Alkyl chain length effect on construction of copper(II) complexes with tridentate Schiff base ligand and DNA interaction

Article abstract of DOI:10.1080/00958972.2016.1163547

Two tridentate Schiff base ligands were synthesized by condensation of equimolar amounts of benzoylacetone and 2-amino-1-ethanol or 3-amino-1-propanol, H₂L¹ and H₂L², respectively. The reaction of the Schiff bas

Full text of DOI:10.1080/00958972.2016.1163547

Journal of Coordination Chemistry
ISSN: 0095-8972 (Print) 1029-0389 (Online) Journal homepage: http://www.tandfonline.com/loi/gcoo20
Alkyl chain length effect on construction of
copper(II) complexes with tridentate Schiff base
ligand and DNA interaction
Rasoul Vafazadeh, Fariba Jafari, Mohammad Mehdi Heidari & Anthony C.
Willis
To cite this article: Rasoul Vafazadeh, Fariba Jafari, Mohammad Mehdi Heidari & Anthony
C. Willis (2016): Alkyl chain length effect on construction of copper(II) complexes with
tridentate Schiff base ligand and DNA interaction, Journal of Coordination Chemistry, DOI:
10.1080/00958972.2016.1163547
To link to this article: http://dx.doi.org/10.1080/00958972.2016.1163547
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Date: 08 March 2016, At: 18:50

Publisher: Taylor & Francis
Journal: Journal of Coordination Chemistry
DOI: http://dx.doi.org/10.1080/00958972.2016.1163547
Alkyl chain length effect on construction of copper(II) complexes with
tridentate Schiff base ligand and DNA interaction
RASOUL VAFAZADEH*†, FARIBA JAFARI†, MOHAMMAD MEHDI HEIDARI‡ and
ANTHONY C. WILLIS§
†Department of Chemistry, Yazd University, Yazd, Iran
‡Department of Biology, Yazd University, Yazd, Iran
§Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
Two tridentate Schiff base ligands were synthesized by condensation of equimolar amounts of
benzoylacetone and 2-amino-1-ethanol or 3-amino-1-propanol, H2L1 and H2L2, respectively. The
reaction of the Schiff base ligands with Cu(CH3COO)2 in methanol leads to (CuL1)4, 1 and
(CuL2)2, 2. In the tetranuclear cubane species, the tridentate H2L has both a chelating and a
bridging mode, after double deprotonation of the enolic OH groups. The copper(II) centers are
five-coordinate with a NO4 donor set from the ligands. The coordination geometry around each
copper ion is essentially square pyramidal with one nitrogen and two oxygens from one ligand
and two oxygens of adjacent ligands from the next unit of the cubane. In dinuclear 2, H2L2 has
chelating and bridging modes after double deprotonation of the enolic OH groups. The dianionic
form of the Schiff base coordinates forming a six-membered chelate ring with Cu(II). Two such
monomeric CuL2 entities are eventually linked through the alkoxo bridges to produce
dinuclear 2. The absorption spectra strongly suggest that 2 interacts with CT-DNA. Both 1 and 2
appear to be more efficient than the parent compound in DNA cleavage.
Keywords: Copper complex; Tridentate Schiff base; Tetranuclear; Dinuclear; DNA Interaction
*Correspondencing author. Email: rvafazadeh@yazd.ac.ir
1

1. Introduction
The design and synthesis of multinuclear transition metal complexes of Schiff bases have
attracted research due to their relevance in bioinorganic chemistry [1-3], molecular magnetic
materials [4-6], and the development of coordination chemistry [6-12]. There has been growing
interest in using the self-assembly process in synthesis of multinuclear metal complexes. The
most common ligands used for construction of polynuclear complexes by self-assembly are
tridentate Schiff base ligands, which contain potentially bridging phenoxo or hydroxo oxygen
and nitrogen donors [13-16].
Multinuclear copper complexes have attracted attention with interesting structures
[17-20] and because copper is involved in many important biological activities [20-22].
Investigations of the interaction between multinuclear copper(II) complexes and DNA have
attracted interest due to their importance in molecular biology [14, 23-29].
Herein, we report the synthesis, spectroscopic characterization, structural aspects and
DNA interaction of one tetranuclear cubane (CuL1)4 and one dinuclear (CuL2)2 Schiff base
complex. Two Schiff base ligands, H2L1 and H2L2 (with ONO donors), were obtained from 1:1
condensation of benzoylacetone and 2-amino-1-ethanol or 3-amino-1-propanol, respectively
(scheme 1). The coordination sphere around copper(II) can be modified easily and led to a
number of Cu(II) complexes with different nuclearity.
The syntheses of 1 and 2 were previously reported by the template method and their
magnetic properties studied. Molecular weight measurements in chloroform on two different
structures for these complexes have been suggested [30]. However, the synthesis of the free
Schiff base ligands and crystallographic data of the complexes have not been reported.
2. Experimental
2.1. Reagents
All chemicals were used as supplied by Merck and Fluka without purification. Supercoiled
pBR322 DNA was obtained from Sigma–Aldrich and stored at 4 °C. The
tris(hydroxymethyl)aminomethane–HCl (Tris–HCl) buffer was prepared in doubly distilled
water.
2

2.2. Physical measurements
Infrared spectra were taken with an Equinox 55 Bruker FT-IR spectrometer using KBr pellets
from 400-4000 cm-1. Absorption spectra of the ligands were determined in methanol and for 1
and 2 in DMF using a GBC UV-Visible Cintra 101 spectrophotometer with 1 cm quartz cells,
from 200-800 nm at 25 °C. Elemental analyses (C, H and N) were performed using a CHNS-O
2400II PERKIN-ELMER elemental analyzer.
2.3. DNA binding
Interaction of the complexes with calf thymus CT-DNA was studied in Tris–HCl buffer (5.5 mM
Tris–HCl, pH 7.2) containing 50 mM NaCl at room temperature. The solution was kept for over
24 h at 4 °C. The resulting, somewhat viscous, solution was clear and particle-free. CT-DNA in
the buffer medium gave a ratio of UV absorbance at 260 and 280 nm of 1.8:1, indicating that the
DNA was sufficiently free of protein [31]. The DNA concentration was measured from its
absorption intensity at 260 nm using the molar absorption coefficient (ε) value of 6600 M−1 cm−1
as reported [32]. Stock solutions of complexes were freshly prepared by dissolving the
complexes in DMF and diluting them with the buffer. The amount of DMF was kept at 10% (by
volume) for each set of experiments and has no effect on the experimental results. Absorption
spectral titrations were performed while maintaining a constant complex concentration and
varying the nucleic acid concentration. This was achieved by dissolving an appropriate amount
of the complexes (40 μM) and DNA stock solutions (0 - 54 μM) while maintaining the total
volume constant (1 mL). Spectral bands were recorded after successive addition of CT-DNA.
The Tris–HCl buffer was used as a blank to make preliminary adjustments. The intrinsic binding
constant (Kb) of the complex to CT-DNA was determined from the spectral titration data using
the following equation [33]:
[DNA]/(εa - εf) = [DNA]/(εb - εf) + 1/Kb(εb - εf)
(1)
where [DNA] is the concentration of CT-DNA in base pairs and the apparent absorption
coefficients εa, εf and εb correspond to Aobs/[DNA], the absorbance for the free-Cu(II) complex
(unbound), and the absorbance for the fully-bound complex, respectively. A plot of [DNA]/(εa -
εf) versus [DNA] gave a slope 1/(εb - εf) and a intercept 1/Kb(εb - εf), so the value of Kb can be
determined from the ratio of the slope to the intercept.
3

2.4. DNA Cleavage
Cleavage of plasmid DNA was monitored using agarose gel electrophoresis following the
literature method [34-36]. Supercoiled pBR322 DNA (0.1 mg/mL, 1.5 μL) in Tris buffer (pH =
7.2, 5.0 mM Tris-HCl, 20 mM NaCl) with 50 mM NaCl was treated with varying concentrations
of the copper(II) complexes (40-600 μM). The concentration of the complexes in the buffer
corresponded to the quantity after the dilution of the complex stock to the 20 μL final volume
using Tris buffer. The samples were incubated for 2 h at 37 °C, followed by addition of the
loading buffer containing 12.5% bromophenol blue, 25% xylene cyanol, 1% tris, and the solution
was finally loaded on agarose (1%) containing 10 mg/mL ethidium bromide (EB). The
electrophoresis was carried out at 60 V for 3 h in Tris–acetate–EDTA. Afterwards,
electrophoresis bands were visualized by UV light and photographed. The cleavage mechanism
of pBR322 DNA was measured in the presence of H2O2 (oxidizing agent), DMSO (hydroxyl
scavenger) and NaN3 (singlet oxygen scavenger).
2.5. X-ray Crystallography
Diffraction images were measured at 200 K on a Nonius Kappa CCD diffractometer using
Mo Kα, graphite monochromator (λ = 0.71073 Å) and data extracted using the
DENZO/SCALEPACK package [37]. The structures were solved by direct methods with SIR92
2
[38]. The structures were refined on F by full matrix last-squares using the CRYSTALS
program package [39]. Hydrogens were added at calculated positions and then refined with soft
restraints on the bond lengths and angles to regularize their geometry. In 1 the hydrogens were
then refined with riding constraints, whereas in 2 they were allowed to refine freely. Atomic
coordinates, bond lengths and angles, and displacement parameters have been deposited at the
Cambridge Crystallographic Data Centre. Crystallographic details for 1 and 2 are summarized in
table 1.
2.6. Syntheses
2.6.1. Syntheses of H2Ln. Two tridentate Schiff bases, H2L1 and H2L2, were synthesized by a
general method using the condensation of an equimolar amount of benzoylacetone (5 mmol,
0.81 g) and the related amine (5 mmol), 2-amino-1-ethanol (0.3 mL) and 3-amino-1-propanol
4

(0.38 mL) in methanol (30 mL), respectively [40, 41]. Then the mixture was refluxed for 2 h,
giving yellow micro crystals. The micro crystals were filtered off and re-crystallized in
acetone/ethanol at room temperature.
H2L1: Yield: 0.89 g (87%). Anal. Calc. for C12H15NO2 (205.26): C, 70.22; H, 7.37; N,
6.82%. Found: C, 70.51; H, 7.45; N, 6. 66%. IR (KBr, cm−1): νOH 3357, νC=N 1602. Electronic
spectra in CH3OH λmax(nm), (log ε): 237 (3.46), 342 (3.60).
H2L2: Yield: 0.89 g (81%). Anal. Calc. for C13H17NO2 (219.28): C, 71.21; H, 7.81; N,
6.39%. Found: C, 71.48; H, 7.93; N, 6. 27%. IR (KBr, cm−1): νOH 3379, νC=N 1602. Electronic
spectra in CH3OH λmax(nm), (log ε): 243 (3.41), 343 (3.65).
2.6.2. Syntheses of (CuL1)4, 1 and (CuL2)2, 2. Complexes 1 and 2 were synthesized by a
general method. Copper(II) acetate monohydrate (2 mmol 0.396 g) was slowly added to a
methanol solution (30 mL) of the appropriate Schiff base (2 mmol, 0.41 g for 1 and 0.438 g
for 2). The resulting solution was stirred for 2 h at room temperature. The resulting dark-green
precipitates were collected by filtration and dried in air.
(CuL1)4, 1. Dark-green needles suitable for X-ray analysis appeared upon slow
evaporation from dichloromethane/methanol (1:1 v/v). Yield 72% (0.38 g). Anal. Calcd. for
(C12H13CuNO2)4: C, 54.02; H, 4.91; N, 5.25. Found: C, 53.81; H, 4.79; N, 5.21. IR (KBr, cm-1):
νC=N = 1599, νC-O = 1331. Electronic spectra in DMF solvent λmax, nm (log ε): 638 (1.40), 346
(3.64).
(CuL2)2, 2. Dark-green needle-shaped crystals appeared upon slow evaporation from
dichloromethane/methanol (1:1 v/v). Yield 63% (0.35 g). Anal. Calcd. for (C13H15CuNO2)2: C,
55.60; H, 5.38; N, 4.99. Found: C, 55.88; H, 5.50; N, 5.07. IR (KBr, cm-1): νC=N = 1590, νC-O
=
1354. Electronic spectra in DMF solvent λmax, nm (log ε): 584 (1.26), 345 (3.66).
3. Results and discussion
3.1. Synthesis and characterization of the complexes
Syntheses of 1 and 2 are schematically represented in scheme 1. The tridentate Schiff base
ligands were obtained by condensation of benzoylacetone and amino-alcohol under reflux. The
reaction of copper(II) acetate with an equimolar amount of H2L1 in methanol leads to
tetranuclear 1. When H2L2 was used for the synthesis, dinuclear 2 was formed.
5

The most significant IR bands for the ligands and complexes are given in the
Experimental section. The IR spectra of the free Schiff bases show a band at 1602 cm-1, assigned
as νC=N. IR spectra of 1 and 2 show a decrease in νC=N in comparison with the free ligand,
which indicates coordination of the imine nitrogen to copper [12, 41, 42]. The fairly broad band
of medium intensity at 3357 and 3379 cm−1 corresponds to intramolecular hydrogen bonding in
the free ligands, and this band disappears for the copper complexes. The lack of OH groups in
the copper complexes indicates that the OH groups have been deprotonated and are coordinated
to copper.
The electronic spectra of the copper complexes in DMF show a broad band at 638 (for 1,
log ε = 1.40) and 584 nm (for 2, log ε = 1.26) and a sharper signal at 346 (for 1, log ε = 3.64) and
345 nm (for 2, log ε = 3.66), which arise from a spin-allowed d-d transition of the copper(II) ion
(d9 electronic configuration) and a charge transfer transition, respectively [43, 44].
3.2. Crystal structure of 1
The molecular structure of 1 is shown in figure 1. Selected bond lengths and angles as well as
interatomic distances are summarized in table 2. Complex 1 crystallizes in trigonal space
group R3.
In the copper complex, H2L1 has both chelating and bridging modes after double
deprotonation of the enolic OH groups. The dianionic form of the Schiff base ligand coordinates
to the metal center through the imine nitrogen and deprotonated alkoxo oxygens. By a self-
assembly process, four monomeric CuL1 entities are linked through alkoxo bridges to produce
the tetranuclear cubane, 1.
In 1, the four alkoxo oxygens are located at the four corners of the cube and bridge the
metal centers in a μ3-fashion (O2: Cu1, Cu2, Cu3; O4: Cu2, Cu3, Cu4; O6: Cu1, Cu3, Cu4;
O8: Cu1, Cu2, Cu4). Intracluster metal/metal separations of adjacent Cu···Cu distances are
3.105−3.399 Å, which are comparable with values of similar compounds [14, 45, 46]. The
copper(II) centers are five-coordinate with a NO4 donor set from the Schiff base ligands. The
coordination geometry about each copper ion is a square pyramid with one nitrogen and two
oxygens from the Schiff base ligand and two oxygens from the next unit of the cubane.
According to the bond lengths between the copper and coordinating atoms (i.e., four bonds with
short bond distances, 1.908(3)−1.984(3) Å and bonds with long bond distances,
6

2.375(3)−2.622(3) Å) the basal planes for Cu1, Cu2, Cu3 and Cu4 are O1−N1−O2−O8,
O2−N2−O3−O4, O4−N3−O5−O6 and O6−N4−O7−O8, respectively. The Cu ions in the
tetranuclear species deviate from the corresponding mean planes by 0.067−0.144 Å, towards the
apical ligand. The coordination spheres of the copper ions in the complex are best described as a
distorted square pyramid according to the Addison parameter τ values of 0.05−0.19. The
parameter τ is defined as τ = (α-β)/60, α>β, where α and β are the largest angles, with τ = 1 for a
regular trigonal bipyramid and τ = 0 for regular square pyramid [47].
The Cu–O and Cu–N bond lengths in the equatorial plane are 1.908−1.984 Å and
1.920−1.940 Å, respectively (table 2), which agree with similar compounds [12, 14, 41]. The
apical oxygens show longer Cu–O(alkoxo) bond lengths than the equatorial oxygens [43, 48].
The elongation of the Cu–O axial bonds is due to a pseudo-Jahn–Teller distortion of the d9
copper center. The lengths of the bonds between copper and the donor atoms are within the range
of values normally found for such systems [14, 19, 49, 50].
The bridging bond angles of Cu–O(alkoxo)–Cu are 89.26–112.25° (table 2). It is evident
from the different Cu–O body diagonal distances as well as from the unequal metal–metal
distances (table 2) that the cubane core is not a regular one, but distorted. In tetranuclear 1, the
dihedral angles of adjacent phenyl rings are between 51–55°.
In the absence of suitably polar hydrogens there is no hydrogen bonding in the intra- and
intermolecular interaction of the complex. However, the 3D lattice structure of 1 shows the
presence of intermolecular non-covalent C–H ····· N=C and C–H(Ph) ····· C–H(Ph) interactions
(figure 2).
3.3. Crystal structure of 2
The molecular structure of 2 is shown in figure 3. Selected bond lengths and angles as well as
interatomic distances are summarized in table 2; 2 crystallizes in the monoclinic space group
P21/n.
In 2, similar to 1, H2L2 is both chelating and bridging after double deprotonation of the
enolic OH groups. The dianionic form of the ligand coordinates, forming six-membered chelate
rings with Cu(II). Two such monomeric CuL2 entities are eventually linked through the alkoxo
bridges to produce the dinuclear 2.
7

The dinuclear unit is formed by two Cu(II) ions labeled Cu1 and Cu1* (symmetry code:
−x+1, −y+1, -z+1), bridged by the two μ2−alkoxo oxygens of the Schiff base ligands.
Coordination about each copper ion is square planar with one nitrogen and two oxygens from the
Schiff base and one oxygen from the next unit of the dimer. The Cu−O, Cu−N, and two
Cu−Obridge bond distances are 1.9071(12), 1.9455(13), 1.9313(11) and 1.9197(11) Å,
respectively, in agreement with those observed for similar compounds [12, 14, 49, 51]. As
usually found in oxo-bridged Cu(II) complexes [12, 14, 52, 53], the Cu-Obridge bond distances
(1.9313(11) and 1.9197(11) Å) are slightly longer than the chelating Cu-O bond length
(1.9071(12) Å) because of the weaker interaction of the binding alkoxo bridging with copper.
The distance between the two Cu ions is 3.0341(4) Å, indicating the absence of any bond
between the copper centers.
The angles involving the copper ions are different from 90° and 180°. The N–Cu–O trans
angle is 172.87(5)°, the O–Cu–O trans angle is 167.26(5)° and the N–Cu–O cis angles are
95.03(5) and 96.56(5)° (table 2), indicating that there is distortion from ideal square planar
geometry. The geometry of copper is best described as distorted square planar with τ4 index
0.141. The τ parameter is τ4 = [360° − (α + β)]/141°, where a and b are the largest angles around
the central metal in the complex, τ4 = 1 for a regular tetrahedron and τ4 = 0 for a regular square
planar [54]. The copper ions lie 0.043 Å above the NO3 donor planes.
In 2 there are no intramolecular interactions. The crystal packing of the complex shows
the presence of intermolecular non-covalent interactions between CH2 units of the propylene to
form a 2D planar network parallel to the ab plane (figure 4).
3.4. DNA Binding studies
UV–Visible spectroscopy was performed to study the interaction of the complexes with DNA
keeping the complex concentration constant (40 μM) and varying the concentration of CT-DNA
(0 – 54 μM). The change in absorbance values at 260 nm was used to evaluate the intrinsic
binding constant Kb. The absorption spectra of 2 in the absence and presence of CT-DNA (at
constant concentration of complex) are shown in figure 5. Hypochromicity of the DNA at
265 nm without a shift in band position is observed with the addition of CT-DNA.
The hypochromism (decrease of absorbance) indicated a strong interaction between the
complexes and CT-DNA mainly through intercalative binding [55, 56]. The non-covalent
8

intercalative binding of the compound to the DNA helix can lead to large hypochromism due to
the strong stacking interaction between the aromatic moiety of the complex and the base pairs of
DNA [57, 58]. Complex 2 binds to DNA through intercalation and resulted in hypochromism
due to the stacking effect of π electrons which leads to a decrease in the peak intensities in the
UV spectra of complex [59, 60].
Analysis of the spectral data using Equation 1 in the presence of DNA (figure 5) gave a
binding constant, Kb, of (4.10 0.21)×104 M−1 for 2. The Kb value is lower than that observed for
classical intercalator EB (order of magnitude of 106 M−1) [61], indicating that the compounds
bind DNA with less affinity. However, the Kb value for 2 is comparable to that observed for
copper(II) complexes, such as [CuL1](ClO4)2 (L1 = N,N′-bis-pyridin-2-ylmethyl-butane-1,4-
diimine), 2.6×104 M−1 [62] and [Cu(bpy)2(pic)](pic) (pic = picrate), 2.6×104 M−1 [63] and higher
than [CuL2] (HL = N-(3-hydroxybenzyl)-leucine acid), 1.55×103 M−1 [64],
[Cu(bpy)(Gly)Cl]·2H2O, 1.84×103 M−1, [Cu(dpa)(Gly)Cl]·2H2O, 3.10×103 M−1 [65],
[Cu(L)(bpy)Cl] (HL = (E)-3-(2-hydroxyphenylimino)-N-o-tolylbutanamide), 1.55×103 M−1 [66]
[Cu2(μ-Cl)2(O-2-alkoxyethylpyridine-2-carboximidate)2Cl2] complexes, where alkoxy =
methoxy, ethoxy, and butoxy, 1.52×103, 5.59×103, and 6.36×103 M-1, respectively [67].
From the binding constant value, it was clear that 2 had good interaction with CT-DNA.
However, 1 and free ligands H2L1 and H2L2 exhibited no significant DNA bonding.
3.5. Supercoiled pBR322 plasmid DNA cleavage studies
3.5.1. DNA cleavage with 1. DNA cleavage was analyzed by monitoring the conversion of
supercoiled DNA (form I) to nicked circular DNA (form II) and linear DNA (form III). When
circular plasmid DNA is subjected to electrophoresis, a relatively fast migration will be observed
for the intact supercoil form (form I). If scission occurs on one strand (nicking), the supercoil
will relax to generate a slower moving open circular form (form II). If both strands are cleaved, a
linear form (form III) that migrates between form I and form II will be generated [43, 44]. At
micro-molar concentrations, for 3 h incubation periods, in the absence of H2O2 (an oxidizing
agent) 1 exhibited no significant nuclease activity (figure 6, lanes 2-5). However, 1 displays
efficient cleavage of pBR322 circular plasmid DNA converting form I to form II and form III in
the presence of hydrogen peroxide (figure 6, lane 6), indicating an oxidative cleavage process
[36]. Control experiments using only H2O2 (200 μM), copper alone up to 200 μM and copper salt
9

in the presence of H2O2 did not show any significant DNA cleavage under similar experimental
conditions (figure 7, lanes 2, 5 and 6, respectively). The reaction of pBR322 DNA with H2L1
(200 μM) in the absence of H2O2 did not exhibit any cleavage activity (figure 7, lane 3). H2L1,
however, in the presence of H2O2 caused conversion of supercoiled structure into nicked circular
(figure 7, lane 7). The control results suggest the complex to be more efficient than copper salt
and ligand for the chemical nuclease activity.
To elucidate the cleavage mechanism of DNA, the cleavage was investigated in the
presence of hydroxyl radical (OH•) scavenger (DMSO, 1.4 mM) and a singlet oxygen (1O2)
quencher (NaN3, 1 mM). The experiments reveal that NaN3 and DMSO significantly inhibited
the cleavage activity of Cu(II) complex (figure 6, lanes 7 and 8), indicating that singlet oxygen
and hydroxyl radical play significant roles in the DNA cleavage reaction [68, 69]. The result
suggests that 1 cleaves DNA via oxidative cleavage [35, 36].
3.5.2. DNA cleavage with 2. Complex 2 exhibited DNA cleavage activity even in the absence of
an oxidant (figure 8, lanes 2-5). Control experiments using DNA alone (figure 8, lane 1) and
only H2L2 (figure 7, lane 4) in the absence of 2 resulted in no significant cleavage of pBR322
circular plasmid DNA, even after long exposure times. From the observed results, it was
concluded that 2 effectively cleaved the DNA as compared to control DNA and H2L2 alone.
Upon increasing the concentration of 2 more cleavage activity was observed (figure 8, lanes 2-5).
This shows that the concentration of optimal value led to extensive degradations, resulting in
conversion of the supercoiled form (Form I) into a nicked circular form (Form II). The activity
for DNA cleavage without an oxidant is often a hydrolytic rather than an oxidative mechanism
[43].
Also, the nuclease activity of the complex was investigated in the presence of H2O2 as an
oxidant and a hydroxyl radical scavenger, DMSO, and a singlet oxygen quencher, NaN3. In the
presence of H2O2, 2 shows higher activity and supercoiled DNA (Form I) is cleaved to form a
mixture of Form II and Form III (figure 8, lane 6). The presence of DMSO did not significantly
reduce the efficiency of DNA cleavage (figure 8, lane 7), ruling out the possibility of the
involvement of diffusible hydroxyl radicals in the cleavage in the presence of H2O2 [39, 40].
However, the addition of the singlet quencher, NaN3 significantly inhibited the cleavage activity
10

of 2 (figure 8, lane 9). The results suggest that singlet oxygen is responsible for the DNA
cleavage in the presence of H2O2 [43].
4. Conclusion
We have synthesized a tetranuclear copper(II), 1, and a dinuclear, 2, complex with two tridentate
Schiff base ligands. The complexes were structurally characterized by single crystal X-ray
diffraction analysis. Complex 1 has a cubane Cu4O4 core in which copper and oxygen are present
at alternative vertices. The coordination geometry about each copper ion is square pyramidal.
Complex 2 contains a coplanar Cu2O2 core and the coordination geometry around the copper(II)
is four-coordinate square planar. Complex 1 exhibits significant DNA cleavage activity in the
presence of H2O2 as an oxidant. The binding of the complexes with CT-DNA was studied using
UV-Vis spectroscopy. The absorption spectra indicate that 2 interacts with CT-DNA. Both
complexes are more efficient than their parent compound in DNA cleavage.
Supplementary material
The deposition numbers of 1 and 2 are CCDC 1045677 and 1045678, respectively. These data
can be obtained free of charge via www.ccdc.cam.ac.uk/data_request/cif, by emailing
data-request@ccdc.cam.ac.uk, or by contacting the Cambridge Crystallographic Data Centre,
12 Union Road, Cambridge CB2 1EZ, UK; Fax +44 1223 336033.
Acknowledgements
The authors are grateful to the Yazd University and the Australian National University for partial
support of this work.
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15

Figure captions
Figure 1. The molecular structure of tetranuclear 1 with labeling of selected atoms.
Figure 2. Packing diagram of tetranuclear cubane, 1.
Figure 3. The molecular structure of dinuclear 2 with labeling of selected atoms.
Figure 4. Fragment of the molecular packing for 2 along the c axis.
Figure 5. Absorption spectra of 1 (40 μM) in Tris–HCl buffer (pH = 7.2) with increase in molar
ratio of DNA to complex (0–54 μM). Arrow shows the absorbance change upon increase of
DNA concentration. The inset shows plot of [DNA]/(εa − εf) vs. [DNA] for titration of CT−DNA
with the complex.
Figure 6. Agarose gel electrophoresis diagram showing the chemical nuclease activity of 1 using
supercoiled pBR322 plasmid DNA (0.1 mg/mL, 1.5 μL): lane 1, DNA control; lane 2, DNA + 1
(50 μM); lane 3, DNA + 1 (100 μM); lane 4, DNA + 1 (300 μM); lane 5, DNA + 1 (600 μM);
lane 6, DNA + 1 (300 μM) + H2O2 (3.2 mM); lane 7, DNA + 1 (300 μM) + H2O2 (3.2 mM) +
DMSO (1.4 mM); lane 8, DNA + 1 (300 μM) + H2O2 (3.2 mM) + NaN3 (1 mM).
Figure 7. Agarose gel electrophoresis diagram showing the chemical nuclease activity of the
control experiments using Supercoiled pBR322 plasmid DNA (0.1 mg/mL, 1.5 µL): lane 1, alone
DNA control; lane 2, DNA + copper salt (200 µM); lane 3, DNA + H2L1 (200 µM); lane 4, DNA
+ H2L2 (200 µM); lane 5, DNA + H2O2 (3.2 mM); lane 6, DNA + + copper salt (200 µM) + H2O2
(3.2 mM); lane 7, DNA + H2L1 (200 µM) + H2O2 (3.2 mM); lane 8, DNA + H2L2 (200 µM) +
H2O2 (3.2 mM).
Figure 8. Agarose gel electrophoresis diagram showing the chemical nuclease activity of 2 using
supercoiled pBR322 plasmid DNA (0.1 mg/mL, 1.5 μL): lane 1, DNA control; lane 2, DNA + 2
(40 μM); lane 3, DNA + 2 (100 μM); lane 4, DNA + 2 (200 μM); lane 5, DNA + 2 (400 μM);
lane 6, DNA + 2 (200 μM) + H2O2 (3.2 mM); lane 7, DNA + 2 (200 μM) + H2O2 (3.2 mM) +
DMSO (1.4 mM); lane 8, DNA + H2O2 (3.2 mM); lane 9, DNA + 2 (200 μM) + H2O2 (3.2 mM)
+ NaN3 (1 mM).
16

Table 1. Crystallographic data of 1 and 2.
Compound
1
2
Chemical formula
Formula weight
Temperature (K)
Space group
Unit cell dimensions
a (Å)
C48H52Cu4N4O8
1067.15
C26H30Cu2N2O4
561.62
200
200
Trigonal, R3, Z=9
Monoclinic, P21/n, Z=2
30.1694(3)
30.1694(3)
17.7291(1)
90
7.3562(1)
9.9489(1)
16.2141(3)
90
b (Å)
c (Å)
α (°)
90
96.5724(9)
90
β (°)
120
γ (°)
V (Å3)
11610.21(18)
4932
1178.85(3)
580
F(000)
D
Calc (g cm−3)
1.374
1.582
Crystal size (mm)
μ (mm−1)
0.47×0.06×0.01
1.68
0.35×0.18×0.12
1.84
2.7 - 27.5
2.9 - 30.1
θ range (°)
Limiting indices
-39 ≤ h ≤ 39
-33 ≤ k ≤ 33
-18 ≤ l ≤ 19
11670
-10 ≤ h ≤ 10
-13 ≤ k ≤ 14
-22 ≤ l ≤ 22
3435
No. independent reflns.
No. reflns. F2 > 2σ(F2)
No. parameters
9829
3012
578
199
R[F2 > 2σ(F2)]
0.040
0.031
wR(F2) (all data)
S (all data)
0.112*
0.083**
0.94
0.99
* w = [weight]*[1-(deltaF/6*sigmaF)2]2 where [weight] = 1/[A0*T0(x) + A1*T1(x)
+···+ An-1*Tn-1(x)], where Ai = Chebychev coefficients 12.6, 18.1, 9.39, 2.80 and
x = F/Fmax
**w = 1/[σ2(F2) + (0.05P)2 + 0.73P], where P = (max(Fo ,0) + 2Fc2)/3
2
17

Table 2. Selected bond lengths (Å) and angles (°) in 1 and 2a.
Complex 1
Cu1–O1
Cu1–O2
Cu1–O6
Cu1–O8
Cu1–N1
Cu2–O2
Cu2–O3
Cu2–O4
Cu2–O8
Cu2–N2
Cu3–O2
Cu3–O4
Cu3–O5
Cu3–O6
Cu3–N3
Cu4–O4
Cu4–O6
Cu4–O7
Cu4–O8
Cu4–N4
Cu1···Cu2
Cu1···Cu3
1.908(3)
1.963(3)
2.375(3)
1.956(3)
1.934(4)
1.964(3)
1.917(3)
1.972(3)
2.387(3)
1.940(4)
2.622(4)
1.984(3)
1.908(3)
1.960(3)
1.923(4)
2.416(3)
1.966(3)
1.918(3)
1.961(3)
1.920(4)
3.016(1)
3.399(1)
O1–Cu1–O2
O8–Cu1–N1
O6–Cu1–N1
O2–Cu1–O6
O3–Cu2–O4
O2–Cu2–N2
O8–Cu2–N2
O3–Cu2–O8
O5–Cu3–O6
O4–Cu3–N3
O2–Cu3–N3
O4–Cu3–O5
O7–Cu4–O8
O6–Cu4–N4
O4–Cu4–N4
O6–Cu4–O7
Cu3–O2–Cu2
Cu2–O2–Cu1
Cu4–O4–Cu3
Cu2–O4–Cu3
Cu1–O6–Cu4
Cu1–O6–Cu3
179.05(16)
167.92(18)
113.33(16)
83.71(13)
173.36(14)
168.52(15)
109.13(14)
98.43(13)
173.50(16)
170.33(15)
108.15(14)
94.11(14)
174.98(14)
163.41(15)
117.77(15)
96.40(14)
89.26(12)
104.55(15)
91.36(12)
112.25(16)
92.18(12)
102.85(14)
Complex 2
Cu1–O2*
Cu1–O1
Cu1–O2
Cu1–N1
O1–C7
1.9313(11)
1.9071(12)
1.9197(11)
1.9455(13)
1.297(2)
O2*–Cu1–O1
O2*–Cu1–O9
O1–Cu1–O2
O2*–Cu1–N1
O1–Cu1–N1
O2–Cu1–N1
92.01(5)
76.03(5)
167.26(5)
172.87(5)
95.03(5)
96.86(5)
O2–C13
1.4108(19)
Cu1···Cu1*
3.0341(4)
Cu1–O2*–Cu1*–O2
0.0
Cu1–O2*–Cu1*–N1* 179.45(6)
a symmetry code: –x+1, –y+1, –z+1
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