Synthesis, characterization, cytotoxicity, antibacterial and antifungal evaluation of some new platinum (IV) and palladium (II) complexes of thiodiamines

Article abstract of DOI:10.1016/j.ejmech.2007.03.017

Some new platinum (IV) and palladium (II) thiodiamine complexes of type [Pt(L)₂Cl₂] and [Pd(L)Cl₂], [where, L = (cyclohexyl-N-thio)-1,2-ethylenediamine (L¹) and (cyclohexyl-N-thio)-1,3-propanediamine (L²)] have been synthesized. The thiodiamines coordinate as a bidentate N-S ligand. The synthesized platinum (IV) and palladium (II) complexes of the thiodiamines were characterized by elemental analysis, IR, mass, electronic and ¹H NMR spectroscopic studies. These complexes were also screened for cytotoxicity, in vitro antifungal and in vitro antibacterial activities. Thermodynamic parameters such as activation energy (E_a), apparent activation entropy (S^#) and enthalpy change (ΔH) for the dehydration and decomposition reactions of one complex has also been evaluated.

Full text of DOI:10.1016/j.ejmech.2007.03.017

European Journal of Medicinal Chemistry 42 (2007) 1239e1246
http://www.elsevier.com/locate/ejmech
Original article
Synthesis, characterization, cytotoxicity, antibacterial
and antifungal evaluation of some new platinum (IV)
and palladium (II) complexes of thiodiamines
*
A.K. Mishra , N.K. Kaushik
**
Department of Chemistry, University of Delhi, Delhi 110007, India
Received 5 October 2006; received in revised form 12 March 2007; accepted 15 March 2007
Available online 5 April 2007
Abstract
Some new platinum (IV) and palladium (II) thiodiamine complexes of type [Pt(L)2Cl2] and [Pd(L)Cl2], [where, L ¼ (cyclohexyl-N-thio)-1,2-
ethylenediamine (L1) and (cyclohexyl-N-thio)-1,3-propanediamine (L2)] have been synthesized. The thiodiamines coordinate as a bidentate
NeS ligand. The synthesized platinum (IV) and palladium (II) complexes of the thiodiamines were characterized by elemental analysis, IR,
1
mass, electronic and H NMR spectroscopic studies. These complexes were also screened for cytotoxicity, in vitro antifungal and in vitro an-
tibacterial activities. Thermodynamic parameters such as activation energy (Ea), apparent activation entropy (S#) and enthalpy change (DH ) for
the dehydration and decomposition reactions of one complex has also been evaluated.
Ó 2007 Elsevier Masson SAS. All rights reserved.
Keywords: Synthesis; Spectral; Cytototoxicity; Antibacterial; Antifungal and thermal
1. Introduction
testicular, ovarian, bladder carcinomas, head and neck cancers,
lung carcinoma and stomach carcinoma [3e8]. However, the
clinical usefulness of cisplatin has been frequently limited
by its severe side effects such as nephrotoxicity, nausea, oto-
toxicity, neurotoxicity and myelotoxicity [9e11]. Besides,
there is a development in acquired resistance low activity
against breast and colon cancer. Therefore, it is desirable to
develop new platinum based drugs with broader spectrum of
activity, improved clinical efficacy and reduced toxicity, better
than cisplatin [12].
Platinum (IV) and palladium (II) complexes have revealed
significantly greater activity in human than that of cisplatin
[13,14]. The high activity was ascribed to high cellular uptake,
but in vivo reduction alters the pharmacological properties and
thus the effectiveness of the drug. However, platinum (IV) and
palladium (II) complexes have enormous potential as antican-
cer agents. The potential advantages of platinum (IV)
complexes that remain in higher oxidation state in the blood-
stream are their lower reactivity would diminish loss of active
drug and lower the incidence of unwanted side reactions that
lead to toxic side effects [15,16].
The development of potent and effective antineoplastic, an-
tibacterial, antimalarial and antiviral drugs has considerable
interest in chemistry and biology [1,2]. The chemistry of tran-
sition metal complexes of thiodiamines has been largely stud-
ied because of their pharmacological properties [3].
The interest in platinum based antitumour drugs has its or-
igin with the serendipitous discovery by Rosenberg of the
inhibition of the cell division by platinum complexes. Cis-
diaminedichloro-platinum(II) (cis-[PtCl2(NH3)2]) and cis-
diamine-tetrachloro platinum (IV) (cis-[PtCl4(NH3)2]) were
identified as the platinum complexes deducing that platinum
compounds would have uses in cancer treatment with Sarcoma
180 and Leukemia L1210 bearing mice. Cisplatin are widely
applied in the treatment of various types of cancer such as
* Corresponding author. Tel.: þ91 011 27666616.
** Corresponding author.
E-mail
narenderkumar_kaushik@yahoo.co.in (N.K. Kaushik).
addresses:
ajaykmishra1@yahoo.com
(A.K.
Mishra),
0223-5234/$ - see front matter Ó 2007 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2007.03.017

1240
A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
Nowadays, attention is focused on platinum (IV) and palla-
out on Digital Conductometer Model PT-827, India using
DMSO solution. Model Jeol SX102/DA-600 (KV 10MA)
was used for recording mass spectra of the ligands in
dium (II) complexes with bioactive ligands, because of the
lower toxicity and the possibility of oral administration of
some potent platinum (IV) and palladium (II) compounds, as
well as the fact that they can coordinate to DNA.
1
CH3OH solution. H NMR was recorded using d6-DMSO on
Bruker Spectrospin 300 spectrometer. Thermogravimetric
analysis/differential thermal analysis curves for the complexes
were recorded on Shimadzu, model 60 WS Thermal analyzer,
in static air at a heating rate of 10 ꢀC minꢁ1. The platinum cru-
cible was used with alumina as the reference material.
From the studies of platinum complexes in differing cancer
cell lines and DNA binding studies, some important structure
activity rules have previously been summarized [17]. The
three overriding factors in designing platinum drugs appear
to be chain length and flexibility, hydrogen bonding capacity
and charge of linking chain and the geometry of the chloro li-
gands to the linking chain. For the aliphatic chains used by
some workers [17,18], the ideal length of the linker appears
to be eight atoms (two amine and six methylene groups).
This is evident in the dinuclear and trinuclear complexes.
Both are much more active than the other analogues where
the linking chain is either shorter or longer. For the polyamine
platinum complexes, it has been proved that increasing the
charge and the chain length, leads to complexes which are
more active [17e20].
However, flexibility also appears to be a major factor. Studies
show that both the dinuclear [19] and trinuclear [20] complexes
were much less active than their aliphatic equivalents. It has
been observed [20,21] that the complexes containing aromatic
ligands show poorer cytotoxicity. This would indicate that
straight chain aliphatic, or NH3, ligands are preferable, over ar-
omatic ones. The best linking ligands appear to contain both
positive charges and hydrogen bonding capacity in the form of
either a charged platinum centre and/or charged amine groups.
In view of number of applications of the thiodiamines [17e
22] and the well proven clinical utility the platinum metal
complexes, we have prepared, platinum (IV) and palladium
(II) complexes of the thiodiamines. These complexes were
characterized and screened for antibacterial and cytotoxic ac-
tivities. Thermodynamic parameters such as activation energy
(Ea), apparent activation entropy (S#) and enthalpy change
(DH ) for the dehydration and decomposition reactions of the
complexes have also been evaluated.
2.1. Preparation of thiodiamines
The (cyclohexyl-N-thio)-1,2-ethylenediamine and (cyclo-
hexyl-N-thio)-1,3-propanediamine have been prepared by the
modification of the reported method [21e23].
2.1.1. Preparation of (cyclohexyl-N-thio)-1,2-
ethylenediamine (L1)
In a three necked round bottle flask 5.72 mL (0.05 mol) of
cyclohexylamine (density 0.867 g cmꢁ3) was dissolved in
25 mL methanol and chilled it. To this, solution 2.8 g
(0.05 mol) of potassium hydroxide in 1 mL of water and
10 mL of methanol was added and mixed with constant stir-
ring. An ice-cold solution of 3.02 mL (0.05 mol) carbon disul-
phide (density 1.26 g cmꢁ3) in 3 mL methanol was added to
the above mixed solution. The temperature of the reaction
mixture was maintained below 10 ꢀC by keeping flask in
a freezing mixture of common salt and ice. During the process,
a white crystalline precipitate of N-cyclohexyl dithiocarba-
mate was separated. It was filtered, washed with ice-cold aque-
ous methanol (water: methanol; 2:8). The product was then
suspended in 10 mL methanol and treated with freshly pre-
pared potassium chloroacetate. The potassium chloroacetate
was freshly prepared by dissolving 4.73 g chloroacetic acid
in 3 mL ice-cold water and mixing it in 5 mL aqueous solution
of 2.8 g potassium hydroxide. The temperature of the reaction
mixture was kept at about 40 ꢀC for an hour in a water bath
and the contents were left overnight at room temperature
(25 ꢀC). After 24 h methanolic solution of 3.37 mL (0.05 mol)
ethylenediamine (density 0.897 g cmꢁ3) was added to the re-
action mixture. The content was then heated at 40 ꢀC on a wa-
ter bath for about 45 min. When the desired product began to
separate out, it was cooled in ice for 24 h and filtered. (Cyclo-
hexyl-N-thio)-1,2-ethylenediamine thus obtained was recrys-
tallized from methanol and dried under vacuum over CaCl2
at room temperature (25 ꢀC).
2. Materials and methods
All the reagents used were AR grade. The analysis of
CHNS/O contents of ligands and metal complexes were
done on Elementar Analysensysteme Gmbh Vario El-III. IR
spectra were recorded on PerkineElmer spectrum 2000
FTIR spectrometer using KBr disc. Electronic spectra were re-
corded in DMSO solvent on Shimadzu UV-visible spectropho-
tometer Model 1601. Conductance measurements were carried
The reactions taking place in the preparation are shown
below:
S-K+
H O
+
+ CS2 + KOH
NH
C
S
NH2
2
NH
C
S
S-K+ ClCH2COOK
+
NH
C
SCH2COOK + KCl
S

A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
1241
NH CH CH NH
2
NH
C
S
SCH COOK
+
2
HSCH2COOK +
2
2
2
NH
C
S
NHCH2CH2NH2
stirred for 4e5 h. The colour of solution changed from yellow
to yellowish orange. The resulting yellowish orange precipi-
tate obtained was washed with double distilled water several
times and dried in desiccator over CaCl2 under vacuum.
CHNS-analysis; Found (calculated) %: C; 52.91 (53.73), H;
8.69 (9.45), N; 20.17 (20.89), S; 15.92 (16.03).
Mass spectra (CH3OH); m/z: 201.68.
2.2.2. Preparation of thiohydrazide [Pd(L)Cl2] complexes
where L ¼ L1 and L2
2.1.2. Preparation of (cyclohexyl-N-thio)-1,3-
propanediamine (L2)
The corresponding ligand L [where L1 (0.101 g, 0.5 mmol),
L2 (0.108 g, 0.5 mmol)] in methanol and added with constant
stirring to 1 N HCl solution of palladium chloride (0.089 g,
0.5 mmol). The solution was stirred for 4e5 h. The brownish
precipitate appeared immediately, which separated, washed
with double distilled water several times and dried in desicca-
tor over CaCl2 under vacuum.
N-cyclohexyl dithiocarbamate [10.65 g (0.05 mol)] prepared
as earlier was suspended in 18 mL methanol and treated with
freshly prepared potassium chloroacetate. The potassium chlor-
oacetate was freshly prepared by dissolving 4.73 g chloroacetic
acid in 3 mL ice-cold water and mixing it in 5 mL aqueous so-
lution of 2.8 g potassium hydroxide. The temperature of the re-
action mixture was kept at about 40 ꢀC for an hour in a water
bath and the contents were left overnight at room temperature
(25 ꢀC). After 24 h methanolic solution of 4.36 mL (0.05 mol)
1,3-propanediamine (density 0.85 g cmꢁ3) was added to the re-
action mixture. The content was then heated on a water bath at
40 ꢀC for about 45 min. When the desired product began to sep-
arate out, it was cooled in ice for 24 h and filtered. (Cyclohexyl-
N-thio)-1,3-propanediamine thus obtained was recrystallized
from methanol and dried under vacuum over CaCl2 at room tem-
perature (25 ꢀC).
2.3. In vitro antifungal activity
Most of the compounds have been screened in vitro against
Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger.
Among several methods [24] available, the one method [25,26]
that is common in use in recent times has been adopted.
2.3.1. Microbroth dilution assay
The reactions taking place in the preparation are shown
below:
The susceptibility of the fungi to various fractions of com-
pounds was assayed by microbroth dilution method [27]. Sabo-
NH
C
S
S-K+
C
S
+
ClCH2COOK
NH
SCH2COOK + KCl
+
+
HSCH2COOK
C
S
SCH2COOK NH2CH2CH2CH2NH2
NH
NH
C
S
NHCH2CH2CH2NH2
CHNS-analysis; Found (calculated) %: C; 55.23 (55.81), H;
9.89 (9.77), N; 18.73 (19.53), S; 14.88 (13.97).
Mass spectra (CH3OH); m/z: 215.69.
uraud dextrose medium was dissolved in glass double distilled
water and autoclaved at 10 psi for 15 min. A volume of 90 mL
of medium was added to the wells of cell culture plates (Nunc
Nunclon). The different concentrations in the range of
15ꢁ1000 mg/mL of various fractions were prepared in duplicate
wells and then the wells were incubated with 10 mL of conidial
suspension containing 1 ꢂ 104 conidia. The plates were incu-
bated at 37 ꢀC and examined macroscopically after 48 h for the
growth of Aspergillus mycelia. The activity was represented as
ꢁve if growth was there and þve if medium appeared clear with-
out any visible growth of A. fumigatus, A. flavus and A. niger.
2.2. Preparation of complexes
2.2.1. Preparation of thiohydrazide[Pt(L)2Cl2] complexes
where L ¼ L1 and L2
The corresponding ligand L [where L1 (0.101 g, 0.5 mmol),
L2 (0.108 g, 0.5 mmol)] in methanol was added to aqueous so-
lution of H2PtCl6 (0.103 g, 0.25 mmol). The solution was

1242
A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
2.3.2. Spore germination inhibition assay
Two milliliter cultures of gram negative bacterial strain namely
E. coli were prepared and placed in a water bath overnight at
37 ꢀC. The overnight cultures were diluted with Mullere
Hinton broth. The drugs were suspended to a concentration
of 60 mg/disc (in DMSO) with sterile distilled water in a
96-well microplate. A similar two-fold serial dilution of genta-
mycin (Sigma) was used as positive control against each bacte-
rium. Each bacterial culture of 100 mL was added to each well.
The plates were covered and incubated overnight at 37 ꢀC. To
indicate bacterial growth p-iodonitrotetrazolium violet was
added to each well and the plates incubated at 37 ꢀC for
30 min. Bacterial growth in the wells was indicated by a red
colour, whereas clear wells indicate inhibition.
The basic method for spore germination inhibition was
modified and used to evaluate the activity of various test frac-
tions against fungi. The A. fumigatus, A. flavus and A. niger
were grown on Sabouraud dextrose agar plates and their ho-
mogeneous conidial suspension was prepared in the Sabour-
aud maltose broth. The conidia were counted and their
number in the suspension was adjusted to 1 ꢂ 104 mLꢁ1. Var-
ious concentrations of the test samples in 90 mL of culture me-
dium were prepared in 96-well flat bottom micro-culture
plates (Nunc Nunclon) by double dilution method. The wells
were prepared in duplicates for each concentration. The wells
were inoculated with 10 mL of conidial suspension containing
100 ꢃ 5 conidia. The plates were incubated at 37 ꢀC for 10 h
and then examined for spore germination under inverted mi-
croscope (Nikon Diphot). The number of germinated and
non-germinated conidia was recorded. The percent spore ger-
mination inhibition (PSGI) was calculated using following
formula:
2.5. In vitro cell growth inhibition assay
(cytotoxicity test)
Cells were seeded in 96-well plates at a concentration of
0.1e1.0 ꢂ 104 cells/well in 200 mL of complete media and in-
cubated for 24 h at 37 ꢀC in 5% CO2 atmosphere to allow for
cell adhesion. Stock solutions (4 mM) of the compounds made
in DMSO were filter sterilized, then diluted to 1 mM in incom-
plete media. The 1 mM solutions were further diluted to
500 mM and 50 mM incomplete media for treatment against
HeLa cell lines, where 40e4 mL of compound solutions
were added to 160e196 mL, respectively, of fresh medium in
wells to give final concentrations of 100e1 mM. All assays
were performed in two independent sets of quadruplicate tests.
Control group containing no drug as well as equivalent
amounts of DMSO was run in each assay.
Following 48 h of exposure of cells to drug, each well was
carefully rinsed with 200 mL PBS buffer. Cytotoxicity was as-
sessed using MTT (3-[4,5-dimethylthiazol-2yl]-2,5-diphenyl-
2H-tetrazolium bromide). MTT solutions 20 mL (5 mg/mL
dd H2O) along with 200 mL of fresh, complete media were
added to each well and plates were incubated for 4 h. Follow-
ing incubation, the medium was removed and the purple for-
mazan precipitated in each well was solubilized in 200 mL
DMSO. Absorbance was measured using Techman Magellan
microplate reader (molecular device) at 570 nm and the per-
centage (%) cytotoxicity was calculated as:
No: of conidia germinated in drug treated well
PSGI ¼ 100 ꢁ
No:of conidia germinated in control well
ꢂ 100
2.4. In vitro antibacterial activity
Most of the compounds have been screened in vitro against
Escherichia coli (E. coli). Various methods [28e31] are avail-
able for the evaluation of the antibacterial activity of different
types of drugs. However, the most widely used method [31]
consists in determining the antibacterial activity of the drug
is to add in known concentrations to the cultures of the test
organisms.
2.4.1. Disc diffusion assay
The disc diffusion assay was used to determine antibacte-
rial activity of the drug using gram negative strains of bacteria
E. coli. Base plates were prepared by pouring 10 mL of auto-
claved MullereHinton agar (Biolab) into sterile Petri dishes
(9 cm) and allowing them to settle. Molten autoclaved Mul-
lereHinton that had been kept at 48 ꢀC was inoculated with
a broth culture (106ꢁ108 mLꢁ1) of the test organism and
then poured over the base plate. The discs were air dried
and placed on the top of the agar layer. Four replicants of
each drug tested (four disc per plate) with a gentamycin disc
(0.5 mg/disc) as a reference. The plates were then incubated
for 18 h at room temperature. Antibacterial activity is ex-
pressed as a ratio of the inhibition zone produced by the
drug to the inhibition zone produced by the gentamycin
standard.
O:D: in sample well
% Cytotoxicity ¼ 1 ꢁ
ꢂ 100
O:D: in control well
FCS ¼ Fetal calf serum, PBS ¼ Phosphate buffered saline,
O.D. ¼ Optical density (FCS was obtained from Genetix,
DMSO from cell culture tested, MTT from SRL, and
DMEM were purchased from Sigma, USA).
3. Result and discussion
3.1. Elemental analysis
2.4.2. Micro dilution antibacterial assay
The serial dilution technique was performed using 96-well
microplates to determine the minimum inhibitory concentra-
tion (MIC) of the drugs for antibacterial activity was used.
Elemental analysis (Table 1) reveals the purity of the com-
plexes. All the complexes are soluble in DMSO. The molar
conductance values of the isolated complexes measured in

A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
1243
Table 1
Elemental analysis of the complexes
Complexes
Found (calculated) %
C
H
N
S
Cl
Metal
Pt(L1)2Cl2
Pd(L1)Cl2
Pt(L2)2Cl2
Pd(L2)Cl2
32.37 (32.34)
28.55 (28.57)
34.49 (34.48)
30.60 (30.61)
5.70 (5.69)
5.03 (5.03)
6.02 (6.03)
5.36 (5.36)
12.55 (12.57)
11.15 (11.11)
11.99 (12.07)
10.75 (10.71)
9.60 (9.58)
8.45 (8.47)
9.20 (9.19)
8.15 (8.16)
10.61 (10.63)
18.77 (18.78)
10.25 (10.20)
18.12 (18.11)
29.22 (29.19)
28.00 (28.04)
28.02 (28.01)
27.05 (27.04)
DMSO are found to be less than 15 ohmꢁ1 cm2 molꢁ1 suggest-
ing their non-electrolytic nature.
azomethine nitrogen to the metal ions is supported by dis-
placement of the n(NeN) stretching band frequency. Signifi-
cant changes in the ligand band upon complexation include
a shift in n(C]N). These data indicates coordination through
azomethine nitrogen but no interaction between the terminal
amine nitrogen and the metal ions. The thioamide band in
the range of 750e900 cmꢁ1 is due to n(C]S) as major con-
tributor and n(CeN) as minor. This is shifted to lower fre-
quency on complexation indicates the coordination to metal
ion is through thioamide sulphur C]S. This shift is w120e
150 cmꢁ1, if coordination is through thiol sulphur [34] and
30e40 cmꢁ1, if coordination is through the thione sulphur
[35].
In all the platinum (IV) and palladium (II) complexes the
metal nitrogen vibrations, n(MeN) are assigned to the new
bands [36] in the far IR between 460 and 480 cmꢁ1, while
in the region between 375 and 385 cmꢁ1 gives metalesulphur,
n(MeS) band stretching [37]. The band at w285e300 cmꢁ1 is
assigned due to n(PteCl) and n(PdeCl) stretching vibrations.
3.2. Electronic spectra
The electronic spectra (Table 2) of the thiodiamines (L1 and
L2) show spectral bands because of p / p* and n / p* tran-
sitions. On complexation these bands are shifted. Strong
charge transfer transitions may interfere and prevent the obser-
vation of all the expected bands [32,33]. Strong bands
w340 nm is assignable to a combination of metal ligand
charge transfer (M / LCT) and ded band. The very intense
band w390 nm is assignable to combination of sulphur /
metal charge transfer (Lp / MCT) and ded bands. One ab-
sorption band observed at w285 nm is assigned to the p /
p* intraligand electronic transition, N]C]S. This band
shifts on complexation revealing involvement of C]S group
in complexation, in all the complexes. Additional bands ap-
pear in the complexes because of ded and charge transfer
transitions.
3.4. NMR spectra
3.3. IR spectra
1H NMR spectra of ligands and complexes were recorded
in DMSO-d6 taking TMS as internal standards.
In the present studies, the IR spectra (Table 3) of the thiodi-
amine which contain groups, ꢁNHꢁC]S as a potential bond
forming site. The thioamide group displays prominent IR
bands due to n(NeH), n(CeN) and n(C]S). Coordination of
[L1] d(ppm) 1.16e1.8 (m, 11H, ecyclohexyl), 2.42 (t, 2H, e
CH2), 3.8 (br s, 2H, eNH2), 8.9 (br s, 1H, eNH).
[Pt(L1)2Cl2] d(ppm) 1.22e1.73 (m, 22H, ecyclohexyl), 2.8
(t, 4H, eCH2), 4.03 (br s, 4H, eNH2), 8.79 (br s, 2H, -NH).
[Pd(L1)Cl2] d(ppm) 1.18e1.91 (m, 11H, ecyclohexyl), 2.85
(t, 4H, eCH2), 4.1 (br s, 2H, eNH2), 8.76 (br s, 1H, eNH).
[L2] d(ppm) 1.16e1.77 (m, 11H, cyclohexyleH), 1.4 (m, 2H,
eCH2), 2.2 (t, 4H, eCH2), 4.3 (br s, 2H, eNH2), 9.09 (br s,
1H, eNH).
Table 2
Electronic spectra of the complexes
Complexes
L1
lmax (nm)
Log(3)
212
254
285
272
336
392
292
352
411
213
242
287
274
344
402
276
338
397
3.95
3.22
2.70
3.89
2.32
2.06
3.71
2.79
2.26
4.33
3.96
2.56
3.75
2.40
2.23
3.23
2.81
2.42
[Pt(L2)2Cl2] d(ppm) 1.2e1.8 (m, 22H, cyclohexyleH), 1.92
(m, 4H, eCH2), 2.45 (t, 8H, CH2), 5.1 (br s, 4H, eNH2),
9.11 (br s, 2H, eNH).
Pt(L1)2Cl2
Pd(L1)Cl2
L2
Table 3
IR spectra of the complexes
Complexes
nNeN
nC]S
nMeN
nMeS
nMeCl
L1
1025
1028
1019
1028
1030
1026
886
802
788
889
765
803
e
ꢁ
ꢁ
Pt(L2)2Cl2
Pd(L2)Cl2
Pt(L1)2Cl2
Pd(L1)Cl2
L2
461
465
ꢁ
385
389
ꢁ
295
290
ꢁ
Pt(L2)2Cl2
Pd(L2)Cl2
477
480
382
375
285
297

1244
A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
Table 4
Antifungal studies of the complexes
6
5.9
5.8
5.7
5.6
5.5
5.4
S. No. Complexes Aspergillus fumigatus Aspergillus flavus Aspergillus niger
MDA
MIC
(mg/mL) (mg/mL) (mg/mL) (mg/mL) (mg/mL) (mg/mL)
PSGI
MIC
MDA PSGI
MIC MIC
MDA PSGI
MIC MIC
1
2
Pt(L1)2Cl2 250
Pt(L2)2Cl2 125
250
125
5
250
250
5
250
250
5
500
500
5
500
500
5
Amphoterin B
5
MDA ¼ Micro dilution activity and PSGI ¼ Percent spore germination
1.5
1.7
1.9
2.1
2.3
2.5
2.7
2.9
inhibition.
3
1/T x 10
Fig. 1. The curve plotted by CoatseRedfern method for [Pt(L2)2Cl2] for
step I.
Table 5
Antibecterial studies of the complexes
S. No.
Complexes
Zone of inhibition (mm)
MIC (mg/disc)
Escherichia coli
Escherichia coli
1
Pt(L1)2Cl2
Pt(L2)2Cl2
Pd(L2)Cl2
9
10
8
60.0
60.0
60.0
1.0
2
6.48
6.38
6.28
6.18
6.08
3
Gentamycin
16
[Pd(L2)Cl2] d(ppm) 1.18e1.8 (m, 1H, cyclohexyleH), 1.9
(m, 2H, eCH2), 2.56 (t, 4H, eCH2), 5.2 (br s, 2H, e
NH2), 9.13 (br s, 1H, eNH).
0.8
0.9
1
1.1
1.2
1.3
1.4
1.5
1
The H NMR spectrum of thiodiamines [38,39] shows two
3
1/T x 10
signals at d w9.0e10.3 ppm and d w4.0 ppm, due to the pres-
ence of NH protons.
Fig. 2. The curve plotted by CoatseRedfern method for [Pt(L2)2Cl2] for
step II.
3.5. In vitro antifungal study
In the current study, (Table 4) some synthesized complexes
were tested against pathogenic fungal strains such as A. fumiga-
tus, A. flavus and A. niger. Amphotericin B was used as refer-
ence drug for fungi. The minimum inhibitory concentrations
(MICs) by microbroth dilution assays (MDA) and percent spore
germination inhibition assays (PSGIA) are 125ꢁ500 mg/mL.
The complexes show significant activity at a higher concentra-
tion due to the fact that Aspergillii have hard chitinous outer
wall and therefore, higher concentration of fungicidal com-
pounds may be often required to kill the fungi.
Table 7
Kinetic parameters from TGA for [Pt(L2)2Cl2] complex by CoatseRedfern
method: step I
a
1 ꢁ a
T (K)
T2
1/T ꢂ 10ꢁ3
ꢁlogðꢁlnð1ðꢁaÞÞ=T2Þ
0.39
0.58
0.75
0.89
0.95
0.61
0.42
0.25
0.11
0.05
362
413
492
563
633
131044
170569
242064
316969
400689
2.76
2.42
2.03
1.78
1.58
5.78
5.65
5.60
5.52
5.49
3.6. Antibacterial study
In the current study, (Table 5) some synthesized complexes
were tested against pathogenic bacterial strains such as E. coli
Table 8
Kinetic parameters from TG for [Pt(L2)2Cl2] complex by CoatseRedfern
method: step II
a
1 ꢁ a
T (K)
T2
487204
1/T ꢂ 10ꢁ3
ꢁlogðꢁlnð1ðꢁaÞÞ=T2Þ
Table 6
Cytotoxicity studies of the complexes
0.32
0.41
0.59
0.67
0.77
0.84
0.89
0.68
0.59
0.41
0.33
0.23
0.16
0.11
698
749
1.43
1.33
1.18
1.08
1.01
0.90
0.86
6.46
6.39
6.26
6.25
6.19
6.18
6.14
561001
714025
853776
988036
1227664
1340964
S. No.
Complexes
Concentration (100 mg/mL)
845
924
1
Pt(L1)2Cl2
Pt(L2)2Cl2
Pd(L1)Cl2
54.5
60.9
e
2
994
3
Cisplatin
1108
1158
72.5

A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
1245
Table 9
Thermal data of the thiodiamine complexes
Complexes
Step no.
TG (CoatseRedfern Method)
DH (kJ/g)
Temperature range (K)
n
Ea (kJ molꢁ1
)
S# (J Kꢁ1 molꢁ1
)
[Pt(L2)2Cl2]
I
301e648
648e1173
1
1
4.5
10.24
0.25
0.79
36.34
147.50
II
50
0
using the disc diffusion method. Gentamycin was used as ref-
erence drug for bacteria. The bacterial strains with the zone of
inhibition were observed, 8e10 mm at minimum inhibitory
concentration (MIC) of 60.0 mg/disc.
3.7. Cytototoxic study
-50
-100
In the present studies (Table 6) the cytotoxic study of three
metal complexes have been determined. The study was used to
test the growth inhibition by MTT assay. Data expressed in
terms of percentage (%) cytotoxicity. The metal complexes
caused w60% inhibition. The result shows according to struc-
ture activity rule. It is found that by increasing chain length
from Pt(L1)2Cl2 to Pt(L2)2Cl2, the cytotoxicity increases.
The importance of such work lays the possibility that the
new complexes might more efficacious drug against tumors.
A thorough investigation regarding the structure activity of
the complexes and their stability is required in order to under-
stand the variation in their biological effects. This could be
helpful in designing more potent antitumour agents for thera-
peutic use.
0
200
400
Temperature (°C)
600
800
1000
Fig. 4. DTA curve of [Pt(L2)2Cl2] Complex.
Cl
Cl
N
N
N
S
Pd
Pt
Cl
S
S
Cl
Fig. 5. Proposed structures of the complexes.
4. Thermogravimetric analysis (TGA)/differential
thermal analysis (DTA) study
by the CoatseRedfern method [40]. From the DTA curves,
the heat of reaction was calculated. Kinetic parameters of
each step from CoatseRedfern method were shown in Figs.
1 and 2 and the thermal data were tabulated in Tables 7e9.
4.1. Thiodiamine complexes
The TGA and DTA studies in air atmosphere have been car-
ried out for one complex. Thermal studies were utilized to elu-
cidate the number of kinetic and thermodynamic parameters.
From TGA curve, order of reaction (n), activation energy
(Ea), and apparent activation entropy (S#) were enumerated
4.1.1. [Pt(L2)2Cl2] complex
TGA curve (Fig. 3) shows two steps decomposition. The
first decomposition step (28e375 ꢀC) corresponds to the loss
of all organic moieties for which observed and calculated
weight losses are 61.46% and 61.78%, respectively. The sec-
ond step starts at 375e900 ꢀC corresponds to the platinum
metal residue for which the observed and calculated weight
losses are 71.98% and 71.79%.
The DTA profile (Fig. 4) shows one exotherm at 360 ꢀC
corresponding to the fusion of the compounds and one exo-
therm at 430 ꢀC corresponding to the oxidation of organic
moieties.
2.5
2
1.5
1
5. Conclusion
0.5
0
The electronic spectra of the complexes suggest that all the
complexes are found to be diamagnetic. The platinum (IV)
complexes must be octahedral and palladium (II) complexes
square planar. Platinum (IV) is d6 system and four bands are
0
200
400
600
Temperature ( °C)
800
1000
1
1
Fig. 3. TGA curve of [Pt(L2)2Cl2] Complex.
expected corresponding to A1g / 3T1g, A1g / 3T2g,

1246
A.K. Mishra, N.K. Kaushik / European Journal of Medicinal Chemistry 42 (2007) 1239e1246
1
1A1g / 1T1g and A1g / 1T2g transitions. Palladium (II) is
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that the complexation takes place to the metal ion is through
nitrogen and sulphur. The in vitro antifungal activity of com-
plexes as compared with standard drug Amphotericin B shows
significant activity. The minimum inhibitory concentrations
(MICs) by microbroth dilution assays (MDA) and percent
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