Solid-phase synthesis of a phytoalexin elicitor pentasaccharide using a 4-azido-3-chlorobenzyl group as the key for temporary protection and catch-and-release purification

Article abstract of DOI:10.1002/ejoc.200300248

A phytoalexin elicitor pentasaccharide of the rice blast disease fungus, Pyricularia oryzae, was synthesized by a new route involving a solid-phase method, in which a 4-azido-3-chlorobenzyl (ClAzb) group was used for temporary protection of the hydroxy functions and catch-and-release purification. Thioglycosides possessing the ClAzb group were used as glycosyl donors and a macroporous polystyrene as a solid support. The saccharide chain was elongated by repeating a set of reactions: removal of the ClAzb group, glycosylation and capping of the unglycosylated hydroxy groups. Cleavage from the solid support by treatment with sodium methoxide afforded a crude pentasaccharide possessing the ClAzb group as a tag. The pentasaccharide was then purified by a catch-and-release procedure based on the specific reaction between the azido group and a solid-supported phosphane. The final deprotection of all benzyl-type protective groups gave the desired phytoalexin elicitor pentasaccharide 1 in overall 15% yield (from a solid-supported monosaccharide 4, average yield of 81% for each of a total of nine steps). The characteristic peaks of the azido group in the infrared spectrum were also useful for monitoring reactions in the solid phase. Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003.

Full text of DOI:10.1002/ejoc.200300248

FULL PAPER
Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide Using a
4-Azido-3-chlorobenzyl Group as the Key for Temporary Protection and
Catch-and-Release Purification
Kenji Egusa,[a] Shoichi Kusumoto,[a] and Koichi Fukase*[a]
Keywords: Glycosylation / Oligosaccharides / Natural products / Solid-phase synthesis
A phytoalexin elicitor pentasaccharide of the rice blast dis-
group as a tag. The pentasaccharide was then purified by a
ease fungus, Pyricularia oryzae, was synthesized by a new catch-and-release procedure based on the specific reaction
route involving a solid-phase method, in which a 4-azido-3-
chlorobenzyl (ClAzb) group was used for temporary protec-
tion of the hydroxy functions and catch-and-release purifica-
tion. Thioglycosides possessing the ClAzb group were used
between the azido group and a solid-supported phosphane.
The final deprotection of all benzyl-type protective groups
gave the desired phytoalexin elicitor pentasaccharide 1 in
overall 15% yield (from a solid-supported monosaccharide 4,
as glycosyl donors and a macroporous polystyrene as a solid average yield of 81% for each of a total of nine steps). The
support. The saccharide chain was elongated by repeating a
set of reactions: removal of the ClAzb group, glycosylation
characteristic peaks of the azido group in the infrared spec-
trum were also useful for monitoring reactions in the solid
and capping of the unglycosylated hydroxy groups. Cleav- phase.
age from the solid support by treatment with sodium methox- ( Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim,
ide afforded a crude pentasaccharide possessing the ClAzb
Germany, 2003)
Introduction
striction. This is because these resins do not require swelling
to permit reactions. In our previous work, α-selective glyco-
sylation was achieved on ArgoPoreTM by virtue of the sol-
vent effect of diethyl ether.[4,6] However, our experiments
revealed that a significant amount of glycosyl acceptors on
the resin remained intact after the glycosylation. This seems
to be due to the presence of smaller pores, where the steric
hindrance of the resin itself interferes significantly in the
glycosylation. Such unchanged acceptors can be capped by
acetylation and excluded from undesirable reactions in the
subsequent steps. Nevertheless, after the final cleavage of
the linker, the incomplete products contaminate the desired
product and often cause difficulty in purification because
their physicochemical properties are similar to those of the
desired product. An efficient purification method with a
simple procedure is required to reduce the problem of the
incomplete products.
We have also previously demonstrated that the ClAzb
group can be used as a new tag for catch-and-release purifi-
cation.[7,8] A compound bearing the ClAzb group was selec-
tively caught by a triphenylphosphane-polystyrene resin by
the specific reaction between the azido group and the poly-
mer-supported phosphane. Removal of the other com-
pounds by simple rinsing of the resin followed by release
from the resin by treatment with DDQ afforded the desired
product with high purity. We expected that this catch-and-
release purification method would be applicable to the final
purification for solid-phase synthesis of oligosaccharides.[9]
In the present study, we combined the above two method-
ologies, i.e., selective protection and catch-and-release puri-
Solid-phase synthesis of oligosaccharides is an important
topic in recent carbohydrate chemistry. Although various
approaches to this methodology have already been re-
ported,[1,2] there is a continuing demand for more efficient
and general procedures aimed at the facile preparation of a
variety of oligosaccharides for functional studies. We have
reported practical, selective protection of hydroxy functions
by the 4-azido-3-chlorobenzyl (ClAzb) group[3] and its ap-
plication to solid-phase syntheses of oligosaccharides.[4] The
ClAzb group is stable under various Lewis acidic conditions
for glycosylation, but can readily be removed by conversion
of the azido function to the corresponding iminophos-
phorane followed by oxidation using 2,3-dichloro-5,6-dicy-
anobenzoquinone (DDQ). In the solid-phase synthesis, thio-
glycosides bearing the ClAzb group for temporary protec-
tion were used as glycosyl donors in combination with an
ester-type linker,[2a,2c,2gϪ2j] and iterative glycosylation reac-
tions and removal of the ClAzb group followed by alkaline
cleavage of the linker afforded the desired oligosaccharides.
A macroporous polystyrene, ArgoPoreTM [5]
as the solid support.
An obvious advantage of macroporous resins is that reac-
tions can be effected successfully in any solvent without re-
,
was employed
[a]
Department of Chemistry, Graduate School of Science, Osaka
University
Machikaneyama 1-1, Toyonaka, Osaka 560-0043, Japan
Fax: (internat.) ϩ 81-6/6850-5419
E-mail: koichi@chem.sci.osaka-u.ac.jp
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DOI: 10.1002/ejoc.200300248
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K. Egusa, S. Kusumoto, K. Fukase
FULL PAPER
fication using the ClAzb group, for efficient multi-step, with phenylthiotrimethylsilane in the presence of zinc iod-
solid-phase synthesis of a complex oligosaccharide, a phy- ide afforded thioglycoside 9 in 84% yield.[14] The benzoyl
toalexin elicitor pentasaccharide 1, from the rice blast dis- groups at the 4- and 6-positions of 9 were selectively re-
ease fungus Pyricularia oryzae.[10Ϫ12]
moved by treatment with sodium methoxide in MeOH/
THF to give diol 10 in 92% yield and subsequent benzyl-
ation gave thioglycoside 2 in 93% yield. The ester-linker was
attached to thioglycoside 2 by coupling with xylylene glycol
using N-bromosuccinimide (NBS) and Sn(OTf)2 as activat-
ing reagents to give glycoside 11 in 75% yield, and sub-
sequent treatment with succinic anhydride afforded the re-
ducing-end unit 12 in 92% yield.
Results and Discussion
The strategy employed for the present solid-phase syn-
thesis of the phytoalexin elicitor pentasaccharide 1 is sum-
marized in Scheme 1. The saccharide was bound to the
solid support, a macroporous polystyrene, at the anomeric
position via an ester-type linker cleavable by alkali treat-
ment. The sugar chain was elongated from the reducing to
the non-reducing end. Thioglycosides 2 and 3 bearing the
ClAzb group were used as glycosyl donors. The ClAzb
group of the glycosyl donor for the final glycosylation step,
namely, thioglycoside 2, was used as a tag for the catch-
and-release purification. The pentasaccharide structure was
constructed by repeating the sequence of glycosylation and
selective cleavage of the ClAzb group on the solid support,
and the ester-type linker was then cleaved to afford penta-
saccharide 5. The crude pentasaccharide 5, which contained
incomplete oligosaccharides, was subjected to the catch-
and-release purification based on the specific reaction of
the azido group and the triphenylphosphane resin. After
the purification, the benzyl groups of the desired pentasac-
charide were removed by catalytic hydrogenation to afford
the phytoalexin elicitor pentasaccharide 1.
Synthesis of the Saccharide Components
Scheme 2. Synthesis of the saccharide units 2 and 12
Synthesis of thioglycoside 2 and the reducing end unit 12
having the ester-linker moiety is summarized in Scheme 2.
1,2:5,6-Diisopropylideneglucofuranose 6 was treated with
Synthesis of thioglycoside 3 is illustrated in Scheme 3.
4-azido-3-chlorobenzyl bromide[3,13] and NaH to afford 7 The 6-position of diol 10 was protected with a trityl group
in 98% yield. Furanose 7 was then hydrolyzed with hydro- using trityl chloride, 2,4,6-collidine and tetrabutylam-
chloric acid, and subsequent benzoylation afforded py- monium perchlorate.[15] Then, 6-O-tritylthioglycoside 13,
ranosyl benzoate 8 (α/β ഠ 1:1) in 97% yield. Reaction of 8 obtained in 98% yield, was benzylated to afford thioglyco-
Scheme 1. Synthetic strategy of the phytoalexin elicitor pentasaccharide 1
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Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide
FULL PAPER
side 14 in 94% yield. The 6-O-trityl group was removed by
successive treatments with trifluoroacetic acid and with an
NaHCO3 solution for hydrolysis of the trifluoroacetate
partly formed to afford 6-O-free thioglycoside 15 in 86%
yield. The resulting 15 was glycosylated with trichloroaceto-
imidate 16,[16] which was prepared from -glucose by ben-
zoylation and selective cleavage of the 1-O-benzoyl group
using methylamine followed by treatment with trichloroace-
tonitrile and Cs2CO3, in the presence of Sn(OTf)2 as an
˚
activating reagent and 4-A molecular sieves at ice/salt tem-
perature to give thioglycoside 3 in 83% yield.
Scheme 3. Synthesis of saccharide unit 3
Solid-Phase Synthesis of the Phytoalexin Elicitor
Pentasaccharide
Among available macroporous polystyrene resins, Di-
[17]
aion HP20 (average pore size: 260 A) was employed as
˚
a solid support in this work instead of ArgoPoreTM used
previously, because the average pore size of Diaion HP20
is larger than that of ArgoPoreTM (average pore size: 90
[18]
˚
A).
As shown in Scheme 4, Diaion HP20 was first amino-
methylated by Merrifield’s method[19] to afford resin 19
(loading of 0.19 mmol/g was determined by Ginsen’s
method[20]). Then, carboxylic acid 12 was introduced onto
the aminomethylated resin 19 using N,N-diisopropylcar-
bodiimide and 1-hydroxybenzotriazole to give solid-sup-
ported monosaccharide 4 in quantitative yield. After cap-
ping of the possibly existing free amino groups on the resin
with acetic anhydride, dimethylaminopyridine and triethyl-
amine, the ClAzb group of solid-supported monosaccharide
4 was removed by treatment with tributylphosphane fol-
Scheme 4. Solid-phase synthesis of the pentasaccharide
These solid-phase reactions were monitored by infrared
lowed by DDQ oxidation for the subsequent glycosylation. (IR) spectroscopy (KBr disks). As shown in Figure 1, intro-
The resulting 21 was glycosylated with thioglycoside 3 using duction of carboxylic acid 12 onto the aminomethylated re-
NBS and Sn(OTf)2 as activating reagents in the presence of sin 19 (Spectrum a) was accompanied by the appearance of
4-A molecular sieves in CH2Cl2 to afford solid-supported characteristic peaks of the azido group around 2120 cmϪ1
˚
trisaccharide 22 in 37% yield from 4.
in the IR spectrum of 4 (Spectrum b). In the spectrum of
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K. Egusa, S. Kusumoto, K. Fukase
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21 after the removal of the ClAzb group of 4, the peaks with triphenylphosphane-NovaGelTM
, a triphenylphos-
attributable to the azido function completely disappeared phane-(polyethylene glycol-polystyrene-copolymer) resin,[21]
(Spectrum c). After glycosylation with the ClAzb-glycosyl to catch pentasaccharide 5 onto the resin. Iminophos-
donor 3, the peaks of the azido group again appeared in phorane 27 bearing the pentasaccharide was then separated
the IR spectrum of 22 (spectrum d).
from the by-products remaining in the liquid phase by fil-
tration. Iminophosphorane 27 was treated with DDQ to
release the saccharide moiety from the resin. Excess DDQ
was removed by Amberlyst A-26 (OHϪ form) after con-
version into the corresponding dihydroquinone using -asc-
orbic acid. 4-Amino-3-chlorobenzaldehyde derived from the
ClAzb group was readily removed by Amberlyst 15 (Hϩ
form) as described in a previous paper[7] to give pentasac-
charide 29, whose yield was, however, unexpectedly low (6%
from 4). Careful inspection led us to conclude that some
portion of iminophosphorane 27 was hydrolyzed during the
‘‘catching’’ process to release the 4-amino-3-chlorobenzyl
derivative 28 in the filtrate as detected by MS analysis. The
filtrate containing 4-amino-3-chlorobenzyl derivative 28
was recovered and treated with DDQ to remove the 4-am-
ino-3-chlorobenzyl group. Purification by reversed-phase
HPLC using the pentasaccharide 29 obtained from the
catch-and-release procedure as a reference substance af-
forded pentasaccharide 29 in 9% yield (from 4) in a pure
Figure 1. IR spectra of the solid-supported saccharides
The weight of resin 22 did not increase significantly after state. Partial hydrolysis of iminophosphorane 27 during the
repeated glycosylation (no more than ca. 50% of the theor- ‘‘catching’’ process suggests that more strictly anhydrous
etical value), nor was the peak area of the azido group in conditions are required to avoid leakage of the desired
the IR spectrum greatly enlarged: this suggests that some product into the filtrate, especially when hydrophilic po-
portion (ca. 50% of 0.19 mmol/g loading) of the acceptor lyhydroxy molecules like 5 are subjected to this purifi-
hydroxy groups on the resin remained unchanged. As de- cation method.
scribed above, this incomplete glycosylation is supposed to
be due to heterogeneity in the pore size of the macroporous
resin, i.e. introduction of the glycosyl donor was impeded
by steric hindrance of the resin itself in smaller pores.[2m,4,6]
Such unchanged hydroxy groups were capped by acety-
lation to avoid undesired glycosylation in the subsequent
steps and the resulting incomplete products were to be re-
moved in the final catch-and-release purification after cleav-
age from the resin.
The saccharide chain was further elongated by repeating
the deprotection and glycosylation. The ClAzb group of
solid-supported trisaccharide 22 was removed as described
above. Subsequent glycosylation with thioglycoside 2 using
NBS and Sn(OTf)2 afforded solid-supported tetrasacchar-
ide 24 in 31% yield (from 4). Capping of the unchanged
hydroxy groups, removal of the ClAzb group and glycosyl-
ation with thioglycoside 2 were repeated once again to af-
ford solid-supported pentasaccharide 26, which was then
treated with 0.1 mol/L sodium methoxide in methanol/THF
(1:1) to cleave the ester-linker and retrieve the saccharide
moiety from the resin. This cleaving step was also moni-
tored by IR spectra and complete cleavage was confirmed
by the disappearance of the azido group peak in the IR
spectrum of the resin (spectrum not shown). The crude
product containing pentasaccharide 5 and by-products was
then subjected to the catch-and-release purification using a
triphenylphosphane resin.
Separation of pentasaccharide 29 is outlined in Scheme 5.
The crude product was dissolved in CH2Cl2 and shaken tion
Scheme 5. Catch-and-release purification and the final deprotec-
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Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide
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Na2SO4 and then concentrated in vacuo. The residue was purified
by silica-gel column chromatography (hexane/ethyl acetate, 3:1) to
give 7 as an orange oil (8.35 g, 98%). TLC (hexane/ethyl acetate,
2:1): Rf ϭ 0.68. [α]2D5 ϭ Ϫ30.1 (c ϭ 1.00, CH2Cl2). 1H NMR
(300 MHz, CDCl3): δ ϭ 1.32 (s, 3 H, CH3), 1.39 (s, 3 H, CH3),
1.43 (s, 3 H, CH3), 1.49 (s, 3 H, CH3), 3.98 (d, J ϭ 3.0 Hz, 1 H, 3-
H), 3.99 (dd, J ϭ 5.7, 8.7 Hz, 1 H, 6-H), 4.09 (dd, J ϭ 3.0, 8.1 Hz,
1 H, 4-H), 4.12 (dd, J ϭ 6.0, 8.7 Hz, 1 H, 6-H), 4.33 (ddd, J ϭ 5.7,
6.0, 8.1 Hz, 1 H, 5-H), 4.56 (d, J ϭ 12.3 Hz, 1 H, ArCHH), 4.58
(d, J ϭ 3.6 Hz, 1 H, 2-H), 4.65 (d, J ϭ 12.3 Hz, 1 H, ArCHH),
5.89 (d, J ϭ 3.6 Hz, 1 H, 1-H), 7.12 (d, J ϭ 8.1 Hz, 1 H, ClAzb),
7.24 (dd, J ϭ 1.8, 8.1 Hz, 1 H, ClAzb), (d, J ϭ 1.5 Hz, 1 H, ClAzb)
ppm. 13C NMR (75 MHz, CDCl3): δ ϭ 25.3 (CH3), 26.2 (CH3),
Finally, the benzyl groups of the combined pentasacchar-
ide 29 were removed by catalytic hydrogenation using
Pd(OH)2 to give elicitor pentasaccharide 1 in quantitative
yield (15% from 4).
Conclusion
We have demonstrated the successful application of a no-
vel methodology to a solid-phase synthesis of oligosacchar-
ides, in which the ClAzb group was used both for tempor-
ary protection and for catch-and-release purification in
combination with thioglycosides as glycosyl donors, a mac- 26.7 (CH3), 26.8 (CH3), 67.4 (C-6), 70.7 (ArCH2), 72.0 (C-5), 81.1
(C-4), 81.4 (C-3), 82.3 (C-2), 105.0 (C-1), 108.9, 111.6, 119.2, 124.7,
126.6, 129.5, 135.2, 136.2 ppm. ESI-MS: m/z ϭ 448.1 [M ϩ Naϩ].
C19H24ClN3O6 (425.86): calcd. C 53.59, H 5.68, N 9.87; found C
53.64, H 5.68, N 9.74.
roporous polystyrene as a solid support and the ester-type
linker. The catch-and-release purification method based on
the specific reaction between the azido group and solid-sup-
ported phosphane facilitated final purification of the prod-
uct after it had been cleaved from the solid support. The
3-O-(4-Azido-3-chlorobenzyl)-2,4,6-tri-O-benzoyl-α/β-D-gluco-
ClAzb group was also useful as a tag for monitoring the pyranosyl Benzoate (8): A mixture of 7 (5.16 g, 12.1 mmol), hydro-
chloric acid (6 mol/L, 15 mL) and 1,4-dioxane (45 mL) was stirred
at room temperature overnight. The mixture was cooled in an ice
bath and neutralized with aqueous NaOH, then 1,4-dioxane was
removed in vacuo. The product was extracted three times with ethyl
acetate and the combined organic layers were washed with brine,
dried with Na2SO4 and then concentrated in vacuo. The residue
was dissolved in pyridine (30 mL) and the solution was cooled in
an ice bath. Benzoyl chloride (6.97 mL, 60.5 mmol) was added to
the solution dropwise and the mixture was stirred in situ for 3 h.
The reaction was quenched by adding cold water (100 mL) and the
product was extracted twice with ethyl acetate. The combined or-
ganic layers were washed with hydrochloric acid (1 mol/L) and
brine, dried with Na2SO4 then concentrated in vacuo. The residue
was crystallized from ethyl acetate and hexane to give the β-anomer
of 8 as pale-yellow crystals (3.30 g, 36%). The mother liquor was
concentrated in vacuo and the residue was purified by silica-gel
column chromatography (hexane/ethyl acetate, 3:1) to give a mix-
ture of two anomers of 8 as an orange gum (5.65 g, 61%, α/β ϭ
5.5:1).
reactions by IR spectroscopy. Even though improvements
in the experimental conditions may increase the efficiency
of the catch-and-release purification, the present method-
ology certainly provides a new and efficient synthesis of
various oligosaccharides.
Experimental Section
General Remarks: All chemicals and solvents used were of reagent
grade. CH2Cl2 used for glycosylations was distilled from calcium
hydride under N2. Analytical TLC was performed on Merck silica-
gel 60 F254 plates (0.25 mm) and spots were visualized with ultra-
violet light (254 nm) followed by spraying with a solution of anisal-
dehyde (0.5 mL) and concd. H2SO4 (1 mL) in acetic acid (50 mL)
and heating. Preparative TLC was performed on Merck silica-gel
60 F254 plates (0.5 mm). Column chromatography was carried out
on Wakogel C-200. Preparative HPLC was carried out using a
Waters 600 system with UV detector 2487. Melting points were
determined with a Yamato MP-21 melting point apparatus and are
uncorrected. Optical rotation was determined with a Horiba SEPA-
300 polarimeter using a 1-dm cell at 25 °C. IR spectra were re-
corded with a PerkinϪElmer Paragon 1000 FT-IR spectrometer.
NMR spectra were recorded with a Varian Mercury plus 300
(300 MHz) and Varian UNITY plus 600 (600 MHz) spectrometers,
using tetramethylsilane (in CDCl3) or H2O (in D2O) as internal
standards. Assignments were made by COSY, TOCSY, ROESY,
HMQC and HMBC experiments; the glucose residues of oligosac-
charides are designated in alphabetical order from the reducing
end. Mass spectra were recorded with a PerkinϪElmer Sciex API-
300 mass spectrometer. Microanalyses were performed in the
Microanalysis Laboratory of the Department of Chemistry, Gradu-
ate School of Science, Osaka University.
8 (α-Anomer): TLC (hexane/ethyl acetate, 2:1): Rf ϭ 0.59. 1H NMR
(300 MHz, CDCl3): δ ϭ 4.38 (dd, J ϭ 4.8, 12.3 Hz, 1 H, 6-H), 4.39
(t, J ϭ 9.9 Hz, 1 H, 3-H), 4.47 (ddd, J ϭ 3.0, 4.8, 9.9 Hz, 1 H, 5-
H), 4.55 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 4.58 (dd, J ϭ 3.0, 12.3 Hz,
1 H, 6-H), 4.63 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 5.56 (dd, J ϭ 3.9,
9.9 Hz, 1 H, 2-H), 5.68 (t, J ϭ 9.9 Hz, 1 H, 4-H), 6.63 (d, J ϭ
8.1 Hz, 1 H, ClAzb), 6.71 (d, J ϭ 3.9 Hz, 1 H, 1-H), 6.94 (dd, J ϭ
2.1, 8.1 Hz, 1 H, ClAzb), 7.07 (d, J ϭ 2.1 Hz, 1 H, ClAzb),
7.34Ϫ7.69 (m, 12 H, ArH), 7.86Ϫ8.16 (m, 8 H, ArH) ppm. 13C
NMR (75 MHz, CDCl3): δ ϭ 62.4 (C-6), 69.9 (C-4), 70.3 (C-5),
71.7 (C-2), 73.4 (ArCH2), 77.7 (C-3), 90.1 (C-1), 119.1, 124.1,
127.0, 127.9, 128.0, 128.10, 128.14, 128.38, 128.47, 128.52, 128.58,
128.76, 129.11, 129.20, 129.27, 129.34, 129.38, 129.42, 129.49,
129.7, 130.1, 132.7, 133.1, 133.2, 133.5, 135.0, 135.9, 163.8 (CϭO),
164.3 (CϭO), 164.6 (CϭO), 165.6 (CϭO) ppm. ESI-MS: m/z ϭ
784.1 [M ϩ Naϩ].
3-O-(4-Azido-3-chlorobenzyl)-1,2:5,6-di-O-isopropylidene-α-D-
glucofuranose (7): NaH (60% suspension in oil, 880 mg, 22.0 mmol)
was added to a solution of 1,2:5,6-di-O-isopropylidene-α--glucof-
8 (β-Anomer): M.p. 140Ϫ141 °C. TLC (hexane/ethyl acetate, 2:1):
uranose (6) (5.20 g, 20.0 mmol) in DMF (25 mL) at 0 °C. After Rf ϭ 0.59. [α]2D5 ϭ Ϫ7.36 (c ϭ 0.41, CH2Cl2). 1H NMR (300 MHz,
stirring for 15 min, 4-azido-3-chlorobenzyl bromide (5.42 g,
22.0 mmol) was added and the mixture was stirred in situ for a
further 3 h. The reaction was quenched by adding cold water
CDCl3): δ ϭ 4.14 (t, J ϭ 8.7 Hz, 1 H, 3-H), 4.24 (ddd, J ϭ 3.0,
4.8, 9.6 Hz, 1 H, 5-H), 4.41 (dd, J ϭ 4.8, 12.0 Hz, 1 H, 6-H), 4.54
(s, 2 H, ArCH2), 4.61 (dd, J ϭ 3.0, 12.0 Hz, 1 H, 6-H), 5.68 (dd,
(50 mL) and the product was extracted twice with ethyl acetate. J ϭ 8.7, 9.6 Hz, 1 H, 4-H), 5.69 (dd, J ϭ 7.8, 8.7 Hz, 1 H, 2-H),
The combined organic layers were washed with brine, dried with 6.16 (d, J ϭ 7.8 Hz, 1 H, 1-H), 6.62 (d, J ϭ 7.8 Hz, 1 H, ClAzb),
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K. Egusa, S. Kusumoto, K. Fukase
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6.94 (dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 7.05 (d, J ϭ 2.1 Hz, 1 H,
(5.00 g, 9.23 mmol) and benzyl bromide (2.41 mL, 20.3 mmol) in
ClAzb), 7.34Ϫ7.61 (m, 12 H, ArH), 7.92Ϫ8.01 (m, 8 H, ArH) ppm. DMF (20 mL) at 0 °C and the mixture was stirred for 2 h. The
13C NMR (75 MHz, CDCl3): δ ϭ 62.6 (C-6), 70.0 (C-4), 71.8 (C-
reaction was quenched by adding cold water (50 mL) and the prod-
2), 72.8 (C-5), 73.0 (ArCH2), 80.3 (C-3), 92.3 (C-1), 119.1, 127.1, uct was extracted twice with ethyl acetate. The combined organic
127.9, 128.12, 128.15, 128.6, 129.1, 129.3, 129.36, 129.40, 129.6, layers were washed with brine, dried with Na2SO4 and then concen-
129.8, 132.6, 133.1, 133.2, 133.4, 134.8, 136.0, 164.2 (CϭO), 164.31
trated in vacuo. The residue was purified by silica-gel column chro-
(CϭO), 164.35 (CϭO), 165.6 (CϭO) ppm. ESI-MS: m/z ϭ 784.1 matography (hexane/ethyl acetate, 5:1 Ǟ 2:1) to give 2 as white
[M ϩ Naϩ]. C41H32ClN3O10 (762.17): calcd. C 64.61, H 4.23, N crystals (6.20 g, 93%). M.p. 118Ϫ119 °C. TLC (hexane/ethyl acet-
1
5.51; found C 64.55, H 4.17, N 5.46.
ate, 2:1): Rf ϭ 0.74. [α]2D5 ϭ ϩ66.9 (c ϭ 0.51, CH2Cl2). H NMR
(300 MHz, CDCl3): δ ϭ 3.59 (m, 1 H, 5-H), 3.70Ϫ3.78 (m, 3 H, 3-
H, 4-H, 6-H), 3.82 (dd, J ϭ 2.1, 10.8 Hz, 1 H, 6-H), 4.48 (d, J ϭ
11.7 Hz, 1 H, ArCHH), 4.55 (d, J ϭ 11.7 Hz, 1 H, ArCHH), 4.62
(d, J ϭ 11.4 Hz, 2 H, ArCHH), 4.66 (d, J ϭ 11.7 Hz, 1 H,
ArCHH), 4.74 (d, J ϭ 9.9 Hz, 1 H, 1-H), 4.75 (d, J ϭ 11.1 Hz, 1
H, ArCHH), 5.19 (dd, J ϭ 9.3, 9.9 Hz,1 H, 2-H), 6.71 (d, J ϭ
8.1 Hz, 1 H, ClAzb), 6.97 (dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 7.07
(d, J ϭ 1.8 Hz, 1 H, ClAzb), 7.16Ϫ7.35 (m, 13 H, ArH), 7.40Ϫ7.47
(m, 4 H, ArH), 7.55Ϫ7.62 (m, 1 H, ArH) 7.92Ϫ7.96 (m, 2 H, ArH)
ppm. 13C NMR (75 MHz, CDCl3): δ ϭ 68.7 (C-6), 71.9 (C-2), 73.4
(ArCH2), 73.8 (ArCH2), 75.0 (ArCH2), 77.8 (C-4), 79.3 (C-5), 84.4
(C-3), 85.9 (C-1), 119.2, 124.3, 127.3, 127.4, 127.5, 127.6, 127.7,
128.2, 128.3, 128.6, 129.4, 129.5, 129.9, 132.4, 133.0, 135.6, 136.0,
137.5, 137.9, 164.7 (CϭO) ppm. ESI-MS: m/z ϭ 744.1 [M ϩ Naϩ].
C40H36ClN3O6S (722.25): calcd. C 66.52, H 5.02, N 5.82; found C
66.47, H 4.95, N 5.76.
Phenyl 3-O-(4-Azido-3-chlorobenzyl)-2,4,6-tri-O-benzoyl-1-thio-β-D-
glucopyranoside (9): A mixture of 8 (5.50 g, 7.22 mmol, α/β ϭ
5.5:1), phenylthiotrimethylsilane (2.05 mL, 10.8 mmol), ZnI2
(1.15 g, 3.61 mmol) and CH2Cl2 (20 mL) was stirred at room tem-
perature overnight. The mixture was filtered and the filtrate was
washed with hydrochloric acid (1 mol/L) and brine, dried with
Na2SO4 and then concentrated in vacuo. The residue was crys-
tallized from hexane/ethyl acetate to give 9 as white crystals (4.55 g,
84%). M.p. 131Ϫ132 °C. TLC (toluene/ethyl acetate, 10:1): Rf ϭ
0.62. [α]2D5 ϭ ϩ13.6 (c ϭ 0.53, CH2Cl2). 1H NMR (300 MHz,
CDCl3): δ ϭ 4.03 (t, J ϭ 9.0 Hz, 1 H, 3-H), 4.05 (ddd, J ϭ 3.0,
6.3, 9.9 Hz, 1 H, 5-H), 4.38 (dd, J ϭ 6.3, 12.3 Hz, 1 H, 6-H), 4.45
(s, 2 H, ArCH2), 4.64 (dd, J ϭ 3.0, 12.3 Hz, 1 H, 6-H), 4.90 (d,
J ϭ 9.9 Hz, 1 H, 1-H), 5.34 (dd, J ϭ 9.0, 9.9 Hz, 1 H, 2-H), 5.47
(dd, J ϭ 9.0, 9.9 Hz, 1 H, 4-H), 6.58 (d, J ϭ 8.1 Hz, 1 H, ClAzb),
6.87 (dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 6.97 (d, J ϭ 2.1 Hz, 1
H, ClAzb), 7.05Ϫ7.10 (m, 2 H, ArH), 7.16Ϫ7.21 (m, 1 H, ArH),
7.39Ϫ7.64 (m, 11 H, ArH), 7.92Ϫ8.02 (m, 6 H, ArH) ppm. 13C
NMR (75 MHz, CDCl3): δ ϭ 63.1 (C-6), 70.4 (C-4), 71.8 (C-2),
73.0 (ArCH2), 75.9 (C-5), 81.8 (C-3), 86.0 (C-1), 119.0, 124.0,
127.1, 127.7, 128.0, 128.12, 128.15, 128.4, 128.6, 129.0, 129.2,
129.3, 129.4, 129.6, 131.7, 132.4, 132.8, 133.1, 133.2, 134.8, 135.9,
164.3 (CϭO), 164.4 (CϭO), 165.6 (CϭO) ppm. ESI-MS: m/z ϭ
772.2 [M ϩ Naϩ]. C40H32ClN3O8S (750.22): calcd. C 64.04, H 4.30,
N 5.60; found C 64.04, H 4.26, N 5.46.
4-(Hydroxymethyl)benzyl 3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-
4,6-di-O-benzyl-β-D-glucopyranoside (11): A mixture of 2 (1.50 g,
2.08 mmol), xylylene glycol (861 mg, 6.23 mmol), Sn(OTf)2
(216 mg, 0.52 mmol), 4-A molecular sieves (ca. 500 mg) and THF
˚
(25 mL) was stirred at room temperature for 15 min, then cooled
to 0 °C. N-Bromosuccinimide (388 mg, 2.18 mmol) was added, the
mixture was stirred for 30 min and then filtered. The filtrate was
concentrated in vacuo and the residue was purified by silica-gel
column chromatography (hexane/ethyl acetate, 2:1) to give 11 as
white crystals (1.17 g, 75%). M.p. 121Ϫ123 °C. TLC (hexane/ethyl
acetate, 1:1): Rf ϭ 0.45. [α]2D5 ϭ ϩ23.4 (c ϭ 0.53, CH2Cl2). 1H
NMR (300 MHz, CDCl3): δ ϭ 3.49 (ddd, J ϭ 2.4, 4.2, 9.6 Hz, 1
H, 5-H), 3.67 (t, J ϭ 9.2 Hz, 1 H, 3-H), 3.72Ϫ3.81 (m, 3 H, 4-H,
6-H, 6-H), 4.47 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 4.49 (d, J ϭ 7.9 Hz,
1 H, 1-H), 4.55Ϫ4.68 (m, 7 H, ArCHH), 4.75 (d, J ϭ 11.1 Hz, 1
H, ArCHH), 4.84 (d, J ϭ 12.6 Hz, 1 H, ArCHH), 5.27 (dd, J ϭ
7.9, 9.2 Hz, 1 H, 2-H), 6.70 (d, J ϭ 7.8 Hz, 1 H, ClAzb), 6.98 (dd,
J ϭ 1.8, 8.1, ClAzb), 7.08 (d, J ϭ 1.8 Hz, 1 H, ClAzb), 7.10Ϫ7.21
(m, 6 H, ArH), 7.25Ϫ7.44 (m, 10 H, ArH), 7.55Ϫ7.61 (m, 1 H,
ArH), 7.84Ϫ7.88 (m, 2 H, ArH) ppm. 13C NMR (75 MHz, CDCl3):
δ ϭ 64.9 (ArCH2O), 68.4 (C-6), 69.7 (ArCH2), 73.2 (C-2), 73.4 (2
C, ArCH2 ϫ 2), 74.8 (ArCH2), 75.0 (C-5), 77.9 (C-4), 82.8 (C-3),
99.1 (C-1), 119.1, 124.0, 126.5, 127.2, 127.3, 127.4, 127.45, 127.52,
127.9, 128.0, 128.1, 129.3, 129.7, 132.7, 135.5, 135.8, 136.1, 137.4,
137.6, 139.8, 164.5 (CϭO) ppm. ESI-MS: m/z ϭ 772.3 [M ϩ Naϩ].
C42H40ClN3O8 (750.24): calcd. C 67.24, H 5.37, N 5.60; found C
67.23, H 5.28, N 5.49.
Phenyl 3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-1-thio-β-D-gluco-
pyranoside (10): Sodium methoxide (243 mg, 4.50 mmol) was added
to a solution of 9 (6.35 g, 8.46 mmol) in a mixture of methanol and
THF (1:2, 75 mL) at room temperature. After stirring for 2 h, the
mixture was neutralized with ion-exchange resin Amberlyst 15
(Hϩ form), then concentrated in vacuo. The residue was crys-
tallized from hexane/ethyl acetate to give 10 as white crystals
(4.22 g, 92%). M.p. 131Ϫ132 °C. TLC (hexane/ethyl acetate, 1:2):
Rf ϭ 0.49. [α]2D5 ϭ ϩ83.5 (c ϭ 0.51, CH2Cl2). 1H NMR (300 MHz,
CDCl3): δ ϭ 1.99 (t, J ϭ 6.8 Hz, 1 H, 6-OH), 2.50 (d, J ϭ 3.3 Hz,
1 H, 4-OH), 3.50 (ddd, J ϭ 3.6, 5.1, 9.0 Hz, 1 H, 5-H), 3.68 (t, J ϭ
9.0 Hz, 1 H, 3-H), 3.78 (dt, J ϭ 3.3, 9.0 Hz, 1 H, 4-H), 3.85 (ddd,
J ϭ 5.1, 6.8, 12.0 Hz, 1 H, 6-H), 3.96 (ddd, J ϭ 3.6, 6.8, 12.0 Hz,
1 H, 6-H), 4.58 (d, J ϭ 11.7 Hz, 1 H, ArCHH), 4.66 (d, J ϭ
11.7 Hz, 1 H, ArCHH), 4.82 (d, J ϭ 10.1 Hz, 1 H, 1-H), 5.20 (dd,
J ϭ 9.0, 10.1 Hz, 1 H, 2-H), 6.85 (d, J ϭ 8.1 Hz, 1 H, ClAzb), 7.05
(dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 7.16 (d, J ϭ 2.1 Hz, 1 H, ClAzb),
7.24Ϫ7.29 (m, 3 H, ArH), 7.38Ϫ7.48 (m, 4 H, ArH), 7.57Ϫ7.63
(m, 1 H, ArH) 7.96Ϫ8.00 (m, 2 H, ArH) ppm. 13C NMR (75 MHz,
CDCl3): δ ϭ 62.2 (C-6), 70.3 (C-4), 71.7 (C-2), 73.4 (ArCH2), 79.1
(C-5), 83.9 (C-3), 86.2 (C-1), 119.2, 124.3, 127.0, 127.7, 128.1,
128.6, 129.1, 129.4, 129.7, 132.0, 132.1, 133.0, 135.3, 136.0, 164.6
(CϭO) ppm. ESI-MS: m/z ϭ 564.0 [M ϩ Naϩ]. C26H24ClN3O6S
(542.00): calcd. C 57.62, H 4.46, N 7.75; found C 57.32, H 4.38,
N 7.64.
4-{[(3-Carboxypropanoyl)oxy]methyl}benzyl 3-O-(4-Azido-3-chloro-
benzyl)-2-O-benzoyl-4,6-di-O-benzyl-β-D-glucopyranoside (12): Suc-
cinic anhydride (195 mg, 1.95 mmol) was added to a solution of 11
(977 mg, 1.30 mmol), dimethylaminopyridine (79 mg, 0.65 mmol)
and triethylamine (0.18 mL, 1.30 mmol) in CH2Cl2 (5 mL) at room
temperature and the mixture was stirred for 2 h, then washed with
hydrochloric acid (0.1 mol/L) and brine, dried with Na2SO4 and
concentrated in vacuo. Crystallization from hexane/ethyl acetate
Phenyl 3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-4,6-di-O-benzyl-
1-thio-β-
D
-glucopyranoside (2): NaH (60% suspension in oil, gave 12 as pale-yellow crystals (1.02 g, 92%). M.p. 125Ϫ128 °C.
812 mg, 20.3 mmol) was added in portions to a solution of 10
TLC (ethyl acetate): Rf ϭ 0.64. [α]2D5 ϭ ϩ26.6 (c ϭ 0.52, CH2Cl2).
3440
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
Eur. J. Org. Chem. 2003, 3435Ϫ3445

Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide
FULL PAPER
1H NMR (300 MHz, CDCl3): δ ϭ 2.65Ϫ2.68 (m, 4 H, COCH2), J ϭ 11.8 Hz, 1 H, ArCHH), 4.56 (d, J ϭ 10.4 Hz, 1 H, ArCHH),
3.50 (ddd, J ϭ 2.3, 3.8, 9.6 Hz, 1 H, 5-H), 3.68 (t, J ϭ 9.2 Hz, 1 4.65 (d, J ϭ 11.8 Hz, 1 H, ArCHH), 4.80 (d, J ϭ 10.1 Hz, 1 H, 1-
H, 3-H), 3.72Ϫ3.81 (m, 3 H, 4-H, 6-H, 6-H), 4.47 (d, J ϭ 7.9 Hz, H), 5.27 (dd, J ϭ 9.0, 10.1 Hz, 1 H, 2-H), 6.70 (d, J ϭ 8.1 Hz, 1
1 H, 1-H), 4.47 (d, J ϭ 12.1 Hz, 1 H, ArCHH), 4.57 (d, J ϭ H, ClAzb), 6.85 (dd, J ϭ 2.1, 8.1 Hz, 2 H, ArH), 6.96 (dd, J ϭ
12.2 Hz, 1 H, ArCHH), 4.60 (d, J ϭ 10.8 Hz, 1 H, ArCHH), 4.61
2.0, 8.1 Hz, 1 H, ClAzb), 7.06 (d, J ϭ 2.0 Hz, 1 H, ClAzb),
(d, J ϭ 12.6 Hz, 1 H, ArCHH), 4.65 (d, J ϭ 12.1 Hz, 1 H, 7.16Ϫ7.30 (m, 14 H, ArH), 7.42Ϫ7.61 (m, 12 H, ArH), 7.94Ϫ7.97
ArCHH), 4.66 (d, J ϭ 12.2 Hz, 1 H, ArCHH), 4.75 (d, J ϭ (m, 2 H, ArH) ppm. 13C NMR (75 MHz, CDCl3): δ ϭ 62.0 (C-6),
10.8 Hz, 1 H, ArCHH), 4.84 (d, J ϭ 12.6 Hz, 1 H, ArCHH), 5.05
(s, 2 H, ArCH2), 5.27 (dd, J ϭ 7.9, 9.2 Hz, 1 H, 2-H), 6.71 (d, J ϭ
8.1 Hz, 1 H, ClAzb), 6.98 (dd, J ϭ 1.8, 8.1, ClAzb), 7.08 (d, J ϭ
71.8 (C-2), 73.8 (ArCH2), 74.9 (ArCH2), 77.7 (C-4), 78.9 (C-5),
84.3 (C-3), 85.8 (Ph3C), 86.3 (C-1), 119.1, 124.1, 126.6, 127.2,
127.5, 127.7, 127.9, 128.0, 128.4, 128.5, 129.30, 129.34, 129.8,
1.8 Hz, 1 H, ClAzb), 7.09Ϫ7.45 (m, 16 H, ArH), 7.56Ϫ7.61 (m, 1 132.3, 132.4, 132.8, 135.4, 135.9, 137.0, 143.3, 164.5 (CϭO) ppm.
H, ArH), 7.85Ϫ7.88 (m, 2 H, ArH) ppm. 13C NMR (75 MHz, ESI-MS: m/z ϭ 896.1[M ϩ Naϩ]. C52H44ClN3O6S (874.45): calcd.
CDCl3): δ ϭ 28.6 (COCH2), 28.9 (COCH2), 66.2 (ArCH2), 68.5 C 71.42, H 5.07, N 4.81; found C 71.10, H 5.01, N 4.73.
(C-6), 69.7 (ArCH2), 73.3 (C-2), 73.5 (2 C, ArCH2 ϫ 2), 74.9
Phenyl
thio-β-
3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-4-O-benzyl-1-
-glucopyranoside (15): Trifluoroacetic acid (90%, 10 mL)
(ArCH2), 75.0 (C-5), 78.0 (C-4), 82.8 (C-3), 99.2 (C-1), 119.2, 124.2,
127.3, 127.5, 127.6, 127.61, 127.64, 127.7, 128.0, 128.1, 128.20,
128.25, 129.5, 129.8, 133.0, 134.8, 135.7, 136.0, 137.0, 137.5, 137.7,
164.7 (CϭO), 171.6 (CϭO) ppm. ESI-MS: m/z ϭ 872.2 [M ϩ
Naϩ], 848.2 [M Ϫ H]Ϫ. C46H44ClN3O11 (850.32): calcd. C 64.98,
H 5.22, N 4.94; found C 64.95, H 5.19, N 4.78.
D
was added to a solution of 14 (2.62 g, 3.00 mmol) in CH2Cl2
(10 mL) and the mixture was stirred at room temperature for 2 h.
After concentration in vacuo, the residue was dissolved in ethyl
acetate and washed with satd. aqueous NaHCO3 and brine, then
concentrated again. The residue was dissolved in THF (20 mL).
Aqueous NaHCO3 (84 mg, 2.0 mmol/2 mL) was added and the
mixture was stirred at room temperature for 30 min, then concen-
trated in vacuo. The residue was dissolved in ethyl acetate, washed
with brine, dried with Na2SO4 and then concentrated in vacuo. The
Phenyl 3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-6-O-(triphenyl-
methyl)-1-thio-β-D-glucopyranoside (13): 2,4,6-Collidine (0.85 mL,
6.46 mmol) was added to a solution of 10 (2.50 g, 4.61 mmol), tri-
phenylmethyl chloride (1.80 g, 6.46 mmol) and tetrabutylam-
monium perchlorate (394 mg, 1.15 mmol) in CH2Cl2 (20 mL) and residue was purified by silica-gel column chromatography (hexane/
the mixture was stirred at room temperature for 2 h. The mixture ethyl acetate, 3:1) to give 15 as white crystals (1.63 g, 86%). M.p.
was washed with an aqueous solution of citric acid (10%) and
brine, dried with Na2SO4, then concentrated in vacuo. The residue
109Ϫ111 °C. TLC (toluene/ethyl acetate, 10:1): Rf ϭ 0.28. [α]2D5
ϭ
ϩ79.8 (c ϭ 0.51, CH2Cl2). 1H NMR (300 MHz, CDCl3): δ ϭ 1.88
was purified by silica-gel column chromatography (hexane/ethyl (m, 1 H, 6-OH), 3.49 (ddd, J ϭ 2.4, 4.2, 9.0 Hz, 1 H, 5-H), 3.70 (t,
acetate, 4:1) to give 13 as a pale-yellow gum (3.55 g, 98%). TLC
J ϭ 9.0 Hz, 1 H, 4-H), 3.70Ϫ3.78 (m, 1 H, 6-H), 3.78 (t, J ϭ
9.0 Hz, 1 H, 3-H), 3.90Ϫ3.97 (m, 1 H, 6-H), 4.51 (d, J ϭ 11.7 Hz,
(hexane/ethyl acetate, 2:1): Rf ϭ 0.63. [α]2D5 ϭ ϩ37.9 (c ϭ 0.57,
1
CH2Cl2). H NMR (300 MHz, CDCl3): δ ϭ 2.43 (d, J ϭ 3.0 Hz,1 1 H, ArCHH), 4.69 (d, J ϭ 11.7 Hz, 2 H, ArCHH ϫ 2), 4.80 (d,
H, 4-OH), 3.40Ϫ3.54 (m, 3 H, 5-H, 6-H, 6-H), 3.59 (t, J ϭ 9.0 Hz, J ϭ 10.2 Hz, 1 H, 1-H), 4.81 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 5.19
1 H, 3-H), 3.80 (dt, J ϭ 3.0, 9.0 Hz, 4-H), 4.53 (d, J ϭ 12.0 Hz, 1 (dd, J ϭ 9.0, 10.2 Hz, 1 H, 2-H), 6.72 (d, J ϭ 8.1 Hz, 1 H, ClAzb),
H, ArCHH), 4.67 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 4.75 (d, J ϭ
6.98 (dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 7.08 (d, J ϭ 2.1 Hz, 1 H,
9.9 Hz, 1 H, 1-H), 5.18 (dd, J ϭ 9.0, 9.9 Hz, 1 H, 2-H), 6.79 (d, ClAzb), 7.22Ϫ7.47 (m, 12 H, ArH), 7.57Ϫ7.66 (m, 1 H, ArH),
J ϭ 8.1 Hz, 1 H, ClAzb), 7.00 (dd, J ϭ 2.1, 8.1 Hz, 1 H, ClAzb), 7.92Ϫ7.95 (m, 2 H, ArH) ppm. 13C NMR (75 MHz, CDCl3): δ ϭ
7.14 (d, J ϭ 2.1 Hz, 1 H, ClAzb), 7.18Ϫ7.33 (m, 12 H, ArH), 61.7 (C-6), 71.9 (C-2), 73.7 (ArCH2), 75.0 (ArCH2), 77.4 (C-4),
7.41Ϫ7.50 (m, 10 H, ArH), 7.57Ϫ7.62 (m, 1 H, ArH), 7.95Ϫ7.98
79.4 (C-5), 84.0 (C-3), 85.8 (C-1), 119.1, 124.1, 127.2, 127.6, 127.7,
(m, 2 H, ArH) ppm. 13C NMR (75 MHz, CDCl3): δ ϭ 63.4 (C-6), 128.0, 128.2, 128.6, 129.1, 129.3, 129.7, 132.0, 132.2, 132.9, 135.3,
71.5 (C-2), 71.7 (C-4), 73.2 (ArCH2), 78.1 (C-5), 83.6 (C-3), 85.8 135.9, 137.2, 164.5 (CϭO) ppm. ESI-MS: m/z ϭ 654.0 [M ϩ Naϩ].
(Ph3C), 86.8 (C-1), 119.1, 124.1, 126.8, 127.1, 127.5, 127.6, 128.0,
C33H30ClN3O6S (632.13): calcd. C 62.70, H 4.78, N 6.65; found C
128.2, 128.4, 129.2, 129.3, 129.7, 132.1, 132.3, 132.9, 135.5, 135.8, 62.53, H 4.69, N 6.66.
143.1, 164.5 (CϭO) ppm. ESI-MS: m/z ϭ 806.1 [M ϩ Naϩ].
C45H38ClN3O6S (784.32): calcd. C 68.91, H 4.88, N 5.36; found C
68.80, H 5.01, N 5.16.
2,3,4,6-Tetra-O-benzoyl-α-D-glucopyranosyl Benzoate (17): Benzoyl
chloride (6.91 mL, 60.0 mmol) was added dropwise to a suspension
of -glucose (1.80 g, 10.0 mmol) in pyridine (50 mL) at 0 °C and
the mixture was stirred for 2 h. The reaction was quenched by ad-
ding cold water (100 mL) and the product was extracted twice with
ethyl acetate. The combined organic layers were washed with hydro-
chloric acid (1 mol/L) and brine, dried with Na2SO4 and then con-
Phenyl 3-O-(4-Azido-3-chlorobenzyl)-2-O-benzoyl-4-O-benzyl-6-O-
(triphenylmethyl)-1-thio-β-D-glucopyranoside (14): NaH (60% sus-
pension in oil, 190 mg, 4.74 mmol) was added in portions to a solu-
tion of 13 (2.86 g, 3.65 mmol) and benzyl bromide (0.56 mL,
4.74 mmol) in DMF (20 mL) at 0 °C and the mixture was stirred centrated in vacuo. The residue was crystallized from hexane/ethyl
for 2 h. The reaction was quenched by adding cold water (50 mL)
and the product was extracted twice with ethyl acetate. The com-
bined organic layers were washed with brine, dried with Na2SO4
and concentrated in vacuo. The residue was purified by silica-gel
column chromatography (hexane/ethyl acetate, 5:1) to give 14 as
white crystals (3.00 g, 94%). M.p. 162Ϫ164 °C. TLC (hexane/ethyl
acetate, 3:1): Rf ϭ 0.64. [α]2D5 ϭ ϩ46.5 (c ϭ 0.50, CH2Cl2). 1H
acetate to give 17 as white crystals (6.66 g, 95%). M.p. 177Ϫ179
°C. TLC (hexane/ethyl acetate, 2:1): Rf ϭ 0.49. [α]2D5 ϭ ϩ136.8 (c ϭ
1.00, CH2Cl2). 1H NMR (300 MHz, CDCl3): δ ϭ 4.48 (dd, J ϭ
5.1, 12.6 Hz, 1 H, 6-H), 4.58Ϫ4.64 (m, 2 H, 5-H, 6-H), 5.67 (dd,
J ϭ 3.9, 9.9 Hz, 1 H, 2-H), 5.85 (t, J ϭ 9.9 Hz, 1 H, 4-H), 6.31 (t,
J ϭ 9.9 Hz, 1 H, 3-H), 6.84 (d, J ϭ 3.9 Hz, 1 H, 1-H), 7.24Ϫ7.56
(m, 14 H, ArH), 7.62Ϫ7.68 (m, 1 H, ArH), 7.86 (m, 4 H, ArH),
NMR (300 MHz, CDCl3): δ ϭ 3.29 (dd, J ϭ 4.1, 10.1 Hz, 1 H, 6- 7.93 (m, 2 H, ArH), 8.01 (m, 2 H, ArH), 8.15 (m, 2 H, ArH) ppm.
H), 3.54 (ddd, J ϭ 1.5, 4.1, 9.0 Hz, 1 H, 5-H), 3.65 (dd, J ϭ 1.5,
13C NMR (75 MHz, CDCl3): δ ϭ 62.3 (C-6), 68.6 (C-4), 70.3 (3 s,
10.1 Hz, 1 H, 6-H), 3.69 (t, J ϭ 9.0 Hz, 1 H, 3-H), 3.86 (t, J ϭ 3 C, C-2, C-3, C-5), 89.8 (C-1), 127.99, 128.02, 128.2, 128.3, 128.4,
9.0 Hz, 1 H, 4-H), 4.31 (d, J ϭ 10.4 Hz, 1 H, ArCHH), 4.45 (d, 128.6, 129.1, 129.35, 129.39, 129.45, 129.49, 129.6, 132.7, 132.9,
Eur. J. Org. Chem. 2003, 3435Ϫ3445
www.eurjoc.org
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3441

K. Egusa, S. Kusumoto, K. Fukase
FULL PAPER
133.1, 133.5, 163.9 (CϭO), 164.6 (CϭO), 164.8 (CϭO), 165.4 (Cϭ
0.18 mmol) was added and the mixture was stirred for 2 h, then
O), 165.6 (CϭO) ppm. ESI-MS: m/z ϭ 723.2 [M ϩ Na]ϩ. filtered. The filtrate was concentrated in vacuo and the residue was
C41H32O11 (700.69): calcd. C 70.28, H 4.60; found C 70.11, H 4.54.
purified by silica-gel column chromatography (toluene/ethyl acetate
10:1) to give 3 as white crystals (1.51 g, 83%). M.p. 164Ϫ167 °C.
TLC (toluene/ethyl acetate, 10:1): Rf ϭ 0.47. [α]2D5 ϭ ϩ38.6 (c ϭ
0.51, CH2Cl2). 1H NMR (300 MHz, CDCl3): δ ϭ 3.45 (dd, J ϭ
9.0, 9.3 Hz, 1 H, 4a-H), 3.59 (ddd, J ϭ 1.8, 5.6, 9.3 Hz, 1 H, 5a-
H), 3.65 (t, J ϭ 9.0 Hz, 1 H, 3a-H), 3.83 (dd, J ϭ 5.6, 11.8 Hz, 1
H, 6a-H), 4.06 (ddd, J ϭ 3.1, 5.0, 9.6 Hz, 1 H, 5b-H), 4.15 (dd,
J ϭ 1.8, 11.8 Hz, 1 H, 6a-H), 4.38 (d, J ϭ 11.7 Hz, 1 H, ArCHH),
4.46 (d, J ϭ 10.9 Hz, 1 H, ArCHH), 4.49 (dd, J ϭ 5.0, 12.0 Hz, 1
H, 6b-H), 4.53 (d, J ϭ 10.9 Hz, 1 H, ArCHH), 4.55 (d, J ϭ
11.7 Hz, 1 H, ArCHH), 4.63 (dd, J ϭ 3.1, 12.0 Hz, 1 H, 6b-H),
4.66 (d, J ϭ 10.0 Hz, 1 H, 1a-H), 4.92 (d, J ϭ 7.8 Hz, 1 H, 1b-H),
5.11 (dd, J ϭ 9.0, 10.0 Hz, 1 H, 2a-H), 5.56 (dd, J ϭ 7.8, 9.6 Hz,
1 H, 2b-H), 5.66 (t, J ϭ 9.6 Hz, 1 H, 4b-H), 5.85 (d, J ϭ 9.6 Hz,
1 H, 3b-H), 6.70 (d, J ϭ 8.1 Hz, 1 H, ClAzb), 6.91 (dd, J ϭ 2.1,
8.1 Hz, 1 H, ClAzb), 7.07 (d, J ϭ 2.1 Hz, 1 H, ClAzb), 7.09Ϫ7.13
(m, 2 H, ArH), 7.23Ϫ7.61 (m, 23 H, ArH), 7.80Ϫ7.94 (m, 8 H,
ArH), 7.99Ϫ8.03 (m, 2 H, ArH) ppm. 13C NMR (75 MHz, CDCl3):
δ ϭ 62.9 (C-6b), 67.7 (C-6a), 69.5 (C-4b), 71.6 (C-2b), 71.7 (C-2a),
72.1 (C-5b), 72.4 (C-3b), 73.5 (ArCH2), 74.8 (ArCH2), 77.6 (C-
4a), 78.9 (C-5a), 84.1 (C-3a), 85.7 (C-1a), 100.7 (C-1b), 119.0,
124.0, 127.1, 127.3, 127.6, 127.8, 127.9, 127.98, 128.03, 128.1,
128.4, 128.6, 128.8, 129.17, 129.20, 129.3, 129.36, 129.39, 129.7,
132.0, 132.4, 132.7, 132.8, 133.0, 135.3, 135.8, 137.1, 164.4 (CϭO),
164.5 (CϭO), 164.7 (CϭO), 165.3 (CϭO), 165.6 (CϭO) ppm. ESI-
MS: m/z ϭ 1232.3 [M ϩ Naϩ]. C67H56ClN3O15S (1210.70): calcd.
C 66.47, H 4.66, N 3.47; found C 66.20, H 4.58, N 3.51.
2,3,4,6-Tetra-O-benzoyl-α/β- -glucopyranose (18): A solution of
D
methylamine in methanol (40%, 2 mL) was added to a solution of
17 (2.57 g, 3.66 mmol) in THF (20 mL) at room temperature and
the mixture was stirred for 2 h. The mixture was concentrated in
vacuo and the residue was purified by silica-gel column chromatog-
raphy (hexane/ethyl acetate, 3:1) to give 18 as a colorless solid
(1.81 g, 83%, α/β ϭ 4:1). TLC (hexane/ethyl acetate, 2:1): Rf ϭ
1
0.36. H NMR (300 MHz, CDCl3): α-anomer: δ ϭ 4.44 (dd, J ϭ
4.5, 12.0 Hz, 1 H, 6-H), 4.65 (dd, J ϭ 3.0, 12.0 Hz, 1 H, 6-H), 4.67
(ddd, J ϭ 3.0, 4.5, 9.9 Hz, 1 H, 5-H), 5.32 (dd, J ϭ 3.3, 9.9 Hz, 1
H, 2-H), 5.73 (t, J ϭ 9.9 Hz, 1 H, 4-H), 5.75 (d, J ϭ 3.3 Hz, 1 H,
1-H), 6.25 (t, J ϭ 9.9 Hz, 1 H, 3-H), 7.24Ϫ7.61 (m, 12 H, ArH),
7.83Ϫ8.08 (m, 8 H, ArH); β-anomer: δ ϭ 4.19 (ddd, J ϭ 3.0, 5.1,
9.7 Hz, 1 H, 5-H), 4.49 (dd, J ϭ 5.1, 12.3 Hz, 1 H, 6-H), 4.67 (dd,
J ϭ 3.0, 12.3 Hz, 1 H, 6-H), 5.06 (d, J ϭ 8.1 Hz, 1 H, 1-H), 5.35
(dd, J ϭ 8.1, 9.7 Hz, 1 H, 2-H), 5.70 (t, J ϭ 9.7 Hz, 1 H, 4-H),
5.96 (t, J ϭ 9.7 Hz, 1 H, 3-H), 7.24Ϫ7.61 (m, 12 H, ArH),
7.83Ϫ8.08 (m, 8 H, ArH) ppm. 13C NMR (75 MHz, CDCl3): α-
anomer: δ ϭ 62.7 (C-6), 67.6 (C-5), 69.2 (C-4), 69.9 (C-3), 72.0 (C-
2), 90.2 (C-1), 127.9, 128.00, 128.03, 128.07, 128.50, 128.54, 128.7,
129.2, 129.3, 129.4, 129.5, 132.7, 133.0, 133.1, 164.8 (CϭO), 165.4
(2 C, CϭO ϫ 2), 165.8 (CϭO) ppm. ESI-MS: m/z ϭ 619.1 [M ϩ
Naϩ]. C34H28O10 (596.59): calcd. C 68.45, H 4.73; found C 68.06,
H 4.73.
O-(2,3,4,6-Tetra-O-benzoyl-α/β-D-glucopyranosyl) Trichloroacetim-
idate (16): Cs2CO3 (164 mg, 0.50 mmol) was added to a solution of
18 (1.50 g, 2.51 mmol) and trichloroacetonitrile (2.52 mL,
25.1 mmol) in CH2Cl2 (5 mL) at room temperature and the mixture
was stirred for 2 h, then filtered. The filtrate was concentrated in
vacuo and the residue was purified by silica-gel column chromatog-
raphy (hexane/ethyl acetate, 3:1) to give 16 as a white solid (1.70 g,
91%, α/β ϭ 5:1). TLC (hexane/ethyl acetate, 2:1): Rf ϭ 0.56, 0.44
(α- and β-anomer, respectively). 1H NMR (300 MHz, CDCl3): α-
anomer: δ ϭ 4.48 (dd, J ϭ 5.7, 12.9 Hz, 1 H, 6-H), 4.61Ϫ4.66 (m,
2 H, 5-H, 6-H), 5.61 (dd, J ϭ 3.8, 10.0 Hz, 1 H, 2-H), 5.81 (t, J ϭ
10.0 Hz, 1 H, 4-H), 6.27 (t, J ϭ 10.0 Hz, 1 H, 3-H), 6.83 (d, J ϭ
3.8 Hz, 1 H, 1-H), 7.24Ϫ7.58 (m, 12 H, ArH), 7.84Ϫ8.03 (m, 8 H,
ArH), 8.62 (s, 1 H, NH); β-anomer: δ ϭ 4.38 (ddd, J ϭ 3.5, 4.8,
9.5 Hz, 1 H, 5-H), 4.54 (dd, J ϭ 4.8, 11.9 Hz, 1 H, 6-H), 4.68 (dd,
J ϭ 3.5, 11.9 Hz, 1 H, 6-H), 5.82 (dd, J ϭ 7.5, 9.0 Hz, 1 H, 2-H),
5.82 (dd, J ϭ 9.0, 9.5 Hz, 1 H, 4-H), 5.97 (t, J ϭ 9.0 Hz, 1 H, 3-
H), 6.23 (d, J ϭ 7.5 Hz, 1 H, 1-H), 7.24Ϫ7.56 (m, 12 H, ArH),
7.85Ϫ8.04 (m, 8 H, ArH), 8.70 (s, 1 H, NH) ppm. 13C NMR
(75 MHz, CDCl3): α-anomer: δ ϭ 62.3 (C-6), 68.5 (C-4), 70.0 (C-
Aminomethylated Resin 19: A suspension of Diaion HP20 (5.23 g),
N-(hydroxymethyl)phthalimide (185 mg, 1.05 mmol), trifluorome-
thanesulfonic acid (0.47 mL, 5.23 mmol) in trifluoroacetic acid/
CH2Cl2 (1:1, 50 mL) was shaken at room temperature for 5 h. The
reaction was monitored by IR spectroscopy of washed resin
samples (KBr pellets of 2Ϫ3 beads) and proceeded until no further
change was observed around 1720 cmϪ1. The mixture was filtered
and the resin washed with methanol and CH2Cl2, then dried in
vacuo. The resin was suspended in ethanol (45 mL), hydrazine hy-
drate (5 mL) was added and the mixture was refluxed for 16 h. The
reaction was monitored by IR spectroscopy of washed resin
samples (KBr pellets of 2Ϫ3 beads) and proceeded until the peak
around 1720 cmϪ1 disappeared. The mixture was filtered while hot,
and the resulting resin was washed with methanol and dried in
vacuo to give 19 (5.59 g, 0.19 mmol/g loading) as faintly yellow be-
ads.
Determination of the Loading Rate of Aminomethyl Group: The re-
sin 19 (ca. 100 mg, exactly weighed) was washed with a solution of
3), 70.5 (2 s, 2 C, C-2, C-5), 92.8 (C-1), 127.96, 127.99, 128.05, diisopropylethylamine in CH2Cl2 (5%, 5 mL ϫ 2) and CH2Cl2
128.17, 128.23, 128.5, 129.2, 129.3, 129.4, 129.5, 132.7, 132.9, (5 mL ϫ 2). Then, the resin was suspended in a solution of picric
133.2, 160.0 (CϭNH), 164.7 (CϭO), 164.9 (CϭO), 165.1 (CϭO), acid in CH2Cl2 (0.1 mol/L, 5 mL) and the suspension was shaken
165.5 (CϭO); β-anomer: δ ϭ 62.6 (C-6), 68.9 (C-4), 70.5 (C-2),
72.4 (C-3), 72.8 (C-5), 95.6 (C-1), 127.96, 128.03, 128.05, 128.3,
128.6, 129.2, 129.37, 129.42, 129.44, 132.7, 133.0, 133.1, 160.5 (Cϭ
NH), 164.3 (CϭO), 164.6 (CϭO), 165.1 (CϭO), 165.6 (CϭO) ppm.
at room temperature for 1 h. The resin was filtered and washed
with CH2Cl2 (5 mL ϫ 5) and methanol (5 mL ϫ 2). The picrate
on the resin was eluted with a solution of diisopropylethylamine in
CH2Cl2 (5%, 5 mL ϫ 2) and CH2Cl2 (4 mL ϫ 2). The filtrate con-
ESI-MS: m/z ϭ 762.1 [M ϩ Naϩ]. C36H28Cl3NO10 (740.97): calcd. taining the picrate was combined and diluted with CH2Cl2 to
C 58.35, H 3.81, N 1.89; found C 58.47, H 3.80, N 1.98.
20 mL. This solution was further diluted with ethanol (1:50) and
subjected to UV measurements. The concentration was calculated
using a standard straight line prepared with solutions of known
amounts of picric acid.
Phenyl (2,3,4,6-Tetra-O-benzoyl-β- -glucopyranosyl)-(1Ǟ6)-3-O-(4-
D
azido-3-chlorobenzyl)-2-O-benzoyl-4-O-benzyl-1-thio-β- -gluco-
D
pyranoside (3): A mixture of imidate 16 (1.33 g, 1.80 mmol), thio-
˚
glycoside 15 (948 mg, 1.50 mmol) and 4-A molecular sieves (ca. Solid-Supported Monosaccharide 4: A mixture of carboxylic acid
500 mg) in CH2Cl2 (10 mL) was stirred at room temperature for
15 min, then cooled to ice/salt temperature. Sn(OTf)2 (75 mg,
12 (187 mg, 0.22 mmol), resin 19 (1.053 g, 0.2 mmol), N,N-diiso-
propylcarbodiimide (38 µL, 0.24 mmol), 1-hydroxybenzotriazole
3442
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
Eur. J. Org. Chem. 2003, 3435Ϫ3445

Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide
FULL PAPER
(HOBt) (30 mg, 0.22 mmol) and CH2Cl2 (5.5 mL) was shaken at
room temperature for 16 h. After filtration, the resin was washed
with methanol and CH2Cl2, then dried in vacuo to give solid-sup-
ported monosaccharide 4 as a slightly yellowish powder (1.232 g,
quant., calculated from weight gain of the resin).
molecular sieves (bead type, ca. 0.5 g) were added and the mixture
was shaken at room temperature for 15 min. The reaction was
started by addition of NBS (112 mg, 0.63 mmol) and the mixture
was shaken at room temperature for 1 h, then filtered. The resin
was suspended in CH2Cl2 (5 mL) and allowed to settle to remove
molecular sieves. The resin was further washed with methanol
(10 mL ϫ 2) and CH2Cl2 (10 mL ϫ 2), and dried in vacuo to give
solid-supported trisaccharide 22 as a slightly yellowish powder
(1.325 g). IR measurement (KBr disk) was carried out on the resin
to confirm the introduction of ClAzb-donor onto the resin (peak
of the azido group at 2126 cmϪ1). If necessary, the procedure was
repeated to complete the glycosylation.
4-(Hydroxymethyl)benzyl 3-O-(4-Azido-3-chlorobenzyl)-4,6-di-O-
benzyl-β-D-glucopyranoside (20). General Procedure for Cleavage:
Solid-supported monosaccharide 4 (245 mg, 0.04 mmol) was sus-
pended in a mixture of methanol and THF (1:1, 5 mL). Sodium
methoxide (27 mg, 0.5 mmol) was added and the mixture was
shaken at room temperature for 4 h. The resin was filtered off and
washed with methanol (5 mL ϫ 2). Complete cleavage was con-
firmed by IR measurement (KBr disk) on a small portion of the
resin (disappearance of the peak of the azido group around 2120
cmϪ1). If necessary, the procedure was repeated to complete the
cleavage. The filtrate and washings were combined and neutralized
with Amberlyst 15 (Hϩ form), then concentrated in vacuo. The
residue was purified by preparative TLC (hexane/ethyl acetate, 1:1)
to give 20 as white crystals (25 mg, quant.). M.p. 145Ϫ146 °C. TLC
(hexane/ethyl acetate, 1:1): Rf ϭ 0.42. [α]2D5 ϭ Ϫ36.2 (c ϭ 0.52,
4-(Hydroxymethyl)benzyl β-
3-chlorobenzyl)-4,6-di-O-benzyl-β-
benzyl-β- -glucopyranoside (23): Solid-supported trisaccharide 22
D-Glucopyranosyl-(1Ǟ6)-3-O-(4-azido-
D-glucopyransyl-(1Ǟ3)-4,6-di-O-
D
(331 mg) was treated according to the General Procedure for cleav-
age and the product was purified by silica-gel column chromatogra-
phy (CH2Cl2/methanol, 98:2) to give trisaccharide 23 as a colorless
solid (20 mg, 37% from 4). TLC (CH2Cl2/methanol, 9:1): Rf ϭ 0.35.
1H NMR (300 MHz, CDCl3 ϩ 5% CD3OD): δ ϭ 2.94 (m, 1 H,
5c-H), 3.11 (dd, J ϭ 7.5, 8.7 Hz, 1 H, 2c-H), 3.28 (t, J ϭ 8.7 Hz,
1 H, 3c-H), 3.33 (t, J ϭ 8.7 Hz, 1 H, 4c-H), 3.40Ϫ3.67 (m, 10 H),
3.68 (dd, J ϭ 4.2, 10.9 Hz, 1 H, 6a-H), 3.75 (dd, J ϭ 2.1, 10.9 Hz,
1 H, 6a-H), 3.89 (overlapped, 1 H, 6b-H), 3.91 (t, J ϭ 9.5 Hz, 1 H,
3a-H), 4.14 (d, J ϭ 7.5 Hz, 1 H, 1c-H), 4.33 (d, J ϭ 7.8 Hz, 1 H,
1b-H), 4.50 (d, J ϭ 10.2 Hz, 1 H, ArCHH), 4.54 (d, J ϭ 12.3 Hz,
1 H, ArCHH), 4.58 (d, J ϭ 11.1 Hz, 2 H, ArCHH), 4.61 (s, 2 H,
ArCH2O), 4.61 (d, J ϭ 12.3 Hz, 1 H, ArCHH), 4.65 (d, J ϭ 9.3 Hz,
1 H, 1a-H), 4.68 (d, J ϭ 11.7 Hz, 1 H, ArCHH), 4.77 (d, J ϭ
11.1 Hz, 1 H, ArCHH), 4.86 (d, J ϭ 11.7 Hz, 1 H, ArCHH), 4.89
(d, J ϭ 10.2 Hz, 1 H, ArCHH), 4.90 (d, J ϭ 11.1 Hz, 1 H,
ArCHH), 7.04 (d, J ϭ 8.1 Hz, 1 H, ClAzb), 7.19Ϫ7.37 (m, 21 H,
ArH) ppm. 13C NMR (75 MHz, CDCl3 ϩ 5% CD3OD): δ ϭ 61.4
(6c-C), 64.5 (ArCH2OH), 68.0 (C-6b), 68.5 (C-6a), 69.4 (4c-C), 70.4
(ArCH2), 72.9, 73.1 (2c-C), 73.4 (ArCH2), 73.6 (ArCH2), 74.6 (4
C), 74.8 (ArCH2), 75.1 (5c-C), 75.9 (3c-C), 76.0, 77.5, 82.6 (C-3a),
84.3 (C-3b), 100.7 (C-1b), 102.0 (C-1a), 102.7 (1c-C), 119.2, 124.4,
126.9, 127.0, 127.46, 127.50, 127.57, 127.60, 128.0, 128.1, 128.2,
128.3, 129.7, 135.8, 136.0, 136.2, 137.3, 137.4, 137.5, 140.5 ppm.
ESI-MS: m/z ϭ 1082.4 [M ϩ Naϩ]. C54H62ClN3O17 (1060.5).
1
CH2Cl2). H NMR (300 MHz, CDCl3): δ ϭ 1.61 (t, J ϭ 5.7 Hz, 1
H, CH2-OH), 2.31 (d, J ϭ 2.1 Hz, 1 H, 2-OH), 3.46 (ddd, J ϭ 2.4,
4.2, 8.4 Hz, 1 H, 5-H), 3.55 (t, J ϭ 8.4 Hz, 1 H, 3-H), 3.60 (ddd,
J ϭ 2.1, 7.7, 8.4 Hz, 1 H, 2-H), 3.61 (t, J ϭ 8.4 Hz, 1 H, 4-H), 3.71
(dd, J ϭ 4.2, 10.8 Hz, 1 H, 6-H), 3.76 (dd, J ϭ 2.4, 10.8 Hz, 1 H,
6Ј-H), 4.32 (d, J ϭ 7.7 Hz, 1 H, 1-H), 4.54 (d, J ϭ 12.3 Hz, 1 H,
ArCHH), 4.54 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 4.61 (d, J ϭ
11.4 Hz, 1 H, ArCHH), 4.63 (d, J ϭ 12.3 Hz, 1 H, ArCHH), 4.69
(d, J ϭ 5.7 Hz, 2 H, ArCH2OH), 4.71 (d, J ϭ 11.7 Hz, 1 H,
ArCHH), 4.75 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 4.85 (d, J ϭ
11.7 Hz, 1 H, ArCHH), 4.94 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 7.05
(d, J ϭ 8.4 Hz, 1 H, ClAzb), 7.13Ϫ7.16 (m, 2 H, ArH), 7.20Ϫ7.35
(m, 14 H, ArH) ppm. 13C NMR (75 MHz, CDCl3): δ ϭ 64.8 (Ar-
CH2OH), 68.5 (C-6), 70.6 (ArCH2), 73.3 (ArCH2), 73.4 (ArCH2),
74.6 (C-2), 74.8 (ArCH2), 75.0 (C-5), 77.2 (C-4), 84.1 (C-3), 101.3
(C-1), 119.1, 124.4, 126.7, 126.9, 127.3, 127.4, 128.00, 128.03, 128.1,
129.7, 135.9, 136.0, 136.3, 137.5, 137.6, 140.3 ppm. ESI-MS: m/z ϭ
668.2 [M ϩ Naϩ]. C35H36ClN3O7 (646.13): calcd. C 65.06, H 5.62,
N 6.50; found C 64.81, H 5.55, N 6.46.
Solid-Supported Trisaccharide 22
Solid-Supported Tetrasaccharide 24: Solid-supported trisaccharide
22 (994 mg) was treated according to the General Procedures for
capping of free hydroxy groups and removal of the ClAzb groups,
then dried in vacuo. The resin (979 mg) was treated with thioglyco-
side 2 according to the General Procedure for the solid-phase glyco-
sylation to give solid-supported tetrasaccharide 24 as a slightly yel-
lowish powder (1.025 g).
1. General Procedure for Capping of Free Hydroxy Groups: Resin 4
(1.232 g, 0.2 mmol) was suspended in CH2Cl2 (5 mL). Triethyl-
amine (28 µL, 0.2 mmol), acetic anhydride (19 µL, 0.2 mmol) and
dimethylaminopyridine (6 mg, 0.05 mmol) were added and the mix-
ture was shaken at room temperature for 1 h. The resin was then
filtered off, washed with methanol (10 mL ϫ 2) and CH2Cl2
(10 mL ϫ 2), then subjected to the subsequent deprotection step.
4-(Hydroxymethyl)benzyl 3-O-(4-Azido-3-chlorobenzyl)-4,6-di-O-
2. General Procedure for Removal of the ClAzb Groups: Resin 4
(0.2 mmol) was suspended in CH2Cl2 (5 mL). Tributylphosphane
(148 µL, 0.6 mmol) was added and the mixture was shaken at room
temperature for 1 h. The resin was then filtered off, washed with
CH2Cl2, then suspended in THF (5 mL) again. DDQ (68 mg,
0.3 mmol), acetic acid (50 µL) and H2O (50 µL) were added and
the mixture was shaken at room temperature for 2 h, then filtered
off. The resin was washed with DMF (10 mL), a solution of tri-
ethylamine in DMF (5%, 10 mL ϫ 2), methanol (10 mL ϫ 2) and
CH2Cl2 (10 mL ϫ 2), and dried in vacuo to give resin 21 as a
slightly yellowish powder (1.201 g).
benzyl-β-
di-O-benzyl-β-
D
-glucopyranosyl-(1Ǟ3)-[β-
D
-glucopyranosyl-(1Ǟ6)]-4,6-
-gluco-
D
-glucopyranosyl-(1Ǟ3)-4,6-di-O-benzyl-β-
D
pyranoside (25): Solid-supported tetrasaccharide 24 (342 mg) was
cleaved according to the General Procedure for cleavage and the
product was purified by silica-gel column chromatography
(CH2Cl2/methanol, 98:2) to give tetrasaccharide 25 as a colorless
solid (22 mg, 31% from 4). For NMR measurement, tetrasacchar-
ide 25 was purified by reversed-phase HPLC (column: Waters Prep
Nova-Pak HR C18, 6 µm, 25 ϫ 100 mm, mobile phase: aceto-
nitrile/water, 7:3, flow rate: 7.0 mL/min, detection: UV 210 nm, re-
tention time ϭ 17.9 min). TLC (CH2Cl2/methanol, 9:1): Rf ϭ 0.32.
1H NMR (600 MHz, CDCl3 ϩ 5% CD3OD): δ ϭ 2.92 (ddd, J ϭ
3.0, 4.8, 9.0 Hz, 1 H, 5cЈ-H), 3.14 (dd, J ϭ 7.8, 9.0 Hz, 1 H, 2cЈ-
3. General Procedure for the Solid-Phase Glycosylation: Resin 21
(1.201 g, 0.2 mmol) was suspended in CH2Cl2 (5 mL). Glycosyl do-
˚
nor 3 (701 mg, 0.6 mmol), Sn(OTf)2 (63 mg, 0.15 mmol) and 4-A H), 3.29 (t, J ϭ 9.0 Hz, 1 H, 3cЈ-H), 3.34 (t, J ϭ 9.0 Hz, 1 H, 4cЈ-
Eur. J. Org. Chem. 2003, 3435Ϫ3445
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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3443

K. Egusa, S. Kusumoto, K. Fukase
FULL PAPER
H), 3.38 (ddd, J ϭ 2.1, 3.3, 9.0 Hz, 1 H, 5c-H), 3.42 (dd, J ϭ 2.1, pentasaccharide 28 was concentrated in vacuo and the residue was
10.8 Hz, 1 H, 6c-H), 3.45Ϫ3.48 (m, 2 H, 5b-H, 5a-H), 3.50Ϫ3.69 dissolved in THF (5 mL). DDQ (23 mg, 0.1 mmol), acetic acid (50
(m, 11 H), 3.70 (dd, J ϭ 3.0, 11.4 Hz, 1 H, 6cЈ-H), 3.75 (dd, J ϭ
µL) and H2O (50 µL) were added and the mixture was stirred at
2.2, 10.8 Hz, 1 H, 6a-H), 3.82 (t, J ϭ 9.0 Hz, 1 H, 3b-H), 3.86 (dd, room temperature for 2 h, then quenched by addition of a meth-
J ϭ 2.1, 9.6 Hz, 1 H, 6b-H), 3.90 (t, J ϭ 9.0 Hz, 1 H, 3a-H), 4.13 anolic solution of -ascorbic acid (18 mg, 0.1 mmol in 5 mL). The
(d, J ϭ 7.8 Hz, 1 H, 1cЈ-H), 4.31 (d, J ϭ 12.0 Hz, 1 H, ArCHH), mixture was passed through two columns of ion-exchange resins
4.36 (d, J ϭ 7.8 Hz, 1 H, 1a-H), 4.44 (d, J ϭ 12.0 Hz, 1 H,
ArCHH), 4.48 (d, J ϭ 10.2 Hz, 1 H, ArCHH), 4.52 (d, J ϭ form, 1 ϫ 5 cm) in this order, then concentrated in vacuo. The
11.4 Hz, 1 H, ArCHH), 4.50 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 4.59 residue was purified by reversed-phase HPLC (column: Waters
(d, J ϭ 11.4 Hz, 1 H, ArCHH), 4.61 (d, J ϭ 12.0 Hz, 1 H, Prep Nova-Pak HR C18, 6 µm, 25 ϫ 100 mm, mobile phase:
Amberlyst A-26 (OHϪ form, 2 ϫ 5 cm) and Amberlyst 15 (Hϩ
ArCHH), 4.62 (d, J ϭ 7.8 Hz, 1 H, 1c-H), 4.63 (s, 2 H, ArCH2OH),
4.64 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 4.70 (d, J ϭ 7.6 Hz, 1 H, 1b-
acetonitrile/water, 7:3, flow rate: 8.0 mL/min, detection: UV
210 nm, retention time: 16.5 min) using pentasaccharide 29 ob-
H), 4.71 (d, J ϭ 11.4 Hz, 1 H, ArCHH), 4.73 (d, J ϭ 12.0 Hz, 1 tained by the catch-and-release procedure as a reference substance
H, ArCHH), 4.88 (d, J ϭ 12.0 Hz, 1 H, ArCHH), 4.92 (d, J ϭ
11.4 Hz, 1 H, ArCHH), 4.97 (d, J ϭ 10.2 Hz, 1 H, ArCHH), 5.15
(d, J ϭ 11.4 Hz, 1 H, ArCHH), 7.07Ϫ7.15 (m, 3 H, ArH),
7.21Ϫ7.43 (m, 29 H, ArH) ppm. 13C NMR (151 MHz, CDCl3ϩ
10% CD3OD): δ ϭ 61.4 (C-6cЈ), 64.1 (ArCH2OH), 68.0 (6c-C),
68.1 (C-6b), 68.6 (C-6a), 69.7 (C-4cЈ), 70.5 (ArCH2), 73.2 (ArCH2),
73.3 (4 C, C-2a, C-2cЈ, ArCH2 ϫ 2), 73.4 (ArCH2), 74.2 (ArCH2),
74.5 (2 C, C-5a, C-5b), 74.6 (2c-C), 74.7 (C-2b), 74.7 (ArCH2), 75.0
(5c-C), 75.2 (C-5cЈ), 75.4 (C-3cЈ), 75.9 (C-4a), 76.2 (C-4b), 76.4 (4c-
C), 84.3 (C-3a), 84.5 (3c-C), 86.6 (C-3b), 101.0 (C-1a), 103.1 (2 C,
C-1b, C-1cЈ), 105.5 (1c-C), 119.3, 126.8, 127.2, 127.3, 127.4, 127.5,
127.6, 127.7, 127.9, 128.1, 128.2, 129.8, 135.9, 136.0, 136.6, 137.6,
137.8, 137.9, 138.3, 140.6 ppm. ESI-MS: m/z ϭ 1424.6 [M ϩ Naϩ].
C74H84ClN3O22 (1402.9).
to give 29 as a colorless solid (14 mg, 9% from 4).
Pentasaccharide 28: Detected by MS analysis of the filtrate. ESI-
MS: m/z ϭ 1740.9 [M ϩ Naϩ]. C94H108ClNO27 (1719.3).
4-(Hydroxymethyl)benzyl 4,6-Di-O-benzyl-β-
4,6-di-O-benzyl-β- -glucopyranosyl-(1Ǟ3)-[β-
anosyl-(1Ǟ6)]-4,6-di-O-benzyl-β- -glucopyranosyl-(1Ǟ3)-4,6-di-O-
benzyl-β-
-glucopyranoside (29): 1H NMR (600 MHz, CDCl3
D-glucopyranosyl-(1Ǟ3)-
D
D
-glucopyr-
D
D
ϩ
10% CD3OD): δ ϭ 2.79 (ddd, J ϭ 3.0, 4.8, 9.0 Hz, 1 H, 5cЈ-H),
3.06 (dd, J ϭ 7.7, 9.0 Hz, 1 H, 2cЈ-H), 3.20 (t, J ϭ 9.0 Hz, 1 H,
3cЈ-H), 3.23 (t, J ϭ 9.0 Hz, 1 H, 4cЈ-H), 3.28Ϫ3.31 (m, 2 H, 5c-H,
5d-H), 3.34 (dd, J ϭ 7.8, 9.0 Hz, 1 H, 2d-H), 3.35 (dd, J ϭ 1.7,
11.1 Hz, 1 H, 6c-H), 3.36Ϫ3.42 (m, 3 H, 6d-H, 5a-H, 5b-H),
3.43Ϫ3.54 (m, 11 H), 3.56 (t, J ϭ 9.0 Hz, 1 H, 4c-H), 3.58 (dd, J ϭ
4.4, 10.8 Hz, 1 H, 6a-H), 3.60 (dd, J ϭ 3.0, 12.0 Hz, 1 H, 6cЈ-H),
3.65 (dd, J ϭ 1.8, 10.8 Hz, 1 H, 6a-H), 3.70 (t, J ϭ 9.1 Hz, 1 H,
3c-H), 3.73 (t, J ϭ 8.8 Hz, 1 H, 3b-H), 3.82 (dd, J ϭ 1.7, 11.0 Hz,
1 H, 6b-H), 3.84 (t, J ϭ 9.0 Hz, 1 H, 3a-H), 4.07 (d, J ϭ 7.7 Hz,
1 H, 1cЈ-H), 4.20 (d, J ϭ 12.1 Hz, 1 H, ArCHH), 4.23 (d, J ϭ
12.1 Hz, 1 H, ArCHH), 4.29 (d, J ϭ 8.0 Hz, 1 H, 1a-H), 4.32 (d,
J ϭ 12.0 Hz, 1 H, ArCHH), 4.35 (d, J ϭ 12.1 Hz, 1 H, ArCHH),
4.37 (d, J ϭ 10.7 Hz, 1 H, ArCHH), 4.38 (d, J ϭ 11.0 Hz, 1 H,
ArCHH), 4.45 (d, J ϭ 12.4 Hz, 1 H, ArCHH), 4.45 (d, J ϭ
10.7 Hz, 1 H, ArCHH), 4.51 (d, J ϭ 12.0 Hz, 2 H, ArCHH ϫ 2),
4.53 (s, 2 H, ArCH2O), 4.54 (d, J ϭ 7.8 Hz, 1 H, 1d-H), 4.57 (d,
J ϭ 11.8 Hz, 1 H, ArCHH), 4.60 (d, J ϭ 8.0 Hz, 1 H, 1c-H), 4.64
(d, J ϭ 8.0 Hz, 1 H, 1b-H), 4.77 (d, J ϭ 10.7 Hz, 1 H, ArCHH),
4.82 (d, J ϭ 12.1 Hz, 1 H, ArCHH), 4.90 (d, J ϭ 10.7 Hz, 1 H,
Pentasaccharide 5: Solid-supported tetrasaccharide 24 (683 mg)
was treated according to the General Procedures for capping of
free hydroxy groups and removal of the ClAzb groups, then dried
in vacuo. The resin (675 mg) was treated with thioglycoside 2 ac-
cording to the General Procedure for the solid-phase glycosylation
to give solid-supported pentasaccharide 26 as a slightly yellowish
powder (701 mg). The resin was then treated according to the Gen-
eral Procedure for cleavage to give a mixture containing pentasac-
charide 5. The product was subjected to the catch-and-release puri-
fication. ESI-MS: m/z ϭ 1766.8 [M ϩ Naϩ]. C54H62ClN3O17
(1745.3).
Catch-and-Release Purification of Pentasaccharide 5: The crude
product containing pentasaccharide 5 was dispersed in CH2Cl2
(5 mL) and the insoluble substance was filtered off. The filtrate ArCHH), 5.03 (d, J ϭ 11.0 Hz, 1 H, ArCHH), 5.07 (d, J ϭ
was concentrated and dried in vacuo. The residue was dissolved in
CH2Cl2 (2 mL). To this solution was added triphenyl-
phosphaneϪNovaGelTM resin (Novabiochem, 0.48 mmol/g,
417 mg, 0.2 mmol, pre-washed with CH2Cl2) and the mixture was
shaken at room temperature for 2 h. The resin was then filtered
and washed with methanol (5 mL ϫ 2) and CH2Cl2 (5 mL ϫ 2).
The filtrate and the washings (containing pentasaccharide 28) were
combined and subjected to the recovery procedure (vide infra). The
resin was suspended in THF (2 mL), then DDQ (45 mg, 0.2 mmol),
acetic acid (50 µL) and H2O (50 µL) were added. The mixture was
shaken at room temperature for 2 h, then filtered and the resin was
11.3 Hz, 1 H, ArCHH), 7.10Ϫ7.30 (m, 39 H, ArH) ppm. 13C NMR
(151 MHz, CDCl3ϩ 10% CD3OD): δ ϭ 61.2 (C-6cЈ), 63.7 (Ar-
CH2OH), 68.0 (C-6b), 68.1 (6c-C), 68.2 (6d-C), 68.5 (C-6a), 69.6
(C-4cЈ), 70.3 (ArCH2), 72.9 (ArCH2), 73.0 (ArCH2), 73.1 (ArCH2),
73.2 (C-2cЈ), 73.4 (C-2a), 74.0 (ArCH2), 74.1 (ArCH2), 74.2
(ArCH2), 74.35 (ArCH2), 74.42 (C-5b), 74.5 (C-5a), 74.6 (C-2b),
74.7 (2 C, 5c-C, 5d-C), 74.77 (2c-C), 74.84 (2d-C), 75.1 (C-5cЈ),
75.6 (4c-C), 75.8 (C-3cЈ), 76.06 (C-4a), 76.09 (C-4b), 76.6 (3d-C),
77.1 (4d-C), 84.3 (C-3a), 86.6 (3c-C), 86.8 (C-3b), 100.9 (C-1a),
102.9 (C-1b), 103.1 (C-1cЈ), 104.5 (1c-C), 104.9 (1d-C), 126.5,
127.0, 127.2, 127.25, 127.28, 127.41, 127.44, 127.6, 127.7, 127.77,
washed with methanol (5 mL ϫ 2) and CH2Cl2 (5 mL ϫ 2). The 127.84, 127.9, 135.9, 137.48, 137.52, 137.8, 137.9, 138.20, 138.24,
filtrate and washings were combined and added to a methanolic
solution of -ascorbic acid (35 mg, 0.2 mmol in 5 mL). After stir-
ring for 5 min, the solution was passed through two columns of
ion-exchange resins Amberlyst A-26 (OHϪ form, 2 ϫ 5 cm) and
Amberlyst 15 (Hϩ form, 1.2 ϫ 4 cm) in this order. The solvent
was removed in vacuo to give pentasaccharide 29 as a colorless
solid (9 mg, 6% from 4).
140.4 ppm. ESI-MS: m/z ϭ 1601.7 [M ϩ Naϩ]. C87H102O27
(1579.8).
β-
D
-Glucopyranosyl-(1Ǟ3)-β-
D
-glucopyranosyl-(1Ǟ3)-[β-
D-gluco-
pyranosyl-(1Ǟ6)]-β-
D
-glucopyranosyl-(1Ǟ3)-α/β-
D
-glucopyranose
(1): Pd(OH)2/C (20%, 20 mg) was added to a solution of pentasac-
charide 29 (9.2 mg, 5.8 µmol) in methanol (3 mL) and the mixture
was stirred under 5 atm of hydrogen at room temperature over-
night. The Pd catalyst was removed by filtration and the filtrate
was concentrated in vacuo to give pentasaccharide 1 as a colorless
Recovery of Pentasaccharide from the Filtrate of the Catch-and-Re-
lease Purification: The filtrate of the ‘‘catching’’ process containing
3444
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
Eur. J. Org. Chem. 2003, 3435Ϫ3445

Solid-Phase Synthesis of a Phytoalexin Elicitor Pentasaccharide
FULL PAPER
[2i]
[2j]
solid (5.2 mg, quant., α/β ϭ 1:2). 1H NMR (600 MHz, D2O): α-
anomer (selected data): δ ϭ 5.05 (d, J ϭ 4.2 Hz, 1 H, 1a-H); β-
anomer: δ ϭ 3.11 (dd, J ϭ 7.8, 9.6 Hz, 1 H, 2cЈ-H), 3.17 (dd, J ϭ
8.4, 9.6 Hz, 1 H, 2d-H), 3.20 (t, J ϭ 9.0 Hz, 1 H, 4cЈ-H), 3.21 (t,
J ϭ 9.6 Hz, 1 H, 4d-H), 3.24 (dd, J ϭ 7.8, 9.0 Hz, 1 H, 2a-H), 3.26
(ddd, J ϭ 2.1, 4.2, 8.4 Hz, 1 H, 5cЈ-H), 3.27Ϫ3.35 (m, 7 H, 5a-H,
5c-H, 5d-H, 3cЈ-H, 3d-H, 4a-H, 4c-H), 3.36 (dd, J ϭ 8.4, 9.6 Hz,
1 H, 2c-H), 3.39 (t, J ϭ 8.4 Hz, 1 H, 2b-H), 3.42 (t, J ϭ 8.4 Hz, 1
H, 4b-H), 3.49Ϫ3.57 (m, 6 H, 5b-H, 6d-H, 6cЈ-H, 6c-H, 6a-H, 3a-
H), 3.59 (t, J ϭ 9.6 Hz, 1 H, 3c-H), 3.62 (t, J ϭ 8.4 Hz, 1 H, 3b-
H), 3.69Ϫ3.75 (m, 5 H, 6a-H, 6b-H, 6c-H, 6cЈ-H, 6d-H), 4.03 (m,
1 H, 6b-H), 4.33 (d, J ϭ 7.8 Hz, 1 H, 1cЈ-H), 4.49 (d, J ϭ 7.8 Hz,
1 H, 1a-H), 4.55 (d, J ϭ 8.4 Hz, 1 H, 1b-H), 4.70 (d, J ϭ 8.4 Hz,
1 H, 1d-H), 4.75 (d, J ϭ 8.4 Hz, 1 H, 1c-H) ppm. 13C NMR
(151 MHz, D2O): α-anomer (selected data): δ ϭ 91.8 (C-1a); β-
anomer: δ ϭ 60.5 (4 C, C-6a, 6c-C, C-6cЈ, 6d-C), 67.8Ϫ68.0 (3 C,
C-4a, C-4b, 4c-C), 68.5 (C-6b), 69.4 (2 C, C-4cЈ, 4d-C), 72.9Ϫ73.3
(4 C, C-2b, 2c-C, C-2cЈ, 2d-C), 74.1 (C-5b), 75.4Ϫ75.8 (7 C, C-2a,
C-3cЈ, 3d-C, C-5a, 5c-C, C-5cЈ, 5d-C), 83.6 (C-3b), 84.0 (2 C, C-
3a, 3c-C), 95.5 (C-1a), 102.3 (1c-C), 102.6 (3 C, C-1b, C-1cЈ, 1d-C)
ppm. ESI-MS: m/z ϭ 851.4 [M ϩ Naϩ]. C30H52O26 (828.72).
M. Grathwohl, R. R. Schmidt, Synthesis 2001, 2263Ϫ2272.
X. Wu, M. Grathwohl, R. R. Schmidt, Angew. Chem. 2002,
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[2k]
O. J. Plante, E. R. Palmacci, P. H. Seeberger, Science 2001,
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[3]
[4]
K. Egusa, K. Fukase, S. Kusumoto, Synlett 1997, 675Ϫ676.
K. Egusa, K. Fukase, Y. Nakai, S. Kusumoto, Synlett 2000,
27Ϫ32.
Commercially available from Argonaut Technologies, San
Carlos, California, http://www. argotech.com/resins/index.htm
K. Fukase, Y. Nakai, K. Egusa, J. A. Porco Jr., S. Kusumoto,
Synlett 1999, 1074Ϫ1078.
K. Egusa, S. Kusumoto, K. Fukase, Synlett 2001, 777Ϫ780.
For a review of catch-and-release purification and other poly-
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2002, 102, 3693Ϫ3716. [8b] L. A. Thompson, Curr. Opin. Chem.
Biol. 2000, 4, 324Ϫ337.
For other approaches to polymer-assisted purification in solid-
phase syntheses of oligosaccharides, see: refs.[2d,2l]
T. Yamaguchi, A. Yamada, N. Shibuya, Plant Cell 2000, 12,
817Ϫ826.
[5]
[6]
[7]
[8]
[9]
[10]
[11]
Synthesis of 1 had already been reported: T. Amaya, H.
Tanaka, T. Yamaguchi, N. Shibuya, T. Takahashi, Tetrahedron.
Lett. 2001, 42, 9191Ϫ9194 (by solution-phase synthesis) and
ref.[2c] (by solid-phase synthesis).
Acknowledgments
This work was financially supported in part by Grants-in-Aid for
Scientific Research No. 11558080 and ‘‘Research for the Future’’
Program No. 97L00502 from the Japanese Society for the Pro-
motion of Science and by Special Coordination Funds and Grants-
in-Aid for Creative Scientific Research ‘‘In vivo Molecular Science
for Discovery of New Biofunctions and Pharmaceutical Drugs’’
No. 13NP0401 from the Ministry of Education, Culture, Sports,
Science and Technology, Japan.
[12]
Other examples of solid-phase syntheses of oligosaccharides
[12a]
expressing phytoalexin elicitor activity:
R. Verduyn, P. A.
M. van der Klein, M. Douwes, G. A. van der Marel, J. H. van
Boom, Recl. Trav. Chim. Pays-Bas 1993, 112, 464Ϫ466. [12b] K.
C. Nicolaou, N. Winssinger, J. Pastor, F. Deroose, J. Am. Chem.
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Li, J. Pastor, N. Winssinger, Angew. Chem. 1998, 110,
1636Ϫ1638; Angew. Chem. Int. Ed. 1998, 37, 1559Ϫ1561, and
ref.[2k]
.
[13]
[14]
[15]
[16]
[17]
[18]
Commercially available from Wako Pure Chemicals Industries,
Ltd., http://www.wako-chem.co.jp/
L.-X. Wang, N. Sakairi, H. Kuzuhara, J. Chem. Soc., Perkin
Trans. 1 1990, 1677Ϫ1682.
M. P. Reddy, J. B. Rampal, S. L. Beaucage, Tetrahedron Lett.
1987, 28, 23Ϫ26.
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[1a]
For a review, see:
K. Fukase, in Glycoscience, vol. II (Eds.:
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Seeberger, W.-C. Haase, Chem. Rev. 2000, 100, 4349Ϫ4393.
H. M. I. Osborn, T. H. Khan, Tetrahedron 1999, 55,
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R. R. Schmidt, J. Michel, M. Roos, Liebigs Ann. Chem. 1984,
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[2]
[2a]
For recent advances, see:
J. 2001, 7, 2382Ϫ2389.
T. Zhu, G.-J. Boons, Chem. Eur.
[2b]
T. Takahashi, H. Inoue, Y. Yama-
Commercially available from Mitsubishi Chemical Corpor-
ation, http://www.diaion.com/
For investigations of other highly crosslinked macroporous re-
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Janda, Bioorg. Med. Chem. Lett. 1998, 8, 2363Ϫ2368.
A. R. Mitchell, S. B. H. Kent, M. Engelhard, R. B. Merrifield,
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B. F. Ginsin, Anal. Chim. Acta 1972, 58, 248Ϫ249.
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mura, T. Doi, Angew. Chem. 2001, 113, 3330Ϫ3333; Angew.
[2c]
Chem. Int. Ed. 2001, 40, 3230Ϫ3233.
T. Takahashi, A. Ok-
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H. Ando, S. Manabe, Y. Nakahara, Y. Ito, Angew. Chem. 2001,
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[2e]
T. Kanemitsu, C.-H. Wong, O. Kanie, J. Am. Chem. Soc.
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Org. Chem. 2000, 2803Ϫ2808.
Schmidt, Eur. J. Org. Chem. 2001, 2067Ϫ2073.
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[21]
L. Knerr, R. R. Schmidt, Eur. J.
[2g]
F. Roussel, L. Knerr, R. R.
[2h]
F. Roussel,
Received April 23, 2003
Eur. J. Org. Chem. 2003, 3435Ϫ3445
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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3445