Development of an off-on selective fluorescent sensor for the detection of Fe3+ ions based on Schiff base and its Hirshfeld surface and DFT studies

Article abstract of DOI:10.1016/j.molliq.2019.111814

Herein, a Schiff base, 4-(2-hydroxybenzylideneamino) benzoic acid (HBBA) was synthesized and characterized by FT-IR, ¹H NMR and single crystal X-ray. The geometrical parameters calculated for the structure prepared using DFT methods were in clo

Full text of DOI:10.1016/j.molliq.2019.111814

Journal Pre-proof
3+
Development of an off-on selective fluorescent sensor for the detection of Fe ions
based on Schiff base and its Hirshfeld surface and DFT studies
Gurpreet Singh, Jayant Sindhu, Manisha, Vinod Kumar, Vishal Sharma, Suresh
Kumar Sharma, Surinder Kumar Mehta, Mater H. Mahnashi, Ahmad Umar, Ramesh
Kataria
PII:
S0167-7322(19)34077-2
DOI:
https://doi.org/10.1016/j.molliq.2019.111814
MOLLIQ 111814
Reference:
To appear in: Journal of Molecular Liquids
Received Date: 21 July 2019
Revised Date: 20 September 2019
Accepted Date: 24 September 2019
Please cite this article as: G. Singh, J. Sindhu, Manisha, V. Kumar, V. Sharma, S.K. Sharma, S.K.
Mehta, M.H. Mahnashi, A. Umar, R. Kataria, Development of an off-on selective fluorescent sensor for
3+
the detection of Fe ions based on Schiff base and its Hirshfeld surface and DFT studies, Journal of
Molecular Liquids (2019), doi: https://doi.org/10.1016/j.molliq.2019.111814.
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Graphical Abstract

Development of an off-on selective fluorescent sensor for the
3
+
detection of Fe ions based on Schiff base and its Hirshfeld
Surface and DFT studies
a,b,#
c,#
d
d
e
Gurpreet Singh , Jayant Sindhu , Manisha , Vinod Kumar , Vishal Sharma ,
b
a
f
Suresh Kumar Sharma , Surinder Kumar Mehta , Mater H. Mahnashi, Ahmad
g
a
Umar * and Ramesh Kataria *
a
Department of Chemistry, Panjab University, Chandigarh-160014, India
b
Department of Chemistry, D. A. V. College, Abohar-152116, India
c
Department of Chemistry, COBS&H, CCS Haryana Agricultural University, Hisar-
1
25004, India
d
Department of Chemistry, C. U. Haryana, Jant-Pali, Mahendergarh-123031, India
Institute of Forensic Science & Criminology, Panjab University, Chandigarh-160014,
e
India
f
Department of Pharmaceutical Chemistry, College of Pharmacy, Najran University,
Najran-11001, Saudi Arabia
g
Department of Chemistry, Faculty of Science and Arts and Promising Centre for
Sensors and Electronic Devices (PCSED), Najran University, Najran-11001, Saudi
Arabia
#
*
Authors contributed equally
Corresponding e-mails: rkataria@pu.ac.in, ahmadumar786@gmail.com

ABSTRACT
Herein, a Schiff base, 4-(2-hydroxybenzylideneamino) benzoic acid (HBBA) was
1
synthesized and characterized by FT-IR, H NMR and single crystal X-ray. The
geometrical parameters calculated for the structure prepared using DFT methods
were in close agreement with the experimentally observed values. Hirshfeld surface
analysis performed on the crystal structure to explore various intramolecular
interactions in the molecule. The band gap and molecular electrostatic potential were
calculated using DFT method in order to support charge transfer. Hyperconjugative
interactions and electron densities of donor (i) and acceptor (j) were explored using
NBO analysis. Photophysical studies revealed the dependency of absorption profile
on the pH of the medium. Thermodynamic properties calculated using DFT method
revealed a correlations of heat capacity (C), entropy (S) and enthalpy changes (H)
3
+
with temperatures. The sensor is specific towards Fe ions with turn-off response
and reversibility in its fluorescence profile upon addition of EDTA. The proposed
3
+
was successfully applied for the real time detection of Fe in water samples with
good accuracy. The limit of detection observed for the developed sensor was lower
than the USEPA guidelines (5.37 μM) for drinking water.
3
+
Keywords: Schiff base, DFT, Electrostatic potential, HOMO, LUMO, Ferric ion (Fe )

1
.
Introduction
Industrial waste has significantly contaminated the environment via different
organic to inorganic pollutants [1-3]. Heavy metals in industrial waste water are
toxic to human and animal life.[2] The removal of these contaminants from water
bodies is necessary to deal with this issue. Functionalised materials have widely
been used to remove these pollutants [4-11]. In addition to this, functionalized
materials have broad range of applications in medical and industrial field.[12-23]
The researcher utilizes the different materials as functional sensing elements owing
to their remarkable optical, mechanical and chemical properties. Materials based on
organic framework received considerable attention due to applications in thin-film
transistors, solar cells, sensors and photorefractive devices.[24] The applications of
sensors based on soft materials also extended to sensing of temperature, stress,
pressure and strain.[25-27] The incorporation of sensing techniques in wearable has
led to the development of multifunctional soft materials for various sensing
techniques.[28-31] The real time monitoring of metal ions using fluorescent sensors
has attracted a great deal of attention. [32-44] Molecular theory is extensively used to
describe chemical behaviour, charge transfer, molecular electronics, photoexcitation
and magnetism associated with a compound. Materials with large HOMO-LUMO
gaps are more stable than those with small gap.[45] Quantum chemical methods can
be used for designing materials that can act as dyes, sensors and molecular
transistors.[46]

3
+
The role of Fe is very crucial in storage and transport of oxygen. [47-49] The
deficiency of iron leads to anemia, low blood pressure and decreased immunity in
human beings and its excess in the body may also equally harmful.[50] The level of
iron should be maintained in human being for proper biological functioning. [51-53]
3
+
Hence, a convenient and rapid method for Fe detection in biological samples is of
great interest. A typical chemosensor contains a receptor/ligand unit, which binds
specific metal ion. A concomitant change in one or more properties of the system,
such as color or fluorescence is the indicator of ligand metal binding due to
complexation, hydrolysis/substitution, or oxidation/ring transformation. A variety of
3
+
efficient chemosensors reported in the literature for the selective detection of Fe .[50,
5
4-59] The selectivity of ligand towards metal ion depends upon the binding affinity
of metal ion. Molecular sensors particularly based on organic chromophores with
colorimetric and fluorescence responses have been used due to their cost-
effectiveness and real time analysis. [60, 61] The ease of synthesis and biological
properties should be considered during the development of molecular sensors.
Recently, a number of research groups have proposed various chemosensors based
on Schiff base and other organic chromophores for the detection of metal ions, such
3+
2+
2+
2+
2+
3+
as Al , Mn , Zn , Cu , Hg and Fe ions.[62-67] Based on the aforementioned facts
and in continuation of our research interests [68-71] we, herein have reported a
3
+
Schiff base ligand (HBBA) as a chemosensor for the selective detection of Fe ions
with turn-off response and reversibility in its fluorescence profile upon addition of
3
+
EDTA. The developed sensor is highly sensitive and selective for Fe ions and

3
+
applied for the real time detection of Fe in water samples with a good detection
limit.
2
. Experimental details
2
.1. Materials and instrumentation
All chemicals were obtained from commercial suppliers and used without further
1
purification. H NMR spectra were measured on the BRUKER AVANCE II 400 NMR
spectrometer using TMS as an internal standard. IR spectra were recorded on a
-1
Nicolet IS-50 FTIR spectrometer with absorption reported in cm . UV-Vis absorption
spectra were determined on a Hitachi UH-5300 spectrophotometer. Fluorescence
spectra were recorded on a Hitachi F-7000 spectrophotometer equipped with quartz
cuvettes of 1 cm path length.
2
.2. Synthesis of HBBA
A mixture of 4-aminobenzoic acid (0.9780 g, 8 mmol) and salicylaldehyde (1.096 g, 8
mmol) was dissolved in CH OH (10 mL) in a 50 mL round bottom flask. The content
3
of the flask was stirred at room temperature for 4 h. The progress of the reaction was
monitored using TLC, the yellow precipitates obtained on completion of the reaction
1
was filtered and recrystallized using ethanol to afford HBBA.[72] HNMR (400MHz,
DMSO-d
6
): δ 13.00 (s, 1H, H11), 12.72 (s, 1H, H10), 9.00 (s, 1H, H ), 8.02 (d, 2H, H9+6),
5
7
.71 (m, 1H, H
4
), 7.49 (d, 2H, H7+8), 7.45 (m, 1H, H ), 6.99 (m, 2H, H1+3).
2

2
.3. X-ray crystallography
Crystals of HBBA suitable for single crystal were selected and mounted on a
SuperNova, single source at offset/far, HyPix3000 diffractometer. Cryo loop is used
to mount the crystal. The crystals were kept at 293(2) K during data collection. The
crystal structure was solved using Olex2[73]. Direct method was used in with
SIR2004 [74] and refinement was done by ShelXL [75] refinement package using
Least Squares minimization.
2
.4. Solution preparation
Stock solutions of Schiff base (HBBA) with a concentration of 1 mM were prepared
-1
by dissolving 0.241g L in MeOH. The stock solution was further diluted to prepare
-5
-6
solutions of 10 M and 10 M concentration for photophysical studies. The solution of
metal ions of concentration 10 mM was prepared by dissolving respective amounts
of corresponding salts in distilled water.
2
.5. Computational details
The Density functional theory (DFT) with hybrid density function B3LYP
(Becke-Lee-Young-Parr composite of exchange-correction functional) method and
the standard 6-311G++(d,p) basis set were used for structure optimization for
HBBA.[76] All theoretical calculations were performed using Gaussian 09 program
package.[77] Gauss view and Avogadro 2.0 were used for the visualization of results

obtained from Gaussian 09 program.[78] Initially, geometry generated from
geometrical parameters was minimized without any constraint in the potential
energy surface at DFT level. The optimized structural parameters were used in the
vibrational wavenumber calculations at the DFT level to characterize all stationary
points as minima. The vibrational modes were assigned based on potential energy
distribution (PED) using VEDA4 program. [79] Hirshfeld surface maps and
fingerprint plots were generated from the crystallographic data using the Crystal
Explorer 3.1.[80] Natural bond orbital (NBO) analysis was done using NBO 3.1
implemented in Gaussian 09 package. The NBO analysis was performed to explore
various second-order interactions between two subsystems. The second-order
interactions are a measure of intermolecular and intramolecular delocalization.
3
. Results and discussion
3
.1. Synthesis and characterizations
The compound 4-(2-hydroxybenzylideneamino)benzoic acid (HBBA) was prepared
by condensation of 4-aminobenzoic acid (0.9780 g, 8 mmol) and salicylaldehyde
(
1.096 g, 8 mmol) using CH OH (10 mL) as a solvent in a 50 mL round bottom
3
flask.[81] The reaction was attempted at room temperature for 4 h and resulted
HBBA in 85% yield (Scheme 1). The structure of HBBA was confirmed by using FT-
1
IR, H NMR and by Single Crystal X-ray analysis.

Scheme 1: Synthesis of 4-(2-hydroxybenzylideneamino)benzoic acid (HBBA)
3
.2. Molecular structure and geometry
The single crystals were obtained from the methanolic solution of HBBA. The
single crystal X-ray analysis reveals that HBBA crystalizes in monoclinic cell system
with P21/c space group. The CCDC search details showed that the crystal structure is
already reported by Uekusa and co-workers.[81] The crystal structure of HBBA is
crystallizes as monomer as depicted in Figure 1a. The carboxylic group of HBBA
…
…
…
forms hydrogen-bonded dimer structure via O
1
H1
O (see Figure 1b) with  D-H A
2
of 167° as suggested by PLATON.[82] These 1D dimeric units are further connected
to each other through the various types of hydrogen bonding interactions and finally
adapted 2D hydrogen bonding network (Figure 1c). The details of the structure
refinement and hydrogen bonding interactions are compiled in Table S1 and S2 of
the supporting information. However, the selected bond lengths and angles are
listed in Table 1.
The molecular structure for HBBA was optimized by B3LYP/6-311++G(d,p)
method and was shown in Figure 2. The computed optimized structure for both
monomer and dimer showed C1 point group symmetry with ground state energy of
-820.79 and -1641.61 a.u and dipole moment of 4.0039 and 4.5810 Debye respectively.

The computed geometrical parameters for monomer and dimer along with the
experimental ones obtained from XRD were listed in Table 1. The recorded XRD
value for C14-O showed a bond distance of 1.259 Å, however the computed bond
2
distance for monomer and dimer was 1.209 and 1.230 Å respectively. The higher
value of bond length obtained from XRD data might be attributed to the resonance
effect of the carboxylate moiety. On the other hand C
7
-N has a double bond
1
character as evident form computed bond length of 1.291 and 1.289 Å for monomer
and dimer respectively. The computed bond length for dimer is in close agreement
with the experimentally observed values of 1.274 Å. In case of dimer the dihedral
o
angle (C
8
-N
1
-C
7
-C
6
) is 176.9 , while in case of monomer the dihedral angle (C
8
-N
1
-C -
7
o
o
C
6
) is 180.0 which is in close agreement with the XRD obtained value of 179.4 which
supports the planarity of two phenyl rings.

Figure 1: Molecular structure of HBBA (a) Monomer; (b) Dimer; (c) 2D-hydrogen
bonded network structure along crystallographic a-axis.
The computed O
dimer it was found to be 0.991 Å which is in close agreement with the
experimentally determined value of 0.983 Å. The computed bond length of O -H for
dimer and monomer was found to be 0.968 and 0.999 Å respectively. The
experimentally observed bond length for O -H bond was 0.983 Å which reveals
3
-H3A bond distance for monomer was 0.994 Å, however for
1
1
1
1
close agreement of experimentally determined value with computed bond length in
case of dimer. It has been revealed from above that the structure is a hydrogen
bonded dimer even in the solid phase. The computed C-H bond length in case of
aromatic system varies from 1.082 Å to 1.085 Å which is also in close agreement with
the experimentally determined value of 1.083 Å. The small deviation in the

computed bond length for C-H bond might be attributed due to the low scattering
factor of hydrogen atoms involved in the X-ray diffraction experiment. In addition,
the geometrical data for most of the bond lengths calculated using DFT/B3LYP
method are higher than the experimental values obtained from experimental XRD
results owing to the fact that XRD were performed in solid state however the
theoretical calculations were carried out on isolated molecules in gas phase. The
crystal and refinement data for the HBBA has been provided in Table S1.
Figure 2: Optimized structure of HBBA (a) Monomer; (b) Dimer.
Table 1: Geometrical parameters for HBBA in its monomeric and dimeric form
Bond length/ Bond angle Experimental
Å]
Monomer
Calculated [Å]
1.338
1.422
1.400
1.386
1.402
1.382
Dimer
Calculated [Å]
1.340
1.421
1.400
1.386
1.402
1.383
[
C(1)-O(3)
C(1)-C(6)
C(1)-C(2)
C(2)-C(3)
C(3)-C(4)
C(4)-C(5)
1.340(2)
1.392(2)
1.396(2)
1.356(3)
1.374(3)
1.371(2)

C(5)-C(6)
C(6)-C(7)
C(7)-N(1)
C(8)-C(13)
C(8)-C(9)
C(8)-N(1)
C(9)-C(10)
C(10)-C(11)
C(11)-C(12)
C(11)-C(14)
C(12)-C(13)
C(14)-O(2)
C(14)-O(1)
1.385(2)
1.451(2)
1.268(2)
1.373(2)
1.403(2)
1.421(19)
1.378(2)
1.381(2)
1.381(2)
1.485(2)
1.388(2)
1.260(2)
1.2674(19)
1.409
1.445
1.291
1.404
1.406
1.406
1.386
1.401
1.399
1.483
1.386
1.209
1.359
1.408
1.446
1.289
1.403
1.404
1.403
1.388
1.399
1.401
1.482
1.385
1.230
1.323
O(3)-C(1)-C(6)
O(3)-C(1)-C(2)
C(6)-C(1)-C(2)
C(3)-C(2)-C(1)
C(2)-C(3)-C(4)
C(5)-C(4)-C(3)
C(4)-C(5)-C(6)
C(5)-C(6)-C(1)
C(5)-C(6)-C(7)
C(1)-C(6)-C(7)
N(1)-C(7)-C(6)
C(13)-C(8)-C(9)
C(13)-C(8)-N(1)
C(9)-C(8)-N(1)
C(10)-C(9)-C(8)
C(9)-C(10)-C(11)
C(12)-C(11)-C(10)
C(12)-C(11)-C(14)
C(10)-C(11)-C(14)
C(11)-C(12)-C(13)
C(8)-C(13)-C(12)
O(2)-C(14)-O(1)
O(2)-C(14)-C(11)
O(1)-C(14)-C(11)
C(7)-N(1)-C(8)
121.18(15)
119.54(17)
119.28(18)
120.54(19)
121.05(18)
118.70(19)
122.07(19)
118.34(15)
120.35(16)
121.31(16)
122.69(17)
119.19(14)
117.32(14)
123.49(15)
119.58(16)
120.92(16)
119.56(14)
120.69(15)
119.75(15)
119.83(16)
120.91(15)
123.57(14)
118.97(15)
117.46(15)
122.05(14)
121.8
118.7
119.4
120.2
121.0
119.0
121.3
118.8
119.6
121.5
121.1
118.5
116.0
125.3
121.9
118.5
119.4
120.2
121.0
119.0
121.3
118.8
119.6
121.4
122.6
119.1
118.1
122.6
120.3
120.4
119.3
121.5
119.0
120.3
120.3
179.9
122.2
114.4
121.1
120.4
120.5
119.2
118.4
122.3
120.2
120.9
121.8
125.0
113.0
123.3
3
.3. Hirshfeld surface analysis
The Hirshfeld surfaces analysis for HBBA was performed using Crystal
explorer 3.1 software. The Hirshfeld surface of a molecule is mapped using the

descriptor dnorm which includes two factors: d
e
and d . The distance of any surface
i
points nearest to the internal atoms and exterior atoms are represented by d
e
and d
i
respectively.[83] The intermolecular interaction of the compound HBBA can be
explored using this study from valuable graphical images. The Hirshfeld surfaces
are over dnorm (-0.745 to 1.155 Å), d
i
(0.645 to 2.491 Å), d (0.647 to 2.372 Å), shape
e
index (-0.991 to 0.993 Å) and curvedness (-4.327 to 0.178 Å). In Hirshfeld surfaces of
the HBBA, there are OH hydrogen boding interactions which are visualized as red
spots referring (dnorm) in Figure 3. The presence of highlighted red and blue triangles
in the shape index revealed the existence of π-π interaction in the crystal structure of
the title compound. In addition, the carbon atoms which are located outside the
surface are represented by red triangles while those located inside the surface are
represented by blue color triangles.
Figure 3: Hirshfeld surfaces mapped over (a) d
i
; (b) d ; (c) dnorm; (d) shape index and
e
(e) curvedness for HBBA.
Other visible spots in the Hirshfeld surfaces correspond to C---H and H---H
contacts. The intermolecular interactions of the title compound are shown in the 2D
fingerprint plots shown in Figure 4. H---H (41.6%) contacts make the largest

contribution to the Hirshfeld surfaces. O---H/H---O (22.3%), interactions are
represented by left-side blue spikes, top and bottom. The purple C---C/C---C (8.1%)
interactions are in between the C---H regions while the green C---H/H---C
interactions (18.9%) are outside of H---H regions.
Figure 4: Two-dimensional fingerprint plots of HBBA
3
.4. Vibrational analysis
The number of fundamental vibrations which are potentially active for a non-
linear molecule is equal to (3N-6), where N is the number of atoms. The total number
of atoms present in compound HBBA is 29, which has 81 normal modes of
vibrations. The vibrational bands calculated for the synthesized compound were
assigned by with the help of potential energy distribution (PED) using VEDA
program [84]. The computed vibrational frequencies (Unscaled and Scaled) along
with the experimentally determined values are tabulated in Table S3.

3
.5. Molecular electrostatic potential (MEP)
The reactive sites for the electrophilic and nucleophilic attack in a chemical
reaction can be predicted using the molecular electrostatic potential (MEP) which
further helps in the recognition of hydrogen bonding sites. The molecular properties
such as electronegativity, dipole moment, partial charges of molecules and chemical
reactivity are related to total charge densities and hence can also be predicted using
MEP. The different values of electrostatic potential over the surface of the molecule
are represented by different colors (Figure 5). The region of negative electron density
is by red color, while the positive is represented by blue color, and the white color
indicates the zero potential region. The localization of red colored sites mainly on
oxygen and nitrogen represents the electron-acceptor behavior of the both these
atoms in the hydrogen bonding. Thereby, the oxygen atoms served as an electron-
acceptor in the title compound both intramolecular and intermolecular hydrogen
bonding (Figure 5).

Figure 5: Molecular electrostatic potential diagram for HBBA.
3
.6. Electrostatic results
The chemical stability of a molecule can be best explained on the basis
quantum parameters such as highest occupied molecular orbital (HOMO) and
lowest unoccupied molecular orbital (LUMO).[85] The optical and electrical
properties can be explored using these molecular orbitals. In order to explore these
properties the HOMO and LUMO energies of the compound (3) were calculated at
B3LYP/6-311++G(d, p) level in gas phase and 3D plots of the HOMO and LUMO
were illustrated in Figure 6. The calculated energies of the HOMO and LUMO for
the compound (3) were found to be -6.40 eV and -2.69 eV respectively. The HOMO
of the studied compound is localized on the -OH substituted benzene, while LUMO
is concentrated more towards benzene rings substituted with -COOH group. In
addition, both of the HOMO and LUMO frontier orbitals are localized on
azomethine group. As shown in Figure 6, the energy gap (∆E) between the HOMO

and LUMO was determined as -2.45 eV. According to Koopmans theorem the
ionization energy (I) and electron affinity (A) can be best explained by using EHOMO
and ELUMO values as I = -EHOMO and A = -ELUMO. The chemical hardness is associated
with the stability and reactivity of a system and measures the resistance to change in
the electron distribution of a molecule.[86] It is well known that molecular stability
and reactivity are characterized by other important parameters, the electronic
chemical potential (μ) and global hardness (η), global electrophilicity index (ω) and
softness (s) which are also related to EHOMO and ELUMO. The low chemical hardness
(1.85 eV) with high negative value of chemical potential (-4.54 eV) indicates that
HBBA is a soft molecule with high polarizability. The high electrophilicity (5.55) of
HBBA indicates its high power to withdraw electrons and hence to act as an
electrophile. The obtained global reactivity data are given in Table 2.
Table 2: Calculated quantum chemical parameters for HBBA.
HOMO LUMO Ionization
Electron
affinity
Chemical
potential
(μ)a
Global
Hardness
(η)a
Global
Softness
(s)a
(
eV)
(eV)
energy (I)
electrophilicity
index (ω)a
5.55
(
A)
-
6.40
-2.69
6.40
2.69
-4.54
1.85
0.54
a
2
μ=1/2(EHOMO+ELUMO); η = -1/2(EHOMO – ELUMO); ω= μ /2η; s = 1/η

Figure 6: Selected frontier molecular orbitals of compound HBBA with energy gap.
The number of states in a unit energy interval can be best represented by
density-of-states (DOS) as proposed in solid physics. The values derived from
density-of-states provide valuable information regarding the distribution, occupancy
and transition rates in a state upon absorbing and emitting light. The total density of
states (TDOS) analysis of HBBA was performed at B3LYP/6-311++G(d, p) level of
theory in the gas phase. The DOS spectra were created by convoluting the molecular
orbital information from Gaussian curves of unit height and Full width at half
maximum (FWHM) of 0.3 eV using the Gauss Sum 2.2 program. [87] The DOS
analysis curve for compound HBBA is shown in Figure S1 (see ESI). It can be
observed from Figure S1 that the maximum density of states lies over occupied
orbitals with energy range of -6.3 eV to -19.84 eV.

Fukui functions are the most important local reactivity descriptors with a
significant role in the prediction of reactivity and selectivity. These are the derivative
of electron density with respect to variation in the number of electrons without
changing nuclei position. Despite these variations, Fukui functions also give
information about the distortion of electron density at a given position upon
accepting or donating electrons. Fukui functions in terms of condensed Fukui
functions have been calculated for HBBA in order to explore the positions prone to
undergo nucleophilic, electrophilic and radical attack within the molecule. The
condensed Fukui function for nucleophilic, electrophilic and radical attack can be
ꢂ
ꢃ
ꢈ
expressed as ꢀ , ꢀ ꢄꢅꢆꢇꢄꢀ ꢄrespectively. The value related with these functions can
ꢁ
ꢁ
ꢁ
be assessed as mentioned below: -
ꢂ
ꢁ
ꢁ
ꢀ
= ꢉ ꢊꢋ + 1ꢌ − ꢉ ꢊꢋꢌ
ꢁ
ꢃ
ꢁ
ꢁ
ꢀ
= ꢉ ꢊꢋꢌ − ꢉ ꢊꢋ − 1ꢌ
ꢁ
ꢈ
ꢁ
ꢁ
ꢀ
= ꢊꢉ ꢊꢋ + 1ꢌ − ꢉ ꢊꢋ − 1ꢌꢌ/2ꢄ
ꢁ
th
The electronic population at k atom is represented by qk in the neutral (N), anionic
(N+1) or cationic (N-1) chemical species. The electronic population used in this
study was calculated using NBO analysis where natural population analysis (NPA)
was also done in order to access the charge present on each atom. A new dual
descriptor f(r) was proposed by Morrel et al. to differentiate the site for nucleophilic
and electrophilic attack present within a molecule which is defined by the equation.
ꢂ
ꢃ
ꢍꢀꢊꢎꢌ = ꢊꢀ − ꢀ ꢌ
ꢁ
ꢁ

The atomic site with ꢄꢍꢀꢊꢎꢌ > 0 is prone to electrophilic attack, while a site
favourable for nucleophilic attack should haveꢄꢍꢀꢊꢎꢌ < 0. The value of Fukui
functions calculated for HBBA are summarised in
Table 3.
Table 3: Condensed Fukui function for HBBA
k
k
k
-
+
0
Atom
q (N)
q (N-1) q (N+1) f
k
f
k
f
k
Δf
C
3
4
5
6
1
2
7
-0.076
-0.127
-0.067
-0.093
0.190
-0.128
0.084
-0.271
0.080
-0.111
-0.074
-0.085
-0.071
-0.094
-0.339
0.393
-0.192
-0.102
-0.174
-0.006
0.110
-0.123
-0.162
-0.316
0.016
-0.166
-0.080
-0.173
-0.120
-0.113
-0.370
0.350
-0.069
-0.158
-0.032
-0.132
0.153
-0.124
0.106
-0.308
0.064
-0.129
-0.055
-0.111
-0.050
-0.113
-0.358
0.392
0.115
-0.025
0.107
-0.086
0.080
-0.005
0.246
0.046
0.064
0.054
0.006
0.088
0.048
0.019
0.031
0.044
0.063
0.016
0.008
-0.032
0.034
-0.039
-0.038
0.004
0.022
-0.037
-0.016
-0.018
0.019
-0.026
0.021
-0.018
-0.019
-0.001
0.009
0.007
0.061
-0.028
0.071
-0.063
0.021
-0.001
0.134
0.004
0.024
0.018
0.012
0.031
0.035
0.000
0.006
0.021
0.036
0.011
-0.108
-0.007
-0.073
0.048
-0.118
0.009
-0.224
-0.083
-0.081
-0.072
0.014
-0.113
-0.027
-0.037
-0.050
-0.045
-0.054
-0.010
C
C
C
C
C
C
N
1
C
C
C
C
C
C
O
C
O
O
8
13
12
11
10
9
3
14
2
1
-0.300
-0.346
-0.363
-0.363
-0.291
-0.340
3
.7. Natural bond orbital (NBO) analysis
Hyper-conjugation interactions and electron transfer between two subsystems
can be effectively analyzed using DFT calculations for natural bond orbital (NBO).
The extent of delocalization and hyperconjugation interactions can be quantified by
various second order interactions between the filled and vacant orbitals of a system.
The hyperconjugation interaction energy can be obtained from the second order
perturbation method as follows:

ꢐ
<
ꢒ ∣ ꢓ ∣ ꢒ >
ꢓ
ꢕꢖ
ꢏꢐ = −ꢆꢑ
= ꢆꢒ
∗
ꢔꢒ − ꢔꢒ
∆ꢏ
where q
i
is the donor orbital occupancy, ε
i
and ε are the diagonal elements
j
and F (i, j) is the off diagonal NBO Fock matrix element. The various possible donors
and acceptors in the molecule along with their occupancy can be localized using
NBO analysis. In addition to this, NBO analysis also provides various possible
transitions among these donors and acceptors. The intensity of interaction between
these donor and acceptors groups depends upon the value of E . The larger the
2
values the more intense will be the interaction and hence greater will be the extent of
conjugation in the system. Full NBO analysis and second order Fock matrix
perturbation theory analysis was carried for molecule under study using B3LYP/6-
3
11++G(d, p) level of theory. The results of complete NBO analysis with the
percentage of the NBO on each hybrid, and a hybrid label showing the hybrid orbital
x
(
sp ) composition along with second-order perturbation theory analysis of Fock-
matrix in NBO basis for compound HBBA are shown in Table S4 and Table S5
respectively (see ESI). It has been revealed from second order Fock matrix that
strong intramolecular hyperconjugative interactions exist in compound which
results in stabilization of the system.
The intramolecular hyper-conjugative interaction of the π(C
1
-C ) distribute to
6
π*(C
2
-C
3
) and π*(C
4
-C ) and leads to stabilization of 16.06 and 23.79 Kcal/mol
5
respectively. The interaction of π(C
2
-C
3
) to π*(C
1
-C
6
) and (C
4
-C ) leads to stabilization
5
of 21.78 and 14.95 kcal/mol respectively. The distribution of π(C
4
-C
5
) to σ*(C
1
-C ),
6

π*(C
2
-C
3
) and π*(C
7
-N ) provides stabilization of 14.07, 22.19 and 23.71 Kcal/mol
8
respectively. There is a strong interaction between σ(C -O15) and σ*(C17-O19) with a
5
stabilization energy of 154.85 Kcal/mol. The conjugation of π(C
9
-C14) to σ*(C
7
-N ),
8
π*(C10-C11) and π*(C12-C13) leads to stabilization of 14.52, 17.52 and 23.8 Kcal/mol
respectively. This strong conjugation provides stability to the C=N linkage in HBBA.
The π electron delocalization of π(C10-C11) with anti-bonding π*(C
9
-C14) and σ*(C12-
C
13) leads to stabilization of 21.24 and 17.18 Kcal/mol respectively as shown in Table
S5.
This enhanced π*(C
4
-C
5
) and π*(C
1
-C
6
) NBO further conjugates with π*(C
2
-C ),
3
resulting in an enormous stabilization energy of 229.99 and 210.77 kcal/mol.
Similarly, the NBO π*(C -N ) and π*(C -C ) further conjugates with π*(C -C14) leads
7
8
4
5
9
to stabilization of 123.38 and 50.03 Kcal/mol respectively as shown in Table S4 (see
ESI). A strong intramolecular hyperconjugative interaction occurs from (LP) O15 to
σ*(C17-O19), π*(C
4
-C
72.05 kcal/mol, respectively. All other less intense interactions have been tabulated
in Table S5.
5
) and σ*(C17-O19), leading to stabilization of 123.52, 37.81 and
1
3
.8. Photo-physical studies
The spectral characteristics of HBBA such as absorption maxima (λmax),
emission maxima (λem) and stoke shift were measured in different solvents of
varying polarity to study the effect of solvent polarity on the spectral characteristic
of HBBA. HBBA shows two absorption band in the region of 280-295 nm and 340-
3
50 nm in all solvents. The absorption spectrum for HBBA in different solvents is

shown in Figure 7A. Furthermore, the fluorescence spectrum for HBBA was also
measured in different solvents like CH CN, benzene, CHCl , DCM, DMF, DMSO,
EtOAc, EtOH, H O and MeOH. It has been found that the absorption and emission
3
3
2
spectrum of HBBA was not affected much by solvent polarity which indicates that
the ground and excited state of the molecule are quite same. The emission spectrum
for HBBA in different solvents is shown in Figure 7B. The stoke shift and absorption
and emission maxima for HBBA is shown in Table 4.
The absorption spectrum for compound HBBA was further investigated for
the change in pH of the medium. The absorption spectrum for the compound was
studied by the addition of base and acid in the MeOH solution of the compound.
The decline in the absorption intensity observed for band at 340-350 nm was more
dramatic than that observed for a band at 280-295 nm with increasing acid
concentration. An overall decrease in the absorption intensity was observed with
increasing acid concentration. The effect of HCl (0.1 M) concentration on the
absorption spectrum of compound HBBA is shown in Figure 8A.

-5
Figure 7: (A) Absorption; (B) Emission spectrum for compound HBBA (1 x10 M for
-6
absorbance and 1 x10 M for emission spectra) in different solvents of varying
polarity.

Table 4: Absorption maxima (λmax), emission maxima (λem) and stoke shift for HBBA
in different solvents.
-
1
(cm-1)
) (cm-1) (υ
) (cm-1)
+υ
)/2 (cm-1)
S.No Solvent
υ
f
(cm )
υ
a
(υ
a
- υ
f
a
+υ
f
(υ
a
f
1
2
3
4
5
6
7
8
9
1
MeOH
EtOAc
DCM
29515.94
29850.75
30284.68
29655.99
29002.32
37174.72
36563.07
35842.29
34129.69
33670.03
35211.27
35842.29
34188.03
31055.90
30959.75
7658.78
6712.32
5557.61
4473.70
4667.71
4611.51
6062.66
4390.65
2103.96
1685.75
66690.66
29850.75
66126.97
63785.68
33670.03
65811.03
65621.92
63985.41
60007.84
60233.75
33345.33
14925.38
33063.49
31892.84
16835.02
32905.52
32810.96
31992.71
30003.92
30116.88
DMF
DMSO
Benzene 30599.76
CH
3
Cl
29779.63
29797.38
28951.94
29274.00
ACN
EtOH
0
H2O
Further, the absorption spectrum of compound HBBA was also investigated
at increasing concentration of NaOH. Ratiometric behavior with a bathochromic
shift of 50 nm was observed when the absorption profile for compound HBBA was
investigated at increasing NaOH (0.1 M) concentration. The effect of increasing HCl
and NaOH concentration on the absorption profile of compound HBBA is shown in
Figure 8B.

Figure 8: (A) Effect of increasing HCl and (B) NaOH concentration on the absorption
-5
spectrum for compound HBBA (1 x 10 M).

In order to investigate the chemosensor behavior of compound HBBA, its
fluorescence spectrum was explored by mixing it with thirteen different metal ions
2
+
2+
2+
2+
+3
2+
2+
2+
2+
2+
2+
2+
(
10 equiv.) namely Ba , Ca , Cd , Co , Cr , Cu , Fe , Mg , Mn , Ni , Pb , Zn
3
+
and Fe in methanol. Methanol was chosen as a solvent for metal ion investigation
N
because of its high polarizaibility (E
T
) and hydrogen bond donor acidity (α or SA)
value. It can easily stabilize the charge separation, which makes it a solvent of
choice. The effect of various metal ions on fluorescence spectra of HBBA is shown in
Figure 9A. It can be easily inferred from Figure 9A that chemosensor HBBA
exhibited excellent fluorescence in absence of metal ions in methanol. Addition of
2
+
2+
2+
2+
+3
2+
2+
2+
2+
2+
2+
2+
metal ions Ba , Ca , Cd , Co , Cr , Cu , Fe , Mg , Mn , Ni , Pb and Zn (10
3
+
equiv.) did not affect the fluorescence spectra of HBBA. However, addition of Fe
(10 equiv.) affected the fluorescence spectra dramatically and resulted in quenching
of fluorescence. The fluorescence intensity of HBBA was decreased by more than 10
fold like turn off sensing response (Figure 9A).

Figure 9: (A) Effect of various metal ion on the fluorescence spectrum of compound
-7
3+
HBBA (1 x 10 M); (B) Effect of different concentration of Fe ions on the
-7
fluorescence spectrum of compound HBBA (1 x10 M)