A.A. Abdel Aziz, H.A. Elbadawy / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 404–415
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attention [1–4]. Additionally, it has been demonstrated that free
radicals can damage proteins, lipids, and DNA of bio-tissues, lead-
ing to increased rates of cancer [5]. Fortunately, antioxidants can
prevent this damage, due to their free radical scavenging activity
[6]. Hence, it was very important to develop compounds with both
strong antioxidant and DNA-binding properties for effective cancer
therapy. Transition metal complexes of Schiff bases have been
widely exploited to develop synthetic binding and cleavage agents
for DNA [7–9].
Ruthenium’s properties are well suited towards pharmacologi-
cal oxidation states (II–IV) under physiologically relevant condi-
tions [10]. Also, the energy barrier for interconversion between
these oxidation states is relatively low, allowing for ready
oxidation state changes when inside the cell [11]. Furthermore,
ruthenium tends to form octahedral complexes, which gives the
chemist two more ligands to exploit compared with platinum(II)
complexes, which adopt a square planar geometry and can also
form strong chemical bonds with a range of different elements of
varying chemical ‘hardness’ and electronegativities, meaning that
ruthenium can bind to a range of biomolecules, not just DNA [12].
Ru(II) and Ru(III) complexes are presently an object of great
attention in the field of medicinal chemistry, as antitumor agents
with selective antimetastatic properties and low systemic toxicity.
Ruthenium compounds appear to penetrate reasonably well the
tumor cells and bind effectively to DNA [13,14].
The well-developed synthetic chemistry of ruthenium, particu-
larly with imine ligands provides for many approaches to innova-
tive new metallopharmaceuticals [15]. Advantages of utilizing
ruthenium imine complexes in drug development include; (i) reli-
able preparations of stable complexes with predictable structures,
(ii) the ability to tune ligand affinities, electron transfer, substitu-
tion rates, and reduction potentials, and (iii) an increasing knowl-
edge of the biological effects of ruthenium complexes.
However little work was focused on binding ability and antiox-
idant activity of ruthenium(III) Schiff bases complexes and as a
result of the continuing quest for new complexes of ruthenium,
in this work, we are reporting the synthesis, spectral and electro-
chemical characterization of a series of new Ru(III) complexes con-
taining tetradentate Schiff base ligands derived from condensation
reactions of 4,5-dimethyl-1,2-phenylendiammine and 4,5-di-
chloro-1,2-phenylendiammine with salicylaldehyde and o-vanillin.
Furthermore, the interaction of calf thymus DNA (CT-DNA) with
the novel Ru(III) complexes was investigated by UV–Vis. Spectro-
photometry, fluorescence quenching and viscosity measurements.
Furthermore, the antioxidant activity of the complexes was deter-
mined by superoxide and hydroxyl radical scavenging method
in vitro. In addition, the antibacterial activity of the reported com-
pounds was studied and the results were compared with standard
antibiotics.
complexes was estimated by using 1-nitroso-2-naphthol reagent
[16] by adopting spectrophotometric extraction technique [17].
The chloride content of the complexes was determined by photo-
metric method [18]. The FT-IR spectra of the samples in the
4000–400 cmꢂ1 region were obtained in KBr discs on a Unicam-
Mattson 1000 FT-IR. The molar conductivities of the complexes
(1 ꢃ 10ꢂ3 M) in dimethylformamide (DMF) solution were mea-
sured at room temperature by using Jenway 4010 conductivity me-
ter. Room temperature (298 K) magnetic susceptibilities were
measured using a Sherwood Scientific balance using Hg[Co(SCN)4]
as a calibrant. Diamagnetic corrections calculated from Pascal’s
constants [19] were used to obtain the molar paramagnetic sus-
ceptibilities. Electron spin resonance (ESR) measurements of solid
state Ru(III) complexes were recorded at room temperature
(298 K) and liquid nitrogen temperature (77 K) on Bruker EPR
spectrometer at 9.706 GHz (X-band), the microwave power was
(1.0 mW) with 4.0 G modulation amplitude, using 2,2-diph-
enylpyridylhydrazone (DPPH) as standard (g = 2.0037). Cyclic vol-
tammetric measurements were carried out using a Princeton EG
and GPARC model potentiostat using glassy carbon working elec-
trode and all the potentials were referred to Ag/AgCl. Thermogravi-
metric analyses (TGA) were carried out using a Shimadzu DT-50
thermal analyzer under nitrogen atmosphere with a heating rate
10 °C/min. The UV–Vis spectra were recorded on a Shimadzu UV
1800 spectrophotometer. Fluorescence spectra were recorded on
a Jenway 6270 fluorimeter at room temperature.
Syntheses
Microwave assisted solvent-free synthesis of the Schiff base ligands
(L1–4H2)
The ligands L1–4H2 were previously synthesized by Ref. [20].
However, microwave assisted solvent-free synthesis method is
used to enhance the yield and reduce the time. 0.1 mol of the dia-
mine derivative and 0.2 mol of aldehyde were mixed well in a
50 ml Pyrex beaker and the mixture was irradiated in a microwave
oven for one minute. The yellow product obtained was separated,
dried and recrystallized from ethanol and the purity of the ligands
was checked by TLC.
Synthesis of Ru(III) complexes (1–4)
All Ru(III) complexes were synthesized according to the general
procedure: a stoichiometric amount of RuCl3ꢁ3H2O (10 mmol) in
ethanol was added to a hot ethanol solution of the desired ligand
(10 mmol) and the reaction mixture was boiled under reflux with
stirring for 3 h. On cooling the desired complex was obtained as
powder. In some cases, complete precipitation was achieved by
the addition of diethyl ether to the cold reaction mixture. The sol-
vent was evaporated on a vacuum line. The residue was washed
several times with hot petroleum ether (60–80 °C) and recrystal-
lized from benzene/ethanol to give reddish-brown crystals. The
products were finally dried in vacuo over P2O5. Synthetic route of
[Ru(L1–4)(H2O)2]Cl complexes (1–4) is shown in Scheme 1.
Experimental
Materials
DNA-binding studies
4,5-Dimethyl-1,2-phenylendiammine, 4,5-dichloro-1,2-pheny-
lendiammine, o-vanillin, salicylaldehyde and RuCl3ꢁ3H2O were
supplied from Aldrich. Calf thymus DNA (CT-DNA) and ethidium
bromide (EB) were purchased from Sigma Chemicals Co. All
solvents used were of analytical reagent grade and used without
further purification.
All experimental involving CT-DNA were performed in HCl/NaCl
(5:50 mM) buffer solution (pH = 7.24). Tris–HCl was prepared
using deionized and sonicated triple distilled water and kept at
4 °C for 3 days. The absorption ratio of CT-DNA solutions A260
/
A280 was 1.8:1.9, indicating that the CT-DNA was sufficiently free
from protein [21]. The CT-DNA concentration was determined via
absorption spectroscopy using the molar absorption coefficient of
6600 Mꢂ1 cmꢂ1 (260 nm) [22]. Stock solutions of metal complexes
were prepared by dissolving them in dimethylformamide (DMF)
and suitably diluting them with the corresponding buffer to the
required concentrations for all experiments. The extent of DMF
Instruments
Carbon, hydrogen and nitrogen were determined using Perkin
Elmer 2400 CHN elemental analyzer. Ruthenium content of the