Development of a borondipyrromethene-based Zn2+ fluorescent probe: Solvent effects on modulation sensing ability

Article abstract of DOI:10.1039/c1dt10797f

A borondipyrromethene-based Zn²⁺ fluorescent probe BODPAQ was designed and synthesized. The chelators in BODPAQ, 2,2′-dipicolylamine (DPA) and 8-aminoquinoline (AQ), coordinate to Zn²⁺ in a synergic manner. As a result, BODPAQ displa

Full text of DOI:10.1039/c1dt10797f

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PAPER
Development of a borondipyrromethene-based Zn2+ fluorescent probe: solvent
effects on modulation sensing ability†
Chunchang Zhao,* Yulin Zhang, Peng Feng and Jian Cao
Received 29th April 2011, Accepted 3rd October 2011
DOI: 10.1039/c1dt10797f
A borondipyrromethene-based Zn2+ fluorescent probe BODPAQ was designed and synthesized. The
chelators in BODPAQ, 2,2¢-dipicolylamine (DPA) and 8-aminoquinoline (AQ), coordinate to Zn2+ in a
synergic manner. As a result, BODPAQ displays high Zn2+ selectivity with a dramatic enhanced
emission accompanied by a notable hypsochromic shift due to the binary inhibition effect of PET and
ICT mechanisms, enabling the detection of Zn2+ by both ratiometric and normal turn-on fluorescence
methods in acetonitrile. Interestingly, the sensitivity of BODPAQ towards Zn2+ changes upon varying
the compositions of buffer solutions. In 3-morpholinopropanesulfonic acid (MOPS) buffer aqueous
solution (50% CH3CN), BODPAQ displays the highest sensitivity for Zn2+, while in citrate–phosphate
buffer, BODPAQ shows no response to Zn2+.
exhibit a spectral shift upon binding to Zn2+ can eliminate most
or all ambiguities by self-calibration of two emission bands.
To date, only a few ratiometric probes for Zn2+ have been
reported.10
Introduction
Fluorescent probes are expected to be powerful tools for the
analysis of many biologically relevant heavy transition-metal
ions because of the simplicity, high sensitivity and high spa-
tial resolution of fluorescence.1 Among heavy transition-metal
ions, Zn2+ is an attractive target in the design of fluorescent
probes due to the significant biological roles of zinc. Zinc plays
vital roles in catalytic functions of proteins, gene expression,
apoptosis, neurotransmission, and so forth.2 It is also known
that disorders of Zn2+ metabolism are associated with physical
growth retardation and neurological diseases such as cerebral
ischemia and Alzheimer’s disease.3 Therefore, measurement of
Zn2+ is essential but challenging due to the biological significance
and the interference of other metals in biological systems. High
sensitivity and selectivity are essential properties that an ideal
fluorescent probe for zinc should possess. Accordingly, there have
been considerable efforts to develop fluorescent sensors for Zn2+.4
The most currently available small molecule fluorescent probes
detect Zn2+ by an increase or decrease of the emission intensity,5
derived mainly from aryl sulfonamides,6 and bulk xanthenone
fluorophores.7,8 However, as the change in fluorescence intensity
is the only detection signal, factors such as the probe concen-
tration, instrumental efficiency, and environmental conditions
can interfere with the signal output.9 It is therefore desirable to
eliminate the effects of these factors. Ratiometric sensors that
Although a variety of fluorescent sensors for Zn2+ have been
developed successfully, based on photo-induced electron transfer
(PET) or intramolecular charge transfer (ICT) mechanisms, few
fluorescent sensors with both PET and ICT sensing behaviors have
been constructed. Given the successful application of PET or ICT
in designing fluorescent probes,5–10 we envisioned that combination
of the two sensing behaviors could be exploited as an interesting
platform for fluorescent Zn2+ probes. Here, we decided to employ a
combination of the key features of ICT and PET in our ratiometric
probe design. Based on this strategy, a new borondipyrromethene-
based ratiometric probe BODPAQ with both PET and ICT sensing
behaviors was reported. The probe was designed by combining two
chelators, 2,2¢-dipicolylamine (DPA) and 8-aminoquinoline (AQ),
into one borondipyrromethene molecule. Given the successful
application of DPA7,8d,8e and 8-aminoquinoline7g,11 derivatives in
Zn2+ sensing, we expected a combination of DPA and quinoline to
be advantageous. These two receptors are expected to coordinate
to metals in a synergic manner, thus improve the Zn2+ selectivity.
The binding of Zn2+ to BODPAQ induced not only emission en-
hancement but also an emission shift due to the binary inhibition
effect of PET and ICT mechanisms, enabling the detection of Zn2+
by both ratiometric and normal turn-on fluorescence methods. It is
known that compositions of solutions affect the binding ability of
fluorescent probes to analytes. Changing the solvent from organic
to aqueous solutions, the response can be significantly affected by
the compositions of buffer solutions and the emission response can
completely vanish. The present study also demonstrates in detail
how solvent effects modulate the binding affinity and selectivity
of BODPAQ towards Zn2+.
Key Laboratory for Advanced Materials and Institute of Fine Chemicals,
East China University of Science & Technology, Shanghai, 200237, P. R.
China. E-mail: zhaocchang@ecust.edu.cn; Tel: +86 6425 2388
† Electronic supplementary information (ESI) available: List of materials
and general experimental methods; NMR spectra of the new compounds;
Absorption and emission spectra of BODPAQ in aqueous solutions;
Ratiometric calibration curve. See DOI: 10.1039/c1dt10797f
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NMR (100 MHz, CDCl3) d 161.4, 157.5, 147.60, 136.8, 135.5,
134.7, 133.7, 133.0, 132.3, 131.1, 129.7, 128.4, 128.0, 122.3, 121.8,
110.1, 58.2, 58.1, 17.2, 14.8, 12.3, 11.4; HRMS (ESI) calcd for
C31H29N5BF2: 520.2484. Found: 520.2474 [M - H]-.
Experimental
General information
All chemicals were purchased from commercial suppliers unless
otherwise specified. Et3N, chloroform and 1,2-dichloroethane
were used as received without further purification. Anhydrous
N,N-dimethylformamide (DMF) and toluene were dried and
distilled immediately prior to use. 2-chloro-5-benzoyl-pyrrole12
and 2,4-dimethyl-3-ethylpyrrole12 were prepared according to
literature procedures.
1H NMR and 13C NMR spectra were recorded on a spectrome-
ter operating at 400 MHz and 100 MHz, respectively. Deuterated
chloroform was used as the solvent and TMS as the internal
standard. Mass spectra were measured on an HP 1100 LC-MS
spectrometer. UV-vis spectra were measured using a Shimadzu
UV-2550 spectrophotometer. Fluorescence spectroscopic mea-
surements were conducted on a Varian Cary Eclipse fluorescence
spectrophotometer.
Synthesis of 2-formyl-3-(4-(bis(pyridine-2-ylmethyl)amino))-5,7-
dimethyl-6-ethyl-8-phenyl-BODIPY (4). A mixture of DMF
(10 mL) and POCl3 (10 mL) was stirred in an ice bath for 5 min
under argon. After warming to room temperature, it was stirred
for an additional 1 h. A solution of 3 (0.47 g, 0.90 mmol) in 10 mL
chloroform was then added, and the resulting mixture was stirred
for 2 h at room temperature. The reaction mixture was cooled
to room temperature and slowly poured into saturated aqueous
Na2CO3 (30 mL) under ice-cold conditions. After warming to
room temperature, the reaction mixture was further stirred for
30 min and washed with water. The organic layers were dried over
anhydrous Na2SO4, and evaporated in vacuo. The crude product
was further purified using column chromatography (alumina gel,
EtOAc/hexane = 1 : 3) to give 4 (0.45 g, 91%). 1H NMR (400 MHz,
CDCl3) d 9.67 (s, 1H), 8.53 (d, 2H), 7.73–7.67 (m, 2H), 7.58 (s,
1H), 7.56 (s, 1H), 7.52–7.46 (m, 3H), 7.36–7.33 (m, 2H), 7.21–7.16
(m, 2H), 6.73 (s, 1H), 4.88 (s, 4H), 2.67 (s, 3H), 2.38 (q, 2H), 1.45
(s, 3H), 1.03 (t, 3H); 13C NMR (100 MHz, CDCl3) d 185.0, 157.8,
149.3, 140.9, 140.3, 136.7, 133.6, 131.6, 131.5, 129.4, 129.1, 128.5,
128.4, 124.0, 123.9, 122.8, 122.3, 59.3, 59.2, 17.2, 14.2, 13.4, 12.2;
HRMS (ESI) calcd for C32H29N5OBF2: 548.2433. Found: 548.2457
[M - H]-.
For absorption or fluorescence measurements, compounds were
dissolved in DMSO to obtain stock solutions (2–5 mM). These
stock solutions were diluted with CH3CN or aqueous solutions to
the desired concentration.
Synthesis of 3-chloro-5,7-dimethyl-6-ethyl-8-phenyl-BODIPY
(2).
A
mixture of 2-chloro-5-benzoyl-pyrrole (0.555 g,
2.71 mmol) and POCl3 (0.75 mL) in 30 mL CH2Cl2 was stirred
for 1 h at room temperature. To this reaction mixture was added
2,4-dimethyl-3-ethylpyrrole (1.000 g, 8.12 mmol) in CH2Cl2, and
the mixture was further stirred for 36 h. The reaction mixture was
slowly poured into saturated aqueous NaHCO3 (50 mL) under ice-
cold conditions, washed with water, dried over anhydrous Na2SO4,
and evaporated in vacuo. The residue was dissolved in toluene
before Et3N (1 mL) was added and the mixture was stirred for
1 h at room temperature. BF3·OEt2 (0.9 mL, 6.62 mmol) was
Synthesis of 2-(quinolin-8-ylaminomethyl)-3-(4-(bis(pyridine-2-
ylmethyl)amino))-5,7-dimethyl-6-ethyl-8-phenyl-BODIPY (BOD-
PAQ). To 20 mL 1,2-dichloroethane was added 4 (0.55 g,
1 mmol) and 8-aminoquinoline (0.15 mg, 1.03 mmol). The reaction
was stirred for 24 h at room temperature. A portion (0.26 g,
1.23 mmol) of NaB(OAc)3H was then added and the reaction
mixture was stirred overnight. The solvent was removed under
vacuum, and the residue was purified by flash chromatography
(alumina gel, eluent: hexane/EtOAc 5 : 1) to afford BODPAQ
(0.170 g, 25%). 1H NMR (400 MHz, CDCl3) d 8.65–8.62 (d, 1H),
8.49 (d, 2H), 8.06–8.02 (m, 1H), 7.70 (s, 1H), 7.68 (s, 1H), 7.64–
7.58 (m, 2H), 7.41–7.27 (m, 7H), 7.10 (t, 2H), 7.04 (d, 1H), 6.53
(d, 1H), 6.39 (s, 1H), 4.85 (s, 4H), 4.14 (s, 2H), 2.62 (s, 3H), 2.36
(q, 2H), 1.38 (s, 3H), 1.01 (t, 3H); 13C NMR (100 MHz, CDCl3) d
159.1, 158.5, 154.0, 149.1, 146.7, 144.5, 138.2, 137.9, 137.1, 136.7,
135.9, 134.7, 133.2, 131.4, 130.6, 130.4, 129.4, 128.7, 128.5, 128.10,
127.7, 124.3, 122.9, 122.1, 121.2, 114.3, 105.7, 58.6, 41.0, 17.2, 14.6,
12.8, 11.8; HRMS (ESI) calcd for C41H39N7BF2: 678.3328. Found:
678.3338 [M + H]+.
◦
added via syringe and the reaction was stirred at 100 C for 6 h.
After cooling, the reaction mixture was diluted with chloroform
and washed with saturated aqueous NaHCO3 solution and brine,
dried over Na2SO4, and evaporated. The residue was purified by
column chromatography (silica gel, 1 : 50 EtOAc/hexane) to give 2
(0.336 mg, 42%) as an orange crystalline solid. 1H NMR (CDCl3,
400 MHz): d 7.55–7.47 (m, 3H), 7.35–7.30 (m, 2H), 6.24 (d, 1H),
6.21 (d, 1H), 2.63 (s, 3H), 2.36 (q, 2H), 1.43 (s, 3H), 1.04 (t, 3H);
13C NMR (100 MHz, CDCl3) d 163.0, 142.0, 140.4, 136.7, 136.4,
136.3, 133.6, 133.5, 133.3, 129.4, 128.9, 128.5, 126.0, 114.9, 114.8,
17.13, 14.2, 13.3, 12.3; HRMS (ESI) calcd for C19H19N2BClF2:
359.1298. Found: 359.1285 [M + H]+.
Synthesis of 3-(4-(bis(pyridine-2-ylmethyl)amino))-5,7-dimethyl-
6-ethyl-8-phenyl-BODIPY (3). To a solution of 2 (0.62 g
1.73 mmol) in 30 mL CH3CN was added DPA (0.62 g, 2.60 mmol)
and Et3N (5 mL), and the reaction mixture was refluxed for 12
h. Excess CH3CN was removed under vacuum, and the residue
was dissolved in ethyl acetate, washed with H2O and dried over
Na2SO4. The crude product was purified by flash chromatography
(silica gel, eluent: hexane/EtOAc 3 : 1) to afford 3 (0.630 g, 69%).
1H NMR (400 MHz, CDCl3) d 8.52 (s, 1H), 8.51 (s, 1H), 7.69–7.63
(m, 2H), 7.48 (s, 1H), 7.46 (s, 1H), 7.44–7.39 (m, 3H), 7.33–7.30
(m, 2H), 7.19–7.15 (m, 2H), 6.35 (d, 1H), 5.94 (d, 1H), 5.09 (s,
4H), 2.49 (s, 3H), 2.34 (q, 2H), 1.36 (s, 3H), 0.98 (t, 3H); 13C
Results and discussion
Synthesis
The designed probe BODPAQ was synthesized in 5 steps as
shown in Scheme 1. DPA was firstly introduced into the BODIPY
core by SN2 nucleophilic substitution. Subsequent formylation of
the 2-position of the BODIPY core was achieved by Vilsmeier–
Haack reactions using DMF/POCl3 in chloroform.13 Conden-
sation of 1 equiv of 8-aminoquinoline with monoaldehyde 4 in
1,2-dichloroethane gave essentially pure intermediate imine, and
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The pH effect on the fluorescence spectra was investigated in
acetonitrile/water (50 : 50, v/v) solution (Fig. 1). The fluorescence
spectrum of BODPAQ exhibits an emission band with a maximum
at 579 nm, a small (2 nm) blue shift compared to that in CH3CN.
The emission intensity at 579 nm displays little change between pH
6 and 10. However, a distinct emission hypsochromic shift from
579 nm to 560 nm can be observed when increasing the acidity from
pH 6 to 5. This phenomenon could be attributed to the protonation
of the tertiary amine in DPA, which inhibits the ICT interaction
and results in a blue-shift in emission. Further increasing the
acidity from pH 5 to 3, a dramatic emission enhancement occurs
due to the reduced PET process by protonation of the aromatic
amine in AQ. A similar protonation process has been observed
in the DPA-based sensors.9d,10c The process was further studied in
detail by 1H NMR titration experiments of BODPAQ with HCl in
CD3CN (Fig. 2). As shown in Fig. 2, protonation with 1 equiv. HCl
triggers a clear downfield shift up to Dd = 0.31 ppm for Ha. The
chemical shift of Hc, in the ortho position of the pyridine ring,
also underwent a downfield shift from 8.54 to 8.60 ppm. These
results hint that the proton should firstly bind to the nitrogen on
the DPA group. Notably, upon increasing HCl to more than 5
equiv., the chemical shift of Hd¢, adjacent to the amino group in
quinoline, moved from 6.52 to 7.13 ppm (Dd = 0.61 ppm), which
clearly confirms the protonation of the aromatic amine in AQ.
Scheme 1 Synthesis of BODPAQ.
reduction of the imine under mild conditions using NaBH(OAc)3
in DCE gave BODPAQ in 25% yield.7g,14
Spectroscopic properties of BODPAQ and its pH dependence
The spectroscopic evaluation of BODPAQ was firstly performed
in organic solvents of varying polarity. The optical features
are characteristics of the BODIPY platform. As is evident
from Table 1, BODPAQ shows almost identical absorption and
fluorescence spectra in the various solvents.15 The main S0–S1
absorption transition band, centered between 553 and 563 nm in
the pure solvents, shows minor solvent-dependent variation and
the maximum shifts of the absorbance are only 10 nm. Similar
to the absorption spectra, the emission spectra also show minor
solvent-dependent shifts. The maximum emission wavelength of
BODPAQ is in the 578–587 nm range and the emission shows
low fluorescence quantum yield (<0.03), especially in acetonitrile
and methanol, where the fluorescence quantum yield is lower than
0.007.
The spectroscopic characteristics of BODPAQ were then eval-
uated in a series of aqueous solutions with varying acetonitrile
content (Fig. S1, ESI†). BODPAQ exhibits a strong absorption
band in the visible region centered at 560 nm in acetonitrile,
and the maximum undergoes a slightly blue-shift from 560 nm
to 552 nm with decreasing concentrations of CH3CN in aqueous
solutions from 100% to 10%. Emission spectral changes caused
by the concentration of acetonitrile are in good agreement with
the hypsochromic shift in the absorption spectra. Upon excitation
at 560 nm, an emission spectrum with a maximum at 581 nm
was observed in CH3CN, and the emission band is finally shifted
to 562 nm when excited at 552 nm in aqueous solution (10 mM
Tris-HCl, 10% CH3CN, 0.1 M KNO3).
Fig. 1 Effect of pH on the fluorescence of BODPAQ (5 mM) in
acetonitrile/water (50 : 50, v/v) solution, excitation wavelength: 530 nm.
Spectroscopic response of BODPAQ to Zn2+ in organic solvents
The spectra of BODPAQ are stable at around pH 7.0, so the
titration experiments of BODPAQ with Zn2+ were carried out at
pH 7.4. In acetonitrile (Fig. 3 and Fig. S2, ESI†), upon gradual
addition of Zn2+, a decrease in the absorption band at 560 nm and
a concomitant increase of a new band at 520 nm were observed
with a distinct isosbestic point at 530 nm; at the same time
the color of the solution turned from purple to light pink. A
significant blue shift of 40 nm of the absorption wavelength implies
the coordination of the tertiary amino group in DPA to Zn2+,
which decreases the electron-donating ability of the tertiary amino
group and leads to a blue-shift in absorption.9d The absorption
bands at 560 nm and 520 nm change linearly up to a 1 : 1 ratio
(Zn2+/BODPAQ), which is consistent with the formation of a
1 : 1 complex. The Kd of a 1 : 1 Zn2+/BODPAQ complex was
determined to be 1.81 ¥ 105 using Benesi–Hildebrand plots (see
ESI†).
Table 1 Spectroscopic data for BODPAQ in various solvents
Solvent
labs (nm)
lem (nm)
UF
Methanol
Acetonitrile
Acetone
DMSO
Dichloromethane
Benzene
553
560
558
562
559
563
560
575
581
582
587
586
587
582
0.005
0.007
0.015
0.013
0.019
0.027
0.022
Hexane
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of the newly formed complex is blue-shifted by 60 nm and is
responsible for the change in color from purple to light orange.
This color change can be used for a “naked-eye” detection of Zn2+
in acetonitrile (Fig. 2c). Moreover, the ratios of the absorbance
at 498 and 560 nm showed sigmoidal dependence on the [Zn2+]
concentration, with a 227-fold ratiometric enhancement (Fig. 2b).
This indicates the capability of BODPAQ for detecting [Zn2+] by
absorption ratiometry.
In the fluorescence spectra (Fig. 4), free BODPAQ exhibits
fluorescence with a maximum at 581 nm and quantum yield of
0.007, which is lower than that of compound 3 (0.02) because of
the PET quenching from the AQ amino group. Upon addition
of Zn2+ from 0 to 1 equiv., the lmax undergoes a blue shift to
em
549 nm accompanied by a remarkable fluorescence enhancement
(quantum yield 0.04), indicating that a inhibited PET and
ICT process occurred with consequent ratiometric and turn-on
fluorescence signals although both Ufree and UZn are not high.
Both the emission intensity at 549 nm and the intensity ratio, R
(F549/F581, Fig. S3, ESI†), increase upon gradual addition of Zn2+,
which allows the detection of Zn2+ by both ratiometric and normal
turn-on fluorescence methods. Continuous addition of Zn2+ from
1 to 4 equiv. leads to further dramatic enhancement of the emission
intensity, and the fluorescence turns out to be stable when 4 equiv.
Zn2+ was added; higher [Zn2+] does not lead to any change. As
[Zn2+] increases from 1 to 4 equiv., the emission wavelength is also
hypsochromically shifted from 549 nm to 535 nm, resulting from
the stronger interaction of Zn2+ with the tertiary amino group in
Fig. 2 Part of the 1H NMR spectra of BODPAQ and BODPAQ + HCl
(1, 4, 6 equiv.) in CD3CN.
Fig. 3 (a) Absorption spectra of BODPAQ (5 mM) upon addition of Zn2+
at 0, 0.2, 0.4, 0.6, 0.8, 1.0 equiv. with respect to BODPAQ in CH3CN. (b)
Absorption spectra of BODPAQ (5 mM) upon addition of Zn2+ at 1.0,
1.2, 1.4, 1.6, 1.8, 2.0, 2.4, 2.8, 3.5, 4.0 equiv. with respect to BODPAQ in
CH3CN. (c) Ratio of absorbance at 498 nm and absorbance at 560 nm as a
function of Zn2+ concentration. Inset: Color of solutions of BODPAQ in
the presence of Zn2+ (right: BODPAQ, middle: BODPAQ+Zn2+ (1 equiv.),
left: BODPAQ+Zn2+ (4 equiv.).
With higher [Zn2+] (1–4 equiv.), a significant decrease at 520 nm
was observed accompanied by the hypsochromic shift to 498 nm.
These results can be explained in terms of the strength of the
interaction between Zn2+ and the tertiary amino group in DPA.
Higher [Zn2+], greater than 1 equiv., causes stronger interaction
and leads to a further blue-shift in the spectra, suggesting another
type of complex other than the 1 : 1 type is formed. The lmax
Fig. 4 (a) Emission spectra of BODPAQ (5 mM) upon addition of Zn2+
(0, 0.2, 0.4, 0.6, 0.8, 1.0 equiv. with respect to BODPAQ) in CH3CN; (b)
Emission spectra of BODPAQ (5 mM) upon addition of Zn2+ (0, 0.4, 1.0,
1.2, 1.6, 2.6, 3.2, 3.6, 4.0 equiv. with respect to BODPAQ) in CH3CN.
Excitation wavelength: 530 nm.
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the newly formed complex other than the 1 : 1 complex. It is worth
noting that the newly formed complex is highly fluorescent with
quantum yield around 0.20, which makes BODPAQ a promising
turn-on fluorescent probe for [Zn2+].
Other organic solvents were also used to study the binding
properties of BODPAQ with Zn2+. BODPAQ responds to Zn2+
in the same way in methanol or ethanol as it does in CH3CN (Fig.
S4–S5, ESI†). Upon titration with Zn2+, the absorption spectra
show the evident hypsochromic shift from 552 nm to 514 nm with
[Zn2+] up to 1 equiv. More [Zn2+] also induced the new absorption
band at 514 nm to shift to 505 nm, but the shift is less pronounced.
With increasing [Zn2+], the lmaxem also undergoes a blue shift from
575 nm to 544 nm, accompanied by a significant fluorescence
enhancement (quantum yield changes from 0.005 to 0.1).
The possible 1 : 1 binding model of Zn2+ and BODPAQ in
acetonitrile therefore can be deduced and is shown in Scheme
2, which is further confirmed by mass spectrometry. The ESI mass
spectrum of Zn2+ (1 equiv.) + BODPAQ has a major peak with m/z
-
of 840.2211 ([Zn2+ + BODPAQ + ClO4 ]+), which corresponds to
a 1 : 1 complex (Fig. S6, ESI†).
Fig. 5 Part of the 1H NMR spectra of BODPAQ in CD3CN in the absence
and presence of Zn2+. (a) free BODPAQ; (b) BODPAQ + 1 equiv. Zn2+; (c)
BODPAQ + 4 equiv. Zn2+
.
Scheme 2 Proposed binding model of Zn2+ with BODPAQ.
To validate the nature of the interaction of BODPAQ and Zn2+,
1H NMR titrations in CD3CN were carried out, as shown in Fig.
5. Upon addition of 1 equiv. Zn2+, all the protons of DPA and
AQ experience significant changes. The coordination triggers a
clear change of protons Ha and Hb from two singlet signals to
six doublet signals, indicating that the nitrogen atom attached
to the methylene group strongly bonds with Zn2+. Protons of
pyridines show downfield or upfield shifts as a result of the N-metal
coordination effect, such as Hc at the ortho position, one from 8.54
to 9.15, and another from 8.54 to 8.45. Meanwhile, similar shifts
are also observed for the protons of quinoline, suggesting the direct
interaction between the quinoline platform and metal ions. More
Zn2+ does not lead to any evident change in the 1H NMR spectrum.
All these results indicate a 1 : 1 binding model between BODPAQ
and Zn2+. Unfortunately, we cannot give direct evidence by mass
and 1H NMR spectra to explain the phenomenon that a new type
of complex other than a 1 : 1 complex is formed by introducing
more than 1 equiv. Zn2+, as observed by UV-vis and fluorescence
spectra.
Fig. 6 Structure of Zn2+/BODPAQ complex estimated by density
functional theory calculations. Hydrogen atoms, tetrahedral perchlorate
and solvent molecules are omitted for clarity.
coordinated to Zn2+, thus the dipole moment is distinctly changed
in the Zn2+ binding process, therefore the PET and ICT process
from coordination atoms to the fluorophore is blocked.
The selectivity of fluorescence response of BODPAQ to metal
ions was then investigated by titration of different ions in CH3CN.
As illustrated in Fig. 7, only Zn2+ and Cd2+ enhance the emission of
BODPAQ accompanied by a notable hypsochromic shift. Alkali
and alkaline earth metal ions such as Ca2+, Mg2+, Na+, and
K+ do not induce any emission change. Most heavy transition
metal ions such as Fe2+, Hg2+, and Mn2+ have little influence
The coordination of BODPAQ to Zn2+ was also explored by
molecular modeling. The conformation of the Zn2+/BODPAQ
complex was optimized by density functional theory (DFT)
calculations at the B3LYP/6-311G level (Gaussian 2009).16 As
shown in Fig. 6, the zinc ion in Zn2+/BODPAQ is coordinated
by the three nitrogen atoms of one DPA arm, two N atoms
-
of aminoquinoline, and a solvent molecule or a ClO4 anion
to form the distorted six-coordination geometry. The theoretical
study shows that the amines both in DPA and AQ are directly
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The influence of solvent systems on Zn2+ binding affinity and
selectivity
The binding properties of BODPAQ with Zn2+ obtained from
the studies in pure acetonitrile do not necessarily correspond to
those that apply in aqueous solutions. Therefore, the influence of
the solvent systems on the binding property and selectivity was
studied in different buffer solutions.
The binding property and selectivity were firstly investigated
in 3-morpholinopropanesulfonic acid (MOPS) buffer aqueous
solution (50 mM, 50% CH3CN, pH = 7.3). Compared to the
spectral response of BODPAQ to Zn2+ in 100% acetonitrile, the
measured absorption spectra of free BODPAQ exhibits a strong
absorption band centered at 557 nm in MOPS buffer aqueous
solution with the lmax undergoing a slight blue-shift (Fig. 8a).
When Zn2+ was added, a distinct decrease of the absorption band
at 557 nm can be observed accompanied by a hypsochromic shift
to 511 nm with a distinct isobestic point at 520 nm, which is
consistent with the presence of only two species, free ligand and
Zn2+-ligand complex. The titration profile shows the absorption
at 557 nm descends almost linearly until a ligand-to-metal ratio
of 1 : 1 is reached. Further addition of Zn2+ does not affect the
absorption spectrum, suggesting a 1 : 1 stoichiometry for the zinc
Fig. 7 (a) Fluorescence spectra of BODPAQ (5 mM) in the presence of
various metal ions (4 equiv. of Zn2+, Cd2+, Co2+, Cu2+, Fe2+, Hg2+, Mn2+
,
and Ni2+ and 100 equiv. of K+, Ca2+, and Mg2+) in CH3CN. (b) Histogram
of F/F0 induced by various metal ions. (c) The ratio fluorescence response
(F535/F0) of BODPAQ (5 mM) containing 4 equiv. Zn2+ to the selected
metal ions.
on the fluorescence intensity of BODPAQ while Cu2+, Co2+
and Ni2+ decrease the emission intensity. Therefore, BODPAQ
shows distinct selectivity for Zn2+ over other cations except for
Cd2+.
To further check the Zn2+-specific amplified fluorescence change
of BODPAQ, titration experiments of BODPAQ-Zn2+ complex
(1 : 4) were conducted by the addition of 100 equiv. of Ca2+,
Mg2+, Na+, K+ and 4 equiv. of Fe2+, Hg2+, and Mn2+ Cu2+,
Co2+, Ni2+. The experimental results indicate that the emission
profile of the BODPAQ-Zn2+ complex is not influenced by
these metal ions, indicating the high affinity and selectivity for
Zn2+.
Fig. 8 (a) Absorption spectra of BODPAQ (5 mM) upon addition of Zn2+
(0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.2, 1.4, 1.6, 1.8, 2.0, 3.0 equiv.
with respect to BODPAQ in CH3CN/buffer (50 : 50); Inset is the titration
profile according to the absorbance at 557 nm. (b) Emission spectra of
BODPAQ (5 mm) upon addition of Zn2+ (0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7,
0.8, 0.9, 1.0, 1.2, 1.4, 1.6, 1.8, 2.0, 3.0 equiv. with respect to BODPAQ in
CH3CN/buffer (50 : 50), Inset is the titration profile according to F/F0,
the fluorescence intensity ratio at 544 nm.
836 | Dalton Trans., 2012, 41, 831–838
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complex. The binding constant was determined to be 0.94 ¥ 105
using Benesi–Hildebrand plots (see ESI†).
fluorescence intensity ratio (F/F0) of the BODPAQ-Zn2+ complex,
although Cu2+, Ni2+ and Co2+ slightly quench the fluorescence
and Cd2+ induces a slight fluorescence enhancement. These results
suggest that BODPAQ has good selectivity for Zn2+ under the
experimental conditions.
Fluorescence titration of BODPAQ by Zn2+ in CH3CN/MOPS
buffer (50 : 50) aqueous solution exhibits a distinct emission
enhancement (Ufree = 0.005, UZn = 0.20) with the concentration
of Zn2+ up to a molar ratio of 1 : 1, which is confirmed by Job’s
plot (Fig. 8b). An enhancement factor of 9 was observed. Higher
concentrations of Zn2+ do not lead to noticeable fluorescence
changes, suggesting that only a 1 : 1 Zn2+-BODPAQ complex
formed. The same as in acetonitrile, the Zn2+-induced emission
enhancement is caused by the PET blocking effect and inhibited
ICT induced by the Zn2+ coordination to the AQ amino and
tertiary amino groups in DPA. In addition, BODPAQ responds
to metal ions in the same way as in methanol aqueous solutions
(50 mM MOPS, 50% Methanol, pH 7.3) (Fig. S9, ESI†).
The fluorescence titration experiments were also carried out
in other types of buffer solutions (HEPES, Tris-HCl, citrate–
phosphate). As shown in Fig. 10, Zn2+ induced the highest ratio
of F/F0 in MOPS buffer solution. In other words, the highest
fluorescence off/on ratio can be obtained and thus the highest
sensitivity can be achieved in MOPS. When the buffer solutions
were switched to HEPES (50 mM, 100 M KNO3, 50% CH3CN),
Tris-HCl (10 mM Tris-HCl, 50% CH3CN, 0.1 M KNO3), and
citrate–phosphate, the fluorescence ratio decreases and finally
no response of BODPAQ to Zn2+ occurred in citrate–phosphate
buffer. The sensitivity of BODPAQ to Zn2+ in different buffer
solutions may indicate that the different environment around
BODPAQ and different ionic strength could affect the binding
affinity of the probe to metal ions.17
The fluorescence response of BODPAQ to various cations and
its selectivity for Zn2+ were also examined in CH3CN/MOPS
buffer (50 : 50) aqueous solution (Fig. 9). Fluorescence enhance-
ment of BODPAQ was not observed upon addition of 100 equiv.
of alkali and alkaline earth metals including Na+, K+, Mg2+, and
Ca2+. As for heavy transition metal ions, Fe2+, Hg2+, Mn2+, and
Pb2+ have little influence on the fluorescence intensity of BODPAQ
while Cu2+, Co2+ and Ni2+ obviously quench the fluorescence
to some extent. Only when Zn2+ and Cd2+ were added was a
remarkable fluorescence change detected. Note that Zn2+ induced
much stronger enhancement of the fluorescence than Cd2+. The
competition experiments were then investigated by addition of
various metal ions to the solution of BODPAQ-Zn2+ complex
(1 : 4). Other metal ions promote no significant variation in the
Fig. 10 Fluorescence intensity ratio of BODPAQ towards Zn2+ in
different buffer solutions containing 50% CH3CN as cosolvent, (a) F/F0
is the fluorescence intensity ratio at 544 nm in MOPS buffer solution, (b)
F/F0 at 542 nm in HEPES buffer solution, (c) F/F0 at 547 nm in Tris-HCl
buffer solution, (d) F/F0 at 577 nm in citrate–phosphate buffer solution.
Conclusion
A new borondipyrromethene-based fluorescent probe BOD-
PAQ with both PET and ICT sensing behaviors has been
synthesized. In this probe, two chelators, 2,2¢-dipicolylamine
(DPA) and 8-aminoquinoline (AQ), are combined into one
borondipyrromethene molecule, which was designed to optimize
the coordination to metals of the two chelators in a synergic
manner, thus improve the Zn2+ selectivity. The interaction between
the DPA group and Zn2+ inhibits the electron-donating ability
of the amino group, and the interaction between the 8-AQ
group and Zn2+ blocks the PET process. Therefore, the binding
of Zn2+ to BODPAQ induced not only emission enhancement
but also an emission shift due to the binary inhibition effect
of the PET and ICT mechanisms. In acetonitrile, BODPAQ
displays distinct fluorescence selectivity for Zn2+ with a dramatic
enhanced emission accompanied by a notable hypsochromic shift
among competing metal ions, although Cd2+ also induces an
emission enhancement. More interestingly, BODPAQ displays
the highest sensitivity for Zn2+ in MOPS among several buffer
Fig. 9 (a) Fluorescence spectra of BODPAQ in the presence of various
metal ions (4 equiv. of Zn2+, Cd2+, Co2+, Cu2+, Fe2+, Hg2+, Mn2+, and Ni2+
and 100 equiv. of K+, Ca2+, and Mg2+) in CH3CN/MOPS buffer (50 : 50);
(b) Histogram of F/F0 (544 nm) induced by 4 equiv. of various metal ions.
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solutions used in this study. When the buffer solutions were then
switched to HEPES, Tris-HCl, or citrate–phosphate respectively,
the fluorescence sensitivity begins to decrease, and finally no
response of BODPAQ to Zn2+ occurred in citrate–phosphate
buffer. It should be noted that Zn2+ induced much stronger
enhancement of the fluorescence than Cd2+, however, it is still
hard to distinguish successfully between Zn2+ and Cd2+ in aqueous
solutions. To improve the Zn2+ specific amplified fluorescence in
aqueous solutions, chelators with high affinity for Zn2+ should
be introduced into the borondipyrromethene core. Strategies
including introduction of N,N,N¢-tri(pyridine-2-ylmethyl)ethane-
1,2-diamine (TPEA)5h,9d,18 instead of DPA are in progress.
Forbes, R. F. Seamark, R. Borlinghaus, W. H. Betts, S. F. Lincoln and
A. D. Ward, Chem. Biol., 1993, 1, 153–161.
7 (a) S. C. Burdette, G. K. Walkup, B. Spingler, R. Y. Tsien and S. J.
Lippard, J. Am. Chem. Soc., 2001, 123, 7831–7841; (b) S. C. Burdette,
C. J. Frederickson, W. Bu and S. J. Lippard, J. Am. Chem. Soc., 2003,
125, 1778–1787; (c) E. M. Nolan, S. C. Burdette, J. H. Harvey, S. A.
Hilderbrand and S. J. Lippard, Inorg. Chem., 2004, 43, 2624–2635;
(d) C. J. Chang, E. M. Nolan, J. Jaworski, S. C. Burdette, M. Sheng
and S. J. Lippard, Chem. Biol., 2004, 11, 203–210; (e) C. J. Chang, E.
M. Nolan, J. Jaworski, K.-I. Okamoto, Y. Hayashi, M. Sheng and S. J.
Lippard, Inorg. Chem., 2004, 43, 6774–6779; (f) C. C. Woodroofe, R.
Masalha, K. R. Barnes, C. J. Frederickson and S. J. Lippard, Chem.
Biol., 2004, 11, 1659–1666; (g) E. M. Nolan, J. Jaworski, K.-I. Okamoto,
Y. Hayashi, M. Sheng and S. J. Lippard, J. Am. Chem. Soc., 2005, 127,
16812–16823; (h) G. K. Walkup, S. C. Burdette, S. J. Lippard and R.
Y. Tsien, J. Am. Chem. Soc., 2000, 122, 5644–5645; (i) X. Zhang, K. S.
Lovejoy, A. Jasanoff and S. J. Lippard, Proc. Natl. Acad. Sci. U. S. A.,
2007, 104, 10780–10785; (j) B. A. Wong, S. Friedle and S. J. Lippard,
J. Am. Chem. Soc., 2009, 131, 7142–7152; (k) X. Zhang, D. Hayes, S.
J. Smith, S. Friedle and S. J. Lippard, J. Am. Chem. Soc., 2008, 130,
15788–15789; (l) E. Tomat, E. M. Nolan, J. Jaworski and S. J. Lippard,
J. Am. Chem. Soc., 2008, 130, 15776–15777.
8 (a) Z.-X. Han, X.-B. Zhang, Z. Li, Y.-J. Gong, X.-Y. Wu, Z. Jin, C.-M.
He, L.-X. Jian, J. Zhang, G.-L. Shen and R.-Q. Yu, Anal. Chem., 2010,
82, 3108–3113; (b) K. Hanaoka, K. Kikuchi, H. Kojima, Y. Urano
and T. Nagano, Angew. Chem., Int. Ed., 2003, 42, 2996–2999; (c) K.
Hanaoka, K. Kikuchi, H. Kojima, Y. Urano and T. Nagano, J. Am.
Chem. Soc., 2004, 126, 12470–12476; (d) T. Hirano, K. Kikuchi, Y.
Urano, T. Higuchi and T. Nagano, J. Am. Chem. Soc., 2000, 122, 12399–
12400; (e) T. Hirano, K. Kikuchi, Y. Urano and T. Nagano, J. Am.
Chem. Soc., 2002, 124, 6555–6562.
Acknowledgements
We gratefully acknowledge the financial support by the National
Science Foundation of China (grant no.: 20902021), the Scientific
Research Foundation for the Returned Overseas Chinese Scholars
(State Education Ministry), Key Laboratory of Photochemical
Conversion and Optoelectronic Materials (TIPC, CAS) and
the East China University of Science and Technology research
funds. We also gratefully acknowledge Prof. Fu Wei for help in
optimizing the Zn2+/BODPAQ complex by density functional
theory calculations.
9 (a) D. Srikun, E. W. Miller, D. W. Domaille and C. J. Chang, J. Am.
Chem. Soc., 2008, 130, 4596–4597; (b) K. Komatsu, Y. Urano, H.
Kojima and T. Nagano, J. Am. Chem. Soc., 2007, 129, 13447–13454;
(c) W. Lin, L. Long, B. Chen and W. Tan, Chem.–Eur. J., 2009, 15,
2305–2309; (d) F. Qian, C. Zhang, Y. Zhang, W. He, X. Gao, P. Hu and
Z. Guo, J. Am. Chem. Soc., 2009, 131, 1460–1468.
Notes and references
1 (a) A. P. de Silva, H. Q. N. Gunaratne, T. Gunnlaugsson, A. J. M.
Huxley, C. P. McCoy, J. T. Rademacher and T. E. Rice, Chem. Rev.,
1997, 97, 1515–1566; (b) B. Valeur, Molecular Fluorescence: Principles
and Applications; Wiley-VCH: Weinheim, NY, 2002.
2 (a) J. M. Berg and Y. Shi, Science, 1996, 271, 1081–1085; (b) A. C.
Burdette and S. J. Lippard, Proc. Natl. Acad. Sci. U. S. A., 2003, 100,
3605–3610; (c) B. L. Vallee and K. H. Falchuk, Psychol. Rep., 1993, 73,
79–118.
3 (a) D. W. Choi and J. Y. Koh, Annu. Rev. Neurosci., 1998, 21, 347–375;
(b) J. H. Weiss, S. L. Sensi and J. K. Koh, Trends Pharmacol. Sci., 2000,
21, 395–401; (c) J. Y. Koh, S. W. Suh, B. J. Gwag, Y. Y. He, C. Y. Hsu
and D. W. Choi, Science, 1996, 272, 1013–1016.
4 Recent reviews for zinc sensors: (a) E. L. Que, D. W. Domaille and
C. J. Chang, Chem. Rev., 2008, 108, 1517–1549; (b) E. M. Nolan and
S. J. Lippard, Acc. Chem. Res., 2009, 42, 193–203; (c) Z. Dai and J.
W. C a n a r y , New J. Chem., 2007, 31, 1708–1718; (d) P. Jiang and J.
Guo, Coord. Chem. Rev., 2004, 248, 205–229; (e) P. Carol, S. Sreejith
and A. Ajayaghosh, Chem.–Asian J., 2007, 2, 338–348; (f) N. C. Lim,
H. C. Freake and C. Bru¨ckner, Chem.–Eur. J., 2005, 11, 38–49; (g) D.
W. Domaille, E. L. Que and C. J. Chang, Nat. Chem. Biol., 2008, 4,
168–175.
5 (a) T. W. Kim, J. Park and J.-I. Hong, J. Chem. Soc., Perkin Trans. 2,
2002, 923–927; (b) K. R. Gee, Z.-L. Zhou, W.-J. Qian and R. Kennedy,
J. Am. Chem. Soc., 2002, 124, 776–778; (c) N. C. Lim, L. Yao, H. C.
Freake and C. Bru¨ckner, Bioorg. Med. Chem. Lett., 2003, 13, 2251–
2254; (d) T. Koike, T. Watanabe, S. Aoki, E. Kimura and M. Shiro,
J. Am. Chem. Soc., 1996, 118, 12696–12703; (e) S. Aoki, S. Kaido,
H. Fujioka and E. Kimura, Inorg. Chem., 2003, 42, 1023–1030; (f) T.
Hirano, K. Kikuchi, Y. Urano, T. Higuchi and T. Nagano, Angew.
Chem., Int. Ed., 2000, 39, 1052–1054; (g) A. Czarnik, Acc. Chem. Res.,
1994, 27, 302–308; (h) K. Hanaoka, Y. Muramatsu, Y. Urano, T. Terai
and T. Nagano, Chem.–Eur. J., 2010, 16, 568–572; (i) Z. Li, M. Yu, L.
Zhang, M. Yu, J. Liu, L. Wei and H. Zhang, Chem. Commun., 2010, 46,
7169–7171; (j) X. Peng, X. Tang, W. Qin, W. Dou, Y. Guo, J. Zheng,
W. Liu and D. Wang, Dalton Trans., 2011, 40, 5271–5277.
10 (a) A. Caballero, R. Martinez, V. Lioveras, I. Ratera, J. Vidal-Gancedo,
K. Wurst, A. Tarraga, P. Molina and J. Veciana, J. Am. Chem. Soc.,
2005, 127, 15666–15667; (b) K. Kiyose, H. Kojima, Y. Urano and T.
Nagano, J. Am. Chem. Soc., 2006, 128, 6548–6549; (c) Z. Xu, K.-H.
Baek, H. N. Kim, J. Cui, X. Qian, D. R. Spring, I. Shin and J. Yoon, J.
Am. Chem. Soc., 2010, 132, 601–610; (d) L. Xue, Q. Liu and H. Jiang,
Org. Lett., 2009, 11, 3454–3457; (e) Z. Liu, C. Zhang, Y. Li, Z. Wu,
F. Qian, X. Yang, W. He, X. Gao and Z. Guo, Org. Lett., 2009, 11,
795–798; (f) S. Maruyama, K. Kikuchi, T. Hirano, Y. Urano and T.
Nagano, J. Am. Chem. Soc., 2002, 124, 10650–10651; (g) M. Taki, J. L.
Wolford and T. V. O’Halloran, J. Am. Chem. Soc., 2004, 126, 712–713;
(h) C. C. Woodroofe and S. J. Lippard, J. Am. Chem. Soc., 2003, 125,
11458–11459; (i) C. J. Chang, J. Jaworkski, E. M. Nolan, M. Sheng and
S. J. Lippard, Proc. Natl. Acad. Sci. U. S. A., 2004, 101, 1129–1134;
(j) L. Xue, C. Liu and H. Jiang, Chem. Commun., 2009, 1061–1063.
11 (a) D. A. Pearce, N. Jotterand, I. S. Carrico and B. Imperiali, J.
Am. Chem. Soc., 2001, 123, 5160–5161; (b) G. Xue, J. S. Bradshaw,
N. K. Dalley, P. B. Savage, R. M. Izatt, L. Prodi, M. Montalti and
N. Zaccheroni, Tetrahedron, 2002, 58, 4809–4815; (c) Y. Mikata, M.
Wakamatsu and S. Yano, Dalton Trans., 2005, 545–550.
12 (a) S. Petruso and S. Caronna, J. Heterocycl. Chem., 1992, 29, 355–357;
(b) S. Mula, A. K. Ray, M. Banerjee, T. Chaudhuri, K. Dasgupta and
S. Chattopadhyay, J. Org. Chem., 2008, 73, 2146–2154.
13 L. Jiao, C. Yu, J. Li, Z. Wang, M. Wu and E. Hao, J. Org. Chem., 2009,
74, 7525–7528.
14 L. Xue, C. liu and H. Jiang, Org. Lett., 2009, 11, 1655–1658.
15 (a) H. Sunahara, Y. Urano, H. Kojima and T. Nagano, J. Am. Chem.
Soc., 2007, 129, 5597–5604; (b) K. Rurack, M. Kollmannsberger, U.
Resch-Genger and J. Daub, J. Am. Chem. Soc., 2000, 122, 968–969;
(c) M. Kollmannsberger, K. Rurack, U. Resch-Genger and J. Daub, J.
Phys. Chem. A, 1998, 102, 10211–10220.
16 M. Frisch, G. Trucks, J. Cheeseman, G. Scalmani, F. Clemente, H.
Hratchian, M. Caricato, A. Izmaylov, J. Hess, C. Adria, et al. Gaussian
09, Revision A1[M], Wallingford CT: Gaussian Inc., 2009.
6 (a) C. J. Frederickson, E. J. Kasarskis, D. Ringo and R. E. Frederickson,
J. Neurosci. Methods, 1987, 20, 91–103; (b) M. C. Kimber, I. B.
Mahadevan, S. F. Lincoln, A. D. Ward and E. R. T. Tiekink, J. Org.
Chem., 2000, 65, 8204–8209; (c) C. J. Fahrni and T. V. O’Halloran,
J. Am. Chem. Soc., 1999, 121, 11448–11458; (d) P. D. Zalewski, I. J.
17 T. Cheng, T. Wang, W. Zhu, Y. Yang, B. Zeng, Y. Xu and X. Qian,
Chem. Commun., 2011, 47, 3915–3917.
18 E. Kawabata, K. Kikuchi, Y. Urano, H. Kojima, A. Odani and T.
Nagano, J. Am. Chem. Soc., 2005, 127, 818–819.
838 | Dalton Trans., 2012, 41, 831–838
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