Synthesis, characterization and in vitro anticancer activity of highly cytotoxic trithiolato diruthenium complexes of the type [(ν6-p-MeC 6H4iPr)2Ru2 (m 2-SR1)2 (m2-SR2)] + containing different thiolato bridges

Article abstract of DOI:10.1016/j.jorganchem.2013.04.049

A series of cationic dinuclear p-cymene ruthenium complexes containing three thiolato bridges with different substituents at the sulfur atoms, [(ν6-p-MeC₆H₄ ⁱPr) ₂Ru ₂ (m₂-SR¹)₂ (m₂-SR ²)]+ (R¹ = CH₂Ph, R² = Ph: 4; R ¹ = CH₂Ph, R² = p-C₆H₄ ⁱPr: 5; R¹ = CH₂Ph, R² = p-C ₆H₄^tBu: 6; R¹ = CH₂Ph, R² = p-C₆H₄ OH: 7; R¹ = CH ₂Ph, R² = p-C₆H₄ Br: 8; R ¹ = CH₂Ph, R² = p-C₆H₄ F: 9; R¹ = CH₂CH₂Ph, R² = Ph: 10; R¹ = CH₂CH₂Ph, R² = p-C ₆H₄ ⁱPr: 11; R¹ = CH ₂CH₂Ph, R² = p-C₆H₄ ^tBu: 12; R¹ = CH₂CH₂Ph, R ² = p-C₆H₄ OH: 13; R¹ = CH ₂CH₂Ph, R² = p-C₆H₄ Br: 14; R¹ = CH₂CH₂Ph, R² = p-C ₆H₄ F: 15; R¹ = CH₂C ₆H₄ -p-tBu, R² = Ph: 16; R¹ = CH₂C₆H₄ -p-tBu, R² = p-C ₆H₄ ⁱPr: 17; R¹ = CH ₂C₆H₄ -p-^tBu, R² = p-C₆H₄ ^t Bu: 18; R¹ = CH ₂C₆H₄ -p-tBu, R² = p-C ₆H₄ OH: 19; R¹ = CH₂C ₆H₄ -p-^tBu, R² = p-C ₆H₄ Br: 20; R¹ = CH₂C ₆H₄ -p-tBu, R² = p-C₆H₄ F: 21), have been obtained from the reaction of the neutral dithiolato intermediates [(ν6-MeC₆H₄ ⁱPr) ₂Ru₂Cl₂ (m₂-SR¹) ₂] (R¹ = CH₂Ph: 1; R¹ = CH ₂CH₂Ph: 2; R¹ = CH₂C ₆H₄ -p-^tBu: 3) with the corresponding thiophenol R²SH. All cationic complexes have been isolated as their chloride salts and fully characterized by spectroscopic and analytical methods. All complexes are highly cytotoxic against human ovarian cancer cells, the IC50 values being in the submicromolar range. The highest activity is shown by complex 6 with IC50 values of 48 nM against the A2780 cell line and 42 nM against the cisplatin-resistant line A2780cisR. This family of cationic trithiolato complexes belongs to the most cytotoxic ruthenium compounds ever reported. The catalytic activity selected representatives for the oxidation of glutathione (GSH) to GSSG has been investigated by NMR spectroscopy.

Full text of DOI:10.1016/j.jorganchem.2013.04.049

Journal of Organometallic Chemistry 744 (2013) 41e48
Contents lists available at SciVerse ScienceDirect
Journal of Organometallic Chemistry
journal homepage: www.elsevier.com/locate/jorganchem
Synthesis, characterization and in vitro anticancer activity of highly
cytotoxic trithiolato diruthenium complexes of the type
i
[(h
6-p-MeC6H4 Pr)2Ru2(m2-SR1)2(m2-SR2)]þ containing
different thiolato bridges
,
b
,
c
Federico Giannini a, Julien Furrer a , Georg Süss-Fink , Catherine M. Clavel ,
**
*
Paul J. Dyson c
a Department für Chemie und Biochemie, Universität Bern, CH-3012 Bern, Switzerland
b Institut de Chimie, Université de Neuchâtel, CH-2000 Neuchâtel, Switzerland
c Institut des Sciences et Ingénierie Chimiques, Ecole Polytechnique Fédérale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of cationic dinuclear p-cymene ruthenium complexes containing three thiolato bridges with
different substituents at the sulfur atoms, [(
h
6-p-MeC6H4 iPr)2Ru2(m2-SR1)2(m2-SR2)]þ (R1 ¼ CH2Ph,
Received 31 January 2013
Received in revised form
22 April 2013
R2 ¼ Ph: 4; R1 ¼ CH2Ph, R2 ¼ p-C6H4 iPr: 5; R1 ¼ CH2Ph, R2 ¼ p-C6H4 tBu: 6; R1 ¼ CH2Ph, R2 ¼ p-C6H4OH:
7; R1 ¼ CH2Ph, R2 ¼ p-C6H4Br: 8; R1 ¼ CH2Ph, R2 ¼ p-C6H4F: 9; R1 ¼ CH2CH2Ph, R2 ¼ Ph: 10;
R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4 iPr: 11; R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4 tBu: 12; R1 ¼ CH2CH2Ph, R2 ¼ p-
C6H4OH: 13; R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4Br: 14; R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4F: 15; R1 ¼ CH2C6H4ep-tBu,
R2 ¼ Ph: 16; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4 iPr: 17; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6Ht4Bu: 18;
R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4OH: 19; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4Br: 20; R1 ¼ CH2C6H4ep-tBu,
Accepted 28 April 2013
Keywords:
Arene ruthenium compounds
Dinuclear complexes
Thiolato complexes
Anticancer compounds
R2 ¼ p-C6H4F: 21), have been obtained from the reaction of the neutral dithiolato intermediates [(
h
6-p-
MeC6H4 iPr)2Ru2Cl2(m2-SR1)2] (R1 ¼ CH2Ph: 1; R1 ¼ CH2CH2Ph: 2; R1 ¼ CH2C6H4ep-tBu: 3) with the
corresponding thiophenol R2SH. All cationic complexes have been isolated as their chloride salts and
fully characterized by spectroscopic and analytical methods. All complexes are highly cytotoxic against
human ovarian cancer cells, the IC50 values being in the submicromolar range. The highest activity is
shown by complex 6 with IC50 values of 48 nM against the A2780 cell line and 42 nM against the
cisplatin-resistant line A2780cisR. This family of cationic trithiolato complexes belongs to the most
cytotoxic ruthenium compounds ever reported. The catalytic activity selected representatives for the
oxidation of glutathione (GSH) to GSSG has been investigated by NMR spectroscopy.
Ó 2013 Elsevier B.V. All rights reserved.
1. Introduction
other metals. Nowadays ruthenium complexes are among the
most studied non-platinum metal drugs for new therapies [3].
Cisplatin, [cis-PtCl2(NH3)2], has become the most widely used
metal-based anticancer drug ever since its introduction in 1978.
Cisplatin was shown to attack DNA in the cells, preferentially
binding to N7 of guanine, which ultimately triggers apoptosis
(programmed cells death) [1]. However, its use in cancer therapy is
limited by two factors: the established general toxicity of platinum
compounds and the development of resistance of some tumors after
an initial treatment of cisplatin [2]. These limitations have moti-
vated new studies on alternative anticancer drugs containing
Ruthenium forms strong complexes with numerous ligands [4],
different oxidation states are accessible under physiological condi-
tions [5], and it can potentially mimic iron in binding to important
carrier proteins [6,7]. Ruthenium(III) complexes such as NAMI-A,
with an imidazole ring coordinated to the ruthenium center, and
KP1019 with two indazole heterocycles coordinated to the metal
center, have already entered clinical trials [8]. Studies have shown that
KP1019 is efficiently taken up into cancer cells, possibly via interaction
with transferrin [9], where they appear to induce Fenton type redox
processes and intracellular radicals, leading to apoptosis [10]. Ruth-
enium(II) compounds are also interesting for the development of new
anticancer drugs, in particular arene ruthenium complexes, such as the
* Corresponding author. Tel.: þ41 (0) 32 718 2405; fax: þ41 (0) 32 718 2511.
** Corresponding author. Tel.: þ41 (0) 31 631 4383; fax: þ41 (0) 31 631 4887.
E-mail addresses: julien.furrer@dcb.unibe.ch (J. Furrer), georg.suess-fink@
unine.ch, georg.suess-fink@ich.unine.ch (G. Süss-Fink).
so-called RAPTA compounds containing
a
1,3,5-triaza-7-
phosphaadamantane (pta) ligand, the best characterized of the series
0022-328X/$ e see front matter Ó 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.jorganchem.2013.04.049

42
F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
being RAPTA-C [(
h
6-p-MeC6H4 iPr)RuCl2(pta)]. This compound has only
R2 ¼ p-C6H4F: 15; R1 ¼ CH2C6H4ep-tBu, R2 ¼ Ph: 16; R1 ¼ CH2C6H4e
p-tBu, R2 ¼ p-C6H4 iPr: 17; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4 tBu:
18; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4OH: 19; R1 ¼ CH2C6H4ep-tBu,
R2 ¼ p-C6H4Br: 20; R1 ¼ CH2C6H4ep-tBu, R2 ¼ p-C6H4F: 21), which
are isolated as orange to red air stable chloride salts in good to
excellent yields, see Scheme 1. The analytical data are given in the
Experimental section.
moderate in vitro anticancer activity, but significant activity with regard
to reducing the number and weight of solid metastases, although not
affecting the primary tumor [11].
Most studies on arene ruthenium compounds are concerned
with mononuclear complexes, although highly active dinuclear
arene ruthenium complexes have been reported: the dinuclear
arene ruthenium complexes (p-iPrC6H4Me)Ru(O,O-C6H5O2N(-
CH2)nNC6H5O2-O,O)Ru(p-iPrC6H4Me) containing a pyridone-deri
ved linker show relevant cytotoxic effects against various cancer
cell lines [12,13]. Recently, we have developed a new family of
cationic dinuclear arene ruthenium complexes of general formula
All compounds have been fully characterized by spectroscopic
analytical methods, the 1H and 13C NMR data providing a fingerprint
for these complexes. For instance, the1H NMR spectrum ofcomplex 4
(R1 ¼ CH2Ph and R2 ¼ Ph) exhibits three multiplets in the aromatic
region between 7.8 and 7.3 ppm, the first one at 7.78 ppm being
attributed to the two ortho protons of the R2 ligand and the second
one at 7.33 ppm being assigned to the three remaining protons of the
R2 ligand. Interestingly, the ten aromatic protons of the two R1 li-
gands appear as a single multiplet at 7.45 ppm. The two CH2 benzyl
groups show up as two singlets at 3.67 and 3.50 ppm, indicating that
the two R1 ligands are non-equivalent. As expected, their frequencies
are slightly shifted low-field, because of the presence of the sulfur
atom. The low-field shift of the signals of the aromatic protons of the
p-cymenegroup isalso characteristicforsuch complexes. Inthis case,
the coordination to the ruthenium center causes a chemical shift of
thefourcharacteristic doubletstolower frequencies between5.2and
4.5 ppm with a coupling constant of 6 Hz, thus confirming that the
two R1 ligands are not equivalent. The two isopropyl groups of the p-
cymene ligands give rise to a single septuplet at 1.9 ppm and to two
doublets at 0.97 and 0.91 ppm for the two equivalent dimethyl
groups. The two remaining methyl substituents of the two p-cymene
ligands appear as a singlet at 1.76 ppm. The 1H NMR spectrum of
complex 10 (R1 ¼ CH2CH2Ph and R2 ¼ Ph) reveals nearly the same
resonances as 4, the first multiplet at 7.71 ppm being assigned to the
two ortho protons of the R2 substituent and the multiplet at 7.30 ppm
being attributed to the three remaining protons of R2. The ten aro-
matic protons of the two R1 substituents appear as a single multiplet
at 7.31 ppm. Typical for the complexes containing R1 ¼ CH2CH2Ph are
four triplets in the region between 3.15 and 2.60 ppm, each with a
couplingconstantof7.6 Hz, reflectingthe fourdifferentCH2 groupsof
thetwo R1 substituents. In this case, the eight aromatic protons of the
two p-cymene rings collapse into a single multiplet at 5.2 ppm. The
1H NMR spectrum of complex 16 with R1 ¼ CH2ep-C6H4 tBu and
R2 ¼ Ph is similar, the resonances of the aromatic protons of the R1
ligands being displayed as two defined multiplets at 7.47 and
7.42 ppm, thus indicating the non-equivalence of the two groups.
This isfurtherconfirmed by two distinct singlets at 1.37 and1.33 ppm
for the two tert-butyl protons and by two singlets at 3.62 and
3.44 ppm for the two benzyl CH2 groups.
[(h
6-p-MeC6H4i Pr)2Ru2(m2-SR)3]þ (R being alkyl or aryl), in which
the two ruthenium atoms are bridged by three thiophenolato li-
gands. The corresponding chloride salts are highly cytotoxic toward
the human ovarian cancer cell line A2780 and its cisplatin-resistant
mutant A2780cisR, the IC50 values being in the submicromolar
range [14]. Incubation with biological targets such as nucleotides
and aminoacids, monitored by NMR spectroscopy, revealed in-
teractions only with cysteine and glutathione, causing their
oxidation to cystine and oxidized glutathione (GSSG), respectively.
The complex could be recovered intact after the oxidation, which
prompted us to postulate a catalytic role of the ruthenium complex
as mode of action in the cytotoxic activity [15]. Extension of the
series of dinuclear trithiophenolato compounds of general formula
[(h
6-p-MeC6H4 iPr)2Ru2(m2-S-p-C6H4-X)3]Cl, where X are various
functional groups showed that the cytotoxicity of the complexes is
clearly influenced by the lipophilicity and the Hammett’s constants
of the corresponding thiols, although a direct correlation between
cytotoxicity, catalytic oxidation activity and redox potentials could
not be established [16], which is perhaps not surprizing based on
the different levels of cellular uptake of the compounds.
Here we report the synthesis, the in vitro anticancer activity and
the catalytic glutathione oxidation activity of a new series of
eighteen dinuclear p-cymene ruthenium complexes of the general
formula [(h
6-p-MeC6H4 iPr)2Ru2(m2-SR1)2(m2-SR2)]þ, where R1 is an
aliphatic substituent and R2 an aromatic substituent.
2. Results and discussion
2.1. Synthesis and characterization
The neutral dichlorido dithiolato intermediates 1e3 are acces-
sible from the reaction of the p-cymeneeruthenium dichloride
dimer with 2 equivalents of phenylmethanethiol (for 1), 2-
phenylethanethiol (for 2) and (4-tert-butylphenyl)methanethiol
(for 3) in ethanol at 0 ꢀC, according to the published method [17].
These intermediates react in refluxing ethanol during 12e15 h with
an excess (usually 6 equivalents) of the corresponding thiol to give
2.2. Cytotoxicity of 4e21
the trithiolato complexes [(
h
6-p-MeC6H4 iPr)2Ru2(m2-SR1)2(m2
-
The in vitro anticancer activity of the trithiolato complexes was
evaluated against the human ovarian cancer cell line A2780 and the
cisplatin-resistant mutant A2780cisR, using the MTT [3-(4,5-
SR2)]þ (R1 ¼ CH2Ph, R2 ¼ Ph: 4; R1 ¼ CH2Ph, R2 ¼ p-C6H4 iPr: 5;
R1 ¼ CH2Ph, R2 ¼ p-C6H4 tBu: 6; R1 ¼ CH2Ph, R2 ¼ p-C6H4OH: 7;
R1 ¼ CH2Ph, R2 ¼ p-C6H4Br: 8; R1 ¼ CH2Ph, R2 ¼ p-C6H4F: 9;
R1 ¼ CH2CH2Ph, R2 ¼ Ph: 10; R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4 iPr: 11;
R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4 tBu: 12; R1 ¼ CH2CH2Ph, R2 ¼ p-
C6H4OH: 13; R1 ¼ CH2CH2Ph, R2 ¼ p-C6H4Br: 14; R1 ¼ CH2CH2Ph,
dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide]
assay,
which measures mitochondrial dehydrogenase activity as an indi-
cation of cell viability. The cytotoxicity of the complexes, expressed
as IC50 values which represent the drug concentration required to
-
+
R1
S
R1
S
Cl
Cl
Cl
Cl
Cl
Ru
Ru
Ru
S
R2
Ru
Ru
Ru
Cl
Cl
S
S
2 R1SH 2 HCl
R2SH
HCl
R1
R1
Scheme 1. Synthesis of complexes 4e21 as the chloride salts.

F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
43
complex 17 (R2 ¼ C6H4 iPr) and then by complex 18 (R2 ¼ C6H4 tBu).
For R1 ¼ CH2CH2Ph a relationship between the aliphatic nature of
the R2 substituent and the cytotoxicity is not apparent. Indeed,
complexes 11 and 12 with R2 ¼ C6H4 iPr and C6H4 tBu, respectively,
exhibit IC50 values lower than complex 10 where R2 ¼ Ph. When
the R2 substituent contains a heteroatom such as O, Br or F, the
nature of the R1 ligand has practically no influence on the bio-
logical activity, these complexes showing almost the same cyto-
toxicity, except complex 13 with R1 ¼ CH2CH2Ph and R2 ¼ p-
C6H4OH, which is by far the less cytotoxic of this series.
Table 1
Nanomolar cytotoxicities of complexes 4e21 toward A2780 and A2780cisR human
ovarian cancer cells.
Complex
IC50 [nM] A2780
IC50 [nM] A2780cisR
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
128 ꢁ 4
72.0 ꢁ 0.8
47.8 ꢁ 3
142 ꢁ 6
66.9 ꢁ 2.2
42.9 ꢁ 1
134 ꢁ 16.0
172 ꢁ 4
196 ꢁ 11
151 ꢁ 9
124 ꢁ 2.5
120 ꢁ 6
113.8 ꢁ 0.3
118.5 ꢁ 3.4
46.8 ꢁ 0.3
49.9 ꢁ 1.9
372 ꢁ 13
120.9 ꢁ 3.5
108 ꢁ 7
57.8 ꢁ 1.6
74.4 ꢁ 2.8
298 ꢁ 11
125.5 ꢁ 1.6
141 ꢁ 8
The subtle balance between hydrophobicity and lipophilicity of
R1 and R2 for this family of complexes is consistent with the results
we found for the series of trithiolato-bridged dinuclear ruthenium
arene complexes containing three identical R groups. In this case,
t
the cytotoxicity increases in the order H < Me < iPr < Bu [16].
55.5 ꢁ 0.9
153 ꢁ 8
41.4 ꢁ 0.7
218 ꢁ 14
159.5 ꢁ 1.3
118.3 ꢁ 2.1
100.7 ꢁ 1
123 ꢁ 4
163 ꢁ 8
2.3. Correlations between cytotoxicity and Hammett’s constants
and lipophilicity
132 ꢁ 3.3
97.3 ꢁ 0.6
139 ꢁ 10
The Hammett constants (sp) related to the para-substituted phenyl
rings of the R1 and R2 substituents reflect the electronic influence of
the substituents [20] and the partition coefficients (log P) of the thiols
R1SH and R2SH reflect the lipophilicity of the substituents. These
calculated values are given in Table 2. For clarity, the complexes have
been classified as a function of the R1 substituent. As the trithiolato-
bridged diruthenium core remains the same for all complexes, the
lipophilicity of these complexes should vary only as a function of the
R1SH and R2SH log Pparameters. Thepartitioncoefficients (log P)were
calculated using the ACD/ChemSketch software [21,22].
From Table 2, some interesting correlations between the Ham-
mett constants (sp) of the para-aryl substituted groups of R1 and R2,
the lipophilicity (log P) of the thiol ligands and the IC50 values for
the A2780 and A2780cisR cell lines can be extracted. Taking the
combined influences for the log P parameters for both R1 and R2
groups into account, it can be clearly seen that complexes with both
ligands having log P coefficients inferior to 3.5 have IC50 values
higher than 100 nM. In contrast, if one of the two thiols has a log P
value between 3.5 and 4.5, and the other one between 2.5 and 3.5,
then the biological activity against both cellular lines is enhanced,
presumably due to increased uptake as the more lipophilic com-
pounds can more readily traverse cell membranes.
cause 50% inhibition of cancer cell growth relative to control cells,
are reported in Table 1.
All the complexes are highly cytotoxic against both cell lines with
IC50 values in the nanomolar range. Complex 6, where R1 ¼ CH2Ph
and R2 ¼ C6H4 tBu, exhibits the highest cytotoxic effect against both
cell lines, with IC50 values of about 40 nM. These complexes are
amongst the most active anticancer ruthenium compounds ever
reported. To date, the most cytotoxic ruthenium complexes reported
include dimeric complexes with pyridone-derived linkers [12,13],
supramolecular arene ruthenium cages containing thiolato linkers
[18], and dendritic systems terminated with arene ruthenium
groups [19]. However, it should be noted that the supramolecular
and dendrimer systems contain multiple ruthenium centers and per
ruthenium the compounds reported herein are more cytotoxic.
The differences in cytotoxicity observed for these complexes
can be related to the R1 and R2 ligands. In the series where
R1 ¼ CH2Ph, the aliphatic character as well as the lipophilicity of
the substituent group in the p-position of the arene ligand grad-
ually increases in going from R2 ¼ Ph (4), to p-C6H4 iPr (5) and
p-C6H4 tBu (6), resulting in an enhanced cytotoxicity, see Table 2.
Interestingly, the other complexes do not follow the same ten-
dency, and for R1 ¼ CH2C6H4 tBu the trend is even reversed. Thus,
complex 16 (R2 ¼ Ph) exhibits the lowest IC50 value, followed by
From the data shown in Table 2, we were also able to extract a
relationship between the Hammett’s constants of the substituents
and the cytotoxicities. The complexes, in which one of the two R
Table 2
Comparison of cytotoxicities of 4e21 with physicochemical data for the thiols underlying the substituents R1 and R2. Calculated log P values are related to the thiols R1SH and
R2SH. Tabulated Hammett constants (sp) correspond to the p-C6H4X, where X is the substituent in para position of the phenyl ring of the R1 and R2 ligands.
Complex
R1
R2
Log P (R1SH)
sp-R1
Log P (R2SH)
sp-R2
IC50 [nM] A2780
IC50 [nM] A2780cisR
4
CH2Ph
Ph
Ph
Ph
2.74
2.87
4.43
2.74
2.87
4.43
2.74
2.87
4.43
2.74
2.87
4.43
2.74
2.87
4.43
2.74
2.87
4.43
0.00
0.00
2.52
2.52
2.52
3.86
3.86
3.86
4.21
4.21
4.21
1.68
1.68
1.68
3.53
3.53
3.53
2.81
2.81
2.81
0.00
0.00
0.00
128
120
55.5
72
142
10
16
5
11
17
6
12
18
7
13
19
8
14
20
9
15
21
CH2CH2Ph
CH2C6H4 tBu
CH2Ph
CH2CH2Ph
CH2C6Ht4Bu
CH2Ph
CH2CH2Ph
CH2C6H4 tBu
CH2Ph
CH2CH2Ph
CH2C6H4 tBu
CH2Ph
118.5
41.4
66.9
46.8
218
42.9
49.9
159.5
196
372
118.3
151
120.9
100.7
113.8
108
ꢂ0.20
C6H4 iPr
C6H4 iPr
C6H4 iPr
C6H4 tBu
C6H4 tBu
C6H4 tBu
C6H4OH
C6H4OH
C6H4OH
C6H4Br
C6H4Br
C6H4Br
C6H4F
0.00
ꢂ0.15
ꢂ0.15
ꢂ0.15
ꢂ0.20
ꢂ0.20
ꢂ0.20
ꢂ0.37
ꢂ0.37
ꢂ0.37
0.23
0.00
57.8
153
47.8
74.4
163
134
298
132
172
125.5
97.3
124
141
139
ꢂ0.20
0.00
0.00
ꢂ0.20
0.00
0.00
ꢂ0.20
0.00
CH2CH2Ph
CH2C6H4 tBu
CH2Ph
0.00
0.23
0.23
0.06
0.06
ꢂ0.20
0.00
CH2CH2Ph
C6H4F
C6H4F
0.00
ꢂ0.20
CH2C6H4 tBu
0.06
123

44
F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
groups (R1 or R2) has a sp value between 0 and ꢂ0.20, while the
other one has a sp ¼ 0.00, display the best IC50 values of the series.
All deviations from this optimal range of Hammett’s constants lead
to higher IC50 values.
COO-
N
COO-
N
-OOC
H2N
H
O
H
O
O
H
Interestingly, the two optimal ranges for both the Hammett’s
constants and the log P parameters coincide for the best cytotox-
icities. These results are in agreement with those obtained for our
O
H
2 e-
O
S-
N
N
H
O
N
S
S
2
previous series of ruthenium compounds [(h
6-p-MeC6H4 iPr)2
-
Ru2(m2-SR)3]Cl, the lowest IC50 values being obtained for complexes
having log P coefficients between 3.0 and 4.2 for the A2780 cell line,
and between 2.8 and 4.3 for the A2780cisR cell line, while the
Hammett constants were in the range between ꢂ0.20 and 0.00 [16].
In contrast to our previously reported series of cationic dinu-
clear p-cymene ruthenium trithiophenolato complexes of the type
2 e-
H
N
H2N
H2N
COO-
COO-
-OOC
[(
differences in the in vitro anticancer activity [16], all complexes of
the present series [(
6-p-MeC6H4 iPr)2Ru2(m2-SR1)2(m2-SR2)]þ are
h
6-p-MeC6H4 iPr)2Ru2(m2-SC6H4-p-X)3]þ, which showed large
(GSH)
(GSSG)
h
Scheme 2. Oxidation of glutathione (GSH) to give GSSG.
highly cytotoxic against human ovarian cancer cells. Our previous
results suggested that complexes possessing Hammett constants in
p-tBu) with GSH in a ratio 1:100, in a solution of D2O/DMSO-d6
(99:1), at pD 7 and 37 ꢀC and in aerobic atmosphere (the GSH
autoxidation in the presence of O2 being less than 5% in 24 h).
the range ꢂ0.2 <
sp < 0 and Log P values above 3.0 have the lowest
IC50 values, i.e. in the nanomolar region [16]. The cytotoxicities
found for the present series, which are much less varied, can be
explained by the fact that the physicochemical properties of the
compounds can be easily fine-tuned, since complexes 4e21 have
two different substituents R1 and R2 in the thiolato ligands. Thus, a
less favorable substituent R1 can be counterbalanced by a better
substituent R2. As a result, the overall cytotoxic properties of all
complexes are at least acceptable. Typical examples are complexes
9, 15, and 21 all containing a p-C6H4F substituent with non-optimal
physicochemical properties. Their IC50 values are still in the range
108e141 nM, thanks to the presence of a fluorine-free substituent
As evidenced by the disappearance of the
GSH at w3.0 ppm and the simultaneous appearance of two new
signals at w3.1 ppm and w 3.4 ppm, the six experiments led to
b-CH2 resonances of
d
d
d
the complete oxidation of GSH to GSSG within 24 h. The TOF50
values, which correspond to the turnover frequencies for each
complex as catalyst at about 50% conversion of GSH to GSSG, are
reported in Table 3, in comparison with the corresponding IC50
values.
A direct correlation of IC50 values with the TOF50 values cannot
be established. Indeed, the TOF50 of the most cytotoxic complex 6
(IC50 < 50 nM for both cell lines) is only 4.59 hꢂ1, whereas it in-
creases to 6.44 hꢂ1 for the less cytotoxic complex 8 (IC50 > 150 nM
for both cell lines) and goes to 7.23 hꢂ1 for complex 11 which has a
comparable biological activity as 6. While there is not a direct
correlation between catalytic oxidation of GSH and cytotoxicity, it is
not unexpected as the uptake of the complexes into cancer cells
depends on the log P values and therefore the amount of complex
in the cells differs for each complex. Thus, although it remains to be
proven that the mechanism of cell death is due to the catalytic
oxidation of GSH, such a mechanism cannot be excluded, at least in
part. Moreover, since the complexes show comparable cytotoxic-
ities in both the A2780 and cisplatin resistant A2780R cell lines a
mode of action different to that of cisplatin, i.e. primarily DNA
binding, can be excluded to some extent.
in the other thiolato bridges, as compared to [(h
6-p-MeC6H4 iPr)2
-
Ru2(m2-SC6H4-p-F)3]þ which is much less cytotoxic (IC50 ¼ 660 nM
for A2780 and IC50 ¼ 1050 nM for A2780cisR) [16].
2.4. Catalytic oxidation of glutathione
The dinuclear arene ruthenium trithiolato cations are inert to
substitution of both, arene ligands and thiolato bridges; therefore,
the question of their mode of action in living cell arises. Since we
had observed earlier that the complex [(h -
6-p-MeC6H4 iPr)2Ru2(m2
SC6H4-p-Me)3]þ catalyzes the oxidation of glutathione [16], it is
reasonable to assume that this might be at least one of the reasons
for the high cytotoxicity of this type of complexes.
The tripeptide glutathione (GSH) is found in all living cells with
a concentration of approximately 5 mM. It is derived from the three
aminoacids L-cysteine, L-glutamic acid and glycine. GSH represents
the major endogenous antioxidant in living cells, preventing
damage to important cellular components caused by reactive ox-
ygen species such as free radicals and peroxides [23]. In healthy
cells, more than 90% of the total glutathione pool is present in the
reduced form (GSH) and less than 10% exist in the oxidized disulfide
form (GSSG) [24,25].
3. Conclusion
To the best of our knowledge, the new diruthenium trithiolato
complexes, all obtained in good to excellent yields, are among the
most cytotoxic arene ruthenium compounds ever reported.
Recently, we studied the interaction of GSH with a series of
highly cytototoxic compounds of the general formula [(h
6-p-
Table 3
MeC6H4 iPr)2Ru2(SR)3]Cl (where R are p-substituted phenyl groups)
and found them to catalyze the oxidation of glutathione to GSSG in
water. However, a direct correlation between their cytotoxicity
(IC50) and their turnover frequency (TOF50) for their catalytic ac-
tivity of glutathione oxidation was not observed [16].
Comparison of cytotoxicities (IC50) with catalytic activity (TOF50) of the most and the
least cytotoxic complexes.
Complex
R1
R2
TOF50
IC50 [nM]
A2780
IC50 [nM]
A2780cisR
(hꢂ1
)
6
8
11
13
16
18
CH2Ph
CH2Ph
CH2CH2Ph
CH2CH2Ph
CH2C6H4 tBu
CH2C6H4 tBu
C6H4 tBu
C6H4Br
C6H4 iPr
C6H4OH
Ph
4.59
6.44
7.23
7.02
5.64
6.40
47.8
172
57.8
298
55.5
163
42.9
151
46.8
372
41.4
159.5
For this new series of complexes, we also studied the catalytic
activity for the oxidation of GSH to GSSG (Scheme 2) by NMR
spectroscopy. To evaluate the catalytic activity, we incubated the
most (lowest IC50) and the least (highest IC50) cytotoxic complex of
each of the three groups (R1 ¼ CH2Ph, CH2CH2Ph and CH2C6H4e
C6H4 tBu

F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
45
Interestingly, all complexes show comparable effects on both,
cisplatin-sensitive and cisplatin-resistant human ovarian cancer
cells. It can be assumed that the catalytic oxidation of GSH to GSSG
plays a role in the biological activity of these complexes, but other
modes of action such as interactions with enzymes and/or DNA,
may also be involved. The cytotoxicity does not appear to correlate
to the catalytic oxidation, but this may be due to the fact that the
catalytic oxidation was studied in NMR tubes, while the cytotoxicity
was determined with cancer cells. Therefore, the catalytic GSH
oxidation activity cannot be ruled out as mode of action of these
highly cytotoxic complexes, since the cytotoxicity correlates well
with lipophilicity, and hence uptake of the complexes into cancer
cells is likely to differ significantly.
From the results obtained, we can confirm that the lipophilicity
plays an important role for this kind of drugs, as also shown for
other types of arene ruthenium compounds [26,27]. The optimal
range of partition coefficients and Hammett’s constants leading to
the best IC50 values follows the same tendency found previously.
Overall, these results help to identify the structural features for
increasing the in vitro anticancer activity, the cellular uptake and
the selectivity for a particular biological target.
p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 139.8, 139.7, 137.7, 132.7, 129.6, 129.4, 129, 128.8, 128.6, 128.5,
128.2, 128.1, 107.2, 100.1, 83.8, 82.4, 40.2, 39.9, 30.9, 23.2, 22.5,
18.1 ppm.
4.2.2. Data for [5]Cl
Yield: 79.4 mg (87%). C43H53ClRu2S3 (904.11): calcd. C 57.15, H
5.91; found C 57.44, H 6.02. ESI MS (MeOH þ CH2Cl2 þ CH3CN): m/
z ¼ 868.6 [M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.61 (d, 3J ¼ 8.4 Hz,
2H, S-p-C6H4 iPr), 7.42 (m, 10H, SCH2C6H5), 7.12 (d, 3J ¼ 8.4 Hz, 2, S-
p-C6H4 iPr), 5.07 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.94 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.83 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 4.60 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.60 (s, 2H,
SCH2C6H5), 3.42 (s, 2H, SCH2C6H5), 2.86 [sept, 3J ¼ 6.8 Hz, 1H, S-p-
C6H4CH(CH3)2], 1.81 [sept, 3J ¼ 7.2 Hz, 2H, p-MeC6H4CH(CH3)2], 1.76
(s, 6H, p-CH3C6H4 iPr), 1.22 [d, 3J ¼ 6.8 Hz, 6H, S-p-C6H4CH(CH3)2],
0.92 [d, 3J ¼ 7.2 Hz, 6H, p-MeC6H4CH(CH3)2], 0.86 [d, 3J ¼ 7.2 Hz, 6H,
p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 147.7, 139.9, 139.8, 134.4, 132.6, 129.6, 129.4, 128.8, 128.7, 128.2,
128.0, 127.1, 106.7, 100.5, 84.6, 83.5, 83.4, 82.5, 40.3, 40.0, 33.8, 30.7,
23.9, 23.3, 22.4, 18.2 ppm.
4. Experimental
4.2.3. Data for [6]Cl
Yield: 89.9 mg (97%). C44H55ClRu2S3$3/4 CH2Cl2 (981.4): calcd. C
4.1. Materials and methods
54.77,
H
5.80; found
C
54.40,
H
6.12. ESI MS (MeOH
þ
CH2Cl2 þ CH3CN): m/z ¼ 882.7 [M]þ. 1H NMR (400 MHz, CDCl3):
The starting material [(
three neutral dithiolato complexes [(
SeCH2Ph)2] (1) [(
6-p-MeC6H4 iPr)2Ru2Cl2(m2-SeCH2CH2Ph)2] (2)
and [(
6-p-MeC6H4 iPr)2Ru2Cl2(m2-SeCH2-p-C6H4 tBu)2] (3) were
h
6-p-MeC6H4 iPr)2Ru2Cl4] and the
6-p-MeC6H4 iPr)2Ru2Cl2(m2
d
¼ 7.65 [d, 3J ¼ 8.4 Hz, 2H, S-p-C6H4C(CH3)3], 7.43 (m, 10H,
h
-
SCH2C6H5), 7.30 [d, 3J ¼ 8.4 Hz, 2H, S-p-C6H4C(CH3)3], 5.13 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.97 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 4.88 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.63 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.65 (s, 2H, SCH2C6H5), 3.47 (s, 2H,
SCH2C6H5), 1.79 [sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.77 (s,
6H, p-CH3C6H4 iPr), 1.30 [s, 9H, S-p-C6H4C(CH3)3], 0.91 [d,
3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.86 [d, 3J ¼ 7.2 Hz, 6H, p-
h
h
prepared according to published methods [16]. All other reagents
were commercially available and were used without further
purification.
NMR spectra were recorded with a Bruker 400 MHz spectrom-
eter. Electrospray mass spectra were obtained in positive- or
negative-ion mode with an LCQ Finnigan mass spectrometer. Mi-
croanalyses were performed by the Mikroelementaranalytisches
Laboratorium, ETH Zürich (Switzerland).
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 151.9,
139.9, 139.8, 134.2, 132.3, 129.6, 129.5, 128.8, 128.6, 128.2, 128.0,
125.9, 106.5, 100.6, 84.8, 83.4, 83.3, 82.6, 40.3, 40.1, 34.7, 31.2, 30.7,
23.4, 22.4, 18.2 ppm.
4.2. Synthesis of complexes [(
h
6-p-MeC6H4 iPr)2Ru2(m2-SR1)2(m2
-
4.2.4. Data for [7]Cl
SR2)]þ (4e21)
Yield: 75.3 mg (85%). C40H47ClORu2S3 (877.6): calcd. C 54.74, H
5.40; found C 54.55, H 5.70. ESI MS (MeOH þ CH2Cl2): m/z ¼ 842.3
After dissolution of the neutral dithiolato complex [1 (80 mg,
0.101 mmol), 2 (80 mg, 0.098 mmol), 3 (80 mg, 0.089 mmol)] in
technical-grade EtOH (50 ml) under reflux, the corresponding thiol
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 10.31 (s, 1H, S-p-C6H4OH), 7.41
(m, 14H, SCH2C6H5 þ S-p-C6H4OH), 5.06 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 4.94 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.71 (m, 4H, p-
MeC6H4 iPr), 3.62 (s, 2H, SCH2C6H5), 3.45 (s, 2H, SCH2C6H5), 2.04
[sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.73 (s, 6H, p-
CH3C6H4 iPr), 1.05 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.99 [d,
3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz,
(6 equiv.; SR ¼ SPh: 62.5
91.4 l, 83
l; S-p-C6H4 tBu: 105
76.8 mg, 74.2 mg, 67.4 mg; S-p-C6H4Br: 115 mg,111.2 mg,101 mg; S-
p-C6H4F: 64.9 l, 62.6 l, 56.9 l) was added to the hot solution,
ml, 60.4
m
l, 54.8
m
m
l; S-p-C6H4 iPr: 94.7
l, 92 l; S-p-C6H4OH:
ml,
m
m
ml, 101.4
m
m
m
m
which was then refluxed for 15 h. After evaporation of the solvent,
the residue was purified by column chromatography on silica gel
with dichloromethane/ethanol (9:1) as eluent. The yellow to
brownish products were isolated as chloride salts and dried under
vacuum.
CDCl3):
d
¼ 159.9,139.9,139.7,133.3,129.5,129.2,128.8,128.7,128.2,
128.1, 124.0, 117.1, 107.5, 99.7, 84.1, 83.7, 83.2, 82.0, 39.9, 39.5, 31.0,
23.1, 22.7, 18.0 ppm.
4.2.5. Data for [8]Cl
Yield: 85.5 mg (90%). C40H46BrClRu2S3$½CH2Cl2$EtOH (1029):
calcd. C 49.55, H 5.01; found C 49.85, H 5.16. ESI MS (MeOH): m/
4.2.1. Data for [4]Cl
Yield: 76.6 mg (88%). C40H47ClRu2S3$½ CH2Cl2 (904.05): calcd. C
53.81, H 5.35; found C 54.02, H 5.56. ESI MS (MeOH): m/z ¼ 826.6
z ¼ 904.8 [M]þ.1H NMR (400 MHz, CDCl3):
d
¼ 7.72 (d, 3J ¼ 8.4 Hz, 2H,
S-p-C6H4Br), 7.53 (m, 5H, SCH2C6H5), 7.48 (d, 3J ¼ 8.4 Hz, 2H, S-p-
C6H4Br), 7.41 (m, 5H, SCH2C6H5), 5.19 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 5.08(d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.90(d, 3J ¼ 6.0 Hz,
2H, p-MeC6H4 iPr), 4.75 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.67 (s, 2H,
SCH2C6H5), 3.50 (s, 2H, SCH2C6H5), 1.96 [sept, 3J ¼ 6.8 Hz, 2H, p-
MeC6H4CH(CH3)2],1.78 (s, 6H, p-CH3C6H4 iPr),1.00 [d, 3J ¼ 6.8 Hz, 6H,
p-MeC6H4CH(CH3)2], 0.94 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2]
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.78 (m, 2H, SC6H5), 7.45
(m, 10H, CH2C6H5), 7.33 (m, 3H, SC6H5), 5.16 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 5.05 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.88 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.71 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 3.67 (s, 2H, SCH2), 3.50 (s, 2H, SCH2) 1.9 [sept,
3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.76 (s, 6H, p-CH3C6H4 iPr),
0.97 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.91 [d, 3J ¼ 6.8 Hz, 6H,
ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 139.7, 139.6, 137.3, 134.3,

46
F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
131.9, 129.5, 129.3, 128.8, 128.6, 128.2, 128.1, 122.6, 107.3, 100.0, 84.0,
83.9, 83.8, 82.4, 40.2, 40.0, 31.0, 25.3, 23.1, 22.5, 18.1 ppm.
7.47 (d, 3J ¼ 8.8 Hz, 2H, S-p-C6H4OH), 7.39 (m, 10H, SCH2CH2C6H5),
7.16 (m, 2H, S-p-C6H4OH), 5.18 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr),
5.15 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.11 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 5.05 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.07 (m, 4H,
SCH2CH2), 2.83 (t, 3J ¼ 7.6 Hz, 2H, SCH2CH2), 2.66 (t, 3J ¼ 7.6 Hz, 2H,
SCH2CH2), 2.16 [sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.80 (s,
6H, p-CH3C6H4 iPr), 1.10 [m, 12H, p-MeC6H4CH(CH3)2] ppm. 13C{1H}
4.2.6. Data for [9]Cl
Yield: 72.8 mg (82%). C40H46ClFRu2S3$¼ CH2Cl2 (900.8): calcd. C
53.67, H 5.20; found C 53.32, H 5.41. ESI MS (MeOH): m/z ¼ 845.5
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.74 (m, 2H, S-p-C6H4F), 7.38
(m, 10H, SCH2C6H5), 6.98 (m, 2H, S-p-C6H4F), 5.10 (d, 3J ¼ 6.0 Hz,
2H, p-MeC6H4 iPr), 4.99 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.82 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.65 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 3.58 (s, 2H, SCH2C6H5), 3.41 (s, 2H, SCH2C6H5), 1.83
[sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.69 (s, 6H, p-
CH3C6H4 iPr), 0.91 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.85 [d,
3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm. 19F NMR (376 MHz,
NMR (100 MHz, CDCl3):
126.8, 125.2, 117.0, 107.1,100.0, 84.5, 84.0, 83.4, 82.9, 40.4, 40.0, 38.8,
38.7, 30.9, 23.2, 22.7, 17.8 ppm.
d
¼ 159.2, 139.9, 133.5, 128.8, 128.7, 126.9,
4.2.11. Data for [14]Cl
Yield: 80.7 mg (85%). C42H50BrClRu2S3$½ CH2Cl2 (1011.01):
calcd. C 50.49, H 5.08; found C 50.32, H 5.23. ESI MS (MeOH): m/
CDCl3):
d
¼ ꢂ112.0 ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 164.0,
z ¼ 932.8 [M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.63 (d, 3J ¼ 8.4 Hz,
161.5, 139.6, 134.5, 133.0, 129.5, 129.3, 128.7, 128.6, 128.2, 128.0,
116.1, 115.9, 107.1, 100.0, 83.9, 83.8, 83.7, 82.3, 40.1, 39.9, 30.8, 23.1,
22.4, 18.0 ppm.
2H, S-p-C6H4Br), 7.40 (m, 10H, SCH2CH2C6H5), 7.34 (m, 2H, S-p-
C6H4Br), 5.21 (m, 8H, p-MeC6H4 iPr), 3.12 (m, 2H, SCH2CH2), 3.06 (m,
2H, SCH2CH2), 2.94 (m, 2H, SCH2CH2), 2.66 (m, 2H, SCH2CH2), 2.10
[sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.85 (s, 6H, p-
CH3C6H4 iPr), 1.05 [m, 12H, p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR
4.2.7. Data for [10]Cl
Yield: 81.1 mg (93%). C42H51ClRu2S3$½ CH2Cl2 (932.1): calcd. C
54.76, H 5.62; found C 54.79 H 5.62. ESI MS (MeOH): m/z ¼ 854.19
(100 MHz, CDCl3):
128.6, 126.7, 122.4, 107.1, 100.0, 84.1, 83.8, 83.6, 83.0, 41.3, 39.9, 38.7,
38.6, 30.9, 22.8, 17.8 ppm.
d
¼ 140, 139.7, 137.7, 134.2, 131.8, 131.6, 128.7,
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.71 (m, 2H, SC6H5), 7.39 (m,
10H, SCH2CH2C6H5), 7.30 (m, 3H, SC6H5), 5.2 (m, 8H, p-MeC6H4 iPr),
3.11 (m, 4H, SCH2CH2), 2.93 (m, 2H, SCH2CH2), 2.69 (m, 2H,
SCH2CH2), 2.10 [sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2], 1.86 (s,
6H, p-CH3C6H4 iPr), 1.07 [m, 12H, p-MeC6H4CH(CH3)2] ppm. 13C{1H}
4.2.12. Data for [15]Cl
Yield: 80.9 mg (91%). C42H50ClFRu2S3$¼ CH2Cl2 (928.88): calcd.
C 54.63, H 5.48; found C 54.33, H 5.72. ESI MS (MeOH): m/z ¼ 872.1
NMR (100 MHz, CDCl3):
128.5, 126.9, 107.0, 100.3, 84.4, 84.1, 83.9, 83.2, 41.2, 40.2, 38.8, 38.7,
31.0, 23.3, 22.5, 17.9 ppm.
d
¼ 139.8, 138.3, 132.6, 128.9, 128.8, 128.7,
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.75 (m, 2H, S-p-C6H4F), 7.40
(m, 10H, SCH2CH2C6H5), 7.04 (m, 2H, S-p-C6H4F), 5.24 (m, 8H, p-
MeC6H4 iPr), 3.13 (m, 2H, SCH2CH2), 3.09 (m, 2H, SCH2CH2), 2.93 (m,
2H, SCH2CH2), 2.69 (m, 2H, SCH2CH2), 2.13 [sept, 3J ¼ 6.8 Hz, 2H, p-
MeC6H4CH(CH3)2], 1.87 (s, 6H, p-CH3C6H4 iPr), 1.08 [m, 12H, p-
4.2.8. Data for [11]Cl
Yield: 78.5 mg (86%). C45H57ClRu2S3$¼ CH2Cl2 (952.95): calcd. C
57.03, H 6.08; found C 57.09, H 6.32. ESI MS (MeOH): m/z ¼ 897.6
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 164.1,
161.6, 139.8, 134.5, 134.4, 128.9, 128.8, 128.7, 126.9, 116.1, 115.9, 107.1,
100.3, 84.5, 84.3, 83.8, 83.2, 41.4, 40.2, 38.8, 38.7, 31.0, 23.2, 22.5,
18.0 ppm.
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.53 [d, 3J ¼ 8.4 Hz, 2H, S-
p-C6H4CH(CH3)2], 7.43 (m, 10H, SCH2CH2C6H5), 7.08 [d, 3J ¼ 8.4 Hz,
2H, S-p-C6H4CH(CH3)2], 5.16 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.11
(d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.10 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 5.06 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.0 (m, 4H,
SCH2CH2), 2.82 [sept, 3J ¼ 6.8 Hz, 1H, S-p-C6H4CH(CH3)2], 2.78 (m,
2H, SCH2CH2), 2.62 (t, 3J ¼ 7.6 Hz, 2H, SCH2CH2), 1.95 [sept,
3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2],1.90 (s, 6H, p-CH3C6H4 iPr),1.29
4.2.13. Data for [16]Cl
Yield: 78 mg (90%). C48H63ClRu2S3$¼ CH2Cl2 (995.17): calcd. C
58.24, H 6.43; found C 58.24 H 6.76. ESI MS (MeOH): m/z ¼ 939.3
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.76 (m, 2H, SC6H5), 7.47
(m, 4H, S-p-CH2C6H4 tBu), 7.42 (m, 4H, S-p-CH2C6H4 tBu), 7.32 (m,
3H, SC6H5) 5.11 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.01 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.91 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 4.61 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.62 (s, 2H, S-p-
CH2C6H4 tBu), 3.44 (s, 2H, S-p-CH2C6H4 tBu), 1.88 [sept, 3J ¼ 6.8 Hz,
2H, p-MeC6H4CH(CH3)2], 1.76 (s, 6H, p-CH3C6H4 iPr), 1.37 [s, 9H,
SCH2C6H5C(CH3)3], 1.33 [s, 9H, SCH2C6H5C(CH3)3], 0.93 [d,
3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.88 [d, 3J ¼ 6.8 Hz, 6H, p-
[d, 3J
¼
6.8 Hz, 6H, S-p-C6H4CH(CH3)2], 1.00 [m, 12H, p-
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 149.7,
132.6, 128.9, 128.8, 127, 106.7, 100.6, 84.1, 83.9, 83.6, 83.4, 33.8, 30.8,
23.9, 23.4, 22.4, 18.0 ppm.
4.2.9. Data for [12]Cl
Yield: 77.8 mg (84%). C46H59ClRu2S3$¼ CH2Cl2 (966.99): calcd. C
57.45, H 6.20; found C 57.37, H 6.02. ESI MS (MeOH): m/z ¼ 911.6
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 151.7,
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.54 [d, 3J ¼ 8.0 Hz, 2H, S-
151.6, 137.8, 136.7, 132.7, 129.3, 129.2, 129.0, 128.5, 125.6, 125.4,
106.9, 100.4, 84.1, 83.7, 82.4, 40.0, 39.5, 34.8, 34.7, 31.4, 30.8, 23.1,
22.7, 18.2 ppm.
p-C6H4C(CH3)3], 7.35 (m, 10H, S-p-CH2CH2C6H5), 7.24 [m, 2H, S-p-
C6H4CH(CH3)2], 5.10 (m, 8H, p-MeC6H4 iPr), 3.04 (m, 2H, SCH2CH2),
2.97 (m, 2H, SCH2CH2), 2.82 (m, 2H, SCH2CH2), 2.58 (m, 2H,
SCH2CH2), 1.90 [m, 2H, p-MeC6H4CH(CH3)2], 1.80 (s, 6H, p-
CH3C6H4 iPr), 1.28 [s, 9H, S-p-C6H4C(CH3)3], 0.94 [m, 12H, p-
4.2.14. Data for [17]Cl
Yield: 78.6 mg (87%). C51H69ClRu2S3$2 CH3(CH2)4CH3
(1188.23): calcd. C 63.15, H 8.08; found C 63.41, H 8.32. ESI MS
(MeOH þ CH2Cl2): m/z ¼ 980.2 [M]þ. 1H NMR (400 MHz, CDCl3):
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
139.9,134.7,132.2,128.9,128.8,128.7,126.9,125.8,106.5,100.7, 84.5,
83.9, 83.7, 83.4, 41.2, 40.2, 38.9, 38.8, 34.7, 31.2, 30.7, 23.4, 22.3,
18.0 ppm.
d
¼ 151.9,
d
¼ 7.70 [d, 3J ¼ 8.4 Hz, 2H, S-p-C6H4CHiPr], 7.50 (m, 4H, S-p-
CH2C6H4 tBu), 7.46 (m, 4H, S-p-CH2C6H4 tBu), 7.20 [d, 3J ¼ 8.4 Hz,
2H, S-p-C6H4CHiPr], 5.16 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.03 (d,
4.2.10. Data for [13]Cl
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.97 (d, 3J
¼
6.0 Hz, 2H,
Yield: 78.1 (88%). C42H51ClORu2S3$¼ CH2Cl2 (923.83): calcd. C
54.75, H 5.60; found C 54.72, H 5.80. ESI MS (MeOH): m/z ¼ 870.8
p-MeC6H4 iPr), 4.60 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.65 (s, 2H,
S-p-CH2C6H4 tBu), 3.47 (s, 2H, S-p-CH2C6H4 tBu), 2.95 [sept,
3J ¼ 6.8 Hz, 1H, S-p-C6H4CH(CH3)2], 1.85 [sept, 3J ¼ 6.8 Hz, 2H,
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
¼ 9.79 (s, 1H, S-p-C6H4OH),
d

F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
47
p-MeC6H4CH(CH3)2], 1.82 (s, 6H, p-CH3C6H4 iPr), 1.41 [s, 9H, S-p-
CH2C6H4C(CH3)3], 1.37 [s, 9H, S-p-CH2C6H4C(CH3)3], 1.26 [m, 6H, S-
p-C6H4CH(CH3)2], 0.94 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2],
0.90 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR
4.88 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.57 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 3.56 (s, 2H, S-p-CH2C6H4 tBu), 3.38 (s, 2H, S-p-
CH2C6H4 tBu),1.85 [sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2],1.71 (s,
6H, p-CH3C6H4 iPr), 1.34 [s, 9H, SCH2C6H4C(CH3)3], 1.31 [s, 9H,
SCH2C6H4C(CH3)3], 0.89 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.85
[d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm.13C{1H} NMR (100 MHz,
(100 MHz, CDCl3):
d
¼ 151.7, 151.6, 136.8, 136.7, 134.5, 134.4, 132.5,
129.3, 129.1, 127.0, 125.6, 125.4, 106.6, 100.7, 84.6, 83.4, 83.3, 82.5,
34.8, 34.7, 33.8, 31.4, 30.6, 23.9, 23.1, 22.5, 18.2 ppm.
CDCl3):
d
¼ 164.1, 161.6, 151.7, 151.6, 134.4, 133.2, 129.2, 129.0, 125.5,
125.4, 116.1, 115.9, 106.9, 100.4, 84.0, 83.7, 83.6, 82.4, 39.9, 39.4, 34.7,
31.4, 30.8, 23.0, 22.6, 18.2 ppm.
4.2.15. Data for [18]Cl
Yield: 87 mg (95%). C52H71ClRu2S3$1/8 CH2Cl2 (1040.53): calcd.
C 60. 17, H 6.90; found C 59.03, H 7.02. ESI MS (MeOH): m/z ¼ 995.4
4.3. Cell culture and inhibition of cell growth
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.66 (d, 3J ¼ 8.4 Hz, 2H, S-
p-C6H4 tBu), 7.47 (m, 4H, S-p-CH2C6H4 tBu), 7.41 (m, 4H, S-p-
CH2C6H4 tBu), 7.32 (d, 3J ¼ 8.4 Hz, 2H, S-p-C6H4 tBu), 5.11 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.99 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6H4 iPr), 4.95 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.55 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 3.61 (s, 2H, S-p-CH2C6H4 tBu), 3.42 (s,
Human A2780 and A2780cisR ovarian cancer cells were ob-
tained from the European Collection of Cell Cultures (Salisbury,
UK). Cells were grown routinely in RPMI-1640 medium supple-
mented with 10% foetal calf serum (FCS) and antibiotics at 37 ꢀC
and 5% CO2. Cytotoxicity was determined using the MTT assay
(MTT ¼ 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium
bromide). Cells were seeded in 96-well plates as monolayers with
2H, S-p-CH2C6H4 tBu), 1.81 [sept, 3J
¼
6.8 Hz, 2H, p-
MeC6H4CH(CH3)2], 1.79 (s, 6H, p-CH3C6H4 iPr), 1.37 [s, 9H, S-p-
C6H4C(CH3)3], 1.33 [s, 9H, S-p-CH2C6H4C(CH3)3], 1.32 [s, 9H, S-p-
CH2C6H4C(CH3)3], 0.89 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.86
[d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR
100 mL of cell solution (approximately 20 000 cells) per well and
pre-incubated for 24 h in medium supplemented with 10% FCS.
Compounds were prepared as DMSO solution, then dissolved in
the culture medium and serially diluted to the appropriate con-
(100 MHz, CDCl3):
129.3, 129.2, 125.9, 125.7, 125.6, 125.4, 106.4, 100.9, 84.9, 83.3, 83.2,
82.6, 40.1, 39.6, 34.8, 34.7, 31.4, 31.3, 30.6, 23.2, 22.5, 18.3 ppm.
d
¼ 151.9, 151.7, 151.5, 136.8, 136.7, 134.3, 132.3,
centration to give a final DMSO concentration of 0.5%. 100 ml of
drug solution was added to each well and the plates were incu-
bated for another 72 h. Subsequently, MTT (5 mg mLꢂ1 solution)
was added to the cells and the plates were incubated for a further
2 h. The culture medium was aspirated, and the purple formazan
crystals formed by the mitochondrial dehydrogenase activity of
vital cells were dissolved in DMSO. The optical density, directly
proportional to the number of surviving cells, was quantified at
540 nm using a multiwell plate reader and the fraction of surviving
cells was calculated from the absorbance of untreated control cells.
Evaluation is based on means from two independent experiments,
each comprising three microcultures per concentration level.
4.2.16. Data for [19]Cl
Yield: 82.8 mg (94%). C48H63ClORu2S3$¼ CH2Cl2 (1011.02): calcd.
C 57.32, H 6.33; found C 57.46, H 6.60. ESI MS (MeOH): m/z ¼ 954.9
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 10.33 (s,1H, S-p-C6H4OH), 7.50
(m, 2H, S-p-C6H4OH), 7.48 (m, 8H, S-p-CH2C6Ht4Bu), 7.25 (d,
3J ¼ 8.4 Hz, 2H, S-p-C6H4OH), 5.03 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6Hi4Pr),
4.93 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4i Pr), 4.75 (d, 3J ¼ 6.0 Hz, 2H, p-
MeC6Hi4Pr), 4.62 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6Hi4Pr), 3.58 (s, 2H, S-p-
CH2C6Ht4Bu), 3.40 (s, 2H, S-p-CH2C6H4t Bu), 2.01 [sept, 3J ¼ 6.8 Hz, 2H,
p-MeC6H4CH(CH3)2], 1.73 (s, 6H, p-CH3C6Hi4Pr), 1.40 [s, 9H, S-p-
CH2C6H4C(CH3)3], 1.38 [s, 9H, S-p-CH2C6H4C(CH3)3], 1.02 [d,
3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2], 0.97 [d, 3J ¼ 6.8 Hz, 6H, p-
4.4. Glutathione oxidation
MeC6H4CH(CH3)2] ppm. 13C{1H} NMR (100 MHz, CDCl3):
d
¼ 159.9,
NMR data were acquired at 37 ꢀC using a Bruker Avance II 500-
MHz NMR spectrometer equipped with an inverse dual channel
(1H, X) z-gradient probe head (broadband inverse) or using a Bruker
Avance II 400-MHz NMR spectrometer equipped with an inverse
dual channel (1H, X) z-gradient probe head (broadband inverse).
One-dimensional 1H NMR data were acquired with 16e64 tran-
sients as 32 768 data points over a width of 12 ppm using a classical
presaturation to eliminate the water resonance. A relaxation delay
of 6 s was applied between the transients. All NMR data were
processed using Topspin (version 2.1 or 3.0, Bruker, Switzerland).
151.8, 151.7, 136.8, 136.6, 133.3, 129.2, 129.0, 125.5, 125.4, 124.1, 117.1,
107.3, 100.1, 83.9, 83.5, 83.4, 82.1, 39.7, 39.0, 34.8, 34.7, 31.4, 30.9,
23.1, 22.8, 18.0 ppm.
4.2.17. Data for [20]Cl
Yield: 77.7 mg (83%). C48H62BrClRu2S3$½ CH2Cl2 (1095.15): calcd.
C53.19, H 5.80; found C 52.99, H 5.95. ESI MS (MeOH): m/z ¼ 1017.8
[M]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.72 (m, 2H, S-p-C6H4Br), 7.50
(m, 8H, S-p-CH2C6H4 tBu), 7.48 (m, 2H, S-p-C6H4Br), 5.17 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 5.08 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr),
4.96 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.66 (d, 3J ¼ 6.0 Hz, 2H,
p-MeC6H4 iPr), 3.64 (s, 2H, S-p-CH2C6H4 tBu), 3.46 (s, 2H, S-p-
CH2C6H4 tBu),1.94 [sept, 3J ¼ 6.8 Hz, 2H, p-MeC6H4CH(CH3)2],1.79 (s,
6H, p-CH3C6H4 iPr), 1.39 [s, 9H, S-p-CH2C6H4C(CH3)3], 1.36 [s, 9H, S-
p-CH2C6H4C(CH3)3], 0.97 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2],
0.92 [d, 3J ¼ 6.8 Hz, 6H, p-MeC6H4CH(CH3)2] ppm. 13C{1H} NMR
The 1H
d scale was referenced to the residual water signal at
4.637 ppm (37 ꢀC).
To evaluate the catalytic performance of the complexes for the
oxidation of the reduced form of GSH to GSSG, the complexes
(approximately 0.2 mM) were dissolved in D2O/DMSO-d6 (99:1) and
100 equiv of GSH was added to the solution. The samples were
subsequently analyzed by 1H NMR spectroscopy. For the six com-
plexes, the 1H NMR spectra were recorded immediatelyafter sample
preparation, and then every 30 min until complete disappearance of
the original resonances of GSH. The TOF50 values were obtained
from each catalytic run by fitting the turnover numbers (TON) as a
function of time with the exponential expression Y ¼ a ꢂ bcx for all
complexes. The turnover numbers were calculated according to the
following equation: {IGSSG/(IGSH þ IGSSG)} ꢃ {[GSH]0/[complex]},
where IGSSG and IGSH are the integral intensities of the signals of
GSSG and GSH respectively. The turnover frequencies were obtained
as a derivative of the fitting function for x ¼ 2 (after 2 h incubation
(100 MHz, CDCl3):
129.1, 125.6, 125.4, 122.6, 107.2, 100.4, 83.9, 83.8, 83.7, 82.4, 40.0,
39.5, 34.8, 34.7, 31.4, 30.9, 23.0, 22.7, 18.2 ppm.
d
¼ 151.8, 151.6, 137.4, 136.6, 134.2, 131.9, 129.3,
4.2.18. Data for [21]Cl
Yield: 78.5mg(89%). C48H62ClFRu2S3$1/3 CH2Cl2 (1020.1): calcd. C
57.64, H 6.32; found C 57.43, H 6.74. ESI MS (MeOH): m/z ¼ 957.6
[M þ H]þ. 1H NMR (400 MHz, CDCl3):
d
¼ 7.75 (m, 2H, S-p-C6H4F),
7.41 (m, 8H, S-p-CH2C6H4 tBu), 7.00 (m, 2H, S-p-C6H4F), 5.08 (d,
3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr), 4.99 (d, 3J ¼ 6.0 Hz, 2H, p-MeC6H4 iPr),

48
F. Giannini et al. / Journal of Organometallic Chemistry 744 (2013) 41e48
[12] M.-G. Mendoza-Ferri, C.G. Hartinger, R.E. Eichinger, N. Stolyarova, K. Severin,
M.A. Jakupec, A.A. Nazarov, B.K. Keppler, Organometallics 27 (2008) 2405e
2407.
corresponding approximately to 50% conversion of GSH to GSSG)
and are reported considering only one ruthenium atom.
[13] M.-G. Mendoza-Ferri, C.G. Hartinger, A.A. Nazarov, W. Kandioller, K. Severin,
B.K. Keppler, Appl. Organomet. Chem. 22 (2008) 326e332.
[14] M. Gras, B. Therrien, G. Süss-Fink, O. Zava, P.J. Dyson, Dalton Trans. 39 (2010)
10305e10313.
Acknowledgments
This work was financially supported by the Swiss National Sci-
ence Foundation (projects no 200020-131844 and 200020-143254)
and by the Berne University Research Foundation.
[15] F. Giannini, G. Süss-Fink, J. Furrer, Inorg. Chem. 50 (2011) 10552e10554.
[16] F. Giannini, J. Furrer, A.F. Ibao, G. Süss-Fink, B. Therrien, O. Zava, M. Baquie,
ꢀ
ꢀ
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P.J. Dyson, P. Stepnicka, J. Biol. Inorg. Chem. 17 (2012) 951e960.
[17] A.F. Ibao, M. Gras, B. Therrien, G. Süss-Fink, O. Zava, P.J. Dyson, Eur. J. Inorg.
Chem. 2012 (2012) 1531e1535.
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