Target-triggered deprotonation of 6-hydroxyindole-based BODIPY: Specially switch on NIR fluorescence upon selectively binding to Zn2+

Article abstract of DOI:10.1039/c2cc35080g

Based on 6-hydroxyindole BODIPY with a Schiff-base structure, NIR fluorescence with impressively high selectivity is triggered by deprotonation of the phenol group upon binding with Zn²⁺ due to the chelation-enhanced fluorescence effect, thus realizing a promising application in bioimaging of Zn²⁺. The Royal Society of Chemistry 2012.

Full text of DOI:10.1039/c2cc35080g

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Target-triggered deprotonation of 6-hydroxyindole-based BODIPY:
specially switch on NIR fluorescence upon selectively binding to Zn2+w
Jian Cao,ab Chunchang Zhao,*a Xuzhe Wang,a Yanfen Zhanga and Weihong Zhu*a
Received 16th July 2012, Accepted 21st August 2012
DOI: 10.1039/c2cc35080g
Based on 6-hydroxyindole BODIPY with a Schiff-base structure,
NIR fluorescence with impressively high selectivity is triggered by
deprotonation of the phenol group upon binding with Zn2+ due to the
chelation-enhanced fluorescence effect, thus realizing a promising
for Zn2+ (Scheme 1). Upon titration with Zn2+, probe 1-OH
undergoes a significant increase in NIR fluorescence. The
enhanced fluorescence might be attributed to the Zn2+ binding
with the Schiff-base ligand, whereupon it would deprotonate
the phenol group and rigidify the CQN bond. Interestingly, the
resulting complex of Zn(1-O)2 (Scheme 1) exhibits a long
emission wavelength at 680 nm, thus making the determining
wavelength fall in the desirable NIR region. Furthermore,
the selectivity of 1-OH towards Zn2+ is impressively high,
especially with little interference from Cd2+. As demonstrated,
1-OH can be successfully applied for sensing Zn2+ in aqueous
media and living cells, showing high promise in clarifying the
roles of Zn2+ in biological processes.
application in bioimaging of Zn2+
.
Zinc is a vital component in biological systems since it plays vital
roles in catalytic functions of proteins, gene expression, apoptosis,
neurotransmission and so forth.1 As an active research field, rapid
progress has been made in fluorescent probes for Zn2+ 2–4
Near
.
Infra-red (NIR) fluorescent sensors are highly desirable due to
the induction of minimum background fluorescence and less
photodamage as well as deep penetration into tissue.5,6 The
commonly employed approach to construct NIR Zn2+ probes
is to install responsive receptor units into chromophores with
extended p-conjugation moieties so as to shift the absorption
or emission to long wavelengths. Alternatively, sensors based
on the chelation-enhanced fluorescence (CHEF) effect are
relatively convenient with incorporation of proper ligands into
the chromophore. Phenol-based Schiff bases (imines) are known
as good p-conjugated ligands for metal ions.7 In general, dyes
containing acyclic CQN bonds are poorly fluorescent due to the
isomerization of CQN bonds in excited states, but dyes with
cyclic CQN bonds are highly fluorescent.8 Both the imine
nitrogen and phenol oxygen can serve as good ligand donors
for metal ions, giving rise to structural rigidity via coordination
with metal ions. Thus, the isomerization of the CQN bond can be
inhibited, resulting in the specific CHEF effect. More importantly,
the phenol group is deprotonated in the coordination process,
which can be seen in a characteristic emission of phenolate.9
Accordingly, construction of phenol-based Schiff bases on the
basis of the CHEF effect would be expected to provide
opportunities to the further development of NIR probes.
Herein, we report a novel BODIPY derivative (1-OH) with
salicylaldehyde benzoyl hydrazone as a specific tridentate
chelating unit to develop a turn-on NIR fluorescence probe
As shown in Scheme 1, 1-OH and the reference compound
1-OMe were synthesized straightforwardly. Formylation
in the 5-position via a standard Vilsmeier–Haack reaction
generated aldehyde 2 with 68% isolated yield. BODIPY based
salicylaldehyde 3 was then obtained by removal of the methoxy
protecting group. Both 2 and 3 were treated with benzohydrazide
in methanol to afford 1-OMe and 1-OH, respectively. Their
chemical structures were fully characterized by 1H NMR,
13C NMR and HRMS (ESIw).
To demonstrate the potential of 1-OH as a turn-on NIR
fluorescent probe, we firstly evaluated the binding property of
1-OH toward Zn2+ in CH3CN. Upon titration of Zn2+ with
1-OH, distinct spectral changes were observed (Fig. 1). The
intense absorption band at 548 nm was found to decrease
while a new band at 617 nm increased with a well-defined
isosbestic point at 567 nm, consistent with the presence of two
species, free 1-OH and the resulting complex between Zn2+
a Shanghai Key Laboratory of Functional Materials Chemistry, 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, whzhu@ecust.edu.cn
b Shanghai University of Engineering Science, Shanghai 201620,
P. R. China
Scheme 1 Synthesis of 1-OH and reference compound 1-OMe:
(i) DMF, POCl3, CH2Cl2, r. t., 68% yield; (ii) BBr3, CH2Cl2, À78 1C,
40% yield; (iii) benzohydrazide, EtOH, reflux.
w Electronic supplementary information (ESI) available: The synthesis
and spectroscopic properties of 1-OH. See DOI: 10.1039/c2cc35080g
c
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Based on the fluorescence titration experiments, the detection
limit was evaluated to be 9.7 Â 10À7 M.
The coordination properties of 1-OH with Zn2+ were also
examined by means of 1H NMR spectra (Fig. S2, ESIw). Upon
addition of Zn2+, the phenolic proton signal at 11.13 ppm
disappeared, and the aromatic protons on the phenol ring
underwent a significant downfield shift, suggesting that the
coordination occurred between the deprotonated phenolic
group and Zn2+. Moreover, the downfield shift from 11.88
to 12.69 ppm, corresponding to the amide signal, is indicative
of the coordination of Schiff-base ligand to Zn2+. The possible
1 : 2 binding model of Zn2+ and 1-OH (Scheme 1) was further
confirmed by the ESI mass spectrum of Zn2+ (0.5 equiv.) +
1-OH (Fig. S3, ESIw). A peak found at 1163.5160 was consistent
with the calculated data for [Zn(1-O)2 + H]+ (1163.3916).
The Zn2+-binding ability of 1-OH was also investigated in
aqueous solution (0.05 M Tris-HCl, 50% CH3CN, pH = 7.5).
1-OH showed almost no fluorescence with a fluorescent
quantum yield of 0.003 upon excitation at 617 nm, suggesting
that 1-OH is a promising probe with very low background
fluorescence. The large fluorescence enhancement was observed
upon addition of Zn2+ to the solution of 1-OH. More inter-
estingly, 1-OH can only sense Zn2+ in aqueous solution with
high selectivity (Fig. 3). For instance, other cations such as
Fig. 1 Absorption spectra of 1-OH (5.0 mM) in the presence of
different concentrations 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, 2.2, 2.4, 2.6, 2.8 and 3.0 equiv.) in
CH3CN. Inset: the color change of 1-OH in the absence and presence
of Zn2+
.
and 1-OH. Also, the solution color changed from pink to blue
upon addition of Zn2+
.
Next, we investigated the Zn2+-binding ability of 1-OH by
fluorescence titration (Fig. 2). When excited at the isosbestic
wavelength (567 nm) or the newly formed band (617 nm), the
fluorescence signal at 680 nm increased significantly upon
addition of Zn2+. The fluorescence quantum yield (Ff)
increased from 0.006 for free 1-OH to 0.19 in the presence
of excess Zn2+ upon excitation at 617 nm, showing a 32-fold
enhancement in Ff. This is indicative of a sufficient fluores-
cence response of 1-OH to Zn2+ to serve as a NIR turn-on
probe for Zn2+. Moreover, the Job plot indicated that the
coordination between Zn2+ and 1-OH has a 1 : 2 binding
stoichiometry (Fig. 2b), exhibiting a binding constant of 3.5 Â
Ca2+, Mg2+, Na+, K+, Li+, Fe2+, Hg2+, Mn2+, Cu2+
,
Co2+ and Ni2+ gave no observable change. Even Cd2+, which
interferes with Zn2+ detection in most reported Zn2+ sensors,
has little effect on the fluorescence of 1-OH.
We finally sought to apply 1-OH for the bioimaging of
Zn2+ in living Breast Cancer MCF-7 cells (Fig. 4). Upon
incubation with 5.0 mM of 1-OH for 30 min at 37 1C, the cells
displayed a faintly intracellular fluorescence image. When
further treated with Zn2+ in the culture medium for 30 min,
and washed with phosphate buffered saline to remove extracellular
Zn2+, the bright red fluorescence image was then observed in these
106
M
À2, from the theoretical nonlinear fit of experimental
data to a 1 : 2 binding model. In contrast, control experiments
with reference compound 1-OMe under the same conditions
showed no change in the spectra of absorption and fluores-
cence upon addition of Zn2+ (Fig. S1, ESIw), hinting that the
ortho-hydroxy group in 1-OH plays a vital role in the coordi-
nation of Zn2+. Obviously, such a large enhancement in NIR
fluorescence could be attributed to the inhibition of CQN
isomerization and the deprotonation of the phenolic group
conjugated to the BODIPY core upon stable chelation of
1-OH with Zn2+ (Scheme 1), which is a typical CHEF effect.
Fig. 3 Fluorescence response of 5.0 mM 1-OH to various metal ions
in aqueous solution (CH3CN/Tris-HCl = 1 : 1 v/v, pH = 7.5) upon
excitation at 617 nm: (a) free 1-OH, (b) Zn2+, (c) Cu2+, (d) Pb2+
(e) Ni2+, (f) Hg2+, (g) Cd2+, (h) Mg2+, (i) Li+, (j) Na+, (k) Co2+
(l) Fe2+, (m) K+, (n) Ba2+, (o) Mn2+, (p) Ca2+, (q) Ag+, (r) Cr2+
,
,
.
Fig. 2 (a) Fluorescence spectra of 1-OH (5.0 mM) in CH3CN upon
different concentrations 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, 2.2, 2.4, 2.6, 2.8 and 3.0 equiv.) upon
excitation at 617 nm. (b) Job plot of 1-OH obtained from fluorescence
titration in CH3CN. The total concentration of 1-OH and Zn(ClO4)2
is 10.0 mM.
Note: pink bars for selectivity experiments with 1-OH and 4.0 equiv.
of metal ions; blue bars for interfering experiments which were
conducted in the presence of 100 equiv. of Ca2+, Mg2+, Na+, K+
,
and 4 equiv. of Fe2+, Hg2+, Mn2+, Cu2+, Co2+, Ni2+, Cd2+, with
the subsequent addition of 1 equiv. of Zn2+
.
c
9898 Chem. Commun., 2012, 48, 9897–9899
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Fig. 4 Fluorescence confocal images of Breast Cancer MCF-7 cells:
top, (a–c) cells incubated with 5.0 mM 1-OH for 30 min; bottom, (d–f)
cells incubated with 5.0 mM 1-OH for 30 min, then treated with Zn2+
for another 30 min and washed with PBS. Overlap field (a and d),
fluorescence image (b and e), bright field (c and f). The samples were
excited with 638 nm, under observation between 662–737 nm.
cells. The overlap of fluorescence and bright-field images
revealed that the fluorescence signals were localized in the
perinuclear area of the cytosol, indicative of a subcellular
distribution and good cell membrane permeability of 1-OH.
These results suggest that 1-OH can be explored for monitoring
Zn2+ within living cells.
In summary, we reported the synthesis and photophysical
evaluation of a NIR turn-on fluorescent probe 1-OH for Zn2+
with high selectivity. The coordination of 1-OH with Zn2+
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Schiff-base ligand on the basis of the CHEF effect, resulting in
the deprotonation of the phenol group and causing the CQN
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NIR region, 1-OH can be explored for the bioimaging of Zn2+
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This work was financially supported by NSFC/China,
National 973 Program, the Oriental Scholarship, the Funda-
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Chem. Commun., 2012, 48, 9897–9899 9899