First synthesis of 5,6-branched galacto-hexasaccharide, the dimer of the trisaccharide repeating unit of the cell-wall galactans of Bifidobacterium catenulatum YIT 4016

Article abstract of DOI:10.1016/j.tetasy.2004.12.014

α-d-Galactopyranosyl-(1→6)-[β-d-galactofuranosyl-(1→5) ]-β-d-galactofuranosyl-(1→6)-β-d-galactofuranosyl-(1→5) -[α-d-galactopyranosyl-(1→6)]-β-d-galactofuranose, the dimer of the trisaccharide repeating unit of the cell-wall galactans of Bifidobacterium c

Full text of DOI:10.1016/j.tetasy.2004.12.014

Tetrahedron:
Asymmetry
Tetrahedron: Asymmetry 16 (2005) 733–738
First synthesis of 5,6-branched galacto-hexasaccharide, the dimer
of the trisaccharide repeating unit of the cell-wall galactans of
Bifidobacterium catenulatum YIT 4016
Guohua Zhang, Mingkun Fu and Jun Ning*
Research Center for Eco-Environmental Sciences, Academia Sinica, PO Box 2871, Beijing 100085, PR China
Received 7 December 2004; accepted 24 December 2004
Available online 26 January 2005
Abstract—a-D-Galactopyranosyl-(1!6)-[b-D-galactofuranosyl-(1!5)]-b-D-galactofuranosyl-(1!6)-b-D-galactofuranosyl-(1!5)-[a-
D-galactopyranosyl-(1!6)]-b-D-galactofuranose, the dimer of the trisaccharide repeating unit of the cell-wall galactans of Bifidobac-
terium catenulatum YIT 4016, has been synthesized as its dodecyl glycoside 2 by coupling of 2,3,4,6-tetra-O-benzyl-a-D-galactopyr-
anosyl-(1!6)-[6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl-(1!5)]-2-O-acetyl-3-O-benzyl-b-D-galactofuranosyl trichloro-
acetimidate 14 with dodecyl 2,3,4,6-tetra-O-benzyl-a-D-galactopyranosyl-(1!6)-[2,3,5-tri-O-benzoyl-b-D-galactofuranosyl-(1!5)]-
2-O-acetyl-3-O-benzyl-b-D-galactofuranoside 16. The trisaccharide trichloroacetimidate donor 14 and trisaccharide acceptor 16
were regiospecifically prepared by employing 3-O-benzyl-1,2-O-isopropylidene-a-D-galactofuranose 4 as the glycosyl acceptor,
and isopropyl 2,3,4,6-tetra-O-benzyl-1-thio-b-D-galactopyranoside 5 and 6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl tri-
chloroacetimidate 9 as glycosyl donors.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
3,6-branched
saccharides.7,8
and
5,6-branched
galacto-oligo-
Recent demonstrations that oligosaccharides play
important roles in diverse biological events have resulted
in renewed interest in the synthesis of oligosacchar-
ides.1,2 However, compared with other biopolymers
such as peptides and nucleic acids, the role of oligosac-
charide structure in function has been minimally studied
owing to the wide structural and stereochemical diversi-
fication. Although many advances have been made over
recent decades,3 the clear facts remain that we have sin-
gularly failed to develop a general solution and glycosyl-
ation chemistry is still not predictable or generally
accessible. To facilitate the synthesis of target oligosac-
charides, regioselective glycosylation of glycosyl donors
with unprotected or partially protected sugar acceptors
has been extensively studied. Employing this strategy,
in the last few years, we have prepared a lot of oligosac-
charides with various structures present in natural
sources such as 3,6-branched gluco-oligosaccharides,4
2,6-branched manno-oligosaccharides,5 2,6-branched,6
Bifidobacterium, a bacteria living in the mammalian
intestines is thought to be helpful in the protection of
hosts from bacterial infections, suppression of enteric
putridity and the supply of several kinds of vitamin,
etc.9 As the major components of the cell wall, the galac-
tans play important roles in biological activities. In
1996, the structure of the galactan 1 from the cell wall
of Bifidobacterium catenulatum YIT 4016 was elucidated
by Nagaoka et al. (Fig. 1).10 This galactan is composed
6
1 5
1
Galf
)
(
Galf
β
β
β
6
α
1
Galp
1
5
1
5
1
6
β
1
Galf
Galf
Galf
Galf
ODodecyl
β
β
6
6
α
α
1
Galp
1
Galp
2
*
Figure 1. Cell-wall galactan of Bifidobacterium catenulatum YIT 4016
1 and the synthesized hexasaccharide 2.
Corresponding author. Tel.: +86 10 6284 9157; fax: +86 10 6292
3563; e-mail: jning@mail.rcees.ac.cn
0957-4166/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2004.12.014

734
G. Zhang et al. / Tetrahedron: Asymmetry 16 (2005) 733–738
of b(1!5) linked galactofuranose residues, while the
branches consist of galactopyranosyl residues. From a
synthetic standpoint, this structure is extremely compli-
cated because both the 5- and 6-positions of the reduc-
ing galactofuranose component are linked with
galactofuranosyl and galactopyranosyl residues, and
the glycosidic linkage of the galactopyranose moiety
is a.
product for another donor–acceptor pair. Furthermore,
the stereochemical outcome is often difficult to predict.
In our synthesis, 1,2:5,6-di-O-isopropylidene-a-D-galac-
tofuranose 3 was a key starting material, which was pre-
pared by our improved method.14 Benzylation of 3 in
DMF with PhCH2Br at rt, followed by selective 5,6-O-
deacetonation with 90% AcOH at 40 ꢁC afforded 3-O-
benzyl-1,2-O-isopropylidene-a-D-galactofuranose
4
(Scheme 1). For stereoselectively forming a-galactopyr-
anosides in glycosylation reactions, a general method
was using the glycosyl donor possessing a benzyl group
at the C-2 position. However, when we used the same
rule to synthesize our desired a-(1!6)-linked galacto-
disaccharide we found that there was nearly no stereose-
lectivity. Coupling of 4 with isopropyl 2,3,4,6-tetra-O-
benzyl-1-thio-b-D-galactopyranoside 515 gave a mixture
consisting of a-(1!6)-linked and b-(1!6)-linked disac-
charides in a ratio of 1:1.25. What made the thing more
difficult was that the mixture of the two disaccharides
could not be separated. Fortunately, we found that after
acetylation of the a- and b-linked disaccharides, the
desired product, 2,3,4,6-tetra-O-benzyl-a-D-galactopyr-
anosyl-(1!6)-5-O-acetyl-3-O-benzyl-1,2-O-isopropylid-
ene-a-D-galactofuranose 6 could be isolated in a poor
yield (27%). Through comparison with the spectrum of
its isomer, 2,3,4,6-tetra-O-benzyl-b-D-galactopyranosyl-
(1!6)-5-O-acetyl-3-O-benzyl-1,2-O-isopropylidene-a-D-
galactofuranose 7, the structure of compound 6 was
unambiguously identified by 1H NMR data. The charac-
teristic resonances due to the anomeric protons H-1a,
H-1b were located as a doublet at 4.94 ppm with
J1,2 = 3.7 Hz and as a doublet at 4.40 ppm with
J1,2 = 7.6 Hz, respectively. Deacylation of 6 in ammo-
nia-saturated methanol furnished 8. Coupling of 6-O-
acetyl-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl trichlo-
roacetimidate 916–18 and 8 with TMSOTf as catalyst in
CH2Cl2 at rt gave 2,3,4,6-tetra-O-benzyl-a-D-galacto-
pyranosyl-(1!6)-[6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-
galactofuranosyl-(1!5)]-3-O-benzyl-1,2-O-isopropylid-
In addition to the extremely complex structure, there are
other interests promoting us to synthesize the oligomer
of this trisaccharide repeating unit. First, the backbone
of the b(1!5) linked galactofuranose chain is of immu-
nological importance. For example, studies on Aspergil-
lus cell wall antigens showed that the b(1!5) linked
galactofuranose chains were regarded to be the immu-
nodominant etitopes.11 Second, the fact that galacto-
furanose residues have not been found in mammalian
glycoconjugates implied that these oligosaccharides
could be potential diagnostic agent and therapeutic drug
for intestinal disease that should have limited side effect
in mammalian cells.12 Third, the synthesis of target com-
pound is useful for further elucidating the molecular
structure responsible for the biological activities. These,
together with the fact that synthesis of this kind of oligo-
saccharide has not been reported so far, promote us to
develop a method for the synthesis of the dimer hexasac-
charide of the trisaccharide repeating unit of the cell-
wall galactans of Bifidobacterium catenulatum YIT 4016.
2. Results and discussion
Preparation of oligosaccharides is one of the most diffi-
cult and complex tasks in organic chemistry. Most gly-
cosylation methods are extremely sensitive to
structural variations in the glycosyl donor–acceptor
pairs.13 Reaction conditions that provide excellent yields
with one donor–acceptor pair may give virtually no
O
O
OBn
O
OH
O
a
O
O
O
O
OH
OH
4
3
O
O
BnO
BnO
BnO
BnO
BnO
OBn
OBn
O
O
c
O
O
O
O
BnO
OAc
OH
BnO
BnO
OBn
OBn
BnO
O
O
O
4
O
OBn
6
8
S
b
OBn
O
BnO
BnO
BnO
O
O
OBn
7
5
OAc
O
Scheme 1. Reagents and conditions: (a) i. PhCH2Br, NaH, DMF, rt, overnight; ii. 90% HOAc, 40 ꢁC, 24 h, 92% (over two steps); (b) i. 4, CH2Cl2,
TMSOTf (cat.), NIS, rt, 1 h; ii. Ac2O, pyridine, rt, 2 h, 27% for 6 and 34% for 7 (over two steps); (c) CH2Cl2–CH2OH saturated with ammonia, rt,
12 h, 92%.

G. Zhang et al. / Tetrahedron: Asymmetry 16 (2005) 733–738
735
O
BnO
BnO
OR1
O
OBn
O
OCNHCCl3
OBz
BnO
BnO
BnO
O
O
OBn
O
O
O
BnO
O
OBz
OBn
b
OR2
O
O
a
O
OBn
BzO
O
OBz
OAc
BzO
BzO
BzO
AcO
BzO
BzO
R3O
9
10
11 R1=H, R2=H, R3=Ac
12 R1= R2=R3=Ac
c
d
e
13 R1=H, R2=R3=Ac
14 R1=CNHCCl3, R2=R3=Ac
15 R1=C12H25, R2=R3=Ac
16 R1=C12H25, R2=Ac,R3=H
f
g
OC12H25
O
BnO
BnO
BnO
OBn
O
OAc
O
O
OBn
O
BzO
h
BzO
O
a
2
14
16
+
BzO
O
BnO
BnO
OBn
O
OAc
O
BnO
O
OBn
O
BzO
BzO
BzO
AcO
17
Scheme 2. Reagents and conditions: (a) CH2Cl2, TMSOTf (cat.), rt, 1 h, 95% for 10, 91% for 17; (b) 10:1 CHCl3–CF3COOH, rt, 2 h; (c) Ac2O,
pyridine, rt, 2 h; (d) benzylamine, THF, under darkness, rt, 24 h; (e) CCl3CN, CH2Cl2, K2CO3, rt, 12 h, 75% (over four steps); (f) C12H25OH,
TMSOTf (cat.), CH2Cl2, rt, 1 h, 97%; (g) 0.5% HCl in MeOH, rt, 10 h, 56%; (h) i. 1:4 MeOH/EtOAc, Pd/C (5%), rt, 24 h; ii. saturated with dry NH3,
rt, 24 h, 80% (over two steps).
ene-a-D-galactofuranose 10 in 95% yield. The structure
the chemical shifts of anomeric carbons of 17 revealed
by 13C NMR spectrum were at 106.34, 106.20, 106.07,
105.75, 97.94 and 97.81 ppm. Finally, deprotection of
17 yielded the target compound 2. The 13C NMR spec-
trum of 2 gave six signals for C-1 (108.03, 107.51,
107.40, 107.11, 98.50 and 98.50 ppm) confirming the
structure of 2.
1
of 10 was confirmed by H NMR and 13C NMR data.
1H NMR revealed three anomeric proton signals at
5.82 ppm as a doublet with J1,2 = 4.1 Hz, 4.84 ppm as
a doublet with J1,2 = 3.6 Hz and 5.66 ppm as a singlet,
meanwhile the 13C NMR spectrum revealed anomeric
carbons at 105.16, 104.63 and 97.88 ppm (Scheme 2).
De-isopropylidenation of 10 in 10:1 CHCl3–CF3COOH
(v/v) at rt offered 11. Acetylation of 11 with acetic anhy-
dride in pyridine gave 12. Selective 1-O-deacetylation of
12 with PhCH2NH2 in THF gave 13. After treating 13
with trichloroacetonitrile in the presence of K2CO3,
the desired trisaccharide glycosyl donor 14 was ob-
tained. Coupling of 14 with C12H25OH gave 15. There
was no difficulty for selective 6-O-deacetylation of 15
in MeOH solution containing 0.5% HCl to give the gly-
cosyl acceptor 16 in an acceptable yield (56%). Conden-
sation of 14 and 16 offered dodecyl 2,3,4,6-tetra-O-
benzyl-a-D-galactopyranosyl-(1!6)-[6-O-acetyl-2,3,5-tri-
O-benzoyl-b-D-galactofuranosyl-(1!5)]-2-O-acetyl-3-O-
benzyl-b-D-galactofuranosyl-(1!6)-2,3,5-tri-O-benzoyl-
b-D-galactofuranosyl-(1!5)-[2,3,4,6-tetra-O-benzyl-a-D-
galactopyranosyl-(1!6)]-2-O-acetyl-3-O-benzyl-b-D-
galactofuranoside 17. The 1H NMR spectrum of 17
contained structurally characteristic information: three
acetyl signals (d 1.89, 1.80 and 1.70 ppm). In addition,
3. Conclusion
In summary, synthesis of the dimer of the trisaccharide
repeating unit of the cell-wall galactans of Bifidobacte-
rium catenulatum YIT 4016 has been achieved for the
first time by regioselective glycosylation using partially
protected sugars as the acceptor. This should promote
the studies on fundamental biochemical properties and
biological functions about this oligosaccharide.
4. Experimental section
4.1. General methods
Optical rotations were determined at 25 ꢁC with a digital
polarimeter. The NMR spectra were recorded in CDCl3
with TMS internal standard or D2O with ethanol as

736
G. Zhang et al. / Tetrahedron: Asymmetry 16 (2005) 733–738
standard on ARX 400 MHz. Mass spectra were re-
corded on an autospec mass spectrometer using ESI
technique to introduce the sample.
1H NMR (400 MHz, CDCl3): d 7.35–7.25 (m, 25H,
5PhH), 5.72 (d, 1H, J = 4.1 Hz, H-10), 5.32–5.28 (m,
1H, H-50), 4.94 (d, 1H, J = 3.7 Hz, H-1), 4.93–4.25 (m,
12H, H-30, H-40, 5PhCH2), 4.05–4.01 (dd, 1H,
J = 3.7 Hz, 9.8 Hz, H-6a0), 3.93–3.86 (m, 4H, H-6b0,
H-20, H-2, H-4), 3.83–3.79 (dd, 1H, J = 6.0 Hz,
10.8 Hz, H-6a), 3.69–3.64 (dd, 1H, J = 5.6 Hz, 11.0 Hz,
H-6b), 3.50–3.48 (m, 2H, H-5, H-3), 1.98 (s, CH3CO),
1.53 (s, 3H, (CH3)C), 1.35 (s, 3H, (CH3)C). Anal. Calcd
for C52H58O12: C, 71.38; H, 6.68. Found: C, 71.83; H,
6.77.
Elemental analyses were done on elemental analyzer
model 1108 EA. Thin-layer chromatography (TLC)
was performed on silica gel HF254 with detection by
charring with 30% (v/v) H2SO4 in MeOH or in some
cases by a UV detector. Column chromatography was
conducted by elution of a column (10 · 240 mm,
18 · 300 mm, 35 · 400 mm) of silica gel (100–200 mesh)
with EtOAc–petroleum ether (60–90 ꢁC) as the eluent.
Solutions were concentrated at <60 ꢁC under diminished
pressure. Dry solvents were distilled over CaH2 and
stored over molecular sieves.
4.4. 2,3,4,6-Tetra-O-benzyl-b-D-galactopyranosyl-(1!6)-
5-O-acetyl-3-O-benzyl-1,2-O-isopropylidene-a-D-galac-
tofuranose 7
4.2. 3-O-Benzyl-1,2-O-isopropylidene-a-D-galactofura-
nose 4
[a]D = +10.8 (c 1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): d 7.37–7.30 (m, 25H, 5PhH), 5.82 (d, 1H,
J = 4.0 Hz, H-10), 5.44–5.43 (m, 1H, H-50), 4.98–4.41
(m, 11H, 5PhCH2, H-40), 4.40 (d, 1H, J = 7.6 Hz, H-
1), 4.15–4.11 (m, 2H, H-4, H-20), 3.97–3.94 (m, 2H, H-
2, H-30), 3.87–3.84 (dd, 1H, J = 6.9 Hz, 10.6 Hz, H-
6a), 3.75–3.72 (dd, 1H, J = 6.9 Hz, 10.6 Hz, H-6b),
3.63–3.53 (m, 4H, H-3, H-5, 2H-60), 1.97 (s, 3H,
CH3CO), 1.60 (s, 3H, CH3(CH)), 1.41 (s, 3H,
CH3(CH)). Anal. Calcd for C52H58O12: C, 71.38; H,
6.68. Found: C, 71.89, H, 6.78.
To a solution of 3 (3.0 g, 12 mmol) in dry DMF (80 mL)
was added PhCH2Br (1.5 mL, 13 mmol) and NaH
(0.50 g) at 0 ꢁC. After stirring the mixture overnight at
rt, TLC (4:1 petroleum ether–EtOAc) indicated that
the reaction was complete. Water (10 mL) was added
to the reaction mixture, and then the mixture was
diluted with EtOAc and washed with water. The organic
phase was concentrated, and the resulting residue was
directly dissolved in 90% acetic acid solution (80 mL).
The mixture was kept at 40 ꢁC for 24 h and then concen-
trated to a residue under reduced pressure. The residue
was purified by column chromatography (3:1 petroleum
ether–EtOAc) to afford 4 (3.3 g, 92% for two steps):
[a]D = ꢀ11.2 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.36–7.25 (m, 5H, PhH), 5.91 (d, 1H,
J = 4.1 Hz, H-1), 4.68 (d, 1H, J = 3.5 Hz, H-3), 4.67–
4.54 (m, 2H, PhCH2), 4.14–4.11 (m, 1H, H-4), 4.00 (d,
1H, J = 2.8 Hz, H-2), 3.80–3.77 (m, 1H, H-5), 3.70–
3.67 (dd, 1H, J = 3.7 Hz, 11.7 Hz, H-6a), 3.60–3.56
(dd, 1H, J = 4.8 Hz, 11.7 Hz, H-6b), 2.55 (s, 1H, OH),
2.13 (s, 1H, OH), 1.52 (s, 3H, (CH3)C), 1.34 (s, 3H,
(CH3)C). Anal. Calcd for C16H22O6: C, 61.92; H, 7.15.
Found: C, 61.52; H, 7.34.
4.5. 2,3,4,6-Tetra-O-benzyl-a-D-galactopyranosyl-(1!6)-
3-O-benzyl-1,2-O-isopropylidene-a-D-galactofuranose 8
Compound 6 (2.4 g, 2.7 mmol) was dissolved in an
ammonia-saturated solution of 1:10 CH2Cl2–CH3OH
(55 mL) saturated with NH3 at rt. After 12 h, TLC
(2:1 petroleum ether–EtOAc) indicated that the reaction
was complete. The reaction mixture was concentrated,
and the residue was passed through a silica gel column
with 3:1 petroleum ether–EtOAc as the eluent to give
1
8 (2.1 g, 92%): [a]D = +14.7 (c 1.0, CHCl3); H NMR
(400 MHz, CDCl3): d 7.35–7.23 (m, 25H, 5PhH), 5.85
(d, 1H, J = 4.1 Hz, H-10), 4.93–4.36 (m, 13H, 5PhCH2,
H-1, H-30, H-40), 4.11–3.91 (m, 5H, H-6a0, H-6b, 2H-
2, H-4), 3.75–3.72 (dd, 1H, J = 6.0 Hz, 10.8 Hz, H-6a),
3.60–3.56 (dd, 1H, J = 4.3 Hz, 11.0 Hz, H-6b0), 3.52–
3.44 (m, 2H, H-3, H-5), 2.96 (d, 1H, J = 5.6 Hz, H-50),
1.53 (s, 3H, C(CH3)), 1.34 (s, 3H, C(CH3)). Anal. Calcd
for C50H56O11: C, 72.10; H, 6.78. Found: C, 71.86; H,
6.69.
4.3. 2,3,4,6-Tetra-O-benzyl-a-D-galactopyranosyl-(1!6)-
5-O-acetyl-3-O-benzyl-1,2-O-isopropylidene-a-D-galacto-
furanose 6
A solution of isopropyl 2,3,4,6-tetra-O-benzyl-1-thio-b-
D-galactopyranoside 5 (6.3 g, 10.5 mmol) and 3-O-benz-
yl-1,2-O-isopropylidene-a-D-galactofuranose
10.3 mmol) in anhydrous CH2Cl2 (40 mL) was stirred
4
(3.2 g,
4.6. 2,3,4,6-Tetra-O-benzyl-a-D-galactopyranosyl-(1!6)-
[6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl-
(1!5)]-3-O-benzyl-a-1,2-O-isopropylidene-D-galacto-
furanose 10
˚
with activated 4 A molecular sieves (2 g) and NIS
(2.3 g, 9.4 mmol) at rt for 20 min, and then TMSOTf
(15 lL) was added. The mixture was stirred for 1 h, at
the end of which time TLC (3:1 petroleum ether–
EtOAc) indicated that the reaction was complete. The
reaction mixture was neutralized with Et3N and filtered,
and the filtrate was concentrated. The residue was acet-
ylated with acetic anhydride (3 mL) in pyridine (30 mL)
for 2 h at rt and the mixture was concentrated to dry-
ness. Purification of the residue by silica gel column
chromatography (6:1 petroleum ether–EtOAc) gave
solid 6 (2.5 g, 27%): [a]D = +12.8 (c 1.0, CHCl3);
A solution of 6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galac-
tofuranosyl trichloroacetimidate 9 (1.7 g, 2.5 mmol)
and 8 (2.0 g, 2.4 mmol) in anhydrous CH2Cl2 (40 mL)
˚
was stirred with activated 4 A molecular sieves at rt
for 20 min, and then TMSOTf (15 lL) was added. The
mixture was stirred for 1 h, at the end of which time
TLC (2.5:1 petroleum ether–EtOAc) indicated that the
reaction was complete. The reaction mixture was neu-
tralized with Et3N, filtered and then the filtrate was con-

G. Zhang et al. / Tetrahedron: Asymmetry 16 (2005) 733–738
737
centrated. The resultant residue was subjected to the col-
umn chromatography with 2.5:1 petroleum ether–
EtOAc as the eluent to give 10 (3.1 g, 95%):
[a]D = +2.5 (c 1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): d 8.08–7.11 (m, 40H, 8PhH), 6.00–5.96 (m,
1H, H-500), 5.82 (d, 1H, J = 4.1 Hz, H-10), 5.66 (s, 1H,
H00-1), 5.52 (d, 1H, J = 9.2 Hz, H-300), 5.45 (s, 1H, H-
200), 4.92–4.84 (dd, 1H, J = 2.7 Hz, 5.3 Hz, H-400), 4.88
(d, 1H, J = 8.4 Hz, H-30), 4.85 (d, 1H, J = 3.6 Hz, H-
1), 4.74–4.36 (m, 13H, 5PhCH2, H-50, H-2, H-4), 4.14–
3.81 (m, 7H, 4H-6, H-40, H-3, H-20), 3.60–3.47 (m,
3H, H-5, 2H-6), 1.97 (s, 3H, CH3CO), 1.61 (s, 3H,
C(CH3)), 1.34 (s, 3H, C(CH3)). Anal. Calcd for
C79H80O20: C, 70.31; H, 5.98. Found: C, 70.62; H, 6.15.
TLC (3:1 petroleum ether–EtOAc) indicated that the
reaction was complete. Then the mixture was neutral-
ized with Et3N and filtered, and the filtrate was concen-
trated. Purification of the residue by column
chromatography (3:1 petroleum ether–EtOAc) gave 15
1
(2.0 g, 97%) as a syrup: [a]D = ꢀ8.6 (c 1.0, CHCl3); H
NMR (400 MHz, CDCl3): d 8.05–7.17 (m, 40H,
8PhH), 5.92–5.88 (m, 1H, H-500), 5.70 (s, 1H, H-10),
5.54 (s, 1H, H-100), 5.52 (d, 1H, J = 9.2 Hz, H-300), 5.10
(s, 1H, H-2), 4.95 (s, 1H, H-200), 4.88–4.21 (m, 16H),
4.11–4.00 (m, 3H), 3.94–3.86 (m, 3H), 3.61–3.25 (m,
5H, 3H-6, CH2C12H23), 1.98 (s, 3H, CH3CO), 1.96 (s,
3H, CH3CO), 1.59–0.86 (m, 23H, CH2C11H23). Anal.
Calcd for C90H102O21: C, 71.13; H, 6.76. Found: C,
70.92; H, 6.98.
4.7. 2,3,4,6-Tetra-O-benzyl-a-D-galactopyranosyl-(1!6)-
[6-O-acetyl-2,3,5-tri-O- benzoyl-b-D-galactofuranosyl-
(1!5)]-2-O-acetyl-3-O-benzyl-b-D-galactofuranosyl tri-
chloroacetimidate 14
4.9. Dodecyl 2,3,4,6-tetra-O-benzyl-a-D-galactopyrano-
syl-(1!6)]-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl-
(1!5)]-2-O-acetyl-3-O-benzyl-b-D-galactofuranoside 16
Compound 10 (3.1 g, 2.3 mmol) was treated with 10:1
CHCl3–CF3COOH (50 mL) at rt for 2 h, at the end of
which time TLC (2:1 petroleum ether–EtOAc) indicated
that the reaction was complete. The solution was diluted
with toluene (100 mL), and the mixture was concen-
trated under vacuum, and then the residue was acetyl-
ated with acetic anhydride (5 mL) in pyridine (50 mL)
for 2 h at rt. The resultant trisaccharide and benzyl-
amine (5 mL) in anhydrous THF (100 mL) was kept
under darkness at rt for 24 h, at the end of which time
TLC (2:1 petroleum ether–EtOAc) indicated that the
reaction was complete. The mixture was diluted with
CH2Cl2 (100 mL), and was washed with 1 N HCl and
saturated aq NaHCO3. The combined organic phrase
was dried (Na2SO4) and concentrated to dryness. The
resultant residue without purification was dissolved in
CH2Cl2 (50 mL), and then CCl3CN (0.55 mL, 5.5 mmol)
and K2CO3 (2.5 g) were added. The mixture was stirred
for 12 h at rt, and the solid material was filtrated off.
Concentration of the filtrate, followed by purification
on a silica gel column with 2:1 petroleum ether–EtOAc
as the eluent, gave the trisaccharide donor 14 (2.6 g,
Acetyl chloride (0.25 mL) was added to a solution of
compound 15 (1.5 g, 0.99 mmol) in CH3OH (50 mL)
and CH2Cl2 (2 mL), and the reaction was carried out
at rt for 10 h. TLC (2:1 petroleum ether–EtOAc) indi-
cated that the reaction was complete. After neutraliza-
tion and concentration, the residue was subjected to
column chromatography on silica gel using petroleum
ether–EtOAc (2:1) as the eluent to give 16 (0.82 g,
56%): [a]D = ꢀ15.6 (c 1.0, CHCl3); 1H NMR
(400 MHz, CDCl3): d 8.09–7.20 (m, 40H, 8PhH), 5.72
(s, 1H, H-10), 5.62–5.59 (m, 2H, H-5, H-1), 5.53 (d,
1H, J = 9.2 Hz, H-300), 5.11 (s, 1H, H-20), 4.98–3.26
(m, 28H), 2.00 (s, 3H, CH3CO), 1.96 (s, 3H, CH3CO),
1.59–0.84 (m, 23H, CH2C11H23). Anal. Calcd for
C88H100O20: C, 71.52; H, 6.82. Found: C, 71.78; H, 6.76.
4.10. Dodecyl 2,3,4,6-tetra-O-benzyl-a-D-galactopyrano-
syl-(1!6)-[6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galacto-
furanosyl-(1!5)]-2-O-acetyl-3-O-benzyl-b-D-galactofur-
anosyl-(1!6)-2,3,5-tri-O-benzoyl-b-D-galactofuranosyl-
(1!5)-[2,3,4,6-tetra-O-benzyl-a-D-galactopyranosyl-
(1!6)]-2-O-acetyl-3-O-benzyl-b-D-galactofuranoside 17
1
75%): [a]D = ꢀ1.5 (c 1.0, CHCl3); H NMR (400 MHz,
CDCl3): d 8.50 (s, 1H, O(CNH)CCl3), 7.92–7.14 (m,
40H, 8PhH), 6.28 (s, 1H, H-10), 5.91 (m, 1H, H-500),
5.66 (s, 1H, H-100), 5.65 (d, 1H, J = 8.4 Hz, H-300), 5.50
(s, 1H, H-200), 5.47 (d, 1H, J = 8.4 Hz, H-30), 5.38 (s,
1H, H-20), 4.87–4.25 (m, 15H), 4.05–3.87 (m, 6H, 4H-
6, H-4, H-3), 3.58–3.33 (m, 3H, H-5, 2H-6), 2.04 (s,
3H, CH3CO), 1.97 (s, 3H, CH3CO). Anal. Calcd for
C80H78Cl3NO21: C, 64.24; H, 5.26. Found: C, 64.67;
H, 5.41.
A solution of 14 (0.55 g, 0.37 mmol) and 16 (0.50 g,
0.34 mmol) in anhydrous CH2Cl2 (50 mL) was stirred
˚
with activated 4 A molecular sieves (2 g) at rt for
20 min, and then TMSOTf (12 lL) was added. The mix-
ture was stirred for 1 h, at the end of which time TLC
(1.5:1 petroleum ether–EtOAc) indicated that the reac-
tion was complete. The reaction mixture was neutralized
with Et3N and filtered, and the filtrate was concentrated.
Purification of the residue by column chromatography
(1.5:1 petroleum ether–EtOAc) gave 17 (0.87 g, 91%):
[a]D = ꢀ13.4 (c 1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): d 8.03–7.01 (m, 80H, 16PhH), 5.85 (m, 1H,
H-5), 5.74 (m, 1H, H-5), 5.70 (s, 1H, H-1), 5.69 (s, 1H,
H-1), 5.67–5.57 (m, 2H, H-1, H-2), 5.50–5.48 (m, 2H,
H-1, H-2), 5.12–3.20 (m, 56H), 1.89 (s, 3H, CH3CO),
1.80 (s, 3H, CH3CO), 1.70 (s, 3H, CH3CO), 1.26–0.87
(m, 23H, CH2C11H23); 13C NMR (100 MHz, CDCl3):
d 170.52, 169.96, 169.40 (3CH3CO), 165.68, 165.68,
165.59, 165.51, 165.17, 165.17 (6PhCO), 106.34,
106.20, 106.07, 105.75 (4b-C-1), 97.94, 97.81 (2a-C-1).
4.8. Dodecyl 2,3,4,6-tetra-O-benzyl-a-D-galactopyrano-
syl-(1!6)]-6-O-acetyl-2,3,5-tri-O-benzoyl-b-D-galacto-
furanosyl-(1!5)]-2-O-acetyl-3-O-benzyl-b-D-galactofur-
anoside 15
A solution of 14 (2.0 g, 1.3 mmol) and C12H25OH
(0.37 g, 2.0 mmol) in anhydrous CH2Cl2 (40 mL) was
˚
stirred with activated 4 A molecular sieves (2 g) at rt
for 20 min, and then TMSOTf (15 lL) was added. The
mixture was stirred for 1 h, at the end of which time

738
G. Zhang et al. / Tetrahedron: Asymmetry 16 (2005) 733–738
Anal. Calcd for C166H176O40: C, 70.92; H, 6.31. Found:
C, 70.58; H, 6.47.
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4.11. Dodecyl a-D-galactopyranosyl-(1!6)-[b-D-galacto-
furanosyl-(1!5)]-b-D-galactofuranosyl-(1!6)-b-D-galac-
tofuranosyl-(1!5)-[a-D-galactopyranosyl-(1!6)]-b-D-
galactofuranoside 2
To a solution of 17 (0.50 g, 0.18 mmol) in 1:4 MeOH/
EtOAc (40 mL) was added Pd/C (5%, 40 mg). The reac-
tion mixture was stirred for 24 h at rt in hydrogen atmo-
sphere, at the end of which time TLC indicated that the
debenzylation of 17 was complete. Then the mixture was
filtered, and the filtrate was concentrated under reduced
pressure to dryness. The residue was dissolved in a sat-
urated solution of NH3 in anhydrous CH3OH
(10 mL). After 24 h at rt, the reaction mixture was con-
centrated to about 5 mL, and then CH2Cl2 (40 mL) was
added. The resultant precipitate was filtered and washed
with CH2Cl2 (4 · 5 mL) to afford 2 (0.16 g, 80%):
[a]D = +2.4 (c 1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): d 5.26–4.74 (m, 6H, 6H-1), 1.28–1.12 (m,
23H, CH2C11H23); 13C NMR (100 MHz, D2O): d
108.03, 107.51, 107.40, 107.11 (4b-C-1), 98.50, 98.50
(2a-C-1). Anal. Calcd for C48H86O31: C, 49.73; H,
7.48. Found: C, 49.97; H, 7.35. MALDI-TOF MS:
Calcd for C48H86O31, 1159.18 [M]. Found: 1182.30
(M+Na)+.
15. Du, Y.; Zhang, M.; Yang, F.; Gu, G. J. Chem. Soc.,
Perkin Trans. 1 2001, 3122–3127.
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296.
This work was supported by the National Key Project
for Basic Research (2003CB114400), the Beijing Natural
Science Foundation (6021004) and The Ministry of Sci-
ence and Technology (2001AA246014).