Novel platinum(II) and palladium(II) complexes of thiosemicarbazones derived from 5-substitutedthiophene-2-carboxaldehydes and their antiviral and cytotoxic activities

Article abstract of DOI:10.1016/j.ejmech.2011.09.031

A series of thiosemicarbazones and their platinum(II) and palladium(II) complexes have been synthesized. The chemical structures of ligands and their complexes were characterized by UV-Vis, IR, ¹H NMR, ¹³C NMR, MS spectra, elemental analysis and TGA. The antiviral and cytotoxic activities of all compounds have been tested. Results of broad antiviral evaluation showed that none of the compounds evaluated endowed with anti-DNA or -RNA virus activity at subtoxic concentrations except for the palladium complex 1b. This compound exhibited slightly selective inhibition against cytomegalovirus. The platinum complex 4a exhibited the best cytostatic activities against human cervix carcinoma. Ligands 2, 4 and 5 showed cytostatic potential. The palladium complexes were in general less cytostatic than the corresponding platinum complexes or unliganded congeners.

Full text of DOI:10.1016/j.ejmech.2011.09.031

European Journal of Medicinal Chemistry 46 (2011) 5616e5624
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Novel platinum(II) and palladium(II) complexes of thiosemicarbazones derived
from 5-substitutedthiophene-2-carboxaldehydes and their antiviral
and cytotoxic activities
a
a
a
,
b
_
*
ꢀ
Ays¸egül Karaküçük-Iyidogan , Demet Tas¸ demir , Emine Elçin Oruç-Emre , Jan Balzarini
a Department of Chemistry, Faculty of Arts and Sciences, Gaziantep University, 27310 Gaziantep, Turkey
b Laboratory of Virology and Chemotherapy, Rega Institute for Medical Research, Katholieke Universtiteit Leuven, 3000 Leuven, Belgium
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of thiosemicarbazones and their platinum(II) and palladium(II) complexes have been synthe-
sized. The chemical structures of ligands and their complexes were characterized by UVeVis, IR, 1H NMR,
13C NMR, MS spectra, elemental analysis and TGA. The antiviral and cytotoxic activities of all compounds
have been tested. Results of broad antiviral evaluation showed that none of the compounds evaluated
endowed with anti-DNA or -RNA virus activity at subtoxic concentrations except for the palladium
complex 1b. This compound exhibited slightly selective inhibition against cytomegalovirus. The plat-
inum complex 4a exhibited the best cytostatic activities against human cervix carcinoma. Ligands 2, 4
and 5 showed cytostatic potential. The palladium complexes were in general less cytostatic than the
corresponding platinum complexes or unliganded congeners.
Received 26 May 2011
Received in revised form
16 September 2011
Accepted 20 September 2011
Available online 1 October 2011
Keywords:
Thiosemicarbazones
Platinum(II) complexes
Palladium(II) complexes
Cytotoxic activity
Antiviral activity
Ó 2011 Elsevier Masson SAS. All rights reserved.
1. Introduction
of metal containing agents, from main group elements to early
transition metals, have been evaluated [12e14]. Especially organo-
Since the discovery of the importance of metal containing
compounds used in cancer treatment, reports on the use of metals
are increasing [1e5]. In recent years new metal complexes have been
identified as a very promising class of anticancer active compounds
[6e8]. Metals bound to atoms such as N, O and S can form a chelate
ring that binds the metal more tightly when compared to the non-
chelate form. Large biological molecules (proteins, enzymes, DNA
etc.) are electron-rich but metal ions are electron-deficient. There-
fore, interactions occur between metal ions and many important
biological molecules. This event has led to the use of metals or metal-
containing agents to modulate biological systems [9]. Organometallic
compounds have gained importance as enzyme inhibitors due to the
capability of binding large biological molecules more strongly than
metal-free organic compounds [10].
platinum compounds such as cisplatin, carboplatin and oxaliplatin
are metal-based drugs that are among the most active and widely
used clinical drugs in cancer chemotherapy [15e17]. These plat-
inum complexes react in vivo, binding to and causing crosslinking of
DNA which finally activates programmed cell death [18]. However,
the clinical utility of these drugs is limited to a relatively narrow
range of tumors (sarcomas, small cell lung cancer, ovarian cancer,
lymphomas and germ cell tumors) because of primary resistance
and the development of resistance secondary to the initial treat-
ment [19,20]. Therefore, unconventional platinum complexes, that
could be used in cisplatin-resistant tumors are made by different
research groups [21e24].
Lipophilicity that controls the rate of entry into the cell is altered
by metal-coordination and some side-effects can be reduced by
complexation. Moreover, metal complexes can exhibit biological
activities more than free ligand [6]. Recently, thiosemicarbazones and
their metal complexes have achieved importance due to their appli-
cation in pharmaceutical chemistry and proved to be chemothera-
peutic agents potentially useful for inhibiting the activities of cancer
cells [25,26]. For example, 3-aminopyridine-2-carboxaldehyde thio-
semicarbazone (TriapineÒ; Vion Pharmaceuticals Inc. New Haven, CT)
inhibit the biosynthesis of DNA in leukemia L1210 cells by blocking
Metal complexes can inhibit metalloenzymes by chelate
substitution and also inhibit non-metalloenzymes by coordinating
to their active site. The other property of metals is to catalyze the
formation of reactive oxygen species [11]. The anticancer potential
* Corresponding author. Tel.: þ90 342 317 18 30; fax: þ90 342 360 10 32.
E-mail address: oruc@gantep.edu.tr (E.E. Oruç-Emre).
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.09.031

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A. Karaküçük-Iyidogan et al. / European Journal of Medicinal Chemistry 46 (2011) 5616e5624
5617
activity of ribonucleotide reductase [27]. Many heterocyclic thio-
H
N
semicarbazone derivatives and their platinum and palladium comp-
lexes have a wide range of pharmacological activities, such as anti-
tuberculosis [28], antibacterial [29], antitumor [30], antiprotozoal
[31], antimalarial [32], antimicrobial [33], antiviral [34], antifungal
[35], anticonvulsant [36] and anti-trypanosomal [37] activities.
In our present work, the synthesis and characterization of
thiosemicarbazones derived from 5-substitutedthiophene-2-
carboxaldehydes and their platinum(II) and palladium(II)
complexes are reported. The results of in vitro antiviral and
cytostatic/toxic activities of ligands and their platinum and
palladium complexes have been evaluated.
N
S
R'
N
S
R
M
Cl
Cl
M=Pt(II) or Pd(II)
Fig. 1. The structures of Pt(II) and Pd(II) complexes (1ae5a and 1be5b).
2. Chemistry
3. Results and discussion
The precursors phenyl isothiocyanate and 4-nitrophenyl iso-
thiocyanate were synthesized from aniline and 4-nitroaniline
according to the method described in the literature [38]. N-phe-
nylhydrazinecarbothioamide and N-(4-nitrophenyl)hydrazine-
carbothioamide were prepared by stirring isothiocyanates with
hydrazine monohydrate in diethyl ether at room temperature
according to Ref. [39]. The ligands 1e5 used in this work were ob-
tained by refluxing in methanol (20e30 mL) an equimolar amount
of 5-substituted-2-thiophene carboxaldehydes with thiosemicarba-
zides according to Refs. [35,40e42]. The chemical structures of the
ligands are given in Scheme 1. All thiosemicarbazone derivatives
synthesized were characterized by UVeVis, IR,1H NMR,13C NMR, MS
spectral data and elemental analysis.
3.1. Synthesis
All of the newly synthesized metal complexes 1a-5a and 1b-5b
are solids and decomposed above ca. 200 ꢀC. They are insoluble in
organic solvents such as acetone, chloroform and methanol but
soluble in DMF and DMSO. The elemental analyses data of the
thiosemicarbazones and their platinum and palladium complexes
(Table 1) were compatible with the structures of the ligands shown
in Scheme 1. and the formulas of the complexes are shown in Fig. 1.
3.2. Spectral studies
The platinum(II) and palladium(II) complexes of thiosemicar-
bazone derivatives 1ae5a and 1be5b were prepared by exposing
a solution of the K2PtCl4 or K2PdCl4 in water to a solution of the
appropriate ligand in ethanol in a 1:1 M:L molar ratio [43]. The
structures of Pt(II) and Pd(II) complexes were confirmed by
elemental analysis, UVeVis, IR, 1H NMR, MS spectroscopies, eleme-
ntal analyses and TGA thermogram and are given in Fig. 1. The
elemental analysis data of the ligands and their Pt(II) and Pd(II)
complexes are presented in Table 1.
3.2.1. Electronic spectral studies
The study of the electronic spectra in the ultraviolet and visible
(UVeVis) ranges for the ligand and metal complexes was carried
out in DMF. Solid-state electronic spectra of all thiosemicarbazone
derivatives reveal similar patterns, exhibiting two bands in the
433e344 nm and 28e258 nm regions. An intense band at ca.
433e344 nm is attributed to the n /
C]N group and thiophene ring, which are overlapped. The
p
* transitions of C]S group,
p*
p
/
H
N
H
N
NCS
NH2
NH2NH2.H2O
Et2O, rt
CSCl2, NaOH
CHCl3, rt
NH2
S
R
R
R
O
S
MeOH, reflux
H
R'
H
N
H
N
S
R'
N
S
R
1 R=H, R'=H
2 R=H, R'=Cl
3 R=H, R'=C6H5
4 R=H, R'=NO2
5 R=NO2, R'=H
Scheme 1. General synthesis of thiosemicarbazones (1e5).

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5618
Table 1
Physical data of thiosemicarbazone derivatives and their Pt(II) and Pd(II) complexes.
Comp. no
Molecular formulas
Colour
Yield (%)
Mp/Dec. temp. (ꢀC)
Found (calcd.)
C
H
N
S
1
C12H11N3S2
Yellow
85
69
58
67
53
62
83
56
59
82
52
54
83
62
61
188e190
212
55.14
(55.29)
27.38
(27.43)
32.93
(32.27)
47.86
(47.72)
25.65
(25.43)
30.53
(30.28)
64.06
(64.72)
35.89
(35.62)
42.08
(42.19)
47.04
(47.22)
25.23
(25.43)
29.86
(29.43)
47.04
(47.16)
25.23
(25.61)
29.86
(29.67)
4.24
(4.28)
1.91
(1.98)
2.30
(2.35)
3.21
(3.41)
1.79
(1.58)
1.92
(1.81)
4.48
(4.41)
2.34
(2.02)
2.75
(2.82)
3.29
(3.34)
1.59
(1.19)
1.88
(1.89)
3.29
(3.25)
1.59
(1.65)
1.88
(1.75)
16.08
(16.16)
7.98
(7.82)
9.60
(9.53)
12.69
(12.21)
7.48
(7.82)
8.90
(8.62)
12.45
(12.21)
6.97
(6.73)
8.18
(8.33)
18.29
(18.28)
9.81
(9.66)
11.61
(11.66)
18.29
(18.32)
9.81
24.54
(24.32)
12.18
(12.67)
14.65
(14.45)
22.62
(22.58)
11.41
(11.67)
13.58
(13.85)
19.00
(19.28)
10.64
(10.48)
12.48
(12.82)
20.93
(20.79)
11.22
(11.83)
13.29
(13.72)
20.93
(20.91)
11.22
(11.43)
13.29
(13.34)
1a
1b
2
C12H10N3S2Cl2Pt
C12H10N3S2Cl2Pd
C12H10N3S2Cl
Orange
Brown
272
Yellow
130e132
243
2a
2b
3
C
12H9N3S2Cl3Pt
Dark yellow
Brown
C12H9N3S2Cl3Pd
C18H15N3S2
250
Yellow
182e184
256
3a
3b
4
C18H14N3S2Cl2Pt
Orange
Brown
C
18H14N3S2Cl2Pd
307
C12H10N4O2S2
Yellow
190e192
220
4a
4b
5
C12H9N4O2S2Cl2Pt
C12H9N4O2S2Cl2Pd
Dark brown
Dark brown
Yellow
302
C
12H10N4O2S2
188e190
225
5a
5b
C12H9N4O2S2Cl2Pt
C12H9N4O2S2Cl2Pd
Orange
Brown
(9.96)
11.61
(11.84)
296
transitions of the thiophene ring and thiosemicarbazone imine
function are rather weak, and observed at ca. 281e258 nm. These
two bands are shifted to lower energies (bathochromic shift) after
complexation (467e371 nm, 344e264 nm). Such observations have
also been reported earlier in other platinum(II) and palladium(II)
complexes of similar ligand systems [44].
Although thiosemicarbazone derivatives are capable of inter-
acting with metals in different ways, they usually behave as neutral
NS donor bidentate ligands. The thiosemicarbazones can exhibit
thione-thiol tautomerism in solution because of the thioamide
(NHeC]S) function. The n
(SH) band at ca. The 2500e2600 cmꢁ1
region is absent from the IR spectra of the ligands but the v(NH)
band at 3115e3140 cmꢁ1 and the strong band due to v(C]S) at
783e825 cmꢁ1 are present, suggesting that in the solid phase the
thiosemicarbazones remain as the thione form [45]. These bands
are shifted to lower wavenumbers (13e66 cmꢁ1) indicating the
coordination of metal through thione/thiol sulfur in all complexes.
This coordination is approved by the presence of two new bands at
the 502e521 and 400e418 cmꢁ1 region due to v(PteN, PdeN and
PteS, PdeS), respectively [46]. The band due to v(CeSeC) of the
thiophene ring remains unaltered in complexes (1ae5a and
1be5b), indicating non-participation of the thiophene ring in
3.2.2. IR spectral studies
IR spectral data of all thiosemicarbazones and their Pt(II) and
Pd(II) complexes showed a holding the 5-substitutedthiophene-2-
carboxaldehyde thiosemicarbazone moiety in all complexes. IR
spectra were compared with free ligands and their Pt(II) and Pd(II)
complexes and given in Table 2.
Table 2
Main characteristic IR vibrational bands of thiosemicarbazones and their plati-
num(II) and palladium(II) complexes.
coordination. The azomethine stretching vibrations n(C]N), char-
acteristic of a Schiff base are observed at 1582e1595 cmꢁ1 [7]. The
negative shift (30e78 cmꢁ1) of the C]N band was observed in all
complexes which exhibit the involvement of azomethine nitrogen
upon complexation. On the other hand, the shift to higher wave-
numbers of the v(NeN) band, observed for the platinum and
palladium complexes, have also been related to the electronic
delocalization and occurred as a consequence of coordination
through the azomethine nitrogen atom and deprotonation of thi-
osemicarbazones [47]. Finally, the ligands 1e5 coordinate to plati-
num(II) and palladium(II) as NS bidentate.
Compound
vmax (cmꢁ1
NeH
)
C]N
NeN
C]S
MeN
MeS
1
3295
3272
3242
3316
3298
3205
3304
3288
3241
3342
3320
3255
3360
3289
3262
1588
1510
1527
1591
1524
1533
1592
1528
1527
1595
1565
1538
1582
1533
1547
1042
1095
1073
1064
1088
1105
1028
1049
1072
1110
1046
1099
1108
1172
1181
783
748
717
787
747
748
804
791
749
825
807
778
800
776
769
e
e
1a
1b
2
2a
2b
3
3a
3b
4
4a
4b
5
521
518
e
400
403
e
513
511
e
402
405
e
507
502
e
418
412
e
3.2.3. NMR spectral analysis
518
510
e
411
408
e
The chemical structures of all thiosemicarbazones and their
complexes were characterized by 1H NMR spectroscopy and given
in Table 3. Platinum(II) and palladium(II) complexes exhibited
similar 1H NMR spectra and behavior.
5a
5b
516
509
409
410

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5619
Table 3
major fragmentation pathway involved the leaving of PtCl2 or
PdCl2 [53].
1H NMR chemical shift values of (
complexes.
d) ligands and their platinum(II) and palladium(II)
Compound
d (ppm)
3.2.5. Thermal decomposition
TGA thermograms of complexes (1ae5a and 1be5b) were
recorded under nitrogen with a heating rate of 10 ꢀC/min between
30 ꢀC and 900 ꢀC. All complexes did not lose weight up to 230 ꢀC.
Further increment of the temperature causes decomposition of
these complexes in two steps [54]. The first step being 250e415 ꢀC
for the palladium(II) and platinum(II) complexes where losses of
mixed fragments are observed. The second step starts after the first
step and continues until complete decomposition of the ligands
and formation of MS [where, M ¼ Pd(II) and Pt(II)]. The total weight
loss of all complexes depends on the loss of the respective ligand,
after one sulfur atom moving to the metal ion and residues
conversion to the palladium sulfide and platinum sulfide.
CSNH
PhNH
N]CH
AreH
1
11.83
e
9.81
9.94
9.83
9.86
9.85
9.80
9.84
9.81
9.79
10.19
10.12
10.27
10.23
10.69
10.26
8.36
8.48
8.66
8.24
8.62
8.55
8.33
8.62
8.64
8.31
8.63
8.36
8.40
9.09
8.91
7.70e7.15
8.00e6.92
8.02e7.09
7.54e7.16
7.92e6.93
7.68e7.18
7.73e7.21
7.78e6.98
8.00e7.33
8.11e7.24
8.18e7.02
8.31e7.20
8.25e7.17
8.12e7.64
8.38e7.22
1a
1b
2
2a
2b
3
3a
3b
4
4a
4b
5
e
11.89
e
e
11.90
e
e
12.19
e
e
11.91
e
5a
5b
e
3.3. Biological activity studies
The biological activity of the newly synthesized complexes were
tested in terms of cytotoxic and antiviral activity compared with
the free ligands.
Results of the 1H NMR integrations and signal multiplicities
were in agreement with the proposed structures and other spectral
data. 1H NMR spectra of all thiosemicarbazones (1e5) recorded in
3.3.1. Cytostatic/toxic activity
DMSO-d6 exhibited a sharp singlet peak at
the PhNH proton and at
11.83e12.19 ppm due to the ¼ NeNH
proton, which indicate that even in a polar solvent they remain in
the thione form. A singlet peak at 8.24e8.40 ppm due to the HC]
d
9.81e10.23 ppm due to
The all compounds were examined for their cytostatic activity
against human lymphocyte (CEM), human cervix carcinoma (HeLa),
Crandell feline kidney (CRFK), Madin Darby canine kidney (MDCK)
and human lung fibroblast (HEL) cells and their activity results
were given Table 4.
Among all ligands and their platinum and palladium
complexes evaluated the platinum complex of 5-nitrothiophene-
2-carboxaldeyde-N(4)phenyl thiosemicarbazone 4a showed the
d
d
N
group was in accordance with the literature [48e50].
The ¼ NeNH proton signals disappeared in the 1H NMR spectra of
the platinum(II) and palladium(II) complexes indicating the
deprotonation of the ¼ NeNH group, while the signals of the other
NH (PhNH) protons were observed at the same field
(d
highest cytostatic activity against HeLa (IC50: 1.7
mM IC50: 0.22 mM
9.80e10.69 ppm). The signals of the HC]N protons shift to
downfield in all complexes and appeared at 8.48e9.09 ppm. This
information indicates the coordination of the metal center to the
azomethine nitrogen and the thioamide sulfur and the PhNH group
is not coordinated to the metal ion [51]. All aromatic protons were
for cisplatinum). However this compound was cytostatic against
proliferating HEL fibroblasts (IC50: 3.1
m
M), CRFK cells (IC50
:
:
7.5
2.2
m
m
M), MDCK cells and (IC50: 3.4
M for cisplatinum). Furthermore, the ligands 2, 4 and 5
m
M) CEM cells (IC50: 8.1 M IC50
m
showed moderately cytostatic activity against in cell cultures
(IC50: 7.1e57.8 M for compound 2, IC50: 3.6e52.1 M for
compound 4 and IC50: 2.4->100 M for compound 5). Other
observed at the expected region (
d
7.15e8.25 ppm) in the NMR
m
m
spectrum of the thiosemicarbazone ligands. These signals do not
afford relevant changes in the chemical shifts for the platinum(II)
and palladium(II) complexes.
m
ligands and complexes did not show appreciable cytostatic action
against the evaluated confluent monolayer cell cultures (Table 4).
Furthermore, the 13C NMR spectra of the ligands were recorded
in DMSO-d6 and gave the spectral signals in good agreement with
the probable chemical structure. All ligands exhibited two impor-
Regarding toxicity, it is striking that the ligands 2e5 (IC50
2.4e163 M)andtheirplatinumcomplexes1ae5a(IC50:1.7e105 M)
were consistently more cytostatic than palladium complexes 1be5b
(IC50: 13.3e385 M). Also, these compounds are clearly cytostatic
:
m
m
tant signals at
to the thioamide (C]S) and imine (C]N) carbon atoms, respec-
tively [52]. The signals at 123.60e156.22 ppm were observed in
d
174.22e176.21 and
d
137.28e140.65 ppm assigned
m
(inhibition of cellproliferation) ratherthancytotoxic (microscopically
visible alteration of cell morphology).
d
the spectra due to the aromatic carbons.
3.3.2. Antiviral activity
3.2.4. Mass analysis
The compounds 2e5, platinum(II) complexes 1ae5a and palla-
dium (II) complexes 1be5b were also evaluated for their antiviral
activity against a wide variety of DNA and RNA viruses, including
herpes simplex virus type 1 (HSV-1) (strain KOS), HSV-2 (strain G),
vaccinia virus, vesicular stomatitis virus (VSV), varicella-zoster
virus (VZV) strains OKA and 07/1 and human cytomegalovirus
(HCMV) strains AD-169 and Davis in HEL cell cultures, VSV, Cox-
sackie B4 and respiratory syncytial virus (RSV) in HeLa cell cultures,
parainfluenza-3, reovirus-1, Sindbis virus, Coxsackie B4 and Punta
Toro virus in Vero cell cultures, influenza A (H1N1, H3N2) influenza
B virus in MDCK cell cultures and feline coronavirus (FIPV) and
feline herpes virus in CrFK cell cultures and their activities were
compared with those of ganciclovir and cidofovir. Unfortunately,
none of the compounds tested were endowed with appreciable
inhibitory activity at subtoxic concentrations except for compound
The MS spectra of the free ligands (1e5) and their complexes
(1ae5a and 1be5b) were recorded by electro spray impact (ESI) in
positive ion mode. The mass fragmentation routes of compound 2 is
given in Scheme 2.
The mass spectrum of compound 2 confirms the proposed
molecular structure by showing a peak at m/z 296 corresponding to
the molecular ion [M þ H]þ. The other thiosemicarbazone deriva-
tives also showed the similar fragmentation pathway as given in
Fig. 2.
A general splitting pathway with the characteristic peaks was
observed by the platinum(II) and palladium(II) complexes 1ae5a
and 1be5b. The spectra of all complexes showed several infor-
mative fragment ions confirming their molecular weights. The
molecular ion peaks were conformed to [M]þ or [M þ H]þ and the

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.
+
H
N
H
.
+
N
NH2
N
N
S
m/z = 120
m/z = 215
+
S
-
-
H
.
H
N
+
H
H
N
H
NH2
+
N
N
S
S
m/z = 152
m/z = 216
.
S+
+H
-
HC
H
N
H
H
N
.
+
.
+
NH
N
S
N
S
S
m/z = 151
m/z = 261
.
Cl
-
H
N
H
N
.
+
S
N
Cl
S
m/z = 296
HC
-
CH
S
-
H
N
H
N
.
+
S
.
+
N
H
S
Cl
HC
.
S
+
N
N
N
S
Cl
Cl
m/z = 270
H
m/z = 131
+2H
m/z = 264
C H
.
.
-
6
5
.
+
.
+H
S
Cl
H3C
H
N
.
+
N
S
m/z = 133
N
H
Cl
H
m/z = 188
.
Cl
-
H
N
N
N
S
H
+
H
m/z = 152
Scheme 2. The mass fragmentation routes of compound 2.
1b that showed the highest antiviral activity against HCMV (EC50
:
2.9e15.3 M). By comparison of the ganciclovir potency against
m
HCMV we can conclude that 1b was 3-fold less active against HCMV
AD-169 strain and almost equally potent against Davis strain as
B
D
C
A
H
H
N
ganciclovir (EC50: 5.0
strain). However, it should be mentioned that this compound was
also cytostatic at CC50 value of 13.3 M in HEL cell cultures.
mM for AD-169 strain, EC50: 3.4 mM for Davis
S
R
N
m
N
E
Therefore, the observed anti-HCMV activity may well be due to an
indirect cytostatic activity of the compounds rather than a specific
antiviral activity. No specific antiviral effect was noted for any
compounds evaluated against other viruses (data not given).
S
R'
Fig. 2. Proposed mass fragmentation pathway for ligands.

_
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A. Karaküçük-Iyidogan et al. / European Journal of Medicinal Chemistry 46 (2011) 5616e5624
5621
Table 4
(5.98 mmol) in methanol (5 mL) during 15 min. The reaction
mixture was refluxed and the progress of the reaction was followed
by TLC (about 10 h) and cooled. After cooling the product was
filtered and the filtrate was concentrated under reduced pressure.
The crude product was recrystallized from methanol.
Cytotoxic and cytostatic activity of the ligands and their platinum(II) and palladiu-
m(II) complexes.
MICa
HEL
(
m
M)
IC50
(
mM)
b
Compound
HeLa
MDCK
HEL
6.0
CrFK
HeLa
CEM
1a
1b
2
2a
2b
3
3a
3b
4
4a
4b
5
ꢂ20
100
100
>100
100
100
>100
ꢂ20
100
100
ꢂ20
20
10.8
>100
57.8
12.6
>100
>100
60.7
100
10.9
3.4
55.3
>100
>100
>100
6.7
37
22
7.1
37
180
43
105
254
3.6
1.7
21
55
34
10
36
60
163
53
168
6.4
8.1
30
27
26
5.1.2. Thiophene-2-carboxaldehyde-N(4)phenyl thiosemicarbazone
(1)
13.3
10.4
26.2
42.8
32.1
52.7
100
52.1
3.1
91.9
10.1
15.6
>100
>100
65.0
>100
3.8
7.5
39.1
2.4
>100
>100
ꢂ100
60
Yellow solid (methanol). Yield: 85%. m.p.: 188e190 ꢀC. Anal. calc.
(C12H11N3S2): C, 55.14; H, 4.24; N,16.08; S, 24.54; found: C, 55.29; H,
4.28; N, 16.16; S, 24.32%. ES-MS (m/z) 261 [Mþ], 262 [M þ H]þ. UV/
Vis lmax(nm): 260, 344. IR vmax(cmꢁ1): 3295, 3132 (NeH); 1588
(C]N); 1170, (CeN); 1042 (NeN); 783 (C]S). 1H NMR (DMSO-d6,
100
ꢂ100
100
ꢂ100
ꢂ20
20
ꢂ100
ꢂ20
ꢂ100
>100
d
ppm): 11.83 (s, 1H, CSNHN); 9.81 (s, 1H, PhNH); 8.36 (s, 1H, HC]
100
20
>100
>100
14.6
3.2
34
N); 7.70 (d,1H, J ¼ 4.99 Hz, C2 proton of thiophene ring); 7.58 (d, 2H,
J ¼ 7.82 Hz, ArH ortho position); 7.54 (d, 1H, J ¼ 3.29 Hz C4 proton of
thiophene ring); 7.37 (t, 2H, ArH meta position); 7.20 (t, 1H, ArH
para position); 7.15 (dd, 1H, J1 ¼ 3.73 Hz, J2 ¼ 4.85 Hz, C3 proton of
12
46
385
5a
5b
51.7
152
a
Minimal inhibitory concentration or compound concentration required to alter
thiophene ring). 13C NMR (DMSO-d6,
d ppm): 174.22 (C]S), 144.68
microscopically visible morphology of the confluent cell cultures.
b
50% Inhibitory concentration or compound concentration requred to inhibit
(ArC), 140.65 (C]N), 138.52, 130.05, 129.13, 128.44, 127.91, 127.16,
126.55 (ArC).
MDCK (Madin Darby canine kidney), HEL (human lung fibroblast), CrFK (Cranell
feline kidney), HeLa (human cervic carcinoma) or CEM (human lymphocyte) cells by
50%.
5.1.3. 5-Chlorothiophene-2-carboxaldehyde-N(4)phenyl
thiosemicarbazone (2)
Yellow solid (methanol). Yield: 67%. m.p.: 130e132 ꢀC. Anal. calc.
(C12H10ClN3S2): C, 47.86; H, 3.21; N, 12.69; S, 22.62; found: C, 47.72;
H, 3.41; N, 12.21; S, 22.58%. ES-MS (m/z) 296 [M þ Hþ]. UV/Vis
lmax(nm): 258, 351. IR nmax(cmꢁ1): 3316, 3132 (N-H), 1591 (C]N),
4. Conclusions
We have prepared
a series of 5-substitutedthiophene-2-
carboxaldeyhde thiosemicarbazones and their platinum(II) and
palladium(II) complexes. In conclusion, none of the new compounds
showed antiviral potency at subtoxic concentrations except for 1b
that showed slight inhibitory activity against HCMV at 2.9e15.3
Several compounds in particular 4a were endowed with significant
cytostatic potency (IC50: 1.7 M for HeLa cell cultures) and can be
viewed as new lead compounds for further modifications. The other
ligands and their platinum complexes were showed only moderate
cytostatic activity in the lower micromolar range.
1198 (CeN), 1064 (NeN), 787 (C]S). 1H NMR (DMSO-d6,
d ppm):
11.89 (s, 1H, CSNHN); 9.86 (s,1H, PhNH); 8.24 (s,1H, HC]N); 7.54 (t,
2H, J ¼ 8.42 Hz, ArH meta position) 7.40 (d,1H, J ¼ 4.05 Hz, C3 proton
of thiophene ring); 7.36 (d, 2H, J ¼ 8.23 Hz, ArH ortho position);
7.20 (t, 1H, ArH para position); 7.16 (d, 1H, J ¼ 3.94 Hz, C4 proton of
mM.
m
thiophene ring). 13C NMR (DMSO-d6,
d ppm): 176.14 (C]S), 156.22
(ArC), 139.42 (C]N), 138.30, 137.90, 137.65, 134.37, 133.78, 131.59,
131.20, 128.55, 126.08 (ArC).
5. Experimental
5.1.4. 5-Phenylthiophene-2-carboxaldeyde-N(4)phenyl
thiosemicarbazone (3)
5.1. Chemicals and instruments
Yellow solid (methanol). Yield: 83%. m.p.: 182e184 ꢀC. Anal. calc.
(C18H15N3S2): C, 64.06; H, 4.48; N, 12.45; S, 19.00; found: C, 64.72;
H, 4.41; N, 12.21; S, 19.28%. ES-MS (m/z) 337 [Mþ]. UV/Vis lmax(nm):
266, 378. IR nmax(cmꢁ1): 3304, 3115 (NeH); 1592 (C]N); 1160
All reactions were monitored by thin layer chromatography
(TLC) using Merck silica gel 60 F254 plates. Chemicals and solvents
purchased from Aldrich, Fluka and Riedel de Haen. Melting points
were determined by EZ-Melt melting point apparatus and were
uncorrected. Electronic spectra were recorded in DMF on a PG
Instruments T80þ UVevisible Spectrophometer. IR spectra on KBr
disks were recorded on a Perkin Elmer 1620 model FT-IR spectro-
photometer. Elemental analyses (C,H,N,S) were performed on
a VarioMICRO elemental analyzer. 1H and 13C NMR spectra were
(CeN); 1028 (NeN); 804 (C]S). 1H NMR (DMSO-d6,
d ppm): 11.90
(s, 1H, CSNHN); 9.84 (s, 1H, PhNH); 8.33 (s,1H, HC]N); 7.73 (d, 2H,
J ¼ 7.29 Hz, ArH, ortho position of phenyl bearing to thiophene
ring); 7.60 (d, 2H, J ¼ 7.45 Hz, ArH, ortho position); 7.58 (d, 1H,
J ¼ 3.87 Hz, C3 proton of thiophene ring); 7.55 (d,1H, J ¼ 3.90 Hz, C4
proton of thiophene ring); 7.34e7.48 (m, 5H, ArH, meta position
and para and meta position phenyl bearing to thiophene ring); 7.21
obtained at room temperature with
a
Brucker AVANC-DPX-
( t, 1H, ArH, para position). 13C NMR (DMSO-d6,
d ppm): 175.42 (C]
400 MHz NMR spectrometer in DMSO-d6 using TMS as an internal
standard. The chemical shift values are given in ppm and following
abbreviations were used: s ¼ singlet, d ¼ doublet, dd ¼ double
doublet, t ¼ triplet, m ¼ multiplet. The coupling constants (J) are
given in Hertz. The mass spectra of all compounds were recorded
on an Agilent 1100 MSD spectrometer in the electro spray mode.
Thermogravimetric analysis of the complexes were performed by
a Shimadzu TGA 50 thermogravimetric analyzer under nitrogen
atmosphere with a heating rate of 10 ꢀC/min.
S), 147.56 (ArC), 137.28 (C]N), 136.91, 133.46, 132.54, 131.17, 129.02,
128.75, 128.43, 126.12, 125.89, 124.38, 123.60 (ArC).
5.1.5. 5-Nitrothiophene-2-carboxaldeyde-N(4)phenyl
thiosemicarbazone (4)
Yellow solid (methanol). Yield: 82%. m.p.: 190e192 ꢀC. Anal. calc.
(C12H10N4O2S2): C, 47.04; H, 3.29; N,18.29; S, 20.93; found: C, 47.22;
H, 3.34; N, 18.28; S, 20.79%. ES-MS (m/z) 306 [Mþ]. UV/Vis
lmax(nm): 281, 433. IR nmax(cmꢁ1): 3342, 3140 (NeH); 1595 (C]N);
1348 (eNO2); 1162 (CeN); 1110 (NeN); 825 (C]S). 1H NMR (DMSO-
5.1.1. Synthesis of thiosemicarbazones (1e5): a general method
To a hot solution of 4-substituted-N-phenylthiosemicarbazide
(5.98 mmol) in methanol (30 mL) was added dropwise a solution of
the corresponding 5-substituted-2-thiophene carboxaldehydes
d6, d ppm): 12.19 (s, 1H, CSNHN); 10.19 (s, 1H, PhNH); 8.31 (s,1H,
HC]N); 8.11 (d, 1H, J ¼ 4.36 Hz, C3 proton of thiophene ring); 7.62
(d, 1H, J ¼ 4.40 Hz, C4 proton of thiophene ring); 7.51 (d, 2H,
J ¼ 7.42 Hz, ArH ortho position); 7.38 (t, 2H, J1 ¼ 7.55 Hz ve

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A. Karaküçük-Iyidogan et al. / European Journal of Medicinal Chemistry 46 (2011) 5616e5624
5622
J2 ¼ 8.17 Hz, ArH meta position); 7.24 (t, 1H, J ¼ 8.46 Hz, ArH para
position). 13C NMR (DMSO-d6,
ppm): 175.84 (C]S), 144.22 (ArC),
138.25 (C]N), 137.29, 130.48, 129.36, 128.34, 127.98, 127.02, 126.13
(ArC).
5.1.12. Pt(C12H9N4O2S2)Cl2 (5a)
d
Orange solid (ethanol). Yield 62%. m.p.: 225 ꢀC (with decom-
position). Anal. calc. (C12H9N4O2S2Cl2Pt): C, 25.23; H, 1.59; N, 9.81;
S, 11.22; found: C, 25.61; H, 1.65; N, 9.96; S, 11.43%. ES-MS (m/z) 571
[Mþ]. UV/Vis lmax(nm): 344, 419. IR nmax(KBr, cmꢁ1): 3289 (NeH);
1533 (C]N); 1172 (NeN); 776 (CeS); 516 (PteN); 409 (PteS). 1H
5.1.6. Thiophene-2-carboxaldehyde-N(4)nitrophenyl
thiosemicarbazone (5)
NMR (DMSO-d6, d ppm): 10.69 (s, 1H, PhNH); 9.09 (s, 1H, HC]N);
Yellow solid (methanol). Yield: 83%; m.p.: 188e190 ꢀC. Anal.
calc. (C12H10N4O2S2): C, 47.04; H, 3.29; N, 18.29; S, 20.93; found: C,
47.16; H, 3.25; N, 18.32; S, 20.91%. ES-MS (m/z) 306 [Mþ]. UV/Vis
lmax(nm): 260, 353. IR: nmax(cmꢁ1): 3360, 3142 (NeH); 1582 (C]
N); 1342 (eNO2); 1171 (CeN); 1108 (NeN); 800 (C]S). 1H NMR
8.12e7.64 (m, 7H, ArH).
5.1.13. Pd(C12H10N3S2)Cl2 (1b)
Brown solid (ethanol). Yield: 58%. m.p.: 272 ꢀC (with decom-
position). Anal. calc. (C12H10N3S2Cl2Pd): C, 32.93; H, 2.30; N, 9.60; S,
14.65; found: C, 32.27; H, 2.35; N, 9.53; S, 14.45%. ES-MS (m/z) 438
[Mþ]. UV/Vis lmax(nm): 262, 368. IR nmax(KBr, cmꢁ1): 3242 (N-H);
1527 (C]N); 1073 (NeN); 717 (CeS); 518 (PdeN); 403 (PdeS). 1H
(DMSO-d6,
d ppm): 11.91 (s, 1H, CSNHN); 10.23 (s,1H, PhNH); 8.40
(s,1H, HC]N); 8.25 (d, 2H, J ¼ 7.15 Hz, ArH ortho position of nitro
group); 8.04 (d, 2H, J ¼ 7.11 Hz, ArH, meta position of nitro group);
7.74 (d, 1H, J ¼ 4.99 Hz, C2 proton of thiophene ring); 7.58 (d, 1H,
J ¼ 3.60 Hz, C4 proton of thiophene ring); 7.17 (dd, 1H, J1 ¼ 3.64 Hz,
J2 ¼ 4.93 Hz, C3 proton of thiophene ring). 13C NMR (DMSO-d6,
NMR (DMSO-d6,
d ppm): 9.83 (s, 1H, PhNH); 8.66 (s, 1H, HC]N);
8.02e7.09 (m, 8H, ArH).
d
ppm): 176.21 (C]S), 145.18 (ArC), 139.42 (C]N), 138.76, 131.17,
5.1.14. Pd(C12H9ClN3S2)Cl2 (2b)
130.49, 129.01, 128.00, 127.51, 126.88, 126.25 (ArC).
Brown solid (ethanol). Yield: 62%. m.p.: 250 ꢀC (with decom-
position). Anal. calc. (C12H9N3S2Cl3Pd): C, 30.53; H, 1.92; N, 8.90; S,
13.58; found: C, 30.28; H, 1.81; N, 8.62; S, 13.85%. ES-MS (m/z) 438
[MeCl]þ. UV/Vis lmax(nm): 261, 372. IR nmax(KBr, cmꢁ1): 3205
(NeH); 1533 (C]N); 1105 (NeN); 748 (CeS); 511 (PdeN); 405
5.1.7. Preparation of Pt(II) and Pd(II) complexes (1ae5a): a general
method
A solution of K2PtCl4 or K2PdCl4 (0.38 mmol) in 10 mL distilled
water was added to appropriate thiosemicarbazone derivative
(0.38 mmol) in 10 mL ethanol. The reaction mixture was stirred for
24 h at room temperature. The solid precipitated was filtered and
washed with EtOH and diethyl ether and dried in vacou over silica
gel.
(PdeS). 1H NMR (DMSO-d6,
d ppm): 9.80 (s, 1H, PhNH); 8.55 (s, 1H,
HC]N); 7.68e7.18 (m, 7H, ArH).
5.1.15. Pd(C18H14N3S2)Cl2 (3b)
Brown solid (ethanol). Yield: 59%. m.p.: 307 ꢀC (with decom-
position). Anal. calc. (C18H14N3S2Cl2Pd): C, 42.08; H, 2.75; N, 8.18; S,
12.48; found: C, 42.19; H, 2.82; N, 8.33; S, 12.82%. ES-MS (m/z) 514
[Mþ]. UV/Vis lmax(nm): 270, 389. IR nmax(KBr, cmꢁ1): 3241 (N-H);
1527 (C]N); 1072 (NeN); 749 (CeS), 502 (PdeN), 412 (PdeS). 1H
5.1.8. Pt(C12H10N3S2)Cl2 (1a)
Orange solid (ethanol). Yield: 69%. m.p.: 212 ꢀC (with decom-
position). Anal. calc. (C12H10N3S2Cl2Pt): C, 27.38; H, 1.91; N, 7.98; S,
12.18; found: C, 27.43; H, 1.98; N, 7.82; S, 12.67%. ES-MS (m/z) 526
[Mþ]. UV/Vis lmax(nm): 265, 371. IR nmax(KBr, cmꢁ1): 3272 (N-H);
1510 (C]N); 1095 (N-N); 748 (C]S); 521 (PteN); 400 (PteS). 1H
NMR (DMSO-d6,
d ppm): 9.79 (s, 1H, PhNH); 8.64 (s, 1H, HC]N);
8.00e7.33 (m, 12H, ArH).
NMR (DMSO-d6,
8.00e6.92 (m, 8H, ArH).
d
ppm): 9.94 (s, 1H, PhNH); 8.48 (s, 1H, HC]N);
5.1.16. Pd(C12H9N4O2S2)Cl2 (4b)
Dark brown solid (ethanol). Yield: 54%. m.p.: 302 ꢀC (with
decomposition). Anal. calc. (C12H9N4O2S2Cl2Pd): C, 29.86; H, 1.88;
N, 11.61; S, 13.29; found: C, 29.43; H, 1.89; N, 11.66; S, 13.72%. ES-MS
(m/z) 483 [Mþ]. UV/Vis lmax(nm): 294, 452. IR nmax(KBr, cmꢁ1):
3255 (N-H); 1538 (C]N); 1320 (eNO2); 1099 (NeN); 778 (CeS),
5.1.9. Pt(C12H9ClN3S2)Cl2 (2a)
Dark yellow solid (ethanol). Yield: 53%. m.p.: 243 ꢀC (with
decomposition). Anal. calc. (C12H9N3S2Cl3Pt): C, 25.65; H, 1.79; N,
7.48; S,11.41; found: C, 25.43; H,1.58; N, 7.82; S,11.67%. ES-MS (m/z)
561 [Mþ]. UV/Vis lmax(nm): 264, 386. IR nmax(KBr, cmꢁ1): 3298 (N-
H); 1524 (C]N); 1088 (NeN); 747 (CeS); 513 (PteN); 402 (PteS).
510 (PdeN), 408 (PdeS). 1H NMR (DMSO-d6,
PhNH); 8.36 (s, 1H, HC]N); 8.31e7.20 (m, 7H, ArH).
d ppm): 10.27 (s, 1H,
1H NMR (DMSO-d6,
d
ppm): 9.85 (s, 1H, PhNH); 8.62 (s, 1H, HC]N);
5.1.17. Pd(C12H9N4O2S2)Cl2 (5b)
7.92e6.93 (m, 7H, ArH).
Brown solid (ethanol). Yield 61%. m.p.: 296 ꢀC (with decompo-
sition). Anal. calc. (C12H9N4O2S2Cl2Pd): C, 29.86; H, 1.88; N, 11.61; S,
13.29; found: C, 29.67; H, 1.75; N, 11.84; S, 13.34%. ES-MS (m/z) 483
[Mþ]. UV/Vis lmax(nm): 335, 412. IR nmax(KBr, cmꢁ1): 3262 (NeH);
1547 (C]N); 1329 (eNO2); 1181 (NeN); 769 (CeS), 509 (PdeN), 410
5.1.10. Pt(C18H14N3S2)Cl2 (3a)
Orange solid (ethanol). Yield: 56%. m.p.: 256 ꢀC (with decom-
position). Anal. calc. (C18H14N3S2Cl2Pt): C, 35.89; H, 2.34; N, 6.97; S,
10.64; found: C, 35.62; H, 2.02; N, 6.73; S, 10.48%. ES-MS (m/z) 602
[Mþ]. UV/Vis lmax(nm): 272, 403. IR nmax(KBr, cmꢁ1): 3288 (NeH);
1528 (C]N); 1049 (NeN); 791 (CeS); 507 (PteN); 418 (PteS). 1H
(PdeS). 1H NMR (DMSO-d6,
d ppm): 10.26 (s, 1H, PhNH); 8.91 (s, 1H,
HC]N); 8.38e7.22 (m, 7H, ArH).
NMR (DMSO-d6,
d
ppm): 9.81 (s, 1H, PhNH); 8.62 (s, 1H, HC]N);
5.2. Biological assays
7.78e6.98 (m, 12H, ArH).
The antiviral assays were based on the inhibition of virus-
induced cytopathicity in confluent cell cultures, and the cytostatic
assays on inhibition of tumor cell proliferation in exponentially
growing tumor cell cultures according to previously described
methodology [55e57].
5.1.11. Pt(C12H9N4O2S2)Cl2 (4a)
Dark brown solid (ethanol). Yield: 52%. m.p.: 220 ꢀC (with
decomposition). Anal. calc. (C12H9N4O2S2Cl2Pt): C, 25.23; H, 1.59; N,
9.81; S, 11.22; found: C, 25.43; H, 1.19; N, 9.66; S, 11.83%. ES-MS (m/
z) 571 [Mþ]. UV/Vis lmax(nm): 295, 467. IR nmax(KBr, cmꢁ1): 3320
(N-H); 1565 (C]N); 1046 (NeN); 807 (CeS); 518 (PteN); 411 (Pt-S).
5.2.1. Antiviral activity assays
The antiviral assays, other than the anti-HIV assays, were based
on inhibition of virus-induced cytopathicity in HEL [herpes simplex
1H NMR (DMSO-d6,
8.18e7.02 (m, 7H, ArH).
d
ppm): 10.12 (s,1H, PhNH); 8.63 (s,1H, HC]N);

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A. Karaküçük-Iyidogan et al. / European Journal of Medicinal Chemistry 46 (2011) 5616e5624
5623
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titer plate containing appropriate dilutions of the test compounds.
After 4 days of incubation at 37 ꢀC, HIV-induced CEM giant cell
formation was examined microscopically. Antiviral activity was
expressed as the concentration required to reduce virus-induced
cytopathogenicity by 50% (EC50).
5.2.2. Cytostatic/toxic activity assays
Cytostatic activity against CEM (human T-lymphocytes) and
HeLa (human cervix carcinoma) cell lines were measured essen-
tially as originally described for the mouse leukemia cell lines [8].
Human lymphocyte CEM and human cervix carcinoma HeLa cells
were seeded in 96-well microtiter plates at 75,000 (CEM) or 20,000
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The 50% cytostatic concentration (CC50) was defined as the comp-
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50%.
Cytotoxicity measurements were based on the inhibition of
MDCK, HEL, CRFK, Vero and HeLa cell growth. Cells were seeded at
5 ꢃ 103 cells/well into 96-well microtiter plates. Then, medium
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(MCC) or the compound concentration that causes a microscopi-
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This work was supported by Scientific Research Projects Gov-
erning Unit Council of Scientific Research Projects (Grant no. FEF.
09.09), Gaziantep, Turkey and the Concerted Action of the K.U.
Leuven (GOA no. 10/014). We thank Mrs. Leentje Persoons, Mrs.
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Uyttersprot for excellent technical asistance.
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