Rapid synthesis of oligosaccharide moieties of globotriaosylceramide using fluorous protective group

Article abstract of DOI:10.1016/S0040-4039(03)00091-1

The use of the Bfp (bisfluorous chain type propanoyl) group as a fluorous protective group made it possible to rapidly synthesize galabiose and the Gb3 oligosaccharide derivatives by a simple fluorous-organic extraction purification. The fluorous oligosaccharide synthesis using the Bfp group is an excellent strategic alternative to solid phase oligosaccharide synthesis, and removes some of the disadvantages of the solid phase method.

Full text of DOI:10.1016/S0040-4039(03)00091-1

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 44 (2003) 1819–1821
Rapid synthesis of oligosaccharide moieties of
globotriaosylceramide using fluorous protective group
Tsuyoshi Miura and Toshiyuki Inazu*
The Noguchi Institute, 1-8-1 Kaga, Itabashi-ku, Tokyo 173-0003, Japan
Received 26 November 2002; revised 7 January 2003; accepted 10 January 2003
Abstract—The use of the Bfp (bisfluorous chain type propanoyl) group as a fluorous protective group made it possible to rapidly
synthesize galabiose and the Gb3 oligosaccharide derivatives by a simple fluorous–organic extraction purification. The fluorous
oligosaccharide synthesis using the Bfp group is an excellent strategic alternative to solid phase oligosaccharide synthesis, and
removes some of the disadvantages of the solid phase method. © 2003 Elsevier Science Ltd. All rights reserved.
Globotriaosyl ceramide (Gb3), known as a Pk antigen
in the P blood-group system,1 is a natural ligand of
Shiga-like toxins (Stx-I and Stx-II, or verotoxins) pro-
duced by Escherica coli O-157:H-7.2 Shiga-like toxins
mainly recognize a galactobiosyl a(1–4)-linkage (gal-
abiose moiety) in Gb3, and induce carbohydrate-medi-
ated internalization into the host cell.2 In order to
develop verotoxin inhibitors and detect the verotoxin,
galabiose and the Gb3 trisaccharide play an important
role as a key molecule.3 The synthesis of galabiose and
the Gb3 trisaccharide have already been accomplished
by several groups.4 However, each synthetic method
consumes much time and cost due to the purification
procedure such as column chromatography in multi-
steps. To provide a rapid synthesis, the solid phase
synthesis of oligosaccharides has been actively studied.5
The solid phase method suffers from some serious
disadvantages, such as the difficulty of a large scale
synthesis, reduced reactivity, and the inability to moni-
tor the reaction by TLC, NMR, and MS. Recently,
fluorous synthesis has been developed for use in several
fields by Curran et al.6 A highly fluorinated compound
is readily separated from nonfluorinated compounds by
a simple fluorous-organic phase separation. A highly
fluorinated compound is also soluble in common
organic solvents and can be measured by NMR and
MS as a single compound. Therefore, fluorous synthesis
has become an excellent strategic alternative to solid
phase synthesis. Recently, we reported the fluorous
oligosaccharide synthesis using the Bfp (bisfluorous
chain type propanoyl) group as a novel fluorous protec-
tive group.7 Use of the Bfp group made it possible to
rapidly synthesize a simple oligosaccharide (Glcb(1–
6)Glc linkage) by purification using fluorous-organic
solvent extraction. To demonstrate the usefulness of the
Bfp group as a fluorous tag, we attempted to synthesize
the oligosaccharide moieties of Gb3 as a more complex
bioactive carbohydrate molecule. We would like to
report the rapid synthesis of the oligosaccharide moi-
eties of Gb3 using the Bfp group as a fluorous protec-
tive group in this communication.
We first synthesized the galabiose derivative 8 as shown
in Scheme 1. The Bfp group was introduced to the two
hydroxyl functions of the galactose derivative 2 using
N,N%-dicyclohexylcarbodiimide (DCC) and 4-(dimethyl-
amino)pyridine (DMAP) to give 3.8 The benzylidene
group of 3 was removed by treatment of the camphor-
sulfonic acid (CSA) in MeOH-CHCl3 to afford 4.8 The
* Corresponding author. Tel.: +81-3-5944-3214; fax: +81-3-5944-3214;
e-mail: inz@noguchi.or.jp
0040-4039/03/$ - see front matter © 2003 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(03)00091-1

1820
T. Miura, T. Inazu / Tetrahedron Letters 44 (2003) 1819–1821
Scheme 1. Reagents and conditions: (a) 1a, DCC, DMAP, CH2Cl2, rt, 4 h; (b) CSA, CHCl3–MeOH, rt, 5 h; (c) 1a, DCC, DMAP,
,
CH2Cl2–EtOC4F9, −5°C, 21 h; (d) TMS–OTf, 4 A molecular sieves, ether–EtOC4F9, 0°C, 30 min; (e) NaOMe, Et2O–MeOH, rt,
1 h, 27% (in 5 steps); (f) Ac2O, py, rt, 12 h, 95%.
Bfp group was selectively introduced to the primary
hydroxyl function of 4 using DCC and DMAP to give
the fluorous glycosyl acceptor 5.8 The reaction of the
fluorous glycosyl acceptor 5 with an excess of the
glycosyl donor 6 (6 equiv.) in the presence of TMS-OTf
14 were extracted with FC-72 by being partitioned
between FC-72 and an organic solvent (toluene or
methanol), and were purified without silica gel column
chromatography. The Bfp group of 14 was removed by
treatment with sodium methoxide in MeOH-ether to
afford 15, which was extracted with MeOH by being
partitioned between FC-72 and MeOH. Finally, the
pure trisaccharide 15 was obtained by only one silica
gel column chromatography purification in the final
step for a 34% total yield from 10 (5 steps). The
trisaccharide 15 was introduced to 16 by the treatment
with acetic anhydride in pyridine, and its structure was
identified.
9
in ether–EtOC4F9 afforded selectively the a-linked
fluorous disaccharide 7.8 (No b-isomer could be
detected.) The fluorous intermediates 3, 4, 5, and 7 were
extracted with FC-7210 by being partitioned between
FC-72 and an organic solvent (toluene or methanol),
and were purified without silica gel column chromato-
graphy. The Bfp group of 7 was removed by treatment
with sodium methoxide in MeOH-ether to afford the
crude 8, which was extracted with MeOH by being
partitioned between FC-72 and MeOH. The methyl
ester 1b, which was obtained from the FC-72 layer, was
treated with aqueous NaOH to give 1a that was able to
be reused as the fluorous protective reagent. Finally,
the pure galabiose derivative 8 was obtained by only
one silica gel column chromatography purification in
the final step for a 27% total yield from 2 (5 steps). The
galabiose derivative 8 was introduced to the acetate 9
by the treatment with acetic anhydride in pyridine, and
its structure was identified.
In conclusion, the use of the Bfp group as a fluorous
protective group made it possible to rapidly synthesize
galabiose and the Gb3 oligosaccharide derivatives by a
fluorous-organic extraction purification. The fluorous
oligosaccharide synthesis can be applied to large scale
synthesis due to the liquid phase synthesis. Because
each synthetic intermediate containing the Bfp group
was monitored by TLC, NMR, and MS, the reaction
conditions in each synthetic step were able to be rapidly
optimized. The fluorous intermediates were also able to
be purified by silica gel column chromatography in the
case which needed purification. After optimization of
the reaction conditions in each step, the synthesis in
multisteps was accomplished by fluorous-organic parti-
tion purification without column chromatography. The
only final compounds, which were removed the Bfp
group, were purified by column chromatography on
silica gel. Therefore, the fluorous oligosaccharide syn-
thesis using the Bfp group is an excellent strategic
alternative to solid phase oligosaccharide synthesis, and
removes some of the disadvantages of the solid phase
method. Further application to the synthesis of a bioac-
tive carbohydrate and glycoconjugate is now in
progress.
Next, we synthesized the Gb3 trisaccharide derivative
15 as shown in Scheme 2. The Bfp group was intro-
duced to the four hydroxyl functions of the lactose
derivative 10 using DCC and DMAP to give 11.8 The
benzylidene group and trityl (Tr) group of 11 were
removed by treatment with HCl in AcOEt–EtOC4F9 to
afford 12.8 The benzoyl (Bz) group was selectively
introduced to the two primary hydroxyl functions of 12
to afford the fluorous glycosyl acceptor 13.8 The reac-
tion of the fluorous glycosyl acceptor 13 with an excess
of the glycosyl donor 6 (5 equiv.) in the presence of
TMS-OTf in ether-EtOC4F9 afforded selectively the
a-linked fluorous trisaccharide 14.8 (No b-isomer could
be detected.) The fluorous intermediates 11, 12, 13, and

T. Miura, T. Inazu / Tetrahedron Letters 44 (2003) 1819–1821
1821
f
Scheme 2. Reagents and conditions: (a) 1a, DCC, DMAP, CH2Cl2, rt, 3 h; (b) HCl, AcOEt–EtOC4F9, 0°C, 2 h; (c) BzCl, Et3N,
,
CH2Cl2–EtOC4F9, −20°C, 7 h; (d) TMS–OTf, 4 A molecular sieves, ether–EtOC4F9, 0°C, 20 min; (e) NaOMe, Et2O–MeOH, rt,
1 h, then silica gel chromatography, 34% from 10 (in 5 steps); (f) Ac2O, Py, rt, 12 h, 95%.
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This work was partly supported by a Grant-in-Aid for
Scientific Research (C) (No. 13680680), a Grant-in-Aid
for Encouragement of Young Scientists (No. 13771349)
from the Japan Society for the Promotion of Science, and
Takeda Science Foundation. This work was performed
through the Noguchi Fluorous Project by our institute.
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