Reduction of some Pt(iv) complexes with biologically important sulfur-donor ligands

Article abstract of DOI:10.1039/c3dt50751c

The reduction of the Pt(iv) complexes [PtCl₄(bipy)], [PtCl ₄(dach)] and [PtCl₄(en)] by glutathione (GSH), l-cysteine (l-Cys) and l-methionine (l-Met) was investigated by stopped-flow spectrophotometry at pH 2.0 (in 0.01 M perchloric acid) and at pH 7.2 (in 25 mM Hepes buffer). Kinetic measurements were performed under pseudo-first order conditions with an excess of the reducing agent. The order of the reactivity of the studied complexes was [PtCl₄(bipy)] > [PtCl₄(dach)] > [PtCl₄(en)], and reactivity of investigated reducing agents followed the order GSH > l-Cys > l-Met. All the reactions between the selected Pt(iv) complexes and the sulfur donor biomolecules proceeded by a reductive elimination process that included nucleophilic attack by the reducing agent on one of the mutually trans-coordinated chloride ligands, which led to a two-electron transfer process. The final products of the redox reactions were the corresponding reduced Pt(ii) complexes and the oxidized form of the reducing agents.

Full text of DOI:10.1039/c3dt50751c

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To be submitted to Dalton Transactions
Reduction of some Pt(IV) complexes with biologically important sulfurꢀ
donor ligands
Snežana Jovanović,a Biljana Petrović,a Živadin D. Bugarčića* and Rudi van Eldikb*
a Department of Chemistry, Faculty of Science, University of Kragujevac, R. Domanovića
12, P. O. Box 60, 34000 Kragujevac, Serbia
b Inorganic Chemistry, Department of Chemistry and Pharmacy, University of
ErlangenꢀNürnberg, Egerlandstrasse 1, 91058 Erlangen, Germany
* Corresponding authors: Prof. Dr. Živadin D. Bugarčić
Department of Chemistry, Faculty of Science,
University of Kragujevac
R. Domanovića 12, P. O. Box 60
34000 Kragujevac, Serbia
Tel: +381(0)34300262
Fax: +381(0)34335040
eꢀmail: bugarcic@kg.ac.rs
Prof. Dr. Rudi van Eldik
Inorganic Chemistry
Department of Chemistry and Pharmacy
University of ErlangenꢀNürnberg, Egerlandstrasse 1,
91058 Erlangen, Germany
Tel: +49ꢀ9131ꢀ8528383
Fax: +49ꢀ9131ꢀ8527387
eꢀmail: vaneldik@chemie.uniꢀerlangen.de
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Abstract
The reduction of the Pt(IV) complexes [PtCl4(bipy)], [PtCl4(dach)] and [PtCl4(en)] by
glutathione (GSH), Lꢀcysteine (LꢀCys) and Lꢀmethionine (LꢀMet) was investigated by
stoppedꢀflow spectrophotometry at pH 2.0 (in 0.01 M perchloric acid) and at pH 7.2 (in
25 mM Hepes buffer). Kinetic measurements were performed under pseudoꢀfirst order
conditions with an excess of the reducing agent. The order of the reactivity of the studied
complexes was [PtCl4(bipy)] > [PtCl4(dach)] > [PtCl4(en)], and reactivity of investigated
reducing agents followed the order GSH > LꢀCys > LꢀMet. All the reactions between the
selected Pt(IV) complexes and the sulfur donor biomolecules proceeded by a reductive
elimination process that included nucleophilic attack by the reducing agent on one of the
mutually transꢀcoordinated chloride ligands, which led to a twoꢀelectron transfer process.
The final products of the redox reactions were the corresponding reduced Pt(II)
complexes and the oxidized form of the reducing agents.
Keywords: Pt(IV) complexes; reduction; glutathione; Lꢀcysteine; Lꢀmethionine
2

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Introduction
It is well known that the clinical efficiency of cisplatin, cisꢀ[PtCl2(NH3)2], is limited by
many toxic side effects, such as doseꢀlimiting nephrotoxicity, drug resistance of the
tumor cells and a narrow range of activity.1 Further studies in this field revealed a number
of different Pt(II) complexes, structurally similar to cisplatin, with reduced toxicity, but
still not suitable for longer application.2ꢀ4 To date, about 3000 different platinum
complexes have been synthesized and investigated in an attempt to improve the antitumor
activity, to lower the toxicity and to design a drug that is able to overcome cell resistance.
However, only about 30 platinum complexes have hitherto entered clinical trials.5ꢀ7
Significant among them are Pt(II) complexes with a transꢀgeometry, dinuclear Pt(II)
complexes and Pt(IV) complexes.1,2,4
Platinum(IV) complexes have shown considerable promise as antitumor agents.
Their octahedral geometry introduces two additional ligand sites, and the high kinetic
inertness of the complexes lowers their reactivity as well as the prospect of side reactions.
Thus, the potential advantages of Pt(IV) complexes are based on the fact that they remain
in the higher oxidation state in the bloodstream due to their lower reactivity. This can
diminish the loss of the active drug and lower the number of side reactions that lead to
toxic side effects.8ꢀ10
Compared to the Pt(II) complexes, some complexes of Pt(IV) can be orally
administrated since the amount of the complex that is lost or deactivated on the way to
the target cells is relatively small. For example, satraplatin is the first orally applicable Pt
drug in a clinical trial.11 On the other hand, the lipophilicity of the Pt(IV) complexes is an
important factor for their oral administration.8,12
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Pt(IV) complexes are inert in ligand substitution reactions relative to their Pt(II)
analogues.13ꢀ15 Upon entering into the cell, there are two metabolic pathways for Pt(IV)
complexes, viz. reduction by agents present in the cell, such as glutathione, Lꢀcysteine, Lꢀ
methionine and ascorbic acid, or direct interaction with DNA molecules in the cell
nucleus. The first pathway leads to the wellꢀknown reactions of Pt(II) complexes,16
whereas the second pathway involves the formation of an adduct between Pt(IV) and
DNA.17,18
Elding and coꢀworkers19ꢀ23 studied in detail the reduction of various Pt(IV)
complexes,
such
as
trans,trans,transꢀ[PtCl2(OH)2(NH3)(cha)],
cis,trans,cisꢀ
[PtCl2(OH)2(NH3)(cha)], transꢀ[PtCl4(NH3)(thiazole)], transꢀ[PtCl4(NH3)(cha)], cisꢀ
[PtCl4(NH3)(cha)], cisꢀ[PtCl4(NH3)2], and trans,cis,cisꢀ[Pt(OCOCH3)2Cl2(NH3)(cha)],
where cha = cyclohexylamine, with different thiols and ascorbic acid as reducing agents.
The most of these reactions followed an innerꢀsphere reductiveꢀelimination mechanism
that involved attack of the reducing agent on the halide coordinated trans to the good
leaving group, leading to a twoꢀelectron transfer process. The final products of these
reactions were the corresponding reduced Pt(II) complexes and the oxidized form of the
reducing agents.
Although most evidence indicates that Pt(IV) compounds are reduced to Pt(II) by
potential cellular reducing agents, the underlying reaction mechanism is still poorly
understood. The relation between the structures of the Pt(IV) complexes and the rates and
mechanisms of their reduction should be strongly connected. Published results for the
reduction of different Pt(IV) complexes showed that their reactivity depended on their
reduction potential.8,17,18,24,25 For diammine complexes of platinum, the structure
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variation of the diammine ligand had less effect on the reduction potential, whereas the
nature of the axial ligands exerted a stronger influence.21,22,26 When the axial ligands were
chloride, the reduction was very fast when compared to those of carboxylate or hydroxide
complexes.27
In this study, the reactions of [PtCl4(bipy)], [PtCl4(dach)] and [PtCl4(en)] with
glutathione (GSH), Lꢀcysteine (LꢀCys) and Lꢀmethionine (LꢀMet) were investigated by
stoppedꢀflow spectrophotometry. The influence of acidity on the rate of reduction of the
Pt(IV) complexes could reveal more information on the relationship between their
structure and chemical behavior due the interaction with the selected biomolecules. The
structures of the studied complexes and reducing agents are presented schematically in
Fig. 1.
Fig. 1
Results and discussion
The redox reactions of [PtCl4(bipy)], [PtCl4(dach)] and [PtCl4(en)] with GSH, LꢀCys and
LꢀMet were studied at pH 2.0 (0.01 M HClO4) in the presence of 0.2 M NaCl, by
following the changes in absorbance at suitable wavelengths as a function of time.
Singleꢀexponential traces were obtained under all experimental conditions. The
calculated values for the pseudoꢀfirst order rate constants, kobsd, are given in Tables S1ꢀS3
(ESI). The suggested pathways for the reduction of the selected Pt(IV) complexes by the
thiols and thioether at pH 2.0 are presented in Scheme 1. In the first step, the reductive
elimination by RSH results in the production of RSCl, which hydrolyzes rapidly in the
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subsequent step to form RSOH, which in turn reacts with another RSH molecule to form
the final product RSSR.21 The first step of this reaction sequence is the rateꢀdetermining
process.
k2
[PtCl2(NꢀN)] + RSCl + H+ + Clꢀ
slow
[PtCl4(NꢀN)] + RSH
RSCl + H2O
RSOH + H+ + Clꢀ
RSSR + H2O
fast
RSOH + RSH
k2
[PtCl2(NꢀN)] + MeS(O)R + 2H+ + 2Clꢀ
[PtCl4(NꢀN)] + MeSR + H2O
NꢀN = bipy, dach, en; RSH = GSH, LꢀCys; MeSR = LꢀMet
Scheme 1
The dependence of the observed rate constants, kobsd, on the concentration of the reducing
agents can be express by Eqn. (1), where k2 represents the secondꢀorder rate constants for
the rateꢀdetermining step of all reaction systems:
kobsd = k2[Reducing agent]
(1)
All plots of kobsd vs. reducing agent concentration were linear and passed through the
origin. This indicates that possible parallel or back reactions are insignificant or absent
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(see Fig. 2, Figs. S1 and S2 (ESI)). The secondꢀorder rate constants (k2) obtained from
the slope of the observed plots are summarized in Table 1.
Fig. 2
Table 1
The obtained results clearly show that the reactivity of the studied complexes followed
the order: [PtCl4(bipy)] > [PtCl4(dach)] > [PtCl4(en)]. The order of reactivity of the
selected reducing agents was: GSH > LꢀCys > LꢀMet.
To determine the influence of added chloride on the reduction rate at
pH 2.0, the same reactions were investigated in the absence of NaCl. The calculated rate
constants are also given in Table 1, whereas the linear plots of kobsd vs. reducing agent
concentration are shown in Figs. S3ꢀS5 (ESI). The values of kobsd are given in Tables
S4ꢀS6 (ESI). It was found that the rate constant for reduction, k2, was independent of the
chloride concentrations, which is in agreement with previously published results.23
Since Pt(IV) compounds are in general inert toward substitution, reduction of
Pt(IV) complexes by thiols and thioether occurs by the attack of a sulfur atom on one of
the trans coordinated ligands (innerꢀsphere mechanism). The reduction of Pt(IV) halide
compounds by reducing agents follows a reductive elimination process. It was proposed
that this mechanism involves the attack of the reductant on one of the mutually trans
coordinated chlorides in the Pt(IV) complex, which leads to a twoꢀelectron transfer via a
chlorideꢀbridged complex. Furthermore, the reduction is followed by loss of two trans
chloride ligands and the formation of the corresponding Pt(II) complex.8,20,21,28,29 The rate
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of the reduction of Pt(IV) complexes strongly depends on the type of ligands in the axial
and equatorial positions. The axial bridging ligands are usually halide ions, which
correlates with their electronegativity. More electronegative ligands promote
destabilization of the Pt(IV) complexes, which resulted in a faster reduction process.26 In
the case of complexes that do not have axial chloride ligands but some others, such as
acetate ligands, the reduction was slower and followed an outerꢀsphere mechanism.22,28
On the other hand, bulky inert equatorial ligands destabilized sixꢀcoordinated Pt(IV)
complexes and allowed a faster reduction to the fourꢀcoordinated Pt(II) complexes.26,30
This was confirmed by the results obtained in the present study since ethylenediamine is
less bulky than the other two chelates, bipy and dach, and the reduction of the [PtCl4(en)]
complex was the slowest, whereas [PtCl4(bipy)] showed the fastest reduction rate.
The rate of reduction of Pt(IV) complexes depended on their cathodic reduction
potential.17,18,24ꢀ26 Namely, the cathodic potential depends on the electronꢀwithdrawing
power of the axial ligands as well as on the bulkiness of the inert equatorial ligands. The
published value for the cathodic potential for [PtCl4(en)] was ꢀ160 mV and was ꢀ90 mV
for [PtCl4(dach)].26 This trend is in good agreement with the obtained order of reactivity
in the studied case, since the higher value for the cathodic redox potential should
correlate with a faster reduction process.8,26
The lowest reactivity observed for the thioether (LꢀMet) of all studied reducing
agents can be accounted for since thioethers are in general less powerful reducing agents
than thiols. Furthermore, another possible reason is the steric hindrance of the methyl
group, which is directly linked to the sulfur atom.8
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The reduction of [PtCl4(bipy)], [PtCl4(dach)], [PtCl4(en)] with the selected
biomolecules was also investigated at pH 7.2 (25 mM Hepes buffer) in the presence of
0.2 M NaCl by stoppedꢀflow spectrophotometry at 310 K. Singleꢀexponential traces were
obtained for all the reactions. The calculated values for the pseudoꢀfirst order rate
constants, kobsd, are reported in Tables S7ꢀS9 (ESI). A linear dependence of kobsd on the
reducing agent concentration was observed for all studied reactions (Fig. 3, Figs. S6 and
S7, ESI), which can be described by Eqn. (2).
Fig. 3
k
obsd = k1 + k2[Reducing agent]
(2)
From the slopes of the observed lines, the secondꢀorder rate constants, k2, for the redox
reactions were calculated, see Table 1. The obtained results showed the same order of
reactivity for the studied complexes and reducing agents at pH 7.2 as was observed for
the reactions at pH 2.0. However, there was a significant difference in that all the
observed dependencies at pH 7.2 did not pass through the origin and showed significant
intercepts (k1). In order to investigate this observation further, the reactions between the
studied complexes and glutathione were studied at different chloride concentrations. The
values for the pseudoꢀfirst order rate constants obtained under such conditions are
reported in Tables S10–S12 (ESI), and the plots of kobsd as a function of glutathione
concentration for the [PtCl4(bipy)] complex in the presence of different concentrations of
chloride are reported in Fig. S8 (ESI). The values of the intercept (k1) and slope (k2) for
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the studied reactions as a function of chloride concentration are summarized in the Table
2. Both these rate constants increased strongly with increasing chloride concentration.
The dependences of the rate constants k1 and k2 on the chloride concentration are shown
in Figs. S9 and S10 (ESI), respectively, from which it follows that both rate constants
depended linearly on the chloride concentration. This suggests that the higher chloride
concentration mediates the reductive elimination process via both reaction paths
represented by k1 and k2. Thus, Eqn, (2) can be extended to Eqn. (3) for the reaction
between [PtCl4(bipy)] and glutathione, where k3 and k4 were determined from the slopes
of the plots in Figs. S9 and S10 and had the values (2.4 ± 0.1) × 102 Mꢀ1 sꢀ1 and (5.7 ±
0.5) × 103 Mꢀ2 sꢀ1, respectively.
Table 2
kobsd = k3[Clꢀ] + k4[Clꢀ][glutathione]
(3)
The proposed pathway for the reaction at pH 7.2 is shown in Scheme 2.
k2
ꢀ
[PtCl3(H2O)(NꢀN)]+ + RSꢀ + Clꢀ
[PtCl2(NꢀN)] + RSCl + Cl + H2O
slow
RSCl + H2O
RSOH + RSꢀ
RSOH + H+ + Clꢀ
RSSR + OHꢀ
fast
k2
[PtCl2(NꢀN)] + MeS(O)R + 2Clꢀ + 2H+
[PtCl3(H2O)(NꢀN)]+ + MeSR + Clꢀ
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where:
[PtCl3(H2O)(NꢀN)]+ + Clꢀ
[PtCl4(NꢀN)] + H2O
Cl
Pt
Cl
Cl
Pt
Cl
Cl
Cl
N
N
Cl
Cl
Cl
Cl
Cl
Cl
N
N
N
k1
N
N
+ Clꢀ
Pt
+
+
Pt
N
Cl
Cl
Pt
Cl
Cl
Cl
N
N
Cl
Cl
Cl
Cl
Cl
N
N
N
+ Clꢀ
+
Pt
Pt
N
Cl
Cl
N
Cl
Pt
N
Cl
NꢀN = bipy, dach, en; RSꢀ = deprotonated GSH, Lꢀ Cys; MeSR = LꢀMet
Scheme 2
In the first step, the rateꢀdetermining step, the aqua complex, [PtCl3(H2O)(NꢀN)]+ is
present in the system that reacts with chloride ions to give the [PtCl4(NꢀN)] complex.
This complex reacts further with the reducing agents leading to the formation of the
corresponding Pt(II) complexes. The higher chloride concentration in the solution, the
greater is the stability of the [PtCl4(NꢀN)] complex.
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In order to confirm the hydrolysis of the complexes under the investigated
experimental conditions, the change of absorbance as a function of wavelength was
monitored for a series of solutions of the [PtCl4(en)] complex (1 × 10ꢀ3 M) in the absence
of chloride and in the presence of different concentrations of chloride. Three NaCl
concentrations were investigated: 0.05, 0.1 and 0.2 M (Fig. S11, ESI). The spectra of
each solution of the complex was recorded after 1 h, 3 h, 4 h and 1 day, and then
compared. It can be seen that the absorbance significantly decreased when chloride were
absent from the solution and hydrolysis of the [PtCl4(en)] complex was rapid, while
higher concentrations of chloride suppresed the hydrolysis process.
In a separate experiment, an aliquot (3 ml) of the solution of complex
[PtCl3(H2O)(en)]+, was transferred into a cuvvete and UVꢀVis spectrum was recorded.
An appropriate amount of solid NaCl was added in the cuvvete to the concentration
0.2 M NaCl, and UVꢀVis spectrum was recorded, after 1 day (Fig. S12, ESI). Comparing
the spectra it can be seen that the absorbance increases with addition of chloride in a
solution of complex [PtCl3(H2O)(en)]+, and this complex is converted to the [PtCl4(en)].
The appearance of the intercept could be explained by a dinuclear transition state
between Pt(II) and Pt(IV) complexes through chloride as a bridging ligand. The aboveꢀ
mentioned transition state, as a result of a twoꢀelectron transfer between the divalent and
the tetravalent metal ions, the formerly Pt(IV) species loses the two axial chloride
ligands, leading to the reduced Pt(II) species. However, the Pt(II) species will be oxidized
to Pt(IV) species that have two chloride ligands in the axial positions30,31 (Scheme 2). As
the formation of a bridge between complexes of Pt(II) and Pt(IV) occurs through a
chloride ligand, higher chloride concentrations in the solution could facilitate the
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formation of the bridge. The reduction to the Pt(II) complexes is faster at pH 7.2 than at
pH 2.0, which leads to the formation of larger amounts of complex Pt(II). In addition, due
to the faster reduction, the chlorides were released faster at pH 7.2 than at pH 2.0, which
led to higher chloride concentrations in the solution.
The obtained results also showed that the rate of reduction strongly depended on
the acidity of the solution. At pH 7.2, the reductions were about 5ꢀ30 times faster than at
pH 2.0. This observation is connected with the structures of the ligands and their
deprotonation constants. The acid dissociation constants for GSH are pKa1 = 2.05, pKa2 =
3.40, pKa3 = 8.79 and pKa4 = 9.49, for LꢀCys, they are pKa1 = 1.9 pKa2 = 8.1 and pKa3 =
10.9, and for LꢀMet, they are pKa1 = 2.65 and pKa2 = 9.08.32
The employed amino acids and peptide represent different protolytic forms
depending on the pH (Figs. S13ꢀS15, ESI). At pH 2.0, the formation of a bridged
activated complex is very difficult, but at pH 7.2, deprotonation of the thiol group
increased the reaction rate by a factor of between 5 and 30. Elding et al. defined the
reduction rates of Pt(IV) complexes that include all protolytic constants of the thiols.21
Their results for GSH and LꢀCys are in very good agreement with the present data in
Table 1, viz., the reactivity of the thiolates (RSꢀ) was directly related to the proton
basicity, whereas the steric factor had little influence on the reaction rate. The oxidation
of the thioether, LꢀMet, is also favored at higher pH. Transformation of thioether to
sulfoxide occurs through attack of a hydroxyl group on the sulfur atom and further
deprotonation of the obtained product. At pH 2.0, the occurrence of this process is more
difficult, which could be the explanation for the higher reactivity of LꢀMet at neutral pH.
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To confirm that the reduction of the Pt(IV) complexes leads to the formation of
Pt(II) analogues through the reductive elimination pathway, the reaction between
[PtCl4(en)] and GSH was studied by 1H NMR in D2O at 295 K and at pD 6.4 (pD = pH +
0.4). The spectra of (a) free glutathione, (b) oxidized glutathione and (c) the product of
the reduction of [PtCl4(en)] by GSH are summarized in Fig. 4. The formation of GSSG
during the reaction could be verified by the disappearance of the multiple peaks at 3 ppm,
assigned to the CysꢀßCH2 group of GSH, and the appearance of doublet of doublets at
3.06 ppm and 3.36 ppm, typical for free GSSG. In this region, the signal of the
corresponding Pt(II) complex appeared at 3.35 ppm, while the signal at 3.37 ppm was
ascribed to the remaining Pt(IV) complex in solution.
Fig. 4
Experimental
Materials
Potassium tetrachloridoplatinate(II) (K2PtCl4) was purchased from Strem Chemicals. The
ligands ethylenediamine (en) (Merck), (1R,2R)ꢀ1,2ꢀdiaminocyclohexane (dach) (Acros
Organics) and 2,2′ꢀbipyridyl (Aldrich), and the nucleophiles glutathione (Acros
Organics), Lꢀcysteine and Lꢀmethionine (Aldrich) were used without further purification.
Hepes buffer (4ꢀ(2ꢀhydroxyethyl)ꢀ1ꢀpiperazineethanesulfonic acid) was obtained from
Aldrich. All other chemicals, such as NaCl (Lachema), AgClO4 (Aldrich), HClO4
(Reanal), EDTA (Acros Organics) and D2O (Aldrich) were of the highest commercially
available purity. The complexes [PtCl4(bipy)], [PtCl4(dach)] and [PtCl4(en)] were
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prepared according to published procedures.33,34 All solutions were prepared in doubly
distilled water.
The aqua complex, [PtCl3(H2O)(en)]+, was prepared by the addition of 1
equivalent AgClO4 into the solution of the complex [PtCl4(en)] in perchloric acid
(1 × 10ꢀ3 M) at pH 3.0, under stirring in the dark at room temperature, for 2 h. The
precipitated AgCl was filtered off using Milipore filters. The obtained aqua complex was
used as proof for the hydrolysis of the [PtCl4(en)] complex.
Instrumentation
The UVꢀVis spectra were recorded on Shimadzu UV 250 and HewlettꢀPackard 8452A
diodeꢀarray spectrophotometers with thermostated 1.00 cm quartz Suprasil cells. The
kinetic measurements were performed on an Applied Photophysics SX.18MV stoppedꢀ
flow instrument coupled to an onꢀline data acquisition system. Nuclear magnetic
resonance (1H NMR) spectra were acquired on a Varian Geminiꢀ200 spectrometer at 295
K. All chemical shifts are referenced to trimethylsilylpropionic acid (TSP).
Kinetic measurements
The redox reactions were investigated by stoppedꢀflow spectrophotometry at pH 2.0
(0.01 M HClO4 solution) and at pH 7.2 (25 mM Hepes buffer) in the presence of 0.2 M
NaCl. To determine the effect of chloride on the reduction rate, the same reactions were
investigated at pH 2.0 in the absence of chloride. All kinetic measurements were
performed by following the change in absorbance at suitable wavelengths as a function of
time at 310 K. The selected wavelength for each system was determined by recording the
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spectra of the reaction mixture over the wavelength range between 220 and 500 nm and is
given in Tables S1–S9 (ESI). Reactions were initiated by mixing equal volumes of both
solutions of the complex and reducing agent directly in the stoppedꢀflow instrument and
were followed for at least five halfꢀlives of the reaction. Kinetic measurements were
performed under pseudoꢀfirst order conditions (10ꢀ30 fold excess of the reducing agent).
Higher concentrations of the reducing agent could not be used due to differences
observed in the recorded spectra that pointed to a more complex reaction sequence (see
Fig. S16, ESI). The pseudoꢀfirst order rate constants, kobsd, were calculated as the average
value from five to seven independent kinetic runs. Experimental data are reported in
Tables S1–S9 (ESI).
It is known that molecules that contain thiol groups are sensitive to autoxidation
catalyzed by metal ions such as Cu(II) and Fe(III).35 The rate of autoxidation increases
with increasing pH.23 In order to check whether autoxidation had an effect on the present
kinetic measurements, control experiments were conducted for the reactions of the
[PtCl4(bipy)] complex with glutathione at pH 7.2. In these experiments, both the complex
and glutathione solutions contained 0.5 mM EDTA in order to eliminate the possible
catalytic effect of any traces of metal ions. Before the reactions were initiated in the
stoppedꢀflow instrument, solutions of the [PtCl4(bipy)] complex and glutathione were
flushed with nitrogen for at least 30 min in order to remove dissolved oxygen. No
significant discrepancy between the values of kobsd obtained in these experiments and
those performed in airꢀsaturated solutions was observed (see data in Table S7, ESI). This
finding is in agreement with the results reported by Elding et al.23 For the calculation of
the kinetic data, Microsoft Excel and Origin 6.1 programs were used.
16

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Conclusion
The present results show that the reactivity of the studied complexes follows the order
[PtCl4(bipy)] > [PtCl4(dach)] > [PtCl4(en)]. The reactivity depends of the structure of the
spectator ligands, because bulky inert equatorial ligands destabilize sixꢀcoordinated
Pt(IV) complexes and enable faster reduction to the corresponding Pt(II) complex. The
obtained order of reactivity of the investigated ligands is GSH > LꢀCys > LꢀMet. LꢀMet
is a less powerful reductant than the thiols. The steric hindrance of the methyl group
could be a further reason for the lower reactivity.
The rate of reduction strongly depends on the acidity of the solution. At higher pH
values, the process is much faster due to the easier formation of the bridged intermediate
with the deprotonated form of the thiols. In addition, the thioether converts to the
sulfoxide faster in neutral than in acidic medium. At pH 7.2, hydrolysis of the starting
complex occurs and the formed aqua complex, [PtCl3(H2O)(NꢀN)]+, reacts with chloride
ions forming [PtCl4(NꢀN)]. Furthermore, an increase in chloride concentration leads to
increasing of intercepts, due to the facilitated formation of a bridge between Pt(IV) and
Pt(II) complexes. All these results confirm the fact that the reduction of complexes Pt(IV)
involves a twoꢀelectron transfer process and the loss of axial ligands to form the
corresponding squareꢀplanar Pt(II) complex.
17

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Acknowledgements
The authors gratefully acknowledge the financial support from the Ministry of Education,
Science and Technological Development of the Republic of Serbia (Project No. 172011),
the Deutsche Forschungsgemeinschaft and DAAD.
References
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20

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Cl
Cl
Pt
Cl
Pt
N
N
Cl
Cl
NH2
NH2
NH2
NH2
Cl
Cl
Cl
Cl
Pt
Cl
Cl
Cl
[PtCl4(dach)]
[PtCl4(bipy)]
[PtCl4(en)]
SH
O
H
N
HOOC
COOH
N
H
NH2
O
Glutathione (GSH)
H2N
COOH
H2N
COOH
SH
L-Cysteine (L-Cys)
S
CH3
L-Methionine (L-Met)
Fig. 1 Schematic structures of the investigated Pt(IV) complexes and reducing agents.

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8
7
6
5
4
3
2
1
0
kobsd/s-1
GSH
L-Cys
L-Met
103[L]/M
0
5
10
15
20
25
30
35
Fig. 2 Pseudo-first order rate constants plotted as a function of reducing agent (L)
concentration for the reactions of the [PtCl4(bipy)] complex at pH 2.0 (0.01 M HClO4) in
the presence of 0.2 M NaCl at 310 K.

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90
80
70
60
50
40
30
20
10
0
k
obsd/s-1
GSH
L-Cys
L-Met
103[L]/M
0
5
10
15
20
25
30
35
Fig. 3 Pseudo-first order rate constants plotted as a function of reducing agent (L)
concentration for the reactions of the [PtCl4(bipy)] complex at pH 7.2 (25 mM Hepes
buffer) in the presence of 0.2 M NaCl at 310 K.

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a)
b)
c)
Fig. 4 1H NMR spectra recorded in D2O at 295 K and pD 6.4 (pD = pH + 0.4) of a) 4 mM
GSH, b) 2 mM oxidized glutathione (GSSG), c) the final spectrum of the reaction
between 2 mM [PtCl4(en)] and 4 mM GSH.

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Table 1 Second-order rate constants for the reactions of the selected Pt(IV) complexes
with GSH, L-Cys and L-Met at different pH at 310 K.
Complex
pH
GSH
L-Cys
L-Met
k2/M-1s-1
[PtCl4(bipy)]
2.0a
2.0b
7.2c
7.2d
2.0a
2.0b
7.2c
7.2d
210 ± 20
190 ± 20
980 ± 40
10 ± 1
118 ± 6
107 ± 5
350 ± 50
-
50 ± 4
48 ± 5
150 ± 20
-
[PtCl4(dach)]
62 ± 3
57 ± 3
54 ± 1
340 ± 40
-
39 ± 3
35 ± 2
134 ± 5
-
60 ± 3
920 ± 70
1.5 ± 0.5
31 ± 2
[PtCl4(en)]
2.0a
2.0b
20 ± 1
14 ± 1
30 ± 2
19 ± 2
13 ± 1
7.2c
7.2d
870 ± 90
330 ± 40
130 ± 20
1.1 ± 0.1
-
-
a 0.01 M HClO4 in the presence 0.2 M NaCl; b 0.01 M HClO4 in the absence of NaCl; c 25
d
mM Hepes buffer in the presence 0.2 M NaCl; 25 mM Hepes buffer in the absence of
NaCl

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Table 2 The values of the rate constants k1 and k2 for the reactions between the Pt(IV)
complexes and glutathione (Scheme 2) in the presence of different concentrations of
chloride at pH 7.2 and 310 K.
[Cl-]/M
k1/s-1
0.51 ± 0.02
4.9 ± 0.2
k2/M-1 s-1
10 ± 1
In the absence of
NaCl
[PtCl4(bipy)]
With 0.02
With 0.05
With 0.10
With 0.15
With 0.20
114 ± 9
230 ± 40
450 ± 20
760 ± 80
980 ± 40
1.5 ± 0.5
91 ± 7
10.8 ± 0.5
17.7 ± 0.5
25 ± 2
42.8 ± 0.5
0.038 ± 0.005
3.7 ± 0.2
In the absence of
NaCl
[PtCl4(dach)]
[PtCl4(en)]
With 0.02
With 0.20
35.2 ± 0.5
920 ± 70
1.1 ± 0.1
In the absence of
NaCl
0.0034 ± 0.0005
2.8 ± 0.1
27 ± 2
87 ± 8
With 0.02
870 ± 90
With 0.20

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Graphical Contents Entry
Reduction of some Pt(IV) complexes with biologically important sulfurꢀ
donor ligands
Snežana Jovanović,a Biljana Petrović,a Živadin D. Bugarčića* and Rudi van Eldikb*
The reduction of the Pt(IV) complexes [PtCl4(bipy)], [PtCl4(dach)] and [PtCl4(en)] by
glutathione (GSH), L-cysteine (L-Cys) and L-methionine (L-Met) was investigated by
stopped-flow spectrophotometry at pH 2.0 (in 0.01 M perchloric acid) and at pH 7.2 (in
25 mM Hepes buffer).
8
k
obsd/s-1
7
6
5
4
3
2
1
0
GSH
L-Cys
L-Met
103[L]/M
0
5
10
15
20
25
30
35