Synthesis, characterization, theoretical calculations and biochemical evaluation of a novel oxime ligand with complexes

Article abstract of DOI:10.1016/j.molliq.2019.03.169

A novel amine containing ketooxime ligand (HBOX) and its Cu(II) and Mn(II) complexes were synthesized, characterized and tested for some of their biological activities. Structural characterization was carried out by elemental analysis, ICP-OES, ¹H and ¹³C NMR, UV–Vis, FT-IR, XRD, TG-DTG, magnetic susceptibility and molar conductivity measurements. Elemental analyses, stoichiometric and spectroscopic data of the metal complexes indicated that the metal:ligand ratio was found to be 1:2 and the metal ions were coordinated to the oxime oxygen and amine nitrogen atoms. Furthermore, DFT/B3LYP method with 6-311G(d,p) and LANL2DZ basis sets were used for full optimization of molecular geometries of the ligand and complexes, respectively. The vibrational frequencies, isotropic chemical shifts (¹H and ¹³C NMR), electronic transition absorption wavelengths, HOMO and LUMO analyses and molecular electrostatic potential (MEP) properties of the synthesized molecules have been calculated. The results obtained experimentally were confirmed by the theoretical data which are in good agreement. Inhibitory capacity of the HBOX was investigated against neoangiogenic factors, vascular endothelial growth factor receptor-2 (VEGFR-2) and cyclooxygenase-2 (COX-2) by molecular docking studies. HBOX bound to COX-2 protein with 2 hydrogen bonds at the lowest energy level which indicates the most stabilized form of the protein ligand complex. Complexes were also tested for their catecholase and phenoxazinone synthase-like activities using spectrophotometric procedures. Catecholase and phenoxazinone synthase-like enzyme activities were spectrophotometrically followed by the increase in absorbance at 400 and 433 nm resulted from the oxidation reaction of 3,5-di-tert-butylcatechol and 2-aminophenol to 3,5-di-tert-butylquinone and 2-aminophenoxazine-3-one, respectively. According to the calculated k_obs values, the Mn(II) complex was found to be more active for both enzymes compared to Cu(II).

Full text of DOI:10.1016/j.molliq.2019.03.169

Accepted Manuscript
Synthesis, characterization, theoretical calculations and
biochemical evaluation of a novel oxime ligand with complexes
Güvenç Görgülü, Bülent Dede
PII:
S0167-7322(19)30664-6
DOI:
https://doi.org/10.1016/j.molliq.2019.03.169
MOLLIQ 10710
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Accepted date:
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11 March 2019
29 March 2019
Please cite this article as: G. Görgülü and B. Dede, Synthesis, characterization, theoretical
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Date
: 03 Februaty 2019
Author Name : Dr. Bülent DEDE
University
Address
: Süleyman Demirel University
: Süleyman Demirel University Faculty of Arts & Sciences, Department of Chemistry 32260
Isparta TURKEY
Tel
: +90 246 2114153
E-mail
: bulentdede@sdu.edu.tr
I am enclosing herewith a manuscript entitled “SYNTHESIS, CHARACTERIZATION, THEORETICAL
CALCULATIONS AND BIOCHEMICAL EVALUATION OF A NOVEL OXIME LIGAND WITH COMPLEXES” for
possible evaluation in “JOURNAL OF MOLECULAR LIQUIDS”. We expect, our findings about molecular
docking studies, enzymatic activities and DFT calculations of a novel aminoketooxime ligand and its
Mn(II) and Cu(II) complexes are likely to be of great interest to the vision scientists, researchers, and
trainees who read your journal. The corresponding author of this manuscript is (Dr. Bülent DEDE) and co-
author has been mentioned below according to their contribution.
1.
Dr. Güvenç GÖRGÜLÜ
With the submission of this manuscript, I would like to undertake that:

All authors of this research paper have directly participated in the planning, execution & analysis
of this study.




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
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Dr. Bülent DEDE
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Corresponding Author)
Potential Reviewers
1. Dr. Tanurima Bhaumik e-mail: tanurimabhaumik@gmail.com
Department of Chemistry, Jadavpur University, Kolkata, India
2. Dr. Zbigniew Kałuża e-mail: zbigniew.kaluza@icho.edu.pl
Institute of Organic Chemistry Polish Academy of Sciences, Warsaw, Poland
3. Dr. Jan Veselý e-mail: jxvesely@natur.cuni.cz
Department of Organic Chemistry, Charles University, Prague, Czech Republic
1

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SYNTHESIS, CHARACTERIZATION, THEORETICAL
CALCULATIONS AND BIOCHEMICAL EVALUATION OF A NOVEL
OXIME LIGAND WITH COMPLEXES
1
2
Güvenç GÖRGÜLÜ , Bülent DEDE *
1
Mehmet Akif Ersoy University, Faculty of Education, Department of Science Education,
Burdur, Turkey
2
Süleyman Demirel University, Faculty of Sciences and Arts, Department of Chemistry,
Isparta, Turkey
*
e-mail: bulentdede@sdu.edu.tr
Tel: +90 246 2114153
2

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ABSTRACT
A novel amine containing ketooxime ligand (HBOX) and its Cu(II) and Mn(II) complexes
were synthesized, characterized and tested for some of their biological activities. Structural
1
13
characterization was carried out by elemental analysis, ICP-OES, H- and C-NMR, UV-Vis,
FT-IR, XRD, TG-DTG, magnetic susceptibility and molar conductivity measurements.
Elemental analyses, stoichiometric and spectroscopic data of the metal complexes indicated
that the metal:ligand ratio was found to be 1:2 and the metal ions were coordinated to the
oxime oxygen and amine nitrogen atoms. Furthermore, DFT/B3LYP method with 6-
3
11G(d,p) and LANL2DZ basis sets were used for full optimization of molecular geometries
of the ligand and complexes, respectively. The vibrational frequencies, isotropic chemical
1
13
shifts ( H- and C-NMR), electronic transition absorption wavelengths, HOMO and LUMO
analyses and molecular electrostatic potential (MEP) properties of the synthesized molecules
have been calculated. The results obtained experimentally were confirmed by the theoretical
data which are in good agreement. Inhibitory capacity of the HBOX was investigated against
neoangiogenic factors, vascular endothelial growth factor receptor-2 (VEGFR-2) and
cyclooxygenase-2 (COX-2) by molecular docking studies. HBOX bound to 1CX2 protein
with 2 hydrogen bonds at the lowest energy level which indicates the most stabilized form of
the protein ligand complex. Complexes were also tested for their catecholase and
phenoxazinone synthase-like activities using spectrophotometric procedures. Catecholase and
phenoxazinone synthase-like enzyme activities were spectrophotometrically followed by the
increase in absorbance at 400 and 433 nm resulted from the oxidation reaction of 3,5-di-tert-
butylcatechol and 2-aminophenol to 3,5-di-tert-butylquinone and 2-aminophenoxazine-3-one,
respectively. According to the calculated kobs values, the Mn(II) complex was found to be
more active for both enzymes compared to Cu(II).
3

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Keywords: Oxime, metal complexes, DFT, molecular docking, catecholase-like activity,
phenoxazinone synthase-like activity
4

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1
. INTRODUCTION
The stability and function of a metal complex basically depend on its design and synthesis,
besides, the electronic configuration of added metal ion(s) and electron delocalization of
added functional groups. Since the existence of an oxime group in the ligand increase
stability, strength and biological activity of the complex, they used extensively in industry,
medicine and environmental protection [1]. Illuminating the structures of these ligands and
metal complexes, therefore, become important to understand their functioning related to
configuration [2]. Besides spectroscopic and analytical techniques, DFT gives valuable
information on structural, energetic, molecular, electronic, vibrational and magnetic behaviour
which makes an important contribution for the characterization of the interested compound
[3].
In addition to their antioxidant, antibacterial, antifungal and antiviral properties, oxime
compounds are also used as potential agents for the inhibition of tumour growth [4-11]. It is
still critically important to inhibit molecules increasing in pathological conditions like chronic
inflammation, tumour growth and metastasis [12]. Pathological angiogenesis plays a key role
in tumour growth and metastasis with a cascade mechanism [13]. Vascular endothelial growth
factor-2 (VEGFR-2) is often used as a parameter for being a potent metastatic factor due to its
angiogenic and lymphangiogenic effects [14]. It also promotes the angiogenic process by
inducing cyclooxygenase (COX-2) and inducible nitric oxide synthase (iNOS) [15,16].
Molecular docking research focuses on computationally simulating the molecular
recognition process. It aims to achieve an optimized conformation with the best orientation
between the target protein and ligand such that the free energy of the overall system is
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minimized [17]. The application of docking in a targeted drug-delivery system is a huge
benefit. One can study the size, shape, charge distribution, polarity, hydrogen bonding, and
hydrophobic interactions of both the ligand (prospective drug) and the receptor (target site)
[18].
It is not possible to use natural enzymes as drugs due to their instability in solution and
delivery problems, while metal complexes are better candidates so far for medical uses [19-
2
2]. Oxime containing complexes have the potential use as the catalysts due to their natural
enzyme mimicking abilities [23-26]. Oxidoreductase family of enzymes involves in various
life-sustaining reactions ranging from the simplest organism to human beings. Catecholase
(catechol oxidase: EC 1.10.3.1) and phenoxazinone synthase (o-aminophenol oxidase: EC
1
.10.3.4) are the members of this group both carrying a Cu(II) atom in their active site as a
cofactor which is one of the factors we considered during the design of the complexes [27].
In this study, an oxime containing ligand (HBOX: Hydrogen Biphenyl OXime) and its
mononuclear Cu(II) and Mn(II) complexes were synthesized and characterized
spectrophotometrically and theoretically. The structural and spectroscopic data of the HBOX
and its metal complexes in the ground state were calculated by using DFT/B3LYP/6-
3
11G(d,p) and DFT/B3LYP/LANL2DZ levels, respectively. Molecular docking studies were
performed to understand the antiangiogenic efficiency of the HBOX ligand. In addition,
enzymatic potentials of both complexes were tested for catecholase and phenoxazinone
synthase-like activities.
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2
. EXPERIMENTAL
2
.1. Physical Measurements
All chemicals acquired commercially in their highest purity grade were used as they received.
1
13
The instruments used in our experiments were as follows: H- and C-NMR spectra of the
ligand were recorded on a JEOL NMR-400 MHz spectrometer, using TMS as an internal
standard and chloroform as a solvent. Spectrophotometric measurements were applied with a
PG T80+ UV-Vis spectrometer. FT-IR spectra of the synthesized ligand and its metal
complexes were measured using a Schimadzu IRPrestige-21 FT-IR spectrophotometer within
-
1
the range 4000-400 cm , using KBr disc pellets. Powder X-ray diffraction (XRD) pattern of
the complex was recorded with a Bruker AXS D8 Advance X-ray diffractometer. In addition,
LECO 932 CHNS analyzer was used to determine C, H and N proportion of the compounds.
Perkin Elmer Optima 5300 DV ICP-OES spectrometer was used to obtain metal contents of
-
3
the complexes. Also, molar conductance of the complexes in DMF (10 M solution) were
measured on an Optic Ivymen System conductivity meter at room temperature. Melting points
were determined with an Electrothermal model IA 9100 digital instrument. Magnetic moment
value measurements were carried out at room temperature on a Sherwood Scientific Magnetic
Susceptibility Balance (Model MX1).
2
.2. Synthesis of Ligand [2-([1,1'-biphenyl]-4-yl)-N-(3,4-dimethylphenyl)-N'-hydroxy-2-
oxoacetimidamide (HBOX)]
1
-(biphenyl-4-yl)ethanone and 2-(biphenyl-4-yl)-2-oxoacetaldehyde oxime was synthesized
according to the literature [28-30]. As a starting material, 2 mmol, 0.519 g of 2-(biphenyl-4-
yl)-N-hydroxy-2-oxoacetimidoyl chloride was dissolved in 20 mL EtOH. The mixture was
7

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cooled and kept at -5°C during dropwise addition of ethanol solution of 4 mmol 3,4-dimethyl
aniline to the solution of chloroketooxime over 15 min. Expected precipitation and color
change were observed in the reaction medium just after the addition. The reaction mixture left
stirring 2 hours at minus temperatures. Then it was allowed to stir at ambient temperature at
least for 2 more hours. The resulting precipitation was filtered, washed by aqueous sodium
bicarbonate (1% w/v), distilled water and ethanol and then, dried on P2O5.
2
.2. Synthesis of Complexes [Cu(BOX)2H2O and Mn(BOX)2H2O]
For the synthesis of the complexes, 0.30 mmol Cu(CH3COO)2.H2O (0.029 g) or
Mn(CH3COO)2.4H2O (0.036 g) in 30 mL ethanol was added to previously prepared 30 mL
o
ethanol solution of the ligand HBOX. After stirring in water bath at 40 C for a few minutes,
the pH was adjusted to 5.5-6 with KOH solution containing 1% of ethanol. Stirring continued
o
for 2 more hours and the mixture transferred to +4 C after cooling and kept overnight. Final
solution was filtered, the precipitation was rinsed with water and ethanol and left to dry.
2
.3. Theoretical Calculations
Theoretical calculations were carried out by the Gaussian 09 software [31] with the DFT level
of theory. Optimizations and frequency calculations were performed by B3LYP functionals
with 6-311G(d,p) and LANL2DZ basis set [32,33] in the gas phase. Files obtained from the
optimization were introduced as the starting file for succeeding calculations. For graphics and
other simulations, GaussView 5.0.9 program was used [34]. Uv-Vis spectrums and electronic
properties like HOMO and LUMO energies were obtained by TD-DFT method with the same
set. Molecular orbital contributions were calculated by GaussSum program [35]. Finally, the
1
13
H- and C-NMR data were calculated by GIAO method [36,37].
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2
.4. Molecular Docking
Molecular docking studies were performed on SwissDock web server using EADock DSS
algorithm [38]. High resolution crystal structures of VEGFR-2 (PDB ID: 2XIR) and COX-2
(PDB ID:1CX2) were obtained from protein data bank (https://www.rcsb.org). COX-2
subunit A was used for simplicity and a better display among the four subunits for docking
and visualizing. All images in molecular docking studies were drawn with the UCSF Chimera
package [39].
2
.5. Enzymatic Activity Studies
2
.5.1. Catecholase-Like Enzyme Activity
Catecholase-like activity of the complexes were measured by the enzymatic conversion of
-
3
substrate 3,5-di-tert-butylcatechol (3,5-DTBC, 5x10 M) to quinone product 3,5-di-tert-
-
4
butyl-o-benzoquinone (3,5-DTBQ) by Cu(II) and Mn(II) complexes (1x10 M each). The
increase in product was followed spectrophotometrically by the increase in absorbance at 400
nm with 30 sec. intervals. The method was modified from Reim and Krebs [40].
2
.5.2. Phenoxazinone Synthase-Like Enzyme Activity
Cu(II) and Mn(II) complexes were tested for phenoxazinone-like activity. For the reaction to
-
4
-3
occur, 1.67x10 M of complex as the enzyme with 12.5x10 M 2-aminophenol (OAPH) as
the substrate were mixed and final volume was adjusted to 25 mL with DMF. The formation
of 2-aminophenoxazine-3-one (APX) as the product was followed spectrophotometrically by
the increase in absorbance at 433 nm with 30 sec. intervals [41]. The rate constants (kobs) for
both enzymatic reactions, catalysed by both complexes were calculated from the equation
below.
ln(A∞/A∞-At)=kt
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A∞ and At are the absorbances of the formed product at time = ∞ and time = t
3
. RESULTS AND DISCUSSION
The route for the synthesis of the HBOX ligand is shown in Fig. 1. Numerous physical and
spectroscopic methods have been used for the characterization of compounds. Measured and
calculated physical properties of the compounds are given in Table 1.
Figure 1. Synthetic plan for the 2-([1,1'-biphenyl]-4-yl)-N-(3,4-dimethylphenyl)-N'-hydroxy-
2
-oxoacetimidamide ligand (HBOX)
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Table 1. Some physical data and elemental analysis results of the synthesized compounds
Color
Yield
Melting
Point
( C)
Found (Calculated) %
μeff
Conductivity
-1 2 -1
Compound
HBOX
(
B.M.) ( cm mol )
o
C
H
N
Metal
(% )
Bright
yellow
7
(
6.58
76.72)
5.73
8.34
-
-
178
280
-
(5.85)
(8.13)
(81)
Dark
brown
6
(
8.96
68.78)
5.48
7.06
8.52
(8.27)
Cu(BOX)2H2O
Mn(BOX)2H2O
1.78
5.94
9.5
6.7
(5.25)
(7.29)
(59)
Black
44)
69.28
(69.56)
5.56
7.14
7.47
(7.23)
>300
(
(5.31)
(7.37)
1
13
3
.1. H- and C-NMR spectra
1
13
H- and C-NMR spectra were recorded in chloroform and calculated by the use of gauge-
including atomic orbital (GIAO) method (Fig. S1 and S2) and both chemical shift data are
1
shown in Table 2 and 3. The H-NMR spectrum of the HBOX showed a D2O exchangeable
proton (H35) at 8.14 ppm due to the oxime group. This chemical shift value in the lowest field
of the spectrum was calculated as 8.65 ppm. The C-H protons of the aromatic groups were
recorded in 6.56-7.69 ppm range as multiplet peaks. These values were calculated between
6
.47-8.55 ppm. The chemical shift for the N-H proton of the amine group in the HBOX was
appeared at 5.92 ppm. The H40 proton, bound to the highly electronegative nitrogen atom,
was calculated in a slightly lower area than the experimental result (calc. 6.27 ppm). Since the
nitrogen was adjacent to the aromatic ring, this shift might be affected by the structures in
resonance. Additionally, signals observed at the 2.13 and 1.59 ppm were attributable to the
H41-H46 protons of the HBOX. Their corresponding calculated chemical shift appeared at
2
.37 and 1.98 ppm, respectively.
11

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1
Table 2. Recorded and calculated chemical shift values of the HBOX in H-NMR spectra
δ(ppm)
Experimental
in CDCl3)
.59
Theoretical
(in CDCl3)
Atom
(
1
H41-H46
(s, 3H)
2.13
1.98
2.37
(aliphatic)
(s, 3H)
H40
5.92
(s, 1H)
6
.27
6.47-8.55
.65
(
amine)
H27-H34
H36, H37-H39
6
(
.56-7.69
m, 12H)
(
aromatic)
H35
oxime)
8.14
(s, 1H)
8
(
s:singlet, m:multiplet
1
3
By considering the C-NMR spectra of the HBOX molecule, the peak observed at 186.58
ppm was assigned to the carbonyl carbon (C13) which was calculated as 170.41 ppm at the
mentioned level of theory. The signal at 149.24 ppm assigned to the C14 carbon of the oxime
group was computed as 135.06 ppm. The oxime carbon was also observed in a low region
such as carbonyl carbon. High electronegative nitrogen and oxygen atoms adjacent to the
oxime carbon caused such a downfield shift. The chemical shifts for the aromatic ring carbons
(C1-C12 and C19-C24) were appeared in the range of 118.29-146.66 ppm and the calculated
range for these carbons was between 102.20-134.80 ppm. The highly shielded methyl carbons
(C25, C26) of the HBOX molecule showed signals at 19.18 and 19.98 ppm and calculated as
1
13
1
1.23 and 11.97 ppm, respectively. Comparison of experimental and theoretical H- and C-
NMR spectra revealed that, the chemical shifts were found to be compatible with each other
and in good agreement with the data reported in the other oxime molecule researches [42-48].
The theoretical chemical shifts seem to appear in lower region approximately 15 ppm
1
3
compared to the experimental shifts for C-NMR.
12

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1
3
Table 3. Recorded and calculated chemical shift values of the HBOX in C-NMR spectra
δ(ppm)
Experimental
in CDCl3)
Theoretical
(in CDCl3)
Atom
(
C25, C26
1
9.18, 19.98
11.23, 11.97
(aliphatic)
C1-C12,
C19-C24
(aromatic)
118.29-146.66
102.20-134.80
C14
oxime)
C13
carbonyl)
1
1
49.24
86.58
135.06
170.41
(
(
3
.2. FT-IR spectra
The calculated vibrational wavenumbers were scaled as 1.0119 for frequencies lower than
-
1
-1
1
800 cm and 0.9682 for frequencies higher than 1800 cm [49]. Some experimental and
calculated wavenumbers for the HBOX and its mononuclear Cu(II) and Mn(II) complexes are
given in Table 4. Comparative experimental and calculated spectra are also given for HBOX,
Cu(BOX)2H2O and Mn(BOX)2H2O in Fig. S3, S5 and S6, respectively. FT-IR spectra of the
-
1
HBOX showed a broad band at 3231 cm which was assigned to (O-H) stretching vibration
-
1
of oxime group. The theoretical frequency of this band was found to be as 3126 cm . This
band was disappeared with the formation of the complex and new bands were appeared at
-
1
-1
3
645 cm (calc. 3716) and 3638 cm (calc. 3702) for the Cu(BOX)2H2O and Mn(BOX)2H2O,
respectively. The disappearance of the band belonging to the O-H of oxime group showed that
the HBOX ligand participated to the coordination via the oxime oxygen atom. The broad
-
1
bands around 3600 cm can be assigned to the coordination water in the complexes. Since the
theoretical calculations were done in gaseous phase, the calculated frequencies for the
-
1
-1
coordination water deviated about 70 cm . The peak observed at 3382 cm was assigned to
-
1
-1
the ν(N-H) and its computed wavenumber was found as 3451 cm . The band at 3382 cm
-
1
underwent a shift to lower frequencies about 60 cm by the complexation which indicates the
13

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coordination of the nitrogen atom of amine with metal ions. Coordination of the oxime
oxygen and the amine nitrogen atoms in the complexes was proved by the appearance of new
-
1
bands at about 545 and 450 cm , which were assigned to ν(M-O) and ν(M-N), respectively
[50,51]. Theoretically computed M-O and M-N stretching vibrations were appeared at about
-
1
5
80 and 470 cm , respectively.
-
1
Table 4. Some selected band frequencies (cm ) of the HBOX and its Cu(II) and Mn(II)
complexes
Compounds
HBOX
O-H(water) O-H(oxime) N-H
C=N N-O
3382m 1602s 1384s 1510s
3451 1628 1387 1524
C-N M-O M-N
Exp.
-
-
3231b
3126
-
-
-
-
Theo.
Exp.
Theo.
Exp.
3645b
3716
-
-
-
3323b 1599s 1453m 1503s 547w 445w
3364 1611 1458 1518 582 471
3327b 1598s 1452m 1492m 545w 449w
Cu(BOX)
2
2
H O
3638b
Mn(BOX)
2
H O
2
Theo.
3702
-
3371
1612
1464
1513
577
475
b:broad, s:sharp, m:medium, w:weak
-
1
The C=N vibration of oxime group of the HBOX was observed as a sharp band at 1602 cm .
-
1
The same vibration appeared at 1628 cm in theoretical spectrum. Since the oxime nitrogen
was not directly involved in coordination, no significant shift was observed in the stretching
-
1
vibration of this band with complexation. However, the stretching vibration at 1384 cm ,
-
1
belonging to the N-O bond of the ligand oxime, shifted to 1453 cm (calc. 1458) for
-
1
Cu(BOX)2H2O and 1452 cm (calc. 1464) for Mn(BOX)2H2O with the complex formation.
These shifts to higher wavenumbers also supported that the ligand was participated in
14

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coordination through the oxygen atom of the oxime group [42]. Data obtained from the FT-IR
spectra showed that the experimental frequencies were quite compatible with the theoretical
ones.
3
.3. Uv-vis spectra and Frontier Molecular Orbital Analysis
Uv-vis spectra of the HBOX and its mononuclear Cu(II) and Mn(II) complexes were recorded
in ethanol solvent. Electronic parameters such as the oscillator strengths, excitation energies
and absorption wavelengths were calculated using TD-B3LYP/6-311G(d,p) level of theory in
same solvent. Experimentally obtained and computed Uv-vis spectra of the HBOX are shown
in Fig. S4 using the same scale and electronic parameters are also given in Table 5 and 6. The
experimental Uv-vis absorption spectrum of the free HBOX ligand showed three maximum
*
wavelength values at 295, 316 and 339 nm. The band at 295 nm was due to π→π transitions
in the benzene ring [52]. Its corresponding calculated wavelength as the most intense band
was observed at 318 nm (f=0.5822) with HOMO-1→LUMO (76%) and HOMO-6→LUMO
-
1
(
20%) contributions. No shifting at 295 cm band by the complex formation indicated that the
benzene ring was not participated in the coordination.
15

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Table 5. Electronic transitions, wavelengths, oscillator strengths and major contributions for
HBOX in ethanol solvent
Theoretical
Transitions
Experimental
B3LYP/6-311G(d,p)
Major
λ (nm)
E (eV) λ (nm) E (eV)
f
contributions
H-1→L (76%)
H-6→L (20%)
H-6→L (65%),
H-1→L (21%)
H→L (99%)
*
π π (benzene)
295
4.20
318
3.90
0.5822
*
π π (imine)
316
339
3.92
3.66
332
434
3.73
2.86
0.1149
0.0198
*
n π
The band at 316 nm can be attributed to π→π* transitions of the imine moiety of oxime group
calc. 332 nm). The band calculated at 332 nm was composed of the HOMO-6→LUMO and
(
HOMO-1→LUMO with 65% and 21% contributions, respectively. The imine group was also
affected by the binding of oxime oxygen to the metal ions, which was shifted to the lower
wavelengths for Cu(BOX)2H2O complex (300 nm) and to higher wavelengths for the
*
Mn(BOX)2H2O complex (328 nm). The band appeared at 339 nm was attributed to the n→π
-
1
transitions of the free ligand and its corresponding wavelength was calculated as 434 cm
which was occurred between HOMO and LUMO with 99% contribution. The band at 339 nm
corresponding to the n→π* transitions of the HBOX was observed at 310 and 360 nm in the
Cu(BOX)2H2O and Mn(BOX)2H2O spectra, respectively. These shifts were indicated that the
ligand was coordinated through the oxime oxygen. The weak bands observed at 504 and 695
nm in the spectra of the complexes, which can’t be seen in the spectrum of HBOX, were
originated from the d→d transitions of the metal ions [52]. These transitions revealed that
both Cu(BOX) H O and Mn(BOX) H O complexes were in distorted square-pyramidal
2
2
2
2
geometry [53].
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Table 6. Electronic transitions and wavelengths of metal complexes of the
HBOX in ethanol solvent
*
*
*
π π (benzene) π π (imine) n π dd
Complexes
2
2
95
95
300
328
310
360
504
695
Cu(BOX)2H2O
Mn(BOX)2H2O
It is well known that the energy gap between the highest occupied molecular orbital (HOMO)
and the lowest unoccupied molecular orbital (LUMO) gives valuable information about the
chemical reactivity, stability and polarizability of the molecules [54,55]. 3D plot with energy
gap value between HOMO and LUMO of HBOX and its Cu(II) and Mn(II) complexes are
given in Fig. 2-4. Calculated energy gap value for the HBOX between the ground state and
the first exited state was found to be 3.504 eV. This calculated energy gap was smaller than
the conventional oxime bearing compounds in the literature [56,57]. The lower the energy gap
corresponds to the higher the reactivity of the HBOX molecule. As can be seen from Fig 2.,
the majority of HOMO was localized on dimethlybenzene moiety whereas the LUMO was
mainly on the biphenyl region.
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Figure 2. Molecular orbital surface and HOMO-LUMO gap of the HBOX computed at
B3LYP/6-311G(d,p) level
Figure 3. Molecular orbital surface and HOMO-LUMO gap of the Cu(BOX)2H2O computed
at B3LYP/LANL2DZ level
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Figure 4. Molecular orbital surface and HOMO-LUMO gap of the Mn(BOX)2H2O computed
at B3LYP/LANL2DZ level
3
.4. Thermal Analysis
The thermogravimetry (TG) curves give information on the thermal stability and the products
formed on heating. Thermal analyses for the HBOX and its metal complexes were performed
o
within the temperature range from ambient temperature up to 1000 C. In TG, the changes in
the weight of the molecule were recorded as a function of the temperature during heating. The
TG curve was also supported by the derivative thermogravimetry (DTG) curves.
The thermal behaviour of both complexes was almost the same. For this reason, only
Cu(BOX)2H2O complex were discussed here in detail. The thermogram of the HBOX and
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[Cu(BOX)2H2O] are shown in Fig 5 and 6, respectively. The critical data obtained from the
investigation of the thermograms are given in Table 7.
2
00.0
5
5
5.00
0.00
11.40
1
80.0
1
60.0
45.00
40.00
35.00
30.00
1
1.20
140.0
11.00
120.0
100.0
10.80
2
2
5.00
0.00
8
0.0
0.0
0.0
0.0
1
0.60
6
4
2
1
5.00
10.40
1
0.00
10.20
5
.00
.00
0
.0
0
100
200
300
400
500
Temp Cel
600
700
800
900
1000
o
Figure 5. Thermogram of the HBOX taken between room temperature and 1000 C
180.0
4
4
3
.500
.000
.500
1
.400
160.0
140.0
1.200
120.0
1.000
100.0
0
0
0
0
0
.800
.600
.400
.200
.000
8
0.0
0.0
0.0
0.0
3.000
6
4
2
2
.500
2.000
100
200
300
400
500
Temp Cel
600
700
800
900
1000
o
Figure 6. Thermogram of the [Cu(BOX)2H2O] taken between room temperature and 1000 C
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The HBOX ligand with the molecular formula C22H20N2O2 was thermally decomposed in two
o
successive degradation steps. The first step occurs between 170 and 510 C with a mass loss of
9
.21% (calc. 9.29%) which is reasonably accounted for the loss of 1 mole of hydroxyl and 1
o
mole of methyl groups. The second degradation step started at 510 C but the decomposition
o
was not completed at 1000 C.
The thermal decomposition of the [Cu(BOX)2H2O] with the molecular formula
[C44H40CuN4O5] was taken place in four succeeding stages. In the first stage, from 20 to
o
1
2
95 C, one mole of coordinated H2O molecule degraded with a 2.32% mass loss (calc.
o
o
.34%). The second stage started at 195 C and ended at 215 C in which the corresponding
mass loss of 11.72% was due to the degradation of 2 moles of oxime groups confirming the
o
calculated value (calc. 10.93%). The DTG curve showed a peak at 215 C for this stage. The
third estimated mass loss of 39.52% (calculated mass loss, 39.82%), occurred within the
o
temperature range of 215-550 C, can be attributed to the liberation of 2 moles of biphenyl
o
groups. In the DTG curve, three peaks at 290, 350 and 380 C were observed for this region.
o
o
The last decomposition stage started at 550 C was still in progress at 1000 C. By considering
the thermal analyses results, found and calculated mass loss percentages obtained from the
related decomposition stages were in good agreement.
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Table 7. Thermoanalytical results (TG, DTG) of the HBOX and Cu(BOX)2H2O
Estimated
(calculated) %
TG range
DTGmax
Assignment
Residue
Compounds
o
o
(
C)
( C)
Total mass
loss
Mass loss
1
mole of
1
2
2
3
80,
10,
60,
40
hydroxyl
group
1
9.21
(9.29)
170-510
mole of
HBOX
methyl group
Decomposition
is in progress
510-
1
mole of
2
.32
20-195
-
coordinated
water
(2.34)
2
mole of
oxime
1
1.72
195-215
215
(10.93)
groups
Cu(BOX)2H2O
2
90,
350,
80
2 mole of
biphenyl
groups
3
9.52
215-550
(39.82)
3
Decomposition
is in progress
550-
3
.5. X-ray Diffraction
X-ray diffraction measurement was carried out to obtain further evidence of the structure of
o
the Mn(BOX)2H2O. X-ray pattern of the title complex was recorded at 2θ = 10-90 range and
shown in Fig. 7. It was found that the Mn(BOX)2H2O complex showed sharp peaks. The
XRD patterns indicated the crystalline nature of the studied complex and this behaviour is due
to the incorporation of water molecules into the coordination sphere [58,59]. Attempts to
prepare the single crystal synthesis of the both complexes were unsuccessful.
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Figure 7. XRD diffraction pattern of the Mn(BOX)2H2O complex
Magnetic Susceptibility
The magnetic susceptibilities of the [Cu(BOX)2H2O] and [Mn(BOX)2H2O] were measured at
room temperature using Gouy method and the results were given in Table 1. The results
revealed that both complexes were paramagnetic at ambient temperature. Magnetic moment
values of the mononuclear complexes were 1.78 and 5.94 B.M. for [Cu(BOX)2H2O] and
9
[
Mn(BOX)2H2O], respectively. By considering the [Cu(BOX)2H2O] complex, Cu(II) ion (3d )
had one unpaired electron in its 3d shell, therefore its compounds were considered to have
magnetic moments close to the spin-only value of 1.73 B.M. According to the magnetic
moment value of [Cu(BOX)2H2O], the environment around the Cu(II) was square-pyramidal
geometry [60].
On the other hand, the magnetic moment value of [Mn(BOX)2H2O] was close to 5,92 B.M.
5
of high-spin penta-coordinated manganese(II) ion with d electron configuration. It can be
concluded from these data that the metal(II) ion in the complexes were coordinated with the
23

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oxime and amine groups in the HBOX and completed five-coordinated geometry with one
mole of coordination water.
3
.6. Molar Conductivity
The investigation of the molar conductivity values of the complexes was highly supportive of
their structures. Molar conductivity measurements were performed in N,N-
dimethylformamide solutions at room temperature. Obtained conductivity values for the
-
1
2
-1
[
Cu(BOX)2H2O] and [Mn(BOX)2H2O] were found to be 9.5 and 6.7 Ω cm mol ,
respectively. The molar conductance of the synthesized Cu(II) and Mn(II) complexes in DMF
revealed that the complexes had a non-electrolyte nature [61].
Figure 8. The proposed structure of synthesized complexes
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3
.7. Computational Details
.7.1. Molecular Structure
3
The optimized molecular structure of the HBOX and its mononuclear Cu(II) and Mn(II)
complexes were calculated by using B3LYP/6-311G(d,p) and B3LYP/LANL2DZ level of
theory, respectively. The numbering schemes of the compounds are given in Fig. 9-11. The
optimized parameters (band lengths, bond angles and dihedral angles) of the HBOX and metal
complexes calculated by mentioned levels are listed in Table 8 and 9, respectively. The
calculated C4-C7 bond length between two benzo rings in biphenyl moiety was found as
1
.484 Å. The C10-C13 and C13-C14 bond lengths between biphenyl-carbonyl and carbonyl-
oxime groups were computed as 1.489 and 1.518 Å, respectively. In addition, the C14-N15
and N15-O17 bond lengths in the oxime group were found to be 1.299 and 1.419 Å,
respectively. When the calculated bond length values of the HBOX molecule were examined
collectively it was found that the aliphatic C-C bonds were longer than the aromatic ones and
the aromatic ones were longer from the double bonds as expected. The HBOX molecule can
be divided into four parts: biphenyl, carbonyl, oxime and amine. The dihedral angles for the
C11-C10-C13-C14, O16-C13-C14-N15 and N15-C14-N18-H40 related biphenyl-carbonyl,
o
o
o
carbonyl-oxime and oxime-amine moieties were calculated as -167.1 , -134.7 and 8.6 ,
respectively. This result revealed that the molecular geometry of the HBOX was nonplanar
and four parts of the title molecule were not in the same plane.
25

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Figure 9. The optimized geometric structure of the HBOX computed at B3LYP/6-311G(d,p)
level
o
C5-C4-C7-C8 dihedral angle was obtained as 143.1 in mentioned level which was revealed
that the two benzo rings in biphenyl group was not in same plane, too.
o
Table 8. Some selected calculated bond lengths (Å), bond and dihedral angles ( ) for the
HBOX computed at B3LYP/6-311G(d,p) level
Parameters
Bond lengths (Å)
B3LYP/6-311G(d,p)
C5-C6
1.394
C2-C3
1.394
1.086
1.484
1.391
1.387
1.085
1.489
1.222
1.518
C1-H9
C4-C7
C8-C9
C11-C12
C11-H36
C10-C13
C13-O16
C13-C14
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C14-N15
1.299
1.419
0.966
1.368
1.410
1.393
1.395
1.510
1.510
1.098
N15-O17
O17-H35
C14-N18
N18-C20
C19-C24
C21-C22
C23-C26
C22-C25
C26-H46
o
Bond angles ( )
C6-C1-C2
119.5
118.2
120.9
118.0
118.7
117.4
121.8
109.8
129.2
118.9
119.4
121.2
C5-C4-C3
C3-C4-C7
C8-C7-C12
C9-C10-C11
C11-C10-C13
C10-C13-O16
C14-N15-O17
C14-N18-C20
C19-C20-C21
C20-C19-C24
C22-C23-C26
o
Dihedral angles ( )
C1-C6-C5-C4
-0.1
C5-C4-C7-C8
143.1
-167.1
-134.7
-174.2
-157.9
-147.1
C11-C10-C13-C14
O16-C13-C14-N15
C14-N15-O17-H35
C14-N18-C20-C21
C10-C13-C14-N18
27

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Bond lengths, bond angles and dihedral angles of the Cu(BOX)2H2O and Mn(BOX)2H2O
complexes were quite similar. Slight elongations were observed in the aromatic and aliphatic
carbon-carbon bond lengths, with the formation of the complexes. But the C14-N18 (1.368 Å)
bond between the oxime and amine groups elongated significantly with the complex
formation. For both complexes, C14-N18 bond length was 1.485 Å and 1.497 Å, respectively.
Due to the electron donation of N18 to the metal ion, the charge density decreased and the
bond with the oxime carbon was weakened and elongated. In addition, the N15-O17 bond
length of 1.419 Å in the HBOX ligand was shortened in the complexes and was calculated as
1
.358 Å (N43-O52) and 1.361 Å (N43-O52) for Cu(BOX)2H2O and Mn(BOX)2H2O
complexes, respectively. The reason of this shortening in the nitrogen-oxygen bond was
interpreted as backbonding. With the backbonding interaction, the metal ions in the
complexes donated the electron density back to the ligand and the nitrogen-oxygen bond in
oxime group was shortened by strengthening.
From the angles around the Cu(II) and Mn(II), two oxime oxygen and amine nitrogen atoms
were not lying in same plane. The coordination water was bound perpendicular to the metal
ion. The resulting bond distances and angles for the Cu(BOX)2H2O and Mn(BOX)2H2O
complexes were consistent with a pentacoordinate distorted square-pyramidal structure
involving the amine nitrogen atom, deprotonated oxime oxygen atom and coordination water.
28

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Figure 10. The optimized geometric structure of the Cu(BOX)2H2O computed at
B3LYP/LANL2DZ level
29