A novel quinoline-based two-photon fluorescent probe for detecting Cd 2+in vitro and in vivo

Article abstract of DOI:10.1039/c2dt30192j

A new two-photon fluorescent Cd²⁺ probe APQ is developed by introducing a N¹,N¹-dimethyl-N²-(pyridin-2- ylmethyl)ethane-1,2-diamine binding group and a 4-methoxyphenylvinyl conjugation-enhancing group to the 2- and 6-positions of quinoline. This probe shows a large red shift and good emission enhancement under Cd²⁺ binding. It also exhibits a high ion selectivity for Cd²⁺ (especially over Zn²⁺) and a large two-photon absorption cross section at 710 nm. Two-photon microscopy imaging studies reveal that the new probe is non-toxic and cell-permeable and can be used to detect intracellular Cd²⁺ under two-photon excitation.

Full text of DOI:10.1039/c2dt30192j

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PAPER
A novel quinoline-based two-photon fluorescent probe for detecting Cd2+
in vitro and in vivo†
Yiming Li,‡a,b Hanbao Chong,‡a Xiangming Meng,*a Shuxin Wang,a Manzhou Zhua and Qingxiang Guob
Received 26th January 2012, Accepted 5th April 2012
DOI: 10.1039/c2dt30192j
A new two-photon fluorescent Cd2+ probe APQ is developed by introducing a N1,N1-dimethyl-N2-
(pyridin-2-ylmethyl)ethane-1,2-diamine binding group and a 4-methoxyphenylvinyl conjugation-
enhancing group to the 2- and 6-positions of quinoline. This probe shows a large red shift and good
emission enhancement under Cd2+ binding. It also exhibits a high ion selectivity for Cd2+ (especially over
Zn2+) and a large two-photon absorption cross section at 710 nm. Two-photon microscopy imaging
studies reveal that the new probe is non-toxic and cell-permeable and can be used to detect intracellular
Cd2+ under two-photon excitation.
Introduction
As one of the most toxic metal pollutants, cadmium has been
widely used in industry, agriculture, and many other fields.1–3
This element is not essential for life and listed by the U.S.
Environmental Protection Agency as one of 126 priority pollu-
tants. There have been many reports on the toxicity of Cd2+ to
Scheme 1 The design of APQ for Cd2+ detection.
bones, kidneys, the nerve system, and tissues, which result in
many serious health problems.4–6 In particular, Cd2+ can replace
Zn2+ in many zinc enzymes, thereby impairing their catalytic
activities.7 Thus, the detection and quantification of Cd2+ in vitro
developed for the detection of various analytes, very few studies
have been reported about the development of two-photon probes
and in vivo is a highly needed technology for a number of
research areas. Although several approaches including atomic
for Cd2+. Herein, we design and synthesize a 6-substitued
quinoline-based Cd2+ two-photon probe APQ (Scheme 1). This
absorption and inductively coupled plasma have been previously
developed for the detection of Cd2+,8–10 these methods are rela-
design is inspired by our recent success in developing a two-
photon Zn2+ probe derived from 7-hydroxyquinoline.30 The
reason for using the quinoline skeleton is that quinoline exhibits
tively expensive and unsuitable for real-time monitoring. In com-
parison, fluorescent probes often provide a better choice to
good two-photon spectrum properties, good water solubility, and
detect metal ions in biology and the environment because of
low cytotoxicity.31–33 To enhance the ion selectivity and sensi-
their high selectivity and sensitivity.11–13 Earlier fluorescent
Cd2+ probes are mostly designed based on single-photon fluor-
tivity, the N,N-dimethyl-N-(pyridin-2-ylmethyl)methanediamine
group is incorporated as a Cd2+ receptor. Moreover, we put a
escence technology.3,6,14–18 Most of these earlier fluorescent
Cd2+ probes also suffer from relatively low selectivity between
4-methoxyphenylvinyl conjugation-enhancing group at the
Cd2+ and Zn2+.19–22
6-position to promote intramolecular charge transfer (ICT) to
improve the fluorescence properties. This ICT involves a charge
transfer process from an electron donor to an electron acceptor
Recently, two-photon microscopy (TPM) has evolved into a
widely used tool in biomedical research because this technology
within a fluorophore, thereby causing a red-shift emission upon
shows less phototoxity, better spatial localization, increased
excitation.34–36
specimen penetration, and lower cellular autofluorescence.23–29
Although many two-photon fluorescent probes have been
Experimental section
aDepartment of Chemistry, Anhui University, Hefei 230039, P. R. China.
All reagents and solvents were commercially purchased. 1H
Fax: +86-551-5107342; Tel: +86-551-5108970
bDepartment of Chemistry, University of Science and Technology of
China, Hefei 230026, P. R. China. E-mail: mengxm@ahu.edu.cn
†Electronic supplementary information (ESI) available: Details of
photophysical measurements and cell imaging; NMR, MALDI-TOF MS
spectra data. See DOI: 10.1039/c2dt30192j
NMR spectra were recorded on Bruker-400 MHz spectrometers
and 13C NMR spectra were recorded on 100 MHz spectrometers.
Fluorescence spectra were obtained using a HITACHI F-2500
spectrometer. The detection of metal ions was operated at pH
7.4, maintained with HEPES buffer (25 mM HEPES/0.1 M
‡These authors contributed equally to this work.
This journal is © The Royal Society of Chemistry 2012
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NaNO3). UV-vis absorption spectra were recorded on a Tech-
comp UV 1000 spectrophotometer. Fluorescence responses were
recorded on a FL2500. MS spectra were conducted by Bruker
MALDI TOF. The two-photon cross section was tested in
methanol with 1 mM APQ.
8.18–8.25 (2H, dd, J = 18.1 Hz), 10.21 (1H, s). 13C NMR
(100 MHz, CDCl3, ppm): δ 55.37, 114.33, 117.88, 125.11,
125.15, 128.19, 128.30, 129.41, 130.54, 130.64, 131.37, 136.97,
138.71, 147.44, 151.91, 159.97, 193.51.
(E)-N1-((6-(4-Methoxystyryl)quinolin-2-yl)methyl)-N2,N2
dimethyl-N1-(pyridin-2-ylmethyl)ethane-1,2-diamine (APQ)
6-Bromo-2-methylquinoline (1)
1.5 g of compound 3 (5.2 mmol) and 1.0 g of compound 4
(5.2 mmol) were mixed in 50 mL of dichloromethane and stirred
for 1 h. Then, 1.33 g of NaBH(OAc)3 (6.24 mmol) was added
gradually in 1 h. After 4 h, the solution was washed by 5 mL of
water twice. The organic layer was dried by sodium sulfate and
evaporated to dryness. The crude product was purified by
column chromatography by using a mixed eluent DCM–MeOH
(10 : 1). 1H-NMR (400 MHz, DMSO, ppm): δ 8.48–8.47 (1H, d,
J = 4.1 Hz), 8.28–8.26(1H, d, J = 8.5 Hz), 8.04–8.02 (1H, dd,
J = 1.6, 8.9 Hz), 7.97 (1H, s), 7.93–7.91(1H, d, J = 8.8 Hz),
7.79–7.74 (1H, td, J = 1.7, 7.7, 7.7 Hz), 7.70–7.68 (1H, d, J =
8.49 Hz), 7.60–7.58 (2H, d, J = 8.7 Hz), 7.56–7.54 (1H, d, J =
7.8 Hz), 7.39–7.34 (1H, d, J = 16.4 Hz), 7.28–7.25 (1H, d, J =
12.2 Hz), 7.24–7.22 (1H, d, J = 7.5 Hz), 6.98–6.96 (2H, d, J =
8.7 Hz), 3.93 (2H, s), 3.81 (2H, s), 3.78 (3H, s), 2.63–2.60
(2H, t, J = 6.9 Hz), 2.44–2.40 (2H, t, J = 6.8 Hz), 2.06 (6H, s).
13C NMR (100 MHz, DMSO, ppm) δ 160.04, 159.34, 159.14,
148.65, 146.46, 136.44, 136.05, 135.18, 129.51, 129.27,128.77,
127.93, 127.25, 127.10, 125.42, 125.21, 122.68, 122.05, 121.33,
114.21, 60.73, 60.10, 56.94, 55.12, 51.49, 45.27.
A mixture of 4-bromobenzenamine (14.59 g, 84.8 mmol) and
HCl (6 N, 60 mL) was heated to 100 °C. Then, crotonaldehyde
(11.9 g, 170 mmol) was added slowly. The resulting mixture was
refluxed until TLC showed no raw material. After being cooled
to room temperature, 200 mL of H2O was added. The mixture
was extracted with EtOAc (100 mL × 2) to remove unreacted
crotonaldehyde. The aqueous phase was neutralized with
aqueous ammonia and then extracted with EtOAc (50 mL × 2).
The organic phase was dried by Na2SO4 and evaporated to give
the crude product. The target product was recrystallized in
EtOAc–PE to give 13.46 g of 1 (60.64 mmol, yield = 75.6%).
1H NMR (400 MHz, CDCl3, ppm): δ 2.73 (3H, s), 7.29–7.31
(1H, d, J = 8.4 Hz), 7.73–7.75 (1H, d, J = 9.0 Hz), 7.87–7.96
(3H, m). 13C NMR (400 MHz, CDCl3, ppm): δ 25.18, 119.57,
122.91, 127.64, 129.54, 130.12, 133.03, 135.49, 146.05, 159.45.
(E)-6-(4-Methoxystyryl)-2-methylquinoline (2)
A mixture of 1 (6.03 g, 27.15 mmol), 1-methoxy-4-vinyl-
benzene (4.37 g, 32.58 mmol), PdCl2(PPh3)2 (77 mg,
0.97 mmol), K2CO3 (11 g, 79.7 mmol) and DMF (25 mL) was
heated at 160 °C for 24 h. After being cooled to room tempera-
ture, the mixture was filtered to remove salts, and then 100 mL
of H2O was added. The resulting mixture was extracted by
dichloromethane (50 mL × 3). The organic phase was combined,
dried over Na2SO4, and evaporated to give the crude product.
The target product was recrystallized in EtOAc–PE to give
Measurement of two-photon cross section (δ)
Two-photon excitation fluorescence (TPEF) spectra were
measured using femtosecond laser pulse and Ti: sapphire system
(680–1080 nm, 80 MHz, 140 fs, Chameleon II) as the light
source. All measurements were carried out in air at room temp-
erature. Two-photon absorption cross sections were measured
using the two-photon-induced fluorescence measurement tech-
nique. The two-photon absorption cross sections (δ) were deter-
mined by comparing their TPEF to that of fluorescein in
different solvents, according to the following equation:
1
6.77 g of 2 (24.63 mmol, yield = 90.7%). H-NMR (400 MHz,
CDCl3, ppm): δ 2.75 (3H, s), 3.84 (3H, s), 6.91–6.93 (2H, d, J =
8.6 Hz), 7.08–7.21 (2H, q, J = 16.3 Hz), 7.25–7.27 (1H, d, J =
8.3 Hz), 7.48–7.50 (2H, d, J = 8.6 Hz), 7.73 (1H, s), 7.91–7.93
(1H, d, J = 8.9 Hz), 8.01–8.03 (2H, d, J = 8.5 Hz). 13C-NMR
(100 MHz, CDCl3, ppm): δ 25.11, 55.35, 114.23, 122.39,
125.19, 125.79, 126.81, 127.39, 127.87, 128.59, 129.36, 129.90,
135.26, 136.35, 147.07, 158.43, 159.54.
Φ
ref cref nref
F
δ ¼ δref
Φ
c
n Fref
In the equation, the subscript ref stands for the reference mol-
ecule. δ is the two-photon adsorption cross-section value, c is
the concentration of solution, n is the refractive index of the sol-
ution, F is the TPEF integral intensities of the solution emitted at
the exciting wavelength, and Φ is the fluorescence quantum
yield. The δref value of reference was taken from the literature.37
(E)-6-(4-Methoxystyryl)quinoline-2-carbaldehyde (3)
A solution of compound 2 (2.00 g, 7.27 mmol) in dioxane
(20 mL) was heated at 60 °C. SeO2 (8.00 mmol, 0.889 g) was
added to this solution. Then the reaction temperature was
increased to 80 °C. After 2.5 h, the mixture was cooled to room
temperature. Precipitates were filtered off and washed with
dioxane (5 mL × 2). The organic phase was combined and con-
centrated to give the crude product. The crude material was
purified by column chromatography (dichloromethane as the
Cytotoxicity assays
To test the cytotoxic effect of the probe in cells for over a 24 h
period, MTT (5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium
bromide) assay was performed as previously reported.38 HeLa
cells were passed and plated to ca. 70% confluence in 96-well
plates 24 h before treatment. Prior to APQ treatment, DMEM
1
eluent) to give 1.818 g of 3 (6.61 mmol, yield = 90.9%). H
NMR (400 MHz, CDCl3, ppm): δ 3.85 (3H, s), 6.92–6.94 (2H,
d, J = 8.6 Hz), 7.11–7.30 (2H, dd, J = 16.1 Hz), 7.50–7.52 (2H,
d, J = 8.6 Hz), 7.82 (1H, s), 7.99–8.05 (2H, dd, J = 16.4 Hz),
6190 | Dalton Trans., 2012, 41, 6189–6194
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(Dulbecco’s Modified Eagle Medium) with 10% FCS (Fetal Calf
Serum) was removed and replaced with fresh DMEM, and ali-
quots of APQ stock solutions (5 mM DMSO) were added to
obtain final concentrations of 10, 30, and 50 μM, respectively.
The treated cells were incubated for 24 h at 37 °C under 5%
CO2. Subsequently, the cells were treated with 5 mg mL−1 MTT
(40 μL per well) and incubated for an additional 4 h (37 °C, 5%
CO2). Then, the cells were dissolved in DMSO (150 μL per
well), and the absorbance at 570 nm was recorded. The cell via-
bility (%) was calculated according to the following equation:
Cell viability% = OD570(sample)/OD570(control) × 100, where
OD570(sample) represents the optical density of the wells treated
with various concentrations of APQ and OD570(control) rep-
resents that of the wells treated with DMEM plus 10% FCS. The
percent of cell survival values is relative to untreated control
cells.
show that upon coordination to Cd2+ in DMSO-d6 the proton at
the ortho-position of pyridine shifts downfield from 3.34 to
3.62 ppm. Job’s plot analysis indicates when the Cd2+ molar
fraction reaches 0.5, the maximum fluorescence intensity is
obtained. This indicates a 1 : 1 bonding model between the
probe and Cd2+. Furthermore, the MALDI-TOF MS spectrum of
the probe increased by 148 Da upon the addition of Cd2+. This
observation confirms the formation of the APQ–Cd2+ complex
with a 1 : 1 binding mode (Fig. 1) (ESI†).
Absorption spectra studies
To investigate the binding property of APQ toward Cd2+, the
UV-vis spectra of APQ are measured at room temperature in the
Tris-HCl (50 mM, pH = 7.4) ethanol–H2O (v/v = 1 : 9) buffer in
the presence of Cd2+. As shown in Fig. 2, the UV-vis spectrum
of APQ exhibits two strong absorption peaks at 290 and
340 nm, respectively. Upon addition of Cd2+ (0 to 1.5 equiv.),
the absorption spectrum of APQ exhibits a bathochromic shift
with a clear isosbestic point at 310 nm. Besides, APQ shows
clear absorption shifts in the visible light range 325–375 nm,
indicating the electron-donating effect of the 6-methoxylphenyl-
vinyl group.
Cell culture and two-photon fluorescence microscopy imaging
For two-photon bio-imaging, HeLa cells were cultured in
DMEM supplemented with 10% FCS, penicillin (100 μg mL−1),
and streptomycin (100 μg mL−1) at 37 °C in a humidified atmos-
phere with 5% CO2 and 95% air. Cytotoxicity assays show that
APQ is safe enough for two-photon bio-imaging at low concen-
trations, so that the cells were incubated with 15 μM APQ at
37 °C under 5% CO2 for 30 min, washed once and bathed in
DMEM containing no FCS prior to imaging and/or Cd2+
addition. Then 20 μM Cd2+ was added in the growth medium for
0.5 h at 37 °C, washed 3 times with PBS buffer. Then, cells
were imaged on a confocal microscope (Zeiss LSM 510 Meta
NLO). Two-photon fluorescence microscopy images of labeled
cells were obtained by exciting the probe with a mode-locked
titanium-sapphire laser source set at wavelength 800 nm.
Fluorescence spectra responses
As shown in Fig. 3, APQ exhibits a weak fluorescence emission
at 475 nm when excited at 360 nm. The addition of Cd2+ causes
Results and discussion
As shown in Scheme 2, APQ can be easily synthesized in four
steps with good yields. With the probe in hand, we then examine
1
its complex with Cd2+. The H NMR spectra of APQ (ESI†)
Fig. 1 Job’s plot for binding with Cd2+ (λex = 360 nm, λem = 520 nm).
Scheme 2 Synthesis of APQ.
Fig. 2 The UV-vis spectra of APQ in the presence of Cd2+
.
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Fig. 4 Fluorescence response of APQ to different metal ions (1–16:
Cd2+, Zn2+, Ca2+, Co2+, Cr3+, Cu+, Cu2+, Fe2+, Fe3+, Hg2+, K+, Mg2+
,
Mn2+, Na+ Ni2+, Pb2+) in Tris buffer (pH = 7.4, λex = 360 nm). Insert:
emission spectra of APQ upon addition of Cd2+ and other metal ions
(λex = 360 nm).
Fig. 3 Fluorescence emission spectra of APQ (25 μM), with the exci-
tation at 360 nm, upon the titration of Cd2+ (0–1.5 equiv) in methanol–
water solution (1 : 9, v/v, 50 mM HEPES buffer, pH = 7.4).
the maximum emission of APQ to shift from 475 to 520 nm
with a significant red shift of 45 nm. Such a large red shift is
rather rare among the fluorescent probes reported for Cd2+ detec-
tion to date.18 This red shift also indicates that the nitrogen atom
of the quinoline platform is coordinated with Cd2+ causing an
enhanced ICT effect. Such a large red shift provides a good
opportunity for fluorescent Cd2+ detection using this new probe.
Note that the maximum of fluorescence emission intensity is
obtained when the Cd2+ concentration equals to that of the
probe. This result further verifies that the APQ–Cd2+ complex is
formed by 1 : 1 binding mode.
As to the quantum yield, we find that APQ exhibits a
quantum yield enhancement of nearly 4-fold (form Φfree = 0.057
Fig. 5 pH sensitivity of APQ and its Cd2+ complex.
to ΦCd = 0.2124) upon binding to Cd2+. The dissociation con-
2+
stant Kd of APQ with Cd2+ was tested in PBS (phosphate buf-
fered saline) buffer and calculated to be 2.72 × 10−9 M,
suggesting high sensitivity for Cd2+. We conclude that APQ can
detect free Cd2+ in the nanomolar range, and therefore affords
sufficient sensitivity for Cd2+ detection in living cells.
Ion selectivity and pH stability
Metal ion selectivity is another important character of the fluor-
escent probe. As shown in Fig. 4, APQ can selectively respond
between Cd2+ and other metal ions in both the fluorescence
intensity and emission wavelength shift. Upon addition of 1
equiv. Cd2+, the fluorescence intensity increases by 8.3 fold.
Simultaneously, the maximal emission wavelength of APQ
shifts from 475 nm to 520 nm and this phenomenon is similar as
previously reported for quinoline based probes.39,40 The ion
competitive experiments indicate that Ca2+, Cr3+, Cu+, Fe2+,
Fe3+, Hg2+, K+, Mg2+, Mn2+, Na+, Pb2+, and more importantly
Zn2+, exert little effect on the emission intensity. Moreover,
Co2+, Cu2+ and Ni2+ quench the fluorescence.28,30 Note that pre-
vious Cd2+ probes often suffer from low selectivity between
Cd2+ and Zn2+. By comparison, APQ can be considered as a
good candidate for monitoring Cd2+ flux in biological systems.
Furthermore, we examined the pH-dependence of APQ and
its Cd2+ complex (Fig. 5). In the biologically relevant pH range
Fig. 6 Two-photon excitation spectra of APQ with and without Cd2+
.
(e.g. 5–9), the fluorescence intensity of APQ and the APQ–Cd2+
complex are found to be almost pH insensitive. This phenom-
enon indicates that pH changes do not affect the fluorescence
signal output of APQ-Cd2+ under physiological conditions and,
therefore, expands its application to physiological studies.
Two-photon cross sections
We further determine the two-photon cross sections of APQ and
its Cd2+ complexes using the two-photon induced fluorescence
measurement technique. As shown in Fig. 6, APQ shows the
δmax value of 206 GM at 720 nm. Upon addition of 1.2 equiv. of
6192 | Dalton Trans., 2012, 41, 6189–6194
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Cd2+, the δmax value increases by about 200 GM. It is critical to
point out that the above two-photon cross section values are sig-
nificantly larger than the previously reported ones for other Cd2+
probes. Therefore, APQ with an enlarged two-photon cross
section should act as a valuable tool to image Cd2+ flux in living
systems under two-photon excitation.
Cell cytotoxicity and two-photon bio-imaging
Cytotoxicity is a potential side effect of many organic probes
when used in living cells or tissues. The use of a fluorescent
probe in biological systems must meet the criterion that the
probe should not interact or interfere with the biological system.
To this end, we conduct cytotoxic assays in HeLa cells for APQ.
As shown in Fig. 7, MTT assay demonstrates that the cell viabi-
lity remains more than 90% after being treated with 10 μM
APQ. As a result, APQ show little cytotoxicity for long period
incubation and should be safe when used for two-photon bio-
imaging.
Finally, we investigate the utility of APQ for monitoring the
intracellular Cd2+ flux under two-photon excitation. HeLa cells
are cultured and stained with APQ within 30 min and washed
once in DMEM containing no FCS prior to imaging or Cd2+
addition. TPM images are obtained by exciting the probe at
wavelength 800 nm. As shown in Fig. 8, because APQ can
penetrate the cell membrane, before the addition of Cd2+ APQ-
labeled HeLa cells display very weak intracellular fluorescence.
The system then exhibits a greatly increased fluorescence both
from optical windows at 450–480 nm and 510–540 nm upon the
addition of Cd2+ (20 μM) within 0.5 h. These findings clearly
demonstrate that our new probe can readily reveal the variation
of intracellular Cd2+ flux under two-photon excitation.
Fig. 7 Cytotoxicity data of APQ (HeLa cells incubated for 24 h).
Note that we recently also developed two highly selective
two-photon zinc probes 6-MPVQ and 6-MPQ (Fig. 9a). They
were designed and synthesized based on an identical lumophore
coupled to a different ligand.41 To explain the selectivity
observed for the different probes, we have tested the selectivity
of APQ between Cd2+ and Zn2+ by MALDI-TOF mass. The
Fig. 8 Two-photon image of Hela cells labeled with 20 μM APQ after
30 min of incubation, washed with PBS buffer. λex = 800 nm. (A1)
Emission wavelength from 450 to 480 nm; (B1) Emission wavelength
from 510 to 540 nm; (C1) Bright-field of HeLa cells; (D1) The overlay
of A1 and B1. TPF image of APQ labeled HeLa after 0.5 h treated with
20 μM CdCl2 water solution, λex = 800 nm. (A2) Emission wavelength
from 450 to 480 nm; (B2) Emission wavelength from 510 to 540 nm;
(C2) Bright-field of HeLa cells; (D2) The overlay of A2 and B2.
Fig. 9 (a) Structure of 6-MPQ, 6-MPVQ, APQ and APPQ; (b) fluor-
escence response of APPQ to different metal ions (1–16: Ca2+, Cd2+
Co2+, Cr3+, Cu+, Fe2+, Fe3+, Hg2+, K+, Mg2+, Mn2+, Na+ Ni2+, Pb2+
Zn2+, Ag+) in Tris buffer (pH = 7.4, λex = 360 nm).
,
,
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results verified that APQ favors Cd2+ over Zn2+, consistent with
the binding experiments. Specifically, as shown in Fig. S3 in the
ESI,† APQ is fully converted to the APQ–Cd2+ complex in the
presence of Cd2+ and Zn2+. On the other hand, the binding
between APQ and Zn2+ is relatively weak when only Zn2+ is
added (Fig. S4†).
6 M. Mameli, M. C. Aragoni, M. Arca, C. Caltagirone, F. Demartin,
G. Farruggia, G. De Filippo, F. A. Devillanova, A. Garau, F. Isaia,
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455.
To further understand the ion selectivity, we have synthesized
another compound APPQ with an identical lumophore coupled
to a more flexible binding group. As shown in Fig. 9b, APPQ
can bind to Ag2+ the most strongly. Cd2+ and Zn2+ can also bind
to APPQ, but both with lower strengths. Therefore, the structure
of the binding group indeed exerts a strong influence on ion
selectivity.
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In theory, the atomic radius of Zn2+ is smaller than that of
Cd2+ and therefore, Cd2+ is more sensitive to the steric effect
than Zn2+. On the other hand, the d orbitals of Cd2+ are more
diffused than those of Zn2+, making Cd2+ more electron-
deficient and more sensitive to the electronic effects than Zn2+.
Thus, Zn2+ tends to coordinate with a rigid ligand like 6-MPQ,
whereas Cd2+ favors the coordination with the more electron-
donating and more flexible APQ ligand.
Conclusion
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Soc., 2004, 126, 13377.
In conclusion, we have designed and developed a 6-substituted
quinoline-based two-photon Cd2+ probe (APQ) and investigated
its two-photon activities in vitro and in vivo. The new probe
shows a large red shift upon Cd2+ binding with 8-fold emission
enhancement. Importantly, APQ exhibits high ion selectivity and
sensitivity for Cd2+, especially to discriminate Cd2+ from Zn2+.
Compared with previous reported Cd2+ probes, APQ exhibits a
large two-photon cross section of ca. 400 GM. Finally, in vivo
two-photon microscopy imaging demonstrates that the new
probe is cell permeable and can act as a good tool to monitor the
Cd2+ flux in living cells.
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Acknowledgements
This study was supported by NSFC (21102002, 20932006 and
21102083), Natural Science Foundation of Education Depart-
ment of Anhui Province (KJ2010A028, KJ2011A018), and 211
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