NIR sensing of Zn(II) and subsequent dihydrogen phosphate detection by a benzothiazole functionalized ninhydrin based receptor

Article abstract of DOI:10.1039/c4ra10556g

A benzothiazole functionalized ninhydrin based chemosensor (L₁), exhibits selective naked eye detection of biologically important zinc ions (from light yellow to orange) accompanied by a 'turn-on' fluorescence emission response in the near infra-red (NIR) region. Most importantly, Zn²⁺ ion induced 'turn-on' fluorescence emission is also preserved in the presence of Cd²⁺ and most of the other competing metal ions which clearly suggests the high sensitivity of the chemosensor towards Zn²⁺. The Job's plot suggests 1:1 binding of the L₁ with the Zn²⁺ ion with a detection limit of 6 nM. Detailed ¹H NMR titrations were also conducted to understand the binding behaviour of L₁ towards the Zn²⁺ ion. The 'L₁-Zn²⁺ ensemble' further shows a ratiometric response to H₂PO₄^- among other competitive anions and nucleotides in the same experimental conditions.

Full text of DOI:10.1039/c4ra10556g

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PAPER
NIR sensing of Zn(II) and subsequent dihydrogen
phosphate detection by a benzothiazole
functionalized ninhydrin based receptor†
Cite this: RSC Adv., 2014, 4, 55689
Abhijit Gogoi and Gopal Das*
1
A benzothiazole functionalized ninhydrin based chemosensor (L ), exhibits selective naked eye detection of
biologically important zinc ions (from light yellow to orange) accompanied by a ‘turn-on’ fluorescence
2+
emission response in the near infra-red (NIR) region. Most importantly, Zn ion induced ‘turn-on’
2+
fluorescence emission is also preserved in the presence of Cd and most of the other competing metal
2+
ions which clearly suggests the high sensitivity of the chemosensor towards Zn . The Job's plot
Received 16th September 2014
Accepted 17th October 2014
2+
1
suggests 1 : 1 binding of the L
1
with the Zn ion with a detection limit of 6 nM. Detailed H NMR
2+
titrations were also conducted to understand the binding behaviour of L
1
towards the Zn ion. The ‘L
1
–
DOI: 10.1039/c4ra10556g
2+
4ꢀ
Zn ensemble’ further shows a ratiometric response to H
2
PO among other competitive anions and
www.rsc.org/advances
nucleotides in the same experimental conditions.
anions, phosphate and molecules featuring this group are
widely studied because of their omnipresent presence in a range
Introduction
The design and development of new uorogenic receptors for of life processes. Again among them, dihydrogen phosphate
metal ions is a promising research area in chemistry due to plays a pivotal role in signal transduction, energy storage, and
9
their essential roles in ecology, biology and clinical applica- construction of the backbone of DNA and RNA. However, only a
ꢀ
1
tions. Fluorescent chemosensors are attractive because of their few H
PO
ion based receptors are known till now in literature
2
4
intrinsic high sensitivity, ease of handling, and real-time which responds to the aforementioned anion either with
2
10
monitoring with a fast response time. Among the metal ions, enhancement or quenching of uorescence. But a ratiometric
2
+
selective and sensitive detection of Zn ion is an intriguing task sensor is always superior from practical point of view as they are
due to the similar spectroscopic properties also exhibited by the avoid of normal quenching process which occurred due to
toxic Cd ion. Zn ion is the second most abundant transition higher sensor concentration, photobleaching, microenviron-
metal ion in the human body aer iron. The total concen- ment around the sensor molecule etc. Various approaches
tration of Zn in mammalian cells is estimated to be in the utilizing hydrogen bonding, anion–p interaction, and the
2
+
3
2+
4,5
11
2
+
range of 100 to 500 mM; the largest fraction is tightly bound to chelation mode of interactions have been adopted to the
6
metallo-proteins.
development of uorescent chemosensors for pyrophosphate,
0
Again among the various features of a chemosensor, highly phosphate ions, ATP/guanosine 5 -triphosphate (GTP), or
12
selective ‘turn-on’ emission at higher wavelength is a desirable phosphorylated peptide. Among these techniques, the metal-
property and mostly cited in recent years. Chemosensors which ligand complexes are turning out as most promising and
exhibit larger Stoke's shis and emit in the near infra-red (NIR) attractive tool for anion recognition and sensing. Anions have a
region (650 nm–900 nm) helps them to overcome the auto usual tendency for the metal–ligand complexes and their

uorescence occurring from biological samples, photo damage structural and geometrical exibility can provide an excellent
7
etc. Again, the investigation of new anion sensors is an another way of organizing anion binding groups for optimal host guest
2
+
2+
2+
hot research area of host–guest chemistry as the anions are interactions. In this regard, Zn , Cd and Cu complexes are
ubiquitous in nature and the anionic species are of funda- extensively studied because of their normal attraction towards
1
3
mental importance in many chemical, biological, medicinal, phosphorylated anions.
8
environment and industrial processes. In particular among all
In our continuing effort to design and synthesis of new
14
sensor molecules, herein we report a benzothiazole function-
alized ninhydrin derived chemosensor (E)-2-(2-(11H-indeno[1,2-
b]quinoxalin-11-ylidene) hydrazinyl) benzo[d]thiazole (L1),
which exhibits highly selective and sensitive ‘turn-on’ uores-
cence response only to Zn ion in longer wavelength NIR
region. Zn responsive chemosensors which show ‘turn-on’
Department of Chemistry, Indian Institute of Technology Guwahati, Guwahati 781039,
India. E-mail: gdas@iitg.ernet.in; Fax: + 91 361 2582349; Tel: +91 361 2582313
†
Electronic supplementary information (ESI) available. CCDC 1000228. For ESI
and crystallographic data in CIF or other electronic format see DOI:
0.1039/c4ra10556g
2
+
2
+
1
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uorescence response in the NIR region are very limited.
Ninhydrin is known for its widespread applications in the elds
of biochemical, analytical and forensic works. Again, the
Paper
15

Results and discussion
Designing aspects of L
1
ninhydrin derived chemosensors exhibited beautiful color- The chemosensor consists of a signalling unit (chromophore/
metric response to various anions and metal ions in aqueous/ uorophore) and a guest binding unit (receptor) integrated
mixed aqueous solution, but there are very few references into one species through an imine bond. The benzothiazole
16
2+
known till date. The only Zn ion induced Stoke's shi of unit thus increases the conjugation of the system and also act
nearly 130 nm was also maintained in physiological medium. as good chelator for the selective binding of a metal ion. Again,
Further the zinc–chemosensor ensemble could successfully the quinoxaline core also acts as chelating uorophore. To
ꢀ
distinguish H PO4 among other competitive phosphorylated understand the importance of the benzothiazole substituent
2
anions.
in the sensing process, a non-chelating substituent phenyl is
2
attached to achieve chemosensor L .
Experimental section
General information and materials
Synthesis of the sensor molecules
All of the materials for synthesis were purchased from
commercial suppliers. The absorption spectra were recorded
on a Perkin-Elmer Lamda-25 UV-Vis spectrophotometer using
The synthetic scheme of the probe (L
1
) and the control
molecule (L ) are shown in Scheme 1. The structure of the L
2
1
is fully conrmed by NMR, mass and single crystal X-ray
diffraction.
1
0 mm path length quartz cuvettes in the range 250–700 nm
wavelengths, while the uorescence measurements were
carried on a Horiba Fluoromax-4 spectrouorometer using 10
mm path length quartz cuvettes with a slit width of 5 nm at 298
Room temperature stirring of ninhydrin (1.02 mmol) and o-
phenylene diamine (1.5 mmol) in methanol with few drops of
concentrated sulphuric acid for 3 hours give a appy orange
like precipitate (L). To a solution of L (1.02 mmol) in ethanol,
K. The mass spectra of L
1
obtained using Waters Q-ToF
Premier mass spectrometer. The NMR spectra were recorded
on a Varian FT-400 MHz instrument and the chemical shis
were presented in parts per million (ppm) on the scale. The
following abbreviations were used to describe spin multiplic-
2
-hydrazinylbenzothiazole (1.5 mmol) and few drops of
concentrated sulphuric acid are added. The resulting solution
is stirred for 4 hours at room temperature. It give an orange
red type precipitate which then ltered and wash with ethanol
and nally with water to give pure L
of L with phenyl hydrazine gives L2.
1
ities in H NMR spectra: s ¼ singlet; d ¼ doublet; t ¼ triplet; m
1
. Similarly condensation
¼
multiplet. The IR spectra were recorded on a Perkin Elmer-
Spectrum One FT-IR spectrometer with KBr disks in the range
Important spectroscopic data for receptor L : yield ¼ 70%;
1
ꢀ
1
1
4
000–450 cm .
H NMR (400 MHz, CDCl , TMS): 13.471 (s, 1H), 8.250 (d, 1H),
3
8
7
1
1
1
.151 (t, 2H), 7.992 (d, 1H), 7.781 (m, 4H), 7.568 (m, 2H),
1
3
.419 (t, 1H), 7.254 (d, 1H). C: 166.872, 153.509, 151.984,
47.317, 141.952, 140.411, 138.082, 137.558, 135.077,
31.950, 131.609, 130.865, 130.395, 129.803, 129.727,
29.660, 126.297, 123.231, 122.457, 121.357, 120.742, 94.653.
X-ray crystallography
The intensity data were collected using a Bruker SMART APEXII
CCD diffractometer, equipped with a ne focus 1.75 kW sealed
˚
tube Mo-Ka radiation (l ¼ 0.71073 A) at 298 (2) K, with
ESI-MS: m/z Calculated for C22H13N S [M] ¼ 379.04, found
5
ꢁ
increasing u (width of 0.3 per frame) at a scan speed of 3 s per
+
[
M + H ] ¼ 380.1019.
frame. The SMART soware was used for data acquisition. Data
Important spectroscopic data for receptor L : yield ¼ 75%,
2
integration and reduction were undertaken with SAINT and
1
3
H NMR (400 MHz, CDCl , TMS): 12.874 (s, 1H), 8.180 (d, 1H),
17
XPREP soware. Multi-scan empirical absorption corrections
8.139 (m, 2H), 7.795 (d, 1H), 7.7541 (m, 4H), 7.561 (t, 2H), 7.480
1
8
were applied to the data using the program SADABS. Struc-
tures were solved by direct methods using SHELXS-97 and were
rened by full-matrix least squares on F2 using SHELXL-97
1
3
(t, 1H), 7.060 (t, 1H), C: 1153.212, 147.645, 143.511, 141.437,
1
1
40.583, 140.011, 132.476, 131.851, 129.921, 129.822, 129.647,
29.365, 129.304, 128.495, 122.455, 120.350, 114.173, 66.058.
19
program package. In the crystal structure, non-hydrogen
atoms were rened anisotropically. Hydrogen atoms attached
to all carbon atoms were geometrically xed. Structural illus-
trations have been generated using ORTEP-3 and MERCURY 1.3
ESI-MS: m/z calculated for C H N [M] ¼ 322.12, found [M +
2
1
14 4
+
H ] ¼ 323.1382.
20
for Windows. Crystal data, as well as details of data collection
and renement for L are summarized below.
Empirical formula: C22
1
H
13
N
5
S, M
w
: 379.44, T ¼ 298 (2) K,
˚
ꢁ
monoclinic, space group: P2(1)/c, a ¼ 9.9373 (12) A, b ¼ 13.5639
˚
˚
ꢁ
ꢀ3
ꢁ
ꢀ1
(
¼
16) A, c ¼ 13.7420 (17) A, a ¼ 90 , b ¼ 102.302 (6) , g ¼ 90.00 , V
3
˚
1809.73 A , Z ¼ 4, Dx (g cm ) ¼ 1.393, m ¼ 0.197 mm
,
F(000) ¼ 784, reections collected/unique ¼ 25 344/4521 [R
¼
int
0
0
.0325], R
1
¼ 0.0435, wR
2
¼ 0.1315 [I > 2s(I)], R
1
¼ 0.0570, wR
2
¼
.1517 (all data), GOF(F2) ¼ 0.920.
Scheme 1 Synthesis of the probes.
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UV-vis and uorescence spectral studies
Result and discussion
ꢀ
3
ꢀ1
Stock solutions of various ions (1 ꢂ 10 mol L ) are prepared
in deionized water. Chloride or nitrate salts are used for metal
ions while tetrabutyl, tetraethyl or sodium salts of the corre-
sponding anions and nucleotides are used for the preparation
Self-assembly behaviour of L
1
in solid state
Block shaped single crystals of L
1
were grown from slow evap-
oration of its propanol solution. It crystallizes in monoclinic
system with P21/c space group (Z ¼ 4). The intramolecular N2–
of anion stock solutions. The stock solution of L
1
and L
2
(5 ꢂ
˚
ꢀ
3
ꢀ1
ꢀ1
H/N4 (2.084 A) hydrogen bonding made the crystal to adopt a
1
1
0
0
mol L ) are prepared in dry DMF and then diluted to 10 ꢂ
˚
˚
ꢀ6
trans-amine form (C15/N1 ¼ 1.305 A, C16/N2 ¼ 1.364 A,
mol L with acetonitrile. For the titration experiments,
˚
ꢀ
3
ꢀ5
ꢀ1
C16/N3 ¼ 1.309 A) and thus forced to adopt a planer geometry.
each time a 1 ꢂ 10 M solution of L (1 ꢂ 10 mol L ) in a
1
This was further helpful to decorates a number of p/p inter-
actions in the packing gure.
quartz optical cell of 1 cm optical path length is titrated with the
escalating concentration of stock solutions by using a micro-
pipette. For the competitive selectivity experiment, uorescence
Each of the receptor molecule interacts with two other
receptor molecules through p/p interactions, where the qui-
noxaline framework was sandwich between quinoxaline
framework of one and benzothiazole functionality of the other
2
+
emission of the L
and presence of other competitive metal ions in an excess
50 equiv.) in the aforementioned experimental medium.
1
–Zn ensembles are collected in the absence
(
˚
receptor. A short C–H/p (2.665 A) interaction from a third
receptor molecule to the benzothiazole group extends the
packing further (Fig. 1).
Evaluation of the apparent binding constant
Receptor L
with an effective concentration of 10.0 ꢂ 10 M is
used for the emission titration studies with a Zn solution
ꢀ6
1
Absorption based selectivity and sensitivity study with various
metal ions
2
+
ꢀ
3
2+
(
0.2 ꢂ 10
M). The effective Zn concentration is varied
ꢀ
5
The sensing ability of the probe L
1
was investigated in aceto-
between 0 and 60 ꢂ 10 M for this titration.
2
+
2+
2+
nitrile solution with various metal ions such as Zn , Cd , Co ,
2
+
2+
2+
3+
3+
3+
2+
+
2+
2+
2+
Pb , Ni , Cu , Fe , Al , Cr , Hg , Ag , Mg , Ba , Mn ,
+
+
2+
1
Na , K , Ca etc. The probe L , exhibited two characteristic
Calculations for the apparent binding constants using
spectrophotometric titration data
absorption peaks nearly at 380 nm and at 442 nm in acetonitrile
solution originating from p–p* transitions and long conjuga-
tion present in the polyaromatic system. However, as evident
from Fig. 2a, a noticeable change in UV-Vis spectral pattern was
The apparent binding constant for the formation of the
respective complexes are evaluated using the Benesi–Hilde-
21
brand (B–H) plot (eqn (1)).
2+
2+
22
manifested only in presence of Zn and Cu ions. Interaction
2
+
of the probe with the Zn ion leads to emerge new absorption
peaks at 536 nm and 418 nm. This was probably due to Zn
1
/(A ꢀ A
0
) ¼ 1/{K(Amax ꢀ A
0
)C} + 1/(Amax ꢀ A
0
)
(1)
2+
1
coordination induced increase in conjugation of L which in
A is the absorbance of L at maximum, A is the observed
0
1
turn leads to concomitant visual colour change of the solution
from light yellow to orange (inset Fig. 2b). Further, on titration
absorbance at that particular wavelength in the presence of a
certain concentration of the analyte (C), Amax is the maximum
absorbance value that was obtained at l ¼ 536 nm during
titration with varying analyte concentration, K is the apparent
2+
the probe solution with increasing Zn ion concentration
showed a progressive decrease in absorbance of the character-
istic 380 nm peak with concomitant increase in absorbance at
ꢀ
1
binding constant (M ) and was determined from the slope of
the linear plot.
5
4
36 nm (Fig. 2b and S10†). The well-dened isobestic point at
2
+
1
68 nm also indicates the formation of a new Zn –L complex.
2
+
Again, addition of higher equivalents of Cu ion (10 equiv.)
Finding the detection limit
The detection limit was calculated on the basis of the uores-
cence titration. The uorescence emission spectrum of L was
1
measured 10 times, and the standard deviation of blank
measurement was achieved. To gain the slope, the ratio of the

uorescence emission at 654 nm was plotted as a concentration
2
+
of Zn .
So the detection limit was calculated with the following
equation
Detection limit ¼ 3s/k
(2)
where s is the standard deviation of blank measurement, and k
is the slope between the ratio of uorescence emission versus
respective analyte concentration.
Fig. 1 (a) ORTEP presentation of L
hydrogen atoms are omitted for clarity). (b) Various non-covalent
interactions in the crystal.
1
with 30% ellipsoid probability
(
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maximum at 654 nm with 15.5 fold enhancement in intensity
and Stoke's shi of 130 nm from a 10 mM solution (uorescence
2
+
2+
quantum yield (F) ¼ 0.11). Other metal ions, such as Cd , Co ,
2
+
2+
3+
3+
3+
2+
+
2+
2+
2+
+
+
Pb , Ni , Fe , Al , Cr , Hg , Ag , Mg , Ba , Mn , Na , K ,
2
+
Ca etc. caused no uorescence change. Similarly, the coloured
2
+
Cu solution was also remain uorescence inactive. Further,
the titration spectra (Fig. 3b) of the chemosensor L
1
, with
2
+
increasing Zn
ion concentration manifested a linear
enhancement in intensity nearly at l ¼ 654 nm while the 545
nm peak was hardly got effected. A plot of uorescent intensity
Fig. 2 Changes in absorption spectra of sensor molecule L
1
(10 mM)
2+
as a function of Zn ion concentration revealed that close to 20
ꢀ4
with the addition of (a) various guest metal ions (c ¼ 10 M) and (b)
2
+
2+
mM concentration of Zn ion the uorescent intensity increase
sharply, and aer that, the rate of enhancement was reduced
varying concentration of Zn ion in acetonitrile. Inset: the visual
2+
2+
1
colour changes after addition of Zn and Cu ions to L in aceto-
2
nitrile solution.
(Fig. 3c). Again, the linear (R ¼ 0.9902) relationship of log(FI)
2
+
(
uorescence intensity) with log[Zn ] (Fig. S11†) also indicates
2
+
that chemosensor L
1
can be used to determine Zn ion
produces a new peak at 454 nm and a red shied absorption
maxima at 550 nm. The new isobestic points at 460 nm and 366
nm were the indicative of the formation of a new Cu –L
complex. This also leads to a visual change in colour of the
concentration. And based on the uorescence titration experi-
ment, the calculated detection limit was 6 nM.
Rapid isomerisation around C]N and photo electron
transfer (PET) from N of benzothizole to quinoxaline uo-
rophore are the probable cause of weak uorescence of L .
1
While the imine N is essential for the chelation with most metal
ions, to verify the importance of benzothiazole group, a control
2
+
1
2
+
solution from light yellow to light pink. More importantly, Cd
2
+
ion, the electronic congener of Zn ion, do not produce any
signicant change in UV-vis pattern.
2 2
compound L was synthesized. As expected, L do not show any
Fluorescence spectroscopic selectivity study of L with various
metal ions
selectivity to any of the aforementioned cations under the same
experimental condition (Fig. S12†). It gives the idea of involve-
ment of imine N, benzothiazole N/S and quinoxaline N (ref. 23)
1
When excited at 440 nm, the probe L showed a distinct emis-
1
2
+
in the chelation with Zn ion which thus cease the isomer-
isation process and the uorescence increases as a result of
chelation enhanced emission (CHEF) effect. Since the border
sion maximum at 545 nm (quantum yield <0.002) with a Stoke's
shi of 103 nm in acetonitrile. In good agreement with the
2
+

nding for absorption spectral studies, only Zn ion promotes
2
+
line Zn ion can interact with both hard and so binding sites
and the chemosensor L can provide required number of proper
binding sites, thus it shows high selectivity for Zn ions.
signicant uorescent enhancement to chemosensor
L
1
2
+
1
(Fig. 3a). Addition of Zn ion leads to emerge a new emission
2
+
2
+
The selectivity of the chemosensor for Zn over other
competing metal ions was preserved in most of the cases
(
Fig. 3d). However, the switch ON red uorescence of L with
1
2
+
Zn ion was interfered by the copper ion only which switches
off the uorescence probably due to its paramagnetic nature.
2
+
Thus this uorish ‘L

1
–Zn ensemble’ can also be used for
2
+
uorescent based sensing of Cu . Pandey et al. have used
similar approach to report ‘on–off–on’ type sensing of Cu and
Ag ions with a uorescent Zn(II) complex. Anyway, the inter-
ference from the Cu ion can be removed by with the addition
2
+
+
24
2
+
2
ꢀ
of S ion which restores the uorescence based on the well-
2
ꢀ
2+
known strong affinity of S for Cu . However, under similar
2
ꢀ
experimental condition, interaction of S anion with ‘chemo-
sensor-Zn ensemble’ caused no change in the uorescence
output signal (Fig. S13†). Again, Cu ion would have little
inuence in vivo, as they exist at very low concentrations. Other
paramagnetic metal ions like Co and Ni do not affect the
2
+
2
+
2
+
2+
2
+

uorescence emission of the ‘chemosensor-Zn ensemble’.
Considering an environmental application, the uorescence
1
Fig. 3 Changes in fluorescence emission response of L (10 mM) with
ꢀ
4
2+
(
a) various metal ions (c ¼ 10 M); (b) increasing concentration of Zn
ion in acetonitrile. (c) The I654/conc. plot showing the change in
experiment was repeated in an optimized mixed aqueous
2+
emission intensity at the l ¼ 654 nm with varying Zn concentration
and (d) the blue bars represents the intensity plot of L
at l ¼ 654 nm
with various metal ions while the red bars indicate the same after
2+
solution. The receptor could also sense the Zn ion in presence
1
of other interfering metal ions in an acetonitrile-buffer (HEPES)
2+
7.4 (8 : 2) mixtures with nearly 6 fold enhancement in intensity
addition of Zn ion to the respective L
1
+ metal ion containing
solution (lex ¼ 440 nm, slit width 5 nm/5 nm).
(Fig. S14†) at the same wavelength.
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2
+
1
Application of the ‘L –Zn ensemble’
2
+
We explore the potential of this pre-synthesized L –Zn
1
ensemble as sensor for common anionic analytes and nucleo-
ꢀ
tides (Fig. 4a). This revealed that among the tested anions; F ,
ꢀ
ꢀ
ꢀ
ꢀ
ꢀ
2
Cl , Br , I , HSO
emissive property of the L
displayed an interesting concentration dependent ratiometric
uorescence response while AMP, ADP, ATP had no effect. To
4
and NO
3
do not affect the uorescence
+
ꢀ
1
–Zn ensemble. However, H
2
PO
4

gain a better insight into the uorescence behaviour, titration
experiment was carried out with a xed concentration of L –
1
2
+
ꢀ
Zn ensemble and a varying amount of H
systematic variation of the amount of H
continuous blue shiing of the emission peak from 654 nm to
2
PO
4
(Fig. 4b). The
2
PO
4ꢀ
displayed a
1
2+
Fig. 5 H NMR stack plot of L
ion in CDCl
1
with increasing concentration of Zn
3
at room temperature with the probable binding mode.
5
H
84 nm with increasing intensity. Aer 2.0 mole equivalents of
ꢀ
2+
2
PO
4
1
ions, the L –Zn ensemble displayed a 22 fold
enhancement in uorescence emission intensity at 584 nm.
However, addition of higher mole equivalents of the same
further quenched the emission and thus a discernible colour
change from orange to yellow was observed. The initial uo-
rescence enhancement was probably due to formation of
Binding mode and composition of metal complex
For better understanding of the sensing mechanism and the
resulting metal ion–sensor complex, Job's plot and H NMR
1
titration were carried out. The apparent binding constant (K) for
ꢀ
2+
2
+
H
2
PO
4
1
complex with the L –Zn ensemble where the addi-
1
the formation of L –Zn complex was calculated by using B–H
tional hydrogen bonding with the receptor might be helpful for
method from the absorption titration spectra. A 1 : 1 binding
stoichiometry was suggested by the Job's plot on the basis of
change in absorbance at 536 nm and the apparent binding
n+
the anion to interact with the L–M complex. Such type of
emission enhancement was also described by Kaur et al. with a
2
+
10d
4
ꢀ1
hexaphenylbenzene derivative-Zn ensemble. Further addi-
constant value was found to be 8 ꢂ 10 M (Fig. S17†).
However, due to the poor solubility of the sensor molecule in
CD CN, the NMR titration was carried out in CDCl solution. It
ꢀ
tion of H PO4 anion may result in the de-complexation via
2
initial coordination of the anion to the metal centre which was
manifested by overall quenching of the uorescence.
3
3
showed a singlet in the very downeld region of the spectra
Further to understand more about the anion binding
behaviour, we have also analyzed the changes in absorption
spectra of the same L –Zn ensemble with the aforementioned
1
which might be the NH proton of the benzothiazole unit due to
2
+
the conjugation in the system. However, addition of Zn ion
causes disappearance of this signal (Fig. 5). A close inspection
in the titration stack plot also revealed that the CH(1) proton
undergoes a downeld shi from 8.250 ppm to 8.286 ppm upon
2
+
anions and nucleotides. This revealed the resemblance of the
ꢀ
spectra obtained aer addition of excess H PO anion to the
2
4
2
+
2
+
L1–Zn ensemble with the free L (Fig. S15†). The absorption
1
addition of 1 equivalent of Zn ion, while CH(2) proton shied
upeld from 8.151 to 8.132 and CH(3) proton shied from 7.991
ppm to 7.961 ppm. Negligible changes were observed for the
rest of the protons which means that these CHs were not bound
to zinc ion. Based on these a plausible binding scheme is also
represented in Fig. 5, where the nitrogen atoms of benzothia-
zole, quinoxaline and hydrazide functionalities are the most
probable chelating atoms.
titration spectra (Fig. S16†), revealed a consistent decrease in
absorbance of the peak at 536 nm with concomitant increase in
absorbance at 380 nm, which was the cause of the observed
colour change from orange to very light yellow. Similar to the

uorescence results, other anions and nucleotides caused no
2
+
change to the absorption pattern of ‘L –Zn ensemble’.
1
Conclusion
In conclusion, we have synthesized a novel benzothiazole
functionalized ninhydrin based chemosensor L , which impels
1
excellent colorimetric and ‘turn-on’ uorometric response in
2
+
near infra-red region (NIR) only towards Zn ion. These spec-
tral changes are signicant enough to enable naked eye detec-
2
+
tion of Zn
ion in physiological medium also. Most
2
+
importantly, the sensitivity of L
in presence of its congener Cd and most of the other tested
metal ions even when present in excess. The Job's plot from the
titration data suggests a 1 : 1 (L –Zn ) binding stoichiometry
and the calculated detection limit was 6 nM. Nature of the Zn
1
2
for Zn ion was also preserved
2+
+
1
Fig. 4 Changes in fluorescence intensity of the L –Zn ensemble
ꢀ4
with (a) various anions and nucleotides (c ¼ 10 M) and (b) with
ꢀ
varying concentration of H
2
PO
4
anion in acetonitrile. The broken red
2
+
1
lines refer to the change in emission upto 2 equiv. and the result of
2
+
ꢀ
excess H
2
PO
4
anion are presented by blue lines (lex ¼ 440 nm, slit
1
width 5 nm/5 nm).
binding mode with L
was also analysed with H NMR titration.
1
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2
+
Finally, L
1
–Zn ensemble could selectivity sense and differen- 11 (a) S. Das, S. Karmakar, S. Mardanya and S. Baitalik, Dalton
ꢀ
tiate H PO4 from pyrophosphate, ATP, ADP etc. nucleotides
and other common anions.
Trans., 2014, 43, 3767–3782; (b) Z. Liu, C. Zhang, X. Wang,
W. He and Z. Guo, Org. Lett., 2012, 14, 4378–4381; (c)
W.-C. Lin, S.-K. Fang, J.-W. Hu, H.-Y. Tsai and K.-Y. Chen,
Anal. Chem., 2014, 86, 4648–4652.
2
Acknowledgements
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