Synthesis of a pentasaccharide repeating unit corresponding to the cell wall O-antigen of Escherichia coli O59 using iterative glycosylations in one pot

Article abstract of DOI:10.1016/j.tet.2016.06.025

Synthesis of a pentasaccharide repeating unit corresponding to the cell wall O-antigen of Escherichia coli O59 has been achieved by orthogonal glycosylation and two iterative glycosylations in one pot. Synthesis of a β-D-mannosidic linkage present in the molecule has been successfully achieved with satisfactory yield by the activation of thioglycoside with a combination of 1-benzenesulfinyl piperidine (BSP) and triflic anhydride (Tf₂O). The α-D-glucosaminyl moiety was achieved in moderate yield from the α-D-mannosyl moiety by azidolysis of C-2 hydroxy group with inversion of configuration. TEMPO mediated selective oxidation of the primary hydroxyl group has been carried out at the late stage of the synthetic strategy.

Full text of DOI:10.1016/j.tet.2016.06.025

Tetrahedron 72 (2016) 4435e4441
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Synthesis of a pentasaccharide repeating unit corresponding to the
cell wall O-antigen of Escherichia coli O59 using iterative
glycosylations in one pot
*
Anshupriya Si, Anup Kumar Misra
Bose Institute, Molecular Medicine Division, P-1/12, C.I.T. Scheme VII M, Kolkata 700054, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Synthesis of a pentasaccharide repeating unit corresponding to the cell wall O-antigen of Escherichia coli
Received 13 April 2016
Received in revised form 25 May 2016
Accepted 7 June 2016
O59 has been achieved by orthogonal glycosylation and two iterative glycosylations in one pot. Synthesis
of a
yield by the activation of thioglycoside with a combination of 1-benzenesulfinyl piperidine (BSP) and
triflic anhydride (Tf2O). The -glucosaminyl moiety was achieved in moderate yield from the
b-D-mannosidic linkage present in the molecule has been successfully achieved with satisfactory
Available online 8 June 2016
a-
D
a-D-
mannosyl moiety by azidolysis of C-2 hydroxy group with inversion of configuration. TEMPO mediated
selective oxidation of the primary hydroxyl group has been carried out at the late stage of the synthetic
strategy.
Keywords:
Pentasaccharide
One-pot
Iterative glycosylation
Orthogonal glycosylation
Ó 2016 Elsevier Ltd. All rights reserved.
b-D-mannoside
TEMPO oxidation
1. Introduction
bacterial strains are serious concern. As an alternative approach,
development of glycoprotein conjugate vaccines12e15 related to the
Diarrheal outbreaks and gastrointestinal disorders are serious
health concern all over the world and particularly in the developing
countries because of the lack of adequate sanitation.1 Consump-
tions of the contaminated food and water are the major causes of
bacterial infections leading to the diarrheal outbreaks.2 Among
several bacteria responsible for the diarrheal infections, Escherichia
coli, Salmonella, Shigella are predominant.3e5 Even though, E. coli
strains exist as commensal organism in the human gastrointestinal
flora, they often acquire virulence and cause several infections such
as diarrhea,6 septicemia7 and urinary tract infections.8 E. coli strains
responsible for the diarrheal infections are further divided into
several subgroups based on their mode of pathogenic actions, such
as (a) enteropathogenic E. coli (EPEC); (b) enteroinvasive E. coli
(EIEC); (c) enterohemorrhagic E. coli (EHEC); (d) enteroaggregative
E. coli (EAEC); (e) enterotoxigenic E. coli (ETEC) etc.9 E. coli (E. coli)
O59 belongs to EHEC strains and responsible for several diarrheal
outbreaks in Europe.10 Initially this strain was isolated from a pa-
tient suffering from bloody diarrhea and hemolytic uremic syn-
drome.11 Although several therapeutics has been developed for
controlling diarrheal infections, emergence of multidrug resistant
cell wall polysaccharides have been found useful to eradicate the
bacterial infections because of the fact that capsular poly-
saccharides (CPC) or lipopolysaccharides (LPSs) usually construct
the bacterial cell and are responsible for the host-bacterial in-
teractions. It is thus of immense importance to develop glyco-
conjugates of bacterial cell wall CPS or LPS for understanding their
biological roles and also for manipulating or controlling of their
survival ability. In this context, synthesis of oligosaccharides related
to bacterial polysaccharides or lipopolysaccharides remains always
important for unveiling the above facts. Recently, the structure of
the repeating unit of the cell wall O-antigen of E. coli O59 has been
reported by Widmalm and co-workers,16 which is a penta-
saccharide composed of one
a-N-acetyl-
D-glucosamine, two a-D-
mannose, one -mannose and one a-D
b
-
D
-galacturonic acid in which
3 and 4-hydroxy groups are acetylated. In continuation of an on-
going program for the preparation of glycoprotein conjugates,
synthesis of a pentasaccharide containing a
a-D-galacturonic acid
and a -mannose in its native form corresponding to the cell wall
b-D
O-antigen of E. coli O59 has been carried out in minimum number
of steps by applying modern synthetic methodologies (Fig. 1). A
concise chemical synthetic strategy for the synthesis of a penta-
saccharide as its 2-aminoethyl glycoside is reported herein apply-
ing two iterative stereoselective glycosylations in one pot.
* Corresponding author. Tel.: þ91 (33) 25693240; fax: þ91 (33) 2355 3886;
e-mail address: akmisra69@gmail.com (A.K. Misra).
http://dx.doi.org/10.1016/j.tet.2016.06.025
0040-4020/Ó 2016 Elsevier Ltd. All rights reserved.

4436
A. Si, A.K. Misra / Tetrahedron 72 (2016) 4435e4441
a-D-mannopyranoside (5) in 69% yield. A small amount of 3-O-
benzylated product (10%) and 2,3-di-O-benzylated product (5%)
were also formed under the reaction condition, which were sep-
arated by chromatography (Scheme 1).
OH
O
HO
HO
OH
O
HO
HO
O
AcHN
A
NH2
B
D
O
O
HO
HO
O
HO
O
O
AcO
AcO
E
C
HO
HO
O
OH
O
OH
O
1
OH
O
OAc
O
Ph
HO
HO
HO
O
O
OAc
O
Ph
a, b
NHCbz
OH
O
OBn
O
HO
Ph
NHCbz
OBn
OBn
O
Ph
O
O
O
O
HO
O
O
O
NHCbz
HO
AllO
2
3
O
3
OPMP
SEt
5
6
OBn
O
Ph
Ph
O
O
AcO
AcO
OBn
O
Ph
c
O
O
O
O
O
O
HO
HO
SEt
HO
OBn
OPMP
OPMP
SEt
4
5
8
7
Scheme 1. Reagents: (a) PhCH(OCH3)2, I2, CH3CN, room temperature, 1.5 h; (b) (i)
CH3C(OC2H5)3; Camphorsulfonic acid, DMF, room temperature, 1 h; (ii) H2O, room
temperature, 1 h, 74% in two steps; (c) benzyl bromide, 10% NaOH, CH2Cl2, TBAB, room
temperature, 30 h, 69%.
Fig. 1. Structure of the synthesized pentasaccharide and its synthetic intermediates.
2. Results and discussion
The target pentasaccharide
aminoethyl glycoside keeping the O-acetyl groups attached to the
-galacturonic acid moiety at the non-reducing end. The penta-
1 was synthesized as its 2-
Although several stereoselective glycosylation techniques have
been developed in the recent past, only a few reaction conditions
D
can be used for the preparation of b-D-mannosidic linkage. In the
present synthetic strategy, following Crich et al.25 a combination
saccharide derivative 16 was synthesized by the stereoselective
glycosylation of a number of suitably protected monosaccharide
intermediates 3, 5, 6,17 718 and 8,19 which were prepared from the
naturally available reducing sugars applying a number of reaction
of 1-benzenesulfinyl piperidine (BSP) and triflic anhydride (Tf2O)
has been used as glycosyl promoter for the activation of
nosyl thioglycoside donor towards the formation of
nosidic linkage. Activation of the
using a combination of BSP and Tf2O and stereoselective cou-
pling with compound 3 resulted in the formation of -man-
nosidic disaccharide derivative 9 in 55% yield. Although BSP
mediated activation of -mannosyl thioglycoside generally fur-
nishes -mannosidic linkage, in this case a small amount (20%)
of -mannosylated disaccharide was also formed, which was
D
-man-
b
-
D
-man-
conditions reported earlier.17e19 Presence of a
b
-D
-mannosidic
D
-mannose thioglycoside 617
linkage and an a-D-glucosamine linkage makes the synthesis of the
target compound quite challenging because of the difficulties as-
sociated with the formation of these two glycosyl linkages. The
b-D
noteworthy features of the synthetic strategy include (a)
mannosidic glycoside formation, (b) -glucosaminyl glycoside
formation by inversion of C-2 position of -mannoside, (c) two
b-D-
D
a-D
b-D
a-D
a-D
step iterative glycosylations in one pot using orthogonal glycosyl-
ation, (d) use of p-methoxyphenyl (PMP) group as oxidative re-
movable temporary anomeric protecting group, (e) use of
perchloric acid on silica (HClO4eSiO2) as a non-corrosive solid acid
substitute of protonic acid for the activation of glycosyl tri-
chloroacetimidate and thioglycoside derivatives, (f) direct trans-
formation of azido group into acetamido group by the treatment
with thioacetic acid, (g) selective removal of benzyl ethers in the
presence of benzylidene acetal under catalytic hydrogenation
condition, (h) TEMPO mediated selective oxidation of the primary
hydroxyl group into carboxylic acid at the late stage of the syn-
thesis, (i) anomeric 2-aminoethyl group at the reducing end can
serve as a readily available amino group for the conjugation of the
synthetic pentasaccharide with a suitable protein to make glyco-
conjugate derivative.
separated by the chromatography. The rest of the glycosyl donor
and acceptor got decomposed during the reaction. Formation of
compound 9 was confirmed from NMR spectral analysis [signals
at
d d 99.7
4.86 (br s, H-1A), 4.74 (br s, H-1B) in the 1H NMR and at
(JC-1/H-1¼155.0 Hz, C-1B), 98.7 (JC-1/H-1¼170.0 Hz, C-1A) in the 13C
NMR spectra]. Appearance of JC1/H1 values 170 Hz (a-D-mannosyl)
and 155 Hz (b-D
-mannosyl) in the gated 1H coupled 13C NMR
spectrum confirmed26 the formation of compound 9. Removal of
O-acetyl group of compound 9 using sodium methoxide27 fur-
nished compound 10 in 92% yield. The axially oriented 2-hydroxy
group (
D
-mannose moiety) of compound 10 was converted to
-glucosamine moiety) to give
equatorially oriented azido group (
D
compound 11 in 37% yield by the formation of triflate derivative
using triflic anhydride and pyridine followed by SN2 substitution
of triflate group by azido group using sodium azide at elevated
temperature. The displacement of triflate group with the azide
group suffered from the formation of elimination product causing
lower yield of the required compound. In earlier reports, this fact
has been well explained considering the unfavorable torsional
strain and dipolar interactions in the transition state of the
reaction.28e31 Although, earlier such conversion in a disaccharide
was reported with 13% yield,31 in our case the yield was better
(37%) possibly due to the presence of a 2-(N-benzyloxycarbonyl)
aminoethyl group at the anomeric position instead of a sugar
moiety. Formation of compound 11 was confirmed from its NMR
Treatment of 2-(N-benzyloxycarbonyl)aminoethyl
a-D-man-
nopyranoside (2)20 with benzaldehyde dimethylacetal in the
presence of molecular iodine21 furnished 4,6-O-benzylidene acetal
derivative, which was selectively 2-O-acetylated to give 2-(N-
benzyloxycarbonyl)aminoethyl 2-O-acetyl-4,6-O-benzylidene-a-D-
mannopyranoside (3) in 74% yield by the treatment with triethyl
orthoacetate in the presence of camphorsulfonic acid via the for-
mation of an orthoester intermediate followed by its hydrolysis.22
In another experiment, p-methoxyphenyl 4,6-O-benzylidene-a-D-
mannopyranoside (4)23 was treated with benzyl bromide and 10%
sodium hydroxide solution in the presence of tetrabutylammo-
nium bromide (TBAB) in a biphasic phase transfer reaction con-
dition24 to give p-methoxyphenyl 2-O-benzyl-4,6-O-benzylidene-
spectral analysis [signals at
d
5.02 (d, J¼3.5 Hz, H-1A), 4.85 (br s,
H-1B) in the 1H NMR and at
d
102.2 (C-1B), 98.8 (C-1A) in the 13C
NMR spectra]. Removal of the allyl group from compound 11

A. Si, A.K. Misra / Tetrahedron 72 (2016) 4435e4441
4437
using palladium chloride32 furnished disaccharide acceptor 12 in
76% yield (Scheme 2).
trisaccharide thioglycoside derivative 15, which was not isolated
from the reaction. The disaccharide acceptor 12 was then added to
the reaction mixture and allowed to react with the in situ generated
trisaccharide thioglycoside derivative 15 by the addition of NIS to
the reaction mixture to furnish pentasaccharide derivative 16 in
64% in two steps. Formation of compound 16 was supported by its
3
6
+
a
NMR spectral analysis [signals at
s, H-1D), 5.28 (br s, H-1C), 4.90 (d, J¼3.0 Hz, H-1A), 4.85 (br s, H-1B)
in 1H NMR and at
102.1 (C-1B), 99.3 (2 C, C-1D, C-1C), 98.8 (C-1A),
d
5.45 (d, J¼3.5 Hz, H-1E), 5.37 (br
d
OBn
O
Ph
O
97.4 (C-1E) in the 13C NMR spectra]. Compound 16 was subjected to
a series of functional group transformations. The azido group of
compound 16 was converted to the acetamido group by the treat-
ment with thioacetic acid in pyridine.40 Partial hydrogenolysis of
the pentasaccharide derivative over Pearlman’s catalyst in a limited
period of time resulted in the selective removal of benzyl ethers
keeping the benzylidene acetals unaffected.41 The partially hydro-
genolized pentasaccharide was subjected to the TEMPO mediated
selective oxidation42,43 of the primary hydroxyl group using
TEMPO, sodium hypochlorite and sodium chlorite in a biphasic
reaction condition. Finally, deprotection of Cbz group and benzy-
lidene acetals under catalytic hydrogenation44 using hydrogen over
Pearlman’s catalyst furnished compound 1 in 50% over all yield.
NMR spectroscopic analysis of compound 1 confirmed its formation
O
NHCbz
O
B
A
AllO
O
O
O
OR
O
Ph
b
9: R = Ac
10:
R = H
c, d
OBn
O
Ph
O
O
RO
O
N3
NHCbz
B
O
A
O
O
O
Ph
[signals at
d
5.13 (d, J¼4.0 Hz, H-1E), 4.97 (br s, H-1D), 4.92 (br s, H-
11: R = All
12: R = H
1C), 4.62 (d, J¼3.0 Hz, H-1A), 4.57 (br s, H-1B) in 1H NMR and at
e
d
102.5 (C-1C), 102.2 (C-1D), 100.7 (C-1E), 100.2 (C-1B), 97.3 (C-1A) in
the 13C NMR spectra] (Scheme 4).
Scheme 2. Reagents: (a) BSP, TTBP, Tf2O, ꢀ60 ꢁC, 1 h, then addition of compound 3,
ꢀ78 ꢁC, 2 h, 55% (and 20%
a-glycoside); (b) 0.1 M CH3ONa, CH3OH, 1 h, 92%; (c) Tf2O,
pyridine, CH2Cl2, ꢀ10 ꢁC, 1 h; (d) NaN3, DMF, TBAB, 70 ꢁC, 5 h, 37% in two steps; (e)
PdCl2, CH3OH, 0 ꢁC, room temperature, 3 h, 76%.
Ph
OBn
O
O
O
In another experiment, iodonium ion mediated stereoselective
C
HO
1,2-cis glycosylation of
D
-galactose thioglycoside 718 with
D-man-
SEt
8
nose acceptor 5 in the presence of a combination of N-iodosucci-
nimide and HClO4eSiO2 in diethyl etheredichloromethane solvent
furnished disaccharide PMP glycoside 13 in 68% yield.33,34 Spectral
Ph
BnO
OBn
O
O
O
D
NH
O
a
O
CCl3
AcO
E
analysis of compound 13 confirmed its formation [signals at
d
5.50
Orthogonal glycosylation
O
(d, J¼3.0 Hz, H-1E), 5.46 (br s, H-1D) in the 1H NMR and at
d
97.7 (C-
14
AcO
OBn
1E), 97.5 (C-1D) in the 13C NMR spectra]. Oxidative removal of PMP
group from compound 13 by the treatment with ammonium cer-
ium(IV) nitrate35 in acetonitrileewater furnished hemiacetal de-
rivative which on treatment with trichloroacetonitrile in the
presence of DBU36 furnished disaccharide trichloroacetimidate
derivative 14 in 58% yield in two steps (Scheme 3).
OBn
O
Ph
O
O
O
D
SEt
BnO
E
O
AcO
AcO
C
O
O
O
O
Ph
15
OBn
OBn
OBn
O
Ph
O
O
N3
Successive Iterative
O
NHCbz
Ph
BnO
OBn
O
B
O
HO
O
O
A
glycosylations in one pot
O
O
D
O
O
b
a
Ph
12
5
7
+
AcO
E
OR
O
OBn
O
Ph
Ph
BnO
OBn
O
O
O
O
AcO
N3
O
NHCbz
OBn
13: R = PMP
O
B
D
O
O
O
A
O
O
O
O
O
AcO
AcO
Ph
b, c
C
E
O
O
OBn
14: R = C(NH)CCl3
O
Ph
OBn
Scheme 3. Reagents: (a) NIS, HClO4eSiO2, MS 4 A, Et2OeCH2Cl2 (3:1), ꢀ10 ꢁC, 1 h, 68%;
(b) Cerium(IV) ammonium nitrate, CH3CNeH2O (4:1), 0 ꢁC, 2 h; (c) CCl3CN, DBU,
CH2Cl2, ꢀ10 ꢁC, 1 h, 58%.
ꢀ
16
c, d, e, f
Having equipped with tailor made disaccharide acceptor 12 and
disaccharide donor 14, attempts were made to develop a one pot
reaction condition for two stereoselective iterative glycosylations
exploiting the orthogonal property of thioglycoside derivative 8.19
In an one pot set up, stereoselective orthogonal glycosylation37 of
1
Scheme 4. Reagents: (a) HClO4eSiO2, CH2Cl2, ꢀ10 ꢁC, 20 min; (b) compound 12, NIS,
ꢀ10 ꢁC, 30 min, 64% in two iterative steps; (c) CH3COSH, pyridine, room temperature,
24 h; (d) 20% Pd(OH)2eC, H2, CH3OH, room temperature, 6 h; (e) (i) (i) NaBr, TBAB,
TEMPO, NaHCO3, NaOCl, CH2Cl2, H2O, 0e5 ꢁC, 3 h; (ii) tert-butanol, 2-methyl-but-2-
ene, NaClO2, NaH2PO4, room temperature, 3 h; (f) H2, 20% Pd(OH)2eC, CH3OH, room
temperature, 24 h over all 50% in four steps.
thioglycoside acceptor 8 with disaccharide trichloroacetimidate
38,39
derivative 14 in the presence of HClO4eSiO2
furnished the

4438
A. Si, A.K. Misra / Tetrahedron 72 (2016) 4435e4441
The synthesized pentasaccharide 1 containing a 2-aminoethyl
68.6 (C-6), 67.2 (OCH2), 67.1 (C-3), 63.6 (C-5), 40.7 (NCH2), 20.9
(COCH3); ESI-MS [MþNa]þ, found 510.1, C25H29NO9 requires 487.18.
linker attached to the reducing end could be conjugated to an ap-
propriate protein using standard reaction conditions for the func-
tionalization of glycan and protein and their conjugation to furnish
glycoprotein conjugate derivative.45
4.1.2. p-Methoxyphenyl
2-O-benzyl-4,6-O-benzylidene-a-D-man-
nopyranoside (5). To a solution of compound 423 (2 g, 5.34 mmol) in
CH2Cl2 (10 mL) were added benzyl bromide (1.2 mL, 10.08 mmol),
TBAB (100 mg) and 10% aq NaOH (10 mL) and the reaction mixture
was allowed to stir briskly at room temperature for 30 h. The re-
action mixture was diluted with CH2Cl2 (100 mL), the organic layer
was washed with H2O (100 mL), dried (Na2SO4) and concentrated.
The crude product was purified by flash chromatography (20%
EtOAc/hexane) to give pure compound 5 (1.7 g, 69%) as a white
solid; [Found C, 69.62; H, 6.22. C27H28O7 requires C, 69.81; H,
6.08%]; Rf (20% EtOAc/Hexane) 0.3; mp 118e120 ꢁC [EtOH];
3. Conclusions
In summary, synthesis of the 2-aminoethyl glycoside of the
pentasaccharide repeating unit of the O-antigen of E. coli O59 strain
has been achieved using orthogonal glycosylation and iterative
glycosylations in one pot in minimum number of steps. The
mannosidic linkage was prepared by the activation of -mannosyl
thioglycoside using a combination of BSP and Tf2O whereas the
glucosaminyl moiety was derived from -mannosyl moiety by
b-D-
D
a-D-
[a
]
25ꢀ6.5 (c 1.0, CHCl3); nmax (KBr): 2800, 3020, 2990, 1371, 1232,
D
a-D
1047 cmꢀ1; 1H NMR (500 MHz, CDCl3):
d
7.47e7.26 (10 H, m, AreH),
the azidolysis with inversion of configuration at C-2 hydroxy group
in moderate yield. A late stage TEMPO mediated selective oxidation
6.80 (2 H, d, J¼9.0 Hz, AreH), 6.91 (2 H, d, J¼9.0 Hz, AreH), 5.55
(1 H, s, PhCH), 5.40 (1 H, br s, H-1), 4.77 (2 H, br s, PhCH2), 4.28e4.23
(1 H, m, H-5), 4.18 (1 H, dd, J¼10.5, 4.5 Hz, H-6a), 4.00e3.98 (1 H, m,
H-2), 3.96 (1 H, t, J¼9.5 Hz, H-4), 3.91 (1 H, dd, J¼9.5, 3.0 Hz, H-3),
3.80e3.77 (1 H, m, H-6b), 3.76 (3 H, s, OCH3); 13C NMR (125 MHz,
of the primary group resulted in the formation of a
acid moiety in the molecule.
D-galacturonic
4. Experimental
CDCl3): d 155.1e114.6 (AreC), 102.1 (PhCH), 97.3 (C-1), 79.3 (C-4),
78.3 (C-2), 73.9 (PhCH2), 68.7 (C-3), 68.6 (C-6), 64.1 (C-5), 55.5
4.1. General methods
(OCH3); ESI-MS [MþNa]þ, found 487.1, C27H28O7 requires 464.18.
All reactions were monitored by thin layer chromatography over
silica gel coated TLC plates. The spots on TLC were visualized by
warming ceric sulfate (2% Ce(SO4)2 in 2N H2SO4) sprayed plates in
hot plate. Silica gel 230e400 mesh was used for column chroma-
tography. NMR spectra were recorded on Bruker Avance 500 MHz
using CDCl3 as solvent and TMS as internal reference unless stated
4.1.3. 2-(N-Benzyloxycarbonyl)aminoethyl O-(3-O-allyl-2-O-benzyl-
4,6-O-benzylidene-b-D-mannopyranosyl)-(1/3)-2-O-acetyl-4,6-O-
benzylidene-a-D-mannopyranoside (9). To a solution of compound
617 (1 g, 2.26 mmol) in dry CH2Cl2 (10 mL) were added BSP (570 mg,
ꢀ
2.72 mmol), TTBP (850 mg, 3.42 mmol) and MS 4 A (2 g) and the
reaction mixture was cooled to ꢀ60 ꢁC under argon. To the cooled
reaction mixture was added triflic anhydride (0.5 mL, 2.97 mmol)
and the reaction mixture was stirred at same temperature for 1 h.
After complete consumption of the starting material the reaction
mixture was further cooled to ꢀ78 ꢁC. To the cold reaction was
added compound 3 (1 g, 2.05 mmol) and the reaction was contin-
ued for another 2 h at same temperature. The reaction mixture was
warmed to the room temperature and diluted with CH2Cl2 (50 mL).
The organic layer was washed with satd. NaHCO3 (50 mL), H2O
(50 mL), dried (Na2SO4) and concentrated. The crude product was
purified by flash chromatography (30% EtOAc/hexane) to give pure
compound 9 (980 mg, 55%) as colorless oil; [Found: C, 66.25; H,
6.34. C48H53NO14 requires C, 66.42; H, 6.15%]; Rf (30% EtOAc/Hex-
otherwise. Chemical shift value is expressed in
d ppm. The com-
plete assignment of proton and carbon spectra was carried out by
using a standard set of NMR experiments, e.g., 1H NMR, 13C NMR,
13C DEPT 135, 2D COSY and 2D HSQC etc. MALDI-MS were recorded
on a Bruker mass spectrometer. Optical rotations were recorded in
a Jasco P-2000 spectrometer. Elemental analyses were carried out
in Euro-3000 elemental analyzer. Commercially available grades of
organic solvents of adequate purity are used in all reactions.
HClO4eSiO2 was prepared following the reported method.34
4.1.1. 2-(N-Benzyloxycarbonyl)aminoethyl 2-O-acetyl-4,6-O-benzyli-
dene-a-D
-mannopyranoside (3). To a solution of compound 220 (2 g,
5.60 mmol) in dry CH3CN (10 mL) were added PhCH(OCH3)2 (1 mL,
6.66 mmol) and iodine (150 mg, 0.59 mmol) and the reaction
mixture was stirred at room temperature for 1.5 h. The solvents
were removed under reduced pressure and co-evaporated with
toluene (3ꢂ50 mL). To a solution of the crude product in anhydrous
DMF (10 mL) were added CH3C(OC2H5)3 (1.5 mL, 8.18 mmol) and
camphorsulfonic acid (300 mg) and the reaction mixture was stir-
red for 1 h. After complete consumption of the starting material and
formation of a faster moving product, H2O (5 mL) was added to the
reaction and allowed to stir for 1 h. The solvents were removed
under reduced pressure and the crude product was purified by flash
chromatography (50% EtOAc/hexane) to give pure compound 3 (2 g,
74%) as a white solid; [Found C, 61.42; H, 6.20. C25H29NO9 requires
C, 61.59; H, 6.00%]; Rf (50% EtOAc/Hexane) 0.3; mp 158e160 ꢁC
ane) 0.2; [
a]
25ꢀ6.5 (c 1.0, CHCl3); nmax (neat): 3436, 2854, 1715,
D
1611, 1515, 1455, 1391, 1222, 912, 723, 697 cmꢀ1; 1H NMR (500 MHz,
CDCl3):
d 7.63e7.26 (20 H, m, Ar-H), 6.02e5.95 (1 H, m, CH]CH2),
5.72 (1 H, s, PhCH), 5.61 (1 H, s, PhCH), 5.45 (1 H, br s, H-2A),
5.41e5.25 (2 H, m, CH]CH2), 5.24e5.21 (1 H, m, NH), 5.20 (2 H, br s,
Cbz), 4.92 (1 H, d, J¼12.0 Hz, PhCH), 4.87 (1 H, d, J¼12.0 Hz, PhCH),
4.86 (1 H, br s, H-1A), 4.74 (1 H, br s, H-1B), 4.41 (1 H, dd, J¼10.0,
3.5 Hz, H-3A), 4.36e4.30 (3 H, m, H-6abB, OCH2eCH]), 4.22e4.16
(2 H, m, H-4B, OCH2eCH]), 4.07 (1 H, t, J¼9.5 Hz, H-4A), 3.97e3.90
(4 H, m, H-5A, H-6abA, OCH), 3.87 (1 H, d, J¼3.0 Hz, H-2B), 3.64e3.63
(1 H, m, OCH), 3.61 (1 H, dd, J¼9.5, 3.0 Hz, H-3B), 3.60e3.43 (2 H, m,
NCH2), 3.42e3.37 (1 H, m, H-5B), 2.15 (3 H, s, COCH3); 13C NMR
(125 MHz, CDCl3):
d 169.7 (COCH3), 156.1e126.1 (AreC, CH]CH2),
116.5 (CH]CH2),101.7 (PhCH),101.3 (PhCH), 99.7 (JC-1/H-1¼155.0 Hz,
C-1B), 98.7 (JC-1/H-1¼170.0 Hz, C-1A), 78.6 (C-4B), 78.0 (C-3B), 77.3 (C-
4A), 77.1 (C-2B), 74.7 (PhCH2), 72.7 (C-3A), 71.5 (OCH2eCH]), 69.0
(C-2A), 68.5 (2 C, C-6A, C-6B), 67.7 (C-5B), 67.1 (Cbz), 66.9 (OCH2),
64.2 (C-5A), 40.6 (NCH2), 20.8 (COCH3); MALDI-MS [MþNa]þ,
found: 890.3, C48H53NO14 requires 867.35.
[EtOH]; [
a]
25ꢀ9 (c 1.0, CHCl3); nmax (KBr): 3031, 2924, 1959, 1715,
D
1611, 1515, 1452, 1390, 1079, 912, 873, 725, 697 cmꢀ1
;
1H NMR
(500 MHz, CDCl3): d 7.47e7.28 (10 H, m, AreH), 5.57 (1 H, s, PhCH),
5.19 (1 H, d, J¼2.5 Hz, H-2), 5.10 (2 H, br s, Cbz), 5.08e5.06 (1 H, m,
NH), 4.77 (1 H, br s, H-1), 4.26e4.24 (1 H, m, H-5), 4.19 (1 H, dd,
J¼9.5 Hz, 3.0 Hz, H-3), 3.89 (1 H, t, J¼9.5 Hz, H-4), 3.82e3.77 (3 H,
m, H-6ab, OCH), 3.55e3.50 (1 H, m, OCH), 3.48e3.36 (2 H, m, NCH2),
4.1.4. 2-(N-Benzyloxycarbonyl)aminoethyl O-(3-O-allyl-2-O-benzyl-
2.16 (3 H, s, COCH3); 13C NMR (125 MHz, CDCl3):
d
170.2 (COCH3),
4,6-O-benzylidene-
b-D-mannopyranosyl)-(1/3)-4,6-O-benzylidene-
137.0e126.2 (AreC), 102.2 (PhCH), 98.6 (C-1), 78.9 (C-4), 71.9 (C-2),
a-D-mannopyranoside (10). A solution of compound 9 (900 mg,

A. Si, A.K. Misra / Tetrahedron 72 (2016) 4435e4441
4439
1.04 mmol) in 0.1 M CH3ONa in CH3OH (15 mL) was stirred at room
temperature for 1 h, neutralized with Dowex 50W X8 (Hþ) resin,
filtered and concentrated. The crude product was purified by flash
chromatography (40% EtOAc/hexane) to give pure compound 10
(790 mg, 92%) as colorless oil; [Found: C, 66.75; H, 6.40. C46H51NO13
0
ꢁC for 3 h. The solvents were removed under reduced pressure
and the crude product was purified by flash chromatography (25%
EtOAc/hexane) to give pure compound 12 (400 mg, 76%) as white
solid; [Found: C, 63.52; H, 5.90. C43H46N4O12 requires C, 63.69; H,
5.72%]; Rf (25% EtOAc/Hexane) 0.3; mp 78e80 ꢁC [EtOH]; [
a
]
25ꢀ10
D
requires C, 66.90; H, 6.22%]; Rf (40% EtOAc/Hexane) 0.25; [
a
]
25ꢀ6.5
(c 1.0, CHCl3); nmax (KBr): 3432, 2932, 2111, 1777, 1455, 1390, 1237,
D
(c 1.0, CHCl3); nmax (neat): 3032, 1712, 1529, 1252, 1073, 874, 751,
1097, 1044, 994, 916 cmꢀ1; 1H NMR (500 MHz, CDCl3):
d 7.50e7.15
722, 697 cmꢀ1
;
1H NMR (500 MHz, CDCl3):
d
7.60e7.37 (20 H, m,
(20 H, m, AreH), 5.50 (1 H, s, PhCH), 5.35 (1 H, s, PhCH), 5.23e5.15
(1 H, m, NH), 5.08 (2 H, br s, Cbz), 4.98 (1 H, d, J¼12.0 Hz, PhCH),
4.84 (1 H, d, J¼3.5 Hz, H-1A), 4.78 (1 H, br s, H-1B), 4.65 (1 H, d,
J¼11.5 Hz, PhCH), 4.27e4.19 (2 H, m, H-3A, H-6aB), 4.15e4.08 (1 H,
m, H-6bB), 3.92 (1 H, d, J¼3.5 Hz, H-2B), 3.87e3.73 (2 H, m, H-4A, H-
5A), 3.72e3.62 (4 H, m, H-3B, H-6abA, OCH), 3.60e3.52 (2 H, m, H-
4B, OCH), 3.51e3.35 (2 H, m, NCH2), 3.25 (1 H, dd, J¼9.5 Hz, 3.5 Hz,
H-2A), 3.24e3.18 (1 H, m, H-5B); 13C NMR (125 MHz, CDCl3):
AreH), 6.02e5.97 (1 H, m, CH]CH2), 5.68 (1 H, s, PhCH), 5.61 (1 H,
s, PhCH), 5.42e5.27 (2 H, m, CH]CH2), 5.22 (2 H, br s, Cbz),
5.20e5.17 (1 H, m, NH), 5.05 (1 H, d, J¼11.5 Hz, PhCH), 4.95 (1 H, br s,
H-1A), 4.93 (1 H, d, J¼11.5 Hz, PhCH), 4.87 (1 H, br s, H-1B),
4.40e4.35 (2 H, m, H-3A, OCH2eCH]), 4.30e4.19 (5 H, m, H-4B, H-
6
abB, H-6aA, OCH2eCH]), 4.03 (1 H, d, J¼3.0 Hz, H-2B), 4.00 (1 H, br
s, H-2A), 3.98e3.88 (4 H, m, H-4A, H-5A, H-6bA, OCH), 3.67 (1 H, dd,
J¼10.0 Hz, 3.0 Hz, H-3B), 3.65e3.63 (1 H, m, OCH), 3.60e3.45 (2 H,
m, NCH2), 3.40e3.33 (1 H, m, H-5B); 13C NMR (125 MHz, CDCl3):
d
156.1e126.2 (AreC), 101.9 (C-1B), 101.7, 101.5 (2 C, 2 PhCH), 98.9
(C-1A), 80.2 (C-4B), 79.3 (C-4A), 78.6 (C-2B), 75.7 (C-3A), 75.4
(PhCH2), 70.6 (C-3B), 68.6 (C-6A), 68.5 (C-6B), 67.9 (OCH2), 67.1 (C-
5B), 66.8 (Cbz), 63.3 (C-5A), 63.2 (C-2A), 40.7 (NCH2); MALDI-MS
[MþNa]þ, found 833.3, C43H46N4O12 requires 810.31.
d
156.1e126.1 (AreC, CH]CH2), 116.5 (CH]CH2), 101.8, 101.4 (2
PhCH), 100.2 (C-1A), 99.8 (C-1B), 78.8 (C-4B), 78.1 (C-3B), 77.3 (C-4A),
76.3 (C-2B), 75.8 (C-3A), 74.6 (PhCH2), 71.9 (OCH2eCH]CH2), 69.9
(C-2A), 68.7 (C-6A), 68.6 (C-6B), 67.5 (C-5B), 66.9 (2 C, OCH2, Cbz),
63.8 (C-5A), 40.7 (NCH2); MALDI-MS [MþNa]þ, found: 848.3,
4.1.7. p-Methoxyphenyl O-(3,4-di-O-acetyl-2,6-di-O-benzyl- -gal-
a-D
C
46H51NO13 requires 825.34.
actopyranosyl)-(1/3)-2-O-benzyl-4,6-O-benzylidene-a-D-man-
nopyranoside (13). To a solution of compound 5 (1 g, 2.15 mmol)
and compound 718 (1.1 g, 2.25 mmol) in Et2OeCH2Cl2 (15 mL; 3:1)
were added MS 4 A (2 g) and the reaction mixture was cooled to
4.1.5. 2-(N-Benzyloxycarbonyl)aminoethyl O-(3-O-allyl-2-O-benzyl-
4,6-O-benzylidene- -mannopyranosyl)-(1/3)-2-azido-4,6-O-ben-
zylidene-2-deoxy- -glucopyranoside (11). To a solution of com-
ꢀ
b
-
D
a-D
ꢀ10 ꢁC under argon. To the cooled reaction mixture were added NIS
(510 mg, 2.26 mmol) and HClO4eSiO2 (20 mg) and it was stirred at
same temperature for 1 h. The reaction mixture was diluted with
CH2Cl2 (50 mL) and filtered. The filtrate was successively washed
with 5% Na2S2O3 (50 mL), satd. NaHCO3 (50 mL) and H2O (50 mL),
dried (Na2SO4) and concentrated. The crude product was purified
by flash chromatography (20% EtOAc/hexane) to give pure com-
pound 13 (1.3 g, 68%) as white solid; [Found: C, 68.58; H, 6.30.
pound 10 (700 mg, 0.85 mmol) in dry CH2Cl2 (5 mL), was added
pyridine (0.5 mL) and triflic anhydride (0.3 mL, 1.78 mmol) and the
reaction mixture was allowed to stir at ꢀ10 ꢁC for 1 h. The reaction
mixture was diluted with CH2Cl2 (25 mL) and the organic layer was
washed with satd. NaHCO4 (25 mL), H2O (25 mL), dried (Na2SO4)
and concentrated. To a solution of the crude triflate derivative in
anhydrous DMF (5 mL) were added NaN3 (300 mg, 4.61 mmol) and
TBAB (50 mg) and the reaction mixture was stirred at 70 ꢁC for 5 h.
The reaction mixture was poured into water and extracted with
CH2Cl2 (25 mL). The organic layer was washed with H2O (25 mL),
dried (Na2SO4) and concentrated. The crude product was purified
by flash chromatography (30% EtOAc/hexane) to give pure com-
pound 11 (270 mg, 37%) as white solid; [Found: C, 64.76; H, 6.10.
C
51H54O14 requires C, 68.75; H, 6.11%]; Rf (20% EtOAc/Hexane) 0.25;
mp 128e130 ꢁC [EtOH]; [
a
]
25þ2 (c 1.0, CHCl3); nmax (KBr): 3060,
D
2933, 2876, 1746, 1613, 1587, 1369, 1217, 1094, 985, 912, 797 cmꢀ1
;
1H NMR (500 MHz, CDCl3):
d 7.50e7.10 (18 H, m, AreH), 6.99e6.90
(2 H, m, AreH), 6.88 (2 H, d, J¼9.0 Hz, AreH), 6.75 (2 H, d, J¼9.0 Hz,
AreH), 5.50 (1 H, d, J¼3.0 Hz, H-1E), 5.46 (1 H, br s, H-1D), 5.45 (1 H,
s, PhCH), 5.35 (1 H, dd, J¼10.5, 3.0 Hz, H-3E), 5.30 (1 H, d, J¼3.5 Hz,
H-4E), 4.83 (1 H, d, J¼12.0 Hz, PhCH), 4.77 (1 H, d, J¼12.0 Hz, PhCH),
4.52e4.43 (3 H, m, 2 PhCH, H-3D), 4.38e4.30 (2 H, m, PhCH, H-4D),
4.25 (1 H, d, J¼12.5 Hz, PhCH), 4.16e4.14 (1 H, m, H-6aD), 4.05 (1 H,
br s, H-2D), 4.01e3.95 (1 H, m, H-5D), 3.88e3.80 (2 H, m, H-5E, H-
6D), 3.79 (3 H, s, OCH3), 3.77e3.71 (1 H, m, H-2E), 3.43e3.40 (1 H, m,
C
46H50N4O12 requires C, 64.93; H, 5.92%]; Rf (40% EtOAc/Hexane)
0.35; mp 60e62 ꢁC [EtOH]; [
a]
25ꢀ10 (c 1.0, CHCl3); nmax (KBr): 3421,
D
2933, 2109, 1777, 1497, 1455, 1256, 1043, 993, 698 cmꢀ1
;
1H NMR
(500 MHz, CDCl3):
d 7.65e7.25 (19 H, m, Ar-H), 6.00e5.91 (1 H, m,
CH]CH2), 5.64 (1 H, s, PhCH), 5.61 (1 H, s, PhCH), 5.40e5.35 (1 H, m,
CH]CH2), 5.32e5.27 (1 H, m, NH), 5.26e5.23 (1 H, m, CH]CH2),
5.22 (2 H, br s, Cbz), 5.02 (1 H, d, J¼3.5 Hz, H-1A), 4.97 (2 H, dd,
J¼11.5, 3.5 Hz, PhCH2), 4.85 (1 H, br s, H-1B), 4.39e4.28 (3 H, m, H-
H-6aE), 3.32e3.29 (1 H, m, H-6bE), 2.00, 1.99 (6 H, 2 s, 2 COCH3); 13
NMR (125 MHz, CDCl3): 169.9, 169.7 (2 COCH3), 155.1e114.6
C
d
3A, H-6aA
,
OCH2eCH]), 4.28e4.13 (3 H, m, H-6aB
,
H-4A,
(AreC), 102.4 (PhCH), 97.7 (C-1E), 97.5 (C-1D), 79.1 (C-4D), 76.9 (C-
2D), 73.4 (PhCH2), 73.3 (C-3D), 73.1 (PhCH2), 72.4 (C-2E), 70.5
(PhCH2), 69.1 (C-3E), 69.0 (C-4E), 68.8 (2 C, C-6D, C-6E), 67.9 (C-5E),
64.7 (C-5D), 55.6 (OCH3), 20.8, 20.6 (2 COCH3); MALDI-MS
[MþNa]þ, found 913.3, C51H54O14 requires 890.35.
OCH2eCH]), 4.09 (1 H, d, J¼3.0 Hz, H-2B), 3.99e3.86 (3 H, m, H-5A,
H-6aB, OCH), 3.85e3.78 (1 H, m, H-6bB), 3.74 (1 H, t, J¼9.0 Hz, H-4B),
3.70e3.65 (1 H, m, OCH), 3.62 (1 H, dd, J¼9.5, 3.5 Hz, H-3B),
3.61e3.45 (2 H, m, NCH2), 3.38 (1 H, dd, J¼9.5, 3.5 Hz, H-2A),
3.35e3.29 (1 H, m, H-5B); 13C NMR (125 MHz, CDCl3):
d 156.1e126.1
(AreC, CH]CH2), 116.5 (CH]CH2), 102.2 (C-1B), 101.4 (2 C, 2 PhCH),
98.8 (C-1A), 80.3 (C-4B), 78.6 (C-4A), 78.1 (C-3B), 77.3 (C-2B), 76.3 (C-
3A), 75.1 (PhCH2), 71.6 (OCH2eCH]CH2), 68.6 (C-6A), 68.5 (C-6B),
67.8 (OCH2), 67.6 (C-5B), 66.8 (Cbz), 63.2 (C-5A), 63.1 (C-2A), 40.7
(NCH2); MALDI-MS [MþNa]þ, found 873.3, C46H50N4O12 requires
850.34.
4.1.8. (3,4-Di-O-acetyl-2,6-di-O-benzyl-
a
-
D
-galactopyranosyl)-
tri-
(1/3)-2-O-benzyl-4,6-O-benzylidene- -mannopyranosyl
a-D
chloroacetimidate (14). A solution of compound 13 (1 g,
1.12 mmol) in CH3CNeH2O (15 mL; 4:1) was cooled to 0 ꢁC. To the
cooled reaction mixture was added CAN (1.2 g, 2.19 mmol) and it
was stirred at same temperature for 2 h. The reaction mixture was
diluted with H2O (50 mL) and extracted with CH2Cl2 (50 mL). The
organic layer was successively washed with satd. NaHCO3 (50 mL)
and H2O (50 mL), dried (Na2SO4) and concentrated. The crude
product was passed through a short pad of SiO2 using hex-
aneeEtOAc (2:1) as eluent to give disaccharide hemiacetal de-
rivative. To a solution of the hemiacetal derivative (545 mg) in
4.1.6. 2-(N-Benzyloxycarbonyl)aminoethyl
benzylidene- -mannopyranosyl)-(1/3)-2-azido-4,6-O-benzyli-
dene-2-deoxy- -glucopyranoside (12). To a solution of com-
O-(2-O-benzyl-4,6-O-
b-D
a-D
pound 11 (550 mg, 0.65 mmol) in dry CH3OH (8 mL) was added
PdCl2 (50 mg, 0.28 mmol) and the reaction mixture was stirred at

4440
A. Si, A.K. Misra / Tetrahedron 72 (2016) 4435e4441
anhydrous CH2Cl2 (5 mL) was added CCl3CN (0.5 mL, 4.98 mmol)
was added 20% Pd(OH)2eC (100 mg) and the reaction mixture was
stirred at room temperature under a positive pressure of hydrogen
for 6 h. The reaction mixture was filtered through a Celite bed,
washed with CH3OH (20 mL) and concentrated. To the solution of
the partially hydrogenated product in CH2Cl2 (10 mL) and H2O
(1.5 mL) were added 1M aq NaBr (2 mL),1M aq TBAB (2 mL), TEMPO
(50 mg, 0.32 mmol), satd aq NaHCO3 (10 mL) and 4% aq NaOCl
(15 mL) successively and the reaction mixture was stirred at 0e5 ꢁC
for 3 h. The reaction mixture was neutralized with the addition of
1 N aq HCl solution. To the reaction mixture were added tert-bu-
tanol (10 mL), 2-methyl-but-2-ene (10 mL; 2 M solution in THF),
20% aq NaClO2 (10 mL) and 20% aq NaH2PO4 (5 mL) and the reaction
mixture was stirred at room temperature for 3 h. The reaction
mixture was concentrated and the crude mass was extracted with
CHCl3 (50 mL). The organic layer was washed with water (20 mL),
dried (Na2SO4) and concentrated to dryness. The crude product was
passed through a short pad of SiO2 using CH2Cl2eCH3OH (2:1) as
eluent. To a solution of the oxidized product in CH3OH (5 mL) was
added 20% Pd(OH)2eC (100 mg) and the reaction mixture was
stirred at room temperature under a positive pressure of hydrogen
for 24 h. The reaction mixture was filtered through a Celite bed,
washed with CH3OHeH2O (10 mL; 2:1) and concentrated. The size
exclusion chromatography of the crude product through a Sepha-
dex LH-20 column (25% CH3OH/H2O) gave pure compound 1
(130 mg, 50%) as white powder; [Found: C, 44.95; H, 6.40.
and it was cooled to ꢀ10 ꢁC. DBU (20
mL) was added to the cooled
reaction mixture and it was allowed to stir at same temperature for
1 h. The solvents were removed under reduced pressure and the
crude product was purified over SiO2 using hexaneeEtOAc (8:1) to
give pure compound 14 (600 mg, 58%) which was used in next step
immediately without further analytical characterization.
4.1.9. 2-(N-Benzyloxycarbonyl)aminoethyl O-(3,4-di-O-acetyl-2,6-
di-O-benzyl-a-D-galactopyranosyl)-(1/3)-O-(2-O-benzyl-4,6-O-
benzylidene-
benzylidene-
a
-
-
D
-mannopyranosyl)-(1/3)-O-(2-O-benzyl-4,6-O-
-mannopyranosyl)-(1/3)-O-(2-O-benzyl-4,6-O-
a
D
benzylidene-
b
-
D
-mannopyranosyl)-(1/3)-2-azido-4,6-O-benzyli-
dene-2-deoxy-
a-D-glucopyranoside (16). To a solution of compound
819 (200 mg, 0.49 mmol) in anhydrous CH2Cl2 (5 mL) was added
trichloroacetimidate derivative (14; 500 mg, 0.54 mmol) in CH2Cl2
(5 mL) and the reaction mixture was cooled to ꢀ10 ꢁC under argon.
To the cooled reaction was added HClO4eSiO2 (30 mg) and it was
stirred at same temperature for 20 min. After consumption of
compound 14 and formation of a new compound (checked by TLC),
compound 12 (350 mg, 0.43 mmol) was added to the reaction to-
gether with NIS (100 mg, 0.44 mmol) and stirring was continued for
another 30 min at same temperature. The reaction mixture was
diluted with CH2Cl2 (50 mL) and the organic layer was successively
washed with 5% Na2S2O3 (50 mL), satd. NaHCO3 (50 mL) and H2O
(50 mL), dried (Na2SO4) and concentrated. The crude product was
purified by flash chromatography (25% EtOAc/hexane) to give pure
compound 16 (600 mg, 64%) as colorless oil; [Found: C, 66.84; H,
6.10. C107H112N4O29 requires C, 67.00; H, 5.89%]; Rf (25% EtOAc/
C
38H62N2O29 requires C, 45.15; H, 6.18%]; Rf (CHCl3eCH3OHeH2O;
10:5:1.5) 0.3; [
a
]
25ꢀ19 (c 0.5, H2O); nmax (KBr): 2920, 1746, 1377,
D
1221, 1066, 766 cmꢀ1
;
1H NMR (500 MHz, D2O):
d
5.13 (1 H, d,
J¼4.0 Hz, H-1E), 4.97 (1 H, br s, H-1D), 4.92 (1 H, br s, H-1C), 4.62
(1 H, d, J¼3.0 Hz, H-1A), 4.57 (1 H, br s, H-1B), 4.13 (1 H, br s, H-2D),
4.07 (1 H, br s, H-2C), 3.97e3.92 (2 H, m, H-2A, H-5E), 3.88e3.83
(4 H, m, H-2B, H-3C, H-3D, H-4E), 3.80e3.73 (4 H, m, H-3A, H-4D, H-
Hexane) 0.3; [
a]
25ꢀ9 (c 1.0, CHCl3); nmax (neat): 3444, 3019, 2920,
D
2855, 2116, 1742, 1670, 1455, 1386, 1216, 978, 927, 699 cmꢀ1
;
1H
NMR (500 MHz, CDCl3):
d 7.50e6.92 (50 H, m, Ar-H), 5.57 (1 H, s,
PhCH), 5.51 (1 H, s, PhCH), 5.45 (1 H, d, J¼3.5 Hz, H-1E), 5.37 (1 H, br
s, H-1D), 5.36 (1 H, s, PhCH), 5.28 (2 H, br s, PhCH, H-1C), 5.25 (1 H,
dd, J¼9.5, 3.0 Hz, H-3E), 5.20 (2 H, m, H-4E, NH), 5.08 (2 H, br s, Cbz),
4.93 (1 H, d, J¼12.0 Hz, PhCH), 4.90 (1 H, d, J¼3.0 Hz, H-1A), 4.55
(1 H, d, J¼12.0 Hz, PhCH), 4.85 (1 H, br s, H-1B), 4.81 (1 H, d,
J¼12.0 Hz, PhCH), 4.48 (2 H, t, J¼10.5 Hz, PhCH2), 4.35 (1 H, d,
J¼12.0 Hz, PhCH), 4.19e4.08 (12 H, m, H-3A, H-3C, H-3D, H-4C, H-4D,
H-6abB, H-6aC, 4 PhCH2), 4.07e4.39 (2 H, m, H-4A, H-6bC), 3.98 (1 H,
d, J¼3.0 Hz, H-2B), 3.92e3.78 (6 H, m, H-2C, H-2D, H-3B, H-5A, H-5E,
6abC), 3.72e3.63 (8 H, m, H-2E, H-3E, H-4C, H-5A, H-5D, H-6aB, H-6aD,
OCH), 3.62e3.55 (6 H, m, H-3B, H-5C, H-6abA, H-6bB, H-6bD),
3.54e3.50 (1 H, m, H-4B), 3.48e3.43 (1 H, m, H-4A), 3.42e3.37 (1 H,
m, OCH), 3.29e3.25 (1 H, m, H-5B), 2.88e2.80 (2 H, m, NCH2), 1.90
(s, 3 H, COCH3), 1.75 (6 H, 2 s, 2 COCH3); 13C NMR (125 MHz, D2O):
d
175.1 (COOH), 174.2 (3 COCH3), 102.5 (C-1C), 102.2 (C-1D), 100.7 (C-
1E), 100.2 (C-1B), 97.3 (C-1A), 80.7 (2 C, C-3A, C-3B), 78.4 (C-3C), 78.2
(C-3D), 76.1 (C-5B), 73.5 (C-5A), 73.4 (C-5D), 71.7 (C-5C), 71.5 (2 C, C-
3E, C-5E), 70.6 (C-4E), 69.8 (C-2D), 69.7 (C-2C), 69.3 (2 C, C-2B, C-4A),
68.7 (C-4C), 68.4 (2 C, C-2E, C-4D), 66.1 (OCH2), 65.8 (2 C, C-4B, C-6D),
60.9 (C-6B), 60.6 (C-6C), 60.5 (C-6A), 52.5 (C-2A), 39.7 (NCH2), 23.3,
21.9, 20.0 (3 COCH3); MALDI-MS [MþNa]þ, found 1033.3,
C38H62N2O29 requires 1010.34.
H-6aA), 3.75e3.62 (8 H, m, H-2E, H-4B, H-5C, H-5D, H-6bA, H-6abD
,
OCH), 3.60e3.55 (1 H, m, OCH), 3.54e3.35 (2 H, m, NCH2), 3.28 (1 H,
dd, J¼9.5, 3.5 Hz, H-2A), 3.26e3.21 (2 H, m, H-5B, H-6aE), 3.18 (1 H,
dd, J¼10.0, 3.5 Hz, H-6bE), 1.97, 1.91 (6 H, 2 s, 2 COCH3); 13C NMR
(125 MHz, CDCl3):
d 169.5, 169.3 (2 COCH3), 156.1e126.1 (AreC),
102.1 (C-1B),102.0,101.9,101.8,101.4 (4 PhCH), 99.3 (2 C, C-1D, C-1C),
98.8 (C-1A), 97.4 (C-1E), 80.3 (C-4B), 79.3 (C-4A), 79.0 (C-4C), 78.6 (C-
4D), 78.0 (C-2B), 77.2 (C-2D), 76.6 (C-2C), 76.0 (C-3A), 75.6 (C-3D),
75.1 (PhCH2), 73.0 (PhCH2), 72,6 (2 C, C-3B, C-3C), 72.5 (C-2E), 72.2 (2
PhCH2), 70.3 (PhCH2), 69.0 (C-4E), 68.9 (C-3E), 68.8 (C-6A), 68.6 (C-
6B), 68.5 (C-6C), 68.4 (C-6D), 67.9 (C-6E), 67.5 (OCH2), 67.4 (2 C, C-5B,
C-5E), 66.8 (Cbz), 64.6 (C-5D), 64.5 (C-5C), 63.1 (C-5A), 63.0 (C-2A),
40.8 (NCH2), 20.7, 20.5 (2 COCH3); MALDI-MS [MþNa]þ, found
1939.7, C107H112N4O29 requires 1916.74.
Acknowledgements
A.S. thanks CSIR, New Delhi for providing Senior Research Fel-
lowship. The work is supported by SERB, New Delhi (Project No.
EMR/2015/000282 dated 17.09.2015) and Bose Institute. The au-
thors sincerely thank reviewers for their valuable comments to
improve the quality of the manuscript.
Supplementary data
4.1.10. 2-Aminoethyl O-(3,4-di-O-acetyl-
acid)-(1/3)-O-( -mannopyranosyl)-(1/3)-O-(
anosyl)-(1/3)-O-( -mannopyranosyl)-(1/3)-2-acetamido-2-
deoxy- -glucopyranoside (1). To a solution of compound 16
(500 mg, 0.26 mmol) in pyridine (1 mL) was added CH3COSH
(0.2 mL, 2.80 mmol) and the reaction mixture was stirred at room
temperature for 24 h. The solvents were evaporated and co-
evaporated with toluene (3ꢂ20 mL) under reduced pressure. The
crude product was passed through SiO2 using hexaneeEtOAc (1:1)
as eluent. To solution of the acetamido product in CH3OH (5 mL)
a
-
D
-galactopyranosyluronic
Supplementary data associated with this article can be found in
the online version, at http://dx.doi.org/10.1016/j.tet.2016.06.025.
a
-
D
a-D
-mannopyr-
b-D
a-D
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