Synthesis and characterization of sulfated gal-β-1,3/4-GlcNAc disaccharides through consecutive protection/glycosylation steps

Article abstract of DOI:10.1002/asia.201201204

We have developed an expeditious procedure to yield large amounts of orthogonally protected Gal-β1,3/4-GlcNAc, which allowed for the systematic introduction of a sulfate group onto the C3/C6 positions of Gal and/or the C6 position of GlcNAc. In particular

Full text of DOI:10.1002/asia.201201204

FULL PAPER
DOI: 10.1002/asia.201201204
Synthesis and Characterization of Sulfated Gal-b-1,3/4-GlcNAc
Disaccharides through Consecutive Protection/Glycosylation Steps
Zhijay Tu,[a] Hsiao-Wu Hsieh,[a, b] Chih-Ming Tsai,[c] Chia-Wei Hsu,[a, b] Shy-Guey Wang,[a]
Kuan-Jung Wu,[a] Kuo-I Lin,*[c] and Chun-Hung Lin*[a, b, c]
Abstract: We have developed an expe-
ditious procedure to yield large
amounts of orthogonally protected
Gal-b1,3/4-GlcNAc, which allowed for
the systematic introduction of a sulfate
group onto the C3/C6 positions of Gal
and/or the C6 position of GlcNAc. In
particular, the disaccharide precursors
were prepared in five or six steps and
high overall yield from para-tolyl-6-O-
tert-butyldiphenylsilyl-1-thio-b-d-galac-
topyranoside. After deprotection and
sulfation steps, the final products were
characterized by using several NMR
methods to unambiguously confirm the
location of each introduced sulfate
group and they were examined for
their binding specificity of human ga-
lectin-1 and galectin-8.
Keywords: carbohydrates · galectin ·
glycosylation · protecting groups ·
sulfates
Introduction
date how the specific position of a sulfate affects the binding
and activity of the corresponding receptors (lectins). Several
groups have prepared sulfated carbohydrates. For example,
the groups of Hsieh–Wilson and Mallet prepared chondroi-
tin sulfate (CS) oligosaccharides and their analogues to
access their functional role in the modulation of neuron
growth and binding interactions with hepatocyte growth-
factor/scatter-factor and CXCL12 (a GAG-binding pro-
tein).[4] Others have established efficient methods for the
synthesis of heparin and heparan sulfate oligosaccharides.[5–9]
Kiso and co-workers were the first to synthesize 6-sulfo
sialyl Lewis x gangliosides and analogues as potent l-selec-
tin ligands. Pratt and Bertozzi recently established divergent
chemoenzymatic syntheses for a similar purpose.[10]
Laborious protection/deprotection steps are major im-
pediments to the development of efficient methods for the
synthesis of carbohydrates.[11] Specifically, the design of pro-
tection/deprotection steps that involve sulfated sugars is
troublesome because sulfates are labile and must be added
at a late stage in the synthesis. If sulfates are to be added at
different sites on a sugar, different synthetic precursors and
protection strategies must be employed, which inevitably
lengthens the procedure. Herein, we report efficient synthe-
ses of fully protected Gal-b1,3/4-GlcNAc disaccharides by
using consecutive protection and glycosylation steps. Subse-
quent deprotection and sulfation steps afforded two non-sul-
fated and six mono-sulfated saccharides (compounds 1–8,
Scheme 1). We introduced a sulfate group onto the C3 or
C6 positions of Gal, or the C6 position of GlcNAc, either of
which could be found at the non-reducing terminus of N- or
O-glycans. These glycan products were further examined for
their binding interactions with human galectins-1 and -8.
Our aim was to prepare common disaccharide precursors
that contained orthogonal protecting groups, which would
allow for the incorporation of one or more sulfate groups at
the desired position(s) and allow the prompt assembly of
Sulfate-containing glycans are involved in myriad physiolog-
ical processes, including cell morphogenesis and develop-
ment, viral invasion, tumor metastasis, inflammation, and
neural disorders.[1] For instance, the sulfated forms of a2,3-
sialylated, a1,3-fucosylated glycans, such as the sialyl Lewis
x determinant, are essential for recognition by l-selectin (a
calcium-ion-dependent lectin that is expressed in most leu-
kocytes).[2] Sulfation often occurs at the O6 position of
a Gal or GlcNAc residue. Lymphocytes in mice that lack
the two sulfotransferases that are responsible for sulfation
cannot target high endothelial venules.[2] Several carbohy-
drate-binding proteins, including selectins and galectins (b-
galactoside-binding lectins), have been shown to bind sulfat-
ed sugars.[3] However, the structure–activity relationships in
sulfated glycans are unclear, that is, it is important to eluci-
[a] Dr. Z. Tu, H.-W. Hsieh, C.-W. Hsu, Dr. S.-G. Wang, K.-J. Wu,
Prof. Dr. C.-H. Lin
Institute of Biological Chemistry
Academia Sinica
No. 128, Academia Road Section 2
Nan-Kang, Taipei, 11529 (Taiwan)
Fax : (+886)2-26514705
E-mail: chunhung@gate.sinica.edu.tw
[b] H.-W. Hsieh, C.-W. Hsu, Prof. Dr. C.-H. Lin
Department of Chemistry
National Taiwan University
Taipei (Taiwan)
[c] C.-M. Tsai, Prof. Dr. K.-I. Lin, Prof. Dr. C.-H. Lin
Genomics Research Center
Academia Sinica
Taipei (Taiwan)
Fax : (+886)2-27899931
E-mail: kuoilin@gate.sinica.edu.tw
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/asia.201201204.
Chem. Asian J. 2013, 8, 1536 – 1550
1536
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
Gal are protected as their TBDPS ether and Lev ester (i.e.,
compound 12), respectively, thus making it possible to spe-
cifically introduce a sulfate group in a later stage of the syn-
thesis. Acetylation at the O2 position was essential for con-
trolling the stereoselectivity during the glycosylation reac-
tion, thus resulting in the formation of a 1,2-trans-glycosidic
bond. Moreover, we also prepared glycosyl acceptors with
different protecting groups (Table 1), including amino
groups at the C2 position and 4,6-benzylidene (for b1,3-gly-
cosylation) and 3,6-dibenzyl ether groups (for b1,4-glycosy-
lation). These protecting groups are commonly used in car-
bohydrate chemistry and can be selectively removed.[7b,8c,d]
Because three protecting groups are installed on the donor
and two/three groups are installed on acceptors (Table 1), it
is necessary to optimize these glycosylation reactions.
Scheme 1. Structures of target disaccharides 1–8.
longer glycan chains by rapidly converting the disaccharide
precursors into glycosyl donors and acceptors. The modular
construction of orthogonally protected disaccharides has
greatly simplified and accelerated these synthetic processe-
s.[7a,12]
To optimize the glycosylation reaction of compound 12,
we prepared several glucosamine-type acceptors.[15] Com-
pound 12 was activated by using Ph2SO, 2,4,6-tri-tert-butyl-
pyrimidine (TTBP), and Tf2O, mainly for b1,3-glycosylation
(Table 1, conditions A). The same compound was treated
with N-iodosuccinimide (NIS) and TfOH to form a b1,4-gly-
cosidic bond (conditions B). Several acceptors gave moder-
ate-to-good yields, despite the presence of different amino-
protecting groups at the C2 position, including azido (N3),
2,2,2-trichloroethoxycarbonyl (Troc), and trichloroacetyl
(TCA) groups (Table 1). Interestingly, among the four ac-
ceptors that we examined (compounds 13–16; Table 1, en-
tries 1–5), only compound 13 was successfully glycosylated
with compound 12 to generate compound 17 in 86% yield
under the preactivation conditions. The O3-Lev protection
Results and Discussion
The primary hydroxy group of para-tolyl-1-thio-b-d-galacto-
pyranoside (which was directly derived from peracetate 9)
was masked by the formation of the tert-butyldiphenylsilyl
(TBDPS) ether.[13] Then, the product (10) was reacted with
trimethyl orthoacetate in the presence of a catalytic amount
of camphor sulfonic acid (CSA) to form orthoesters at the
C3 and C4 hydroxy groups. The resulting product was sub-
jected to acetylation and selective unmasking of the C3-hy-
droxy group under acidic conditions to generate compound
11 (87% yield). Further esterification with levulinic acid at
the O3 position afforded the fully protected thiogalactoside
12 (94% yield). Because the re-
action conditions that were
used to form compound 12
from compound 10 were com-
patible, the four-step synthesis
was performed consecutively
without chromatographic purifi-
cation of the intermediate prod-
ucts and it was only interrupted
by the removal of that solvents
and a simple workup between
each reaction (Scheme 2A).
After final purification, com-
pound 12 was obtained in 78%
yield (>50 g). Similar proce-
dures have been reported for
the preparation of elaborated
gluco-, glucosamino-, galacto-,
and digitoxosides.[14]
Our orthogonal protection
strategy offers several advan-
tages in the subsequent synthe-
ses. In general, regio- and ste-
reoselective glycosylations can
be achieved by the introduction
of various protecting groups.
Scheme 2. Consecutive operations in the syntheses of A) thiogalactoside 12 and B) b1,3-disaccharide 18 and
The O3 and O6 positions of b1,4-disaccharide 26. Each thick, blue arrow represents one reaction. M.S.=molecular sieves.
Chem. Asian J. 2013, 8, 1536 – 1550
1537
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
Table 1. Glycosylation reactions of b-galactosyl donor 12 with several GlcNAc-type acceptors.
For convenient product char-
acterization, the reaction in
Table 1, entry 1 was followed by
b-selective acetylation to afford
compound 18 in 80% yield
(2 steps). This preactivation
method was also applied to the
synthesis of acetate analogue 19
(Table 1, entry 6), albeit in
a lower yield. Compound 22,
which was directly derived from
Entry
Acceptor
Glycosylation
conditions[a]
Product
Yield
[%][b]
3-OH acceptor for b1,3-glycosylation
1
A
86
[c]
compound
13
in
three
2
A
B
–
steps,[8d,17] was coupled with
[c,d]
compound 12 under the same
glycosylation
3
4
14
–
conditions
(Table 1, entry 11) in 80%
yield, whereas the use of NIS/
TfOH as a promoter afforded
the product in 84% yield
(Table 1, entry 12). Acceptors
that contained a linker at the
reducing terminus (20, 21, and
23) were also tolerated in the
glycosylation reactions to afford
their corresponding disacchar-
ides in good yields (Table 1, en-
tries 8, 9, and 13).
[c]
A
A
–
[c]
5
–
6
7
A
A
56
70
19
18
8
B
71
The results listed in Table 1
show several useful findings.
First, galactosyl donor 12 suc-
cessfully reacted with six differ-
ent acceptors that were protect-
ed with two/three different pro-
tecting groups. Second, better
yields were obtained under con-
ditions B compared to those
9
A
B
57
77
10
21
25
26
4-OH acceptor for b1,4-glycosylation
11
A
B
80
84
under conditions
A (Table 1,
12
13
22
entries 6, 7, 9–12). In addition,
compound 13 was a diol and its
corresponding glycosylation re-
action showed complete regio-
selectivity at the O3 position.
The possible reaction mecha-
nism is different from that re-
ported for the base-mediated
esterification reaction.[18] When
B
71
[a] Glycosylation conditions: A) Ph2SO, Tf2O, TTBP, 3 ꢁ M.S., CH2Cl2/toluene/MeCN or CH2Cl2, ꢀ608C, 1 h;
B) NIS/TfOH, CH2Cl2, 3 ꢁ M.S. (for temperature and reaction time, see the Experimental Section). [b] Yield
of isolated product. [c] No glycosylated product was formed, owing to the hydrolysis of donor 12. [d] The ac-
ceptor (14) was even insoluble in MeCN and 1,4-dioxane.
of compound 12 appeared to be critical because the O3-ace-
tate analogue only gave low yields in the glycosidation reac-
tions, owing to the accompanying side reactions of acetyl
transfer or donor hydrolysis.[14f,16] In the reactions of accept-
ors 14–16, we observed significant hydrolysis of donor 12.
One possible explanation of this result is the poor solubility
of these acceptors. We had no success on trying to dissolve
these compound in several different solvents (such as
CH2Cl2, MeCN, 1,4-dioxane, and the mixture CH2Cl2/tolu-
ene/MeCN).
donor 12 forms the oxocarbenium intermediate through the
preactivation method, the 3-OH group of diol 13 can reacts
with the intermediate to produce compound 17 because of
its higher nucleophilicity than that of hemiacetal-OH group.
In contrast, the base-mediated esterification reaction was
found to predominantly occur at the hemiacetal-OH group.
The hydroxy group has a higher tendency for deprotonation
under basic conditions, likely owing to its lower pKa value.
Note that compound 17 is fully characterized (see the Ex-
perimental Section and the Supporting Information, S22–28)
and, in its HMBC spectrum, the resonance of the C3 atom
Chem. Asian J. 2013, 8, 1536 – 1550
1538
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
of the GlcNAc group correlates with the H1’ atom of the
Gal group (3J
(C,H) coupling) and vice versa.
30 in the presence of TMSOTf afforded tetrasaccharide 31
in 67% yield without further optimization. In comparison
with other published syntheses of Gal-b1,4-GlcNAc-repeat-
ing oligosaccharides,[20] our method not only decreases the
time that is needed to prepare a variety of disaccharide pre-
cursors, but also incorporates orthogonal protecting groups
that meet the requirement to modify the sugars, for exam-
ple, by sulfation or oxidation, at a specific position(s). In ad-
dition, we have achieved an eight-step synthesis of bianten-
nary heptasaccharide 32, in which the major step was the
glycosidation of donor 29 with a trimannoside acceptor
(Scheme 3B).[21]
ACHTUNGTRENNUNG
Thus, we selected compounds 13 and 22[19] for further in-
vestigation. More than 50 g of each of compounds 12, 13,
and 22 could be obtained in an effective manner. The afore-
mentioned synthesis of compound 12 was combined with
each of the b1,3- and b1,4-glycosylation reactions so that
disaccharides 18 and 26 were synthesized in successive oper-
ations starting from compound 10 (Scheme 2B). As a conse-
quence, compound 18 was obtained in 48% yield (6 steps)
and compound 26 was obtained in 54% yield (5 steps).
Furthermore, our synthetic method can also be extended
to the construction of longer-chain oligosaccharides in an ef-
ficient manner by using modular glycosyl donors and accept-
The conversion of b1,3- and b1,4-disaccharides 33 and 34
into non-sulfated (1 and 5) and mono-sulfated compounds
(2–4 and 6–8), respectively, was
easily achieved by using various
combinations of deprotection
and sulfation steps (Scheme 4).
For instance, compound 3,
which contained a sulfate group
at the C6 position of the Gal
moiety was derived from com-
pound 33 through the following
five steps:[22] The treatment of
compound 33 with an 80%
aqueous solution of AcOH at
508C removed the benzylidene
protecting group to afford the
free diol and was followed by
acetylation (in the presence of
pyridine, Ac2O, and a catalytic
amount of dimethylaminopyri-
dine). The next two steps in-
volved cleavage of the silyl
ether (treatment with TBAF)
and
sulfation
by
using
NMe3·SO3. Finally, NaOMe/
MeOH was used to simultane-
ously remove the acetate and
levulinate ester groups to afford
compound 3 in 39% overall
yield in five steps. The other
disaccharides were obtained in
a similar manner. Moreover, it
was not trivial to purify these
final sulfated products. When
using column chromatography
on silica gel, ammonium hy-
droxide or triethylamine was
added to the eluent (CHCl3/
Scheme 3. A) Synthesis of tetrasaccharide 31 from the disaccharide precursor (26). B) Application of disac-
charide donor 29 to the synthesis of heptasaccharide 32.
ors. For example, disaccharide 26 can be converted into
donor 29 and acceptor 30 (Scheme 3A). Triethyl phosphine
(PEt3) was utilized to reduce the azide at the C2 position
and to hydrolyze the anomeric acetate at the same time, fol-
lowed by selective N-trichloroacetylation (to give compound
28) and 1-O-imidation to afford glycosyl donor 29. Notably,
the glycosylation reaction that involved compounds 29 and
MeOH or EtOAc/MeOH) to avoid delayed retention of the
desired product. Excessive salts can be removed by means
of a Zeba spin desalting column. Ion-exchange chromatog-
raphy was performed on a Hitrap Q column to obtain the
product or the desired counterion in high purity.
All of the final products (1–8) were characterized in
1
detail by using several NMR experiments, including H-1H
Chem. Asian J. 2013, 8, 1536 – 1550
1539
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
Scheme 4. Compounds 1–8 were obtained by using different combina-
tions of deprotection and sulfation steps: a) 80% AcOH, 508C; b) 1.0m
TBAF in THF, AcOH, THF, 08C to RT; c) MeONa, MeOH, RT;
d) SO3·TMA, DMF, RT or 508C; e) pyridine, Ac2O, cat. DMAP, 08C to
RT; f) N2H4·AcOH, THF, 08C to RT.
COSY, 1H-13C HMQC, 1D-selective TOCSY, and DEPT
NMR spectroscopy. In general, the anomeric protons of
these products were identified first because they appeared
1
as the most downfield-shifted signals in the H NMR spec-
tra. The HMBC method can be applied to pinpoint the H1
proton of GlcNAc, because only this proton correlates with
those of the 6-azidohexyl group (the reducing-end linker).
1
The subsequent use of H-13C HMQC is useful to not only
confirm the previous assignment but also to identify/distin-
guish between the two anomeric carbon atoms. 1H-1H
COSY, in conjunction with 1D-selective TOCSY, is able to
assign all of the protons (Figure 1A). In particular, for sul-
fated disaccharides 2–4 and 6–8, the desirable position of
each sulfate was rigorously determined. For instance, com-
pound 7 had a sulfate group at the C6 position of the Gal
moiety. The corresponding protons (H6a and H6b) appeared
at d=4.20 and 4.16 ppm, which were 0.48–0.52 ppm down-
field shifted in comparison with those of its non-sulfated an-
alogue (5, Table 2). Likewise, the sulfated carbon atom of
compound 7 had a chemical shift of d=68.1 ppm, which was
5.4 ppm higher than that of compound 5 (Figure 1B and
Table 2). The trend of higher chemical shifts was observed
in all of the sulfated carbon atoms and their adjacent pro-
tons (Figure 1).
Figure 1. A) All of the protons of the Gal and GlcNAc groups in com-
1
pound 7 were assigned by H-1H COSY NMR spectroscopy. The red and
black lines connect the Gal and GlcNAc resonances, respectively. B) A
comparison of the 13C NMR and the corresponding DEPT-135 spectra of
compound 7 (top two spectra) and its non-sulfated analogue (5, bottom
two spectra). Only a region of the spectra is shown to include the signals
of the carbon atoms of the sugar ring.
-8 were utilized as the counterparts because they represent
two main galectin groups, that is, galectin-1, as a prototypical
galectin, whereas galectin-8 is a tandem-repeat galectin.[2]
Both galectins are essential for the differentiation of mature
B cells into immunoglobulin-secreting plasma cells.[24] The
preliminary result of binding analysis showed two interesting
features: First, that the incorporation of a sulfate group at
a specific position of Gal-b1,3/4-GlcNAc increased the bind-
ing of galectin. Compounds 4 and 8, which contained a sul-
fate group at the C3 position of Gal, bound more tightly to
galectin-1 than their respective non-sulfated analogues (1
Furthermore, compounds 1–8 were examined for their
lectin-binding specificities, in which we employed the previ-
ously developed AlphaScreen method, which employed in-
solution proximity binding to photosensitizers,[23] for qualita-
tive measurements (the major reagents were commercially
available from Perkin–Elmer Inc.). Human galectins-1 and
Chem. Asian J. 2013, 8, 1536 – 1550
1540
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
Table 2. Proton (top) and carbon (bottom) chemical shifts of Gal/GlcNAc in the final compounds 1–8.[a]
d (1H NMR) [ppm] of Gal
d (1H NMR) [ppm] of GlcNAc
H1’
4.23
4.28
4.31
4.42
4.33
4.50
4.40
4.49
H2’
3.50
3.54
3.54
3.66
3.49
3.52
3.53
3.72
H3’
3.42
3.57
3.50
4.19
3.43
3.51
3.55
4.24
H4’
3.76
3.81
3.87
4.19
3.76
3.82
3.87
4.22
H5’
3.51
3.50
3.86
3.61
3.53
3.63
3.89
3.64
H6a’
3.72
3.77
4.18
3.75
3.72
3.76
4.20
3.76
H6b’
3.65
3.70
4.17
3.67
3.64
3.69
4.16
3.69
H1
H2
H3
H4
H5
H6a
3.84
4.41
3.88
3.85
3.86
4.36
3.92
3.92
H6b
3.62
4.10
3.73
3.71
3.80
4.30
3.88
3.87
1
2
3
4
5
6
7
8
4.44
4.47
4.45
4.46
4.36
4.41
4.44
4.39
3.66
3.75
3.77
3.75
3.65
3.71
3.75
3.72
3.63
3.70
3.68
3.71
3.58
3.63
3.63
3.64
3.37
3.62
3.40
3.40
3.56
3.62
3.61
3.62
3.26
3.55
3.32
3.30
3.35
3.63
3.43
3.40
d (13C NMR) [ppm] of Gal
d (13C NMR) [ppm] of GlcNAc
C1’
C2’
72.5
72.5
72.3
70.7
728
73.0
72.5
71.1
C3’
74.8
74.8
74.5
81.7
75.0
75.0
74.7
82.0
C4’
70.4
70.4
74.8
68.7
70.5
70.6
70.1
68.7
C5’
77.3
77.7
70.0
76.8
77.3
77.2
74.9
77.0
C6’
62.6
62.6
67.9
62.5
62.7
62.6
68.1
62.6
C1
C2
C3
C4
C5
C6
1
2
3
4
5
6
7
8
105.7
105.7
105.7
105.2
105.3
104.9
105.7
105.0
102.5
102.5
102.6
102.6
102.9
102.8
102.6
103.0
56.5
56.5
56.3
56.2
56.9
56.9
56.7
56.8
85.1
85.1
85.2
84.9
74.3
74.4
75.2
74.4
70.7
70.7
71.0
70.6
81.3
80.7
81.8
81.4
77.7
77.2
77.5
77.6
76.7
74.7
76.5
76.7
62.9
68.7
63.0
62.8
62.2
67.7
62.1
62.1
1
[a] The assignments of the chemical shifts are based on H, 13C, DEPT-90, DEPT-135, HMQC, HMBC, and 1D-selective TOCSY NMR experiments. The
chemical shifts shown in bold and underlined denote the signals of sulfated carbon or proton atoms adjacent to the sulfate groups.
construct longer glycans by modular assembly. This method
could potentially be used for the synthesis of other sulfated
oligosaccharides, such as keratan sulfate oligosaccharides,
which are glycosaminoglycans and contain sulfated N-acetyl-
lactosamine-repeating disaccharides. The method could also
be applied to the preparation of uronic-acid-containing gly-
cans if the C6-protecting group on Gal or GlcNAc can be
selectively removed and oxidized. We are currently prepar-
ing other sulfated oligosaccharides and analogues for com-
parison of their galectin-binding specificities.
Figure 2. Relative binding specificities of compounds 1–8 with galectins-
1 and -8. The results were averaged from at least three independent
assays. A blank experiment was performed in the absence of the synthe-
sized product.
Experimental Section
General Considerations
All of the reactions were performed under a nitrogen atmosphere in
oven-dried glassware unless otherwise indicated. The reaction products
and 5, respectively; Figure 2). Moreover, galectins-1 and -8
exhibited dissimilar binding patterns towards disaccharides
were purified by using column chromatography on silica gel.[26] Anhy-
drous solvents and moisture-sensitive materials were transferred by using
1–8. Specifically, galectin-8 preferentially bound to b1,3-
linked disaccharides. Our results are consistent with previ-
ous reports,[25] including data that were released by the Con-
sortium for Functional Genomics (CFG)[25b] and those ob-
tained by using a flow cytometry study in which splenic B
cells were stained with galectin-1-FITC or galectin-8-FITC
and then treated with compounds 1–8.[24a]
an oven-dried syringe or cannula through a rubber septum. Organic solu-
tions were concentrated under reduced pressure in a water bath (<
408C). Analytical TLC was performed on pre-coated SiliaFlash P60 glass
plates (230–400 mesh, 250 mm thick, Silicyle) and was detected by UV
visualization (254 nm) and/or by staining with reagents that contained
phosphomolybdic acid hydrate (for general use), para-anisaldehyde (for
carbohydrates), or ninhydrin (for amino-group-containing samples).
Column chromatography on silica gel was performed on SiliaFlash G60
(70–230 mesh) or SiliaFlash P60 (230–400 mesh) from Silicyle. Purifica-
tion of the bulk building blocks were either performed on a Teledyne
Isco CombiFlash Rf that was equipped with a UV detector (for com-
pounds with UV-active chromophores) or on a Grace Davison Reveleris
flash system that was equipped with an Evaporative Light Scattering De-
tector (ELSD, for compounds without UV-active chromophores). Gel-fil-
tration chromatography (Sephadex LH-20, G-10 and G-25), ion-exchange
chromatography (GE HiTrap Q HP Column), and spin desalting column
chromatography (Thermo Zeba spin desalting column, 7K MWCO) were
Conclusions
Herein, we have presented an efficient procedure for the
preparation of protected Gal-b1,3/4-GlcNAc disaccharides.
Both saccharide precursors were used to systematically in-
troduce sulfate groups at specific positions and to rapidly
1
used to achieve satisfactory purities of the final disaccharide products. H
Chem. Asian J. 2013, 8, 1536 – 1550
1541
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
and 13C NMR spectra were recorded on Bruker AV-400 (400 MHz) or
AVII-500 (500 MHz) spectrometers by using tetramethylsilane (dH =
0.00 ppm), CDCl3 (dH =7.26 ppm), or CD3OD (dH =3.31 ppm, central
line of a quintet) as internal standards. 13C NMR spectra were recorded
on Bruker AV-400 (100 MHz) or AVII-500 (125 MHz) spectrometers by
using CDCl3 (dC =77.2 ppm, central line of a triplet) or CD3OD (dC =
49.2 ppm, central line of a septet) as internal standards. 2D NMR spectra
(including 1H-1H COSY, 1H-13C HMQC, and NOESY) were acquired on
Bruker AV-400 or AVII-500 spectrometers. High-resolution mass spec-
troscopy (HRMS) was performed on Bruker Bio-TOF III (ESI-TOF) or
Bruker Ultraflex (MALDI-TOF/TOF) spectrometers and is reported as
mass/charge (m/z) ratios with percentage relative abundance. Optical ro-
tations were measured at the sodium D-line (589 nm) at 258C on a Perki-
6-Azidohexyl (6-O-sulfonato-b-d-galactopyranosyl)-(1!3)-2-acetamido-2-
deoxy-b-d-glucopyranoside Sodium Salt (3)
The following four paragraphs describe the detailed procedure for the
synthesis of compound 3 from compound 33. For the structures of these
compounds, see the Supporting Information, Scheme 5.
To a solution of compound 33 (250 mg, 0.25 mmol) was added an 80%
aqueous solution of HOAc (5 mL). The resulting mixture was heated at
508C for 2 h and then co-evaporated with toluene three times to give
a white solid. The solid was dissolved in anhydrous pyridine (3 mL) at
08C under a nitrogen atmosphere and treated with Ac2O (1 mL) and
a catalytic amount of DMAP (5 mg). After removing the ice bath, the re-
action mixture was stirred at RT for 4 h, diluted with EtOAc, washed
with a 1m aqueous solution of HCl (three times), a saturated aqueous so-
lution of NaHCO3, water, and brine, dried over MgSO4, filtered, and con-
centrated in vacuo. The residue was purified by column chromatography
on silica gel (EtOAc/hexanes, 1:1) to provide compound 35 (197 mg,
79% yield over two steps) as a white amorphous foam.
25
nElmer Model 341 Polarimeter. Specific rotations were reported as ½aꢁD
after dividing the observed values by the sample concentration (C, in
gmLꢀ1) and the path length (l, in dm).
The solvents for extraction and chromatography were of ACS grade.
CH2Cl2, MeCN, and THF were pre-dried by using molecular sieves and
then percolated through an active Al2O3 column.[27] Anhydrous DMF and
MeOH were purchased from Acros Co. in an AcroSeal package. Diphe-
nylsulfoxide (Ph2SO), 2,4,6-tri-tert-butylpyrimidine, and triethylphosphine
(1.0m in THF) were purchased from Aldrich Chemical Co. All of the
chemicals were used without further purification unless otherwise speci-
fied. Celite 545 and 3 ꢁ molecular sieves (powder<50 mm) were pur-
chased from Acros Co. 1,2,3,4,6-Penta-O-acetyl-b-d-galactopyranoside,
d-glucosamine hydrochloride, and 1-ethyl-3-(3-dimethylaminopropyl)car-
bodiimide hydrochloride (EDC) were purchased from Carbosynth Ltd.
in a bulk package. Tetra-n-butylammonium fluoride (TBAF, 1.0m in
THF) was purchased from Alfa Aesar.
To a solution of compound 35 (150 mg, 0.15 mmol) in anhydrous THF
(3 mL) were sequentially added HOAc (25.5 mL, 0.45 mmol) and TBAF
(1.0m in THF, 0.45 mL, 0.45 mmol) at 08C under a nitrogen atmosphere.
The reaction mixture was gradually warmed to RT and stirred for 15 h.
The reaction mixture was diluted with EtOAc and then washed with a sa-
turated aqueous solution of NaHCO3, water, and brine. The organic layer
was dried over anhydrous MgSO4, filtered, and concentrated under re-
duced pressure to give a dry residue that was purified by column chroma-
tography on silica gel (EtOAc) to afford compound 36 (83.3 mg, 73%
yield) as a white amorphous solid.
Compound 36 (70.0 mg, 90.3 mmol) was dissolved in anhydrous DMF
(2 mL) and treated with sulfur trioxide trimethylamine complex
(SO3·TMA, 62.9 mg, 0.45 mmol) at RT under a nitrogen atmosphere. The
reaction was heated at 508C for 3 h, stopped by the addition of MeOH at
RT, and co-evaporated with toluene three times. The resulting residue
was dissolved in MeOH (5 mL) and subjected to ion exchange [Na+] on
the IR-120 resin to generate the sodium salt. After the filtration, the fil-
trate was concentrated under reduced pressure and purified by column
chromatography on silica gel (MeOH/CHCl3, 1:5) to give compound 37
(63.4 mg, 80% yield) as a white amorphous foam.
General Procedure for Measuring the Binding Affinities between Synthetic
Sulfated Sugars and Galectins-1 and -8
The binding affinity of the synthesized sugars to galectins-1 and -8 was
measured by using a previously reported AlphaScreen-based solution car-
bohydrate array.[23] The measurements were performed on a PerkinElmer
Envision instrument. The donor beads and acceptor beads were pur-
chased from PerkinElmer Life Sciences, Inc. (Boston, MA, USA). Re-
combinant galectin-1 and galectin-8 were prepared according to a litera-
ture procedure.[24b]
Compound 37 (50 mg, 57.0 mmol) was dissolved in MeOH (2 mL) and
treated with a catalytic amount of NaOMe (1 mg) at RT under a nitrogen
atmosphere. After stirring for 2.5 h, the reaction was neutralized by the
addition of Amberlite IR-120 [H+] resin and then filtered. The filtrate
was concentrated in vacuo and purified by column chromatography on
silica gel (EtOAc/MeOH/H2O, 8:2:1 with 1% Et3N) to afford the product
as a white foam. Further ion-exchange chromatography was performed
on a GE HiTrap Q HP column (gradient elution with NH4HCO3) to give
the final product (3, 29.6 mg, 85% yield) as white powder. Rf =0.70 (IPA/
H2O/NH4OH, 7:2:1 v/v/v); ½aꢁ2D5 =ꢀ2.4 (c=1.3, MeOH); 1H NMR
(500 MHz, CD3OD): d=4.46 (d, J=8.4 Hz, 1H), 4.32 (d, J=7.3 Hz, 1H),
4.18 (d, J=6.2 Hz, 1H), 4.91–4.85 (m, 4H), 3.79–3.67 (m, 3H), 3.57–3.47
(m, 3H), 3.40 (t, J=9.5 Hz, 1H), 3.35–3.32 (m, 1H), 3.28 (t, J=6.9 Hz,
CH2N3, 2H), 1.98 (s, 3H; COCH3), 1.60–1.57 (m, 4H; alkyl chain), 1.40–
1.38 ppm (m, 4H; alkyl chain); 13C NMR (125 MHz, CD3OD): d=174.3
(C=O), 105.7 (CH), 102.6 (CH), 85.2 (CH), 77.5(CH), 74.8 (CH), 74.5
(CH), 72.3 (CH), 71.0 (CH), 70.6 (CH2), 70.0 (CH), 67.9 (CH2), 63.0
(CH2), 56.3 (CH), 52.5 (CH2), 30.6 (CH2), 30.0 (CH2), 27.6 (CH2), 26.8
(CH2), 23.4 ppm (CH3); HRMS (ESI-TOF): m/z calcd for
C20H35N4NaO14SNa: 633.1660 [M+Na]+; found: 633.1678.
6-Azidohexyl (b-d-galactopyranosyl)-(1!3)-2-acetamido-2-deoxy-b-d-
glucopyranoside (1)
White amorphous solid in 54% overall yield (3 steps from compound
32). Rf =0.55 (IPA/H2O/NH4OH, 7:2:1 v/v/v); ½aꢁ2D5 =+1.1 (c=0.3,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.47 (d, J=8.0 Hz, 1H), 4.27
(d, J=7.5 Hz, 1H), 3.91–3.88 (m, 2H), 3.80 (d, J=2.7 Hz, 1H), 3.78–3.65
(m, 6H), 3.56–3.52 (m, 2H), 3.50–2.40 (m, 3H), 3.28 (t, J=7.0 Hz, 2H;
CH2N3), 1.96 (s, 3H; COCH3), 1.59–1.57 (m, 4H; alkyl chain), 1.41–
1.39 ppm (4H, m; alkyl chain); 13C NMR (125 MHz, CD3OD): d=174.3,
105.8, 102.5, 85.1, 77.7, 77.3, 74.8, 72.5, 70.7, 70.6, 70.4, 62.8, 62.7, 56.5,
52.5, 30.6, 30.1, 27.7, 26.8, 23.3 ppm; HRMS (ESI-TOF): m/z calcd for
C20H36N4O11Na: 531.2273 [M+Na]+; found: 531.2265.
6-Azidohexyl (b-d-galactopyranosyl)-(1!3)-2-acetamido-2-deoxy-6-O-
sulfonato-b-d-glucopyranoside Sodium Salt (2)
White amorphous solid in 37% overall yield (4 steps from compound
32). Rf =0.70 (IPA/H2O/NH4OH, 7:2:1 v/v/v); ½aꢁ2D5 =ꢀ2.9 (c=0.7,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.50 (d, J=8.2 Hz, 1H), 4.43
(dd, J=10.8, 1.9 Hz, 1H), 4.33 (d, J=7.4 Hz, 1H), 4.13 (dd, J=10.8,
6.3 Hz, 1H), 3.91–3.87 (m, 1H), 3.83–3.77 (m, 3H), 3.75–3.59 (m, 5H),
3.56–3.49 (m, 3H), 3.43 (dd, J=9.7, 8.2 Hz, 1H), 3.30 (t, J=6.7 Hz, 2H;
CH2N3), 2.01 (s, 3H; COCH3), 1.62–1.58 (m, 4H; alkyl chain), 1.42–
1.40 ppm (m, 4H; alkyl chain); 13C NMR (125 MHz, CDCl3): d=174.5,
105.6, 102.4, 84.8, 77.2, 75.7, 74.8, 72.5, 70.8, 70.7, 70.5, 68.6, 62.7, 56.5,
52.5, 30.6, 30.0, 27.6, 26.8, 23.2 ppm; HRMS (ESI-TOF): m/z calcd for
C20H35N4NaO14SNa: 633.1660 [M+Na]+; found: 633.1655.
6-Azidohexyl (3-O-sulfonato-b-d-galactopyranosyl)-(1!3)-2-acetamido-2-
deoxy-b-d-glucopyranoside Sodium Salt (4)
White amorphous solid in 26% overall yield (6 steps from compound
32). Rf =0.70 (IPA/H2O/NH4OH, 7:2:1 v/v/v); ½aꢁ2D5 =ꢀ33 (c=0.1,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.70 (t, J=4.0 Hz, 1H), 4.40
(dd, J=10.0, 7.5 Hz, 1H), 4.30 (t, J=9.6 Hz, 1H), 3.96 (d, J=8.2 Hz,
1H), 3.94 (d, J=7.5 Hz, 1H), 3.91 (dd, J=10.0, 4.0 Hz, 1H), 3.85 (dd, J=
10.0, 8.0 Hz, 1H), 3.84–3.82 (m, 3H), 3.82–3.80 (m, 1H), 3.79 (dd, J=9.0,
8.2 Hz, 1H), 3.77–3.75 (m, 1H), 3.70 (t, J=6.5 Hz, 2H), 3.58 (m, 1H),
Chem. Asian J. 2013, 8, 1536 – 1550
1542
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
3.34 (t, J=7.1 Hz, 1H), 3.32 (t, J=7.1 Hz, 1H), 2.04 (s, 3H; COCH3),
1.63 (m, 4H; alkyl chain), 1.45 ppm (m, 4H; alkyl chain); 13C NMR
(125 MHz, CD3OD): d=172.8, 103.7, 101.1, 83.4, 80.1, 76.0, 75.3, 69.2,
69.1, 67.1, 61.3, 61.0, 54.7, 51.0, 47.9, 47.8, 47.6, 47.5, 47.3, 47.1, 29.0, 28.5,
26.1, 25.2, 21.8 ppm; HRMS (MALDI-TOF): m/z calcd for
C20H35N4NaO14SNa: 633.1660 [M+Na]+; found: 633.1619.
4-Methylphenyl 2,3,4,6-tetra-O-acetyl-1-thio-b-d-galactopyranoside (9)[28]
To a solution of 1,2,3,4,6-penta-O-acetyl-b-d-galactopyranoside (50.0 g,
128 mmol) and para-toluenethiol (19.3 g, 154 mmol) in anhydrous CH2Cl2
(300 mL) at 08C under a nitrogen atmosphere was added dropwise
BF3·OEt2 (32.6 mL, 256 mmol). The reaction mixture was gradually
warmed to RT and stirred for 16 h. The color of the solution changed to
red when the reaction was complete. The reaction was quenched by the
portion-wise addition of a saturated aqueous solution of NaHCO3 at 08C
with vigorous stirring, washed with water and brine (three times each),
and dried over MgSO4. The filtrate was evaporated under reduced pres-
sure to give a dry residue, which was purified by recrystallization from
hot EtOH to give the desired product (9, 43.3 g, 86% yield) as a white
solid. Rf =0.50 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =+0.6 (c=1.3, CHCl3);
1H NMR (500 MHz, CDCl3): d=7.41 (d, J=8.1 Hz, 2H; ArH), 7.13 (d,
J=8.1 Hz, 2H; ArH), 5.40 (d, J=3.3 Hz, 1H; H4), 5.21 (t, J=10.0 Hz,
1H; H2), 5.04 (dd, J=9.9, 3.4 Hz, 1H; H3), 4.65 (d, J=10.0 Hz, 1H;
H1), 4.18 (dd, J=11.3, 6.9 Hz, 1H; H6a), 4.11 (dd, J=11.4, 6.4 Hz, 1H;
H6b), 3.91 (t, J=6.6 Hz, 1H; H5), 2.35 (s, 3H; PhCH3), 2.11 (s, 3H;
COCH3), 2.10 (s, 3H; COCH3), 2.04 (s, 3H; COCH3), 1.97 ppm (s, 3H;
COCH3); 13C NMR (100 MHz, CDCl3): d=170.4, 170.2, 170.1, 169.4,
138.5, 133.2, 129.6, 128.6, 87.0, 74.4, 72.1, 67.3, 67.3, 61.6, 21.1, 20.8, 20.65,
20.60, 20.56 ppm; HRMS (ESI-TOF): m/z calcd for C21H26O9SNa:
477.1190 [M+Na]+; found: 477.1199.
6-Azidohexyl (b-d-galactopyranosyl)-(1!4)-2-acetamido-2-deoxy-b-d-
glucopyranoside (5)
White amorphous solid in 67% overall yield (2 steps from compound
33). Rf =0.60 (IPA/H2O/NH4OH=7:2:1 v/v/v); ½aꢁ2D5 =ꢀ3.0 (c=0.7,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.40 (d, J=8.4 Hz, 1H), 4.38
(d, J=8.1 Hz, 1H), 3.92–3.86 (m, 3H), 3.81 (d, J=2.4 Hz, 1H), 3.78–3.64
(m, 4H), 3.62–3.52 (m,4H), 3.45–3.44 (m, 2H), 3.40–3.38 (m, 1H), 3.28
(t, J=6.9 Hz, 2H; CH2N3), 1.97 (s, 3H; COCH3), 1.59–1.56 (m, 4H; alkyl
chain), 1.39–1.38 ppm (m, 4H; alkyl chain); 13C NMR (125 MHz,
CD3OD): d=173.6, 105.3, 102.9, 81.2, 77.3, 76.7, 75.0, 74.4, 72.8, 70.6,
70.5, 62.7, 62.1, 56.9, 52.6, 30.6, 30.1, 27.2, 28.8, 23.1 ppm; HRMS (ESI-
TOF): m/z calcd for C20H36N4O11Na: 531.2273 [M+Na]+; found: 531.2280.
6-Azidohexyl (b-d-galactopyranosyl)-(1!4)-2-acetamido-2-deoxy-6-O-
sulfonato-b-d-glucopyranoside Sodium Salt (6)
White amorphous solid in 34% overall yield (3 steps from compound
33). Rf =0.70 (IPA/H2O/NH4OH, 7:2:1 v/v/v); ½aꢁ2D5 =ꢀ1.2 (c=0.4,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.51 (d, J=7.5 Hz, 1H), 4.43
(d, J=8.3 Hz, 1H), 4.37–4.30 (m, 2H), 3.88–3.83 (m, 2H), 3.79–3.61 (m,
8H), 3.54–3.46 (m, 3H), 3.28 (t, J=6.8 Hz, 2H; CH2N3), 2.01 (s, 3H;
COCH3), 1.60–1.55 (m, 4H; alkyl chain), 1.40–1.38 ppm (m, 4H; alkyl
chain); 13C NMR (125 MHz, CDCl3): d=174.1, 104.8, 102.6, 80.6, 77.2,
74.9, 74.7, 74.3, 72.9, 70.7, 70.6, 67.6, 62.7, 57.0, 52.5, 30.6, 30.0, 27.6, 26.7,
22.9 ppm; HRMS (ESI-TOF): m/z calcd for C20H35N4NaO14SNa:
633.1660 [M+Na]+; found: 633.1669.
4-Methylphenyl 6-O-tert-butyldiphenylsilyl-1-thio-b-d-galactopyranoside
(10)[29]
For details of the reactions, see the Supporting Information, Scheme 1.
To a solution of thiogalactoside 9 (10.0 g, 22.0 mmol) in anhydrous
MeOH/CH2Cl2 (1:1, 200 mL) was added a catalytic amount of NaOMe
(120 mg, 10 mol% yield). The reaction mixture was stirred at RT for 1 h
under a nitrogen atmosphere. Amberlite IR-120 [H+] resin was added to
quench the reaction and to adjust the pH value to 6. The resulting mix-
ture was filtered, concentrated, and co-evaporated with toluene three
times. The resulting white solid was dissolved in anhydrous DMF
(200 mL) and imidazole (3.00 g, 44.0 mmol) at 08C, followed by the addi-
tion of TBDPSCl (6.9 mL, 26.4 mmol). The mixture was stirred at RT for
a further 2.5 h under a nitrogen atmosphere. The reaction was quenched
by the addition of MeOH (10 mL). After stirring at RT for 10 min, the
solvent was removed under reduced pressure to give a dry residue that
was purified by column chromatography (EtOAc/hexanes=2:1 v/v) to
give compound 10 as a white solid (10.6 g, 92% yield). Rf =0.50 (EtOAc/
hexanes, 2:1 v/v); ½aꢁD25 =ꢀ14 (c=1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): d=7.72 (d, J=6.8 Hz, 2H; ArH), 7.69 (d, J=6.6 Hz, 2H; ArH),
7.45–7.37 (m, 8H; ArH), 7.06 (d, J=7.9 Hz, 2H; ArH), 4.43 (d, J=
9.1 Hz, 1H; H1), 4.13–3.99 (m, 1H; H4), 3.96–3.94 (m, 2H; H6a,b), 3.64–
3.54 (m, 3H; H2, H3, H5), 2.81 (br, 1H; OH), 2.64 (d, J=5.5 Hz, 1H;
OH), 2.47 (br, 1H; OH), 2.32 (s, 3H; PhCH3), 1.07 ppm (s, 9H; tBu);
13C NMR (CDCl3, 100 MHz): d=138.2, 135.7, 135.6, 133.0, 132.9, 132.7,
129.9, 129.8, 128.3, 127.8, 88.8, 78.1, 74.9, 69.9, 69.4, 63.8, 26.8, 21.1,
19.1 ppm; HRMS (ESI-TOF): m/z calcd for C29H36O5SSiNa: 547.1945
[M+Na]+; found: 547.1930.
6-Azidohexyl (6-O-sulfonato-b-d-galactopyranosyl)-(1!4)-2-acetamido-2-
deoxy-b-d-glucopyranoside Sodium Salt (7)
White amorphous solid in 55% overall yield (3 steps from compound
33). (IPA/H2O/NH4OH=7/2/1 v/v/v) Rf =0.70; ½aꢁ2D5 =ꢀ1.3 (c=0.9,
MeOH); 1H NMR (500 MHz, CD3OD): d=4.44 (d, J=8.4 Hz, 1H; H1),
4.40 (d, J=7.2 Hz, 1H; H1’), 4.20 (dd, J=11.0, 7.7 Hz, 1H; H6a’) 4.16
(dd, J=11.0, 5.0 Hz, 1H; H6b’), 3.93–3.86 (m, H4’, H6ab, 4H; OCH2),
3.77–3.71 (m, 1H; H2), 3.64–3.60 (m, 2H; H3, H4), 3.57–3.53 (m, 2H;
H3’, H2’), 3.50 (dt, J=9.7, 6.6 Hz, 1H; OCH2), 3.44–3.42 (m, 1H; H5),
3.28 (t, J=6.9 Hz, 2H; CH2N3), 1.99 (s, 3H; NCOCH3), 1.60–1.56 (m,
4H; alkyl chain), 1.40–1.36 ppm (m, 4H; alkyl chain); 13C NMR
(125 MHz, CDCl3): d=174.2 (C=O), 105.3 (CH), 102.6 (CH), 81.8 (CH),
76.4 (CH), 75.2 (CH), 74.9 (CH), 74.7 (CH), 72.5 (CH), 70.7 (CH2), 70.1
(CH), 68.1 (CH2), 62.1 (CH2), 56.7 (CH), 52.5 (CH2), 30.6 (CH2), 30.0
(CH2), 27.7 (CH2), 26.8 (CH2), 23.2 ppm (CH3); HRMS (ESI-TOF): m/z
calcd for C20H35N4NaO14SNa [M+Na]+ 633.1660; found: 633.1679.
6-Azidohexyl (3-O-sulfonato-b-d-galactopyranosyl)-(1!4)-2-acetamido-2-
deoxy-b-d-glucopyranoside Sodium Salt (8)
White amorphous solid in 34% overall yield (4 steps from compound
4-Methylphenyl 2,4-di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-
b-d-galactopyranoside (12)
33). Rf =0.20 (EtOAc/MeOH/H2O, 8:2:1 v/v/v); ½aꢁ2D5 =ꢀ36 (c=1.0,
1
MeOH); H NMR (500 MHz, CD3OD): d=4.49 (d, J=7.8 Hz, 1H; H1’),
To a solution of 2,3,4-triol 10 (5.0 g, 9.53 mmol) and a catalytic amount of
CSA (221 mg, 0.95 mmol) in MeCN (100 mL) at RT under a nitrogen at-
mosphere was added trimethyl orthoacetate (3.7 mL, 28.6 mmol). After
stirring for 30 min, Et3N was added to quench the reaction and the reac-
tion mixture was dried under reduced pressure. After dissolving in
CH2Cl2 (100 mL), to the resulting solution (which contained the O3,O4-
orthoformate product) were sequentially added Et3N (4.0 mL,
28.6 mmol), acetic anhydride (1.8 mL, 19.1 mmol), and DMAP (116 mg,
0.95 mmol). After stirring for 30 min, the solvent was removed under re-
duced pressure to give a dry residue and then poured into an 80% aque-
ous solution of AcOH (100 mL) with vigorous stirring for 30 min. The
solvent was removed by evaporation and the residue was extracted with
EtOAc. The collected organic layer was washed with an ice-cold saturat-
ed solution of aqueous NaHCO3, water, and brine, and dried over
4.39 (d, J=8.4 Hz, 1H; H1), 4.24 (dd, J=9.7, 3.2 Hz, 1H; H3’), 4.22 (d,
J=3.1 Hz, 1H; H4’), 3.92 (dd, J=12.1, 2.6 Hz, 1H; H6a), 3.91–3.85 (m,
2H; H6b, OCH2), 3.76 (dd, J=11.6, 7.6 Hz, 1H; H6a’), 3.73–3.67 (m,
3H; H2, H2’, H6b’), 3.65–3.59 (m, 3H; H3, H5’, H4), 3.47 (dt, J=9.7,
6.6 Hz, 1H; OCH2), 3.40–3.38 (m, 1H; H5), 3.28 (t, J=6.9 Hz, 2H;
CH2N3), 1.97 (s, 3H; NCOCH3), 1.60–1.53 (m, 4H), 1.42–1.38 ppm (m,
4H); 13C NMR (125 MHz, CD3OD): d=173.6 (C=O), 105.0 (CH), 103.0
(CH), 82.0 (CH), 81.4 (CH), 77.0 (CH), 76.7 (CH), 74.4 (CH), 71.1 (CH),
70.6 (CH2), 68.7 (CH), 62.6 (CH2), 62.1 (CH2), 56.8 (CH), 52.6 (CH2),
30.6 (CH2), 30.0 (CH2), 27.7 (CH2), 26.8 (CH2), 23.1 ppm (CH3); HRMS
(ESI-TOF): m/z calcd for C20H35N4NaO14SNa: 633.1660 [M+Na]+; found:
633.1619.
Chem. Asian J. 2013, 8, 1536 – 1550
1543
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
MgSO4. The filtrate was evaporated under reduced pressure. To a solution
that contained the resulting product (compound 11) in anhydrous CH2Cl2
(100 mL) and levulinic acid (1.66 g, 14.3 mmol) were added EDC (2.74 g,
14.3 mmol) and 4-dimethylaminopyridine (DMAP, 116 mg, 0.95 mmol) at
08C under a nitrogen atmosphere. The ice bath was removed after the
addition had been completed and the mixture was stirred at RT for 1 h,
evaporated, redissolved in EtOAc, washed with a saturated aqueous solu-
tion of NaHCO3, water, and brine and dried over anhydrous MgSO4. The
filtrate was concentrated under reduced pressure to give a dry residue
that was purified by flash column chromatography (EtOAc/hexanes, 1:2
v/v) to afford compound 12 (5.25 g, 78% yield) as a white amorphous
foam. Rf =0.45 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =+1.6 (c=2.5, CHCl3);
1H NMR (500 MHz, CDCl3): d=7.63–7.60 (m, 4H; ArH), 7.59–7.35 (m,
8H; ArH), 7.06 (d, J=7.8 Hz, 2H; ArH), 5.51 (d, J=2.3 Hz, 1H; H4),
5.18 (t, J=9.9 Hz, 1H; H2), 5.06 (dd, J=9.9, 2.4 Hz, 1H; H3), 4.64 (d,
J=9.9 Hz, 1H; H1), 3.78–3.77 (m, 2H; H5, H6a), 3.68–3.63 (m, 1H;
H6b), 2.79–2.73 (m, 1H; Lev-CH2), 2.63–2.51 (m, 2H; Lev-CH2), 2.41–
2.37 (m, 1H; Lev-CH2), 2.31 (s, 3H; PhCH3), 2.15 (s, 3H; COCH3), 2.12
(s, 3H; COCH3), 1.99 (s, 3H; COCH3), 1.02 ppm (s, 9H; tBu); 13C NMR
(125 MHz, CDCl3): d=206.2, 171.6, 171.1, 170.1, 169.7, 138.1, 135.6,
132.9, 132.8, 132.7, 129.8, 129.7, 129.6, 129.1, 127.9, 127.8, 127.5, 87.2,
72.6, 67.4, 67.3, 61.5, 60.4, 51.8, 37.9, 37.6, 36.6, 29.8, 29.6, 27.8, 26.7, 24.7,
21.1, 21.0, 20.8, 20.6, 19.0, 14.7 ppm; HRMS (ESI-TOF): m/z calcd for
C38H46O9SSiNa: 729.2524 [M+Na]+; found: 729.2529.
1.7H (a); H2), 3.37 (dd, J=9.5, 7.7 Hz, 1.7H (b); H2), 3.03 (d, J=
2.6 Hz, 1H (a); OH), 2.78 (d, J=2.3 Hz, 1H (a); OH), 2.76 ppm (d, J=
2.3 Hz, 1.7H (b); OH); HRMS (ESI-TOF): m/z calcd for C13H15N3O5Na:
316.0904 [M+Na]+; found: 316.0915.
(2,4-Di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-b-d-
galactopyranosyl)-(1!3)-2-azido-4,6-O-benzylidene-2-deoxy-ab-d-
glucopyranoside (17)
A
solution of thiogalactoside 12 (200 mg, 0.28 mmol) in anhydrous
CH2Cl2/toluene (2:1, 9 mL) was mixed with 2,4,6-tri-tert-butylpyrimidine
(TTBP, 73.0 mg, 0.29 mmol) and diphenylsulfoxide (Ph2SO, 62 mg,
0.31 mmol). The reaction mixture was stirred in the presence of 3 ꢁ mo-
lecular sieves (400 mg, freshly dried prior to use) at RT for 30 min under
a nitrogen atmosphere and subjected to the addition of Tf2O (51 mL,
0.31 mmol) after cooling to ꢀ608C. Upon consumption of the glycosyl
donor (by TLC, 10 min), a solution of glycosyl acceptor 13 (123 mg,
0.42 mmol) in anhydrous CH2Cl2/MeCN (2:1, 9 mL) was added to the re-
action mixture. After stirring at ꢀ608C for a further 1 h, the reaction was
stopped with the addition of Et3N and filtered through a pad of Celite.
The filtrate was concentrated in vacuo to give a crude residue, which was
purified by flash column chromatography (EtOAc/hexanes, 1:2) to give
compound 17 (213 mg, 86% yield) as an amorphous foam (H1-anomeric
isomers, H1a/H1b 1:1.1). Rf =0.37 (EtOAc/hexanes=1:1 v/v); ½aꢁ2D5 =ꢀ18
(c=0.5, CHCl3); 1H NMR (500 MHz, CDCl3): d=7.54 (d, J=7.3 Hz,
4.2H; ArH
16.8H; ArH
4.2H; ArH(a+b)), 5.55 (s, 2.1H; H4’
5.31 (br, 1H; H1(a)), 5.23 (t, J=8.6 Hz, 2.1H; H2’
10.6 Hz, 2.1H; H3’(a+b)), 4.77 (d, J=8.6 Hz, 1H; H1’(a)), 4.75 (d, J=
8.6 Hz, 1.1H; H1’(b)), 4.69 (t, J=6.7 Hz, 1.1H; H1(b)), 4.27 (dd, J=10.0,
4.3 Hz, 1.1H; H6a(b)), 4.22 (dd, J=10.0, 3.8 Hz, 1H; H6a(a)), 4.15 (t,
J=10.0 Hz, 1H; H3(a)), 4.09–4.01 (m, 1H; H5(a)), 3.75–3.71 (m, 2.1H;
G
ACHTUNGTREN(NUNG a+b)), 7.42–7.33 (m,
2-Azido-4,6-O-benzylidene-2-deoxy-ab-d-glucopyranoside (13)[17]
R
ACHTUNGTRENNUNG
G
R
ACHTUNGTRENNUNG
For details of the reactions, see the Supporting Information, Scheme 2.
AHCTUNGTRENNUNG
In situ preparation of TfN3: To a vigorously stirring solution of sodium
azide (NaN3, 98 g, 1.50 mol) in H2O/CH2Cl2 (1:1, 300 mL) was dropwise
added trifluoromethanesulfonic anhydride (Tf2O, 50 mL, 0.30 mol) at
08C over a period of 30 min. The resulting mixture turned into white
cloudy solution and was stirred at 08C for a further 1.5 h. The organic
layer was washed with water and a saturated aqueous solution of Na2CO3
was added until the pH value of the aqueous layer was above 7. The re-
sulting solution of TfN3 (in about 250 mL of CH2Cl2) was directly used in
the next step without further purification.
CuII-catalyzed diazo transfer: The freshly prepared solution of TfN3 (in
CH2Cl2) was added to a solution of d-glucosamine hydrochloride (33 g,
0.15 mol), potassium carbonate (25 g, 0.18 mol), and a catalytic amount
of CuSO4·5H2O (0.38 g, 1 mol% yield) in water (100 mL) at 08C. Then,
the ice bath was removed and MeOH (400 mL) was added to make the
reaction homogeneous. After vigorous stirring at RT for 18 h, the mixture
was passed through a pad of Celite and the filtrate was concentrated
under reduced pressure to give a dry residue that was purified by column
chromatography on silica gel (acetone) to give the azide product.
AHCTUNGTRENNUNG
H5’
ACHTUNGTRENNUNG
H6b(a)), 3.66–3.59 (m, 4.3H; H3(b), H4(b), H6ab’AHCTUNGTRENNUNG
9.9 Hz, 1H; H4(a)), 3.51 (br, 1.1H; OH(b)), 3.38–3.31 (m, 3.2H; H5(b),
H2(b), H2(a)), 2.99 (br, 1H; OH(a)), 2.80–2.75 (m, 2.1H; Lev-CH2
A
(a+b)), 2.42–2.39 (m, 2.1H; Lev-
3A(a+b)),
E
N
CHTUNGTRENNUNG
AHCTUNGTRENNUNG
172.0, 170.4, 170.23, 170,21, 170.14, 137.13, 137.06, 135.97, 135.95, 135.76,
133.03, 133.01, 132.8, 130.09, 130.02, 129.2, 128.3, 128.0, 125.91, 125.86,
101.76, 101.72, 101.3, 101.1, 97.1, 93.1, 80.4, 79.7, 79.3, 77.4, 77.25, 77.23,
73.2, 73.1, 71.9, 71.8, 69.70, 69.66, 68.9, 68.5, 67.7, 66.91, 66.87, 66.7, 63.8,
62.9, 60.6, 60.5, 37.9, 29.9, 28.1, 26.9, 21.0, 20.8, 19.11, 19.08 ppm; HRMS
(ESI-TOF): m/z calcd for C44H53N3O14SiNa: 898.3189 [M+Na]+; found:
898.3194.
4,6-Benzylidene protection: Benzaldehyde dimethyl acetal (46.1 mL, 0.31
mol) was added to a solution of the aforementioned azide product (33 g,
0.11 mol) and a catalytic amount of camphorsulfonic acid (CSA, 8.89 g,
38.3 mmol) in anhydrous MeCN (300 mL) at RT under a nitrogen atmos-
phere. After stirring at RT for 5 h, the reaction was quenched by the ad-
dition of Et3N, concentrated in vacuo, redissolved in EtOAc, sequentially
washed with a saturated aqueous solution of NaHCO3, water, and brine,
and dried over MgSO4. The filtrate was concentrated under reduced pres-
sure to give a dry residue that was purified by column chromatography
on silica gel (EtOAc/hexanes, 1:2). The desired fractions were concentrat-
ed in vacuo to afford a yellowish foam that was solidified from CHCl3/
Et2O/hexanes (1:5:20, v/v/v) to obtain compound 13 (28.7 g) as white
powder (as a mixture of a/b-anomers, 1:1.7) in 64% yield (of the last two
steps). Rf =0.39 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =ꢀ87 (c=1.3, MeOH);
1H NMR (500 MHz, CDCl3): d=7.50–7.47 (m, 5.4H (a+b); ArH), 7.41–
7.37 (m, 7.1H (a+b); ArH), 5.549 (s, 1H (a); PhCH), 5.545 (s, 1.7H (b);
PhCH), 5.32 (t, J=3.1 Hz, 1H (a); H1), 4.72 (dd, J=7.7, 4.6 Hz, 1.7H
(b); H1), 4.33 (dd, J=10.3, 5.1 Hz, 1.7H (b); H6a), 4.28 (dd, J=10.3,
5.0 Hz, 1H (a); H6a), 4.24 (dd, J=9.5, 1.7 Hz, 1H (a); H4), 4.10 (ddd,
J=15.0, 5.0, 5.0 Hz, 1H (a); H5), 3.78 (t, J=10.3 Hz, 1.7H (b); H6b),
3.73 (t, J=10.3 Hz, 1H (a); H6b), 3.70 (dd, J=9.4, 2.0 Hz, 1.7H (b);
H4), 3.59 (t, J=9.4 Hz, 1.7H (b); H3), 3.54 (t, J=9.5 Hz, 1H (a); H3),
3.45 (ddd, J=14.5, 5.0, 4.9 Hz, 1.7H (b); H5), 3.39 (dd, J=9.5, 3.1 Hz,
Acetyl (2,4,-Di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-b-d-
galactopyranosyl)-(1!3)-2-azido-4,6-O-benzylidene-2-deoxy-b-d-
glucopyranoside (18)
The following glycosylation reaction was performed immediately after
the sequential preparation of compound 12 without purification.
A solution of crude thiogalactoside 12 (which was obtained from a 4-
step, consecutive procedure without purification; for details, see
Scheme 2; 500 mg, 0.95 mmol of compound 10 were used at the start of
the synthesis) in anhydrous CH2Cl2/toluene (2:1, 30 mL) was mixed with
2,4,6-tri-tert-butylpyrimidine (TTBP, 199 mg, 0.80 mmol) and diphenyl-
sulfoxide (Ph2SO, 170 mg, 0.84 mmol). The reaction mixture was stirred
in the presence of 3 ꢁ molecular sieves (freshly dried prior to use, 1.0 g)
at RT for 30 min under a nitrogen atmosphere and subjected to the addi-
tion of Tf2O (139 mL, 0.84 mmol) after cooling to ꢀ608C. Upon consump-
tion of the glycosyl donor (by TLC, 10 min), a solution of glycosyl accept-
or 13 (334 mg, 1.14 mmol) in anhydrous CH2Cl2/MeCN (2:1, 30 mL) was
added to the reaction mixture. After stirring at ꢀ608C for a further 1 h,
the reaction was stopped by the addition of Et3N and filtered through
a pad of Celite. The filtrate was concentrated in vacuo to give a dry resi-
due that was redissolved in anhydrous CH2Cl2 (3.0 mL), Et3N (0.64 mL,
Chem. Asian J. 2013, 8, 1536 – 1550
1544
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
4.56 mmol) and Ac2O (0.21 mL, 2.28 mmol) were added, and the solution
was stirred at 48C under a nitrogen atmosphere. After stirring at 48C for
18 h, the acetylation reaction was stopped by the addition of MeOH,
evaporated in vacuo, redissolved in EtOAc, washed with a saturated
aqueous solution of NaHCO3 and brine, and dried over MgSO4. The fil-
trate was concentrated and purified by column chromatography on silica
gel (EtOAc/hexanes, 2:3) to give compound 18 (419 mg, 48% overall
yield in 6 steps from compound 10) as a white amorphous foam.
CDCl3): d=154.7, 137.2, 129.6, 128.6, 126.6, 102.1, 101.2, 95.7, 81.7, 77.5,
70.9, 70.3, 68.8, 66.3, 59.1, 51.6, 29.6, 28.9, 26.6, 25.7 ppm; HRMS (ESI-
TOF): m/z calcd for C22H29Cl3N4O7Na: 589.0994 [M+Na]+; found:
589.0958.
5-(Methoxycarbonyl)pentyl 2-trichloroacetamido-4,6-O-benzylidene-2-
deoxy-b-d-glucopyranoside (21)
For details of the reactions, see the Supporting Information, Scheme 4.
Preactivation method:
A solution of thiogalactoside 12 (200 mg,
To a solution of d-glucosamine hydrochloride (10.0 g, 47.3 mmol) in an-
hydrous MeOH (100 mL) were sequentially added NaOMe (9 mL, 30%
w/w in MeOH, 47.3 mmol), Et3N (7.8 mL, 56.8 mmol), and trichloroace-
tyl chloride (TCACl, 6.2 mL, 56.8 mmol) at 08C. The reaction mixture
was warmed to RT and stirred for 12 h. The reaction mixture was filtered
through a pad of Celite and the filtrate was concentrated under reduced
pressure. The resulting yellowish residue was redissolved in anhydrous
CH2Cl2 (100 mL) and DMAP (30 mg, 0.24 mmol), Et3N (13.6 mL,
98.3 mmol), and Ac2O (9.0 mL, 98.3 mmol) were added at 08C. After stir-
ring for 3 h, the reaction was stopped by the addition of MeOH and sub-
jected to extraction with EtOAc. The collected organic layer was washed
with a saturated aqueous solution of NaHCO3, dried over MgSO4, fil-
tered, concentrated in vacuo, and purified by column chromatography on
silica gel (EtOAc/hexanes, 1:2) to afford the peracetylated product
(15.4 g, 66% yield) as a white solid.
0.28 mmol) in anhydrous CH2Cl2/toluene (2:1, 9 mL) was mixed with
2,4,6-tri-tert-butylpyrimidine (TTBP, 73.0 mg, 0.29 mmol) and diphenyl-
sulfoxide (Ph2SO, 62 mg, 0.31 mmol). The reaction mixture was stirred in
the presence of 3 ꢁ molecular sieves (freshly dried prior to use, 400 mg)
at RT for 30 min under
a nitrogen atmosphere and Tf2O (51 mL,
0.31 mmol) was added after cooling the mixture to ꢀ608C. Upon con-
sumption of the glycosyl donor (by TLC, 10 min), a solution of glycosyl
acceptor 19 (141 mg, 0.42 mmol) in anhydrous CH2Cl2 (9 mL) was added
to the reaction mixture. After stirring at ꢀ608C for a further 1 h, the re-
action was stopped by the addition of Et3N and filtered through a pad of
Celite. The filtrate was concentrated in vacuo to give a dry residue that
was purified by column chromatography on silica gel (EtOAc/hexanes,
2:3) to give compound 18 (145 mg, 56% yield) as a white amorphous
foam.
The NIS/TfOH method: A suspension of purified thiogalactoside 12
(379 mg, 0.54 mmol), acceptor 19 (120 mg, 0.36 mmol), and freshly dried
3 ꢁ molecular sieves (500 mg) in anhydrous CH2Cl2 (3.6 mL) was stirred
at RT for 30 min and then cooled to 08C under a nitrogen atmosphere.
NIS (161 mg, 0.72 mmol) and TfOH (3.2 mL, 36 mmol) were added to the
reaction and the mixture was stirred at the same temperature for a further
0.5 h. The reaction was stopped by the addition of Et3N and filtered
through a pad of Celite. The filtrate was sequentially washed with a satu-
rated aqueous solution of NaHCO3, a saturated aqueous solution of
Na2S2O3, water, and brine, dried over MgSO4, and concentrated under re-
duced pressure. The residue was purified by column chromatography on
silica gel (EtOAc/hexanes, 2:3) to give compound 18 (230 mg, 70%
yield) as a pale-yellow amorphous foam. Rf =0.30 (EtOAc/hexanes, 2:3 v/
v); ½aꢁ2D5 =ꢀ29 (c=1.6, CHCl3); 1H NMR (500 MHz, CDCl3): d=7.53–
7.51 (m, 2H; ArH), 7.49–7.48 (m, 2H; ArH), 7.42–7.37 (m, 2H; ArH),
7.32 (t, J=7.4 Hz, 6H; ArH), 7.14 (t, J=7.2 Hz, 1H; ArH), 7.04 (d, J=
7.2 Hz, 2H; ArH), 5.91 (d, J=8.8 Hz, 1H; H1), 5.53 (t, J=3.0 Hz, 1H;
H4’), 5.51 (s, 1H; PhCH), 5.15 (dd, J=10.0, 8.0 Hz, 1H; H2’), 5.05 (dd,
J=10.0, 3.0 Hz, 1H; H3’), 4.75 (d, J=8.0 Hz, 1H; H1’), 4.26 (m, 2H;
H6ab), 3.79–3.77 (m, 1H; H5’), 3.75 (dd, J=10.0, 8.8 Hz, H2), 3.65 (dd,
J=10.0, 3.7 Hz, H3), 3.63 (t, J=3.7 Hz, 1H; H4), 3.60–3.52 (m, 3H; H5,
H6ab’), 2.77–2.71 (m, 1H; Lev-CH2), 2.60–2.48 (m, 2H; Lev-CH2), 2.39–
2.33 (m, 1H; Lev-CH2), 2.11 (s, 3H; COCH3), 2.08 (s, 3H; COCH3), 2.05
(s, 3H; COCH3), 1.96 (s, 3H; COCH3), 1.96 (s, 3H; COCH3), 0.97 ppm
(s, 9H; tBu); 13C NMR (125 MHz, CDCl3): d=206.4, 171.9, 170.3, 170.0,
169.9, 138.4, 136.9, 136.0, 135.8, 133.0, 132.8, 130.1, 130.0, 129.3, 128.3,
128.0, 125.9, 101.6, 101.2, 93.4, 79.5, 77.5, 77.2, 77.1, 73.3, 71.8, 69.8, 68.3,
67.3, 66.9, 65.2, 60.7, 37.9, 29.8, 28.1, 26.9, 21.1, 21.0, 20.8, 19.1 ppm;
HRMS (ESI-TOF): m/z calcd for C46H55N3O15SiNa: 940.3295 [M+Na]+;
found: 940.3295.
The peracetylated product (10.0 g, 20.3 mmol) was mixed with para-tol-
uenethiol (5.0 g, 40.3 mmol) in anhydrous CH2Cl2 (100 mL) at 08C under
a
nitrogen atmosphere and subjected to the dropwise addition of
BF3·OEt2 (6.40 mL, 50.5 mmol). The reaction mixture was gradually
warmed to RT and stirred for a further 48 h. The reaction was stopped by
the portion-wise addition of a saturated aqueous solution of NaHCO3 at
08C with vigorous stirring. The organic layer was washed with water and
brine, dried with anhydrous MgSO4, filtered, and concentrated under re-
duced pressure to give a dry residue that was purified by recrystallization
from hot EtOH to give the desired thioglycoside (7.0 g, 62% yield) as
a white powder.
A suspension of the thioglycoside (1.0 g, 1.79 mmol), methyl 6-hydroxy-
hexanoate (395 mg, 370 mL, 2.70 mmol), and 3 ꢁ molecular sieves (1.5 g,
freshly dried prior to use) in anhydrous CH2Cl2 (10 mL) was stirred at
RT for 30 min, cooled to 08C under a nitrogen atmosphere, and sequen-
tially treated with NIS (800 mg, 3.58 mmol) and TfOH (16 mL, 89 mmol).
The reaction mixture was stirred at 08C for a further 30 min, quenched
by the addition of Et3N, and filtered through a pad of Celite. The filtrate
was sequentially washed with a saturated aqueous solution of NaHCO3,
a saturated aqueous solution of Na2S2O3, water, and brine, dried over
MgSO4, and concentrated under reduced pressure. The residue was puri-
fied by column chromatography on silica gel (EtOAc/hexanes, 1:2) to
give the linker-tethered product (1.10 g, quantitative yield) as a white
solid.
The product (200 mg, 0.35 mmol) was dissolved in anhydrous MeOH
(2 mL) and treated with a catalytic amount of NaOMe (about 6 mg)
under a nitrogen atmosphere. After stirring for 1 h at RT, Amberlite IR-
120 [H+] resin was added to neutralize the mixture. The resulting mixture
was filtered, concentrated in vacuo, and then dissolved in anhydrous
MeCN (2 mL) for the next step of the protection reaction. A catalytic
amount of CSA (40 mg, 0.17 mmol) and benzaldehyde dimethyl acetal
(0.1 mL, 0.69 mmol) were added at 08C under a nitrogen atmosphere.
The mixture was warmed to RT and stirred for 5 h. The reaction was
stopped by the addition of Et3N (about 0.5 mL), concentrated under re-
duced pressure, washed with water (3ꢂ5 mL), and extracted with Et2O
to afford compound 21 (170 mg, 90% yield) as a white powder. Rf =0.25
(EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =ꢀ31 (c=1.2, CHCl3); 1H NMR
(400 MHz, CDCl3): d=7.50–7.40 (m, 2H; ArH), 7.39–7.27 (m, 2H;
ArH), 7.03 (d, J=7.1 Hz, 1H; NH), 5.55 (s, 1H; PhCH), 4.93 (d, J=
8.6 Hz, 1H; H1), 4.83 (t, J=10.2 Hz, 1H; H3), 4.39–4.34 (m, 1H; H6a),
3.87 (td, J=9.7, 6.2 Hz, 1H; alkyl chain), 3.79 (t, J=9.7 Hz, 1H; H6b),
3.67 (s, 3H; CO2CH3), 3.58 (m, 4H; H2, H4, H5, alkyl chain), 2.31 (d, J=
3.2 Hz, 1H; C3-OH), 2.30 (t, J=7.4 Hz, 2H; alkyl chain), 1.65–1.62 (m,
4H; alkyl chain), 1.41–1.28 ppm (m, 2H; alkyl chain); 13C NMR
(100 MHz, CDCl3): d=174.3, 162.4, 137.1, 129.56, 128.6, 126.5, 102.1,
Acetyl 2-Azido-4,6-O-benzylidene-2-deoxy-b-d-glucopyranoside (19)[17,18]
Compound 19 was synthesized by the regio- and stereoselective acetyla-
tion of compound 13 according to a literature procedure.
6-Azidohexyl 4,6-O-benzylidene-2-deoxy-2-(2,2,2-
trichloroethoxycarbonylamino)-b-d-glucopyranoside (20)
Rf =0.25 (EtOAc/hexanes, 1:2 v/v); ½aꢁ2D5 =ꢀ30 (c=1.0, CHCl3); H NMR
1
(500 MHz, CDCl3): d=7.48–7.45 (m, 2H; ArH), 7.38–7.33 (m, 3H;
ArH), 5.50 (s, 1H; PhCH), 5.32 (d, J=8.0 Hz, 1H; NH), 4.74–4.69 (m,
2H; OCH2CCl3), 4.57 (s, 1H; H1), 4.32–4.29 (m, 1H; H6a), 4.09 (s, 1H;
H3), 3.85–3.81 (m, 1H; OCH2), 3.77–3.72 (t, J=8.0 Hz, 1H; H6b), 3.52
(t, J=8.0 Hz, 1H; H4), 3.46–4.42 (m, 2H; OCH2, H5), 3.34–3.35 (m, 1H;
H2), 3.25 (t, J=7.0 Hz, 2H; CH2N3), 3.16 (s, 1H; OH), 1.57 (m, 4H;
alkyl chain), 1.35 ppm (m, 4H; alkyl chain); 13C NMR (125 MHz,
Chem. Asian J. 2013, 8, 1536 – 1550
1545
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
100.2, 92.7, 81.9, 70.3, 69.8, 68.8, 66.4, 59.9, 51.7, 34.1, 29.3, 25.7,
24.8 ppm; HRMS (ESI-TOF): m/z calcd for C22H28Cl3NO8Na: 562.0787
[M+Na]+; found: 562.0773.
gel (EtOAc/hexanes, 2:3) to give compound 23 (92 mg, 76% yield) as
a white solid. Rf =0.18 (EtOAc/hexanes, 1:2 v/v); ½aꢁ2D5 =ꢀ7.7 (c=1.7,
CHCl3); 1H NMR (500 MHz, CDCl3): d=7.37–7.27 (m, 10H; ArH), 6.96
(d, J=7.8 Hz, 1H; NH), 4.88 (d, J=8.3 Hz, 1H; H1), 4.78 (ABq, 2H;
PhCH2), 4.60 (ABq, 2H; PhCH2), 4.03 (dd, J=10.4, 8.3 Hz, 1H; H3),
3.87 (td, J=9.4, 6.2 Hz, 1H; OCH2), 3.77 (dd, J=9.9, 7.3 Hz, 1H; H6a),
3.76 (dd, J=9.9, 7.3 Hz, 1H; H6b), 3.71 (dd, J=8.8, 2.0 Hz, 1H; H4),
3.66 (s, 3H; CO2CH3), 3.56–3.51 (m, 1H; H5), 3.27 (td, J=9.4, 6.2 Hz,
1H; alkyl chain), 3.45 (dd, J=9.8, 7.8 Hz, 1H; H2), 2.79 (d, J=2.6 Hz,
1H; OH), 2.29 (t, J=2.6 Hz, 2H; CH2CO2Me), 1.63–1.57 (m, 4H; alkyl
chain), 1.37–1.63 ppm (m, 2H; alkyl chain); 13C NMR (125 MHz, CDCl3):
d=174.3, 162.0, 138.3, 137.8, 128.8, 128.7, 128.3, 128.8, 128.1, 128.0, 99.5,
92.8, 79.8, 74.9, 74.0, 73.9, 73.8, 70.8, 69.9, 58.7, 51.7, 34.1, 29.4, 25.7,
24.8 ppm; HRMS (ESI-TOF): m/z calcd for C29H36Cl3NO8Na: 656.1375
[M+Na]+; found: 656.1409.
Acetyl 2-Azido-3,6-di-O-benzyl-2-deoxy-b-d-glucopyranoside (22)[8d]
For details of the reactions, see the Supporting Information, Scheme 2.
Triethylamine (28.6 mL, 205 mmol) was added to a solution of compound
13 (20.0 g, 68.2 mmol) and acetic anhydride (7.1 mL, 75.0 mmol) in anhy-
drous CH2Cl2 (500 mL) at 08C under a nitrogen atmosphere. Then, the
mixture was gradually warmed to RT and stirred overnight. The reaction
was quenched by the addition of MeOH (20 mL), stirred for 10 min, con-
centrated under reduced pressure, redissolved in EtOAc, washed with
a saturated aqueous solution of NaHCO3 and brine, and dried over
MgSO4. The filtrate was evaporated in vacuo to give a dry residue, which
was purified by column chromatography on silica gel (EtOAc/hexanes,
1:2) to afford the O1-acetate (18.6 g, 82% yield).
6-Azidohexyl (2,4-di-O-acetyl-6-O-tert-butylphenylsilyl-3-O-levulinoyl-b-
d-galactopyranosyl)-(1!3)-4,6-O-benzylidene-2-deoxy2-(2,2,2-
trichloroethoxycarbonylamino)-b-d-glucopyranoside (24)
A mixture of the aforementioned O1-acetate (10.0 g, 29.8 mmol) and
Ag2O (13.8 g, 59.6 mmol) in anhydrous CH2Cl2 (150 mL) was treated
with benzyl bromide (6.1 mL, 50.7 mmol) at 08C under a nitrogen atmos-
phere. The reaction was gradually warmed to RT and stirred for a further
2 days. The resulting mixture was filtered through a pad of Celite. The fil-
trate was concentrated in vacuo to give a dry residue that was purified by
column chromatography on silica gel (EtOAc/hexanes, 1:3) to afford the
benzyl ether (11.2 g, 88% yield) as a white solid.
A suspension solution of thiogalactoside 12 (158 mg, 0.22 mmol), accept-
or 20 (98 mg, 0.17 mmol), and 3 ꢁ molecular sieves (550 mg, freshly
dried prior to use) in anhydrous CH2Cl2 (6 mL) was stirred at RT for
30 min under a nitrogen atmosphere, cooled to 08C, and treated with
NIS (100 mg, 0.45 mmol) and TfOH (1.5 mL, 17 mmol). The reaction mix-
ture was stirred at 08C for 30 min, quenched by the addition of Et3N, and
filtered through a pad of Celite. The filtrate was sequentially washed
with a saturated aqueous solution of NaHCO3, a saturated aqueous solu-
tion of Na2S2O3, water, and brine, dried over MgSO4, and concentrated
under reduced pressure. The residue was purified by column chromatog-
raphy on silica gel (EtOAc/hexanes, 1:2) to give compound 24 (140 mg,
71% yield) as a pale-yellow amorphous foam. Rf =0.30 (EtOAc/hexanes,
1:2 v/v); ½aꢁ2D5 =ꢀ14 (c=1.1, CHCl3); 1H NMR (500 MHz, CDCl3): d=
7.63–7.3 (m, 12H; ArH), 7.09 (d, J=7.0 Hz, 2H; ArH), 7.01 (m, 2H;
ArH), 5.49 (d, J=2.0 Hz, 1H; H4’), 5,41 (s, 1H; PhCH), 5.25 (s, 1H;
NH), 5.17 (t, J=10.0 Hz, 1H; H2’), 4.99 (dd, J=10.0 Hz, 3.0 Hz, 1H;
H3’), 4.80 (d, J=11.0 Hz, 2H; H1, OCH2CCl3), 4.68 (m, 2H;
H1’,OCH2CCl3), 4.32 (m, 1H; H3), 4.28(dd, J=11.0 Hz, 5.0 Hz, 1H;
H6a), 3.83–3.80 (m, 1H; OCH2), 3.74–3.66 (m, 2H; H5’, H6b), 3.60–3.50
(m, 3H; H4, H6a, H6b), 3.46–3.37 (m, 2H; H5, OCH2), 3.24 (t, J=
7.0 Hz, 2H,CH2N3), 3.19 (d, J=8.0 Hz, 1H; H2), 2.78–2.72 (m, 1H; Lev-
CH2), 2.60–2.48 (m, 2H; Lev-CH2), 2.38–2.32 (m, 1H; Lev-CH2), 2.12 (s,
3H; COCH3), 2.06 (s, 3H; COCH3), 1.88 (s, 3H; COCH3), 1.55–1.54 (m,
4H; alkyl chain), 1.36–1.33 (m, 4H; alkyl chain), 0.95 ppm (s, 9H; tBu);
13NMR (125 MHz, CDCl3): d=206.4, 171.8, 170.3, 169.9, 154.0, 137.2,
136.0, 135.8, 133.0, 132.7, 130.1, 130.0, 129.8, 129.1, 128.2, 128.0, 125.9,
101.4, 101.1, 100.4, 95.6, 80.1, 77.5, 73.0, 71.8, 70.3, 69.6, 68.7, 66.9, 66.3,
60.5, 58.6, 51.5, 37.8, 29.8, 29.5, 28.9, 28.0, 26.9, 26.6, 25.7, 21.0, 20.8,
19.0 ppm; HRMS (ESI-TOF): m/z calcd for C53H67Cl3N4O16SiNa:
1171.3285 [M+Na]+; found: 1171.3279.
To a solution of the benzyl ether (6.0 g, 14.1 mmol) and triethylsilane
(Et3SiH, 13.5 mL, 84.6 mmol) in anhydrous CH2Cl2 (120 mL) at 08C were
dropwise added trifluoroacetic anhydride (TFAA, 2.0 mL, 14.1 mmol)
and trifluoroacetic acid (TFA, 5.4 mL, 70.5 mmol). After stirring at 08C
for 2 h, the reaction mixture was poured into a saturated aqueous solu-
tion of NaHCO3 and extracted twice with EtOAc. The combined organic
layer was washed with brine and dried over MgSO4. The filtrate was con-
centrated under reduced pressure to give a dry residue that was purified
by column chromatography on silica gel (EtOAc/hexanes, 1:4) to give
compound 20 (5.18 g, 86% yield) as a colorless oil. Rf =0.45 (EtOAc/hex-
anes, 1:3 v/v); ½aꢁ2D5 =ꢀ23 (c=2.1, CHCl3); 1H NMR (500 MHz, CDCl3):
d=7.41–7.26 (m, 10H; ArH), 5.47 (d, J=8.5 Hz, 1H; H1), 4.90 (d, J=
11.0 Hz, 1H; PhCH), 4.82 (d, J=11.0 Hz, 1H; PhCH), 4.60 (d, J=
12.0 Hz, 1H; PhCH), 4.51 (d, J=12.0 Hz, 1H; PhCH), 3.78–3.73 (m,
2H), 3.68 (dd, J=4.4, 10.2 Hz, 1H), 3.55–3.49 (m, 2H), 3.38 (t, J=
9.0 Hz, 1H), 2.64 (d, J=2.6 Hz, 1H; OH), 2.16 ppm (s, 3H; COCH3);
13C NMR (125 MHz, CDCl3): d=169.0, 137.8, 137.4, 128.6, 128.5, 128.2,
128.1, 128.0, 127.9, 92.9, 82.5, 75.3, 74.5, 73.8, 71.8, 69.5, 64.5, 21.0 ppm;
HRMS (ESI-TOF): m/z calcd for C22H25N3O6Na: 450.1636 [M+Na]+;
found: 450.1637.
5-(Methoxycarbonyl)pentyl 2-trichloroacetamido-3,6-di-O-benzyl-2-deoxy-
b-d-glucopyranoside (23)
For details of the reactions, see the Supporting Information, Scheme 4.
To a solution of compound 21 (1.0 g, 1.93 mmol) in anhydrous DMF
(20 mL) was added sodium hydride (60% NaH in mineral oil, 0.14 g,
5.79 mmol) at 08C under a nitrogen atmosphere. The mixture was stirred
for 10 min at 08C and then benzyl bromide (BnBr, 470 mL, 3.85 mmol)
and a catalytic amount of tetra-n-butylammonium iodide (TBAI, 70 mg,
0.19 mmol) were added at 08C. The reaction was stirred for a further
30 min and quenched by the slow addition of water (50 mL). The mixture
was extracted with EtOAc (3ꢂ50 mL), washed with water and brine,
dried over anhydrous MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by column chromatography on silica
gel (EtOAc/hexanes, 1:2) to afford the benzyl ether (850 mg, 73% yield)
as a white solid.
5-(Methoxycarbonyl)pentyl (2,4-di-O-acetyl-6-O-tert-butylphenylsilyl-3-O-
levulinoyl-b-d-galactopyranosyl)-(1!3)-2-trichloroacetamido-4,6-O-
benzylidene-2-deoxy-b-d-glucopyranoside (25)
A suspension of thiogalactoside 12 (170 mg, 0.24 mmol), acceptor 21
(100 mg, 0.19 mmol), and 3 ꢁ molecular sieves (300 mg, freshly dried
prior to use) in anhydrous CH2Cl2 (2 mL) was stirred at RT for 30 min
under a nitrogen atmosphere, cooled to 08C, and sequentially treated
with NIS (108 mg, 0.48 mmol) and TfOH (1.7 mL, 19 mmol). The reaction
mixture was stirred at 08C for 30 min, quenched by the addition of Et3N,
and filtered through a pad of Celite. The filtrate was sequentially washed
with a saturated aqueous solution of NaHCO3, a saturated aqueous solu-
tion of Na2S2O3, water, and brine, dried over MgSO4, and concentrated
under reduced pressure. The residue was purified by column chromatog-
raphy on silica gel (EtOAc/hexanes, 1:2) to give compound 25 (142 mg,
77% yield) as a yellowish amorphous foam. Rf =0.28 (EtOAc/hexanes,
1:1 v/v); ½aꢁ2D5 =ꢀ20 (c=1.3, CHCl3); 1H NMR (400 MHz, CDCl3,): d=
7.56–7.53 (m, 4H; ArH), 7.53–7.34 (m, 8H; ArH), 7.19 (t, J=7.4 Hz, 1H;
ArH), 7.07 (t, J=7.64 Hz, 2H; ArH), 7.02 (d, J=7.4 Hz, 1H; NH), 5.51
(d, J=3.24 Hz, 1H; H4’), 5.44 (s, 1H; PhCH), 5.16 (dd, J=10.4, 7.9 Hz,
The benzyl ether (120 mg, 0.19 mmol) was dissolved in anhydrous CH2Cl2
(1 mL) and triethylsilane (0.18 mL, 1.14 mmol), TFAA (24.6 mL,
0.19 mmol), and TFA (70.7 mL, 0.95 mmol) were sequentially added at
08C under a nitrogen atmosphere. The reaction mixture was stirred at
08C for 5 h and then stopped by the addition of a saturated aqueous solu-
tion of NaHCO3. The organic layer was washed with water and brine,
dried over anhydrous MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by column chromatography on silica
Chem. Asian J. 2013, 8, 1536 – 1550
1546
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
1H; H2’), 4.97 (d, J=8.5 Hz, 1H; H1), 4.94 (dd, J=10.4, 3.2 Hz, 1H;
H3’), 4.68 (d, J=8.0 Hz, 4H; H1), 4.53 (t, J=9.4 Hz, 1H; H3), 4.30 (dd,
J=10.5, 4.9 Hz, 1H; H6a), 3.86 (dt, J=9.6, 6.2 Hz, 1H; alkyl chain), 3.74
(t, J=10.3 Hz, 1H; H6b), 3.66 (s, 3H; CO2CH3), 3.65–3.63 (m, 2H;
H6’ab), 3,60 (t, J=9.5 Hz, 2H; H4, H5’), 3.48 (dt, J=9.7, 6.2 Hz, 2H;
H5, alkyl chain), 3.39 (dt, J=9.8, 9.7 Hz, 1H; H2), 2.78–2.73 (m, 1H;
Lev-CH2), 2.64–2.49 (m, 2H; Lev-CH2), 2.41–2.34 (m, 1H; Lev-CH2),
2.29 (t, J=7.6 Hz, 2H; CH2CO2Me), 2.18 (s, 3H; COCH3), 2.04 (s, 3H;
COCH3), 1.92 (s, 3H; COCH3), 1.66–1.52 (m, 4H; alkyl chain), 1.43–
1.1.30 (m, 2H; alkyl chain), 0.99 ppm (s, 9H; tBu); 13C NMR (100 MHz,
CDCl3): d=207.2, 206.4, 174.35, 171.9, 170.3, 169.9, 162.1, 137.1, 135.9,
135.8, 132.9, 132.7, 130.1, 129.2, 128.2, 128.0, 127.9, 126.1, 101.1, 100.3,
99.5, 92.6, 79.6, 76.0, 73.0, 71.9, 70.4, 69.4, 68.7, 66.9, 66.4, 60.6, 59.6, 51.7,
37.8, 34.0, 31.1, 29.9, 29.2, 28.0, 26.9, 25.7, 24.8, 21.1, 20.8, 19.1 ppm;
HRMS (ESI-TOF): m/z calcd for C53H66Cl3NO17SiNa: 1146.3045
[M+Na]+; found: 1146.3024.
H2’), 4.93 (d, J=10.1 Hz, 1H; PhCH2), 4.82 (dd, J=3.5, 10.5 Hz, 1H;
H3’), 4.76 (d, J=12.0 Hz, 1H; PhCH2), 4.59 (d, J=10.0 Hz, 1H; PhCH2),
4.43 (d, J=8.0 Hz, 1H; H1’), 4.38 (d, J=12.0 Hz, 1H; PhCH2), 4.02 (t,
J=9.5 Hz, 1H; H5), 3.75 (dd, J=2.6, 11.2 Hz, 1H; H6a’), 3.69 (dd, J=
1.3, 11.0 Hz, 1H; H6b’), 3.59 (dd, J=5.3, 9.6 Hz, 1H; H3), 3.48–3.44 (m,
2H; H2, H6a), 3.41–3.36 (m, 3H; H4, H5’, H6b), 2.84–2.78 (m, 1H; Lev-
CH2), 2.66–2.59 (m, 1H; Lev-CH2), 2.57–2.51 (m, 1H; Lev-CH2), 2.42–
2.36 (m, 1H; Lev-CH2), 2.18 (s, 3H; COCH3), 2.15 (s, 3H; COCH3), 2.02
(s, 3H; COCH3), 1.98 (s, 3H; COCH3), 1.02 ppm (s, 9H; tBu); 13C NMR
(125 MHz, CDCl3): d=206.2, 171.5, 170.0, 169.4, 169.1, 137.7, 137.4,
135.7, 135.5, 132.8, 129.9, 126.7, 128.4, 128.3, 128.2, 127.9, 127.8, 127.6,
99.8, 92.7, 81.1, 75.6, 75.5, 75.0, 73.7, 73.0, 71.5, 69.5, 67.0, 66.8, 64.2, 60.2,
37.7, 29.6, 27.8, 26.7, 21.0, 20.8, 20.6, 19.0 ppm; HRMS (ESI-TOF): m/z
calcd for C53H63N3O15SiNa: 1032.3921 [M+Na]+; found: 1032.3969.
5-(Methoxycarbonyl)pentyl (2,4-di-O-acetyl-6-O-tert-butylphenylsilyl-3-O-
levulinoyl-b-d-galactopyranosyl)-(1!4)-2-trichloroacetamido-3,6-O-
benzyl-2-deoxy-b-d-glucopyranoside (27)
Acetyl (2,4-Di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-b-d-
galactopyranosyl)-(1!4)-2-azido-3,6-di-O-benzyl-2-deoxy-b-d-
glucopyranoside (26)
A suspension of thiogalactoside 12 (170 mg, 0.24 mmol), acceptor 23
(75.0 mg, 0.12 mmol), and 3 ꢁ molecular sieves (300 mg, freshly dried
prior to use) in anhydrous CH2Cl2 (2 mL) was stirred at RT for 30 min
under a nitrogen atmosphere, cooled to 08C, and sequentially treated
with NIS (69.0 mg, 0.31 mmol) and TfOH (1.6 mL, 19 mmol). The reaction
was stirred at 08C for 30 min, stopped by the addition of Et3N, and fil-
tered through a pad of Celite. The filtrate was sequentially washed with
a saturated aqueous solution of NaHCO3, a saturated aqueous solution
of Na2S2O3, water, and brine, dried over MgSO4, and concentrated under
reduced pressure. The residue was purified by column chromatography
on silica gel (EtOAc/hexanes, 1:2) to give compound 27 (90 mg, 76%
yield) as a yellowish amorphous foam. Rf =0.45 (EtOAc/hexanes, 1:1 v/
v); ½aꢁ2D5 =ꢀ12 (c=1.1, CHCl3); 1H NMR (400 MHz, CDCl3,): d=7.57–
7.54 (m, 4H; ArH), 7.48–7.27 (m, 11H; ArH), 7.27–7.20 (m, 2H; ArH),
7.17–7.08 (m, 1H; ArH), 7.03 (d, J=7.5 Hz, 1H; NH), 7.02–6.98 (m, 2H;
ArH), 5.52 (d, J=3.45 Hz, 1H; H4’), 5.09 (dd, J=10.4, 7.9 Hz, 1H; H2’),
4.91 (dd, J=10.43, 3.45 Hz, 1H; H3’), 4.86 (d, J=7.53 Hz, 1H; H1), 4.83
(d, J=10.3 Hz, 1H; PhCH2), 4.93 (d, J=12.1 Hz, 1H; PhCH2), 4.50 (d,
J=7.9 Hz, 1H; H1’), 4.48 (d, J=9.5 Hz, 1H; PhCH2), 4.47 (d, J=
12.7 Hz, 1H; PhCH2), 4.30 (t, J=8.4 Hz, 1H; H3), 3.95 (t, J=8.0 Hz,
1H; H4), 3.74 (dt, J=9.7, 6.1 Hz, 1H; alkyl chain), 3.77–3.75 (m, 2H;
H6ab), 3.66 (s, 3H; CO2CH3), 3,60 (dd, J=7.6, 3.5 Hz, 1H; H6’a), 3.52–
3.47 (m, 5H; H2, H5, H5’, H6’b, alkyl chain), 2.80–2.78 (m, 1H; Lev-
CH2), 2.67–2.52 (m, 2H; Lev-CH2), 2.43–2.41 (m, 1H; Lev-CH2), 2.30 (t,
J=7.5 Hz, 1H; CH2CO2Me), 2.19 (s, 3H; COCH3), 2.06 (s, 3H; COCH3),
1.98 (s, 3H; COCH3), 1.56–1.53 (m, 4H; alkyl chain), 1.41–1.31 (m, 2H;
alkyl chain), 1.02 ppm (s, 9H; tBu); 13C NMR (100 MHz, CDCl3): d=
174.3, 171.8, 170.19, 169.7, 161.8, 138.1, 138.0, 135.9, 135.7, 133.0, 130.1,
128.8, 128.3, 128.2, 128.0, 127.8, 100.4, 99.4, 92.7, 76.4, 75.3, 74.8, 73.8,
73.2, 71.6, 69.8, 69.7, 68.2, 67.0, 60.5, 57.7, 51.7, 37.9, 34.1, 29.9, 29.3, 28.0,
27.0, 25.8, 24.8, 21.0, 20.9, 19.2 ppm; HRMS (ESI-TOF): m/z calcd for
C60H74Cl3NO17SiNa: 1238.3674 [M+Na]+; found: 1238.3715.
The following glycosylation reaction was performed immediately after
the sequential preparation of compound 12 without purification.
A solution of crude thiogalactoside 12 (which was obtained from a 4-
step, consecutive procedure without purification; for details, see
Scheme 2; 500 mg, 0.95 mmol of compound 10 were used at the start of
the synthesis), acceptor 22 (487 mg, 1.14 mmol), and freshly activated
3 ꢁ molecular sieves (2.0 g) in CH2Cl2 (10 mL) was stirred at RT for
30 min and then cooled to ꢀ458C under a nitrogen atmosphere. NIS
(426 mg, 1.90 mmol) and TfOH (8.9 mL, 0.10 mmol) were sequentially
added to the reaction and the resulting mixture was stirred at ꢀ458C for
a further 1 h. The reaction was quenched by the addition of Et3N, diluted
with EtOAc, and filtered through a pad of Celite. The filtrate was
washed with a saturated aqueous solution of Na2S2O3, a saturated aque-
ous solution of NaHCO3, water, and brine, dried over anhydrous MgSO4,
filtered, and concentrated under reduced pressure. The resulting residue
was purified by column chromatography on silica gel (EtOAc/hexanes,
2:3) to give compound 24 (520 mg, 54% overall yield for 5 steps starting
from compound 10) as a pale-yellow amorphous foam.
Preactivation method: A solution of purified thiogalactoside 12 (200 mg,
0.28 mmol), TTBP (73.0 mg, 0.29 mmol), and Ph2SO (62 mg, 0.31 mmol)
in anhydrous CH2Cl2/toluene (2:1, 6 mL) was stirred in the presence of
3 ꢁ molecular sieves (400 mg, freshly dried prior to use) at RT for
30 min, cooled to ꢀ608C under a nitrogen atmosphere, and treated with
Tf2O (51 mL, 0.31 mmol). Upon consumption of the glycosyl donor (by
TLC), a solution of glycosyl acceptor 22 (180 mg, 0.42 mmol) in anhy-
drous CH2Cl2 (6 mL) was added to the reaction and the resulting mixture
was stirred at ꢀ608C for a further 1 h. The reaction was stopped by the
addition of Et3N and filtered through a pad of Celite. The filtrate was
concentrated in vacuo and purified by column chromatography on silica
gel (EtOAc/hexanes, 2:3) to afford compound 26 (234 mg, 80% yield) as
a white amorphous foam.
(2,4-Di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-b-d-
galactopyranosyl)-(1!4)-2-trichloroacetamido-3,6-di-O-benzyl-2-deoxy-
ab-d-glucopyranosyl N-phenyltirfluoroacetimidate (29)
The NIS/TfOH method: A suspension of purified thiogalactoside 12
(744 mg, 1.05 mmol), acceptor 22 (300 mg, 0.70 mmol), and freshly dried
3 ꢁ molecular sieves (2.0 g) in anhydrous CH2Cl2 (7 mL) was stirred at
RT for 30 min and then cooled to ꢀ458C under a nitrogen atmosphere.
NIS (471 mg, 2.10 mmol) and TfOH (9.8 mL, 0.11 mmol) were added to
the reaction and the mixture was stirred at the same temperature for
a further 1 h. The reaction was stopped by the addition of Et3N and fil-
tered through a pad of Celite. The filtrate was sequentially washed with
a saturated aqueous solution of NaHCO3, a saturated aqueous solution
of Na2S2O3, water, and brine, dried over MgSO4, and concentrated under
reduced pressure. The residue was purified by column chromatography
on silica gel (EtOAc/hexanes, 2:3) to give compound 26 (596 mg, 84%
yield) as a pale-yellow amorphous foam. Rf =0.30 (EtOAc/hexanes, 2:3 v/
v); ½aꢁ2D5 =ꢀ44 (c=1.1, CHCl3); 1H NMR (500 MHz, CDCl3): d=7.56–
7.53 (m, 4H; ArH), 7.48–7.42 (m, 2H; ArH), 7.40–7.30 (m, 11H; ArH),
7.11–7.08 (m, 1H; ArH), 7.03–7.00 (m, 2H; ArH), 5.50 (d, J=3.3 Hz,
1H; H4’), 5.34 (d, J=8.5 Hz, 1H; H1), 5.06 (dd, J=8.1, 10.5 Hz, 1H;
A solution of compound 26 (300 mg, 0.30 mmol) in anhydrous CH2Cl2/
MeOH (1:1, 3 mL) was treated with a fresh 1.0m solution of PEt3
(0.36 mL, 0.36 mmol in THF) at 08C under a nitrogen atmosphere. After
the addition had been completed, the ice bath was removed and the mix-
ture was gradually warmed to RT, accompanied by the release of gas.
The reaction was stirred at RT for a further 1 h and then concentrated in
vacuo. The residue was kept under high vacuum for at least 2 h to
remove most of the phosphine and redissolved in anhydrous CH2Cl2
(3 mL) at 08C under a nitrogen atmosphere. To the resulting solution
were sequentially added trichloroacetic chloride (0.15 mL, 1.35 mmol)
and Et3N (0.25 mL, 1.80 mmol). The mixture was stirred at 08C for
15 min and then poured into ice water for extraction. The combined or-
ganic layers were washed with a saturated aqueous solution of NaHCO3
and brine, dried over MgSO4, filtered, and concentrated in vacuo. The
residue was purified by column chromatography on silica gel (EtOAc/
Chem. Asian J. 2013, 8, 1536 – 1550
1547
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
hexanes, 3:2) to give the trichloroacetamide-protected product 28
(171 mg, 53% overall yield in 3 steps) as a white amorphous foam.
10.4 Hz, 1H; PhCH2), 4.47 (d, J=7.8 Hz, 1H; H1’), 4.47 (ABq, J=
11.9 Hz, 1H; PhCH2), 3.99 (t, J=8.3 Hz, 1H; H3), 3.95 (t, J=7.8 Hz,
1H; H4), 3.84 (dt, J=9.9, 6.2 Hz, 1H; OCH2), 3.81–3.76 (m, 2H; H6a,b’),
3.68 (dt, J=10.1, 4.2 Hz, 1H; H3’), 3.65 (s, 3H; COOCH3), 3.61 (dd, J=
9.6, 5.5 Hz, 1H; H6a), 3.55–3.42 (m, 5H; H5, H6b, OCH2, H2, H5’),
2.41–2.39 (br, 1H; OH), 2.27 (t, J=7.5 Hz, 2H; CH2COOCH3), 2.08 (s,
3H; COCH3), 2.01 (s, 3H; COCH3), 1.64–1.53 (m, 6H; alkyl chain),
1.38–1.32 (m, 2H; alkyl chain), 1.03 ppm (s, 9H; tBu); 13C NMR
(125 MHz, CDCl3): d=174.3, 171.5, 170.8, 161.9, 138.2, 135.9, 135.7,
133.1, 133.0, 130.12, 130.08, 128.7, 128.3, 128.2, 128.0, 127.8, 100.3, 99.5,
92.7, 76.5, 75.4, 74.6, 73.8, 73.5, 73.4, 72.0, 69.8, 69.7, 68.4, 60.8, 57.5, 51.7,
34.1, 29.3, 27.0, 25.8, 24.9, 21.2, 21.0, 19.3 ppm; HRMS (ESI-TOF): m/z
calcd for C55H68Cl3NO15SiNa: 1138.3316 [M+Na]+; found: 1138.3366.
A solution of compound 28 (150 mg, 0.14 mmol) in anhydrous CH2Cl2
(3 mL) was treated with Cs2CO3 (91 mg, 0.28 mmol) and 2,2,2-trifluoro-
N-phenylacetimidoyl chloride (87 mg, 0.42 mmol) at 08C under a nitrogen
atmosphere, gradually warmed to RT, and stirred for 16 h. The resulting
mixture was filtered through a pad of Celite. The filtrate was concentrat-
ed in vacuo and purified by column chromatography on silica gel
(EtOAc/hexanes, 1:2) to afford compound 29 (110 mg, 63% yield) as
a white amorphous foam (a,b-mixture, a/b 1.8:1). Rf =0.65 (EtOAc/hex-
anes, 1:1 v/v); ½aꢁ2D5 =+26 (c=1.2, CHCl3); 1H NMR (500 MHz, CDCl3):
d=7.56–7.52 (m, 11.2H; ArH
ACHUTNGTRENNUG(a+b)), 7.46–7.30 (m, 30.8H; ArHCAHTUNGTRENNUNG
7.28–7.25 (m, 5.6H; ArH(a+b)), 7.22–7.08 (m, 16.8H; ArH+NHACHTUNGTRENNUNG
G
6.76 (d, J=7.7 Hz, 3.6H; ArH(a)), 6.50 (d, J=7.6 Hz, 2H; ArH(b)), 6.45
(br, 1.8H; H1(a)), 6.27 (d, J=7.4 Hz, 1H; H1(b)), 5.52 (d, J=3.4 Hz,
1H; H4’(b)), 5.52 (d, J=3.5 Hz, 1.8H; H4ꢃ (a)), 5.10 (dd, J=10.4, 8.3 Hz,
1.8H; H2’(a)), 5.07 (dd, J=10.4, 7.8 Hz, 1H; H3’(b)), 4.91 (dd, J=10.4,
3.5 Hz, 1H; H3’(b)), 4.87 (dd, J=10.4, 3.5 Hz, 1.8H; H3’(a)), 4.89 (ABq,
J=10.9 Hz, 1.8H; PhCH2(a)), 4.75 (ABq, J=12.0 Hz, 1.8H; PhCH2(a)),
4.66 (ABq, J=12.4 Hz, 1H; PhCH2(b)), 4.56 (ABq, J=10.9 Hz, 1.8H;
PhCH2(a)), 4.56 (ABq, J=10.8 Hz, 1H; PhCH2(b)), 4.49 (ABq, J=
12.4 Hz, 1H; PhCH2(b)), 4.49 (ABq, J=10.8 Hz, 1H; PhCH2(b)), 4.47 (d,
J=8.5 Hz, 1.8H; H-1(a)), 4.45 (ABq, J=12.0 Hz, 1.8H; PhCH2(a)), 4.38
(d, J=8.0 Hz, 1H , H-1’(b)), 4.37 (td, J=7.4, 1.0 Hz, 1H; H2(b)), 4.19 (t,
J=2.3 Hz, 1H; H6a(b)), 4.15–4.11 (m, 1.8H; H2(a)), 4.12 (t, J=7.2 Hz,
1H; H4(b)), 4.10 (t, J=9.3 Hz, 1.8H; H4(a)), 3.98–3.94 (m, 1H; H5(b)),
3.79–3.71 (m, 5.6H (a+b)), 3.67–3.41 (m, 9.2H (a+b)), 2.86–2.75 (m,
2.8H (a+b); Lev-CH2), 2.66–2.50 (m, 5.6H; Lev-CH2), 2.42–2.36 (m,
5-(Methoxycarbonyl)pentyl (2,4-di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-
O-levulinoyl-b-d-galactopyranosyl)-(1!4)-(2-trichloroacetamido-3,6-di-
O-benzyl-2-deoxy-b-d-glucopyranosyl)-(1!3)-(2,4-di-O-acetyl-6-O-tert-
butyldiphenylsilyl-b-d-galactopyranosyl)-(1!4)-2-trichloroacetamido-3,6-
di-O-benzyl-2-deoxy-b-d-glucopyranoside (31)
A suspension of glycosyl donor 29 (100 mg, 79.5 mmol), glycosyl acceptor
27 (59 mg, 53.0 mmol), and 3 ꢁ molecular sieves (320 mg, freshly dried
prior to use) in anhydrous CH2Cl2 (1 mL) was stirred at RT for 30 min
under a nitrogen atmosphere, cooled to 08C, and a catalytic amount of
TMSOTf (2.9 mL, 15.9 mmol) was added. The reaction was stirred at 08C
for a further 1 h, quenched by the addition of Et3N, and filtered through
a pad of Celite. The filtrate was concentrated under reduced pressure
and purified by column chromatography on silica gel (EtOAc/hexanes,
1:2) to give compound 31 (64.3 mg, 62% yield) as a white amorphous
foam. Rf =0.42 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =ꢀ4.4 (c=0.7, CHCl3);
1H NMR (500 MHz, CDCl3): d=7.59 (d, J=7.6 Hz, 4H; ArH), 7.55–7.52
(m, 4H; ArH), 7.46–7.27 (m, 22H; ArH), 7.21 (d, J=8.9 Hz, 2H; ArH),
7.16 (d, J=7.1 Hz, 2H; ArH), 7.09–7.01 (m, 5H; ArH, NH), 6.98–6.95
(m, 3H; ArH, NH), 5.514 (d, J=3.6 Hz, 1H; H4’’’), 5.508 (d, J=3.6 Hz,
1H; H4’), 5.06 (dd, J=10.4, 7.7 Hz, 1H; H2’’’), 5.06 (dd, J=10.4, 8.3 Hz,
1H; H2’ overlapped with H2’’’ and H1’’), 5.03 (d, J=8.3 Hz, 1H; H1’’),
4.95 (dd, J=10.4, 3.6 Hz, 1H; H3’’’), 4.80 (ABq, J=10.4 Hz, 1H;
PhCH2), 4.76 (d, J=7.2 Hz, 1H; H1), 4.72 (ABq, J=10.9 Hz, 1H;
PhCH2), 4.70 (ABq, J=11.7 Hz, 1H; PhCH2), 4.65 (ABq, J=11.7 Hz,
1H; PhCH2), 4.54 (d, J=7.7 Hz, 1H; H1’’’), 4.52 (ABq, J=10.9 Hz, 1H;
PhCH2), 4.49 (ABq, J=11.7 Hz, 1H; PhCH2), 4.46 (ABq, J=11.7 Hz,
1H; PhCH2), 4.44 (ABq, J=10.4 Hz, 1H; PhCH2), 4.35 (d, J=8.3 Hz,
1H; H1’), 3.98–3.92 (m, 3H; H4, H3’’, H4’’), 3.89 (t, J=7.2 Hz, 1H; H3),
3.85–3.74 (m, 5H; H6a’, alkyl chain, H6a,b, H3’), 3.71 (dd, J=10.4,
4.0 Hz, 1H; H6b’), 3.64 (s, 3H; COOCH3), 3.62–3.59 (m, 1H; H2), 3.60–
3.37 (m, 9H; H5, H5’, H5’’, H5’’’, H6a,b, H6a,b’’, alkyl chain), 3.36–3.32
(m, 1H; H2’’), 2.81 (ddd, J=18.2, 6.1, 5.3 Hz, 1H; Lev-CH2), 2.65–2.52
(m, 2H; Lev-CH2), 2.39 (dt, J=17.3, 6.1 Hz, 1H; Lev-CH2), 2.26 (t, J=
7.5 Hz, 2H; CH2COOCH3), 2.17 (s, 3H; CH2COCH3), 2.06 (s, 3H;
COCH3), 2.05 (s, 3H; COCH3), 2.03 (s, 3H; COCH3), 1.93 (s, 3H;
COCH3), 1.88 (s, 3H; COCH3), 1.63–1.54 (m, 6H; alkyl chain), 1.39–1.32
(m, 2H; alkyl chain), 1.01 (s, 9H; tBu), 1.00 ppm (s, 9H; tBu); 13C NMR
(125 MHz, CDCl3): d=206.5, 174.3, 171.8, 170.2, 169.8, 169.6, 169.5,
161.85, 161.81, 138.2, 138.12, 138.08, 135.9, 135.7, 133.2, 132.96, 132.94,
130.1, 130.03, 130.01, 128.7, 128.3, 128.25, 128.21, 128.13, 128.09, 128.04,
128.0, 127.97, 127.93, 127.7, 100.6, 100.5, 99.6, 98.8, 92.7, 92.6, 77.3, 76.5,
76.3, 75.7, 75.66, 75.4, 74.33, 74.29, 74.0, 73.8, 73.78, 73.3, 71.8, 71.6, 69.7,
69.0, 68.7, 68.6, 67.0, 61.5, 60.4, 57.6, 56.4, 51.7, 37.9, 34.1, 29.9, 29.3, 28.0,
27.0, 26.96, 25.8, 24.9, 21.4, 21.0, 20.9, 20.8, 19.3, 19.2 ppm; HRMS
(MALDI-TOF): m/z calcd for C108H128Cl6N2O29Si2Na: 2209.6158
[M+Na]+; found: 2209.6160.
2.8H; Lev-CH2), 2.17 (s, 5.6H; COCH
2.06 (s, 5.6H; COCH3(a)), 2.02 (s, 3H; COCH3(b)), 1.97 (s, 5.6H;
COCH3(a)), 1.93 (s, 3H; COCH3(b)), 1.01 ppm (s, 25.2H; tBu(a+b));
3ACHTUNGRTEN(NUNG a+b)), 2.16 (s, 3H; COCH3(b)),
AHCTUNGTRENNUNG
13C NMR (125 MHz, CDCl3): d=206.44, 206.41, 171.84, 171.79, 170.3,
170.2, 169.7, 169.2, 162.9, 162.0, 143.2, 137.98, 137.92, 137.8, 137.6, 135.86,
135.82, 135.76, 135.70, 133.03, 133.0, 132.8, 130.2, 130.13, 130.1, 130.08,
128.99, 128.96, 128.8, 128.62, 128.61, 128.5, 128.49, 128.3, 128.2, 128.17,
128.05, 128.00, 127.98, 127.5, 119.6, 104.3, 102.3, 100.4, 92.3, 86.5, 77.6,
76.2, 75.7, 74.9, 74.0, 73.73, 73.71, 73.3, 73.2, 72.1, 71.7, 71.6, 71.1, 69.7,
69.1, 68.8, 67.1, 67.05, 66.9, 66.3, 60.8, 60.6, 54.0, 37.9, 37.89, 29.88, 29.86,
28.07, 28.02, 27.0, 26.9, 21.1, 20.9, 20.84, 20.81, 19.23, 19.19 ppm; HRMS
(ESI-TOF): m/z calcd for C61H65Cl3F3N2O15Si: 1255.3166 [MꢀH]+; found:
1255.3258.
5-(Methoxycarbonyl)pentyl (2,4-di-O-acetyl-6-O-tert-butyldiphenylsilyl-b-
d-galactopyranosyl)-(1!4)-2-trichloroacetamido-3,6-di-O-benzyl-2-deoxy-
b-d-glucopyranoside (30)
A suspension of glycosyl donor 29 (150 mg, 0.12 mmol), methyl 6-hydrox-
yhexanoate (26 mg, 0.18 mmol), and 3 ꢁ molecular sieves (350 mg, fresh-
ly dried prior to use) in anhydrous CH2Cl2 (2 mL) was stirred at RT for
30 min, cooled to 08C under a nitrogen atmosphere, and treated with
a catalytic amount of TMSOTf (6.5 mL, 36.0 mmol). The reaction was
stirred at 08C for 1 h, quenched by the addition of Et3N, and filtered
through a pad of Celite. The filtrate was washed with a saturated aqueous
solution of NaHCO3 and brine and dried over anhydrous MgSO4. The fil-
trate was evaporated in vacuo, redissolved in anhydrous THF/MeOH
(9:1, 2 mL), and treated with hydrazine monoacetate (22 mg, 0.24 mmol)
at 08C under a nitrogen atmosphere. Then, the ice bath was removed
and the mixture was stirred at RT for a further 2 h. The solution was ex-
tracted with EtOAc and the combined organic layer was washed with
a saturated aqueous solution of NaHCO3, water, and brine, dried over
MgSO4, filtered, and concentrated in vacuo. The residue was purified by
column chromatography on silica gel (EtOAc/hexanes, 1:1) to give com-
pound 30 (95.9 mg, 72% in two steps) as a white amorphous foam. Rf =
0.28 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =ꢀ20 (c=1.1, CHCl3); 1H NMR
(500 MHz, CDCl3): d=7.57–7.56 (m, 4H; ArH), 7.47–7.42 (m, 2H;
ArH), 7.39–7.36 (m, 4H; ArH), 7.34–7.29 (m, 5H; ArH), 7.22–7.20 (m,
2H; ArH), 7.09–7.06 (m, 1H; ArH), 7.03 (d, J=7.2 Hz, 1H; NH), 7.03–
6.99 (m, 2H; ArH), 5.47 (d, J=3.3 Hz, 1H; H4’), 4.87 (dd, J=10.1,
7.8 Hz, 1H; H2’), 4.85 (d, J=7.50 Hz, 1H; H1), 4.81 (ABq, J=10.4 Hz,
1H; PhCH2), 4.71 (ABq, J=11.9 Hz, 1H; PhCH2), 4.48 (ABq, J=
6-Azidohexyl (2,4,-di-O-acetyl-6-O-tert-butyldiphenylsilyl-3-O-levulinoyl-
b-d-galactopyranosyl)-(1!3)-2-azido-4,6-O-benzylidene-2-deoxy-b-d-
glucopyranoside (33)
Rf =0.40 (EtOAc/hexanes, 1:1 v/v); ½aꢁ2D5 =ꢀ16 (c=1.6, CHCl3); H NMR
1
(500 MHz, CDCl3): d=7.53 (d, J=6.9 Hz, 2H; ArH), 7.48 (d, J=6.9 Hz,
2H; ArH), 7.45–7.40 (m, 2H; ArH), 7.37–7.32 (m, 6H; ArH), 7.24–7.14
(m, 3H; ArH), 5.69 (d, J=6.9 Hz, 1H; NH), 5.50 (d, J=3.4 Hz, 1H;
H4’), 5.41 (s, 1H; PhCH), 5.16 (d, J=8.5 Hz, 1H; H1), 5.14 (dd, J=10.2,
Chem. Asian J. 2013, 8, 1536 – 1550
1548
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
7.9 Hz, 1H; H2’), 4.99 (dd, J=10.2, 3.4 Hz, 1H; H3’), 4.76 (d, J=7.9 Hz,
1H; H1’), 4.63 (dd, J=9.3, 4.2 Hz, 1H; H3), 4.28 (dd, J=10.3, 4.5 Hz,
1H; H6a), 3.82 (td, J=10, 7 Hz, 1H; OCH2), 3.71–3.68 (m, 1H; H6b),
3.66 (t, J=4.2 Hz, H4), 3.64–3.56 (m, 3H; H5 and H6ab’), 3.51 (t, J=
7 Hz, 1H; OCH2), 3.50–3.47 (m, 1H; H5’), 3.26 (t, J=6.9 Hz, 2H;
CH2N3), 2.99 (dd, J=9.3, 8.5 Hz, 1H; H2), 2.82–2.75 (m, 1H; Lev-CH2),
2.63–2.50 (m, 2H; Lev-CH2), 2.41–2.36 (m, 1H; Lev-CH2), 2.16 (s, 3H;
COCH3), 2.09 (s, 3H; COCH3), 2.00 (s, 3H; COCH3), 1.90 (s, 3H;
COCH3), 1.60–1.57 (m, 4H; alkyl chain), 1.40–1.37 (m, 4H; alkyl chain),
0.98 ppm (s, 9H; tBu); 13C NMR (125 MHz, CDCl3): d=206.3, 171.7,
170.6, 170.1, 169.7, 137.1, 135.8, 135.6, 132.8, 132.5, 129.9, 129.1, 128.6,
127.8, 127.7, 125.8, 101.0, 100.5, 99.3, 80.9, 77.3, 77.0, 76.8, 72.7, 71.7, 70.0,
69.8, 68.7, 66.7, 65.9, 60.5, 58.8, 37.7, 31.9, 29.7, 29.7, 28.8, 27.8, 26.7, 26.4,
25.5, 23.7, 22.7, 20.9 ppm; HRMS (ESI-TOF): m/z calcd for
C52H68N4O15Si: 1017.1996 [M+Na]+; found: 1017.1990.
[4] a) C. I. Gama, S. E. Tully, N. Sotogaku, P. M. Clark, M. Rawat, N.
Vaidehi, W. A. Goddard, III, A. Nishi, L. C. Hsieh-Wilson, Nat.
Chem. Biol. 2006, 2, 467–473; b) S.-G. Lee, J. M. Brown, C. J.
Rogers, J. B. Matson, C. Krishnamurthy, M. Rawat, L. C. Hsieh-
Wilson, Chem. Sci. 2010, 1, 322–325; c) G. Despras, C. Bernard, A.
Perrot, L. Cattiaux, A. Prochiantz, H. Lortat-Jacob, J.-M. Mallet,
Chem. Eur. J. 2012, 19, 531–540.
[5] For representative reviews on heparan sulfate (HS) synthesis, see:
a) S. B. Dulaney, X. Huang, Adv. Carbohydr. Chem. Biochem. 2012,
67, 95–136 and references therein; b) C. Noti, P. H. Seeberger, in
Chemistry and Biology of Heparin and Heparan Sulfate (Eds.: H. G.
Garg, R. J. Linhardt, C. A. Hales), Elsevier, Oxford, 2005, pp. 79–
142; c) M. Petitou, C. A. A. van Boeckel, Angew. Chem. 2004, 116,
3180–3196; Angew. Chem. Int. Ed. 2004, 43, 3118–3133 and referen-
ces therein; d) J. D. C. Codꢆe, H. S. Overkleeft, G. A. van der Marel,
C. A. A. van Boeckel, Drug Discovery Today Technol. 2004, 1, 317–
326; e) L. Poletti, L. Lay, Eur. J. Org. Chem. 2003, 2999–3024;
f) N. A. Karst, R. J. Linhardt, Curr. Med. Chem. 2003, 10, 1993–
2031.
[6] a) H. A. Orgueira, A. Bartolozzi, P. Schell, P. H. Seeberger, Angew.
Chem. 2002, 114, 2232–2235; Angew. Chem. Int. Ed. 2002, 41, 2128–
2131; b) C. Noti, J. L. de Paz, L. Polito, P. H. Seeberger, Chem. Eur.
J. 2006, 12, 8664–8686; c) A. Adibekian, P. Bindschꢄdler, M. S. M.
Timmer, C. Noti, N. Schꢇtzenmeister, P. H. Seeberger, Chem. Eur. J.
2007, 13, 4510–4522.
[7] a) S. Arungundram, K. Al-Mafraji, J. Asong, F. E. Leach III., I. J.
Amster, A. Venot, J. E. Turnbull, G.-J. Boons, J. Am. Chem. Soc.
2009, 131, 17394–17405; b) A. Prabhu, A. Venot, G.-J. Boons, Org.
Lett. 2003, 5, 4975–4978.
[8] a) M. M. L. Zulueta, S.-Y. Lin, Y.-T. Lin, C.-J. Huang, C.-C. Wang,
C.-C. Ku, Z. Shi, C.-L. Chyan, D. Irene, L.-H. Lim, T.-I. Tsai, Y.-P.
Hu, S. D. Arco, C.-H. Wong, S.-C. Hung, J. Am. Chem. Soc. 2012,
134, 8988–8995; b) Y.-P. Hu, S.-Y. Lin, C.-Y. Huang, M. M. L. Zu-
lueta, J.-Y. Liu, W. Chang, S.-C. Hung, Nat. Chem. 2011, 3, 557–563;
c) J.-C. Lee, X.-A. Lu, S. S. Kulkarni, Y.-S. Wen, S.-C. Hung, J. Am.
Chem. Soc. 2004, 126, 476–477; d) L.-D. Lu, C.-R. Shie, S. S. Kulkar-
ni, G.-R. Pan, X.-A. Lu, S.-C. Hung, Org. Lett. 2006, 8, 5995–5998.
[9] a) B. Yang, K. Yoshida, Z. Yin, H. Dai, H. Kavunja, M. H. El-Dak-
douki, S. Sungsuwan, S. B. Dulaney, X. Huang, Angew. Chem. 2012,
124, 10332–10336; Angew. Chem. Int. Ed. 2012, 51, 10185–10189;
b) G. Tiruchinapally, Z. Yin, M. El-Dakdouki, Z. Wang, X. Huang,
Chem. Eur. J. 2011, 17, 10106–10112.
6-Azidohexyl (2,4-di-O-acetyl-6-O-tert-butylphenylsilyl-3-O-levulinoyl-b-
d-galactopyranosyl)-(1!4)-2-acetamido-3,6-di-O-acetyl-2-deoxy-b-d-
glucopyranoside (34)
Rf =0.40 (EtOAc/hexanes, 2:1 v/v); ½aꢁ2D5 =ꢀ28 (c=1.5, CHCl3); H NMR
1
(500 MHz, CDCl3): d=7.62–7.56 (m, 4H; ArH), 7.46–7.36 (m, 6H;
ArH), 5.64 (d, J=9.3 Hz, 1H), 5.54 (d, J=2.7 Hz, 1H), 5.09–4.96 (m,
3H), 4.48 (dd, J=12.1, 3.1 Hz, 1H), 4.43 (d, J=7.4 Hz, 1H), 4.37 (d, J=
7.4 Hz, 1H), 4.09 (dd, J=5.1, 12.1 Hz, 1H), 4.01–3.96 (m, 1H), 3.84–3.76
(m, 1H; OCH2), 3.74–3.65 (m, 3H), 3.60–3.55 (m, 2H), 3.18–3.37 (m,
1H; OCH2), 3.25 (d, J=7.0 Hz, 2H), 2.82–2.76 (m, 1H; Lev-CH2), 2.65–
2.51 (m, 2H; Lev-CH2), 2.42–2.37 (m, 1H; Lev-CH2), 2.16 (s, 3H;
COCH3), 2.09 (s, 3H; COCH3), 2.08 (s, 3H; COCH3), 2.02 (s, 3H;
COCH3), 1.92 (s, 3H; COCH3), 1.76 (s, 3H; COCH3), 1.59–1.51 (m, 4H),
1.36–1.31 (m, 4H), 1.02 ppm (s, 9H; tBu); 13C NMR (125 MHz, CDCl3):
d=206.1, 171.6, 170.4, 170.3, 169.9, 169.7, 135.5, 135.4, 132.6, 132.5,
129.94, 129.93, 127.8, 127.76, 127.68, 100.9, 100.8, 75.6, 73.1, 72.5, 72.4,
71.3, 69.1, 66.5, 62.3, 60.6, 52.8, 51.3, 37.6, 29.6, 29.2, 28.7, 27.7, 26.6, 26.3,
25.4, 23.1, 20.8, 20.6, 20.56, 20.51, 18.9 ppm; HRMS (ESI-TOF): m/z
calcd for C49H68N4O17SiNa: 1035.4241 [M+Na]+; found: 1035.4244.
Acknowledgements
This work was supported by the Academia Sinica (AS-99-TP-AB4 to
K.I.L. and C.H.L.) and by the National Science Council of Taiwan (NSC
99-3112-B-001-004 to K.I.L. and NSC 99-2113M-001-006-MY3 to
C.H.L.).
[10] a) S. Komba, H. Ishida, M. Kiso, A. Hasegawa, Carbohydr. Res.
1996, 285, C1–C8; b) M. R. Pratt, C. R. Bertozzi, Org. Lett. 2004, 6,
2345–2348.
[11] a) A. V. Demchenko in General Aspects of the Glycosidic Bond For-
mation in Handbook of Chemical Glycosylation (Ed.: A. V. Dem-
chenko), Wiley-VCH, Weinheim, 2008, pp. 1; b) X. Zhu, R. R.
Schmidt, Angew. Chem. 2009, 121, 1932–1967; Angew. Chem. Int.
Ed. 2009, 48, 1900–1934.
[12] a) H. A. Orgueira, A. Bartolozzi, P. Schell, R. E. J. N. Litjens, E. R.
Palmacci, P. H. Seeberger, Chem. Eur. J. 2003, 9, 140–169;
b) J. D. C. Codꢆe, B. Stubba, M. Schiattarella, H. S. Overkleeft,
C. A. A. van Boeckel, J. H. van Boom, G. A. van der Marel, J. Am.
Chem. Soc. 2005, 127, 3767–3773.
[1] a) U. Hꢄcker, K. Nybakken, N. Perrimon, Nat. Rev. Mol. Cell Biol.
2005, 6, 530–541; b) R. Sasisekharan, Z. Shriver, G. Venkataraman,
U. Narayanasami, Nat. Rev. Cancer 2002, 2, 521–528; c) H. E.
Conrad, in Heparin Binding Proteins, Academic Press, New York,
1998; d) D. Van Vactor, D. P. Wall, K. G. Johnson, Curr. Opin. Neu-
robiol. 2006, 16, 40–51; e) J. R. Bishop, M. Schuksz, J. D. Esko,
Nature 2007, 446, 1030–1037; f) A. Ori, M. C. Wilkinson, D. G.
Fernig, Front. Biosci. 2008, 13, 4309–4338; g) Z. Johnson, A. E.
Proudfoot, T. M. Handel, Cytokine Growth Factor Rev. 2005, 16,
625–636; h) C. R. Parish, Nat. Rev. Immunol. 2006, 6, 633–643;
i) L. R. Zacharski, A. Y. Lee, Expert Opin. Invest. Drugs 2008, 17,
1029–1037; j) H. Kawashima, Trends Glycosci. Glycotechnol. 2010,
22, 211–225; k) K. Sakamoto, K. Kadomatsu, Trends Glycosci. Gly-
cotechnol. 2011, 23, 212–220.
[13] See the Supporting Information, Scheme 1.
[14] a) B. Mukhopadhyay, R. A. Field, Carbohydr. Res. 2003, 338, 2149–
2152; b) C.-C. Wang, J.-C. Lee, S.-Y. Luo, S. S. Kulkarni, Y.-W.
Huang, C.-C. Lee, K.-L. Chang, S.-C. Hung, Nature 2007, 446, 896–
899; c) A. FranÅais, D. Urban, J.-M. Beau, Angew. Chem. 2007, 119,
8816–8819; Angew. Chem. Int. Ed. 2007, 46, 8662–8665; d) Y.
Vohra, M. Vasan, A. Venot, G.-J. Boons, Org. Lett. 2008, 10, 3247–
3250; e) M. Zhou, G. A. OꢃDoherty, Curr. Top. Med. Chem. 2008, 8,
114–125; f) M. Guillemineau, F.-I. Auzanneau, J. Org. Chem. 2012,
77, 8864–8878.
[2] P. Stanley, R. D. Cummings in Essentials of Glycobiology, 2nd ed.
(Eds.: A. Varki, R. D. Cummings, J. D. Esko, H. H. Freeze, P. Stan-
ley, C. R. Bertozzi, G. W. Hart, M. E. Etzler), Cold Spring Harbor
Laboratory Press, New York, 2008, pp. 196.
[15] For recent reviews of glucosamie-based glycosylation reactions, see:
a) A. F. G. Bongat, A. V. Demchenko, Carbohydr. Res. 2007, 342,
374–406; b) R. Enugala, L. C. R. Carvalho, M. J. Dias Pires,
M. M. B. Marques, Chem. Asian J. 2012, 7, 2482–2501.
[3] a) H. J. Allen, H. Ahmed, K. L. Matta, Glycoconjugate J. 1998, 15,
691–695; b) H. Ideo, A. Seko, T. Ohkura, K. L. Matta, K. Yamashi-
ta, Glycobiology 2002, 12, 199–208; c) C. T. ꢅberg, A.-L. Noresson,
H. Leffler, U. J. Nilsson, Chem. Eur. J. 2011, 17, 8139–8144.
Chem. Asian J. 2013, 8, 1536 – 1550
1549
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemasianj.org
Kuo-I Lin, Chun-Hung Lin et al.
[16] R. J. Williams, N. W. McGill, J. M. White, S. J. Williams, J. Carbo-
hydr. Chem. 2010, 29, 236–263 and references therein.
[17] M. Palme, A. Vasella, Helv. Chim. Acta 1995, 78, 959–961.
[18] X.-A. Lu, C.-H. Chou, C.-C. Wang, S.-C. Hung, Synlett 2003, 1364–
1366.
[24] a) C.-M. Tsai, C.-H. Guan, H.-W. Hsieh, T.-L. Hsu, Z. Tu, K.-J. Wu,
C.-H. Lin, K.-I. Lin, J. Immunol. 2011, 187, 1643–1652; b) C.-M.
Tsai, Y.-K. Chiu, T.-L. Hsu, I.-Y. Lin, S.-L. Hsieh, K.-I. Lin, J. Im-
munol. 2008, 181, 4570–4579.
[25] a) H. Ideo, T. Matsuzaka, T. Nonaka, A. Seko, K. Yamashita, J.
Biol. Chem. 2011, 286, 11346–11355; b) S. R. Stowell, C. M. Arthur,
K. A. Slanina, J. R. Horton, D. F. Smith, R. D. Cummings, J. Biol.
Chem. 2008, 283, 20547–20559; c) T. Horlacher, M. A. Oberli, D. B.
Werz, L. Krçck, S. Bufali, R. Mishra, J. Sobek, K. Simons, M. Hira-
shima, T. Niki, P. H. Seeberger, ChemBioChem 2010, 11, 1563–1573.
[26] W. C. Still, M. Kahn, A. Mitra, J. Org. Chem. 1978, 43, 2923–2925.
[27] a) Drying was performed on the Solvent Purification System SD-300
(Asiawong Enterprise Co. Ltd.); b) A. B. Pangborn, M. A. Giardel-
lo, R. H. Grubbs, R. K. Rosen, F. J. Timmers, Organometallics 1996,
15, 1518–1520.
[19] For the detailed synthesis of compounds 13 and 20, see the Support-
ing Information, Scheme 3.
[20] a) K.-K. T. Mong, C.-Y. Hung, C.-H. Wong, J. Org. Chem. 2003, 68,
2135–2142; b) G. Srivastava, O. Hindsgaul, J. Carbohydr. Chem.
1991, 10, 927–933; c) H. Shimizu, Y. Ito, O. Kanie, T. Ogawa,
Bioorg. Med. Chem. Lett. 1996, 6, 2841–2846; d) K. M. Koeller, C.-
H. Wong, Chem. Eur. J. 2000, 6, 1243–1251; e) J. Alais, A. Veyr-
ieꢈres, Carbohydr. Res. 1990, 207, 11–31; f) G. Kretzschmara, W.
Stahl, Tetrahedron 1998, 54, 6341–6358.
[21] S.-M. Chang, Z. Tu, H.-M. Jan, J.-F. Pan, C.-H. Lin, Chem.
Commun. 2013, DOI: 10.1039/C2CC37099A.
[28] C.-A. Tai, S. S. Kulkarni, S.-C. Hung, J. Org. Chem. 2003, 68, 8719–
8722.
[29] C.-H. Wong, X.-S. Ye, Z.-Y. Zhang, J. Am. Chem. Soc. 1998, 120,
7137–7138.
[22] a) We indeed encountered some difficulties in the subsequent depro-
tection and sulfation steps and we had to make extra effort to syn-
thesize compounds 33 and 34 to afford the final sulfated disacchar-
ides; see the synthetic procedures in the Supporting Information,
pp. S8–9. b) For the synthetic procedure, see the Supporting Infor-
mation, Scheme 5.
Received: December 19, 2012
Revised: March 6, 2013
Published online: May 2, 2013
[23] C.-F. Chang, J.-F. Pan, C.-N. Lin, I.-L. Wu, C.-H. Wong, C.-H. Lin,
Glycobiology 2011, 21, 895–902.
Chem. Asian J. 2013, 8, 1536 – 1550
1550
ꢀ 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim