Colorimetric sugar sensing method useful in "neutral" aqueous media

Article abstract of DOI:10.1246/cl.2000.856

Azobenzene derivatives bearing one or two aminomethylphenylboronic acid groups were synthesized: they were confirmed to be useful for practical colorimetric saccharide sensing in "neutral" aqueous media.

Full text of DOI:10.1246/cl.2000.856

856
Chemistry Letters 2000
Colorimetric Sugar Sensing Method Useful in “Neutral”Aqueous Media
Kazuya Koumoto and Seiji Shinkai*
Department of Chemistry and Biochemistry, Graduate School of Engineering, Kyushu University, Fukuoka 812-8581
(Received April 21, 2000; CL-000380)
Azobenzene derivatives bearing one or two aminomethyl-
phenylboronic acid groups were synthesized: they were con-
firmed to be useful for practical colorimetric saccharide sensing
in “neutral” aqueous media.
The specific interaction between phenylboronic acids and
saccharides or related compounds has been attracting increasing
attention as a novel force for sugar recognition in an aqueous
system.1–8 Among them, the most successful example is based
on fluorometric sensing coupled with photoinduced electron-
transfer (PET).1 Compound 1 is the typical example: the
boronic acid group has an sp3-hybridized orbital suitable to the
saccharide-binding due to the intramolecular B–N interaction
and the fluorescence quenching efficiency of the nitrogen base
is sensitively affected by the saccharide-binding.1,9 In fact, 1
acts as a practical saccharide sensor useful in ‘neutral’ aqueous
media. Judging from convenience and simplicity of the read-
ing-out system, however, colorimetric sensing seems to have
many advantages over fluorometric sensing. To the best of our
knowledge, there exist only a few colorimetric sensing methods
using the boronic acid function. Compound 2, in which the
B–N interaction affects the intramolecular charge-transfer band
in response to the saccharide-binding, changes it color in ‘neu-
tral’ pH region, but the color change is not so large.10 This is
due to the low basicity of the amino group acting as an electron-
donor group. To avoid this drawback compound 3 was devel-
oped, in which the pyridine nitrogen acting as an electron-accep-
tor group can interact with the boronic acid group more
strongly.11 As expected, a large color change was in fact
induced in this intermolecular system11 but the reformation of
this system into a more convenient intramolecular system is not
yet achieved. More recently, Strongin et al.12 reported an inter-
esting finding that a boronic acid-containing resorcin[4]arene
results in various colors when it is heated with saccharides.
However, this reaction is irreversible and cannot be applied as a
saccharide sensor.
Compounds 4 and 5 were synthesized from 4-nitro-N-
methylacetanilide according to Scheme 1. The products were
1
identified by IR, H NMR and Mass (ESI TOF) spectral evi-
dence and elemental analyses13. When the absorption spectra
of these compounds were measured in MeOH/water mixtures at
25 °C, the distinct spectral change was observed in water-rich
region. This spectral change is attributed to the aggregation
equilibria. We thus employed a MeOH/water (pH 7.50 with 50
mmol dm–3 phosphate buffer) = 1:1 (v/v) mixture where the
spectral changes satisfied the Lambert–Beer’s law ( [4 or 5] =
(0–5.00) × 10–5 mol dm–3).
Taking these backgrounds into consideration, we newly
designed compounds 4 and 5: as the pKa of the amino groups
should be higher than that in 2, one may expect the sufficient
B–N interaction which eventually induces a large color change.
Compound 6 was used as a reference compound.
Copyright © 2000 The Chemical Society of Japan

Chemistry Letters 2000
857
becomes CD-active.1,7–10 Under the present spectral measurement
conditions, no CD-active species was recognized. The finding
implies that both 1:1 and 1:2 complexes with 4 are noncyclic
species. Thus, the K1 for the formation of 1:1 complexes (=
[4·saccharide]/[4] [saccharide]) and K2 for the formation of 1:2
complexes from 1:1 complexes (= [4·(saccharide)2]/[4·saccharide]
[saccharide]) were estimated by a nonlinear least-squares compu-
tation method, assuming that the ε values of the 1:1 complexes are
approximated by those of the 5·saccharide complexes and that the
1:2 complexes are formed at the plateau region in phototitration of
4: K1 = 797 74 and K2 = 320 29 for D-fructose, K1 = 16 4 and
K2 (too small to determine) for D-glucose, and K1=355 10 and
K2 = 69 2 for D-ribose. Again, both the K1 and K2 values appear
in the order of D-fructose > D-ribose > D-glucose. Although
observed saccharide selectivity is common between 4 and 5, one
can raise a merit of 4 over 5: as seen from Figure 2, ∆A at pH 7.5
induced by the saccharide addition in 4 (e.g., 0.13 at maximum
wavelength for D-fructose) is greater than that in 5 (0.05). This
leads to the high sensitivity in colorimetric saccharide sensing.
In conclusion, this paper demonstrates the first example for
the practical colorimetric sensing system useful even in “neutral”
aqueous media. The breakthrough has been brought forth owing
to the preparation of an electron-rich dye-conjugated nitrogen for
the B–N interaction. We believe that this basic concept can be
further elaborated to “selective” saccharide sensing by a colori-
metric method.
The typical example of the pH-dependent spectral change in
5 is shown in Figure 1. A plot of A500 vs pH (Figure 2) was
obtained from this spectral change. Thus, the pKa1 (deprotona-
tion of Me2NH+) and pKa2 [conversion of B(OH)2 to B–(OH)3]
were estimated to be 2.8 and 8.9, respectively. Phototitration of 4
gave rise to a similar pH-A500 profile with pKa1 = 3.1 and pKa2
=
8.7. In phototitration of 5 a few tight isosbestic points (e.g., at
457 nm) appeared in pKa2 region whereas in phototitration of 4
they were significantly divergent. The difference indicates that
in 4 two boronic acid groups form the OH–-adducts in a stepwise
manner. However, the difference in their pKa2 values is so small
that the titration curve is apparently analyzable as a single acid
dissociation process.
In the presence of 0.100 mol dm–3 D-fructose (which shows
the highest affinity with monoboronic acids1–4,7–11), the pKa2 val-
ues largely shifted to lower pH region: pKa2 = 5.8 for 4 and 5.9 for
5 (Figure 2). Also in the presence of D-fructose, phototitration of
5 resulted in a few tight isosbestic points whereas that of 4 result-
ed in significantly divergent isosbestic points. The difference
implies that the saccharide-binding to the two boronic acid groups
in 4 also occurs in a stepwise manner. It is clearly seen from
Figure 2 that large spectral changes are induced by the saccharide-
binding at pH 6–9. The findings support the view that these
boronic acid-containing azobenzene derivatives are useful for
practical colorimetric saccharide sensing in “neutral” pH region.
We have found that a distinct color change (from yellow to pink-
ish red) is observable upon addition of saccharides. In particular,
both of the saccharide-induced pKa2 shift and the absorbance
change are much larger in diboronic acid-containing 4 than in
monoboronic acid-containing 5. For 6 which has no boronic acid
group, the perceptible spectral change was not induced at pH 4–11
by the saccharide addition (Figure 2).
To estimate the association constants with monosaccharides
the absorption spectra were measured as a function of saccharide
concentrations. The spectral change in 5 again held tight isos-
bestic points. The plots of A480 vs [saccharide] are shown in
Figure 3. Assuming the formation of 1:1 complexes, the associa-
tion constants (K1 / dm3mol–1) were estimated to be 433 8 for
D-fructose, 13 0.2 for D-glucose, and 127 3 for D-ribose.
The K1 values appear in the order of D-fructose > D-ribose > D-
glucose, which is in line with the general affinity order of
monoboronic acids for monosaccharides.1,2,4,14 The absorption
spectral change in 4 again featured the divergent isosbestic
points, suggesting that the saccharide-binding occurs in a step-
wise manner. It is known that when a diboronic acid forms a
macrocyclic 1:1 structure with a saccharide, the resultant complex
References and Notes
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