← Back
Versatile Coordination Behavior of Salicylaldehydethiosemicarbazone in Ruthenium(II) Carbonyl Complexes: Synthesis, Spectral, X-ray, Electrochemistry, DNA Binding, Cytotoxicity, and Cellular Uptake Studies
Article
pubs.acs.org/Organometallics
Versatile Coordination Behavior of
Salicylaldehydethiosemicarbazone in Ruthenium(II) Carbonyl
Complexes: Synthesis, Spectral, X‑ray, Electrochemistry, DNA
Binding, Cytotoxicity, and Cellular Uptake Studies
P. Kalaivani,† R. Prabhakaran,*,† P. Poornima,‡ F. Dallemer,§ K. Vijayalakshmi,† V. Vijaya Padma,‡
and K. Natarajan*,†
†
Department of Chemistry and ‡Department of Biotechnology, Bharathiar University, Coimbatore-641 046, India
Laboratoire Chimie Provence-CNRS UMR6264, Université of Aix-Marseille I, II and III-CNRS, Campus Scientifique de
Saint-Jérôme, Avenue Escadrille Normandie-Niemen, F-13397 Marseille Cedex 20, France
§
S Supporting Information
*
ABSTRACT: The reaction of salicylaldehydethiosemicarbazone,
[H2-(Sal-tsc)], with an equimolar amount of [RuHCl(CO)(PPh3)3]
has afforded two complexes, namely [Ru(H-Sal-tsc)(CO)Cl(PPh3)2] (1) and [Ru(Sal-tsc)(CO)(PPh3)2] (2), in one pot. The
new complexes were separated and characterized by elemental
analyses, various spectroscopic techniques (NMR, UV−vis, IR), Xray crystallography, and cyclic voltammetry. In complex 1, the ligand
coordinated in a bidentate monobasic fashion by forming an unusual
strained NS four-membered ring in 32% yield. However, in 2, the
ligand coordinated in a tridentate dibasic fashion by forming ONS
five- and six-membered rings in 51% yield. Comparative biological
studies such as DNA binding, cytotoxicity (MTT, LDH, and NO),
and cellular uptake studies have been carried out for new
ruthenium(II) complexes (1 and 2). From the DNA binding studies, it is inferred that the complex 1 exhibited electrostatic
binding and 2 exhibited intercalative binding modes. On comparison of the cytotoxicity of the complexes in human lung cancer
cells (A549) and liver cancer cells (HepG2), complex 2 exhibited better activity than 1; this may be due to the strong chelation
and subsequent electron delocalization in 2 increasing the lipophilic character of the metal ion into cells.
■
arise from covalent binding of the drug to DNA.12 The efficacy
of cisplatin, however, is reduced by increasing tumor resistance
and high toxicity. These limitations have aroused interest
toward the design and evaluation of transition-metal complexes
other than platinum-based derivatives for therapeutic use. In
recent years, several ruthenium-based complexes have been
investigated for potential antitumor activity. Among the metal
atoms used in anticancer metal complexes, ruthenium is the
most unique. It is a rare noble metal unknown to living systems
and has strong complexation ability with numerous ligands. In
vitro and in vivo studies reveal that most ruthenium complexes
bind covalently to DNA via the N-7 atom of purines and cause
cytotoxicity by possibly inhibiting cellular DNA synthesis.
Moreover, ruthenium complexes have a stronger affinity for
cancer tissues than normal tissues. This is because ruthenium
binds readily to transferrin molecules in plasma and is
transported to the tumor cells. The first ruthenium compounds
to be studied for cancer activity were chloro ammine
INTRODUCTION
Thiosemicarbazones are versatile ligands and adopt various
binding modes with transition-metal and main-group-metal
ions.1 A number of reasons have been offered as responsible for
their versatility in coordination, such as intramolecular
hydrogen bonding, bulkier coligand, steric crowding on the
azomethine carbon atom, and π−π stacking interactions.2 In
addition, thiosemicarbazones and their metal complexes exhibit
a wide range of applications that extend from their use in
analytical chemistry through pharmacology to nuclear medicine.3−6 Metal complexes that reveal the capacity to bind with
nucleobases, DNA fragments, amino acids, peptides, and
proteins are currently receiving special attention, mainly due
to the clinical use of transition-metal complexes as antitumor
drugs.7−9 Platinum-based drugs have been in clinical use for
cancer treatment for more than 30 years.10 The landmark
discovery of the antitumoral properties of cisplatin by
Rosenberg in 1965 heralded a new area of anticancer research
based on metallopharmaceuticals.11 Although the mechanism
by which cisplatin selectively kills cells is not entirely
understood, it is generally believed that the therapeutic effects
© 2012 American Chemical Society
Received: September 28, 2012
Published: November 28, 2012
8323
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
Vario EL III CHNS instrument at the Department of Chemistry,
Bharathiar University, Coimbatore, India. The electronic spectra of the
complexes have been recorded in dichloromethane using a Systronics
119 spectrophotometer in the 800−200 nm range. 1H NMR, 13C
NMR, and 31P NMR spectra were taken in DMSO at room
temperature with a Bruker 400 MHz instrument with chemical shifts
relative to tetramethylsilane (1H, 13C) and orthophosphoric acid (31P).
Cyclic voltammograms were recorded on a CH instrument by using a
platinum-wire working electrode and platinum-disk counter electrode,
with 0.1 M tetrabutylammonium perchlorate as supporting electrolyte
at a scan rate of 100 mV s−1 in dichloromethane. All potentials were
referenced to the standard Ag/AgCl electrode, and ferrocene was used
as an external standard. Melting points were recorded by using a Lab
India melting point apparatus.
X-ray Crystallography. Single crystals of [Ru(H-Sal-tsc)(CO)Cl(PPh3)2] (1) and [Ru(Sal-tsc)(CO)(PPh3)2] (2) were obtained from
a CH2Cl2/CH3CN mixture and DMF, respectively. The data sets for
the single-crystal X-ray studies for the new complexes were collected
with Mo Kα (λ = 0.71073 Å) radiation on a Bruker SMART 1000
CCD diffractometer.32 All calculations were performed using the
SHELXS-97 and SHELXL-97 programs.33,34
DNA Binding Study. CT DNA solutions of various concentrations
(0.05−0.5 μM) dissolved in a phosphate buffer (pH 7) were added to
the new ruthenium(II) complexes (10 μM dissolved in a DMSO/H2O
mixture). Absorption spectra were recorded after equilibrium at 20 °C
for 10 min. The intrinsic binding constant Kb was determined by using
the Stern−Volmer equation (1).35,36
complexes: Durig et al. had observed in 1976 that the
ruthenium(III) complex fac-[Ru(NH3)3Cl3] induces filamentous growth of E. coli cells, at the same concentration as the
required concentration of cisplatin for the same effect.13 In
1980, this complex as well as the related ruthenium(II) complex
cis-[Ru(NH3)4Cl2] were evaluated for their anticancer properties by Clarke.14 However, although active, these compounds
were not soluble enough for pharmaceutical use.15 In the
following years, a large number of complexes were studied for
their cytotoxic properties.16−25 After extensive preclinical tests,
the compounds [indH]trans-[Ru(N-ind)2Cl4] (KP1019) and
[imiH]trans-[Ru(N-imi)(S-dmso)Cl4] (NAMI-A) entered into
clinical trials.10 Ruthenium has therefore been considered to be
an attractive alternative to platinum, in particular since many
ruthenium compounds are not very toxic and some ruthenium
compounds have been shown to be quite selective for cancer
cells.26,27 Along this line, herein we report the one-pot synthesis
of new ruthenium(II) complexes containing salicylaldehyde
thiosemicarbazone and their interactions with CT-DNA and
anticancer activities (MTT, LDH and NO release) and a
cellular uptake study against human lung (A549) and liver
cancer cells (HepG2).
■
EXPERIMENTAL SECTION
Materials. The ligand [H2-(Sal-tsc)] and the ruthenium metallic
precursor [RuHCl(CO)(PPh3)3] were synthesized according to the
standard literature procedures.28−30 All the reagents used in this study
were analar grade, and the solvents were purified and dried according
to standard procedures.31
Preparation of New Ruthenium(II) Complexes. A solution of
[H2-(Sal-tsc)] (0.021 g; 0.105 mmol) in 10 cm3 of benzene was added
dropwise to a boiling solution of [RuHCl(CO)(PPh3)3] (0.100 g,
0.105 mmol) in benzene (10 cm3). The mixture was heated at reflux
for 5 h and was subjected to thin-layer chromatography. Two spots
were identified and isolated by silica gel column chromatography by
using 9/1 petroleum ether (60−80 °C)/ethyl acetate and 9/1
benzene/methanol mixtures as eluents, respectively, for complexes 1
and 2.
[Ru(H-Sal-tsc)(CO)Cl(PPh3)2] (1). Anal. Calcd for
C45H38ClN3O2P2RuS: C, 61.19; H, 4.34; N, 4.76; S, 3.63. Found: C,
61.13; H, 4.31; N, 4.68; S, 3.57. Yield: 32%. Mp: 182 °C. FT-IR
(cm−1) in KBr: 3447 (νO−H), 1595 (νCN), 1285 (νC−O), 747 (νC−S),
1923 (νC≡O) 1427, 1085, 696 cm−1 (for PPh3). UV−vis (CH2Cl2),
λmax: 251 (15892), 268 (14789), 284 (9604) nm (dm3 mol−1 cm−1)
(intraligand transition); 323 (8380), 371 (4961), 384 (2282), nm
(dm3 mol−1cm−1) (LMCT s → d). 1H NMR (DMSO-d6, ppm): 9.9 (s,
1H, −OH), 8.22 (s, 1H, −CHN), 4.41 (s, −NH2), 6.64−7.73 (m,
aromatic). 13C NMR (DMSO-d6, ppm): 165.9 (C−S), 163.48 (C
N), 154.2 (aromatic PPh3), 149.6 (aromatic PPh3), 131.85 (aromatic
PPh3), 129.93 (2C, aromatic), 126.8 (aromatic), 123.8 (aromatic). 31P
NMR (DMSO-d6, ppm): 36.03.
[Ru(Sal-tsc)(CO)(PPh3)2] (2). Anal. Calcd for C45H37N3SO2P2Ru: C,
63.82; H, 4.40; N, 4.96; S, 3.79. Found: C, 63.78; H, 4.37; N, 4.92; S,
3.71. Yield: 51%. Mp: 216 °C. FT-IR (cm−1) in KBr: 1575 (νCN),
1316 (νC−O), 745 (νC−S), 1949 (νC≡O) 1435, 1089, 694 cm−1 (for
PPh3). UV−vis (CH2Cl2), λmax: 248 (14,426), 269 (13862), 287
(10432) nm (dm3 mol−1cm−1) (intraligand transition); 322 (7478),
378 (6308) nm (dm3 mol−1 cm−1) (LMCT s → d); 432 (4102) nm
(dm3 mol−1 cm−1) forbidden (d → d) transition. 1H NMR (DMSO-d6,
ppm): 8.44 (d, (J = 8.2 Hz), −CHN), 6.68 (d, (J = 8.72 Hz),
−NH2), 7.2−7.63 (m, aromatic). 13C NMR (DMSO-d6, ppm): 166.4
(C−S), 164.24 (CN), 155.2 (aromatic PPh3), 150.2 (aromatic
PPh3), 131.98 (aromatic PPh3), 130.03 (2C, aromatic), 127.2
(aromatic), 124.2 (aromatic). 31P NMR (DMSO-d6, ppm): 42.11.
Measurements. Infrared spectra were measured as KBr pellets on
a Nicolet instrument between 400 and 4000 cm−1. Elemental analyses
of carbon, hydrogen, nitrogen, and sulfur were determined using a
[DNA]/[εa − εf ] = [DNA]/[εb − εf ] + 1/Kb[εb − εf ]
(1)
The absorption coefficients εa, εf, and εb correspond to Aobsd/
[DNA], the extinction coefficient for the free complex and the
extinction coefficient for the complex in the fully bound form,
respectively. The slope and the intercept of the linear fit of the plot of
[DNA]/[εa − εf] versus [DNA] give 1/[εa − εf] and 1/Kb[εb − εf],
respectively. The intrinsic binding constant Kb can be obtained from
the ratio of the slope to the intercept (Table 2).36 Emission
measurements were carried out by using a JASCO FP-6600
spectrofluorometer. Tris buffer was used as a blank to make
preliminary adjustments. The excitation wavelength was fixed, and
the emission range was adjusted before measurements. All measurements were made at 20 °C. For emission spectral titrations, the
complex concentration was maintained constant at 10 μM and the
concentration of DNA was varied from 0.05 to 0.5 μM. The emission
enhancement factors were measured by comparing the intensities at
the emission spectral maxima under similar conditions. In order to
know the mode of attachment of CT DNA, fluorescence quenching
experiments of EB-DNA were also carried out by adding 10 μL
portions of 10 μM ruthenium(II) complexes every time to the samples
containing 12 μM EB, 10 μM DNA, and Tris buffer (pH 7). Before the
measurements, the system was shaken and incubated at room
temperature for ∼5 min. The emission was recorded at 530−750
nm. On the basis of the classical Stern−Volmer equation, the
quenching constant has been analyzed by using eq 2, where I0 and I
represent the fluorescence intensities in the absence and presence of
the complexes, respectively, r is the concentration ratio of the complex
to DNA, and Ksq is a linear Stern−Volmer quenching constant.
I0/I = 1 + K sqr
(2)
MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
Bromide) Assay. The effects of complexes 1 and 2 and the ligand
on the viability of human lung cancer cells (A549) and liver cancer
cells (HepG2) were determined by the 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) assay.37 The cells were
seeded at a density of 10000 cells per well in 200 μL of RPMI 1640
medium and were allowed to attach overnight in a CO2 incubator, and
then the complexes dissolved in DMSO were added to the cells at final
concentrations of 1, 10, 25, and 50 μM in the cell culture media. After
8324
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
Scheme 1. Preparation of New Ruthenium(II) Complexes
48 h, the wells were treated with 20 μL of MTT (5 mg/mL PBS) and
incubated at 37 °C for 4 h. The purple formazan crystals that formed
were dissolved in 200 μL of DMSO and read at 570 nm in a micro
plate reader.
Release of Lactate Dehydrogenase (LDH). LDH activity was
determined by the linear region of a pyruvate standard graph using
regression analysis and expressed as percentage (%) leakage as
described previously.38 Briefly, in a set of tubes, 1 mL of buffered
substrate (lithium lactate) and 0.1 mL of the media or cell extract were
added and the tubes were incubated at 37 °C for 30 min. After 0.2 mL
of NAD solution was added, the incubation was continued for another
30 min. The reaction was then arrested by adding 0.1 mL of DNPH
reagent, and the tubes were incubated for a further 15 min at 37 °C.
After this, 0.1 mL of media or cell extract was added to blank tubes
after arresting the reaction with DNPH. A 3.5 mL portion of 0.4 N
sodium hydroxide was added to all the tubes. The intensity of the
color that developed was measured at 420 nm in a Shimadzu UV/
visible spectrophotometer. The amount of LDH released was
expressed as a percentage.
Nitric Oxide (NO) Assay. The amount of nitrite was determined
by the method of Stueher and Marletta.39 Nitrite reacts with Griess
reagent to give a colored complex measured at 540 nm. To 100 μL of
the medium was added 50 μL of Griess reagent I, and these were
mixed and allowed to react for 10 min. This was followed by 50 μL
addition of Griess reagent II, and the reaction mixture was mixed well
and incubated for another 10 min at room temperature. The pink
color that developed was measured at 540 nm in a microquant plate
reader (Biotek Instruments).
Cellular Uptake Study. Cellular uptakes of complexes 1 and 2 and
the ligand were quantified according to the literature method with a
slight modification.40 Briefly, the lung (A549) and liver (HepG2)
cancer cells were treated with the test compounds for a period of 2 h.
The medium was aspirated, and cells were washed three times with ice
cold PBS. Then the cells were lysed with PBS containing 1% Triton X100. Concentration of the complexes and the ligand in the cell lysates
was measured with a fluorescence spectrophotometer (Jasco FP 6600)
at their maximum excitation/emission wavelengths of 492/530, 426/
495 and 220/291 nm, respectively, for complexes 1, 2 and ligand. To
offset the background fluorescence from the cellular components,
separate standardization curves were prepared using cellular lysates
containing series of known concentrations of different complexes and
the intracellular concentrations were determined using the standard
curve.
complexes 1 and 2, this band has been observed at 1575 and
1595 cm−1, indicating the coordination of the azomethine
nitrogen atom to the ruthenium.41,42 A sharp band that
appeared for the ligand at 826 cm−1 corresponding to a νCS
vibration disappeared completely in both of the complexes, and
a new band appeared at 745 and 747 cm−1 corresponding to a
possible νC−S vibration, indicating coordination of a thiolate
sulfur atom after enolization followed by deprotonation.43,44 A
broad band corresponding to a νOH vibration that appeared at
3439 cm−1 in the ligand has been observed at 3447 cm−1 in
complex 1, indicating the nonparticipation of the phenolic
oxygen atom in coordination.45 However, this band completely
disappeared in complex 2, indicating the involvement of
phenolic oxygen in coordination. This was further supported by
an increase in the phenolic C−O stretching frequency,
appearing at 1316 cm−1. Sharp bands at 1923 and 1949 cm−1
account for the terminal carbonyl group in 1 and 2,
respectively. In addition, vibrations corresponding to the
presence of triphenylphosphine also appeared in the expected
region.46 The electronic spectra of the complexes have been
recorded in dichloromethane, and they displayed six bands in
the region around 248−432 nm. The bands appearing in the
region 248−287 nm have been assigned to intraligand
transitions,47 the bands around 322−384 nm have been
assigned to ligand to metal charge transfer transitions, and
the band 432 nm has been assigned to a forbidden (d → d)
transition. The 1H NMR spectra of [H2-(Sal-tsc)] and
complexes 1 and 2 showed a complex overlap of signals in
the δ 6.3−7.6 ppm range corresponding to aromatic protons of
the ligand and coordinated triphenylphosphine.42 A sharp
singlet appearing for complex 1 at δ 8.21 ppm corresponds to
azomethine protons of the ligand.43 For complex 2, a doublet
corresponding to the azomethine group was observed at δ 8.44
ppm, due to the coupling with the phosphorus atom of the
triphenylphosphine.48 In the spectra of the ligand the singlet
appearing at δ 9.52 ppm is assigned to the N(2)HCS group.49
However, in the spectra of the complexes there was no
resonance attributable to N(2)H, indicating the coordination of
ligand in the anionic form upon deprotonation at N(2). A sharp
singlet corresponding to the phenolic −OH group appeared at
δ 11.25 ppm in the free ligand. In the spectra of 1, the
appearance of a singlet at δ 9.9 ppm indicated the nonparticipation of the phenolic oxygen in coordination,50 and this
signal completely disappeared in 2, confirming the involvement
of the phenolic oxygen in coordination. Though the spectra of
1 showed a singlet at δ 4.41 ppm corresponding to NH2
protons, the same has been observed as a doublet at δ 6.68 ppm
for 2, which may be due to the restricted CN bond rotation
of the ligand.48 In the 13C{1H} NMR spectra of the complexes,
the thioamide (C−S) carbon resonates at 166 and 167.0 ppm.
The azomethine carbon resonance is observed at 164.0 and
163.48 ppm. In both complexes, aromatic carbon atoms of the
■
RESULTS AND DISCUSSION
Reaction of [H2-(Sal-tsc)] with an equimolar amount of
[RuHCl(CO)(PPh3)3] resulted in two different entities with
different structural features in a single reaction (Scheme 1).
The new complexes are soluble in common organic solvents
such as dichloromethane, chloroform, benzene, acetonitrile,
ethanol, methanol, dimethyl sulfoxide, and dimethylformamide.
The analytical data of the complexes agreed well with the
proposed molecular formulas.
Spectroscopic Studies. The IR spectrum of [H2-(Sal-tsc)]
shows a sharp band at 1611 cm−1 corresponding to the νCN
vibration of the azomethine group. In the IR spectra of the new
8325
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
phenoxy group observed around 129.4, 126.8, 123.8 ppm
(complex 1) and 130.02, 127.2, 124.2 ppm (complex 2) are
comparable to the literature values.51 The CO carbon
resonating at 208 ppm (1) and 207 ppm (2) is comparable with
earlier observations. 4 In both complexes three signals
corresponding to the presence of triphenylphosphine observed
at 154.2, 149.6, and 131.28 ppm (complex 1) and 155.4, 150.2,
and 131.98 ppm (complex 2) are in the range of the reported
values.52 In order to confirm the presence of triphenylphosphine, 31P NMR spectra were recorded. The singlet observed at
36.03 and 42.11 ppm in complexes 1 and 2, respectively,
suggested the presence of two magnetically equivalent
triphenylphosphines trans to each other (Figure S1 and S2,
Supporting Information).
X-ray Crystallography. Complexes 1 and 2 crystallized in
monoclinic space group C2/c. The crystallographic data of the
complexes are given in Table S1 (Supporting Information).
The molecular structures and hydrogen-bonding diagrams are
depicted in Figures 1−4. In complex 1, the ligand [H2-(Sal-
Figure 2. ORTEP diagram of 1 with hydrogen bonds leading to a
pseudo-hydroxo-bridged binuclear structure.
Figure 1. ORTEP diagram of [Ru(H-Sal-tsc)(CO)Cl(PPh3)2] (1)
(hydrogen atoms are omitted for clarity). Bond lengths (Å): Ru(1)−
N(1), 2.192(2); Ru(1)−P(1), 2.3815(7); Ru(1)−P(2), 2.3703(7);
Ru(1)−S(1), 2.4012(7); Ru(1)−Cl(1), 2.4247(7); Ru(1)−C(37),
1.845(3). Bond angles (deg): C(9)−Ru(1)−N(2), 166.4(2); C(9)−
Ru(1)−P(2), 90.1(1); N(2)−Ru(1)−P(2), 89.7(1); C(9)−Ru(1)−
P(1), 91.4(1); N(2)−Ru(1)−P(1), 89.9(1); P(2)−Ru(1)−P(1),
178.48(5); C(9)−Ru(1)−S(1), 101.3(1); N(2)−Ru(1)−S(1),
65.1(1); P(2)−Ru(1)−S(1), 90.68(4); P(1)−Ru(1)−S(1), 89.42(4);
C(9)−Ru(1)−Cl(1), 94.5(1); N(2)−Ru(1)−Cl(1), 99.2(1); P(2)−
Ru(1)−Cl(1), 89.10(5); P(1)−Ru(1)−Cl(1), 90.40(5); S(1)−
Ru(1)−Cl(1), 164.23(5).
Figure 3. ORTEP diagram of [Ru(Sal-tsc)(CO)(PPh3)2] (2)
(hydrogen atoms are omitted for clarity). Bond lengths (Å):
Ru(1)−N(2), 2.061(6); Ru(1)−P(1), 2.406(2); Ru(1)−P(2),
2.399(2); Ru(1)−S(1), 2.344(2); Ru(1)−O(1), 2.069(5); Ru(1)−
C(37), 1.893(8). Bond angles (deg): C(45)−Ru(1)−O(1), 95.0(3);
C(45)−Ru(1)−N(2), 174.0(3); O(1)−Ru(1)−N(2), 90.6(2);
C(45)−Ru(1)−S(1), 93.3(2); O(1)−Ru(1)−S(1), 171.8(1); N(2)−
Ru(1)−S(1), 81.2(2); C(45)−Ru(1)−P(1), 87.4(2); N(2)−Ru(1)−
P(1), 90.7(2); S(1)−Ru(1)−P(1), 94.29(7); C(45)−Ru(1)−P(2),
91.2(2); O(1)−Ru(1)−P(2), 92.0(1); N(2)−Ru(1)−P(2), 90.9(2);
S(1)−Ru(1)−P(2), 87.73(7); P(1)−Ru(1)−P(2), 177.58(7).
tsc)] is coordinated to ruthenium ion through the N(2)
nitrogen and thiolate sulfur atoms, forming a more strained
four-membered chelate ring with a bite angle N(2)−Ru(1)−
S(1) of 65.1(1)°. The Ru(1)−N(2) bond distance is 2.208(5)
Å, and the Ru(1)−S(1) distance is 2.413(1) Å. The other four
sites are occupied by phosphorus atoms of two triphenylphosphine ligands which are mutually trans to each other with
Ru(1)−P(1) and Ru(1)−P(2) distances of 2.384(1) and
2.380(1) Å and one chloride and a carbonyl group with
Ru(1)−Cl(1) and Ru(1)−C(9) distances of 2.427(1) and
1.835(5) Å, respectively. The observed bond distances are
comparable with those found in other reported ruthenium
complexes containing PPh3.53 The cis angles N(2)−Ru(1)−
P(2) = 89.7(1)°, N(2)−Ru(1)−P(1) = 88.9(1)°, P(1)−
Ru(1)−S(1) = 89.42(4)°, and P(2)−Ru(1)−Cl(1) =
89.10(5)° are acute, whereas the other cis angles C(9)−
Ru(1)−P(1) = 91.4(1)°, C(9)−Ru(1)−P(2) = 90.1(1)°,
C(9)−Ru(1)−S(1) = 101.3(1)°, P(2)−Ru(1)−S(1) =
90.68(4)°, C(9)−Ru(1)−Cl(1) = 94.5(1)°, N(2)−Ru(1)−
Cl(1) = 99.2(1)°, and P(1)−Ru(1)−Cl(1) = 90.40(5)° are
obtuse. The trans angles C(9)−Ru(1)−N(2) = 166.4(2)°,
P(2)−Ru−P(1) = 178.48(5)°, and S(1)−Ru(1)−Cl(1) =
164.23(5)° deviate from linearity. The variations in bond
lengths and angles lead to a significant distortion from an ideal
octahedral geometry for the complex.
In addition, complex 1 contains one intramolecular hydrogen
bond through the hydrogen atom of the hydroxy group with
the hydrazinic nitrogen (N3) of thiosemicarbazone moiety with
an O1(A)−H(A)···N3(A) distance of 2.680 Å and one
intermolecular hydrogen bond through the hydrogen atom of
the amine nitrogen (N1) of one molecule with the oxygen
8326
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
octahedral geometry of the complex. In this complex, one of
the hydrogen atoms of each amino group of three independent
molecules (N3, N6, and N9) are engaged in intermolecular
hydrogen bonding with the hydrazinic nitrogen atoms (N1, N7,
and N4) of a second set of three molecules: N3(A)−
H(A)···N1(B) and N3(B)−H(B)···N1(A), N6(A)−H(A)···N7(B) and N9(B)−H(B)···N4(A), N9(A)−H(A)···N4(B) and N6(B)−H(B)···N7(A) (Table 1). This intermolecular
hydrogen bonding creates a pseudo-binuclear structure (Figure
4).
Salicylaldehyde thiosemicarbazone exists in thiol and thione
forms as indicated by II and III, and they can form stable fiveand six-membered rings as in VI and VII. However, the
coordination behavior of salicylaldehyde thiosemicarbazone was
reported by Bhattacharya et al., who stated that the formation
of a five-membered ring is an impossible mode of binding and a
four-membered ring is possible.50,53 They supported the
formation of a four-membered ring as in VIII by offering a
couple of reasons: intramolecular hydrogen bonding and a
bulky coligand or bulky substitution on the azomethine carbon
atom. In contrast to this, we observed both five- and fourmembered-ring products from a single reaction between
salicylaldehyde thiosemicarbazone and [RuHCl(CO)(PPh3)3]
with different yield and coordination behavior. NS coordination
of the ligand with the formation of an unusual four-membered
ring (VIII) was found in complex 1, and ONS coordination
(VI) with five- and six-membered-ring formation was found in
complex 2. According to the report of Bhattacharya et al., if the
intramolecular hydrogen bonding and the bulkiness were the
only responsible factors, we could have obtained only a product
of type VIII.50,53 However, we obtained a complex of type VI in
higher yield as compared with type VIII. From this, it is
concluded that the aforementioned factors are not the only
responsible factors in determining the coordination behavior of
thiosemicarbazones and there may be some other factors and/
or their collective influence in directing them.
Figure 4. ORTEP diagram of 2 with hydrogen bonds leading to a
pseudo-binuclear structure.
atom of the hydroxyl group (O1) of another molecule with an
N1(A)−H(A)···O1(B) distance of 3.054 Å, leading to the
formation of a pseudo-hydroxo-bridged binuclear complex
(Table 1, Figure 2).
Table 1. Hydrogen Bonds for Complexes 1 and 2 (Å and
deg)
D−H···A
O1(A)−H1(A)···N3(A)
N1(A)−H1(A)···O1(B)
N1(B)−H1(B)···O1(A)
N3(A)−H(A)···N1(B)
N3(B)−H(B)···N1(A)
N9(A)−H(A)···N4(B)
N6(B)−H(B)···N7(A)
N6(A)−H(A)···N7(B)
N9(B)−H(B)···N4(A)
d(D−H)
d(H···A)
Complex 1a
0.850
1.95(2)
0.860
2.27(4)
0.860
2.27(4)
Complex 2b
0.861
2.16(2)
0.861
2.16(2)
0.859
2.17(9)
0.860
2.15(1)
0.860
2.15(1)
0.859
2.17(9)
d(D···A)
∠(DHA)
2.680
3.05(4)
3.05(4)
143.16
150.81
150.81
2.98(8)
2.98(8)
2.99(3)
2.99(5)
2.99(5)
2.99(3)
160.84
160.84
158.18
167.20
167.20
158.18
Symmetry operation: (x, y, z); (1 − x, y, 1.5 − z). bSymmetry
operation: (x, y, z); (1 − x, y, 1/2 − z).
a
Suitable crystals of complex 2 were obtained from
dimethylformamide. Complex 2 crystallized in the monoclinic
crystal system with three independent molecules in the unit
cell. In this complex, the thiosemicarbazone ligand coordinated
to ruthenium in an ONS fashion by utilizing its phenolic
oxygen, N1 hydrazinic nitrogen, and thiolate sulfur atoms with
the formation of one six-membered ring and another fivemembered ring with a bite angle N(2)−Ru(1)−S(1) of
81.2(2)°.
However, on comparison with complex 1, the angular
distortion observed in 2 is much less, which may be due to
strong ONS chelation of the ligand. The Ru(1)−N(2) bond
distance is 2.061(6) Å, the Ru(1)−O(1) bond distance is
2.069(5) Å, and the Ru(1)−S(1) distance is 2.344(2) Å. The
other three sites are occupied by phosphorus donor atoms of
two triphenylphosphines with Ru(1)−P(1) and Ru(1)−P(2)
distances of 2.406(2) and 2.399(2) Å, respectively, and one
carbonyl group with a Ru(1)−C(45) distance of 1.893(8) Å.
These values are comparable with those found for complexes
reported earlier.54 The trans angles C(45)−Ru(1)−N(2) =
174.0(3)°, O(1)−Ru(1)−S(1) = 171.8(1)°, and P(1)−Ru−
P(2) = 177.58(7)° indicate a significant distortion in the
Electrochemistry. The complexes 1 and 2 were electroactive in the sweep range ±2.00 V. The cyclic voltammogram of
1 showed a reversible one-electron-oxidation response E1/2(oxi)
at 0.725 V with a peak to peak separation of 50 mV and quasireversible reduction at −0.480 V with a peak to peak separation
of 200 mV (Table S2, Supporting Information). These quasireversible reduction peaks can be assigned to a Ru(II)/Ru(I)
process. In addition, complex 1 showed both quasi-reversible
ligand oxidation and reduction with E1/2 at 1.125 and −1.385 V
and peak to peak separations of 250 and 130 mV, respectively
(Figures S3 and S4, Supporting Information). Complex 2
exhibited quasi-reversible oxidation at 0.549 V with a peak to
peak separation of 332 mV and reversible reduction
corresponding to Ru(II)/Ru(I) at −0.460 V with a peak to
peak separation of 80 mV. The reason for the quasi-reversible
electron transfer process may be due to slow electron transfer
or the adsorption of the complex onto the electrode surface.55
8327
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
at 404 nm showed hypochromism (A = 0.0501−0.0426)
without a wavelength shift in the absorption maxima. As shown
in Figure 5, the addition of CT-DNA to complex 2 also led to
isosbestic spectral changes with an isosbestic point at 304 nm.
The observed hyperchromic effect with blue shift in the
intraligand band suggested that the new ruthenium(II)
complexes bind to CT-DNA by external contact, possibly due
to electrostatic binding.56
The intrinsic binding constant Kb is a useful tool to
determine the magnitude of the binding strength of compounds
with CT-DNA. It can be determined by monitoring the
changes in the absorbance in the IL band at the corresponding
λmax value with increasing concentration of DNA and is given
by the ratio of slope to the y intercept in plots of [DNA]/(εa −
εf) versus [DNA] (insets in Figure 5). From the binding
constant values (Table 2), it is inferred that both the complexes
bind with CT-DNA more efficiently.
From current electrochemical investigations, it is concluded
that the ligand [H2-(Sal-tsc)] can stabilize the lower and higher
oxidation states of the metal.
DNA Binding Studies. UV absorption titration experiments were carried out to study the DNA binding properties of
the new Ru(II) complexes 1 and 2. The absorption spectra of
the new complexes at constant concentration (10 μM) in the
presence of different concentrations of CT-DNA (0.05−0.50
μM) are given in Figure 5. The absorption spectra of complex 1
Table 2. Binding Constant for Interaction of Complexes
with CT-DNA
system
Kb (105 M−1)
CT-DNA + 1
CT-DNA + 2
5.5977
5.5815
Emission spectral studies were carried out to know more on
the binding nature of metal complexes to DNA. Complex 1 had
a fluorescence emission at 412 nm (Figure 6), while addition of
CT DNA to the complex solution resulted in hyperchromism
with an increase in intensity (I = 45.21−79.38) without any
shift in the absorption maxima. However, in complex 2,
hypochromism was observed with a decrease in intensity at 469
nm (41.73−33.73). The enhanced fluorescence intensity
observed for complex 1 symbolizes electrostatic binding of
DNA. However, the marked decrease in the fluorescence
intensity of complex 2 indicates the intercalative binding mode
of DNA.
The results obtained from the above experiments suggested
that both the compounds can bind with CT-DNA. However,
the exact mode of binding cannot be proposed by these studies.
Hence, ethidium bromide displacement studies were carried
out. Ethidium bromide competitive binding experiments using
new ruthenium(II) complexes 1 and 2 as quenchers may give
further information about the binding of these complexes to
DNA. EB emits intense fluorescence light in the presence of
DNA, due to its strong intercalation between adjacent DNA
base pairs. The quenching extent of fluorescence of EB bound
to DNA is used to determine the extent of binding of metal
complexes to DNA. When complexes 1 and 2 were added to
DNA pretreated with EB, the DNA induced emission intensity
at 602 nm was decreased (Figure 7). This indicated that the
complexes could replace EB from the DNA-EB system. The
Stern−Volmer quenching constants Ksv, obtained as a slope
from the plot of I0/I vs [Q] (Figure 7, inset), were found to be
1.65 × 103 and 8.72 × 103 M−1, respectively, for complexes 1
and 2. The results obtained suggested that the complex 2 has
relatively high magnitude of binding than complex 1.
Further, the apparent DNA binding constants (Kapp) were
calculated using eq 3, where [complex] is the value at 50%
Figure 5. Absorption titration spectra of 1 and 2 with increasing
concentrations (0.05−0.5 μM) of CT-DNA (phosphate buffer, pH 7).
The inset shows binding isotherms with CT-DNA.
mainly consist of two resolved bands (intraligand (IL) and CT
transitions) centered at 262 nm (IL) and 360 nm (CT). As the
DNA concentration is increased, a hyperchromism (A =
0.2598−1.0963) with a blue shift of 2 nm was observed in the
intraligand band. The CT band at 360 nm showed
hypochromism (A = 0.2525−0.1571) with a 5 nm blue shift
in the absorption maxima. In addition, the binding of complex
1 to CT DNA led to isosbestic spectral changes with an
isosbestic point at 338 nm. For complex 2, upon addition of
DNA, the intraligand band at 267 nm exhibited hyperchromism
(A = 0.1264−1.0167) with a blue shift of 7 nm. The CT band
KEB[EB] = K app[complex]
8328
(3)
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
Figure 6. Changes in the emission spectra of 1 and 2 with increasing concentrations (0.05−0.5 μM) of CT-DNA (phosphate buffer, pH 7).
reduction in the fluorescence intensity of EB, KEB (1.0 × 107
M−1) is the DNA binding constant of EB, and [EB] is the
concentration of EB (12 μM). Kapp values were 1.98 × 105 and
10.46 × 105 M−1 for complexes 1 and 2, respectively. From
these experimental data, it is seen that the ruthenium(II)
complex 2 replaces EB more effectively than 1, which is in
agreement with the results observed from the electronic
absorption spectra. Since these changes indicate only one
kind of quenching process, it may be concluded that both
complexes can bind to DNA via the intercalation mode.
Furthermore, the observed quenching constants and binding
constants of the new complexes 1 and 2 suggest that the
interaction of both complexes with DNA should be
intercalative.57
MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium Bromide) Assay. The cytotoxic effect of the new
ruthenium(II) complexes 1 and 2 and the ligand on the
proliferation of human lung cancer and liver cancer cell lines
A549 and HepG2 were assayed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay). From the
results, it is found that complex 2 exhibited higher
antiproliferative activity than 1 (Figure S5, Supporting
Information). The IC50 values were found to be 26 ± 1.03
and 23 ± 0.99 for A549 and 23 ± 1.01 and 20 ± 1.21 for
HepG2 cells, respectively, for complexes 1 and 2. In
comparison with the conventional standard cisplatin, complex
2 was found to have a lower IC50 value for the A549 cell line.
However, for HepG2, both complexes have IC50 values higher
than that for cisplatin. In order to assess whether the complexes
exhibited their cytotoxic activity through the induction of
oxidative stress, we analyzed the cell viability in the presence of
an antioxidant, glutathione (Figure 8). The results of the
present study indicated that the cell viability was significantly
increased in the presence of glutathione in the cells treated with
the complexes. This in turn supports the notion that the newly
synthesized complexes exhibited their cytotoxic activities
through ROS generation and thereby oxidative stress.
Lactate Dehydrogenase Release. When cancer cell lines
A549 and HepG2 were treated with the new ruthenium(II)
complexes treated for a period of 48 h, a significant increase of
LDH release in the culture medium was observed (Figure 9).
This indicates the efficiency of new complexes in inducing cell
death by collapsing the membrane integrity. LDH is a stable
cytoplasmic enzyme that is released into the culture medium
following loss of membrane integrity and serves as a general
means to assess cytotoxicity resulting from chemical compounds or environmental toxic factors. The significant increase
of LDH level in the culture supernatant confirmed the cytotoxic
effect of the newly synthesized complexes on lung and liver
cancer cell lines. This study has highlighted how the complexes
enhance cell death via a mechanism which is dependent on the
induction of oxidative stress. Oxidative stress damages the
membrane lipids, thereby leading to the destruction of
membrane integrity. This in turn may lead to the LDH release
from the cells treated with the complexes. The induction of
LDH release was found to be higher for complex 2 than for 1.
These results are comparable with our earlier reports and those
of Alia et al., concerning the significant release of LDH leakage
into the culture medium, which confirms the cytotoxic effect
induced by the complexes.48,58−60
Nitric Oxide Assay. The nitric oxide (NO) assay is also an
important measure of cytotoxicity, as NO has been shown to
directly inhibit methionine adenosyl transferase, leading to
glutathione depletion, and its reaction with superoxide
generates the strong oxidant peroxynitrite, which can initiate
lipid peroxidation or cause a direct inhibition of the
mitochondrial respiratory chain.61 In the present study NO
release by the new ruthenium(II) complexes was evaluated
using A549 and HepG2 cells. The quantification of the nitrite
produced in the cell media by the Griess assay is an indirect but
cost-effective measurement of the amount of NO produced by
the cells. It is interesting to note that both complexes were
found to release more NO than the control, and 2 was found to
be the more effective complex (Figure 10). The results of the
nitric oxide assay support the concept that the complex-induced
cell death is mediated by reactive oxygen species generation.
Cellular Uptake Study. The intracellular uptake of a
specific drug plays a vital role in ameliorating several diseases.
Since the IC50 values are critical when comparing normal cells
in the human body, the present study was focused on the
8329
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
Figure 9. Percentage of lactate dehydrogenase released by the human
cancer cell lines A549 and HepG2 after an incubation period of 48 h
with complexes 1 and 2. Error bars represent the standard mean error
(n = 6).
Figure 10. Nitrite released (in nmol) by the human cancer cell lines
A549 and HepG2 after an incubation period of 48 h with complexes 1
and 2. Error bars represent the standard mean error (n = 6).
respectively, for A549 and HepG2 cell lines (Figure 11). It is
obvious from the results that, even at low concentrations of the
Figure 7. Emission spectra of EB bound to DNA in the presence of
complexes 1 and 2 in Tris-HCl buffer (pH 7). Arrows indicate the
intensity changes upon increasing concentration of the complexes.
Inset: fluorescence quenching curve of DNA-bound EB with the
complexes.
Figure 11. Percentage of intracellular uptake of complexes 1 and 2 by
human cancer cell lines A549 and HepG2 after an incubation period of
2 h. Error bars represent the standard mean error (n = 6).
Figure 8. Cell viability assay of the complexes in presence of
glutathione.
complexes, they are cytotoxic to the lung and liver cancer cells
when they are completely absorbed by the cell; in particular,
the uptake level of complex 1 by both cell lines is higher than
that of 2. The uptake levels were dependent on the dose of each
complex used. This also indicates that their cytotoxicities as
determined by the MTT assay were not disproportionately
influenced by the complexes having different cellular uptake
concentrations which showed 50% inhibition for A549 and
HepG2 cell lines. The intracellular concentrations of complexes
1 and 2 were determined as described in the Experimental
Section. The intracellular concentrations of complexes 1, 2 and
ligand after an incubation period of 4 h were found to be 61.53
and 69.56%, 54.34 and 62.5% and 27.35 and 31.25%,
8330
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
■
levels. Results in their percentage uptake are shown as mean ±
SD (n = 9), in three separate experiments performed in
triplicate.
CONCLUSIONS
In an effort to determine the coordination behavior of
thiosemicarbazones, the reaction of salicylaldehydethiosemicarbazone [H2-(Sal-tsc)] with [RuHCl(CO)(PPh3)3] has been
carried out and discussed in this article. The stoichiometric
reaction afforded two different complexes having different
structural features, and they were characterized by various
spectral, analytical, X-ray crystallographic, and cyclic voltammetric methods. It is interesting to note that one of the
complexes obtained (1) has thiosemicarbazone coordinated as
NS through the N(2) nitrogen and thiolate sulfur by forming
an unusual four-membered ring, whereas in complex 2, the
same ligand coordinated as an ONS dibasic tridentate donor.
Since these two new ruthenium complexes containing a
biologically active thiosemicarbazone moiety have different
coordination modes, an attempt was made to compare their
modes of chelation with their potential biological activities. For
that purpose, they were subjected to CT-DNA binding,
cytotoxicity (MTT, lactate dehydrogenase (LDH), and NO
release in human carcinoma cell lines A549 and HepG2), and
cellular uptake studies. The results showed that complex 2 has
an activity higher than that of 1, and this may be due to the
strong chelation and increased electron delocalization in 2
increasing the lipophilic character of the metal ion into the cells
in comparison to 1. Further studies remain to determine the
coordination behavior of thiosemicarbazones and exact
molecular mechanism of cytotoxicity.
ASSOCIATED CONTENT
S Supporting Information
*
CIF files, tables, and figures giving crystallographic data for
[Ru(H-Sal-tsc)(CO)Cl(PPh3)2] (1) and [Ru(Sal-tsc)(CO)(PPh3)2] (2), 31P NMR spectra and cyclic voltammograms of
1 and 2, and results of a cell proliferation (MTT) assay. This
material is available free of charge via the Internet at http://
pubs.acs.org. Crystallographic data for 1 and 2 have also been
deposited at the Cambridge Crystallographic Data Centre as
supplementary publications (CCDC Nos. 857206 and 857205).
The data can be obtained free of charge at www.ccdc.cam.ac.
uk/conts/retrieving.html or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ,
U.K. (fax, +44-1223/336-033; e-mail, deposit@ccdc.cam.ac.uk).
■
REFERENCES
(1) (a) Campbell, M. J. M. Coord. Chem. Rev. 1975, 15, 297.
(b) Casas, J. S.; Garcia-Tasende, M. S.; Sordo, J. Coord. Chem. Rev.
2000, 209, 197.
(2) (a) Basuli, F.; Peng, S. M.; Bhattacharya, S. Inorg. Chem. 1997, 36,
5645. (b) Basuli, F.; Ruf, M.; Pierpont, C. G.; Bhattacharya, S. Inorg.
Chem. 1998, 37, 6113. (c) Basuli, F.; Ruf, M.; Pierpont, C. G.;
Bhattacharya, S. Inorg. Chem. 2000, 39, 1120. (d) Pal, I.; Basuli, F.;
Mak, T. C. W.; Bhattacharya, S. Angew. Chem., Int. Ed. Engl. 2001, 113,
3007. (e) Archaryya, R.; Basuli, F.; Peng, S. M.; Lee, G. H.; Falvello, L.
R.; Bhattacharya, S. Inorg. Chem. 2006, 45, 1252. (f) Ze-hua, L.;
Chung-Ying, D.; Ji-hui, L.; Young-jiang, L.; Yu-hua, M.; X-Zeng, Y.
New J. Chem. 2000, 24, 1057. (g) Prabhakaran, R.; Kalaivani, P.;
Renukadevi, S. V.; Huang, R.; Senthilkumar, K.; Karvembu, R.;
Natarajan, K. Inorg. Chem. 2012, 51, 3525.
(3) West, D. X.; Swearingen, J. K.; Valdes-Martinez, J.; HernandezOrtega, S.; El-Sawaf, A. K.; van Meurs, F.; Castineiras, A.; Garcia, I.;
Bermejo, E. Polyhedron 1999, 18, 2919.
(4) Tarasconi, P.; Capacchi, S.; Pelosi, G.; Cornia, M.; Albertini, R.;
Bonati, A.; Dall’Aglio, P.; Lunghi, P.; Pinelli, S. Bioorg. Med. Chem.
2000, 8, 157.
(5) Ghazy, S. E.; Kabil, M. A.; El-Asmy, A. A.; Sherief, Y. A. Anal. Lett.
1996, 29, 1215.
(6) Dilworth, J. R.; Cowley, A. H.; Donnelly, P. S.; Gee, A. D.;
Heslop, J. M. Dalton Trans. 2004, 2404.
(7) Bruijnincx, P. C. A.; Sadler, P. J. Adv. Inorg. Chem. 2009, 61, 1.
(8) Dyson, P. J.; Sava, G. Dalton Trans. 2006, 1929.
(9) Allardyce, C. S.; Dorcier, A.; Scolaro, C.; Dyson, P. J. Appl.
Organomet. Chem. 2005, 19, 1.
(10) Bratsos, I.; Jedner, S.; Gianferrara, T.; Alessio, E. Chimia 2007,
61, 692.
(11) Rosenberg, B.; Vancamp, L.; Trosko, J. E.; Mansour, V. H.
Nature 1969, 222, 385.
(12) Brown, S. J.; Chow, C. S.; Lippard, S. J. In Encyclopedia of
Inorganic Chemistry; King, R. B., Ed.; Wiley: West Sussex, England,
1994; p 3305.
(13) Durig, J. R.; Danneman, J.; Behnke, W. D.; Mercer, E. E. Chem.Biol. Interact. 1976, 13, 287.
(14) Clarke, M. J. Met. Ions Biol. Syst. 1980, 11, 231.
(15) Clarke, M. J.; Zhu, F.; Frasca, D. R. Chem. Rev. 1999, 99, 2511.
(16) Novakova, O.; Kasparkova, J.; Vrana, O.; Vanvliet, P. M.;
Reedijk, J.; Brabec, V. Biochemistry 1995, 34, 12369.
(17) Vanvliet, P. M.; Toekimin, S. M. S.; Haasnoot, J. G.; Reedijk, J.;
Novakova, O.; Vrana, O.; Brabec, V. Inorg. Chim. Acta 1995, 231, 57.
(18) Vilaplana, R.; Romero, M.; Quiros, M.; Salas, J.; GonzalezVilches, F. Met.-Based Drugs 1995, 2, 211.
(19) Chatterjee, D.; Mitra, A.; De, G. S. Platinum Met. Rev. 2006, 50,
2.
(20) Alessio, E.; Mestroni, G.; Nardin, G.; Attia, W. M.; Calligaris,
M.; Sava, G.; G. Zorzet, G. Inorg. Chem. 1988, 27, 4099.
(21) Sava, G.; Pacor, S.; Zorzet, S.; Alessio, E.; Mestroni, G.
Pharmacol. Res. 1989, 21, 617.
(22) Coluccia, M.; Sava, G.; Loseto, F.; Nassi, A.; Bocarelli, A.;
Giordano, D.; Alessio, E.; Mestroni, G. Eur. J. Cancer 1993, 29, 1873.
(23) Velders, A. H.; Kooijman, H.; Spek, A. L.; Haasnoot, J. G.; de
Vos, D.; Reedijk, J. Inorg. Chem. 2000, 39, 2966.
(24) Hotze, A. C. G.; Caspers, S. E.; de Vos, D.; Kooijman, H.; Spek,
A. L.; Flamigni, A.; Bacac, M.; Sava, G.; Haasnoot, J. G.; Reedijk, J. J.
Biol. Inorg. Chem. 2004, 9, 354.
(25) Kelman, A. D.; Clarke, M. J.; Edmonds, S. D.; Peresie, H. J. J.
Clin. Hematol. Oncol. 1977, 7, 274.
(26) Allardayce, C. S.; Dyson, P. J. Platinum Met. Rev. 2001, 45, 62.
(27) Kostova, J. Curr. Med. Chem. 2006, 13, 1085.
(28) Purohit, S.; Kolay, A. P.; Prasad, L. S.; Manoharan, P. T.; Ghosh,
S. Inorg. Chem. 1989, 28, 3735.
(29) Klayman, D. L.; Brtosevich, J. F.; Griffin, T. S.; Mason, C. J.;
Scovill, J. P. J. Med. Chem. 1997, 22, 855.
(30) Ahmed, N.; Levision, J. J.; Robinson, S. D.; Uttley, M. F. Inorg.
Synth. 1974, 15, 48.
■
■
Article
AUTHOR INFORMATION
Corresponding Author
*Tel: +91-422-2428319. Fax: +91-422-2422387. E-mail:
rpnchemist@gmail.com (R.P.); k_natraj6@yahoo.com (K.N.).
Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS
We gratefully acknowledge the Council of Science and
Industrial Research, New Delhi, India, and Department of
Science and Technology, New Delhi, India, for financial
assistance.
8331
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332
Organometallics
Article
(31) Vogel, A. I. Textbook of Practical Organic Chemistry, 5th ed.;
Longman: London, 1989; p 268.
(32) (a) Blessing, R. H. Acta Crystallogr. 1995, A51, 33. (b) Blessing,
R. H. Cryst. Rev. 1987, 1, 3. (c) Blessing, R. H. J. Appl. Crystallogr.
1989, 22, 396.
(33) Sheldrick, G. M. SHELXTL Version 5.1, An Integrated System for
Solving, Refining and Displaying Crystal Structures from Diffraction Data;
Siemens Analytical X-ray Instruments, Madison, WI, 1990.
(34) Sheldrick, G. M. SHELXL-97, A Program for Crystal Structure
Refinement, Release 97-2; Institut fur Anorganische Chemie der
Universitat Gottingen, Tammanstrasse 4, D-3400 Gottingen, Germany, 1998.
(35) Wolfe, A.; Shimer, G. H.; Meehan, T. Biochemistry 1987, 26,
6392.
(36) Cohen, G.; Eisenberg, H. Biopolymers 1969, 8, 45.
(37) Mossman, T. J. Immunol. Methods 1983, 65, 55.
(38) Wacker, W. E. C.; Ulmer, D. D.; Valee, B. L. J. Med. 1956, 255,
449.
(39) Stueher, D. J.; Marletta, M. A. J. Immunol. 1987, 139, 518.
(40) Xiong, X. B.; Ma, Z.; Lai, R.; Lavasanifar, A. Biomaterials 2010,
31, 757.
(41) Prabhakaran, R.; Jayabalakrishnan, C.; Krishnan, V.; Pasumpon,
K.; Sukanya, D.; Bertagnolli, H.; Natarajan, K. Appl. Organomet. Chem.
2006, 20, 203.
(42) Prabhakaran, R.; Karvembu, R.; Hashimoto, T.; Shimizu, K.;
Natarajan, K. Inorg. Chim. Acta 2005, 358, 2093.
(43) Prabhakaran, R.; Renukadevi, S. V.; Karvembu, R.; Huang, R.;
Mautz, J.; Huttner, G.; Subhaskumar, R.; Natarajan, K. Eur. J. Med.
Chem. 2008, 43, 268.
(44) Prabhakaran, R.; Renukadevi, S. V.; Karvembu, R.; Huang, R.;
Zeller, M.; Natarajan, K. Inorg. Chim. Acta 2008, 361, 2547.
(45) Prabhakaran, R.; Kalaivani, P.; Jayakumar, R.; Zeller, M.;
Hunter, A. D.; Renukadevi, S. V.; Ramachandran, E.; Natarajan, K.
Metallomics 2011, 3, 42.
(46) Karvembu, R.; Hemalatha, S.; Prabhakaran, R.; Natarajan, K.
Inorg. Chem. Commun. 2003, 6, 486.
(47) Tojal, J. G.; Dizarro, J. L.; Orad, A. G.; Sanz, A. R. P.; Ugalda,
M.; Diaz, A. A.; Serra, J. L.; Arriortua, M. I.; Rojo, T. J. Inorg. Biochem.
2001, 86, 627.
(48) Kalaivani, P.; Prabhakaran, R.; Dallemer, F.; Poornima, P.;
Vaishnavi, E.; Ramachandran, E.; Vijaya Padma, V.; Renganathan, R.;
Natarajan, K. Metallomics 2012, 4, 101.
(49) Rodrigues, C.; Batista, A. A.; Aucélio, R. Q.; Teixeira, L. R.;
Visentin, L. C.; Beraldo, H. Polyhedron 2008, 27, 3061.
(50) Basuli, F.; Ruf, M.; Pierpont, C. G.; Bhattacharya, S. Inorg. Chem.
1998, 37, 6113.
(51) (a) Chellan, P.; Land, K. M.; Shokar, A.; Au, A.; Clavel, C. M.;
Dyson, P.; Kock, C.; Smith, P. J.; Chibale, K.; Smith, G. S.
Organometallics 2012, 31, 5791. (b) Ali, A. A.; Nimir, H.; Aktas, C.;
Huch, V.; Rauch, U.; Schafer, K. H.; Veith, M. Organometallics 2012,
31, 2256. (c) Carreira, M.; Sanjuan, R. C.; Sanau, M.; Marzo, I.; Cont,
M. Organometallics 2012, 31, 5772.
(52) Prabhakaran, R.; Anantharaman, S.; Thilagavathi, M.; Kaveri, M.
V.; Kalaivani, P.; Karvembu, R.; Dharmaraj, N.; Bertagnolli, H.;
Dallemer, F.; Natarajan, K. Spectrochim. Acta, Part A 2011, 78, 844−
853.
(53) (a) Basuli, F.; Peng, S. M.; Bhattacharya, S. Inorg. Chem. 1997,
36, 5645. (b) Basuli, F.; Peng, S. M.; Bhattacharya, S. Inorg. Chem.
2000, 39, 1120. (c) Basuli, F.; Peng, S. M.; Bhattacharya, S. Inorg.
Chem. 2001, 40, 1126.
(54) Prabhakaran, R.; Huang, R.; Karvembu, R.; Jayabalakrishnan, C.;
Natarajan, K. Inorg. Chim. Acta 2007, 360, 691.
(55) Wallace, A. W.; Murphy, W. R.; Peterson, J. D. Inorg. Chim. Acta
1989, 166, 47.
(56) (a) Long, E. C.; Barton, J. K. Acc. Chem. Res. 1990, 23, 271.
(b) Pasternack, R. F.; Gibbs, E. J.; Villafranca, J. J. Biochemistry 1983,
22, 251.
(57) Kalaivani, P.; Prabhakaran, R.; Ramachandran, E.; Dallemer, F.;
Paramaguru, G.; Renganathan, R.; Poornima, P.; Vijaya Padma, V.;
Natarajan, K. Dalton Trans. 2012, 41, 2486.
(58) Li, Y.; Yang, Z. Y.; Wu, J. C. Eur. J. Med. Chem. 2010, 45, 5692.
(59) Prabhakaran, R.; Kalaivani, P.; Poornima, P.; Dallemer, F.;
Paramaguru, G.; Vijaya Padma, V.; Renganathan, R.; Huang, R.;
Natarajan, K. Dalton Trans. 2012, 41, 9323.
(60) Alia, M.; Ramos, S.; Mateos, R.; Serrano, A. B. G.; Bravo, L.;
Goya, L. Toxicol. Appl. Pharmacol. 2006, 212, 110.
(61) Yu, L.; Gengaro, P. E.; Niederberger, M.; Burke, T. J.; Schriert,
R. W. Proc. Natl. Acad. Sci. U.S.A. 1994, 91, 1691.
■
NOTE ADDED AFTER ASAP PUBLICATION
In the version of this paper published on Nov 28, 2012, ref 44
had the wrong volume and page information. In addition, ref 58
was a duplicate of ref 48. In the version published on Dec 10,
2012, ref 44 is correct and ref 58 has been deleted; the
references following the original ref 58 have been renumbered
and their citations in the text updated.
8332
dx.doi.org/10.1021/om300914n | Organometallics 2012, 31, 8323−8332