Synthesis, spectral characterization, in-vitro microbiological evaluation and cytotoxic activities of novel macrocyclic bis hydrazone

Article abstract of DOI:10.1016/j.ejmech.2009.03.032

A macrocyclic hydrazone Schiff base was synthesized by reacting 1,4-dicarbonyl phenyl dihydrazide with 2,6-diformyl-4-methyl phenol and a series of metal complexes with this new Schiff base were synthesized by reaction with Co(II), Ni(II) and Cu(II) metal salts. The Schiff base and its complexes have been characterized by elemental analyses, IR, ¹H NMR, UV-vis, FAB mass, ESR spectra, fluorescence, thermal, magnetic and molar conductance data. The analytical data reveal that the Co(II), Ni(II) and Cu(II) complexes possess 2:1 metal-ligand ratios. All the complexes are non-electrolytes in DMF and DMSO due to their low molar conductance values. Infrared spectral data suggest that the hydrazone Schiff base behaves as a hexadentate ligand with NON NON donor sequence towards the metal ions. The ESR spectral data shows that the metal-ligand bond has considerable covalent character. The electrochemical behavior of the copper(II) complex was investigated by cyclic voltammetry. The Schiff base and its complexes have also been screened for their antibacterial (Escherichia coli, Staphylococcus aureus, Shigella dysentery, Micrococcus, Bacillus subtilis, Bacillus cereus and Pseudomonas aeruginosa) and antifungal activities (Aspergillus niger, Penicillium and Candida albicans) by MIC method. The brine shrimp bioassay was also carried out to study their in-vitro cytotoxic properties.

Full text of DOI:10.1016/j.ejmech.2009.03.032

European Journal of Medicinal Chemistry 44 (2009) 3552–3559
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis, spectral characterization, in-vitro microbiological evaluation
and cytotoxic activities of novel macrocyclic bis hydrazone
,
Prakash Gouda Avaji a, C.H. Vinod Kumar b, Sangamesh A. Patil c, K.N. Shivananda d, C. Nagaraju b
*
a Department of Organic Chemistry, Indian Institute of Science, Bangalore 560012, Karnataka, India
b Department of Chemistry, S.V. University, Tirupathi 517502, Andhra Pradesh, India
c Department of Chemistry, Karnatak University, Dharwad 580003, Karnataka, India
d Department of Chemistry, Bangalore University, Bangalore 560001, Karnataka, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A macrocyclic hydrazone Schiff base was synthesized by reacting 1,4-dicarbonyl phenyl dihydrazide with
2,6-diformyl-4-methyl phenol and a series of metal complexes with this new Schiff base were synthe-
sized by reaction with Co(II), Ni(II) and Cu(II) metal salts. The Schiff base and its complexes have been
characterized by elemental analyses, IR, 1H NMR, UV–vis, FAB mass, ESR spectra, fluorescence, thermal,
magnetic and molar conductance data. The analytical data reveal that the Co(II), Ni(II) and Cu(II)
complexes possess 2:1 metal–ligand ratios. All the complexes are non-electrolytes in DMF and DMSO due
to their low molar conductance values. Infrared spectral data suggest that the hydrazone Schiff base
behaves as a hexadentate ligand with NON NON donor sequence towards the metal ions. The ESR
spectral data shows that the metal–ligand bond has considerable covalent character. The electrochemical
behavior of the copper(II) complex was investigated by cyclic voltammetry. The Schiff base and its
complexes have also been screened for their antibacterial (Escherichia coli, Staphylococcus aureus, Shigella
dysentery, Micrococcus, Bacillus subtilis, Bacillus cereus and Pseudomonas aeruginosa) and antifungal
activities (Aspergillus niger, Penicillium and Candida albicans) by MIC method. The brine shrimp bioassay
was also carried out to study their in-vitro cytotoxic properties.
Received 17 August 2008
Received in revised form
26 February 2009
Accepted 12 March 2009
Available online 5 April 2009
Keywords:
Synthesis
Characterization
Biological
Toxicity
Hydrazone
Ó 2009 Elsevier Masson SAS. All rights reserved.
1. Introduction
mental disorder [1]. Tuberculostatic activity is attributed to the
formation of stable chelates with transition metals present in the
Hydrazones are a special group of compounds in the Schiff base
family. They are characterized by the presence of pC]N–N]Co.
The presence of two inter-linked nitrogen atoms was separated
from imines, oximes, etc. Hydrazone Schiff bases of acyl, aroyl and
heteroacroyl compounds have an additional donor sites like C=O.
The additional donor sites make them more flexible and versatile.
This versatility has made hydrazones good polydentate chelating
agents that can form a variety of complexes with various transition
and inner transition metals and have attracted the attention of
many researchers.
Various hydrazones are obtained depending on the experi-
mental conditions; which have application as biologically active
compounds [1] and as analytical reagents [2]. As biologically active
compounds, hydrazones find applications in the treatment of
diseases such as anti-tumor [3], tuberculosis [4], leprosy and
cell. Thus many vital enzymatic reactions catalyzed by these tran-
sition metals cannot take place in the presence of hydrazones [5–7].
Hydrazones also act as herbicides, insecticides, nematocides,
rodenticides and plant growth regulators.
Hydrazones are used as plasticizers and stabilizers for polymers,
polymerization initiators, antioxidants, etc., they act as intermedi-
ates in preparative chemistry. In analytical chemistry, hydrazones
find application in detection, determination and isolation of
compounds containing the carbonyl group. More recently, they
have been extensively used in detection and determination of
several metals. They also find applications as indicators and spot
test reagents [2].
Recently, a number of attempts have been made to obtain
Mn(III), Fe(III) and Co(III) complexes with bis hydrazone formed by
condensation of isatin monohydrazone with 2-hydroxy-1-naph-
thaldehyde, which have been well characterized by elemental
analyses and spectral data [7]. Fe(III), Co(II), Ni(II), Cu(II), Zn(II) and
UO2(VI) complexes have been synthesized by the condensation of
* Corresponding author. Fax: þ91 877 2249611.
E-mail address: cnraju.962@rediffmail.com (C. Nagaraju).
0223-5234/$ – see front matter Ó 2009 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2009.03.032

P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
3553
2-amino-4-hydrazino-6-methyl pyrimidine with o-hydrox-
yacetophenone [8] and Mn(II) complexes have been synthesized
from the bis-Schiff bases of 2,6-diacetylpyridine and anthraniloyl
hydrazide/benzoyl hydrazide/salicyloyl hydrazide [9]. All these
compounds have been characterized by elemental analyses,
magnetic, thermal, IR, 1H NMR, UV–vis, FAB mass, ESR and X-ray
spectral data.
hydrazone Schiff base derived from 1,4-dicarbonyl phenyl dihy-
drazide with 2,6-diformyl-4-methyl phenol. This Schiff base has
donor sites with the NON NON sequence and varied coordination
abilities. This nature of the Schiff base (Scheme 1) has attracted our
attention and aroused our interest in elucidating the structure of
Co(II), Ni(II) and Cu(II) complexes. These are evaluated for their
antibacterial and antifungal properties against various pathogenic
bacterial strains using the minimum inhibitory concentration
method.
A survey of the literature reveals that no work has been carried
out on the synthesis of metal complexes with macrocyclic
O
O
EtOH
H2N-HN-C
C-NH-NH2
2
NH2NH2. H2O
C2H5OOC
Diethyl terephthalate
COOC2H5
+
Reflux, 3-4h
Hydrazinhydrate
CH3
EtOH
O
O
O
C
Reflux, 3 h
O
C
OH
H
H
HN
N
NH
N
HCl (3 drops)
H3C
OH
CH3
OH
N
N
HN
C
O
C
O
NH
Co
Ni
/
Cl H O
2 6
2
Reflux, 3 h
EtOH
Cl
2 6
O
O
H
H
/
2
2
Cu
2
Cl2
CH3
CH3
O
O
NH
HN
NH
C
N
II
N
HN
C
N
OH2
N
Cl
Cl
M
O
O
C
O
C
O
O
.
(H2O)2
II
.
Ni
(H2O)2
Ni
M
Cl
C
O
O
C
C
O
C
OH2
Cl
N
NH
HN
N
NH
N
HN
N
O
O
M= Co(II) & Cu(II)
CH3
CH3
Scheme 1.

3554
P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
2. Chemistry
3. Pharmacology
2.1. Methods
3.1. In-vitro antibacterial and antifungal assays
2.1.1. Synthesis of diethyl terephthalate
The biological activities of the newly synthesized Schiff base
and its metal complexes have been studied for their antibacterial
and antifungal activities by disc diffusion method [13,14]. The
antibacterial activities were done by using the following organ-
isms (Escherichia coli, Staphylococcus aureus, Shigella dysentery,
Micrococcus species, Bacillus subtilis, Bacillus cereus and Pseudo-
monas aeruginosa). These bacterial strains were chosen as they
are the known pathogens of human body and Aspergillus niger,
Penicillium and Candida albicans were used for antifungal activi-
Terephthalic acid (1.6 g) in super dry ethanol (60 mL) containing
2–3 drops of concentrated H2SO4 (AR) was refluxed till it dissolves.
Then, the reaction mixture was poured onto ice cold water,
immediately a solid started separating from the clear solution. To
this a solution of sodium bicarbonate was added till the efferves-
cence seized. The ester thus obtained was filtered and washed with
water for several times (mp 44 ꢀC).
2.1.2. Synthesis of dihydrazide of terephthalic acid
ties at 10, 25, 50 and 100 m
g mLꢁ1 concentrations in solvent DMF.
A mixture of diethyl ester of terephthalic acid (2.22 g) and
hydrazine hydrate (98% 2 cc) in ethanol was refluxed for 4–5 h. The
reaction mixture was allowed to cool to room temperature then,
the cooled solution was poured onto ice cold water. The dihy-
drazide of terephthalic acid thus obtained was filtered and
recrystallized from ethanol.
The bacteria were subcultured in agar medium. The Petri dishes
were incubated for 24 h at 37 ꢀC. Standard antibacterial drug
(Gentamycin) was also screened under similar conditions for
comparison. The fungi were subcultured in potato dextrose agar
medium. Standard antifungal drug (Fluconazole) was used for
comparison. The Petri dishes were incubated for 48 h at 37 ꢀC
(Figs. 1 and 2).
2.1.3. Preparation of 2,6-diformyl-4-methyl phenol
2,6-Diformyl-4-methylphenol was prepared by reported
method [10–12].
3.1.1. Minimum inhibitory concentration (MIC)
Compounds showing promising antibacterial/antifungal activity
were selected for minimum inhibitory concentration studies. The
minimum inhibitory concentration was determined by assaying at
2.1.4. Synthesis of macrocyclic terephthalic acid hydrazone
Schiff base
100, 50 and 25 m
g mLꢁ1 concentrations along with standards at the
2,6-Diformyl-4-methylphenol (2 mmol) in ethanol (20 mL) was
added to an ethanolic solution of 1,4-dicarbonyl phenyl dihydrazide
(2 mmol, 30 mL) containing a few drops of concentrated HCl. The
reaction mixture was refluxed for 3 h. The mixture was cooled to
room temperature and the solvent removed under reduced pres-
sure until a solid product was formed that was washed with cold
ethanol and dried under vacuum. Yield 70–75%.
same concentrations.
3.1.2. Pharmacology results
The microbial results of Schiff base and its Co(II), Ni(II) and Cu(II)
complexes are systematized in Table 1. The biological activity of the
Schiff base exhibited a considerable enhancement on coordination
with the metal ions against all fungal strains. However, the metal
complexes showed good antifungal activity against A. niger,
Penicillium and C. albicans. It was evident from the data that this
activity significantly increased on coordination. This enhancement
in the activity may be rationalized on the basis that their structures
mainly possess an additional C]N bond. It has been suggested that
the Schiff base with nitrogen and oxygen donor systems inhibit
enzyme activity, since the enzymes which require these groups of
their activity appear to be especially more susceptible to deacti-
vation by metal ions on coordination. Moreover, coordination
2.1.5. Synthesis of Co(II), Ni(II) and Cu(II) complexes
A mixture of ligand (0.01 mol) and metal chloride (0.02 mol)
was refluxed for an hour in alcohol. To the reaction mixture sodium
acetate (0.02 mol) in water was added and the reflection was
continued for another 2 h. The complex was precipitated by adding
distilled water. The complex separated was filtered, washed with
water, then with hot alcohol and finally dried in vacuum desiccator
over P2O5 (yield 60–70%).
Fig. 1. Photograph (Bacillus subtilis) showing antibacterial screening of Schiff base and Gentamycin.

P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
3555
Fig. 2. Photograph (A. niger) showing antifungal screening of Schiff base and Fluconazole.
reduces the polarity [15,16] of the metal ion mainly because of the
partial sharing of its positive charge with the donor groups [17,18]
within the chelate ring system formed during coordination. This
process, in turn, increases the lipophilic nature of the central metal
atom, which favors its permeation more efficiently through the
lipid layer of the microorganism [19–21] thus destroying them
more aggressively.
Some important factors such as the nature of the metal ion,
nature of the ligand, coordinating sites, geometry of the complex,
concentration, hydrophilicity, lipophilicity and presence of co-
ligands have considerable influence on antibacterial activity.
Certainly, steric and pharmacokinetic factors also play a decisive
role in deciding the potency of an antimicrobial agent. Apart from
this, the mode of action of these compounds may also invoke
hydrogen bond through the pC]N–N]CH– group with the active
centers and thus interfere with normal cell process. The presence of
lipophilic and polar substituents is expected to enhance antibac-
terial activity. Heterocyclic ligands with multifunctionality have
a greater chance of interaction either with nucleoside bases (even
after complexation with metal ion) or with biologically essential
metal ions present in the biosystem and can be promising candi-
dates as bactericides since they always tend to interact especially
with some enzymatic functional groups, in order to achieve higher
coordination numbers [22]. Thus antibacterial property of metal
complexes cannot be ascribed to chelation alone, but it is an
intricate blend of several contributions.
3.1.3. In-vitro cytotoxicity
The synthesized Schiff base and its Co(II), Ni(II) and Cu(II)
complexes were screened for their cytotoxicity (brine shrimp
bioassay) by using the protocol of Meyer et al. [17]. Brine shrimp
(Artemia salina leach) eggs were hatched in a shallow rectan-
gular plastic dish (22 ꢂ 32 cm) filled with artificial seawater,
which was prepared with a commercial salt mixture and double
distilled water. An unequal partition was made in the plastic dish
with the help of a perforated device. Approximately 50 mg of
eggs were sprinkled into the large compartment, which was
darkened while the minor compartment was open to ordinary
light. After two days nauplii were collected by a pipette from the
lighted side.
A sample of the test compound was prepared by dissolving
20 mg of each compound in 2 mL of DMF. From this stock solution
100, 50 and 10
m
g mLꢁ1 were transferred to nine vials (three for
each dilutions were used for each test sample and LD50 is the
Table 1
Bacteriological results of Schiff base and its Co(II), Ni(II) and Cu(II) complexes at 10
m
g mLꢁ1 concentration.
Empirical formula
Activity against bacteria (mm)
Activity against fungi (mm)
E. coli
P. aeruginosa
B. cereus
S. aureus
S. dysentery
Micrococcus
B. subtilis
A. niger
Penicillium
C. albicans
C34H28N8O6
16
10
14
12
20
–
14
16
12
14
20
–
16
16
14
10
20
–
10
12
10
12
20
–
16
14
16
16
20
14
12
16
10
20
20
12
16
14
20
22
20
20
18
–
20
18
18
20
–
16
18
18
20
[Co(C34H26N8O6)Cl2]2H2O
[Ni(C34H26N8O6)Cl2$2H2O]2H2O
[Cu(C34H26N8O6)Cl2]2H2O
Gentamycin
Fluconazole
DMF
24
12
24
12
24
12
12
12
12
12
12
12
12
Key for interpretation: less than 12 mm – inactive; 12–14 mm – weakly active; 14–16 mm – moderately active; above 16 mm – highly active.

3556
P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
mean of three values) and one vial was kept as control having
2 mL of DMF only. The solvent was allowed to evaporate over-
night. After two days, when shrimp larvae were ready, 1 mL of
seawater and 10 shrimps were added to each vial (30 shrimps/
dilution) and the volume was adjusted with seawater to 5 mL per
vial. After 24 h the number of survivors was counted. Data were
analyzed by a Finney computer program to determine the LD50
values [23]. In the case of cytotoxic activity it was observed that
the Schiff base and its Co(II), Ni(II) and Cu(II) complexes displayed
weak cytotoxic activity against A. salina, therefore they can be
considered non-cytotoxic.
in the region 6.80–7.75 ppm due to aromatic protons. A signal in the
region 2.50–2.82 ppm is assigned to methyl protons [30]. All these
observations support the infrared conclusions.
4.3. Electronic spectral studies
The electronic spectra of Co(II) complex exhibit absorption
bands in the region 11015, 17212 and 20 032 cmꢁ1 which can be
4
4
assigned to the transitions 4A2 þ E / 4B1, 4A2 þ E / 4E (P) and
4A2 þ E / 4A2 (P) respectively which are characteristic of square
4
pyramidal geometry [31]. The Ni(II) complex shows three bands
around 10 882, 16 752 and 26 342 cmꢁ1 which are assigned to 3A2g
3
3
4. Results and discussion
(F) / 3T2g (F) (n1), A2g (F) / 3T1g (F) (n2) and A2g (F) / 3T1g (P)
(n3) transition respectively, indicating octahedral geometry [31].
All the Co(II), Ni(II) and Cu(II) complexes are colored non-
hygroscopic solids, stable in air. They are sparingly soluble in
common organic solvents, but soluble in DMF and DMSO. The
elemental analyses show that the Co(II), Ni(II) and Cu(II) complexes
have stoichiometry of the type [M2LX2]$2H2O (M ¼ Co(II), Cu(II)
and X ¼Cl) and [Ni2LX2(H2O)2]$2H2O. The molar conductance
values are too low to account for any dissociation of non-electro-
lytes in DMF.
The ligand field parameters are listed in Table 2. The Cu(II) complex
exhibited a high intensity band at 27 523 cmꢁ1 in the UV region.
Appearance of this band is due to
p /
p* transition associated with
the azomethine linkage and L / M charge transfer transition. The
electronic spectrum of Cu(II) complex shown three bands at 10 952,
14798 and 23 228 cmꢁ1, these bands have been assigned to the
transition 2B1 / 2A1 n1), 2B1 / 2B2 n2) and 2B1 / 2E (n3) respec-
( (
tively. These transitions are characteristic of square pyramidal
geometry [32].
4.1. IR spectra
4.4. Magnetic studies
The important infrared frequencies of Schiff base and its Co(II),
Ni(II) and Cu(II) complexes. It is well established that the Schiff base
having O-hydroxy group either on aldehyde or on aniline residue
can form intramolecular hydrogen bond [24,25]. This has direct
The magnetic moments of the Co(II), Ni(II) and Cu(II) complexes
obtained at room temperature are listed in Table 3. The Co(II) and
Ni(II) complexes shown magnetic moment values around 4.28 and
3.34 BM respectively which are lower than the respective spin
values only and indicates weak antiferromagnetic coupling inter-
action between the metal ions which further confirm the dinuclear
nature of the complexes. The Cu(II) complex showed a magnetic
moment around 1.45 BM which is considerably lower than the spin
only value for Cu(II) complexes. The low value of the magnetic
moment is attributed to the antiferromagnetic coupling interaction
between two metal ions. This fact suggests the dinuclear nature of
the Cu(II) complexes [33].
impact on the n(OH) vibration, and shifts to the lower frequency
with broadening. The extent of shift depends on the strength of
hydrogen bonding.
The broad band at 3439 cmꢁ1, a strong band at 1626 and
1687 cmꢁ1 in the IR spectrum of the Schiff base are assigned to H-
bonded –OH stretching,
n
n(C]N) of azomethine and carbonyl
(C]O) vibrations, respectively. An intense band at 3100 cmꢁ1 is
due to the –NH– vibrations of the hydrazine group, a broad medium
intense band was observed at 2921 cmꢁ1 due to methyl groups and
the band at 1068 cmꢁ1 is assigned to hydrazinic
ligand.
n(N–N) of the free
4.5. FAB mass spectrum
For the Co(II), Ni(II) and Cu(II) complexes, we observed the
following changes; the high intense band due to phenolic –OH
appeared in the region at 3439 cmꢁ1 in the Schiff base was dis-
appearedinthecomplexes. Theseobservationssupporttheformation
of M–O bonds via deprotonation. So the H-bonded –OH groups have
been replaced by the metal ion. The presence of broad stretching
vibrations in the 3598–3570 cmꢁ1 region can be attributed to coor-
dinated or lattice water molecules in all these complexes.
The FAB mass spectrum of the Schiff base has been depicted in
Fig. 3. The spectrum showed a molecular ion peak at m/z 645 which
is equivalent to its molecular weight [L þ H]þ.
The FAB mass spectrum of Cu(II) complex is depicted in Fig. 4.
The spectrum showed a molecular ion peak Mþ at m/z 876, which is
equivalent to its molecular weight of the Cu(II) complex. The
molecular ion peak fragmentation with the loss of two water
molecules, gave a peak A1 at m/z 840 due to the fragment ion
[Cu2(C34H26N8O6)Cl2]þ. Further, the fragments leading to the
formation of the species [Cu2L]þ which undergo demetallation to
form the species [L þ H]þ gave fragment ion at m/z 644. All these
fragmentation patterns are well observed in the FAB mass spec-
trum. These entire fragmentation patterns are well observed in the
FAB mass spectrum (Fig. 4). It clearly indicates dinuclear nature of
the complex and two Cu(II) ions are held in the macrocyclic
compartment of the Schiff base. The metal ions are bonded to two
phenoxo bridges, which endogenously coordinated to the metal
ions and the other coordinating sites in the ligand are the azome-
thine nitrogen atoms. Both phenoxide and azomethine groups
surround the two metal ions, which are in close proximity within
the ligand molecule to form a square base. A chloride ion coordi-
nates to each metal ion from opposite sides to give square pyra-
midal configuration to the metal ions. The FAB mass spectrum
confirms the dinuclear nature of the metal complex.
The medium intense band at 1601 cmꢁ1 due to
n
(C ¼ N) indi-
cates that the C]N of the ligand coordinates to the metal through
nitrogen. A broad band at 3565 cmꢁ1 in addition to a medium to
high intense band at 850 cmꢁ1 is attributed to the symmetric and
antisymmetric-OH stretching modes of coordinated water [26]. The
unaltered position of the
involvement in coordination. The
been assigned in the region 540–550 cmꢁ1 and 425–450 cmꢁ1
n
(C]O) (carbonyl) confirms non-
n
(M–O) and (M–N) bands have
n
,
respectively [27,28].
4.2. 1H NMR spectra
The Schiff base exhibits signals at 13.42 ppm due to NH protons
[29]. These protons are D2O exchangeable and confirm the
assignment. It also exhibits resonance due to phenolic –OH protons
around 10.20 ppm [30]. The other characteristic resonance due to
azomethine proton in the Schiff base appears at 8.20 ppm. Signals

P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
3557
Fig. 3. FAB mass spectrum of Schiff base.
4.6. Thermal studies
molecules, two aldehyde moieties and two bis hydrazine moieties
respectively.
TG and DTG studies were carried out for Co(II), Ni(II) and Cu(II)
complexes. These complexes decompose gradually with the
formation of respective metal oxide above 500 ꢀC. The nature of
proposed chemical change with the temperature range and the
percentage of metal oxide obtained are given in Table 4. The
thermal decomposition of respective Co(II) and Cu(II) complexes
takes place in three steps as indicated by DTG peaks around
110–120, 200–210 and 240–250 ꢀC corresponding to the mass loss
of two coordinated water molecules, two aldehyde moieties and
two bis hydrazine moieties respectively, Fig. 5. Whereas, one
representative Ni(II) complex decomposes in four steps as indicated
by DTG peaks around 200–210, 230–235, 280–290 and 410–450 ꢀC
corresponding to the mass loss of two coordinated/adhered water
4.7. ESR studies
The X-band ESR spectrum of Cu(II) complex was recorded at
room temperature using DPPH as a reference standard. The g and
gt values have been found to be 2.25 and 2.09 respectively. In
general, dinuclear Cu(II) complexes give broad ESR peaks and the
broadening is assigned to a dipolar interaction [34]. The observed
ESR spectrum is characteristic of square pyramidal geometry. The
k
gav value was calculated to be 2.14. The existence of g > gt
k
suggests that dx2ꢁy2 orbital is in the ground state and d9 configu-
ration is (eg)4(a1g)2(b2g)2(b1g)1. The ‘g’ values are related to the axial
symmetry and g > gt suggests square pyramidal geometry of
k
Fig. 4. FAB mass spectrum of Cu(II) complex.

3558
P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
Fig. 5. TG/DTG spectrum of Ni(II) complex.
Cu(II) complex. The axial symmetry parameter G ¼ 2.542 which has
less than 4.0 indicates considerable exchange interaction between
metal ions in the solid complex [35] which further supports the
dinuclear nature of the Cu(II) complex.
10ꢁ3 M solution was recorded at room temperature in the potential
range ꢁ3.0 V to þ2.4 V with a scan rate 100 mV sꢁ1. The complex
shows a redox process corresponding to the Cu(II) / Cu(I) couple at
Epa ¼ þ1.40 V and associated cathodic peak at Epc ¼ ꢁ0.60 V. This
couple is found to be quasi-reversible as the peak separation
between the anodic and cathodic potential is very high. But the ratio
between the anodic and cathodic currents suggests that the process
is simple one-electron transfer, quasi-reversible process [36,37]. On
comparing the cyclic voltammograms, we observed that the varia-
tion in oxidation and reduction potential may be due to distortion in
the geometry of the complex which arises due to different anions
coordinated to the metal ion [38].
4.8. Cyclic voltammetry
The electrochemical behavior of Cu(II) complex was examined
by employing glassy carbon as working electrode, Ag/AgCl as
reference electrode and platinum wire as auxiliary electrode (Fig. 6).
The working media consisted of DMF and n-Bu4NþClOꢁ as sup-
4
porting electrolyte. The cyclic voltammogram of Cu(II) complex in
4.9. Fluorescence studies
The emission spectrum of Schiff base was investigated in various
solvents viz., DMF, DMSO, MeCN and THF.
The Schiff base characterized by emission band around 470 nm
in above solvents (DMF, DMSO, THF, and MeCN) is due to phenoxide
anion in the Schiff base. On addition of aqueous alkali (2% NaOH) to
all the above solutions, we observed that, the intensity of 422 nm
band was reduced. The changes in this spectrum of Schiff base in all
the solutions clearly indicate that, proton transferred (H-bonded
ion pair) species exist in equilibrium. In the case of DMF solution as
the addition of aqueous alkali (2% NaOH) the lmax of Schiff base
undergoes red shift in DMF may be due to the hydrogen bond
formation.
5. Conclusion
The newly synthesized compound acts as hexadentate Schiff
base. The metals are coordinated to azomethine nitrogen and
phenolic oxygen atoms. The analytical, spectral, magnetic, and
Fig. 6. CV spectrum of Cu(II) complex.

P.G. Avaji et al. / European Journal of Medicinal Chemistry 44 (2009) 3552–3559
3559
thermal studies confirm the bonding of Schiff base to metal ions.
References
Electrochemical study of Cu(II) complex can provide the degree of
the reversibility of one-electron transfer reaction and they have
a quasi-reversible character. Schiff base was found potentially
active towards microbial strains. All these observations put
together lead us to propose the following structure (Scheme 1) in
which the Ni(II) complex shows octahedral geometry and Co(II) and
Cu(II) complexes exhibit square pyramidal geometry.
[1] J.P. Corhnelissen, J.H. Van Diemen, L.R. Groeneveld, J.G. Haasnoot, A.L. Spek,
J. Reedisk, Inorg. Chem. 31 (1992) 198.
[2] S.N. Dubey, B. Kaushik, Indian J. Chem. 24A (1985) 950.
[3] D.R. Richardson, P.V. Bernhardt, J. Biol. Inorg. Chem. 4 (1999) 266–273.
[4] L. Ramamohan, R.K. Shikkaragol, S.D. Angadi, V.H. Kulkarni, Asian J. Pure Appl.
Chem. 1 (2) (1995) 86.
[5] M.S. Yadawe, S.A. Patil, Transition Met. Chem. 22 (1997) 220.
[6] G. Darnell, D.R. Richardson, Blood 94 (2) (1999) 781.
[7] B. Murukan, K. Mohanan, J. Enzyme Inhib. Med. Chem. 22 (1) (2007)
65–70.
[8] S.M.E. Khalil, H.S. Seleem, B.A. El-Shetary, M. Shebl, J. Coord. Chem. 55 (8)
(2002) 883–899.
6. Experimental protocol
[9] S. Naskar, D. Mishra, S. Kumar Chattopadhyay, M. Corbella, A.J. Blake, Dalton
Trans. (2005) 2428–2435.
[10] D.A. Denton, N. Suschitzky, J. Chem. Soc. (1963) 4741.
[11] R.R. Gange, C.L. Spiro, T.J. Smith, C.A. Hamann, W.R. Thies, A.K. Shiemke, J. Am.
Chem. Soc. 103 (1981) 4073.
[12] F. Ullmann, K. Brittner, Chem. Ber. 42 (1909) 2539.
[13] E.O. Offiong, S. Martelli, Il Farmaco 49 (1994) 513.
[14] J.G. Collee, J.P. Duguid, A.G. Farser, B.D. Marmion (Eds.), Practical Medical
Microbiology, Churchill Livingstone, New York, 1989.
[15] C.J. Balhausen, An Introduction to Ligand Field, McGraw Hill, New York, 1962.
[16] A.B.P. Lever, Inorganic Electronic Spectroscopy, Elsevier, Amsterdam, 1984,
p. 23.
6.1. Chemistry
All chemicals and solvents used were of AR grade. All metal(II)
salts were used as their chlorides. Further remaining pure reagents
were purchased from Ranbaxy Chemicals. The metal contents were
determined gravimetrically by the known method. The results of
elemental analyses, molar conductance and magnetic data are lis-
ted in Table 3.
The IR spectra of the Schiff base and its Co(II), Ni(II) and Cu(II)
complexes were recorded on a HITACHI-270 IR spectrophotometer
in the 4000–350 cmꢁ1 region in KBr discs. The electronic spectra of
the complexes were recorded in DMF on a VARIAN CARY 50-BIO UV
spectrophotometer in the 200–1100 nm region. The 1H NMR spectra
of Schiff base and complexes were recorded in DMSO-d6 on
a BRUKER-300 MHz spectrometer at room temperature using TMS
(Me4Si) as an internal reference. FAB mass spectra were recorded on
a JEOL SX 102/DA-6000 mass spectrometer/data system using argon/
xenon (6 kV,10 A) as the FAB gas. The accelerating voltage was 10 kV
and the spectra were recorded at room temperature; m-nitrobenzyl
alcohol was used as the matrix. The mass spectrometer was operated
in the positive ion mode. Thermogravimetric data were measurꢁed1
[17] B.N. Meyer, N.R. Ferrigni, J.E. Putnam, L.B. Jacobsen, D.E. Nichols,
J.L. McLauhglin, Planta Med. 45 (1982) 31.
[18] Z.H. Chohan, A. Scozzafava, C.T. Supran, J. Enzyme Inhib. Med. Chem. 17 (2003)
261.
[19] Z.H. Chohan, H. Pervez, M.K. Khan, A. Rauf, C.T. Supuran, J. Enzyme Inhib. Med.
Chem. 19 (2004) 85.
[20] Z.H. Chohan, K.M. Khan, C.T. Supuran, Appl. Organomet. Chem. 18 (2004) 305.
[21] Z.H. Chohan, Synth. React. Inorg. Met.-Org. Chem. 34 (2004) 833.
[22] W. Levingson, P. Mikeleus, J. Jackson, W. Kaska, in: G.N. Schranzer (Ed.),
Inorganic and Nutritional Aspects of Cancer, Plenum Press, New York, 1978.
[23] A.W. Bauer, W.M. Kirby, J.C. Sherris, M. Turck, Am. J. Clin. Pathol. 45 (1966)
493.
[24] H.H. Freedman, J. Am. Chem. Soc. 83 (1956) 2900.
[25] A.W. Baker, A.T. Shulgin, J. Am. Chem. Soc. 83 (1959) 1523.
[26] K. Nakamoto, Infrared Spectra of Inorganic and Coordination Compounds,
Wiley-Interscience, New York, 1970.
[27] B.B. Mohapatra, S.K. Saraf, J. Indian Chem. Soc. 80 (2003) 696.
[28] C.K. Choudhary, R.K. Choudray, L.K. Mishra, J. Indian Chem. Soc. 80 (2003) 693.
[29] M.S. Gunthkal, T.R. Goudar, S.A. Patil, Oriental J. Chem. 16 (2000) 151–154.
[30] Sayaji Rao, Asian J. Chem. 17 (2005) 2663–2668.
[31] D.N. Satyanarayana, Electronic Absorption Spectroscopy and Related Tech-
niques, University Press, Hyderabad, 2001.
[32] N. Sengottuvelan, J. Mohanmani, M. Kondaswamy, Polyhedron 21 (2002).
[33] R. Kannappan, R. Mahalakshmy, T.M. Rajchdiran, R. Venkateshan,
P. Sambashiva Rao, Proc. Indian Acad. Sci. (Chem. Sci.) (2003) 115.
[34] M.C. Jain, A.K. Shrivastava, P.C. Jain, Inorg. Chim. Acta 23 (1977) 199.
[35] A. Jaggi, S. Chandra, K.K. Sharma, Polyhedron 4 (1958) 163.
[36] Z. Shirin, R.M. Mukherjee, Polyhedron 11 (1992) 2625.
[37] A. Shyamala, A.R. Chakravarty, Polyhedron 12 (1993) 1545.
[38] S. Chandra, L.K. Gupta, Sangeetika, Synth. React. Inorg. Met.-Org. Chem. 34
(2004) 1591.
from room temperature to 1000 ꢀC at a heating rate of 10 ꢀC min
.
The data were obtained by using a Perkin–Elmer Diamond TG/DTG
instrument. Molar conductivity measurements were recorded on an
Elico-CM-82 T conductivity bridge with a cell having cell constant
0.51; magnetic moments were determined on a Faraday balance.
Appendix. Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.ejmech.2009.03.032.