Monatsh Chem (2014) 145:17271735
DOI 10.1007/s00706-014-1262-1
ORIGINAL PAPER
Kinetics and mechanism of reactions of some essential amino acids
with cis-diaqua(1,3-diaminopropane)-platinum(II) ion: a potent
anticancer agent
Sumon Ray Debabrata Nandi Animesh Chattopadhyay
Roshni Sarkar Parnajyoti Karmakar Alak K. Ghosh
Received: 11 March 2014 / Accepted: 10 June 2014 / Published online: 30 July 2014
Springer-Verlag Wien 2014
Abstract The substitution of both coordinated water
molecules of cis-diaqua(1,3-diaminopropane)platinum(II)
complex by a series of amino acids, viz. L-asparagine,
L-glutamic acid, and glycine was investigated as a function of
ligand concentration, temperature, and pH and found to occur
in two consecutive reaction steps. The mechanism of the
substitution reactions appears associative in nature as sup-
ported by the large and negative values of DS=. The kinetic
study has been substantiated by product isolation, UV, IR,
and ESI-MS spectroscopic analysis and rate parameters have
been evaluated under different reaction conditions.
Keywords Platinum(II)  1,3-Diaminopropane 
Amino acids  Kinetics
Introduction
In the twenty-first century, the number of patients suffering
from cancer is increasing worldwide. Cancer, which is
expected to be the most significant disease in the twenty-
first century [1, 2], is generally known to be the unre-
strained growth of a tumor or cells. In the last half of the
twentieth century, several compelling platinum-containing
anticancer complexes were developed. Unfortunately, these
complexes have some toxic side effects, which are intim-
idating because of the development of many severe
secondary complications. Today, widespread effort is being
made to look for new anticancer agents with high potency
and little or no side effects.
S. Ray  D. Nandi  A. Chattopadhyay  R. Sarkar 
P. Karmakar  A. K. Ghosh (&)
Department of Chemistry, The University of Burdwan,
Burdwan 713104, West Bengal, India
e-mail: alakghosh2002@yahoo.co.in
The kinetic and mechanistic description of the square
planar platinum(II) complexes attracted continued attention
due to their intrinsic chemical and bio-medical applications
[3]. In particular, cisplatin and its structural analogs are
used in cancer chemotherapy. But the major drawback of
these chloro derivatives of N,N-chelated platinum(II)
complexes was their nephrotoxicity. However, the aqua
variety was found to be superior to others [35].
Nuclear DNA has been recognized as the principal target
for Pt and certain platinated DNA adducts elicit DNA deg-
radation and apoptosis (programmed cell death) [6, 7]. Even
though intensive efforts of various research groups in recent
past have contributed appreciably towards understanding of
the anticancer bustle of platinum complexes, most of the
studies paying attention on the structural identification of the
binding of the DNA constituents to the metal center by means
of NMR, HPLC, ESI-MS, and X-ray technique. Signifi-
cantly, very less is known about the mechanistic details with
regard to the various steps of adduct formation and the fac-
tors controlling such adduct formation.
This paper reports a thorough study of the concentration
of the ligand, temperature, and pH dependencies of the rates
of substitution of the title complex with amino acids in
aqueous medium. We preferred the above-mentioned amino
acids seeing that platinum complexes can also interact with
other molecules in a cellular environment, like peptide
structures or RNAs. Appraisal of the modes of action of
metal-based antitumor drugs therefore requires the study of
their interactions with a range of feasible biological targets
including amino acids, hormones, peptides, and proteins.
The study of model species such as essential amino acids
can give a hand in the elucidation of more amalgamated
systems. The results of this study, together with those of the
previous investigations, shed light on the kinetic behavior
of the N,N-chelated cationic platinum(II) complexes.
123
1728 S. Ray et al.

Results and discussion
Spectroscopic analysis
The electronic transitions observed in the spectra produced
by UVVis spectroscopy were comparable with what was
expected for platinum(II) coordination complexes (Fig. 1).
The IR spectra of the compounds were measured in the
region 4,000400 cm-1 (Figs. 2, 3, 4) and showed
characteristic changes when compared to those of the free
ligands. The platinumligand vibrations are expected to
occur in a very low frequency range (below 600 cm-1) [8].
FT-IR spectra of L-asparagine (L1H), L-glutamic acid
(L2H), and glycine (L3H)-substituted products, the strong
bands at 437, 443, and 505 cm-1, respectively, are
assigned to the PtN stretching vibrations. The PtO
stretching vibrations are assigned to the bands observed at
497, 526, and 605 cm-1, respectively. The strong bands at

Fig. 1 Absorbance differences

between complex 1, asparagine
(L1H)-substituted product 2,
glutamic acid (L2H)-substituted
product 3, and glycine (L3H)-
substituted product 4; [1]
= 2 9 10-4 mol dm-3, [amino
acids] = 4 9 10-3 mol dm-3,
pH = 4.0, and cell
used = 1 cm quartz

80

437.84
70

497.63
1041.56
1109.07

705.95
1462.04

60
1290.38
1406.11

1238.30

50
%T

40
3261.63

30
3417.86

1681.93

20

10

0
4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

Wavenumber/cm-1

Fig. 2 IR spectrum of the L1H-substituted product

123
Reactions of essential amino acids 1729

90

443.63
1045.42

596.00

526.57
80

1195.87
70

1404.18
1707.00
%T

1666.50
60
3192.19
3234.62
3435.22

50

40

30
4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500
Wavenumber/cm-1

Fig. 3 IR spectrum of the L2H-substituted product

90

80

777.31
927.76
70

605.65
1195.87
1126.43
2611.62

688.59
891.11
1039.63

60
%T

505.35
3437.15

2966.52

1330.88

50
1500.62

1408.04
3132.40

40
1598.99

30

20
4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500
Wavenumber/cm-1

Fig. 4 IR spectrum of the L3H-substituted product

3,261, 3,234, and 3,132 cm-1 in the experimental FT-IR
spectrum are assigned to the NH stretching vibrations of
the amino groups. The asymmetric COO stretching fre-
quency of the amino acids occurs at 1,5801,660 cm-1
when the group is coordinated to metals, whereas a non-
coordinated COO group has the masym (COO) stretch at
lower frequency [9]. The band at 1,682, 1,666, and
1,599 cm-1 is therefore assigned to the masym (COO) of the
metal-bound carboxyl group. From the individual assign-
ments of different bands it can be presumed that the final

123
1730 S. Ray et al.

Fig. 5 Mass spectrum of the L1H-substituted product

Fig. 6 Mass spectrum of the L2H-substituted product

123
Reactions of essential amino acids
Fig. 7 Mass spectrum of the L3H-substituted product
products are (O,N)-coordinated chelates and amino acids
behave like a bidentate ligand in the experimental pH. ESI-
MS of the resulting solutions, obtained from the 1:1 mix-
ture of the substrate complex 1 and three amino acids are
shown in Figs. 5, 6, and 7. It is clear from these spectra that
the ions at m/z = 400.07, 415.07, and 343.05 (major peak)
for L1H, L2H, and L3H are the precursor ion species in the
mixture solutions and this is cautiously attributed to [amino
acids ? Pt ? (1,3-diaminopropane)] the relative abun-
dance of isotope peaks, which matched with the predictable
values. The precursor ions are made known schematically
(Scheme 1).
We proposed that Pt(II) is likely to be coordinated to
both COO and NH2 considering the strong coordination
ability of these functional groups [1012]. Thus, the
structure proposed here for product ion species, deduced
from ESI-MS and IR are generally consistent with those
derived from other experimental methods.
Kinetic studies
The pKA values of the amino acids L1H, L2H, and L3H
are 2.02 a(COOH), 8.84 a(NH3?); 2.19 a(COOH),
1731
6.04(COOH), 9.67 a(NH3?) and 2.34 a(COOH), 9.60
a(NH3?) [13], respectively, at 25 C. From the pKA
values of the ligands it can be concluded that at pH 4.0,
ligands L1H, L2H, and L3H remain in the zwitterionic
forms, which take part in the reaction. The pK1 and pK2
values for cis-[Pt(1,3-diaminopropane)(H2O)2]2? have
been evaluated by Irving-Rossotti method [14] and found
to be 6.14 and 7.67, respectively. Hence it can be
assumed that at pH 4.0, the reactant complex exists as
diaqua ion. The reactions between complex 1 and amino
acids involve a two-step consecutive route; we put for-
ward that in the first step one aqua ligand is replaced
from cis-[Pt(1,3-diaminopropane)(OH2)2]2? by the LH.
The second step is slower, where the remaining aqua
ligand is substituted with chelate ring closure. The rate
constant for such a process can be evaluated by assuming
the following scheme:
k1 k2
A ! B ! C 1
where A is the diaqua species 1, B is the single substituted
intermediate, and C is the final product 2, [Pt(1,3-diami-
nopropane)(L)]?. Formation of C from B is predominant
after some time has elapsed.
123
1732 S. Ray et al.

Scheme 1 Table 1 103 k1(obs)/s-1 values for different concentrations of LH at

different temperatures
Ligand Temp./C 103 [LH]/mol dm-3
(0.1 C)
2.00 3.00 4.00 6.00 10.00

L1H 35 1.03 1.46 1.95 2.99 5.21

40 1.30 1.95 2.48 3.76 6.61
45 1.68 2.52 3.48 5.04 8.58
50 2.15 3.32 4.14 6.60 10.91
55 2.91 4.36 5.69 8.84 14.52
L2H 35 0.87 1.32 1.78 2.53 4.44
40 1.11 1.54 2.18 3.38 5.57
45 1.45 2.17 2.96 4.49 7.28
50 1.95 3.05 3.95 5.68 9.75
55 2.67 4.00 5.49 7.81 13.21
L3H 35 0.59 0.84 1.21 1.79 2.99
40 0.78 1.28 1.52 2.28 3.79
45 1.04 1.62 2.23 3.21 5.35
50 1.39 2.14 2.70 4.21 7.01
55 1.81 2.78 3.62 5.61 9.13
-4 -3 -3
[1] = 2 9 10 mol dm , pH = 4.0, l = 0.1 mol dm NaClO4

k1obs k1 LH; 4

where k1 is the second-order rate constant for the first aqua

ligand substitution. A plot of k1(obs)/s-1 versus [LH] should
be linear, passing through the origin. This was found to be
. so at all temperatures studied (Figs. 8, 9, 10). The second-
order rate constants k1 calculated from the slopes of the
Calculation of k1 plots of k1(obs)/s-1 versus [LH]/mol dm-3 at different
temperatures are given in Table 2.
The rate constant for the first phase of the reaction A ? B
was calculated from the absorbance data using the Weyh Calculation of k2
and Hamm equation, as discussed in our previous work
[15]. The k1(obs) values for different ligand concentrations The B ? C step is assigned to ring closure in which the
at different temperatures are given in Table 1. The fol- NH2 group of the amino acids binds the metal center
lowing mechanism is proposed: through N. This chelation step is independent of ligand
 2 k1 ; slow concentration. Slower reaction rate seems to be due to the
Pt1; 3-diaminopropaneH2 O2 LH !
Substitution steric hindrance. At each temperature, the k2 values were
Pt1; 3-diaminopropaneH2 OL H3 O calculated from the limiting linear portion (when t is large)
2 of the ln(A? -At) versus t curves and are collected in
k2
Pt1; 3-diaminopropaneH2 OL ! Table 2. Unlike k1, k2 was found to be independent of
Chelation
Pt1; 3-diaminopropaneL H2 O ligand concentration at each of the temperatures studied.

where LH is the zwitterionic form of amino acids. Based Effects of pH on reaction rate
on the above scheme, a rate expression Eq. (4) can be
derived for the A ? B step: The reaction was studied at four different pH values (2.8,
k1 Pt1;3-diaminopropaneH2 O2 3.4, 4.0, and 4.6). The k(obs) values increased with increase
dB=dt 2 total LH 3
in pH; at fixed concentration of 2 9 10-4 mol dm-3 of
where [Pt(1,3-diaminopropane)(H2O)2? 2 ]total is the complex 1, 6 9 10-3 mol dm-3 glutamic acid, and
concentration of the unreacted complex. Hence, we can 0.1 mol dm-3 NaClO4 ionic strength. The increase in rate
write may be explained based on the acid dissociation equilibria

123
Reactions of essential amino acids 1733

Fig. 8 Plot of k1(obs)/s-1 versus [L1H] 16 E

at different temperatures: A = 35 C,
B = 40 C, C = 45 C, D = 50 C,
and E = 55 C
12 D

-1
10 k1(obs) /s
C
8
B

3
A
4

0
0 2 4 6 8 10
3 -3
10 [Asparagine]/mol dm

Fig. 9 Plot of k1(obs)/s-1 versus E

[L2H] at different temperatures:
A = 35 C, B = 40 C,
C = 45 C, D = 50 C, and 12
E = 55 C D
-1
10 k1(obs)/s

8 C

B
3

A
4

0
0 2 4 6 8 10
3 -3
10 [Glutamic acid]/mol dm

8
D
-1

C
10 k1(obs)/s

B
3

4
A

0
0 2 4 6 8 10
3 -3
10 [Glycine]/mol dm

Fig. 10 Plot of k1(obs)/s-1 versus [L3H] at different temperatures: A = 35 C, B = 40 C, C = 45 C, D = 50 C, and E = 55 C

123
1734 S. Ray et al.

of the amino acids and the complex. In the studied pH Conclusion

range, the complex starts to switch into hydroxoaqua spe-
cies from diaqua species. The enhancement in rate
therefore explained by the deprotonation and increased
donor ability of the amino acids and reactant complex. The
characteristic pH dependence for the substitution reaction
can be theoretically explained by considering the following
derived rate expression:
kobs ka Ka Lt =Ka H ;
where ka, Ka, and [L]t are the rate constant, acid dissoci-
ation constant of the amino acid carboxylate group, and
total amino acid concentration, respectively. In subsequent
kinetic runs, the substitution reactions were followed at a
constant pH of 4.0 to avoid complications from an addi-
tional parameter of [H?] to the rate equation.
Effects of temperature on reaction rate
These reactions were monitored at five different tempera-
tures for diverse ligand concentrations and the substitution
rate constants for both A ? B (k1) and B ? C (k2) steps
are arranged in Table 2. The activation parameters calcu-
lated from Eyring plots are tabulated in Table 3.
Table 2 k1 and k2 values for the substitution reaction
Ligands Temp./C k1/mol-1 dm3 s-1 105 k2/s-1
In summary, from the pKA values, it is clear that all three
amino acids exist as zwitterion at pH 4.0. The reaction with
amino acids proceed first via substitution of a water mol-
ecule by the oxygen donor, followed by a second
substitution step, which is independent of the nucleophile
concentration, thus, indicating a ring-closure process.
Further investigations confirmed that a five-membered O,N
chelate ring is formed under liberation of a proton. The
succeeding step is the chelate ring closure, which is slower
than the first step is independent of ligand concentration
but depends on pH and exhibits very large negative values
for DS= compared to the first step.
From a comparison of the studied amino acids, the
following reactivity order for the first step can be
formulated:
Asparagine L1 H [ Glutamic acid L2 H [ Glycine L3 H:
The first step is related with the deprotonation of
COOH group. So decrease in rate from asparagine to
glycine is associated with the pKA values of the ligands.
The chelation reaction also shows different rates for
different amino acids. The rate constant for this reaction
k2 has been found to be in the order:
Asparagine L1 H [ Glycine L3 H [ Glutamic acid L2 H:

L1H 35 0.52 2.65 Chelation step is accompanied by deprotonation of the

40 0.66 3.54 a-NH3? of the amino acids. So rate constant values for the
45 0.86 4.74 second step are in accordance with pKA of a-NH3? group.
50 1.09 6.39 The DH= = =
1 and DH2 values and negative DS1 and DS2
=

55 1.45 8.75 values entail a good extent of ligand participation in the

L2H 35 0.44 1.45 transition state.
40 0.56 2.07
45 0.73 2.93
50 0.97 4.08 Experimental
55 1.32 5.78
L3H 35 0.30 1.65 K2PtCl4 and 1,3-diaminopropane were purchased from
40 0.38 2.32 Sigma-Aldrich and amino acids were purchased from Sisco
45 0.54 3.27 Research Laboratory. All other reagents used in this
50 0.70 4.59
research were obtained from commercial sources and used
55 0.92 6.37
without purification. Solution of the above-mentioned
complex and other reagents used for this work were pre-
pared freshly in double-distilled water before use.
Table 3 Activation parameters for complex 1 by L1H, L2H, and L3H The pH measurements were carried out with the help of
in aqueous medium at pH 4.0 a Sartorius Digital pH meter (model PB11) with an accu-
Ligands DH= DS= -1
DH= DS= -1 racy of 0.01 units. Electronic spectra were measured on a
1 / 1 /J K 2 / 2 /J K
kJ mol-1 mol-1 kJ mol-1 mol-1 Shimadzu UV 2450) spectrophotometer. Infrared (IR)
spectroscopy on KBr pellets was performed on a Shimadzu
L1H 39.90 2.7 -132.5 7.0 49.67 3.8 -211.3 9.4
FT-IR model IR Prestige 21 infrared spectrophotometer
L2H 42.80 2.0 -124.9 6.1 54.41 7.2 -198.8 9.6 from 4,000 to 400 cm-1. ESI-MS were recorded using a
L3H 44.89 3.2 -113.7 9.1 52.67 4.9 -203.5 11.6 micromass Q-Tof microTM mass spectrometer in positive

123
Reactions of essential amino acids
1.0
0.9
0.8
0.7
Abs
0.6
0.5
0.4
0.3
0.0 0.2 0.4 0.6 0.8 1.0
[L1H] / ([L1H] + [M])
Fig. 11 A typical Jobs plot for the reaction between complex 1 and
L1H at pH = 4.0 and ionic strength = 0.1 mol dm-3 NaClO4
ion mode. Kinetic measurements were monitored on a
Shimadzu spectrophotometer (UV-2450) equipped with a
Shimadzu TCC 240A cell temperature controller (accuracy
0.1 C).
cis-Dichloro(1,3-diaminopropane)platinum(II) was syn-
thesized according to an optimized procedure, based on a
previously reported method for Pt(en)Cl2 [16]. The reactant
complex cis-[Pt(1,3-diaminopropane)(OH2)2](ClO4)2 (1)
was prepared from cis-dichloro(1,3-diaminopropane)plati-
num(II) by hydrolysis in the presence of two molar
equivalents of silver perchlorate. The above-stated reactant
complex 1 was then characterized spectrophotometrically.
The products of the reaction between complex 1 and amino
acids were separately prepared by mixing the reactants at
pH 4.0 in different molar ratios, namely 1:1, 1:2, 1:3, 1:4,
and 1:5, and thermostating the mixtures at 60 C for 48 h.
The absorption spectra of the resulting solutions were
recorded and each set was found to exhibit almost identical
absorbance at the characteristic wavelengths irrespective of
their different molar ratios. The difference in spectra
between the product complexes and the substrate complex
is shown in Fig. 1. The pH of the solutions was adjusted by
adding NaOH/HClO4 at pH 4.0 so that reactant complex 1
exists as diaqua species in the reaction mixture. According
to Jobs method of continuous variation, metalligand
compositions were found to be 1:1 in each case (Fig. 11).
1735
All three amino acids were separately mixed with cis-
[Pt(1,3-diaminopropane)(OH2)]2?(1) in 1:1 ratio at the
experimental pH and three different products were isolated
and characterized by IR and mass spectroscopy. The
kinetic measurements of these systems were done accord-
ing to our previous work [17]. Origin software was used for
computational analysis. Rate data, represented as an aver-
age of duplicate runs, were reproducible to within 4 %.
Acknowledgments One of the authors (Sumon Ray) is grateful to
the University Grants Commission, New Delhi, India, for providing
financial assistance and authors are also very much gratified to the
Department of Chemistry, University of Burdwan, West Bengal,
India, for infrastructural amenities.
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