Synthesis, spectroscopic, antimicrobial, DNA binding and cleavage studies of some metal complexes involving symmetrical bidentate N, N donor Schiff base ligand

Article abstract of DOI:10.1016/j.saa.2011.07.031

The Schiff base ligand, N,N′-bis-(4-isopropylbenzaldimine)-1,2- diaminoethane (L), obtained by the condensation of 4-isopropylbenzaldehyde and 1,2-diaminoethane, has been used to synthesize the complexes of the type [ML₂X₂] [M = Co(I

Full text of DOI:10.1016/j.saa.2011.07.031

Spectrochimica Acta Part A 82 (2011) 191–199
Contents lists available at ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
Synthesis, spectroscopic, antimicrobial, DNA binding and cleavage studies of
some metal complexes involving symmetrical bidentate N, N donor Schiff base
ligand
D. Arish, M. Sivasankaran Nair∗
Department of Chemistry, Manonmaniam Sundaranar University, Tirunelveli 627 012, India
a r t i c l e i n f o
a b s t r a c t
The Schiff base ligand, N,Nꢀ-bis-(4-isopropylbenzaldimine)-1,2-diaminoethane (L), obtained by the
condensation of 4-isopropylbenzaldehyde and 1,2-diaminoethane, has been used to synthesize the com-
plexes of the type [ML2X2] [M = Co(II), Ni(II) and Zn(II); X = Cl and OAc]. The newly synthesized ligand (L)
and its complexes have been characterized on the basis of elemental analyses, mass, 1H and 13C-NMR,
molar conductance, IR, UV–vis, magnetic moment, CV and thermal analyses, powder XRD and SEM. IR
spectral data show that the ligand is coordinated to the metal ions in a bidentate manner. The geometrical
structures of these complexes are found to be octahedral. Interestingly, reaction with Cu(II) ion with this
ligand undergoes hydrolytic cleavage to form ethylenediamine copper(II) complex and the correspond-
ing aldehyde. The antimicrobial results indicate that the chloro complexes exhibit more activity than the
acetato complexes. The complexes bind to CT–DNA by intercalation modes. Novel chloroform soluble
ZnL2Cl2 complex exhibits tremendous antimicrobial, DNA binding and cleaving properties.
© 2011 Elsevier B.V. All rights reserved.
Article history:
Received 12 February 2011
Received in revised form 4 June 2011
Accepted 7 July 2011
Keywords:
Schiff base
IR
CV
Antimicrobial activity
DNA
1. Introduction
2. Experimental
The chelating structures, moderate electron donation and easy
tunable electronic and steric effects proved Schiff bases as versatile
ligands capable of stabilizing different metals in various oxida-
tion states with unusual structural features [1–7]. The chemistry
of Schiff base ligands and their metal complexes have attracted
a lot of interest due to their facile synthesis and wide range of
applications including antifungal, antibacterial, anticancer and cat-
alytic fields [8–11]. The interaction of transition metal compounds
with DNA has been extensively studied due to the unusual bind-
ing, cleaving properties and general photoactivity [12,13]. These
coordination compounds have been found to be suitable candidates
as DNA secondary structure probes, photocleavers and antitumor
drugs [14,15]. Further, for the development of metallo drug as
chemotherapeutic agents, it is essential to explore the interactions
of metal complexes with DNA [16]. In this paper, we report the
synthesis, characterization and biological studies of Co(II), Ni(II)
and Zn(II) complexes with N,Nꢀ-bis-(4-isopropylbenzaldimine)-
1,2-diaminoethane Schiff base ligand.
2.1. Materials
4-isopropylbenzaldehyde was obtained from Himedia. 1,2-
diaminoethane was purchased from Merck and used without
further purification. Calf Thymus (CT) and pUC19 DNA was
obtained from Genei, Bangalore. Co(II), Ni(II), Cu(II) and Zn(II) chlo-
rides/acetates were Merck samples. All other reagents and solvents
were purchased from commercial sources and were of analytical
grade.
2.2. Synthesis of Schiff base ligand
To a solution of 4-isopropylbenzaldehyde (2 mmol) in methanol
(20 ml), 1,2-diaminoethane (1 mmol) in methanol (20 ml) was
added dropwise. The above mixture was magnetically stirred for
about 6 h and the resulting yellow colour solution was kept in a
beaker. After one day, yellow colored crystals were separated out.
It was washed with cold alcohol, ether and recrystallized from
ethanol. Yellow solid; Yield: 92%. Anal. Calcd. for C22H28N2: C 82.45,
H 8.81, N 8.74; found C 82.56, H 8.72, N 8.72; GC–MS (m/z): 321.2
[M + H+]; 1H-NMR (CDCl3, ppm): –CH3 (12H, d, ␦ 1.288), –CH– (2H,
m, ␦ 2.956), –CH2– (4H, s, 3.941), –CH– (aromatic) (4H, d, ␦ 7.249;
4H, d, ␦ 7.605), –CH N– (2H, s, ␦ 8.512); 13C-NMR (CDCl3, ppm):
–CH3 (4C, ␦ 23.746), –CH– (2C, ␦ 34.022), –CH– (aromatic) (12C, ␦
∗
Corresponding author. Tel.: +91 9443540046; fax: +91 462 23334363.
E-mail address: msnairchem@rediffmail.com (M.S. Nair).
1386-1425/$ – see front matter © 2011 Elsevier B.V. All rights reserved.
doi:10.1016/j.saa.2011.07.031

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D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
110-154), –CH N– (2C, ␦ 162.427); IR (ꢀ, cm−1): –C N– (1648);
UV–vis (ꢁmax, nm): 236, 295.
KBr discs on a JASCO FT/IR-410 spectrometer in the 4000–400 cm−1
region. The electronic spectra were recorded on a Perkin Elmer
Lambda-25 UV–vis spectrometer. Room temperature magnetic
measurements were performed on a Guoy’s balance by making dia-
magnetic corrections using Pascal’s constant. Cyclic voltammetric
measurements were carried out in a Bio-Analytical System (BAS)
model CV-50W electrochemical analyzer. The three-electrode cell
comprised of a reference Ag/AgCl, auxiliary platinum and work-
ing glassy electrodes. Tetrabutylammonium perchlorate was used
as supporting electrolyte. Thermal analysis was carried out on a
Perkin-Elmer thermal analyzer with a heating rate of 20 ◦C/min
using N2 atmosphere. Powder XRD was recorded on a Rigaku Dmax
2.3. Synthesis of the Schiff base metal(II) complexes
Schiff base ligand (2 mmol) was dissolved in methanol (20 ml)
and metal(II) chloride/acetates (1 mmol) in methanol (20 ml) was
added dropwise with constant stirring. The above mixture was
magnetically stirred for about 6–8 h. The solid product formed was
filtered off, washed several times with ether and cooled in vacuum
desiccator over fused anhydrous calcium chloride.
[CoL2Cl2]: violet solid; yield: 78%: anal. calcd. for
˚
X-ray diffractometer with Cu-K␣ radiation (ꢁK␣ = 1.5406 A). SEM
images were recorded in a Hitachi SEM analyzer.
C
44H56N4Cl2Co: C 68.56, H 7.32, N 7.27, Cu 7.65; found C 68.33,
H 7.52, N 7.58,−C1o 7.08; m.p: >250 ◦C; DART–MS (m/z):770.6542
[M+]; IR (ꢀ, cm ): –C N– (1638), M–N (452); UV–vis (ꢁmax, nm):
590, 659; ꢂc (ꢃ−1 cm2 mol−1): 12; ꢄeff (B.M): 4.91.
2.5. Antimicrobial activities
[CoL2(OAc)2]: violet solid; yield: 81%: anal. calcd. for
Antibacterial and antifungal activities of the ligand and its com-
plexes were tested in vitro against the bacterial species Escherichia
coli, Bacillus subtilis, Pseudomonas aeruginosa and Staphylococcus
aureus; fungal species, Aspergillus niger, Aspergillus flavus and Can-
dida albicans by the disc diffusion method [18]. All the species used
were of ATCC. The test organisms were grown on nutrient agar for
anti bacterial and potato dextrose agar medium for antifungal in
petri plates. The compounds were prepared in DMF and soaked in
filter paper disc of 5 mm diameter and 1 mm thickness. The discs
were placed on the previously seeded plates and incubated at 37 ◦C
and the diameter of inhibition zone around each disc was mea-
sured after 24 h for antibacterial and 72 h for antifungal activities.
The standard error for the experiment is 0.001 cm and the experi-
ment is repeated three times under similar conditions. DMF is used
as negative control and Amikacin, Ofloxacin and Ciprofloxacin were
used as positive standards for antibacterial and Nystatin for anti-
fungal activities. The minimum inhibitory concentration (MIC) was
determined by serial dilution technique.
C
48H62N4O4Co: C 70.48, H 7.64, N 6.85, Cu 7.20; found C 70.41,
H 7.69, N 6.67, Co 7.27; m.p: >250 ◦C; DART–MS (m/z): 817.9197
[M+]; IR (ꢀ, cm−1): –C N– (1640), COO− (1610, 1402), M–N (449),
M–O (510); UV–vis (ꢁmax, nm): 592, 650; ꢂc (ꢃ−1 cm2 mol−1):
8.0; ꢄeff (B.M): 5.13.
[NiL2Cl2]: green solid; yield: 64%: anal. calcd. for C44H56N4Cl2Ni:
C 68.73, H 7.24, N 7.27, Ni 7.65; found C 68.33, H 7.52, N 7.58, Ni
7.57; m.p: >250 ◦C; DART–MS (m/z):770.5743 [M+]; IR (ꢀ, cm−1):
–C N– (1640), M–N (438); UV–vis (ꢁmax, nm): 378, 710, 1100; ꢂc
(ꢃ−1 cm2 mol−1): 4.0; ꢄeff (B.M): 2.87.
[NiL2(OAc)2]: light green solid; yield: 80%: anal. calcd. for
C48H62N4O4Ni: C 70.50, H 7.64, N 6.85, Ni 7.18; found C 70.71, H
7.61, N 6.64, Ni 7.32; m.p: >250 ◦C; DART–MS (m/z): 817.6523 [M+];
IR (ꢀ, cm−1): –C N– (1640) COO− (1604, 1392), M–N (451), M–O
(530); UV–vis (ꢁmax, nm): 390, 728, 1100; ꢂc (ꢃ−1 cm2 mol−1):
7.0; ꢄeff (B.M): 2.93.
[ZnL2Cl2]: white solid; yield: 92%. anal. calcd. for
C44H56N4Cl2Zn: C 67.99, H 7.26, N 7.21, Zn 8.41; found C 67.93,
H 7.64, N 7.28, Ni 7.98; m.p: >250 ◦C; DART–MS (m/z): 777.3645
[M+]; 1H-NMR (CDCl3, ppm): –CH3 (12H, d, ␦ 1.279), –CH– (2H,
m, ␦ 2.974), –CH2– (4H, s, 4.029), –CH– (aromatic) (4H, d, ␦ 7.418;
4H, d, ␦ 7.944), –CH N– (2H, s, ␦ 8.675); 13C-NMR (CDCl3, ppm):
–CH3 (4C, ␦ 23.590), –CH– (2C, ␦ 34.436), –CH2– (2C, ␦ 59.815),
–CH– (aromatic) (12C, ␦ 126–156), –CH N– (2C, ␦ 169.468); IR (ꢀ,
cm−1): –C N– (1642), M–N (461); UV–vis (ꢁmax, nm):237, 290;
ꢂc (ꢃ−1 cm2 mol−1): 12.0.
[ZnL2(OAc)2]: white solid; yield: 92%: anal. calcd. for
C48H62N4O4Zn: C 69.93, H 7.58, N 6.80, Ni 7.93; found C 70.11,
H 7.70, N 6.64, Ni 8.12; m.p: >250 ◦C; DART–MS (m/z): 824.6312
[M+]; 1H-NMR (CDCl3, ppm): –CH3 (12H, d, ␦ 1.278), –CH– (2H,
m, ␦ 2.971), –CH2– (4H, s, 4.039), –CH– (aromatic) (4H, d, ␦ 7.412;
4H, d, ␦ 7.938), –CH N– (2H, s, ␦ 8.677); 13C-NMR (CDCl3, ppm):
–CH3 (4C, ␦ 23.590), –CH– (2C, ␦ 34.436), –CH2– (2C, ␦ 59.815),
–CH– (aromatic) (12C, ␦ 126–156), –CH N– (2C, ␦169.732); IR (ꢀ,
cm−1): –C N– (1640) COO− (1612, 1418), M–N (437), M–O (517);
UV–vis (ꢁmax, nm):236, 289; ꢂc (ꢃ−1 cm2 mol−1): 9.0.
2.6. DNA binding experiments
A solution of CT–DNA in 0.5 mM NaCl/5 mM Tris-HCl (pH 7.0)
gave a ratio of UV absorbance at 260 and 280 nm (A260/A280)
of 1.8–1.9, indicating that the DNA was sufficiently free of pro-
teins. A concentrated stock solution of DNA was prepared in 5 mM
Tris-HCl/50 mM NaCl in water at pH 7.0 and the concentration of
CT–DNA was determined per nucleotide by taking the absorption
coefficient (6600 dm3 mol−1 cm−1) at 260 nm [19]. Stock solutions
were stored at 4 ◦C and were used only for a maximum of 4 days.
Doubly distilled water was used to prepare buffer solutions. Solu-
tions were prepared by mixing the complex and CT–DNA in DMF
medium. Absorption titrations were performed by keeping the con-
centration of the complex constant (10 ␮M) and by varying the
concentration of CT–DNA from 0 to 25 ␮M. The binding constant
(Kb) for the complexes have been determined from the following
equation [19]
[DNA]
(εA − εF)
[DNA]
(εB − εF)
1
=
+
Kb(εB − εF)
2.4. Physical measurements
where εA, εB and εF correspond respectively to the apparent,
bound and free metal complex extinction coefficients. A plot of
[DNA]/(εA − εF) versus [DNA] gave a slope of 1/(εB − εF) and a Y-
intercept equal to 1/Kb(εB − εF), where Kb is the ratio of slope to the
intercept.
Elemental analyses were done using a Perkin-Elmer elemen-
tal analyzer. The GC mass spectrum of the Schiff base ligand was
recorded on a GC–MS/MS Varian Saturn 2200 GC-Varian CP 3800
spectrometer. The DART–MS was recorded on a JEOL-AccuTOF JMS-
T100LC mass spectrometer having a DART (Direct Analysis in Real
Time) source. The metal contents in the complexes were deter-
mined by standard EDTA titration [17]. Molar conductance of the
complexes were measured in MeOH (10−3 M) solutions using a
coronation digital conductivity meter. IR spectra were recorded in
2.7. Gel electrophoresis
The pUC19 DNA (1 ␮g), 10 ␮M metal complex, 50 ␮M H2O2
in 50 mM Tris-HCl buffer (pH 7.1) were mixed. The contents

D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
193
Fig. 1. (a) GC-Mass spectrum of the Schiff base ligand (L). (b) DART-Mass spectrum of [ZnL2Cl2] complex.
were incubated for 30 min at 37 ◦C and loaded on 0.8% Agarose
3.1. Mass spectra
gel after mixing 5 ␮l of loading buffer (0.25% bromophenol
blue + 0.25% Xylene cyanol + 30% glycerol sterilized distilled water).
Electrophoresis was performed at constant voltage (100 V) until
the bromophenol blue reached to one third of the gel. The gel was
stained for 10 min by immersing in an ethidium bromide solution.
The gel was then detained for 10 min by keeping in sterilized dis-
tilled water and the plasmid bands visualized by photographing
the gel under a UV transilluminator. The efficiency of DNA cleavage
was measured by determining the ability of the complex to form
open circular (OC) or nicked circular (NC) DNA from its super coiled
(SC) form.
The GC–MS of Schiff base (Fig. 1a) shows a well-defined
molecular ion peak at 321.2 (M + H+), which coincides with for-
mula weight of the Schiff base. The DART mass spectrum of
CoL2Cl2, NiL2Cl2, ZnL2Cl2, CoL2(OAc)2, NiL2(OAc)2 and ZnL2(OAc)2
complexes shows a peak at m/z 770.6542(4%), 770.5743(2.1%),
777.3645(2.8%), 817.9197(4%), 817.6523(4%) and 824.6312(2.2%),
respectively, corresponding to their molecular weights. The mass
spectrum of all the complexes indicates that the complexes are
monomeric confirming the metal to ligand ratio to be 1:2 in the
complexes. The representative mass spectrum of ZnL2Cl2 com-
plexes is shown in Fig. 1b.
3. Results and discussion
3.2. NMR spectra
The analytical data and physical properties of the ligand and
its complexes are given in experimental Sections 2.2 and 2.3. The
elemental analyses data confirm the assigned composition of the
ligand and its complexes. The Schiff base is soluble in all common
organic solvents except water. All the complexes are stable at room
temperature, insoluble in water, but soluble in MeOH, DMF, DMSO
and MeCN. The Zn(II) complexes are highly soluble in chloroform.
The low molar conductivity values of the metal complexes are given
in experimental Section 2.3 suggests their non-electrolytic nature
[20].
The NMR spectral data of the ligand and its Zn(II) com-
plexes are given in experimental Sections 2.2 and 2.3. The 1H
and 13C-NMR spectrum of the ligand and its ZnL2Cl2 is shown
in Figs. 2a and b and 3a and b. As shown in Fig. 2a, it is observed that
the signal for azomethine proton (>CH N–) in the ligand appears
as a singlet at 8.512 ppm. The peaks at 1.288 and 2.596 ppm are
assignable to the isopropyl –CH– and –CH3 protons, respectively.
The aromatic ring protons are observed in the 7–8 ppm range as
expected. The 13C-NMR spectrum of Schiff base ligand shows a peak
at 162.427 ppm characteristic of azomethine carbon. In the 1H and

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D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
Fig. 2. (a) 1H-NMR spectrum of the Schiff base ligand (L). (b) 13C-NMR spectrum of the Schiff base ligand (L).
13C-NMR spectrum of Zn(II) complexes, the azomethine (–CH N–)
proton and carbon peaks are shifted to downfield which indicates
the metal to coordinate the ligand through azomethine nitrogen
atom.
3.3. IR spectra
The IR spectrum of the free ligand exhibits a sharp band
at 1648 cm−1, due to the azomethine group vibration. On com-

D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
195
Fig. 3. (a) 1H-NMR spectrum of [ZnL2Cl2] complex. (b) 13C-NMR spectrum of [ZnL2Cl2] complex.
plexation this band was shifted to lower frequency in the
3.4. Electronic spectra and magnetic measurements
∼1642–1638 cm−1 range indicating the coordination of the azome-
thine nitrogen atom to the metal ion. In addition several strong
sharp bands at 2990–2940 cm−1 range can be ascribed to ꢀ(C–H)
vibration of the isopropyl group. In the acetato complexes, the
bands at 1612–1605 and 1418–1392 cm−1 can be attributed
to the asymmetric and symmetric stretching vibration of the
acetate group. The large difference between the ꢀas(CH3COO−)
and ꢀs(CH3COO−) value of ∼200 cm−1 indicates the monoden-
tate binding nature of the acetato group [21,22]. The weak bands
observed at 461–430 cm−1 can be assigned to the ꢅ(M–N) vibration
[22].
In the electronic spectrum of the ligand, the broad band cen-
tered at 295 nm is assigned to ␲–␲* transition of the azomethine
(>C N) chromophore. In addition, the other intense absorption
broad band centered at 236 nm is due to the ␲–␲* and n–␲*
transition of the benzene ring of the Schiff base ligand. The elec-
tronic spectrum of Co(II) complexes show two absorption bands at
550–700 nm region (experimental Section 2), which are assignable
4
4
to 4T1g(F) → A2g(F) and 4T1g(F) → T2g(F) transitions in an octahe-
dral environment [23]. The magnetic susceptibility value is found
to be 4.91 and 5.13 B.M for CoL2Cl2 and CoL2(OAc)2, respectively.

196
D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
Fig. 4. TG–DTA curves of (a) [CoL2Cl2]; (b) [CoL2(OAc)2] complexes.
This is comparable with that of 4.3–5.2 B.M expected for octa-
hedral Co(II) complexes, indicative of octahedral geometry for
the present Co(II) complexes [24]. The electronic spectrum of the
Ni(II) complexes show three bands in the region ∼1100, ∼700 and
250–260 ◦C corresponds to the loss of chloride ions. Above this tem-
perature, a successive decomposition step was observed which can
be attributed to the partial decomposition of the Schiff base ligand.
This step is accompanied by exothermic peaks. Whereas, in acetato
complexes, the first thermal degradation that occurs between 250
and 400 ◦C corresponds to the loss of acetato groups and partial
decomposition of the Schiff base ligand. This step is accompanied
by exothermic and endothermic peaks. Based on the above results,
the proposed structure of the complexes is given in Fig. 5.
3
3
3
3
∼400 nm, attributable to A2g(F) → T2g(F), A2g(F) → T1g(F) and
3A2g(F) → T1g(P) transitions, respectively, suggesting an octahe-
3
dral geometry around Ni(II) atom [24]. The NiL2Cl2 and NiL2(OAc)2
complexes reported herein are found to have a room temperature
magnetic moment value of 2.87 and 2.93 B.M respectively, which
are in the range of 2.9–3.3 B.M observed for octahedral Ni(II) com-
plexes [24].
3.7. Powder XRD and SEM
Powder XRD pattern of the Schiff base complexes were recorded
over the 2Â = 0–80◦. The CoL2Cl2 and CoL2(OAc)2 complexes do
not exhibit well defined crystalline peak due to their amorphous
3.5. Electrochemistry
The cyclic voltammograms were recorded in acetonitrile solu-
tion at a scan rate of 100 mV s−1 in the potential range +2.0
to −2.0 V. The cyclic voltammogram of CoL2Cl2 shows a well
defined redox process corresponding to the formation of the
quasi-reversible Co(II)/Co(I) couple. The cathodic peak at −689 mV
versus Ag/AgCl and the associated anodic peak at −240 mV cor-
respond to the Co(II)/Co(I) couple. The peak-to-peak separation
(
Ep = 449 mV) indicates quasi-reversible one electron transfer
process. The redox property of CoL2(OAc)2 displayed electro-
chemically reversible Co(II)/Co(I) reduction peak at −703 mV and
the associated oxidation peak at −258 mV corresponds to the
Co(II)/Co(I) quasi-reversible process. The peak-to-peak separation
(
Ep) is 445 mV indicating the process to be quasi-reversible. The
electrochemistry of NiL2Cl2 is similar to that of NiL2(OAc)2, anodic
waves are seen at −810 and at −838 mV and the associated cathodic
peaks at −692 and −678 mV respectively, corresponding to the for-
mation of the quasi-reversible one-electron reduction Ni(II)/Ni(I)
couple. The cyclic voltammogram of Zn(II) complexes did not show
any characteristic peak potential in this potential range under sim-
ilar conditions.
3.6. Thermal study
The thermal stabilities of the complexes were investigated using
TG and DTA at a heating rate of 10 ◦C min−1 in N2 from 35 to 500 ◦C.
The representative TG–DTA curves of CoL2Cl2 and CoL2(OAc)2 com-
plexes are shown in Fig. 4a and b. All the complexes display almost
similar decomposition steps. The thermograms showed no weight
loss up to 250 ◦C, indicating stability of the complexes and the
absence of water molecule in the complex. In the chloro complexes
(Fig. 4a), it is observed that the first thermal degradation that occurs
between 250 and 300 ◦C with an endothermic peak in the range
Fig. 5. Proposed structure of the complexes.

D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
197
Fig. 6. SEM images of (a) Schiff base ligand (L); (b) [CoL2Cl2]; (c) [CoL2(OAc)2]; (d) [NiL2Cl2]; (e) [NiL2(OAc)2]; (f) [ZnL2Cl2] and (g) [ZnL2(OAc)2] complexes.
nature whereas other complexes display sharp crystalline peaks
indicating their crystalline nature. The average crystallite sizes of
the complexes dXRD were calculated using Scherrer formula [25].
The [NiL2Cl2], [NiL2(OAc)2], [ZnL2Cl2] and [ZnL2(OAc)2] complexes
have an average crystallite size of 34, 48, 64 and 53 nm, respec-
tively.
The SEM micrographs of the Schiff base ligand (L) and its metal
complexes are shown in Fig. 6a–g. The SEM image of Schiff base
Scheme 1. Complex formation of Cu(II) ion with Schiff base ligand [L].

198
D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
Table 1
Minimum inhibitory concentration (MIC) results of the synthesized compounds ((g/mL).
Compounds
Bacterial species
Fungal species
E. coli
B. subtilis
P. aereuguinosa
S. aureus
A. niger
A. flavus
C. albicans
L
>100
25
>100
15
>100
10
45
15
15
08
–
95
30
95
25
95
05
25
10
10
10
–
>100
28
95
25
40
20
15
10
10
05
–
95
35
90
85
90
20
75
05
10
10
–
80
15
95
80
80
04
75
–
>100
35
>100
>100
>100
04
10
–
–
–
85
15
>100
20
>100
05
10
–
–
–
08
[CoL2Cl2]
[CoL2(OAc)2]
[NiL2Cl2]
[NiL2(OAc)2]
[ZnL2Cl2]
[ZnL2(OAc)2]
Amikascina
Ciprofloxacina
Ofloxacina
Nystatina
–
–
10
08
a
Standard.
ligand shows (Fig. 6a) irregularly shaped micro flake like surface
morphology. The CoL2Cl2 complex (Fig. 6b) display agglomer-
ated particles and presence of random distribution of voids in
their structure. Whereas [CoL2(OAc)2] complex exhibits (Fig. 6c)
irregular shaped secondary crystalline object with primary tiny
particles. From the SEM image (Fig. 6d and e), it is observed that
the crystalline objects are rod-shaped with lateral dimensions in
nanometer range but with a considerable length of 50 to few hun-
dred micrometer in the Ni(II) complexes. The SEM image of Zn(II)
complexes (Fig. 6f and g) show a gorgeous surface morphology with
nano needles and rods, and the presence of particles are randomly
distributed.
cleavage reaction [26]. In general, coordination to metal can cause
weakening or strengthening of the –C N– group [27]. Thus, the
Cu(II) ion coordinates to the imine nitrogen atom causing the weak-
ening of the –C N– bond. Consequently, the –C N– group is highly
polarized and nucleophilic attack on the carbon atom of the –C N–
group can occur. This is followed by the presence of water molecule
bringing cleavage [27].
4. Biological studies
4.1. Antimicrobial activity
The minimum inhibitory concentration (MIC) values of the
compounds are given in Table 1. From the in vitro antimicrobial
screening results (Table 1), it is observed that the ligand is mod-
erately active against bacteria, E. coli, S. aureus and fungi, A. niger.
The complexes have shown higher activity in comparison with the
ligand against both bacteria and fungi used. The activity of the lig-
and has enhanced on complexation, which can be explained on the
basis of Overtone’s concept and Tweedy’s chelation theory [28].
Among the complexes, ZnL2Cl2 complex has the highest poten-
tial against all the microorganisms, which is even more than the
standard drugs used. [CoL2(OAc)2] and [NiL2(OAc)2] complexes
showed lower activity against E. coli and C. albicans as compared
with the free ligand. As seen in Table 1, it can be concluded that
the chloro complexes have higher antimicrobial activity than the
acetato complexes. This may be due to the presence of labile chlo-
ride ions in the complexes [29].
3.8. Effect of coordination on Cu(II)
Cu(II) reacts with the Schiff base ligand (L) under the nor-
mal reaction condition, and undergoes hydrolytic cleavage to form
Cu(II)-ethylenediamine complex and the corresponding aldehyde
(Scheme 1). The hydrolytic products were isolated and charac-
terized using mass and IR spectroscopic techniques. The cleavage
mechanism was not correctly known. However, it is likely that
under the conditions of the reaction, an activated nucleophile has
been generated at the metal and is responsible for the resultant
4.2. DNA binding studies
In the UV region, all the complexes exhibit an intense band at
290 nm which is attributed to a ␲–␲* transition. Upon the addition
of CT–DNA to the complexes, a decrease in the molar absorptivity
(hypochromism, 23–52%) of the ␲–␲* absorption band with red
shifts is observed, which indicates strong binding of the complexes
to DNA. The extent of hypochromism is commonly consistent
with the strength of DNA interaction. The observed order of
decrease in hypochromism, ZnL2Cl2 (Kb = 5.2 × 105 M−1) > NiL2Cl2
(Kb = 1.33 × 104 M−1) > CoL2Cl2 (Kb = 6.8 × 103 M−1) > ZnL2(OAc)2
(Kb = 5.3 × 103 M−1) > NiL2(OAc)2 (Kb = 5.1 × 103 M−1) > CoL2(OAc)2
(Kb = 1.25 × 102 M−1), reflects the decreasing DNA binding affinities
of the complexes in this order. This is consistent with the higher
hypochromism and the higher red shift observed for ZnL2Cl2.
Interestingly, chloro complexes exhibit higher binding affinity
than acetato complexes which is also reflected in antimicrobial
activity. Complexes bound to DNA through intercalation usu-
ally result in hypochromism and red shift due to the intercalative
mode involving a strong stacking interaction between the aromatic
Fig. 7. Absorption spectra of [ZnL2Cl2] complex in the presence of increas-
ing amounts of CT DNA. Curve 1: 0 ␮M DNA + 10 ␮M [ZnL2Cl2]; curve 2: 5 ␮M
DNA + 10 ␮M [ZnL2Cl2]; curve 3: 10 ␮M DNA + 10 ␮M [ZnL2Cl2]; curve 4: 15 ␮M
DNA + 10 ␮M [ZnL2Cl2]; curve 5: 20 ␮M DNA + 10 ␮M [ZnL2Cl2]; curve 6: 25 ␮M
DNA + 10 ␮M [ZnL2Cl2].

D. Arish, M.S. Nair / Spectrochimica Acta Part A 82 (2011) 191–199
199
Fig. 8. Gel electrophoresis diagram for the reaction done in the control experiment with an incubation time of 60 min at 30 ◦C in dark condition: Lane 1: DNA + H2O2;
lane 2: DNA + Ligand + H2O2; lane 3: DNA + [CoL2Cl2] + H2O2; lane 4: DNA + [CoL2(OAc)2] + H2O2; lane 5: DNA + [NiL2Cl2] + H2O2; lane 6: DNA + [NiL2(OAc)2] + H2O2; lane 7:
DNA + [ZnL2Cl2] + H2O2; lane 8: DNA + [ZnL2(OAc)2] + H2O2.
chromophore and the base-pairs of DNA [30]. Thus, the above
phenomena imply that the present complexes interact with DNA
by intercalating mode. The absorption spectrum of the ZnL2Cl2
complex in the absence and presence of CT–DNA is shown in Fig. 7
shaped structure. Coordination with Cu(II) ion with this ligand
undergoes hydrolytic cleavage to form ethylenediamine Co(II)
complex and the corresponding aldehyde. The complexes can
effectively cleave plasmid DNA in the presence of H2O2 as an
oxidant. Among the complexes, chloroform soluble ZnL2Cl2 com-
plex exhibits a good antimicrobial, DNA binding and cleaving
properties.
4.3. DNA cleavage studies
Gel electrophoresis experiments using pUC19 DNA were per-
formed with ligand and its metal complexes in the presence of
H2O2 as an oxidant. At micro molar concentrations for 2 h incu-
bation periods, the ligand (Lane 2) exhibits no significant cleavage
activity in the presence of H2O2. The nuclease activity is greatly
enhanced by the incorporation of metal ion in the respective Schiff
base ligand. From Fig. 8, it is evident that the complexes cleave DNA
more efficiently in the presence of oxidant, which may be due to
the formation of hydroxyl free radicals. These hydroxyl free radicals
participate in the oxidation of the deoxyribose moiety, followed by
the hydrolytic cleavage of the sugar phosphate backbone. The pro-
duction of hydroxyl radical due to the reaction between the metal
complex and oxidant can be explained as shown below [31].
Acknowledgment
One of the authors (D.A) is grateful to National Testing Service-
India, CIIL, Mysore for providing the fellowship.
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In
Summary,
a
new
Schiff
base,
N,Nꢀ-bis-(4-
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and the geometrical structures of these complexes are found to
be octahedral. The thermograms showed no weight loss up to
250 ◦C, indicating the absence of water molecule in the complex.
The SEM image of Zn(II) complexes show nano needles and rod
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