A synthetic strategy to xylose-containing thioglycoside tri- and tetrasaccharide building blocks corresponding to Cryptococcus neoformans capsular polysaccharide structures

Article abstract of DOI:10.1039/c5ob00766f

As part of an ongoing project aimed at developing vaccine candidates against Cryptococcus neoformans the preparation of tri- and tetrasaccharide thioglycoside building blocks, to be used in construction of structurally defined part structures of C. neofor

Full text of DOI:10.1039/c5ob00766f

Organic &
Biomolecular Chemistry
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A synthetic strategy to xylose-containing
thioglycoside tri- and tetrasaccharide building
blocks corresponding to Cryptococcus neoformans
capsular polysaccharide structures†
Cite this: Org. Biomol. Chem., 2015,
13, 6598
Lorenzo Guazzelli, Rebecca Ulc, Lina Rydner and Stefan Oscarson*
As part of an ongoing project aimed at developing vaccine candidates against Cryptococcus neoformans
the preparation of tri- and tetrasaccharide thioglycoside building blocks, to be used in construction of
structurally defined part structures of C. neoformans GXM capsular polysaccharide, was investigated.
Using a naphthalenylmethyl (NAP) ether as a temporary protecting group and trichloroacetimidate donors
in optimized glycosylations the target building blocks, ethyl 6-O-acetyl-2,4-di-O-benzyl-3-O-
(2-naphthalenylmethyl)-α-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)]-4,6-di-
O-benzyl-1-thio-α-D-mannopyranoside (16) and ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-
di-O-benzyl-3-O-(2-naphthalenylmethyl)-α-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyra-
nosyl-(1→2)]-6-O-acetyl-4-O-benzyl-1-thio-α-D-mannopyranoside (21), were efficiently prepared. These
synthesized thiosaccharide building blocks were then used as donors in high-yielding (∼90%) DMTST pro-
moted glycosylations to a spacer-containing acceptor to, after deprotection, afford GXM polysaccharide
part structures ready for protein conjugation to give vaccine candidates. Also, the NAP groups in the build-
ing blocks were removed to obtain tri- and tetrasaccharide acceptors suitable for further elongation
towards larger thiosaccharide building blocks.
Received 16th April 2015,
Accepted 12th May 2015
DOI: 10.1039/c5ob00766f
www.rsc.org/obc
ing whole polysaccharide in such conjugate vaccines. Conse-
quently, a new approach of using synthetic oligosaccharides is
Introduction
Cryptococcus neoformans is a fungal pathogen causing severe attractive since it includes the possibility to focus the antibody
infections in immunocompromised individuals, e.g., it is now response on epitopes that elicit protective immunity.4,5 Such
the most prevalent infection in AIDS-patients in Africa.1 The synthetic structures would provide unique tools to investigate
discovery that monoclonal antibodies (mAbs) can protect the structure–activity relationship (SAR) between the immune
against cryptococcal infections in mice encouraged the devel- response and specific GXM structures and to investigate the
opment of vaccines against the infection.2–5 Cryptococcus neo- epitope specificity of protective and non-protective mAbs.
formans is surrounded by capsular polysaccharides, of which Structural analysis indicates that GXM are heteropolymers
the major one is the GXM-polysaccharide containing gluc- comprised of different triads linked together (Fig. 1),8 why the
uronic acid, xylose and mannose residues, and vaccines candi- GXM polysaccharide is highly variable and heterogeneous and
dates have been preferentially based on these structures. The can’t be used in SAR studies. Given this complexity of the GXM
polysaccharides themselves, in contrast to most bacterial poly- and that there are no enzymes available to regioselectively
saccharides, are not immunogenic, but glycoconjugate vac- cleave GXM, synthetic structures provide the only means for
cines composed of the polysaccharide linked to a carrier the planned studies. The aim is to explore the hypothesis that
protein have been tested and proven promising.6,7 However, it is possible to focus an immune response to make only pro-
the discovery that the polysaccharide contained epitopes that tective antibodies. We have already established the converse,
could elicit both protective, non-protective and even disease- that certain oligosaccharides can focus the response to non-
enhancing antibodies2,3 raised serious concerns about employ- protective antibodies.9–11 There is evidence that some protec-
tive epitopes are conformational.12 Hence, synthesizing longer
oligosaccharides that can attain conformations like those
found in the native polysaccharide is a major target. To
efficiently perform these syntheses, larger building blocks
Center for Synthesis and Chemical Biology, University College Dublin, Belfield,
Dublin 4, Ireland. E-mail: stefan.oscarson@ucd.ie
†Electronic supplementary information (ESI) available. See DOI: 10.1039/
than the disaccharides we have so far been using are desired.
c5ob00766f
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yields in the rearrangement reaction to give the enol ether
were poor. Various catalysts, e.g. PdCl2 and Wilkinson’s cata-
lyst, that had been successful with O-glycoside substrates,
failed with the thioglycoside. Our own methodology using
SmI2,17 which earlier had worked well on thioglycosides, also
proved unsuccessful. Performing the rearrangement with a
strong base finally afforded the 3-OH acceptor but only in a
moderate yield (∼40%). These experiences forced us to develop
a new strategy including a more suitable temporary protecting
group. Before the selection of the allyl group, other protecting
groups had been tested, mainly a TBDMS group and a p-meth-
oxybenzyl (pMBn) group, but both these were found to be
inferior to the allyl group, the TBDMS group due to problems
with migration as well as low yields in glycosylation and
benzylidene opening reactions and the pMBn groups due to its
acid lability.18 However, since there is not that many good tem-
porary protecting groups and most acyl protecting groups are
not compatible with the presence of acetates in the target
structures, routes using the TBDMS and the pMBn groups
Fig. 1 Suggested structures of C. neoformans GXM serotype triads.
Herein we describe the development of a strategy that allows were revisited. Again the TBDMS group was found to give low
construction of larger donor structures and its application to yields in a number of reactions but the problem with the acid
the synthesis of xylose-containing tri- and tetrasaccharide thio- lability of the pMBn group could be overcome, at least in most
glycoside building blocks and their subsequent transformation reactions.
into acceptors as well as their use as donors in synthesis
Hence, 3-O-pMBn protected disaccharide building blocks
of spacer-containing tetra-and pentasaccharide GXM part could be prepared in a five-step synthesis starting from benzo-
structures.
bromoxylose and ethyl 4,6-O-benzylidene-3-O-(4-methoxy-
benzyl)-1-thio-α-D-mannopyranoside19 in around an overall
yield of 35%.20 These could then be effectively converted to
bromosugar donors or 3-OH acceptors through treatment with
Br2 or DDQ, respectively (Scheme 1).20
Results and discussion
As mentioned in the introduction the structure of the GXM is
However, when these acceptors and donors were tested in
believed to be built up of triads, i.e., substituted trisaccharide orthogonal glycosylation reactions the yields were again low.
mannans, as depicted in Fig. 1, and the serotyping depending Not only hydrolysis of the pMBn ether was again a problem
on the amount of xylose substituents. GXM is also hetero- but also hydrolysis of the glycosyl bromide was observed. With
geneously acetylated. The acetylation is crucial for virulence, these reoccurring problems with the p-methoxybenzyl group,
knock-out mutants that can’t introduce the acetates are not our attention was turned to the 2-naphthalenylmethyl (NAP)
virulent.13 The acetates are positioned at the 6-OH of the group, which is removed like the p-methoxybenzyl group by
mannose backbone but not present in the residues carrying DDQ or CAN treatment but considered to be more acid
4-O-xylose substitutions and in serotype A and D there are stable.21 We recently used this protecting group in the syn-
about 2 acetates per mannose triad. Serotype A and D are the thesis of glucuronic acid containing disaccharide GXM build-
most prevalent in humans and these structures (both without ing blocks.22 The NAP group was introduced regioselectively
4-O-xylose substitutions) are therefore our primary targets. The on 223 by known tin activation methodology24 in an 88% yield
A/D donor building blocks, synthesized and used in vaccine (Scheme 2). The coupling of 125 and 3 was then carried out
candidate syntheses, so far have been ethyl thioglycosides with using TBDMSOTf in the presence of commercial acid-washed
benzyl ethers and esters as permanent protecting groups, and molecular sieves to prevent orthoester formation. Compound 4
a 3-O-allyl group as a temporary protecting group.14–16 This
approach has worked well and allowed efficient synthesis of up
to heptasaccharide structures with variant acetylation pat-
terns.9 However, when these blocks were to be combined to
yield larger thioglycoside structures through orthogonal glyco-
sylations, issues were encountered with the allyl protecting
group. The unsaturated allyl function precluded the transform-
ation of the thioglycoside into a bromo sugar donor using
bromine and, more unexpectedly, problems were experienced
in the removal of the allyl group to create a thioglycoside
Scheme 1 Reagents and conditions: (a) Br2, CH2Cl2, 0 °C; (b) DDQ,
acceptor. Many deallylation methods were tested, but the CH2Cl2/H2O 20 : 1, 0 °C.
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Scheme 2 Reagents and conditions: (a) Bu2SnO, Bu4NI, NA PBr, toluene, reflux 88%; (b) TBDMSOTf, CH2Cl2, AW-300 MS, −78 °C to 20 °C, over
night, 86%; (c) NaOMe, MeOH, 20 °C, 88%; (d) NaH, BnBr, DMF, 0 °C to 20 °C, 88% for 6, 99% for 9; (e) Bu2BOTf, BH3 (1 M in THF), 94% for 7, 84%
for 11; (f) Ac2O, pyridine, 20 °C, 98% for 8, 96% for 12.
was obtained in an 86% yield. At this stage benzoyl groups, t-BuOH. This second solvent system gave slightly better yields
which ensured the stereoselective course of the glycosylation, (75% and 70% respectively) and reduced the benzyl removal
were exchanged for benzyl groups affording 6 in a 77% yield side reaction which has been reported by Crich and Yao.28
over two steps. The regioselective opening of the benzylidene
Considering our earlier experiences with the orthogonal
26
ring using Bu2BOTf/BH3 afforded compound 7 (94%), which silver triflate-promoted glycosylations using the pMBn-
was either acetylated (→8, 98%) or benzylated (→9, 99%). The protected bromo sugar donors, where major side products
monosaccharide building block 12, required for the construc- were due to hydrolysis of the glycosyl bromide in addition to
tion of serotype D structures, was prepared from 1027 by the pMBn group, we decided to try trichloroacetimidates as gly-
opening of the benzylidene ring, employing the same con- cosyl donors instead. The imidate donors were obtained from
ditions used for 7 (→11, 84%) followed by acetylation (→12, 9 and 12 after hydrolysis of the thioethyl glycoside with NIS
96%).
and trifluoroacetic acid in a vigorously stirred CH2Cl2/H2O
From this set of mono- and disaccharide building blocks mixture and subsequent activation of the anomeric position
the synthesis of the target tri- and tetrasaccharide building (DBU, Cl3CCN in CH2Cl2).
blocks, 16 (serotype D) and 21 (serotype A), respectively, was
Initially, couplings with the new trichloroacetimidate
now attempted. The same strategy was used for both serotypes donors 14 and 19 proceeded in very low yields (Schemes 3 and
(reported in Schemes 3 and 4) and involves two steps prior to 4). Optimizing the conditions for tetrasaccharide 21, catalysts
the glycosylation reaction: (1) preparation of the thioglycoside TMSOTf and TBDMSOTf were found to be the best, but in
acceptor by removing the orthogonal protecting group; and (2) CH2Cl2 they still produced 21 in a maximum yield of only
conversion of the thioglycoside donor into a suitable one for 25%. Changing the solvent to either diethyl ether or aceto-
the orthogonal glycosylation.
nitrile gave no product at all, but when toluene was tried (at
Acceptors 15 and 20 were obtained by removing the NAP −20 °C) a better yield was obtained (∼50%). Further optimiz-
group with DDQ in a mixture of CH2Cl2/H2O or CH2Cl2/ ation established that employing the inverse glycosylation pro-
Scheme 3 Reagents an d conditions: (a) NIS, TFA, CH2Cl2/H2O (20 : 1), 0 °C, 1 h, 86% (α/β = 9 : 1); (b) Cl3CCN, DBU, CH2Cl2, 0 °C, 9 0 min, 90%; (c)
DDQ, CH2Cl2/H2O (10 : 1), 3.5 h min, 65% or DDQ, CH2Cl2/t-BuOH (10 : 1), 90 min, 75%; (d) TMSOTf, toluene, A W-300 MS, −20 °C, 16 : 17 =
65% : 18% or TMSOTf, toluene, AW-3 00 MS, rt, 16 : 17 = 75% : 9%.
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Scheme 4 Reagents and conditions: (a) NIS, TFA, CH2Cl2/H2O (20 : 1), 0 °C, 90 min, 8 5% (α/β = 5 : 1); (b) Cl3CCN, DBU, CH2Cl2, 0 °C, 90 min,
quant.; (c) DDQ, CH2Cl2/H2O ( 20 : 1), 90 min, 70%; (d) TBDMSOTf, toluene, AW-300 MS, −40 °C, 95%.
cedure29 to these couplings, i.e., adding the donor to a premix deprotected by means of hydrogenolysis to afford spacer-
of the acceptor and the promoter, was a most high-yielding equipped GXM part structures 27 and 28 in a 72% and a 69%
and reliable methodology to the target structures. Tetra- yield respectively (Scheme 6).
saccharide 21 was reproducibly obtained in over 90% yield
from acceptor 20 and donor 19 (1.2 equiv.). In the synthesis of
the trisaccharide 16 a 3.5 : 1 α/β-mixture was obtained in an
Conclusions
83% yield using the same conditions. The anomeric configur-
1
1
ations were unambiguously confirmed by the H–13C J coup-
ling constant value (16, JC1′–H1′ = 175 Hz; 17, JC1′–H1′ = 155 Hz).
Higher α-selectivity, 8 : 1 (84% yield), was obtained running
the glycosylation reaction at room temperature instead of at
−20 °C. Similar temperature effects on the stereoselectivity of
glycosylations have been reported.30
In conclusion, an efficient and reliable strategy to tri- and
tetrasaccharide thioglycoside building blocks corresponding to
Cryptococcus neoformans GXM oligosaccharide structures has
been developed. The introduction of a 2-naphthalenylmethyl
ether as a temporary protecting group allowed efficient trans-
formation of synthesized thioglycoside mono- and disacchar-
ides into both 3-OH acceptors and other types of glycosyl
donors. Use of trichloroacetimidate donors in couplings
employing the inverse glycosylation protocol with TBDMOTf or
TMSOTf as promoters in toluene was found to be an efficient
methodology to construct tri- and tetrasaccharide thioglyco-
side building blocks, which can be converted into even larger
building blocks by reiteration of the developed approach. The
synthesized thioglycoside building blocks were shown to be
excellent donors in model glycosylation reactions proceeding
with complete α-selectivity in ∼90% yield and were also effec-
tively transformed into acceptor structures.
The new thiosaccharide building blocks 16 and 21 were
then tested as donors in dimethyl(methylthio)sulfonium tri-
flate (DMTST)31 promoted glycosylations with acceptor 2214
(Scheme 5). Both glycosylations proceeded with complete
α-stereoselectivity and derivatives 23 and 24 were obtained in
89% and 92% yield respectively.
Also, compounds 16 and 21 were converted, through DDQ-
based removal of the NAP group, to two new 3-OH acceptors
(Scheme 5), the trisaccharide 25 (72%) and the tetrasaccharide
26 (68%), to allow continued synthesis of even larger thioglyco-
side building blocks. Compounds 23 and 24 were completely
Scheme 5 Reagents and conditions: (a) DDQ, CH2Cl2/H2O or t-BuOH (10 : 1), 72% for 25, 68% for 26; (b) DMTST, Et2O, MS 4 Å, 0 °C to 20 °C, 89%
for 23, 92% for 24.
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Stark trap). After 3 h, the mixture was concentrated to half-
volume, (Bu)4NI (20.75 g, 56.18 mmol) and NAPBr (10.12 g,
45.77 mmol) were added, and the reaction mixture was
refluxed for another 3 h (TLC, toluene/EtOAc, 1 : 3). The reac-
tion mixture was diluted with EtOAc (400 mL), and the organic
layer was washed sequentially with H2O (3 × 300 mL), 10% aq.
KF-solution (3 × 300 mL) and brine (1 × 300 mL), dried over
MgSO4, and concentrated in vacuo to an orange oily residue.
Purification by flash column chromatography (SiO2, 350 mL,
7.5 cm, pentane→pentane–Et2O, 3 : 1→2 : 1→1 : 1→1 : 2→1 : 3)
gave 3 (16.66 g, 88%) as a pale yellow syrup; Rf 0.50 (toluene/
EtOAc 3 : 1); [α]2D0 +143.4 (c 1.0, CHCl3); 1H NMR (500 MHz,
CDCl3): δ = 7.82–7.71 (m, 4H, Ar-H), 7.51–7.36 (m, 8H, Ar-H),
5.62 (s, 1H), 5.37 (s, 1H), 4.98 (benzylic d, 1H, Jgem = 11.5 Hz),
4.87 (benzylic d, 1H, Jgem = 11.5 H), 4.25–4.21 (m, 2H),
4.19–4.15 (m, 2H), 3.96–3.93 (dd, 1H, J = 3.5 Hz, J = 9.5 Hz),
3.88 (m, 1H), 2.87 (s, 1H), 2.66–2.53 (m, 2H,), 1.27 (t, 3H,
SCH2CH3); 13C NMR (125 MHz, CDCl3): δ = 137.5, 135.2, 133.2,
133.1, 128.9, 128.3, 128.2, 127.9, 127.7, 126.6, 126.1, 126.0,
125.6, 101.7, 84.2, 79.1, 75.9, 73.0, 71.4, 68.7, 63.8, 24.9, 14.8.
HRMS (ESI): [M + Na]+ calcd for C26H28O5NaS, 475.1555.
Found: 475.1534; anal. calcd for C26H28O5S: C, 69.00; H, 6.24;
S, 7.09. Found: C, 68.93; H, 6.06; S, 7.34%.
Ethyl 2,3,4-tri-O-benzoyl-β-D-xylopyranosyl-(1→2)-4,6-O-
benzylidene-3-O-(2-naphthalenylmethyl)-1-thio-α-D-
mannopyranoside (4)
Scheme 6 Reagents and conditions: (a) H2/Pd-C (30 bar), EtOAC, H2O,
AcOH, 72% for 27, 69% for 28.
A catalytic amount of TBDMSOTf (450 μL, 1.96 mmol) was
added to a solution of donor 1 (11.90 g, 19.62 mmol) and
acceptor 3 (8.88 g, 19.62 mmol) in dry CH2Cl2 (250 mL) con-
taining crushed acid-washed molecular sieves (Aldrich
AW-300) kept at −78 °C in an atmosphere of nitrogen. The
temperature was allowed to rise to 20 °C overnight (TLC,
toluene/EtOAc 6 : 1). The reaction mixture was neutralised with
Et3N (1.64 mL, 11.77 mmol), the solids were removed by fil-
tration, and the filtrate was concentrated in vacuo to
a yellowish foam. Purification by flash column chromato-
Experimental section
TLC was carried out on precoated 60 F254 silica gel alumina
plates (Merck) using UV-light and/or 8% H2SO4 and/or AMC-
solution (ammonium molybdate, cerium(IV) sulphate, 10%
H2SO4 [5 : 0.1 : 100, w/w/v] for visualization. Flash column
chromatography was performed on silica gel (Merck, pore size
60 Å, particle size 40–63 μm). NMR spectra were recorded in
CDCl3 (internal Me4Si d = 0.00 ppm) or D2O (standardized
against the residual solvent peak, d = 4.79) at 25 °C on a
Varian instrument (500 MHz for 1H and 125 MHz for 13C or
graphy
(SiO2,
400
mL,
7.5
cm,
toluene/EtOAc,
98 : 2→96 : 4→94 : 6→92 : 8→90 : 10→88 : 12→ 86 : 14) gave 4
(15.09 g, 86%) as a colourless amorphous solid; Rf 0.58
(toluene/EtOAc 9 : 1); [α]2D0 −0.3 (c 1.0, CHCl3); 1H NMR
(500 MHz, CDCl3): δ = 8.22 (m, 2H), 8.07 (m, 2H, Ar-H), 7.98
(m, 2H, Ar-H), 7.86 (m, 1H, Ar-H), 7.81–7.79 (m, 2H, Ar-H),
7.71–7.70 (m, 1H, Ar-H), 7.58–7.36 (m, 15H, Ar-H), 7.28–7.25
(m, 2H, Ar-H), 5.63 (t, 1H, J = 4.0 Hz), 5.36 (t, 1H, J = 3.0 Hz),
5.32 (s, 1H), 5.15 (d, 1H, J = 1.5 Hz), 5.10 (m, 2H), 5.04
1
600 MHz for H and 150 MHz for 13C). Coupling constants are
given in Hertz (Hz). HRMS spectra were recorded on a micro-
mass LCT instrument using electrospray ionisation (ESI) in
either the positive or negative modes. Optical rotations were
measured with a Perkin-Elmer 343 polarimeter at the sodium
D-line (589 nm) at 20 °C using a 1 dm cell. All reactions con-
taining air- and moisture-sensitive reagents were carried out
under an argon atmosphere. Organic phases were dried over
MgSO4 before evaporation, which was performed under
reduced pressure at temperatures not exceeding 40 °C.
(benzylic d, 1H, Jgem = 12.4 Hz), 4.98 (benzylic d, 1H, Jgem
=
12.4 Hz), 4.91 (dd, 1H, J = 2.5 Hz, J = 12.5 Hz), 4.36 (s, 1H),
4.12–4.07 (m, 1H), 4.03–4.00 (m, 3H), 3.78 (dd, 1H, J = 3.0 Hz,
J = 13.0 Hz), 3.51 (t, 1H, J = 10.0 Hz), 2.61–2.49 (m, 2H), 1.21 (t,
3H); 13C NMR (125 MHz, CDCl3): δ = 165.6, 165.2, 164.9, 137.7,
135.8, 133.4, 133.3, 133.0, 130.3, 130.0, 129.9, 129.6, 129.3,
128.9, 128.5, 128.4, 128.3, 128.1, 127.9, 127.7, 126.3, 126.2,
Ethyl 4,6-O-benzylidene-3-O-(2-naphthalenylmethyl)-1-thio-
α-D-mannopyranoside (3)
A solution of compound 2 (13.00 g, 41.62 mmol) and (Bu)2SnO 126.1, 125.8, 125.6, 101.4, 95.9, 82.3, 78.9, 76.1, 75.4, 73.1,
(13.98 g, 56.18 mmol) in anhydrous toluene (420 mL) was 68.5, 68.3, 67.9, 67.4, 64.5, 59.4, 25.5, 14.9; HRMS (ESI): [M +
heated under reflux with continuous removal of water (Dean- Na]+ calcd for C52H48O12NaS, 919.2764; found, 919.2803; anal.
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calcd for C52H48O12S: C, 69.63; H, 5.39; S, 3.57. Found: C, 4.71 (benzylic d, 1H, Jgem = 12.0 Hz), 4.61 (benzylic d, 1H, Jgem
=
69.09; H, 5.20; S, 3.64%.
12.0 Hz), 4.45 (d, 1H, J = 7.0 Hz), 4.28 (dd, 1H, J = 2.5 Hz, J = 1.0
Hz), 4.26–4.19 (m, 3H), 4.00 (dd, 1H, J = 5.0 Hz, J = 12.0 Hz),
3.97–3.94 (m, 1H), 3.84–3.80 (m, 1H), 3.69–3.65 (m, 1H), 3.60 (t,
1H, J = 8.7 Hz), 3.58–3.54 (m, 1H), 3.25 (dd, 1H, J = 11.6 Hz, J =
Ethyl β-D-xylopyranosyl-(1→2)-4,6-O-benzylidene-3-O-
(2-naphthalenylmethyl)-1-thio-α-D-mannopyranoside (5)
A catalytic amount of NaOMe (7 mg, 0.12 mmol) was added to 9.7 Hz), 2.62–2.51 (m, 2H), 1.22 (t, 3H); 13C NMR (125 MHz,
a solution of 4 (1.12 g, 1.25 mmol) in dry MeOH (20 mL). The CDCl3): δ = 138.7, 138.4, 138.1, 137.7, 135.8, 133.3, 132.9, 128.8,
mixture was stirred at 20 °C overnight (TLC, CH2Cl2/MeOH, 128.5, 128.4, 128.3, 128.2, 127.9, 127.9, 127.9, 127.8, 127.8,
9 : 1). After complete conversion, Dowex® (H+) acidic ion 127.6, 127.6, 126.3, 126.1, 125.9, 125.7, 125.6, 103.2, 101.6, 83.8,
exchange resin was added for neutralisation, the resin was fil- 83.3, 81.4, 78.8, 77.8, 75.5, 75.1, 74.7, 73.3, 71.8, 68.8, 64.6, 64.1,
tered off, washed with MeOH and the filtrate was concentrated 25.6, 15.0; HRMS (ESI): [M + Na]+ calcd for C52H54O9NaS,
in vacuo. Purification by flash column chromatography 877.3386; found, 877.3403; anal. calcd for C52H54O9S: C, 73.04;
on silica gel (SiO2, 200 mL, 4.5 cm, CH2Cl2/MeOH, H, 6.37; S, 3.75. Found: C, 73.11; H, 6.33; S, 3.91%.
99 : 1→98 : 2→97 : 3→96 : 4→95 : 5) gave 5 (0.64 g, 88%) as a
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-3-O-(2-
naphthalenylmethyl)-4-O-benzyl-1-thio-α-D-mannopyranoside (7)
colourless, amorphous solid; Rf 0.39 (CH2Cl2/MeOH 9 : 1);
[α]2D0 +37.4 (c 1.0, CHCl3); 1H NMR (500 MHz, CDCl3): δ =
7.81–7.73 (m, 4H, Ar-H), 7.52–7.50 (m, 2H, Ar-H), 7.47–7.43 (m, A 1 M solution of BH3 in THF (138.0 mL, 138.0 mmol) was
3H, Ar-H), 7.39–7.37 (m, 3H, Ar-H), 5.65 (s, 1H), 5.29 (d, J = 1.1 added to a solution of 6 (11.80 g, 13.80 mmol) in dry CH2Cl2
Hz, 1H), 5.03 (benzylic d, 1H, Jgem = 12.0 Hz), 4.86 (benzylic d, (200 mL) kept at 0 °C in an atmosphere of nitrogen. After
1H, Jgem = 12.0 Hz), 4.58 (d, 1H, J = 6.0 Hz), 4.24–4.20 (m, 4H), 5 min, a 1 M solution of Bu2BOTf (13.80 mL, 13.80 mmol) was
4.05 (dd, 1H, J = 4.5 Hz, J = 12.0 Hz), 4.01 (dd, 1H, J = 9.5 Hz, added dropwise at 0 °C, and the reaction mixture was stirred
J = 3.4 Hz), 3.93 (d, 1H, J = 3.5 Hz), 3.90–3.86 (m, 1H), for 90 min (TLC, toluene/EtOAc 6 : 1). After complete consump-
3.62–3.61 (m, 1H), 3.53–3.48 (m, 2H), 3.31 (d, 1H, J = 2.5 Hz), tion of the starting material, the reaction was quenched with
3.26 (dd, 1H, J = 8.5 Hz, J = 12.0 Hz), 2.79 (d, 1H, J = 3.5 Hz), Et3N, followed by dropwise addition of MeOH at 0 °C. The
2.67–2.55 (m, 2H), 1.26 (t, 3H, J = 7.5 Hz); 13C NMR (125 MHz, mixture was concentrated in vacuo, and the residue was re-
CDCl3): δ = 137.5, 134.7, 133.2, 133.1, 129.0, 128.3, 128.28, dissolved and co-concentrated with MeOH (3 × 200 mL). The
128.0, 127.7, 127.3, 126.2, 126.1, 126.0, 101.7, 100.7, 84.3, 79.7, syrupy residue was re-dissolved in MeOH, the solution was fil-
75.0, 74.2 (2C), 73.8, 69.9, 69.4, 68.6, 64.9, 64.4, 25.6, 15.0. tered through a short pad of Celite, and the filtrate was con-
HRMS (ESI): [M + H]+ calcd for C31H37O9S, 585.2158; found, centrated in vacuo to a yellow oil. Purification by flash column
585.2150; anal. calcd for C31H36O9S: C, 63.68; H, 6.21; S, 5.48. chromatography (SiO2, 500 mL, 7.5 cm, toluene→toluene/
Found: C, 60.97; H, 5.96; S, 5.00%.
EtOAc, 97 : 3→96 : 4→91 : 9→88 : 12→85 : 15→82 : 18→ 79 : 21)
gave 7 (10.98 g, 94%) as a colourless foam; Rf 0.38 (toluene/
EtOAc 6 : 1); [α]2D0 +42.0 (c 1.0, CHCl3); 1H NMR (500 MHz,
CDCl3): δ = 7.83–7.81 (m, 2H, Ar-H), 7.77–7.75 (m, 1H, Ar-H),
7.72–7.70 (m, 1H, Ar-H), 7.50 (dd, 1H, J = 2.0 Hz, J = 9.0 Hz, Ar-
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-O-
benzylidene-3-O-(2-naphthalenylmethyl)-1-thio-α-D-
mannopyranoside (6)
NaH (2.83 g, 70.81 mmol, 60% oil dispersion) was washed H), 7.48–7.44 (m, 2H, Ar-H), 7.38–7.27 (m, 20H, Ar-H), 5.39 (d,
with pentane (3 × 80 mL) prior to use. NaH was added portion- 1H, J = 1.5 Hz), 5.02 (benzylic d, 1H, Jgem = 10.5 Hz), 4.99
wise to a solution of 5 (9.20 g, 15.74 mmol) in dry DMF (benzylic d, 1H, Jgem = 11.0 Hz), 4.92 (benzylic d, 1H, Jgem
=
(100 mL) at 0 °C in an atmosphere of nitrogen. After 15 min, 12.0 Hz), 4.86 (benzylic ABq, 2H, J = 13.2 Hz), 4.74–4.71 (m,
BnBr (7.48 mL, 62.96 mmol) was added dropwise at 0 °C 2H), 4.62 (benzylic d, 1H, Jgem = 11.7 Hz), 4.60 (benzylic d, 1H,
under vigorous stirring. The temperature was then allowed to Jgem = 10.9 Hz), 4.42 (d, 1H, J = 7.5 Hz), 4.22 (s, 1H), 4.00–3.94
rise 20 °C over 2 h (TLC, toluene/EtOA 6 : 1). After complete (m, 2H), 3.92–3.91 (m, 2H), 3.77–3.65 (m, 3H), 3.59 (t, 1H, J = 8.8
consumption of the starting material, residual NaH was Hz), 3.56–3.52 (m, 1H), 3.23 (dd, 1H, J = 11.4 Hz, J = 10.2 Hz),
quenched with MeOH, and then with H2O (300 mL). The 2.60–2.49 (m, 2H), 1.23 (t, 3H, J = 7.5 Hz); 13C NMR (125 MHz,
resulting mixture was extracted once with EtOAc (400 mL), the CDCl3): δ = 138.7, 138.6, 138.5, 138.2, 135.4, 133.3, 133.1, 128.5,
layers were separated, and the organic layer was washed with 128.4, 128.4, 128.3, 128.0, 128.0, 127.9, 127.9, 127.9, 127.7, 127.
brine (1 × 300 mL), dried over MgSO4 and concentrated 7, 127.6, 127.5, 127.2, 126.4, 126.0, 125.8, 103.2, 83.9, 82.2, 81.2,
in vacuo. Purification by flash column chromatography 78.7, 77.5, 76.5, 75.5, 75.2, 74.8, 73.4, 72.2, 71.4, 64.2, 62.5, 25.5,
(SiO2,
600
mL,
7.5
cm,
toluene→toluene/EtOAc, 14.9; HRMS (ESI): [M + Na]+ calcd for C52H56O9NaS, 879.3543;
98 : 2→96 : 4→94 : 6→ 92 : 8→90 : 10→88 : 12) gave 6 (11.80 g, found, 879.3524; anal. calcd for C52H56O9S: C, 72.87; H, 6.59; S,
88%) as a colourless, amorphous solid; Rf 0.58 (toluene/EtOAc 3.74. Found: C, 72.71; H, 6.57; S, 3.83%.
1
5 : 1); [α]2D0 −1.6 (c 1.0, CHCl3); H NMR (500 MHz, CDCl3): δ =
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-6-O-acetyl-4-O-
7.85 (m, 1H, Ar-H), 7.81–7.79 (m, 1H, Ar-H), 7.76–7.75 (m, 1H,
benzyl-3-O-(2-naphthalenylmethyl)-1-thio-α-D-mannopyranoside (8)
Ar-H), 7.68–7.66 (m, 1H, Ar-H), 7.52–7.26 (m, 23H, Ar-H), 5.60
(s, 1H), 5.41 (d, J = 0.8 Hz, 1H), 5.01 (benzylic d, 1H, Jgem = 10.0 Ac2O (5.71 mL, 60.29 mmol) was added to a solution of 7
Hz), 4.92–4.84 (m, 4H), 4.72 (benzylic d, 1H, Jgem = 10.0 Hz), (6.46 g, 7.54 mmol) in dry pyridine (100 mL), and the mixture
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was stirred at 20 °C for 5 h (TLC, toluene/EtOAc, 9 : 1). The J = 12.0 Hz), 3.92 (dd, 1H, J = 3.0 Hz, J = 9.0 Hz), 3.78 (dd, 1H,
reaction mixture was concentrated in vacuo, and then re-dis- J = 4.0 Hz, J = 10.5 Hz), 3.69–3.64 (m, 2H), 3.58–3.51 (m, 2H),
solved and co-evaporated with toluene (3 × 100 mL). Purifi- 3.22 (m, 1H), 2.64–2.52 (m, 2H), 1.23 (t, 3H); 13C NMR
cation by flash column chromatography (SiO2, 200 mL, 4.5 cm, (125 MHz, CDCl3): δ = 138.9, 138.7, 138.6, 138.3, 138.2, 135.5,
toluene→toluene/EtOAc, 97 : 3→96 : 4→91 : 9→88 : 12→85 : 15) 133.3, 133.1, 129.0, 128.5, 128.3, 128.2, 128.1, 128.0, 128.0,
gave 8 (6.64 g, 98%) as a colourless oil; Rf 0.59 (toluene/EtOAc 127.9, 127.9, 127.9, 127.7, 127.6, 127.5, 127.4, 127.3, 127.2,
6 : 1); [α]2D0 +46.0 (c 1.0, CHCl3); 1H NMR (500 MHz, CDCl3): δ = 126.5, 125.9, 125.8, 103.5, 84.0, 82.3, 81.1, 78.9, 77.4, 76.9,
7.83–7.81 (m, 2H, Ar-H), 7.77 (benzylic d, 1H, J = 8.5 Hz, Ar-H), 75.5, 75.2, 75.0, 74.7, 73.4, 73.2, 72.0, 71.4, 69.5, 64.2, 25.5,
7.72–7.70 (m, 1H, Ar-H), 7.51 (benzylic dd, 1H, J = 1.0 Hz, J = 15.0; HRMS (ESI): [M + Na]+ calcd for C59H62O9NaS, 969.4012;
8.5 Hz, Ar-H), 7.48–7.45 (m, 2H, Ar-H), 7.40–7.25 (m, 20H, Ar- found, 969.4032; anal. calcd for C59H62O9S: C, 74.81; H, 6.60;
H), 5.38 (s, 1H), 5.10 (benzylic d, 1H, Jgem = 10.0 Hz), 4.98 S, 3.39. Found: C, 74.70; H, 6.56; S, 3.28%.
(benzylic d, 1H, Jgem = 10.5 Hz), 4.94–4.85 (m, 3H), 4.75–4.71
(m, 2H), 4.64–4.61 (m, 2H), 4.55 (benzylic d, 1H, Jgem = 10.5
Hz), 4.40 (d, 1H, J = 7.5 Hz), 4.34 (dd, 1H, J = 4.3 Hz, J = 11.9
Hz), 4.27 (dd, 1H, J = 1.9 Hz, J = 11.9 Hz), 4.22 (m, 1H),
4.20–4.17 (m, 1H), 4.00–3.95 (m, 2H), 3.91 (dd, 1H, J = 3.5 Hz,
J = 9.5 Hz), 3.71–3.66 (m, 1H), 3.60–3.54 (m, 2H), 3.23 (t, 1H,
J = 10.5 Hz), 2.60–2.55 (m, 2H), 1.73 (s, 3H), 1.23 (t, 3H); 13C
NMR (125 MHz, CDCl3): δ = 170.6, 138.8, 138.3, 138.2, 138.1,
135.3, 133.3, 133.1, 128.9, 128.5, 128.4, 128.3, 128.1, 128.0,
127.9, 127.8, 127.7, 127.6, 127.3, 126.5, 126.0, 125.9, 103.5,
84.0, 82.5, 81.3, 78.8, 77.4, 76.7, 75.6, 75.2, 75.1, 74.2, 73.4,
71.4, 70.0, 64.2, 63.4, 25.7, 20.5, 15.0. HRMS (ESI): [M + Na]+
calcd for C54H58O10NaS, 921.3648; found, 921.3619; anal. calcd
for C54H58O10S: C, 72.14; H, 6.50; S, 3.57. Found: C, 71.99; H,
6.45; S, 3.43%.
Ethyl 2,4-di-O-benzyl-3-O-(2-naphthalenylmethyl)-1-thio-
α-D-mannopyranoside (11)
Compound 10 (1.30 g, 2.40 mmol) was dissolved in 1 M BH3 in
THF (24 mL, 24 mmol) and the mixture was stirred at 0 °C for
5 min before adding drop wise 1 M Bu2BOTf in CH2Cl2
(2.4 mL, 2.4 mmol). After 2 h, Et3N was carefully added fol-
lowed by drop wise addition of MeOH until gas evolution
ceased. The solution was co-concentrated three times with
MeOH and the crude was purified by flash column chromato-
graphy on silica gel (toluene/EtOAc 7 : 3) to give pure 11
(1.10 g, 2.02 mmol, 84%) as a syrup; Rf 0.44 (toluene/EtOAc
1
7 : 3); [α]2D0 +65.3 (c 1.35, CHCl3); H NMR (500 MHz, CDCl3): δ
= 7.83–771 (m, 4H, Ar-H), 7.48–7.22 (m, 13H, Ar-H), 5.31 (d,
1H, J = 1.3 Hz), 4.97 (benzylic d, 1H, Jgem = 11.0 Hz), 4.75–4.70
(m, 4H), 4.68 (benzylic d, 1H, Jgem = 11.0 Hz), 4.06–3.98 (m,
2H), 3.92 (dd, 1H, J = 3.0, J = 8.7 Hz), 3.88 (dd, 1H, J = 1.3, J =
3.0 Hz), 3.85–3.77 (m, 2H), 2.61–2.48 (m, 2H), 1.98 (t, 1 H, J =
6.5 Hz), 1.21 (t, 3H, J = 7.4 Hz); 13C NMR (125 MHz, CDCl3): δ =
138.6, 138.2, 135.8, 133.4, 133.1, 128.5, 128.5, 128.2, 128.1,
128.0, 128.0, 127.9, 127.8, 127.8, 126.5, 126.2, 126.0, 125.9,
82.4, 80.6, 76.8, 75.3, 75.1, 72.6, 72.5, 72.3, 62.5, 25.5, 15.0;
HRMS (ESI): [M + Na]+ calcd for C33H36NaO5S, 567.2181;
found, 567.2162.
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-
benzyl-3-O-(2-naphthalenylmethyl)-1-thio-α-D-
mannopyranoside (9)
NaH (734 mg, 18.34 mmol, 60% oil dispersion) was washed
with pentane (3 × 30 mL) prior to use. NaH was added portion-
wise to a solution of 7 (4.49 g, 5.24 mmol) in dry DMF
(100 mL) at 0 °C in an atmosphere of nitrogen. After 15 min,
BnBr (1.87 mL, 15.72 mmol) was added dropwise at 0 °C
under vigorous stirring. The temperature was then allowed to
rise 20 °C over 4 h (TLC, toluene/EtOA 9 : 1). After complete
consumption of the starting material, residual NaH was
quenched with MeOH, and then with H2O (200 mL). The
Ethyl 6-O-acetyl-2,4-di-O-benzyl-3-O-(2-naphthalenylmethyl)-
1-thio-α-D-mannopyranoside (12)
resulting mixture was extracted once with EtOAc (400 mL), the Ac2O (10 mL) was added to a solution of 11 (1.02 g,
layers were separated, and the organic layer was washed with 1.87 mmol) in pyridine (20 mL). The mixture was stirred at
brine (1 × 200 mL), dried over MgSO4 and concentrated room temperature overnight and then co-evaporated with
in vacuo. Purification by flash column chromatography toluene (3 × 40 mL). The crude was purified by flash column
(SiO2,
200
mL,
4.5
cm,
toluene→toluene/EtOAc, chromatography on silica gel (cyclohexane/EtOAc 9 : 1) to give
98 : 2→96 : 4→94 : 6→92 : 8 →90 : 10) gave 9 (5.02 g, >99%) as a pure 12 (1.05 g, 1.80 mmol, 96%) as a white foam; Rf 0.17 (9 : 1
colourless syrup. Rf 0.59 (toluene/EtOAc 9 : 1); [α]D20 +44.0 (c 1.0, cyclohexane/EtOAc); [α]D20 +61.2 (c 1.03, CHCl3); 1H NMR
1
CHCl3); H NMR (500 MHz, CDCl3): δ = 7.82–7.81 (m, 2H, Ar- (500 MHz, CDCl3): δ = 7.84–7.70 (m, 4H, Ar-H), 7.48–7.21 (m,
H), 7.76–7.74 (m, 1H, Ar-H), 7.70–7.68 (m, 1H, Ar-H), 7.51–7.12 13H, Ar-H), 5.37 (d, 1H, J = 1.1 Hz), 4.96 (benzylic d, 1H, Jgem
=
(m, 28H, Ar-H), 5.41 (d, 1H, J = 1.0 Hz), 5.10 (benzylic d, 1H, 10.9 Hz), 4.74–4.65 (m, 4H), 4.60 (benzylic d, 1H, Jgem = 10.9
Jgem = 11.0 Hz), 4.97 (benzylic d, 1H, Jgem = 10.5 Hz), 4.92 Hz), 4.39 (dd, 1H, J = 11.9 Hz, J = 5.0 Hz), 4.32 (dd, 1H, J = 2.2,
(benzylic d, 1H, Jgem = 11.5 Hz), 4.90 (benzylic d, 1H, Jgem
=
11.9 Hz), 4.18 (ddd, 1H, J = 2.2, 5.0, 9.4 Hz), 3.97 (t, 1H, J = 9.3
11.5 Hz), 4.83 (benzylic d, 1H, Jgem = 11.5 Hz), 4.74–4.71 (m, Hz), 3.91 (dd, 1H, J = 3.0, 9.3 Hz), 3.89 (dd, 1H, J = 1.1, 3.0 Hz),
2H), 4.61 (benzylic d, 1H, Jgem = 12.0 Hz), 4.54 (benzylic d, 1H, 2.64–2.50 (m, 2H), 2.04 (s, 3H), 1.24 (t, 3H, J = 7.4 Hz); 13C
Jgem = 10.5 Hz), 4.53 (benzylic d, 1H, Jgem = 10.0 Hz), 4.41 (d, NMR (125 MHz, CDCl3): δ = 170.9, 138.3, 138.1, 135.6, 133.4,
1H, J = 7.5 Hz), 4.39 (m, 2H), 4.21 (m, 1H), 4.13 (dd, 1H, J = 3.0 133.1, 128.5, 128.5, 128.2, 128.1, 128.0, 127.9, 127.8, 127.8,
Hz, J = 9.5 Hz), 4.05 (t, 1H, J = 9.5 Hz), 3.96 (dd, 1H, J = 5.0 Hz, 126.6, 126.2, 126.0, 125.9, 82.1, 80.5, 76.4, 75.2, 74.8, 72.2,
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72.1, 70.4, 63.6, 25.6, 20.9, 15.1; HRMS (ESI): [M + Na]+ calcd (benzylic d, 1H, Jgem = 12.1 Hz), 4.46 (benzylic d, 1H, Jgem
=
for C35H38NaO6S, 609.2287; found, 609.2303.
12.1 Hz), 4.35 (d, 1H, J = 7.7 Hz), 4.13 (ddd, 1H, J = 9.8 Hz, J =
4.5 Hz, J = 1.4 Hz), 4.03 (dd, 1H, J = 1.3 Hz, J = 3.4 Hz),
3.99–3.91 (m, 2H), 3.80 (dd, 1H, J = 10.9 Hz), 3.76–3.69 (m,
2H), 3.64–3.58 (m, 1H), 3.56 (t, 1H, J = 8.9 Hz), 3.42–3.38 (m,
6-O-Acetyl-2,4-di-O-benzyl-3-O-(2-naphthalenylmethyl)-1-thio-
D-mannopyranose (13)
NIS (702,6 mg, 3.12 mmol) and TFA (0.24 mL, 3.12 mmol) 1H), 3.20 (dd, 1H, J = 10.3, 11.5 Hz), 3.08 (d, 1H, J = 9.2 Hz),
were added to a vigorously stirred solution of 12 (1.83 g, 2.64–2.50 (m, 2H), 1.23 (t, 3H, J = 7.4 Hz); 13C NMR (125 MHz,
3.12 mmol) in CH2Cl2 (26 mL) and H2O (2.6 mL) at 0 °C. After CDCl3): δ = 138.8, 138.8, 138.6, 138.4, 138.2, 128.7, 128.6,
1 h, the reaction was quenched by adding 10% aq. Na2S2O3 128.4, 128.4, 128.4, 128.3, 128.1, 128.0, 128.0, 128.0, 127.9,
solution (10 mL). The reaction mixture was extracted with 127.7, 127.7, 127.6, 127.5, 104.1, 83.8, 82.7, 82.3, 81.2, 77.5,
CH2Cl2 (20 mL), the layers were separated, and the organic 77.2, 75.8, 75.1, 75.0, 73.6, 73.4, 71.7, 71.7, 69.5, 64.3, 25.3,
layer was washed with sat. NaHCO3 (aq., 20 mL), then dried 15.0; HRMS (ESI): [M + Na]+ calcd for C48H54O9NaS, 829.3386;
over MgSO4, filtered and concentrated. The crude was purified found, 829.3346.
by flash column chromatography on silica gel (cyclohexane/
Ethyl 6-O-acetyl-2,4-di-O-benzyl-3-O-(2-naphthalenylmethyl)-
EtOAc 3 : 2) to give pure 13 (1.45 g, 2.68 mmol, 86%) as 9 : 1 α:β
1
α-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-
(1→2)]-4,6-di-O-benzyl-1-thio-α-D-mannopyranoside (16) and
ethyl 6-O-acetyl-2,4-di-O-benzyl-3-O-(2-naphthalenylmethyl)-
β-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-
(1→2)]-4,6-di-O-benzyl-1-thio-α-D-mannopyranoside (17)
mixture; Rf 0.25 (cyclohexane/EtOAc 3 : 2); H NMR (500 MHz,
CDCl3 α-compound): δ = 7.84–7.70 (m, 4H, Ar-H), 7.48–7.42
(m, 3H, Ar-H), 7.38–7.22 (m, 10H, Ar-H), 5.23 (dd, 1H, J = 1.2,
3.2 Hz), 4.96 (benzylic d, 1H, Jgem = 10.9 Hz), 4.79–4.68 (m,
4H), 4.62 (benzylic d, 1H, Jgem = 10.9 Hz), 4.38 (dd, 1H, J = 1.8,
11.8 Hz), 4.25 (dd, 1H, J = 5.2, 11.8 Hz), 4.06–4.00 (m, 2H),
General procedure for 14. To a solution of 13 (266 mg,
3.97–3.93 (m, 1H), 3.84–3.82 (m, 1H), 3.14 (d, 1 H, J = 3.2 Hz), 0.5 mmol) in dry CH2Cl2 (5 mL), trichloroacetonitrile
2.02 (s, 3H); 13C NMR (125 MHz, CDCl3): δ = 171.1, 138.4, (0.295 mL, 2.99 mmol) and DBU (1.5 μL, 0.01 mmol) were
138.3, 135.9, 133.4, 133.1, 128.5, 128.5, 128.2, 128.2, 128.0, added at 0 °C under nitrogen atmosphere. After 1.5 h, the
127.9, 127.8, 127.8, 126.4, 126.2, 126.0, 125.9, 92.8, 79.9, 75.3, mixture was concentrated in vacuo and filtered through a short
74.9, 74.8, 72.8, 72.4, 70.4, 63.9, 21.0; HRMS (ESI): [M + Na]+ column of silica gel (cyclohexane/EtOAc 1 : 1) affording 14
calcd for C33H34NaO7, 565.2202; found, 565.2230.
(315 mg) which was used in the subsequent glycosylation reac-
tion without any characterisation.
Method A – To a solution of 15 (180 mg, 0.22 mmol) in dry
toluene (5 mL) powdered molecular sieves 4 Å (50 mg) and
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-
benzyl-1-thio-α-D-mannopyranoside (15)
Method A – To a solution of 9 (640 mg, 0.68 mmol) in CH2Cl2/ TMSOTf (4 μL, 0.022 mmol) were added at −20 °C under N2
H2O 10 : 1 (12 mL) DDQ (463 mg, 2.04 mmol) was added. The atmosphere. After stirring for 10 min, a solution of 14
reaction mixture was stirred at room temperature for 3.5 h and (230 mg, 0.33 mmol) in dry toluene (2 mL) was added drop
then quenched by addition of sat. Na2S2O3 (aq., 20 mL). The wise. After 1 h, the reaction was quenched by addition of Et3N
organic layer was diluted with CH2Cl2 (20 mL), washed three (1 mL). The reaction mixture was filtered through a pad of
times with sat. NaHCO3 (aq., 3 × 20 mL), dried (MgSO4), Celite, concentrated in vacuo and the crude was purified by
filtered and concentrated. The crude was purified by flash column chromatography on silica gel (toluene/EtOAc
flash column chromatography on silica gel (cyclohexane/EtOAc 9.5 : 5 → 9 : 1) to give pure 16 (194 mg, 0.15 mmol, 65%) and
9 : 1) to give pure 15 (355 mg, 0.44 mmol, 65%) as a white pure 17 (53 mg, 0.04 mmol, 18%).
foam.
Method B – To a solution of 15 (150 mg, 0.19 mmol) in dry
Method B – To a solution of 9 (136 mg, 0.14 mmol) in toluene (4 mL) powdered molecular sieves 4 Å (40 mg) and
CH2Cl2/t-BuOH 10 : 1 (2.2 mL) DDQ (65 mg, 0.28 mmol) was TMSOTf (3.5 μL, 0.019 mmol) were added at room temperature
added. The reaction mixture was stirred at room temperature under N2 atmosphere. After stirring for 10 min, a solution of
for 1.5 h and then quenched by addition of sat. Na2S2O3 (aq., 14 (191 mg, 0.27 mmol) in dry toluene (1.5 mL) was added
5 mL). The organic layer was diluted with CH2Cl2 (8 mL), drop wise. After 1 h, the reaction was quenched by addition of
washed three times with sat. NaHCO3 (aq., 3 × 8 mL), dried Et3N (0.8 mL). The reaction mixture was filtered through a pad
(MgSO4), filtered and concentrated. The crude was purified by of Celite, concentrated and the crude was purified by flash
flash column chromatography on silica gel (cyclohexane/EtOAc column chromatography on silica gel (toluene/EtOAc 9.5 : 5 →
9 : 1) to give pure 15 (87 mg, 0.11 mmol, 75%); Rf 0.12 (cyclo- 9 : 1) to give pure 16 (186 mg, 0.14 mmol, 75%) and pure 17
hexane/EtOAc 3 : 2); [α]2D0 +60.4 (c 1.05, CHCl3); 1H NMR (22 mg, 0.02 mmol, 9%).
(500 MHz, CDCl3): δ = 7.47–7.13 (m, 25H, Ar-H), 5.37 (bs, 1H,
Compound 16
H-1), 4.99 (benzylic d, 1H, Jgem = 10.5 Hz), 4.94 (benzylic d, 1H,
Jgem = 10.5 Hz), 4.90 (benzylic d, 1H, Jgem = 11.1 Hz), 4.85 Rf 0.59 (toluene/EtOAc 9 : 1); [α]2D0 +25.7 (c 1.01, CHCl3); 1H
(benzylic d, 1H, Jgem = 11.1 Hz), 4.72 (benzylic d, 1H, Jgem
=
NMR (500 MHz, CDCl3): δ = 7.80–7.72 (m, 4H, Ar-H), 7.47–7.49
11.2 Hz), 4.65 (benzylic d, 1H, Jgem = 11.2 Hz), 4.60 (benzylic d, (m, 38H, Ar-H), 5.37 (s, 1H), 5.18 (s, 1H, H), 5.08 (benzylic d,
1H, Jgem = 11.0 Hz), 4.59 (benzylic d, 1H, Jgem = 11.0 Hz), 4.53 1H, Jgem = 10.2 Hz), 4.98 (benzylic d, 1H, Jgem = 10.2 Hz), 4.87
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(benzylic d, 1H, Jgem = 11.1 Hz), 4.83 (benzylic d, 1H, Jgem
=
6 : 1). Data for the a-anomer: 1H NMR (500 MHz, CDCl3): δ =
11.1 Hz), 4.77 (benzylic d, 1H, Jgem = 11.8 Hz), 4.67 (benzylic d, 7.82–7.80 (m, 2H, Ar-H), 7.76–7.74 (m, 1H, Ar-H), 7.68 (m, 1H,
1H, Jgem = 11.8 Hz), 4.64–4.59 (m, 2H), 4.51–4.34 (m, 8H), Ar-H), 7.51 (m, 1H, Ar-H), 7.44–7.18 (m, 22H, Ar-H), 5.31 (d,
4.32–4.25 (m, 4H), 4.17–4.10 (m, 2H), 4.09–4.06 (m, 2H), 1H), 5.05 (benzylic d, 1H, Jgem = 10.5 Hz), 4.97–4.89 (m, 3H),
4.02–3.95 (m, 2H), 3.86 (t, 1H, J = 9.4 Hz), 3.74 (dd, 1H, J = 4.1, 4.83 (benzylic d, 1H, Jgem = 12.0 Hz), 4.75 (benzylic d, 1H, Jgem
10.7 Hz), 3.70 (dd, 1H, J = 1.7, 2.3 Hz), 3.63 (dd, 1H, J = 1.2, = 12.5 Hz), 4.70 (benzylic d, 1H, Jgem = 11.5 Hz), 4.59 (benzylic
10.7 Hz), 3.52 (t, 1H, J = 8.8 Hz), 3.48–3.41 (m, 2H), 3.11 (dd, d, 1H, Jgem = 12.0 Hz), 4.54–4.48 (m, 2H), 4.45–4.37 (ABq, 2H),
1H, J = 10.1, 11.6 Hz), 2.63–2.51 (m, 2H), 2.11 (s, 3H), 1.23 (t, 4.28 (d, 1H, J = 7.0 Hz), 4.09 (m, 1H), 4.07–4.00 (m, 2H), 3.90
3H, J = 7.4 Hz); 13C NMR (125 MHz, CDCl3): δ = 171.2, 139.0, (dd, 1H, J = 5.5, 12.0 Hz), 3.84 (t, 1H, J = 9.8 Hz), 3.72–3.68 (m,
138.8, 138.7, 138.7, 138.4, 138.3, 138.1, 136.1, 133.4, 133.0, 1H), 3.66–3.59 (m, 2H), 3.54 (t, 1H, J = 8.5 Hz), 3.50 (m, 1H),
129.0, 128.5, 128.4, 128.4, 128.4, 128.3, 128.2, 128.1, 128.1, 3.07–3.01 (m, 1H); 13C NMR (125 MHz, CDCl3): δ = 138.9,
128.0, 127.9, 127.8, 127.7, 127.7, 127.6, 127.6, 127.5, 127.5, 138.7, 138.6, 138.3, 137.8, 135.8, 133.3, 133.0. 129.3, 128.9,
127.5, 126.8, 126.5, 126.2, 126.0, 104.1 (JC,H = 155 Hz), 99.9 (JC, 128.4, 128.3, 128.3, 128.2, 128.0, 128.0, 127.9, 127.9, 127.8,
= 170 Hz), 84.0, 82.6 (JC,H = 165 Hz), 81.4, 80.5, 80.1, 77.6, 127.8, 127.8, 127.7, 127.6, 127.6, 127.5, 127.4, 127.0, 126.5,
H
77.0, 75.9, 75.7, 75.5, 75.4, 75.2, 75.1, 74.5, 73.5, 72.9, 72.6, 125.9, 125.8, 103.4, 92.5, 83.9, 81.4, 77.8, 77.5, 75.5, 75.1, 75.0,
72.4, 72.1, 70.6, 69.4, 64.5, 63.6, 25.4, 21.2, 15.0; HRMS (ESI): 74.9, 74.8, 73.4, 73.3, 71.5, 71.4, 69.9, 64.0. Selected data for
[M
+
Na]+ calcd for C81H86O15NaS, 1353.5585; found, the β-anomer: 13C NMR (125 MHz, CDCl3): δ 105.6, 94.6, 83.4,
1353.5609.
81.5, 80.4, 78.7, 76.1, 75.6, 75.3, 75.2, 74.1, 73.6, 73.4, 71.6,
69.3, 64.4; HRMS (ESI): [M + Na]+ calcd for C57H58O10Na,
925.3928; found, 925.3905; anal. calcd for C57H58O10: C, 75.81;
Compound 17
Rf 0.43 (toluene/EtOAc 9 : 1); [α]2D0 −2.8 (c 1.26, CHCl3); 1H H, 6.47. Found: C, 75.76; H, 6.32%.
NMR (500 MHz, CDCl3): δ = 7.80–7.65 (m, 4H, Ar-H), 7.45–7.16
(m, 38H, Ar-H), 5.42 (d, J = 2.2 Hz, 1H), 5.08 (benzylic d, 1H,
Jgem = 10.5 Hz), 5.02–4.96 (m, 3H), 4.94–4.89 (m, 2), 4.83
(benzylic d, 1H, Jgem = 11.1 Hz), 4.69 (s, 1H), 4.66–4.60 (m, 4H),
1-O-[2,3,4-Tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-benzyl-
3-O-(2-naphthalenylmethyl)-D-mannopyranosyl]
trichloroacetimidate (19)
4.52 (benzylic d, 1H, Jgem = 11.7 Hz), 4.45–4.38 (m, 5H), Trichloroacetonitrile (300 μL, 3.00 mmol) and DBU (8 μL,
4.32–4.25 (m, 2H), 4.18–4.13 (m, 2H), 4.11 (d, 1H, J = 2.8 Hz), 0.05 mmol) were added to an ice-cooled solution of 18
3.95–3.89 (m, 2H), 3.77–3.69 (m, 2H), 3.66 (dd, 1H, J = 1.8, 10.8 (455 mg, 0.50 mmol) in dry CH2Cl2 (20 mL) in an atmosphere
Hz), 3.58–3.53 (m, 2H), 3.52–3.47 (m, 2H), 3.43–3.38 (m, 1H), of nitrogen (TLC, toluene/EtOAc 5 : 1). After 60 min, the
3.14 (dd, 1H, J = 10.2, 11.4 Hz), 2.69–2.55 (m, 2H), 1.97 (s, 3H), mixture was concentrated in vacuo (water bath temperature
1.26 (t, 3H, J = 7.4 Hz); 13C NMR (125 MHz, CDCl3): δ = 171.1, of rotary evaporator: <30 °C). Purification by flash column
139.3, 138.9, 138.8, 138.5, 138.5, 138.3, 138.2, 135.6, 133.4, chromatography (SiO2, neutralised with 1% Et3N per column
133.1, 129.0, 128.6, 128.5, 128.4, 128.4, 128.3, 128.3, 128.2, volumed, 20 mL, 2.3 cm, toluene→toluene/EtOAc, 98 : 2
128.2, 128.2, 128.0, 128.0, 128.0, 127.9, 127.9, 127.8, 127.8, →95 : 5→90 : 10→85 : 15) gave an anomeric mixture of 19
127.8, 127.7, 127.7, 127.6, 127.5, 127.3, 126.3, 126.0, 125.6, (527 mg) as a colourless foam; Rf 0.71 (toluene/EtOAc 5 : 1).
103.2 (JC,H = 160 Hz), 98.8 (JC,H = 155 Hz), 83.9, 82.7, 81.9 (JC,H
Donor 19 was prepared immediately prior to use in the sub-
= 165 Hz), 81.3, 77.9, 76.8, 75.8, 75.7, 75.3, 75.0, 74.7 (2C), sequent inverse glycosylation with acceptor 20.
74.2, 74.0, 74.0, 73.9, 73.4, 73.3, 71.5, 71.3, 69.7, 64.2, 64.0,
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-6-O-acetyl-4-
O-benzyl-1-thio-α-D-mannopyranoside (20)
25.5, 21.0, 15.0 (SCH2CH3); HRMS (ESI): [M + Na]+ calcd for
C81H86O15NaS, 1353.5585; found, 1353.5565.
DDQ (242 mg, 1.06 mmol) was added to a vigorously stirred
mixture of 7 (600 mg, 0.67 mmol) in CH2Cl2/H2O (34.1 mL,
10 : 1) at 20 °C. An additional amount of DDQ (182 mg,
2,3,4-Tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-benzyl-3-O-
(2-naphthalenylmethyl)-D-mannopyranose (18)
NIS (47 mg, 0.21 mmol) and TFA (16 μL, 0.21 mmol) were 0.80 mmol) was added after 20 min. More DDQ (182 mg,
added to vigorously stirred solution of (200 mg, 0.80 mmol) was added after 40 min. The progress of the reac-
a
8
0.21 mmol) in CH2Cl2/H2O (10 mL, 20 : 1) at 20 °C (TLC, tion was carefully monitored by TLC (toluene/EtOAc 6 : 1).
toluene/EtOAc 6 : 1). After 3 h, the reaction was quenched by After 60 min, the reaction was quenched by the adding 10%
adding 10% aq. Na2S2O3-solution (10 mL). The resulting aq. Na2S2O3-solution (100 mL). The resulting mixture was
mixture was extracted once with CH2Cl2 (50 mL), the layers extracted once with CH2Cl2 (100 mL), the layers were separ-
were separated, and the organic layer was washed sequentially ated, and the organic layer was washed sequentially with sat.
with sat. NaHCO3-solution (3 × 25 mL), and brine (1 × 25 mL), NaHCO3-solution (3 × 100 mL) and brine (1 × 100 mL), dried
dried over MgSO4 and concentrated in vacuo. Purification by over MgSO4 and concentrated in vacuo. The residue was puri-
flash column chromatography (SiO2, 75 mL, 4.5 cm, toluene→ fied by flash column chromatography (SiO2, 100 mL, 4.5 cm,
toluene/EtOAc,
93 : 7→90 : 10→87 : 13→84 : 16→81 : 19 toluene→toluene/EtOAc,
96 : 4→93 : 7→90 : 10→87 : 13→
→75 : 25→70 : 30) gave an anomeric mixture of 18 (163 mg, 84 : 16→80 : 20). The first fraction to appear in the eluate con-
85%, α : β = 5 : 1) as a colourless syrup. Rf 0.16 (toluene/EtOAc tained recovered starting material (60 mg). Solvent removal of
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the second and main fraction gave 20 (355 mg, 70%) as a col- 20.6, 15.0; HRMS (ESI): [M + Na]+ calcd for C100H106O19NaS,
ourless syrup. Rf 0.45 (toluene/EtOAc 6 : 1); [α]D20 +87.0 (c 1.0, 1665.6947; found, 1665.6919; anal. calcd for C100H106O19S: C,
1
CHCl3); H NMR (500 MHz, CDCl3): δ = 7.38–7.24 (m, 20H, Ar- 73.06; H, 6.50; S, 1.95. Found: C, 72.74; H, 6.61; S, 1.92%.
H), 5.32 (s, 1H), 4.97 (m, 2H), 4.92–4.86 (m, 2H), 4.72 (m, 2H),
2-Azidoethyl 6-O-acetyl-2,4-di-O-benzyl-3-O-(2-naphthalenyl
4.62 (m, 2H), 4.35–4.29 (m, 3H), 4.19–4.17 (m, 1H), 4.02 (m,
methyl)-α-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-
1H), 3.99–3.94 (m, 2H), 3.65–3.61 (m, 2H), 3.57 (t, 1H, J = 8.5
xylopyranosyl-(1→2)]-4,6-di-O-benzyl-α-D-mannopyran-osyl-
Hz), 3.41 (t, 1H, J = 8.3 Hz), 3.21 (t, 1H, J = 10.8 Hz), 3.10 (d,
(1→3)-2,4,6-tri-O-benzyl-α-D-mannopyranoside (23)
1H, J = 9.5 Hz), 2.62–2.49 (m, 2H), 1.84 (s, 3H), 1.23 (t, 3H); 13
C
NMR (125 MHz, CDCl3): δ = 170.6, 138.6, 138.3, 138.0, 137.9, A solution of 16 (58 mg, 0.043 mmol), 22 (15 mg, 0.029 mmol)
128.6, 128.5, 128.4, 128.4, 128.1, 128.0, 127.9, 127.9, 127.9, and 4 Å powdered molecular sieves in dry Et2O (2.5 mL) was
127.8, 127.8, 127.7, 103.9, 83.6, 82.7, 82.0, 81.3, 77.4, 76.5, stirred under N2 at 20 °C. After 30 min, the solution was
75.7, 75.1, 74.8, 73.5, 71.6, 69.6, 64.2, 63.4, 25.3, 20.6, 14.9; cooled to 0 °C and DMTST (23 mg, 0.044 mmol) was added.
HRMS (ESI): [M + Na]+ calcd for C43H50O10NaS, 781.3022; The mixture was gently warmed to room temperature and
found, 781.3024; anal. calcd for C43H50O10S: C, 68.05; H, 6.64; stirred until the disappearance of 22. After 2.5 h, the reaction
S, 4.23. Found: C, 68.09; H, 6.60; S, 4.02%.
was quenched by addition of Et3N (0.8 mL), filtered through a
pad of Celite and concentrated in vacuo. The crude was puri-
fied by flash column chromatography on silica gel (toluene/
EtOAc 95 : 5→9 : 1) to give pure 23 (46 mg, 0.026 mmol, 89%)
as a colourless oil; Rf 0.44 (toluene/EtOAc 9 : 1); [α]2D0 −2.9
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-
benzyl-3-O-(2-naphthalenylmethyl)-α-D-mannopyranosyl-
(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)]-6-O-acetyl-4-
O-benzyl-1-thio-α-D-mannopyranoside (21)
1
(c 1.20, CHCl3); H NMR (600 MHz, CDCl3): δ = 7.78–7.71 (m,
A catalytic amount of TBDMSOTf (3 μL, 12 μmol) was added to 4H, Ar-H), 7.43–7.07 (m, 53H, Ar-H), 5.22 (s, 1H), 5.19 (d, J =
a solution of acceptor 20 (874 mg, 1.15 mmol) in dry toluene 1.3 Hz, 1H), 5.04 (benzylic d, 1H, J = 10.7 Hz), 4.96 (benzylic d,
(30 mL) containing crushed molecular sieves (AW-300) kept at 1H, J = 11.0 Hz), 4.90 (d, J = 1.6 Hz), 4.81–4.59 (m, 9H),
−40 °C in an atmosphere of nitrogen. A solution of donor 19 4.56–4.52 (m, 2H), 4.49 (benzylic d, 1H, J = 12.0 Hz), 4.44–4.34
(1.64 g, 1.56 mmol) in dry toluene (30 mL) was added dropwise (m, 6H), 4.29 (dd, 1H, J = 5.4, 11.9 Hz), 4.26–4.11 (m, 7H), 3.98
over 3 h to the reaction vessel using a syringe pump (rate of (t, 1H, J = 9.4 Hz), 3.94–3.80 (m, 8H), 3.79–3.72 (m, 2H), 3.67
addition: 10 mL per hour). After complete addition, the reac- (dd, 1H, J = 1.3, 10.5 Hz), 3.65–3.64 (m, 1H), 3.58–3.53 (m, 3H),
tion mixture was kept at −40 °C for 1 h (TLC, toluene/EtOAc 3.37–3.29 (m, 3H), 3.28–3.21 (m, 2H), 2.63 (t, 1H, J = 11.0 Hz),
6 : 1). The reaction mixture was neutralised with Et3N (96 μL, 1.98 (s, 3H); 13C NMR (150 MHz, CDCl3): δ = 171.1, 139.0,
0.69 mmol), the solids were removed by filtration, and the fil- 139.0, 138.9, 138.8, 138.5, 138.5, 138.4, 138.3, 138.2, 138.2,
trate was concentrated in vacuo to a yellowish oil. Purification 136.1, 133.4, 133.0, 128.8, 128.7, 128.6, 128.4, 128.4, 128.3,
by flash column chromatography (SiO2, 200 mL, 4.5 cm, tolue- 128.3, 128.2, 128.2, 128.1, 128.1, 128.0, 127.9, 127.8, 127.8,
ne→toluene/EtOAc,
127.7, 127.7, 127.7, 127.6, 127.5, 127.5, 127.5, 127.5, 127.4,
96 : 4→93 : 7→90 : 10→87 : 13→84 : 16→81 : 19) gave 21 (1.80 g, 127.4, 126.7, 126.7, 126.6, 126.2, 126.0, 126.0, 103.9 (JC,H = 155
95%) as a colourless foam; Rf 0.48 (toluene/EtOAc 9 : 1); [α]2D0 Hz), 100.3 (JC,H = 175 Hz), 99.8 (JC,H = 175 Hz), 97.9 (JC,H = 170
+3.0 (c 1.0, CHCl3); 1H NMR (500 MHz, CDCl3): δ = 7.79 (m, Hz), 83.4, 81.3, 80.0, 78.6, 78.5, 77.7, 77.5, 76.2, 76.0, 75.5,
1H, Ar-H), 7.76–7.74 (m, 1H, Ar-H), 7.70–7.66 (m, 2H, Ar-H), 75.4, 75.1 (2C), 74.9, 74.8, 74.4, 74.3, 73.6, 73.5, 72.9, 72.6,
7.49–7.14 (m, 47H, Ar-H), 5.27 (s, 1H), 5.18 (s, 1H), 5.05–4.99 72.5, 72.4, 72,4, 72.3, 70.4, 69.3, 69.1, 66.8, 64.0, 63.2, 50.5,
(m, 3H), 4.87–4.77 (m, 6H), 4.69 (benzylic d, 1H, Jgem = 11.5 21.1; HRMS (ESI): [M + Na]+ calcd for C108H113N3O21Na,
Hz), 4.64 (benzylic d, 1H, Jgem = 10.0 Hz), 4.58–4.54 (m, 3H), 1810.7764; found, 1810.7679.
4.50 (benzylic d, 1H, Jgem = 10.5 Hz), 4.46 (benzylic d, 1H, Jgem
2-Azidoethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-
= 12.0 Hz), 4.41–4.38 (m, 2H), 4.31–4.24 (m, 6H), 4.20–4.17 (m,
O-benzyl-3-O-(2-naphthalenylmethyl)-α-D-manno-pyranosyl-
2H), 4.08 (dd, 1H, J = 3.0, 9.0 Hz), 4.03 (dd, 1H, J = 3.0, 9.0 Hz),
(1→3)-[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)]-6-O-acetyl-4-
3.95–3.87 (m, 4H), 3.83–3.78 (m, 2H), 3.75 (d, 1H, J = 10.0 Hz),
O-benzyl-α-D-mannopyranosyl-(1→3)-2,4,6-tri-O-benzyl-α-D-
mannopyranoside (24)
3.65 (dd, 1H, J = 6.0, 10.5 Hz), 3.58–3.53 (ddd, 1H), 3.49–3.37
(m, 4H), 3.32 (t, 1H, J = 8.8 Hz), 3.04 (m, 1H), 2.79 (t, 1H, J =
11.0 Hz), 2.56–2.45 (m, 2H), 1.82 (s, 3H), 1.20 (t, 3H, J = 7.5 A mixture of 21 (50 mg, 30 μmol), 22 (10 μg, 20 mmol) and
Hz); 13C NMR (125 MHz, CDCl3): δ = 170.6, 139.2, 138.9, 138.8, crushed molecular sieves (4 Å) in dry Et2O (3 mL) was stirred
138.7, 138.5, 138.4, 138.1, 138.0, 135.9, 133.3, 133.0, 128.8, at 20 °C for 30 min. The reaction mixture was cooled to 0 °C,
128.6, 128.6, 128.4, 128.3, 128.3, 128.2, 128.2, 128.2, 127.9, freshly prepared DMTST (16 mg, 62 μmol) was added, and the
127.8, 127.8, 127.8, 127.8, 127.7, 127.7, 127.7, 127.6, 127.6, reaction mixture was stirred at 0 °C for 30 min. The progress of
127.5, 127.5, 127.4, 127.3, 127.1, 126.8, 126.6, 125.9, 125.8, the reaction was carefully monitored by TLC (toluene/EtOAc
103.8 (JC,H = 155 Hz), 103.7 (JC,H = 155 Hz), 100.5 (JC,H = 170 9 : 1). The cooling bath was removed, an additional amount of
Hz), 83.8, 83.5, 82.7 (JC,H = 165 Hz), 81.5, 80.9, 80.3, 78.6, 78.3, DMTST was added (16 mg, 62 mmol), and stirring was contin-
77.6, 77.3, 75.9, 75.6, 75.3, 75.0, 75.0, 74.93, 74.7 (2C), 74.4, ued at 20 °C for 3 h. The reaction was diluted with Et2O, and
73.5, 73.2, 72.5, 72.4, 72.1, 70.1, 70.0, 63.9, 63.4, 63.2, 25.5, quenched with Et3N (28 μL, 0.20 mmol) at 0 °C. The solution
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was filtered through a pad of Celite, and the filtrate was con- 4.20–4.17 (bs, 1H), 4.12–4.03 (m, 5H), 4.00 (dd, J 4.8 Hz, J 11.7
centrated in vacuo. The residue was re-dissolved in CH2Cl2 Hz, 1H), 3.79 (dd, J 4.1 Hz, J 10.8 Hz, 1H), 3.73 (dd, J 1.1 Hz,
(50 mL), and the organic layer was washed sequentially with 1 J 3.6 Hz, 1H), 3.68 (dd, J 1.3 Hz, J 10.7 Hz, 1H), 3.57–3.47 (m,
M aq. HCl-solution (3 × 25 mL), sat. NaHCO3 (2 × 25 mL), and 3H), 3.44 (t, J 8.2 Hz, 1H), 3.09 (dd, J 9.8 Hz, J 11.5 Hz, 1H),
brine (1 × 25 mL), dried over MgSO4 and concentrated in vacuo 2.63–2.51 (m, 2H), 2.28 (d, J 10.0 Hz, 1H), 2.11 (s, 3H), 1.23 (t,
to a yellow oil. Purification by flash column chromatography J 7.4 Hz, 3H); 13C NMR (125 MHz, CDCl3): δ = 171.1, 138.9,
(SiO2,
70
mL,
2.3
cm,
toluene→toluene–EtOAc 138.8, 138.6, 138.6, 138.2, 138.1, 137.7, 128.9, 128.5, 128.4,
94 : 6→93 : 7→92 : 8→91 : 9→90 : 10→ 89 : 11→88 : 12) gave 24 128.3, 128.3, 128.3, 127.9, 127.9, 127.7, 127.7, 127.7, 127.6,
(59 mg, 92%) as a colourless syrup. Rf 0.39 (toluene/EtOAc 127.6, 127.5, 127.5, 126.6, 104.1, 99.0, 83.9, 82.5, 81.1, 80.6,
1
9 : 1); [α]2D0 + 1.3 (c 1.0, CHCl3); H NMR (600 MHz, CDCl3): δ = 78.9, 77.9, 77.7, 77.1, 75.6, 75.4, 75.1, 74.7, 74.5, 73.4, 72.7,
7.77–7.74 (m, 2H, Ar-H), 7.68–7.66 (m, 2H, Ar-H), 7.47–7.45 (m, 72.3, 72.1, 71.7, 69.4, 69.4, 64.5, 63.6, 25.3, 21.2, 15.0. HRMS
1H, Ar-H), 7.42–7.06 (m, 62H, Ar-H), 5.22 (s, 1H), 5.11 (s, 1H), (ESI): [M + Na]+ calcd for C70H78O15NaS, 1213.4959; found,
5.05 (benzylic d, 1H, Jgem = 11.1 Hz), 4.98–4.97 (m, 3H), 1213.4962.
4.82–4.73 (m, 7H), 4.69–4.64 (m, 3H), 4.61 (benzylic d, 1H,
Jgem = 12.1 Hz), 4.56–4.49 (m, 5H), 4.45 (benzylic d, 1H, Jgem
=
11.9 Hz), 4.31–4.26 (m, 4H), 4.19–4.07 (m, 7H), 4.04 (dd, 1H,
J = 3.1, 9.5 Hz), 3.98–3.95 (m, 2H), 3.91 (d, 1H, J = 7.3 Hz),
3.89–3.86 (m, 2H), 3.83–3.78 (m, 5H), 3.76–3.70 (m, 3H),
3.67–3.59 (m, 3H), 3.58–3.49 (m, 3H), 3.44–3.40 (ddd, 1H,
Ethyl 2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)-4,6-di-O-
benzyl-α-D-mannopyranosyl-(1→3)-[2,3,4-tri-O-benzyl-β-D-
xylopyranosyl-(1→2)]-6-O-acetyl-4-O-benzyl-1-thio-α-D-
mannopyranoside (26)
OCH2CHa N3), 3.38–3.30 (m, 3H), 3.29–3.22 (m, 3H), 2.65–2.58
2
(m, 2H), 1.76 (s, 3H); 13C NMR (150 MHz, CDCl3): δ = 170.6, DDQ (142 mg, 0.62 mmol) was added to a vigorously stirred
139.2, 138.9, 138.8, 138.5, 138.5, 138.3, 138.3, 138.2, 138.1, solution of compound 21 (640 mg, 0.38 mmol) in CH2Cl2/H2O
138.1, 137.7, 135.9, 133.2, 132.9, 128.6, 128.5, 128.5, 128.5, (33 mL, 10 : 1) at 20 °C. Additional amounts of DDQ (106 mg,
128.5, 128.5, 128.3, 128.2, 128.2, 128.2, 128.2, 128.0, 127.9, 0.46 mmol) were added after 20 min. More DDQ (106 mg,
127.9, 127.8, 127.7, 127.7, 127.6, 127.5, 127.5, 127.5, 127.4, 0.46 mmol) was added after 40 min. The progress of the reac-
127.4, 127.4, 127.3, 127.3, 127.3, 127.2, 126.7, 126.4, 125.9, tion was carefully monitored by TLC (toluene–EtOAc, 6 : 1).
125.8, 103.6 (JC,H = 160 Hz), 103.5 (JC,H = 160 Hz), 100.5 (JC,H
=
After 60 min, the reaction was quenched by adding 10% aq.
175 Hz), 99.8 (JC,H = 170 Hz), 97.6 (JC,H = 170 Hz), 83.2, 83.2, Na2S2O3-solution (100 mL). The resulting mixture was extracted
81.5, 80.7, 79.6, 78.2, 78.2, 77.7, 77.5, 77.2, 76.1, 75.9, 75.4, once with CH2Cl2 (200 mL), the layers were separated, and the
75.2, 74.9, 74.8 (2C), 74.6, 74.5, 74.5, 74.3, 74.0, 73.4, 73.2, organic layer was washed sequentially with sat. NaHCO3-solu-
73.1, 72.5, 72.4, 72.3, 72.3, 72.1, 70.0, 69.5, 69.1, 66.6, 63.7, tion (3 × 150 mL), and brine (1 × 150 mL), dried over MgSO4
63.2, 63.0, 50.4, 20.5; HRMS (ESI): [M + Na]+ calcd for and concentrated in vacuo to a pale yellow oil. The residue was
C
C
127H133N3O25Na, 2122.9126; found, 2122.9197; anal. calcd for purified by flash column chromatography (toluene→toluene/
127H133N3O25: C, 72.59; H, 6.38; N, 2.00. Found: C, 72.99; H, EtOAc, 96 : 4→93 : 7→90 : 10→87 : 13→84 : 16→81 : 19 →78 : 22).
6.74; N, 1.69%.
The first fraction to appear in the eluate contained recovered
starting material (67 mg). Solvent removal of the second and
main fraction gave 26 (396 mg, 68%) as a colourless foam. Rf
Ethyl 6-O-acetyl-2,4-di-O-benzyl-α-D-mannopyranosyl-(1→3)-
[2,3,4-tri-O-benzyl-β-D-xylopyranosyl-(1→2)]-4,6-di-O-benzyl-1-
thio-α-D-mannopyranoside (25)
1
0.15 (toluene–EtOAc, 9/1); [α]2D0 +38.0 (c 1.00, CHCl3); H NMR
(500 MHz, CDCl3) δ 7.41–7.13 (m, 45H), 5.27 (s, 1H), 5.16 (s,
DDQ (52 mg, 0.23 mmol) was added to a vigorously stirred 1H), 5.02 (d, J 10.0 Hz, 1H), 4.97 (d, J 11.3 Hz, 1H), 4.92 (d,
solution of compound 16 (158 mg, 0.12 mmol) in CH2Cl2/ J 10.8 Hz, 1H), 4.89–4.80 (m, 5H), 4.70 (d, J 11.8 Hz, 1H), 4.65
t-BuOH (20 mL, 10 : 1) at 20 °C. The progress of the reaction (d, J 10.0 Hz, 1H), 4.62–4.48 (m, 6H), 4.35 (d, J 11.5 Hz, 1H),
was carefully monitored by TLC (toluene–EtOAc, 85 : 15). After 4.33–4.26 (m, 5H), 4.23–4.17 (m, 2H), 4.07–4.01 (m, 2H),
1.5 h, the reaction was quenched by adding 10% aq. Na2S2O3- 3.97–3.90 (m, 2H), 3.82–3.74 (m, 4H), 3.66 (dd, J 5.8 Hz, J 10.6
solution (10 mL). The resulting mixture was extracted once Hz, 1H), 3.58–3.35 (m, 6H), 3.29–3.25 (m, 1H), 3.06–2.97
with CH2Cl2 (20 mL), the layers were separated, and the (m, 2H), 2.86 (t, J 11.0 Hz, 1H), 2.55–2.44 (m, 2H), 1.83 (s, 3H),
organic layer was washed sequentially with sat. NaHCO3-solu- 1.20 (t,
J
7.4 Hz, 1H); 13C NMR (126 MHz, CDCl3)
tion (3 × 20 mL), and brine (1 × 20 mL), dried over MgSO4 and δ 170.7, 139.3, 139.0, 138.8, 138.7, 138.5, 138.3 (2C), 138.2
concentrated in vacuo to a pale yellow oil. The residue was puri- (2C), 128.8, 128.8, 128.7, 128.6, 128.4, 128.4, 128.4, 128.4,
fied by flash column chromatography on silica gel (toluene– 128.3, 128.1, 128.0, 128.0, 127.9, 127.8, 127.8, 127.7,
EtOAc, 96 : 4→70 : 30) and afforded 25 (104 mg, 72%) as a col- 127.6, 127.6, 127.5, 127.5, 126.9, 104.3, 104.0, 101.1, 83.9,
ourless foam. Rf 0.59 (toluene/EtOAc 85 : 15); [α]2D0 +36.0 83.4, 82.9, 81.5, 80.9, 80.8, 80.7, 78.9, 77.8, 77.4, 77.3,
(c 1.00, CHCl3);1H NMR (500 MHz, CDCl3): δ = 7.43–7.11 (m, 75.6 (2C), 75.1, 74.9, 74.9, 74.7 (2C), 73.6, 73.5, 72.8, 71.9,
35H), 5.39 (s, 1H), 5.25 (s, 1H), 5.07 (d, J 10.1 Hz, 1H), 4.93 (d, 70.9, 70.3, 69.9, 64.1, 63.6, 63.3, 25.7, 20.7, 15.1. HRMS
J 11.2 Hz, 1H), 4.87–4.78 (m, 3H), 4.68 (d, J 11.8 Hz, 1H), (ESI): [M + Na]+ calcd for C89H98O19NaS, 1525.6321; found,
4.60–4.55 (m, 2H), 4.47–4.38 (m, 3H), 4.36–4.22 (m, 6H), 1525.6383.
6608 | Org. Biomol. Chem., 2015, 13, 6598–6610
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Paper
2-Aminoethyl 6-O-acetyl-α-D-mannopyranosyl-(1→3)-
[β-D-xylopyranosyl-(1→2)]-α-D-mannopyranosyl-(1→3)-
α-D-mannopyranoside (27)
Notes and references
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10% w Pd/C (13 mg, 12.3 mmol) was added to a solution of
compound 23 (20 mg, 11.2 mmol) in AcOEt/H2O/AcOH
(4 : 2 : 1, 1.75 mL). The mixture was hydrogenolysed in a high-
pressure reactor (Berghof) at 20 °C (p = 30 bar). After 60 h, the
solids were removed by filtration using a ‘sandwich filter’ (3
frits stacked on top of each other in the following order:
20 μm, 10 μm, 5 μm), rinsed with H2O (3 × 2 mL) and EtOH (3
× 2 mL), and the filtrate was concentrated in vacuo. Purification
by reversed-phase chromatography (C-18, H2O–MeOH,
9 : 1→8 : 2→7 : 3→6 : 4→2 : 8→0 : 10), followed by freeze-drying
gave 27 (5.8 mg, 72%) as a colourless, amorphous solid. [α]D20
1
+26.2 (c 0.45, H2O); H NMR (600 MHz, D2O): δ = 5.12 (s, 1H),
4.95 (s, 1H), 4.74 (s, 1H), 4.33 (d, J 11 Hz, 1H), 4.22 (d, J 8 Hz,
1H), 4.11 (dd, J 6 Hz, 12 Hz, 1H), 4.08–4.02 (m, 1H), 3.99 (s,
2H), 3.93 (1H, s), 3.88 (dd, J 3 Hz, 9 Hz, 1H), 3.86–3.81 (m,
3H), 3.76–3.71 (m, 2H), 3.71–3.54 (m, 8H), 3.54–3.45 (m, 2H),
3.26 (t, J 9 Hz, 1H), 3.18–3.04 (m, 4H), 2.02 (s, 3H); 13C anome-
ric signals taken from HSQC: 103.2, 103.0, 100.0, 99.8. HRMS
(ESI): [M + Na]+ calcd for C27H47NO21Na, 744.2538; found,
744.2559.
2-Aminoethyl β-D-xylopyranosyl-(1→2)-α-D-manno-pyranosyl-
(1→3)-[β-D-xylopyranosyl-(1→2)]-6-O-acetyl-α-D-
mannopyranosyl-(1→3)-α-D-mannopyranoside (28)
10% w Pd/C (17 mg, 16.1 mmol) was added to a solution of
compound 24 (26 mg, 12.4 mmol) in AcOEt/H2O/AcOH
(4 : 2 : 1, 1.75 mL). The mixture was hydrogenolysed in a high-
pressure reactor (Berghof) at 20 °C (p = 30 bar). After 60 h, the
solids were removed by filtration using a ‘sandwich filter’
(3 frits stacked on top of each other in the following order:
20 μm, 10 μm, 5 μm), rinsed with H2O (3 × 2 mL) and EtOH
(3 × 2 mL), and the filtrate was concentrated in vacuo. Purifi-
cation by reversed-phase chromatography (C-18, H2O-MeOH,
9 : 1→8 : 2→7 : 3→6 : 4→2 : 8→0 : 10), followed by freeze-drying
gave 28 (7.3 mg, 69%) as a colourless, amorphous solid.
1
[α]2D0 +17.6 (c 0.40, H2O); H NMR (600 MHz, D2O): δ = 5.09 (s,
1H), 5.04 (s, 1H), 4.76 (s, 1H), 4.32–4.21 (m, 4H), 4.11 (1H, s),
4.01 (1H, s), 3.99–3.96 (m, 2H), 3.93–3.87 (m, 1H), 3.87–3.56
(m, 13H), 3.55–3.45 (m, 4H), 3.32–3.22 (m, 2H), 3.21–3.07 (m,
6H), 1.99 (s, 3H); 13C anomeric signals taken from HSQC:
102.9, 102.3, 100.5, 99.9, 99.4. HRMS (ESI): [M + Na]+ calcd for
C32H55NO25Na, 876.2961; found, 876.2949.
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Acknowledgements
We thank Science Foundation Ireland (Grants 08/IN.1/B2067
and 13/IA/1959) and Marie Curie-Intra-European Fellowships 24 S. David and S. Hanessian, Tetrahedron, 1985, 41, 643–663.
(FP7-PEOPLE-2011-IEF, project number 299710) for financial 25 Z.-H. Jiang and R. R. Schmidt, Liebigs Ann. Chem, 1992,
support.
975–982.
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Paper
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