Variant synthetic pathway to glucuronic acid-containing di- and trisaccharide thioglycoside building blocks for continued synthesis of Cryptococcus neoformans capsular polysaccharide structures

Article abstract of DOI:10.1016/j.carres.2007.11.026

An alternative pathway to glucuronic acid-containing di- and trisaccharide thioglycoside building blocks, suitable for the synthesis of Cryptococcus neoformans capsular polysaccharide structures, has been developed. As opposed to our earlier synthesis, this approach features the introduction of the glucuronic acid motif at the di- and trisaccharide level through oxidation of a glucose residue. This approach circumvents problems encountered in glycosylations with glucuronic acid donors and benzylation of glucuronic acid-containing derivatives. Selective protection of primary alcohols was obtained at the di- and trisaccharide stage using TBDMS or trityl protecting groups, respectively. After benzylation of the secondary hydroxyl groups and subsequent removal of the TBDMS or trityl group, oxidation of the free primary alcohols to carboxylic acids was performed in high yield using the TEMPO-BAIB reagent mixture, which does not tend to oxidize thioglycosides. The new approach requires a number of extra steps, but has proven to be more reliable and easily reproducible.

Full text of DOI:10.1016/j.carres.2007.11.026

Available online at www.sciencedirect.com
Carbohydrate Research 343 (2008) 2200–2208
Note
Variant synthetic pathway to glucuronic acid-containing
di- and trisaccharide thioglycoside building blocks for
continued synthesis of Cryptococcus neoformans
capsular polysaccharide structures
Jan Vesely, Lina Rydner and Stefan Oscarson*
Department of Organic Chemistry, Arrhenius laboratory, Stockholm University, S-106 91 Stockholm, Sweden
Received 4 October 2007; received in revised form 12 November 2007; accepted 25 November 2007
Available online 4 December 2007
Presented at Eurocarb 14th Lubeck, Germany, September 2007
¨
Abstract—An alternative pathway to glucuronic acid-containing di- and trisaccharide thioglycoside building blocks, suitable for the
synthesis of Cryptococcus neoformans capsular polysaccharide structures, has been developed. As opposed to our earlier synthesis,
this approach features the introduction of the glucuronic acid motif at the di- and trisaccharide level through oxidation of a glucose
residue. This approach circumvents problems encountered in glycosylations with glucuronic acid donors and benzylation of gluc-
uronic acid-containing derivatives. Selective protection of primary alcohols was obtained at the di- and trisaccharide stage using
TBDMS or trityl protecting groups, respectively. After benzylation of the secondary hydroxyl groups and subsequent removal of
the TBDMS or trityl group, oxidation of the free primary alcohols to carboxylic acids was performed in high yield using the
TEMPO–BAIB reagent mixture, which does not tend to oxidize thioglycosides. The new approach requires a number of extra steps,
but has proven to be more reliable and easily reproducible.
Ó 2007 Elsevier Ltd. All rights reserved.
Keywords: Oligosaccharide synthesis; Thioglycosides; Oxidation
We had earlier published synthetic pathways to donor
building blocks suitable for the synthesis of Crypto-
coccus neoformans CPS structures, including those
corresponding to Xylp-(1?2)-Manp and Xylp-(1?2)-
[Xylp-(1?4)]-Manp di- and trisaccharides1 as well as
GlcpA-(1?2)-Manp and GlcpA-(1?2)-[Xylp-(1?4)]-
Manp di- and trisaccharides.2 These compounds were
subsequently used in the synthesis of larger structures
(up to heptasaccharides) with variant acetylation pat-
terns, which have been used in antibody binding- and
immunizing experiments.3
of the glucuronic acid-containing building blocks is
not always reliable and is difficult to perform on a large
scale. This is due to a number of reasons: the low
stereoselectivity in couplings with GlcpA-donors with
non-participating groups, the low reactivity and yields
in couplings with GlcpA-donors with participating
groups and difficulties in benzylating glucuronic acid
derivatives in reproducibly high yield. These are all
known problems frequently experienced in the synthesis
of uronic acid-containing oligosaccharides.4 An often
used solution to these problems is to instead start with
a suitably protected aldose donor and to perform
glycosylations and alkylations prior to oxidation to the
6-carboxylic acid function present in uronic acids.4 This
approach has earlier been applied to the synthesis of a
Cryptococcus pentasaccharide O-glycosidic structure by
van Boom and co-workers.5 We herein report on our
Although the synthesis of Xylp-(1?2)-Manp di- and
trisaccharide building blocks is practical, the synthesis
*
Corresponding author at present address: Centre for Synthesis and
Chemical Biology, University College Dublin, Belfield, Dublin 4,
Ireland; e-mail: stefan.oscarson@ucd.ie
0008-6215/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2007.11.026

J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
2201
experiences in employing this approach in the synthesis
of the di- and trisaccharide thioglycoside building blocks
9 and 17.
(e.g., PDC) oxidation often occurs. However, it has been
shown that oxidations with TEMPO–BAIB7 generally
do not affect thioglycosides.8 Other features of this
reagent are that it is most selective for primary positions
and that the carboxylic acid is obtained. Oxidation of 5
using this reagent mixture gave the desired uronic acid
derivative 6 in high yield (80%). Treatment with
PhCHN2 then gave the known benzyl ester 7 in 76%
yield. We were primarily interested in the 6-O-acetylated
disaccharide building block 9, and this was obtained
from 7 using an improved pathway. Regioselective
The same acceptor used in our earlier building block
syntheses, ethyl 2-O-allyl-4,6-O-benzylidene-1-thio-a-D-
mannopyranoside,1 was employed in these studies.
Glycosylation of this alcohol with 2,3,4,6-tetra-O-benzyl-
D-glucopyransosyl bromide promoted by silver triflate
gave a good yield (63%) of the (1?2)-b-linked disaccha-
ride 1 (Scheme 1). Deacylation by treatment with
sodium methoxide yielded the tetraol 2 in 80% yield.
Regioselective protection of the primary position, to
allow later deprotection and oxidation at C-60, was
performed using two different protecting groups, either
a TBDMS group or a trityl group. The former gave a
better yield in the selective protection and deprotection
whereas the latter gave a better yield in the intermediate
benzylation (no migration observed). Thus, silylation
using TBDMSCl and DMAP in pyridine gave an 89%
yield of the 60-O-silyl compound 3, which was benzyl-
ated (BnBr, NaH, DMF) to afford compound 4 in
67% yield together with 13% of the 4-O-TBDMS isomer.
Attempts to change conditions to avoid silyl migration
during the alkylation reaction were not successful. Desi-
lylation with TBAF afforded the 60-OH derivative 5
almost quantitatively (97%).
9
opening of the benzylidene acetal with Et3SiH–PhBCl2
gave the primary alcohol 8, which was acetylated to
afford 9 in an overall 81% yield. As compared to the
earlier synthesis of this building block, this route is
longer (9 steps as compared to 5 steps), but more reliable
and reproducible. The overall yields of both routes are
comparable (ꢀ15%).
In the synthesis of the trisaccharide, the fact that
xylopyranosides have no primary hydroxyl group could
be utilized (Scheme 2). First, the benzylidene acetal of
disaccharide 7 was again regioselectively opened, but
now to yield the 4-hydroxy derivative 10 by the use of
the NaCNBH3–HCl reagent system.10 Silver triflate-pro-
moted glycosylation of this acceptor with 2,3,4-tri-O-
benzyl-xylopyranosyl bromide afforded the b-(1?4)-
linked trisaccharide 11 in 85% yield. Deacylation
(NaOMe–MeOH, ?12, 80%) followed by regioselective
protection of the primary alcohol, this time using trityl
chloride, gave the 60-O-trityl derivative 13 (65%), which
was benzylated to afford 14 (92%). Detritylation using
p-TsOH in CHCl3–MeOH gave the 60-hydroxy derivative
Thioglycosides are stable to almost any protecting
and functional group manipulation and interconver-
sions.6 More or less only two types of reactions consti-
tute a problem: oxidation and catalytic hydrogenolysis.
With regard to oxidations, DMSO-based oxidations
do not oxidize the sulfur, whereas with other reagents
BzO
O
TBDMSO
BnO
BnO
R
RO
O
BzO
BzO
O
O
O
O
BnO
BnO
RO
RO
RO
RO
O
O
O
BnO
O
BzO
v
iii
i
viii
85%
OBn
OR
OR
OBn
Br
RO
BnO
AllO
97%
89%
O
O
Ph
O
Ph
63% Ph
O
O
O
O
O
OH
O
O
O
O
Ph
AllO
AllO
AllO
1 R = Bz
O
AllO
SEt
8 R = H
SEt
2 R = H (80%)
SEt
SEt
5 R = CH2OH
ix
3 R = H
vi
vii
iv
ii
9 R = Ac (95%)
SEt
6 R = COOH (81%)
7 R = COOBn (76%)
4 R = Bn (67%)
Scheme 1. Reagents: (i) AgOTf, DTBP, CH2Cl2; (ii) NaOMe, MeOH; (iii) TBDMSCl, DMAP, pyridine; (iv) BnBr, NaH, DMF; (v) TBAF, CHCl3/
MeOH, (vi) TEMPO, BAIB, CH2Cl2/H2O; (vii) PhCHN2, EtOAc; (viii) Et3SiH, PhBCl2, CH2Cl2; (ix) Ac2O, pyridine.
O
BzO
BzO
OBz
O
OBz
OTr
R
OR
O
BnO
BzO
i
BzO
BzO
BzO
O
O
iv
67%
O
Br
vi
65%
BnO
BnO
RO
RO
O
RO
RO
O
O
O
O
OBz
OBz
ii, 85%
OBn
OR
BnO
OR
O
O
BnO
O
Ph
O
BnO
O
78%
O
O
BnO
O
BnO
RO
O
RO
HO
O
O
O
O
BnO
RO
SEt
AllO
RO
AllO
AllO
AllO
BnO
AllO
OR
OR
10
7
SEt
SEt
15 R = CH2OH
SEt
SEt
vii
viii
11 R = Bz
12 R = H (80%)
13 R = H
14 R = Bn (92%)
iii
v
16 R = COOH (83%)
17 R = COOBn (49%)
Scheme 2. Reagents: (i) NaCNBH3, HCl/Et2O, THF; (ii) AgOTf, DTBP, CH2Cl2; (iii) NaOMe, MeOH; (iv) TrCl, DMAP, pyridine; (v) BnBr, NaH,
DMF; (vi) pTsOH, CHCl3/MeOH, (vii) TEMPO, BAIB, CH2Cl2/H2O; (viii) PhCHN2, EtOAc.

2202
J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
15 (67%), which was oxidized, again using TEMPO–
BAIB, to yield the glucuronic acid 16, once more in a
high yield (83%). Transformation into the benzyl ester
using phenyl diazomethane then gave the known tri-
saccharide building block 17. For the synthesis of the
trisaccharide block, this new approach is better. The
disadvantage of the three extra steps needed (protection,
deprotection and oxidation of the primary alcohol) is
more than compensated by the higher yields in and the
reproducibility of the glycosylation and the benzylation
reactions performed.
(1H, dd, J = 4.4, J = 10.0 Hz, H-60I), 3.97 (1H, m,
H-5I), 4.01 (1H, t, J = 9.6 Hz, H-4I), 4.13 (2H, dd,
J = 1.6, J = 5.6 Hz, OCH2CH@CH2), 4.18 (1H, ddd,
J = 3.2, 5. J = 6, 9.6 Hz, H-5II), 4.24 (1H, dd, J = 1.2,
J = 3.2 Hz, H-2I), 4.51 (1H, dd, J = 6.0, J = 12.0 Hz,
H-6II), 4.65 (1H, dd, J = 2.8, J = 12.0 Hz, H-60 ),
II
4.96 (1H, d, J = 7.6 Hz, H-1II), 5.02 (1H, dq, J = 1.6,
J = 10.8 Hz, OCH2CH@CH2), 5.18 (1H, d, J =
0.8 Hz, H-1I), 5.22 (1H, dq, J = 1.6, J = 17.2 Hz,
OCH2CH@CH2), 5.47 (1H, s, CHC6H5), 5.65 (1H, dd,
J = 7.6, J = 10.0 Hz, H-2II), 5.68 (1H, t, J = 10.0 Hz,
H-3II), 5.81 (1H, dddt, J = 1.6, J = 5.6, J = 10.8,
J = 17.2 Hz, OCH2CH@CH2), 5.93 (1H, t, J = 9.6 Hz,
H-4II), 7.11–8.01 (25H, m, Ar-H); 13C NMR
(100.6 MHz, CDCl3): d 14.9 (q, SCH2CH3), 25.6 (t,
SCH2CH3), 63.5 (t, C-6II), 64.6 (d, C-5I), 68.5 (t, C-6),
69.9 (d, C-3II), 71.0 (t, OCH2CH@CH2), 71.8 (d,
C-2II), 72.7 (d, C-4II), 72.8 (d, C-5II), 74.2 (d, C-3I),
78.3 (d, C-2I), 78.6 (d, C-4I), 82.7 (d, C-1), 100.2 (d,
C-1II), 101.7 (d, CHC6H5), 117.2 (t, OCH2CH@CH2),
126.2–129.9 (24C) (Ar-C), 131.7 (d, CHC6H5), 133.2,
133.3, 133.4, 133.6 (4 ꢂ s, 4 ꢂ COC6H5), 134.7 (d,
OCH2CHCH2), 137.7 (s, CHC6H5), 164.9, 165.3,
166.0, 166.2 (4 ꢂ s, 4 ꢂ COC6H5); HRMS (ESI):
[M+Na]+ calcd for C52H50O14S, 953.2813; found,
953.2793.
In conclusion, efficient and reproducible pathways to
glucuronic acid-containing derivatives 9 and 17 have been
developed. These thioglycosides are important building
blocks in the synthesis of C. neoformans CPS structures.
1. Experimental
1.1. General methods
TLC was carried out on Merck precoated 60 F254 plates
using AMC (ammonium molybdate–cerium(IV) sul-
fate–10% sulfuric acid 100 g:2 g:2 L) or 8% H2SO4 for
visualization. Column chromatography was performed
on silica gel (0.040–0.063 mm, Amicon) or reversed phase
gel (C18 60A 40–63 lm). NMR spectra were recorded in
CDCl3 (Me4Si, d 0.00 ppm) or D2O (acetone 13C d
30.89 ppm, 1H = 2.22 ppm) at 25 °C on a Varian
300 MHz or 400 MHz instrument. Optical rotations were
measured on a Perkin–Elmer 241 polarimeter. Organic
solutions were dried over Na2SO4 before concentration,
which was performed under reduced pressure.
1.3. Ethyl b-D-glucopyranosyl-(1?2)-3-O-allyl-4,6-O-
benzylidene-1-thio-a-D-mannopyranoside (2)
Compound 1 (1.8 g, 1.93 mmol) was dissolved in dry
MeOH (80 mL), 1 M methanolic NaOMe (1.5 mL)
was added and the mixture was stirred overnight at
room temperature. The progress of the reaction was
followed by TLC (CH2Cl2–MeOH, 9:1). Acetic acid
(15 drops) was added, the stirring was continued for
30 min, then the mixture was concentrated and purified
by silica gel chromatography (CH2Cl2–MeOH, 15:1) to
1.2. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyl-(1?2)-
3-O-allyl-4,6-O-benzylidene-1-thio-a-D-mannopyranoside
(1)
1
Silver triflate (2.33 g, 9.08 mmol) dissolved in dry
toluene was added at ꢁ50 °C to a stirred mixture
of 2,3,4,6-tetra-O-benzoyl-D-glucopyranosyl bromide
(5.99 g, 9.08 mmol), ethyl 3-O-allyl-4,6-O-benzylidene-
1-thio-a-D-mannopyranoside (1.60 g, 4.54 mmol) and
2,6-di-tert-butylpyridine (510 lL, 2.27 mmol) in distilled
yield 2 (0.8 g, 80%); [a]D +9.8 (c 0.2, DMSO-d6); H
NMR (400 MHz, DMSO): d 1.28 (3H, dt, J = 0.8,
J = 7.6 Hz, SCH2CH3), 2.40 (2H, dq, J = 7.0,
J = 7.2 Hz, SCH2CH3), 3.07 (1H, br s, OH), 3.38–3.42
(1H, m, H-5II), 3.50–3.66 (3H, m, H-2II, H-3II, H-4II),
3.82–3.91 (4H, H-6I, H-6II, H-60 , H-3I), 4.11–4.26
II
CH2Cl2 (150 mL) containing crushed 4 A molecular
(5H, m, H-2I, H-4I, H-5I, H-60 , OCH2CH@CH2), 4.31
˚
I
sieves (1.5 g). The reaction mixture was allowed to attain
room temperature overnight. The progress of the reac-
tion was followed by TLC (toluene–EtOAc, 6:1). After
18 h, Et3N (1.5 mL) was added, the mixture was diluted
with CH2Cl2 (150 mL), filtered through Celite, concen-
trated and purified by silica gel chromatography (tolu-
ene–EtOAc, 20:1) to give 1 (2.7 g, 63%); [a]D +32.8 (c
(1H, dd, J = 6.0 Hz, J = 12.8 Hz, OCH2CH@CH2),
4.48 (1H, br s, OH), 4.53 (1H, d, J = 8.0 Hz, H-1II),
5.21 (1H, br d, J = 10.4 Hz, OCH2CH@CH2), 5.31
(1H, br d, J = 18.4 Hz, OCH2CH@CH2), 5.35 (1H, s,
H-1I), 5.60 (1H, s, CHC6H5), 5.90 (1H, ddt, J = 6.0,
J = 10.4, J = 16.8 Hz, OCH2CH@CH2), 7.33–7.39
(3H, m, Ar-H), 7.46–7.50 (2H, m, Ar-H); 13C NMR
(100.6 MHz, DMSO): d 15.2 (q, SCH2CH3), 25.7 (t,
SCH2CH3), 62.2 (t, C-6II), 64.7 (d, C-5I), 68.8 (t, C-6I),
70.0 (d, C-4II), 71.5 (d, C-2II), 72.6 (t, OCH2CH@CH2),
74.7 (d, C-3I), 75.8 (d, C-4I), 76.1 (d, C-3II), 76.4 (d,
1
1.0, CHCl3); H NMR (400 MHz, CDCl3): d 1.10 (3H,
br t, J = 7.6 Hz, SCH2CH3), 2.40 (2H, dq, J = 7.0,
J = 7.2 Hz, SCH2CH3), 3.46 (1H, t, J = 10.0 Hz,
H-6I), 3.77 (1H, dd, J = 3.2, J = 9.6 Hz, H-3I), 3.83

J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
2203
C-5II), 79.1 (d, C-2I), 84.9 (d, C-1), 101.6 (d, CHC6H5),
102.0 (d, C-1II), 118.7 (t, OCH2CH@CH2), 126.1 (d,
2C), 128.4 (d, 2C), 129.1 (d) (Ar-C), 134.2 (d,
OCH2CH@CH2), 137.6 (s) (Ar-C); MALDI-TOF
MS: [M+Na]+ calcd for C24H34O10S, 514.59; found,
537.58.
tion was followed by TLC (toluene–EtOAc, 6:1). MeOH
(1 mL) was carefully added and the mixture was diluted
with toluene (100 mL), washed with water (2 ꢂ 50 mL),
dried and concentrated. Purification on a silica gel col-
umn (toluene–EtOAc, 30:1) gave 4 (440 mg, 67%). 13%
of the migrated product ethyl 2,3,6-tri-O-benzyl-4-
O-tert-butyldimethylsilyl-b-D-glucopyranosyl-(1?2)-3-
O-allyl-4,6-O-benzylidene-1-thio-a-D-mannopyranoside
was also isolated. 4: [a]D +38.6 (c 1.0, CHCl3); 1H NMR
(400 MHz, CHCl3): d 0.09 (3H, (CH3)3C(CH3)2Si), 0.11
(3H, (CH3)3C(CH3)2Si), 0.93 (9H, (CH3)3C(CH3)2Si),
1.32 (3H, dt, J = 1.2, J = 7.2 Hz, SCH2CH3), 2.59–
2.74 (2H, m, SCH2CH3), 3.33–3.38 (1H, m, H-5II),
3.59–3.72 (3H, m, H-2II, H-3II, H-4II), 3.76–3.92 (3H,
1.4. Ethyl 6-O-tert-butyldimethylsilyl-b-D-glucopyran-
osyl-(1?2)-3-O-allyl-4,6-O-benzylidene-1-thio-a-D-
mannopyranoside (3)
tert-Butyldimethylchlorosilane (282 mg, 1.03 mmol) was
added to a solution of 2 (440 mg, 0.86 mmol) in pyridine
(11 mL) at room temperature and the mixture was stir-
red overnight. The progress of the reaction was followed
by TLC (CH2Cl2–MeOH, 9:1). The mixture was concen-
trated and purified by silica gel chromatography (tolu-
ene–EtOAc, 1:2) to afford 3 (480 mg, 89%); [a]D +12.1
(c 0.3, CHCl3); 1H NMR (400 MHz, CHCl3): d 0.09
(3H, (CH3)3C(CH3)2Si), 0.10 (3H, (CH3)3C(CH3)2Si),
0.91 (9H, (CH3)3C(CH3)2Si), 1.28 (3H, dt, J = 0.8,
J = 7.2 Hz, SCH2CH3), 2.55–2.69 (2H, m, SCH2CH3),
2.86 (1H, br s, OH), 3.19 (1H, br s, OH), 3.38–3.43
(1H, m, H-5II), 3.46–3.51 (1H, m, H-2II), 3.58–3.64
(2H, m, H-3I, H-3II), 3.81 (1H, dd, J = 5.2,
J = 10.8 Hz, H-6II), 3.85–3.89 (2H, m, H-4II, H-6I),
3.92 (1H, dd, J = 4.4, J = 10.8 Hz, H-60II), 4.12–4.26
m, H-6I, H-60 , H-6II), 3.96 (1H, dd, J = 3.2, J =
I
10.0 Hz, H-3I), 4.17–4.34 (5H, m, H-4I, H-5I, H-60 ,
II
OCH2CH@CH2), 4.37 (1H, d, J = 3.6 Hz, H-2I),
4.52 (1H, d, J = 7.6 Hz, H-1II), 4.70 (1H, d, J =
10.8 Hz), 4.73 (1H, d, J = 10.4 Hz), 4.84 (1H, d,
J = 11.2 Hz), 4.89 (1H, d, J = 10.8 Hz), 4.97 (1H, d,
J = 11.2 Hz), 5.14 (1H, d, J = 10.4 Hz) (OCH2Ph),
5.21 (1H, dq, J = 1.2, J = 11.6 Hz, OCH2CH@CH2),
5.38 (1H, dq, J = 1.2, J = 17.2 Hz, OCH2CH@CH2),
5.52 (1H, s, H-1I), 5.58 (1H, s, CHC6H5), 5.91–6.01
(1H, m, OCH2CH@CH2), 7.29–7.40 (16H, m), 7.44–
7.46 (2H, m), 7.51–7.55 (2H, m) (Ar-H); 13C NMR
(100.6 MHz, CDCl3): d ꢁ5.1 (q, (CH3)3C(CH3)2Si),
ꢁ5.3 (q, (CH3)3C(CH3)2Si), 15.1 (q, SCH2CH3), 18.4
(s, (CH3)3C(CH3)2Si), 25.7 (t, SCH2CH3), 26.1 (q, 3C,
(CH3)3C(CH3)2Si), 62.5 (t, C-6II), 64.6 (d, C-5I), 68.8
(t, C-6I), 70.5 (t, OCH2CH@CH2), 73.7 (d, C-3I), 75.1
(t, CH2Ph), 75.3 (t, CH2Ph), 75.8 (t, CH2Ph), 76.5 (d,
2C, C-2I, C-5II), 77.5 (d, C-4II), 78.9 (d, C-4I), 82.2 (d,
C-2II), 82.9 (d, C-1I), 84.9 (d, C-3II), 101.6 (d, C-1II),
101.7 (d, CHC6H5), 117.6 (t, OCH2CH@CH2), 126.1
(d, 2C), 127.7 (d), 127.9 (d, 2C), 128.0 (d, 4C), 128.2
(d, 2C), 128.5 (d, 2C), 128.5 (d, 4C), 128.6 (d, 2C),
128.9 (d) (Ar-C), 134.8 (d, OCH2CH@CH2), 137.7,
138.4, 138.5, 138.7 (4 ꢂ s) (Ar-C); HRMS (ESI):
[M+Na]+ calcd for C51H66O10SSi, 921.4038; found,
921.4080.
(5H, m, H-2I, H-4I, H-5I, H-60 , OCH2CH@CH2),
I
4.39 (1H, ddt, J = 1.2, J = 5.6, J = 12.4 Hz, OC-
H2CH@CH2), 4.58 (1H, d, J = 8.0 Hz, H-1II), 5.23
(1H, dq, J = 1.6, J = 10.4 Hz, OCH2CH@CH2), 5.28
(1H, s, H-1I), 5.29 (1H, dq, J = 1.6, J = 17.2 Hz,
OCH2CH@CH2), 5.60 (1H, s, CHC6H5), 5.86–5.96
(1H, m, OCH2CH@CH2), 7.34–7.40 (3H, m, Ar-H),
7.46–7.50 (2H, m, Ar-H); 13C NMR (100.6 MHz,
CDCl3): d ꢁ5.3 (q, 2C, (CH3)3C(CH3)2Si), 15.2 (q,
SCH2CH3), 18.4 (s, (CH3)3C(CH3)2Si), 25.7 (t,
SCH2CH3), 26.0 (q, 3C, (CH3)3C(CH3)2Si), 64.3 (t,
C-6II), 64.5 (d, C-5I), 68.8 (t, C-6I), 70.1 (d, C-2II),
72.4 (d, C-3I), 73.5 (t, OCH2CH@CH2), 73.7 (d, C-2I),
74.9 (d, C-4II), 75.5 (d, C-5II), 75.9 (d, C-3II), 79.9 (d,
C-4I), 85.8 (d, C-1), 101.5 (d, C-1II), 101.7 (d, CHC6H5),
119.1 (t, OCH2CH@CH2), 126.1 (d, 2C), 128.4 (d, 2C),
129.1 (d) (Ar-C), 133.8 (d, OCH2CH@CH2), 137.6 (s)
(Ar-C); HRMS (ESI): [M+Na]+ calcd for C30H48O10-
SSi, 651.2630; found, 651.2608.
1.6. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyl-(1?2)-
3-O-allyl-4,6-O-benzylidene-1-thio-a-D-mannopyranoside
(5)
TBAF (180 mg, 0.570 mmol) was added to a solution of
4 (379 mg, 0.421 mmol) in THF (30 mL) and the
mixture was stirred at room temperature for 1 h. The
progress of the reaction was followed by TLC (tolu-
ene–EtOAc, 6:1). The reaction mixture was then concen-
trated and purified by silica gel chromatography
(toluene–EtOAc, 10:1) to yield 5 (320 mg, 97%); [a]D
+46.4 (c 0.8, CHCl3); H NMR (400 MHz, CHCl3): d
1.25 (3H, t, J = 7.6 Hz, SCH2CH3), 2.06 (1H, br s,
OH), 2.55–2.64 (2H, m, SCH2CH3), 3.35–3.39 (1H, m,
1.5. Ethyl 2,3,4-tri-O-benzyl-6-O-tert-butyldimethylsilyl-
b-D-glucopyranosyl-(1?2)-3-O-allyl-4,6-O-benzylidene-
1-thio-a-D-mannopyranoside (4)
A 60% oil dispersion of sodium hydride (132 mg,
3.29 mmol) was added to a solution of 3 (460 mg,
0.73 mmol) and benzyl bromide (350 lL, 2.94 mmol)
in DMF (30 mL) at room temperature and the reaction
mixture was stirred overnight. The progress of the reac-
1

2204
J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
H-5II), 3.54–3.61 (2H, m, H-2II, H-4II), 3.62–3.72 (2H,
d 15.1 (q, SCH2CH3), 25.8 (t, SCH2CH3), 64.7 (d,
C-5I), 68.8 (t, C-6I), 72.0 (t, OCH2CH@CH2), 74.3 (t,
CH2Ph), 74.6 (d, C-3I), 75.1 (d, C-5II), 75.2 (t, CH2Ph),
75.3 (t, CH2Ph), 79.2 (d, C-4I), 79.4 (d, C-2I), 79.8 (d,
C-4II), 80.2 (d, C-2II), 82.8 (d, C-3II), 83.6 (d, C-1I),
101.8 (d, CHC6H5), 102.9 (d, C-1II), 118.2 (t,
OCH2CH@CH2), 126.2 (d, 2C), 127.9 (d, 3C), 128.1
(d), 128.2 (d), 128.4 (d, 4C), 128.6 (d, 8C), 129.1 (d)
(Ar-C), 134.2 (d, OCH2CH@CH2), 137.5 (2C), 138.0,
138.3, (4 ꢂ s) (Ar-C); HRMS (ESI): [M+Na]+ calcd
for C45H50O11S, 821.2966; found, 821.2977.
m, H-3II, H-6II), 3.76–3.84 (2H, m, H-6I, H-60 ), 3.86
II
(1H, dd, J = 3.2, J = 9.6 Hz, H-3I), 4.14 (1H, t,
J = 9.6 Hz, H-4I), 4.18–4.29 (5H, m, H-2I, H-60 , H-5I,
I
OCH2CH@CH2), 4.49 (1H, d, J = 8.0 Hz, H-1II), 4.62
(1H, d, J = 10.8 Hz), 4.70 (1H, d, J = 10.0 Hz), 4.82
(1H, d, J = 11.2 Hz), 4.84 (1H, d, J = 10.8 Hz), 4.94
(1H, d, J = 11.2 Hz), 5.02 (1H, d, J = 10.0 Hz)
(OCH2Ph), 5.18 (1H, dq, J = 1.2, J = 10.4 Hz,
OCH2CH@CH2), 5.33 (1H, dq, J = 1.2, J = 17.2 Hz,
OCH2CH@CH2), 5.40 (1H, s, H-1I), 5.56 (1H, s,
CHC6H5), 5.86–5.96 (1H, m, OCH2CH=CH2), 7.23–
7.48 (20H, m, Ar-H); 13C NMR (100.6 MHz, CDCl3):
d 15.2 (q, SCH2CH3), 25.8 (t, SCH2CH3), 62.3 (t,
C-6II), 64.7 (d, C-5I), 68.9 (t, C-6I), 71.6 (t,
OCH2CH@CH2), 74.7 (d, C-3I), 75.2 (t, CH2Ph), 75.3
(t, CH2Ph), 75.9 (t, CH2Ph), 76.5 (d, C-5II), 77.7 (d,
C-4II), 79.2 (d, 2C, C-2I,C-4I), 81.0 (d, C-2II), 83.7 (d,
C-1I), 84.6 (d, C-3II), 101.8 (d, CHC6H5), 103.2 (d,
C-1II), 117.2 (t, OCH2CH@CH2), 126.2 (d, 2C), 127.8
(d), 128.0 (d, 2C), 128.1 (d, 2C), 128.3 (d, 2C), 128.4
(d, 2C), 128.6 (d, 6C), 128.7 (d, 2C), 129.0 (d) (Ar-C),
134.9 (d, OCH2CH@CH2), 137.6, 138.0, 138.3, 138.7
(4 ꢂ s) (Ar-C); HRMS (ESI): [M+Na]+ calcd for
C45H52O10S, 807.3173; found, 807.3208.
1.8. Ethyl benzyl (2,3,4-tri-O-benzyl-b-D-glucopyran-
osyl)uronate-(1?2)-3-O-allyl-4,6-O-benzylidene-1-
thio-a-D-mannopyranoside (7)
A solution of 6 (235 mg, 0.294 mmol) in EtOAc (50 mL)
was titrated with PhCHN2 (0.3 M solution in Et2O,
3 mL, 0.9 mmol) at room temperature until full conver-
sion was observed by TLC (toluene–EtOAc, 6:1). After
2 h, the reaction was concentrated and purified by silica
gel chromatography (toluene–EtOAc, 20:1) to afford 7
1
(200 mg, 76%); [a]D +26.7 (c 0.8, CHCl3), lit.2 +22; H
NMR (400 MHz, CHCl3): d 1.26 (3H, t, J = 7.6 Hz,
SCH2CH3), 2.54–2.64 (2H, m, SCH2CH3), 3.62–3.64
(2H, m, H-2II, H-3II), 3.77 (1H, t, J = 11.6 Hz, H-6I),
3.83–3.92 (3H, m, H-3I, H-4II, H-5II), 4.03–4.15 (3H,
m, H-4I, OCH2CH@CH2), 4.21–4.27 (3H, m, H-2I,
1.7. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyluronic
acid-(1?2)-3-O-allyl-4,6-O-benzylidene-1-thio-a-D-
mannopyranoside (6)
H-5I, H-60 ), 4.47 (1H, d, J = 10.4 Hz) (OCH2Ph), 4.48
I
(1H, d, J = 7.6 Hz, H-1II), 4.67 (1H, d, J = 10.4 Hz),
4.71 (1H, d, J = 10.4 Hz), 4.77 (1H, d, J = 11.2 Hz),
4.90 (1H, d, J = 11.2 Hz), 5.03 (1H, d, J = 10.4 Hz),
5.12 (2H, s), (OCH2Ph), 5.10 (1H, dt, J = 1.6,
J = 10.4 Hz, OCH2CH@CH2), 5.28 (1H, dt, J = 1.6,
J = 17.2 Hz, OCH2CH@CH2), 5.40 (1H, d, J = 1.2 Hz,
H-1I), 5.53 (1H, s, CHC6H5), 5.78–5.88 (1H, m,
OCH2CH@CH2), 7.07–7.47 (25H, m, Ar-H); 13C
NMR (100.6 MHz, CDCl3): d 15.1 (q, SCH2CH3),
25.9 (t, SCH2CH3), 64.7 (d, C-5I), 67.5 (t, CH2Ph),
68.9 (t, C-6I), 70.5 (t, OCH2CH@CH2), 74.2 (d, C-3I),
74.9 (d, C-4II), 75.2 (t, CH2Ph), 75.4 (t, CH2Ph), 76.0
(t, CH2Ph), 77.7 (d, C-2I), 78.8 (d, C-4I), 79.1 (d,
C-5II), 81.4 (d, C-2II), 83.3 (d, C-1I), 84.1 (d, C-3II),
101.8 (d, CHC6H5), 102.8 (d, C-1II), 117.3 (t,
OCH2CH@CH2), 126.2 (d, 2C), 127.8 (d), 127.9 (d,
3C), 128.0 (d, 2C), 128.1 (d), 128.3 (d, 2C), 128.5 (d,
2C), 128.6 (d, 4C), 128.7 (d, 3C), 128.8 (d, 4C), 129.1
(d) (Ar-C), 134.7 (d, OCH2CH@CH2), 135.5, 137.7,
138.0, 138.2, 138.5, (5 ꢂ s) (Ar-C), 167.9 (s, C-6II);
HRMS (ESI): [M+Na]+ calcd for C52H56O11S,
911.3436; found, 911.3477.
To
a vigorously stirred mixture of 5 (314 mg,
0.40 mmol) in CH2Cl2 (20 mL) and H2O (10 mL), TEM-
PO (250 mg, 1.60 mmol) and BAIB (1.29 g, 4.0 mmol)
were added at room temperature for 2 h. The progress
of the reaction was followed by TLC (hexane–EtOAc–
HOAc, 16:3:1). The reaction was quenched by the addi-
tion of 10% solution of Na2S2O3 (25 mL). The mixture
was extracted with EtOAc (2 ꢂ 80 mL), the combined
organic layers were dried and concentrated. Purification
of the residue on a silica gel column (hexane–EtOAc–
HOAc, 16:3:1) gave 6 (260 mg, 81%); [a]D +9.1 (c 0.5,
1
CHCl3); H NMR (400 MHz, CHCl3): d 1.23 (3H, t,
J = 7.6 Hz, SCH2CH3), 2.52–2.62 (2H, m, SCH2CH3),
3.57 (1H, t, J = 8.0 Hz, H-2II), 3.66 (1H, t, H-3II), 3.80
(1H, t, J = 11.6 Hz, H-6I), 3.88 (1H, dd, J = 3.2,
J = 10.0 Hz, H-3I), 4.11 (1H, t, J = 7.2 Hz, H-4II), 4.11
(1H, t, J = 9.6 Hz, H-4I), 4.14 (1H, d, J = 7.2 Hz,
H-5II), 4.19–4.26 (5H, m, H-2I, H-5I, H-60I,
OCH2CH@CH2), 4.65 (1H, d, J = 7.6 Hz, H-1II), 4.69
(2H, s), 4.70 (1H, d, J = 10.0 Hz), 4.75 (1H, d,
J = 11.2 Hz), 4.82 (1H, d, J = 11.2 Hz), 4.88 (1H, d,
J = 10.0 Hz) (OCH2Ph), 5.15 (1H, br d, J = 10.4 Hz,
OCH2CH@CH2), 5.29 (1H, br d, J = 17.2 Hz,
OCH2CH@CH2), 5.33 (1H, s, H-1I), 5.57 (1H, s,
CHC6H5), 5.83–5.92 (1H, m, OCH2CH@CH2), 7.26–
7.48 (20H, m, Ar-H); 13C NMR (100.6 MHz, CDCl3):
PhCHN2 is potentially explosive and may burn violently when
exposed to air. This compounds can be redistilled under high
vacuum.11

J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
2205
1
1.9. Ethyl benzyl (2,3,4-tri-O-benzyl-b-D-glucopyran-
osyl)uronate-(1?2)-3-O-allyl-4-O-benzyl-1-thio-a-D-
mannopyranoside (8)
(48 mg, 95%); [a]D +28.8 (c 1.0, CHCl3), lit.2 +22; H
NMR (400 MHz, CHCl3): d 1.29 (3H, t, J = 7.6 Hz,
SCH2CH3), 1.70 (3H, s, COCH3), 2.59–2.70 (2H, m,
SCH2CH3), 3.63–3.72 (2H, m, H-2II, H-3II), 3.81 (1H,
dd, J = 3.2, J = 9.2 Hz, H-3I), 3.89–4.02 (4H, m, H-4I,
H-5I, H-4II, OCH2CH@CH2), 4.16–4.29 (4H, m, H-2I,
H-6I, H-5II, OCH2CH@CH2), 4.38 (1H, dd, J = 4.0,
A solution of 7 (178 mg, 0.200 mmol) in CH2Cl2
˚
(20 mL), containing crushed 4 A molecular sieves
(0.5 g), was cooled to ꢁ78 °C and Et3SiH (97 lL,
0.607 mmol) and PhBCl2 (100 lL, 0.762 mmol) were
added. The reaction mixture was stirred at ꢁ78 °C for
2 h. Then Et3N (0.8 mL) was added, the mixture was
allowed to attain room temperature, concentrated and
purified by silica gel chromatography (toluene–EtOAc,
12:1) to yield 8 (152 mg, 85%); [a]D +14.9 (c 0.5,
J = 12.0 Hz, H-60 ), 4.50–4.55 (2H, m, H-1II, OCH2Ph),
I
4.56 (1H, d, J = 10.8 Hz), 4.63 (1H, d, J = 10.0 Hz),
4.75 (1H, d, J = 10.8 Hz), 4.79 (1H, d, J = 11.2 Hz),
4.94 (1H, d, J = 11.2 Hz), (OCH2Ph), 5.13–5.20 (2H,
m, OCH2Ph, OCH2CH@CH2), 5.16 (2H, s), (OCH2Ph),
5.30 (1H, dt, J = 1.2, J = 17.2 Hz, OCH2CH@CH2),
5.41 (1H, s, H-1I), 5.91 (1H, ddt, J = 6.0, J = 10.4,
J = 17.2 Hz, OCH2CH@CH2), 7.11–7.42 (25H, m,
Ar-H); 13C NMR (100.6 MHz, CDCl3): d 15.1 (q,
SCH2CH3), 20.6 (q, COCH3), 25.9 (t, SCH2CH3), 63.5
(t, C-6I), 67.5 (t, CH2Ph), 70.0 (d, C-5I), 70.1 (t,
OCH2CH@CH2), 74.1 (d), 74.8 (d), (C-4I, C-4II), 75.2
(t, 2C, CH2Ph), 75.4 (t, CH2Ph), 75.9 (t, CH2Ph), 76.5
(d, C-2I), 78.4 (d, C-3I), 78.9 (d, C-5II), 81.2 (d, C-2II),
82.5 (d, C-1I), 84.1 (d, C-3II), 102.9 (d, C-1II), 118.1 (t,
OCH2CH@CH2), 127.8 (d), 127.9 (d, 3C), 128.1 (d,
2C), 128.2 (d, 3C), 128.5 (d, 9C), 128.7 (d, 3C), 128.9
(d, 2C), 129.2 (d, 2C) (Ar-C), 134.7 (d, OCH2C-
H@CH2), 135.1, 137.9, 138.0, 138.4, 138.6 (5 ꢂ s) (Ar-
C), 167.9 (s, C-6II), 170.8 (s, COCH3); HRMS (ESI):
[M+Na]+ calcd for C54H60O12S, 955.3698; found,
955.3740.
1
CHCl3); H NMR (400 MHz, CHCl3): d 1.28 (3H, t,
7.2 Hz, SCH2CH3), 2.56–2.66 (2H, m, SCH2CH3),
3.63–3.76 (4H, m, H-2II, H-3II, H-6I, H-60 ), 3.79 (1H,
I
dd, J = 3.2, J = 9.6 Hz, H-3I), 3.90 (1H, t, J = 9.6 Hz,
H-4I), 3.90–3.94 (2H, m, H-4II, H-5II), 3.95–4.02 (2H,
m, H-5I, OCH2CH@CH2), 4.14–4.18 (1H, m,
OCH2CH@CH2), 4.23 (1H, dd, J = 1.2, J = 3.2 Hz,
H-2I), 4.51–4.53 (2H, m, H-1II, OCH2Ph), 4.59 (1H, d,
J = 11.2 Hz), 4.70 (1H, d, J = 10.8 Hz), 4.74 (1H, d,
J = 10.8 Hz), 4.78 (1H, d, J = 10.8 Hz), 4.90 (1H, d,
J = 10.4 Hz), 5.06 (1H, d, J = 10.4 Hz), 5.15 (2H, s),
(OCH2Ph), 5.15 (1H, dt, J = 0.8, J = 10.4 Hz,
OCH2CH@CH2), 5.27 (1H, dt, J = 0.8, J = 17.2 Hz,
OCH2CH@CH2), 5.40 (1H, s, H-1I), 5.84–5.94 (1H, m,
OCH2CH@CH2), 7.11–7.39 (25H, m, Ar-H); 13C
NMR (100.6 MHz, CDCl3): d 15.0 (q, SCH2CH3),
25.7 (t, SCH2CH3), 62.6 (t, C-6I), 67.5 (t, CH2Ph),
70.3 (t, OCH2CH@CH2), 72.2 (d, C-5I), 74.7 (d, C-4I),
74.9 (d, 2C, C-4II, CH2Ph) 75.2 (t, CH2Ph), 75.4 (t,
CH2Ph), 75.9 (t, CH2Ph), 76.4 (d, C-2I), 78.3 (d, C-3I),
78.9 (d, C-5II), 81.2 (d, C-2II), 82.2 (d, C-1I), 84.1 (d,
C-3II), 102.6 (d, C-1II), 118.0 (t, OCH2CH=CH2),
127.8 (d), 127.9 (d, 2C), 128.0 (d, 5C), 128.1 (d, 2C),
128.2 (d, 2C), 128.5 (d, 6C), 128.6 (d, 2C), 128.7 (d),
128.8 (d, 2C), 128.9 (d, 2C) (Ar-C), 134.8 (d, OC-
H2CH@CH2), 135.2, 138.0, 138.4, 138.5, 138.6 (5 ꢂ s)
(Ar-C), 167.9 (s, C-6II); HRMS (ESI): [M+Na]+ calcd
for C52H58O11S, 913.3592; found 913.3624.
1.11. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyl-(1?2)-
3-O-allyl-6-O-benzyl-1-thio-a-D-mannopyranoside (10)
To a mixture of 7 (931 mg, 1.00 mmol) and NaCNBH3
(380 mg, 6.05 mmol) in THF (40 mL) containing
˚
crushed 3 A molecular sieves (0.5 g) a solution of HCl
in Et2O was added (to pH 1–2) at room temperature.
The progress of the reaction was followed by TLC (tol-
uene–EtOAc, 6:1). After 2 h, Et3N (4 mL) was added,
the mixture was diluted with CH2Cl2 (150 mL), filtered
through Celite, concentrated and purified by silica gel
chromatography (toluene–EtOAc, 7:1) to yield 10
(730 mg, 78%); [a]D +22.1 (c 1.0, CHCl3); 1H NMR
1.10. Ethyl benzyl (2,3,4-tri-O-benzyl-b-D-glucopyran-
osyl)uronate-(1?2)-6-O-acetyl-3-O-allyl-4-O-benzyl-1-
thio-a-D-mannopyranoside (9)
(400 MHz, CDCl3):
d
1.14 (3H, t, J = 7.6 Hz,
SCH2CH3), 2.39–2.54 (2H, m, SCH2CH3), 3.26 (1H,
dd, J = 6.8, J = 10.8 Hz, H-6I), 3.54–3.54 (2H, m,
Ac2O (1 mL, 10.6 mmol) was added at room tempera-
ture to a solution of 8 (48 mg, 0.054 mmol) in pyridine
(10 mL), and the mixture was stirred overnight. The
progress of the reaction was observed by TLC (tolu-
ene–EtOAc, 6:1). Then toluene (40 mL) and 1 M HCl
(30 mL) were added, the organic layer was separated
and washed with 1 M HCl (30 mL), saturated solution
of NaHCO3 (30 mL) and water (40 mL), dried and
concentrated. Purification of the residue by silica gel
H-3I, H-60 ), 3.74 (1H, t, J = 9.6 Hz, H-4I), 3.91–3.97
I
(2H, m, H-5I, OCH2CH@CH2), 4.15–4.22 (3H, m,
H-2I, H-5II, OCH2CH@CH2), 4.25 (2H, s, OCH2Ph),
4.49 (1H, dd, J = 5.6, J = 12.0 Hz, H-6II), 4.64 (1H,
dd, J = 2.8, J = 12.0 Hz, H-60II), 4.95 (1H, d,
J = 8.0 Hz, H-1II), 5.06 (1H, br d, J = 10.0 Hz,
OCH2CH@CH2), 5.22 (1H, dq, J = 1.6, J = 17.2 Hz,
OCH2CH@CH2), 5.23 (1H, s, H-1I), 5.59 (1H, dd,
J = 8.0, J = 9.6 Hz, H-2II), 5.65 (1H, t, J = 10.4 Hz,
H-4II), 5.79–5.89 (1H, m, OCH2CH@CH2), 5.91 (1H,
chromatography (toluene–EtOAc, 12:1) afforded
9

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J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
t, J = 10.0 Hz, H-3II), 7.14–8.00 (25H, m, Ar-H); 13C
NMR (100.6 MHz, CDCl3): d 14.8 (q, SCH2CH3),
25.4 (t, SCH2CH3), 63.4 (t, C-6II), 68.0 (d, C-4I), 69.8
(d, C-4II), 69.9 (t, OCH2CH@CH2), 70.9 (t, C-6), 71.7
(d, C-5I), 71.8 (d, C-2II), 72.6 (d, C-5II), 72.9 (d, C-3II),
73.4 (t, CH2Ph), 75.6 (d, C-2I), 77.6 (d, C-3I), 81.3 (d,
C-1), 99.8 (d, C-1II), 118.1 (t, OCH2CH@CH2), 127.5–
129.9 (25C) (Ar-C), 133.1, 133.3, 133.4, 133.6 (4 ꢂ s,
4 ꢂ COC6H5), 134.5 (d, OCH2CHCH2), 138.4 (s, Ar-
C), 164.9, 165.3, 165.9, 166.2 (4 ꢂ s, 4 ꢂ COC6H5);
HRMS (ESI): [M+Na]+ calcd for C52H52O14S,
955.2970; found, 955.2949.
30 min, then the mixture was concentrated and purified
by silica gel chromatography (CH2Cl2–MeOH, 9:1) to
give 12 (322 mg, 80%); [a]D +5.1 (c 1.0, CHCl3); H
1
NMR (400 MHz, DMSO): d 4.20 (1H, d, J = 7.6 Hz,
H-1III), 4.31 (1H, d, J = 7.6 Hz, H-1II), 5.39 (1H, s,
H-1I); 13C NMR (100.6 MHz, DMSO): d 14.9 (q,
SCH2CH3), 24.6 (t, SCH2CH3), 61.4 (t, C-5III), 65.8
(t, C-6II), 68.8 (t, C-6I), 69.6 (d, C-5I), 70.0 (t,
OCH2CH@CH2), 70.1 (d), 71.7 (d), 72.1 (t, CH2Ph),
72.9 (d, C-2III), 73.7 (d, C-2II), 74.2 (d, C-4I), 74.7 (d,
C-2I), 76.6 (d, C-3I), 76.6 (2C, 2 ꢂ d), 77.2 (d), 81.8 (d,
C-1), 101.4 (d, C-1II), 103.8 (d, C-1III), 116.4 (t,
OCH2CH@CH2), 127.3 (d), 127.4 (2C, 2 ꢂ d), 128.2
(2C, 2 ꢂ d) (Ar-C), 135.5 (d, OCH2CHCH2), 138.6 (s,
Ar-C); HRMS (ESI): [M+Na]+ calcd for C78H72O21S,
1399.4179; found, 1399.4151.
1.12. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyl-(1?2)-
2,3,4-tri-O-benzyl-b-D-xylopyranosyl-(1?4)-3-O-allyl-6-
O-benzyl-1-thio-a-D-mannopyranoside (11)
Silver triflate (475 mg, 1.85 mmol) dissolved in dry tolu-
1.14. Ethyl 2,3,4-tri-O-benzyl-6-O-trityl-b-D-glucopyran-
osyl-(1?2)-2,3,4-tri-O-benzyl-b-D-xylopyranosyl-(1?4)-
3-O-allyl-6-O-benzyl-1-thio-a-D-mannopyranoside (14)
ene was added at ꢁ50 °C to a stirred mixture of 2,3,4-
tri-O-benzoyl-D-xylopyranosyl
bromide
(971 mg,
1.85 mmol), 10 (690 mg, 0.74 mmol) and 2,6-di-tert-
butylpyridine (330 lL, 1.48 mmol) in distilled CH2Cl2
(100 mL) containing crushed 4 A molecular sieves
Compound 12 (650 mg, 1.0 mmol) was dissolved in dry
pyridine (10 mL), trityl chloride (670 mg, 2.4 mmol)
and N,N-dimethylaminopyridine (24 mg, 0.2 mmol)
were added and the mixture was stirred at 70 °C for
4 h. The progress of the reaction was followed by TLC
(CH2Cl2–MeOH, 9:1). Then the mixture was concen-
trated and purified by silica gel chromatography
(CH2Cl2–MeOH, 10:1) to yield ethyl 6-O-trityl-b-D-gluco-
pyranosyl-(1?2)-b-D-xylopyranosyl-(1?4)-3-O-allyl-6-
O-benzyl-1-thio-a-D-mannopyranoside (13, 580 mg,
65%). 1H NMR (400 MHz, CDCl3): d 4.44 (1H, d,
J = 7.2 Hz, H-1III), 4.44 (1H, d, J = 7.2 Hz, H-1II),
5.36 (1H, br s, H-1I); 13C NMR (100.6 MHz, CDCl3):
d 15.2 (q, SCH2CH3), 25.9 (t, SCH2CH3), 63.9 (t,
C-5III), 65.6 (t, C-6II), 69.1 (t, C-6I), 69.8 (d, C-5I),
71.4 (d), 71.8 (2C, 2 ꢂ d), 71.8 (t, OCH2CH@CH2),
73.5 (d, C-2III), 73.6 (d, C-2I), 73.8 (t, CH2Ph), 74.2
(d), 74.8 (d), 76.1 (d), 76.3 (d), 77.3 (d, C-3I), 83.3 (d,
C-1), 87.2 (s, CPh3), 100.7 (d, C-1II), 103.4 (d, C-1III),
119.2 (t, OCH2CH@CH2), 127.4 (4C, 4 ꢂ d), 128.1
(6C, 6 ꢂ d), 128.2 (2C, 2 ꢂ d), 128.6 (2C, 2 ꢂ d), 128.8
(6C, 6 ꢂ d) (Ar-C), 134.0 (d, OCH2CHCH2), 137.8 (s),
143.7 (s) (Ar-C).
˚
(1.0 g). The temperature was allowed to go up to
ꢁ20 °C. The progress of the reaction was followed by
TLC (toluene–EtOAc, 3:1). After 3 h, Et3N (1.5 mL)
was added (pH 8), the mixture was diluted with CH2Cl2
(150 mL), filtered through Celite, concentrated and puri-
fied by silica gel chromatography (toluene–EtOAc, 22:1)
to afford 11 (870 mg, 85%); [a]D +22.1 (c 1.0, CHCl3);
1H NMR (400 MHz, CDCl3): 4.91 (1H, d, J = 6.5 Hz,
H-1III), 4.99 (1H, d, J = 8.0 Hz, H-1II), 5.20 (1H, d,
J = 2.4 Hz, H-1I); 13C NMR (100.6 MHz, CDCl3): d
14.7 (q, SCH2CH3), 25.2 (t, SCH2CH3), 61.9 (t, C-
5III), 63.3 (t, C-6II), 69.4 (t, C-6I), 69.6 (d, C-4III), 69.8
(d, C-4II), 70.9 (t, OCH2CH@CH2), 71.3 (2C, 2 ꢂ d,
C-2III, C-3III), 71.5 (d, C-5I), 71.9 (d, C-2II), 72.6 (d,
C-5II), 72.8 (t, CH2Ph), 73.1 (d, C-3II), 75.8 (d, C-4I),
76.7 (d, C-2I), 77.0 (d, C-3I), 81.3 (d, C-1), 100.4 (d,
C-1II), 100.7 (d, C-1III), 117.7 (t, OCH2CH@CH2),
127.4–130.0 (40C) (Ar-C), 133.0–133.6 (7C, 7 ꢂ s,
7 ꢂ COC6H5), 134.9 (d, OCH2CHCH2), 138.7 (s, Ar-
C), 164.9, 165.3 (2C), 165.6, 165.7, 166.0, 166.2 (7C,
7 ꢂ s, 4 ꢂ COC6H5); HRMS (ESI): [M+Na]+ calcd for
C52H52O14S, 955.2970; found, 955.2949.
A 60% oil dispersion of sodium hydride (160 mg,
4.00 mmol) was added at room temperature to a solu-
tion of 13 (360 mg, 0.40 mmol) and benzyl bromide
(480 lL, 4.00 mmol) in DMF (15 mL) and the reaction
mixture was stirred overnight. The progress of the reac-
tion was followed by TLC (toluene–EtOAc, 6:1). When
the reaction was complete, MeOH (1 mL) was carefully
added and the mixture was diluted with toluene
(100 mL), washed with water (2 ꢂ 50 mL), dried and
concentrated. Purification on a silica gel column (tolu-
ene–EtOAc, 25:1) gave 14 (530 mg, 92%); [a]D +24.0 (c
1.13. Ethyl b-D-glucopyranosyl-(1?2)-b-D-xylopyran-
osyl-(1?4)-3-O-allyl-6-O-benzyl-1-thio-a-D-manno-
pyranoside (12)
Compound 11 (860 mg, 0.62 mmol) was dissolved in dry
MeOH (48 mL), 1 M methanolic NaOMe (0.7 mL) was
added and the mixture was stirred overnight at room
temperature. The progress of the reaction was followed
by TLC (CH2Cl2–MeOH 9:1). Next, acetic acid (20
drops) was added, the stirring was continued for
1
1.0, CHCl3); H NMR (400 MHz, CDCl3): 4.43 (1H,

J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
2207
d, J = 7.6 Hz, H-1III), 4.56 (1H, d, J = 7.2 Hz, H-1II),
5.51 (1H, s, H-1I); 13C NMR (100.6 MHz, CDCl3):
d 15.2 (q, SCH2CH3), 25.7 (t, SCH2CH3), 63.2 (t,
C-5III), 63.8 (t, C-6II), 69.1 (t, C-6I), 70.8 (t,
OCH2CH@CH2), 72.0 (d), 73.0 (t, CH2Ph), 73.2 (t,
CH2Ph), 74.1 (t, CH2Ph), 75.0 (t, CH2Ph), 75.1 (t,
CH2Ph), 75.2 (d), 75.3 (d), 75.6 (t, CH2Ph), 75.8 (t,
CH2Ph), 75.8 (d, C-3I), 76.0 (d, C-2I), 78.0 (d, C-3III),
78.3 (d), 81.7 (d), 81.9 (d, C-1I), 82.5 (d, C-2III), 84.3
(d), 85.1 (d), 86.7 (s, CPh3), 101.9 (d, C-1II), 103.4 (d,
C-1III), 117.4 (t, OCH2CH@CH2), 126.3–129.2 (50C,
50 ꢂ d) (Ar-C), 135.1 (d, OCH2CHCH2), 137.9 (s),
138.4 (s), 138.5 (s), 138.8 (2C, 2 ꢂ s), 138.9 (2C, 2 ꢂ s),
144.0 (3C, 3 ꢂ s) (Ar-C); HRMS (ESI): [M+Na]+ calcd
for C48H58O14S, 913.3439; found, 913.3473.
EtOAc, 6:1). Then the mixture was diluted with CH2Cl2
(20 mL), washed with water (10 mL), dried, concentrated
and purified by silica gel chromatography (hexane–
EtOAc–HOAc, 14:7:1) to yield 16 (30 mg, 83%); [a]D
1
+46.3 (c 0.5, CHCl3); H NMR (400 MHz, CDCl3): d
4.42 (1H, d, J = 7.6 Hz, H-1III), 4.65 (1H, d,
J = 8.0 Hz, H-1II), 5.35 (1H, s, H-1I); 13C NMR
(100.6 MHz, CDCl3): d 15.0 (q, SCH2CH3), 25.5 (t,
SCH2CH3), 63.8 (t, C-5III), 68.6 (t, C-6I), 72.3 (t,
OCH2CH@CH2), 72.2 (d, C-5I), 73.1 (2C, 2 ꢂ t,
2 ꢂ CH2Ph), 74.4 (t, CH2Ph), 75.1 (2C, 2 ꢂ t,
2 x CH2Ph), 75.2 (t, CH2Ph), 75.6 (t, CH2Ph), 76.7 (d,
C-4I), 76.7 (d, C-4III), 77.0 (d), 78.3 (d, C-3I), 78.8 (d),
79.6 (d, C-4II), 80.3 (d, C-2II), 82.4 (d, C-1I), 82.4 (d,
C-2III), 83.1 (d, C-3II), 84.2 (d, C-3III), 102.9 (d, C-1II),
103.5 (d, C-1III), 117.8 (t, OCH2CH@CH2), 128.8–
127.5 (35C, 35 ꢂ d) (Ar-C), 134.7 (d, OCH2CHCH2),
137.6 (s), 138.2 (s), 138.3 (2C, 2 ꢂ s), 138.4 (s), 138.7
(s), 138.8 (s) (Ar-C), 170.2 (s, C-6II); HRMS (ESI):
[M+Na]+ calcd for C71H78O15S, 1225.4954; found,
1225.4962.
1.15. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyl-(1?2)-
2,3,4-tri-O-benzyl-b-D-xylopyranosyl-(1?4)-3-O-allyl-6-
O-benzyl-1-thio-a-D-mannopyranoside (15)
Compound 14 (60 mg, 0.04 mmol) was dissolved in
CHCl3–MeOH (2:1, 6 mL), p-TsOH (2 mg, 0.01 mmol)
was added and the mixture was stirred at room temper-
ature overnight. The progress of the reaction was fol-
lowed by TLC (toluene–EtOAc, 6:1). Then the mixture
was diluted with CHCl3 (10 mL), washed with saturated
solution of NaHCO3 (5 mL), water (10 mL), dried, con-
centrated and purified by silica gel chromatography (tol-
uene–EtOAc, 10:1) to yield 15 (36 mg, 72%); [a]D +31.8
(c 1.0, CHCl3); 1H NMR (400 MHz, CDCl3): d 4.43
(1H, d, J = 7.6 Hz, H-1III), 4.51 (1H, d, J = 7.6 Hz, H-
1II), 5.38 (1H, s, H-1I); 13C NMR (100.6 MHz, CDCl3):
d 15.1 (q, SCH2CH3), 25.6 (t, SCH2CH3), 62.3 (t,
C-5III), 63.9 (t, C-6II), 69.0 (t, C-6I), 71.8 (t,
OCH2CH@CH2), 72.2 (d), 73.0 (d), 73.1 (t, CH2Ph),
73.3 (t, CH2Ph), 75.1 (t, CH2Ph), 75.2 (2C, 2 ꢂ t,
2 ꢂ CH2Ph), 75.5 (d, C-5I), 75.6 (t, CH2Ph), 75.8 (t,
CH2Ph), 77.2 (d, C-3I), 77.6 (d), 78.0 (d, C-2I), 78.3 (d,
C-4III), 81.8 (d), 82.4 (d, C-1I), 82.4 (d, C-2III), 84.3 (d,
C-3III), 84.7 (d), 103.2 (d, C-1II), 103.8 (d, C-1III),
116.7 (t, OCH2CH@CH2), 126.4–130.6 (35C, 35 ꢂ d)
(Ar-C), 135.5 (d, OCH2CHCH2), 138.1 (s), 138.4 (2C,
2 ꢂ s), 138.6 (s), 138.7 (s), 138.8 (2C, 2 ꢂ s) (Ar-C);
HRMS (ESI): [M+Na]+, calcd for C90H94O14S,
1453.6256; found 1453.6279.
1.17. Ethyl benzyl (2,3,4-tri-O-benzyl-b-D-glucopyran-
osyl)uronate-(1?2)-2,3,4-tri-O-benzyl-b-D-xylopyran-
osyl-(1?4)-3-O-allyl-6-O-benzyl-1-thio-a-D-manno-
pyranoside (17)
A solution of 16 (30 mg, 0.025 mmol) in EtOAc (6 mL)
was titrated with PhCHN2 (0.3 M solution in Et2O,
3 mL, 0.9 mmol) at room temperature until full conver-
sion was observed by TLC (toluene–EtOAc, 6:1). After
2 h, the reaction was concentrated and purified by silica
gel chromatography (toluene–EtOAc, 20:1) to yield 17
1
(16 mg, 49%); [a]D +37.9 (c 1.0, CHCl3), lit.2 +27; H
NMR (400 MHz, CDCl3): d 4.44 (1H, d, J = 8.0 Hz,
H-1III), 4.47 (1H, d, J = 8.8 Hz, H-1II), 5.20 (1H, d,
J = 1.6 Hz, H-1I); 13C NMR (100.6 MHz, CDCl3): d
15.1 (q, SCH2CH3), 25.7 (t, SCH2CH3), 63.9 (t,
C-5III), 67.4 (t, CH2Ph), 69.1 (t, C-6I), 70.8 (t,
OCH2CH@CH2), 71.9 (d), 73.0 (t, CH2Ph), 73.2 (t,
CH2Ph), 74.6 (d, C-4I), 74.9 (d), 75.1 (t, CH2Ph), 75.2
(2C, 2 ꢂ t, 2 ꢂ CH2Ph), 75.6 (t, CH2Ph), 75.8 (t,
CH2Ph), 76.4 (d, C-3I), 76.7 (d, C-2I), 78.4 (d), 78.9
(d), 81.1 (d), 82.0 (d, C-1I), 82.5 (d, C-2III), 84.2 (d),
84.4 (d, C-3III), 102.7 (d, C-1II), 103.4 (d, C-1III), 117.1
(t, OCH2CH@CH2), 129.3–127.4 (40C, 40 ꢂ d) (Ar-C),
135.3 (d, OCH2CHCH2), 137.9 (s), 138.1 (s), 138.4 (s),
138.5 (2C, 2 ꢂ s), 138.7 (s), 138.8 (s), 138.9 (s) (Ar-C),
168.0 (s, C-6II).
1.16. Ethyl 2,3,4-tri-O-benzyl-b-D-glucopyranosyluronic
acid-(1?2)-2,3,4-tri-O-benzyl-b-D-xylopyranosyl-(1?4)-
3-O-allyl-6-O-benzyl-1-thio-a-D-mannopyranoside (16)
To a mixture of 15 (36 mg, 0.03 mmol) in CH2Cl2 (2 mL)
and water (1 mL), BAIB (24 mg, 0.08 mmol) and TEM-
PO (5 mg, 0.03 mmol) were added and the mixture was
stirred at room temperature. The same amount of BAIB
and TEMPO was added after 2 h, and after 4 h. The pro-
gress of the reaction was followed by TLC (toluene–
Acknowledgements
We thank the Swedish Research Council and EU
(MRTN-CT-2004-005645 GlycoGold) for financial
support.

2208
J. Vesely et al. / Carbohydrate Research 343 (2008) 2200–2208
6. Oscarson, S. Glycosylation Methods. Thioglycosides. In
Oligosaccharides in Chemistry and Biology: A Comprehen-
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