A multinuclear NMR perspective on the complexation between bisboronic acids and bisbenzoxaboroles with: Cis -diols

Article abstract of DOI:10.1039/c7nj04143h

The interaction between boronic acids and cis-diols is an intense area of research, with numerous applications in sensing, separation, drug delivery, and materials science. Several analytical tools have been proposed over the past 20 years to probe these interactions, but most of them are limited by shortcomings such as high concentration requirements and/or the lack of clear evidence of the nature of the species present in solution. These drawbacks are all the more problematic in the case of compounds bearing multiple binding sites. In this work, a multinuclear (¹H, ¹¹B, ¹⁹F) NMR spectroscopy approach has been used to investigate for the first time the equilibria between bisboronic acids and bisbenzoxaboroles (as well as their monofunctional counterparts) with representative cis-diols, including catechol derivatives. The developed method is shown to provide valuable and straightforward insight into the nature of the species in solution, and the way in which the equilibria can be affected by the complexation media. In particular, the interest of using a ¹⁹F-tag on the cis-diol is emphasized.

Full text of DOI:10.1039/c7nj04143h

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Jason B. Benedict et al.
The role of atropisomers on the photo-reactivity and fatigue of
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A Multinuclear NMR Perspective on the Complexation between
Bisboronic acids and Bisbenzoxaboroles with cis-Diols
Adèle Larcher,a,b Aurélien Lebrun,b Michael Smietana,*,b and Danielle Laurencin*,a,†
Received 00th January 20xx,
Accepted 00th January 20xx
The interaction between boronic acids and cis-diols is an intense area of research, with numerous applications in sensing,
separation, drug delivery, and materials science. Several analytical tools have been proposed over the past 20 years to
probe these interactions, but most of them are limited by shortcomings such as high concentration requirements and/or
the lack of clear evidence of the nature of the species present in solution. These drawbacks are all the more problematic in
the case of compounds bearing multiple binding sites. In this work, a multinuclear (1H, 11B, 19F) NMR spectroscopy
approach has been used to investigate for the first time the equilibria between bisboronic acids and bisbenzoxaboroles (as
well as their monofunctional counterparts) with representative cis-diols, including catechol derivatives. The developed
method is shown to provide valuable and straightforward insight into the nature of the species in solution, and the way in
which the equilibria can be affected by the complexation media. In particular, the interest of using of a 19F-tag on the cis-
DOI: 10.1039/x0xx00000x
www.rsc.org/
diol
is
emphasized.
been explored to further enhance the binding strength to
receptors.8 However, while many bisboronic acid scaffolds
have been synthesized to enhance the selectivity towards
saccharidic targets,7c,9 the development of bisbenzoxaborole
architectures is still in its infancy. Indeed, although
benzoxaborole chemistry is developing rapidly,6,10 only a
handful of non-polymeric multibenzoxaborole architectures
have been described so far.6,11 Most surprisingly, no specific
study concerning the reactivity of these multivalent derivatives
towards diols has been reported, even for the most-simple
candidates, bisbenzoxaboroles.
In this context, we decided to investigate the spontaneous
reaction of bisbenzoxaboroles with cis-diols in solution, in view
of developing rapid and simple methodologies for comparing
the reactivity of these molecules to those of simple
benzoxaboroles, as well as mono- and bisboronic acid
analogues. Indeed, developing such tools could offer unique
opportunities for tailoring the organoboron molecule / cis-diol
equilibria in view of the construction of novel supramolecular
assemblies. Among the various analytical techniques
developed to measure the binding constants between boronic
acids or benzoxaboroles and diols,6,7b,12 the fluorescence assay
introduced by Springsteen and Wang (vide infra) has long been
the most frequently used.13 However, this assay intrinsically
presents limitations, especially when several equilibria occur in
solution, and it has been a matter of debate in the
community.12a In contrast, solution NMR spectroscopy
methods have been shown to present attractive features,12c,14-
16 especially for identifying the nature of the species present in
the medium, but their use has so far been much scarcer. With
the current increase in sensitivity of NMR experiments, thanks
to the development of cryoprobes and higher magnetic field
Introduction
The reversible interaction of boronic acids with cis-1,2- and
1,3-diols, allowing the formation of five- or six-membered
cyclic boronate esters, is an intense area of research. Indeed,
as shown in the numerous reviews published recently, it allows
boronic acid moieties to be used for biomedical,1
(nano)materials,2 sensing3 and supramolecular recognition4
applications. This remarkable success undeniably comes from
two main parameters: (i) the ability of boronic acids to operate
in water, and (ii) the reactivity of their boron atom, which can
switch from sp2 to sp3 configurations upon addition of a Lewis
base.
Because of the high pKa of boronic acids (> 9 in many
cases),5 rational designs of boronic acid molecules have been
proposed to increase the Lewis acidity at the boron center,
thus allowing effective boronate ester formation at neutral pH.
In particular, the substitution of a phenylboronic acid by an
ortho hydroxymethyl group leads to the benzoxaborole
scaffold, which possesses a unique reactivity toward diols
under physiological conditions.6 For this reason,
benzoxaboroles are being exploited in an increasing number of
biological applications,6-7 including the design of new drugs.
Multivalent binding is another attractive approach which has
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magnets, the routine study of solutions of very low to determine the apparent complexation constants between
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concentration (< 1 mM) has become possible, and hence it small organoboron molecules (PBADOI:a1n0d.1039B/CB7zNxJ)041a4n3Hd
appeared to us as timely to explore in more depth the representative sugars (D-fructose and catechol). The
potential of such analyses for studying the binding of complexation values measured are reported in Table 1. The
multifunctional organoboron molecules.
Kapp,2 constants obtained here with D-fructose were consistent
In this manuscript, we present the first comprehensive with those reported in the literature in similar media (see
study of the interaction between a small bisbenzoxaborole and Table S3, supporting information).6,7b,12b,13b,13d Moreover, for
the related bisboronic acid (noted respectively bisPBBzx and both PBA and BBzx, the binding strength of catechol was much
PBB, Figure 1) with a selection of cis-diols of interest, using not higher than that of D-fructose, in line with previous studies
only the fluorescence assay, but mainly multinuclear (1H, 11B comparing different saccharides and catechol-like cis-diols
and 19F) solution NMR spectroscopy measurements. The focus (including ARS).6,13b
was purposely set on these bifunctional organoboron
Applying the same method to molecules bearing two
molecules, because they appear as strategic building blocks for organoboron functions (PBB and bisPBBzx) was found to be
the construction of supramolecular architectures and porous more problematic. First, due to solubility issues (especially for
materials like COFs (Covalent Organic Frameworks). As part of the bisPBBzx molecule), measurements had to be performed in
the investigation, the monofunctional counterparts (noted presence of DMSO (dimethylsulfoxide), using a 60/40 volume
respectively BBzx and PBA, Figure 1) were also studied. The ratio between DMSO and the aqueous phosphate buffer. Kapp,2
objective is to emphasize the complementarity of these constants were hence also remeasured for PBA and BBzx in
different NMR characterization techniques for analyzing the this medium. In all cases, the presence of DMSO led to higher
various complexes that exist when bisbenzoxaboles and values than in pure PBS. However, the main difficulty arose
bisboronic acids interact with cis-diols in solution, and to from the fact that the reactions of diols with the two
propose new ways of using 19F NMR for this purpose.
organoboron moieties of bifunctional molecules like PBB and
bisPBBzx are not independent. Indeed, a simple treatment of
the fluorescence data assuming that both boron functions
were independent led to unsatisfactory fits, as illustrated for
PBB in Figures S2 and S3 (supporting information). Moreover,
attempts to determine the stoichiometry of binding of PBB and
bisPBBzx with ARS and cis-diols using Job-plots were
inconclusive. Although the limitation of Job-plots as indicators
of stoichiometry has been raised,17 this could imply that
several different complexes are simultaneously present in the
medium, such as 1:1 and 1:2 complexes between the
organoboron molecule and the diols. Another issue concerns
the relative values obtained for mono and bi-functional
derivatives. Indeed, the observed stability constants for PBA
and PBB are increased by a factor of 1.9 and 2.9 with fructose
and catechol respectively, and by a factor of 2.2 for BBzx and
bisPBBzx with catechol. However, the stability constants
decrease by a factor of 1.6 for BBzx and bisPBBzx with
fructose. This counterintuitive observation is difficult to
rationalize and cannot be explained by a hypothetical loss of
cooperative action, and hence it points to the fact that the
equilibria are more complex.
BBzx
PBA
bisPBBzx
PBB
D-fructose
ARS
catechol
F-catechol
Figure 1. Structures of the organoboron compounds and cis-diols studied herein:
benzoxaborole (BBzx), phenylboronic acid (PBA), bisbenzoxaborole (bisPBBzx),
1,4-phenylenediboronic acid (PBB), Alizarin-Red S (ARS), D-fructose, catechol and
3-fluorocatechol (F-catechol).
Results and discussion
Limitations of the fluorescence titration method.
Overall, these experiments clearly show that although
fluorescence spectroscopy measurements are very sensitive,
there are several other limitations to the ARS binding assay
described in Figure 1. Indeed, it cannot provide unambiguous
information regarding the complexation of small multi-boronic
or multi-benzoxaborole anions with diols. The stoichiometry of
the complexes formed with ARS and the other cis-diols thus
needs to be determined by other means, as well as the way in
which the complexation of a diol on one side of the
organoboron molecule may affect the binding on the other
A simplified representation of the equilibria involved in the
“standard” fluorescence assay13a,b is shown in Figure 2. The
overall idea is (i) to form a fluorescent complex between the
organoboron molecule and Alizarin-Red
S (ARS) (see
supporting information, Figure S1), and then (ii) to displace the
equilibrium by adding a second diol in the medium. Both steps
are followed by fluorescence spectroscopy, in order to extract
the binding constants related to each equilibrium, and hence
derive the apparent complexation constant between the
benzoxaborole and the diol of interest (referred to here as
Kapp,2, Figure 2b).
site. Moreover, even for
a molecule bearing a single
organoboron function, there are several constraints underlying
each measurement of an apparent complexation constant: (i)
for a given diol of interest, two sets of fluorescence
In the course of the present work, this fluorescence assay
was first carried out in a phosphate buffer (pH = 7.4) in order
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DOI: 10.1039/C7NJ04143H
a/
c/
Fluorescent complex
λmax
60000
50000
40000
30000
20000
10000
0
IF
[BBzx]
Kapp,1
l (nm)
725
475
525
575
625
675
d/
88,0.0
7,0
[diol]/P
6.0
6,0
5,0
4.0
4,0
b/
3,0
y = 3.2065x - 7.4496E-01
R² = 0.999
2.0
2,0
1,0
Kapp,2
0,0
0.0
Q
3
1.0
1
2.0
2
0.0
0
0,5
1,5
2,5
Figure 2. a/ Main equilibria involved in the fluorimetric titration assay used to measure complexation constants between organoboron molecules (here BBzx) and
diols;18 b/ Benzoxaborole/ diol equilibrium corresponding to the Kapp,2 binding constant; c/ Increase in fluorescence observed upon formation of the
ARS/benzoxaborole complex in an aqueous phosphate buffer (pH 7.4) (see Figure S1 for an example of determination of Kapp,1); d/ Illustration of the linear
extrapolation of experimental data used to determine the benzoxaborole/D fructose complexation constant Kapp,2 from fluorescence measurements (see supporting
information, page S3, for details regarding the equations used to process the data).13b
measurements actually need to be performed in order to
derive Kapp,2, (ii) prior to the fluorescence measurements,
preliminary experiments are also needed to determine the
excitation wavelength and to estimate the kinetics of
complexation, using for example UV-visible spectroscopy (see
Figure S1, supporting information, for an illustration of these
experiments in the case of PBA). Hence, with the increasing
number of organoboron molecules currently being synthesized
in view of exploiting their reactivity with respect to cis-diols, it
would be preferable to have alternative tools which may
inform in a simple and rapid way on their relative affinity for a
given diol. This prompted us to look into the possibility of
using multinuclear NMR spectroscopy in a more systematic
way to study these complexation reactions. Here, we set our
focus on catechol and its derivatives, these diols being of great
interest due to their high affinity with boronic acids and
benzoxaboroles.20
Catechol
PBA
BBzx
900c
1470
NDd
2200
10660
6450
PBB
bisPBBzx
NDd
24100
a Aqueous potassium phosphate buffer (pH = 7.4; 40 mM). b DMSO/phosphate buffer mixture (60/40
v/v), prepared using an aqueous potassium phosphate buffer (pH = 7.4 ; 40 mM). c See supporting
information (Table S3) for comparisons with published data. dNot determined due to solubility issues.
Multinuclear NMR studies of boronic acid / benzoxaborole –
catechol complexations
1H NMR spectroscopy
1H NMR analyses were first performed on the monofunctional
molecules PBA and BBzx, using solvents and buffers analogous
to the ones employed in the fluorescence assays, namely a
phosphate buffer at pH ~ 7.4 and a 60/40 DMSO/phosphate
buffer mixture, both prepared here in a deuterated version.
1
Very distinct H resonances were observed for the complexed
species, as illustrated in Figure S4 (supporting information) in
the case of the BBzx-catechol system. Moreover, these
measurements confirmed that the DMSO/phosphate buffer
mixture is more favorable to the complexation reactions than
the phosphate buffer alone, in line with the fluorescence
study.
Table 1. Apparent complexation constants Kapp,2 determined by fluorimetry between
organoboron molecules (PBA, BBzx, PBB and bisPBBzx) and cis-diols (D-fructose and
catechol). The values are the average of at least three independent measurements. The
data reported for the bifunctional molecules (PBB and bisPBBzx) are in italic, because
they were derived as a first approximation from fits based on the equations reported in
supporting information (page S3), which is unsatisfactory (as shown in Figures S2 and
S3 for PBB – and discussion in the text).
Concerning the complexation of catechol with the
bifunctional molecules PBB and bisPBBzx (Figure 3), several
new H resonances were observed on the spectra, which can
be assigned to 1:1 and 1:2 complexes. In the case of PBB, the
formation of a 1:1 complex leads to the disymmetrization of
the aromatic protons, and hence to the appearance of 2
Organoboron
molecule
Phosphate
buffera
DMSO/
phosphate
bufferb
1020
1
D-Fructose
PBA
BBzx
180c
560c
NDd
NDd
4000
PBB
1940
bisPBBzx
2410
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1-1 & 1-2
complexes
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DOI: 10.1039/C7NJ04143H
1-1 & 1-2
1-1
1-2
complex
complexes
1-2
complex
1-2
complex
1-1
complex
1-1
complex
complex
25:75
40:60
*
*
*
*
*
*
7.8
7.6
7.4
7.2
7.0
6.8
6.6
6.4
6.2
7.6 7.4 7.2 7.0 6.8 6.6 6.4
5.0 4.8 4.6
δ(1H) /ppm
δ(1H) / (ppm)
Figure 3. 1H NMR analyses of the complexation reactions of PBB (left) and bisPBBzx (right) with catechol in the DMSO/phosphate buffer mixture. For the solutions containing both
molecules, the spectra shown here correspond to ~40:60 and ~25:75 molar ratios between the boron-molecule and catechol. “*” symbols correspond to unidentified species.
1
doublets on the H NMR spectrum, which are shifted to lower
1H NMR spectroscopy was also found to be useful in the
frequencies with respect to the signal of the free molecule. case of another catechol derivative, ARS. Indeed, it was
1
Upon binding of a second catechol, the equivalence of these observed that its H resonances are sufficiently resolved from
protons is restored, leading to a singlet, which is shifted those of the four organoboron molecules (PBA, PBB, BBzx,
towards even lower frequencies. This assignment was further bisPBBzx) to be able to identify the complexed species. In
confirmed by looking at the changes in relative intensities of particular, a distinct singlet around 8 ppm corresponding to
the aromatic signals of PBB as a function of the amount of the isolated aromatic proton of ARS is present on the spectra,
catechol introduced. Similar observations can be made from which shifts towards lower frequencies upon complexation, as
the comparison of the 1H NMR spectra of the shown in Figure 4 for PBB and in Figure S5 for bisPBBzx.
bisbenzoxaborole / catechol system. Indeed, the methylene
As shown above, the key advantage of the 1H NMR
and aromatic resonances of bisPBBzx both become analyses lies in the possibility of providing a more complete
inequivalent upon formation of a 1:1 complex, while for the picture of the composition of the complexation medium.
1:2 complex their equivalence is restored, and all resonances However, while the spectra shown above were relatively
are shifted to even lower frequencies (Figure 3, left). It is simple to interpret, given the high symmetry of the
worth noting that a higher proportion of 1:2 complex is organoboron molecules involved, such 1H NMR studies are
observed for the bisbenzoxaborole in comparison to PBB, as expected to be far less straightforward for more complex
would be expected on the basis of the higher affinity of these molecules, requiring the use multidimensional analyses to
molecules with respect to catechols.
resolve the different resonances. Moreover, the need to work
This 1H NMR study has several implications. First, it with deuterated media is constraining when trying to
confirms that both 1:1 and 1:2 complexes can be determine how complexation reactions of organoboron
simultaneously present in solution, thereby explaining the molecules are affected by different solvents or pH conditions.
difficulties in analyzing the fluorescence data on the basis of In this context, the use of heteronuclear NMR spectroscopy
simple analogies with the monofunctional compounds (Figure analyses appears as a more promising alternative.
2). Second, it demonstrates that bisbenzoxaboroles like
bisPBBzx have a better ability to bind together two catechol-
like molecules than bisboronic acids, meaning that they can be
seen as very interesting building units for the construction of
supramolecular architectures. Finally, it also shows that the
equilibria taking place in solution can actually be even more
complex than the simple formation of 1:1 and 1:2 complexes,
because additional minority species (yet unidentified) are
present on the spectra of PBB-catechol mixtures (species
marked with “*” symbols in Figure 3).
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Overview of heteronuclear NMR spectroscopy appVirewoaArctihcleesOnline
11
+
DOI: 10.1039/C7NJ04143H
B NMR spectroscopy has already been put forward as an
interesting tool for analyzing complexation reactions between
boronic acids / benzoxaboroles and cis-diols in solution,12d,21
due to the high natural abundance of boron-11. However, it
suffers from a lack of resolution. Indeed, although the 11B
chemical shift is known to move towards lower frequencies
upon switching from
a
planar to
a
tetrahedral-like
environment,22 the quadrupolar nature of 11B implies that the
peaks are much broader than in 1H NMR, meaning that several
planar (or tetrahedral) resonances may overlap when working
with bi- (or multi-) functional organoboron molecules. Thus,
although 11B NMR analyses were performed as part of this
work (vide infra), they did not appear to us as the most
suitable approach to elucidate the complexity of the equilibria.
13C and 17O NMR spectroscopy analyses have also been
shown in the past to be useful for studying boronic acid /
8.2
8.0
7.8
7.6
7.4
δ(1H) / ppm
Figure 4. 1H NMR analysis of the complexation reactions of PBB with ARS in the
DMSO/phosphate buffer mixture. The solution containing both molecules corresponds
to a PBB:ARS molar ratio ~ 1.5. On the spectra of the ARS molecule alone (middle) and
of the mixture (top), the 1H resonance of the “red” aromatic proton of ARS is
highlighted by an arrow, showing its shift to low frequencies upon formation of the
complex.
a/
19F NMR
in DMSO / phosphate buffer
Free
F-catechol
Complexed
F-catechol
19F NMR
in phosphate buffer
b/
-136
-138
-140
-142
-144
-138
-140
-142
-144
δ(19F) / ppm
δ(19F) / ppm
Free
F-catechol
Complexed
F-catechol
Free
F-catechol
Complexed
F-catechol
11B NMR in
DMSO / phosphate buffer
11B NMR in
DMSO / phosphate buffer
c/
40
32
24
16
8
0
40
32
24
16
8
0
δ(11B) / ppm
δ(11B) / ppm
Figure 5. a/ Equilibrium of complexation between the simplest benzoxaborole (BBzx) and F-catechol; b/ 19F NMR analyses of the complexation
reactions of PBA, PBB, BBzx and bisPBBzx with F-catechol in phosphate buffer and DMSO/phosphate buffer mixtures (50:50 molar ratio between
the 2 molecules); c/ 11B NMR analyses of the complexation reactions of PBA, PBB, BBzx and bisPBBzx with F-catechol in the DMSO/phosphate
buffer mixture (50:50 molar ratio between the 2 molecules). Additional 19F NMR spectra for these systems are shown in Figure S7.
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cis-diol equilibria in solution.15,16 However, due to the much
poorer sensitivity of these nuclei, they were not attempted.
Indeed, the concentrations used here for the solution NMR
studies (and the fact that we were working in aqueous
solutions for 17O) would have been an obstacle to observing
the weaker 13C or 17O resonances at natural abundance in
reasonable time.
19F NMR analyses were first performed toVileowoAkrticaletOntlhinee
complexation between F-catechol and PBA or BBzx (Figure 5a).
DOI: 10.1039/C7NJ04143H
The same media as previously used for the fluorescence
spectroscopy analyses were tested (phosphate buffer at
pH ~ 7.4 and 60/40 DMSO/phosphate buffer mixture). As
shown in Figure 5b, the complexation of F-catechol leads to a
new 19F NMR resonance, which is shifted towards lower
frequencies with respect to the free molecule, and whose
relative intensity increases when switching from the simple
phosphate buffer to the DMSO/phosphate buffer mixture. This
resonance was unambiguously assigned to complexed F-
catechol (i) by performing additional 11B NMR analyses of the
reaction media (Figure 5c), which demonstrate that the boron
is in a more tetrahedral-like environment after addition of F-
catechol (signal centered at ~ 11 ppm for PBA and ~ 15 ppm
for BBzx), and (ii) by recording the 19F NMR spectra of mixtures
of p-fluorobenzeneboronic acid with F-catechol and looking at
the relative intensities of the different resonances (see
supporting information, Figure S6). It is worth noting that a
similar shift had been observed by 19F NMR spectroscopy upon
complexation of fluorinated aryl-boronic acids and
benzoxaboroles with cis-diols.14a
An alternative approach for high resolution heteronuclear
NMR analyses of organoboron / cis-diol equilibria consists in
using 19F NMR spectroscopy. Indeed, through the introduction
of
spectroscopy
a
19F-tag on the organoboron molecule, 19F NMR
analyses
have
provided
valuable
information,12c,14 because the 19F chemical shift of fluorine
substituents on the aromatic cycle of aryl-boronic acids and
benzoxaboroles (simple F- atom or –CF3 group) vary upon
complexation of the organoboron function. Based on these
observations, our idea was to use a 19F-tag on the cis-diol
(rather than on the organoboron molecule), as a new means to
compare the complexation properties of a given set of boronic
acids and benzoxaboroles in different media. Although the
absolute complexation constants of the fluorinated cis-diol are
expected to be different from those of the non-fluorinated
analogue, their relative order is supposed to stay the same
when switching from one organoboron molecule to another,
making this approach potentially attractive for scaling the
relative affinity of different organoboron molecules for diols.
Most interestingly, in the case of the bifunctional
organoboron molecules (PBB and bisPBBzx), several
resonances are observed by 19F NMR spectroscopy in the
“complexed F-catechol” region (Figure 5b), whose relative
intensity varies depending on the [F-catechol/organoboron
molecule] molar ratio: the lower frequency signal (pointed by a
red arrow in Figure 5b) increases at higher F-catechol
concentrations (see Figure S7). Such observations would be
consistent with the presence of both (1:1) and (1:2) complexes
in solution. This is confirmed by complementary 1H NMR
analyses of these solutions (Figure 6), which lead to similar
observations as for the non-fluorinated catechol (Figure 3),
19F NMR spectroscopy: F-catechol as a new means for studying the
complexation equilibria in solution
Considering the stronger binding of catechols (vide supra),
we decided to use a fluorinated derivative of this molecule,
3-fluorocatechol (F-catechol), as a probe molecule for 19F NMR
spectroscopy.
1
and also by additional H DOSY NMR experiments (Diffusion
Ordered SpectroscopY
–
see Figure S8). Indeed, new
1-1
1-2
1-1
complex
complex
1-1 & 1-2
complexes
1-2
complex
1-1
complex
1-1 & 1-2
complexes
complex
1-2
complex
35:65
50:50
*
*
*
*
*
*
*
*
*
*
*
*
.
5.0
4.8
4.6
4.4
7.4 7.2 7.0 6.8 6.6 6.4
7.6
7.4
7.2
7.0
6.8
6.6
6.4
6.2
δ(1H) /ppm
δ(1H) /ppm
Figure 6. 1H NMR analyses of the complexation reactions of PBB (left) and bisPBBzx (right) with F-catechol in the DMSO/phosphate buffer mixture (for the solutions containing
both molecules, the spectra shown here correspond to ~50:50 and ~35:65 molar ratios between the organoboron molecule and F-catechol). Additional 1H DOSY (Diffusion
Ordered SpectroscopY) analyses were carried on the bisPBBzx-F-catechol system, which are consistent with the spectral assignments (see supporting information Figure S8).
6 | J. Name., 2012, 00, 1-3
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1
resonances at lower frequencies are present on the H NMR which reaction parameters (solvent, pH, temperature, ionic
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DOI: 10.1039/C7NJ04143H
spectra, which are consistent with the formation of (1:1) and strength etc..) may best favour the formation of 1:1 vs 1:2
(1:2) adducts.
entities.
These results highlight that heteronuclear NMR (ii) which molecules will more strongly interact with catechol-
spectroscopy analyses involving 19F NMR are far more sensitive like diols.
and informative than 11B NMR for studying complexation
In both cases, this will be highly valuable for tuning the
reactions between organoboron molecules and fluorinated cis- formation of supramolecular architectures based on
diols. Indeed, while different complexed F-catechol benzoxaborole (or boronic acids) / diol interactions, and even
environments are resolved by 19F NMR, only one is observed for determining the optimal conditions for dynamic
by 11B NMR (Figure 5c), most probably due to the strong combinatorial chemistry approaches. Moreover, while the
similarity in 11B NMR parameters of the 1:1 and 1:2 complexed concept was exemplified with F-catechol, it could obviously be
forms. Hence, this makes 19F NMR a highly valuable method extended to other fluoro-diols as a way to probe precisely
for shedding light on these complexation equilibria, which simple and multiple boronic acids/benzoxaboles and cis-diol
1
could turn out to be particularly useful in situations where H interactions. In this sense, we foresee that this work will lead
NMR may not be as straightforward to apply.
the way to the development and use of other fluoro-diols (e.g.
catechol derivatives bearing –CF3 substituent on the
a
aromatic cycle or fluorinated sugars) for studying these
complexation reactions.
Conclusion
In this manuscript, the complexation of simple bifunctional
organoboron molecules with cis-diols was studied from a
multinuclear NMR spectroscopy perspective. This approach
Experimental
was shown to present many advantages to compare the Reagents
binding affinity of mono- or multivalent boronic acids or
Phenylboronic acid (PBA, C6H7BO2, 98%, Alfa Aesar), benzene-
benzoboroxazoles for a defined class of cis-diol compounds.
The obtained results notably indicate that cooperativity is very
limited with PBB and bisPBBzx. The repulsive effect resulting
from proximal negative charges on the two organoboron
groups might explain this observation, because for both
molecules, these charges will merely be separated by a
benzene ring.
1,4-diboronic acid (PBB, C6H8B2O4, 95%, Aldrich),
2-(hydroxymethyl)phenylboronic acid cyclic monoester (BBzx,
C7H7BO2, 97%, Aldrich) and p-fluorophenylboronic acid
(C6H6BFO2, 97%, Alfa Aesar) were purchased from commercial
sources and used as received (after verifying the lack of boric
acid by 11B solution NMR). Alizarin red S (ARS, C14H7NaO7S,
Acros Organics), D-fructose (C6H12O6, 99%, Fluka BioChemika),
catechol (C6H6O2, 99%, Aldrich) and 3-fluorocatechol (C6H5FO2,
Alfa Aesar, 99%) were purchased from commercial sources and
used as received. Phenyl-bisbenzoxaborole (noted here
bisPBBzx) was synthesized by adapting a previously published
protocol.11e
The solvents for the fluorimetric experiments (DMSO,
99.9%, and HPLC grade ultra-pure water) were purchased from
Aldrich. The phosphate buffer (pH = 7.4, 40 mM) was prepared
by dissolving 1.361 g (10 mmol) of KH2PO4 in 250 mL of ultra-
pure water, and then adjusting the pH with a 3 M NaOH
solution. This buffer was then stored at 4 °C and used over a
period of two weeks (maximum).
1H NMR spectroscopy analyses were demonstrated to be
highly informative, allowing a complete picture of the
composition of the medium to be reached. This is a major
advantage in comparison to the “traditional” fluorescence
assay (and more generally optical spectroscopies), but also to
other methods which are currently being put forward for
studying these complexation equilibria, such as Isothermal
Titration Calorimetry (ITC)12b and Capillary Electrophoresis.12a
Indeed, none of the latter two techniques, on their own, would
allow gaining such insight for multifunctional organoboron
molecules such as those studied here, nor quantifying
unambiguously the relative proportions of 1:1 and 1:2 species.
1
Beyond the H NMR analyses, a new method based on 19F
Deuterated solvents (DMSO-d6, 99.8% and D2O, 99.9%)
NMR spectroscopy was also developed as a new means to help
investigate and compare the complexation properties of
were purchased from Eurisotop.
A deuterated sodium
hydroxide solution (NaOD – 40 wt% in D2O, 99.98%) and a
deuterated phosphate monobasic salt (KD2PO4, Aldrich; 98%
atom% D) were purchased from Aldrich. The deuterated
phosphate buffer (pH = 7.4, 40 mM) was prepared by
dissolving 55.2 mg of KD2PO4 (0.4 mmol) in 9.979 mL of D2O,
and the pH was adjusted at with 21.0 µL of the NaOD solution.
This buffer was then stored at 4 °C and used over a maximum
period of two weeks.
different organoboron molecules. By proposing the use of a 19
F
tag on the cis-diol (with F-catechol as a binding agent), 19F
NMR was found to be sufficiently sensitive and resolved to
bring evidence of the presence of several species in solution,
and hence could act as a complementary analytical tool for
future studies of the complexation equilibria. More
specifically, we foresee that the 19F NMR analysis of the
binding to F-catechol should allow determining in
straightforward way
a
Fluorescence measurements
(i) which media are the most appropriate for favouring the
complexation (as exemplified herein when comparing the
phosphate buffer to the DMSO/buffer mixture), and hence
The methodology used for the fluorescence studies was based
on the work by Wang et al. (see supporting information for
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further details).13b All fluorescence measurements were here, a recycle delay of 6 s was used, and the number of
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DOI: 10.1039/C7NJ04143H
performed on an Edinburgh Instruments, Xe 900, NIR 300/2, transients acquired ranged from 160 to 320. Chemical shifts
apparatus, using cuvettes made of Quartz Suprazil (light path were internally referenced to CF3COOH: after 19F NMR
of 10 × 10 mm). Emission scans were recorded at room acquisition of each organoboron molecule/F-catechol mixture,
temperature from *λexct°+15 nm+ to 750 nm (λexct° depending on a solution of this molecule was added directly in the NMR
the organoboron molecule/ARS complex and the medium, as tube, and a 19F NMR spectrum was then re-recorded; the 19F
shown in Table 2), using a step size of 1 nm and a dwell time of resonance of CF3COOH was set to −76.5 ppm in the phosphate
1 s; each spectrum was acquired three times. The organoboron buffer and −75 ppm in the DMSO/buffer mixture. These 19F
molecule/diol complexation constants were all measured in chemical shifts were referenced externally to a solution of
triplicate starting from independent solution preparations.
trifluorotoluene in CDCl3 (at −63.7 ppm).
11B NMR spectra were recorded at 298 K on a Bruker
Avance III 500 MHz NMR spectrometer equipped with a CP-
BBO cryoprobe. A single-pulse experiment was used (with an
additional background substraction in some cases – “zgbs”
sequence in the Bruker library). Quartz NMR tubes were used
for the measurements. The recycle delay was set to 0.2 to 0.5
s, and the number of transients acquired ranged from 2k to
10k, depending on the solution. Spectra were referenced
externally to a solution of BF3-Et2O (at 0 ppm).
Table 2. Excitation wavelengths (λexct°, in nm) used in the fluorescence measurements,
as determined by UV-vis spectroscopy (see supporting information for further details
regarding UV-vis measurements).
Phosphate buffer
DMSO/phosphate
buffer (60/40 v/v)
BBzx
PBA
bisPBBzx
PBB
459
468
465
483
466
479
Spectra were processed and analyzed using TopSpin 3.2
and Mestrenova 7.0.3 programs.
NMR analyses
1H NMR spectra were recorded at 298 K on a Bruker Avance III
600 MHz NMR spectrometer, using QXI and BBFO probes. All
1H NMR spectra were calibrated to the DMSO resonance (set
at 2.50 ppm). One-dimensional 1H NMR spectra were recorded
using the “zgpr” sequence of the Bruker library, as it allows
presaturating intense solvent resonances (in our case water).
For each sample, the following optimizations were performed
in order to be able to derive quantitative data from the
spectra: (i) the saturation scheme for the water signals was
optimized, to ensure that it did not affect the intensities of the
peaks of interest (especially the methylene resonances of
benzoxaboroles); (ii) the recycle delay was set to a value
Conflicts of interest
There are no conflicts to declare.
Acknowledgements
Dr Cédric Totée (ICGM) is thanked for his assistance in the
setting-up of the 19F NMR experiments. The LABEX
“Chemisyst” ANR 2011-01-05 is acknowledged for financial
support.
1
ensuring a full relaxation of all H resonances of interest, and
(iii) it was verified that an equilibrium between the different
species had been reached inside the NMR tube. The recycle
delays ranged from 18 to 28 s, and the number of transients
acquired from 64 to 80.
References
1
H. Fu, H. Fang, J. Sun, H. Wang, A. Liu, J. Sun and Z. Wu, Curr.
Med. Chem., 2014, 21, 3271.
For the two-dimensional 1H DOSY (Diffusion Ordered
SpectroscopY) experiments, the “ledbpgp2spr” pulse sequence
of the Bruker library was used, which corrects for Eddy current
imperfections, and also allows presaturating intense solvent
resonances (in our case water). In addition to the previously
mentioned optimizations, the gradient parameters were
optimized (“big delta” ~ 0.1 s and “little delta” ~ 3.6 ms), and
the NMR tube was spun to 20 Hz. In the 2D DOSY experiments,
32 spectra were acquired (with 16 to 64 transients per
spectrum, depending on the sample), and then processed
under standard conditions in Topspin.
2
(a) W. L. A. Brooks and B. S. Sumerlin, Chem. Rev., 2016, 116
,
1375; (b) R. F. Guan, H. H. Chen, F. X. Cao, D. X. Cao and Y. L.
Deng, Inorg Chem Commun, 2013, 38; (c) P. C. Trippier and C.
McGuigan, MedChemComm, 2010, 1, 183.
3
4
(a) W. Zhai, X. Sun, T. D. James and J. S. Fossey, Chem. Asian
J., 2015, 10, 1836; (b) J. S. Hansen, J. B. Christensen, J. F.
Petersen, T. Hoeg-Jensen and J. C. Norrild, Sensors and
Actuators B, 2012, 161, 45 ; (c) B. Akgun, C. Li, Y. Hao, G.
Lambkin, R. Derda and D. G Hall, J. Am. Chem. Soc. 2017,
139, 14285.
K. Lacina, P. Skladal and T. D. James, Chem. Cent. J., 2014, 8,
60.
5
6
D. G. Hall, Boronic acids, Wiley-VCH, 2011.
19F NMR spectra were recorded at 298 K on a Bruker
Avance III 400 MHz NMR spectrometer, using a SmartProbe
optimized for 19F. The “zgig” sequence from the Bruker library
A. Adamczyk-Wozniak, K. M. Borys and A. Sporzynski, Chem.
Rev., 2015, 115, 5224.
7
(a) M. Berube, M. Dowlut and D. G. Hall, J. Org. Chem., 2008,
73, 6471; (b) M. Dowlut and D. G. Hall, J. Am. Chem. Soc.,
2006, 128, 4226; (c) Y. Liu, X. Zhang, H. Guo, Y. Wu, Q. Li, L.
Liu and J. Zhao, J. Fluoresc., 2011, 21, 1979; (d) S. Sene, J.
McLane, N. Schaub, S. Begu, P. H. Mutin, L. Ligon, R. J. Gilbert
1
was used. H decoupling powers were optimized, and for each
sample, different recycle delays were tested to ensure full
relaxation of the different 19F signals. For the spectra shown
and D. Laurencin, J. Mat. Chem. B, 2016,
4, 257; (e) V.
8 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 9 of 11
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Please do not adjust margins
Journal Name
ARTICLE
Alterio, R. Cadoni, D. Esposito, D. Vullo, A. Di Fiore, S. M. 19 J. Yan, G. Springsteen, S. Deeter and B. Wang,VTieewtArartihcleedOrnolinne,
Monti, A. Caporale, M. Ruvo, M. Sechi, P. Dumy, C. T.
2004, 60, 11205.
Supuran, G. De Simone and J. Y. Winum, Chem. Commun., 20 (a) E. Faure, C. Falentin-Daudre, C. JDerOoI:m10e.,10J3.9L/Cys7kNaJw04a1,43DH.
2016, 52, 11983.
J. D. Badjic, A. Nelson, S. J. Cantrill, W. B. Turnbull and J. F.
Stoddart, Acc. Chem. Res., 2005, 38, 723.
(a) X. Wu, Z. Li, X. X. Chen, J. S. Fossey, T. D. James and Y. B.
Jiang, Chem. Soc. Rev., 2013, 42; (b) S. Craig, Bioorg. Chem.,
2012, 40, 137; (c) R. Zhang, J.-P. Miao, C.-H. Ge, M.-Y. Zhang, 21 (a) T. Okamoto, A. Tanaka, E. Watanabe, T. Miyazaki, T.
Fournier, P. Woisel and C. Detrembleur, Prog. Polym. Sci.,
2013, 38, 236; (b) F. Coumes, P. Woisel and D. Fournier,
Macromolecules, 2016, 49, 8925; c) A. Adamczyk-Wozniak, K.
M. Borys, I. D. Madura, A. Pawełko, E. Tomecka and K.
Zukowski, New J. Chem. 2013, 37, 188.
8
9
L.-X. Wang and X.-D. Zhang, Sensor Actuat. B-Chem, 2014,
198, 260; (d) S. Jin, Y. Cheng, S. Reid, M. Li and B. Wang,
Med. Res. Rev., 2010, 30, 171; (e) Y. Ooyama, K. Uenaka, A.
Sugaya, S. Iwatsuki, M. Inamo, H. D. Takagi, A. Odani and K.
Ishihara, Eur. J. Inorg. Chem., 2014, 14, 2389; (b) Y. Suzuki,
M. Shimizu, T. Okamoto, T. Sugaya, S. Iwatsuki, M. Inamo, H.
Matsugasako, Y. Harima and J. Ohshita, RSC Adv, 2013,
W. Zhai, L. Male and J. S. Fossey, Chem. Commun., 2017, 53
2218; (g) T. Wu, X. Xiao, Z. Zhang and M. Zhao, Chem. Sci.,
2015,
Tetrahedron, 2009, 65, 109;
3
; (f)
D. Takagi, A. Odani and K. Ishihara, ChemistrySelect, 1, 5141;
,
(c) J. W. Tomsho and S. J. Benkovic, J. Org. Chem., 2012, 77,
2098.
6
, 1206; (h) P. J. Duggan and P. A. Offermann, 22 (a) M. Sanjoh, D. Iizuka, A. Matsumoto and Y. Miyahara, Org.
Lett., 2015, 17, 588; (b) S. Sene, S. Begu, C. Gervais, G.
Renaudin, A. Mesbah, M. E. Smith, P. H. Mutin, A. van der
Lee, J. M. Nedelec, C. Bonhomme and D. Laurencin, Chem.
Mater., 2015, 27, 1242.
10 C. T. Liu, J. W. Tomsho and S. J. Benkovic, Bioorg. Med.
Chem., 2014, 22, 4462.
11 (a) A. Adamczyk-Wozniak, K. M. Borys, I. D. Madura, S.
Michalek and A. Pawelko, Tetrahedron, 2013, 69, 8936; (b) A.
Pal, M. Bérubé and D. G. Hall, Angew. Chem., Int. Ed., 2010,
49, 1492; (c) D. Claes, M. Holzapfel, N. Clausen and W.
Maison, Eur. J. Org. Chem., 2013, 2013, 6361; (d) J. F. Cairns
and H. R. Snyder, J. Org. Chem., 1964, 29, 2810; (e) R. Berger,
A. Giannakopoulos, P. Ravat, M. Wagner, D. Beljonne, X.
Feng and K. Müllen, Angew. Chem., Int. Ed., 2014, 53, 10520;
(f) A. Adamczyk-Wozniak, K. Ejsmont, B. Gierczyk, E.
Kaczorowska, A. Matuszewska, G. Schroeder, A. Sporzynski
and B. Zarychta, J. Organomet. Chem., 2015, 788, 36.
12 (a) C. C. Lu, H. Y. Li, H. Y. Wang and Z. Liu, Anal. Chem., 2013,
85; (b) S. Schumacher, M. Katterle, C. Hettrich, B. R. Paulke,
D. G. Hall, F. W. Scheller and N. Gajovic-Eichelmann, J. Mol.
Recogn., 2011, 24, 953; (c) J. Axthelm, H. Görls, U. S.
Schubert and A. Schiller, J. Am. Chem. Soc., 2015, 137
,
15402; (d) J. W. Tomsho and S. J. Benkovic, J. Org. Chem.,
2012, 77, 11200.
13 (a) G. Springsteen and B. Wang, Chem Commun, 2001, 17; (b)
G. Springsteen and B. Wang, Tetrahedron, 2002, 58, 5291; (c)
A. Adamczyk-Wozniak, K. M. Borys, I. D. Madura, A. Pawełko,
E. Tomecka and K. Zukowski, New. J. Chem. 2013, 37, 188; (d)
A. Mahalingam, A. R. Geonnotti, J. Balzarini and P. F. Kiser,
Mol. Pharmaceutics, 2011, 8, 2465.
14 (a) L. Iannazzo, E. Benedetti, M. Catala, M. Etheve-
Quelquejeu, C. Tisne and L. Micouin, Org. Biomol. Chem.,
2015, 13, 8817; (b) S. Vshyvenko, M. L. Clapson, I. Suzuki and
D. G. Hall, ACS Med. Chem. Lett., 2016,
7, 1097; (c) J.
Axthelm, S. H. C. Askes, M. Elstner, U. Reddy G, H. Görls, P.
Bellstedt and A. Schiller, J. Am. Chem. Soc. 2017, 139, 11413;
(d) R. E. London and S. A. Gabel, J. Am. Chem. Soc. 1994, 116
,
2562; (e) S. L. Yeste, M. E. Powell, S. D. Bull and T. D. James,
J. Org. Chem. 2009, 74, 427.
15 (a) J. C. Norrild and H. Eggert, J. Am. Chem. Soc. 1995, 117
,
1479; (b) J. C. Norrild and H. Eggert, J. Chem. Soc. Perkin
Trans 2, 1996, 2583; (c) M. Bielecki, H. Eggert, J. C. Norrild, J.
Chem. Soc. Perkin Trans 2, 1999, 449; (d) J. C. Norrild and I.
Sotofte, J. Chem. Soc. Perkin Trans 2, 2002, 2, 303; (e) S. P.
Draffin, P. J. Duggan, S. A. M. Duggan and J. C. Norrild,
Tetrahedron, 2003, 59, 9075.
16 B. Gierczyk, M. Kazmierczak, G. Schroeder and A. Sporzynski,
New J. Chem. 2013, 37, 1056.
17 D. B. Hibbert and P. Thordarson, Chem. Comm. 2016, 52,
12792.
18 Kinetic studies of such complexation reactions by Benkovic et
al and Ishihara et al have shown that intermediate species
also form (see ref. 21 below), which are not represented
here.
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A table of contents entry text.
A new way of using solution NMR (especially 19F NMR) to study organoboron molecule/cis-diol
equilibria is presented

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