Fluorescence sensing and intracellular imaging of Pd2+ ions by a novel coumarinyl-rhodamine Schiff base

Article abstract of DOI:10.1039/c8nj06511j

Coumarinyl-rhodamine, HCR, served as an extremely selective sensor for Pd²⁺ ions in ethanol/H₂O (8:2, v/v, HEPES buffer, pH 7.2) solution and the limit of detection (LOD) was 18.8 nM (3σ method). The free sensor, HCR, was weakly emissive and in the presence of Pd²⁺, the colour changed from straw to pink with very strong emission at 598 nm in the presence of eighteen other cations. A plausible mechanism involved opening of the spirolactam ring of rhodamine on interaction with Pd²⁺, which was justified by structure optimization and transition energy calculations using the DFT technique. HCR underwent 1:1 complexation with Pd²⁺, which was confirmed via the Job's plot, mass spectra and Benesi-Hildebrand plot (association constant K_a, 9.1 × 10⁴ M^-1). A separate in vitro experiment showed that HCR could specifically sense Pd²⁺ in MCF7 (human breast adenocarcinoma) cell lines.

Full text of DOI:10.1039/c8nj06511j

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Fluorescence sensing and intracellular imaging of Pd2+ ion by a novel
coumarinyl-rhodamine Schiff base†
Arup Kumar Adak,a,b Rakesh Purkait , Saikat Manna, Bankim Chandra Ghosh,d Sudipta
b
c
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Pathak and Chittaranjan Sinha *
a
Department of Chemistry, Bidhannagar College, EB-2, Sector –1, Salt Lake, Kolkata‒700064,
West Bengal, India.
b
Department of Chemistry, Jadavpur University, Kolkata‒700032, West Bengal, India.
cDepartment of Chemistry, Haldia Govt. College, Debhog, Haldia, Purba Medinipur‒721657,
West Bengal, India.
dDepartment of Chemistry, Durgapur Govt. College, J. N. Avenue, Durgapur, Paschim
Barddhaman ‒713214, West Bengal, India.
Abstract
2
+
Coumarinyl-rhodamine, HCR, serves as extremely selective sensor to Pd ion in ethanol/H O
2
(8:2, v/v, HEPES buffer, pH 7.2) solution and the limit of detection (LOD) is 18.8 nM (3
2+
method). The free sensor, HCR, is weakly emissive and in presence of Pd , the colour changes
from straw to pink with very strong emission at 598 nm in presence of eighteen other cations.
The plausible mechanism includes opening of the spirolactam ring of rhodamine upon interaction
2+
with Pd . This is justified by a structure optimization and transition energy calculation by DFT
2
+
technique. HCR undergoes 1:1 complexation with Pd that has been confirmed via Job’s plot,
4
1
mass spectra and Bensei-Hildebrand plot (association constant Ka, 9.110 M ). A separate in

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vitro experiment shows that HCR can specifically perceive Pd2+ in MCF7 (Human breast
adenocarcinoma) cell lines.
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Keywords: Coumarinyl-rhodamine, Pd -sensor, spirolactam ring opening, live cell imaging,
DFT computation.
_
_________________
Corresponding Author: crsjuchem@gmail.com (C. Sinha)
Electronic supplementary information (ESI) available. See DOI: xxxxxxxxxxxx
†
1. Introduction
During the past few years, the development of methods for the identification and sensing of
platinum group metals (PGMs) has received amazing interest because of their enormous
biological, environmental, industrial and chemical significance. 1-4 Among these, palladium, a
precious metal, has become one of the most attractive sensing targets in recent years because of
its wide catalytic use in synthesis of organic and pharmaceutical molecules, fuel cells, dental
appliances, medical devices, electrical equipments etc. 5,6 In the Pd-catalysed reactions, it may
apprehend that Pd(0) oxidation state may temporarily oxidize or those of Pd(IV) may reduce to
the intermediate oxidation state Pd(II) as the reaction proceeds. The final product is often
contaminated with impurity of Pd(II) even after rigorous purification. Such contamination in
ultratrace level in the industrial products may cause serious health problem. Due to its thiophilic
nature, palladium can bind with DNA, proteins and other macromolecules, and disturb a variety
of cellular processes.7, 8 Palladium also hampers the activity of many enzymatic reactions such as
alkaline phosphatase, creatine kinase and prolyl hydroxylase (Hypoxia-inducible factor) etc.9,10
For these reasons, various analytical techniques are used for the quantitative analysis of

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palladium. Popular techniques are inductively coupled plasma-mass spectrometry (ICP-MS),
atomic absorption spectrometry (AAS), solid-phase micro-extraction high performance liquid
chromatography (SPME-HPLC) and X-ray fluorescence spectroscopy.11-13 However; these
methods require sophisticated highly expensive equipments, extensive sample preparation steps,
time-consuming and high salaried experts. Among these different techniques for the analysis of
palladium ions, colorimetric and fluorometric techniques are more convenient and dependable
for the speedy and sensitive detection of palladium both qualitatively and quantitatively in light
of their simplicity, selectivity and sensitivity. Hence, the development of palladium selective
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fluorescent probes is extremely essential. Pd , being a heavy transition metal ion with an open
1
4
15
shell electronic configuration, is a typical fluorescence quencher. Based upon the ON–OFF
16,17
or OFF–ON
mechanism some fluorescent chemosensors and chemodosimeters are
synthesized for the identification of palladium species. Towards the design of sensor, the
important objectives are the long-wavelength emission and ecofriendly availability of fluorescent
chemosensors. With this views Rhodamine functionalized chemosensors are of increasing
1
8
interest in current years. For the last couple of years we have taken a strategy to design
Rhodamine derivatives for the identification of trace level of Pd; allyl ether Schiff base of
1
9
Rhodamine has been tested for detection of total Pd at 50 nM while allyl ether hydrazone
2
0
Rhodamine is capable to detect as low as 95 nM concentration at pH 7.2. The performance of
Rhodamine derivatives has inspired us to synthesize newer chemosensor those could able to
detect even lower concentration of Pd(II). For further accomplishment, we have selected two
fluorogenic units, coumarin and rhodamine, to combine together in a single molecule, (E)-3',6'-
bis(diethylamino)-2-(2-(((7-hydroxy-4-methyl-2-oxo-2H-chromen-8-yl)methylene)amino)ethyl)
spiro[isoindoline-1,9'-xanthen]-3-one (HCR) (Scheme 1) and has been used for the detection of

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2+
2+
Pd . HCR is nearly non-emissive while its complexation with Pd makes powerfully
fluorescent motif within the red wavelength region (598 nm) that may be due to opening of
spirolactam ring of rhodamine. The potential application of the ligand (HCR) has been carried
out by intracellular imaging in the MCF7 (human breast adenocarcinoma) cell line in presence of
exogenous palladium ions.
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Me
Me
O
HO
O
O
O
N
Pd2+
O
O
O
N
N
N
Pd2+
N
O
N
N
O
N
CR-Pd]+
[
HCR
2
+
Scheme 1: Proposed sensing mechanism of Pd ‒selective sensor HCR (  598 nm for the
em
complex).
2
2
. Experimental section
.1 Materials and methods
Rhodamine B (Sigma-Aldrich) and resorcinol, ehtylene diamine, ethyl acetoacetate
(
Spectrochem) were used as received. All other organic chemicals and inorganic salts were
acquired from Merck and utilized without further purification. Purified water was collected from
Milli-Q water (Millipore). Following instruments were used for different physicochemical
measurements : Perkin-Elmer 2400 Series-II CHN analyzer, Perkin Elmer, USA for elemental
analyses; Perkin Elmer Lambda 25 spectrophotometer for absorption spectral measurements;

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Perkin Elmer spectrofluorimeter model LS55 for emission spectral studies; Perkin Elmer LX-1
FTIR spectrophotometer used for IR spectral collection; Bruker (AC) 300 MHz FT-NMR
spectrometer utilizing TMS as an internal standard for NMR spectra; Water HRMS model
XEVO-G2QTOF#YCA351 spectrometer used for mass spectra. Data were collected at room
temperature.
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The test sample and reference were excited at the identical wavelength. From the area of
the fluorescence spectra (using available software in the instrument) the fluorescence quantum
yield of the experimental sample was calculated using the equation,
2
S  F   A   
S
R
S
2
R



…(1)

   


R

FR
   
AS

Where, R and S are the fluorescence quantum yield of the reference and samples; F and FS
R
are the respective areas under emission spectra of the reference and sample respectively. A and
R
AS are the absorbance of the reference and sample at the excitation wave length. R and S are
the refractive index of the solvent used for the reference and the sample. Fluorescein (reported
21
quantum yield, R  0.79 in 0.1M NaOH was used as reference sample.
2
.2 Synthesis of probe, HCR
4
-Methyl-7-hydroxy-8-formylcoumarin (A)22 and N-(rhodamine-B)lactam-1,2-ethylenediamine
(
B) 23 were synthesized by reported methods. A suspension of B (1.0 equiv) in dry EtOH (10 ml)
was stirred at 80C with 4-methyl-7-hydroxy-8-formyl Coumarin (A) (1.0 equiv) in the same
solvent (10 ml) under inert (N ) environment for 2 h; and an orange-yellow colored crystalline
2
product was separated. The solvent extraction using ethyl acetate/brine water system was
performed followed by chromatographic separation over silica gel using ethyl acetate/petroleum

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ether (1:1 v/v) solvent mixture to elute the desired product, HCR. Yield: 75%, m.p.: 145-147˚C,
1
H NMR (300 MHz, CDCl ) δ 8.62 (s, 1H), 7.91 (d, J = 5 Hz, 1H), 7.44-7.42 (m, 3H), 7.09 (t, J
3
= 3 Hz, 1H), 6.70 (d, J = 9 Hz, 1H), 6.32-6.45 (m, 6H), 5.98 (s, 1H), 3.51 (t, J = 6Hz, 2H), 3.32-
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†
13
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.34 (m, 10H), 2.42 (s, 3H), 1.16 (t, J = 6.81 Hz, 12H) (ESI , Fig. S1); C NMR (75 MHz,
CDCl ) δ 172.4, 168.3, 162.3, 160.6, 160.5, 155.0, 153.6, 153.4, 153.3, 148.9, 132.6, 130.8,
3
1
1
29.5, 128.7, 128.1, 123.8, 123.0, 117.7, 109.2, 108.3, 105.2, 105.0, 97.7, 64.9, 53.5, 44.4, 40.1,
†
+
8.9, 12.6. (ESI , Fig. S2); MS ES , m/z calculated for C H N O : 670.32; found: 671.16
4
1
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4
5
+
†
−1
†
[M+H] (ESI , Fig. S3); IR (KBr pellet,cm ): 3180, 1570 (ESI , Fig. S4).
2
.3 UV–vis and fluorescence spectral studies
Ethanol solution (5 ml) of HCR (3.35 mg, 0.001 mmol) was used as stock solution; 100

L of this stock of HCR was diluted utilizing 2 ml ethanol/H O (8:2, v/v) containing HEPES
2
2
+
buffer (pH 7.2). Independently, a Pd solution was prepared by warming PdCl (1.77 mg, 0.001
2
mmol) in minimum volume of DMSO and diluted to 10 ml by adding water. Microlitre quantities
of Pd2+ solution was slowly poured to the HCR solution prepared above for spectral
measurement. Excitation wavelength of 545 nm (excitation slit = 5.0 and emission slit = 5.0) was
used for fluorescence study.
2
.4 Theoretical computation
-
2+ +
Optimized molecular geometries of HCR and [CR -Pd ] complex were generated by
DFT/B3LYP technique exploiting Gaussian 09 software.24-27 For C, H, N, O basis set of 6-311G
were used and for heavy atom Pd the LanL2DZ basis set was used.28,29 Vbrational frequency
calculations were checked with the optimized geometries which could represent the structures of

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the species closer to the actual structure. TD-DFT technique in MeOH using conductor-like
polarizable continuum model (CPCM) 30-32 was employed for theoretical evaluation of UV-vis
spectra. The fractional contributions of various groups to the molecular orbital were checked by
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GAUSSSUM program. 33
2.5 Biological studies
2
.5.1 Cell culture
The cells of MCF7, human breast adenocarcinoma cell line, (NCCS, Pune, India) were cultured
o
in CO incubator (5%) at 37 C using DMEM containing fetal bovine serum (FBS) (10%),
2
-
Kanamycin sulfate (110 mg/L), penicillin (50 units/ml) and streptomycin (50 µg/ml). The cell
detachment during cell splitting program was checked by Trypsin-EDTA (1X) solution was used
for cell detachment during cell splitting.
2
.5.2 Cell viability assay of tetramer peptides
Human breast adenocarcinoma cell line, MCF7 cells were taken in DMEM culture media with
0% fetal bovine serum. Using different concentrations of HCR ligand in DMEM medium cell
viability assay was performed within ninety six well plates. MTT (3-(4,5-dimethylthiazol-2-yl)-
,5-diphenyltetrazolium bromide) is a colorimetric technique where yellow tetrazole compound
1
2
gets reduced to purple coloured formazan by reductase enzymes present in living cells while the
dead cells are unable to interfere. Formazan was further dissolved in DMSO/MeOH (1:1 (v/v))
and absorbance of each well was measured at 550 nm by micro-plate ELISA reader. Percent of
viability was measured from absorbance value. Percentage viability has been calculated as
{
A550 (treated cell) A550 (background)}

100 .
{
A550 (untreated cell) A550 (background)}

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.5.3 Cellular uptake study by microscopic imaging
Approximately, 2000 MCF7 cells were seeded in DMEM medium containing 10% fetal
bovine serum on cover glass bottom disc for overnight before the treatment. Then, the media was
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removed and 5 µM HCR in DMEM containing media was added and incubated for 3 h (DMSO
2
+
concentration maintained 0.4%). After 3 h of incubation, 5 μM of Pd was added and incubated
for another 30 min followed by the addition of 4% formaldehyde in PBS buffer for 30 min to fix
the cell in each cover glass. Formaldehyde solution was then removed and washed with PBS
buffer. Cell imaging was performed by a spinning disc confocal microscope with 40X objective
(Olympus) equipped with AndoriXon 3897 EMCCD camera in 568 nm wavelength light.
3
3
. Results and discussions
.1 Synthesis and characterization of HCR
The
probe
(E)-3',6'-bis(diethylamino)-2-(2-(((7-hydroxy-4-methyl-2-oxo-2H-chromen-8-
yl)methylene)amino)ethyl)spiro[isoindoline-1,9'-xanthen]-3-one (HCR) is synthesized by the
reaction of 4-methyl-7-hydroxy-8-formyl-Coumarin (A) and N-(rhodamine-B)lactam-1,2-
†
-1
ethylenediamine (B) (ESI , Scheme S1). Disappearance of (CHO) of A at 1651 cm and
-1

(NH ) of B in the FTIR spectrum of HCR and the strong stretch at 1624 cm refers to (C=N)
2
-1
−1
along with lactone ν(COO), 1729 cm and (OH), 3444 cm stretching have confirmed the
+
synthesis of the probe. The mass spectral peak at 671.16 matches to [M+H] (calculated m/z for
1
C H N O is 670.32) and supports the elemental composition of HCR. The H NMR spectrum
4
1
42
4
5
of (300 MHz, CDCl ) shows singlet at 2.42 ppm which is assigned to methyl protons at
3
coumarinyl back bone. Signals at δ 3.53-3.32 ppm refer to methylene protons which are attached

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to nitrogen centres. The most deshielded protons at δ 8.62 ppm is assigned to imine proton which
proves the formation of Schiff base. The triplet signal at δ 1.16 ppm is due to the presence of –
CH protons of ethyl group. All other aromatic-Hs are usual and have been compared with
3
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3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
literature data.34 13C NMR spectrum is difficult to assign due to large number of overlapping C
centres. Ester carbonyl and amide carbonyl appear at 172.4 and 168.3 ppm respectively. Peaks at
1
8.9, 12.6 ppm are assigned to two types of methyl groups. The carbons which are attached to
nitrogen centers appear at 64.9, 53.5, 44.4, 40.1 ppm. The quaternary carbon appears as highest δ
value (64.9 ppm) among said four carbons and has been compared with literature report.35
3
.2 Absorption spectroscopic studies
HCR exhibits absorption in ethanol-water (8:2, v/v; HEPES buffer, pH, 7.2)) at 272, 317, 350
+
+
2+
2+
2+
3+
and 414 nm. The addition of following metal ions, for example, Na , K , Ca , Mg , Ba , Al ,
4
2
+
2+/3+
3+
2+
2+
2+
2+
2+
+
2+
2+
Mn , Fe
, Co , Ni , Cu , Zn , Cd , Hg , Ag , Pb , Pt , ( 10 M ) to HCR solution
does not show any significant change in visible color under indistinguishable conditions, while
6
5
2
+
on adding Pd at even lower concentration ( 10 to 10 M ) shows a reasonable improvement
2
+
of absorbance at 565 nm and a visible change of colour (Fig. 1). Addition of Pd to EtOH/H O
2
solution (8:2, v/v, HEPES buffer, pH 7.2) of HCR, pale yellow colour of free probe changes to
pink with a strong absorption band centered at 565 nm (Fig. 2). It is unanimously explained the
unique selectivity of HCR to Pd2+ over other competitive metal ions and the experiment
2
+
demands easy sensitive naked eye probe for Pd .

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0
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
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5
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5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Fig. 1: UV-vis responses of HCR to various metal ions in EtOH/H O (8:2, v/v, HEPES buffer,
2
5
2
5
4
pH 7.2). [HCR]  2.010 M
,
[Pd ] 1.010 M
,
[other ions] 10 M
.
(
a)
(b)
Pd2+

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1
1
2
3
4
5
6
7
8
9
5
Fig. 2: (a) Variation of UV-vis spectrum of sensor HCR ( 2.010 M ) in EtOH/H O (8:2 v/v,20
2
6
2
+
mM HEPES buffer, pH 7.2) medium on adding of Pd ion (0 to 9.510 M with increment of
7
5
.010 M ) gradually. (b) Magnification of the relevant portion of (a). Inset: Naked eye view
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2+
of the sensor solution before and after addition of Pd ion.
3
.3 Emission spectroscopic studies
The sensor, HCR, bears two fluorogenic units, Coumarin and Rhodamine-B; so, our
apprehension is that it could be utilized as powerful fluorogenic dyad. With the end goal to
2
+
additionally explore the selectivity of HCR to Pd , the fluorescence spectral response of HCR
2
+
to the previously mentioned metal ions have been analyzed. Upon addition of Pd to the solution
of HCR a brilliant fluorescence is observed (Fig. 3) at 598 nm with high quantum yield of the
-
2+ +
[
CR -Pd ] complex (Φ, 0.68; fluorescein standard) while HCR is very weakly emissive (Fig.
2
+
3
). This demonstrates that HCR is extremely selectivity to Pd ion. During the titration of HCR
2
+
solution by adding Pd ion, the fluorescence intensity at 598 nm gradually increases; excitation
at 545 nm enhances the fluorescence intensity by 270 times (Fig. 4). The experimental detection
2
+
limit (LOD) of Pd is 18.8 nM (3σ method, ESI†, Fig. S5). Use of a number of the rhodamine
functionalized sensor for quantitative identification of Pd2+ has been reported; for example,
34
rhodamine bonded azophenol shows LOD 450 nM , rhodamine-nitro-salicaldehyde Schiff base
exhibits LOD 50 nM , rhodamine-ethynyl-benzaldehyde gives LOD 191 nM 35, rodamine
1
9
3
6
appended oxime is accounted for with LOD 15 nM , rodamine-benzimidazile gives LOD 21
nM. 37 The present probe, HCR is astoundingly high sensitive (LOD, 18.8 nM) and it is a
2+
potential competitor in Pd sensor’s club.

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2
1
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3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
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5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2+
Fig. 3: Emission spectra of HCR on adding different metal ions and Pd in EtOH/H O (8:2
2
5
2
5
v/v,20 mM HEPES buffer, pH 7.2). [HCR]  2.010 M
,
[Pd ] 1.010 M ,
4
[
other ions] 10 M
.
-
2+ +
The complex [CR ‒Pd ] shows better excited state stability (complex, 1.62 ns) than that of free
ligand, HCR only (HCR, 0.15 ns) (Fig. 5). Enhancement of life time on binding with Pd2+
2
+
supports the stabilization at excited state by sharing orbitals of metal ion. Hence, Pd induced
cleavage of the C–O bond with consequent chelation and opening of the spirolactam ring 38-45
(Scheme 1).
The pH sensitivity of the emission of HCR shows that there is almost no change in
emissivity of the ligand in the pH range 7.0 - 12.0 while in acidic pH 3.0 – 7.0 a substantial
intensity change is observed. In presence of Pd2+ the ligand does not show any change of
2
+
emission in the pH range 2.0-12.0 (Fig. 6). Hence sensing of Pd by HCR through fluorescence

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1
3
1
2
3
4
5
6
7
8
9
enhancement is most suitable in the pH range of 7.0‒12.0, which may be attributed solely to a
spirolactam ring opening of the rhodamine moiety caused by Pd2+ ion. Thus, Pd2+ can be
detected in approximately physiological pH ranges in living cells using HCR.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
+
Fig. 4: Alteration in the fluorescence spectra of HCR upon the addition of Pd at an emission
slit of 5 nm; inset: naked eye view of the sensor solution (placed in a UV chamber) in presence
2+
and absence of Pd ions.

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4
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
+
Fig. 5 (a) Time resolved fluorescence decay of HCR in the absence and presence of Pd in the
medium already mentioned; (b) corresponding residual plot (λ =450 nm).
ex
2
+
To further explore the selectivity of HCR for Pd , competition experiments are performed in the
2
+
presence of Pd mixed with 2.0 equiv. of each of the guest cations. Fig. 7 shows that these free
cations would have no such influence towards HCR and do not hamper the fluorogenic detection
2
+
2+
of Pd . Thus, coexisting metal ions do not significantly influence the recognition of Pd by
HCR.

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5
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
-
2+ +
Fig.6 Variation of normalized fluorescence intensity of (HCR) and [CR -Pd ] in EtOH/ H O
2
(8:2, v/v) (ex  545 nm , em  598 nm) with change in pH.
2+
Fig. 7 Fluorescence intensity change of 20 µM HCR in EtOH/ H O (8:2, v/v) by Pd ion at pH
2
7
.2 in presence of competing metal ions.

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6
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
3
.4 Proposed binding mode
Job’s method of continuous variation has yielded, from the absorption spectral data, a 1:1 (HCR:
2
+
Pd ) stoichiometry for the complex (ESI†, Fig. S6). The association constant (K ) of the sensor
a
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
with Pd2+ ion has been found to be 9.1×10 M from Benesi–Hildebrand equation, using
4
-1
emission titration data (ESI†, Fig. S7). In FTIR spectrum of the sensor a broad peak at 3444 cm-1
-
2+ +
for –OH (HCR) disappears in the complex [CR -Pd ] (ESI†, Fig. S8) which refers to the
binding of ligand to Pd(II) through oxygen of –OH via deprotonation. In the ESI-MS experiment,
2
+
free HCR shows a signal at m/z = 671.16 [M+H]+; after completion of the reaction of Pd with
HCR, ESI-MS spectrum showed a clear base peak at 775.1075 [M]+ (ESI†, Fig. S9); and which
-
2+ +
certainly corresponds to the formation of the complex, [CR -Pd ] . All these results suggest that
the coordination cavity generated by coumarin-rhodamine dyad with hydroxy-O and imine-N
2
+
-
2+ +
chelator causes the binding of Pd resulting in the four-coordinated [CR -Pd ] complex by
inducing opening of the spirolactam ring (Scheme 1). It is reported45-48 that the opening of
spirolactam ring of rhodamine moiety enhances the emission intensity. This is happened due to
the increase in conjugation in the ring-open form than in the ring-closed form which decreases
the HOMO-LUMO energy gap from the free sensor to its Pd(II) complex and thereby the
proposed mechanism will be as shown in Scheme 1.
3
.5 Theoretical studies
-
2+ +
The geometry generated by DFT for [CR -Pd ] shows distorted square planar about
Pd2+ ion where HCR acts as O,N,N,O chelator. The calculated Pd–O (rhodamine), Pd–N
(
imine), Pd–O (Coumarinyl) and Pd–N (rhodamine) distances are 2.48, 1.97, 2.20 and 2.48 Å
respectively and are comparable with similar reported structure. 49,50 Upon coordination the
shortening of C–O (coumarinyl) distance, 1.38 to 1.32 Å, and elongation of the distances C–O

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7
1
2
3
4
5
6
7
8
9
(
spirolactam) 1.25 to 1.30 Å and imine C=N, 1.27 to 1.30 Å have been calculated. Upon
2
+
chelation of Pd ion with the HCR, the energy of HOMO and LUMO are decreased relative to
those of free HCR.( ESI†, Fig. S1, S2). The HOMO–LUMO energy gap in HCR is
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
-
2+ +
2
.91 eV ( 426 nm) while that in the complex [CR -Pd ] is 2.32 eV ( 535 nm). These are
well in agreement with the experimentally observed longest wavelength absorption bands of
-
2+ +
HCR and [CR -Pd ] respectively (Fig . 2). This supports the fact that the appearance of the
2+
intense 565 nm absorption band upon addition of Pd is due to the formation of an entirely new
-
2+ +
species, the [CR -Pd ] complex.
2.91eV
2.32eV
-
2+ +
Fig. 8: HOMO–LUMO transitions of HCR and [CR -Pd ] complex.
.6 Intracellular imaging of Pd2+
3

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8
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
2+
Above experiments establish the unique selectivity and extreme sensitivity of HCR to Pd with
very low detection limit (18.8 nM), we were interested to check the potential utility of the sensor
in living cells. Therefore, we have first checked the toxicity of the ligand in MCF-7 (breast
cancer cell line) upto 200 μM. Sensor shows toxicity after 25 μM (Fig. 9) and can be used as an
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
+
anti-cancer agent. Following fluorescence microscopic technique the detection of Pd at very
low concentration has been examined in living cells. Palladium (0, II, IV) is a useful catalyst in
organic synthesis and the final product may contain traces of Pd(II) even after thorough repeated
purification. Such contamination of Pd is known to cause irritation of skin, eye, respiratory tract;
5
1
Pd(II) complexes are reportedly toxic and carcinogenic. For Pd, with a lethal dose of 5 – 10
ppm 51-53, the WHO recommendation for maximum uptake is restricted to ~1.5 – 15 μg/day.
Thus development of highly specific Pd2+ detection technique in live cells is an important
requirement of current research. In the present work, the cells were incubated with 5.0 μM of
o
HCR for 3h at 37 C and then treated with PdCl (5 μM) for 30 min at the same temperature,
2
which resulted in red fluorescence (Fig 10). The red emission indicates that HCR is easily
entered into cell membrane and capable to image Pd2+ in living cells. It may, therefore, be
2
+
concluded that in both in vitro and in vivo sensing and in whole cell imaging of Pd , the HCR is
a potential analytical reagent. Moreover, a control experiment has also been performed to check
whether the spiro ring remains intact in the cell culture medium or not. The cells were treated
with HCR (5M) for 24 h but did not get any sign of red coloration from the cells. This result
suggests that even in the complicated biological medium, the spiro ring of HCR does not open
(ESI†, Fig. S10).

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9
1
2
3
4
5
6
7
8
9
120
00
1
80
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
60
4
0
0
0
2
Control 6.25 12.5
25
50
100
200
Concentraton (M)
Fig. 9: Cell viability test by MTT assay by adding HCR to the media.
2
+
Fig. 10: Image of MCF7 Cells after the treatment with compound with Pd .
4. Conclusion
The synthesized coumarinyl-rhodamine dyad (HCR) has served as highly efficient fluorescent
2
+
sensor ( , 598 nm) for quantitative detection of Pd in semi-aqueous solution in presence of
em
many other metal ions with a very low detection limit of 18.8 nM. The selectivity has been
demonstrated by fluorescence, absorption and mass spectroscopy. The fluorescence intensity of

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0
1
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3
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6
7
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9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
the sensor has been enhanced 270 times by adding Pd2+ to it via opening of ring of the
spirolactam form of rhodamine moiety. This sensor acts as a dual probe for detecting visually
2
+
2+
Pd ions via the change in fluorescence and color. Due to high selectivity of HCR to Pd in
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
physiological medium at intracellular level, it can be used for identification of breast cancer cell.
Acknowledgements
Financial support from the Council of Scientific and Industrial Research (CSIR, Sanction no.
0
1(2814)/17/EMR-II,), New Delhi, India is gratefully acknowledged. Dr. Madhumita Manna,
Principle, Bidhannagar College has acknowledged for her enormous moral support and
encouragement. One of the authors (R.P.) is thankful to Department of Science and Technology
(DST), Govt. of India for providing DST-INSPIRE research fellowship.
References:
1
(a) L. Pu, Chem. Rev., 2004, 104, 1687-1716. (b) V. Amendola, L. Fabbrizzi, F. Forti, M.
Licchelli, C. Mangano, P. Pallavicini, A. Poggi, D. Sacchi and A. Taglieti, Coord. Chem. Rev.,
2
2
006, 205, 273-299. (c) E. M. Nolan and S. J. Lippard, Chem. Rev., 2008, 108, 3443-3480.
(a) J. F. Zhang, Y. Zhou, J. Yoon and J. S. Kim, Chem. Soc. Rev., 2011, 40, 3416-3429.
(
b) Z. Xu, X. Chen, H. N. Kim and J. Yoon, Chem. Soc. Rev., 2010, 39, 127-137
(
c) R. Balamurugan, C.C. Chien, K.M. Wu, Y.-H. Chiu and Jui-Hsiang Liu, Analyst, 2013, 138
1
3
564-1569.
(a) H. Kim, K. S. Moon, S. Shim and J. Tae, Chem. Asian J. 2011, 6, 1987-1991.(b) H. Li, J.
Fan, J. Du, K. Guo, S. Sun, S. Liu and X. Peng, Chem. Commun., 2010, 46, 1079-1081.
4
1
(a)H. Li, J. Fan, F. Song, H. Zhu, J. Du, S.Sun and X. Peng, Chem.–Eur. J., 2010, 16, 12349-
2356. (b) D. Keum, S. Kim and Y. Kim, Chem. Commun., 2014, 50, 1268-1270.

Page 21 of 26
New Journal of Chemistry
View Article Online
DOI: 10.1039/C8NJ06511J
2
1
1
2
3
4
5
6
7
8
9
5
(a) M. Ware, Platinum Metals Review, 2005, 49, 190-195. (b) T. Iwasawa, M. Tokunaga, Y.
Obora and Y. Tsuji, J. Am. Chem. Soc., 2004, 126, 6554-6555.(c) K. Ko¨ hler, W. Kleist and S.
S. Pro¨ cki, Inorg. Chem., 2007, 46, 1876-1879.(d) C.M.Crudden, M. Sateesh and R. Lewis, J.
Am. Chem. Soc., 2005, 127, 10045-10050.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
6
2
5
7
2
(a) S. L. Buchwald, C. Mauger, G. Mignani and U. Scholzc, Adv. Synth. Catal., 2006, 348,
3-29. (b) X. Chen, K. M. Engle, D. H. Wang and J. Q. Yu, Angew. Chem., 2009, 212, 5196-
217.
(a) J. Kielhorn, C. Melber, D. Keller and I. Mangelsdorf, Int. J. Hyg. Environ. Health, 2002,
05, 417-432.(b) C. D Spicer, T. Triemer and B. G. Davis, J. Am. Chem. Soc., 2012, 134, 800-
803.
8
T.Gebel, H. Lantzsch, K. Plebow and H. Dunkelberg, Mutat. Res. Genet. Toxicol. Environ.
Mutagen, 1997, 389, 183.
9 T. Z. Liu, S. D. Lee and R. S. Bhatnagar, Toxicol. Lett., 1979, 4, 469-473.
1
4
1
1
0 J. Kielhorn, C. Melber, D. Keller and I. Mangelsdorf, Int. J. Hyg. Environ. Health, 2002, 205,
17-432.
1 B. Dimitrova, K. Benkhedda, E. Ivanova and F. Adams, J. Anal. At. Spectrom., 2004, 19,
394-1396.
1
2 C. Locatelli, D. Melucci and G. Torsi, Anal. Bioanal. Chem., 2005, 382, 1567-1573..
1
3 K. Van Meel, A. Smekens, M. Behets, P. Kazandjian and R. Van Grieken, Anal. Chem.,
2007, 79, 6383-6389.
4 (a) H. Li, J. Fan and X. Peng, Chem. Soc. Rev., 2013, 42, 7943-7962. (b) J. Du, M. Hu, J.
Fan and X. Peng, Chem. Soc. Rev., 2012, 41, 4511-4535.
1

New Journal of Chemistry
Page 22 of 26
View Article Online
DOI: 10.1039/C8NJ06511J
2
2
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
5 (a) L. Duan, Y. Xu and X. Qian, Chem. Commun., 2008, 0, 6339-6341. (b) J. R. Matthews,
F. Goldoni, H. Kooijman, A. L. Spek, A. P. H. J. Schenning and E. W. Meijer, Macroomol Rapid
Commun., 2007, 28, 1809-1815.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
6 (a) A. K. Mahapatra, S. K.Manna, K. Maiti, S.Mondal, R. Maji, D. Mandal, S.Mandal, M. R.
Uddin, S. Goswami, C.K. Quah and H. Fun, Analyst , 2015, 140, 1229-1236. (b) S. Goswami, D.
Sen, N.K. Das, H. Fun and C. K. Quah, Chem. Commun., 2011, 47, 9101–9103.
1
3
1
7 G. Wei, L. Wang, J. Jiao, J. Hou, Y. Cheng and C. Zhu, Tetrahedron Lett., 2012, 53, 3459–
462.
8. (a) Y. K. Yang, Yook, K. J.; Tae, J. J. Am. Chem. Soc., 2005, 127, 16760-16761. (b) J. Y.
Kwon, Y. J. Jang, Y. J. Lee, K. M. Kim, M. S. Seo, W. Nam, J. Yoon, J. Am. Chem.
Soc., 2005, 127, 10107-10111. (c) V. Dujols, F. Ford, A. W. Czarnik, J. Am. Chem.
Soc.,1997, 119, 7386-7387. (d) M. Wang, X. Liu, H. Lu, H. Wang, and Z. Qin, ACS Appl.
Mater. Interfaces 2015, 7, 1284–1289. (e) R. Balamurugan, J. –H. Liu, B. –T. Liu, Coord. Chem.
Rev., 2018, 376, 196-224. (f) N. Kumari, N. Dey, K. Kumar and S. Bhattacharya, Chem. Asian J.
2
1
014, 9, 3174 – 3181. (g) H. Li, J. Cao, H. Zhu, J. Fan, X. Peng, Tet. Lett., 2013, 54, 4357–4361
9 A. K. Bhanja, S. Mishra, K. Das Saha and C. Sinha, Dalton Trans., 2017, 46, 9245–9252.
20. A. K. Bhanja, S. Mishra, K. Kar, K. Naskar, S. Maity, K. Das Saha and C. Sinha, New J.
Chem., 2018, 42, 17351-17358.
2
1 Jacob Q. Umberger and Victor K. LaMer, J. Am. Chem. Soc., 1945, 67, 1099–1109.
22 K. Das, U. Panda, A. Datta, S. Roy, S. Mondal, C. Massera, T. Askun, P. Celikboyun, E.
Garribba, C. Sinha, K. Anand, T. Akitsu and K. Kobayashi, New J. Chem., 2015, 39,7309-7321.
2
3 J.-S. Wu, I.-C. Hwang, K. S. Kim and J. S. Kim, Org. Lett., 2007, 9, 907–910.

Page 23 of 26
New Journal of Chemistry
View Article Online
DOI: 10.1039/C8NJ06511J
2
3
1
2
3
4
5
6
7
8
9
2
4 M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria, M. A. Robb, J. R. Cheeseman, G.
Scalmani, V. Barone, B. Mennucci, G. A. Petersson, H. Nakatsuji, M. Caricato, X. Li, H. P.
Hratchian, A. F. Izmaylov, J. Bloino, G. Zheng, J. L. Sonnenberg, M. Hada, M. Ehara, K.
Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T.
Vreven, J. A. Montgomery Jr., J. E. Peralta, F. Ogliaro, M. Bearpark, J. J. Heyd, E. Brothers, K.
N. Kudin, V. N. Staroverov, R. Kobayashi, J. Normand, K. Raghavachari, A. Rendell, J. C.
Burant, S. S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J. M. Millam, M. Klene,J. E. Knox, J.
B.Coss, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R. E. Stratmann, O. Yazyev, A. J.
Austin, R. Cammi, C. Pomelli, J. W. Ochterski, R. L. Martin, K. Morokuma, V. G. Zakrzewski,
G. A. Voth, P. Salvador, J. J. Dannenberg, S. Dapprich, A. D. Daniels, Ö. Farkas,J. B. Foresman,
J. V. Ortiz, J. Cioslowski and D. J. Fox, Gaussian 09, Revision D.01, Gaussian Inc., Wallingford,
CT, 2009.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
25 A. D. Becke, J. Chem. Phys., 1993, 98, 5648-5652.
2
2
2
2
3
3
6 C. Lee, W. Yang and R. G. Parr, Phys. Rev. B: Condens. Matter, 1988, 37, 785–789.
7 P. J. Hay and W. R. Wadt, J. Chem. Phys., 1985, 82, 270–283.
8 W. R. Wadt and P. J. Hay, J. Chem. Phys., 1985, 82, 284–298.
9 P. J. Hay and W. R. Wadt, J. Chem. Phys., 1985, 82, 299–305.
0 V. Barone and M. Cossi, J. Phys. Chem. A, 1998, 202, 1995– 2001.
1 M. Cossi and V. Barone, J. Chem. Phys., 2001, 115, 4708–4717.
32 M. Cossi, N. Rega, G. Scalmani and V. Barone, J. Comput. Chem., 2003,24, 669-681.
3
3
3 N. M. O’Boyle, A. L. Tenderholt and K. M. Langner, J. Comput. Chem., 2008, 29, 839–845.
4 A. K. Mahapatra, S. K. Manna, K. Maiti, S. Mondal, R. Maji, D. Mandal, S. Mandal, Md. R.
Uddin, S. Goswami, C. K. Quah and H. K. Fun, Analyst, 2015, 140, 1229–1236.

New Journal of Chemistry
Page 24 of 26
View Article Online
DOI: 10.1039/C8NJ06511J
2
4
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
3
3
5 F. Liu, J. Du, M. Xu and G. Sun, Chem. Asian J., 2016, 11, 43-48.
6 Q. Huang, Y. Zhou, Q. Zhang, E. Wang, Y. Min, H. Qiao, J. Zhang and T. Ma, Sens.
Actuators B:, 2015, 208, 22–29.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
3
3
3
1
4
7 Y. Chen, B. Chen and Y. Han, Sens. Actuators B:, 2016, 237, 1–7.
8 V. Sharma, A. K. Sainib and S. M. Mobin, J. Mater. Chem. B, 2016, 4, 2466–2476.
9 J. Panchompoo, L. Aldous, M. Baker, M. I. Wallace and R. G. Compton, Analyst, 137 (2012,
37, 2054–2062.
0 W. Liu, J. Jiang, C. Chen, X. Tang, J. Shi, P. Zhang, K. Zhang, Z. Li, W. Dou, L. Yang and
W. Liu, Inorg. Chem., 2014, 53, 12590–12594.
41 F. Song, A. L. Garner and K. Koide, J. Am. Chem. Soc., 2007, 129, 12354–12355.
4
4
2 D. Keum, S. Kim and Y. Kim, Chem. Commun., 2014, 50, 1268–1270.
3 P. Kumar, V. Kumar and R. Gupta, RSC Adv., 2017, 7, 7734–7741.
44 W. X. Ren, T. Pradhan, Z. Yang, Q. Y. Cao and J. S. Kim, Sens. Actuators B:, 2012, 171,
1
4
4
9
277–1282.
5 S. Sun, B. Qiao, N. Jiang, J. Wang, S. Zhang and X. Peng, Org. Lett., 2014, 16, 1132–1135.
6 S. Goswami, D. Sen, N. Kumar Das, H. K. Fun and C. K. Quahb, Chem. Commun., 2011,47,
101-9103.
4
7 M. A. Bennett, S. K. Bhargava, M. A. Keniry, S. H. Prive´r, P. M. Simmonds, J. Wagler and
A. C. Willis, Organometallics, 2008, 27, 5361-5370.
48 U. Reddy G., F. Ali, N. Taye, S. Chattopadhyay and A. Das, Chem. Commun., 2015, 51,
3
649-3652.

Page 25 of 26
New Journal of Chemistry
View Article Online
DOI: 10.1039/C8NJ06511J
2
5
1
2
3
4
5
6
7
8
9
4
9 B. Liu, H. Wang, T. Wang, Y. Bao, F. Du, J. Tian, Q. Li and R. Bai, Chem. Commun., 2012,
4
8, 2867–2869.
50 X. Wang, Z. Guo, S. Zhu, H. Tian and W. Zhu, Chem. Commun., 2014, 50, 13525–13528.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
5
5
5
1 C. E. Garrett, K. Prasad, Adv. Synth. Catal., 2004, 346, 889–900.
2 D. G. Cho and J. L. Sessler, Chem. Soc. Rev., 2009, 38, 1647–1662.
3 F. Song, A.L. Garner and K. Koide, J. Am. Chem. Soc., 2007, 129, 12354–12355.

New Journal of Chemistry
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Pictorial abstract
2
+
Fluorescence sensing and intracellular imaging of Pd ion by a novel
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coumarinyl-rhodamine Schiff base†
a
b
c
d
c
Arup Kumar Adak, Rakesh Purkait , Saikat Manna, Bankim Chandra Ghosh, Sudipta Pathak
b
and Chittaranjan Sinha
Pd2+
pH 7.2
λex=545 nm
λem=598 nm