Biomimetic complexes of Mn(II), Fe(III), Co(II), and Ni(II) with 1,10-phenanthroline and a salen type ligand: tailored synthesis, characterization, DFT, enzyme kinetics, and antibacterial screening

Article abstract of DOI:10.1080/00958972.2018.1536267

A symmetrical tetradentate Schiff base, o-HACPHENEN, was prepared by condensation of 1,2-bis(ethylenediamine) with o-hydroxyacetophenone in 1:2 molar ratio. The Schiff base ligand and 1,10-phenanthroline were used for the tailored synthesis of four mixed-

Full text of DOI:10.1080/00958972.2018.1536267

Journal of Coordination Chemistry
ISSN: 0095-8972 (Print) 1029-0389 (Online) Journal homepage: http://www.tandfonline.com/loi/gcoo20
Biomimetic complexes of Mn(II), Fe(III), Co(II) and
Ni(II) with 1,10-phenanthroline and a salen type
ligand: Tailored synthesis, characterization, DFT,
enzyme kinetics and antibacterial screening
Satyendra N. Shukla, Pratiksha Gaur, Preeti Vaidya, Bhaskar Chaurasia &
Sangeeta Jhariya
To cite this article: Satyendra N. Shukla, Pratiksha Gaur, Preeti Vaidya, Bhaskar Chaurasia
& Sangeeta Jhariya (2018): Biomimetic complexes of Mn(II), Fe(III), Co(II) and Ni(II) with
1,10-phenanthroline and a salen type ligand: Tailored synthesis, characterization, DFT,
enzyme kinetics and antibacterial screening, Journal of Coordination Chemistry, DOI:
10.1080/00958972.2018.1536267
To link to this article: https://doi.org/10.1080/00958972.2018.1536267
View supplementary material
Accepted author version posted online: 16
Oct 2018.
Submit your article to this journal
View Crossmark data
Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=gcoo20

Biomimetic complexes of Mn(II), Fe(III), Co(II) and Ni(II) with
1,10-phenanthroline and a salen type ligand: Tailored synthesis,
characterization, DFT, enzyme kinetics and antibacterial screening
SATYENDRA N. SHUKLA*, PRATIKSHA GAUR, PREETI VAIDYA, BHASKAR CHAURASIA and
SANGEETA JHARIYA
Coordination Chemistry Research Lab, Department of Chemistry, Government Science College,
Jabalpur (M.P.) 482001, INDIA
A symmetrical tetradentate Schiff base, o-HACPHENEN, was prepared by condensation of
1,2-bis(ethylenediamine) with o-hydroxyacetophenone in 1:2 molar ratio. The Schiff base ligand
and 1,10-phenanthroline were used for the tailored synthesis of four mixed-ligand complexes,
[Mn(o-HACPHENEN)(1,10-phen)]
(1),
[Fe(o-HACPHENEN)(1,10-phen)]Cl
(2),
[Co(o-HACPHENEN)(1,10-phen)] (3) and [Ni(o-HACPHENEN)(1,10-phen)] (4). The ligand
and complexes were characterized on the basis of elemental analyses, ESI-MS, molar
1
conductance, electronic, FT-IR, H-NMR and 13C-NMR spectra. DFT study was used to
optimize the geometry of the investigated compounds. Using Gaussian09 molecular modelling,
HOMO-LUMO study, bond lengths, bond angles, molecular electrostatic potential map (MEP)
and Mulliken charge were also evaluated. Complexes displayed remarkable catalase-like activity
in the disproportion reactions of hydrogen peroxide. The kinetics of the activity were
investigated and data are fitted in Lineweaver Burk plot, revealing Michaelis Menten behavior.
The catalase activity of ligand and complexes were found in the order 1 > 2 > 4 > 3 >
o-HACPHENEN. Compounds were screened for their antibacterial activity against gram-
negative bacteria Escherichia coli in comparison to standard drugs. Complexes were more potent
than Schiff base. The MIC of complexes was also studied.
Keywords: Symmetrical Schiff base ligand; Biomimetic; Catalase activity; DFT Study; Enzyme
kinetics
*Corresponding author. Emails: sns1963_1@rediffmail.com, ccrl_2004@rediffmail.com

1. Introduction
Molecular oxygen present in the air is a clean, abundant and environmentally green reagent,
ideally suited for substrate oxidation processes including selective C–H bond functionalization.
But in the tissues in particular condition, it sometimes changes into hydrogen peroxide like
reactive oxygen species (ROS) which are actually responsible for “oxidative stress” reactions
leading to aging and other irreversible damage to the body [1-3]. Catalase is an enzyme that
disproportionates hydrogen peroxide into H2O and O2, and thereby protects living organisms
from ROS. In other words, it is the most abundant peroxisomal protein which protects
peroxisomes and cells from the toxic effects of peroxides [4].
Mixed-ligand complexes containing nitrogen and oxygen atoms have significant
importance not only due to their various biological activities but also owing to their catalytic
activities of industrial importance, i.e. hydrogenation, hydroformylation, oxidative hydrolysis of
olefins and carboxylation of methanol [5-7]. 1,10-Phenanthroline is a strong-field bidentate
ligand with a rigid framework, displaying superb ability to chelate many metal ions via two
nitrogen donors. It also shows potential for technological applications, due to high charge
transfer mobility, bright light-emission, good electrochemical and photoactive properties as well
as in vitro antimicrobial activity against a broad-spectrum of bacteria [8-12].
Schiff base complexes with a variety of metals have been extensively used as efficient
homogeneous and heterogeneous catalysts, sensors, corrosion inhibitors and model compounds
for active centers of metalloenzymes [13-16]. Metal salen complexes have attracted great
attention due to their interesting geometry and vivid biomimetic activity [17, 18]. In octahedral
M(salen) complexes, three potential configurations are possible, trans (planar), cis-α and cis-β.
Due to the relatively rigid planarity of salen skeleton, trans geometry is more stable than cis
geometry and that is the main reason that most complexes adopt trans geometry [19]. But in the
presence of bulky group on donor atom or on its nearby group, complexes can adopt a cis-
configuration. For ONNO-tetradentate ligand, cis-β geometry is more common [20]. Some
model compounds like salen-manganese complexes are known to be promising because they
exhibit dual biomimetic functions, i.e. superoxide dismutase, peroxidase, cytochrome P-450 and
catalase mimetic activities [21-23].
In anticipation of interesting chemistry, biomimetic and biological activity, we have
synthesized a salen type ligand and its four mixed-ligand complexes of Mn(II), Fe(III), Co(II)

and Ni(II) with 1,10-phenanthroline. Ligand and complexes were characterized by spectroscopic
techniques and screened for their biological and biomimetic activity. Since density functional
theory (DFT) is a simulated quantum mechanical technique which provides information about
compounds before its synthesis in the lab, we have employed DFT in optimizing the structure of
the ligand and complexes.
2. Experimental
2.1. Materials and methods
Ethylenediamine (Koch Light Lab.), o-hydroxyacetophenone, MnCl2·4H2O, FeCl3, CoCl2·6H2O,
NiCl2·6H2O and 1,10-phenanthroline (all E. Merck) were purchased from the market and used as
received. Analytical reagent grade methyl alcohol, ethyl alcohol (distilled before use),
acetonitrile, acetone, dimethylformamide, dimethylsulphoxide and other solvents (all E. Merck)
and Mueller Hinton Agar (Himedia) were used as received. Elemental analyses were estimated
on a EuroVector E 3000 Elemental Analyser and metal contents were analyzed gravimetrically
by the literature procedure [24]. Conductivity measurements were carried out in DMSO at 25 °C
on an EI-181 conductivity bridge with dipping type cell. ESI-MS spectra were recorded on an
Agilent-6520(Q-TOF) mass spectrometer. FT-IR spectra were recorded in KBr pellets on a
Shimadzu-8400 PC spectrometer. Electronic absorption spectra were recorded in the range 800-
200 nm with an EI-2371 double beam spectrophotometer equipped with a PC. Guoy’s method
1
was employed for magnetic susceptibility measurements. H- and 13C-NMR spectra were
recorded in DMSO-d6 on a Bruker Avance 400 (FT NMR) spectrophotometer (400 MHz).
2.2. Preparation of Schiff base ligand, 2-[2-(E)-(2-
hydroxyphenyl)ethylidene]aminoethyl)ethanimidoyl]phen, (o-HACPHENEN)
o-HACPHENEN was prepared according to the method reported [25], by the condensation of
1,2-ethylenediamine with o-hydroxyacetophenone in 1:2 molar ratio in methanol. The solid
obtained was recrystallized from hot methanol to yield yellow crystals, which were dried over
anhydrous calcium chloride in a desiccator under vacuum. Yield: 2.582 g (84.36%); m.p. =
175 °C; Anal. Calc. for C18H20N2O2 (MW = 296.36): C, 72.95; H, 6.80; N, 9.45%. Found: C,
72.12; H, 6.75; N, 9.32%. Selected infrared absorption (KBr, cm-1): ν(O-H), 3365(w);
ν(>C=N)imine, 1608(s); ν(CH2-CH2), 949 (s). Electronic spectra (λmax in nm, ε in M-1cm-1) in

DMF: 260 (700), 320 (1661). 1H-NMR spectra (δ, in ppm): 3.330 (s, 6H), (-CH3); 3.943 (s, 4H),
(-CH2); 6.769 (m, 4H, J=1.6 Hz, 1.2 Hz), 7.275 (sextet, 2H, J=1.6 Hz) and 7.675 (qrt, 2H,
J=1.6 Hz), (Ar-H)benzene; 16.034 (s, 2H), (O-H)phenolic.
13C-NMR spectra (δ, in ppm): 14.66,
(-CH3); 49.42, (CH2-CH2); 116.91(C5), 117.82(C3), 119.05(C1), 128.73(C2) and 132.26(C4),
(Ar-C)benzene; 163.01, (C-OH); 173.12, (>C=N)imine. ESI-Mass spectra, m/z: [C9H10NO + H+]+ =
149.0833, [C10H12NO + H+]+ = 163.0952, [C18H20N2O2 + H+]+ = 297.1589, MW = 296.36. The
overall reaction scheme can be represented as shown in scheme 1.
2.3. Synthesis of complexes
Metal chloride (0.001 mol) and 1,10-phen (0.180 g, 0.001 mol) were dissolved in 25 mL ethanol
by stirring for 30 min. This mixture was added to a solution of Schiff base o-HACPHENEN
(0.296 g, 0.001 mol), dissolved in 25 mL hot ethanol and a few drops of acetonitrile in a flat
bottom flask and stirred for 1-2 h in an inert atmosphere. A solid was obtained in each case after
refluxing the reaction mixture for 2-11 h, was filtered and washed several times with acetone.
The solid obtained was recrystallized from hot methanol or an appropriate solvent mixture of
methanol:ethanol:water (v/v) to yield crystalline solid, which was dried in a desiccator over
anhydrous calcium chloride under vacuum at room temperature. The overall reaction scheme can
be represented as shown in scheme 2.
2.3.1. [Mn(o-HACPHENEN)(1,10-phen)] (1). Color = black, Yield: 0.473 g (74.25%); m.p. >
300 °C; Anal. Calc. for C30H26MnN4O2 (MW = 529.14): C, 68.05; H, 4.95; N, 10.58; Mn,
10.38%. Found: C, 68.01; H, 4.93; N, 10.54; Mn, 10.25%. eff = 2.21 BM. Molar conductance
Ʌm at 25 °C (Ω–1 cm2 mol–1): 11 in DMSO. Selected infrared absorption (KBr, cm-1): ν(C-H)arom
,
3050(w); ν(-CH3), 2835(w); ν(>C=N)cyclic, 1635(m) ν(>C=N)imine, 1519(s); ν(C-O), 1251(s);
ν(CH2-CH2), 941(m); ν(M-O), 569(w); ν(M-N), 445(s). Electronic spectra (λmax in nm, ε in
M-1cm-1) in DMF: 560 (15), 620 (50), 720 (20). ESI-Mass spectra, m/z: [C12H8N2 + H+]+ =
181.0721, [C12H8MnN2 + H+]+ = 236.0102, [C18H18N2O2 + 3H+]+ = 297.1601, [C18H18MnN2O2]+
= 350.0782, [C30H26MnN4O2 + H+]+ = 530.1470, MW = 529.14.
2.3.2. [Fe(o-HACPHENEN)(1,10-phen)]Cl (2). Color = black, Yield: 0.543 g (85.24%); m.p. >
300 °C; Anal. Calc. for C30H26ClFeN4O2 (MW = 565.11): C, 63.68; H, 4.63; N, 9.90; Fe, 9.87%.

Found: C, 63.60; H, 4.60; N, 9.85; Fe, 9.80%. eff = 2.62 BM. Molar conductance Ʌm at 25 °C
(Ω–1 cm2 mol–1): 112 in DMSO. Selected infrared absorption (KBr, cm-1): ν(C-H)arom, 3057(m);
ν(-CH3), 2983(w); ν(>C=N)cyclic, 1649(s); ν(>C=N)imine, 1599(s); ν(C-O), 1234(s); ν(CH2-CH2),
950(s); ν(M-O), 532(m); ν(M-N), 449(s). Electronic spectra (λmax in nm, ε in M-1cm-1) in DMF:
320 (1500), 420 (612), 500 (559). ESI-Mass spectra, m/z: [C12H8N2 + H+]+ = 181.0721,
[C18H18N2O2 + 3H+]+ = 297.1601, [C18H18FeN2O2]+ = 350.0705, [C30H26FeN4O2 + H+]+ =
531.3745, [C30H26FeN4O2Cl + 2CH3CN]+ = 647.4388, MW = 565.11.
2.3.3. [Co(o-HACPHENEN)(1,10-phen)] (3). Color = black, Yield: 0.415 g (58.20%); m.p. >
300 °C; Anal. Calc. for C30H26CoN4O2 (MW = 533.14): C, 67.54; H, 4.91; N, 10.50; Co,
11.05%. Found: C, 67.50; H, 4.88; N, 10.42; Co, 10.95%. eff = 2.1 BM. Molar conductance Ʌm
at 25 °C (Ω–1 cm2 mol–1): 8.5 in DMSO. Selected infrared absorption (KBr, cm-1): ν(C-H)arom
,
3057(w); ν(-CH3), 2958(w); ν(>C=N)cyclic, 1657(s); ν(>C=N)imine, 1604(m); ν(C-O), 1236(w);
ν(CH2-CH2), 960(m); ν(M-O), 528(w); ν(M-N), 480(w). Electronic spectra (λmax in nm, ε in
M-1cm-1) in DMF: 520 (30). ESI-Mass spectra, m/z: [C12H8N2 + H+]+ = 181.0721, [C12H8CoN2 +
H+]+ = 240.0053, [C18H18N2O2 + 3H+]+ = 297.1601, [C18H18CoN2O2]+ = 353.0700,
[C30H26CoN4O2 + H+]+ = 534.1421, MW = 533.14.
2.3.4. [Ni(o-HACPHENEN)(1,10-phen)] (4). Color = Brown, Yield: 0.441 g (61.85%); m.p. >
300 °C; Anal. Calc. for C30H26NiN4O2 (MW = 532.14): C, 67.57; H, 4.91; N, 10.51; Ni, 11.01%.
Found: C, 67.55; H, 4.89; N, 10.48; Ni, 10.91%. eff = 3.01 BM. Molar conductance Ʌm at 25 °C
(Ω–1 cm2 mol–1): 10.7 in DMSO. Selected infrared absorption (KBr, cm-1): ν(C-H)arom, 3074(w);
ν(-CH3), 2953(m); ν(>C=N)cyclic, 1657(s); ν(>C=N)imine, 1599(m); ν(C-O), 1246(s); ν(CH2-CH2),
925(m); ν(M-O), 520(m); ν(M-N), 447(s). Electronic spectra (λmax in nm, ε in M-1cm-1) in DMF:
420 (120), 560 (60). ESI-Mass spectra, m/z: [C12H8N2 + H+]+ = 181.0721, [C12H8NiN2 + H+]+ =
239.9995, [C18H18N2O2 + 3H+]+ = 297.1601, [C18H18NiN2O2]+ = 352.0722, [C30H26NiN4O2 +
H+]+ = 533.1409, MW = 532.14.
2.4. DFT calculation
Gaussian09 software package was employed to carry out all the quantum chemical calculations
[26]. The DFT calculations were performed using the B3LYP parameter density function, which

includes Becke’s gradient exchange correction with the Lee, Yang, Parr correlation functional
[27, 28]. Full geometry optimization of the o-HACPHENEN and its complexes 1-4 were carried
out using DFT method of B3LYP with 6-311++G(d,p) or 3-21+G* basic set for all nonmetallic
atoms and Los Alamos National Laboratory 2 double zeta (LANL2DZ) basic set for the central
metal atoms in the gas phase [29]. The optimized structural parameters such as bond lengths and
angles of all the compounds were determined with the atom numbering scheme of the molecule.
The electronic properties such as electronic absorption wavelengths and the representation of the
HOMO, LUMO energy distributions and HOMO-LUMO energy gap of all compounds have
been calculated using the TD-SCF, DFT with 6-311++G(d,p)/LANL2DZ basic set of the B3LYP
level of the theory [30]. The quantum chemical parameters such as separation energies (E),
Mulliken electronegativity (), absolute hardness (), absolute softness (σ), chemical potential
(Pi), global softness (S), global electrophilicity (), additional electronic charge (Nmax), dipole
moment (μ) and total energy (ETD-HF/ETD-KS) were calculated after geometrical optimization
of the structures of all compounds. The distribution of electronegativity/electropositivity over a
large number of atoms was presented by the molecular electrostatic potential (MEP) maps and
were deduced using the same method [31].
2.5. Biological studies
2.5.1. Preparation of yeast catalase. For the preparation of catalase from dry yeast, 5 g dry
yeast was mixed with 50 mL of warm water to make a suspension. The temperature of the water
should not be too hot, but close to body temperature (37 °C). Suspension of the yeast in water
was kept at rest for at least 5-10 min. This homogenized stock suspension was used for further
study.
2.5.2. Catalase-like activity.
H2O2 Disproportionation reactions and its kinetics. The catalase-like activity was observed by
measuring the volume of O2 produced during the reactions of the ligand/complex 1-4 with H2O2
and compared with yeast catalase. The mini reactor was a Kitasato flask (25 mL) magnetically
stirred and closed with a rubber septum. The Kitasato flask was connected to a tank filled with
water. This was arranged in a manner that the water will be pushed out as oxygen fills the tank.
Volumes of evolved dioxygen were measured for 20 min at two-minute time intervals. In the

first set of experiment, which was control set of reaction, 5 mL yeast suspension (0.005 mol)
dissolved in distilled water was added to the reactor through the septum. Thereafter, 10 mL
aqueous solution of H2O2 (0.008 mol, commercial 30%) was injected into it through the rubber
septum with a syringe. Similarly, in other experiments 5 mL ligand/complex solution in DMSO
(0.005 mol) was added to the reactor, followed by 10 mL (0.008 mol, commercial 30% aqueous
solution) of H2O2 through the rubber septum with the help of a syringe. The experiment was
repeated for the highly efficient complex at different concentration. All the experiments were
performed at room temperature of 25 °C.
In order to study enzyme kinetics in characteristic disproportionation testing, the
concentration of the H2O2 was varied to determine the dependence of the initial rate on the
dioxygen evolution. The velocity for the catalase-like activity of the metal enzyme was obtained
by time-dependent measurements of decomposition of hydrogen peroxide to water and evolved
dioxygen.
2.5.3. In-vitro antibacterial assay. o-HACPHENEN and 1-4 were screened for antibacterial
activity against gram-negative bacteria Escherichia coli (ATCC no. 35218, E. coli) at different
concentration using Agar well method [32, 33]. The culture medium was prepared by mixing
2.8 g of Mueller-Hinton agar in 100 mL of water in a sterile conical flask and stirred for 15 min
to ensure complete dissolution. Thereafter, it was autoclaved at 15 psi pressure at 120 °C for
15 min and poured into a sterilized petri plate and allowed to settle. The bacterial culture was
inoculated and spread homogeneously over the culture medium. The inoculant was prepared by
4-5 similar colonies of E. coli. In the particular inoculants of petri plate, the 0.05 mL of 5 mM
(dissolved in DMSO) solutions of the compounds, o-HACPHENEN and 1-4 were added to each
well with the help of a micropipette. These plates were incubated at 37 °C for 24 h in refrigerated
incubator shakers [34]. Standard antibacterial drugs Amoxicillin and Chloramphenicol were used
as references to estimate the effectiveness of the tested compounds under the same conditions.
2.5.4. Minimum Inhibitory Concentration (MIC). Since the appearance of resistance is a
prevalent phenomenon in pathogens, it was obvious to report sensitivity pattern accurately and
rapidly. Therefore, the four complexes were evaluated for MIC. Successive dilution method was
employed for MIC evaluation [35]. In tubes A and B, 1 mL of test solution (1-4) was added;

tube B was stirred and 1 mL mixture of it was taken out and transferred to tube C. This
successive transference was repeated until tube K. The 0.1 mL suspension of inoculums E. coli
having ~5,00,000 cells per μL were added to all tubes, except the tube K. Incubation at 37±1 °C
temperature was developed in 24-48 h. The MIC is the concentration of the highest dilution tube,
in which bacterial growth was absent.
3. Results and discussion
3.1. Characterization of ligand
The elemental analysis data of o-HACPHENEN was in full agreement with the empirical
formula. In ESI-MS (figure S1 in supplementary file), the peak at m/z 297.1589 was a pseudo-
molecular ion peak attributed for [M + H+]+. FT-IR spectrum (figure S2) exhibits a broad
shoulder at ~3365 cm-1 and a sharp peak at 1608 cm-1 assigned to ν(O-H) and ν(>C=N–),
respectively [36]. UV-Vis spectra (figure S3) exhibit two bands at 280 nm and 340 nm assigned
1
to π→ π* and n→ π* transition. H-NMR spectrum of o-HACPHENEN (figure S4) exhibits a
multiplet centered at δ = 6.769 ppm assigned for four aromatic protons associated with C3 and
C6 carbon with coupling constant (J) 1.6 Hz and 1.2 Hz. A quartet and sextet centered at δ =
7.675 ppm and δ = 7.275 ppm were attributed to two protons each, associated with C4 and C5
aromatic carbon with J = 1.6 Hz. Two singlets observed at 3.943 ppm for four protons and
3.330 ppm for six protons were attributed for methylenic and methyl group, respectively. A
singlet observed at 16.034 ppm for two protons was assigned for phenolic (-OH) proton.
The 13C-NMR spectrum of the ligand (figure S5) exhibits a signal at δ = 173.12 ppm
assigned for imine (>C=N–) carbon. The signal at 163.01 ppm was attributed to the carbon
linked with the hydroxyl group. All aromatic carbons display signals between 116.91-
132.26 ppm. The signals at 49.42 ppm and 14.66 ppm were assigned for methylenic carbon and
methyl carbons, respectively [37]. Thus, spectroscopic characterization of the ligand confirms
the structure suggested in scheme 1.
3.2. Characterization of complexes
Stoichiometries of the complexes were in agreement with elemental analyses data. Molar
conductance values for 1, 3 and 4 (10-3 M in DMSO) were 11.0, 8.5 and 10.7 Ω-1cm2 mol-1
,
respectively, indicating their non-ionic nature. However, 2 exhibits high molar conductance

(112 Ω-1cm2mol-1), suggesting ionic nature as 1:1 electrolyte in DMSO solution [38]. ESI-Mass
spectra of all four complexes exhibit several peaks depending on the fragmentation pattern.
Complexes 1, 3 and 4 exhibit several peaks such as at m/z = 181.0721, 236.0102, 240.0053,
239.9995, 297.1601, 350.0782, 353.0700, 352.0722, 530.1470, 534.1421 and 533.1409
attributed for [1,10-phen + H+]+, [C12H8MnN2 + H+]+, [C12H8CoN2 + H+]+, [C12H8NiN2 + H+]+,
[o-HACPHENEN
+
3H+]+, [C18H18MnN2O2]+, [C18H18CoN2O2]+, [C18H18NiN2O2]+,
[C30H26MnN4O2 + H]+, [C30H26CoN4O2 + H]+ and [C30H26NiN4O2 + H]+, respectively, including
[M + H+]+, a pseudo-molecular ion peak. However, 2 (figure S6) exhibits peaks at m/z =
181.0721, 297.1601, 350.0705, 531.3745 and 647.4388 attributed for [C12H8N2 + H+]+,
[C18H18N2O2 + 3H+]+, [C18H18FeN2O2]+, [C30H26FeN4O2 + H]+ and [C30H26FeN4O2Cl +
2CH3CN]+, respectively. A signal observed for [M+] confirms 1:1 electrolytic nature of 2,
probably due to dissociation in solution as below:
[Fe(o-HACPHENEN)(1,10-phen)]Cl → [Fe(o-HACPHENEN)(1,10-phen)]+ + Cl¯
The presence of peak for [M + H+]+, a pseudomolecular ion peak in 1, 3 and 4 confirms
molecular mass of compounds. Similarly, in 2 the presence of signal for M+ and
[C30H26FeN4O2Cl + 2CH3CN]+ indicates its molecular mass. In FT-IR spectra of complexes, a
new peak observed in between 1635-1657 cm-1 was attributed to cyclic ν(>C=N–) moiety of
phenanthroline. A strong peak at 1608 cm-1 in the ligand for imine ν(>C=N–) groups was shifted
downwards in the complexes and observed in between 1570-1599 cm-1, indicating the
coordination of imine nitrogen to the metal. This was supported by the appearance of new signals
between 445-480 cm-1 in the IR spectra of the complexes due to M-N coordination [39]. A band
of medium-to-low intensity appearing at 3365 cm-1 in the IR spectra of the ligand assigned to
ν(O–H) vanished completely in the IR spectra of the complexes and at the same time a new peak
of medium intensity appeared between 520-569 cm-1 was assigned to new M-O bond, indicative
of the bonding of metal to the oxygen by removal of phenolic –H [40]. Complexes also exhibit
medium/sharp signals between 925-960 cm-1 attributed to ν(CH2–CH2) group of Schiff base
ligands. FT-IR spectrum of 2 is given in figure S7.
The electronic spectra of the complexes were measured in DMF. The free ligand
exhibited a strong absorption band in the range 280 nm, which was due to π→π* transition of π

electrons of the aromatic ring, and this band remains almost unchanged in the spectra of
complexes. Absorption bands in the range 340-380 nm were attributed to the n→π* transition
associated with (>C=N–) imine nitrogen. The bathochromic shift of this absorption in complexes
was due to the donation of a lone pair of electrons to the metal ion, indicating the coordination of
imine nitrogen [41, 42].
Complex 1 exhibits magnetic moment of 2.21 BM, which indicate that complex was low
spin with one unpaired electron in Mn(II) probably due to the presence of a strong-field ligand
1,10-phen. The complex exhibited three weak absorption bands at 720, 620 and 560 nm
6
6
6
4
attributed to A1g→4T1g (ν1), A1g→4T2g (ν2) and A1g→4Eg, A1g (ν3) transitions, respectively,
indicating octahedral geometry around Mn(II) ion [43]. Complex 2 exhibits a magnetic moment
of 2.62 BM which is lower than the expected value of the high-spin octahedral arrangements,
suggesting an antiferromagnetic interaction. The electronic spectra (figure S8) shows a band at
500 nm indicating an octahedral geometry around the metal ion and the band at 420 nm may be
due to charge transfer. However, it was not possible to identify the type of the d-d transition. The
magnetic moment of 3 was 2.10 BM corresponding to one unpaired electron in Co(II). The
electronic spectrum of this complex exhibits a band at 520 nm which may be assigned to the
4T1g(F)→4T1g(P) (ν3) transition in an octahedral geometry. The other two transitions
4
4T1g(F)→4A2g(F) (ν2) and T1g(F)→4T2g(F) (ν1) expected for octahedral geometry were not
observed. Transition (ν2) was not observed, being formally a two-electron transition and (ν1) was
probably due to its position beyond the scope (>800 nm) of the instrument. But the position of
the band ν3 suggests an octahedral environment around the Co(II) ion. Complex 4 exhibits a
magnetic moment of 3.01 BM indicating the presence of two unpaired electrons in Ni(II) ion
consistent with octahedral geometry with antiferromagnetic interactions. The electronic spectrum
3
of 4 displayed two bands at 425 nm and 560 nm assigned for A1g→3T1g(P) (ν3) and
3A2g→3T1g(F) (ν2) d-d transitions indicative of Ni(II) ion in an octahedral environment [44].
Since complexes were paramagnetic, NMR spectra of 1-4 were complicated and not
much helpful. Due to unpaired electron-nucleus interaction and its consequences resonances
were broad and shifted.

3.3. Molecular modelling and quantum chemical calculations
Determination of the molecular structure of complexes in the absence of single-crystal structure
by using full geometry optimizations of o-HACPHENEN and 1-4 were performed with G09
using DFT methods of B3LYP with 6-311++G(d,p) or 3-21+G* (nonmetallic atoms)/LANL2DZ
(metallic atoms) basic set in the gas phase.
3.3.1. Mulliken atomic charge analysis. The Mulliken population analyses of o-HACPHENEN
and 1-4 were done by using the method B3LYP/6-311++G(d,p) with LANL2DZ basis set in the
gas phase. The calculated Mulliken atomic charge values of selected atoms are listed in table S1.
The Mulliken charge is directly related to the vibrational properties of the molecule and indicates
how the positive and negative charges present in a molecule increase or decrease the bond
length. Distribution of Mulliken atomic charges affects dipole moment, polarizability, electronic
structure and structural parameters. The illustration of atomic charges for o-HACPHENEN and 2
is shown in figure 1 as bar diagrams. Bar diagrams of the other complexes are displayed in
figure S9. The result shows that in o-HACPHENEN, more positive charges are on C23
(+0.1563) and C24 (+0.1378) carbon due to their direct connectivity with electronegative imine
nitrogen N29 (–0.3713) and N30 (–0.3085). It was observed that more positively charged carbon
C2 (+0.1554) and hydrogen H31 (+0.3838) were connected with the more electronegative
phenolic oxygen O21 (–0.5874). In the same way, another carbon C16 (+0.18635), and hydrogen
H32 (+0.3566) were attached with other electronegative phenolic O22 (–0.5672). It is obvious
that highly shielded atoms such as oxygen and nitrogen attached to less electronegative or
positively charged atoms such as carbon and hydrogen acquire a high negative charge. However,
Mn(II), Fe(III), Co(II) and Ni(II) ion carry positive charge of +0.1536, +0.8122, +0.8110 and
+0.1788, respectively. Therefore, more electronegative two phenolic oxygen atoms and two
imine nitrogen atoms of o-HACPHENEN as well as two nitrogen atoms of 1,10-phen were most
probable coordinating points with an electropositive metal center. The carbon atoms in the
complexes mostly exhibit electropositive behavior due to the electron delocalization in the
aromatic and unsaturated bonds in the backbone of the organic ligand. In complexes, Mn, Fe, Co
and Ni metal ions of 1-4 bestow positive charge of +1.4672, +1.3245, +1.3319 and +1.3405,
respectively. Mulliken charge values of electropositive carbon and electronegative
oxygen/nitrogen were also higher than the ligand, probably due to the occurrence of back

bonding of π electrons during complexation. This result was consistent with the molecular
electrostatic potential (MEP) map [45, 46].
3.3.2. Molecular electrostatic potential analysis (MEP) and contour maps. The molecular
electrostatic potential (MEP) is related to the electron density and a very useful descriptor in
understanding sites for electron donation and electron acceptance as well as hydrogen bonding
interactions. The MEP mapped surface of the molecule was calculated by the B3LYP method
with the 6-31++G(d,p) basis set. The MEP maps of o-HACPHENEN and 2 are shown in
figure 2. MEP maps of the other complexes are given in figure S10. As can be easily observed
from the MEP map of the o-HACPHENEN, the negative regions in MEP are related to the
electronegative, while the positive regions are related to the electropositive region. The negative
regions were mainly over the oxygen atoms (deep red/yellow) on each of the O-H groups and
nitrogen atoms of imine groups. The hydrogen and carbon atoms bear the maximum region of
positive charge and the most positive regions (blue/green) were observed around the hydrogen
atoms of phenolic O-H groups as well as carbon atoms of imine groups. The color codes for this
map were in the ranges between –0.082 a.u. (deep red) to +0.082 a.u. (deep blue) in
o-HACPHENEN. These data indicate that the ligand has four possible sites for coordination. On
the other hand, metal atoms and some carbon/hydrogen atoms of the complexes show
electropositive behavior. In 1-4, MEP maps were in the range between –0.079 a.u. to +0.079 a.u.,
–0.085 a.u. to +0.085 a.u., –0.081 a.u. to +0.081 a.u. and –0.082 a.u. to +0.082 a.u., respectively.
The different values of the electrostatic potential at the surface are represented by different
colors. The potential increases in the order red < orange < yellow < green < blue [47].
3.3.3. Quantum chemical parameters. The calculated values of the following quantum
chemical parameters: the highest occupied molecular orbital energy (EHOMO), the lowest
unoccupied molecular orbital energy (ELUMO), the difference between HOMO and LUMO energy
levels, (E), Mulliken electronegativity (), absolute hardness (), absolute softness (σ),
chemical potential (Pi), global softness (S), global electrophilicity (), additional electronic
charge (Nmax), dipole moment (μ) and total energy (ETD-HF/ETD-KS) after geometrical
optimization of the structures of compounds are listed in table S2 in supplementary file.
From the above data, we can conclude that:

1. o-HACPHENEN coordinates to different metal ions through the nitrogen of imine,
deprotonated oxygen of the hydroxyl group and the two nitrogens of phenanthroline moiety.
These atoms carry more charge confirming active sites for coordination.
2. The energy difference between values of absolute softness (σ) in o-HACPHENEN and
complexes indicate that o-HACPHENEN has a good tendency to chelate metal ions.
3. The negative values of chemical potential (Pi) in all complexes indicate that energy decreases
upon accepting electronic charge form o-HACPHENEN. The value of Pi depends on the
Mulliken electronegativity ().
4. The increase in the global electrophilicity () value in all complexes, especially in 3 and 4, is
attributed to higher electron accepting capability.
3.3.4. Electronic and frontier molecular orbitals analysis. The calculated energies of the
HOMO and LUMO show that:
1. Both the EHOMO and ELUMO of 2 and 4 were attributed to beta molecular orbitals, while EHOMO
and ELUMO of 1 and 3 were attributed to beta and alpha molecular orbitals, respectively.
2. The difference of FMOs energies in complexes were such that; for EHOMO, 3β > 1α > 2 > 1β >
4 > 3α, and for ELUMO, 4 > 1β > 2 > 3α > 1α > 3β. Therefore, it is suggested that 3α has a higher
tendency of donating its HOMO electron to electron-accepting species in comparison to other
metal complexes. However, 4 was more vulnerable to electron acceptance from electron-rich
species than others.
3. Complexes 3 and 4 have low values of energy gap (E), suggesting high reactivity.
4. Complex 2 exhibits a high value of the dipole moment, which may favor their dipole-dipole
interactions with high dipole moment species, mainly in biological systems.
3.3.5. Bond length and bond angle calculations. The optimized structures of o-HACPHENEN
and 1-4 with the atomic numbering scheme have been determined. The bond lengths and angles
obtained from the energy minimum optimized structures of o-HACPHENEN and 1-4 are given
in tables S3 and S4. The molecular structures of 1-4 exhibit almost octahedral geometry around
the metal center as revealed by the calculated bond lengths and angles. The C=N, C-O and C-C
bond lengths become slightly longer in complexes as o-HACPHENEN coordinates via imine
nitrogen and phenolic oxygen of ligand. By the shifting of electron density of imine nitrogen to

metal, M-N bond was formed. The C-O bond distance in all complexes becomes longer due to
the formation of M-O bond, which makes the C-O bond weaker. This elongation in M-N and
M-O bond lengths causes a slight distortion from the regular octahedral geometries.
Additionally, the N,N′-bidentate coordination of phen in all complexes form another five-
membered metallocycle.
After coordination, some angular changes in the imine and phen moiety were observed,
i.e. widening of the C-C-O, C-N=Cimine and C-N-Cphen angles. The bond angle distortions were
observed in the coordination sphere of the complex, such as cis-Nphen-M-Nimine, Nphen-M-O and
Nimine-M-O were smaller than trans-Nphen-M-Nimine, Nphen-M-O and Nimine-M-O revealing the
octahedral geometry of the compounds [48, 49].
In an octahedral M(salen) complex, there are three probable configurations, trans
(planar), cis-α and cis-β. Due to the relatively rigid planarity of the salen skeleton, the ligand of
trans-geometry is more stable than that of cis-geometry and the complexes usually adopt trans-
configuration. It is well established here that M(salen) complexes could adopt a cis-configuration
under certain conditions, e.g. the coordination of a bidentate ligand such as o-phenanthroline
forces a M(salen) complex to adopt a cis-configuration. Although two cis-configurations (cis-α
and cis-β) are possible, the cis-β is more probable in ONNO-tetradentate ligand [20].
On the basis of the elemental analysis, ESI-MS, molar conductance, magnetic
susceptibility, electronic spectra, FT-IR and DFT, following Chem Draw and DFT optimized
structures of o-HACPHENEN and 1-4 are proposed in figures 3 and 4.
3.4. Biological applications
3.4.1. Experimental study of in vitro catalase-like activity. In a typical experiment, the
catalytic decomposition of H2O2 was exhibited by 1-4. It was observed that after 20 min of
reaction, 11.8 mL, 10.6 mL, 9.6 mL, 8.9 mL, 8.3 mL and 6.1 mL of water was displaced
corresponding to 5.20×10-4 mmol, 4.67×10-4 mmol, 4.23×10-4 mmol, 3.92×10-4 mmol,
3.66×10-4 mmol and 2.69×10-4 mmol of evolved O2 for 1-4 and o-HACPHENEN, respectively.
The progression profile of displaced water and evolved dioxygen are given in figure 5. It was
clearly observed that 1 and 2 displayed more effective catalysis than others. During the
experiment, it was also observed that solution of 1 in DMSO was brown initially, but after the
addition of H2O2, it becomes colorless which may be due to the formation of Mn(III)

intermediate [50]. Moreover, the presence of the bidentate chelating N-N′ donor ligand 1,10-phen
in the coordination sphere of the metal probably enhances the ability of the metal to
disproportionate H2O2. The electron-transfer mechanism for the catalytic reaction, the
conjugating and chelating effect of the phen ligand may make a possible formation and
stabilization of the Mn(III) intermediate [51]. Interestingly, it can be noticed easily that 2, which
contains Fe(III), was second highest in catalysis. The order of catalytic disproportionation of
H2O2 is 1 > 2 > 3 > 4.
In the second set of the experiment, the concentration of 1 was varied (from 0.003 M to
0.15 M in DMSO) and concentration of H2O2 (0.008 M) remained the same for each experiment
at room temperature. The successive profile of evolved dioxygen is given in figure 6. As it was
observed from the graph, the amount of evolved dioxygen increases with increase in the
concentration of 1. After 20 min, the maximum amount of evolved dioxygen was
5.51×10-4 mmol measured for the 0.15 M concentration of 1.
In the last set of the experiment, the concentration of H2O2 was varied (from 0.002 M to
0.008 M) and concentration of 1 (0.005 M) remained the same for each experiment at room
temperature. From this experiment, the effect of catalyst (1) on the substrate (H2O2)
concentration could be observed. The experiment conducted by using 1 as a catalyst in DMSO
with the constant concentration of [EMn] = 0.005 M and different concentrations of [H2O2] =
0.002 M, 0.004 M, 0.006 M and 0.008 M shows that after 20 min, the amount of evolved
dioxygen at different concentrations of H2O2 was 3.48×10-4 mmol, 4.27×10-4 mmol,
5.33×10-4 mmol and 5.90×10-4 mmol, respectively, as displayed in figure 7.
3.4.2. Kinetics of the H2O2 disproportionation reactions. The kinetics of H2O2
disproportionation reactions were explained by the Michaelis-Menten equation. In a
characteristic experiment, the concentration of the H2O2 was varied to determine the dependence
of the initial rate on the dioxygen evolution. The velocity for the catalase-like activity of the
metal catalysts was obtained by monitoring the decomposition of hydrogen peroxide to
molecular water by time-dependent measurements of evolved dioxygen. Michaelis-Menten
kinetic behavior of 1 was monitored by recording the rate of reaction (V) at various
concentrations of [H2O2] = 0.002-0.008 M with keeping the concentration of 1 constant at [E] =
0.005 M and evolution of dioxygen is measured at different time intervals,

ν0 = KM [S]/Vmax [S]
(1)
where [S] is the concentration of H2O2, KM is Michaelis constant and Vmax is the maximum
reaction rate measured using Lineweaver Burk plot, based on Eq. 2.
1/ν0 = Vmax/KM × 1/[S]+ 1/Vmax
(2)
Catalytic constant kcat or turnover frequency of an enzyme which demonstrates the maximum
number of substrates converting to a product by an active site at a unit of time was obtained
using Eq. 3,
kcat = Vmax/[E]
(3)
where [E] is concentration of 1.
The catalytic efficiency ε (epsilon) of an enzyme is the ratio of catalytic constant kcat and
Michaelis constant KM. The value of catalytic efficiency was calculated by Eq. 4.
ε = kcat/KM
(4)
The higher value of ε shows the more efficiency of the enzyme [51].
The enzyme kinetic parameters were determined by Lineweaver-Burk plot. A plot of 1/v0
against 1/[S] gave a straight line with the slope of KM/Vmax, at y-intercept of 1/v0 and an
x-intercept at -1/KM (figure 8). The experimental data could be fitted to the Michaelis Menten
equation, for 1, and the catalytic constant or catalytic turnover number kcat calculated was
6.7610-2 s-1 and the Michaelis constant KM was 7.1110-4 M. Since higher the value of  the
more is the efficiency of the catalyst from Eq. 4, complex 1 with  = 9.50101 shows the
efficient catalytic activity for disproportionation of H2O2. The value of KM = 7.1110-4 M for 1
was more than the values of the natural enzymes catalase (KM = 2.510-4 M). However, KM of 1
was lower in comparison to other reported enzymes i.e., Thermus thermophilus (KM
8.3010-2 M), Tricholoma album (KM = 1.5010-2 M) and Lactobacillus plantarum (KM
=
=
3.5010-1). The value of catalytic constant kcat = 6.7610-2 s-1 was low when compared to the
natural enzymes catalase (kcat = 1.0105 s-1), T. thermophilus (kcat = 2.6105 s-1), T. Album (kcat
=

2.0105 s-1) and L. plantarum (kcat = 2.6104 s-1), but found to be quite satisfactory when
compared with other reported manganese and iron complexes. Moreover the value of  =
9.50101 M-1s-1 of 1 was comparatively lower than natural enzymes catalase ( = 5.0108 M-1s-1),
T. thermophilus ( = 3.13106 M-1s-1), T. Album ( = 1.73106 M-1s-1) and L. plantarum ( =
0.57106 M-1s-1), but found to be quite similar with reported other synthetic models of
manganese and iron. The resulting values of all calculated parameters are displayed and also
compared to most of the published models in table S5 [52, 53].
3.4.3. Antibacterial assessments. The results of in vitro antibacterial activity of the Schiff base
ligand o-HACPHENEN, 1,10-phen and its 1-4 complexes are presented in table 1. Amoxicillin
and Chloramphenicol were used as positive standards and DMSO was used as negative control
for this antibacterial activity. In general, metal complexes have been shown to be more effective
than the free ligand. Complex 1 displayed the highest bactericidal activity against E. coli, which
was almost the same as the standard drugs used for comparison [58]. The other complexes show
lower activity. However, other complexes exhibit less antibacterial activity. The activity order of
the synthesized compounds was 1 > 4 > 2 > 3 > o-HACPHENEN. The higher activity of the
metal complexes may be due to the effect of metal ions on the normal cell membrane. The
variation in the activity of different complexes against E. coli depends either on the
impermeability of cells of the microbes or differences in the ribosomes of microbial cells. The
result could be explained on the basis of the chelation theory, which suggests that chelation could
promote the ability of such complexes to pass through a cell membrane [59]. The MIC of the
complexes and ligands is also displayed in the same table. Since, the MIC is the concentration of
the highest dilution tube, in which bacterial growth was absent. It was observed that 1-4 have
exhibited MIC in between 39-48 μg/mL, which was higher than standard drug chloramphenicol
but lower than amoxicillin. MIC of the complexes follows trend as 1 > 4 > 2 > 3. Antibacterial
results of some similar complexes have also been mentioned in the table. Results show that 1, 2
and 4 exhibit higher inhibition zone than the reported similar model complexes. However, the
reported model complexes have displayed lower MIC value than 1-4.

4. Conclusion
One Schiff base ligand o-HACPHENEN and four mixed-ligand mononuclear Mn(II), Fe(III),
Co(II) and Ni(II) complexes containing 1,10-phenanthroline as N,N′-donor and Schiff base as
N,O-donor have been synthesized by tailored synthesis and characterized by spectroscopic
techniques. To achieve better insight into molecular structure several quantum chemical
calculations have been performed, i.e. Mulliken charge study, the molecular electrostatic
potential (MEP), HOMO, LUMO, bond parameters and geometry of complexes was optimized.
From spectroscopy and molecular modeling, it was further ascertained that complexes have
adopted an octahedral cis-β configuration with slight distortion in both bond lengths and bond
angles. The theoretical structural parameters have shown an excellent agreement with the
experimental results and hence proved the fact of the reliability of the employed level of theory.
The synthesized compounds were tested as catalysts for H2O2 decomposition at different
concentration. Enzyme kinetics study performed on these complexes has clearly indicated that to
some extent, in solution complex 1 exactly mimics catalase-like activity. Antibacterial activity of
ligand and complexes indicate that the activity of ligand becomes more pronounced when
coordinated with the metal ions. Hence, from all these extensive observations, it is concluded
that some of these complexes could be exploited for the design of novel antibacterial drugs as
well as catalytic material.
Supplementary information
Figures S1-S10 and tables S1-S5.
Acknowledgement
The authors are grateful to Principal, Government Science College, Jabalpur and Head,
Chemistry Department, for providing necessary laboratory facilities. We sincerely thank SAIF,
1
13
CDRI, Lucknow, for recording elemental analysis, ESI-MS, H-NMR and C-NMR. One of us
(Preeti Vaidya) is also grateful to the UGC-New Delhi for financial support through RGNF
(Award letter no. F1-17.1/2015-16/RGNF-2015-17-SC-MAD-13266/(SAIII/Website)). We
gratefully acknowledge the help rendered by Dr. Usha Masram, Assistant Professor, Department
of English for checking and rechecking the manuscript for its English.

References
[1]
[2]
[3]
[4]
A.E.M. Ramadan. J. Coord. Chem., 65, 1417 (2012).
R.S. Balaban, S. Nemoto, T. Finkel. Cell, 120, 483 (2005).
F.J. Giordano. J. Clin. Invest., 115, 500 (2005).
J.M. Bolivar, S. Schelch, M. Pfeiffer, B. Nidetzky. J. Mol. Catal. B: Enzym., 134, 302
(2016).
[5]
[6]
[7]
M.E. Moustafa. Spectroscopy Lett., 38, 23 (2005).
V.S. Shivankar, N.V. Thakkar. Acta Pol. Pharm., 60, 45 (2003).
R.P. Ajay, K.J. Donde, S.S. Raut, V. Patil, R.S. Lokhande. J. Chem. Pharm. Res., 4, 1413
(2012).
[8]
[9]
P.G. Sammes, G. Yahioglu. Chem. Soc. Rev., 23, 327 (1994).
M. Munakata, S. Kitagawa, A. Ashara, H. Masuda. Bull. Chem. Soc. Jpn., 60, 1927
(1987).
[10] M. Serb, B. Calmuschi-Cula, F. Dumitru, T. Dols, U. Englet, C. Guran. Acta
Crystallogr., Sect. A, 63, 1292 (2007).
[11] N. Raman, R. Mahalakshmi. Inorg. Chem. Commun., 40, 157 (2014).
[12] F.H. Haghighi, H. Hadadzadeh, F. Darabi, H. Farrokhpour, M. Daryanavard, H.A.
Rudbari. J. Mol. Struct., 1040, 98 (2013).
[13] D. Moreno, V. Daier, C. Palopoli, J. Tuchagues, S. Signorella. J. Inorg. Biochem., 104,
496 (2010).
[14] K.C. Gupta, A.K. Sutar. Coord. Chem. Rev., 252, 1420 (2008).
[15] M.N. Esfahani, M. Montazerozohori, P. Akhlaghi. Bull. Kor. Chem. Soc., 30, 1583
(2009).
[16] S. Miriyala, I. Spasojevic, A. Tovmasyan, D. Salvemini, Z. Vujaskovic, D.S. Clair, I.B.
Haberle. Biochim. Biophys. Acta, 1822, 794 (2012).
[17] S. Sinorella, C. Hureau. Coord. Chem. Rev., 256, 1229 (2012).
[18] A. Erxleben. Inorg. Chim. Acta, 472, 40 (2018).
[19] M. Nakamura, H. Okawa, T. Inazu, S. Kida. Bull. Chem. Soc. Jpn., 55, 2400 (1982).
[20] K. Matsumoto, B. Saito, T. Katsuki. Chem. Commun., 0, 3619 (2007).
[21] T. Kurahashi. Inorg. Chem., 54, 8356 (2015).
[22] C. Rocha, O.R. Nascimento, M.D.C. Ferreira. J. Coord. Chem., 47, 479 (1999).

[23] Y. Watanabe, A. Namba, N. Umezawa, M. Kawahata, K. Yamaguchi, T. Higuchi. Chem.
Commun., 4958 (2006).
[24] G.H. Jeffery, J. Bassett, J. Mendham, R.C. Denney, Vogel’s Textbook of Quantitative
Inorganic Analysis, 5th Edn., John Wiley & Sons, Inc., New York (1989).
[25] M. Salehi, M. Amirnasr, S. Meghdadi, K. Mereiter, H.R. Bijanzadeh, A. Khaleghian.
Polyhedron, 81, 90 (2014).
[26] M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman, G.
Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato, X. Li,
H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M. Hada, M.
Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao,
H. Nakai, T. Vreven, J.A. Montgomery, Jr., J.E. Peralta, F. Ogliaro, M. Bearpark, J.J.
Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, R. Kobayashi, J. Normand, K.
Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J.M.
Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo, J. Jaramillo, R.
Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C. Pomelli, J.W.
Ochterski, R.L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth, P. Salvador, J.J.
Dannenberg, S. Dapprich, A.D. Daniels, O. Farkas, J.B. Foresman, J.V. Ortiz, J.
Cioslowski, D.J. Fox, Gaussian Inc., Wallingford, CT (2009).
[27] A.D. Becke. J. Chem. Phys., 98, 5648 (1993).
[28] C. Lee, W. Yang, R.G. Parr. Phys. Rev. B: Condens. Matter, 37, 785 (1988).
[29] M.G. Russo, E.G.V. Hissi, A.C. Rizzi, C.D. Brondino, Á.G.S. Ibañez, A.E. Vega, H.J.
Silva, R. Mercader, G.E. Narda. J. Mol. Struct., 1061, 5 (2014).
[30] M. Khajehzadeh, N. Sadeghi. J. Mol. Liq., 249, 281 (2017).
[31] F.A.I. Al-Khodir, M.S. Refat. J. Mol. Struct., 1094, 22 (2015).
[32] M.J. Pelczar, E.C.S. Chan, N.R. Krieg, Text Book of Microbiology, 5th Edn., McGraw-
Hill Publishing Company Ltd., New Delhi, India (2001).
[33] S.N. Shukla, P. Gaur, N. Rai. Appl. Nanosci., 5, 583 (2015).
[34] S.K. Bajpai, P. Jyotishi, M. Bajpai. Carbohydr. Polym., 154, 223 (2016).
[35] L.A. Saghatforoush, R. Mehdizadeh, F. Chalabian. Transition Met. Chem., 35, 903
(2010).

[36] R.M. Silverstein, G.C. Bassler, T.C. Morrill, Spectrometric Identification of Organic
Compound, 4th Edn., John Wiley & Sons Ltd. Inc, New York (1991).
[37] C. Arunagiri, A. Subashini, M. Saranya, P. Thomas Muthiah, K. Thanigaimani, I. Abdul
Razak. Spectrochim. Acta, Part A, 135, 307 (2015).
[38] W.J. Geary. Coord. Chem. Rev., 7, 81 (1971).
[39] S.M. Abdallah, M.A. Zayed, G.G. Mohamed. Arab. J. Chem., 3, 103 (2010).
[40] M.S. Sujamol, C.J. Athira, Y. Sindhu, K. Mohanan. Spectrochim. Acta, Part A, 75, 106
(2010).
[41] L. Antolini, L. Menabue, G.C. Pellacani, M. Saladini, L.P. Battaglia, A.B. Corradi, G.
Marcotrigiano. J. Chem. Soc., Dalton Trans., 2325 (1984).
[42] S. Budagumpi, M.P. Sathisha, N.V. Kulkarni, G.S. Kurdekar, V.K. Revankar. J. Incl.
Phenom. Macrocycl. Chem., 66, 327 (2010).
[43] V.A. Sawant, S.N. Gotpagar, B.A. Yamgar, S.K. Sawant, R.D. Kankariya. Spectrochim.
Acta, Part A, 72, 663 (2009).
[44] M. Shebl, S.M.E. Khalil, F.S. Al-Gohani. J. Mol. Struct., 980, 78 (2010).
[45] P.S. Raj, D. Shoba, S. Ramalingam, S. Periandy. Spectrochim. Acta, Part A, 147, 293
(2015).
[46] M. Khajehzadeh, N. Sadeghi. J. Mol. Liq., 249, 181 (2017).
[47] I. Rajaei, S.N. Mirsattari. Polyhedron, 102, 479 (2015).
[48] A.C. Ekennia, A.A. Osowole, L.O. Olasunkanmi, D.C. Onwudiwe, O.O. Olubiyi, E.E.
Ebenso. J. Mol. Struct., 1150, 279 (2017).
[49] T.A. Yousef, G.M. Abu El-Reash, R.M. El-Morshedy. J. Mol. Struct., 1045, 145 (2013).
[50] I. Kani, Ö. Atlier, K. Güven. J. Chem. Sci., 128, 523 (2016).
[51] L. Xu, S. Zhang, E.-B. Wang, R.-D. Huang. J. Coord. Chem., 56, 1329 (2003).
[52] P. Atkins, J. de Paula, Physical Chemistry, 8th Edn., W. H. Freeman and Company, New
York (2006).
[53] C. Palopoli, N. Bruzzo, C. Hureau, S. Ladeira, D. Murgida, S. Signorella. Inorg. Chem.,
50, 8973 (2011).
[54] E. Buxbaum, Fundamentals of Protein Structure and Function, 2nd Edn., Springer,
Switzerland (2015).
[55] M. Shank, V. Barynin, G.C. Dismukes. Biochemistry, 33, 15433 (1994).

[56] E.J. Larson, V.L. Pecoraro. J. Am. Chem. Soc., 113, 3810 (1991).
[57] B.M. Pires, D.M. Silva, L.C. Visentin, V. Drago, N.M.F. Carvalho, R.B. Faria, O.A.C.
Antunes. Inorg. Chim. Acta, 407, 69 (2013).
[58] A.A.A. Aziz, A.N.M. Salem, M.A. Sayed, M.M. Aboaly. J. Mol. Struct., 1010, 130
(2012).
[59] G.G. Mohamed, C.M. Sharaby. Spectrochim. Acta, Part A, 66, 949 (2007).
[60] I.P. Ejidike, P.A. Ajibade. Molecules, 20, 9788 (2015).
[61] M. Azam, S.I. Al-Resayes, S.M. Wabaidur, M. Altaf, B. Chaurasia, M. Alam, S.N.
Shukla, P. Gaur, N.T.M. Albaqami, M.S. Islam, S. Park. Molecules, 23, 813 (2018).
[62] S.M. Jadhav, V.A. Shelke, A.S. Munde, S.G. Shankarwar, V.R. Patharkar, T.K.
Chondhekar. J. Coord. Chem., 63, 4153 (2010).

Figure captions
Figure 1. Mulliken atomic charge plot of (a) o-HACPHENEN and (b) 2.
Figure 2. Molecular Electrostatic Potential (MEP) of (a) o-HACPHENEN and (b) 2.
Figure 3. (a) DFT optimized structure of o-HACPHENEN, (b) Chem Draw structure of 1,
(c) DFT optimized structure of 1, (d) Chem Draw structure of 2 and (e) DFT optimized structure
of 2.
Figure 4. (a) Chem Draw structure of 3, (b) DFT optimized structure of 3, (c) Chem Draw
structure of 4 and (d) DFT optimized structure of 4.
Figure 5. Time courses (sec) of dioxygen evolution (mmol) in the disproportionation of H2O2 by
catalase, o-HACPHENEN and 1-4 in DMSO. [Complex] = (0.005 M), [H2O2] = (0.008 M), T =
25 °C.
Figure 6. Time courses (sec) of dioxygen evolution (mmol) in the disproportionation of H2O2 by
1 in DMSO. [Complex] = 0.003 M, 0.005 M, 0.008 M, 0.05 M, 0.08 M, and 0.15 M, [H2O2] =
0.008 M, T = 25 °C.
Figure 7. Time (sec) dependence of O2 evolution (mmol) at different concentrations of H2O2.
[H2O2] = 0.002 M, 0.004 M, 0.006 M, and 0.008 M, [Complex] = 0.005 M, T = 25 C.
Figure 8. Lineweaver–Burk plot, a H2O2 saturation kinetics for 1 fitted to the Michaelis–Menten
equation with Vmax = 3.3810-4 M and KM = 7.1110-4 M.