Synthesis and molecular tumbling properties of Sialyl Lewis X and derived neoglycolipids

Article abstract of DOI:10.1002/1521-3765(20020603)8:11<2454::AID-CHEM2454>3.0.CO;2-U

The sialyl Lewis X (sLe^x) epitope has become a prominent target for biological studies because of its role in inflammation through binding to selectins. This epitope is located at the terminal end in glycosphingolipids and a lactose unit serves

Full text of DOI:10.1002/1521-3765(20020603)8:11<2454::AID-CHEM2454>3.0.CO;2-U

FULL PAPER
Synthesis and Molecular Tumbling Properties of Sialyl Lewis X and Derived
Neoglycolipids
Christian Gege,[a] Armin Geyer,*[b] and Richard R. Schmidt*[a]
Dedicated to Professor Lutz F. Tietze on the occasion of his 60th birthday
Abstract: The sialyl Lewis X (sLeX)
epitope has become a prominent target
for biological studies because of its role
in inflammation through binding to
selectins. This epitope is located at the
terminal end in glycosphingolipids and a
lactose unit serves as spacer to the
ceramide moiety. This paper focuses on
the influence of the spacer structure and
spacer length in regard to the mobility of
the sLeX epitope. To this end sLeX
neoglycolipids 1a c, with one, two, or
three lactose units as spacer between the
sLeX tetrasaccharide epitope and the
membrane anchor, were synthesized.
The synthetic strategy was also applied
to the synthesis of the corresponding
Lewis X (LeX) derivatives. The glycoli-
pids were inserted in model membranes,
and the tumbling frequencies of the sLeX
tetrasaccharide epitopes were then ana-
lysed by NMR spectroscopy. A nona-
ethylene glycol spacer decouples the
carbohydrate moiety from the mem-
brane mobility while (oligo-)lactoses
act as more rigid distance keepers be-
tween the Lewis epitope and the surface
of the membrane. Quantification of the
different degrees of decoupling was
possible by analysis of rotational corre-
lation times.
Keywords: glycolipids
dynamics NMR spectroscopy
sialyl Lewis X ¥ synthesis design
¥ molecular
¥
¥
of the terminal sLeX tetrasaccharide epitope. In this manu-
script, the dependence of the epitope dynamics of sLeX Aa,[5]
of the membrane-anchored sLeX neoglycolipids Bc,[6] and
1a c on spacer structure and spacer length is quantified,
because these neoglycolipids were found to be highly effective
mimics of the natural sLeX gangliosides in dynamic cell rolling
studies.[6, 7] The synthesis of compounds 1a c, that have one,
two, or three lactose units as spacer, is also described in this
paper.
Introduction
Glycosphingolipids, found on the surface of living cells, are
functional elements in cell cell interactions.[1] The amplifi-
cation of relatively weak monovalent carbohydrate lectin
affinities to strong polyvalent contacts is a prerequisite of
many carbohydrate-based recognition processes.[2] High local
concentrations of the carbohydrate ligands on membrane
surfaces and an adequate presentation of the sugar epitopes
result in strong polyvalent receptor affinities.[3] The protein-
independent homophilic carbohydrate carbohydrate inter-
action is another cell-adhesion process that depends on the
pertinent organization of sugar head groups, as has been
found by recent studies with the Lewis X (LeX) antigen
family.[4] The LeX and sialyl Lewis X (sLeX) sphingolipids
occur as oligomeric glycoforms on cellular surfaces
(Scheme 1, Aa, Ab). Fucosylated N-acetyllactosamine trisac-
charide units and one lactose as a spacer increase the mobility
Results and Discussion
Synthesis of Compounds 1a c: For the synthesis of com-
pounds 1a c, we employed a modification of our previously
reported strategy for the formation of the sLeX structures.[6]
To this end a 3b-O-unprotected 2a-O-benzoyl-lactose build-
ing block was used to which, in three successive steps, the
glucosamine residue and then a fucosyl residue was attached.
Finally a Neu5Aca(2-3)Gal-disaccharide building block[8] was
connected.
3b-O-Unprotected 2a-O-benzoyl-lactoside 4 (Scheme 2)
was readily obtained from known 2a-O-unprotected 3b-O-
allyl-lactoside 2.[9] 2a-O-Benzoylation (!3) and then removal
of the 3b-O-allyl group under palladium(ii) chloride catalysis
in aqueous acetic acid[10] afforded acceptor 4 in high yield.
Glycosylation with known glucosamine donor 5[11] under
standard conditions, that is, with catalytic amounts of
[a] Prof. Dr. R. R. Schmidt, Dr. C. Gege
Fakult‰t f¸r Chemie M 725, Universit‰t Konstanz
78457 Konstanz (Germany)
Fax : (‡49)7531-88-3135
E-mail: richard.schmidt@uni-konstanz.de
[b] Prof. Dr. A. Geyer
Institut f¸r Organische Chemie, Universit‰t Regensburg
93040 Regensburg (Germany)
Fax : (‡49)941-943-4627
E-mail: armin.geyer@chemie.uni-regensburg.de
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Chem. Eur. J. 2002, 8, No. 11

2454 2463
Scheme 1. Natural sialyl Lewis X and derived neoglycolipids.
trimethylsilyl trifluoromethanesulfonate (TMSOTf) as acti-
vator, led to exclusive b(1-3)-linkage affording trisaccharide 6
in practically quantitative yield (1H NMR: J1c,2c ˆ 8.3 Hz;
13C NMR: C-1c, d ˆ 101.51). Selective removal of the O-acetyl
in this compound (1H NMR: J1e,2e ˆ 8.1 Hz, 13C NMR: C-1e,
d ˆ 98.97). Hydrogenolytic O-debenzylation and then O-
acetylation gave O-acyl-protected 14. The anomeric O-acetyl
group could be selectively removed with an excess of
piperidinium acetate[15] in THF at 508C to give the 1-O-
unprotected derivative 15. Treatment with trichloroacetoni-
trile in the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene
(DBU) as base afforded the desired trichloroacetimidate 16 as
hexasaccharide donor.
¬
groups under Zemplen conditions at 08C (!7) and then
4c,6c-O-benzylidenation with benzaldehyde dimethylacetal
in the presence of p-toluenesulfonic acid (pTsOH) as
catalyst gave 3c-O-unprotected acceptor 8. Fucosylation of 8
with known donor 9[12] under standard conditions furnished
exclusively the desired a-linked tetrasaccharide, which
was immediately subjected to reductive opening of
the 4c,6c-O-benzylidene ring with sodium cyanoborohydride
in HCl and diethyl ether,[13] thus affording 4c-O-un-
protected tetrasaccharide 10 as acceptor. It is worth mention-
ing that the fucosylation of the corresponding N-acetyl-
protected glucosamine trisaccharide under the same condi-
tions gave only an a/b-mixture (4:1) and moderate yields. This
fact demonstrates the good acceptor properties of the
trichloroethoxycarbonyl (Troc) residue as amino protecting
group.
The synthesis of compounds 1a c was completed with the
help of dialkylglycerols 17a c (Scheme 4); 17a was obtained
as previously described.[6] Its reaction with known donor 18[16]
led, after acid catalyzed removal of the isopropylidene group,
to disaccharide neoglycolipid 17b.[17] Ensuing glycosylation
with donor 18 and then de-O-isopropylidenation led to
tetrasaccharide neoglycolipid 17c.[17] Glycosylation of accep-
tors 17a c with donor 16 furnished hexa-, octa- and
decasaccharides 19a c; only b-linkages were formed
(1H NMR, 19a: J1a,2a ˆ 7.9 Hz ; 19b: J1c,2c ˆ 8.0 Hz, 19c:
J1e,2e ˆ 7.6 Hz). Removal of all O-acyl groups from 19a
c
¬
Glycosylation of 10 with the known Neu5Aca(2-3)Gal
donor 11[6, 14] and TMSOTf as catalyst furnished hexasacchar-
ide 12 in very good yield (Scheme 3). The Troc group in 12 was
removed under reductive conditions, and the liberated amino
group was acetylated, thus furnishing acetylamino derivative
13. The new anomeric linkage could be readily assigned
under Zemplen conditions and then saponification of the
methyl ester of the Neu5Ac moiety furnished target mole-
cules 1a c in very good yields. The structural assignments
were confirmed by the NMR data.
With building block 10, the branching to give the Lewis X
derivatives is possible at a late stage of the synthetic route.
Chem. Eur. J. 2002, 8, No. 11
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R. R. Schmidt et al.
FULL PAPER
Here with 17a in hand, the LeX
analogue of 1a was prepared.
To this end, tetrasaccharide 10
was glycosylated with known
galactosyl donor 20[18] under
borontrifluoride ether catalysis;
the product was treated with
zinc in acetic acid/acetic anhy-
dride under sonication, thus
furnishing N-acetylated penta-
saccharide 21 (Scheme 5). The
HMQC-spectrum exhibits
large coupling constant for the
1e-H signal at d ˆ 4.59 (J1e,2e
a
>
7 Hz); this indicates its b-link-
age. Hydrogenolytic removal of
all O-benzyl groups and then
O-acetylation afforded the per-
O-acylated intermediate 22.
Regioselective 1-O-deacetyla-
tion with piperidinium acetate
(!23) and then treatment with
trichloroacetonitrile in the
presence of DBU as base fur-
nished trichloroacetimidate 24
as pentaosyl donor. Reaction
with 17a as acceptor afforded
the desired b-linked (1H NMR:
J1a,2a ˆ 7.9 Hz) LeX neoglycoli-
pid 25, which on de-O-acylation
¬
under Zemplen conditions fur-
nished target molecule 26. The
structural assignments were
again confirmed by the NMR
data collected for compounds
25 and 26.
Scheme 2. Synthesis of tetrasaccharide intermediate 10.
Molecular tumbling rates of
membrane-anchored sLeX gly-
colipids: Deuterated sodium
dodecylsulfate ([D25]SDS) mi-
celles form a simple membrane-
mimetic environment for the
NMR analysis of amphiphilic
biomolecules.[19] The tumbling
frequency of an SDS micelle is
characterized by an average
rotational correlation time (tc,
the average time taken for the
molecule to rotate through one
radian) of only 5 ns, much
shorter than phospholipid-
based cell membranes. Yet, this
correlation time is long enough
to separate the tc of the micelle
and the tc of the carbohydrate
head group of
a glycolipid
which is inserted into the model
membrane. Changing the na-
Scheme 3. Synthesis of hexasaccharide donor 16.
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Chem. Eur. J. 2002, 8, No. 11

Sialyl Lewis X Neoglycolipids
2454 2463
Scheme 4. Synthesis of target molecule 1a c.
ture of the spacer between the carbohydrate epitope and the
hydrophobic membrane anchor then quantifies the rigidity of
the spacer: the flexible spacer decouples the epitope tumbling
rate from the membrane mobility while the rigid spacer does
not.
between the cross-signal (Ic) and diagonal-signal (Id) inten-
sities according to the formula:[21]
ꢀ
ꢁ
1
1 ‡ x
1 À x
Ic
Id
j sij jˆ
ln
x ˆ
2tmix
Molecular tumbling rates influence the NMR relaxation
behaviour. There are several methods for connecting molec-
ular tumbling rates with spin-relaxation properties.[20] Yet, not
all of them can be applied to glycolipids that are not isotope
enriched and that show line-broadening due to their micelle-
anchoring. Dipolar cross relaxation was selected for two
reasons, firstly, because of its high sensitivity as a proton-
detected method and secondly because of its model-free data
analysis. For a fixed interproton distance, the NOE intensity
depends only on the reorientation frequency of the corre-
sponding interproton vector relative to the applied magnetic
field. Thus, each cross-signal intensity in a NOESY spectrum
describes the dynamics of a selected molecular moiety. The
strong dependence of the NOE intensity on the molecular
tumbling rate in the region of sign inversion of the NOE
quantifies the correlation times of individual molecular
groups. By this, the nanosecond timescale is analyzed in a
measuring window between approximately 0.1 and 10 w0 ¥ tc
Cross-relaxation rates between proton pairs of fixed
interatom distances were then analyzed with the formula:
2 ꢂ
ꢀ
ꢁꢃ
g4h
m0
6tc
sij ˆ
2 À tc
2
10 r6ij ꢀ4p† 1 ‡ ꢀ2wtc†
which contains tc as the only variable term. The rigid chair
conformations of pyran rings constrain several short inter-
proton distances, which therefore exhibit negligible time
1
averaging. Well-separated H NMR resonance signals were
observed for the sLeX tetrasaccharide moieties of 1a c, Aa
and Bc. We selected three intraglycosidic NOE contacts of
galactose (Gal), fucose (Fuc), and sialic acid (Sia) and one
interglycosidic contact to N-acetylglucosamine (GlcNAc)
Fuc-H1 GlcNAc-H3 for the analysis. The data are listed in
Table 1.
A gradation of correlation times was detected within the
sLeX tetrasaccharide units of 1a c, Aa and Bc. The highest
mobility (shortest tc) was detected for sialic acid which always
tumbles at least twice as fast as the LeX trisaccharide unit. The
tc×s of Fuc are shorter than the tc of Gal and describe internal
dynamics within the LeX trisaccharide units. This stepwise
decrease of correlation times within the columns of Table 1
(w0 ˆ spectrometer frequency). For short mixing times (tmix
ˆ
70 ms), each cross-signal intensity in a 2D NOESY spectrum
depends on a single interproton distance, and the correspond-
ing cross-relaxation rate sij is calculated from the ratio
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R. R. Schmidt et al.
FULL PAPER
Neoglycolipid 1a and the
natural sLeX ganglioside Aa
have the same hexasaccharide
moiety, the structural differen-
ces are restricted to the hydro-
phobic membrane anchors.
Their similar correlation times
prove that the tumbling behav-
iour of the sugar head group in
the micellar environment is
hardly influenced by the mem-
brane anchor. Probably, the
aglyconic methylene group acts
as a flexible joint between the
spacer and the hydrophobic
lipid anchor. Two or three lac-
tose units double or triple the
distance between the mem-
brane surface and the sLeX
moiety, respectively. Yet the
average tc×s of the sLeX tetra-
saccharide moieties decrease
only gradually by approximate-
ly 30% between 1a and 1c.
Although the molecular weight
increases from 1a to 1c, the
average correlation time of the
terminal LeX tetrasaccharide
decreases in the same direction.
Thus, the oligolactose spacer
partly decouples the tumbling
of the terminal epitope from
the micelle dynamics. The
nonaethylene glycol spacer in
Bc[6] doubles the tumbling fre-
quency of the sLeX head
group–it divides the correla-
tion times by a factor of two–
and thus increases the overall
Scheme 5. Synthesis of LeX neoglycolipid 26.
mobility of the terminal tetra-
saccharide. Thus, the different
describes the internal dynamics of each sLeX tetrasaccharide
unit, while the rows in indicate the influence of the spacer
unit.
mobilities of the sLeX tetrasaccharide units of 1c and Bc are
due to the effective decoupling of the carbohydrate tumbling
from the micelle tumbling by the nonaethylene spacer. A
quantitative description of the degree of decoupling by an
oligoethylene spacer is given here for the first time. The
tumbling rates are the basis for the interpretation of receptor
binding data of 1a 1c and Bc described in refs. [6, 7]. The
data were measured with isolated glycolipids, which were
inserted into model membranes. For the design of multivalent
ligands,[22] it has to be taken into account that the membrane
anchor also causes differences in the supramolecular arrange-
ment (microdomains) of the glycolipids in the membrane.[7]
Table 1. The cross-relaxation rates sij were determined from NOESY
spectra and then transformed into tumbling rates tc with the formula given
in the text. The structures of naturally occurring sLeX glycosphingolipid Aa
and its analogues 1a c and Bc are given in Scheme 1.
Aa
1a
1b
1c
Bc
sij [sÀ1
]
Gal-H1 Gal-H3
Fuc-H1 Fuc-H2
Fuc-H1 GlcNAc-H3
Sia-H3ax H3eq
À 0.84
À 0.56
À 0.61
À 2.30
À 0.71
À 0.50
À 0.63
À 2.30
À 0.69
À 0.34
À 0.53
À 1.66
À 0.61
À 0.34
À 0.61
À 1.86
À 0.36
n.d.
À 0.33
À 1.10
tc [10À9 s]
Gal-H1 Gal-H3
Fuc-H1 Fuc-H2
Fuc-H1 GlcNAc-H3
Sia-H3ax H3eq
3.6
1.8
2.1
1.45
3.0
1.8
2.2
1.45
3.0
1.3
1.8
1.1
2.6
1.3
2.1
1.2
1.6
n.d.
1.2
0.8
Conclusion
In conclusion, an efficient route for the synthesis of LeX and
sLeX trichloroacetimidate donors 16 and 24 has been devel-
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Sialyl Lewis X Neoglycolipids
2454 2463
H), 7.05 8.08 (m, 35H; 7 C6H5); elemental analysis calcd (%) for
oped. Glycosylation of dialkylglycerols 17a c furnished the
target molecules 1a c with one to three lactose units as
spacer after deprotection. We could separate the influence of
the spacer on the presentation of the carbohydrate epitope by
quantifying molecular tumbling rates of the sLeX tetrasac-
charide head groups. These results underline the dominating
influence of the spacer in the presentation of carbohydrate
epitopes and thus on molecular recognition phenomena.
C61H62O12 ¥ 1³4 H2O (991.65): C 73.88, H 6.35; found: C 73.73, H 6.32.
Benzyl O-(3,4,6-tri-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylami-
no)-b-d-glucopyranosyl)-(1 ! 3)-(2,4,6-tri-O-benzyl-b-d-galactopyrano-
syl)-(1 ! 4)-2-O-benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside (6): A sol-
ution of 4 (4.10 g, 4.15 mmol) and 5[11] (2.81 g, 4.49 mmol) in dry dichloro-
methane (25 mL) with molecular sieves (AW-300) was treated at room
temperature with trimethylsilyl trifluoromethanesulfonate (46 mL,
259 mmol). After being stirred for 15 min, the mixture was neutralized
with triethylamine and concentrated in vacuo. The residue was purified by
flash chromatography (petroleum ether/ethyl acetate 2:1 to 3:2) to give 6
(5.78 g, 95%) as a colourless foam. Rf ˆ 0.24 (petroleum ether/ethyl acetate
2:1); [a]D ˆ À13.8 (c ˆ 1.0 in CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ
2.00, 2.07, 2.09 (3s, 9H; 3COCH3), 3.42 3.55 [m, 4H; HMQC: 3.43 (5a-H),
3.44 (6b-H), 3.51 (5b-H), 3.53 (6'b-H)], 3.67 (ddd, J(4,5) ˆ 10.1, J(5,6) ˆ 2.5,
J(5,6') ˆ 5.1 Hz, 1H; 5c-H), 3.70 3.81 [m, 5H; HMQC: 3.72 (3b-H), 3.74
(3a-H), 3.77 (2b-H), 3.78 (6a-H), 3.78 (2c-H)], 3.89 (dd, J(5,6) ˆ 4.0, 2J ˆ
11.1 Hz, 1H; 6'a-H), 3.92 (d, J(3,4) ˆ 3.1 Hz, 1H; 4b-H), 4.18 4.21 [m, 2H;
HMQC: 4.19 (4a-H), 4.19 (6c-H)], 4.27 (d, 2J ˆ 11.7 Hz, 1H; CHHPh), 4.32
(dd, J(5,6') ˆ 5.1, 2J ˆ 12.3 Hz, 1H; 6'c-H), 4.36 (brd, 1H; NH), 4.39 (d,
2J ˆ 11.7 Hz, 1H; CHHPh), 4.50 4.65 [m, 9H; HMQC: 4.51 (1b-H), 3.55
Experimental Section
General techniques: Solvents were purified according to standard proce-
dures. Flash chromatography was performed on Baker silica gel 60 (0.040
0.063 mm) at a pressure of 0.4 bar. Thin layer chromatography was
performed on Merck silica gel plastic plates, 60F254 or Merck silica gel glass
plates, HPTLC 60F254; compounds were visualized by treatment with a
solution of (NH4)6Mo7O24 ¥ 4H2O (20 g) and Ce(SO4)2 (0.4 g) in 10%
sulfuric acid (400 mL) and heating at 1508C. Optical rotations were
measured on a Perkin-Elmer polarimeter 241 in a 1 dm cell at 228C. NMR
measurements were recorded at 228C on a Bruker AC250 Cryospec or a
Bruker DRX600. TMS or the resonance of the deuterated solvent was used
as internal standard; solvents: CDCl3, d ˆ 7.26; D2O, d ˆ 4.63. Target
2
(1a-H), CH2CCl3, 5CHHPh], 4.75 (d, J ˆ 12.1 Hz, 1H; CHHPh), 4.78 (d,
J(1,2) ˆ 8.3 Hz, 1H; 1c-H), 4.81 (dd, J(2,3) ˆ J(3,4) ꢁ 10.0 Hz, 1H; 3c-H),
4.87 4.99 (m, 4H; 4CHHPh), 5.05 (dd, J(3,4) ˆ J(4,5) ˆ 9.7 Hz, 1H; 4c-H),
5.36 (dd, J(1,2) ˆ J(2,3) ꢁ 8.7 Hz, 1H; 2a-H), 7.01 8.01 (m, 35H; 7C6H5);
13C NMR (151 MHz, CDCl3, excerpt): d ˆ 56.18 (2c-C), 62.17 (6c-C), 68.04
(6a-C), 68.19 (6b-C), 68.75 (4c-C), 71.74 (5c-C), 72.04 (3c-C), 73.09 (2a-C),
73.36 (5b-C), 75.53 (5a-C), 76.00 (4b-C), 76.24 (4a-C), 80.21 (3b-C), 80.41
(3a-C), 80.94 (2b-C), 99.36 (1a-C), 101.51 (1c-C), 102.56 (1b-C); elemental
analysis calcd (%) for C76H80Cl3NO21 ¥ 1³2 H2O (1458.83): C 62.57, H 5.60, N
0.93; found: C 62.40, H 5.43, N 0.67.
molecules (max. 6 mg) were measured in
a 320 mmol solution of
[D25]sodiumdodecyl sulfate ([D25]SDS) in 0.5 mL D2O. At this concen-
tration, each micelle bears approximately one or two glycolipids. NOESY
spectra were acquired with different mixing times, of which only the
shortest tmix (70 ms) were used for the data analysis. MALDI-mass spectra
were recorded on a Kratos Kompact MALDII instrument using a 2,5-
dihydroxybenzoic acid matrix.
Benzyl O-(2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-b-d-glucopy-
ranosyl)-(1 ! 3)-(2,4,6-tri-O-benzyl-b-d-galactopyranosyl)-(1 ! 4)-2-O-
benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside (7): Compound 6 (8.81 g,
6.04 mmol) was dissolved in dry methanol (100 mL), and sodium meth-
oxide (100 mg, 1.85 mmol) was added at 08C. After the mixture had been
stirred for 1.5 h at 08C, TLC showed that there was no more starting
Benzyl O-(3-O-allyl-2,4,6-tri-O-benzyl-b-d-galactopyranosyl)-(1 ! 4)-2-
O-benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside (3): A solution of com-
pound 2[9] (13.3 g, 14.4 mmol) and benzoyl chloride (2.5 mL, 22 mmol) in
dry pyridine (50 mL) was heated at 458C for 18 h. After addition of some
ethanol the mixture was concentrated in vacuo. After addition of dichloro-
methane (300 mL), the solution was washed with 1m hydrochloric acid and
saturated NaHCO3 solution. The organic layer was dried over sodium
sulfate and concentrated in vacuo. Purification was accomplished by flash
chromatography (petroleum ether/ethyl acetate 5:1 to 4:1) to furnish 3
(13.5 g, 91%) as a colourless oil. Rf ˆ 0.32 (petroleum ether/ethyl acetate
4:1); [a]D ˆ À5.7 (c ˆ 1.0 in CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ 3.33
(dd, J(2,3) ˆ 9.7, J(3,4) ˆ 3.0 Hz, 1H; 3b-H), 3.36 3.53 [m, 4H; HMQC:
3.37 (6b-H), 3.39 (5b-H), 3.46 (5a-H), 3.51 (6'b-H)], 3.69 3.73 [m, 2H;
HMQC: 3.71 (3a-H), 3.72 (2b-H)], 3.78 (dd, J(5,6) ˆ 1.7, 2J ˆ 10.9 Hz, 1H;
6a-H), 3.85 (dd, J(5,6') ˆ 4.5, 2J ˆ 10.9 Hz, 1H; 6'a-H), 3.87 (d, J(3,4) ˆ
3.1 Hz, 1H; 4b-H), 4.09 (dd, J(3,4) ˆ J(4,5) ˆ 9.2 Hz, 1H; 4a-H), 4.17 (m,
‡
material. The solution was neutralized with Amberlite IR120 (H ), filtered
and evaporated. Flash chromatography (toluene/acetone/methanol
66:33:1) yielded 7 (7.00 g, 86%) as a colourless foam. Rf ˆ 0.21 (toluene/
acetone/methanol 66:33:1); [a]D ˆ À9.3 (c ˆ 1.0 in CHCl3); 1H NMR
(250 MHz, CDCl3): d ˆ 3.14 4.97 (m, 35H; 31-H, 2b-H, 2c-H, 33-H, 34-H,
35-H, 66-H, NH, CH2CCl3, 6CH2Ph), 5.31 (dd, J(1,2) ˆ 8.1, J(2,3) ˆ 9.3 Hz,
1H; 2a-H), 6.93 7.98 (m, 35H; 7C6H5); elemental analysis calcd (%) for
C70H74Cl3NO18 ¥ H2O (1341.73): C 62.66, H 5.71, N 1.04; found: C 62.52, H
5.75, N 0.77.
Benzyl O-(4,6-di-O-benzylidene-2-deoxy-2-(2,2,2-trichloroethoxycarbonyl-
amino)-b-d-glucopyranosyl)-(1 ! 3)-(2,4,6-tri-O-benzyl-b-d-galactopyran-
osyl)-(1 ! 4)-2-O-benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside (8): Com-
pound 7 (3.74 g, 2.79 mmol) was dissolved in dry acetonitrile (30 mL) with
molecular sieves (AW-300), then benzaldehyde dimethylacetal (510 mL,
3.39 mmol) and p-toluenesulfonic acid (108 mg, 560 mmol) was added at
room temperature. After 3 h, the solution was neutralized with triethyl-
amine, filtrated and concentrated in vacuo. The residue was purified by
flash chromatography (petroleum ether/ethyl acetate 3:1 to 2:1) to give 8
(3.70 g, 94%) as a colourless foam. Rf ˆ 0.24 (petroleum ether/ethyl acetate
2:1); [a]D ˆ À25.4 (c ˆ 1.0 in CHCl3); 1H NMR (250 MHz, CDCl3): d ˆ
3.37 5.01 (m, 35H; 31-H, 2b-H, 2c-H, 33-H, 34-H, 35-H, 66-H, NH,
CH2CCl3, 6CH2Ph), 5.31 (dd, J(1,2) ˆ 8.1, J(2,3) ˆ 9.3 Hz, 1H; 2a-H), 5.55
(s, 1H; CHPh), 7.04 7.98 (m, 40H; 8C6H5); elemental analysis calcd (%)
for C77H78Cl3NO18 (1411.82): C 65.51, H 5.57, N 0.99; found: C 62.53, H 5.69,
N 0.81.
2H; OCH2-CH CH2), 4.24 4.96 [m, 14H; HMQC: 4.25, 4.35 (2d, 2J ˆ
ˆ
11.8 Hz, CH2Ph), 4.45 (1b-H), 4.55 (1a-H), 5 CH2Ph], 5.19 (dd, 2J ˆ 1.3,
3
ˆ
J ˆ 10.5 Hz, 1H; OCH2-CH CHH), 5.32 5.35 [m, 2H; HMQC: 5.33
ˆ
ˆ
(OCH2-CH CHH), 5.33 (2a-H)], 5.93 (m, 1H; OCH2-CH CH2), 6.98
7.99 (m, 35H; 7C6H5); 13C NMR (151 MHz, CDCl3, excerpt): d ˆ 68.18 (6a-
C, 6b-C), 73.04 (5b-C), 73.15 (2a-C), 73.44 (4b-C), 75.59 (5a-C), 76.79 (4a-
C), 79.94 (2b-C), 80.56 (3a-C), 82.24 (3b-C), 99.43 (1a-C), 102.84 (1b-C),
165.15 (COPh); elemental analysis calcd (%) for C64H66O12 (1027.21): C
74.83, H 6.48; found: C 74.91, H 6.52.
Benzyl O-(3-O-allyl-2,4,6-tri-O-benzyl-b-d-galactopyranosyl)-(1 ! 4)-2-
O-benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside (4): Compound 3 (13.5 g,
13.1 mmol) was dissolved in acetic acid (130 mL) and water (13 mL), then
sodium acetate (3.2 g) and palladium chloride (2.7 g) were added. After
24 h, the solution was filtered over celite and washed with diethyl ether.
After evaporation in vacuo, the residue was dissolved in diethyl ether
(400 mL), and the solution was washed with saturated NaHCO3 solution.
The organic layer was dried over sodium sulfate and concentrated in vacuo.
Flash chromatography (petroleum ether/ethyl acetate 3:1 to 2:1, added in
Benzyl O-(3,4-di-O-acetyl-2-O-benzyl-a-l-fucopyranosyl)-(1 ! 3)-(6-O-
benzyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-b-d-glucopyrano-
syl)-(1 ! 3)-(2,4,6-tri-O-benzyl-b-d-galactopyranosyl)-(1 ! 4)-2-O-benzo-
yl-3,6-di-O-benzyl-b-d-glucopyranoside (10): A solution of acceptor 8
(3.63 g, 2.57 mmol) and donor 9[12] (1.86 g, 3.86 mmol) in dry dichloro-
methane (40 mL) with molecular sieves (AW-300) was treated at 08C with
trimethylsilyl trifluoromethanesulfonate (19 mL, 106 mmol). After 30 min,
the mixture was neutralized with triethylamine and concentrated in vacuo.
The residue was purified by flash chromatography (petroleum ether/ethyl
toluene) furnished
4
(10.7 g, 83%) as
a
colourless foam. Rf ˆ 0.42
(petroleum ether/ethyl acetate 2:1); [a]D ˆ À1.3 (c ˆ 1.0 in CHCl3);
1H NMR (250 MHz, CDCl3): d ˆ 3.47 5.01 (m, 23H; 21-H, 2b-H, 23-H,
24-H, 25-H, 46-H, 6CH2Ph), 5.43 (dd, J(1,2) ˆ 8.0, J(2,3) ˆ 9.3 Hz, 1H; 2a-
Chem. Eur. J. 2002, 8, No. 11
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0811-2459 $ 20.00+.50/0
2459

R. R. Schmidt et al.
FULL PAPER
acetate 2:1 to 3:2 to 1:1) to give the tetrasaccharide as a colourless foam.
This compound was dissolved in dry tetrahydrofuran (70 mL) with
molecular sieves (AW-300), and sodium cyanoborohydride (1.6 g, 80 mmol)
was added. The mixture was treated dropwise under cooling with a
saturated solution of hydrochloride in diethyl ether until the solution
remained acidic. After 30 min, sodium hydrogen carbonate was added, the
mixture was filtrated and diluted with a saturated NaHCO3 solution. After
extraction with dichloromethane (3 Â 100 mL), the combined organic
layers were dried over sodium sulfate and concentrated in vacuo. Flash
chromatography (toluene/acetone 9:1 to 8:1 to 7:1) yielded 10 (3.33 g,
75%) as a colourless foam. Rf ˆ 0.31 (toluene/acetone 7:1); [a]D ˆ À34.0
(c ˆ 1.0 in CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ 1.08 (d, J(5,6) ˆ
6.5 Hz, 3H; 6d-CH3), 1.92, 2.09 (2s, 6H; COCH3), 3.22 (brs, 1H; 3c-H),
3.32 (m, 1H; 5a-H), 3.39 (dd, J(5,6) ˆ 4.6, 2J ˆ 8.4 Hz, 1H; 6b-H), 3.42
3.80 [m, 12H; HMQC: 3.43 (5c-H), 3.46 (5b-H), 3.49 (6'b-H), 3.52 (4c-H),
3.56 (2c-H), 3.65 (3a-H), 3.68 (6a-H), 3.68 (3b-H), 3.70 (2b-H), 3.72 (6c-H),
(6d-C), 37.57 (3f-C), 49.08 (5f-C), 53.16 (OCH3), 57.37 (2c-C), 62.40 (9f-C),
64.33 (5d-C), 67.14 (7f-C), 67.22 (8f-C), 67.38 (6e-C), 67.81 (4e-C), 68.08 (6a-
C), 68.53 (6b-C), 69.02 (6c-C), 69.41 (4f-C), 70.22 (3d-C), 70.52 (2e-C),
71.65 (3e-C), 71.89 (6f-C), 71.94 (4d-C), 72.09 (5e-C), 72.9 (2d-C, 3c-C),
73.11 (2a-C), 73.34 (5b-C), 74.88 (5c-C), 75.5 (4c-C, 5a-C), 76.2 (4a-C, 4b-
C), 79.59 (2b-C), 80.52 (3a-C), 82.36 (3b-C), 96.49 (1d-C), 96.99 (2f-C),
98.97 (1e-C), 99.39 (1a-C), 101.40 (1c-C), 102.54 (1b-C), 167.72 (1f-C);
elemental analysis calcd (%) for C130H148N2O42 (2410.59): C 64.77, H 6.19, N
1.16; found: C 64.56 H 5.89, N 0.95.
O-acyl-protected hexasaccharide (14): Compound 13 (575 mg, 239 mmol)
was dissolved in methanol (20 mL) and acetic acid (3 drops). Palladium on
charcoal (100 mg, 10% Pd) was added, and the solution was stirred
vigorously under a hydrogen atmosphere overnight. The catalyst was
filtered off and washed with methanol. After evaporation in vacuo the
residue was dissolved in pyridine (15 mL), and acetic anhydride (15 mL)
was added. After 16 h, the solvent was evaporated, and the residue was
coevaporated with toluene (3 Â ) and purified by flash chromatography
(toluene/acetone 1:1) to give 14 (422 mg, 93%) as a colourless foam. Rf ˆ
0.30 (toluene/acetone 1:1); 1H NMR (250 MHz, CDCl3): d ˆ 1.18 (d,
J(5,6) ˆ 6.5 Hz, 3H; 6d-CH3), 1.64 2.20 (m, 58H; 3f-Hax, 19 COCH3), 2.59
(dd, 2J ˆ 12.6, J(3eq,4) ˆ 4.6 Hz, 1H; 3f-Heq), 3.11 6.43 (m, 45H; 51-H, 52-
H, 53-H, 64-H, 65-H, 96-H, 7f-H, 8f-H, 29f-H, CO2CH3, 2NH), 7.41 7.99
(m, 5H; C6H5); elemental analysis calcd (%) for C85H112N2O51 (1977.80): C
51.62, H 5.71, N 1.42; found: C 51.31 H 5.75, N 1.07.
2
3.79 (6'a-H), 3.79 (2d-H)], 3.82 (dd, J(5,6) ˆ 2.9, J ˆ 10.6 Hz, 1H; 6'c-H),
3.91 (s, 1H; 4b-H), 4.10 (dd, J(3,4) ˆ J(4,5) ˆ 9.2 Hz, 1H; 4a-H), 4.22 (d,
2J ˆ 11.8 Hz, 1H; CHHPh), 4.33 4.89 [m, 23H; HMQC: 4.34 (5d-H), 4.41
(1b-H), 4.47 (1a-H), 4.50 (NH), 4.75 (1c-H), 4.89 (1d-H), 15 CHHPh,
CH2CCl3], 5.22 5.25 [m, 2H; HMQC: 5.23 (3d-H), 5.24 (4d-H)], 5.30 (dd,
J(1,2) ˆ J(2,3) ꢁ 8.7 Hz, 1H; 2a-H), 6.93 8.01 (m, 45H; 9C6H5); 13C NMR
(151 MHz, CDCl3, excerpt): d ˆ 15.88 (6d-C), 56.30 (2c-C), 65.75 (5d-C),
67.90 (6a-C), 68.35 (6b-C), 69.84 (6c-C, 3d-C), 70.41 (4c-C), 71.28 (4d-C),
73.24 (2a-C), 73.41 (2d-C), 73.64 (5b-C), 74.77 (5c-C), 75.42 (5a-C), 76.23
(4a-C, 4b-C), 80.16 (3b-C), 80.40 (3a-C), 80.90 (2b-C), 83.59 (3c-C), 97.79
(1d-C), 99.32 (1a-C), 101.60 (1c-C), 102.49 (1b-C); elemental analysis calcd
(%) for C94H100Cl3NO24 (1734.18): C 65.10, H 5.88, N 0.81; found: C 64.87, H
5.91, N 0.73.
Unprotected hexasaccharide (15): Piperidinium acetate (580 mg) was
added to a solution of compound 14 (1.06 g, 535 mmol) in dry tetrahy-
drofuran (40 mL), and the mixture was stirred for 4 h at 508C. After
evaporation of the solvent, the residue was purified by flash chromatog-
raphy (toluene/acetone 1:1) to give 15 (954 mg, 92%) as a colourless foam.
Rf ˆ 0.42 (toluene/acetone 2:3); 1H NMR (250 MHz, CDCl3): d ˆ 1.18 (d,
J(5,6) ˆ 6.5 Hz, 3H; 6d-CH3), 1.53 2.20 (m, 55H; 3f-Hax, 18COCH3), 2.59
(dd, 1H; 3f-Heq), 3.15 5.57 (m, 45H; 51-H, 42-H, 53-H, 64-H, 65-H, 96-
H, 7f-H, 8f-H, 29f-H, CO2CH3, 2NH, OH), 5.73 (dd, J(1,2) ˆ J(2,3) ˆ
9.7 Hz, 1H; 2a-H), 7.42 8.03 (m, 5H; C6H5); elemental analysis calcd
(%) for C83H110N2O50 (1935.76): C 51.50, H 5.73, N 1.45; found: C 51.95 H
6.02, N 1.74.
Hexasaccharide (12): A solution of acceptor 10 (1.73 g, 1.00 mmol) and
donor 11[6] (1.52 g, 1.56 mmol) in dry dichloromethane (15 mL) with
molecular sieves AW-300 was treated at 08C with trimethylsilyl trifluoro-
methanesulfonate (18 mL, 100 mmol). After being stirred for 2 h, the
mixture was neutralized with triethylamine, filtrated and concentrated in
vacuo. The residue was purified by flash chromatography (toluene/acetone
3:1) to give 12 (1.95 g, 77%) as colourless foam. Remaining acceptor 10 can
be recovered. Rf ˆ 0.33 (toluene/acetone 2:1); [a]D ˆ À30.2 (c ˆ 1.0 in
CHCl3); 1H NMR (600 Hz, CDCl3): d ˆ 1.20 (d, J(5,6) ˆ 6.5 Hz, 3H; 6d-
CH3), 1.73 (dd, J(3ax,4) ˆ 2J ˆ 12.4 Hz, 1H; 3f-Hax), 1.85, 1.86, 1.95, 2.00,
2.07, 2.07, 2.08, 2.08, 2.22 (9s, 27H; 9COCH3), 2.56 (dd, 2J ˆ 12.6, J(3eq,4) ˆ
4.7 Hz, 1H; 3f-Heq), 3.30 5.30 (m, 63H; 51-H, 52-H, 53-H, 64-H, 65-H,
96-H, 29-H, CO2CH3, 2NH, CH2CCl3, 9CH2Ph), 5.38 (dd, J(6,7) ˆ 2.8,
J(7,8) ˆ 9.4 Hz, 1H; 7f-H), 5.56 (m, 1H; 8f-H), 6.94 7.98 (m, 50H;
10C6H5); elemental analysis calcd (%) for C131H147Cl3N2O43 (2543.95): C
61.85, H 5.82, N 1.10; found: C 61.83 H 5.73, N 0.71.
Trichloroacetimidate (16): Trichloroacetonitrile (320 mL, 3.2 mmol) and
1,8-diazabicyclo[5.4.0]undec-7-ene (4 mL) were added to a solution of 15
(614 mg, 317 mmol) in dry dichloromethane (20 mL). After 3 h, the mixture
was concentrated in vacuo. Flash chromatography (toluene/acetone
1:1‡1% triethylamine) furnished 16 (607 mg, 92%) in a a/b ratio of
>19:1 as a pale yellow foam. Rf ˆ 0.53 (toluene/acetone 2:3); 1H NMR
(250 MHz, CDCl3): d ˆ 1.18 (d, J(5,6) ˆ 6.5 Hz, 3H; 6d-CH3), 1.64 2.19
(m, 55H; 3f-Hax, 18COCH3), 2.58 (dd, 1H; 3f-Heq), 3.10 5.78 (m, 45H;
41-H, 52-H, 53-H, 64-H, 65-H, 96-H, 7f-H, 8f-H, 29f-H, CO2CH3, 2NH),
6.62 (d, J(1,2) ˆ 3.7 Hz, 1H; 1a-H), 7.38 7.99 (m, 5H; C6H5), 8.56 (s, 1H;
Acetylamino hexasaccharide (13): A solution of 12 (1.95 g, 767 mmol) in
tetrahydrofuran/acetic anhydride/acetic acid (6:2:1, 24 mL) was treated
with activated zinc powder (1.5 g, activation with 2% CuSO4 in water for
5 min, then subsequent washing with water, methanol and dry diethyl
ether). The mixture was stirred for 4 h at room temperature with sonication
(5 Â 30 min) and then filtered and washed with diethyl ether (400 mL).
After careful addition of a saturated sodium bisulfate solution, the mixture
was stirred vigorously for 2 h. The organic layer was separated, dried over
sodium sulfate, and evaporated under reduced pressure. Purification by
flash chromatography (toluene/acetone 5:2 to 2:1) gave 13 (1.81 g, 93%) as
a colourless foam. Rf ˆ 0.52 (toluene/acetone 1:1); [a]D ˆ À35.2 (c ˆ 1.0 in
CHCl3); 1H NMR (600 MHz, CDCl3, main conformer): d ˆ 1.10 (d,
J(5,6) ˆ 6.5 Hz, 3H; 6d-CH3), 1.50 (s, 3H; COCH3), 1.72 (dd, 2J ˆ
J(3ax,4) ˆ 12.4 Hz, 1H; 3f-Hax), 1.85, 1.86, 1.87, 2.00, 2.04, 2.07, 2.07, 2.09,
2.21 (9s, 27H; 9COCH3), 2.57 (dd, 2J ˆ 12.6, J(3eq,4) ˆ 4.7 Hz, 1H; 3f-Heq),
3.32 3.44 [m, 4H; HMQC: 3.32 (6b-H), 3.33 (5a-H), 3.36 (6'b-H), 3.38 (5b-
H)], 3.57 3.76 [m, 55H; HMQC: 3.57 (6e-H), 3.57 (6'e-H), 3.61 (6f-H),
3.63 (3b-H), 3.64 (3a-H), 3.66 (6a-H), 3.69 (2b-H), 3.69 (5c-H), 3.72 (5e-H),
3.73 (6'a-H), 3.73 (2c-H), 3.81 (2d-H), 3.82 (6c-H), 3.82 (6'c-H), 3.84
(COOCH3), 3.89 (9f-H), 3.96 (4b-H), 4.06 (4a-H), 4.05 (5f-H), 4.19 (3c-H),
4.19 (4c-H), 4.22 (9'f-H), 4.41 (1b-H), 4.48 (1a-H), 4.59 (3e-H), 4.65 (5d-H),
4.81 (1e-H, J(1,2) ꢁ 8.1 Hz), 4.90 (4f-H), 4.93 (2e-H), 4.99 (1c-H), 5.03
(NfH), 5.08 (4e-H), 5.18 (3d-H), 5.22 (1d-H), 5.23 (4d-H), 9CH2Ph], 5.28 (d,
J(1,2) ˆ J(2,3) ꢁ 8.7 Hz, 1H; 2a-H), 5.33 (dd, J(6,7) ˆ 2.8, J(7,8) ˆ 9.5 Hz,
1H; 7f-H), 5.54 (d, J(2,NH) ˆ 8.3 Hz, 1H; NcH), 5.61 (m, 1H; 8f-H), 6.93
7.98 (m, 50H; 10C6H5); 13C NMR (151 MHz, CDCl3, excerpt): d ˆ 15.86
ˆ
NH); elemental analysis calcd (%) for C85H110N3Cl3O50 ¥ H2O (2098.17): C
48.66, H 5.38, N 2.00; found: C 48.97 H 5.90, N 2.02.
Hexasaccharide (19a): A solution of donor 16 (96 mg, 46 mmol) and lipid
17a[6] (50 mg, 92 mmol) in dry dichloromethane (3 mL) with molecular
sieves (AW-300) was treated at 08C with trimethylsilyl trifluoromethane-
sulfonate (3.3 mL, 18 mmol). After being stirred for 3 h, the mixture was
neutralized with triethylamine, filtrated and concentrated in vacuo. The
residue was purified by flash chromatography (toluene/acetone 2:1 to 3:2)
to give 19a (87 mg, 76%) as a colourless foam. Rf ˆ 0.56 (toluene/acetone
1:1, HPTLC); [a]D ˆ À22.1 (c ˆ 1.0 in CHCl3); 1H NMR (600 MHz,
CDCl3): d ˆ 0. 88 (t, 6H; 2CH3), 1.18 1.31 (m, 55H; 26CH2, 6d-CH3),
1.36 1.43 (q, 4H; 2OCH2CH2), 1.68 (dd, 2J ˆ J(3ax,4) ˆ 12.4 Hz, 1H; 3f-
Hax), 1.85, 1.92, 1.94, 1.96, 2.00, 2.05, 2.07, 2.07, 2.08, 2.09, 2.09, 2.10, 2.11,
2.12, 2.13, 2.14, 2.15, 2.20 (18s, 54H; 18COCH3), 2.58 (dd, 2J ˆ 12.5,
J(3eq,4) ˆ 4.4 Hz, 1H; 3f-Heq), 3.08 3.17 [m, 3H; HMQC: 3.11, 3.15
2
(OCH2), 3.17 (2c-H)], 3.24 (dd, J ˆ 10.2, J(2',3') ˆ 5.3 Hz, 1H; 3'-H), 3.32
(dd, 2J ˆ 10.2, J(2',3') ˆ 4.2 Hz, 1H; 3'-H), 3.38 3.67 [m, 7H; HMQC: 3.40
(OCH2), 3.44 (2'-H), 3.46 (5c-H), 3.57 (1'-H), 3.63 (6f-H), 3.65 (5a-H)], 3.71
(dd, J(2,3) ˆ 9.9, J(3,4) ˆ 3.5 Hz, 1H; 3b-H), 3.75 (dd, J(4,5) ˆ J(5,6) ꢁ
6.5 Hz, 1H; 5b-H), 3.80 3.89 [m, 7H; HMQC: 3.81 (4a-H), 3.83 (1'-H),
3.84 (5e-H), 3.86 (COOCH3), 3.87 (4c-H)], 3.99 4.06 [m, 4H; HMQC:
4.01 (6b-H, 6'b-H), 4.03 (6c-H), 4.03 (5f-H)], 4.08 (dd, 2J ˆ 12.9, J(8,9) ˆ
3.4 Hz, 1H; 9f-H), 4.16 (dd, 2J ˆ 11.8, J(8,9) ˆ 5.2 Hz, 1H; 6a-H), 4.21
4.39 [m, 5H; HMQC: 4.22 (3c-H), 4.22 (6e-H), 4.25 (9'f-H), 4.36 (1b-H),
4.37 (6'e-H)], 4.47 (d, 2J ˆ 10.6, 1H; 6'a-H), 4.52 (dd, J(2,3) ˆ 10.1,
2460
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0811-2460 $ 20.00+.50/0
Chem. Eur. J. 2002, 8, No. 11

Sialyl Lewis X Neoglycolipids
2454 2463
J(3,4) ˆ 3.3 Hz, 1H; 3e-H), 4.64 (d, J(1,2) ˆ 7.9 Hz, 1H; 1a-H), 4.77 (d,
J(1,2) ˆ 8.2 Hz, 1H; 1e-H), 4.80 (d, 2J ˆ 11.0 Hz, 1H; 6'c-H), 4.86 5.51 [m,
17H; HMQC: 4.88 (1c-H), 4.88 (4f-H), 4.89 (2e-H), 4.95 (2d-H), 4.95 (4e-
H), 4.97 (2b-H), 4.99 (5d-H), 5.12 (NfH), 5.15 (2a-H), 5.19 (3d-H), 5.31 (4d-
H), 5.32 (1d-H), 5.33 (4b-H), 5.35 (3a-H), 5.43 (7f-H), 5.47 (NcH), 5.50 (8f-
H)], 7.41 8.00 (m, 5H; C6H5); 13C NMR (151 MHz, CDCl3, excerpt): d ˆ
15.80 (6d-C), 37.61 (3f-C), 49.07 (5f-C), 53.15 (OCH3), 58.4 (2c-C), 60.92
(6c-C), 61.37 (6e-C), 61.52 (6b-C), 61.66 (9f-C), 62.17 (6a-C), 64.10 (5d-C),
66.58 (7f-C), 67.35 (4e-C), 67.57 (8f-C), 67.95 (3d-C), 68.80 (2d-C), 69.01
(4b-C, 1'-C), 69.41 (4f-C), 69.87 (2e-C), 70.04 (3'-C), 70.49 (OCH2), 70.93
(5e-C), 71.19 (5b-C), 71.23 (2b-C), 71.36 (3e-C), 71.51 (4d-C, OCH2), 71.86,
71.91 (2a-C, 6f-C), 72.29 (3a-C), 72.4 (3c-C), 72.87 (5a-C), 73.11 (5c-C),
74.23 (4c-C), 75.66 (4a-C), 75.96 (3b-C), 77.31 (2'-C), 95.26 (1d-C), 96.82
(2f-C), 99.39 (1c-C), 99.88 (1e-C), 100.69 (1b-C), 101.11 (1a-C), 165.11
H), 3.60 (5e-H), 3.63 (6k-H), 3.68 (3f-H), 3.69 (3d-H), 3.70 (4e-H), 3.71 (5f-
H), 3.73 (1'-H), 3.78 (4b-H), 3.83 (5i-H), 3.85 (4g-H), 3.85 (COOCH3), 3.92
(6'b-H), 3.92 (4c-H), 3.95 (4d-H), 3.96 (6f-H), 3.96 (6'f-H), 4.01 (6g-H), 4.03
(4a-H), 4.03 (5k-H), 4.05 (9k-H), 4.16 (6'd-H), 4.20 (3g-H), 4.23 (9k-H),
4.20 4.27 (6a-H, 6c-H, 6e-H, 6i-H), 4.31 (1f-H), 4.34 4.45 (6'a-H, 6'c-H,
6'e-H, 6'i-H), 4.41 (1b-H), 4.48 (1d-H), 4.51 (3i-H)], 4.61 (d, J(1,2) ˆ 7.7 Hz,
2H; 1a-H, 1c-H), 4.64 (d, J(1,2) ˆ 7.6 Hz, 1H; 1e-H), 4.68 5.51 [m, 24H;
HMQC: 4.76 (1i-H), 4.78 (6'g-H), 4.87 (2i-H), 4.88 (1g-H), 4.88 (4k-H), 4.92
(2f-H), 4.94 (2h-H), 4.95 (4i-H), 5.00 (5h-H), 5.04 (NkH), 5.07 (2e-H), 5.19
(3h-H), 5.20 (3e-H), 5.24 (2c-H), 5.30 (2b-H), 5.30 (2d-H), 5.30 (4f-H), 5.30
(4h-H), 5.31 (2a-H), 5.31 (1h-H), 5.37 (NgH), 5.43 (7k-H), 5.48 (3c-H), 5.50
(8k-H)], 5.59 (dd, J(2,3) ˆ J(3,4) ˆ 9.4 Hz, 1H; 3a-H), 7.02 8.08 (m, 55H;
11C6H5); 13C NMR (151 MHz, CDCl3, excerpt): d ˆ 15.8 (6h-C), 37.4 (3k-
C), 49.1 (5k-C), 53.1 (OCH3), 58.4 (2g-C), 60.8 (6g-C), 61.4 (6f-C), 61.6 (9k-
C), 62.7 (6b-C), 62.9 (6d-C), 64.1 (5h-C), 66.5 (7k-C), 67.3 (4i-C), 67.6 (8k-
C), 67.8 (4b-C), 67.9 (3h-C), 67.9 (4d-C), 68.8 (1'-C), 68.8 (2h-C), 68.9 (4f-C),
69.3 (2i-C), 69.8 (4k-C), 70.1 (3'-C), 70.3 (2b-C), 70.3 (2d-C), 70.4 (OCH2),
70.9 (5i-C), 71.1 (2f-C), 71.2 (5f-C), 71.3 (2e-C), 71.3 (3i-C), 71.4 (2c-C), 71.4
(OCH2), 71.5 (4h-C), 71.6 (2a-C), 71.9 (3e-C), 71.9 (6k-C), 72.1 (3c-C), 72.2
(5b-C), 72.3 (3g-H), 72.4 (5d-C), 72.5 (3a-C), 73.0 (5a-C), 73.0 (5c-C), 73.0
(5e-C), 73.1 (5g-C), 74.2 (4g-C), 75.2 (4c-C), 75.3 (4a-C), 75.9 (4e-C), 75.9
(3f-C), 77.2 (2'-C), 80.9 (3b-C), 80.9 (3d-C), 95.2 (1h-C), 99.3 (1g-C), 99.8
(1i-C), 100.4 (1b-C), 100.4 (1d-C), 100.7 (1f-C), 101.0 (1e-C), 101.1 (1a-C),
101.1 (1c-C); elemental analysis calcd (%) for C212H260N2O82 ¥ 3H2O
(4202.40): C 60.59, H 6.38, N 0.67; found: C 60.52 H 6.29, N 0.74.
(COPh), 167.84 (1f-C); elemental analysis calcd (%) for C118H180N2O52
¥
2H2O (2494.73): C 56.81, H 7.43, N 1.12; found: C 56.57 H 7.32, N 0.91.
Octasaccharide (19b): A solution of donor 16 (337 mg, 161 mmol) and lipid
17b[17] (445 mg, 321 mmol) in dry dichloromethane (5 mL) with molecular
sieves (AW-300) was treated at room temperature with trimethylsilyl
trifluoromethanesulfonate (12 mL, 64 mmol). After being stirred for 3 h, the
mixture was neutralized with triethylamine, filtrated and concentrated in
vacuo. The residue was purified by flash chromatography (toluene/acetone
8:5 to 1:1) to give 19b (254 mg, 47%) and unreacted 17b (301 mg) as
colourless foams. Rf ˆ 0.45 (toluene/acetone 1:1, HPTLC); [a]D ˆ ‡2.0
(c ˆ 1.0 in CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ 0.88 (t, 6H; 2CH3),
1.16 (d, 3H; 6f-CH3), 1.25 1.30 (brs, 52H; 26CH2), 1.55 (q, 4H;
2OCH2CH2), 1.65 1.69 (m, 1H; 3h-Hax), 1.78 2.19 (m, 54H; 18COCH3),
2.58 (dd, 1H; 3h-Heq), 3.03 3.16 [m, 5H; HMQC: 3.05, 3.08 (OCH2CH2),
3.15 (5e-H)], 3.19 (dd, J(2',3') ˆ 5.4, 2J ˆ 10.3 Hz, 1H; 3'-H), 3.26 (dd,
J(2',3') ˆ 4.2, 2J ˆ 10.3 Hz, 1H; 3'-H), 3.31 3.50 [m, 6H; HMQC: 3.32
(OCH2), 3.39 (2'-H), 3.43 (5e-H), 3.47 (1'-H), 3.48 (5b-H)], 3.59 3.72 [m,
8H; HMQC: 3.59 (6b-H), 3.63 (5a-H), 3.62 (5c-H), 3.63 (6 h-H), 3.68 (3b-
H), 3.68 (3d-H), 3.70 (4c-H), 3.70 (5d-H)], 3.76 (dd, J(1',2') ˆ 4.8, 2J ˆ
10.5 Hz, 1H; 1'-H), 3.82 4.44 [m, 22H; HMQC: 3.83 (5g-H), 3.85 (4e-
H), 3.85 (COOCH3), 3.93 (4b-H), 3.93 (6d-H), 3.97 (6'd-H), 4.01 (6e-H),
4.02 (5h-H), 4.04 (6c-H), 4.06 (4a-H), 4.07 (9h-H), 4.19 (6'b-H), 4.21 (3e-H),
4.21 (6g-H), 4.23 (9h-H), 4.30 (6a-H), 4.32 (1d-H), 4.37 (6'g-H), 4.39 (6'a-
H), 4.42 (6'c-H)], 4.49 (d, J(1,2) ˆ 8.1 Hz, 1H; 1b-H), 4.51 (dd, J(2,3) ˆ 10.1,
J(3,4) ˆ 3.5 Hz, 1H; 3g-H), 4.64 (d, J(1,2) ˆ 8.0 Hz, 2H; 1a-H, 1c-H), 4.75
5.36 [m, 19H; HMQC: 4.75 (1g-H), 4.78 (6'e-H), 4.88 (1e-H), 4.88 (2g-H),
4.88 (4h-H), 4.92 (2d-H), 4.94 (2f-H), 4.95 (4g-H), 4.99 (5f-H), 5.05 (NhH),
5.07 (2c-H), 5.18 (3f-H), 5.19 (3c-H), 5.30 (4d-H), 5.30 (4f-H), 5.31 (1f-H),
5.33 (2b-H), 5.34 (2a-H), 5.35 (NeH)], 5.43 (dd, J(6,7) ˆ 2.9, J(7,8) ˆ 9.6 Hz,
1H; 7h-H), 5.50 (ddd, J(7,8) ˆ 9.5, J(8,9) ˆ J(8,9') ꢁ 3.3 Hz, 1H; 8f-H), 5.62
(dd, J(2,3) ˆ J(3,4) ˆ 9.5 Hz, 1H; 3a-H), 7.08 7.97 (m, 30H; 6C6H5);
13C NMR (151 MHz, CDCl3, excerpt): d ˆ 16.22 (6f-C), 37.75 (3h-C), 49.45
(5h-C), 53.61 (OCH3), 58.80 (2e-C), 61.25 (6e-C), 61.64 (6g-C), 61.79 (6d-
C), 62.09 (6c-C), 62.09 (9h-C), 62.87 (6a-C), 63.16 (6b-C), 64.49 (5f-C),
66.91 (7h-C), 67.68 (4g-C), 67.89 (8h-C), 68.31 (4b-C), 69.20 (1'-C), 69.22
(2f-C), 69.33 (4f-C), 69.80 (4h-C), 70.45 (3'-C), 70.12 (2g-C), 70.68 (2b-C),
71.30 (5g-C), 71.43 (2d-C), 71.59 (5d-C), 71.68 (2c-C), 71.70 (3g-C), 71.86
(4d-C), 72.18 (2a-C), 72.25 (6h-C), 72.39 (3c-C), 72.67 (3e-C), 72.74 (5b-C),
72.97 (3a-C), 73.29 (5a-C), 73.29 (5c-C), 73.48 (5e-C), 74.54 (4e-C), 75.89
(4c-C), 75.78 (4a-C), 76.27 (3d-C), 77.56 (2'-C), 81.31 (3b-C), 95.61 (1f-C),
99.76 (1e-C), 100.21 (1g-C), 100.94 (1b-C), 101.04 (1d-C), 101.44 (1a-C, 1c-
C); elemental analysis calcd (%) for C165H220N2O67 ¥ H2O (3321.55): C 59.67,
H 6.74, N 0.84; found: C 59.51 H 6.71, N 0.80.
Neoglycolipid 1a: Compound 19a (249 mg, 97.7 mmol) was dissolved in dry
methanol (50 mL), and sodium methoxide (126 mg, 3.3 mmol) was added.
After being stirred for 3 d at room temperature, the solution was
‡
neutralized with Amberlite IR120 (H ), filtered and evaporated. The
residue was dissolved in dioxane/water (1:1, 30 mL), and an aqueous
solution of potassium hydroxide (0.2m, 1 mL) was added. After overnight
stirring, carbon dioxide was added, and the solution was evaporated in
vacuo. Flash chromatography (CHCl3/methanol/water/triethylamine
70:30:5:1) yielded 1a (153 mg, 83%) as
a colourless powder after
lyophilization from water. Rf ˆ 0.23 (CHCl3/methanol/0.2% aqueous CaCl2
1
65:35:8, HPTLC); H NMR (600 MHz, [D25]SDS/D2O): d ˆ 0.79 (m, 6H;
2CH3), 1.16 (d, J(5,6) ˆ 6.4 Hz, 3H; 6d-CH3), 1.21 1.31 (m, 61H; 26CH2,
N(CH2CH3)3), 1.57 (brs, 4H; 2OCH2CH2), 1.79 (t, 1H; 3f-Hax), 2.02, 2.03
(2s, 6H; 2COCH3), 2.75 (dd, 1H; 3f-Heq), 3.20 (q, 6H; N(CH2Me)3), 3.34
(dd, J(1,2) ˆ J(2,3) ˆ 8.4 Hz, 1H; 2a-H), 3.48 4.14 (m, 42H; 3a-H, 4a-H,
5a-H, 26a-H, 2b-H, 3b-H, 4b-H, 5b-H, 26b-H, 2c-H, 3c-H, 4c-H, 5c-H, 26c-
H, 2d-H, 3d-H, 4d-H, 2e-H, 3e-H, 4e-H, 5e-H, 26e-H, 4f-H, 5f-H, 6f-H, 7f-
H, 8f-H, 29f-H, 5H glycerol, 2O(CH2)(CH2)14), 4.43 (d, J(1,2) ˆ 7.8 Hz,
1H; 1a-H), 4.45 (d, J(1,2) ˆ 7.8 Hz, 1H; 1b-H), 4.51 (d, J(1,2) ˆ 7.8 Hz, 1H;
1e-H), 4.71 (1c-H (in HDO-signal)), 4.82 (q, J(5,6) ˆ 6.9 Hz, 1H; 5d-H),
5.12 (d, J(1,2) ˆ 3.9 Hz, 1H; 1d-H); FAB-MS (positive Mode, NBA/
‡
‡
glycerol/MeOH 1:1:1
HNEt3‡2Na]; C84H157N3O35 (1769.16).8888
‡
NaI): 1690 [M À NEt3‡Na], 1712 [M
À
Neoglycolipid 1b: Compound 19b (72 mg, 21.6 mmol) was treated as
described above to furnish 1b (39 mg, 86%) as a colourless powder after
flash chromatography (CHCl3/methanol/water/triethylamine 70:30:4:1 !
65:35:8:1). Rf ˆ 0.18 (CHCl3/methanol/0.2% aqueous CaCl2 65:35:8,
HPTLC); 1H NMR (600 MHz, [D25]SDS/D2O): d ˆ 0.82 (m, 6H; 2CH3),
1.17 (d, J(5,6) ˆ 6.4 Hz, 3H; 6d-CH3), 1.22 1.32 (m, 61H; 26CH2,
N(CH2CH3)3), 1.58 (brs, 4H; 2OCH2CH2), 1.80 (t, 1H; 3h-Ha), 2.03 (s,
6H; 2COCH3), 2.76 (d, 1H; 3h-He), 3.18 (q, 6H; N(CH2Me)3), 3.34 4.21
(m, 56H; 2a-H, 3a-H, 4a-H, 5a-H, 26a-H, 2b-H, 3b-H, 4b-H, 5b-H, 26b-H,
2c-H, 3c-H, 4c-H, 5c-H, 26c-H, 2d-H, 3d-H, 4d-H, 5d-H, 26d-H, 2e-H, 3e-
H, 4e-H, 5e-H, 26e-H, 2f-H, 3f-H, 4f-H, 2g-H, 3g-H, 4g-H, 5g-H, 26g-H,
3h-H, 4h-H, 5h-H, 6h-H, 7h-H, 8h-H, 29h-H, 5H glycerol,
2O(CH2)(CH2)14), 4.44 (d, 2H; 1a-H, 1d-H), 4.54 (m, 2H; 1b-H, 1f-H),
4.73 (brs, 2H; 1c-H, 1e-H (in HDO-signal)), 4.82 (q, 1H; 5f-H), 5.13 (brs,
1H; 1f-H); FAB-MS (positive mode, NBA/glycerol/MeOH 1:1:1 ‡ NaI):
Decasaccharide 19c: A solution of donor 16 (282 mg, 134 mmol) and lipid
17c[17] (609 mg, 271 mmol) in dry dichloromethane (6 mL) with molecular
sieves (AW-300) was treated at room temperature with trimethylsilyl
trifluoromethanesulfonate (10 mL, 54 mmol). After being stirred for 31³2 h,
the mixture was neutralized with triethylamine, filtrated and concentrated
in vacuo. The residue was purified by flash chromatography (toluene/
acetone 3:1 to 3:2) to give 19c (194 mg, 34%) and unreacted 17c (380 mg)
as colourless foams. Rf ˆ 0.46 (toluene/acetone 1:1, HPTLC); [a]D ˆ ‡14.2
(c ˆ 1.0 in CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ 0.88 (t, 6H; 2CH3),
1.17 (d, 3H; 6h-CH3), 1.25 1.33 (m, 52H; 26CH2), 1.53 1.69 (m, 5H;
2OCH2CH2, 3k-Hax), 1.77 2.19 (m, 54H; 18COCH3), 2.58 (dd, 1H; 3k-
Heq), 3.02 4.52 [m, 54H; HMQC: 3.04, 3.08 (OCH2), 3.15 (2g-H), 3.17 (3'-
H), 3.25 (3'-H), 3.31 (OCH2), 3.32 (5b-H), 3.37 (2'-H), 3.43 (5g-H), 3.46 (1'-
H), 3.47 (6b-H), 3.52 (5d-H), 3.55 (3b-H), 3.58 (6d-H), 3.60 (5a-H), 3.60 (5c-
‡
‡
1993 [M À NEt3‡H], 2014 [M À NEt3‡Na]; C96H177N3O45 (2093.45).
Neoglycolipid 1c: Compound 19c (151 mg, 35.9 mmol) was treated as
described above to furnish 1c (80 mg, 92%) as a colourless powder after
flash chromatography (CHCl3/methanol/water/triethylamine 70:30:4:1 !
65:35:8:1). Rf ˆ 0.13 (CHCl3/methanol/0.2% aqueous CaCl2 65:35:8,
1
HPTLC); H NMR (600 MHz, [D25]SDS/D2O, excerpt): d ˆ 0.75 (m, 6H;
2CH3), 1.12 (d, J(5,6) ˆ 6.4 Hz, 3H; 6d-CH3), 1.17 1.27 (m, 61H; 26CH2,
N(CH2CH3)3), 1.53 (brs, 4H; 2OCH2CH2), 1.75 (t, 1H; 3j-Ha), 1.97, 1.98 (2
Chem. Eur. J. 2002, 8, No. 11
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0811-2461 $ 20.00+.50/0
2461

R. R. Schmidt et al.
FULL PAPER
s, 6H; 2COCH3), 2.72 (dd, 1H; 3j-He), 3.16 (q, 6H; N(CH2Me)3), 3.25
4.15 (m, 68H; 2a-H, 3a-H, 4a-H, 5a-H, 26a-H, 2b-H, 3b-H, 4b-H, 5b-H,
26b-H, 2c-H, 3c-H, 4c-H, 5c-H, 26c-H, 2d-H, 3d-H, 4d-H, 5d-H, 26d-H, 2e-
H, 3e-H, 4e-H, 5e-H, 26e-H, 2f-H, 3f-H, 4f-H, 5f-H, 26f-H, 2g-H, 3g-H,
4g-H, 5g-H, 26g-H, 2h-H, 3h-H, 4h-H, 2i-H, 3i-H, 4i-H, 5i-H, 2 6i-H, 3j-H,
4j-H, 5j-H, 6j-H, 7j-H, 8j-H, 29j-H, 5H glycerol, 2O(CH2)(CH2)14), 4.39 (d,
2H; 1a-H, 1f-H), 4.45 4.49 (m, 1H; 1b-H, 1d-H, 1i-H), 4.68 (brs, 3H; 1c-
H, 1e-H, 1g-H (in HDO-signal)), 4.77 (q, 1H; 5h-H), 5.12 (d, 1H; 1h-H);
MALDI-MS (negative mode, 4-nitroaniline, MeOH): 2314 [MÀ À HNEt3],
2412 [MÀ À H]; C108H197N3O55 (2417.73).
J(5,6) ˆ 6.2 Hz, 3H; 6d-CH3), 1.94 2.18 (m, 42H; 14COCH3), 3.24 5.74
(m, 37H; 51-H, 52-H, 53-H, 54-H, 55-H, 86-H, CO2CH3, NH), 7.42 8.02
(m, 5H; C6H5); elemental analysis calcd (%) for C65H85NO39 ¥ H2O
(1522.39): C 51.28, H 5.76, N 0.92; found: C 51.25 H 5.80, N 0.85.
O-(2,4,6-Tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-[(2,3,4-tri-O-acetyl-
a-l-fucopyranosyl)-(1 ! 3)]-(2-acetamido-6-O-acetyl-2-deoxy-b-d-gluco-
pyranosyl)-(1 ! 3)-(2,4,6-tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-3,6-
di-O-acetyl-2-O-benzoyl-a/b-d-glucopyranosyl Trichloracetimidate (24):
Trichloroacetonitrile (760 mL, 7.6 mmol) and 1,8-diazabicyclo[5.4.0]undec-
7-ene (2 drops) were added to a solution of 23 (1.15 g, 756 mmol) in dry
dichloromethane (30 mL). After 2 h, the mixture was concentrated in
vacuo. Flash chromatography (toluene/acetone 3:2‡1% triethylamine)
furnished 24 (1.22 g, 95%) in a a/b ratio of >19:1 as a colourless foam. Rf ˆ
0.20 (toluene/acetone 2:1); 1H NMR (250 MHz, CDCl3): d ˆ 1.20 (d,
J(5,6) ˆ 6.6 Hz, 3H; 6d-CH3), 1.58 2.19 (m, 42H; 14COCH3), 3.06 (m,
1H; 2c-H), 3.42 5.77 (m, 35H; 41-H, 42-H, 53-H, 54-H, 55-H, 86-H,
CO2CH3, NH), 6.63 (d, J(1,2) ˆ 3.7 Hz, 1H; 1a-H), 7.27 7.98 (m, 5H;
ˆ
Benzyl O-(2,3,4,6-Tetra-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-[(3,4-di-
O-acetyl-2-O-benzyl-a-l-fucopyranosyl)-(1 ! 3)]-(2-acetamido-6-O-ben-
zyl-2-deoxy-b-d-glucopyranosyl)-(1 ! 3)-(2,4,6-tri-O-benzyl-b-d-galacto-
pyranosyl)-(1 ! 4)-2-O-benzoyl-3,6-di-O-benzyl-b-d-glucopyranoside
(21): A solution of acceptor 10 (1.89 g, 1.09 mmol) and donor 20[18] (1.07 g,
2.18 mmol) in dry dichloromethane (30 mL) with molecular sieves AW-300
was treated at room temperature with borontrifluoride diethyl etherate
(27 mL, 0.2 equiv). After being stirred for 1 h, the mixture was neutralized
with triethylamine, filtrated and concentrated in vacuo. The residue was
purified by flash chromatography (petroleum ether/ethyl acetate 2:1 !
3:2 ! 1:1) to afford an impure derivative, which was dissolved in
tetrahydrofuran/acetic anhydride/acetic acid (6:2:1, 27 mL) and treated
with activated zinc powder (1.2 g). The mixture was stirred for 4 h at room
temperature with sonication (5 Â 30 min) and then filtered and washed with
diethyl ether (400 mL). After careful addition of a saturated sodium
bisulfate solution, the mixture was stirred vigorously for 1 h. The organic
layer was separated, dried over sodium sulfate, and evaporated under
reduced pressure. Purification by flash chromatography (petroleum ether/
ethyl acetate 1:1 ! 4:5) gave 21 (1.82 g, 86%) as a colourless foam. Rf ˆ
0.30 (petroleum ether/ethyl acetate 1:1); [a]D ˆ À39.5 (c ˆ 1.0 in CHCl3);
1H NMR (600 MHz, CDCl3, main conformer): d ˆ 1.14 (d, J(5,6) ˆ 6.4 Hz,
3H; 6d-CH3), 1.39, 1.90, 1.97, 1.98, 2.01, 2.08, 2.15 (7 s, 21H; 7COCH3),
3.30 3.80 [m, 14H; HMQC: 3.30 (5a-H), 3.36 (6b-H), 3.42 (5b-H), 3.44
(6'b-H), 3.45 (2c-H), 3.47 (5c-H), 3.55 (3b-H), 3.56 (5e-H), 3.65 (3a-H), 3.66
(6a-H), 3.68 (6c-H), 3.70 (2b-H), 3.75 (6'a-H), 3.78 (6'c-H)], 3.84 (dd,
J(1,2) ˆ 3.6, J(2,3) ˆ 10.5 Hz, 1H; 2d-H), 3.99 (d, J(3,4) ˆ 2.3 Hz, 1H; 4b-
H), 4.03 5.11 [m, 29H; HMQC: 4.06 (4c-H), 4.07 (4a-H), 4.23 (6e-H), 4.28
(3c-H), 4.30 (6'e-H), 4.42 (1b-H), 4.48 (1a-H), 4.59 (1e-H), 4.78 (5d-H), 4.81
(3e-H), 5.01 (2e-H), 5.09 (1c-H), 5.10 (1d-H), 8CH2Ph], 5.18 (dd, J(2,3) ˆ
10.6, J(3,4) ˆ 3.2 Hz, 1H; 3d-H), 5.27 5.31 [m, 3H; HMQC: 5.28 (4d-H),
5.29 (2a-H), 5.31 (4e-H)], 5.34 (d, J(2,N) ˆ 7.7 Hz, 1H; NH), 6.95 7.97 (m,
40H; 8C6H5); 13C NMR (151 MHz, CDCl3, excerpt): d ˆ 15.83 (6d-C),
58.58 (2c-C), 60.77 (6e-C), 64.28 (5d-C), 66.86 (4e-C), 68.06, 68.15 (6a-C, 6c-
C), 68.38 (6b-C), 69.19 (2e-C), 70.27 (3d-C), 70.87 (3e-C, 5e-C), 71.98 (4d-
C), 73.23 (2a-C), 73.30 (5b-C), 73.36 (2d-C), 73.79 (4c-C), 74.77 (5c-C),
75.51 (5a-C), 76.27 (4b-C), 76.39 (4a-C), 79.53 (2b-C), 80.47 (3a-C), 82.34
(3b-C), 96.83 (1d-C), 99.42 (1a-C), 99.52 (1e-C), 100.93 (1c-C), 102.67 (1b-
C); elemental analysis calcd (%) for C107H119NO32 (1931.10): C 66.55 H 6.21
N 0.73; found: C 66.26 H 6.20 N 0.59.
C6H5), 8.56 (s, 1H;
NH); elemental analysis calcd (%) for
C67H85N2Cl3O39 ¥ 2H2O (1684.79): C 47.76, H 5.32, N 1.66; found: C 47.83
H 5.29, N 1.65.
(1,2-Di-O-hexadecyl-sn-3-glyceryl) O-(2,4,6-Tri-O-acetyl-b-d-galactopyra-
nosyl)-(1 ! 4)-[(2,3,4-tri-O-acetyl-a-l-fucopyranosyl)-(1 ! 3)]-(2-acet-
amido-6-O-acetyl-2-deoxy-b-d-glucopyranosyl)-(1 ! 3)-(2,4,6-tri-O-acet-
yl-b-d-galactopyranosyl)-(1 ! 4)-3,6-di-O-acetyl-2-O-benzoyl-b-d-gluco-
pyranoside (25): A solution of donor 24 (154 mg, 91.4 mmol) and lipid 17a[6]
(101 mg, 187 mmol) in dry dichloromethane (8 mL) with molecular sieves
(AW-300) was treated at room temperature with BF3 ¥ Et2O (4.8 mL,
36 mmol). After being stirred for 90 min, the mixture was neutralized with
triethylamine, filtrated and concentrated in vacuo. The residue was purified
by flash chromatography (toluene/acetone 3:1) to give 25 (125 mg, 66%) as
a colourless foam. Rf ˆ 0.44 (toluene/acetone 2:1); [a]D ˆ À19.5 (c ˆ 1.0 in
CHCl3); 1H NMR (600 MHz, CDCl3): d ˆ 0.88 (t, 6H; 2CH3), 1.20 1.31
(m, 55H; 26CH2, 6d-CH3), 1.36 1.43 (m, 4H; 2OCH2CH2), 1.93, 1.94, 1.97,
1.98, 2.00, 2.01, 2.04, 2.08, 2.08, 2.10, 2.11, 2.14, 2.16, 2.19 (14s, 42H;
14COCH3), 3.05 3.17 [m, 3H; HMQC: 3.04 (2c-H), 3.10, 3.14 (OCH2)],
2
3.23, 3.31 (2dd, J(2',3') ˆ 5.4, J ˆ 10.3 Hz, 2H; 23'-H), 3.38 3.46 [m, 4H;
HMQC: 3.40 (OCH2), 3.42 (5c-H), 3.44 (2'-H)], 3.57 (dd, J(1',2') ˆ 5.9, 2J ˆ
10.5 Hz, 1H; 1'-H), 3.65 (m, 1H; 5a-H), 3.71 (dd, J(2,3) ˆ 9.9, J(3,4) ˆ
3.6 Hz, 1H; 3b-H), 3.76 3.89 [m, 5H; HMQC: 3.77 (5b-H), 3.79 (4c-H),
3.82 (4a-H), 3.84 (1'-H), 3.87 (5e-H)], 3.93 (dd, J(5,6) ꢁ 1.9, 2J ꢁ 12.0 Hz,
1H; 6c-H), 4.02, 4.06 (2dd, 2H; 6b-H, 6'b-H), 4.16 (dd, J(5,6) ˆ 5.2, 2J ˆ
11.8 Hz, 1H; 6a-H), 4.28 4.34 [m, 2H; HMQC: 4.29 (3c-H), 4.32 (6e-H)],
2
4.36 (d, J(1,2) ˆ 7.9 Hz, 1H; 1b-H), 4.46 (d, J ˆ 10.4 Hz, 1H; 6'a-H), 4.52
2
(dd, J(5,6) ˆ 5.7, J ˆ 11.4 Hz, 1H; 6'e-H), 4.61 (d, J(1,2) ˆ 8.1 Hz, 1H; 1e-
H), 4.65 (d, J(1,2) ˆ 7.9 Hz, 1H; 1a-H), 4.94 4.97 [m, 5H; HMQC: 4.94
(5d-H), 4.95 (2b-H), 4.95 (1c-H), 4.95 (2d-H), 4.96 (6'c-H)], 5.01 (dd,
J(2,3) ˆ 10.4, J(3,4) ˆ 3.4 Hz, 1H; 3e-H), 5.09 (dd, 1H; 2e-H), 5.14 5.19
[m, 2H; HMQC: 5.15 (2a-H), 5.18 (3d-H)], 5.30 (d, J(3,4) ˆ 3.2 Hz, 1H; 4b-
H), 5.32 5.37 [m, 4H; HMQC: 5.32 (1d-H), 5.34 (NH), 5.35 (3a-H), 5.36
(4d-H)], 5.41 (d, J(3,4) ˆ 3.1 Hz, 1H; 4e-H), 7.41 8.00 (m, 5H; C6H5);
13C NMR (151 MHz, CDCl3, excerpt): d ˆ 16.14 (6d-C), 59.19 (2c-C), 60.11
(6c-C), 60.86 (6e-C), 61.87 (6b-C), 62.58 (6a-C), 64.45 (5d-C), 67.05 (4e-C),
68.35 (3d-C), 69.20 (2d-C), 69.41 (2e-C), 69.42 (1'-C), 69.52 (4b-C), 70.45
(3'-C), 70.87 (OCH2), 71.33 (3e-C), 71.42 (5e-C), 71.49 (2b-C), 71.54 (5b-C),
71.78 (4d-C), 71.91 (OCH2), 72.27 (2a-C), 72.34 (3c-C), 72.67 (3a-C), 73.24
(5a-C), 73.42 (5c-C), 74.75 (4c-C), 76.06 (4a-C), 76.34 (3b-C), 77.74 (2'-C),
95.65 (1d-C), 99.47 (1c-C), 100.93 (1e-C), 101.07 (1b-C), 101.54 (1a-C);
elemental analysis calcd (%) for C100H155NO41 ¥ 2H2O (2063.34): C 58.21, H
7.77, N 0.68; found: C 58.19 H 7.57, N 0.84.
Acetyl O-(2,4,6-Tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-[(2,3,4-tri-O-
acetyl-a-l-fucopyranosyl)-(1 ! 3)]-(2-acetamido-6-O-acetyl-2-deoxy-b-d-
glucopyranosyl)-(1 ! 3)-(2,4,6-tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-
3,6-di-O-acetyl-2-O-benzoyl-a/b-d-gluco-pyranoside (22): Compound 20
(1.82 g, 942 mmol) was treated as described above for compound 14 to
furnish 22 (1.20 g, 83%) as a colourless foam after flash chromatography
(toluene/acetone 5:2 ! 2:1‡1% methanol). Rf ˆ 0.55 (toluene/acetone
1:1); 1H NMR (250 MHz, CDCl3): d ˆ 1.21 (d, J(5,6) ˆ 6.4 Hz, 3H; 6d-
CH3), 1.94 2.20 (m, 45H; 15COCH3), 3.11 (m, 1H; 2c-H), 3.38 5.69 (m,
3H; 41-H, 42-H, 53-H, 54-H, 55-H, 86-H, CO2CH3, NH), 5.82 (d, J(1,2) ˆ
1
1
8.0 Hz, ³2 H; 1a-Hb), 6.42 (d, J(1,2) ˆ 3.6 Hz, ³2 H; 1a-Ha), 7.41 7.99 (m,
5H; C6H5); elemental analysis calcd (%) for C67H87NO40 (1546.41): C 52.04,
H 5.67, N 0.91; found: C 52.25 H 5.94, N 1.16.
(1,2-Di-O-hexadecyl-sn-3-glyceryl)
O-(b-d-Galactopyranosyl)-(1 ! 4)-
[(a-l-fucopyranosyl)-(1 ! 3)]-(2-acetamido-2-deoxy-b-d-glucopyranosyl)-
(1 ! 3)-(b-d-galactopyranosyl)-(1 ! 4)-b-d-glucopyranoside (26): Com-
pound 25 (106 mg, 51.3 mmol) was dissolved in dry methanol/dichloro-
methane (2:1, 30 mL), and sodium methoxide (54 mg, 1 mmol) was added.
After being stirred overnight at room temperature, the solution was
O-(2,4,6-Tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-[(2,3,4-tri-O-acetyl-
a-l-fucopyranosyl)-(1 ! 3)]-(2-acetamido-6-O-acetyl-2-deoxy-b-d-gluco-
pyranosyl)-(1 ! 3)-(2,4,6-tri-O-acetyl-b-d-galactopyranosyl)-(1 ! 4)-3,6-
di-O-acetyl-2-O-benzoyl-a/b-d-glucopyranose (23): A solution of com-
pound 22 (1.20 g, 780 mmol) in dry tetrahydrofuran (10 mL) was treated
with piperidinium acetate (1.1 g) and then stirred for 4 h at 508C. After
evaporation of the solvent, the residue was purified by flash chromatog-
raphy (toluene/acetone 9:7) to give 23 (1.15 g, 97%) as a colourless foam.
Rf ˆ 0.38 (toluene/acetone 1:1); 1H NMR (400 MHz, CDCl3): d ˆ 1.19 (d,
‡
neutralized with Amberlite IR120 (H ), filtered and evaporated at low
temperature to avoid premature foaming. Flash chromatography (CHCl3/
methanol/water 70:30:4) yielded 26 (64.0 mg, 89%) as a colourless powder
after lyophilization from water/dioxane. Rf ˆ 0.29 (CHCl3/methanol/water
65:35:8, HPTLC); [a]D ˆ À29.7 (c ˆ 1.0 in CHCl3/MeOH 3:1); 1H NMR
(600 MHz, [D25]SDS/D2O): d ˆ 0.76 (brs, 6H; 2CH3), 1.12 (d, J(5,6) ˆ
2462
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Chem. Eur. J. 2002, 8, No. 11

Sialyl Lewis X Neoglycolipids
2454 2463
6.4 Hz, 3H; 6d-CH3), 1.17 1.27 (m, 52H; 26CH2), 1.53 (brs, 4H;
2OCH2CH2), 2.00 (s, 3H; COCH3), 3.30 (dd, J(1,2) ˆ J(2,3) ꢁ 8.3 Hz, 1H;
2a-H), 3.44 4.09 (m, 35H; 3a-H, 4a-H, 5a-H, 26a-H, 2b-H, 3b-H, 4b-H, 5b-
H, 26b-H, 2c-H, 3c-H, 4c-H, 5c-H, 26c-H, 2d-H, 3d-H, 4d-H, 2e-H, 3e-H,
4e-H, 5e-H, 26e-H, 5H glycerol, 2OCH2CH2), 4.37 4.42 (m, 3H; 1a-H,
1b-H, 1e-H), 4.69 (d, J(1,2) ˆ 8.3 Hz, 1H; 1c-H), 4.76 (q, J(5,6) ꢁ 6.6 Hz,
1H; 5d-H), 5.09 (d, J(1,2) ˆ 3.3 Hz, 1H; 1d-H); MALDI-MS (positive
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[8] A related strategy was previously employed: A. Kameyama, H.
Ishida, M. Kiso, A. Hasegawa, Carbohydr. Res. 1991, 209, C1 - C4; J.
Carbohydr. Chem. 1991, 10, 549 560.
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Konstanz (Germany) 1992.
[10] T. Ogawa, S. Nakabayashi, T. Kitajima, Carbohydr. Res. 1983, 114,
225 236.
‡
‡
mode, DHB, CHCl3/methanol): 1398.7 [M ‡Na], 1415.2 [M ‡K];
C67H125NO27 (1376.72).
[11] W. Dullenkopf, J. C. Castro-Palomino, L. Manzoni, R. R. Schmidt,
Carbohydr. Res. 1996, 296, 135 147.
[12] R. Windm¸ller, Diploma Thesis, University of Konstanz (Germany)
1990.
[13] P. J. Garegg, H. Hultberg, S. Wallin, Carbohydr. Res. 1982, 108, 97
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Acknowledgement
This work was supported by the Deutsche Forschungsgemeinschaft and the
Fonds der Chemischen Industrie. We are grateful to Anke Friemel
(University Konstanz) for her help in the structural assignments by NMR
experiments.
[15] W. Flitsch, H.-G. Kneip, Liebigs Ann. Chem. 1985, 1895 1903.
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Received: November 2, 2001 [F3651]
Chem. Eur. J. 2002, 8, No. 11
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0947-6539/02/0811-2463 $ 20.00+.50/0
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