A regenerative chemodosimeter based on metal-induced dye formation for the highly selective and sensitive optical determination of Hg2+ ions

Article abstract of DOI:10.1002/anie.200500583

(Chemical Equation Presented) Into the blue: Hg²⁺-ion-triggered formation of a brightly fluorescent squaraine dye is used as a novel approach for the design of highly selective and sensitive chromofluorogenic probes for the analysis of Hg²

Full text of DOI:10.1002/anie.200500583

Angewandte
Chemie
Analytical Chemistry
species, leading to the formation of a fluorescent or colored
product.[7,8] A disadvantage of dosimeters, however, is that
the reactions are mostly irreversible and thus provide only
single-use assays.
A Regenerative Chemodosimeter Based on
Metal-Induced Dye Formation for the Highly
Selective and Sensitive Optical Determination
of Hg2+ Ions**
To take advantage of the favorable features of chemo-
dosimeters and also reuse them, it is necessary to install a
procedure that allows the molecular reporter to be regen-
erated. We thus designed a method where the indicator dye is
passivated first in a chemical addition reaction with a small
organic compound, the “spectroscopic inhibitor” that
“switches off” the color and fluorescence of the indicator.
This addition product is the chemodosimeter. The target ion
then reacts with the inhibitor, liberating the dye, that is,
signaling is accomplished by “metal-induced dye release”
methods. With the proper choice of dye scaffold and
passivation reaction, it should thus be possible to generate
drastic chromo- and fluorogenic changes that result in a true
“switching-on” behavior rather than a modulation of already
existing signals or minor shifts of absorption or emission
bands.
We have applied this newly designed procedure to the
selective analysis of Hg2+ ions. The method is based on the
thiophilic affinity of the Hg2+ ion and its reactivity with 2,4-
bis(4-dialkylaminophenyl)-3-hydroxy-4-alkylsulfanylcyclo-
but-2-enone (APC) derivatives. As we have recently shown,
APC derivatives can be elegantly obtained by a simple
reaction of squaraines—the indicator dye—with thiols, the
inhibitor.[9] A representative compound (1) is shown in
Scheme 1. Compound 1 contains two independent subchro-
JosØ V. Ros-Lis, M. Dolores Marcos, Ramón Mµrtinez-
Mµæez,* Knut Rurack,* and Juan Soto
The Hg2+ ion is one of the environmentally most important
metal ions whose toxicity, even at very low concentrations, has
long been recognized and is a problem of primary concern.[1]
Moreover, despite a reduction of its industrial use as a result
of stricter regulations, high concentrations of mercury are still
present in many environmental compartments and it can still
be found in many products of daily life such as paints,
electronic equipment, and batteries.[2] Accordingly, the need
for analytical methods for the sensitive and selective deter-
mination of mercury is of topical interest, especially in
situations where conventional techniques are not appropriate,
for instance in many on-site or in situanalyses and for rapid
screening applications. In these fields, optical and electro-
chemical sensing devices play a leading role, in particular
utilizing molecular probes that generate and transduce an
analytical signal as a response to the binding event.[3]
Although several examples of redox-active,[4] fluorogenic,[5]
and chromogenic[6] chemosensors for Hg2+ ions have been
reported recently, the combined “binding site and signaling
subunit” approach that is commonly used with such probes
often harbors disadvantages for analysis in realistic media.
Many signaling units suffer through the strong hydrogen-
bonding ability of water, fluoroionophores often undergo
unspecific fluorescence quenching upon binding to heavy-
metal ions, and the number of Hg2+-ion-selective receptors is
limited.
A particularly attractive alternative presented herein are
chemodosimeters that indicate an analyte through a specific
chemical reaction between dosimeter molecule and target
Scheme 1. Derivatives 1 and 2.
[*] J. V. Ros-Lis, Dr. M. D. Marcos, Prof. R. Mµrtinez-Mµæez, Dr. J. Soto
Centro de Investigación en Química Molecular Aplicada
Departamento de Química
mophores that are electronically separated and chemically
bound through an sp3-hybridized carbon atom. Accordingly, 1
shows two overlapping bands in the UV spectral region; one
at approximately 265 nm which is characteristic for the
dialkylanilino moiety and a second, 1.3-fold more intensive
band centered at around 305 nm (Figure 1). The latter band
can be ascribed to the dialkylaminophenylhydroxycyclobut-2-
enone (PC) group. Quantum chemical calculations at the
semiempirical level are in agreement with the experimental
results and reveal the existence of two independent tran-
sitions of moderate oscillator strength, the HOMO–LUMO
transition that is located on the PC fragment and the
energetically higher-lying HOMO-1–LUMO + 1 transition
localized on the anilino group.
Universidad PolitØcnica de Valencia
Camino de Vera s/n, 46071 Valencia (Spain)
Fax: (+34)96-387-9349
E-mail: rmaez@qim.upv.es
Dr. K. Rurack
Div. I.3
Bundesanstalt für Materialforschungund -prüfung(BAM)
Richard-Willstätter-Strasse 11, 12489 Berlin (Germany)
Fax: (+49)30-8104-5005
E-mail: knut.rurack@bam.de
[**] We thank the Ministerio de Ciencia y Tecnología (projects REN2002-
04237-C02-01 and MAT2003-08568-C03-02) for support. J.V.R.L. also
thanks the Ministerio de Educación, Cultura y Deporte for a
Doctoral Fellowship.
Addition of Hg2+ ions to solutions of 1 results in a
dramatic change of color owing to the appearance of a new
Supportinginformation for this article is available on the WWW
under http://www.angewandte.org or from the author.
Angew. Chem. Int. Ed. 2005, 44, 4405 –4407
DOI: 10.1002/anie.200500583
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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Communications
As the coordination with thiols is not an exclusive feature
of mercury, the reactivity of 1 in the presence of other metal
ions was studied. In a typical experiment, up to 10 equivalents
of Hg2+, Cu2+, Fe3+, Pb2+, Ni2+, Cd2+, Zn2+, Al3+, and Tl+ were
added to a solution of 1 in water/acetonitrile (4:1, c1 = 6 ”
10À6 m, pH 9.6, CHES 0.01m). The results are shown in
Figure 2. There is a remarkably selective development of
Figure 1. Absorption spectra of 1 (c) and 3 (a) in water/acetoni-
trile (4:1 v/v), before and after the reaction of 1 with Hg2+ ions. The
gray bars denote the region of absorption of the single chromophores:
A=anilino moiety (this band is also present as a higher transition in
3), PC=aminophenylhydroxycyclobut-2-enone group and SQ=squar-
aine.
and intense absorption band at 642 nm that is typical for a
squaraine dye (Figure 1).[9,10] This Hg2+-ion triggered forma-
tion of squaraines is shown in Scheme 2. The reaction product
Figure 2. Absorption spectra of 1 (6”10À6 m) in water/acetonitrile (4:1
v/v, pH 9.6, CHES 0.01m) upon addition of 0.5 equivalents of various
metal ions. In the photograph, from left to right: Ni2+, Zn2+, Tl+, Fe3+
Hg2+, Pb2+, Al3+, Cd2+, Cu2+, and no metal ion.
,
the blue color only in the presence of Hg2+, while the other
potential competitors remain entirely silent. This chromo-
genic indication reaction allows Hg2+ ions to be detected
down to 20 ppb by using a conventional spectrophotometer
and standard conditions (such as 10-mm cells).
Despite the inherent interest of the colorimetric response,
fluorometric methods can reach much lower detection limits
with very low-cost and widely used instrumentation. Consid-
ering the high brightness (the product of e and Ff) of
squaraine dyes, the present system should be ideal for use in
conjunction with fluorescence detection. Thus, similar studies
Scheme 2. The analytical cycle: Hg2+ -ion-induced formation of 3 and 4
and regeneration of 1 and 2 by reaction with propanethiol.
to those described above were performed with c1 = 1 ” 10À7
m
on a conventional fluorometer. Figure 3 shows the fluores-
3 has a comparatively large molar absorptivity (lge = 5.05)
and a moderately high fluorescence quantum yield (Ff =
0.023) in water/acetonitrile (4:1). This dual chromo- and
fluorogenic sensing of Hg2+ ions is remarkable as both
absorption and emission are found at the red end of the
visible spectral window where usually the interference arising
from matrix absorption or autofluorescence is negligible.
Moreover, this dramatic hyper- and bathochromic shift upon
mercury-induced transformation of 1 into 3 allows for
straightforward “naked-eye” monitoring of the Hg2+-ion-
detection process.[11]
cence titration spectra of 1 with Hg2+ ions, revealing that the
To optimize the method, different pH ranges were tested
and a basic pH (9.6, buffered with 2-(cyclohexylamino)eth-
anesulfonic acid (CHES) 0.01m) was found to yield the best
performance of the chromofluorogenic dosimeter. Further-
more, these studies revealed that a maximum signal can be
observed at a molar ratio of 1/Hg2+ = 2:1, suggesting the
formation of Hg(SR)2 species (see Scheme 2).
Figure 3. Fluorescence titration spectra of 1 (10À7 m) with Hg2+ ions in
water/acetonitrile (4:1 v/v, pH 9.6, CHES 0.01m, lexc =642 nm). The
inset shows the titration curve obtained from the emission intensities
at 670 nm.
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Chemie
detection of less than 2 ppb of Hg2+, which is the U.S.
Environmental Protection Agencyꢀs limit for drinking water,
is clearly possible by the Hg2+-ion-induced release of the
highly fluorescent squaraine 3.
Motivated by these favorable features of the solution-
based chemistry, we took a step towards the use of metal-
triggered reactions for in situsensing and rapid screening
Keywords: chemodosimeter · dyes · fluorometric analysis ·
mercury · spectrophotometric analysis
.
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applications. For this purpose, the APC derivative
2
(Scheme 1) containing hydrophobic n-butyl chains was pre-
pared and adsorbed on powdered silica. The butyl chains offer
the required hydrophobicity to prevent the leaching of the
dye in aqueous samples. This rather simple method allowed
an accurate control of the amount of dye that is physisorbed
onto the solid. The loading is a key parameter for fluoro-
metric applications as too high loadings can easily lead to self-
quenching for dyes with narrow, intense, and weakly Stokes-
shifted absorption and fluorescence spectra.
In a series of experiments we found that a suitably
sensitive solid able to show dual chromofluorogenic signaling
should typically contain 2 in a concentration of 10À6 mmolgÀ1.
The powder was then fixed on a polyethyleneterephthalate
film for a first prototype of a dip-stick assay that was tested by
simply dipping the films into aqueous samples containing
Hg2+ ions. Once the sticks come into contact with the sample
solutions, the solid turns blue (labs = 642 nm) and begins to
fluoresce (lem = 670 nm). The response time is on the order of
a few seconds and the behavior is consistent with the reaction
shown in Scheme 2. Compound 2 reacts with the mercuric ion
to give the corresponding squaraine derivative 4 that remains
adsorbed on the solid, resulting in a chromo- and fluorogenic
“switching-on” process indicative of the presence of Hg2+
ions. Finally, having accomplished the reaction, the chemical
reaction described above[9] can be used to regenerate the
chemodosimetrical solid quantitatively with propanethiol
(Scheme 2). Upon addition of the “spectroscopic inhibitor”,
the conjugation in the squaraine is interrupted and the color
immediately turns from blue back to colorless; the device is
ready for the next cycle (see the Supporting Information).
In summary, we have designed a highly selective and
sensitive dual chromofluorogenic chemodosimeter system for
the determination of Hg2+ ions in aqueous environments
[5] For recent examples see: a) X. Guo, X. Qian, L. Jia, J. Am.
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[11] The ethoxyethyl substituents at the amino nitrogen of 1 and 3
were employed to increase the solubility of the compounds in
aqueous solution. The moiety itself does not respond to the
metal ions tested in this work in a classical fluoroionophore-type
fashion.
using a very simple and specific reaction, that is, Hg2+
-
triggered formation of a squaraine dye. The method allows
the selective detection of Hg2+ ions below 2 ppb in aqueous
solutions, shows a fluorescence enhancement, and displays a
very large shift in absorption with the development of a blue
color from colorless solutions. The possibility to adsorb or
anchor the probe on suitable supports allowed prototypes of
reusable dip-stick assays to be designed for rapid screening
for the target analyte. Additionally, we believe that this
procedure, based on guest-induced dye release methods using
suitable dye scaffoldings and passivation reactions, might be
of interest as a new route for the design of new and improved
regenerative sensor molecules for the rapid colorimetric
screening of a number of other target guests.
Received: February 16, 2005
Revised: April 8, 2005
Published online: June 9, 2005
Angew. Chem. Int. Ed. 2005, 44, 4405 –4407
www.angewandte.org
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
4407