Synthesis of N-(β-d-glycuronopyranosyl)alkanamides and 1-(β-d-glycuronopyranosyl)-4-phenyl-[1,2,3]-triazoles as N-glycoprotein linkage region analogs: Examination of the effect of C5 substituent on the N-glycosidic torsion (ΦN) based on X-ray c

Article abstract of DOI:10.1016/j.carres.2013.06.023

The torsion angle around the N-glycoprotein linkage region (GlcNAc-Asn) is an important factor for presenting sugar on the cell surface which is crucial for many biological processes. Earlier studies using model and analogs showed that this important tors

Full text of DOI:10.1016/j.carres.2013.06.023

Carbohydrate Research 380 (2013) 1–8
Contents lists available at SciVerse ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Synthesis of N-(b-D-glycuronopyranosyl)alkanamides
and 1-(b-D-glycuronopyranosyl)-4-phenyl-[1,2,3]-triazoles as
N-glycoprotein linkage region analogs: examination of the effect
of C5 substituent on the N-glycosidic torsion (UN) based
on X-ray crystallography
a,
Manoharan Mathiselvam a, , Duraikkannu Loganathan , Babu Varghese b
⇑
a Department of Chemistry, Indian Institute of Technology Madras, Chennai 600036, India
b Sophisticated Analytical Instrumentation Facility, Indian Institute of Technology Madras, Chennai 600036, India
a r t i c l e i n f o
a b s t r a c t
Article history:
The torsion angle around the N-glycoprotein linkage region (GlcNAc-Asn) is an important factor for pre-
senting sugar on the cell surface which is crucial for many biological processes. Earlier studies using
model and analogs showed that this important torsion angle is greatly influenced by substitutions in
the sugar part. In the present work, uronic acid alkanamides and triazole derivatives have been designed
and synthesized as newer analogs of N-glycoprotein linkage region to understand the influence of the
Received 30 April 2013
Received in revised form 20 June 2013
Accepted 26 June 2013
Available online 6 July 2013
carboxylic group on linkage region torsion as well as on molecular packing. Crystal structure of N-(b-
galacturonopyranosyl)acetamide is solved with the space group of P22121. Comparison of the torsion
angle and molecular packing of this compound with N-(b- -galactopyranosyl)acetamide showed that
changing the C6-hydoxymethyl group to the carboxylic acid group has minimum influence on the N-gly-
cosidic torsion angle, UN and significant influence on the molecular packing.
Ó 2013 Elsevier Ltd. All rights reserved.
D-
Keywords:
N-Glycoprotein
Linkage region
N-Glycosidic torsion
Uronic acid
D
X-ray crystallography
1. Introduction
of the N-glycoprotein linkage region to examine the effect of struc-
tural variation in the linkage region. The above studies revealed
Glycoproteins are involved in a variety of functions due to enor-
mous number of structural possibilities in the glycan part.1 They
are constituents of the cell wall, plasma membrane, connective tis-
sues and also found in gastrointestinal mucus secretions. N-Glyco-
proteins are one of the major types of eukaryotic glycoproteins and
that the linkage region torsion angle, UN is influenced to a larger
extent by the structural variation of the sugar part than that of
the aglycon part. Ab initio quantum chemical calculations along
with X-ray crystallographic studies on various sugar alkanamides
showed that the C2-acetamide of the linkage GlcNAc influences
the conformation of the glycan-peptide linkage at C-1 and also
controls the rotational freedom around the C10–C20 bond which
helps to ensure minimal variation with respect to v2 (Fig. 1).11
Influence of the sugar ring substituents at C2 and C5 on the confor-
mational preference of the UN torsion angle was also studied by ab
initio quantum chemical calculations which showed that neighbor-
ing groups and environmental factors mainly inter- and intramo-
lecular interactions involving regular hydrogen bonds and the
weak C–Hꢀ ꢀ ꢀO interactions influence the energy preference of the
UN torsion angle.12 Uronic acids are biologically relevant aldohex-
oses in which the C6-hydroxymethyl group is oxidized to carbox-
ylic acid13–15 and they are found in natural products such as
glycosaminoglycans and pectins.16 In the present study, we have
designed uronic acid alkanamides as N-glycoprotein linkage region
analogs (Fig. 2) to understand the effect of carboxylic group on
N-glycosidic torsion, UN as well as on molecular assembly. We
their linkage region consists of 2-deoxy-2-acetamino-b-
D-gluco-
pyranose (GlcNAc) as a linkage sugar and
L-asparagine (Asn) as a
linkage amino acid (Fig. 1).2,3
Change in the conformation of the N-glycoprotein linkage (Glc-
NAc-Asn) can influence the presentation of the glycan chain on the
cell surface and local conformation of the protein. However, the
inherent conformational flexibility in the glycan portion of glyco-
protein and the difficulty in getting enough quantity from natural
sources are limiting factors for conformational analysis. Thus,
investigation using small models of the N-glycoprotein linkage re-
gion became essential. Earlier we have reported the results of a
systematic crystallographic4–10 investigation of model and analogs
⇑
Corresponding author. Tel.: +91 44 22575221; fax: +91 44 22574202.
E-mail address: mathi24@gmail.com (M. Mathiselvam).
Deceased on Feb 09, 2013.
0008-6215/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.carres.2013.06.023

2
M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
ONa
O6
OH
HO
O
H
N
O
H
N
O
HO
HO
i
HO
R
R
C6
HO
O5
O
OH
ω
OH
3
R = CH3
O
H
O
1
N
R = CH3
HO
HO
C1'
O
C2'
C5
2 R = CH2CH3
4 R = CH2CH3
COO
N
N1
C3'
C1
2
Scheme 1. Synthesis of N-(b-D-glucuronopyranosyl)alkanamides (route A).
Reagents and conditions: (i) TEMPO, NaOCl, KBr, THF/H2O, 0 °C, 48 h.
NH
NH3
O
triazole derivative. De-protection of this triazole derivative using
LiOH19 gave a mixture of compounds. Therefore, another method
was followed wherein the per-O-acetylated b-D-glycopyranosyl
CH3
azides20 (23 & 24) were de-O-acetylated and the primary hydroxyl
group was selectively oxidized using TEMPO to give the respective
uronic acid azides (25 & 26). These azides were then treated with
acetic anhydride in the presence of iodine to afford the 3,6-lactones
(27 & 28) which were hydrolyzed to the partially acetylated gly-
curonopyranosyl azides (29 & 30). These azides were reacted with
phenyl acetylene under click condition21 to give the corresponding
triazole derivatives (31 & 32) in moderate yield (Scheme 4). Finally,
de-O-acetylation of compounds 29 was performed by treatment
Figure 1. Schematic representation of the linkage region (GlcNAcbAsn) of the N-
glycoproteins with the depiction of torsion angles,
N1–C10, WN = C1–N1–C10–C20 and
x = O5–C5–C6–O6, UN = O5–C1–
v
2 = N1–C10–C20–C30.
HOOC
HO
HOOC
H
O
O
N
N
N
(HO)n
N
HO
R
R'
O
R = H, CH3, Cl
R' = OH, NHAc
with sodium methoxide to give 1-(b-D-glucuronopyranosyl)-4-
Figure 2. Molecular structures of uronic acid derived analogs of N-glycoprotein
linkage region.
phenyl-[1,2,3]-triazole (33). In the case of compound 32, de-O-
acetylation gave an inseparable mixture of products. The 1H NMR
spectrum of compound 33 displayed a singlet at 8.59 ppm due to
the triazole methine proton. The anomeric proton (H-1) signal
was seen as a doublet at 5.74 ppm with a coupling constant of
9.2 Hz, indicating the presence of a b-anomeric linkage.
have also designed uronic acid derivatives of 1-(b-D-glycopyrano-
syl)-4-phenyl-[1,2,3]-triazole as linkage region analogs, wherein
the amide bond is replaced by the bioisosteric triazole group.
2. Results and discussion
2.2. X-Ray crystallographic analysis of N-(b-D-
galacturonopyranosyl)acetamide (21)
2.1. Synthesis of N-(b-
D-glycuronopyranosyl)alkanamides and
uronic acid triazole derivatives
2.2.1. Structure description
Target N-(b-
sized from the corresponding N-(b-
or per-O-acetylated methyl 1-deoxy-1-azido-b-
D
-glycuronopyranosyl)alkanamides can be synthe-
-glycopyranosyl)alkanamides
-glycuronates. At
Among all the uronic acid derivatives synthesized, only N-(b-D-
D
galacturonopyranosyl)acetamide (21) crystallized with one mole-
cule of methanol in P22121 space group from the methanol med-
ium. ORTEP representation of the structure with atom numbering
is shown in Figure 3. Pyranose ring of this molecule adopts 4C1 con-
formation. The chair conformation is similar to that found in other
galactopyranose structures. A list of selected bond lengths and
D
first N-(b-D-glucopyranosyl)alkanamides (1 & 2) were prepared
from glucose following the literature procedure.4,9 The primary
hydroxymethyl group was selectively oxidized using NaOCl in
the presence of TEMPO to give the corresponding N-(b-D-glycuron-
bond angles of 21, N-b-D
-(galactopyranosyl)acetamide (34)8 and
opyranosyl)alkanamides (3–4) (Scheme 1) with excess of NaCl as
byproduct.17 Since the separation of the above uronic acid alkana-
mides from NaCl by acetylation was not successful, another meth-
od was followed wherein fully protected uronic acid azides (5–7)
related simplest monosaccharide uronic acid derivatives reported
earlier (35–37)22–24 are provided in Table 1. Comparison of the
pyranose ring C–O bond lengths showed the absence of consis-
tency in compounds 21 and 34–37. The C1–O5 bond length of com-
pound 21 was longer than the C5–O5, which was also observed in
were prepared from per-O-acetylated b-D-glycopyranosyl azides
by following the literature procedure.18 These azides were reduced
to amines which were acylated by using anhydrides to give the
corresponding fully protected alkanamides (8–14) in good yield.
Finally, the ester protections were removed by hydrolysis using
LiOH19 to give uronic acid alkanamides (15–21) (Scheme 2). The
chloroacetamide (10) was treated with sodium azide in acetone/
water (3:1) at 60 °C to give the corresponding azidoacetamide
(22) (Scheme 3) which is a good ligating agent. All these com-
pounds (8–14 & 15–22) were characterized by physical and spec-
tral methods. In 1H NMR spectrum of fully protected
alkanamides (8–14), a triplet resonating around 5.2 ppm with a
coupling constant of 9.6 Hz corresponds to the H1 proton indicat-
ing the b-anomeric configuration. The doublet resonating around
4.2 ppm with a coupling constant of 10 Hz accounts for the H5 pro-
ton which is the characteristic of uronic acid derivatives.
a
-GalUAꢀH2O (35)22 and Na-b-GlcUAꢀH2O (37)24 whereas the
reverse trend was noted for GalbNHAc (34)8 and Me(4-OMe)-
a-
GlcUA (36).23 The C6–O6A bond length is longer than that of C6–
O6B in compound 21 and a similar trend was noticed for other
uronic acid derivatives. The angle C4–C5–C6 is larger than O5–
C5–C6 angle in the compounds 21, 35 and 36 but the reverse
was noted in compounds 34 and 37. The value of the torsion angle
O5–C5–C6–O6B was found to be 12.5° which slightly deviates from
the values of ꢁ7.0° and 2.4° reported for galacturonic acid mono-
hydrate (35)22 and its methyl ester,25 respectively. The minimum
deviation observed for carboxylic group (O5–C5–C6–O6B) confor-
mation in galacturonic acid derivatives could be due to the pres-
ence of an axial hydroxyl group at the C4 position. In contrast, a
wide range of deviations are observed among the glucuronic acid
derivatives.23,24,26–29
Replacement of amide group with triazole is a well known con-
cept in medicinal chemistry. So we have planned to synthesize
uronic acid triazole derivatives from the per-O-acetylated methyl
2.2.2. Linkage region conformation
1-deoxy-1-azido-b-
phenyl acetylene under click condition to give the corresponding
D-glycuronates. The azide (7) was reacted with
The conformation of the N-glycosidic linkage is defined by the
torsion angles, UN and WN (Fig. 1). Selected torsion angles of

M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
3
O
O
O
HO
MeO
iii
i, ii
MeO
O
O
H
N
H
N
O
(HO)n
(AcO)n
R
R
(AcO)n
N3
O
O
15
, GlcUA, R = CH3 (52%)
5,
8
,
Glc
GlcUA, R = CH3 (95%)
16, GlcUA, R = CH2CH3 (43%)
17, GlcUA, R = CH2Cl (43%)
6, GlcNAc
9, GlcUA, R = CH2CH3 (92%)
10, GlcUA, R = CH2Cl (90%)
7,
Gal
18
, GlcNAcUA, R = CH3 (58%)
11
, GlcNAcUA, R = CH3 (91%)
19, GlcNAcUA, R = CH2CH3 (47%)
20, GlcNAcUA, R = CH2Cl (24%)
12, GlcNAcUA, R = CH2CH3 (93%)
13, GlcNAcUA, R = CH2Cl (94%)
21
, GalUA, R = CH3 (21%)
14
, GalUA, R = CH3 (95%)
Scheme 2. Synthesis of N-(b-
THF, 0 °C, 0.5 h.
D-glycuronopyanosyl)alkanamides (route B). Reagents and conditions: (i) H2, Pd/C, DCM; (ii) anhydride, pyridine, 0 °C; (iii) LiOH, MeOH/H2O/
O
O
MeO
i
MeO
O
O
H
N
H
N
AcO
AcO
AcO
AcO
N3
Cl
OAc
OAc
10
O
O
22 (98 %)
Scheme 3. Synthesis of methyl 1-N-azidoacetamido-2,3,4-tri-O-acetyl-b-
ronate. Reagents and conditions: (i) NaN3, acetone/H2O (2:1) 50 °C.
D-glucu-
GalUAbNHAc (21) and other reported N-(b-D-galactopyrano-
syl)alkanamides (34 & 38) are presented in Table 2. The UN values
of GalbNHAc (34), GalbNHCOCH2Cl (38) and GalUAbNHAc (21) are
ꢁ105.2°, ꢁ111.5°, and ꢁ109.7°, respectively, and the difference be-
tween the values for GalbNHAc (35) and GalUAbNHAc (21) is 4.5°
which is considerably small. The WN value of GalUAbNHAc (21) is
ꢁ176.0 which is comparable with that of GalbNHAc (34) and taken
together with the UN value revealed the Z-anti conformation of the
amide aglycon moiety. This study reveals that change in the func-
tional group from hydroxymethyl to carboxylic acid has limited
influence on the N-glycosidic torsion, UN
.
Figure 3. ORTEP representation (40% probability) of GalUAbNHAcꢀMeOH (21) with
atom numbering.
2.2.3. Molecular packing
Molecular packing in the crystal structure of the compound 21
is stabilized by several regular hydrogen bonds and C–Hꢀ ꢀ ꢀO inter-
actions listed in Tables 3 and 4. The direct hydrogen bond between
the amide N1–H and carbonyl oxygen (N1–Hꢀ ꢀ ꢀO10) present in the
N-glycoprotein linkage region model compound GlcNAcbNHAc is
absent in GalUAbNHAcꢀMeOH (21) (Fig. 4). Similar observation
was earlier noticed for GalbNHAc (34). The carbonyl oxygen O10
is acting as a bifurcated acceptor for O3-H and O4-H. A finite chain
of hydrogen bonds was also seen that begins with N1, passes
through O2, and ends in O6B. Another finite chain starts with
O6A as a donor running through O1M, O4, and ends in O10. Molec-
ular packing is further stabilized by two C–Hꢀ ꢀ ꢀO interactions (C3–
H3ꢀ ꢀ ꢀO5 & C5–H5ꢀ ꢀ ꢀO6B). The carbonyl oxygen (O6B) acts as a
bifurcated acceptor involving one regular hydrogen bond and one
C–Hꢀ ꢀ ꢀO interaction (O2–H2Oꢀ ꢀ ꢀO6Bꢀ ꢀ ꢀH5–C5). All the above non-
covalent interactions of GalUAbNHAcꢀMeOH (21) are completely
O
O
N3
AcO
iii
i, ii
NaO
HO
O
R
O
AcO
AcO
O
R
O
N3
N3
HO
OAc
R
23
R = OAc
25
R = OAc
27 R = OAc
24 R = NHAc
26 R = NHAc
28
R = NHAc
iv
O
O
OH
O
N3
HO
HO
HO
O
N
N
OH
N
vi
O
v
N
N
N
O
AcO
HO
HO
OH
33
R
OAc
R
29 R = OAc (24 %)
30 R = NHAc (23 %)
31 R = OAc (52 %)
32
R = NHAc (46 %)
Scheme 4. Synthesis of uronic acid triazole derivatives. Reagents and conditions: (i) CH3ONa, CH3OH; (ii) TEMPO, NaOCl, KBr, THF/H2O, 0 °C, 48 h; (iii) Ac2O, I2, 0 °C to rt; (iv)
H2O, 0 °C to rt; (v) phenyl acetylene, Cu(OAc)2, sodium ascorbate, acetone/water, rt; (vi) CH3ONa, CH3OH, 0 °C to rt.

4
M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
Table 1
Selected bond lengths (Å) and bond angles (°) of GalUAbNHAcꢀMeOH (21), GalbNHAc (34), a-GalUAꢀH2O (35), Me(4OMe)-a-GlcUA (36), and Na-b-GlcUAꢀH2O (37)
Parameter
GalUAbNHAcꢀMeOH (21)
GalbNHAc8 (34)
a
-GalUAꢀH2O22 (35)
Me(4-OMe)-
a
-GlcUA23 (36)
Na-b-GlcUAꢀH2O24 (37)
C1–O5
C5–O5
C6–O6a
C6–O6b
1.440(4)
1.420(4)
1.306(5)
1.197(5)
1.447(5)
1.347(5)
1.347(5)
112.5(4)
107.6(3)
105.8(3)
110.3(3)
122.3(4)
ꢁ167.9(3)
12.5(5)
1.418(4)
1.432(3)
1.422(4)
—
1.432(4)
1.358(4)
1.219(4)
103.8(3)
106.7(3)
107.6(3)
110.9(3)
122.0(3)
—
1.425(3)
1.424(3)
1.313(3)
1.212(3)
—
1.4361(16)
1.4396(18)
1.332(2)
1.189(3)
—
1.436
1.429
1.254
1.244
—
—
—
106.65
108.06
—
C1–N1
C10–N1
—
—
—
—
C10–O10
C4–C5–C6
O5–C5–C6
O5–C1–N1
C2–C1–N1
N1–C10–O10
O5–C5–C6– O6a
O5–C5–C6– O6b
108.9(2)
106.7(2)
—
—
—
174.75
ꢁ7.03
111.59(13)
103.62(12)
—
—
—
66.8(2)
ꢁ110.3(3)
—
—
156.76
ꢁ28.03
—
Table 2
HAcꢀMeOH (21) leads to the formation of a solvent channel along
Selected torsion angles of GalbNHAc (34), GalbNHCOCH2Cl (38), and GalUAbN-
HAcꢀMeOH (21)
the crystallographic axis.
Compound code
UN
WN
x
3. Summary and conclusions
GalbNHAc (34)
GalbNHCOCH2Cl (38)
GalUAbNHAcꢀMeOH (21)
ꢁ105.2(4)
ꢁ111.5
ꢁ178.4(3)
153.3
ꢁ176.3(3)
60.5(3)
61.7(3)
—
In the present study, N-(b-
D-glycuronopyranosyl)alkanamides
ꢁ109.7(4)
of various sugars have been synthesized starting from b-
D
-glyco-
pyranosyl azides in moderate yield. The 1-(glycuronopyranosyl)-
4-phenyl-[1,2,3]-triazoles were also synthesized as N-glycoprotein
linkage region analogs, where the naturally occurring amide link-
age was replaced by a bioisosteric triazole group. The crystal struc-
ture of GalUAbNHAc was solved and compared with those reported
Table 3
Regular hydrogen bond parameters for GalUAbNHAcꢀMeOH (21)
D–Hꢀ ꢀ ꢀA Hꢀ ꢀ ꢀA (Å) Dꢀ ꢀ ꢀA (Å) D–Hꢀ ꢀ ꢀA (°) Symmetry
O6A–H6Aꢀ ꢀ ꢀO1M 1.659
O1M–H4Mꢀ ꢀ ꢀO4 1.88
2.46
165.08
x, y, z
for N-(b-D-galactopyranosyl)alkanamides. This study showed that
2.697(6) 171.9
3.043(5) 167(3)
2.788(4) 151(5)
2.720(5) 111(4)
2.752(5) 157(6)
2.718(5) 148.2
x, ꢁy, ꢁz + 1
in monosaccharide analogs of the N-glycoprotein linkage region,
change of the C6-hydroxymethyl group to carboxylic acid has lim-
ited impact on glycosidic torsion, UN but noteworthy influence on
the molecular packing.
N1–H1Nꢀ ꢀ ꢀO2
O4–H4Oꢀ ꢀ ꢀO10
O4–H4Oꢀ ꢀ ꢀO3
O2–H2Oꢀ ꢀ ꢀO6B
O3–H3Oꢀ ꢀ ꢀO10
2.08(4)
1.98(6)
2.28(6)
2.07(5)
1.99
2x ꢁ 1, y, z
ꢁx+2, y + 1/2, ꢁz + 1/2
x, y, z
ꢁx + 2, y + 1/2,ꢁz + 1/2
4ꢁx + 3, y + 1/2,ꢁz + ½
4. Experimental section
Table 4
C–Hꢀ ꢀ ꢀO hydrogen bonding pattern of GalUAbNHAcꢀMeOH (21)
4.1. Materials and methods
D–Hꢀ ꢀ ꢀA
Hꢀ ꢀ ꢀA (Å)
2.545
2.666
Dꢀ ꢀ ꢀA (Å)
3.522
3.470
D–Hꢀ ꢀ ꢀA (°)
175
139
Symmetry
Thin-layer chromatograms were performed on 25 mm E. Merck
silica gel plates (60F-254). Detection was done by spraying the
plates with 10% sulfuric acid in ethanol and heating on a hot plate.
Optical rotations were measured at 30 °C on a JASCO- DIP 200 dig-
ital polarimeter using a cell of 10 mm length. NMR spectra were re-
corded on a Bruker AV400 spectrometer. IR spectrum was recorded
using JASCO FT/IR-4100 spectrometer. ESI-MS spectra were mea-
sured on a Micromass Q-Tof mass spectrometer.
C3–H3ꢀ ꢀ ꢀO5
1 + x, y, z
1 + x, y, z
C5–H5ꢀ ꢀ ꢀO6B
4.2. Crystal structures solution and refinement
Single crystals of compound 21 (GalUAbNHAcꢀMeOH) were ob-
tained from methanol at room temperature by the slow evapora-
tion method. X-ray data collection was performed with Bruker
AXS Kappa Apex II CCD diffractometer equipped with graphite
monochromated Mo (Ka) (k = 0.7107 Å) radiation. Crystal fixed at
the tip of the glass fiber was mounted on the goniometer head with
the aid of a video microscope. The automatic cell determination
routine, with 32 frames at three different orientations of the
detector was employed to collect reflections and the program
APEX2-SAINT30 was used for finding the unit cell parameters.
Four-fold redundancy per reflection was utilized for achieving
good absorption corrections using a multi-scan procedure. Besides
absorption, Lorentz, polarization and decay correction were ap-
plied during data reduction. The program SADABS was used for
absorption correction using multi-scan procedure. The structures
Figure 4. Regular (red) and C–Hꢀ ꢀ ꢀO hydrogen bonding (green) in GalUAbN-
HAcꢀMeOH (21).
different from the interactions present in the GalbNHAc (34). Thus,
the GalUAbNHAc.MeOH (21) and GalbNHAc (34) differ in their
molecular packing (Fig. 5). Presence of methanol in GalUAbN-

M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
5
Figure 5. Molecular packing in unit cell (view along a-axis). (a) GalUAbNHAcꢀMeOH (21) (b) GalbNHAc (34).
were solved by direct methods using SIR9231 and refined by full-
matrix least squares techniques using SHELXL-9732 computer pro-
grams. All hydrogen could be located in difference Fourier maps at
chemically meaningful positions. However, all hydrogen belonging
to carbon atoms were fixed at geometrically idealized positions
and riding model refinement was carried out for them. The hydo-
gen belonging to nitrogen and oxygen atoms were initially refined
with isotropic thermal parameters. The refinement showed: O2–
H2O = 0.73(5) Å, O4–H4O = 0.88(6) Å. Positions of H3O, H6O, and
H4M refined with bad thermal parameters. Hence, these O–H
hydrogen were geometrically fixed with Fourier peak positions as
starting points. O–H distances were fixed at 0.82 Å and were al-
lowed to ride during refinement. The N–H hydrogen was refined
with isotropic thermal parameters. Molecular graphics were drawn
using ORTEP3233 and Mercury programs.34 Crystal data and refine-
ment statistics are summarized in the Table 5. Complete structural
data of the 21 (GalUAbNHAcꢀMeOH) have been deposited at the
Cambridge Crystallographic Data Centre (CCDC 936098), and can
be obtained free of charge via www.ccdc.cam.ac.uk/data_request/
cif (or from the Director, Cambridge Crystallographic Data Centre,
12 Union Road, Cambridge CB2 1EZ, UK; fax: +44 1223 336033;
or email: deposit@ccdc.cam.ac.uk).
2H, H-3 & H-4), 4.96 (t, 1H, J = 8.8 Hz, H-2), 4.72 (d, 1H,
J = 8.8 Hz, H-1), 4.12 (d, 1H, J = 9.6 Hz, H-5), 3.78 (s, 3H, –OCH3),
2.08, 2.03, 2.03 (3s, 9H, 3 ꢃ –COCH3).
4.3.2. Methyl 1-deoxy-1-azido-2-deoxy-2-acetamido-3,4-di-O-
acetyl-b-D-glucuronate (6)
White solid; mp 153 °C, lit.18 175–177 °C; ½a D30
ꢁ10.2° (c 0.8,
ꢂ
CHCl3), lit.18
½
a 3D0
ꢂ
ꢁ41.8° (c 1.2, CHCl3); 1H NMR (CDCl3) d 5.67
(d, 1H, J = 8.6 Hz, –NH), 5.35 (t, 1H, J = 9.6 Hz, H-3), 5.22 (t, 1H,
J = 9.6 Hz, H-4), 4.91 (d, 1H, J = 8.6 Hz, H-1), 4.13 (d, 1H,
J = 9.6 Hz, H-5), 3.92–3.82 (m, 1H, H-2), 3.78 (s, 3H, –OCH3), 2.06,
2.04, 1.99 (3s, 9H, 3 ꢃ –COCH3).
4.3.3. Methyl 1-deoxy-1-azido-2,3,4-tri-O-acetyl-b-D-
galacturonate (7)
White solid; mp 95 °C; ½a D30
ꢂ
+2.7° (c 1, CHCl3), lit.18
½
a 3D0
+7.0° (c
ꢂ
1.1, CHCl3); 1H NMR (CDCl3) d 5.75 (d, 1H, H-4), 5.24–5.15 (m, 1H,
H-2), 5.09 (dd, 1H, J = 3.6, 10.4 Hz, H-3), 4.68 (d, 1H, J = 8.4 Hz, H-1),
4.42 (s, 1H, H-5), 3.79 (s, 3H, –OCH3), 2.13, 2.09, 2.01 (3s, 9H, 3 ꢃ –
COCH3).
4.4. General procedure for the synthesis of per-O-acetylated
methyl 1-N-alkanamido-b-
D
-glycuronate (8–14)
4.3. Per-O-acetylated methyl 1-deoxy-1-azido-b-D-glycuronate
(5–7)
Per-O-acetylated methyl 1-deoxy-1-azido-b-
D-glycuronate (5–
7) (1 mmol) and Pd/C (10 wt%, 35 mg) were taken in a 50 mL
two necked RB flask. To this, dry dichloromethane (5 mL) was
added under inert atmosphere (N2) and then changed to hydrogen
atmosphere. The solution was stirred vigorously for 6 h at room
temperature. The progress of the reaction was monitored by TLC.
After the disappearance of the starting glycosyl azide, anhydride
4.3.1. Methyl 1-deoxy-1-azido-2,3,4-tri-O-acetyl-b-D-
glucuronate (5)
White solid; mp 150 °C, lit.18 152 °C; ½a D30
ꢁ11.04° (c 1, CHCl3),
ꢂ
lit.18
½
a 3D0
ꢂ
ꢁ21.3° (c 1.1, CHCl3); 1H NMR (CDCl3) d 5.32–5.19 (m,
(1.5 mmol) and pyridine (78 lL, 1.5 mmol) were added at 0 °C
and allowed to stir at room temperature for 10 h. Then, the reac-
tion mixture was filtered through a celite bed and the organic layer
was washed with water (10 mL). The aqueous layer was extracted
with CHCl3 (3 ꢃ 10 mL), brine (10 mL), dried over Na2SO4, and con-
centrated to afford the crude product which was purified by col-
umn chromatography on silica gel (100–200 mesh) using a
mixture of hexane and ethyl acetate to give the per-O-acetylated
Table 5
Data collection and refinement statistics for GalUAbNHAc (21)
Parameter
GalUAbNHAcꢀMeOH (21)
Empirical formula
Formula weight
Wavelength
C9H17NO8
267.24
0.71073 Å
Crystal system
Space group
Orthorhombic
P22121
methyl 1-N-alkanamido-b-D-glycuronate (8–14).
Cell dimensions
a = 5.0520 (7) Å
b = 10.2882 (12) Å
c = 23.745 (3) Å
1234.2(3)
4, 1.438
0.128
4.4.1. Methyl 1-N-acetamido-2,3,4-tri-O-acetyl-b-D-glucuronate
Volume (Å3)
(8)
Z, calculated density (Mg/m3)
Absorption coefficient (mmꢁ1
F(000)
White solid; mp 146–148 °C; ½a D30
ꢂ
13.7° (c 1.0, MeOH); IR (KBr):
)
3380, 3054, 2952, 2925, 2855, 1760, 1702, 1639, 1532, 1442, 1373,
568
Crystal size (mm)
Theta range (°)
0.3 ꢃ 0.3 ꢃ 0.2
1255, 1225, 1103, 1088, 1039, 997, 937, 896, 780, 631, 598,
2.62–24.99
554 cmꢁ1 1H NMR (CDCl3) d 6.47 (d, 1H, J = 9.6 Hz, –NH), 5.38 (t,
;
Index ranges
ꢁ6 6 h 6 4, ꢁ8 6 6 12, ꢁ28 6 l 6 22
4485/2140 [R(int) = 0.0399]
2140/1/180
1H, J = 9.6 Hz, H-3), 5.29 (d, 1H, J = 9.6 Hz, H-1), 5.14 (t, 1H, H-4),
4.95 (t, 1H, J = 9.6 Hz, H-2), 4.16 (d, 1H, J = 10.0 Hz, H-5), 3.74 (s,
3H, –COOCH3), 2.06, 2.04, 1.99 (3s, 12H, 4 ꢃ –COCH3); 13C NMR
(CDCl3) d 171.1, 170.6, 169.8, 169.7, 167.3 (5 ꢃ CO), 78.1 (C-1),
74.0, 70.4, 69.8, 53.1 (–COOCH3), 23.5, 20.8, 20.7, 20.6
Reflections collected/unique
Data/restraints/parameters
Goodness-of-fit on F^2
Final R indices [I >2sigma (I)]
R indices (all data)
1.010
R1 = 0.0535, wR2 = 0.1151
R1 = 0.1060, wR2 = 0.1374

6
M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
(4 ꢃ –COCH3); ESI-MS (m/z): calcd for C15H22NO10 ([M+H]+):
H-3), 5.07 (t, 1H, J = 9.6 Hz, H-1), 4.32–4.22 (m, 1H, H-2), 4.13 (d,
1H, J = 9.6 Hz, H-5), 4.04 & 3.97 (ABq, 2H, –CH2Cl), 3.75 (s, 3H, –
COOCH3), 2.10, 2.06, 1.97 (3s, 9H, 3 ꢃ –COCH3); 13C NMR (CDCl3)
d 172.4, 170.7, 169.8, 168.2, 167.6 (5 ꢃ CO), 79.0 (C-1), 73.7, 71.9,
69.6, 52.7, 51.9, 41.9 (–COOCH3), 22.3, 20.3, 20.2 (3 ꢃ –COCH3);
ESI-MS (m/z): calcd for C15H22N2O9 ([M+H]+): 409.1014, found:
409.1027.
376.1244, found: 376.1237.
4.4.2. Methyl 1-N-propionamido-2,3,4-tri-O-acetyl-b-D-
glucuronate (9)
White solid; mp 66–69 °C; ½a D30
ꢂ
12.6° (c 1.0, MeOH); IR (KBr):
3599, 3552, 3368, 3279, 3071, 2984, 2955, 2890, 1753, 1677,
1550, 1442, 1375, 1227, 1104, 1039, 981, 896, 778, 723, 686,
638, 600, 560 cmꢁ1 1H NMR (CDCl3) d 6.36 (d, 1H, J = 9.2 Hz, –
;
4.4.7. Methyl 1-N-acetamido-2,3,4-tri-O-acetyl-b-D-
NH), 5.39 (t, 1H, J = 9.6 Hz, H-3), 5.31 (t, 1H, J = 9.6 Hz, H-1), 5.14
(t, 1H, H-4), 4.96 (t, 1H, J = 9.6 Hz, H-2), 4.17 (d, 1H, J = 10.4 Hz,
H-5), 3.73 (s, 3H, –COOCH3), 2.28–2.16 (m, 2H, –CH2CH3), 2.05,
2.04 (2s, 9H, 3 ꢃ –COCH3), 1.12 (t, 1H, J = 7.6 Hz, 3H, –CH2CH3;
13C NMR (CDCl3) d 174.4, 170.7, 169.7, 169.5, 167.3 (5 ꢃ CO),
77.8 (C-1), 73.7, 72.1, 70.3, 69.6, 52.8 (–COOCH3), 29.4 (–CH2CH3),
20.5, 20.5, 20.4 (3 ꢃ –COCH3), 9.1 (–CH2CH3); ESI-MS (m/z): calcd
for C16H24NO10 ([M+H]+): 390.1400, found: 390.1386.
galacturonate (14)
White solid; mp 79–81 °C; ½a D30
ꢂ
50.6° (c 1.0, MeOH); IR (KBr):
3380, 3054, 2952, 2925, 2855, 1760, 1702, 1639, 1532, 1442,
1373, 1247, 1226, 1132, 1084, 1054, 954, 914, 885, 752, 703,
600, 555 cmꢁ1 1H NMR (CDCl3) d 6.57 (d, 1H, J = 9.6 Hz, –NH),
;
5.75 (m, 1H, H-4), 5.32–5.25 (m, 1H, H-1), 5.22–516 (m, 2H, H-2
& H-3), 4.46 (d, 1H, J = 1.2 Hz, H-5), 3.75 (s, 3H, –COOCH3), 2.13,
2.06, 2.00 (3s, 12H, 4 ꢃ –COCH3); 13C NMR (CDCl3) d 171.5, 170.4,
170.0, 169.7, 166.9 (5 ꢃ CO), 78.2 (C-1), 73.9, 71.1, 68.5, 67.6,
53.0 (–COOCH3), 23.2, 20.8, 20.7, 20.6 (4 ꢃ –COCH3); ESI-MS (m/
z): Calcd for C15H22NO10 ([M+H]+): 376.1244, Found: 376.1241.
4.4.3. Methyl 1-N-chloroacetamido-2,3,4-tri-O-acetyl-b-D-
glucuronate (10)
White solid; mp 144–146 °C; ½a D30
ꢂ
10.6° (c 1.0, MeOH); IR (KBr):
3470, 3378, 3045, 2996, 2953, 2113, 1753, 1709, 1536, 1441, 1372,
1251, 1039, 977, 928, 897, 781, 708, 680, 641, 598, 554 cmꢁ1 1H
;
4.5. General procedure for the synthesis of N-(b-D-
glycuronopyranosyl)alkanamides (15–21)
NMR (CDCl3) d 7.40 (d, 1H, –NH), 5.41 (t, 1H, J = 9.6 Hz, H-3),
5.25 (t, 1H, J = 9.4 Hz, H-1), 5.17 (t, 1H, J = 9.6 Hz, H-4), 5.05 (t,
1H, J = 9.4 Hz, H-2), 4.19 (d, 1H, J = 10.0 Hz, H-5), 4.07 & 4.04
(ABq, 2H, –CH2Cl), 3.74 (s, 3H, –COOCH3), 2.07, 2.05, 2.05 (3s, 9H,
Per-O-acetylated methyl 1-N-alkanamido-b-D-glycuronate (8–
14) (1 mmol) was dissolved in MeOH–H2O–THF (2.5:1.0:0.5) mix-
ture (4 mL). The solution was cooled to 0 °C and LiOH (144 mg,
6 mmol) was added. After complete deprotection based on TLC
analysis, the solution was diluted with water (2 mL) and the pH
was adjusted to 7 with Amberlite IR120 (H+) ion exchange resin.
Amberlite was removed by filtration and MeOH and THF were re-
moved under reduced pressure. The remaining aqueous solution
3 ꢃ –COCH3); 13C NMR (CDCl3)
d 170.7, 169.8, 169.6, 167.1
(5 ꢃ CO), 78.3 (C-1), 74.8, 70.0, 69.6, 53.1 (–COOCH3), 42.3 (–CH2-
Cl), 20.6, 20.5 (3 ꢃ –COCH3); ESI-MS (m/z): calcd for C15H21NO10
([M+H]+): 410.0854, found: 410.0854.
4.4.4. Methyl 1-N-acetamido-2-deoxy-2-acetamido-3,4-di-O-
was lyophilized to obtain the N-(b-
mides (15–21).
D-glycuronopyranosyl)alkana-
acetyl-b-D-glucuronate (11)
White solid; mp 231–233 °C; ½a D30
ꢂ
14.7° (c 1.0, MeOH); IR (KBr):
3303, 3086, 2955, 2898, 2859, 1755, 1667, 1535, 1437, 1376, 1239,
1176, 1116, 1043, 984, 899, 731, 697, 645, 599, 555 cmꢁ1; 1H NMR
(CDCl3) d 7.14 (d, 1H, J = 8.4 Hz, –NH), 6.18 (d, 1H, J = 8.4 Hz, –NH),
5.21–5.05 (m, 3H), 4.23–4.06 (m, 2H), 3.74 (s, 3H, –COOCH3), 2.09,
2.05, 1.98, 1.96 (4s, 12H, 4 ꢃ –COCH3); 13C NMR (CDCl3) d 172.1,
172.0, 171.1, 169.6, 167.4 (5 ꢃ CO), 80.1 (C-1), 74.0, 72.2, 69.4,
53.1 (–COOCH3), 23.5, 23.2, 20.9, 20.6 (4 ꢃ –COCH3); ESI-MS (m/
z): calcd for C15H23N2O9 ([M+H]+): 375.1404, found: 375.1404.
4.5.1. N-(b-D-glucuronopyranosyl)acetamide (15)
White solid; mp 81–83 °C; ½a D30
ꢁ3.1° (c 1.5, MeOH); IR (KBr):
ꢂ
3427, 3093, 2925, 2858, 1736, 1655, 1551, 1433, 1376, 1289,
1256, 1163, 1084, 1024, 949, 697, 629, 579, 532 cmꢁ1 1H NMR
;
(D2O) d 4.95 (d, 1H, J = 9.2 Hz, H-1), 3.98 (d, 1H, J = 9.2 Hz, H-5),
3.57–3.45 (m, 2H, H-3 & H-4), 3.42–3.35 (m, 1H, H-2), 2.01 (s,
3H, –COCH3); 13C NMR (D2O) d 175.4, 172.6 (2 ꢃ CO), 79.0 (C-1),
76.1, 75.9, 71.2, 71.1, 21.9 (–COCH3); ESI-MS (m/z): Calcd for C8H13-
NO7Na ([M+Na]+): 258.0590, Found: 258.0583.
4.4.5. Methyl 1-N-propionamido-2-deoxy-2-acetamido-3,4-di-
O-acetyl-b-D-glucuronate (12)
White solid; mp 223–226 °C; ½a D30
ꢂ
14.8° (c 1.0, MeOH); IR (KBr):
4.5.2. N-(b-D-glucuronopyranosyl)propionamide (16)
White solid; mp 99–101 °C; ½a D30
ꢁ4.2° (c 1.0, MeOH); IR (KBr):
ꢂ
3297, 3087, 2947, 2927, 2855, 1752, 1667, 1535, 1439, 1376, 1299,
1237, 1113, 1107, 1043, 986, 898, 774, 726, 689, 604, 554 cmꢁ1; 1H
NMR (CDCl3) d 7.04 (d, 1H, J = 8.4 Hz, –NH), 6.03 (d, 1H, J = 8.0 Hz, –
NH), 5.22–5.05 (m, 3H, H-1, H-3 & H-4), 4.23–4.14 (m, 1H, H-2),
4.11 (d, 1H, J = 9.6 Hz, H-5), 3.74 (s, 3H, –COOCH3), 2.30–2.94 (m,
2H, –CH2CH3), 2.10, 2.06, 1.95 (3s, 9H, 3 ꢃ –COCH3), 1.11 (t, 3H, –
CH2CH3); 13C NMR (CDCl3) d 175.0, 172.1, 171.7, 169.6, 167.5
(5 ꢃ CO), 79.9 (C-1), 73.9, 72.2, 69.6, 53.1 (–COOCH3), 29.8 (–CH2-
CH3), 23.2, 20.8, 20.6 (3 ꢃ –COCH3), 9.3 (–CH2CH3); ESI-MS (m/z):
calcd for C16H25N2O9 ([M+H]+): 389.1560, found: 389.1552.
3385, 2983, 2922, 2486, 1734, 1662, 1550, 1436, 1378, 1320, 1253,
1161, 1084, 1020, 942, 906, 693, 675, 579, 527 cmꢁ1 1H NMR
;
(D2O) d 4.95 (d, 1H, J = 9.2 Hz, H-1), 3.94 (d, 1H, J = 8.8 Hz, H-5),
3.56–3.44 (m, 2H, H-3 & H-4), 3.38 (t, 1H, H-2), 2.27 (q, 2H,
J = 7.6 Hz, –CH2CH3) 1.05 (t, J = 7.6 Hz, –CH2CH3); 13C NMR (D2O)
d 179.2, 173.0 (2 ꢃ CO), 79.0 (C-1), 76.4 (C-5), 75.9, 71.2, 71.1,
28.9 (–CH2CH3), 8.9 (–CH2CH3); ESI-MS (m/z): calcd for C9H14NO7
([M+H]ꢁ): 248.0770, found: 248.0770.
4.5.3. N-(b-D-glucuronopyranosyl)chloroacetamide (17)
White solid; mp 128–130 °C; ½a D30
ꢁ4.1° (c 1.5, MeOH); IR (KBr):
ꢂ
4.4.6. Methyl 1-N-chloroacetamido-2-deoxy-2-acetamido-3,4-
3379, 3085, 2922, 2521, 1733, 1685, 1550, 1415, 1362, 1346, 1237,
1085, 1023, 950, 930, 791, 663, 582, 528, 501 cmꢁ1; 1H NMR (D2O):
d 5.00 (d, 1H, J = 8.8 Hz, H-1), 4.14 (s, 2H, –CH2Cl), 3.96 (d, 1H,
J = 8.8 Hz, H-5) 3.57–3.39 (m, 3H, H-2, H-3 & H-4); 13C NMR
(D2O) d 173.0, 170.8 (2 ꢃ CO), 79.3 (C-1), 76.5, 75.9, 71.2, 71.1,
42.0 (–CH2Cl); ESI-MS (m/z): calcd for C8H12NO7NaCl ([M+Na]+):
292.0200, found: 292.0203.
di-O-acetyl-b-D-glucuronate (13)
White solid; mp 223–225 °C; ½a D30
ꢂ
12.9° (c 1.0, MeOH); IR (KBr):
3404, 3319, 3284, 3088, 3004, 2953, 2927, 2855, 2747, 1757, 1688,
1665, 1541, 14.37 1411, 1376, 1300, 1237, 1184, 1119, 1104, 1045,
981, 898, 793, 774, 729, 683, 641, 6065, 548, 530 cmꢁ1 1H NMR
;
(CDCl3+CD3OD)
d 7.97 (d, 1H, J = 8.0 Hz, –NH), 6.02 (d, 1H,
J = 8.0 Hz, –NH), 5.21 (t, 1H, J = 9.6 Hz, H-4), 5.13 (t, 1H, J = 9.6 Hz,

M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
7
4.5.4. N-(2-Deoxy-2-acetamido-b-
D
-
167.2 (5 ꢃ CO), 77.9 (C-1), 76.8, 74.0, 71.8, 70.1, 69.6, 53.1 (–
COOCH3), 52.5 (–CH2Cl), 20.7, 20.5 (3 ꢃ –COCH3); ESI-MS (m/z):
calcd for C15H21N4O10 ([M+H]+): 417.1245, found: 417.1245.
glucuronopyranosyl)acetamide (18)
White solid; mp 151–153 °C; ½a D30
ꢂ
1.7° (c 1.0, MeOH); IR (KBr):
3427, 2925, 2858, 1736, 1655, 1551, 1431, 1377, 1310, 1254, 1163,
1082, 971, 600, 567, 508 cmꢁ1
;
1H NMR (D2O) d 5.05 (d, 1H,
4.7. General procedure for the synthesis of azides 29 & 30
J = 9.6 Hz, H-1), 3.95 (d, 1H, J = 8.8 Hz, H-5), 3.82 (t, 1H, J = 9.6 Hz,
H-2), 3.63–3.50 (m, 2H, H-3 & H-4), 1.96, 1.94 (2s, 6H, 2 ꢃ –
NHCOCH3); 13C NMR (D2O) d 174.9, 174.8, (3 ꢃ CO), 78.4 (C-1),
76.2 (C-5), 73.7, 71.3, 53.7, 22.0, 21.9 (2 ꢃ –COCH3); ESI-MS (m/
z): calcd for C10H17N2O7 ([M+H]+): 277.1036, found: 277.1035.
Per-O-acetylated b-
13.4 mmol) was converted to sodium 1-deoxy-1-azido-b-
D
-glycopyranosyl azide (23
&
24) (5 g,
-glycu-
D
ronate (25 & 26) following the literature procedure.17 The crude
product obtained with byproduct NaCl was suspended in acetic
anhydride (36 mL) and stirred at 0 °C. Iodine (19 mg, 0.87 mmol)
was added in portions and stirring was continued at 0 °C for 2 h
and at 25 °C for 10 h. The solution was poured into ice water and
extracted with dichloromethane (3 ꢃ 20 mL). The organic layer
was washed with sodium thiosulphate solution (0.5 M, 50 mL),
dried over Na2SO4, and concentrated. The resultant residue was
purified by crystallization from methanol to obtain the pure
product.
4.5.5. N-(2-Deoxy-2-acetamido-b-D-
glucuronopyranosyl)propionamide (19)
White solid; mp 84–86 °C; ½a D30
ꢂ
2.9° (c 1.2, MeOH); IR (KBr):
3404, 3095, 2977, 2925, 2857, 2491, 1737, 1656, 1549, 1434,
1377, 1311, 1237, 1161, 1078, 701, 601, 563, 527 cmꢁ1 1H NMR
;
(D2O) d 5.07 (d, 1H, J = 10.0 Hz, H-1), 3.99 (d, 1H, J = 9.6 Hz, H-5),
3.82 (t, 1H, J = 10.0 Hz, H-2), 3.64–3.51 (m, 2H, H-3 & H-4), 2.26–
2.16 (m, 2H, –CH2CH3), 1.93 (s, 3H, –NHCOCH3), 1.0 (t, 1H,
J = 7.6 Hz, –CH2CH3); 13C NMR (D2O)
d
178.8, 174.7, 172.3
4.7.1. 1-Deoxy-1-azido-2,4-di-O-acetyl-b-
D-glucuronate (29)
(3 ꢃ CO), 78.2 (C-1), 76.2 (C-5), 73.6, 71.3, 53.8, 29.0 (–CH2CH3),
21.8 (–COCH3), 9.1 (–CH2CH3); ESI-MS (m/z): calcd for
C11H17N2O7 ([M+H]+): 289.1036, found: 289.1031.
White solid; mp 61–63 °C; ½a D30
ꢂ
ꢁ43.6° (c 1.0, MeOH); IR (KBr):
3611, 3472, 3384, 3323, 3060, 2986, 2952, 2893, 2458, 2952, 2892,
2458, 2115, 1824, 1754, 1433, 1375, 1311, 1237, 1124, 1053, 992,
914, 876, 816, 739, 677, 596, 558. cmꢁ1; 1H NMR (CDCl3) d 5.45 (s,
1H, H-1), 5.08 (t, 1H, H-3), 4.97–4.91 (m, 2H, H-2, H-4), 4.37 (d, 1H,
J = 3.2 Hz, H-5), 2.20, 2.12 (2s, 6H, 2 ꢃ –COCH3); 13C NMR (CDCl3) d
170.1, 168.9, 168.9 (3 ꢃ CO), 88.2 (C1), 71.3, 68.7, 67.4, 67.2, 20.5,
20.5, 20.4 (3 ꢃ –COCH3); ESI-MS (m/z): calcd for C10H13N3O8Na
([M+Na]+): 326.0600, found: 326.0586.
4.5.6. N-(2-Deoxy-2-acetamido-b-D-
glucuronopyranosyl)chloroacetamide (20)
White solid; mp 114–116 °C; ½a D30
ꢂ
1.3° (c 1.3, MeOH); IR (KBr):
3367, 3084, 2925, 2858, 1730, 1657, 1549, 1425, 1376, 1314, 1256,
1163, 1084, 964, 788, 713, 699, 608, 558 cmꢁ1 1H NMR (D2O) d
;
5.01 (d, 1H, J = 9.6 Hz, H-1), 4.10 & 4.48 (ABq, 2H, –CH2Cl), 4.00
(d, 1H, J = 9.2 Hz, H-5), 3.88 (t, 1H, J = 9.6 Hz, H-2), 3.66–3.53 (m,
2H, H-3 & H-4), 1.94 (s, 3H, –NHCOCH3); 13C NMR (D2O) d 174.8,
172.5, 170.4 (3 ꢃ CO), 78.6 (C-1), 76.3 (C-5), 73.4, 71.3, 53.7, 41.9
(–CH2ClCCH2Cl2Cl), 21.8 (–COCH3); ESI-MS (m/z): calcd for C10H15-
N2O7NaCl ([M+Na]+): 333.0465, found: 333.0457.
4.7.2. 1-Deoxy-1-azido-2-deoxy-2-acetamido-4-O-acetyl-b-D-
glucuronate (30)
White solid; mp 123–125 °C; ½a D30
ꢁ10.7° (c 1.0, MeOH); IR
ꢂ
(KBr): 3359, 3090, 2951, 2888, 2862, 2460, 2118, 1757, 1664,
1533, 1435, 1372, 1233, 1172, 1111, 1048, 980, 895, 736, 690,
640, 595, 551 cmꢁ1 1H NMR (CDCl3) d 6.35 (d, 1H, J = 9.6 Hz, –
;
4.5.7. N-(b-
D
-galacturonopyranosyl)acetamide (21)
NH), 5.34 (s, 1H, H-1), 5.08–5.02 (m, 1H, H-4), 4.77 (t, 1H, H-3),
4.54 (dd, 1H, J = 3.4, 9.6 Hz, H-2), 4.41 (d, 1H, J = 3.2 Hz, H-5),
2.28, 2.05 (2s, 6H, 2 ꢃ –COCH3); 13C NMR (CDCl3) d 169.3, 169.1,
168.3 (3 ꢃ CO), 90.7 (C-1), 72.7, 68.2, 48.5, 23.3, 20.8 (2 ꢃ –
COCH3); ESI-MS (m/z): calcd for C10H15N4O7 ([M+H]+): 303.0941,
found: 303.0932.
White solid; mp 122–124 °C; ½a D30
ꢁ2.6° (c 1.0 MeOH); IR (KBr):
ꢂ
3402, 2927, 2862, 1746, 1656, 1553, 1436, 1375, 1290, 1236, 1144,
1087, 1043, 699, 617, 576, 545 cmꢁ1 1H NMR (D2O) d 4.94 (d, 1H,
;
J = 9.2 Hz, H-1), 4.30 (d, 1H, J = 2.4 Hz, H-5) 3.85–3.74 (m, 2H, H-3 &
H-4), 3.63 (t, 1H, J = 9.2 Hz, H-2), 2.07 (s, 3H, –NHCOCH3); 13C NMR
(D2O) d 175.5, 172.6 (2 ꢃ CO), 79.2 (C-1), 75.4, 72.8, 69.6, 68.7, 22.1
(–COCH3); ESI-MS (m/z): calcd for C8H14NO7 ([M+H]+): 236.0770,
found: 236.0774.
4.8. General procedure for the synthesis of compounds 31 & 32
To a solution of uronic acid azide (29 & 30) (0.5 mmol) in ace-
tone (1 mL), phenyl acetylene (1.05 equiv) was added. To this,
Cu(OAc)2 (20 mol %), sodium ascorbate (40 mol %), and water
(0.5 mL) were added and the reaction mixture was stirred at room
temperature for 2 h. The white precipitate formed was filtered off
and washed with water (10 mL), ethyl acetate (10 mL) to give the
product as a white solid.
4.6. Preparation of methyl 1-N-azidoacetamido-2,3,4-tri-O-
acetyl-b-
D
-glucuronate (22)
Methyl
1-N-chloroacetamido-2,3,4-tri-O-acetyl-b-D-glucuro-
nate (10) (410 mg, 1 mmol) was dissolved in acetone (15 mL). To
this solution, NaN3 (325 mg, 5 mmol) in water (7 mL) was added.
The reaction mixture was refluxed for 12 h. Acetone was removed
under vacuum and the aqueous layer was extracted with ethyl ace-
tate (2 ꢃ 15 mL). The combined organic layer was washed with
water (2 ꢃ 15 mL), brine solution (30 mL), dried over Na2SO4, and
concentrated to give the pure product (22) as a white solid in
quantitative yield.
4.8.1. 1-(2,4-Di-O-acetyl-b-D-glucuronopyranosyl)-4-phenyl-
[1,2,3]-triazole (31)
White solid; mp 195–197 °C; ½a D30
ꢁ8.4° (c 1.0, DMSO); IR (KBr):
ꢂ
3412, 3134, 2925, 2856, 1743, 1632, 1443, 1408, 1361, 1313, 1281,
1248, 1217, 1184, 1085, 1034, 982, 912, 842, 768, 692, 601, 54,
White solid; mp 148–150 °C; ½a D30
ꢂ
10.5° (c 1.1, MeOH); IR (KBr):
3408, 3023, 2960, 2936, 2214, 2116, 1755, 1700, 1532, 1437, 1374,
502 cmꢁ1 1H NMR (CD3OD)
; d 8.66 (s, 1H), 7.84 (d, 2H,
J = 7.6 Hz), 7.44 (t, 2H, J = 7.2 Hz), 7.34 (t, 1H, J = 7.2 Hz), 6.09 (d,
1H, H-1), 5.46 (t, 1H, J = 9.2 Hz, H-2), 5.23 (t, 1H, J = 9.2 Hz, H-4),
4.45 (d, 1H, J = 9.2 Hz, H-5), 4.11 (t, 1H, J = 9.2 Hz, H-3), 2.12, 1.92
(2s, 6H, 2 ꢃ –COCH3); 13C NMR (CD3OD) d 171.6, 170.8 (3 ꢃ CO),
149.3, 131.3, 130.0, 129.6, 126.8, 121.1, 86.8 (C-1), 73.9, 73.4,
72.5, 20.8, 20.3 (2 ꢃ –COCH3); ESI-MS (m/z): calcd for C18H19N3O8-
Na ([M+Na]+): 428.1070, found: 428.1081.
1327, 1223, 1103, 1039, 982, 897, 650, 603, 556, 519 cmꢁ1 1H
;
NMR (CDCl3) d 7.20 (d, 1H, –NH), 5.40 (t, 1H, J = 9.6 Hz, H-3),
5.26 (t, 1H, J = 9.6 Hz, H-1), 5.16 (t, 1H, J = 9.6 Hz, H-4), 5.01 (t,
1H, J = 9.6 Hz, H-2), 4.17 (d, 1H, J = 10.0 Hz, H-5), 4.01 & 3.95
(ABq, 2H, –CH2N3), 3.74 (s, 3H, –COOCH3), 2.07, 2.05, 2.04 (3s,
9H, 3 ꢃ –COCH3); 13C NMR (CDCl3) d 170.8, 169.8, 169.6, 167.9,

8
M. Mathiselvam et al. / Carbohydrate Research 380 (2013) 1–8
4.8.2. 1-(4-O-Acetyl-2-deoxy-2-acetamido-b-
D
-
Centre (CCDC), United Kingdom, for making the program Mercury
3.0 available for use.
glucuronopyranosyl)-4-phenyl-[1,2,3]-triazole (32)
White solid; mp 209–211 °C; ½a D30
ꢁ15.8° (c 1.0, DMSO); IR
ꢂ
(KBr): 3379, 3121, 3088, 2954, 2922, 2853, 2504, 2041, 1720,
1662, 1521, 1466, 1442, 1378, 1274, 1246, 1214, 1184, 1127,
1092, 1049, 985, 963, 937, 914, 843, 820, 765, 697, 667, 612,
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.carres.2013.06.023.
560, 521 cmꢁ1 1H NMR (CD3OD) d 8.63 (s, 1H), 7.84 (d, 2H), 7.44
;
(t, 2H, J = 7.2 Hz), 7.34 (t, 1H, J = 7.2 Hz), 6.01 (d, 1H, J = 10.0 Hz,
H-1), 5.17 (t, 1H, J = 9.2 Hz, H-4), 4.44–4.32 (m, 2H, H-2, H-5),
4.04 (t, 1H, J = 9.2 Hz, H-3), 2.12, 1.80 (2s, 6H, 2 ꢃ –COCH3); 13C
NMR (DMSO-d6) d 169.3, 169.0 (3 ꢃ CO), 146.1, 130.3, 128.7,
127.8, 125.1, 120.1, 85.3 (C-1), 71.7, 70.6, 54.5, 39.5, 22.4, 20.6
(2 ꢃ –COCH3); ESI-MS (m/z): calcd for C18H20N4O7Na ([M+Na]+):
427.1230, found: 427.1232.
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1-(2,4-Di-O-acetyl-b-D-glucuronopyranosyl)-4-phenyl-[1,2,3]-
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ꢂ
(KBr): 3435, 3137, 2924, 2858, 1738, 1634, 1436, 1393, 1355,
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We are grateful to the Indo-French Centre for Promotion of Ad-
vanced Research (IFCPAR), New Delhi, for the financial support and
encouragement. The funding provided by the Department of Sci-
ence and Technology (DST), New Delhi for the purchase of the
400 MHz NMR under IRHPA Scheme and ESI-MS under the FIST
program to the Department of Chemistry, IIT Madras is gratefully
acknowledged. The authors thank the single crystal XRD facility,
Chemistry Department, IIT Madras for the X-ray data collection.
We are thankful to Mr. V. Ram Kumar for X-ray data collection.
One of us (M.M.) is thankful to the Council of Scientific and Indus-
trial Research (CSIR), New Delhi, for the award of a Senior Research
Fellowship. We also thank the Cambridge Crystallographic Data
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