (125 MHz, CDCl3) d 22.9, 24.7, 25.2, 26.2, 39.7,
54.7, 55.5, 59.5, 66.3, 68.4, 69.6, 70.7, 71.0, 71.7, 73.7,
96.5, 108.9, 109.6, 114.1, 127.9, 128.7, 130.6, 131.1,
138.1, 159.0, 173.8. Anal. Calcd for C39H50N2O10: C,
66.27; H, 7.13; N, 3.96. Found: C, 66.50; H, 7.11; N,
3.93.
0
0
To a stirring solution of 7, prepared as already re-
ported,13 (1.04 g, 2.00 mmol) in MeOH (13 mL) was
added dropwise aq NaOH (1.0 M solution in H2O,
4.0 mL) at 0 ꢁC. The reaction, allowed to warm to rt,
was stirred for 6.5 h and then it was diluted with EtOAc
(100 mL). The organic phase was treated with a solution
of HCl (3 M, 2 · 100 mL), washed with brine (50 mL)
and dried over Na2SO4. The solvent was removed under
reduced pressure and the residue was purified by flash
5.6. 1-{[(1S,2S)-3-(Benzyloxy)-2-[di(4-methoxybenzyl)-
amino]-1-[(1,2:3,4-di-O-isopropylidene-6-deoxy-a-L-
galactopyranos-6-yl)carbamoyl]propyl} 4-methyl succi-
nate (10)
chromatography (CHCl3/MeOH, 9:1) to afford
8
25
(0.90 g, 92%) as a pale yellow oil. ½aꢁD +44.0 (c 0.9,
CHCl3); 1H NMR (500 MHz, CDCl3): d 3.45 (ddd,
1H, J3,2 11.0, J3,4a 9.6, J3,4b 3.4 Hz, H-3), 3.82 (s, 6H,
2 · OCH3), 3.84 (d, 1H, J2,3 11.0 Hz, H-2), 3.89 (d,
2H, Ja,b 12.0 Hz, 2 · CHaPMB), 3.98 (dd, 1H, J4a,4b
11.3, J4a,3 9.6 Hz, Ha-4), 4.10 (dd, 1H, J4b,4a 11.3, J4b,3
3.4 Hz, Hb-4), 4.22 (d, 2H, Jb,a 12.0 Hz, 2 · CHbPMB),
4.59 (d, 1H, Ja,b 11.7 Hz, CHaPh), 4.73 (d, 1H, Jb,a
11.7 Hz, CHbPh), 6.89 (d, 4H, Jortho 8.8 Hz, ArH),
7.25 (d, 4H, Jortho 8.8 Hz, ArH), 7.38–7.48 (m, 5H,
PhH); 13C NMR (125 MHz, CDCl3) d 54.2, 55.1, 60.1,
64.1, 66.2, 73.9, 114.6, 128.1, 128.6, 131.6, 136.9,
160.2, 175.0. Anal. Calcd for C27H31NO6: C, 69.66; H,
6.71; N, 3.01. Found: C, 69.42; H, 6.69; N, 3.03.
3-Carbomethoxy propionyl chloride (0.17 g, 1.16 mmol)
and anhydrous pyridine (94 lL, 1.16 mmol) were added
to a solution of compound 9 in anhydrous CH2Cl2
(11 mL) at 0 ꢁC. The stirring solution was allowed to
warm slowly to rt and after 4 h most of the solvent
was evaporated under reduced pressure and replaced
by EtOAc. The organic phase was washed with brine
(2 · 30 mL), dried (Na2SO4), and concentrated under re-
duced pressure. The residue oil was purified by silica gel
chromatography (hexane/EtOAc, 8:2!6:4) to give the
25
oily product 10 (0.56 g, 90%). ½aꢁD +18.0 (c 0.8, CHCl3);

36
S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
1H NMR (500 MHz, CDCl3) d 1.29, 1.32, 1.43, 1.45 (4s,
12H, 4 · CH3), 2.58–2.68 (m, 4H, CH2–CH2), 3.05 (ddd,
5.8. Methyl 3-{[(1S,2S)-2-hydroxy-1-(hydroxymethyl)-2-
([1,2:3,4-di-O-isopropylidene-6-deoxy-a-L-galactopyr-
anos-6-yl]carbamoyl)ethyl]carbamoyl}propanoate (12)
1H, J6 a;6 b 13:9; J6 a;5 8:8; J6 a;NH 3:5 Hz, Ha-60), 3.51–
3.56 (m, 1H, H-2), 3.60 (d, 2H, Ja,b 13.1 Hz,
2 · CHaPMB), 3.65 (d, 2H, Jb,a 13.1 Hz, 2 · CHbPMB),
3.66 (s, 3H, CH3OCO), 3.67–3.80 (m, 10H, Hb-60, H-50,
0
0
0
0
0
A solution of compound 11 (0.20 g, 0.35 mmol) in
MeOH (4 mL) was added to a stirring suspension of
5% palladium on carbon (0.07 g) in the same solvent
(5 mL) and then was hydrogenated (1 atm) at 40 ꢁC.
The flask was immersed in an ultrasound cleaning bath
filled with water and sonicated for 22 h. Then the sus-
pension was filtered through Celiteꢂ and the solid
washed twice with MeOH (2 · 5 mL). The organic phase
was evaporated down under reduced pressure to afford
0
0
0
0
2 · OCH3, 2 · H-3), 4.13 (dd, 1H, J4 ;3 7:8; J4 ;5 1:6 Hz,
H-40), 4.27 (dd, 1H, J2 ;1 4:9; J2 ;3 2:5 Hz, H-20), 4.44
0
0
0
0
0
0
0
0
(s, 2H, CH2Ph), 4.56 (dd, 1H, J3 ;4 7:8; J3 ;2 2:5 Hz,
H-30), 5.46 (d, 1H, J1 ;2 4:9 Hz, H-10), 5.51 (d, 1H, J1,2
0
0
0
0
4.3 Hz, H-1), 6.54 (dd, 1H, JNH;6 b 8:0; JNH;6 a 3:5 Hz,
NH), 6.82 (d, 4H, Jortho 8.7 Hz, ArH), 7.27 (d, 4H, Jortho
8.7 Hz, ArH), 7.30–7.39 (m, 5H, PhH); 13C NMR
(100 MHz, CDCl3) d 24.8, 25.4, 26.1, 26.4, 29.2,
29.6, 39.6, 52.2, 54.4, 55.6, 58.3, 67.2, 67.8, 71.0,
71.2, 71.9, 72.9, 73.3, 96.6, 109.2, 109.8, 114.0,
127.9, 128.1, 128.7, 130.5, 132.1, 138.7, 159.0,
169.4, 171.1, 173.1. Anal. Calcd for C44H56N2O13: C,
64.38; H, 6.88; N, 3.41. Found: C, 64.19; H, 6.85; N,
3.43.
25
the oily product 12 (0.16 g, 93%). ½aꢁD +0.4 (c 0.5,
CHCl3); 1H NMR (500 MHz, CDCl3) d 1.33, 1.36,
1.47, 1.49 (4s, 12H, 4 · CH3), 2.51–2.57 (m, 2H, CH2–
CH2), 2.63–2.69 (m, 2H, CH2–CH2), 3.37 (ddd, 1H,
J6 a;6 b 13:4; J6 a;5 8:3; J6 a;NH 4:4 Hz, Ha-60), 3.65–3.72
(m, 4H, Hb-60, CH3OCO), 3.74 (dd, 1H, JHa,Hb
11.9, JHa,1 4.9 Hz, CHaOH), 3.87 (ddd, 1H,
0
0
0
0
0
J5 ;6 a 8:3; J5 ;6 b 3:9; J5 ;4 1:9 Hz, H-50), 4.05 (dd, 1H,
JHb,Ha 11.9, JHb,1 1.5 Hz, CHbOH), 4.15–4.18 (m, 1H,
0
0
0
0
0
0
5.7. Methyl 3-{[(1S,2S)-1-[(benzyloxy)methyl]-2-
hydroxy-2-([1,2:3,4-di-O-isopropylidene-6-deoxy-a-L-
galactopyranos-6-yl]carbamoyl)ethyl]carbamoyl}pro-
panoate (11)
H-1), 4.19 (dd, 1H, J4 ;3 7:8; J4 ;5 1:9 Hz, H-40), 4.27–
0
0
0
0
0
0
0
0
4.30 (m, 1H, H-2), 4.31 (dd, 1H, J2 ;1 4:9; J2 ;3 2:4 Hz,
H-20), 4.61 (dd, 1H, J3 ;4 7:8; J3 ;2 2:4 Hz, H-30), 5.20
0
0
0
0
(br s, 1H, OH), 5.50 (d, 1H, J1 ;2 4:9 Hz, H-10), 5.90
(br s, 1H, OH), 6.77 (d, 1H, JNH,1 6.3 Hz, NHserine),
7.54–7.64 (m, 1H, NHgalactosamine); 13C NMR
(50 MHz, CDCl3) d 22.7, 23.2, 24.3, 27.4, 28.0, 28.7,
38.1, 50.2, 54.0, 60.5, 64.4, 68.8, 69.1, 70.1, 75.5, 94.6,
107.1, 107.9, 171.6, 173.2, 173.4. Anal. Calcd for
C21H34N2O11: C, 51.42; H, 6.99; N, 5.71. Found: C,
51.28; H, 6.96; N, 5.73.
0
0
A solution of CAN (4.9 mL, 2.9 mmol) in H2O was
added to a stirring solution of compound 10 (0.48 g,
0.58 mmol) in MeCN at 0 ꢁC. The reaction was kept
to 0 ꢁC for 1 h and then was allowed to warm to rt. After
18 h, the reaction mixture was quenched by the addition
of a saturated NaHCO3 solution (50 mL) and extracted
with CHCl3. The organic layer was washed with brine
until neutral, dried (Na2SO4), and concentrated under
reduced pressure. The residue, after chromatography
on silica gel (CHCl3/MeOH, 95:5), gave the oily product
5.9. 3-{[(1S,2S)-2-Hydroxy-1-(hydroxymethyl)-2-
([1,2:3,4-di-O-isopropylidene-6-deoxy-a-L-galactopyr-
anos-6-yl]carbamoyl)ethyl]carbamoyl}propanoic acid (13)
25
1
11 (0.20 g, 90%). ½aꢁD ꢂ11.0 (c 0.6, CHCl3); H NMR
(500 MHz, CDCl3) d 1.31, 1.33, 1.45, 1.47 (4s, 12H,
4 · CH3), 2.50 (t, 2H, J1,2 6.8 Hz, CH2–CH2), 2.65 (t,
2H, J2,1 6.8 Hz, CH2–CH2), 3.28 (ddd, 1H,
One molar solution of aq NaOH (0.78 mL, 0.78 mmol)
was added to a stirring solution of compound 12
(0.13 g, 0.26 mmol) in MeOH (13 mL) at 0 ꢁC. The reac-
tion mixture was allowed to warm slowly to rt and after
3 h (TLC monitoring; CHCl3/CH3OH, 8:2) was
quenched with some drops of acetic acid until neutral.
The precipitate was filtered and the organic phase was
evaporated under reduced pressure to afford the oily
J6 a;6 b 13:9; J6 a;5 8:8; J6 a;NH 4:0 Hz, Ha-60), 3.61–3.68
(m, 5H, Hb-60, CHaPMB, CH3OCO), 3.84–3.90
(m, 2H, H-50, CHbPMB), 4.16 (dd, 1H,
0
0
0
0
0
J4 ;3 7:9; J4 ;5 1:8 Hz, H-40), 4.23 (br s, 1H, H-1), 4.30
0
0
0
0
(dd, 1H, J2 ;1 4:9; J2 ;3 2:4 Hz, H-20), 4.35–4.40 (m,
1H, H-2), 4.48 (d, 1H, Ja,b 11.7 Hz, CHaPh), 4.52
(d, 1H, Jb,a 11.7 Hz, CHbPh), 4.59 (dd, 1H,
0
0
0
0
25
product 13 (0.11 g, 91%). ½aꢁD +9.0 (c 0.8, CH3OH);
J3 ;4 7:9; J3 ;2 2:4 Hz, H-30), 5.51 (d, 1H, J1 ;2 4:9 Hz,
H-10), 6.31 (br d, 1H, JNH,1 6.8 Hz, NHserine), 7.22–
7.26 (m, 1H, NHgalactosamine), 7.30–7.40 (m, 5H, PhH);
13C NMR (100 MHz, CDCl3) d 24.8, 25.3, 26.3, 26.4,
29.6, 31.2, 40.0, 52.2, 53.7, 66.6, 69.7, 70.9, 71.2, 72.1,
73.5, 96.7, 109.1, 109.9, 128.2, 128.4, 128.9, 137.8,
171.9, 173.3, 173.6. Anal. Calcd for C28H40N2O11: C,
57.92; H, 6.94; N, 4.82. Found: C, 58.14; H, 6.91; N,
4.84.
1H NMR (400 MHz, CD3OD) d 1.33, 1.36, 1.43, 1.47
(4s, 12H, 4 · CH3), 2.40–2.52 (m, 4H, CH2–CH2),
0
0
0
0
0
0
3.35 (dd, 1H, J6 a;6 b 14:0; J6 a;5 8:1 Hz, Ha-60), 3.48
0
0
0
0
(dd, 1H, J6 b;6 a 14:0; J6 b;5 4:4 Hz, Hb-60), 3.61–3.66
(m, 2H, CH2OH), 3.94–3.99 (m, 1H, H-50), 4.20 (br
d, 1H, J2,1 3.7 Hz, H-2), 4.24–4.31 (m, 2H, H-40,
0
0
0
0
H-1), 4.34 (dd, 1H, J2 ;1 5:0; J2 ;3 2:5 Hz, H-20), 4.63
0
0
0
0
(dd, 1H, J3 ;4 8:0; J3 ;2 2:5 Hz, H-30), 5.48 (d, 1H,
0
0
0
0
J1 ;2 5:0 Hz, H-10); 13C NMR (125 MHz, CD3OD)
0
0

S. Pedatella et al. / Carbohydrate Research 343 (2008) 31–38
37
d 23.2, 24.5, 25.1, 26.3, 34.0, 34.6, 40.7, 55.3, 61.1,
67.4, 71.9, 72.2, 72.9, 73.2, 97.8, 109.9, 110.5,
174.7, 176.3, 181.0. Anal. Calcd for C20H32N2O11: C,
50.42; H, 6.77; N, 5.88. Found: C, 50.28; H, 6.79; N,
5.89.
Post-processing of the trajectories (torsional, distance
and energy analysis) was performed using the CARNAL
module of AMBER 7.0 package.
Acknowledgements
5.10. 3-{[(1S,2S)-2-Hydroxy-1-(hydroxymethyl)-2-([6-
deoxy-L-galactopyranos-6-yl]carbamoyl)ethyl]carbam-
oyl}propanoic acid (2)
S.P. is grateful to Professor Steve Hanessian, who intro-
duced her to the fascinating sLex research field. We
thank Oto Miedico for collecting valuable results while
performing his master thesis work and Professor Romu-
aldo Caputo for useful discussions. We are grateful to
Dr Cristina de Castro for her help in the purification
A solution of glycoaminoacid 13 (0.11 g, 0.23 mmol) in
TFA/H2O (9:1, 5 mL) was stirred at rt. After 3 h, the
reaction solvent was evaporated under reduced pressure
to afford a crude that was purified by Sephadex G-10
column leading to the mimic 2 as a mixture of b and a
anomers in 2:1 ratio (0.067 g, 80%). 1H NMR
(500 MHz, D2O) d 2.33–2.41 (m, 4H, CH2–CH2),
3.30–3.38 (m, 2.66H, H-20b, 2 · H-60b, 2 · H-60a), 3.52
1
protocol of the final product. H and 13C NMR spectra
were performed at Centro Interdipartimentale di Met-
odologie Chimico-Fisiche (CIMCF), Universita‘ di Na-
poli Federico II. Varian Inova 500 MHz spectrometer is
the property of Naples Laboratory of Consorzio Inter-
universitario Nazionale La Chimica per l’Ambiente
(INCA).
(dd, 0.66H, J3 ;2 10:0; J3 ;4 3:3 Hz, H-30b), 3.53–3.60
0
0
0
0
0
0
(m, 2H, CH2OH), 3.63 (br dd, 0.66H, J5 ;6 a 7:4;
J5 ;6 b 5:0 Hz, H-50b), 3.68 (dd, 0.34H, J2 ;3 10:3;
0
0
0
0
J2 ;1 3:6 Hz, H-20a), 3.72 (br dd, 0.34H, J3 ;2 10:3;
0
0
0
0
J3 ;4 3:1 Hz, H-30a), 3.76 (br d, 0.66H, J4 ;3 3:3 Hz,
0
0
0
0
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