Synthesis of sialyl Le(x) ganglioside analogues modified at C-6 of the galactose residue to elucidate the mechanism of selection recognition

Article abstract of DOI:10.1016/S0008-6215(98)00013-5

Sialyl Lewis X ganglioside analogues modified at C-6 of the galactose residue, having 6-O-methoxy, 6-acetamido and 6-amino functional groups, have been synthesized. Treatment of 2-(trimethylsilyl)ethyl (methyl 5-acetamido- 4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2- nonulopyranosylonate)-(2→3)-2-O-benzoyl-β-D-galactopyranoside with trimethyloxonium tetrafluoroborate or Tf₂O, followed by NAN₃, gave the 6- O-methyl and 6-azido derivatives, respectively, which were converted into their respective methyl 1-thioglycoside derivatives. The glycosyl donors obtained were coupled with 2-(trimethylsilyl)ethyl 2-acetamido-6-O-benzyl-2- deoxy-3-O-(4-methoxybenzyl)-β-D-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl- β-D-galactopyranosyl-(1→4)-2,3,6-tri-O-benzyl-β-D-galactopyranoside in the presence of dimethyl(methylthio)sulfonium triflate (DMTST) to give their respective pentasaccharides. The glycosylation of the pentasaccharide acceptors derived from their precursors by removal of the 4-methoxybenzyl group, with phenyl 1-thioglycoside derivative of L-fucose using N- iodosuccinimide TfOH afforded the corresponding hexasaccharides, which were transformed in good yield, via reductive removal of the benzyl group, the simultaneous transformation of azide to amine and acetamide, O-deacylation and saponification of the methyl ester into the target compounds without the ceramide groups. On the other hand, the proper manipulation of the protecting group of the hexasaccharides, including a transformation of azide into acetamide and trifluoroacetamide, gave the hexasaccharide imidates, which were coupled with (2S,3R,4E)-2-azido-3-O-tert-butyldiphenylsilyl-4- octadecene-1,3-diol. Selective reduction of the azido group, N-acylation with octadecanoic acid, and the complete removal of the protecting groups gave the desired sialyl Le(x) ganglioside analogues.

Full text of DOI:10.1016/S0008-6215(98)00013-5

Carbohydrate Research 306 (1998) 517±530
Synthesis of sialyl Lex ganglioside analogues
modi®ed at C-6 of the galactose residue to
elucidate the mechanism of selection
recognition1
Nobumasa Otsubo, Hideharu Ishida, Makoto Kiso*, Akira Hasegaway
Department of Applied Bioorganic Chemistry, Gifu University, Gifu 501-11, Japan
Received 2 December 1997; accepted 23 December 1997
Abstract
Sialyl Lewis X ganglioside analogues modi®ed at C-6 of the galactose residue, having 6-
O-methoxy, 6-acetamido and 6-amino functional groups, have been synthesized. Treatment
of 2-(trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-ꢀ-D-
galacto-2-nonulopyranosylonate)-(2!3)-2-O-benzoyl-ꢁ-D-galactopyranoside with trimethylox-
onium tetra¯uoroborate or Tf2O, followed by NaN3, gave the 6-O-methyl and 6-azido derivatives,
respectively, which were converted into their respective methyl 1-thioglycoside derivatives. The
glycosyl donors obtained were coupled with 2-(trimethylsilyl)ethyl 2-acetamido-6-O-benzyl-2-
deoxy-3-O-(4-methoxybenzyl)-ꢁ-D-glucopyranosyl-(1!3)-2,4,6-tri-O-benzyl-ꢁ-D-galactopyranosyl-
(1!4)-2,3,6-tri-O-benzyl-ꢁ-D-galactopyranoside in the presence of dimethyl(methylthio)sulfonium
tri¯ate (DMTST) to give their respective pentasaccharides. The glycosylation of the penta-
saccharide acceptors derived from their precursors by removal of the 4-methoxybenzyl group, with
phenyl 1-thioglycoside derivative of L-fucose using N-iodosuccinimide±TfOH a€orded the corre-
sponding hexasaccharides, which were transformed in good yield, via reductive removal of the
benzyl group, the simultaneous transformation of azide to amine and acetamide, O-deacylation
and saponi®cation of the methyl ester into the target compounds without the ceramide groups. On
the other hand, the proper manipulation of the protecting group of the hexasaccharides, including
a transformation of azide into acetamide and tri¯uoroacetamide, gave the hexasaccharide imi-
dates, which were coupled with (2S,3R,4E)-2-azido-3-O-tert-butyldiphenylsilyl-4-octadecene-1,3-
diol. Selective reduction of the azido group, N-acylation with octadecanoic acid, and the complete
removal of the protecting groups gave the desired sialyl LeX ganglioside analogues. # 1998 Elsevier
Science Ltd. All rights reserved
Keywords: Sialyl Lex; Selectin; Ganglioside
1
Synthetic studies on sialoglycoconjugates, Part 106. For Part 105, see Ref. [1].
* Corresponding author.
y
Deceased 10 October 1996.
0008-6215/98/$19.00 # 1998 Elsevier Science Ltd. All rights reserved
PII S0008-6215(98)00013-5

518
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
modi®cation at C-6 of the D-galactose residue, such
1. Introduction
as the 6-O-methoxy, 6-acetamido and 6-amino
derivatives, which are useful probes to prove the
hypothesis described above.
Selectins (E-, P-, and L-selection) are a family of
calcium-dependent adhesion molecules [2±5] that
are demonstrated to recognize sialyl Lewis X (sLex)
as a ligand [6±8]. In our continuing e€orts [9±11]
to elucidate the mechanism of the selectin±sLeX
interaction, we have synthesized sLeX ganglioside
derivatives that are sulfated at C-6 of either the D-
galactose and/or the N-acetyl-D-glucosamine resi-
due [12] and have examined their potency as
ligands for selectins [13]. It is of interest that the
introduction of a sulfo group to the C-6 of D-
galactose completely abolished the ligand activity
to E-selectin, and that this could be explained by a
repulsion between the sulfo group of sLeX and the
negatively charged residue, Gln 80 and Asp 100,
of E-selectin [14].
2. Results and discussion
For the systematic synthesis of the galactose-
modi®ed sialyl LeX hexasaccharides, we employed
2-(trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,9-
tetra-O-acetyl-3,5-dideoxy-D-glycero-ꢀ-D-galacto-
2-nonulopyranosylonate)-(2!3)-2-O-benzoyl-ꢁ-D-
galactopyranoside [12] (1) as the common inter-
mediate (Scheme 1).
Treatment [15] of 1 with 2,6-di-tert-butyl-4-
methylpyridine and trimethyloxonium tetra-
ꢀ
In view of these facts, we describe herein the
synthesis of the sLeX ganglioside analogues with
¯uoroborate in dichloromethane for 4 h at 0 C
a€orded the 6-O-methyl derivative 2 in 86% yield,
Scheme 1.

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
519
while 6-O-tri¯ation of 1 and subsequent treatment
with sodium azide gave the 6-azido derivative 3 in
60% yield. The compounds 2 and 3 were treated
with benzoic anhydride in the presence of 4-dime-
thylaminopyridine to give 4-benzoates 4 (92%)
and 5 (93%), which, on treatment [16] with boron
tri¯uoride etherate in toluene±acetic anhydride,
gave the ꢁ-1-acetates 6 (91%) and 7 (84%). Con-
version of 6 and 7 into the corresponding methyl
ꢁ-thioglycosides 8 (94%) and 9 (75%) was achieved
by treatment [17] with methylthiotrimethylsilane
and boron tri¯uoride etherate in dichloromethane.
Glycosylation of 10 [18] with 1.5 equiv of 8 in
10 and 9 gave the corresponding pentasaccharide
12 (83%).
Removal [20] of the 4-methoxybenzyl group in
11 and 12 in dichloromethane in the presence of
tin(II) chloride, anisole and chlorotrimethylsilane
for 7 h at 20 ꢀC a€orded the desired glycosyl
acceptors 13 (90%) and 14 (96%). The glycosyla-
tion of 13 and 14 with phenyl 2,3,4-tri-O-benzyl-1-
thio-ꢁ-D-fucopyranoside [12] (15) in the presence of
N-iodosuccinimide (NIS)±tri¯uoromethane- sul-
fonic acid (TfOH) [21,22] and 4A MS in benzene
ꢀ
for 7 h at 7 C gave the desired hexasaccharide 16
(96%) and 17 (89%), respectively. The azide of 17
was converted into the tri¯uoroacetamide 18 (46%)
by successive treatments with triphenylphosphine
in benzene±water and tri¯uoroacetic anhydride in
dichloromethane±pyridine, to serve an inter-
mediate of the 6-amino derivative 40 (Scheme 2).
ꢀ
dichloromethane for 21 h at 0 C in the presence
of 8.0 equiv of dimethyl(methylthio)sulfonium
tri¯ate (DMTST) [19] and powdered 4A molecular
sieves (4A MS) gave the expected pentasaccharide
derivative 11 (90%). In the same way coupling of
Scheme 2.

520
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
Hydrogenolytic removal of the benzyl groups in
unexpectedly lost their binding activity to E-selectin.2
We have already reported that the binding activity
of sLeX ganglioside to E-selectin was completely
abolished, not only by a sulfation [13], but also by
a deoxygenation [11,27] of C-6 of the galactose
residue. Taking account of these results, it is
strongly suggested that E-selectin recognizes the
C-6 of the galactose residue of sLeX in a very
severe manner.
16 over 10% Pd±C in acetic acid gave 19 (53%),
and a similar hydrogenation of 17 in the presence
of acetic anhydride gave the 6-acetamido derivative
20 (57%), both of which were converted by O-
deacylation with sodium methoxide in methanol
and subsequent saponi®cation of the methyl ester,
into the target compounds 21 (99%) and 22 (98%),
while O-deacylation and saponi®cation of 17 (91%
in two steps), followed by hydrogenation (77%)
over Pd(OH)2, gave the 6-amino derivative 24.
Removal of the benzyl groups from 16, 17, and
18 by catalytic hydrogenolysis over Pd(OH)2 and
subsequent acetylation gave the per-O-acylated
hexasaccharides 25 (78%), 26 (77%), and 27
(80%). The azido group in 17 was simultaneously
transformed into the acetamido, groups as descri-
bed for 20. For the selective removal of the 2±(tri-
methylsilyl)ethyl group, the fully acylated oligo-
saccharides 25, 26 and 27 were treated [23] with
tri¯uoroacetic acid in dichloromethane for 3 h at
room temperature to give the 1-hydroxy compounds,
which, on further treatment [24] with trichloro-
acetonitrile in the presence of 1,8-diazabicy-
clo[5.4.0]undec-7-ene (DBU) in dichloromethane
for 2 h at 0 ꢀC, gave the corresponding tri-
chloroacetimidates 28 (81%), 29 (88%), and 30
(84%), respectively.
3. Experimental
General procedures.ÐSpeci®c rotations were
determined with a Union PM-201 polarimeter at
ꢀ
1
25 C, and H NMR spectra were recorded at
400 MHz with a Varian Inova 400, or 200 MHz
with a Varian Gemini-2000 spectrometer. Pre-
parative TLC was performed on Silica Gel 60 (E.
Merck), and column chromatography on silica gel
(Fuji Silysia Co., 300 mesh) was accomplished
with the solvent systems (v/v) speci®ed. Con-
centrations and evaporations were conducted in
vacuo.
2-(Trimethylsilyl)ethyl (methyl 2-acetamido-4,7,
8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-O-benzoyl-6-O-
methyl-b-D±galactopyranoside (2).ÐTo a solution
of 1 (ꢀ400 mg, 0.47 mmol) in CH2Cl2 (2 mL), cooled
to 0 C, was added 2,6-di-tert-butyl-4-methylpyr-
idine (288 mg, 1.4 mmol) and trimethyloxonium
tetra¯uoroborate (173 mg, 1.2 mmol), and the mix-
ture was stirred for 4 h at 0 ꢀC. After completion of
the reaction, MeOH (2 mL) was added to the mix-
ture, which was then concentrated. Column chro-
matography (35:1 CH2Cl2±MeOH) of the residue
on silica gel gave 2 (352 mg, 86%) as an amor-
phous mass: [ꢀ]D+2.9ꢀ (c 0.9, CHCl3); IR (®lm)
Glycosylation of (2S,3R,4E)-2-azido-3-O-(tert-
butyldiphenylsilyl)-4-octadecene-1,3-diol [25] (31)
with the glycosyl donors 28±30 which was carried
out in the presence of trimethylsilyl tri-
¯uoromethanesulfonate and 4A MS (AW-300) for
ꢀ
24 h at 0 C, a€orded the desired ꢁ-glycosides 32
(33%), 33 (58%) and 34 (65%), respectively.
Selective reduction [26] of the azido group in 32±
34 with triphenylphosphine in 20:1 benzene±water
gave the amine, which, on condensation with stearic
acid using 1-(3-dimethylaminopropyl)-3-ethylcar-
bodiimide hydrochloride in dichloromethane, gave
the fully protected sialyl LeX ganglioside derivatives
35 (81%), 36 (70%), and 37 (48%), respectively.
Finally, removal [25] of the tert-butyldiphenyl-
silyl group in 35±37 with 1.0 M tetrabutylammonium
¯uoride in acetonitrile, O-deacylation, and simul-
taneous deprotection of N-tri¯uoroacetyl group
with sodium methoxide in 5:1 methanol±tetra-
hydrofuran for 24 h at 40 ꢀC, and subsequent
saponi®cation of the methyl ester a€orded the
desired sialyl LeX ganglioside analogues 38 (98%),
39 (87%) and 40 (93%) that are modi®ed at C-6 of
the galactose residue. All of the products 38±40
1
3550, 3350, 2950, 1750, 1680, 860, 840, 700 cm ;
1H NMR (CDCl3): ꢂ 7.57±8.27 (m, 5 H, Ph), 5.62
(m, 1 H, H-8), 5.42 (dd, 1 H, H-2a), 4.84 (m, 1 H,
H-4b), 4.52 (dd, 1 H, J2,3 9.9 Hz, J3,4 2.9 Hz, H-3a),
4.43 (dd, 1 H, Jgem 12.5, J8,9 2.4 Hz, H-90), 3.91 (s,
0
3 H, COOMe), 3.53 (s, 3 H, MeO), 2.69 (dd, 1
H, Jgem 12.8, J3eq,4, 4.6 Hz, H-3beq), 2.19, 2.09,
1.93, 1.69, (4 s, 12 H, 4 AcO) 1.38 (s, 3 H, AcN),
0.94 (m, 2H, Me3SiCH2CH2). Anal. Calcd for
C39H57NO19Si (871.96): C, 53.72; H, 6.59; N, 1.61.
Found: C, 53.45; H, 6.42; N, 1.44.
2
The detailed results of the biological studies will be
published by H. Kondo et al. elsewhere.

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
521
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,
7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-gal-
acto-2-nonulopyranosylonate)-(2!3)-6-azido-2-O-
benzoyl-6-deoxy-b-D-galactopyranoside (3).ÐTo a
solution of 1 (100 mg, 0.12 mmol) in pyridine
Jgem 12.9, J3eq,4, 4.5 Hz, H-3beq), 2.23, 2.08, 1.92,
1.78, (4 s, 12 H, 4 AcO) 1.44 (s, 3 H, AcN),
0.95 (m, 2 H, Me3SiCH2CH2). Anal. Calcd for
C44H59NO20Si (950.03): C, 55.63; H, 6.26; N, 1.47.
Found: C, 55.57; H, 5.98; N, 1.40.
ꢀ
(3 mL) and CH2Cl2 (3 mL), cooled to 20 C, was
added tri¯uoromethanesulfonic anhydride (39 ꢃL,
0.23 mmol), and the mixture was stirred for 1 h at
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-gal-
acto-2-nonulopyranosylonate)-(2!3)-6-azido-2,4-
di-O-benzoyl-6-deoxy-b-D-galactopyranoside (5).Ð
To a solution of 3 (670 mg, 0.759 mmol) in pyr-
idine (6 mL) containing 4-dimethylaminopyridine
(28 mg, 0.23 mmol) was added benzoic anhydride
(344 mg, 1.5 mmol), and the mixture was stirred for
20 h at room temperature. Workup as described for
4 gave 5 (679 mg, 93%) as an amorphous mass: [ꢀ]D
+44.5ꢀ (c 1.4, CHCl3); IR (®lm) 3350, 2950, 2130,
ꢀ
20 C then extracted with CH2Cl2. The extract
was washed with 2 M HCl, M Na2CO3 and water,
dried (Na2SO4) and concentrated. The residue was
dissolved in DMF (5 mL) and treated with sodium
ꢀ
azide (76.0 mg, 1.2 mmol) for 12 h at 10 C, then
extracted with EtOAc. The extract was washed
with water, dried (Na2SO4) and concentrated.
Column chromatography (70:1 CH2Cl2±MeOH)
of the residue on silica gel gave 3 (64.1 mg, 60%)
as an amorphous mass: [ꢀ]D+7.7ꢀ (c 0.7, CHCl3);
IR (®lm) 3550, 3350, 2950, 2130,1750, 1680, 860,
1
1750, 1680, 860, 840, 700 cm ; 1H NMR
(CDCl3): ꢂ 7.38±8.30 (m, 10 H, 2 Ph), 5.64 (m, 1
H, H-8), 5.55 (dd, 1 H, J2,3 10.1 Hz, H-2a), 4.82
(d, 1 H, J1,2 8.0 Hz, H-1a), 4.44 (dd, 1 H, Jgem
1
1
840, 700 cm ; H NMR (CDCl3): ꢂ 7.50±8.27 (m,
5 H, Ph), 5.64 (m, 1 H, H-8), 4.89 (m, 1 H, H-4b),
4.56 (dd, 1 H, J2,3=9.8 Hz, J3,4 2.9 Hz, H-3a), 4.41
12.1, J8,9 2.4 Hz, H-90), 3.88 (s, 3 H, COOMe),
0
2.69 (dd, 1 H, Jgem 12.7, J3eq,4 4.4 Hz, H-3beq),
2.32, 2.17, 1.99, 1.86 (4 s, 12 H, 4 AcO), 1.56 (s, 3
H AcN), 0.97 (m, 2 H, Me3SiCH2CH2). Anal.
Calcd for C43H56N4O19Si (961.02): C, 53.74; H,
5.87; N, 5.83. Found: C, 53.59; H, 5.75; N, 5.55.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-
dideoxy-D-glycero-a-D-galacto-2-nonulopyranosylon-
ate-(2!3)-1-O-acetyl-2,4-di-O-benzoyl-6-O-methyl-
b-D-galacto-pyranose (6).ÐTo a solution of 4
(197 mg, 0.201 mmol) in toluene (2 mL) was added
acetic anhydride (0.27 mL, 2.9 mmol) and boron
(dd, 1 H, Jgem 12.5, J8,9 1.6 Hz, H-90), 3.90 (s, 3 H,
0
COOMe), 2.62 (dd, 1 H, Jgem 12.7, J3eq4, 4.6 Hz, H-
3beq), 2.26, 2.18, 2.08, 1.92, (4 s, 12 H, 4 AcO) 1.72
(s, 3 H, AcN), 0.98 (m, 2 H, Me3SiCH2CH2). Anal.
Calcd for C38H54 N4O18Si (882.95): C, 51.69; H,
6.16; N, 6.35. Found: C, 51.57; H, 5.94; N, 6.27.
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra±O-acetyl-3,5-dideoxy-D-glycero-a-D-
galacto-2-nonulopyranosylonate)-(2!3)-2,4-di-O-
benzoyl-6-O-methyl-b-D-galactopyranoside (4).Ð
To a solution of 2 (352 mg, 0.44 mmol) in pyridine
(3 mL) containing 4-dimethylaminopyridine (15.0 mg,
0.12 mmol) was added benzoic anhydride (183 mg,
0.81 mmol), and the mixture was stirred for 24 h
at room temperature. At the end of this time the
mixture was concentrated and extracted with
CH2Cl2. The extract was successively washed with
2 M HCl and water, dried (Na2SO4) and con-
centrated. Column chromatography (60:1 CH2Cl2±
MeOH) of the residue on silica gel gave 4 (350 mg,
92%) as an amorphous mass: [ꢀ]D +44.5ꢀ (c 1.4,
CHCl3); IR (®lm) 3350, 2950, 1750, 1680, 860, 840,
.
tri¯uoride etherate (BF3 OEt2, 48.7 ꢃL, 0.18 mmol),
and the mixture was stirred for 3 h at room tem-
perature and then extracted with CH2Cl2. The
extract was successively washed with M Na2CO3
and water, dried (Na2SO4), and concentrated. Col-
umn chromatography (EtOAc) of the residue on
silica gel gave 6 (170 mg, 91%) as an amorphous
mass: [ꢀ]D +62.5ꢀ (c 0.8, CHCl3); IR (®lm) 3350,
1
1
2950, 1750, 1680, 700 cm ; H NMR (CDCl3): ꢂ
7.38±8.13 (m, 10 H, 2 Ph), 6.12 (d, 1 H, H-1a), 5.59
(m, 1 H, H-8), 5.52 (dd, 1 H, J1,2 8.3 Hz, H-2a),
5.31 (d, 1 H, J3,4 2.9 Hz, H-4a), 5.19 (dd, 1 H, J6,7
2.8, J7,8 9.2 Hz, H-7), 5.04 (d, 1 H, J5,NH 9.9 Hz,
NH), 4.70 (dd, 1 H, J2,3 9.8 Hz, J3,4 2.4 Hz, H-3a),
1 1
700 cm ; H NMR (CDCl3): ꢂ 7.45±8.27 (m, 10 H,
2 Ph), 5.64 (m, 1 H, H-8), 5.39 (dd, 1 H, J1,2 7.9,
J2,3 10.1 Hz, H-2a), 5.24 (d, 1 H, J3,4 3.1 Hz, H-4a),
5.19 (dd, 1 H, J6,7 2.6, J7,8 9.3 Hz, H-7), 5.06 (d, 1
H, J5,NH 10.1 Hz, NH), 4.82 (d, 1 H, J1,2 8.1 Hz, H-
1a), 4.76 (dd, 1 H, J2,3 10.1 Hz, J3,4 3.4 Hz, H-3a),
4.33 (dd, 1 H, Jgem 12.3, J8,9 2.4 Hz, H-90), 3.92 (s,
0
3 H, COOMe), 3.30 (s, 3 H, MeO), 4.6 Hz, H-
3beq), 2.48 (dd, 1 H, Jgem 12.6, J3eq,4, 2.21, 2.10, 1.98,
1.92 (5 s, 15 H, 5 AcO), 1.78, 1.45 (s, 3H, AcN).
Anal. Calcd for C41H49NO21 (891.83): C, 55.22; H,
5.54; N, 1.57. Found: C, 55.03; H, 5.38; N, 1.46.
4.35 (dd, 1 H, Jgem 12.5, J8,9 2.4 Hz, H-90), 3.90 (s,
0
3 H, COOMe), 3.33 (s, 3 H, MeO), 2.62 (dd, 1 H,

522
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
.
(2.5 mL) was added TMS SMe (0.14 mL,
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3, 5-
.
0.99 mmol) and BF3 OEt2 (0.11 mL, 0.37 mmol),
dideoxy-D-glycero-a-D-galacto-2-nonulopyranosyl-
onate-(2!3)-1-O-acetyl-6-azido-2,4-di-O-benzoyl-
6-deoxy-b-D-galactopyranose (7).ÐTo a solution of
5 (460 mg, 0.478 mmol) in toluene (3 mL) were added
ꢀ
and the mixture was stirred for 7.5 h at 50 C.
Workup as described for 8 gave 9 (261 mg, 75%) as
an amorphous mass [ꢀ]D +55.0ꢀ (c 5.1, CHCl3); IR
1
1
.
acetic anhydride (0.61 mL, 6.5 mmol) and BF3 OEt2
(®lm) 3350, 2950, 2130, 1750, 1680, 700 cm ; H
NMR (CDCl3): ꢂ 7.24±8.24 (m, 10 H, 2 Ph), 5.60
(m, 1 H, H-8), 5.55 (t, 1 H, H-2a), 4.79 (dd, 1 H,
J2,3 9.8 Hz, J3,4 3.3 Hz, H-3a), 4.78 (d, 1 H, J1,2
(0.12 mL, 0.45 mmol), and the mixture was stirred
for 3 h at room temperature. Workup as described
for 6 gave 7 (375 mg, 84%) as an amorphous mass:
[ꢀ]D +53.0ꢀ (c 1.7, CHCl3); IR (®lm) 3350, 2950,
0
9.9 Hz, H-1a), 4.35 (dd, 1 H, Jgem 12.5, J8,9 2.4 Hz,
1
1
2130, 1750, 1680, 700 cm ; H NMR (CDCl3):ꢂ,
7.28±8.14 (m, 10 H, 2 Ph), 6.09 (d, 1 H, H-1a), 5.47
(dd, 1 H, J1,2 8.2 Hz, H-2a), 4.98 (dd, 1 H, J2,3
10.2 Hz, J3,4 3.3 Hz, H-3a), 4.79 (m, 1 H, H-4b),
H-90), 3.88 (s, 3 H, COOMe), 2.52 (dd, 1 H, Jgem
12.5, J3eq,4 4.5 Hz, H-3beq), 2.25 (s, 3 H, SMe),
2.22, 2.09, 1.91, 1.78 (4 s, 12 H, 4 AcO), 1.49 (s, 3
H, AcN). Anal. Calcd for C41H48N4O18S (916.91):
C, 53.71; H, 5.28; N, 6.11. Found: C, 53.70; H,
5.18; N, 5.96.
4.34 (dd, 1 H, Jgem 12.2, J8,9 2.2 Hz, H-90), 3.9 (s, 3
0
H, COOMe), 4.3 Hz, H-3beq), 2.49 (dd, 1 H, Jgem
12.6, J3eq,4, 2.22, 2.11, 1.99, 1.92 (5 s, 15 H, 5 AcO),
1.78, 1.48 (s, 3 H, AcN). Anal. Calcd for
C42H48N4O20 (928.85): C, 54.31; H, 5.21; N, 6.03.
Found: C, 54.28; H, 5.16; N, 5.78.
Methyl (methyl 5-acetamido-4,7,8,9-tetra-O-
acetyl-3,5-dideoxy-D-glycero-a-D-galacto-2-nonulo-
pyranosylonate)-(2!3)-2,4-di-O-benzoyl-6-O-methyl-1-
thio-b-D-galactopyranoside (8).ÐTo a solution of 6
(130 mg, 0.145 mmol) in toluene (2 mL) was added
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-
galacto-2-nonulopyranosylonate)-(2!3)-2,4-di-O-
benzoyl-6-O-methyl-b-D-galactopyranosyl-(1!4)-
2-acetamido-6-O-benzyl-2-deoxy-3-O-(4-methoxy-
benzyl) - b - D - glucopyranosyl - (1!3) - 2,4,6-tri-O-
benzyl -b -D- galactopyranosyl -(1!4) -2,3,6-tri - O -
benzyl-b-D-glucopyranoside (11).ÐTo a solution of
8 (110 mg, 0.125 mmol) and 10 (118 mg, 84 ꢃmol)
in dry CH2Cl2 (3 mL) was added powdered 4A
(MS 500 mg), and the mixture was stirred for 6 h at
.
(methylthio)trimethylsilane (TMS SMe; 49.8 ꢃL,
.
0.35 mmol) and BF3 OEt2 (37.8 ꢃL, 0.14 mmol),
ꢀ
ꢀ
and the mixture was stirred for 2.5 h at 50 C and
room temperature, then cooled to 0 C. Dimethyl-
(methylthio)sulfonium tri¯ate (DMTST; 165 mg,
0.64 mmol) was added to the mixture, which was
extracted with CH2Cl2. The extract was succes-
sively washed with M Na2CO3 and water, dried
(Na2SO4), and concentrated. Column chromato-
graphy (30:1 CH2Cl2±MeOH) of the residue on
silica gel gave 8 (124 mg, 94%) as an amorphous
mass:[ꢀ]D +59.1ꢀ (c 2.4, CHCl3); IR (®lm) 3350,
ꢀ
stirred for 21 h at 0 C. The solids were ®ltered o€
and washed with CH2Cl2. The combined ®ltrate
and washings was washed with M Na2CO3 and
water, dried (Na2SO4), and concentrated. Column
chromatography (40:1 CH2Cl2±MeOH) of the
residue on silica gel gave 11 (170 mg, 90%) as an
amorphous mass: [ꢀ]D 41 +7.3ꢀ (c 1.1, CHCl3); IR
1
1
2950, 1750, 1680, 700 cm ; H NMR (CDCl3): ꢂ
7.31±8.16 (m, 10 H, 2 Ph), 5.59 (m, 1 H, H-8), 5.52
(t, 1 H, J1,2 = J2,3 = 9.9 Hz, H-2a), 5.31 (d, 1 H,
J3,4 2.9 Hz, H-4a), 5.21 (dd, 1 H, J6,7 2.6, J7,8
9.2 Hz, H-7), 4.86 (dd, 1 H, J2,3 9.8 Hz, J3,4 3.0 Hz,
H-3a), 4.78 (d, 1 H, J1,2 9.7 Hz, H-1a), 4.34 (dd, 1
1
(®lm) 3350, 2950, 1750, 1680, 860, 840, 700 cm ;
1H NMR (CDCl3): ꢂ 7.05±8.24 (m, 49 H, 9 Ph,
MeOPh, 5.67 (m, 1 H, H-8), 5.46 (t, 1 H, J2,3
10.9 Hz, H-2d), 5.22 (dd, 1 H, J6,7 2.1, J7,8 8.1 Hz,
H-7), 4.59 (d, 1 H, J1,2 10.9 Hz, H-1d), 3.89 (s, 3 H,
COOMe), 3.64 (s, 3 H, MeOPh), 3.39 (s, 3 H,
MeO), 2.43 (dd, 1 H, Jgem 12.3, J3eq,4 4.4 Hz, H-
3eeq), 2.17, 1.98, 1.92, 1.78 (4 s, 12 H, 4 AcO), 1.49,
1.22 (2 s, 6 H, 2 AcN), 0.99 (m, 2 H, Me3SiCH2CH2).
Anal. Calcd for C123H144N2O36Si (2254.57): C,
65.53; H, 6.44; N, 1.24. Found: C, 65.26; H, 6.32;
N, 0.95.
H, Jgem 12.5, J8,9 2.3 Hz, H-90), 3.91 (s, 3 H,
0
COOMe), 3.32 (s, 3 H, MeO), 2.47 (dd, 1, H, Jgem
12.8, J3eq,4 4.6 Hz, H-3beq), 2.26 (s, 3 H, SMe),
2.22, 2.08, 1.92, 1.78 (4 s, 12 H, 4 AcO), 1.47 (s, 3
H, AcN). Anal. Calcd for C40H49NO19S (879.89):
C, 54.60; H, 5.61; N, 1.59. Found: C, 54.46; H,
5.48; N,1.34.
Methyl (methyl 5-acetamido-4,7,8,9-tetra-O-
acetyl-3,5-dideoxy-D-glycero-a-D-galacto-2- nonulo-
pyranosylonate)-(2!3)-azido-2,4-di-O-benzoyl-6-
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-
galacto-2-nonulopyranosylonate)-(2!3)-6-azido-
deoxy-1-thio-b-D-galactopyranoside (9).ÐTo
a
solution of 7 (350 mg, 0.377 mmol) in toluene

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
523
2,4-di-O-benzoyl-6-deoxy-b-D-galactopyranosyl-
1!4)-2-acetamido-6-O-benzyl-2-deoxy-3-O-(4- metho-
xybenzyl)-b-D-glucopyranosyl-(1!3)-2,4,6-tri-O-
benzyl-b-D -galactopyranosyl-(1!4)-2,3,6-tri-O-
benzyl-b-D-glucopyranoside (12).ÐGlycosylation
galacto-2-nonulopyranosylonate)-(2!3)-6-azido-
2,4-di-O-benzoyl-6-deoxy-b-D-galactopyranosyl-
(1!4)-2-acetamido-6-O-benzyl-2-deoxy-b-D-gluco-
pyranosyl-(1!3)-2,4,6-tri-O-benzyl-b-D-galacto-
pyranosyl-(1!4)-2,3,6-tri-O-benzyl-b-D-glucopyr-
anoside (14).ÐTo a solution of 12 (24 mg, 11 ꢃmol)
in dry CH2Cl2 (3 mL) were added SnCl2 (0.1 mg,
0.5 ꢃmol) and anisole (1.7 ꢃL, 16 ꢃmol), and the
mixture was cooled to 0 ꢀC. TMSCl (4 ꢃL,
32 ꢃmol) was added to the mixture, which was
stirred for 7 h at 20 ꢀC. Workup as described for 13
gave 14 (22.6 mg, 96%) as a syrup: [ꢀ]D +27.2ꢀ (c
0.4, CHCl3); IR (®lm) 3550, 3350, 2950, 2130,
of 10 (102 mg, 72 ꢃmol) with
9
(100 mg,
0.109 mmol) in dry CH2Cl2 (3 mL) in the presence
of DMTST (141 mg, 0.55 mmol) and 4A MS
(500 mg) for 14 h at 0 ꢀC, then workup as described
for 11, gave 12 (137 mg, 83%) as an amorphous
mass: [ꢀ]D +11.6ꢀ (c 3.2, CHCl3); IR (®lm) 3350,
1
1
2950, 2130, 1750, 1680, 860, 840, 700 cm ; H
NMR (CDCl3): ꢂ 7.08±8.31 (m, 49 H, 9 Ph,
MeOPh), 5.67 (m, 1 H, H-8), 5.45 (t, 1 H, J2,3
10.6 Hz, H-2d), 4.64 (d, 1 H, J1,2 10.6 Hz, 3.87 (s, 3
H, COOMe), 3.67 (s, 3 H, MeOPh), 2.46 (dd, 1 H,
Jgem = 12.4, J3eq,4 = 4.4 Hz, H-3eeq), 2.19 (4 s, 12
H, 4 AcO), 1.98, 1.92, 1.79, 1.49 (2 s, 6 H, 2 AcN),
1.33, 0.92 (m, 2 H, Me3SiCH2CH2). Anal. Calcd
for C122H141N5O35Si (2265.56): C, 64.68; H, 6.27;
N, 3.09. Found: C, 64.51; H, 6.15; N, 3.09.
1
1750, 1680, 860, 840, 700 cm ; 1H NMR (CDCl3):
ꢂ 7.09±8.33 (m, 45 H, 9 Ph), 5.63 (m, 1 H, H-8),
5.44 (dd, 1 H, J2,3 8.1 Hz, H-2d), 4.82 (d, 1 H, J1,2
8.9 Hz, H-1d), 3.87 (s, 3 H, COOMe), 2.59 (dd, 1
H, Jgem 12.8, J3eq,4 4.4 Hz, H-3eeq), 2.22, 2.03, 1.96,
1.78 (4 s, 12 H, 4 AcO), 1.54, 1.22 (2 s, 6 H, 2
AcN), 1.00 (m, 2 H, Me3SiCH2CH2). Anal. Calcd
for C114H133N5O34Si (2145.41): C, 63.82; H, 6.25;
N, 3.26. Found: C, 63.52; H, 6.13; N, 3.06.
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,9-
tetra-O-acetyl-3,5-dideoxy±D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-2,4-di-O-benzoyl-
6-O-methyl-b-D-galactopyranosyl-(1!4)-2-acet-
amido-6-O-benzyl-2-deoxy-b-D-glucopyranosyl-(1!3)-
2,4,6-tri-O-benzyl-b-D-galactopyranosyl-(1!4)-
2,3,6-tri-O-benzyl-b-D-glucopyranoside (13).ÐTo
a solution of 11 (167 mg, 74 ꢃmol) in dry CH2Cl2
(3 mL) were added SnCl2 (0.7 mg, 3.7 ꢃmol) and
anisole (12 ꢃL, 0.11 mmol), and the mixture was
cooled to 0 ꢀC. Trimethylchlorosilane (TMSCI;
28 ꢃL, 0.22 mmol) was added to the mixture,
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-
galacto-2-nonulopyranosylonate)-(2!3)-2,4-di-O-
benzoyl-6-O-methyl-b-D-galactopyranosyl-(1!4)-
[2,3,4-tri-O-benzyl-a-L-fucopyranosyl-(1!3)]-2-
acetamido-6-O-benzyl-2-deoxy-b-D-glucopyranosyl-
(1!3)-2,4,6-tri-O-benzyl-b-D-galactopyranosyl-
(1!4)-2,3,6-tri-O-benzyl-b-D-glucopyranoside (16).Ð
To a solution of 13 (220 mg, 0.10 mmol) and 15
(81.4 mg, 0.15 mmol) in dry benzene (4 mL) was
added powdered 4A MS (500 mg), and the mixture
was stirred for 5 h at room temperature, then
ꢀ
which was stirred for 7 h at 20 C and extracted
ꢀ
with CH2Cl2. The extract was washed with M
Na2CO3 and water, dried (Na2SO4) and con-
centrated. Column chromatography (40:1 CH2Cl2±
MeOH) of the residue on silica gel gave 13 (142 mg,
90%) as a syrup: [ꢀ]D +38.3ꢀ (c 3.4, CHCl3); IR
(®lm) 3550, 3350, 2950, 1750, 1680, 860, 840,
cooled to 0 C. N-Iodosuccinimide (NIS; 104 mg,
0.46 mmol) and tri¯uoromethanesulfonic acid
(TfOH; 11 ꢃL, 0.12 mmol), were added to the mix-
ꢀ
ture, which was stirred for 7 h at 7 C, then neu-
tralized with Et3N. The solids were ®ltered o€ and
washed with CH2Cl2. The combined ®ltrate and
washings was successively washed with M Na2CO3
and water, dried (Na2SO4) and concentrated. Col-
umn chromatography (45:1 CH2Cl2±MeOH) of the
residue on silica gel gave 16 (245 mg, 96%) as a
syrup: [ꢀ]D 19.7ꢀ (c 2.3, CHCl3); IR (®lm) 3350,
1 1
700 cm ; H NMR (CDCl3): ꢂ 7.02±8.28 (m, 45 H,
9 Ph), 5.59 (m, 1 H, H-8), 5.20 (dd, 1 H, J6,7 2.4 J7,8
8.4 Hz, H-7), 4.66 (d, 1 H, J1,2 10.6 HZ, H-1d), 4.47
(dd, 1 H, Jgem 12.5, J8,9 2.4 Hz, H-90), 3.87 (s, 3 H,
COOMe), 3.41 (s, 3 H, MeO), 2.46 (dd, 1 H, Jgem
12.4, J3eq,4 4.3 Hz, H-3eeq), 2.21, 2.03, 1.91, 1.78 (4
s, 12 H, 4 AcO), 1.71, 1.43 (2 s, 6 H, 2 AcN), 1.00
(m, 2 H, Me3SiCH2CH2). Anal. Calcd for
C115H136N2O35Si (2134.42): C, 64.71; H, 6.42; N,
1.31. Found: C, 64.43; H, 6.36; N, 1.07.
1
1
2950, 1750, 1680, 860, 840, 700 cm ; H NMR
(CDCl3): ꢂ 7.14±8.20 (m, 60 H, 12 Ph), 5.55 (m, 1
H, H-8), 5.24 (dd, 1 H, J6,7 2.4, J7,8 8.4 Hz, H-7),
3.84 (s, 3 H, COOMe), 3.45 (s, 3 H, MeO), 2.46
(dd, 1 H, Jgem 12.4, J3eq,4 4.3 Hz, H-3eeq), 2.17, 1.97,
1.92, 1.79 (4 s, 12 H, 4 AcO), 1.46, 1.40 (2 s, 6 H, 2
AcN), 1.20 (d, 3 H, J5,6 6.4 Hz, H-6f), 1.02 (m, 2 H,
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-

524
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
Me3SiCH2CH2). Anal. Calcd for C142H164N2O39Si
(2550.94): C, 66.86; H, 6.48; N, 1.10. Found: C,
66.71; H, 6.36; N, 0.87.
[ꢀ]D 13.5ꢀ (c 1.4, CHCl3); IR (®lm) 3350, 2950,
1
1750, 1680, 860, 840, 700 cm ; 1H NMR (CDCl3):
ꢂ 7.08±8.21 (m, 60 H 12 Ph), 5.65 (m, 1 H, H-8),
3.71 (s, 3 H, COOMe), 2.41 (dd, 1 H, Jgem 12.6,
J3eq,4, 14.7 Hz, H-3eeq), 2.11, 1.92, 1.77, 1.58
(4 s, 12 H, 4 AcO), 1.38, 1.26 (2 s, 6 H, 2 AcN)
(d, 3 H, J5,6 6.3 Hz, H-6f), 1.16, 1.01 (m, 2 H,
Me3SiCH2CH2), Anal. Calcd for C143H162F3N3-
O39Si (2631.93): C, 65.26; H, 6.20; N, 1.60. Found:
C, 65.06; H, 6.03; N, 1.38.
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-
4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-
galacto-2-nonulopyranosylonate)-(2!3)-6-azido-
2,4-di-O-benzoyl-6-deoxy-b-D-galactopyranosyl-
(1!4)-[2,3,4-tri- O -benzyl -a -L -fucopyranosyl-
(1!3)]-2-acetamido-6-O-benzyl-2-deoxy-b-D-gluco-
pyranosyl-(1!3)-2,4,6-tri-O-benzyl-b-D-galacto-
pyranosyl-(1!4)-2,3,6-tri-O-benzyl-b-D-glucopyr-
anoside (17).ÐTo a solution of 14 (148 mg,
69.1 ꢃmol) and 15 (54.5 mg, 0.10 mmol) in dry
benzene (4 mL) was added powdered 4A MS
(500 mg), and the mixture was stirred for 5 h at
room temperature, then cooled to 0 ꢀC. NIS
(69.8 mg, 0.31 mmol) and TfOH (7.3 ꢃL, 83 ꢃmol)
were added to the mixture, which was stirred for
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,
8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-2,4-di-O-benzoyl-
6-O-methyl-b-D-galactopyranosyl-(1!4)-[a-L-fuco-
pyranosyl-(1!3)]-2-acetamido-2-deoxy-b-D-gluco-
pyranosyl-(1!)-b-D-galactopyranosyl-(1!4)-b-D-
glucopyranoside (19).ÐA solution of 16 (103 mg,
40 ꢃmol) in HOAc (2 mL) was hydrogenated over
ꢀ
ꢀ
7 h at 7 C, then neutralize with Et3N. Workup as
described for 16 gave 17 (158 mg, 89%) as a syrup:
[ꢀ]D 9.4ꢀ (c 3.1, CHCl3); IR (®lm) 3350, 2950,
10% Pd±C (150 mg) for 48 h at 40 C, then ®ltered
and concentrated. Column chromatography (15:1
CH2Cl2±MeOH) of the residue on silica gel gave 19
(34.9 mg, 53%) as an amorphous mass: [ꢀ]D 18.7ꢀ
(c 0.7, 1:1 CHCl3±MeOH); IR (®lm) 3550, 3350,
1
1
2130, 1750, 1680, 860, 840, 700 cm ; H NMR
(CDCl3): ꢂ 7.01±8.23 (m, 60 H, 12 Ph), 5.63 (m, 1
H, H-8), 5.20 (t, 1 H, H-2d), 4.49 (d, 1 H, J1,2
7.9 Hz, H-1d), 3.81 (s, 3 H, COOMe), 2.49 (dd, 1
H, Jgem 12.8, J3eq,4 4.6 Hz, H-3eeq), 2.19, 1.96, 1.92,
1.78 (4 s, 12 H, 4 AcO), 1.46, 1.22 (2 s, 6 H, 2 AcN)
(d, 3 H, J5,6 6.4 Hz, H-6f), 1.18, 1.01 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C141H161N5O38Si
(2561.92): C, 66.10; H, 6.33; N, 2.73. Found: C,
66.04; H, 6.30; N, 2.52.
1
1
2950, 1750, 1680, 860, 840, 700 cm ; H NMR
(CD3OD): ꢂ 7.47±8.23 (m, 10 H, 2 Ph),5.62 (m, 1
H, H-8), 5.32 (d, 1 H, J5,NH 9.9 Hz, NH), 5.05 (d, 1
H, J1,2 7.9 Hz, H-1d), 4.55 (d, 1 H, J1,2 = 3.1 Hz,
H-1f), 4.43 (dd, 1 H, Jgem 12.5, J8.9, 2.4 Hz, H-90),
3.80 (s, 3 H, COOMe), 3.33 (s, 3 H, MeO), 2.42
(dd, 1 H, Jgem 12.8, J3eq,4, 4.6 Hz, H-3eeq), 2.21,
2.06, 1.92, 1.84 (4 s, 12 H, 4 AcO), 1.70, 1.53 (2 s,
6 H, 2 AcN), 1.17 (d, 3 H, J5,6 5.7 Hz, H-6f),
0.94 (m, 2 H, Me3SiCH2CH2), Anal. Calcd for
C72H104N2O39Si (1649.69): C, 52.42; H, 6.35; N,
1.70. Found: C, 52.28; H, 6.16; N, 1.65.
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,
8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-2,4-di-O-benzoyl-
6-deoxy-6-tri¯uoroacetamido-b-D-galactopyranosyl-
(1!4)-[2,3,4-tri-O-benzyl-a-L-fucopyranosyl-(1!3)]-
2-acetamido-6-O-benzyl-2-deoxy-b-D-glucopyrano-
syl-(1!3)-2,4,6-tri-O-benzyl-b-D-galactopyranosyl-
(1!4)-2,3,6-tri-O-benzyl-b-D-glucopyranoside (18).
ÐTo a solution of 17 (135 mg, 52 ꢃmol) in ben-
zene (1.5 mL) were added triphenylphosphine
(55.3 mg, 0.21 mmol) and water (0.1 mL), and the
mixture was stirred for 5.5 h at room temperature.
The mixture was concentrated, and the residue was
treated with tri¯uoroacetic anhydride (30 ꢃL,
0.21 mmol) in CH2Cl2 (2 mL) and pyridine
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,
9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-6-acetamido-2,4-
di-O-benzoyl-6-deoxy-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 (20).ÐA solution of
17 (49.0 mg, 19 ꢃmol) in HOAc (2 mL), EtOH
(2 mL) and Ac2O (0.2 mL) was hydroꢀgenated over
10% Pd±C (80 mg) for 48 h at 40 C, then ®l-
tered and concentrated. Column chromatography
(12:1 CH2Cl2±MeOH) of the residue on silica gel
gave 20 (18.1 mg, 57%) as an amorphous mass:
[ꢀ]D 14.3ꢀ (c 0.3, 1:1 CHCl3±MeOH); IR (®lm)
ꢀ
(0.4 mL) for 6 h at 0 C. After completion of the
reaction, the mixture was diluted with CH2Cl2
(50 mL) successively washed with M Na2CO3, 2 M
HCl and water, dried (Na2SO4), and concentrated.
Column chromatography of the residue on silica
gel gave 18 (63.5 mg, 46%) as an amorphous mass:
1
3550, 3350, 2950, 1750, 1680, 860, 840, 700 cm ;
1H NMR (CD3OD): ꢂ 7.44±8.21 (m, 10 H, 2 Ph),
5.67 (m, 1 H, H-8), 5.24 (d, 1 H, J5,NH 9.8 Hz, NH),

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
525
5.08 (d, 1 H, J1,2 7.7 Hz, H-1d), 4.41 (dd, 1 H, Jgem
12.4, J8.9 2.4 Hz, H-90) 3.78 (s, 3 H, COOMe), 2.39
[2,3,4-tri-O-benzyl-a-L-fucopyranosyl-(1!3)]-2-
acetamido-6-O-benzyl-2-deoxy-b-D-glucopyranosyl-
(1!3)-2,4,6-tri-O-benzyl-b-D-galactopyranosyl-
(1!4)-2,3,6-tri-O-benzyl-b-D-glucopyranoside (23).Ð
To a solution of 17 (46.1 mg, 18 ꢃmol) in MeOH
(2 mL) and THF (1 mL) was added NaOMe
(10ꢀmg), and the mixture was stirred for 48 h at
40 C, then water (0.5 mL) was added. After com-
pletion of the reaction (24 h), the mixture was neu-
tralized with Amberlite IR-120 (H+) resin and
®ltered. The resin was washed with MeOH, and
combined ®ltrate and washings was concentrated.
Column chromatography (5:1 CH2Cl2±MeOH) of
the residue on silica gel gave 23 (35.2 mg, 91%) as
an amorphous mass: [ꢀ]D 29.1ꢀ (c 0.7, 1:1 CHCl3±
MeOH); IR (®lm) 3550, 3350, 2950, 2130, 1750,
1680, 860, 840, 700 cm ; H NMR (CD3OD): ꢂ
6.92±7.51 (m, 50 H, 10 Ph), 5.47 (d, 1 H, J5,NH
10.1 Hz, NH), 2.47 (dd, 1 H, Jgem 12.9, J3eq,4
4.6 Hz, H-3eeq), 2.01, 1.68 (2 s, 6 H, 2 AcN), 1.14
(d, 3 H, J5,6 5.7 Hz, H-6f), 0.97 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C118H143N5O32Si
(2171.53): C, 65.27; H, 6.64; N, 3.23. Found: C,
65.19; H, 6.62; N, 3.22.
0
(dd, 1 H, Jgem 2.8, J3eq,4, 1.1 Hz, H-3eeq) 2.06, 2.00,
1.95, 1.92 (4 s, 12 H, 4 AcO), 1.81, 1.70, 1.42 (3 s, 9
H, 3AcN), 1.26 (d, 3 H, J5,6 5.3 Hz, H-6f), 0.89
(m, 2 H, Me3SiCH2CH2), Anal. Calcd for
C73H105N3O39Si (1676.71): C, 52.29; H, 6.31; N,
2.51. Found: C, 52.03; H, 6.02; N, 2.41.
2-(Trimethylsilyl)ethyl 5-acetamido-3,5-dideoxy-
D-glycero-a-D-galacto-2-nonulopyranosylonic acid-
(2!3)-6-O-methyl-b-Dgalactopyranosyl-(1!4)-
[a-L-fucopyranosyl(1!3)]-2-acetamido-2-deoxy-
b-D-glucopyranosyl-(1!3)-b-D-galactopyranosyl-
(1!4)-b-D-glucopyranoside (21).ÐTo a solution
of 19 (33.2 mg, 20 ꢃmol) in MeOH (2 mL) and
THF (1 mL) was added NaOMe (10 mg), and the
mixture was stirred for 48 h at 40 ꢀC, and for
another 24 h after addition of water (0.5 mL). The
mixture was neutralized with Amberlite IR-120
(H+) resin and ®ltered. The resin was washed with
MeOH, and the combined ®ltrate and washings
were concentrated. Column chromatography
(MeOH) of the residue on Sephadex LH-20 (40 g)
gave 22 (25.1 mg, 99%) as an amorphous mass:
1
1
ꢀ
1
[ꢀ]D 29.4 (c 0.5, MeOH); H NMR (CD3OD): ꢂ
3.35 (s, 3 H, MeO), 2.38 (dd, 1 H, Jgem 12.7, J3eq,4
4.6 Hz, H-3eeq), 1.98, 1.94 (2 s, 6 H, 2 AcN), 1.26
(d, 3 H, J5,6 5.7 Hz, H-6f), 0.97 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C49H86N2O33Si
(1259.29): C, 46.74; H, 6.88; N, 2.22. Found: C,
46.71; H, 6.85; N, 2.09.
2-(Trimethylsilyl)ethyl 5-acetamido-3,5-dideoxy-
D-glycero-a-D-galacto-2-nonulopyranosylonic acid-
(2!3)-6-amino-6-deoxy-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 (24).ÐA solution of
21 (29.8 mg, 14 ꢃmol) in HOAc (2 mL) was hydro-
ꢀ
2-(Trimethylsilyl)ethyl 5-acetamido-3,5-dideoxy-
D-glycero-a-D-galacto-2-nonulopyranosylonic acid-
(2!3)-6-acetamido-6-deoxy-b-D-galactopyranosyl-
(1!4)-[a-L-fucopyranosy-(1!3)]-2-acetamido-2-
deoxy-b-D-glucopyranosyl-(1!3)-b-D-galactopyrano-
syl-(1!4)-b-D-glucopyranoside (22).ÐTo a solution
of 20 (20.1 mg, 12 ꢃmol) in MeOH (2 mL) and
THF (1 mL) was added NaOMe (10 mg), and the
mixture was stirred for 48 h at 40 ꢀC, and for
another 24 h after addition of water (0.5 mL).
Workup as described for 21 gave 22 (14.9 mg,
98%) as an amorphous mass: [ꢀ]D 22.1ꢀ (c 0.2,
genated over Pd(OH)2 (150 mg) for 48 h at 40 C,
then ®ltered and concentrated. Column chromato-
graphy (MeOH) of the residue on Sephadex LH-20
(40 g) gave 24 (13.1 mg, 77%) as an amorphous
1
mass: [ꢀ]D 22.7 (c 0.2, MeOH); H NMR (CD3
OD): ꢂ 2.39 (dd, 1 H, Jgem 12.5, J3eq,4 4.2 Hz, H-
3eeq), 1.96, 1.95 (2 s, 6 H, 2 AcN), 1.18 (d, 3 H, J5,6
6.4 Hz, H-6f), 0.93 (m, 2 H, Me3SiCH2CH2). Anal.
Calcd for C48H85N3O32Si (1244.28): C, 46.33; H,
6.89; N, 3.38. Found: C, 46.09; H, 6.79; N, 3.24.
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,
9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-2,4-di-O-benzoyl-
6-O-methyl-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)-2,3,6-tri-
O-acetyl-b-D-glucopyranoside (25).ÐA solution of
16 (86.8 mg, 34 ꢃmol) in EtOH (2 mL) and HOAc
(2 mL) was hyꢀdrogenated over Pd(OH)2 (90 mg)
for 24 h at 40 C, then ®ltered and concentrated.
1
MeOH); H NMR (CD3OD): ꢂ 5.02 (d, 1 H, J5,NH
0.1 Hz, NH), 1.98, 1.95, 1.94 (3 s, 9 H, 3 AcN), 1.19
(d, 3 H, J5,6 5.6 Hz, H-6f), 0.94 (m, 2 H,
Me3SiCH2CH2. Anal. Calcd for C50H87N3O33Si
(1286.32): C, 46.69; H, 6.82; N, 3.27. Found: C,
46.59; H, 6.54; N, 3.02.
2-(Trimethylsilyl)ethyl 5-acetamido-3,5-dideoxy-
D-glycero-a-D-galacto-2-nonulopyranosylonic acid-
(2!3)-6-azido-6-deoxy-b-D-galactopyranosyl-(1!4)-

526
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
ꢀ
The residue was acetylated with Ac2O (0.5 mL) and
pyridine (3 mL) for 24 h at room temperature. The
solution was diluted with CH2Cl2 (50 mL), and the
solution was washed with 2 M HCI and water,
dried (Na2SO4), and concentrated. Column chro-
matography (40:1 CH2Cl2±MeOH) of the residue
on silica gel gave 25 (54.9 mg, 78%) as an amor-
phous mass: [ꢀ]D 15.6ꢀ (c 2.3, CHCl3); IR (®lm)
3350, 2950, 1750, 1680, 860, 840, 700 cm ; H
NMR (CDCl3): ꢂ 7.41±8.22 (m, 10 H, 2 Ph), 5.63
(m, 1 H, H-8), 3.88 (s, 3 H, COOMe), 3.37 (s, 3 H,
MeO), 2.39 (dd, 1 H, Jgem 12.2, J3eq,4 4.6 Hz, H-
3eeq), 1.55±2.22 (16 s, 48 H, 2 AcN, 14 AcO), 1.18
(d, 3 H, J5,6 6.6 Hz, H-6f), 0.94 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C92H124N2O49Si
(2070.06): C, 53.38; H, 6.04; N, 1.35. Found: C,
53.17; H, 5.86; N, 1.28.
Pd(OH)2 (65 mg) for 24 h at 40 C, then ®ltered
and concentrated. The residue was acetylated with
Ac2O (1 mL) and pyridine (1 mL) for 24 h at room
temperature. Workup as described for 25 gave 26
(41.5 mg, 80%) as an amorphous mass: [ꢀ]D 16.3ꢀ
(c 1.0, CHCl3); IR (®lm) 3350, 2950, 1750, 1680,
860, 840, 700 cm ; 1H NMR (CDCl3): ꢂ 7.28±8.18
1
(m, 10 H, 2 Ph),5.61 (m, 1 H, H-8), 3.78 (s, 3 H,
COOMe), 2.38 (dd, 1 H, Jgem 12.5, J3eq,4 4.6 Hz, H-
3eeq), 1.43±2.18 (16 s, 48 H, 2 AcN, 14 AcO), 1.17
(d, 3 H, J5,6 6.7 Hz, H-6f), 0.91 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C93H122F3N3-
O49Si (2151.05): C, 51.93; H, 5.72; N, 1.95. Found:
C, 51.76; H, 5.67; N, 1.69.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-
dideoxy-D-glycero-a-D-galacto-2-nonulopyranosyl-
onate-(2!3)-2,4-di-O-benzoyl-6-O-methyl-b-D-gal-
actopyranosyl-(1!4)-[2,3,4-tri-O-acetyl-a-L-fuco-
pyranosyl-(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)-2,3,6-tri-O-acetyl-b-D-gluco-
pyranosyl trichloroacetimidate (28).ÐTo a solution
of 25 (97.3 mg, 47 ꢃmol) in CH2Cl2 (1.2 mL),
1
1
2-(Trimethylsilyl)ethyl (methyl 5-acetamido-4,7,8,
9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate)-(2!3)-6-acetamido-2,4-
di-O-benzoyl-6-deoxy-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)-
ꢀ
cooled to 0 C, was added CF3COOH (0.7 mL),
2,3,6-tri-O-acetyl-b-D-glucopyranoside
(26).ÐA
and the mixture was stirred for 3 h at room tem-
perature and concentrated. The product was pur-
i®ed by column chromatography on silica gel with
35:1 CH2Cl2±MeOH to give the 1-hydroxy com-
pound (83.0 mg). To a solution of the residue in
CH2Cl2 (1 mL) cooled to 0 C were added tri-
chloroacetonitrile (0.13 mL, 1.3 mmol) and DBU
(6.3 ꢃL, 42 ꢃmol). The mixture was stirred for 2 h
solution of 17 (191 mg, 75 ꢃmol) in EtOH (3 mL),
HOAc (3 mL) and Ac2O (0.5 mL) was hydro-
ꢀ
genated over Pd(OH)2 (200 mg) for 48 h at 40 C,
then ®ltered and concentrated. The residue was
acetylated with Ac2O (1 mL) and pyridine (2 mL)
for 24 h at room temperature. Workup as described
for 25 gave 26 (121 mg, 77%) as an amorphous
mass: [ꢀ]D 16.4ꢀ (c 2.4, CHCl3); IR (®lm) 3350,
ꢀ
ꢀ
at 0 C, and the progress of the reaction was mon-
1
1
2950, 1750, 1680, 860, 840, 700 cm ; H NMR
(CDCl3): ꢂ 7.41±8.22 (m, 10 H, 2 Ph), 5.67 (m, 1 H,
H-8), 4.48 (d, 1 H, J1,2 3.4 Hz, H-1f), 3.88 (s, 3 H,
COOMe), 2.36 (dd, 1 H, Jgem 12.7, J3eq,4 4.7 Hz, H-
3eeq), 1.58±2.22 (17 s, 51 H, 3 AcN, 14 AcO), 1.19
(d, 3 H, J5,6 6.6 Hz, H-6f), 0.93 (m, 2 H,
Me3SiCH2CH2). Anal. Calcd for C93H125N3O49Si
(2097.08): C, 53.27; H, 6.01; N, 2.00. Found: C,
53.25; H, 5.81; N, 1.94.
itored by TLC. After completion of the reaction,
the mixture was concentrated. Column chromato-
graphy (30:1 CH2Cl2±MeOH) of the residue on
silica gel gave 28 (80.1 mg, 81%) as an amorphous
mass: [ꢀ]D +10.2ꢀ (c 0.8, CHCl3); IR (®lm) 3350,
2950, 1750, 1680, 700 cm ; H NMR (CDCl3): ꢂ
8.65 (s, 1 H, C=NH), 7.28±8.22 (m, 10 H, 2 Ph),
6.43 (d, 1 H, J1,2 3.5 Hz, H-1a), 5.68 (m, 1 H, H-8),
3.88 (s, 3 H, COOMe), 3.37 (s, 3 H, MeO), 2.39
(dd, 1 H, Jgem 12.6, J3eq,4 4.5 Hz, H-3eeq), 1.59±
2.24 (16 s, 48 H, 2 AcN, 14 AcO), 1.18 (d, 3 H, J5,6
6.3 Hz, H-6f). Anal. Calcd for C89H112Cl3N3O49Si
(2114.21): C, 50.56; h, 5.34; N, 1.99. Found: C,
50.30; h, 5.28; N, 1.80.
1
1
2-(Trimethylsilyl)ethyl methyl 5-acetamido-4,7,8,
9-tetra-O-acetyl-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonate-(2!3)-2,4-di-O-benzoyl-6-
deoxy-6-tri¯uoroacetamido-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-glucopyrano-
syl-(1!3)-2,4,6-tri-O-acetyl-b-D-galactopyranosyl-
(1!4)-2,3,6-tri-O-acetyl-b-D-glucopyranoside (27).Ð
A solution of 18 (63.5 mg, 24 ꢃmol) in EtOH
(2 mL) and HOAc (2 mL) was hydrogenated over
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-
dideoxy-D-glycero-a-D-galacto-2-nonulopyranosyl-
onate-(2!3)-6-acetamido-2,4-di-O-benzoyl-6-deoxy
-b-D-galactopyranosyl-(1!4)-[2,3,4-tri-O-acetyl-a-
L-fucopyranosyl-(1!3)]-2-acetamido-6-O-acetyl-2-

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
527
deoxy-b-D-glucopyranosyl-(1!3)-2,4,6-tri-O-acetyl-
b-D-galactopyranosyl-(1!4)-2,3,6-tri-O-acetyl-b-D -
glucopyranosyl trichloroacetimidate (29).ÐSelective
removal of the 2-(trimethylsilyl)ethyl group in 26
(62.1 mg, 30 ꢃmol) with CF3COOH (0.7 mL) in
CH2Cl2 (1 mL) for 2 h at room temperature, and
subsequent reaction of the product with tri-
chloroacetonitrile (83 ꢃL, 0.83 mmol) in CH2Cl2
(1 mL) in the presence of DBU (4.1 ꢃl, 27 ꢃmol)
for 2 h at 0 ꢀC as described for 28, gave 29 (56.5 mg,
88%) as an amorphous mass: [ꢀ]D +1.9ꢀ (c 1.1,
CHCl3); IR (®lm) 3350, 2950, 1750, 1680,
actopyranosyl-(1!4)-(2,3,6-tri-O-acetyl-b-D-gluco-
pyranosyl)-(1!1)-(2S,3R,4E)-2-azido-3-O-(tert-
butyldiphenylsilyl)-4-octadecene-1,3-diol (32).ÐTo
a
solution of 28 (37.9 mg, 18 ꢃmol) and
(2S,3R,4E)-2-azido-3-O-(tert-butyldiphenylsilyl)-4-
octadecene-1,3-diol (31; 20.2 mg, 36 ꢃmol) in dry
CH2Cl2 (0.5 mL) was added 4A MS (AW-300,
300 mg), and the mixture was stirred for 12 h at
room temperature, then cooled to 0 ꢀC. Tri-
methylsilyl tri¯uoromethanesulfonate (TMSOTf,
1 ꢃL, 5.2 ꢃmol) waꢀs added, and the mixture was
stirred for 24 h at 0 C. The solids were ®ltered o€
and washed with CH2Cl2. The combined ®ltrate
and washings were washed with M Na2CO3 and
water, dried (Na2SO4) and concentrated. Column
chromatography of the residue on silica gel gave 30
(14.8 mg, 33%) as an amorphous mass: [ꢀ]D 10.5ꢀ
(c 0.7, CHCl3); IR (®lm) 3350, 2950, 2130, 1750,
1
700 cm ; 1H NMR (CDCl3): ꢂ 8.65 (s, 1 H,
C=NH), 7.28±8.20 (m, 10 H, 2 Ph), 6.47 (d, 1 H,
J1,2 3.7 Hz, H-1a), 5.62 (m, 1 H, H-8), 5.19 (dd, J6,7
2.5 Hz, J7,8 8.4 Hz, H-7), 3.86 (s, 3 H, COOMe),
2.34 (dd, 1 H, Jgem 12.8, J3eq,4 4.5 Hz, H-3eeq),
1.88±2.21 (17 s, 51 H, 3 AcN, 14 AcO), 1.19 (d, 3
H, J5,6 6.4 Hz, H-6f). Anal. Calcd for
C90H113Cl3N4O49Si (2141.23): C, 50.48; H, 5.32; N,
2.62. Found: C, 50.47; H, 5.27; N, 2.36.
1
1680, 700 cm ; 1H NMR (CDCl3): ꢂ 7.16±8.21 (m,
20 H, 4 Ph), 3.81 (s, 3 H, COOMe), 3.39 (s, 3 H,
MeO), 2.38 (dd, 1 H, Jgem 12.7, J3eq,4 4.5 Hz, H-
3eeq), 1.81±2.22 (16 s, 48 H, 2 AcN, 14 AcO), 1.26
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-
dideoxy-D-glycero-a-D-galacto-2-nonulopyranosyl-
onate-(2!3)-2,4-di-O-benzoyl-6-deoxy-6-tri¯uoro-
acetamido-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)-2,3,6-tri-O-
acetyl-b-D-glucopyranos trichloroacetimidate (30).Ð
Selective removal of the 2-(trimethylsilyl) ethyl
group in 27 (41.1 mg, 19 ꢃmol) with CF3 COOH
(0.7 mL) in CH2Cl2 (1 mL) for 2 h at room tem-
perature, and subsequent reaction of the product
with trichloroacetonitrile (63 ꢃL, 0.63 mmol) in
CH2Cl2 (0.7 mL) in the presence of DBU (2.8 ꢃL,
(s, 22 H, 11 CH2), 0.88 (t, 3 H, JMe,CH 6.8 Hz,
2
MeCH2). Anal. Calcd for C121H143N5O50Si
(2495.55): C, 58.24; h, 5.78; N, 2.81. Found: C,
58.07; H, 5.57; N, 2.74.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-6-acetamido-2,4-di-O-benzoyl-6-deoxy-b-D-
galactopyranosyl-(1!4)-[2,3,4-tri-O-acetyl-a-L-fuco-
pyranosyl-(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)-2,3,6-tri-O-acetyl-b-D-gluco-
pyranosyl-(1!1)-(2S,3R,4E)-2-azido-3-O-(tert-
butyldiphenylsilyl)-4-octadecene-1,3-diol (33).Ð
Coupling of 31 (26.2 mg, 12 ꢃmol) with 29
(13.6 mg, 24 ꢃmol) in CH2Cl2 (0.4 mL) in the pre-
sence of TMSOTf (0.2 ꢃL, 1.0 ꢃmol) and 4A MS
(AW-300, 100 mg) as described for 32 gave 33
(18.2 mg, 58%) as an amorphous mass: [ꢀ]D 27.3ꢀ
(c 0.4, CHCl3); IR (®lm) 3350, 2950, 2130, 1750,
ꢀ
18 ꢃmol) for 1.5 h at 0 C as described for 28 gave
30 (35.2 mg, 84%) as an amorphous mass: [ꢀ]D
+4.9ꢀ (c 0.6, CHCl3); IR (®lm) 3350, 2950, 1750,
1
1
1680, 700 cm ; H NMR (CDCl3): ꢂ 8.65 (s, 1 H,
C=NH), 7.22±8.21 (m, 10 H, 2 Ph), 6.45 (d, 1 H,
J1,2 3.8 Hz, H-1a), 5.60 (m, 1 H, H-8), 3.88 (S, 3
H, COOMe), 2.36 (dd, 1 H, Jgem 12.7, J3eq,4
4.6 Hz, H-3eeq), 1.83±2.20 (16 s, 48 H, 2 AcN, 14
AcO), 1.19 (d, 3 H, J5,6 6.6 Hz, H-6f). Anal. Calcd
for C90H110Cl3F3N4O49Si (2195.20): C, 49.24; H,
5.05; N, 2.55. Found: C, 49.08; H, 4.82; N, 2.55.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-2,4-di-O-benzoyl-6-O-methyl-b-D-galactopyr-
anosyl-(1!4)-[2,3,4-tri-O-acetyl-a-L-fucopyrano-
syl-(1!3)]-2-acetamido-6-O-acetyl-2-deoxy-b-D-
glucopyranosyl-(1!3)-2,4,6-tri-O-acetyl-b-D-gal-
1
1680, 700 cm ; 1H NMR (CDCl3): ꢂ 7.22±8.24 (m,
20 H, 4 Ph), 5.64 (m, 1 H; H-8), 4.49 (d, 1 H, J1,2
7.6 Hz, H-1a), 3.89 (s, 3 H, COOMe), 2.39 (dd, 1
H, Jgem 12.4, J3eq,4 4.6 Hz, H-3eeq), 1.70±2.26 (17 s,
51 H, 3 AcN, 14 AcO), 1.25 (s, 22 H, 11 CH2), 0.88
(t, 3 H, JMe,CH 6.8 Hz, MeCH2). Anal. Calcd for
C122H144N6O50Si (2522.57): C, 58.09; H, 5.75; N,
3.33. Found: C, 57.91; H, 5.56; N, 3.05.
2
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-2,4-di-O-benzoyl-6-deoxy-6-tri¯uoroace-

528
N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
tamido-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)-2,3,6-tri-O-
acetyl-b-D-glucopyranosyl-(1!1)-(2S,3R,4E)-2-
azido-3-O-(tert-butyldiphenylsilyl)-4-octadecene-1,3-
diol (34).ÐCoupling of 30 (26.0 mg, 12 ꢃmol) with
31 (13.3 mg, 23 ꢃmol) in CH2Cl2 (0.4 mL) in the
presence of TMSOTf (0.2 ꢃL, 1.1 ꢃmol) and
4A MS (AW-300, 100 mg) as described for 32 gave
34 (19.8 mg, 65%) as an amorphous mass: [ꢀ]D
17.4 (c 0.3, CHCl3); IR (®lm) 3350, 2950, 2130,
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-2,4-di-O-benzoyl-6-deoxy-6-tri¯uoroacetam-
ido-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-acet-
yl-b-D-galactopyranosyl-(1!4)-(2,3,6-tri-O-acetyl-
b-D-glucopyranosyl)-(1!1)-(2S,3R,4E)-3-O-tert-
butyldiphenylsilyl)-2-octadecamido-4-octadecene-1,3-
diol (36).ÐSelective reduction of the azido group
in 33 (17.9 mg, 7.1 ꢃmol) with triphenylphosphine
(3.7 mg, 14 ꢃmol) in benzene (1 mL) and water
(50 ꢃL), followed by coupling of the product with
octadecanoic acid (6 mg, 21.3 ꢃmol) in the presence
of WSC (1.3 mg, 21.2 ꢃmol) and work up as
described for 35 gave 36 (13.8 mg, 70%) as an
amorphous mass: [ꢀ]D 24.3 (c 0.4, CHCl3); IR
1
1750, 1680, 700 cmq; H NMR (CDCl3): ꢂ 7.23±
8.18 (m, 20 H, 4 Ph), 5.63 (m, 1 H, H-8), 4.40 (d, 1
H, J1,2 7.7 Hz, H-1a), 3.79 (s, 3 H, COOMe), 2.38
(dd, 1 H, Jgem 2.7, J3eq,4 12.7 Hz, H-3eeq), 1.45±
2.18 (16 s, 48 H, 2 AcN, 14 AcO), 1.23 (s, 22 H, 11
1
1
CH2), 0.83 (t, 3 H, JMe,CH 6.8 Hz, MeCH2). Anal.
(®lm) 3350, 2950, 1750, 1680, 700 cm ; H NMR
(CDCl3): ꢂ 7.24±8.26 (m, 20 H, 4 Ph), 3.88 (s, 3 H,
COOMe), 2.36 (dd, 1 H, Jgem 12.4 J3eq,4 4.4 Hz, H-
3eeq), 1.67±2.27 (17 s, 51 H, 3 AcN, 14 AcO), 1.27
2
Calcd for C122H141F3N6O50Si (2576.54): C, 56.87;
H, 5.52; N, 3.26. Found: C, 56.60; H, 5.49; N, 3.20.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-2,4-di-O-benzoyl-6-O-methyl-b-D-galactopyr-
anosyl-(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-galactopyr-
anosyl-(1!4)-(2,3,6-tri-O-acetyl-b-D-glucopyrano-
syl)-(1!1)-(2S,3R,4E)-3-O-tert-butyldiphenylsilyl)-
2-octadecamido-4-octadecene-1,3-diol (35).ÐTo a
solution of 32 (14.8 mg, 5.9 ꢃmol) in benzene
(0.6 mL) and water (25 ꢃL) was added triphenyl-
phosphine (4.6 mg, 18 mmol), and the mixture was
stirred for 24 h with the progress of the reaction
being monitored by TLC. The mixture was con-
centrated, and the residue was stirred with octade-
canoic acid (5.0 mg, 18 ꢃmol) and 1-(3-dimethyl-
aminopropyl)-3-ethylcarbodiimide hydrochloride
(WSC, 3.4 mg, 18 ꢃmol) in dry CH2Cl2 (0.8 mL)
for 24 h at room temperature. The mixture was
diluted with CH2Cl2 (25 mL), washed with water,
dried (Na2SO4) and concentrated. Column chro-
matography (20:1 CH2Cl2±MeOH) of the residue
gave 35 (13.1 mg, 81%) as an amorphous mass:
[ꢀ]D 6.1ꢀ (c 0.2, CHCl3); IR (®lm) 3350, 2950,
(s, 52 H, 26 CH2), 0.88 (t, 6 H, JMe,CH 6.8 Hz, 2
2
MeCH2). Anal. Calcd for C140H200N4O51Si
(2783.20): C, 60.42; H, 7.24; N, 2.01. Found: C,
60.34; H, 7.17; N, 1.92.
Methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dide-
oxy-D-glycero-a-D-galacto-2-nonulopyranosylonate-
(2!3)-2,4-di-O-benzoyl-6-deoxy-6-tri¯uoroacetam-
ido-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-acet-
yl-b-D-galactopyranosyl-(1!4)-(2,3,6-tri-O-acetyl-
b-D-glucopyranosyl-(1!1)-(2S,3R,4E)-3-O-tert-
butyldiphenylsilyl)-2-octadecamido-4-octadecene-1,3-
diol (37).ÐSelective reduction of azido group in 34
(16.7 mg, 6.5 ꢃmol) with triphenylphosphine
(5.1 mg, 19 ꢃmol) in benzene (0.6 mL) and water
(50 ꢃL), followed by coupling of the product with
octadecanoic acid (5.5 mg, 19 ꢃmol) in the presence
of WSC (3.7 mg, 19 ꢃmol) and workup as descri-
bed for 35, gave 37 (8.8 mg, 48%) as an amorphous
mass: [ꢀ]D 11.4ꢀ (c 0.2, CHCl3); IR (®lm) 3350,
1
1
2950, 1750, 1680, 700 cm ; H NMR (CDCl3): ꢂ
7.24±8.17 (m, 20 H, 4 Ph), 3.78 (s, 3 H, COOMe),
2.38 (dd, 1 H, Jgem 2.7, J3eq,4 4.4 Hz, H-3eeq), 1.41±
2.17 (16 s, 48 H, 2 AcN, 14 AcO), 1.24 (s, 52 H, 26
1
1
1750, 1680, 700 cm ; H NMR (CDCl3): ꢂ 7.29±
8.19 (m, 20 H, 4 Ph), 3.84 (s, 3 H, COOMe), 3.39
(s, 3 H, MeO), 2.35 (dd, 1 H, Jgem 12.8, J3eq,4 4.4 Hz,
H-3eeq),1.82±2.23 (16 s, 48 H, 2 AcN, 14 AcO),
CH2), 0.83 (t, 6 H, JMe,CH 6.8 Hz, 2 MeCH2).
2
Anal. Calcd for C140H197F3N4O51Si (2837.17): C,
59.27; H, 7.00; N, 1.97. Found: C, 59.03; H, 6.89;
N, 1.69.
5-Acetamido-3,5-dideoxy-D-glycero-a-D-galacto-
2-nonulopyranosylonic acid-(2!3)-6-O-methyl-b-
1.25 (s, 52 H, 26 CH2), 0.89 (t, 6 H, JMe,CH 6.9 Hz,
2
2 MeCH2), Anal. Calcd for C139H199N3O51Si
(2756.18): C, 60.57; H, 7.28; N, 1.52. Found: C,
60.39; H, 7.15; N, 1.41.

N. Otsubo et al./Carbohydrate Research 306 (1998) 517±530
529
D-galactopyranosyl-(1!4)-[a-L-fucopyranosyl-(1!3)]-
2-acetamido-b-D-glucopyranosyl-(1!3)-b-D-galacto-
pyranosyl-(1!4)-b-D-glucopyranosyl-(1!1)-(2S,3R,
4E)-2-octadecamido-4-octadecene-1,3-diol (38).Ð
To a solution of 35 (8 mg, 2.9 ꢃmol) in MeCN
(0.5 mL), cooled to 0 ꢀC, was added M tetra-
butylammonium ¯uoride (TBAF, 80 ꢃL), and the
mixture was stirred for 24 h at room temperature,
then concentrated. The residue was treated with
NaOMe (10 mg) in MeOH (0.5 mL) and THF
(0.1 mL) for 24 h at 40 ꢀC, then water (0.1 mL) was
added. After completion of the reaction (24 h), the
mixture was neutralized with Amberlite IR-120
(H+) resin and ®ltered. The resin was washed
with MeOH, and the combined ®ltrate and wash-
ings were concentrated. Column chromatography
(5:4 CHCl3±MeOH) of the residue on Sephadex
LH-20 (40 g) gave 38 (5.1 mg, 98%) as an amor-
phous mass: [ꢀ]D 14.3ꢀ (c 0.1, 1:1 CHCl3±MeOH);
1H NMR (CDCl3): ꢂ 3.35 (s, 3 H, MeO), 2.01 (2 s,
6 H, 2 AcN), 1.91, 1.26 (s, 52 H, 26 CH2), 0.88 (t, 6
(80 ꢃL), and the mixture was stirred for 24 h at
room temperature, then concentrated. The residue
was treated with NaOMe (10 mg) in MeOH
(0.5 mL) and THF (0.1 mL) for 24 h at 40ꢀC, then
water (0.1 mL) was added. Workup as described
for 38 gave 40 ꢀ(5.2 mg, 93%) as an amorphous
1
mass: [ꢀ]D 9.1 (c 0.1, 1:1 CHCl3±MeOH); H
NMR (CDCl3): ꢂ 1.98, 1.61 (2 s, 6 H, 2 AcN), 1.29
(s, 52 H, 26 CH2), 0.83 (t, 6 H, JMe,CH 6.8 Hz, 2
2
MeCH2). Anal. Calcd for C89H146N4O34 (1816.14):
C, 58.86; H, 8.16; N, 3.09. Found: C, 58.85; H,
8.06; N, 3.02.
Acknowledgements
This work was supported in part by Grant-in-
Aid Scienti®c Research on Priority Areas (No.
09240101) from the Ministry of Education, Science
and Culture of Japan.
H, JMe,CH 7.1 Hz, 2 MeCH2). Anal. Calcd for
C90H147N3O35 (1831.15): C, 59.03; H, 8.09; N,
2.29. Found: C, 58.99; H, 8.02; N, 2.14.
2
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