Synthesis of natural β-D-(1→3)-glucopyranosyl oligosaccharides

Article abstract of DOI:10.1016/S0008-6215(02)00276-8

β-D-(1→3)-Glucan core structure derivatives corresponding to schizophyllan, epiglucan and lentinan were synthesized efficiently. 4,6-O-Benzylidenated glucopyranosyl acceptors were found to be helpful in the attempted β-D-(1→3) bond formation. The epiglucan pentasaccharide showed a weak anti-tumor activity in preliminary mice tests.

Full text of DOI:10.1016/S0008-6215(02)00276-8

Carbohydrate Research 337 (2002) 1673–1678
www.elsevier.com/locate/carres
Synthesis of natural b- -(1“3)-glucopyranosyl oligosaccharides
D
Hongmei He, Feng Yang, Yuguo Du*
Research Center for Eco-En6ironmental Sciences, Chinese Academy of Sciences, PO Box 2871, Beijing 100085, PR China
Received 10 July 2002; accepted 7 August 2002
Abstract
b-
D
-(1“3)-Glucan core structure derivatives corresponding to schizophyllan, epiglucan and lentinan were synthesized effi-
-(1“3) bond formation.
ciently. 4,6-O-Benzylidenated glucopyranosyl acceptors were found to be helpful in the attempted b-
D
The epiglucan pentasaccharide showed a weak anti-tumor activity in preliminary mice tests. © 2002 Elsevier Science Ltd. All
rights reserved.
Keywords: Glycosylation; Oligosaccharides; Regioselectivity
1. Introduction
Schizophyllan,2 scleroglucan,3 epiglucan4 and lentinan5
are the most well-known members of this group of
polysaccharides. It has been suggested that the im-
A family of glucans which contain a main chain of
b-
D
-(1“3)-glucopyranosyl units, and are substituted at
-glucopyranosyl side chains
munopharmacological activity of soluble (1“3)-b-D-
O-6 by a single unit of b-
D
glucans are related to the organization of the
(1“3)-b-linked backbone into a triple helix, the fre-
quency and the complexity of side-branching and to
their molecular weight.6 However, Tsuzuki and co-
workers7 found that the conformation of the glucans,
either single or triple helix, is independent of the hema-
topoietic response. These results encourage chemists to
prepare the structures of the minimum size of the
natural polysaccharides and investigate their structure–
activity relationships.8
have received considerable attention because of their
antitumor activity (immunomodulating action).1
In our previous synthesis of the sanqi hexasaccharide
via a 3+3 strategy,9 we obtained an unexpected a-
product even using an O-2 acetylated donor under
standard glycosylation conditions. Further investi-
gation10 showed that this a priority is especially general
in complex carbohydrate coupling reactions for at-
tempted 1“3 glycosidic bond formation. To find an
easy way of preparing branched b- -(1“3)-glucan core
D
Scheme 1. Reagents and conditions: (a) NIS, TMSOTf,
CH2Cl2; 45% for 5; 39% for 7; (b) TMSOTf, CH2Cl2; 68% for
5; 85% for 7; 84% for 10; (c) Ac2O, Pyr; (d) 50% TFA, 2:1
CH3CN–CH2Cl2, 35 °C; 88%; (e) NBS, CH2Cl2, H2O;
Cl3CCN, DBU; 73% for two steps; (f) NaOMe, MeOH; 79%.
structures, we tried to use a 4,6-O-benzylidenated
glucopyranosyl acceptor in the glycosylation and have
gotten some positive results. We now report the synthe-
sis of schizopyllan, epiglucan and lentinan core struc-
ture derivatives, which contain
a
b-D-(1“3)-
glucopyranosyl backbone, using 4,6-O-benzylidene-pro-
tected glucopyranosyl acceptors under usual glycosyla-
tion conditions.
* Corresponding author. Tel.: +86-10-62914475; fax: +
86-10-62923563
E-mail address: ygdu@mail.rcees.ac.cn (Y. Du).
0008-6215/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(02)00276-8

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H. He et al. / Carbohydrate Research 337 (2002) 1673–1678
J2,3 9.8 Hz) and l 4.40 ppm (d, H-1II, J 8.0 Hz) in the
1H NMR spectrum of its acetylated derivative, 6. When
methyl 2-O-benzoyl-4,6-O-benzylidene-a- -glucopyran-
D
oside (4) was used as acceptor in the above reaction,
b-(1“3)-linked schizophyllan tetrasaccharide 7 was ob-
tained in 39%. The yield of this reaction improved
significantly when using the corresponding trisaccharide
imidate donor 2 (85%). Hydrolysis of 4,6-O-benzyli-
dene of 7 with 50% trifluoroacetic acid in 2:1 CH3CN–
CH2Cl2 at 35 °C gave tetrasaccharide diol 8, which was
further subjected to the regioselective glycosylation with
9, furnished fully protected pentasaccharide 10 in a
total yield of 74%. Removal of the acyl groups from 10
with NaOMe in MeOH accomplished the synthesis of
the epiglucan core structure derivative 11. 13C NMR
spectroscopy of 11 showed the reducing end a C-1I at
99.94 ppm, and all other b C-1s (C-1II-V) appeared at
103.48 ppm. It is worth noting that direct condensation
of 1 or 2 with disaccharide acceptor 12 under standard
glycosylation conditions gave no coupled products. In-
terestingly, when trichloroacetimidate 2 was coupled
Scheme 2. Reagents and conditions: (a) NIS, TMSOTf,
CH2Cl2; (b) TMSOTf, CH2Cl2; 85% for 14; (c) Ac2O, Pyr.
with isopropyl 4,6-O-benzylidene-1-thio-b-D-glucopy-
ranoside (13) in CH2Cl2 with TMSOTf as catalyst, the
b-(1“3)-linked tetrasaccharide 14 was obtained in high
yield (Scheme 2). The expected regio- and stereoselec-
tivity of this reaction was further confirmed by its
acetylated analog 15 showing H-2I at l 4.88 ppm (J 9.6,
10.4 Hz), while H-1II appeared at l 4.32 ppm (J1,2 8.0
Hz).
Encouraged by this model study, we envisaged a
facile synthesis of the lentinan hexsaccharide derivative
as shown in Scheme 3. TMSOTf-catalyzed condensa-
tion of 4,6-O-benzylidene-2,3-di-O-benzoyl-a-
pyranosyl trichloroacetimidate (16) and 1,2:5,6-di-O-
isopropylidene-a- -glucopyranose (17) in CH2Cl2 at
D-gluco-
D
0 °C gave disaccharide 18, which was deacylated with
NaOMe in MeOH to furnish the diol 19 in 56% yield.
Critical glycosylation of trisaccharide imidate 2 and
disaccharide diol 19 using the method as described in
the model study gave desired pentasaccharide 20 as the
predominant products. Acetylation of 20 with acetic
anhydride in pyridine followed by CeCl3·7H2O cata-
lyzed regioselective O-deisopropylidenation12 gave pen-
tasaccharide diol 21 in a good yield. The primary
hydroxyl group favored the glycosylation of 21 with 9
in CH2Cl2 at −15 °C to furnish the lentinan hexasac-
charide derivative 22 in excellent yield. 2D COSY ex-
periments showed C-1III at l 100.11 ppm in the 13C
NMR, while the corresponding H-1III appeared at l
Scheme 3. Reagents and conditions: (a) TMSOTf, CH2Cl2;
56% for 18; 78% for 20; 87% for 22; (b) NaOMe, MeOH; (c)
Ac2O, Pyr; then CeCl3·7H2O; 78% for two steps; (d) 90%
TFA, rt, 30 min; then NaOMe, MeOH; 53%.
2. Results and discussion
We have shown that the coupling reaction of trisac-
charide thioglycoside 1 or trichloroacetimidate donor 2
and octyl 2,4,6-tri-O-acetyl-b-D-glucopyranoside gave
a,b-mixed tetrasaccharide.10 To take the advantage of
our easy entry11 to the trisaccharide donor 1 in the
1
preparation of b-
tried the glycosylation of thioglycoside 1 and methyl
4,6-O-benzylidene-a- -glucopyranoside (3) in the pres-
D
-(1“3)-glucan core structures, we
4.33 ppm (d, J 8.0 Hz) in the H NMR, indicating a b
linkage between sugar residue II and III in 22. Full
deprotection of 22 with 90% TFA, followed by deac-
ylation with MeONa in MeOH, afforded hexasaccha-
ride 23 in 53% yield.
D
ence of NIS (2.5 equiv) and TMSOTf (0.3 equiv), and
obtained the undesired b-(1“2) product 5 in a modest
yield (Scheme 1). The b-(1“2) linkage in 5 was confi-
rmed by the peaks at l 3.68 ppm (dd, H-2I, J1,2 3.5 Hz,
The antitumor activity of epiglucan pentasaccharide
11 was preliminarily studied according to the method

H. He et al. / Carbohydrate Research 337 (2002) 1673–1678
1675
described by Sasaki and co-workers.5b Kun-min mice
weighing about 20 g and seven days Sarcoma-180
ascites (5×106 cells) were used for the bioassay.
Lentinan (imported from Japan for medical usage) and
Cisplatin® (CDDP) were selected as the positive con-
trols in the parallel tests. The epiglucan pentasaccharide
and lentinan were injected daily for 14 days, while
CDDP was given every other day. The tumor inhibition
ratios for 11, lentinan and CDDP were 21–25%
(dosage: 1.0–10 mg/kg/day), 31–36% (dosage: 1 mg/
Kg/day) and 55–57% (dosage: 2 mg/kg/every other
day), respectively.
In conclusion, we have described the synthesis of the
core structure derivatives of schizophyllan, epiglucan
and lentinan. From our experiments, 4,6-O-benzylide-
nated glucopyranosyl acceptors seem to be helpful for b
bond formation in complex oligosaccharide synthesis. It
would appear that the 3-OH groups in the current
acceptors are less hindered compared to those in their
4,6-diacylated counterparts. Apparently the orthoester
intermediate is formed and rearranges smoothly under
standard glycosylation conditions to give a high yield of
the product.
Hz, H-3III), 7.25–8.03 (m, 40 H, Ph). Anal. Calcd for
C81H74O25S: C, 65.76; H, 5.04. Found: C, 65.42; H,
5.09.
Methyl 2,3,4,6-tetra-O-benzoyl-i-
(1“3)-[2,3,4,6-tetra-O-benzoyl-i- -glucopyranosyl-(1
“6)]-2,4-di-O-acetyl-i- -glucopyranosyl-(1“2)-3-O-
acetyl-4,6-O-benzylidene-h- -glucopyranoside (6).—(a)
D-glucopyranosyl-
D
D
D
Method A: To a solution of 1 (300 mg, 0.203 mmol)
and 3 (56 mg, 0.2 mmol) in anhyd CH2Cl2 (6 mL) was
added NIS (114 mg, 0.507 mmol) and TMSOTf (11 mL,
0.06 mmol) at −15 °C. The mixture was stirred under
these conditions for 2 h and neutralized with Et3N. The
solution was concentrated and subjected to the column
chromatography (2:3 EtOAc–petroleum ether) to give
syrupy 5 (153 mg, 45%). (b) Method B: To a solution of
2 (314 mg, 0.2 mmol) and 3 (51 mg, 0.18 mmol) in
anhyd CH2Cl2 (4 mL) was added TMSOTf (6 mL, 0.03
,
mmol) in the presence of 4 A molecular sieves under an
N2 atmosphere at −15 °C. The mixture was stirred
under these conditions for 1.5 h, then neutralized with
Et3N. The solution was concentrated and subjected to
column chromatography (2:3 EtOAc–petroleum ether)
to give syrupy 5 (207 mg, 68%), which was further
acetylated with AC2O in pyridine for 4 h, to give 6 (267
1
mg, 99%) as a syrup: [h]D +45° (c 2.4, CHCl3); H
NMR: 1.89, 1.90, 2.04 (3 s, 9 H, 3 CH3CO), 3.21 (s, 3
H, CH3O), 3.58–3.63 (m, 2 H, H-5I, H-5II), 3.68 (dd, 1
H, J 3.5, 9.8 Hz, H-2I), 3.78–3.90 (m, 5 H, 2 H-6I, 2
H-6II, H-3II), 4.12–4.20 (m, 3 H, H-5IV, H-5III, H-4I),
4.40 (d, 1 H, J 8.0 Hz, H-1II), 4.48, 4.50 (2 d, 2 H, J 5.2
Hz, 2 H-6), 4.60, 4.64 (2 dd, 2 H, J 3.0, 12.1 Hz, 2 H-6),
4.71 (d, 1 H, J 3.5 Hz, H-1I), 4.76 (t, 1 H, J 9.5 Hz,
H-4II), 4.81 (dd, 1 H, J 8.0, 9.2 Hz, H-2II), 4.86 (d, 1 H,
J 7.8 Hz, H-1III), 5.03 (d, 1 H, J 7.9 Hz, H-1IV), 5.39
(dd, 1 H, J 8.9, 9.2 Hz, H-2IV), 5.43 (t, 1 H, J 9.8 Hz,
H-3I), 5.49 (dd, 1 H, J 8.0, 9.5 Hz, H-2III), 5.50 (s, 1 H,
PhCH), 5.65 (dd, 1 H, J 3.2, 9.7 Hz, H-4IV), 5.69 (dd,
1 H, J 3.0, 9.6 Hz, H-4III), 5.88 (t, 1 H, J 9.6 Hz,
H-3III), 5.91 (t, 1 H, J 9.6 Hz, H-3IV). 13C NMR: 20.44,
20.50, 20.96, 55.46, 62.21, 62.99, 63.12, 68.04, 68.75,
68.94, 69.45, 69.54, 70.48, 71.92 (2 C), 72.00, 72.49,
72.55, 72.91, 74.30, 77.55, 78.42, 79.32, 99.92 (C-1I),
100.77 (PhCH), 101.02, 101.16, 101.30 (C-1II,III,IV),
164.95, 165.08, 165.14, 165.65, 165.82, 166.02 (2 C),
168.11, 169.19, 169.44 (CO). MALDITOF-MS Calcd
for C94H86O32: 1726 [M]. Found 1749.5 [M+Na]+.
3. Experimental
General methods.—Optical rotations were deter-
mined at 25 °C with a Perkin–Elmer Model 241-Mc
automatic polarimeter. H, 13C NMR and H–1H, H–
13C COSY spectra were recorded with a Bruker ARX
400 spectrometer for solutions in CDCl3 or D2O.
Chemical shifts are given in ppm downfield from inter-
nal Me4Si. Mass spectra were measured using either
MALDITOF-MS with a-cyano-4-hydroxycinnamic
acid (CCA) as the matrix or a VG PLATFORM mass
spectrometer using the ESI technique. Thin-layer chro-
matography (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 lamp.
1
1
1
Isopropyl
2,3,4,6-tetra-O-benzoyl-i-D-glucopyran-
osyl-(1“3)-[2,3,4,6-tetra-O-benzoyl-i-
osyl-(1“6)]-2,4-di-O-acetyl-1-thio-i-
D
-glucopyran-
-glucopyran-
D
oside (1).—Compound 1 was prepared according to
our previous general method.11 [h]D −5° (c 4, CHCl3);
1H NMR: 0.92, 1.02 (2 d, 6 H, 2 CH3), 1.90, 1.92 (2 s,
6 H, 2 CH3CO), 2.73–2.81 (m, 1 H, SCH), 3.50 (br t, 1
H, H-5I), 3.66 (dd, 1 H, J 8.0, 11.2 Hz, H-6aI), 3.86–
3.90 (m, 2 H, H-3I, H-6bI), 4.10–4.15 (m, 2 H, H-5III,
H-5II), 4.25 (d, 1 H, J 9.6 Hz, H-1I), 4.45, 4.50, 4.60,
4.65 (4 dd, 4 H, 2 H-6II, 2 H-6III), 4.76 (t, 1 H, H-2I),
4.81 (t, 1 H, J 9.6 Hz, H-4I), 4.92 (d, 1 H, J 8.7 Hz,
H-1II), 4.95 (d, 1 H, J 8.0 Hz, H-1III), 5.39 (dd, 1 H, J
8.0, 9.7 Hz, H-2III), 5.51 (dd, 1 H, J 8.7, 9.4 Hz, H-2II),
5.64 (t, 1 H, J 9.7 Hz, H-4III), 5.68 (t, 1 H, J 9.4 Hz,
H-4II), 5.85 (t, 1 H, J 9.4 Hz, H-3II), 5.90 (t, 1 H, J 9.7
Methyl 2,3,4,6-tetra-O-benzoyl-i-
(1“3)-[2,3,4,6-tetra-O-benzoyl-i- -glucopyranosyl-(1
“6)]-2,4-di-O-acetyl-i- -glucopyranosyl-(1“3)-2-O-
benzoyl-4,6-O-benzylidene-h- -glucopyranoside (7).—
D-glucopyranosyl-
D
D
D
To a solution of 2 (1.1 g, 0.7 mmol) and 4 (258 mg, 0.67
mmol) in anhyd CH2Cl2 (10 mL) was added TMSOTf
,
(15 mL, 0.08 mmol) in the presence of 4 A molecular
sieves under an N2 atmosphere at 0 °C. The mixture
was stirred under these conditions for 1.5 h, then
neutralized with Et3N and concentrated. The residue
was subjected to column chromatography (2:3 EtOAc–

1676
H. He et al. / Carbohydrate Research 337 (2002) 1673–1678
petroleum ether) to give 7 (1.068 g, 85%) as a syrup:
[h]D −16° (c 4, CHCl3); H NMR: 1.49, 1.93 (2 s, 6 H,
and the filtrate was then concentrated. The residue was
purified on an LH-20 column to give 11 (336 mg, 79%)
as an amorphous solid: [h]D +19° (c 1, H2O); 13C
NMR (100 MHz, D2O): 55.95 (C-6), 61.43 (2 C-6),
68.57 (C-6I), 68.75 (C-6II), 70.32 (3 C-4), 71.16 (C-5),
75.14 (C-5), 76.28 (2 C-5), 76.35 (C-5), 76.57 (C-3IV,
C-3V), 76.70 (C-3III), 83.62 (C-3I), 84.69 (C-3II), 99.94
(C-1I), 103.48 (4 C-1). ESIMS Calcd for C31H54O26: 842
[M]. Found 841.3 [M−H]+.
1
2 CH3CO), 2.91 (dt, 1 H, J 9.9, 2.5 Hz), 3.38 (s, 3 H,
OCH3), 3.42–3.55 (m, 3 H), 3.64 (dd, 1 H, J 2.0, 11.9
Hz), 3.70–3.75 (m, 2 H), 3.80–3.95 (m, 3 H), 4.40 (dd,
1 H, J 10.8, 4.7 Hz), 4.47 (d, 1 H, J 8.0 Hz, H-1II), 4.52
(d, 1 H, J 8.0, 9.4 Hz, H-1III), 4.56–4.65 (m, 4 H), 4.84
(t, 1 H, J 9.4 Hz, H-2II), 4.95 (t, 1 H, J 9.5 Hz, H-4II),
4.92–4.96 (m, 2 H, H-1I, H-2I), 5.30 (d, 1 H, J-
8.1 Hz, H-1IV), 5.31 (dd, 1 H, J 8.0, 9.4 Hz, H-2III), 5.45
(dd, 1 H, J 8.0, 9.6 Hz, H-2IV), 5.54 (t, 1 H, J 9.5 Hz,
H-4III), 5.59 (t, 1 H, J 9.6 Hz, H-4IV), 5.64 (s, 1 H,
PhCH), 5.75 (t, 1 H, J 9.6 Hz, H-3IV), 5.77 (t, 1 H, J 9.6
Hz, H-3III), 7.18–8.24 (m, 50 H, Ph). Anal. Calcd for
C99H88O32: C, 66.44; H, 4.96. Found: C, 66.79; H, 5.05.
Isopropyl
osyl-(1“3)-[2,3,4,6-tetra-O-benzoyl-i-
osyl - (1“6)] - 2,4 - di - O - acetyl - i -
“3)-2-O-benzoyl-4,6-O-benzylidene-1-thio-i-
2,3,4,6-tetra-O-benzoyl-i-
D
-glucopyran-
-glucopyran-
D
D
- glucopyranosyl-(1
-gluco-
D
pyranoside (15).—Compounds 2 (150 mg, 0.096 mmol)
and 13 (31 mg, 0.095 mmol) were coupled as described
in the preparation of 5 (Method B) to afford 14, which
was further acetylated with acetic anhydride in pyridine
to give 15 as a syrup (143 mg, 85% for two steps): [h]D
Methyl 2,3,4,6-tetra-O-benzoyl-i-
(1“3)-[2,3,4,6-tetra-O-benzoyl-i- -glucopyranosyl-(1
“6)]-2,4-di-O-acetyl-i- -glucopyranosyl-(1“3)-[2,3,
4,6-tetra-O-benzoyl-i- -glucopyranosyl-(1“6)]-2-O-
benzoyl-h- -glucopyranoside (10). Compound 7 (900
D-glucopyranosyl-
D
D
D
1
−18° (c 0.3, CHCl3); H NMR: 1.32, 1.38 (2 d, 6 H, J
D
6.6 Hz, CH3), 1.90, 1.95, 1.96 (3 s, 9 H, 3 CH3CO), 2.97
(dt, 1 H, J 9.8, 2.5 Hz, H-5I), 3.24–3.31 (m, 1 H, SCH),
3.43 (br t, 1 H, H-5II), 3.49, 3.57 (2 dd, 2 H, J 2.5, 9.8
Hz, H-6I), 3.68 (t, 1 H, J 9.8, H-4I), 3.73 (br d, 1 H, J
12.4, B1 Hz, H-6), 3.82 (d, 1 H, J 9.6 Hz, H-1I), 3.85
(dd, 1 H, J 3.8, 12.9 Hz, H-6), 3.89 (t, 1 H, J 10.4 Hz,
H-3II), 4.08–4.24 (m, 3 H, H-5III, 3I, 5IV), 4.32 (d, 1 H,
J 8.0, H-1II), 4.45–4.61 (m, 3 H, H-6), 4.81 (d, 1 H, J
7.9 Hz, H-1IV), 4.88 (dd, 1 H, J 9.6, 10.4 Hz, H-2I), 4.98
(dd, 1 H, J 8.0, 9.8 Hz, H-2II), 5.02 (d, 1 H, J 8.0 Hz,
H-1III), 5.08 (d, 1 H, J 10.4, H-6), 5.30–5.35 (m, 2 H,
H-4II, 2III), 5.41 (dd, 1 H, J 8.0, 9.7 Hz, H-2IV),
5.56–5.73 (m, 4 H, H-4III, 4IV, 3III, PhCH), 5.87 (t, 1 H,
J 9.7 Hz, H-3IV), 7.20–8.24 (m, 45 H, Ph). MALDI-
TOF-MS Calcd for C96H90O31S: 1770.5 [M]. Found
1793.3 [M+Na]+.
mg, 0.5 mmol) was treated with 50% TFA (3 mL) in 2:1
CH3CN–CH2Cl2 (15 mL) at 35 °C for 3 h, then con-
centrated and subjected to column chromatography
(3:2 EtOAc–petroleum ether) to give 8 (753 mg, 88%).
Compounds 8 (790 mg, 0.46 mmol) and 9 (363 mg, 0.49
mmol) were reacted as described in the preparation of 7
to give syrupy 10 (889 mg, 84%): [h]D −4° (c 12,
1
CHCl3); H NMR: 1.33, 1.89 (2 s, 6 H, 2 CH3CO), 3.19
(s, 3 H, CH3CO), 3.40–3.45 (m, 2 H, H-3, H-5), 3.60 (t,
1 H, J 9.4 Hz, H-3), 3.65–3.72 (m, 2 H, 2 H-6),
3.90–4.00 (m, 3 H, H-4I,5,6), 4.04–4.10 (m, 2 H, 2
H-5), 4.20–4.23 (m, 1 H, H-5), 4.24 (d, 1 H, J 8.0 Hz,
H-1), 4.36 (d, 1 H, J 10.0 Hz, H-1), 4.40–4.55 (m, 5 H,
5 H-6), 4.58 (d, 1 H, J 7.8 Hz, H-1), 4.60–4.78 (m, 4 H,
H-2II, H-4II, 2 H-6), 4.91–4.94 (m, 2 H, J 3.6, 7.8 Hz,
H-1I, H-2I), 5.31 (d, 1 H, J 8.0 Hz, H-1), 5.33 (dd, 1 H,
J 8.0, 9.6 Hz, H-2), 5.50 (dd, 1 H, J 8.0, 9.8 Hz, H-2),
5.55–5.63 (m, 3 H, H-2, 2 H-4), 5.68 (t, 1 H, J 10.0 Hz,
H-4), 5.75 (t, 1 H, J 9.6 Hz, H-3), 5.87 (t, 1 H, J 9.6 Hz,
H-3), 5.93 (t, 1 H, J 10.0 Hz, H-3), 7.22-8.10 (m, 65 H,
Ph); 13C NMR: 19.71, 20.93, 54.62, 62.84, 62.97, 63.29,
66.81, 68.12, 69.41, 69.49, 69.74, 70.07, 71.39, 71.79,
71.80, 71.95, 71.96, 72.00, 72.52, 72.62, 72.67, 72.98,
75.01, 77.20, 78.30, 81.19, 96.61 (C-1I), 99.93, 100.79,
101.01, 102.38 (C-1II-V), 164.89, 164.96, 164.98, 165.05
(2C), 165.14, 165.17, 165.55, 165.65, 165.76, 165.94,
166.06, 166.12, 167.50, 169.27. Anal. Calcd for
4,6-O-Benzylidene-i-
D
-glucopyranosyl-(1“3)-1,2:5,
6-di-O-isopropylidene-h-
D
-glucopyranose (19).—Com-
pounds 16 (768 mg, 1.2 mmol) and 17 (260 mg, 1
mmol) were reacted as described in the synthesis of 7 to
give 18, along with some contaminants. The above
residue was treated with NaOMe (2 mL, 0.5 M in
MeOH) in MeOH (20 mL) for 4 h and neutralized with
Dowex-50W (H+) resin. After filtration, the filtrate was
concentrated and subjected to column chromatography
(1:1 EtOAc–petroleum ether) to give 19 as a syrup (286
1
mg, 56% for two steps): [h]D −22° (c 1, CHCl3); H
NMR: 1.33, 1.38, 1.46, 1.51 (4 s, 12 H, 2 C(CH3)2), 3.51
(ddd, 1 H, J 9.3, 4.7, 6.1 Hz, H-5II), 3.56 (t, 1 H, J 9.3
Hz, H-4II), 3.67 (dd, 1 H, J 9.3, 8.2 Hz, H-2II), 3.82 (dd,
1 H, J 10.1, 6.1 Hz, H-6aII), 3.84 (dd, 1 H, J 10.1, 4.7
Hz, H-6bII), 4.07 (dd, 1 H, J 4.4, 9.0 Hz, H-4I),
4.11–4.18 (m, 2 H, H-3II, H-6aI), 4.34–4.40 (m, 3 H,
H-3I, H-5I, H-6bI), 4.64 (d, 1 H, J 3.6 Hz, H-2I), 4.67
(d, 1 H, J 8.2 Hz, H-1II), 5.54 (s, 1 H, PhCH), 5.90 (d,
1 H, J 3.6 Hz, H-1I), 7.36–7.51 (m, 5 H, Ph). Anal.
C
126H110O41: C, 66.37; H, 4.86. Found: C, 66.11; H,
4.92.
Methyl i-
osyl-(1“6)]-i-
pyranosyl-(1“6)]-h-
D
-glucopyranosyl-(1“3)-[i-
-glucopyranosyl-(1“3)-[i-
-glucopyranoside (11).—To a so-
D
-glucopyran-
D
D
-gluco-
D
lution of 10 (1.15 g, 0.504 mmol) in MeOH (50 mL)
was added NaOMe (5 mL, 0.5 M in MeOH). The
mixture was stirred at rt overnight, neutralized with ion
exchange resin [Amberlite IR120 (H+)] and filtered,

H. He et al. / Carbohydrate Research 337 (2002) 1673–1678
1677
Calcd for C25H34O11: C, 58.81; H, 6.71. Found: C,
58.59; H, 6.80.
4.38 (d, 1 H, J 3.3 Hz, H-3I), 4.39 (dd, 1 H, J 5.6, 10.9
Hz, H-6bII), 4.48 (dd, 1 H, J 4.3, 11.7 Hz, H-6aV), 4.50
(dd, 1 H, J 5.1, 12.0 Hz, H-6aVI), 4.60 (dd, 1 H, J 2.6,
12.2 Hz, H-6bV), 4.67 (dd, 1 H, J 3.9, 11.7 Hz, H-6bVI),
4.84 (d, 1 H, J 7.5 Hz, H-1IV), 4.85–4.90 (m, 3 H, J 8.8
Hz, H-1II, H-2II, H-4III), 4.95 (t, 1 H, J 9.5 Hz, H-2III),
5.02 (d, 1 H, J 7.9 Hz, H-1VI), 5.21 (d, 1 H, J 8.0 Hz,
H-1V), 5.30 (t, 1 H, H-2V), 5.42 (dd, 1 H, J 7.5, 9.6 Hz,
H-2IV), 5.52 (s, 1 H, PhCH), 5.55–5.72 (m, 5 H, H-2VI,
H-4IV, H-3V, H-4V, H-4VI), 5.79 (d, 1 H, J 3.3 Hz,
H-1I), 5.89 (t, 1 H, J 9.6 Hz, H-3IV), 5.91 (t, 1 H, J 9.2
Hz, H-3VI), 7.25–8.00 (m, 65 H, Ph). Selected 13C
NMR: 99.71 (C-1II), 100.11 (C-1III), 100.31 (C-1V),
101.29 (C-1IV), 101.54 (C-1VI), 105.16(C-1I). Anal.
Calcd for C136H124O46: C, 65.48; H, 5.01. Found: C,
65.80; H, 5.07.
2,3,4,6-Tetra-O-benzoyl-i-
[2,3,4,6-tetra-O-benzoyl-i-
2,4-di-O-acetyl-i- -glucopyranosyl-(1“3)-2-O-acetyl-
4,6-O-benzylidene-i- -glucopyranosyl-(1“3)-1,2-O-
isopropylidene-h- -glucofuranose (21).—Compounds 2
D
-glucopyranosyl-(1“3)-
D
-glucopyranosyl-(1“6)]-
D
D
D
(595 mg, 0.38 mmol) and 19 (180 mg, 0.35 mmol) were
glycosylated under the conditions described in the
preparation of 14 to give the crude product (20, 570
mg), which was acetylated with Ac2O (1 mL) in pyri-
dine (5 mL) at 40 °C for 2 h. The mixture was concen-
trated to dryness with the help of toluene. To the above
residue (560 mg, 0.286 mmol) in CH3CN (10 mL) was
added CeCl3·7H2O (115 mg, 0.5 mmol) and two drops
of water. The mixture was refluxed for 6 h, at the end
of which time TLC showed the complete consumption
of the starting material. Concentration and purification
of the residue by silica gel column chromatography (2:1
EtOAc–petroleum ether) gave compound 21 (428 mg,
i -
“6)]-i-
(1“3)-[i-
D
- Glucopyranosyl - (1“3) - [i-
-glucopyranosyl-(1“3)-i-
-glucopyranosyl-(1“6)]-
D
-glucopyranosyl-(1
-glucopyranosyl-
-glucopyranose
D
D
D
D
(23).—Compounds 22 (230 mg, 0.092 mmol) was
treated with 90% trifluoroacetic acid for 30 min, con-
centrated and then co-concentrated with toluene, dis-
solved in methanol (40 mL), deacylated with sodium
methoxide (0.5 M, 1.2 mL), neutralized with Amberlite
IR-120 (H+), filtered and concentrated. The residue
was purified on a Sephadex LH-20 column (EtOAc,
then MeOH) to yield 23 (48 mg, 53%): [h]D +11° (c 1,
1
78%): [h]D −30° (c 1, CHCl3); H NMR: 1.24, 1.30 (2
s, 6 H, 2 CH3), 1.90, 1.94, 2.00 (3 s, 9 H, 3 CH3CO),
3.04–3.08 (m, 1 H, H-5IV), 3.22 (br s, 2 H, OH),
3.46–3.49 (m, 1 H, H-5III), 3.63–3.76 (m, 4 H, H-6aII,
H-6bII, H-6aIV, H-6aI), 3.77–3.87 (m, 5 H, H-3II, H-3III,
H-6aIII, H-6bIII, H-6bI), 3.90–3.93 (m, 1 H, H-5I),
4.02–4.15 (m, 3 H, H-4III, H-6bIV, H-5V), 4.20 (dd, J
3.3, 7.8 Hz, H-4I), 4.34 (d, 1 H, J 7.9 Hz, H-1II), 4.39
(d, 1 H, J 4.5 Hz, H-2I), 4.43 (d, 1 H, J 3.3 Hz, H-3I),
4.44–4.47 (m, 2 H, H-5II, H-6aV), 4.60 (dd, 1 H, J 2.7,
12.0 Hz, H-6bV), 4.83 (d, 1 H, J 8.1 Hz, H-1V), 4.87 (t,
1 H, J 9.9 Hz, H-4III), 4.92–4.97 (m, 3 H, H-1III, H-2II,
H-2III), 5.21 (d, 1 H, J 8.1 Hz, H-1IV), 5.28 (dd, 1 H, J
8.1, 9.3 Hz, H-2IV), 5.41 (dd, 1 H, J 8.1, 9.2 Hz, H-2V),
5.58–5.69 (m, 4 H, J 9.6, 9.3 Hz, H-4IV, PhCH, H-4V,
H-3IV), 5.85–5.88 (m, 2 H, H-1I, H-3V), 7.23–8.06 (m,
45 H, Ph). Anal. Calcd for C102H98O37: C, 63.95; H,
5.16. Found: C, 64.22; H, 5.17.
1
H2O). Selected H NMR (D2O): 5.21 (d, 0.7 H, J 3.2
Hz, H-1I of a isomer), 4.61–4.70 (m, 5.3 H, H-1I of b
isomer and H-1II,III,IV,V,VI). Selected 13C NMR (100
MHz, D2O): 96.35, 103.22, 103.38 (C-1I-VI). ESIMS
Calcd for C36H62O31: 990 [M]. Found 989.2 [M−H]+.
Acknowledgements
This work was supported by NNSF of China
(Projects 29972053, 39970179) and RCEES of CAS.
The authors thank Dr. Hongyan Hu (General Hospital
of AP) and Professor Liqin Wang (Medical School,
Beijing University) for performing the mice tests.
2,3,4,6-Tetra-O-benzoyl-i-
[2,3,4,6-tetra-O-benzoyl-i-
2,4-di-O-acetyl-i-
4,6-O-benzylidene-i-
tetra-O-benzoyl-i- -gluco pyranosyl-(1“6)]-1,2-O-iso-
propylidene-h- -glucofuranose (22).—Compounds 21
D
-glucopyranosyl-(1“3)-
D
-glucopyranosyl-(1“6)]-
D
-glucopyranosyl-(1“3)-2-O-acetyl-
-glucopyranosyl-(1“3)-[2,3,4,6-
D
D
D
(360 mg, 0.188 mmol) and 9 (148 mg, 0.2 mmol) were
coupled as described in the preparation of 5 (Method
B) to give 22 (430 mg, 87%) as a white solid: [h]D −12°
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