Synthesis, characterization & biological studies of Mn(II), Fe(III) and Co(II) complexes of (Z)-1, 5-dimethyl-4-(2-(2-oxopropylidene) hydrazinyl)-2-phenyl-1H-pyrazol-3(2H)-one

Article abstract of DOI:10.1016/j.molstruc.2019.127110

Mn(II), Fe(III) and Co(II) complexes of the hydrazone derived from 4-aminoantipyrine and ethylacetoacetate has been synthesized and characterized by elemental analysis, electrical conductance in non-aqueous solvents, spectroscopic data including FT-IR, electronic, Electrospray ionization mass and M?ssbauer as well as by thermogravimetric analysis. In all the complexes, the compound acts as neutral bidentate ligand, coordinating through the pyrazolone oxygen and azomethine nitrogen. A high spin octahedral geometry assigned to the Mn(II), Fe(III) and Co(II) complexes were further confirmed by magnetic moment data. Elemental analysis showed that Fe(III) complex composed of metal and ligand in a molar ratio of 1:1 whereas metal to ligand ratio in Mn(II) and Co(II) complexes were found to be 1:2. The complexes have also been screened for their antimicrobial and antioxidant activity. The MTT assay of the Co(II) complex was also carried out. Antiviral docking studies of the ligand was done against the native and mutant HIV-1 Reverse Transcriptase protein (PDB ID: 1RT2 and 1FK9) and the ligand showed significant inhibitory activity against both forms of the protein.

Full text of DOI:10.1016/j.molstruc.2019.127110

Journal Pre-proof
Synthesis, characterization & biological studies of Mn(II), Fe(III) and Co(II) complexes
of (Z)-1, 5-dimethyl-4-(2-(2-oxopropylidene) hydrazinyl)-2-phenyl-1H-pyrazol-3(2H)-
one
R.S. Sreepriya, Shubha S. Kumar, Sadasivan V, Biju S, S.S. Meena
PII:
S0022-2860(19)31216-5
DOI:
https://doi.org/10.1016/j.molstruc.2019.127110
MOLSTR 127110
Reference:
To appear in: Journal of Molecular Structure
Received Date: 17 July 2019
Revised Date: 16 September 2019
Accepted Date: 21 September 2019
Please cite this article as: R.S. Sreepriya, S.S. Kumar, S. V, B. S, S.S. Meena, Synthesis,
characterization & biological studies of Mn(II), Fe(III) and Co(II) complexes of (Z)-1, 5-dimethyl-4-(2-(2-
oxopropylidene) hydrazinyl)-2-phenyl-1H-pyrazol-3(2H)-one, Journal of Molecular Structure (2019), doi:
https://doi.org/10.1016/j.molstruc.2019.127110.
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Synthesis, characterization & biological studies of Mn(II), Fe(III) and Co(II) complexes of
(Z)-1, 5-dimethyl-4-(2-(2-oxopropylidene) hydrazinyl)-2-phenyl-1H-pyrazol-3(2H)-one.
Sreepriya R.Sa, Shubha S.Kumara, Sadasivan Va*, Biju Sa, S.S. Meenab
a)University College, M.G.Road, Palayam, Thiruvananthapuram-695034,Kerala,India.
b)Solid State Physics Division, Bhabha Atomic Research Centre, Mumbai 400 085, India.
Abstract
Mn(II), Fe(III) and Co(II) complexes of the hydrazone derived from 4-aminoantipyrine and
ethylacetoacetate has been synthesized and characterized by elemental analysis, electrical
conductance in non-aqueous solvents, spectroscopic data including FT-IR, electronic,
Electrospray ionization mass and Mӧssbauer as well as by thermo gravimetric analysis. In all the
complexes, the compound acts as neutral bidentate ligand, coordinating through the pyrazolone
oxygen and azomethine nitrogen. A high spin octahedral geometry assigned to the Mn(II),
Fe(III) and Co(II) complexes were further confirmed by magnetic moment data. Elemental
analysis showed that Fe(III) complex composed of metal and ligand in a molar ratio of 1:1
whereas metal to ligand ratio in Mn(II) and Co(II) complexes were found to be 1:2. The
complexes have also been screened for their antimicrobial and antioxidant activity. The MTT
assay of the Co(II) complex was also carried out. Antiviral docking studies of the ligand was
done against the native and mutant HIV-1 Reverse Transcriptase protein (PDB ID: 1RT2 and
1FK9) and the ligand showed significant inhibitory activity against both forms of the protein.
Key words: Hydrazones; manganese; iron; cobalt complexes; Mӧssbauer spectroscopy;
antioxidant activity; MTT assay; Docking studies.

1. INTRODUCTION
Pyrazole is a 5-membered heterocycle containing two adjacent nitrogen atoms and pyrazolone is
a derivative of pyrazole having an additional keto group. It is an active moiety as a
pharmaceutical ingredient especially in non-steroidal anti-inflammatory drugs used for the
treatment of arthritis and other musculoskeletal and joint disorders. Among the pyrazolone
derivative, 4-aminoantipyrine is a remarkable reagent due to its importance in biological,
pharmacological, clinical & analytical applications. Derivatives of 4-aminoantipyrine are known
to exhibit a wide range of interesting biological activities like antioxidant, anti-inflammatory,
anticonvulsant, analgesic, antimicrobial, anticancer, antiprotozoal, antiparasitic, antiplatelet,
cardioprotective, antihelminthic, antidiabetic, antitubercular, anti-HIV and so forth [1].
Selection of ligands is most important in determining the properties of coordination compounds.
A ligand system having electronegative atoms like nitrogen and oxygen increases the denticity
and thus enhances the coordination possibilities. The architectural beauty of coordination
compounds is due to the interesting ligand systems containing different donor sites say ONO,
NNO, NO and NNS [2]. Among the N-O donor ligands, hydrazones possess a special place due
to their ease of synthesis, easily tunable electronic properties, denticity and formation of wide
variety of complexes with chemical, structural, biological and industrial importance [3].
Hydrazones are a special class of organic compounds in the Schiff base family having basic
structure, R1R2C=NNH2. The physical and chemical properties of hydrazones are attributed to
the active centers of hydrazone, that is, carbon and nitrogen [4]. The remarkable biological
activity of aroyl hydrazones & their mode of chelation with transition metal ions present in the
living systems have been of significant interest.
*Corresponding Author
Email id:sadasivan.v@gmail.com

In this work, we present the synthesis and spectroscopic characterization of a new hydrazone of
pyruvaldehyde with the hydrazine, 4-hydrazinyl-1,5-dimethyl-2-phenyl-1H-pyrazole-3(2H)-one,
prepared via an indirect method involving Japp-Klingemann reaction and its complexes with
Mn(II), Co(II) and Fe(III) metal salts. Also present the evaluation of their antioxidant,
antimicrobial and anti tumour activities. In silico antiviral study of hydrazone against HIV-1
Reverse transcription protease have also been carried out.
2. Experimental
2.1 Materials and Methods.
Reagent grade chemicals were used without further purification unless otherwise noted.
All chemicals used were of highest available purity. They include ethyl acetoacetate and 4-
aminoantipyrine (Merck). All other reagents and solvents used were of analytical grade (AR).
2.2 Method of preparation of ligand
The ligand (Z)-1, 5-dimethyl-4-(2-(2-oxopropylidene) hydrazinyl)-2-phenyl-1H-pyrazol-3(2H)-
one (EAP) was synthesized via the Japp-Klingemann reaction [5] (Scheme 1). About 28 g of
KOH (0.5 M) wasadded to 6.4 ml ethyl acetoacetate (0.05M) with stirring and the mixture was
allowed to stand at room temperature for 24 hrs. 4-amino antipyrine was diazotized using NaNO2
and con.HCl at 0°C. The diazonium salt was then added to the solution of acidified potassium
acetoacetate at 0°C slowly with stirring. The solid that separated was collected by vacuum
filtration and dried in a dessicator. The yield of the crude product was found to be 86%.
Purification was effected by recrystallization from ethanol and crystals of suitable size
wereselected and used for X-ray diffraction studies.
*Corresponding Author
Email id:sadasivan.v@gmail.com

O
O
O
O
HCl
KOH
H3C
OK
H3C
OEt
Ethyl acetoacetate ethyl 3ꢀoxobutanoate
Potassium acetoacetate
C6H5
O
N
N
N2+Cl-
O
O
O
H3C
CH3
COOH
H3C
OH
H3C
3ꢀoxobutanoic acid
O
C6H5
N HC
N
N
N
C
CH3
O
H
H3C
CH3
(Z)ꢀ1,5ꢀdimethylꢀ4ꢀ(2ꢀ(2ꢀ
oxopropylidene)hydrazinyl)ꢀ2ꢀphenylꢀ1Hꢀpyrazolꢀ
3(2H)ꢀone
Scheme 1. Synthesis of the ligand EAP.
2.3. Preparation of complexes.
Complexes were prepared by refluxing aqueous ethanolic solutions of MnCl2.4H2O,
FeCl3.6H2O and CoCl2.6H2O (0.05 mol) with that of ligand (0.05 mol) for 4 hrs. On cooling, the
*Corresponding Author
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solid complex separated were filtered and washed with ethanol and petroleum ether. It was then
dried in a dessicator.
2.4. Physical methods of analysis.
The elemental analysis (C, H, N and S) were made using Vario El III Carlo Erba 1108 elemental
analyzer. The molar conductance of the complexes (10-3 M solutions) in DMSO and acetonitrile
were measured at room temperature using a Systronic Direct Reading Conductivity meter. The
complexes were analyzed for their metal and halide content by standard methods[6]. The melting
points of the ligand and complexes were determined by open capillary method. The magnetic
moments of the complexes were measured at room temperature on a Sherwood scientific
magnetic susceptibility meter. IR spectra of the ligand and the complexes wererecorded using
KBr discs on a SHIMADZU IR Prestige-21 Spectrometer in the range 400-4000 cm-1. The
electronic spectra were recorded on a Perkin Elmer Lambda 25 UV-Visible spectrophotometer in
the range 200-800 nm. The mass spectral analysis of the ligand was done on Thermo Scientific
Q-Exactive (ESI mode) and that of complexes were done on Thermo Finnigan LCQ Advantage
max LC-MS/MS Ion Trap Spectrometer. TG and DTG curves were recorded using Perkin Elmer
STA 6000 thermal analyzer. Mӧssbauer spectrum of the iron(III) complexwas recorded using a
Mӧssbauer spectrometer operated in constant acceleration mode (triangular wave) in
transmission geometry at room temperature. The SXRD study of the ligand was done on Bruker
Kappa Apex II diffractometer making use of soft wares SAINT (APEX II) for frame integration
and SHELXTL for structure solution and refinement.
2.5. Biological studies.
*Corresponding Author
Email id:sadasivan.v@gmail.com

The in vitro antibacterial and antifungal activity of the ligand EAP and its Mn(II), Fe(III) and
Co(II) complexes were performed through the Agar well diffusion method using Streptomycin
and Clotrimazole as positive control for bacteria and fungi respectively. The two bacteria
selected were Staphylococcus aureus and Escherichia coli and the pathogenic fungi were
Candida albicans and Aspergillus niger.
The antioxidant properties of the free ligand and the Mn(II), Fe(III) and Co(II) complexes
werestudied using DPPH (2, 2-diphenyl-1-picrylhydrazyl) assay. The viability of cells was
evaluated by direct observation of HeLa cells by Inverted phase contrast microscope and
followed by MTT assay method using mononuclear cells. The binding conformations between
the EAP and the protein targets (PDB ID: 1RTZ, 1JLB, 3BGR, 1FK9) were analyzed with the
help of Auto Dock 4.0 Tools.
RESULTS AND DISCUSSION
The EAP and its complexes are non-hygroscopic solids, stable in air and soluble in common
organic solvents. The purity of the hydrazone is ascertained by elemental analysis. The micro
analytical data obtained is found to be C (61.68%), H (5.43%), N (20.40%). This agrees well
with the theoretical data C (61.99%), H (5.53%), N (20.66%) and corresponds to the molecular
formula C14H15N4O2. The analytical data of the complexes along with their molar conductance
values are given in Table 1. The molar conductance of the above complexes in DMSO and
acetonitrile shows that all are electrolytes. The Fe(III) complex is of 1:1 type and the Mn(II) and
Co(II) complexes are of 1:2 type electrolyte. The melting point of the ligand is found to be 165°C
and that of complexes are found to be more than 200°C.
*Corresponding Author
Email id:sadasivan.v@gmail.com

Table 1.Analytical and conductance data of the obtained complexes.
Complex
Yi
eld
%
C
H
N
Cl
Metal Ω-1cm2 mol-1
DMSO Aceto
nitrile
76 61.68 5.43 20.40
61.99 5.53 20.66
-
-
-
-
-
-
EAP
Calculated
-
-
85 48.75 4.84 16.38 10.22 8.09
75
133
[Mn(EAP)2Cl(H2O)]Cl
Calculated
48.99 4.66 16.33 10.35 8.00
81 35.49 4.15 11.90 22.52 11.87 89
35.79 4.05 11.93 22.69 11.90
152
155
[Fe(EAP)(Cl)2(H2O)2]Cl
Calculated
85 48.59 4.58 16.48 10.12 8.33
91
[Co(EAP)2Cl(H2O)]Cl
Calculated
48.72 4.64 16.24 10.29 8.51
3.1. IR SPECTRA
The IR spectrum of the free ligand EAP, exhibits a strong band at 1686 cm-1which corresponds
to the stretching vibration of the ν(C=O).This band is shifted to a lower wave number of about 30
cm-1 in all the three complexes indicating that the carbonyl group of the pyrazolone ring is
coordinated to the transition metal ion.The strong band observed at 1635 cm-1 in EAP is
attributed to the ν(C=N) mode of the azomethine group. This undergoes a blue shift of 40 cm-1 in
complexes which suggests that the azomethine group is co-ordinated to the metal ion in the
complexes [7]. The appearance of ν(C-H) and δ(C-H) of the –CH2 group at lower frequency in the
*Corresponding Author
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complexes compared to their positions in the free ligand could also be taken as evidence for the
co-ordination nature of the azomethine nitrogen. The selected IR bands are given in Table 2.
A weak broad band around 3061 cm-1due to the stretching frequency of intramolecularly bonded
–NH in ligand is found to appear without much shift in complexes [8]. This shows that
complexation occurs without deprotonation and hence EAP acts as a neutral bidentate ligand in
the chloro complexes of Mn(II), Fe(III) and Co(II).The presence of coordinated water is
suggested by the very broad absorption band centered at 3435, 3390 and 3458cm-1. It is further
confirmed from the presence of rocking and wagging vibrational modes of coordinated water
around 950-974 cm-1 and 760-780 cm-1. Bands in the region 550-640 cm-1 are associated with M-
O stretching vibrations. The M-N stretching vibrations in complexes are seen in the region of
450-505 cm-1 [9].
Table 2.Important IR spectral bands (cm-1) of the complexes.
EAP [Mn(EAP)2Cl(H2O)]Cl [Fe(EAP)(Cl)2(H2O)2]Cl
[Co(EAP)2Cl(H2O)]Cl Assignment
3435
3390
3458
-
ꢀ(OH)
1686
1635
1654
1583
1668
1590
1656
1575
ꢀ(C=O)
ꢀ(C=N)
3.2. Electronic spectra
The electronic spectra of the ligand and complexes were recorded at 300K in ethanol in the range
200-800 nm. The n-π* transition which is observed at 345 nm in ligand undergoes a blue shift in
*Corresponding Author
Email id:sadasivan.v@gmail.com

complexes which indicates that the lone pair of electrons of oxygen and nitrogen are coordinated
to the metal ion [10]. The weak shoulder found in the UV-Vis spectrum of Mn(II) complex may
be due to the overlapping of one spin forbidden d-d transition with the strong and broad
absorption of the ligand [11]. The Fe(III) complex showed a weak and broad band at 630 nm
6
which is assignable to A1g→4T2g transition. In Co(II) complex, two bands of low intensity
4
4
observed at 530 nm and 670 nm are attributed to T1g(F)→4T1g(P) and T1g(F)→4A2g(F)
respectively. The analysis of the absorption bands of the above complexes support an octahedral
geometry around the corresponding metal ions in their complexes [12]. The absorption regions,
assignments and the proposed geometry of the complexes are given in Table 3.
Table 3.Electronic spectral data and magnetic moments of the obtained complexes.
Magnetic
Absorption
moment
Compound
region
(nm)
Band Assignment
Geometry
(BM)
345
471
n-π*
-
EAP
423
470
420
630
409
530
670
n-π*
d-d transition
n-π*
[Mn(EAP)2Cl (H2O)]Cl
[Fe(EAP) Cl2 (H2O)2]Cl
[Co(EAP)2Cl(H2O)]Cl
5.72
5.84
3.72
Octahedral
Octahedral
Octahedral
6A1g →4T2g
n-π*
4T1g(F) →4T1g(P)
4T1g(F) →4A2g(F)
3.3. Mӧssbauer spectrum of [Fe(EAP)(H2O)2Cl2]Cl.
*Corresponding Author
Email id:sadasivan.v@gmail.com

The most valuable information concerning bond properties, anisotropy, electron
population of molecular orbitals and local structure may be extracted from the quadrupole
splitting.The quadrupole splitting (QS) of the resonance lines arises from the electric field
gradient at the nucleus, as a consequence of asymmetry in the electronic environment. Therefore,
in principle, the QS values are useful for elucidating the geometry of the complexes and the
values assist in examining the donor and acceptor properties of the ligand. The quadrupole
splitting decreases with increase in the symmetry of the central iron atom [13]. TheMӧssbauer
parameters of the synthesized Fe(III) complex are given in Table 4.
The spectrum of the present Fe(III) complex shows a doublet which indicates the
presence of single species.The quadrupole splitting at room temperature suggest that an electric
field gradient is persisting around due to the assymetric ligand environment around the central
metal ion. It is interesting to note that the quadrupole splitting in this complex is high. This
seems plausible in view of the asymmetric geometry of the Fe(III) ion. The investigation of this
complex has been carried out to understand the position of the metal (Mӧssbauer nucleus) within
the complex under investigation Fig.1. In complex, the Mӧssbauer spectrum reveals that the
surrounding ligands are distorted and, therefore, appears as quadrupole splitting.
Table 4 . Mӧssbauer parameters
Sample
Fe site
IS
QS
LW(mm/s) Relative Fitting
(mm/s) (mm/s)
area
(%)
100
quality
parameter(χ2)
1.04552
[Fe(EAP)Cl2(H2O)2]Cl Doublet 0.364
0.673
0.434
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.
Fig.1.Mӧssbauer spectrum of [Fe(EAP) Cl2(H2O)2]Cl recorded at room temperature.
The value of isomer shift (IS) (0.364mm/s) for the complex is consistent with the
formulation of iron atom as high spin ferric ion [Fe3+] and well within the range observed for
related high spin ferric complexes [14].The magnitude of the isomer shift depends on the total ‘s’
electron density at the resonant iron nucleus, which is related to the degree of covalency of the
bonds. The increase in the s electron density is attributed to increased back bonding from the iron
atom to the ligand that is compensated by a synergistic flow of electron density to the iron. In
iron compounds the isomer shift decreases with an increase in the electron density at the nucleus.
The more covalent complexes have lower isomer shift. For the complexes where the iron(III)
ligand bonding is more ionic an isomer shift of 0.5 mm/swith respect to iron metal is observed
[15]. Thus the lower values of isomer shift indicates substantial covalency for metal ligand bond
in Fe(III) complex. For the present Fe(III) complex, the value of isomer shift lie at 0.364 mm/s
with respect to natural iron, typically observed for other high spin Fe(III) complexes.
3.4.Thermal Analysis.
The Fe(III) complex undergoes a three stage decomposition process as shown in Fig. 2.
The complex is thermally stable up to 245°C. The first stage of decomposition starts at 245°C
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and ends at 280°C with a mass loss of 8.89%. The observed mass loss is due to the
decomposition of two water molecules which is in agreement with a calculated mass loss of
7.65%.The maximum rate of mass loss occurs at 254°C as indicated by the DTG peak. The
second stage starts at 280°C and ends at 390°C with the DTG peak at 340°C.The corresponding
mass loss of 29.6% is attributed to the decomposition of part of ligand molecule and two chlorine
atoms. The third stage starts at 430°C and comes to an end at 495°C with DTG peak at 480°C
and the corresponding mass loss of 42.01% is due to decomposition of remaining portion of
ligand molecule. The mass loss at second and third stages is in agreement with a calculated mass
loss of 29.69% and 43.2% respectively. The decomposition completes at 510°C and the final
residue weight found (14.78%) may be due to the anhydrous metal oxide and the remaining can
be accounted for the volatilization of the residue of anhydrous metal chloride [16].
1
100
0
-1
80
-2
60
40
20
0
-3
-4
-5
-6
-7
-8
100
200
300
400
500
600
700
Temperature (°C)
*Corresponding Author
Email id:sadasivan.v@gmail.com

Fig. 2.TG-DTG curve of [Fe(EAP)Cl2(H2O)2]Cl
The Co(II) and Mn(II) complexes decompose in two stages. A dip within 100°C in the
TG-DTG curve of Co(II) complex (Fig. 3) can be due to the loss of lattice water and 2% water
loss in the range 140-175°C can be due to the removal of a coordinated water molecule.There is
a characteristic mass loss in the range 220-260°C due to the loss of two chlorine atoms and is in
agreement with the calculated mass loss of 10.01%.The decomposition from 460-550°C with a
mass loss of 80.3% and a DTG peak at 518°C can be attributed to the loss of two ligand
molecules and is in agreement with the calculated mass loss of 78.72%. The decomposition
completes at 570°C and the final residue of 10.16% corresponds to the metal oxide (CoO)
against the calculated value of 10.84%.
0.0
100
-0.2
80
-0.4
-0.6
-0.8
-1.0
-1.2
-1.4
60
40
20
0
100
200
300
400
500
Temperature (°C)
Fig.3. TG-DTG Curve of [Co(EAP)2Cl(H2O)]Cl
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The decomposition of Mn(II) complex within 100°Cindicates the presence of
lattice water, whereas a 2% weight loss in the range from 135-150°C is indicative of the
loss of coordinated water molecule. The decomposition range from 220°C-270°C
corresponds to a mass loss of 10.12% with a DTG peak at 253°C. This is in agreement
with the calculated value of 10.34% and may be accounted for by the evolution of two
chlorine atoms. The second stage starts from 460°C and ends at 540°C with a DTG peak
at 525°C.This is due to the loss of two ligand molecules and corresponds to a mass loss
of 80% which very well agrees with the theoretical value of 79.01%. The total mass loss
upto 575 °C leaves a 9.82% residual weight, which is in agreement with the final product
(cal. 10.34%) at the end of the Mn(II) complex decomposition.The TG-DTG curve of
Mn(II) complex is as shown in Fig. 4.
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Fig.4. TG-DTG curve of [Mn(EAP)2Cl (H2O)]Cl
3.5. Mass spectra
The FAB mass spectrum of synthesized ligand is recorded and the obtained molecular ion
peak confirms the proposed formulae.The mass spectrum of EAP shows molecular ion peak at
m/z 304, corresponding to [C14H16N4O2+MeOH]+.[Fig. 5].Also the spectrum exhibits the
fragments at m/z 272 corresponding to [C14H16N4O2]+., m/z 217 corresponding to [C11H12N4O]+.
which is an [M+H] peak and m/z 202 to give [C11H12N3O]+. moiety.
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Fig.5.Mass spectrum of EAP.
The M+ peak corresponding to m/z 652 of Mn(II) complex obtained after the loss of chlorine
atom outside the coordination sphere is in agreement with the formula [Mn(EAP)2Cl(H2O)]+.
[Fig. 6]. The peak at m/z 380 corresponds to the loss of a ligand molecule [Mn(EAP)Cl(H2O)]+.
whereas the peak at m/z 216 is obtained for [C11H12N4O]+. obtained after the loss of manganese,
water, chlorine and (CH3-CO-CH-) group. Finally the peak at m/z 202 corresponds to the
[C11H12N3O]+. moiety.
*Corresponding Author
Email id:sadasivan.v@gmail.com

Fig.6. Mass spectrum of [Mn(EAP)2Cl(H2O)]Cl.
The spectrum of Fe(III) complex shows molecular ion peak at m/z 517, which is an adduct peak
of [C14H20N4O4FeCl2+2Na+2H2O] + and corresponds to [M+H] peak. The other fragment peaks
are seen at m/z 480, 388, 215& 201 which corresponds to [C14H20N4O4FeCl2+2Na]+.
[C14H20N4O4Fe+Na]+.,[C11H11N4O]+.and
[C11H11N3O]+. ie:
4-aminoantipyrine
moiety
respectively. The m/z of all the fragments of ligands and their complexes confirm the
stoichiometry of the Fe(III) complex as [Fe(EAP)Cl2(H2O)2]Cl [Fig: 7 ].
*Corresponding Author
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Fig.7. Mass spectrum of [Fe(EAP)Cl2 (H2O)2]Cl
The M+ peak in cobalt(II) complex corresponds to m/z 656.5 which is in agreement with the
formula [Co(EAP)2Cl(H2O)]+.. The other important peaks of appreciable intensity are seen at m/z
values 451.15 and 389.19. The peak at m/z 215 is for [C11H11N4O]+. obtained after the loss of
cobalt, water, chlorine and (CH3-CO-CH-) group and the peak at m/z 201 is for [C11H11N3O]+.
moiety [Fig. 8].
*Corresponding Author
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.
Fig.8.Mass spectrum of [Co(EAP)2Cl(H2O)]Cl
3.6. SXRD study of ligand.
The ligand crystallized by slow evaporation from ethanol is found belonging to monoclinic P21/c
space group, and unit cell parameters of a = 10.5492(8) Å, b = 8.8669(6) Å, c = 15.5125(10) Å,
α= γ= 90°, β
=104.868 (3)o V = 1402.44(18) Å3, and Z = 4. A summary of the crystallographic
data and details of the data collection of EAP is shown in the Table 5. Atom coordinates, thermal
parameters, and torsion angles are provided as supporting information. Selected bond lengths and
angles are presented in Table 6. All bond lengths and angles were within normal ranges.
Table 5.Crystal data and structure refinement for EAP.
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Identification code
Empirical formula
Formula weight
Temperature
EAP
C14 H16 N4 O2
272.31
296(2) K
Wavelength
0.71073 Å
Crystal system
Space group
Monoclinic
P21/c
Unit cell dimensions
a = 10.5492(8) Å
b = 8.8669(6) Å
c = 15.5125(12) Å
α
β
= 90°
= 104.868(3) °
γ
= 90°
Volume
3
1402.44(18) Å
4
Z
Density (calculated)
3
1.290 Mg/m
Absorption coefficient
-1
0.090 mm
F(000)
576
Crystal size
3
0.200 x 0.150 x 0.100 mm
2.669 to 24.997°.
-12<=h<=12,
Theta range for data collection
Index ranges
-10<=k<=10,
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-18<=l<=18
Reflections collected
16181
Independent reflections
Completeness to theta = 24.997°
Absorption correction
2478 [R (int) = 0.0388]
100.0 %
Semi-empirical
equivalents
from
Max. and min. transmission
Refinement method
0.7455 and 0.6686
Full-matrix least-squares on
2
F
Data / restraints / parameters
2478 / 0 / 185
1.063
2
Goodness-of-fit on F
Final R indices [I>2 sigma(I)]
R1 = 0.0527,
wR2 = 0.1177
R1 = 0.0896,
wR2 = 0.1483
n/a
R indices (all data)
Extinction coefficient
Largest diff. peak and hole
-3
0.326 and -0.193 e.Å
Table 6. Selected bond lengths and bond angles
Bond lengths (Å)
N (3)-N (4)
1.33(3)
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N (3)-H (3A)
0.93(3)
Bond angles (º)
C (9)-C (8)-N (3)
130.1(2)
119.1(3)
120.5
N (4)-C (12)-C (13)
N (4)-C (12)-H (12)
N (4)-N (3)-C (8)
119.8(2)
119(2)
N (4)-N (3)-H (3A)
C (8)-N (3)-H (3A)
121(2)
C (12)-N (4)-N (3)
118.4(2)
-179.2(3)
178.5(3)
-176.6(3)
-178.0(3)
2.0(5)
C (4)-C (5)-C (6)-N (1)
C (2)-C (1)-C (6)-N (1)
O (1)-C (7)-C (8)-C (9)
N (4)-C (12)-C (13)-O (2)
N (4)-C (12)-C (13)-C (14)
C (9)-C (8)-N (3)-N (4)
C (7)-C (8)-N (3)-N (4)
C (13)-C (12)-N (4)-N (3)
C (8)-N (3)-N (4)-C (12)
-25.6(5)
158.0(3)
179.2(3)
176.9(3)
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Fig.9. ORTEP Diagram of EAP
The ORTEP diagram of the ligand is shown in Fig. 9. The distance between N3 and C8
[1.394(3) Å] is intermediate between the classical lengths of single [C-N (1.43 Å)] and double
[C=N (1.28 Å)] bonds, which describes the imine bridged moiety as an effective electron
conjugated substructure found to be within the normal reported range [17]. The C7-O1 bond
length of 1.237(3) Å is found to be within the normal reported range. The distance between C8
and C9 in the pyrazolone ring is 1.356(4) Å, indicating a double bond character. The dihedral
angle between the planes of the antipyrine and phenyl rings (C5-C6-N1-C7) is -61.3(4)o, and the
dihedral angles of C5-C6 -N1-N2 and C1-C6-N1-N2 are 138.4(3)oand -41.8(4)°, respectively.
The O1 atom is slightly displaced from the antipyrine mean plane and the dihedral angles of O1-
C7-C8-C9 is -176.6(3)o. The dihedral angle of C(11)-C(9)-N(2)-C(10) is found to be 41.9(4)o.
The azomethine double bond C(12)-N(4) is found to be 1.290(3) Å which is in close agreement
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with the normal value. The dihedral angles of N(4)-C(12)-C(13)-O(2) and N(4)-C(12)-C(13)-
C(14) are -178.0(3)° and 2.0(5)° respectively. These deviations from planarity are believed to
have been caused by intermolecular interactions in the crystal lattice [18].
Table 7. Hydrogen bonds for EAP [Å and °]
D-H...A
d(D-H) d(H...A) d(D...A)
<(DHA) Symmetry
codes
C(10)-H(10B)...O(2) 10.96
C(11)-H(11B)...O(2) 20.96
2.55
2.35
3.392(4)
3.298(4)
2.849(3)
146.9
171.2
172(3)
x,-y+1/2
-x,-y,-z+1
-x+1,-y,-+1
N(3)-H(3A)...O(1) 30.93(3) 1.93(3)
The crystal is mainly stabilized by intermolecular hydrogen bonds involving N-H and methyl
protons with oxygen atoms. It is given in Table 7. C(10)-H(10B)...O(2), C(11)-H(11B)...O(2),
N(3)-H(3A)...O(1) with interatomic distance of 3.392 (4) Å (x,-y+1/2) , 3.298(4) Å (-x,-y,-z+1),
2.849 (3) Å (-x+1,-y,-+1 ). As expected, the N–H⋅⋅⋅O bonds are stronger as compared to the C–
H⋅⋅⋅O bonds [19]. Based on the above studies, the proposed structures of the Fe(III), Mn(II) and
Co(II) complexes are given in Fig.10 & 11.
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Cl
H3C
H3C
N
H2O
N
O
Fe
N
H
N
H
H2O
O
C
C
Cl
CH3
Fig.10. Structure of [Fe(EAP)Cl2(H2O)2]Cl.
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Fig.11. Structure of [M(EAP)2Cl(H2O)]Cl (M=Mn2+ or Co2+).
3.7. Antimicrobial assessments
The in vitro antimicrobial activity of the investigated ligand and its selected metal complexes
were tested against the bacteria, S. aureus and E. coli and fungi C. albicans and A. niger through
the Agar well diffusion method [20]
.
The Mn(II), Fe(III) and Co(II) complexes have more
The ligand does not have any activity
antibacterial and antifungal activity than the ligand
.
whereas the complex has notable activity against C. albicans under the same experimental
conditions. However, both the ligand and the complexes do not have any activity against A.
niger. The antimicrobial activity of the ligand and complexes are shown in the Table 8.
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Table 8.Antimicrobial activities of EAP and complexes.
Compound
Bacteria
S. aureus
Fungi
E. coli
13
C. albicans
EAP
NZ
15
18
16
NZ
9
[Mn(EAP)2ClH2O]Cl
[Fe(EAP)Cl2(H2O)2]Cl
[Co(EAP)2ClH2O]Cl
14
16
13
11
15
3.8. Antioxidant Studies
The results of the antioxidant studies were compared with ascorbic acid, the standard antioxidant
[21] and shown in Table 9. The antioxidant activity of the Mn(II), Fe(III), Co(II) complexes of
EAP are found to be less when compared to the activity of free ligand[22]. Among the prepared
complexes, cobalt complex is found to have more anti-oxidative property than the corresponding
Mn(II) and Fe(III) complexes (Fig. 12). The percentage of inhibition of protein denaturation
iscalculated as follows:
ꢁAbsꢂꢃꢄꢅꢆꢃꢇ– Absꢈꢉꢊꢋꢇꢌ
Absꢂꢃꢄꢅꢆꢃꢇ
ꢍ
Percentage inhibition =
× 100
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Fig.12.Graphical representation of antioxidant activities of EAP and complexes
Table9.Antioxidant activity of EAP and complexes.
Compound
Concentration(ꢀg/ml)
500 750
250
1000
EAP
0.419
30.59
0.278
51.74
0.240
58.39
0.230
61.12
OD
% of inhibition
[Mn(EAP)2ClH2O]Cl
OD
0.916
29.46
0.739
37.31
0.672
49.52
0.573
55.50
% of inhibition
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