Facile synthesis of the heptasaccharide repeating unit of O-deacetylated GXM of C. neoformans serotype B

Article abstract of DOI:10.1016/j.bmc.2004.09.049

A heptasaccharide, β-d-Xylp-(1→2)-α-d-Manp-(1→3)- [β-d-Xylp-(1→2)]-α-d-Manp-(1→3)-[β-d-GlcpA-(1→2)] [β-d-Xylp-(1→4)]-α-d-Manp, the repeating unit of the exopolysaccharide from Cryptococcus neoformans serovar B, was synthesized as its methyl glycoside. Thu

Full text of DOI:10.1016/j.bmc.2004.09.049

Bioorganic & Medicinal Chemistry 13 (2005) 121–130
Facile synthesis of the heptasaccharide repeating unit of
O-deacetylated GXM of C. neoformans serotype B
Wei Zhao and Fanzuo Kong*
Research Center for Eco-Environmental Sciences, Academia Sinica, PO Box 2871, Beijing 100085, PR China
Received 9 September 2004; revised 28 September 2004; accepted 28 September 2004
Abstract—A heptasaccharide, b-D-Xylp-(1!2)-a-D-Manp-(1!3)-[b-D-Xylp-(1!2)]-a-D-Manp-(1!3)-[b-D-GlcpA-(1!2)][b-D-
Xylp-(1!4)]-a-D-Manp, the repeating unit of the exopolysaccharide from Cryptococcus neoformans serovar B, was synthesized as
its methyl glycoside. Thus 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-3,4,6-tri-O-benzoyl-a-D-mannopyranosyl trichloroacetimi-
date (7) and allyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-4,6-di-O-benzoyl-a-D-mannopyranoside (8), readily obtained from
the corresponding monosaccharide derivatives via simple transformation, were coupled to give a (1!3)-linked tetrasaccharide 9.
Deallylation of 9 followed by trichloroacetimidate formation produced the tetrasaccharide donor 11. Condensation of methyl
2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!4)-2-O-acetyl-6-O-benzoyl-a-D-mannopyranoside (18) with 11 followed by selective
deacetylation yielded hexasaccharide acceptor 20. Coupling of 20 with methyl 2,3,4-tri-O-acetyl-a-D-glucopyranosyluronate bro-
mide (21) and subsequent deprotection furnished the target heptaoside. A hexasaccharide fragment, a-D-Manp-(1!3)-[b-D-Xylp-
(1!2)]-a-D-Manp-(1!3)-[b-D-GlcpA-(1!2)][b-D-Xylp-(1!4)]-a-D-Manp, was also similarly synthesized as its methyl glycoside.
ꢀ 2004 Elsevier Ltd. All rights reserved.
1. Introduction
side,4 and a hexasaccharide fragment5 of the GXM of
C. neoformans serotype B. Earlier, the synthesis of tri-
saccharide and tetrasaccharide fragments6 correspond-
ing to structures in capsular polysaccharides of C.
neoformans and the synthesis of a pentasaccharide7—
the repeating unit of the polysaccharide in C. neofor-
mans serovar D were reported. We now report a conver-
gent synthesis of the heptasaccharide repeating unit of
O-deacetylated GXM of C. neoformans serotype B,
and a hexasaccharide fragment of the GXM of serotypes
B and C.
C. neoformans, the etiological agent of cryptococcosis, is
an encapsulated yeast. The major component of the cap-
sular envelop of C. neoformans is composed of an acidic
hetero-polysaccharide, glucuronoxylomannan (GXM),
consisting of mannose, xylose, glucuronic acid, and O-
acetyl groups. Initially, 12 strains of C. neoformans were
classified into three antigenic types (A, B, and C) on the
basis of agglutinin titers.1 A fourth, type D, was intro-
duced later.2 Of the four major serotypes A–D for
GXM, D has the simplest pentaose structure while C
has the most complex octaose structure. All the four sero-
types are composed of a linear a-1,3-linked mannosyl
backbone with b-glucopyranosyluronic acid, b-xylopy-
ranosyl, and 6-O-acetyl substituents (Fig. 1).3
2. Results and discussion
In our previous work,5 a trial for the synthesis of methyl
glycoside of the frame-shifted heptaoside, b-D-GlcpA-
(1!2)-[b-D-Xylp-(1!4)]-a-D-Manp-(1!3)-[b-D-Xylp-
(1!2)]-a-D-Manp-(1!3)-[b-D-Xylp-(1!2)]-a-D-Manp,
was not successful, since the coupling of the hexasaccha-
ride acceptor, methyl 2,3,4-tri-O-benzoyl-b-D-xylopy-
ranosyl-(1!4)-3,6-di-O-benzoyl-a-D-mannopyranosyl-
(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-
4,6-di-O-benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-tri-
O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-benzoyl-
a-D-mannopyranoside, with glucuronate donors either
We have reported the successful syntheses of the hexa-
saccharide repeating unit of O-deacetylated GXM of
C. neoformans serotype A and its frame-shifted hexao-
Keywords: Mannose; Xylose; Glucuronic acid; Regio- and stereo-
selective synthesis.
*
Corresponding author. Tel.: +86 1062936613; fax: +86
1062923563; e-mail: fzkong@mail.rcees.ac.cn
0968-0896/$ - see front matter ꢀ 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.09.049

122
W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
β
β
β
-D-Xylp1
β-D-Xylp1
↓2
-D-Xylp1
β-D-GlcpA1
↓2
-D-Xylp1
β-D-GlcpA1
↓
2
2
2
2
↓
↓
↓
→
→
→
→
→
→
→3)-α-D-Manp(1 3)-α-D-Manp(1 3)-α-D-Manp(1
3)-α-D-Manp(1 3)-α-D-Manp(1 3)-α-D-Manp(1
→
↑
4
A
B
β-D-Xylp1
β
β-D-Xylp1
β-D-Xylp1
↓2
β-D-GlcpA1
-D-Xylp1
β-D-GlcpA1
↓2
↓2
↓2
2
↓
→
→
→
→
α
3)-α-D-Manp(1→3)- -D-Manp(1 3)-α-D-Manp(1
→3)-α-D-Manp(1→3)- -D-Manp(1→3)-α-D-Manp(1
α
C
↑4
↑4
D
β-D-Xylp1
β-D-Xylp1
Figure 1. Structures of deacetylated GXM of C. neoformans serotypes A–D.
gave transacetylated hexasaccharide product when the
acetylated glucuronate trichloroacetimidate was used,
or did not occur when the acetylated glucuronate bro-
mide was used. We suppose that, a serious steric hin-
drance at C-2 of the upstream mannose residue of the
hexasaccharide acceptor restricted the coupling. Thus,
we expected that a hexasaccharide acceptor with C-2
free hydroxyl group at the downstream mannose residue
would be suitable for its coupling with the glucuronate
donor due to smaller steric hindrance. The experimental
results verified the expectation, and a successful synthe-
sis of the title heptaoside was achieved.
firming the 3-O-allylation of 12, and the 1H NMR spec-
trum of 16 gave H-4 in the range of d 4.18–3.87ppm
indicating the selective 6-O-benzoylation of 15. Conden-
sation (90%) of 16 with 2 followed by deallylation (85%)
produced the downstream disaccharide acceptor 18.
Coupling of 18 with 11 went smoothly giving the hexasac-
charide 19 (70%) with the 13C NMR spectrum showing
three signals at d 102.6, 99.7, and 98.6ppm with JC1-H1
163.4–164.1Hz for Xylp-b-C-1, and 98.5, 97.7, and
97.0ppm with JC1-H1 173.4–176.1Hz for Manp-a-C-1.
Selective removal of 2-O-acetyl group of 19 by methano-
lysis9 with MeCOCl/MeOH/CH2Cl2 (3.5/40/10mL)
afforded the hexasaccharide acceptor 20 in acceptable
yield (45%). Condensation of 20 with methyl 2,3,4-tri-
O-acetyl-a-D-glucopyranosyluronate bromide (21) in
the presence of AgOTf furnished the heptaoside 22 in
good yield (78%), and the 13C NMR spectrum of 22 gave
GlcpA-b-C-1 at d 95.7ppm with JC1-H1 166.8Hz. Depro-
tection of 22 in ammonia-saturated methanol gave the
target free heptaoside 23.
Scheme 1 outlined the synthesis of the target heptaoside.
Allyl
3-O-benzoyl-4,6-O-benzylidene-a-D-mannopyr-
anoside5 (1) was chosen as the starting material since it
was obtained in nearly quantitative yield from 3-O-selec-
tive benzoylation of 4,6-O-benzylidene-a-D-mannopyr-
anoside.5 Glycosylation (85%) of allyl 3-O-benzoyl-4,6-
O-benzylidene-a-D-mannopyranoside5 (1) with 2,3,4-tri-
O-benzoyl-D-xylopyranosyl trichloroacetimidate (2)
followed by debenzylidenation (90%) and benzoylation
(80%) gave the disaccharide 5. Subsequent deallylation
(85%) with PdCl2 and trichloroacetimidate formation
(89%) with trichloroacetonitrile8 afforded the upstream
disaccharide donor 7. Condensation (62%) of allyl
2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-4,6-di-O-
benzoyl-a-D-mannopyranoside5 (8) with the disaccharide
donor 7 yielded the tetrasaccharide 9 that showed two
signals in its 13C NMR spectrum at d 100.0 and
99.3ppm with JC1-H1 163.4Hz for Xylp-b-C-1, and two
signals at d 99.1ppm (JC1-H1 176Hz) and 96.7ppm (JC1-H1
172Hz) for Manp-a-C-1. Deallylation (89%) of 9 and
subsequent trichloroacetimidate formation (89%) pro-
duced the upstream tetrasaccharide donor 11. The down-
stream disaccharide was readily prepared by the
following way. Selective 3-O-allylation (85%) of methyl
4,6-O-benzylidene-a-D-mannopyranoside (12) through
dibutyltin complex, followed by acetylation (90%),
debenzylidenation (90%), and selective 6-O-benzoylation
(90%) furnished 16. The 1H NMR spectrum of 14 showed
H-2 at d 5.31ppm with J1,2 = 1.1Hz, J2,3 = 3.5Hz, and
H-3 at d 3.91ppm with J2,3 = 3.5Hz, J3,4 = 9.9Hz con-
A hexasaccharide fragment of the GXM of serotypes B
and C was also synthesized similarly as shown in Scheme
2. Thus, coupling (95%) of the disaccharide acceptor 8
with 2,3,4,6-tetra-O-benzoyl-a-D-mannopyranosyl tri-
chloroacetimidate (24) followed by deallylation (86%)
and trichloroacetimidate formation (89%) afforded the
trisaccharide donor 27. Condensation (80%) of the disac-
charide acceptor 18 with 27, followed by selective
deacetylation (42%) gave the pentasaccharide acceptor
29. Coupling (76%) of 29 with 21 followed by deacylation
produced the hexasaccharide fragment 31.
3. Conclusion
In summary, a convergent synthesis of complex glucur-
ono-xylo-mannosyl oligosaccharide was achieved
through a regio- and stereoselective manner with readily
accessible materials. The described method is suitable
for preparation of other oligosaccharide fragments of
the deacetylated GXM of serotypes A, B, and D.

W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
123
BzO
BzO
BzO
BzO
BzO
O
OBz
O
O
OR
BzO
O
O
BzO
O
Ph
BzO
BzO
O
HO
BzO
O
BzO
R = C(NH)CCl3
Ph
b
O
BzO
d
O
O
2
BzO
BzO
RO
RO
O
O
O
O
a
OAll
BzO
BzO
OR
BzO
OAll
OAll
3
1
4
R = H
6 R = H
c
e
5 R=Bz
7 R = C(NH)CCl3
BzO
BzO
BzO
BzO
O
BzO
O
BzO
BzO
BzO
O
O
O
9 R = All
10 R = H
d
e
BzO
BzO
BzO
O
O
a
BzO
BzO
BzO
O
O
11 R = C(NH)CCl3
BzO
HO
O
BzO
OAll
RO
OR
O
8
OBz
OR
Ph
BzO
BzO
BzO
OAc
O
O
O
BzO
OAc
BzO
RO
HO
AllO
O
b
O
2
R = C(NH)CCl3
O
O
RO
BzO
R1O
a
OMe
OMe
OMe
12
13
R = H R1=H
R = H R1=All
15 R = H
R = All
R = H
17
18
f
c
d
R = Bz
16
g
14
R=Ac
R1=All
OBz
BzO
O
O
COOMe
O
AcO
AcO
BzO
BzO
OBz
BzO
BzO
O
BzO
O
BzO
AcO
Br
O
O
O
OR
21
BzO
BzO
O
BzO
O
O
a
BzO
i
11
+
18
BzO
OBz
h
MeO
19 R= Ac
R = H
20
COOMe
AcO
AcO
COOH
HO
HO
OBz
OH
BzO
O
O
HO
O
O
O
O
O
AcO
BzO
OBz
O
HO
HO
j
OH
BzO
BzO
O
O
O
HO
HO
BzO
OBz
O
HO
HO
OH
O
O
O
BzO
O
O
BzO
BzO
O
O
O
HO
HO
O
O
BzO
BzO
O
O
HO
HO
HO
BzO
OBz
MeO
22
MeO
OH
23
Scheme 1. Reagents and conditions: (a) TMSOTf (0.01–0.05equiv), CH2Cl2, ꢀ20 to 0ꢁC, 2–4h, 85% for 3, 62% for 9, 90% for 17, 70% for 19,
respectively; (b) 90% HOAc–H2O, 70ꢁC, 2h, 90% for 4, 90% for 15; (c) BzCl–pyridine, 80% for 5; 90% for 16; (d) PdCl2, CH3OH, rt, 4h, 85% for 6,
89% for 10, 85% for 18; (e) CCl3CN, DBU, CH2Cl2, 3h, 89% for 7, 89% for 11; (f) (i) MeOH, Bu2SnO, reflux, (ii) AllBr, toluene; (g) Ac2O–pyridine,
90% for 14; (h) MeCOCl/MeOH/CH2Cl2 (3.5/40/10mL), rt, 48h, 45% for 20; (i) 2,4-lutidine, silver triflate, CH2Cl2, ꢀ20 to 0ꢁC, 2–4h, 78% for 22;
(j) satd NH3–MeOH, rt, 72h, 65% for 23.

124
W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
BzO
BzO
O
BzO
BzO
O
O
BzO
BzO
BzO
BzO
BzO
BzO
O
BzO
BzO
BzO
O
a
BzO
BzO
O
O
BzO
BzO
BzO
+
O
OC(NH)CCl3
O
HO
OAll
OR
24
8
R = All
R = H
25
26
b
c
27
R = C(NH)CCl3
OBz
BzO
O
O
COOMe
O
BzO
BzO
OBz
AcO
AcO
O
BzO
O
BzO
BzO
O
BzO
AcO
21
Br
O
OR
BzO
a
O
18
27
+
O
e
BzO
BzO
OBz
MeO
28
Ac
R=
d
29 R = H
COOMe
AcO
AcO
HO COOH
HO
OBz
OH
BzO
O
O
HO
O
O
O
O
O
BzO
BzO
AcO
OBz
BzO
f
O
HO
HO
OH
BzO
O
O
HO
O
HO
O
O
BzO
HO
HO
OH
BzO
O
BzO
HO
O
O
O
O
O
O
BzO
BzO
HO
HO
OBz
MeO
MeO
30
OH
31
Scheme 2. Reagents and conditions: (a) TMSOTf (0.01–0.05equiv), CH2Cl2, ꢀ20 to 0ꢁC, 2–4h, 95% for 25, 80% for 28, respectively; (b) PdCl2,
CH3OH, rt, 4h, 86% for 26; (c) CCl3CN, DBU, CH2Cl2, 3h, 89% for 27; (d) MeCOCl/MeOH/CH2Cl2 (3.5/40/10mL), rt, 48h, 42% for 29; (e) 2,4-
lutidine, silver triflate, CH2Cl2, ꢀ20 to 0ꢁC, 2–4h, 76% for 30; (f) satd NH3–MeOH, rt, 72h, 64% for 31.
4. Experimental
4.1. General methods
of a column (16 · 240mm, 18 · 300mm, 35 · 400mm)
of silica gel (100–200 mesh) with EtOAc–petroleum
ether (60–90ꢁC) as the eluent. Solutions were concen-
trated at <60ꢁC under reduced pressure.
Optical rotations were determined at 25ꢁC with a Per-
kin–Elmer Model 241-Mc automatic polarimeter. 1H
NMR and 13C NMR spectra were recorded with Bruker
4.2. General procedure for the glycosylations
1
ARX 400 spectrometers (400MHz for H, 100MHz for
13C) for solutions in CDCl3 or D2O as indicated. Chem-
ical shifts are given in ppm downfield from internal
Me4Si. Mass spectra were measured using MALTI-
TOF-MS with CCA as matrix or recorded with a VG
PLATFORM mass spectrometer using the ESI mode.
Thin layer chromatography (TLC) was performed on
silica gel HF254 with detection by charring with 30%
(v/v) H2SO4 in MeOH or in some cases by a UV detec-
tor. Column chromatography was conducted by elution
The mixture of donor and acceptor was dried together
under high vacuum for 2h, then dissolved in anhyd
CH2Cl2. TMSOTf (0.05equiv) was added dropwise at
ꢀ20ꢁC with nitrogen protection. The reaction mixture
was stirred for 3h, during which time the temperature
was gradually raised to ambient temperature. Then the
mixture was neutralized with Et3N. Concentration of
the reaction mixture, followed by purification on a silica
gel column, gave the desired products.

W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
125
4.3. Allyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-
3-O-benzoyl-4,6-O-benzylidene-a-D-mannopyranoside (3)
5.33–5.20 (m, 2H, CH2@CHCH2O), 5.12 (m, 1H, H-4
of Xylp), 4.99 (d, 1H, J1,2 = 1.6Hz, H-1 of Manp),
4.98 (d, 1H, J1,2 = 3.6Hz, H-1 of Xylp), 4.50 (dd, 1H,
J1,2 = 1.6Hz, J2,3 = 3.3Hz, H-2 of Manp), 4.40–4.33
(m, 2H, H-5 of Xylp), 4.30 (ddd, 1H, J4,5 = 10.1Hz,
J5,6a = 6.3Hz, J5,6b = 4.6Hz, H-5 of Manp), 4.20 (m,
1H, CH2@CHCH2O), 4.06 (dd, 1H, J5,6a = 6.3Hz,
J6a,6b = 12.0Hz, H-6a of Manp), 4.00 (m, 1H,
As described in the general procedure, 1 (5.14g,
12.50mmol) and 2 (8.00g, 13.16mmol) were coupled,
and the product was purified by column chromatogra-
phy with 2.5:1 petroleum ether–EtOAc as the eluent to
give 3 (9.02g, 85%) as a foamy solid. [a]D ꢀ38.9 (c 1.0,
1
CHCl3); H NMR (400MHz, CDCl3): d 8.13–7.25 (m,
CH2@CHCH2),
3.33
(dd,
1H,
J5,6b = 4.6Hz,
25H, 5Ph), 5.87–5.77 (m, 1H, –CH2–CH@CH2), 5.66
(dd, 1H, J3,4 = J2,3 = 4.9Hz, H-3 of Xylp), 5.57 (dd,
1H, J2,3 = 3.3, J3,4 = 10.1Hz, H-3 of Manp), 5.38 (dd,
1H, J1,2 = 4.0Hz, J2,3 = 4.9Hz, H-2 of Xylp), 5.31 (s,
1H, PhCH), 5.27–5.17 (m, 2H, –CH2–CH@CH2), 5.17
(m, 1H, H-4 of Xylp), 4.89 (d, 1H, J1,2 = 4.0Hz, H-1
of Xylp), 4.84 (d, 1H, J1,2 = 1.3Hz, H-1 of Manp),
4.47 (dd, 1H, J1,2 = 1.3Hz, J2,3 = 3.3Hz, H-2 of Manp),
4.30–3.43 (m, 8H, H-5 of Xylp, H-4, 5, 6 of Manp,
–CH2–CH@CH2). Anal. Calcd for C49H44O14:
C, 68.69; H, 5.14. Found: C, 68.77; H, 5.10.
J6a,6b = 12.0Hz, H-6b of Manp). Anal. Calcd
for C56H48O16: C, 68.85; H, 4.92. Found: C, 68.78; H,
5.00.
4.6. 2,3,4-Tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-3,4,6-
tri-O-benzoyl-a-D-mannopyranose (6)
To a solution of 5 (4.29g, 5.31mmol) in anhyd MeOH
(25mL) was added PdCl2 (20mg). After stirring the mix-
ture at rt for 2h, TLC (2:1 petroleum ether–EtOAc)
indicated that the reaction was complete. The mixture
was filtered, the solution was concentrated to dryness,
and the resultant residue was purified by flash chroma-
tography (2:1 petroleum ether–EtOAc) to give 6
(4.22g, 85%) as an amorphous solid. [a]D ꢀ23.7 (c 1.3,
4.4. Allyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-
3-O-benzoyl-a-D-mannopyranoside (4)
1
A mixture of 3 (8.69g, 10.15mmol) and 90% HOAc–
H2O (100mL) was stirred for 2h at 70ꢁC, then concen-
trated to dryness. Purification of the residue by column
chromatography (1:1 petroleum ether–EtOAc) gave 4
(7.02g, 90%) as an amorphous solid. [a]D ꢀ78.6 (c 1.0,
CHCl3); H NMR (400MHz, CDCl3): d 8.19–7.30 (m,
30H, 6Ph), 5.94 (dd, 1H, J3,4 = J4,5 = 10.0Hz, H-4 of
Manp), 5.70 (dd, 1H, J2,3 = 3.3Hz, J3,4 = 10.0Hz, H-3
of Manp), 5.63 (dd, 1H, J2,3 = J3,4 = 4.9Hz, H-3 of
Xylp), 5.40 (d, 1H, J1,2 = 1.4Hz, H-1 of Manp), 5.32
(dd, 1H, J1,2 = 3.8Hz, J2,3 = 4.9Hz, H-2 of Xylp), 5.14
(m, 1H, H-4 of Xylp), 4.94 (d, 1H, J1,2 = 3.8Hz, H-1
of Xylp), 4.50 (dd, 1H, J1,2 = 1.4Hz, J2,3 = 3.3Hz, H-2
of Manp), 4.40–3.33 (m, 5H, H-5 of Xylp, H-5, 6 of
Manp). Anal. Calcd for C53H44O16: C, 67.95; H, 4.70.
Found: C, 68.04; H, 4.72.
1
CHCl3); H NMR (400MHz, CDCl3): d 8.13–7.35 (m,
20H, 4Ph), 5.83–5.76 (m, 1H, –CH2–CH@CH2), 5.76
(dd, 1H, J2,3 = J3,4 = 5.0Hz, H-3 of Xylp), 5.46 (dd,
1H, J1,2 = 4.1Hz, J2,3 = 5.0Hz, H-2 of Xylp), 5.42 (dd,
1H, J2,3 = 3.3Hz, J3,4 = 10.1Hz, H-3 of Manp), 5.28
(m, 1H, H-4 of Xylp), 5.27–5.10 (m, 2H, –CH2–
CH@CH2), 4.75 (d, 1H, J1,2 = 4.1Hz, H-1 of Xylp),
4.69 (d, 1H, J1,2 = 1.2Hz, H-1 of Manp), 4.27 (dd, 1H,
J1,2 = 1.2Hz, J2,3 = 3.3Hz, H-2 of Manp), 4.30–3.33
(m, 8H, H-5 of Xylp, H-4, 5, 6 of Manp, –CH2–
CH@CH2). Anal. Calcd for C42H40O14: C, 65.63; H,
5.21. Found: C, 65.58; H, 5.23.
4.7. 2,3,4-Tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-3,4,6-
tri-O-benzoyl-a-D-mannopyranosoyl trichloroacetimidate
(7)
A mixture of 6 (4.02g, 4.30mmol), trichloroacetonitrile
(1.0mL, 10.0mmol) and 1,8-diazabicyclo[5.4.0]-unde-
cene (DBU) (0.5mL) in dry CH2Cl2 (50mL) was stirred
under nitrogen for 3h and then concentrated. The resi-
due was purified by flash chromatography (3:1 petro-
leum ether–EtOAc) to give 7 (3.71g, 89%) as an
4.5. Allyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-
3,4,6-tri-O-benzoyl-a-D-mannopyranoside (5)
Compound 4 (6.25g, 8.14mmol) was dissolved in pyr-
idine (30mL), and benzoyl chloride (3.5mL, 30mmol)
was added dropwise within 30min. The mixture was stir-
red overnight at rt, and TLC (2:1 petroleum ether–
EtOAc) indicated that the reaction was complete. Ice
water was added, and the mixture was diluted with
dichloromethane, washed with 1M HCl, water, and satd
aq sodium bicarbonate subsequently. The organic layer
was combined, dried, and concentrated. Purification by
column chromatography (2:1 petroleum ether–EtOAc)
gave 5 (5.26g, 80%) as an amorphous solid. [a]D ꢀ23.7
1
amorphous solid. [a]D ꢀ17.5 (c 1.0, CHCl3); H NMR
(400MHz, CDCl3): d 8.73 (s, 1H, CNHCCL3), 8.18–
7.12 (m, 30H, 6Ph), 6.44 (d, 1H, H-1 of Manp), 6.03
(dd, 1H, J3,4 = J4,5 = 10.1Hz, H-4 of Manp), 5.68 (dd,
1H, J2,3 = 3.2Hz, J3,4 = 10.1Hz, H-3 of Manp), 5.66
(dd, 1H, J2,3 = J3,4 = 5.1Hz, H-3 of Xylp), 5.40 (dd,
1H, J1,2 = 3.8Hz, J2,3 = 5.1Hz, H-2 of Xylp), 5.13 (m,
1H, H-4 of Xylp), 5.09 (d, 1H, J1,2 = 3.8Hz, H-1 of
Xylp), 4.75 (dd, 1H, J1,2 = 0.6Hz, J2,3 = 3.2Hz, H-2 of
Manp), 4.50 (ddd, 1H, J4,5 = 10.1Hz, J5,6a = 6.3Hz,
J5,6b = 4.6Hz, H-5 of Manp), 4.44 (dd, 1H,
J4,5a = 2.8Hz, J5a,5b = 12.1Hz, H-5a of Xylp), 4.34 (dd,
1H, J4,5b = 3.2Hz, J5a,5b = 12.1Hz, H-5b of Xylp), 4.16
(dd, 1H, J5,6a = 5.8Hz, J6a,6b = 12.1Hz, H-6a of Manp),
3.44 (dd, 1H, J5,6b = 5.0Hz, J6a,6b = 12.1Hz, H-6b of
1
(c 1.3, CHCl3); H NMR (400MHz, CDCl3): d 8.19–
7.30 (m, 30H, 6Ph), 5.94 (m, 1H, CH2@CHCH2O),
5.92 (dd, 1H, J3,4 = J4,5 = 10.1Hz, H-4 of Manp), 5.66
(dd, 1H, J2,3 = 3.3Hz, J3,4 = 10.1Hz, H-3 of Manp),
5.63 (dd, 1H, J2,3 = J3,4 = 4.9Hz, H-3 of Xylp), 5.34
(dd, 1H, J1,2 = 3.6Hz, J2,3 = 4.9Hz, H-2 of Xylp),

126
W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
Manp). Anal. Calcd for C55H44Cl3NO16: C, 61.09; H,
4.10. Found: C, 61.19; H, 4.18.
4.10. 2,3,4-Tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-
3,4,6-tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-
tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-ben-
zoyl-a-D-mannopyranosyl trichloroacetimidate (11)
4.8. Allyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-
3,4,6-tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-
tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-ben-
zoyl-a-D-mannopyranoside (9)
A mixture of 10 (1.5g, 0.86mmol), trichloroacetonitrile
(200lL, 2mmol) and 1,8-diazabicyclo[5.4.0]-undecene
(DBU) (200lL) in dry CH2Cl2 (30mL) was stirred
under nitrogen for 3h and then concentrated. The residue
was purified by flash chromatography (2:1 petroleum
ether–EtOAc) to give 11 (1.47g, 89%) as an amorphous
As described in the general procedure, 7 (2.33g,
2.44mmol) and 8 (2.27g, 2.60mmol) were coupled,
and the product was purified by column chromatogra-
phy with 2:1 petroleum ether–EtOAc as the eluent to
give 9 (2.72g, 62%) as an amorphous solid. [a]D ꢀ92.9
1
solid. [a]D ꢀ71.2 (c 1.3, CHCl3); H NMR (400MHz,
CDCl3): d 8.70 (s, 1H, CNHCCL3), 8.13–7.31 (m,
55H, 11Ph), 6.40 (s, 1H, H-1, Manp), 5.82 (dd, 1H,
1
J3,4 = J4,5 = 9.7Hz, H-4, Manp), 5.78 (dd, 1H, J2,3
3.2Hz, J3,4 = 9.8Hz, H-3, Manp), 5.74 (dd, 1H,
J1,2 = J2,3 = 4.1Hz, H-2, Xylp), 5.72 (dd, 1H, J3,4
=
(c 1.2, CHCl3); H NMR (400MHz, CDCl3): d 8.16–
7.31 (m, 55H, 11Ph), 5.85 (m, 1H, CH2@CHCH2O),
5.84 (dd, 1H, J3,4 = J4,5 = 9.9Hz, H-4 of Manp), 5.76–
5.62 (m, 3H, H-4 of Manp, H-3 of Xylp), 5.49 (m, 1H,
H-4 of Xylp), 5.45 (dd, 1H, J1,2 = J2,3 = 5.7Hz, H-2 of
Xylp), 5.35–5.07 (m, 3H, H-3 of Manp, H-2 of Xylp,
H-4 of Xylp), 5.10 (d, 1H, J1,2 = 0.7Hz, H-1 of Manp),
5.07 (d, 1H, J1,2 = 4.2Hz, H-1 of Xylp), 4.87 (d, 1H,
J1,2 = 0.9Hz, H-1 of Manp), 4.43 (d, 1H, J1,2 = 5.1Hz,
H-1 of Xylp), 4.35–3.88 (m, 15H, H-5 of Xylp, H-2, 3,
5 of Manp, CH2@CHCH2O), 2.92–2.81 (m, 2H, H-6a,
H-6b of Manp); 13C NMR (100MHz, CDCl3) d 166.2,
166.1, 165.6, 165.6, 165.5, 165.4, 165.3, 165.3, 165.1,
164.8, 164.7 (11C, 11PhCO), 118.3 (CH2@CHCH2O),
100.0 (JC1-H1 163.4Hz), 99.3 (JC1-H1 163.4Hz), 99.1
(JC1-H1 176Hz), 96.7 (JC1-H1 172Hz) (4C, 4C-1), 78.2,
76.0, 75.9, 70.3, 69.8, 69.8, 69.7, 69.3, 68.8, 68.7, 68.7,
68.6, 68.1, 67.6, 64.0, 63.7, 60.6, 60.5 (C-2 to C-6). Anal.
Calcd for C102H86O30: C, 68.37; H, 4.84. Found: C,
68.52; H, 4.89.
=
J4,5 = 9.8Hz, H-4, Manp), 5.51 (m, 1H, H-4, Xylp),
5.45 (dd, 1H, J1,2 = J2,3 = 4.0Hz, H-2, Xylp), 5.36 (dd,
1H, J2,3 = 4.0Hz, J3,4 = 6.5Hz, H-3, Xylp), 5.16 (d,
1H, J1,2 = 4.1Hz, H-1, Xylp), 5.06 (d, 1H,
J1,2 = 1.7Hz, H-1, Manp), 5.04 (m, 1H, H-4, Xylp),
4.58 (d, 1H, J1,2 = 4.0Hz, H-1, Xylp); 13C NMR
(100MHz, CDCl3) d 166.0, 166.0, 165.4, 165.4, 165.4,
165.3, 165.2, 165.1, 165.0, 164.6, 164.5 (11C, 11PhCO),
100.0, 99.0, 98.9, 94.9, (4C, 4C-1), 75.9, 75.6, 75.3,
71.4, 69.9, 69.5, 69.5, 69.5, 69.2, 69.2, 68.8, 68.4, 68.2,
68.0, 67.9, 64.0, 63.4, 60.5 (C-2 to C-6). Anal. Calcd
for C101H82Cl3NO30: C, 63.98; H, 4.36. Found: C,
63.75; H, 4.41.
4.11. Methyl 3-O-allyl-4,6-O-benzylidene-a-D-mannopyr-
anoside (13)
To a solution of 12 (4.35g, 21.2mmol) in CH3OH
(50mL) was added Bu2SnO (5.22g, 21.2mmol), and
the mixture was heated under reflux for 2h, then concen-
trated to dryness. The residue was diluted with toluene
(60mL), and then allyl bromide (17.1mL, 200mmol),
Bu4NI (7.38g, 20.0mmol) were added to the mixture.
The reaction was carried out at 60ꢁC for 24h, at which
time TLC (1:1 petroleum ether–EtOAc) showed that the
reaction was complete. The solution was concentrated to
dryness. The residue was passed through a short silica
gel column to give 13 (5.22g, 85%) as an amorphous
4.9. 2,3,4-Tri-O-benzoyl-b-D-xylopyranosyl-(1!2)-3,4,6-
tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-tri-O-
benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-benzoyl-a-D-
mannopyranose (10)
Deallylation of tetrasaccharide 9 (2.69g, 1.5mmol)
under the same conditions as that used for preparation
of 6 from 5 gave a residue, which was purified by flash
chromatography (1.5:1 petroleum ether–EtOAc) to give
10 (2.36g, 89%) as an amorphous solid. [a]D ꢀ69.5 (c
1
solid. [a]D +12.7 (c 1.0, CHCl3); H NMR (400MHz,
1
1.0, CHCl3); H NMR (400MHz, CDCl3): d 8.17–7.21
CDCl3): d 7.49–7.35 (m, 5H, 1Ph), 5.96–5.86 (m, 1H,
–CH2–CH@CH2), 5.59 (s, 1H, PhCH), 5.32–5.17 (m,
2H, –CH2–CH@CH2), 4.77 (d, 1H, J1,2 = 1.2Hz, H-1),
4.31–3.80 (m, 8H, H-2, H-3, H-4, H-5, H-6, –CH2–
CH@CH2), 3.38 (s, 3H, OCH3). Anal. Calcd for
C17H22O6: C, 63.35; H, 6.83. Found: C, 63.44; H, 6.80.
(m, 55H, 11Ph), 5.82 (dd, 1H, J3,4 = J4,5 = 9.9Hz, H-4
of Manp), 5.79–5.67 (m, 3H), 5.51 (m, 1H, H-4 of Xylp),
5.40 (dd, 1H, J1,2 = J2,3 = 5.7Hz, H-2 of Xylp), 5.33–
5.15 (m, 3H), 5.20 (d, 1H, J1,2 = 0.7Hz, H-1 of Manp),
5.09 (d, 1H, J1,2 = 4.2Hz, H-1 of Xylp), 5.06 (m, 1H,
H-4 of Xylp), 5.00 (d, 1H, J1,2 = 0.9Hz, H-1 of Manp),
4.87 (dd, 1H, J2,3 = 5.4Hz, J3,4 = 6.0Hz, H-3 of Xylp),
4.36 (d, 1H, J1,2 = 5.1Hz, H-1 of Xylp), 4.36–2.86 (m,
13H), 2.92–2.81 (dd, 2H, H-6a H-6b of Manp); 13C
NMR (100MHz, CDCl3) d 166.2, 166.1, 165.6, 165.5,
165.5, 165.3, 165.2, 165.2, 165.1, 164.7, 164.6 (11C,
11PhCO), 99.5, 99.1, 98.2, 92.2 (4C, 4C-1), 77.9, 77.3,
75.6, 75.8, 70.0, 69.8, 69.7, 69.6, 69.4, 69.2, 68.6, 68.3,
68.0, 67.9, 64.2, 63.9, 60.5, 60.0 (C-2 to C-6). Anal.
Calcd for C99H82O30: C, 67.89; H, 4.69. Found: C,
67.99; H, 4.63.
4.12. Methyl 2-O-acetyl-3-O-allyl-4,6-O-benzylidene-a-
D-mannopyranoside (14)
Compound 13 (4.14g, 12.86mmol) was dissolved in pyr-
idine (30mL), and Ac2O (6.00mL, 50mmol) was added.
The mixture was stirred at rt for 12h, then was concen-
trated to give a residue. Purification of the residue by col-
umn chromatography (3:1 petroleum ether–EtOAc) gave
14 (4.17g, 90.0%) as a syrup. [a]D +10.7 (c 1.0, CHCl3);
1H NMR (400MHz, CDCl3): d 7.50–7.33 (m, 5H, 1Ph),

W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
127
5.90–5.82 (m, 1H, –CH2–CH@CH2), 5.59 (s, 1H, PhCH),
5.31 (dd, 1H, J1,2 = 1.1Hz, J2,3 = 3.5Hz, H-2), 5.32–5.13
(m, 2H, –CH2–CH@CH2), 4.66 (d, 1H, H-1), 3.91 (dd,
1H, J3,4 = 9.9Hz, H-3), 4.66–3.92 (m, 6H, H-4, H-5, H-
6, –CH2–CH@CH2), 3.37 (s, 3H, OCH3), 2.14 (s, 3H,
CH3CO). Anal. Calcd for C19H24O7: C, 62.64; H, 6.59.
Found: C, 62.81; H, 6.52.
J1,2 = 6.4Hz, H-1 of Xylp), 4.62 (d, 1H, H-1 of Manp),
4.60 (dd, 1H, J5,6a = 1.6Hz, J6a,6b = 12.0Hz, H-6a of
Manp), 4.47 (dd, 1H, J4,5a = 4.8Hz, J5a,5b = 12.0Hz, H-
5a of Xylp), 4.36 (dd, 1H, J5,6b = 4.0Hz, J6a,6b = 12.0Hz,
H-6b of Manp), 4.20–4.06 (m, 2H, CH2@CHCH2O),
4.14 (dd, 1H, J3,4 = J4,5 = 9.3Hz, H-4 of Manp), 3.90
(d, 1H, J2,3 = 3.4Hz, H-3 of Manp), 3.81 (m, 1H, H-5
of Manp), 3.60 (dd, 1H, J4,5a = 8.2Hz, J5a,5b = 12.0Hz,
H-5b of Xylp), 3.32 (s, 3H, OCH3), 2.03 (s, 3H, CH3CO).
Anal. Calcd for C45H44O15: C, 65.53; H, 5.34. Found: C,
65.67; H, 5.32.
4.13. Methyl 2-O-acetyl-3-O-allyl-a-D-mannopyranoside
(15)
A mixture of 14 (4.1g, 11.26mmol) and 90% HOAc–H2O
(50mL) was stirred for 2h at 70ꢁC, then concentrated to
dryness. Purification of the residue by column chromato-
graphy (1:1 petroleum ether–EtOAc) gave 15 (2.95g,
90%) as an amorphous solid. [a]D +14.6 (c 1.0, CHCl3);
1H NMR (400MHz, CDCl3): d 5.93–5.83 (m, 1H,
4.16. Methyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!4)-2-O-acetyl-6-O-benzoyl-a-D-mannopyranoside
(18)
Deallylation of tetrasaccharide 17 (5.22g, 6.33mmol)
under the same conditions as that used for the prepara-
tion of 6 from 5 gave a residue. Then the residue was
passed through a short silica gel column to give 18
(4.21g, 85%) as an amorphous solid. [a]D ꢀ14.5 (c 1.3,
H2O); 1H NMR (400MHz, CDCl3): d 7.96–7.27 (m,
20H, 4PhH), 5.85 (dd, 1H, J2,3 = J3,4 = 9.3Hz, H-3 of
Xylp), 5.55 (dd, 1H, J1,2 = 7.3Hz, H-2 of Xylp), 5.40
(m, 1H, H-4 of Xylp), 5.17 (dd, 1H, J1,2 = 1.7Hz, H-2
of Manp), 4.86 (d, 1H, J1,2 = 7.3Hz, H-1 of Xylp),
4.68 (d, 1H, H-1 of Manp), 4.50 (dd, 1H,
J4,5a = 5.2Hz, J5a,5b = 11.7Hz, H-5a of Xylp), 4.36–
4.30 (m, 2H, H-6a, H-6b of Manp), 4.12 (dd, 1H,
J2,3 = 3.6Hz, H-3 of Manp), 3.90 (dd, 1H,
J3,4 = J4,5 = 9.7Hz, H-4 of Manp), 3.83 (m, 1H, H-5 of
Manp), 3.67 (dd, 1H, J4,5b = 9.6Hz, J5a,5b = 11.7Hz,
H-5b of Xylp), 3.33 (s, 3H, OCH3), 2.08 (s, 3H,
CH3CO). Anal. Calcd for C42H40O15: C, 64.29; H,
5.10. Found: C, 64.10; H, 5.15.
–CH2–CH@CH2), 5.26 (dd, 1H, J1,2 = 1.4Hz, J2,3
=
3.3Hz, H-2), 5.31–5.19 (m, 2H, –CH2–CH@CH2), 4.68
(d, 1H, H-1), 3.69 (dd, 1H, J3,4 = 9.9Hz, H-3), 4.66–
3.92 (m, 6H, H-4, H-5, H-6, –CH2–CH@CH2), 3.38 (s,
3H, OCH3), 2.12 (s, 3H, CH3CO). Anal. Calcd for
C12H20O7: C, 52.17; H, 7.25. Found: C, 52.30; H, 7.20.
4.14. Methyl 2-O-acetyl-3-O-allyl-6-O-benzoyl-a-D-
mannopyranoside (16)
Compound 15 (2.90g, 10.51mmol) was dissolved in
anhyd CH2Cl2 (30mL) containing pyridine (4.1mL,
50mmol), then under N2 protection and stirring, a solu-
tion of benzoyl chloride (1.23mL, 10.51mmol) in anhyd
dichloromethane (6mL) was added dropwise within
20min at 0ꢁC. The reaction temperature slowly raised
to rt. After stirring the mixture for 8h, TLC (3:1 petro-
leum ether–EtOAc) indicated that the reaction was com-
plete. The mixture was concentrated to dryness.
Purification by column chromatography (3:1 petroleum
ether–EtOAc) gave 16 (3.60g, 90%) as an amorphous
4.17. Methyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)-3,4,6-tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-
benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-tri-O-ben-
zoyl-b-D-xylopyranosyl-(1!4)]-2-O-acetyl-6-O-benzoyl-
a-D-mannopyranoside (19)
1
solid. [a]D +13.5 (c 1.0, CHCl3); H NMR (400MHz,
CDCl3): d 8.07–7.40 (m, 5H, 1Ph), 5.93–5.83 (m, 1H,
–CH2–CH@CH2), 5.27 (dd, 1H, J1,2 = 1.6Hz,
J2,3 = 3.2Hz, H-2), 5.31–5.18 (m, 2H, –CH2–CH@CH2),
4.71 (d, 1H, H-1), 4.64–4.61 (m, 2H, H-6a, H-6b), 3.73
(dd, 1H, J3,4 = 9.0Hz, H-3), 4.18–3.87 (m, 4H, H-4, H-
5, –CH2–CH@CH2), 3.39 (s, 3H, OCH3), 2.08 (s, 3H,
CH3CO). Anal. Calcd for C19H24O8: C, 60.00; H, 6.32.
Found: C, 60.18; H, 6.30.
As described in the general procedure, 11 (1.00g,
0.53mmol) and 17 (0.60g, 1.00mmol) were coupled,
and the product was purified by silica gel column chro-
matography with 1:1 petroleum ether–EtOAc as the elu-
ent to give 19 (0.91g, 70%) as an amorphous solid. [a]D
1
4.15. Methyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!4)-2-O-acetyl-3-O-allyl-6-O-benzoyl-a-D-mannopyr-
anoside (17)
ꢀ46.1 (c 0.5, CHCl3); H NMR (CDCl3, 400MHz): d
8.14–7.03 (m, 75H, 15PhH), 5.90 (dd, 1H,
J2,3 = 3.2Hz, J3,4 = 10.2Hz, H-3 of Manp), 5.86 (dd,
1H, J3,4 = J4,5 = 5.8Hz, H-3 of Xylp), 5.85 (dd, 1H,
As described in the general procedure, 16 (3.00g,
7.89mmol) and 2 (4.86g, 8.00mmol) were coupled, and
the product was purified by column chromatography
with 2.5:1 petroleum ether–EtOAc as the eluent to give
17 (5.85g, 90%) as an amorphous solid. [a]D ꢀ25.5 (c
J3,4 = J4,5 = 7.0Hz, H-3 of Xylp), 5.84 (dd, 1H, J3,4
J4,5 = 10.2Hz, H-4 of Manp), 5.71 (dd, 1H, J3,4
=
=
J4,5 = 10.0Hz, H-4 of Manp), 5.69 (m, 1H, H-4 of Xylp),
5.57–5.46 (m, 4H), 5.32 (m, 1H, H-4 of Xylp), 5.30 (d,
1H, J1,2 = 5.0Hz, H-1 of Xylp), 5.28 (s, 1H, H-1 of
Manp), 5.19 (d, 1H, J1,2 = 4.9Hz, H-1 of Xylp), 5.17
(s, 1H, H-1 of Manp), 5.02 (m, 1H, H-4 of Xylp), 5.00
(dd, 1H, H-2 of Manp), 4.68 (d, 1H, J1,2 = 4.8Hz, H-1
of Xylp), 4.56 (s, 1H, H-1 of Manp), 4.75–2.87 (m,
20H), 3.26 (s, 3H, OCH3), 1.81 (s, 3H, COCH3); 13C
NMR (100MHz, CDCl3): 170.3 (COCH3), 166.2,
1
1.0, CHCl3); H NMR (400MHz, CDCl3): d 7.98–7.17
(m, 20H, 4PhH), 5.96–5.87 (m, 1H, CH2@CHCH2O–),
5.78 (dd, 1H, J2,3 = J3,4 = 8.3Hz, H-3 of Xylp), 5.43
(dd, 1H, J1,2 = 6.4Hz, H-2 of Xylp), 5.32 (m, 1H, H-4
of Xylp), 5.26 (dd, 1H, J1,2 = 1.6Hz, H-2 of Manp),
5.34–5.18 (m, 2H, CH2@CHCH2O), 5.03 (d, 1H,

128
W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
166.0, 165.9, 165.7, 165.5, 165.4, 165.3, 165.3, 165.3,
165.3, 165.2, 164.6, 164.6, 164.5, 164.5 (15C, 15COPh),
102.6 (JC1-H1 163.4Hz), 99.7 (JC1-H1 163.4Hz), 98.6
(JC1-H1 164.1Hz), 98.5 (JC1-H1 173.4Hz), 97.7 (JC1-H1
174.2Hz), 97.0 (JC1-H1 176.1Hz) (6C, C-1), 78.9, 77.6,
75.2, 74.7, 74.6, 74.6, 74.1, 72.4, 71.6, 71.0, 70.3, 70.2,
69.9, 69.7, 69.7, 69.6, 69.3, 69.2, 68.7, 68.4, 67.7, 63.8,
63.2, 63.1, 62.6, 60.5, 59.7 (C-2 to C-6), 54.9 (OCH3),
20.51 (COCH3). Anal. Calcd for C141H120O44: C,
67.25; H, 4.77. Found: C, 67.50; H, 4.82.
centrated to dryness. Purification of the residue by silica
gel column chromatography (1:1.3 petroleum ether–
EtOAc) gave 22 (210mg, 78%) as an amorphous solid.
[a]D ꢀ61.7 (c 1.3, CHCl3); 1H NMR (400MHz, CDCl3):
d
8.20–6.90 (m, 75H, 15PhH), 6.09 (dd, 1H,
J3,4 = J4,5 = 9.8Hz, H-4, Manp), 6.00 (dd, 1H, J3,4
=
J4,5 = 9.2Hz, H-4, Manp), 5.92 (dd, 1H, J2,3 = 3.3Hz,
J3,4 = 9.8Hz, H-3 of Manp), 5.27 (d, 1H, J1,2 = 1.2Hz,
H-1, Manp), 5.16 (d, 1H, J1,2 = 1.0Hz, H-1, Manp),
5.10 (d, 1H, J1,2 = 1.1Hz, H-1, Manp), 4.88 (d, 1H,
J1,2 = 7.5Hz, H-1, Xylp), 4.80 (d, 1H, J1,2 = 7.0Hz, H-1,
Xylp), 4.72 (d, 1H, J1,2 = 7.1Hz, H-1, Xylp), 4.44 (dd,
1H, J1,2 = 1.1Hz, J2,3 = 3.3Hz, H-2, Manp), 4.13 (dd,
1H, J1,2 = 1.0Hz, J2,3 = 3.2Hz, H-2, Manp), 4.00
4.18. Methyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)-3,4,6-tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-
benzoyl-a-D-mannopyranosyl-(1!3)-[2,3,4-tri-O-ben-
zoyl-b-D-xylopyranosyl-(1!4)]-6-O-benzoyl-a-D-manno-
pyranoside (20)
(d, 1H, J1,2 = 7.0Hz, H-1, GlcpA), 3.79 (dd, 1H, J1,2
=
1.2Hz, J2,3 = 2.9Hz, H-2, Manp), 3.65 (s, 3H, COOCH3),
3.24 (s, 3H, OCH3), 2.13, 2.12, 2.03 (3s, 9H, 3COCH3).
13C NMR (100MHz, CDCI3): 171.2, 171.0, 169.8, 169.2
(4C, 3COCH3, COOMe), 167.3, 167.2, 166.1, 166.0,
165.8, 165.8, 165.4, 165.3, 165.3, 165.3, 165.3, 165.2,
165.2, 165.1, 164.6 (15C, 15COPh), 102.7 (JC1-H1
163.2Hz), 99.3 (JC1-H1 164.1Hz), 99.0 (JC1-H1 163.2Hz),
97.9 (JC1-H1 176.2Hz), 96.4 (JC1-H1 173.8Hz), 96.4 (JC1-
To a solution of 19 (600mg, 0.24mmol) in anhyd CH2Cl2
(10mL) was added anhyd MeOH (40mL), then acetyl
chloride (3.5mL) was added to the reaction mixture at
0ꢁC. The mixture was stirred at rt for 3days, TLC (1:1
petroleum ether–EtOAc) showed that the starting mate-
rial disappeared. The solution was neutralized with
Et3N, then concentrated to dryness. Purification of the
residue by silica gel column chromatography (1:1.2
petroleum ether–EtOAc) gave 20 (210mg, 45%) as an
173.4Hz), 95.7 (JC1-H1 166.8Hz) (7C, 7C-1), 54.8
H1
(OCH3), 52.8 (COOC3), 20.8, 20.8, 20.5 (3C, 3COCH3).
Anal. Calcd for C152H134O52: C, 65.13; H, 4.85. Found:
C, 65.32; H, 4.91; Negative-ESI MS calcd for
C152H134O52: [M] 2790.1. Found: [M+Na] 2812.7,
[M+K] 2828.7.
1
amorphous solid. [a]D ꢀ47.1 (c 1.0, CHCl3); H NMR
(400MHz, CDCl3): d 8.17–7.05 (m, 75H, 15PhH), 5.93
(dd, 1H, J3,4 = J2,3 = 5.6Hz, H-3 of Xylp), 5.92 (dd,
1H, J2,3 = 3.3Hz, J3,4 = 10.1Hz, H-3 of Manp), 5.89
(dd, 1H, J3,4 = J4,5 = 7.2Hz, H-3 of Xylp), 5.82 (dd,
1H, J2,3 = J3,4 = 10.2Hz, H-4 of Manp), 5.73 (dd, 1H,
J3,4 = J4,5 = 10.1Hz, H-4 of Manp), 5.71–5.48 (m, 5H),
5.39–5.33 (m, 2H), 5.27 (s, 1H, H-1 of Manp), 5.08 (d,
1H, J1,2 = 4.8Hz, H-1 of Xylp), 5.08–5.00 (m, 2H, H-4
of Xylp), 4.65 (d, 1H, J1,2 = 4.9Hz, H-1 of Xylp), 4.64
(s, 1H, H-1 of Manp), 4.53 (d, 1H, J1,2 = 4.9Hz, H-1 of
Xylp), 4.51 (s, 1H, H-1 of Manp), 4.55–2.98 (m, 19H),
3.16 (s, 3H, OCH3); 13C NMR (100MHz, CDCl3):
166.3, 166.0, 166.0, 165.5, 165.4, 165.4, 165.4, 165.3,
165.2, 165.2, 165.2, 164.8, 164.8, 164.6, 164.6 (15C,
15COPh), 102.4, 100.0, 99.1, 99.0, 98.0, 97.3 (6C, C-1),
77.2, 76.2, 74.8, 74.8, 74.8, 72.1, 71.5, 70.9, 70.3, 70.2,
70.1, 69.8, 69.7, 69.6, 69.5, 69.2, 69.1, 69.0, 68.7, 68.3,
67.6, 64.3, 64.1, 63.3, 62.9, 60.2, 59.9 (C-2 to C-6), 54.8
(OCH3). Anal. Calcd for C139H118O43: C, 67.42; H,
4.77. Found: C, 67.68; H, 4.72.
4.20. Methyl b-D-xylopyranosyl-(1!2)-a-D-mannopy-
ranosyl-(1!3)-[b-D-xylopyranosyl-(1!2)]-a-D-manno-
pyranosyl-(1!3)-[(b-D-glucopyranosyluronic acid)-
(1!2)][b-D-xylopyranosyl-(1!4)]-a-D-mannopyranoside,
ammonium salt (23)
Heptasaccharide 22 (200mg, 0.07mmol) was dissolved
in a satd methanolic ammonia (50mL). After 96h at
rt, water (1.0mL) was added to the mixture to cleave
the methyl ester. After stirring at rt for 5h, the reaction
mixture was concentrated, and purified on a Bio-Gel P2
column (eluent: water), affording the target heptasaccha-
ride 23 (63mg, 64.5%) as an amorphous solid. [a]D +56.9
1
(c 0.5, H2O); H NMR (D2O, 400MHz): d 5.05 (s, 1H,
H-1, Manp), 4.97 (s, 1H, H-1, Manp), 4.65 (s, 1H, H-
1, Manp), 4.59 (d, 1H, J1,2 = 8.0Hz, H-1, GlcpA), 4.38
(d, 1H, J1,2 = 8.7Hz, H-1, Xylp), 4.34 (d, 1H,
J1,2 = 8.5Hz, H-1, Xylp), 4.30 (d, 1H, J1,2 = 8.6Hz, H-
1, Xylp), 3.31 (s, 3H, OCH3); 13C NMR (100MHz,
D2O): 173.9 (–COONH4), 103.5, 103.5, 103.3, 102.7,
100.6, 99.1, 96.0 (7C, 7C-1), 78.4, 78.2, 76.2, 76.1,
76.0, 75.7, 74.1, 73.5, 73.4, 72.8, 72.2, 70.5, 70.2, 69.4,
66.9, 66.5, 66.2, 65.3, 65.3, 61.5, 60.4, 60.4 (C-2 to C-
6), 54.5 (OCH3). MALDI-TOF MS Calcd for the
ammonium salt of 22, C40H69NO34: 1107.9 [M]. Found:
1107.9 (M); 1113.0 (M ꢀ NHþ4 þ Naþ).
4.19. Methyl 2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)-3,4,6-tri-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-4,6-di-O-
benzoyl-a-D-mannopyranosyl-(1!3)-[methyl 2,3,4-tri-O-
acetyl-b-D-glucopyranosyluronate-(1!2)][2,3,4-tri-O-
benzoyl-b-D-xylopyranosyl-(1!4)]-6-O-benzoyl-a-D-
mannopyranoside (22)
To a cooled solution (0ꢁC) of 20 (200mg, 0.09mmol), 21
(92mg, 0.24mmol), and 2,4-lutidine (10lL, 0.10mmol) in
anhyd CH2Cl2 (20mL), was added silver triflate (45mg,
0.24mmol). The mixture was stirred at this temperature
for 6h, TLC (1:1.5 petroleum ether–EtOAc) showed that
the starting material disappeared. The solution was con-
4.21. Allyl 2,3,4,6-tetra-O-benzoyl-a-D-mannopyranosyl-
(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-
4,6-di-O-benzoyl-a-D-mannopyranoside (25)
As described in the general procedure, 24 (2.00g,
2.70mmol) and 8 (2.26g, 2.60mmol) were coupled,

W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
129
and the product was purified by column chromatogra-
phy with 2.5:1 petroleum ether–EtOAc as the eluent to
give 25 (3.60g, 95%) as an amorphous solid. [a]D
J2,3 = 3.2Hz, J3,4 = 9.7Hz, H-3 of Manp), 5.79 (dd,
1H, J2,3 = J3,4 = 5.4Hz, H-3 of Xylp), 5.76 (dd, 1H,
J3,4 = J4,5 = 9.8Hz, H-4 of Manp), 5.52 (d, 1H, H-2 of
Manp), 5.52 (m, 1H, H-4 of Xylp), 5.48 (dd, 1H,
J1,2 = 4.0Hz, H-2 of Xylp), 5.39 (s, 1H, H-1 of Manp),
5.25 (d, 1H, J1,2 = 4.0Hz, H-1 of Xylp), 4.80–4.03
(m, 10H, CH2@CHCH2O, H-5 of Xylp, H-2, 3, 5, 6 of
Manp). 13C NMR (100MHz, CDCl3): 166.1, 165.9,
165.6, 165.4, 165.3, 165.2, 165.1, 165.0, 164.8, (9C,
9COPh), 99.8, 98.6, 94.9 (3C, 3C-1), 75.6, 75.4, 71.7,
70.3, 69.7, 69.5, 69.8, 68.6, 68.0, 67.3, 63.3, 62.9, 60.3,
60.2 (C-2 to C-6). Anal. Calcd for C82H66Cl3NO24: C,
63.28; H, 4.24. Found: C, 63.43; H, 4.19.
1
ꢀ82.9 (c 1.2, CHCl3); H NMR (400MHz, CDCl3): d
8.06–7.21
(m,
45H,
9Ph),
6.00
(dd,
J3,4 = J4,5 = 9.9Hz, H-4 of Manp), 5.98 (dd, 1H, J2,3
1H,
=
J3,4 = 5.1Hz, H-3 of Xylp), 5.80 (m, 1H,
CH2@CHCH2O), 5.80–5.69 (m, 2H, H-3 of Manp, H-
2 of Xylp), 5.50 (m, 1H, H-4 of Xylp), 5.42–5.39 (m,
2H, H-4 of Manp, H-2 of Manp), 5.34 (s, 1H, H-1 of
Manp), 5.12 (d, 1H, J1,2 = 4.2Hz, H-1 of Xylp), 5.20–
5.11 (m, 2H, CH2@CHCH2O), 4.96 (d, 1H,
J1,2 = 0.9Hz, H-1 of Manp), 4.86–3.86 (m, 12H,
CH2@CHCH2O, H-5 of Xylp, H-2, 3, 5, 6 of Manp);
13C NMR (100MHz, CDCl3) d 166.2, 166.0, 165.7,
165.4, 165.3, 165.3, 165.1, 164.9, 164.4 (9C, 9PhCO),
118.2 (CH2@CHCH2O), 99.8, 98.6, 96.4 (3C, 3C-1),
77.6, 76.2, 75.9, 70.3, 69.8, 69.4, 69.0, 68.9, 68.6, 68.1,
67.6, 63.9, 63.9, 63.3 (C-2 to C-6). Anal. Calcd for
C83H70O24: C, 68.69; H, 4.83. Found: C, 68.62; H, 4.90.
4.24. Methyl 2,3,4,6-tetra-O-benzoyl-a-D-mannopyrano-
syl-(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)]-4,6-di-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!4)]-2-O-
acetyl-6-O-benzoyl-a-D-mannopyranoside (28)
As described in the general procedure, 27 (1.21g,
0.78mmol) and 17 (0.72g, 0.92mmol) were coupled,
and the product was purified by silica gel column chro-
matography with 1:1 petroleum ether–EtOAc as the elu-
ent to give 28 (1.36g, 80%) as an amorphous solid. [a]D
4.22. 2,3,4,6-Tetra-O-benzoyl-a-D-mannopyranosyl-
(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-
4,6-di-O-benzoyl-a-D-mannopyranose (26)
1
Deallylation of tetrasaccharide 25 (3.22g, 2.22mmol)
under the same conditions as that used for preparation
of 6 from 5 gave a residue, which was purified by flash
chromatography (2:1 petroleum ether–EtOAc) to give
26 (2.66g, 86%) as an amorphous solid. [a]D ꢀ30.4 (c
1.0, CHCl3); 1H NMR (CDCl3, 400MHz): d 8.06–
7.14 (m, 45H, 9 PhH), 6.03 (dd, 1H, J2,3 = 3.2Hz,
J3,4 = 10.1Hz, H-3 of Manp), 5.96 (dd, 1H,
J3,4 = J4,5 = 10.1Hz, H-4 of Manp), 5.77 (dd, 1H,
ꢀ16.9 (c 0.5, CHCl3); H NMR (CDCl3, 400MHz): d
8.16–7.04 (m, 65H, 13PhH), 6.15 (dd, 1H,
J3,4 = J4,5 = 10.0Hz, H-4 of Manp), 6.05 (dd, 1H,
J2,3 = 3.2Hz, J3,4 = 10.0Hz, H-3 of Manp), 5.98 (dd,
1H, J2,3 = J3,4 = 6.1Hz, H-3 of Xylp), 5.85 (dd, 1H,
J2,3 = J3,4 = 8.0Hz, H-3 of Xylp), 5.81 (dd, 1H,
J3,4 = J4,5 = 10.0Hz, H-4 of Manp), 5.66 (m, 1H, H-4
of Xylp), 5.62 (dd, 1H, J1,2 = 7.2Hz, J2,3 = 8.0Hz, H-2
of Xylp), 5.56 (dd, 1H, J1,2 = 4.0Hz, J2,3 = 6.1Hz, H-2
of Xylp), 5.53 (dd, 1H, H-2 of Manp), 5.50 (d, 1H,
J1,2 = 4.0Hz, H-1 of Xylp), 5.40 (m, 1H, H-4 of Xylp),
5.40 (s, 1H, H-1 of Manp), 5.35 (s, 1H, H-1 of Manp),
5.24 (dd, 1H, H-2 of Manp), 4.77 (d, 1H, J1,2 = 7.2Hz,
H-1 of Xylp), 4.55 (s, 1H, H-1 of Manp), 4.95–3.45 (m,
17H, H-5 of Xylp, H-2, 3, 4, 5, 6 of Manp), 3.15 (s,
3H, OCH3), 2.16 (s, 3H, COCH3); 13C NMR
(100MHz, CDCl3): 170.1 (COCH3), 166.1, 166.0,
165.9, 165.7, 165.6, 165.4, 165.3, 165.3, 165.2, 164.8,
164.5, 164.5, 164.3, 5 (13C, 13COPh), 102.8, 99.9, 98.2,
98.2, 97.5 (5C, 5C-1), 77.1, 77.1, 76.1, 74.7, 72.3, 71.6,
71.2, 70.5, 70.4, 70.0, 69.8, 69.6, 69.4, 69.0, 68.9, 68.7,
68.5, 66.9, 63.9, 63.3, 62.5, 62.1, 60.3 (C-2 to C-6), 54.9
(OCH3), 20.0 (COCH3). Anal. Calcd for C122H104O38:
C, 67.28; H, 4.78. Found: C, 67.18; H, 4.81.
J2,3 = J3,4 = 5.1Hz, H-3 of Xylp), 5.76 (dd, 1H, J3,4
=
J4,5 = 10.0Hz, H-4 of Manp), 5.52 (m, 1H, H-4 of Xylp),
5.44 (dd, 1H, H-2 of Manp), 5.38 (dd, 1H, J1,2 = 6.4Hz,
H-2 of Xylp), 5.37 (s, 1H, H-1 of Manp), 5.34 (d, 1H,
J1,2 = 6.4Hz, H-1 of Xylp), 4.94 (d, 1H, J1,2 = 1.2Hz,
H-1 of Manp), 4.97–3.35 (m, 12H, CH2@CHCH2O,
H-5 of Xylp, H-2, 3, 5, 6 of Manp); 13C NMR
(100MHz, CDCl3): 166.1, 165.7, 165.5, 165.2, 165.1,
165.0, 164.9, 164.9, 164.8, 164.6, (10C, 10COPh), 99.6,
98.3, 92.1 (3C, 3C-1), 75.6, 75.1, 70.4, 69.6, 69.4, 69.0,
68.8, 69.3, 68.0, 67.7, 63.9, 63.4, 60.4, 60.2 (C-2 to C-
6). Anal. Calcd for C80H66O24: C, 68.09; H, 4.68.
Found: C, 68.25; H, 4.72.
4.23. 2,3,4,6-Tetra-O-benzoyl-a-D-mannopyranosyl-
(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!2)]-
4,6-di-O-benzoyl-a-D-mannopyranosyl trichloroacetim-
idate (27)
4.25. Methyl 2,3,4,6-tetra-O-benzoyl-a-D-mannopyrano-
syl-(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)]-4,6-di-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!4)]-6-O-
benzoyl-a-D-mannopyranoside (29)
A mixture of 26 (2.60g, 1.84mmol), trichloroacetonitrile
(0.4mL, 4.0mmol) and 1,8-diazabicyclo[5.4.0]-undecene
(DBU) (0.3mL) in dry CH2Cl2 (10mL) was stirred
under nitrogen for 3h and then concentrated. The resi-
due was purified by flash chromatography (2:1 petro-
leum ether–EtOAc) to give 27 (2.58g, 89%) as an
Deacetylation of 28 (1.00g, 0.46mmol) under the
same conditions as that used for preparation of 20
from 19 gave the crude product, which was purified by
flash chromatography (1:1 petroleum ether–EtOAc) to
furnish 29 (390mg, 42%) as an amorphous solid. [a]D
+17.2 (c 1.0, CHCl3); 1H NMR (400MHz, CDCl3):
1
amorphous solid: [a]D ꢀ71.2 (c 1.3, CHCl3); H NMR
(400MHz, CDCl3): d 8.70 (s, 1H, CNHCCl3), 8.05–
7.19 (m, 45H, 9Ph), 6.46 (s, 1H, H-1, Manp), 6.10 (dd,
1H, J3,4 = J4,5 = 9.7Hz, H-4 of Manp), 6.04 (dd, 1H,
d
8.11–6.89 (m, 65H, 13Ph), 6.14 (dd, 1H,

130
W. Zhao, F. Kong / Bioorg. Med. Chem. 13 (2005) 121–130
J3,4 = J4,5 = 10.0Hz, H-4 of Manp), 5.95 (dd, 1H,
J2,3 = 3.3Hz, J3,4 = 10.0Hz, H-3 of Manp), 5.67 (dd,
1H, J2,3 = 4.5Hz, J3,4 = 5.3Hz, H-3 of Xylp), 5.58 (dd,
1H, J3,4 = J4,5 = 9.5Hz, H-4 of Manp), 5.55 (dd, 1H,
J2,3 = 4.5Hz, J3,4 = 5.3Hz, H-3 of Xylp), 5.51 (dd, 1H,
J1,2 = J2,3 = 5.1Hz, H-2 of Xylp), 5.44 (dd, 1H, H-2 of
Manp), 5.30 (m, 1H, H-4 of Xylp), 5.26 (dd, 1H,
J1,2 = J2,3 = 4.5Hz, H-2 of Xylp), 5.11 (m, 1H, H-4 of
Xylp), 5.04 (d, 1H, J1,2 = 1.1Hz, H-1 of Manp), 5.01
(d, 1H, J1,2 = 0.6Hz, H-1 of Manp), 5.02 (d, 1H,
4.27. Methyl a-D-mannopyranosyl-(1!3)-[b-D-xylo-
pyranosyl-(1!2)]-a-D-mannopyranosyl-(1!3)-[(b-D-
glucopyranosyluronic acid)-(1!2)][b-D-xylopyranosyl-
(1!4)]-a-D-mannopyranoside, ammonium salt (31)
Hexasaccharide 30 (245mg, 0.10mmol) was dissolved in
a satd methanolic ammonia (50mL). After 96h at rt,
water (1.0mL) was added to the mixture to cleave the
methyl ester. After stirring at rt for 5h, the reaction mix-
ture was concentrated, and purified on a Bio-Gel P2 col-
umn (eluent: water), affording the target hexasaccharide
31 (63mg, 64.0%) as an amorphous solid. [a]D +56.9 (c
J1,2 = 4.5Hz, H-1 of Xylp), 4.53 (d, 1H, J1,2
=
0.9Hz, H-1 of Manp), 4.50 (dd, 1H, J2,3 = 2.9Hz,
J3,4 = 9.9Hz, H-3 of Manp), 4.44 (d, 1H, J1,2 = 5.1Hz,
H-1 of Xylp), 4.42–3.86 (m, 18H, H-5 of Xylp, H-2,
3, 4, 5, 6 of Manp), 3.16 (s, 3H, OCH3); 13C
NMR (100MHz, CDCl3) d 166.3, 166.0, 166.0, 165.6,
165.6, 165.4, 165.4, 165.4, 165.3, 164.9, 164.9, 164.7,
164.5 (13C, 13PhCO), 102.5, 100.1, 99.6, 98.8, 98.1
(5C, 5C-1), 76.1, 75.2, 72.1, 71.6, 70.6, 70.3, 70.0, 69.9,
69.6, 69.5, 69.3, 69.2, 69.1, 68.7, 66.9, 65.5, 64.2, 63.0,
62.9, 62.3, 60.7 (C-2 to C-6), 54.9 (OCH3). Anal. Calcd
for C120H102O37: C, 67.48; H, 4.78. Found: C, 67.62; H,
4.70.
1
0.5, H2O); H NMR (D2O, 400MHz): d 5.00 (s, 1H,
H-1, Manp), 4.94 (s, 1H, H-1, Manp), 4.63 (s, 1H, H-
1, Manp), 4.50 (d, 1H, J1,2 = 8.2Hz, H-1, GlcpA), 4.38
(d, 1H, J1,2 = 8.8Hz, H-1, Xylp), 4.20 (d, 1H,
J1,2 = 8.4Hz, H-1, Xylp), 3.20 (s, 3H, OCH3); 13C
NMR (100MHz, D2O): 173.9 (–COONH4), 103.5,
103.5, 102.7, 100.6, 99.1, 97.0 (6C, 6C-1), 78.4, 78.2,
76.2, 76.1, 76.0, 75.7, 74.1, 73.5, 73.4, 72.8, 72.2, 70.5,
70.2, 69.4, 66.9, 66.5, 66.2, 65.3, 65.3, 61.5, 60.4, 60.4
(C-2 to C-6), 55.0 (OCH3). MALDI-TOF MS calcd
for the ammonium salt of 31, C35H61O30N: 975.8 [M].
Found: 975.8 (M); 980.9 (M ꢀ NHþ4 þ Naþ).
4.26. Methyl 2,3,4,6-tetra-O-benzoyl-a-D-mannopyrano-
syl-(1!3)-[2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-
(1!2)]-4,6-di-O-benzoyl-a-D-mannopyranosyl-(1!3)-
[methyl 2,3,4-tri-O-acetyl-b-D-glucopyranosyluronate-
(1!2)][2,3,4-tri-O-benzoyl-b-D-xylopyranosyl-(1!4)]-6-
O-benzoyl-a-D-mannopyranoside (30)
Acknowledgements
This work was supported by The Chinese Academy of
Sciences (KZCX3-J-08) and by The National Natural
Science Foundation of China (Projects 30070185 and
39970864).
To a cooled solution (0ꢁC) of 29 (200mg, 0.11mmol), 21
(96mg, 0.26mmol), and 2,4-lutidine (7lL, 0.07mmol) in
anhyd CH2Cl2 (20mL), was added silver triflate (50mg,
0.26mmol). The mixture was stirred at this temperature
for 6h, TLC (1:1.5 petroleum ether–EtOAc) showed
that the starting material disappeared. The solution
was concentrated to dryness. Purification of the residue
by flash column chromatography (1:1.2 petroleum
ether–EtOAc) gave 30 (208mg, 76%) as an amorphous
References and notes
1. Evans, E. E. Proc. Soc. Exp. Biol. Med. 1949, 71, 644–
652.
2. Wilson, D. E.; Bennett, G. E.; Bailey, J. W. Proc. Soc. Exp.
Biol. Med. 1968, 127, 820–827.
3. (a) Bhattacharjee, A. K.; Kwon-Chung, K. J.; Glaudemans,
C. P. J. Carbohydr. Res. 1979, 73, 183–192; (b) Bhatta-
charjee, A. K.; Kwon-Chung, K. J.; Glaudemans, C. P. J.
Carbohydr. Res. 1981, 95, 237–245; (c) Bhattacharjee, A.
K.; Kwon-Chung, K. J.; Glaudemans, C. P. J. Carbohydr.
Res. 1980, 82, 103–111; (d) Bhattacharjee, A. K.; Kwon-
Chung, K. J.; Glaudemans, C. P. J. Mol. Immunol. 1979, 16,
531–540.
4. (a) Zhang, J.; Kong, F. Tetrahedron Lett. 2003, 1839–1850;
(b) Zhang, J.; Kong, F. Bioorg. Med. Chem. 2003, 11, 4027–
4037; (c) Zhang, J.; Kong, F. Carbohydr. Res. 2003, 338,
1719–1725.
1
solid. [a]D ꢀ66.7 (c 1.3, CHCl3); H NMR (400MHz,
CDCl3): d 8.24–6.80 (m, 65H, 13PhH), 6.25 (dd,
1H, J1,2 = J2,3 = 7.7Hz, H-2 of Xylp), 6.11 (dd, 1H,
J3,4 = J4,5 = 9.6Hz, H-4 of Manp), 5.74 (dd, 1H,
J3,4 = J4,5 = 9.9Hz, H-4 of Manp), 5.21 (d, 1H, J1,2
=
1.1Hz, H-1 of Manp), 5.13 (d, 1H, J1,2 = 0.8Hz, H-1
of Manp), 5.11 (d, 1H, J1,2 = 1.1Hz, H-1 of Manp),
4.80 (d, 1H, J1,2 = 7.9Hz, H-1 of Xylp), 4.73 (d, 1H,
J1,2 = 7.7Hz, H-1 of Xylp), 4.50 (dd, 1H, H-2 of Manp),
4.33 (dd, 1H, J1,2 = 1.1Hz, J2,3 = 3.2Hz, H-2 of Manp),
4.14 (dd, 1H, J1,2 = 0.8Hz, J2,3 = 2.8Hz, H-2 of Manp),
4.06 (d, 1H, J1,2 = 7.2Hz, H-1 of GlcpA), 3.55 (s, 3H,
COOCH3), 3.28 (s, 3H, OCH3), 2.14, 2.12, 2.01 (3s,
9H, 3COCH3). 13C NMR (100MHz, CDCl3): 171.2,
171.0, 169.5, 169.4 (4C, 3COCH3, COOMe), 167.2,
167.2, 166.1, 166.1, 165.8, 165.4, 165.4, 165.3, 165.3,
165.2, 165.1, 165.1, 164.2 (13C, 13COPh), 102.6, 100.6,
100.6, 99.6, 98.9, 98.2 (6C, 6C-1), 54.9 (OCH3), 52.6
(COOCH3), 20.5, 20.4, 20.1 (3C, 3COCH3). Anal. Calcd
for C133H118O46: C, 65.13; H, 4.85. Found: C, 65.32; H,
4.90. Negative-ESI MS calcd for C133H118O46: [M]
2450.2. Found: [M+Na] 2472.7. [M+K] 2488.6.
5. Zhao, W.; Kong, F. Carbohydr. Res. 2004, 339, 1779–
1786.
6. (a) Garegg, P. J.; Olsson, L.; Oscarson, S. J. Carbohydr.
Chem. 1993, 12, 195–203; (b) Garegg, P. J.; Olsson, L.;
Oscarson, S. Bioorg. Med. Chem. 1996, 4, 1867–1878.
7. Zegelaar-Jaarsveld, K.; Smits, S. A. W.; van der Marel, G.
A.; van Boom, J. H. Bioorg. Med. Chem. 1996, 4, 1819–
1823.
8. Schmidt, R. R.; Kinzy, W. Adv. Carbohydr. Chem. Bio-
chem. 1994, 50, 21–125.
9. (a) Byramova, N. E.; Ovchinnikov, M. V.; Backinowsky, L.
V.; Kochetkov, N. K. Carbohydr. Res. 1983, 124, c8; (b)
Zhu, Y.; Kong, F. Chin. J. Chem. 2001, 19, 119–123.