New Chromenylium?cyanine based dual channel chemosensors for copper and hypochlorite sensing

Article abstract of DOI:10.1016/j.dyepig.2020.108445

Based on a change in optical properties of chromenylium?cyanine Schiff base derivatives (5a-c), a series of colorimetric dual sensors have been for the first time designed, synthesized and characterized to monitor Cu²⁺ and OCl^? ions at near-IR (NIR) region in aqueous samples. The colorimetric responses of the sensors toward Cu²⁺ and OCl^? ions were evaluated for aqueous samples within a series of the competitive anions and cations at pH 7.2. The sensors detected Cu²⁺ selectively via Cu²⁺-promoted spirocyclic ring–opening reaction while photophysical change during OCl^? recognition is due to the spirocyclic ring?opening reaction by coordination Cu²⁺ ion generated from reaction between Cu⁺ and hypochlorite ions in analysis media. Mass and IR data suggest a 1:1 complex formation between Cu²⁺ and the receptor via phenolic O atom, N atom from Schiff base and O atom from spirocycle form. The detection limits of the dual sensors (5a-c) were determined to be 3.3 × 10^?8 M, 1.93 × 10^?8 M and 2.36 × 10^?8 for Cu²⁺ determination and 2.83 × 10^?8 M, 2.10 × 10^?8 M and 2.60 × 10^?8 M for OCl^? determination, indicating a high sensitivity of the sensors for Cu²⁺ and OCl^? detection Additionally, we present the first single-crystal structure analysis of a chromenylium-cyanine Schiff base in this study.

Full text of DOI:10.1016/j.dyepig.2020.108445

Journal Pre-proof
New Chromenylium−cyanine based dual channel chemosensors for copper and
hypochlorite sensing
Kaan Karaoglu, Kerem Kaya, Ismail Yilmaz
PII:
S0143-7208(20)30693-8
DOI:
https://doi.org/10.1016/j.dyepig.2020.108445
DYPI 108445
Reference:
To appear in: Dyes and Pigments
Received Date: 17 March 2020
Revised Date: 9 April 2020
Accepted Date: 10 April 2020
Please cite this article as: Karaoglu K, Kaya K, Yilmaz I, New Chromenylium−cyanine based dual
channel chemosensors for copper and hypochlorite sensing, Dyes and Pigments (2020), doi: https://
doi.org/10.1016/j.dyepig.2020.108445.
This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition
of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of
record. This version will undergo additional copyediting, typesetting and review before it is published
in its final form, but we are providing this version to give early visibility of the article. Please note that,
during the production process, errors may be discovered which could affect the content, and all legal
disclaimers that apply to the journal pertain.
© 2020 Published by Elsevier Ltd.

CRediT authorship contribution statement
Kaan Karaoglu: Conceptualization, Methodology, Validation, Investigation, Writing -
Original Draft
Kerem KAYA: Investigation- single crystal X-ray diffraction.
Ismail Yilmaz: Conceptualization, Supervision, Writing - Review & Editing

New Chromenylium−cyanine Based Dual Channel Chemosensors for Copper and
Hypochlorite Sensing
Kaan KARAOGLU1*, Kerem KAYA2, Ismail YILMAZ2
1Recep Tayyip Erdogan University, Vocational School of Technical Sciences, Department of
Chemistry and Chemical Process Technology, 53100, Rize, Turkey
2Istanbul Technical University, Department of Chemistry, 34469, Maslak, Istanbul, Turkey
Abstract
Based on a change in optical properties of chromenylium−cyanine Schiff base derivatives
(5a-c), a series of colorimetric dual sensors have been for the first time designed, synthesized
and characterized to monitor Cu2+ and OCl- ions at near-IR (NIR) region in aqueous samples.
The colorimetric responses of the sensors toward Cu2+ and OCl- ions were evaluated for
aqueous samples within a series of the competitive anions and cations at pH 7.2. The sensors
detected Cu2+ selectively via Cu2+-promoted spirocyclic ring–opening reaction while
photophysical change during OCl- recognition is due to the spirocyclic ring−opening reaction
by coordination Cu2+ ion generated from reaction between Cu+ and hypochlorite ions in
analysis media. Mass and IR data suggest a 1:1 complex formation between Cu2+ and the
receptor via phenolic O atom, N atom from Schiff base and O atom from spirocycle form. The
detection limits of the dual sensors (5a-c) were determined to be 3.3×10-8 M, 1.93×10-8 M and
2.36×10-8 for Cu2+ determination and 2.83×10-8 M, 2.10×10-8 M and 2.60×10-8 M for OCl-
determination, indicating a high sensitivity of the sensors for Cu2+ and OCl- detection
Additionally, we present the first single-crystal structure analysis of a chromenylium-cyanine
Schiff base in this study.
Keywords: Hypochlorite sensing, Near-IR chemosensor, Chromenylium-cyanine
1

*Corresponding author, Tel.: +90(464)223 6126
E-mail address: kaan.karaoglu@erdogan.edu.tr (K. KARAOGLU)
2

1. Introduction
Hypochlorite (OCl-) is one of the most used oxidizing agents in a wide range from industrial
processes to household cleaning. Although the human immune system generates OCl- from
hydrogen peroxide and chloride ion by myeloperoxidase enzyme to protect the body from
pathogens, the accumulation of OCl- to the body can result in illnesses such as cardiovascular
diseases, inflammatory reactions, arthritis, and cancer. Thus, the development of sensitive and
selective sensors for OCl- sensing in water samples is of great importance [1–3]. One of the
sophisticated methods for OCl- detection includes the use of rhodamine-based (Cu2+) sensors
which generate an optical response based on the spirocyclic ring-opening reaction [4–6]. In
this method, Cu2+ is generated by oxidation of Cu+ ions upon the addition of OCl−, then an
absorption band is observed corresponding to Cu2+–promoted spirocyclic ring–opening
reaction [7,8]. To this end, various rhodamine B-based sensors have been developed in recent
years, and these derivatives exhibit a characteristic intense absorption band with a maximum
at about 550 nm [9–12]. In order to decrease photodamage in vivo, chromenylium-cyanine
based chemosensors have been designed for metal ion sensing [13–17].
However, to the best of our knowledge, chromenylium-cyanine based OCl- sensing has
been never reported. In this study, we design and synthesize some chromenylium-cyanine
chemosensors to detect selectively OCl- ion in aqueous samples, via Cu2+ promoted the ring-
opening reaction. In order to achieve this goal, first, we investigated the selectivity of the
sensors to Cu2+ ion, and then performed a series of experiments in order to evaluate OCl-
sensing properties of the sensors. The spectrometric assays showed that the sensor is highly
sensitive and selective to OCl- in the near-IR region. The chemical structures of chemosensors
have been characterized by FT–IR, UV−Vis, 1H– and 13C–NMR, mass spectrometry. We also
present the first single-crystal structure analysis of a chromenylium-cyanine Schiff base.
3

2. Experimental
2.1. Materials and apparatus
The solutions of Na+, K+, Mg2+, Ca2+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Al3+, Cd2+, Hg2+ and
Pb2+ ions in water were prepared freshly for each measurement from their nitrate salts except
for Mn2+ was prepared acetate salt. Aqueous solutions of sodium salts of F- Cl-, Br-, I-,
2-
-
-
3-
-
2-
CH3COO-, CO3 , ClO-, NO2 , NO3 , PO4 , HSO3 and SO4 at 1×10-3 M were prepared in
deionized water and stored in amber glass bottles. 3-Diethylaminohenol, phthalic anhydride,
hydrazine
hydrate,
2,4–dihydroxybenzaldehyde
and
(benzotriazol-1-
yloxy)tris(dimethylamino)phosphonium hexafluorophosphate were obtained from Sigma
Aldrich.
Fourier transform infrared (FT-IR) spectra were recorded on a Perkin Elmer Spectrum
100 spectrometer equipped with an ATR apparatus. Mass spectra were recorded using
Thermo Sci. TSQ Quantum Access MAX Triple Stage Quadrupole mass spectrometer
equipped with heated electrospray ionization (H-ESI) probe at the Central Research
Laboratory of Recep Tayyip Erdogan University. 1H and 13C NMR spectra were recorded on
an Agilent Technologies 400/54 spectrometer at the Istanbul Technical University. UV–Vis
spectra were recorded on a Perkin Elmer Lambda 35 UV–Vis spectrophotometer. HPLC
analysis were carried out on a Thermo Scientific Finnigan Surveyor HPLC system equipped
with photodiode array (PDA) detector. The injection volume was 20 μL and the flow rate
maintained at 1.2 mL min−1 . A C18 column was applied for the chromatographic separation
at 25 °C and the elution solvents were A and B containing acetonitrile-water (70:30, v/v) and
acetic acid-water (2:98, v/v), respectively.
2.2. Synthesis of chemosensor (5a, 5b, and 5c)
The starting hydrazide, (4'Z)–2–amino–6'–(diethylamino)–4'–{2–[(2Z)–1,3,3–trimethyl–2,3–
dihydro–1H–indol–2–ylden]ethyliden}–1',2,2',3,3',4'–hexahydrospyro[isoindol–1,9'–
4

xanthene]–3–on, was synthesized by literature procedures [18,19]. The hydrazide (0.573 g, 1
mmol) was dissolved in absolute ethanol (10 mL) and solution purged by argon for 30 min.
Then aldehyde (1.1 mmol) was added into the solution, and the mixture was stirred at room
temperature for appropriate time controlled by thin–layer chromatography (TLC) (Scheme 1).
The solvent was evaporated by rotary evaporator under reduced pressure to dryness, and
crude product was purified by column chromatography on silica gel (EtOH/CH2Cl2, 1:20).
Finally, an aliquot of the isolated fraction was then used to verify the purity by HPLC (Figure
S. 1).
5a: Yield: 73%. Melting point: 244 °C. Color: Yellow. Rf: 0.57, EtOH/CH2Cl2, (1:20, v/v).
Analytical data for C44H44N4O3, ESI-MS (m/z) 676.85 [M+H]+ (676.85 calc.). HPLC purity
98.4% (tR=30.06 min). FT-IR (ATR, cm-1): 3079, 2969, 2929, 1702, 1619, 1589, 1310, and
1
1124. H NMR (δ, ppm): 11.13, 9.14, 7.95, 7.57−7.53, 7.49, 7.25, 7.23−7.19, 7.17, 6.90,
6.86, 6.82, 6.63, 6.46, 6.29, 6.46, 5.39, 3.35, 3.160, 2.67, 2.46, 1.76, 1.79, 1.63, 1.26, 1.18.
13C NMR (δ, ppm): 164.7, 158.8, 157.8, 152.5, 149.8, 148.9, 148.3, 145.4, 138.9, 133.3,
131.4, 131.3, 129.8, 128.5, 127.7, 121.5, 120.2, 119.3, 118.6, 117.1, 108.5, 105.7, 104.8,
103.3, 97.9, 92.2, 68.2, 45.5, 44.3, 29.1, 28.4, 25.3, 22.2, 12.6.
5b: Yield: 26%. Melting point: 195 °C. Color: Yellow. Rf: 0.36, EtOH/CH2Cl2, (1:20, v/v).
Analytical data for C44H44N4O4, ESI-MS (m/z) 693.25 [M+H]+ (692.84 calc.). HPLC purity
96.8% (tR= 30.14 min). FT-IR (ATR, cm-1): 3056, 2969, 2929, 1687, 1622, 1589, 1313, and
1
1115. H NMR (δ, ppm): 11.34, 9.69, 9.05, 7.93, 7.46−7.55, 7.25, 7.18, 7.01, 6.86, 6.62,
13
6.46, 6.39−6.33, 6.28, 5.39, 3.35, 3.16, 2.61, 2.46, 1.79, 1.76, 1.63, 1.26, 1.18. C NMR (δ,
ppm): 194.2, 164.6, 160.8, 159,1, 157.7, 153.5, 152.5, 149.8, 148.9, 148.1, 145.3, 138.9,
135.1, 129.9, 128.5, 127.7, 123.8, 123.5, 121.5, 120.2, 120.0, 119.3, 112.2, 108.5, 107.5,
105.7, 104.9, 103.5, 98.0, 92.1, 68.3, 45.5, 44.3, 31.2, 28.3, 25.3, 22.1, 12.6.
5

5c: Yield: 36%. Melting point: 155 °C. Color: Yellow. Rf: 0.41, EtOH/CH2Cl2, (1:20, v/v).
Analytical data for C48H53N5O4, ESI-MS (m/z) 748.25 [M+H]+ (747.97calc.). HPLC purity
97.9% (tR=33.11 min). FT-IR (ATR, cm-1): 3066, 2968, 2928, 1694, 1624, 1593, 1314, and
1126. 1H NMR (δ, ppm): 11.24, 9.08, 7.91, 7.51-7.47, 7.19-7.18, 6.99, 6.85, 6.62, 6.37, 6.27,
6.16, 6.13, 5.39, 3.33, 3.15, 2.56, 2.48, 1.86, 1.83, 1.77, 1.75, 1.18, 1.14. 13C NMR (δ, ppm):
164.2, 160.8, 157.4, 154.8, 152.5, 150.5, 149.7, 148.7, 148.0, 145.4, 139.0, 132.9, 132.7,
130.5, 128.3, 127.7, 127.6, 123.4, 123.0, 121.5, 120.6, 119.4, 119.1, 108.4, 107.4, 105.6,
105.2, 104.0, 103.3, 98.2, 98. 0, 92.3, 68.1, 45.4, 44.5, 29.1, 28.4, 25.3, 22.9, 22.2, 12.6.
Scheme 1. Synthesis scheme of the sensor. i: Toluene, 3h, 35% NaOH, 10,0 M HCl; ii:
H2SO4, cyclohexanone, 2h; iii: 2-(1,3,3-trimethylindolin-2-ylden)acetaldehyde; iv: hydrazine
hydrate, BOP reagent; v: 2-hydroxybenzaldehyde, vi: 2,4-dihydroxybenzaldehyde, vii: 4-
diethylamino-2-hydroxybenzaldehyde.
6

2.3. Sensing Experiments
Nitrate salts of the Na+, K+, Mg2+, Ca2+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Al3+, Cd2+, Hg2+
and Pb2+ were used to evaluate cation sensing properties of the sensors. Stock solutions (3×10-
3
2-
-
-
3-
-
M) of the sodium salts of F- Cl-, Br-, I-, CH3COO-, CO3 , ClO-, NO2 , NO3 , PO4 , HSO3
2-
and SO4 were prepared in deionized water, and 3×10-3 M solutions of the sensors were
prepared in acetonitrile (MeCN). All the solutions were stored in amber bottles at room
temperature. All the spectroscopic measurements were performed in triplicate and data are
presented as the mean, with error bars reflecting the standard deviation.
Detection limit based on the standard deviation of the response and the slope by
IUPAC was determined from the Eq. (1).
ꢀꢁꢂ = ꢃ.ꢃ×ꢄꢅ
(1)
ꢄ
where Sd is the standard deviation of the blank measurements (n=10) and S is the slope of the
calibration curve.
2.4. Single Crystal X-Ray Crystallography
The single crystal of 5a with dimensions 0.03x0.10x0.20mm was grown by slow evaporation
of ethanol solution. The crystal was mounted on a micromount and attached to a goniometer
head on a Bruker D8 VENTURE diffractometer equipped with PHOTON100 detector and
measured with graphite monochromated Mo-Kα radiation (λ = 0.71073 Å) using 1.0° of Ω
and ϕ rotation frames at room temperature (295K). The structure has been solved by intrinsic
method SHELXS-1997 [20] and refined by SHELXL-2014/7 [21]. Molecular drawings are
generated using OLEX2. Ver. 1.2-dev [22].
7

3. Results and Discussion
In order to evaluate effect of electron-donating/withdrawing group for complexation,
Schiff bases of chromenylium-cyanine (5a-5c) from 2-hydroxybenzaldehyde, 2,4-
hydroxybenzaldehyde and 4-diethylamino-2-hydroxybenzaldehyde were synthesized. Spectral
data obtained for the sensors were in accordance with our previous study [17]. Characteristic
C=O stretching modes of spirocycle were observed at about 1690 cm-1 for the sensors in FTIR
spectra. Bands at about 1620 cm-1 were also assigned to C=N stretching of imine bond. The
disappearance of the carbonyl band and the shift of the imino stretching to ~1600 cm-1 clearly
suggest complexation of the Cu2+ (Figure S. 2–Figure S. 5) [23]. The chemical structures of
1
the sensors were also confirmed by NMR spectra. In the H NMR spectra of the 5a, –OH
proton and imine proton signals were observed as a singlet at 11.132 and 9.140 ppm,
respectively. Proton signals of H46 and H34 were observed at 5.391 and 6.459 ppm, for atom
labels see structure in Figure S. 6. The signals of methyl protons of H1, H39, H37 and H38
were observed as singlets at 1.1180, 3.160, 1.759, and 1.788 ppm, respectively. A doublet of
quartet (J=5.6 Hz, J=1.6 Hz) appeared at 3.351 ppm which assigned to H2 and H5 protons.
1
The signals of aromatic protons were in the range of 6.339−7.952 ppm. H NMR signals of
the H16, H17 and H17 were observed at 2.607–2.456, 1.256 and 1.627 ppm, respectively
1
[24]. The Н NMR spectra of 5b and 5c are similar that of 5a and differ only in substituted
group at the para-position of the imine moiety (Figure S. 6−Figure S. 9). The characteristic
13C NMR signals of carbonyl (C32) and the C12 corresponding to spirocycle closed form of
5a were observed at 164.378 and 68 ppm, respectively (Figure S. 6−Figure S. 9). The similar
spectral features were observed for 5b and 5c. The mass peaks of positive ions were detected
at m/z 677.16, 693.25 and 748.25, corresponding to the protonated molecular ions for 5a, 5b
and 5c, respectively (Figure S. 10–Figure S. 12). Molecular structure of the 5a-Cu2+ complex
has been also investigated by ESI mass spectra (Figure S. 13). The molecular ion peak for the
8

complex structure was observed at 769.77 m/z, which confirmed the formation of a 1:1
complex of 5a with Cu2+. Mass data were consistent with IR data that suggest coordination
between Cu2+ and phenolic O atom, N atom from Schiff base and O atom from spirocycle
(Figure S. 14).
Compound 5a was structurally characterized by single crystal X-ray diffraction, and thermal
ellipsoids are plotted in Figure 1. Figure 2 shows the crystal packing of the structure showing
π—π interaction and Figure 3 shows the intra- and intermolecular hydrogen bonding of the
crystal lattice.
Figure 1. Thermal ellipsoids of 5a are drawn at 50% probability. Hydrogen atoms are omitted
for clarity.
9

Figure 2. Face-to-face π−π stacking occurring between phenoxy units of structure 5a.
Figure 3. Intra- and intermolecular hydrogen bonding of the crystal structure 5a.
10

The crystal data and structure refinement parameters of 5a are given in Table 1. Table S. 1
contains the selected bond lengths, bond and torsion angles for 5a. Table S. 2 contains the
hydrogen bonding geometries. CCDC 1985102 contains the supplementary crystallographic
data for 5a. Further details on crystal data, data collection, and refinements can be found in
the supporting information.
11

Table 1. Crystal data and structure refinement parameters for 5a.
5a
C44H44N4O3
Empirical formula
676.83
295(0)
0.71073
Triclinic
P - 1
Formula weight (g/mol)
T(K)
λ(Å)
Crystal system
Space group
Unit cell dimensions: (Å, º)
11.8062(15)
12.3990(16)
12.8593(16)
1838.3(4)
a
b
c
V(Å3)
92.222(3)
102.121(3)
91.095(3)
2
α
β
γ
Z
Absorption coefficient (mm-1)
Dcalc (g/ cm3)
0.077
1.223
720
F(000)
0.03 x 0.10 x 0.20
2.13 to 25.00
-13≤h≤14
-14≤k≤14
-15≤l≤15
Crystal size (mm)
θ range for data collection (º)
Index ranges
48124
6471
Reflections collected
Independent reflections
99.9
Coverage of independent reflections (%)
Data/parameters
6471/ 465
0.998 - 0.991
R1 = 0.0685
wR2 = 0.1243
R1 = 0.1576
wR2 = 0.1584
1.009
Max. and min. transmission
Final R indices [I≥2σ(I)]
R indices (all data)
Goodness-of-fit on F2
R1: agreement between the calculated and observed models, wR2: squared F-values.
5a crystallized in a triclinic crystal system with P-1 space group. Each unit cell contains two
molecules (Z=2). The crystal lattice is strongly stabilized by intra- and intermolecular
hydrogen bonding, also by π—π stacking which can be seen in Figure 2 and Figure 3,
12

respectively. Xanthene moiety and isoindole unit having imine functionality are nearly
perpendicular to each other with a dihedral angle of 86.54o. On the other hand, xanthene and
isoindole having quaternized ammonium are nearly coplanar with a dihedral angle of 2.03o.
C34-C45 single bond of 1.44(6) Å is much shorter than average carbon-carbon single bond of
1.54 Å which shows the extended π-conjugation of the system. Carbon atoms of the
diethylamine groups contain minor disorders. The intramolecular hydrogen bonding occurs
between phenoxy hydrogen and imine nitrogen. The intermolecular hydrogen bonding occurs
between phenyl hydrogen and carbonyl oxygen of a neighbour molecule. A strong face-to-
face π—π interaction occurs between neighbour phenyl units with 3.92 Å centroid-to-centroid
distance and a shift of 0.93 Å. C39-N4 distance of 1.44(4) Å belonging to the ammonium unit
is shorter than similar crystals found in the literature.5-7 This also proves the extended π-
system of the molecule. N1-N2 distance of 1.38(4) Å and C25-O2 double bond distance of
1.22(5) Å belonging to the carbonyl unit shows that the keto tautomer is dominant in the
structure.
3.1. Response of the chemosensors to cations and anions
Sensing properties of the sensors were tested within a series of mono-, di- and trivalent metal
ions in MeCN/HEPEs buffer (2:1, v/v, pH 7.2). In order to determine selectivity of the sensors
at room temperature, 1×10-5 M solution of the sensor was used each time in the presence of 5
equivalents of different metal ions. The UV−Vis. spectra of the sensors show a very weak
absorption band between 625 and 750 nm, and an absorption band located at 711 nm was
significantly enhanced upon addition of Cu2+ ion (Figure 4A) [25,26]. Other metal ions, Na+,
K+, Mg2+, Ca2+, Cr2+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Al3+, Cd2+, Hg2+ and Pb2+, did not
cause any significant changes under the identical conditions. The sensors detected Cu2+
selectively via Cu2+-promoted spirocyclic ring–opening reaction.
13

To investigate the selectivity of the sensor in the presence of competitive metal ions,
such as Na+, K+, Mg2+, Ca2+, Cr2+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Al3+, Cd2+, Hg2+ and
Pb2+, the experiments were performed by addition of 1.0 equiv. of Cu2+ to solutions
containing 5.0 equivalent of other metal ions in MeCN/HEPEs buffer (2:1, v/v, pH 7.2)
media. The selectivity experiments for mono–, di–, and trivalent cations showed that none of
the competitive metal ions had no obvious interference with the detection of Cu2+, and
indicated the good selectivity of the sensor for Cu2+ ion (Figure 4B). Details of the sensing
properties of 5a and 5c were given in Supplementary materials (Figure S. 21–Figure S. 22)
14

A
B
Figure 4. UV−vis. experiments of 5b (1.0×10-5 M) in MeCN/H2O (2:1, v/v, pH 7.2); A)
absorption spectra of the sensor in the presence of 5×10-5 M metal ions B) Absorbance of the
sensor (1.0×10-5 M) with Cu2+, followed by 5 equivalent of competitive metal ions.
The instant response of the sensor to Cu2+ enables determination of the detection limit
of the sensor by titration experiments. The titration experiments were performed by
15

successive additions of 2 μL solution of Cu2+ ion to 1×10-5 M sensor solution at pH 7.2 in
HEPES buffer media (Figure 5)
The detection limits of the 5a, 5b and 5c were determined to be 3.3×10-8 M, 1.93×10-8
M and 2.36×10-8 M, respectively, indicating a high sensitivity of the sensors for Cu2+. For
comparison, literature data are given in Table 2. These results indicated that the sensors 5a-c
can serve as a potential candidate of a “naked-eye” chemosensor for Cu2+ in aqueous media.
A
B
Figure 5. UV−vis titration experiments of 5b (1.0×10-5 M) in MeCN/H2O (2:1, v/v, pH 7.2)
A) with Cu2+ B) the titration curve derived from the titration with Cu2+ at 711 nm.
Table 2. A comparison table about the detection limits for Cu2+.
Structure
Working range, mol L-1
1.0×10-5 to 3.0×10-4
NA
LOD, mol L-1
3.42×10-6
1.63×10-6
5.00×10-8
2.36×10-8
1.50×10-8
Reference
[27]
Rhodamine B
Rhodamine B
[28]
Chromenylium-cyanine
Chromenylium-cyanine
Chromenylium-cyanine
0 to 5.0×10-6
3.3×10-7 to 1.0×10-5
1.0×10-7 to 6.0×10-7
[29]
This work
[30]
NA: Not available
It was found that the sensors formed colorless solutions with cuprous ions in 1:2(v/v)
water/ MeCN at pH = 7.2. Upon addition of hypochlorite to these solutions, an absorption
peak centered at 711 nm appeared as a result of the formation of Cu2+ by in situ oxidation of
16

Cu+ ion (Figure 6). In order to evaluate the hypochlorite sensing properties of the sensors via
Cu+→Cu2+ oxidation, the same experimental procedure adopted for metal sensing properties
2-
-
-
were performed using common anions (F- Cl-, Br-, I-, CH3COO-, ClO-, CO3 , NO2 , NO3 ,
3-
-
2-
PO4 , HSO3 and SO4 ) and H2O2 , one of the reactive oxygen species, in water (Figure 7).
Figure 6. Proposed spirocyclic ring-opening reaction induced by Cu+/Cu2+ oxidation.
Absorption behaviors of blank samples were investigated in both some common
solvents (Figure S. 18) and different temperatures (Figure S. 19). It was found by
spectrophotometric analysis that stability of the cuprous ions was higher in MeCN.
Temperature dependencies of the sensors were also investigated in the same media, and
significant absorption changes for blank samples were observed by increasing temperature.
Thus, all the spectrophotometric experiments were performed at 25 °C in MeCN/water
mixture. Sodium ascorbate was used to stabilize cuprous ions in analysis media, and
ascorbate:Cu+ ratio was optimized as shown in Figure S. 20. The optimum concentrations of
Cu+ and sodium ascorbate were determined as 1.0×10-4 M and 4.0×10-6 M, respectively. The
optical responses of the sensors toward ClO- were analyzed in MeCN/H2O (2:1, v/v, pH 7.2)
at pH 7.2 applying 5 minutes incubation time. In order to determine selectivity of the sensors
17

at room temperature, 1×10-5 M solution of the sensors was used each time in the presence of 5
2-
-
-
3-
-
equivalents of F- Cl-, Br-, I-, CH3COO-, ClO-, H2O2, CO3 , NO2 , NO3 , PO4 , HSO3 and
2-
SO4 . The colorimetric responses of the 5a, 5b and 5c to OCl– ion were given in Figure 7.
The more intense absorption bands were obtained for OCl– ion for all the sensors while the
other anions, showed negligible absorption bands. However, selectivity experiments indicated
that the presence of 5-fold excess of H2O2 caused interference by oxidation of Cu+ ion to
Cu2+. The sensors responded to H2O2, although not as well as in the case of ClO-. Because of
weaker oxidation capacity of the H2O2 than ClO-, the observed absorbance chance is much
lower than that in the presence of ClO- [7].
18

A
B
C
Figure 7. Absorption changes of sensor (1.0×10-5 M) upon the addition of various anions and
H2O2 (5 equiv) in water/MeCN (2:1, v/v) media at pH = 7.2. A: 5a, B: 5b and C: 5c.
The competition experiments were conducted in the presence of ClO– mixed with 10
2-
-
equivalents of the other relevant anions, such as F- Cl-, Br-, I-, CH3COO-, ClO-, CO3 , NO2 ,
-
3-
-
2-
NO3 , PO4 , HSO3 and SO4 . Competitive experiments performed in triplicate showed that
5b yielded the higher recovery and lower RSD values than those of 5a and 5c (Figure 8).
Although, sensors showed some selectivity toward H2O2, experimental result indicated that
H2O2 has had no obvious interference for the detection of ClO-.
19

A
B
C
Figure 8. Effect of the competitive anions (5.0×10-5 M) on the interaction between sensors
(1.0×10-5 M) in water/MeCN (2:1, v/v) media at pH = 7.2. A: 5a, B: 5b and C: 5c
20

The linear working ranges of the sensors were obtained by successive additions of solution of
hypochlorite into analysis media (Figure 9). Then calibration curves were established in order
to calculate detection limit of the sensors. The experiment was performed three times, with
triplicate samples in the individual experiments. A calibration curve for OCl– concentration vs
absorption was obtained for sensing experiments, and error bars given in Figure 10 represents
the standard error for triplicate samples. The detection limits were calculated as 2.83×10-8 M,
2.10×10-8 M and 2.60×10-8 M for 5a, 5b and 5c, respectively. The results are tabulated and
compared with literature data in Table 3. As seen in Table 3, 5a-c displayed highly selective
and sensitive response toward OCl-. A detailed spectrophotometric analysis for 5a and 5c is
given in Figure S. 23 in Supplementary file.
Table 3. A comparison table about the detection limits for ClO-.
Working range, mol L-1 LOD, mol L-1 Reference
Cyanine
Quinolone
Chromenylium-cyanine
Coumarine
NA
1.00×10-7
8.90×10-8
2.10×10-8
0.90×10-9
[31]
[32]
This work
[33]
8.00×10-7 to 1.25×10-5
8.30×10-6 to 1.7×10-6
NA
NA: Not available
A
B
Figure 9. Optical responses of the 5b to hypochlorite ion sensors in water/MeCN (2:1, v/v)
media at pH = 7.2. A, varying in terms of molar concentration between 4.2×10-7 and 1.3×10-6
M .
21

Figure 10. The calibration curve for OCl- sensing experiments was calculated by a least-
squares curve-of-best-fit procedure for 5b.
Accuracy and precision of the sensors were tested in tap water samples. OCl- ion was
not detected in tap water, and all experiments were performed on contaminated water
samples. Recovery values higher than 94% were obtained for all the sensors. As seen in Table
4, 5a and 5c showed lower repeatability for concentrations lower than 5.00 µM of the OCl-.
The RSD data calculated for 5b showed that values of repeatability were acceptable for the
concentration range of 1.67–6.70 µM. The higher repeatability could be related to polarization
of imine double bond by electron–withdrawing hydroxyl group on aromatic ring, which
would favor the complexation of Cu2+ ion [34].
22

Table 4. Results for the determination of hypochlorite in tap water samples.
Sensor
Added (μM)
Recovery (%)
100.82
99.16
RSD(n=3)
1.67
5.03
5.00
2.24
5a
6.70
100.41
105.42
94.60
0.97
1.67
2.85
5.00
3.81
5b
5c
6.70
102.71
102.74
97.28
1.67
1.67
16.87
1.12
5.00
6.70
101.37
3.76
3.2. Theoretical Calculations
The molecular structures of the sensors have been investigated by density functional theory
(DFT) in the gas phase. Theoretical calculations have been performed using the GAMESS
2018 program [35], and data were visualized by Gabedit [36]. The Becke-3-Lee-Yang-Parr
(B3LYP) exchange-correlation functional with a 6-31G(d, p) basis set for the H, C, N and O
and LanL2Dz for the Cu atom were used geometric optimizations [37,38]. No imaginary
frequency was obtained in vibrational frequency calculations for optimized geometries, which
demonstrated that all of the theoretical structures at a minimum on the potential energy
surface. DFT calculations showed that sensors have almost planar geometry with close to 0°
dihedral angle between chromenylium-cyanine framework and indole moiety (Figure 11A). In
addition, DFT calculations revealed that the spirolactam ring was perpendicular to the
23

chromenylium-cyanine framework. It indicated that the theoretical results were quite
compatible with the single crystal X-ray diffraction analysis.
The sensor is likely to coordinate with Cu2+ via phenolic O, amido N and carbonyl O
atom resulting in the ring opening reaction of the spirocyclic unit, as seen Figure 11B [39].
The geometry of 5b-Cu2+ complex was optimized by using different basis set and different
functional combinations and computed electronic transitions for corresponding geometries
were compared with the experimental results (Table S. 3). Calculations showed that
SDD/B3LYP yields accurate prediction for electronic spectra. According to DFT data, the
low-energy optical transitions can be assigned to π→π* transition localized on the Schiff base
unit and chromenylium rings, and shoulder at 650 nm in experimental spectrum could be
assigned to ligand to metal charge transfer transition (LMCT) [40] (see Figure S. 17 for
detailed analysis).
A
B
Figure 11. DFT optimized geometries of 5b and its Cu2+ complex.
24

4. Conclusion
In summary, we developed a series of chromenylium−cyanine based Schiff base sensor to
monitor OCl− ion in aqueous media. Colorimetric response of the sensors to OCl− ion was
obtained by Cu2+ induced spirocyclic ring opening of the chromenylium−cyanine. An
absorption enhancement was observed at near-IR region a result of complexation of the
sensors with Cu2+ that generated by oxidation of Cu+ ions upon the addition of OCl− in
analysis media. The results indicated that sensors exhibit excellent selectivity toward OCl−
and could be used as a selective colorimetric probe for detecting OCl− ion in aqueous samples.
The detection performances of the sensors were also tested by means of RSD% in tap water,
and it was found that the sensors seem useful for the determination of the OCl− in real
samples.
Acknowledgement
This work is supported by The Scientific and Technological Research Council of Turkey
(TÜBİTAK) project number 217Z250. The numerical calculations reported in this paper were
fully performed at TUBITAK ULAKBIM, High Performance and Grid Computing Center
(TRUBA resources).
References
[1] Q. Xu, K.-A. Lee, S. Lee, K.M. Lee, W.-J. Lee, J. Yoon, A Highly Specific Fluorescent
Probe for Hypochlorous Acid and Its Application in Imaging Microbe-Induced HOCl
Production,
J.
Am.
Chem.
Soc.
135
(2013)
9944–9949.
https://doi.org/10.1021/ja404649m.
[2] D. Wu, L. Chen, Q. Xu, X. Chen, J. Yoon, Design Principles, Sensing Mechanisms,
and Applications of Highly Specific Fluorescent Probes for HOCl/OCl –, Acc. Chem.
25

Res. 52 (2019) 2158–2168. https://doi.org/10.1021/acs.accounts.9b00307.
[3] W. Ma, Y. Ding, M. Zhang, S. Gao, Y. Li, C. Huang, G. Fu, Nature-inspired chemistry
toward hierarchical superhydrophobic, antibacterial and biocompatible nanofibrous
membranes for effective UV-shielding, self-cleaning and oil-water separation, J.
Hazard. Mater. 384 (2020) 121476. https://doi.org/10.1016/j.jhazmat.2019.121476.
[4] S. Goswami, S. Das, K. Aich, P.K. Nandi, K. Ghoshal, C.K. Quah, M. Bhattacharyya,
H.-K. Fun, H.A. Abdel-Aziz, A rhodamine-quinoline based chemodosimeter capable of
recognising endogenous OCl- in human blood cells, RSC Adv. 4 (2014) 24881–24886.
https://doi.org/10.1039/c4ra03200d.
[5] Y.-R. Zhang, X.-P. Chen, Jing-Shao, J.-Y. Zhang, Q. Yuan, J.-Y. Miao, B.-X. Zhao, A
ratiometric fluorescent probe for sensing HOCl based on a coumarin-rhodamine dyad.,
Chem. Commun. (Camb). 50 (2014) 14241–4. https://doi.org/10.1039/c4cc05976j.
[6] H.J. Lee, M.J. Cho, S.K. Chang, Ratiometric Signaling of Hypochlorite by the
Oxidative Cleavage of Sulfonhydrazide-Based Rhodamine-Dansyl Dyad, Inorg. Chem.
54 (2015) 8644–8649. https://doi.org/10.1021/acs.inorgchem.5b01284.
[7] X. Lou, Y. Zhang, J. Qin, Z. Li, Colorimetric hypochlorite detection using an
azobenzene acid in pure aqueous solutions and real application in tap water, Sensors
Actuators, B Chem. 161 (2012) 229–234. https://doi.org/10.1016/j.snb.2011.10.024.
[8] X. Lou, Y. Zhang, Q. Li, J. Qin, Z. Li, A highly specific rhodamine-based colorimetric
probe for hypochlorites: a new sensing strategy and real application in tap water,
Chem. Commun. 47 (2011) 3189–3191. https://doi.org/10.1039/C0CC04911E.
[9] Y. Huang, M. Wang, Z. Yang, M. She, S. Wang, High efficient probes with Schiff base
functional receptors for hypochlorite sensing under physiological conditions, Chinese
Chem. Lett. 25 (2014) 1077–1081. https://doi.org/10.1016/j.cclet.2014.05.011.
[10] S. Goswami, A.K. Das, A. Manna, A.K. Maity, P. Saha, C.K. Quah, H.K. Fun, H.A.
26

Abdel-Aziz, Nanomolar detection of hypochlorite by a rhodamine-based chiral
hydrazide in absolute aqueous media: Application in tap water analysis with live-cell
imaging, Anal. Chem. 86 (2014) 6315–6322. https://doi.org/10.1021/ac500418k.
[11] X. Chen, X. Wang, S. Wang, W. Shi, K. Wang, H. Ma, A highly selective and sensitive
fluorescence probe for the hypochlorite anion, Chem. - A Eur. J. 14 (2008) 4719–4724.
https://doi.org/10.1002/chem.200701677.
[12] Y. Long, J. Zhou, M.P. Yang, X.J. Liu, M. Zhang, B.Q. Yang, Highly selective,
sensitive and naked-eye fluorescence probes for the direct detection of hypochlorite
anion and their application in biological environments, Sensors Actuators, B Chem.
232 (2016) 327–335. https://doi.org/10.1016/j.snb.2016.03.157.
[13] Y. Wei, D. Cheng, T. Ren, Y. Li, Z. Zeng, L. Yuan, Design of NIR Chromenylium-
Cyanine Fluorophore Library for “switch-ON” and Ratiometric Detection of Bio-
Active
Species
in
Vivo,
Anal.
Chem.
88
(2016)
1842–1849.
https://doi.org/10.1021/acs.analchem.5b04169.
[14] X. Zhu, L. Yuan, X. Hu, L. Zhang, Y. Liang, S. He, X.-B. Zhang, W. Tan,
Construction of a fluorine substituted chromenylium-cyanine near-infrared fluorophore
for ratiometric sensing, Sensors Actuators B Chem. 259 (2018) 219–225.
https://doi.org/10.1016/j.snb.2017.12.008.
[15] R. Flores-Cruz, R. López-Arteaga, L. Ramírez-Vidal, F. López-Casillas, A. Jiménez-
Sánchez, Unravelling the modus-operandi of chromenylium-cyanine fluorescent
probes: a case study, Phys. Chem. Chem. Phys. 21 (2019) 15779–15786.
https://doi.org/10.1039/C9CP03256H.
[16] X. Jiao, C. Liu, S. He, L. Zhao, X. Zeng, Highly selective and sensitive ratiometric
near-infrared fluorescent probe for real-time detection of Hg2+ and its bioapplications
in
live
cells,
Dye.
Pigment.
160
(2019)
86–92.
27