Synthesis of a set of di- and tri-sulfated galabioses

Article abstract of DOI:10.1016/S0008-6215(01)00222-1

Among cell-adhesion molecules, L-selectin recognizes sulfated sLe^x with relatively low affinity. Here, we aimed at artificial mimics by synthesizing a set of di- and tri-sulfated galabioses, which may surpass the affinity of sulfated sLe^x. As a strategy to obtain 3′,6′,6-tri-O-sulfogalabioses, regioselective reductive cleavage of 4,6- and 4′,6′-di-O-benzylidenegalabioses was employed. Two suitably protected galactose precursors were conjugated to yield α and β anomers (48 and 18%, respectively) by using a pentenyl galactoside donor and iodinium di-sym-collidine perchlorate as the catalyst. For synthesizing the 3′,6-di-O-sulfogalabiose, however, a trichloroacetimidate donor was superior (52%) to the pentenyl one (30%).

Full text of DOI:10.1016/S0008-6215(01)00222-1

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Carbohydrate Research 335 (2001) 167–180
Synthesis of a set of di- and tri-sulfated galabioses
Tomoaki Yoshida,a,* Taku Chiba,b Takashi Yokochi,a Kikuo Onozaki,b
Tsuyoshi Sugiyama,a Izumi Nakashimac
aDepartment of Microbiology and Immunology, Aichi Medical Uni6ersity, Aichi 480-1195, Japan
bFaculty of Pharmaceutical Sciences, Nagoya City Uni6ersity, Nagoya 467-8603, Japan
cDepartment of Immunology, Nagoya Uni6ersity School of Medicine, 65 Tsurumai-cho, Showa-ku,
Nagoya 466, Japan
Received 8 January 2001; accepted 25 July 2001
Abstract
Among cell-adhesion molecules,
L
-selectin recognizes sulfated sLex with relatively low affinity. Here, we aimed at
artificial mimics by synthesizing a set of di- and tri-sulfated galabioses, which may surpass the affinity of sulfated
sLex. As a strategy to obtain 3%,6%,6-tri-O-sulfogalabioses, regioselective reductive cleavage of 4,6- and 4%,6%-di-O-ben-
zylidenegalabioses was employed. Two suitably protected galactose precursors were conjugated to yield a and b
anomers (48 and 18%, respectively) by using a pentenyl galactoside donor and iodinium di-sym-collidine perchlorate
as the catalyst. For synthesizing the 3%,6-di-O-sulfogalabiose, however, a trichloroacetimidate donor was superior
(52%) to the pentenyl one (30%). © 2001 Elsevier Science Ltd. All rights reserved.
Keywords: Galabiose; a Anomer; b Anomer; Regio-selective sulfation; 3-(2-Aminoethylthio)propyl sugars
1. Introduction
sulfated sLex is reported to bind
-selectin at
L
250 mM of IC50,11 which is relatively weaker
than the intrinsic ligand GlyCAM-1, 108
mM.12 To circumvent such low affinity in
Increasing numbers of biomolecules have
been shown to recognize anionic sugar
molecules. Among them, the selectin family of
cell-adhesion molecules is known to bind sia-
lylated oligosaccharides on the cell surface,1–3
to mediate the primary weak interactions of
lymphocytes, monocytes or granulocytes to
vascular endothelial cells, such that the rolling
phenomenon takes place.4 While the intrinsic
ligands for all three members of the selectin
family share the core structure of sialyl Lex,5
monovalent form, -selectin, whose expression
L
is more concentrated on the pili of the cell
surface,13 apparently exploits the clustered
presence of O-glycosylation sites on the natu-
ral ligands, GlyCAM-114 or CD34.15 Indeed,
previous studies in vitro have shown remark-
able enhancement of the affinity by using
polyvalent oligosaccharide structures.16,17
When the regioselectively monosulfated
galactoses were clustered on polyglutamic
acid, they showed barely detectable affinities
(mM as galactose), whereas the individual
monosaccharide sulfates showed no detectable
affinity (\100 mM).18 Since the overall affini-
ties of the clustered galactose sulfates were far
below the practical value, we next considered
the
L
-selectin requires additional O-sulfation
on the O-6 of the galactose or N-acetylglu-
cosamine residue.6–10 The monovalent 6%-O-
* Corresponding author. Fax: +81-561-639187.
E-mail address: tomo@aichi-med-u.ac.jp (T. Yoshida).
0008-6215/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(01)00222-1

168
T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
2. Results and discussion
sulfated disaccharide structures that possess a
primary amino group on the aglycone for
subsequent formation of clusters. In our expe-
rience with sulfated glucose polymers, the
affinity of the (1“3) linkage was superior to
that of (1“4) or (1“6) linkages.17 The well
Throughout this work, all galactose precur-
sors were suitably protected prior to glycosi-
dation to enable precise regioselective
sulfation to optimize the glycosidation step.
Two different methods were employed accord-
ing to the stability of the donor and the
efficacy of the reaction.
To obtain the 3%,6,6%-tri-O-sulfated com-
pounds 23, 27, a combination of pentenyl
donor, iodinium di-sym-collidine perchlorate
as the catalyst, and 1:4 CH2Cl2–Et2O as the
solvent was optimal in terms of glycosidation
yield, giving the anomeric 2-O-benzyl-4,6-O-
benzylidene-3-O-(2-O-benzyl-4,6-O-benzyli-
known unnatural ligand for
L
-selectin, fu-
coidan, also consists of a-(1“3) linked fu-
cose.19 Thus we constructed a series of a- and
b-(1“3) linked galabioses as possible artificial
mimics, having selective sulfation at the 3%, 6
or 6% positions. The 3%-sulfate here acts as a
substitute for the 3%-sialic acid residue.20 The
rationale for the 6- and 6%-sulfates is based on
the fact that the monosaccharide components
of the natural ligand oligosaccharides consist
of 6-O-sulfated galactose, fucose, sialic acid,
N-acetylglucosamine, and 6-O-sulfated N-
acetylglucosamine.14
dene-3-O-methoxybenzyl-a/b-
D
-galactopyrano
syl)-b- -galactopyranoside (20 (a) 48%; 21 (b)
D
18%).21 To increase the ratio of b anomer, a
switch of the solvent to acetonitrile only de-
creased the efficacy of glycosidation to 24%,
and did not materially affect the anomeric
ratio (20:21=1.5:1).22,23 Use of trichloroace-
timidate as the donor, with the same protect-
ing groups as 4, gave less than 30% of
glycosidation product. Thus, the a/b mixture
obtained by using the pentenyl donor 4 in 1:4
CH2Cl2–Et2O was purified by repeated chro-
matography and crystalization from EtOH.
The position of glycosidation on the acceptor
was confirmed by the downfield shift of C-3
(72.4 to 74.5 (a) or 75.1 (b) ppm).24 In the next
step, regioselective reductive cleavage of two
4,6-O-benzylidene groups by LiAlH4–AlCl3
afforded an acceptable yield of 6,6%-dihydroxy
derivatives (22, 33%; 27, 35%) from 20 and
2125 (Schemes 1–3).
The strategy for 3%,6-dihydroxygalabiose,
the reductive cleavage of 2-O-benzyl-4,6-O-
benzylidene - 3 - O - (2,4,6 - tri - O - benzyl - 3 - O-
methoxybenzyl- a -
D
- galactopyranosyl)- b - -
D
galactopyranoside, obtained by pentenyl gly-
cosidation, gave exclusively 3%,4-dihydroxy
derivative. Thus, avoiding cleavage of the ben-
zylidene group, compound 15 was obtained as
the selectively protected acceptor. When the
pentenyl 2,4,6-tri-O-benzyl-3-O-methoxyben-
zyl-a- -galactopyranosyl donor reacted with
D
15, the glycosidation yield was less than 30%.
The trichloroacetimidate donor having the
same protective groups gave an unsatisfactory
Scheme 1.

T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
169
Scheme 2.
Scheme 3.

170
T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
NMR analyses of the sulfated products
showed noteworthy downfield shifts of the
geminal protons of the 6-sulfate groups and the
carbon atoms connecting to the sulfate groups
(Tables 1–5), indicating that the sulfate groups
had been introduced at the desired positions,
and no desulfation had taken place during the
deprotection step.29–31
result because of its extreme lability. Finally,
3-O-acetyl-2,4,6-tri-O-benzyl-b- -galactopy-
D
ranosyl trichloroacetimidate (19),26 which is
reasonably stable, in reaction with 15 improved
the glycosidation yield to 52%. The downfield
chemical shift of the C-3 acceptor was from
74.2 to 77.7 ppm.24
To access the 3%,6%-dihydroxy compound, 37,
compounds 6 and 11 were coupled in the
presence of iodinium di-sym-collidine perchlo-
rate as described for 20 and 21, followed by
transesterification with NaOMe, to give the
desired product, 37.
The primary spacer arm attached to gal-
abiose was the 2-propenyl group so that it
could be substituted with cysteamine27 after
sulfation at the open hydroxy groups. There
was a possible option of starting with another
spacer that has a latent amino group at the end.
However, when we tried the N-benzyloxycar-
bonylaminohexyl group, the protection of the
O-2 position of the acceptor galactoside, leav-
ing OH-3 open was extremely difficult, pre-
sumably because of steric hindrance.18 The
random glycosidation of the 2,3-dihydroxy ac-
ceptor was unsuccessful because of its poor
solubility in the reaction solvent 1:4 CH2Cl2–
Et2O.
Although cysteamine could be linked at the
double bond of the 2-propenyl group almost
quantitatively with a 10-fold excess under UV
irradiation, the subsequent steps of purification
and deprotection required improvement. Sepa-
ration on a Sephadex LH-20 column, which is
a good alternative to the conventional silica gel
chromatography to avoid loss of sulfated sug-
ars,18 was poor unless 0.5% Et3N was added to
the eluant. This amine, however, was bound
tightly to sulfate esters and resisted removal by
a cation-exchange resin in either the Na+ or
H+ form. Thorough evaporation in the pres-
ence of a 2-fold excess of NaHCO3 was re-
quired to replace the Et3N counter-ion. The
sodium form is required because the Et3N or
pyridine forms substantially destabilize the sul-
fate esters.18 For the final deprotection, the
thioether introduced by cysteamine made hy-
drogenolysis over palladium impossible. We
therefore used trifluoroacetic acid–bro-
motrimethylsilane–thioanisole for final re-
moval of the benzyl protective groups.28
In this way, we constructed anionic gal-
abioses to be tested as artificial ligands for
L
-selectin. The rationale for galabiose was that
the (1“3) linkage, which is most commonly
observed in Gal-Gal or Gal-GalNAc structures
in mammals, showed the highest affinity
among glucose polymers of various configura-
tions.17 Interestingly, the binding affinities of
the trisulfated galabioses, 26 and 30, were 120
mM and 160 mM kD, respectively, toward
L
-selectin in our liquid phase system (data not
shown), which are reasonably high compared
with the affinity of the intrinsic ligand.
3. Experimental
General methods.—Melting points were de-
termined by a Yanagimoto micro melting point
apparatus and uncorrected. Optical rotations
were measured with a Jasco automatic digital
polarimeter, Model DIP-140, in a 0.5 dm tube.
,
All solvents were dried over 4 A molecular
,
sieves except for MeOH (3 A molecular sieves)
and CHCl3 (Na2SO4) before use. Aqueous solu-
tions of H2SO4 and NaHCO3 used for washing
the organic solution were 1 M and saturated,
respectively. H and 13C NMR spectra were
1
obtained with a Jeol Model JNM-GSX400 or
JNM-a 500 spectrometer at 22 °C. The internal
standards for H NMR and 13C NMR mea-
1
surement were Me4Si for CDCl3 and CD3OD,
or acetone (2.200 ppm) for measurements in
D2O. All evaporations were done with a rotary
evaporator. TLC was performed on Silica Gel
60 F254-precoated aluminum sheets (EM Indus-
tries, Inc., Gibbstown, NJ) and compounds
were detected by UV absorbance (254 nm),
ninhydrin reaction or charring (after spraying
with 15% H2SO4 in 50% EtOH). Column chro-
matography was performed on Silica Gel 60
(EM Industries, Inc) or Sephadex LH-20
(Pharmacia Biotech. Inc. Uppsala, Sweden).
All solvent compositions are given as v/v.

T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
171
4-Pentenyl
methoxybenzyl)-i-
4,6-O-benzylidene-3-O-(4-
-galactopyranoside (3).—
D
A mixture of 1 (8.02 g, 32.3 mmol),32
benzaldehyde (17.5 mL, 167 mmol) and for-
mic acid (17.5 mL, 464 mmol) was allowed to
react at 22 °C for 1 h and then neutralized
with aqueous NaHCO3. By the addition of
petroleum ether to the mixture, amorphous
solid 233 was isolated and crystallized from
EtOH.34 The mother liquor was chro-
matographed on a column of silica gel, eluted
with 1:1 toluene–EtOAc. The fractions were
concentrated in vacuo and crystallized from
EtOH to give additional 2. The total yield was
5.85 g (52%); m.p. 160–162 °C; Rf 0.26 (1:4
toluene–EtOAc). Compound 2 (6.12 g, 18.2
mmol) was dissolved in toluene (70 mL) and
azeotropically distilled in the presence of
Bu2SnO (4.75 g, 1.05 fold excess) for 50 min.
Tetrabutylammonium iodide (9.40 g, 1.4 fold
excess) and 4-methoxybenzyl chloride (3.52
mL, 1.4 fold excess) were added and the mix-
ture was stirred at 90 °C overnight, diluted
with CHCl3 and washed with aqueous
NaHCO3. The organic layer was dried with
Na2SO4, concentrated in vacuo and crystal-
lized from EtOH to afford the title compound
3. The mother liquor was chromatographed
on a column of silica gel, eluted with 2:1
hexane–CHCl3 to yield additional 3; total
yield 4.40 g (53%); mp 135.5–136.5 °C; [h]D27
+42.1° (CHCl3); Rf 0.38 (2:1 toluene–
EtOAc). Anal. Calcd for C26H32O7: C, 68.40;
H, 7.06. Found: C, 68.23; H, 7.13.
4-Pentenyl 2-O-benzyl-4,6-O-benzylidene-3-
O-(4-methoxybenzyl)-i- -galactopyranoside
D
(4).—To a stirred solution of 3 (2.67 g, 5.86
mmol) and NaH (60%, 1.4 g) in 26 mL dry
THF, benzyl bromide (4.18 mL, 35.1 mmol)
was added dropwise and allowed to react at
24 °C overnight. After quenching with MeOH,
the suspension was evaporated in vacuo, di-
luted with toluene, and washed twice with
ice-cold water. The toluene layer was dried
with Na2SO4, concentrated, and subjected to
column chromatography on silica gel using 2:1
hexane–EtOAc as eluant. Crystallization from
EtOH afforded 4 (2.93g, 92%); mp 117–
118 °C; [h]2D8 +32.8° (CHCl3). Anal. Calcd for
C33H38O7: C, 72.51; H, 7.01. Found: C, 72.32;
H, 6.78.

Table 2
1H NMR data for propenyl galactosides 8-12 and 14-17, and galactosyl imidate 19 a
H-1
(d, J1,2
H-2
(dd, J2,3
H-3
(dd, J3,4
H-4
(bd)
H-5
H-6
(dd, J6,6
H-6
(dd)
OꢀCH2
(m)
CH
(m)
CH2
(m)
CH3O
(s)
PhꢀCH
(s)
PhꢀCH2
(d)
)
)
)
)
8
4.35
(7.3)
4.36
(8.1)
4.43
(7.7)
4.46
(7.3)
4.27
(7.7)
4.41
(7.7)
4.40
(7.7)
4.40
(8.0)
4.47
(7.7)
6.52
(3.6)
3.79
(9.5)
4.01
(9.5)
3.85
(9.9)
3.67
(9.5)
3.67
(9.5)
3.84
(9.9)
3.65
3.71
(3.7)
3.47
(3.7)
3.53
(3.7)
3.74
(3.7)
3.34
(3.3)
3.50
(2.9)
3.66
4.22
4.12
4.06
4.23
3.99
3.73
3.78
3.85
3.94
4.18
3.49
3.35
3.29
3.45
4.35
(12.5)
4.31
(12.8)
4.29
(12.5)
4.34
(12.5)
3.75
(12.1)
3.76
(11.5)
4.26
(11.4)
3.58
4.09
4.04
3.99
4.09
3.72
3.48
4.09
3.58
3.61
3.56
4.45
4.14
4.42
4.14
4.45
4.13
4.47
4.15
4.36
4.14
4.40
4.13
4.41
4.12
4.40
4.10
4.41
4.12
5.97
5.97
5.96
5.96
5.96
5.94
5.94
5.92
5.93
5.33
5.24
5.32
5.21
5.33
5.18
5.35
5.21
5.32
5.15
5.31
5.17
5.32
5.20
5.30
5.16
5.31
5.18
5.56
5.46
5.48
5.56
9
3.80
3.78
4.70/4.68
/
10
11
12
14
15
16
17
19
4.93/4.77, 4.70/4.68
5.00/4.73
3.42
(m)
3.35
(bt)
3.60
3.77
3.81
4.68/4.59
4.95/4.65, 4.94/4.77,
4.73/4.67
3
4.99/4.67, 4.85/4.66
)
3.81
(9.6)
3.79
(10.3)
4.20
(10.6)
3.49
(3.0)
4.89
3.51
3.67
4.28
3.78
4.93/4.60, 4.92/4.76
4.66/4.65, 4.43/4.41
4.88/4.64, 4.59/4.50
4.49/4.44
167
3.61
180
5.32
(3.3)
3.60
(9.5)
4.67/4.63, 4.61/4.52
4.50/4.42
a Values in parentheses are the associated coupling constants reported in Hz.
The solvent employed was CDCl3, except for 12, which was measured in CD3OD.

Table 3
1H NMR data for the unsulfated and sulfated galabioses 20–21, 23–25 and 27–40 a
H-1
H-2
H-3
H-4
H-5
H-6
H-6
H-1%
H-2%
H-3%
H-4%
H-5%
H-6%
H-6%
(J1,2
)
(J2,3
)
(J3,4
)
(J6,6
)
(J1%,2%
)
(J2%,3%
)
(J3%,4%
)
(J6%,6%)
20
21
23
24
25
26
27
28
29
4.48
(7.5)
4.43
(7.3)
4.41
(7.3)
4.45
(7.9)
4.41
(7.9)
4.67
(7.9)
4.40
(7.0)
4.42
(7.7)
4.37
(7.9)
4.64
4.29
(7.3)
4.32
(7.3)
4.23
(7.9)
4.16
(7.3)
4.47
(7.9)
5.45
(7.7)
4.48
(7.3)
4.56
(7.3)
4.53
(7.9)
4.64
(7.9)
3.90
(9.9)
3.93
(9.8)
3.83
(10.4)
3.68
(9.8)
3.66
3.80
(3.5)
3.99
(3.7)
3.76
4.29
3.56
4.34
(12.5)
4.30
4.06
5.22
(3.1)
4.99
(7.9)
5.20
(3.1)
5.25
(3.6)
5.26
(3.1)
5.04
(3.1)
4.86
(7.3)
4.87
(7.9)
4.85
(7.3)
4.72
5.28
(3.6)
5.26
(3.7)
5.29
(3.6)
5.28
(3.7)
5.20
(3.7)
5.39
(3.6)
5.22
(3.7)
5.27
(3.6)
5.30
(3.7)
5.20
(3.7)
4.06
(10.1)
3.85
(9.8)
3.86
(10.4)
4.06
(10.4)
4.03
(10.4)
4.02–3.99
4.00
(3.5)
3.42
(3.6)
4.02
(3.1)
5.10
(3.1)
5.09
3.85
4.01
3.49
4.26
4.26
4.39
3.82
4.39
4.39
3.73
3.97
(12.5)
4.19
3.51
4.36
3.80
3.37
3.38
3.73
3.75
4.05
3.51
3.11
4.04
4.42
4.41
3.94
3.15
3.75
(11.0)
4.09
3.39
(11.0)
4.00
(9.2)
4.04
3.85
(2.4)
3.88
(2.4)
3.78
4.03
4.08
3.86
4.04
4.10
4.08
3.86
(10.4)
3.62
(9.2)
4.20–4.14
4.20–4.14
3.86
4.20–4.14
4.20–4.14
3.49
4.62
4.63
4.20–4.14
3.63
3.91–3.86
3.91–3.86
3.40
3.81
3.82
3.74
(11.4)
4.14
(10.3)
4.16
(10.7)
4.22–4.16
4.18
(11.0)
3.71
3.66
(11.0)
3.73
(9.9)
3.75
3.40
3.76
3.78
3.75
(11.4)
4.14
/
3.68
(9.5)
3.64
(9.8)
3.64
3.84
(10.3)
3.82
(9.8)
3.68
(10.4)
3.62
3.92
(3.3)
3.92
3.99
4.10
4.55
(3.3)
4.55
4.14
4.0
4.07
4.17
4.16
(9.8)
30
31
3.82
3.75
4.22–4.16
4.36
5.48
4.22–4.16
3.46
(9.9)
3.51
(9.8)
3.41
(11.0)
3.41
(11.0)
3.76–3.71
4.22–4.16
3.80
3.80
3.41
3.30
3.99
4.19
(10.3)
3.85
(10.4)
4.10
(10.4)
4.10
(10.4)
4.01
3.90
3.65
4.27
4.27
4.35
4.30
3.55
4.30
4.31
4.39
4.42
4.37
3.24
32
33
34
35
36
37
38
39
40
3.73
3.83
3.44
4.08
4.07
5.21
5.18
4.59
5.39
4.06
5.03
5.03
4.62
3.37
2.88
5
(11.6)
4.09
0
4.04
3.65
4.40
3.82
4.01
3.64
4.08
4.23–4.16
4.34
4.07
4.38
2.91
167
180
3.64
3.78
4.23–4.16
ꢀ3.85
4.27
3.76–3.71
3.35
4.23–4.16
4.07
4.23–4.16
4.17
3.76–3.71
ꢀ3.84
3.14
3.79
(9.9)
3.90
(9.9)
3.78
(10.4)
3.76
(9.8)
3.61
3.97
4.06
ꢀ3.86
3.85
3.39
4.34
(12.1)
4.19
4.07
3.85
(9.9)
3.97
(10.4)
4.00
(9.8)
4.01
(10.4)
3.88
3.41
(11.4)
4.06
(9.8)
4.08
3.94
4.50
3.52
4.14
4.23
3.89
ꢀ3.98
3.77–3.71
4.51
3.54
4.19
4.16
4.28
3.90
(9.8)
4.21–4.13
4.21–4.13
3.67
3.77–3.71
3.77–3.71
4.42
4.21–4.13
(9.8)
a Values in parentheses are the associated coupling constants reported in Hz.
The solvents employed were CDCl3 for 20–21, 23, 27, 32 and 37, D2O for 26, 30, 35 and 40, and CD3OD for the others.

Table 4
13C NMR data for 3, 6, 10–12, 14, 15, 17, and 19 a
C-1
C-2
C-3
C-4
C-5
C-6
O-CH2
CH2CH2 CH2-CH
CH
CH2
Ph-CH
Ph-CH2
CH3O
55.3
CH3CO
3
6
103.0
103.9
102.7
102.5
104.0
103.2
102.8
102.8
94.6
70.0
76.7
78.5
79.3
71.8
79.7
79.3
77.0
73.2
78.7
75.0
78.9
72.4
82.1
82.0
74.2
75.0
72.2
73.2
73.8
74.1
75.5
67.2
73.0
74.7
74.5
74.9
66.7
71.7
66.5
66.5
76.5
74.6
72.2
73.1
71.3
69.3
62.2
69.2
69.1
62.5
62.0
62.8
68.3
67.7
30.2
30.2
70.2
70.2
71.0
70.4
70.1
70.2
28.6
28.9
69.1
69.5
138.2
138.0
134.3
134.1
135.9
134.3
134.0
134.0
114.8
115.0
117.1
117.3
117.4
117.1
117.3
117.2
101.1
71.1
74.9, 74.7
75.3, 71.7
74.9
10
11
12
14
15
17
19
101.4
101.4
55.2
72.3
55.7
55.3
75.3, 74.1, 73.1
74.9, 74.7
75.0, 74.7, 73.5
75.3, 73.4, 72.8
20.8
20.8
21.0
/
a The solvent employed was CDCl3, except for 12, which was measured in CD3OD.
Table 5
13C NMR data for the unsulfated and sulfated galabioses 20–21, 23–25, 27–29, 32–34 and 37–39 a
C-1
C-2 C-3 C-4 C-5 C-6 C-1%
C-2% C-3% C-4% C-5% C-6% O-CH2 CH
(CH2) b
CH2 (CH2-S) b Ph-CH
Ph-CH2
CH3O
3
20 102.8 77.3 74.5 71.8 66.4 69.3
21 102.7 79.5 75.1 76.1 66.7 69.0 103.2 78.6 78.3 74.3 66.4 69.2 70.2
94.0 75.2 75.0 74.9 62.5 69.2 70.4
134.2
134.2
134.1
135.7
31.1
134.1
135.5
31.4
134.2
135.6
29.9
134.1
135.7
30.8
117.2
117.2
117.3
117.4
29.1
117.3
117.4
29.1
117.1
117.4
29.0
117.3
117.3
29.0
101.3, 101.0
101.1, 100.6
75.3, 72.5, 71.7
75.1, 74.8, 71.6
55.2
55.3
(
23 103.3 78.5 77.7 72.4 70.1 62.3
24 104.6 79.2 78.8 74.3 74.1 67.5
25 105.2 79.1 78.4 74.0 73.9 67.5
27 103.1 80.0 79.6 75.0 74.6 62.0 104.1 79.7 74.2 75.3 74.5 61.8 70.4
28 104.0 81.2 81.1 78.0 74.1 67.9 105.1 79.2 81.8 76.1 73.7 67.1 71.3
29 105.1 81.1 81.4 78.2 74.2 68.4 105.2 79.2 81.8 76.1 73.8 67.3 69.5
32 103.4 78.4 77.9 72.0 74.4 62.0
33 104.7 79.3 77.8 73.7 74.2 67.4
34 105.6 79.2 77.5 73.7 74.0 68.0
37 102.8 77.5 76.2 71.7 66.3 69.3
38 104.0 78.5 75.4 72.8 67.7 70.3
39 104.6 78.5 75.3 72.8 67.8 70.4
94.9 77.4 70.6 77.3 74.4 62.0 70.4
97.3 75.8 79.3 77.3 69.9 67.6 71.4
96.6 75.6 79.3 77.2 69.7 67.6 69.2
75.4, 75.1, 74.4, 74.1
76.4, 76.2, 76.1, 75.5
76.4, 76.1, 76.0, 75.5
75.0, 74.8, 74.6, 74.2
76.4, 76.2, 76.0, 75.9
76.3, 76.1, 76.0, 75.9
75.2, 75.2
76.6, 76.1, 76.1, 75.7
76.6, 76.1, 76.1, 76.0, 73.0
75.4, 74.9, 71.4
167
180
95.2 77.6 70.3 77.3 68.9 68.8 70.4
96.2 75.9 79.6 77.3 69.3 70.5 71.4
96.0 75.8 79.5 77.0 69.1 70.5 69.5
92.1 73.8 70.3 76.5 70.0 62.7 70.2
94.2 74.9 78.2 77.0 70.1 67.9 71.4
94.1 75.0 78.3 77.0 70.1 68.2 68.6
101.5
102.3
102.1
76.3, 76.3, 73.0
76.4, 76.3, 73.1
a The solvents employed were CDCl3 for 20–21, 23, 27, 32 and 37, and CD3OD for the others.
b The formulas in the parentheses apply to 25, 29 and 34 due to the substitution to cysteamine.

T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
175
pound 9. The mother liquor was chro-
matographed on silica gel (8:1
4-Pentenyl 3,6-di-O-acetyl-2,4-di-O-benzyl-
i- -galactopyranoside (6).—To a stirred solu-
D
toluene–EtOAc) to afford additional 9. The
total yield was 4.63 g (81%); mp 165–
165.5 °C; [h]2D7 +32.8° (CHCl3); Rf 0.21 (2:1
toluene–EtOAc). Anal. Calcd for C24H28O7:
C, 67.28; H, 6.59. Found: C, 67.10; H, 6.49.
2-Propenyl 2-O-benzyl-4,6-O-benzylidene-3-
tion of 4 (1.60 g, 2.93 mmol) in 20 mL of dry
1:1 CH2Cl2 –Et2O, LiAlH4 (890 mg) was
added portionwise and allowed to react for 5
min at 22 °C, followed by the dropwise addi-
tion of an AlCl3 (2.93 g) suspension in dry
Et2O (7 mL) over 7 min. The mixture was
boiled under reflux for 1 h.25 After quenching
with EtOAc and ice, the suspension was
filtered through Celite and the residual solid
rinsed several times with acetone. The residue
was diluted with CHCl3, washed with water,
and chromatographed on a column of silica
gel (5:1 toluene–EtOAc) to yield a mixture of
3,4-OH and 3,6-OH (5) derivatives (0.583 g,
47%, Rf 0.51 (1:4 toluene–EtOAc)). Acetyla-
tion of the resultant mixture with Ac2O (3.5
mL) and dry Py (3.5 mL) overnight, permitted
separation by column chromatography on sil-
ica gel (8:1 hexane–EtOAc), affording the title
compound 6 (0.184 g, 26%, Rf 0.21 (4:1 tolu-
ene–EtOAc)) and 3,4-O-acetylated isomer
(0.48 g, 69%, Rf 0.38 (4:1 toluene–EtOAc)). 6;
[h]2D1 +14.6° (CHCl3). Anal. Calcd for
C29H36O8: C, 67.95; H, 7.08. Found: C, 67.82;
H, 7.12.
O-(4-methoxybenzyl)-i- -galactopyranoside
D
(10).—To a stirred solution of 9 (5.15 g, 12.0
mmol) in 50 mL dry THF, NaH (60%, 2.88 g,
72 mmol) and benzyl bromide (8.29 mL, 69.7
mmol) were added sequentially and allowed to
react overnight at 22 °C. The mixture was
processed as for 4, chromatographed on a
column of silica gel and eluted with 2:1 tolu-
ene–CHCl3. The residue was crystallized from
EtOH to afford the title product 10 (5.46 g,
88%); mp 115–116 °C; [h]2D5 +38.7° (CHCl3);
Rf 0.39 (4:1 toluene–EtOAc). Anal. Calcd for
C31H34O7: C, 71.80; H, 6.61. Found: C, 71.77;
H, 6.76.
2-Propenyl 2-O-benzyl-4,6-O-benzylidene-
i- -galactopyranoside (11).—A solution of 10
D
(4.91 g, 9.47 mmol) and 2,3-dichloro-5,6-di-
cyano-1,4-benzoquinone (2.57 g, 11.3 mmol)
in 50 mL 25:1 CH2Cl2–H2O, was stirred for 1
h at 22 °C. The mixture was diluted with
CHCl3 and washed twice with NaHCO3. The
organic layer was dried with Na2SO4, concen-
trated and applied to a column of silica gel
eluted by 8:1 toluene–EtOAc. The residue was
evaporated in vacuo and crystallized from tol-
uene at 4 °C to afford 11 (2.93 g, 78%); mp
73.5–74 °C; [h]2D7 +12.6° (CHCl3); Rf 0.32
(2:1 toluene–EtOAc). Anal. Calcd for
C23H26O6: C, 66.33; H, 6.78. Found: C, 66.35;
H, 6.72.
2-Propenyl
methoxybenzyl)-i-
4,6-O-benzylidene-3-O-(4-
-galactopyranoside (9).—
D
A mixture of 7 (9.95 g, 45.2 mmol),35
benzaldehyde (32 mL, 305 mmol) and formic
acid (32 mL, 848 mmol) was stirred for 0.5 h
at 22 °C. Addition of Et2O gave crystalline 8.
The residual solution was neutralized with
solid NaHCO3 and Et2O and petroleum ether
were added so that more 8 could be obtained
as an amorphous solid. The solid was recrys-
tallized from MeOH and Et2O to afford 8
(9.69g, 70%); mp 175–176 °C (Lit.: 178–
179 °C36); Rf 0.23 (1:4 toluene–EtOAc). A
suspension of 8 (4.13 g, 13.4 mmol) in toluene
(40 mL) was azeotropically distilled in the
presence of Bu2SnO (3.66 g, 1.1 fold excess) in
the same manner as for 3. To the mixture,
tetrabutylammonium iodide (6.43 g, 1.3 fold
excess) and 4-methoxybenzyl chloride (2.41
mL, 1.3 fold excess) were added and the mix-
ture was refluxed for 30 min. The solution was
diluted with CHCl3 and washed with water.
The organic layer was dried with Na2SO4 and
crystallized from EtOH to yield the title com-
2-Propenyl
3-O-(4-methoxybenzyl)-i- -
D
galactopyranoside (12).—Compound 9 (3.95
g, 9.22 mmol) was treated with 80% aqueous
AcOH (20 mL) for 50 min at 86 °C. The
solution was evaporated in vacuo (toluene).
The residue was made neutral with aqueous
NaHCO3, the solution was evaporated, and
the residue extracted with acetone several
times for chromatography on a silica gel
column (1:4 toluene–EtOAc). The residue was
crystallized from EtOH and toluene to afford
the title compound 12 (1.85 g, 59%); mp 115–
116 °C; [h]2D4 +16.6° (MeOH); Rf 0.52 (8:1

176
T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
(10.5 mL) and processed as for 11. After
column chromatography on silica gel (4:1 hex-
ane–EtOAc), the residue was acetylated with
Ac2O and Py to afford the title compound 17
(1.04 g, 89%); [h]2D7 +40.7° (CHCl3); Rf 0.44
(4:1 toluene–EtOAc). Anal. Calcd for
C32H36O7: C, 72.16; H, 6.81. Found: C, 71.84;
H, 7.03.
EtOAc–MeOH). Anal. Calcd for C17H24O7:
C, 59.99; H, 7.11. Found: C, 59.90; H, 7.10.
2-Propenyl 2,4-di-O-benzyl-3-O-(4-methoxy-
benzyl)-i- -galactopyranoside (14).—A mix-
D
ture of 12 (0.34 g, 1 mmol) and Ph3CCl (0.56
g, 2 mmol) in dry Py (4 mL) was heated for 18
h at 37 °C. Ice was added, and the mixture
concentrated in vacuo, and the residue chro-
matographed on silica gel (2:1 hexane–
EtOAc) to yield 13 as a syrup (0.505 g, 86.7%,
Rf 0.36 (2:1 toluene–EtOAc)). Compound 13
was 2,4-O-benzylated in dry DMF and pro-
cessed in the same way as 4, employing a
6-fold excess of NaH and a 5-fold excess of
benzyl bromide. The resultant mixture was
treated with 80% aqueous AcOH (4 mL) for 1
h at 90 °C, concentrated in vacuo and sub-
jected to chromatography on silica gel, eluting
with 3:1 hexane–EtOAc. The residue was
crystallized from EtOH to afford the title
compound 14 (0.279 g, 62%, as needles); mp
94.5–96.0 °C; [h]2D4 +18.5° (CHCl3); Rf 0.29
(2:1 toluene–EtOAc). Anal. Calcd for
C31H36O7: C, 71.52; H, 6.97. Found: C, 71.25;
H, 6.97.
3-O-Acetyl-2,4,6-tri-O-benzyl-i- -galacto-
D
pyranosyl trichloroacetimidate (19).—A mix-
ture of 17 (1.04 g, 1.95 mmol) and
RhCl(PPh3)3 (0.37 g) in 7:3:1 EtOH–toluene–
water (22 mL) was refluxed for 20 h and
treated with aqueous 2 N HCl (1 mL) under
reflux for 1 h.37 After being neutralized with 1
M NaHCO3 (2 mL), the mixture was evapo-
rated, diluted with CHCl3 and washed with
aqueous NaHCO3. The residue was chro-
matographed on a column of silica gel, eluted
with 3:1 hexane–EtOAc, to yield an a/b mix-
ture of 18 (0.773 g, 81%, Rf 0.33, 0.37, (2:1
toluene–EtOAc)). To a solution of 18 (0.773
g, 1.57 mmol) in dry CH2Cl2 (15 mL),
trichloroacetonitrile (0.647 mL) and NaH
(60%, 96 mg) were added sequentially, and the
solution was stirred for 1 h at 22 °C.38 Chro-
matography on silica gel, eluting with 15:1
hexane–EtOAc plus 0.1% Et3N, gave the title
compound 19 (0.748 g, 75%); Rf 0.49 (8:1
2-Propenyl 6-O-acetyl-2,4-di-O-benzyl-i- -
D
galactopyranoside (15).—Compound 14 (0.399
g, 0.766 mmol) was acetylated conventionally
with Ac2O (4 mL) in dry Py (4 mL). The
residue was dissolved in 20:1 CH2Cl2 –H2O
(5.25 mL) and treated with 2,3-dichloro-5,6-
dicyano-1,4-benzoquinone (0.191 g) as de-
scribed for 11. After washing with aqueous
NaHCO3 and concentrating, the mixture was
chromatographed using 3:1 hexane–EtOAc to
afford the title compound 15 (0.283 g, 96%);
[h]2D7 –0.6° (CHCl3); Rf 0.23 (4:1 toluene–
EtOAc). Anal. Calcd for C25H30O7: C, 67.86;
H, 6.83. Found: C, 67.65; H, 6.75.
toluene–EtOAc).
Anal.
Calcd
for
C31H32O7Cl3N·H2O: C, 56.85; H, 5.23; N,
2.14. Found: C, 56.59; H, 4.84; N, 2.27.
2-Propenyl 2-O-benzyl-4,6-O-benzylidene-3-
O-[2-O-benzyl-4,6-O-benzylidene-3-O-(4-
methoxybenzyl) - h -
D
- galactopyranosyl] - i - -
D
galactopyranoside (20) and 2-Propenyl 2-O-
benzyl-4,6-O-benzylidene-3-O-[2-O-benzyl-4,
6-O-benzylidene-3-O-(4-methoxybenzyl)- i-
galactopyranosyl]-i- -galactopyranoside (21).
D
-
D
2-Propenyl 3-O-acetyl-2,4,6-tri-O-benzyl-i-
-galactopyranoside (17).—A stirred solution
—The donor compound 4 (0.599 g, 1.10
mmol) and acceptor compound 11 (0.398 g,
1.0 mmol) were dissolved in 10 mL of dry 1:4
D
of 12 (0.749 g, 2.20 mmol) in dry DMF (8
mL), chilled on ice, was treated with NaH
(60%, 0.555 g) and benzyl bromide (1.57 mL,
13.2 mmol) was added dropwise. After stirring
overnight at 24 °C, the reaction was quenched
with MeOH, the solution evaporated in vacuo
and the residue processed as described for 4.
The product 16 (Rf 0.51, 4:1 toluene–EtOAc)
was treated with 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone (550 mg) in 20:1 CH2Cl2 –water
,
CH2Cl2–Et2O in the presence of ground 4 A
molecular sieves. Iodinium di-sym-collidine
perchlorate (0.93 g) freshly prepared accord-
ing to the established method39 was added to
the solution under stirring. After 20 h, the
mixture was filtered through Celite, diluted
with CHCl3, and washed sequentially with
aqueous H2SO4 and NaHCO3. The residue
was subjected to chromatography on silica gel,

T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
177
water). Anal. Calcd. For C43H47O20S3Na3·
8H2O: C, 43.29; H, 5.32, Found: C, 43.30; H,
4.90.
eluting stepwise with 2:3, 1:2, and 1:4 hexane–
CHCl3 such that the a and b anomers (20 and
21, Rf 0.56 and 0.48 (2:1 toluene–EtOAc),
respectively) were separated efficiently. Each
anomer was crystallized from EtOH to yield
20 (0.412 g, 48%) and 21 (0.155 g, 18%).
Compound 20 had mp 171.5–172.5 °C; [h]D27
+123° (CHCl3). Anal. Calcd for C51H55O13:
C, 71.31 H, 6.34. Found: C, 71.03; H, 6.27.
Compound 21 had mp 190–191.5 °C; [h]D19
+46.2° (CHCl3). Anal. Calcd for C51H55O13:
C, 71.31 H, 6.34. Found: C, 71.17; H, 6.24.
2-Propenyl 2,4-di-O-benzyl-3-O-(2,4-di-O-
3-(2-Aminoethylthio)propyl 2,4-di-O-benzyl-
3-O-(2,4-di-O-benzyl-3,6-di-O-sulfo-h-
D
-gal-
actopyranosyl)-i- -galactopyranoside trisod-
D
ium salt (25).—Compound 24 (42.9 mg, 40.9
mmol) was dissolved in 1:1 MeOH–water (1
mL) and allowed to react with cysteamine
HCl (48 mg) for 1.5 h at 24 °C, under UV
irradiation (264 nm). Chromatography of the
mixture on a column of Sephadex LH-20 (1×
48 cm), eluted with MeOH supplemented with
0.5% Et3N, afforded the tri(Et3N) salt (49.0
mg, 88%). The counter-ion was changed into
Na by the addition of 108 mL of 1 M
NaHCO3 and evaporation to give the title
compound 25; Rf 0.11 (6:2:1 EtOAc–AcOH–
water). FAB-MS data: (+) 1127 (M+H),
1105 (M+2H−Na); (−) 1103 (M−Na).
3-(2-Aminoethylthio)propyl-6-O-sulfo-3-O-
benzyl-h-
D
-galactopyranosyl)-i- -galactopy-
D
ranoside (23).—To a stirred solution of 20
(0.389 g, 0.453 mmol) in 9 mL dry 1:1
CH2Cl2–Et2O, was added LiAlH4 (0.138 g)
portionwise and allowed to react at 26 °C for
10 min, followed by dropwise addition of
AlCl3 (478 mg) suspended in dry Et2O (3 mL).
The mixture was refluxed for 30 min,
quenched sequentially with EtOAc and
MeOH, and the residue filtered off through
Celite, washing the solid with MeOH several
times. Chromatography on silica gel gave 22
(0.128 g, 33%, Rf 0.48 (1:4 toluene–EtOAc)),
as well as de-3%-O-methoxybenzylated 23
(0.017 g, 5.1%, Rf 0.43 (1:4 toluene–EtOAc)),
and the 6-O-benzyl derivative (0.052 g, 13%,
Rf 0.69 (1:4 toluene–EtOAc)). Compound 22
(0.102 g, 0.119 mmol) was treated with 2,3-
dichloro-5,6-dicyano-1,4-benzoquinone as de-
scribed for 11 to afford the title compound 23
(0.069 g, 78%); [h]2D4 +86.5° (CHCl3). Anal.
Calcd for C43H50O11·0.5H2O: C, 68.69; H,
6.84. Found: C, 68.51; H, 7.00.
(3,6-di-O-sulfo-h-
D
-galactopyranosyl)-i- -
D
galactopyranoside (26).—Compound 25 (30.7
mg, 27.3 mmol) was dried, placed on ice and
ice-chilled trifluoroacetic acid (0.3 mL), bro-
motrimethylsilane (52.8 mL, 1 M final) and
thioanisole (46.8 mL, 1 M final) were added.
After 30 min on ice, the reaction was stopped
with 3.84 mL 1 M NaHCO3. The desired
compound (20.5 mg, 98%) was isolated on a
column of Sephadex LH-20 column (1×47
cm) eluted with 1:1 MeOH–water, 0.5% Et3N;
Rf 0.11 (3:2:1 EtOAc–AcOH–water). FAB-
MS data: (+) 745 (M+2H−Na).
2-Propenyl 2,4-di-O-benzyl-3-O-(2,4-di-O-
benzyl-i-
D
-galactopyranosyl)-i- -galactopy-
D
2-Propenyl 2,4-di-O-benzyl-3-O-(2,4-di-O-
ranoside (27).—Reductive cleavage of the ben-
zylidene groups in 21 (0.327 g, 0.381 mmol)
was performed as for 20, employing LiAlH4
(0.118 g, 8-fold excess) and AlCl3 (0.411 g). In
this step, the 3%-O-methoxybenzyl group was
removed simultaneously, as determined by
NMR and elemental analysis. After quenching
the reaction with EtOAc and MeOH, the mix-
ture was filtered through a layer of Celite, and
washed with MeOH. The filtrate and washings
were concentrated in vacuo and chro-
matographed on a column of silica gel (1:2
toluene–EtOAc) to yield 27 (100 mg, 35%)
along with other positional isomers (81 mg,
benzyl-3,6-di-O-sulfo-h-
D
-galactopyranosyl)-
6-O-sulfo-i-
D
-galactopyranoside trisodium salt
(24).—A mixture of 23 (68.9 mg, 0.0928
mmol) and sulfur trioxide–Py (133 mg, 3 fold
excess) in 1:1 dry Py–DMF (1 mL) was
heated for 18 h at 56 °C. The reaction was
quenched with 1.67 mL of 1 M NaHCO3 and
the solvent was evaporated in vacuo. The
residue was dissolved in MeOH and filtered
through Celite to remove some salt. Chro-
matography on a Sephadex LH-20 column (30
cm), eluted with MeOH, afforded the title
compound 24 (74 mg, 76%); [h]1D5 +69.7°
(MeOH); Rf 0.42 (3:2:1 EtOAc–2-propanol–

178
T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
29%). Compound 27; [h]2D0 −4.9° (CHCl3); Rf
0.26 (1:4 toluene–EtOAc). Anal. Calcd. for
C43H50O11·H2O: C, 67.88; H, 6.89. Found: C,
68.13; H, 6.78.
10% solution of BF3–Et2O complex (1.07 mL)
in CH2Cl2 was added dropwise in the presence
,
of ground molecular sieves (4A). After stirring
at for 18 h 24 °C, the mixture was filtered
through Celite and chromatographed on a
column of silica gel (5:1 hexane–EtOAc) to
afford 31 (342 mg, 52%, Rf 0.29 (4:1 toluene–
EtOAc)). Compound 31 was deacetylated in
41 mM NaOCH3 in dry MeOH (3 mL) to give
the title compound 32 (275 mg, 89%); [h]D25
+49.5° (CHCl3); Rf 0.51 (1:1 toluene–
EtOAc). Anal. Calcd. for C50H56O11: C, 72.10;
H, 6.78. Found: C, 71.86; H, 6.80.
2-Propenyl 2,4-di-O-benzyl-3-O-(2,4-di-O-
benzyl-3,6-di-O-sulfo-i-
D
-galactopyranosyl)-
6-O-sulfo-i-
D
-galactopyranoside trisodium salt
(28).—Compound 27 (90.3 mg, 121.6 mmol)
was O-sulfated by sulfur trioxide–Py (174 mg,
3-fold excess) in dry 1:1 Py–DMF (4 mL) for
16 h at 56 °C. After quenching with 2.19 mL
of 1 M NaHCO3, the residue was processed as
for 24. Chromatography on a column (1×19
cm) of Sephadex LH-20 with MeOH gave the
2-Propenyl 2,4-di-O-benzyl-3-O-(2,4,6-tri-
title compound 28 (126.8 mg, 99%); [h]D16
+
O-benzyl-3-O-sulfo-h-
O-sulfo-i-
D
-galactopyranosyl)-6-
D
-galactopyranoside disodium salt
15.9° (MeOH); Rf 0.38 (3:2:1 EtOAc–iso-
propanol–water). Anal. Calcd. For
(33).—Syrupy 32 (30.5 mg, 36.6 mmol) was
dissolved in dry 1:1 Py–DMF (0.6 mL) and
subjected to O-sulfation by sulfur trioxide–Py
(35 mg, 3-fold excess) for 20 h at 56 °C. The
resulting mixture was processed as for 24, to
afford the title compound 33 (31 mg, 82%);
[h]1D5 +58.2° (MeOH); Rf 0.38 (6:2:1 EtOAc–
C43H47Na3O20S3·6H2O: C, 44.64; H, 5.14,
Found: C, 43.84; H, 4.77.
3-(2-Aminoethylthio)propyl 2,4-di-O-benzyl-
3 -O-(2,4-di-O-benzyl-3,6-di-O-sulfo-i-
galactopyranosyl)-6-O-sulfo-i-
D
-
D
-galactopy-
ranoside trisodium salt (29).—Compound 28
(29.5 mg, 28.1 mmol) was dissolved in 1:1
MeOH–water (1 mL) and allowed to react
with cysteamine HCl (33.8 mg) for 1 h at
24 °C, under UV irradiation (254 nm). The
mixture was subjected to chromatography on
a Sephadex LH-20 column (1×47 cm), eluted
with MeOH supplemented with 0.5% Et3N, to
afford the tri(Et3N) salt (30.5 mg, 94%). The
ion form was converted as for 25 to afford the
title compound 29; Rf 0.12 (6:2:1 EtOAc–
AcOH–water). FAB-MS data: (+) 1127
(M+H), 1105 (M+2H−Na); (−) 1103
(M−Na).
isopropanol–water).
Anal.
Calcd.
for
C50H54Na2O17S2·3H2O: C, 55.04; H, 5.54.
Found: C, 55.00; H, 5.28.
3-(2-Aminoethylthio)propyl 2,4-di-O-benzyl-
3-O-(2,4,6-tri-O-benzyl-3-O-sulfo-h-
D
-galac-
topyranosyl)-6-O-sulfo-i-
D
-galactopyranoside
disodium salt (34).—A solution of 33 (27.2
mg, 26.2 mmol) in 2:1 MeOH–water (1 mL)
was allowed to react with cysteamine HCl (48
mg) for 2 h at 24 °C under UV irradiation
(254 nm). The mixture was chromatographed
and handled as described for 25 to afford the
tri(Et3N) salt (32.1 mg, 96%). The ion form
was converted as for 25 to give the title com-
pound 34; Rf 0.41 (6:2:1 EtOAc–AcOH–wa-
ter). FAB-MS data: (−) 1091 (M−Na).
3-(2-Aminoethylthio)propyl 3-O-(3-O-sulfo-
3-(2-Aminoethylthio)propyl 3-O-(3,6-di-O-
sulfo-i-
D
-galactopyranosyl)-6-O-sulfo-i- -
D
galactopyranoside trisodium salt (30).—Com-
pound 29 (29.5 mg, 26.2 mmol) was depro-
tected as for 26. The desired compound (16.8
mg, 84%) was isolated by using a column
(1×47 cm) of Sephadex LH-20 eluted with
1:1 MeOH–water, 0.5% Et3N; Rf 0.11 (3:2:1
EtOAc–AcOH–water). FAB-MS data: (+)
745 (M+2H−Na).
h-
D
-galactopyranosyl)-6-O-sulfo-i- -galacto-
D
pyranoside disodium salt (35).—Debenzylation
of 34 (27.8 mg, 25 mmol) was performed in the
same way as for 26. The yield was 13.6 mg
(82%); Rf 0.15 (3:2:1 EtOAc–AcOH–water).
FAB-MS data: (−) 641 (M−Na).
2-Propenyl 2-O-benzyl-4,6-O-benzylidene-3-
2-Propenyl 2,4-O-benzyl-3-O-(2,4,6-tri-O-
O-(2,4-di-O-benzyl-h- -galactopyranosyl)-i-
D
benzyl-h-
D
-galactopyranosyl)-i- -galactopy-
D
D
-galactopyranoside (37).—Glycosidation be-
ranoside (32).—A mixture of 15 (262 mg,
0.592 mmol) and 19 (504 mg, 0.791 mmol) in
CH2Cl2 (10 mL) was chilled at −80 °C, and a
tween 6 (174 mg, 0.340 mmol) and 11 (113
mg, 0.283 mmol) was performed in dry 1:4
CH2Cl2–Et2O (10 mL) using freshly prepared

T. Yoshida et al. / Carbohydrate Research 335 (2001) 167–180
179
iodinium di-sym-collidine perchlorate (0.20 g)
as described for 20 and 21. The resultant
mixture was processed as for 20, and chro-
matographed on a column of silica gel eluted
with 1:1 toluene–EtOAc, to afford 36 (78 mg,
33%); Rf 0.40 (2:1 toluene–EtOAc), while
leaving a significant amount of unreacted 11
(60 mg, 53%). Compound 36 was deacetylated
by conventional transesterification to give 37
in quantitative yield; [h]1D5 +99.0° (CHCl3); Rf
0.56 (1:4 toluene–EtOAc). Anal. Calcd. for
C43H48O11·H2O: C, 68.05; H, 6.64. Found: C,
68.07; H, 6.42.
are indebted to Dr Y. C. Lee, the Johns
Hopkins University, Baltimore MD, for his
great academic advice and support.
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