Combinatorial ionic catch-and-release oligosaccharide synthesis (combi-ICROS)

Article abstract of DOI:10.1039/c2cc37164b

A combinatorial ionic-liquid-supported catch-and-release strategy for oligosaccharide synthesis (combi-ICROS) is reported. A series of β-(1→6) glucan di-, tri- and tetra-saccharides were synthesized in one pot to showcase the versatility of the strategy.

Full text of DOI:10.1039/c2cc37164b

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Combinatorial ionic catch-and-release oligosaccharide
synthesis (combi-ICROS)†‡
Cite this: Chem. Commun., 2013,
49, 4217
Imke Sittel, Anh-Tuan Tran, David Benito-Alifonso and M. Carmen Galan*
Received 1st October 2012,
Accepted 3rd January 2013
DOI: 10.1039/c2cc37164b
www.rsc.org/chemcomm
A combinatorial ionic-liquid-supported ‘‘catch-and-release’’ strat- synthetic and enzymatic applications.6–11 The unique physical
egy for oligosaccharide synthesis (combi-ICROS) is reported. A and chemical properties of ILs, which can be tuned by altering
series of b-(1-6) glucan di-, tri- and tetra-saccharides were synthe- the cation or the anion structure, make this class of molecules
sized in one pot to showcase the versatility of the strategy.
particularly useful as new vehicles for the immobilisation of
reagents.12
Methodologies that can provide rapid access to structurally
More recently, strategies based on the use of ILs as soluble
defined oligosaccharides for biological screening are greatly functional supports, termed ITags, applied to oligosaccharide
lacking.1,2 The challenges associated with carbohydrate syn- synthesis have been developed.13–19 The IL-based approaches
thesis, including laborious protecting group manipulation, the have shown great promise as they combine the ability to carry
need for regio- and stereoselective glycosylation reactions and out reactions in solution with the additional advantage of fast,
most notably the requirement for purification after each step, chromatography-free purification. Ionic-liquid-labeled compounds
normally accomplished by chromatography, are primarily (ITag-compounds) are soluble in polar solvents used in glycosyla-
responsible for the lack of more intensive efforts. Combi- tion reactions (i.e. dichloromethane, acetonitrile) but in the
natorial chemistry has become an important tool in modern absence of the polar solvent, the ITag-materials become insoluble
drug development and although carbohydrate-based compounds in less polar solvents such as diethyl ether, isopropyl ether or
hold great potential as therapeutic agents due to their involve- hexane, which means that non-ITag-materials (excess reagents,
ment in a myriad of physiological and pathological processes,3 unreacted material) can be removed from the ITag-products by
the application of combinatorial chemistry to this class of simple biphasic extraction or by precipitation.
biomolecules remains limited. Nature offers the best example
Our group has recently reported the first example of ionic-
of combinatorial chemistry, where glycan chains are assembled liquid-supported ‘‘catch-and-release’’ oligosaccharide synthesis
by glycosyltransferases with exquisite regio- and stereocontrol, (ICROS), where the ionic-liquid-purification labels (ITags) are
resulting in remarkably heterogeneous glycoforms.4
introduced at the anomeric position of the reducing end of the
Herein, we present a novel ionic-liquid based ‘‘catch-and- oligosaccharide target.13,14,20 The methodology was shown
release’’ strategy for the one-pot combinatorial synthesis of to be compatible with both chemical and enzymatic processes
oligosaccharides. This method is based on the use of covalently (Fig. 1).
attached ionic-liquid tags (ITags) as soluble supports that allows
fast, chromatography-free product purification and in situ reac-
tion monitoring. We demonstrate the versatility and generality of
this technique in the synthesis of a series of b-(1-6) glucans,
which are biologically relevant fungal cell-wall components.5
Ionic liquids (ILs) have attracted enormous interest over the
past few years because of their use in a broad number of
School of Chemistry, University of Bristol, Cantock’s Close, Bristol, BS8 1TS, UK.
E-mail: M.C.Galan@bristol.ac.uk; Fax: +44 (0)1179298611; Tel: +44 (0)1179287654
† This article is part of the ChemComm ‘Emerging Investigators 2013’ themed
issue.
‡ Electronic supplementary information (ESI) available: Experimental proce-
dures, characterization data for new compounds and NMR traces. See DOI:
10.1039/c2cc37164b
Fig. 1 ICROS methodology.
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Scheme 2 One-pot combinatorial synthesis of b-(1-6) glucans.
Encouraged by these results and having shown that purifica-
tion of ITag-products from non-ITag-materials is fast and simple,
we decided to apply this methodology to the one-pot combina-
torial synthesis of a series of b-(1-6) glucans (Scheme 2).
To that end, glycosylation of glucoside acceptor 2 with 1 in
the presence of 0.2 equiv. of TMSOTf in DCM was monitored by
MALDI-TOF, HPLC and 1H-NMR until a 1 : 2 ratio of ITag-
compounds 2 ([M+] 677.2; Rt = 2.8 min; H-1 d 5.05 ppm, J 8.1 Hz)
and 3 ([M+] 1307.5; Rt = 16.8 min; H-1A d 5.00 ppm, J 7.9 Hz and
H-1B d 4.90 ppm, J 8.0 Hz) was obtained.24 The solvent was then
removed and the non-ITag materials were washed away with a
diethyl ether–hexane (1/1, v/v) solvent mixture. Selective O-TIPS
removal with HCl in MeOH afforded a 1 : 2 mixture of 2 and 4
([M+] 1151.4; Rt = 5.5 min; H-1A d 4.92 ppm, J 8.0 Hz and H-1B d
4.73 ppm, J 8.0 Hz), which after purification by simple non-
polar solvent washes was ready to be further glycosylated.
The reaction sequence of glycosylation with 1, 6-OH selective
deprotection and solvent washes was performed with careful
monitoring by MALDI-TOF and HPLC for another two cycles
(see Fig. 3) until the final mixture contained a 1 : 5 : 3 ratio of
disaccharide 4, trisaccharide 6 ([M+] 1625.5; Rt = 12.4 min; H-1A
d 4.88 ppm, J 7.9 Hz, H-1B d 4.98 ppm, J 8.0 Hz and H-1C d 4.99
ppm, J 7.9 Hz) and tetrasaccharide 8 ([M+] 2099.6, Rt = 15.2 min;
H-1A d 4.89 ppm, J 8.0 Hz, H-1B d 4.96 ppm, J 7.7 Hz, H-1C d 5.01
ppm, J 8.1 Hz and H-1D d 5.04 ppm, J 8.1 Hz) as the major
components (as confirmed by NMR).24
Scheme 1 Sequential ICROS.
A key feature of ICROS, in addition to the fast and chroma-
tography-free purification, is that this strategy allows for simple
and in situ reaction progress monitoring by mass spectroscopy
(MS), HPLC and NMR, which is a great advantage over other
traditional supported methodologies.21–23
Based on our previous investigations, we envisioned that
ICROS would be ideally suited for the combinatorial synthesis
of oligosaccharides in one pot.
In order to test our hypothesis, a series of b-(1-6) glucan
linear mono-, di- and trisaccharides 2–5 were synthesized sequen-
tially by ICROS and individually purified and characterized.
In this instance a benzyl derived IL-tag (ITag)15 was chosen as
the cleavable purification label (Scheme 1).
ITagged-glycoside acceptor 2, which was prepared in 2 steps
from 1 in 84% yield,15 was glycosylated with trichloroacetimidate
glycosyl donor 1 in the presence of TMSOTf (0.3 equiv.) affording
disaccharide 315 in 94% yield, exclusively as the b-anomer due to the
presence of the acyl group at C-2, which ensured neighbouring
group participation. Selective 6-OH unmasking of 3 by O-TIPS
removal using a mixture of HCl in MeOH provided pure acceptor
4 in 97% yield after simple washes. Subsequent glycosylation with 2
under the same catalytic conditions as before afforded trisaccharide
5 in 94% yield. Each individual compound was purified at each
stage by simple phase extraction and fully characterized by NMR
and MS (see ESI‡ for details). Compounds 2–5 were subjected to
HPLC analysis and it was determined that the retention time (Rt) of
the different species, while still covalently linked to the IL, was
sufficiently different to be individually monitored by HPLC (Fig. 2).
The different size oligomers were separated at the final stage
of the synthesis using size-exclusion chromatography. Thus,
the mixture of compounds was subjected to purification on a
Sephadex LH-20, and targets 4, 6 and 8 were isolated in 10%,
30% and 14% yield, respectively, from starting material 2.
Fig. 3 Typical HPLC traces of
a
one-pot b-(1-6)-glucan reaction mixture
Fig. 2 HPLC traces of individual ITag-oligosaccharides.
4218 Chem. Commun., 2013, 49, 4217--4219
between glycosylation cycles.
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purification in between steps. Efforts are currently underway to
apply the ICROS methodology to more complex targets.
We gratefully acknowledge financial support from EPSRC
and MCG thanks The Royal Society DHF and EPSRC CAF for her
fellowships.
Notes and references
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Scheme 3 Deconvolution of an oligosaccharide mixture.
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Global deprotection of the ester groups was accomplished
using NaOMe in MeOH followed by cleavage of the ionic-liquid-
component in the presence of Pd/C in a mixture of water–methanol
and 5% HClaq to afford the hemiacetals 10, 12 and 14 in yields of
85–95% over 2 steps (Scheme 3).
6 J. P. Hallett and T. Welton, Chem. Rev., 2011, 111, 3508.
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The use of a benzyl type linker as a cleavable functionality
between the glycoside and the ionic liquid provides the ITag with
enough chemical stability to withstand a wide range of chemical
conditions, while product release remains efficient and in a form
suitable for further oligosaccharide manipulation.
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advantages over other supported methods: (a) purification is
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sylation reactions are performed under conditions typically used
for solution-phase chemistry and reaction progress can be moni-
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oligosaccharide elongation process.
We have shown here that ICROS is ideally suited for the
combinatorial synthesis of small libraries of oligosaccharides
and we exemplified this strategy in the preparation of a series of
b-1,6-glucan oligosaccharides in one pot. Moreover, we demon-
strated that HPLC in combination with MALDI-TOF and NMR
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This journal is The Royal Society of Chemistry 2013
Chem. Commun., 2013, 49, 4217--4219 4219