An easy access to a 3,6-branched mannopentaoside bearing one terminal [1-13C]-labeled D-mannopyranose residue1

Article abstract of DOI:10.1007/s11172-005-0396-z

Methyl 2,4-di-O-benzoyl-α-D-mannopyranoside was used as a key intermediate in the synthesis of 3,6-branched mannopentaoside bearing one terminal D-[1-¹³C]mannopyranose residue, viz., methyl 6-O-[3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranosyl)

Full text of DOI:10.1007/s11172-005-0396-z

Russian Chemical Bulletin, International Edition, Vol. 54, No. 5, pp. 1287—1293, May, 2005
1287
An easy access to a 3,6ꢀbranched mannopentaoside bearing
one terminal [1ꢀ13C]ꢀlabeled Dꢀmannopyranose residue*
ꢀ
P. I. Abronina, L. V. Backinowsky, A. A. Grachev, S. L. Sedinkin, and N. N. Malysheva
N. D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,
47 Leninsky prosp., 119991 Moscow, Russian Federation.
Fax: +7 (095) 135 5328. Eꢀmail: leon@ioc.ac.ru
Methyl 2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoside was used as a key intermediate in the
synthesis of 3,6ꢀbranched mannopentaoside bearing one terminal Dꢀ[1ꢀ13C]mannopyranose
residue, viz., methyl 6ꢀOꢀ[3,6ꢀdiꢀOꢀ(αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranosyl)ꢀ3ꢀOꢀ
{αꢀDꢀ[1ꢀ13C]mannopyranosyl}ꢀαꢀDꢀmannopyranoside.
Key words: methyl 2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoside, 3,6ꢀbranched mannooligoꢀ
saccharides, Dꢀ[1ꢀ13C]mannopyranose, [1ꢀ13C]ꢀlabeled mannopentaoside, selective glycoꢀ
sylation, selective deacylation.
It is well known that carbohydrates play an important
role in various recognition processes, for example, in inꢀ
tercellular interactions and immune reactions. Detailed
investigation of binding of carbohydrates to protein reꢀ
ceptors may be of considerable importance for the design
of potential modulators of such interactions. The strucꢀ
tures of specific carbohydrateꢀprotein complexes can be
studied by the siteꢀdirected replacement of amino acids of
the receptor or modifications of monosaccharide residues
of the ligand followed by calorimetry1 or by Xꢀray diffracꢀ
tion.2 NMR spectroscopy is a much more popular and
accessible method, which is widely used for the determiꢀ
nation of the conformations of carbohydrates in free state
and after their binding to proteins.3—5 This method is
particularly useful in combination with enrichment of oliꢀ
gosaccharides with stable isotopes, for example, with the
13C isotope.6,7
Results and Discussion
Mannoseꢀbinding proteins (MBP) are widespread in
plants and animals. Mannoseꢀspecific mammalian lectins
Stepwise and block syntheses of the aboveꢀmenꢀ
tioned mannopentaoside with natural 13C isotope abunꢀ
dance have been described repeatedly (see, for example,
Refs 12—15). Our approach to the synthesis of selectively
13Cꢀlabeled mannopentaoside is based on the use of readily
accessible methyl 2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoꢀ
side (2) as a key intermediate and its successive transforꢀ
mation into a (1—6)ꢀlinked disaccharide by selective
mannosylation, then into a 3,6ꢀbranched trisaccharide
containing the Dꢀ[1ꢀ13C]mannopyranose residue, and, fiꢀ
nally, into a pentasaccharide through bisꢀmannosylation.
Compound 2 was prepared by a oneꢀpot method
(Scheme 1) analogous to that described earlier for the
synthesis of octyl and tetradecyl αꢀDꢀmannopyranoside
derivatives.16 Treatment of methyl αꢀDꢀmannopyranoside
are involved in various immune processes, including inꢀ
nate immunity.8—10 Recently,11 antitumor activity of
MBP has been revealed. Synthetic analogs of natural carꢀ
bohydrate structures can be useful as ligands in elucidatꢀ
ing the character of binding of the latter to receptors. In
the present study, we synthesized one of such comꢀ
pounds, 3,6ꢀbranched mannopentaoside containing one
terminal Dꢀ[1ꢀ13C]mannopyranose residue, viz., methyl
6ꢀOꢀ[3,6ꢀdiꢀOꢀ(αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyraꢀ
nosyl]ꢀ3ꢀOꢀ{αꢀDꢀ[1ꢀ13C]mannopyranosyl}ꢀαꢀDꢀmannoꢀ
pyranoside (1ꢀ13C).
* Dedicated to Academician N. K. Kochetkov on the occasion
of his 90th birthday.
Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 5, pp. 1250—1255, May, 2005.
1066ꢀ5285/05/5405ꢀ1287 © 2005 Springer Science+Business Media, Inc.

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Russ.Chem.Bull., Int.Ed., Vol. 54, No. 5, May, 2005
Abronina et al.
Scheme 2
with triethyl orthobenzoate in the presence of TsOH and
CF3COOH in MeCN afforded 2,3:4,6ꢀbis(ethyl orthoꢀ
benzoate), which was subjected to acid hydrolysis without
isolation. The target compound 2 and its isomer, viz.,
methyl 2,6ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoside, which
differ substantially in chromatographic mobilities, were
isolated in 30 and 45% yields, respectively. Their 1H NMR
spectroscopic data are similar to those of analogous octyl
and tetradecyl αꢀDꢀmannopyranoside derivatives.16
Scheme 1
the synthesis of an analog of mannopentaoside 1ꢀ13C conꢀ
taining three terminal nonreducing Dꢀ[1ꢀ13C]mannoꢀ
pyranose residues.20
Reagents and conditions: i. PhC(OEt)3, H+; ii. H3O+.
Before the synthesis of mannopentaoside 1ꢀ13C, we
prepared its unlabeled analog. For this purpose, disacchaꢀ
ride 4 was initially transformed into trisaccharide 5 in
78% yield by the reaction with 2,3,4,6ꢀtetraꢀOꢀbenzoylꢀ
αꢀDꢀmannopyranosyl bromide in the presence of silver
trifluoromethanesulfonate (AgOTf) (Scheme 3). The seꢀ
lective removal of the Oꢀacetyl protective groups under
the conditions of mild acidic methanolysis where the
Oꢀbenzoyl groups remained intact19 afforded diol 6 in a
nearly quantitative yield.
In the synthesis of disaccharide, methyl 2,4ꢀdiꢀOꢀbenꢀ
zoylꢀαꢀDꢀmannopyranoside (2) served as both the glycoꢀ
syl acceptor and a precursor of a glycosyl donor. Acetolyꢀ
sis of glycoside 2 (Ac2O—AcOH—H2SO4, 40 °C) afforded
triacetate 3, which was then transformed into the corꢀ
responding glycosyl bromide according to a standard
procedure.17 The Helferich condensation (MeCN,
Hg(CN)2 + HgBr2) of this glycosyl donor with diol 2
occurs regioselectively to give (1—6)ꢀlinked disaccharide 4
in 57% yield (Scheme 2).
The presence of the 1—6 linkage in disaccharide 4 was
established by 13C NMR spectroscopy. The C(6a) atom
resonates at substantially lower field (δ 66.49) than the
C(6c) atom (δ 62.56) due to the α effect of glycosylation
(see Ref. 18).* The presence of a free hydroxy group at the
C(3a) atom in the derivative of disaccharide 4 provides
the possibility to perform the siteꢀdirected introduction of
the 13Cꢀlabeled mannopyranose residue, while selective
Oꢀdeacetylation19 of the resulting 3,6ꢀbranched trisacꢀ
charide allows one to carry out bisꢀmannosylation at the
O(3c) and O(6c) positions. It should be noted that we
have used selective Oꢀdeacetylation of disaccharide 4 in
Diol 6 was used as the glycosyl acceptor in condensaꢀ
tion with 1.8 equiv. of 2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀ
mannopyranosyl bromide in a CH2Cl2—toluene mixture
in the presence of AgOTf (Scheme 4). The yield of the
bisꢀmannosylation product, viz., fully protected mannoꢀ
pentaoside 7, was ~70%.
The structures of compounds 5—7 were confirmed by
NMR spectroscopy. In particular, the NMR spectrum of
diacetate 5 shows signals of acetyl groups (δH 1.90
and 1.80, δC 20.59 and 20.48 (Me); 169.74 and 170.40
(CO)), which disappear in the spectrum of diol 6 In addiꢀ
tion, the signals for H(6c) and H(6´c) in the spectrum of
diol 6 are shifted upfield (δH 4.22 → 3.67 and 3.60),
whereas the signals for C(2c), C(4c), and C(5c) are
shifted downfield (δC 70.20 → 72.81, 66.99 → 70.27, and
68.78 → 70.72, respectively) (Tables 1 and 2).
* The αꢀeffects of glycosylation revealed in the cited study18
refer to unprotected monosaccharide units. The signs of these
effects remain unchanged for the Oꢀacylated sugars, although
they can differ in magnitude.
The assignment of the signals for the protons and carꢀ
bon atoms in the NMR spectra of perbenzoate 7 was

3,6ꢀBranched 13Cꢀlabeled mannopentaoside
Russ.Chem.Bull., Int.Ed., Vol. 54, No. 5, May, 2005
1289
Scheme 3
Table 1. 1H NMR spectroscopic data
Scheme 4
Comꢀ Reꢀ
δ
poꢀ
und
siꢀ
due
H(1)
H(2)
H(3)
H(4) H(5) H(6)H(6´)
5
6
7
а
b
c
a
b
c
a
b
c
d
e
5.03 5.70—5.80 4.66
5.84 4.24 4.09 3.77
5.36
5.08
5.03
5.33
5.14
5.02
5.32
5.23
5.37
4.83
5.37 5.70—5.80 6.00 4.55 4.63 4.41
5.60 5.70—5.80 4.38 4.22
5.71 4.61—4.64 5.96 4.18 4.04 3.77
5.39 5.76 6.01 4.57 4.60 4.40
5.52 4.61—4.64 5.54 4.00 3.67 3.60
5.71
5.34
5.82
5.45
5.63
4.66
5.69
4.73
5.74
5.94
6.04 4.34 4.24 3.84
5.94 4.55 4.60 4.38
6.00 4.33 4.05 3.50
6.06 4.45 4.56 4.32
6.09 4.43 4.50 4.30
Table 2. 13C NMR spectroscopic data
Comꢀ Resiꢀ
pound due
δ
C(1) C(2) C(3) C(4) C(5) C(6)
made using 2D NMR techniques (COSY, ROESY, and
HSQC) with account of the spectroscopic data for fully
protected 3,6ꢀbranched mannooligosaccharides.14 Of sigꢀ
nals for the anomeric H and C atoms, those of the units
involved in 1—3 linkages (b and d) are observed at the
lowest field, whereas the signal of the terminal unit with
the 1—6 linkage (e) appears at the highest field. The
chemical shift of the anomeric proton of the latter unit
(δH 4.83) is close to that observed earlier20 (δH 4.87) for
an analogous unit in the spectrum of a mannopentaoside
containing three terminal nonreducing Dꢀ[1ꢀ13C]mannoꢀ
pyranose residues.
5
6
7
а
b
c
a
b
c
a
b
c
d
e
98.61 71.89 76.06 68.78 69.51 66.80
99.55 70.13 69.34 66.65 69.64 62.74
97.23 70.20 69.13 66.99 68.78 62.74
98.61 71.75 76.02 68.47 69.38 66.64
99.40 70.27 69.38 66.64 69.63 62.70
97.76 72.81 68.62 70.27 70.72 61.22
98.62 71.95 76.14 68.38 69.37 66.53
99.50 70.13 69.57 66.61 69.49 62.72
97.54 71.95 77.10 68.10 69.37 66.20
99.94 70.13 69.65 66.29 69.49 62.53
97.36 70.13 70.20 66.46 69.32 62.42

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Abronina et al.
The Zemplén debenzoylation (MeONa in MeOH) of
perbenzoate 7 afforded a free mannopentaoside having
structure 1 with natural 13C isotope abundance. The
13C NMR spectrum of the latter was identical to the specꢀ
trum of the pentasaccharide synthesized earlier.14
The same sequence of reactions, including mannoꢀ
sylation of glycosyl acceptor 4, Oꢀdeacetylation, and bisꢀ
mannosylation of the trisaccharide glycosyl acceptor,
was used for the synthesis of the target mannopentaꢀ
oside 1ꢀ13C.
Naturally, the first glycosylation was carried out with
2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀ[1ꢀ13C]mannopyranosyl
bromide prepared from the corresponding pentabenzoate
by treatment with hydrogen bromide in AcOH. Condenꢀ
sation of this glycosyl donor with disaccharide derivaꢀ
tive 4 in the presence of AgOTf under the conditions of
the synthesis of unlabeled trisaccharide 5 afforded trisacꢀ
charide 5ꢀ13C in 69% yield.
The chemical shifts of the anomeric H atoms in the
1H NMR spectrum of the latter compound (δ 5.04, 5.37,
and 5.09 for H(1a), H(1b), and H(1c), respectively) are
virtually identical to those observed in the spectrum of the
unlabeled analog (cf. Table 1). The signal for the anomeric
H atom of the [1ꢀ13C]ꢀlabeled mannopyranose residue,
1
H(1b), appears as a doublet with JH,C = 172 Hz, which
proves the α configuration of the resulting mannosidic
bond. In other respects, the 1H NMR spectrum of trisacꢀ
charide 5ꢀ13C is identical to that of 5.
The 13C NMR spectra of these trisaccharides are also
similar to each other. The spectrum of compound 5ꢀ13C
is characterized by high intensity of the signal for the
anomeric C atom of the residue b (δ 99.61).
a broadened singlet in the spectrum measured with the
use of broadꢀband {13C}ꢀ1H heteronuclear decoupling (b).
The latter spectrum is virtually identical to the specꢀ
trum (c) of the authentic sample of mannopentaoside 1
with natural 13C isotope abundance.
To summarize, we synthesized the branched 13Cꢀlaꢀ
beled mannopentaoside starting from methyl 2,4ꢀdiꢀOꢀ
benzoylꢀαꢀDꢀmannopyranoside as a key intermediate. This
mannopentaoside can be used as a probe for NMR studies
of interactions of mannoseꢀbinding proteins with ligands.
Deacetylation of diacetyl derivative 5ꢀ13C with HCl in
MeOH gave rise to labeled trisaccharide diol 6ꢀ13C in
96% yield.
The latter compound was bisꢀmannosylated under the
conditions described above for the synthesis of mannoꢀ
pentaoside 7. The glycosylation product 7ꢀ13C was isoꢀ
1
lated in 68% yield. The H NMR spectrum of this comꢀ
pound differed from the spectrum of the unlabeled
perbenzoylated mannopentaoside only in that it had a
Experimental
1
doublet for H(1b) (δ 5.34, JH,C = 173 Hz). The correꢀ
sponding chemical shifts of all other H atoms of both
compounds are the same within 0.02 ppm. The 13C NMR
spectra of these compounds are also virtually identical
(the differences in the chemical shifts are at most 0.1 ppm).
The target mannopentaoside 1ꢀ13C was prepared by
debenzoylation of the corresponding perbenzoate, viz.,
compound 7ꢀ13C. In our opinion, the structure of this
pentasaccharide is convincingly supported by a compariꢀ
son of the 1H NMR spectrum of labeled mannopentaoside
1ꢀ13C with the spectrum of authentic mannopentaoside 1
(see Fig. 1). As can be seen from Fig. 1, the doublet for
the anomeric proton H(1b) in the spectrum of mannoꢀ
pentaoside 1ꢀ13C (a) (1JH,C = 172 Hz) is transformed into
The optical rotation was measured on a PUꢀ07 digital polaꢀ
1
rimeter (Russia) in chloroform. The H and 13C NMR spectra
were recorded on ACꢀ200, WMꢀ250, AMꢀ300, and DRXꢀ500
spectrometers (Bruker) in CDCl3 with Me4Si as the internal
standard (the spectra of compound 1ꢀ13C were measured in D2O
with (CH3)2CO as the internal standard). The assignment of the
signals in the spectra was made using the COSY, ROESY, and
HSQC techniques. The NMR spectroscopic data are given in
Tables 1 and 2; the chemical shifts of the protons of the benzoyl
residues and MeO groups, the multiplicities of the signals (in the
tables), and the coupling constants characteristic of mannoꢀ
pyranosides are omitted.
Column chromatography was carried out on Silpearl silica
gel (Czech Republic), and TLC analysis was performed on Merck

3,6ꢀBranched 13Cꢀlabeled mannopentaoside
Russ.Chem.Bull., Int.Ed., Vol. 54, No. 5, May, 2005
1291
c
b
1a
1e
1d
1c
1b
a
1b
172 Hz
1d
1e
1a
1c
5.2
5.0
4.8
4.6
4.4
4.2
4.0
3.8
3.6
δ
Fig. 1. 1H NMR spectra (D2O) of (a) mannopentaoside 1ꢀ13C, (b) of the same pentasaccharide after broadꢀband {13C}ꢀ1H heteronuclear
decoupling, and (c) of a sample of mannopentaoside 1 with natural 13C isotope abundance.14 The notations 1a—1e refer to the signals
for the anomeric H atoms of the residues a—e, respectively.
DCꢀAlufolien Kieselgel 60254 plates; toluene—ethyl acetate mixꢀ
tures were used as eluents for column chromatography and solꢀ
vents for TLC; visualization was performed with UV light and by
spraying plates with dilute H2SO4 followed by heating at ∼150 °C.
Toluene was distilled over Na. Dichloromethane and acetoꢀ
nitrile were distilled successively over P2O5 and CaH2. Comꢀ
mercial methyl αꢀDꢀmannopyranoside (Sigma), triethyl
orthobenzoate (Aldrich), and Dꢀ[1ꢀ13C]mannose (Omicron
Biochemicals, Inc.) were used.
2,3,4,6ꢀTetraꢀOꢀbenzoylꢀαꢀDꢀmannopyranosyl bromide and
its [1ꢀ13C]ꢀlabeled analog were synthesized in virtually quantiꢀ
tative yields by the reaction of hydrogen bromide (20 equiv.,
a 40% solution in AcOH, which was prepared from AcBr and
the calculated amount of H2O in AcOH) with solutions of the
corresponding pentabenzoates in chloroform or dichloromethane
for 2 days followed by standard work up.17
Methyl 2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoside (2) (cf. lit.
data16). Triethyl orthobenzoate (2.9 g, 2.9 mL, 13 mmol) and a
solution of TsOH (60 mg) in MeCN (6.5 mL) and CF3COOH
(0.03 mL) were added with stirring to a suspension of methyl
αꢀDꢀmannopyranoside (0.97 g, 5 mmol) in MeCN (80 mL). The
reaction mixture was stirred at ∼40 °C until complete dissolution
was achieved (∼1 h), and then the solvent was evaporated. The
residue was dissolved in MeCN (45 mL), after which 90%
CF3COOH (4 mL) was added. The solution was kept at ∼20 °C
for 20 min and concentrated. Column chromatography of the
residue afforded isomers, viz., compound 2 and methyl 2,6ꢀdiꢀ
OꢀbenzoylꢀαꢀDꢀmannopyranoside.
Compound 2, the yield was 0.6 g (30%), [α]D –35.4 (c 1.3).
Found (%): C, 63.01; H, 5.87. C21H22O8. Calculated (%):
1
C, 62.68; H, 5.51. H NMR, δ: 5.50 (t, 1 H, H(4)); 5.41 (dd,
1 H, H(2)); 4.91 (br.s, 1 H, H(1)); 4.41 (dd, 1 H, H(3)); 3.95
(m, 1 H, H(5)); 3.84 (dd, 1 H, H(6)); 3.75 (dd, 1 H, H(6´)).
13C NMR, δ: 98.44 (C(1)); 72.68 (C(2)); 70.31 (C(4)); 70.17
(C(5)); 68.34 (C(3)); 61.32 (C(6)).
Methyl 2,6ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranoside, the yield
was 0.9 g (45%), [α]D +11.8 (c 1.33). Found (%): C, 62.68;
1
H, 5.71. H NMR (200 MHz, c 0.5 g mL–1), δ: 5.32 (dd, 1 H,
H(2)); 4.79 (br.s, 1 H, H(1)); 4.15 (dd, 1 H, H(3)); 4.02 (t, 1 H,
H(4)); 3.86 (m, 1 H, H(5)); 4.60 (m, 2 H, H(6), H(6´)).
13C NMR, δ: 98.47 (C(1)); 72.29 (C(2)); 70.38 (C(5)); 69.76
(C(3)); 67.60 (C(4)); 63.48 (C(6)).
1,3,6ꢀTriꢀOꢀacetylꢀ2,4ꢀdiꢀOꢀbenzoylꢀDꢀmannopyranose (3).
A cold solution of concentrated H2SO4 (0.1 mL) in Ac2O
(2.5 mL) was added to a solution of methyl glycoside 2 (0.34 g,
0.81 mmol) in AcOH (2.5 mL) and Ac2O (2.5 mL) cooled to 0 °C.
The mixture was heated at 40 °C for 2 h. After completion of the
reaction (TLC control), the solution was cooled to ∼20 °C,
water was added dropwise, and the solution was concentrated
in vacuo. The residue was dissolved in toluene and the solution

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Abronina et al.
was concentrated. A solution of the residue in chloroform was
washed with a NaHCO3 solution. Acetate 3 was obtained in a
yield of 0.41 g (98.5%), [α]D –39.1 (c 2.55). Found (%): C, 61.21;
H, 5.24. C26H26O10. Calculated (%): C, 60.70; H, 5.09.
1H NMR, δ: 6.30 (br.s, 1 H, H(1)); 5.80 (t, 1 H, H(4)); 5.68 (dd,
1 H, H(3)); 5.57 (dd, 1 H, H(2)); 4.40—4.20 (m, 3 H, H(5),
H(6), H(6´)). 13C NMR, δ: 90.63 (C(1)); 70.66 (C(5)); 68.94
(C(3)); 68.74 (C(2)); 66.43 (C(4)); 62.49 (C(6)).
αꢀDꢀ[1ꢀ13C]mannopyranosyl bromide (210 mg, 0.318 mmol) in
a yield of 208 mg (69%).
Methyl 2,4ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀ(2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀ
mannopyranosyl)ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀmannoꢀ
pyranosyl)ꢀαꢀDꢀmannopyranoside (6). A ∼0.6 M hydrogen chloꢀ
ride solution (8 mL) in MeOH, which was prepared by the
addition of acetyl chloride (0.4 mL) to methanol (10 mL) at 0 °C,
was added to a solution of diacetate 5 (110 mg, 0.077 mmol) in
CH2Cl2 (2 mL). The solution was kept at ∼20 °C for ∼14 h. After
evaporation of the solvent, diol 6 was obtained in a yield of
100 mg (96%), [α]D –46.6 (c 0.7). Found (%): C, 65.96; H, 5.13.
C75H66O24. Calculated (%): C, 66.66; H, 4.92. The NMR specꢀ
troscopic data are given in Tables 1 and 2.
Under analogous conditions, methyl 2,4ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀ
(2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranosyl)ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀ
OꢀbenzoylꢀαꢀDꢀ[1ꢀ13C]mannopyranosyl)ꢀαꢀDꢀmannopyranoside
(6ꢀ13C) was prepared from trisaccharide diacetate 5ꢀ13C (188 mg,
0.131 mmol) in a yield of 170 mg (96%).
Methyl 6ꢀOꢀ[3,6ꢀdiꢀOꢀ(αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannoꢀ
pyranosyl]ꢀ3ꢀOꢀ(αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranoside
(1). Calcined 4 Å molecular sieves were added to a solution of
trisaccharide diol 6 (150 mg, 0.11 mmol) and 2,3,4,6ꢀtetraꢀOꢀ
benzoylꢀαꢀDꢀmannopyranosyl bromide (263 mg, 0.4 mmol) in
CH2Cl2 (3 mL). The reaction mixture was stirred at 0 °C for 1 h
and a solution of AgOTf (257 mg, 0.44 mmol) in toluene (2 mL)
was added. Then the reaction mixture was stirred at ∼20 °C
for 3 h, diluted with chloroform, filtered through a layer of
Hyflo superꢀcel, washed with a Na2S2O3 solution, and concenꢀ
trated. Column chromatography afforded methyl 2,4ꢀdiꢀOꢀbenꢀ
zoylꢀ6ꢀOꢀ[2,4ꢀdiꢀOꢀbenzoylꢀ3,6ꢀdiꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀ
αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranosyl]ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀ
OꢀbenzoylꢀαꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranoside (7) in a
yield of 202 mg (71.5%), [α]D –43.5 (c 0.5). The NMR spectroꢀ
scopic data are given in Tables 1 and 2.
The fully protected pentasaccharide (100 mg) was dissolved
in dry pyridine (2 mL). Then anhydrous MeOH (1 mL) and
0.5 M MeONa (2 mL) were added. The reaction mixture was
kept at ∼20 °C for 2 days and neutralized with Dowexꢀ50 (H+)
cationꢀexchange resin, which was prewashed with methanol.
The resin was filtered off, the filtrate was concentrated, the
residue was dissolved in water, the solution was washed with
chloroform, and the aqueous layer was concentrated. The target
mannopentaoside was isolated by gelꢀpermeation chromatoꢀ
graphy (Fractogel TSK HWꢀ40 (S), 1.1×80 cm, 0.1 M AcOH as
the eluent, a Knauer (Germany) differential refractometer as
the detector), the yield was 30 mg (91%), [α]D +99.8 (c 2.0,
water) (cf. lit data: [α]D +108.1,12 +87.5,14 +100 20).
Methyl 2,4ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀ(3,6ꢀdiꢀOꢀacetylꢀ2,4ꢀdiꢀOꢀ
benzoylꢀαꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranoside (4). Acetic
acid (1.5 mL), then AcBr (0.32 mL, 4.25 mmol, 5 equiv.), and
aqueous AcOH (80 µL of H2O in 1.6 mL of AcOH) were added
to a solution of acetate 3 (0.44 g, 0.85 mmol) in chloroform
(1.5 mL) cooled to 0 °C. The mixture was kept at ∼20 °C for
∼16 h and diluted with chloroform. The solution was washed
with ice water, a NaHCO3 solution cooled to 0 °C, and water,
dried by filtration through a layer of cotton, and concentrated.
3,6ꢀDiꢀOꢀacetylꢀ2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannopyranosyl broꢀ
mide was obtained in a yield of 0.42 g (92%), [α]D –10.1 (c 1.09).
A solution of 3,6ꢀdiol 2 (0.24 g, 0.6 mmol), Hg(CN)2
(152 mg, 0.6 mmol), and HgBr2 (43 mg, 0.12 mmol) in MeCN
(3 mL) was stirred with ground 3 Å molecular sieves for 1 h and
then a solution of the glycosyl bromide obtained in MeCN (2 mL)
was added. After 30 min (disappearance of the glycosyl bromide,
TLC control), the reaction mixture was filtered and the filtrate
was diluted with chloroform. The solution was washed with
water and a KI solution and concentrated. The residue was
chromatographed. Disaccharide derivative 4 was obtained in a
yield of 293 mg (57%), [α]D –19.3 (c 1.77). Found (%): C, 63.76;
H, 5.32. C45H44O17. Calculated (%): C, 63.08; H, 5.18.
1H NMR, δ: 5.76 (dd, 1 H, H(3c)); 5.67 and 5.68 (both t,
1 H each, H(4a), H(4c)); 5.60 (dd, 1 H, H(2c)); 5.47 (dd, 1 H,
H(2a)); 5.06 (br.s, 1 H, H(1c)); 4.98 (br.s, 1 H, H(1a)); 4.41
(dd, 1 H, H(3a)); 4.23 (m, 1 H, H(5a)); 4.17 (m, 1 H, H(5c));
4.07 (dd, 1 H, H(6c)); 4.04 (dd, 1 H, H(6´c)); 4.01 (dd, 1 H,
H(6a)); 3.74 (dd, 1 H, H(6´a)). 13C NMR, δ: 98.70 (C(1a));
97.32 (C(1c)); 72.88 (C(2a)); 70.15 (C(4a)); 70.06 (C(2c)); 69.15
(C(3a)); 69.11 (C(3c)); 69.00 (C(5a)); 68.78 (C(5c)); 66.81
(C(4c)); 66.49 (C(6a)); 62.56 (C(6c)).
Methyl 2,4ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀ(3,6ꢀdiꢀOꢀacetylꢀ2,4ꢀdiꢀOꢀ
benzoylꢀαꢀDꢀmannopyranosyl)ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀ
Dꢀmannopyranosyl)ꢀαꢀDꢀmannopyranoside (5). A solution of the
monohydroxy disaccharide derivative 4 (205 mg, 0.24 mmol)
and 2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀmannopyranosyl bromide
(237 mg, 0.36 mmol) in freshly distilled CH2Cl2 (2 mL) was
stirred with 4 Å molecular sieves at ∼20 °C for 1 h and cooled
to 0 °C. Then a solution of AgOTf (0.1 g, ∼0.4 mmol) in toluene
(2 mL) was added. After 1 h, the reaction mixture was diluted
with chloroform. The resulting solution was filtered through a
layer of Hyflo superꢀcel, washed with water and a Na2S2O3
solution, and concentrated. Column chromatography afforded
fully protected trisaccharide 5 in a yield of 269 mg (78%),
[α]D –43.6 (c 0.5). Found (%): C, 66.52; H, 5.01. C79H70O26.
Calculated (%): C, 66.10; H, 4.92. The NMR spectroscopic
data are given in Tables 1 and 2.
Condensation of trisaccharide glycosyl acceptor 6ꢀ13C
(123 mg, 0.091 mmol) with 2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀ
mannopyranosyl bromide (180 mg, 0.273 mmol) in the presence
of AgOTf (77 mg, 0.3 mmol) and 4 Å molecular sieves in a
mixture of CH2Cl2 (2 mL) and toluene (1 mL) was performed
analogously to the aboveꢀdescribed synthesis of protected
mannopentaoside 7 to give methyl 2,4ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀ[2,4ꢀ
diꢀOꢀbenzoylꢀ3,6ꢀdiꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀmannoꢀ
pyranosyl)ꢀαꢀDꢀmannopyranosyl]ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀ
αꢀDꢀ[1ꢀ13C]mannopyranosyl)ꢀαꢀDꢀmannopyranoside (7ꢀ13C) in a
yield of 160 mg (68%).
Under analogous condensation conditions, methyl 2,4ꢀdiꢀ
Oꢀbenzoylꢀ6ꢀOꢀ(3,6ꢀdiꢀOꢀacetylꢀ2,4ꢀdiꢀOꢀbenzoylꢀαꢀDꢀmannoꢀ
pyranosyl)ꢀ3ꢀOꢀ(2,3,4,6ꢀtetraꢀOꢀbenzoylꢀαꢀDꢀ[1ꢀ13C]mannoꢀ
pyranosyl)ꢀαꢀDꢀmannopyranoside (5ꢀ13C) was prepared from diꢀ
saccharide 4 (180 mg, 0.21 mmol) and 2,3,4,6ꢀtetraꢀOꢀbenzoylꢀ
Deacylation of this benzoate (50 mg) under the aboveꢀdeꢀ
scribed conditions afforded the target methyl 6ꢀOꢀ[3,6ꢀdiꢀ

3,6ꢀBranched 13Cꢀlabeled mannopentaoside
Russ.Chem.Bull., Int.Ed., Vol. 54, No. 5, May, 2005
1293
Oꢀ(αꢀDꢀmannopyranosyl)ꢀαꢀDꢀmannopyranosyl]ꢀ3ꢀOꢀ(αꢀDꢀ
[1ꢀ13C]mannopyranosyl)ꢀαꢀDꢀmannopyranoside (1ꢀ13C) in a yield
of 7 mg (43%). The 1H NMR spectrum is shown in Fig. 1.
9. T. Kawasaki, Biochim. Biophys. Acta, 1999, 1473, 186.
10. D. C. Kilpatrick, Biochim. Biophys. Acta, 2002, 1572, 401.
11. Y. Ma, K. Uemura, S. Oga, Y. Kozutsumi, N. Kawasaki,
and T. Kawasaki, Proc. Natl. Acad. Sci. USA, 1999, 99, 371.
12. T. Ogawa and K. Sasajima, Carbohydr. Res., 1981, 97, 205.
13. Y. Du, M. Zhang, and F. Kong, Tetrahedron, 2001, 57, 1757.
14. L. V. Backinowsky, P. I. Abronina, A. S. Shashkov, A. A.
Grachev, N. K. Kochetkov, S. A. Nepogodiev, and J. F.
Stoddart, Chem. Eur. J., 2002, 8, 4412.
This study was financially supported by the Russian
Foundation for Basic Research (Project No. 04ꢀ03ꢀ32855)
and the Council on Grants of the President of the Russian
Federation (Program for Support of Leading Scienꢀ
tific Schools of the Russian Federation, Grant
NShꢀ1557.2003.3).
15. N. Smilijanic, S. Halila, V. Moreau, and F. DjedaïniꢀPilard,
Tetrahedron Lett., 2003, 44, 8999.
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247, 323.
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in revised form April 18, 2005