Synthesis of a trisaccharide and a tetrasaccharide from the cell-wall lipopolysaccharides of Azospirillum brasilense S17

Article abstract of DOI:10.1055/s-0029-1217401

A trisaccharide containing a D-mannosamine moiety and a tetrasaccharide containing an L-rhamnan chain that are found in the cell walls of Azospirillum brasilense S17 were synthesized concisely and in excellent yields. The key features of the synthetic str

Full text of DOI:10.1055/s-0029-1217401

2584
PAPER
Synthesis of a Trisaccharide and a Tetrasaccharide from the Cell-Wall
Lipopolysaccharides of Azospirillum brasilense S17
O
ligosaccharid
h
e
s
of Azospi
a
rillum bras
s
ilense hi Pandey,a Samir Ghosh,b Anup Kumar Misra*b
a
Division of Medicinal and Process Chemistry, Central Drug Research Institute, Lucknow 226001, India
b
Division of Molecular Medicine, Bose Institute, A.J.C. Bose Centenary Campus, P-1/12, C.I.T. Scheme VII-M, Kolkata 700054, India
Fax +91(33)23553886; E-mail: akmisra69@rediffmail.com
Received 11 March 2009
Abstract: A trisaccharide containing a D-mannosamine moiety and
a tetrasaccharide containing an L-rhamnan chain that are found in
the cell walls of Azospirillum brasilense S17 were synthesized con-
cisely and in excellent yields. The key features of the synthetic strat-
egy were stereoselective glycosylations and a minimum number of
protecting-group manipulations.
OH
AcHN
A
HO
O
O
O
HO
HO
B
NHAc
O
OMe
O
C
HO
1
HO
OMe
Key words: oligosaccharides, glycosylations, D-mannosamine,
Azospirillum brasilense, rhizobacteria
SEt
N3
BzO
HO
O
O
BnO
Azospirilla are Gram-negative rhizobacteria that promote
plant growth through their association with various plants
in the rhizosphere.1 Although azospirilla do not associate
with particular plant species, they are widely distributed
in soils and establish close relationships with the roots of
grasses, cereals, and other nonleguminous plants.2 They
fix atmospheric nitrogen and have a positive effect on
plant growth through the action of pectolytic enzymes and
by excreting phytohormones, vitamins, and other biologi-
cally active substances into the rhizosphere.2,3 Macromol-
BnO
SEt
BnO
OMe
OMe
7
10
OAc
O
AcO
AcO
NPhth
11
Figure 1 Structures of trisaccharide methyl glycoside 1 and its syn-
thetic precursors
ecules
such
as
exopolysaccharides,
capsular
OMe
polysaccharides, and lipopolysaccharides that are present
in the bacterial cell surface are actively involved in the in-
teractions of azospirilla with plants.4 It is therefore essen-
tial to understand the roles of the cell-surface
lipopolysaccharides of various strains of azospirilla in the
interactions of the bacteria with plant roots.
O
A
HO
O
OH
O
B
HO
O
HO
OH
OH
O
D
O
O
C
HO
HO
HO
OH
2
For a detailed biological study on the role of cell-surface
oligosaccharides of such bacteria with plants, it will be
necessary to obtain greater quantities of the relevant oli-
gosaccharides than can be obtained by isolation from the
natural source. The development of concise strategies for
the chemical syntheses of these oligosaccharides in large
quantities for use in studies on plant–bacterial interactions
is therefore a highly desirable goal. A trisaccharide con-
taining a D-mannosamine moiety and tetrasaccharide con-
taining an L-rhamnan chain from Azospirillum brasilense
S17, which were recently isolated and characterized by
Zdorovenko and co-workers,5 were synthesized as their
methyl glycosides (1 and 2, respectively; Figure 1 and
Figure 2) in high overall yields by a sequence of reactions
SEt
OMe
O
O
BnO
BnO
AcO
HO
O
OAc
OBn
SEt
OAc
O
AcO
AcO
SEt
AcO
AcO
OAc
OAc
20
Figure 2 Structure of tetrasaccharide methyl glycoside 2 and its
synthetic precursors
starting from monosaccharide intermediates prepared
from commercially available reducing sugars.
Methyl 4,6-O-benzylidene-a-D-glucopyranoside (3)6 was
selectively benzylated to give the 3-O-benzyl derivative
47 through formation of the stannylidene acetal. Glycoside
SYNTHESIS 2009, No. 15, pp 2584–2590
Advanced online publication: 22.06.2009
DOI: 10.1055/s-0029-1217401; Art ID: P04109SS
© Georg Thieme Verlag Stuttgart · New York
x
x.
x
x
.
2
0
0
9

PAPER
Oligosaccharides of Azospirillum brasilense
2585
4 was treated with triflic anhydride in the presence of py- imide and trimethylsilyl trifluoromethanesulfonate gave
ridine, and the resulting triflate derivative was treated the trisaccharide derivative 16 in 89% yield. The stereose-
with sodium azide to give methyl 2-azido-3-O-benzyl-2- lective formation of compound 16 was confirmed by its
deoxy-4,6-O-benzylidene-a-D-mannopyranoside (5) in one- and two-dimensional NMR spectra [d = 5.10 (br s, H-
82% yield. Acidic hydrolysis of the benzylidene acetal in 1C), 4.81 (d, J = 8.3 Hz, H-1B), 4.55 (br s, H-1A) in the 1H
compound 5 with 80% aqueous acetic acid, followed by NMR spectrum, and d = 100.2 (2 C, C-1B, C-1C), 99.5 (C-
selective benzoylation of the primary hydroxy group of 1A) in the 13C NMR spectra] and by mass spectral analy-
the ring-opened product 6 with benzoyl cyanide gave sis. Hydrogenolysis over Pearlmann’s catalyst17 followed
methyl 2-azido-6-O-benzoyl-3-O-benzyl-2-deoxy-a-D- by deacetylation gave the required trisaccharide 1 as its
mannopyranoside (7) in 90% yield. In another experi- methyl glycoside in 81% yield; this was purified over LH-
ment, ethyl 4-O-benzyl-1-thio-a-L-rhamnopyranoside 20 Sephadex by using MeOH–H2O (4:1) as the eluant
(8),8 derived from L-rhamnose, was selectively benzylated (Scheme 2). The presence of signals at d = 4.94 (br s, 1 H,
via its stanylidene acetal to give the monodeprotected H-1C), 4.63 (br s, 1 H, H-1A), and 4.56 (d, J = 7.5 Hz, 1 H,
compound 9,9 which was methylated under alkaline con- H-1B) in 1H NMR spectrum and at d = 102.4 (C-1B), 101.4
dition to give ethyl 3,4-O-benzyl-2-O-methyl-1-thio-a-L- (C-1A), and 94.5 (C-1C) in the 13C NMR spectrum con-
rhamnopyranoside (10) in 90% yield (Scheme 1). Com- firmed the formation of compound 1.
pounds 11,10 17,11 18,12 19,13 and 2014 were prepared in
excellent yields by following the reaction conditions re-
7
11
+
ported in the relevant literature.
a
OAc
R
BzO
O
N3
O
Ph
O
O
R1O
AcO
AcO
Ph
O
O
BnO
O
BnO
NPhth
O
O
b
OMe
R2O
OMe
12: R = N3
b
13: R = NHAc
OMe
3: R1 = R2 = H
4: R1 = Bn, R2 = H
5
a
c
N3
R1O
R2O
OAc
AcHN
O
AcO
O
RO
c
O
O
AcO
AcO
BnO
NHAc
OMe
OMe
6: R1 = R2 = H
7: R1 = Bz, R2 = H
14: R = Bn
d
d
15: R = H
10
OAc
SEt
e
SEt
e
O
O
AcHN
BnO
BnO
AcO
O
HO
BnO
O
OH
OR
AcO
AcO
O
8
9: R = H
10: R = Me
O
NHAc
f
OMe
O
BnO
Scheme 1 Reagents and conditions: (a) (i) see ref 7; (b) triflic an-
hydride, py, CH2Cl2, –10 °C, 1 h then NaN3, HMPT–DMF (1:1),
70 °C, 24 h, 82%; (c) 80% aq AcOH, 80 °C, 2 h, quant.; (d) BzCN,
py, CH2Cl2, 2 h, 90%; (e) see ref. 9; (f) MeI, NaH, DMF, 90%.
BnO
OMe
16
f
1
Iodonium ion-promoted stereoselective glycosylation of
compound 7 with thioglycoside donor 11 in the presence
of N-iodosuccinimide and trimethylsilyl trifluoro-
methanesulfonate15 gave the disaccharide derivative 12,
which on treatment with triethylsilane16 and 20% palladi-
um(II) hydroxide/carbon followed by conventional acety-
lation gave the disaccharide derivative 13 in 91% yield.
Scheme 2 Reagents and conditions: (a) NIS, TMSOTf, CH2Cl2,
–20 °C, 1.5 h, 90%; (b) Et3SiH, 20% Pd(OH)2/C, MeOH–MeOH
(5:1), r.t., 4 h, 91%; (c) (i) H2N(CH2)2NH2, BuOH, 110 °C, 10 h; (ii)
Ac2O, py, 2 h, r.t., 85%; (d) H2, 20% Pd(OH)2/C, r.t., 8 h, 92%; (e)
NIS, TMSOTf, CH2Cl2, –10 °C, 2 h, 89%; (f) H2, 20% Pd(OH)2/C,
r.t., 8 h then MeONa, MeOH, r.t., 12 h, 81%.
1
Stereoselective glycosylation of monodeprotected com-
pound 17 with the thioglycoside donor 18 in the presence
of N-iodosuccinimide and trimethylsilyl trifluoro-
methanesulfonate gave the disaccharide derivative 21 in
90% yield; this was then deacetylated by sodium methox-
ide to give disaccharide diol derivative 22 quantitatively.
The formation of compound 21 was confirmed by the
presence of signals at d = 98.9 (C-1A) and 98.4 (C-1B) in
the 13C NMR spectrum.
The presence of signals in the H NMR spectrum at d =
5.72 (d, J = 8.5 Hz, H-1B) and 4.58 (d, J = 1.6 Hz, H-1A)]
and in the 13C NMR spectrum at d = 98.9 (C-1A) and 98.5
(C-1B) confirmed the formation of compound 12. Treat-
ment of disaccharide 13 with ethane-1,2-diamine in bu-
tan-1-ol followed by conventional acetylation gave di-
saccharide derivative 14 in 85% yield. Disaccharide 14
was hydrogenolyzed over 20% palladium(II) hydroxide/
carbon17 to give disaccharide acceptor 15 in 92% yield.
Stereoselective glycosylation of compound 15 with
thioglycoside donor 10 in the presence of N-iodosuccin-
Selective allylation of compound 22 under phase-transfer
reaction conditions gave the disaccharide acceptor 23 in
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York

2586
S. Pandey et al.
PAPER
84% yield. Iodonium ion-promoted coupling of disaccha- All the reactions were monitored by TLC on silica gel-coated
plates; the spots were visualized by spraying the plates with 2%
ride 23 with thioglycoside 19 in the presence of N-iodo-
Ce(SO4)2 in 4 M H2SO4 and then warming the plates on a hotplate.
succinimide and trimethylsilyl trifluoromethanesulfonate
Silica gel (230–400 mesh) was used for column chromatography.
gave trisaccharide derivative 24 in 89% yield. The forma-
1H and 13C NMR, 2D COSY, and HSQC spectra were recorded on
tion of trisaccharide 24 was confirmed by the presence of
a Bruker Avance DPX at 300 MHz using CDCl3 and D2O as sol-
signals in at d = 99.3 (C-1B), 99.2 (C-1C), 98.5 (C-1A) in
the 13C NMR spectrum. Removal of the allyl group from
compound 24 by using palladium chloride in methanol
gave trisaccharide acceptor 25 in 87% yield. Glycosyla-
tion of this product with thioglycoside 20 in the presence
of N-iodosuccinimide and trimethylsilyl trifluoro-
methanesulfonate gave the tetrasaccharide derivative 26
in 83% yield. This was deprotected by hydrogenation and
saponification to give the tetrasaccharide methyl glyco-
side 2 in 79% overall yield. Tetrasaccharide 2 was puri-
fied by passage through a column of LH-20 Sephadex
using methanol–water (4:1) as the eluant. The formation
of compound 2 was confirmed by the presence of signals
at d = 5.04 (br s, H-1A), 5.01 (br s, H-1C), 4.76 (d, J = 7.8
Hz, H-1D), and 4.57 (br s, H-1B) in the 1H NMR and at d
= 104.2 (C-1D), 104.0 (C-1A), 103.9 (C-1C), 102.6 (C-1B)]
in the 13C NMR spectrum (Scheme 3).
vents and TMS as the internal reference, unless otherwise stated.
Chemical shifts are expressed in d (ppm). ESI-MS were recorded on
a Micromass Quattro II triple quadrupole mass spectrometer. Ele-
mentary analysis was carried out on Carlo ERBA-1108 analyzer.
Optical rotations were measured at 25 °C on a Rudolf Autopol III
polarimeter. Commercially available grades of organic solvents of
adequate purity were used in most reactions.
Methyl 2-Azido-3-O-benzyl-4,6-O-benzylidene-2-deoxy-a-D-
mannopyranoside (5)
Pyridine (5 mL) was added to a soln of compound 4 (1.7 g, 4.57
mmol) in anhyd CH2Cl2 (10 mL) and the mixture was cooled to
–10 °C. Triflic anhydride (1.6 mL, 9.51 mmol) was added and the
cold mixture was stirred at –10 °C for 1 h. The solvents were re-
moved under reduced pressure and the crude product was used di-
rectly in the next step.
NaN3 (1.5 g, 23 mmol) was added to a soln of the crude product in
HMPT (5 mL), and the mixture was stirred at 70 °C for 24 h. The
mixture was then poured into cold H2O and extracted with EtOAc
(100 mL). The organic layer was washed with sat. aq NaHCO3,
dried (Na2SO4), and concentrated under reduced pressure. The
crude product was purified by chromatography [hexane–EtOAc
(3:1)] to give pure 5 as a yellow oil; yield: 1.5 g (82%); [a]D +93 (c
1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.47–7.17 (m, 10 H, Ar-H), 5.59
(s, 1 H, PhCH) 4.89–4.85 (d, J = 12.2 Hz, 1 H, PhCH2), 4.73–4.69
(d, J = 12.2 Hz, 1 H, PhCH2), 4.62 (s, 1 H, H-1), 4.25–4.20 (dd,
J = 9.4, 3.9 Hz, 1 H, H-6a), 4.12–4.03 (m, 2 H, H-3, H-5), 3.93 (br
s, 1 H, H-2), 3.80 (d, J = 9.6 Hz, 1 H, H-4), 3.85–3.73 (m, 1 H, H-
6b), 3.35 (s, 3 H, OCH3).
In summary, efficient synthetic strategies have been de-
veloped for the synthesis of the methyl glycoside of an
Azospirillum trisaccharide containing a D-mannosamine
moiety and an Azospirillum tetrasaccharide containing an
L-rhamnan chain. All the intermediate steps are high
yielding, and can be used in large-scale preparations.
OMe
O
c
a
BnO
17 + 18
O
OBn
13C NMR (75 MHz, CDCl3): d = 138.1–126.1 (Ar-C), 101.7
(PhCH), 100.2 (C-1), 79.2 (C-3), 75.6 (C-5), 73.4 (PhCH2), 68.7 (C-
6), 63.8 (C-4), 62.8 (C-2), 55.0 (OCH3).
O
BnO
b
RO
OR
21: R = Ac
22: R = H
MS (EI, 70 eV): 420.1 [M + Na]+.
Anal. Calcd for C21H23N3O5: C, 63.46; H, 5.83. Found: C, 63.30; H,
6.0.
OMe
OMe
O
O
O
BnO
BnO
a
O
OBn
OBn
Methyl 2-Azido-6-O-benzoyl-3-O-benzyl-2-deoxy-a-D-manno-
pyranoside (7)
O
O
BnO
BnO
10% HClO4/silica gel (200 mg) was added to a soln of compound 5
(1.5 g, 3.77 mmol) in MeCN (20 mL), and the reaction was stirred
at r.t. for 30 min then filtered through a Celite bed and evaporated
to dryness. The crude mass was dissolved in anhyd CH2Cl2 (10 mL)
then pyridine (5 mL) and BzCN (500 mg, 3.81 mmol) were added
sequentially. The mixture was stirred at r.t. for 2 h and concentrated
under reduced pressure to give a crude product that was purified by
chromatography [silica gel, hexane–EtOAc (5:1)] to give pure com-
pound 7 as a yellow oil; yield: 1.4 g (90%); [a]D +62 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 8.07–7.29 (m, 10 H, Ar-H), 4.76
(d, J = 11.5 Hz, 1 H, PhCH2), 4.70 (br s, 1 H, H-1), 4.64 (d, J = 11.5
Hz, 1 H, PhCH2), 4.61–4.52 (m, 2 H, H-6ab), 3.95–3.79 (m, 4 H, H-
2, H-4, H-5, and H-3), 3.37 (s, 3 H, OCH3).
O
HO
OR
OAllyl
23
O
AcO
AcO
OAc
24: R = Allyl
25: R = H
d
OMe
O
BnO
O
OBn
AcO
e
20
2
O
BnO
a
OAc
OAc
O
O
O
O
AcO
AcO
13C NMR (75 MHz, CDCl3): d = 166.6 (PhCO), 137.5–128.0 (Ar-
C), 99.4 (C-1), 79.2 (C-3), 72.6 (PhCH2), 70.7 (C-5), 66.7 (C-4),
63.8 (C-6), 60.5 (C-2), 55.2 (OCH3).
AcO
OAc
26
Scheme 3 Reagents and conditions: (a) NIS, TMSOTf, CH2Cl2, –30 °C,
1.5 h, 90% for 21, 89% for 24, and 83% for 26; (b) MeONa, MeOH,
r.t., 5 h, quant; (c) allyl bromide, 5% aq NaOH, CH2Cl2, Bu4NHSO4,
r.t., 3 h, 84%; (d) PdCl2, MeOH, r.t., 2 h, 87%; (e) H2, 20% Pd(OH)2/C,
r.t., 12 h then MeONa, MeOH, r.t., 8 h, 79%.
MS (EI, 70 eV): 436.1 [M + Na]+.
Anal. Calcd for C21H23N3O6: C, 61.01; H, 5.61. Found: C, 60.85; H,
5.80.
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York

PAPER
Oligosaccharides of Azospirillum brasilense
2587
Ethyl 3,4-Di-O-benzyl-2-O-methyl-1-thio-a-L-rhamnopyrano-
side (10)
in Ac2O–pyridine (5 mL, 1:1) was kept at r.t. for 2 h. The solvents
were removed under reduced pressure and the crude product was
purified by chromatography [silica gel, hexane–EtOAc (3:1)] to
give pure compound 13 as a yellow oil; yield: 1.2 g (91%); [a]D
+108 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.87–7.27 (m, 14 H, Ar-H), 5.74
(d, J = 8.5 Hz, 1 H, H-1B), 5.75–5.72 (m, 1 H, NHCOCH3), 5.67–
5.60 (dd, J = 9.3, 9.3 Hz, 1 H, H-3B), 5.06 (t, J = 9.5 Hz, 1 H, H-4B),
4.66–4.63 (m, 2 H, H-1A, PhCH2), 4.52–4.44 (m, 3 H, H-2A, H-6aA,
PhCH2), 4.32–4.26 (dd, J = 8.5, 8.5 Hz, 1 H, H-2B), 4.10–4.03 (m,
2 H, H-3A, H-6bA), 3.96–3.91 (m, 3 H, H-4A, H-6abB), 3.85–3.82 (m,
1 H, H-5A), 3.52–3.46 (m, 1 H, H-5B), 3.29 (s, 3 H, OCH3), 2.0,
1.97, 1.92, 1.79 (4 s, 12 H, 4 COCH3).
13C NMR (75 MHz, CDCl3): d = 170.4, 169.9, 169.8, 169.0 (4
COCH3), 167.7, 166.8 (2 CO; Phth), 165.2 (PhCO), 137.8–123.5
(Ar-C), 99.6 (C-1A), 97.5 (C-1B), 75.1 (C-3A), 73.7 (C-4A), 71.8 (C-
5B), 70.8 (C-3B), 70.4 (PhCH2), 68.4 (C-4B), 68.2 (C-5A), 55.1
(OCH3), 55.0 (C-2B), 49.1 (C-2A), 23.1 (NHCOCH3), 20.6, 20.5,
20.3 (3 COCH3).
Crushed NaOH (500 mg, 12.5 mmol) and MeI (1.0 mL, 16 mmol)
were added to a soln of thioglycoside 9 (1.5 g, 3.86 mmol) in anhyd
THF (20 mL), and the mixture was stirred at 0–5 °C for 4 h. The
mixture was then diluted with H2O and extracted with CH2Cl2 (100
mL). The organic layer was washed with H2O, dried (Na2SO4), and
concentrated to dryness. The crude product was purified by chroma-
tography [silica gel, hexane–EtOAc (10:1)] to give pure compound
10 as a yellow oil; yield: 1.4 g (90%); [a]D –134 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.36–7.24 (m, 10 H, Ar-H), 5.27
(br s, 1 H, H-1), 4.93–4.89 (d, J = 11.0 Hz, 1 H, PhCH2), 4.71–4.66
(m, 2 H, PhCH2), 4.62–4.59 (d, J = 11.0 Hz, 1 H, PhCH2), 4.03–
3.91 (m, 1 H, H-5), 3.77–3.73 (dd, J = 9.4, 3.1 Hz, 1 H, H-3), 3.53–
3.50 (m, 2 H, H-2, H-4), 3.47 (s, 3 H, OCH3), 2.66–2.52 (m, 2 H,
SCH2CH3), 1.30–1.25 (m, 6 H, SCH2CH3, CH3).
13C NMR (75 MHz, CDCl3): d = 138.7–127.5 (Ar-C), 81.2 (C-1)
80.6 (C-4), 80.2 (PhCH2), 79.8 (C-2), 75.3 (PhCH2), 72.3 (C-3),
68.3 (C-5), 58.5 (OCH3), 25.3 (SCH2CH3), 17.9 (CCH3), 15.0
(SCH2CH3).
MS (EI, 70 eV): 869.3 [M + Na]+.
MS (EI, 70 eV): 425.2 [M + Na]+.
Anal. Calcd for C43H46N2O16: C, 60.99; H, 5.48. Found: C, 60.82;
H, 5.64.
Anal. Calcd for C23H30O4S: C, 68.62; H, 7.51. Found: C, 68.45; H,
7.72.
Methyl 4-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-b-D-glucopy-
ranosyl)-2-acetamido-6-O-acetyl-3-O-benzyl-2-deoxy-a-D-man-
nopyranoside (14)
Methyl 4-(3,4,6-Tri-O-acetyl-2-deoxy-2-N-phthalimido-b-D-
glucopyranosyl)-2-azido-6-O-benzoyl-3-O-benzyl-2-deoxy-a-D-
mannopyranoside (12)
H2N(CH2)2NH2 (0.8 mL, 12 mmol) was added to a soln of aceta-
mide 13 (1.1 g, 1.3 mmol) in BuOH (20 mL), and the mixture was
stirred at 110 °C for 6 h. The solvents were removed under reduced
pressure and a soln of the crude product in Ac2O–py (10 mL, 1:1)
was kept at r.t. for 2 h. The solvents were removed under reduced
pressure, and the crude product was purified by chromatography
[silica gel, hexane–EtOAc (3:1)] to give pure compound 14 as a yel-
low oil; yield: 770 mg (85%); [a]D +28 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.38–7.28 (m, 5 H, Ar-H), 5.92 (t,
J = 9.2 Hz, 2 H, 2 NHCOCH3), 5.24 (t, J = 10.2 Hz, 1 H, H-3B),
5.05–4.89 (m, 2 H, H-1B, H-4B), 4.74–4.71 (m, 2 H, H-1A, PhCH2),
4.59–4.55 (m, 1 H, H-2A), 4.49 (d, J = 11.4 Hz, 1 H, PhCH2), 4.41–
4.38 (dd, J = 11.8, 1.6 Hz, 1 H, H-6bA), 4.31–4.25 (dd, J = 11.8, 5.6
Hz, 1 H, H-6bA), 4.07–4.02 (m, 2 H, H-3A, H-6aB), 3.92–3.83 (m, 2
H, H-3A, H-6bB), 3.77–3.69 (m, 2 H, H-2B, H-5A), 3.47–3.44 (m, 1
H, H-5B), 3.35 (s, 3 H, OCH3), 2.13, 2.02, 1.99, 1.87 (4 s, 18 H, 6
COCH3).
A soln of azide 7 (1.0 g, 2.42 mmol) and thioglycoside 11 (1.4 g,
2.92 mmol) in anhyd CH2Cl2 (20 mL) was treated with 4-Å MS (2
g), and the mixture was stirred under argon for 1 h. The mixture was
then cooled to –20 °C, and NIS (850 mg, 3.77 mmol) and TMSOTf
(15 mL) were added sequentially. The mixture was stirred at –20 °C
for 1.5 h then diluted with CH2Cl2 (100 mL) and filtered through a
Celite bed. The organic layer was washed successively with sat. aq
NaHCO3 and H2O, dried (Na2SO4), and concentrated under reduced
pressure. The crude product was purified by chromatography [silica
gel, hexane–EtOAc (3:1)] to give pure disaccharide derivative 12 as
a yellow oil; yield: 1.8 g (90%); [a]D +152 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.80–7.28 (m, 14 H, Ar-H), 5.72
(d, J = 8.5 Hz, 1 H, H-1B), 5.66 (d, J = 9.5 Hz, 1 H, H-3B), 5.11 (t,
J = 9.5 Hz, 1 H, H-4B), 4.84 (d, J = 12.3 Hz, 1 H, PhCH2), 4.79 (d,
J = 12.3 Hz, 1 H, PhCH2), 4.58 (d, J = 1.6 Hz, 1 H, H-1A), 4.77–
4.43 (dd, J = 12.3, 1.6 Hz, 1 H, H-6aA), 4.31–4.24 (dd, J = 8.5 Hz
each, 1 H, H-2B), 4.14 (d, J = 9.1 Hz, 1 H, H-4A), 4.10–4.01 (m, 2
H, H-3A, H-6aB), 3.94–3.83 (m, 2 H, H-6bA, H-6bB), 3.79–3.74 (m,
2 H, H-2A, H-5A), 3.57–3.54 (m, 1 H, H-5B), 3.28 (s, 3 H, OCH3),
2.04, 1.98, 1.81 (3 s, 9 H, 3 COCH3).
13C NMR (75 MHz, CDCl3): d = 170.7, 170.6, 170.5, 170.3, 169.3,
169.1 (6 COCH3), 137.8–127.3 (Ar-C), 99.9 (C-1B), 99.7 (C-1A),
75.3 (C-3A), 74.4 (C-5A), 72.2 (C-3B), 71.7 (C-5B), 70.8 (PhCH2),
68.5 (C-4A), 68.2 (C-4B), 63.3 (C-6A), 61.7 (C-6B), 55.3 (C-2B), 55.1
(OCH3), 49.3 (C-2A), 23.3, 23.2 (2 NHCOCH3), 20.9, 20.6, 20.5 (2
C) (4 COCH3).
13C NMR (75 MHz, CDCl3): d = 170.3, 169.8, 169.1 (3 COCH3),
167.6, 166.7 (2 CO; Phth), 165.3 (PhCO), 137.9–123.6 (Ar-C), 98.9
(C-1A), 98.5 (C-1B), 78.1 (C-3A), 74.7 (C-4A), 71.9 (PhCH2), 71.7
(C-5B), 70.7 (C-3B), 68.9 (C-5A), 68.4 (C-4B), 62.5 (C-6A), 61.3 (C-
6B), 60.6 (C-2A), 55.2 (C-2B), 55.0 (OCH3), 20.6, 20.5, 20.3 (3 C,
COCH3).
MS (EI, 70 eV): 719.3 [M + Na]+.
Anal. Calcd for C32H44N2O15: C, 55.17; H, 6.37. Found: C, 55.0; H,
6.55.
MS (EI, 70 eV): 853.2 [M + Na]+.
Methyl 4-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-b-D-gluco-
pyranosyl)-2-acetamido-6-O-acetyl-2-deoxy-a-D-mannopyra-
noside (15)
Anal. Calcd for C41H42N4O15: C, 59.27; H, 5.10. Found: C, 59.10;
H, 5.26.
20% Pd(OH)2-C (100 mg) was added to a soln of disaccharide 14
(750 mg, 1.07 mmol) in MeOH (10 mL), and the mixture was stirred
under a positive pressure of H2 at r.t. for 10 h. The mixture was then
filtered through a Celite bed and the solvents were evaporated to
dryness. The crude product was purified by chromatography [silica
gel, hexane–EtOAc (2:1)] to give pure compound 15 as a yellow oil;
yield: 600 mg (92%); [a]D +7.7 (c 1.2, CHCl3).
Methyl 4-(3,4,6-Tri-O-acetyl-2-deoxy-2-N-phthalimido-b-D-
glucopyranosyl)-2-acetamido-6-O-benzoyl-3-O-benzyl-2-
deoxy-a-D-mannopyranoside (13)
20% Pd(OH)2/C (100 mg) and TES (3.2 mL, 50 mmol) were added
to a soln of azide 12 (1.3 g, 1.56 mmol) in MeOH–CHCl3 (5:1) un-
der argon, and the mixture was stirred at r.t. for 6 h, filtered through
a Celite bed, and evaporated to dryness. A soln of the crude product
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York

2588
S. Pandey et al.
PAPER
1H NMR (300 MHz, CDCl3): d = 6.32 (d, J = 8.8 Hz, 1 H, NHCH3),
5.76 (d, J = 6.51 Hz, 1 H, NHCOCH3), 5.31–5.24 (m, 1 H, H-3B),
5.00 (t, J = 9.6 Hz, 1 H, H-4B), 4.83 (br s, 1 H, H-1A), 4.70 (d,
J = 8.4 Hz, 1 H, H-1B), 4.44–4.41 (m, 1 H, H-6aB), 4.33–4.31 (m, 1
H, H-2A), 4.27–4.19 (m, 2 H, H-6aA, H-6bB), 4.15–4.07 (m, 2 H, H-
3A, H-6bA), 3.97–3.93 (m, 1 H, H-2B), 3.84–3.77 (m, 2 H, H-4A, H-
5B), 3.35 (s, 3 H, OCH3), 3.38–3.32 (m, 1 H, H-5A), 2.16, 2.08, 2.03,
2.02, 1.97 (5 s, 18 H, 6 COCH3).
13C NMR (75 MHz, CDCl3): d = 171.4, 170.8, 170.7, 169.3, 169.1,
169.0 (6 COCH3), 101.4 (C-1B), 99.4 (C-1A), 80.2 (C-5A), 72.1 (C-
3B), 71.9 (C-4A), 68.6 (C-4B), 67.5 (C-5B), 67.3 (C-3A), 62.7 (C-6A),
62.1 (C-6B), 55.1 (OCH3), 54.6 (C-2B), 52.3 (C-2A), 23.3, 23.1 (2
NHCOCH3), 20.9, 20.6, 20.5, 20.4 (4 COCH3).
ed to dryness to give crude target compound 1, which was purified
as an amorphous powder by chromatography [LH-20 Sephadex,
MeOH–H2O (4:1)]; yield: 360 mg (81%); [a]D +20 (c 1.0, H2O).
1H NMR (300 MHz, CDCl3): d = 4.94 (br s, 1 H, H-1C), 4.63 (br s,
1 H, H-1A), 4.56 (d, J = 7.5 Hz, 1 H, H-1B), 4.50–4.49 (m, 1 H, H-
2A), 4.31–4.21 (m, 1 H, H-5C), 4.04–4.00 (m, 1 H, H-3A), 3.97–3.84
(m, 5 H, H-4A, H-4B, H-6abB, H-6aA), 3.70–3.65 (m, 2 H, H-2B, H-
5B), 3.62–3.59 (m, 3 H, H-3C, H-5B, H-6bA), 3.46 (s, 3 H, OCH3),
3.37 (s, 3 H, OCH3), 3.34–3.22 (m, 3 H, H-2C, H-4C, H-5A), 2.05,
2.02 (2 s, 6 H, 2 COCH3), 1.31 (d, J = 6.3 Hz, 3 H, CCH3).
13C NMR (75 MHz, CDCl3): d = 173.7, 173.4 (2 COCH3), 102.4 (C-
1B), 101.4 (C-1A), 94.5 (C-1C), 82.0 (C-2C), 77.6 (C-5A), 75.3 (C-
5B), 74.5 (C-4C), 73.1 (C-3C), 72.6 (C-3B), 72.4 (C-3A) 71.6 (2 C, C-
4A, C-4B), 69.5 (C-5C), 62.3 (C-6B), 61.4 (C-6A), 59.1 (OCH3), 58.0
(C-2B), 55.4 (OCH3), 49.9 (C-2A), 22.9, 22.6 (2 COCH3), 17.8
(CCH3).
MS (EI, 70 eV): 629.2 [M + Na]+.
Anal. Calcd for C25H38N2O15: C, 49.50; H, 6.31. Found: C, 49.34;
H, 6.50.
MS (EI, 70 eV): 621.7 [M + Na]+.
Methyl (2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-b-D-glucopyr-
anosyl)-(1→4)-[(3,4-di-O-benzyl-2-O-methyl-a-L-rhamnopyra-
nosyl)-(1→3)]-2-acetamido-6-O-acetyl-2-deoxy-a-D-
mannopyranoside (16)
Anal. Calcd for C24H42N2O15: C, 48.16; H, 7.07. Found: C, 47.97;
H, 7.24.
Methyl 3-(2,3-Di-O-acetyl-4-O-benzyl-a-L-rhamnopyranosyl)-
2,4-di-O-benzyl-a-L-rhamnopyranoside (21)
A soln of acetamide 15 (550 mg, 0.9 mmol) and thioglycoside 10
(540 mg, 1.34 mmol) in anhyd CH2Cl2 (10 mL) was treated with 4-
Å MS (1 g), and the mixture was stirred at r.t. under argon for 1 h.
The mixture was then cooled to –10 °C then NIS (360 mg, 1.6
mmol) and TMSOTf (5 mL) were added successively. The mixture
was stirred at –10 °C for 1.5 h, diluted with CH2Cl2 (100 mL), and
filtered through a Celite bed. The organic layer was washed succes-
sively with sat. aq NaHCO3 and H2O, dried (Na2SO4), and concen-
trated under reduced pressure. The crude product was purified by
chromatography [silica gel, hexane–EtOAc (3:1)] to give pure com-
pound 16 as a yellow oil; yield: 760 mg (89%); [a]D –1.4 (c 1.2,
CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.40–7.26 (m, 10 H, Ar-H), 6.32
(d, J = 8.8 Hz, 1 H, NHCOCH3), 5.83 (d, J = 6.8 Hz, 1 H,
NHCOCH3), 5.19 (t, J = 9.6 Hz, 1 H, H-3B), 5.10 (br s, 1 H, H-1C),
4.96–4.89 (m, 2 H, H-4B, PhCH2), 4.81 (d, J = 8.3 Hz, 1 H, H-1B),
4.71–4.62 (m, 3 H, PhCH2), 4.55 (br s, 1 H, H-1A), 4.53–4.48 (m, 1
H, H-6aB), 4.33 (br s, 2 H, H-2A, H-3A), 4.17–4.15 (m, 1 H, H-2B),
4.12–4.08 (m, 1 H, H-6bB), 3.96–3.88 (m, 1 H, H-5C), 3.85–3.76 (m,
4 H, H-3C, H-4A, H-6abA), 3.64–3.59 (m, 2 H, H-4C, H-5B), 3.54–
3.84 (m, 1 H, H-5A), 3.50 (s, 3 H, OCH3), 3.41 (br s, 1 H, H-2C), 3.35
(s, 3 H, OCH3), 2.16, 2.02, 2.0, 1.95, 1.79 (5 s, 18 H, 6 COCH3),
1.30 (d, J = 6.4 Hz, 3 H, CCH3).
A soln of monodeprotected compound 17 (1.4 g, 3.9 mmol) and
thioglycoside 18 (1.8 g, 4.7 mmol) in anhyd CH2Cl2 (20 mL) was
treated with 4-Å MS (2 g), and the mixture was stirred at r.t. under
argon for 1 h. The mixture was then cooled to –10 °C, and NIS (1.3
g, 5.77 mmol) and TMSOTf (20 mL) were added sequentially. The
mixture was stirred at –10 °C for 1.5 h then diluted with CH2Cl2
(100 mL) and filtered through a Celite bed. The organic layer was
washed successively with sat. aq NaHCO3 and H2O, dried
(Na2SO4), and concentrated under reduced pressure. The crude
product was purified by chromatography [silica gel, hexane–EtOAc
(4:1)] to give pure compound 21 as a yellow oil; yield: 2.4 g (90%);
[a]D –47 (c 1.1, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.41–7.18 (m, 15 H, Ar-H), 5.43–
5.37 (m, 2 H, H-2B, H-3B), 5.02 (s, 1 H, H-1B), 4.81–4.57 (m, 7 H,
H-1A, 3 PhCH2), 4.04–4.01 (m, 1 H, H-5A), 3.89–3.80 (m, 1 H, H-
5B), 3.67–3.62 (m, 3 H, H-2A, H-3A, H-4B), 3.47–3.41 (t, J = 9.4 Hz,
1 H, H-4A), 3.29 (s, 3 H, OCH3), 2.01, 1.98 (2 s, 6 H, 2 COCH3),
1.30, 1.27 (2 d, J = 6.3 Hz each, 6 H, 2 CCH3).
13C NMR (75 MHz, CDCl3): d = 169.4, 169.3 (2 COCH3), 138.5–
127.3 (Ar-C), 98.9 (C-1A), 98.4 (C-1B), 80.7 (C-4B), 78.7 (C-4A),
77.7 (C-2B), 77.4 (C-3A), 75.1 (C-2A), 74.6 (PhCH2), 72.6 (PhCH2),
71.5 (PhCH2), 70.2 (C-3B), 68.0 (C-5A), 67.9 (C-5B), 54.5 (OCH3),
20.7, 20.6 (2 COCH3), 18.0, 17.9 (2 CCH3).
13C NMR (75 MHz, CDCl3): d = 170.7, 170.4 (2 C), 169.9, 169.8,
169.0 (6 COCH3), 138.6–127.6 (Ar-C), 100.2 (2 C, C-1B, C-1C),
99.5 (C-1A), 80.6 (C-4C, C-5A), 79.2 (C-3B), 77.6 (C-2C), 75.6
(PhCH2), 72.4 (C-5B), 71.9 (2 C, C-3A, C-4A), 71.5 (PhCH2), 69.3
(C-3C), 69.1 (C-4B), 67.7 (C-5C), 63.2 (C-6A), 62.3 (C-6B), 58.9
(OCH3), 55.1 (OCH3), 54.5 (C-2B), 49.0 (C-2A), 23.2, 23.1 (2
NHCOCH3), 20.8, 20.7 (2 C), 20.6 (4 COCH3), 17.9 (CCH3).
MS (EI, 70 eV): 701.3 [M + Na]+.
Anal. Calcd for C38H46O11: C, 67.24; H, 6.83. Found: C, 67.07; H,
7.0.
Methyl 3-(2-O-Allyl-4-O-benzyl-a-L-rhamnopyranosyl)-2,4-di-
O-benzyl-a-L-rhamnopyranoside (23)
MS (EI, 70 eV): 969.3 [M + Na]+.
A soln of compound 21 (2.3 g, 3.38 mmol) in 0.1 M methanolic
NaOMe (50 mL) was stirred at r.t. for 3 h and then neutralized with
Dowex 50W-X8 (H+) resin. The mixture was filtered and evaporat-
ed to dryness. To a soln of the crude product 22 in CH2Cl2 (25 mL)
were added 5% aq NaOH (15 mL), allyl bromide (440 mL, 5.08
mmol), and Bu4NHSO4 (100 mg), and the biphasic reaction mixture
was stirred briskly at r.t. for 3 h. The mixture was diluted with
CH2Cl2 (100 mL), and the organic layer was washed with H2O,
dried (Na2SO4), and concentrated under reduced pressure. The
crude product was purified by chromatography [silica gel, hexane–
EtOAc (4:1)] to give pure compound 23 as a yellow oil; yield: 1.8 g
(84%); [a]D –58 (c 1.2, CHCl3).
Anal. Calcd for C46H62N2O19: C, 58.34; H, 6.60. Found: C, 58.17;
H, 6.72.
Methyl (2-Acetamido-2-deoxy-b-D-glucopyranosyl)-(1→4)-[(2-
O-methyl-a-L-rhamnopyranosyl)-(1→3)]-2-acetamido-2-
deoxy-a-D-mannopyranoside (1)
20% Pd(OH)2/C (150 mg) was added to a soln of the protected com-
pound 16 (700 mg, 0.74 mmol) in MeOH (10 mL), and the mixture
was stirred under a positive pressure of H2 at r.t. for 10 h. The mix-
ture was then filtered through a Celite bed and the solvents were
evaporated to dryness. A soln of the crude product in 0.1 M metha-
nolic NaOMe (5 mL) was stirred at r.t. for 6 h and then neutralized
with Dowex 50W X-8 (H+). The mixture was filtered and evaporat-
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York

PAPER
Oligosaccharides of Azospirillum brasilense
2589
1H NMR (300 MHz, CDCl3): d = 7.39–7.22 (m, 15 H, Ar-H), 5.78–
5.65 (m, 1 H, CH2=CH), 5.09 (br s, 1 H, H-1B), 5.08–5.02 (m, 2 H,
CH=CH2), 4.93–4.89 (d, J = 11.3 Hz, 1 H, PhCH2), 4.81–4.78 (d,
J = 11.7, Hz, 1 H, PhCH2), 4.73–4.63 (m, 7 H, H-1A, 3 PhCH2),
4.09–4.05 (dd, J = 12.3, 3.0 Hz, 1 H, H-3A), 4.01–3.93 (m, 1 H, H-
3B), 3.82–3.57 (m, 7 H, H-2A, H-2B, H-4A, H-5A, H-5B, OCH2–
CH=CH2), 3.31 (s, 3 H, OCH3), 3.30–3.24 (m, 1 H, H-4B), 1.32,
1.26 (2 d, J = 6.4 Hz each, 6 H, 2 CCH3).
13C NMR (75 MHz, CDCl3): d = 138.8–126.8 (Ar-C), 117.3 (–CH=CH2),
98.9 (C-1A), 98.7 (C-1B), 82.2 (C-4B), 80.9 (C-4A), 78.8 (C-2B), 77.8
(C-3A), 76.6 (C-2A), 74.8 (PhCH2), 74.7 (PhCH2), 72.7 (PhCH2),
71.6 (OCH2–CH=CH2), 71.5 (C-3B), 68.1 (C-5A), 67.7 (C-5B), 54.6
(OCH3), 18.0, 17.9 (2 CCH3).
(m, 1 H, H-2A), 3.63–3.20 (m, 2 H, H-3B, H-4A), 3.46 (t, J = 9.3 Hz,
1 H, H-4B), 3.31 (s, 3 H, OCH3), 2.08, 2.06, 1.97 (3 s, 9 H, 3
COCH3), 1.29, 1.24, 1.10 (3 d, J = 6.6 Hz each, 9 H, 3 CCH3).
13C NMR (75 MHz, CDCl3): d = 169.7, 169.6, 169.5 (3 COCH3),
138.2–127.6 (Ar-C), 101.3 (C-1B), 99.3 (C-1C), 98.4 (C-1A), 80.9
(C-4A), 80.1 (C-4B), 80.0 (C-5A), 78.3 (C-5C), 77.9 (C-2A), 75.3 (2
C, PhCH2), 72.6 (PhCH2), 71.3 (C-3A), 70.9 (C-3C), 69.8 (C-2C),
69.2 (C-4C), 68.2 (C-3B), 68.0 (C-5B), 67.1 (C-2B), 54.7 (OCH3),
20.8, 20.7 (2 C) (3 COCH3), 18.1, 18.0, 17.4 (3 CCH3).
MS (EI, 70 eV): 889.4 [M + Na]+.
Anal. Calcd for C46H58O16: C, 63.73; H, 6.74. Found: C, 63.55; H,
6.90.
MS (EI, 70 eV): 657.3 [M + Na]+.
Methyl (2,3,4-Tri-O-acetyl-a-L-rhamnopyranosyl)-(1→3)-
[(2,3,4,6-tetra-O-acetyl-b-D-glucopyranosyl)-(1→2)]-(4-O-ben-
zyl-a-L-rhamnopyranosyl)-(1→3)-2,4-di-O-benzyl-a-L-rham-
nopyranoside (26)
Anal. Calcd for C37H46O9: C, 70.01; H, 7.30. Found: C, 69.84; H,
7.48.
Methyl (2,3,4-Tri-O-acetyl-a-L-rhamnopyranosyl)-(1→3)-(2-O-
allyl-4-O-benzyl-a-L-rhamnopyranosyl)-(1→3)-2,4-di-O-ben-
zyl-a-L-rhamnopyranoside (24)
A soln of monodeprotected derivative 25 (1.4 g, 1.61 mmol) and
thioglycoside 20 (950 mg, 2.42 mmol) in anhyd CH2Cl2 (20 mL)
was treated with 4-Å MS (2 g), and the soln was stirred under argon
for 1 h. The mixture was cooled to –10 °C then NIS (650 mg, 2.88
mmol) and TMSOTf (10 mL) were added successively. The mixture
was stirred at –10 °C for 1.5 h, diluted with CH2Cl2 (100 mL), and
filtered through a Celite bed. The organic layer was washed succes-
sively with sat. aq NaHCO3 and H2O, dried (Na2SO4), and concen-
trated under reduced pressure. The crude product was purified by
chromatography [silica gel, hexane–EtOAc (4:1)] to give pure com-
pound 26 as a yellow oil; yield: 1.6 g (83%); [a]D –12 (c 1.2,
CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.40–7.22 (m, 15 H, Ar-H), 5.63
(d, J = 5.1 Hz, 1 H, H-2D), 5.31–5.28 (m, 1 H, H-2C), 5.21–5.17 (m,
2 H, H-3C, H-3D), 5.15 (br s, 1 H, H-2C), 5.01–4.94 (m, 2 H, H-1B,
H-4C), 4.88–4.60 (m, 8 H, H-1A, H-1D, 3 PhCH2), 4.44–4.42 (m, 1
H, H-4D), 4.22–4.17 (m, 3 H, H-3B, H-6abD), 4.04–4.01 (m, 1 H, H-
3A), 3.94–3.75 (m, 5 H, H-2A, H-2B, H-4B, H-5B, H-5C), 3.69–3.52
(m, 3 H, H-4A, H-5A, H-5D), 3.34 (s, 3 H, OCH3), 2.14, 2.12, 2.10,
2.08, 2.04, 1.96 (6 s, 21 H, 7 COCH3), 18.2, 18.0, 17.5 (3 d, J = 6.4
Hz each, 9 H, 3 CCH3).
A soln of allyl derivative 23 (1.5 g, 2.36 mmol) and thioglycoside
19 (1.0 g, 3.0 mmol) in anhyd CH2Cl2 (20 mL) was added 4-Å MS
(2 g), and the soln was stirred under argon for 1 h. The reaction mix-
ture was cooled to –10 °C, and NIS (560 mg, 2.48 mmol) and TM-
SOTf (10 mL) were added sequentially. The mixture was stirred at
–10 °C for 1.5 h then diluted with CH2Cl2 (100 mL) and filtered
through a Celite bed. The organic layer was washed successively
with sat. aq NaHCO3 and H2O, dried (Na2SO4), and concentrated
under reduced pressure. The crude product was purified by chroma-
tography [silica gel, hexane–EtOAc (4:1)] to give pure compound
24 as a yellow oil; yield: 1.9 g (89%); [a]D –35 (c 1.2, CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.36–7.19 (m, 15 H, Ar-H), 5.77–
5.72 (m, 1 H, CH2=CH–), 5.34–5.27 (m, 2 H, H-2C, H-4C), 5.15–
4.96 (m, 5 H, H-1B, H-1C, H-3C, CH2=CH–), 4.81–4.58 (m, 7 H, H-
1A, 3 PhCH2), 4.04–3.99 (m, 2 H, H-3B, H-5C), 3.97–3.94 (m, 1 H,
H-4A), 3.90–3.84 (m, 1 H, H-3A), 3.74–3.62 (m, 5 H, H-2A, H-2B, H-
5B, OCH2–CH=CH2), 3.59–3.53 (m, 2 H, H-4B, H-5A), 3.31 (s, 3 H,
OCH3), 2.07, 2.02, 1.97 (3 s, 9 H, 3 COCH3), 1.27, 1.24, 1.11 (3 d,
J = 6.4 Hz each, 9 H, 3 CCH3).
13C NMR (75 MHz, CDCl3): d = 170.3, 169.7, 169.5, 169.4, 169.2,
168.8, 168.7 (7 COCH3), 138.2–127.4 (Ar-C), 98.6 (C-1B), 98.0 (C-
1C), 97.2 (C-1D), 96.9 (C-1A), 81.1 (C-5A), 80.0 (C-4B), 78.3 (C-4A),
74.7 (C-5D), 73.3 (2 C, C-5B, C-5C), 72.9 (C-3A), 72.4, 72.3, 70.8 (3
PhCH2), 69.9 (C-2C), 69.7 (C-4D), 69.5 (C-2A), 68.9 (C-3C), 68.2 (C-
3D), 68.1 (C-4C), 68.0 (C-3B), 67.1 (C-2B), 67.0 (C-2D), 63.2 (C-6D),
54.6 (OCH3), 20.8 (2 C), 20.7 (2 C), 20.5 (2 C), 20.4 (7 COCH3),
18.3, 18.1, 17.6 (3 CCH3).
13C NMR (75 MHz, CDCl3): d = 169.6, 169.5, 169.4 (3 COCH3),
138.5–126.8 (Ar-C), 99.3 (C-1B), 99.2 (C-1C), 98.5 (C-1A), 80.8 (C-
5A), 80.5 (C-4B), 78.8 (C-3A), 78.3 (C-3B), 78.1 (C-5C), 78.0 (C-5B),
75.2 (PhCH2), 74.7 (PhCH2), 72.7 (PhCH2), 71.5 (OCH2-
CH = CH2), 71.1 (C-1C), 69.9 (C-4C), 69.1 (C-2C), 68.7 (C-2B), 68.1
(C-2A), 66.8 (C-4A), 54.7 (OCH3), 20.8, 20.7, 20.6 (3 COCH3), 18.1,
18.0, 17.5 (3 CCH3).
MS (EI, 70 eV): 929.4 [M + Na]+.
MS (EI, 70 eV): 1219.4 [M + Na]+.
Anal. Calcd for C49H62O16: C, 64.89; H, 6.89. Found: C, 64.72; H,
7.08.
Anal. Calcd for C60H76O25: C, 60.19; H, 6.40. Found: C, 60.0; H,
6.54.
Methyl (2,3,4-Tri-O-acetyl-a-L-rhamnopyranosyl)-(1→3)-(4-O-
benzyl-a-L-rhamnopyranosyl)-(1→3)-2,4-di-O-benzyl-a-L-
rhamnopyranoside (25)
Methyl (a-L-Rhamnopyranosyl)-(1→3)-[(b-D-glucopyranosyl)-
(1→2)]-(a-L-rhamnopyranosyl)-(1→3)-a-L-rhamnopyranoside
(2)
PdCl2 (180 mg, 1.0 mmol) was added to a soln of allyl derivative 24
(1.8 g, 1.98 mmol) in anhyd MeOH (25 mL), and the mixture was
stirred at r.t. for 3 h. The mixture was filtered through a Celite bed
and evaporated to dryness. The crude product was purified by chro-
matography [silica gel, hexane–EtOAc (4:1)] to give pure com-
pound 25 as a yellow oil; yield: 1.5 g (87%); [a]D –49 (c 1.2,
CHCl3).
1H NMR (300 MHz, CDCl3): d = 7.35–7.19 (m, 15 H, Ar-H), 5.31–
5.28 (m, 2 H, H-2C, H-4C), 5.04 (br s, 1 H, H-1B), 5.01–4.98 (m, 2
H, H-1C, H-3C), 4.80–4.59 (m, 7 H, H-1A, 3 PhCH2), 4.03–3.95 (m,
4 H, H-2B, H-3A, H-5A, H-5C), 3.79–3.76 (m, 1 H, H-5B), 3.69–3.66
20% Pd(OH)2/C (250 mg) was added to a soln of protected deriva-
tive 26 (1.5 g, 1.25 mmol) in MeOH (20 mL), and the mixture was
stirred under a positive pressure of H2 at r.t. for 10 h. The mixture
was filtered through a Celite bed, and the solvents were evaporated
to dryness. A soln of the crude product in 0.1 M methanolic NaOMe
(20 mL) was stirred at r.t. for 6 h and neutralized with Dowex 50W
X-8 (H+). The mixture was filtered and evaporated to dryness to
give the target compound 2, which was purified as an amorphous
powder by chromatography [LH-20 Sephadex, MeOH-H2O (4:1)];
yield: 625 mg (79%); [a]D –20 (c 1.0, H2O).
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York

2590
S. Pandey et al.
PAPER
1H NMR (300 MHz, CDCl3): d = 5.04 (br s, 1 H, H-1A), 5.01 (br s,
1 H, H-1C), 4.76 (d, J = 7.8 Hz, 1 H, H-1D), 4.57 (br s, 1 H, H-1B),
4.08–4.05 (m, 1 H, H-2C), 4.0 (br s, 1 H, H-2A), 3.89–3.65 (m, 9 H,
H-2B, H-3A, H-3B, H-3C, H-3D, H-4B, H-5A, H-5C, H-6aD), 3.63–3.49
(4 H, H-4A, H-4C, H-5B, H-6bD), 3.44–3.28 (6 H, H-2D, H-4D, H-5D,
OCH3), 18.1, 18.0, 17.7 (m, 9 H, 3 CCH3).
13C NMR (75 MHz, CDCl3): d = 104.2 (C-1D), 104.0 (C-1A), 103.9
(C-1C), 102.6 (C-1B), 79.9 (C-5A), 79.4 (C-5C), 78.5 (C-5D), 76.8 (C-
2D), 76.5 (C-3D), 74.1 (C-4B), 73.2 (C-2B), 73.0 (C-3B), 72.2 (C-2C),
72.1 (C-3C), 72.4 (C-4D), 71.9 (C-2A), 71.8 (C-3A), 70.3 (C-4A), 70.2
(C-4C), 70.1 (C-5B), 62.6 (C-6D), 55.4 (OCH3), 18.3, 18.2, 18.0 (3
CCH3).
(2) Bashan, Y.; Holgium, G. Can. J. Microbiol. 1997, 43, 103.
(3) Tien, T. M.; Gaskins, M. H.; Hubbell, D. H. Appl. Environ.
Microbiol. 1979, 37, 1016.
(4) (a) Jofré, E.; Lagares, A.; Mori, G. FEMS Microbiol. Lett.
2004, 231, 267. (b) Burdman, S.; Okon, Y.; Jurkevitch, E.
Crit. Rev. Microbiol. 2000, 26, 91. (c) De Troch, P.;
Vanderleyden, J. Microbiol. Ecol. 1996, 32, 149.
(d) Konnova, S. A.; Makarov, O. E.; Skvortsov, I. M.;
Ignatov, V. V. FEMS Microbiol. Lett. 1994, 118, 93.
(5) Fedonenko, Y. P.; Konnova, O. N.; Zdorovenko, E. L.;
Konnova, S. A.; Zatonsky, G. V.; Shashkov, A. S.; Ignatov,
V. V.; Knirel, Y. A. Carbohydr. Res. 2008, 343, 810.
(6) Hall, D. M. Carbohydr. Res. 1980, 86, 158.
(7) Boken, H. B.; Ekborg, G.; Lonngren, J. Acta Chem. Scand.,
Ser. B. 1975, 29, 1085.
MS (EI, 70 eV): 655.2 [M + Na]+.
Anal. Calcd for C25H44O18: C, 47.47; H, 7.01. Found: C, 47.30; H,
7.20.
(8) Zuurmond, H. M.; van der Klein, P. A. M.; van der Marel,
G. A.; van Boom, J. H. Tetrahedron 1993, 49, 6501.
(9) Sajtos, F.; Hajkó, J.; Kövér, K. E.; Lipták, A. Carbohydr.
Res. 2001, 334, 253.
Acknowledgment
(10) Sarkar, K.; Mukherjee, I.; Roy, N. J. Carbohydr. Chem.
2003, 22, 95.
(11) Garegg, P. J.; Hultberg, H.; Wallin, S. Carbohydr. Res.
1982, 108, 97.
(12) Veeneman, G. H.; Van Leeuwen, S. H.; Zuurmond, H.;
van Boom, J. H. J. Carbohydr. Chem. 1990, 9, 783.
(13) Veeneman, G. H.; Gomes, L. J. F.; van Boom, J. H.
Tetrahedron 1989, 45, 7433.
S.G. thanks UGC, New Delhi, for providing a Junior Research Fel-
lowship. This work was supported by a Ramanna Fellowship
(AKM), Department of Science and Technology, New Delhi
(SR/S1/RFPC-06/2006).
References
(14) Das, S. K.; Roy, N. Carbohydr. Res. 1996, 296, 275.
(15) (a) Veeneman, G. H.; van Leeuwen, S. H.; van Boom, J. H.
Tetrahedron Lett. 1990, 31, 1331. (b) Konradsson, P.;
Udodong, U. E.; Fraser-Reid, B. Tetrahedron Lett. 1990, 31,
4313.
(16) Mandal, P. K.; McMurray, J. S. J. Org. Chem. 2007, 72,
6599.
(17) Pearlman, W. M. Tetrahedron Lett. 1967, 8, 1663.
(1) (a) Okon, Y.; Vanderleyden, J. ASM News 1997, 63, 366.
(b) Fedonenko, Y. P.; Konnova, O. N.; Zatonsky, G. V.;
Konnova, S. A.; Kocharova, N. A.; Zdorovenko, E. L.;
Ignatov, V. V. Carbohydr. Res. 2005, 340, 1259.
(c) Fedonenko, Y. P.; Konnova, O. N.; Zatonsky, G. V.;
Shashkov, A. S.; Konnova, S. A.; Zdorovenko, E. L.;
Ignatov, V. V.; Knirel, Y. A. Carbohydr. Res. 2004, 339,
1813. (d) Fedonenko, Y. P.; Zatonsky, G. V.; Konnova, S.
A.; Zdorovenko, E. L.; Ignatov, V. V. Carbohydr. Res. 2002,
337, 869.
Synthesis 2009, No. 15, 2584–2590 © Thieme Stuttgart · New York