A new, improved synthesis of the trisaccharide repeating unit of the O-antigen from Xanthomonas campestris pv. campestris 8004

Article abstract of DOI:10.1016/j.tet.2008.01.093

Recent studies have revealed that lipid-A and core fragments of the lipopolysaccharide from Xanthomonas campestris pv. campestris 8004 (Xcc), a phytopathogenic Gram-negative bacterium, are able to elicit plant immunity with two independent mechanisms. To

Full text of DOI:10.1016/j.tet.2008.01.093

Available online at www.sciencedirect.com
Tetrahedron 64 (2008) 3381e3391
www.elsevier.com/locate/tet
A new, improved synthesis of the trisaccharide repeating unit of the
O-antigen from Xanthomonas campestris pv. campestris 8004
*
Daniela Comegna, Emiliano Bedini , Michelangelo Parrilli
Dipartimento di Chimica Organica e Biochimica, Universita` di Napoli ‘Federico II’, Complesso Universitario Monte Santangelo,
Via Cintia 4, 80126 Napoli, Italy
Received 15 November 2007; received in revised form 7 January 2008; accepted 24 January 2008
Available online 31 January 2008
Dedicated to Professor Matteo Adinolfi on the occasion of his 70th birthday
Abstract
Recent studies have revealed that lipid-A and core fragments of the lipopolysaccharide from Xanthomonas campestris pv. campestris 8004
(Xcc), a phytopathogenic Gram-negative bacterium, are able to elicit plant immunity with two independent mechanisms. To date, nothing is
known about the effect of the O-antigen portion. Since its separation from the core region by selective chemical degradation is very difficult,
the chemical synthesis of related oligosaccharides is strictly necessary. In this paper a new, improved synthesis of the O-antigen repeating unit is
presented. The main improvements in the synthesis are: (1) a shorter, high-yielding preparation of an efficient glycosyl donor of the rare sugar
3-acetamido-3,6-dideoxy-D-galactopyranose (3-acetamido-D-fucose, D-Fucp3NAc); (2) a new protecting group pattern, which is demonstrated to
open a path to the future synthesis of higher oligomers.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: 3-Acetamido-D-fucose; D-Rhamnose; Glycosylation; Xanthomonas
1. Introduction
that the recognition mechanism works analogously to the in-
nate immunity system of animals, which is based on the per-
ception of pathogen-associated molecular patterns (PAMPs),3
characteristic structures of the pathogen indispensable for its
growth within the host.4 Since LPSs cover a wide part of the
outer bacterial membrane, they might be a group of general
elicitors that can be recognized by plants to trigger a defence
response. Actually, it was shown that the lipid-A moiety acts
as PAMP in plants;5 recently, the core moiety was recognized
to elicit plant innate immunity too. This study revealed that,
when Arapidopsis thaliana leaves were inoculated with the
core or the lipid-A from the phytopathogenic bacterium Xan-
thomonas campestris pv. campestris 8004 (Xcc), a noteworthy
up-regulation of defense-related genes was observed. In addi-
tion, the transcript levels showed an accumulation after 12 h in
response to the core and after 24 h in response to the lipid-A,
which highly resembled the values obtained in an experiment
where the leaves were inoculated with the whole lipooligosac-
charide.6 These data strongly suggested for the first time that
The principal components of the Gram-negative outer
membrane cell surface are the lipopolysaccharides (LPSs),
amphiphilic macromolecules usually consisting of three differ-
ent domains: a lipid part (lipid-A), an oligosaccharide region
(core) and a long-chain polysaccharide portion (O-chain, O-
antigen). LPSs are highly involved in bacterial pathogenesis
both in animals and plants.
The mechanism of interaction between eukaryotic cells and
Gram-negative bacteria, which are pathogenic to humans and
animals, has been studied in great extent,1 whereas only a few,
non-exhaustive studies have been accomplished about plante
bacteria interactions.2 Indeed, to date, very little is known
about LPS perception by plants. It has been suggested recently
* Corresponding author. Tel.: þ39 81 674153; fax: þ39 81 674393.
E-mail address: ebedini@unina.it (E. Bedini).
0040-4020/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.01.093

3382
D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
two distinct and independent mechanisms are involved in the
perception of the core and lipid-A, even if the receptor compo-
nents involved in recognition and transmembrane signalling
still remain unknown.7 The role of the O-antigen portion of
the Xcc lipopolysaccharide was not elucidated since its separa-
tion from the core region by selective chemical degradation is
very difficult. The chemical structure of this O-antigen con-
sists of a trisaccharide repeating unit, that has a D-rhamnose
disaccharide backbone with a 3-acetamido-3,6-dideoxy-D-gal-
actopyranose (3-acetamido-D-fucose, D-Fucp3NAc) as branch
(Fig. 1).8
yield. Therefore, we looked for a high-yielding synthesis of a
new, more efficient D-Fucp3NAc donor.
Firstly, we planned to perform the a,b-epoxytrichloroacet-
imidate intramolecular cyclization on a compound with a b-
configured thioalkyl group already installed at the anomeric
position, to avoid the low b-stereoselectivity in the conversion
of hemiacetal 12 to thioglycoside 5 (Scheme 2, route A).13
Thus, b-thioglycoside 13 was obtained in high yield (78%)
and excellent stereoselectivity (b/a¼19:1) from D-fucose via
a thiouronium intermediate generated from fucosyl iodide.15
Compound 13 was then deacetylated and regioselectively pro-
´
We have recently demonstrated that synthetic oligo-
rhamnans, which resemble the most general structure of
O-antigen backbones from phytopathogenic bacteria,9 are
recognized by plant cells and elicit plant innate immunity.10
From this result one could hypothesize that the ‘rhamnose-
rich’ Xcc O-antigen could be involved in the mechanisms of
elicitation of plant innate immunity. This could be proved by
synthesizing the repeating unit of this O-antigen as well as
its higher oligomers and then testing them in Arabidopsis
thaliana defense-related genes up-regulation experiments. En
route to this goal, we present here a new synthesis of the tri-
saccharide repeating unit. The synthetic strategy was carefully
planned in order to use a protecting group pattern, which will
enable the oligomerization of the repeating unit.
tected at positions O-2 and O-4 in four steps one-pot (Zemplen
deacylation, orthoesterification, acetylation, orthoester hydro-
lysis) to afford alcohol 14 (71% over four steps from 13)
(Scheme 2, route B). Unfortunately, treatment of triflate deriv-
ative 15 with sodium methoxide did not afford the desired 2,3-
epoxide derivative 16. Instead a complex mixture was obtained
probably due to the trans-configuration of a good leaving group
at position O-2 and the adjacent thioalkyl moiety that was prone
to give an intramolecular displacement of the epoxide.16 This
generated a glycosyl oxonium ion, whose uncontrollable reac-
tivity could explain the complex mixture of products obtained
in this reaction. Alternatively, the introduction of a nitrogen
function at position O-3 was planned via a Lattrell-Dax17,18 ep-
imerization followed by an SN2 azide displacement. However,
the treatment of triflate 15 with KNO2 in strictly anhydrous
DMF at 50 ꢀC19 afforded the desired 6-deoxy-gulose (antiar-
ose) derivative in only moderate yield (57%); therefore, this
route was not further developed. Inspired by the recent synthe-
sis of ravidosamine (L-Fucp3NMe2)20 and by the synthesis of
the unnatural 3-acetamido-3,6-dideoxy-L-galactopyranose (L-
Fucp3NAc),21 we planned to insert the nitrogen function at
position O-3 by oxime reduction. Thus, thioglycoside 13 was
deacetylated and the resulting triol was regioselectively sily-
lated at position O-322 to give diol 17 (79%) (Scheme 2, route
C). Benzylation (93%) and subsequent cleavage of TBS group
(98%) afforded 19 that gave in turn compound 20 (78%) after
oxidation of the alcohol and oximation of the resulting ketone.
Several reduction conditions were screened on oxime 20.
Red-AlÒ in THF at 0 ꢀC was found to give best yield and stereo-
selectivity, affording the desired galacto-configured amine 21
together with the gulo-configured compound 22 in 88% yield
(21/22¼10:1). After separation of the two epimers by column
chromatography, the amino moiety of 21 was protected as
Troc to give the D-Fucp3NAc donor 6. The overall yield of
the new synthetic path (route C) was 27% over 11 steps, which
was much better than the former one (route A, 3% over 15
steps). Besides, the coupling of the new donor 6 with the rham-
nose disaccharide 23 proceeded smoothly and in better yield
than coupling of donor 5 with acceptor 23 (Scheme 3). Actually,
in the latter case trisaccharide 3 was obtained in modest yield
(40%) together with unreacted acceptor recovered in 27%
yield: the separation of disaccharide 23 and trisaccharide 3
was not easily achievable and required HPLC.13 Difficulties
in introducing an amino-fucose residue on a sterically crowded
acceptor was already reported in literature.23 Nevertheless the
coupling between the new D-Fucp3NAc donor 6 and 23 under
2. Results and discussion
The chemical synthesis of the Xcc O-antigen trisaccharide
repeating unit was challenging owing to two features: (1) its
monosaccharide constituentsdD-rhamnose and D-Fucp3NAcd
are rather unusual and therefore not commercially available:
they had to be synthesized from other sugar precursors; (2)
two of the three linkages present in the repeating unit are
cis-configured and one of these has a b-manno configuration,
which is the most difficult to synthesize stereoselectively.11,12
The synthesis of this trisaccharide repeating unit as the methyl
glycoside 1 was already reported by us (Scheme 1).13 Unfor-
tunately, the overall yield of the synthesis was not high enough
to allow further manipulation of the trisaccharide towards
higher oligomers. Actually, the ‘Achilles heel’ of that synthe-
sis was the overly long and low-yielding path (3% overall yield)
for the conversion of commercially available D-fucose into the
D-Fucp3NAc donor 5. The synthetic strategy required 15 reac-
tions, with the intramolecular cyclization of the a,b-epoxytri-
chloroacetimidate 10 as key step for the introduction of the
nitrogen moiety at position O-3 (Scheme 2, route A).14 Besides,
the coupling of 5 with the sterically crowded D-rhamnose disac-
charide 23 (see Scheme 3) gave trisaccharide 3 in only 40%
3)-β-D-Rhap-(1 3)-α-D-Rhap-(1
2
1
α-D-Fucp3NAc
Figure 1. Repeating unit of the O-antigen from Xanthomonas campestris pv.
campestris 8004.

D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
3383
Scheme 1. Former13 and new retrosynthetic analysis for the target trisaccharide.
NIS/AgOTf activation at ꢁ20 ꢀC in 1:1 v/v CH2Cl2/Et2O
proceeded in good yield (77%) and pure trisaccharide 24
was obtained simply by column chromatography. The a-
A further goal was to open a synthetic way towards O-an-
tigen higher oligomers. For this purpose, we had to test if the
trisaccharide building block 24 could be transformed into both
a glycosyl donor and a glycosyl acceptor. As first step towards
a glycosyl donor, we planned the conversion of the anomeric
methoxy group into an acetate by acetolysis. Even when it
was conducted in mild conditions (1:1:0.05 v/v/v AcOH/
Ac2O/TFA at 70 ꢀC) as recently reported on a disaccharide
case,24 a complex mixture of mono-, di- and trisaccharides
configuration of the new glycosidic bond was ascertained by
1
a JH-1 H-2
,
value of 3.3 Hz, typical of a 1,2-cis-configuration
in the case of a galacto-configured residue. In conclusion,
a new, high-yielding synthetic path to a protected form of the
trisaccharide repeating unit of the Xcc O-antigen has been
accomplished.
Scheme 2. Synthesis of D-Fucp3NAc donors 513 and 6. Reagents and conditions: i: (a) Ac2O, py, rt, (b) I2, Et3SiH, CH2Cl2, reflux, (c) thiourea, CH3CN, 60 ꢀC then
EtI, Et3N, rt, 74% over three steps; ii: (a) t-BuOK, MeOH, 0 ꢀC, (b) CSA, 7:2 v/v MeC(OMe)3/DMF, 100 mbar, rt, (c) Ac2O, py, rt, (d) 80% AcOH, rt, 82% over
four steps; iii: Tf2O, 1:1 v/v py/CH2Cl2, 0 ꢀC; iv: Na, 1:1 v/v MeOH/CH2Cl2, 0 ꢀC; v: (a) t-BuOK, MeOH, 0 ꢀC, (b) TBSCl, ImH, DMF, 0 ꢀC, 79% over two steps;
vi: BnBr, NaH, DMF, 0 ꢀC, 93%; vii: TBAF, THF, rt, 98%; viii: (a) 2:1 v/v DMSO/Ac2O, rt, (b) NH2OH$HCl, NaOAc, MeOH, rt, 78% over two steps; ix: Red-
AlÒ, THF, 0 ꢀC, 21: 80%, 22: 8%; x: TrocCl, py, 82%.

3384
D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
OBn
O
TrocHN
BnO
BnO
BnO
HO
O
O
O
O
O
6, ii.
5, i.
BnO
BnO
AllO
BnO
BnO
AllO
3
O
O
OMe
OMe
23
24
Scheme 3. Glycosylation reactions of D-Fucp3NAc donors 513 and 6 with rhamnose disaccharide 23. Reagents and conditions: i: 40%: see Ref. 13; ii: NIS, AgOTf,
ꢀ
˚
AW-300 4 A MS, 1:1 v/v CH2Cl2/Et2O, ꢁ20 C, 77%.
was obtained, due to the simultaneous cleavage of the intergly-
cosidic bonds. The explanation is that in our case both the two
glycosidic linkages and the methoxy group are rather activated
towards acid-mediated cleavage due to the electron-donating
protecting groups at position O-2 of all three residues. The
mild acetolysis reaction was unable to select between the three
similarly activated linkages; a non-regioselective cleavage was
observed.25 At this point, the synthesis of a new trisaccharide
building block, possessing an orthogonal protecting group at
the anomeric position, was mandatory. We chose a p-methoxy-
phenyl (MP) as new anomeric protecting group: its use as
an effective anomeric protecting group in D-rhamnose oligo-
saccharide synthesis was very recently reported.26,27 Thus,
the new D-rhamnose acceptor 8 was obtained from the
former one 713 by acetolysis, p-methoxyphenylation and Zem-
´
plen deacetylation (62% yield over three steps) (Scheme 4). N-
Phenyl trifluoroacetimidate 913 was used as D-rhamnose donor
to have a b>a stereoselectivity in the glycosylation, since the
benzylsulfonyl group at 2-O-atom was already exploited in
b-stereoselective mannosylations.28 Unfortunately, the coupling
between 8 and 9 afforded disaccharide 25 in high yield
(88%) but without the desired stereoselectivity (a/b¼3:2).
Since the coupling between 9 and the former acceptor 7
gave disaccharide 26 in quantitative yield (99%) and higher
b-stereoselectivity,13 we decided to insert the MP group at
the ‘disaccharide level’. Mild acetolysis of 26b proceeded
smoothly to give the disaccharide with an acetyl group at
the anomeric position. This result was explained by the strong
BnO
BnO
BnO2SO
BnO
O
O
O
i.
9, ii.
O
BnO
HO
BnO
HO
BnO
AllO
BnO
O
OMe
7
8
OMP
25
OMP
OMP
9, ii.
BnO2SO
BnO
HO
BnO
O
O
O
O
iii.
iv.
BnO
AllO
BnO
27
BnO
AllO
BnO
25
O
O
26
OMe
6, v.
OBn
TrocHN
OBn
TrocHN
O
O
BnO
BnO
vi.
BnO
BnO
O
O
O
O
O
O
BnO
O
BnO
BnO
AllO
BnO
AllO
O
OH
OMP
29
4
vii.
OBn
TrocHN
O
BnO
viii.
2
BnO
O
O
O
BnO
BnO
HO
O
OMP
28
Scheme 4. Completion of the synthesis. Reagents and conditions: i: (a) 1:1:0.1 v/v/v Ac2O/AcOH/TFA, 70 ꢀC, (b) p-methoxyphenol, BF3$OEt2, CH2Cl2, 0 ꢀC, (c)
ꢀ
ꢀ
˚
t-BuOK, MeOH, rt, 62% over three steps; ii: TMSOTf, AW-300 4 A MS, CH2Cl2, ꢁ50 C to ꢁ25 C, 25: 88% (a/b¼3:2), 26 (see Ref. 13): 99% (a/b¼2:3); iii: (a)
1:1:0.1 v/v/v Ac2O/AcOH/TFA, 70 ꢀC, (b) p-methoxyphenol, BF3$OEt2, CH2Cl2, 0 ꢀC, 65% over two steps; iv: NaNH2, DMF, rt, 52%; v: NIS, AgOTf, AW-300
ꢀ
˚
4 A MS, 1:1 v/v CH2Cl2/Et2O, ꢁ20 C, 65%; vi: CAN, 2:2:1 v/v/v toluene/CH3CN/H2O, rt, 68% (a/b¼4.5:1); vii: PdCl2, 2:1 v/v MeOH/CH2Cl2, rt, 86%; viii: (a)
Zn/Cu, 2:1 v/v AcOH, Ac2O, rt, (b) Pd/C, 9:1 v/v MeOH/HCOOH, ultrasound bath, 55% over two steps.

D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
3385
electron-withdrawing character of the benzylsulfonyl group,
which disfavoured the acid-catalyzed cleavage of the intergly-
cosidic bond; on the contrary the electron-donating benzyl
group at position O-2A activated the anomeric methoxy group
towards acetolysis.25 After subsequent p-methoxyphenylation,
disaccharide 25b was obtained in good yield (65% after two
steps). Cleavage of the benzylsulfonyl group on 25b with so-
dium amide in DMF for 48 h proceeded in 52% yield. At-
tempts to shorten the reaction time by using microwave
irradiation29 considerably lowered the yield. In this case, b-
elimination of PhCH2SO2H competed with benzylsulfonyl
cleavage, as observed by MALDI mass spectrum of the crude
reaction mixture. Disaccharide acceptor 27 was coupled with
donor 6 under reaction conditions already used for the synthe-
sis of 24. Glycosylation proceeded uneventfully to give trisac-
higher oligomers of the repeating unit of Xcc O-antigen. To
this purpose, work is currently underway.
4. Experimental section
4.1. General methods
1H and 13C NMR spectra were recorded on Varian XL-200
(1H: 200 MHz, 13C: 50 MHz), Varian Gemini-300 (1H:
300 MHz, 13C: 75 MHz), Bruker DRX-400 (1H: 400 MHz,
13C: 100 MHz) or Varian INOVA 500 (1H: 300 MHz, 13C:
125 MHz) instruments in CDCl3 (CHCl3 as internal standard,
1H: CHCl3 at d 7.26; 13C: CDCl3 at d 77.0) and in D2O (ace-
tone as internal standard, 1H: (CH3)2CO at d 2.22; 13C:
(CH3)2CO at d 31.5). Assignment of proton chemical shifts
were based on 1D HOHAHA and COSY experiments. Assign-
ment of proton and carbon chemical shifts of the deprotected
trisaccharide 2 was based on 2D NMR experiments such as
COSY, TOCSY and HSQC. Positive ESI-MS spectra were re-
corded on a Finnigan LCQ-DECA ion trap mass spectrometer.
Positive MALDI-MS spectra were recorded on a Applied Bio-
system Voyager DE-PRO MALDI-TOF mass spectrometer in
the positive mode: compounds were dissolved in the appropriate
solvent at a concentration of 1 mg/mL and 1 mL of these solu-
tions were mixed with 1 mL of a 20 mg/mL solution of 2,5-dihy-
droxybenzoic acid in 7:3 CH3CN/water. Optical rotations were
measured on a JASCO P-1010 polarimeter at 294 K. Analytical
thin layer chromatography (TLC) was performed on aluminium
plates precoated with Merck Silica Gel 60 F254 as the adsorbent.
The plates were developed with 5% H2SO4 ethanolic solution
and then heated to 130 ꢀC. Column chromatography was per-
formed on Merck Kieselgel 60 (63e200 mesh). Gel-filtration
chromatography was performed on a Sephadex G-10 column
(2.0ꢂ90 cm) with water as eluant.
charide 4 in 65% yield. The a-configuration of the new
1
glycosidic bond was ascertained by a JH-1 H-2
,
value of
3.2 Hz, typical of a 1,2-cis-configuration in the case of a gal-
acto-configured residue. The protecting group pattern on 4 was
demonstrated to be orthogonal enough to open a path towards
higher oligomers of the repeating unit of Xcc O-antigen. Actu-
ally, compound 4 was smoothly transformed into trisaccharide
acceptor 28 (86%) by chemoselective de-O-allylation with
PdCl2 in 2:1 v/v MeOH/CH2Cl2. Selective deprotection of
the anomeric MP group was accomplished by treating 4
with CAN in 2:2:1 toluene/CH3CN/water: hemiacetal 29 was
obtained in 68% yield (a/b¼4.5:1) and can be activated as
a trisaccharide donor. The optimization of the [3þ3] glycosyl-
ation is currently underway and the synthesis of higher oligo-
mers of the repeating unit of Xcc O-antigen will be reported
elsewhere. Finally, global deprotection of trisaccharide 28
was accomplished in two steps: firstly, the NHTroc group
was converted into an acetamido group by treatment with
Zn/Cu in 2:1 v/v AcOH/Ac2O, then de-O-benzylation was per-
formed by transfer hydrogenation under Perlin conditions30 to
give deprotected trisaccharide 2. 1H and 13C NMR data of the
target compound 2 are reported in Table 1.
4.1.1. Ethyl 2,3,4-tri-O-acetyl-1-thio-b-D-fucopyranoside31
(13)
A solution of D-fucose (2.5 g, 15.3 mmol) in 1:1 v/v Ac2O/
pyridine (20 mL) was stirred overnight, after that, it was di-
luted with CH2Cl2 (200 mL) and washed with 1 M HCl
(200 mL), 1 M NaHCO3 (200 mL) and water (200 mL). The
organic layer was collected, dried over anhydrous Na2SO4, fil-
tered and concentrated. The obtained residue was dissolved in
CH2Cl2 (20 mL) and treated with iodine (5.5 g, 21.7 mmol)
and Et3SiH (3.47 mL, 21.7 mmol). The mixture was heated
at reflux for few minutes, then diluted with CH2Cl2
(200 mL) and washed with 1:1 v/v 1 M NaHCO3/10%
Na2S2O3 (200 mL) and water (200 mL). The organic layer
was collected, dried over anhydrous Na2SO4, filtered and
3. Conclusions
In conclusion, a new, improved synthesis of the trisaccha-
ride repeating unit of the O-antigen from Xanthomonas cam-
pestris pv. campestris 8004 was accomplished. The main
improvements are: (1) a shorter, high-yielding synthesis of
a very efficient D-Fucp3NAc donor; (2) the choice of a new
protecting group pattern, which was demonstrated to be or-
thogonal enough to open a path to both a trisaccharide accep-
tor and a trisaccharide donor. This will enable the synthesis of
Table 1
1H and 13C NMR (italic) chemical shifts (d in ppm) of target trisaccharide 2
Sugar residue
H-1/C-1
H-2/C-2
H-3/C-3
H-4/C-4
H-5/C-5
H-6/C-6
A
B
C
5.57/100.1
4.96/98.1
5.28/101.1
4.44/68.1
4.24/79.1
3.94/69.5
4.22/79.1
3.78/74.2
4.36/52.2
3.68/71.4
3.58/74.3
3.82/71.2
3.94/69.3
3.54/73.5
4.64/68.1
1.32/18.8
1.44/18.7
1.28/18.0

3386
D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
concentrated. The residue was mixed with thiourea (1.75 g,
23.0 mmol), suspended in CH3CN (20 mL) and then heated
to 60 ꢀC. After 20 min the reaction mixture was cooled to rt,
and EtI (2.5 mL, 31.3 mmol) and Et3N (8.5 mL, 61.0 mmol)
were added. The mixture was stirred for additional 20 min
and then concentrated. The residue was subjected to column
chromatography (15e30% ethyl acetate in petroleum ether)
to give 1331 (3.99 g, 74%). [a]D21 ꢁ30 (c 0.5, CH2Cl2) [lit.:31
and TBSCl (2.25 g, 14.8 mmol). The mixture was stirred at
0 ꢀC for 30 min and then diluted with ethyl acetate
(200 mL) and washed with water (200 mL). The organic layer
was collected, dried over anhydrous Na2SO4, filtered and con-
centrated to afford a residue that was subjected to a column
chromatography (10e20% ethyl acetate in petroleum ether)
to give 17 (2.888 g, 79%) as a yellowish oil. [a]2D1 ꢁ13 (c
1.0, CH2Cl2). 1H NMR (CDCl3, 200 MHz) d 4.21 (d,
J1,2¼8.4 Hz, 1H, H-1), 3.66e3.50 (m, 4H, H-2, H-3, H-4,
H-5), 2.81e2.64 (m, 2H, SCH2CH3), 2.55 (br s, 1H, OH),
2.36 (br s, 1H, OH), 1.32 (d, J6,5¼6.6 Hz, 3H, H-6), 1.26 (t,
Jvic¼6.8 Hz, 3H, SCH2CH3), 0.87 (s, 9H, SiC(CH3)3), 0.12
(s, 3H, SiCH3), 0.10 (s, 3H, SiCH3); 13C NMR (CDCl3,
50 MHz) d 85.8 (C1), 75.9, 74.2, 72.3, 69.7 (C2, C3, C4, C5),
25.6 (SiC(CH3)3), 23.9 (SCH2CH3), 18.0 (C6), 15.1
(SCH2CH3), ꢁ4.5, ꢁ5.1 (Si(CH3)2). ESI-MS for C14H30O4SSi
(m/z): Mr (calcd) 322.16, Mr (found) 345.33 (MþNa)þ. Anal.
Calcd: C 52.13, H 9.38. Found: C 52.26, H 9.28.
1
ꢁ28 (c 1, CHCl3)]. H NMR (CDCl3, 300 MHz) d 5.28 (dd,
[a]2D0 J4,3¼3.3 Hz, J4,5¼1.2 Hz, 1H, H-4), 5.22 (t, J2,3¼J2,1
¼
9.9 Hz, 1H, H-2), 5.06 (dd, J3,2¼9.9 Hz, J3,4¼3.3 Hz, 1H, H-
3), 4.49 (d, J1,2¼9.9 Hz, 1H, H-1), 3.86 (dq, J5,6¼6.6 Hz,
J5,4¼1.2 Hz, 1H, H-5), 2.81e2.67 (m, 2H, SCH2CH3), 2.19,
2.08, 1.99 (3s, 9H, 3Ac), 1.29 (t, Jvic¼7.5 Hz, 3H, SCH2CH3),
1.23 (d, J6,5¼6.6 Hz, 3H, H-6); 13C NMR (CDCl3, 75 MHz)
d 170.4, 169.8, 169.4 (3CO), 83.2 (C1), 72.9, 72.1, 70.2, 67.1
(C2, C3, C4, C5), 23.8 (SCH2CH3), 20.5, 20.3 (2COCH3),
16.1 (C6), 14.5 (SCH2CH3). ESI-MS for C14H22O7S (m/z): Mr
(calcd) 334.11, Mr (found) 356.93 (MþNa)þ.
4.1.4. Ethyl 2,4-di-O-benzyl-3-O-tert-butyldimethylsilyl-1-
thio-b-D-fucopyranoside (18)
4.1.2. Ethyl 2,4-di-O-acetyl-1-thio-b-D-fucopyranoside (14)
A solution of 13 (0.512 g, 1.75 mmol) in MeOH (5.0 mL)
was treated at 0 ꢀC with t-BuOKuntil it was strongly basic. After
30 min stirring at rt, it was neutralized with Amberlyst-15 (Hþ),
then filtered and concentrated to give a residue that was dis-
solved in 7:2 v/v MeC(OMe)3/DMF (9.0 mL). CSA (80 mg,
0.34 mmol) was then added and the solution was evacuated at
100 mbar for 20 min, after that pyridine (7.0 mL) and Ac2O
(7.0 mL) were sequentially added. The solution was stirred
overnight at rt, then coevaporated four times with toluene
(10 mL each). The residue was dissolved in 80% AcOH
(10 mL) and the solution was stirred at rt for 10 min, after that
it was coevaporated two times with toluene (5 mL each). The
residue was subjected to column chromatography (40% ethyl
acetate in petroleum ether) to give 14 (0.893 g, 82%) as a white
A solution of 17 (2.888 g, 8.97 mmol) in DMF (10 mL) was
cooled to 0 ꢀC and then treated with BnBr (1.97 mL, 19.6 mmol)
and NaH (60% oil suspension; 1.808 g, 53.8 mmol). The mix-
turewas stirred at 0 ꢀC for 1 h, after that water (10 mL) was care-
fully added. The mixture was diluted with ethyl acetate
(200 mL) and washed with water (200 mL). The organic layer
was collected, dried over anhydrous Na2SO4, filtered and con-
centrated to afford a residue, that was chromatographed (4%
ethyl acetate in petroleum ether) to give 18 (4.188 g, 93%) as
white amorphous crystals. [a]2D1 ꢁ14 (c 0.8, CH2Cl2). 1H NMR
(CDCl3, 300 MHz) d 7.47e7.24 (m, 10H, H-Ar), 5.15 (d,
Jgem¼11.7, 1H, OCHHPh), 4.94 (d, Jgem¼10.5, 1H, OCHHPh),
4.80 (d, Jgem¼10.5, 1H, OCHHPh), 4.68 (d, Jgem¼11.7, 1H,
OCHHPh), 4.44 (d, J1,2¼9.0 Hz, 1H, H-1), 3.81 (dd,
J3,2¼9.3 Hz, J3,4¼3.0 Hz, 1H, H-3), 3.73 (t, J2,3¼J2,1¼9.3 Hz,
1H, H-2), 3.61 (q, J5,6¼6.6 Hz, 1H, H-5), 3.51 (d,
J4,3¼3.0 Hz, 1H, H-4), 2.86e2.66 (m, 2H, SCH2CH3), 1.34 (t,
Jvic¼7.5 Hz, 3H, SCH2CH3), 1.29 (d, J6,5¼6.6 Hz, 3H, H-6),
1.02 (s, 9H, SiC(CH3)3), 0.19 (s, 3H, SiCH3), 0.14 (s, 3H,
SiCH3); 13C NMR (CDCl3, 75 MHz) d 138.8, 138.3 (2Cipso),
128.3e127.2 (C-Ar), 84.8 (C1), 80.6, 78.4 (C2, C4), 77.7,
75.3, 75.1, 74.2 (C3, C5, 2OCH2Ph), 26.0 (SiC(CH3)3), 24.5
(SCH2CH3), 18.0 (C6), 14.9 (SCH2CH3), ꢁ4.1, ꢁ4.6
(Si(CH3)2). ESI-MS for C28H42O4SSi (m/z): Mr (calcd)
502.26, Mr (found) 525.41 (MþNa)þ. Anal. Calcd: C 66.89, H
8.42. Found: C 66.76, H 8.68.
solid. [a]2D1 ꢁ2.4 (c 1.0, CH2Cl2). Mp¼120.4e121.3 ꢀC. H
1
NMR (CDCl3, 200 MHz)
d
5.17 (dd, J4,3¼3.2 Hz,
J4,5¼0.8 Hz, 1H, H-4), 4.97 (t, J2,3¼J2,1¼9.6 Hz, 1H, H-2),
4.37 (d, J1,2¼10.0 Hz, 1H, H-1), 3.79 (dd, J3,2¼9.6 Hz,
J3,4¼3.2 Hz, 1H, H-3), 3.71 (dq, J5,6¼6.4 Hz, J5,4¼0.8 Hz,
1H, H-5), 2.76e2.58 (m, 2H, SCH2CH3), 2.14, 2.07 (2s, 6H,
2Ac), 1.23 (t, Jvic¼7.2 Hz, 3H, SCH2CH3), 1.16 (d,
J6,5¼6.4 Hz, 3H, H-6); 13C NMR (CDCl3, 50 MHz) d 171.3,
171.0 (2CO), 83.1 (C1), 73.4, 73.1, 72.4, 71.0 (C2, C3, C4, C5),
24.1 (SCH2CH3), 20.9, 20.8 (2COCH3), 16.6 (C6), 14.7
(SCH2CH3). ESI-MS for C12H20O6S (m/z): Mr (calcd) 292.10,
Mr (found) 315.21 (MþNa)þ. Anal. Calcd: C 49.30, H 6.90.
Found: C 49.35, H 6.85.
4.1.5. Ethyl 2,4-di-O-benzyl-1-thio-b-D-fucopyranoside (19)
A 1 M solution of TBAF in THF (20 mL, 20 mmol) was
added to a solution of 18 (3.500 g, 6.97 mmol) in THF
(20 mL), and the resulting solution was stirred at rt for 3 h.
After concentration, column chromatography (10e30% ethyl
acetate in petroleum ether) afforded 19 (2.650 g, 98%) as a
4.1.3. Ethyl 3-O-tert-butyldimethylsilyl-1-thio-b-D-
fucopyranoside (17)
To a solution of 13 (3.800 g, 11.4 mmol) in MeOH (25 mL)
at 0 ꢀC, t-BuOK was added until the solution was strongly
basic. After 30 min stirring at rt, the mixture was neutralized
with Amberlyst-15 (Hþ), then filtered and concentrated to
give a residue that was dissolved in DMF (7.0 mL), cooled
to 0 ꢀC and then treated with imidazole (2.04 g, 29.6 mmol)
1
yellowish oil. [a]D21 þ7 (c 0.7, CH2Cl2). H NMR (CDCl3,
300 MHz) d 7.49e7.25 (m, 10H, H-Ar), 5.01 (d, Jgem¼10.8,
1H, OCHHPh), 4.88e4.76 (m, 3H, 3OCHHPh), 4.44 (d,

D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
3387
J1,2¼9.0 Hz, 1H, H-1), 3.75 (dd, J3,2¼9.3 Hz, J3,4¼3.0 Hz,
1H, H-3), 3.63 (t, J2,3¼J2,1¼9.3 Hz, 1H, H-2), 3.57 (d,
J4,3¼3.0 Hz, 1H, H-4), 3.54 (q, J5,6¼6.6 Hz, 1H, H-5),
2.91e2.78 (m, 3H, SCH2CH3, OH), 1.39 (t, Jvic¼7.5 Hz,
3H, SCH2CH3), 1.33 (d, J6,5¼6.6 Hz, 3H, H-6); 13C NMR
(CDCl3, 75 MHz) d 138.1, 138.0 (2Cipso), 128.0e127.4 (C-
Ar), 84.3 (C1), 79.0, 78.9 (C2, C4), 75.5, 75.1, 74.9, 74.3
(C3, C5, 2OCH2Ph), 24.5 (SCH2CH3), 16.9 (C6), 14.7
(SCH2CH3). ESI-MS for C22H28O4S (m/z): Mr (calcd)
388.17, Mr (found) 411.38 (MþNa)þ. Anal. Calcd: C 68.01,
H 7.26. Found: C 68.36, H 7.20.
J5,4¼2.0 Hz, 1H, H-5), 3.66 (dd, J2,1¼9.6 Hz, J2,3¼3.6 Hz,
1H, H-2), 3.64 (dd, J4,3¼3.6 Hz, J4,5¼2.0 Hz, 1H, H-4), 3.47
(t, J3,4¼J3,2¼3.6 Hz, 1H, H-3), 2.77e2.66 (m, 2H, SCH2CH3),
1.28 (t, Jvic¼8.0 Hz, 3H, SCH2CH3), 1.20 (d, J6,5¼6.4 Hz, 3H,
H-6); 13C NMR (CDCl3, 75 MHz) d 138.3, 138.1 (2Cipso),
128.4e127.7 (C-Ar), 80.6, 80.0, 75.6, 72.6, 70.6, 62.7 (C1, C2,
C4, C5, 2OCH2Ph), 49.3 (C3), 24.4 (SCH2CH3), 16.5 (C6),
14.9 (SCH2CH3). ESI-MS for C22H29NO3S (m/z): Mr (calcd)
387.19, Mr (found) 410.41 (MþNa)þ. Anal. Calcd: C 68.18, H
7.54, N 3.61. Found: C 68.06, H 7.66, N 3.57.
Second eluted compound 21 (1.726 g, 80%) was recovered
as a yellowish oil. [a]2D1 þ18 (c 0.3, CH2Cl2). 1H NMR
(CDCl3, 300 MHz) d 7.40e7.28 (m, 10H, H-Ar), 5.04 (d,
Jgem¼10.8 Hz, 1H, OCHHPh), 4.76 (d, Jgem¼11.4 Hz, 1H,
OCHHPh), 4.65 (d, Jgem¼11.4 Hz, 1H, OCHHPh), 4.43 (d,
Jgem¼10.8 Hz, 1H, OCHHPh), 4.34 (d, J1,2¼9.6 Hz, 1H, H-
1), 3.61e3.57 (m, 2H, H-4, H-5), 3.30 (t, J2,3¼J2,1¼9.6 Hz,
1H, H-2), 2.84e2.69 (m, 3H, H-3, SCH2CH3), 1.33 (t,
Jvic¼7.5 Hz, 3H, SCH2CH3), 1.31 (d, J6,5¼6.3 Hz, 3H, H-6);
13C NMR (CDCl3, 75 MHz) d 138.2, 138.0 (2Cipso), 128.5e
127.7 (C-Ar), 85.5 (C1), 81.0, 80.5 (C2, C4), 76.1, 76.0, 75.3
(C5, 2OCH2Ph), 58.0 (C3), 24.9 (SCH2CH3), 17.2 (C6), 14.9
(SCH2CH3). ESI-MS for C22H29NO3S (m/z): Mr (calcd)
387.19, Mr (found) 410.29 (MþNa)þ. Anal. Calcd: C 68.18,
H 7.54, N 3.61. Found: C 68.00, H 7.58, N 3.55.
4.1.6. Ethyl 2,4-di-O-benzyl-6-deoxy-1-thio-b-D-xylo-hexo-
pyranosid-3-ulose (E)-oxime (20)
Alcohol 19 (2.796 g, 7.20 mmol) was dissolved in 2:1 v/v
DMSO/Ac2O (15 mL) and stirred at rt for 3 h. The solution
was then diluted with ethyl acetate (300 mL) and washed with
water (300 mL). The organic layer was collected, dried over an-
hydrous Na2SO4, filtered and concentrated to afford a residue
that was dissolved in MeOH (25 mL) and treated with NaOAc
(847 mg, 10.1 mmol) and NH2OH$HCl (750 mg, 10.1 mmol).
After stirring at rt for 2 h, the solution was concentrated and
the obtained residue was subjected to column chromatography
(10e30% ethyl acetate in petroleum ether) to afford 20
(2.252 g, 78%) as a yellowish oil. [a]2D1 ꢁ38.8 (c 1.1, CH2Cl2).
1H NMR (CDCl3, 300 MHz) d 7.36e7.20 (m, 10H, H-Ar),
4.83 (d, J4,5¼1.8 Hz, 1H, H-4), 4.81 (d, Jgem¼11.2 Hz, 1H,
OCHHPh), 4.71 (d, Jgem¼11.2 Hz, 1H, OCHHPh), 4.54 (d,
J1,2¼9.8 Hz, 1H, H-1), 4.51 (d, Jgem¼12.2 Hz, 1H, OCHHPh),
4.34 (d, Jgem¼12.2 Hz, 1H, OCHHPh), 4.27 (d, J2,1¼9.8 Hz,
1H, H-2), 3.58 (dq, J5,6¼6.6 Hz, J5,4¼1.8 Hz, 1H, H-5),
2.89e2.58 (m, 2H, SCH2CH3), 1.31 (d, J6,5¼6.6 Hz, 3H, H-
6), 1.29 (t, Jvic¼7.5 Hz, 3H, SCH2CH3); 13C NMR (CDCl3,
75 MHz) d 154.8 (CN), 137.6, 137.1 (2Cipso), 128.7e127.7
(C-Ar), 86.7 (C1), 76.0, 74.7, 73.9, 71.2, 70.2 (C2, C4, C5,
2OCH2Ph), 24.4 (SCH2CH3), 16.4 (C6), 14.8 (SCH2CH3).
ESI-MS for C22H27NO4S (m/z): Mr (calcd) 401.17, Mr (found)
424.42 (MþNa)þ. Anal. Calcd: C 65.81, H 6.78, N 3.49. Found:
C 65.95, H 6.59, N 3.43.
4.1.8. Ethyl 2,4-di-O-benzyl-3-deoxy-1-thio-3-(2,2,2-tri-
chloroethoxycarbonylamino)-b-D-fucopyranoside (6)
A solution of 21 (138 mg, 0.356 mmol) in pyridine (2.0 mL)
was treated with 2,2,2-trichloroethyl chloroformate (122 mL,
0.890 mmol) and stirred at rt for 2 h, after that MeOH (10 mL)
was added and the solution was concentrated. The residue was
subjected to a column chromatography (1% methanol in di-
chloromethane) to afford 6 (164 mg, 82%) as a yellowish pow-
1
der. [a]2D1 þ25 (c 0.8, CH2Cl2). H NMR (CDCl3, 300 MHz)
d 7.40e7.28 (m, 10H, H-Ar), 4.86 (d, Jgem¼10.8 Hz, 1H,
OCHHPh), 4.76 (d, Jgem¼11.3 Hz, 1H, OCHHPh), 4.63 (s,
2H, OCH2CCl3), 4.58 (d, Jgem¼11.3 Hz, 1H, OCHHPh), 4.48
(d, J1,2¼9.3 Hz, 1H, H-1), 4.44 (d, Jgem¼10.8 Hz, 1H,
OCHHPh), 3.84 (dt, J3,2¼J3,NH¼9.9 Hz, J3,4¼3.0 Hz, 1H, H-
3), 3.66e3.57 (m, 2H, H-4, H-5), 3.42 (t, J2,3¼J2,1¼9.9 Hz,
1H, H-2), 2.84e2.71 (m, 2H, SCH2CH3), 1.37e1.32 (m, 6H,
H-6, SCH2CH3); 13C NMR (CDCl3, 75 MHz) d 154.0 (CO),
137.7, 137.5 (2Cipso), 128.6e127.9 (C-Ar), 95.4 (CCl3), 85.6
(C1), 79.4, 76.3, 75.9, 75.0, 74.6, 74.3 (C2, C4, C5, 2OCH2Ph,
OCH2CCl3), 57.1 (C3), 25.0 (SCH2CH3), 17.1 (C6), 14.9
(SCH2CH3). ESI-MS for C25H30Cl3NO5S (m/z): Mr (calcd)
561.09, Mr (found) 584.29 (MþNa)þ. Anal. Calcd: C 53.34, H
5.37, N 2.49. Found: C 53.38, H 5.35, N 2.48.
4.1.7. Ethyl 3-amino-2,4-di-O-benzyl-3-deoxy-1-thio-b-D-
fucopyranoside (21) and ethyl 3-amino-2,4-di-O-benzyl-
3,6-dideoxy-1-thio-b-D-gulopyranoside (22)
A solution of 20 (2.231 g, 5.56 mmol) in THF (25 mL) was
cooled to 0 ꢀC and then treated with a 70% solution of Red-
AlÒ in toluene (7.16 mL, 27.8 mmol). The solution was stirred
at 0 ꢀC for 5 h, then water (10 mL) was added dropwise. The
mixture was diluted with ethyl acetate (200 mL) and washed
with water (200 mL). The organic layer was collected, dried
over anhydrous Na2SO4, filtered and concentrated. The residue
was subjected to column chromatography (1% methanol in di-
chloromethane) to afford 22 (168 mg, 8%) as a yellowish oil.
[a]2D1 ꢁ12 (c 0.4, CH2Cl2). 1H NMR (CDCl3, 300 MHz)
d 7.36e7.25 (m, 10H, H-Ar), 4.90 (d, J1,2¼9.6 Hz, 1H, H-1),
4.71 (d, Jgem¼11.6 Hz, 1H, OCHHPh), 4.55 (d, Jgem¼11.6 Hz,
1H, OCHHPh), 4.53 (d, Jgem¼12.0 Hz, 1H, OCHHPh), 4.49
(d, Jgem¼12.0 Hz, 1H, OCHHPh), 4.57 (dq, J5,6¼6.4 Hz,
4.1.9. Methyl 2,4-di-O-benzyl-3-deoxy-3-(2,2,2-trichloro-
ethoxycarbonylamino)-a-D-fucopyranosyl-(1/2)-3-O-
allyl-4-O-benzyl-b-D-rhamnopyranosyl-(1/3)-2,4-di-O-
benzyl-a-D-rhamnopyranoside (24)
A mixture of acceptor 23 (34 mg, 0.054 mmol) and donor 6
(60 mg, 0.107 mmol) was coevaporated three times with toluene
(2 mL). The residue was mixed with freshly activated AW-300

3388
D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
ꢀ
˚
4 A molecular sieves, cooled to ꢁ40 C, suspended in 1:1 Et2O/
CH2Cl2 (2.0 mL) under Ar atmosphere and then treated with
NIS (30 mg, 0.134 mmol) and AgOTf (12 mg, 0.047 mmol).
The temperature was allowed to rise gradually to ꢁ20 ꢀC. After
1 h stirring at ꢁ20 ꢀC, the mixture was diluted with CH2Cl2
(30 mL) and washed with 10% Na2S2O3 (30 mL) and then
with 1 M NaHCO3 (30 mL). The organic layer was collected,
dried over anhydrous Na2SO4, filtered and concentrated to
give a residue that, after column chromatography (10e30%
ethyl acetate in petroleum ether), afforded 24 (47 mg, 77%) as
a yellowish oil. [a]D21 þ145 (c 0.7, CH2Cl2). 1H NMR (CDCl3,
600 MHz) d 7.49e7.18 (m, 25H, H-Ar), 5.98e5.92 (m, 1H,
OCH2CH]CH2), 5.73 (d, J1,2¼3.3 Hz, 1H, H-1C), 5.34 (d,
J¼16.8 Hz, 1H, trans OCH2CH]CHH), 5.24 (d, J¼10.3 Hz,
1H, cis OCH2CH]CHH), 5.14 (d, J¼10.3 Hz, 1H, OCHHPh),
5.07 (d, J¼12.1 Hz, 1H, OCHHPh), 5.01 (d, JH,NH¼5.1 Hz, 1H,
NH), 4.81e4.46 (m, 12H, H-1A, H-5C, OCH2CCl3, 8OCHHPh),
4.38 (br s, 1H, H-1B), 4.26e4.20 (m, 3H, H-2B, H-3C,
OCHHCH]CH2), 4.17e4.11 (m, 3H, H-3A, H-4C,
OCHHCH]CH2), 3.78 (dd, J2,3¼11.1 Hz, J2,1¼3.3 Hz, 1H,
H-2C), 3.74e3.67 (m, 2H, H-2A, H-5A), 3.56 (t,
J4,5¼J4,3¼8.8 Hz, 1H, H-4A), 3.50 (t, J4,5¼J4,3¼9.3 Hz, 1H,
H-4B), 3.35 (s, 3H, OCH3), 3.29 (dd, J3,4¼9.3 Hz,
J3,2¼2.3 Hz, 1H, H-3B), 3.21 (dq, J5,4¼9.3 Hz, J5,6¼6.0 Hz,
1H, H-5B), 1.41 (d, J6,5¼6.2 Hz, 3H, H-6A), 1.26e1.23 (m,
6H, H-6B, H-6C); 13C NMR (CDCl3, 150 MHz) d 153.9
(CO), 138.9, 138.6, 138.5, 138.1, 138.0 (5Cipso-Bn),
134.6 (OCH2CH]CH2), 128.5e126.8 (C-Ar), 117.2
(OCH2CH]CH2), 99.0, 97.8, 95.7, 95.1 (C1A, C1B, C1C, CCl3),
83.6, 80.4, 80.2, 78.9, 76.5, 76.3, 75.1, 74.8, 74.1, 73.5, 73.4,
72.4, 72.0, 71.7, 71.1, 69.9, 67.2, 66.0 (C2A, C2B, C2C, C3A, C3B,
C4A, C4B, C4C, C5A, C5B, C5C, 5OCH2Ph, OCH2CH]CH2,
OCH2CCl3), 54.6, 52.0 (C3C, OCH3), 18.5, 17.8, 16.8 (C6A, C6B,
C6C). MALDI-MS for C60H70Cl3NO14 (m/z): Mr (calcd)
1133.39, Mr (found) 1156.42 (MþNa)þ. Anal. Calcd: C 63.46,
H 6.21, N 1.23. Found: C 63.30, H 6.30, N 1.21.
subjected to column chromatography (8e16% ethyl acetate in
petroleum ether) to give 8 (56 mg, 62%) as a colourless oil. 1H
NMR (CDCl3, 300 MHz) d 7.42e7.29 (m, 10H, H-Ar), 6.97
(d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3), 6.84 (d, Jortho¼9.0 Hz,
2H, C6H2H2OCH3), 5.45 (br s, 1H, H-1), 4.95 (d, Jgem¼11.1 Hz,
1H, OCHHPh), 4.82 (d, Jgem¼11.4 Hz, 1H, OCHHPh), 4.70 (d,
Jgem¼11.1 Hz, 1H, OCHHPh), 4.68 (d, Jgem¼11.4 Hz, 1H,
OCHHPh), 4.21e4.14 (m, 1H, H-3), 3.94 (dd, J2,3¼3.9 Hz,
J2,1¼1.2 Hz, 1H, H-2), 3.84 (dq, J5,4¼9.3 Hz, J5,6¼6.3 Hz, 1H,
H-5), 3.78 (s, 3H, C6H4OCH3), 3.44 (t, J4,5¼J4,3¼9.3 Hz, 1H,
H-4), 2.45 (d, JH,OH¼7.0 Hz, 1H, OH), 1.34 (d, J6,5¼6.3 Hz,
3H, H-6); 13C NMR (CDCl3, 75 MHz) d 154.7, 150.2 (2Cipso
-
MP), 138.3, 137.5 (2Cipso-Bn), 128.5e127.6 (C-Ar), 117.4,
114.5 (C-Ar MP), 95.9 (C1), 82.1, 78.3 (C2, C4), 75.0, 73.1,
71.4, 67.8 (C3, C5, 2OCH2Ph), 55.6 (OCH3), 18.0 (C6).
ESI-MS for C27H30O6 (m/z): Mr (calcd) 450.20, Mr (found)
472.97 (MþNa)þ. Anal. Calcd: C 71.98, H 6.71. Found: C
71.66, H 6.61.
4.1.11. 4-Methoxyphenyl 3-O-allyl-2-O-benzenesulfonyl-4-
O-benzyl-D-rhamnopyranosyl-(1/3)-2,4-di-O-benzyl-a-D-
rhamnopyranoside (25)
From compounds 8 and 9da mixture of acceptor 8 (40 mg,
89.7 mmol) and donor 9 (89 mg, 143.5 mmol) was coevaporated
three times with toluene (2 mL). The residue was mixed with
˚
freshly activated AW-300 4 A molecular sieves, cooled to
ꢁ50 ꢀC and suspended in CH2Cl2 (3.0 mL) under Ar atmo-
sphere. A 144 mM solution of TMSOTf in CH2Cl2 (70 mL,
9.9 mmol) was added. The temperature was allowed to rise grad-
ually to ꢁ25 ꢀC. After 2 h stirring at ꢁ25 ꢀC, some drops of
Et3N were added. The mixture was filtered over a Celite pad,
then concentrated to give a residue that was subjected to column
chromatography (1e3% ethyl acetate in toluene) to afford 25a
(42 mg, 53%) as a yellowish oil. As second eluted compound
25b (28 mg, 35%) was recovered as a yellowish oil.
From compound 26bdcompound 26b (1.066 g, 1.35 mmol)
was dissolved in 1:1:0.1 v/v/v AcOH/Ac2O/TFA (42 mL). The
yellow solution was heated to 70 ꢀC and stirred overnight, after
that it was cooled to 0 ꢀC and water (40 mL) was very carefully
added. The solution was diluted with CH2Cl2 (500 mL) and
washed twice with water (500 mL) and then with 0.1 M
NaHCO3 (500 mL). The organic layer was collected, dried
over anhydrous Na2SO4, filtered and concentrated to give a res-
idue that was mixed with p-methoxyphenol (336 mg,
2.71 mmol) and then dissolved at 0 ꢀC in CH2Cl2 (26 mL) under
Ar atmosphere. The solution was then treated with BF3$OEt2
(50 mL, 0.410 mmol), stirred at 0 ꢀC for 2 h, then diluted with
CH2Cl2 (200 mL) and washed with 1 M NaHCO3 (200 mL).
The organic layer was collected, dried over anhydrous
Na2SO4, filtered and concentrated to give a residue that was sub-
jected to a column chromatography (1e3% ethyl acetate in tol-
uene) to afford 25b (772 mg, 65%) as a yellowish oil.
4.1.10. 4-Methoxyphenyl 2,4-di-O-benzyl-a-D-rhamno-
pyranoside (8)
Alcohol7(90 mg, 0.254 mmol)wasdissolvedin1:1:0.1v/v/v
Ac2OH/Ac2O/TFA (2.2 mL). The yellow solution was stirred
at 70 ꢀC overnight, after that the solution was cooled to 0 ꢀC
and water (10 mL) was very carefully added. The solution was
diluted with CH2Cl2 (50 mL) and washed twice with water
(50 mL) and then with 0.1 M NaHCO3 (50 mL). The organic
layer was collected, dried over anhydrous Na2SO4, filtered
and concentrated to give a residue that was mixed with p-me-
thoxyphenol (63 mg, 0.508 mmol) and then dissolved at 0 ꢀC
in CH2Cl2 (2.5 mL) under Ar atmosphere. The solution was
then treated with BF3$OEt2 (9.4 mL, 76.2 mmol), stirred at
0 ꢀC for 2 h, then diluted with CH2Cl2 (40 mL) and washed
with 1 M NaHCO3 (40 mL). The organic layer was collected,
dried over anhydrous Na2SO4, filtered and concentrated to
give a residue that was dissolved in MeOH (3.0 mL) and
t-BuOK was added until the solution was strongly basic. The
solution was stirred at rt for 40 min, then neutralized with Am-
berlyst-15 (Hþ), filtered and concentrated. The residue was
25a: [a]2D1 þ30 (c 0.7, CH2Cl2). 1H NMR (CDCl3, 400 MHz)
d 7.55e7.15 (m, 20H, H-Ar), 6.93 (d, Jortho¼9.2 Hz, 2H,
C6H2H2OCH3), 6.80 (d, Jortho¼9.2 Hz, 2H, C6H2H2OCH3),
5.98e5.86 (m, 1H, OCH2CH]CH2), 5.35 (d, J2,3¼3.0 Hz,
1H, H-2B), 5.28 (d, Jvic¼17.2 Hz, 1H, trans OCH2CH]CHH),

D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
3389
5.18e5.08 (m, 3H, H-1A, H-1B, cis OCH2CH]CHH), 4.93 (d,
Jgem¼11.0 Hz, 1H, OCHHPh), 4.82 (d, Jgem¼10.8 Hz, 1H,
OCHHPh), 4.72 (s, 2H, 2OCHHPh), 4.64 (d, Jgem¼11.0 Hz,
1H, OCHHPh), 4.60 (d, Jgem¼10.8 Hz, 1H, OCHHPh), 4.46
(d, Jgem¼14.0 Hz, 1H, OSO2CHHPh), 4.37 (d, Jgem¼14.0 Hz,
concentrated. The residue was dissolved in CH2Cl2 (300 mL)
and washed with 1 M NaHCO3 (300 mL) and brine (300 mL).
The organic layer was collected, dried over anhydrous
Na2SO4, filtered and concentrated to give a residue that, after
column chromatography (10e50% ethyl acetate in petroleum
ether), afforded 27 (333 mg, 52%) as a yellowish oil. [a]D21
1H,
OSO2CHHPh),
4.25e4.14
(m,
2H,
H-3A,
1
OCHHCH]CH2), 4.06 (dd, 1H, Jgem¼12.6 Hz, Jvic¼5.4 Hz,
OCHHCH]CH2), 3.91e3.76 (m, 7H, H-2A, H-3B, H-5A, H-
5B, C6H4OCH3), 3.63 (t, J4,5¼J4,3¼9.2 Hz, 1H, H-4A), 3.39 (t,
J4,5¼J4,3¼9.4 Hz, 1H, H-4B), 1.28e1.24 (m, 6H, H-6A, H-6B);
13C NMR (CDCl3, 100 MHz) d 154.9, 150.3 (2Cipso-MP),
138.4, 137.9, 137.8 (3Cipso-Bn), 134.3 (OCH2CH]CH2),
130.9 (Cipso-SO2Bn), 128.1e126.3 (C-Ar), 120.5, 117.8,
114.6 (C-Ar MP, OCH2CH]CH2), 99.2, 96.4 (C1A, C1B),
84.6, 80.4, 79.7, 78.2, 77.5, 76.3, 76.2, 75.3, 72.8, 71.4, 68.7,
68.6 (C2A, C2B, C3A, C3B, C4A, C4B, C5A, C5B, 3OCH2Ph,
OCH2CH]CH2), 57.6, 55.6 (OCH3, OSO2CH2Ph), 18.0, 17.9
(C6A, C6B). MALDI-MS for C50H56O12S (m/z): Mr (calcd)
880.35, Mr (found) 903.41 (MþNa)þ. Anal. Calcd: C 68.16, H
6.41. Found: C 68.04, H 6.38.
þ24.0 (c 1.0, CH2Cl2). H NMR (CDCl3, 300 MHz) d 7.44e
7.26 (m, 15H, H-Ar), 6.98 (d, Jortho¼9.0 Hz, 2H,
C6H2H2OCH3), 6.80 (d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3),
6.16e6.00 (m, 1H, OCH2CH]CH2), 5.46 (d, J1,2¼2.4 Hz,
1H, H-1A), 5.39 (d, Jvic¼17.3 Hz, 1H, trans OCH2CH]CHH),
5.27 (d, Jvic¼11.2 Hz, 1H, cis OCH2CH]CHH), 5.00 (d,
Jgem¼12.0 Hz, 1H, OCHHPh), 4.97 (d, Jgem¼11.7 Hz, 1H,
OCHHPh), 4.87 (d, Jgem¼12.6 Hz, 1H, OCHHPh), 4.73e4.64
(m, 3H, OCHHPh), 4.46e4.43 (m, 2H, H-3A, H-1B), 4.28
(dd, Jgem¼12.0 Hz, Jvic¼6.0 Hz, 1H, OCHHCH]CH2), 4.15
(dd, Jgem¼12.0 Hz, Jvic¼6.0 Hz, 1H, OCHHCH]CH2),
3.98e3.95 (m, 2H, H-2A, H-2B), 3.88 (dq, J5,4¼9.0 Hz,
J5,6¼6.0 Hz, 1H, H-5A), 3.79 (s, 3H, OCH3), 3.68 (t,
J4,5¼J4,3¼9.0 Hz, 1H, H-4A), 3.53 (t, J4,5¼J4,3¼9.0 Hz, 1H,
H-4B), 3.38 (dd, J3,4¼9.0 Hz, J3,2¼2.7 Hz, 1H, H-3B), 3.34
(dq, J5,4¼9.0 Hz, J5,6¼6.0 Hz, 1H, H-5B), 1.36e1.33 (m, 6H,
H-6B, H-6A); 13C NMR (CDCl3, 50 MHz) d 154.4, 149.7
(2Cipso-MP), 137.8, 137.7, 137.3 (3Cipso-Bn), 134.3
(OCH2CH]CH2), 127.9e127.1 (C-Ar), 117.2, 116.3, 114.0
(C-Ar MP, OCH2CH]CH2), 96.9, 96.3 (C1A, C1B), 80.8,
79.3, 79.0, 75.6, 74.9, 74.3, 74.1, 72.1, 71.1, 70.0, 68.2, 67.8
(C2A, C2B, C3A, C3B, C4A, C4B, C5A, C5B, 3OCH2Ph,
OCH2CH]CH2), 55.1 (OCH3), 17.6, 17.4 (C6A, C6B). ESI-
MS for C43H50O10 (m/z): Mr (calcd) 726.34, Mr (found)
749.57 (MþNa)þ. Anal. Calcd: C 71.05, H 6.93. Found: C
71.10, H 6.95.
25b: [a]2D1 þ2.8 (c 1.0, CH2Cl2). 1H NMR (CDCl3, 400 MHz)
d 7.39e7.10 (m, 20H, H-Ar), 6.91 (d, Jortho¼9.0 Hz, 2H,
C6H2H2OCH3), 6.76 (d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3),
6.01e5.89 (m, 1H, OCH2CH]CH2), 5.44 (d, J2,3¼2.9 Hz,
1H, H-2B), 5.35 (d, Jvic¼17.3 Hz, 1H, trans OCH2CH]CHH),
5.19 (d, Jvic¼11.2 Hz, 1H, cis OCH2CH]CHH), 5.08 (d,
J1,2¼2.2 Hz, 1H, H-1A), 4.93 (d, Jgem¼11.0 Hz, 1H, OCHHPh),
4.89 (d, Jgem¼10.9 Hz, 1H, OCHHPh), 4.79 (d, Jgem¼12.1 Hz,
1H, OCHHPh), 4.68 (d, Jgem¼12.1 Hz, 1H, OCHHPh), 4.58
(d, Jgem¼10.9 Hz, 1H, OCHHPh), 4.52e4.47 (m, 4H, H-1B,
OCHHPh, OSO2CH2Ph), 4.35e4.25 (m, 2H, H-3A,
OCHHCH]CH2), 4.05 (dd, Jgem¼12.4 Hz, Jvic¼5.6 Hz, 1H,
OCHHCH]CH2), 3.95 (t, J2,1¼J2,3¼2.2 Hz, 1H, H-2A), 3.82
(dq, J5,4¼9.0 Hz, J5,6¼6.3 Hz, 1H, H-5A), 3.70 (s, 3H,
C6H4OCH3), 3.60 (t, J4,5¼J4,3¼9.0 Hz, 1H, H-4A), 3.39 (dd,
J3,4¼9.0 Hz, J3,2¼2.9 Hz, 1H, H-3B), 3.35 (t, J4,5¼J4,3¼9.0 Hz,
1H, H-4B), 3.28 (dq, J5,4¼9.0 Hz, J5,6¼6.0 Hz, 1H, H-5B), 1.27
4.1.13. 4-Methoxyphenyl 2,4-di-O-benzyl-3-deoxy-3-(2,2,2-
trichloroethoxycarbonylamino)-a-D-fucopyranosyl-(1/2)-
3-O-allyl-4-O-benzyl-b-D-rhamnopyranosyl-(1/3)-2,4-di-
O-benzyl-a-D-rhamnopyranoside (4)
(d, J6,5¼6.0 Hz, 3H, H-6B), 1.19 (d, J6,5¼6.3 Hz, 3H, H-6A); 13
C
Acceptor 27 (179 mg, 0.246 mmol) was mixed with donor
6 (281 mg, 0.499 mmol). The mixture was coevaporated three
times with toluene (5 mL), then mixed with freshly activated
NMR (CDCl3, 100 MHz) d 154.4, 152.8 (2Cipso-MP), 137.8,
137.5, 137.3 (3Cipso-Bn), 133.6 (OCH2CH]CH2), 130.4
(Cipso-SO2Bn), 128.1e127.1 (C-Ar), 117.5, 115.5, 114.3
(C-Ar MP, OCH2CH]CH2), 97.1, 96.6 (C1A, C1B), 79.8,
78.9, 77.2, 76.6, 76.0, 75.3, 75.1, 73.3, 72.3, 71.4, 70.5, 67.8
(C2A, C2B, C3A, C3B, C4A, C4B, C5A, C5B, 3OCH2Ph,
OCH2CH]CH2), 57.2, 55.3 (OCH3, SO2CH2Ph), 17.8, 17.4
(C6A, C6B). MALDI-MS for C50H56O12S (m/z): Mr (calcd)
880.35, Mr (found) 903.54 (MþNa)þ. Anal. Calcd: C 68.16, H
6.41. Found: C 67.98, H 6.36.
ꢀ
˚
AW-300 4 A molecular sieves, cooled to ꢁ40 C and sus-
pended in 1:1 v/v Et2O/CH2Cl2 (4.0 mL) under Ar atmo-
sphere. The mixture was treated with NIS (141 mg,
0.627 mmol) and AgOTf (54 mg, 0.209 mmol). The tempera-
ture was allowed to rise gradually to ꢁ20 ꢀC. After 90 min
stirring at ꢁ20 ꢀC, the mixture was diluted with CH2Cl2
(100 mL) and washed with 10% Na2S2O3 (100 mL) and then
with 1 M NaHCO3 (100 mL). The organic layer was collected,
dried over anhydrous Na2SO4, filtered and concentrated to
give an oily residue. After column chromatography (10e
30% ethyl acetate in petroleum ether), 4 (196 mg, 65%) was
4.1.12. 4-Methoxyphenyl 3-O-allyl-4-O-benzyl-b-D-rhamno-
pyranosyl-(1/3)-2,4-di-O-benzyl-a-D-rhamnopyranoside
(27)
Disaccharide 25b (776 mg, 0.882 mmol) was dissolved in
dry DMF (15 mL) under Ar atmosphere and then NaNH2
(690 mg, 17.6 mmol) was added. The mixture was stirred at
rt for 48 h, after that we added MeOH (60 mL) and then, drop-
wise, AcOH to neutralize the solution, that was then
1
obtained as a yellowish oil. [a]2D1 þ130 (c 0.5, CH2Cl2). H
NMR (CDCl3, 500 MHz) d 7.54e7.33 (m, 25H, H-Ar), 7.02
(d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3), 6.88 (d, Jortho¼9.0 Hz,
2H, C6H2H2OCH3), 6.06e5.98 (m, 1H, OCH2CH]CH2),
5.78 (d, J1,2¼2.8 Hz, 1H, H-1C), 5.43 (d, J1,2¼3.2 Hz, 1H,
H-1A), 5.39 (d, Jvic¼17.5 Hz, 1H, trans OCH2CH]CHH),

3390
D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
5.29 (d, Jvic¼10.0 Hz, 1H, cis OCH2CH]CHH), 5.15 (d,
Jgem¼12.4 Hz, 1H, OCHHPh), 5.12 (d, Jgem¼12.5 Hz, 1H,
OCHHPh), 5.05 (d, JH,NH¼4.5 Hz, 1H, NH), 4.87 (d,
Jgem¼12.5 Hz, 1H, OCHHPh), 4.79e4.53 (m, 11H, H-1B, H-
5C, OCH2CCl3, 7OCHHPh), 4.36e4.18 (m, 6H, H-2B, H-3A,
H-3C, H-4C, OCH2CH]CH2), 3.99 (br s, 1H, H-2A), 3.95
(dq, J5,4¼9.9 Hz, J5,6¼6.0 Hz, 1H, H-5A), 3.87e3.83 (m,
4H, H-2C, OCH3), 3.68 (t, J4,5¼J4,3¼9.9 Hz, 1H, H-4A),
3.59 (t, J4,5¼J4,3¼9.0 Hz, 1H, H-4B), 3.39 (dd, J3,4¼9.0 Hz,
J3,2¼2.4 Hz, 1H, H-3B), 3.33 (dq, J5,4¼9.0 Hz, J5,6¼6.0 Hz,
1H, H-5B), 1.41 (d, J6,5¼6.0 Hz, 3H, H-6C), 1.33 (d,
J6,5¼6.0 Hz, 3H, H-6B), 1.27 (d, J6,5¼6.0 Hz, 3H, H-6A);
13C NMR (CDCl3, 50 MHz) d 154.9, 153.9, 150.6 (CO,
2Cipso-MP), 138.8, 138.6, 138.5, 138.0, 137.9 (5Cipso-Bn),
134.6 (OCH2CH]CH2), 129.0e125.3 (C-Ar), 118.2, 117.2,
114.5 (OCH2CH]CH2, C-Ar MP), 98.4, 98.1, 95.7, 95.1
(C1A, C1B, C1C, CCl3), 83.6, 80.8, 80.4, 79.0, 77.1, 76.2,
75.4, 75.1, 74.2, 73.5, 73.2, 72.6, 72.1, 71.6, 71.0, 70.0,
1185.38, Mr (found) 1208.43 (MþNa)þ. Anal. Calcd: C
63.72, H 5.94, N 1.18. Found: C 63.63, H 6.02, N 1.16.
4.1.15. 2,4-Di-O-benzyl-3-deoxy-3-(2,2,2-trichloroethoxy-
carbonylamino)-a-D-fucopyranosyl-(1/2)-3-O-allyl-4-O-
benzyl-b-D-rhamnopyranosyl-(1/3)-2,4-di-O-benzyl-a-D-
rhamnopyranose (29)
A solution of 4 (130 mg, 0.106 mmol) in 2:2:1 v/v/v toluene/
CH3CN/water (15 mL) was treated with CAN (290 mg,
0.530 mmol) and stirred at rt overnight. The solution was then
diluted with ethyl acetate (100 mL) and washed with 1 M
NaHCO3 (100 mL) and then with water (100 mL). The organic
layer was collected, dried over anhydrous Na2SO4, filtered and
concentrated to give a residue that, after column chromatogra-
phy (10e50% ethyl acetate in petroleum ether), afforded 29
1
(81 mg, 68%; a/b¼4.5:1) as a yellowish oil. a-anomer: H
NMR (CDCl3, 400 MHz) d 7.42e7.18 (m, 25H, H-Ar), 6.02e
5.90 (m, 1H, OCH2CH]CH2), 5.73 (d, J1,2¼2.8 Hz, 1H, H-
1C), 5.34 (d, Jvic¼17.2 Hz, 1H, trans OCH2CH]CHH), 5.24
(d, Jvic¼10.3 Hz, 1H, cis OCH2CH]CHH), 5.18 (d,
J1,2¼2.3 Hz, 1H, H-1A), 5.12 (d, Jgem¼10.5 Hz, 1H, OCHHPh),
5.07 (d, Jgem¼12.0 Hz, 1H, OCHHPh), 5.01 (d, JH,NH¼6.3 Hz,
1H, NH), 4.78e4.13 (m, 18H, H-1B, H-2B, H-3A, H-3C, H-4C,
H-5C, 8OCHHPh, CH2CCl3, OCH2CH]CH2), 3.97 (dq,
J5,4¼7.9 Hz, J5,6¼6.2 Hz, 1H, H-5A), 3.81e3.74 (m, 2H, H-
2A, H-2C), 3.59 (t, J4,5¼J4,3¼7.9 Hz, 1H, H-4A), 3.53 (t,
J4,5¼J4,3¼9.4 Hz, 1H, H-4B), 3.31 (dd, J3,4¼9.4 Hz,
J3,2¼1.8 Hz, 1H, H-3B), 3.23 (dq, J5,4¼9.4 Hz, J5,6¼6.2 Hz,
1H, H-5B), 1.39 (d, J6,5¼6.1 Hz, 3H, H-6C), 1.29e1.25 (m,
6H, H-6A, H-6B); 13C NMR (CDCl3, 100 MHz) d 153.9 (CO),
138.8, 138.5, 138.4, 138.0, 137.9 (5Cipso), 134.5
(OCH2CH]CH2), 128.7e126.8 (C-Ar), 117.2 (OCH2
CH]CH2), 97.9, 95.6, 95.0, 92.6 (C1A, C1B, C1C, CCl3), 83.5,
80.3, 78.9, 77.6, 76.3, 76.1, 75.3, 75.0, 74.1, 73.5, 73.3, 72.3,
68.0, 66.0 (C2A, C2B, C2C, C3A, C3B, C4A, C4B, C4C, C5A
C
,
5B, C5C, 5OCH2Ph, OCH2CH]CH2, OCH2CCl3), 55.6,
52.0 (C3C, OCH3), 18.7, 17.8, 16.8 (C6A, C6B, C6C).
MALDI-MS for C66H74Cl3NO15 (m/z): Mr (calcd) 1225.41,
Mr (found) 1248.64 (MþNa)þ. Anal. Calcd: C 64.57, H
6.08, N 1.14. Found: C 64.45, H 6.03, N 1.12.
4.1.14. 4-Methoxyphenyl 2,4-di-O-benzyl-3-deoxy-3-(2,2,2-
trichloroethoxycarbonylamino)-a-D-fucopyranosyl-(1/2)-
4-O-benzyl-b-D-rhamnopyranosyl-(1/3)-2,4-di-O-benzyl-
a-D-rhamnopyranoside (28)
A solution of 4 (39 mg, 31.7 mmol) in 2:1 v/v MeOH/CH2Cl2
(2.1 mL) was treated with PdCl2 (1.7 mg, 9.5 mmol). The mix-
ture was vigorously stirred at rt overnight and then filtered over
a Celite pad, diluted with CH2Cl2 (30 mL) and washed with wa-
ter (30 mL). The organic layer was collected, dried over anhy-
drous Na2SO4, filtered and concentrated. The residue was
subjected to column chromatography (10% ethyl acetate in tol-
uene) to give 28 (32 mg, 86%) as a colourless oil. [a]2D1 þ54.4 (c
71.9, 71.6, 71.1, 69.9, 67.5, 65.9 (C2A, C2B, C2C, C3A, C3B
C
,
4A, C4B, C4C, C5A, C5B, C5C, 5OCH2Ph, OCH2CCl3,
OCH2CH]CH2), 51.9 (C3C), 18.5, 17.7, 16.7 (C6A, C6B
,
6C). MALDI-MS for C59H68Cl3NO14 (m/z): Mr (calcd)
C
1.5, CH2Cl2). H NMR (CDCl3, 400 MHz) d 7.39e7.12 (m,
1119.37, Mr (found) 1142.73 (MþNa)þ. Anal. Calcd: C 63.18,
1
25H, H-Ar), 6.95 (d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3), 6.81
(d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3), 5.37 (d, J1,2¼2.7 Hz,
1H, H-1A), 5.23 (d, J1,2¼3.2 Hz, 1H, H-1C), 4.92 (d,
Jgem¼10.4 Hz, 1H, OCHHPh), 4.84e4.24 (m, 14H, 9OCHHPh,
H-1B, H-3C, CH2CCl3, NH), 3.92e3.86 (m, 3H, H-2A, H-3A, H-
5C), 3.77 (s, 3H, OCH3), 3.73 (dd, J2,3¼10.9 Hz, J2,1¼3.2 Hz,
1H, H-2C), 3.65e3.55 (m, 2H, H-4C, H-5A), 3.47 (t,
J4,5¼J4,3¼8.8 Hz, 1H, H-4A), 3.34 (br s, 1H, H-2B), 3.27e
3.20 (m, 3H, H-3B, H-4B, H-5B), 1.34 (d, J6,5¼6.2 Hz, 3H, H-
6C), 1.28e1.25 (m, 6H, H-6A, H-6B); 13C NMR (CDCl3,
100 MHz) d 155.0, 153.9, 150.5 (CO, 2Cipso-MP), 138.7,
138.6, 138.0, 137.7, 136.7 (5Cipso-Bn), 129.1e125.3 (C-Ar),
H 6.11, N 1.25. Found: C 63.04, H 6.18, N 1.24.
4.1.16. 4-Methoxyphenyl 3-acetamido-3-deoxy-a-D-fuco-
pyranosyl-(1/2)-b-D-rhamnopyranosyl-(1/3)-a-D-
rhamnopyranoside (2)
A solution of 28 (9.0 mg, 7.6 mmol) in 2:1 v/v AcOH/Ac2O
(900 mL) was treated with Zn/Cu couple (23 mg) and vigor-
ously stirred for 3 days. The mixture was filtered over a Celite
pad, then diluted with ethyl acetate (15 mL) and washed with
1 M NaHCO3 (15 mL) and water (15 mL). The organic layer
was collected, dried over anhydrous Na2SO4, filtered and con-
centrated to give a residue that was dissolved in 9:1 v/v
MeOH/HCOOH (1.0 mL). The solution was treated with a cat-
alytic amount of 10% Pd/C under argon atmosphere. After 2 h
in an ultrasound bath, the mixture was filtered over a Celite
pad and concentrated. The residue was subjected to a Sephadex
G-10 column chromatography with water as eluant. After
lyophilization, compound 2 (2.5 mg, 55%) was recovered as
117.9, 114.6 (C-Ar MP), 99.8, 98.8, 97.8, 95.6 (C1A, C1B
C1C, CCl3), 82.1, 80.8, 80.6, 80.4, 76.6, 76.5, 76.0, 75.7,
75.6, 75.2, 74.4, 74.3, 73.4, 72.8, 71.6, 68.0, 66.8 (C2A, C2B
C2C, C3A, C3B, C4A, C4B, C4C, C5A, C5B, C5C, 5OCH2Ph,
OCH2CCl3), 55.6, 52.2 (C3C, OCH3), 18.4, 18.0, 16.7 (C6A
6B, C6C). MALDI-MS for C63H70Cl3NO15 (m/z): Mr (calcd)
,
,
,
C

D. Comegna et al. / Tetrahedron 64 (2008) 3381e3391
3391
a white waxy solid. [a]D21 ꢁ224 (c 0.2, water). 1H NMR (D2O,
400 MHz) (see also Table 1) d 7.20 (d, Jortho¼9.0 Hz, 2H,
C6H2H2OCH3), 7.08 (d, Jortho¼9.0 Hz, 2H, C6H2H2OCH3),
3.89 (s, 3H, OCH3), 1.98 (s, 3H, Ac); 13C NMR (D2O,
100 MHz) (see also Table 1) d 176.0 (CO), 150.0, 147.5
(2Cipso-MP), 120.3, 116.4 (C-Ar MP), 56.8 (OCH3), 24.1
(COCH3). MALDI-MS for C27H41NO14 (m/z): Mr (calcd)
603.25, Mr (found) 626.26 (MþNa)þ. Anal. Calcd: C 53.72,
H 6.85, N 2.32. Found: C 53.50, H 6.99, N 2.29.
10. Bedini, E.; De Castro, C.; Erbs, G.; Mangoni, L.; Dow, J. M.; Newman,
M.-A.; Parrilli, M.; Unverzagt, C. J. Am. Chem. Soc. 2005, 129, 2414e
2416.
11. Gridley, J. J.; Osborn, H. M. I. J. Chem. Soc., Perkin Trans. 1 2000,
1471e1479.
12. Crich, D. ACS Symp. Ser. 2007, 960, 60e72.
13. Bedini, E.; Carabellese, A.; Barone, G.; Parrilli, M. J. Org. Chem. 2005,
70, 8064e8070.
14. Bedini, E.; Iadonisi, A.; Carabellese, A.; Parrilli, M. Tetrahedron Lett.
2004, 45, 4445e4448.
`
15. Valerio, S.; Iadonisi, A.; Adinolfi, M.; Ravida, A. J. Org. Chem. 2007, 72,
6097e6106.
Acknowledgements
16. Hou, D.; Lowary, T. L. Org. Lett. 2007, 9, 4487e4490.
17. Lattrell, R.; Lohaus, G. Justus Liebigs Ann. Chem. 1974, 901e920.
18. Albert, R.; Dax, K.; Link, R. W.; Stuetz, A. E. Carbohydr. Res. 1983, 118,
C5eC6.
NMR and MS facilities of CIMCF (Centro Interdipartimen-
tale di Metodologie Chimiche Fisiche) are acknowledged.
19. Dong, H.; Pei, Z.; Ramstrom, O. J. Org. Chem. 2006, 71, 3306e3309.
¨
20. Hsu, D.-S.; Matsumoto, T.; Suzuki, K. Synlett 2005, 801e804.
21. Adinolfi, M.; Barone, G.; Corsaro, M. M.; Lanzetta, R.; Monaco, P.
J. Carbohydr. Chem. 1995, 14, 913e928.
References and notes
22. Hua, Y.; Gu, G.; Du, Y. Carbohydr. Res. 2004, 339, 867e872.
23. van den Bos, L. J.; Boltje, T. J.; Provoost, T.; Mazurek, J.; Over-
kleeft, H. S.; van der Marel, G. A. Tetrahedron Lett. 2007, 48,
2697e2700.
1. Endotoxins in Health and Disease; Brade, H., Opal, S. M., Vogel, S.,
Morrison, D. C., Eds.; Marcel Dekker: New York, NY, 1999.
2. Zipfel, C.; Felix, G. Curr. Opin. Plant Biol. 2005, 8, 353e360.
3. Parker, J. Trends Plant Sci. 2003, 8, 245e247.
´
´
´
24. Fekete, A.; Gyergyoi, K.; Kover, K. E.; Bajza, I.; Liptak, A. Carbohydr.
4. Nurnberger, T.; Brunner, F. Curr. Opin. Plant Biol. 2002, 5, 318e324.
¨
¨
Res. 2006, 341, 1312e1321.
25. Bedini, E.; Comegna, D.; Di Nola, A.; Parrilli, M., submitted.
´
26. Faure, R.; Shiao, T. C.; Damerval, S.; Roy, R. Tetrahedron Lett. 2007, 48,
5. Zeidler, D.; Zahringer, U.; Gerber, I.; Dubery, I.; Hartung, T.; Bors, W.;
¨
Hutzler, P.; Durner, J. Proc. Natl. Acad. Sci. U.S.A. 2004, 101,
15811e15816.
2385e2388.
´
6. Silipo, A.; Molinaro, A.; Sturiale, L.; Dow, J. M.; Erbs, G.; Lanzetta, R.;
Newman, M.-A.; Parrilli, M. J. Biol. Chem. 2005, 280, 33660e33668.
7. Newman, M.-A.; Erbs, G.; Tandrup Jensen, T.; Dow, J. M.; Molinaro, A.;
Parrilli, M. Biology of Plant-Microbe Interactions; Sanchez, F., Quinto, C.,
Lopez Lara, I.-M., Geiger, O., Eds.; APS: St. Paul, 2006; Vol. 5,
pp 231e235.
`
27. Faure, R.; Shiao, T. C.; Lagnoux, D.; Giguere, D.; Roy, R. Org. Biomol.
Chem. 2007, 5, 2704e2708.
28. Abdel-Rahman, A. A.-H.; Jonke, S.; El Ashry, E. S. H.; Schmidt, R. R.
Angew. Chem., Int. Ed. 2002, 41, 2972e2974.
29. Tanaka, H.; Yoshizawa, A.; Takahashi, T. Angew. Chem., Int. Ed. 2007,
46, 2505e2507.
8. Molinaro, A.; Silipo, A.; Lanzetta, R.; Newman, M.-A.; Dow, J. M.;
Parrilli, M. Carbohydr. Res. 2003, 338, 277e281.
30. Rao, V. S.; Perlin, A. S. Carbohydr. Res. 1980, 83, 175e177.
31. Joosten, J. A. F.; Evers, B.; van Summeren, R. P.; Kamerling, J. P.;
Vliegenthart, J. F. G. Eur. J. Org. Chem. 2003, 3569e3586.
9. Corsaro, M. M.; De Castro, C.; Molinaro, A.; Parrilli, M. Recent Res.
Devel. Phytochem. 2001, 5, 119e138.