Concise synthesis of a tetrasaccharide related to the repeating unit of the cell wall O-antigen of Salmonella enterica O60

Article abstract of DOI:10.1016/j.tet.2020.131412

Synthesis of a tetrasaccharide related to the repeating unit of the cell wall O-antigen of Salmonella enterica O60 has been achieved by sequential glycosylations in very good yield. Use of p-methoxybenzyl group (PMB) as an in situ removable protecting gro

Full text of DOI:10.1016/j.tet.2020.131412

Journal Pre-proof
Concise synthesis of a tetrasaccharide related to the repeating unit of the cell wall O-
antigen of Salmonella enterica O60
Arin Gucchait, Pradip Shit, Anup Kumar Misra
PII:
S0040-4020(20)30577-9
https://doi.org/10.1016/j.tet.2020.131412
TET 131412
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 9 April 2020
Revised Date: 26 June 2020
Accepted Date: 14 July 2020
Please cite this article as: Gucchait A, Shit P, Misra AK, Concise synthesis of a tetrasaccharide related
to the repeating unit of the cell wall O-antigen of Salmonella enterica O60, Tetrahedron (2020), doi:
https://doi.org/10.1016/j.tet.2020.131412.
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Graphical Abstract
Leave this area blank for abstract info.
Concise synthesis of a tetrasaccharide
related to the repeating unit of the cell wall
O-antigen of Salmonella enterica O60
Arin Gucchait, Pradip Shit and Anup Kumar Misra
*
NIS, HClO4-SiO2
or NIS, TMSOTf
Suitably functionalized
-mannose, 3-amino-3-deoxy-
fucose thioglycoside donors and
functionalized
acceptor
D
-glucose,
HO
D
D
-
O
OH
HO
HO
HO
HO
O
HO
O HO
AcHN
O
D
-glucosamine
O
HO
O
O
OPMP
NHAc
OH
1

1
Tetrahedron
TETRAHEDRON
Pergamon
Concise synthesis of a tetrasaccharide related to the repeating
unit of the cell wall O-antigen of Salmonella enterica O60
Arin Gucchait, Pradip Shit and Anup Kumar Misra*
Bose Institute, Molecular Medicine Division, P-1/12, C.I.T. Scheme VII M, Kolkata 700054
Abstract— Synthesis of a tetrasaccharide related to the repeating unit of the cell wall O-antigen of Salmonella enterica O60 has been
achieved by sequential glycosylations in very good yield. Use of p-methoxybenzyl group (PMB) as an in situ removable protecting group
allowed obtaining the desired compound in less number of steps. Synthesis of a beta-
D-mannosidic linkage present in the molecule has
been successfully achieved using -mannosyl thioglycoside donor having a p-methoxybenzyl (PMB) group at remote C-3 position. A
D
combination of N-iodosuccinimide (NIS) and perchloric acid supported over silica (HClO4-SiO2) has been used as thiophilic glycosylation
promoter in glycosylation reactions. Thioglycoside of 3-amino-3-deoxy-D-fucose has been prepared and used in the synthetic scheme for
its incorporation in α-glycosidic linkage.
Keywords: Tetrasaccharide; glycosylation; beta-mannoside; 3-amino-3-deoxy-D-fucose; Salmonella enterica O60.
*Corresponding author. Tel.: (+91)(-33) 25693240; Fax: (+91)(-33) 2355-3886; E-mail: akmisra69@gmail.com

Tetrahedron
2
1. Introduction
obtained by fermentation process. Recently,
glycoconjugate vaccine candidates have been reported
against Salmonella infections using chemically
synthesized oligosaccharides.19,20 In this context, a
concise synthesis of the tetrasaccharide related to the
O-antigen of Salmonella enterica O60 reported herein
Food borne illness and diarrhoeal outbreaks caused by
bacterial infections are serious concern worldwide.1
The common causes for gastrointestinal infections
include intake of contaminated food and water as well
as lack of adequate sanitation.2,3 Salmonella has been
identified as one of the major pathogens among several
diarrhoea causing bacteria such as Shigella, E. coli,
Vibrio cholerae, Proteus strains causing millions of
deaths annually.4 The Salmonella infections are
commonly termed as salmonellosis5 and treated with
antimicrobial agents. Due to the emergence of the
multi-drug resistant bacterial strains 6,7 it is essential to
develop alternate mechanism for controlling of
salmonellosis. Based on the structure of the cell wall
polysaccharides or O-antigens Salmonella enterica has
been classified in several strains.8 Since the cell wall
O-polysaccharides are responsible for the virulence
properties of Salmonella, they have been considered as
important target for the development of glycoconjugate
based vaccine candidates.9,10 Although a number of
reports appeared in the past on the effective
vaccinations against bacterial infections such as
pneumococcal infections,11 influenza,12 meningitis,13
Shigellosis,14 Cholera15 etc. using bacterial cell wall
glycoconjugates, there is no such vaccine available
against salmonellosis.16 Consequently, structures of the
cell wall polysaccharides of several Salmonella
enterica strains have been reported earlier.8 The
structure of the repeating unit of the cell wall O-
antigen of Salmonella enterica O60 has been reported
by Perepelov et al.17 It is a tetrasaccharide repeating
by applying
glycosylation conditions. Presence of a rare sugar, 3-
amino-3-deoxy- -fucosamine with α-glycosidic
linkage as well as a β- -mannosidic linkage in the
a
number of recently reported
D
D
tetrasaccharide poses extra challenge in the synthetic
strategy, which was competently overcome during the
synthesis (Figure 2).
HO
O
OH
D
HO
HO
O
HO
HO
HO
AcHN
O
O
HO
O
HO
C
B
A
O
O
OPMP
NHAc
1
OH
OBn
O
Ph
OBn
O
O
AcO
HO
O
O
AcO
HO
OPMP
AllO
SEt
OPMP
NPhth
3
OAc
N3
2
4
BnO
N3
All: allyl
Ph
OBn
O
O
PMP: -methoxyphenyl
p
O
O
PMB: -methoxybenzyl
p
PMBO
STol
Phth: phthaloyl
OBn
STol
5
Tol: -methylphenyl
p
6
Figure 2: Structure of the synthesized tetrasaccharide and its
possible synthetic intermediates.
2. Results and discussion
unit comprising β-linked
glucose, β-linked -mannose and α-linked 3-amino-3-
deoxy- -fucosamine moieties (Figure 1).
D-glucosamine, β-linked D-
D
The synthesis of the target tetrasaccharide (1) as its p-
methoxyphenyl glycoside was achieved using a
sequential glycosylation approach. The synthetic
strategy of the target tetrasaccharide involved a
challenging stereoselective beta-glycosylation of a D-
mannose moiety as well as preparation of a suitably
D
α-
D
-Fucp3NFo
1
↓
3
functionalized
3-amino-3-deoxy-
D
-fucose
moiety
→2)-β-D-Manp-(1→3)-β-D-Glcp-(1→3)-β-D-GlcpNAc-(1→
together with other constituents such as
D
-glucose and
Fucp3NFo: 3-formamido-3,6-dideoxygalactose
D-glucosamine intermediates.
3-Amino-3-deoxy-D-fucose is rarely found in nature. It
Figure 1: Structure of the repeating unit of the cell
wall polysaccharide of Salmonella enterica O60.
can be found as a part of the polysaccharide chain of
cell wall in few bacterial strains. Being essential
components of a variety of antibiotics, deoxy sugars
play critical roles in their biological function. Dube and
co-workers21 summarized biological roles of a variety
of deoxy amino sugars in the bacterial polysaccharides
and their possible links in the pathogenesis. Although
the deoxyamino sugars are biosynthesized in nature, it
is quite difficult to synthesize them chemically. A
number of reports appeared from Kulkarni and co-
workers22,23 and Andreana and co-workers24 on the
synthetic aspects of rare deoxy aminosugars.22-24
However, there is no report available in the literature
Conventionally, the oligosaccharides can be obtained
by bacterial fermentation with some inherent
drawbacks, such as handling of living organisms, lack
of adequate purity in terms of biological
contamination, insufficient quantity and lack of
homogeneity.18 Therefore, development of concise
chemical synthesis is the best option for getting access
to the sufficient quantity of required oligosaccharides
with adequate purity avoiding the above mentioned
shortcomings. It has been demonstrated that
glycoconjugates
prepared
using
synthetic
on the synthesis of 3-amino-3-deoxy- -fucose
D
oligosaccharide fragments of the bacterial cell wall
could serve as effective vaccine candidate similar to
the vaccines prepared using cell wall polysaccharides
derivative for its use in the glycosylation. Suitably
functionalized monosaccharide intermediates 2,25 3, 4,

Tetrahedron
3
526 and 6 were prepared using reaction conditions
available in the literature. Preparation of compounds
225 and 526 were carried out using the earlier reported
reaction conditions. Compounds 3, 4 and 6 were
synthesized from earlier reported monosaccharide
derivatives 7,27 828 and 929 respectively.
followed by functionalization of the remaining
hydroxyl groups and inversion of C-3 hydroxyl group
using minimum number of steps. However, it was
observed that no inversion product was obtained in
case of 2-O-benzylated derivative except formation of
a complex mixture. Earlier, similar observation was
reported by Ramström and co-workers.35,36 The
inversion reaction of triflate group proceeded smoothly
only when an ester group was present at the carbon
adjacent to the carbon atom carrying the leaving triflate
group. On the contrary, a complex mixture was formed
rapidly without formation of the inversion product
when benzyl groups were employed. These results
suggested that a neighboring ester group is essential to
induce or activate the inversion reaction, whereas an
ether derivative is unable to produce this effect. It was
also shown35,36 that the inversion reaction proceeded
smoothly regardless of the triflate configuration.
Therefore, it was decided to re-design the synthetic
scheme by functionalizing the C-2 hydroxy group with
a benzoyl group to achieve better yield of the inversion
steps, although the number of steps increased.
Compound 9 was subjected to a series of reactions
comprising (a) treatment of 2-naphthylmethyl (NAP)
bromide in the presence of sodium hydride;37 (b)
removal of isopropylidene group using 80% aq. acetic
acid;38 (c) selective benzylation of the axial hydroxyl
group at C-4 using a phase transfer reaction condition;
(d) esterification of the remaining hydroxyl group at C-
3 using levulinic acid in the presence of DIC and
DMAP;30 (e) oxidative removal of the NAP group
using DDQ;39 (f) benzoylation of the free hydroxyl
group followed by (g) removal of the levulinic ester
using hydrazine acetate32 to give tolyl 2-O-benzoyl-4-
p-Methoxyphenyl 4-O-acetyl-6-O-benzyl-2-deoxy-2-
N-phthalimido-β-
D
-glucopyranoside
(3)
was
synthesized from p-methoxyphenyl 4,6-O-benzylidene-
2-deoxy-2-N-phthalimido-β-
D
-glucopyranoside (7)27
using a sequence of reactions, involving esterification
of the C-3 hydroxyl group using levulinic acid in the
presence of DIC and DMAP;30 reductive ring opening
of 4,6-O-benzylidene acetal using a combination of
triethylsilane and trifluoroacetic acid;31 acetylation of
the resulting C-4 hydroxyl group and finally removal
of the levulinic ester using hydrazine acetate32 in 62%
over all yield. NMR spectroscopic analysis confirmed
the formation of compound 3 (Scheme 1).
BnO
AcO
HO
Ph
O
a, b, c, d
O
O
O
OPMP
NPhth
OPMP
NPhth
HO
3
7
Scheme 1: Preparation of 2-deoxy-2-N-phthalimido-D-glucose
intermediate as glycosyl acceptor. Reagents and conditions: (a)
levulinic acid, DIC, DMAP, CH2Cl2, r t, 5 h; (b) Et3SiH, TfOH,
CH2Cl2, 0 °C, 3 h; (c) Ac2O, pyridine, r t, 2 h; (d) AcOH,
NH2NH2·H2O, CH3OH-CH2Cl2 (1:1), 0 °C, 8 h, 62% in four
steps.
p-Methoxyphenyl 4-O-acetyl-2-azido-6-O-benzyl-2-
deoxy-β-D-glucopyranoside (4) was synthesized from
p-methoxyphenyl 2-azido-4,6-O-benzylidene-2-deoxy-
β- sequence of
D
-glucopyranoside (8)28 using
a
O-benzyl-1-thio-β-D-fucopyranoside (10) in 45% over
reactions, involving allylation using allyl bromide in
the presence of sodium hydride;33 reductive ring
all yield. The free hydroxyl group at C-3 in compound
10 was epimerized by the treatment of triflic anhydride
in the presence of pyridine followed by SN2
substitution of the triflate group by hydroxyl group in
the presence of sodium nitrite40 and removal of benzoyl
group using sodium methoxide furnished compound 11
in 58% over all yield. The equatorial C-2 hydroxyl
group in the diol derivative 11 was selectively
benzylated via stannylidene acetal41 formation. The C-
3 hydroxyl group was treated with triflic anhydride in
the presence of pyridine to give triflate derivative
which was subjected to the SN2 inversion using sodium
azide42 to furnish compound 6 in 60% over all yield.
NMR spectroscopic analysis confirmed the formation
of compound 6 (Scheme 3).
opening of 4,6-O-benzylidene acetal using
a
combination of triethylsilane and trifluoroacetic acid;31
acetylation of the resulting C-4 hydroxyl group and
finally removal of the allyl ether using palladium
chloride34 in 53% over all yield. NMR spectroscopic
analysis confirmed the formation of compound 4
(Scheme 2).
Ph
BnO
AcO
HO
O
a, b, c, d
O
O
O
HO
OPMP
OPMP
N3
N3
4
8
Scheme 2: Preparation of 2-azido-2-deoxy-
D
-glucose
intermediate as glycosyl acceptor. Reagents and conditions: (a)
Allyl bromide, NaH, DMF, 0 °C to r t, 2 h; (b) Et3SiH, TfOH,
CH2Cl2, 0 °C, 3 h; (c) Ac2O, pyridine, r t, 2 h; (d) PdCl2,
CH3OH, r t, 3 h, 53%.
Tolyl
3-azido-2,4-di-O-benzyl-3-deoxy-1-thio-β-D-
fucopyranoside (6) was synthesized from literature
known
tolyl
3,4-O-isopropyl-1-thio-β-D-
fucopyranoside (9)29 in thirteen steps using functional
groups manipulation of the hydroxyl groups. In order
to achieve compound 6, initially it was decided to
benzylate the C-2 hydroxy group of compound 9

Tetrahedron
4
Table 1: Glycosylation of glycosyl donor 2 with glycosyl
activator 3 using different glycosylation activator.
O
BnO
HO
O
a, b, c, d, e, f, g
O
O
STol
STol
OH
Sl.
No.
1
Activator
Temp.
(°C)
−40
−15
−40
−40
0
Time
(min)
60
Yield
OBz
(%)a
9
10
NIS/HClO4-SiO2
NIS/HClO4-SiO2
NIS/TfOH
0
0
0
0
0
h, i, j
O
2
60
BnO
3
60
BnO
N3
k, l, m
4
5
DMTST
CH3OTf
300
300
O
STol
STol
OH
a: Acceptor fully recovered and 1,1-self coupled product of
donor obtained (20%).
OBn
HO
11
6
Scheme 3: Preparation of 3-azido-3,6-dideoxy-
D
-galactose
thioglycoside intermediate as glycosyl donor. Reagents and
conditions: (a) 2-naphthylmethyl (NAP) bromide, NaH, DMF, 0
°C to r t, 2 h; (b) 80% aq. AcOH, 80 °C, 2 h; (c) benzyl bromide,
5% NaOH, CH2Cl2, TBAB, r t, 3 h; (d) levulinic acid, DIC,
DMAP, CH2Cl2, r t, 5 h; (e) DDQ, CH2Cl2-H2O (5:1), r t, 6 h; (f)
benzoyl chloride, pyridine, r t, 2 h; (g) AcOH, NH2NH2·H2O,
CH3OH-CH2Cl2 (1:1), 0 °C-r t, 3 h, 45% in seven steps; (h)
Tf2O, pyridine, CH2Cl2, − 10 °C, 2 h; (i) NaNO2, DMF, 60 °C,
12 h; (j) 0.1 M CH3ONa, CH3OH, r t, 3 h, 58%; (k) (i) Bu2SnO,
CH3OH, 65 °C, 3 h; (ii) benzyl bromide, CsF, DMF, 65 °C, 6 h;
(l) Tf2O, pyridine, CH2Cl2, − 10 °C, 2 h; (m) NaN3, DMF, 60
°C, 8 h, 60% in three steps.
Stereoselective glycosylation of thioglycoside 2 with
compound 4 in the presence of a combination43 of NIS
and HClO4-SiO2 as thiophilic glycosylation activator
44
furnished disaccharide derivative 12 in 52% yield. Use
of other glycosylation activators such as DMTST,45
NIS-TfOH46 did not improve the yield of the reaction.
Spectroscopic data of compound 12 confirmed its
formation and the stereochemistry of the newly formed
glycosidic linkage. Compound 12 was subjected to a
series of functional group modifications involving (a)
one-pot de-O-acetylation and benzylation using benzyl
bromide in the presence of sodium hydroxide and
tetrabutylammonium bromide (TBAB);48 (b) one-pot
conversion of 4,6-O-benzylidene acetal into di-O-
acetate derivative using acetic anhydride in the
Having required functionalized monosaccharide
intermediates, attempts were made to couple them
using stereoselective glycosylations conditions.
Initially, thioglycoside 2 was allowed to glycosylate
with glycosyl acceptor 3 in the presence of a
49
presence of HClO4-SiO2 and (c) removal of allyl
group using palladium chloride34 to give disaccharide
acceptor 13 in 66% over all yield. The synthetic
strategy faced another challenge at this point because
combination43 of N-iodosuccinimide (NIS) and HClO4-
44
SiO2
as thiophilic glycosylation activator.
Unfortunately formation of glycosylation product was
not observed after several attempts using different
glycosylation promoters e.g. DMTST,45 NIS-TfOH,46
methyl triflate47 except the formation of 1,1-linked self
coupled disaccharide of compound 2 (~20%) (Scheme
4, Table 1). Presumably the presence of 2-N-
phthalimido group posed extra steric hindrance to the
incoming glycosyl donor towards C-3 hydroxyl group
of compound 3. Hence it was decided to use the
glycosyl acceptor 4 in which the N-phthaloyl group
was replaced by a smaller non-interactive azido group.
of the need of incorporation of a β-D-mannosidic
linkage in the molecule. In the recent past, a number of
seminal reports appeared in the literature for the
construction of the β-D-mannosidic linkage using
mannosyl donors, which include Crich’s two stage
activation of thiomannosides,50 intramolecular
aglycone delivery (IAD),51 specially designed glycosyl
donor and acceptor,52 application of thiourea ligand53
etc. However, none of these methods can be considered
as general approach. Recently, a convenient NIS-
TMSOTf mediated β-mannosylation has been
OBn
O
Ph
O
developed using
D-mannosyl thioglycosides having
O
AcO
HO
O
OPMP
AllO
SEt
+
remotely placed hydrogen bond mediating p-
methoxybenzyl (PMB) group.54 This elegant reaction
condition has been successfully applied for
incorporation of β-mannosidic linkage in this synthetic
scheme. Stereoselective 1,2-cis-glycosylation of
OAc
NPhth
2
3
a
No expected glycosylation product
compound 13 with D-mannosyl thioglycoside 5 in the
Ph
O
AcO
presence of a combination46, 54 of NIS and TMSOTf at
low temperature followed by removal of the PMB
group55 from the newly formed trisaccharide derivative
in one-pot by raising the temperature of the reaction
condition after glycosylation furnished trisaccharide
acceptor 14 in 68% yield. Use of other glycosylation
O
OAll
O
O
O
AllO
O
AcO
O
Ph
(20%)
Scheme 4: Attempted glycosylation between glycosyl donor 2
with acceptor 3. Reagents and conditions: NIS, HClO4-SiO2 or
NIS, TfOH or DMTST, CH2Cl2, −40 °C to −10 °C.

Tetrahedron
5
activators such as a combination43 of NIS and HClO4-
Ph
OBn
O
O
O
SiO2,44 DMTST45 did not furnish satisfactory yield of
the reaction. Spectral analysis of compound 14
confirmed its formation and the stereochemistry of the
O
AcO
AllO
+
SEt
a
OPMP
HO
OAc
N3
2
4
newly formed β-D-mannosidic linkage [signals at δ
4.67 (s, H-1C) and 3.23-3.18 (m, H-5C) in 1H NMR and
at δ 102.5 (JC1,H1 = 158 Hz; C-1C) in 13C NMR
spectra]. The JC1,H1 = 158 Hz value of the anomeric
carbon of the mannosidic moiety in the 1H coupled 13C
NMR spectrum unambiguously confirmed its
stereochemistry.56
OBn
O
Ph
O
O AcO
O
OAc
O
A
B
AllO
OPMP
N3
12
b, c, d
OBn
O
AcO
AcO
HO
Stereoselective 1,2-cis glycosylation of compound 14
with thioglycoside 6 in the presence of a combination43
of NIS and HClO4-SiO2 furnished the tetrasaccharide
derivative together with hemiacetal derivative of the
glycosyl donor. De-O-acetylated tetrasaccharide
derivative 15 was obtained in 70% yield by the
treatment of the product mixture with sodium
methoxide. Spectral analysis of compound 15
confirmed its formation and the stereochemistry of the
newly formed α-glycosidic linkage [signals at δ 5.33
(d, J = 3.5 Hz, H-1D), 4.85 (d, J = 7.5 Hz, H-1A), 4.69
O BnO
O
OBn
A
B
OPMP
N3
13
Ph
OBn
O
O
O
e
PMBO
STol
5
OBn
O
Ph
OBn
O
AcO
AcO
O
O
O BnO
O
B
A
C
HO
O
OPMP
N3
OBn
14
1
BnO
N3
(s, H-1C), 4.68 (d, J = 8.0 Hz, H-1B) in H NMR].
O
f
Formation of β-glycosidic product was not observed
under the reaction condition. It is noteworthy to
mention that, this is the first report for the
STol
OBn
6
g
BnO
N3
glycosylation using 3-azido-3-deoxy-D-fucopyranosyl
O
D
donor. Finally, compound 15 was treated with
thioacetic acid57 in presence of pyridine to transform
azido groups into acetamido group followed by
hydrogenolized under a positive pressure of hydrogen
over 20% Pd(OH)2-C (Pearlman’s catalyst)58 to furnish
compound 1 in 65% over all yield (Scheme 5; Table 2).
The spectroscopic analysis of compound 1 confirmed
the stereochemistry of the glycosidic linkages present
in it [signals at δ 5.13 (d, J = 3.5 Hz, H-1D), 5.00 (d, J
= 8.5 Hz, H-1B), 4.87 (br s, H-1C), 4.49 (d, J = 8.0 Hz,
OBn
Ph
OBn
O
HO
HO
O
O
BnO
O BnO
O
OBn
O
O
A
B
C
O
OPMP
N3
15
h, i
1
Scheme 5: Synthesis of tetrasaccharide
1 by sequential
glycosylations and functionalization of the intermediate
glycoside derivatives. Reagents and conditions: (a) NIS, HClO4-
SiO2, MS 4Å, CH2Cl2, −10 °C, 2 h, 52%; (b) benzyl bromide,
NaOH, DMF, TBAB, r t, 3 h; (c) acetic anhydride, HClO4-SiO2,
r t, 10 min; (d) PdCl2, CH3OH, r t, 3 h, 66% in three steps; (e)
NIS, TMSOTf, CH2Cl2, −40 °C, 3 h, then at 20 °C for 30 min,
68%; (f) NIS, HClO4-SiO2, CH2Cl2, −10 °C, 1 h; (g) 0.1 M
CH3ONa, CH3OH, r t, 3 h, 70% in two steps; (h) CH3COSH,
pyridine, r t, 12 h; (i) H2, 20% Pd(OH)2-C, CH3OH, r t, 24 h,
60% in two steps.
1
H-1A) in H NMR and at δ 102.9 (JC1,H1 = 158 Hz; C-
1A), 100.4 (JC1,H1 = 156 Hz; C-1C), 100.3 (JC1,H1 = 159
Hz; C-1B), 100.2 (JC1,H1 = 172 Hz; C-1D), in 13C NMR
spectra]. The stereochemistry of the anomeric carbons
of the monosaccharide moieties in compound 1 were
unambiguously confirmed from their JC1,H1 values.56
Table 2: Comparative yields of glycosylation reactions used in the synthetic Scheme 5.
Sl. No.
Donor
Acceptor
Activator
NIS/ HClO4-SiO2
NIS/ TfOH
Time (h)
Yield (%)
Temp (°C)
−10
1
2
3
4
5
6
7
8
2
2
5
3
3
6
1
5
52
50
40
56
68
40
70
40
2
2
2
5
5
5
6
6
4
4
4
13
13
13
14
14
−10
−5
−40
−40
−10
−10
0
DMTST
NIS/ HClO4-SiO2
NIS/ TMSOTf
DMTST
NIS/ HClO4-SiO2
DMTST

Tetrahedron
6
through a Sephadex LH-20 column using CH3OH-H2O
(3:1) as eluant to give pure compound 1 (175 mg,
60%). White powder; [α]D − 17 (c 1.0, H2O); 1H NMR
(500 MHz, D2O): δ 6.99-6.88 (m, 4 H, Ar-H), 5.13 (d,
J = 3.5 Hz, 1 H, H-1D), 5.00 (d, J = 8.5 Hz, 1 H, H-1B),
4.87 (br s, 1 H, H-1C), 4.49 (d, J = 8.0 Hz, 1 H, H-1A),
4.19-4.17 (m, 3 H, H-5D, H-3D, H-2C), 4.02 (t, J = 9.0
Hz, 1 H, H-2B), 3.88-3.78 (m, 5 H, H-6abB, 6abC, H-3B),
3.77-3.71 (m, 2 H, H-6aA,H-2D), 3.73 (s, 3 H, OCH3),
3.70-3.64 (m, 5 H, H-4D, H-3A, H-4B, H-3C, H-6bA),
3.58-3.52 (m, 2 H, H-4A, H-5C), 3.50-3.42 (m, 2 H, H-
5A, H-4C), 3.38 (t, J = 8.5 Hz, 2 H, H-2A, H-5B), 1.98
(s, 3 H, NHCOCH3), 1.96 (s, 3 H, NHCOCH3), 1.14 (d,
J = 6.5 Hz, 1 H, CCH3);13C NMR (125 MHz, D2O): δ
174.8 (NHCOCH3), 174.4 (NHCOCH3), 154.8-115.05
3. Conclusions
In summary, a concise synthetic strategy has been
developed for the synthesis of a tetrasaccharide related
to the repeating unit of the cell wall O-antigen of
Salmonella enterica O60 in very good yield. A
combination of NIS and HClO4-SiO2 has been used as
glycosylation activator. The rare sugar intermediate, 3-
amino-3-deoxy-
(6) has been prepared from
and used in the synthetic strategy for the construction
of the α-linked 3-amino-3-deoxy- -fucose moiety in
D
-fucosyl thioglycoside intermediate
D-fucose for the first time
D
the molecule. A recently developed PMB group
directed glycosylation condition has been used to
achieve excellent yield of β-mannosidic linkage.
Presence of the in situ removable PMB group in the
mannosyl thioglycoside intermediate (5) allowed
reducing the number of reaction steps.
(Ar-C), 102.9 (JC1,H1 = 158 Hz; C-1A), 100.4 (JC1,H1
=
156 Hz; C-1C), 100.3 (JC1,H1 = 159 Hz; C-1B), 100.2
(JC1,H1 = 172 Hz; C-1D), 84.3 (C-3A), 82.6 (C-3B), 80.7
(C-3C), 76.1 (C-5C), 75.6 (C-5B), 75.4 (C-5A), 72.7 (C-
2A), 70.6 (C-3D), 70.3 (C-4D), 68.4 (C-4A), 68.0 (C-4C),
67.2 (C-2C), 66.4 (C-2D), 65.9 (C-4B), 60.9 (C-6A), 60.6
(C-6B), 60.5 (C-6C), 55.8 (OCH3), 54.4 (C-2B), 51.1 (C-
5D), 22.0 (NHCOCH3), 21.9 (NHCOCH3), 15.3
(CCH3); HRMS (ESI) calcd. for C35H54N2O21
(838.3219): [M+H]+ 839.3297; found: 839.3290.
4. Experimental
General methods: All reactions were monitored by
thin layer chromatography over silica gel coated TLC
plates. The spots on TLC were visualized by warming
ceric sulphate (2% Ce(SO4)2 in 2N H2SO4) sprayed
plates in hot plate. Silica gel 230-400 mesh was used
for column chromatography. NMR spectra were
recorded on Bruker Avance 500 MHz using CDCl3 as
solvent and TMS as internal reference unless stated
otherwise. Chemical shift value is expressed in δ ppm.
The complete assignment of proton and carbon spectra
p-Methoxyphenyl 4-O-acetyl-6-O-benzyl-2-deoxy-2-
N-phthalimido-β-D-glucopyranoside (3): To
a
solution of compound 7 (2 g, 3.97 mmol) were added
levulinic acid (450 µL, 4.41 mmol), DIC (750 µL, 4.79
mmol) and DMAP (485 mg, 3.97 mmol) and the
reaction mixture was stirred at room temperature for 5
h. The reaction mixture was diluted with H2O (50 mL)
and extracted with CH2Cl2 (100 mL). The organic layer
was washed with 2 M HCl (50 mL), H2O (100 mL),
dried (Na2SO4) and concentrated. To a solution of the
crude product in CH2Cl2 (25 mL) were added Et3SiH
(3.82 mL, 23.96 mmol) at 0 °C followed by the
addition of TfOH (1.5 mL, 19.96 mmol) and the
reaction mixture was stirred at the same temperature
for 3 h. The reaction mixture was diluted with CH2Cl2
(50 mL), washed with saturated NaHCO3 solution
(2×50 mL), dried (Na2SO4), and concentrated. To the
solution of the crude product in pyridine (10 mL) was
added Ac2O (5 mL) and the reaction mixture was
stirred at room temperature for 2 h. The solvents were
removed and co-evaporated with toluene (2×25 mL)
under reduced pressure. To a solution of the acetylated
product in CH3OH-CH2Cl2 (20 mL; 1:1 v/v) were
added AcOH (2.5 mL), hydrazine monohydrate (950
µL, 19 mmol) and the reaction mixture was stirred at 0
°C for 8 h. The reaction mixture was quenched by
adding acetone (50 mL) and evaporated to dryness to
give the crude product which was purified over SiO2
using hexane-EtOAc (2:1) as eluant to give pure
was carried out by using a standard set of NMR
experiments, e.g. H NMR, 13C NMR, 13C DEPT 135,
1
2D COSY and 2D HSQC etc. ESI-MS were recorded
on a Thermo Scientific Orbitrap Velos Pro TM mass
spectrometer. Optical rotations were recorded in a
Jasco P-2000 spectrometer. Commercially available
grades of organic solvents of adequate purity are used
in all reactions. HClO4-SiO2 was prepared following
the reported method.44
p-Methoxyphenyl
(3-acetamido-3-deoxy-α-
D-
fucopyranosyl)-(1→3)-(β-
D
-mannopyranosyl)-
(1→3)-(β-
D
-glucopyranosyl)-(1→3)-2-acetamido-2-
-glucopyranoside (1): To a solution of
deoxy-β-
D
compound 15 (500 mg, 0.35 mmol) in pyridine (2 mL)
was added CH3COSH (0.5 mL) and the reaction
mixture was stirred at room temperature for 12 h. The
solvents were removed and co-evaporated with toluene
(3 x 20 mL) under reduced pressure and the crude
product was passed through a short pad of SiO2. To a
solution of the N-acetylated product in CH3OH (5 mL)
was added 20% Pd(OH)2-C (50 mg) and the reaction
mixture was allowed to stir at room temperature under
a positive pressure of H2 for 24 h. The reaction mixture
was filtered through a Celite bed, washed with
CH3OH-H2O (20 mL; 2:1 v/v) and concentrated under
reduced pressure. The deprotected product was passed
compound 3 (1.35 g, 62%). Colorless oil; [α]D − 11 (c
1
1.0, CHCl3); H NMR (500 MHz, CDCl3): δ 7.87-6.69
(m, 13 H, Ar-H), 5.73 (d, J = 8.0 Hz, 1 H, H-1), 5.07 (t,
J = 9.0 Hz, 1 H, H-4), 4.61 (d, J = 11.5 Hz, 1 H,
PhCH), 4.53 (d, J = 12.0 Hz, 1 H, PhCH), 4.48-4.47
(m, 2 H, H-2, H-3), 3.88-3.85 (m, 1 H, H-5), 3.73 (s, 3

Tetrahedron
7
H, OCH3), 3.70-3.64 (m, 2 H, H-6ab), 2.03 (s, 3 H,
The organic layer was washed with H2O (50 mL), dried
(Na2SO4) and concentrated under reduced pressure. A
solution of the crude product in 80% aq. AcOH (50
mL) was stirred at 80 °C for 2 h. The solvents were
removed under reduced pressure and co-evaporated
with toluene (3 x 20 mL). To a solution of the crude
product in CH2Cl2 (50 mL) were added 5% aq. NaOH
(10 mL), benzyl bromide (1.3 mL; 10.94 mmol) and
TBAB (50 mg) and the biphasic reaction mixture was
stirred vigorously at room temperature for 3 h. The
reaction mixture was diluted with H2O (50 mL) and
extracted with CH2Cl2 (50 mL). The organic layer was
washed with H2O (50 mL), dried (Na2SO4) and
concentrated under reduced pressure. To a solution of
the crude product in dry CH2Cl2 (25 mL) were
successively added levulinic acid (1.1 mL; 10.8 mmol),
DIC (1.5 g, 11.88 mmol) and DMAP (1 g, 8.2 mmol)
and the reaction mixture was stirred at room
temperature for 5 h. The reaction mixture was diluted
with H2O (50 mL) and extracted with CH2Cl2 (100
mL). The organic layer was washed with 2 M HCl (50
mL), H2O (100 mL), dried (Na2SO4) and concentrated.
To a solution of the crude product in CH2Cl2 (25 mL)
was added a solution of DDQ (2.3 g, 10.13 mmol) in
H2O (5 mL) and the biphasic reaction mixture was
stirred at room temperature for 6 h. The reaction
mixture was diluted with H2O (50 mL) and extracted
with CH2Cl2 (50 mL). The organic layer was washed
with H2O (50 mL), dried (Na2SO4) and concentrated.
To a solution of the crude product in pyridine (15 mL)
was added benzoyl chloride (1.2 mL, 10.33 mmol) at 0
°C and the reaction mixture was stirred at room
temperature for 2 h. The reaction mixture was
quenched by addition of aq. NH4Cl (10 mL), diluted
with H2O (50 mL) and extracted with CH2Cl2 (100
mL). The organic layer was successively washed with
2N HCl (50 mL), H2O (50 mL), dried (Na2SO4) and
concentrated. To a solution of the crude product in
CH3OH-CH2Cl2 (15 ml; 1:1 v/v) were added AcOH
(600 µL, 10.5 mmol) and NH2NH2·H2O (510 µL, 10.5
mmol) and the reaction mixture was stirred at room
temperature for 2 h. The reaction mixture was diluted
with CH2Cl2 (50 mL), successively washed with 2N
HCl (50 mL), H2O (50 mL), dried (Na2SO4) and
concentrated. The crude product was purified over
SiO2 using hexane-EtOAc (8:1) as eluant to give pure
compound 10 (2 g, 45%). A solution of compound 10
(2 g, 4.30 mmol) in dry CH2Cl2 (25 mL) was cooled to
−10 °C. To the cooled reaction mixture were added
pyridine (1 mL) and Tf2O (1.5 mL, 8.93 mmol) and it
was stirred at same temperature for 2 h. The solvents
were removed and co-evaporated with toluene (2 x 20
mL) under reduced pressure. To a solution of the crude
product in dry DMF (10 mL) was added NaNO2 (2 g,
29 mmol) and it was stirred at 60 °C for 12 h. The
reaction mixture was diluted with H2O (50 mL) and
extracted with CH2Cl2 (50 mL). The organic layer was
washed with water (50 mL), dried (Na2SO4) and
concentrated. A solution of the crude product in 0.1 M
CH3ONa in CH3OH (10 mL) was stirred at room
temperature for 3 h, neutralized with Amberlite IR-120
13
COCH3); C NMR (125 MHz, CDCl3): δ 170.9 (CO),
155.5-114.4 (Ar-C), 97.5 (C-1), 73.7 (C-4), 73.5
(CH2Ph), 72.8 (C-5), 70.4 (C-3), 68.8 (C-6), 57.1 (C-
2), 55.4 (OCH3), 20.8 (COCH3); HRMS (ESI) calcd.
for C30H29NO9 (547.1842): [M+H]+ 548.1920; found:
548.1930.
p-Methoxyphenyl 4-O-acetyl-2-azido-6-O-benzyl-2-
deoxy-β-D-glucopyranoside (4):
A
solution of
compound 8 (2 g, 5.0 mmol) in dry DMF (5 mL) was
cooled to 0 °C. To the cooled reaction mixture was
added NaH (60% oil coated; 450 mg) followed by allyl
bromide (0.8 mL, 9.2 mmol) and the reaction mixture
was allowed to stir at room temperature for 2 h. The
reaction mixture was quenched with aq. NH4Cl, diluted
with H2O (50 mL) and extracted with CH2Cl2 (50 mL).
The organic layer was washed with H2O (50 mL), dried
(Na2SO4) and concentrated under reduced pressure by
co-evaporating with toluene (2×20 mL). To the
solution of the crude in dry CH2Cl2 (25 mL) were
added triethylsilane (4.8 mL, 30 mmol) at 0 °C
followed by the addition of trifluoroacetic acid (1.75
mL, 22.8 mmol) and the reaction mixture was stirred at
the same temperature for 3 h. The reaction mixture was
diluted with CH2Cl2 (30 mL), washed with saturated
NaHCO3 solution (2×50 mL), dried (Na2SO4), and
concentrated. To the solution of the crude product in
pyridine (5 mL) was added Ac2O (5 mL) and the
reaction mixture was stirred at room temperature for 2
h. The solvents were removed and co-evaporated with
toluene (2×25 mL) under reduced pressure. To a
solution of the acetylated product in anhydrous CH3OH
(10 mL) was added PdCl2 (180 mg, 1.0 mmol) and the
reaction mixture was stirred at room temperature for 3
h. The reaction mixture was filtered through a Celite
bed and washed with CH3OH (50 mL) and
concentrated under reduced pressure to give the crude
product which was purified over SiO2 using hexane-
EtOAc (as eluant to give pure compound 4 (1.2 g,
1
53%). Colorless oil; [α]D − 7 (c 1.0, CHCl3); H NMR
(500 MHz, CDCl3): δ 7.30-6.76 (m, 9 H, Ar-H), 4.93
(t, J = 9.5 Hz, 1 H, H-4), 4.73 (d, J = 8.0 Hz, 1 H, H-1),
4.56 (d, J = 12.0 Hz, 1 H, PhCH), 4.50 (d, J = 12.0 Hz,
1 H, PhCH), 3.76 (s, 3 H, OCH3), 3.63-3.53 (m, 5 H,
H-2, H-3, H-5, H-6ab), 2.01 (s, 3 H, COCH3); 13C NMR
(125 MHz, CDCl3): δ 170.5 (CO), 155.7-114.5 (Ar-C),
101.4 (C-1), 73.7 (C-4), 73.6 (CH2Ph), 73.5 (C-3), 71.2
(C-2), 68.9 (C-6), 66.2 (C-5), 55.4 (OCH3), 20.8
(COCH3); HRMS (ESI) calcd. for C22H25N3O7
(443.1693): [M+H]+ 444.1771; found: 444.1780.
p-Methylphenyl 3-azido-2,4-di-O-benzyl-3-deoxy-1-
thio-β-D-fucopyranoside (6): A solution of compound
9 (3 g, 9.66 mmol) in dry DMF (10 mL) was cooled to
0 °C. To the cooled reaction mixture was added NaH
(60% oil coated; 500 mg) followed by NAP bromide
(2.2 g, 9.95 mmol) and the reaction mixture was
allowed to stir at room temperature for 2 h. The
reaction mixture was quenched with aq. NH4Cl, diluted
with H2O (50 mL) and extracted with CH2Cl2 (50 mL).

Tetrahedron
8
(H+) resin, filtered and concentrated. The crude product
was purified over SiO2 using hexane-EtOAc (2:1) as
eluant to give pure compound 11 (900 mg, 58%). To a
solution of compound 11 (900 mg, 2.49 mmol) in
CH3OH (30 mL) was added Bu2SnO (1.5 g, 6.02
mmol) and the reaction mixture was stirred at 65 °C for
3 h. The solvents were evaporated and co-evaporated
with toluene (3 x 20 mL) under reduced pressure. To a
solution of the crude product in dry DMF (10 mL)
were added benzyl bromide (1 mL, 8.42 mmol), CsF
(380 mg, 2.5 mmol) and the reaction mixture was
stirred at 65 °C for 6 h. The reaction mixture was
diluted with H2O (50 mL) and extracted with EtOAc
(50 mL). The organic layer was successively washed
with 2N HCl (50 mL) and H2O (50 mL), dried
(Na2SO4) and concentrated. A solution of the crude
product in dry CH2Cl2 (15 mL) was cooled to −10 °C.
To the cooled reaction mixture were added pyridine
(0.5 mL) and Tf2O (850 µL, 5.06 mmol) and it was
stirred at same temperature for 2 h. The solvents were
removed and co-evaporated with toluene (2 x 20 mL)
under reduced pressure. To a solution of the crude
product in dry DMF (5 mL) was added NaN3 (1.5 g, 23
mmol) and it was stirred at 60 °C for 12 h. The reaction
mixture was diluted with H2O (50 mL) and extracted
with CH2Cl2 (50 mL). The organic layer was washed
with water (50 mL), dried (Na2SO4) and concentrated.
The crude product was purified over SiO2 using
hexane-EtOAc (5:1) to give pure compound 6 (710 mg,
60%). Colorless oil; [α]D + 26 (c 1.0, CHCl3); 1H NMR
(500 MHz, CDCl3): δ 7.43-7.00 (m, 14 H, Ar-H), 4.92
(d, J = 10.5 Hz, 2 H, PhCH2), 4.67 (d, J = 10.0 Hz, 1
H, PhCH), 4.62 (d, J = 11.5 Hz, 1 H, PhCH), 4.53 (d, J
= 9.5 Hz, 1 H, H-1), 3.84 (t, J = 9.5 Hz, 1 H, H-2),
3.55-3.50 (m, 3 H, H-3, H-4, H-5), 2.31 (s, 3 H, CH3),
1.24 (d, J = 6.5 Hz, 3 H, CCH3);13C NMR (125 MHz,
CDCl3): δ 138.0-127.7 (Ar-C), 88.2 (C-1), 77.9 (C-4),
77.2 (C-2), 75.3 (CH2Ph), 75.1 (CH2Ph), 75.0 (C-3),
67.6 (C-5), 21.1 (CH3), 17.1 (CCH3); HRMS (ESI)
calcd. for C27H29N3O3S (475.1930): [M+H]+ 476.2008;
found: 476.2000.
PhCH), 5.24 (d, J = 17.5 Hz, 1 H, C=CH), 5.13 (d, J =
9.5 Hz, 1 H, C=CH), 4.93-4.87 (m, 2 H, H-2B, H-4A),
4.76, 4.71 (2 d, J = 8.0 Hz each, 2 H, H-1A, H-1B), 4.51
(s, 2 H, CH2Ph), 4.31 (s, 2 H, H-6aB, OCH), 4.10 (d, J
= 7.5 Hz, 1 H, OCH), 3.82 (s, 3 H, OCH3), 3.76-3.54
(m, 7 H, H-2A, H-3A, H-3B, H-4B, H-5A, H-6abA, H-6B),
3.39 (s, 1 H, H-5B), 2.15 (s, 3 H, COCH3), 1.95 (s, 3 H,
COCH3); 13C NMR (125 MHz, CDCl3): δ 169.0
(COCH3), 168.9 (COCH3), 155.7-114.6 (Ar-C), 102.0
(C-1B), 101.5 (C-1A), 101.2 (PhCH), 81.2 (C-3A), 79.7
(C-3B), 78.6 (C-2A), 73.7 (C-4B), 73.6 (CH2Ph), 73.1
(OCH2), 72.8 (C-2B), 69.4 (C-4A), 69.3 (C-6B), 68.6
(C-6A), 66.1 (C-5B), 65.9 (C-5A), 55.5 (OCH3), 20.81
(COCH3), 20.8 (COCH3); HRMS (ESI) calcd. for
C40H45N3O13 (775.2952): [M+H]+ 776.3030; found:
776.3022.
p-Methoxyphenyl (4,6-di-O-acetyl-2-O-benzyl-β-
D-
glucopyranosyl)-(1→3)-2-azido-4,6-di-O-benzyl-2-
deoxy-β-D-glucopyranoside (13): To a solution of
compound 12 (1 g, 1.29 mmol) in DMF (10 mL) were
added powdered NaOH (200 mg, 5.0 mmol), benzyl
bromide (400 µL, 3.36 mmol) and TBAB (50 mg) and
the reaction mixture was stirred at room temperature
for 3 h. The reaction mixture was diluted with H2O (50
mL) and extracted with CH2Cl2 (50 mL). The organic
layer was washed with H2O (50 mL), dried (Na2SO4)
and concentrated. To a solution of the crude product in
acetic anhydride (5 mL) was added HClO4-SiO2 (50
mg) and it was stirred at room temperature for 10 min.
The reaction mixture was filtered and concentrated
under reduced pressure. To a solution of the crude
product in CH3OH (10 mL) was added PdCl2 (25 mg,
0.14 mmol) and it was stirred at room temperature for
3 h. The reaction mixture was filtered and concentrated
under reduced pressure. The crude product was
purified over SiO2 using hexane-EtOAc (5:1) as eluant
to give pure compound 13 (705 mg, 66%). Colorless
1
oil; [α]D − 18 (c 1.0, CHCl3); H NMR (500 MHz,
CDCl3): δ 7.38-6.78 (m, 19 H, Ar-H), 5.02 (d, J = 11.0
Hz, 1 H, PhCH), 4.99 (d, J = 12.0 Hz, 1 H, PhCH),
4.97 (d, J = 9.0 Hz, 1 H, H-1B), 4.94 (t, J = 10.0 Hz, 1
H, H-4B), 4.77 (d, J = 11.5 Hz, 1 H, PhCH), 4.72 (d, J
= 8.0 Hz, 1 H, H-1A), 4.59 (d, J = 12.5 Hz, 1 H,
PhCH), 4.53 (d, J = 12.0 Hz, 1 H, PhCH), 4.47 (d, J =
12.0 Hz, 1 H, PhCH), 4.24 (dd, J = 12.5 Hz, 4.5 Hz, 1
H, H-6aB), 4.01 (dd, J = 12.0 Hz, 2.0 Hz, 1 H, H-6bB),
3.76 (s, 3 H, OCH3), 3.75 -3.71 (m, 3 H, H-3A, H-3B,
H-6aA), 3.69 (dd, J = 11.0 Hz, 5.0 Hz, 1 H, H-6aB), 3.63
(t, J = 8.5 Hz, 1 H, H-2A), 3.62 (t, J = 7.5 Hz, 1 H, H-
4A), 3.58-3.54 (m, 1 H, H-5B), 3.51-3.50 (m, 1 H, H-
5A), 3.33 (t, J = 8.0 Hz, 1 H, H-2B); 13C NMR (125
MHz, CDCl3): δ 170.4 (COCH3), 170.0 (COCH3),
155.6-114.6 (Ar-C), 102.6 (C-1B), 101.7 (C-1A), 81.9
(C-2B), 80.5 (C-3A), 75.6 (C-2A), 75.0 (CH2Ph), 74.9
(H-5A), 74.8 (CH2Ph), 74.2 (C-3B), 73.5 (CH2Ph), 71.8
(C-5B), 69.9 (C-4B), 68.6 (C-6A), 66.4 (C-4A), 62.2 (C-
6B), 55.5 (OCH3), 20.8 (COCH3), 20.6 (COCH3);
HRMS (ESI) calcd. for C44H49N3O13 (827.3265):
[M+H]+ 828.3343; found: 828.3336.
p-Methoxyphenyl
benzylidene-β- -glucopyranosyl)-(1→3)-4-O-acetyl-
2-azido-6-O-benzyl-2-deoxy-β- -glucopyranoside
(2-O-acetyl-3-O-allyl-4,6-O-
D
D
(12): To a solution of compound 2 (1.3 g, 3.29 mmol)
and compound 4 (1.2 g, 2.70 mmol) in anhydrous
CH2Cl2 (20 mL) was added MS 4Å (1 g) and the
reaction mixture was cooled to − 10 °C under argon.
To the cooled reaction mixture were added NIS (750 g,
3.33 mmol) and HClO4-SiO2 (50 mg) and it was stirred
at same temperature for 2 h. The reaction mixture was
filtered through a Celite bed, washed with CH2Cl2 (50
mL). The combined organic layer was successively
washed with 5% Na2S2O3 (100 mL), satd. NaHCO3 (50
mL), H2O (50 mL), dried (Na2SO4) and concentrated.
The crude product was purified over SiO2 using
hexane-EtOAc (4:1) as eluant to give pure compound
12 (1.1 g, 52%). Colorless oil; [α]D − 15 (c 1.0,
1
CHCl3); H NMR (500 MHz, CDCl3): δ 7.44-6.77 (m,
14 H, Ar-H), 5.82-5.81 (m, 1 H, CH), 5.51 (s, 1 H,

Tetrahedron
9
p-Methoxyphenyl (4,6-O-benzylidene-2-O-benzyl-β-
-mannopyranosyl)-(1→3)-(4,6-di-O-acetyl-2-O-
benzyl-β- -glucopyranosyl)-(1→3)-2-azido-4,6-di-O-
benzyl-2-deoxy-β- -glucopyranoside (14): To
temperature for 1 h. The reaction mixture was filtered
through a Celite bed, washed with CH2Cl2 (50 mL).
The combined organic layer was successively washed
with 5% Na2S2O3 (50 mL), satd. NaHCO3 (50 mL),
H2O (50 mL), dried (Na2SO4) and concentrated. A
solution of the crude product in 0.1 M CH3ONa in
CH3OH (10 mL) was stirred at room temperature for 3
h, neutralized with Amberlite IR-120 (H+) resin,
filtered and concentrated. The crude product was
purified over SiO2 using hexane-EtOAc (6:1) as eluant
D
D
D
a
solution of compound 13 (700 mg, 0.85 mmol) and
compound 5 (600 mg, 1.02 mmol) in anhydrous
CH2Cl2 (5 mL) was added MS 4Å (0.5 g) and the
reaction mixture was cooled to − 40 °C under argon.
To the cooled reaction mixture were added NIS (230
mg, 1.02 mmol) and TMSOTf (20 µL) and it was
stirred at same temperature for 3 h. After consumption
of the starting materials (TLC), the reaction mixture
was allowed to stir at 20 °C for 30 min. The reaction
mixture was filtered through a Celite bed, washed with
CH2Cl2 (50 mL). The combined organic layer was
successively washed with 5% Na2S2O3 (50 mL), satd.
NaHCO3 (50 mL), H2O (50 mL), dried (Na2SO4) and
concentrated. The crude product was purified over
SiO2 using hexane-EtOAc (6:1) as eluant to give pure
to give pure compound 15 (560 mg, 70%). Colorless
1
oil; [α]D − 10 (c 1.0, CHCl3); H NMR (500 MHz,
CDCl3): δ 7.39-6.73 (m, 39 H, Ar-H), 5.33 (d, J = 3.5
Hz, 1 H, H-1D), 5.23 (s, 1 H, PhCH), 4.92 (d, J = 11.0
Hz, 1 H, PhCH), 4.91 (d, J = 10.5 Hz, 1 H, PhCH),
4.85 (d, J = 7.5 Hz, 1 H, H-1A), 4.83 (d, J = 12.5 Hz, 1
H, PhCH), 4.76 (d, J = 11.5 Hz, 1 H, PhCH), 4.69 (s, 1
H, H-1C), 4.68 (d, J = 8.0 Hz, 1 H, H-1B), 4.64 (d, J =
11.5 Hz, 1 H, PhCH), 4.62 (d, J = 12.0 Hz, 1 H,
PhCH), 4.56 (d, J = 12.0 Hz, 1 H, PhCH), 4.49 (d, J =
12.0 Hz, 2 H, 2 PhCH), 4.44 (d, J = 11.0 Hz, 1 H,
PhCH), 4.43 (d, J = 12.0 Hz, 1 H, PhCH), 4.20 (d, J =
12.0 Hz, 1 H, PhCH), 4.18 (dd, J = 9.0 Hz, 3.0 Hz, 1
H, H-6aC), 4.10 (t, J = 9.0 Hz, 1 H, H-4D), 3.76-3.72
(m, 4 H, H-2D, H-3D, H-4A, H-6bC), 3.70 (s, 3 H,
OCH3), 3.71-3.65 (m, 4 H, H-4C, H-6aA, H-6abB), 3.59-
3.54 (m, 4 H, H-2B, H-3A, H-3C, H-5B), 3.47-3.39 (m, 3
H, H-2C, H-5A, H-6bA), 3.37-3.40 (m, 2 H, H-3B, H-5D),
3.29 (t, J = 8.5 Hz, 1 H, H-2A), 3.51-3.21 (m, 2 H, H-
4B, H-5C), 0.90 (d, J = 6.5 Hz, 1 H, CCH3); 13C NMR
(125 MHz, CDCl3): δ 155.7-114.6 (Ar-C), 103.1 (2C,
C-1A, C-1C), 102.2 (PhCH), 101.8 (C-1B), 96.5 (C-1D),
86.5 (C-3A), 81.0 (C-2A), 80.4 (C-4C), 78.6 (C-4D),
78.4 (C-4A), 78.2 (C-4B), 75.7 (PhCH2), 75.6 (PhCH2),
75.3 (2C, 2 PhCH2), 75.2 (H-3C), 75.1 (H-2C), 75.0 (H-
5C), 74.6 (C-3D), 73.8 (C-2D), 73.5 (CH2Ph), 70.4
(CH2Ph), 69.8 (C-3B), 68.5 (C-6A), 68.2 (C-6C), 67.4
(C-5D), 66.3 (C-5A), 66.2 (C-2B), 62.7 (C-6B), 60.8 (C-
5B), 55.5 (OCH3),16.6 (CCH3); HRMS (ESI) calcd. for
C80H86N6O19 (1434.5948): [M+H]+ 1435.6026; found:
1435.6017.
compound 14 (675 mg, 68%). Colorless oil; [α]D − 6 (c
1
1.0, CHCl3); H NMR (500 MHz, CDCl3): δ 7.42-6.78
(m, 29 H, Ar-H), 5.44 (s, 1 H, PhCH), 5.06 (d, J =
11.0 Hz, 1 H, PhCH), 5.01 (t, J = 8.5 Hz, 1 H, H-4B),
4.97 (d, J = 12.0 Hz, 1 H, PhCH), 4.97 (d, J = 8.0 Hz, 1
H, H-1A), 4.88 (d, J = 11.5 Hz, 1 H, PhCH), 4.74 (d, J
= 8.0 Hz, 1 H, H-1B), 4.67 (s, 1 H, H-1C), 4.62 (d, J =
11.5 Hz, 1 H, PhCH), 4.59 (d, J = 11.5 Hz, 1 H,
PhCH), 4.53 (d, J = 12.0 Hz, 1 H, PhCH), 4.49 (d, J =
11.0 Hz, 1 H, PhCH), 4.34 (d, J = 11.0 Hz, 1 H,
PhCH), 4.28 (dd, J = 10.0 Hz, 5.5 Hz, 1 H, H-6aC),
4.26 (dd, J = 16.5 Hz, 4.5 Hz, 1 H, H-6aB), 4.05 (dd, J
= 12.0 Hz, 2.0 Hz, 1 H, H-6bB), 3.79 (t, J = 10.0 Hz, 1
H, H-3B), 3.76 (s, 3 H, OCH3), 3.75 (t, J = 9.0 Hz, 1 H,
H-3A), 3.72-3.60 (m, 7 H, H-2B, H-2C, H-4A, H-5B, H-
6bA, H-6abC), 3.52-3.48 (m, 1 H, H-5A), 3.44 (t, J = 9.0
Hz, 1 H, H-2A), 3.41 (dd, J = 9.0 Hz, 2.0 Hz, 1 H, H-
3C), 3.34 (d, J = 3.5 Hz, 1 H, H-4C), 3.23- 3.18 (m, 1 H,
H-5C), 2.00 (s, 3 H, COCH3), 1.95 (s, 3 H, COCH3);
13C NMR (125 MHz, CDCl3): δ 170.4 (COCH3), 169.3
(COCH3), 155.7-114.6 (Ar-C), 103.1 (JC1,H1 = 159 Hz;
C-1A), 102.5 (JC1,H1 = 158 Hz; C-1C), 101.9 (PhCH),
101.7 (JC1,H1 = 156 Hz; C-1B), 82.5 (C-3C), 80.7 (C-
3B), 79.7 (C-3A), 79.4 (C-4A), 78.4 (C-4C), 75.7 (C-2B),
75.5 (CH2Ph), 75.4 (CH2Ph), 75.0 (C-5A), 74.9
(CH2Ph), 73.5 (CH2Ph), 71.7 (C-2C), 70.6 (C-2A), 68.6
(2C, C-6A, C-6C), 68.1 (C-4B), 66.9 (C-5C), 66.4 (C-
5B), 62.3 (C-6B), 55.5 (OCH3), 20.8 (COCH3), 20.7
(COCH3); HRMS (ESI) calcd. for C64H69N3O18
(1167.4576): [M+H]+ 1168.4654; found: 1168.4664.
Acknowledgements
A.G. and P.S. thank CSIR, New Delhi for providing
Senior Research Fellowship. The work is supported by
SERB, New Delhi (Project No. CRG/2019/000352
dated 23.01.2020) (AKM) and Bose Institute.
Conflicts of Interest
There are no conflicts to declare.
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Concise synthesis of a tetrasaccharide related to the
repeating unit of the cell wall O-antigen of Salmonella
enterica O60
Arin Gucchait, Pradip Shit and Anup Kumar Misra*
Bose Institute, Molecular Medicine Division, P-1/12, C.I.T. Scheme VII M, Kolkata 700054
Highlights
Tetrasaccharide related to the cell wall of Salmonella enterica O60 was synthesized.
•
•
•
•
•
A combination of NIS and HClO4-SiO2 has been used as glycosylation activator.
3-Amino-3-deoxy-D-fucosyl thioglycoside has been prepared from D-fucose.
PMB group directed glycosylation has been used to achieve
β-mannosidic linkage.
The number of reaction steps was reduced by using in situ removable PMB group.
*Corresponding author. Tel.: (+91)(-33) 25693240; Fax: (+91)(-33) 2355-3886; E-mail: akmisra69@gmail.com

Conflicts of Interest
There are no conflicts to declare.