New ruthenium(II)/phosphines/diimines complexes: Promising antitumor (human breast cancer) and Mycobacterium tuberculosis fighting agents

Article abstract of DOI:10.1016/j.poly.2013.01.004

The synthesis and characterization of ruthenium compounds of the type [RuCl₂(P)₂(N-N)] [(P)₂ = (PPh₃) ₂, dppb = 1,4-bis(diphenylphosphino)butano; dppp = 1,3-bis(diphenylphosphino)propane; N-N = 5,5′-dimethyl-2,2′dipyridyl (5,5′-mebipy) or 4,4′-dimethyl-2,2′dipyridyl (4,4′-mebipy)] are described. The complexes were characterized using elemental analysis, UV-Vis and infrared spectroscopies, cyclic voltammetry, and X-ray crystallography. In vitro evaluation of the complexes, using the MTT methodology, revealed their cytotoxic activities in a range of 5.4-15.7 μM against the MDA-MB-231 breast tumor cells and showed that, in this case, they are more active than the reference metallodrug cisplatin. The in vitro antimycobacterial activities of the complexes had their Minimum Inhibitory Concentration (MIC) for MTB cell growth measured, by the REMA method. The MICs for these complexes were found to be between 12.5 and 25.0 μg/mL. The results are comparable with the second line drug cycloserine (MIC = 12.5-50.0 μg/mL), commonly used in the treatment of TB.

Full text of DOI:10.1016/j.poly.2013.01.004

Polyhedron 51 (2013) 292–297
Contents lists available at SciVerse ScienceDirect
Polyhedron
journal homepage: www.elsevier.com/locate/poly
New ruthenium(II)/phosphines/diimines complexes: Promising antitumor
(human breast cancer) and Mycobacterium tuberculosis fighting agents
Edjane R. dos Santos a, Melina A. Mondelli a, Lucas V. Pozzi a, Rodrigo S. Corrêa a,
Heloisa S. Salistre-de-Araújo b, Fernando R. Pavan c, Clarice Q.F. Leite c, Javier Ellena d, Valéria R.S. Malta e,
Sérgio P. Machado f, Alzir A. Batista a,
⇑
a Departamento de Química, Universidade Federal de São Carlos, CEP 13565-905 São Carlos, SP, Brazil
b Departamento de Ciências Fisiológicas, Universidade Federal de São Carlos, CP 676, CEP 13565-905 São Carlos, SP, Brazil
c Departamento de Ciências Biológicas, Faculdade de Ciências Farmacêuticas, UNESP, CEP 14800-900 Araraquara, SP, Brazil
d Instituto de Física de São Carlos, Universidade de São Paulo, CEP 13560-970 São Carlos, SP, Brazil
e Departamento de Química,Universidade Federal de Alagoas, CEP 57072-970 Maceio, AL, Brazil
f Departamento de Química Inorgânica, Instituto de Química, Universidade Federal do Rio de Janeiro, CEP 21941-909 Rio de Janeiro, RJ, Brazil
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis and characterization of ruthenium compounds of the type [RuCl2(P)2(N–N)] [(P)2 = (PPh3)2,
dppb = 1,4-bis(diphenylphosphino)butano; dppp = 1,3-bis(diphenylphosphino)propane; N–N = 5,50-
dimethyl-2,20dipyridyl (5,50-mebipy) or 4,40-dimethyl-2,20dipyridyl (4,40-mebipy)] are described. The
complexes were characterized using elemental analysis, UV–Vis and infrared spectroscopies, cyclic vol-
tammetry, and X-ray crystallography. In vitro evaluation of the complexes, using the MTT methodology,
Received 31 October 2012
Accepted 7 January 2013
Available online 17 January 2013
Keywords:
revealed their cytotoxic activities in a range of 5.4–15.7 lM against the MDA-MB-231 breast tumor cells
Ruthenium(II) complexes
Phosphine/diimine complexes
Cytotoxity
and showed that, in this case, they are more active than the reference metallodrug cisplatin. The in vitro
antimycobacterial activities of the complexes had their Minimum Inhibitory Concentration (MIC) for MTB
cell growth measured, by the REMA method. The MICs for these complexes were found to be between
Antimycobacterial activity
12.5 and 25.0
lg/mL. The results are comparable with the ‘‘second line’’ drug cycloserine (MIC = 12.5–
50.0 g/mL), commonly used in the treatment of TB.
l
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
general structural composition [Ru(g
6-arene)X2(PTA)] (PTA =
1,3,5-triaza-7-phosphaadamantane) (RAPTA) showed antimeta-
static properties and generally low toxicity comparable to those
observed for NAMI-A [3].
Since the discovery of the cisplatin as an antitumor agent in the
sixties, extensive research with transition metal ions has been
done and ruthenium has attracted a considerable interest as the
basis for new potential metallodrugs to treat cancer diseases [1–
3]. Metallopharmaceuticals have enormous potential to help over-
come the limitations of current therapies in cancer due to their
variety of geometries and chemical reactivities [4]. The activities
of these compounds depend on the metal ion, their ligands and
the structure of the compounds. Thus, NAMI-A, imidazolium
trans-[tetrachloro(dimethylsulfoxide)(1H-imidazole)ruthenate(III)],
and KP1019, indazolium trans-[tetrachlorobis(1H-indazole) ruthe-
nate(III)], are potential antitumor drugs that are in clinical trials
having successfully completed phase I. The success of these two
compounds reinforces the scope to continue searching for new
drugs that may have the cure, or for diminishing unwanted effects
of cancer treatment. Also, the bifunctional complexes of the
The interest in synthesizing new metal compounds is in dimin-
ishing the diverse side effects, as nephrotoxicity, toxicity, nausea
and vomiting, in cancer patients treated with the cisplatin. In the
last five years we have been synthesizing some ruthenium/phos-
phine/diimine complexes, which are showing promising activity
against cancer and also against Mycobacterium tuberculosis [MT]
[5–9]. The bacterium M. tuberculosis is the tuberculosis agent,
which is responsible for the death of about 2 million people annu-
ally [10]. The treatment of the illness is expensive, long, causes
some side effects and encounters resistance. Therefore many
researchers are looking for new alternatives in the treatment of
this illness such as some ruthenium and gold complexes that have
been found to be very promising, showing minimum inhibitory
concentrations (MICs) comparable to or even better than some ref-
erence drugs used in the treatment of tuberculosis [7–9,10].
In this work, four new ruthenium complexes were synthesized,
characterized, their X-rays structures were determined, and their
⇑
Corresponding author.
E-mail address: daab@ufscar.br (A.A. Batista).
0277-5387/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.poly.2013.01.004

E.R. dos Santos et al. / Polyhedron 51 (2013) 292–297
293
citotoxicity tested against MDA-MB-231 (human breast cancer)
cancer cells and against MT.
2.2.3. Synthesis of cis-[RuCl2(dppp)(N–N)]; N–N = 5,50-mebipy (3) or
(4,40-mebipy) (4)
These complexes were synthesized by reacting 0.300 g
(0.340 mmol) of [RuCl2(PPh3)2(N–N)], [N–N = 5,50-mebipy (3) or
4,40-mebipy (4)] in 30.0 mL of a mixture of dichlorometane:ben-
zene (1:1) with 0.210 g (0.510 mmol) of dppp [1,3-bis(diphenil-
phosphine) propane]. For the syntheses of the complex (3) and
(4) the solutions were refluxed for 4 days. (3): Yield: 0.223 g
(85%). Experimental Anal. Calc. for C39H38Cl2N2P2Ru: C, 60.90; H,
5.27; N, 3.27. Found: C, 60.94; H, 4.98; N, 3.64%. Molar conductance
2. Experimental
2.1. Materials and measurements
All manipulations were carried out under purified argon using
standard Schlenk techniques. Reagent grade solvents were appro-
priately distilled and dried before use. All chemicals used were of
reagent grade or comparable purity. All chemicals were purchased
from Aldrich or Fluka and were used as received. The precursors
[RuCl2(PPh3)3] and [RuCl2(dppb)(PPh3)] were prepared using pub-
lished procedures [11–13].
(l
S/cm, CH2Cl2) 1.7. 31P{1H} NMR (162 MHz, CH2Cl2/D2O) d(ppm)
2
38, (d) and 30.2 (d) Jp–p(Hz) 42,2. Selected IR (cmꢀ1): 1618 [w,
m
(C@N)], 1483 [s,
444 [w, (Ru–N)], 301 and 274 [w,
kmax/nm (
/Mꢀ1 cmꢀ1): 292 (13885), 424 (1954).
m
(C@N)], 1433 [s,
m
(C@N)], 517 [s,
m (Ru–P)],
m
e
m(Ru–Cl)]. UV–Vis CH2Cl2,
The IR spectra of the complexes were recorded on a FTIR Bo-
mem-Michelson 102 spectrometer in the 4000–200 cmꢀ1 region
using solid samples pressed in CsI pellets. The NMR spectra of
31P{1H} were recorded using a BRUKER, DRX400 tesla equipment,
in a BBO 5 mm probe at room temperature, and dichloromethane
was used as solvent, using a capillary containing D2O. The molar
(4): Yield: 0.236 g (90%). Experimental Anal. Calc. for C39H38Cl2-
N2P2Ru: C, 61.05; H, 5.01; N, 3.40. Found: C, 60.94; H, 4.98; N, 3.64%.
Molar conductance (
l
S/cm, CH2Cl2) 2.0. 31P{1H} NMR (162 MHz,
2
CH2Cl2/D2O) d(ppm) 39.3 (d) and 28.8 (d) Jp–p(Hz) 42.0.Selected
IR(cmꢀ1): 1585 [w,
515 [s, (Ru–P)], 442 [w,
UV–Vis CH2Cl2 kmax./nm (
/Mꢀ1 cmꢀ1): 306 (14252), 424 (1947).
m
(C@N)], 1481[s,
m
(C@N)], 1433 [s,
m(C@N)],
m
m(Ru–N)], 303 e 276 [w,
m(Ru–Cl)].
conductance measurements (K) were carried out in dichlorometh-
e
ane at 25 °C, using concentrations of 1.0 ꢁ 10ꢀ3 M for the com-
plexes. Cyclic voltammetry (CV) experiments were carried out at
room temperature in CH2Cl2 containing 0.100 M Bu4NClO4 (TBAP)
(Fluka Purum) using a BAS-100B/W Bioanalytical Systems Instru-
ment. The working and auxiliary electrodes were stationary Pt
foils; the reference electrode was of Ag/AgCl, in a Luggin capillary
probe. The electronic spectra were obtained through a scanning
done on a Hewlett–Packard diode array, model 8452A, spectropho-
tometer. The microanalyses were performed at the Microanalytical
Laboratory at the Federal University of São Carlos, São Carlos, São
Paulo, using a FISIONS CHNS, mod. EA 1108 micro analyzer.
2.3. Cell culture assay in MDA-MB-231
In vitro cytotoxicity assays on cultured human tumor cell lines
still represent the standard method for the initial screening of anti-
tumoral agents. Thus, as a first step in assessing their pharmaco-
logical properties, the new ruthenium complexes were assayed
against a human breast tumor cell line MDA-MB-231 (ATCC No.
HTB-26). The cells were routinely maintained in Dulbecco’s Modi-
fied Eagle’s medium (DMEM) supplemented with 10% fetal bovine
serum (FBS), at 37 °C in a humidified 5% CO2 atmosphere. After
reaching confluence, the cells were detached by trypsinization
and counted. For the cytotoxicity assay, 5 ꢁ 104 cells wellꢀ1 were
2.2. Synthesis
seeded in 200 lL of complete medium in 96-well assay microplates
(Corning Costar). The plates were incubated at 37 °C in 5% CO2 for
24 h to allow cell adhesion, prior to drug testing. All tested com-
pounds were dissolved in sterile DMSO (stock solution with max-
imum concentration of 20 mM) and diluted to 5, 2, 1, 0.5, 0.2,
2.2.1. Synthesis of [RuCl2(PPh3)2(5,50-mebipy)] (1)
The complex was obtained by reacting 0.500 g (0.520 mmol) of
[RuCl2(PPh3)3] dissolved in 20.0 mL of dichloromethane, previously
deaerated, and after that 0.106 g (0.570 mmol) of 5,50-mebipy was
added. For the syntheses of the complex (1) and (2) the solutions
were refluxed for 2 h, after that the volumes were diminished until
2 mL and diethyl ether was added for the precipitation of the de-
sired product. The solids were filtered off, well rinsed with diethyl
ether (3 ꢁ 5 mL) and dried in vacuum. Yield 0.413 g (90%) Experi-
mental Anal. Calc. for C48H42P2N2Cl2Ru: C, 65.84; H, 4.66; N, 3.22.
0.02 and 0.002 mM. From each of these dilute samples 2
lL ali-
quots were added to 200 L medium (without FBS) giving a final
l
concentration of DMSO of approximately 1% and a final concentra-
tion of the complex diluted about 100 times. Cells were exposed to
the compounds for a 24 h-period. Cell respiration, as an indicator
of cell viability, was determined by the mitochondrial-dependent
reduction of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetra-
zolium bromide] to formazan [14]. MTT solution (0.5 mg/mL) was
Found: C, 65.46; H, 4.81; N, 3.18%. Molar conductance (
Cl2) 3.2. 31P{1H} NMR (162 MHz, CH2Cl2/D2O) d(ppm) 27.6 (s) Se-
lected IR(cmꢀ1; w = weak; s = strong): 1604 [w,
(C@N)], 1481[s,
(C@N)], 1435 [s, (C@N)], 517 [s, (Ru–P)], 461 [w, (Ru–N)],
303 e 287 [w, (Ru–Cl)]. UV–Vis, CH2Cl2 kmax/nm (
/Mꢀ1 cmꢀ1):
lS/cm, CH2-
added to cell cultures and incubated for 3 h, after which 100 lL of
m
isopropanol was added to dissolve the precipitated formazan crys-
tals. The conversion of MTT to formazan by metabolically viable
cells was monitored in an automated microplate reader at
570 nm. The percent cell viability was calculated by dividing the
average absorbance of the cells treated with the test compounds
by that of the control; % cell viability was plotted against drug con-
centration (logarithmic scale) to determine the IC50 (drug concen-
tration at which 50% of the cells are viable relative to the control),
with the error estimated from the average of three trials.
m
m
m
m
m
e
272 (22391), 306 (21361), 460 (1458).
2.2.2. Synthesis of cis-[RuCl2(dppb)(5,50-mebipy)] (2)
The complex was obtained by reacting 0.300 g (0.350 mmol) of
[RuCl2(dppb)(PPh3)] dissolved in 20.0 mL of dichloromethane, pre-
viously deaerated, and after that 0.071 g (0.380 mmol) of 5,50-me-
bipy was added. Yield 0.246 g (90%). Experimental Anal. Calc. for
C
40H40Cl2N2P2Ru: C, 61.24; H, 5.22; N, 3.68. Found: C, 61.38; H,
5.15; N, 3.58%. Molar conductance (
S/cm, CH2Cl2) 4.5. 31P{1H}
NMR (162 MHz, CH2Cl2/D2O) d(ppm) 45.4 (d) and 31.8 (d)
2Jp–p(Hz) 32.7. Selected IR(cmꢀ1): 1637 [w,
(C@N)], 1475 [s,
(C@N)], 1433 [s, (C@N)], 519 [s, (Ru–P)], 436 [w, (Ru–N)],
301 e 267 [w, (Ru–Cl)]. UV–Vis CH2Cl2 kmax/nm (
/Mꢀ1 cmꢀ1):
312 (16594), 424 (2330).
2.4. Antimycobacterial activity assay
l
Antimycobacterial activities of each tested compound and of
the standard drug isoniazide (Difco laboratories, Detroit, MI, USA)
were determined in triplicate in 96 sterilized flat bottomed mi-
cro-plates (Falcon 3072; Becton Dickinson, Lincoln Park, NJ, USA)
and Middlebrook 7H9 Broth (Difco) supplemented with oleic
m
m
m
m
m
m
e

294
E.R. dos Santos et al. / Polyhedron 51 (2013) 292–297
Table 1
Crystal data and structure refinement for the complexes (2) cis-[RuCl2(dppb)(5,50-mebipy)], (3) cis-[RuCl2(dppp)(5,50-mebipy)] and (4) cis-[RuCl2(dppp)(4,40-mebipy)].
Empirical formula
Formula weight
Temperature(K)
Crystal system
Space group
Unit cell dimensions
(a) (Å)
C40H40Cl2N2P2Ru(2)
782.65
298(2)
Monoclinic
P21/c
C39H38Cl2N2P2RuꢂCH2Cl2(3)
C39H38Cl2N2P2Ru(4)
768.62
298(2)
Monoclinic
P21/c
853.55
298(2)
Triclinic
P-1
17.2615(2)
12.5182(1)
17.9871(2)
90
10.3775(9)
11.797(1)
16.787(1)
11.7965(1)
13.0803(2)
24.0323(4)
90
(b) (Å)
(c) (Å)
(a
) (°)
80.466(4)
(b) (°)
111.949(1)
90
3604.98(6)
4
1.442
0.703
75.158(3)
78.196(4)
1930.6(3)
2
1.468
0.797
872
0.20 ꢁ 0.11 ꢁ 0.10
3.08–26.39
ꢀ12 6 h 6 12,
ꢀ14 6 k 6 14,
ꢀ20 6 l 6 20
14706
97.8870(10)
90
3673.15(9)
4
1.39
0.689
(c
) (°)
Volume (Å3)
Z
Density (calc) (Mg/m3)
Absorption coefficient (mmꢀ1
F(000)
)
1608
1576
Crystal size (mm3)
0.50 ꢁ 0.17 ꢁ 0.07
3.02–26.71
ꢀ2l 6 h 6 2l,
ꢀ15 6 k 6 15,
ꢀ22 6 l 6 22
27579
0.08 ꢁ 0.17 ꢁ 0.18
3.00–26.73
ꢀ14 6 h 6 14,
ꢀ16 6 k 6 16,
-29 6 l 6 30
26976
Theta range for data collection (°)
Index ranges
Reflections collected
Independent reflections
Maximum and minimum transmission
7570, R(int) (0.0381)
0.956 and 0.643
R1 = 0.0365,
wR2 = 0.0955
R1 = 0.0448,
wR2 = 0.1012
0.500 and ꢀ0.740
1.034
7843, R(int) (0.0296)
0.923 and 0.864
R1 = 0.0505,
wR2 = 0.1162
R1 = 0.0624,
wR2 = 0.1254
0.576 and ꢀ0.704
1.134
7759, R(int) (0.0491)
0.9544 and 0.9006
R1 = 0.0393,
wR2 = 0.0911
R1 = 0.0602,
wR2 = 0.0966
0.396 and ꢀ0.789
1.003
Final R indices [I > 2
R indices (all data)
r(I)]
Largest diffraction in peak and hole (e Åꢀ3
)
Goodness-of-fit (GOF) on F2
acid–albumin–dextrose–catalase (OADC) enrichment (BBL/Becton
Dickinson, Sparks, MD, USA). The concentrations of the tested com-
Table 2
Selected bond distances (Å) and bond angles (°) for the (2), (3) and (4) complexes.
pound ranged from 0.15 to 250
lg/mL and the isoniazide ranged
Distance (°)
(2)
(3)
(4)
from 0.015 to 1.0 g/mL. The micro-plate Alamar Blue assay [11]
l
Ru–N(1)
2.115(2)
2.134(2)
2.3016(6)
2.3189(6)
2.4237(6)
2.4614(6)
77.25(8)
98.85(5)
91.40(6)
102.40(5)
173.24(6)
95.31(2)
165.41(6)
83.43(5)
89.08(6)
85.53(6)
86.42(2)
87.72(2)
90.52(2)
2.095(3)
2.110(3)
2.2686(9)
2.3150(9)
2.4361(8)
2.4882(9)
77.55(11)
94.14(8)
91.84(8)
107.29(8)
173.24(8)
92.50(3)
169.49(8)
84.08(8)
91.94(8)
83.50(8)
85.86(3)
83.20(3)
95.09(3)
2.071(2)
2.107(2)
2.2719(7)
2.3278(7)
2.4378(7)
2.4940(7)
77.89(8)
95.61(6)
91.53(6)
102.19(6)
173.34(6)
95.08(3)
170.58(6)
83.49(6)
92.72(6)
83.94(6)
85.13(3)
89.45(3)
95.03(3)
(MABA) was used to measure the minimal inhibitory concentration
(MIC) for the tested compounds (minimum concentration neces-
sary to inhibit 90% growth of M. tuberculosis H37Rv ATCC 27294).
The development of the color pink, in the wells, was taken as an
indicator of bacterial growth and the maintenance of the color blue
as the contrary. Thus, the MIC was assumed to be the lowest con-
centration able to inhibit the change of color from blue to pink. MIC
values were determined by fluorescence measured on a SPECTRA-
fluor Plus microfluorimeter (TecanÒ), with excitation at 530 nm
and emission at 590 nm.
Ru–N(2)
Ru–P(1)
Ru–P(2)
Ru–Cl(1)
Ru–Cl(2)
N(1)–Ru–N(2)
N(1)–Ru–P(1)
N(2)–Ru–P(1)
N(1)–Ru–P(2)
N(2)–Ru–P(2)
P(1)–Ru–P(2)
N(1)–Ru–Cl(1)
N(1)–Ru–Cl(2)
N(2)–Ru–Cl(1)
N(2)–Ru–Cl(2)
P(1)–Ru–Cl(1)
P(2)–Ru–Cl(2)
Cl(1)–Ru–Cl(2)
2.5. X-ray diffraction data
The crystals of the complexes were grown by slow evaporation
of the solutions in dichloromethane/diethyl ether. Suitable yellow
crystals of complexes were grown by the diffusion of diethyl ether
into a dichloromethane solution of the complex. Crystals were
measured on an Enraf–Nonius KappaCCD diffractometer, using a
wavelength of 0.71073 (Å).
2.6. Theoretical calculations
The geometry optimization was performed using a i-7 com-
puter, with a GAUSSIAN 09 programXX with the B3LYP hybrid density
functional combined with the 6-31G(d,p) and LACVP⁄ basis set and
for Ru atoms. The calculation of frequencies used to find the min-
imum geometries does not have imaginary values [21].
The final unit cell parameters were based on all reflections. Data
collections were made using the COLLECT program [15]; integration
and scaling of the reflections were performed with the HKL DENZO–
SCALEPACK system of programs [16]. Absorption corrections were car-
ried out using the Gaussian method [17]. The structures were solved
by direct methods using the SHELXS-97 [18]. The model was refined
by using the least-squares full-matrix on F2 by means of SHELXL-97
[19]. The ORTEP views of the structures were prepared using the
ORTEP-3 for WINDOWS [20]. All the hydrogen atoms were stereochemi-
cally positioned and refined with the riding model. All H atoms were
located on stereochemical grounds. Data collections and experi-
mental details for the complexes are summarized in Table 1.
Relevant interatomic bond lengths and angles are listed in Table 2.
3. Results and discussion
3.1. Syntheses of the complexes
The molar conductance measurements for the prepared com-
plexes (1–4) were carried out in dichloromethane and the results
are consistent with neutral type compounds [22].

E.R. dos Santos et al. / Polyhedron 51 (2013) 292–297
295
3.3. Spectroscopical characterization
The 31P{1H} NMR spectra of the new compounds present a typ-
ical AX pattern, which corresponds to the observation of two dou-
blets in the spectrum. Then, differently from the precursor
complex, which the NMR 31P{1H} presents a singlet in 27.6 ppm,
the new diphosphine derivatives have magnetic nonequivalent
phosphorus atoms (P trans Cl and P trans L). It is suggested that
the most unshielded signal refers to the phosphorous trans to the
nitrogen of the pyridines, and the most shielded signal, which is
close to that observed for similar complexes, refers to the phospho-
rous trans to Cl atom (see Table 3) [28].
The IR spectra of the new complexes confirm the presence of
the ligands coordinated to the metal. The presence of two
m(Ru–Cl) bands in the IR spectra of the complexes supports the
cis position of the chlorine in their structures, as shown by the
X-ray technique. The electronic spectra of the complexes show a
band [two bands for the complex (1)] in the UV region, which
Fig. 1. ORTEP View of cis-[RuCl2(dppb)(5,50-mebipy)] (2) showing the atom-labeling
scheme and the 30% probability thermal ellipsoids.
can be attributed to the intra-ligand transitions (p?
p⁄) from its
comparison to the UV spectra of free ligands (PPh3, dppp, dppb,
4,40-mebipy and 5,50-mebipy), and a second band that corresponds
to the charge transfer transition (MLCT) of the metal to the ligand,
at lower energies [29,30].
3.2. Structural studies
The ORTEP diagrams of the complexes are shown in Figs. 1–3. In
the structures, the metal displays the expected distorted octahe-
dral coordination geometry.
3.4. Electrochemical behavior
For all three structures the distances of Ru–P(2) (phosphorus
atoms trans to nitrogen) are longer than the distances of Ru–(P1)
(phosphorus atoms trans to chloride). This can be a consequence
of the competitive effect between the phosphorus and the nitrogen
The results of the cyclic voltammetry measurements for the
complexes (1–4), in CH2Cl2 solutions, at room temperature, are
presented in Table 4 and Fig. 4. The complexes (1–4) showed qua-
si-reversible processes, corresponding to a one-electron redox pro-
cess RuIII/RuII.
atoms, two
p acceptor species, for the electronic density of the
ruthenium. This has also been observed in other ruthenium com-
plexes [23]. Also, the distances Ru–Cl(2) trans to phosphorus atoms
are longer than the ones in which the chlorides are trans to the
nitrogen atoms, Ru–Cl(1), which is a consequence to the strong
trans effect of the phosphorus atoms [24,25]. Another observation
is with respect to the trans Ru–Cl bond length, where the chloride
is trans to the phosphorus atoms. Thus, for the cis-RuCl2
(dppp)(5,50-mebipy), Ru(1)–Cl(2) = 2.4882(9) Å and for the cis-
A general trend is that complexes containing monodentate
phosphine ligands show lower oxidation potentials when com-
pared to similar complexes containing bidentate ligands, with sim-
ilar basicity. This can be explained by the strong competition
between the trans
p acceptor atoms for the same d electrons of
the metallic center (trans competitive effect) [30–32].
As can be seen in Table 4 the redox potentials of the complexes
follow the basicity of the bipyridine or of the diphosphine ligands,
and in Fig. 5 there is a good correlation between the values plotted
for the redox potentials of the complexes and the Mulliken charges
on the ruthenium atom (% of Ru d-orbitals): ꢀ0.104, ꢀ0.099 and
ꢀ0.091, for cis-[RuCl2(dppb)(5,50-mebipy)], cis-[RuCl2(dppp)(5,50-
mebipy)] and cis-[RuCl2(dppp)(4,40-mebipy)], respectively.
RuCl2(dppp)(4,40-mebipy),
Ru(1)–Cl(2) = 2.4940(7) Å
these
distances are only slightly longer than the Ru–Cl distance in the
cis-RuCl2(dppb)(5,50-mebipy), Ru(1)–Cl(2) = 2.4614(6) Å, but con-
sistent with the higher basicity of the dppb (pKa = 4.72), when
compared with the basicity of the dppp (pKa = 4.50) [26,27].
Fig. 2. ORTEP View of cis-[RuCl2(dppp)(5,50-mebipy)]ꢂCH2Cl2 (3), showing the atom-labeling scheme and the 30% probability thermal ellipsoids.

296
E.R. dos Santos et al. / Polyhedron 51 (2013) 292–297
300
200
100
0
703
-100
-200
518
400
0
200
600
800
1000
1200
Potential (mV)
Fig. 3. ORTEP View of cis-[RuCl2(dppp)(4,40-mebipy)] (4) showing the atom-label-
Fig. 4. The cyclic voltammogram of the cis-[RuCl2(dppp)(5,50-mebipy)] complex:
ingscheme and the 30% probability thermal ellipsoids.
TBAP 0.1 mol Lꢀ1; CH2Cl2; Ag/AgCl; scan rate of 100 mV sꢀ1
.
Table 3
31P{1H} NMR data of ruthenium containing compounds, in CH2Cl2
.
r= 0.997
760
Complexes
(d)
2Jp-p(Hz)
cis,trans-[RuCl2(PPh3)2(5,50-mebipy)]
cis-[RuCl2(dppb)(5,50-mebipy)]
cis-[RuCl2(dppp)(5,50-mebipy)]
cis-[RuCl2(dppp)(4,40-mebipy)]
27.6
–
740
720
700
680
660
640
620
45.4 e 31.8
39.3 e 28.8
38.8 e 30.2
32.7
42.0
42.2
Table 4
Electrochemical data for the complexes (1–4), scan rate of 100 mV/s⁄.
Complex
E1/2
Epa
(mV)
Epc
(mV)
Ipa/
Ipc
(mV)
cis,trans-[RuCl2(PPh3)2(5,50-
mebipy)]
cis-[RuCl2(dppb)(5,50-mebipy)]
cis-[RuCl2(dppp)(5,50-mebipy)]
cis-[RuCl2(dppp)(4,40-mebipy)]
340
379
303
1.1
625
610
600
670
703
754
580
518
445
1.0
1.0
1.3
-0.106 -0.104 -0.102 -0.100 -0.098 -0.096 -0.094 -0.092 -0.090
% of Ru d-orbitals
pKa of the ligands: dppb = 4.72
mebipy = 5.01.
;
dppp = 4.50 ; ;
4,40-mebipy = 4.92 5,50-
Fig. 5. Relationship between the oxidation potential [Epa(mV)] values of the
complexes vs. Mulliken charges on their metal centers.
3.5. Antitumor activity
Table 5
The MDA-MB-231 tumor cell line (human breast carcinoma)
was exposed to (2–4) compounds for a period of 48 h in order to
allow them to reach their biological target. The IC50 (drug concen-
tration at which 50% of the cells are viable relative to the control)
values, calculated from the dose-survival curves obtained after
48 h of drug treatment with an MTT assay, are shown in Table 5.
For comparison purposes, the cytotoxicity of cisplatin was evalu-
ated under the same experimental conditions.
The complexes showed a good activity against human MDA-
MB-231 cell lines, compared to cisplatin these complexes are 4
to 10-fold more active in fighting MDA-MB-231, indicating their
potentiality as anti-tumor agents.
IC50 values for the MDA-MB-231 cell line of Ru(II) complexes (1–4), and the cisplatin
measured in a DMSO solution.
Complex
IC50 l
mol Lꢀ1
cis,trans-[RuCl2(PPh3)2(5,50-mebipy)]
10.31 0.21
5.41 0.71
14.60 1.81
15.64 0.44
27.2 0.49
10.59 0.91
>200
>200
>200
63.0 5.0
cis-[RuCl2(dppb)(5,50-mebipy)]
cis-[RuCl2(dppp)(5,50-mebipy)]
cis-[RuCl2(dppp)(4,40-mebipy)]
PPh3
dppp
dppb9
4,40-Mebipy9
5,50-Mebipy
Cycloserine
3.6. Antimicobacterial activity
and cis-[RuCl2(dppp)(4,40-mebipy)], were tested against M. tuber-
culosis H37Rv ATCC 27294, the observed MICs are reported in
Table 6. The values obtained were comparable to cycloserine
The in vitro anti-M. tuberculosis properties of the free ligands
dppp, dppb and the precursors used to synthesize the complexes
cis-[RuCl2(dppb)(5,50-mebipy)],
cis-[RuCl2(dppp)(5,50-mebipy)]
(MIC 12.5–50.0 lg/mL), an actually used ‘‘second line’’ drug.

E.R. dos Santos et al. / Polyhedron 51 (2013) 292–297
297
Table 6
[3] P. Nowak-Sliwinska, J.R. van Beijnum, A. Casini, A.A. Nazarov, G. Wagnières, H.
van den Bergh, P.J. Dyson, A.W. Griffioen, J. Med. Chem. 54 (2011) 3895.
[4] S. Ahmad, A.A. Isab, S. Ali, A.R. Al-Arfaj, Polyhedron 25 (2006) 1633.
[5] C. Golfeto, G. Von Poelhsitz, H.S. Selistre-de-Araújo, M.P. de Araujo, J. Ellena,
E.E. Castellano, L.G.L. França, I.S. Moreira, A.A. Batista, J. Inorg. Biochem. 104
(2010) 489.
[6] T.A. Heinrich, G. Von Poelhsitz, R.I. Reis, E.E. Castellano, A. Neves, M.
Lanznaster, S.P. Machado, A.A. Batista, C.M. Costa-Neto, Eur. J. Med. Chem. 46
(2011) 3616.
[7] F.R. Pavan, G. Von Poelhsitz, F.B. do Nascimento, S.R.A. Leite, A.A. Batista, Victor
M. Deflon, Daisy N. Sato, S.G. Franzblau, C.Q.F. Leite, Eur. J. Med. Chem. 45
(2010) 598–601.
[8] F.B. Nascimento, G. Von Poelhsitz, F.R. Pavan, D.N. Sato, C.Q.F. Leite, H.S.S.
Araujo, J. Ellena, E.E. Castellano, V.M. Deflon, A.A. Batista, J. Inorg. Biochem. 102
(2008) 1783.
MIC values of anti-M. tuberculosis activity of ruthenium complexes, free ligands and
reference drugs.
Complex
MIC (lg/mL)
cis,trans-[RuCl2(PPh3)2(5,50-mebipy)]
10.17
25.00
12.50
12.50
25
>50
>50
25
27
cis-[RuCl2(dppb)(5,50-mebipy)]
cis-[RuCl2(dppp)(5,50-mebipy)]
cis-[RuCl2(dppp)(4,40-mebipy)]
PPh3
dppp
dppb9
4,40-Mebipy9
5,50-Mebipy
Cycloserine
Isoniazide
[9] F.R. Pavan, G. Von Poelhsitz, M.I.F. Barbosa, S.R.A. Leite, A.A. Batista, J. Ellena,
L.S. Sato, S.G. Franzblau, V. Moreno, D. Gambino, C.Q.F. Leite, Eur. J. Med. Chem.
46 (2011) 5099.
12.5–50.0
0.03
[10] Global Alliance for TB Drug Development. www.tballiance.org (accessed
19.07.12).
[11] T.A. Stephenson, G. Wilkinson, J. Inorg. Nucl. Chem. 28 (1966) 945.
[12] S.L. Queiroz, M.P. de Araujo, A.A. Batista, K.S. MacFarlane, B.R. James, J. Chem.
Educ. 78 (2001) 87.
4. Conclusions
Four new ruthenium(II)/phosphines/diimines compounds were
synthesized, characterized and their biological activity evaluated.
The IC50 values for the MDA-MB-231 cell line of Ru(II) complexes
showed better activity than the cisplatin (reference drug), and
the MIC values of anti-M. tuberculosis activity obtained for the com-
plexes showed an activity comparable to cycloserine, a second line
drug used in the treatment of the illness. The crystal structures of
the [RuCl2(dppb)(5,50-mebipy)], [RuCl2(dppp)(5,50-mebipy)] and
[RuCl2(dppb)(4,40-mebipy)] complexes were determined and for
all of them the chlorines are in the cis position to each other, as
suggested by 31P{1H} NMR experiments.
[13] M. Bressan, P. Rigo, Inorg. Chem. 14 (1975) 2286.
[14] T. Mosmann, J. Immunol. Methods 65 (1983) 55.
[15] Enraf-Nonius, Collect, Nonius BV, Delft, The Netherlands, 1997–2000.
[16] Z. Otwinowski, W. Minor, Macromol. Crystallogr. Part A 276 (1997) 307.
[17] R.H. Blessing, Acta Crystallogr., Sect. A 51 (1995) 33.
[18] G.M. Sheldrick, SHELXS-97 Program for Crystal Structure Resolution, University
of Göttingen, Göttingen, Germany, 1997.
[19] G.M. Sheldrick, ShelXL-97, University of Göttingen, Göttingen, Germany,
Program for Crystal Structures Analysis, 1997.
[20] L.J. Farrugia, J. Appl. Crystallogr. 30 (1997) 565.
[21] M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Heeseman,
G. Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato,
X. Li, H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M.
Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y.
Honda, O. Kitao, H. Nakai, T. Vreven, J. A. Montgomery Jr., J.E. Peralta, F. Ogliaro,
M. Bearpark, J.J. Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, R. Kobayashi, J.
Normand, K. Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M.
Cossi, N. Rega, N.J. Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo, J.
Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C.
Pomelli, J.W. Ochterski, R.L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth,
P. Salvador, J.J. Dannenberg, S. Dapprich, A.D. Daniels, Ö. Farkas, J.B. Foresman,
J.V. Ortiz, J. Cioslowski, D.J. Fox, GAUSSIAN, Inc., Wallingford CT, 2009.
[22] W.J. Geary, Coord. Chem. Rev. 7 (1971) 81.
Acknowledgment
We thank CNPq, CAPES, FAPESP, FAPERJ and CYTED, for financial
support.
Appendix Supplementary. material
[23] G. Von Poelhsitz, M.P. de Araujo, L.A.A. de Oliveira, S.L. Queiroz, J. Ellena, E.E.
Castellano, A.G. Ferreira, A.A. Batista, Polyhedron 21 (2002) 2221.
[24] E.M.A. Valle, B.A.V. Lima, A.G. Ferreira, F.B. do Nascimento, Victor M. Deflon,
I.C.N. Diógenes, U. Abram, J. Ellena, E.D. Castellano, A.A. Batista, Polyhedron 28
(2009) 3473.
CCDC 906486, 906487 and 906488 contain the supplementary
crystallographic data for cis-[RuCl2(dppb)(5,50-mebipy)], cis-
[RuCl2(dppp)(5,50-mebipy)] and cis-[RuCl2(dppp)(4,40-mebipy)]
complexes. These data can be obtained free of charge via http://
www.ccdc.cam.ac.uk/conts/retrieving.html, or from the Cambridge
Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ,
UK; fax: +44 1223-336-033; or e-mail: deposit@ccdc.cam.ac.uk.
[25] A.A. Batista, L.A.C. Cordeiro, G. Oliva, O.R. Nascimento, Inorg. Chim. Acta 258
(1997) 131.
[26] S.J. Berners-Price, R.E. Norman, P.J. Sadler, J. Inorg. Biochem. 31 (1987) 197.
[27] J.R. Sowa, R.J. Angelici, J. Inorg. Chem. 30 (1991) 3534.
[28] S.L. Queiroz, A.A. Batista, G. Oliva, M.T. do P. Gambardella, R.H.A. Santos, K.S.
MacFarlane, S.J. Rettig, B.R. James, Inorg. Chim. Acta 267 (1998) 209.
[29] J. Chakravarty, S. Bhattacharya, Polyhedron 13 (1994) 2671.
[30] B. Sullivan, D. Salmon, T. Meyer, Inorg. Chem. 17 (1978) 3334.
[31] A.A. Batista, M.O. Santiago, C.L. Donnici, I.S. Moreira, P.C. Helay, S.J. Berners-
Price, S.L. Queiroz, Polyhedron 20 (2001) 2123.
References
[1] A. Bergamo, G. Sava, Dalton Trans. 40 (2011) 7817.
[2] A. Bergamo, A. Masi, A.F.A. Peacock, A. Habtemariam, P.J. Sadler, G. Sava, J.
Inorg. Biochem. 104 (2010) 79.
[32] A.A. Batista, L.R.V. Olmo, M.R.M. Fontes, G. Oliva, J. Braz. Chem. Soc. 7 (1996)
257.