First synthesis of the β-D-rhamnosylated trisaccharide repeating unit of the O-antigen from Xanthomonas campestris pv. campestris 8004

Article abstract of DOI:10.1021/jo051153d

The trisaccharide repeating unit of the O-antigen of the lipopolysaccharide from Xanthomonas campestris pv. campestris 8004, a pathogen of cruciferous crops, presents some structural features that renders it a challenging synthetic target: the presence of

Full text of DOI:10.1021/jo051153d

First Synthesis of the â-D-Rhamnosylated Trisaccharide Repeating
Unit of the O-Antigen from Xanthomonas campestris pv.
campestris 8004
Emiliano Bedini,* Antonella Carabellese, Gaspare Barone, and Michelangelo Parrilli
Dipartimento di Chimica Organica e Biochimica, Universita` di Napoli “Federico II”,
Complesso Universitario Monte Santangelo, Via Cintia 4, 80126 Napoli, Italy
ebedini@unina.it
Received June 8, 2005
The trisaccharide repeating unit of the O-antigen of the lipopolysaccharide from Xanthomonas
campestris pv. campestris 8004, a pathogen of cruciferous crops, presents some structural features
that renders it a challenging synthetic target: the presence of a â-D-rhamnosidic linkage, the steric
crowd on a 1,2-cis-diglycosylated D-rhamnose, and finally the noncommercial availability of its
monosaccharide constituents. The synthesis of this trisaccharide as methyl glycoside has been
accomplished by exploiting a strategy whose key steps were the sequential â-D-rhamnosylation
with a 2-O-benzylsulfonyl-N-phenyltrifluoroacetimidate donor, debenzylsulfonylation, and coupling
with a D-Fucp3NAc thioglycoside donor.
Introduction
far from being completely elucidated. A recent work
showed that the lipid A moiety may be at least partially
responsible for LPS perception by plant cells;4 neverthe-
less, oligosaccharides, in particular synthetic oligorham-
nans mimicking the general structure of the O-chains
from phytopathogenic bacteria, have also been proved to
prevent the hypersensitive response (HR).5 The aim to
investigate deeper the molecular basis of HR and LIR
effects on plants prompted the synthesis of model oli-
gosaccharides related to the O-chains from phytopatho-
genic bacteria. These are typically constituted by a
repeating unit with a rhamnanic backbone, which usually
bears a single monosaccharide as branch.6 One of the
most interesting O-chain structure is that from Xanth-
omonas campestris pv. campestris (Xcc) strain 8004, a
Almost 80% of the Gram-negative outer membrane cell
surface is covered by lipopolysaccharides (LPSs), which
are amphiphilic macromolecules consisting in three dif-
ferent domains: a lipid part (Lipid-A), an oligosaccharide
region (Core), and a polysaccharide portion (O-specific
chain, or simply O-chain).1 LPSs are highly involved in
bacterial pathogenesis both in animals and in plants: the
mechanisms of interaction between bacteria and eukary-
otic hosts cells have been addressed by several studies
on Gram-negative bacteria that are pathogenic for ani-
mals and humans,2 whereas very little is known about
LPS-plant interactions to date. One of the most widely
studied effects of LPSs on plant cells is the ability,
induced by avirulent bacteria, to prevent the hypersensi-
tive response (HR), a programmed cell death response
triggered by live bacteria. The mechanism of this effect,
usually named as localized induced resistance (LIR),3 is
(3) Erbs, G.; Newman, M.-A. Mol. Plant Pathol. 2003, 4, 421-425.
(4) Zeidler, D.; Za¨hringer, U.; Gerber, I.; Dubery, I.; Hartung, T.;
Bors, W.; Hutzler, P.; Durner, J. Proc. Natl. Acad. Sci. U.S.A. 2004,
101, 15811-15816.
(5) 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, 2414-2416.
(6) Corsaro, M. M.; De Castro, C.; Molinaro, A.; Parrilli, M. Recent
Res. Dev. Phytochem. 2001, 5, 119-138.
* Corresponding author. Phone: +39-81-674146. Fax: +39-81-
674393.
(1) Caroff, M.; Karibian, D. Carbohydr. Res. 2003, 338, 2431-2447.
(2) Endotoxins in health and disease; Brade, H., Opal, S. M., Vogel,
S., Morrison, D. C., Eds.; Marcel Dekker: New York, 1999.
10.1021/jo051153d CCC: $30.25 © 2005 American Chemical Society
Published on Web 09/01/2005
8064
J. Org. Chem. 2005, 70, 8064-8070

Synthesis of the Repeating Unit of O-Antigen from Xcc
FIGURE 1. Structure of the repeating unit of the O-antigen from Xcc and of the related compounds that have been synthesized.
pathogen of cruciferous crops that is the causative agent
of black rot, a disease of worldwide importance.7 Ad-
ditionally, a very recent study has demonstrated the
effectiveness of both Lipid-A and oligosaccharides ex-
tracted from this bacterium to be active in LIR triggering
and, moreover, with two independent mechanisms.8
The trisaccharide repeating unit of the O-chain from
Xcc 8004 consists of a D-rhamnose disaccharide backbone
with a 3-acetamido-3,6-dideoxy-D-galactopyranose (D-
Fucp3NAc) unit as branch (Figure 1).9 The presence of a
â-rhamnosidic linkage, the “steric crowd” on the 1,2-cis-
diglycosylated D-rhamnose unit, and finally the noncom-
mercial availability of both D-rhamnose and D-Fucp3NAc
surely render the synthesis of oligosaccharides related
to this O-chain challenging. In this article is reported the
synthesis of compound 1, the methyl glycoside of the
trisaccharide repeating unit of the O-antigen from Xcc,
from the fully protected trisaccharide 2. The latter was
equipped with a suitable protecting-group pattern, which
might allow its further elongation to higher oligosaccha-
rides.
Recently, Crich developed three different â-rhamnosyla-
tion methods,11 one of which is specific for D-rhamnose
and employs a (2-(2-iodophenyl)ethylthiocarbonyl)ben-
zylidene-protected D-mannose donor, which is coupled
with high â-stereoselectivity; the benzylidene cycle is
then reductively cleaved to give regioselectively the
6-deoxy functionality of â-D-rhamnose.11b Even if this
protocol has been already successfully used for the
synthesis of a tetrasaccharide containing two units of
D-rhamnose,12 we preferred to explore the possibility to
apply a “nonbenzylidene requiring” method of â-D-man-
nosylation to the D-rhamnose series. Among such proto-
cols,13 the attention was focused on the use of a 2-O-
sulfonate group whose electron-withdrawing effect was
already demonstrated to be â-directing in glycosylation
with L-rhamno-chlorides14 and more recently extended
to thioglycosides.11c Among the several different sulfonate
groups already reported for this purpose, the benzylsul-
fonyl has been recently exploited by the Schmidt group
on a â-glycosylation of a thioglycoside and a trichloro-
acetimidate mannosyl donor.13f Since a benzylsulfonyl
group can be very easily installed on a hydroxyl function
and selectively cleaved in the presence of ether-based
protecting groups,15 its use as both â-directing and
Results and Discussion
The 1,2-cis-diglycosylated moiety present on the â-D-
rhamnose unit clearly suggested a synthetic approach in
which the â-D-rhamnosidic linkage is first built up to give
a rhamnose disaccharide with an orthogonal protecting-
group pattern that allows the selective deprotection on
the O-2B position and the subsequent R-coupling with a
suitable D-Fucp3NAc donor. The manno configuration of
D-rhamnose makes its â-stereoselective coupling part of
a synthetic challenge on which several research groups
focused their attention during the last two decades.10
Nevertheless, many protocols for â-mannosylation re-
quire the use of 4,6-benzylidene-protected donor and
therefore are not applicable to the rhamnose series.
(11) (a) Crich, D.; Vinod, A. U.; Picione, J. J. Org. Chem. 2003, 68,
8453-8458. (b) Crich, D.; Yao, Q. Org. Lett. 2003, 5, 2189-2191. (c)
Crich, D.; Picione, J. Org. Lett. 2003, 5, 781-784.
(12) Crich, D.; Yao, Q. J. Am. Chem. Soc. 2004, 126, 8232-8236.
(13) (a) De Meo, C.; Kamat, M. N.; Demchenko, A. V. Eur. J. Org.
Chem. 2005, 706-711. (b) Crich, D.; Hutton, T. K.; Banerjee, A.;
Jayalath, P.; Picione, J. Tetrahedron: Asymmetry 2005, 16, 105-119.
(c) Chayajarus, K.; Chambers, D. J.; Chughtai, M. J.; Fairbanks, A. J.
Org. Lett. 2004, 6, 3797-3800. (d) Toshima, K.; Nagai, H.; Kasumi,
K.; Kawahara, K.; Matsumura, S. Tetrahedron 2004, 60, 5331-5339.
(e) Hashihayata, T.; Mandai, H.; Mukaiyama, T. Chem. Lett. 2003, 32,
442-443. (f) Abdel-Raman, A. A.-H.; Jonke, S.; El Ashry, E. S. H.;
Schmidt, R. R. Angew. Chem., Int. Ed. 2002, 41, 2972-2974. (g) Aloui,
M.; Chambers, D. J.; Cumpstey, I.; Fairbanks, A. J.; Redgrave, A. J.;
Seward, C. M. P. Chem.-Eur. J. 2002, 8, 2608-2621. (h) Nagai, H.;
Kawahara, K.; Matsumura, S.; Toshima, K. Tetrahedron Lett. 2001,
42, 4159-4162. (i) Chung, S.-K.; Park, K.-H. Tetrahedron Lett. 2001,
42, 4005-4007. (j) Singh, G.; Vankayalapati, H. Tetrahedron: Asym-
metry 2000, 11, 125-138. (k) Plante, O. J.; Palmacci, E. R.; Seeberger,
P. H. Org. Lett. 2000, 2, 3841-3843. (l) Hodosi, G.; Kovac, P. J. Am.
Chem. Soc. 1997, 119, 2335-2336. (m) Kim, W.-S.; Sasai, H.; Shiba-
saki, M. Tetrahedron Lett. 1996, 37, 7797-7800.
(7) Vauterin, L.; Hoste, B.; Yang, P.; Alvarez, A.; Kersters, K.;
Swings, J. G. In Xanthomonas; Swings, J. G., Civerolo, E. L., Eds.;
Chapman and Hall: London, 1993; pp 157-192.
(8) Silipo, A.; Molinaro, A.; Sturiale, L.; Dow, J. M.; Erbs, G.;
Lanzetta, R.; Newman, M.-A.; Parrilli, M. J. Biol. Chem., in press.
(9) Molinaro, A.; Silipo, A.; Lanzetta, R.; Newman, M.-A.; Dow, J.
M.; Parrilli, M. Carbohydr. Res. 2003, 338, 277-281.
(10) Gridley, J. J.; Osborn, H. M. I. J. Chem. Soc., Perkin Trans. 1
2000, 1471-1491
(14) Srivastava, V. K.; Schuerch, C. Carbohydr. Res. 1982, 100, 411-
417.
(15) Awad, L. F.; El Ashry, E. S.; Schuerch, C. Bull. Chem. Soc. Jpn.
1986, 59, 1587-1592.
J. Org. Chem, Vol. 70, No. 20, 2005 8065

Bedini et al.
to be â-directing glycosyl donor,13k hemi-acetal 5 was
converted into the diphenyl phosphate donor 8 by treat-
ment with diphenyl chlorophosphate in CH2Cl2 at -10
°C in the presence of DMAP.19 Analogously to 6, 8 was
demonstrated to be highly unstable by TLC analysis and
chromatography on neutral alumina support, which did
not allow the recovery of any glycosyl phosphate. Thus,
crude 8 (R/â ) 2.5:1) was directly subjected to glycosy-
lation reaction without any chromatographic purifica-
tion: upon coupling 8 and 7 with stoichiometric TMSOTf
in CH2Cl2 at -78 °C, the desired disaccharide 10 was
obtained in 58% yield. The stereoselectivity of the coup-
ling was quite low: the â-disaccharide 10â was recovered
in 31% yield, whereas 10R was recovered in 27% yield.
The configuration of the new glycosidic bond in 10R and
10â was ascertained by comparing the chemical shifts
values of H-3B and H-5B, which are upfield shifted in 10â
(H-3: 3.34 ppm; H-5: 3.24 ppm) with respect to 10R
(H-3, H-5: 3.84 ppm). To enhance the yield of the
coupling, an N-phenyltrifluoroacetimidate was chosen as
alternative leaving group on the anomeric position,20
since it leads to glycosyl donors that are more stable and
sometimes also more effective in glycosylation reactions
than trichloroacetimidate ones.21 Hemi-acetal 5 was
therefore treated with CF3C(NPh)CCl and NaH22 to give
9, after chromatography on neutral alumina, in a rather
better yield (69%; R/â ) 1:1) than 6. Coupling of 9 with
7 in CH2Cl2 at -25 °C using catalytic TMSOTf gave 10
in excellent yield and acceptable ratio of anomeric
glycosides (99%; R/â ) 2:3; 59% of isolated 10â). A slight
modification in the solvent mixture (addition of hexane
to enhance the SN2 character of the glycosyl acceptor
attack on the supposed intermediate glycosyl triflate/
oxacarbenium ion)13f afforded 10 in slightly lower yield
(see Table 1, entry 5). Cleavage of the benzylsulfonyl
protecting group on 10â with sodium amide in DMF
afforded the disaccharide acceptor 11 (62%).
The installation of the D-Fucp3NAc unit was first
attempted with the known N-phenyltrifluoroacetimidate
12, the sole D-Fucp3NAc donor reported to date.23 The
glycosylation with TMSOTf in an R-stereodirecting ter-
nary solvent mixture (4:1:1 dioxane/DME/toluene)24 af-
forded the trisaccharide 13 in only 17% yield (see Table
2, entry 1).
Since 12 has been already reported to glycosylate
selectively armed acceptors,21b its coupling with 11, whose
sterical crowd around the hydroxyl function renders it a
quite disarmed acceptor, proceeds not surprisingly with
low yield. A different D-Fucp3NAc donor was therefore
required. Since thiofucosides have been already reported
to act as efficient donors in glycosylations in which
glycosyl trihaloacetimidates failed,25 the synthesis of a
D-Fucp3NAc thioglycoside was attempted (Scheme 2).
SCHEME 1. Synthesis of 11a
a (a) i. BuSnO, 10:1 benzene/methanol, 60 °C, 90 min; ii. TBAB,
AllBr, toluene, 65 °C, 60 min, 82% over two steps; (b) i. BnSO2Cl,
py, rt, 45 min; ii. 73:26:1 Ac2O/AcOH/H2SO4, rt, 3 h; iii. hydrazine
acetate, DMF, rt, 2 h, 57% over three steps (R/â ) 3:1); (c) Cl3CCN,
DBU, CH2Cl2, rt, 2 h, 35%; (d) (PhO)2POCl, DMAP, CH2Cl2, -30
to -10 °C, overnight; (e) CF3C(NPh)Cl, NaH, CH2Cl2, 0 °C, 4 h,
69% (R/â ) 1:1); (f) see Table 1, entry 3; (g) see Table 1, entries 4
and 5; (h) NaNH2, DMF, rt, 4 days, 62%.
temporary protecting group could be advantageous in the
synthesis of the target compound 1.
Thus, to prepare a suitable 2-O-benzylsulfonylated
D-rhamnosyl donor, the known methyl 4-O-benzyl-R-D-
rhamnopyranoside 316 was regioselectively allylated at
position O-3 with the stannylidene method, giving 4 in
82% yield (Scheme 1), and this alcohol was then subjected
to benzylsulfonylation with BnSO2Cl in pyridine; without
any intermediate chromatography, subsequent acetolysis
and cleavage of the anomeric acetate gave the hemi-
acetal 5 in 57% yield after three steps.
Conversion of the hemi-acetal into trichloroacetimidate
surprisingly proceeded with low yield (35%). This result
was explained with the high instability of compound 6,
which was degraded during the chromatographic puri-
fication on a neutral alumina support. Despite this low
stability, we attempted the coupling of 6 with the
D-rhamnose acceptor 7, which was synthesized in one
step from 3 according to the known phase-transfer
procedure,16,17 but the total consumption of the donor was
observed giving no disaccharide product (Table 1). To
have a glycosyl donor that was effective in glycosylate 7
and that did not degrade too quickly, alternative glyco-
sylation procedures were investigated. Gin dehydrative
coupling18 between hemi-acetal 5 and acceptor 7 was first
tested, but it did not proceed at all: no product was
detected by TLC analysis, even when the reaction was
conducted for 2 days. Actually, ESI-MS analysis revealed
the presence of a small peak related to disaccharide
formation, which was quantified in less than 10% yield
by NMR analysis. Since glycosyl phosphates are known
(19) Sabesan, S.; Neira, S. Carbohydr. Res. 1992, 223, 169-185.
(20) Yu, B.; Tao, H. Tetrahedron Lett. 2001, 42, 2405-2407; J. Org.
Chem. 2002, 67, 9099-9102.
(21) (a) Tanaka, H.; Iwata, Y.; Takahashi, D.; Adachi, M.; Takahashi,
T. J. Am. Chem. Soc. 2005, 127, 1630-1631. (b) Bedini, E.; Carabellese,
A.; Schiattarella, M.; Parrilli, M. Tetrahedron 2005, 61, 5439-5448.
(22) Adinolfi, M.; Barone, G.; Iadonisi, A.; Schiattarella, M. Tetra-
hedron Lett. 2002, 43, 5573-5577.
(23) Bedini, E.; Iadonisi, A.; Carabellese, A.; Parrilli, M. Tetrahedron
Lett. 2004, 45, 4445-4448.
(24) Adinolfi, M.; Iadonisi, A.; Ravida`, A.; Schiattarella, M. Tetra-
hedron Lett. 2004, 45, 4485-4488.
(16) Zou, W.; Sen, A. K.; Szarek, W. A.; MacLean, D. B. Can. J.
Chem. 1993, 71, 2194-2200.
(17) Fang, Y.; Kong, F.; Wang, Q. J. Carbohydr. Chem. 1987, 6, 169-
179.
(18) Garcia, B. A.; Gin, D. Y. J. Am. Chem. Soc. 2000, 122, 4269-
4279.
8066 J. Org. Chem., Vol. 70, No. 20, 2005

Synthesis of the Repeating Unit of O-Antigen from Xcc
TABLE 1. Glycosylation Reactions of Acceptor 7 To Give Disaccharide 10
entry
donor
acceptor
solvent
activator
temperature
yielda (R/â)b
1
2
3
4
5
5
6
8
9
9
7
7
7
7
7
CH2Cl2/toluene 3:1
CH2Cl2
Tf2O/Ph2SO/DTBMP
TMSOTf
-78 °C to room temperature
-50 °C to room temperature
-78 °C to -15 °C
traces
no product
58% (1:1.1)
99% (2:3)
95% (1.1:1)
CH2Cl2
TMSOTf
CH2Cl2
CH2Cl2/hexane 1:1
TMSOTf
TMSOTf
-60 °C to -25 °C
-50 °C
a Isolated yield. b Measured after isolation of the two anomers.
TABLE 2. Glycosylation Reactions of Disaccharide Acceptor 11
entry
donor
acceptor
solvent
activator
temperature
yield
product
1
2
3
4
12
11
11
11
11
dioxane/DME/toluene 4:1:1
CH2Cl2/Et2O 1:1
TMSOTf
0 °C to room temperature
17%
13
13
2
18
NIS/TfOH
NIS/TfOH
NIS/TfOH
-20 °C
-20 °C
-20 °C
traces
15%
19r
19â
CH2Cl2/Et2O 1:1
CH2Cl2/Et2O 1:1
40% (55%)a
2
a Yield calculated on reacted acceptor.
SCHEME 2. Synthesis of 1a
a (a) i. CSA, 2:7 DMF/MeC(OMe)3, 100 mbar, rt, 20 min; ii. Ac2O, py, rt, overnight; iii. 80% AcOH, rt, 10 min, 82% over three steps; (b)
Tf2O, 1:1 py/CH2Cl2, 0 °C, 45 min; (c) Na, 1:1 MeOH/CH2Cl2, rt; (d) see Table 2, entry 1; (e) i. Ac2O, py, rt, overnight; ii. EtSH, BF3‚OEt2,
rt, overnight, 79% over two steps (R/â ) 1:1); (f) AcCl, DIPEA, CH2Cl2, rt, overnight, 91%; (g) see Table 2, entries 3 and 4; (h) i. PdCl2,
1:1 CH2Cl2/MeOH, rt, overnight; ii. 0.4 M NaOMe, 1:1 CH2Cl2/MeOH, rt, 3 h; iii. H2, Pd/C, MeOH, rt, 4 days then HCOOH, ultrasound
bath, rt, 3 h, 84% over three steps.
Thus, compound 1426 was subjected to a one-pot
sequence of three reactions (orthoesterification, acetyla-
tion, and orthoester regioselective opening; 82% over
three steps) to afford the alcohol 15. Unfortunately, the
treatment of the triflate derivative of 15 with sodium in
methanol gave a complex mixture in which we detected
only traces of the desired 2,3-epoxide 16, which was
required for the subsequent insertion of the 3-amino
functionality via the intramolecular cyclization of an
epoxytrichloroacetimidate.23 For this reason, it was de-
cided to install a thioalkyl group directly on a D-Fucp3NAc
building block, whose position 3 is namely already
aminated. Thus, hemi-acetal 1723 was acetylated and
then treated with EtSH/BF3‚OEt2 in CH2Cl2 to give the
thioglycoside 18 (79%; R/â ) 1:1 as an inseparable
mixture). The NIS/TfOH mediated coupling of this donor
and acceptor 11 in 1:1 CH2Cl2/Et2O afforded only traces
of the desired R-trisaccharide 13. To enhance the yield,
a more efficient D-Fucp3NAc donor was required. Since
it has been reported that the failure of a glycosylation
can be sometimes ascribed to the inhibitory effect of a
NHAc group on the glycosyl donor27 or acceptor,28 com-
pound 18 was treated with AcCl/DIPEA in CH2Cl2 to give
(25) (a) Joosten, J. A. F.; Evers, B.; van Summeren, R. P.; Kamerling,
J. P.; Vliegenthart, J. F. G. Eur. J. Org. Chem. 2003, 3569-3586. (b)
Wei, G.; Du, Y.; Linhardt, R. J. Tetrahedron Lett. 2004, 45, 6895-
6898.
(26) Sjo¨lin, P.; George, S. K.; Bergqvist, K.-E.; Roy, S.; Svensson,
A.; Kihlberg, J. J. Chem. Soc., Perkin Trans. 1 1999, 1731-1742.
(27) Schmidt, R. R.; Kinzy, W. Adv. Carbohydr. Chem. Biochem.
1994, 50, 21-123.
(28) (a) Lucas, R.; Hamza, D.; Lubineau, A.; Bonnaffe´, D. Eur. J.
Org. Chem. 2004, 2107-2117. (b) Liao, L.; Auzanneau, F.-I. Org. Lett.
2003, 5, 2607-2610.
J. Org. Chem, Vol. 70, No. 20, 2005 8067

Bedini et al.
3-O-Allyl-2-O-benzensulfonyl-4-O-benzyl-D-rhamnopy-
ranose (5). Compound 4 (0.577 g, 1.87 mmol) was dissolved
in pyridine (12 mL), and then BnSO2Cl (0.899 g, 4.71 mmol)
was added. The solution was stirred for 45 min at room
temperature, and after that water (10 mL) was added. The
mixture was diluted with CH2Cl2 and washed with water. The
organic layer was collected, dried, and concentrated to give a
brown oil that was dissolved in Ac2O (10 mL) and cooled to 0
°C. A 25:20:0.5 v/v/v mixture of Ac2O/AcOH/H2SO4 (15 mL) was
added. The solution was allowed to gradually warm to room
temperature, and after 3 h it was diluted with CH2Cl2 and
washed with water, 1 M NaHCO3, and water again. The
organic layer was collected, dried, and concentrated to give a
residue that was dissolved in DMF (5 mL). The solution was
treated with hydrazine acetate (0.488 g, 5.10 mmol) and then
stirred for 2 h at room temperature. Then it was diluted with
CH2Cl2, washed with 5 N NaCl, dried, and concentrated. The
residue was subjected to column chromatography (4:1 to 2:1
petroleum ether/ethyl acetate) to give 5 (0.478 g, 57%; R/â )
6:1) as a yellowish oil. 1H NMR (CDCl3, 200 MHz) (R-anomer)
δ 7.50-7.27 (m, 10H), 5.96 (m, 1H), 5.36 (dd, 1H, Jvic ) 17.2
Hz, Jgem ) 1.6 Hz), 5.23 (dd, 1H, Jvic ) 10.5 Hz, Jgem ) 1.6
Hz), 5.14 (d, 1H, J1,2 ) 1.6 Hz), 4.99 (dd, 1H, J2,3 ) 2.8 Hz, J1,2
the N,N-diacetylated thioglycoside 19 in a ca. 1:1 R/â
mixture, which was then easily separated by standard
silica gel chromatography (19R: 44%; 19â: 47%). The
R-anomer gave the R-trisaccharide 2 in 15% yield,
whereas compound 19â afforded the same coupling
product in higher yield (40%) together with a 27%
recovery of unreacted acceptor 11 (55% yield based on
reacted 11) and 10% of compound 13, whose formation
is probably due to an acidic cleavage of the diamide
function to NHAc group. The R-configuration of the newly
3
formed glycosydic bond was ascertained by the JH1-H2
value (3.4 Hz). In comparison with the NHAc group, the
presence of an N,N′-diacetyl protecting group in 2 does
not increase the number of the required deprotection
steps, since conventional transesterification with NaOMe
on an NAc2 group retains one N-acetyl functionality,
which occurs in the natural repeating unit of the O-
antigen from Xcc. Thus, after a first de-O-allylation step
with PdCl2 in 1:1 MeOH/CH2Cl2, Zemple`n deacetylation
and subsequent hydrogenolysis afforded the target com-
pound 1 (84%). Interestingly, hydrogenolysis with Pd/C
in MeOH under H2 atmosphere allowed the cleavage of
only two benzyl groups, even after a prolonged period of
several days. The complete debenzylation was, however,
accomplished by transfer hydrogenation under Perlin’s
conditions.29
) 1.6 Hz), 4.92 (d, 1H, Jgem ) 11.2 Hz), 4.64 (d, 1H, Jgem
)
11.2 Hz), 4.54 (AB d, 1H, Jgem ) 14.6 Hz), 4.45 (AB d, 1H, Jgem
) 14.6 Hz), 4.23 (m, 2H), 3.91 (m, 2H), 3.37 (t, 1H, J4,3 ) J4,5
) 9.6 Hz), 1.28 (d, 3H, J6,5 ) 6.2 Hz); 13C NMR (CDCl3, 50
MHz) (R-anomer) δ 138.2, 134.2, 130.8-127.8, 117.7, 92.3, 79.8,
78.0, 75.6, 75.4, 71.5, 67.7, 57.4, 17.9. ESI-MS for C23H28O7S
(m/z): Mr (calcd) 448.16, Mr (found) 471.41 (M + Na)+. Anal.
Calcd: C, 61.59; H, 6.29. Found: C, 61.70; H, 6.26.
Conclusion
3-O-Allyl-2-O-benzensulfonyl-4-O-benzyl-D-rhamnopy-
ranosyl Trichloroacetimidate (6). Hemi-acetal 5 (0.459 g,
1.02 mmol) was dissolved under argon in CH2Cl2 (11 mL), and
Cl3CCN (0.565 mL, 5.60 mmol) and DBU (30 µL, 0.20 mmol)
were sequentially added. The solution was stirred at room
temperature for 2 h, and then it was concentrated to give a
residue, which, after neutral alumina (Brockman grade 1)
column chromatography (10:1 petroleum ether/ethyl acetate),
afforded 6 (0.214 g, 35%) as a colorless oil. [R]D +6.1 (c 1.0,
CH2Cl2). 1H NMR (CDCl3, 200 MHz) δ 8.65 (s, 1H), 7.44-7.20
(m, 10H), 6.14 (bs, 1H), 5.92 (m, 1H), 5.31 (dd, 1H, Jvic ) 17.2
Hz, Jgem ) 1.6 Hz), 5.19 (dd, 1H, Jvic ) 10.5 Hz, Jgem ) 1.6
Hz), 5.05 (bd, 1H, J2,3 ) 2.8 Hz), 4.89 (d, 1H, Jgem ) 11.1 Hz),
4.61 (d, 1H, Jgem ) 11.1 Hz), 4.52 (d, 1H, Jgem ) 14.7 Hz), 4.47
(m, 3H), 4.35-4.10 (m, 5H), 3.87 (m, 2H), 3.44 (m, 3H), 1.29
(d, 3H, J6,5 ) 6.2 Hz), 1.23 (d, 1H, J6,5 ) 6.2 Hz); 13C NMR
(CDCl3, 50 MHz) 159.7, 137.7, 133.9, 130.7-127.5, 118.4, 95.0,
78.8, 76.5, 75.6, 75.4, 71.5, 70.9, 57.6, 17.7. Anal. Calcd: C,
50.64; H, 4.76; N, 2.36. Found: C, 50.58; H, 4.73; N, 2.37.
3-O-Allyl-2-O-benzensulfonyl-4-O-benzyl-D-rhamnopy-
ranosyl N-Phenyltrifluoroacetimidate (9). A mixture of
5 (0.544 g, 1.21 mmol) and freshly activated 4 Å molecular
sieves was suspended under argon in CH2Cl2 (25 mL) and
cooled to 0 °C. CF3C(NPh)Cl (195 µL, 1.58 mmol) and NaH
(60% oil suspension; 86 mg, 2.14 mmol) were sequentially
added. The mixture was stirred at 0 °C for 4 h, and then it
was filtered over a Celite pad and concentrated. Neutral
alumina (Brockman grade 1) column chromatography (13:1
petroleum ether/ethyl acetate) on the residue afforded 9 (0.514
g, 69%; R:â ) 1:1) as a yellowish oil. 1H NMR (CDCl3, 200 MHz)
δ 7.45-6.79 (m, 15H), 6.08-5.90 (m, 3H), 5.79 (bm, 1H), 5.35
(2 dd, 2H, Jvic ) 17.4 Hz, Jgem ) 1.6 Hz), 5.21 (m, 3H), 5.11
(dd, 1H), 4.95 (d, 1H, Jgem ) 10.8 Hz), 4.91 (d, 1H, Jgem ) 10.8
Hz), 4.67 (d, 1H, Jgem ) 10.8 Hz), 4.62 (d, 1H, Jgem ) 10.8 Hz),
4.57 (d, 1H, Jgem ) 14.4 Hz), 4.47 (m, 3H), 4.35-4.10 (m, 5H),
3.87 (m, 2H), 3.44 (m, 3H), 1.29 (d, 3H, J6,5 ) 6.2 Hz), 1.23 (d,
1H, J6,5 ) 6.2 Hz); 13C NMR (CDCl3, 50 MHz) δ 135.9, 130.9,
130.7, 128.7-123.7, 119.3, 119.2, 94.0, 93.4, 78.8, 78.7, 77.5,
75.6, 75.5, 75.1, 72.9, 71.7, 71.1, 70.6, 57.6, 17.7. ESI-MS for
C31H32F3NO7S (m/z): Mr (calcd) 619.19, Mr (found) 619.65 (M
In conclusion, the first synthesis of a methyl trisac-
charide corresponding to the repeating unit of the O-
antigen from Xanthomonas campestris pv. campestris
8004 has been reported. It is noteworthy that the
proposed synthetic approach yields the orthogonally
protected trisaccharide building block 2, whose allyl
protecting group could chemoselectively be cleaved to give
a trisaccharide acceptor. This one would be ready for
successive glycosylations to higher oligosaccharide frag-
ments of the O-antigen from Xcc, suitable, as 1, for
phytopathological structure-activity studies.
Experimental Section
Methyl 3-O-Allyl-4-O-benzyl-r-D-rhamnopyranoside (4).
Diol 315 (1.337 g, 4.99 mmol) was dissolved in 10:1 benzene/
methanol (34 mL), and Bu2SnO (1.565 g, 6.29 mmol) was then
added. After being stirred at 60° C for 90 min, the solvent was
evaporated. Bu4NBr (1.609 g, 4.99 mmol) was added to the
residue under argon. The mixture was suspended in toluene
(22 mL), AllBr (4.63 mL, 54.8 mmol) was then added, and
stirring was conducted at 65 °C for 60 min, after that the
solvent was evaporated. A column chromatography (6:1 pe-
troleum ether/ethyl acetate) on the residue afforded 4 (1.258
g, 82%) as a yellowish oil. [R]D +54.0 (c 0.9, CH2Cl2). 1H NMR
(CDCl3, 200 MHz) δ 7.33 (m, 5H), 5.93 (m, 1H), 5.32 (dd, 1H,
Jvic ) 17.0 Hz, Jgem ) 1.6 Hz), 5.20 (dd, 1H, Jvic ) 10.4 Hz,
Jgem ) 1.6 Hz), 4.87 (d, 1H, Jgem ) 10.8 Hz), 4.69 (bs, 1H),
4.62 (d, 1H, Jgem ) 10.8 Hz), 4.16 (m, 2H), 3.99 (bd, 1H, J2,3
)
3.0 Hz), 3.68 (m, 2H), 3.40 (t, 1H, J4,3 ) J4,5 ) 9.6 Hz), 3.34 (s,
3H), 2.48 (s, 1H), 1.31 (d, 3H, J6,5 ) 6.2 Hz); 13C NMR (CDCl3,
50 MHz) δ 138.4, 134.5, 128.3-127.7, 117.3, 100.0, 79.8, 79.6,
75.3, 70.9, 68.6, 67.0, 54.7, 17.9. ESI-MS for C17H24O5 (m/z):
Mr (calcd) 308.16, Mr (found) 331.39 (M + Na)+. Anal. Calcd:
C, 66.21; H, 7.84. Found: C, 66.50; H, 7.87.
(29) Rao, V. S.; Perlin, A. S. Carbohydr. Res. 1980, 83, 175-177.
8068 J. Org. Chem., Vol. 70, No. 20, 2005

Synthesis of the Repeating Unit of O-Antigen from Xcc
+ Na)+. Anal. Calcd: C, 60.09; H, 5.21; N, 2.26. Found: C,
74.7, 72.5, 71.6, 70.5, 68.8, 67.5, 54.7, 18.1, 17.9. ESI-MS for
C37H46O9 (m/z): Mr (calcd) 634.31, Mr (found) 657.47 (M +
Na)+. Anal. Calcd: C, 70.01; H, 7.30. Found: C, 69.90; H, 7.30.
60.16; H, 5.22; N, 2.25.
Methyl (3-O-Allyl-2-O-benzensulfonyl-4-O-benzyl-â-D-
rhamnopyranosyl)-(1 f 3)-2,4-di-O-benzyl-r-D-rhamnopy-
ranoside (10). A mixture of acceptor 7 (0.182 g, 0.51 mmol)
and donor 9 (0.362 g, 0.58 mmol) was coevaporated three times
with toluene (5 mL). The residue was mixed with freshly
activated AW-300 4 Å molecular sieves and suspended under
argon in CH2Cl2 (20 mL). The mixture was cooled to -60 °C,
and an 80 µM solution of TMSOTf in CH2Cl2 (75 µL, 6.0 µmol)
was added. The temperature was allowed to gradually rise to
-25 °C. After 4 h the mixture was neutralized by adding Et3N,
then filtered over Celite, and concentrated to give a residue
that after column chromatography (9:1 to 6:1 petroleum ether/
ethyl acetate) afforded, as first eluted compound, 10R (0.165
Ethyl 2,4-Di-O-acetyl-1-thio-â-D-fucopyranoside (15).
Triol 14 (0.775 g, 3.72 mmol) was dissolved in 2:7 v/v DMF/
MeC(OMe)3 (9.0 mL), CSA (80 mg, 0.34 mmol) was then added,
and the solution was evacuated at 100 mbar for 20 min. Then
pyridine (7.0 mL) and Ac2O (7.0 mL) were sequentially added.
The solution was stirred overnight at room temperature, then
coevaporated four times with toluene (10 mL each). The
residue was dissolved in 80% AcOH, and the solution was
stirred at room temperature for 10 min. Then it was coevapo-
rated two times with toluene (5 mL each). The residue was
subjected to column chromatography (5:2 petroleum ether/
ethyl acetate) to give 15 (0.893 g, 82%) as a white solid. [R]D
1
1
g, 41%) as a yellowish oil. [R]D -6.8 (c 1.0, CH2Cl2). H NMR
-2.4 (c 1.0, CH2Cl2). H NMR (CDCl3, 200 MHz) δ 5.17 (dd,
(CDCl3, 200 MHz) δ 7.40-7.24 (m, 20H), 5.91 (m, 1H), 5.27
1H, J4,3 ) 3.2 Hz, J4,5 ) 0.8 Hz), 4.97 (t, 1H, J2,3 ) J2,1 ) 9.6
Hz), 4.37 (d, 1H, J1,2 ) 10.0 Hz), 3.79 (dd, 1H, J3,2 ) 9.6 Hz,
J3,4 ) 3.2 Hz), 3.71 (dq, J5,6 ) 6.4 Hz, J5,4 ) 0.8 Hz), 2.67 (dq,
2H, Jvic ) 7.2 Hz, Jgem ) 3.2 Hz), 2.14, 2.07 (2s, 6H), 1.23 (t,
3H, Jvic ) 7.2 Hz), 1.16 (d, 3H, J6,5 ) 6.4 Hz); 13C NMR (CDCl3,
50 MHz) δ 171.3, 171.0, 83.1, 73.4, 73.1, 72.4, 71.0, 24.1, 20.9,
20.8, 16.6, 14.7. ESI-MS for C12H20O6S (m/z): Mr (calcd) 292.10,
Mr (found) 292.21 (M + Na)+. Anal. Calcd: C, 49.30; H, 6.90.
Found: C, 49.35; H, 6.85.
(dd, 1H, Jvic ) 17.4 Hz, Jgem ) 1.6 Hz), 5.14 (dd, 1H, Jvic
)
10.6 Hz, Jgem ) 1.6 Hz), 5.07 (bd, 1H, J2,3 ) 2.7 Hz), 4.91 (d,
1H, Jgem ) 11.1 Hz), 4.80 (d, 1H, Jgem ) 10.8 Hz), 4.66 (d, 1H,
J1,2 ) 1.5 Hz), 4.64 (bs, 1H), 4.62 (d, 1H, Jgem ) 11.1 Hz), 4.57
(d, 1H, Jgem ) 10.8 Hz), 4.42 (d, 1H, Jgem ) 14.1 Hz), 4.35 (d,
1H, Jgem ) 14.1 Hz), 4.17 (m, 1H), 4.02 (m, 2H, H-3A), 3.84 (m,
2H), 3.70 (bd, 1H, J2,3 ) 2.7 Hz), 3.63 (dq, 1H, J5,4 ) 9.4 Hz,
J5,6 ) 6.2 Hz), 3.57 (t, 1H, J4,5 ) J4,3 ) 9.4 Hz), 3.36 (t, 1H, J4,5
) J4,3 ) 9.8 Hz), 3.30 (s, 3H), 1.26 (m, 6H); 13C NMR (CDCl3,
50 MHz) δ 138.4, 138.0, 134.3, 130.9, 128.7-127.8, 117.7, 99.1,
98.3, 80.5, 79.7, 78.4, 77.4, 77.3, 75.3, 75.2, 72.7, 71.3, 68.6,
67.9, 57.5, 54.7, 17.9. ESI-MS for C44H52O11S (m/z): Mr (calcd)
788.32, Mr (found) 810.91 (M + Na)+. Anal. Calcd: C, 66.98;
H, 6.64. Found: C, 66.88; H, 6.62.
Ethyl 3-Acetamido-4-O-acetyl-2-O-benzyl-1-thio-D-fu-
copyranoside (18). Hemi-acetal 1722 (100 mg, 297 µmol) was
dissolved in pyridine (1.5 mL), and Ac2O (2.0 mL) was added.
The solution was stirred overnight at room temperature, then
coevaporated twice with toluene (10 mL). The residue was
dissolved in CH2Cl2, washed with 1 M HCl and 0.2 M NaHCO3,
dried, and concentrated to give a residue that was then
dissolved in CH2Cl2 (2.0 mL) and treated with EtSH (25 µL,
0.34 mmol) and BF3‚OEt2 (76 µL, 0.60 mmol). After being
stirred overnight at room temperature the mixture was diluted
with CH2Cl2 (40 mL), washed with 1 M KOH (50 mL) and
water (50 mL), dried, and concentrated. The residue was
subjected to column chromatography (1:1 petroleum ether/
ethyl acetate) to afford 18 (90 mg, 79%; R/â ) 1:1) as a white
foam. 1H NMR (CDCl3, 200 MHz) δ 7.36 (m, 10H), 5.57 (d,
1H, J1,2 ) 5.0 Hz), 5.33 (d, 1H, J4,3 ) 2.2 Hz), 5.20 (d, 1H, J4,3
) 2.4 Hz), 5.02-4.87 (m, 2H), 4.86 (d, 1H, Jgem ) 11.4 Hz),
Second eluted compound 10â (0.237 g, 59%) was recovered
as a yellowish oil. [R]D -22.1 (c 0.8, CH2Cl2). 1H NMR (CDCl3,
200 MHz) δ 7.40-7.25 (m, 20H), 5.99 (m, 1H), 5.39 (dd, 1H,
Jvic ) 17.4 Hz, Jgem ) 1.8 Hz), 5.23 (dd, 1H, Jvic ) 10.5 Hz,
Jgem ) 1.8 Hz), 5.04 (d, 1H, J2,3 ) 2.1 Hz), 4.97 (d, 1H, Jgem
10.8 Hz), 4.92 (d, 1H, Jgem ) 10.8 Hz), 4.79 (d, 1H, Jgem
)
)
12.0 Hz), 4.75 (bs, 1H), 4.65 (d, 1H, Jgem ) 12.0 Hz), 4.61 (d,
1H, Jgem ) 10.8 Hz), 4.52 (d, 3H), 4.44 (bs, 1H), 4.34 (dd,
1H, Jgem ) 14.4 Hz, Jvic ) 6.6 Hz), 4.11 (m, 2H, H-3A), 3.75 (t,
1H, J2,1 ) J2,3 ) 5.8 Hz), 3.63 (dq, 1H, J5,4 ) 9.6 Hz, J5,6
)
6.2 Hz), 3.55 (t, 1H, J4,5 ) J4,3 ) 9.6 Hz), 3.34 (m, 5H), 3.24
(dq, 1H, J5,4 ) 9.6 Hz, J5,6 ) 6.2 Hz), 1.28 (d, 6H, J6,5 ) 6.2
Hz); 13C NMR (CDCl3, 50 MHz) δ 138.8, 138.3, 134.3, 131.0,
130.9, 128.6-127.4, 117.7, 99.1, 96.8, 80.0, 79.9, 79.7, 78.1,
75.9, 75.7, 74.0, 72.8, 72.1, 71.2, 67.7, 57.9, 54.9, 18.2, 17.9.
ESI-MS for C44H52O11S (m/z): Mr (calcd) 788.32, Mr (found)
810.91 (M + Na)+. Anal. Calcd: C, 66.98; H, 6.64. Found: C,
67.11; H, 6.58.
4.78 (d, 1H, Jgem ) 12.0 Hz), 4.57 (m, 2H), 4.46 (q, 1H, J5,6
)
6.0 Hz), 4.38 (d, 1H, Jgem ) 12.0 Hz), 4.33-4.16 (m, 2H), 3.87
(dd, 1H, J2,3 ) 11.2 Hz, J2,1 ) 5.0 Hz), 3.76 (q, 1H, J5,6 ) 6.6
Hz), 3.397 (t, 1H, J2,3 ) J2,1 ) 9.8 Hz), 2.79, 2.58 (2q, 4H, Jvic
) 6.8 Hz), 2.09, 2.08 (2s, 6H), 1.82, 1.73 (2s, 6H), 1.33 (t, 6H,
Jvic ) 6.8 Hz), 1.13 (d, 3H, J6,5 ) 6.0 Hz), 1.07 (d, 3H, J6,5
)
6.6 Hz); 13C NMR (CDCl3, 50 MHz) δ 170.1-169.9, 137.6,
137.4, 128.4-128.0, 85.8, 82.8, 75.4, 74.1, 73.6, 72.6, 72.2, 72.1,
71.2, 64.9, 53.3, 49.8, 25.2, 23.6, 23.0, 20.6, 16.7, 16.1, 14.9,
14.8. ESI-MS for C19H27NO5S (m/z): Mr (calcd) 381.16, Mr
(found) 404.36 (M + Na)+. Anal. Calcd: C, 59.82; H, 7.13; N,
3.67. Found: C, 59.89; H, 7.19; N, 3.68.
Methyl (3-O-Allyl-4-O-benzyl-â-D-rhamnopyranosyl)-(1
f 3)-2,4-di-O-benzyl-r-D-rhamnopyranoside (11). A mix-
ture of 10 (0.229 g, 0.29 mmol) and NaNH2 (127 mg, 3.26
mmol) was suspended in DMF (5 mL) and stirred at room
temperature. After 24 and 48 h additional aliquots of NaNH2
(127 mg, 3.26 mmol) were added. After 4 days we added
methanol (30 mL) and then, dropwise, AcOH (3 mL). The
mixture was concentrated to give a residue that was dissolved
in CH2Cl2, washed with 1 M NaHCO3 and 5 M NaCl, dried,
and concentrated. Column chromatography (6:1 petroleum
ether/EtOAc) on the residue afforded 11 (0.114 g, 62%) as a
yellowish oil. [R]D -11.1 (c 1.0, CH2Cl2). 1H NMR (CDCl3, 200
Ethyl 3,3-Diacetamido-4-O-acetyl-2-O-benzyl-1-thio-D-
fucopyranoside (19). Compound 18 (71 mg, 186 µmol) was
dissolved under argon in CH2Cl2 (2.0 mL). This solution was
treated with DIPEA (148 µL, 0.86 mmol) and then, dropwise,
with AcCl (183 µL, 2.58 mmol). The solution was stirred
overnight at room temperature, then diluted with CH2Cl2,
washed with 1 M NaHCO3, dried, and concentrated. The
residue was subjected to column chromatography (5:1 to 3:1
petroleum ether/ethyl acetate) to give, as first eluted com-
pound, 19R (35 mg, 44%) as a white foam. [R]D +20.3 (c 2.0,
MHz) δ 7.39-7.26 (m, 15H), 5.98 (m, 1H), 5.34 (dd, 1H, Jvic
17.2 Hz, Jgem ) 1.6 Hz), 5.21 (dd, 1H, Jvic ) 10.4 Hz, Jgem
)
)
1.6 Hz), 4.95 (d, 1H, Jgem ) 10.8 Hz), 4.90 (d, 1H, Jgem ) 10.8
Hz), 4.77 (d, 1H, Jgem ) 12.4 Hz), 4.72 (d, 1H, J1,2 ) 1.9 Hz),
4.61-4.56 (m, 3H), 4.28 (bs, 1H), 4.21 (m, 2H), 4.11 (m, 1H),
3.88 (d, 1H, J2,3 ) 3.0 Hz), 3.73-3.64 (m, 2H), 3.56 (t, 1H, J4,3
) J4,5 ) 8.9 Hz), 3.45 (t, 1H, J4,3 ) J4,5 ) 9.3 Hz), 3.33 (s, 3H),
3.28 (dd, 1H, J3,4 ) 9.3 Hz, J3,2 ) 3.0 Hz), 3.19 (dq, J5,4 ) 9.3
Hz, J5,6 ) 6.2 Hz), 1.35 (d, 3H, J6,5 ) 6.2 Hz), 1.27 (d, 3H, J6,5
) 6.2 Hz); 13C NMR (CDCl3, 50 MHz) δ 138.5, 138.0, 134.8,
134.7, 128.3-127.5, 117.2, 98.6, 97.1, 81.4, 79.7, 75.5, 75.0,
1
CH2Cl2). H NMR (CDCl3, 200 MHz) δ 7.30 (m, 5H), 5.55 (d,
1H, J1,2 ) 4.0 Hz), 5.11 (d, 1H), 4.68 (m, 2H), 4.58 (d, 1H, Jgem
) 10.6 Hz), 4.47 (q, 1H, J5,6 ) 6.6 Hz), 4.26 (d, 1H, Jgem ) 10.6
Hz), 2.51 (q, 2H, Jvic ) 7.2 Hz), 2.21 (s, 6H), 2.12 (s, 3H), 1.27
(t, 3H, Jvic ) 7.2 Hz), 1.15 (d, 3H, J6,5 ) 6.6 Hz); 13C NMR
(CDCl3, 50 MHz) δ 173.9, 171.3, 137.1, 128.3-127.9, 83.6, 71.6,
71.3, 70.8, 65.7, 57.9, 27.0, 23.7, 21.0, 16.1, 14.9. ESI-MS for
C21H29NO6S (m/z): Mr (calcd) 423.17, Mr (found) 446.41 (M +
J. Org. Chem, Vol. 70, No. 20, 2005 8069

Bedini et al.
Na)+. Anal. Calcd: C, 59.55; H, 6.90; N, 3.31. Found: C, 59.56;
H, 6.87; N, 3.29.
Methyl 3-Acetamido-r-D-fucopyranosyl-(1 f 2)-â-D-
rhamnopyranosyl-(1 f 3)-r-D-rhamnopyranoside (1). To
a solution of 2 (8.6 mg, 8.6 µmol) in 1:1 CH2Cl2/MeOH (400
µL) PdCl2 (0.6 mg, 3.4 µmol) was added, and the mixture was
vigorously stirred at room temperature overnight. Then it was
filtered over a Celite pad, diluted with CH2Cl2, washed with 5
N NaCl, dried, and concentrated. The residue was then
dissolved in 1:1 CH2Cl2/MeOH (800 µL) and treated with a
0.4 M methanolic solution of NaOMe (30 µL). After 3 h of being
stirred at room temperature, the solution was neutralized with
Amberlist-15 (H+), filtered, and concentrated. The residue was
dissolved in MeOH (1.5 mL) and then added to a suspension
of 10% Pd/C (catalyst amount) in MeOH (0.5 mL). After being
stirred at room temperature for 4 days under a hydrogen
atmosphere, HCOOH (100 µL) was added and the mixture was
kept in an ultrasound bath for 3 h. Then it was filtered on
Celite and concentrated to give 1 (3.7 mg, 84% yield). [R]D +26
(c 0.2, H2O). 1H NMR (D2O, 600 MHz) δ 5.18 (d, 1H, J1,2 ) 3.6
Hz, H-1C), 4.81 (s, 1H, H-1B), 4.74 (s, 1H, H-1A), 4.54 (q, 1H,
J5,6 ) 6.4 Hz, H-5C), 4.27 (dd, 1H, J3,2 ) 9.5 Hz, J3,4 ) 3.2 Hz,
H-3C), 4.14 (bs, 1H, H-2A), 4.12 (d, 1H, J2,3 ) 3.0 Hz, H-2B),
3.92 (dd, 1H, J3,4 ) 9.8 Hz, J3,2 ) 3.0 Hz, H-3A), 3.87 (dd, 1H,
J2,3 ) 9.5 Hz, J2,1 ) 3.6 Hz, H-2C), 3.72 (m, 3H, H-3B, H-4C,
H-5A), 3.53 (t, 1H, J4,3 ) J4,5 ) 9.5 Hz, H-4A), 3.50 (t, 1H, J4,3
) J4,5 ) 9.3 Hz, H-4B), 3.41 (m, 4H, H-5B, OMe), 2.06 (s, 3H,
The second eluted compound, 19â (37 mg, 47%), was
recovered as a white foam. [R]D -45.3 (c 0.9, CH2Cl2). 1H NMR
(CDCl3, 200 MHz) δ 7.30 (m, 5H), 5.12 (bs, 1H), 4.99 (d, 1H,
Jgem ) 11.0 Hz), 4.50 (m, 3H), 4.33 (d, 1H, Jgem ) 11.0 Hz),
3.82 (q, 1H, J5,6 ) 6.0 Hz), 2.79 (dq, 2H, Jvic ) 7.6 Hz, Jgem
)
2.0 Hz), 2.23 (s, 6H), 2.10 (s, 3H), 1.33 (t, 3H, Jvic ) 7.6 Hz),
1.19 (d, 3H, J6,5 ) 6.0 Hz); 13C NMR (CDCl3, 50 MHz) δ 174.2,
171.5, 138.1, 128.3-127.4, 87.2, 74.4, 73.7, 73.5, 72.0, 62.5,
27.2, 25.0, 20.9, 16.6, 14.9. ESI-MS for C21H29NO6S (m/z): Mr
(calcd) 423.17, Mr (found) 446.39 (M + Na)+. Anal. Calcd: C,
59.55; H, 6.90; N, 3.31. Found: C, 59.59; H, 6.86; N, 3.28.
Methyl (3,3-Diacetamido-4-O-acetyl-2-O-benzyl-r-D-fu-
copyranosyl)-(1 f 2)-(3-O-allyl-4-O-benzyl-â-D-rhamnopy-
ranosyl)-(1 f 3)-2,4-di-O-benzyl-r-D-rhamnopyranoside
(2). A mixture of acceptor 11 (19.7 mg, 31.0 µmol) and donor
19â (28 mg, 66 µmol) was coevaporated three times with
toluene (1 mL). The residue was mixed with freshly activated
AW-300 4 Å molecular sieves and suspended under argon in
1:1 v/v CH2Cl2/Et2O (800 µL). NIS (16 mg, 71 µmol) was then
added under argon, the mixture was cooled to -20 °C, and a
0.60 mM solution of TfOH in CH2Cl2 (20 µL, 12 µmol) was
added. After 90 min of being stirred at -20 °C, the mixture
was filtered over Celite, diluted with CH2Cl2, washed with 10%
Na2S2O3 and 1 M NaHCO3, dried, and concentrated. The
residue was then subjected first to column chromatography
(6:1 petroleum ether/ethyl acetate) and then to HPLC (Phe-
nomenex Proteo 90A C-18 column, 250 × 10 mm; eluent:
MeOH/CH3CN/H2O 2:2:1) to afford a first eluted fraction,
containing 11 (5.4 mg, 27%), and a second fraction, which
contained 2 (12.3 mg, 40%) as a white foam. [R]D +5 (c 0.3,
CH2Cl2). 1H NMR (CDCl3, 400 MHz) δ 7.39-7.10 (m, 20H),
NHAc), 1.33 (d, 3H, J6,5 ) 6.2 Hz, H-6A), 1.31 (d, 3H, J6,5
)
6.2 Hz, H-6B), 1.18 (d, 3H, J6,5 ) 6.4 Hz, H-6C); 13C
NMR (CDCl3, 150 MHz) 174.4 (NHCOCH3), 100.5 (C-1A), 100.1
(C-1C), 96.6 (C-1B), 78.7 (C-2B), 77.0 (C-3A), 73.5 (C-3B), 72.5
(C-4B), 72.4 (C-5B), 70.5 (C-4A), 70.3 (C-4C), 68.5 (C-5A), 67.1
(C-2A, C-5C), 66.6 (C-2C), 54.7 (OMe), 51.2 (C-3C), 22.0
(NHCOCH3), 16.7, 16.6 (C-6A, C-6B), 15.3 (C-6C). ESI-MS for
C21H37NO13 (m/z): Mr (calcd) 511.23, Mr (found) 533.71 (M +
Na)+. Anal. Calcd: C, 49.31; H, 7.29; N, 2.74; Found: C, 48.79;
H, 7.47; N, 2.67.
5.92 (m, 1H), 5.76 (d, 1H, J1,2 ) 3.4 Hz), 5.29 (d, 1H, Jgem
)
18.0 Hz), 5.21 (d, 1H, Jgem ) 10.4 Hz), 5.13 (bs, 1H), 5.04 (m,
2H), 4.85 (q, 1H, J5,6 ) 6.4 Hz), 4.75 (d, 1H, Jgem ) 12.0 Hz),
4.71 (m, 2H), 4.64 (d, 1H, Jgem ) 12.0 Hz), 4.52 (m, 2H), 4.39
(m, 2H), 4.28 (d, 1H, Jgem ) 11.1 Hz), 4.24-4.07 (m, 4H), 3.67
Acknowledgment. We thank MIUR, Rome (Pro-
getti di Ricerca di Interesse Nazionale 2004, M.P.), for
the financial support. E.B. thanks Dr. Alfonso Iadonisi
for proofreading. The NMR spectra were performed at
CIMCF (Centro di Metodologie Chimico-Fisiche) of
University “Federico II” of Naples.
(m, 2H), 3.49 (t, 1H, J4,3 ) J4,5 ) 9.5 Hz), 3.44 (t, 1H, J4,3
)
J4,5 ) 9.3 Hz), 3.32 (s, 3H), 3.24 (m, 2H), 2.11 (s, 3H), 2.01 (s,
6H), 1.35 (d, 3H, J6,5 ) 6.0 Hz), 1.28 (d, 3H, J6,5 ) 6.2 Hz),
1.16 (d, 3H, J6,5 ) 6.4 Hz); 13C NMR (CDCl3, 100 MHz) 174.3,
171.5, 138.8, 138.5, 138.3, 138.0, 134.5, 130.8-126.9, 117.6,
98.9, 98.4, 95.8, 83.2, 80.6, 80.5, 75.1, 75.0, 73.3, 72.5, 72.4,
72.2, 72.1, 72.0, 71.9, 71.5, 70.2, 68.2, 65.7, 56.7, 54.7, 23.8,
21.0, 18.5, 17.9, 16.2. ESI-MS for C56H69NO15 (m/z): Mr (calcd)
995.47, Mr (found) 1018.50 (M + Na)+. Anal. Calcd: C, 67.52;
H, 6.98; N, 1.41; Found: C, 67.44; H, 7.02; N, 1.40.
Supporting Information Available: 1H and 13C NMR
spectra for all new compounds. This material is available free
of charge via the Internet at http://pubs.acs.org.
JO051153D
8070 J. Org. Chem., Vol. 70, No. 20, 2005