Modulation of the solubility properties of arene ruthenium complexes bearing stannyl ligands as potential anti-cancer agents

Article abstract of DOI:10.1016/j.jorganchem.2019.04.002

Cleavage of the known ruthenium dimer [RuCl₂(η⁶-C₆H₅OCH₂CH₂OH)]₂ (1), bearing a hydrophilic substituent on the η⁶ coordinated aromatic ring, with the phosphine ligands

Full text of DOI:10.1016/j.jorganchem.2019.04.002

Accepted Manuscript
Modulation of the solubility properties of arene ruthenium complexes bearing stannyl
ligands as potential anti-cancer agents
Clara Berg, Suviti Chari, Kaste Jurgaityte, Alice Laurora, Mateusz Naldony, Frances
Pope, Dario Romano, Thato Medupe, Sharon Prince, Siyabonga Ngubane, Judith
Baumgartner, Burgert Blom
PII:
S0022-328X(19)30128-7
DOI:
https://doi.org/10.1016/j.jorganchem.2019.04.002
JOM 20752
Reference:
To appear in: Journal of Organometallic Chemistry
Received Date: 23 January 2019
Revised Date: 2 April 2019
Accepted Date: 3 April 2019
Please cite this article as: C. Berg, S. Chari, K. Jurgaityte, A. Laurora, M. Naldony, F. Pope, D. Romano,
T. Medupe, S. Prince, S. Ngubane, J. Baumgartner, B. Blom, Modulation of the solubility properties
of arene ruthenium complexes bearing stannyl ligands as potential anti-cancer agents, Journal of
Organometallic Chemistry (2019), doi: https://doi.org/10.1016/j.jorganchem.2019.04.002.
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ACCEPTED MANUSCRIPT
Modulation of the Solubility Properties of Arene Ruthenium
Complexes Bearing Stannyl Ligands as Potential Anti-Cancer
Agents
Clara Berg,a Suviti Chari,a Kaste Jurgaityte,a Alice Laurora,a Mateusz Naldony,a Frances Pope,a
Dario Romano,b,c Thato Medupe,d Sharon Prince,e Siyabonga Ngubane,d,∗ Judith Baumgartner,f
∗∗
Burgert Bloma,
aMaastricht Science Programme, Faculty of Science and Engineering, Maastricht University,
Kapoenstraat 2, PO Box 616, 6200 MD, Maastricht, The Netherlands
bDepartment of Biobased Materials, Faculty of Science and Engineering, Maastricht University,
Brightlands Chemelot Campus, Urmonderbaan 22, Geleen, 6167 RD, The Netherlands
cAachen-Maastricht Institute of Biobased Materials, Brightlands Chemelot Campus,
Urmonderbaan 22, Geleen, 6167 RD, The Netherlands
dUniversity of Cape Town, Department of Chemistry, Rondebosch 7701, Cape Town, South Africa
eUniversity of Cape Town, Department of Human Biology, Observatory 7935, Cape Town, South
Africa
fInstitut für Anorganische Chemie, Technische Universität Graz, Stremayrgasse 9, A-8010 Graz,
Austria
Abstract
Cleavage of the known ruthenium dimer [RuCl (η6-C6H5OCH2CH2OH)]2 (
1
), bearing a hydrophilic substituent on
2
the η6 coordinated aromatic ring, with the phosphine ligands: triphenyl phosphine, triphenyl phosphite, trimethyl
phosphite, and 1,3,5-triaza-7-phosphaadamantane (PTA) afforded the known
C6H5OCH2CH2OH)(PPh3)] 2a),
[RuCl2(η6-C6H5OCH2CH2OH){P(OPh)3}]
C6H5OCH2CH2OH){P(OMe3)}] (2c), and [RuCl2 (η6-C6H5OCH2CH2OH)(PTA)] (
). The reaction of the known
complexes [RuCl2(η6-
2b),
[RuCl2(η6-
(
(
4
complex 2a with SnCl2 afforded, by facile insertion of the SnCl2 moiety into the Ru-Cl bond, the novel complex
[RuCl(η6-C6H5OCH2CH2OH)(PPh3)(SnCl3)] (3a). Similarly, the reaction of complex 2b with SnCl2 afforded the
novel complex [RuCl(η6-C6H5OCH2CH2OH){P(OPh)3}(SnCl3)] (3b). Complexes 3a and 3b were fully characterized
1
31
by spectroscopy (Infrared (IR) -spectroscopy, H, P and 119Sn Nuclear Magnetic Resonance (NMR) spectroscopy,
UV-Vis spectroscopy and high resolution ESI-MS) and their thermal behaviour elucidated by Thermogravimetric
Analysis (TGA). Density Functional Theory (DFT) calculations (Level of theory B3LYP, basis set for H, C, P, O, N
and Cl is 6-31+G(d,p) and for Ru and Sn is DGDZVP) for complex 3a, 3b and 4 were also carried out, in particular to
elucidate the bonding situation between Ru and Sn in complexes. The hitherto unprecedented anti-cancer activity of
the complexes 2a – 2c as well as the novel stannyl complexes 3a and 3b were evaluated against MCF-7 (oestrogen
receptor positive) human breast adenocarcinoma cell lines. All complexes show activity active against MCF-7 cell
lines, indicating potential application as an anti-tumor agent.
Keywords: Ruthenium, Solubility, Stannyl ligands, Arene, Anti-cancer, MCF-7 Human Breast
Adenocarcinoma Cells.
∗Corresponding author. ∗∗Corresponding author. Email addresses: burgert.blom@maastrichtuniversity.nl(Burgert Blom)
siyabonga.ngubane@uct.ac.za (Siyabonga Ngubane),

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1. Introduction
Cis-diamminedichloridoplatinum(II), commonly known as cisplatin, is administered intravenously to
treat patients suffering from numerous types of cancers including testicular, cervical and certain lung
cancers [1, 2]. In an attempt to increase the bioavailability and limit the side-effects of chemotherapeutic
drugs, ruthenium complexes have been investigated as an alternative to platinum [3-8]. A detailed
mechanistic study of the complex [Ru(p-cymene)(NHC)Cl2] (NHC=1,3-bis(4-(tert-butyl)benzylimidazol-
2-ylidene), hasbeenreported andshows antiproliferate activity of EC109 cells is accompanied by up-regulation
of p53 and p21 proteins and a down-regulation of cyclin D1 [9]. The aforementioned proteins are involved
in the cell cycle, which can cause cell death if not properly regulated [9]. Ruthenium has potentially
important pharmacological advantages, including its access to a range of oxidation states (II, III and IV)
[6]. The energy barriers between each state are relatively low, allowing a change in oxidation state under
relevant physiological conditions when inside the cell [6]. Despite the flexible oxidation states, ligand
exchange rates in water are relatively slow and correlate with the time-scale of cellular reproduction [10].
If a ruthenium ion binds to a biomolecular target in the cell, it will most likely remain bound for the
remainder of that cell’s lifetime, which can be crucial for cytotoxic activity [10]. Research has been done
on the cytotoxic activity of ruthenium arene complexes, giving promising results [3, 8]. One such example is
RAPTA-C (Fig. 1) [11-13]. This piano-stool ruthenium arene complex bears a 1,3,5-triaza-7-
phosphaadamantane ligand and presents promise, similar to the drug NAMI-A, another active agent (Fig.
1) [14, 15]. Interesting chlorambucil derivatives of RAPTA-C have also been reported more recently in
the literature [13].
-
H
N
O
S
Cl
Cl
Cl
Cl
HN
Ru
N
Ru
Cl
P
Cl
HN
N
N
N
NAMI - A
RAPTA - C
Figure 1: The drug candidates RAPTA-C and NAMI-A are undergoing clinical trials for the treatment of secondary
tumors [11, 14].
Previous work has shown that tin groups possess cytotoxic properties [16], and the effectiveness of a
tin dichloride (SnCl2) insertion on ruthenium arene complexes facilitates an improvement in cytotoxicity
[17]. In this vein, we have recently reported neutral complexes of the type [RuCl(η6-C H )(PR (SnCl )] (R
)
6
6
-
3
3
= OPh, Ph, OMe), the ionic complexes [Ru(η6-C H )(PPh )(SnCl3)(DMAP)]+BF4 and [(RuCl(η6-
6
6
3
C H )(PPh )(DMAP)]+BF4 ; the latter exhibit promising cytotoxic activity towards the cell lines A2780
-
6
6
3
(human ovarian cancer) and A2780cisR (human ovarian cisplatin-resistant cancer) (Fig. 2) [18]. In
-
particular, [Ru(η6-C6H6) (PPh3)(SnCl )(DMAP)]+BF4 exhibits some cancer cell selectivity. The neutral
3
complexes [RuCl(η6-C H )(PR (SnCl )] were shown to be less cytotoxic and exhibited in some cases low
)
3
6
6
3
cytotoxicity. In a further study, complexes of the type [RuCl(η6-Arene)(PR3)(GeCl3)] were shown to
exhibit very low cytotoxicity towards A2780 which we proposed was linked to the rapid aquation of the
complexes [19]. Possibly, the lower than expected cytotoxicity in the complexes [RuCl(η6-
C6H6)(PR3)(SnCl3)] could be linked to the their extreme insolubility in aqueous or non-polar media.
Hence, solubility in aqueous media seemed to play a distinctive role in the cytotoxic activity: very insoluble

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complexes exhibited low activity, despite having SnCl3 groups which could be expected to enhance
cytotoxicity [18]. Inspired by these findings, and as part of our ongoing investigation into elucidating the
effect of incorporating a stannyl group into the ligand sphere of a Ru (II) arene complex and its effect on
cytotoxicity and cancer cell selectivity, we synthesized a series of both known and novel complexes of the
type [RuClX(η6-C6H5OCH2CH2OH)(PR3)] (X = Cl or SnCl3, R = OMe, Ph, OPh). These complexes bear
a hydrophilic tail on the arene moiety, which might enhance solubility in aqueous media. The cytotoxicity
towards MCF-7 (oestrogen receptor positive) human breast adenocarcinoma cells is reported here.
Compounds are benchmarked against the cisplatin clinical drug (CDDP).
Figure 2: Ruthenium arene complexes exhibiting promising cytotoxic activity towards the cell lines A2780 (human ovarian
cancer) and A2780cisR (human ovarian cisplatin-resistant cancer) [18].
2. Results and Discussion
2.1. Synthesis of the complexes
The precursor selected for this synthesis is the easily accessible ruthenium dimer [RuCl2(η6-
C6H5OCH2CH2OH)]2 (1) [20], which enables facile entryto the known piano-stool complexes 2a, 2b and 2c
through simple ligand cleavage reactions with the appropriate phosphine (Scheme 1) [20, 21]. For the
synthesis of the known complexes 2a and 2b a modified procedure from Lastra-Barreira et al. was used
[20]. Amendments were made by stirring the solution for an extended time period and eliminating the
necessity of column chromatography to afford a simplified procedure with a higher overall yield (77.9 %
and 91.0 % respectively). Subsequent reaction of complexes 2a and 2b with SnCl2 affords the novel
complexes
C6H5OCH2CH2OH){P(OPh)3}(SnCl3)] (3b) respectively by insertion of SnCl2 into the Ru-Cl bond. An
analogous complex to RAPTA-C was synthesized utilizing a PTA ligand, affording the known complex
[RuCl(η6-C6H5OCH2CH2OH)(PPh3)(SnCl3)]
(
3a
)
and
[RuCl(η6-
4
.

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Scheme 1: Overview of complexes synthesized in this study. (i) 2 equiv. PPh3/P(OPh)3/P(OMe)3 in DCM, r.t., 17 h; (ii) 1
equiv. SnCl2 in DCM,reflux,4.5h;(iii) 2 equiv. PTA in DCM : methanol (1:1), 50°C, 30 min.
2.2. Spectroscopic characterization of the novel complexes
2.2.1. [RuCl(η6-C6H5OCH2CH2OH)(PPh3)(SnCl3)] (3a)
31
A downfield shift of 3 ppm can be observed in the P{1H} NMR spectrum of complex 3a in
comparison to the precursor 2a.This is in close analogy to our earlier observations, however, in contrast to
our earlier findings [18], no 119Sn or 117Sn satellites were observed. Due to the insertion of SnCl2 into the
Ru-Cl bond of the precursor, the Ru center is rendered chiral and this results in loss of symmetry of the
1
arene ring: in the H NMR spectrum several resonance signals are observed corresponding to the
asymmetric nature of the arene ring. This is also in close analogy to our earlier reported findings [18, 19].
Indeed, starting from the achiral precursor, 2a, the insertion of SnCl2 induces a chiral center at Ru which
should afford a racemic mixture of both enantiomers (rac-3a). This would be expected to afford one set
of resonance signals in all NMR spectra (we represent, for simplicity in all our schemes, one of these
stereoisomers for 3a and 3b). The IR spectrum of 3a shows a very weak and broad stretching vibration at
3078 cm-1, which confirms the presence of the OH group. TGA shows an onset of 3.84% (29.26 g/mol) at
75 °C which may be due to trace impurities. The greatest mass loss of 14.22% (108.35 g/mol) occurred at
the thermal event at temperatures in the range 321.01-342.26 °C. The weight of the leftover sample at the
temperature of 493.45 °C corresponded to 44.40% (339.30 g/mol) which could be attributed to the Ru metal
center and Sn. The observed decomposition point at 205 °C is in accordance with the thermal event in the
range 128 -237 °C. The high resolution mass spectrum (ESI) of the complex shows a complex
fragmentation pattern, with the base peak corresponding to the loss of the trichlorostannyl ligand from the
parent ion i.e. [M-SnCl3]+. This suggests that the stannyl ligand is somewhat weakly bound to the metal
centre, which is supported by our DFT calculations showing a Wiberg Bond Index of 0.723 (see below).
This fragmentation is in analogy to our previously reported analogous germyl complexes [19].
2.2.2. [RuCl(η6-C6H5OCH2CH2OH)({P(OPh)3}(SnCl3)] (3b)
The chirality at the ruthenium center results in different environments for each proton on the arene ring

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1
visible in the H NMR spectrum in analogy to 3a. The OH group can be assigned to a distinct doublet of
doublets at δ = 4.51 ppm. This is a result of the adjacent diastereotopic CH2 hydrogens which appear to be
in distinct environments. The OCH2 attached to the arene ring appears as an unresolved broad peak (δ =
3.93-3.83 ppm) as a result of complex 3J coupling to each of the diastereotopic H atoms on the adjacent
CH2. When comparing the resonance signal in the 31P{1H} NMR to the precursor 2b a downfield shift (by
7.3 ppm) upon the tin insertion is found. This is comparable to what is observed for complex 3a and its
precursor 2a and our earlier findings [18]. In the 119Sn {1H} NMR spectrum of 3b a doublet resonance
signal is observed at δ = -169.3 ppm confirming the attachment of the SnCl3 moiety to the Ru (II) center,
as the doublet arises from a 2
JSn,P coupling. This confirms additionally, that also in solution, no fluxional
process is occurring, and the SnCl3 is statically bound to the Ru center, as any decomplexation equilibrium
would quench the coupling to P (Fig. 3). In close analogy to 3a, complex 3b exhibits a high resolution mass
spectrum (ESI) which has a complex fragmentation pattern, with one of the high intensity signals
corresponding to the loss of SnCl3 from the parent ion.
Figure 3: 119Sn NMR spectrum of complex 3b
.
In the IR spectrum recorded for this sample a very weak and broad peak at 3000 cm-1 denotes the
presence of the OH group as in complex 3a. The analysis using UV-Vis spectroscopy shows the highest
absorbance at the λmax = 454 nm. When compared to the precursor 2b, a blue shift of 30 nm has been
observed, which is in agreement with the visible colour change. The TGA analysis of complex 3b reveals
the initial mass loss of 3.27 % (26.50 g/mol) at a temperature of 135 °C. The biggest mass decrease of 18.40
% (149.00g/mol)wasrecordedat thethermaleventattemperaturesintherange225.24–339.11°C.
2.2.3 Water solubility
By the addition of a hydrophilic substituent on the arene ligand an improved water solubility of the
complexes was intended. Simple tests were conducted to assess the relative water solubilities of the
complexes. Complexes 3a and
4 were found to be somewhat water soluble at 20 °C (0.03 mg/mL and 10
mg/mL respectively). However, 2b and 3b were found to be insoluble.

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2.2.4 Single Crystal X-Ray Investigation of 3b
Crystals, suitable for single crystal X-ray diffraction analysis, of complex 3b were obtained by slow
evaporation of a dilute sample of 3b in dichloromethane at 7 °C (Fig. 4), providing additional evidence of
SnCl2 insertion into the Ru-Cl bond of its precursor 2b. Complex 3b has features typical of piano-stool
Ru(II) arene complexes of this type (see [18] and references therein). Complex 3b features a Ru-Sn bond
length of 2.5627(19) Å, which is comparable to the complex [(η6-C6H6)RuCl(SnCl3)(P(OPh)3)] (2.5686
Å) reported by us earlier [18]. The complex exhibits a slightly distorted tetrahedral geometry about the Sn
center evidenced by the slight deviations from a perfect tetrahedral geometry: Cl(1)-Sn(1)-Ru(1)
120.30(16), Cl(3)-Sn(1)-Ru(1) 118.9(4), Cl(2)-Sn(1)-Ru(1) 115.34(17). Similarly, the geometry around
the Ru centre can be described as distorted tetrahedral.
Figure 4: ORTEP representation of the X-ray structure of complex rac-3b at the 30 % probability level. Disorder at
Cl3 not represented. Selected bond lengths [Å]: Sn(1)-Ru(1) 2.5627(19), Cl(3)-Sn(1)
2.409(9), Sn(1)-
Cl(2) 2.351(5), Sn(1)-Cl(1) 2.377(7), Ru(1)-P(1) 2.246(5). Selected bond angles [o]: P(1)-Ru(1)-Sn(1)
86.56(13), Cl(4)-Ru(1)-Sn(1) 85.46(14), P(1)-Ru(1)-Cl(4) 85.80(17), Cl(1)-Sn(1)-Ru(1) 120.30(16), Cl(3)-
Sn(1)-Ru(1) 118.9(4), Cl(2)-Sn(1)-Ru(1) 115.34(17). (Other metric parameters found in the supporting
information).
2.3. Cytotoxic testing
The anti-cancer activity of Ru(II) arene complexes (5 & 10 µM prepared in DMSO) on the MCF-7
(oestrogen-receptor positive) human breast adenocarcinoma, was evaluated using the MTT colorimetric
assay (Fig. 5) [22]. In all assays, DMSO diluted in free medium was included as a negative control and
the clinical drug cisplatin (CDDP) was included as a positive control with an IC50 value of 5.47
figure S9), comparable to that reported in literature for the same cell-line (5.75 M) [23]. The % cell
viability against the Ru(II) arene complexes is herein reported in figure 5. Complexes 2a – 2c 3a and
µM (see
µ
,
4

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show activity against the MCF-7 cells, at both concentrations tested with complex 3b showing low
activity. Complex 2b is the most active (<50% cell survival) at 10 µM which may be attributed to the
presence of the triphenyl phosphate ligand. This possibly enhances the solubility of the drug molecule
upon direct contact or interaction with the human albumin serum (HSA) protein enabling its
transportation to the biologically active target at enhanced rates [24]. Compound 4 is suspected to undergo
an aquation reaction in solution to yield its complex cation and the respective counter ion which may account
for the observed trend specifically for this compound. The tin compounds 3a and 3b exhibit some cytotoxicity
as compared with the complexes reported earlier of the type [RuCl(η6-C6H6)(PR3)(SnCl3)] which were
largely inactive. This provides some evidence that attachment of the hydrophilic tail increases the water
solubility and hence the cytotoxic activity.

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Figure 5: MTT cell viability assays for MCF-7 cells treated for 24 hours with Ru arene organometallic complexes 2a-c
,
3a, 3b & 4 prepared at (a) 5 & 10 µM and (b) 0 - 10 µM or vehicle (DMSO) or Cisplatin (CDDP). Graphs show mean cell
viability as a percentage of vehicle control ± SEM for each concentration of complexes determined from three independent
experiments performed in quadruplicate [*p<0.5; **p<0.01; ***p<0.001].
Although compound 2b was insoluble in water, it was soluble in DMSO and is the most active
against the MCF-7 cells in this series. Compound 3b was also insoluble in water and only partially
soluble in DMSO and it showed only some activity against the MCF-7 cells. The hydrophobicity of 3b
may be attributed to the presence of the trichlorostannyl ligand coordinated to the ruthenium metal center,
impeding the solubility of the compound in DMSO. This phenomenon was also observed and reported for
the half-sandwich ruthenium complex, [(η6-C6H6)Ru(SnCl3)2(P(OMe)3)], bearing two trichloro stannyl

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ligand moieties [18], for which the antiproliferative activity was measured against ovarian cancer cells
after 72 hours of continuous drug action.
In the present study, the compound was used after centrifugation of the stock solution to help aid
solubility before dilution with the supplemented RPMI medium. Complexes 2a, 3a and 3b at 10 µM
indicate an increase in the cell viability, possibly due to the uptake limit of the complexes. A multi-dose
cell viability of compounds 3a and 3b is shown in figure 5b indicating an approximate cell death of 30 %
at 8 µM, beyond which an increase in the cell viability is observed.
2.4 Density Functional Calculations
The optimized structures of complexes 3a, 3b and 4 are shown in figure 6.
Figure 6: Density Functional Theory optimized structures for complexes 3a (left), 3b (middle) and
4 (right). Level of
theory B3LYP, basis set for H, C, P, O, N and Cl is 6-31+G(d,p) and for Ru and Sn is DGDZVP.
In all the optimized structures, the Ru atoms show pseudo-tetrahedral geometry. For complexes 3a
and 3b, the Sn atoms also show pseudo-tetrahedral geometry; in addition, the optimized structures of 3a
and 3b are very similar in the arene and SnCl3 moieties. A visual representation of the highest occupied
energy orbitals (HOMO) for complexes 3a, 3b and 4 can be found in figure 7; while a visual representation of
the lowest unoccupied energy orbitals (LUMO) is depicted in figure 8.
Figure 7: Graphical representation of the highest occupied energy orbitals (HOMO) for complexes 3a (left), 3b (middle)
and
4 (right). Level of theory B3LYP, basis set for H, C, P, O, N and Cl is 6-31+G(d,p) and for Ru and Sn is DGDZVP.
For complex 3b, the DFT-optimized structure matches the crystallographic structure obtained by X-
ray analysis within 8 % difference according to selected values of bond length and angles reported in

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table 1.
Table 1: Comparison of the DFT-optimized structure and the X-ray crystallographic data
X-Ray
Bond Angles
DFT optimized
Bond
Lengths
[Å]
Angles
Difference
Lengths Angles
Angles
[Å]
[o]
[o]
(%)
2
3
-1
3
3
Sn1-Cl1
Sn1-Cl2
Sn1-Cl3
Sn1-Ru1
Ru1-Cl4
Ru1-P1
2.377
2.351
2.409
2.563
2.379
2.246
2.432
2.434
2.384
2.638
2.447
2.276
1
Ru1-Sn1-Cl4
Ru1-Sn1-P1
Cl1-Sn1-Cl2
Cl1-Sn1-Cl3
Cl2-Sn1-Cl3
85.46
86.56
96.30
108.00
92.50
85.47
88.77
99.34
101.86
100.77
0
2
3
-6
8
For complexes 3a and 3b, the HOMO is mainly located on the Ru-Sn bond and the Ru and Cl atoms.
This indicates that the oxygen atoms connected to the phosphorous do not significantly influence the
HOMO of complex 3b with respect to complex 3a. For complex 4, the HOMO is mainly located on the
Cl atoms and along the Ru-arene coordination.
Figure 8: Graphical representation of the lowest unoccupied energy orbitals (LUMO) for complexes 3a (left), 3b
(middle) and
DGDZVP.
4 (right). Level of theory B3LYP, basis set for H, C, P, O, N and Cl is 6-31+G(d,p) and for Ru and Sn is
For complexes 3a and 3b the LUMO is mainly located on the Sn-Cl bonds as well as on the Ru, Ru-
Cl bond and delocalized over the arene. Likewise, the analysis of the HOMOs, the LUMOs of the
optimized structures for 3a and 3b are very similar, denoting a negligible effect of the oxygens bound to
the phosphorous on the LUMO. Similar results are obtained for the LUMO of complex
4 where the
LUMO is located on the Ru-Cl bonds and delocalized on the arene. Natural bond analysis (NBO) was
performed to establish the nature of bonds with particular focus on the ruthenium-tin bonds and atoms of

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complexes 3a and 3b. Table 2 summarizes the DFT results of the natural bond order analysis for
complexes 3a and 3b
.
Table 2: Natural bond analysis calculation for complexes 3a and 3b
.
Atom Bond Polariz. s-character p-character d-character Mulliken Wiberg
Charge
Bond
Index
(%)
(%)
(%)
(%)
Complex 3a
27.73
50.47
49.53
23.52
82.12
48.75
0.37
-0.709
0.602
0.723
0.711
Ru
Sn
17.51
Complex 3b
25.93
52.62
47.38
22.87
80.27
51.20
0.41
-0.895
0.648
Ru
Sn
19.32
The calculation results show no significant differences between the bond polarizations and the
contributions of the different atomic orbitals of Ru and Sn of complexes 3a and 3b. Complex 3b shows a
slightly more negative Mulliken charge on the Ru atom most likely due to an increased electron density
generated by the presence of the oxygen atoms directly connected to the P atoms coordinated to the Ru
centre. The Wiberg bond index for the Ru-Sn bond is 0.723 for Complex 3a and 0.711 for complex 3b
,
which is in close agreement with our earlier reported findings for a series of analogous Germyl complexes
[19].
3. Experimental Section
3.1 Materials and methods
All reactions were carried out under nitrogen overpressure. Analysis of all complexes proceeded in air
as the complexes were air and moisture stable. All commercially available chemicals used were obtained
from Sigma-Aldrich and used without further purification. Dimer
1 was prepared according to
Soleimannejad et al. [25]. Complex 2c was prepared following a published procedure from Lastra-
Barreira et al. [20]. For complexes 2a and 2b the same procedure was used with only minor amendments.
Complex 4 was prepared using a procedure reported by Matsina et al. [21]. NMR spectra were recordedona
Bruker Ultrashield 300 or in the case of 119Sn NMR (111.8 MHz): a Varian Unity INOVA 300
spectrometer. IR spectra were recorded on a Shimadzu MIRacle IR (ATR), UV-Vis on a Shimadzu UV
3600, and TGA spectra were recorded on a TGA Q-500 at the University of Maastricht Brightlands
Chemelot Campus, Netherlands. The NMR signals are reported with reference to their solvent residual
signals. The peaks in the IR spectra were reported according to their relative intensities: s: strong; w: weak;
m: medium; vs: very strong; vw: very weak. The high resolution ESI-MS of 3a and 3b was recorded at
the University of Stellenbosch Central Anlytical Facilities on a Waters Synapt G2 spectrometer with a
cone voltage of 15 V.

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X-ray structure determination: For X-ray structure analyses the crystals were mounted onto the
tip of glass fibers, and data collection was performed with a BRUKER-AXS SMART APEX CCD
diffractometer using graphite-monochromated Mo Kα radiation (0.71073 Å). The data were
reduced to F2 and corrected for absorption effects with SAINT [26] and SADABS [27, 28],
0
respectively. Structures were solved by direct methods and refined by full-matrix least-squares
method (SHELXL97 and SHELX2013) [29, 30]. All non-hydrogen atoms were refined with
anisotropic displacement parameters. Hydrogen atoms were placed in calculated positions to
correspond to standard bond lengths and angles. All diagrams were drawn with 30 % probability
thermal ellipsoids and all hydrogen atoms were omitted for clarity.
Crystallographic data for the structure of compounds 3b reported in this paper has been
deposited with the Cambridge Crystallographic Data Center as supplementary publication no.
CCDC 1585436 (3b). Copies of data can be obtained free of charge at:
http://www.ccdc.cam.ac.uk/products/csd/request/. Figures of solid state molecular structures were
generated using Ortep-3 as implemented in WINGX [31] and rendered using POV-Ray 3.6 [32].
3.2 Cell culture
RPMI 1640 medium (Highveld Biological, Lyndhurst, UK) was used to maintain the human breast
adenocarcinoma MCF-7 (oestrogen receptor positive) cell line. The medium was supplemented with 10%
Foetal Bovine Serum (FBS), 100 U/ml penicillin and 100 µg/ml streptomycin. Cells were maintained at
37 °C in a 5 % CO2-95 % air-humidified incubator.
3.3 In vitro cytotoxicity evaluation-MTT assay protocol
The complex solutions were prepared by dissolving compounds in DMSO to give 10 mM stock
solutions which were used on the same day of treatment. MCF-7 cells were seeded in 96 well plates at a
density of 4.5 x 103 cells per well. After 48 hrs, cells were treated for 24 hours with medium (200 µL)
containing the test compounds (0, 5 and 10 µM) or vehicle (10 µM DMSO). The 3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyyltetrazolium bromide (MTT) assay (11465007001, Sigma-Aldrich) [22] was used to
determine the cell viability according to the manufacturer’s instructions. Briefly, MTT (10 µL) solution
was added to each well and the plates were incubated for 4 h at 37 °C. After this period, the solubilizing
buffer (10% SDS in 0.01 M HCl) was added to each well and the plates were incubated overnight at 37
°C. Absorbance (595 nm) measurements were determined for each well. Mean cell viability was
calculated as a percentage of the mean vehicle control. At least three independent experiments in
quadruplicate were performed.
3.4 Statistical analysis
Statistical analysis was conducted with the acquisition of the data represented as mean values and
SEM (standard error of the means) of three independent experiments. The t-test (a student’s statistical
hypothesis test) was used to compare the two experimental frames and a value of p < 0.5 was accepted as
statistically significant.
3.5 Density Functional Theory calculations
DFT calculations were performed to model the complexes 3a, 3b and 4. The Guassian09 software
package was used. The level of theory used for all calculations is B3LYP with the basis set 6-31+G(d,p)
for H, C, O, P, Cl and N atoms; while for Ru and Sn atoms the DGDZVP basis set was used. Geometry

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optimizations were calculated without any constraints. All the optimized geometries show non imaginary
frequency. Energies and Natural Bond Order analyses were conducted on the optimized structures [33-
36].
3.6 [RuCl(η6-C6H5OCH2CH2OH)(PPh3)(SnCl3)] (3a
)
To a solution of 2a (0.300 g, 0.524 mmol) in DCM (35 mL), anhydrous tin dichloride (0.109 g, 0.576
mmol) was added. The mixture was refluxed for 4.5 h. The solution was filtered, and the solvent removed
in vacuo affording a dark red solid. Yield: 0.319 g (0.418 mmol, 79.8 %). 1H NMR (300.1 MHz, DMSO-d6
,
3
296.2 K, ppm) δ 7.66-7.41 (m, 15H, PPh3), 6.40 (t, JH,H = 5.9 Hz, 1H, C
H
), 6.09 (d, 3JH,H = 5.9 Hz,
6 5
5), 5.02 (broad s, 1H, O
2OH). 13C{1H} NMR (75.5 MHz,
DMSO-d6, 296.7 K, ppm) δ 143.4 (d, Cipso, PPh3), 133.9 (d, Cmeta, PPh3), 131.4 (d, Cpara, PPh3), 129.3 (d,
Cortho, PPh3), 77.9 (d, 2JC,P = 9.5 Hz Cmeta
C6H5), 72.2 (s, Cpara 6H5), 62.0 (s, H2O), 59.4 (s, Cortho
6H5) 55.4 (s, 6, 298.2 K,
H2OH) (C1 of C6H5 ring not visible). 31P {1H} NMR (121.5 MHz, DMSO-
1H, C6
4.17 (m, 1H, C6
H
5), 5.94 (d, 3JH,H = 6.3 Hz, 1H, C6
5), 4.12-4.05 (m, 2H, C
H
5), 5.59-5.54 (ps q, 1H, C6
H
H), 4.24-
H
H2O), 3.71 (broad s, 2H, CH
,
,
C
C
,
C
C
d
ppm) δ 33.2 (s). FTIR 3078 (w, broad), 2959 (vvw), 1528 (m), 1464 (w), 1435 (m), 1265 (m, broad), 1092
−1
(m), 1072-999 (w, broad), 910 (w, broad), 845 (w), 800 (w, broad), 746 (m), 692 (s), 667 (m) cm . UV-
◦
◦
◦
Vis (nm) λmax = 376. TGA: (Weight % decrease) 80 C – 128.17 C (3.847 %), 128.17 C – 237.71 (6.69
◦
◦
◦
◦
◦
◦
%), 237.71 C – 283.11 C (6.52 %), 283.11 C – 321.01 C (11.13 %), 321.01 C – 342.26 C (14.22 %),
◦
◦
◦
◦
◦
◦
20
342.26 C – 375.16 C (7.55 %), 375.16 C – 493.45 C (5.70 %). Melting point: 205 C (dec.). S
C
(H O) soluble, 0.03 mg/mL. ESI-MS 537.0331 (found), 537.0342 (calcd.) ([M-SnCl3]+, 100 %).
2
3.7 [RuCl(η6-C6H5OCH2CH2OH){P(OPh)3}(SnCl3)] (3b
)
To a solution of 2b (0.300g, 0.484 mmol) in DCM (35 mL), anhydrous tin dichloride (0.100 g, 0.532
mmol) was added. The mixture was refluxed for 4.5 h. The orange solution was filtered, and the solvent
removed in vacuo affording a viscous red oil. The crude product was left to crystallize from a mixture of
1
◦
diethyl ether and DCM at -25 C obtaining bright red crystals. Yield: 0.212 g (0.261 mmol, 54.0%). H
x
NMR (300.1 MHz, CDCl , 296.2 K, ppm) δ 7.38-7.25 (m, 15H, P(OPh)3), 6.33 (t, JH,H = 5.4 Hz, 1H,
3
C6
H
5), 5.76-5.74 (m, 1H, C6
H
5), 5.60-5.55 (m, 1H, C6
H
5), 4.52 (dd, 3
J
H,H = 2.03 Hz, 3
2O), 3.71 (ps t, 3
J
J
H,H = 3.35 Hz, 1H,
H,H = 6.0
OH
), 4.16-4.10 (m, 1H, C6
H
5), 4.04-3.99 (m, 1H, C6
H
5), 3.93-3.86 (m, 2H, C
H
Hz, 1H, CH2AOH), 2.45 (t, 1H, CH2BOH). C{1H} NMR (75.5 MHz, DMSO-d6, 295.2 K, ppm) δ 150.9
13
(d, 2
J
C,P= 10.0 Hz, Cipso, P(OPh)3), 130.0 (s, Cmeta, P(OPh)3), 125.9 (s, Cpara, P(OPh)3 ), 121.7 (d, 3
J
C,P= 4.4
Hz Cortho, P(OPh)3 ), 94.6 (s, Cmeta
,
C6H5), 92.2 (s, Cpara
,
C
6H5), 72.7 (s,
C
H2O), 72.3 (s, Cortho
,
C
6H5), 60.4
31
(s,
C
H2OH) (C1 of C6H5 ring not visible). P{1H} NMR (121.5 MHz, CDCl , 296.2 K, ppm) δ 123.7 (s,
3
2J119
J117
). 119Sn {1H} NMR (111.8 MHz, CDCl3, 298.2 K, ppm) δ -169.4
P = 896 Hz, 2
P = 849 Hz
Sn,
31
31
Sn,
2
(d, J119
). FTIR 3000 (vw, broad), 1540 (vw), 1485 (vw), 1270 (vw, broad), 1190 (vw),
−1
31
= 974 Hz
P
Sn,
1150 (vw), 1080 (vw), 1020 (vw), 940 (vw), 900 (w, broad), 760 (w), 690 (w) cm . UV-Vis (nm) λmax
◦
◦
◦
◦
= 452. TGA (Weight % decrease) 150 C – 177.74 C (3.27 %), 177.74 C – 225.24 (8.59 %), 225.24 C
◦
◦
◦
◦
◦
– 339.11 C (18.40 %), 399.11 C – 404.81 C (12.89 %), 404.81 C – 485.32 C (14.81 %). Melting
(H O) insoluble. ESI-MS 584.9194 (found), 585.0172 (calcd.) ([M-SnCl3]+,
◦
◦
20
point: 169 C (dec.). S
27 %).
C
2

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4. Conclusion
Two novel half-sandwich Ru(II) compounds bearing trichlorostannyl ligands were successfully
synthesized as potential anti-cancer agents: [RuCl(η6-C6H5OCH2CH2OH)(PPh3)(SnCl3)] (3a) and
[RuCl(
known complexes: [RuCl2(η6-C6H5OCH2CH2OH)(PPh3)] (2a), [RuCl2(η6-C6H5OCH2CH2OH){P(OPh)3}]
2b), [RuCl2(η6-C6H5OCH2CH2OH){P(OMe3)}] (2c), and [RuCl2 (η6-C6H5OCH2CH2OH)(PTA)] (
).
η
6-C6H5OCH2CH2OH){P(OPh)3}(SnCl3)] (3b) and their cytotoxicity compared with the related
(
4
Both new complexes were characterized using IR-spectroscopy, multinuclear NMR spectroscopy, UV-
Vis spectroscopy and TGA as well as high resolution ESI-MS. Complex 3b was also characterized by
single crystal X-ray diffraction analysis. The aim to increase the bioavailability via increasing
hydrophilicity was gauged using a simple water solubility test finding the complexes 3a and
4 to be water
soluble, while 3b was insoluble. Compounds 2(a-c), 3(a) and is somewhat cytotoxic to MCF-7 breast
4
cancer cells at the concentrations tested. The low cytotoxicity of compound 3b is likely due to the only
partial solubility of the compound in DMSO, thus offsetting the compound’s capability to induce cellular
apoptosis. Hence in this study the complexes 2a and 2b are more cytotoxic than their trichlorotin
analogues 3a and 3b, suggesting that despite attachment of a hydrophilic substituent to the η
6-arene ring,
insolubility caused by the trichlorostannyl ligand still might impede cytotoxicity of the latter. An
investigation into the solubility of these compounds in a synthetic buffer like that of the human body may
provide more accurate results.
Acknowledgements
The authors wish to thank the Maastricht Science Programme (MSP) and Maastricht University for
financial support of this research. The University of Cape Town acknowledges financial support from the
National Research Foundation (NRF), Cancer Association of South Africa (CANSA) and the South African
Medical Research Council (SAMRC). We would like to thank Dr. Jade Peres and Mrs. Alexis Neumann-
Mufweba who kindly assisted with the cell culture and cytotoxicity assays described in this study.
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Highlights
•
Modulating solubility of potential anti-cancer agents by attachment of a hydrophilic tail
trichlorostannyl complexes.
•
•
In vitro testing against MCF-7 (oestrogen receptor positive) human breast
adenocarcinoma cell lines
DFT (density functional theory) results.