Coumarin based colorimetric and fluorescence on-off chemosensor for F?, CN? and Cu2 + ions

Article abstract of DOI:10.1016/j.saa.2017.09.071

(E)-4-Chloro-3-[{2-(4-nitrophenyl)hydrazono}methyl]-2H-chromen-2-one (C), a coumarin derivative has been studied toward its ion sensing properties for F^?, CN^? and Cu^{2 +}. A proton-transfer mechanism for F^? sensing has been deduced with the help of ¹H NMR titration alongwith from the changes in the absorption and emission spectra of C in the presence of F^?. C formed 1:1 stoichiometric complex with each of these analytes. Sensing of C toward Cu^{2 +} is poor, but interestingly in the presence of F^? or CN^? the sensing ability of Cu^{2 +} gets enhanced many folds, and C can act as F^? or CN^? mediated off-on sensor for Cu^{2 +}. Moreover, colorimetric strip (pre-coated with the coumarin derived compound) tests for F^? and CN^? from their DMSO solution at high temperature (~ 100 °C) opens up the door for easiest naked eye recognition and distinction of these ions, and also for naked-eye detection of F^? and CN^? from its aqueous solution at high temperature (~ 100 °C).

Full text of DOI:10.1016/j.saa.2017.09.071

Accepted Manuscript
Coumarin based colorimetric and fluorescence on-off
chemosensor for F−, CN− and Cu2+ ions
Debashis Roy, Arijit Chakraborty, Rina Ghosh
PII:
DOI:
Reference:
S1386-1425(17)30798-9
doi:10.1016/j.saa.2017.09.071
SAA 15505
Spectrochimica Acta Part A: Molecular and Biomolecular
Spectroscopy
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Received date: 17 June 2017
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26 September 2017
Please cite this article as: Debashis Roy, Arijit Chakraborty, Rina Ghosh , Coumarin
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Coumarin based colorimetric and fluorescence on-off chemosensor
for F, CN and Cu2+ ions
Debashis Roya, Arijit Chakrabortyb,*, Rina Ghosha,*
a Department of Chemistry, Jadavpur University, Kolkata 700032, India.
b Department of Chemistry, Acharya B. N. Seal College, Cooch Behar, West Bengal 736101, India.
*E-mails of the Corresponding authors: arijit_chak2002@yahoo.com (A. Chakraborty),
ghoshrina@yahoo.com (R. Ghosh)
ABSTRACT
(E)-4-Chloro-3-[{2-(4-nitrophenyl)hydrazono}methyl]-2H-chromen-2-one (C),
a
coumarin
derivative has been studied toward its ion sensing properties for F, CN and Cu2+. A proton-
transfer mechanism for F sensing has been deduced with the help of 1H NMR titration alongwith
from the changes in the absorption and emission spectra of C in the presence of F. C formed 1:1
stoichiometric complex with each of these analytes. Sensing of C toward Cu2+ is poor, but
interestingly in the presence of For CN the sensing ability of Cu2+ gets enhanced many folds,
and C can act as For CN mediated off-on sensor for Cu2+. Moreover, colorimetric strip (pre-
coated with the coumarin derived compound) tests for Fand CN from their DMSO solution at
high temperature (~100 C) opens up the door for easiest naked eye recognition and distinction of
these ions, and also for naked-eye detection of F and CN from its aqueous solution at high
temperature (~100 C).

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Key words: Chemosensor; Coumarin derived sensor; Fluoride sensing; Cyanide sensing; Copper
(II) sensing; Aqueous fluoride estimation.
1. Introduction
Recently owing to their immense impact in biological, medicinal, environmental and
industrial fields studies on chemical sensors have gained much research interest [1-2]. Ions,
being crucial to physiological functions, play vital role in our daily life. Fluoride ion is used
in the treatment of dental cavity and also for osteoporosis, but in high doses or persistent
ingestion in lower doses of fluoride may cause fluorosis with over accumulation of F in
bone [3], and also may lead to nephrotoxic changes as well as urolithiasis in human [4].
Cyanide ion is used, in chemical industry for resins, fibers, herbicides and in gold extraction
process [5]. Cyanide ion is a fast acting poison that can be lethal to human body [6-9]. The
World Health Organization (WHO) has set an upper limit of 1.9 µM for cyanide in drinking
water [9-10]. Cyanide inhibits the process of cellular respiration in mammals by interacting
very strongly with a heme unit in the active site of cytochrome a3 [11]. Cu2+ ion plays vital
roles in many fundamental physiological processes in organisms. Various Cu2+ dependent
enzymes act as catalysts to provide energy for biochemical reactions, e.g. transformation
of melanin for pigmentation of the skin and assistance of the formation of crosslinks in
collagen and elastin [12]. Unregulated overloading of Cu2+ can induce severe
neurodegenerative diseases including Alzheimer’s, Parkinson’s and prion diseases [13].
Spectro-photometric sensing of F is based on interaction with Lewis acids or
through utilizing H-bonding interaction or by reaction with F or based on interaction with
mesoporous silica [14]. So far only a few coumarin-derived F sensors have been reported

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[3, 4, 15-17]. A variety of CN sensing reagents have been there in the literature; of these
whereas mostly provide an indirect way for cyanide detection, a few are based on H-bond
mediated sensing [18, 19], others through utilization of cyanide complexes [11, 20-24] and
some exploit nucleophilic reaction with CN [8, 11]. A few of the reported CN sensors
contain coumarin skeleton [5, 9, 11, 25, 26]. Among the metal ions, Cu2+ is one of the
mostly researched metal sensing ions [2, 13, 25-29]; a few of these have used coumarin
moiety [13, 25, 26, 28, 29].
Inspite of the literature reports on F/ CN/ Cu2+ ion sensors; there is still scope for
development of simple as well as stable chemical sensor that can be used for detection of
these analytes spectrophotometrically as well as by naked–eye in low level without
requiring any instrument, with an emphasis particularly on the estimation of aqueous F.
Herein, we are reporting our recent studies on coumarin derived hydrazone as a
chemosensor of F, CN and Cu2+ ions.
2. Experimental
2.1. General Information
All the solvents and reagents used were of analytical grade and spectroscopic grade.
1H NMR and 13C NMR measurements were performed on a Bruker DPX-300 (300 and 75
MHz, respectively) spectrometer, and chemical shifts were recorded in ppm. FT-IR data
were taken from PerkinElmer Spectrum Version 10.03.09. A High-resolution mass
spectrum was obtained on a Waters Micromass Q-tof Micro mass spectrometer using
electrospray ionization method. Melting points were determined on a LabX India digital
melting point apparatus. UV-vis spectra were recorded at room temperature using a

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PerkinElmer UV Lambda 25 spectrometer, and fluorescence data were collected from
PerkinElmer fluorescence Spectrometer LS 55.
2.2. Synthesis of 4-chloro-3-{(2-(4-nitrophenyl)hydrazono)methyl}-2H-chromen-2-one (C)
4-Chloro-3-formylcoumarin [30] (1.043 g, 5 mmol) was dissolved in absolute
ethanol (22 mL). p-Nitro phenylhydrazine (0.736 g, 5 mmol) was dissolved in this solution,
and the resulting mixture was stirred vigorously for an hour at room temperature. An orange
coloured precipitate separated from the mixture. The reaction mixture was then filtered off,
and the precipitate was recrystallised from N,N-DMF. Yield 1.598 g (93%). M.P. = 250 °C.
1H NMR (DMSO-d6, 300MHz): δ (ppm) = 7.17 (2H, d, J = 8.52, Ar); 7.45-7.49
(2H, m, Ar); 7.66-7.71 (1H, t, J=7.67, Ar); 8.00 (1H, d, J = 7.94, Ar) 8.15 (2H, m, J = 9.14,
Ar); 8.22 (1H, s, =CH–); 11.69 (1H, br. s, NH). 13C-NMR (DMSO-d6, 75 MHz): δ (ppm)
= 112.5, 116.9, 119.1, 119.5, 125.8, 126.2, 126.6, 133.6, 135.3, 139.9, 142.7, 150.3, 151.2
and 158.4. IR spectrum (cm-1, KBr pellet): 753, 840, 1328, 1568, 1603, 1702, 3271. HRMS:
calcd for molecular formula C16H10ClN3O4N’: 366.0258 [M + Na] +; found 366.0260.
2.3. UV-vis Titration of C with F
C
(3.6 mg, 10.5 μmol) was dissolved in dimethylsulfoxide (DMSO, 1 mL), and 2
(10.5 mM) was diluted to 2 mL acetonitrile. Tetrabutylammonium fluoride
μL of the
C
(TBAF) (2.3 mg, 7.3 μmol) was dissolved in acetonitrile (1mL). 1μL of the TBAF solution
(7.3 mM) was added each time to the solution. After mixing these homogeneously, UV-
C
vis absorption spectra were run at room temperature. This process was continued for several
times upto saturation.

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2.4. UV-vis Titration of C with CN
(3.5 mg, 10.1 μmol) was dissolved in DMSO (1 mL), and 2 μL of the
C
C (10.1
mM) was diluted to 2 mL acetonitrile. KCN (4 mg, 61.2 μmol) was dissolved in 1 ml water:
acetonitrile (1: 9) mixture. 1μL of the KCN solution (61.2 mM) was added each time to the
C
solution. After mixing these homogeneously, UV-vis absorption spectra were run at room
temperature. This process was run for several times upto saturation.
2.5. UV-vis Titration of C with Cu2+
C
(2.1 mg, 6.2 μmol) was dissolved in DMSO (1 mL), and 2 μL of the C (6.2 mM)
was diluted to 2 mL acetonitrile. Cu(NO3)2. 3H2O (4 mg, 16.5 μmol) was dissolved in
acetonitrile (1 mL), and 1 μL of the Cu2+ solution (16.5 mM) was added each time to the
C
solution. After mixing homogeneously, UV-vis absorption spectra were recorded at room
temperature. This process was repeated for several times upto saturation.
2.6. Fluorescence Spectroscopic Titration of C with F
C
(3.6 mg, 10.5 μmol) was dissolved in DMSO (2 mL), and 2 μL of the
mM) was diluted to 2 mL acetonitrile. TBAF (4.6 mg, 14.6 μmol) was dissolved in
acetonitrile (1 mL). 1 μL of the TBAF solution (14.6 mM) was added each time to the
solution. After mixing these homogeneously, fluorescence spectra were taken at room
C (10.5
C
temperature by exciting
saturation.
C at 425 nm. This process was continued for several times upto

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2.7. Fluorescence Spectroscopic Titration of C with CN
(5 mM) was prepared in DMSO, and 3.7 μL of the
acetonitrile. KCN (5 mM) solution was prepared in water: acetonitrile mixture (1: 9). 2 μL
of the KCN solution was added each time to the solution. After mixing homogeneously,
C
C (5 mM) was diluted to 2 mL
C
fluorescence spectra were recorded at room temperature by exciting
process was run for several times upto saturation.
C at 425 nm. This
2.8. Fluorescence Spectroscopic Titration of C with Cu2+
C
(3.6 mg, 10.5 μmol) was dissolved in DMSO (2 mL), and 2 μL of the C (10.5
mM) was diluted to 2 mL acetonitrile. Cu(NO3)2. 3H2O (6.8 mg, 28.1 μmol) was dissolved
in acetonitrile (1 mL), and 8 μL of the Cu2+ solution (28.1 mM) was added each time to
the
C solution. After mixing these homogeneously, fluorescence spectra were run at room
temperature by exciting
saturation.
C at 425 nm. This process was repeated for several times upto
2.9. Job Plot Measurements of C with F, CN and Cu2+
(5 mM) solution was prepared in DMSO, and the analyte ion solution (5 mM)
(tetrabutyl ammonium fluoride, KCN, and Copper nitrate trihydrate) was prepared in
acetonitrile. Then 1, 2, 3, 4, 5, 6, 7, 8 and 9 μL of the solution (5 mM) were taken and
transferred to vials. 9, 8, 7, 6, 5, 4, 3, 2 and 1 μL of the analyte ion solution (5 mM) was
added to each solution so that the mole fraction of became 0.1, 0.2, 0.3, 0.4, 0.5, 0.6,
0.7, 0.8 and 0.9, respectively. Each vial was diluted with acetonitrile to make a total volume
of 2 mL. After shaking the vials for a few seconds to form -analyte ion complex, UV-vis
C
C
C
C
C

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absorption spectra were taken at 25°C. Similar process was run for blank test with
solution only.
C
2.10. Colorimetric Strip Test toward F and CN
C
(1.4 mg, 4 μmol) was dissolved in DMSO (2 mL).
C
coated strips were prepared
by immersing filter papers into
C solution (2 mM) and then dried in an oven. The test strips
prepared above were dipped into different anion solutions (8 mM). Only F (DMSO
solution) and CN (in DMSO: water 4: 1) could change the color of the strips from yellow
to bluish green. For detection of aqueous fluoride and cyanide ion by naked-eye, the strip
(
C coated paper strips dipped into the aqueous fluoride or cyanide solution) must be kept
into an oven at ~100°C.
2.11. EDTA Reversibility Test for C
2.69 μL 5 mM solution in DMSO of
C
was diluted with 2 ml acetonitrile. Then 2.69
. After mixing the solution, UV-
μL 5 mM Cu2+ ion solution was added to this solution of
C
vis spectrum was taken at room temperature. 5 mM disodium dihydrated salt of EDTA was
dissolved in 1: 1 = water: acetonitrile solution, and 2.69 μL of the EDTA solution was
added to the solution of (
C
+ Cu2+) complex (5 mM) as prepared above. After mixing
homogeneously UV-vis spectrum was run. For the reversibility study, another 2.69 μL of
the Cu2+ ion solution (5 mM) was mixed to the above solution. After mixing it, UV-vis
spectrum was taken at room temperature. The same procedure was repeated for 3 times
more in the same system.

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2.12. pH Dependency of C
A series of buffer solutions with pH values ranging from 1 to 10 were prepared
according to G. Gomori’s method by using pH-meter where pH 1 and pH 2 were of KCl-
HCl buffer (by mixing 0.2 M KCl solution and 0.2 M HCl solution), pH 3 to pH 7 were of
Citrate-Phosphate buffer (by mixing 0.1 M citric acid solution and 0.2 M disodium
hydrogenphosphate solution), pH 8 to pH 9 were of Borax-Boric acid buffer (by mixing
0.05 M Borax and 0.2 M Boric acid solution), and pH 10 solution was made of carbonate-
bicarbonate buffer (by mixing 0.2 M sodium carbonate solution and 0.2 M Sodium
bicarbonate solution). After making the desired pH, 2 μL 5 mM solution of
C (prepared in
DMSO) was added to 2 ml DMSO/ buffer (v/v, 1:1) mixture of each pH. After mixing
homogeneously, fluorescence spectra (excited at 436 nm) were run for all pH ranging from
1 to 10 at room temperature.
For UV-vis and colour response experiments of
C in a series of buffers with pH
values ranging from 1 to 10, 10 μL 5 mM solution of (prepared in DMSO) was added to
C
2 ml DMSO/ buffer (v/v, 1:1) mixture of each pH. After mixing homogeneously, absorption
spectra were run for all pH ranging from 1 to 10 at room temperature.
2.13. Preparation of Commercial Toothpaste Solution
0.4 to 0.5 g of toothpaste was weighed into a 2 ml eppendrof, homogenised by
stirring in 1.0 mL of deionised water. The suspension was centrifuged (5000 r.p.m, 10 min,
room temperature) to remove the abrasive. The supernatant liquid was taken to determine
the total soluble fluoride in the toothpaste sample.

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3. Results and Discussion
3.1. Synthesis and Characterisation
The chemosensor was synthesised from the 4-hydroxy coumarin (
literature procedure [30] with some modification (Scheme 1); a much better yield of
obtained by this modification. In the first step 4-chloro-2-oxo-2H-chromene-3-
carbaldehyde ( ) was synthesised from by using Vilsmeier–Haack reaction. On coupling
of C
C
A
) following
was
C
B
A
B
with p-nitrophenylhydrazine in absolute ethanol at room temperature compound
was characterised by 1H NMR, 13C NMR, FT-
was prepared with 96% yield. Compound
C
IR and high resolution mass spectrometry.
Scheme 1. Synthesis of Chemosensor C.
3.2. Colorimetric and Spectral Studies of Chemosensor C toward Analytes
3.2.1. Anion Sensing
The colorimetric sensing abilities of
C were investigated in acetonitrile solution (6.0 μM,
흀max = 425 nm, molar extinction co-efficient (ε) = 4.6x104 M-1cm-1) upon the treatment with
3


various kinds of anions such as C6H5COO, CH3COO, AsO2 , CN, BO3 , F, Cl, Br,
2





I, SCN
,
H2PO4 , HPO4 , HSO4 , NO2 and NO3 (Fig. 1). Anions F, Cl, Br, I and
CH3COO were used as their tetrabutyl ammonium salts dissolved in acetonitrile; CN was


used as its potassium salt in water-acetonitrile mixture (1:9); NO2 and NO3 ions were used

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2


as their sodium salts in acetonitrile, and C6H5COO, AsO2 , SCN, H2PO4 , HPO4  and

HSO4 ions were used as their sodium salts in 1:1 water-acetonitrile [26].
Fig. 1. (a) The color changes of C (50 μM) upon addition of various anions (1 equiv.) in MeCN.
3


A= Free C, B = C+ C6H5COO, C = C+ CH3COO, D = C+ AsO2 , E = C+CN, F = C+ BO3 ,
2


G = C+F, H = C+ Cl, I = C+ Br, J = C+ I, K = C+ SCN , L = C+ H2PO4 , M = C+ HPO4
,



N = C+ HSO4 , O = C+ NO2 , P = C+ NO3 . (b) Absorption changes of C (6.0 μM) upon addition
of various anions (4 equiv.) in MeCN.

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Fig. 2. (a) UV-vis spectral changes of C (10.5 μM) upon addition of F in MeCN at room
temperature. Inset: Absorbance at 641 nm as a function of fluoride ion concentration. (b) UV-vis
spectral changes of C (10.0 μM) upon addition of CN in MeCN at room temperature. Inset:
Absorbance-1 at 425 nm as a function of CN ion concentration.
Whereas upon the addition of F to the solution of C, a change of bright yellow to brilliant blue
color was observed, a bright yellow to greenish blue color change was witnessed through naked
eye on similar addition of CN ion, and other anions did not produce any visual change.
Subsequently a distinct change in the UV-vis absorption peak with a red-shift by Δ흀max of 216 nm
giving isosbestic points at 470 nm and 345 nm for F, and with a red-shift at Δ흀max of 213 nm
giving isosbestic points at 470 nm and 336 nm for CN, were noticed (Fig.1b and 2). Thus
compound C is no-doubt a potential candidate for “naked-eye” chemosensor of F and CN ions.
To obtain fluorescent properties of
C toward above mentioned anions, the emission
changes were investigated. Among the various anions studied
C selectively responded to
F and CN in acetonitrile medium. By gradual addition of F ion to the solution of
C
, a
dramatic fluorescent quenching was observed with quenching efficiency = 80% (Fig. 3)
[31], but a slight quenching (with quenching efficiency = 13%) was noticed for CN

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suggesting that
C
shows a specific fluorescence response to F ion. For collisional
quenching the decrease in intensity was described by the well-known Stern-Volmer
equation (eqn.1) [32]:
퐼
0 = 1 + KS-V [A]
(1)
퐼
In this equation I0 and I are the fluorescence intensities, respectively, in the absence
of and at the intermediate of the interaction of analyte ions; [A] is the concentration of
analyte ions; KS-V is the Stern-Volmer quenching constant with values 1.0 x 105 M-1 for F
and 8.1 x 104 M-1 for CN.
Fig. 3. (a) Fluorescent spectra of C (10.5 μM) upon addition of F (upto 4 equiv) in MeCN. Inset:
Intensity of C at 496 nm as a function of fluoride ion concentration. (b) Stern-Volmer plot for

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calculation of quenching constant of C for F ion. (c) Fluorescent spectra of C (9.2 μM) upon
addition of CN (upto 3.2 equiv.) in MeCN. Inset: Intensity of C at 496 nm as a function of CN
concentration. (d) Stern-Volmer plot for calculation of quenching constant of C for CN.
3.2.2. Cation Sensing
To examine the cation sensing property of
C [26] a solution of C (6.2μM ) was
further treated with various kinds of cations such as Na+, K+, Mg2+, Al3+, Ca2+, Cr3+, Mn2+,
Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Zr4+, Ag+, Cd2+, Ba2+ and Pb2+ (Fig. 4). Except MnSO4
and Fe2+ (Mohr’s salt) all cations were used as their nitrate salts.
Fig. 4. (a) Absorbance changes of C (6.2 μM) upon addition of various cations (20 equiv.) in
MeCN at room temperature. (b) UV-vis spectral changes of C (6.2 μM) upon addition of Cu2+
(checked upto 17 equiv.) in MeCN at room temperature. Inset: Absorbance at 425 nm as a function
of Cu2+ ion concentration.
Among the above mentioned cations, the change in UV-vis absorption spectra with
isosbestic points at 335 nm and 260 nm in case of Cu2+ (Fig. 4b) were witnessed, which
indicate the selectivity of
C
for Cu2+, and this was also ensured by the corresponding
fluorescence titration (Fig. 5), but there was no visible change with Cu2+. The association

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constant (Ka) determined from this absorption titration curve for Cu2+ ion (using eqn. 2,
vide section 3.2.3) was found to be Ka
=
4.5 x 102 M-1.
Fig. 5. (a) Fluorescent spectra of C (8.7 μM) upon addition of Cu2+ ion (checked upto 62 equiv.)
in MeCN. Inset: Intensity of C at 496 nm as a function of Cu2+ ion concentration. (b) Stern-Volmer
plot for calculation of quenching constant of C for Cu2+ ion.
3.2.3. Determination of Association Constant
The extent of the association of the chemosensor
C
to the ions F, CN and Cu2+ are
obtained from an examination of the absorbance data using the Benesi–Hildebrand relation
(eqn. 2) [33]. According to this relation
1
1
1
=
+
퐾
푎
[ ]
퐼 (퐴 −퐴 )
퐴−퐴
퐴 −퐴
0
푓
0
푓
0
(퐴 −퐴 ) = 1 +
1
푓
0
[ ]
퐾 퐼
푎
(퐴−퐴 )
0
log ( 퐴−퐴0 ) = log[I ] + log Ka
(2)
퐴 −퐴
푓
0
in which A0, A and Af are the absorbances, respectively, in the absence of, at the
intermediate and at the saturation of the interaction of analyte (F, CN and Cu2+), and [I]

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is the concentration of analyte added. The association constant (Ka) was determined by
linear fitting of absorption titration curve (Fig. 6).
Fig. 6. (a) Benesi-Hildebrand plots for calculation of association constant of C for F ion. (b) B-H
plots for calculation of association constant of C for CN ion. (c) B-H plots for calculation of
association constant of C for Cu2+.
3.2.4. Job Plot Analysis
The Job plot analysis (Fig. 7) revealed a 1:1 stoichiometric ratio between the
chemosensor
C
and the corresponding analyte ion (F , CN and Cu2+) when difference in
separately [26].
absorption maxima were plotted against mole fraction of chemosensor
C
Fig. 7. Job plots for the binding of C with F, CNand Cu2+, respectively. (a) (A-A0) at 641 nm
was plotted as a function of the molar ratio [C]/ ([C] + [F]). (b) (A-A0) at 638 nm was plotted as
a function of the molar ratio [C]/ ([C] + [CN]). (c) (A-A0) at 425 nm was plotted as a function of
the molar ratio [C]/ ([C] + [Cu2+]). Where A and A0 are the absorption maxima of chemosensor C
with analyte ions and without analyte ions, respectively.

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3.2.5. Detection Limits
Using UV-vis titration experiments of
the respective detection limits were calculated using the equation: detection limit = 3
where is the standard deviation of the blank solution and m is the slope of the
C
with F, CN and Cu2+ (Fig. 2a, 2b and 4b),
/m,


C
calibration curve [34], O.D. versus analyte ion concentration. The detection limits of
C
with F, CN and Cu2+ are 9.0 x 10-9 M, 1.5 x 10-8 M and 8.0 x 10-6 M, respectively. This
method allows a sensitive readout of chemosensor
C within a wide linear range of (5– 2105)
x 10-8 M at 흀max of 425 nm.
3.2.6. Quantum Yield Measurements
Quantum yield of chemosensor
C (Table 1) was estimated by a secondary method
[35] using fluorescence spectra of
C
in the absence and in the presence of F, CN and Cu2+
in acetonitrile medium utilising eqn. 3, where 푂퐷푠 and 푂퐷푟푒푓 are the absorbances, As and
Aref are integrated emission areas of chemosensor and standard, and ηS and ηref are the
C
refractive index of water and acetonitrile at 25ºC, respectively. Quinine sulfate dihydrate
(Quantum yield in 0.1 N H2SO4 is 0.54) has been used as a standard [36].
2
푂퐷
푟푒푓
. 퐴 . 휂
푠
푠
Φs =
Φ푟푒푓
(3)
2
푟푒푓
푂퐷 . 퐴
. 휂
푠
푟푒푓
Table 1. Quantum Yields of free C and C with analyte ions
Compound in solution
C in MeCN
Quantum Yield at 25ºC
0.0053
0.0062
0.0071
C + TBAF in MeCN
C + KCN in MeCN

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C + Cu2+ in MeCN
0.0019
3.2.7. pH Dependency of C and Measurement of pKa Value
We further investigated the effect of pH on the fluorescence and UV-vis response
(Figs. 8a, 8b and 8c) alongwith colour response (Fig. 8d) of
C in a series of buffers with
pH values ranging from 1 to 10 [37]. Although appreciable intensity change was not
witnessed from pH 1 to 7, but there was a sharp decrease in the intensity at 525 nm (Figs.
8a and 8b) and increase in the absorbance at 575 nm (inset, Fig. 8c) beyond pH 7; a similar
trend of colour response was also witnessed (Fig. 8d). In aqueous medium
absorption maximum at 436 nm, hence to get the intensity changes of , it was excited at
436 nm for all tested pH values. pKa of was 8.1 as determined from the plot of Intensity
at 525 nm (emission maxima) in the pH range 1 to 10, vs pH (Fig. 8b).
C shows
C
C

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Fig. 8. (a) Intensity of chemosensor C (10 μM) at various pH ranges (1 to 10) in buffer solutions
excited at 436 nm. (b) Determination of pKa value of C, (c) UV-vis response of C (24.9 μM) at
various pH ranges (1 to 10) in buffer solutions; Inset: Absorption changes of C at 575 nm as a

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function of pH. (d) Colour response of C (24.9 μM) at various pH ranges (1 to 10) in buffer
solutions.
3.2.8. NMR Titration [38]
1H NMR spectra obtained after the concomitant addition of F as their
tetrabutylammonium salt to the 2.4×10−2 M solution of the
C in d6-DMSO (Fig. 9)
exhibited the transformation of the initial sharp peak of –NH proton (at δ 11.69 ppm) to a
broad one, which ultimately disappeared upon addition of 1.6 equivalent F; this indicates
involvement of NH in this phenomenon. Although, a respective change in the position of
chemical shift of protons of phenyl ring of
C were expected, but these were practically
remaining almost unchanged on gradual increase of F up to 1.6 equivalents; this finding is
not however clear to us.

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Fig. 9. Partial Stack plot of 1H NMR titration spectra of C (2.4 × 10-2 M) upon addition of TBAF
(0 – 1.6 equiv.) in d6-DMSO at 25oC.
3.2.9. Turn On- Off- On Property Studies [17]
Upon addition of TBAF solutions in the solution of
C in acetonitrile, the color of C
changed from yellow to brilliant blue (turn off). When Cu2+ solution in acetonitrile was
added to this blue solution, the yellow color of
in acetonitrile turned into greenish blue on addition of CN and returned
back to yellow again upon addition of Cu2+. On the other hand fluorescence quenching of
occurred in the presence of F (turn off) and on addition of Cu2+ to the quenched solution,
C returned back (turn on). Similarly the
yellow color of
C
C
it became flourished (turn on) again (Fig. 10). The experiment was repeated by the
sequential addition of F and Cu2+ ions in the same solution, and a reproducible result was
observed in three cycles (Fig. 11b). Thus the compound
C, in association with fluoride ion,
can be considered to be a good turn off- turn on fluorescence chemosensor for Cu2+ ion.

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Fig. 10. (a) UV-vis spectra of 4 μL 2.5 mM C in 2ml MeCN + 130 μL 5 mM F solution which
was titrated gradually with 14.07 mM Cu2+ solution. (b) UV-vis spectra of 4 μL 5 mM C in 2 ml
MeCN + 10 μL 61.4 mM CN solution which was titrated gradually with 14.9 mM Cu2+ solution.
(c) Fluorescence spectra of 4 μL 2.5 mM C in 2 ml MeCN + 6 μL 0.2 M TBAF solution + 24 μL
14.1 mM Cu2+ solution. Inset 1: The visual fluorescence emissions of C (50 μM) by using a UV
lamp upon addition of TBAF (1 equiv.) in MeCN, then upon addition of Cu2+ (1 equiv.). Inset 2:
Relative intensities of C, C + F and C + F + Cu2+.
3.2.10. Mechanism of Binding Mode of Analytes
1
Based on H NMR spectra of chemosensor
C
with different concentration of F
(Fig. 9) and also on the other spectrophotometric experiments, the mechanism of sensing
has been developed (Fig. 11a). The 1H-NMR results indicate that the F definitely combines
with the –NH proton of
C in solution either generating a negative charge on the nitrogen

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atom or forming a strong hydrogen bond to the NH proton. The simultaneous red shift in
흀max value (Fig. 2a) is apparently due to the increase in conjugation after the blending of F
ion with the receptor. The decrease in fluorescence intensity (Fig. 3a) is the result of the
decrease in the probability of relaxation in the excited state, due to the availability of free
electrons in the form of non bonded electron pair. Thus the ultimate negative charge
developed on the receptor would inflect both UV-Vis and fluorescence spectra due to the
change in intramolecular charge transfer (ICT). On addition of Cu2+ ion to the system,
formation of a 1:1 complex with the Cu2+ ion leads to the disappearance of blue color and
regeneration of fluorescence (Fig. 10a). The turn-on fluorescence after addition of Cu2+ ion
is probably due to the coordination of the anionic receptor with the Cu2+ ion by re-
establishing the ICT in the system.

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Fig. 11. (a) Proposed binding mode of C with F and Cu2+. (b) Reversible changes in intensity of
C (4.995 μM) (excitation at 425 nm) after the sequential addition of F and Cu2+solution at room
temperature.
3.2.11. EDTA Reversibility [26]
Weak binding between C and Cu2+ spurred us to examine the reversibility of Cu2+ ion
sensing. When disodium salt of ethylenediaminetetraacetic acid (Na2EDTA) was added to the
mixed solution of C and Cu2+ the absorption intensity decreases and the repetition of the
experiment shows the same result (Fig. 12). Hence C could be recovered after the detection of
Cu2+ ion.
Fig. 12. Reversible changes in absorbance of C (6.8 μM) at 288 nm after the sequential addition
of Cu2+ and Na2EDTA solution at room temperature.
3.3. Practical Application
3.3.1. Colorimetric Test Strips

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For instant detection of F and CN ions, in aqueous solution, a very simple
inexpensive method has been developed. Due to very high affinity of F towards water, it
is difficult to detect F ion directly in aqueous medium. This strip test may be a solution of
this problem. A
C coated yellow paper strip was completely immersed into the aqueous
solution of F, and then the strip was kept into a hot air oven at around 100

C by which
the water absorbed by the strip was vaporised. The color of this paper strip changed from
with F to a large extent.
coated paper strip turned into blue color in case of aqueous CN solution also.
yellow to blue within a few minutes due to an interaction of
C
Similarly
C
It is interesting to note that although after immersing paper strips, pre-coated with
C,
separately in F and CN solution (in DMSO for the former and in DMSO/H2O v/v, 4:1 for
the latter) the yellow color of the strip transformed immediately into bluish green in both
of these cases, but after heating these individually into an oven at ~100 C for 2-3 minutes
the color of the former strip changed into dark blue again but that of the latter into yellow.
Thus F and CN can be distinguished by this simple pre-coated strip tests. Similar strip
tests were also performed for other anions, but none of these exhibit any color change.
3.3.2. Quantitative Estimation of Unknown Aqueous Commercial F
Upon gradual addition of aqueous fluoride ion (0.005 M TBAF in deionised water)
to a solution of chemosensor
C (10 μM) in DMSO [having two absorption peaks at 447 nm
and 658 nm], the absorbance at 447 nm decreases gradually while that at 658 nm increases
gradually (Fig. 13). The rate of increase in absorption intensity at 658 nm was
comparatively greater than the rate of decrease in absorption intensity at 447 nm [39]. The
absorbance of
C
(at 658 nm) against the added known concentration of aqueous F were
plotted, and the best fit straight line passing through the origin was drawn (Inset of Fig. 13).

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Fig. 13. UV-vis spectral changes of
C
(10 μM) upon addition of totally aqueous F
(toothpaste solution) in DMSO at room temperature. Inset: Absorbance at 658 nm as a
function of aqueous F ion concentration.
Table 2. Determination of total dissolved fluoride in various toothpaste samples
Reported F (mg kg-1)
by the toothpaste brand found by linear calibration
curve
Measured F (mg kg-1)
Sample
Fluoridated
agent
Toothpaste A
Toothpaste B
Toothpaste C
NaF
Na2PFO3
NaF
1000
7600
2400
1178 ± 13
7588 ±20
2512 ±10
A measured quantity of the prepared aqueous toothpaste solution was added to the 10 μM
solution of in DMSO and the absorbance at 658 nm was recorded. By plotting the
C
absorbance value to the calibration curve (Inset of Fig. 13) the concentration of F ion in
toothpaste samples were estimated. The calculated F ion concentrations are consistent with
the reported data [40] by the toothpaste brand as listed in Table 2, establishing that the
chemosensor
C
can quantitatively determine the F ions from toothpaste samples.

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4. Conclusion
We have developed a new colorimetric sensor
C
for the detection of two most important
anions, fluoride ion and cyanide ion, consequently the chemosensor can be used as a
selective fluorescence on-off sensor for fluoride ion. Additionally, from aqueous solution
F can be estimated directly spectrophotometrically and also can be detected by strip tests.
Method for distinguishing F and CN by strip tests has also been presented. Moreover, a
unique Cu2+ ion sensing technique using
C
+ F composite was also fabricated.
Acknowledgements
AC and RG are thankful to SERB, DST, India (SR/FT/CS-116/2010) and CSIR, India
(02/0186/14/EMR-II), respectively for financial supports. FIST-DST and UGC-CAS, India
for providing with the Instrumental facilities in the Department of Chemistry, Jadavpur
University and Department of Chemistry, Maulana Azad College, Kolkata for supporting
with some instrumental facilities are also gratefully acknowledged.

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