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Synthesis, Characterization, Biological and
Antitumor Activity of Co(II) , Ni (II) , Cu(II)
and Zn (II) Complexes of
Ali M Hassan1, Zaghloul I Elbialy2 and Khalid M Wahdan3*
1Department of Chemistry, Al Azhar University, Egypt
2Egyptian Organization for Standardization and Quality
3 Researcher at the Faculty of Science, Al Azhar University, Egypt
Submission: May 20, 2020; Published: June 04, 2020
*Corresponding author: Khalid M Wahdan, Researcher at the Faculty of Science, Al Azhar University, Egypt
How to cite this article: Ali M H, Zaghloul I , Khalid M W. Synthesis, Characterization, Biological and Antitumor Activity of Co(II) , Ni (II) , Cu(II) and Zn
(II) Complexes of N-(2-Chlorophenyl)-N’-Benzoyl Thiourea. Organic & Medicinal Chem IJ. 2020; 9(4): 555767. DOI: 10.19080/OMCIJ.2019.09.555767
A derivative of thiourea ligand N-(2-chlorophenyl)-N’-Benzoyl thiourea (CBT) in equimolar ratio 1:1and its transition metal complexes CoII, NiII, CuII and ZnII were synthesized by microwave (green chemistry). The structure of ligand and its complexes have been characterized by using elemental analysis, mass Spectroscopy, FT-IR, UV-Vis,1HNMR and 13CNMR. The geometry of the proposed structures of the chelates based on their electronic spectra, electron spin resonance (ESR) and magnetic susceptibility. The stability of complexes was studied by TGA analysis (Thermal studies). The free derivative thiourea ligand (CBT) and its complexes were Studied for antibacterial and antifungal activity. Also the anticancer activity of ligand and its complexes were studied against breast cell cancer (MCF-7). It was noticed that the NiII complex showed the highest cytotoxic activity (IC50=122) more than the free CBT ligand.
Thiourea (NH2)2C=S is a compound where the oxygen atom of urea compound replaced by a Sulphur atom. The compounds bearing carbonyl and thiocarbonyl groups are used as potential donor ligand for the preparation of complexes [1,2]. Among these, Thiourea and its derivatives are versatile ligands that coordinate to form stable compounds. They are able to coordinate with metal either as neutral or monoanion or dianion ligand [3,4]. These thiourea ligands and their metal complexes exhibit a wide range of biological activity. They are reported to act as antimicrobial, antibacterial, antifungal, antimalarial, antituberculosis and anticancer activities [5,6]. They also form a variety of complexes of different symmetries with various metal ions [7,8]. In view of the importance of thiourea and their derivatives it was worth interesting to synthesize N-substituted thiourea ligand and their complexes with transition metal elements because it was observed that this activity was enhanced by complexion with certain transition metal elements [3,9].
Materials and physical measurements: All purchased chemicals were of Annular AR grade and were obtained from Sigma Aldrich and all Metals salts were purchased from ADWIC .The Microwave-assisted synthesis was carried out in a domestic microwave energy output 900W. Purity of Schiff base ligand and its complexes were detected by using thin-layer chromatography (TLC) technique. Melting points were recorded in open capillaries with Barnstead Thermolyne Mel-temp 1001D Electrothermal Melting Point. Elemental analysis was done on automatic analyzer CHNS Vario El III-Elementar, Germany. The FT-IR spectra samples were ground with (CsBr) powder. Then pressed into a disk and recorded on Shimadzu FTIR spectrometer. Mass spectra were determined by using Mass GC-2010 Shimdazu instrument. Metals content were determined by complexometric titration using xylene orange (XO) as indicator and hexamine as a buffer (pH = 6).
Electronic absorption spectra in DMF were measured using
automated UV/Vis-NIR 3101 PC Shimadzu spectrophotometer
ranged from 200-900nm. 1HNMR spectra for Schiff base ligand
was recorded in 300MHz Varian-Oxford Mercury in DMSO-d6 as
solvent and the chemical shifts were recorded in ppm relative to
TMS as an internal standard. Magnetic susceptibility of complexes
was measured on powdered samples using the faraday method.
Thermal analysis measurements (TGA) were carried out with
Shimadzu thermal analyzer model 50 at Micro analytical .The
ESR spectra of the powdered Cu(II) complex recorded at room
temperature by X-band EMX spectrometer (Bruker, Germany)
using a standard rectangular cavity of ER 4102 with 100KHz
frequency.. Schiff base ligand and their metal complexes were
screened for in-vitro antibacterial activity against two species of
Gram-positive bacteria and two species of Gram-negative bacteria
as well as two species of fungi. Also, Cytotoxicity evaluation was
applied against breast cell cancer (MCF-7). All of these were
carried out in faculty of Science, Cairo University.
All consumed chemicals were from analytical grade and were
used as received without further purifications. Chemicals used
are Benzoyl chloride, ammonium thiocyanate, o-chloro analin,
acetone, cobalt acetate, nickel acetate, copper acetate, zinc acetate
0.1m of ammonium thiocyanate(7.6gm) dissolved in 50ml
of acetone then added drop by drop to 0.1m of benzoyl chloride
(14.06gm) (11.62ml) taken in 3 neck flask with continuous
stirring. The mixture is refluxed for 1 hour with continuous stirring.
After 45 minutes white ppt (ammonium chloride) appeared and
then disappeared at 1hr. The mixture left in room temp until
the precipitate appears again completely. Filtration done and
precipitate washed by acetone to get all the filtrate (Benzoyl
thiocyanate). The filtrate added drop by drop in a 3-neck flask
contains 0.1m (10.5ml) of ortho chloro analin with continuous
stirring. The mixture refluxed for 6hrs with continuous stirring.
The mixture transferred to a baker and covered for two days for
complete precipitation. Then the Precipitate was filtrated and
washed by ethanol and acetone to give the pure ligand (CBT).
The prepared ligand and the acetate salts of the metal
Co(CH3COO)2.4H2O, Ni (CH3COO)2.4 H2O,Cu(CH3COO)2.H2O and Zn
(CH3COO)2.2H2O were mixed in (1:1) ratio. The reaction mixtures
were then irradiated by the microwave oven by using drops of
methanol as a solvent. The reaction was completed in a short time
(3-5min) with higher yields. The resulting product washed by
hot methanol and ether and finally dried under reduced pressure
over anhydrous CaCl2 in a desiccator. The progress of the reaction
and purity of the product was monitored by TLC using silica gel
(yield: 80-85%). The synthetic route of the prepared compounds
is illustrated in Scheme 1.
The structure of the prepared ligand N-(2-chlorophenyl)-
N’-Benzoyl thiourea (CBT) was characterized by melting point,
elemental analysis, IR, mass spectra, UV-vis and NMR studies.
This data is compatible with the required product. Analytical and
physical properties of prepared compounds tabulated in Table 1.
The characteristic IR bands of all thiourea ligands showed the
expected frequencies of υ(C=O), υ (N-H), υ(C-N) and υ(C=S). The
coordinative behavior of present ligand, towards Zn(II) and Ni (II) ions to form complexes is difficult to establish, as this ligand is
capable of exhibiting three tautomeric forms as clear from Scheme
2 due to presence of [‒NH‒C(=S)] and [‒NH‒C(=O)] functional
groups. However, the lack of the characteristic vibrations of υ(S–H)
around 2500‒2600cm-1  and presence of a peak at 3312cm-1
characteristic of υ(N–H)  confirmed the absence of tautomeric
form (N=C‒SH) and (N=C-OH). Two sharp intense bands observed
at 1669 and 1329 cm-1can be ascribed to the stretching vibration
of carbonyl group υ(C=O) and thionyl group υ(C=S) respectively
These observations confirmed the ketonic-thion form of the
ligand in the solid-state . Moreover, the υ(C–Cl) stretching
frequency was observed at 753cm-1, while this band appearing at
678-686cm-1 assigned to the usual modes of phenyl ring vibration,
respectively [15,16]. On the other hand and upon coordination
of the metal center to ligand, the characteristic bands of υ(C=O)
and υ(C=S) present in the spectrum of the free ligand at 1669
and 1329cm-1 were found to be shifted to a lower frequency and
appear from 1597 to 1666 for C=O and from 1242 to 1288cm-1 for
C=S. This finding may be taken as an evidence for the coordination
of the carbonyl oxygen and thionyl sulphur atoms with the metal
ions. The IR spectra for ligand and complexes are shown in Figure
1-3 and the data are tabulated in Table 2.
The recorded mass spectra of the ligand and its complexes
(Scheme 3) showed a peak at m/z (relative abundance)
2902.7(39.57%) which is the molecular peak of (C14HH1N2OSCl).
The ion peak at m/e = m/z 212.55(42%) is due to M+(C8H5N2OSCl),
while the ion peak at m/e = 145.79 (100%) corresponds to
M+(C6H6SCl) which is the base peak, the ion peak at m/e =
120.17 (47%) corresponds to M+(C7H6ON), the ion peak at m/e
= 171 (56.75%) is due to M+(C7H5NSCl), the ion peak at m/e =
113.88 (22.77 %) points to M+(C6H5CL). The ion peak at m/e =
50.68(38.79%) refers to M+ (C4H2), the ion peak at m/e = 78.9
(21.5%) corresponds to M+ (C6H6). The fragmentation pattern
of N-(2-chlorophenyl)-N’-Benzoyl thiourea is shown in Scheme 3
and Figure 2.
The mass spectra of the CBT-Zn complex (Scheme 4) showed
peak at m/z (relative abundance) 510.9 (21.06%) as the molecular
peak of (Zn.C18H21N2O7S). The ion peak at m/e = m/z 400 (45.9
%) is due to M+(Zn.C16H14NO3S), while the ion peak at m/e 325
(35.48%) corresponds to M+(ZN.C10H9O3S), the ion peak at m/e =
385.5 (55.5%) corresponds to M+(Zn.C16H17ON2S), the ion peak at
m/e = 336.89 (49.9%) is due to M+(Zn.C15H14ON2S), the ion peak
at m/e = 319.34 (49.6 %) points to M+( Zn.C14H11ON2S). The ion
peak at m/e = 243(48.9%) refers to M+ (Zn.C8H6ON2), the ion peak
at m/e = 97.2(38.58%) corresponds to M+ (Zn-S) M+ (C6H6SCl),
the ion peak at m/e =52.6(100%) REFERS TO (C2N2) which is the
base peak, the ion peak at m/e =44.7(35.63) REFERS TO (CS). The
fragmentation pattern of ZnII complex is shown in Scheme 4 and
The assignments of the main single in the  HNMR spectra
of the ligand given in Figure 5 recorded in DMSO-d6 as a solvent
with TMS internal standard, displays some groups of signals
corresponding to the various protons . The chemical shifts
were expressed in ppm. The spectrum of the ligand (CBT) give two singlet signals, one at δ 11.78ppm assigned to (S,1H, N8-H) and
the other at δ 12.742ppm assigned to(S,1H, N10-H)  (which
were also identified by D2O exchange) . The multiplets observed
at δ 7.293 - 8.098ppm are attributed to the phenyl protons .
The 13CNMR spectra of the synthesized ligand (CBT) showed
peaks at 180.219 ppm assigned to (C=O), 168.556ppm assigned to
(C=s), 135.357ppm (C11) 133.239ppm(C16), 131.858 ppm (C5),
129.483ppm(C2) and at 128.793ppm(C12), 128.261ppm(C13),
128.110ppm(C1 and C3), 127.935 ppm(C4 and C6), 127.222
The stereochemistry of the metal ions in the complexes can
be assigned via the electronic spectral measurements. The diffuse
solid reflectance spectra of the ligand and its complexes in solidstate
showed a number of bands in the UV -Vis region (200-
Three absorption bands were observed at 257.5, 275 and
310nm. The first band can be assigned to n −π * transitions
originated from aromatic moieties, and the third band can be
assigned to n −π * transitions originated from C=O and C=S groups.
The latter band is due to the intermolecular charge transfer
interaction from aromatic groups to C=O and C=S group [20,21].
The electronic spectra of the copper complexes revealed bands
from 425 to 750nm assigned to the transitions ( ) 2 2 2 B → E . The
measured magnetic moments, 1.78B.M, falls in the range reported
for tetrahedral geometry.
Absorption bands from 560 to 720 (nm) which can be assigned
to metal ligand charge transfer MLCT in a low spin tetrahedral
geometry of Zn (II) complex confirmed by the diamagnetic
properties. The assignments of the observed electronic transitions
apart from that together with the geometry and the magnetic
moment values [23, 24] are listed in Table 3.
The most essential features of the spectrum of the complex are
entirely different when compared to that of the free ligand. Upon
interaction of the ligand with metal ions, there are some shifts in
bands positions in the spectra of the complexes. This change can
be taken as a positive evidence of complex formation. In addition,
appearance of new bands at longer wavelength assigned to ligand
metal charge transfer (LMCT) and d-d transitions is another
confirmation to complex formation, which gives evidence for the
coordination of the ligands to the metal ions. All spectra data were
tabulated in Table 3 and represented in Figure 6.
ESR spectroscopy is a technique for studying chemical species
that have one or more unpaired electrons through measuring the
absorption of electromagnetic radiation by a molecular system
containing one or more unpaired electron, such as organic and
inorganic free radicals or inorganic complexes possessing a
transition metal ion and rare earth metals. The spectra of copper
complex exhibit single broad signal with hyperactive structure
indicating the contribution of free acetate ligand with complex
formation. The spectra showed broad signals with two “g” values
(g\\, g┴) Figure 7. For all complexes the value of g\\ < g┴ < 2.3,
characteristic of complexes with 2B1 (dx2-y2) orbital ground state.
The average g values were calculated according to the equation
gav = 1/3[g\\ +2g┴] and it was equal to 1.397 for CBT-Cu.
TGA data of the thermal decomposition of the prepared
complexes are shown in Table 4, Figures 8- 10 and Scheme 5. The
TGA curves indicate that the loss of weight starts around 205°C
and continues to about 300°C at which point most of the organic
part of the compounds have been lost. This sharp decomposition
period brings about 78% weight loss in the complexes and led to
the complete formation of sulphide.
Some chelates exhibited a moderate inhibitory activity of
complexes than that of the corresponding free ligands. The free
ligand (CBT) and its metal complex ZnII in addition to the standard
drugs were screened separately for their antibacterial activity
against Staphylococcus aureus (ATCC:6538), streptococcus
mutans (ATCC:25175)(Gram-positive bacteria), Escherichia Coli
(ATCC:9637) and Klebsiella Pneumonia (ATCC:10031) (Gramnegative
bacteria) and antifungal activity against Aspergillus
Nigar (ATCC:32856) and Candida albicans (ATCC:6538)
fungi. The antimicrobial activity against the growth of various
microorganisms was determined by measuring the inhibition
zone in millimetersaround the well, also the activity index data
was calculated . The result is recorded in Table 5.
As one can see from the observation results that the growth
of microorganism was more inhibited by the metal complexes as
compared with the organic ligand (CBT) and the standard drugs.
The metal complexes act as more powerful bactericides and
fungicides agents and they may serve as a vehicle for activation
of ligand where the metal ions being more hypersensitive against
the microbial cells. This behavior of the metal complexes may
be a result of modification in structure upon coordination and
formation of metal organic framework and can be explained based
on the overtone concept and chelation theory. In general, the
easy penetration of the metal complexes into lipid membranes,
disturbance of the respiration process of the cell and blocking the
synthesis of proteins are restrict further growth of the organism
and lead to enhance of activity of metal complexes compared with
the organic ligand [26,27].
The cytotoxicity activities of free ligand (CBT) and Ni (II)
complex were tested against (MCF-7) human tumor cell lines. For
comparison purpose, the cytotoxicity of Doxorubicin as stander
antitumor drug was evaluated and produced (IC50 ml) under the
same conditions. The reported results in terms of IC50 value was
recorded in Table 6. The cytotoxic activity was evaluated by using
different concentrations and reported in Table 7 & Figure 11.
As depicted, the ligand and its complex has very good cytotoxic
activity compared with the standard. These finding could be
explained by the solubility effect as good relationship could be
seen between activity and solubility of compounds. The activity
of the complex could be explained by its greater solubility and
lipophilicity increase with increasing bulkiness and may be
facilitate transport through the cellular membrane .
To conclude, complexes were successfully synthesized and
fully characterized by chemical and spectroscopic methods. Then
the antibacterial and antifungal effect studied and compared
between the ligand itself and the ZnII complex and the anticancer
effect studied of its NiII complex and compared with the ligand and
standard drugs. The study showed that the ligand and its metal
complexes possess an appreciable activity and can consider as
an effective inhibitor towards the different microbial strains and
cancer. Generally, such activity enhanced upon complexation
where metal complexes show better activity than their parent
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