Synthesis and biological activities of octyl 2,3,4-tri-O-sulfo-α-L- fucopyranosyl-(1 → 3)-2,4-di-O-sulfo-α-L-fucopyranosyl-(1 → 3)-2,4-di-O-sulfo-α-L-fucopyranosyl-(1 → 3)-2,4-di-O-sulfo-β-L- fucopyranoside

Article abstract of DOI:10.1016/j.carres.2003.12.014

An efficient method for the regioselective 3-O-silylation of β-thiofucopyranoside was disclosed. Based on this discovery, we described a high-yielding strategy for the synthesis of the natural core structure of L-fucan and its fully sulfated derivative. The bioassay suggested that octyl 2,3,4-tri-O-sulfo-α-L-fucopyranosyl-(1→3)-2,4-di-O-sulfo-α-L- fucopyranosyl-(1→3)-2,4-di-O-sulfo-α-L-fucopyranosyl-(1→3)-2, 4-di-O-sulfo-β-L-fucopyranoside presented better antitumor activities than that of the free tetramer based on Sarcoma 180 cells and Lewis lung carcinoma model studies.

Full text of DOI:10.1016/j.carres.2003.12.014

Carbohydrate
RESEARCH
Carbohydrate Research 339 (2004) 867–872
Synthesis and biological activities of octyl 2,3,4-tri-O-sulfo-a-
fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-a- -fucopyranosyl-(1 fi 3)-
2,4-di-O-sulfo-a- -fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-
b- -fucopyranoside
L
-
L
L
L
Yuxia Hua, Guofeng Gu and Yuguo Du*
Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, PO Box 2871, Beijing 100085, PR China
Received 20 November 2003; accepted 20 December 2003
Abstract—An efficient method for the regioselective 3-O-silylation of b-thiofucopyranoside was disclosed. Based on this discovery,
we described a high-yielding strategy for the synthesis of the natural core structure of
bioassay suggested that octyl 2,3,4-tri-O-sulfo-a- -fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-a-
a- -fucopyranoside presented better antitumor activities than that of the free tetramer
L
-fucan and its fully sulfated derivative. The
L
L
-fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-
L
-fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-b-
L
based on Sarcoma 180 cells and Lewis lung carcinoma model studies.
Ó 2004 Elsevier Ltd. All rights reserved.
Keywords: Regioselective silylation; Fucan; Sulfated oligosaccharide; Antitumor activity; Glycosylation
1. Introduction
by specific patterns of sulfation.7 A comprehensive
review article with respect to structures, functions, and
biological properties of sulfated fucans has been pub-
lished very recently.7b Curious about the bioactivities of
sulfated fucan fragments, we launched a project on the
synthesis of a series of sulfated fucoidan oligosaccha-
rides. Here we would like to report the synthesis and
Sulfated fucans are among the most widely studied of all
the sulfated polysaccharides of nonmammalian origin
that exhibit biological activities in mammalian systems.
Such compounds have been isolated from the cell walls
of marine brown algae,1 the jelly coat from sea urchin
eggs,2 and the sea cucumber body wall.3 These polyan-
ionic molecules show anticoagulant activity and are
potent activators of both antithrombin III and heparin
cofactor II.4 They are inhibitors of retroviral infection,
blocking the infection of human cell lines with, for
example, HIV, herpes, and cytomegalovirus.5 They also
can act as anti-angiogenic agents and can block selectin-
mediated cell–cell binding.6 The structural components
of sulfated fucans necessary for all these biological
activities have not yet been determined. However, most
of them have a common and simple linear backbone,
antitumor activities of octyl 2,3,4-tri-O-sulfo-a-
pyranosyl-(1 fi 3)-2,4-di-O-sulfo-a- -fucopyranosyl-
(1 fi 3)-2,4-di-O-sulfo-a- -fucopyranosyl-(1 fi 3)-2,4-di-
O-sulfo-a- -fucopyranoside.
L-fuco-
L
L
L
2. Results and discussion
(1 fi 3)-a-
Dejter-Juszynski and Flowers using the Koenigs–Knorr
reaction.8 More recently, a (1 fi 3)-a-
-fucopyranosyl
L-Fucobioside was previously synthesized by
L
trisaccharide derivative was prepared in a moderate yield
by reiterative glycosylation using 3,4-di-O-benzoyl-2-O-
benzylfucosyl bromide and regioselective 3-O-fucosyl-
ation in the presence of Hg(CN)2 and HgBr2.9a A simi-
lar strategy was also applied to the preparation of
that of a (1 fi 3)-linked a-
L-fucopyranoside that differs
* Corresponding author. Tel.: +86-10-62914475; fax: +86-10-629235-
63; e-mail: duyuguo@mail.rcees.ac.cn
0008-6215/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2003.12.014

868
Y. Hua et al. / Carbohydrate Research 339 (2004) 867–872
(1 fi 2)-linked fucopyranosyl oligomers.9b It is obvious
that a latent 2,4-O-benzylated fucosyl donor, such as
thioglycoside 6, would significantly simplify the synthesis
5.13 ppm, respectively, from around 3.60 ppm in com-
pound 2. It is worth noting that ethyl 1-thio-a- -fuco-
pyranoside (4) did not give acceptable yield of 5 under
the same reaction conditions, but a mixture of the 2-O-
silyl and 3-O-silyl derivation in a ratio of 2:3.
L
of (1 fi 3)-a-
prepared. Attempts at regioselective benzylation of alkyl
a- -fucopyranoside afforded 2,4- and 3,4-dibenzyl ethers
in the ratio of 3:2.10 Tin complex-assisted 3-O-allyation
of alkyl a- -fucopyranoside presented a better way to
L-fucose oligomers, if 6 could be easily
L
Encouraged by this basic finding, we envisioned a
practical strategy to synthesize the (1 fi 3)-linked fuco-
idan framework, an octyl fucotetroside 15 and its sul-
fated derivative 16 (Scheme 2). Thus, compound 2 was
benzylated with BnBr and NaHin DMF to give 6 in
97% yield. Compound 6 was condensed with 1-octanol
in CH2Cl2 at )20 °C using N-iodosuccinimide/trimeth-
ylsilyl trifluoromethanesulfonate (NIS/TMSOTf) as
catalysts to give octyl fucoside (7), followed by tetra-
butylammonium fluoride (TBAF)-catalyzed desilylation
affording acceptor 8 in 88% isolated yield for two steps.
1HNMR analysis surprisingly found that this major
fucoside has the b configuration. Doublet (J 7.7 Hz) at d
4.37 ppm (H-1) clearly indicated a b linkage in 8. Gly-
cosylation of 6 and 8 as described in the preparation of 7
L
discriminate between the 3-OHand 2,4-OHs, but it is
toxic and expensive.11 We found that direct silylation of
ethyl 1-thio-b-L-fucopyranoside (1) with tert-butyldi-
methylsilyl chloride (TBSC1) and imidazole in N,N-di-
methylformamide (DMF) at 0 °C gave an excellent yield
of monosilylated 2 (92% isolated yield, Scheme 1). Reg-
ioselective substitution on C-3 of 2 was further confirmed
from its acetylated compound 3, that is, the chemical
shifts of H-2 and H-4 in 3 moved downfield to 5.11 and
1
gave the a linked disaccharide 9 in 89% yield. H–1H
COSY of 9 showed a doublet (J 3.9 Hz) at d 5.04 ppm
confirming the a linkage between the sugar residues.
Reiteration of the desilylation with TBAF and glyco-
sylation with 6 transformed disaccharide 9 into trisac-
charide acceptor 12 in 59% overall yield. Warning
should be given here that trifluoroacetic acid-promoted
desilylation on 9 and 11 caused bond breakage between
the fucose residues.12 Final coupling reaction of 12 and
thioglycosyl donor 1313 furnished the full benzylated
Scheme 1. Reagents and conditions (yields): (a) TBSC1, DMF, Im,
0 °C (92%); (b) Ac2O, Pyr (100%).
Scheme 2. Reagents and conditions (yields): (a) BnBr, NaH, DMF (97%); (b) NIS, TMSOTf, CH2Cl2, )20 °C (97% for 7; 89% for 9; 80% for 11; 80%
for 14); (c) TBAF, THF (89% for 8; 76% for 10; 97% for 12); (d) Pd–C, H2 1:1 MeOH–EtOAc, 85%; (e) SO3ÆPyr, Pyr; 3 N NaOH(92%).

Y. Hua et al. / Carbohydrate Research 339 (2004) 867–872
869
tetrasaccharide 14 (80%). HMQC assigned four ano-
meric protons that appeared at d 4.30 ppm (J 7.4 Hz),
5.07 ppm (J 3.5 Hz), 5.09 ppm (J 3.4 Hz), and 5.10 ppm
(J 3.4 Hz), respectively, supporting the structure of
compound 14. Catalytic hydrogenolysis of 14 with H2
on Pd(OH)2 gave the free tetrasaccharide 15, which was
subsequently sulfated with the sulfur trioxide–pyridine
complex in pyridine at 55 °C for 72 h, giving 16 in an
excellent yield (92%) after Sephadex LH20 purification
and lyophilization of the eluate.
The antitumor activities of the free fucotetroside 15
and the corresponding sulfated derivative 16 were pre-
liminarily tested in vivo according to the method
described by Sasaki and Takasuka.14 Kun-min mice
weighing about 20 g and Lewis lung carcinoma cells
(1.5 · 106) or Sarcoma-180 cells (5 · 106) were used for
the bioassay. Cisplatinꢀ (CDDP) was selected as the
positive control in parallel tests. The mice were injected
with compound 15 or 16 (daily dose, 2 mg/kg each; iv)
from day 3 to 14 after inoculation with the tumor cells,
while CDDP was given every other day.
spectra were recorded with ARX 400 spectrometers for
solutions in CDCl3 or D2O. Chemical shifts are given in
ppm downfield from internal Me4Si or TSP. Mass
spectra were measured using MALDITOF or ESIMS
with dihydroxybenzoic acid (DHB) as the matrix. Thin-
layer chromatography (TLC) was performed on silica
gel HF254 with detection by charring with 30% (v/v)
H2SO4 in MeOHor in some cases by a UV detector.
3.2. Ethyl 3-O-tert-butyldimethylsilyl-1-thio-b-L-fucopyr-
anoside (2)
To a solution of compound 1 (1.9 g, 9.13 mmol) in N,N-
dimethylformamide (DMF, 15 mL) was added imidaz-
ole (1.595 g, 23.42 mmol) and tert-butylchlorodimethyl-
silane (1.775 g, 11.67 mmol) at 0 °C. The mixture was
stirred under these conditions for 30 min and then at
room temperature for another 2 h. The reaction mixture
was diluted with cold water (50 mL) and extracted with
EtOAc (3 · 50 mL). The organic phase was dried over
anhydrous MgSO4 and concentrated. Purification of the
Control mice were injected with saline alone. The
tumor volumes were estimated according to the fol-
lowing formula: tumor volume ðcm3Þ ¼ 4pðxyzÞ=3,
where x, y, and z are the three perpendicular diameters
of the tumor. Inhibition ratio was thus calculated based
on tumor volume: ðVcontrol À VtestÞ=Vcontrol, where Vcontrol is
the average tumor volume of saline group, Vtest is the
average tumor volume for the evaluating group. The
tumor inhibition ratios for 15, 16, and CDDP on Sar-
coma-180 cells were 20%, 50%, and 68%, while on Lewis
lung carcinoma they were 16%, 48%, and 60%, respec-
tively. It was thus deduced that sulfated fucose oligo-
mers might enhance the antitumor activities.15 Different
structure of fucoidans with specific sulfation patterns are
currently under construction, and the related synthesis
and bioactivities will be reported in due course.
residue on a silica gel column (7:1 petroleum ether–
25
D
EtOAc) gave 2 (2.709 g, 92%) as a syrup; ½aŠ +11° (c 1,
1
CHCl3); HNMR (400 MHz, CDCl 3): d 4.24 (d, 1H, J
9.6 Hz, H-1), 3.55–3.66 (m, 4H, H-2, H-3, H-4, H-5),
2.70–2.76 (m, 2H, SCH2), 1.37 (d, 3H, J 6.4 Hz, H-6),
1.30 (t, 3H, J 7.6 Hz, SCH2CH3), 0.92 (s, 9H,
SiC(CH3)3), 0.16, 0.13 (2s, 2 · 3H, Si(CH3)2). Anal.
Calcd for C14H30O4SSi: C, 52.13; H, 9.38. Found: C,
51.91; H, 9.45.
3.3. Ethyl 2,4-di-O-acetyl-3-O-tert-butyldimethylsilyl-1-
thio-b-L-fucopyranoside (3)
Compound 2 (52 mg, 0.16 mmol) was acetylated with
Ac2O (0.5 mL) in pyridine (1 mL) giving 3 as a syrup
25
D
quantitatively; ½aŠ +36° (c 2, CHCl3); 1HNMR
In conclusion, we have disclosed a simple method for
the efficient regioselective 3-O-silylation of b-thiofuco-
side. Based on this discovery, we have described a high-
yielding strategy for the synthesis of the core structure of
(400 MHz, CDCl3): d 5.13 (d, 1H, J 3.2 Hz, H-4), 5.11 (t,
1H, J 9.8 Hz, H-2), 4.36 (d, 1H, J 9.8 Hz, H-l), 3.82 (dd,
1H, J 3.2, 9.8 Hz, H-3), 3.73 (q, 1H, J 6.6 Hz, H-5),
2.68–2.77 (m, 2H, SCH2), 2.15, 2.08 (2s, 6H, 2CH3CO),
1.26 (t, 3H, J 7.6 Hz, SCH2CH3), 1.20 (d, 3H, J 6.6 Hz,
H-6), 0.82 (s, 9H, SiC(CH3)3), 0.07, 0.05 (2s, 2 · 3H,
Si(CH3)2). MALDITOF-MS: calcd for C18H34O6SSi,
406.18; found, 429 (M+Na)þ.
natural
L-fucan and its fully sulfated derivative. The
antitumor activities were preliminarily tested in vivo.
The results suggested that sulfated analogue 16 pre-
sented higher tumor growth inhibition ratios based on
studies using Sarcoma-180 cells and Lewis lung carci-
noma model studies.
3.4. Ethyl 2,4-di-O-benzyl-3-O-tert-butyldimethylsilyl-1-
thio-b-L-fucopyranoside (6)
3. Experimental
3.1. General
To a solution of compound 2 (1.114 g, 3,46 mmol) in
DMF (15 mL) at 0 °C was added NaH(50% content,
1.040 g, 21.67 mmol) and BnBr (0.90 mL, 7.55 mmol),
respectively. The mixture was stirred at 0 °C for 30 min,
then at room temperature for 2 h. It was then poured
into ice-cold water (50 mL) and extracted with EtOAc
(3 · 50 mL). The organic phase was dried over
Optical rotations were determined at 25 °C with a Per-
kin–Elmer Model 241-Mc automatic polarimeter at
1
1
k ¼ 254 nm. H, 13C, H–1HCOSY, and HMQC NMR

870
Y. Hua et al. / Carbohydrate Research 339 (2004) 867–872
anhydrous MgSO4 and concentrated. Purification of the
residue by silica gel column chromatography (1:1
petroleum ether–EtOAc) gave 6 (1.678 g, 97%) as a
3H, J 7.0 Hz, CH3). Anal. Calcd for C28H40O5 for
compound 8: C, 73.65; H, 8.83. Found: C, 73.32; H,
8.89.
25
syrup: ½aŠ +10° (c 6, CHCl3); 1HNMR (400 MHz,
D
CDCl3): d 7.47–7.28 (m, 10H, Ph), 5.12, 4.78 (2d, 2H, J
11.5 Hz, PhCH2), 4.91, 4.66 (2d, 2H, J 10.4 Hz, PhCH2),
4.42 (d, 1H, J 9.3 Hz, H-1), 3.78 (dd, 1H, J 2.8, 9.3 Hz,
H-3), 3.70 (t, 1H, J 9.3 Hz, H-2), 3.57 (dq, 1H J 0.5,
6.4 Hz, H-5), 3.49 (dd, 1H, J 0.5, 2.8 Hz, H-4), 2.82–2.69
(m, 2H, SCH2), 1.32 (t, 3H, J 7.4 Hz, SCH2CH3), 1.23
(d, 3H, J 6.4 Hz, H-6), 1.00 (s, 9H, SiC(CH3)3), 0.17,
0.11 (2s, 2 · 3H, Si(CH3)2). Anal. Calcd for
C28H42O4SSi: C, 66.89; H, 8.42. Found: C, 67.13; H,
8.36.
3.7. Octyl 2,4-di-O-benzyl-3-O-tert-butyldimethylsilyl-
L-fucopyr-
anoside (9)
a-L
-fucopyranosyl-(1 fi 3)-2,4-di-O-benzyl-b-
To a solution of compound 6 (549 mg, 1.09 mmol) and
compound 8 (468 mg, 1.03 mmol) in dry CH2Cl2 (3 mL)
ꢀ
was added 4 A molecular sieves (1 g) at )20 °C under an
N2 atmosphere. After 20 min, NIS (270 mg, 1.10 mmol)
and TMSOTf (15 lL, 0.08 mmol) were added to the
above stirred mixture. The reaction mixture was stirred
under these conditions for 60 min, quenched by Et3N
(two drops) and concentrated. The residue was purified
by silica gel column chromatography (20:1 petroleum
3.5. Octyl 2,4-di-O-benzyl-3-O-tert-butyldimethylsilyl-
b-L-fucopyranoside (7)
ether–EtOAc) to give 9 (819 mg, 89%) as a white foam:
25
D
½aŠ )103° (c 0.3, CHCl3); 1HNMR (400 MHz, CDCl 3):
d 7.36–7.18 (m, 20H, Ph), 5.08, 5.02 (2d, 2H, J 11.2 Hz,
PhCH2), 5.04 (d, 1H, J 3.6 Hz, H-10), 4.95, 4.76, 4.65,
4,60, 4.55, 4.46 (6d, 6H, J 10.7 Hz, PhCH2), 4.28 (d, 1H,
J 7.5 Hz, H-1), 4.23 (dd, 1H, J 2.8, 9.9 Hz, H-30), 4.17 (q,
1H, J 6.6 Hz, H-50), 3.94 (dd, 1H, J 3.3, 9.9 Hz, H-20),
3.91–3.87 (m, 1H, OCH2), 3.71 (dd, 1H, J 7.5, 10.0 Hz,
H-2), 3.64 (dd, 1H, J 2.6, 10.0 Hz, H-3), 3.50 (d, 1H, J
2.6 Hz, H-4), 3.45–3.42 (m, 1H, OCH2), 3.38 (q, 1H, J
6.8 Hz, H-5), 3.30 (br d, 1H, J 2.8 Hz, H-40), 1.39–1.23
(m, 15H, CH2 and H-60), 0.94 (s, 9H, SiC(CH3)3), 0.87
(d, 3H, J 6.6 Hz, H-6), 0.85 (t, 3H, J 7.0 Hz, CH2CH3),
0.13, 0.12 (2s, 2 · 3H, Si(CH3)2). Anal. Calcd
for C54H76O9Si: C, 72.28; H, 8.54. Found: C, 72.66; H,
8.48.
To a solution of compound 6 (512 mg, 1.02 mmol) and
1-octanol (194 lL, 1.22 mmol) in dry CH2Cl2 (3 mL) was
ꢀ
added 4 A molecule sieves (1 g) at )20 °C under a N2
atmosphere. The mixture was stirred under these con-
ditions for 20 min, then NIS (370 mg, 1.51 mmol) and
TMSOTf (19.4 lL, 0.11 mmol) were added. The reac-
tion was monitored by TLC until the starting materials
disappeared (ca. 60 min), then the mixture was
neutralized with Et3N (two drops) and concentrated.
The residue was purified by silica gel column chroma-
tography (1:1 petroleum ether–EtOAc) to give (569 mg,
97%) as a white foam. This compound was directly
used for the next reaction without further characteriza-
tion.
3.8. Octyl 2,4-di-O-benzyl-a-
L
-fucopyranosyl-(1 fi 3)-
2,4-di-O-benzyl-b- -fucopyranoside (10)
L
3.6. Octyl 2,4-di-O-benzyl-b-L-fucopyranoside (8)
To a solution of compound 9 (800 mg, 0.89 mmol) in
THF (5 mL) was added TBAF (890 mg, 0.28 mmol) at
room temperature. The mixture was stirred at room
temperature until TLC indicated that the reaction was
complete. Concentration and purification of the residue
by silica gel column chromatography (7:1 petroleum
ether–EtOAc) gave 10 (533 mg, 76%) as a white foam:
To a solution of compound 7 (468 mg, 0.82 mmol) in
THF (15 mL) was added tetrabutylammonium fluoride
monohydrate (TBAF, 1.327 g, 4.21 mmol) at room
temperature. The mixture was stirred under these con-
ditions for 2 h, then concentrated and purified by silica
gel column chromatography (1:1 petroleum ether–
EtOAc) to give 8 (334 mg, 89%) as a white foam. To
confirm the structure, compound 8 (30 mg, 0.066 mmol)
was acetylated with Ac2O (0.1 mL) in pyridine (1 mL)
25
1
½aŠ )94° (c 1.3, CHCl3); HNMR (400 MHz, CDCl 3):
D
d 7.35–7.23 (m, 20H, Ph), 5.18 (d, 1H, J 3.3 Hz, H-10),
5.05, 4.52 (2d, 2H, J 11.6 Hz, PhCH2), 4.95, 4.58 (2d,
2H, J 10.4 Hz, PhCH2), 4.71–4.60 (m, 4H, PhCH2), 4.29
(d, 1H, J 7.0 Hz, H-1), 4.17 (q, 1H, J 6.5 Hz, H-50), 4.02
(ddd, 1H, J 10.0, 3.0, 6.1 Hz, H-30), 3.93–3.80 (m, 1H,
OCH2), 3.80 (dd, 1H, J 10.1, 3.4 Hz, H-3), 3.75 (dd, 1H,
J 7.0, 10.1 Hz, H-2), 3.69 (dd, 1H, J 10.0, 3.3 Hz, H-20),
3.56 (d, 1H, J 3.4 Hz, H-4), 3.44–3.39 (m, 2H, one
proton of OCH2 and H-5), 3.29 (d, 1H, J 3.0 Hz, H-40),
2.02 (d, 1H, J 6.1 Hz, OH), 1.65–1.55 (m, 2H, CH2),
1.40–1.20 (m, 10H, CH2), 1.14 (d, 3H, J 6.3 Hz, H-6),
affording octyl 3-O-acetyl-2,4-di-O-benzyl-b-L-fucoside:
25
D
½aŠ )40° (c, 0.4, CHCl3); 1HNMR (400 MHz, CDCl 3):
d 7.26–7.35 (m, 10H, Ph), 4.88, 4.65, 4.62, 4.58 (4d, 4H,
J 11.8 Hz, PhCH2), 4.86 (dd, 1H, J 3.2, 10.2 Hz, H-3),
4.37 (d, 1H, J 7.7 Hz, H-1), 3.97–3.91 (m, 1H, OCH2),
3,74 (dd, 1H, J 7.7, 10.2 Hz, H-2), 3.65 (d, 1H, J 3.0 Hz,
H-4), 3.60 (q, 1H, J 6.4 Hz, H-5), 3.51–3.44 (m, 1H,
OCH2), 1.92 (s, 3H, COCH3), 1.68–1.58 (m, 2H), 1.40–
1.22 (m, 10H, CH2), 1.21 (d, 3H, J 6.4 Hz, H-6), 0.87 (t,

Y. Hua et al. / Carbohydrate Research 339 (2004) 867–872
871
0.90 (d, 3H, J 6.5 Hz, H-60), 0.85 (t, 3H, J 7.0 Hz,
CH2CH3). Anal. Calcd for C48H62O9: C, 73.63; H, 7.98.
Found: C, 73.29; H, 8.06.
3.11. Octyl 2,3,4-tri-O-benzyl-a-
2,4-di-O-benzyl-a-
benzyl-a-
pyranoside (14)
L
-fucopyranosyl-(1 fi 3)-
L
-fucopyranosyl-(1 fi 3)-2,4-di-O-
-fucopyranosyl-(1 fi 3)-2,4-di-O-benzyl-b-
L
L-fuco-
3.9. Octyl 2,4-di-O-benzyl-3-O-tert-butyldimethylsilyl-a-
L
-fucopyranosyl-(1fi 3)-2,4-di-O-benzyl-a-
L
-fucopyranos-
Glycosylation of compounds 13 (139 mg, 0.29 mmol)
and 12 (270 mg, 0.24 mmol) as described in the prepa-
yl-(1 fi 3)-2,4-di-O-benzyl-b- -fucopyranoside (11)
L
25
D
ration of 9 gave 14 (295 mg, 80%) as a white foam: ½aŠ
1
Compounds 6 (288 mg, 0.57 mmol) and 10 (393 mg,
0.50 mmol) were glycosylated as described in the prep-
)90° (c 0.8, CHCl3); HNMR (400 MHz, CDCl 3): d
7.35–7.10 (m, 45H, Ph), 5.12 (d, 1H, J 3.4 Hz, H-100),
5.10 (d, 1H, J 3.4 Hz, H-10), 5.09 (d 1H, J 11.4 Hz,
PhCH2), 5.07 (d, 1H, J 3.2 Hz, H-1000), 5.05 (d, 1H, J
11.4 Hz, PhCH2), 4.92, 4.90, 4.80 (3d, 3H, J 11.6 Hz,
PhCH2), 4.70–4.64 (m, 5H, PhCH2), 4.62, 4.60, 4.58,
4.56, 4.54, 4.40, 4.37 (7d, 7H, PhCH2), 4.31–4.26 (m,
3H, H-1, H-5, H-50), 4.200 (q, 1H, J 6.3 Hz, H-5000), 4.15–
aration of compound 9 to give 11 (488 mg, 80%) as a
25
D
white foam: ½aŠ )76° (c 0.8, CHCl3); 1HNMR
(400 MHz, CDCl3): d 7.35–7.16 (m, 30H, Ph), 5.10, 5.02
(2d, 2H, J 11.6 Hz, PhCH2), 5,08 (d, 1H, J 3.3 Hz, H-
100), 5.06 (d, 1H, J 3.4 Hz, H-10), 5.03, 4.92, 4.79, 4.71,
4.69, 4.64, 4,59, 4.58, 4.49, 4.33 (10d, 10H, J 10.9 Hz,
PhCH2), 4.32–4.23 (m, 4H, H-1, H-50, H -5, H -3), 4.07
(dd, 1H, J 3.3, 10.4 Hz, H-200), 4.02 (dd, 1H, J 3.4,
10.0 Hz, H-20), 3.98 (q, 1H, J 6.4 Hz, H-5), 3.95–3.87 (m,
1H, OCH2), 3.72–3.65 (m, 2H, H-3, H-30), 3.51 (br s,
1H, H-4), 3.46–3.39 (m, 2H, H-2 and one proton of
OCH2), 3.38 (br s, 1H, H-40), 3.31 (br s, 1H, H-400), 1.65–
1.58 (m, 2H, CH2), 1.40–1.19 (m, 10H, CH2), 1.17 (d,
3H, J 6.4 Hz, H-6), 1.03 (d, 3H, J 6.5 Hz, H-60), 0.92 (s,
9H, SiC(CH3)3), 0.85 (t, 3H, J 7.0 Hz, CH3), 0.80 (d, 3H,
J 6.4 Hz, H-600), 0.10, 0.08 (2s, 2 · 3H, Si(CH3)2). Anal.
Calcd for C74H98O13Si: C, 72.63; H, 8.07. Found: C,
72.97; H, 8.11.
4.09 (m, 3H, H-2000, H -3, H -3), 4.04 (dd, 1H, J 3.5,
00
00
00
10.2 Hz, H-200), 3.95 (q, 1H, J 6.5 Hz, H-500), 3.94–3.85
(m, 2H, H-200, one proton of OCH2), 3.71–3.65 (m, 2H,
H-3, H-3000), 3.53 (br s, 1H, H-4000), 3.47–3.40 (m, 2H, one
proton of OCH2, H-2), 3.35–3.30 (m, 3H, H-4, H-40, H -
400), 1.64–1.56 (m, 2H), 1.40–1.20 (m, 10H, CH2), 1.18
(d, 3H, J 6.4 Hz, H-6), 0.96 (d, 3H, J 7.6 Hz, H-60), 0.90–
00
0.82 (m, 6H, CH2CH3, H -6), 0.80 (d, 3H, J 6.4 Hz,
H-6000); 13C NMR (100 MHz, CDCl3): d 139.02, 138.92,
138.90, 138.82, 138.65, 138.56, 138.31, 138.13, 138.07,
103.98 (C-1), 96.23 (C-1000), 95.02 (C-10), 94.87 (C-100),
79.47, 78.67, 78.43, 77.32, 77.01, 76.50, 76.30, 76.00,
75.62, 75.53, 75.18, 74.86, 74.78 (3 C), 74.39, 74.30,
74.18 (2 C), 73.94, 72.46, 70.12 (C-5000), 69.91, 66.80 (C-
500), 66.50 (C-5), 66.23 (C-50), 31.79, 29.73, 29.40, 29.20,
26.14, 22.66, 16.78 (C-6), 16.48 (C-60), 16.43 (C-6000),
16.25 (C-600), 14.10. Anal. Calcd for C95H112O17: C,
74.78; H, 7.40. Found: C, 75.16; H, 7.46.
3.10. Octyl 2,4-di-O-benzyl-a-
2,4-di-O-benzyl-a- -fucopyranosyl-(1 fi 3)-2,4-di-O-
benzyl-b- -fucopyranoside (12)
L
-fucopyranosyl-(1 fi 3)-
L
L
To a solution of compound 11 (308 mg, 0.25 mmol) in
THF (2 mL) was added TBAF (890 mg, 0.28 mmol) at
room temperature. The mixture was stirred at room
temperature until TLC indicated that the reaction was
complete. Concentration and purification of the residue
by silica gel column chromatography (7:1 petroleum
ether–EtOAc) gave 12 (270 mg, 97%) as a white foam:
3.12. Octyl a-
L
-fucopyranosyl-(1 fi 3)-a-
L-fucopyranos-
yl-(1 fi 3)-a-
(15)
L
-fucopyranosyl-(1 fi 3)-b- -fucopyranoside
L
25
1
½aŠ )19° (c 1, CHCl3); HNMR (400 MHz, CDCl 3: d
To a solution of compound 14 (212 mg, 0.14 mmol) in
1:1 MeOH–EtOAc (20 mL) was added 20% Pd(OH)2 on
charcoal (209 mg, 0.14 mmol). The mixture was bubbled
with H2 under the flow rate of 100 mL/min for 70 h. The
D
7.36–7.17 (m, 30H, Ph), 5.19 (d, 1H, J 3.4 Hz, H-100),
5.12 (d, 1H, J 3.2 Hz, H-10), 5.10, 5.02, 4.94 (3d, 3H, J
11.3 Hz, PhCH2), 4.73–4.64 (m, 6H, PhCH2), 4.57, 4.51,
4.39 (3d, 3H, J 11.3 Hz, PhCH2), 4.31–4.25 (m, 3H, H-1,
reaction mixture was filtered, and the filtrate was con-
centrated to give 15 (84 mg, 85%) as a white foam; ½aŠ
25
D
H-50, H -5), 4.08 (dd, 1H, J 3.4, 10.5 Hz, H-200), 4.04
00
1
(ddd, 1H, J 3.0, 10.5, 6.3 Hz, H-300), 4.02 (q, 1H, J
6.4 Hz, H-5), 3.95–3.89 (m, 1H, OCH2), 3.84 (dd, 1H, J
3.2, 10.3 Hz, H-20), 3.75–3.68 (m, 2H, H-3, H-30), 3.54
(br s, 1H, H-4), 3.46–3.41 (m, 2H, H-2 and one proton
of OCH2), 3.33 (br s, 1H, H-40), 3.31 (br d, 1H, J 3.0 Hz,
H-400), 2.02 (d, J 6.3 Hz, OH), 1.65–1.56 (m, 2H, CH2),
1.40–1.20 (m, 10H, CH2), 1.16 (d, 3H, J 6.4 Hz, H-6),
1.03 (d, 3H, J 6.7 Hz, H-600), 0.87 (t, 3H, J 7.1 Hz,
CH2CH3), 0.80 (d, 3H, J 6.4 Hz, H-60). Anal. Calcd for
C68H84O13: C, 73.62; H, 7.63. Found: C, 73.39; H, 7.57.
+5° (c 0.4, H2O); HNMR (400 MHz, CDCl 3): d 5.00
(br s, 3H, 00H0 -10, H -1, H -1 ), 4.25 (br s, 4H, H-5, H-50,
00
000
00
000
H-500, H -5 ), 3.94–3.85 (m, 8H, H-4, H-40, H -4, H -4 ,
H-1, H-3000, H-2, OCH2), 3.26 (br s, 3H, H-3, H-30, H -
300), 3.57–3.25 (m, 4H, H-20, H -2, H -2 , OCH2), 1.55
(br s, 2H), 1.20–1.13 (m, 22H), 0.79 (br s, 3H); 13C NMR
(100 MHz, D2O): d 102.85 (C-1), 95.70 (C-100, C-1000),
95.44 (C-10), 77.80 (C-3), 74.97 (C-30, C-300), 71.91 (C-
4000), 70.45 (C-3000, OCH2), 69.46 (C-4), 68.74 (C-2000),
68.52 (C-20, C-40, C-400), 68.03 (C-200), 67.62 (C-5000), 66.95
00
000

872
Y. Hua et al. / Carbohydrate Research 339 (2004) 867–872
(C-5), 66.47 (C-2, C-50, C-500), 31.50, 29.06, 28.93 (2C),
25.43, 22.34, 15.84, 15.59, 15.48, 15.33 (4 C-6), 13.70;
ESIMS (negative-ion): calcd for C32H58O17, 714.37;
found, 713.3 (M)H)þ.
References
1. Percival, E.; McDowell, R. H. Chemistry and Enzymology
of Marine Algal Polysaccharides; Academic: New York,
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~
2. Alves, A.-P.; Mulloy, B.; Diniz, J. A.; Mourao, P. A. S.
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~
3. Pereira, M. S.; Mulloy, B.; Mourao, P. A. S. J. Biol. Chem.
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3.13. Octyl 2,3,4-tri-O-sulfo-a-
2,4-di-O-sulfo-a- -fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-
a- -fucopyranosyl-(1 fi 3)-2,4-di-O-sulfo-b- -fucopyr-
anoside (16)
L
-fucopyranosyl-(1 fi 3)-
L
L
L
~
E. S.; Valente, A.-P.; Mourao, P. A. S. Carbohydr. Res.
2002, 337, 2231–2238.
To a solution of compound 15 (83 mg, 0.12 mmol) in
pyridine (4 mL) was added sulfur trioxide–pyridine
complex (530 mg, 3.26 mmol). The mixture was stirred at
55 °C for 72 h, then cooled to 4 °C to generate a pre-
cipitate. The pyridine phase was sucked off, and the
precipitate was dissolved in water (1 mL). Sodium
hydroxide (3 N) was added until pH10. The water phase
was washed with CH2Cl2 (2 · 1 mL), then loaded on to a
Sephadex LH20 column and eluted with water. The
5. (a) McClure, M. O.; Moore, J. P.; Blanc, D. F.; Scotting,
P.; Cook, G. M.; Keynes, R. J.; Weber, J. N.; Davies, D.;
Weiss, R. A. AIDS Res. Human Retrovir. 1992, 8, 19–26;
(b) Yoshida, T.; Hatanaka, K.; Uryu, T.; Kaneko, Y.;
Suzuki, E.; Miyano, H.; Mimura, T.; Yoshida, O.;
Yamamoto, N. Macromolecules 1990, 23, 3717–3722.
6. (a) Hahnenberger, R.; Jakobson, A. M. Glycoconjugate J.
1991, 8, 350–353; (b) Khoo, K.-H.; Sarda, S.; Xu, X.;
Caulfield, J. P.; McNeil, M. R.; Homans, S. W.; Morris,
H. R.; Dell, A. J. Biol. Chem. 1995, 270, 17114–17123.
7. (a) Mulloy, B.; Ribeiro, A.-C.; Alves, A.-P.; Vieira, R. P.;
desired fractions were combined and freeze dried to give
25
16 (174 mg, 92%) as an amorphous white solid; ½aŠ
~
Mourao, P. A. S. J. Biol. Chem. 1994, 269, 22113–22123;
D
1
(b) Berteau, O.; Mulloy, B. Glycobiology 2003, 13, 29R–
40R.
)75° (c 0.5, H2O); HNMR (400 MHz, CDCl 3): d 5.30
(br s, 3H, H-10, 100, 1000), 4.90–4.85 (m, 5H, H-3000, H-4,40,
400, 4000), 4.76 (d, 1H, J 6.7 Hz, H-1), 4.56–4.45 (m, 8H, H-
2, 20, 200, 2000, H-5, 50, 500, 5000), 4.30–4.18 (m, 3H, H-3, 30,
300), 4.05–3.91 (m, 1H, OCH2), 3.80–3.70 (m, 1H,
OCH2), 1.54–1.49 (m, 2H), 1.30–1.10 (m, 22H, H-6 and
CH2), 0.79 (t, 3H, J 7.2 Hz, CH3); Selected 13C NMR
(100 MHz, D2O): d 103.80, 99.71 (2C), 97.54 (4C-1).
Anal. Calcd for C32H49Na9O44S9Æ3H2O: C, 22.76; H,
3.26; S, 17.07. Found: C, 22.39; H, 3.32; S, 17.18.
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Nifantiev, N. E. J. Carbohydr. Chem. 2003, 22, 109–122,
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This work was supported by NNSF of China (Project
20372081, 30330690) and RCEES of CAS. The authors
thank Professor Huimin Lu (Division of Toxicology,
CDC of China) for performing the mice tests.