Synthesis of oligosaccharide structures from the lipopolysaccharide of Moraxella catarrhalis

Article abstract of DOI:10.1021/jo960789p

The synthesis of the octasaccharide [p-(trifluoroacetamido)phenyl]ethyl 4-O-[2-O-(2-acetamido-2-deoxy-α-D-glucopyranosyl)-β-D-glucopyranosyl] -6-O-[2-O-[4-O-(4-O-α-D-galactopyranosyl-β-D-galactopyranosyl)- α-D-glucopyranosyl]-α-D-glucopyranosyl]-3-O-β-D-glucopyranosyl- α-D-glucopyranoside, representing the outer part of the lipooligosaccharide from Moraxella catarrhalis serotype A, is described, together with a hepta-, a hexa-, and a pentasaccaride, composing parts thereof with shorter oligosaccharide chains substituted in the 6-position of the central 3,4,6-branched glucose moiety. The versatility of the use of thioglycosides in oligosaccharide synthesis is shown, since throughout the synthesis thioglycosides are used as glycosyl donor precursors, either directly in dimethyl(methylthio)sulfonium triflate (DMTST)-promoted coupling reactions or after conversion to the corresponding glycosyl bromide in silver triflate-promoted couplings. The effects of different protecting groups, anomeric leaving groups, and solvents used in the various coupling reactions are often substantial, which necessitates the use of easily convertible intermediates.

Full text of DOI:10.1021/jo960789p

J . Org. Chem. 1996, 61, 7711-7718
7711
Syn th esis of Oligosa cch a r id e Str u ctu r es fr om th e
Lip op olysa cch a r id e of Mor a xella ca ta r r h a lis
Kerstin Ekelo¨f and Stefan Oscarson*
Department of Organic Chemistry, Arrhenius Laboratory, Stockholm University,
S-106 91 Stockholm, Sweden
Received April 29, 1996 (Revised Manuscript Received August 27, 1996X)
The synthesis of the octasaccharide [p-(trifluoroacetamido)phenyl]ethyl 4-O-[2-O-(2-acetamido-2-
deoxy-R-D-glucopyranosyl)-â-D-glucopyranosyl]-6-O-[2-O-[4-O-(4-O-R-D-galactopyranosyl-â-D-galac-
topyranosyl)-R-D-glucopyranosyl]-â-D-glucopyranosyl]-3-O-â-D-glucopyranosyl-R-D-glucopyrano-
side, representing the outer part of the lipooligosaccharide from Moraxella catarrhalis serotype A,
is described, together with a hepta-, a hexa-, and a pentasaccaride, composing parts thereof with
shorter oligosaccharide chains substituted in the 6-position of the central 3,4,6-branched glucose
moiety. The versatility of the use of thioglycosides in oligosaccharide synthesis is shown, since
throughout the synthesis thioglycosides are used as glycosyl donor precursors, either directly in
dimethyl(methylthio)sulfonium triflate (DMTST)-promoted coupling reactions or after conversion
to the corresponding glycosyl bromide in silver triflate-promoted couplings. The effects of different
protecting groups, anomeric leaving groups, and solvents used in the various coupling reactions
are often substantial, which necessitates the use of easily convertible intermediates.
In tr od u ction
Resu lts a n d Discu ssion
In the retrosynthetic analysis of the target compounds,
care has to be taken of the order of introduction of the
different substituents on the branched central glucose
moiety, since in the synthesis of oligosaccharides there
is always the possibility to encounter problems with
unreactive glycosyl acceptors, especially when the struc-
ture is large and/or highly branched. The knowledge
about the conformational changes in the native struc-
tures further emphasized this possibility. The decision
was therefore taken to introduce the 3- and 4-substituent
first and then lastly the primary, hopefully more acces-
sible, 6-O-substituent. This scheme would also imply the
shortest way to derivatives with different length of the
6-O-chain by the introduction of various substituent
donors to a common intermediate. Benzyl groups were
chosen as permanent protection groups, whereas silyl
groups and, from the start, benzoyl groups were chosen
as temporary protecting groups. The known disaccharide
derivative 1,5 used in the earlier synthesis of the central
branched tetrasaccharide and already containing the 3-O-
substituent, was chosen as precursor.
Moraxella catarrhalis has been increasingly recognized
as a major pathogen in a number of respiratory diseases,
especially in children.1 The bacterium is divided into
three serogroups, A, B, and C. The structures of the cell-
surface lipopolysaccharide (LPS) from Moraxella ca-
tarrhalis strains of serotype A and C have recently been
determined.2-4 They lack the extended polymeric O-
antigenic side chains, and the structure without the lipid
A part of one of the strains of serotype A is shown in
Chart 1.
Some heterogeneity in the LPS within the serogroups
is known, e.g., different lengths of the oligosaccharide
chain in the primary position of the central glucose
moiety. In serogroup C elongation of the 4-O-substituent
is often found. To investigate the antigenicity of different
parts of the LPS structure synthesis was necessary. As
a first approach the tetrasaccharide and the penta-
saccharide shown in Chart 2 were synthesized,5 both
containing the central branched structure common for
all three serotypes of M. catarrhalis.
The benzoyl groups in the starting disaccharide de-
rivative 1 were changed to permanent benzyl groups (f2,
78%), whereafter the spacer arm, necessary for the
formation of glycoconjugates, was introduced via a di-
methyl(methylthio)sulfonium triflate6 (DMTST)-promot-
ed coupling in diethyl ether between the thioglycoside 2
and [p-(trifluoroacetamido)phenyl]ethanol to give the
R-linked O-glycoside 3 (61%) (Scheme 1). To allow
introduction of the 4-O-substituents and later the 6-O-
substituents, the benzylidene acetal was removed (87%)
and the resulting diol 4 was regioselectively silylated at
the primary hydroxyl group to give compound 5 (93%).
Two ways were considered for the entry of the 4-O-linked
disaccharide R-D-GlcNAc-(1f2)-â-D-Glc: a block synthe-
sis, in which a disaccharide donor is constructed first and
then coupled to acceptor 5, or a consecutive stepwise
synthesis. Although the R-coupling of the glucos-
In the structural elucidation, NMR experiments showed
anomalities in some of the NOE values, indicating a
conformational change in the oligosaccharide as the
substituents on the central glucose moiety grew longer.
To investigate when this change takes place and to
further evaluate the antigenicity of various structures
of the LPS, syntheses of larger structures have been
performed, including the complete hexose-containing
octasaccharide of serotype A and derivatives thereof with
shorter 6-O-substituents.
X Abstract published in Advance ACS Abstracts, October 1, 1996.
(1) Doern, G. V. Diagn. Microbiol. Infect. Dis. 1986, 4, 191.
(2) Masoud, H.; Perry, M. B.; Brisson, J .-R.; Uhr´ın, D.; Richards, J .
C. Can J . Chem. 1994, 72, 1466.
(3) Edebrink, P.; J ansson, P.-E.; Rahman, M. M.; Widmalm, G.;
Holme, T.; Rahman, M.; Weintraub, A. Carbohydr. Res. 1994, 257, 269.
(4) Edebrink, P.; J ansson, P.-E.; Mahbur, R.; Widmalm, G.; Holme,
T.; Rahman, M Carbohydr. Res. 1995, 266, 237.
(5) Ekelo¨f, K.; Oscarson, S. Carbohydr. Res. 1995, 278, 289.
(6) Fu¨gedi, P.; Garegg, P. J . Carbohydr. Res. 1986, 149, c9.
S0022-3263(96)00789-X CCC: $12.00 © 1996 American Chemical Society

7712 J . Org. Chem., Vol. 61, No. 22, 1996
Ekelof and Oscarson
Ch a r t 1
Ch a r t 2
Sch em e 1a
a
Key: (a) NaOMe, MeOH (89%); (b) BnBr, NaH, DMF (88%); (c) p-CF3CONHPhCH2CH2OH, DMTST, diethyl ether (61%); (d) HOAc
(aqueous), CH3CN (87%); (e) TBDMSCl, imidazole, DMF (93%).
acetamide moiety was expected to be the most difficult
and, thus, should be performed as early as possible in
the synthesis, the latter strategy was chosen, since this
would allow the use of a 2-O-participating group and
accordingly stereoselective â-coupling of the glucose
donor. As donor, the 2-O-benzoylated derivative 7,
obtained from the known derivative ethyl 2-O-acetyl-
3,4,6-tri-O-benzyl-1-thio-â-D-glucopyranoside7 through
deacetylation (f 6, 96%) and benzoylation (93%), was
chosen since benzoates are known to give less side
products in the coupling reaction.8 Using, once more,
DMTST as promoter, the trisaccharide 9 was cleanly
produced in 93% yield (Scheme 2). The debenzoylation
to the 2′′-OH derivative proved to be time-consuming, due
to the often noticed unreactivity of the 2-position of
glucose in oligosaccharides. Thus, treatment of deriva-
tive 9 with methanolic sodium methoxide required 14
days for complete deacylation, and during that time the
trifluoroacetamide group was also cleaved. Therefore,
subsequent N-trifluoroacetylation had to be performed
in two steps to finally give compound 10 (89%). As a way
to circumvent this delay the 2-O-chloroacetyl donor 8 was
synthesized and converted to the glycosyl bromide by
treatment with bromine and then coupled to acceptor 5
using silver triflate as promotor to give trisaccharide 11
(86%), which was found to rapidly (1 h) form 10 after
treatment with sodium methoxide. If the thioglycoside
8 was used directly as donor in a DMTST-promoted
coupling, only moderate yields of tetrasaccharide 11 were
obtained.
to be constructed, ethyl 2-azido-3,4,6-tri-O-benzyl-2-deoxy-
1-thio-â-D-glucopyranoside, with the nonparticipating
azido group as amino precursor in the 2-position, was
selected as donor. Different promoters (DMTST, NIS/
silver triflate) and different solvents (CH2Cl2, diethyl
ether) were tested, and when all these couplings failed,
different anomeric leaving group in the donor (bromide,
trichloroacetimidate) were also tested without success.
In all these experiments the reactivity of the donor was
shown to be poorly matched to that of the aglycon.9 The
high reactivity of the donor caused decomposition before
coupling could occur to the unreactive acceptor. There-
fore, the protection groups were changed from benzyl
groups to acetyl groups to decrease the reactivity of the
donor.10 This time, the bromo sugar donor (obtained from
the thioglycoside 12) promoted by silver triflate stereo-
selectively gave a tetrasaccharide product (Scheme 2),
which was shown to be the desired R-glycoside 13 by
NMR [δ 5.53 (d, J 1′′′,2′′′ ) 3.3 Hz, H-1′′′)]. The yield, 54%,
was acceptable, especially when taking into consideration
the stereoselectivity and the fact that 27% of the acceptor
could be recovered (74% coupling yield calculated on
consumed acceptor). In this coupling donor 12 could also
be used in a DMTST-promoted coupling with similar
results (see Experimental Section).
The silyl group was removed from compound 13 by acid
treatment to give derivative 14 (85%), with a free 6-OH
group. In the elongation at 6-OH it was again decided
to proceed via a stepwise procedure to be able to use a
stereoauxiliary group in position 2 of the donor to create
the â-linkage to the primary hydroxyl group in the
acceptor. The necessity of using acetyl groups as protect-
The introduction of the glucosamine motif caused, as
expected, problems to start with. Since an R-linkage was
(7) Garegg, P. J .; Ha¨llgren, C. J . Carbohydr. Chem. 1992, 11, 425.
(8) Garegg, P. J .; Konradsson, P.; Kvarnstro¨m, I.; Norberg, T.;
Svensson, S. C. T.; Wigilius, B. Acta Chem. Scand. 1985, B 37, 569.
(9) Paulsen, H. Angew. Chem., Int. Ed. Engl. 1982, 21, 155.
(10) Konradsson, P.; Udodong, U.; Fraser-Reid, B. Tetrahedron Lett.
1990, 31, 4313.

Oligosaccharide Structures from M. catarrhalis
J . Org. Chem., Vol. 61, No. 22, 1996 7713
Sch em e 2a
a
Key: (a) RCl, pyridine, CH2Cl2 (93 or 81%); (b) 7, DMTST, CH2Cl2 (94%) or 8, Br2, CH2Cl2 and then AgTf (86%); (c) (i) NaOMe,
MeOH, (ii) TFAA, pyridine, CH2Cl2, (iii) NaOMe, MeOH (89%); (d) NaOMe, MeOH; (e) Br2, CH2Cl2; (f) AgTf, CH2Cl2 (54%); (g) HOAc
(aqueous) (85%); (h) Br2, CH2Cl2; (i) AgTf, CH2Cl2 (79%).
Sch em e 3a
a
Key: (a) BnBr, NaH, DMF (90%); (b) PhCH(OMe)2, p-TsOH; (c) BnBr, NaH, DMF (78%); (d) NaCNBH3, HCl, THF (56%); (e) 2,3,4,6-
tetra-O-benzyl-R-D-galactopyranosyl chloride, AgTf, diethyl ether (62%).
ing groups in the azido donor precluded the use of
benzoates as temporary protecting groups, and therefore,
the monochloroacylated compound 8 was selected as
donor. In a DMTST-promoted coupling this donor once
more gave only moderate yields of coupling product, this
time together with the orthoester product (total 83%
yield). Various ways to avoid orthoester formation in
glycosidations with participating acetyl groups have been
described.11 Here, once more the thioglycoside was
transformed into the bromo sugar before coupling. Silver
triflate was used as promoter, and the â-linked penta-
saccharide product 15 was obtained in 79% yield (Scheme
2). Selective removal of the monochloroacetyl group in
the presence of the acetyl groups was accomplished with
hydrazine dithiocarbonate12 to give a key intermediate
16 (79%), with a free 2-hydroxyl group ready for elonga-
tion with various glycosyl donors. Pentasaccharide 15
was also completely deprotected through Zemple´n de-
acylation and catalytic hydrogenolysis to remove benzyl
groups and reduce the azido function. Selective N-
acetylation of the obtained amino group then afforded
the first target product 17 (Scheme 4).
The donors to be used in the subsequent couplings were
synthesized from the known methyl 1-thio-â-D-lacto-
pyranoside (18)13 through benzylation to give the lactose
donor 19 and via benzylidenation, benzylation, reductive
benzylidene opening, and a silver triflate-promoted cou-
pling with 2,3,4,6-tetra-O-benzyl-R-D-galactopyranosyl
chloride14 to yield the globotriose donor 22 (Scheme 3).
The R-configuration in the latter coupling was evident
from NMR [δ 5.19 (1H, d, J 1′′,2′′ ) 2.9 Hz, H-1′′)].
(11) Wilstermann, M.; Magnusson, G. Carbohydr. Res. 1995, 272,
1.
(13) Leontein, K.; Nilsson, M.; Norberg, T. Carbohydr. Res. 1985,
144, 231.
(12) van Boeckel, C. A. A.; Beetz, T. Tetrahedron Lett. 1983, 24,
3775.
(14) Iversen, T.; Bundle, D. R. Carbohydr. Res. 1982, 103, 29.

7714 J . Org. Chem., Vol. 61, No. 22, 1996
Ekelof and Oscarson
Sch em e 4a
a
Key: (a) (i) NaOMe, MeOH, (ii) H2, Pd-C, (iii) Ac2O; (b) (i) 2,3,4,6-tetra-O-benzyl-R-D-glucopyranosyl bromide or 19, Br2 or 22, Br2,
(ii) AgTf, diethyl ether.
As a model donor, which also would yield the hexa-
saccharide target compound, a 2,3,4,6-tetra-O-benzylated
glucose derivate was chosen. Once again, problems were
encountered with the low reactivity of the 2-position of
glucose, here in acceptor 16, and this time the stereo-
selectivity in the coupling was found to be a further
complication. Initially, the glucopyranosyl bromide15 was
tried as donor in a silver triflate-promoted coupling in
CH2Cl2, which gave a hexasaccharide product, but only
in 47% yield and and as an R/â-mixture (ratio 1/1).
Halide-assisted conditions were tried in an attempt to
improve the R/â-ratio but were found to be too mild to
yield any coupling product. The â-anomeric trichloro-
acetimidate donor16 and the ethyl thioglycoside donor15
analogs were also tested with various promoters, but with
even worse results than in the initial coupling using the
bromide as donor. A nonparticipating group is required
at the 2-position of the donor to enable the formation of
the desired R-linkage. Perhaps a modulation of the
reactivity of the donor by the introduction of acyl groups
at other positions would have improved the yields of
coupling product, but this would at the same time
considerably complicate the synthesis of the donors.
Fortunately, a simple solution to the problem was found,
as diethyl ether was tried as solvent in the initial silver
triflate-promoted coupling with the bromide as donors.
This solvent has previously been used to improve the
amount of R-glycosidically linked product formed, inter
alia, by us in DMTST-promoted couplings with thiogly-
coside donor.17 Here, it also seemed to have a stabilizing
effect on the activated donor, and a high yield (75%) of
the pure R-linked hexasaccharide 23 was obtained.
These coupling conditions were found to function as well
with the lactose and the globotriose donors. Couplings
with the bromo sugar obtained from 19 or 22 with
acceptor 16 gave stereoselectively the heptasaccaride 24
(69%) and the octasaccharide 25 (63%), respectively
(Scheme 4).
Deprotection of the coupling products 23, 24, and 25,
using the same protocol as for compound 15 above, gave
the final target compounds 26, 27, and 28, respectively
(Scheme 4).
This synthesis shows the versatility of thioglycoside
donors. They are easy to synthesize, are stable under
almost any protecting group manipulations, often func-
tion well as glycosyl donors using thiophilic promotors,
and if these couplings fail are easy to transform into most
other types of donors commonly used, even unstable
benzylated oligosaccharide glycosyl bromides as described
here.
Exp er im en ta l Section
Gen er a l Rem a r k s. Melting points are corrected. Organic
solutions were dried over MgSO4 before concentrating, which
was performed under reduced pressure at <40 °C (bath
temperature). NMR spectra were recorded at 25 °C at 270
MHz (1H) or 67.5 MHz (13C) in CDCl3 with Me4Si as internal
standard (δ ) 0 ppm) or in D2O with acetone as internal
standard (13C: δ ) 31.0 ppm; 1H: δ ) 2.225 ppm), unless
(15) Weygandt, F.; Ziemann, H. Liebigs Ann. Chem. 1962, 657, 179.
(16) Schmidt, R. R.; Michel, J .; Roos, M. Liebigs Ann. Chem. 1984,
1343.
(17) Garegg, P. J .; Oscarson, S.; Ritze´n, H.; Szo¨nyi, M. Carbohydr.
Res. 1992, 228, 121.

Oligosaccharide Structures from M. catarrhalis
J . Org. Chem., Vol. 61, No. 22, 1996 7715
otherwise stated. TLC was performed on Silica Gel F254 (E.
Merck) with detection by UV light and/or by charring with 8%
sulfuric acid. Silica gel (0.040-0.063 mm, Amicon) was used
for column chromatography.
mg, 1.9 mmol) were added to a solution of 4 (1.22 g, 1.2 mmol)
in DMF (10 mL). The solution was stirred for 1.5 h and then
diluted with toluene, washed with water, dried, and concen-
trated. Purification by silica gel chromatography (toluene-
EtOAc 15:1) gave 5 (1.26 g, 93%): [R]D +49° (c 1.1, CHCl3);
13C NMR (CDCl3) δ -5.2, 18.5, 26.0, 35.3, 62.9, 68.1, 68.6, 68.9,
72.3, 72.9, 73.5, 74.3, 74.5, 75.0, 75.7, 77.6, 79.1, 82.0, 82.9,
84.6, 96.4, 103.5, 120.6-138.6. Anal. Calcd for C63H74O12F3-
NSi: C, 67.42; H, 6.65; N, 1.25. Found: C, 67.18; H, 6.50; N,
1.39.
Eth yl 3,4,6-Tr i-O-ben zyl-1-th io-â-D-glu copyr an oside (6).
A solution of ethyl 2-O-acetyl-3,4,6-tri-O-benzyl-1-thio-â-D-
glucopyranoside7 (0.80 g, 1.5 mmol) in MeOH (25 mL) was
treated with a catalytic amount of 1 M sodium methoxide. The
solution was stirred at rt for 5 days and then neutralized with
Dowex-50 (H+) resin, filtered, concentrated, and purified by
silica gel chromatography (toluene-EtOAc 20:1) to give 6 (0.71
g, 96%): mp 78-79 °C (EtOAc-n-hexane); [R]D -12° (c 1.0,
CHCl3); 13C NMR (CDCl3) δ 15.4, 24.2, 69.0, 73.3, 73.4, 75.0,
75.2, 77.5, 86.0, 127.5-128.5. Anal. Calcd for C29H34O5S: C,
70.41; H, 6.93. Found: C, 70.39; H, 7.01.
Eth yl 2-O-Ben zyl-4,6-O-ben zylid en e-3-O-(2,3,4,6-tetr a -
O-b en zoyl-â-D-glu cop yr a n osyl)-1-t h io-â-D-glu cop yr a n o-
sid e (1). Silver triflate was added at -30 °C to a stirred
mixture of ethyl 2-O-benzyl-4,6-O-benzylidene-1-thio-â-D-glu-
copyranoside5 (200 mg, 0.50 mmol) and 2,3,4,6-tetra-O-benzoyl-
R-D-glucopyranosyl bromide18 (500 mg, 0.76 mmol) in CH2Cl2
(5 mL) containing molecular sieves (4 Å). The temperature
was allowed to rise to -5 °C during 1.5 h. Triethylamine (1
mL) was added, and stirring was continued for 20 min. The
mixture was diluted with CH2Cl2, filtered through Celite,
concentrated, and purified by silica gel chromatography (tolu-
ene-EtOAc 30:1) to give 1 (420 mg, 86%): [R]D +2.5° (c 1.0,
CHCl3); 13C NMR (CDCl3) δ 14.9, 24.9, 63.0, 68.6, 69.7, 70.3,
71.9, 72.3, 73.2, 75.4, 79.2, 80.9, 81.8, 85.5, 100.4, 101.3, 125.2-
137.7, 165.0, 165.1, 165.7, 165.9. Anal. Calcd for
C56H52O14S: C, 68.56; H, 5.34. Found: C,68.49; H, 5.45.
Eth yl 2-O-Ben zyl-4,6-O-ben zylid en e-3-O-(2,3,4,6-tetr a -
O-b en zyl-â-D-glu cop yr a n osyl)-1-t h io-â-D-glu cop yr a n o-
sid e (2). A solution of 1 (3.5 g, 3.6 mmol) in methanol (200
mL) was treated with a catalytic amount of 1 M sodium
methoxide. The solution was stirred overnight and then
neutralized with Dowex-50 (H+) resin, filtered, and concen-
trated. The residue was crystallized from MeOH, and the
mother liquor was purified by silica gel chromatography
(CHCl3-MeOH 30:1) to give more ethyl 2-O-benzyl-4,6-O-
benzylidene-3-O-(â-D-glucopyranosyl)-1-thio-â-D-glucopyrano-
side (total 1.8 g, 89%). To a suspension of sodium hydride
(60%, 411 mg, 10.3 mmol) in DMF (2 mL) was added a solution
of the above compound (966 mg, 1.71 mmol) in DMF (7 mL)
at rt, followed by a solution of benzyl bromide (1.1 mL, 9.58
mmol) in DMF (5 mL). After 3 h, MeOH (5 mL) was added,
and stirring was continued for 30 min. The mixture was
diluted with toluene, washed with water, dried (Na2SO4), and
concentrated. The residue was purified by silica gel chroma-
tography (light petroleum bp 40-60 °C-EtOAc 6:1) to give 2
(1.4 g, 88%): mp 155-156 °C (diethyl ether-n-hexane); [R]D
-1.9° (c 1.1, CHCl3); 13C NMR (CDCl3) δ 15.1, 25.1, 68.6, 68.9,
70.5, 73.5, 74.8, 74.9 75.0, 75.2, 75.5, 78.0, 79.2, 79.9, 81.8,
82.9, 84.8, 85.7, 101.1, 102.1, 126.1-138.5. Anal. Calcd for
C56H60O10S: C, 72.71; H, 6.54. Found: C, 72.93; H, 6.58.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-4,6-
O-b e n zylid e n e -3-O-(2,3,4,6-t e t r a -O-b e n zyl-â-D -glu co-
p yr a n osyl)-r-D-glu cop yr a n osid e (3). DMTST (2.18 g, 8.44
mmol) was added at rt to a stirred mixture of 2 (1.98 g, 2.14
mmol) and 2-[4-(trifluoroacetamido)phenyl]ethanol (0.59 g,
2.55 mmol) in dry diethyl ether (25 mL) containing molecular
sieves (4 Å). After 44 h triethylamine (2 mL) was added, and
stirring was continued for 20 min. The mixture was diluted
with CH2Cl2, filtered through Celite, concentrated, and purified
by silica gel chromatoraphy (toluene-EtOAc 10:1) to give 3
(1.44 g, 61%): 13C NMR (CDCl3) δ 35.4, 62.2, 68.7, 68.8, 69.0,
73.4, 74.7, 74.8, 75.5, 76.1 77.9, 80.2, 80.4, 82.8, 84.8, 97.3,
101.3, 102.5, 121.0-138.8.
Eth yl 2-O-Ben zoyl-3,4,6-tr i-O-ben zyl-1-th io-â-D-glu co-
p yr a n osid e (7). Benzoyl chloride (1.9 mL, 17 mmol) was
added to a solution of 6 (4.11 g, 8.31 mmol) in pyridine (50
mL). The mixture was stirred at rt for 19 h and then
concentrated. Purification of the residue by silica gel chro-
matography (toluene-EtOAc 20:1) gave 7 (4.65 g, 93%): mp
107-109 °C (EtOAc-n-hexane); [R]D +33° (c 0.9, CHCl3); 13C
NMR (CDCl3) δ 14.9, 23.8, 68.9, 72.4, 73.5, 75.1, 75.2, 77.9,
79.6, 83.5, 84.3, 127.6-138.2, 165.3. Anal. Calcd for
C36H39O6S: C, 72.20; H, 6.40. Found: C, 72.01; H, 6.50.
E t h yl 3,4,6-Tr i-O-b en zyl-2-O-ch lor oa cet yl-1-t h io-â-D-
glu cop yr a n osid e (8). Chloroacetyl chloride (0.74 mL, 9.30
mmol) was added to a solution of 6 (2.31 g, 4.67 mmol) in
CH2Cl2 (60 mL) and pyridine (4 mL). The mixture was stirred
for 1 h and then diluted with CH2Cl2 and washed with water.
The organic phase was dried and concentrated, and the residue
was purified by silica gel chromatography (toluene:EtOAc 20:
1) to give 8 (2.15 g, 81%): mp 72-73 °C (diethyl ether-light
petroleum bp 40-60 °C); [R]D +8.1° (c 1.2, CHCl3); 13C NMR
(CDCl3) δ 14.9, 23.7, 40.7, 68.7, 73.2, 73.5, 75.1, 75.3, 77.9,
79.5, 83.0, 84.0, 127.6-138.1, 166.1. Anal. Calcd for
C31H35O6SCl: C, 65.19; H, 6.18. Found: C, 65.24; H, 6.08.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a -O-ben zyl-â-D-glu cop yr a n osyl)-4-O-(3,4,6-tr i-
O-b e n zyl-2-O-b e n zoyl-â-D -glu cop yr a n osyl)-6-O-(t er t -
bu tyldim eth ylsilyl)-r-D-glu copyr an oside (9). DMTST (0.97
g, 3.74 mmol) was added to a mixture of 5 (1.05 g, 0.94 mmol)
and 7 (0.67 g, 1.12 mmol) in CH2Cl2 (7 mL) containing
molecular sieves (4 Å). After 18 h, triethylamine (1 mL) was
added, and the stirring was continued for 20 min. The mixture
was diluted with CH2Cl2, filtered through Celite, concentrated,
and purified by silica gel chromatography (toluene-EtOAc 15:
1) to give 9 (1.46 g, 94%): [R]D +36° (c 1.0, CHCl3); 13C NMR
(CDCl3) δ -5.2, -5.0, 18.4, 26.0, 35.2, 62.1, 67.9, 69.2 (2 C),
71.3, 72.4, 73.1, 73.3, 73.5, 74.4 (2 C), 74.6 (3 C), 74.7 (2 C),
75.0, 75.5, 78.1, 78.5, 82.3, 82.7, 82.8, 84.9, 95.9, 97.2, 102.2,
120.7-138.9, 154.4, 165.3. Anal. Calcd for C97H106O18NF3Si:
C, 70.22; H, 6.44; N, 0.84. Found: C, 69.67; H, 6.43; N,0.91.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a -O-ben zyl-â-D-glu cop yr a n osyl)-4-O-(3,4,6-tr i-
O-b en zyl-â-D-glu cop yr a n osyl)-6-O-(ter t-b u t yld im et h yl-
silyl)-r-D-glu cop yr a n osid e (10). A solution of 9 (100 mg,
0.06 mmol) in MeOH (5 mL) was treated with a catalytic
amount of 1 M sodium methoxide. The solution was stirred
for 14 days, neutralized with TFA, and concentrated. The
residue was dissolved in pyridine/CH2Cl2 (6:1, 21 mL), and
trifluoroacetic anhydride (24 µL, 0.17 mmol) was added. After
20 h, methanolic sodium methoxide was added until pH 9. The
solution was stirred for 10 min and then neutralized with
Dowex-50 (H+) resin, filtered, concentrated, and purified by
silica gel chromatography (toluene-EtOAc 20:1) to yield 10
(84 mg, 89%): [R]D +40° (c 1.0, CHCl3); 13C NMR (CDCl3) δ
-5.1, 18.4, 26.0, 35.2 , 63.8, 67.8, 68.6, 69.2, 70.3, 72.0, 73.2,
73.3 (2 C), 73.6, 74.9, 75.0, 75.1, 75.2, 75.3, 75.4, 75.5, 75.7,
76.7, 77.9, 81.7, 82.8, 84.8, 84.9, 95.4, 101.1, 102.1, 120.6-
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-t et r a -O-b en zyl-â-D-glu cop yr a n osyl)-r-D-glu co-
p yr a n osid e (4). Aqueous acetic acid (70%, 3 mL) was added
to a solution of 3 (485 mg, 0.44 mmol) in CH3CN (1 mL). The
solution was stirred at 70 °C for 4 h and then concentrated
and purified by silica gel chromatography (toluene-EtOAc 2:1)
to give 4 (390 mg, 87%): mp 145-146 °C (EtOAc-n-hexane);
[R]D +51° (c 0.9, CHCl3); 13C NMR (CDCl3) δ 35.3, 63.0, 68.5,
68.9, 70.2, 70.9, 73.0, 73.5, 74.3, 74.5, 75.0, 75.7, 77.7, 78.9,
82.0, 82.8, 84.6, 96.7, 103.5, 120.6-138.5, 154.3, 154.9. Anal.
Calcd for C57H60O12F3N: C, 67.91; H, 6.00; N 1.39. Found:
C, 67.86; H, 5.99; N, 1.41.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-t e t r a -O -b e n z y l-â-D -g lu c o p y r a n o s y l)-6-O -(t -
bu tyld im eth ylsilyl)-r-D-glu cop yr a n osid e (5). tert-Butyl-
dimethylsilyl chloride (237 mg, 1.6 mmol) and imidazole (132
(18) Ness, R. K.; Fletcher, J r., H. G.; Hudson, C. S. J . Am. Chem.
Soc. 1950, 72, 2200.

7716 J . Org. Chem., Vol. 61, No. 22, 1996
Ekelof and Oscarson
139.2. Anal. Calcd for C90H102017NF3Si: C,69.52; H, 6.61;
N,0.90. Found: C, 69.02; H, 6.44; N, 0.98.
1) to give 13 (534 mg, 61%). Starting material 10 (212 mg,
29%) could be recovered by further eluation (toluene:EtOAc
9:1).
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a -O-ben zyl-â-D-glu cop yr a n osyl)-4-O-[3,4,6-tr i-
O-ben zyl-2-O-(ch lor oacetyl)-â-D-glu copyr an osyl]-6-O-(ter t-
bu tyld im eth ylsilyl)-r-D-glu cop yr a n osid e (11). Bromine
(48 µL, 0.93 mmol) was added at rt to a solution of 8 (492 mg,
0.86 mmol) in CH2Cl2 (10 mL). After 10 min, toluene was
added, the solution was concentrated, and the residue was
coevaporated twice with toluene. The residue in CH2Cl2 (4
mL) was added to a solution of 5 (745 mg, 0.66 mmol) in
CH2Cl2 (4 mL) containing molecular sieves (4 Å). The mixture
was stirred under argon for 30 min at rt, and then the
temperature was lowered to -20 °C and silver triflate (290
mg, 1.1 mmol) was added. After 30 min triethylamine was
added and the stirring was continued for 20 min. The mixture
was diluted with CH2Cl2, filtered through Celite, and concen-
trated. The residue was purified by silica gel chromatography
(toluene:EtOAc 20:1) to give 11 (935 mg, 86%): 13C NMR
(CDCl3) δ -5.2, -5.1, 18.3, 26.0, 35.2, 40.7, 61.9, 68.0, 69.1,
69.2, 71.4, 73.0, 73.1, 73.5, 74.4, 74.7, 74.8, 74.9, 75.4, 75.7,
76.6, 77.1, 77.6, 78.1, 78.2, 82.0, 82.7, 84.9, 96.0, 97.6, 102.2,
120.9-138.9, 166.0.
Eth yl 3,4,6-tr i-O-a cetyl-2-a zid o-2-d eoxy-1-th io-â-D-glu -
cop yr a n osid e (12). Acetic anhydride (20 mL) was added to
a solution of ethyl 2-azido-2-deoxy-1-thio-â-D-glucopyranoside19
(1.1 g, 4.4 mmol) in pyridine (30 mL). After 1 h, the solution
was concentrated and the residue was purified by silica gel
chromatography (toluene:EtOAc 12:1) to yield 12 (1.81 g,
92%): mp 69-70 °C (diethyl ether-n-hexane); [R]D -40° (c
1.1, CHCl3); 13C NMR (CDCl3) δ 15.0, 20.6, 20.7, 25.1, 62.2,
63.6, 68.3, 74.5, 75.8, 84.5, 169.6, 169.9, 170.6. Anal. Calcd
for C14H21O7N3S: C,44.80; H, 5.64; N, 11.19. Found: C, 44.75;
H, 5.51; N, 11.11.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
t r i-O-b en zyl-â-D-glu cop yr a n osyl]-r-D-glu cop yr a n osid e
(14). 13 (620 mg, 0.33 mmol) was dissolved in CH3CN (4 mL),
HOAc (80%, aqueous, 8 mL) was added, and the solution was
stirred at 40 °C for 18 h, concentrated, and then purified by
silica gel chromatography (toluene:EtOAc 4:1) to give 14 (492
mg, 85%): [R]D +73 ° (c 1.2, CHCl3); 13C NMR (CDCl3) δ 20.5,
20.7, 20.8, 35.3 , 60.8, 61.1, 61.9, 67.0, 67.9, 68.6, 69.1, 69.4,
70.2, 71.3, 72.5, 72.8, 73.3 (2 C), 73.6 (2C), 74.3, 74.8, 74.9,
75.1 (2 C), 75.6, 76.4, 78.3, 78.5, 82.3, 82.8, 83.0, 84.9, 96.3,
96.4, 99.0, 102.2, 120.5-139.1, 154.3, 154.9, 169.6, 170.0, 170.5.
Anal. Calcd for C96H103O24N4F3: C,65.74; H, 5.92; N,3.19.
Found: C, 65.57; H, 5.90; N,3.23.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-(2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
tr i-O-ben zyl-â-D-glu cop yr a n osyl)-6-O-(3,4,6-tr i-O-ben zyl-
2-O -(c h lo r o a c e t y l)-â-D -g lu c o p y r a n o s y l)-r-D -g lu c o -
p yr a n osid e (15). Bromine (12 µL, 0.24 mmol) was added at
0 °C to a solution of 8 (120 mg, 0.21 mmol) in CH2Cl2 (4 mL).
After 10 min, toluene was added, the solution was concen-
trated, and the residue was coevaporated twice with toluene.
The residue in CH2Cl2 (5mL) was added to a solution of 14
(244 mg, 0.14 mmol) in CH2Cl2 (5 mL) containing molecular
sieves (4 Å). The mixture was stirred under argon for 30 min
at rt, and then the temperature was lowered to -35 °C and
silver triflate (71 mg, 0.28 mmol) was added. After 30 min,
triethylamine was added, and the stirring was continued for
20 min. The mixture was diluted with CH2Cl2, filtered through
Celite, and concentrated. Purification of the residue by silica
gel chromatography (toluene:EtOAc 8:1) gave 15 (250 mg,
79%): [R]D +66 ° (c 1.0 , CHCl3); 13C NMR (CDCl3) δ 20.5, 20.6,
20.8, 35.2, 40.8, 60.5, 61.2, 67.2, 67.9, 68.3, 68.5, 68.9, 69.1,
69.6, 70.0, 71.8, 72.3, 73.5, 73.9, 74.3, 74.6, 74.8, 74.9, 75.2,
75.3, 75.6, 76.0, 78.1, 78.4, 82.2, 82.7, 83.0 83.1, 85.0, 96.1 (2C),
97.7, 101.6, 102.5, 120.6-138.8, 154.3, 154.9, 165.9, 169.5,
169.9, 170.6. Anal. Calcd for C125H132O30N4F3Cl: C, 66.35;
H, 5.88; N, 2.48. Found: C, 66.27; H, 5.88; N, 2.46.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
tr i-O-ben zyl-â-D-glu cop yr a n osyl]-6-O-(3,4,6-tr i-O-ben zyl-
â-D-glu cop yr a n osyl)-r-D-glu cop yr a n osid e (16). Hydrazine
dithiocarbonate12 (100 mg, 1.3 mmol) in DMF (10 mL) was
added to a solution of 15 (600 mg, 0.27 mmol) in DMF (10 mL).
After 30 min, the solution was diluted with CH2Cl2 and washed
with 1 M H2SO4, NaHCO3, and water. The organic phase was
dried and concentrated, and the residue was purified by silica
gel chromatography (toluene:EtOAc 6:1) to give 16 (459 mg,
79%): [R]D +56° (c 1.0, CHCl3); 13C NMR (CDCl3) δ 21.5, 21.7,
21.8, 36.3, 61.9, 62.1, 68.1, 69.0, 69.5, 69.9, 70.0, 70.4, 71.1,
71.3, 73.3, 73.8, 74.4, 74.6, 75.2, 75.5, 75.8, 76.0, 76.1, 76.7,
79.4, 79.9, 83.0, 83.8, 84.1, 85.5, 85.9, 97.3, 97.6, 99.8, 103.2,
104.5, 121.6-140.0, 170.5, 171.0, 171,5. Anal. Calcd for
C123H131O29N4F3: C, 67.57; H, 6.04; N, 2.56. Found: C, 67.55;
H, 6.03; N, 2.52.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
t r i -O -b e n z y l-â-D -g lu c o p y r a n o s y l]-6-O -(t er t -b u t y l-
d im eth ylsilyl)-r-D-glu cop yr a n osid e (13). Bromine (34 µL,
0.66 mmol) was added to a solution of 12 (113 mg, 0.30 mmol)
in CH2Cl2 (4 mL). After 30 min, toluene was added, the
mixture was concentrated, and the residue was coevaporated
twice with toluene. The residue in CH2Cl2 (5 mL) was added
to a solution of 10 (236 mg, 0.15 mmol) in CH2Cl2 (5 mL)
containing molecular sieves (4 Å). The mixture was stirred
under argon for 30 min at rt, after which time the temperature
was lowered to -30 °C and silver triflate (97 mg, 0.38 mmol)
was added. After 30 min, triethylamine was added, and the
stirring was continued for 20 min. The mixture was diluted
with CH2Cl2, filtered through Celite, and concentrated. The
residue was purified by silica gel chromatography (toluene:
EtOAc 12:1) to give 13 (152 mg, 54%). Starting material 10
(64 mg, 27%) was recovered by further elution (toluene:EtOAc
9:1). 13: [R]D +67° (c 0.6, CHCl3); NMR (CDCl3) 13C, δ -5.3,
-5.0, 18.4, 20.5, 20.7, 20.8, 26.0, 35.3, 60.9, 61.2, 62.1, 66.9,
68.0, 68.4, 69.1, 69.5, 70.2, 71.7, 72.7 (2 C), 72.9, 73.1, 73.3,
73.6, 74.4, 74.9 (2 C), 75.1 (3 C), 75.5, 76.3, 78.4, 78.6, 82.4,
83.0 (2 C), 84.9, 96.0 (J C,H ) 164.0 Hz), 96.1 (J C,H ) 166.1 Hz),
99.0 (J C,H ) 166.1 Hz), 102.1 (J C,H ) 170.6 Hz), 120.5-139.1,
1
154.3, 154.9, 169.6, 170.0, 170.5; H, δ 3.00 (dd, J 1′′′,2′′′ ) 3.3
[p-(Tr iflu or oa ceta m id o)p h en yleth yl 4-O-[2-O-(2-Acet-
a m id o-2-d eoxy-r-D-glu cop yr a n osyl)-â-D-glu cop yr a n osyl]-
3-O-â-D-glu cop yr a n osyl-6-O-â-D-glu cop yr a n osyl-r-D-glu -
cop yr a n osid e (17). A solution of 15 (250 mg, 0.11 mmol) in
CH2Cl2 (2 mL) and MeOH (10 mL) was treated with a catalytic
amount of 1 M sodium methoxide. After 6 h, the mixture was
neutralized with Dowex 50 (H+) resin, filtered, concentrated,
and purified by silica gel chromatography (toluene:EtOAc 2:1).
The residue was dissolved in EtOAc/MeOH/HOAc (2:2:1, 15
mL) and hydrogenolyzed over 10% Pd/C (50 mg) at 120 psi for
20 h. The mixture was filtered and concentrated and the
residue treated with acetic anhydride (38 µL, 0.41 mmol) in
MeOH (2 mL). After 2 h the solution was concentrated,
dissolved in water, and washed with diethyl ether. The water
phase was concentrated and the residue desalted on a BioGel
Hz, J 2′′′,3′′′ ) 10.6 Hz 1H, H-2′′′), 5.53 (d, J 1′′′,2′′′ ) 3.3 Hz, 1H,
H-1′′′). Anal. Calcd for C102H117O24N4F3Si: C, 65.58; H, 6.31;
N, 3.05. Found: C, 66.92; H, 6.23; N, 3.23.
To a stirred mixture of 10 (0.73 g, 0.47mmol) and 12 (0.26
g, 0.70 mmol) in diethyl ether (30 mL) containing molecular
sieves (4 Å) were added NIS (0.18 g, 0.79 mmol) and silver
tfiflate (24 mg, 0.09 mmol). After 3 h, triethylamine was
added, and the mixture was diluted with CH2Cl2, filtered
through Celite, and washed with Na2S2O3, NaHCO3, and
water. The organic phase was dried, filtered, concentrated,
and purified by silica gel chromatography (toluene:EtOAc 12:
(19) Buskas, T.; Garegg, P. J .; Konradsson, P.; Maloisel, J .-L.
Tetrahedron: Assymetry, 1994, 5, 2187.

Oligosaccharide Structures from M. catarrhalis
J . Org. Chem., Vol. 61, No. 22, 1996 7717
P-2 column to give after lyophilization 17 (64 mg, 53%): [R]D
+67° (c 1.0, H2O); NMR (D2O) 13C, δ 23.1, 35.3, 54.4, 54.6, 61.4,
67.3, 69.1, 70.1, 70.2, 70.3, 70.5, 70.7, 71.7, 72.2, 72.4, 72.5,
72.6, 73.6, 74.0, 75.2, 76.0, 76.2, 76.5, 76.7, 76.8, 77.7, 96.6,
98.7, 101.2, 102.7, 102.9, 122.9, 130.5, 133.9, 138.3, 174.5; 1H,
δ 2.08 (3H, s), 2.96-4.13 (32H, m), 4.44 (1H, d, J 1,2 ) 8.1 Hz),
4.75 (1H, d, J 1,2 ) 8 Hz), 4.80 (1H, d, J 1,2 ) 8 Hz), 4.93 (1H, d,
J 1,2 ) 3.7 Hz), 5.22 (1H, d, J 1,2 ) 3.3 Hz), 7.37 (2H, d, J ) 8.1
Hz), 7.51 (2H, d, J ) 8.1 Hz); HRMS calcd for C42H63O27N2F3
[M - H]+ 1083.3492, found 1083.3496.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
tr i-O-ben zyl-â-D-glu cop yr a n osyl]-6-O-[3,4,6-tr i-O-ben zyl-
2-O -(2,3,4,6-t e t r a -O -b e n zy l-r-D -g lu c o p y r a n o s y l)-â-D -
glu cop yr a n osyl]-r-D-glu cop yr a n osid e (23). Bromine (17
µL, 0.34 mmol) was added at 0 °C to a solution of ethyl 2,3,4,6-
tetra-O-benzyl-1-thio-â-D-glucopyranoside15 (188 mg, 0.31 mmol)
in CH2Cl2 (10 mL). After 10 min, toluene was added, the
solution was concentrated, and the residue was coevaporated
twice with toluene. The residue in diethyl ether (5 mL) was
added to a solution of 16 (226 mg, 0.10 mmol) in diethyl ether
(5 mL) containing molecular sieves (4 Å). The mixture was
stirred under argon for 30 min at rt, whereafter the temper-
ature was lowered to -35 °C and silver triflate (106 mg, 0.41
mmol) was added. After 30 min, triethylamine was added,
and the stirring was continued for 20 min. The mixture was
diluted with CH2Cl2, filtered through Celite, and concentrated.
Purification of the residue by silica gel chromatography
(toluene:EtOAc 10:1) gave 23 (210 mg, 75%): [R]D +76° (c 1.0,
CHCl3); 13C NMR (CDCl3) δ 20.5, 20.6, 20.8, 34.9, 60.7, 60.9,
67.0, 67.9, 68.6, 68.7, 69.1, 69.6, 69.9, 70.2, 71.3, 71.8, 72.3,
73.0, 73.1, 73.6, 73.7, 74.4, 74.6, 74.9, 75.2, 75.6, 76.1, 77.0,
77.8, 78.0, 78.7, 80.3, 81.8, 82.6, 82.8, 82.9, 83.3, 84.9, 94.4,
95.3, 96.6, 97.7, 102.2, 103.5, 120.7-139.0, 169.5, 170.0, 170.4.
Anal. Calcd for C157H165O34N4F3: C, 69.61; H, 6.14; N, 2.07.
Found: C, 69.50; H, 6.01; N, 2.07.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
tr i-O-ben zyl-â-D-glu cop yr a n osyl]-6-O-[3,4,6-tr i-O-ben zyl-
2-O-[4-O-(2,3,4,6-t et r a -O-b en zyl-â-D-ga la ct op yr a n osyl)-
2,3,6-t r i -O -b e n z y l-r-D -g lu c o p y r a n o s y l]-â-D -g lu c o -
p yr a n osyl]-r-D-glu cop yr a n osid e (24). The bromosugar of
19 (215 mg, 0.21 mmol) was prepared as described and coupled
to 16 (241 mg, 0.11 mmol) as described for compound 23 above
to give, after silica gel chromatography twice (toluene-EtOAc
10:1 then light petroleum bp 40-60 °C-EtOAc 5:2), 24 (240
mg, 69%): [R]D +58° (c 1.1, CHCl3); 13C NMR (CDCl3) δ 20.5,
20.6, 20.7, 35.1, 60.3, 60.7, 60.8, 67.0, 67.9, 68.6, 69.0, 69.3,
69.9, 70.3, 71.2, 71.8, 72.3, 72.7, 72.8, 73.2, 73.5, 73.6, 73.8,
74.1, 74.5, 74.7, 75.1, 75.3, 75.5, 76.8, 78.0, 78.5, 78.6, 79.1,
79.8, 80.1, 82.5, 82.9, 84.9, 94.0, 95.6, 96.5, 97.7, 102.1, 103.0,
103.6, 121.2-139.4, 154.2, 154.8, 169.4, 169.9, 170.3. Anal.
Calcd for C184H193O39N4F3: C, 70.35; H, 6.19; N, 1.78. Found:
C, 70.33; H, 6.21; N, 1.76.
Meth yl 2,3,6-Tr i-O-ben zyl-4-O-(2,3,4,6-tetr a -O-ben zyl-
â-D-ga la ctop yr a n osyl)-1-th io-â-D-glu cop yr a n osid e (19). A
solution of methyl 1-thio-â-D-lactoside13 (18, 1.15 g, 3.09 mmol)
in DMF (10 mL) was added to a suspension of sodium hydride
(60%, 1.3 g, 32.5 mmol) in DMF (5 mL) followed by a solution
of benzyl bromide (3.6 mL, 30.3 mmol) in DMF (10 mL). After
2 h, MeOH (10 mL) was added, and stirring was continued
for 30 min. The mixture was diluted with toluene, washed
with water, dried, and concentrated. Purification of the
residue by silica gel chromatography (light petroleum bp 40-
60 °C-EtOAc 5:1) gave 19 (2.80 g, 90%): mp 82-84 °C (diethyl
ether-n-hexane); [R]D +6.5° (c 1.1, CHCl3); 13C NMR (CDCl3)
δ 12.6, 68.0, 68.3, 72.6, 73.0, 73.4, 73.6, 74.7, 75.3, 75.5, 76.4,
79.5, 80.4, 82.5, 84.8, 85.2, 102.7, 127.1-139.9. Anal. Calcd
for C62H66O10S: C,74.23; H, 6.63. Found: C, 74.10; H, 6.70.
Meth yl 2,3,6-Tr i-O-ben zyl-4-O-(4,6-O-ben zylid en e-â-D-
ga la ctop yr a n osyl)-1-th io-â-D-glu cop yr a n osid e (20). A so-
lution of methyl 4-O-(4,6-O-benzylidene-â-D-galactopyranosyl)-
1-thio-â-D-glucopyranoside13 (1.18 g, 2.56 mmol) in DMF (20
mL) was added to a suspension of sodium hydride (60%, 0.77
g, 19.2 mmol) in DMF (3 mL) followed by a solution of benzyl
bromide (2.1 mL, 17.9 mmol) in DMF (5 mL). After 2 h, MeOH
(5 mL) was added, and stirring was continued for 30 min. The
mixture was diluted with toluene, washed with water, dried,
and concentrated. Purification of the residue by silica gel
chromatography (light petroleum bp 40-60 °C:EtOAc 4:1) gave
20 (1.83 g, 78%): mp 124-126 °C (EtOAc-n-hexane); [R]D
+7.2° (c 1.0, CHCl3); 13C NMR (CDCl3) δ 12.7, 66.4, 68.3, 68.9,
71.6, 72.9, 73.7, 75.4, 75.5, 76.0, 77.2, 78.8, 79.4, 79.6, 80.5,
84.9, 85.2, 101.4, 102.7, 126.6-138.8. Anal. Calcd for
C55H58O10S: C,72.50; H, 6.42. Found: C, 72.43; H, 6.54.
Met h yl 2,3,6-Tr i-O-b en zyl-4-O-(2,3,6-t r i-O-b en zyl-â-D-
ga la ctop yr a n osyl)-1-th io-â-D-glu cop yr a n osid e (21). So-
dium cyanoborohydride (0.91, 14.6 mmol) was added at 0 °C
to a stirred mixture of 20 (1.31g, 1.40 mmol) and molecular
sieves (4 Å) in THF (20 mL), followed by HCl in diethyl ether
(saturated) until gas evolution ceased. After 20 min, CH2Cl2
(4 mL) and water (2 mL) were added, and the mixture was
filtered through Celite. The organic layer was washed with
water, NaHCO3, and water, dried, and concentrated. Purifica-
tion of the residue by silica gel chromatography (toluene:EtOAc
15:1) gave 21 (0.74 g, 56%): mp 65-67 °C (diethyl ether-n-
hexane); [R]D +18° (c 1.0, CHCl3); NMR (CDCl3) 13C δ 12.6,
66.1, 68.2, 68.4, 72.0, 72.8, 73.0, 73.5, 75.2, 75.4, 75.5, 76.2,
79.4 (2 C), 79.6, 80.4, 84.7, 85.1, 102.4, 127.3-138.9 Anal.
Calcd for C55H60O10S: C, 72.34; H, 6.62. Found: C, 72.24; H,
6.67.
Meth yl 2,3,6-Tr i-O-ben zyl-4-O-[2,3,6-tr i-O-ben zyl-4-O-
(2,3,4,6-tetr a -O-ben zyl-r-D-ga la ctop yr a n osyl)-â-D-ga la c-
top yr a n osyl]-1-th io-â-D-glu cop yr a n osid e (22). Silver tri-
flate (159 mg, 0.62 mmol) was added at -60 °C to a stirred
mixture of 2,3,4,6-tetra-O-benzyl-R-D-galactopyranosyl chlo-
ride14 (330 mg, 0.59 mmol) and 20 (270 mg, 0.30 mmol) in
diethyl ether (15 mL) containing molecular sieves (4 Å). After
1 h at -60 °C, the reaction was left to attain -10 °C during 3
h, and then triethylamine (1 mL) was added and the stirring
was continued for 20 min. The mixture was diluted with
CH2Cl2, filtered through Celite, and concentrated. Purification
of the residue by silica gel chromatography twice (toluene-
EtOAc 25:1 and then light petroleum bp 40-60 °C-EtOAc 5:1)
gave 2 (262 mg, 62%): [R]D+32° (c 1.2, CHCl3); NMR (CDCl3)
13C, δ 12.6, 67.7, 67.8, 68.3, 69.5, 72.1, 72.4, 73.0, 73.2, 73.3,
73.7, 74.8, 75.1, 75.3, 75.4, 76.6, 76.8, 79.4, 80.3, 81.6, 84.5,
85.2, 100.7, 102.7, 127.1-139.0; 1H, δ 5.19 (1H, d, J 1′′,2′′ ) 2.9
Hz, H-1′′), 5.27 (1H, d, J 1′,2′ ) 10.6 Hz, H-1′). Anal. Calcd for
C89H94O15S: C, 74.45; H, 6.60. Found: C, 73.92; H, 6.51.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 2-O-Ben zyl-3-O-
(2,3,4,6-tetr a-O-ben zyl-â-D-glu copyr an osyl)-4-O-[2-O-(3,4,6-
t r i-O-a cet yl-2-a zid o-2-d eoxy-r-D-glu cop yr a n osyl)-3,4,6-
tr i-O-ben zyl-â-
D-glu cop yr a n osyl]-6-O-[3,4,6-tr i-O-ben zyl-
2-O -[4-O -[4-O -(2,3,4,6-t e t r a -O -b e n z y l-r-
D -g a la c t o -
p yr a n osyl)-2,3,6-tr i-O-ben zyl-â-D-ga la ctop yr a n osyl]-2,3,6-
tr i-O-ben zyl-r-D-glu cop yr a n osyl]-â-D-glu cop yr a n osyl]-r-
D-glu cop yr a n osid e (24). The bromosugar of 22 (346 mg, 0.24
mmol) was prepared and coupled to 16 (275 mg, 0.13 mmol)
as described for compound 23 above to give, after silica gel
chromatography twice (toluene-EtOAc 10:1 then light petro-
leum bp 40-60°C-EtOAc 3:1), 25 (282 mg, 63%): [R]D +64°
(c 0.8, CHCl3); 13C NMR (CDCl3) δ 20.5, 20.6, 20.8, 35.0, 60.7,
60.9, 67.0, 67.7, 67.9, 68.4, 68.5, 68.7, 69.0, 69.3, 69.4, 69.9,
70.1, 70.3, 71.2, 71.9, 72.2, 72.9, 73.1, 73.5, 73.6, 73.8, 74.1,
74.4, 74.6, 74.7, 74.9, 75.2, 75.3, 75.6, 76.0, 76.9, 77.3, 78.0,
78.4, 78.6, 79.5, 79.7, 80.1, 81.5, 82.5, 82.9, 83.0, 84.9, 94.0,
95.6, 96.5, 97.6, 100.9, 102.1, 103.1, 103.7, 121.2-139.5, 154.2,
154.7, 169.5, 169.9, 170.4. Anal. Calcd for C211H221O44N4F3:
C, 70.91; H, 6.23; N, 1.57. Found: C, 70.98; H, 6.27; N, 1.44.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 4-O-[2-O-(2-Acet-
a m id o-2-d eoxy-r-D-glu cop yr a n osyl)-â-D-glu cop yr a n osyl]-
3-O-â-D-glu cop yr a n osyl-6-O-[2-O-(r-D-glu cop yr a n osyl)-â-
D-glu cop yr a n osyl]-r-D-glu cop yr a n osid e (26). 23 (190 mg,
0.07 mmol) was deprotected and desalted as described for
compound 17 above to give 26 (46 mg, 53%): [R]D +73 ° (c 0.6,
H2O); NMR (D2O) 13C, δ 23.4, 35.4, 54.4, 61.0, 61.5, 61.7, 69.2,
69.7, 70.3, 70.5, 70.9, 72.3, 72.5, 72.7, 73.0, 73.5, 73.8, 74.4,
75.2, 75.4, 76.5, 76.6, 76.8, 80.2 97.4, 98.9 (3 C), 103.1, 103.7,
122.9, 130.5, 134.1, 137.9, 157.3, 157.9, 174.9; 1H, δ 4.57 (1H,

7718 J . Org. Chem., Vol. 61, No. 22, 1996
Ekelof and Oscarson
d, J 1,2 ) 7.7 Hz), 4.92 (1H, d, J 1,2 ) 3.7 Hz), 4.93 (1H, d, J 1,2
)
6-O-[2-O-[4-O-(4-O-r-D -ga la ct op yr a n osyl-â-D -ga la ct o-
p yr a n osyl)-r-D-glu cop yr a n osyl]-3-O-â-D-glu cop yr a n osyl-
â-D-glu cop yr a n osyl]-r-D-glu cop yr a n osid e (28). 25 (272
mg, 0.08 mmol) was deprotected and desalted as described for
17 to give 28 (43 mg, 36%): [R]D +84° (c 1.0, H2O); NMR (D2O)
13C, δ 23.4, 35.4 , 54.4, 60.9, 61.1, 61.3, 61.5, 61.7, 69.3, 69.7,
69.9, 70.3, 70.5, 70.9, 71.2, 71.5, 71.6, 72.1, 72.2, 72.7, 72.9,
73.8, 74.4, 75.1, 75.3, 76.2, 76.4, 76.5, 76.6, 76.8,78.1, 79.8, 80.1
97.1, 98.9, 101.0, 103.2, 103.6, 104.0, 122.9, 130.5, 134.0, 137.9,
7.7 Hz), 4.99 (1H, d, J 1,2 ) 7.3 Hz), 5.12 (1H, d, J 1,2 ) 3.3 Hz),
5.34 (1H, d, J 1,2 ) 3.7 Hz); HRMS calcd for C48H73O32N2F3 [M
- H]+ 1245.4021, found 1245.4003.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 4-O-[2-O-(2-Acet-
a m id o-2-d eoxy-r-D-glu cop yr a n osyl)-â-D-glu cop yr a n osyl]-
6-O-[2-O-(4-O-â-D-ga la ctop yr a n osyl-r-D-glu cop yr a n osyl)-
â-D-glu cop yr a n osyl]-3-O-â-D-glu cop yr a n osyl-r-D-glu co-
p yr a n osid e (27). 24 (210 mg, 0.07 mmol) was deprotected
and desalted as described for compound 17 above to yield 27
(34 mg, 36%): [R]D +75 ° (c 0.9, H2O); NMR (D2O)13C, δ 23.4,
35.4 , 54.4, 60.9, 61.5, 61.7, 61.8, 69.3, 69.6, 70.3, 70.5, 70.9,
71.1, 72.0, 72.2, 72.7, 73.0, 73.3, 73.7, 74.4, 75.1, 75.3, 76.1,
76.5, 76.8, 79.5, 80.1, 97.1, 98.9 (3 C), 103.1, 103.6, 103.8, 122.9,
1
174.9; H, δ 4.45 (1H, d, J 1,2 ) 7.7 Hz), 4.58 (1H, d, J 1,2 ) 7.7
Hz), 4.94 (3H, m), 4.98 (1H, d, J 1,2 ) 7.7 Hz), 5.12 (1H, d, J 1,2
) 3.3 Hz), 5.34 (1H, d, J 1,2 ) 3.7 Hz); FABMS calcd for
C
60H93O42N2F3 [M + H]+ 1570.5, found 1571.1.
1
130.5, 134.0, 137.9, 157.2, 174.9; H, δ 4.38 (1H, d, J 1,2 ) 7.7
Ack n ow led gm en t. We thank Professor Per J . Gar-
Hz), 4.58 (1H, d, J 1,2 ) 7.7 Hz), 4.92 (1H, d, J 1,2 ) 7.3 Hz),
4.93 (1H, d, J 1,2 ) 3.7 Hz), 4.98 (1H, d, J 1,2 ) 7.3 Hz), 5.12
(1H, d, J 1,2 ) 3.3 Hz), 5.34 (1H, d, J 1,2 ) 4.0 Hz); HRMS calcd
for C54H83O37N2F3 [M - H]+ 1407.4549, found 1407.4548.
[p-(Tr iflu or oa ceta m id o)p h en yl]eth yl 4-O-[2-O-(2-Acet-
a m id o-2-d eoxy-r-D-glu cop yr a n osyl)-â-D-glu cop yr a n osyl]-
egg for his interest in this work and the Swedish
Research Council for Engineering Studies and Lederle-
Praxis Biologicals for financial support.
J O960789P