Mechanistic scrutiny of the oxidations of thiol-containing drugs cysteamine and d-penicillamine by cis-diamminetetrachloroplatinum(IV)

Article abstract of DOI:10.1002/kin.21464

Cysteamine (CA) and d-penicillamine (Pen) are the thiol-containing drugs and good antioxidants. Their reactions with a cisplatin Pt(IV) prodrug cis-diamminetetrachloroplatinum(IV) (cis-[Pt(NH₃)₂Cl₄]) were investigated by use of rapid scan, stopped-flow, and mass spectral techniques. The kinetic traces are biphasic in nature, encompassing a faster reduction of cis-[Pt(NH₃)₂Cl₄] to cisplatin followed by slow substitutions on cisplatin. The reduction reactions were demonstrated to follow overall second-order kinetics over a wide pH range. The observed second-order rate constants versus pH profiles were established at 25.0°C and 1.0?M ionic strength, indicating a huge increase of reaction rate with the increase of pH. However, the oxidations of CA and Pen by cis-[Pt(NH₃)₂Cl₄] displayed different reaction stoichiometric ratios as revealed by the spectrophotometric titration experiments. Accordingly, CA was oxidized to CA-disulfide while Pen-sulfinic acid and Pen-disulfide were identified as the major products in the case of Pen via mass spectral analysis. The above similarities and differences are rationalized in terms of the proposed reaction mechanisms, which encompass similar rate-determining reactions for both CA and Pen, but involve disparate and faster followed-up reactions. Rate constants of the rate determining were derived at 25.0°C and 1.0?M ionic strength. A consequent species reactivity analysis revealed that the species -SCH₂CH₂NH₃⁺ of CA and the species ⁺H₃NCH(COO^?)CMe₂S^? of Pen played a predominant role toward the reduction of cis-[Pt(NH₃)₂Cl₄] from pH 5 to 8, which also is a critical pH region for most of drugs.

Full text of DOI:10.1002/kin.21464

Received: 12 June 2020
DOI: 10.1002/kin.21464
Revised: 17 November 2020
Accepted: 30 November 2020
A RT I C L E
Mechanistic scrutiny of the oxidations of thiol-containing
drugs cysteamine and d-penicillamine by
cis-diamminetetrachloroplatinum(IV)
Li Zhou1
Tiejian Li2,3
Lingli Kong1
Ying Sun2
Hongwu Tian3
Cunxiu Gao2
Chunli Liu1
Guimin Zhang2,3
Tiesheng Shi1
1 College of Chemistry, Chemical
Engineering and Materials Science,
Zaozhuang University, Zaozhuang,
Shandong Province 277160, China
2 National Engineering Technology
Center of Chirality Pharmaceuticals,
Lunan Pharmaceutical Group Co., Ltd.,
Linyi, Shandong Province 276006, China
Abstract
Cysteamine (CA) and d-penicillamine (Pen) are the thiol-containing drugs
and good antioxidants. Their reactions with a cisplatin Pt(IV) prodrug cis-
diamminetetrachloroplatinum(IV) (cis-[Pt(NH3)2Cl4]) were investigated by use
of rapid scan, stopped-flow, and mass spectral techniques. The kinetic traces
are biphasic in nature, encompassing a faster reduction of cis-[Pt(NH3)2Cl4] to
cisplatin followed by slow substitutions on cisplatin. The reduction reactions
were demonstrated to follow overall second-order kinetics over a wide pH range.
The observed second-order rate constants versus pH profiles were established at
25.0◦C and 1.0 M ionic strength, indicating a huge increase of reaction rate with
the increase of pH. However, the oxidations of CA and Pen by cis-[Pt(NH3)2Cl4]
displayed different reaction stoichiometric ratios as revealed by the spectropho-
tometric titration experiments. Accordingly, CA was oxidized to CA-disulfide
while Pen-sulfinic acid and Pen-disulfide were identified as the major products
in the case of Pen via mass spectral analysis. The above similarities and differ-
ences are rationalized in terms of the proposed reaction mechanisms, which
encompass similar rate-determining reactions for both CA and Pen, but involve
disparate and faster followed-up reactions. Rate constants of the rate determin-
ing were derived at 25.0◦C and 1.0 M ionic strength. A consequent species reac-
3 Shandong New Time Pharmaceutical
Co., Ltd., Feixian, Shandong Province
273400, China
Correspondence
Guimin Zhang, NationalEngineering
Technology Center ofChiralityPhar-
maceuticals, Lunan Pharmaceutical
Group Co., Ltd., Linyi276006, Shandong
Province, China.
Email:lnzhangguimin@lunan.cn
Tiesheng Shi, CollegeofChemistry, Chem-
ical Engineering and MaterialsScience,
ZaozhuangUniversity, Zaozhuang 277160,
ShandongProvince, China.
Email:rock@uzz.edu.cn
LiZhouand Tiejian Licontributedequally
to this work.
+
tivity analysis revealed that the species -SCH2CH2NH3 of CA and the species
+H3NCH(COO‒)CMe2S‒ of Pen played a predominant role toward the reduction
of cis-[Pt(NH3)2Cl4] from pH 5 to 8, which also is a critical pH region for most of
drugs.
Fundinginformation
Critical R&DPlans of ShandongProvince
(Major ScientificInnovationProjects),
Grant/AwardNumber: 2019JZZY010516;
Natural Science FoundationofShan-
dongProvince, Grant/AwardNumber:
ZR2019MB015
K E Y WO R D S
cis-[Pt(NH3)2Cl4], cysteamine, d-penicillamine, kinetics, mechanism, reduction
1
INTRODUCTION
of cystinosis; it removes cystine that builds up in cells
of people with the disease.1–3 In addition, it has been in
clinical trials for Parkinson’s disease, malaria, radiation
sickness, neurodegenerative disorders, neuropsychiatric
Among the thiol-containing drugs, cysteamine (CA) bear-
ing the simplest structure was approved for the treatment
Int J Chem Kinet. 2020;1–10.
wileyonlinelibrary.com/journal/kin
© 2020 Wiley Periodicals LLC
1

2
ZHOU et al.
disorders, and cancer treatment.3 Biologically, it has a
function of promoting the transport of l-cysteine into cells
that can be further used to synthesize glutathione, which
is one of the most potent intracellular antioxidants.3 d-
Penicillamine (Pen) is also a thiol-containing drug, but
its thiol group is structurally hindered by the two adja-
cent methyl groups. Pen is a medication used primarily
for the treatment of Wilson’s disease;4,5 it is also used for
people with kidney stones, rheumatoid, and various heavy
metal poisonings.4,5 The anticancer effect of Pen has also
been exploited.6 Moreover, the antioxidative properties of
CA and Pen related to some pharmacological processes
have been pursued as exemplified by the research works
in references7–9; this is not hard to understand since both
CA and Pen are good antioxidants due to the thiol group in
the drugs.
Cl
Pt
Cl
NH2
NH3
Cl
Cl
NH2
NH3
Cl
Cl
Pt
cis-[Pt(NH3)2Cl4]
cis-[Pt(NH3)2Cl2] (cisplatin)
H
NH2
CH3
C
HS-CH2-CH2-NH2
Cysteamine (CA)
C
COOH
HS
CH3
D-Penicillamine (Pen)
Not surprisingly, a number of kinetic and mechanistic
investigations on the oxidations of CA and Pen by various
types of oxidants have been performed in order to have a
better understanding of their antioxidative properties.10–15
These investigations have clearly unraveled that the kinetic
rate laws and the derived reaction mechanisms are totally
diversified, depending largely upon the nature of the
oxidants10–15; and based on the investigations, it is almost
impossible to predict an oxidation mechanism when a new
oxidant is considered. In the last few years, we have been
focusing on the mechanistic elucidations for some redox
reactions concerning a few important drugs.16–20 We thus
performed a detailed kinetic analysis of oxidation reactions
of CA and Pen by cis-diamminetetrachloroplatinum(IV)
(cis-[Pt(NH3)2Cl4]) and divulge the results in the present
work. cis-[Pt(NH3)2Cl4] is an anticancer active Pt(IV) com-
pound and is in fact a cisplatin prodrug since it can be eas-
ily reduced to cisplatin (cis-[Pt(NH3)2Cl2])21–23; its reduc-
tive activation processes by some biologically important
molecules and by an electrochemical approach have been
studied.17,19,23–25 On the other hand, it was reported that
mechanistically, a combination of Pen and platinum anti-
cancer drugs synergistically inhibited tumor growth.26 The
above-mentioned impetuses promoted us to undertake the
present study, enabling us to have a mechanistic scrutiny
on the oxidation reactions and to unravel the mecha-
nistic similarities and differences. The structures of cis-
[Pt(NH3)2Cl4], cis-[Pt(NH3)2Cl2], CA and Pen are shown
in Figure 1.
F I G U R E
1
Structures of cis-[Pt(NH3)2Cl4] and cis-
[Pt(NH3)2Cl2] and of thiol-containing drugs cysteamine and
d-penicillamine
from Sigma–Aldrich (St. Louis, MO, USA). Acetic acid,
sodium acetate, sodium chloride, hydrochloric acid, con-
centrated o-phosphoric acid (ca 85% solution), sodium
dihydrogen-phosphate, disodium hydrogen phosphate,
trisodium phosphate dodecahydrate, sodium bicarbonate,
sodium carbonate, sodium perchlorate, and 0.100 M NaOH
standard solution were purchased as the purest forms
available either from Fisher Scientific (Fisher Branch in
Shanghai, China) or from Alfa Aesar (Alfa Aesar Branch
in Shanghai, China). These chemicals were used directly
without further purification. For pH meter calibrations,
standard buffers of pH 4.00, 7.00, and 10.00 were pur-
chased from Fisher Scientific. Doubly distilled water was
employed for preparations of all the solutions used in this
work.
2.2
Buffer solutions
Buffering pairs of H3PO4/NaH2PO4, HAc/NaHc,
NaH2PO4/Na2HPO4, NaHCO3/Na2CO3, and Na2HPO4/
Na3PO4 (0.15-0.2 M) were prepared to cover a pH range
of 2.47 ≤ pH ≤ 11.81. All the buffer solutions contained
0.10 M of NaCl and 2.0 mM of EDTA; sodium perchlorate
was used to adjust the ion strength (µ) of buffers to 1.0 M.
The addition of NaCl was to suppress the hydrolysis of the
Pt(IV) complex. The role of EDTA in the buffers was to
complex trace amounts of metal ions such as Cu(II) and
Fe(III), eliminating or minimizing the catalytic effects
of the metal ions in the thiol oxidation processes.13,27–29
An Accumet Basic AB150 Plus pH meter equipped with
an Accumet combination pH electrode (Fisher Scientific,
2
EXPERIMENTAL
Materials
2.1
Cysteamine hydrochloride (CA), Pen, cis-[Pt(NH3)2Cl4]),
and cis-[Pt(NH3)2Cl2] in their purest forms were obtained

ZHOU et al.
3
Pittsburgh, PA, USA) was utilized to measure the pH of
buffer solutions; the electrode was calibrated just before
the measurements by use of the standard buffers of pH
4.00, 7.00 and 10.00.
2.3
Rapid scan spectra and kinetic data
collection
An Applied Photophysics SX-20 stopped-flow spectrom-
eter (Applied Photophysics Ltd., Leatherhead, UK) was
employed for kinetic measurements and for recording the
rapid scan spectra. The stopped-flow spectrometer was
connected to a water bath circulation from a thermostat
(Lauda Alpha RA8; Delran, NJ, USA); temperature could
be controlled to 0.1◦C. A stock solution of 1.0 mM cis-
[Pt(NH3)2Cl4] was prepared daily by dissolving the desired
amount of the Pt(IV) complex in a solution containing
0.90 M NaClO4, 0.09 M NaCl, and 0.01 M HCl; these solu-
tions were only used for a few hours. A stock solution of
CA/Pen (20-30 mM) was prepared just before the kinetic
measurements by dissolving a certain amount of CA/Pen
in a specific buffer; the stock solution was bubbled with
nitrogen for 10 min and was only used for a couple of hours.
Solutions of CA/Pen and of the Pt(IV) complex for kinetic
measurements were prepared, respectively, by adding an
appropriate amount of the Pt(IV) stock solution and of
CA/Pen stock solution to the buffer of the same pH. Those
solutions were bubbled for 5 min with nitrogen before load-
ing onto the stopped-flow machine. Reactions were started
by mixing equal volumes of CA/Pen and Pt(IV) solutions
directly in the stopped-flow machine; pseudo first-order
conditions were fulfilled by utilizing CA/Pen being at least
10-fold excess. The software provided by the Applied Pho-
tophysics in the control system was employed to simulate
the kinetic traces.
F I G U R E
2
A kinetic trace acquired at 265 nm for the reac-
tion between cis-[Pt(NH3)2Cl4] and CA by the stopped-flow spec-
trometer under the reaction conditions: [Pt(IV)] = 0.20 mM,
[CA]tot = 2.00 mM, pH 4.52 buffer, 25.0◦C, and 1.0 M ionic strength.
Insert: The kinetic trace is enlarged to show the initial phase between
0 to 50 s; the red curve was resulted from a theoretical simulation of
the experimental data by Equation (1) [Color figure can be viewed at
wileyonlinelibrary.com]
Germany) with an ESI of positive ionization mode. On the
other hand, an Agilent 1200/6310 ion trap mass spectrome-
ter with an ESI was employed for recording the mass spec-
tra for a reaction mixture of 1.0 mM cis-[Pt(NH3)2Cl4] with
8 mM Pen in 10 mM acetic acid after a reaction time of
about 0.5; both positive and negative modes of ionization
were run.
3
RESULTS AND DISCUSSION
Kinetic traces
2.4
Electronic spectra and absorption
measurements
3.1
Electronic spectra and absorption measurements at room
temperature were performed on a TU-1900 spectropho-
tometer and 1.00 cm quartz cells were used (all from Bei-
jing Persee, Inc., Beijing, China).
As a control, the electronic spectra of 0.10 mM cis-
[Pt(NH3)2Cl4], 0.10 mM cis-[Pt(NH3)2Cl2], 1.0 mM CA,
and 1.0 mM Pen recorded in an HAc/NaAc buffer of pH
4.42 are given in Figure S1 in Supplementary Materials
(SM). The general kinetic traces under pseudo first-order
reaction conditions, that is, the thiol drug total concen-
trations [thiol]tot being ≥ 10∙[Pt(IV)], were investigated. A
kinetic trace for the oxidation of CA by cis-[Pt(NH3)2Cl4]
under a set of reaction conditions is shown in Figure 2 dis-
playing a biphasic nature. The initial absorption decline of
the kinetic trace, being enlarged and shown in the insert of
Figure 2, is designated to the reduction of cis-[Pt(NH3)2Cl4]
2.5
Mass spectral measurements
A reaction mixture of 1.0 mM cis-[Pt(NH3)2Cl4] with
10 mM CA in 10 mM acetic acid after a reaction time of
about 0.5 h was run on a high resolution mass spectrom-
eter (Apex Ultra 7.0T FT-ICR; Bruker Daltonik, Bremen,

4
ZHOU et al.
to cisplatin; while the second phase, that is, the subsequent
absorption increase, is much slower, which can be assigned
to the substitution of the coordination chloride(s) in cis-
platin by the excess CA in the reaction mixture.19 In the
oxidation of Pen by the Pt(IV) compound, similar biphasic
kinetic traces were also observed, cf Figures S2 and S3 in
the SM. Since the kinetic trace is basically separable on the
time scale, the initial kinetic phase was well-simulated by
a single exponential equation (cf Equation 1), where At, A0,
and A∞ pertain to the
ꢀꢁ = ꢀ − ꢀ exp −ꢂ
ꢁ
+ ꢀ∞
(1)
(
)
(
)
0
∞
obsd
absorbances at time t, zero and infinity, respectively.30 In
the equation, kobsd stands for the pseudo first-order rate
constant, and the value A∞ can hardly be acquired directly
from the kinetic trace due to the subsequent reaction, but
is obtainable through the simulation. The simulated result
is also shown in the insert of Figure 2, indicating that the
reduction phase is indeed first-order in [Pt(IV)]. Since the
substitution reactions on cisplatin proceed slowly,31 and
are not directly related to the antioxidative properties of
the drugs, our attention in this work is paid to the redox
processes.
F I G U R E 3 Time-resolved spectra in a wavelength range from
220 to 340 nm. The reaction conditions are: [Pt(IV)] = 0.20 mM,
[CA]tot = 2.00 mM, buffer of pH 4.42, 25.0◦C, and 1.0 M ionic strength.
Spectra were acquired at 3.3, 20, 40, 70, 100, 150, 200, 300, 400, 600,
800, 1000, 1500, 1995 ms [Color figure can be viewed at wileyonlineli-
brary.com]
the observed second-order rate constant.
3.2
Rapid scan spectra and rate law
[
]
[
]
−d Pt NH Cl4 ∕dt = ꢂobsd Pt NH Cl4
(
)
(
)
3
3
2
2
To shed more light on the course of the redox reactions,
time-resolved spectra were acquired in an HAc/NaAc
buffer of pH 4.42 by use of the stopped-flow spectrometer,
cf Figure 3. Characterized attributes in the spectra are that
the absorption peak at about 236 nm and absorption shoul-
der between 260 and 280 nm retained unchanged and no
new absorption band emerged, suggesting that neither a
rapid substitution reaction on the Pt(IV) compound nor a
strong association between CA and cis-[Pt(NH3)2Cl4] took
place in the course of the redox reaction.
[
]
′
= ꢂ thiol
[
Pt NH Cl4
(2)
]
(
)
3
tot
2
Values of k′ at different pHs were evaluated from the lin-
ear plots by a least-squares method and are summarized in
Table S1 in the SM and are also shown in Figures 5 and 6.
3.3
Stoichiometric ratios for the redox
reactions
The influence of changing [thiol]tot on the reaction rates
was investigated in an extended series of buffer solutions.
In each buffer solution, the variation region of [thiol]tot
was amended within the buffering capacity of that particu-
lar buffer, thus not causing any pH change of the buffered
solutions. Consequently, a large body of kinetic data was
collected for the oxidation reactions of both CA and Pen.
Plots of kobsd versus [Pen]tot in several buffers of differ-
ent pH are shown in Figure 4, whereas the kobsd versus
[CA]tot plots are given in Figure S4 in the SM. Undoubt-
edly, these plots are linear and do not possess any signifi-
cant intercepts within the experimental errors, revealing
A spectrophotometric titration approach was opted to
study the reaction stoichiometry of the present redox
processes.32–34 A series of reaction mixtures of cis-
[Pt(NH3)2Cl4] with CA in a phosphate buffer of pH 6.29
were prepared, in which [Pt(IV)] = 0.20 mM was kept
constant while [CA]tot was varied from 0 to 0.80 mM.
Moreover, the absorption measurement at 265 nm for a
reaction time of about 2 min was controlled for each reac-
tion mixture avoiding the interference from the subse-
quence substitution reactions (vide supra). The measured
absorption value as a function of [CA]tot is shown in Fig-
ure S5 in the SM; unambiguously the data points follow
two straight crossing lines. A reaction ratio of ∆[Pt(IV)]:
∆[CA]tot = 0.20 mM:(0.43 0.01) mM = 1:(2.15 0.05)
that the redox reactions are also first-order in [thiol]tot
.
Therefore, the redox reactions follow overall second-order
kinetics as expressed by Equation (2), where k′ represents

ZHOU et al.
5
F I G U R E
5
Observed second-order rate constant k′ (in loga-
rithmic scale) as a function of pH for the oxidation of CA by cis-
[Pt(NH3)2Cl4] at 25.0◦C and 1.0 M ionic strength. The solid curve
was obtained by simulation of Equation (6) to the experimental data
using a weighted nonlinear least-squares method [Color figure can
be viewed at wileyonlinelibrary.com]
sulfinic acid were not observed, which is similar to the
oxidations of glutathione and DL-homocysteine by Pt(IV)
anticancer prodrugs.18,19
A similar spectrophotometric titration for the Pen oxida-
tion was also carried out; Figure 7 shows the result together
with the detailed experimental conditions. No doubt, the
data points in the figure also follow two crossing straight
lines, the intersection point renders a ratio of ∆[Pt(IV)]:
∆[Pen]tot = 0.30 mM:(0.163 0.006) mM = 1:(0.54 0.02).
This ratio, differentiating largely with the one obtained
above for the CA oxidation, is in fact close to a 2:1 sto-
ichiometry. Equation (4) is inferred to correspond to an
ideal 2:1 stoichiometry:
F I G U R E 4 Plots of the pseudo first-order constant kobsd versus
[Pen]tot at 25.0◦C and 1.0 M ionic strength [Color figure can be viewed
at wileyonlinelibrary.com]
was estimated from the intersection point. Essentially, the
estimated ratio is very close to a theoretical stoichiom-
etry of 1:2 if CA was assumed to be oxidized to CA
disulfide (H2NCH2CH2SSCH2CH2NH2 or CA-disulfide)
as expressed by Equation (3):
H2NCH(COOH)CMe2SH + 2[Pt(NH3)2Cl4]
+2H2O → H2NCH(COOH)CMe2SO2H
[
]
[
]
Pt NH Cl4 + 2H NCH CH SH → Pt NH Cl2
(
)
(
)
3
2
2
2
3
2
2
+2[Pt(NH3)2Cl2] + 4HCl
(4)
+H2NCH2CH2SSCH2CH2NH2 + 2HCl
(3)
Equation
(4)
hypothesizes
Pen-sulfinic
acid
A high-resolution mass spectrum, acquired for a reac-
tion mixture of 10 mM CA and 1.0 mM cis-[Pt(NH3)2Cl4]
in 1.0 mM HAc after a reaction time of about 0.5 h, is given
in Figure S6 in the SM. From the peak assignments given
in the figure legend, CA-disulfide was indeed the primary
oxidation product of CA, justifying Equation (3). Under the
pseudo first-order conditions with an excess of CA, deeper
oxidations of CA probably yielding sulfenic acid and/or
(H2NCH(COOH)CMe2SO2H or Pen-SO2H) being a
major oxidation product of Pen. In order to confirm this
hypothesis, a reaction mixture of 8 mM Pen with 1 mM
cis-[Pt(NH3)2Cl4] was subjected to an ESI mass spectral
analysis; both positive and negative ionization modes
were performed. Figure S7 in the SM gives rise to the mass
spectra together with the peak analysis in the figure leg-
end. The analytic results unravel that two major oxidation

6
ZHOU et al.
products of Pen were generated in the reaction mix-
ture: Pen-sulfinic acid and Pen-disulfide (Pen-S-S-Pen).
However, the percentages of the two products cannot be
evaluated from their peak heights since their mass spectral
sensitivities are very different (the sulfinic acid form has a
much lower sensitivity based on our experience). Hence,
deeper oxidations than a disulfide were confirmed in the
Pen oxidation in this work, which was not observed in the
oxidations of other sterically unhindered thiol compounds
by Pt(IV) compounds.18,19
3.4
Mechanistic elucidations
All the reaction characters observed in the oxidation of
CA by cis-[Pt(NH3)2Cl4] (including time-resolved spec-
tra, overall second-order kinetics, and the reaction stoi-
chiometry) are similar to those found in the oxidations of
glutathionine and dl-homocysteine by Pt(IV) anticancer
prodrugs.18,19 Thus, a similar reaction mechanism is sug-
gested as delineated in Scheme 1. In the scheme, reac-
tions described by k1–k3 are the rate-determining steps,
which have been interpreted in terms of a Cl+ transfer
from the Pt(IV) compound to the attacking sulfur atom of
CA, resulting in formation of transient species chloroth-
iol and sulfenylchloride.17–19 These transient species can be
trapped rapidly by another CA molecule leading to forma-
tion of CA-disulfide (vide infra).18,19
F I G U R E 6 Observed second-order rate constant k′ (in loga-
rithmic scale) as a function of pH for the oxidation of Pen by cis-
[Pt(NH3)2Cl4] at 25.0◦C and 1.0 M ionic strength. The solid curve
was acquired by simulation of Equation (8) to the experimental data
using a weighted nonlinear least-squares method [Color figure can
be viewed at wileyonlinelibrary.com]
In the oxidation of Pen by cis-[Pt(NH3)2Cl4], deeper oxi-
dations engender a disparate stoichiometric ratio and two
oxidation products of Pen. Except for these salient differ-
ences, the other kinetic characters are all similar to those
found in the CA oxidation. When all these attributes are
put together, a reaction mechanism is proposed as depicted
in Scheme 2 for the Pen oxidation process involving rate-
determining steps (i)-(iv). A transient species chloroth-
iol or sulfenylchloride is produced in each of the rate-
determining steps. Each transient species is anticipated to
be followed by a few fast reactions,35,36 which are steps (v)-
(viii) in Scheme 2 (with +H3NCH(COO‒)CMe2SCl (or Pen-
SCl in short) generated in the reaction (iii) as an exam-
ple). Specifically, step (v) is a trapping reaction of Pen-SCl
by excess Pen in reaction solution35,36; this type of trap-
ping reaction was very fast in the case of CA, but could be
much slower in the case of Pen due to the highly sterically
congested nature around the sulfur atom in both Pen-SCl
and Pen. On the one hand, the trapping reaction brought
about the product Pen-disulfide; on the other, a big por-
tion of Pen-SCl had a good chance to undergo a hydrolytic
reaction, leading to formation of another reactive species
+H3NCH(COO‒)CMe2SOH (Pen-sulfenic acid), that is, the
step (vi) in Scheme 2.35,36 The doom of Pen-sulfenic acid
was controlled by steps (vii) and (viii); step (vii) is another
F I G U R E
7
Spectrophotometric titration experiment: absorp-
tion value at 265 nm as function of [Pen]tot for a series of reaction
mixtures of cis-[Pt(NH3)2Cl4] with Pen in a phosphate buffer of pH
6.29. In the reaction mixtures, [Pt(IV)] = 0.30 mM was maintained
constant and [Pen]tot was varied from 0 to 0.50 mM; a reaction time
of about 2 min was controlled for each of the reaction mixtures before
the absorption measurement [Color figure can be viewed at wileyon-
linelibrary.com]

ZHOU et al.
7
k1
+
+
+
HSCH2CH2NH3
HSCH2CH2NH3
_
Cl
[Pt(NH3)2Cl2] + Cl
Ka1
H+
_
k2
+
+
[Pt(NH3)2Cl4] +
SCH2CH2NH3
ClSCH2CH2NH3
H2NCH2CH2S-SCH2CH2NH2
_
_
Cl + nH+
CA
[Pt(NH3)2Cl2] + Cl
Ka2
H+
k3
_
ClSCH2CH2NH2
SCH2CH2NH2
_
[Pt(NH3)2Cl2] + Cl
S C H E M E 1 The reaction mechanism suggested for the oxidation of cysteamine by cis-[Pt(NH3)2Cl4] [Color figure can be viewed at wiley-
onlinelibrary.com]
k1
+
+
+H3NCH(COOH)CMe2SH
(i)
[Pt(NH3)2Cl4] + H3NCH(COOH)CMe2SH
_
Cl
Ka1
[Pt(NH3)2Cl2] + Cl
H+
k2
+
_
_
+H3NCH(COO )CMe2SH
Cl
(ii)
+H3NCH(COO )CMe2SH
[Pt(NH3)2Cl4] +
_
[Pt(NH3)2Cl2] + Cl
Ka2
H+
k3
_
_
_
+
[Pt(NH3)2Cl4] + H3NCH(COO )CMe2S
+H3NCH(COO )CMe2SCl
(iii)
(iv)
_
[Pt(NH3)2Cl2] + Cl
Ka3
H+
k4
_
_
_
H2NCH(COO )CMe2SCl
[Pt(NH3)2Cl4] + H2NCH(COO )CMe2S
_
[Pt(NH3)2Cl2] + Cl
_
_
_
+H3NCH(COO )CMe2SCl + +H3NCH(COO_ )CMe2S
Pen-S-S-Pen + Cl
(v)
_
_
+H3NCH(COO )CMe2SCl + H2O
+H3NCH(COO )CMe2SOH + HCl
(vi)
(vii)
_
_
_
_
+H3NCH(COO )CMe2SOH + +H3NCH(COO )CMe2S
Pen-S-S-Pen + OH
_
_
+H3NCH(COO )CMe2SOH + [Pt(NH3)2Cl4] + H2O
+H3NCH(COO )CMe2SO2H + 2HCl (viii)
[Pt(NH3)2Cl2]
S C H E M E 2 A schematic representation of the proposed reaction mechanism for the oxidation of d-penicillamine by cis-[Pt(NH3)2Cl4],
in which reactions (i)-(iv) are the rate-determining steps [Color figure can be viewed at wileyonlinelibrary.com]
trapping reaction while step (viii) is a fast oxidation by
another Pt(IV) molecule. By analogy, the highly steric con-
gestion of Pen could make the rate of step (vii) slower sig-
nificantly than that of step (viii). Step (vii) may only play
a very minor role in the proposed mechanism but can-
not be totally eliminated. It is thus understandable that
the percentages of Pen-disulfide and Pen-sulfinic acid are
largely determined by the relative rates of steps (v) and
(viii). Nevertheless, step (viii) was apparently an open reac-
tion route giving rise to a high portion of Pen-sulfinic acid
and accounting for the stoichiometric ratio measured (vide
supra).

8
ZHOU et al.
TA B L E 1 Rate constants of the rate-determining steps
evaluated for the oxidations of cysteamine and d-penicillamine by
cis-[Pt(NH3)2Cl4] at 25.0◦C and ionic strength of 1.0 M
late the k′-pH dependence data in Table S1 in the SM and
the pKa values were treated as the fixed constants, it turned
out that the k1 value was indeterminate (or not observed).
Consequently, Equation (7) was reduced to Equation (8) by
exclusion of the k1-term in Equation (7).
Thiol drug
CA
km
k1
Value (M–1 s–1)
0.40 0.05
k2
k3
k1
k2
k3
k4
(1.85 0.05) × 105
(2.62 0.09) × 105
Not observed
2
ꢂ2ꢄꢃ1ꢃH + ꢂ3ꢄꢃ1ꢄꢃ2ꢃH + ꢂ4ꢄꢃ1ꢄꢃ2ꢄꢃ3
′
ꢂ
=
(8)
3
2
ꢃH + ꢄꢃ1ꢃH + ꢄꢃ1ꢄꢃ2ꢃH + ꢄꢃ1ꢄꢃ2ꢄꢃ3
Pen
0.43 0.08
The simulation of the data by Equation (8) was then exe-
cuted, offering an essentially excellent fit, cf Figure 6; the
k2-k4 values secured by the simulation are also summa-
rized in Table 1.
(1.04 0.05) × 105
(3.53 0.09) × 105
The excellent fits in Figures 5 and 6 lend a strong
support to the reasonableness of the proposed reaction
mechanisms. The rate-determining steps in Schemes 1
and 2 are anticipated to take place via a bridge forma-
tion between one of the trans-coordinated chlorides in
cis-[Pt(NH3)2Cl4] and the attacking sulfur atom of CA or
Pen whereas the bridge formation leads consequently to a
Cl+ transfer from the Pt(IV) center to the sulfur atom.17,35
Moreover, recent theoretical calculations on the reductions
of trans-dichloro-platinum(IV) compounds clearly bolster
the above Cl+ transfer mechanism.39,40 Among the oxi-
dants exploited for the oxidations of the two drugs,10–15
the chloramine was the only one in its reaction mecha-
nism involving a Cl+ transfer as that in the present reac-
tion systems,11 but the chloramine oxidations were slower
in rates.
3.5
Calculations of rate constants of the
rate-determining steps
Equation (5) as the exact rate law was derived from
Scheme 1, where Ka1 and Ka2 are the acid dissociation con-
stants of CA and aH stands for the proton activity, cor-
responding to the measured pH values (cf derivations of
Equations 5-7 in the SM).
−d Pt IV ∕dt
[
(
)]
2
ꢂ1ꢃH + ꢂ2ꢄꢃ1ꢃH + ꢂ3ꢄꢃ1ꢄꢃ2
=
CA
[
Pt IV
(
]
[
)]
tot
2
ꢃH + ꢄꢃ1ꢃH + ꢄꢃ1ꢄꢃ2
(5)
When Equation (5) is matched with Equation (2), Equa-
tion (6) is affordable:
3.6
An alternative reaction mechanism
2
ꢂ1ꢃH + ꢂ2ꢄꢃ1ꢃH + ꢂ3ꢄꢃ1ꢄꢃ2
′
ꢂ =
(6)
2
ꢃH + ꢄꢃ1ꢃH + ꢄꢃ1ꢄꢃ2
An alternative reaction mechanism for the oxidation of CA
by cis-[Pt(NH3)2Cl4] is also suggested, as delineated in Fig-
ure S8 in the SM. This mechanism involves formation of
free radicals on the sulfur atom from CA species and gen-
eration of a Pt(III) intermediate in the rate determining-
steps (a)-(c); Pt(III) is hardly holding the same config-
uration as cis-[Pt(NH3)2Cl4] and is thus presumed as
[Pt(NH3)2Cl3]. If we assume that the Pt(III) intermedi-
ate is very active, it rapidly reacts with the thiol species
Simulation of Equation (6) to the k′-pH dependence data
in Table S1 was performed by use of a weighted nonlin-
ear least-squares method. In the simulation, k1-k3 were
opted as adjustable parameters while the reported pKa val-
ues of CA (pKa1 = 8.37 and pKa2 = 10.44 at 25.0◦C and
µ = 0.50 M37) were used as the direct inputs. The simu-
lation conferred a good fit (cf Figure 5), concurrently gen-
erating the values of k1-k3, which are listed in Table 1.
According to Scheme 2, the rate expression for k′ was
deduced as Equation (7) (the derivation process is demon-
strated in the SM):
‒
in excess; with SCH2CH2NH2 as an example, this rapid
reaction is reaction (d) in Figure S8 generating another
free radical∙SCH2CH2NH2. Reaction (e) in Figure S8 is the
rapid combination of two free radicals leading to formation
of final product. Based on the reaction mechanism in Fig-
ure S8 and on the assumptions, the rate Equation (5) is also
obtainable. In such case, the values of rate constants k1-k3
in Table 1 have a different meaning.
3
2
ꢂ1ꢃH + ꢂ2ꢄꢃ1ꢃH + ꢂ3ꢄꢃ1ꢄꢃ2ꢃH + ꢂ4ꢄꢃ1ꢄꢃ2ꢄꢃ3
′
ꢂ =
3
2
ꢃH + ꢄꢃ1ꢃH + ꢄꢃ1ꢄꢃ2ꢃH + ꢄꢃ1ꢄꢃ2ꢄꢃ3
(7)
In fact, the assumptions of formation of the Pt(III)
intermediate and of Pt(III) being very reactive in the
mechanism are hardly finding supports from the experi-
The dissociation constants of Pen were reported to be
pKa1 = 1.9, pKa2 = 7.92, and pKa3 = 10.50 at 25.0◦C and
µ = 1.0 M.38 When Equation (7) was employed to simu-

ZHOU et al.
9
ments in this work. If the rate-determining steps involve
single-electron transfer processes, the speeds of these
processes are expected to be closely related to the
redox potentials of the Pt(IV) complexes; the higher the
redox potential of Pt(IV) complex, the higher of the
electron-transfer speed. Our earlier works demonstrated
that cis,trans-[Pt(NH3)2Cl2Br2] and trans-[PtBr2(CN)4]2‒,
having lower redox potentials than cis-[Pt(NH3)2Cl4]
and trans-[PtCl2(CN)4]2‒, respectively, are reduced much
faster.17,32 Hence, our earlier findings are against the reac-
tion mechanism illustrated in Figure S8.17,32
is also a medically critical pH region for functions for most
of drugs.
C O N F L I C T O F I N T E R E S T
The authors declare no conflict of interest.
DATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are openly
available in at https://doi.org/[doi], reference number [ref-
erence number].
O RC I D
TieshengShi https://orcid.org/0000-0002-0767-6324
3.7
Analysis of reactivity of the drug
species
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How to cite this article: Zhou L, Li T, Sun Y,
et al. Mechanistic scrutiny of the oxidations of
thiol-containing drugs cysteamine and
D-penicillamine by
cis-diamminetetrachloroplatium(IV). Int J Chem
Kinet. 2020;1-10. https://doi.org/10.1002/kin.21464