Highly selective on-off fluorescence recognition of Fe3+ based on a coumarin derivative and its application in live-cell imaging

Article abstract of DOI:10.1016/j.saa.2017.07.068

A novel coumarin chemosensor, 7-hydroxy-2-oxo-N-(pyridin-2-ylmethyl)chromene-3-carboxamide (Probe 1), demonstrated significant selectivity towards Fe³⁺ ions. Probe 1 exhibited high fluorescence emission profile at 447?nm, excellent selectivity towards Fe³⁺ over other biologically important metal ions (Al³⁺, Ba²⁺, Co²⁺, Cu²⁺, Zn²⁺, Cd²⁺, Hg²⁺, Pb²⁺ and Sn²⁺). Interestingly, there was ~?30-fold decrease in fluorescence intensity upon Fe³⁺ binding. The limit of detection of Fe³⁺ was found to be 0.76?μM (~?40?ppb). Probe 1 also exhibited high potential as an intracellular chemosensor for Fe³⁺.

Full text of DOI:10.1016/j.saa.2017.07.068

Accepted Manuscript
Highly selective on-off fluorescence recognition of Fe3+ based on
a coumarin derivative and its application in live-cell imaging
Sona Warrier, Prashant S. Kharkar
PII:
S1386-1425(17)30628-5
doi: 10.1016/j.saa.2017.07.068
SAA 15355
DOI:
Reference:
Spectrochimica Acta Part A: Molecular and Biomolecular
Spectroscopy
To appear in:
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Revised date:
Accepted date:
24 April 2017
15 July 2017
30 July 2017
Please cite this article as: Sona Warrier, Prashant S. Kharkar , Highly selective on-off
fluorescence recognition of Fe3+ based on a coumarin derivative and its application in
live-cell imaging, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy
(2017), doi: 10.1016/j.saa.2017.07.068
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ACCEPTED MANUSCRIPT
3
+
Highly selective on-off fluorescence recognition of Fe based on a coumarin derivative and
its application in live-cell imaging
Sona Warrier and Prashant S. Kharkar*
Department of Pharmaceutical Chemistry, Shobhaben Pratapbhai Patel School of Pharmacy
and Technology Management, SVKM’s NMIMS,
V. L. Mehta Road, Vile Parle (West), Mumbai-400 056. India.

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Abstract
A
novel coumarin chemosensor, 7‐hydroxy‐2‐oxo‐N‐(pyridin‐2‐ylmethyl)chromene‐3-
3
+
carboxamide (Probe 1), demonstrated significant selectivity towards Fe ions. Probe 1
3
+
exhibited high fluorescence emission profile at 447 nm, excellent selectivity towards Fe over
3+
2+
2+
2+
2+
2+
2+
2+
2+
other biologically important metal ions (Al , Ba , Co , Cu , Zn , Cd , Hg , Pb and Sn ).
3
+
Interestingly, there was ~30-fold decrease in fluorescence intensity upon Fe binding. The limit
3
+
of detection of Fe was found to be 0.76 μM (~ 40 ppb). Probe 1 also exhibited high potential
3
+
as an intracellular chemosensor for Fe .
Keywords
3
+
Coumarin, fluorescent chemosensor, fluorescence quenching, Fe probes, live-cell imaging
1
. Introduction
The role of iron as a micronutrient in cellular functions is crucial for various life processes. Iron
is essential for cellular metabolism as it forms a part of the structure of many enzymes and
proteins [1]. Iron is distributed in hemochrome which consists of > 50% of total iron in the body.
Nevertheless, up-regulation (hyperferremia) or down-regulation (hypoferremia) of iron
concentrations in the body gives rise to disease conditions related to iron trafficking and storage
[
2]. High concentrations of iron cause nausea, abdominal pain, gastric problems and
hyperferremic and hypotransferrinemic disorders. Likewise, its deficiency causes anaemia and
related discrepancies such as impaired cognitive function, lethargy, low immunity, etc. When
2
+
3+
blood comes in contact with atmospheric air, Fe gets oxidised to Fe that causes a colour
change from red to brown. Although iron is absorbed and largely used in its iron (II) oxidation
state, it is stored in ferritin as the iron (III) state. Homeostasis of iron is a significant aspect in
neuro-inflammation and progression of Alzheimer’s disease [3-5]. Although the human body can

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balance iron concentrations to some extent, it is important to utilize analytical techniques for
sensitive detection of iron for biomedical purposes [6].
Various techniques are available for the detection of metal ions, which include atomic absorption
spectroscopy (AAS), voltammetry, colorimetry and electrochemical methods. The downside of
such techniques is that they require sophisticated equipments, tedious sample preparation and
specialized skills [7-8]. A recent advancement in analytical technology is the field of
chemosensors. These are fluorescent molecules which exhibit changes in their photophysical
properties in presence of a desired analyte. Their popularity can be attributed to advantages such
as sensitivity, selectivity, non-destructiveness, ease of handling and rapid visual detection [9-11].
Such chemosensors can be designed by systematic optimization of their electronic structure by
modification/addition of functional groups to achieve desired properties, which in turn are
favourable for the detection of trace levels of metal ions in biological fluids [12].
The present work utilizes coumarins as chemosensors due to their excellent spectroscopic
properties, light stability and less toxicity [13]. The design of the probe is based on fluorescence
switching mechanism wherein the photophysical property of the molecule is altered upon
interaction with the desired analyte [14-16]. According to the concept of hard-soft acid-base
theory, probes attached to the recognition moiety with N and O atoms can display good affinity
3
+
towards Fe [17]. Reflecting on these ideas, a novel chemosensor (Probe 1) was synthesized by
3
+
a facile method that detects Fe by fluorescence quenching mechanism (Scheme 1). This
molecule offers several advantages such as ease of synthesis, highly sensitive detection, good
aqueous solubility, stability and cell-staining behaviour.

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2
. Experimental
2
.1. General
Unless stated, all reagents and solvents are of analytical grade and used without further
purification. Nuclear magnetic resonance (NMR) spectra were obtained on Brucker NMR 300
instrument, and chemical shifts were given in ppm from tetramethylsilane (TMS) as internal
standard. Mass spectra (MS) were recorded on a Shimadzu 8040 LC-MS/MS system.
Fluorescence emission spectra were conducted on a JASCO FP-8000 Series fluorescence
spectrofluorometer. UV-Vis spectra were obtained on a Perkin Elmer Lambda 25 UV-Vis
spectrometer.
2
.2. Synthesis
The
synthetic
route
for
Probe
1
is
shown
in
Scheme
1.Ethyl
7
‐hydroxy‐2‐oxochromene‐3‐carboxylate (1) was prepared from 2,4-dihydroxybenzaldehyde
according to literature method [18]. Compound 1 was further hydrolysed using 0.5 M NaOH to
give 7‐hydroxy‐2‐oxochromene‐3‐carboxylic acid (2) which was further coupled with 2-
picolylamine to obtain Probe 1.
Scheme 1. Synthesis of Probe 1
Synthesis of 7‐hydroxy‐2‐oxo‐N‐(pyridin‐2‐ylmethyl) chromene‐3- carboxamide (Probe 1).
To a stirred solution of 2-picolylamine (280 mg, 2.59 mmol) and 2 (750 mg, 3.88 mmol) in
anhydrous DMF (4 mL), DIEA (2.2 mL, 12.91 mmol) was added slowly. With stirring, HATU
(1.6 g, 5.0 mmol) was added and the reaction was warmed to 50°C and stirred further for 3 h.

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The reaction mixture was then diluted with water and extracted with EtOAc (3x20 mL). The
combined organic layers were dried with anhydrous NaSO and concentrated in vacuo to yield
4
the crude product which was purified by column chromatography using EtOAc : pet ether (7:3)
1
as eluent to obtain Probe 1 as yellow solid (820 mg, 76%). H NMR (400MHz, DMSO-d6)
(
δppm): 4.65 (d, 2H, -NH-CH ), 6.9 (dd, 2H, coumarin-H), 7.3 (m, 2H, coumarin-H, Pyr-H), 7.7
2
(m, 2H, coumarin-H, Pyr-H), 8.55 (d, 1H, Pyr-H), 8.8 (s, 1H, -OH), 9.4 (t, 1H, Pyr-H), 11.08(s,
+
1
H, -NH-); LC-MS (ESI): 297 [M+H] .
2
.3. UV–Vis and fluorescence spectral measurements
Aqueous stock solutions (10 mM) of the appropriate metal salts were prepared. Stock
solution of Probe 1 (1 mM) was prepared in DMSO. Further, working standards were freshly
prepared. For all fluorescence measurements, excitation was set at 385 nm with excitation and
emission slit widths set at 5.0 nm. UV/Vis and fluorescence titration experiments were
performed using 10 µM and 1 µM of Probe 1, respectively, in water with varying concentrations
of the metal salts. The metal salts used were AlCl , BaCl , CuCl .2H O, CoCl . 6H O, CdCl ,
3
2
2
2
2
2
2
FeCl , HgCl , PbCl , MgCl , ZnCl , and SnCl . 2H O.
3
2
2
2
2
2
2
3
+
2
.4. Fluorescence titration involving Fe
3
+
The titration experiments were performed at Probe 1 concentration 1 μM and Fe concentrations
at 0.0, 0.2, 0.4, 0.5, 0.8, 1, 2, 4, 5, 10 μM. Fluorescence spectra were recorded at RT.
2
.5. Solubility and effect on pH
Direct kinetic solubility was performed with Probe 1 using standard procedures [19] to confirm
the aqueous solubility of the probe as well as to explore its behaviour in different pH solutions.

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2
.6. Effect of solvents
The fluorescence behaviour of Probe 1 in the presence of different solvents with varying
polarity was investigated. 1 µM solution of the Probe 1 was prepared in MeOH, EtOAc, EtOH,
DMF and DMSO and their fluorescence spectra were recorded.
2
.7. Cell imaging
LLC-MK2 cell lines were used for fluorescence imaging. Cell lines were cultured in Dulbecco’s
Modified Eagle’s medium (Gibco BRL, USA), supplemented with 10% (v/v) heat-inactivated
fetal calf serum (FCS) (cell Clone), 100 µg/mL penicillin,100 µg/mL streptomycin, and 0.25
mM L-glutamine, at 37 °C in 5% CO humidified air. As soon as the cells reached confluency,
2
5
cell density was adjusted to 1.0 × 10 cells/well in culture media. To determine the cell
3
+
permeability of Fe , the cells were incubated with Probe 1 (1µM) for 30 mins at 37 °C. Upon
3
+
addition of 10 equivalents of Fe onto the cells, the images were observed under phase-contrast
microscope with DAPI filter.
2
.8. MTT Assay
4
LLC-MK2 cell in the concentration of 5 × 10 cells/mL were seeded into a 96-well, microplate
Nunc, DNK). 100μL of complete medium was added to all wells and plates were incubated in
% CO and 37 °C. After 24 hours, the cells were treated with Probe 1 (1, 10, 50, 100, 200 µM)
(
5
2
then incubated under 5% CO at 37 °C. Post-24 hrs, cells were washed with DPBS followed by
2
addition of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] solution
(
5mg/mL, Sigma) and again incubated under 5% CO at 37 °C for 4 hrs. The MTT solution was
2
removed and 150 µL of DMSO was added to dissolve the crystals and incubated for 10 mins.
The optimal density (OD) value was measured by a microplate reader (Epoch 2 Biotek) at 570

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nm. The cell viability was calculated as (mean OD in treated wells/mean OD in control wells) ×
1
3
3
00.
. Results and Discussion
.1. UV–Vis spectral measurements
3
+
2+
2+
2+
2+
2+
2+
3+
2+
When 10 equivalents of various metal ions (Al , Ba , Co , Cu , Zn , Cd , Hg , Fe , Pb
2
+
2+
2+
3+
and Sn ) were added to Probe 1 (10 μM), only Cu , Co and Fe produced an instant color
change (colourless to light yellow). The interaction of Probe 1 with various metal ions was
initially studied by UV–Vis spectroscopy (Figure 1). The absorbance of Probe 1 decreased
3
+
significantly, compared to other metal ions in the presence of Fe ions. It was observed that the
3
+
interaction between Probe 1 and Fe was very fast and efficient.
0
0
0
.25
Probe 1+Al
Probe 1+Ba
Probe 1+Cd
Probe 1+Co
Probe 1+Cu
Probe 1+Fe
Probe 1+Hg
Probe 1+Pb
Probe 1+Sn
Probe 1+Zn
Probe 1 Std
0
.2
.15
0
.1
.05
0
3
20
370
420
470
520
-
0.05
Wavelength (nm)
Figure 1.
UV–Vis absorption spectra of Probe 1 (10 μM) upon addition of respective metal
ions (100 μM) in water.

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3
.2. Fluorescence spectral measurements
Furthermore, the photophysical properties of Probe 1 in the presence of a wide range of
3
+
2+
2+
2+
2+
environmentally and physiologically significant metal ions such as Al , Zn , Cu , Ba , Co ,
2
+
2+
2+
2+
3+
Sn , Cd , Pb and Hg as well as Fe were investigated. Probe 1 (1μM) showed a significant
emission upon excitation at 385 nm (Figure 2a). There was no obvious spectral change in the
intensity of Probe 1 when other cations were added. However, major quenching of fluorescence
3
+
3+
2+
2+
intensity occurred with Fe ions. There was slight quenching observed with Al , Sn and Hg
while other cations revealed almost insignificant response (Figure 2b). Thus, it can be
3
+
established that the selectivity of Probe 1 for Fe ions over other cations, in aqueous media, is
remarkably high.
a)
b)

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1
1
.4
.2
1
0
0
0
0
.8
.6
.4
.2
0
Free
Al
Ba
Cd
Co
Cu
Fe
Hg
Pb
Sn
Zn
Probe
Figure 2.
a) Fluorescence spectra of Probe 1 (1 μM) upon addition of metal salts (10
3
+
2+
2+
2+
2+
2+
2+
3+
2+
2+
equiv.) Al , Ba , Co , Cu , Zn , Cd , Hg , Fe , Pb and Sn at  of 385 nm, b) relative
ex
responses at 447 nm ( ) of Probe 1 upon addition of various metal ions upon excitation at 385
em
nm.
To further validate the utility of Probe 1, fluorescence titration experiment was carried out in the
3
+
3+
presence and absence of Fe by mixing Probe 1 (1μM) with increasing concentrations of Fe
(0.2, 0.4, 0.5, 0.8, 1, 2, 4, 5, 10 equivalents) (Figure 3).The signals were measured within one
3
+
3+
minute of addition of Fe ions. As Fe concentration increased, the high fluorescence intensity
of Probe 1 at 447 nm decreased almost 30-fold when excited at 385 nm ). The fluorescence
ex
quenching occurred in a linear fashion as the concentration of the analyte increased.

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0
equivalent
1
0 equivalents
Figure 3.
Fluorescence titration spectra of Probe 1 (1 μM, λ = 385 nm) upon addition of
ex
3
+
increasing concentrations of Fe ions metal chloride (0, 0.2, 0.4, 0.5, 0.8, 1, 2, 4, 5, 10 μM) at
room temperature.
3
.3. Determination of detection limit
To analyse the quenching process of the fluorophore-metal system, Stern-Volmer equation
Equation 1) was used for determining the type of quenching. Linear plot of Stern-Volmer
(
establishes that the quenching is a static type of quenching and not dynamic quenching [20,21].
I /I= 1 + K [Q]...................................................................................... (1)
0
SV
3
+
where I and I are the fluorescence intensities at 447 nm in the absence and presence of Fe
0
ions, respectively. KSV is the Stern–Volmer fluorescence quenching constant, which is related to
3
+
the quenching efficiency of the quencher, and [Q] is the concentration of the quencher, [Fe ].
In the presence of different concentrations of the quencher, fluorescence intensity was reduced
3
+
from I to I. The fluorescence data were then analysed by plotting a graph of I /I versus [Fe ].
0
0
The Stern–Volmer plot of I /I versus [Q] (Figure 4) showed a linear relationship between the
0

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3
+
-6
-6
fluorescence response and [Fe ] in the range of 0.2x10 M to 10x10 M with a correlation
coefficient of 0.995. The linear Stern–Volmer plot indicated that the quenching was static for
3
+
3+
Probe 1-Fe system. From the linearity graph, the limit of detection (LOD) for Fe was
calculated to be 0.76 μM, which is much lower than WHO (World Health Organization)
3
+
2
guidelines (5 μM) for Fe ions in drinking water.
25
20
15
10
5
0
y = 2.0127x + 0.2129
R² = 0.9956
0
2
4
6
8
10
12
3
+
[
Fe ] (μM)
3
+
Figure 4.
.4, 0.5, 0.8, 1, 2, 4, 5, 10 µM) are normalized between the minimum (0.0 µM Fe ) and the
maximum emission intensities.
Emission (at 447 nm) of Probe 1 (1 µM) at different concentrations of Fe (0.2,
3
+
0
3
.4.Selectivity of Probe 1
3
+
The interference of other metal cations that could possibly coexist with Fe was explored by
3
+
carrying out competition experiments. To utilize Probe 1 as a selective sensor for Fe ions, the
fluorescence responses of Probe 1 was investigated in the presence of co-existing metal ions
3
+
2+
2+
2+
2+
2+
2+
2+
2+
+
+
2+
2+
2+
such as Al , Ba , Cd , Cu , Co , Hg , Pb , Sn , Zn , Na , K , Mg , Mn , Ca . As
shown in Figure 5, the presence of other metal ions did not interfere the quenching of Probe 1

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3
+
3+
by Fe and the intensity changes due to Fe were not affected by the addition of co-existing
3
+
metal ions. The results show that Probe 1 was a selective On-Off sensor for Fe .
3
+
Figure 5. Fluorescence emission responses of Probe 1 containing Fe and other co-existing
metal ions; λ = 385 nm.
ex
3
+
3
.5. Ratiometric measurements of Probe 1 in the presence of Fe ions
3
+
Since there is no shift in the emission wavelength of Probe 1 upon addition of Fe ions,
ratiometric measurements were carried out by employing another fluorescent probe having a
3
+
different λmax and that has Fe chelating properties. 8-hydroxyquinoline (8-HQ) is extensively
3
+
used in iron chelation therapy and has affinity towards Fe ions. Therefore, 8-hydroxyquinoline
was employed for ratiometric calculations (λmax= 255 nm). The UV-Vis absorbances of Probe 1
3
+
(
λmax=385 nm) and 8-HQ (λmax= 255 nm) were measured in the presence of Fe ions. Upon the
3
+
addition of increasing amounts of Fe , the absorbance ratio at 385 nm and 255 nm (A nm/
385
3
+
A255 nm) also decreased linearly with the concentration of Fe (0-10 equiv) (Figure 6). These

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ratiometric measurements were performed to allow correction of subtle variations independent
of the sample conditions (e.g. concentration of the probe, sample thickness, intensity of
illumination).
3
+
Figure 6. Ratiometric calibration curve A385nm/ A255nm as a function of Fe concentration.
3
.6.Aqueous solubility and the effect of pH
Aqueous solubility and stability forms an integral part in the discovery of fluorescent probes
since it has a great influence on assays involving biological systems such as cells. The solubility
of Probe 1 was explored by direct "kinetic" solubility conditions. The probe was also subjected
to different pH conditions for 2 days. The probe was found to have a aqueous solubility of more
than 10 μM and that it was stable over a range of pH (1-8).
3
.7. Effect of solvents
To get a view of the practical application potential, particularly in material sciences, we
examined the fluorescence intensity of Probe 1 in different solvents. Since, in many cases, the
influence of solvents is significant for the fluorescence spectra of complexes, fluorescence
emissions of Probe 1 (1μM) in different solvents were explored (Figure 5) [22-25]. There was

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no significant change in the fluorescence intensity of Probe 1 in different solvents like MeOH,
EtOH, EtOAc, DMF and DMSO. Thus, it can be deduced that Probe 1 can be employed with a
variety of solvents without significant changes in its photophysical properties.
Figure 7. Fluorescence emission spectra of Probe 1 (1μM) in different solvents.
3
.8. Mechanism of fluorescence quenching
The presence of a 7-OH group and 3-CONH- group in Probe 1 make it highly fluorescent due to
the transfer of electrons within the molecule, like a push-pull system. Therefore, when chelation
occurs, there is a transfer of charges within the Probe 1-metal system which in turn causes a
decrease in the fluorescence intensity of the molecule [26,27]. Consequently, it would be rational
3
+
to infer that the mechanism of fluorescence quenching of Probe 1 in the presence Fe could be
attributed to ligand-metal charge transfer (LMCT) mechanism. There occurs a charge transfer
3
+
which is initiated by the binding of Fe ions to Probe 1. This hypothesis is supported by the fact
3
+
that Fe is known to be paramagnetic with an unfilled d shell that would take part in the energy
3
+
and/or electron transfer processes leading to quenching of the fluorescence [28-30]. When Fe
comes in contact with Probe 1, the fluorophore opens a nonradiative deactivation channel

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induced by unfilled d shell and paramagnetism, to cause electron transfer that is, resulting in
3
+
quenching of fluorescence. The interaction between Probe 1 and Fe could occur at C=O, –NH
and the nitrogen in pyridine ring which could be identified as binding to transition and post-
transition metals [31,32]. Hence, the mechanism of LMCT could occur rapidly due to strong
3
+
paramagnetic quenching property of Fe that covers all other mechanisms for fluorescence
quenching.
3
.9. Cell imaging and MTT assay
Further, the practical application of Probe 1 was explored as an intracellular chemosensor for
3
+
Fe (Figure 6). The imaging of cells by Probe 1 was assessed by phase-contrast microscopy
with DAPI fluorescence filter. As seen from the bright-field image, LLC-MK2 cells incubated
with 1 μM of Probe 1 for 30 min at 37 °C show cell viability (Figure 6a) with strong
intracellular background fluorescence (Figure 6b). The cells that were pre-incubated with Probe
3
+
1
were supplemented with 10 μM of Fe for 10 min at 37 °C that reduced the intracellular
background fluorescence (Figure 6c). These results demonstrated the potential utility of Probe 1
3
+
as cell- permeable, chemosensor of Fe in living cells.
a
b
c
3
+
Figure 8.
Live-cell imaging of intracellular Fe : (a) Bright field transmission image of
LLC-MK2 cells incubated with 1 μM of Probe 1 (b) Fluorescence image of LLC-MK2 cells
supplemented with 1 μM of Probe 1 in the growth media for 30 min at 37°C (c) LLC-MK2 cells

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incubated with 1 μM of Probe 1 in the growth media for 30 min at 37°C and then further
incubated with 10 μM FeCl for 10 min at 37°C.
3
Consecutively, the toxicity evaluation of Probe 1 was carried out by MTT assay that exhibited
dose-dependent decrease in cell viability. There was no significant loss of viable cells at lower
concentrations indicating relatively non-toxic nature of Probe 1. This observation supports
potential utility of Probe 1 for cell biology applications (Figure 7).
Figure 9.
Cytotoxicity of Probe 1 on LLC-MK2 cells. Cell viability was assayed by the
MTT method (values: mean ± s.d.)
3
.10. Performance comparison
3
+
The analytical performance of the probe proposed in the present work towards Fe was
compared with other reported fluorescent probes based on various fluorophore derivatives as
shown in Table S1 (ESI). Although it can be seen that most of these probes have good selectivity
3
+
2+
2+
towards Fe , some have dual-sensing mechanisms that involves Fe and Cu that would make
3
+
2+
them less selective towards Fe , in the presence of Cu [21,34,38]. The fluorescent probe
3
+
developed in this work exhibits selectivity only towards Fe and has no interference from other
transition metals. Some of the fluorescent probes listed in the table do not mention their

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detection limits [7,20,37], while some probes have higher detection limits than the probe
presented in this work [21,36]. Therefore, overcoming the disadvantages of expensive starting
materials [20], lower yields and narrow linear range, this small molecule fluorescent probe aims
to provide a valuable sensor for further biomedical applications.
4
. Conclusion
In conclusion, a novel coumarin chemosensor was synthesized by a simplistic method using
3
+
inexpensive starting materials and evaluated for its sensing abilities towards Fe ions. Probe 1
3
+
demonstrated highly selective fluorescence turn-off response only in the presence of Fe .
LMCT mechanism is known to facilitate the quenching processes by the fluorophore. Probe 1
shows no interference from other metal ions. The limit of detection for Probe 1 was found to be
3
+
0
.76 μM. Probe 1 can also be utilized in studying the cellular functions involving Fe in various
disease and pathological conditions.
Conflict of Interest
The authors confirm that this article content has no conflict of interest.
Acknowledgements
Ms. Sona would like to acknowledge SVKM’s NMIMS for PhD fellowship during the course of
this work. The authors acknowledge the help and valuable assistance of Dr. Nishigandha Naik
and Ms. Amruta Naik from Haffkine Institute for Training, Research and Testing,Parel, Mumbai
for providing facility for fluorescence spectrophotometer.

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Graphical Abstract

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Highlights
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Probe 1 demonstrated highly selective on-off fluorescence response in the presence of Fe
ions.
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The coumarin-based fluorescent probe exhibited ‘ligand-metal charge transfer’ mechanism.
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The detection limit for Fe was found to be 0.76 μM.
Probe 1 can potentially be used for live cell imaging for Fe detection.
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