Synthesis of N-acetyllactosamine-containing oligosaccharides, galectin ligands

Article abstract of DOI:10.1134/S1068162007010141

The following spacered oligosaccharides were synthesized: GlcNAcβ1-3Galβ1-4GlcNAcβ-sp, GlcNAcβ1-6Galβ1- 4GlcNAcβ-sp, GlcNAcβ1-3(GlcNAcβ1-6)Galβ1-4GlcNAcβ-sp, Galβ1-4GlcNAcβ1-3Galβ1-4GlcNAcβ-sp, Galβ1- 4GlcNAcβ1-6Galβ1-4GlcNAcβ-sp, Galβ1-4GlcNAcβ1- 3(Galβ1

Full text of DOI:10.1134/S1068162007010141

ISSN 1068-1620, Russian Journal of Bioorganic Chemistry, 2007, Vol. 33, No. 1, pp. 122–138. © Pleiades Publishing, Inc., 2007.
Original Russian Text © V.V. Severov, I.M. Belyanchikov, G.V. Pazynina, N.V. Bovin, 2007, published in Bioorganicheskaya Khimiya, 2007, Vol. 33, No. 1, pp. 131–147.
SYNTHETIC
STUDIES
Synthesis of N-Acetyllactosamine-Containing Oligosaccharides,
Galectin Ligands
V. V. Severov, I. M. Belyanchikov, G. V. Pazynina, and N. V. Bovin1
Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences,
ul. Miklukho-Maklaya 16/10, Moscow, 117997 Russia
Received April 16, 2006; in final form, May 24, 2006
Abstract—The following spacered oligosaccharides were synthesized: GlcNAcβ1-3Galβ1-4GlcNAcβ-sp,
GlcNAcβ1-6Galβ1-4GlcNAcβ-sp,
GlcNAcβ1-3(GlcNAcβ1-6)Galβ1-4GlcNAcβ-sp,
Galβ1-4GlcNAcβ1-
3Galβ1-4GlcNAcβ-sp, Galβ1-4GlcNAcβ1-6Galβ1-4GlcNAcβ-sp, Galβ1-4GlcNAcβ1-3(Galβ1-4GlcNAcβ1-
6)Galβ1-4GlcNAcβ-sp, GlcNAcβ1-3(Galβ1-4GlcNAcβ1-6)Galβ1-4GlcNAcβ-sp, and Galβ1-4GlcNAcβ1-
3(GlcNAcβ1-6)Galβ1-4GlcNAcβ-sp (sp = O(CH2)2NH2). They represent N-acetyllactosamines substituted with
N-acetylglycosamine or N-acetyllalctosamine residue at O3, O6, or at both positions of galactose. Glycosyla-
tion was achieved by coupling with N-trichloroethoxycarbonyl-protected glucosamine bromide in the presence
of silver triflate.
Key words: oligo-N-acetyllactosamines, oligosaccharide synthesis
DOI: 10.1134/S1068162007010141
INTRODUCTION
Galβ1-4GlcNAcβ1-6
Galβ1-4GlcNAcβ-sp,
(IV)
GlcNAcβ1-3
Oligolactosamine and poly(lactosamine) fragments
are the characteristic elements of architecture of com-
plex N-chains of glycoproteins.2 The specific interac-
tion with proteins of galectin family is one of the func-
tions of oligolactosamines [2]. The carbohydrate spec-
ificity was determined only for several galectins of the
family consisting of 14 proteins. Even those galectins
(first of all, galectin-1 and -3) that have been investi-
gated better than others require additional studies, as
there is no total consensus on their carbohydrate speci-
ficity in the studies reported. The further studies of
galectins require new molecular tools, such as glyco-
conjugates that are usually synthesized from oligosac-
charides. We describe in this work the synthesis of a
series of linear and branched oligolactosamines, which
are used in the studies of galectins in the form of indi-
vidual soluble proteins and also as constituents of nor-
mal and transformed cells [3].
GlcNAcβ1-6
Galβ1-4GlcNAcβ1-3
Galβ1-4GlcNAcβ1-3Galβ1-4GlcNAcβ-sp,
Galβ1-4GlcNAcβ-sp,
(V)
(VI)
Galβ1-4GlcNAcβ1-6Galβ1-4GlcNAcβ-sp, (VII)
Galβ1-4GlcNAcβ1-6
Galβ1-4GlcNAcβ-sp,
(VIII)
Galβ1-4GlcNAcβ1-3
sp = O(CH2)2NH2.
RESULTS AND DISCUSSION
The goal of this study is the synthesis of oligosac-
charides: two linear oligolactosamines (VI) and (VII),
branched trilactosamine (VIII), and also their agalacto
analogues (I)–(V). Glycosylation with N-Troc deriva-
tive of lactosamine or glucosamine by the Koenigs–
Knorr method that allows the reaction to proceed ste-
reospecifically and in a high yield [4] is a key stage in
the synthesis. It should also be noted that the transfor-
mation of the TrocNH-fragment to the AcNH moiety
does not involve other protective groups [4]. The use of
these donors in the Koenigs–Knorr reaction has already
been approved in the synthesis of another series of oli-
gosaccharides, the fragments of glycoprotein é-chains
[5, 6].
(I)
(II)
GlcNAcβ1-3Galβ1-4GlcNAcβ-sp,
GlcNAcβ1-6Galβ1-4GlcNAcβ-sp,
GlcNAcβ1-6
Galβ1-4GlcNAcβ-sp,
GlcNAcβ1-3
(III)
1
Corresponding author; phone: +7(495) 330-7138; fax: +7(495)
330-5592; e-mail: bovin@carbohydrate.ru.
Abbreviations: AgOTf, silver triflate; Bn, benzyl; LacNAc,
N-acetyllactosamine; TMM, tetramethylurea; and Troc, 2,2,2-
trichloroethoxycarbonyl.
2
2-Azidoethyl-spacered lactosamine (IX) [7] was
used as the starting compound for the construction of
122

SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
123
OH
OH
OH
OH
HO
HO
HO
HO
a, b
O
O
O
O
O
O
O
HO
O
HO
N3
NHCOCF
3
OH
NHAc
OH
NHAc
(IX)
(X)
c, d, e, d
OH
OBn
OAc
OAc
AcO
AcO
AcO
a
O
O
O
O
O
O
O
O
AcO
AcO
OAc
NHCOCF3
AcO
OAc
NHCOCF3
NHAc
NHAc
(XII)
(XI)
f, g, d, h
OBn
OAc
AcO
HO
O
O
O
O
AcO
OAc
NHCOCF3
NHAc
(XIII)
Scheme 1. Reagents: ‡: 10% Pd/C, H ; b: CF COOCH , NEt , MeOH; c: PhCH(OCH ) , TsOH, MeCN; d: Ac O/Py;
2
3
3
3
3
3 2
2
e: NaCNBH , HCl/Et O; f: MeONa/MeOH; g: CH C(OEt) , TsOH, MeCN; h: 80% AcOH.
3
2
3
the core part of oligosaccharides (Scheme 1). Hydroge- in 84% yield. A comparison of the 1H NMR spectrum
nation of the azide group resulted in an amine, which of product (XI) with that of its benzylidene precursor
was protected with the N-trifluoroacetyl group by the proved the opening of the benzylidene acetal with the
treatment with methyl trifluoroacetate in methanol [8]. formation of 6-é-benzyl ether. The signal of the proton
The substitution of 2-trifluoroacetamidoethyl spacer at C4 of galactose was shifted downfield (δ 5.460 ppm
for the azidoethyl moiety was executed at the initial in lactosamine (XI) and 4.369 ppm in the benzylidene
step of the synthesis, because the 2-trifluoroacetamido- derivative), whereas the signals of H6' and H6" were
ethyl protective group is stable in the further synthetic shifted upfield (δ 3.521 and 3.437 ppm) as compared to
procedures, particularly, at hydrogenolysis and, at the those at δ 4.068 and 4.301 ppm in the benzylidene pre-
same time, can be quantitatively removed at the final cursor. Furthermore, the signals of benzyl protons
step by an aqueous base.
(δ 4.533 and 4.425 ppm) in the form of two doublets
with the coupling constant Jhem 11.7 Hz were observed.
Of two possible strategies for the creation of branch-
ing at the galactose residue, we have chosen the
approach that included glycosylation of lactosamine
first at position 3 of the galactose residue and then at
position 6, because the primary hydroxy group is more
reactive than the secondary hydroxy and its effective
glycosylation is possible in the presence of the bulky
substituent already introduced in position 3.
Compound (XI) was deacetylated by Zemplen in
86% yield. Introduction of the 3',4'-orthoester protec-
tive group [10] with triethyl orthoacetate in the pres-
ence of p-toluenesulfonic acid in acetonitrile followed
by acetylation and opening of the orthoester cycle with
acetic acid permitted the obtaining of derivative (XIII)
in 70% yield. The presence of free OH group at C3 of
The benzyl group obtained from 4,6-é-benzylidene the galactose residue is indicated by an upfield shift of
derivative was chosen for the selective protection of the signal of H3 (δ 3.810 ppm, J3,éç 6.1 Hz) as com-
hydroxyl at C6 of galactose residue. The benzylidene pared to that in (XI) acetylated at C3 of galactose (δ
protective group was introduced into lactosamine (X) 4.985 ppm). The lactosamine derivative with the free
by treatment with dimethoxytoluene in the presence of OH group in position 4 of galactose (δ 4.147 ppm, H4)
p-toluenesulfonic acid. The resulting benzylidene and acetyl group at C3 (δ 4.896 ppm, H3), which was
derivative was acetylated with acetic anhydride in pyri- apparently generated upon the chromatography on sil-
dine without additional purification; the total yield after ica gel due to migration of the acetyl group (it was not
the two steps was 87%. The use of sodium cyanoboro- observed in the reaction mixture according to TLC),
hydride [9] allowed a selective opening of the ben- was the main by-product of this transformation. The
zylidene cycle and, thus, the obtaining of the necessary proportion of the 4-OH derivative can be decreased by
derivative with the benzyl protective group at C6. the addition of 0.5% of pyridine in the eluent for chro-
Acetylation and chromatographic isolation led to (XI) matography.
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

124
SEVEROV et al.
OAc
AcO
AcO
O
+ (XIII)
Br
NHTroc
(XIV)
a
OAc
OAc
OBn
OBn
OH
AcO
O
AcO
AcO
O
O
O
O
O
AcO
O
O
NHCOCF3
(XV) (58%)
NHAc
NHTroc
OAc
b
OAc
OAc
OAc
OAc
AcO
O
AcO
AcO
O
O
AcO
O
O
NHCOCF3
(XVI) (74%)
NHAc
NHAc
OAc
c
AcO
O
AcO
AcO
O
O
AcO
O
O
NHCOCF3
(XVII) (85%)
NHAc
NHAc
OAc
d
OAc
OAc
OAc
AcO
O
AcO
AcO
O
O
O
O
O
AcO
NHCOCF3
NHAc
NHAc
OAc
(XVIII) (85%)
e
(I) (98%)
Scheme 2. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: 10% Pd/C, H ; d: Ac O/Py;
2
2
2
2
e: MeONa/MeOH, then NaOH, H O.
2
Oligosaccharides (I), (III), and (IV) with the yield of the target isomer (XV) of approximately 30%
GlcNAcβ(1-3')LacNAc fragment were obtained from after acetylation and 50% recovery of disaccharide
(XIII) by its glycosylation with the bromide of glu- (XIII) in the form of 3-acetate. The use of the trichlo-
cosamine Troc derivative (XIV), which was synthe- roacetimidate of Troc-protected glucosamine as the
sized from the corresponding peracetate by treatment glycosyl donor instead of bromide (XIV) also did not
with hydrogen bromide in acetic acid (Scheme 2). Gly- give the expected result: the yield of compound (XV)
cosylation was carried out in dichloromethane in the was 38%. Thus, the target product was obtained in the
presence of AgOTf, TMM, and molecular sieves 4 Å. highest yield upon glycosylation of glycosyl acceptor
The yield of trisaccharide (XV) was 58%, 38% of the (XIII) bearing one OH group with glycosyl bromide
starting disaccharide (XIII) being recovered. Note that (XIV) using AgOTf as the promoter. The presence of
this result was obtained using twofold excess of donor glycosyl substituent at C3 of the Gal residue was con-
(XIV) relative to acceptor (XIII). In the case of (XIV) : firmed by the position of the signal of H3b in the spec-
(XIII) ratio equal 1.5, the yield of (XV) was 46%. Gly- trum of (XV) (δ 3.935 ppm) as compared with the deriv-
cosylation of the corresponding 3,4-diol resulted in the atives bearing OAc group in this position (δ ~ 5.0 ppm),
mixture of mono- (1–3 and 1–4) and diadducts with e.g., δ 4.985 ppm in (XI). The configuration of the ter-
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SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
125
(XIV)
(XVII)
+
a
AcO
AcO
OAc
O
TrocHN
OAc
O
OAc
O
AcO
AcO
AcO
O
O
OAc
O
O
O
O
AcO
NHCOCF3
NHAc
NHAc
OAc
(XIX) (85%)
b
AcO
AcO
O
AcNH
OAc
OAc
O
AcO
O
AcO
AcO
O
O
AcO
O
O
NHCOCF3
NHAc
NHAc
OAc
(XX) (51%)
c
(III) (97%)
Scheme 3. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: MeONa/MeOH, then NaOH, H O.
2
2
2
minal glucosamine unit was confirmed by the value of to C6 of the galactose fragment (~10%) (the value of
coupling constant J1c,2c 8.3 Hz. The presence of Troc the chemical shift of H4 in the product of migration was
moiety in the resulting compound was confirmed by 4.343 ppm, whereas that of trisaccharide (XVII) was
two doublets (δ 4.975 and 4.506 ppm, Jhem 12.5 Hz). 5.488 ppm). The amount of the migration product
increased with the time of hydrogenolysis and chro-
matographic purification.
Glycosylation was hereinafter achieved according to a
similar method varying the excess of a glycosyl donor
within the range of 1.5–2 in dependence on the
expected activity of glycosyl acceptor.
Acetylation of (XVII) resulted in peracetate
(XVIII) in 85% yield. The necessity of this step is sub-
stantiated by the facts that oligosaccharides in the form
of peracetates are convenient for additional chromato-
graphic purification and their 1H NMR spectra are more
informative than those of partially protected carbohy-
drates. The subsequent deacetylation of peracetate
(XVIII) and removal of the N-trifluoroacetyl protective
group resulted in the target trisaccharide (I) in 98%
yield.
Despite of the literature data [11] on the stability of
the Troc protective group under the conditions of
hydrogenolysis in neutral medium, our attempt to
remove the benzyl protective group in (XV) in the pres-
ence of palladium on carbon resulted in a multicompo-
nent mixture. Therefore, we first substituted N-acetyl
protection for the N-Troc protective group by the treat-
ment of (XV) with zinc in acetic acid–acetic anhydride
1
to get product (XVI) in 74% yield. Its H NMR spec-
Glycosylation of trisaccharide (XVII) with the gly-
cosyl bromide of Troc derivative of glucosamine (XIV)
(Scheme 3) and lactosamine (XXI) (Scheme 4) resulted
in tetrasaccharide (XIX) (in 85% yield) and in pen-
trum exhibited no doublets of Troc group; instead, nine
singlets of acetyl groups were present [one more than in
lactosamine (XV)]. The subsequent hydrogenolysis of
benzyl derivative (XVI) resulted in the formation of tasaccaride (XXII) (in 65% yield), respectively. Com-
(XVII), containing one OH group at C6 of galactose pound (XIX) that appeared to be homogeneous accord-
unit, in 85% yield. We also isolated from the reaction ing to TLC contained in fact ~20% of impurity accord-
1
mixture a product of migration of acetyl group from C4 ing to H NMR spectroscopy. In this and subsequent
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126
SEVEROV et al.
OAc
OAc
AcO
AcO
O
O
+
O
Br
(XVII)
AcO
OAc
NHTroc
(XXI)
OAc
AcO
a
AcO
O
O
AcO
AcO
OAc
O
TrocHN
OAc
OAc
O
AcO
O
AcO
AcO
O
O
O
O
O
AcO
NHCOCF3
NHAc
NHAc
OAc
OAc
(XXII) (65%)
AcO
b
AcO
O
O
AcO
OAc
AcO
OAc
O
AcHN
OAc
O
AcO
O
AcO
AcO
O
O
O
O
O
AcO
NHCOCF3
NHAc
NHAc
OAc
(XXIII) (49%)
c
(IV) (95%)
Scheme 4. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: MeONa/MeOH, then NaOH, H O.
2
2
2
cases, the total assigning of the signals in the 1H NMR glycosyl bromide of lactosamine Troc derivative (XXI)
spectra of oligosaccharide Troc-derivatives was not
achieved, since these compounds usually contained
impurities (mainly α-anomers) and, furthermore, the
spectra of N-acetyl derivatives are easier to interpret.
Then, the N-acetyl protective group was substituted for
Troc by treatment with zinc in acetic acid–acetic anhy-
dride to give peracetates (XX) and (XXIII) in 51 and
59% yields, respectively.
(Scheme 5) to give tetrasaccharide (XXIV) in 59%
yield. The subsequent substitution of the N-acetyl pro-
tective group for Troc moiety resulted in (XXV) in 73%
yield. Its 1H NMR spectrum corresponded to the struc-
ture expected: the four H1 signals were characterized
by the coupling constants J1, 2 of 7.8–8.3 Hz, the forma-
tion of the GlcN(1-3)Gal links was confirmed by the
position of H3 (δ 3.67–3.83 ppm) in the glycosylated
galactose residue (see above).
Deacetylation and removal of N-trifluoroacetyl pro-
tective group resulted in the target compounds (III) and
(IV) in 97 and 95% yields, respectively.
Compound (XXV) was hydrogenolyzed to (XXVI)
with a free OH group at C6 of the internal galactose res-
Oligosaccharides (V), (VI), and (VIII) containing
the LacNAcβ(1–3')LacNAc fragment were obtained as
1
idue in 73% yield, which was proved by the H NMR
follows. Compound (XIII) was glycosylated with the spectroscopy: the signals of H6' and H6"[a5] are shifted
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SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
127
(XXI)
(XIII)
+
a
OBn
OAc
OAc
OAc
OAc
OAc
OAc
AcO
AcO
AcO
O
O
O
O
O
O
O
O
AcO
NHCOCF3
AcO
OAc
NHAc
OAc
NHTroc
(XXIV) (59%)
b
OBn
AcO
AcO
AcO
O
O
O
O
O
O
O
O
AcO
NHCOCF3
AcO
OAc
NHAc
OAc
NHAc
(XXV) (73%)
c
OH
OAc
OAc
OAc
OAc
OAc
OAc
AcO
AcO
AcO
O
O
O
O
O
O
O
O
AcO
NHCOCF3
AcO
OAc
NHAc
OAc
NHAc
d
(XXVI) (73%)
OAc
AcO
AcO
AcO
O
O
O
O
O
O
O
O
AcO
NHCOCF3
AcO
OAc
NHAc
OAc
NHAc
(XXVII) (90%)
e
(VI) (96%)
Scheme 5. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: 10% Pd/C, H ; d: Ac O/Py;
2
2
2
2
e: MeONa/MeOH, then NaOH, H O.
2
downfield (δ 3.55–3.77 ppm) as compared to those of gosaccharides (V) and (VIII) in 92 and 93% yield,
respectively.
the benzyl derivative (XXV) (δ 3.43–3.51 ppm).
To obtain lactosamine derivatives (II) and (VII)
Acetylation of (XXVI) gave peracetate (XXVII) in
90% yield. The subsequent deacetylation and removal
of N-trifluoroacetyl protective group resulted in tet-
rasaccharide (VI) in 96% yield.
nonsubstituted in position-3 of the galactose unit, lac-
tosamine derivative (XI) was hydrogenolyzed to disac-
charide (XII) (see Scheme 1) with OH group at C6 of
the terminal galactose unit (H4, δ 5.557; H6', δ 3.799;
and H6", δ 3.655 ppm) in 69% yield. The lactosamine
derivative with OH group at C4 of the galactose residue
(H4, δ 4.152 ppm, H6' and H6", δ 4.399 ppm) was
obtained as a by-product in 14% yield.
Glycosylation of (XXVI) with the glycosyl bromide
of Troc derivative of glucosamine (XIV) (Scheme 6)
and lactosamine (XXI) (Scheme 7) afforded pentasac-
caride (XXVIII) in 50% yield and hexasaccharide
(XXX) in 53% yield, respectively. The subsequent sub-
stitution of N-acetyl protective group for the Troc pro-
tective group resulted in peracetate (XXIX) in 58%
yield and peracetate (XXXI) in 46% yield, respectively.
Deacetylation of peracetates and removal of N-trifluo-
Glycosylation of lactosamine derivative (XII) with
glycosyl bromide (XIV) resulted in compound
(XXXII) in 79% yield (the purity was ~80% according
1
to H NMR spectroscopy) (Scheme 8). Glycosylation
of compound (XII) with glycosyl bromide (XXI)
roacetyl protective group resulted in the target oli- resulted in (XXXIV) in 61% yield (Scheme 9). The fur-
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128
SEVEROV et al.
(XIV) (XXVI)
+
a
AcO
AcO
OAc
O
TrocHN
OAc
OAc
OAc
O
AcO
AcO
AcO
O
O
O
O
O
AcO
O
O
O
AcO
NHCOCF3
OAc
NHAc
NHAc
OAc
(XXVIII) (50%)
b
AcO
AcO
OAc
O
AcHN
OAc
OAc
OAc
O
AcO
AcO
AcO
O
O
O
O
O
AcO
O
O
O
AcO
NHCOCF3
OAc
NHAc
NHAc
OAc
(XXIX) (58%)
c
(V) (92%)
Scheme 6. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: MeONa/MeOH, then NaOH, H O.
2
2
2
ther substitution of the N-acetyl protective group for the osylation are due to a low conversion degree rather than
Troc moiety resulted in peracetates (XXXIII) and side processes taking place (taking into account the
(XXXV) in 59 and 68% yield, respectively. Deacetyla- recovery of glycosyl acceptors, the yields exceed 80%).
tion of peracetates and removal of the N-trifluoroacetyl
protective group gave the target oligosaccharides (II)
and (VII) in 99 and 95% yield, respectively.
The strategy of block synthesis used in this study is
one of the most convenient approaches to the synthesis
of complex oligosaccharides. This approach is espe-
cially effective for the synthesis of a series of structures
containing an identical fragment. As compared to the
strategy of sequential chain lengthening, this method
allows obtaining the same oligosaccharides in a lesser
number of synthetic steps [12].
The structures of the compounds were confirmed by
1
mass spectrometry and H NMR spectroscopy. The
total assignment of the signals for the peracetates of all
oligosaccharides synthesized was achieved by 2D-1H,
1H-COSY experiments.
Thus, glycosylation with lactosamine block (XXI)
is a convenient method for the synthesis of oligolac-
tosamines. Synthesis of some linear lactosamines is
reported in [13–16]. We have herein demonstrated that
this approach also gives good results for the synthesis
of the branched oligolactosamine chains.
We should note that the glycosylation was not β-ste-
reospecific. Glycosylation of the hydroxy group at C6
of the galactose residue of lactosamine (XII) with glu-
cosamine (XIV) resulted in the formation of 18% of α-
anomer, whereas the use of lactosamine (XXI) as the
glycosyl donor gave 7% of the α-anomer. Glycosyla-
tion of the hydroxyl group at C3 of lactosamine (XIII)
with glucosamine (XIV) yielded 5% of α-anomer. In
other cases, α-anomers were generated in the amounts
not exceeding 2%. The separation of the anomer mix-
EXPERIMENTAL
Values of optical rotation were measured on a Jasco
ture was more effective after the substitution of the N- DIP-360 digital polarimeter at 25°ë. 1H NMR spectra
acetyl protective group for the Troc protective group. were registered on a Bruker WM spectrometer
Also note that low yields of the target products of glyc- (500 MHz) at 25°ë. Values of chemical shifts (δ, ppm)
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SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
129
(XXI)
OAc
(XXVI)
+
a
AcO
AcO
O
O
AcO
AcO
OAc
O
TrocHN
OAc
OAc
OAc
O
AcO
AcO
AcO
O
OAc
O
O
O
O
AcO
O
O
O
AcO
NHCOCF
3
NHAc
NHAc
OAc
OAc
(XXX) (53%)
b
AcO
AcO
O
O
AcO
AcO
OAc
O
AcHN
OAc
OAc
OAc
O
AcO
AcO
AcO
O
OAc
O
O
O
O
AcO
O
O
O
AcO
NHCOCF
3
NHAc
NHAc
OAc
(XXXI) (46%)
c
(VIII) (93%)
Scheme 7. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: MeONa/MeOH, then NaOH, H O.
2
2
2
are given using residual signals of solvents: D2O Solvents were removed in a vacuum at 30–40°ë. Sol-
vents for glycoside synthesis were dried by distillation
from drying agents and stored over molecular sieves.
Solid reagents were dried for 2 h in a vacuum (0.1 mm
Hg) at 20–40°ë.
(δ 4.750), CDCl3 (δ 7.270), or DMSO-d6 (δ 2.500) as
internal standards; values of coupling constants are
given in Hz. Signals in 1H NMR spectra were assigned
using the technique of the suppression of spin coupling
(double resonance) and 2D-1H,1H-COSY experiments.
Mass spectra were registered on a Vision-2000
(Thermo Bioanalysis, UK) MALDI TOF mass spec-
trometer using dihydroxybenzoic acid as a matrix. TLC
was carried out on silica gel 60 precoated plates
(Merck, Germany). The spots were visualized by spray-
ing with 5% aqueous orthophosphoric acid and subse-
quent heating at 150°ë in the case of carbohydrates or
by soaking in a ninhydrin solution (3 g/l in 30 : 1
butanol–acetic acid) in the case of amines. Column
chromatography was carried out on silica gel 60
(0.040–0.063 µm, Merck, Germany) and gel chroma-
Deacetylation was executed according to Zemplen
in anhydrous methanol by addition of 2 M sodium
methylate in methanol up to pH 8−9 to a solution of
acetylated compound. After the reaction was over, Na+
ions were removed with cation exchange resin Dowex
50X-400 (H+ form, Acros, Belgium); the solution was
evaporated in a vacuum.
Glycosyl bromide (XIV). Acetyl bromide
(0.256 ml, 3.2 mmol) was added to a solution of per-
acetylated Troc derivative of glucosamine (0.21 mmol)
in anhydrous dichloromethane (2 ml) and acetic acid
(0.5 ml), the mixture was cooled to 0°ë, and methanol
tography, on Sephadex LH-20 (Pharmacia, Sweden). (0.128 ml, 3.2 mmol) was added. The reaction mixture
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

130
SEVEROV et al.
(XIV)
(XII)
+
a
AcO
AcO
OAc
O
TrocHN
OAc
O
AcO
AcO
O
O
O
O
AcO
NHCOCF3
NHAc
OAc
(XXXII) (79%)
AcO
AcO
b
OAc
O
AcHN
OAc
O
AcO
AcO
O
O
OAc
O
O
AcO
NHCOCF3
NHAc
(XXXIII) (59%)
c
(II) (99%)
Scheme 8. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH, c: MeONa/MeOH, then NaOH, H O.
2
2
2
was kept for 1 h at room temperature and poured on product was isolated by column chromatography on si-
crashed ice. The organic layer was diluted with chloro- lica gel.
form; subsequently washed with water, saturated
Acetylation was carried out by treatment with 2 : 1
sodium bicarbonate, and water; filtered through a cot-
pyridine–acetic anhydride at 20°ë for 12–24 h. After
ton layer; concentrated in a vacuum; and coevaporated
the reaction was over, the reagents were removed by
with toluene. The residue was used for glycosylation
coevaporation with toluene.
without additional purification on the same day.
Hydrogenolysis was achieved in methanol or 1 : 1
Glycosyl bromide (XXI). Bromine (18 µl, methanol–water at atmospheric pressure for 1–3 h in the
0.37 mmol) was added to a solution of thioglycoside of presence of 10% Pd/C (Merck, Germany). The catalyst
peracetylated Troc derivative of lactosamine (270 mg, was taken at a catalyst–substrate mass ratio of 2 : 1.
0.332 mmol) in anhydrous dichloromethane (10 ml)
Removal of Troc group and N-acetylation.
cooled to 0°ë. The reaction mixture was kept for 1 h at
Freshly activated zinc powder (4 g) and then thriethy-
0°ë, concentrated, coevaporated with toluene, and used
lamine (0.5 ml) were added to a solution of a Troc
without additional purification on the same day, consid-
derivative (0.5 mmol) in acetic acid (40 ml) and acetic
ering the yield to be quantitative; Rf 0.60 (10 : 1 chlo-
anhydride (2 ml). The reaction mixture was stirred for
roform–isopropanol).
20–30 h and filtered, the solvents were removed, and
the residue was chromatographed on a Sephadex LH-
A general glycosylation procedure. A mixture of a
20 column (elution with 1 : 1 chloroform–methanol).
glycosyl acceptor (0.1 mmol), AgOTf (0.15 mmol),
TMM (0.15 mmol), and freshly calcined molecular
Deacetylation with simultaneous removal of the
sieves 4Å (300 mg) in anhydrous dichloromethane N-trifluoroacetyl protective group. Sodium methy-
(5 ml) was stirred for 30 min at room temperature in the late (50 µl of 2 M solution in methanol) was added to a
dark. Additional portion of molecular sieves 4Å solution of a peracetate (0.05 mmol) in anhydrous
(100 mg) was added and a solution of a glycosyl bro- methanol (2 ml). The solution was kept for 1 h and con-
mide (0.15 mmol) in anhydrous dichloromethane (2 ml) centrated in a vacuum. Water (2 ml) was added, and the
was added. The mixture was stirred for 15–20 h and fil- mixture was kept for 3 h and then applied onto a col-
tered, the filtrate was concentrated in a vacuum, and the umn with cation exchange resin Dowex 50 (H+). The
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
131
(XXI)
(XII)
+
a
OAc
O
AcO
AcO
O
AcO
AcO
OAc
O
TrocHN
OAc
O
AcO
AcO
O
O
O
O
AcO
NHCOCF3
NHAc
OAc
(XXXIV) (61%)
OAc
O
AcO
b
AcO
O
AcO
AcO
OAc
O
AcHN
AcO
AcO
OAc
O
O
O
OAc
O
O
AcO
NHCOCF
3
NHAc
(XXXV) (68%)
c
(VII) (95%)
Scheme 9. Reagents: a: AgOTf, TMM, molecular sieves 4Å, CH Cl ; b: Zn, AcOH; c: MeONa/MeOH, then NaOH, H O.
2
2
2
target compound was eluted with 1 M ammonia, the added, and the mixture was stirred for 2.5 h until the
eluate was concentrated in a vacuum, and the residue negative test for amine. The reaction mixture was
was lyophilized.
diluted with anhydrous methanol (50 ml) and quenched
with Dowex 50 (ç+) up to the neutral reaction of
medium. The solution was filtered and then concen-
trated in a vacuum. The residue was purified by gel fil-
tration on Sephadex LH-20 (elution with 0.5% aqueous
acetic acid) to give 3.91 (85%) of (X) (85%), Rf 0.89
(3 : 1 : 1 : 1 ethanol–water–pyridine–acetic acid); Rf
0.40 (2 : 3 : 1 isopropanol–ethanol–water); [α]D –15.8
(Ò 1, water); MS, m/z: 545 (522 + 23) M+ + Na+.
2-Trifluoroacetamidoethyl 2-acetamido-2-deoxy-
4-é-(b-D-galactopyranosyl)-b-D-glucopyranoside
(X). A solution of azide (IX) (4.82 g, 10.65 mmol) in
1 : 1 methanol–water (150 ml) was subjected to hydro-
genolysis over 10% Pd/C (2.4 g) for 1.5 h. The reaction
mixture was then filtered; the cation exchange resin
Dowex 50 (H+) (35 ml) was added to the filtrate. Then
the resin was filtered off and washed with 1 : 1 metha-
nol–water (4 × 20 ml). The product was eluted with 1 M
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
ammonia, and the eluate was concentrated in a vacuum é-acetyl-2-deoxy-4-é-(2,3,4-tri-é-acetyl-6-é-benzyl-
to give 3.77 g (8.84 mmol, 83%) of 2-aminoethyl deriv- b-D-galactopyranosyl)-b-D-glucopyranoside
(XI).
ative of N-acetyllactosamine; Rf 0.60 (3 : 1 : 1 : 1 etha- α,α-Dimethoxytoluene (2.2 ml, 12 mmol) and
nol–water–pyridine–acetic acid). The resulting product p-toluenesulfonic acid (0.63 g) (to pH 2–3) were added
was dissolved in anhydrous methanol (150 ml), methyl to a solution of disaccharide (X) (3.91 g, 7.48 mmol) in
trifluoroacetate (6 ml) and thriethylamine (2 ml) were anhydrous acetonitrile (200 ml). A gradual precipita-
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

132
SEVEROV et al.
tion of the product was observed. After 24 h, the reac-
tion mixture was quenched with pyridine, concentrated,
and coevaporated with toluene. The dry residue was
acetylated with 2 : 1 pyridine–acetic anhydride (45 ml).
Chromatography on silica gel (elution with 4 : 2 : 1 hex-
ane–chloroform–isopropanol) resulted in 5.08 g (87%)
of 2-trifluoroacetamidoethyl 2-acetamido-3,6-di-O-
acetyl-2-deoxy-4-O-(2,3-di-O-acetyl-4,6-O-benzylidene-
β-D-galactopyranosyl)-β-D-glucopyranoside; Rf 0.21
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
O-acetyl-2-deoxy-4-é-(2,4-di-é-acetyl-6-é-benzyl-
b-D-galactopyranosyl)-b-D-glucopyranoside (XIII).
Lactosamine derivative (XI) (4.13 g, 5 mmol) was
deacetylated by Zemplen. Anhydrous acetonitrile
(100 ml), triethyl orthoacetate (1.2 ml, 6.5 mmol), and
p-toluenesulfonic acid (150 mg) were added to the
deacetylation product. The mixture was stirred for 1 h
at room temperature (the mixture becomes homoge-
nous after 10 min) and quenched with pyridine. The
solution was concentrated, the resulting orthoacetate
was acetylated with 2 : 1 pyridine–acetic anhydride
(30 ml), and the resulting compound was treated with
80% aqueous acetic acid (10 ml). After 4 h, the solution
was concentrated in a vacuum. Chromatography on sil-
ica gel (elution with 10–20% isopropanol in chloro-
form) resulted in 2.34 g (70%) of the product of the
orthoacetate opening (XIII); Rf 0.29 (10 : 1 chloro-
form–methanol); [α]D –17.8 (Ò 1, CHCl3); MS, m/z:
1
(4 : 2 : 1 hexane–chloroform–isopropanol); H NMR
(CDCl3): 1.952, 2.051, 2.054, 2.099, and 2.120 (5 × 3
H, 5 s, 5 Ac), 3.414–3.584 (4 H, m, CH2N, H5a, H5b),
3.686 (1 H, m, OCH), 3.797 (1 H, dd ≈ t, J3,4 8.8, J4,5
9.1, H4a), 3.824 (1 H, m, OCH), 4.068 (1 H, dd, J5,6'
1.5, J6',6" 12.7, H6'b), 4.082–4.159 (2 H, m, H6'a, H2a),
4.301 (1 H, dd, J5,6'' 1.5, J6',6'' 12.5, H6"b), 4.369 (1 H,
dd ≈ d, J3,4 3.4, J4,5 < 1, H4b), 4.433 (1 H, d, J1,2 8.1,
H1a), 4.501 (1 H, d, J1,2 7.8, H1b), 4.546 (1 H, dd, J5,6''
2.0, J6',6'' 11.9, H6"a), 4.914 (1 H, dd, J2,3 10.4, J3,4 3.4,
H3b), 5.093 (1 H, dd, J2,3 10.3, J3,4 8.8, H3a), 5.234 (1
H, dd, J1,2 7.8, J2,3 10.4, H2b), 5.495 (1 H, s, PhCH),
6.291 (1 H, d, J2,NH 9.1, NHAc), 7.179(1 H, m,
NHCOCF3), 7.376–7.389 (3 H, m, Ph), 7.442–7.461
(2 H, m, Ph).
1
802 (780 + 23) M + + Na+; H NMR (CDCl3): 1.959,
2.012, 2.096, 2.102, and 2.131 (5 × 3 H, 5 s, 5 Ac),
2.537 (1 H, d, J3,OH 6.1, 3-OH), 3.42–3.50 (1 H, m,
CHN), 3.472 (1 H, dd, J5,6' 6.7, J6',6'' 9.5, H6'b), 3.524 (1
H, dd, J5,6'' 5.9, J6',6'' 9.5, H6"b), 3.59–3.67 (2 H, m,
CHN, H5a), 3.71–3.84 (4 H, m, H5b, OCH, H4a, H3b),
3.881 (1 H, m, OCH), 4.040 (1 H, ddd, J1,2 7.8, J2,3 10.0,
J2,NH 9.0, H2a), 4.141 (1 H, dd, J5,6' 5.4, J6',6'' 12.0,
H6'a), 4.414 (1 H, d, J1,2 7.8, H1a), 4.435 (1 H, d, J1,2
8.1, H1b), 4.454 and 4.522 (2 × 1 H, 2 d, JAB 11.7,
CH2Ph), 4.501 (1 H, dd, J5,6'' 2.2, J6',6'' 11.7, H6"a),
4.872 (1 H, dd, J1,2 8.1, J2,3 9.8, H2b), 5.036 (1 H, dd,
J2,3 10.0, J3,4 8.8, H3a), 5.383 (1 H, dd ≈ d, J3,4 3.7,
A mixture of the resulting disaccharide (4.67 g,
6 mmol) and freshly calcined molecular sieves 3Å
(12 g) in anhydrous THF (100 ml) was stirred for
30 min. Then sodium cyanoborohydride (3.77 g,
60 mmol) was added portionwise, the saturated HCl in
anhydrous Et2O was added dropwise for 1 h (up to pH
2–3), and the mixture was stirred for 3 h. The reaction
mixture was quenched with thriethylamine and filtered,
the precipitate was washed with chloroform, and the
combined filtrate was concentrated in a vacuum. The
dry residue was dissolved in chloroform (300 ml),
washed with water (2 × 300 ml), dried with sodium sul-
fate, and concentrated. The resulting product was
acetylated with 2 : 1 with pyridine–acetic anhydride
(45 ml). Chromatography on silica gel (elution with 5–
10 % isopropanol in chloroform) resulted in 0.64 g
(8%) of the starting benzylidene derivative and 4.13 g
(84%) of benzyl derivative (XI) (84%). Rf 0.26 (4 : 2 : 1
J
4,5 < 1, H4b), 5.755 (1 H, d, J2,NH 9.0, NHAc), 7.192
(1 H, m, NHCOCF3), 7.29–7.39 (5 H, m, Ph).
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-O-
acetyl-2-deoxy-4-é-[2,4-di-é-acetyl-6-é-benzyl-3-é-
(3,4,6-tri-é-acetyl-2-deoxy-2-(2,2,2-trichloroethoxy-
carbonylamido)-b-D-glucopyranosyl)-b-D-galactopy-
ranosyl]-b-D-glucopyranoside (XV). Glycosylation of
lactosamine derivative (XIII) (390 mg, 0.5 mmol) with
glycosyl bromide (XIV) resulted in 522 mg (1.0 mmol)
of the corresponding peracetate. Chromatography on
silica gel (elution with 3–10% isopropanol in chloro-
form) gave 150 mg (38%) of the starting disaccharide
(XIII) and 360 mg (58%) of trisaccharide (XV); Rf 0.29
1
hexane–chloroform–isopropanol); H NMR (CDCl3):
1.962, 1.975, 2.018, 2.055 (×2), and 2.100 (6 × 3 H, 6
s, 6 Ac), 3.437 (1 H, dd, J5,6' 7.1, J6',6'' 9.3, H6'b), 3.439–
3.491 (1 H, m, CHN), 3.521 (1 H, dd, J5,6'' 5.6, J6',6'' 9.3,
H6"b), 3.595–3.655 (2 H, m, CHN, H5a), 3.733 (1 H,
m, OCH), 3.779–3.821 (2 H, m, H4a, H5b), 3.877 (1 H,
m, OCH), 4.038 (1 H, ddd, J1,2 8.1, J2,3 9.8, J2,NH 8.8,
H2a), 4.099 (1 H, dd, J5,6' 5.1, J6',6'' 12.0, H6'a), 4.408 (1
H, d, J1,2 7.8, H1b), 4.425 and 4.553 (2 × 1 H, 2 d, JAB
11.9, CH2Ph), 4.498 (1 H, d, J1,2 8.1, H1a), 4.501 (1 H,
dd, J5,6'' 2.7, J6',6'' 11.9, H6"a), 4.985 (1 H, dd, J2,3 10.3,
J3,4 3.4, H3b), 5.036 (1 H, dd, J2,3 9.8, J3,4 8.3, H3a),
5.095 (1 H, dd, J1,2 7.8, J2,3 10.3, H2b), 5.460 (1 H, dd
≈d, J3,4 3,4, J4,5 < 1, H4b), 5.701 (1 H, d, J2,NH 8.8,
NHAc), 7.136 (1 H, m, NHCOCF3), 7.255–7.360 (5 H,
m, Ph).
1
(10 % isopropanol in chloroform); H NMR (DMSO-
d6): 1.730, 1.872, 1.881, 1.956, 1.969, 1.997, 2.011, and
2.050 (8 × 3 H, 8 s, 8 Ac), 3.913 (1 H, m, H5b), 3.935
(1 H, dd, J2,3 10.5, J3,4 3.4, H3b), 4.010 (1 H, dd, J5,6'
2.2, J6',6" 12.0, H6'a), 4.026–4.070 (2 H, m, H6'c, H6"c),
4.297 (1 H, dd, J5,6" 1.0, J6',6" 12.0, H6"a), 4.426 and
4.464 (2 × 1 H, 2 d, JAB 12.0, CH2Ph), 4.490 (1 H, d, J1,2
7.8, H1b), 4.506 and 4.975 (2 × 1 H, 2 d, JAB 12.5,
CH2CCl3), 4.546 (1 H, d, J1,2 8.6, H1a), 4.662 (1 H, d,
J
1,2 8.3, H1c), 4.750 (1 H, dd, J1,2 7.8, J2,3 10.5, H2b),
4.796 (1 H, dd ≈t, J3,4 9.8, J4,5 9.3, H4c), 4.908 (1 H, dd,
J
2,3 8.3, J3,4 10.0, H3a), 5.092 (1 H, dd ≈ t, J2,3 10.0, J3,4
9.8, H3c), 5.340 (1 H, dd ≈ d, J3,4 3.4, J4,5 < 1, H4b),
7.268–7.381 (5 H, m, Ph), 7.794 (1 H, d, J2,NH 9.0,
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
133
NHAc), 7.822 (1 H, d, J2,NH 9.3, NHTroc), 9.292 (1 H, (87 mg, 0.085 mmol) and subsequent chromatography
m, NHCOCF3).
on silica gel (elution with 10% isopropanol in ethyl ace-
tate) resulted in 77 mg (85%) of peracetate (XVIII); Rf
0.35 (9 : 1 ethyl acetate–isopropanol); [α]D +15.1 (Ò 1,
chloroform); MS, m/z: 1084 (1061 + 23) M+ + Na+; 1H
NMR (CDCl3): 1.907, 1.962, 2.020, 2.028, 2.076,
2.101 (×2), 2.115, 2.119, and 2.125 (10 × 3 H, 10 s, 10
Ac), 3.315 (1 H, dd, J1,2 8.1, J2,NH 7.8, J2,3 10.5, H2c),
3.491 (1 H, m, CHN), 3.609 (2 H, m, CHN, H5a), 3.677
(1 H, m, H5c), 3.740 (1 H, dd ≈ t, J3,4 8.4, J4,5 8.8, H4a),
3.749 (1 H, m, OCH), 3.801 (1 H, ddd ≈t, H5b), 3.815
(1 H, dd, J3,4 3.5, J2,3 9.9, H3b), 3.893 (1 H, m, OCH),
4.036 (1 H, ddd, H2a), 4.065 (3 H, m, H6"c, H6'b,
H6"b), 4.121 (1 H, dd, J6',6'' 11.9, J5,6'' 5.2, H6"a), 4.393
(2 H, m, H1b, J1,2 7.9, H6'c), 4.444 (1 H, d, J1,2 7.8,
H1a), 4.484 (1 H, dd, J6',6'' 11.0, J5,6' 2.2, H6'a), 5.013
(1 H, dd, J1,2 7.9, J2,3 9.9, H2b), 5.030 (1 H, dd, J2,3 9.9,
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-O-
acetyl-2-deoxy-4-é-[2,4-é-acetyl-6-é-benzyl-3-é-(2-
acetamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopy-
ranosyl)-b-D-galactopyranosyl]-b-D-glucopyrano-
side (XVI). The treatment of Troc derivative (XV)
(517 mg, 0.416 mmol) with zinc in acetic acid and sub-
sequent purification by chromatography on silica gel
(elution with 5–10% methanol in chloroform) resulted
in 343 mg (74%) of N-acetyl derivative (XVI) (74%),
1
Rf 0.31 (10% isopropanol in ethyl acetate); H NMR
(CDCl3): 1.903, 1.957, 2.006, 2.016, 2.023, 2.067,
2.078, and 2.095 (×2) (9 × 3 H, 9Ò, 9Ac), 3.360 (1 H,
ddd, J1,2 8.1, J2,3 10.5, J2,NH 7.8, H2c), 3.483 (2 H, m,
H6'b, H6"b), 3.578–3.751 (6 H, m, CH2N, OCH, H5a,
H5b, H5c), 3.774 (1 H, dd ≈ t, J3,4 ≈ J4,5 ≈ 8.6, H4a),
3.828 (1 H, dd, J2,3 10.0, J3,4 3.7, H3b), 3.879 (1 H, m,
OCH), 4.045 (1 H, ddd, J1,2 7.8, J2,3 9.3, J2,NH 9.0, H2a),
4.096–4.143 (2 H, m, H6'a, H6'c), 4.295 (1 H, dd, J5,6''
2.6, J6',6'' 12.4, H6"c), 4.378 (1 H, d, J1,2 8.1, H1b),
4.397 (1 H, d, J1,2 7.8, H1a), 4.458 (1 H, dd, J5,6'' 2.7,
J6',6'' 12.0, H6"a), 4.473 and 4.507 (2 × 1 H, 2 d, JAB
11.7, CH2Ph), 4.990–5.058 (4 H, m, H1c, H2b, H3a,
H4c), 5.414 (1 H, dd ≈ d, J3,4 3.7, J4,5 < 1, H4b), 5.454
(1 H, dd, J2,3 10.5, J3,4 9.5, H3c), 5.485 (1 H, d, J2,NH
7.8, NHAc-c), 5.806 (1 H, d, J2,NH 9.0, NHAc-a), 7.225
(1 H, m, NHCOCF3), 7.339 (5 H, m, Ph).
J3,4 8.4, H3a), 5.032 (1 H, d, J1,2 7.9, H1c), 5.054 (1 H,
dd ≈ t, J3,4 9.3, J4,5 10.1, H4c), 5.360 (1 H, dd ≈ d, J3,4
3.5, J4,5 < 1, H4b), 5.473 (1 H, dd, J2,3 10.5, J3,4 9.3,
H3a), 5.510 (1 H, d, J2,NH 7.8, NHAc-c), 5.849 (1 H, d,
J2,NH 8.7, NHAc-a), 7.207 (1 H, m, NHCOCF3).
2-Aminoethyl 2-acetamido-2-deoxy4-é-[3-é-(2-
acetamido-2-deoxy-b-D-glucopyranosyl)-b-D-galac-
topyranosyl]-b-D-glucopyranoside (I). De-O-acety-
lation and removal of the N-trifluoroacetamide protec-
tive group in derivative (XVIII) (71 mg, 0.067 mmol)
resulted in 41.3 mg (98%) of trisaccharide (I); Rf 0.56
(4 : 1 : 1 : 1 ethanol–water–pyridine–acetic acid); [α]D
–15.1 (Ò 0.5, water); MS, m/z: 652 (629 + 23) M+ + Na+;
1H NMR (D2O): 2.052 and 2.062 (2 × 3 H, 2 s, 2 Ac),
3.238 (2 H, m, CH2N), 3.42–3.51 (2 H, m), 3.55–3.66
(3 H, m), 3.69–3.83 (9 H, m), 3.854 (1 H, dd, J6',6'' 12.2,
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-O-
acetyl-2-deoxy-4-é-[2,4-di-é-acetyl-3-é-(2-acetamido-
3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopyranosyl)-b-D-
galactopyranosyl]-b-D-glucopyranoside
(XVII).
Hydrogenolysis of 6'-O-benzyl derivative (XVI)
(333 mg, 0.3 mmol) and subsequent chromatography
on silica gel (elution with 7% methanol in chloroform)
resulted in 260 mg (85%) of trisaccharide (XVII);
Rf 0.21 (10% isopropanol in ethyl acetate); MS, m/z:
J5,6'' 5.2, H6"a), 3.911 (2 H, m, OCH, H6'c), 4.010 (1 H,
dd, J6',6'' 12.2, J5,6' 2.0, H6'a), 4.073 (1 H, m, OCH),
4.168 (1 H, dd, J3,4 3.2, J4,5 < 1, H4b), 4.478 (1 H, d, J1,2
8.0, H1b), 4.594 (1 H, d, J1,2 8.3, H1a), 4.701 (1 H, d,
J1,2 8.4, H1c).
1043 (1019 + 23) M+ + Na+; H NMR (3 : 1 CDCl3–
1
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-é-
CD3OD): 2.026, 2.067, 2.149, 2.167, 2.186, 2.233,
2.244, 2.267, and 2.271 (9 × 3 H, 9 s, 9 Ac), 3.56–3.67
(4 H, m, CH2N, H2c, H6'b), 3.69–3.83 (3 H, m, H6"b,
H5a, H5b), 3.804 (1 H, m, OCH), 3.851 (1 H, ddd, J4,5
10.0, J5,6' 2.4, J5,6'' 4.2, H5c), 3.932 (1 H, dd ≈ t, J3,4 8.7,
acetyl-2-deoxy-4-é-{2,4-é-acetyl-3-é-[2-acetamido-
3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopyranosyl]-6-é-
[2-acetamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopy-
ranosyl]-b-Db-D-glucopyranoside (XX). Glycosyla-
tion of trisaccharide (XVII) (177 mg, 0.174 mmol) with
glycosyl bromide (XIV) obtained from the correspond-
ing peracetate (136 mg, 0.261 mmol) and subsequent
chromatography on silica gel (elution with 7% isopro-
panol in ethyl acetate) yielded 219 mg (85%) of tet-
rasaccharide (XIX), Rf 0.44 (10% isopropanol in ethyl
acetate). The substance that seemed to be individual
according to TLC appeared to contain about 20% of
impurity according to 1H NMR spectroscopy. The treat-
ment of Troc derivative (XIX) with zinc in acetic acid
and three chromatographic purifications on silica gel
J4,5 9.3, H4a), 3.973 (1 H, dd, J2,3 10.0, J3,4 3.4, H3b),
4.010 (1 H, m, OCH), 4.086 (1 H, ddd, J1,2 8.1, J2,3 10.3,
J2,NH 9.0, H2a), 4.256 (1 H, dd, J5,6' 5.6, J6',6'' 12.0,
H6'a), 3.955 (1 H, dd, J5,6' 4.2, J6',6'' 12.3, H6'c), 4.381
(1 H, dd, J5,6'' 2.4, J6',6'' 12.3, H6"c), 4.563 (1 H, d, J1,2
7.8, H1b), 4.576 (1 H, d, J1,2 8.1, H1a), 4.602 (1 H, dd,
J
5,6'' 2.3, J6',6'' 12.0, H6"a), 5.086 (1 H, d, J1,2 8.3, H1c),
5.10–5.18 (3 H, m, H4c, H2b, H3a), 5.488 (1 H, dd ≈ d,
3,4 3.4, J4,5 < 1, H4b), 5.510 (1 H, dd, J2,3 10.5, J3,4 9.3,
H3c).
J
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di- (elution with 7% methanol in chloroform, 6 : 1 ethyl
é-acetyl-2-deoxy-4-é-[2,4,6-tri-é-acetyl-3-é-(2- acetate–isopropanol, and then 8 : 1 chloroform–isopro-
acetamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopy- panol) yielded 100 mg (51%) of peracetylated tetrasac-
ranosyl)-b-D-galactopyranosyl]-b-D-glucopyrano-
charide (XX); Rf 0.25 (6 : 1 ethyl acetate–isopropanol);
side (XVIII). Acetylation of trisaccharide (XVII) 1H NMR (3 : 1 CDCl3–CD3OD): 2.019, 2.070, 2.077,
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

134
SEVEROV et al.
2.145, 2.164 (×2), 2.180, 2.186, 2.231 (×2), 2.245, tive (XXII) with zinc in acetic acid and subsequent
2.255, and 2.266 (13 × 3 H, 13 s, 13 Ac), 3.55–3.69 chromatography on silica gel (twofold elution with 10–
(2 H, m, CH2N), 3.605 (1 H, dd, J1,2 8.3, J2,3 10.5, H2c), 20% isopropanol in chloroform and then with 15% iso-
3.682 (1 H, dd, J6',6'' 10.5, J5,6'' 7.0, H6"b), 3.767 (1 H, propanol in ethyl acetate) yielded 124 mg (49%) of per-
m, H5a), 3.79–3.85 (2 H, m, OCH, H5c), 3.813 (1 H, acetate (XXIII); Rf 0.17 (10% isopropanol in ethyl ace-
1
dd, J1,2 8.3, J2,3 10.7, H2d), 3.86–3.93 (3 H, m, H4a, tate); H NMR (3 : 1 CDCl3–CD3OD): 2.024, 2.072,
H5b, H5d), 3.938 (1 H, dd, J6',6'' 10.5, J5,6' 4.6, H6'b), 2.086, 2.126, 2.148, 2.168, 2.183, 2.218 (×2), 2.221,
3.975 (1 H, dd, J3,2 10.0, J3,4 3.7, H3b), 4.018 (1 H, m, 2.230, 2.233, 2.253, 2.266, 2.276, and 2.300 (16 × 3 H,
OCH), 4.077 (1 H, dd, J1,2 8.3, J2,3 10.0, H2a), 4.279 (2 16 s, 16 Ac), 3.57–3.70 (4 H, m, J1,2 8.3, J2,3 10.5,
H, m, H6"a, H6"c), 4.322 (1 H, dd, J6',6'' 12.2, J5,6'' 2.2, CH2N, H6"b, H2c), 3.75–3.98 (10 H, m), 3.995 (1 H,
H6"d), 4.410 (1 H, dd, J6',6'' 12.2, J5,6' 4.7, H6'd), 4.422 dd, J2,3 10.0, J3,4 3.4, H3b), 4.024 (1 H, m, OCH), 4.057
(1 H, dd, J6',6'' 12.2, J5,6' 2.4, H6'c), 4.534 (1 H, d, J1,2 (1 H, dd, J1,2 8.3, J2,3 10.0, H2a), 4.115 (1 H, t, H5a),
7.8, H1b), 4.575 (1 H, dd, J6',6'' 12.0, J5,6' 2.0, H6'a),
4.590 (1 H, d, J1,2 8.3, H1a), 4.952 (1 H, d, J1,2 8.3,
H1d), 5.041 (1 H, d, J1,2 8.3, H1c), 5.094 (1 H, dd, J1,2
7.8, J2,3 10.0, H2b), 5.13–5.20 (3 H, m, H3a, H4c,
H4d), 5.471 (1 H, dd, J3,4 3.7, J4,5 < 1, H4b), 5.498 (1
H, dd, J2,3 10.7, J3,4 9.5, H3d), 5.521 (1 H, dd, J2,3 10.5,
4.23–4.33 (5 H, m), 4.532 (1 H, d, J1,2 8.1, H1b), 4.575
(1 H, dd, J6',6'' 12.2, J5,6' 2.0, H6'a), 4.608 (1 H, d, J1,2
8.1, H1a), 4.727 (1 H, dd, J6',6'' 11.7, J5,6' 2.0, H6'd),
4.744 (1 H, d, J1,2 7.8, H1c), 4.757 (1 H, d, J1,2 8.1,
H1d), 5.043 (1 H, d, J1,2 8.1, H1c), 5.096 (1 H, dd, J1,2
8.1, J2,3 10.0, H2b), 5.153 (1 H, dd, J3,4 9.3, J4,5 9.8,
H4c), 5.157 (1 H, dd, J2,3 10.5, J3,4 3.0, H3e), 5.193 (1
H, dd, J2,3 10.0, J3,4 9.1, H3b), 5.235 (1 H, dd, J1,2 7.8,
J3,4 9.3, H3c).
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{3-é-[2-
J
2,3 10.5, H2a), 5.324 (1 H, dd, J2,3 10.0, J3,4 9.1, H3d),
5.485 (1 H, dd, J3,4 3.4, J4,5 < 1, H4b), 5.497 (1 H, dd,
2,3 10.5, J3,4 9.3, H3c), 5.514 (1 H, dd, J3,4 3.0, J4,5 < 1,
H4a).
acetamido-2-deoxy-b-D-glucopyranosyl]-6-é-[2-ace-
tamido-2-deoxy-b-D-glucopyranosyl]-b-D-galactopy-
ranosyl}-b-D-glucopyranoside (III). De-é-acetylation
and removal of the N-trifluoroacetyl protective group
from oligosaccharide (XX) (100 mg, 0.074 mmol)
yielded 60 mg (97%) of tetrasaccharide (III); Rf 0.57
(3 1 : 1 : 1 ethanol–water–pyridine–acetic acid); [α]D –
16.0 (c 0.5, water); MS, m/z: 855 (832 + 23) M+ + Na+;
1H NMR (D2O): 2.050 and 2.073 (×2) (3 × 3 ç, 3Ò,
3Ac), 3.243 (2 H, m, CH2N), 3.42–3.51 (4 H, m), 3.53–
3.65 (4 H, m), 3.66–4.02 (16 H, m), 4.079 (1 H, m,
OCH), 4.158 (1 H, dd, J3,4 3.2, J4,5 < 1, H4b), 4.471 (1
H, d, J1,2 7.8, H1b), 4.598 (2 H, d, J1,2 8.4, H1a, H1c),
4.696 (1 H, d, J1,2 8.5, H1d).
J
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{3-é-
[2-acetamido-2-deoxy-b-D-glucopyranosyl]-6-O-[2-
acetamido-2-deoxy-4-é-(b-D-galactopyranosyl)-b-
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu-
copyranoside (IV). De-é-acetylation and removal of
the N-trifluoroacetamide protective group from perace-
tate (XXIII) (124 mg, 0.076 mmol) yielded 71.4 mg
(95%) of pentasaccharide (IV); Rf 0.54 (3 : 1 : 1 :1 eth-
anol–water–pyridine–acetic acid); [α]D –11.8 (Ò 1,
water); MS, m/z: 1018 (995 + 23) M+ + Na+; 1H NMR
(D2O): 2.043 and 2.063 (×2) (3 × 3 ç, 3Ò, 3ÄÒ), 3.235
(2 H, m, CH2N), 3.42–3.46 (2 H, m), 3.53–3.64 (5 H,
m), 3.66–4.02 (23 H, m), 4.073 (1 H, m, OCH), 4.153
(1 H, dd, J3,4 3.4, J4,5 < 1, H4b), 4.466 (1 H, d, J1,2 7.8,
H1b), 4.475 (1 H, d, J1,2 7.8, H1e), 4.589 (1 H, d, J1,2
8.2, H1a), 4.656 (1 H, d, J1,2 7.5, H1d), 4.690 (1 H, d,
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-3-é-[2-ace-
tamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopyra-
nosyl]-6-é-[3,6-di-é-acetyl-2-deoxy-2-(2,2,2-trichlo-
roethoxycarbonylamido)-4-é-(2,3,4,6-tetra-é-acetyl-
b-D-galactopyranosyl)-b-D-glucopyranosyl]-b-D-
J1,2 8.4, H1c).
galactopyranosyl}-b-D-glucopyranoside
(XXII).
Glycosylation of trisaccharide (XVII) (222 mg,
0.218 mmol) with glycosyl bromide (XXI) obtained
from the corresponding thioglycoside (270 mg, 0.332
mmol) and subsequent chromatography on silica gel
(elution with 5–15% isopropanol in chloroform)
resulted in 57 mg (26%) of a mixture of starting glyco-
syl acceptor (XVII) with its isomer (the product of
migration of the OAc group from C4 to C6 of the galac-
tose unit) and 250 mg (65%) of pentasaccaride (XXII);
Rf 0.37 (10% isopropanol in ethyl acetate).
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
O-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-6-é-benzyl-
3-é-[3,6-di-é-acetyl-2-deoxy-2-(2,2,2-trichloroet-
hoxycarbonylamido)-4-é-(2,3,4,6-tetra-é-acetyl-b-
D-galactopyranosyl)-b-D-glucopyranosyl]-b-D-galac-
topyranosyl}-b-D-glucopyranoside (XXIV). Glyco-
sylation of lactosamine derivative (XIII) (516 mg,
0.66 mmol) with glycosyl bromide (XXI) obtained
from the corresponding thioglycoside (1.073 g, 1.32
mmol) and two subsequent chromatographic purifica-
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di- tions on silica gel (elution with 6–8% isopropanol in
é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-3-é-[2-ace- chloroform and 1–10 % isopropanol in chloroform)
tamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopyrano- yielded 119 mg (23%) of the starting disaccharide
syl]-6-é-[2-acetamido-3,6-di-é-acetyl-2-deoxy-44- (XIII), 48 mg (9%) of its isomer (the product of migra-
é-(2,3,4,6-tetra-O-acetyl-b-D-galactopyranosyl)-b- tion of OAc group from C4 to C3 of the galactose unit)
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu- and 601 mg (59%) of tetrasaccharide (XXIV); Rf 0.44
copyranoside (XXIII). The treatment of Troc deriva- (10% isopropanol in chloroform).
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
135
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di- 5.477 (1 H, dd ≈ d, J3,4 3.4, J4,5 < 1, H4b), 5.499 (1 H,
é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-6-é-benzyl- dd ≈ d, J3,4 3.2, J4,5 < 1, H4d).
3-é-[2-acetamido-3,6-di-é-acetyl-2-deoxy-4-é-
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
(2,3,4,6-tetra-é-acetyl-b-D-galactopyranosyl)-b-
é-acetyl-2-deoxy-4-é-{2,4,6-tri-é-acetyl-3-é-[2-ace-
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu-
tamido-3,6-di-é-acetyl-2-deoxy-4-é-(2,3,4,6-tetra-é-
copyranoside (XXV). The treatment of Troc derivative
acetyl-b-D-galactopyranosyl)-b-D-glucopyranosyl]-b-
(XXIV) (601 mg, 0.39 mmol) with zinc in acetic acid
D-galactopyranosyl}-b-D-glucopyranoside (XXVII).
and three subsequent chromatographic purifications on
Acetylation of tetrasaccharide (XXVI) (42 mg,
silica gel (elution with 9% isopropanol in chloroform,
0.032 mmol) and the subsequent chromatography on
and then twice with 10 : 1 ethyl acetate–isopropanol)
silica gel (7 : 1 chloroform–isopropanol) yielded 39 mg
yielded 401.6 mg (73%) of tetrasaccharide (XXV);
(90%) of peracetate (XXVII); Rf 0.35 (7 : 1 chloro-
Rf 0.45 (9 : 1 ethyl acetate–isopropanol); MS, m/z: 1420
form–isopropanol); [α]D +4.73 (Ò 1, chloroform); MS,
(1397 + 23) M+ + Na+; 1H NMR (CDCl3): 1.907, 1.955,
1.977, 2.004, 2.055, 2.066, 2.073, 2.080, 2.083, 2.092,
2.095, and 2.154 (12 × 3 H, 12 s, 12 Ac), 3.43–3.51 (3
H, m, CHN, H6'b, H6"b), 3.537 (1 H, m, H5c), 3.618 (3
H, m, CHN, H2c, H5a), 3.67–3.83 (5 H, m, OCH, H3b,
H5b, H4a, H4c), 3.880 (2 H, m, OCH, H5d), 4.019 (1
H, dd, J6',6'' 11.9, J5,6'' 3.8, H6"c), 4.043 (1 H, m, H2a),
4.103 (2 H, m, H6'd, H6"d), 4.131 (1 H, dd, J6',6'' 11.5,
1
m/z: 1372 (1349 + 23) M+ + Na+; H NMR (CDCl3):
1.903, 1.960, 1.978, 2.052, 2.072 (×3), 2.087, 2.114,
2.122, 2.124, 2.154, and 2.157 (13 × 3 H, 13 s, 13 Ac),
3.44–3.66 (5 H, m, CH2N, H5a, H5c, H2c), 3.738 (1 H,
dd ≈ t, J3,4 8.4, J4,5 8.8, H4a), 3.745 (1 H, m, OCH),
3.759 (1 H, dd, J2,3 10.1, J3,4 3.5, H3b), 3.802 (1 H, ddd
≈ t, H5b), 3.807 (1 H, dd ≈ t, J3,4 8.8, J4,5 9.4, H4c),
3.883 (2 H, m, H5d, OCH), 3.985 (1 H, dd, J6',6'' 11.9,
J
5,6'' 6.6, H6"a), 4.369 (1 H, d, J1,2 7.9, H1b), 4.392 (1
J5,6'' 3.3, H6"c), 4.035 (1 H, ddd, J1,2 7.9, J2,NH 8.8, J2,3
H, d, J1,2 7.6, H1a), 4.460 (1 H, dd, J6',6'' 11.5, J5,6' 2.3,
H6'a), 4.474 and 4.519 (2 × 1 H, 2 d, JAB 11.6, CH2Ph),
4.536 (1 H, d, J1,2 7.8, H1d), 4.677 (1 H, d, J1,2 7.5,
H1c), 4.687 (1 H, dd, J6',6'' 11.9, J5,6' 2.2, H6'c), 4.985 (1
H, dd, J2,3 10.5, J3,4 3.4, H3d), 5.004 (1 H, dd, J1,2 7.9,
9.7, H2a), 4.04–4.16 (5 H, m, H6"a, H6'b, H6"b, H6'd,
H6"d), 4.383 (1 H, d, J1,2 7.9, H1b), 4.439 (1 H, d, J1,2
7.9, H1a), 4.474 (1 H, dd, J6',6'' 11.9, J5,6' 2.3, H6'a),
4.555 (1 H, d, J1,2 7.9, H1d), 4.693 (1 H, d, J1,2 7.7,
H1c), 4.781 (1 H, dd, J6',6'' 11.9, J5,6' 2.3, H6'c), 4.992 (1
H, dd, J1,2 7.9, J2,3 10.1, H2b), 4.993 (1 H, dd, J2,3 10.5,
J3,4 3.5, H3d), 5.024 (1 H, dd, J2,3 9.7, J3,4 8.4, H3a),
5.118 (1 H, dd, J1,2 7.9, J2,3 10.5, H2d), 5.192 (1 H, dd,
J
2,3 10.1, H2b), 5.010 (1 H, dd, J2,3 9.6, J3,4 8.2, H3a),
5.118 (1 H, dd, J1,2 7.8, J2,3 10.5, H2d), 5.165 (1 H, dd,
2,3 9.8, J3,4 8.8, H3c), 5.361 (1 H, dd, J3,4 3.4, J4,5 < 1,
J
H4b), 5.394 (1 H, dd, H4d), 5.400 (1 H, d, J2,NH 8.7,
NHAc-c), 5.777 (1 H, d, J2,NH 8.8, NHAc-a), 7.230 (1
H, m, NHCOCF3), 7.331 (5 H, m, Ph).
J
2,3 10.1, J3,4 8.8, H3c), 5.335 (1 H, dd ≈ d, J3,4 3.5, J4,5
< 1, H4b), 5.361 (1 H, dd ≈ d, J3,4 3.5, J4,5 < 1, H4d),
5.417 (1 H, d, J2,NH 8.4, NHAc-c), 5.831 (1 H, d, J2,NH
2-Trifluoroacetamidoethyl) 2-acetamido-3,6-di- 8.8, NHAc-a), 7.204 (1 H, m, NHCOCF3).
é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-3-é-[2-ace-
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{3-é-[2-
tamido-3,6-di-é-acetyl-2-deoxy-4-é-(2,3,4,6-tetra-
acetamido-2-deoxy-4-é-(b-D-galactopyranosyl)-b-
é-acetyl-b-D-galactopyranosyl)-b-D-glucopyrano-
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu-
syl]-b-D-galactopyranosyl}-b-D-glucopyranoside
copyranoside (VI). De-é-acetylation and removal of
(XXVI). Hydrogenolysis of (XXV) (372 mg,
the N-trifluoroacetamide protective group from
0.266 mmol) and the subsequent chromatographic
(XXVII) (98 mg, 0.073 mmol) resulted in 55 mg (96%)
purification of product on silica gel (elution with 9%
of tetrasaccharide (VI), Rf 0.46 (4 : 1 : 1 : 1 ethanol–
isopropanol in chloroform containing 0.5% pyridine
water–pyridine–acetic acid), [α]D –8.67 (Ò 0.5, water).
and then with 9 : 1 ethyl acetate–isopropanol contain-
MS, m/z: 1948 (1925 + 23) M+ + Na+; 1H NMR (D2O):
ing 0.5% pyridine) yielded 255 mg (73%) of the corre-
2.051 and 2.064 (2 × 3 H, 2 s, 2 Ac), 3.240 (2 H, m,
sponding 6b-OH derivative (XXVI); Rf 0.36 (5 : 1 chlo-
CH2N), 3.559 (1 H, dd, J1,2 7.9, J2,3 9.8, H2d), 3.607 (1
roform–isopropanol); 1H NMR (3 : 1 CDCl3–CD3OD):
H, dd, J1,2 7.9, J2,3 9.8, H2b), 3.58–3.66 (2 H, m), 3.687
2.023, 2.066, 2.118, 2.185, 2.197, 2.205, 2.213, 2.229,
(1 H, dd, J2,3 9.8, J3,4 3.4, H3d), 3.70–3.83 (13 H, m),
2.259, 2.265, 2.294, and 2.296 (112 × 3 H, 12 s, 12 Ac),
3.83–3.89 (2 H, m), 3.914 (1 H, m, OCH), 3.946 (1 H,
3.55–3.77 (8 H, m, CH2N, H6'b, H6"b, H5b, H5a, H5c,
dd, J3,4 3.4, J4,5 < 1, H4d), 3.972 (1 H, dd, J6',6'' 12.2, J5,6'
H2c), 3.803 (1 H, m, OCH), 3.918 (3 H, m, H3b, H4a,
2.0, H6'c), 4.012 (1 H, dd, J6',6'' 12.1, J5,6' 1.7, H6'a),
H4c), 4.009 (1 H, m, OCH), 4.088 (2 H, m, J1,2 8.3, J2,3
4.076 (1 H, m, OCH), 4.173 (1 H, dd, J3,4 3.4, J4,5 < 1,
10.3, H2a, H5d), 4.149 (1 H, dd, J6',6'' 11.9, J5,6'' 3.9,
H4b), 4.480 and 4.500 (2 H, 2 d, J1,2 7.8, H1b and H1d),
H6"c), 4.256 (3 H, m, H6'a, H6'd, H6"d), 4.549 (1 H, d,
4.596 (1 H, d, J1,2 8.2, H1a), 4.724 (1 H, d, J1,2 8.4,
J
1,2 7.9, H1b), 4.571 (1 H, d, J1,2 8.3, H1a), 4.593 (1 H,
H1c).
dd, J6',6'' 11.9, J5,6'' 2.0, H6"a), 4.721 (1 H, d, J1,2 7.8,
H1d), 4.778 (1 H, dd, J6',6'' 11.9, J5,6' 2.0, H6'c), 4.854 (1
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
H, d, J1,2 8.1, H1c), 5.101 (1 H, dd, J1,2 7.9, J2,3 9.9, é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-3-é-[2-ace-
H2b), 5.140 (1 H, dd, J3,4 3.4, J2,3 10.5, H3d), 5.152 (1 tamido-3,6-di-é-acetyl-2-deoxy-4-é-(2,3,4,6-tetra-
H, dd, J2,3 10.3, J3,4 8.8, H3a), 5.219 (1 H, dd, J1,2 7.8, é-acetyl-b-D-galactopyranosyl)-b-D-glucopyrano-
J2,3 10.5, H2d), 5.307 (1 H, dd, J2,3 10.3, J3,4 8.9, H3c), syl]-6-é-[2-acetamido-3,4,6-tri-é-acetyl-2-deoxy-b-
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

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SEVEROV et al.
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu- syl]-6-é-[2-acetamido-3,6-di-é-acetyl-2-deoxy-4-
copyranoside (XXIX). Glycosylation of tetrasaccha- é-(2,3,4,6-tetra-é-acetyl-b-D-galactopyranosyl)-b-
ride (XXVI) (142 mg, 0.109 mmol) with glycosyl bro- D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu-
mide (XIV) prepared from the corresponding perace- copyranoside (XXXI). Glycosylation of tetrasaccha-
tate (114 mg, 0.217 mmol) and subsequent isolation by ride (XXVI) (148 mg, 0.113 mmol) with glycosyl bro-
chromatography on silica gel (10 : 1chloroform–iso- mide (XXI) prepared from the corresponding thiogly-
propanol) yielded 35 mg (24%) of the starting (XXVI) coside (139 mg, 0.171 mmol) and repeated
and 96 mg (50% or 65% taking into account the recov- chromatographic purification on silica gel (10 : 1 chlo-
ery of the starting compound) of a mixture of the prod- roform–isopropanol) yielded 39 mg (26%) of the start-
ucts with the same chromatographic mobility, Rf 0.45 ing (XXVI) and 122.6 mg (53%) of the target (XXX);
1
(10 : 1 ethyl acetate–isopropanol). According to H Rf 0.22 (10 : 1 chloroform–methanol). Troc derivative
NMR, pentasaccharide (XXVIII) was the main compo- (XXX) was treated with zinc in acetic acid and sub-
nent of the mixture. The resulting mixture was treated jected to five subsequent chromatographic purifications
with zinc in acetic acid and separated by repeated chro- on silica gel (elution with 9% isopropanol in ethyl ace-
matography on silica gel (elution with 17% isopropanol tate, twice with 10% isopropanol in ethyl acetate, and
in chloroform and with 12% isopropanol in chloro- twice with 17% isopropanol in chloroform) to give
form) to give 51.4 mg (58%) of peracetate (XXIX); Rf 53 mg (46%) of peracetate (XXXI); Rf 0.30 (7 : 1 ethyl
0.27 (7 : 1 ethyl acetate–isopropanol); [α]D +9.8 (Ò 1, acetate–isopropanol); [α]D –6.4 (Ò 1, chloroform); MS,
1
chloroform); MS, m/z: 1660 (1637 + 23) M+ + Na+; 1H m/z: 1947 (1924 + 23) M+ + Na+; H NMR (CDCl3):
NMR (3 : 1 CDCl3–CD3OD): 2.020, 2.072, 2.081, 1.912, 1.963, 1.971, 1.977, 1.986, 2.052, 2.063 (×4),
2.121, 2.170, 2.185, 2.187, 2.199, 2.209, 2.216, 2.227, 2.082, 2.085, 2.089, 2.100, 2.108, 2.143, 2.153, 2.158,
2.230, 2.246, 2.269, 2.299, and 2.320 (16 × 3 H, 16 s, and 2.161 (19 × 3 H, 19 s, 19 Ac), 3.48–3.76 (10 H, m),
16 Ac), 3.56–3.66 (2 H, m, CH2N), 3.66–3.84 (7 H, m, 3.806 (4 H, m, H2e, H4e, H3b, H4c), 3.84–3.94 (4 H,
OCH, H2c, H2e, H5a, H5c, H6'b, H6"b), 3.85–3.97 (5 m, OCH, H4a, H5d, H5f), 4.003 (1 H, dd, J6',6'' 11.9, J5,6''
H, m, H3b, H4a, H4c, H5e, H5b), 4.020 (1 H, m, OCH), 3.1, H6"c), 4.009 (1 H, ddd, H2a), 4.05–4.23 (6 H, m,
4.077 (1 H, dd, J1,2 7.9, J2,3 9.8, H2a), 4.095 (1 H, ddd ≈ H6"a, H6"e, H6'd, H6"d, H6'f, H6"f), 4.378 (1 H, d, J1,2
t, J4,5 < 1, J5,6' ≈ J5,6'' ≈ 6.0, H5d), 4.154 (1 H, dd, J6',6'' 8.2, H1b), 4.449 (1 H, dd, J6',6'' 11.9, J5,6' 2.2, H6'a),
11.9, J5,6'' 3.8, H6"c), 4.260 (2 H, m, H6'd and H6"d), 4.530 (1 H, d, J1,2 7.9, H1a), 4.537 (2 H, 2 d, J1,2 7.9,
4.282 (1 H, dd, J6',6'' 12.1, J5,6'' 5.5, H6"a), 4.412 (1 H, H1d and H1f), 4.594 (1 H, dd, J6',6'' 12.0, J5,6' 2.6, H6'e),
dd, J6',6'' 12.3, J5,6' 4.7, H6'e), 4.528 (1 H, d, J1,2 8.1, 4.665 (1 H, d, J1,2 7.9, H1c), 4.700 (1 H, dd, J5,6' 1.8,
H1b), 4.574 (1 H, dd, J6',6'' 12.0, J5,6' 2.0, H6'a), 4.595 (1
J6',6'' 12.0, H6'c), 4.738 (1 H, d, J1,2 7.0, H1e), 4.958 (1
H, d, J1,2 7.9, H1a), 4.735 (1 H, d, J1,2 7.7, H1d), 4.825 H, dd, J1,2 7.9, J2,3 9.7, H2b), 4.983 and 5.005 (2 × 1 H,
(2 H, m, J1,2 8.1, H6'c, H1c), 4.981 (1 H, d, J1,2 8.3, 2 dd, J2,3 11.0, J3,4 3.4, H3d and H3f), 5.115 (2 H, 2 dd,
H1e), 5.072 (1 H, dd, J1,2 8.1, J2,3 10.0, H2b), 5.152 (1 H2d and H2f), 5.158 (2 H, 2dd ≈ t, H3a and H3c), 5.218
H, dd, J2,3 10.5, J3,4 3.5, H3d), 5.172 (2 H, dd ≈ t, J2,3
≈
(1 H, dd ≈ t, J3,4 8.3, J2,3 8.8, H3e), 5.299 (1 H, dd ≈ d,
J3,4 ≈ J4,5 ≈ 9.5, H3a, H4e), 5.223 (1 H, dd, J2,3 10.1, J3,4
J3,4 3.4, J4,5 < 1, H4b), 5.358 and 5.377 (2 × 1 H, 2 dd ≈
9.0, H3c), 5.465 (1 H, dd ≈ d, J3,4 3.4, J4,5 < 1, H4b), d, J3,4 3.4, J4,5 < 1, H4d and H4f), 5.407 (1 H, d, J2,NH
5.501 (1 H, dd ≈ d, J3,4 3.5, J4,5 < 1, H4d), 5.564 (1 H, 8.6, NHAc-c), 6.281 (1 H, d, J2,NH 8.8, NHAc-e), 6.324
dd, J2,3 10.1, J3,4 9.5, H3e).
(1 H, d, J2,NH 8.6, NHAc-a), 7.335 (1 H, m, NHCOCF3).
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{3-é-[2-
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{3-é-
[2-acetamido-2-deoxy-4-é-(b-D-galactopyranosyl)- acetamido-2-deoxy-4-é-(b-D-galactopyranosyl)-b-
b-D-glucopyranosyl]-6-é-[2-acetamido-2-deoxy-b- D-glucopyranosyl]-6-é-[2-acetamido-2-deoxy-4-
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu- é-(b-D-galactopyranosyl)-b-D-glucopyranosyl]-b-
copyranoside (V). De-é-acetylation and removal of D-galactopyranosyl}-b-D-glucopyranoside (VIII).
the N-trifluoroacetamide protective group from De-é-acetylation and the removal of N-trifluoroaceta-
(XXIX) (50.4 mg, 0.031 mmol) yielded 28.1 mg (92%) mide protective group from (XXXI) (53 mg, 0.028
of pentasaccaride (V); Rf 0.44 (4 : 1 : 1 : 1 ethanol– mmol) yielded 29.4 mg (93%) of hexasaccharide
water–pyridine–acetic acid); [α]D –12.5 (Ò 0.5, water); (VIII); Rf 0.39 (4 : 1 : 1 : 1 ethanol–water–pyridine–ace-
MS, m/z: 1017 (994 + 23) M+ + Na+; 1H NMR (D2O): tic acid); [α]D +7.15 (Ò 0.5, water); MS, m/z: 1179
2.047 and 2.073 (×2) (3 × 3 H, 3c, 3Ac), 3.242 (2 H, m, (1156 + 23) M+ + Na+; 1H NMR (D2O): 2.046 and 2.069
CH2N), 3.461 (2 H, m), 3.53–3.65 (5 H, m), 3.66–3.89 (×2) (3 × 3 H, 3c, 3Ac), 3.241 (2 H, m, CH2N), 3.52–3.69
(18 H, m), 3.89–4.03 (6 H, m), 4.078 (1 H, m, OCH), (6 H, m), 3.66–3.89 (24 H, m), 3.90–4.03 (7 H, m),
4.162 (1 H, dd, J3,4 3.1, J4,5 < 1, H4b), 4.470 (1 H, d, J1,2 4.079 (1 H, m, OCH), 4.165 (1 H, dd, J3,4 2.4, J4,5 < 1,
7.9, H1b), 4.495 (1 H, d, J1,2 7.7, H1d), 4.596 (2 H, 2 d, H4b), 4.484 (3 H, m, H1b, H1d, and H1f), 4.594 (1 H,
J
1,2 8.4, H1a and H1e), 4.716 (1 H, d, J1,2 8.3, H1c).
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
d, H1a, J1,2 8.3), 4.641 (1 H, d, J1,2 7.2, H1e), 4.716 (1 H,
d, J1,2 8.3, H1c).
é-acetyl-2-deoxy-4-é-{2,4-di-é-acetyl-3-é-[2-ace-
tamido-3,6-di-é-acetyl-2-deoxy-4-é-(2,3,4,6-tetra- acetyl-2-deoxy-4-é-(2,3,4-tri-é-acetyl-b-D-galactopy-
é-acetyl-b-D-galactopyranosyl)-b-D-glucopyrano- ranosyl)-b-D-glucopyranoside (XII). Hydrogenolysis
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-é-
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

SYNTHESIS OF N-ACETYLLACTOSAMINE-CONTAINING OLIGOSACCHARIDES
137
of lactosamine derivative (XI) (975 mg, 1.13 mmol) galactopyranosyl]-b-D-glucopyranoside (II). De-é-
and the subsequent chromatography on silica gel (elu- acetylation and removal of the N-trifluoroacetamide
tion with 7–15% isopropanol in chloroform containing protective group from (XXXIII) (153 mg, 0.144 mmol)
0.5% pyridine) yielded 572 mg (69%) of 6b-OH deriv- yielded 90 mg (99%) of trisaccharide (II); Rf 0.61 (3 :
ative (XII); Rf 0.45 (10% isopropanol in ethyl acetate); 1 : 1 : 1 ethanol–water–pyridine–acetic acid); [α]D
[α]D –17.4 (Ò 1, 3 : 1 CHCl3–CH3OH); MS, m/z: 754 −32.5 (Ò 1, water); MS, m/z: 652 (629 + 23) M+ + Na+;
(732 + 23) M+ + Na+; 1H NMR (3 : 1 CDCl3–CD3OD): 1H NMR (D2O): 2.069 (×2) (2 × 3 H, 2 s, 2 Ac), 3.238
2.074, 2.120, 2.211, 2.220, 2.269, and 2.292 (6 × 3 H, (2 H, m, CH2N), 3.42–3.49 (2 H, m, H4c), 3.544 (1 H,
6 s, 6 Ac), 3.55–3.69 (3 H, m, CH2N, 3.655, J6',6'' 11.1, dd, J1,2 7.8, J2,3 10.0, H2b), 3.560 (1 H, dd, J2,3 10.4, J3,4
J5,6' 6.4, H6'b), 3.74–3.84 (3 H, m, OCH, H5a, 3.799, 8.4, H3c), 3.629 (1 H, m, H5a), 3.666 (1 H, dd, J2,3 10.0,
dd, J6',6'' 11.1, J5,6'' 6.6, H6"b), 3.871 (1 H, m, H5b), J3,4 3.3, H3b), 3.689 (1 H, dd, J3,4 8.2, J4,5 9.7, H4a),
3.970 (1 H, dd, J3,4 8.8, J4,5 9.5, H4a), 4.014 (1 H, m, 3.699 (1 H, dd, J1,2 8.4, J2,3 10.4, H2c), 3.738 (1 H, dd,
OCH), 4.095 (1 H, dd, J1,2 8.3, J2,3 10.3, H2a), 4.248 (1
J2,3 10.2, J3,4 8.2, H3a), 3.74–3.78 (1 H, m, H5c), 3.817
H, dd, J6',6'' 12.0, J5,6' 5.4, H6'a), 4.589 (1 H, d, J1,2 8.3, (1 H, dd, J1,2 8.4, J2,3 10.2, H2a), 3.82–4.02 (9 H, m),
H1a), 4.639 (1 H, dd, J6',6'' 12.0, J5,6'' 2.1, H6"a), 4.689 4.074 (1 H, m, OCH), 4.477 (1 H, d, J1,2 7.8, H1b),
(1 H, d, J1,2 7.8, H1b), 5.144 (1 H, dd, J3,4 3.2, J2,3 10.3, 4.595 (2 H, d, J1,2 8.4, H1a, H1c).
H3b), 5.173 (1 H, dd, J2,3 10.3, J3,4 8.8, H3a), 5.211 (1
H, dd, J1,2 7.8, J2,3 10.3, H2b), 5.557 (1 H, dd, J3,4 3.2,
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di-
é-acetyl-2-deoxy-4-é-{2,3,4-tri-é-acetyl-6-é-[2-
acetamido-3,6-di-O-acetyl-2-deoxy-4-é-(2,3,4,6-
J4,5 < 1, H4b).
2-Trifluoroacetamidoethyl 2-acetamido-3,6-di- tetra-é-acetyl-b-D-galactopyranosyl)-b-D-glucopy-
é-acetyl-2-deoxy-4-é-[2,3,4-tri-é-acetyl-6-é-(2-ace- ranosyl]-b-D-galactopyranosyl}-b-D-glucopyrano-
tamido-3,4,6-tri-é-acetyl-2-deoxy-b-D-glucopyrano-
syl)-b-D-galactopyranosyl]-b-D-glucopyranoside
side (XXXV). Glycosylation of disaccharide (XII)
(210 mg, 0.287 mmol) with glycosyl bromide (XXI)
(XXXIII). Glycosylation of disaccharide (XII) obtained from the corresponding thioglycoside
(250 mg, 0.342 mmol) with glycosyl bromide (XIV) (301 ;mg, 0.370 mmol) and the subsequent chromatog-
prepared from the corresponding peracetate (267 mg, raphy on silica gel (elution with 5–10 % isopropanol in
0.512 mmol) and subsequent chromatography on silica chloroform) yielded 63 mg (30%) of the starting (XII)
gel (elution with 5–10% isopropanol in chloroform) and 258 mg (61%) of tetrasaccharide (XXXIV), Rf 0.27
yielded 37 mg (15%) of the mixture of the starting (10% isopropanol in chloroform). The treatment of
(XII), its isomer (the product of migration of the OAc Troc derivative (XXXIV) with zinc in acetic acid and
group from C6 to C4 of the galactose unit), and 323 mg four subsequent chromatographic purifications on sil-
(79%) of the mixture of the products with the same ica gel (elution with 5–7% isopropanol in ethyl acetate)
chromatographic mobility, Rf 0.30 (10% isopropanol in yielded 159 mg (68%) of N-acetyl derivative (XXXV)
chloroform). According to 1H NMR data, trisaccharide (68%), Rf 0.48 (10% isopropanol in ethyl acetate); [α]D
(XXXII) was the main component. The resulting mix- –7.2 (Ò 1, chloroform); MS, m/z: 1372 (1349 + 23) M+ +
1
ture was treated with zinc in acetic acid. Four chro- Na+; H NMR (CDCl3): 1.961, 1.964, 1.975, 1.981,
matographic purifications on silica gel (elution with 8– 2.053, 2.064, 2.065, 2.089, 2.095, 2.114, 2.135, 2.143,
15% isopropanol in chloroform and with 8% isopro- and 2.157 (13 × 3 H, 13 s, 13 Ac), 3.47–3.64 (2 H, m,
panol in ethyl acetate) yielded 160 mg (59%) of perac- CH2N), 3.64–3.77 (5 H, m, OCH, H6'b, H6"b, H5a,
etate (XXXIII); Rf 0.34 (10% isopropanol in ethyl ace- H5c), 3.794 (1 H, dd ≈ t, J3,4 ≈ J4,5 ≈ 8.1, H4c), 3.843 (1
1
tate); H NMR (3 : 1 CDCl3–CD3OD): 2.073, 2.077, H, ddd ≈ t, J4,5 < 1, J5,6' ≈ J5,6'' ≈ 5.6, H5b), 3.865–3.946
2.104, 2.166, 2.180, 2.207, 2.211, 2.249, and 2.275 (4 H, m, éëç, H2c, H5d, H4a), 4.015 (1 H, ddd, J2,NH
(×2) (10 × 3 ç, 10Ò, 10ÄÒ), 3.56–3.69 (2 H, m, CH2N), 8.8, J1,2 8.1, J2,3 8.7, H2a), 4.111 (2 H, m, H6'd and
3.770 (1 H, dd, J1,2 8.3, J2,3 10.5, H2c), 3.75–3.79 (1 H, H6"d), 4.177 (2 H, m, H6"a and H6"c), 4.491 (5 H, m,
m, H5a), 3.814 (1 H, m, OCH), 3.85–3.91 (3 H, m, H5, H1a, H1b, H1d, H6'a, H6'c), 4.671 (1 H, d, J1,2 7.0,
H6'b and H6"b), 3.933 (1 H, dd, J3,4 8.8, J4,5 9.5, H4a), H1c), 4.985 (1 H, dd, J2,3 10.4, J3,4 3.4, H3b), 5.000 (1
3.99–4.05 (2 H, m, OCH, H5b), 4.081 (1 H, dd, J1,2 8.3, H, dd, J2,3 10.4, J3,4 3.4, H3d), 5.078 (1 H, dd, J1,2 7.8,
J2,3 10.3, H2a), 4.260 (1 H, dd, J6',6'' 11.7, J5,6' 5.6, H6'a),
J2,3 10.4, H2b), 5.112 (1 H, dd, J1,2 7.8, J2,3 10.4, H2d),
4.298 (1 H, dd, J6',6'' 12.5, J5,6'' 2.2, H6"c), 4.412 (1 H, 5.141 (1 H, dd ≈ t, J3,4 8.1, J2,3 8.8, H3c), 5.170 (1 H,
dd, J6',6'' 12.5, J5,6' 4.7, H6'c), 4.606 (1 H, d, J1,2 8.3, dd ≈ t, J2,3 ≈ J3,4 ≈ 9.0, H3a), 5.361 (1 H, dd ≈ d, J3,4 3.4,
H1a), 4.628 (1 H, dd, J6',6'' 11.7, J5,6'' 2.2, H6"a), 4.703 J4,5 < 1, H4b), 5.373 (1 H, dd ≈ d, J3,4 3.4, J4,5 < 1, H4d),
(1 H, d, J1,2 7.8, H1b), 4.980 (1 H, d, J1,2 8.3, H1C), 6.046 (1 H, d, J2,NH 9.0, NHAc-c), 6.236 (1 H, d, J2,NH
5.066 (1 H, dd, J2,3 10.3, J3,4 3.4, H3b), 5.155 (1 H, dd ≈ 8.8, NHAc-a), 7.371 (1 H, m, NHCOCF3).
t, J3,4 ≈ J4,5 ≈ 9.3, H4c), 5.191 (1 H, dd, J1,2 7.8, J2,3 10.3,
H2b), 5.213 (1 H, dd, J2,3 10.3, J3,4 8.8, H3a), 5.488
(1 H, dd, J2,3 10.5, J3,4 9.3, H3C), 5.511 (1 H, dd, J3,4
3.4, J4,5 < 1, H4b).
2-Aminoethyl 2-acetamido-2-deoxy-4-é-[6-é- N-trifluoroacetamide protective group from peracetate
(2-acetamido-2-deoxy-b-D-glucopyranosyl)-b-D- (XXXV) (262 mg, 0.194 mmol) yielded 146 mg (95%)
2-Aminoethyl 2-acetamido-2-deoxy-4-é-{6-é-[2-
acetamido-2-deoxy-4-é-(b-D-galactopyranosyl)-b-
D-glucopyranosyl]-b-D-galactopyranosyl}-b-D-glu-
copyranoside (VII). De-é-acetylation and removal of
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 33 No. 1 2007

138
SEVEROV et al.
Den Otter, W., Breast Cancer Research and Treatment,
of tetrasaccharide (VII); Rf 0.59 (3 : 1 : 1 : 1 ethanol–
water–pyridine–acetic acid); [α]D –23.9 (Ò 0.5, water);
2005, vol. 91, pp. 227–241.
1
MS, m/z: 814 (791 + 23) M+ + Na+; H NMR (D2O):
4. Windholz, T.M. and Jonston, B.R., Tetrahedron Lett.,
1967, p. 2555.
2.071 and 2.076 (2 × 3 H, 2 s, 2 Ac), 3.246 (2 H, m,
CH2N), 3.557 (2 H, m, H2b and H2d), 3.59–3.94 (18 H,
m), 3.934 and 3.946 (2 H, 2 dd, J3,4 3.4, J4,5 < 1, H4b
and H4d), 3.96–4.03 (3 H, m), 4.082 (1 H, m, OCH),
4.486 (2 H, 2 d, J1,2 7.9, H1b and H1d), 4.601 (1 H, d,
5. Pazynina, G.V. and Bovin, N.V., Mendeleev Commun.,
2000, pp. 132–134.
6. Pazynina, G.V., Severov, V.V., and Bovin, N.V., Men-
deleev Commun., 2002, pp. 183–184.
7. Blixt, O., Brown, J., Schur, M.J., Wakarchuk, W., and
Paulson, J.C., J. Org. Chem., 2001, vol. 66, pp. 2442–
2448.
J1,2 8.2, H1a), 4.645 (1 H, d, J1,2 7.2, H1c).
ACKNOWLEDGMENTS
8. Curphey, T.J., J. Org. Chem., 1979, vol. 44, pp. 2805–
2807.
We are grateful to O. Blixt (the Scripps Research
9. Garegg, P. and Hultberg, H., Carbohydr. Res., 1982, vol.
Institute, San Diego, United States) for N-acetyllac-
108, pp. 97–101.
tosamine azidoethyl glycoside granted.
10. Mukhopadhyay, B. and Robert, A., Carbohydr. Res.,
This work was supported by the National Institute of
Health (grant no. 5 U54 GM062116-05) and by the
Russian Foundation for Basic Research (project no. 03-
04-48926).
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