Chemical synthesis of β-arabinofuranosyl containing oligosaccharides derived from plant cell wall extensins

Article abstract of DOI:10.1039/c3ob40958a

Extensins are plant-derived glycoproteins that are densely modified by oligo-arabinofuranosides linked to hydroxyproline residues. These glycoproteins have been implicated in many aspects of plant growth and development. Here, we describe the chemical syn

Full text of DOI:10.1039/c3ob40958a

Organic &
Biomolecular Chemistry
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Chemical synthesis of β-arabinofuranosyl containing
oligosaccharides derived from plant cell wall
extensins†
Cite this: Org. Biomol. Chem., 2013, 11,
5136
Sophon Kaeothip and Geert-Jan Boons*
Extensins are plant-derived glycoproteins that are densely modified by oligo-arabinofuranosides linked to
hydroxyproline residues. These glycoproteins have been implicated in many aspects of plant growth and
development. Here, we describe the chemical synthesis of a tetrameric β(1–2)-linked arabinofuranoside
that is capped by an α(1–3)-arabinofuranoside and a similar trisaccharide lacking the capping moiety. The
challenging β(1–2)-linked arabinofuranosides were installed by using an arabinofuranosyl donor pro-
tected with 3,5-O-(di-tert-butylsilane) and a C-2 2-methylnaphthyl (Nap) ether. It was found that the
cyclic silane-protecting group of the glycosyl donor greatly increased β-anomeric selectivity. It was,
however, imperative to remove the silane-protecting group of an arabinosyl acceptor to achieve optimal
anomeric selectivities. The anomeric linker of the synthetic compounds was modified by a biotin moiety
for immobilization of the compounds to microtiter plates coated with streptavidine. The resulting micro-
titer plates were employed to screen for binding against a panel of antibodies elicited against plant cell
wall polysaccharides.
Received 6th May 2013,
Accepted 4th June 2013
DOI: 10.1039/c3ob40958a
www.rsc.org/obc
structures of the oligo-arabinosides of Nicotina tabacum have
been determined, and it was found that they are mainly com-
Introduction
Hydroxyproline-rich glycoproteins are ubiquitous components posed of a tetrameric β(1–2)-linked arabinofuranoside that is
of the growing plant cell wall that have been implicated in all capped by an α(1–3)-arabinofuranoside (compound 1, Fig. 1).
aspects of plant growth and development including stress In addition, a trisaccharide was detected lacking the capping
responses.1 This family of glycoproteins, which include moiety (compound 2, Fig. 1).7
proline-rich proteins, arabinogalactans and extensins, gener-
Recently, a collection of 180 plant cell wall glycan-binding
ally consist of four- to six amino acid repeating motifs that are monoclonal antibodies (mAbs) was reported to facilitate an in-
extensively O-glycosylated at hydroxyproline by arabinosides or depth analysis of plant cell wall structure, function, dynamics,
arabinogalactans.2,3 Occasionally, serine residues can also be and biosynthesis.8 An enzyme-linked immunosorbent assay-
glycosylated, mainly by a single galactoside.
based screen against a diverse panel of 54 plant polysacchar-
The conservation and widespread occurrence of only a few ides was used to characterize the binding patterns of these
hydroxyproline-rich glycoprotein modules indicate that they mAbs. We are generating well-defined synthetic oligosacchar-
represent functional units whose roles depend on the identity ides derived from plant cell wall polysaccharides to determine
and precise arrangement of the pendent oligosaccharides. the fine epitope specificities of the mAbs.9,10 As part of this
Indeed, for most hydroxyproline-rich glycoproteins, the
peptide backbone is completely shielded by carbohydrates,
very similar to what is observed for eukaryotic mucins,4,5 and
therefore their interactive molecular surface is probably deter-
mined by carbohydrate structures.
Plant derived extensins6 are modified by linear L-arabino-
furanosides, which are linked to hydroxyproline residues. The
Complex Carbohydrate Research Center, The University of Georgia, 315 Riverbend
Road, Athens, GA 30602, USA. E-mail: gjboons@ccrc.uga.edu; Fax: +1 706-542-4412
†Electronic supplementary information (ESI) available: 1H NMR spectra of syn-
thetic compounds. See DOI: 10.1039/c3ob40958a
Fig. 1 Arabinofuranosides of plant elastins.
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project, we report here the chemical synthesis of two arabino-
furanosides derived from plant cell wall extensins.
Results and discussion
In general, α-arabinofuranosides can be obtained in a straight-
forward manner by neighboring group participation of an acyl
ester at C-2 of a furanosyl donor. On the other hand, the
stereoselective introduction of β-arabinofuranosides, which
requires glycosyl donors having a non-participating protecting
group at C-2, often leads to the formation of mixtures of α/β
anomers.11,12 Furanosides exhibit weak anomeric effects,
which differ little between the α- and β-anomer, making it
difficult to exploit in situ anomerization protocols for control-
ling anomeric selectivities.13,14 Furthermore, furanosides are
inherently flexible due to their ability to assume several twist
and envelope conformations, which can interconvert via
pseudo-rotational itineraries.12 As a result, furanosides can
glycosylate through different transition states, which may com-
promise anomeric selectivity.15
Recently, we reported a practical approach for the stereo-
selective introduction of β-arabinofuranosides by locking a
donor in a conformation in which attack from the β-face is
favored.16 The glycosyl donor was designed by analyzing opti-
mized geometries of low energy conformers of the arabinofura-
nosyl oxa-carbenium ion. The Newman projection of the E3
conformer indicated that nucleophilic attack from the α-face is
disfavored because an eclipsed H-2 will be encountered. On
the other hand, an approach from the β-face was expected to
be more favorable because it will experience only staggered
substituents. The arabinofuranosyl oxa-carbenium ion could
be locked in the E3 conformation by employing a 3,5-O-(di-tert-
butylsilane)-protecting group, which places C-5 and O-3 in a
pseudo-equatorial orientation resulting in a perfect chair con-
formation of the protecting group. We have shown that the
new glycosyl donor gives excellent β-anomeric selectivities in
glycosylations with glycosyl acceptors having primary and sec-
ondary alcohols. Several other methods have been introduced
to conformationally constrain the five-membered ring
structure of arabinofuranosyl donors to control β-anomeric
selectivity.17–24
We envisaged that target oligosaccharides 27 and 28
(Scheme 3, spacer modified analogs of 1 and 2), which contain
multiple β-arabinofuranosides, could readily be prepared by
using arabinofuranosyl donor 3 (Scheme 1). This compound
contains a 3,5-O-(di-tert-butylsilane) protecting group that was
expected to control the β-anomeric selectivity of the glycosyla-
tions. Furthermore, the 2-methylnaphthyl (Nap) ether at C-2
can be removed by oxidation using 2,3-dichloro-5,6-dicyano-
1,4-benzoquinone (DDQ)25,26 to reveal an alcohol that can be
glycosylated to install additional β(1–2)-linked arabinofurano-
sides of the target compounds. Furthermore, donors 420 and
516 were also employed to explore the importance of the 3,5-O-
(di-tert-butylsilane) protecting group for the stereoselective
outcome of the glycosylations.
Scheme 1 Installation of an anomeric linker.
First, attention was focused on the installation of an
anomeric linker, which is required for the conjugation of the
oligosaccharides to carrier proteins or biotin in order to facili-
tate immunological studies. Thus, a glycosylation of confor-
mationally constrained glycosyl donor 3 with 3-bromopropan-
1-ol in the presence of the powerful thiophilic promoter
system NIS/AgOTf27 in DCM at −40 °C gave spacer modified
6 in an excellent yield (92%) as predominantly the β-anomer
(α/β 1/4.5). On the other hand, similar glycosylations with the
more flexible glycosyl donors 4 and 5 provided spacer-linked
derivatives 7 and 8, respectively, with lower β-anomeric selec-
tivities (α/β 1/2.5 and 1/2.1, respectively). The anomeric con-
figuration of the products was readily assigned by
a
combination of chemical shift and coupling constant data.
In this respect, α-arabinofuranosides are characterized by
3JH-1,H-2 = 1–3 Hz and δ C-1 = 104–110 ppm whereas similar
3
β-glycosides have JH-1,H-2 = 4–5 Hz and δ C-1 = 97–104 ppm
values.16,21,28 The 13C chemical shift of the anomeric carbon
and geminal coupling constant of H-1 of the major product of
6 was in the expected region for a β-anomer (δ C-1 = 100.8 ppm
and 3JH-1,H-2 = 5.1 Hz).
Compound 6 was treated with sodium azide in DMF to
convert the bromide into an azide to give compound 9. The
Nap ether of 9 was removed by oxidation with DDQ in a
mixture of dichloromethane and methanol to provide glycosyl
acceptor 10 in 84% yield. In addition, glycosyl acceptors 14
and 15 were prepared by removal of the di-tert-butylsilane pro-
tecting group of 9 with HF-pyridine in dichloromethane29 to
give 11, which was benzylated or benzoylated using standard
conditions to provide 12 and 13, followed by oxidation with
DDQ to remove the Nap ethers.
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Scheme 2 Protecting groups of the glycosyl donor and acceptor influence the
anomeric outcome of arabinofuranosylations.
Interestingly, glycosylations of silane protected donor 3
with glycosyl acceptors 10, 14 and 15 gave, in each case, the
respective coupling products 16, 17 and 18 in high yield but
with differing anomeric selectivities (Scheme 2). In particular,
the use of di-tert-butylsilane protected glycosyl acceptor 10
gave disaccharide 16 as a 1/1 mixture of α/β anomers and
probably in this specific case, unfavorable steric interactions
between the bulky silane protecting groups of the acceptor and
donor in the transition state leading to the β-anomer are
responsible for the low anomeric selectivity.30 The use of C-3
and C-5 benzoylated and benzylated acceptors 14 and 15 led to
the formation of the corresponding disaccharides 17 and 18,
respectively, with reasonable to good β-anomeric selectivities,
and in particular, the benzylated acceptor behaved very well
giving spacer modified 18 as a 1/6.5 mixture of α/β anomers
that could readily be separated by silica gel column chromato-
graphy. As expected, a glycosylation of the conformational flex-
ible donor 5 with glycosyl acceptor 15 in the presence of NIS/
AgOTf gave disaccharide 19 as a 1/1.7 mixture of α/β anomers,
highlighting that appropriate protection of the glycosyl donor
and acceptor is critical for achieving optimal β-anomeric
selectivity.
Scheme 3 Assembly of arabinofuranosides.
using benzoic acid in the presence of 2-chloro-1-methyl-
pyridinium iodide (CMPI) and 1,4-diazabicyclo[2.2.2]octane
(DABCO). Finally, a glycosylation of 24 with 25 in the presence
of NIS/AgOTf27 gave tetrasaccharide 26 in an excellent yield of
87% as the only α-anomer due to the neighboring group par-
ticipation of the C-2 benzoyl ester of donor 25.
Deprotection of 26 and 23 to afford the target compounds
27 and 28, respectively, was accomplished by a two-step pro-
cedure involving saponification of the benzoyl esters using
sodium methoxide in methanol, followed by catalytic hydro-
genolysis over Pd/C to remove the benzyl and Nap ethers and
convert the azido moiety into an amine. NMR and MS analyses
confirmed the structural integrity of tetrasaccharide 27 (Araf,
3JH-1,H-2 = 4.5 Hz, δ C-1 = 100.0 ppm, Araf′, JH-1,H-2 = 4.5 Hz,
3
3
δ C-1 = 99.9 ppm, Araf″, JH-1,H-2 = 4.5 Hz, δ C-1 = 97.7 ppm,
Araf′′′, H-1, singlet, δ C-1 = 108.1 ppm). Finally, the amine of
the anomeric linker of compounds 28 and 29 was exploited to
install a biotin moiety by reaction with biotin N-hydroxysuccin-
imide ester (NHS-Biotin) in phosphate buffer at pH 7.4 to give,
after purification by reverse phase C-18 column chromato-
graphy, biotin modified derivatives 29 and 30, respectively.
The compounds were immobilized on microtiter plates
coated with streptavidine, which were subsequently used to
probe for the binding of a panel of 180 anti-plant polysacchar-
ide mAbs. Surprisingly, none of the antibodies exhibited
The process of removal of the di-tert-butylsilane protecting
group using HF-pyridine followed by protection of the result-
ing diol as benzyl ethers and oxidative removal of the Nap
ether was repeated to give glycosyl acceptor 21 (Scheme 3). An
NIS/AgOTf promoted glycosylation of the latter compound
with glycosyl donor 3 gave trisaccharide 22 as a separable
1/4.5 mixture of α/β anomers. Next, trisaccharide 22 was con-
verted into glycosyl acceptor 24 having a free C-3″ hydroxyl by
removal of the di-tert-butylsilane protecting group (→23) fol-
lowed by regioselective benzoylation of the primary hydroxyl
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binding for compounds 29 and 30, indicating that the array of spectra were obtained on a Bruker model Ultraflex MALDI-TOF
antibodies cannot detect oligo-arabinofuranosides. Current mass spectrometer.
efforts are focused on conjugation of the oligosaccharides to a
carrier protein, and the use of the resulting conjugates for methyl-1-thio-α-L-arabinofuranoside (3). To
mouse immunizations and subsequent mAb generation using phenyl 3,5-O-(di-tert-butylsilanediyl)-1-thio-α-L-arabinofurano-
Phenyl
3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)-
a
solution of
conventional hybridoma technology.
side (4.0 g, 10.4 mmol) in DMF (30 mL) at 0 °C was added
2-(bromomethyl)naphthalene (3.47 g, 15.7 mmol) and NaH
(60% dispersion in oil, 0.62 g, 15.7 mmol) in four portions.
The reaction mixture was kept stirring at 0 °C for 3 h. Upon
completion, the reaction mixture was poured into ice-water
(50 mL), stirred until cessation of H2 evolution, and then
extracted with ethyl acetate–diethyl ether (1/1, v/v, 3 × 100 mL).
The combined organic phase was washed with cold water
(200 mL), separated, dried (MgSO4) and concentrated in vacuo.
The residue was purified by silica gel column chromatography
(ethyl acetate–hexane gradient elution) to afford the title com-
pound as a colorless syrup (4.28 g, 75%). Analytical data for 3:
Rf = 0.57 (ethyl acetate–hexane, 1.2/8.8, v/v); [α]2D2 = −119.7
(c 1.0, CHCl3); 1H NMR (300 MHz, CDCl3): δ, 7.89–7.86 (m, 4H,
ArH), 7.57–7.42 (m, 5H, ArH), 7.25–7.23 (m, 3H, ArH), 5.49 (d,
J1,2 = 5.4 Hz, H-1), 5.05–4.93 (dd, J = 12.3 Hz, 2H, CH2Nap),
4.38 (m, 1H, H-5b), 4.23 (dd, J3,4 = 6.9 Hz, 1H, H-3), 4.05 (dd,
1H, J2,3 = 5.4 Hz, H-2), 3.99 (m, 2H, H-2, 5a), 1.12 (s, 9H, t-Bu),
1.02 (s, 9H, t-Bu) ppm; 13C NMR (75 MHz, CDCl3): δ, 135.20,
134.63, 133.56, 133.37, 131.52, 129.14, 128.47, 128.19, 127.97,
127.23, 126.36, 126.21, 126.19, 90.14, 86.82, 81.52, 73.98,
72.50, 67.55, 27.74, 27.35, 22.88, 20.36 ppm; MALDI HR-MS
calc. for C30H38NaO4SSi [M + Na]+: 545.2158, found 545.2163.
Conclusions
A concise synthetic strategy has been developed for the prepa-
ration of 1,2-linked β-arabinofuranosides commonly found in
plant glycoproteins. The difficult to introduce β-arabinofurano-
sides were installed using an arabinofuranosyl donor protected
by a cyclic 3,5-O-(di-tert-butylsilane). A temporary Nap ether at
C-2 of the donor made it possible to further extend the glycosy-
lation products by 1,2-arabinofuranosides. The silane-protect-
ing group of the acceptors, however, needed to be removed to
achieve optimal β-anomeric selectivities. The use of confor-
mationally more flexible donors led to glycosylation products
having poor anomeric ratios. Thus, it has been found that
proper protection of the arabinofuranosyl donor and acceptor
is critical for achieving high β-anomeric selectivity. A panel of
180 plant cell wall glycan-binding monoclonal antibodies that
is commonly employed for in-depth analysis of the plant cell
wall structure, function, dynamics, and biosynthesis did,
unfortunately, not recognize the synthetic oligosaccharides.
Phenyl
3,5-O-(tetraisopropyldisiloxane-1,3-diyl)-2-O-(2-
(4). To
naphthyl)methyl-1-thio-α-L-arabinofuranoside
a
solution of phenyl 1-thio-α-L-arabinofuranoside (0.43 g,
1.77 mmol) and imidazole (0.53 g, 7.81 mmol) in DMF (7 mL)
Experimental
General procedures
was
added
dropwise
dichlorotetraisopropyldisiloxane
Column chromatography was performed on silica gel 60 (EM (0.63 mL, 1.95 mmol). After 4 h, excess silylation agent was
Science, 70–230 mesh). Reactions were monitored by thin-layer decomposed by the addition of methanol (2 mL), which was
chromatography (TLC) on Kieselgel 60 F254 (EM Science), and followed by the addition of ethyl acetate (25 mL). The solution
compounds were detected by examination under UV light and was poured into saturated aqueous sodium chloride (30 mL)
by charring with 10% sulfuric acid in MeOH. Solvents were and extracted with ethyl acetate (3 × 30 mL). The combined
removed under reduced pressure at <40 °C. MeOH was dried organic layers were dried (MgSO4), and the solvents were
by refluxing with magnesium methoxide and then was distilled removed in vacuo. The residue was purified by column
and stored under argon. Pyridine was dried by heating under chromatography on silica gel (ethyl acetate–hexane gradient
refluxing over CaH2 and then distilled and stored over mole- elution) to afford phenyl 3,5-O-(tetraisopropyldisiloxane-1,3-
cular sieves (3 Å). CH2Cl2 was freshly distilled from calcium diyl)-1-thio-α-L-arabinofuranoside as a colorless syrup (0.60 g,
hydride under nitrogen prior to use. AgOTf was co-evaporated 70%). MALDI HR-MS calc. for C23H40NaO5SSi2 [M + Na]+:
with toluene and dried in vacuo for 2–3 h prior to application. 545.2158, found 545.2163. Phenyl 3,5-O-(tetraisopropyldisilox-
Molecular sieves (4 Å) were flame activated under vacuum ane-1,3-diyl)-1-thio-α-L-arabinofuranoside (0.35 g, 0.72 mmol)
prior to use. All reactions were carried out under an argon was dissolved in DMF (8 mL) and 2-(bromomethyl)naphtha-
atmosphere. 1H NMR and 13C NMR spectra were recorded with lene (0.23 g, 1.08 mmol) and NaH (60% dispersion in oil,
Varian spectrometers (models Inova300 and Inova500) 43 mg, 1.08 mmol) were added. The reaction mixture was kept
equipped with Sun workstations. 1H NMR spectra were stirring at 0 °C for 3 h. Upon completion, the reaction was
recorded in CDCl3 and referenced to residual CHCl3 at poured into ice-water (20 mL), stirred until cessation of H2
7.24 ppm, and 13C NMR spectra were referenced to the central evolution, and then extracted with ethyl acetate–diethyl ether
peak of CDCl3 at 77.0 ppm. Assignments were made by stan- (1/1, v/v, 3 × 30 mL). The combined organic phase was washed
dard gCOSY and gHSQC. Optical rotations were measured with cold water (75 mL), separated, dried (MgSO4) and concen-
using a ‘Jasco P-1020’ polarimeter and the specific rotations trated in vacuo. The residue was purified by column chromato-
are provided in degrees cm3 dm−1 g−1. High resolution mass graphy on silica gel (ethyl acetate–hexane gradient elution) to
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afford the title compound as a colorless syrup (0.34 g, 77%). room temperature and stirring was continued for 15 min. The
Analytical data for 4: Rf = 0.57 (ethyl acetate–hexane, 1.2/8.8, reaction was quenched by the addition of Et3N or pyridine.
v/v); [α]2D2 = −67.2 (c 1.0, CHCl3); 1H NMR (300 MHz, CDCl3): δ, The suspension was diluted with CH2Cl2 (15 mL) and filtered
7.81 (m, 4H, ArH), 7.47 (m, 5H, ArH), 7.25 (m, 3H, ArH), 5.51 through a pad of Celite, and the filtrate was washed succes-
(d, 1H, J1,2 = 4.2 Hz, H-1), 4.91–4.81 (dd, 2H, J = 12.0 Hz, sively with 10% Na2S2SO3 (10 mL) and brine (10 mL). The
CH2Nap), 4.38 (m, 1H, H-3), 4.11 (dd, 1H, J2,3 = 5.7 Hz, H-2), organic layer was dried (MgSO4) and concentrated in vacuo to
4.01 (m, 3H, H-4, 5a, 5b), 1.02–1.09 (m, 28H, CH(CH3)2) ppm; give a residue. The residue was purified by silica gel chromato-
13C NMR (75 MHz, CDCl3): δ, 135.32, 131.07, 129.06, 128.28, graphy (ethyl acetate–hexane gradient elution) to afford the
128.14, 127.90, 127.23, 126.85, 126.29, 126.11, 126.00, 89.67, corresponding oligosaccharide.
89.36, 80.38, 76.13, 73.05, 61.34, 17.68, 17.55, 17.37, 17.35,
17.29, 17.22, 13.77, 13.36, 13.06, 12.78 ppm; MALDI HR-MS methyl-β-L-arabinofuranoside (6). Analytical data for 6: Rf = 0.53
calc. for C34H48NaO5SSi2 [M + Na]+: 647.2659, found 647.2659. (ethyl acetate–hexane, 2/8, v/v); [α]D22 = +57.9 (c 1.0, CHCl3);
Phenyl
3,5-di-O-benzyl-2-O-(2-naphthyl)methyl-1-thio-α-L- 1H NMR (300 MHz, CDCl3): δ, 7.86–7.81 (m, 4H, ArH),
3-Bromopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)-
arabinofuranoside (5). Hydrogen fluoride–pyridine (85 µL, 7.56–7.47 (m, 3H, ArH), 4.97 (dd, 2H, J = 12.0 Hz, CH2Nap),
3.28 mmol) was carefully diluted with pyridine (5 mL) at 0 °C. 4.93 (d, 1H, J1,2 = 5.1 Hz, H-1), 4.37 (dd, 1H, J4,5a = 9.0 Hz,
The resulting solution was added slowly to a stirred at 0 °C H-4), 4.30 (dd, 1H, J2,3 = 5.1 Hz and J3,4 = 9.0 Hz, H-3),
suspension of 3 (0.45 g, 0.82 mmol) in CH2Cl2 (5 mL) and the 4.00–3.87 (m, 2H, H-2, 5b), 3.83 (m, 1H, 1/2CH2-linker),
reaction was allowed to proceed for 2 h at 0 °C. The reaction 3.68–3.48 (m, 4H, H-5a, 1/2CH2-linker, CH2-linker), 2.15 (m,
mixture was diluted with CH2Cl2 (15 mL), washed with water 2H, CH2-linker), 1.10 (s, 9H, t-Bu), 1.01 (s, 9H, t-Bu) ppm;
(10 mL) and NaHCO3 (10 mL), dried (MgSO4) and concentrated 13C NMR (75 MHz, CDCl3): δ, 135.51, 133.45, 13.29, 128.37,
in vacuo. The crude product was purified by flash chromato- 138.12, 127.93, 127.16, 126.34, 126.23, 126.16, 100.80, 80.77,
graphy on silica gel (EtOAc–hexane, 1/1, v/v) to give a diol inter- 78.95, 73.80, 72.18, 68.71, 66.49, 32.91, 30.74, 27.79, 27.42,
mediate (0.28 g, 81%). MALDI HR-MS calc. for C22H22NaO4S 22.83, 20.34 ppm; MALDI HR-MS calc. for C27H39BrNaO5Si
[M + Na]+: 428.1034, found 428.1041. The diol (0.25 g, [M + Na]+: 573.1648, found 573.1639.
0.62 mmol) and benzyl bromide (0.22 mL, 1.85 mmol) were
3-Bromopropyl 2-O-(2-naphthyl)methyl-3,5-O-(tetraisopropyl-
dissolved in DMF (5 mL), cooled to 0 °C, and treated with NaH siloxane-1,3-diyl)-β-L-arabinofuranoside (7). Analytical data for
(60% dispersion in oil, 74 mg, 1.85 mmol, four portions). The 7: Rf = 0.55 (ethyl acetate–hexane, 2/8, v/v); [α]2D2 = +62.9 (c 1.0,
1
reaction mixture was stirred for 2 h, then poured into ice-water CHCl3); H NMR (300 MHz, CDCl3): δ, 7.82 (m, 4H, ArH), 7.48
(20 mL), stirred for 10 min, and extracted with ethyl acetate– (m, 3H, ArH), 4.87–4.76 (dd, 2H, J = 12.0 Hz, CH2Nap), 4.70 (d,
diethyl ether (1/1 v/v, 3 × 30 mL). The combined organic phase 1H, J1,2 = 4.2 Hz, H-1), 4.51 (dd, 1H, J3,4 = 5.1 Hz, H-3), 3.98
was washed with cold water (50 mL), separated, dried (MgSO4) (dd, 1H, J2,3 = 7.5 Hz, H-2), 3.94 (dd, 1H, J = 9.0 Hz, H-5b),
and concentrated in vacuo. The residue was purified by 3.83–3.78 (m, 2H, H-4, H5a), 3.74 (m, 1H, 1/2CH2-linker),
column chromatography on silica gel (ethyl acetate–hexane 3.58–3.37 (m, 3H, 1/2CH2-linker, CH2-linker), 2.10 (m, 2H,
gradient elution) to afford the title compound as a colorless CH2-linker), 1.12–1.02 (m, 28H, CH(CH3)2) ppm; 13C NMR
syrup (0.27 g, 79%). Analytical data for 5: Rf = 0.47 (ethyl (75 MHz, CDCl3): δ, 135.65, 133.43, 133.26, 128.33, 128.07,
1
acetate–hexane, 2/8, v/v); [α]2D2 = −86.9 (c 1.0, CHCl3); H NMR 127.91, 126.85, 126.36, 126.16, 126.05, 99.99, 84.92, 82.12,
(300 MHz, CDCl3): δ, 7.80–7.74 (m, 4H, ArH), 7.49–7.39 (m, 5H, 78.19, 73.02, 66.86, 65.50, 32.79, 30.76, 17.80, 17.70, 17.67,
ArH), 7.29–7.20 (m, 13H, ArH), 4.76 (d, 1H, J = 12.0 Hz, 17.63, 17.37, 17.31, 17.25, 17.23, 13.73, 13.52, 13.10,
1/2CH2Nap), 4.62 (d, 1H, J = 12.0 Hz, 1/2CH2Nap), 4.58–4.42 12.72 ppm; MALDI HR-MS calc. for C31H49BrNaO6Si [M + Na]+:
(m, 4H, 2 × CH2Ph), 4.39 (m, 1H, H-4), 4.17 (dd, 1H, J2,3
=
675.2149, found 675.2155.
3.0 Hz, H-3), 4.08 (dd, 1H, J3,4 = 6.9 Hz, H-3), 3.71–3.62 (dd, J =
3-Bromopropyl 3,5-di-O-benzyl-2-O-(2-naphthyl)methyl-β-L-
3.90 Hz and 6.90 Hz, H-5a, 5b) ppm; 13C NMR (75 MHz, arabinofuranoside (8). Analytical data for 8: Rf = 0.56 (ethyl
CDCl3): δ, 138.41, 138.02, 135.20, 135.07, 133.54, 133.42, acetate–hexane, 3/7, v/v); [α]2D2 = +35.2 (c 1.0, CHCl3); H NMR
1
131.58, 129.25, 128.73, 128.69, 128.63, 128.30, 128.15, 128.07, (300 MHz, CDCl3): δ, 7.81 (m, 2H, ArH), 7.47 (m, 2H, ArH),
127.96, 127.47, 127.29, 126.58, 126.42, 126.20, 90.66, 88.82, 7.33–7.28 (m, 8H, ArH), 4.89 (d, 1H, J1,2 = 2.1 Hz, H-1),
83.73, 80.84, 73.67, 72.62, 72.60, 68.34 ppm; MALDI HR-MS 4.76–4.50 (m, 6H, CH2Nap, 2 × CH2Ph), 4.12 (m, 3H, H-2, 3, 4),
calc. for C36H34NaO4S [M + Na]+: 585.2075, found 585.2083.
3.76 (m, 1H, 1/2CH2-linker), 3.53 (m, 2H, J = 5.1 Hz, H-5a,
H5b), 3.49–3.39 (m, 3H, 1/2CH2-linker, CH2-linker), 2.03 (m,
2H, CH2-linker) ppm; 13C NMR (75 MHz, CDCl3): δ, 138.41,
138.25, 135.44, 133.43, 133.31, 128.59, 128.58, 128.47, 128.13,
General glycosylation procedure for the synthesis of mono-
and disaccharides
A mixture of a thioglycoside (0.125 mmol), an alcohol 128.07, 127.92, 127.86, 127.07, 126.41, 126.26, 126.13, 101.05,
(0.10 mmol) and freshly activated molecular sieves (4 Å, 84.49, 83.38, 80.45, 73.56, 72.83, 72.66, 72.60, 65.49, 32.69,
350 mg) in CH2Cl2 (2.0 mL) was stirred under argon at room 30.83 ppm; MALDI HR-MS calc. for C33H35BrNaO5 [M + Na]+:
temperature for 30 min. After the mixture was cooled (−40 °C), 613.1566, found 613.1572.
NIS (0.1875 mmol) followed by AgOTf (0.0375 mmol) were
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)-
added. The reaction mixture was allowed to warm slowly to methyl-β-L-arabinofuranoside (9). Compound
6
(0.67 g,
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1.21 mmol) and sodium azide (0.79 g, 12.1 mmol) were dis- (0.29 mL, 2.54 mmol) was added dropwise. The reaction
solved in DMF (10 mL). The reaction mixture was stirred at mixture was stirred at room temperature for 2 h. The reaction
60 °C for 16 h and then concentrated in vacuo. The resulting mixture was diluted with CH2Cl2 (30 mL) and then washed
residue was diluted with ethyl acetate (50 mL), washed with with 1 N HCl (20 mL), aqueous saturated NaHCO3 (20 mL) and
H2O (30 mL), dried (MgSO4), and concentrated in vacuo. The then H2O (20 mL). The organic phase was dried (MgSO4) and
residue was purified by column chromatography on silica gel concentrated in vacuo. The residue was purified by column
(ethyl acetate–hexane gradient elution) to afford the title com- chromatography on silica gel (ethyl acetate–hexane gradient
pound as a colorless syrup (0.57 g, 92%). Analytical data for 9: elution) to afford the title compound as a colorless syrup
Rf = 0.5 (ethyl acetate–hexane, 2/8, v/v); [α]2D2 = +63.2 (c 1.0, (0.37 g, 87%). Analytical data for 12: Rf = 0.45 (ethyl acetate–
CHCl3); 1H NMR (300 MHz, CDCl3): δ, 7.84 (m, 3H, ArH), hexane, 3/7, v/v); [α]D22 = +33.6 (c 1.0, CHCl3); 1H NMR
7.56–7.47 (m, 4H, ArH), 5.02–4.90 (m, 3H, J1,2 = 3.90 Hz, H-1, (300 MHz, CDCl3): δ, 8.11–8.04 (m, 4H, ArH), 7.95 (d, 1H, J =
CH2Nap), 4.39 (dd, 1H, J4,5 = 9.30 Hz, H-4), 4.30 (dd, 1H, J3,4
4.2 Hz, H-3), 3.97 (dd, 1H, J2,3 = 5.4 Hz, H-2), 3.92 (dd, 1H, J = 5.64 (dd, 1H, J3,4 = 5.7 Hz, H-3), 5.02 (d, 1H, J1,2 = 4.2 Hz, H-1),
6.3 Hz, H-5b), 3.78 (m, 1H, 1/2CH2-linker), 3.64 (m, dd, J = 4.89–4.78 (dd, 2H, J = 12.0 Hz, CH2Nap), 4.74 (dd, 1H, J5a,5b
=
8.4 Hz, ArH), 7.80–7.72 (m, 4H, ArH), 7.59–7.36 (m, 8H, ArH),
=
5.1 Hz, H-5a), 3.54 (m, 1H, 1/2CH2-linker), 3.40 (t, 2H, J = 11.7 Hz, H-5b), 4.57 (dd, 1H, J4,5a = 7.5 Hz, H-5a), 4.36 (dd, 1H,
5.7 Hz, CH2-linker), 1.88 (m, 2H, CH2-linker) ppm; 13C NMR J2,3 = 6.9 Hz, H-2), 4.28 (m, 1H, H-4), 3.88 (m, 1H, 1/2CH2-
(75 MHz, CDCl3): δ, 135.50, 133.44, 133.30, 128.36, 128.10, linker), 3.51–3.31 (m, 3H, 1/2CH2-linker, CH2-linker), 1.85 (m,
127.94, 127.22, 126.35, 126.26, 126.17, 100.77, 80.73, 79.00, 2H, CH2-linker) ppm; 13C NMR (75 MHz, CDCl3): δ, 166.49,
73.77, 72.19, 68.68, 65.79, 48.57, 29.41, 27.7, 27.42, 22.83, 166.21, 135.07, 133.99, 133.68, 133.41, 133.39, 133.27, 130.43,
20.33 ppm; MALDI HR-MS calc. for C27H39N3NaO5Si 130.16, 130.04, 129.50, 128.71, 128.64, 128.56, 128.11, 127.98,
[M + Na]+: 536.2557, found 536.2553.
127.04, 126.50, 126.35, 125.93, 101.05, 81.92, 79.59, 77.93,
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-β-L-arabinofurano- 72.90, 66.4, 65.26, 48.50, 29.13 ppm; MALDI HR-MS calc. for
side (10). Compound 9 (0.25 g, 0.48 mmol) was dissolved in a C33H31N3NaO7 [M + Na]+: 604.2060, found 604.2057.
mixture of CH2Cl2 and MeOH (4 : 1, 5 mL) and freshly crystal-
3-Azidopropyl
3,5-di-O-benzyl-2-O-(2-naphthyl)methyl-β-
lized DDQ (0.22 g, 0.97 mmol) was added. The reaction arabinofuranoside (13). Hydrogen fluoride–pyridine (250 µL,
mixture was stirred at room temperature for 16 h. The mixture 9.8 mmol) was carefully diluted with pyridine (2.0 mL) at 0 °C.
was diluted with CH2Cl2 (20 mL), and the organic phase was The resulting solution was added slowly to a stirred at 0 °C
washed with saturated NaHCO3 (10 mL), dried (MgSO4) and suspension of 9 (1.25 g, 2.43 mmol) in CH2Cl2 (12 mL) and the
concentrated in vacuo. The residue was purified by column reaction was allowed to proceed for 2 h at 0 °C. The reaction
chromatography on silica gel (ethyl acetate–hexane gradient mixture was diluted with CH2Cl2 (30 mL), washed with water
elution) to afford the title compound as a colorless syrup (20 mL) and NaHCO3 (20 mL), dried (MgSO4) and concentrated
(0.15 g, 84%). Analytical data for 10: Rf = 0.40 (ethyl acetate– in vacuo. The crude product was purified by flash chromato-
hexane, 4/6, v/v); [α]2D2 = +25.9 (c 1.0, CHCl3); 1H NMR graphy on silica gel (EtOAc–hexane, 1/1, v/v) to give the diol
(300 MHz, CDCl3): δ, 4.54 (d, 1H, J1,2 = 6.9 Hz, H-1), 4.14 (dd, intermediate 11 (0.73 g, 81%). MALDI HR-MS calc. for
J2,3 = 4.2 Hz and J3,4 = 9.6 Hz, H-3), 4.01 (dd, 1H, J = 1.8 Hz and C19H23N3NaO5 [M + Na]+: 396.1535, found 396.1503. The diol
9.0 Hz, H-5b), 3.90–3.66 (m, 5H, H-2, 4, 5a, CH2-linker), (0.73 g, 1.95 mmol) and benzyl bromide (0.70 mL, 5.86 mmol)
3.48–3.43 (m, 5H, OCH3, CH2-linker), 2.62–2.45 (dd, 1H, J = were dissolved in DMF (10 mL), cooled to 0 °C, and treated
4.8 Hz, –OH), 1.89 (m, 2H, CH2-linker), 1.03 (s, 9H, t-Bu), 0.99 with NaH (60% dispersion in oil, 0.23 g, 5.86 mmol, four por-
(s, 9H, t-Bu) ppm; 13C NMR (75 MHz, CDCl3): δ, 103.55, 77.06, tions). The reaction mixture was stirred at 0 °C for 2 h, then
70.20, 68.83, 66.08, 62.87, 55.93, 48.68, 29.46, 27.66, 27.31, poured into ice-water (30 mL), stirred for 10 min, and extracted
22.91, 20.42 ppm; MALDI HR-MS calc. for C16H31N3NaO5Si with ethyl acetate–diethyl ether (1/1, v/v, 3 × 50 mL). The com-
[M + Na]+: 396.1931, found 396.1910.
3-Azidopropyl 3,5-di-O-benzoyl-2-O-(2-naphthyl)methyl- separated, dried (MgSO4) and concentrated in vacuo. The
bined organic phase was washed with cold water (100 mL),
β-arabinofuranoside (12). Hydrogen fluoride–pyridine (90 µL, residue was purified by column chromatography on silica gel
3.50 mmol) was carefully diluted with pyridine (0.5 mL) at (ethyl acetate–hexane gradient elution) to afford 13 as a color-
0 °C. The resulting solution was added slowly to a stirred at less syrup (0.81 g, 74%). Analytical data for 7: Rf = 0.50 (ethyl
1
0 °C suspension of 9 (0.45 g, 0.87 mmol) in CH2Cl2 (5 mL) and acetate–hexane, 3/7, v/v); [α]2D2 = +24.2 (c 1.0, CHCl3); H NMR
the reaction was allowed to proceed for 2 h at 0 °C. The reac- (300 MHz, CDCl3): δ, 7.86 (m, 4H, ArH), 7.53 (m, 3H, ArH),
tion mixture was diluted with CH2Cl2 (20 mL), washed with 7.35 (m, 10H, ArH), 4.92 (d, 1H, J1,2 = 3.3 Hz, H-1), 4.80 dd,
water (10 mL), NaHCO3 (10 mL), dried (MgSO4) and concen- 2H, J = 12.6 Hz, CH2Ph), 4.76 (dd, 2H, J = 11.7 Hz, CH2Nap),
trated in vacuo. The crude product was purified by flash 4.59 dd, 2H, J = 12.6 Hz, CH2Ph), 4.17 (m, 3H, H-2, H5a, H5b),
chromatography on silica gel (EtOAc–hexane, 1/1, v/v) to give 3.77 (m, 1H, 1/2CH2-linker), 3.58 (m, 2H, H-3, H-4), 3.44–3.26
the diol intermediate 11 (0.27 g, 81%). MALDI HR-MS calc. for (m, 3H, 1/2CH2-linker, CH2-linker), 1.84 (m, 2H, CH2-linker)
C19H23N3NaO5 [M + Na]+: 396.1535, found 396.1503. The ppm; 13C NMR (75 MHz, CDCl3): δ, 138.47, 138.20, 133.44,
resulting intermediate (0.27 g, 0.72 mmol) was dissolved in 133.50, 128.66, 128.65, 128.52, 128.18, 128.14, 128.00, 127.93,
dry pyridine (10 mL), cooled to 0 °C, and benzoyl chloride 127.12, 126.50, 126.34, 126.19, 100.97, 84.54, 83.39, 80.48,
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73.59, 72.90, 72.65, 72.62, 64.76, 48.58, 29.20 ppm; MALDI 18H), 0.91 (s, 18H) ppm; 13C NMR (300 MHz, CDCl3): δ,
HR-MS calc. C33H35N3NaO5 for [M + Na]+: 576.2475, found 136.27, 135.70, 133.45, 133.26, 133.11, 128.37, 128.13, 128.06,
576.2480.
127.97, 127.92, 127.91, 126.94, 126.38, 126.33, 126.25, 126.15,
3-Azidopropyl 3,5-di-O-benzoyl-β-L-arabinofuranoside (14). 126.04, 125.96, 104.70, 101.39, 101.37, 81.34, 80.61, 79.05,
Compound 12 (0.35 g, 0.60 mmol) was dissolved in a mixture 78.71, 78.48, 75.24, 76.70, 75.24, 74.02, 73.64, 71.86, 71.49,
of CH2Cl2 and MeOH (4 : 1, v/v, 8 mL) and freshly crystallized 68.66, 66.14, 65.16, 55.39, 48.65, 34.89, 31.14, 29.51, 27.28,
DDQ (0.27 g, 1.2 mmol) was added. The reaction mixture was 27.74, 27.71, 27.64, 27.42, 27.28, 27.25, 27.22, 22.87, 22.76,
stirred at room temperature for 5 h. The mixture was diluted 22.74, 22.64, 21.32, 20.35, 20.33, 20.20 ppm; MALDI HR-MS
with CH2Cl2 (20 mL), and the organic phase was washed with calc. for C40H63N3NaO9Si2 [M
saturated NaHCO3 (15 mL), dried (MgSO4) and concentrated 808.3993.
+
Na]+: 808.4001, found
in vacuo. The residue was purified by column chromatography
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)-
on silica gel (ethyl acetate–hexane gradient elution) to afford methyl-β-L-arabinofuranosyl-(1→2)-3,5-di-O-benzoyl-β-L-arabino-
the title compound as a colorless syrup (0.22 g, 81%). Analyti- furanoside (17). Analytical data for 17: Rf = 0.50 (ethyl acetate–
cal data for 14: Rf = 0.44 (ethyl acetate–hexane, 4/6, v/v); [α]D22
=
hexane, 3/7, v/v); [α]2D2 = +58.2 (c 1.0, CHCl3); 1H NMR
+51.9 (c 1.0, CHCl3); 1H NMR (300 MHz, CDCl3): δ, 8.05 (m, (300 MHz, CDCl3): δ, 8.05–7.99 (m, 3H, ArH), 7.84–7.80 (m, 4H,
4H, ArH), 7.62–7.52 (m, 3H, ArH), 7.47–7.37 (m, 3H, ArH), 5.45 ArH), 7.56–7.25 (m, 10H, ArH), 5.71 (dd, 1H, J3,4 = 7.2 Hz, H-3),
(dd, J3,4 = 6.3 Hz, H-3), 5.09 (d, J1,2 = 4.8 Hz, H-1), 4.66 (dd, 1H, 5.08 (d, 1H, J1,2 = 4.5 Hz, H-1), 5.04 (d, 1H, J1′,2′ = 5.1 Hz, H-1′),
J = 4.5 Hz and 11.4 Hz, H-5b), 4.57–4.45 (m, 2H, H-2, H-5a), 4.94 (dd, 2H, J = 12.0 Hz, CH2Ph), 4.71 (dd, 1H, J4,5b = 4.20 Hz
4.18 (m, 1H, H-4), 3.93 (m, 1H, 1/2CH2-linker), 3.60 (m, 1H, and 11.7 Hz, H-5b), 4.57 (dd, 1H, J4,5a = 7.5 Hz and J5a,5b
=
1/2CH2-linker), 3.37 (m, 1H, CH2-linker), 2.90 (d, 1H, J = 11.7 Hz, H-5a), 4.49 (dd, 1H, J2,3 = 7.2 Hz, H-2), 4.43 (dd, 1H,
8.7 Hz, –OH), 1.85 (dd, 2H, J = 6.6 Hz, CH2-linker) ppm; J3′,4′ = 9.3 Hz, H-3′), 4.34 (m, 1H, H-4′), 4.08 (dd, 1H, J = 5.1 Hz,
13C NMR (75 MHz, CDCl3): δ, 166.65, 166.35, 133.75, 133.35, 1/2CH2-linker), 3.98 (dd, 1H, J2′,3′ = 9.0 Hz), 3.86 (m, 1H, H-4′),
130.06, 130.01, 129.93, 129.36, 128.69, 128.58, 101.87, 79.92, 3.76 (dd, 1H, J5a′,5b′ = 9.0 Hz, H-5b′), 3.60 (m, 1H, 1/2CH2-
79.45, 76.77, 65.98, 65.79, 48.55, 29.02 ppm; MALDI HR-MS linker), 3.49 (m, 1H, H-5a′), 3.35–3.19 (m, 2H, CH2-linker), 1.74
calc. for C22H23N3NaO7 [M + Na]+: 464.1434, found 464.1437.
(m, 2H, CH2-linker), 1.02 (s, 9H, t-Bu), 0.96 (s, 9H, t-Bu) ppm;
3-Azidopropyl 3,5-di-O-benzyl-β-L-arabinofuranoside (15). 13C NMR (75 MHz, CDCl3): δ, 166.40, 165.97, 135.51, 133.53,
Compound 13 (0.78 g, 1.40 mmol) was dissolved in a mixture 133.43, 133.28, 133.19, 130.14, 129.98, 129.93, 129.70, 128.65,
of CH2Cl2 and MeOH (4 : 1, v/v, 20 mL) and freshly crystallized 128.51, 128.33, 128.08, 127.93, 127.27, 126.35, 126.16, 101.03,
DDQ (0.64 g, 2.80 mmol) was added. The reaction mixture was 99.97, 80.53, 80.52, 79.39, 78.49, 77.34, 74.27, 71.90, 68.48,
stirred at room temperature for 3 h. The mixture was diluted 66.34, 65.24, 48.20, 29.10, 27.71, 27.36, 22.76, 20.76 ppm;
with CH2Cl2 (50 mL), and the organic phase was washed with MALDI HR-MS calc. for C46H55N3NaO11Si [M + Na]+: 876.3504,
saturated NaHCO3 (50 mL), dried (MgSO4) and concentrated found 876.3489.
in vacuo. The residue was purified by column chromatography
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)-
on silica gel (ethyl acetate–hexane gradient elution) to afford methyl-β-L-arabinofuranosyl-(1→2)-3,5-di-O-benzyl-β-L-arabino-
the title compound as a colorless syrup (0.45 g, 77%). Analyti- furanoside (18). Analytical data for 18: Rf = 0.47 (ethyl acetate–
cal data for 15: Rf = 0.44 (ethyl acetate–hexane, 4/6, v/v); [α]D22
=
hexane, 2.5/7.5, v/v); [α]2D2 = +89.7 (c 1.0, CHCl3); 1H NMR
+52.8 (c 1.0, CHCl3); 1H NMR (300 MHz, CDCl3): δ, 7.32 (m, (300 MHz, CDCl3): δ, 7.84–7.76 (m, 4H, ArH), 7.54–7.45 (m, 4H,
10H, ArH), 4.94 (d, 1H, J1,2 = 4.8 Hz, H-1), 4.76 (d, 1H, J = ArH), 7.30–7.24 (m, 10H, ArH), 5.07 (d, 1H, J1′,2′ = 5.1 Hz, H-1),
12.0 Hz, 1/2CH2Nap), 4.61 (d, 1H, J = 12 Hz, 1/2CH2Nap), 5.54 5.00 (m, 2H, J1,2 = 4.5 Hz and J = 12.0 Hz, H-1, 1/2CH2Ph), 4.89
(dd, 2H, J = 12.0 Hz, CH2Nap), 4.24 (m, 1H, J2,3 = 5.7 Hz, H-2), (d, J = 12.0 Hz, 1/2CH2Ph), 4.75 (d, J = 12.0 Hz, 1/2CH2Ph), 4.60
4.12 (dd, 1H, J4,5a = 11.1 Hz, H-4), 3.83 (m, 2H, H-3, 1/2CH2- (d, J = 12.0 Hz, 1/2CH2Ph), 4.51–4.45 (m, 3H, H-3′, CH2Ph),
linker), 3.55–3.48 (m, 3H, H-5a, 5b, 1/2CH2-linker), 3.28 (m, 4.39 (dd, 1H, J2,3 = 6.6 Hz, H-2), 4.30 (ds, 1H, J3,4 = 9.0 Hz,
2H, CH2-linker), 2.57 (d, J = 9.6 Hz, –OH), 1.80 (m, 2H, CH2- H-3), 4.18–4.11 (m, 2H, H-5a′, H-5b′), 4.01 (dd, 1H, J2′,3′
=
linker) ppm; 13C NMR (75 MHz, CDCl3): δ, 138.18, 128.61, 9.0 Hz, H-2′), 3.89 (dd, 1H, J5a,5b = 10.5 Hz, H-5b), 3.7–3.65 (m,
128.01, 127.96, 127.93, 101.95, 84.62, 80.95, 78.09, 73.53, 2H, H-4′, 1/2CH2-linker), 3.50 (m, 2H, H-4, H-5a), 3.39 (m, 1H,
72.14, 71.97, 65.50, 48.61, 29.11 ppm; MALDI HR-MS calc. for 1/2CH2-linker), 3.19 (m, 2H, CH2-linker), 1.68 (m, 2H, CH2-
C22H27NaN3O5 [M + Na]+: 436.1849, found 436.1833.
linker), 1.06 (s, 9H, t-Bu), 0.99 (s, 9H, t-Bu) ppm; 13C NMR
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)- (75 MHz, CDCl3): δ, 138.25, 135.55, 133.43, 133.28, 128.63,
methyl-α/β-L-arabinofuranosyl-(1→2)-3,5-O-(di-tert-butylsilane- 128.57, 128.32, 128.07, 128.03, 127.94, 127.90, 127.88, 127.23,
diyl)-β-L-arabinofuranoside (16). Analytical data for 16 126.34, 126.31, 126.15, 100.52, 99.14, 82.20, 81.08, 80.44,
(500 MHz, CDCl3): Rf = 0.52 (ethyl acetate–hexane, 2/8, v/v); α/β 80.36, 78.68, 74.31, 73.50, 72.50, 72.34, 71.66, 68.73, 64.52,
1H NMR: δ, 7.81–7.71 (m, 4H, ArH), 7.48–7.38 (m, 3H, ArH), 48.32, 29.15, 27.72, 27.39, 22.85, 20.34 ppm; MALDI HR-MS
5.33 (d, 1H, J = 3.0 Hz), 5.05 (d, 1H, J = 3.0 Hz), 4.94–4.79 (m, calc. for C46H59N3NaO9Si [M + Na]+: 848.3919, found 848.3929.
4H, 2 × CH2Ph), 4.55 (d, 1H, J = 4.5 Hz), 4.31–4.15 (m, 3H),
3-Azidopropyl
3,5-O-di-benzyl-2-O-(2-naphthyl)methyl-β-L-
4.07 (m, 1H), 3.92 (m, 2H), 3.82–3.70 (m, 3H), 3.59–3.48 (m, arabinofuranosyl-(1→2)-3,5-di-O-benzyl-β-L-arabinofuranoside
2H), 3.36–3.30 (m, 2H), 3.17 (m, 2H), 1.68 (m, 2H), 1.01 (s, (19). Analytical data for 19: Rf = 0.42 (ethyl acetate–hexane,
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3/7, v/v); [α]2D2 = +83.8 (c 1.0, CHCl3); 1H NMR (300 MHz, quenched by the addition of Et3N or pyridine. The suspension
CDCl3): δ, 7.82–7.75 (m, 4H, ArH), 7.51–7.46 (m, 3H, ArH), was diluted with CH2Cl2 (20 mL) and filtered through a pad of
7.29–7.19 (m, 20H, ArH), 5.17 (d, 1H, J1′,2′ = 3.0 Hz, H-1′), 5.07 Celite, and the filtrate was washed successively with 10%
(d, 1H, J1,2 = 4.2 Hz, H-1), 4.91 (d, 1H, J = 11.4 Hz, 1/2CH2Ph), Na2S2SO3 (20 mL) and brine (20 mL). The organic layer was
4.71–4.57 (m, 5H, 2 × CH2Ph, 1/2CH2Ph), 4.51 (dd, 2H, J = dried (MgSO4) and concentrated in vacuo. The residue was pur-
12.0 Hz, CH2Ph), 4.46 (dd, 1H, J2,3 = 5.7 Hz, H-2), 4.42 (d, 1H, ified by column chromatography on silica gel (ethyl acetate–
J = 12.0 Hz, 1/2CH2Ph), 4.31 (d, 1H, J = 12.0 Hz, 1/2CH2Ph), hexane gradient elution) to give the trisaccharide 22 (432 mg,
4.18–4.13 (m, 6H, H-2′, H-5a′, H-5b′, H-3, H-5a, H-5b), 3.75 (m, 69%). Analytical data for 22: Rf = 0.42 (ethyl acetate–hexane,
1H, 1/2CH2-linker), 3.61–3.39 (m, 5H, H-3′, H-4′, H-3, CH2- 3/7, v/v); [α]2D3 = +84.2° (c 1.0, CHCl3); 1H NMR (500 MHz,
linker), 3.15 (m, 2H, CH2-linker), 1.68 (m, 2H, CH2-linker) CDCl3): δ, 7.83 (s, 1H, ArH), 7.73 (d, 1H, J = 8.5 Hz, ArH), 7.65
ppm; 13C NMR (75 MHz, CDCl3): δ, 138.34, 138.30, 138.24, (d, 2H, J = 8.0 Hz, ArH), 7.51 (dd, 1H, J = 8.5 Hz, ArH),
135.57, 133.47, 133.28, 128.58, 128.55, 128.53, 128.39, 128.10, 7.42–7.38 (m, 2H, ArH), 7.33–7.22 (m, 14H, ArH), 7.15–7.11 (m,
128.07, 128.00, 127.98, 127.94, 127.90, 127.86, 127.83, 127.76, 4H, ArH), 6.88 (dd, 2H, J = 6.5 Hz, ArH), 5.38 (d, 1H, J =
126.88, 126.83, 126.19, 126.13, 100.36, 98.77, 84.25, 83.18, 4.5 Hz), 5.12 (d, 1H, J = 4.5 Hz), 5.10 (d, 1H, J = 11.5 Hz), 5.04
82.50, 81.02, 80.57, 79.37, 73.52, 73.32, 72.64, 72.52, 72.46, (d, 1H, J = 4.5 Hz), 4.78 (d, 1H, J = 12.0 Hz), 4.70 (dd, 2H, J =
72.01, 64.56, 48.36, 29.28 ppm; MALDI HR-MS calc. for 11.5 Hz), 4.67 (d, 1H, J = 12.0 Hz), 4.52–4.49 (m, 2H), 4.46 (d,
C52H55N3NaO9 [M + Na]+: 888.3836, found 888.3821.
3-Azidopropyl 3,5-di-O-benzyl-β-L-arabinofuranosyl-(1→2)- 9.0 Hz), 4.31 (d, 1H, J = 12.5 Hz), 4.18 (m, 3H), 4.09 (d, 1H, J =
1H, J = 10 Hz), 4.41 (m, 1H), 4.35 (dd, 1H, J = 5.0 Hz and
3,5-di-O-benzyl-β-L-arabinofuranoside (21). Compound 19 11.5 Hz), 4.05–4.00 (m, 3H), 3.90 (dd, 1H, J = 10.5 Hz),
(530 mg, 0.61 mmol) was dissolved in a mixture of DCM and 3.82–3.74 (m, 3H), 3.40 (m, 2H), 3.44–3.35 (m, 3H), 3.20 (m,
MeOH (4 : 1, v/v, 15 mL) and freshly crystallized DDQ (278 mg, 2H), 1.75 (m, 2H, CH2-linker), 1.05 (s, 9H), 1.00 (s, 9H) ppm;
1.22 mmol) was added. The reaction mixture was stirred at 13C NMR(75 MHz, CDCl3): δ, 138.35, 138.17, 138.15, 138.03,
room temperature for 4 h. The mixture was diluted with 136.04, 133.47, 133.22, 128.63, 128.49, 128.35, 128.21, 128.08,
CH2Cl2 (30 mL), and the organic phase was washed with satu- 127.98, 127.93, 127.77, 127.70, 127.64, 127.51, 126.65, 126.30,
rated NaHCO3 (20 mL), dried (MgSO4) and concentrated 126.24, 126.04, 100.09, 98.19, 97.69, 83.12, 81.85, 81.19, 80.95,
in vacuo. The residue was purified by column chromatography 80.41, 79.88, 79.12, 78.21, 74.61, 73.50, 73.22, 72.71, 72.56,
on silica gel (ethyl acetate–hexane gradient elution) to give the 72.40, 71.91, 71.43, 69.03, 64.81, 48.42, 29.36, 27.73, 27.40,
alcohol as a colorless syrup (315 mg, 71%). Analytical data for 22.92, 20.35 ppm; MALDI HR-MS calc. for C65H79N3NaO13Si
21: Rf = 0.47 (ethyl acetate–hexane, 4/6, v/v); [α]2D2 = +86.4 (c 1.0, [M + Na]+: 1160.5280, found 1160.5261.
CHCl3); 1H NMR (300 MHz, CDCl3): δ, 7.32–7.22 (m, 20H,
ArH), 5.00 (d, 1H, J1′,2′ = 4.8 Hz, H-1′), 4.97 (d, 1H, J1,2 = 4.2 Hz, furanosyl-(1→2)-3,5-di-O-benzyl-2-O-naphthyl-β-L-arabino-
H-1), 4.78 (d, 1H, J = 12.0 Hz, 1/2CH2Ph), 4.64 (dd, 2H, J = furanosyl-(1→2)-3,5-di-O-benzyl-β-L-arabinofuranoside (24).
3-Azidopropyl 5-O-benzoyl-2-O-(2-naphthyl)methyl-β-L-arabino-
11.7 Hz, CH2Ph), 4.53 (dd, 2H, J = 12.3 Hz, CH2Ph), 4.98 (d, Hydrogen fluoride–pyridine (32 µL, 1.24 mmol) was carefully
3H, J = 12.0 Hz, 1/2CH2Ph, CH2Ph), 4.43–4.35 (m, 3H, H-2, diluted with pyridine (200 µL) at 0 °C. The resulting solution
CH2Ph), 4.24 (m, 1H, H-2′), 4.15–4.09 (dd, 2H, J = 5.7 Hz, H5a′, was added slowly to a stirred at 0 °C suspension of 22 (360 mg,
H-5b′), 3.87 (dd, 1H, J3′,4′ = 6.3 Hz, H-3′), 3.77 (m, 1H, H-4′), 0.31 mmol) in CH2Cl2 (5 mL) and the reaction was allowed to
3.56 (dd, 2H, J = 5.7 Hz, H-4, H-5b), 3.48 (dd, 2H, J = 6.0 Hz, proceed for 2 h at 0 °C and another 3 h at room temperature.
H-5a, 1/2CH2-linker), 3.41 (m, 1H, 1/2CH2-linker), 3.29 (m, 2H, The reaction mixture was diluted with CH2Cl2 (10 mL), washed
CH2-linker), 3.09 (d, 1H, J = 8.7 Hz, OH), 1.77 (m, 2H, CH2- with water (5 mL) and saturated NaHCO3 (5 mL), dried
linker) ppm; 13C NMR (75 MHz, CDCl3): δ, 138.21, 138.20, (MgSO4) and concentrated in vacuo. The crude product was
138.15, 126.61, 128.58, 128.54, 128.03, 127.96, 127.89, 127.82, purified by flash chromatography on silica gel (EtOAc–hexane,
102.46, 100.38, 84.31, 82.47, 82.42, 81.22, 80.49, 78.07, 73.55, 1/1, v/v) to give diol 23 (280 mg, 88%). MALDI HR-MS calc. for
73.40, 72.45, 72.30, 72.05, 72.02, 64.79, 48.42, 28.99 ppm; C57H63NaN3O13 [M + Na]+: 1020.4259, found 1020.4230. The
MALDI HR-MS calc. for C41H47N3NaO9 [M + Na]+: 748.3210, resulting diol (280 mg, 0.28 mmol) was dissolved in dry
found 748.3520.
CH2Cl2 (2 mL), cooled to 0 °C, and treated with benzoic acid
3-Azidopropyl 3,5-O-(di-tert-butylsilanediyl)-2-O-(2-naphthyl)- (0.44 g, 7.4 mmol) and 2-chloromethyl pyridinium iodide
methyl-β-L-arabinofuranosyl-(1→2)-3,5-di-O-benzyl-2-O-naphthyl- (CMPI) (2.36 g, 9.25 mmol). The mixture was stirred for
β-L-arabinofuranosyl-(1→2)-3,5-di-O-benzyl-β-L-arabinofurano- 15 minutes at room temperature followed by the addition of
side (22). A mixture of the glycosyl donor 3 (360 mg, 1,4 diazabicyclo[2,2,2]octane (DABCO) (0.980 g, 8.75 mmol).
0.68 mmol), glycosyl acceptor 21 (400 mg, 0.55 mmol), and Stirring was continued until TLC indicated consumption of
freshly activated 4 Å molecular sieves (1 g) in CH2Cl2 (15 mL) starting material (∼1.5 h). The reaction mixture was filtered
was stirred at room temperature for 30 min and then cooled to through Celite, diluted with EtOAc (40 mL), and washed with
−40 °C. NIS (235 mg, 1.03 mmol) followed by AgOTf (53 mg, brine (2 × 20 mL). The combined organic layers were dried
0.20 mmol) was added. The reaction mixture was stirred at (MgSO4) and concentrated in vacuo. The residue was purified
−40 °C for 2 h and then warmed slowly to room temperature by column chromatography on silica gel (ethyl acetate–hexane
and stirring was continued for 30 min. The reaction was gradient elution) to give the trisaccharide acceptor 24 (161 mg,
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52%). Analytical data for 22: Rf = 0.54 (ethyl acetate–hexane, organic layer was dried (MgSO4) and concentrated in vacuo.
1/1, v/v); [α]2D3 = +78.1° (c 1.0, CHCl3); 1H NMR (500 MHz, The residue was purified by column chromatography on silica
CDCl3): δ, 8.06 (d, 2H, J = 7.5 Hz), 8.86 (s, 1H, ArH), 7.73 (d, gel (ethyl acetate–hexane gradient elution) to give the tetra-
1H, J = 8.0 Hz, ArH), 7.52 (m, 2H, ArH), 7.45–7.39 (m, 4H, ArH), saccharide 26 (183 mg, 87%). Analytical data for 26: Rf = 0.45
7.29–7.09 (m, 18H, ArH), 6.85 (d, 2H, J = 7.0 Hz, ArH), 5.51 (d, (ethyl acetate–hexane, 4/6, v/v); [α]2D4 = +55.5° (c 1.0, CHCl3); 1H
1H, J = 4.5 Hz), 5.16 (d, 1H, J = 4.0 Hz), 5.07 (d, 1H, J = 11.0 Hz), NMR (500 MHz, CDCl3): δ, 8.03 (d, 2H, J = 7.5 Hz, ArH), 7.96
5.05 (d, 1H, J = 4.0 Hz), 4.65 (dd, 1H, J = 12.0 Hz), 4.61–4.56 (m, (dd, 4H, J = 7.5 Hz, ArH), 7.76 (s, 1H, ArH), 7.66 (d, 1H, J =
3H, J = 4.5 Hz), 4.49 (m, 2H, J = 7.0 Hz), 4.44–4.36 (m, 3H), 4.30 8.0 Hz, ArH), 7.55 (m, 4H, ArH), 7.48–7.10 (m, 31H, ArH), 6.90
(d, 1H, J = 12.0 Hz), 4.25 (dd, 1H, J = 5.0 Hz), 4.18 (m, 3H), (d, 2H, J = 7.0 Hz, ArH), 5.68 (s, 1H), 5.57 (s, 1H), 5.56 (d, 1H,
4.07–3.98 (m, 4H), 3.80 (m, 2H), 3.49–3.37 (m, 4H), 3.31 (dd, J = 4.0 Hz), 5.52 (d, 1H, J = 4.0 Hz), 5.17 (d, 1H, J = 4.0 Hz),
1H, J = 5.0 Hz and 10.0 Hz), 3.26 (m, 2H), 1.79 (m, 2H) ppm; 5.07 (d, 1H, J = 4.0 Hz), 5.01 (dd, 1H, J = 11.5 Hz), 4.79 (dd, 1H,
13C NMR (75 MHz, CDCl3): δ, 166.61, 138.31, 138.11, 138.04, J = 4.5 Hz and 11 Hz), 4.72–4.39 (m, 14H), 4.30 (d, 1H, J =
137.99, 135.55, 133.47, 133.34, 133.31, 130.02, 128.67, 128.60, 12.5 Hz), 4.25 (dd, 1H, J = 4.0 Hz and 7.0 Hz), 4.20–412 (m,
128.58, 128.52, 128.48, 128.33, 128.17, 128.06, 127.94, 127.87, 4H), 4.05 (m, 2H), 3.85 (dd, 1H, J = 7.0 Hz), 3.81 (m, 1H), 3.50
127.75, 127.70, 127.61, 127.38, 126.92, 126.44, 126.24, 99.59, (dd, 1H, J = 4.0 Hz and 9.5 Hz), 3.44 (m, 3H), 3.37 (dd, 1H, J =
98.31, 98.17, 84.33, 83.17, 82.20, 81.04, 80.35, 80.10, 79.48, 5.0 Hz and 10.0 Hz), 3.26 (m, 2H), 1.78 (m, 2H) ppm; 13C NMR
79.19, 76.27, 73.45, 73.20, 72.62, 72.58, 72.29, 72.26, 72.01, (75 MHz, CDCl3): δ, 166.36, 166.26, 165.84, 165.57, 138.30,
67.01, 64.95, 48.51, 29.36 ppm; MALDI HR-MS calc. for 138.12, 138.05, 135.65, 133.72, 133.69, 133.41, 133.25, 133.19,
C64H67N3NaO14 [M + Na]+: 1124.4521, found 1124.4507.
130.19, 130.15, 130.11, 130.08, 130.00, 129.94, 129.89, 129.33,
Phenyl 2,3,5-tri-O-benzoyl-1-thio-α-L-arabinofuranoside (25). 129.23, 129.17, 128.72, 128.59, 128.53, 128.50, 128.48, 129.36,
To a solution of phenyl 1-thio-α-L-arabinofuranoside (0.65 g, 128.12, 128.30, 127.93, 127.88, 127.77, 127.73, 127.64, 127.42,
2.68 mmol) in pyridine (15 mL) at 0 °C was added dropwise 126.60, 126.30, 126.07, 105.75, 100.02, 98.42, 98.24, 84.15,
benzoyl chloride (1.25 mL, 10.75 mmol). The reaction mixture 83.21, 82.52, 82.37, 81.60, 81.14, 81.04, 80.37, 79.47, 79.40,
was stirred at room temperature for 2 h. The reaction mixture 79.23, 77.78, 77.56, 73.48, 73.23, 72.59, 72.53, 72.48, 72.34,
was then diluted with CH2Cl2 (50 mL) and then washed with 72.29, 71.88, 48.47, 29.34, 27.68 ppm; MALDI HR-MS calc. for
1 N HCl (30 mL), saturated NaHCO3 (30 mL) and then H2O C90H87N3NaO21 [M + Na]+: 1568.5730, found 1568.5741.
(30 mL). The organic phase was dried (MgSO4) and concen-
3-Aminopropyl α-L-arabinofuranosyl-(1→3)-β-L-arabinofura-
trated in vacuo. The residue was purified by column chromato- nosyl-(1→2)-β-L-arabinofuranosyl-(1→2)-β-L-arabinofuranoside
graphy on silica gel (ethyl acetate–hexane gradient elution) to (27). To a solution of 26 (100 mg, 0.103 mmol) in methanol
afford the title compound as a colorless syrup (1.26 g, 85%). (1.0 mL) was added 1 M NaOMe until pH ∼ 10 (∼0.1 mL) was
Analytical data for 25: Rf = 0.62 (ethyl acetate–hexane, 4/6, v/v); achieved. The reaction mixture was stirred for 4 h at room
[α]D22 = −52.8 (c 1.0, CHCl3); 1H NMR (300 MHz, CDCl3): δ, 8.15 temperature and then neutralized with Dowex (H+), filtered,
(d, J = 7.2 Hz, 2H, ArH), 8.02 (dd, J = 7.2 Hz, 3H, ArH), and concentrated in vacuo. The crude residue was dissolved in
7.64–7.25 (m, 15H, ArH), 5.84 (s, 1H, H-1), 5.72 (dd, 1H, J = 1.5 MeOH–H2O–HOAc (5 mL, 3/1/1, v/v/v), and 10% Pd–C (20 mg)
Hz, H-2), 5.66 (dd, 1H, J3,4 = 4.5 Hz, H-3), 4.86 (m, 1H, J4,5 = 8.7 was added. The reaction mixture was stirred under an atmos-
Hz, H-4), 4.76–4.83 (m, 2H, J = 8.5 Hz, H-5a, 5b) ppm; 13C phere of H2 for 15 h. The catalyst was then filtered-off, washed
NMR (75 MHz, CDCl3): δ, 166.38, 165.80, 165.57, 133.90, with methanol and H2O, and the combined filtrates were con-
133.83, 133.67, 133.31, 132.53, 130.27, 130.13, 129.99, 129.90, centrated under reduced pressure. The residue was passed
129.30, 129.18, 129.09, 128.80, 128.77, 128.55, 128.08, 91.66, through a small amount of latrobeads and slowly eluted with
82.78, 81.38, 78.27, 63.74 ppm; MALDI HR-MS calc. for H2O. Fractions containing the product were collected and con-
C32H26NaO7S [M + Na]+: 577.1297, found 577.1301.
3-Azidopropyl 2,3,5-tri-O-benzoyl-α-L-arabinofuranosyl-(1→3)- ide 25 (33.2 mg, 85% for 2 steps). Analytical data for 27: [α]2D6
centrated in vacuo to afford the fully deprotected tetrasacchar-
=
5-O-benzoyl-2-O-naphthyl-β-L-arabinofuranosyl-(1→2)-3,5-O-di- +58.2° (c 1.0, H2O); 1H NMR (500 MHz, D2O): δ, 5.15 (d, 1H, J =
benzyl-2-O-(2-naphthyl)methyl-β-L-arabinofuranosyl-(1→2)-3,5- 4.5 Hz), 5.87 (s, 1H), 5.04 (d, 1H, J = 4.5 Hz), 4.99 (d, 1H, J =
di-O-benzyl-β-L-arabinofuranoside (26). A mixture of glycosyl 4.5 Hz), 4.22 (m, 2H, J = 4.5 Hz), 4.15 (dd, 1H, J = 4.5 Hz and
donor 25 (114 mg, 0.20 mmol), glycosyl acceptor 24 (150 mg, 8.0 Hz), 4.10 (dd, 1H, J = 7.5 Hz), 4.08–4.05 (m, 2H, J = 7.0 Hz),
0.14 mmol), and freshly activated 4 Å molecular sieves 4.02 (m, 1H), 3.95–3.90 (m, 2H), 3.85 (m, 1H), 3.83–3.77 (m,
(500 mg) in CH2Cl2 (7.5 mL) was stirred at room temperature 5H), 3.73–3.67 (m, 3H), 3.65–3.55 (m, 5H), 3.54–3.40 (m, 2H),
for 30 min. The mixture was cooled to −20 °C, and NIS (70 mg, 3.00 (m, 2H), 1.84 (m, 1H) ppm; 13C NMR (75 MHz, D2O): δ,
0.31 mmol) and AgOTf (16 mg, 0.06 mmol) were added. The 108.09, 99.90, 97.72, 99.98, 84.44, 81.88, 81.68, 81.57, 81.38,
reaction mixture was stirred at −20 °C for 2 h and then 80.64, 80.44, 76.78, 76.22, 72.96, 72.59, 66.27, 62.75, 62.72,
warmed slowly to room temperature and stirring was contin- 62.66, 61.42, 38.29, 26.59 ppm; MALDI HR-MS calc. for
ued for 1 h. The reaction was quenched by the addition of pyri- C23H41NNaO17 [M + Na]+: 626.2272, found 626.2265.
dine. The suspension was diluted with CH2Cl2 (15 mL) and
N-Biotinyl-3-aminopropyl
α-L-arabinofuranosyl-(1→3)-β-L-
filtered through a pad of Celite, the filtrate was washed succes- arabinofuranosyl-(1→2)-β-L-arabinofuranosyl-(1→2)-β-L-arabino-
sively with 10% Na2S2SO3 (10 mL) and brine (10 mL). The furanoside (29). Compound 27 (5 mg, 8.2 µmol) was dissolved
5144 | Org. Biomol. Chem., 2013, 11, 5136–5146
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in PBS buffer pH 7.4 (0.5 mL) and NHS-Biotin (2.8 mg, 8.2
µmol) in PBS buffer pH 7.4 (0.2 mL) was added. The resulting
mixture was stirred for 2 h at room temperature. The mixture
was directly transferred to a reverse phase C-18 column and
purified by eluting with 0–10% MeOH–H2O. The appropriate
fractions were collected and concentrated in vacuo to afford 29
References
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1
(5.6 mg, 81%) as a white solid. Analytical data for 29: H NMR
(500 MHz, D2O): δ, 5.14 (d, 1H, J = 4.0 Hz), 5.09 (s, 1H), 5.07
(d, 1H, J = 5.0 Hz), 5.05 (d, 1H, J = 4.5 Hz), 4.50 (m, 1H), 4.32
(dd, 1H, J = 4.5 Hz), 4.24–4.16 (m, 4H), 4.08–4.03 (m, 3H),
3.95–3.92 (m, 2H), 3.87 (m, 1H), 3.84–3.79 (m, 3H), 3.73–3.57
(m, 8H), 3.51 (dd, 1H, J = 7.0 Hz and 12.0 Hz), 3.43 (m, 1H),
3.26–3.18 (m, 3H), 3.10 (dd, 1H, J = 6.5 Hz), 2.89 (dd, 1H, J =
5.0 Hz), 2.68 (dd, 1H, J = 7.0 Hz), 2.16 (dd, 1H, J = 7.0 Hz), 2.10
(dd, 1H, J = 7.5 Hz), 1.71 (dd, 1H, J = 7.0 Hz), 1.68–1.44 (m,
5H), 1.30 (m, 2H) ppm; 13C NMR (HSQC, 125 MHz, D2O): δ,
108.96, 99.83, 98.15, 99.50 (four anomeric carbons) ppm;
MALDI HR-MS calc. for C33H55N3NaO20S [M + Na]+: 868.2998,
found 868.4810.
N-Biotinyl-3-aminopropyl
β-L-arabinofuranosyl-(1→2)-β-L-
arabinofuranosyl-(1→2)-β-L-arabinofuranoside (30). To a solution
of 23 (70 mg, 70.2 µmol) in MeOH–H2O–HOAc (3 mL/1 mL/
1 mL) was added 10% Pd–C (14 mg). The reaction mixture was
stirred under an atmosphere of H2 for 15 h. The catalyst was 10 Y. Rao, T. Buskas, A. Albert, M. A. O’Neill, M. G. Hahn and
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lected and concentrated in vacuo to afford the fully deprotected
trisaccharide 28 (23.5 mg, 79%). MALDI HR-MS calc. for 14 J. B. Houseknecht, T. L. Lowary and C. M. Hadad, J. Phys.
C18H33NNaO13 [M + Na]+: 494.1850, found 494.1854. The fully
Chem. A, 2003, 107, 5763–5777.
deprotected trisaccharide (5 mg, 10.6 µmol) was dissolved in 15 A. Imamura and T. L. Lowary, Trends Glycosci. Glycotechnol.,
PBS buffer pH 7.4 (0.5 mL), and NHS-Biotin (3.6 mg, 10.6 2011, 23, 134–152.
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was directly transferred to a reverse phase C-18 column and 17 R. R. Gadikota, C. S. Callam and T. L. Lowary, Org. Lett.,
purified by eluting with 0–10% MeOH–H2O. The appropriate 2001, 3, 607–610.
fractions were collected and concentrated in vacuo to afford 30 18 C. S. Callam, R. R. Gadikota, D. M. Krein and T. L. Lowary,
2340–2347.
1
(6.7 mg, 89%) as a white solid. Analytical data for 30: H NMR
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(500 MHz, D2O): δ, 5.12 (d, 1H, J = 4.5 Hz), 5.04 (d, 1H, J = 4.5 19 R. R. Gadikota, C. S. Callam, T. Wagner, B. Del Fraino
Hz), 5.02 (d, 1H, J = 3.0 Hz), 4.48 (dd, 1H, J = 5.0 Hz and 8.0
Hz), 4.30 (dd, 1H, J = 4.5 Hz and 8.0 Hz), 4.20–4.13 (m, 3H),
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Acknowledgements
This research was supported by the NSF (DBI 0421683).
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