A Ratiometric Fluorescent Probe for Gold and Mercury Ions

Article abstract of DOI:10.1002/chem.201502411

A fluorescent probe that displays a ratiometric fluorescence response towards gold and mercury ions has been devised. Emitting at a relatively longer wavelength, the conjugated form of the fluorescent dye transforms in the presence of the gold or mercury

Full text of DOI:10.1002/chem.201502411

DOI: 10.1002/chem.201502411
Communication
&
Sensors
A Ratiometric Fluorescent Probe for Gold and Mercury Ions
[
a]
Muhammed ÜÅüncü, Erman Karaku s¸ , and Mustafa Emrullaho g˘ lu*
Notably, most existing gold ion sensors are based on specific
chemical reactions that exploit the exceptional catalytic behav-
iour of gold species. In general, the optical signal is recognized
as either an increase (“turn-on”) or a decrease (“turn-off”) in
emission intensity without any noticeable change in emission
wavelength. Importantly, measurements based on intensity
changes are easily influenced by a host of environmental fac-
tors, including concentration variations and intensity of excita-
tion.
Abstract: A fluorescent probe that displays a ratiometric
fluorescence response towards gold and mercury ions has
been devised. Emitting at a relatively longer wavelength,
the conjugated form of the fluorescent dye transforms in
the presence of the gold or mercury ions into a new dye,
the molecular structure of which lacks the conjugation
and consequently emits at a distinctly shorter wavelength.
By contrast, measuring optical signals as intensity ratios at
two different wavelengths provides a built-in correction for the
environmental effects and may assuage many of the problems
associated with intensity-based sensors. Interestingly, the ratio-
metric recognition of gold ions by a single fluorescent probe
Gold catalysis has recently become a highly popular subject in
synthetic chemistry. Its popularity stems from the unique cata-
lytic properties of certain gold ion species, which have been in-
crementally exploited in the synthesis of complex molecular
[
7a,8b]
structure is currently uncommon.
Two of those examples
[
1]
structures. Though the unparalleled contribution of gold cat-
alysis to synthetic chemistry remains unquestionable, employ-
ing metal species such as gold as catalysts in chemical process-
es raises important health issues concerning the toxicity of
in recent literature are smart extensions of intensity-based sen-
sors that benefit from the FRET (i.e., fluorescence resonance
[
9]
energy transfer) principle to achieve a ratiometric response.
In response, we herein present the design, synthesis, spec-
tral behaviour, and living cell application of a fluorescent
probe, BURAK-1, which displays a sensitive, highly selective ra-
[
2]
gold ion species.
In contrast to gold’s elemental form, its ionic forms (i.e., Au
+
3
+
2+
and Au ) are sensitive, extremely reactive, and are able to in-
teract and bind with biomolecules, such as enzymes, proteins,
and DNA, thereby disturbing a series of cellular processes and
precipitating serious health problems. For instance, it has been
tiometric response to gold ions and, surprisingly, Hg ions as
well.
In our sensing approach, we drew inspiration from a gold-
catalyzed intramolecular cyclization reaction presented years
[
10]
documented that the intake of AuCl causes damage to vital
ago by Larock et al. (Scheme 1). It was reported that eny-
3
human organs, including the kidney and liver, as well as the
[2]
peripheral nervous system.
Considering the deleterious effects of gold species on living
organisms and their increasing role as catalysts in the chemical
industry, it is crucial for researchers to be able to assess the
levels of gold species in certain chemical, environmental, and
biological samples.
Recently, fluorescence-based techniques for sensing and
monitoring target species in solutions as well as in living envi-
[3]
ronments have received a great deal of attention. In this con-
text, several types of fluorescent probes have been devised for
[4]
analysing gold species. By extension, a variety of fluorophore
[
5]
core units such as rhodamine,
boron-dipyrromethene
[
6]
[7]
[8]
(
BODIPY), fluorescein, and naphthalimide have been judi-
Scheme 1. Work of both our and the Larock group.
ciously modified with specific molecular motifs to recognize
gold species through a distinct optical output: colorimetric
and/or fluorometric change.
none (1) in the presence of a catalytic amount of AuCl trans-
3
forms rapidly into a new furan (2) derivative. By extension, our
attention focused chiefly on exploiting this unique chemical
transformation as a signal-transducing event for the recogni-
tion of gold ions.
[
a] M. ÜÅüncü, E. Karaku s¸ , Prof. Dr. M. Emrullaho g˘ lu
Department of Chemistry, Faculty of Science
˙I zmir Institute of Technology, Urla, 35430, ˙I zmir (Turkey)
E-mail: mustafaemrullahoglu@iyte.edu.tr
It is well-known that extending the conjugation within a fluo-
rophore–chromophore structure dramatically affects the
Supporting information for this article is available on the WWW under
http://dx.doi.org/10.1002/chem.201502411.
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Communication
HOMO/LUMO levels of molecules and results in a dramatic red-
analysis. The green emissive compound, clearly followed on
the TLC plate, was isolated and further characterized by NMR
spectroscopy and HRMS as BOD-FUR, the cyclization product
[11]
shift of its absorption and emission band. With this in mind,
we integrated the enynone scaffold to a BODIPY-based fluoro-
phore dye with the expectation of generating a highly conju-
gated BODIPY derivative emitting at a wavelength distinctly
longer than its unmodified form (Scheme 1). We envisioned
that only in the presence of gold species would the sensor
structure transform itself into another BODIPY derivate lacking
the extended conjugation. Gold-ion-triggered structural modifi-
cation of the fluorescent dye was then anticipated to enable
us to recognize the presence of gold species as a ratio of two
distinct wavelengths, which is a primary principle of ratiomet-
ric sensing.
[
13]
3+
of BURAK-1 (Scheme 3). Evidently, the recognition of Au
3
+
was based on an Au -mediated cyclization reaction that re-
sulted in the formation of a highly emissive BODIPY-furane de-
rivative (BOD-FUR).
3
+
The systematic titration of BURAK-1 with Au revealed that
the ratiometric change of emission intensities at both wave-
lengths linearly correlated with the increased concentration of
3
+
Au in the range of 0.005–10 mm. At the same time, our kinet-
ic study showed that the spectral response toward the addi-
3
+
tion of Au was rapid (<1 min) and that emission intensity at
The title compound BURAK-1 investigated in this study
was prepared according to the synthetic route outlined in
Scheme 2. Following a Sonogashira coupling protocol, the
516 nm plateaued within 40 min due to the addition of
3
+
2 equiv of Au , which thereby enhanced intensity at 516 nm
3
+
by more than 70-fold. Moreover, the minimum amount of Au
detectable was evaluated to be
[
13]
8
nm, one of the lowest detec-
tion limits reported in fluores-
[
4a]
cence-based gold ion sensing.
Meanwhile, similar spectral
a
trend was observed in the pres-
+
ence of Au species, revealing
that the probe operates effi-
ciently for both oxidation states
of gold in a nondiscriminative
manner.
Scheme 2. Synthesis of BURAK-1.
acetylene derivative of BODIPY (BODIPY-AC) prepared in three
[
12]
[10]
individual steps was coupled with 2-iodocyclohex-2-enone
to give the desired probe structure in a moderate yield. The
structure of the probe was clearly confirmed by NMR spectro-
[13]
scopic and HRMS analysis.
The probe’s spectroscopic behaviour toward the added
metal species was systematically investigated with the aid of
ultraviolet (UV) and fluorescence spectroscopy. As depicted in
Figure 1a, the UV/Vis spectrum of free BURAK-1 (phosphate
buffer/ethanol 6:4, pH 7.0) displays a maximum absorption
band at 526 nm, while its fluorescence spectrum collected
upon excitation at 460 nm exhibits an intense emission band
at 562 nm, which belongs to the BODIPY chromophore.
Our investigation resumed with an evaluation of the optical
3
+
behaviour of BURAK-1 in response to the addition of Au
ions (e.g., AuCl ). The spectral changes of the probe in the ab-
3
3
+
sence and presence of Au ions appear in Figure 1. As shown,
3
+
the addition of Au (2 equiv) to BURAK-1 prompted the ap-
pearance of a new emission band at 516 nm, with a concomi-
tant decrease in the emission band at 562 nm. As anticipated,
when the probe solution was treated with AuCl , the orange-
3
emitting probe solution became distinctly green, as was clearly
visible to the naked eye (see Figure S28 in the Supporting In-
formation). The dramatic change in colour of the solution was
attributed to a change of the BODIPY dye structure. Moreover,
green emission was evidence of the existence of a nonconju-
gated BODIPY derivative.
Figure 1. a) Absorbance and b) fluorescence titration spectra of BURAK-1
This suggestion was supported by the outcome of the reac-
3+
(5 mm) + Au (0.005 to 10 mm) in 0.1m phosphate buffer/EtOH (pH 7.0, v/v,
3
+
tion of BURAK-1 mediated by Au , controlled by using TLC
6:4).
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Further experiments on im-
proving the selectivity of the
probe toward one distinct metal
3
+
species, that is, either Au
or
Hg , revealed that the selectivi-
2
+
2
+
ty toward Hg could be signifi-
cantly improved by adjusting
the nature of the sensing media.
Remarkably, in an alternative sol-
vent system, namely, a HEPES/
acetonitrile (pH 7.0) buffer (6:4
Scheme 3. Gold-mediated cyclization of BURAK-1.
2
+
(
v/v)), the probe displayed exceptional selectivity toward Hg
ions. Importantly, in HEPES/acetonitrile, particularly at high
3
+
contents of water (e.g. 6/4; v/v), Au ions were rapidly re-
duced to gold black (aggregates of elemental gold), which in-
dicates considerable loss of catalytic activity and accounts for
2
+
the selectivity towards Hg . Under these sensing conditions,
2
+
[13]
the detection limit for Hg was evaluated to be 250 nm.
Furthermore, no cross-talk with other metal species in the de-
2
+
tection of Hg was detected. Lastly, BURAK-1 shows condi-
tion-dependent selectivity toward Hg species and can be
used on-demand as a sensitive, selective fluorescent probe for
2
+
2
+
Hg ions.
As consistent with findings reported in the literature, the
3
+
2+
recognition of Au or Hg is suggested to proceed by a con-
certed process initiated by the activation of the triple bond by
the metal species (Scheme 4). This process follows a conjugate
addition of an oxygen nucleophile that promotes intramolecu-
lar cyclization and yields a new BODIPY structure appended
with a furan motive, which displays the distinct colour and
emission of the solution.
3
+
Figure 2. Fluorescence spectra of BURAK-1 (5 mm), BURAK-1 (5 mm) + Au
Au /Hg (25 mm, 5 equiv), BURAK-1 (5 mm) + other metal ions (50 mm,
/
+
2+
1
0 equiv) in 0.1m potassium phosphate buffer, pH 7.0/EtOH (v/v, 6:4)
(
lex =460 nm, at 258C). Inset: Bar graph notation.
The selectivity profile of BURAK-1 was surveyed by screening
Since BURAK-1 constitutes all of the desirable features nec-
essary for tracking species in a living milieu, we assessed its
sensing capacity in living cells. To this end, human colon carci-
noma cells (A-549) were incubated first with the probe
2
+
the spectral response toward metal species, including Zn
,
,
2
+
2+
2+
+
+
2+
2+
2+
3+
2+
2+
Cd , Ba , Cu , Li , K , Ni , Cr , Mg , Fe , Pb , Hg
2
+
+
2+
Co , and Ag (Figure 2). Surprisingly, in the presence of Hg
3
+
2+
ions the probe displayed the exact sensing behaviour as in the
detection of gold species. Similar to the gold ion sensing event,
(10 mm), to which was added Au or Hg (10 mm) to be incu-
bated for another 60 min. The cells were also stained with a nu-
cleus-staining dye (DAPI) for another 10 min. With the aid of
2
+
with the addition of Hg
a new peak band appeared at
5
16 nm, while the band at 562 nm decreased with an
2
+
increased concentration of Hg ions. This unexpect-
2
+
ed observation was reasonable, for Hg ions similar
3
+
to Au species are known to have high affinities to
2
+
alkynes. Notably, the detection limit for Hg
measured to be slightly higher than that for Au
was
3
+
[13]
ions (60 nm) yet still at nanomolar levels. From the
competition experiment, BURAK-1 was ultimately
shown to display a dual nature and to efficiently op-
+
3+
2+
erate for two metal species: Au /Au and Hg ions.
Having clarified the nature of detecting both metal
species, we next assessed the possible interference of
3
+
other metal species in the detection of Au and
2
+
Hg . As shown in Figures S7 and S14 (Supporting In-
formation), the response of BURAK-1 toward both
species remained unaffected in the presence of other
competitive metal species. These results established
3
+
that BURAK-1 can also properly detect Au
and
2
+
Hg ions in mixtures of other related species.
Scheme 4. Proposed reaction mechanism for the detection of gold ions.
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fluorescence microscopy, the fluorescence images of the cells
were taken before and after the addition of the metal species.
As Figure 3 clearly shows, the cells incubated with BURAK-
Experimental Section
General method
1
emitted a red fluorescence in the absence of the metal spe-
[PdCl (PPh ) ] (3.51 mg, 0.05 equiv), CuI (1.9 mg, 0.1equiv), and
2 3 2
2
+
3+
cies, while following Hg or Au accumulation they emitted
BODIPY-AC (69.6 mg, 2.0 equiv) were added to a mixture of 2-
iodo-2-cyclohexen-1-one (22.1 mg, 0,1 mmol) in THF at 08C. Then,
diisopropylamine (42 mL, 3.0 equiv) was added and the resulting
mixture was stirred at 08C for 1 h. After completion of the reaction,
THF was removed under vacuum and the resulting residue extract-
ed three times with CH Cl (3”30 mL). The organic layer was dried
2
2
over MgSO , filtered, and concentrated. The resultant residue was
4
purified by column chromatography (4:1 (hexane/ethyl acetate)) to
1
afford BURAK-1 as a red solid (31 mg, 70% yield). H NMR
(
400 MHz, CDCl ): d=7.49–7.48 (m, 3H), 7.27–7.22 (m, 2H), 6.02 (s,
3
1
6
H), 2.66 (s, 3H), 2.56 (s, 3H), 2.52–2.45 (m, 4H), 2.04 (quint. J=
.4 Hz, 2H), 1.47 (s, 3H), 1.18 ppm (s, 3H); C NMR (100 MHz,
13
CDCl ): d=195.1, 157.2, 156.5, 152.2, 144.4, 142.9, 141.9, 134.4,
3
1
2
32.1, 130.0, 129.0, 127.6, 125.3, 121.8, 114.7, 90.0, 84.7, 37.9, 26.2,
2.2, 14.5, 14.2, 13.2, 12.9 ppm.
Acknowledgements
˙
˙
We thank Izmir Institute of Technology (IZTECH) and TUBITAK
for financial support.
Keywords: BODIPY · fluorescent sensors · gold · mercury ·
ratiometric
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7
Received: June 22, 2015
Published online on July 29, 2015
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