Novel tetranuclear distorted open-cubane copper complex containing oximate bridges: Synthesis, crystal structure, DNA binding and cleavage activity

Article abstract of DOI:10.1016/j.ica.2012.03.042

The synthesis, molecular structure, DNA binding and nuclease activity of Cu₄O₄ open-cubane tetranuclear copper(II) complex with 3-[2-[(ethyl amino)ethyl]imino]-2-butanoneoxime (HL) are reported for the first time. The neutral tetranuclear [Cu₄L₄(ClO ₄)₄] complex crystallizes in tetragonal space group P4?2₁c with the unit cell parameters; a = 13.798(4) ?, b = 13.798(4) ?, c = 14.119(6) ?, V = 2688(16) ?³, Z = 8, R = 0.0636. Symmetrically equivalent copper atoms exhibit a CuN ₃O₃ elongated distorted octahedral coordination environment, with three nitrogen atoms of the L^- ligand and one oxime-oxygen atom of second L^- ligand at equatorial positions, one oxime-oxygen atom of the third L^- ligand and perchlorate oxygen at axial positions. The complex shows quasireversible cyclic voltammetric response at 0.805 V (ΔE_p = 277 mV) at 100 mV s^-1 in DMF for the Cu(II)/Cu(I) redox couple. The binding study of the complex with calf-thymus DNA has been investigated using absorption spectrophotometry. The complex shows strong nuclease activity on stranded pBR 322 plasmid DNA in the presence of hydrogen peroxide and marginal nuclease activity in the presence of reducing agent (dithiothreitol).

Full text of DOI:10.1016/j.ica.2012.03.042

Inorganica Chimica Acta 392 (2012) 478–484
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Inorganica Chimica Acta
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Note
Novel tetranuclear distorted open-cubane copper complex containing oximate
bridges: Synthesis, crystal structure, DNA binding and cleavage activity
Haribabu Pagonda a, P. Patil Yogesh b, Hussain Reddy Katreddi a, , Nethaji Munirathinam b
⇑
a Department of Chemistry, Sri Krishnadevaraya University, Anantapur 515003, India
b Department of Inorganic and Physical Chemistry, Indian Institute of Science, Bangalore, India
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis, molecular structure, DNA binding and nuclease activity of Cu4O4 open-cubane tetranuclear
copper(II) complex with 3-[2-[(ethyl amino)ethyl]imino]-2-butanoneoxime (HL) are reported for the first
time. The neutral tetranuclear [Cu4L4(ClO4)4] complex crystallizes in tetragonal space group P421c with
Received 24 December 2011
Received in revised form 17 March 2012
Accepted 21 March 2012
Available online 5 April 2012
ꢀ
the unit cell parameters; a = 13.798(4) Å, b = 13.798(4) Å, c = 14.119(6) Å, V = 2688(16) Å3, Z = 8,
R = 0.0636. Symmetrically equivalent copper atoms exhibit a CuN3O3 elongated distorted octahedral
coordination environment, with three nitrogen atoms of the Lꢀ ligand and one oxime-oxygen atom of
second Lꢀ ligand at equatorial positions, one oxime-oxygen atom of the third Lꢀ ligand and perchlorate
oxygen at axial positions. The complex shows quasireversible cyclic voltammetric response at 0.805 V
Keywords:
X-ray crystal structure
Oxime-schiff base ligand
Tetranuclear copper(II) complex
Distorted open-cubane
Nuclease activity
(D
Ep = 277 mV) at 100 mV sꢀ1 in DMF for the Cu(II)/Cu(I) redox couple. The binding study of the complex
with calf-thymus DNA has been investigated using absorption spectrophotometry. The complex shows
strong nuclease activity on stranded pBR 322 plasmid DNA in the presence of hydrogen peroxide and
marginal nuclease activity in the presence of reducing agent (dithiothreitol).
Ó 2012 Elsevier B.V. All rights reserved.
1. Introduction
copper (II) complexes owes their origin to their potentiality for effi-
cient intra-molecular activation of bound dioxygen and to selective
Tetranuclear copper(II) complexes are of current interest in
bioinorganic modeling, multielectron transfer, catalytic and new
extended material investigations [1–5]. The design and synthesis
and characterization of tetranuclear copper(II) complexes have
been carried out in relation to metalloproteins. The cleavage of
nucleic acids may be considered as an enzyme reaction which
comprises of various biological processes as well as the biotechno-
logical manipulation of genetic material. The application of artifi-
cial DNA cleaving agents is manifold: biotechnology, structural
studies of nucleic acids or development of new drugs [6–15].
Compounds showing the property of effective binding as well as
cleaving double stranded DNA under physiological conditions are
of paramount importance since these could be used as diagnostic
agents in medicinal and genomic research [16–31]. Knowing the
very sensitive nature of DNA towards oxidative cleavage, the
majority of the studies on artificial DNA have been centered
around molecules capable of cleaving DNA with an oxidative
mechanism [15]. Several efficient cleaving agents have been devel-
oped in course of time. Transition metal complexes have been
extensively studied because of their nuclease – like activity and
biological role played by them. Large attention on multinuclear
binding to a particular nucleic acid conformation. Studies with
polynuclear copper complexes have been amply reported by Karlin
and co-workers [32–35].
In continuation of our ongoing research on DNA binding and
cleavage activities of transition metal complexes [36–39], herein
we report the synthesis, crystal structure, DNA binding and cleav-
age activity of a new tetra-nuclear copper(II) complex having the
formula [Cu4L4(ClO4)4] (HL = 3-[2[(ethylamino)-ethyl]imino]-2-
butanoneoxime). The complex has distorted Cu4O4 open cubane
core. The complex binds strongly with CT DNA and efficiently
cleaves supercoiled plasmid DNA in the presence of hydrogen per-
oxide. The cleavage of DNA takes place via oxidative path.
2. Experimental
2.1. Materials and measurements
Diacetylmonoxime (AR grade) was purchased from Merck
Chemicals Pvt. Ltd., India and N-ethyl ethylenediamine (AR grade)
from Sigma–Aldrich. All other chemicals were of AR grade and used
as supplied. The solvents were distilled before use. Calf-thymus
DNA and plasmid pBR-322 were purchased from Genie Bio labs,
Bangalore, India. Single crystal X-ray diffraction data were collected
at 298 K on a Bruker AXS KAPPA APEX CCD diffractometer using
⇑
Corresponding author. Tel.: +91 8554 255741.
E-mail address: khussainreddy@yahoo.co.in (H.R. Katreddi).
0020-1693/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.ica.2012.03.042

H. Pagonda et al. / Inorganica Chimica Acta 392 (2012) 478–484
479
graphite monochromated Mo K
a
radiation (k = 0.71073 Å). Mag-
2.4. Synthesis
netic moment measurement of the complex was taken at 298 K
using Faraday’s magnetic susceptibility balance (Sherwood Scien-
tific, Cambridge, UK). High purity pentahydrated copper sulfate
2.4.1. Synthesis of 3-[2-[(ethyl amino)ethyl]imino]-2-butanoneoxime
(HL)
was used as
a
standard. The conductivity measurements at
The ligand was prepared by refluxing N-ethylethylenediamine
(0.53 ml, 5 mmol) and diacetylmonoxime (0.505 g, 5 mmol) in
30 ml of methanol for an hour. The ligand was not isolated, instead
the resulting reddish-yellow methanol solution was subsequently
used in the preparation of complex.
298 2 K in dry and purified DMF were carried out on CM model
162 Conductivity meter (ELICO). The electronic spectra of metal
complex in DMF were recorded with UV lamda50 (Perkin Elmer)
spectrophotometer. The infrared spectra were recorded in the range
4000–400 cmꢀ1 with Perkin Elmer Spectrum 100 spectrophotome-
ter in KBr discs. ESR spectra were recorded on Varian E-112 X-band
spectrophotometer at room temperature and at liquid nitrogen
temperature (LNT) in both solution (DMF) and solid state. The cyclic
voltammetric measurements were taken on a BAS CV-27 assembly
in conjunction with an X–Y recorder. Measurements were made
on the degassed (N2 bubbling for 5 min) solutions in (10ꢀ3 M) con-
taining 0.1 M tetrabutylammonium hexaflourophosphate as the
supporting electrolyte. The three- electrode system consists of
glassy carbon (working), platinum wire (auxiliary) and Ag/AgCl (ref-
erence) electrode. DNA cleavage activities were determined using
UVI-tech-UK gel documentation system.
2.4.2. Synthesis of complex [Cu4L4(ClO4)4]
A 10 ml of methanolic solution of Cu(ClO4)2ꢂ6H2O (1.853 g,
5 mmol) was added to the ligand (HL) solution (5 mmol in 10 ml
methanol), and the resulting reaction mixture was stirred well. Tri-
ethylamine (0.85 ml, 5 mmol) was added drop wise to the reaction
mixture with constant stirring. The resultant solution was filtered
and evaporated at room temperature. A green colored complex was
collected and recrystallised from acetonitrile solvent. Deep-green
crystals of complex suitable for X-ray diffraction were grown by
slow evaporation of a 1:1 acetonitrile–hexane solvent mixture.
Yield: 1.42 g (69%).
2.2. DNA binding experiments
2.5. X-ray crystallography
Interaction of the complex with calf thymus DNA has been
studied by recording electronic absorption spectra. A solution of
CT-DNA in 5 mM Tris–HCl/50 mM NaCl (pH 7.0) gave a ratio of
UV absorbance at 260 and 280 nm (A260/A270) of 1.8–1.9, indicating
that the DNA is sufficiently free of proteins [40]. 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 deter-
mined per nucleotide by taking the absorption coefficient
(6600 dm3 molꢀ1 cmꢀ1) at 260 nm [41]. Stock solutions are stored
at 4 °C and were used after no more than 4 days. Doubly distilled
water was used to prepare buffer solutions. Solutions were pre-
pared by mixing the complex and CT-DNA in DMF medium. After
equilibrium reached (ca. 5 min) the spectra were recorded against
an analogous blank solution containing the same concentration of
DNA.
Single crystal X-ray diffraction measurements were carried out
on a Bruker AXS KAPPA APEX CCD diffractometer. The unit cell
parameters were determined and the data collections were
performed using graphite-monochromated Mo
Ka radiation
(k = 0.71073) radiation. The crystal was found to be tetragonal with
Table 1
Crystal data, data collection and structure refinement parameters of the
[Cu4L4(ClO4)4].
Formula
Formula weight(M)
T (K)
C8H16ClCuN3O5
333.22
293(2)
Wavelength (Mo K
Crystal system
Space group
Lattice constants
a (Å)
a) (Å)
0.71073
tetragonal
ꢀ
P421c
UV-spectral data were fitted into Eq. (1) to obtain the intrinsic
binding constant (Kb)
13.798(4)
b (Å)
13.798(4)
½DNAꢁ=ðea
ꢀ
ef Þ ¼ ½DNAꢁ=ðeb
ꢀ
ef Þ þ 1=Kbðeb
ꢀ
ef
Þ
ð1Þ
c (Å)
14.119(6)
a
(°)
90
where [DNA] is the concentration of DNA in base pairs, ea
,
eb and ef
b (°)
90
are apparent extinction coefficient (Aobs/[M]), the extinction coeffi-
cient for the metal (M) complex in the fully bound form and the
extinction coefficient for free metal (M), respectively. A plot of
c
(°)
90
2688(16)
8
1.647
1.840
1368
V (A3)
Z
Calculated density,
Absorption coefficient
F(000)
Crystal size (mm)
h Range for data collection
Limiting indices
q
(Mg mꢀ3
(mmꢀ1
)
[DNA]/(ea
cept equal to 1/Kb(eb
ꢀ
e
f) versus [DNA] gave a slope of 1/(eb
ꢀ
ef) and Y-inter-
l
)
ꢀ
e
f); Kb is the ratio of the intercept.
0.29 ꢃ 0.27 ꢃ 0.12
2.06–26
2.3. Assay of nuclease activity
ꢀ16 6 h 6 11, ꢀ11 6 k 6 17,
ꢀ17 6 l 6 14
12032
2592[Rint = 0.0405]
99.9
The extent cleavage of pBR 322 DNA by the tetranuclear cop-
per(II) complex was monitored by agarose gel electrophoresis
using. The samples after incubation for 30 min at 37 °C were added
to the loading buffer containing 0.25% bromophenol blue + 0.25%
xylene cynaol + 30% glycerol and solutions were loaded on 0.8%
Reflections collected
Unique reflections
Completeness to h (%)
Maximum and minimum
transmission
Refinement method
Data/restraints/parameters
Goodness-of-fit on F2
Final R indices [I > 2
R indices (all data)
0.8094 and 0.6175
full-matrix least-squares on F2
2592/0/166
agarose gel containing 100 lg of ethidium bromide. Electrophore-
sis was performed at 75 V in TBE buffer until the bromophenol blue
reached to the 3/4th length of the gel. Bands were visualized by
UV-transilluminator and photographed. 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 reactions were carried out under oxida-
tive conditions. Control experiments were done in the presence of
1.102
r
(I)]
R1a = 0.0415, wR2b,c = 0.1042
R1a = 0.0652, wR2b,c = 0.1291
0.529 and ꢀ0.641
Largest difference in peak and hole
(e Åꢀ3
)
P
P
a
b
c
R1
=
(|Fo| ꢀ |Fc|/ |Fo|.
P
P
wR2 = { [w(Fo ꢀ Fc2)2]/ [w(Fo2)2]}1/2
.
2
w = 1/[
r
2(Fo2)+(aP)2 + bP] with P = [Fo2 + 2Fc2]/3, a = 0.0701 and b = 0.2603.
hydroxyl radical scavenger DMSO (4 ll), chelating agent (EDTA).

480
H. Pagonda et al. / Inorganica Chimica Acta 392 (2012) 478–484
Table 2
Selected bond lengths (Å) and bond angles (°) for complex.
Atoms
[Cu4L4(ClO4)4]Complex
Cu(1)–O(1)
1.937(4)
2.575(3)
2.626(4)
2.014(4)
1.944(6)
2.056(4)
173.4(2)
96.7(2)
80.0(2)
99.5(2)
83.8(2)
163.8(2)
114.6(4)
117.2(4)
115.9(4)
117.9(4)
104.0(4)
Cu(1)–O(1ii)
Cu(1)–O(2i)
Cu(1)–N(1)
Cu(1)–N(2)
Cu(1)–N(3)
O(1)–Cu(1)–N(2)
O(1)–Cu(1)–N(1)
N(2)–Cu(1)–N(1)
O(1)–Cu(1)–N(3)
N(2)–Cu(1)–N(3)
N(1)–Cu(1)–N(3)
C(1)–N(1)–Cu(1)
C(2)–N(2)–Cu(1)
C(3)–N(2)–Cu(1)
C(5)–N(3)–Cu(1)
C(4)–N(3)–Cu(1)
Fig. 1. Cental core of four copper centers each having six coordinated atoms.
ꢀ
a P421c space group. Least square refinements of 12032 reflections
were done for the complex. The data collected were reduced using
the SAINT program [42]. The structure was obtained by direct method
[43] using SHELXS-86, which revealed the position of all non-hydro-
gen atoms and refined by full-matrix least squares on F2
(SHELXL-
97) [44]. The graphic tool used was MERCURY for windows [45]. All
non-hydrogen atoms were refined anisotropically, while the hydro-
gen atoms were treated with a mixture of independent and con-
strained refinements.
3. Results and discussion
3.1. Crystal structure of [Cu4L4(ClO4)4]
ꢀ
The title compound crystallize in space group P421c and the
Fig. 2. ORTEP view of the coordination environment around each of four copper
structure contains neutral [Cu4L4(ClO4)4] unit. Crystal data and
structure refinement parameters are shown in Table 1. Central core
of the four copper centers with coordinated atoms is shown in Fig. 1.
Selected bond lengths and bond angles are described in Table 2.
Each CuII center exhibits a (4 + 2) elongated distorted octahedral
CuN3O3 chromophore. The ligand Lꢀ is coordinated to each copper
to form (1) a five-membered chelate ring involving the oxime nitro-
gen and the imine nitrogen and (2) a five-membered chelate ring
involving the imine nitrogen and amine nitrogen donor atoms.
The fourth position in the base of the octahedral is occupied by
the oxygen atom of the second Lꢀ ligand, fifth coordination site of
the copper is occupied by an oxygen atom third Lꢀ ligand resulting
in bridging oxime groups. The oxygen atom of the perchlorate group
is at sixth coordination site of the each copper. Fig. 2 shows coordi-
nation environment around each of the four copper centers in the
complex. The bond length between the copper center and the equa-
torial oxygen [Cu1–O1 = 1.937(4) Å] was significantly shorter than
that of vertical oxygen atoms (Cu1–O1ii = 2.573(3) Å and Cu1–
O2i = 2.628(4) Å). This may be due to the steric repulsion of the che-
late rings and also due to the Jahn–Teller effect of Cu(II). The Cu–N
centers in the complex.
bond lengths [1.945(5)–2.057(4) Å] are within the expected range
[46].
Tetrameric copper cage has two Cu2N2O2 chelate rings with two
diatomic –N–O– bridges between adjacent copper atoms having a
copper. . ..copper separation of 4.415(1) Å and four five-membered
Cu2NO2 chelate rings with one diatomic –N–O– bridge and one
monoatomic –O– bridge having copper. . .copper separation of
3.525(1) Å. In the tetramer four Cu(II) atoms and four oxygen
atoms alternatingly occupy the vertices of a cubane where distance
of eight of twelve edges are longer. Hence, the tetranuclear unit
forms an distorted open-cubane Cu4O4 core in the complex with
idealized S4-symmetry (Fig. 3(a)). The open-cubane is strongly dis-
torted as the Cu–O bond lengths and O–Cu–O bond angles vary
from 1.937(4)–3.019(3) Å and 75.33(1)–88.71(1)°, respectively. In
Cu2NO2 and Cu2N2O2 chelate rings, Cu(1)–O(1)–N(1) bond angles
are 113.8(3)° and 114.5(3)°, respectively which are less than 120°
indicating a variation in sp2 hybridization for the oxime oxygen

H. Pagonda et al. / Inorganica Chimica Acta 392 (2012) 478–484
481
3.2. Characterization of the complex
The reaction of Cu(ClO4)2ꢂ6H2O with HL led to the formation of
complex [Cu4L4(ClO4)4] (Scheme 1). The new compound is green
colored solid insoluble in common organic solvents but readily sol-
uble in acetonitrile, dimethylformamide and dimethylsulfoxide.
The molar conductivity (19
the complex is non-electrolyte in nature [48]. Electronic spectrum
of the complex (in DMF) shows strong intense band at
32051 cmꢀ1 is due to p⁄ transition of >C@N–OH group. The
X
ꢀ1 cmꢀ2 molꢀ1) value indicates that
a
p–
weak intense band at 24155 cmꢀ1 is due to ligand to metal charge
transfer transition (LMCT) which may occur from the p-orbital of
coordinate oxygen/nitrogen to the vancant d-orbital of copper
[49]. A single d–d band at 17301 cmꢀ1 (578 nm) indicates square
based geometry around copper center [50].
The peak due to
m(N–H) vibration of the substituted amino
group is observed at 3267 cmꢀ1 in the IR spectrum of the complex.
Fig. 3a. Distorted open-cubane Cu4O4 core in tetranuclear complex having ideal-
Two strong bands observed at 1662 and 1531 cmꢀ1 are assigned to
ized S4 symmetry.
imino
m
(C@N) and oxime
m
(C@N), respectively. The
m
3 (116ꢀ5, 1106
and 1008 cmꢀ1) and m4 (710 and 624 cmꢀ1) modes of ClO4 anion
ꢀ
undergo splitting. This observation suggests the binding of ClO4
anion to copper(II) [50]. This is confirmed by the X-ray crystal
structure.
The complex [Cu4L4(ClO4)4] in 0.1 M tetrabutyl ammonium
hexaflorophosphate (TBAPF6) supporting electrolyte in DMF shows
quasi irreversible cyclic voltammetric response. The Cu(II)/Cu(I)
reduction peak is observed at 0.804 V versus Ag/AgCl reference
electrode with ic/ia = 0.104 V and
100 mV/S (Fig. 4), respectively. The
D
Ep value 277 mV at a scan rate
G° (695 kcal) value of the
D
complex in DMF indicates that the complex is stable in solution
state.
3.3. Magnetic moment and EPR spectra
The magnetic moment of complex at room temperature is
found to be 1.32 BM. The found value is much less when compared
with the spin only value (1.73 BM). The observed value indicates
the presence of magnetically coupled metal centers. EPR spectra
of the complex were recorded in solid state and in DMF solvent
at room temperature and at liquid nitrogen temperature. X-band
ESR spectra of [Cu4L4(ClO4)4] are shown in Fig. 5. The spin Hamil-
tonian and orbital reduction parameters of the complex are given
in Table 3. The g|| and g\ values are computed from the spectrum
using tetracyanoethylene (TCNE) free radical as ‘g’ marker. From
Fig. 3b. Half chair conformation of Cu2N2O2 chelate ring.
atom resulting from coordination to the second copper atom. The
Cu(1)–N(1)–O(1) bond angle (126.6(3)°) indicate sp2 hybridization
of nitrogen as expected [47]. The two Cu2N2O2 chelate rings have
half chair conformation as shown in Fig. 3(b).
N2
HN3
N1
Cu
O1
O3ClO
HN3
OH
OClO3
Cu
O1
Cu
N1
Cu(ClO4)2.6H2O
N2
N
N2
Et3N,MeOH
N1
N1
O1
O1
N3H
N
N
Cu
O3ClO
OClO3
N2
N3H
HL
Cu4L4(ClO4)4
Scheme 1.

482
H. Pagonda et al. / Inorganica Chimica Acta 392 (2012) 478–484
conditions G values are found to be <4 for the complex. According
to Hathaway, if G value is greater than 4, the exchange interaction
is negligible whereas G value less than 4 indicates considerable ex-
change interaction between metal ions in the solid complex. In the
present case, G value indicates considerable exchange interaction
between Cu(II) ions. The solution ESR spectrum of complex re-
corded in DMF at 300 K shows four-hyperfine signals (giso = 2.097,
Aiso = 70G) which were not observed at 77 K. The appearance of a
four-line pattern in solution at room temperature may be due to
slight loss of magnetic coupling between the copper ions or disso-
ciation of the complex into monomers. At the same time, disap-
pearance of the four hyperfine signals in ESR spectrum at 77 K
can be attributed to the recombination of the monomers into tet-
ramers due to an increase in the viscosity of the solvent [52].
3.4. DNA binding studies
Fig. 4. Cyclic voltammetric profile of complex at scan rates (a) 25, (b) 50, (c) 75, (d)
The binding interaction of the complexes with CT-DNA was mon-
itored by comparing it’s absorption spectra with and without CT-
DNA. Addition of increasing amounts of CT-DNA to the complex
shows a decrease in molar absorptivity (hypochromism 13.8%) of
100 mV sꢀ1
.
the
p–
p⁄ absorption band as well as a red shift of ꢄ2 nm. This obser-
vation indicates the binding of the complex to DNA in different
modes and to different extents. Fig. 6 shows absorption spectra of
[Cu4L4(ClO4)4] in the absence and presence of increasing amounts
of DNA. The change in the absorbance values with increasing
amounts of CT-DNA are used to evaluate the intrinsic binding con-
stant Kb. Intercalation of a complex to DNA generally results in large
hypochromism and significant red shift of the absorption band due
to strong stacking interaction between the aromatic chromophore
of the ligand and the base pairs of the DNA [53–55]. In the absence
of any intercalative ligands in the present complex, the observed red
shift is small. The intrinsic equilibrium DNA binding constant value
of the complex is found to be 1.23 ꢃ 105 Mꢀ1. The low value sug-
gests greater surface aggregation or DNA groove binding in prefer-
ence to intercalative mode of binding [56]. The spectra do not
show any significant shift in the absorbance maximum or any de-
crease in the intensity of band indicating that the complex binds
to only one strand of DNA. This suggests that the complex is a groove
or surface binder to the CT DNA.
3.5. Nuclease activity of complex
Nuclease activity of complex has been studied by agarose gel
electrophoresis using pBR 322 plasmid DNA in Tris–HCl/NaCl
(50 mM/5 mM) buffer (pH 7) in the absence of external agent
and in the presence of H2O2 (an oxidant), DMSO (a free radical
scavenger), EDTA (a chelating agent) and DTT (a reducing agent)
(Fig. 7). In the presence of H2O2 the complex cleaves the super-
coiled DNA (Form I) into nicked DNA (Form II) and further cleaves
to linear DNA (Form III); (lane 4). This may be attributed to the for-
mation of hydroxyl free radicals. Hydroxyl radicals are generated
by a variety of chemical and physical processes. Transition-metal
mediated hydroxyl radical production is known to occur by a num-
ber of routes; one well known pathway is the Fenton mechanism
[57]. These OHꢂ free radicals participate in the oxidation of the
Fig. 5. (A) X-band powder ESR spectrum of [Cu4L4(ClO4)4] at 300 K. (B) At liquid N2
temperature, (C) in DMF solution at 300 K and (D) in DMF at liquid N2 temperature.
the observed values of complex it is clear that g|| > g\ > 2.00 which
suggests the fact that the unpaired electrons lies predominantly in
the dx2
orbital [51] characteristic of square pyramidal or octahe-
ꢀy2
dral geometry for the copper(II) complex. The gav value for the
complex is greater than 2 indicating covalent nature of the me-
tal–ligand bond. In the solid state, similar spectra of the complex
at 77 and 300 K indicate that the geometry around copper(II) ion
is unaffected on cooling to liquid nitrogen temperature. In these
Table 3
Spin Hamiltanian and orbital reduction parameters for the copper(II) complex.
Complex
At 300 and At 77 K (solid state)
At 300 K (solution state)
At 77 K (solution state)
g||
g\
gav
G
giso
Aiso G (10ꢀ3 cmꢀ1
)
g||
g\
G
A||G (10ꢀ2 cmꢀ1
)
A\G (10ꢀ3 cmꢀ1
)
[Cu4L4(ClO4)4]
2.132
2.074
2.093
1.814
2.097
70 (6.8)
2.238
2.054
4.592
170 (1.77)
20.0 (1.92)

H. Pagonda et al. / Inorganica Chimica Acta 392 (2012) 478–484
483
Fig. 6. Absorption spectra of complex II in the absence and presence of increasing amount of DNA. Arrow shows the decrease in absorbance upon increasing conc. of DNA
[0–20 M].
l
Although a few structurally characterized tetranuclear copper
complexes with macrocyclic ligand [1], hydrazone [5] and aziridines
[46] have been reported, yet the present tetranuclear copper(II)
complex with the oxime-schiff base is new in the literature. The
present complex has oximato bridges, which is regarded as a novel
feature. The open-cubane structure of present complex is well com-
parable with the one reported [5] recently.
Acknowledgements
Fig. 7. Agarose gel (0.8%) showing results of electrophoresis of 1
l
l of pBR 322
of
ll of sterilized water; 2 ll of H2O2 (total
Plasmid DNA;
4 ll of Tris–HCl/NaCl (50 mM/5 mM) buffer (pH-7); 2 ll
The authors are thankful to the DST, New Delhi (Sanction No.
SR/S1/IC-37/2007) for financial support and SAIF (RSIC), Indian
Institute of Technology, Bombay, for providing electron spin reso-
nance spectra. Yogesh patil is thankful to CSIR, India for the grant
of research fellowship.
[Cu4L4(ClO4)4] in DMF (1 ꢃ 10ꢀ3 M) ; 11
volume 20 ll) were added, respectively, incubated at 37 °C (30 min); lane 1: DNA
control; lane 2: DNA control + H2O2; lane 3: complex (100
lM) + DNA; lane 4:
complex + DNA + H2O2; lane 5: complex + DNA + DTT;
lane 6: com-
plex + DNA + NaN3; lane 7: complex + DNA + DMSO; lane 8: complex + DNA + EDTA.
deoxyribose moiety, followed by the hydrolytic cleavage of the su-
gar phosphate back bone [36].
Appendix A. Supplementary material
Cleavage activity of the complex increases in the presence of
reducing agent (DTT) due to the formation copper(I) complex by
catalytic reduction (lane5). The chelating agent EDTA efficiently
inhibits the activity of complex. As shown in Fig. 7 (lane 7) nucle-
ase activity of complex is decreased in the presence of DMSO
which acts as scavengers of free radicals. The observation suggests
the involvement of the hydroxyl radical in the cleavage process in
support of oxidative mechanism.
CCDC 827005 contains the supplementary crystallographic data
for this paper. These data can be obtained free of charge from The
Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/
data_request/cif. Supplementary data associated with this article
can be found, in the online version, at http://dx.doi.org/10.1016/
j.ica.2012.03.042.
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