Benzylamidine complexes of platinum(II) derived by nucleophilic addition of primary and secondary amines. X-ray crystal structure of trans-[PtCl2{Z-N(H){double bond, long}C(NHMe)CH2Ph}2]

Article abstract of DOI:10.1016/j.ica.2007.10.054

The reaction of cis- and trans-[PtCl₂(NCCH₂Ph)₂] with a 5-fold excess of MeNH₂ and Me₂NH in CH₂Cl₂ at -10 °C affords in high yield the bis-amidine derivatives cis- and trans-[PtCl₂{Z-N(H){double bond, long}C(NHMe)CH₂Ph}₂] (1a, 2a) and cis- and trans-[PtCl₂{E-N(H){double bond, long}C(NMe₂)CH₂Ph}₂] (3a, 4a), respectively. The complexes were characterized by means of elemental analysis, multinuclear NMR and FT-IR techniques. The X-ray diffraction analysis was carried out for trans-[PtCl₂{Z-N(H){double bond, long}C(NHMe)CH₂Ph}₂] (2a). Moreover, the in vitro cytotoxicity for the new derivatives was evaluated in a wide panel of human tumor cell lines.

Full text of DOI:10.1016/j.ica.2007.10.054

Available online at www.sciencedirect.com
Inorganica Chimica Acta 361 (2008) 3109–3116
www.elsevier.com/locate/ica
Benzylamidine complexes of platinum(II) derived
by nucleophilic addition of primary and secondary amines. X-ray
crystal structure of trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2]
Silvia Mazzega Sbovata a, Frazia Bettio b, Christine Marzano b, Mirto Mozzon a,
Roberta Bertani a, Franco Benetollo c, Rino A. Michelin a,*
a Department of Chemical Processes of Engineering, University of Padova, Via F. Marzolo 9, 35131 Padova, Italy
b Department of Pharmaceutical Sciences, University of Padova, Via F. Marzolo 5, 35131 Padova, Italy
c ICIS – C.N.R., Corso Stati Uniti 4, 35127 Padova, Italy
Received 26 September 2007; accepted 25 October 2007
Available online 1 January 2008
Dedicated to Professor Robert J. Angelici for his many outstanding contributions to inorganic and organometallic chemistry.
Abstract
The reaction of cis- and trans-[PtCl2(NCCH2Ph)2] with a 5-fold excess of MeNH2 and Me2NH in CH2Cl2 at ꢀ10 °C affords in high
yield the bis-amidine derivatives cis- and trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2] (1a, 2a) and cis- and trans-[PtCl2{E-
N(H)@C(NMe2)CH2Ph}2] (3a, 4a), respectively. The complexes were characterized by means of elemental analysis, multinuclear
NMR and FT-IR techniques. The X-ray diffraction analysis was carried out for trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2] (2a).
Moreover, the in vitro cytotoxicity for the new derivatives was evaluated in a wide panel of human tumor cell lines.
Ó 2008 Elsevier B.V. All rights reserved.
Keywords: Amidine complexes; Platinum(II); Cytotoxic activity; Crystal structure
1. Introduction
and trans-[PtCl2{N(H)@C(OR0)R}2] [3,4] (with R = Me,
R0 = Me, Et) and amidine derivatives of the general for-
mula cis- and trans-[PtCl2{N(H)@C(NHR0)R}2] and cis-
and trans-½PtCl2fNðHÞ@CðNR0 ÞRg ꢁ [5–8] (with R = Me,
Organonitrile ligands can be activated toward addition
of a nucleophile [1] by ligating to a metal center which com-
monly results in an enhancement of the electrophilicity of
the unsaturated nitrile carbon atom [2]. Thus, organonitrile
Pt(II) complexes of the type cis- and trans-[PtCl2(NCR)2]
(R = alkyl, phenyl) are of special interest since they are
often excellent starting materials for the synthesis of new
families of platinum(II) derivatives. The addition reactions
of alcohols (R0OH), primary and secondary amines
(R0NH2 and R02NH) to the C„N triple bond of the nitrile,
affording the corresponding iminoethers of the type cis-
2
2
Ph and R0 = Me) have been investigated in the past years.
Such Pt(II) derivatives demonstrated to possess a valuable
pharmacological interest being a new generation of plati-
num antitumor drugs [9].
In particular, the results of the biological activity [10] of
the benzamidine derivatives showed that these species exert
a higher cytotoxicity than the corresponding acetamidines
and, among them, the complex trans-[PtCl2{N(H)@
C(NMe2)Ph}2] appeared the more effective derivative in
the biological assays. It yielded degrees of cytotoxicity
greater than those of cisplatin and circumvented acquired
cisplatin resistance. The results obtained by cellular uptake
indicated that it entered the cells more easily than the
*
Corresponding author. Tel.: +39 0498275522; fax: +39 0498275525.
E-mail address: rino.michelin@unipd.it (R.A. Michelin).
0020-1693/$ - see front matter Ó 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.ica.2007.10.054

3110
S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
reference drug. These results have been related to the more
lipophilic properties of the phenyl ring which enhances the
intracellular penetration of the drug.
platinate, K2PtCl4, was prepared as described in the litera-
ture [16]. Compounds cis- and trans-[PtCl2(NCCH2Ph)2] (1
and 2) were prepared as reported in the literature [11].
Complexes cis- and trans-[PtCl2{Z-N(H)@C(NHMe)CH3}2]
(1b, 2b), cis- and trans-[PtCl2{E-N(H)@C(NMe2)CH3}2]
(3b, 4b), cis- and trans-[PtCl2{Z-N(H)@C(NHMe)Ph}2]
(5b, 6b), cis- and trans-[PtCl2{N(H)@C(NMe2)Ph}2] (7b,
8b) have been synthesized as previously reported [5–8]. Cis-
platin was obtained from Sigma Chemical Co. The amidine
derivatives were dissolved in DMSO (dimethyl sulfoxide)
just before the experiments; calculated amounts of drug
solution were added to the growth medium containing cells
to a final solvent concentration of 0.5% which had no dis-
cernible effect on cell killing.
In our previous work we studied also the synthesis and
characterization of new benzyl iminoether derivatives
along with the evaluation of their anticancer properties.
Surprisingly, the more effective derivative proved to be
cis-[PtCl2{E-N(H)@C(OMe)CH2Ph}2], which was able to
overcome intrinsic cisplatin resistance and to inhibit the
synthesis of DNA in a dose-dependent way [11]. It repre-
sented the first example of an iminoether derivative, hav-
ing cis-geometry, endowed with a marked cytotoxic
potency. The compound was studied also for the in vivo
anticancer efficacy showing an activity higher than that
of cisplatin.
Along with this encouraging results, we explored the
synthesis of new amidine complexes of the type cis- and
trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2] and cis- and
trans-[PtCl2{E-N(H)@C(NMe2)CH2Ph}2]. The complexes
were spectroscopically and crystallographically character-
ized and their cytotoxicity was tested against a wide panel
of human tumor cell lines.
2.2. Synthesis of the complexes
2.2.1. Synthesis of cis-[PtCl2{Z-N(H)@C(NHMe)
CH2Ph}2] (1a)
All complexes were prepared by a similar procedure
which is described below for 1a.
A
suspension of cis-[PtCl2(NCCH2Ph)2] (175 mg,
0.35 mmol) in CH2Cl2 (30 mL) at ꢀ10 °C was treated with
a 5-fold excess of MeNH2 (0.87 mL of a 2 M solution in
THF, 1.75 mmol). The reaction mixture was stirred at
ꢀ10 °C for 5 h and then the volume was concentrated to
ca. 10 mL under reduced pressure. Et2O (40 mL) was
added and the suspension was stirred at ꢀ10 °C for 1 h.
The precipitate was then filtered off at room temperature
and washed with Et2O (3 ꢂ 15 mL). Recrystallization from
CH2Cl2/Et2O gave 1a as a pale yellow solid. Yield 175 mg,
88.5%. Anal. Calc. for C18H24N4Cl2Pt ꢃ 0.5 CH2Cl2: C,
36.74; H, 4.17; N, 9.26. Found: C, 36.80; H, 4.07; N,
9.21%. IR (KBr): m(N–H) 3260 cmꢀ1 (m), m(C@N)
1634 cmꢀ1 (s). IR (PE): m(Pt–Cl) 331, 302 cmꢀ1 (m, br).
2. Experimental
2.1. General procedures and materials
IR spectra were taken on a FT-IR NEXUS of the Nico-
let Instrument Corporation (KBr or polyethylene (PE)
films) spectrophotometer; the wavenumbers (m) are given
in cmꢀ1 1H and 13C {1H} NMR spectra were run at
.
298 K, unless otherwise stated, on a Bruker 200 AC spec-
trometer operating at 200.13 and 50.32 MHz, respectively;
d values are given in ppm. Peak positions are relative to
internal Me4Si. H and 13C NMR spectra were obtained
1
at 298 K as CD2Cl2 solutions on a Bruker DRX-400 spec-
trometer operating at 400.13 and 100.61 MHz, respectively,
and equipped with a BVT2000 temperature controller.
Suitable integral values for the proton spectra were
obtained by a pre-scan delay of 10 s to ensure a complete
relaxation for all the resonances. The proton assignments
were performed by standard chemical shift correlations as
well as by Nuclear Overhauser Enhancement SpectroscopY
(NOESY) experiments. The 13C resonances were attributed
through 2D-heteronuclear correlation experiments (Heter-
onuclear Multiple Quantum Correlation, HMQC, with
Bilinear Rotation-Decoupling, BIRD, sequence [12,13]
and quadrature along F1 achieved using the Time-Propor-
tional receiver Phase Incrementation, TPPI, method [14,15]
for the quaternary ones). The elemental analyses were per-
formed by the Microanalysis Laboratory of the Depart-
ment of Chemical Sciences, University of Padova. All
work was carried out under a N2 atmosphere using stan-
dard Schlenck techniques. All solvents were reagent grade
and were distilled prior to use. Deuterated solvents were
purchased from Cambridge Isotope Laboratories (CIL)
and stored under molecular sieves. Potassium tetrachloro-
3
1H NMR (CD2Cl2): d = 2.86 (d, 6H, JHH = 5.3 Hz, N–
CH3), 3.59 (s, 4H, NC–CH2), 4.92 (br, 2H, PtNH), 7.17
(m, 2H, Ph, o-H), 7.35 (m, 1H, Ph, p-H), 7.38 (m, 2H,
Ph, m-H), 7.45 (br, 2H, NH). 13C {1H} NMR (CD2Cl2):
d = 29.6
(s,
N(H)–CH3),
38.9
(s,
N@CCH2,
3JCPt = 39.7 Hz), 128.12 (s, Ph, p-C), 129.27 (s, Ph, o-C),
129.42 (s, Ph, m-C), 132.9 (s, Ph, Cj), 168.8 (s, C@N).
2.2.2. Synthesis of trans-[PtCl2{Z-N(H)@C(NHMe)
CH2Ph}2] (2a)
Yield 367 mg, 98.9%. Anal. Calc. for C18H24N4Cl2Pt: C,
38.44; H, 4.30; N, 9.96. Found: C, 38.13; H, 4.27; N, 9.77%.
IR (KBr): m(NH) 3277 cmꢀ1 (m), m(C@N) 1625 cmꢀ1 (s),
m(PtCl) 304 cmꢀ1 (m). 1H NMR (CD2Cl2): d = 2.84 (d,
3
6H, JHH = 5.2 Hz, N–CH3), 3.61 (s, 4H, NC–CH2), 5.16
(br, 2H, PtNH), 7.26 (m, 2H, Ph, o-H), 7.36 (m, 1H, Ph,
p-H), 7.41 (m, 2H, Ph, m-H), 7.47 (br, 2H, NH). 13C
{1H} NMR (CD2Cl2): d = 29.5 (s, N(H)–CH3), 39.4 (s,
3
N@CCH2, JCPt = 35.8 Hz), 128.06 (s, Ph, p-C), 129.39
(s, Ph, o-C), 129.63 (s, Ph, m-C), 133.01 (s, Ph, Cj), 168.9
(s, C@N).

S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
3111
2.2.3. Synthesis of cis-[PtCl2{E-N(H)@C(NMe2)
CH2Ph}2] (3a)
2.3. X-ray crystal structure determination of 2a
Yield 270 mg, 91.0%. Anal. Calc. for C20H28N4Cl2Pt: C,
40.68; H, 4.78; N, 9.49. Found: C, 40.59; H, 4.55; N, 9.34%.
IR (KBr): m(NH) 3272 cmꢀ1 (m), m (C@N) 1600 cmꢀ1 (s).
A prismatic yellow crystal of trans-[PtCl2{Z-N(H)@
C(NHMe)CH2Ph}2] of dimensions
0.22 ꢂ 0.24 ꢂ
0.36 mm, was sealed in a Lindemann capillary and used
for the diffractometric experiment. The measurements were
carried out by using a Philips PW1100 diffractometer, with
graphite-monochromated Mo-Ka radiation (k = 0.71073
1
IR (PE): m(PtCl) 328, 318 cmꢀ1 (m). H NMR at 255 K
(CD2Cl2): d = 2.64 (s, 6H, N–CH3), 2.91 (s, 6H, N–CH3),
4.41 (s, 4H, CH2), 6.20 (br, 2H, NH), 7.05–7.08 (m, Ph),
1
˚
7.25–7.26 (m, Ph). H NMR at 298 K (CD2Cl2): d = 2.79
A) following the standard procedures at room temperature.
(s, 12H, N–CH3), 4.49 (s, 4H, CH2), 5.95 (br, 2H, NH),
7.19 (m, 2H, Ph, o-H), 7.28 (m, 1H, Ph, p-H), 7.33 (m,
2H, Ph, m-H). 13C {1H} NMR (CD2Cl2) at 298 K:
d = 39.6 (br, CH3), 41.2 (s, CH2), 126.59 (s, Ph, p-C),
126.60 (s, Ph, o-C), 128.55 (s, Ph, m-C), 135.45 (s, Ph,
Cj), 168.5 (s, N@C).
All intensity data were corrected for Lorentz-polarization
effects and absorption [17]. The structure was solved by
direct methods using SIR-97 [18] and refined by full-matrix
least-squares using anisotropic temperature factors for all
non-hydrogen atoms. Hydrogen atoms were introduced
at calculated positions in their described geometries and
during refinement were allowed to ride on the attached car-
bon atoms with fixed isotropic thermal parameters
(1.2 Ueq of the parent carbon atom), except for the N(1)
and N(2) protons which were found in the final DF2 map
and refined isotropically. The calculations were performed
with the SHELXL-97 program [19], implemented in the
WINGX package [20]. Crystal data are listed in Table 1.
2.2.4. Synthesis of trans-[PtCl2{E-N(H)@C(NMe2)
CH2Ph}2] (4a)
Yield 360 mg, 87.0%. Anal. Calc. for C20H28N4Cl2Pt: C,
40.68; H, 4.78; N, 9.49. Found: C, 40.73; H, 4.46; N, 9.47%.
IR (KBr): m(NH) 3306 cmꢀ1 (m), m(C@N) 1628 cmꢀ1 (s). IR
1
(PE): m(PtCl) 307 cmꢀ1 (m). H NMR at 225 K (CD2Cl2):
d = 2.85 (s, 6H, N–CH3), 2.87 (s, 6H, N–CH3), 4.53 (s,
4H, CH2), 5.08 (br, 2H, NH), 7.20–7.31 (m, Ph), 7.41–
7.45 (m, Ph). H NMR at 298 K (CD2Cl2): d = 2.88 (s,
2.4. Cell cultures
1
12H, CH3), 4.55 (s, 4H, CH2), 5.17 (br, 2H, NH), 7.49 (m,
2H, Ph, o-H), 7.25 (m, 1H, Ph, p-H), 7.31 (m, 2H, Ph,
m-H). 13C {1H} NMR (CD2Cl2) at 298 K: d = 38.2 (br,
CH3), 41.3 (s, CH2), 126.46 (s, Ph, p-C), 128.35 (s, Ph,
o-C), 128.51 (s, Ph, m-C), 134.30 (s, Ph, Cj), 166.5 (s, N@C).
HeLa, LoVo, A549 are human cervix, colon and lung
carcinoma cell lines, respectively, which were obtained
from ATCC (RocKvill, MD) along with the hepatocellular
carcinoma (HepG2), malignant melanoma (A375) and
colon carcinoma (CaCo2) cell lines. HL60 human promye-
locytic leukemia cell line was kindly provided by Prof. F.
Majone, Department of Biology of Padova University,
Italy. Cell lines were maintained in the logarithmic phase
at 37 °C in a 5% carbon dioxide atmosphere using the fol-
lowing culture media: (i) RPMI-1640 medium (Euroclone,
Celbio, Milan, Italy) containing 10% foetal calf serum
(Biochrom-Seromed GmbH & Co., Berlin, Germany) and
supplemented with 25 mM HEPES buffer, l-glutamine
and with antibiotics penicillin (50 units mLꢀ1) and strepto-
mycin (50 lg mLꢀ1) for HL60; (ii) F-12 HAM’S (Sigma
Chemical Co., St. Luis, USA) containing 15% foetal calf
Table 1
Crystallographic data for compound 2a
Compound
trans-[PtCl2{Z-
N(H)@C(NHMe)CH2Ph}2]
Molecular formula
Molecular weight
Temperature (K)
k (A)
Crystal system
C8H18N2Cl2Pt
562.40
293(2)
0.71073
monoclinic
P21/n
10.498(2)
7.193(1)
13.454(3)
93.72(2)
1013.8(3)
2
˚
Space group
˚
a (A)
serum, penicillin (50 units mLꢀ1
)
and streptomycin
˚
b (A)
(50 lg mLꢀ1) for LoVo and HeLa cells; (iii) D-MEM (Dul-
becco’s modified eagle’s) medium (Euroclone, Celbio,
Milan, Italy) supplemented with 10% foetal calf serum
(Euroclone, Celbio, Milan, Italy), penicillin (50 uni-
ts mLꢀ1) and streptomycin (50 lg mLꢀ1) for A549 and
A375 cells; (iv) MEM (Minimum essential eagle’s) medium
(Euroclone) supplemented with 10% foetal calf serum
(Euroclone, Celbio, Milan, Italy), penicillin (50 uni-
ts mLꢀ1) and streptomycin (50 lg mLꢀ1) for HepG2 and
CaCo2 cells.
˚
c (A)
b (°)
3
˚
V (A )
Z
qcalc (g cmꢀ3
F(000)
)
1.842
544
h Range (°)
3–28
7.192
l (mmꢀ1
)
Number of reflections
collected
2524
Number of observed
[I P 2r(I)]
R (F2)a
2078
0.033
0.088
1.288
Rw (F2)b
2.5. Cytotoxicity assay
S
P
P
a
R ¼ ðjF oj ꢀ jF cjÞ= jF oj.
The growth inhibitory effect toward tumor cell
lines was evaluated by means of MTT (tetrazolium salt
P
P
2
2
2
2 2 1=2
b
Rw ¼ ½ wðjF oj ꢀ jF cj Þ = wðjF oj Þ ꢁ
.

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S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
reduction) assay [21]. Briefly, between 3 and 8 ꢂ 103 cells,
dependent upon the growth characteristics of the cell line,
were seeded in 96-well microplates in growth medium
(100 lL). After 24 h, cells were treated for 24 h with the
appropriate concentrations of the compound. Each well
was added with 10 lL of a 5 mg mLꢀ1 MTT (3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide)
saline solution, and after 5 h of incubation, 100 lL of a
sodium dodecylsulfate (SDS) solution in HCl 0.01 M were
added. After overnight incubation, the absorbance of each
well was detected by a microplate reader (Biorad 680,
Milan, Italy) at 570 nm. Mean absorbance for each drug
dose was expressed as a percentage of the control
untreated well absorbance and plotted vs drug concentra-
tion. IC50 values represent the drug concentrations that
reduced the mean absorbance at 570 nm to 50% of those
in the untreated control wells.
3. Results and discussion
3.1. Synthesis and characterization of the complexes
Scheme 1. Synthesis of the benzylamidine Pt(II) complexes with Z
configuration, derived by addition of the primary amine to the starting
nitrile complexes.
The addition reactions of protic nucleophiles to the
C„N triple bond of nitrile Pt(II) complexes, have been
extensively studied [1,2]. Over the past 10 years, in our
group, the reactions of primary and secondary ammines
with nitrile complexes of the type cis- and trans-
[PtCl2(NCR)2] (R = Me, Ph) have been investigated. We
found out that in the case of the acetonitrile ligands, in
cis- and trans-[PtCl2(NCMe)2] complexes, the reaction pro-
ceeds in a selective manner. Thus, primary amines, as
MeNH2, afford amidine complexes exclusively with the Z
configuration, corresponding to the trans addition of the
amine along the C„N triple bond, while secondary amines,
as Me2NH, yield amidine complexes only with E configura-
tion, corresponding to the cis addition of the amine [5–7]. In
contrast with the reactions of the acetonitrile derivatives,
the corresponding reactions of the benzonitrile complexes
cis- and trans-[PtCl2(NCPh)2] with primary and secondary
aliphatic amines afford a complex mixture of isomers Z,
Z0, E, E0 [8].
In this work we investigated the reactions of cis- and
trans-[PtCl2(NCCH2Ph)2] (1 and 2) with a 5-fold excess
of MeNH2 at ꢀ10 °C in CH2Cl2, which afford the corre-
sponding di-amidine derivatives in Z configuration cis-
and trans-[PtCl2{Z-N(H)=C(NHMe)CH2Ph}2] (1a and
2a) as reported in the following scheme (Scheme 1).
On the other hand, the reactions of complexes 1 and 2
with Me2NH, performed under the same experimental con-
ditions, lead to the isolation of the bis-amidine complexes
cis- and trans-[PtCl2{E-N(H)@C(NMe2)CH2Ph}2] (3a and
4a) respectively, bearing the two amidine ligands in the E
configuration (Scheme 2).
Scheme 2. Synthesis of the benzylamidine Pt(II) complexes with E
configuration, derived by addition of the secondary amine to the starting
nitrile complexes.
3260 cmꢀ1, while the C@N vibration appears in the range
1634–1600 cmꢀ1. The Pt–Cl absorptions are found in the
expected range of 331–302 cmꢀ1 [22].
The E or Z conformation of the amidine ligands in
complexes 1a–4a can be readily deduced by 1H NMR
Compounds 1a–4a were characterized by microanalysis,
IR, 1H and 13C NMR spectroscopy (see also Section 2) and
2a also by X-ray diffraction (see later). The IR spectra
(KBr) show the N–H stretching band in the range 3306–

S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
3113
data upon observing the position of the chemical shifts
of the methylene group, since a significant downfield
shift is experienced by protons which come close to plat-
inum [23,24]. In fact, the resonance of the methylenic
group displays in the Z form a singlet resonance in the
range d 3.59–3.61 (1a–2a), while in the case of the E iso-
mers, 3a–4a, these values experience a download shift to
d 4.49–4.55. Moreover, the conformation of the amidine
ligands appears to be confirmed by NOESY experiments.
Spectra recorded for complexes 1a and 2a confirm the Z
configuration of the ligands, allowing to detect an intense
correlation peak between the CH2 group and the Pt–NH
proton, indicating that they are on the same side of the
double C@N bond. Another correlation is observed
between the N–CH3 group and the ortho phenyl protons.
Also for complexes 3a and 4a the NOESY spectra assign
the E configuration since a correlation peak is observed
between the N–CH3 group and the Pt–NH proton
confirming that their arrangement is on the same side
along the double bond. Again correlations between the
N–CH3 group and the ortho phenyl protons are
observed.
The Z configuration of the amidine ligands appears to
be favoured by the formation of strong intramolecular
hydrogen bonds between each chlorine atom and the
amino proton of the NHMe moiety forming a six-member
ring. While the E isomers are the expected kinetically
favoured products [25,26] upon the addition of amine to
coordinated nitriles, which indicate that the reaction pro-
ceeds via a four or a six-center transition state (the latter
involving two amine molecules) formed by the cis-addition
of the amine to the cyano group.
Fig. 1. ORTEP view of the molecule trans-[PtCl2{Z-N(H)@C(NHMe)
CH2Ph}2] (2a).
The square planar coordination geometry around Pt is
characterized by the values of the Pt–N(1) and Pt–Cl
˚
bond distances of 2.024(5) and 2.303(2) A, respectively,
in agreement with the literature data [5–7] for similar
Pt(II) derivatives with coordinated amidine ligands. The
ligands are shown to be in the Z configuration. The
molecular structure is characterized by two intramolecular
0
˚
contacts of 2.55(9) A involving the chlorine atom (Cl )
and the H(2)–N(2) proton, with a N(2)–H(2)ꢃ ꢃ ꢃCl0 angle
of 147°, demonstrating the presence of intramolecular
hydrogen bonds stabilizing the Z configuration of the
ligands as already observed in other amidine Pt(II) com-
plexes [5].
The dimethylamino group of complexes 3a and 4a
appears as a singlet at room temperature at d 2.79 and
2.88, respectively, indicating the presence of free rotation
along the N–Me2 bond. Lowering the temperature at
255 K, it has been observed that the signal is resolved into
two singlets at d 2.64 and 2.91 for compound 3a, with a free
The intramolecular interaction together with the steric
hindrance of the phenyl group may determine the wider
opening observed for the Pt–N(1)–C(1) angle (136.1(5)°),
as already observed in the complex trans-[PtCl2{Z-
N(H)@C(NHBut)Me}2] [8] having bulky t-butyl groups
(138.7(3)°), with respect to that found for the derivatives
trans-[PtCl2{Z-N(H)@C(NHMe)Me}2] [5] of 131.9(7)°.
¼
energy of activation of DG278:1 of 14.53 kcal/mol, value in
agreement with those reported for the rotational barriers
of this kind of compounds [27]. Similarly, the resonance
of the dimethylamino group of compound 4a is split into
two resonances at d 2.87 and 2.85 at 225 K with a free
˚
The N(1)–C(1) bond distance of 1.278(8) A is of the
same order of magnitude of the C(1)–N(2) bond length
˚
of 1.324(9) A, suggesting an extensive electron delocaliza-
¼
energy of activation of DG246 of 13.42 kcal/mol.
tion along the N(1)–C(1)–N(2) moiety with an sp2 hybrid-
ization of both the nitrogen atoms. Similar values for the
same type of distances have been found in the derivative
trans-[PtCl2{Z-N(H)@C(NHMe)Me}2] [5], which shows
In the 13C NMR spectra, the resonance of the C@N car-
bon falls in the range d 166.5–168.9 as expected for amidine
derivatives. The signal for the methyl groups are found in
the range d 29.5–39.6 while the singlets corresponding to
the nitrile CH2 groups are observed in the expected region
(d 38.9–41.3).
˚
equal N(1)–C(1) and C(1)–N(2) values of 1.32(1) A, and
in the crystal structure of trans-[PtCl2{Z-N(H)@C(NHBut)
Me}2] [8] which shows the N(1)–C(1) bond distance of
˚
˚
1.304(4) A and the N(2)–C(1) bond length of 1.321(5) A.
The results reported here seem to suggest that the electron
delocalization is peculiar of this type of ligands, as shown
by the reported examples, where the N–C(R)–N system is
about planar in analogy with the same evidence reported
for the imino ether moiety in the free ligand [28]. The
3.2. X-ray diffraction study of trans-[PtCl2
{Z-N(H)@C(NHMe)CH2Ph}2] (2a)
The ORTEP drawing of 2a is shown in Fig. 1 and the
crystal cell is shown in Fig. 2, while selected bond distances
and angles reported in Table 2.
˚
˚
Pt(1)–Cl(1) (2.303 A) and Pt(1)–N(1) (2.024 A) bond

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S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
Fig. 2. Packing diagram for trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2] (2a).
3.3. Cytotoxic activity
Table 2
˚
Selected bond lengths (A) and angles (°) for compound 2a
The cytotoxic properties of benzylamidine derivatives
cis- and trans-[PtCl2{Z-N(H)@C(NHMe)CH2Ph}2] (1a,
2a), cis- and trans-[PtCl2{E-N(H)@C(NMe2)CH2Ph}2]
(3a, 4a) were evaluated in comparison with cisplatin
against a panel of seven human tumor cell lines containing
examples of cervix (HeLa), colon (LoVo, CaCo2), lung
(A549) and hepatocellular (HepG2) cancer, leukemia
(HL60) and melanoma (A375). In Table 3 are shown the
IC50 values, calculated from the dose-survival curves
obtained after 24 h drug treatment from MTT test, for
the complexes 1a–4a, together with the IC50 values of the
already synthesized complexes cis- and trans-[PtCl2{Z-
Pt–N(1)
Pt–Cl
2.024(5)
2.303(2)
1.278(8)
1.324(9)
1.432(9)
1.513(8)
1.502(9)
N(1)–Pt–Cl0
N(1)–Pt–Cl
96.2(2)
83.8(2)
N(1)–C(1)
N(2)–C(1)
N(2)–C(2)
C(1)–C(3)
C(3)–C(4)
C(1)–N(1)–Pt
C(1)–N(2)–C(2)
N(1)–C(1)–N(2)
N(1)–C(1)–C(3)
N(2)–C(1)–C(3)
C(4)–C(3)–C(1)
136.1(5)
126.4(7)
122.3(6)
120.6(6)
117.1(6)
115.2(5)
distances as well as the Cl(1)–Pt(1)–Cl(1)0 (180°), N(1)–
Pt(1)–N(1)0 (180°) and N(1)–Pt(1)–Cl (83.8(2)–96.2(2)°)
angles have normal values [29,30].
Table 3
Evaluation of in vitro cytotoxic activity of the investigated platinum(II) benzylamidine complexes (1a–4a) toward some established human tumor cell lines
by MTT test after 24 ha
Compound
IC50 [lM] S.D.
HeLa
HL60
LoVo
CaCo2
HepG2
A549
A375
1a
2a
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
3a
4a
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
5b
6b
89.40 2.55
18.53 0.55
77.23 1.55
7.54 0.31
16.64 0.44
96.10 3.99
6.10 0.99
66.10 1.99
4.75 0.44
19.55 0.26
80.87 3.23
31.99 1.77
74.25 1.32
13.26 2.60
55.55 2.66
93.21 2.78
25.33 0.98
60.47 2.55
8.26 1.77
75.01 2.08
83.41 2.08
18.99 1.44
68.98 2.13
10.77 3.16
34.00 3.45
79.09 1.33
32.9 1.66
72.09 2.77
13.07 1.16
38.37 1.79
75.98 0.97
62.88 2.04
64.86 0.44
24.05 1.42
50.05 2.03
7b
8b
Cisplatin
For comparison purposes, the cytotoxicities of the already known benzamidine complexes 5b–8b are reported.
a
IC50 values were calculated by probit analysis (P < 0.05, v2 test). Cells (3–8 ꢂ 104 mLꢀ1) were treated for 24 h with increasing concentrations of the
tested compounds. S.D. = standard deviation.

S.M. Sbovata et al. / Inorganica Chimica Acta 361 (2008) 3109–3116
3115
N(H)@C(NHMe)Ph}2] (5b, 6b) and cis- and trans-
Acknowledgements
[PtCl2{N(H)@C(NMe2)Ph}2] (7b, 8b) reported in a previ-
ous work [10].
This work was financially supported by University of
Padua (Progetto di Ateneo CPDA065113/06) and by
It can be observed that the benzylamidine complexes
1a–4a proved to be ineffective in all tumor cell lines, while
the benzamidine derivatives 5b–8b demonstrated to possess
a significant in vitro antitumor activity. In particular, the
trans-complexes 6b and 8b showed a cytotoxic activity
comparable and even better than cisplatin. The corre-
sponding benzamidine cis-isomers 5b and 7b, instead,
showed an activity significantly lower than that of cisplatin
and of the corresponding trans-isomers.
`
Ministero dell’Istruzione dell’Universita e della Ricerca.
We are grateful to CIRCMSB (Consorzio Interuniversitar-
io di Ricerca in Chimica dei Metalli nei Sistemi Biologici).
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4. Supplementary material
CCDC 665125 contains the supplementary crystallo-
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