Monosaccharides as Versatile Units for Water-Soluble Supramolecular Polymers

Article abstract of DOI:10.1002/chem.201504762

We introduce monosaccharides as versatile water-soluble units to compatibilise supramolecular polymers based on the benzene-1,3,5-tricarboxamide (BTA) moiety with water. A library of monosaccharide-based BTAs is evaluated, varying the length of the alkyl chain (hexyl, octyl, decyl and dodecyl) separating the BTA and saccharide units, as well as the saccharide units (α-glucose, β-glucose, α-mannose and α-galactose). In all cases, the monosaccharides impart excellent water compatibility. The length of the alkyl chain is the determining factor to obtain either long, one-dimensional supramolecular polymers (dodecyl spacer), small aggregates (decyl spacer) or molecularly dissolved (octyl and hexyl) BTAs in water. For the BTAs comprising a dodecyl spacer, our results suggest that a cooperative self-assembly process is operative and that the introduction of different monosaccharides does not significantly change the self- assembly behaviour. Finally, we investigate the potential of post-assembly functionalisation of the formed supramolecular polymers by taking advantage of dynamic covalent bond formation between the monosaccharides and benzoxaboroles. We observe that the supramolecular polymers readily react with a fluorescent benzoxaborole derivative permitting imaging of these dynamic complexes by confocal fluorescence microscopy.

Full text of DOI:10.1002/chem.201504762

DOI: 10.1002/chem.201504762
Full Paper
&
Supramolecular Chemistry
Monosaccharides as Versatile Units for Water-Soluble
Supramolecular Polymers
Christianus M. A. Leenders, Gijs Jansen, Martijn M. M. Frissen, RenØ P. M. Lafleur, Ilja K. Voets,
[a]
Anja R. A. Palmans,* and E. W. Meijer*
Abstract: We introduce monosaccharides as versatile water-
soluble units to compatibilise supramolecular polymers
based on the benzene-1,3,5-tricarboxamide (BTA) moiety
with water. A library of monosaccharide-based BTAs is evalu-
ated, varying the length of the alkyl chain (hexyl, octyl, decyl
and dodecyl) separating the BTA and saccharide units, as
well as the saccharide units (a-glucose, b-glucose, a-man-
nose and a-galactose). In all cases, the monosaccharides
impart excellent water compatibility. The length of the alkyl
chain is the determining factor to obtain either long, one-di-
mensional supramolecular polymers (dodecyl spacer), small
aggregates (decyl spacer) or molecularly dissolved (octyl and
hexyl) BTAs in water. For the BTAs comprising a dodecyl
spacer, our results suggest that a cooperative self-assembly
process is operative and that the introduction of different
monosaccharides does not significantly change the self-
assembly behaviour. Finally, we investigate the potential of
post-assembly functionalisation of the formed supramolec-
ular polymers by taking advantage of dynamic covalent
bond formation between the monosaccharides and benzox-
aboroles. We observe that the supramolecular polymers
readily react with a fluorescent benzoxaborole derivative
permitting imaging of these dynamic complexes by confocal
fluorescence microscopy.
[
22–24]
Introduction
dynamic covalent bonds with boronic acids.
Researchers
have relied on this dynamic covalent bond formation for the
[
25,26]
Saccharide-based amphiphiles, in nature typically found in
development of synthetic lectin mimics,
and dynamic
responsive materi-
[1,2]
[25,27]
membranes as structural or recognition units,
have become
systems with applications in sensing,
[
28–34]
[35–38]
[39]
an important class within the fields of soft matter and materi-
als science because of their excellent water-solubility, biocom-
patibility and the potential for saccharide recognition. Numer-
ous saccharide-based amphiphiles have been synthesised and
als,
drug delivery,
and separation techniques.
Benzoxaborole is of particular interest, because it forms
dynamic covalent bonds with the pyranose form of mono-
[
34,40–44]
saccharides.
[
3–7]
extensively studied, both in bulk and in aqueous solution.
Alkyl glycosides are an appealing subclass because of their
In our previous work, we showed that benzene-1,3,5-
tricarboxamide (BTA) derivatives connected to water-soluble
tetraethylene glycol units through a dodecyl spacer form
responsive supramolecular polymers in water (Figure 1, com-
[8]
straightforward molecular structure. Changes in the relative
size of the alkyl- or saccharide part influence the aggregation
[9]
[45]
in line with the theory of Israelachvili, and so does changing
the spatial orientation of the saccharide head group by modi-
pound 5). The mechanism of exchange of monomers be-
tween different supramolecular polymers was studied in detail
[10,11]
[46]
fying the anomeric centre.
Introducing different epimers
by super resolution fluorescence microscopy, and a detailed
also influences the aggregation behaviour, which is attributed
computational study showed the importance of the formation
of a local hydrophobic domain and suggested stabilisation
[
12–14]
to differences in hydrogen bonding of the alcohol units.
[
47]
The natural role of saccharides as recognition units through
selective binding by lectins provides a strategy for the devel-
opment of responsive systems or post-assembly functional-
through intermolecular hydrogen bond formation. Recently,
the presence of intermolecular hydrogen bonds stabilising the
supramolecular polymers in water was experimentally verified
[
15–21]
[48]
isation.
Furthermore, saccharides are well known to form
by infrared spectroscopy.
Here, we explore the application of monosaccharides as
versatile water-solubilising groups to create BTA-based water-
soluble supramolecular polymers. Two important aspects of
the molecular design are evaluated with respect to the self-as-
sembly behaviour. First, the role of the spacer length is studied
by introducing a dodecyl, decyl, octyl or hexyl spacer connect-
ed to b-d-glucopyranoside (Figure 1, compounds 1a–d). In
a second series the aliphatic spacer is kept constant and either
b-d-glucopyranoside, a-d-glucopyranoside, a-d-galactopyrano-
[
a] Dr. C. M. A. Leenders, G. Jansen, M. M. M. Frissen, R. P. M. Lafleur,
Dr. I. K. Voets, Dr. A. R. A. Palmans, Prof. Dr. E. W. Meijer
Institute for Complex Molecular Systems
Eindhoven University of Technology
P.O. Box 513, 5600 MB, Eindhoven (The Netherlands)
E-mail: A.Palmans@tue.nl
E.W.Meijer@tue.nl
Supporting information and ORCID from the author for this article is avail-
able on the WWW under http://dx.doi.org/10.1002/chem.201504762.
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Scheme 1. Synthesis of BTAs 1a–d.
Figure 1. Chemical structure of BTAs 1–5.
1
13
fully characterised by H NMR-, C NMR- and IR spectroscopy
and MALDI-TOF-MS (see the Supporting Information).
The synthesis of BTA derivatives 2–4 was achieved by
coupling 8a to the corresponding alkyne-functionalised mono-
saccharide by Cu -catalysed azide–alkyne cycloaddition
side, or a-d-mannopyranoside is introduced. This allows us to
assess the effect of changing the anomeric centre (1a vs. 2)
and the effect of changing between different epimers (1a vs. 3
and 4) on the self-assembly behaviour. The monosaccharide is
I
(Scheme 2). Workup by reversed phase column chromatogra-
phy (which was also applied to 1a) effectively removed traces
of salts. BTAs 2–4 were obtained as white solids and were fully
I
connected to the BTA moiety by a Cu-catalysed azide–alkyne
1
13
cycloaddition because of ease of synthesis and functional
group tolerance. The self-assembly of the monosaccharide-
characterised by H NMR-, C NMR-, IR spectroscopy and
MALDI-TOF MS (see the Supporting Information).
1
based BTAs is studied by H NMR spectroscopy, UV/Vis, and
fluorescence spectroscopy, as well as by cryogenic transmission
electron microscopy (cryoTEM) and small angle X-ray scattering
(
SAXS). Our results show that a dodecyl spacer is required to
form supramolecular polymers, and these polymers can be
targeted by benzoxaborole through dynamic covalent bond
formation. Interestingly, the introduction of different mono-
saccharides does not seem to significantly influence the
self-assembly behaviour of the BTAs in water.
Results and Discussion
Scheme 2. Synthesis of BTAs 2–4.
Synthesis of compounds 1–4
The synthetic route towards b-d-glucopyranoside-based BTAs
Exploring the aqueous self-assembly behaviour of 1a–d
1
a–d is shown in Scheme 1. The a,w-dibromoalkanes of re-
quired length were converted into N-(w-azidoalkyl)phthali-
mides 6a–d in a statistic one-pot procedure. The Gabriel syn-
thesis was completed by treating 6a–d with hydrazine mono-
hydrate affording the w-azidoalkyl-a-amines 7a–d in overall
yields of 30–40%. Subsequently, treatment of trimesoyl chlo-
ride with three equivalents of 7a–d provided azide-functional-
ised benzene-1,3,5-tricarboxamides 8a-d in yields of 51–88%.
The acetyl-protected glucose functionalised BTAs 9a–d were
The aggregation behaviour of 1a–d in D O was first assessed
2
1
À3
by H NMR spectroscopy at room temperature (c=1”10 m)
(Figure 2). All samples were heated to 908C during sample
preparation and allowed to equilibrate overnight at room tem-
perature, which promotes HÀD exchange of the amides and al-
cohols. The sharpness of the resonances differs significantly in
the NMR spectra of the different compounds. Whereas the
spectrum of 1c,d shows sharp signals, only one of the aromat-
ic signals is visible in the spectrum of 1b, and all remaining
signals are relatively broad. In the spectrum of 1a, all signals
corresponding to the BTA have disappeared and only the sol-
vent peaks remain visible. These observations suggest that 1d
and 1c do not form aggregates, whereas the absence of
I
produced in high yields by Cu-catalysed cycloaddition of 8a–d
with 2-propynyl-tetra-O-acetyl-b-d-glucopyranoside. Saponifica-
tion of 9a–d with sodium hydroxide resulted in the desired
glucose-functionalised BTAs 1a–d in near quantitative yields.
Compounds 1a–d were obtained as white solids and were
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angle between the amide and central benzene group is typi-
[
49]
cally between 35 and 458. This arrangement of the hydrogen
bonds is reflected by an absorption maximum at 193 nm in
the UV spectrum. In contrast, when these aliphatic BTAs are
molecularly dissolved, the absorption maximum shifts to
[
49,50]
2
08 nm.
As a result, the single absorption maximum at
2
07 nm in the case of 1b and 1c, and 208 nm for 1d suggests
that these BTAs are molecularly dissolved in water. For 1a, the
absorption maximum at 211 nm with a shoulder at around
2
25 nm is very similar to the UV/Vis spectrum we previously
1
À3
Figure 2. H NMR spectra of 1a–d (c=1”10 m) in D
2
O at 208C. The peaks
[45]
observed for aggregates of 5 in water. Recent IR measure-
ments unambiguously show that aggregates formed by 5 are
stabilised by intermolecular hydrogen bonds in water.
at d=7.9 and 8.1 ppm were assigned by 1D ROESY NMR spectroscopy to
the triazole and aromatic core of the BTA, respectively (the Supporting
Information, Figure S1).
[
48]
In
analogy, we believe that also the aggregates formed by 1a are
stabilised by hydrogen bonds. Interestingly, the redshift in the
UV spectra observed for both 5 and 1a in water indicates that
the packing of the BTAs in the aggregated state is different
from those observed for the aliphatic BTA derivatives. We spec-
ulate that either the angle of the amide group with respect to
the central benzene ring is different or that there is an offset
between BTAs within the aggregate resulting in a J-type
a signal for the aromatic core and the broader remaining sig-
nals suggest that 1b does form aggregates. In the spectrum of
1
a all signals are lost, indicative of the formation of large
aggregates.
To study the aggregation behaviour of 1a–d in more detail,
UV/Vis spectroscopy measurements were performed. Samples
were prepared by injection of a concentrated solution in meth-
aggregate. Such offsets have been reported in several crystal
À3
À5
[51]
anol (5 mL, c=5”10 m) into water (2.5 mL, c=1”10 m) and
spectra were recorded at 208C after equilibration of the
samples overnight (Figure 3A). The UV spectra of BTAs 1c and
structures for BTA derivatives.
Although in both cases a
redshift in the UV spectrum is expected, the details of the
hydrogen-bond arrangement in aggregates of 1a and 5 are
not yet fully understood.
1
d perfectly overlap and show an absorption maximum at
2
07 nm. The UV spectrum of 1b shows a maximum at 208 nm,
To assess the formation of a hydrophobic domain upon self-
assembly, experiments with the solvatochromic dye Nile Red
were performed (Figure 3B). Both the fluorescence emission
wavelength and intensity of Nile Red depend on the polarity
and that of 1a shows a maximum at 211 nm with a shoulder
around 225 nm.
The previously studied BTAs comprising aliphatic side chains
[
52]
show a C -symmetrical, helical arrangement of the hydrogen
of its environment. In our experiments, the emission intensi-
ty and lmax are directly related to the presence and polarity of
a hydrophobic pocket. Samples were prepared by injection of
3
bonds stabilising the aggregates in alkane solvents and the
À3
a solution of Nile Red in methanol (5 mL, c=2.5”10 m) into
water (2.5 mL), followed by the desired BTA (5 mL, c=1”
À2
À6
À5
1
0 m, final concentrations: cNR =5”10 m, cBTA =1”10 m).
Solutions of 1c and 1d showed no change in fluorescence
intensity or lmax compared to water, which indicated that no
hydrophobic domains were formed (data not shown). The
emission intensity of Nile Red in a solution of 1a increased
two orders of magnitude and lmax shifted to 625 nm (Fig-
ure 3B). In the solution of 1b, the intensity increase and the
blueshift of lmax to 633 nm was much smaller (Figure 3B).
The results from the Nile Red assay, UV absorption and
1
H NMR experiments show that 1c and 1d do not aggregate
in water at these concentrations. The UV spectrum of 1b (c=
À5
1
”10 m) does not suggest assembly of the BTAs, whereas
the Nile Red study indicates the formation of domains that are
slightly more hydrophobic than water. Furthermore, in the
1
H NMR experiments, measured at two orders of magnitude
higher concentration, compound 1b does form small aggre-
gates. These results may be explained by the amphiphilic
nature of the molecules driving the formation of micelles,
whereas the UV spectrum indicates that within these micellar
aggregates no significant dipole interactions between the BTA
cores occur. In contrast, all techniques indicated assembly of
1a at 208C.
Figure 3. A) UV/Vis spectra of 1a–d (cꢀ1”10À5 m) at 208C in water; the UV
spectra of 1c and 1d perfectly overlap (small deviations in absorption
intensities are attributed to small variations in the sample concentrations);
B) Fluorescence emission spectra of Nile Red in aqueous solutions of 1a and
À5
À6
1
b and in water at 208C (cBTA =1”10 m, cNR =5”10 m, lex =550 nm).
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À4
Figure 4. A) CryoTEM image of 1a in water (c=6.7”10 m); scale bar is
0
.5 mm. Inset: zoom, scale bar is 100 nm; B) SAXS profiles of 1a in aqueous
À3
solution (c=3.2”10 m) measured at 208C (
&
); fitted with the Schurten-
berger–Pedersen form factor for wormlike, self-avoiding chains (c).
=(6.6Æ0.2) nm, rcs =(3.4Æ0.1) nm and L >70 nm (beyond the
experimentally accessible range).
L
k
c
To determine the dimensions of the assemblies formed by
À4
1
a in water, cryoTEM (c=6.7”10 m, Figure 4A) and small-
À3
angle X-ray scattering (SAXS) measurements (c=3.2”10 m,
Figure 4B) were carried out. The cryoTEM experiments re-
vealed fibres of high aspect ratio with lengths of several micro-
metres and a diameter of approximately 5 nm. The fibres did
not appear to form bundles; but this was difficult to assess
due to the high fibre density in the sample. The SAXS profile
was characteristic for one-dimensional fibres with lengths
beyond the experimentally accessible q-range. Data extraction
1
Figure 5. Variable temperature H NMR spectra of 1a (top) and 1b (bottom)
À3
in D
2
O (c=1”10 m) using water presaturation for improved signal-to-
noise.
At 258C, the spectrum of 1b displays broad signals for the
[53]
from the profile using a wormlike chain model yielded a con-
tour length beyond the limit of 70 nm. The Kuhn length (Lk),
a measure for the stiffness of supramolecular polymers, was es-
aliphatic spacer and saccharide end group. Only one signal ap-
pears in the aromatic region at d=7.9 ppm, corresponding to
the triazole resonance. At 408C, a broad signal appears at d=
8.1 ppm, corresponding to the aromatic core of the BTA, and
a signal at d=3.2 ppm emerges which is assigned to the À
timated at (6.6Æ0.2) nm, and the cross-sectional radius (r )
cs
was estimated to be (3.4Æ0.1) nm at 208C. These values are in
good agreement with the cryoTEM images, and suggest that
even at high concentrations these fibres do not tend to form
bundles.
CH À next to the amide by 2D-COSY NMR spectroscopy. Inter-
2
estingly, the shift of the aromatic core in 1a is d=7.8 ppm at
908C, whereas in 1b it is already d=8.1 ppm at 408C. This
suggests p–p stacking of the cores of 1a, even at 908C, and
indicates a stronger interaction of the BTA cores in the aggre-
gates of 1a. The abrupt appearance of signals for 1a com-
pared to the gradual sharpening of signals for 1b indicates
a higher temperature stability of the supramolecular polymers
formed by 1a.
The effect of temperature on the self-assembly behaviour of
1
a and 1b
A major advantage of using saccharides as water-compatibilis-
ing units is that, unlike tetraethyleneglycol, which we used
[
45]
previously, they do not show a lower critical solution tem-
perature (LCST). Therefore, we can investigate the tempera-
ture-dependent behaviour of the supramolecular polymers
formed by 1a and the aggregates formed by 1b with the aim
to elucidate their stability and the mechanism of their self-as-
The normalised integral of the triazole signal plotted as
function of temperature clearly shows the different tempera-
ture response of the aggregates of 1a and 1b in water
(Figure 6). For 1b a plateau is reached at 508C. Based on the
1
sembly process. Variable-temperature H NMR spectroscopic
measurements were performed using pyrazine as an internal
À3
standard with a sharp signal at d=8.6 ppm (c=1”10 m,
1
Figure 5). The H NMR spectrum of 1a shows no signals other
than water at d=1.8 ppm up to 608C. At 708C, a signal for the
triazole at d=7.9 ppm, and broad signals corresponding to the
aliphatic spacer and the saccharide end group appear. At
8
08C, a broad peak at d=7.6 ppm emerges, corresponding to
the aromatic core, which shifts to d=7.8 ppm at 908C.
At 908C, also a broad peak at d=3.2 ppm appears, which is
assigned to the ÀCH À next to the amide by 2D COSY NMR
1
2
Figure 6. Normalised integral of the triazole H NMR signal as function of
À3
spectroscopy.
temperature for 1a and 1b (c=1”10 m) taken from spectra in Figure 5.
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line broadening persisting up to 908C, we hypothesise that
above 508C 1b is present in little clusters of only a few mole-
cules. Taken that the integral of the triazole signal reaches its
maximum value at 508C, we estimate that at 258C the fraction
1c and 1d show no temperature dependency (the Supporting
Information, Figure S2).
By heating a solution of 1b from 20 to 908C, the UV absorp-
tion gradually shifts from 208 to 207 nm accompanied by
a small increase in intensity. The UV spectrum of 1a shows
a more complex temperature-dependent behaviour. At 208C,
the UV spectrum of 1a shows a maximum and a shoulder at
211 and 225 nm, respectively. Increasing the temperature to
628C (Figure 8A) induces a decrease in intensity of the absorp-
tion and a shift to 208 nm. Concurrently, a shoulder at 195 nm
emerges, which is separated by an isosbestic point. By increas-
ing the temperature to 908C (Figure 8B), the shoulder at
195 nm diminishes and the absorption maximum at 208 nm
further increases, again through an isosbestic point. The tem-
perature behaviour is completely reversible and upon cooling
to 208C the initial spectrum is obtained again.
1
of 1b in aggregates too large to be detected by H NMR spec-
troscopy is roughly f=0.5. For 1a, the normalised integral of
the triazole signal sharply increases above 608C. Based on the
shape of the curve, it seems that a plateau is not yet reached
even at 908C. For 1a, we estimate f=0.5 is reached between
6
5 and 708C. The sharpness of the transition in 1a suggests
[54]
a cooperative supramolecular polymerisation process.
More insight into the nature of the supramolecular polymeri-
sation processes of 1a and 1b was gathered by variable-
temperature UV measurements. The UV spectra of 1a and, to
a much lesser extent, of 1b change upon increasing the
temperature (Figure 7 and Figure 8), whereas the UV spectra of
The transition between 20 and 628C has a non-linear tem-
perature dependency (see below) and appears to be a transi-
tion from one aggregated state to another, indicated by the
upcoming band at 195 nm and the presence of an isosbestic
point. Previously, the opposite transition has been observed
[
45]
after injection of 5 from methanol into water. Also, an ab-
sorption maximum at 194 nm has previously been attributed
to BTA self-assembly into columnar helical stacks stabilised by
[
49]
threefold hydrogen bonding in organic solvents, and to heli-
[
45]
cal BTA assembly in water. The absorption band at 208 nm
may indicate that a part of the BTAs is in an unordered confor-
mation, however, the 1,2,3-triazole unit also has an absorption
À5
[55]
Figure 7. UV/Vis spectra of 1b (c=1”10 m) in water measured between 0
and 908C with 108C intervals.
maximum at 210 nm in water, which may contribute to the
overall intensity at this wavelength. Taking the results from the
1
H NMR experiment into account, the observed transition
between 20 and 608C seems to be accompanied by a
reduction in the size of the aggregates. The intensity increase
at 208 nm indicates partial disruption of the aggregates at
high temperatures.
The differences in the temperature-dependent behaviour of
the assemblies of 1a and 1b were further assessed by probing
the hydrophobic domains with Nile Red at various tempera-
tures. In both cases, an initial shift towards shorter wave-
lengths was observed at low temperatures, which is attributed
to the temperature dependency of the Nile Red emission. The
same shift was reproduced in variable temperature measure-
ments of Nile Red both in ethanol and acetonitrile (the Sup-
porting Information, Figure S3). By heating a solution of 1a to
7
08C the Nile Red emission shows a sharp decrease and shift
to longer wavelengths (Figure 9A), in agreement with the vari-
able temperature UV measurements. In solutions of 1b only
a gradual intensity decrease and shift of lmax is observed (Fig-
ure 9B). Interestingly, at 808C lmax is more blueshifted for 1a
than for 1b. This indicates that at 808C hydrophobic domains
are still present in case of 1a, which further supports that the
sharp transition is one between different aggregation types.
À5
Figure 8. UV/Vis spectrum of 1a in water (c=1”10 m) measured at
temperatures between 20 and 908C displaying two different transitions.
A) 20 to 628C; B) 64 to 908C.
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À4
Figure 10. CryoTEM images of 3 and 4 in water (c=3.4”10 m); scale bar is
0.5 mm. Inset: zoom, scale bar is 100 nm.
right inset) does not clearly show the same feature, possibly
due to poorer contrast or focus. Both 3 and 4 show some
bundling, but this appears to be a result of the relatively high
concentration rather than a general effect.
Figure 9. A) Nile Red fluorescence emission in an aqueous solution of 1a
Temperature stability of assemblies of 1a and 2–4
À5
À6
(
cBTA =5”10 m, cNR =5”10 m) measured at temperatures between 10 and
8
08C (108C intervals) displaying an abrupt shift between 60 and 708C;
The influence of the configuration of the monosaccharides on
the self-assembly behaviour was further explored by variable-
B) Nile Red fluorescence emission in an aqueous solution of 1b
À5
À6
(
c
BTA =5”10 m, cNR =5”10 m) measured at temperatures between 10 and
temperature UV spectroscopy. The UV spectra of 2–4 in water
8
08C (108C intervals) displaying a gradual temperature-dependent trend in
À5
(
c=1”10 m) were recorded at temperatures between 20 and
the Nile Red emission.
9
08C and compared to those obtained for 1a (Figure 8). Sam-
ples were prepared by addition of water to the solid material
and upon gentle heating the solid material dissolved readily,
resulting in a clear solution.
The role of the monosaccharides in the self-assembly
process
At 208C, compounds 2–4 show the same typical UV spec-
trum as 1a and the same transitions occur when increasing
the temperature to 908C (the Supporting Information, Fig-
ure S4). Interestingly, the changes and transitions in the UV
spectra of 1a and 2–4 occur at almost identical temperatures.
Only the shape and intensity change of the shoulder at
195 nm varies slightly between 1a–4. Possibly, this is due to
the solvent cut-off of water, which is 190 nm at 208C and
shifts with temperature, making this part of the spectrum sen-
sitive to small deviations of the background correction. Never-
theless, in all cases the same trend is observed. Interestingly,
for 2, a transition in the absorption spectrum is observed di-
rectly after sample preparation and over the course of 105 min
(the Supporting Information, Figure S4 inset). The absorption
maximum below 200 nm diminishes while simultaneously
a maximum and shoulder at 211 and 225 nm, respectively,
After assessing the importance of an adequately long
hydrophobic spacer, the role of the configuration of the
monosaccharides in the BTA self-assembly was explored. To
that purpose, enantiomerically pure 2–4, all comprising a dode-
cyl spacer, were prepared in addition to 1a and studied with
UV and circular dichroism (CD) measurements at room temper-
À5
[56]
ature (c=1”10 m). Like l-proline,
monosaccharides may
induce a helical preference in the BTA-based supramolecular
polymers. Although the UV spectra were indicating that all
derivatives were self-assembled (see below), no CD effect was
observed in any of the samples, indicating that no helical
bias was induced by the monosaccharides.
CryoTEM measurements were performed to visualise the
aggregates formed by 2–4 in aqueous solution and to assess
their shape and dimensions (Figure 10). Similar to 1a, both 3
and 4 form long, fibrillar aggregates of micrometres in length
[
45]
emerge similar to what was reported for 5 previously. We
expect that this process may occur for all BTA derivatives 1a
and 2–4. It appears that heating the solution to 608C induces
the reversed transition.
and a diameter of approximately 5 nm at a concentration of
À4
3
.4”10 m (Figure 10). For 2 no fibres could be observed, pos-
sibly due to poor-quality ice formation during the vitrification
process. However, based on our results (see below) we expect
To visualise the trends more clearly, the normalised absorp-
tion intensity of 1a and 2–4 at 225 nm is plotted as function
of temperature (Figure 11). BTA derivatives 1a and 2–4 all dis-
play a very similar temperature profile. A gradual decrease of
the intensity is observed going from 90 to 608C, followed by
an apparent cooperative intensity increase between 60 and
208C. The final absorption intensity is reached fastest by 1a
and 4, whereas 2 deviates slightly. The gradual decrease
2
to form supramolecular polymers similar to 1a, 3 and 4. In-
terestingly, the supramolecular polymers of 3 display a periodic
variation in diameter and intensity, as can be seen at a higher
magnification (Figure 10, left inset). A similar feature was ob-
served in the cryoTEM images of 5 and is tentatively attributed
to
a
non-circular cross-section of the supramolecular
[
45]
polymers. The higher magnification image of 4 (Figure 10,
Chem. Eur. J. 2016, 22, 4608 – 4615
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1
a shows a mixture of fibres longer than 10 mm and short
fibres. For 3, mostly long fibres of around 10 mm long are ob-
served. Instead, short fibres are present in the solution of 4,
but some fibres reach up to 10 mm in length. To rule out non-
specific or hydrophobic interactions, fluorescein-labelled ben-
zoxaborole 10 was added to a pre-equilibrated solution of 5 in
water. In this solution, no aggregates could be visualised with
fluorescent microscopy. This strongly suggests selective dy-
namic covalent-bond formation to occur between the benzox-
aborole and the monosaccharides in 1a, 3 and 4.
Figure 11. Normalised UV/Vis absorption of aqueous solutions of 1a–4
Although cryoTEM measurements showed that 1a, 3 and 4
form fibres of similar dimensions of several micrometres in
length, the fluorescence microscopy measurements suggest
that the average length of the fibres varies. Possibly, the
length scale of the cryoTEM is too small to detect these differ-
ences. However, the observed differences may also result from
complexation of benzoxaborole with the supramolecular poly-
mers, which may influence the self-assembly and exert a differ-
ent effect on the different monosaccharides. Importantly, these
measurements show the proof of principle of a post-assembly
dynamic step to functionalise, label, or target the supramolec-
ular polymers, and preparation of multivalent benzoxaborole
derivatives may lead to higher affinity. Furthermore, in a
multicomponent system, this approach may allow for the
development of selective cross-linkers, or recruiters to gain
spatiotemporal control on the population distribution over the
À5
(
c=1”10 m) at 225 nm plotted as a function of temperature.
during cooling from 90 to 608C corresponds to the formation
1
of small aggregates according to the H NMR experiments.
Below 608C the aggregation into long supramolecular poly-
mers seems highly cooperative. Possibly, the small aggregates
formed at high temperature act as nucleus for elongation. The
minor difference between the temperatures at which the elon-
gation occurs for 1a and 2–4 is surprising because a strong
relation between the introduction of different anomers or
epimers and the critical micelle concentration (CMC),
solubility, and aggregate shape has been observed in alkyl
[
10–14]
glycosides.
[
57]
length of the supramolecular polymer.
Targeting the saccharide units through dynamic covalent
bond formation with benzoxaborole
Conclusion
As a versatile strategy for post-assembly functionalisation, the
potential for selectively targeting the saccharides in 1a and 2–
A series of BTA derivatives functionalised with monosaccharide
end groups has been synthesised and studied. A dodecyl
spacer separating the BTA core from the saccharide end group
is required for the formation of supramolecular polymers of
high aspect ratio. The introduction of different monosacchar-
ides has little effect on the self-assembly behaviour. When
increasing the temperature, the supramolecular polymers are
disrupted at almost identical temperature. This is in sharp con-
trast to alkyl glycosides, which show a strong dependency of
the solubility and CMC on the configuration of the saccharide,
and indicates these BTAs do not behave simply as saccharide-
based amphiphiles. The formation of long supramolecular
polymers appears to be a cooperative process. However, differ-
ent species are present at different temperature regimes. Possi-
bly, the small aggregates present at high temperature act as
nucleus for elongation, although this needs to be studied in
more detail. Finally, we show that the monosaccharides in the
supramolecular polymers can be selectively targeted through
dynamic covalent bond formation. This strategy may provide
a powerful method to functionalise or label the supramolecular
polymers in a post-assembly stage.
4
by dynamic covalent bond formation with benzoxaborole
[
40,41]
was assessed.
Fluorescein-labelled benzoxaborole 10
À7
(Figure 12,
c=5”10 m)
was
added to pre-equilibrated solutions
À5
of 1a, 3 and 4 (3”10 m) and
the binding was qualitatively as-
sessed by fluorescence microscopy
(Figure 13).
The snapshots from imaging the
solutions (see the Supporting In-
formation for the movie files) show
fluorescent fibrillar objects, al-
though with some differences be-
tween 1a, 3 and 4. The sample of
Figure 12. Fluorescein-la-
belled benzoxaborole 10.
Acknowledgements
À5
Figure 13. Fluorescence microscopy images of 1a, 3, and 4 (c=3”10 m) in
the presence of fluorescein-labelled benzoxaborole 10 (c=5”10 m)
showing fibrillar objects in all cases. Scale bar is 10 mm.
À7
We gratefully acknowledge financial support from the Dutch
Ministry of Education, Culture and Science (Gravity program
Chem. Eur. J. 2016, 22, 4608 – 4615
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0
2
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Keywords: boronic acids
·
fluorescence spectroscopy
·
polymers · self-assembly · supramolecular chemistry
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2
Received: November 25, 2015
Published online on February 17, 2016
2
Chem. Eur. J. 2016, 22, 4608 – 4615
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4615
ꢀ 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim