Synthetic sulfated glucuronosyl paragloboside (SGPG) and its use for the detection of autoimmune peripheral neuropathies

Article abstract of DOI:10.1016/j.tet.2005.10.021

The sulfated glucuronosyl paragloboside SO₄-3-GlcA(β1-3) Gal-(β1-4)GlcNAc(β1-3)Gal-(β1-4)Glcβ(1-1′)Cer (SGPG) is a specific constituent of the peripheral nervous system involved in human dysimmune neuropathies. Although it is highly immunogenic in human pathology, it is not a major constituent of peripheral nerve in humans. Pure SGPG is very difficult to obtain from natural sources, but is needed for the design of efficient and reproducible immunoassays. We describe a new chemical synthesis of this important glycolipid with a stearic tail, including its complete characterization by mass spectrometry and its use in the detection of autoimmune neuropathies.

Full text of DOI:10.1016/j.tet.2005.10.021

Tetrahedron 62 (2006) 563–577
Synthetic sulfated glucuronosyl paragloboside (SGPG) and its use
for the detection of autoimmune peripheral neuropathies
a
b,c
Reynald Chevalier, Benoıt Colsch, Carlos Afonso,c Nicole Baumann,b
ˆ
Jean-Claude Tabetc and Jean-Maurice Malleta,
*
a
´
Ecole Normale Superieure, UMR CNRS 8642, 24, rue Lhomond, 75231 Paris cedex 05, France
`
b
ˆ ˆ
Laboratoire de Neurochimie, UMR INSERM 711, Hopital de la Salpetriere, 47 bd de l’hopital, 75651 Paris cedex 13, France
ˆ
c
´
Universite Pierre et Marie Curie, UMR CNRS 7613, 4 Place Jussieu, 75252 Paris cedex 05, France
Received 10 August 2005; revised 6 October 2005; accepted 10 October 2005
Available online 7 November 2005
Abstract—The sulfated glucuronosyl paragloboside SO4-3-GlcA(b1-3)Gal-(b1-4)GlcNAc(b1-3)Gal-(b1-4)Glcb(1-10)Cer (SGPG) is a
specific constituent of the peripheral nervous system involved in human dysimmune neuropathies. Although it is highly immunogenic in
human pathology, it is not a major constituent of peripheral nerve in humans. Pure SGPG is very difficult to obtain from natural sources, but is
needed for the design of efficient and reproducible immunoassays. We describe a new chemical synthesis of this important glycolipid with a
stearic tail, including its complete characterization by mass spectrometry and its use in the detection of autoimmune neuropathies.
q 2005 Elsevier Ltd. All rights reserved.
1. Introduction
trisaccharide sequence of the SGPG (glucuronosyl sulfate-
galactosyl N-acetylglucosaminyl) is the HNK-1 epitope,
common to the SGPG lipid, and its derivatives with two
lactosaminyl residues (Fig. 1), and to several adhesion
molecules and myelin proteins, among which is P0, the
major protein of PNS, and MAG (myelin-associated
glycoprotein).1 Clinico-biological studies in neuropathies
have been performed, mainly to search for anti MAG/
SGPG antibodies (reviewed in Ref. 2). Interestingly, the
predominantly sensory demyelinating neuropathies with
monoclonal immunoglobulins of the M type (IgMs) have
antiMAG/SGPG activities, whereas axonal neuropathies,
Sphingoglycolipids are constituents of cellular structures of
the peripheral nervous system (PNS) and the central
nervous system (CNS). Several autoimmune neuropathies
can be characterized, not only on clinical and electro-
physiological grounds, but also on immunochemical
grounds, related to the nature of the antigen recognized
by antiglycolipid antibodies. The sulfated glucuronosyl
paragloboside SO4-3-GlcA(b1-3)Gal-(b1-4)GlcNAc-
(b1-3)Gal-(b1-4)Glcb(1-10)Cer (SGPG) is a specific
constituent of the peripheral nervous system. The terminal
Figure 1.
predominantly motor, or some cases of motor neuron
diseases3 with monoclonal IgMs present only anti SGPG
activity.4 Thus, there is a broad spectrum of dysimmune
neuropathies characterized by anti SGPG antibodies and
this has therapeutic consequences.
Keywords: SGPG; Sphingoglycolipid; Chemical synthesis; Peripheral
nervous system; Neuropathy; Electrospray mass spectrometry.
*
Corresponding author. Tel.: C33 1 44323337; fax: C33 1 44323397;
e-mail: Jean-Maurice.Mallet@eas.fr;
URL: http://www.chimie.ens.fr/glycoscience/
0040–4020/$ - see front matter q 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2005.10.021

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R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
Scheme 1. Retrosynthetic analysis.
SGPG is not commercially available. Furthermore, although
it is highly immunogenic in human pathology, it is not a
major constituent of peripheral nerve in humans and is only
a minor constituent of PNS of several animal species.5 Thus,
its purification remains a hard task. For this reason, it
appears important for clinical purposes to synthesize this
glycolipid.6 In this paper, we describe a new chemical
synthesis of this important glycolipid with a stearic tail,
including its complete spectroscopic characterization and its
use in immunodetections.
the two hydroxyl groups. In contrast to previous results
obtained using a Lewis X trisaccharide donor,7a,b a mixture
of two regioisomers (the expected 3-regioisomer (55%) and
its 4-regioisomer (30%)) has been obtained. In order to
avoid this side reaction, we decided then to protect the
position 4 of 3 by acetylation using the Lemieux method10
yielding 5 (95%) (Scheme 2). The acetate position was
1
checked in H NMR (H-40: 5.4 ppm (J4,3Z3.1 Hz) and
H-30: 3.6 ppm). The glycosylation between 2 and 5 gave,
uneventfully, rise to the formation of 6 (80%) (Scheme 2).
The b-stereochemistry of the newly formed glycosidic bond
1
was checked by H NMR (J1,2Z8.5 Hz).
2. Chemical synthesis
The 1C2C2 synthesis proposed (Scheme 1) is based upon
our previous synthetic work7 on lacto glycolipids. All the
glycosidic linkages are 1,2 trans, this stereochemistry was
easily assured by the use of a participating group at C2 of
each glycosyl donors (acetate, benzoate and phthalimido
(Phth)). The main difficulty was the expected ‘placidness’ of
the two deactivated glycosyl donors: the acylated glucu-
ronic unit (16, 17) and the acetylated pentasaccharide
imidate 20. Three of the four key intermediates have been
prepared previously: the lactose unit 3,8 the lactosamine unit
27a and the azidosphingosine 49 and have been prepared
according to a literature procedure.
We tried first the glycosylation between 2 and 3 using NIS,
TfOH as a promoter in dichloromethane. The hydroxyl
group at position 3 was expected to be the more reactive of
Scheme 2. Reagents: (a) MeC(OEt)3, CSA then AcOH 80%, 81%; (b) NIS,
TfOH, CH2Cl2, K30 8C, 80%.

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
565
This tetrasaccharide 6 was transformed in four steps into a
glycosyl acceptor 8, ready to react with a glucuronosyl
donor (Scheme 3). The hydroxyl group at position 3 of Gal
IV is usually the more reactive of the four hydroxyl group
present in 8.
in two steps from diacetone glucose, then silylated by
terbutyldimethylsilyl chloride in pyridine13 followed by
one-pot acylation with Ac2O to give 10 (or with BzCl to
give 11) (Scheme 4).
Jones oxidation, followed by methylation, afforded the two
methyl 3-O-benzyl uronates in good yields. Exchange of
benzyl to levulinate was at this stage uneventful. The
acylated products 14 and 15 have been then transformed
into glucuronosyl bromides 16 and 17 in good yield using
HBr in acetic acid (Scheme 5).
The [8C16] condensation in dichloromethane promoted by
silver triflate or with Ag2CO3/I2 at room temperature
14
failed to give any pentasaccharide. However, the benzoyl-
ated bromide 17 (2 equiv) finally gave the pentasaccharide
18 in an acceptable yield of 50% (72% based on recovered
8) (Scheme 6). We tried to improve this step using
analogous benzoylated thiophenyl glycosides and trichlor-
oacetimidate, without any success.
The pentasaccharide 18 was then transformed into a
glycosyl donor in four steps (Scheme 7): first removal of
the benzylidene and benzyl groups (H2, Pd/C) followed by
acetylation to give 19. The regioselectivity of the last
glycosylation was easily checked by NMR (H-3 Gal IV dZ
3.8 ppm). The trimethylsilylethyl protecting group was then
removed and the a imidate 20 was prepared using
trichloroacetonitrile and DBU as a base.
Scheme 3. Reagents: (a) MeONa/MeOH, CH2Cl2, 88%; (b) PhCH(OMe)2,
CSA, CH3CN, 82%, (c) N2H4, EtOH, 80 8C, (d) Ac2O, MeOH, 81%
(two steps).
Two glucuronosyl donors have been synthesized (Schemes 4
and 5). They have a levulinoyl group at C3. This cetoester,
selectively removed in presence of acetate or benzoate, is a
classical and reliable choice in sulfated glycosaminoglycan
chemical synthesis.11 We introduced first a levulinate group
at C3 of diacetone D-glucose and then tried without success
to remove the two isopropylidene groups using hot aqueous
AcOH. We moved then to a more stable temporary group,
the benzyl ether. Thus, 3-O-benzyl glucose12 was prepared
Coupling with 4 gave the glycoconjugate 21 with a good
yield (75%) (Scheme 8). Compound 21 was then easily
converted into 1 in a few steps (Scheme 9): reduction of
azide with PPh315 in aqueous THF at 50 8C, introduction of
Scheme 4. Reagents: (a) BnBr, NaH, DMF; (b) H2O, IR 120 (HC), 85% (two steps); (c) TBS-Cl, Pyr., (d) Ac2O, Pyr., 88% (two steps); (e) BzCl, Pyr. 90%
(two steps).
Scheme 5. Reagents (a) CrO3, H2SO4, acetone; (b) MeI, KHCO3, DMF 65% (two steps); (c) H2, Pd/C, MeOH; (d) Lev2O, Pyr., 82% (two steps); (e) HBr,
AcOH, CH2Cl2, 90%; (f) CrO3, H2SO4, acetone; (g) MeI, KHCO3, DMF 68% (two steps); (h) H2, Pd/C, MeOH; (i) Lev2O, Pyr., 80% (two steps); (j) HBr,
AcOH, CH2Cl2, 70%.

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R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
Scheme 6.
Scheme 7. Reagents: (a) H2, Pd/C, MeOH; (b) Ac2O, Pyr.; (c) CF3COOH, CH2Cl2, 0–20 8C; (d) Cl3CCN, DBU, CH2Cl2.
Scheme 8.
stearic acid, with DCC as a coupling agent; removal of
levulinate with hydrazine monoacetate, and sulfatation of
the free hydroxyl group with SO3$NMe3 in DMF at 50 8C.
Final deprotection was done in two steps, that is,
saponification of the methyl ester with lithium hydroxide,
followed by an treatment with MeONa/MeOH/THF. This
3. Mass characterisation of synthetic SGPG
The negative ion ESI mass spectrum of the synthetic SGPG
(Fig. 3a), recorded using the negative ion mode, displayed
various singly charged ions at m/z 1510.1 and m/z 1532.1
and doubly-charged ions at m/z 754.5, that correspond to
the, [MKH]K, [MK2HCNa]K and [MK2H]2K quasi-
molecular species, respectively. From this mass spectrum, it
appears that the molecular weight of the synthetic
compound was 1511.1 u consistent with the expected
1
two steps sequence limits the b elimination. The H NMR
spectrum in DMSO-d6 (see Fig. 2 for ethylenic and
anomeric proton signals) was found in accordance with
reported spectrum for natural SGPG.6b

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
567
Scheme 9. Reagent: (a) PPh3, THF, H2O, 50 8C; (b) DCC, stearic acid, CH2Cl2, 78% (two steps); (c) N2H5$AcO, EtOH, 81%; (d) (Me)3N–SO3, DMF, 50 8C,
80%; (e) LiOH, THF, H2O, 0 8C; (f) MeONa, MeOH, THF, 90% (two steps).
Figure 2. 1H NMR selected data (DMSO-d6, 400 MHz): ethylenic and anomeric protons.
monoisotopic peak as well as with to the natural isotopic
cluster distribution, which is surperimposable on the
calculated cluster distribution (Fig. 3a). Furthermore,
prompt fragmentations are limited since only ions at m/z
1492.1 (i.e., loss of water) and m/z 357.0 raised up to 10% of
base peak (i.e., at m/z 1532.1), under these ESI source
conditions. These quasi-molecular species being sufficiently
abundant they can be studied by MS/MS under low energy
collision excitation (CID processes).
hydroxyl of the neighbouring glucosidic unit (study in
progress) yielding a terminal sulfated glucose in the m/z
1334.1 production. This behaviour makes ambiguity on the
sulfate group location.
In order to obtain more information on the sequence (except
for the sulfate position), in situ consecutive ion excitations
(called ‘tickle excitation’) were applied: (i) to the first
generation product m/z 1430.1 ion and (ii) in second step, to
the second generation product m/z 1412.2 ion (formed by
water release from the previous precursor of the first
generation). The recorded CID spectrum (Fig. 3b) displays a
series of the losses of 158 u (glucuronic unit having lost an
OH group), 162 u (glucosidic unit), 203 u (GlcNAc residue)
produced either by consecutive fragmentations or directly
by competitive cleavages in the ion trap cell. These different
neutral losses give rise to the formation of the following ions
at m/z 1412.3, m/z 1254.2, m/z 1092.1, m/z 889.9, m/z 726.7,
and m/z 564.6 corresponding to the Yi series (without
significant presence of the complementary Bj series)
Firstly, the collision excitation of the deprotonated molecule
(m/z 1510.1) shows this precursor ion was very stable since
only two product ions are displayed at m/z 1430.1 (100%)
and m/z 1334.1 (5%) in the CID spectrum (not reported
herein). These ions are promoted from the deprotonated
molecule carrying out the negative charge at the sulfate site
(the larger acidic group in gas phase) by releasing the SO3
neutral (i.e., 80 u) and the desulfated glucuronic terminal
unit, respectively. The latter loss is generated by sulfate
migration form the C3 position of terminal residue likely to

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R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
Figure 3. Negative ion ESI mass spectrum of synthetic-SGPG (a) and MS2 experiments under CID conditions of singly-deprotonated (b) and doubly-
deprotonated [MK2H]2K synthetic SGPG (m/z 754.5) (c) in comparison with CID spectrum of the doubly deprotonated natural SGPG recorded in same
experimental conditions (d).
according to Domon and Costello’s nomenclature.16 They
are produced after the SO3 release by charge promoted
competitive cleavages of sugar linkages and the charge is
retained at the ceramide moiety. This orientation can be
expected assuming that the charge location occurs at the
ceramide amide group because of its particular gas phase
acidity relative to other acidic sites. This observed series of
product ions is consistent with the proposed sequence
without assuring the sulfate position on glucuronic acid
moiety.
Evidence of its location as well as the total sequence in one
scanning has been achieved from the collision excitation of
the doubly-deprotonated [MK2H]2K molecule (m/z 754.5).

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
569
Figure 4. Main complementary product ions of the deprotonated synthetic-SGPG (C18:0; d18:1) according to Domon and Costello’s nomenclature. These
cleavages are observed from the doubly charged SGPG.
The CID spectrum of this precursor ion (Fig. 3c) displays
different series of singly-charged product ions. These series
are complementary and are noted as Yi and Bi (Fig. 4).
Interestingly, two abundant complementary product ions
(more than 50% of base peak at m/z 638.2) appear at m/z
1254.1 to m/z 254.6 (Fig. 3c) corresponding to the Y4 and B1
ions (Fig. 4) give immediately the evidence on the sulfate
location on the terminal glucuronidic residue. Other Bj/Yi
couples of complementary product ions are too observed at
m/z 417.0 /m/z 1092.0, m/z 620.2/m/z 888.9, m/z 782.3/m/z
726.7, which confirm the SGPG whole sequence as well as
the presence of sulfate on glucuronidic unit. In addition to
the previous series is accompanied by other ions such as the
C4, C3, and C2 product ions corresponding to m/z 800.3, m/z
638.2, m/z 435.3, respectively, (Fig. 4). However, the
complementary product Zi ions are not observed, which
indicates that they decompose into low m/z fragment ions
beyond the product ion m/z ratio range (the parent ion m/z
values must be four times lower than that of its product
ions). This value is related to the low mass cut-off fixed
automatically by the software of the mass spectrometer.
Furthermore, from all the product Bj and Cj ions, a 80 u
neutral loss is released systematically (Table 1).
glycolipid), and the enzymatic reaction was quenched 3 M
HCl (instead of 3 M H2SO4).
We compared the sera of 49 patients with predominantly
sensory peripheral neuropathies associated with the
presence of a monoclonal IgM. We were able, with this
synthetic SGPG (1), to clearly classify them into two
groups: 24 had anti SGPG antibodies, 25 were completely
negative (that is under the detection level). Anti SGPG
antibodies titers of the 24 positives patient are shown in
Figure 5 and confirm the relation between SGPG and the
neuropathy, which is important for therapeutic purpose.18
The titers were extremely variable and no correlation
can be clearly established with the total amount of
monoclonal IgM.
Table 1. Observed consecutive loss of SO3 (80 u) from each product Bj and
Cj ions generated from the precursor doubly-deprotonated [MK2H]2K
molecule
Series of ions
iZ4
iZ3
iZ2
Bj
Cj
782.3/702.3
800.3/720.3
620.2/540.2
638.2/558.2
416.8/336.8
435.0/354.9
Finally, to obtain a definitive confidence in our interpreta-
tion, the same MS/MS experiment was performed from
doubly-deprotonated natural SGPG (Fig. 3). The recorded
fingerprints represented by their respective CID spectra
were superimposable. Furthermore, even the peaks charac-
terized by very weak abundances are present with similar
relative abundances, which confirms the proposed SGPG
structure.
Figure 5. Anti SGPG antibodies titers, in predominantly sensory
demyelinating neuropathy with monoclonal IgM. Negative sera are not
shown.
4. Immunology
Elisa measurements have been done on 96-well micro plates
according to the established procedure for antiglycolipid
antibody Elisa17 with two minor modifications: 100 ng of
SGPG were coated per well (instead of 200 ng of
This technique is much more sensitive (up to 1000 fold) than
the previously used one using crude nerve extracts purified
and immunodetected on a thin-layer chromatography

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R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
plate.19 For instance, patients under 1000 in Figure 5 were
not diagnosed with this technique. This improved technique
´ `
is now in use in routine analysis in Salpetriere hospital for
ejection to the non-linear field at bzZ2/3. The injection low
mass cut off (LMCO) was 70 Th. Negative ion mode
detection was used. The automated ion charge control (ICC)
was set to 10,000 in order to avoid space charge effect.
Sequential MSn experiments were performed under
resonant excitation conditions from precursor ion selected
by application of broadband frequency excitation with a
notch allowed a m/z ratio selection window of 2 Th. In the
case of MSn application (with nO2), when chosen precursor
ion was characterized by a too low abundance, it was
directly excited by application of resonant frequency
without to use the ion selection step. The LMCO used
during the collision induced dissociation (CID) experiments
(related to the excitation at particular bz value) was set to
27% of the mass to charge ratio of the precursor ion.
diagnosis of neuropathies.
5. Conclusion
The synthetic product, although containing a single fatty
acid, has the same specificity for IgM SGPG antibodies as
the natural product (data not shown). The ELISA test that
we have set up for clinical purposes is considerably more
sensitive than immunodetection on thin-layer chromato-
graphy, and thus will allow the detection of dysimmune
neuropathies, which were undiagnosed in the past. This may
have therapeutic consequences, as treatments are becoming
more and more specific and designed to lower antibody
concentration or to interfere with immunopathogenic
mechanisms. It may be that the use of solid-phase
immunoadsorbents containing a reactive oligosaccharide,
will permit a more selective therapy for antibody-mediated
neuropathies. Synthesis of ganglioside epitopes for oligo-
saccharide specific immunoadsorption therapy has been
6.2.1. 2-(Trimethylsilyl)ethyl (2,6-di-O-benzyl-4-O-
acetyl-b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-benzyl-
b-D-glucopyranoside (5). To a stirred solution of 3 (9 g,
10 mmol) in triethyl orthoacetate (30 mL) was added, at
room temperature, CSA (200 mg). After 30 min, aqueous
acetic acid 80% (26 mL) was added dropwise. The mixture
was stirred (15 min) and evaporated in vacuum. The residue
was purified by flash chromatography (eluent, 20% EtOAc
in cyclohexane) to afford 5 (9.3 g, 95%) as a colourless oil.
[a]D K10 (c 1 in chloroform); 1H NMR (400 MHz, CDCl3):
d 7.43–7.30 (m, 25H, arom.), 5.40 (dd, 1H, J4–3Z3.4 Hz,
J4–5Z0.6 Hz, H-4B), 5.03 (d, 1H, JgemZ10.5 Hz, CHPh),
´
suggested for the Guillain–Barre syndrome, with the aim of
removing auto-antibodies either by using soluble blocking
ligands administered systematically or as immunoaffinity
ligands for use as extracorporeal immunoadsorbents.20,21
This type of approach could be used for other antigens such
as SGPG.
4.97 (d, 1H, JgemZ11.0 Hz, CHPh), 4.87 (d, 1H, Jgem
Z
11.4 Hz, CHPh), 4.82 (d, 1H, JgemZ10.5 Hz, CHPh), 4.79
(d, 1H, JgemZ11.0 Hz, CHPh), 4.73 (d, 1H, JgemZ11.4 Hz,
CHPh), 4.65 (d, 1H, JgemZ12.0 Hz, CHPh), 4.53 (d, 1H,
J1–2Z7.7 Hz, H-1B), 4.51 (d, 1H, JgemZ12.0 Hz, CHPh),
6. Experimental
6.1. General procedures
4.49 (d, 1H, JgemZ12.0 Hz, CHPh), 4.45 (d, 1H, J1–2Z
7.7 Hz, H-1A), 4.29 (d, 1H, JgemZ12.0 Hz, CHPh),
4.11–4.08 (m, 1H, O–CH–CH2Si(CH3)3), 4.04 (dd, 1H,
J4–3Z9.0 Hz, J4–5Z9.5 Hz, H-4A), 3.97–3.80 (m, 2H,
H-6A), 3.72–3.67 (m, 1H, H-3B), 3.69–3.65 (m, 1H,
O–CH–CH2Si(CH3)3), 3.63 (t, 1H, J3–2Z9.0 Hz, H-3A),
3.57 (dt, 1H, J5–6Z6.6 Hz, H-5B), 3.48–3.43 (m, 3H, H-2B,
All compounds were homogeneous by TLC analysis and
had spectral properties consistent with their assigned
structures. Melting points were determined in capillary
tubes in a Bu¨chi 510 apparatus, and are uncorrected. Optical
rotations were measured with a Perkin-Elmer Model 241
digital polarimeter at 22G3 8C. Compound purity was
checked by TLC on Silica gel 60 F254 (E. Merck) with
detection by charring with sulfuric acid. Column chromato-
graphy was performed on Silica gel 60 (E. Merck). 1H NMR
spectra were recorded with Bru¨ker AM 250, AM 400
instruments. Elemental analyses were performed by Service
H-2A and H-5A), 3.38 (d, 2H, H-6B), 2.40 (d, 1H, JOH-3
Z
2.4 Hz, OH), 2.09 (s, 3H, O–C]O–CH3), 1.10 (m, 2H,
O–CH2–CH2Si(CH3)3), 0.09 (s, 9H, Si(CH3)3); 13C NMR
(100 MHz): d 170.9 (O–C]O–CH3), 139.0, 138.7, 138.2,
138.1 and 137.9 (5C arom.), 128.4–127.3 (25CH arom.),
103.1 (C-1A), 102.3 (C-1B), 82.7 (C-3A), 81.8 (C-2B), 80.0
(C-2A), 76.5 (C-4A), 75.2 (CH2Ph), 75.0 (C-5A), 74.9
(CH2Ph), 74.9 (CH2Ph), 73.3 (CH2Ph), 73.1 (CH2Ph), 72.4
(C-3B), 71.9 (C-5B), 69.5 (C-4B), 68.2 (C-6A), 67.4
(O–CH2–CH2Si–), 67.1 (C-6B), 20.8 (–O–C]O–CH3),
18.4 (O–CH2–CH2Si–), K1.4 (Si(CH3)3); MS m/z (CI,
NH3): 952.7 (MCNH4)C; Anal. Calcd for C54H66O12Si
(935.204): C 69.35, H 7.11; found C 69.30, H 7.17.
´
d’Analyse de Universite Pierre et Marie Curie. Chemical
ionization and FAB mass spectrometry were recorded with
Jeol MS700: CI (gas: ammonia); FAB (matrix: NBA, NaI).
MALDI-TOF mass spectra were recorded with PerSpective
Biosystems Voyager Elite (Framingham, MA, USA)
equipped with a nitrogen laser (337 nm) using a 2,5-
dihydroxybenzoic acid (2,5-DHB) matrix.
6.2. Mass spectrometry procedure
6.2.2. 2-(Trimethylsilyl)ethyl(2,3,4,6-tetra-O-benzoyl-
b-D-galactopyranosyl)-(1/4)-(6-O-benzyl-2-deoxy-2-
phthalimido-b-D-glucopyranosyl)-(1/3)-(4-O-acetyl-
2,6-di-O-benzyl-b-D-galactopyranosyl)-(1/4)-2,3,6-tri-
O-benzyl-b-D-glucopyranoside (6). To a stirred mixture of
SGPG was analyzed by using an electrospray ionization
(ESI)-ion-trap mass spectrometer22 (Esquire 3000, Bruker)
(Bremen, Germany). The source conditions were as follow:
capillary high voltage 3500 V, capillary exit K50 V,
skimmer 1 K20 V. The ion analysis occurred within a
scan rate of 13,000 Th/s using a mass/charge ratio range of
3000 Th by using analytical scan mode through resonant ion
˚
2 (6.64 g, 6.2 mmol), 5 (5.8 g, 6.2 mmol) and MS-4 A
(15 g) in anhydrous CH2Cl2 (150 mL) were successively
added, at K30 8C and under argon, NIS (2.8 g, 12.4 mmol)
and TfOH (0.165 mL, 1.86 mmol). The mixture was stirred

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
571
for 30 min at K30 8C, neutralized by saturated sodium
hydrogencarbonate and filtered through Celite. The filtrate
was washed with saturated sodium thiosulfate, brine, dried
(MgSO4) and evaporated in vacuum. The residue was
purified by flash chromatography (eluent gradient, 20%
EtOAc in cyclohexane) and precipitated (EtOAc/hexanes)
to afford 6 (9.4 g, 80%) as a white powder. [a]D C48 (c 1 in
(100 mL). The mixture was stirred at room temperature
(15 min), neutralized by solid K2CO3, filtered through
Celite and evaporated in vacuum. The residue was purified
by flash chromatography (eluent, 65% EtOAc in cyclo-
hexane) to afford 7 (4.8 g, 91%) as a white powder. [a]D
K18 (c 1 in chloroform); mp: 95–96 8C (EtOAc/cyclohex-
1
ane); H NMR (400 MHz, CDCl3): d 7.43–7.20 (m, 39H,
arom.), 5.51 (d, 1H, J1–2Z8.4 Hz, H-1C), 5.47 (s, 1H,
1
chloroform); mp: 153–154 8C (EtOAc/hexanes); H NMR
`
(400 MHz, CD3C]OCD3): d 8.24–7.15 (m, 54H, arom.),
6.21 (dd, 1H, J4–3Z2.8 Hz, J4–5Z0.7 Hz, H-4D), 6.03 (m,
2H, H-2D and H-3D), 5.65 (de, 1H, J4–3Z3, 6 Hz, H-4B),
benzylidene), 1.08–1.04 (m, 2H, O–CH2–CH2Si), 0.06 (s,
9H, Si(CH3)3); 13C NMR (100 MHz): d 168.2, 167.9
(2C]O Phth), 139.0, 138.7, 138.4, 138.3, 138.2, 137.7,
137.2 and 131.1 (8C arom.), 133.8–126.2 (CH arom.),
5.59 (d, 1H, J1–2Z8.5 Hz, H-1C), 5.57 (d, 1H, J1–2
Z
`
7.0 Hz, H-1D), 5.10 (d, 1H, JgemZ10.7 Hz, CHPh), 5.00 (d,
1H, JgemZ11.5 Hz, CHPh), 5.00–4.90 (m, 1H, H-5D), 4.90
(dd, 1H, J6b–6aZ11.5 Hz, J6b–5Z4.0 Hz, H-6Db), 4.87 (d,
1H, JOH–3CZ2.1 Hz, OH), 4.85 (d, 1H, JgemZ11.5 Hz,
CHPh), 4.76 (d, 1H, JgemZ10.7 Hz, CHPh), 4.79–4.73 (m,
1H, H-3C), 4.63 (d, 1H, JgemZ12.2 Hz, CHPh), 4.59 (d, 1H,
JgemZ12.2 Hz, CHPh), 4.58 (d, 1H, J1–2Z7.8 Hz, H-1B),
4.57 (d, 1H, JgemZ12.4 Hz, CHPh), 4.58–4.53 (m, 1H,
H-6Da), 4.53 (d, 1H, JgemZ12.2 Hz, CHPh), 4.50 (d, 1H,
J1–2Z7.7 Hz, H-1A), 4.40 (d, 2H, JgemZ12.2 Hz, 2CHPh),
103.6, 102.9, 101.9, 101.1 and 98.7 (benzylidene, C-1A,
C-1B, C1-C and C1-D); MS m/z (CI, NH3): 1541 (MC
NH4)C. Anal. Calcd for C86H97NO22Si (1524.80): C 67.74,
H 6.41, N 0.92; found C 67.62, H 6.52, N 0.93.
6.2.4. 2-(Trimethylsilyl)ethyl (4,6-O-benzylidene-b-D-
galactopyranosyl)-(1/4)-(2-acetamido-6-O-benzyl-2-
deoxy-b-D-glucopyranosyl)-(1/3)-(2,6-di-O-benzyl-
b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-benzyl-b-D-
glucopyranoside (8). NH2NH2$H2O (45 mL) and H2O
(25 mL) were added to a stirred solution of 7 (4.8 g,
3.15 mmol) in ethanol (200 mL). The mixture was stirred
overnight under reflux and then evaporated in vacuum. The
residue was diluted in a mixture of DCM–MeOH (v/vZ1/1,
20 mL). The precipitate was filtered and acetic anhydride
(5 mL, excess) was added to the filtrate at room temperature.
After 2 h, solvents were evaporated in vacuum. The residue
was purified by flash chromatography (eluent, 5% MeOH in
DCM) and precipitated in a mixture of EtOAc/hexanes to
afford 8 (3.74 g, 83%) as a white powder. [a]D K11 (c 1 in
chloroform); mp: 98–102 8C (EtOAc/hexanes); H NMR
(400 MHz, C6D6/CD3OD: 1:1): d 7.80–7.40 (m, 35H,
arom.), 5.59 (s, 1H, benzylidene), 5.43 (d, 1H, Jgem
10.7 Hz, CHPh), 5.17 (d, 1H, JgemZ11.3 Hz, CHPh), 5.03
(s, 2H, CH2Ph), 5.00 (d, 1H, JgemZ10.7 Hz, CHPh), 4.98 (d,
1H, JgemZ11.3 Hz, CHPh), 4.92 (d, 1H, J1–2Z8.4 Hz,
H-1C), 4.81 (d, 1H, J1–2Z8.8 Hz, H-1B), 4.80 (d, 1H,
JgemZ11.8 Hz, CHPh), 4.77 (d, 1H, JgemZ11.8 Hz, CHPh),
4.38 (d, 1H, JgemZ12.2 Hz, CHPh), 4.35 (d, 1H, Jgem
Z
12.4 Hz, CHPh), 4.28 (dd, 1H, J2–3Z10.8 Hz, H-2C),
4.12–4.06 (m, 2H, O–CH–CH2Si– and H-4C), 4.04 (dd,
1H, J4–3Z9.2 Hz, J4–5Z9.8 Hz, H-4A), 3.95 (dd, 1H,
J3–2Z9.6 Hz, H-3B), 3.83–3.79 (m, 1H, H-5C), 3.77–3.70
(m, 5H, H-5B, H-6C, H-6Ab and O–CH–CH2Si), 3.60–3.56
(m, 3H, H-3A, H-6Bb and H-6Aa), 3.50 (dd, 1H, H-2B),
3.46 (dd, 1H, J6a–6bZ10.3 Hz, J6a–5Z6.6 Hz, H-6Ba), 3.39
(ddd, 1H, J5–6aZ3.5 Hz, J5–6bZ1.5 Hz, H-5A), 3.30 (dd,
1H, J2–3Z9.2 Hz, H-2A), 2.14 (s, 3H, O–C]O–CH3),
1.15–1.12 (m, 2H, O–CH2–CH2Si), 0.15 (s, 9H, Si(CH3)3);
13C NMR (100 MHz): d 169.4, 168.9, 168.4, 164.7, 164.5,
164.0 and 163.9 (5O–C]O–CH3/Ph and 2C]O Phth),
138.7, 138.5, 138.2, 138.0, 137.7, 137.6, 2!130.5, 128.5,
128.4 and 128.0 (11C arom.), 128.8–126.0 (54CH arom.),
101.9 (C-1A), 100.8 (C-1B), 100.7 (C-1D), 97.9 (C-1C), 81.6
(C-3A), 81.4 (C-4C), 81.1 (C-2A), 78.2 (C-2B), 77.8 (C-3B),
74.7 (C-4A), 73.9 (CH2Ph), 73.6 (C-5A), 73.4 (CH2Ph), 73.3
(C-5C), 73.3 (CH2Ph), 72.1 (CH2Ph), 72.0 (C-5B), 71.8
(CH2Ph), 71.7 (CH2Ph), 71.1 (C-5D), 70.9 (C-3D), 69.6
(C-4B), 69.0 (C-2D), 68.3 (C-3C), 67.9 (C-4D), 67.8, 67.4,
66.9 and 61.9 (4C-6), 65.5 (O–CH2–CH2Si–), 55.7 (C-2C),
19.2 (–O–C]O–CH3), 17.1 (O–CH2–CH2Si–), K3.0
(Si(CH3)3); MS m/z (CI, NH3): 1912.9 (MCNH4)C. Anal.
Calcd for C109H111NO27Si (1895.17): C 69.08, H 5.90, N
0.74; found C 68.98, H 5.92, N 0.71.
1
Z
4.67 (2d, 2H, JgemZ12.0 Hz, 2CHPh), 4.60 (d, 1H, J1–2
Z
7.8 Hz, H-1D), 4.56 (d, 1H, J1–2Z7.8 Hz, H-1A), 4.50 (d,
1H, JgemZ12.0 Hz, CHPh), 4.49 (d, 1H, JgemZ12.0 Hz,
CHPh), 4.47 (de, 1H, J4–3Z3.3 Hz, H-4B), 4.32 (t, 1H, J4–
3ZJ4–5Z9.8 Hz, H-4A), 4.30 (dd, 1H, J2–3Z10.0 Hz, H-
2C), 4.26–4.14 (m, 4H, H-6Db, –O–CH–CH2–Si and H-6C),
4.13 (de, 1H, J4–3Z3.7 Hz, H-4D), 4.07–4.02 (m, 4H, H-
2D, H-2B, H-6Bb and H-6Ab), 4.02 (t, 1H, J3–4Z10.0 Hz,
H-3C), 3.96 (t, 1H, J4–5Z10.0 Hz, H-4C), 3.94–3.89 (m,
2H, H-6Da and H-6Ba), 3.86 (dd, 1H, J3–2Z9.3 Hz, H-3B),
3.86–3.78 (m, 5H, H-5C, H-3A, H-5B, H-6Aa and –O–CH–
CH2–Si), 3.77 (dd, 1H, J3–2Z9.9 Hz, H-3D), 3.65 (dd, 1H,
J2–3Z9.1 Hz, H-2A), 3.50–3.48 (m, 1H, H-5A), 3.30 (se,
1H, H-5D), 1.96 (S, 3H, –NH–C]O–CH3), 1.23–1.16 (m,
2H, O–CH2–CH2–Si), 0.18 (s, 9H, Si(CH3)3); 13C NMR
(100 MHz): d 173.7 (–NH–C]O–CH3), 140.6, 140.4,
140.2, 139.9, 139.6, 139.6 and 139.3 (7C arom.),
129.9–127.4 (35CH arom.), 105.0 (C-1D), 104.3 (C-1A),
6.2.3. 2-(Trimethylsilyl)ethyl (4,6-O-benzylidene-b-D-
galactopyranosyl)-(1/4)-(6-O-benzyl-2-deoxy-2-phtha-
limido-b-D-glucopyranosyl)-(1/3)-(2,6-di-O-benzyl-
b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-benzyl-b-D-
glucopyranoside (7). Sodium (40 mg, 0.02 M) was added
by portions to a stirred solution of 6 (7.11 g, 3.75 mmol) in
MeOH/CH2Cl2 (80 mL/40 mL). The mixture was stirred for
2 h, neutralized by Amberlite-IR 120 (HC), filtered and
evaporated in vacuum. The residue was purified by flash
chromatography (eluent, 10% MeOH in DCM) to afford a
white powder (4.98 g, 92%), which was directly engaged in
the next step. Benzaldehyde dimethylacetal (1.04 mL,
6.93 mmol) and pTsOH (190 mg, 1 mmol) were added to
a stirred solution of the previous powder in CH3CN
`
104.0 (C-1C), 103.7 (C-1B), 102.0 (benzylidene), 84.0
(C-3A), 83.9 (C-3B), 83.2 (C-2A), 81.4 (C-4C), 80.2
(C-2B), 77.6 (C-4A), 76.9 (C-4D), 76.4 (CH2Ph), 76.2
(C-5A), 76.0 (CH2Ph), 75.9 (CH2Ph), 75.6 (C-5C), 75.0
(C-5B), 74.6 (CH2Ph), 74.5 (CH2Ph), 74.1 (CH2Ph), 73.6

572
R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
(C-3D), 73.5 (C-3C), 71.9 (C-2D), 70.5 (C-6B), 70.1
(C-4B), 69.9 (C-6C), 69.9 (C-6D), 69.3 (C-6A), 68.2
(O–CH2–CH2–Si–), 68.0 (C-5D), 57.0 (C-2C), 23.3
(–NH–C]O–CH3), 19.4 (O–CH2–CH2–Si–), K0.8
(Si(CH3)3); MS m/z (CI): 1436.8 MHC. Anal. Calcd for
C80H97NO21Si (1436.7): C 66.87, H 6.80, N 0.97; found C
66.57, H 6.81, N 0.98.
230 mmol) at 0 8C. The mixture was stirred at room
temperature for 3 h, cooled to 0 8C and anhydrous methanol
(10 mL) was added dropwise. The solvent was removed in
vacuum and the resulting residue was partitioned between
Et2O and water. The organic layer was dried (MgSO4),
filtered and evaporated in vacuum. Amberlite-IR 120 (HC)
(60 g) was added to a suspension of the previous residue in
H2O (300 mL). After one night under reflux, the mixture
was filtered and evaporated in vacuum. The crude
compound was purified by precipitation (acetone/cyclohex-
ane) to afford 9 (24.2 g, 85%) as a white powder. TBDMSCl
(6.42 g, 42.58 mmol) was added to a stirred solution of 9
(10.95 g, 40.55 mmol) in pyridine (100 mL) at 0 8C under
argon. The mixture was immediately warmed to room
temperature and stirred for 2 h. Then benzoyl chloride
(27.86 mL, 240 mmol) was added dropwise to the mixture.
After one night at room temperature, solvent was removed
in vacuum. The residue was diluted with DCM, washed
successively with HCl (1 M), saturated sodium hydro-
gencarbonate, brine, dried (MgSO4) and evaporated in
vacuum. The residue was purified by flash chromatography
(eluent, 20% EtOAc in cyclohexane) to afford a mixture of
11a/b (a/bZ1/3, 25.4 g, 90%) as a white powder. A small
amount of this mixture was selectively recrystallized from
hexanes and gave pure 11b as a white powder.
6.2.5. 1,2,4-Tri-O-acetyl-3-O-benzyl-6-O-tert-butyl-
dimethylsilyl-(a/b)-D-glucopyranose (10a/b). Sodium
hydride (9.2 g, 230 mmol, 60% in mineral oil) was added
to a stirred solution of diacetone glucose (30 g, 115 mmol)
in anhydrous DMF (300 mL) and benzyl bromide (27.4 mL,
230 mmol) at 0 8C. The mixture was stirred at room
temperature for 3 h, cooled to 0 8C and anhydrous methanol
(10 mL) was added dropwise. The solvent was removed in
vacuum and the resulting residue was partitioned between
Et2O and water. The organic layer was dried (MgSO4),
filtered and evaporated in vacuum. Amberlite-IR 120 (HC)
(60 g) was added to a suspension of the previous residue in
H2O (300 mL). After 15 h under reflux, the mixture was
filtered and evaporated in vacuum. The crude compound
was purified by precipitation (acetone/cyclohexane) to
afford 9 (24.2 g, 85%) as a white powder. TBSCl (6.42 g,
42.58 mmol) was added to a stirred solution of 9 (10.95 g,
40.55 mmol) in pyridine (100 mL) at 0 8C under argon. The
mixture was immediately warmed to room temperature and
stirred for 2 h. Then acetic anhydride (37 mL, 365 mmol)
was added dropwise to the mixture. After one night at room
temperature, solvent was removed in vacuum. The residue
was diluted with DCM, washed successively with HCl
(1 M), saturated sodium hydrogencarbonate, brine, dried
(MgSO4) and evaporated in vacuum. The residue was
purified by flash chromatography (eluent, 15% EtOAc in
cyclohexane) to afford a mixture of 10a/b (a/bZ1/1,
18.61 g, 90%) as a white powder.
Compound 11b. [a]D K31 (c 1 in chloroform); mp:
129–130 8C (hexanes); 1H NMR (400 MHz, CDCl3): d
8.08–7.04 (m, 20H, arom.), 6.17 (d, 1H, J1–2Z7.8 Hz, H-1),
5.71 (dd, 1H, J2–3Z8.8 Hz, H-2), 5.58 (t, 1H, J4–3ZJ4–5
Z
8.8 Hz, H-4), 4.68 (d, 1H, JgemZ11.5 Hz, CHPh), 4.64 (d,
1H, JgemZ11.5 Hz, CHPh), 4.20 (t, 1H, H-3), 3.98 (ddd, 1H,
J5–6aZ3.2 Hz, J5–6bZ5.3 Hz, H-5), 3.91 (dd, 1H, J6b–6a
Z
11.4 Hz, H-6b), 3.82 (dd, 1H, H-6a), 0.85 (s, 9H,
SiC(CH3)3), K0.02 and K0.01 (2s, 6H, Si(CH3)2); 13C
NMR (100 MHz): d 164.9, 164.9 and 164.8 (3O–C]O),
137.1, 129.5, 129.3 and 128.7 (4C arom.), 133.6–127.6
(20CH arom.), 92.5 (C-1), 79.3 (C-3), 76.0 (C-5), 73.9
(CH2Ph), 72.1 (C-2), 70.4 (C-4), 62.7 (C-6), 25.7
(SiC(CH3)3), 18.2 (SiC(CH3)3), K5.5 (SiCH3), K5.4
(SiCH3); MS m/z (CI, NH3): 714.4 (MCNH4)C. Anal.
Calcd for C40H44O9Si (696.87): C 68.94, H 6.36; found C
68.98, H 6.38.
1
Compound 10b. H NMR (400 MHz, CDCl3): d 7.37–7.28
(m, 5H, arom.), 5.70 (d, 1H, J1–2Z8.2 Hz, H-1), 5.16 (dd,
1H, J2–3Z9.3 Hz, H-2), 5.13 (t, 1H, J4–3ZJ4–5Z9.5 Hz,
H-4), 4.65 (s, 2H, CH2Ph), 3.78 (t, 1H, H-3), 3.60 (ddd, 1H,
J5–6aZ2.9 Hz, J5–6bZ5.2 Hz, H-5), 3.75–3.65 (m, 2H,
H-6), 2.10, 2.01 and 2.00 (3s, 9H, 3–O–C]O–CH3), 0.90
(s, 9H, SiC(CH3)3), 0.00 (2s, 6H, Si(CH3)2).
1
Compound 11a. H NMR (400 MHz, CDCl3): d 8.08–7.04
(m, 20H, arom.), 6.80 (d, 1H, J1–2Z3.7 Hz, H-1), 5.62
1
Compound 10a. H NMR (400 MHz, CDCl3): d 7.37–7.28
(t, 1H, J4–3ZJ4–5Z9.7 Hz, H-4), 5.56 (dd, 1H, J2–3
Z
(m, 5H, arom.), 6.35 (d, 1H, J1–2Z3.7 Hz, H-1), 5.15 (t, 1H,
J4–3ZJ4–5Z10.0 Hz, H-4), 5.06 (dd, 1H, J2–3Z10.0 Hz,
H-2), 4.68 (d, 1H, JgemZ11.8 Hz, CHPh), 4.65 (d,
1H, JgemZ11.8 Hz, CHPh), 4.00 (t, 1H, H-3), 3.90 (ddd,
1H, J5–6aZ2.7 Hz, J5–6bZ4.1 Hz, H-5), 3.75–3.65 (m, 2H,
H-6), 2.20, 2.03 and 2.00 (3s, 9H, 3–O–C]O–CH3), 0.90
(s, 9H, SiC(CH3)3), K0.03 (2s, 6H, Si(CH3)2).
9.7 Hz, H-2), 4.72 (d, 1H, JgemZ11.4 Hz, CHPh), 4.64 (d,
1H, JgemZ11.5 Hz, CHPh), 4.50 (t, 1H, H-3), 4.28–4.23 (m,
1H, H-5), 3.85–3.82 (m, 2H, H-6), 0.83 (s, 9H, SiC(CH3)3),
K0.03 (2s, 6H, Si(CH3)2).
6.2.7. Methyl 1,2,4-tri-O-acetyl-3-O-benzyl-(a/b)-D-
glucopyranosuronate (12a/b). A solution of the Jones
reagent (44.9 mL of H2SO4 (3.5 M) and 9.7 g of CrO3) was
dropwise added to a stirred solution of 10a/b (12.5 g,
24.5 mmol) in acetone (100 mL) at 0 8C. This mixture was
immediately warmed to room temperature, stirred for 5 h
and then diluted with CH2Cl2 and H2O. The aqueous layer
was extracted three times with CH2Cl2 and the combined
extracts were dried (MgSO4), filtered and the solvent was
removed in vacuum. The residue was dissolved in
DMF (100 mL) and stirred at room temperature. CH3I
MS m/z (CI, NH3): 528 (MCNH4)C. Anal. Calcd for
C40H44O9Si (510.23): C 58.80, H 7.50; found C 58.63,
H 7.52.
6.2.6. 1,2,4-Tri-O-benzoyl-3-O-benzyl-6-O-tert-butyl-
dimethylsilyl-(a/b)-D-glucopyranose (11a/b). Sodium
hydride (9.2 g, 230 mmol, 60% in mineral oil) was added
to a stirred solution of diacetone glucose (30 g, 115 mmol)
in anhydrous DMF (300 mL) and benzyl bromide (27.4 mL,

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
573
(8.2 mL, 122.5 mmol) and KHCO3 (15.3 g, 147 mmol)
were added to the mixture. After one night of stirring, the
suspension was filtered through Celite, the filtrate was
diluted with H2O, extracted three times with Et2O, dried
(MgSO4) and evaporated in vacuum. The residue was
purified by flash chromatography (eluent, 20% EtOAc in
cyclohexane) to afford a mixture of 12a/b (a/bZ1/1,
6.75 g, 65%) as a white powder.
was diluted with DCM, washed with HCl (1 M), saturated
sodium hydrogen carbonate and brine. The organic layer
was dried (MgSO4), filtered and evaporated in vacuum. The
residue was purified by flash chromatography (eluent, 20%
EtOAc in cyclohexane) to afford a mixture of 14a/b (2.44 g,
82%) as a white powder.
Compound 14b. 1H NMR (400 MHz, CDCl3): d 5.77 (d, 1H,
J1–2Z7.9 Hz, H-1), 5.35 (t, 1H, J3–2ZJ3–4Z9.4 Hz, H-3),
5.24 (t, 1H, J4–5Z9.7 Hz, H-4), 5.17 (dd, 1H, H-2), 4.20 (d,
1H, H-5), 3.76 (s, 3H, –CO2CH3), 2.75 and 2.51 (m, 4H,
CH3C]O–CH2–CH2–), 2.17, 2.13, 2.09 and 2.08 (4s, 12H,
3 –C]O–CH3 and CH3C]O–CH2).
1
Compound 12b. H NMR (400 MHz, CDCl3): d 7.47–7.17
(m, 5H, arom.), 5.63 (d, 1H, J1–2Z7.5 Hz, H-1), 5.17 (dd,
1H, J4–3Z9.0 Hz, J4–5Z9.4 Hz, H-4), 5.10 (dd, 1H, J2–3
Z
8.9 Hz, H-2), 4.64 (d, 1H, JgemZ12.0 Hz, CHPh), 4.58 (d,
1H, JgemZ12.0 Hz, CHPh), 4.00 (d, 1H, H-5), 3.73 (t, 1H,
H-3), 3.65 (s, 3H, –CO2CH3), 1.94 and 2!1.93 (2s, 9H, 3
–O–C]O–CH3).
Compound 14a. 1H NMR (400 MHz, CDCl3): d 6.40 (d, 1H,
J1–2Z3.7 Hz, H-1), 5.57 (t, 1H, J3–2ZJ3–4Z9.9 Hz, H-3),
5.24 (t, 1H, J4–5Z9.7 Hz, H-4), 5.16 (dd, 1H, H-2), 4.40 (d,
1H, H-5), 3.77 (s, 3H, –CO2CH3), 2.75 and 2.51 (m, 4H,
CH3C]O–CH2–CH2–), 2.20, 2.18, 2.10 and 2.07 (4s, 12H,
3 –C]O–CH3 and CH3C]O–CH2); MS m/z (CI, NH3):
450 (MCNH4)C. Anal. Calcd for C18H24O12 (432.38): C
50.00, H 5.59; found C 49.91, H 5.61.
1
Compound 12a. H NMR (400 MHz, CDCl3): d 7.47–7.17
(m, 5H, arom.), 6.32 (d, 1H, J1–2Z3.6 Hz, H-1), 5.18 (dd,
1H, J4–3Z9.4 Hz, J4–5Z9.9 Hz, H-4), 5.02 (dd, 1H, J2–3
Z
9.4 Hz, H-2), 4.58 (s, 2H, CH2Ph), 4.26 (d, 1H, H-5), 3.93 (t,
1H, H-3), 3.64 (s, 3H, –CO2CH3), 2.09, 2.03 and 1.91 (3s,
9H, 3 –O–C]O–CH3); MS m/z (CI, NH3): 442 (MC
NH4)C; l. Calcd for C20H24O10 (424.407): C 56.60, H 5.70;
found C 56.52, H 5.72.
6.2.10. Methyl 1,2,4-tri-O-benzoyl-3-O-levulinyl-(a/b)-
D-glucopyranosuronate (15a/b). Hydrogenation and
protection with levulinic anhydride of 15a/b (2.96 g,
4.85 mmol), as described for 15a/b, yielded 15a/b (2.4 g,
80%) as a white powder. A small amount of this mixture
was selectively crystallized from Et2O and gave pure 15b as
a white powder.
6.2.8. Methyl 1,2,4-tri-O-benzoyl-3-O-benzyl-(a/b)-
D-glucopyranosuronate (13a/b). Oxidation and
esterification of 11a/b (12.33 g, 17.70 mmol), as described
for 12a/b, yielded 13a/b (7.35 g, 68%) as a white powder.
1
Compound 13b. H NMR (400 MHz, CDCl3): d 8.08–7.16
Compound 15b. [a]D K9 (c 1 in chloroform); mp:
153–155 8C (Et2O); 1H NMR (400 MHz, CDCl3): d
8.10–7.43 (m, 15H, arom.), 6.26 (d, 1H, J1–2Z7.5 Hz, H-
1), 5.84 (t, 1H, J3–2ZJ3–4Z8.9 Hz, H-3), 5.72 (dd, 1H, H-
2), 5.67 (t, 1H, J4–5Z9.0 Hz, H-4), 4.54 (d, 1H, H-5), 3.66
(s, 3H, –CO2CH3), 2.58 and 2.47 (2t, 4H, CH3C]O–CH2–
CH2–), 1.97 (s, 3H, CH3C]O–CH2); 13C NMR (100 MHz):
d 205.3 (C]O), 171.6, 166.9, 165.2, 164.9 and 164.4
(5O–C]O), 2!128.7 and 128.2 (3C arom.), 133.9–128.5
(15CH arom.), 92.1 (C-1), 73.2 (C-5), 71.1 (C-3), 70.0
(C-2), 69.4 (C-4), 52.9 (–CO2CH3), 37.7 and 27.9
(CH3C]O–CH2–CH2–), 29.3 (CH3C]O–CH2); MS m/z
(CI, NH3): 636.3 (MCNH4)C. Anal. Calcd for C33H30O12
(618.59): C 64.07, H 4.88; found C 63.98, H 4.90.
(m, 20H, arom.), 6.29 (d, 1H, J1–2Z6.2 Hz, H-1), 5.86 (t,
1H, J4–3ZJ4–5Z7.4 Hz, H-4), 5.70 (dd, 1H, J2–3Z7.4 Hz,
H-2), 4.82 (d, 1H, JgemZ11.4 Hz, CHPh), 4.76 (d, 1H,
JgemZ11.4 Hz, CHPh), 4.57 (d, 1H, H-5), 4.29 (t, 1H, H-3),
3.55 (s, 3H, –CO2CH3); 13C NMR (100 MHz): d 167.6,
165.2, 164.9, and164.6 (4O–C]O), 136.9, 2!129.0 and
128.9 (4C arom.), 136.9–127.8 (20CH arom.), 91.5 (C-1),
76.5 (C-3), 73.9 (CH2Ph), 73.1 (C-5), 70.6 (C-2), 70.0 (C-4),
52.8 (–CO2CH3).
1
Compound 13a. H NMR (400 MHz, CDCl3): d 8.08–7.16
(m, 20H, arom.), 6.87 (d, 1H, J1–2Z3.5 Hz, H-1), 5.86 (dd,
1H, J4–3Z9.0 Hz, J4–5Z9.3 Hz, H-4), 5.70 (dd, 1H, J2–3
Z
9.1 Hz, H-2), 4.78 (s, 2H, CH2Ph), 4.70 (d, 1H, H-5), 4.29 (t,
1H, H-3), 3.71 (s, 3H, –CO2CH3); 13C NMR (100 MHz): d
167.6, 165.2, 165.1 and 164.0 (4O–C]O), 136.9, 2!129.0
and 128.9 (4C arom.), 136.9–127.8 (20CH arom.), 89.8
(C-1), 75.6 (C-3), 74.6 (CH2Ph), 71.5 (C-5), 71.0 (C-2), 70.9
(C-4), 52.9 (–CO2CH3); MS m/z (CI, NH3): 628.1 (MC
NH4)C. Anal. Calcd for C35H30O10 (610.62): C 68.84, H
4.95; found C 68.76, H 4.91.
1
Compound 15a. H NMR (400 MHz, CDCl3): d 8.10–7.40
(m, 15H, arom.), 6.85 (d, 1H, J1–2Z3.7 Hz, H-1), 6.06 (t,
1H, J3–2ZJ3–4Z10.0 Hz, H-3), 5.59 (t, 1H, J4–5Z10.0 Hz,
H-4), 5.23 (dd, 1H, H-2), 4.66 (d, 1H, H-5), 3.67 (s, 3H,
–CO2CH3), 2.53 and 2.42 (2t, 4H, CH3C]O–CH2–CH2–),
1.92 (s, 3H, CH3C]O–CH2); 13C NMR (100 MHz): d
205.3 (C]O), 171.8, 167.2, 165.2, 165.1 and 164.0
(5O–C]O), 2!128.7 and 128.2 (3C arom.), 133.9–128.5
(15CH arom.), 89.6 (C-1), 70.8 (C-5), 2!69.7 (C-2 and
C-4), 69.2 (C-3), 53.0 (–CO2CH3), 37.7 and 27.9
(CH3C]O–CH2–CH2–), 29.3 (CH3C]O–CH2).
6.2.9. Methyl 1,2,4-tri-O-acetyl-3-O-levulinyl-(a/b)-D-
glucopyranosuronate (14a/b). A solution of 12a/b (3 g,
7.07 mmol) in MeOH (10 mL) and EtOAc (10 mL) was
hydrogenated in the presence of 10% Pd/C (100 mg) for 1 h
at room temperature, filtered through Celite and concen-
trated. The residue was dissolved in pyridine (20 mL) and
stirred at room temperature. Levulinic anhydride (5.25 g,
21.21 mmol) was added and the mixture was stirred
overnight. Solvent was removed in vacuum. The residue
6.2.11. Methyl 2,4-di-O-acetyl-1-bromo-1-deoxy-3-O-
levulinyl-a-D-glucopyranosyluronate (16). Hydrobromic
acid 33% in AcOH (9 mL, excess) was added to a stirred
solution of 14a/b (1.4 g, 3.23 mmol) in CH2Cl2 (8 mL) at
room temperature. The mixture was stirred for 2 h and

574
R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
neutralized with saturated sodium hydrogencarbonate. The
organic layer was washed with H2O, dried (MgSO4), filtered
and evaporated in vacuum. The residue was purified by flash
chromatography (eluent, 25% EtOAc in cyclohexane) to
afford 16 (1.31 g, 90%) as a white unstable foam; 1H NMR
(400 MHz, CDCl3): d 6.54 (d, 1H, J1–2Z4.0 Hz, H-1), 5.57
H-4A), 3.88 (dd, 1H, J2–3Z9.9 Hz, H-2D), 3.87–3.84 (m,
1H, H-6Bb), 3.81 (dd, 1H, H-3D), 3.78–3.74 (m, 3H, H-6A
and H-6Ba), 3.70 (t, 1H, J4–3ZJ4–5Z10.0 Hz, H-4C), 3.68
(s, 3H, CO2CH3), 3.67–3.62 (m, 2H, H-2C and H-2B),
3.62–3.58 (m, 1H, –CH–CH2Si), 3.72 (t, 1H, J3–2Z9.1 Hz,
H-3A), 3.58–3.52 (m, 3H, H-3C, H-3B and H-5B), 3.53–
3.50 (m, 2H, H-6C), 3.49–3.44 (m, 2H, H-5D and H-5C),
3.42 (dd, 1H, H-2A), 3.40–3.37 (m, 1H, H-5A), 2.53 and 2.42
(t, 1H, J3–2ZJ3–4Z10.0 Hz, H-3), 5.17 (dd, 1H, J4–5
Z
10.3 Hz, H-4), 4.80 (dd, 1H, H-2), 4.50 (d, 1H, H-5), 3.70 (s,
3H, –CO2CH3), 2.70 and 2.45 (m, 4H, CH3C]O–CH2–
CH2–), 2.09, 2.06 and 2.03 (3s, 9H, 3 –C]O–CH3 and
CH3C]O–CH2).
(2t, 4H, JCH –CH2Z6.7 Hz, CH3C]O–CH2–CH2–), 1.90 (s,
2
3H, CH3C]O–CH2), 1.59 (s, 3H, NHAc), 1.06 (m, 2H,
–CH2–CH2Si), 0.05 (s, 9H, Si(CH3)3); 13C NMR (100 MHz):
d 206.0 (C]O), 172.0, 171.4, 167.8, 165.7 and 165.5
(3O–C]O, C]O–O and NHC]O), 139.5, 139.3, 139.2,
138.8, 138.7, 138.2, 138.0, 129.7 and 129.1 (9C arom.),
134.5–126.5 (45CH arom.), 103.9 (C-1D), 103.5 (C-1A),
102.8 (C-1B), 101.8 (C-1C), 100.8 (CH, benzylidene), 100.7
(C-1E), 83.2, 2!82.3, 81.5, 79.8, 78.7, 77.0, 75.7, 75.5, 74.1,
73.4, 73.2, 72.8, 72.2, 72.1, 70.2, 69.8, 68.5, 67.3 and 56.8 (20
CH cycle), 75.8, 75.3, 74.9, 2!73.8 and 73.7 (6CH2Ph), 69.8,
2!69.0 and 68.7 (4C-6), 67.7 (–CH2–CH2Si), 53.3
(–CO2CH3), 38.2 and 28.4 (CH3C]O–CH2–CH2–), 29.7
(CH3C]O–CH2), 23.4 (NH–C]O–CH3), 18.8 (–CH2–
CH2Si), K1.0 (Si(CH3)3); MS m/z (FAB): 1955 (MCNa)C.
Anal. Calcd for C106H121NO31Si (1933.21): C 65.85, H 6.30,
N 0.72; found C 65.80, H 6.31, N 0.69.
6.2.12. Methyl 2,4-di-O-benzoyl-1-bromo-1-deoxy-3-O-
levulinyl-a-D-glucopyranosyluronate (17). Bromination
of 15a/b (2.3 g, 3.72 mmol), as described for 14a/b, yielded
17 (1.7 g, 80%) as a white unstable foam; 1H NMR
(400 MHz, CDCl3): d 8.00–7.40 (m, 10H, arom.), 6.77 (d,
1H, J1–2Z4.0 Hz, H-1), 5.94 (t, 1H, J3–2ZJ3–4Z9.8 Hz,
H-3), 5.48 (dd, 1H, J4–5Z10.2 Hz, H-4), 5.12 (dd, 1H, H-2),
4.70 (d, 1H, H-5), 3.61 (s, 3H, –CO2CH3), 2.46 and 2.35 (2t,
4H, CH3C]O–CH2–CH2–), 1.85 (s, 3H, CH3C]O–CH2);
13C NMR (100 MHz): d 205.2 (C]O), 171.6, 166.7, 165.2
and 165.1 (4O–C]O), 133.9 and 133.7 (2C arom.), 129.9–
128.2 (10CH arom.), 85.5 (C-1), 72.2 (C-5), 70.6 (C-2), 69.2
(C-3), 68.6 (C-4), 52.9 (–CO2CH3), 37.7 and 27.9
(CH3C]O–CH2–CH2–), 29.3 (CH3C]O–CH2).
6.2.14. 2-(Trimethylsilyl)ethyl (methyl 2,4-di-O-benzoyl-
3-O-levulinyl-b-D-glucopyranosyluronate)-(1/3)-(2,
4,6-tri-O-acetyl-b-D-galactopyranosyl)-(1/4)-(2-aceta-
mido-3,6-di-O-acetyl-2-deoxy-b-D-glucopyranosyl)-(1/
3)-(2,4,6-tri-O-acetyl-b-D-galactopyranosyl)-(1/4)-2,
3,6-tri-O-acetyl-b-D-glucopyranoside (19). A solution of
18 (1.2 g, 0.620 mmol) in MeOH (20 mL) was hydrogen-
ated in the presence of 10% Pd/C (200 mg) for 5 h at room
temperature, filtered through Celite and concentrated. The
residue was acetylated with acetic anhydride (15 mL)-
pyridine (30 mL) for 20 h at room temperature. Solvent
was then removed in vacuum. The residue was diluted with
DCM, washed with HCl (1 M), saturated sodium hydro-
gencarbonate and brine. The organic layer was dried
(MgSO4), filtered and evaporated in vacuum. The residue
was purified by flash chromatography (eluent, 2.5% MeOH
in DCM) to afford 19 (0.70 g, 60%) as a white powder.
[a]D C6 (c 0.4 in chloroform); mp: 168–172 8C (DCM/
MeOH); 1H NMR (400 MHz, CDCl3): d 8.00–7.47 (m,
10H, arom.), 5.62 (dd, 1H, J3–2Z8.5 Hz, J3–4Z9.2 Hz,
H-3E), 5.54 (dd, 1H, J4–5Z9.7 Hz, H-4E), 5.50 (d, 1H,
J4–3Z3.1 Hz, H-4D or B), 5.40 (d, 1H, JNHAc-2Z8.8 Hz,
NHAc), 5.30 (d, 1H, J4–3Z3.4 Hz, H-4D or B), 5.26 (dd,
1H, J2–1Z7.3 Hz, H-2E), 5.18 (t, 1H, J3–2ZJ3–4Z9.3 Hz,
H-3A), 5.09 (t, 1H, J3–2ZJ3–4Z9.3 Hz, H-3C), 5.07 (dd,
1H, J2–3Z10.5 Hz, J2–1Z8.0 Hz, H-2D or B), 5.00 (dd,
1H, J2–3Z9.2 Hz, J2–1Z8.0 Hz, H-2D or B), 4.90 (dd, 1H,
J2–1Z8.0 Hz, H-2A), 4.87 (d, 1H, H-1E), 4.68 (de, 1H,
J6b–6aZ10.5 Hz, H-6Cb), 4.59 (d, 1H, J1–2Z7.7 Hz,
6.2.13. 2-(Trimethylsilyl)ethyl(methyl 2,4-di-O-benzoyl-
3-O-levulinyl-b-D-glucopyranosyluronate)-(1/3)-(4,6-
O-benzylidene-b-D-galactopyranosyl)-(1/4)-(2-aceta-
mido-6-O-benzyl-2-deoxy-b-D-glucopyranosyl)-(1/3)-
(2,6-di-O-benzyl-b-D-galactopyranosyl)-(1/4)-2,3,6-tri-
O-benzyl-b-D-glucopyranoside (18). To a stirred mixture
˚
of 8 (1.8 g, 1.25 mmol), 17 (0.73 g, 1.25 mmol) and MS-4 A
(2.5 g) in anhydrous CH2Cl2 (15 mL) was added, at 0 8C and
under argon, AgOTf (0.386 g, 1.5 mmol). After 1 h at 0 8C,
one more equivalent of 17 and 1.2 equiv of AgOTf were
added to the mixture, which was stirred for 2 h at room
temperature, neutralized by Et3N and filtered through
Celite. The filtrate was successively washed with saturated
sodium thiosulfate, brine, dried (MgSO4) and evaporated in
vacuum. The residue was purified by flash chromatography
(eluent gradient, 2% MeOH in DCM) to afford first 18
(1.2 g, 50%) as a white powder and then 8 (0.54 g, 30%).
[a]D C2 (c 0.15 in chloroform); mp: 183–185 8C (DCM/
MeOH); 1H NMR (400 MHz, CDCl3): d 8.03–7.25 (m, 45H,
arom.), 5.68 (t, 1H, J3–2ZJ3–4Z9.5 Hz, H-3E), 5.60 (t, 1H,
`
J4–5Z9.5 Hz, H-4E), 5.48 (s, 1H, benzylidene), 5.43 (dd,
1H, J2–1Z7.4 Hz, H-2E), 5.42 (d, 1H, H-1E), 5.12 (m, 1H,
NHAc), 5.03 (d, 1H, JgemZ10.7 Hz, CHPh), 4.93 (d, 1H,
JgemZ11.1 Hz, CHPh), 4.90 (d, 1H, JgemZ12.4 Hz, CHPh),
4.78 (d, 1H, JgemZ10.7 Hz, CHPh), 4.76 (d, 1H, Jgem
Z
11.1 Hz, CHPh), 4.74 (d, 1H, J1–2Z7.8 Hz, H-1C), 4.62 (d,
1H, JgemZ12.4 Hz, CHPh), 4.61 (d, 1H, JgemZ12.1 Hz,
CHPh), 4.55 (d, 1H, JgemZ12.0 Hz, CHPh), 4.50 (d, 1H,
JgemZ12.0 Hz, CHPh), 4.48 (d, 1H, J1–2Z8.0 Hz, H-1B),
H-1C), 4.49 (d, 1H, H-1A), 4.46 (de, 1H, J6b–6a
Z
4.47 (d, 1H, JgemZ11.8 Hz, CHPh), 4.43 (d, 1H, Jgem
Z
11.9 Hz, H-6Ab), 4.40 (d, 1H, H-1D or B), 4.35 (d, 1H,
H-1D or B), 4.24 (d, 1H, H-5E), 4.16–4.11 (m, 2H, H-6Aa
and H-6Db or H-6Bb), 4.07–4.05 (m, 2H, H-6Db or H-6Bb
and H-6Da or H-6Ba), 4.03–3.99 (m, 1H, H-6Da or
H-6Ba), 3.99–3.95 (m, 1H, –CH–CH2Si), 3.96–3.94
(m, 1H, H-6Ca), 3.90 (dd, 1H, H-3D or B), 3.82 (t, 1H,
J5–6Z6.5 Hz, H-5D or B), 3.78 (t, 1H, J5–6Z6.5 Hz, H-5D
12.1 Hz, CHPh), 4.38 (d, 1H, J1–2Z7.9 Hz, H-1A), 4.36 (d,
1H, J1–2Z7.8 Hz, H-1D), 4.35 (d, 1H, JgemZ11.8 Hz,
CHPh), 4.31 (d, 1H, H-5E), 4.30 (d, 1H, J4–3Z3.3 Hz,
H-4D), 4.22 (de, 1H, J6b–6aZ12.1 Hz, H-6Db), 4.06 (de, 1H,
J4–3Z3.6 Hz, H-4B), 4.04–4.01 (m, 1H, H-6Da), 4.01–3.99
(m, 1H, –CH–CH2Si), 3.98 (t, 1H, J4–3ZJ4–5Z9.1 Hz,

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
575
or B), 3.78–3.71 (m, 3H, H-4C, H-4A and H-3D or H-3B),
3.73 (s, 3H, CO2CH3), 3.65–3.63 (m, 1H, H-5A), 3.59 (dd,
1H, H-2C), 3.56–3.54 (m, 1H, –CH–CH2Si), 3.46–3.43 (m,
1H, H-5C), 2.49 and 2.37 (2t, 4H, JCH –CH Z6:7 Hz,
The residue was purified by flash chromatography (eluent,
2% MeOH in DCM) to afford 21 (258 mg, 75%) as a white
foam. [a]D K10 (c 0.5 in chloroform); mp: 114.5–115 8C
(DCM/MeOH); IR (film): C–N3 2109.3 cmK1; C]O
2
2
CH3–C]O–CH2–CH2–), 2.19–1.91 (12s, 36H, 11
–O–C]O–CH3 and CH3C]O–CH2), 1.68 (s, 3H,
NHAc), 1.00–0.90 (m, 2H, –CH2–CH2Si), 0.03 (s, 9H,
Si(CH3)3); 13C NMR (100 MHz): d 205.3 (C]O), 171.6,
170.5, 170.5, 2!170.4, 170.3, 170.1, 169.8, 2!169.7,
169.5, 168.8, 168.3, 166.7, 164.9 and 164.5 (15O–C]O),
129.0 and 128.8 (2C arom.), 133.5–128.4 (10CH arom.),
100.7 (C-1D or B), 100.6 (C-1E and C-1D or B), 100.4
(C-1C), 99.9 (C-1A), 76.8 (C-3D or B), 75.8 (C-4A), 75.7
(C-3D or B), 75.0 (C-4C), 72.7 (C-5E and C-3A), 72.6
(C-5C), 72.5 (C-5A), 71.6 (C-3C and C-2A), 71.5 (C-3E),
71.4 (C-2E), 71.1 (C-5D or B), 71.0 (C-5D or B), 70.9
(C-2D or B), 70.5 (C-2D or B), 69.5 (C-4E), 68.7 (C-4D or
B), 68.1 (C-4D or B), 67.4 (–CH2–CH2Si), 62.2 (C-6A),
61.7 and 61.5 (C-6D and C-6B), 60.4 (C-6C), 54.4 (C-2C),
52.8 (–CO2CH3), 37.6 and 27.9 (CH3C]O–CH2–CH2–),
29.2 (CH3C]O–CH2), 23.0–20.1 (11 –O–C]O–CH3 and
1 NH–C]O–CH3), 17.8 (–CH2–CH2Si), K1.5 (Si(CH3)3);
MS m/z (CI, NH3): 1783.0 (MCNH4)C. Anal. Calcd for
C79H103NO42Si.1H2O: C 53.16, H 5.93, N 0.78; found C
52.73, H 5.93, N 0.61.
1758.5 cmK1 1H NMR (400 MHz, CDCl3): d 8.08–7.46
;
(m, 15H, arom.), 5.95 (dt, 1H, J5–4Z15.0 Hz, J5-CH Z6.7 Hz,
2
H-5 sphingosine), 5.63 (dd, 1H, J3–2Z9.1 Hz, J3–4Z9.5 Hz,
H-3E), 5.63–5.61 (m, 1H, H-3 sphingosine), 5.61–5.60 (m,
1H, H-4 sphingosine), 5.54 (dd, 1H, J4–5Z9.7 Hz, H-4E),
5.50 (de, 1H, J4–3Z3.4 Hz, H-4D or B), 5.38 (d, 1H,
JNHAc-2Z8.7 Hz, NHAc), 5.30 (d, 1H, J4–3Z3.8 Hz, H-4D or
B), 5.27 (dd, 1H, J2–1Z7.3 Hz, H-2E), 5.18 (t, 1H, J3–2
J3–4Z9.2 Hz, H-3A), 5.09 (dd, 1H, J3–2Z10.0 Hz, J3–4
Z
Z
9.3 Hz, H-3C), 5.07 (dd, 1H, J2–3Z10.0 Hz, J2–1Z8.0 Hz,
H-2D or B), 5.00 (dd, 1H, J2–3Z10.0 Hz, J2–1Z8.0 Hz, H-2D
or B), 4.90 (dd, 1H, J2–1Z7.8 Hz, H-2A), 4.87 (d, 1H, H-1E),
4.69 (dd, 1H, J6b–6aZ12.0 Hz, J6b–5Z2.2 Hz, H-6Cb), 4.59
(d, 1H, J1–2Z7.6 Hz, H-1C), 4.53 (d, 1H, H-1A), 4.46 (dd,
1H, J6b–6aZ11.3 Hz, J6b–5Z1.6 Hz, H-6Ab), 4.40 (d, 1H, H-
1D or B), 4.36 (d, 1H, H-1D or B), 4.25 (d, 1H, H-5E), 4.13
(dd, 1H, J6b–6aZ11.5 Hz, J6b–5Z6.2 Hz, H-6Bb), 4.09–3.99
(m, 4H, H-6Aa, H-6D and H-6Ba), 3.98–3.94 (m, 1H, H-2
sphingosine), 3.92 (dd, 1H, J6a–5Z4.0 Hz, H-6Ca), 3.90 (dd,
1H, H-3D or H-3B), 3.91–3.86 (m, 1H, H-1b sphingosine),
3.83–3.76 (m, 2H, H-5D and H-5B), 3.79–3.73 (m, 3H, H-4C,
H-4A and H-3D or H-3B), 3.73 (s, 3H, CO2CH3), 3.64–3.57
(m, 3H, H-5A, H-2C and H-1a sphingosine), 3.44 (dt, 1H,
J5–4Z9.3 Hz, H-5C), 2.50 and 2.37 (2t, 4H,
JCH –CH Z6:4 Hz, CH3–C]O–CH2–CH2–), 2.19–1.91
6.2.15. (Methyl 2,4-di-O-benzoyl-3-O-levulinyl-b-D-glu-
copyranosyluronate)-(1/3)-(2,4,6-tri-O-acetyl-b-D-
galactopyranosyl)-(1/4)-(2-acetamido-3,6-di-O-acetyl-
2-deoxy-b-D-glucopyranosyl)-(1/3)-(2,4,6-tri-O-acetyl-
b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-acetyl-a-D-
glucopyranosyl) trichloroacetimidate (20). To a stirred
solution of 19 (500 mg, 0.283 mmol) in CH2Cl2 (15 mL)
was added, at 0 8C, trifluoroacetic acid (2.4 mL,
31.3 mmol). After 4 h at room temperature, 1 mL of
trifluoroacetic acid was added. The mixture, after stirring
for 3 h, was diluted with CH2Cl2, neutralized with saturated
sodium hydrogencarbonate. The organic layer was dried
(MgSO4), filtered and evaporated in vacuum. The residue
was dissolved in anhydrous CH2Cl2 (7 mL) and stirred at
0 8C under argon. Trichloroacetonitrile (0.426 mL,
4.25 mmol) and DBU (9 mL, 0.056 mmol) were succes-
sively added and, after stirring for 30 min, the mixture was
directly purified by flash chromatography (eluent, 4%
MeOH in DCM) to afford 20 (476 mg, 92%) as a white
unstable foam; 1H NMR (400 MHz, CDCl3): d 8.60 (s, 1H,
–O–C]NH–CCl3), 6.45 (d, 1H, J1–2Z3.7 Hz, H-1A).
2
2
(12s, 36H, 11 –O–C]O–CH3 and CH3C]O–CH2), 1.70
(s, 3H, NHAc), 1.47–1.21 (m, 24H, CH2 sphingosine), 0.91 (t,
3H, JCH –CH Z7:0 Hz, CH3 sphingosine); 13C NMR
3
2
(100 MHz): d 205.3 (C]O), 171.6, 170.6, 170.5, 2!170.4,
170.3, 170.2, 2!169.8, 169.7, 169.5, 168.8, 168.4, 166.7,
165.0, 164.9 and 164.5 (17O–C]O), 129.8, 129.0 and 128.7
(3C arom.), 133.5–128.4 (15CH arom.), 100.7 (C-1D and C-
1B), 100.6 (C-1E), 100.4 (C-1C), 100.3 (C-1A), 76.8 (C-3D
or C-3B), 75.7 (C-4A or C-4C), 75.6 (C-4C or C-4A), 75.0 (C-
3D or C-3B), 74.6 (C-3 sphingosine), 72.7 (C-5E and C-5A),
72.6 (C-5C), 72.5 (C-3A), 71.6 (C-2D or C-2B), 71.5 (C-2A),
71.4 (C-3E), 71.3 (C-2E), 71.2 (C-5D or C-5B), 71.0 (C-5D or
C-5B), 70.9 (C-2D or C-2B), 70.5 (C-3C), 69.5 (C-4E), 68.7
(C-4D or C-4B), 68.3 (C-1 sphingosine), 68.1 (C-4D or
C-4B), 63.4 (C-2 sphingosine), 61.9 (C-6A), 61.7 and 61.5
(C-6D and C-6B), 60.4 (C-6C), 54.4 (C-2C), 52.8
(–CO2CH3), 37.6 and 27.8 (CH3C]O–CH2–CH2–), 29.2
(CH3C]O–CH2), 32.9, 31.8, 29.6–28.6 and 22.6 (CH2
sphingosine), 23.0–20.1 (11 –O–C]O–CH3 and 1
NH–C]O–CH3), 14.0 (CH3 sphingosine); MS m/z (FAB):
2099.8 (MCNa)C. Anal. Calcd for C99H128N4O44
(2078.159): C 57.21, H 6.20, N 2.69; found C 56.73, H
6.20, N 2.47.
6.2.16. (Methyl 2,4-di-O-benzoyl-3-O-levulinyl-b-
D-glucopyranosyluronate)-(1/3)-(2,4,6-tri-O-acetyl-
b-D-galactopyranosyl)-(1/4)-(2-acetamido-3,6-di-O-
acetyl-2-deoxy-b-D-glucopyranosyl)-(1/3)-(2,4,6-tri-O-
acetyl-b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-acetyl-
b-D-glucopyranosyl)-(1/1)-(2S,3R,4E)-2-azido-3-O-
benzoyl-4-octadecene-1,3-diol (21). To a stirred mixture of
20 (300 mg, 0.165 mmol), acceptor (2S,3R,4E)-2-azido-3-
O-benzoyl-4-octadecene-1,3-diol (180 mg, 0.414 mmol)
6.2.17. (Methyl 2,4-di-O-benzoyl-3-O-levulinyl-b-D-
glucopyranosyluronate)-(1/3)-(2,4,6-tri-O-acetyl-b-D-
galactopyranosyl)-(1/4)-(2-acetamido-3,6-di-O-acetyl-
2-deoxy-b-D-glucopyranosyl)-(1/3)-(2,4,6-tri-O-acetyl-
b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-acetyl-b-D-
glucopyranosyl)-(1/1)-(2S,3R,4E)-2-octadecanamido-
3-O-benzoyl-4-octadecene-1,3-diol (22). Compound 21
(165 mg, 79.4 mmol) and PPh3 (62.4 mg, 238.2 mmol)
were dissolved in a mixture of THF/H2O (3 mL/0.6 mL)
and stirred overnight at 50 8C. Solvents were evaporated in
˚
and MS-4 A (500 mg) in anhydrous CH2Cl2 (5 mL) was
added, at 0 8C and under argon, BF3$Et2O (0.063 mL,
0.495 mmol). After stirring for 3 h at 0 8C, the mixture was
neutralized with saturated sodium hydrogencarbonate and
filtered through Celite. The organic layer was washed with
brine, dried (MgSO4), filtered and evaporated in vacuum.

576
R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
vacuum. The residue was purified by flash chromatography
to afford a foam, which was directly engaged in the next
step. To a stirred solution of this foam in CH2Cl2 (4 mL)
were successively added stearic acid (42 mg, 198.5 mmol)
and DCC (45 mg, 158.8 mmol) at room temperature. After
stirring overnight, solvent was evaporated in vacuum. The
residue was purified by flash chromatography (eluent, 3%
MeOH in CH2Cl2) to afford 22 (143 mg, 78%) as a white
foam. [a]D C12 (c 0.26 in chloroform); mp: 172.5–173 8C
6.2.18. (Methyl 2,4-di-O-benzoyl-3-O-sulfo-b-D-gluco-
pyranosyluronate)-(1/3)-(2,4,6-tri-O-acetyl-b-D-galac-
topyranosyl)-(1/4)-(2-acetamido-3,6-di-O-acetyl-2-
deoxy-b-D-glucopyranosyl)-(1/3)-(2,4,6-tri-O-acetyl-
b-D-galactopyranosyl)-(1/4)-2,3,6-tri-O-acetyl-b-D-
glucopyranosyl)-(1/1)-(2S,3R,4E)-2-octadecanamido-
3-O-benzoyl-4-octadecene-1,3-diol sodium salt (23).
Hydrazine monoacetate (20 mg, 215 mmol) was added to a
stirred solution of 22 (100 mg, 43 mmol) in EtOH (4 mL) at
room temperature. After stirring for 1 h, the mixture was
neutralized with saturated sodium hydrogen carbonate,
washed with brine, dried (MgSO4), filtered and evaporated
in vacuum. The residue was purified by flash chromato-
graphy (eluent, 2% MeOH in DCM) to afford the 3-OH
compound (77 mg, 81%) as a white foam. 1H RMN
(400 MHz, CDCl3): d 4.15–4.1 (m, 1H, H-3E). SO3.Me3N
(72 mg, 520.5 mmol) was added to a stirred solution of the
previous foam (77 mg, 34.7 mmol) in DMF (3 mL). The
mixture was stirred at 40 8C for 20 h. MeOH (0.2 mL) and
CH2Cl2 (0.2 mL) were added, and the solution was applied
to a column of Sephadex LH-20 with 1:1 DCM/MeOH
eluent. Glycolipid containing fractions were concentrated.
Column chromatography (MeOH) of the residue on Dowex-
50*2 (NaC) resin gave 23 (65 mg, 81%) as an amorphous
powder. [a]D C10 (c 0.2 in chloroform); 1H NMR
(400 MHz, CDCl3/CD3OD): d 7.70–7.05 (m, 15H, arom.),
1
(DCM/MeOH); H NMR (400 MHz, CDCl3): d 8.00–7.45
(m, 15H, arom.), 5.89 (dt, 1H, J5–4Z15.0 Hz,
J5ꢀCH Z6:8 Hz, H-5 ceramide), 5.76 (d, 1H, JNH-2
Z
2
9.3 Hz, NH ceramide), 5.63 (t, 1H, J3–4ZJ3–2Z9.4 Hz,
H-3E), 5.58–5.55 (m, 1H, H-3 ceramide), 5.55 (t, 1H, J4–5
9.5 Hz, H-4E), 5.50 (d, 1H, J4–3Z3.4 Hz, H-4D or H-4B),
5.52–5.47 (m, 1H, H-4 ceramide), 5.35 (d, 1H, JNHAc-2
8.7 Hz, NHAc), 5.30 (d, 1H, J4–3Z3.4 Hz, H-4D or H-4B),
5.26 (dd, 1H, J2–1Z7.3 Hz, H-2E), 5.18 (t, 1H, J3–2ZJ3–4
Z
Z
Z
9.5 Hz, H-3A), 5.08 (dd, 1H, J2–3Z10.2 Hz, J2–1Z8.0 Hz,
H-2D or H-2B), 5.08 (t, 1H, J3–2ZJ3–4Z9.2 Hz, H-3C),
5.00 (dd, 1H, J2–3Z10.0 Hz, J2–1Z8.1 Hz, H-2D or H-2B),
4.90 (dd, 1H, J2–3Z9.5 Hz, J2–1Z7.8 Hz, H-2A), 4.88 (d,
1H, H-1E), 4.69 (dd, 1H, J6b–6aZ10.7 Hz, J6b–5Z3.2 Hz,
H-6Cb), 4.57 (d, 1H, J1–2Z7.8 Hz, H-1C), 4.55–4.46 (m,
1H, H-2 ceramide), 4.46 (d, 1H, H-1A), 4.40 (d, 1H, H-1D
or H-1B), 4.33 (de, 1H, J6b–6aZ11.8 Hz, H-6Ab), 4.32 (d,
1H, H-1D or H-1B), 4.24 (d, 1H, H-5E), 4.13 and 4.00 (2dd,
2H, J6b–6aZ11.5 Hz, J6b–5ZJ6a–5Z6.2 Hz, H-6D or H-6B),
4.07–4.05 (m, 2H, H-6D or H-6B), 4.05–4.02 (m, 1H, H-1b
ceramide), 3.98 (dd, 1H, J6a–5Z3.4 Hz, H-6Aa), 3.92 (dd,
1H, J6a–5Z3.2 Hz, H-6Ca), 3.90 (dd, 1H, H-3D or H-3B),
3.82 (t, 1H, J5–6Z6.2 Hz, H-5D or H-5B), 3.78–3.76 (m,
1H, H-5D or H-5B), 3.76–3.70 (m, 3H, H-4C, H-4A and
H-3D or H-3B), 3.73 (s, 3H, CO2CH3), 3.68–3.62 (m, 1H,
H-1a ceramide), 3.62–3.60 (m, 1H, H-2C), 3.61–3.58 (m,
1H, H-5A), 3.44 (dt, 1H, J5–4Z9.2 Hz, H-5C), 2.50 and 2.37
(2t, 4H, JCH –CH Z7:0 Hz, CH3–C]O–CH2–CH2–), 2.19–
5.50 (dt, 1H, J5–4Z13.4 Hz, J
mide), 5.18–5.12 (m, 1H, H-3 ceramide), 5.10 (d, 1H, J4–3
Z6:8 Hz, H-5 cera-
Z
3.4 Hz, H-4D or H-4B), 5.12–5.07 (m, 1H, H-4 ceramide),
5ꢀCH2
5.00 (t, 1H, J4–3ZJ4–5Z9.7 Hz, H-4E), 4.94 (d, 1H, J4–3
3.5 Hz, H-4D or H-4B), 4.84 (dd, 1H, J2–1Z7.8 Hz, J2–3
Z
Z
9.7 Hz, H-2E), 4.74 (t, 1H, J3–2ZJ3–4Z9.4 Hz, H-3A), 4.70
(dd, 1H, J3–2Z9.3 Hz, J3–4Z9.5 Hz, H-3C), 4.61 (t, 1H, H-
3E), 4.59 (dd, 1H, J2–3Z9.5 Hz, J2–1Z8.0 Hz, H-2D or H-
2B), 4.58 (d, 1H, H-1E), 4.52 (dd, 1H, J2–3Z9.9 Hz, J2–1
8.0 Hz, H-2D or H-2B), 4.37 (dd, 1H, J2–1Z8.0 Hz, H-2A),
4.24 (d, 1H, J1–2Z8.2 Hz, H-1C), 4.08 (de, 1H, J6b–6a
Z
Z
9.7 Hz, H-6Cb), 4.17 (d, 1H, H-1A), 4.09–4.04 (m, 1H, H-2
ceramide), 4.06 (d, 1H, H-1D or H-1B), 4.00 (d, 1H, H-1D
or H-1B), 3.83 (d, 1H, J6b–6aZ10.0 Hz, H-6Ab), 3.83 (d,
1H, H-5E), 3.73–3.55 (m, 7H, H-6Aa, H-5D, H-5B, H-6D
and H-6B), 3.63 (dd, 1H, H-3D or H-3B), 3.58–3.55 (m, 1H,
H-1b ceramide), 3.54–3.50 (m, 1H, H-6Ca), 3.39 (dd, 1H,
H-3D or H-3B), 3.38 (t, 1H, H-5D or H-5B), 3.22 (t, 1H,
J4–5Z9.5 Hz, H-4C), 3.28–3.25 (m, 1H, H-1a ceramide),
3.27 (s, 3H, CO2CH3), 3.25–3.22 (m, 1H, H-5A), 3.10 (dd,
1H, H-2C), 2.99 (dt, 1H, H-5C), 1.75–1.58 (11s, 33H,
–O–C]O–CH3), 1.50 (s, 3H, NHAc), 1.82–1.70 and
0.94–0.87 (m, 56H, CH2 ceramide), 0.49 (2t, 6H,
JCH –CH Z7:1 Hz, CH3 ceramide); 13C NMR (100 MHz):
2
2
1.91 (12s, 36H, 11 –O–C]O–CH3 and CH3C]O–CH2),
1.70 (s, 3H, NHAc), 1.70–1.68 and 1.29–1.25 (m, 56H, CH2
ceramide), 0.91 (2t, 6H, JCH –CH Z7:0 Hz, CH3 ceramide);
3
2
13C NMR (100 MHz): d 205.3 (C]O), 172.6, 171.6, 170.6,
170.5, 2!170.4, 170.3, 170.2, 2!169.8, 169.7, 169.6,
168.8, 168.4, 166.7, 165.1, 164.9 and 164.6 (17O–C]O and
1 N–C]O), 137.6 (C-5 ceramide), 133.5–128.4 (15CH
arom.), 130.2, 129.1 and 128.8 (3C arom.), 124.6 (C-4
ceramide), 100.7 (C-1D or C-1B), 100.6 (C-1E and C-1D or
C-1B), 100.4 (C-1C), 100.2 (C-1A), 76.8 (C-3D or C-3B),
75.7 (C-4A), 75.6 (C-4C), 75.0 (C-3D or C-3B), 73.9 (C-3
ceramide), 72.7 (C-5E), 72.6 (C-5A), 72.5 (C-5C), 72.2
(C-3A), 2!71.6 (C-2D or C-2B and C-2A), 71.5 (C-3E),
71.4 (C-2E), 71.2 and 71.0 (C-5D and C-5B), 70.9 (C-2D or
C-2B), 70.5 (C-3C), 69.5 (C-4E), 68.7 (C-4D or C-4B), 68.1
(C-4D or C-4B), 67.3 (C-1 ceramide), 61.9 (C-6A), 61.7 and
61.5 (C-6D and C-6B), 60.4 (C-6C), 54.4 (C-2C), 52.8
(–CO2CH3), 50.5 (C-2 ceramide), 37.6 and 27.9
(CH3C]O–CH2–CH2–), 23.0–20.1 (11 –O–C]O–CH3
and 1 NH–C]O–CH3 and CH3C]O–), 36.8, 33.3, 31.9,
29.7–28.9, 25.7 and 22.6 (CH2 ceramide), 14.1 (CH3
ceramide); MS m/z (FAB): 2340.0 (MCNa)C. Anal.
Calcd for C117H164N2O45 (2318.15): C 60.61, H 7.12, N
1.20; found C 60.68, H 7.31, N 1.11.
3
2
d 174.0, 171.2, 170.4, 2!170.3, 170.2, 170.0, 169.9, 169.7,
169.5, 169.4, 168.8, 168.7, 167.2, 165.3 and 2!164.9
(16O–C]O and 1 N–C]O), 132.4 (C-5 ceramide), 132.2–
127.3 (15CH arom.), 129.5, 129.2 and 128.8 (3C arom.),
123.5 (C-4 ceramide), 100.0 (C-1D and C-1B), 99.8 (C-1E),
99.7 (C-1C), 99.3 (C-1A), 76.9, 76.4, 75.9, 2!75.0, 73.4,
72.2, 72, 71.7, 71.6, 71.5, 71.4, 71.0, 70.3, 70.2, 69.9, 69.8,
69.7, 68.8 and 68.1 (19 CH cycle and C-3 ceramide), 66.7
(C-1 ceramide), 61.9 (C-6C), 61.3 and 61.0 (C-6D and
C-6B), 60.2 (C-6A), 53.9 (C-2C), 51.6 (–CO2CH3), 50.2
(C-2 ceramide), 35.5, 31.5, 31.1, 28.9–28.1, 25.3 and 21.8
(CH2 ceramide), 21.3–18.9 (11 –O–C]O–CH3 and 1
NH–C]O–CH3), 12.8 (CH3 ceramide). MS m/z (FAB):

R. Chevalier et al. / Tetrahedron 62 (2006) 563–577
577
2307.9 MHC. Anal. Calcd for C112H157N2NaO46S (2308.5):
C 57.92, H 6.81, N 1.21; found C 59.13, H 6.96, N 1.10.
1597–1600. (b) Nakano, T.; Ito, Y.; Ogawa, T. Carbohydr.
Res. 1993, 243, 43–69. (c) Isogai, Y.; Ishida, H.; Kiso, M.;
Hasegawa, A. J. Carbohydr. Chem. 1996, 15, 1119–1137.
7. For previous works on lactosaminyl and lewis X glycolipids: (a)
Zhang, Y.; Esnault, J.; Mallet, J.-M.; Sinay¨, P. J. Carbohydr.
6.2.19. (3-O-Sulfo-b-D-glucopyranosyluronic acid)-
(1/3)-(b-D-galactopyranosyl)-(1/4)-(2-acetamido-2-
deoxy-b-D-glucopyranosyl)-(1/3)-(b-D-galactopyrano-
syl)-(1/4)-(b-D-glucopyranosyl)-(1/1)-(2S,3R,4E)-
2-octadecanamido-3-O-benzoyl-4-octadecene-1,3-diol
disodium salt (1). To a stirred solution of 23 (60 mg,
26 mmol) in a mixture of THF/H2O (3.8 mL/0.2 mL) was
added at 0 8C a solution of 1.25 N LiOH (0.26 mL,
1040 mmol). The mixture was stirred 3 h between 0 and
5 8C. Solvents were evaporated in vacuum. The residue was
dissolved in a mixture of THF/MeOH (2 mL/2 mL) and
sodium (cat., 0.15 M) was added at 0 8C. The solution was
stirred for 3 h at room temperature and purified on a column
of Sephadex LH-20 with CHCl3/MeOH/H2OZ6:4:0.8 to
afford SGPG (35 mg, 90%) as a white foam; 1H NMR
(600 MHz, CD3S]OCD3): d 5.54 (dt, 1H, J5–4Z15.2 Hz,
´
Chem. 1999, 18, 419–427. (b) Bregant, S.; Zhang, Y.; Mallet,
J.-M.; Brodzki, A.; Sinay¨, P. Glycoconjugate J. 1999, 16,
757–765. (c) Esnault, J.; Mallet, J.-M.; Zhang, Y.; Sinay¨, P.; Le
Bouar, T.; Pincet, F.; Perez, E. Eur. J. Org. Chem. 2001,
253–260. (d) Gourier, C.; Pincet, F.; Perez, E.; Zhang, Y.;
Mallet, J.-M.; Zhu, Z.; Sinay, P. Angew. Chem., Int. Ed. 2005,
44, 1683–1687.
8. Jansson, K.; Ahfors, S.; Frejd, T.; Kihlberg, J.; Magnusson, G.
J. Org. Chem. 1988, 53, 5629–5647.
9. Schmidt, R. R.; Zimmermann, P. Tetrahedron Lett. 1986, 27,
481–484.
10. Lemieux, R. U.; Driguez, H. J. Am. Chem. Soc. 1975, 97,
4069–4075.
11. Tabeur, C.; Mallet, J.-M.; Bonot, F.; Herbert, J.-M.; Petitou,
M.; Sinay¨, P. Bioorg. Med. Chem. 1999, 7, 2003–2012.
12. (a) Koeppem, B. H. Carbohydr. Res. 1972, 24, 154–158. (b)
Charon, D.; Szabo, L. J. Chem. Soc., Perkin. Trans. 1 1980,
1971–1977. (c) Takeo, K.; Kitamura, S.; Murata, Y.
Carbohydr. Res. 1992, 224, 111–122.
J5ꢀCH Z6:7 Hz, H-5 ceramide), 5.35 (dd, 1H, J4–5
Z
2
15.2 Hz, J4–3Z7.1 Hz, H-4 ceramide), 4.65, 4.54, 4.30,
4.27 and 4.16 (5d, 5H, 5H-1), 3.99 (t, 1H, J3–2ZJ3–4
9.0 Hz, H-3E), 1.82 (s, 3H, NHAc), 0.85 (t, 6H,
Z
JCH –CH Z7:1 Hz, CH3 ceramide); MS (MALDI-TOF):
3
2
Calcd (MCNa)CZ1532.74; foundZ1532.7; Calcd
(MKHC2Na)CZ1510.76; foundZ1510.7.
`
13. Hajko, J.; Borbas, A.; Liptak, A. Carbohydr. Res. 1991, 216,
413–420.
14. Zsiska, M.; Meyer, B. Carbohydr. Res. 1991, 215, 279–292.
15. (a) Brimacombe, J. S. Methods Carbohydr. Chem. 1972, VI,
376. (b) Garegg, P.; Maron, L. Acta Chem. Scand. 1979, B33,
39–41.
Acknowledgements
The authors thank Prof. Jacques Aubry (Nantes) for
initiating this project.
16. Domon, B.; Costello, C. E. Glycoconjugate J. 1988, 5,
397–409.
17. Willison, H. J.; Veitch, J.; Swan, A. V.; Baumann, N.; Comi,
G.; Gregson, N. A.; Illa, I.; Zielasek, J.; Hughes, R. A. Eur.
J. Neurol. 1999, 71–77.
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