Enhanced sialylating activity of O-chloroacetylated 2-thioethyl sialosides

Article abstract of DOI:10.1055/s-2005-868514

It has been shown that O-chloroacetyl protecting groups enhance significantly sialylating activity of 2-thioethyl sialosides in model reactions of α(2-8)-sialylation. Ethyl 4,7,8,9-tetra-O-chloroacetyl-3,5-dideoxy-2- thio-5-trifluoroacetamido-D-glycero-α-

Full text of DOI:10.1055/s-2005-868514

LETTER
1375
Enhanced Sialylating Activity of O-Chloroacetylated 2-Thioethyl Sialosides
E
nhanced Sialylati
u
ng Activity ry E. Tsvetkov,* Nikolay E. Nifantiev
N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Leninsky Prospekt 47, 119991 Moscow, Russian Federation
Fax +7(095)1358784; E-mail: tsvetkov@ioc.ac.ru
Received 21 January 2005
Dedicated to Professor Richard R. Schmidt on the occasion of his 70th birthday
acceptor. The improved glycosyl donor properties of di-
Abstract: It has been shown that O-chloroacetyl protecting groups
N-acetyl8 and N-trifluoroacetyl (N-TFA) derivatives of
neuraminic acid were explained by minimization of side
reactions at the N-acyl moieties due to reducing their
enhance significantly sialylating activity of 2-thioethyl sialosides in
model reactions of a(2-8)-sialylation. Ethyl 4,7,8,9-tetra-O-chloro-
acetyl-3,5-dideoxy-2-thio-5-trifluoroacetamido-D-glycero-a-D-ga-
lacto-non-2-ulopyranoside that combines electron-withdrawing O- nucleophilicity.7,9
chloroacetyl and N-trifluoroacetyl protecting groups displayed the
We supposed that electron-withdrawing N-protecting
best reactivity and enabled the most efficient synthesis of the
Neu5Aca(2-8)Neu5Ac dimer. The GD3 tetrasaccharide was synthe-
sized in stepwise manner using this sialyl donor in the key (2-8)-
coupling, albeit the yield was noticeably lower than in the model
sialylation of monosaccharide acceptors.
groups are able also to increase the positive charge on C-
2 of all cationic species (such as sialyl or sialyl nitrilium10
cations) involved in sialylation and increase thereby their
reactivity towards nucleophiles. If this is the case, elec-
tron-withdrawing O-protecting group would produce the
same effect on the reactivity of sialyl donors. Here we de-
scribe the effect of O-chloroacetyl protecting groups in 2-
thioethyl a-sialosides on their efficiency as sialyl donors
in a(2-8)-sialylation. O-Chloroacetyl groups can be easily
introduced, are stable under sialylation conditions, and
can be removed selectively, if necessary, in the presence
of other acyl protections.
Key words: chloroacetyl protecting group, 2-thioethyl sialoside,
glycosylation, reactivity, GD3 oligosaccharide
The disaccharide sequence Neu5Aca(2-8)Neu5Ac is a
constituent of a number of gangliosides such as GD3,
GT1a, GQ1b, and others, and plays a key role in their bio-
logical activities.1 The simplest representative of this
group, the ganglioside GD3 (1), was shown to be the
human melanoma associated antigen (Figure 1).2
To facilitate NMR analysis of compounds, a-thioethyl
sialoside 3 but not a more readily available a,b-mixture3
was chosen as a starting material for the preparation of O-
chloroacetylated sialyl donors. a-Thiosialosides can be
obtained more easily in anomerically pure form than the
corresponding b-anomers. Compound 3 was synthesized
from chloride 211 according to the modified published
procedure12 followed by deacetylation in 79% yield
(Scheme 1). Since fully acetylated derivatives of Neu5Ac
are conventional sialyl donors, tetraacetate 4a was chosen
as a reference point. Its chloroacetylated analog 4b was
prepared from 3 by reaction with chloroacetic anhydride
in the presence of NaHCO3.13 O-Acetyl and O-chloro-
acetyl N-TFA thioethyl sialoside 7a and 7b14 were synthe-
sized from 3 by acidic de-N-acetylation15 followed by N-
trifluoroacetylation of the amino group in 5 and O-acyla-
tion of 6 formed with Ac2O or (ClCH2CO)2O.
Figure 1
Chemical preparation of the Neu5Aca(2-8)Neu5Ac dimer
is strongly complicated by the low reactivity of the 8-OH
group of neuraminic acid and remains so far one of the
main problems of the oligosaccharide synthesis. The most
of successful syntheses were based on application of mod-
ified sialyl donors bearing an additional function at C-3
which can control the stereochemistry of substitution at
the anomeric center and prevent 2,3-elimination.3 Multi-
step synthetic procedures are required for preparation of
these donors and an additional step is needed for removal
of the auxiliary from the sialylation product.4 An alterna-
tive strategy5 consisting in the use of the Neu5Aca(2-
8)Neu5Ac dimer obtained by mild acid hydrolysis of
colominic acid6 circumvents the problems of the chemical
synthesis of the a(2-8)-linkage but is limited by low avail-
ability of the starting material. Recently, a promising ap-
proach was reported7 that employed N-trifluoroacetylated
derivatives of neuraminic acid as both sialyl donor and
7,8-Diol 14 was used as a sialyl acceptor in model sialyla-
tion reactions. It could be selectively sialylated at 8-OH
because of its much higher reactivity as compared to that
of 7-OH.4c,16 Diol 14 was prepared as follows (Scheme 2).
The starting a-benzyl sialoside 8 was synthesized from
chloride 2 and benzyl alcohol according to the described
procedure17 followed by de-O-acetylation. Treatment of 8
with 2,2-dimethoxypropane and CSA in acetone afforded
8,9-isopropylidene derivative 9 which was selectively
benzoylated at O-4 with benzoyl chloride to give
monobenzoate 10. Removal of the isopropylidene group
followed by treatment of triol 11 with a,a-dimethoxytolu-
ene in the presence of CSA yielded a mixture of 7,9- and
SYNLETT 2005, No. 9, pp 1375–1380
0
1.
0
6.
2
0
0
5
Advanced online publication: 29.04.2005
DOI: 10.1055/s-2005-868514; Art ID: G03405ST
© Georg Thieme Verlag Stuttgart · New York

1376
Y. E. Tsvetkov, N. E. Nifantiev
LETTER
Scheme 1 Reagents and conditions: a) EtSNa, MeCN, r.t.; b) MeO-
Na, MeOH, r.t., 79% over two steps; c) MsOH, MeOH, reflux; d)
Et3N, CF3CO2Et, MeOH, r.t., 84% over two steps; e) Ac2O, pyridine,
r.t., 96%; f) (ClCH2CO)2O, NaHCO3, DMF, r.t., 84% for 4b, 93% for
7b.
8,9-benzylidene derivatives 12 and 13 in a ratio of about
1:1. This mixture was subjected, without separation of the
isomers, to reductive acetal ring-opening with
H3B·NMe3–AlCl3 in the presence of water18 to give 7,8-
diol 14 in high yield.
Scheme 3 Reagents and conditions: a) NIS, TfOH, MS 3 Å, MeCN,
–40 °C; b) (H2N)2CS, MeOH–HOAc (3:1), 7–10 min, r.t.
derivative 17. The transformation 14 → 17 could be
accomplished without chromatographic purification of
the intermediate compounds and afforded diol 17 in 85–
90% overall yield.
For comparison of the sialyl acceptor properties of 14 and
its N-TFA counterpart (cf. ref.7), diol 17 was also prepared
from 14 using the procedure of de-N-acylation of second-
ary amides via intermediate N-Boc-imide formation19
(Scheme 2). Diol 14 was converted into 7,8-O-isopro-
pylidene derivative which was treated, without isolation,
with Boc2O in the presence of DMAP to give imide 15.
Selective removal of the N-acetyl group in 15 with hydra-
zine hydrate afforded Boc-protected amine 16. Simulta-
neous cleavage of acid-labile isopropylidene and Boc
groups followed by N-trifluoroacetylation yielded N-TFA
Then NIS/TfOH-promoted sialylation of acceptors 14 and
17 with two equivalents of thioglycosides 4a,b and 7a,b
was examined20 (Scheme 3). As we reported earlier,21 gly-
cosylation of substrates containing acetamido groups with
ethyl thioglycosides can be accompanied by the transfer
of SEt residue onto these groups resulting in the formation
of EtS(Ac)N-derivatives. This side reaction strongly com-
plicated analysis and separation of reaction mixtures and
Scheme 2 Reagents and conditions: a) BnOH, Ag2CO3, AgClO4, MS 4 Å, CH2Cl2, r.t.; b) MeONa, MeOH, r.t., 90% over two steps; c)
Me2C(OMe)2, CSA, acetone, r.t., 94% for 9; d) BzCl, pyridine–CH2Cl2, 0 °C, 88%; e) 80% aq HOAc, 40° C, 94%; f) PhCH(OMe)2, CSA,
MeCN, r.t.; g) H3B·NMe3, AlCl3, H2O, THF, r.t., 81% over two steps; h) Boc2O, DMAP, THF, reflux, 97% over two steps; i) N2H4·H2O, DMF,
r.t., 95%; j) 90% aq CF3COOH, CH2Cl2, r.t.; k) CF3COOC6F5, Et3N, CH2Cl2, r.t., 91% over two steps.
Synlett 2005, No. 9, 1375–1380 © Thieme Stuttgart · New York

LETTER
Enhanced Sialylating Activity
1377
decreased the yields of target products. It was found that 1H NMR spectra of both 18 and 19 showed that they were
a brief treatment with thiourea in a mixture of MeOH– (2-8)-linked disaccharides.
HOAc22 caused complete conversion of EtS(Ac)N moiety
The known empirical rules for the assignment of the ano-
into the parent acetamide, whereas chloroacetyl groups re-
meric configuration of tetraacetylated Neu5Ac residue3
mained intact under these conditions. To provide a reli-
were found to be also applicable to O-chloroacetyl and N-
TFA sialosides. Although signals for H-4 of the glycosy-
lating residue in N-TFA a-disaccharides 18c–e appeared
at d = 5.0–5.1 (i.e. shifted by ca. 0.2 ppm downfield as
compared to signals for H-4 in N-acetyl derivatives
18a,b), this region did not overlap that of corresponding
able comparison of results of the model reactions,
sialylation reaction mixtures were subjected to the above
procedure. The disaccharide products were separated
from unreacted sialyl acceptors and monosaccharide by-
products by gel permeation chromatography. Pure ano-
mers 18 and 19 were isolated by HPLC. The results are
b-anomers 19c–e (d > 5.40 ppm). The difference in
summarized in Table 1.
Dd{H9¢–H9} values of the glycosylating Neu moiety for
As expected, acetylated donor 4a gave low yield of the a-anomers 18 (0.28–0.37 ppm) and b-anomers 19 (0.45–
corresponding a(2-8)-linked product 18a (entry 1). Re- 0.63 ppm) was not as pronounced as in ‘conventional’ sia-
placement of O-acetyl groups by chloroacetyl ones (donor losides, but nonetheless allowed reliable configuration as-
4b, entry 2) increased two-fold the yield of 18. Essentially signment within each anomeric pair. Coupling constant
the same effect gave rise N-TFA group (donor 7a, entry values J7,8 of the non-reducing Neu residue were also
3). Combination of O-chloroacetyl and N-TFA groups characteristic for its anomeric configuration being 8.0–8.6
(donor 7b, entry 4) resulted in further increase of the yield Hz for a-anomers 18 and 2.8–4.6 Hz for b-anomers 19.
of 18; this yield is comparable with those obtained on sia-
We have found some other regularities in 1H NMR spectra
lylation of 3-OH in many galactose derivatives.3 N-TFA
of 18 and 19 which may be useful for the assignment of
sialyl acceptor 17 (entry 5) displayed higher reactivity7
than the N-acetyl counterpart 14 and allowed preparation
of a-dimer 18e in excellent yield of 55%. This is the high-
est yield of a(2-8)-disaccharide achieved on the use of a
the anomeric configuration in (2-8)-dimers of neuraminic
acid. Thus, irrespective of the protecting groups pattern,
the signals for H-6 in the glycosylating Neu residue ap-
peared at d = 3.95–4.15 for a-anomers 18 and at d = 4.55–
4.89 for b ones 19. The signals for H-8 in the glycosylated
Neu residue located in the region of d = 4.40–4.58 for a-
disaccharides 18 and in the region of d = 4.02–4.07 for b-
non-modified at C-3 sialyl donor. Boons et al. prepared a
similar a(2-8)-dimer in the same yield of 55% but with
three equivalents of sialyl donor.7 It is noteworthy that
yields of the corresponding b-anomers 19 did not change
isomers 19.
much with protecting groups variation, and hence, a-ste-
Recently, Schmidt and co-workers25 described an effi-
cient synthesis of the ganglioside GD3 (1) that was based
on the consecutive introduction of two Neu5Ac residues
into an oligosaccharide chain. A sialyl donor with 3-SPh
reoselectivity also increased upon accumulation of elec-
tron-withdrawing groups in the sialyl donor molecule.
The increase of a-stereoselectivity may be attributed to
the stabilization of b-configurated sialyl nitrilium inter-
participating auxiliary was used in the second sialylation.
We applied sialyl donor 7b, which had displayed the best
reactivity in the model (2-8)-sialylations, to a similar
preparation of 2-aminoethyl glycoside of the GD3 tet-
rasaccharide. To minimize manipulations with protecting
groups in the stage of GM3 trisaccharides, selectively pro-
tected sialyl donors were employed in the first coupling
step. Their N-acetyl and N-TFA precursors 20a,b were
obtained in good overall yields from tetrols 3 and 6 using
the synthetic pathway described above for the transforma-
tion 8 → 14 (Scheme 4). Chloroacetylation of 20a,b
mediates (cf. ref.23) due to the –I effect of protecting
groups.
1
Structure of disaccharides 18 and 19 was proved by H
NMR spectroscopy data.24 Configurations of newly
formed sialoside bonds were deduced from HMBC spec-
tra (optimized for a coupling constant of 8 Hz). A strong
correlation between C-1¢ and H-3¢ax indicated trans-di-
axial orientation of the methoxycarbonyl group and the
proton H-3ax and, hence, a-configuration of 18. The lack
of C-1¢–H-3¢ax correlation in HMBC spectra of 19 proved
their b-configuration. The presence of signals for 7-OH in
Table 1 Effect of O- and N-Protecting Groups on Efficiency of (2-8)-Sialylation
Entry
Donor
4a
Acceptor
Products
18a, 19a
18b, 19b
18c, 19c
18d, 19d
18e, 19e
R1
R2
R3
Yield of 18 (%) Yield of 19 (%)
1
2
3
4
5
14
14
14
14
17
CH3CO
ClCH2CO
CH3CO
ClCH2CO
ClCH2CO
CH3
CH3
CF3
CF3
CF3
CH3
CH3
CH3
CH3
CF3
8
18
20
37
55
7
9
7
8
7
4b
7a
7b
7b
Synlett 2005, No. 9, 1375–1380 © Thieme Stuttgart · New York

1378
Y. E. Tsvetkov, N. E. Nifantiev
LETTER
Scheme 4 Reagents and conditions: a) see steps c–g in Scheme 1, 69% overall for 20a, 60% for 20b; b) ClCH2COCl, pyridine–CH2Cl2, 0 °C,
92% for both 21a,b; c) NIS, TfOH, MS 3 Å, MeCN, –40 °C, 62% for 23a, 72% for 23b; 15% for 25a, 17% for 25b; d) (H2N)2CS, 2,4,6-colli-
dine, MeOH, reflux, 97% for 24a, 93% for 24b; e) 1 M aq NaOH, MeOH, r.t.; f) Ac2O, MeOH, r.t.; g) H2, Pd(OH)2/C, MeOH, r.t.; h) CF3CO2Et,
MeOH, r.t.
afforded sialyl donors 21a,b26 in which chloroacetates are Anomeric configuration of the terminal neuraminic acid
activating and also selectively removable temporary pro- residue in 25a,b was proved by the data of HMBC spectra
tecting groups. NIS/TfOH-promoted sialylation of lacto- (vide supra) and the position of sialylation was confirmed
side 2227 with small excess (1.2 equiv) of 21a,b gave by the presence of signals for 7-OH of internal neuraminic
protected GM3 trisaccharides 23a,b28 in yields of 62% and acid in 1H NMR spectra of both tetrasaccharides. Chemi-
72%, respectively. Only minute amounts of the corre- cal shifts for H-4 (d = 5.05 and 5.07 ppm) and H-6
sponding b-anomers (6% for 23a and 3% for 23b) were (d = 4.12 and 4.08 ppm), and the values of J7,8 (8.8 and 8.2
isolated in both sialylations. Removal of chloroacetyl Hz) and Dd {H9–H9¢} (0.38 and 0.23) confirmed also the
groups from 23a,b with thiourea resulted in triols 24a,b a-configuration of the terminal neuraminic acid residues
almost quantitatively. Thus, the use of the 7,8-di-O-chlo- in 25a,b.
roacetyl-thiosialosides 21a,b provided highly efficient
Transformation of 25a,b into 2-aminoethyl glycoside of
and stereoselective sialylation of lactoside 22 and subse-
GD3 26 was accomplished in 6 steps involving alkaline
quent one-step, high-yielding deprotection of the sialyla-
hydrolysis of all ester and N-TFA groups followed by N-
acetylation; reduction of the spacer azide group by hydro-
tion products 23 into new sialyl acceptors 24.
However, sialylation of N-acetyl and N-TFA acceptors genation and subsequent temporary protection of arising
24a,b with two equivalents of thiosialoside 7b was not as amine by an N-TFA group; hydrogenolysis of benzyl
efficient as could be expected from the results of model ethers; and, finally, alkaline hydrolysis with liberation of
disaccharide syntheses and gave GD3 tetrasaccharide de- the spacer amino group in 79% overall yield. 1H NMR and
rivatives 25a,b29 in modest and practically equal yields of 13C NMR data of 2630 were almost identical to those re-
15% and 17%, respectively. TLC examination of tetrasac- cently published for the GD3 3-azidopropyl glycoside.31
charide fractions obtained after gel permeation
In conclusion, we have found that O-chloroacetylation in-
crease considerably the reactivity of 2-thioethyl sialosides
chromatography20 revealed only traces of products that
might be identified as the corresponding b-anomers.
in sialylation reactions. 2-Thiosialoside 7b, which com-
About 70% of the acceptor was recovered in both cases.
bines O-chloroacetyl and N-TFA protecting groups,
showed the best sialylating activity and provided the most
efficient synthesis of the a(2-8)-linked dimer of
neuraminic acid.
We suppose that such a considerable decrease of efficien-
cy of (2-8)-sialylation compared to model experiments,
and the lack of the difference in reactivity of the N-acetyl
and N-TFA acceptors, may be caused by additional steric
hindrances around the 8-OH group that are created by the
bulky benzylated disaccharide aglycon in the acceptors
Acknowledgment
24.
This work was supported by The Russian Foundation for Basic
Research, Grant No. 03-03-32567. The authors are indebted to Mr.
A. A. Grachev for recording NMR spectra.
Synlett 2005, No. 9, 1375–1380 © Thieme Stuttgart · New York

LETTER
Enhanced Sialylating Activity
1379
chromatography on a column with Bio Beads S-X3 (Bio-
Rad) in toluene. Disaccharide fractions were pooled and
concentrated to give a mixture of 18 and 19. Pure anomers
were isolated by preparative HPLC (LiChrosorb Si-60
column, K-2401 RI detector, both from Knauer) using
toluene–acetone mixtures as eluents.
References
(1) Lloyd, K. O.; Furukawa, K. Glycoconjugate J. 1998, 15,
627.
(2) Nudelman, E.; Hakomori, S.; Kannagi, R.; Levery, S.; Yeh,
M.-Y.; Hellström, K. E.; Hellström, I. J. Biol. Chem. 1982,
257, 12752.
(21) (a) Nifant’ev, N. E.; Tsvetkov, Y. E.; Shashkov, A. S.;
Kononov, L. O.; Menshov, V. M.; Tuzikov, A. B.; Bovin, N.
V. J. Carbohydr. Chem. 1996, 15, 939. (b) Nifant’ev, N. E.;
Sherman, A. A.; Yudina, O. N.; Cheshev, P. E.; Tsvetkov, Y.
E.; Khatuntseva, E. A.; Kornilov, A. V.; Shashkov, A. S.
Pure Appl. Chem. 2004, 76, 1705.
(3) Boons, G.-J.; Demchenko, A. V. Chem. Rev. 2000, 100,
4539.
(4) (a) Ito, Y.; Ogawa, T. Tetrahedron 1990, 46, 89.
(b) Ercegovic, T.; Magnusson, G. J. Org. Chem. 1995, 60,
3378. (c) Castro-Palomino, J. C.; Tsvetkov, Y. E.; Schmidt,
R. R. J. Am. Chem. Soc. 1998, 120, 5434.
(5) Ishida, H.; Ohta, Y.; Tsukada, Y.; Kiso, M.; Hasegawa, A.
Carbohydr. Res. 1993, 246, 75.
(6) Roy, R.; Pon, R. A. Glycoconjugate J. 1990, 7, 3.
(7) De Meo, C.; Demchenko, A. V.; Boons, G.-J. J. Org. Chem.
2001, 65, 5490.
(8) Demchenko, A. V.; Boons, G.-J. Chem.–Eur. J. 1999, 5,
1278.
(9) De Meo, C.; Demchenko, A. V.; Boons, G.-J. Austr. J.
Chem. 2002, 55, 131.
(10) Schmidt, R. R. ACS Symp. Ser. 1994, 560, 276.
(11) Kuhn, R.; Lutz, P.; MacDonald, D. L. Chem. Ber. 1966, 99,
611.
(12) Lubineau, A.; Le Gallic, J. J. Carbohydr. Chem. 1991, 10,
263.
(13) Chittenden, G. J. F.; Regeling, H. Recl. Trav. Chim. Pays-
Bas 1987, 106, 44.
(14) Analytical data for 7b: mp 145–146 °C (EtOAc–hexane);
[a]D +24 (c 1, CHCl3). 1H NMR (500 MHz, CDCl3): d = 1.21
(t, 3 H, J = 7.5 Hz, SCH2CH3), 2.03 (t, 1 H, J3ax,3eq = 12.9 Hz,
(22) Kessler, W.; Iselin, B. Helv. Chim. Acta 1966, 49, 1330.
(23) Yu, C.-S.; Niikura, K.; Lin, C.-C.; Wong, C.-H. Angew.
Chem. Int. Ed. 2001, 40, 2900.
(24) Selected 1H NMR assignments for anomeric (2-8)-
disialosides 18 and 19.
Compound 18a: d = 4.86 (H-4¢), 3.95 (H-6¢), J7¢,8¢ = 8.3 Hz,
4.52 (H-8), Dd{H-9a¢–H-9b¢} = 0.28.
Compound 19a: d = 5.31 (H-4¢), 4.55 (H-6¢), J7¢,8¢ = 4.3 Hz,
4.06 (H-8), Dd{H-9a¢–H-9b¢} = 0.51.
Compound 18b: d = 4.88 (H-4¢), 3.97 (H-6¢), J7¢,8¢ = 8.7 Hz,
4.56 (H-8), Dd{H-9a¢–H-9b¢} = 0.37.
Compound 19b: d = 5.37 (H-4¢), 4.72 (H-6¢), J7,8¢ = 2.8 Hz,
4.07 (H-8), Dd{H-9a¢–H-9b¢} = 0.60.
Compound 18c: d = 5.03 (H-4¢), 4.11 (H-6¢), J7¢,8¢ = 8.0 Hz,
4.53 (H-8), Dd{H-9a¢–H-9b¢} = 0.25.
Compound 19c: d = 5.42 (H-4¢), 4.76 (H-6¢), J7¢,8¢ = 4.6 Hz,
4.03 (H-8), Dd{H-9a¢–H-9b¢} = 0.45.
Compound 18d: d = 5.06 (H-4¢), 4.15 (H-6¢), J7¢,8¢ = 8.6 Hz,
4.58 (H-8), Dd{H-9a¢–H-9b¢} = 0.36.
Compound 19d: d = 5.47 (H-4¢), 4.89 (H-6¢), J7¢,8¢ = 2.8 Hz,
4.02 (H-8), Dd{H-9a¢–H-9b¢} = 0.63.
J3ax,4 = 12.3 Hz, H-3ax), 2.55 (dq, 1 H, Jgem = 12.3 Hz,
SCH2CH3), 2.75 (dq, 1 H, SCH2CH3), 2.85 (dd, 1 H,
J3eq,4 = 4.8 Hz, H-3eq), 3.83 (s, 3 H, CH3O), 3.99, 4.02 (2 d,
2 H, Jgem = 15.0 Hz, ClCH2), 4.07 (m, 4 H, H-5, H-6,
ClCH2), 4.17, 4.20 (2 d, 2 H, Jgem = 14.6 Hz, ClCH2), 4.20,
4.32 (2 d, 2 H, Jgem = 15.2 Hz, ClCH2), 4.25 (dd, 1 H,
J9,9¢ = 12.7 Hz, J9,8 = 5.3 Hz, H-9), 4.58 (dd, 1 H, J9¢,8 = 2.5
Hz, H-9¢), 5.08 (m, 1 H, J4,5 = 10.0 Hz, H-4), 5.38 (d, 1 H,
Compound 18e: d = 5.09 (H-4¢), 4.13 (H-6¢), J7¢,8¢ = 8.3 Hz,
4.40 (H-8), Dd{H-9a¢–H-9b¢} = 0.32.
Compound 19e: d = 5.47 (H-4¢), 4.82 (H-6¢), J7¢,8¢ = 2.8 Hz,
4.03 (H-8), Dd{H-9a¢–H-9b¢} = 0.60.
(25) Castro-Palomino, J. C.; Simon, B.; Speer, O.; Leist, M.;
Schmidt, R. R. Chem.–Eur. J. 2001, 7, 2178.
(26) Analytical data for 21a: [a]D +39 (c 1, CHCl3). 1H NMR
(500 MHz, CDCl3): d = 1.22 (t, 3 H, J = 7.5 Hz, SCH2CH3),
1.78 (s 3 H, CH3CO), 2.11 (t, 1 H, J3ax,3eq = 12.7 Hz,
J7,8 = 8.0 Hz, H-7), 5.50 (m, 1 H, H-8), 6.91 (d, 1 H,
JNH,5 = 9.9 Hz, NH).
(15) Sugata, T.; Kan, Y.; Nagaregava, Y.; Miyamoto, T.;
Higuchi, R. J. Carbohydr. Chem. 1997, 16, 917.
J3ax,4 = 12.2 Hz, H-3ax), 2.58 (dq, 1 H, Jgem = 12.3 Hz,
SCH2CH3), 2.80 (dq, 1 H, SCH2CH3), 2.94 (dd, 1 H,
J3eq,4 = 4.6 Hz, H-3eq), 3.56 (dd, 1 H, J9,9¢ = 11.0 Hz,
(16) (a) Ercegovic, T.; Magnusson, G. J. Org. Chem. 1996, 61,
179. (b) Castro-Palomino, J. C.; Tsvetkov, Y. E.; Schneider,
R.; Schmidt, R. R. Tetrahedron Lett. 1997, 38, 6837.
(17) Hasegawa, A.; Ito, Y.; Ishida, H.; Kiso, M. J. Carbohydr.
Chem. 1989, 8, 125.
(18) Sherman, A. A.; Mironov, Y. V.; Yudina, O. N.; Nifantiev,
N. E. Carbohydr. Res. 2003, 338, 697.
(19) Grehn, L.; Gunnarson, K.; Ragnarsson, U. J. Chem. Soc.,
Chem. Commun. 1985, 1317.
J9,8 = 4.6 Hz, H-9), 3.77 (dd, 1 H, J9¢,8 = 3.5 Hz, H-9¢), 3.86
(s, 3 H, CH3O), 3.96 (dd, 1 H, J6,7 = 1.4 Hz, H-6), 4.06, 4.23
(2 d, 2 H, Jgem = 14.7 Hz, ClCH2), 4.22, 4.38 (2 d, 2 H,
Jgem = 15.0 Hz, ClCH2), 4.29 (q, 1 H, J5,6 = 10.7 Hz, H-5),
4.49, 4.53 (2 d, 2 H, Jgem = 11.8 Hz, PhCH2), 5.07 (m, 1 H,
J4,5 = 11.0 Hz, H-4), 5.41 (d, 1 H, JNH,5 = 9.9 Hz, NH), 5.49
(m, 1 H, H-8), 5.55 (dd, 1 H, J7,8 = 8.8 Hz, H-7), 7.17–7.98
(m, 10 H, arom.).
(20) General Procedure for Model Sialylation.
A mixture of sialyl donor (0.33 mmol), sialyl acceptor
(0.165 mmol) and powdered molecular sieves 3 Å (1.5 g) in
MeCN (4 mL) was stirred at r.t. for 2 h, then cooled to –40
°C. Then, NIS (149 mg, 0.66 mmol) and TfOH (6 mL, 0.07
mmol) were added and the resulting mixture was stirred at
–40 °C for 1.5 h. A drop of pyridine was added, the mixture
was diluted with CHCl3 and filtered through Celite. The
solids were washed with CHCl3, combined filtrates were
washed with 1 M Na2S2O3 solution, H2O, and concentrated.
The residue was dissolved in a mixture of MeOH (3 mL) and
HOAc (1 mL) and thiourea (20 mg) was added. After being
stirred for 7–8 min, the mixture was diluted with CHCl3,
thoroughly washed with H2O, and the solvent was
evaporated. The residue was subjected to gel permeation
(27) Cheshev, P. E.; Khatuntseva, E. A.; Tsvetkov, Y. E.;
Shashkov, A. S.; Nifantiev, N. E. Russ. J. Bioorg. Chem.
2004, 30, 60.
(28) Analytical data for 23a: [a]D +11 (c 1, CHCl3). 1H NMR
(500 MHz, CDCl3): d = 1.78 (s, 3 H, CH3CO), 2.14 (t, 1 H,
J3ax,3eq = 12.6 Hz, J3ax,4 = 11.4 Hz, H-3ax NeuAc), 2.73 (dd,
1 H, J3eq,4 = 4.5 Hz, H-3eq NeuAc), 3.73, 4.02 (2 d, 2 H,
Jgem = 14.9 Hz, ClCH2), 3.81 (s, 3 H, CH3O), 4.15, 4.29 (2 d,
2 H, Jgem = 15.0 Hz, ClCH2), 4.40 (d, 1 H, J1,2 = 8.0 Hz, H-1
Glc), 4.65 (d, 1 H, J1,2 = 7.8 Hz, H-1 Gal), 5.04 (m, 1 H,
J4,5 = 10.3 Hz, H-4 NeuAc), 5.35 (d, 1 H, JNH,5 = 10.1 Hz,
NH), 5.51 (dd, 1 H, J7,6 = 1.7 Hz, J7,8 = 9.0 Hz, H-7 NeuAc),
5.57 (m, 1 H, H-8 NeuAc).
Synlett 2005, No. 9, 1375–1380 © Thieme Stuttgart · New York

1380
Y. E. Tsvetkov, N. E. Nifantiev
LETTER
(29) Analytical data for 25a: [a]D –15 (c 1, CHCl3). 1H NMR (500
MHz, CDCl3): d = 1.92 (s, 3 H, CH3CO), 2.18 (t, 1 H,
(30) Analytical data for 26: [a]D +5 (c 0.5, H2O). 1H NMR (500
MHz, D2O, relative to internal acetone, dH = 2.225);
J
3ax,3eq = 13.2 Hz, J3ax,4 = 11.6 Hz, H-3ax NeuTFA), 2.21 (t,
terminal Neu5Ac: d = 2.03 (CH3CO), 2.78 (dd, J3eq,4 = 4.1
Hz, J3eq,3ax = 12.2 Hz, H-3eq), 1.74 (t, H-3ax), 3.67 (H-4),
3.83 (H-5), 3.61 (H-6), 3.58 (H-7), 3.88 (H-8), 3.65 (H-9),
3.87 (H-9¢); internal Neu5Ac: d = 2.06 (CH3CO), 2.68 (dd,
1 H, J3ax,3eq = 13.1 Hz, J3ax,4 = 11.2 Hz, H-3ax NeuAc), 2.61
(dd, 1 H, J3eq,4 = 4.8 Hz, H-3eq NeuAc), 2.77 (br d, 1 H, 4-
OH Gal), 2.82 (dd, 1 H, J3eq,4 = 4.7 Hz, H-3eq NeuTFA),
3.79 (s, 3 H, CH3O), 3.85 (s, 5 H, CH3O, ClCH2), 4.05 (s, 2
H, ClCH2), 4.12 (dd, 1 H, J6,5 = 10.5 Hz, J6,7 = 1.6 Hz, H-6
NeuTFA), 4.13, 4.16 (2 d, 2 H, Jgem = 15.0 Hz, ClCH2), 4.17,
4.28 (2 d, 2 H, Jgem = 15.1 Hz, ClCH2), 4.19 (dd, 1 H,
J9,9¢ = 12.4 Hz, J9,8 = 6.0 Hz, H-9 NeuTFA), 4.28 (m, 1 H, H-
8 NeuAc), 4.40 (dd, 1 H, J1,2 = 7.8 Hz, H-1 Glc), 4.45 (dd, 1
H, J1,2 = 8.2 Hz, H-1 Gal), 4.55 (dd, 1 H, J9¢,8 = 2.6 Hz, H-9¢
NeuTFA), 5.05 (ddd, 1 H, J4,5 = 10.4 Hz, H-4 NeuTFA),
5.27 (dd, 1H, J7,8 = 8.8 Hz, H-7 NeuTFA), 5.32 (ddd, 1 H,
J4,5 = 10.5 Hz, H-4 NeuAc), 5.52 (m, 1 H, H-8 NeuTFA),
6.00 (d, 1 H, JNH,5 = 8.7 Hz, NH NeuAc), 6.40 (d, 1 H,
JNH,5 = 9.4 Hz, NH NeuTFA).
J3eq,4 = 3.9 Hz, J3eq,3ax = 12.0 Hz, H-3eq), 1.74 (t, H-3ax),
3.61 (H-4), 3.82 (H-5), 3.71 (H-6), 3.86 (H-7), 4.11 (H-8),
3.75 (H-9), 4.17 (dd, J9¢,8 = 2.8 Hz, J9¢,9 = 12.1 Hz, H-9¢);
Gal: d = 4.52 (d, J1,2 = 7.6 Hz, H-1), 3.57 (dd, J2,3 = 9.7 Hz,
H-2), 4.08 (dd, J3,4 = 2.3 Hz, H-3), 3.96 (d, H-4), 3.72 (H-5),
3.73 (H-6), 3.75 (H-6¢); Glc: d = 4.54 (d, J1,2 = 8.2 Hz, H-1),
3.38 (t, J2,3 = 8.5 Hz, H-2), 3.66 (H-3), 3.68 (H-4), 3.62 (H-
5), 3.86 (H-6), 4.00 (dd, J6¢,6 = 12.4 Hz, J6¢,5 = 1.8 Hz, H-6¢),
3.27 (t, J = 4.9 Hz, OCH2CH2NH2), 3.88, 4.12
(OCH2CH2NH2). 13C NMR (125 MHz, D2O, relative to
internal acetone, dC = 31.45); terminal Neu5Ac: d = 176.2
(C-1), 41.7 (C-3), 69.7 (C-4), 53.0 (C-5), 74.0 (C-6), 69.4
(C-7), 72.2 (C-8), 63.9 (C-9), 23.3 (CH3CO), 174.4
(CH3CO); internal Neu5Ac: d = 176.2 (C-1), 41.0 (C-3),
69.0 (C-4), 53.5 (C-5), 75.1 (C-6), 70.5 (C-7), 79.3 (C-8),
62.7 (C-9), 23.5 (CH3CO), 174.4 (CH3CO); Gal: d = 104.0
(C-1), 70.5 (C-2), 76.7 (C-3), 68.7 (C-4), 76.5 (C-5), 62.3
(C-6); Glc: d = 103.2 (C-1), 73.9 (C-2), 75.3 (C-3), 79.2 (C-
4), 76.0 (C-5), 61.1 (C-6); 67.0 (OCH2CH2NH2), 40.7
(OCH2CH2NH2).
(31) Zou, W.; Borelli, S.; Gilbert, M.; Liu, T.; Pon, R. A.;
Jennings, H. J. J. Biol. Chem. 2004, 279, 25390.
Synlett 2005, No. 9, 1375–1380 © Thieme Stuttgart · New York