Photoinduced electron transfer quenching and sugar effects on the electrostatic interaction between an anionic Ru(II) complex and cationic bipyridinium derivatives functionalized with boronic acids

Article abstract of DOI:10.1016/j.ica.2011.06.030

The photoinduced electron transfer quenching of an anionic ruthenium(II) metal-ligand-complex (Ru(dpp(SO₃Na))₂(mcbpy)Cl₂ by two boronic acid functionalized benzyl viologen (BV²⁺) derivatives has been investigated as well as their response to sugar. The electrostatic interaction between these two charge species lead to the formation of static quenching which is removed in presence of sugar due to the formation of a neutral zwitterionic quencher. Spectral data, quenching parameters and sugar titration curves are presented and discussed in term of future developments of optical sensors for sugar.

Full text of DOI:10.1016/j.ica.2011.06.030

Inorganica Chimica Acta 381 (2012) 104–110
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Inorganica Chimica Acta
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Photoinduced electron transfer quenching and sugar effects on the electrostatic
interaction between an anionic Ru(II) complex and cationic bipyridinium
derivatives functionalized with boronic acids
a,
⇑
b
b,
⇑
Md. Jamal Uddin , Nicolas DiCesare , Joseph R. Lakowicz
a
Department of Natural Sciences, Coppin State University, 2500 West North Avenue, Baltimore, MD 21216, USA
Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, University of Maryland at Baltimore, 725 W. Lombard Street, Baltimore, MD 21201, USA
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Available online 24 June 2011
The photoinduced electron transfer quenching of an anionic ruthenium(II) metal–ligand-complex
2+
3 2 2
(Ru(dpp(SO Na)) (mcbpy)Cl by two boronic acid functionalized benzyl viologen (BV ) derivatives has
been investigated as well as their response to sugar. The electrostatic interaction between these two
charge species lead to the formation of static quenching which is removed in presence of sugar due to
the formation of a neutral zwitterionic quencher. Spectral data, quenching parameters and sugar titration
curves are presented and discussed in term of future developments of optical sensors for sugar.
Ó 2011 Elsevier B.V. All rights reserved.
Fluorescence Spectroscopy: from Single
Chemosensors to Nanoparticles Science –
Special Issue
Keywords:
Electron transfer
Quenching
Sugar
Metal complex
Boronic acid
Spectroscopy
1
. Introduction
For a decade, the development of synthetic chemosensors and
[10]. In addition, actual developments in electronic, in light-emit-
ting devices, and solid-state detectors can lead to simple, low-cost
and miniaturized analytical devices that could be used in an auto-
matic and continuously way or used by the patient himself on an
hourly schedule.
sensors for the recognition and analysis of sugar has attracted
much attention [1–6]. Detection and monitoring of glucose is par-
ticularly important for diabetics. The long-term health problems
associated with a high blood glucose level show important reduc-
tion when diabetics can control and maintain a ‘‘normal’’ blood
glucose level. Actual technologies for glucose measurement,
mostly based on enzymes and proteins, show important limita-
tions in their uses [7,8]. For example, an implantable sensor for
continuous glucose monitoring is expected to find useful applica-
tions to help diabetics. To date, none of the actual technologies
have been shown to be robust enough for the development of these
in-vivo applications. Development of new glucose sensitive mate-
rials and new analytical approaches are still needed.
To date, the most promising chelator group used for the devel-
opment of synthetics chemosenors for sugar is the boronic acids.
Boronic acids form a covalent and reversible interaction with
monosaccharides in water. The equilibrium is fast and the affinity
and selectivity against different sugars can be enhanced [3,4,11].
Several interesting fluorescence probes functionalized with boro-
nic acids have been developed over the past decade. Mechanisms
like photoinduced electron transfer (PET), [11] excited-state
charge-transfer [12,13] and molecular rigidification [14], to name
a few, have been investigated. While these results are promising,
the development of fluorescence probes is a long process, optical
responses are hardly predictable and the mechanisms are limited
to few fluorophores. In this way, the development of mechanisms
that are independent of the nature of the fluorophores are greatly
welcome. In this topic, we used change of the boronic acid group,
upon binding saccharides, to induce optical changes based on a
re-equilibrium of the charged species present [15].
Amount analytical approaches, fluorescence spectroscopy is
well known for its promising potential as an analytical tool for
in-vivo medical analysis. In addition to its high sensibility and
selectivity, fluorescence spectroscopy is a non-invasive method
that can be performed in whole blood [9] and through the skin
The interaction of the boronic acids with sugar is shortly illus-
trated in Scheme 1 [16]. Phenyl boronic acids are weak Lewis acids
⇑
Corresponding authors. Tel.: +1 410 951 4118; fax: +1 410 951 4110 (Md.J.
Uddin), tel.: +1 410 706 8409; fax: +1 410 706 8408 (J.R. Lakowicz).
E-mail addresses: juddin@coppin.edu (Md. Jamal Uddin), lakowicz@umbi.um-
d.edu (J.R. Lakowicz).
with pK
a
s ꢀ 9. The interaction with a diol increases the electrophi-
licity of the boron group, and typical pK
a
s ꢀ 6 are than observed. At
0
020-1693/$ - see front matter Ó 2011 Elsevier B.V. All rights reserved.
doi:10.1016/j.ica.2011.06.030

Md. Jamal Uddin et al. / Inorganica Chimica Acta 381 (2012) 104–110
105
Fig. 1. Molecular structure of the benzyl viologen quenchers with boronic acid
groups (p-BV2 and o-BV
+
2+
)
3 2 2
and metal–ligand-complex Ru(II)[(dpp(SO Na) ) -
(
mcbpy)]Cl
2
.
boronic acid groups (p-BV2+ and o-BV2+, Fig. 1) and an anionic
ruthenium(II) metal-ligand-complex (MLC) (Fig. 1). Ru MLC are to-
day known for their good spectrum characteristics (photo-stability,
large Stokes’ shift, long-wavelength emission, long-lifetime, etc.)
and are very promising as active material for the elaboration of
optical sensors [18].
In addition, Ru –polypyridyl complexes show sensitivity to the
presence of bipyridinium derivative through an oxidative electron-
transfer acceptor dipyridium [19–21].
The Stern–Volmer constants (KSV) obtained from steady-state
and time-dependent measurements and super linear quenching
is discussed in terms of static quenching and sphere of action mod-
el. Consequence of the presence of sugar on the KSV and static
quenching is also presented and discussed as well as the effect of
the ionic strength of the solution on these parameters. Sugar titra-
tion curves are presented and discussed in terms of future develop-
ment of optical sensors based on this mechanism.
Scheme 1. The interaction of the boronic acids with sugar moiety.
a pH between 6.5 and 8.5, the boronic acid exists under its neutral
form (A, Scheme 1) while, in presence of sugars, its exists under its
anionic form (B and C, Scheme 1). This change has been first ex-
plored by Singaram and collaborators and recently published in a
brief communication [15]. In their study, they used a dicationic
benzyl viologen quencher functionalized with boroin acids groups
II
o-BV2+, Fig. 1) to quench, by photoinduced electron transfer, the
(
emission of an anionic pyrene derivative following an electrostatic
interaction between the two species. In presence of sugar, a neutral
zwiterionic quencher is formed (C, Scheme 1), which drawback the
formation of complexation between the quencher and the fluores-
cent probe, and then remove the quenching effect. Using this
mechanism, large intensity change can be obtained. Singaram
et al. reported intensity increases up to 5-fold using an anionic pyr-
ene derivative, while we reported intensity increases up to 70-fold
using an anionic conjugated polymer [17].
2. Experimental materials
In this study, we report the photoinduced electron transfer
quenching properties, association constants, and subsequent sugar
effect between two benzyl viologen quencher functionalized with
All solvents (HPLC grade), methyl viologen (MV2+) and starting
chemicals for the synthesis were from Aldrich and used as re-

1
06
Md. Jamal Uddin et al. / Inorganica Chimica Acta 381 (2012) 104–110
ceived.
Sigma. Ru(dpp(SO
D
-fructose,
D
-galactose and
D
-glucose were purchased from
[22], 2-(4-bromomethyl-phe-
(Corning 4303) to remove the long-wavelength emission compo-
3
Na) (mcbpy)Cl
2
)
2
2
nent from the LED. Emission was observed through a combination
of a cut-off (550 nm) and an interference filter (615 nm + 25 nm) to
remove scattered and Raman scattered light. The measurements
were taken in a 1-cm cuvette under an air atmosphere. The fre-
quency intensity profiles were analyzed by nonlinear least squares
in terms of the multi-exponential model:
nyl)-5,5-dimethyl-(1,3,2)dioxaborinane [23] and 2-(2-bromo-
methyl-phenyl)-5,5-dimethyl-(1,3,2)dioxaborinane [11] were
obtained as described in the literature. The ortho derivative, o-
2
+
BV , was synthesized according to a slightly different procedure
than reported in the literature [15].
X
0
+
0
IðtÞ ¼
ai expðꢂt= i
s
Þ
ð3Þ
2
(
.1. 4,4 -N,N -Bis(benzyl-2-boronic acid-bipyridinium dibromide
2
i
o-BV )
where
a
i
is the pre-exponential factor associated with the decay
P
4
,4-Dipyridyl (375 mg, 2.4 mmol) and 2-(2-bromomethyl-phe-
nyl)-5,5-dimethyl-(1,3,2)dioxaborinane (1.5 g, 5.3 mmol) were
heated to reflux in CH CN (25 ml) for 4 h. After cooling to room
time
s
i
, with ia
i
= 1.0. The mean lifetime is given by:
P
2
X
3
a si
i
s
¼ P
¼
f
i
s
i
ð4Þ
temperature, the resulting yellow precipitate was recovered by fil-
tration and washed with acetone. Without further purification, the
solid was dissolved in an H
a
i i
s
2
O/MeOH solution and extracted for
where f is the fractional steady-state intensities of each lifetime
i
two days with ethyl acetate. After separation, the aqueous solution
was concentrated and the product was precipitated by the addition
of acetone, giving 871 mg (62%) of a pale yellow solid. Analytical
and spectroscopic measurements are in accordance with those
published [15].
component.
a
ja
i i
s
j j
f
i
¼ P
ð5Þ
s
Errors of 0.5 and 0.01 on the phase angle and modulation have
been used, respectively.
0
0
2
.2. 4,4 -N,N -Bis(benzyl-4-boronic acid)-bipyridinium dibromide
2
+
(
p-BV
)
4
. Results and discussion
The para derivative, p-BV2+, was synthesized according to the
procedure described above, starting with 2-(4-bromotheyl-phe-
nyl)-5,5-dimethyl-(1,3,2)dioxaborinane, giving yellow solid
74%), mp 236–237 °C (decomp.). (300 MHz; CD OD) 6.00 (4H, s,
CH2), 7.59 (2H, d, J = 7 Hz, PhH), 7.76 (2H,d, J = 7 Hz, PhH), 8.70
Fig. 2 displays the absorption and emission spectra of the
a
Ru-MLC investigated as well as the absorption spectrum of p-
2
+
2+
2+
(
3
BV . Both quenchers, p-BV and o-BV , show identical absorp-
tion profiles. The MLC was excited through its metal-ligand-
charge-transfer (MLCT) transition at 460 nm. At this wavelength,
both benzyl viologen derivatives do not show any significant
absorbance.
+
(
2H, d, J = 6hz, ArH), 9.38 (2H, d, J = 6 Hz, ArH). m/z (EI) 485 (M ,
00%). Anal. Calc. for C24 Br (585.9): C, 49.20; H, 4.13;
N 4.78. Found: C, 48.76; H, 4.5; N, 4.67%.
1
H
24
B
2
2 2 4
N O
For all measurements, the absorption spectrum was recorded
after the complete addition of the quencher, and in all cases, no
change in the spectrum was observed (Fig. 3), except at shorter
wavelengths than 375 nm where the quenchers show absorption.
This result is in contrast with the results reported by Singaram
et al. where they observed changes in the absorption spectrum of
3
. Method and instrumentation
Absorption and emission spectra were recorded with a Cary 50
UV–Vis spectrophotometer (Varian Inc.) and a Eclipse Spectrofluo-
rometer (Varian Inc.), respectively. In all cases, the measurements
were taken at room temperature in a 1-cm quartz cuvette under an
air atmosphere. For all measurements, the absorbance of the solu-
tions were around 0.1 at the excitation wavelength.
an anionic pyrene derivative upon complexation with o-BV2+
.
Obviously, the nature of the MLCT transition is not perturbed upon
complexation with the quenchers.
The quenching studies were done by taking the emission spec-
trum of the solution at a series of quencher concentration and
comparing the photoluminescence intensities (I). Stern–Volmer
constants (KSV) were obtained by linear fitting the experimental
data according to the relation:
I
I
0
¼
1 þ KSV½Cꢁ
ð1Þ
where I
0
is the photoluminescence in absence of quencher and (C)
the concentration of quencher. Titration curves against sugar were
D
fitted and dissociation constant (K ) values were obtained using
the relation:
ꢂ1
n
I
0
þ I
f
K
½Cꢁ
D
I ¼
ð2Þ
ꢂ1
n
1
þ KD ½Cꢁ
0 f
where I and I are the initial (no sugar) and final (plateau) intensi-
ties of the titrations curves. In case of an apparent 1:1 complexation
model, n = 1, and in the case of an apparent 1:2 complexation mod-
el, n = 2.
Frequency domain (FD) measurements were performed using
the instrumentation described previously [24–26]. Excitation was
provided by a blue LED (450 nm, Nishia Co.) through a blue filter
Fig. 2. Absorption and emission spectra of the Ru(II) metal–ligand-complex along
with the absorption spectrum of p-BV2
+
.

Md. Jamal Uddin et al. / Inorganica Chimica Acta 381 (2012) 104–110
107
Fig. 3. Absorption spectra of Ru-MLC, the p-BV2+ quencher (left) and the addition of the Ru-complex + p-BV2+ quencher (right).
Fig. 4. The spectral response of the Ru-MLC as the concentration of o-BV2+ increase.
Fig. 4 shows the spectrum response of the Ru-MLC as the con-
2
+
centration of o-BV increase. Corresponding Stern–Volmer plots
are displayed in Fig. 5A. As the concentration of quencher increase,
we observe a large decrease of the luminescence of the Ru-MLC
with no apparent shift. At low concentration of quencher, the
Stern–Volmer plots show linear relation with the concentration
(Fig. 5B). Both benzyl viologen derivative functionalized with the
boronic acid group show similar Stern–Volmer constant as methyl
2
+
viologen (MV ), showing that the presence of the benzyl and the
boronic acid groups does not alter the quenching property of the
Fig. 5. Stern–volmer plots for Ru-MLC and o-BV2+ quencher in water and PBS at RT
q
viologen moiety. Quenching rate constants (k ) calculated are
(
absence and presence of sugar).
one order higher than the quenching rate constant reported for
2
+
2+
ꢂ9
ꢂ1 ꢂ1
the system Ru(bpy)
3
/MV (k
q
= 0.74 ꢃ 10
m
s ) [22]. Since
diffusion coefficient is expected to be similar in both systems,
the higher quenching rate constant are expected due to the specific
interaction between the anionic Ru-MLC and the cationic quench-
er. This quenching rate constant can be enhanced by increasing the
number of negative charge on the MLC. For example, a Ru-MLC
bearing four negative charges shows a quenching rate constant of
ꢂ9
ꢂ1 ꢂ1
10.1 ꢃ 10
M
S . The MLC reported in this work (Fig. 1) posses
two negative charges, if the carboxylic acid group is considered
non-ionized.
At a higher concentration of quencher, an upward curvature of
the Stern–Volmer plot is observed showing the presence of static

1
08
Md. Jamal Uddin et al. / Inorganica Chimica Acta 381 (2012) 104–110
Stern–Volmer constant was obtained, while in presence of PBS,
5 mM, the quenching constant shows a decrease of about 4-fold.
2
This decrease continues as the concentration of PBS increases as
shown in Fig. 7.
Due to the limited solubility of the quencher at a high concen-
tration of PBS, we were not able to observe whether the upward
curvature at a high concentration is maintained or not.
Dynamic (K
D S
) and static (K ) quenching constants were evalu-
ated using the theory of static quenching, both constants were ob-
tained from the slope and intercept of a plot of ((I
0
/I) ꢂ 1)/(Q) vs (Q)
ꢂ1
[
23]. In water, a dynamic quenching constant of 16200 M was
ꢂ1
obtained with a static quenching constant of 1280 M . Both dy-
namic and static quenching constants were reduced in the pres-
ꢂ1
ꢂ1
D S
ence of PBS 25 mM, K = 3400 M and K = 800 M . Since the
static constant represents the stability constant of the complex
luminophore/quencher, the results show that in the PBS solution
the formation of complexes is greatly reduced.
Fig. 8 shows the frequency domain intensity decays of Ru-MLC is
decreased from 740 ns to 135 ns in H
quencher. We also investigated the effect of adding
2
O by adding 0.43 mM p-BV2+
D-fructose on
the FD intensity decays of Ru-MLC and the quenchers in PBS at RT.
The lifetime of Ru-MLC has decreased to 282 ns in the presence of
4
0 mM D-fructose. The change of the life time is relatively small,
more studies are on the way to link this decrease of lifetime with
electronic properties involved in the molecule following the binding
with sugars and the formation of the anionic form. Fig. 9 shows the
Stern–Volmer plots of F–D intensity decays for Ru-MLC and p-BV2+
2
quencher in H O and PBS. The plots are shown the linearity at lower
concentration. The Dynamic quenching constants obtained using
this model and the steady-state intensity are in agreement with
those obtained with the time-decay measurements, Figs. 8 and 9.
Fig. 6. Stern–volmer plots for Ru-MLC and o-BV2+ quencher in water and PBS at RT.
Note the linear pattern for low concentrations.)
(
5
. Are the dynamic quenching constants supposed to be the
quenching. The Stern–Volmer profile at a high concentration of p-
BV is not shown due to the low solubility of p-BV in phosphate
buffer solution.
Since the amplified quenching comes from an electrostatic
interaction between the luminophore and the quencher, it strongly
depends on the ionic strength of the media.
Fig. 6 shows the effect of the concentration of the phosphate
buffer solution (PBS) on the Stern–Volmer profiles. In water, a large
2
+
2+
same in water and in PBS 25 mM?
In the presence of D-fructose, D-fructose was chosen due to its
higher affinity to monoboronic acid derivative [11], the quenching
properties of the boronic acid functionalized benzyl viologen deriv-
atives are reduced (Fig. 5). Stern–Volmer constants are reduced to
ꢀ
2.8-fold and super linear quenching are suppressed at a higher
concentration of quencher. This reduction of the quenching is
due to the formation of a neutral zwiterionic viologen derivative,
C Scheme 1. The ionic complexation between the anionic Ru-MLC
and the neutral quencher are expected to be removed, resulting
only in quenching from diffusion.
This difference in the quenching property of the boronic acid
functionalized benzyl viologen derivatives in the presence of
sugar can be used to monitor the sugar concentration in the
solution. In this case, at a selected and fixed quencher concentra-
tion, amplified quenching can be observed in the absence of
sugar, while in the presence of sugar, this quenching is removed
and the intensity increases are observed. To study the sugar pro-
file response of both benzyl viologen derivatives, we used a
quencher concentration of 0.5 mM in a PBS pH 7.4 at concentra-
tion 25 mM. While these conditions are far to be optimized to
show the highest sugar response, we chose these conditions, first
because of the relatively low solubility of the quenchers and also
to have a non-zero ionic strength solution since potential appli-
cations will virtually be useful in biological media. Both the
quencher concentration and ionic strength of the solution greatly
affect the optical response of the system. As shown on Fig. 5A,
very large intensity changes, between the absence and the
presence of sugar can be obtained at a higher concentration of
quencher in the upward curvature of the Stern-Volmer plot. Also
as seen in Fig. 6A, this upward curvature increase as the ionic
Fig. 7. Stern–volmer plots for the effect of concentration of PBS at RT.

Md. Jamal Uddin et al. / Inorganica Chimica Acta 381 (2012) 104–110
109
Fig. 8. Frequency domain intensity decays of Ru-MLC and p-BV2+ quencher in water and PBS at RT (absence and presence of sugar).
ous solution have been studied by using two boronic acid function-
2
+
alized benzyl viologen (BV
) derivatives as quenchers. The
subsequent sugar effect is reported between this two quenchers
and ruthenium(II) metal–ligand-complex. The combination of elec-
tron withdrawing and/or donating group and boronic acid group
both directly linked to Ru MLC could lead to the formation of an ex-
3
cited MLCT state. After complexation with a sugar molecule boro-
nic acid changes from the neutral form to the anionic one and a
change in the MLCT properties of the Ru MLC occurs. A drastic
change in the fluorescence intensity has been observed as the con-
centration of quenchers increased. This leads to a new optical and
ratio metric approach for the analysis of sugar molecule using MLC
probes having the boronic acid group.
Acknowledgements
This work has been supported by the Juvenile Diabetes Founda-
tion International, 1-2000-546, and by the NIH National Center for
research Resources, RR-08119.
Fig. 9. Stern–volmer plots of FD intensity decays for Ru-MLC and p-BV2+ quencher
in water and PBS at RT.
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