Syntheses and interfacial behaviour of neoglycolipid analogues of glycosyl ceramides

Article abstract of DOI:10.1016/S0008-6215(01)00255-5

Four glycosyl ceramides analogues having D-galactose or 2-acetamido-2-deoxy-D-glucose moieties linked to enantiomeric lipids have been synthesised to study their interfacial behaviour at the air | water interface. The lipid chains were prepared in two steps by opening 1,2-epoxyhexadecane using Jacobsen kinetic hydrolytic resolution (KHR) followed by an azidosilylation reaction of the diol so obtained. Glycosylation reactions were realised either with 2,3,4,6-tetra-O-benzoyl-α-D-galactopyranosyl trichloroacetimidate or 1,3,4,6-tetra-O-acetyl-2-allyloxycarbonylamino-2-deoxy-β-D-glucopyranose as donors and (2R)- or (2S)-2-azidohexadecanol derivatives as acceptors. Transformation of the azido glycosides into N-acylated products was done by a modified Staudinger reaction in the presence of fatty acyl chlorides. The four neoglycolipids are able to form a condensed monolayer at the air | water interface; their π-A isotherm diagrams are similar to that described for the natural glycosyl ceramides. The detailed analysis of the isotherms, taking into account the chirality of the lipid chains, allowed to determine the contribution of the different parts of the molecule under the monolayer packing.

Full text of DOI:10.1016/S0008-6215(01)00255-5

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Carbohydrate Research 336 (2001) 181–194
Syntheses and interfacial behaviour of neoglycolipid
analogues of glycosyl ceramides
Dominique Lafont,a Marie-Noe¨lle Bouchu,a Agne`s Girard-Egrot,b Paul Boullangera,*
aLaboratoire de Chimie Organique II, Unite´ Mixte de Recherche CNRS 5622, Uni6ersite´ Lyon 1,
Chimie Physique Electronique de Lyon, 43 Boule6ard du 11 No6embre 1918, F-69622 Villeurbanne, France
bLaboratoire de Ge´nie Enzymatique, Unite´ Mixte de Recherche CNRS 5013, Uni6ersite´ Lyon 1,
Chimie Physique Electronique de Lyon, 43 Boule6ard du 11 No6embre 1918, F-69622 Villeurbanne, France
Received 20 July 2001; accepted 18 September 2001
Abstract
Four glycosyl ceramides analogues having
D
-galactose or 2-acetamido-2-deoxy-D-glucose moieties linked to
enantiomeric lipids have been synthesised to study their interfacial behaviour at the air ꢀ water interface. The lipid
chains were prepared in two steps by opening 1,2-epoxyhexadecane using Jacobsen kinetic hydrolytic resolution
(KHR) followed by an azidosilylation reaction of the diol so obtained. Glycosylation reactions were realised either
with 2,3,4,6-tetra-O-benzoyl-a-
D-galactopyranosyl trichloroacetimidate or 1,3,4,6-tetra-O-acetyl-2-allyloxycarbony-
lamino-2-deoxy-b- -glucopyranose as donors and (2R)- or (2S)-2-azidohexadecanol derivatives as acceptors. Trans-
D
formation of the azido glycosides into N-acylated products was done by a modified Staudinger reaction in the
presence of fatty acyl chlorides. The four neoglycolipids are able to form a condensed monolayer at the air ꢀ water
interface; their p-A isotherm diagrams are similar to that described for the natural glycosyl ceramides. The detailed
analysis of the isotherms, taking into account the chirality of the lipid chains, allowed to determine the contribution
of the different parts of the molecule under the monolayer packing. © 2001 Elsevier Science Ltd. All rights reserved.
Keywords: Neoglycosyl ceramides; Kinetic hydrolytic resolution; Glycosylation; Imidate; Staudinger reaction; Monolayers;
Interfacial behaviour; 2D compressibility
1. Introduction
been reviewed recently.2 The biosynthesis of
glycosyl ceramides results of biochemical
events leading to complex structures in which
the lipid moiety is a sphingosine amidified by
a fatty acid. The above structures are embed-
ded at the surface of cells by non-covalent
interactions between phospholipids and the
ceramide part of the glycolipids. The carbohy-
drate is endowed of recognition properties,
modulated by the nature of the lipid moiety
responsible of the self-assembling properties
of the whole. Other lipid structures, more
easily accessible by chemical syntheses than
the natural ceramides, could afford the same
type of assemblies or different assemblies giv-
Glycosphingolipids or glycosyl ceramides
are constituents of animal cell membranes
consisting of various oligosaccharides bound
to ceramides by a glycosidic bound. They
serve as identifying markers and regulate cel-
lular recognition, growth, and development.1
The chemistry of glycosphingolipids-carbohy-
drate molecules of biological significance has
* Corresponding author. Tel.: +33-4-72431162; fax: +33-
4-78898914.
E-mail address: paul.boullanger@univ-lyon1.fr (P. Boul-
langer).
0008-6215/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(01)00255-5

182
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
2. Results and discussion
ing rise to different biochemical properties.
With this purpose in mind, we have already
prepared fucopyranosyl and 2-acetamido-2-
Synthesis.—In the glycosphingolipids, the
ceramide portion consists of a sphingosine
moiety amidified with a fatty acid (usually
C16–C24). The variation in fatty acids, sphin-
gosines and carbohydrates results in a great
variety of distinct glycosphingolipids.6 In the
present paper, we describe a short synthesis of
deoxy- -glucopyranosyl neoglycolipids and
D
studied their self associations at the air ꢀ water
interface, their self organisations with phos-
pholipids and the recognition of their molecu-
lar assemblies by specific lectins.3–5 This paper
deals with a short synthesis of new glycosyl
ceramide analogues having the (R) or (S)
configuration at the b-position of the agly-
cone, and preliminary studies of their self
organisation at the air ꢀ water interface. The
neoglycosyl ceramides with
D
-galactose and
2-acetamido-2-deoxy- -glucose as the carbo-
D
hydrate moieties and (2R)- or (2S)-2-N-ac-
ylaminohexadecane as the lipidic part.
In order to obtain enantiomerically pure
derivatives, we decided to use the asymmetric
catalysis of epoxide ring opening, described
for smaller alkyl chains by Jacobsen and co-
workers.7,8 Thus, treatment of 1,2-epoxyhexa-
decane (1) with water (0.55 equiv) in THF in
the presence of (R,R)-(−)-N,N%-bis(3,5-di-
tert - butylsalicylidene) - 1,2 - cyclohexanedia-
minocobalt(III) acetate afforded the expected
(2S)-hexadecane-1,2-diol (2S) and (2R)-1,2-
epoxyhexadecane (1R) in 37 and 48% yields,
respectively. The high enantioselectivity of the
reaction (ee\95%) was confirmed by com-
parison of the optical rotation of these two
products with that of the literature.9,10 (2R)-
Hexadecane-1,2-diol (2R) was similarly ob-
tained in the presence of (S,S)-(+)-N,N%-bis-
(3,5-di-tert-butylsalicylidene)-1,2-cyclohexane-
diaminocobalt(III) acetate and a slight excess
of water (10%) either from racemic 1,2-epoxy-
hexadecane (1) or from (2R)-1,2-epoxyhexade-
cane (1R) in 37 and 78% yields, respectively
(Scheme 1).
b- -galactopyranosyl head group was chosen
D
because of its widespread occurrence at both
the reducing and non-reducing ends of natural
glycosyl ceramides. The 2-acetamido-2-deoxy-
D
-glucopyranosyl head group, uncommon in
glycosyl ceramides, was chosen for compari-
son with other neoglycolipids in their molecu-
lar associations and molecular recognition by
lectins.3–5 The Langmuir monolayers of chiral
molecules is a unique system to analyse, under
defined conditions, both the interfacial prop-
erties and the importance of chirality-depen-
dent interactions and may help to elucidate
some structural functions.
Treatment of diol 2S with triphenylphos-
phine, diisopropylazodicaboxylate (DIAD)
and azidotrimethylsilane afforded a 20:1 un-
separable mixture of (2R)-2-azido-1-tri-
methylsilyloxyhexadecane (3R) and (2S)-1-
azido-2-trimethylsilyloxyhexadecane (4S) in
high yield. The azidosilylation proceeded via a
cyclic dioxaphospholane intermediate with in-
version of configuration at C-2.11,12 Desilyla-
tion of the mixture 3R/4S was realised in the
presence of an excess of potassium carbonate
in dry methanol affording 5R (86%) and 6S
(5%), that could be separated by column chro-
matography. The same sequence, applied to
diol 2R, afforded a mixture of 3S and 4R
Scheme 1.

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
183
0 °C afforded the desired glycosides 9R and
13
9S in 91 and 92% yields, respectively. The C
NMR spectra for each epimers showed a sin-
gle signal for C-1[l C-1(R) 101.59 ppm or l
C-1(S) 101.92 ppm], C-1% [l C-1%(R) 71.46 ppm
or l C-1%(S) 72.78 ppm] and C-2% [l C-2% (R)
61.34 ppm or l C-2%(S) 62.00 ppm]. 2,3,4,
6-Tetra-O-acetyl-a-
D
-galactopyranosyl
tri-
chloroacetimidate (8)18,19 was previously tried
as the glycosylation donor with the alcohol
5S, but a lower yield was obtained (38%). For
identification purposes, glycosides 10R and
10S were also prepared by de-O-benzoylation
and re-O-acetylation of 9R and 9S. Transfor-
mation of the azidoderivatives 9R and 9S into
the N-acyl derivatives 11R and 11S was ef-
fected via a modification of the Staudinger
reaction involving hexadecanoyl chloride and
triphenylphosphine in dry benzene.20 Deriva-
tives 11R and 11S were then de-O-acylated by
the Zemple´n procedure to give products 12R
and 12S, respectively, in high yields.
2-Acetamido-2-deoxy-
D
-glucopyranosides
were obtained with 1,3,4,6-tetra-O-acetyl-2-al-
lyloxycarbonylamino-2-deoxy-b- -glucopyra-
D
Scheme 2.
nose (13)21,22 as the donor, the silylated mix-
ture 3R/4S or 3S/4R as the acceptor and
trimethylsilyl trifluoromethanesulfonate as the
promotor (Scheme 3). The b-glycosides 14R or
14S were obtained in a pure form in 78 or
79% yields, respectively (after removal of the
minor glycosylation product of 4S or 4R by
whose desilylation afforded 5S and 6R with
similar yields. The high regio- and stereoselec-
tivity of the introduction of the azido group at
the secondary position has also been reported
recently by Bittman and co-workers13 on 1,2-
and 1,3-diols under similar conditions (Mit-
sunobu reaction). Nevertheless, it should be
noted that the above regioselectivity was not
observed previously in the case of partly pro-
tected carbohydrates having a primary and a
secondary free hydroxyl group.14
1
column chromatography). The H and 13C
NMR spectra of the two glycosides confirmed
the presence of one epimer only. Two different
routes were tested for the transformation of
14R and 14S to 17R and 17S, respectively.
The glycoside 14R was first subjected to N-
deallyloxycarbonylation followed by N-acety-
lation to afford product 15R in 73% overall
yield. The azido function of the latter was
reduced by hydrogen transfer and N-acylated
with hexadecanoyl chloride in pyridine to give
the peracetylated glycosyl ceramide analogue
17R in 69% overall yield. On the other hand,
the glycoside 14S was transformed into the
N-hexadecanoyl derivative 16S (55% yield) via
the modified Staudinger reaction already men-
tioned.20 N-Deallyloxycarbonylation and re-
N-acetylation gave the product 17S in 75%
yield. Derivatives 17R and 17S were then O-
Galactosylation was carried out with the
azidoalcohol acceptors 5R and 5S and 2,3,4,
6-tetra-O-benzoyl-a-
D
-galactopyranosyl
tri-
chloroacetimidate (7) as the donor (Scheme 2).
Preparation of 7 was achieved in three steps
starting from penta-O-benzoyl-a- -galactopy-
D
ranose.15 1-O-Debenzoylation with hydrazine
acetate was followed by reaction with tri-
chloroacetonitrile and DBU in CH2Cl2 to af-
ford the known imidate 7.16,17 Glycosylation
of the acceptors 5R and 5S with the donor 7
in the presence of a catalytic amount of
trimethylsilyl trifluoromethanesulfonate at

184
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
Interfacial beha6iour.—The four neoglyco-
syl ceramides (12R, 12S, 18R and 18S) were
spread at the air ꢀ water interface from 2:1
chloroform–methanol solutions. The p-A
isotherms (Fig. 1) did not exhibit any two-di-
mensional phase transition and are character-
istic of monolayers in a liquid-condensed
phase with a high chain order. The isotherm
shapes at 20 °C are comparable with that pre-
viously reported for natural cerebrosides,23
glucosyl cerebrosides24,25 and galactosyl
ceramides26–29 at temperatures ranging from
20 to 24 °C. The characteristic magnitudes
reflecting the monolayer properties, are given
in Table 1. Whatever the glycolipid, the limit-
ing molecular area extrapolated at zero sur-
2
,
face pressure is close to 40 A /molecule
representative of the cross-sectional area of
two hydrocarbon chains in a tightly packed
arrangement. The polar groups orientated
themselves in such a way that they do not
significantly increase the cross-sectional area
over that of the hydrocarbon chains. This
apparently low influence of the polar group is
in accordance with the limiting molecular area
2
,
(41.5 A /molecule), determined by X-ray dif-
fraction at grazing angles incidence, recently
reported for a
D
-erythro C18 ceramide.30 The
Scheme 3.
value of the minimal two-dimensional com-
pressibility31 (Cs, Table 1), determined at the
highest-surface pressure just before collapse,
confirmed the liquid-condensed behavior of
the neoglycosyl ceramides at the air ꢀ water
deacylated by the Zemple´n procedure to give
products 18R and 18S, respectively, in high
yields.
Fig. 1. Pressure–area (p-A) isotherms for the analogues of glycosyl ceramides at 20 °C.

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
185
Table 1
Characteristic magnitudes of the glycosyl-ceramide analogue monolayers
Glycosylceramide
analogue
Limiting molecular area a A0
Apparent collapse pressure yc
(mN/m)
Minimal compressibility Cs (×10−2
(m/mN)
)
2
,
(A /molecule)
12R
12S
18R
18S
41.90
43.30
43.40
41.90
ꢀ41
ꢀ38
ꢀ47
ꢀ40
0.41
0.53
0.46
0.30
a Extrapolated area at zero surface pressure.
on the other hand behave similarly—until 43
interface; this could be related to the high
transition temperature of the gel to liquid
crystalline phase already reported for satu-
rated galactosyl cerebrosides.32 The values of
the apparent collapse-surface pressure, deter-
mined for the neoglycosyl ceramide monolay-
ers, are lower than those reported for natural
galactosyl ceramides.26–29 However, it could
be noted that the galactocerebroside iso-
therms33 and the collapse surface pressures for
monolayers of natural galactosyl ceramides26
are highly dependent on the compression rate.
The common feature with natural glycosyl
ceramides is the poorly defined collapse
pressure26 and the possibility to compress the
film to very low molecular areas (below 40
A /molecule). The increase of pressure at
areas too small to be compatible with that of
a single monolayer argues against a true col-
lapse and about a loss of order of the film. In
a detailed study performed with natural galac-
tocerebrosides of similar behavior, Johnston
and Chapman34 suggested the formation of a
bilayer at smallest areas. Like these authors,
we observed that at very small areas, the
molecules formed a film so rigid that it was
able to physically move the Wilhelmy plate
and eject it from the interface. It is notewor-
thy that a recent AFM study on galactocere-
brosides reported the formation of nanotubes
at the air ꢀ water interface from the beginning
of the compression.33
2
2
,
,
A /molecule for 12S and 18R and 41 A /
molecule for 12R and 18S—could be related
to the limiting molecular areas of the neogly-
cosyl ceramides extrapolated at zero surface
pressure. A0 are identical for 12R and 18S (42
2
2
,
,
A /molecule) and 12S and 18R (43.3 A /
molecule), respectively (Table 1). This value,
corresponding to the smallest area that a
molecule can occupy at the air ꢀ water inter-
face, is correlated to the molecular cross-sec-
tion area (which can be determined by X-ray
scattering). An over-simple explanation could
be a compensation of the higher cross section
of the 2-acetamido-2-deoxy-
group over that of -galactose (displayed in
D
-glucose head
D
2
34
,
the second part of the isotherm) by the higher
cross section of the (R)-lipid moiety over that
of the (S)-lipid (displayed in the third part of
the isotherm). The difference in the packing
effect of the headgroups seems to be partially
compensated by the chiral position of the
N-acyl chain. Therefore, it appears that in the
monolayer weakly compressed, the spatial
configuration of the whole molecule controls
the self-ability of the neoglycosyl ceramides to
pack closely. In the second part (liquid con-
2
,
densed phase, from 41 A /molecule to the
beginning of the collapse), the isotherms of
12R and 12S on one hand and of 18R and 18S
on the other hand, are close together, each
2
,
group being separated by 2 A /molecule. The
A detailed analysis of the isotherms of the
neoglycosyl ceramides (12R, 12S, 18R and
18S) allows to distinguish three regions in the
diagram which could be tentatively explained
taking into account the ‘chiral’ structure of
each glycolipid molecules. The first part (less
condensed monolayer), in which compounds
12R and 18S on one hand, and 12S and 18R
differences can be attributed to the contribu-
tion of the polar group. The packing with a
2-acetamido-2-deoxy-
D
-glucose
headgroup
(18R and 18S) is less condensed than that with
a galactose one (12R and 12S). The smaller,
less hydrated headgroup of galactosyl ce-
ramide would allow a closer intermolecular
approach than is possible with the larger,

186
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
more hydrated glucosamine headgroup.28 In
chirality of the lipid moiety and this investiga-
tion seems of interest in the study of biological
membranes.
the third part, occurring for molecular areas
2
,
lower than 37 A /molecule (apparent collapse
phase), the isotherms of 12R and 18R on one
hand and of 12S and 18S on the other hand,
join at the same surface pressure for molecular
2
4. Experimental
,
areas lower than 20 A /molecule. This could
be interpreted as interdigitation of the alkyl
chains solely dependent on their chirality and
a possible change in molecular arrangements
independent on the nature of the polar group.
The isotherms of 12R and 18R and of 12S and
18S join respectively at the same surface pres-
sure value which is higher for the (R)-configu-
ration than for the (S)-one. It is noteworthy
that for a same N-acyl chain position, the
apparent collapse surface pressure is lower
with a galactose headgroup than with a 2-
General methods.—Pyridine was dried by
boiling with CaH2 prior to distillation.
Dichloromethane was washed twice with wa-
ter, dried with CaCl2 and distilled from CaH2.
Methanol was distilled from magnesium. Tet-
rahydrofurane was distilled from Na–ben-
zophenone. Pyridine, THF and CH2Cl2 were
,
stored over 4 A molecular sieves and MeOH
,
over 3 A molecular sieves. Melting points
were determined on a Bu¨chi apparatus and
were uncorrected. Thin-layer chromatography
was performed on aluminium sheets coated
with Silica Gel 60 F254 (E. Merck). Com-
pounds were visualised by spraying the TLC
plates with dilute 15% aq H2SO4, followed by
charring at 150 °C for a few min. Column
chromatography was performed on Silica-Gel
Geduran Si 60 (E. Merck). Optical rotations
were recorded on a Perkin–Elmer 241 polar-
acetamido-2-deoxy- -glucose one. This fact
D
can be related to the more condensed state
obtained with a galactose headgroup during
the monolayer compression.
3. Conclusion
1
Four different neoglycolipids have been
imeter in a 1 dm cell at 21 °C. H and 13C
synthesized with a b-
D
-galactose or a 2-acet-
NMR spectra were recorded with a Bruker
AC-200 spectrometer working at 200 and 50
MHz, respectively, with Me4Si as the internal
standard. Elemental analyses were performed
by the ‘Laboratoire Central d’Analyses du
CNRS’ (Vernaison, France).
Langmuir monolayer experiments were per-
formed using a computerized KSV 3000 Lang-
muir–Blodgett trough (Finland) enclosed in a
filtered air dry flow cabinet to avoid dust
deposition. The surface pressure was mea-
sured using a platinum Wilhelmy plate at-
tached to a sensitive balance with an accuracy
of 90.3 mN/m. The trough was filled with
Milli-Q ultrapure water (resistivity=18.2
MV/cm) produced by a four-cartridge purifi-
cation system (Millipore, France). The whole
trough was temperature-controlled with a
amido-2-deoxy-
D
-glucose head group and a
2-hexadecanoylaminohexadecyl aglycone of
(R) or (S) configuration. Their interfacial be-
havior has been studied by the Langmuir
monolayer technique. Compared with the nat-
ural glycosyl ceramides, these analogues are
also able to form condensed monolayer at the
air ꢀ water interface. The simplified structure
of the analogue derivatives (with the absence
of both the hydroxyl group at C-3 and the
unsaturation at C-4 of the sphingosine main
chain) allowed us to determine the contribu-
tion of the different parts of the glycosphin-
golipid-like molecule (carbohydrate head and
N-acyl chain) on the packing of the mono-
layer. Differences arise from the nature of the
carbohydrate and the stereochemistry of the
N-acyl linkage; a more precise study on the
influence of the chirality on the molecular
packing through Langmuir monolayer tech-
nique is now under investigation. The molecu-
lar packing of mixtures of glycolipids and
phospholipids could also be affected by the
commercially
system.
available
heating–cooling
Monolayer experiments.—The water sub-
phase was cleaned by suction with a capillary
connected to a water vacuum jet pump. The
cleanliness was checked at surface pressure

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
187
lower than 0.05 mN/m after reducing the
trough area to the minimum. The subphase
was maintained at 20 °C. The spreading solu-
tions were prepared by dissolving each glycol-
ipid into a mixture of analytical grade 2:1
CHCl3–MeOH at a precise concentration
comprised between 5×10−4and 6×10−4 M.
Each solution was stored at 4 °C. The mono-
layer was prepared by spreading an aliquot
(200 mL) of the glycolipid solution at the
air ꢀ water interface, using a 200 mL SGE mi-
crosyringe. After allowing 10 min for a com-
plete solvent evaporation, the monolayer was
symmetrically compressed by two mobile bar-
riers at the rate of 15 cm2/min. The compres-
sion was continuously achieved in a single
step. The interfacial parameters, collapse sur-
face pressure (yc), two-dimensional compress-
ibility of the monolayer (Cs) and limiting
molecular area extrapolated at zero surface
pressure (A0) were determined from the
isotherm. The accuracy of the molecular area
ether). Further elution with 2:1 EtOAc–
petroleum ether as the eluent afforded the
expected derivative 2S which was crystallised
from petroleum ether (0.400 g, 37% yield): mp
83 °C; [h]D −8.9° (c 1.3, EtOH), lit.9 [h]D
−9.0° (c 0.9, EtOH); Rf 0.48 (2:1 EtOAc–
1
13
petroleum ether); H and C NMR data were
in accordance with the literature.9
(2R)-1,2-Hexadecanediol
(2R).—Com-
pound 2R was obtained in 37% yield from
racemic 1,2-epoxyhexadecane 1 and the (S,S)-
(salen)Co(III) complex as described for 2S.
Compound 2R was also prepared in 78% yield
from the (2R)-1,2-epoxyhexadecane (1R) as
previously described, using a slight excess of
water (1.1 equiv.): mp 83 °C, lit.35 mp 84–
84.4 °C; [h]D +9.2° (c 1.0, EtOH).
(2R)-2-Azido-1-trimethylsilyloxy hexade-
cane (3R) and (2S)-1-azido-2-trimethylsilyloxy
hexadecane (4S).—Diisopropylazodicarboxy-
late (0.720 mL, 3.73 mmol) and PPh3 (0.929 g,
3.54 mmol) were successively added under an
argon atmosphere to a suspension of (2S)-1,2-
hexadecanediol (2S) (0.832 g, 3.22 mmol) in
dry THF (7 mL). The mixture became homo-
geneous after a few seconds and was stirred
for 1 h before cooling to 0 °C. After addition
of Me3SiN3 (1.06 mL, 8.06 mmol), the mixture
was allowed to reach rt and stirred overnight.
Excess of Me3SiN3 was destroyed by stirring
with cold satd aq NaHCO3 (30 mL) and the
product was extracted with CH2Cl2 (2×30
mL). The organic phase was dried (Na2SO4)
and concentrated to afford the crude product
which was purified by column chromatogra-
phy (1:25 EtOAc–petroleum ether). Product
3R and a small amount of (2S)-1-azido-2-
trimethylsilyloxy hexadecane (4S) (6% of the
total amount) were recovered as an oil: (0.882
g, 89% yield). Compounds 3R and 4S could
measurements was calculated to be lower than
2
,
90.8 A /molecule on the whole range of pres-
sure. Cs was directly calculated from the p-A
isotherm data by the KSV’s software accord-
ing to the following equation:
Cs= −1/Ay(#Ay/#y)T
where Ay is the area per molecule at the
indicated surface pressure and y is the corre-
sponding surface pressure.
(2R)-1,2-Epoxyhexadecane (1R) and (2S)-
1,2-hexadecanediol (2S).—To a solution of
(R,R)-(−)-N,N%-bis(3,5-di-tert-butylsalicyli-
dene)-1,2-cyclohexanediaminocobalt(III) ace-
tate prepared from the commercially available
precursor (R,R)-(salen)Co(II) complex (25.3
mg, 0.042 mmol) according to Jacobsen et
al.,7,8 and racemic 1,2-epoxyhexadecane 1 (1.0
g, 4.16 mmol) in dry THF (1 mL) was added
at 0 °C over 1 h a mixture of THF (0.5 mL)
and water (41.5 mL, 2.30 mmol, 0.55 equiv).
The mixture was stirred for 6 h at 0 °C, then
allowed to reach rt and stirring was main-
tained overnight. After concentration, the
residue was purified by column chromatogra-
phy. Elution with 1:5 EtOAc–petroleum ether
afforded 1R as an oil (0.480 g, 48% yield): [h]D
+4.7° (c 2.0, CHCl3), lit.10 [h]D +4.84° (c 2.8,
CHCl3); Rf 0.75 (1:10 EtOAc–petroleum
1
not be separated, they were identified by H
and 13C NMR and directly used in further
reactions.
Product 3R: Rf 0.90 (1:25 EtOAc–
1
petroleum ether); H NMR (CDCl3): l 3.70
(dd, 1 H, J1a,2 4.0, J1a,1b 10.5 Hz, H-1a), 3.57
(dd, 1 H, J1b,2 6.9 Hz, H-1b), 3.36 (m, 1 H,
H-2), 1.40–1.26 (m, 26 H, 13 CH2 alkyl
chain), (0.88, t, 3 H, J 6.6 Hz, CH3 alkyl
chain); 13C NMR (CDCl3): l 65.62 (C-1),
63.57 (C-2), 31.80, 30.27, 29.53–29.21, 26.15,

188
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
6R: white solid, mp 44 °C, lit.13 mp 45.8-
46.1 °C; [h]D -3.4° (c 1.0, CHCl3), lit.13 [h]D
−3.3° (c 1.2, CHCl3).
22.73 (13 C, CH2 alkyl chain), 14.12 (CH3
alkyl chain).
Product 4S: 13C NMR (CDCl3): l 71.95
(C-2), 56.63 (C-1).
2,3,4,6-Tetra-O-benzoyl-h- -galactopyrano-
D
osyl trichloroacetimidate (7).—Hydrazine ace-
(2S)-2-Azido-1-trimethylsilyloxy
hexade-
tate (0.945 g, 10.26 mmol) was added to a
cane (3S) and (2R)-1-azido-2-trimethylsilyloxy
hexadecane (4R).—Obtained as a mixture as
described above from (2R)-1,2-hexadecanediol
2R in 88% yield.
solution of 1,2,3,4,6-penta-O-benzoyl-a- -
D
galactopyranose15 (6.00 g, 8.56 mmol) in dry
DMF (50 mL) and the mixture was stirred for
16 h before a new addition of hydrazine ace-
tate (0.630 g, 6.86 mmol) and stirring was
maintained for 24 h. The mixture was poured
into EtOAc (300 mL), washed with brine (50
mL), then with water (50 mL) and dried
(Na2SO4). After concentration, the crude
product was purified by column chromatogra-
phy (2:3 EtOAc–petroleum ether) to afford
the 1-O-debenzoylated derivative as an oily
mixture of anomers (4.10 g, 80% yield). Rf
(2R)-2-Azidohexadecan-1-ol (5R) and (2S)-
1-azidohexadecan-2-ol (6S).—Potassium car-
bonate (0.500 g, 3.62 mmol) was added to a
mixture of 3R/4S (0.850 g, 2.39 mmol) solu-
bilised in MeOH (30 mL) and the suspension
was stirred overnight. After concentration, the
residue was partitioned between CHCl3 (50
mL) and water (50 mL). The aqueous phase
was extracted twice with CHCl3 (2×30 mL)
and the combined organic phases were dried
(Na2SO4) and concentrated; the residue was
purified by column chromatography (1:4
EtOAc–petroleum ether) affording first the
pure product 6S (0.034 g, 5%), then the de-
sired compound 5R (0.583 g, 86%).
1
0.65–0.58; H NMR (CDCl3+water): l 5.88
(d, 0.73 H, J1,2 3.5 Hz, H-1a), 5.09 (d, 0.27 H,
13
J1,2 7.4 Hz, H-1b); C NMR (CDCl3): l 96.32
(C-1b), 91.18 (C-1a).
Trichloroacetonitrile (4.3 mL, 42.9 mmol)
and DBU (1.0 mL, 6.69 mmol) were succes-
sively added to a cooled solution (0 °C) of the
above derivative (4.10 g) in dry CH2Cl2 (24
mL). The mixture was allowed to reach rt and
stirred for 4 h. After concentration, the
residue was purified by column chromatogra-
phy (2:5 EtOAc–petroleum ether containing
0.5% NEt3) to afford the pure product 7
which was obtained as an amorphous solid
(4.07 g, 80% yield). [h]D +138° (c 1.0,
CHCl3), lit.16 [h]D +113° (c 1.0, CHCl3), lit.17
[h]D +141° (c 1.0, CHCl3); Rf 0.70 (2:5
EtOAc–petroleum ether);1H NMR (CDCl3): l
8.12–7.25 (m, 20 H, 4 C6H5), 6.61 (d, 1 H, J1,2
3.0 Hz, H-1), 5.57 (bd, 1 H, J3,4 2.8, J4,5 0.5
Hz, H-4), 5.44 (dd, 1 H, J2,3 10.8 Hz, H-3),
5.37 (dd, 1 H, H-2), 4.45 (bdd, 1 H, J5,6a 6.4,
J5,6b 6.8 Hz, H-5), 4.18 (dd, 1 H, J6a,6b 11.3
Hz, H-6a), 4.09 (dd, 1 H, H-6b).
5R: white solid, mp 46 °C; [h]D −9.5° (c
1
1.0, CHCl3); Rf 0.72; H NMR (CDCl3+
D2O): l 3.71 (dd, 1 H, J1a,2 7.2, J1a,1b 11.5 Hz,
H-1a), 3.59 (dd, 1 H, J1b,2 3.5 Hz, H-1b), 3.45
(m, 1 H, H-2), 1.55–1.26 (m, 26 H, 13 CH2
alkyl chain), 0.88 (t, 3 H, J 6.6 Hz, CH3 alkyl
chain); 13C NMR (CDCl3): l 65.17 (C-1),
64.51 (C-2), 31.99, 30.62, 29.71-29.48, 26.06,
22.71 (13 C, CH2 alkyl chain), 14.09 (CH3
alkyl chain). Anal. Calcd for C16H33N3O
(283.45): C, 67.79; H, 11.74; N, 14.82. Found;
C, 67.66; H, 11.73; N, 14.79.
6S: white solid, mp 45 °C; [h]D +3.1° (c
1
1.0, CHCl3); Rf 0.80; H NMR (CDCl3): l
3.76 (m, 1 H, H-2), 3.39 (dd, 1 H, J1a,2 3.3,
J1a,1b 12.3 Hz, H-1a), 3.25 (dd, 1 H, J1b,2 7.3
Hz, H-1b), 1.92 (m, 1H, OH), 1.50–1.21 (m,
26 H, 13 CH2 alkyl chain), 0.88 (t, 3 H, J 6.6
Hz, CH3 alkyl chain).
(2S)-2-Azidohexadecan-1-ol (5S) and (2R)-
1-azidohexadecan-2-ol (6R).—These product
were obtained in 86 and 5%, respectively, as
described above from the mixture 3S/4R.
5S: white solid, mp 46 °C, lit.13 mp 45–
45.5 °C; [h]D +9.7° (c 1.0, CHCl3), lit.13 [h]D
+9.9° (c 1.2, CHCl3).
(2R)-2-Azidohexadecyl 2,3,4,6-tetra-O-ben-
zoyl-i-
Tetra-O-benzoyl-a-
D
-galactoyranoside
(9R).—2,3,4,6-
tri-
D
-galactopyranosyl
chloroacetimidate (7) (0.741 g, 1.00 mmol),
(2R)-2-azidohexadecanol (5R) (0.272 g, 0.96
,
mmol) and activated 4 A molecular sieves (0.5
g) were added to alcohol-free CH2Cl2 (4 mL).
The reaction mixture was flushed with argon

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
189
30.64, 29.76-29.39, 25.98, 22.77 (CH2 alkyl
chain), 14.22 (CH3 alkyl chain). Anal. Calcd
for C50H59N3O10 (861.99): C, 69.66; H, 6.90;
N, 4.87. Found; C, 69.88; H, 6.86; N, 4.90.
(2R)-2-Azidohexadecyl 2,3,4,6-tetra-O-ace-
while cooling to 0 °C. A solution of TMSOTf
(0.015 mL, 0.07 mmol) in CH2Cl2 (0.5 mL)
was introduced through a syringe over a 1 h
period. The mixture was stirred overnight at
0 °C before filtration over Celite and dilution
with CH2Cl2 (50 mL). The organic phase was
washed with satd aq NaHCO3 and dried
(Na2SO4). Concentration of the solution gave
a residue which was purified by column chro-
matography (1:3 EtOAc–petroleum ether) to
afford pure derivative 9R as an oil (0.753 g,
91% yield): [h]D +72.0° (c 1.0, CHCl3); Rf
tyl-i- -galactoyranoside (10R).—Compound
D
10R was prepared by O-debenzoylation of 9R
(0.259 g, 0.30 mmol) in MeOH containing a
catalytic amount of sodium. After 16 h, the
solution was concentrated and the residue was
acetylated overnight in a 2:1 pyridine–Ac2O
mixture (10 mL). After concentration and
purification by column chromatography (elu-
ent 2:1 EtOAc–petroleum ether), the product
10R (164 mg, 91% yield) was obtained as an
oil: [h]D −7.6° (c 1.0, CHCl3); Rf 0.42 (5:2
1
0.77; H NMR (CDCl3): l 8.13–7.21 (m, 20
H, 4 C6H5), 6.02 (dd, 1 H, J3,4 3.4, J4,5 0.5 Hz,
H-4), 5.83 (dd, 1 H, J1,2 7.8, J2,3 10.4 Hz, H-2),
5.63 (dd, 1 H, H-3), 4.91 (d, 1 H, H-1), 4.72
(dd, 1 H, J5,6a 6.1 Hz, J6a,6b 10.6 Hz, H-6a),
4.45 (dd, 1 H, J5,6b 6.5 Hz, H-6b), 4.36 (bdd, 1
H, H-5), 4.02 (dd, 1 H, J1%a,2% 5.9, J1%a,1%b 10.3
Hz, H-1%a), 3.66 (dd, 1 H, J1%b,2% 5.0 Hz, H-1%b),
3.48 (m, 1 H, H-2%), 1.50–1.20 (m, 26 H, 13
CH2 alkyl chain), 0.89 (t, 3 H, J 6.3 Hz, CH3
alkyl chain). 13C NMR (CDCl3): l 166.05,
165.65, 165.62, 165.30 (4 C, 4 C6H5CO),
101.59 (C-1), 71.85, 71.54 (2 C, C-3,5), 71.46
(C-1%), 69.86 (C-2), 68.31 (C-4), 62.17 (C-6),
61.34 (C-2%), 32.02, 30.75, 30.64, 29.77–29.36,
25.87, 22.79 (CH2 alkyl chain), 14.24 (CH3
alkyl chain). Anal. Calcd for C50H59N3O10
(861.99): C, 69.66; H, 6.90; N, 4.87. Found; C,
69.50; H, 7.17; N, 5.23.
1
EtOAc–petroleum ether); H NMR (CDCl3):
l 5.40 (dd, 1 H, J3,4 3.3, J4,5 0.5 Hz, H-4), 5.25
(dd, 1 H, J1,2 7.9, J2,3 10.4 Hz, H-2), 5.03 (dd,
1 H, H-3), 4.54 (d, 1 H, H-1), 4.21 (dd, 1 H,
J5,6a 6.9, J6a,6b 11.2 Hz, H-6a), 4.13 (dd, 1 H,
J5,6b 6.8 Hz, H-6b), 3.92 (bdd, 1 H, H-5), 3.91
(dd, 1 H, J1%a,2% 5.6, J1%a,1%b 10.4 Hz, H-1%a), 3.60
(dd, 1 H, J1%b,2% 6.4 Hz, H-1%b), 3.41 (m, 1 H,
H-2%), 2.17, 2.07, 2.06, 1.99 (4s, 12 H, 4
CH3COO), 1.52–1.26 (m, 26 H, 13 CH2 alkyl
chain), 0.88 (t, 3 H, J 6.3 Hz, CH3 alkyl
chain). 13C NMR (CDCl3): l 170.33, 170.22,
170.09, 169.34 (4 C, 4 CH3COO), 101.10 (C-
1), 71.25 (C-1%), 70.91 (C-3), 70.81 (C-5), 68.59
(C-2), 67.04 (C-4), 61.24 (C-6), 61.20 (C-2%),
31.91, 30.50, 29.64-29.35, 25.96, 22.68 (CH2
alkyl chain), 20.74, 20.63, 20.63, 20.54 (4 C, 4
CH3COO), 14.10 (CH3 alkyl chain). Anal.
Calcd for C30H51N3O10 (613.73): C, 58.71; H,
8.38; N, 6.85. Found; C, 58.41; H, 8.58; N,
6.74.
(2S)-2-Azidohexadecyl 2,3,4,6-tetra-O-ben-
zoyl-i- -galactoyranoside (9S).—Obtained as
D
described above from trichoroacetimidate 7
(0.741 g, 1.00 mmol) and (2S)-2-azidohexade-
canol (5S) (0.272 g, 0.96 mmol). Compound
9S was obtained as an oil (0.761 g, 92% yield):
1
[h]D +57.5° (c 1.0, CHCl3); Rf 0.77; H NMR
(2S)-2-Azidohexadecyl
acetyl-i- -galactoyranoside (10S).—2,3,4,6-
Tetra-O-acetyl-a-
2,3,4,6-tetra-O-
(CDCl3): l 8.12–7.22 (m, 20 H, 4 C6H5), 6.01
(dd, 1 H, J3,4 3.4, J4,5 0.5 Hz, H-4), 5.85 (dd, 1
H, J1,2 7.9, J2,3 10.5 Hz, H-2), 5.63 (dd, 1 H,
H-3), 4.90 (d, 1 H, H-1), 4.72 (dd, 1 H, J5,6a
5.8, J6a,6b 10.5 Hz, H-6a), 4.44 (dd, 1 H, J5,6b
6.5 Hz, H-6b), 4.37 (ddd, 1 H, H-5), 4.05 (m,
1 H, H-1%a), 3.65-3.55 (m, 2 H, H-1%b,2%),
1.39–1.27 (m, 26 H, 13 CH2 alkyl chain), 0.88
D
D
-galactopyranosyl trichoro-
acetimidate (8)18,19 (0.616 g, 1.25 mmol) and
(2S)-2-azidohexadecanol (5S) (0.283 g, 1.00
mmol) were treated as described for com-
pounds 9R and 9S. After purification by
column chromatography (2:1 EtOAc–
1
petroleum ether), H NMR of the product
13
(t, 3 H, J 6.3 Hz, CH3 alkyl chain). C NMR
showed the presence of trichloroacetamide
which was eliminated after a new chromatog-
raphy using 20:1 CHCl3–MeOH as the eluent.
Product 10S was obtained as an oil (0.233 g,
38% yield): [h]D −18.6° (c 1.0, CHCl3); Rf
(CDCl3): l 166.08, 165.64, 165.64, 165.30 (4
C, 4 C6H5CO), 101.92 (C-1), 72.78 (C-1%),
71.79, 71.49 (2 C, C-3,5), 69.73 (C-2), 68.16
(C-4), 62.09 (C-6), 62.00 (C-2%), 31.99, 30.91,

190
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
1
CH2 alkyl chains), 0.89 (t, 6 H, J 6.3 Hz, 2
CH3 alkyl chains). 13C NMR (CDCl3): l
172.56 (NHCOO), 165.98, 165.55, 165.50,
165.41 (4 C, 4 C6H5CO), 133.62, 133.55–
133.32, 130.04–128.32 (24 C, 4 C6H5), 101.81
(C-1), 71.56 (3 C, C-1%,3,5), 70.25 (C-2), 68.26
(C-4), 62.14 (C-6), 48.49 (C-2%), 36.52, 31.97,
31.78, 29.76-29.25, 26.06, 25.67, 22.74 (CH2
alkyl chains), 14.17 (CH3 alkyl chains). Anal.
Calcd for C66H91NO11 (1074.40): C, 73.78; H,
8.54; N, 1.30. Found; C, 73.87; H, 8.63 N,
1.44.
0.42 (5:2 EtOAc–petroleum ether); H NMR
(CDCl3): l 5.41 (dd, 1 H, J3,4 3.3, J4,5 0.5 Hz,
H-4), 5.26 (dd, 1 H, J1,2 7.9, J2,3 10.4 Hz, H-2),
5.03 (dd, 1 H, H-3), 4.56 (d, 1 H, H-1), 4.21
(dd, 1 H, J5,6a 6.8 Hz, J6a,6b 11.3 Hz, H-6a),
4.13 (dd, 1 H, J5,6b 6.4 Hz, H-6b), 3.99 (dd, 1
H, J1%a,2% 2.4, J1%a,1%b 9.5 Hz, H-1%a), 3.92 (dd, 1
H, H-5), 3.54 (m, 1 H, H-2%), 3.44 (dd, 1 H,
J1%b,2% 8.8 Hz, H-1%b), 2.17, 2.07, 2.06, 1.99 (4s,
12 H, 4 CH3COO), 1.39–1.26 (m, 26 H, 13
CH2 alkyl chain), 0.88 (t, 3 H, J 6.5 Hz, CH3
alkyl chain). 13C NMR (CDCl3): l 170.37,
170.27, 170.16, 169.48 (4 C, 4 CH3COO),
101.29 (C-1), 72.68 (C-1%), 70.95 (C-3), 70.65
(C-5), 68.65 (C-2), 67.08 (C-4), 62.08 (C-2%),
61.25 (C-6), 31.89, 30.86, 29.62-29.36, 25.88,
22.67 (CH2 alkyl chain), 20.75, 20.63, 20.63,
20.54 (4 C, 4 CH3COO), 14.22 (CH3 alkyl
chain). Anal. Calcd for C30H51N3O10 (613.73):
C, 58.71; H, 8.38; N, 6.85. Found; C, 58.67;
H, 8.37; N, 6.68.
(2S)-2-Hexadecanoylaminohexadecyl 2,3,4,
6-tetra-O-benzoyl-i- -galactoyranoside (11S).
D
—Obtained from 9S in 55% yield as described
for 11R. Compound 11S was obtained as an
amorphous solid: [h]D +36.7° (c 1.0, CHCl3);
Rf 0.27 (1:3 EtOAc–petroleum ether); 1H
NMR (CDCl3): l 8.12–7.25 (m, 20 H, 4
C6H5), 6.02 (dd, 1 H, J3,4 3.3, J4,5 0.5 Hz, H-4),
5.81 (dd, 1 H, J1,2 7.9, J2,3 10.5 Hz, H-2), 5.74
(d, 1 H, J2%,NH 9.0 Hz, NH), 5.66 (dd, 1 H,
H-3), 4.82 (d, 1 H, H-1), 4.67 (dd, 1 H, J5,6a
6.4, J6a,6b 11.0 Hz, H-6a), 4.46 (dd, 1 H, J5,6b
6.4 Hz, H-6b), 4.34 (bdd, 1 H, H-5), 4.01-3.97
(m, 2 H, H-1%a,2%), 3.62 (bd, 1 H, H-1%b), 2.16
(m, 2 H, CH2CO), 1.60–1.26 (m, 52 H, 26
CH2 alkyl chains), 0.89 (t, 6 H, J 6.3 Hz, 2
CH3 alkyl chains). 13C NMR (CDCl3): l
172.74 (NHCOO), 165.71, 165.55, 165.43,
165.37 (4 C, 4 C6H5CO), 133.57, 133.25,
129.97–128.27 (24 C, 4 C6H5), 102.06 (C-1),
71.92 (C-1%), 71.53 (2 C, C-3,5), 70.05 (C-2),
68.37 (C-4), 62.10 (C-6), 48.62 (C-2%), 36.79,
31.95, 31.57, 29.74-29.40, 26.17, 25.82, 22.72
(CH2 alkyl chains), 14.15 (CH3 alkyl chains).
Anal. Calcd for C66H91NO11 (1074.40): C,
73.78; H, 8.54; N, 1.30. Found; C, 73.38; H,
8.51; N, 1.43.
Compound 10S was also prepared from 9S
in 92% yield, following the same procedure as
that used for 10R from 9R.
(2R)-2-Hexadecanoylaminohexadecyl 2,3,4,
6-tetra-O-benzoyl-i- -galactoyranoside (11R).
D
—A solution of PPh3 (0.262 g, 1.00 mmol) in
deoxygenated C6H6 (1 mL) was added drop-
wise over 1 h under an argon atmosphere to a
mixture of glycoside 9R (0.560 g, 0.65 mmol),
hexadecanoyl chloride (0.330 mL, 1.09 mmol)
,
and crushed 4 A molecular sieves (0.5 g) in
dry C6H6. The mixture was stirred for 24 h,
and after filtration over Celite and concentra-
tion of the solution, the product was purified
by two successive column chromatographs
first with 1:3 EtOAc–petroleum ether as the
eluent, then with 24:1 CHCl3–EtOH. Com-
pound 11R was obtained as an oil (0.391 g,
56% yield): [h]D +63.8° (c 1.0, CHCl3); Rf
(2R)-2-Hexadecanoylaminohexadecyl i- -
D
1
0.27 (1:3 EtOAc–petroleum ether); H NMR
galactopyranoside (12R).—The benzoylated
product 11R (0.600 g, 0.558 mmol) was added
to a mixture of MeOH (20 mL) containing a
catalytic amount of MeONa and CH2Cl2 (5
mL). Dissolution occurred after a few min-
utes, then the product started to crystallise.
Stirring was maintained for 16 h and CH2Cl2
was vaporised. The product was recovered by
filtration and washed twice with MeOH (2×5
mL) (0.317 g, 87% yield): mp 143 °C (MeOH).
(CDCl3): l 8.12–7.25 (m, 20 H, 4 C6H5), 6.01
(dd, 1 H, J3,4 3.3, J4,5 0.5 Hz, H-4), 5.78 (dd, 1
H, J1,2 7.5, J2,3 10.4 Hz, H-2), 5.66 (dd, 1 H,
H-3), 5.34 (d, 1 H, J2%,NH 8.8 Hz, NH), 4.79 (d,
1 H, H-1), 4.68 (dd, 1 H, J5,6a 6.5, J6a,6b 10.9
Hz, H-6a), 4.43 (dd, 1 H, J5,6b 6.4 Hz, H-6b),
4.34 (bdd, 1 H, H-5), 4.04 (m, 1 H, H-2%), 3.95
(dd, 1 H, J1%a,2% 1.5 Hz, J1%a,1%b 9.2 Hz, H-1%a),
3.62 (dd, 1 H, J1%b,2% 3.2 Hz, H-1%b), 1.71 (t, 2
H, J 7.5 Hz, CH2CO), 1.58–1.26 (m, 52 H, 26
1
[h]D −6.0° (c 1.0, 1:1 CHCl3–MeOH); H

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
191
12.2 Hz, H-6a), 4.13 (dd, 1 H, J5,6b 2.5 Hz,
H-6b), 3.89 (dd, 1 H, J1%a,2% 6.3, J1%a,1%b 10.4 Hz,
H-1%a), 3.85 (ddd, 1 H, H-5), 3.69 (ddd, 1 H,
H-2), 3.68 (dd, 1 H, J1%b,2% 4.3 Hz, H-1%b), 3.62
(m, 1 H, H-2%), 2.03, 1.99, 1.95 (3 s, 9 H, 3
CH3COO), 1.59–1.29 (m, 26 H, 13 CH2 alkyl
chain), 0.88 (t, 3 H, J 6.7 Hz, CH3 alkyl
chain). 13C NMR (CDCl3): l 170.67, 170.59,
169.45 (3 C, CH3COO), 155.67 (NHCOO),
132.68 (CHꢀ), 117.57 (CH2ꢀ), 100.83 (C-1),
72.13 (C-3), 71.82 (C-5), 71.49 (C-1%), 68.85
(C-4), 65.80 (allyl CH2), 62.13 (C-6), 61.45
(C-2%), 55.99 (C-2), 31.89, 30.59, 29.67-29.34,
25.99, 22.67 (CH2 alkyl chain), 20.72, 20.66,
20.61 (3 C, 3 CH3COO), 14.11 (CH3 alkyl
chain). Anal. Calcd for C32H54N4O10 (654.78):
C, 58.69; H, 8.31; N, 8.56. Found; C, 58.54;
H, 8.27; N, 8.30.
NMR (C5D5N): l 8.23 (d, 1 H, J2%,NH 7.9 Hz,
NH), 7.21, 6.82, 6.55 and 6.44 (4 m, 4 H, 4
OH), 4.87 (d, 1 H, J1,2 7.7 Hz, H-1), 4.58–3.86
(m, 9 H, H-2,3,4,5,6a,6b,1%a,1%b,2%), 2.41 (t, 2
H, J 7.0 Hz, COCH2), 1.79, 1.60–1.10 (m, 52
H, 26 CH2 alkyl chains), 0.87 (m, 6 H, 2 CH3
alkyl chains). Anal. Calcd for C38H75NO7
(657.99): C, 69.36; H, 11.49; N, 2.13. Found;
C, 69.13; H, 11.80; N, 2.12.
(2S)-2-Hexadecanoylaminohexadecyl i- -
D
galactopyranoside (12S).—Product 12S was
prepared in 85% yield from 11S as described
above for 12R: mp 150 °C (MeOH). [h]D
+
1
15.8° (c 1.0, 1:1 CHCl3–MeOH); H NMR
(C5D5N): l 8.25 (d, 1 H, J2%,NH 8.5 Hz, NH),
7.12, 6.82, 6.59 and 6.43 (4 m, 4 H, 4 OH),
4.82 (d, 1 H, J1,2 7.6 Hz, H-1), 4.54–4.08 (m,
9 H, H-2,3,4,5,6a,6b,1%a,1%b,2%), 2.45 (t, 2 H, J
7.0 Hz, COCH2), 1.79, 1.60–1.10 (m, 52 H, 26
CH2 alkyl chains), 0.87 (m, 6 H, 2 CH3 alkyl
chains). Anal. Calcd for C38H75NO7·H2O
(676.00): C, 67.51; H, 11.48; N, 2.07. Found;
C, 67.59; H, 11.47; N, 2.13.
(2S)-2-Azidohexadecyl 3,4,6-tri-O-acetyl-2-
allyloxycarbonylamino-2-deoxy-i- -glucopy-
D
ranoside (14S).—Obtained as described above
from 13 (0.897 g, 2.08 mmol) and the silylated
mixture 3S/4R (0.729 g, 2.05 mmol). Com-
pound 14S (1.060 g, 79% yield) was obtained
as a white solid: mp 119–120 °C (EtOH); [h]D
(2R)-2-Azidohexadecyl 3,4,6-tri-O-acetyl-2-
allyloxycarbonylamino-2-deoxy-i-
ranoside (14R).—Tetra-O-acetyl-2-allyloxy-
carbonylamino-2-deoxy-b- -glucopyranose
D
-glucopy-
1
−1.4° (c 1.0, CHCl3); Rf 0.61; H NMR
(CD3COCD3): l 6.50 (d, 1 H, J2,NH 6.3 Hz,
NH), 5.92 (ddd, 1 H, CHꢀ), 5.26 (bd, 1 H, 0.5
CH2ꢀ), 5.22 (dd, 1 H, J2,3 10.0, J3,4 9.4 Hz,
H-3), 5.13 (dd, 1 H, 0.5 CH2ꢀ), 5.00 (dd, 1 H,
J4,5 9.9 Hz, H-4), 4.87 (d, 1 H, J1,2 8.4 Hz,
H-1), 4.60–4.40 (m, 2 H, allyl CH2), 4.26 (dd,
1 H, J5,6a 4.9, J6a,6b 12.2 Hz, H-6a), 4.13 (dd, 1
H, J5,6b 2.6 Hz, H-6b), 3.96 (d, 1 H, J1%a,1%b 6.9
Hz, H-1%a), 3.85 (ddd, 1 H, H-5), 3.69 (ddd, 1
H, H-2), 3.62 (1 H, H-2%), 3.57 (dd, 1 H, J1%b,2
7.2 Hz, H-1%b), 2.03, 1.99, 1.95 (3 s, 9 H, 3
CH3COO), 1.55–1.29 (m, 26 H, 13 CH2 alkyl
chain), 0.88 (t, 3 H, J 6.7 Hz, CH3 alkyl
chain). 13C NMR (CDCl3): l 170.62, 170.55,
169.46 (3 C, CH3COO), 155.67 (NHCOO),
132.73 (CHꢀ), 117.46 (CH2ꢀ), 100.16 (C-1),
72.76 (C-1%), 72.16 (C-3), 71.75 (C-5), 68.89
(C-4), 65.74 (allyl CH2), 62.11 (2 C, C-6,2%),
56.02 (C-2), 31.89, 30.83, 29.64-29.32, 25.91,
22.65 (CH2 alkyl chain), 20.72, 20.64, 20.59 (3
C, 3 CH3COO), 14.11 (CH3 alkyl chain).
Anal. Calcd for C32H54N4O10 (654.78): C,
58.69; H, 8.31; N, 8.56. Found; C, 59.08; H,
8.39; N, 8.14.
D
(13)21 (1.048 g, 2.43 mmol) and the silylated
mixture 3R/4S (0.850 g, 2.39 mmol) were
added to alcohol-free CH2Cl2. The reaction
mixture was flushed with argon while cooling
to −20 °C. Trimethylsilyl trifluoromethane-
sulfonate (0.470 mL, 2.43 mmol) was intro-
duced through a syringe and the argon flush
maintained for 0.5 h. The mixture was stirred
overnight at −20 °C, then poured into satd
aq NaHCO3 and extracted with CH2Cl2 (2×
50 mL). After drying (Na2SO4), concentration
of the solution gave a residue which was
purified by column chromatography (2:3
EtOAc–petroleum ether) to afford the pure
derivative 14R as a solid (1.24 g, 78% yield):
mp 81–82 °C (EtOH); [h]D +5.9° (c 1.0,
1
CHCl3); Rf 0.61; H NMR (CD3COCD3): l
6.49 (d, 1 H, J2,NH 7.8 Hz, NH), 5.92 (ddd, 1
H, CHꢀ), 5.26 (bd, 1 H, 0.5 CH2ꢀ), 5.24 (dd,
1 H, J2,3 10.0, J3,4 9.5 Hz, H-3), 5.13 (dd, 1 H,
0.5 CH2ꢀ), 5.00 (dd, 1 H, J4,5 9.9 Hz, H-4),
4.87 (d, 1 H, J1,2 8.4 Hz, H-1), 4.57–4.40 (m,
2 H, allyl CH2), 4.27 (dd, 1 H, J5,6a 4.9, J6a,6b

192
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
under argon. The mixture was stirred for 16 h,
(2R)-2-Azidohexadecyl 2-acetamido-3,4,6-
tri - O - acetyl - 2 - deoxy - i - - glucopyranoside
before a new addition of PPh3 (0.105 g, 0.40
mmol) in C6H6 (0.4 mL). Stirring was main-
tained for 24 h, then the solution was concen-
trated and the residue was purified by column
D
(15R).—A solution of palladium tetra-
kis(triphenylphosphine)—prepared under ar-
gon from tris(dibenzylideneacetone)dipal-
ladium(0) (0.015 g, 0.013 mmol) and PPh3
(0.038 g, 0.144 mmol) in dry THF (1.5 mL)—
was added to a solution of compound 14R
(0.753 g, 1.15 mmol) and dimethylmalonate
(1.1 mL) in THF (4 mL). The mixture was
stirred for 16 h, then concentrated and the
products were purified on a short column of
silica-gel (1:1 EtOAc–petroleum ether, fol-
lowed by pure EtOAc) to afford the NH2-free
derivative which was N-reacetylated in a 2:1
pyridine–Ac2O mixture (15 mL). After con-
centration, the residue was purified by column
chromatography
(1:1
EtOAc–petroleum
ether) to afford the pure product 16S (0.354 g,
51% yield): mp 139–140 °C; [h]D −21.5° (c
1.0, CHCl3); Rf 0.50 (4:5 EtOAc–petroleum
1
ether); H NMR (CDCl3): l 5.89 (ddd, 1 H,
CHꢀ), 5.78 (d, 1 H, J2,NH 8.0 Hz, NH), 5.32–
5.11 (m, 3 H, H-3, CH2ꢀ), 5.06 (dd, 1 H, J3,4
8.8, J4,5 9.9 Hz, H-4), 4.88 (d, 1 H, J1,2 8.4 Hz,
H-1), 4.55 (m, 2 H, allyl CH2), 4.48 (d, 1 H,
J2%,NH 8.2 Hz, NH), 4.28 (dd, 1 H, J5,6a 4.5,
J6a,6b 12.2 Hz, H-6a), 4.11 (dd, 1 H, J5,6b 2.0
Hz, H-6b), 4.04 (m, 1 H, H-2%), 3.87 (dd, 1 H,
J1%a,2% 3.3, J1%a,1%b 9.8 Hz, H-1%a), 3.77-3.62 (m, 2
H, H-2,5), 3.56 (dd, 1 H, J1%b,2 2.2 Hz, H-1%b),
2.16 (m, 2 H, CH2CO), 2.09, 2.05, 2.03 (3 s, 9
H, 3 CH3COO), 1.70–1.56 (m, 52 H, 26 CH2
alkyl chains), 0.88 (t, 6 H, J 6.7 Hz, 2 CH3
alkyl chains). 13C NMR (CDCl3): l 172.85
(NHCOCH2), 170.79, 170.59, 169.41 (3 C,
CH3COO), 155.88 (NHCOO), 132.59 (CHꢀ),
117.66 (CH2ꢀ), 102.06 (C-1), 72.29 (C-3),
71.83 (2 C, C-5,C-1%), 68.66 (C-4), 65.77 (allyl
CH2), 62.06 (C-6), 56.02 (C-2), 48.61 (C-2%),
36.87, 31.91–29.35, 26.17, 25.83, 22.67 (CH2
alkyl chains), 20.70, 20.66, 20.55 (3 C, 3
CH3COO), 14.09 (2 C, 2 CH3 alkyl chains).
Anal. Calcd for C48H86N2O11 (867.18): C,
66.48; H, 10.00; N, 3.23. Found; C, 66.28; H,
10.06; N, 3.28.
chromatography
(3:1
EtOAc–petroleum
ether); pure product 15R was obtained as a
white solid (0.515 g, 73% yield): mp 91–92 °C;
[h]D −6.7° (c 1.0, CHCl3); Rf 0.69 (1:3
1
petroleum ether–EtOAc); H NMR (CDCl3):
l 5.67 (d, 1 H, J2,NH 8.5 Hz, NH), 5.35 (dd, 1
H, J2,3 10.1, J3,4 9.7 Hz, H-3), 5.07 (dd, 1 H,
J4,5 9.7 Hz, H-4), 4.82 (d, 1 H, J1,2 8.3 Hz,
H-1), 4.26 (dd, 1 H, J5,6a 4.6, J6a,6b 12.3 Hz,
H-6a), 4.14 (dd, 1 H, J5,6b 2.2 Hz, H-6b), 3.88
(dd, 1 H, J1%a,2 6.0, J1%a,1%b 10.3 Hz, H-1%a), 3.81
(ddd, 1 H, H-2), 3.75 (ddd, 1 H, H-5), 3.64
(dd, 1 H, J1%b,2% 3.7 Hz, H-1%b), 3.38 (m, 1 H,
H-2%), 2.08, 2.03, 2.02, 1.94 (4 s, 12 H, 4
CH3CO), 1.50–1.25 (m, 26 H, 13 CH2 alkyl
chain), 0.88 (t, 3 H, J 6.7 Hz, CH3 alkyl
chain). 13C NMR (CDCl3): l 170.63, 170.63,
170.63, 169.37 (4 C, 3 CH3COO, CH3CON),
100.24 (C-1), 72.29 (C-3), 71.76 (C-5), 71.21
(C-1%), 68.92 (C-4), 62.17 (C-6), 61.38 (C-2%),
54.56 (C-2), 31.88, 30.49, 29.63–29.31, 26.01,
22.64 (CH2 alkyl chain), 23.20 (CH3CON),
20.68, 20.65, 20.57 (3 C, 3 CH3COO), 14.08
(CH3 alkyl chain). Anal. Calcd for
C30H52N4O9 (612.74): C, 58.80; H, 8.55; N,
9.14. Found; C, 58.94; H, 8.66; N, 9.09.
(2R)-2-Hexadecanoylaminohexadecyl 2-ace-
tamido-3,4,6-tri-O-acetyl-2-deoxy-i- -gluco-
D
pyranoside (17R).—Pd(OH)2C (0.150 g) was
added to a solution of the azido derivative
15R (0.490 g, 0.80 mmol) and freshly distilled
cyclohexene (5 mL) in abs EtOH (12 mL). The
suspension was refluxed for 4 h with stirring,
then cooled to rt and the NH2-free product
was recovered as an amorphous solid by filtra-
tion over Celite and concentration of the solu-
tion: Rf 0.50 (1:3 petroleum ether–EtOAc); 1H
NMR (CDCl3): l 6.00 (d, 1 H, J2,NH 8.5 Hz,
NH), 5.26 (dd, 1 H, J2,3 10.4, J3,4 9.5 Hz,
H-3), 5.08 (dd, 1 H, J4,5 9.8 Hz, H-4), 4.70 (d,
1 H, J1,2 8.3 Hz, H-1), 4.30–4.10 (m, 3 H,
H-6a,6b,1%a), 3.93 (ddd, 1 H, H-2), 3.70–3.61
(m, 2 H, H-5,2%), 3.57 (dd, 1 H, J1%b,2% 3.8,
(2S)-2-Hexadecanoylaminohexadecyl 3,4,6-
tri-O-acetyl-2-allyloxycarbonylamino-2-deoxy-
i- -glucopyranoside (16S).—Triphenylphos-
D
phine (0.315 g, 1.20 mmol) in deoxygenated
C6H6 (0.50 mL) was added dropwise over 1 h
to a solution of the glycoside 14S (0.524 g,
0.80 mmol) and hexadecanoyl chloride (0.390
mL, 1.29 mmol) in deoxygenated C6H6 (3 mL)

D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
193
66.95; H, 10.26; N, 3.39. Found; C, 66.88; H,
10.19; N, 3.29.
J1%a,1%b 10.2 Hz, H-1%b), 2.09, 2.04, 2.03, 1.96 (4
s, 12 H, 4 CH3CO), 1.50–1.26 (m, 26 H, 13
CH2 alkyl chain), 0.88 (t, 3 H, J 6.7 Hz, CH3
alkyl chain). 13C NMR (CDCl3): l 170.86,
170.58, 170.54, 169.37 (4 C, 3 CH3COO,
CH3CON), 100.79 (C-1), 73.81 (C-1%), 72.64
(C-3), 71.61 (C-5), 68.84 (C-4), 62.20 (C-6),
54.35 (C-2), 50.82 (C-2%), 33.44, 31.84, 29.62-
29.28, 25.99 (CH2 alkyl chain), 23.22
(CH3CON), 20.67, 20.67, 20.56 (3 C, 3
CH3COO), 14.05 (CH3 alkyl chain).
(2S)-2-Hexadecanoylaminohexadecyl 2-ace-
tamido-3,4,6-tri-O-acetyl-2-deoxy-i- -gluco-
D
pyranoside (17S).—Obtained as described
above for the preparation of 15R from 14R
starting from compound 16S (0.512 g, 0.59
mmol) and tris(dibenzylideneacetone)dipal-
ladium(0) (0.012 g, 0.0104 mmol). The crude
product was purified by column chromatogra-
phy (1:10 CHCl3–EtOAc) affording 17S as a
white solid (0.365 g, 75% yield): mp 157–
158 °C (from EtOAc–petroleum ether); [h]D
Hexadecanoyl chloride (0.380 mL, 1.25
mmol) was added dropwise at 0 °C to a solu-
tion of the NH2-free derivative in dry pyri-
dine. The mixture was allowed to reach rt and
stirring was maintained for 16 h before a new
addition of hexadecanoyl chloride (0.100 mL,
0.33 mmol). After 12 h, the reaction mixture
was cooled to 0 °C and water (0.5 mL) was
added dropwise over 1 h. The solution was
poured into ice-water and extracted with
CH2Cl2 (2×30 mL); the organic phase was
washed with 10% aq HCl, then with satd aq
NaHCO3, dried and concentrated to afford a
residue which was purified by column chro-
matography using 12:1 CHCl3–MeOH as the
eluent. Pure product 17R was recovered as a
solid (0.455 g, 69% yield): mp 168–169 °C
(EtOH); [h]D −16.0° (c 1.0, CHCl3); Rf 0.80
1
−30.5° (c 1.0, CHCl3); Rf 0.63 (EtOAc); H
NMR (CDCl3): l 5.76 and 5.68 (2 d, 2 H,
J2,NH 8.9 Hz, 2 NH), 5.16 (dd, 1 H, J2,3 8.8,
J3,4 9.5 Hz, H-3), 5.08 (dd, 1 H, J4,5 9.7 Hz,
H-4), 4.50 (d, 1 H, J1,2 8.3 Hz, H-1), 4.27 (dd,
1 H, J5,6a 4.6 Hz, J6a,6b 12.3 Hz, H-6a), 4.14
(dd, 1 H, J5,6b 2.3 Hz, H-6b), 4.07–4.00 (m, 2
H, H-2,2%), 3.88 (dd, 1 H, J1%a,2 3.5, J1%a,1%b 9.7
Hz, H-1%a), 3.67 (ddd, 1 H, H-5), 3.53 (dd, 1
H, J1%b,2% 3.7 Hz, H-1%b), 2.16 (2 H, COCH2),
2.08, 2.04, 2.03, 1.95 (4 s, 12 H, 4 CH3CO),
1.62–1.25 (m, 52 H, 26 CH2 alkyl chains),
0.88 (t, 6 H, J 6.7 Hz, 2 CH3 alkyl chains). 13C
NMR (CDCl3): l 172.96, 170.96, 170.58,
170.28, 169.36 (5 C, 3 CH3COO, CH3CON,
CH2CON), 100.66 (C-1), 72.53 (C-3), 71.82
(C-5), 71.75 (C-1%), 68.63 (C-4), 62.06 (C-6),
54.21 (C-2), 48.71 (C-2%), 36.87, 31.91, 31.56,
29.69-29.35, 26.15, 25.88, 22.66 (CH2 alkyl
chains), 23.20 (CH3CON), 20.70, 20.70, 20.58
(3 C, 3 CH3COO), 14.02 (2 C, 2 CH3 alkyl
chains). Anal. Calcd for C46H84N2O10
(825.15): C, 66.95; H, 10.26; N, 3.39. Found;
C, 66.93; H, 10.25; N, 3.46.
1
(12:1 CHCl3–MeOH); H NMR (CDCl3): l
5.93 (d, 1 H, J 8.5 Hz, NH), 5.67 (d, 1 H, J
8.9 Hz, NH), 5.16 (dd, 1 H, J2,3 9.2, J3,4 9.9
Hz, H-3), 5.08 (dd, 1 H, J4,5 9.7 Hz, H-4), 4.63
(d, 1 H, J1,2 8.3 Hz, H-1), 4.28 (dd, 1 H, J5,6a
4.7, J6a,6b 12.3 Hz, H-6a), 4.13 (dd, 1 H, J5,6b
2.2 Hz, H-6b), 4.10 (m, 1 H, H-2%), 3.96 (dd, 1
H, H-2), 3.66 (m, 3 H, H-5,1%a,1%b), 2.16 (t, 2
H, J 7.5 Hz, COCH2), 2.09, 2.04, 2.03, 1.95 (4
s, 12 H, 4 CH3CO), 1.63–1.25 (m, 52 H, 26
CH2 alkyl chains), 0.88 (t, 6 H, J 6.7 Hz, 2
CH3 alkyl chains). 13C NMR (CDCl3): l
173.40, 170.85, 170.61, 170.58, 169.36 (5
C, 3 CH3COO, CH3CON, CH2CON), 100.10
(C-1), 72.97 (C-3), 71.91 (C-5), 70.63 (C-1%),
68.62 (C-4), 62.19 (C-6), 54.19 (C-2), 47.78
(C-2%), 36.91, 31.91, 31.56, 29.70-29.35, 26.02,
25.95, 22.68 (CH2 alkyl chains), 23.29
(CH3CON), 20.72, 20.72, 20.60 (3 C, 3
CH3COO), 14.11 (2 C, 2 CH3 alkyl chains).
Anal. Calcd for C46H84N2O10 (825.15): C,
(2R)-2-Hexadecanoylaminohexadecyl 2-ace-
tamido-2-deoxy-i- -glucopyranoside (18R).—
D
Product 18R was prepared in 86% yield from
17R as described above for 12R. Mp 220 °C
(MeOH). Anal. Calcd for C40H78N2O7·0.5
H2O (708.05): C, 67.85; H, 11.25; N, 3.94.
Found; C, 67.83; H, 11.24; N, 4.24.
(2S)-2-Hexadecanoylaminohexadecyl 2-ace-
tamido-2-deoxy-i- -glucopyranoside (18S).—
D
Obtained in 89% yield from 17S as described
above for 12S. Mp 214 °C (MeOH). Anal.
Calcd for C40H78N2O7·0.5 H2O (708.05): C,
67.85; H, 11.25; N, 3.94. Found; C, 67.82; H,
11.44; N, 4.03.

194
D. Lafont et al. / Carbohydrate Research 336 (2001) 181–194
17. Ivanova, I. A.; Ross, A. J.; Ferguson, M. A. J.; Nikolaev,
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