Synthesis of Neu5Ac- and Neu5Gc-α-(2 → 6′)-lactosamine 3-aminopropyl glycosides

Article abstract of DOI:10.1016/S0008-6215(01)00002-7

In order to prepare 3-aminopropyl glycosides of Neu5Ac-α-(2 → 6′)-lactosamine trisaccharide 1, and its N-glycolyl containing analogue Neu5Gc-α-(2 → 6′)-lactosamine 2, a series of lactosamine acceptors with two, three, and four free OH groups in the galact

Full text of DOI:10.1016/S0008-6215(01)00002-7

www.elsevier.nl/locate/carres
Carbohydrate Research 330 (2001) 445–458
Synthesis of Neu5Ac- and Neu5Gc-a-(2“6%)-lactosamine
3-aminopropyl glycosides
Andrei A. Sherman, Olga N. Yudina, Alexander S. Shashkov, Vladimir M. Menshov,
Nikolay E. Nifant’ev*
N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Leninsky Prospect 47,
Moscow B-334, Russia
Received 6 October 2000; accepted 20 December 2000
Abstract
In order to prepare 3-aminopropyl glycosides of Neu5Ac-a-(2“6%)-lactosamine trisaccharide 1, and its N-glycolyl
containing analogue Neu5Gc-a-(2“6%)-lactosamine 2, a series of lactosamine acceptors with two, three, and four free
OH groups in the galactose residue was studied in glycosylations with a conventional sialyl donor phenyl [methyl
5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-thio-
and a new donor phenyl [methyl 4,7,8,9-tetra-O-acetyl-5-(N-tert-butoxycarbonylacetamido)-3,5-dideoxy-2-thio-
glycero-a- and b- -galacto-2-nonulopyranosid]onates (4), respectively. The lactosamine 4%,6%-diol acceptor was found
D
-glycero-a- and b-
D
-galacto-2-nonulopyranosid]onates (3)
D
-
D
to be the most efficient in glycosylation with both 3 and 4, while imide-type donor 4 gave slightly higher yields with
all acceptors, and isolation of the reaction products was more convenient. In the trisaccharides, obtained by
glycosylation with donor 4, the 5-(N-tert-butoxycarbonylacetamido) moiety in the neuraminic acid could be
efficiently transformed into the desired N-glycolyl fragment, indicating that such protected oligosaccharide deriva-
tives are valuable precursors of sialo-oligosaccharides containing N-modified analogues of Neu5Ac. © 2001 Elsevier
Science Ltd. All rights reserved.
Keywords: N-Acetylneuraminic acid; N-Glycolylneuraminic acid; Sialyllactosamine
1. Introduction
3-oxapentyl spacer, has been described re-
cently.2
Sialo-oligosaccharides play an important
role in the process of cell recognition and act
as receptors of various lectins.1 In this paper
we describe the synthesis of 5-N-acetyl-sialyl-
a-(2“6%)-lactosamine 3-aminopropyl gly-
coside (1) and its N-glycolyl analogue 2 to be
further used as models for the investigations
of carbohydrate specificity of sialyl-binding
lectins — siglecs. Synthesis of a sialyl-a-(2“6%)-
lactosamine derivative, but with 5-amino-
The reported syntheses of sialo-oligosaccha-
rides containing N-modified analogues of
Neu5Ac used neuraminic acid donors in
which the desired N-acyl fragment was al-
ready present.3–5 Alternatively, Ogawa et al.6
used the temporary N-Boc protected sialyl-
donor 5 which allowed varying the acyl moi-
ety at the nitrogen atom of the neuraminic
acid residue after assembling the oligosaccha-
ride backbone.
Recently it was reported7,8 that bis N-acety-
lated imide-type sialyl-donor 6 was signifi-
cantly more reactive and gave higher yields in
glycosylations in comparison with its amide-
* Corresponding author. Fax: +7-095-1358784.
E-mail address: nen@ioc.ac.ru (N.E. Nifant’ev).
0008-6215/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(01)00002-7

446
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
type counterpart 7. Furthermore, no forma-
tion of glycal 12 was observed during glycosy-
lation with the imide donor 6, thus the reac-
tion could be performed with a 1:1 donor–ac-
ceptor ratio. This is in sharp contrast to the
traditionally used conditions by Hasegawa et
al.3,9 in which an approximately twofold ex-
cess of amide donor 3 or 7 is employed and
glycal 11 is a permanent byproduct.
et al.3 sialylation with methylthio donor 7 of
2-trimethylsilylethyl 3-O-benzoyl-b- -galac-
D
topyranoside afforded 70% yield of a-(2“6)-
linked disaccharide solely, while glycosylation
of its 3-O-benzyl counterpart gave approxi-
mately the same yield of a/b mixtures in the
ratio from 3:1 with DMTST as promoter to
2:1 with NIS–TfOH. We employed diol 8,
triol 9 and tetraol 10 in the present study, and
This finding prompted us to investigate a
new imide-type sialyl donor 4, carrying Boc
and Ac moieties at the neuraminic acid nitro-
gen. The donor 4 was chosen because its
application opens ready access to Neu5acyl-
modified analogues of 6%-sialyl-lactosamine via
selective N-deacylation10 of the mixed Ac–N–
Boc imide into the corresponding H–N–Boc
carbamate, subsequent Boc group removal,
and acylation. Similar methodology has al-
ready been applied for the synthesis of (1“5)-
amide-linked amino acid dimers derived from
sialic acids.11 It is worth noting that this ap-
proach seems particularly attractive because it
allows varying the acyl substitutent after as-
sembling the oligosaccharide backbone.
The second aspect which was specially in-
vestigated in this work was evaluation of lac-
tosamine acceptor properties in glycosylation
with donors 3 and 4. A systematic study of the
influence of acceptor structure on the effi-
ciency and stereoselectivity of its sialylation is
currently available for monosaccharide galac-
tose derivatives only: according to Hasegawa
the 3%-O-benzoyl protection in compounds 8
and 9 was chosen based on the above men-
tioned assumption that it would contribute to
a-selectivity.
2. Results and discussion
Bis N-acylated donor 4 was prepared in
85% yield by treatment of 312 with (Boc)2O
and DMAP in refluxing THF.13 Other condi-
tions tested10,13 led to incomplete conversion
of the starting material 3 and afforded 4 in
significantly lower yield. It is worth noting
that the reaction product 4 could not be sepa-
rated from the dark-colored non-carbohydrate
impurities by silica gel column chromatogra-
phy, and thus additional purification by gel-
permeation chromatography on the Bio-Beads
SX-3 in toluene was required. Compound 4
was formed as an inseparable 4:1 mixture of
isomers at the nitrogen atom (NMR data) and
their ratio remained unchanged during the

A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
447
storage at room temperature in either CDCl3
solution or in solid state.
occurred. Thus treatment of 15 with
Me3N·BH3 (4 equiv), AlCl3 (6 equiv), and
water (2 equiv) afforded within hours the 6-O-
Bn derivative 16 in 87% yield. Location of the
benzyloxy group at C-6 in 16 was confirmed
by the downfield shift of this carbon atom
(70.9 ppm) and shielding of the C-4 signal
from 77.6 in 15 to 72.2 ppm. We suppose that
the protic acid formed from AlCl3 and water
is a much more powerful reagent rather than
the equivalent Lewis acid. Other examples of
benzylidene ring opening under such condi-
tions will be published elsewhere.
Initially we designed compound 19 as the
key intermediate for the preparation of lac-
tosamine sialyl acceptors 8, 9, and 10. Appli-
cation of 19 allows ready access to diol 8 via
debenzylidenation, tetraol 10 via further O-
deacylation of 8, and triol 9 by treatment of
19 with methanolic hydrogen chloride under
the conditions of acid-catalyzed deacetyl-
ation.16
Condensation of 16 with thiogalactoside 18,
prepared by acetylation of 17, promoted with
CuBr2–AgOTf–Bu4NBr in MeNO217 gave lac-
tosamine 8, isolated after debenzylidenation of
19, in 37% yield (Scheme 1). The b configura-
tion of the Gal residue in 8 was deduced from
the value of J1%,2% coupling constant 8 Hz, and
no a anomer was formed. The presence of the
AcO group at C-2% position and BzO group at
The synthesis of lactosamine acceptors 8, 9,
and 10 was initiated (Scheme 1) from benzyla-
tion of the known spacer-armed glucosamine
derivative 14.14 In this reaction the excess of
NaH was used in order to convert the
HNC(O)CF3 functionality into amide anion
NaNC(O)CF3 and thereby protect the base-
liable trifluoroacetamido group. The trianion
formed upon treatment of 14 with excess of
NaH was regioselectively O-alkylated with
benzyl bromide at −15 °C and then proto-
nated with acetic acid to give 15 in 90% yield.
The use of only a small excess of benzyl
bromide was essential in order to avoid fur-
ther undesired N-alkylation.
At the next stage the benzylidene acetal ring
in compound 15 was subjected to reductive
opening by treatment with Me3N·BH3 and
anhydrous AlCl3 in dry THF under Ar at
room temperature, i.e. following the known
protocol.15 However, after 24 h the unchanged
starting material 15 and only traces (ꢀ10%)
of target 16 were present in the reaction mix-
ture and no further conversion was observed
in another 24 h. The use of large (ꢀ20-fold)
excess of the reagents did not improve the
yield of 16, while degradation took place upon
heating. Subsequently we found that in the
presence of water, mild reductive ring opening
Scheme 1. Reagents and conditions: (i) Ac2O, Py; (ii) NaH (5 equiv), BnBr (1.2 equiv), DMF, −15 °C, then AcOH; (iii) anhyd
AlCl3 (6 equiv), Me3N·BH3 (4 equiv), water (2 equiv), dry THF, rt; (iv) CuBr2, AgOTf, Bu4NBr, MeNO2, MS-4A, rt; (v) 90% aq
CF3CO2H, CH2Cl2; (vi) AgOTf, CH2Cl2, MS-4A, rt; (vii) MeONa, MeOH; (viii) PhCH(OMe)2, TsOH, DMF; (ix) BzCN, Py.

448
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
Table 1
Glycosylation of acceptors 8, 9, and 10 with donors 3 and 4
Entry
Donor
Acceptor
Product
Yield (%)
a/b ratio
1
2
3
4
5
3
3
4
4
3
8
9
8
9
10
24
25
26
27
79
59
84
60
1.3:1
1.2:1
1.3:1
1:1
complex mixture
n.d.
n.d.
C-3% was confirmed by the downfield location
of H-2% (lH-2% 5.33 ppm) and of H-3% (lH-3% 4.79
ppm). Glycosylation of 16 with 18 under
MeOTf promotion was not effective, whereas
with NIS–TfOH gave orthoesters in low yield
as the only coupling product. Similar moder-
ate efficiency of glycosylation with 2,3-di-O-
was investigated. All the reactions were per-
formed with 2 equiv of either donor 3 or 4 in
MeCN at −35 °C in the presence of NIS,
,
TfOH, and molecular sieves MS-3 A, i.e.,
under the conditions recommended by
Hasegawa et al.9 The trisaccharides 24–27,
obtained after silica gel column chromatogra-
phy were used for further synthetic steps as
anomeric mixtures and separated into individ-
ual isomers by C18 reverse-phase column
chromatography after complete removal of
protecting groups. Therefore the anomeric ra-
tios obtained in glycosylation reactions were
estimated from NMR spectra of purified a/b
mixtures 24–27 and proved after final separa-
tion of deblocked oligosaccharides.
The results of glycosylations (Table 1) indi-
cate that better yields of sialyl-lactosamines
were obtained with diol acceptor 8 (Entries 1
and 3), while triol acceptor 9 was less effective
(Entries 2 and 4). Sialylation of tetraol 10 with
3 (Entry 5) afforded a complex mixture of at
least five trisaccharide products without sig-
nificant prevalence of any of them, thus indi-
cating the absence of regioselectivity of the
reaction. With both acceptors 8 and 9, how-
ever, imide-type donor 4 afforded slightly
higher yields, while the stereoselectivity of the
glycosylation was found to be independent of
both donor and acceptor structure.
acetyl-4,6-O-benzylidene-a- -galactopyranosyl
D
trichloroacetimidate or bromide of 5-azido-3-
oxapentyl 2-acetamido-3,6-di-O-benzyl-2-de-
oxy-b-
recently.2
Preparative synthesis (Scheme 1) of lac-
D
-glycopyranoside has been described
tosamines 8, 9, and 10 was performed by the
reaction sequence involved glycosylation of 16
with benzobromogalactose 20 in the presence
of AgOTf to give 21, NaOMe-catalyzed
debenzoylation into 10, 4%,6%-benzylidenation
to produce 22, regiospecific 3%-O-benzoylation
with BzCN18 to afford 23, and hydrolytic re-
moval of benzylidene acetal to give 9. For the
preparation of diol 8, compound 23 was 2%-O-
acetylated into 19 prior to hydrolysis into 8.
The presence of the Bz group in the 3% posi-
tion in compound 9 was confirmed by the
downfield chemical shift of H-3% (lH-3% 4.89
ppm), and neither 2%-O- nor 2%,3%-di-O-benzoy-
lation was observed.
During glycosylation with the imide-type
donor 4, formation of glycal 13 was observed
in approximately the same quantity as in sialy-
lation with amide-type donor 3. Therefore a
twofold excess of donor 4 was required, and
about one half of unreacted acceptor was
recovered when glycosylation with a 1:1
donor–acceptor ratio was attempted. It was
found that Boc group introduction decreases
the reactivity of the sialyl-donor: while 3 re-
acted within 4 h at −35 °C, 4 was completely
At the next stage of the synthesis of trisac-
charides 1 and 2, the glycosylation of accep-
tors 8, 9, and 10 with sialyl donors 3 and 4

Table 2
1
Chemical shifts (l, ppm) and coupling constants (J, Hz) for carbohydrate ring protons in H NMR spectra of compounds 1, 2, 4 a, 8, 9, 15, 16, 21, 22, 28, 29 in the solvent
specified
Compound
(solvent)
Residue H-1
H-2
(J2,3
H-3eq
(J3eq,4
H-3ax
(J3ax,4
H-4
(J4,5
H-5
(J5,6a
H-6a
(J6a,6b
H-6b
H-7
(J7,8
H-8
(J8,9a
H-9a
(J9a,9b
H-9b
(J9b,8)
(J1,2
)
)
)
)
)
)
)
(J6b,5
)
)
)
)
1 (D2O)
GN
Gal
Neu
GN
Gal
Neu
4.53 (8.2) 3.76
4.45 (7.9) 3.51
3.77
3.63
3.62
3.82
3.81
3.59
3.77
3.86
3.99
3.97
3.69
3.97
3.96
3.79
3.83
3.53
3.64 (3.2) 3.92 (0)
2.67 b (4.5) 1.7 (12.3) 3.62
3.54
3.52
3.89
3.86
3.88
3.84
3.65
3.61
2 (D2O)
4.52 (8)
3.73
3.73
3.63
3.59
3.89
3.79
3.52
4.45 (7.7) 3.50
2.65 c (4.7) 1.69 (12.1) 3.74
4 a (CDCl3)
8 (1:10 CD3OD– GN
2.73 d (4.8) 2.12 (11.2) 5.76 (10) 4.82 (10) 5.42 e
3.56 (9.2) 3.88 (9.2) 3.33 3.69
5.32 (1.7) 5.07 (2)
4.45 (12.3) 4.0 (8.1)
4.34 (7.9) 3.63
3.62
CDCl3)
Gal
4.48 (8)
5.33 (10.1)
4.79 (3)
4.09 (0)
3.34
3.68 (11.9) 3.50 (3.6)
/
9 (CD3OD)
GN
Gal
4.47 (8.1) 3.82
4.54 (7.8) 3.98 (10.2)
4.44 (8.2) 3.60
3.69 (9.5) 4.04 (9.5) 3.57 (3.4) 4.02 (12)
3.86
3.61
4.89 (3.4) 4.11 (0)
3.55 3.70
3.44
3.31 (5)
3.74
15 (1:2 CD3OD–
CDCl3)
4.16 (10.5) 3.65
16 (acetone-d6)
21 (CDCl3)
4.59 (8.9) 3.79 (10.2)
4.53 (7.4) 3.65 (7.2)
4.83 (8.2) 5.73 (10)
4.44 (8.3) 3.71
3.66 (10.2) 3.58
3.87 (7.2) 4.15 (7.2) 3.44
5.47 (2.5) 5.91 (0) 3.99 (6.5) 4.51 (11.3) 4.31 (6.9)
3.68 3.90
3.49 (1.6) 3.85 (10.8) 3.70 (5.8)
GN
Gal
3.72 3.68
22 (2:1 CD3OD– GN
3.44
3.93
3.81
0
CDCl3)
(
Gal
4.43 (7.8) 3.59 (9.3)
4.52 (8.1) 3.77
4.47 (7.9) 3.55
3.42 (3.1) 4.15 (0)
3.75 3.63
3.66 (3.3) 3.93 (0)
1.62 (12.9) 4.07
3.09 (0)
3.65
3.82
3.87
3.60
4.17 (12.2) 3.88 (0)
28 (D2O)
29 (D2O)
GN
Gal
Neu
GN
Gal
Neu
4.02
3.73
3.78
3.99
3.71
3.87
3.85
3.52
4
2.38 f (5)
3.54
3.51
3.85
3.84
3.85
3.83
3.67
3.65
458
4.51 (8)
3.77
3.81
3.62
3.61
3.92
3.82
3.53
4.49 (7.7) 3.53
3.78
3.91
2.35 g (5) 1.58 (12.1) 4.09
a b anomer, major isomer at the nitrogen atom.
b J3eq,3ax 12.3 Hz.
c J3eq,3ax 12.2 Hz.
d J3eq,3ax 13.7 Hz.
e J6,7 1.7 Hz.
f J3eq,3ax 12.9 Hz.
g J3eq,3ax 12.1 Hz.

450
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
consumed only after overnight glycosylation
at −35 °C. It is also worth noting that 4 has
a much more hydrophobic nature in compari-
son with 3 which simplifies greatly chromato-
graphic purification of glycosylation products.
Trisaccharides 24 and 25 were deprotected
by hydrogenolysis followed by saponification
to give after gel-permeation chromatography,
the anomeric mixture 1+28 in 87% overall
yield. Subsequent separation of the mixture by
reverse-phase C18 chromatography afforded 1
(49%) and its b isomer 28 (38%). The structure
of 1 was determined by NMR spectroscopy
(Tables 2 and 3). In particular, the chemical
shifts of H-3eqNeu (l 2.67 ppm), H-3axNeu (l
1.7 ppm), H-4Neu (l 3.62 ppm), together with
C-1Neu (l 174.6 ppm) and C-6Neu (l 73.7
ppm) indicated the anomeric configuration of
the neuraminic acid to be a,19 while the value
of C-6Gal (l 64.5 ppm) proved 6-O-glycosyla-
tion position. In the similar fashion, the values
of H-3eqNeu (l 2.38 ppm), H-3axNeu (l 1.62
ppm), H-4Neu (l 4.07 ppm), together with
C-1Neu (l 176.0 ppm) and C-6Neu (l 71.5
ppm) showed b-Neu linkage in compound
28.19
At the next stage of the synthesis of 2,
selective deblocking of the nitrogen atom in
the neuraminic acid residue was performed
(Scheme 2). Treatment of compounds 26 and
27 with MeONa in dry MeOH followed by
acetylation of the crude reaction mixture af-
forded carbamate 30 in 78% yield. Removal of
the Boc group in 30 with trifluoroacetic acid
in CH2Cl2 afforded amine 31, which was im-
mediately acylated with acetoxyacetyl chloride
and Et3N to give N-glycolyl derivative 32 in
70% overall yield. The intermediate amine 31
was highly prone to O“N acetyl group mi-
gration, which occurred quantitatively even on
a TLC plate when eluted with neutral solvent
system. Therefore elution with acetic acid con-
taining mixtures was essential for TLC analy-
sis of amine 31.
Table 3
Chemical shifts (l, ppm) for carbohydrate ring carbons in 13C NMR spectra of compounds 1, 2, 4, 8, 15, 16, 21, 28, 29 in the
solvent specified.
Compound (solvent)
Residue
C-1
C-2
56.0
C-3
C-4
C-5
C-6
C-7
C-8
C-9
1 (D2O)
GN
Gal
Neu
GN
Gal
Neu
102.2
104.6
174.6
102.2
104.6
174.7
173.7
100.8
99.7
101.3
102.4
101.2
99.8
102.3
104.4
176.0
102.4
104.5
175.9
73.4
73.6
41.2
73.5
73.7
41.4
38.9
79.5
74.2
81.8
83.8
78.2
71.3
73.6
73.8
40.8
75.1
73.9
40.9
81.8
69.5
69.4
81.8
69.6
69.2
66.3
75.5
66.9
77.6
72.2
75.5
68.0
81.2
69.9
67.9
81.4
69.9
67.8
75.6
74.8
53.0
75.7
73.4
52.8
52.8
74.4
75.4
65.8
76.8
74.9
71.3
75.7
74.9
53.3
75.7
73.7
53.1
61.5
64.5
73.7
61.6
64.6
74.9
72.2
67.5
61.5
68.3
70.9
68.3
61.7
61.5
63.5
71.5
61.6
63.6
71.4
71.9
101.3
56.1
71.9
101.4
89.5
54.3
69.8
55.4
55.8
55.0
70.3
56.0
72.0
101.4
56.1
72.1
101.6
69.6
72.8
63.8
2 (D2O)
69.6
68.6
73.0
72.8
63.9
62.4
4 (CDCl3)
8 (1:10 CD3OD–CDCl3)
GN
Gal
15 (1:2 CD3OD–CDCl3)
16 (acetone-d6)
21 (CDCl3)
GN
Gal
GN
Gal
Neu
GN
Gal
Neu
28 (D2O)
29 (D2O)
69.4
69.4
71.3
71.4
64.6
64.7

A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
451
Scheme 2. Reagents and conditions: (i) MeONa, MeOH; (ii) Ac2O, Py; (iii) CF3CO2H, CH2Cl2; (iv) AcOCH2C(O)Cl, Et3N,
CH2Cl2, 0 °C; (v) H2, Pd–C, MeOH; (vi) NaOH, MeOH–water.
3. Experimental
The trisaccharide 32 was deprotected
through the reaction sequence involved cata-
lytic hydrogenolysis followed by alkaline hy-
drolysis of acyl protections. The anomeric
mixture 2+29 obtained after gel-permeation
chromatography in 62% overall yield was fur-
ther separated into individual anomers using
reverse-phase C18 column chromatography to
give 2 (31%) and 29 (31%). The presence of
General methods.—The reagents were pur-
chased from Fluka and Merck, all of the
highest grade available. Compound 1720 was
obtained from Syntesome GmbH, Munich.
Molecular sieves were activated by heating
(180 °C) under vacuum (0.1 mmHg) for 5–8
h. THF was stored over sodium wire and was
freshly distilled from sodium benzophenone
ketyl under Ar prior to use. CH2Cl2 was
washed with 96% H2SO4, water, saturated aq
NaHCO3, dried (MgSO4), distilled twice from
1
N-glycolyl substituent was evident from H
13
and C NMR spectra, where it appeared as a
two-proton singlet (l 4.15 ppm) and a signal
at 62.3 ppm (HOCH2C(O)N), respectively. In
compound 2, the chemical shifts (Tables 2 and
3) of H-3eqNeu (l 2.65 ppm), H-3axNeu (l
1.69 ppm), H-4Neu (l 3.74 ppm) together
with C-1Neu (l 174.7 ppm) and C-6Neu (l
74.9 ppm) indicated a configuration of the
neuraminic acid,19 while b linkage in 29 was
deduced from the values of H-3eqNeu (l 2.35
ppm), H-3axNeu (l 1.58 ppm), H-4Neu (l 4.09
ppm), together with C-1Neu (l 175.9 ppm)
and C-6Neu (l 71.4 ppm).
In conclusion, 3-aminopropyl glycosides of
trisaccharides Neu5Aca-(2“6%)-lactosamine
and Neu5Gca-(2“6%)-lactosamine were syn-
thesized. Examination of sialylation of di-,
tri-, and tetraol acceptors with both 3 and 4
did not show any influence on stereoselectiv-
ity, but glycosylation with imide-type donor 4
was slightly more effective, and isolation of
reaction products was more convenient. Ap-
plication of imide-type donor 4 opens ready
access to oligosaccharides containing modified
N-acyl analogues of Neu5Ac. Preparation of
neoglycoconjugates from compounds 1 and 2
and their use in biological assays will be pub-
lished elsewhere.
,
P2O5 under Ar, and stored over MS-4 A.
DMF was distilled twice from CaH2 and
stored over molecular sieve UOP Type 13 X
(Fluka). Nitromethane was always freshly dis-
tilled from CaH2 under Ar. The catalyst used
for hydrogenolysis was 10% Pd–C, oxide
form, (Merck-Schuchardt). In glycosylation
reactions, molecular sieve Union Carbide type
,
,
4 A (Fluka), or molecular sieve UOP type 3 A
(Fluka) were used. Column chromatography
was performed on silica gel 60 (Fluka, 70–230
mesh). TLC was performed on silica gel 60
F254 (E. Merck, Darmstadt, Germany) with
detection by dipping the chromatograms into
10% H3PO4 in EtOH followed by heating at
150 °C. Amines were also visualized by dip-
ping the chromatograms into 0.017 M
ninhydrin solution in n-butanol–AcOH
(33:1), followed by heating. For TLC ana-
lysis of deblocked oligosaccharides, solvent
systems 1:2:1 n-butanol–n-propanol–0.1 M
aq HCl (BPHCl), 1:1:1 MeCN–MeOH–water
(AMW), and their combination were used.
Optical rotation was measured with JASCO
DIP-360 digital polarimeter at 26–30 °C.
NMR spectra were recorded at 27 °C with
Bruker DRX-500 instrument (500 MHz for 1H

452
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
and 125 MHz for 13C), assignments were
C31H41NO14S: C, 54.46; H, 6.04; N, 2.05; S,
4.69. Found: C, 54.20; H, 6.12; N, 2.15; S,
4.79.
1
aided by APT, COSY, TOCSY, and H–13C
correlation spectroscopy. tert-Butyl alcohol
was used as an internal standard for D2O
solutions (1.24 ppm (1H) and 30.29 ppm (13C))
and Me4Si for other ones. The following nota-
tion was used to define the NMR signals: GN
for glucosamine unit, Gal for galactose unit,
Neu for neuraminic acid unit, and Sp for
spacer unit. Numeration of atoms in the
spacer is Sug-1-2-3. Mass spectra were
recorded using matrix assisted laser desorp-
tion ionization (MALDI)-time of flight (TOF)
on Vision 2000 mass spectrometer. Melting
points were determined with a Kofler appara-
tus and are uncorrected.
3-Trifluoroacetamidopropyl
2-acetamido-
4,6-O-benzylidene-3-O-benzyl-2-deoxy-i-
D
-
glucopyranoside (15).—NaH (60% suspension
in mineral oil, 2.4 g, 60 mmol) was added
portionwise to abs DMF (60 mL) at −20 °C
under Ar (the solvent should be redistilled if
any gas evolution is observed at this stage).
The mixture was stirred until the homoge-
neous suspension was formed and solid 14
(4.62 g, 10 mmol) was added portionwise for
15 min. The stirring was continued until gas
evolution ceased (ca. 1 h at −15 °C), BnBr
(1.3 mL, 11 mmol) was added, and the reac-
tion mixture was stirred at −20/−15 °C for
ca. 1.5 h, then cooled to −40 °C and glacial
AcOH (15 mL, 250 mmol) was carefully
added dropwise. The reaction mixture was
allowed to attain rt and after dilution with
EtOAc (400 mL) the precipitation of 15
started. This biphasic mixture was washed
with water (100 mL), saturated aq NaHCO3,
and the first crop of 15 was filtered off,
washed thoroughly with MeOH, and dried to
give 15 (1 g, 18%). The combined mother
liquor and washings were concentrated and
crystallized from MeOH to afford an addi-
tional lot of 15 (4 g, 72%) as fine powder: Rf
0.7 (EtOAc); mp (dec) 279–281 °C; [h]D −46°
Phenyl [methyl 4,7,8,9-tetra-O-acetyl-5-(N-
tert-butoxycarbonylacetamido)-3,5-dideoxy-2-
thio-
D
-glycero-h- and i- -galacto-2-nonu-
D
lopyranosid]onate (4).—A solution of 3 (500
mg, 0.86 mmol, 1:2.3 a/b), Boc2O (500 mg, 2.3
mmol) and 4-dimethylaminopyridine (DMAP)
(70 mg, 0.57 mmol) in dry THF (12 mL) was
refluxed under Ar and new portions of Boc2O
(250 mg, 1.15 mmol) and DMAP (35 mg,
0.028 mmol) were added every 6 h. After a
total of 24 h, the reaction mixture was cooled
to rt, diluted with CH2Cl2 (50 mL), washed
with 0.5 M aq H2SO4, water, saturated aq
NaHCO3, dried and concentrated. Chro-
matography (petroleum ether“1:1 petroleum
ether–EtOAc) on a column of silica gel (50 g)
gave the starting 3 (40 mg, 8%) and 600 mg of
impure 4 as a yellow syrup. The syrup was
subjected to gel-permeation chromatography
on a 3×70 cm column of Bio-Beads SX3
(Bio-Rad Laboratories) in toluene to afford 4
(500 mg, 85%) as white foam: Rf 0.8 (3:4
petroleum ether–EtOAc); NMR spectra
showed a 4.6:1 ratio of isomers at the nitrogen
atom; NMR data for b anomer, major isomer
1
(c 1, DMF); NMR (1:2 CD3OD–CDCl3): H,
see Table 2 for carbohydrate ring protons; l
7.35–7.1 (m, 10 H, Ph), 5.43 (s, 1 H, PhCH),
4.68 (d, 1 H, J 11.8 Hz, PhCH2), 4.49 (d, 1 H,
PhCH2), 3.73 (m, H-1aSp), 3.37 (m, 1 H,
H-1bSp), 3.29 (m, H-3aSp), 3.09 (m, 1 H,
H-3bSp), 1.89 (s, 3 H, NꢀAc), 1.65 (m, 2 H,
H-2aSp, H-2bSp); 13C, see Table 3 for carbo-
hydrate ring carbons; l 136.9 (ipso Ph),
128.6–125.6 (Ph), 100.9 (PhCH), 73.8
(PhCH2), 66.5 (C-1Sp), 36.5 (C-3Sp), 28.2 (C-
2Sp), 22.2 (NC(O)CH3). Anal. Calcd for
C27H31F3N2O7: C, 58.69; H, 5.65; N, 5.07.
Found: C, 58.73; H, 5.62; N, 4.99.
1
at the nitrogen atom (CDCl3): H, see Table 2
for carbohydrate ring protons; l 7.55–7.25
(m, 5 H, Ph), 3.53 (s, 3 H, OMe), 2.35 (s, 3 H,
NꢀAc), 2.03 (s, 6 H, 2 OAc), 1.94 (s, 6 H, 2
3-Trifluoroacetamidopropyl
3,6-di-O-benzyl-2-deoxy-i-
2-acetamido-
-glucopyranoside
13
OAc), 1.71 (s, 9 H, (CH3)3C); C, see Table 3
D
for carbohydrate ring carbons; l 170.4, 170.3,
170.1, 169.7, 168.1, 151.9 (6 CꢁO), 136.1,
129.6, 129.0 (Ph), 85.3 (CH3)3CO), 52.4
(OCH3), 28.2 (CH3)3CO), 26.6 (NꢀC(O)CH3),
20.9, 20.8, 20.7 (OC(O)CH3). Anal. Calcd for
(16).—(a) Compound 15 (570 mg, 1.03 mmol)
was refluxed in anhydrous THF (25 mL) un-
der Ar until complete dissolution and then the
mixture was allowed to attain rt. Me3N·BH3
(301 mg, 4.13 mmol) was added under stirring,

A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
453
0.65 mmol). Stirring was continued overnight
at rt, and the reaction mixture was filtered
through Celite, the filtrate was diluted with
EtOAc, washed with saturated aq NaHCO3, 3
M aq Na2S2O3, dried, and concentrated.
Chromatography (toluene“3:2 toluene–ace-
tone) on a column of silica gel (50 g) afforded
a mixture (187 mg) of 19 (Rf 0.31 in 2:1
toluene–acetone) and unreacted acceptor 16.
This mixture was dissolved in CH2Cl2 (5 mL)
and treated with 90% aq CF3CO2H (0.5 mL)
for 30 min at rt, then coevaporated with tolu-
ene (10 mL) and water (2 mL). Chromatogra-
phy (1:1 toluene–EtOAc“EtOAc) on a
column of silica gel (50 g) afforded amor-
phous 8 (80 mg, 37%): Rf 0.5 (double elution
in EtOAc); [h]D +4° (c 1, EtOAc); NMR
followed by anhyd AlCl3 (822 mg, 6.18
mmol). After the reagents had dissolved, wa-
ter (0.036 mL, 2 mmol) was added dropwise
and stirring was continued for 6 h until the
complete conversion of the starting 15 into the
product 16 (TLC control in 3:2 toluene–ace-
tone). The reaction mixture was quenched by
addition of water (10 mL) followed by 1 M aq
HCl (10 mL) and extracted with EtOAc (2×
100 mL), the extracts were washed with brine,
dried, and concentrated. The residue was dis-
solved in the minimum quantity of EtOAc, the
volume was doubled carefully with toluene,
and the solution was applied to a column of
silica gel (50 g). Gradient elution (toluene“
1:1 toluene–acetone) afforded amorphous 16
(498 mg, 87%): Rf 0.3 (3:2 toluene–acetone);
[h]D −12° (c 1, acetone); NMR (acetone-d6):
1H, see Table 2 for carbohydrate ring protons;
l 7.35–7.18 (m, 10 H, Ph), 4.83 (d, 1 H, J 11.6
Hz, PhCH2), 4.72 (d, 1 H, PhCH2), 4.59 (m, 4
H, H-1GN, 2 PhCH2, NꢀH Sp), 3.87 (m, 2 H,
H-1aSp, H-6aGN), 3.58 (m, 2 H, H-4GN,
H-1bSp), 3.48 (m, 2 H, H-5GN, H-3aSp), 3.32
(m, 1 H, H-3bSp), 1.89 (s, 3 H, NꢀAc), 1.81
(m, 2 H, H-2aSp, H-2bSp); 13C, see Table 3
for carbohydrate ring carbons; l 170.8
(NꢀC(O)CH3), 140.5, 140.0 (2 ipso Bn),
129.2–128.1 (Ph), 74.9, 73.9 (2 PhCH2), 66.9
(C-1Sp), 37.9 (C-3Sp), 29.6 (C-2Sp), 23.5
1
(1:10 CD3OD–CDCl3): H, see Table 2 for
carbohydrate ring protons; l 7.92 (d 2 H, J
7.6 Hz, ortho-protons of Bz), 7.45–7.12 (m,
13 H, Ph), 4.88 (d, 1 H, J 11.1 Hz, PhCH2),
4.61 (d, 1 H, J 12 Hz, PhCH2), 4.51 (d, 1 H,
PhCH2), 4.39 (d, 1 H, PhCH2), 3.77 (m, 1 H,
H-1aSp), 3.38 (m, H-1bSp, H-3aSp), 3.14 (m,
1 H, H-3bSp), 1.80 (s, 3 H, Ac), 1.79 (s, 3 H,
Ac), 1.68 (m, 2 H, H-2aSp, H-2bSp); 13C, see
Table 3 for carbohydrate ring carbons; l
171.4, 169.9, 165.7 (3 CꢁO), 137.6, 137.2 (2
ipso Bn), 133.2–127.6 (Ph), 74.9 (PhCH2),
73.3 (PhCH2), 66.2 (C-1Sp), 36.5 (C-3Sp), 28.1
(C-2Sp), 22.4 (NꢀC(O)CH3), 20.3 (OꢀC(O)-
CH3). Anal. Calcd for C42H49F3N2O14: C,
58.46; H, 5.72; N, 3.25. Found: C, 58.14; H,
5.77; N, 3.12.
(NꢀC(O)CH3);
C27H33F3N2O7: C, 58.48; H, 6.00; N, 5.05.
Found: C, 58.33; H, 5.92; N, 4.88.
Anal.
Calcd
for
(b) Performing of the same experiment
without addition of water gave unchanged 15
and less then 10% of 16 was observed by TLC.
Stirring of the reaction mixture for an addi-
tional 48 h at rt did not result in further
conversion of 15.
(b) Conventional acetylation with Ac2O in
pyridine of compound 23 (91 mg, 0.1 mmol)
followed by debenzylidenation as described in
(a) afforded 8 (75 mg, 87%): identical in all
respect to the material described above.
3-Trifluoroacetamidopropyl (2,3,4,6-tetra-O-
3-Trifluoroacetamidopropyl (2-O-acetyl-3-
O-benzoyl-i-
D
-galactopyranosyl)-(1“4)-2-
-glu-
benzoyl - i -
D
- galactopyranosyl) - (1“4) - 2-
-glu-
acetamido-3,6-di-O-benzyl-2-deoxy-i-
D
acetamido-3,6-di-O-benzyl-2-deoxy-i-
D
copyranoside (8).—(a) Acetylation of 17 (4.16
g, 10 mmol) with Ac2O (5 mL) in Py (10 mL)
and subsequent column chromatography gave
18 (4.53 g, 99%). A solution of 16 (140 mg,
0.25 mmol), 18 (198 mg, 0.43 mmol), and
Bu4NBr (22 mg, 0.07 mmol) in anhyd MeNO2
copyranoside (21).—Acceptor 16 (628 mg,
1.13 mmol) was refluxed in anhyd CH2Cl2 (20
mL) under Ar until complete dissolution and
then the mixture was allowed to attain rt.
Benzobromogalactose 20 (942 mg, 1.43 mmol)
,
was added under Ar, followed by MS-4 A (4.5
,
(12 mL) was stirred with activated MS-4 A (3
g). After stirring for 15 min, powdered AgOTf
(514 mg, 2 mmol) was added, and stirring was
continued overnight at rt. Triethylamine (0.1
g) for 1 h under Ar, and AgOTf (167 mg, 0.65
mmol) was added followed by CuBr2 (145 mg,

454
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
mL) was added, the mixture was filtered
through a pad of Celite, diluted with CH2Cl2
(200 mL), washed with saturated aq NaHCO3,
3 M aq Na2S2O3, dried, and concentrated.
Chromatography (toluene“1:2 toluene–
EtOAc) on a column of silica gel (30 g) af-
forded 21 (906 mg, 70%) as white foam: Rf 0.4
(1:2 toluene–EtOAc); [h]D +16° (c 1,
H, PhCH), 5.18 (d, 1 H, J 11.3 Hz, PhCH2),
4.68 (d, 2 H, 2 PhCH2), 4.58 (d, 1 H, J 11.9
Hz, PhCH2), 3.79 (m, H-1aSp), 3.48 (m, H-
1bSp), 3.42 (m, 1 H, H-3aSp), 3.38 (m, 1 H,
H-3bSp), 1.91 (s, 3 H, NꢀC(O)CH3), 1.82 (m,
2 H, H-2aSp, H-2bSp); 13C, see Table 3 for
carbohydrate
ring
carbons;
l
172.8
(NꢀC(O)CH3), 139.7, 139.2, 139.1 (3 ipso Bn),
104.1, 102.1 (2 C-1), 101.9 (PhCH), 81.7 (C-
3GN), 75.3, 74.1 (2 PhCH2), 69.6, 69.3 (2
C-6), 67.4 (C-1Sp), 56.1 (C-2GN), 37.7 (C-
3Sp), 29.4 (C-2Sp), 23.1 (NꢀC(O)CH3). Anal.
Calcd for C40H47F3N2O12: C, 59.70; H, 5.89;
N, 3.48. Found: C, 59.40; H, 5.84; N, 3.20.
3-Trifluoroacetamidopropyl (3-O-benzoyl-i-
1
CH2Cl2); NMR (CDCl3): H, see Table 2 for
carbohydrate ring protons; l 8.10–7.75 (m, 8
H, ortho-protons of 4 Bz), 7.62–7.15 (m, 22
H, Ph), 5.92 (d, 1 H, JN-H,2 10 Hz, NꢀH GN),
4.95 (d, 1 H, J 11.8 Hz, PhCH2), 4.75 (d, 1 H,
PhCH2), 4.67 (d, 1 H, J 11.9 Hz, PhCH2), 4.39
(d, 1 H, PhCH2), 3.77 (m, H-1aSp, H-3aSp),
3.28 (m, 2 H, H-1bSp, H-3bSp), 1.99 (s, 3 H,
NꢀAc), 1.72 (m, 2 H, H-2aSp, H-2bSp); 13C,
see Table 3 for carbohydrate ring carbons; l
170.8 (NꢀC(O)CH3), 165.5 (PhC(O)O), 138.5,
137.8 (2 ipso Bn), 133.7–127.9 (Ph), 73.6
(PhCH2), 67.3 (C-1Sp), 37.6 (C-3Sp), 28.1 (C-
2Sp), 23.4 (NꢀC(O)CH3). Anal. Calcd for
C61H59F3N2O16: C, 64.66; H, 5.25; N, 2.47.
Found: C, 64.64; H, 5.25; N, 2.37.
D
- galactopyranosyl) - (1“4) - 2 - acetamido-
3,6-di-O-benzyl-2-deoxy-i- -glucopyranoside
D
(9).—To a solution of 22 (100 mg, 0.12 mmol)
in Py (3 mL), BzCN (150 mg, 1.2 mmol) was
added portionwise at rt until TLC indicated
complete consumption of the starting mate-
rial. Methanol (1 mL) was added, and after 30
min the reaction mixture was concentrated to
give crude 23, Rf 0.54 (EtOAc), which was
debenzylidenated as described for the prepara-
tion of 8. Chromatography (EtOAc“10:1
EtOAc–MeOH) of the residue on a column of
silica gel (20 g) afforded amorphous 9 (82 mg,
80% overall): Rf 0.27 (EtOAc); [h]D −3° (c 2,
3-Trifluoroacetamidopropyl (4,6-O-benzyli-
dene - i -
D
- galactopyranosyl) - (1“4) - 2 - ace-
-gluco-
tamido-3,6-di-O-benzyl-2-deoxy-i-
D
pyranoside (22).—A 0.2 M solution of Me-
ONa in MeOH (0.4 mL, 0.08 mmol) was
added to the solution of 21 (900 mg, 0.79
mmol) in anhyd MeOH (10 mL), and after
stirring overnight at rt, the reaction mixture
was neutralized with KU-2(H+) cation ex-
change resin, filtered, concentrated, and dried
in vacuo to give the crude 10, Rf 0.15 (10:1
EtOAc–MeOH), which was used without fur-
ther purification. A mixture of all obtained 10,
PhCH(OMe)2 (0.2 mL, 1.4 mmol), and TsOH
(10 mg) in DMF (4 mL) was stirred at 50 °C
for 2 h, neutralized with Et3N (0.1 mL), and
coevaporated with toluene (4×20 mL). Chro-
matography in EtOAc of the residue on a
column of silica gel (50 g) afforded 22 (561
mg, 88%) as ca. 4:1 mixture of isomers at the
benzylidene acetal center, Rf 0.47 (major) and
0.53 (minor) in 10:1 EtOAc–MeOH. Crystal-
lization of the mixture from MeOH gave indi-
vidual major isomer: mp 216–218 °C; [h]D
−41° (c 2, MeOH); NMR (2:1 CD3OD–
1
MeOH); NMR (CD3OD): H, see Table 2 for
carbohydrate ring protons; l 8.12 (d, 2 H, J
7.7 Hz, ortho-protons of Bz), 7.65–7.18 (m,
13 H, Ph), 5.11 (d, 1 H, J 10.8 Hz, PhCH2),
4.65 (d, 1 H, J 11.9 Hz, PhCH2), 4.61 (d, 1 H,
J 10.8 Hz, PhCH2), 4.55 (d, 1 H, J 11.9 Hz,
PhCH2), 3.87 (m, H-1aSp), 3.53 (m, H-1bSp),
3.43 (m, H-3aSp), 3.29 (m, 1 H, H-3bSp), 1.95
(s, 3 H, NꢀC(O)CH3), 1.82 (m, 2 H, H-2aSp,
H-2bSp); 13C, l 173.3 (NꢀC(O)CH3), 167.7
(OC(O)Ph), 139.6, 139.5 (2 ipso Bn), 134.3–
128.8 (Ph), 104.2, 102.6 (2 C-1), 82.3 (C-
3GN), 77.8, 77.5, 77.1, 76.6, 76.1, 74.2, 70.5,
69.2, 67.9, 67.7, 62.7 (C-6Gal), 56.2 (C-2GN),
37.9 (C-3Sp), 29.8 (C-2Sp), 23.0 (NꢀC(O)-
CH3). Anal. Calcd for C40H47F3N2O13: C,
58.53; H, 5.77; N, 3.41. Found: C, 58.36; H,
5.84; N, 3.30.
3-Trifluoroacetamidopropyl [methyl (5-acet-
amido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-
glycero-h- and i-
osyl)onate]-(2“6)-(2-O-acetyl-3-O-benzoyl-
D
-
1
CDCl3): H, see Table 2 for carbohydrate ring
D-galacto-2-nonulopyran-
protons; l 7.53–7.15 (m, 15 H, Ph), 5.53 (s, 1

A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
455
and 0.31 (a anomer) in EtOAc (double elu-
tion); integration of the intensities of C-1Neu
in 13C NMR spectrum showed 1.2:1 a/b
anomeric ratio; NMR (1:2 CD3OD–CDCl3):
1H, l 8.05–7.95 (2 d, 2 H, J 7.8 Hz, ortho-
protons of Bz), 7.49–7.12 (m, 13 H, Ph), 3.61
and 3.59 (2 s, OMe), 2.33 (broaden dd, 1 H,
J3eq,4 5.1, J3eq,3ax 12.5 Hz, H-3eqNeu), 2.1–1.7
(m, 12 Ac, H-3axNeu, H-2aSp, H-2bSp); 13C, l
138.2, 137.4 (ipso Bn), 129.5–127.3 (Ph),
103.0 (C-1GN), 100.8 and 100.6 (C-1Gal),
98.6 and 98.1 (C-2Neu), 79.8 and 79.6 (C-
3GN), 68.4 and 68.2 (C-6GN), 66.2 (C-1Sp),
54.7 and 54.4 (C-2GN), 52.5 (OCH3), 36.9
(C-3Neu), 36.5 (C-3Neu, C-3Sp), 28.1 (C-
2Sp), 22.4 and 22.1 (NꢀC(O)CH3), 20.1
(OꢀC(O)CH3). Anal. Calcd for C60H74F3-
N3O25: C, 55.68; H, 5.76; N, 3.25. Found: C,
55.35; H, 5.84; N, 3.02.
i -
D
- galactopyranosyl) - (1“4) - 2 - acetamido-
-glucopyranoside
3,6-di-O-benzyl-2-deoxy-i-
D
(24).—A mixture of donor 3 (163 mg, 0.28
mmol), acceptor 8 (120 mg, 0.14 mmol), and
anhyd MeCN (3 mL) was stirred with freshly
,
activated MS-3 A (1 g) during 1 h at rt under
Ar, NIS (126 mg, 0.56 mmol) was added, and
after it had dissolved, the reaction mixture
was cooled to −35 °C and TfOH (0.005 mL)
was added. After being stirred for 4 h at
−35 °C, the reaction was terminated with
Et3N (0.1 mL), the mixture was filtered
through Celite, diluted with CH2Cl2, washed
with saturated aq NaHCO3, 3 M aq Na2S2O3,
dried, and concentrated. Chromatography
(CH2Cl2 “MeCN) of the residue on a column
of silica gel (30 g) afforded the anomeric
mixture 24 (149 mg, 79%) as white foam: Rf
0.39 (b anomer) and 0.33 (a anomer) in
EtOAc (double elution); integration of the
3-Trifluoroacetamidopropyl {methyl [4,7,8,9-
tetra-O-acetyl-5-(N-tert-butoxycarbonylace-
1
intensities of H-4Gal in H NMR spectrum
showed 1.3:1 a/b anomeric ratio; NMR
tamido)-3,5-dideoxy-
galacto - 2 - nonulopyranosyl]onate} - (2“6)-
(2-O-acetyl-3-O-benzoyl-i- -galactopyran-
osyl)-(1“4)-2-acetamido-3,6-di-O-benzyl-2-
deoxy-i- -glucopyranoside (26).—Glycosyla-
D
-glycero-h- and i- -
D
1
(CDCl3): H, l 8.12–8.01 (2 d, 2 H, ortho-
protons of Bz), 7.19–7.22 (m, 13 H, Ph), 5.49
(dd, 1 H, J2,1 8, J2,3 10.2 Hz, H-2Gal), 5.39–
5.21 (m, H-8Neu, H-7Neu, H-4bNeu), 5.03
(dd, 1 H, J3,4 3.1 Hz, H-3Gal), 4.89–4.49 (m,
4 PhCH2, H-4aNeu, H-1GN, H-1Gal), 4.41
(dd, 1 H, J9a,9b 12.5, J9a,8 3.5 Hz, H-9aNeu),
4.37 (d, H-4Gal of b anomer), 4.25 (d, H-4Gal
of a anomer), 2.51 (broaden dd, 1 H, J3eq,4 5,
J3eq,3ax 12.7 Hz, H-3eqNeu), 2.1–1.7 (m, 14 Ac,
H-3axNeu, H-2aSp, H-2bSp). Anal. Calcd for
C62H76F3N3O26: C, 55.73; H, 5.73; N, 3.14.
Found: C, 55.70; H, 5.68; N, 3.10.
D
D
tion of acceptor 8 (35 mg, 0.04 mmol) with
donor 4 (55 mg, 0.08 mmol) in anhyd MeCN
(3 mL) promoted by NIS (36 mg, 0.16 mmol),
,
TfOH (0.006 mL), and MS-3 A (1 g) for 24 h
at −35 °C as described for the preparation of
24 and subsequent chromatography (tolu-
ene“1:1 toluene–acetone) of the residue on a
column of silica gel (30 g) afforded glycal 13
(22 mg, 50%) and the anomeric mixture 26 (48
mg, 84%) as syrup.
3-Trifluoroacetamidopropyl [methyl (5-acet-
amido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-
glycero-h- and i-
osyl)onate]-(2“6)-(3-O-benzoyl-i-
pyranosyl) - (1“4) - 2 - acetamido - 3,6 - di - O-
benzyl-2-deoxy-i- -glucopyranoside (25).—
D-
Compound 13: Rf 0.88 in EtOAc; NMR
D
-galacto-2-nonulopyran-
-galacto-
data for the main isomer at the nitrogen atom
1
D
(CDCl3): H, l 6.02 (broaden d, 1 H, J3,4
9
Hz, H-3), 5.89 (broaden d 1 H, J4,3 2.8 Hz,
H-4), 5.28 (m, 3 H, H-6, H-7, H-8), 4.91
(broaden m, H-5), 4.61 (broaden d, H-9a),
4.17 (dd, 1 H, J9b,9a 12.5, J9b,8 6.2 Hz, H-9b),
D
Glycosylation of acceptor 9 (106 mg, 0.13
mmol) with donor 3 (163 mg, 0.28 mmol) in
anhyd MeCN (3 mL) promoted by NIS (126
mg, 0.56 mmol), TfOH (0.005 mL), and MS-3
3.79 (s,
3
H, OMe), 2.39 (s,
3
H,
NꢀC(O)CH3), 2.1–1.9 (4 s, 12 H, OC(O)CH3),
1.55 (s, 9 H OC(CH3)3); 13C, l 146.3 (C-2),
109.1 (C-3), 85.2 (OC(CH3)3), 55.5 (OMe),
27.9 (OC(CH3)3), 27.1 (NꢀC(O)CH3), 20.8
(OꢀC(O)CH3). Anal. Calcd for C25H35NO14:
C, 52.35; H, 6.15; N, 2.44. Found: C, 52.12;
H, 5.98; N, 2.18.
,
A (1 g) for 24 h at −35 °C as described for
the preparation of 24 and subsequent chro-
matography (CH2Cl2“MeCN) of the reac-
tion products on a column of silica gel (50 g)
afforded the anomeric mixture 25 (98 mg,
59%) as transparent glass: Rf 0.34 (b anomer)

456
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
and 37.1 (C-3Neu), 28.2 (C-2Sp), 27.7
Compound 26: Rf 0.65 in EtOAc; integra-
13
(OC(CH3)3), 27.5 and 27.0 and 26.6 (Boc–N–
C(O)CH3), 23.3 (NꢀAc), 20.9, 20.7, 20.6, 20.4
(OꢀAc). Anal. Calcd for C65H82F3N3O27: C,
55.99; H, 5.93; N, 3.01. Found: C, 56.27; H,
5.79; N, 2.87.
tion of the intensities of C-1Neu in C NMR
spectrum showed 1.3:1 a/b anomeric ratio;
1
NMR (CDCl3): H, l 8.17–8.06 (m, 2 H,
ortho-protons of Bz), 7.61–7.14 (m, 13 H,
Ph), 5.51 (dd, 1 H, J2,1 8, J2,3 10 Hz, H-2Gal),
5.02 (dd, 1 H, J3,4 3 Hz, H-3Gal), 2.61 (m, 1
H, H-3eqNeu), 2.49–1.75 (m, 24 H, 5 OAc, 2
NAc, H-3axNeu, H-2aSp, H2bSp), 1.59–1.48
(3 s, 9 H, OC(CH3)3); 13C, l 138.3, 137.8 (2
ipso Bn), 133.4–127.9 (Ph), 101.1, 100.2 (C-1
GN, C-1Gal), 99.2 and 98.3 (C-2Neu), 85.3
and 85.0 and 84.7 (OC(CH3)3), 77.8 and 77.5
(C-3GN), 56.3 and 56.1 (C-2GN), 38.3 (C-
3Neu), 37.6 (C-3Sp, C-3Neu), 37.1 (C-3Neu),
29.6 (C-2Sp), 27.8 (OC(CH3)3), 27.6 and 27.2
and 26.7 (Boc–N–C(O)CH3), 23.4 and 23.3
(NꢀAc), 21.1, 20.8, 20.4 (OꢀAc). Anal. Calcd
for C67H84F3N3O28: C, 56.02; H, 5.89; N, 2.93.
Found: C, 55.98; H, 5.98; N, 2.84.
3-Aminopropyl [sodium (5-acetamido-3,5-
dideoxy-
nosyl)onate] - (2“6) - (i -
(1“4) - 2 - acetamido - 2 - deoxy - i -
pyranoside (1) and 3-aminopropyl [sodium (5-
acetamido-3,5-dideoxy- -glycero-i- -galacto-
2-nonulopyranosyl)onate]-(2“6)-(i- -galac-
topyranosyl)-(1“4)-2-acetamido-2-deoxy-i-
-glucopyranoside (28).—To a solution of 24
D
-glycero-h-
D
-galacto-2-nonulopyra-
- galactopyranosyl)-
- gluco-
D
D
D
D
D
D
(123 mg, 0.09 mmol) in MeOH (6 mL), Pd–C
(20 mg) was added, the mixture was degassed
under vacuum with stirring, refilled with hy-
drogen, and stirred for 2 h under H2 at rt. The
reaction mixture was filtered through Celite,
the pad was washed thoroughly with MeOH,
and the filtrate was concentrated. A solution
of the residue in MeOH (4 mL) was treated
with solid MeONa (118 mg, 2.2 mmol), and
after 1 h at rt, water (1 mL) was added, and
the reaction mixture was stirred overnight.
The pH of the solution was carefully made
neutral by addition of AcOH, and the reaction
mixture was concentrated. Chromatography
of the residue on a 1.5×100 cm column of
TSK HW-40S gel (TOYO SODA Manufac-
turing Co.) by elution with 0.1 M aq AcOH
and subsequent lyophilization afforded
anomeric mixture of compounds 1 and 28 (60
mg, 87%). This mixture was subjected to re-
verse-phase column chromatography on a RP
C18 (5 micron particle size) 1×25 cm stain-
less steel column (IBM Instruments, Inc) by
elution with water to give (in order of elution)
b anomer 28 (26 mg, 38%) and a anomer 1 (34
mg, 49%).
Compound 28: Rf 0.34 in 1:1 BPHCl–
AMW; [h]D −25° (c 1, water); NMR (D2O):
1H, see Table 2 for carbohydrate ring protons;
l 4.04 (m, H-1aSp), 3.73 (m, H-1bSp), 3.09
(t, 2 H, J 7.2 Hz, H-3aSp, H-3bSp), 2.04
(s, 6 H, 2 NꢀAc), 1.95 (m, 2 H, H-2aSp,
H-2bSp); 13C, see Table 3 for carbohydrate
ring carbons; l 175.9 (2 NꢀC(O)CH3), 69.0
(C-1Sp), 38.8 (C-3Sp), 27.8 (C-2Sp), 23.3
(2 NꢀC(O)CH3); MALDI-TOF-MS: Calcd for
[C28H48N3NaO19+H]+: 754.3. Found 755.1.
3-Trifluoroacetamidopropyl {methyl [4,7,8,9-
tetra-O-acetyl-5-(N-tert-butoxycarbonylace-
tamido)-3,5-dideoxy-
galacto - 2 - nonulopyranosyl]onate} - (2“6)-
(3-O-benzoyl-i- -galactopyranosyl)-(1“4)-
2-acetamido-3,6-di-O-benzyl-2-deoxy-i- -glu-
D
-glycero-h- and i- -
D
D
D
copyranoside (27).—Glycosylation of acceptor
9 (58 mg, 0.07 mmol) with donor 4 (96 mg,
0.14 mmol) in anhyd MeCN (5 mL) promoted
by NIS (63 mg, 0.16 mmol), TfOH (0.01 mL),
,
and MS-3 A (1.2 g) for 24 h at −35 °C as
described for the preparation of 24 and subse-
quent chromatography (toluene“1:2 tolu-
ene–acetone) of the residue on a column of
silica gel (30 g) afforded glycal 13 (48 mg,
50%) and the anomeric mixture 27 (59 mg,
60%) as foam: Rf 0.3 in 3:2 toluene–acetone;
13
integration of the intensities of C-1Neu in C
NMR spectrum showed 1:1 a/b anomeric ra-
1
tio; NMR (CDCl3): H, l 8.25–8.09 (m, 2 H,
ortho-protons of Bz), 7.65–7.16 (m, 13 H,
Ph), 5.91 (d, 1 H, JN-H,2 8 Hz, NꢀH GN), 5.65
(m, H-4bNeu), 5.35 (m, H-4aNeu), 4.95 (m,
H-3Gal), 2.51 (m, 1 H, H-3eqNeu), 2.49–1.75
(m, 21 H, 4 OAc, 2 NAc, H-3axNeu, H-2aSp,
13
H2bSp), 1.59–1.40 (3 s, 9 H, OC(CH3)3); C,
l 138.5, 137.4 (2 ipso Bn), 133.1–127.9 (Ph),
103.3 (C-1GN), 100.7 (C-1Gal), 98.9 and 98.1
(C-2Neu), 85.1 and 84.9 and 84.6 (OC(CH3)3),
79.2 and 78.9 (C-3GN), 55.9 and 55.8 (C-
2GN), 38.2 (C-3Neu), 37.5 (C-3Sp, C-3Neu)

A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
457
(OꢀC(O)CH3);
Anal.
Calcd
for
Compound 1: Rf 0.4 in 1:1 BPHCl–AMW;
1
C62H82F3N3O28: C, 54.19; H, 6.01; N, 3.06.
Found: C, 54.10; H, 6.08; N, 3.15.
[h]D −21° (c 0.86, water); NMR (D2O): H,
see Table 2 for carbohydrate ring protons; l
4.02 (m, H-1Sp), 3.68 (m, H-1bSp), 3.09 (t, 2
H, J 7 Hz, H-3aSp, H-3bSp), 2.07 (s, 3 H,
N-Ac), 2.03 (s, 3 H, N-Ac), 1.95 (m, 2 H,
H-2aSp, H-2bSp); 13C, see Table 3 for carbo-
hydrate ring carbons; l 176.1, 175.9 (2
NꢀC(O)CH3), 69.1 (C-1Sp), 38.8 (C-3Sp), 27.8
(C-2Sp), 23.4, 23.1 (2 NꢀC(O)CH3); MALDI-
TOF-MS: Calcd for [C28H48N3NaO19+Na]+:
776.3. Found 777.0.
3-Aminopropyl [sodium (3,5-dideoxy-5-hy-
droxyacetamido -
D
- glycero - h -
D
- galacto -2-
-galacto-
pyranosyl)-(1“4)-2-acetamido-2-deoxy-i-
glucopyranoside (2) and 3-aminopropyl (sodium
nonulopyranosyl)onate]-(2“6)-(i-
D
D
-
[3,5-dideoxy-5-hydroxyacetamido-
D-glycero-i-
D
(i-
2-deoxy-i-
- galacto - 2 - nonulopyranosyl]onate) - (2“6)-
D
-galactopyranosyl)-(1“4)-2-acetamido-
-glucopyranoside (29).—To a so-
D
lution of 30 (31 mg, 0.023 mmol) in dry
CH2Cl2 (1 mL), CF3CO2H (0.3 mL) was
added. After 30 min at rt the reaction mixture
was coevaporated with toluene (10 mL) at the
bath temperature ]30 °C, and dried in vacuo
to give amine 31 (Rf 0.5 in 20:1:1 CH2Cl2–
MeOH–AcOH). The residue was dissolved in
CH2Cl2 (2 mL), cooled to 0 °C, and acetoxy-
acetyl chloride (0.025 mL, 0.22 mmol) was
added, followed by Et3N (0.05 mL, 0.36
mmol). After stirring for 1 h the reaction was
quenched with saturated aq NaHCO3 (1 mL),
diluted with CH2Cl2 (30 mL), washed with
saturated aq NaHCO3, dried, and concen-
trated. Chromatography (3:1 EtOAc–tolu-
ene“EtOAc“10:1 EtOAc–MeOH) on a
column of silica gel (20 g) afforded 32 (22 mg,
70%), as white glass, Rf 0.39 (EtOAc). Depro-
tection of compound 32 (20 mg, 0.014 mmol)
and separation of a and b anomers were per-
formed as described for the preparation of 1
and 28 to give (in order of elution) b anomer
29 (3.5 mg, 31%) and a anomer 2 (3.5 mg,
31%).
3-Trifluoroacetamidopropyl {methyl [4,7,8,9-
tetra - O - acetyl - 5 - (N - tert - butoxycarbonyl-
amino)-3,5-dideoxy-
galacto - 2 - nonulopyranosyl]onate} - (2“6)-
(2,3,4 - tri - O - acetyl - i - - galactopyranosyl)-
(1“4)-2-acetamido-3,6-di-O-benzyl-2-deoxy-
i- -glucopyranoside (30).—A 0.16 M solution
D
-glycero-h- and i- -
D
D
D
of MeONa in MeOH (0.5 mL, 0.08 mmol)
was added to the solution of 27 (50 mg, 0.036
mmol) in anhyd MeOH (1 mL), and after
stirring for 1 h at rt under Ar TLC (4:1
EtOAc–BPHCl) indicated deacylation to be
complete. Acetic acid (0.06 mL) was added,
and the reaction mixture was concentrated
and dried in vacuo. The residue was dissolved
in pyridine (2 mL) and Ac2O (0.5 mL) was
added. After stirring overnight at rt, MeOH (1
mL) was added, and the reaction mixture was
concentrated with toluene. Chromatography
(toluene“3:2 toluene–acetone) of the residue
on a column of silica gel (30 g) followed by
gel-permeation chromatography on a 1.5×45
cm column of Bio-Beads SX3 in toluene af-
forded 30 (38.5 mg, 78%) as white glass: Rf
Compound 29: Rf 0.25 in 1:1 BPHCl–
AMW; [h]D −26° (c 0.25, water); NMR
1
0.65 (EtOAc); NMR (CDCl3): H, l 7.51
1
(broaden s, NꢀHSp), 7.41–7.25 (m, 10 H, Ph),
5.15 (m, H-4bNeu), 4.99 (m, 1 H, H-3Gal),
4.81 (m, H-4aNeu), 3.88 (m, H-1aSp), 3.51
(m, H-1bSp), 3.44 (H-3aSp), 3.35 (H-3bSp),
2.51-2.49 (m, 1 H, H-3eqNeu), 2.1–1.9 (m, 7
OAc, 1 N-Ac, H-3axNeu, H-2aSp, H-2bSp),
1.38 (s, 9 H, OC(CH3)3); 13C, l 176.9
(NꢀC(O)CH3), 170.6, 169.9, 169.7 (C-1Neu,
O-C(O)CH3) 155.8 (NꢀC(O)OtꢀBu), 138.2 and
137.8 (2 ipso Bn), 128.9–125.3 (Ph), 101.1 and
100.8 (2 C-1GN), 99.8 and 99.6 (2C-1Gal),
98.9 and 98.7 (2 C-2Neu), 38.0 and 37.2 (2
C-3 Neu), 37.6 (C-3Sp), 29.5 (C-2Sp), 28.2
(OC(CH3)3), 23.3 (NꢀC(O)CH3), 21.4–20.6
(D2O): H, see Table 2 for carbohydrate ring
protons; l 4.15 (s, HOCH2C(O)N), 4.03 (m,
H-1aSp), 3.70 (m, H-1bSp), 3.08 (t, 2 H, J 6.8
Hz, H-3aSp, H-3bSp), 2.03 (s, 3 H, N-Ac),
13
1.91 (m, 2 H, H-2aSp, H-2bSp); C, see Table
3 for carbohydrate ring carbons; l 176.7
(NꢀC(O)CH2OH), 175.9 (NꢀC(O)CH3), 69.1
(C-1Sp), 62.3 (NꢀC(O)CH2OH), 38.9 (C-3Sp),
27.9 (C-2Sp), 23.4 (NꢀC(O)CH3); MALDI-
TOF-MS: Calcd for [C28H48N3NaO20+H]+:
769.3. Found 770.0.
Compound 2: Rf 0.35 in 1:1 BPHCl–AMW;
[h]D −28° (c 0.2, water); NMR (D2O): 1H, see
Table 2 for carbohydrate ring protons; l 4.15

458
A.A. Sherman et al. / Carbohydrate Research 330 (2001) 445–458
4. Tanahashi, E.; Fukunaga, K.; Ozawa, Y.; Toyoda, T.;
Ishida, H.; Kiso, M. J. Carbohydr. Chem. 2000, 19 (6),
747–768.
5. Simeoni, L. A.; Byramova, N. E.; Bovin, N. V. Bioorgan.
Khim. (Rus. J. Bioorg. Chem.; English Transl.) 2000, 26,
206–214.
6. Fujita, S.; Numata, M.; Sugimoto, M.; Tomita, K.;
Ogawa, T. Carbohydr. Res. 1992, 228, 347–370.
7. Demchenko, A. V.; Boons, G. J. Tetrahedron Lett. 1998,
39, 3065–3068.
8. Demchenko, A. V.; Boons, G. J. Chem. Eur. J. 1999, 5,
1278–1283.
(s, 2 H, HOCH2C(O)N), 3.99 (m, H-1aSp),
3.68 (m, H-1bSp), 3.06 (t, 2 H, J 6.8 Hz,
H-3aSp, H-3bSp), 2.06 (s, 3 H, N-Ac), 1.87
(m, 2 H, H-2aSp, H-2bSp); 13C, see Table 3
for carbohydrate ring carbons; l 176.9
(NꢀC(O)CH2OH), 175.9 (NꢀC(O)CH3), 69.2
(C-1Sp and C-4Neu), 62.2 (NꢀC(O)CH2OH),
38.9
(C-3Sp),
27.9
(C-2Sp),
23.5
(NꢀC(O)CH3); MALDI-TOF-MS: Calcd for
[C28H48N3NaO20+H]+: 769.3. Found 769.5.
9. Hasegawa, A.; Nagahama, T.; Ohki, H.; Hotta, K.;
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Acknowledgements
The authors are grateful to Dr L.O.
Kononov (N.D. Zelinsky Institute of Organic
Chemistry, Moscow) for helpful discussions.
This work was supported, in part, by Synte-
some GmbH, Munich, Russian Foundation
for Basic Research (Project No. 99-03-
33047a), and INTAS (Project No. 97-32036).
12. Marra, A.; Sinay¨, P. Carbohydr. Res. 1989, 187, 35–42.
13. Burk, M. J.; Allen, J. G. J. Org. Chem. 1997, 62, 7054–
7057.
14. Bovin, N. V.; Zemlyanukhina, T. V.; Chagiashvili, C. N.;
Khorlin, A. Y. Khim. Prir. Soedin. 1988, 777.
15. Ek, M.; Garegg, P. J.; Hultberg, H.; Oscarson, S. J.
Carbohydr. Chem. 1983, 2, 305–312.
16. Byramova, N. E.; Ovchinnikov, M. V.; Backinowsky, L.
V.; Kochetkov, N. K. Carbohydr. Res. 1983, 124, C8–
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17. Sato, S.; Mori, M.; Ito, Y.; Ogawa, T. Carbohydr. Res.
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