Synthesis of 1-C-linked diphosphate analogues of UDP-N-Ac-glucosamine and UDP-N-Ac-muramic acid

Article abstract of DOI:10.1016/j.tet.2008.07.009

UDP-N-acetyl-glucosamine and UDP-N-acetyl-muramic acid are two important cytoplasmic precursors of bacterial peptidoglycan. The convergent synthesis of their analogues is reported. The α-1-C-linked-N-acetyl-glucosamine was synthesized using microwave-assisted Keck radical allylation. Oxidation of alkene derivatives to the corresponding carboxylic acids allowed attachment to O- and N-sulfamoyluridine giving stable diphosphate mimetics.

Full text of DOI:10.1016/j.tet.2008.07.009

Tetrahedron 64 (2008) 9093–9100
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Synthesis of 1-C-linked diphosphate analogues of UDP-N-Ac-glucosamine
and UDP-N-Ac-muramic acid
,
a
b
b
a,c
ˇ a
ˇ
*
Andrej Babic , Stanislav Gobec , Christine Gravier-Pelletier , Yves Le Merrer , Slavko Pecar
a
ˇ
ˇ
Faculty of Pharmacy, University of Ljubljana, Askerceva 7, 1000 Ljubljana, Slovenia
b Universite´ Paris Descartes, UMR 8601 CNRS, Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, 45 rue des Saints-Pe`res, 75006 Paris, France
c
ꢀ
Jozef Stefan Institute, Jamova cesta 39, 1000 Ljubljana, Slovenia
a r t i c l e i n f o
a b s t r a c t
Article history:
UDP-N-acetyl-glucosamine and UDP-N-acetyl-muramic acid are two important cytoplasmic precursors
of bacterial peptidoglycan. The convergent synthesis of their analogues is reported. The -1-C-linked-N-
acetyl-glucosamine was synthesized using microwave-assisted Keck radical allylation. Oxidation of al-
kene derivatives to the corresponding carboxylic acids allowed attachment to O- and N-sulfamoyluridine
giving stable diphosphate mimetics.
Received 22 March 2008
Received in revised form 12 June 2008
Accepted 3 July 2008
a
Available online 8 July 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
C-Linked
glycosides substrate analogues
Diphosphate isosteres
Microwave-assisted synthesis
1. Introduction
Figure 1. This was based on similar approaches that showed
inhibition of protein glycosylation.7
N-Acetyl-glucosamine (GlcNAc) and N-acetyl-muramic acid
(MurNAc) are present in almost all eubacteria as constituent units
of the bacterial cell wall peptidoglycan.1 In the biosynthesis of
polysaccharides, glycolipids, glycoproteins and the bacterial cell
wall, GlcNAc and MurNAc are usually utilized, in the form of
precursors containing nucleoside diphosphates, by a variety of
enzymes, in particular glycosyltransferases.2–5 These enzymes are
important targets for possible modulation of certain biochemical
processes. We have focused on the enzymes involved in the cyto-
plasmic steps of peptidoglycan biosynthesis.1,6 Since MurA, MurC
and MurG enzymes transfer or modify UDP-GlcNAc or UDP-Mur-
NAc, it was reasoned that analogues of these compounds could
mimic their substrates and serve as useful starting points for
designing new enzyme inhibitors.
The design and synthesis of nucleotide diphosphate sugar ana-
logues are imposing challenges. Replacement of the diphosphate
moiety is not straightforward. Ionic, steric and H-bond forming
characteristics, as well as the distance between nucleoside and
sugar moieties, all play an important role and affect the binding to
the target enzymes. We have introduced the O- and N- sulfamoyl
amide group in lieu of the diphosphate part of UDP as depicted in
To render the target compounds chemically and enzymatically
stable, C-linked glycosides were chosen as a synthetically feasible
alternative to natural
maintaining the -configuration at the anomeric position of the N-
a
-O-glycosides.5,8–10 However, to achieve this,
a
Ac-glucosamine would be crucial. Fortunately, synthetic strategies
for making C-linked derivatives of N-Ac-glucosamine have been
thoroughly investigated.11–18 Keck radical allylation19 was chosen
as the key reaction to introduce the
anomeric position.
a
-carbon–carbon bond at the
O
NH
HO
O
HO
N
O
OH
RO
OH
O
NH
Ac
O
O
P
O
P
HO
OH
O
O
O
NH
HO
O
X = NH, O
HO
RO
N
O
O
H
NH
Ac
N
O
X
R = H,
O
S
O
OH
O
HO
OH
* Corresponding author. Tel.: þ386 1 4769 601; fax: þ386 1 4258 031.
Figure 1. Design of O- and N-sulfamoyl amide UDP-GlcNAc and UDP-MurNAc
analogues.
ˇ
E-mail address: andrej.babic@ffa.uni-lj.si (A. Babic).
0040-4020/$ – see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.07.009

ˇ
A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9094
Here we describe the synthesis of the UDP-GlcNAc and UDP-
O
AcO
AcO
AcO
Ph
Ph
O
O
a,b
O
MurNAc analogues where the uridine and C-1-linked sugar moie-
ties are connected by an O- or N-sulfamoylamide bridge, which acts
as a diphosphate isostere.
HO
Ac
Ac
NH
Ac
NH
3
5
2. Results and discussion
O
O
O
O
Ph
O
c,d
O
e
O
O
O
The target compounds were synthesized using a convergent
strategy. The O- and N-sulfamoyl uridine moieties were synthe-
sized separately from 1-C-linked N-acetyl-a-D-glucosamine de-
rivatives. Both moieties were then joined by a simple coupling
reaction followed by deprotection to afford the end compounds.
NH
Ac
NH
O
OMe
O
OH
OMe
7
6
Scheme 2. Reaction conditions: (a) NaOMe, MeOH, rt, 4 h; (b) PhCHO, TFA, 0–25 ꢀC,
3 h; (c) (R,S) 2-chloro propionic acid, NaH, dioxane, 50 ꢀC, 16 h; (d) MeI, DMF, rt, 24 h;
(e) NaIO4, RuCl3, H2O/dioxane, rt, 4 h.
Scheme 1 presents the synthesis of the desired a-1-C-linked-
GlcNAc. According to known procedures, N-acetyl-glucosamine 1
was first converted to chloride derivative 2 in acetyl chloride sat-
urated with hydrogen chloride gas.14 The reaction proceeds as
previously described albeit the instability of 2 prevents normal
chromatographic purification. In the next reaction step we formed
the desired carbon–carbon bond using Keck radical allylation. De-
spite the fact that this reaction proceeds via a free radical mecha-
nism, it is not devoid of stereoselectivity and a number of authors
have reported this reaction using chloride 2 with different resulting
Acetyl groups were removed with sodium methoxide in abso-
lute methanol at ambient temperature in quantitative yield. After
the usual work up with AmberlystÒ 15 ion exchange resin, the
crude product was reprotected at the 40- and 60-hydroxy position
with benzaldehyde and trifluoroacetic acid as catalyst.14 This gave
benzylidene derivative 5 with the 30-hydroxy group conveniently
unprotected and available for further synthesis.
The muramic acid moiety was introduced to the free 30-hydroxy
group through the Williamson ether synthesis. Racemic (R,S)-2-
chloropropionic acid was used as a reagent, since it was known that
stereoselectivity could be achieved for this reaction.23–25 As in the
a/b a/b anomer ratio as
anomer ratios.13–15,17 To achieve as high an
possible, only 3 equiv of allyltributyltin was used, and only
0.15 equiv of azobisisobutyronitrile (AIBN) initiator, as reported by
Cui and Horton.17 This lowered the yield slightly compared to that
reported by Bouvet and Ben but the 1H NMR spectrum indicated
case of 2-acetamido-1-O-benzyl-4,6-O-benzylidene-2-deoxy-a-D-
glucosamine,25 the (R)-muramic acid derivative 6 was obtained
from 5 in good yield with a diastereoisomeric ratio of 4.3:1. The
ratio was determined from the benzylidene proton signal of the
muramic and isomuramic derivatives mixture at 5.59 and
5.42 ppm, respectively. To ensure that the desired diastereoisomer
with R-lactoyl configuration was the major reaction product,
6 equiv of sodium hydride was used as a base and 5 equiv of race-
mic (R,S)-2-chloropropionic acid, which was added quickly to the
suspension. The obtained crude sodium salt was treated with
methyl iodide in DMF at ambient temperature to give 6 as a di-
astereoisomeric mixture. The diastereoisomeric mixture was
separated using flash chromatography under essentially the same
chromatographic conditions as for the methyl ester of (R,S)-1-O-
benzyl-4,6-O-benzylidene-N-acetyl-muramic acid described pre-
viously.25 Carboxylic acid 7 was obtained in the same way as
compound 4 in dioxane/water with the sodium periodate and ru-
thenium trichloride oxidation system, which left all the protecting
groups intact.
The N- and O-sulfamoyluridine derivatives 12 and 13 were
synthesized from commercially available 20,30-isopropylidene-uri-
dine 8. The 50-N-sulfamoyl derivative 12 was obtained in four re-
action steps (Scheme 3). The 50-hydroxy group was first converted
into a p-toluenesulfonyl ester 9 with p-toluenesulfonylchloride and
pyridine as the catalyst/base. The following nucleophilic sub-
stitution with sodium azide gave azido derivative 10 in high yield.
The ensuing catalytic hydrogenation proceeded to completion
within 3 h, in complete contrast to the report that the azido de-
rivative 10 was entirely resistant to reduction.26 It was established
that a dilute solution of the azido derivative 10 in methanol is re-
quired for fast catalytic hydrogenation. Compound 11 was found to
be pure by 1H NMR spectroscopy and was used without purification
in the next reaction step due to its inherent instability. Treatment of
11 with sulfamoyl chloride27 at 0 ꢀC in dry dichloromethane, with
triethylamine as the base, gave the desired 50-N-sulfamoyl-20,30-O-
isopropylidene-uridine 12 in relatively low yield. On the other
hand, 50-O-sulfamoyl-20,30-O-isopropylidene-uridine 13 was syn-
thesized in a straightforward direct sulfamoylation of 20,30-O-iso-
propylidene-uridine under reaction conditions similar to those
used for 12.
pure
a-anomer of 3. As previously observed very little side products
were formed when THF was used as a solvent in the reaction.14
AcO
HO
HO
HO
a
O
AcNH
1
O
AcO
AcO
Ac
NH
OH
Cl
2
AcO
AcO
AcO
c
O
b
O
AcO
AcO
AcO
AcNH
O
AcNH
OH
3
4
Scheme 1. Reaction conditions: (a) HCl(g), AcCl, 30 ꢀC, 16 h; (b) Bu3Snallyl, AIBN, THF,
10 min, MW; (c) NaIO4, RuCl3, H2O/dioxane, rt, 4 h.
In addition to conventional oil-bath heating, the reaction was
also attempted under microwave-assisted conditions. Using es-
sentially the same amount of reagents and solvent, a significant
reduction in reaction time was achieved, the reaction being com-
plete in only 10 min at 100 ꢀC compared to 7 h at 70 ꢀC under the
conventional procedure. A slight improvement in reaction yield
was also achieved, but more importantly, and to our surprise, only
the a-anomer was detected. Given the very short reaction time and
relatively low temperature needed to achieve total conversion of
the starting material, it appears that this novel use of microwave
irradiation for Keck radical allylation is preferable to conventional
heating.
In the next step, 1-C-allyl-N-acetyl-glucosamine 3 was oxidized
to the corresponding carboxylic acid 4 using the sodium periodate
and ruthenium trichloride oxidation system. Diluted dioxane/water
could be used instead of the tricomponent tetrachlorocarbon,
acetonitrile and water mixture to perform this reaction in very
good yield.21,22
Alkene 3 was used as the starting material for the synthesis of
1-C-linked N-acetyl-muramic acid 7. The standard two-step
deprotection and reprotection protocol for peracetylated alkene
derivative 3 was used (Scheme 2).

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A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9095
O
N
O
O
N
NH
NH
NH
O
O
N
O
O
O
O
O
O
O
d
a
O
O
O
S
S
H2N
HO
O
O
O
O
O
O
8
13
9
O
N
O
N
O
N
NH
NH
NH
O
O
O
O
O
b
c
d
O
O
O
S
N3
N
H
H2N
H2N
O
O
O
O
O
O
10
11
12
Scheme 3. Reaction conditions: (a) TsCl, pyridine, 40 ꢀC, 3 h; (b) NaN3, DMF, 50 ꢀC, 16 h; (c) H2, Pd/C, MeOH, rt, 3 h; (d) ClSO2NH2, TEA, CH2Cl2, 0–20 ꢀC, 16 h.
The 20,30-O-isopropylidene uridine derivatives 12 and 13 were
then coupled with the protected C-linked MurNAc acid 7 and
GlcNAc 4 to give N-sulfamoyl compound 14 and O-sulfamoyl de-
rivatives 15 and 18 (Schemes 4 and 5). The coupling reaction was
tested with several coupling reagents, however, using equimolar
amounts of carboxylic acid and N- or O-sulfamoyl-20,30-O-iso-
propylidene-uridine and small excesses of DCC and DMAP
(1.2 equiv) gave the best yields in short reaction times at room
O
O
NH
NH
AcO
O
AcO
N
O
N
O
O
AcO
AcO
AcO
O
X
AcO
O
O
a
O
H
N
Ac
NH
S
+
X
H2N
AcNH
S
OH
O
O
O
O
O
O
O
O
12
: X=NH
14
: X=NH
15 : X=O
4
O
13 : X=O
NH
HO
HO
HO
O
N
O
O
b,c
H
AcNH
N
X
S
O
O
HO
OH
O
16 : X=NH
17
: X=O
Scheme 4. Reaction conditions: (a) DCC, DMAP, CH2Cl2, RT, 24 h; (b) NaOMe, MeOH, rt, 4 h; (c) 70% TFA, rt, 1 h.
O
N
O
N
NH
NH
O
O
Ph
O
O
O
Ph
O
O
O
O
O
O
O
O
a
O
O
+
H
N
Ac
NH
S
Ac
NH
O
O
OH H2N
O
S
OMe
O
O
OMe
O
O
O
O
O
O
18
7
13
O
N
NH
HO
O
HO
O
O
b,c
O
H
N
AcNH
O
O
O
S
OH
O
HO
OH
O
19
Scheme 5. Reaction conditions: (a) DCC, DMAP, CH2Cl2, rt, 24 h; (b) 70% TFA, rt, 1 h; (c) 1 M LiOH, rt, 2 h.

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A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9096
temperature. However, the use of DCC as the coupling agent made
the isolation and purification difficult. Dicyclohexylurea had to
be removed completely by precipitation and filtration prior to
chromatographic purification. Only then could the products 14, 15
and 18, be isolated in a pure form without the co-elution of
dicyclohexylurea.
The deprotection procedure for C-linked N-acetyl-glucosamine
derivatives 14 and 15 (Scheme 4) proceeded smoothly. Acetyl
groups were removed in the same manner as for compound 4 with
sodium methoxide in dry methanol at room temperature. Then
acidic hydrolysis of the isopropylidene protecting group was per-
formed in 70% trifluoroacetic acid. The crude products were puri-
fied by gel filtration using Sephadex LH-20, which afforded pure
compounds 16 and 17.
In a similar manner the coupling reaction between the C-linked
N-acetyl-muramic acid derivatives 7 and O-sulfamoyl-20,30-O-iso-
propylidene-uridine 13, using DCC and DMAP, gave 18 (Scheme 5).
The benzylidene and isopropylidene protecting groups were re-
moved in 70% trifluoroacetic acid and then the ester group was
hydrolyzed under basic conditions (1 M LiOH). The product 19 was
isolated as the dilithium salt after pH adjustment with Amberlyst
15 and gel filtration on LH-20.
4.3. 3-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-
glucopyranosyl) propene (3)
4.3.1. Method A (classical heating)
2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-glucopyranosyl chlor-
ide (1.2 g, 3.3 mmol) and AIBN (170 mg, 1.1 mmol) were suspended
in allyltributyltin (3.86 mL, 12.6 mmol) and distilled THF (7.0 mL).
The reaction mixture was flushed with argon for 10 min and then
stirred at 70 ꢀC for 7 h after which TLC analysis indicated the ab-
sence of starting material. The solvent was evaporated under re-
duced pressure and the oily residue dissolved in acetonitrile
(50 mL) and extracted with pentane (5ꢂ30 mL). Flash chromato-
graphy (dichloromethane/acetone 10:1) afforded a colourless waxy
solid, which crystallized overnight giving a colourless solid (0.60 g,
1.5 mmol, 49% yield).
4.3.2. Method B (microwave-accelerated)
2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-glucopyranosyl chlor-
ide (365 mg, 1.0 mmol) and AIBN (25 mg, 0.15 mmol) were sus-
pended in allyltributyltin (1.0 mL, 3.0 mmol) and distilled THF
(2.0 mL). The reaction mixture was flushed with argon for 10 min
and then stirred in a microwave reactor at 100 ꢀC for 10 min. The
power was set at 10 W. The reactor was cooled with compressed air
during the reaction to reduce temperature fluctuations. The solvent
was evaporated under reduced pressure and the oily residue dis-
solved in acetonitrile (20 mL) and extracted with pentane
(5ꢂ15 mL). Flash chromatography (dichloromethane/acetone 10:1)
afforded a colourless waxy solid, which crystallized overnight giv-
ing colourless solid (185 mg, 0.51 mmol, 51% yield). The 1H NMR
spectra are identical using method A or B. Mp 104–105 ꢀC. IR (KBr,
3. Conclusions
The synthesis of new 1-C-linked O- and N-sulfamoylamido an-
alogues of UDP-GlcNAc and UDP-MurNAc is presented. The key
compound 3 has been synthesized using the Keck radical allylation
under microwave-accelerated conditions and converted into 1-C-
linked-MurNAc and GlcNAc carboxylic acid derivatives. Coupling
with protected N- and O-sulfamoyl uridine derivatives and the
ensuing deprotection reactions gave new diphosphate analogues of
UDP-GlcNAc and UDP-MurNAc that could provide starting points
for further development of new inhibitors of the bacterial cell wall
biosynthesis.
cmꢁ1): 3420, 2944, 1740, 1667, 1515, 1431, 1386, 1244, 1152, 1093,
20
1035, 988, 936, 924, 814, 668, 608. [
a
]
þ95.1 (c 0.20, DMF). 1H
D
NMR (CDCl3, 300 MHz) -anomer:
a
d
(ppm) 6.23 (d, 1H, J¼8.2 Hz,
NH), 5.85–5.72 (m, 1H, CH), 5.18–5.10 (m, 2H, CH2), 5.06 (t, 1H,
J¼7.8 Hz, H-3g), 4.97 (t, 1H, J¼6.8 Hz, H-4g), 4.36–4.22 (m, 3H, H-2g,
H-6g H-1g), 4,13 (dd, 1H, Jgem¼12.0 Hz, J¼3.7 Hz, H-60g), 3.94–3.88
(dt, 1H, J¼3.7, 6.5 Hz, H-5g), 2.49–2.38 (m, 1H, CHa), 2.34–2.25 (m,
1H, CHb), 2.11 (s, 3H, CH3), 2.10 (s, 3H, CH3), 2.09 (s, 3H, CH3), 1.99 (s,
4. Experimental
4.1. General
3H, CH3). 13C NMR (CDCl3, 62.9 MHz):
d (ppm) 171.10, 170.94,
170.26, 169.47, 133.78, 117.90, 71.57, 70.59, 70.38, 68.57, 62.12, 50.77,
32.13, 23.34, 21.08, 20.99. LRMS (ESI), m/z 372.2 (MþH)þ, 394.2
(MþNa)þ. HRMS (ESI), m/z calcd for C17H26NO8 372.1658 (MþH)þ,
found 372.1665.
Chemicals from Sigma–Aldrich and Fluka were used without
further purification. Analytical TLC was performed on Merck silica
gel (60 F254) plates (0.25 mm) and components visualized with
ultraviolet light and dyed with 20% sulfuric acid in ethanol, rho-
damine G6, 2,4-dinitrophenylhydrazine and ninhydrin. Flash
chromatography was performed using Merck silica gel (0.040–
0.063 mm). Gel filtration was performed using LH-20 stationary
phase and methanol as eluent. 1H, 13C, DEPT-135, gradient COSYand
gradient HSQC NMR spectra were recorded on a Bruker AM250 and
AVANCE DPX300 spectrometers in CDCl3, DMSO-d6, D2O, MeOH-d4
and acetone-d6 solution with TMS as the internal standard. Mi-
croanalyses were performed on a Perkin–Elmer C, H, N analyzer
240C. Mass spectra were obtained using a VG-Analytical Autospec
Q and Q-TOF Premier mass spectrometers.
4.4. 2-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-
glucopyranosyl)acetic acid (4)
3-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-glucopyranosyl)
propene (210 mg, 0.54 mmol) was dissolved in water (20 mL) and
dioxane (20 mL). While being stirred vigorously, sodium periodate
(460 mg, 2.15 mmol) and ruthenium(III) chloride (6 mg, 0.03 mmol)
were added. The same amount of sodium periodate was added
again after 1 h. The reaction was complete in 4 h and the solvents
were evaporated in vacuo. The dark green solid was resuspended in
ethyl acetate and filtered. The crude product obtained after evapo-
ration of ethyl acetate was purified with flash chromatography
(dichloromethane/methanol 15:1 to 8:1). Colourless oil (157 mg,
0.40 mmol, 74% yield), which slowly solidified was obtained. Mp
4.2. 2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-a-D-
glucopyranosyl chloride (2)
Compound 2 was synthesized according to Ref. 14 (
a
/
b
¼4:1).
148–150 ꢀC. IR (KBr, cmꢁ1): 3547, 2939, 2363,1748, 1558, 1374,1239,
20
a
-Anomer: 1H NMR (CDCl3, 300 MHz):
d
(ppm) 6.19 (d, J¼3.7 Hz,
1139, 1088, 1043, 913, 602. [
a
]
þ50.2 (c 0.16, MeOH). 1H NMR
D
1H, H-1g), 5.89 (d, J¼8.7 Hz,1H, NH), 5.33 (t, J¼8.9 Hz,1H, H-3g), 5.20
(t, J¼9.7 Hz,1H, H-4g), 4.58–4.50 (m, 1H, H-5g), 4.30–4.26 (m, 2H, H-
2g, H-6g), 4.14 (d, J¼10.4 Hz, 1H, H-60g), 2.10 (s, 3H, CH3), 2.05 (s, 3H,
CH3), 2.04 (s, 3H, CH3), 1.99 (s, 3H, CH3). 13C NMR (CDCl3, 62.9 MHz):
(CDCl3, 300 MHz):
d
(ppm) 6.53 (d, 1H, J¼8.4 Hz, NH), 5.05 (t, 1H,
J¼6.8 Hz, H-3g), 4.95 (t,1H, J¼6.0 Hz, H-4g), 4.62 (m, 1H, H-1g), 4.39–
4.28 (m, 2H, H-2g, H-6g), 4.22 (dd,1H, J¼11.9, 4.2 Hz, H-60g), 4.00 (dd,
1H, J¼10.4, 5.6 Hz, H-5g), 2.62 (app t, 2H, J¼6.4 Hz, CH2), 2.10 (s, 3H,
CH3), 2.09 (s, 3H, CH3), 2.08 (s, 3H, CH3), 2.00 (s, 3H, CH3). 13C NMR
d
(ppm) 171.76,170.96,170.70,169.49, 94.17, 71.18, 70.36, 68.03, 61.54,
53.64, 23.26, 21.02, 20.92, 20.80. LRMS (ESI), m/z 330.1 (MꢁCl)þ.
(CDCl3, 75 MHz): d (ppm) 173.95, 171.46, 171.31, 170.84, 169.47,

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A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9097
72.04, 69.67, 68.60, 67.88, 61.70, 49.83, 34.76, 23.21, 21.24, 21.21,
J¼10.2, 4.7 Hz, H-3g), 3.99–3.93 (m,1H, H-1g), 3.77 (s, 3H, CH3), 3.74–
3.63 (m, 3H, H-6g, H-2g, H-60g), 3.60–3.51 (m, 1H, H-5g), 2.39–2.33
(m, 2H, CH2), 2.04 (s, 3H, CH3), 1.45 (d, 3H, J¼7.0 Hz, CH3). 13C NMR
21.13. LRMS (ESI), m/z¼390.1 (MþH)þ. HRMS (ESI), m/z calcd for
C
16H24NO10 390.1400 (MþH)þ, found 390.1409. Microanalysis calcd
for C16H23NO10ꢂH2O (%): C, 47.17; H, 6.19; N, 3.44. Found: C, 47.38;
(CDCl3, 75 MHz): d (ppm) 176.06, 171.24, 137.25, 134.15, 128.95,
H, 6.36; N, 3.33.
128.29,125.74,116.98,101.19, 83.80, 75.07, 74.89, 73.75, 69.25, 63.64,
54.11, 52.35, 30.16, 23.10,18.67. LRMS (ESI), m/z 420.2 (MþH)þ, 442.2
(MþNa)þ. HRMS (ESI), m/z calcd for C22H30NO7 420.2022 (MþH)þ,
found 420.2012. Microanalysis calcd for C22H29NO7 (%): C, 62.99; H,
6.97; N, 3.34. Found: C, 63.09; H, 7.20; N, 3.36.
4.5. 3-(2-Acetamido-4,6-O-benzylidene-2-deoxy-a-D-
glucopyranosyl) propene (5)
Compound 5 was synthesized according to Ref. 14.
3-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy- -glucopyranosyl)
a
-D
4.7. 2-(2-Acetamido-4,6-O-benzylidene-2-deoxy-3-((R)-1-
propene (200 mg, 0.54 mmol) was dissolved in dry methanol
(2.0 mL). Under an argon atmosphere 30% sodium methoxide
(methoxycarbonyl)ethoxy)-a-D-glucopyranosyl)acetic acid (7)
(100
m
L) was added and the reaction mixture stirred at ambient
3-(2-Acetamido-4,6-benzylidene-2-deoxy-3-((R)-1-(methoxy-
carbonyl)ethoxy)- -glucopyranosyl) propene (100 mg, 0.24 mmol)
temperature. After 2 h, the reaction was judged complete by TLC
and the solution was neutralized using Amberlyst IR-15 ion ex-
change resin and concentrated to dryness under reduced pressure
a-D
was dissolved in water (20 mL) and dioxane (20 mL). Sodium peri-
odate (440 mg, 2.06 mmol) and ruthenium(III) chloride (5 mg,
0.02 mmol) were thenadded consecutivelyand the dark solutionwas
stirred overnight. The solution was concentrated to dryness under
reduced pressure and the dark green solid resuspended in ethyl ac-
etate and filtered off. Ethyl acetate was evaporated and the crude
product applied to a flash chromatography column. The product was
eluted by gradient elution (dichloromethane/methanol 15:1 to 5:1)
yielding colourless solid (98 mg, 0.22 mmol, 93% yield). Mp 158–
to obtain colourless oil. 3-(2-Acetamido-2-deoxy-a-D-glucopyr-
anosyl) propene was judged pure by 1H and 13C NMR spectroscopy
and was dissolved in benzaldehyde (1.8 mL) and cooled to 0 ꢀC
under argon. Anhydrous trifluoroacetic acid (100 mL) was then
added dropwise and the temperature was raised to ambient. After
3 h, the reaction mixture was evaporated to dryness under reduced
pressure, which gave a colourless solid (168 mg, 0.50 mmol, 92%
yield). Mp >240 ꢀC. IR (KBr, cmꢁ1): 3286, 1996, 1630, 1542, 1375,
160 ꢀC. IR (KBr, cmꢁ1): 3339, 2938,1734,1655,1546,1450,1372,1314,
20
20
1136, 1094, 1039, 993, 757, 697. [
(DMSO-d6, 250 MHz):
a]
D
þ64.1 (c 0.06, DMF). 1H NMR
1278, 1212, 1130, 1093, 1010, 914, 877, 749, 697. [
a
]
þ47.5 (c 0.05,
D
d
(ppm) 7.95 (d, 1H, J¼7.27 Hz, NH), 7.47–7.36
MeOH).1H NMR (CDCl3, 300 MHz):
d
(ppm) 8.01 (d,1H, J¼3.2 Hz, NH),
(m, 5H, Ph–H), 5.81–5.65 (m, 1H, CH), 5.60 (s, 1H, CH–Ph), 5.13 (d,
1H, J¼17 Hz, OH), 5.02 (d, 2H, J¼10.2 Hz, CH2), 4.06–3.84 (m, 3H, H-
3g, H-4g, H-1g), 3.75–3.61 (m, 2H, H-6g, H-2g), 3.55–3.40 (m, 2H, H-
60g, H-5g), 2.55–2.41 (m,1H, CHa), 2.24–2.14 (m,1H, CHb), 1.88 (s, 3H,
7.46–7.36 (m, 5H, Ph–H), 5.58 (s,1H, CH–Ph), 5.21–5.14 (m,1H, H-1g),
4.57 (q, 1H, J¼7.0 Hz, CH), 4.28–4.25 (m, 1H, H-4g), 4.07–3.98 (m, 1H,
H-2g),3.78(s, 3H, CH3),3.74–3.55(m, 4H, H-3g, H-5g, H-6g, H-60g),2.66
(d,1H, J¼2.8 Hz, CHa), 2.63 (br s,1H, CHb), 2.05 (s, 3H, CH3),1.45 (d, 3H,
CH3). 13C NMR (DMSO-d6, 62.9 MHz):
d
(ppm) 170.31, 138.70, 136.27,
J¼7.0 Hz, CH3). 13C NMR (CDCl3, 75 MHz):
d (ppm) 176.53, 172.78,
129.75, 129.72, 129.45, 128.88, 127.29, 117.48, 101.70, 83.83, 75.31,
69.72, 68.00, 64.64, 55.50, 31.19, 24.01. LRMS (CI), m/z 333 (M)þ,
(ESI) m/z 334.2 (MþH)þ. HRMS (ESI), m/z calcd for C18H24NO5
334.1654 (MþH)þ, found 334.1650.
137.62,129.43,128.72,126.26,101.73,83.79,75.58,72.40, 70.46, 69.49,
66.48, 62.79, 54.07, 52.92, 32.81, 23.32, 19.05. LRMS (ESI), m/z 438.2
(MþH)þ, 460.2 (MþNa)þ. HRMS (ESI), m/z calcd for C21H28NO9
438.1764(MþH)þ, found438.1754. Microanalysis calcd forC21H27NO9
(%): C, 57.66; H, 6.22; N, 3.20. Found: C, 57.46; H, 6.12; N, 3.16.
4.6. 3-(2-Acetamido-4,6-O-benzylidene-2-deoxy-3-((R)-1-
(methoxycarbonyl)ethoxy)-
a-
D-glucopyranosyl) propene (6)
4.8. 20,30-O-Isopropylidene-50-O-para-toluenesulfonyl-
uridine (9)
3-(2-Acetamido-4,6-benzylidene-2-deoxy-a-D-glucopyranosyl)
propene (420 mg, 1.26 mmol) was suspended in dioxane (40 mL)
stored over molecular sieves. While being stirred under an argon
atmosphere, sodium hydride (544 mg, 23 mmol) was added care-
fully to the suspension and the temperature raised to 50 ꢀC. After
0.5 h, the reaction mixture was cooled to ambient temperature and
Compound 9 was synthesized according to Ref. 26.
Colourless solid. Mp 93–94 ꢀC. IR (KBr, cmꢁ1): 3220, 1695, 1490,
20
1379, 1273, 1214, 1190, 1177, 1096, 977, 813, 663, 554. [
a
]
D
þ39.3 (c
0.09, DMF).1HNMR(CDCl3,250 MHz):
d
(ppm)9.22(s,1H, NH),7.69(d,
2H, J¼8.1 Hz, Ph–H), 7.26 (d, 2H, J¼8.1 Hz, Ph–H), 7.17 (d,1H, J¼8.0 Hz,
CH), 5.64(d,1H, J¼8.0 Hz, CH), 5.57 (d,1H, J¼1.3 Hz, H-1r), 4.86(dd,1H,
J¼6.4,1.6 Hz, H-2r), 4.72 (dd,1H, J¼6.2, 3.6 Hz, H-3r), 4.30–4.19 (m, 3H,
H-4r, H-5r), 2.37 (s, 3H, CH3–Ph), 1.47 (s, 3H, CH3), 1.26 (s, 3H, CH3).
LRMS (ESI), m/z 439.1 (MþH)þ, 461.1 (MþNa)þ, 477.1 (MþK)þ.
2-chloropropionic acid (580 mL, 6.3 mmol) was added dropwise to
the suspension. The reaction mixture was heated at 50 ꢀC for 4 h at
which time TLC analysis indicated total absence of starting mate-
rial. Water (1.0 mL) was added carefully at ambient temperature to
quench the excess sodium hydride and the reaction mixture was
evaporated to dryness under reduced pressure. The crude product
was suspended in brine (30 mL), cooled on ice for 1 h and the
precipitated solids filtered off. After extensive drying in vacuo for
24 h the sodium salt was dissolved in DMF (15 mL) and methyl
4.9. 50-Azido-50-deoxy-20,30-O-isopropylidene-uridine (10)
Compound 10 was synthesized according to Ref. 26.
Colourless oil. IR (KBr, cmꢁ1): 3227, 2990, 2106,1694,1543,1459,
20
iodide (400
mL, 6.4 mmol) was added in one portion at room tem-
1383,1262,1214,1158,1094, 936, 861, 807, 716, 569, 510. [
a]
þ68.5
D
perature. The reaction mixture was stirred for 24 h. The solvents
were evaporated and the crude product purified by flash chroma-
tography (dichloromethane/hexane/ethyl acetate 1:1:1) yielding
a colourless powder (253 mg, 0.60 mmol, 48% yield). Mp 209–
210 ꢀC. IR (KBr, cmꢁ1): 3311, 3083, 2948, 2873,1758,1736,1651,1553,
(c 0.02, DMF). 1H NMR (CDCl3, 250 MHz):
d (ppm) 8.77 (s, 1H, NH),
7.29 (d, 1H, J¼8.0 Hz, CH), 5.77 (d, 1H, J¼8.0 Hz, CH), 5.66 (d, 1H,
J¼2.0 Hz, H-1r), 5.00 (dd,1H, J¼6.5, 2.0 Hz, H-2r), 4.81 (dd,1H, J¼6.5,
4.2 Hz, H-3r), 4.27–4.21 (m, 1H, H-4r), 3.62 (d, 2H, J¼5.2 Hz, H-5r),
1.57 (s, 3H, CH3), 1.36 (s, 3H, CH3). LRMS (ESI), m/z¼332.1 (MþNa)þ.
1450,1372,1341,1316,1211,1174,1132,1094, 996, 922, 752, 697, 648.
20
[
a]
þ128.9 (c 0.10, DMF). 1H NMR (CDCl3, 300 MHz):
d
(ppm) 7.86
4.10. 50-Amino-50-deoxy-20,30-O-isopropylidene-uridine (11)
D
(br s,1H, NH), 7.45–7.37 (m, 5H, Ph–H), 5.83–5.69 (m,1H, CH), 5.59 (s,
1H, CH–Ph), 5.09 (dd, 1H, J¼17.0, 1.5 Hz, CHa), 5.07 (d, 1H, J¼9.9 Hz,
CHb), 4.78–4.71 (m,1H, H-4g), 4.59 (q,1H, J¼7.0 Hz, CH), 4.22 (dd,1H,
50-Azido-50-deoxy-20,30-O-isopropylidene-uridine
2.1 mmol) was dissolved in methanol (150 mL) and argon was
(650 mg,

ˇ
A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9098
bubbled through the solution for 10 min. Pd/C (50 mg, 10%) was
added and the suspension stirred under hydrogen. After 3 h, TLC
indicated the disappearance of the starting azide and the Pd/C was
filtered off. The solvent was evaporated under reduced pressure at
not more than 40 ꢀC and the colourless solid (585 mg, 2.07 mmol,
98%) dried in vacuo. 1H NMR spectroscopy indicated a high degree
0.15 mmol) and 4-dimethylaminopyridine (20 mg, 0.15 mmol) were
suspended in dry dichloromethane (15 mL). The suspension dis-
solved in 15 min and precipitate started forming after 1 h. The re-
action mixture was left stirring overnight and then concentrated to
3 mL and the precipitate filtered off and washed with the minimal
volume of dichloromethane. The solution was concentrated to dry-
ness. Flash chromatography (dichloromethane/methanol 10:1–7:1)
afforded a colourless solid (49 mg, 0.07 mmol, 51% yield). Mp 148–
of purity so the product was used without purification for further
20
synthesis. Mp 85–87 ꢀC. [
a
]
D
ꢁ22.16 (c 0.16, MeOH). 1H NMR
(DMSO-d6, 300 MHz):
d
(ppm) 7.83 (d, 1H, J¼8.0 Hz, CH), 5.78 (d,
150 ꢀC. IR (KBr, cmꢁ1): 1746, 1692,1647,1561,1458,1378,1236,1150,
þ69.8 (c 0.04, DMF). 1H NMR (CDCl3,
20
1H, J¼2.8 Hz, H-1r), 5.63 (d, 1H, J¼8.0 Hz, CH), 4.94 (dd, 1H, J¼6.5,
2.8 Hz, H-2r), 4.74 (dd, 1H, J¼6.5, 3.9 Hz, H-3r), 3.95 (m, 1H, H-4r),
3.17 (br s, 2H, NH2), 2.77 (d, 2H, J¼5.5 Hz, H-5r), 1.48 (s, 3H, CH3),
1.29 (s, 3H, CH3). LRMS (ESI), m/z 284 (MþH)þ. HRMS (ESI), m/z
calcd for C12H18N3O5 284.1246 (MþH)þ, found 284.1249.
1043, 824, 728, 562. [
300 MHz):
a
]
D
d
(ppm) 10.04 (br s, 1H, NH), 7.32 (d, 1H, J¼7.9 Hz, CH),
6.76 (br s,1H, NH), 6.36 (br s,1H, NH), 5.77 (d,1H, J¼7.9 Hz, CH), 5.47
(app s,1H, H-1r), 5.15–4.96 (m, 4H, H-3g, H-4g, H-2r, H-3r), 4.69–4.61
(m, 1H, H-1g), 4.38–4.21 (m, 6H, H-2g, H-6g, H-60g, H-4r, H-5r, H-50r),
4.15–4.08 (m, 1H, H-5g), 2.79–2.58 (m, 2H, CH2), 2.11 (br s, 9H,
3ꢂCH3), 2.04 (s, 3H, CH3),1.56 (s, 3H, CH3),1.36 (s, 3H, CH3). 13C NMR
4.11. 20,30-O-Isopropylidene-50-N-sulfamoyluridine (12)
(CDCl3, 75 MHz):
d (ppm) 171.79, 171.07, 170.55, 169.57, 169.28,
50-Amino-50-deoxy-20,30-O-isopropylidene-uridine
0.35 mmol) was dissolved in dry dichloromethane (8 mL) and tri-
ethylamine (180 L). Under an argon atmosphere, the solution was
(100 mg,
163.96, 150.61, 144.02, 114.45, 102.67, 85.64, 83.79, 81.55, 77.20,
71.38, 71.34, 69.39, 68.66, 67.90, 61.60, 50.64, 35.48, 27.00, 25.15,
22.94, 20.82, 20.72. LRMS (ESI), m/z 734.2 (MþH)þ. HRMS (ESI), m/z
calcd for C28H40N5O16S 734.2191 (MþH)þ, found 734.2211.
m
cooled to 0 ꢀC in an ice bath. Sulfamoyl chloride (65 mg, 0.56 mmol)
was added to the solution. The temperature was then raised to am-
bient temperature. After being stirred for 5 h, MeOH was used to
quench the reaction mixture, which was concentrated to dryness
under reduced pressure. Flash chromatographyafforded a colourless
4.14. 50-N-(2-(2-Acetamido-2-deoxy-
a-D-gluco-
pyranosyl)acetamidosulfamoyl)uridine (16)
oil (36 mg, 0.10 mmol, 29%). Mp 110–112 ꢀC. IR (KBr, cmꢁ1): 1690,
20,30-O-Isopropylidene-50-O-(2-(2-acetamido-3,4,6-tri-O-acetyl-
20
1460, 1386, 1334, 1273, 1215, 1159, 1089, 860, 816, 550. [
a
]
D
ꢁ3.3 (c
2-deoxy-
0.050 mmol) was dissolved in dry methanol (2.0 mL) and flushed
with argon. Sodium methoxide solution in methanol (150 L, 30%)
a-D-glucopyranosyl)acetamidosulfamoyl) uridine (37 mg,
0.08, DMF). 1H NMR (CD3CN, 250 MHz):
d
(ppm) 9.35 (br s, 1H, NH),
7.49 (d, 1H, J¼8.0 Hz, CH), 5.69 (d, 1H, J¼3.0 Hz, H-1r), 5.67 (d, 1H,
J¼8.0 Hz, CH), 5.55(t,1H, J¼6.2 Hz, NH), 5.35 (brs, 2H, NH2), 5.04 (dd,
1H, J¼6.6, 2.4 Hz, H-2r), 4.84 (dd, 1H, J¼6.6, 4.2 Hz, H-3r), 4.25–4.19
(m, 1H, H-4r), 3.35–3.29 (m, 2H, H-5r), 1.55 (s, 3H, CH3), 1.35 (s, 3H,
CH3). LRMS(ESI), m/z 385.1(MþNa)þ, 413.3(MþK)þ. HRMS(ESI), m/z
calcd for C12H18N4O7SNa 385.0794 (MþNa)þ, found 385.0814.
m
was added dropwise at ambient temperature. After being stirred for
4 h, the reaction mixture was diluted with methanol (5 mL) and
Amberlyst 15 ion exchange resin was added to adjust the pH of the
solution to 7. The resin was filtered off and methanol evaporated
under reduced pressure. The obtained white solid was dissolved in
bidistilled water (800 mL) and trifluoroacetic acid (1850 mL) and
4.12. 20,30-O-Isopropylidene-50-O-sulfamoyluridine (13)
stirred at ambient temperature. After 1 h, the solvents were evap-
orated under reduced pressure and at temperature not exceeding
30 ꢀC. Traces of water were removed by toluene co-evaporation
under reduced pressure, which gave a brownish solid. The crude
product was purified on a Sephadex LH-20 gel filtration columnwith
methanol as mobile phase. The fractions containing the product
were pooled and evaporated under reduced pressure to afford
20,30-O-Isopropylidene-uridine (300 mg, 1.06 mmol) was sus-
pended in dichloromethane (20 mL). After the addition of tri-
ethylamine (1.0 mL), the solution was cooled to 0 ꢀC. Sulfamoyl
chloride (150 mg, 1.3 mmol) was added to the stirred reaction
mixture. After 1 h another portion of sulfamoyl chloride (600 mg,
1.3 mmol) was added and the reaction mixture allowed to warm to
ambient temperature. After stirring overnight, methanol (0.5 mL)
was added and the reaction mixture filtered through a pad of Celite
545 and evaporated to dryness. The crude product was purified by
flash chromatography (ethyl acetate/methanol 20:1), which affor-
ded a colourless solid (220 mg, 0.61 mmol, 57% yield). Mp 90–92 ꢀC.
a colourless solid (17 mg, 0.030 mmol, 60% yield). Mp 158–159 ꢀC. IR
20
(KBr, cmꢁ1): 2930, 2345, 1686, 1467, 1441, 1273, 1160. [
a
]
þ80.0 (c
D
0.04, MeOH). 1H NMR (MeOH-d4, 300 MHz):
d (ppm) 7.72 (d, 1H,
J¼8.1 Hz, CH), 5.81 (d, 1H, J¼4.8 Hz, H-1r), 5.75 (d, 1H, J¼8.1 Hz, CH),
4.64–4.57 (m, 1H, H-1g), 4.25 (t, 1H, J¼5.2 Hz, H-2r), 4.11 (t, 1H,
J¼5.4 Hz, H-3r), 4.06–3.98 (m, 2H, H-4r, H-2g), 3.85–3.73 (m, 2H, H-
6g, H-60g), 3.63–3.58 (m, 2H, H-3g, H-5g), 3.45–3.29 (m, 3H, H-5r, H-
50r, H-4g), 2.68 (dd, 1H, J¼15.3, 9.9 Hz, CHa), 2.49 (dd, 1H, J¼15.3,
IR (KBr, cmꢁ1): 1686, 1560, 1465, 1380, 1274, 1216, 1183, 1070, 992,
20
932, 814, 668, 553. [
a
]
ꢁ4.0 (c 0.14, DMF). 1H NMR (DMSO-d6,
D
300 MHz):
d
(ppm) 11.43, (s, 1H, NH), 7.70 (d, 1H, J¼8.0 Hz, CH), 7.59
4.1 Hz, CHb), 2.01 (s, 3H, CH3). 13C NMR (MeOH-d4, 75 MHz):
d (ppm)
(br s, 2H, NH2), 5.82 (d, 1H, J¼1.9 Hz, H-1r), 5.64 (d, 1H, J¼8.0 Hz,
CH), 5.07 (dd, 1H, J¼6.4, 1.9 Hz, H-2r), 4.80 (dd, 1H, J¼6.4, 3.7 Hz, H-
3r), 4.26–4.21 (m, 2H, H-4r H-5), 4.15 (dd, 1H, J¼11.4, 7.8 Hz, H-50),
1.50 (s, 3H, CH3), 1.31 (s, 3H, CH3). 13C NMR (DMSO-d6, 75 MHz):
173.50, 171.39, 165.95, 152.22, 143.12, 102.83, 91.85, 83.58, 76.51,
74.40, 71.92, 71.72, 71.68, 70.65, 62.19, 53.97, 45.93, 42.38, 35.59,
22.61. LRMS (ESI), m/z 590.1 (MþNa)þ. HRMS (ESI), m/z calcd for
C
19H29N5O13SNa 590.1380 (MþNa)þ, found 590.1390. HPLC: Column
d
(ppm) 163.18, 150.26, 142.95, 113.38, 101.86, 92.81, 84.19, 83.50,
C18 Phenomex Luna 10 m; mobile phase: 20% acetonitrile, 80% tri-
80.78, 68.37, 26.90, 25.11. LRMS (ESI), m/z 364 (MþH)þ. HRMS (ESI),
fluoroacetic acid (0.1%), flow rate 1.0 mL/min; injection volume:
10 L; retention time: 2.46 min (96.47% at 220 nm, 97.38% at
m/z calcd for C12H18N3O8S 364.0815 (MþH)þ, found 364.0828.
m
254 nm).
4.13. 20,30-O-Isopropylidene-50-N-(2-(2-acetamido-3,4,6-
tri-O-acetyl-2-deoxy-
a-
D-glucopyranosyl)acet-
4.15. 4-Dimethylaminopyridine salt of 20,30-O-isopropylidene-
50-O-(2-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-
a-D-
amidosulfamoyl)uridine (14)
glucopyranosyl)acetamidosulfamoyl)uridine (15)
20,30-O-Isopropylidene-50-N-sulfamoyluridine (47 mg, 0.13 mmol),
2-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-
a
-
D
-glucopyranosyl)-
20,30-O-Isopropylidene-50-O-sulfamoyluridine (100 mg, 0.27 mmol),
2-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy- -glucopyranosyl)-
acetic acid (50 mg, 0.13 mmol), dicyclohexylcarbodiimide (32 mg,
a-D

ˇ
A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9099
acetic acid (90 mg, 0.23 mmol), dicyclohexylcarbodiimide (74 mg,
4.17. 20,30-O-Isopropylidene-50-O-(2-(2-acetamido-4,6-
benzylidene-2-deoxy-3-((R)-1-(methoxycarbonyl)ethoxy)-
a-D-glucopyranosyl)acetamidosulfamoyl)uridine (18)
0.36 mmol) and 4-dimethylaminopyridine (44 mg, 0.36 mmol)
were suspended in dry dichloromethane (20 mL). The suspension
dissolved in 15 min and a precipitate started forming after 0.5 h.
The reaction mixture was left stirring overnight. The solvent was
evaporated to concentrate the mixture to 3 mL. Then the pre-
cipitate was filtered off and washed with a minimal volume of
dichloromethane. The solution was concentrated to dryness. Gra-
dient flash chromatography (dichloromethane/methanol 10:1 to
7:1) afforded colourless solid (98 mg, 0.13 mmol, 57% yield). Mp
2-(2-Acetamido-4,6-benzylidene-2-deoxy-3-((R)-1-(methoxy-
carbonyl)ethoxy)- -glucopyranosyl)acetic acid (50 mg, 0.11 mmol),
a-D
20,30-O-isopropylidene-50-O-sulfamoyluridine (42 mg, 0.11 mmol),
dicyclohexylcarbodiimide (26 mg, 0.12 mmol) and 4-dimethyl-
aminopyridine (15 mg, 0.12 mmol) were suspended in dry
dichloromethane (10 mL). The suspension dissolved in 15 min
and a precipitate started forming after 1 h. The reaction mixture
was left stirring overnight and concentrated to 2 mL and the
precipitate filtered off and washed with a minimal volume of
dichloromethane. The solution was concentrated to dryness.
Gradient flash chromatography (dichloromethane/methanol 10:1
to 7:1) afforded colourless solid (60 mg, 0.077 mmol, 70% yield).
124–126 ꢀC. IR (KBr, cmꢁ1): 1747, 1692, 1648, 1561, 1458, 1378,
20
1236, 1151, 1043, 824, 728, 562. [
a
]
þ38.6 (c 0.03, MeOH). 1H
D
NMR (CDCl3, 300 MHz):
d
(ppm) 8.20 (d, 2H, J¼7.5 Hz, CH–Ar), 7.45
(d, 1H, J¼8.0 Hz, CH), 6.69 (d, 2H, J¼7.5 Hz, CH–Ar), 6.64 (d, 1H,
J¼8.5 Hz, NH), 5.68 (d, 1H, J¼8.0 Hz, CH), 5.64 (app s, 1H, H-1r),
5.10 (t, 1H, J¼7.5 Hz, H-3g), 5.06–4.95 (m, 3H, H-2r, H-3r, H-4g),
4.67–4.61 (m, 1H, H-1g), 4.35–4.15 (m, 6H, H-4r, H-5r, H-50r, H-2g,
H-6g, H-60g), 4.01 (m, 1H, H-5g), 3.20 (s, 6H, CH3), 2.71–2.67 (m, 2H,
CH2), 2.06 (s, 3H, CH3), 2.06 (s, 3H, CH3), 2.05 (s, 3H, CH3), 1.96 (s,
3H, CH3), 1.53 (s, 3H, CH3), 1.28 (s, 3H, CH3). 13C NMR (CDCl3,
Mp 193–194 ꢀC. IR (KBr, cmꢁ1): 1691, 1571, 1460, 1380, 1277,
20
1145, 1092, 1001, 832, 760, 698. [
NMR (CDCl3, 300 MHz):
a]
þ49.3 (c 0.16, MeOH). 1H
D
d
(ppm) 10.13 (br s, 1H, NH), 8.31 (s,
1H, NH), 7.48 (d, 2H, J¼7.6 Hz, CH), 7.42–7.33 (m, 5H, Ph–H),
5.83 (d, 1H, J¼7.6 Hz, CH), 5.74 (app s, 1H, H-1r), 5.49 (s, 1H,
CH–Ph), 5.20–5.19 (m, 1H, H-1g), 5.00–4.91 (m, 2H, H-2r, H-3r),
4.51 (q, 1H, J¼7.0 Hz, CH), 4.38–4.21 (m, 4H, H-4g, H-4r, H-5r, H-
50r), 3.91–3.88 (m, 1H, H-2g), 3.76 (s, 3H, CH3), 3.70–3.51 (m,
4H, H-3g, H-5g, H-6g, H-60g), 2.65–2.43 (m, 2H, CH2), 2.08 (s, 3H,
CH3), 1.53 (s, 3H, CH3), 1.40 (d, 3H, J¼7.0 Hz, CH3), 1.32 (s, 3H,
75 MHz):
d (ppm) 178.88, 170.80, 170.71, 169.30, 163.50, 156.93,
150.56, 142.63, 141.11, 114.06, 106.57, 102.55, 94.59, 84.99, 84.39,
80.52, 70.78, 70.18, 68.03, 67.78, 62.04, 50.18, 39.94, 39.19, 27.12,
25.13, 23.03, 20.83, 20.75, 20.72. LRMS (ESI), m/z 735.2 (MþH)þ.
HRMS (ESI), m/z calcd for C28H39N4O17S 735.2031 (MþH)þ, found
735.2061. HPLC: Column C18 Phenomex Luna 10 m; mobile phase:
50% acetonitrile, 50% trifluroacetic acid (0.1%), flow rate 1.0 mL/
min; injection volume: 10 L; retention time: 3.36 min (95.98% at
220 nm, 98.86% at 254 nm).
CH3). 13C NMR (CDCl3, 75 MHz):
d (ppm) 176.03, 173.02, 172.95,
m
164.35, 157.19, 150.40, 140.18, 137.39, 128.87, 128.20, 125.93,
114.13, 106.52, 102.58, 101.24, 84.33, 83.34, 80.61, 77.20, 75.21,
72.72, 68.95, 68.22, 64.59, 53.96, 52.50, 39.99, 27.14, 25.27,
23.08, 18.62. LRMS (ESI), m/z 783.2 (MþH)þ, 805.2 (MþNa)þ.
4.16. 50-O-(2-(2-Acetamido-2-deoxy-
a-D-
glucopyranosyl)acetamidosulfamoyl)uridine (17)
HRMS (ESI), m/z calcd for
found 734.2418.
C
33H43N4O16
S
783.2395 (MþH)þ,
20,30-O-Isopropylidene-50-O-(2-(2-acetamido-3,4,6-tri-O-acetyl-
2-deoxy-
0.088 mmol) was dissolved in dry methanol (4.0 mL) and flushed
with argon. Sodium methoxide solution in methanol (300 L, 30%)
a
-D
-glucopyranosyl)acetamidosulfamoyl) uridine (65 mg,
4.18. (R)-2-Methyl-2-(3-(2-acetamido-1-(50-O-acetamido-
sulfamoyluridine))-2-deoxy-a-D-glucopyranosyl)
m
acetic acid (19)
was added dropwise at ambient temperature. After being stirred for
4 h, the reaction mixture was diluted with methanol (10 mL) and
Amberlyst 15 ion exchange resin was added to adjust the pH of the
solution to 7. The resin was filtered off and methanol evaporated
under reduced pressure. The obtained white solid was dissolved in
20,30-O-Isopropylidene-50-O-(2-(2-acetamido-4,6-benzylidene-2-
deoxy-3-((R)-1-(methoxycarbonyl)ethoxy)- -glucopyranosyl)-
a-D
acetamidosulfamoyl)uridine (20 mg, 0.026 mmol) was suspended
in water (0.5 mL) and trifluoroacetic acid (1.5 mL) at 0 ꢀC. The
suspension dissolved immediately upon the addition of the tri-
fluoroacetic acid and stirred for 1 h. The solvents were evaporated
under reduced pressure using a rotary evaporator at temperature
not exceeding 30 ꢀC. Water and benzaldehyde traces were removed
with toluene co-evaporation (2ꢂ5 mL), affording a colourless oil,
which was dried for several hours in vacuo. Lithium hydroxide
(1 M, 1.0 mL) and methanol (1.0 mL) were added at room temper-
ature and the solution was stirred for 2 h. The solution was diluted
with methanol (2 mL) and the pH adjusted to 7 with AmberlystÒ 15
ion exchange resin. After filtration, the solvent was evaporated
under reduced pressure. The crude product was purified using
Sephadex LH-20 gel filtration column (flow rate 5 mL/h) with
methanol as mobile phase. The fractions containing the product
were pooled and evaporated under reduced pressure to afford
bidistilled water (800 mL) and trifluoroacetic acid (1.85 mL) and
stirred at ambient temperature. After 1 h, the solvents were evap-
orated under reduced pressure and at temperature not exceeding
30 ꢀC. Traces of water were removed by toluene co-evaporation
under reducedpressure, which gave a gummy white solid. The crude
product was purified using Sephadex LH-20 gel filtration column
and methanol as mobile phase. The fractions containing the product
were pooled and evaporated under reduced pressure to afford
a colourless solid (35 mg, 0.062 mmol, 70% yield). Mp 193–196 ꢀC. IR
(KBr, cmꢁ1): 2929, 2345,1686,1547,1467,1390,1273,1220, 814, 766.
20
[
a]
þ44.5 (c 0.08, MeOH).1H NMR (D2O, 300 MHz):
d (ppm) 7.53 (d,
D
1H, J¼8.1 Hz, CH), 5.77 (d,1H, J¼8.1 Hz, CH), 5.73 (d,1H, J¼4.2 Hz, H-
1r), 4.51–4.40 (m, 3H, H-1g, H-5r, H-50r), 4.20–4.11 (m, 3H, H-2r, H-3r,
H-4r), 3.84 (dd, 1H, J¼5.9, 10.6 Hz, H-2g), 3.62–3.51 (m, 3H, H-6g, H-
3g, H-60g), 3.45–3.39 (m, 1H, H-5g), 3.31 (m, 1H, H-4g), 2.68 (dd, 1H,
J¼9.3, 15.6 Hz, CHa), 2.56 (dd, 1H, J¼5.1, 15.6 Hz, CHb), 1.86 (s, 3H,
a colourless solid (16 mg, 0.024 mmol, 94% yield). Mp 211–213 ꢀC.IR
20
(KBr, cmꢁ1): 3422, 2390, 1687, 1459, 1209, 1145, 849. [
a
]
þ38.5 (c
D
CH3). 13C NMR (CDCl3, 75 MHz):
d
(ppm) 175.02, 171.17, 166.38,
0.06, MeOH). 1H NMR (MeOH-d4, 300 MHz):
d (ppm) 7.93 (d, 1H,
151.86,142.04,102.79, 89.95, 80.99, 74.26, 73.40, 71.73, 70.73, 70.58,
70.50, 69.37, 61.02, 53.15, 34.32, 22.16. LRMS (ESI), m/z 569.1
(MþH)þ. HRMS (ESI), m/z calcd for C19H29N4O14S 569.1401 (MþH)þ,
J¼8.1 Hz, CH), 5.95 (d, 1H, J¼4.9 Hz, H-1r), 5.78 (d, 1H, J¼8.1 Hz, CH),
4.92–4.84 (m, 1H, H-1g), 4.48 (q,1H, J¼6.9 Hz, CH) 4.27–4.15 (m, 5H,
H-2r, H-3r, H-2g, H-3g, H-4g), 3.75–3.40 (m, 6H, H-4r, H-5r, H-50r, H-
5g, H-6g, H-60g), 2.53 (dd, 1H, J¼14.5, 10.9 Hz, CHa), 2.31 (dd, 1H,
found 569.1407. HPLC: Column C18 Phenomex Luna 10 m; mobile
phase: 15% acetonitrile, 85% trifluoroacetic acid (0.1%), flow rate
J¼14.5, 3.6 Hz, CHb), 1.99 (s, 3H, CH3), 1.36 (d, 3H, J¼6.9 Hz, CH3). 13
C
1.0 mL/min; injection volume: 10
(98.84% at 254 nm).
mL; retention time: 2.98 min
NMR (MeOH-d4, 75 MHz): d (ppm) 179.65, 173.90, 166.00, 152.36,
142.49, 102.97, 89.49, 83.89, 78.15, 75.17, 75.16, 73.14, 72.67, 72.67,

ˇ
A. Babic et al. / Tetrahedron 64 (2008) 9093–9100
9100
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(MþH)þ, 647.2 (MþLi)þ, 653.2 (Mþ2Li)þ. HRMS (ESI), m/z calcd for
C
C
22H33N4O16S1 641.1612 (MþH)þ, found 641.1617, m/z calcd for
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This work was supported by the European Union FP6 Integrated
Project EUR-INTAFAR (Project no. LSHM-CT-2004-512138) under
the thematic priority Life Sciences, Genomics and Biotechnology
for Health, Institute Charles Nodier and the Ministry of Higher
Education, Science and Technology of the Republic of Slovenia. The
authors thank Dr. Roger H. Pain for critical reading of the
manuscript.
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