Accepted Manuscript

Rhodium(III) and iridium(III) pentamethylcyclopentadienyl complexes with tris(2-

carboxyethyl)phosphine, properties and cytostatic activity

Hanna Pruchnik, Małgorzata Latocha, Aleksandra Zielińska, Florian P. Pruchnik

PII: S0022-328X(16)30326-6
DOI: 10.1016/j.jorganchem.2016.08.005
Reference: JOM 19584

To appear in: Journal of Organometallic Chemistry

Received Date: 4 April 2016

Revised Date: 31 July 2016
Accepted Date: 3 August 2016

Please cite this article as: H. Pruchnik, M. Latocha, A. Zielińska, F.P. Pruchnik, Rhodium(III) and
iridium(III) pentamethylcyclopentadienyl complexes with tris(2-carboxyethyl)phosphine, properties and
cytostatic activity, Journal of Organometallic Chemistry (2016), doi: 10.1016/j.jorganchem.2016.08.005.

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 ACCEPTED MANUSCRIPT

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Rhodium(III) and iridium(III) pentamethylcyclopentadienyl complexes with

tris(2-carboxyethyl)phosphine, properties and cytostatic activity.

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Hanna Pruchnika*, Małgorzata Latochab, Aleksandra Zielińskab, Florian P. Pruchnikc
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Department of Physics and Biophysics, Wroclaw University of Environmental and Life

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Sciences, ul. Norwida 25, 50-375 Wrocław, Poland

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b
Faculty of Pharmacy, Medical University of Silesia, ul. Narcyzów 1, 41-200 Sosnowiec,

Poland

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Faculty of Chemistry, University of Wrocław, ul. Joliot-Curie 14, 50-383 Wrocław, Poland

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*Corresponding author. Tel.: +48 71 3205296.

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E-mail address: hanna.pruchnik@up.wroc.pl (H. Pruchnik)

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Abstract

New pentamethylcyclopentadienyl complexes [M(C5Me5)Cl2(TCEP)] (M =Rh 1, M = Ir 2)

with P(C2H4COOH)3 (TCEP) were investigated using IR, 1H, 13

C, 31
P NMR and ESI-MS

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spectroscopies. Geometry optimization in the gas phase at the B3LYP/3-21G** level

indicated that complex 1 has stable pseudooctahedral structure with large HOMO–LUMO

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gap. Calculated and experimental IR spectra of 1 agree very well. Cytostatic activity of

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compounds 1 and 2 was investigated against melanoma and breast tumor cells. Complexes 1

and 2 show very promising activity towards MDA-MB-231 triple negative breast cancer cells.

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Keywords: Rhodium, Iridium, Phosphine, Antitumor, DFT calculations.
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Introduction

The success of cisplatin in chemotherapy has begun a large number of studies of other metal

complexes as anti-tumour drugs [1]. To the most intensely recently investigated complexes

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belong coordination and organometallic compounds of rhodium [2-15], ruthenium [4, 5, 8,

10] and iridium [8-20]. Rhodium and iridium complexes are amongst the most promising

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class of anti-tumor agents. Cytostatic and anti-cancer activity characterize mainly

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coordination of rhodium and iridium in oxidation states +1, +2 and +3 containing ligands with

nitrogen, sulfur and oxygen donor atoms. The antitumor activity of these compounds was

expected because they have d8, d7 and d6 electronic configuration and are isoelectronic with

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Pt(II), Pt(III) and Pt(IV) complexes applied as antitumor agents. Cytostatic activity of

organometallic complexes of platinum metals was investigated to a lesser extent. Interesting

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antitumor properties were found in the case of ruthenium(II) half-sandwich compounds

[Ru(arene)X2(pta)] (RAPTA complexes) (pta=1,3,4-triaza-7-phosphatricyclo[3.3.1.1]decane).

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These results prompted a switch to investigating cytostatic activity of isoelectronic Rh(III)

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and Ir(III) half-sandwich pentamethylcyclopentadienyl complexes [M(C5Me5)Cl2(pta)],

[M(C5Me5)Cl(pta)2]+ [4,5,9] and compounds [M(C5Me5)Cl(chel)] (chel: various bidentate

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ligands with e.g. N-N, N-C and N-O donor atoms) [6-19]. The cytostatic activity of the

Rh(III) and Ir(III) complexes containing pta is comparable with activity of RAPTA
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compounds, although rather low in comparison with [M(C5Me5)Cl(chel)] ( M = Rh, Ir)

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complexes [6,9]. Iridium(III) complexes with (maleimido)pyridocarbazoles show also

antiangiogenic properties. They inhibit tumor cell induced angiogenesis [20].

Tris(2-carboxyethyl)phosphine P(CH2CH2COOH)3 (TCEP) is readily soluble in water and

stable in air. Thus it can be used for preparation phosphine complexes reasonably soluble in

water. It is frequently used in biochemistry as an efficient reducing agent to break disulfide

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bond in peptides, proteins and other compounds containing S-S bond [21-24]. TCEP is now

often used instead of dithiothreitol (DTT), which is not stable in the reduced form for a long

times. Application of TCEP is very convenient, because it is stable in aqueous solutions,

odourless and stoichiometrically and irreversibly reduces disulfides:

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RSSR + P(CH2CH2COOH)3 + H2O → 2RSH + OP(CH2CH2COOH)3

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TCEP forms complexes with transition metals. Coordination compounds with Zn(II) [25],

Ni(II), Cu(II), Zn(II), Cd(II), Pb(II) [26], Co(III) [27] and Fe(I) [28] were investigated and X-

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ray structures of Zn(II), Cd(II), Co(III) and Fe(I) complexes were determined. In all

it is bonded via P
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complexes TCEP is coordinated via P atom and COO- groups except Fe(I) compound, in

which atom. In platinum(II) compound with 3-{di(2-

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methoxyphenyl)phosphanyl}propionate trans-[PtCl{P(C6H4OMe)2(C2H4COOH)-

κP}{P(C6H4OMe)2(C2H4COO)-κCOO,κP}] one phosphine is a chelating ligand bound via P

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atom and COO- group, while the other is terminal ligand coordinated via phosphorus [29].
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We have found that in trans-[PtCl2(TCEP)2] and cis-[PtCl2(TCEP)2] Pt-P bonds are formed,

however, in aqueous solutions trans complex isomerizes to the cis compound and TCEP-s
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coordinate as chelate ligands via P atom and COO- group [30]. In palladium(II) complexes

trans-[PdCl2(TCEP)2] and trans-[Pd2(µ-Cl)2Cl2(TCEP)2] phospine ligands are coordinated via

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P atom. In aqueous solution dinuclear complex gives polymeric compound with bridging
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phosphine ligand [PdCl{P(RCOO-κO-µ-O’)(RCOOH)2-κP}] [31]. Recently TCEP was used

for reactivity studies between cisplatin, oxaliplatin and model proteins. Due to the presence of

redox active cysteine residue, investigations of interaction of proteins with metal complexes

are typically performed in the presence of TCEP as reducing agent. It was found that cisplatin

interacts with Cu(I) transporters ATP7B, Atox1 [32-34]. Recently discovered that TCEP

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significantly promoted the reaction of cisplatin with Sp1 zinc finger protein [35]. In

Pt(Atox1)(TCEP), the Pt(II) atom has square-planar coordination with two S atoms of Cys in

trans coordinating sites and amide N atom of Cys and TCEP coordinated via P atom. [34].

However, rhodium and iridium complexes with TCEP were not obtained and investigated.

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Here we report complexes [Rh(C5Me5)Cl2(TCEP)] and [Ir(C5Me5)Cl2(TCEP)] and their

properties and cytostatic activity against tumor cells.

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2 Experimental

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2.1. Materials and measurements on physical and chemical properties.

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Reagents and solvents (analytical grade) were purchased from the Polish company POCH,
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Sigma-Aldrich and ABCR Gmbh and were used as received. Infrared spectra (KBr pellets and

nujol) were recorded with a Bruker IFS 113v and Bruker 66/s spectrometers, 1H, 13C and 31P
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NMR spectra on a Bruker AMX 300 and Bruker Avance 500 spectrometers. Proton chemical
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shifts (δ) were reported with reference to the residual protons in D2O, d6-DMSO and CD3OD;
13 13 31
C chemical shifts were given with respect to the natural contents of C in d6-DMSO, P
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chemical shifts were reported with reference to the external 85 % H3PO4. The mass spectra

were recorded on a Bruker MicrOTOF-Q spectrometer (Bruker Daltonik, Bremen, Germany),

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equipped with an Apollo II electrospray ionization source with an ion funnel. The mass
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spectrometer was operated in the negative and positive ion modes. The instrumental

parameters were as follows: scan range m/z 250–2000, dry gas–nitrogen, temperature 200 °C,

ion source voltage 4500 V. The spectra of compounds were recorded for H2O, H2O/DMF,

H2O/CH3OH and CH3OH solutions. Before analysis the instrument was calibrated externally

with the Tunemix™ mixture (Bruker Daltonik, Germany) in the quadratic regression mode.

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The stoichiometry of the analysed ions was confirmed by the isotopic patterns. All elemental

analyses were performed with a Vario EL3 CHN analyzer. The IR, NMR and ESI-MS data

are given in Appendix A.

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2.2. Synthesis of the compounds.

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2.2.1. Dichlorido(pentamethylcyclopentadienyl)[(2-carboxyethyl)phosphine]rhodium(III),

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[Rh(C5Me5)Cl2{P(C2H4COOH)3}], 1.

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The mixture of Rh2(C5Me5)2Cl4 (0.5 mmol, 0.309 g) and P(C2H4COOH)3·HCl (1
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mmol, 0.287 g) in dioxane (10 ml) was stirred at room temperature for 3 h. The mixture was

evaporated to dryness and the red product was washed three times with propan-2-ol and dried
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in air at room temperature. Yield 0.475 g, 85 %. Anal. Calc. for C19H30O6PRhCl2: C 40.81, H

5.41, Cl 12.68. Found: C 40.88, H 5.54, Cl 12.95.

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2.2.2. Dichlorido(pentamethylcyclopentadienyl)[(2-carboxyethyl)phosphine]iridium(III),

[Ir(C5Me5)Cl2{P(C2H4COOH)3}], 2.
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The mixture of Ir2(C5Me5)2Cl4 (0.5 mmol, 0.398 g) and P(C2H4COOH)3·HCl (1 mmol,

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0.287 g) in dioxane (20 ml) was stirred at room temperature for 3 h. The mixture was
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evaporated to dryness and the red product was washed three times with propan-2-ol and dried

in air at room temperature. Yield 0.525 g, 81 %. Anal. Calc. for C19H30O6PIrCl2: C 35.19, H

4.66, Cl 10.93. Found: C 35.58, H 4.33, Cl 10.61.

2.3. Cytostatic Activity

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The melanoma cell lines: SK-mel (human, Caucasian, skin, melanoma), SH-4 (melanotic

melanoma), Colo-829 (human, umbilical metastatis, melanoma) and C-32 (amelanotic

melanoma) and breast cell lines: MCF7, T-47D and MDA-MB-231 were used for the

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proliferation assay. The experiments were repeated in triplicate for each tested compound

concentration. Statistical significance was determined using Student’s t-test (p < 0.05 was

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considered statistically significant). The cell concentration was 6000 cells/well on the 96-well

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microplate. The basic culture medium for tumor cells consisted of RPMI-1640 medium with

10 % fetal calf serum (FCS) and 1 % antibiotic–antimycotic solution. The cultured cells were

maintained at 37 0C in humidified air containing 5 % CO2, for 24 h. The next day the media

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were changed to RPMI with 0.2 % FCS and cells were incubated for a further 24 h. The

concentration of FCS was changed two times to get synchronized cell cultures. After the
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preincubation the experimental media composed with RPMI, 5 % FCS and solution of

investigated complex was added. The concentrations of rhodium and iridium compounds in
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the cultures were µM - mM. The stock solution of coordination compound was prepared in
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DMSO. The tested compound was diluted with culture medium to reach the final

concentrations of the complex. The final concentration of DMSO in the cultured cells was
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0.1–1 % (v/v). The control cell culture was incubated in the standard media enriched with 5 %

FCS and adequate concentration of DMSO (0.1–1 %, v/v). After 72 h of incubation, cell
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viability was quantified by a cell proliferation assay (WST-1; Roche, Basel, Switzerland). The
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amount of WST-1-formazan produced was measured at 450 nm and appropriate calculations

were performed as described previously [31,36-38]. The results of cytotoxic activity in vitro

were expressed as ID50 – the dose of compound that inhibits proliferation rate of the tumor

cells by 50% as compared to control untreated cells.

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2.4. Computational details

DFT calculations were performed using the Gaussian 03 package of programs [39] with

B3LYP functional and 3-21G** basis set. The numerical calculations have been performed in

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part at Wrocław Centre for Networking and Supercomputing. Computed results were

analyzed using Chemcraft program (www.chemcraftprog.com).

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3. Results and discussion

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3.1. Properties and structures of compounds

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Complexes
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[Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1) and

[Ir(C5Me5)Cl2{P(C2H4COOH)3}] (2) were obtained in reactions of [Rh2(C5Me5)2Cl4] and

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[Ir2(C5Me5)2Cl4] with tris(2-carboxyethyl)phosphine hydrochloride (TCEP·HCl) at
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stoichiometric ratios. They are soluble in polar organic solvents and stable in air in solid state.

The 1H, 13C and 31P{1H} NMR spectra of 1 and 2 in CD3OD and d6-DMSO (Tables S1 – S3)
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agree with the proposed formulas and indicate that complexes have pseudooctahedral
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structure, assuming that pentamethylcyclopentadienyl ligand occupies three coordination

sites. The methyl protons of the C5(CH3)5 group in d6-DMSO are observed at 1.555 ppm as a
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doublet (JHP = 2.9 Hz) for 1 and at 1.536 ppm as a singlet for 2. The 31P{1H} NMR spectrum
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of 1 in the same solvent exhibits the doublet at 24.93 ppm (JRhP = 142.8 Hz) and the spectrum
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of 2 shows a singlet at -8.11 ppm. This difference (ca 35 ppm) in the values of the P
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chemical shifts for 1 and 2 is expected because the P resonances of the same phosphine

ligands in Ir(C5Me5)X2(PR3) complexes are shifted upfield for 20 – 40 ppm in comparison

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with Rh(C5Me5)X2(PR3) compounds [5,40-42]. The C NMR spectra (Table S3) can be
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assigned on the basis of J(31P-13C) values. Often 1J(31P-13C) is substantial (20 - 40 Hz),
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J(31P-13C) is much smaller (<5 Hz) and 3J(31P-13C) has intermediate values (7 – 15Hz).

Therefore most frequently C(α) and C(γ) signals are observed as multiplets and C(β) as

singlets [43]. For complexes 1 and 2, signals of CH2P groups were observed as doublets at

20.45 ppm (1JPC = 26.9 Hz) and 18.20 ppm (1JPC = 34.0 Hz), resonances of CH2COO groups

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as singlets at 29.26 ppm and 27.48 ppm and signals COOH groups as doublets at 174.95 (3JPC

= 13.8 Hz) and 173.94 (3JPC = 14.6 Hz), respectively. The complexes 1 and 2 exhibit 13
C

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resonances of C5 atoms at 100.78dd (JRhC = 6.5 Hz, JPC = 2.2 Hz) and 91.67d (JPC = 2.1 Hz)

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and CH3 signals at 9.52s and 8.44s. These values are similar to those found in other

[M(C5Me5)X2(PR3)] complexes [41].

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In solutions in the presence of water Cl ligands relatively easily dissociate from the

complexes 1 and 2 and are substituted by O atoms of COO- groups as evidenced by 1H and
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P NMR spectra in D2O, D2O/d6-DMSO solutions (Tables S1 and S2, Figure S1 and S2). In
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the P NMR spectrum of compound 1 in D2O two doublets at 21.89 ppm and 25.66 ppm

(1JRh-P = 144.3 Hz) (intensity ratio = 2:3) and in D2O/d6-DMSO at 22.80 ppm and 26.43 ppm
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(1JRh-P = 144.3 Hz) (intensity ratio = 3:2) were observed. The doublets with greater chemical
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shifts suggest the presence of [Rh(C5Me5)P(C2H4COO)3]- ion and those with less shifts the

presence of [Rh(C5Me5)P(C2H4COOD)2(C2H4COOD)] complex. The ratio of concentration

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of ionic complex to the neutral compound is greater in D2O solution(3/2) than that in D2O/d6-

DMSO (2/3) as expected, because dissociation constants of RCOOH acids in water are
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considerably greater than in non-aqueous solvents, DMSO, DMF etc. [44,45].

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These conclusions for complexes 1 were confirmed by ESI-MS spectra in H2O,

H2O/DMF (3/1) and CH3OH (Tables S4 and S5). The formulae of the analyzed ions were

established by the isotopic patterns. Concentrations of ions containing Cl ligands in water

solution of complex 1, both in the negative as well as in positive modes, are much lower than

those with Rh-P and Rh-O bonds: [Rh(C5Me5)P(C2H4COO)3]- m/z = 485.06 (100 %),

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[Rh2(C5Me5)2P2(C2H4COO)5(C2H4COOH)]- (44 %), [Rh(C5Me5)ClP(C2H4COO)2

(C2H4COOH)]- m/z = 521.03 (8 %), [Rh(C5Me5)P(C2H4COO)(C2H4COOH)2]+ m/z = 487.07,

(11 %), [Rh2(C5Me5)2P2(C2H4COOH)6(H2O)2]2+ m.z = 506.06 (12 %),

[Rh(C5Me5)P(C2H4COO)2(C2H4COOH)Na]+ m/z = 509.05 (100 %),

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[Rh(C5Me5)ClP(C2H4COOH)3]+ m/z = 523.05 (18 %). Concentration of

[Rh(C5Me5)ClP(C2H4COO)2(C2H4COOH)]- ions, as expected, depends on solvent and

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increases in the series: H2O < H2O/DMF < CH3OH (Table S4).
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Both P NMR and ESI-MS spectra show that in solution of complex 2 chloro ligands

dissociate to a lesser extent. In NMR spectra in D2O two 31P NMR signals at -7.11 ppm and -

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7.08 and in D2O/d6-DMSO (1/1) at -7.57 ppm and -7.51 ppm were observed (Table S2). They

[Ir(C5Me5)P(C2H4COO)3]-
probably indicate the
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presence of

[Ir(C5Me5)ClP(C2H4COO)2(C2H4COOH)]- ions. This conclusion was circumstantiated by

and
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ESI=MS spectra of complex 2 in H2O/MeOH (1/1), H2O/DMF (1/1) and MeOH solutions

(Table S6). The most abundant anions in these solutions are [Ir(C5Me5)P(C2H4COO)3]- m/z =
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575,11 (100 % in H2O/MeOH and H2O/DMF and 76 % in MeOH) and

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[Ir(C5Me5)ClP(C2H4COO)2(C2H4COOH)]- m/z = 611.09 (81 % in H2O/MeOH and 100 % in

MeOH). Similar depiction was found in the case of spectra in the positive mode. The greatest
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abundances were found for [Ir(C5Me5)P(C2H4COO)2(C2H4COOH)Na]+ (for H2O/MeOH , m/z

= 599.13, 100 %) and [Ir(C5Me5)ClP(C2H4COOH)3]+ (for H2O/MeOH , m/z = 613.13, 91 %)

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cations. These data indicate that in solutions concentration of iridium complex with chloro
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ligand is substantial, on the contrary to the solutions of rhodium compound (Figure S2).

Substitution of Cl ligands in the water solution of [Rh2(Cp*)2Cl4] and [Ir2(Cp*)2Cl4] does not

proceed, ions containing aqua ligands were not observed (Tables S5 and S7). The most

abundant are [M2(C5Me5)2Cl5]- anions.

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The pseudooctahedral structure of compounds was confirmed by the calculations

carried out in the gas phase (vacuum) for complex [Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1)

using the DFT method at the B3LYP/3-21G** level. Results of the calculation are presented

in Tables S8 and S9 and Figures 1 and S3 showing optimized structure of the complex 1. The

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bond lengths and angles are given in the Table S9 and Figure S3. The P26-Rh29-Cl27, P26-

Rh29-Cl28 and Cl27-Rh29-Cl28 angles are equal to 89.2o, 82.4o and 96.1o, respectively, and

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confirm pseudooctahedral structure. Average C-C distance in pentadienyl ring and the

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average Rh-C distance are equal to 1.448 Å and 2.281 Å, respectively, and Rh-CCp*(center)

distance is 1.921 Å. These values are very similar to those found in [Rh(C5Me5)Cl2(TPA)]

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(TPA = 1,3,5-triaza-7-phosphaadamantane) [5] and [Rh(C5Me5)Cl2(PPh2R)] (R = N-

containing substituent) [40] complexes.

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Figure 1. Optimized structure of [Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1) at the DFT B3LYP level

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using 3-21G** basis set.

Methyl groups are slightly tilted away from the complex, as evidenced by the greater

Rh-CMe(center) distance (2.014 Å) (Tables S8 and S9). The P-Rh-CCp*(center) and Cl-Rh-

CCp*(center) angles are also very similar to those found experimentally for

[Rh(C5Me5)Cl2(TPA)] and [Rh(C5Me5)Cl2(PPh2R)] complexes [5,40].

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To have a better insight into the nature of rhodium-ligands bonds, frontier molecular

orbitals calculated for complex 1 have been examined. The energy levels of frontier molecular

orbitals (Figure 2) show large HOMO–LUMO gap (3.4806 eV). This indicates that the

compound has a high stability. The spatial plots of frontier orbitals are shown in Figures 3, S4

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and S5 and molecular orbital coefficients for the [Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1) from

the DFT B3LYP/3-21G** calculations are given in Table S10. The HOMO orbital is mainly

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composed of the π(Rh-Cp*), π*(Rh-Cl2) and σ(Rh-P), and LUMO and LUMO+1 orbital have

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predominantly π*(Rh-Cp*), σ*(Rh-Cl2) character. The occupied molecular orbitals HOMO–1

to HOMO–3 are predominantly composed of π*(Rh-Cl2), π(Rh-Cp*), while the HOMO-4 and

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HOMO-5 consist chiefly π(Rh-Cp*), π(Rh-Cl2) orbitals and HOMO-7 has π(Rh-Cl2)

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character. The HOMO-6, LUMO+2 and LUMO+3 are localized on the COO groups (Tables

S10 and S11).

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Figure 2. MO diagram for the [Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1) from the DFT B3LYP/3-

21G** calculations.

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Figure 3. Spatial plots (isovalue = 0.03) of selected frontier molecular orbitals of

[Rh(C5Me5)Cl2{P(C2H4COOH)3}] (1).

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3.2. Vibrational spectra AN
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The IR spectrum of complex 1 was calculated in the gas phase (vacuum) at B3LYP/3-

21G** level and analysis of normal modes in terms of internal coordinates was performed
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using the Gaussian 03 package of programs [39]. Interpretation of spectrum was based on the
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analysis of normal modes and the potential energy distribution (PED) analysis [46] of all 171

fundamental vibration modes. Results of PED analysis are given in the Table S12 and the
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numbering of atoms is given in Figure S6. Table S13 shows assignment of IR spectrum of

complex 1. There is very good agreement of calculated with experimental spectrum for
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vibrations between heavy atoms (Table S13, Figure S7). The calculated values of O-H and C-
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H stretching vibration are considerably greater than those experimental. For ν(CH) vibrations

good agreement was obtained after scaling calculated values using factor of 0.9613. In the

case of ν(OH) vibrations scaled values are greater than experimental ones because O-H groups form

hydrogen bonds in the solid state. The M-ligand stretching vibration for complexes 1 and 2 with

considerable contributions of torsion modes were observed in the range 240-430 cm-1. For

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complex 1 the Rh-ligand stretching vibrations assume values: νas(RhCl2) at 251 and 265,

νs(RhCl2P) at 243 and 284, ν(RhC5) at 375 and ν(RhP) at 428 cm-1 (Tables S12 and S13). In

the case of complex (2) these vibrations were observed at: 268 ν(IrCl), 288 ν(IrCl), 298

ν(IrCl), 378 ν(IrC5) and 414 cm-1 ν(IrP). In the spectrum of complex 1, bands at 1451 and

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1551 cm-1 are caused by δ(CH3)+ν(CC)Cp* vibrations, at 1051 and 1129 by νa(CCO)+

νs(COO)+δ(COH)+δ(HCC) vibrations and at 1701 and 1743 cm-1 by νa(COO)+ δ(COH)

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vibrations. The ν(OH) bands observed at 3266 and 3444 cm-1 and in the range 2580-2760 cm-1

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indicate that in solid compounds strong O-H···OC hydrogen bonds are formed. The IR

spectrum of complex 2 is similar to that of compound 1.

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3.3. Cytostatic activity of complexes.
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The pentamethylcyclopentadienyl ligand (Cp*) is a stabilizing ligand for
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organometallic rhodium(III) and iridium(III) complexes. In the complexes of the type [M(η5-
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C5Me5)(L)Cl2], the Cp* ligand is generally inert, while chlorides can be relatively easily
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substituted by other ligands. This enables interaction of [M(η5-C5Me5)(L)Cl2] complexes with

biomolecules and can induce their cytostatic activity towards diverse tumor cells [4-6,9,20].
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Cytostatic activity of compounds 1 and 2 was investigated against melanoma cells:

SK-mel (human, Caucasian, skin, melanoma), SH-4 (melanotic melanoma), Colo-829

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(human, umbilical metastatis, melanoma) and C-32 (amelanotic melanoma) as well as against
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breast cell lines: MCF7, T-47D and MDA-MB-231. Melanoma is not the most common skin

cancer, however, it is much more dangerous if it is not found in the early stages. It causes the

majority of deaths related to skin cancer. Especially metastatic melanoma is a disease difficult

to treat [47]. The 10-year survival rate for patients with this disease is less than 10%. Recent

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investigations improved methods of therapies against melanomas, however, times of survivals

are still short, therefore it is worthwhile to search for new agents against these tumors.

Activity of complexes 1 and 2 against melanoma cell lines is moderate to low with IC50

values of ~170–510 µM. Considerably greater cytostatic activity towards the same melanoma

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cells show complexes [M(η5-C5Me5)Cl(qol)] (M =Rh and Ir, qol = quinolin-8-olate) [6].

However, similar or lower activities toward HT29 colon carcinoma, A549 lung carcinoma,

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and T-47D breast carcinoma cells were found for analogous phosphine rhodium(III) complex

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[Rh(C5Me5)Cl2(pta)] (pta = 1,3,5-triaza-7-phosphatricyclo[3.3.1.1]decane) and Ru(II) and

Os(II) arene complexes with pta, [MCl2(MeC6H4CHMe2-p)(pta)] [5]. Probably greater

cytostatic activities of [M(η5-C5Me5)Cl(qol)] complexes against melanoma cells result from

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intercalation of quinolin-8-olate ligand between nucleotide bases of DNA.

Complexes 1 and 2 show very promising activity towards triple negative breast cancer cells
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MDA-MB-231, IC50 values are equal to 67 µM and 7.8 µM, respectively (Table 1).
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Table 1. Cytostatic activity of complexes 1 (Rh3P) and 2 (Ir3P) against tumor strains.
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Compound IC50, µM

Sk-mel SH-4 Colo-829 C-32 MCF7 T-47D MDA-MB-231

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1 280 170 310 690 450 500 67±7

2 510 300 480 560 370 450 7.8±1

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TCEP·HCl 580 590 550 > 800 800 620 380

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Cis-Pt 40a 123±15 94±9 20.3±3 65±7 30±4 61±7

a
L. Pendyala, P. J. Creaven, Cancer Res. 53 (1993) 5970-5976.

Complex 2 is significantly more active towards these cells than cisplatin and activity of

compound 1 is comparable with that of cisplatin. The cytotoxicity both compounds against

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triple positive breast cancer lines MCF7 and T-47D is considerably lower than towards triple

negative cells, however somewhat greater than that of [Rh(C5Me5)Cl2(pta)]. In the cells, since

in the presence of water chloro ligands are easily substituted by O atoms of carboxylate

groups, most probably ionic complexes containing chelating phospine ligand

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[M(C5Me5){P(C2H4COO)3}]- and [M(C5Me5)Cl{P(C2H4COO)2(C2H4COOH)}]- are

responsible for cytostatic activity.

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4. Conclusions

New pentamethylcyclopentadienyl complexes [M(C5Me5)Cl2{P(C2H4COOH)3}] (M =Rh 1,

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M = Ir 2) with P(C2H4COOH)3 (TCEP) water soluble and air stable phosphine, an efficient
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reducing agent used in biochemistry to break S-S bond in peptides and proteins, were

characterized using IR, 1H, 13C, 31P NMR and ESI-MS spectroscopies. Geometry optimization
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of complex 1 in the gas phase at the B3LYP/3-21G** level indicated that complex 1 has
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pseudooctahedral structure with pentamethylcyclopentadienyl ligand occupying three

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coordination sites. The energy levels of frontier molecular orbitals show large HOMO–

LUMO gap (3.4806 eV) indicating a high stability of 1. The calculated IR spectrum of
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complex 1 at this level agrees very well with experimental one. The IR, 1H, 13C and 31P NMR

confirmed pseudooctahedral structure for both complexes. The ESI-MS spectra indicate that
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in solutions Cl ligands relatively easily dissociate from the complexes 1 and 2 and are
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substituted by O atoms of COO- groups. Dissociation of Cl ligands in water solutions in the

case of complex 1 proceeds more easily than for compound 2 as evidenced by the peaks:

[Rh(C5Me5)P(C2H4COO)3]- m/z = 485.06 (100 %),

[Rh(C5Me5)ClP(C2H4COO)2(C2H4COOH)]- m/z = 521.03 (8 %) and for 2 in H2O/CH3OH

[Ir(C5Me5)P(C2H4COO)3]- m/z = 575.10 (100 %) and

[Ir(C5Me5)ClP(C2H4COO)2(C2H4COOH)]- 611.09 (81 %). Cytostatic activity of compounds 1

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and 2 against melanoma cell lines is moderate to low, however, they are very promising

antitumor agents towards triple negative breast cancer cells MDA-MB-231. Complex 2 is

significantly more active against these cells than cisplatin and activity of compound 1 is

comparable with that of cisplatin.

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Appendix A. Supplementary data

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Supplementary data to this article can be found online at doi:

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Highlights

Complexes [M(Cp)Cl2L] (M = Rh, Ir) with P(C2H4COOH)3 (TCEP), reductant of S-S bond.

Calculation at B3LYP/3-21G** level show pseudooctahedral structure of complexes.

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Calculated and experimental IR spectra of complexes are in very good agreement.

Complexes [M(Cp)Cl2(TCEP)] (M=Rh,Ir) are very active against MDA-MB-231 cancer cells.

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