Journal of Inorganic Biochemistry 83 (2001) 169179

www.elsevier.nl / locate / jinorgbio

Synthesis, characterisation, X-ray structure and biological activity of three

new 5-formyluracil thiosemicarbazone complexes
a,
a
a
a
Marisa Belicchi Ferrari *, Franco Bisceglie , Giorgio Pelosi , Pieralberto Tarasconi ,
Roberto Albertini b,c , Antonio Bonati b,c , Paolo Lunghi b,c , Silvana Pinelli b,c
a

Dipartimento di Chimica Generale ed Inorganica, Chimica Analitica, Chimica Fisica, Centro di Studio per la Strutturistica Diffrattometrica del
CNR, University of Parma, Parco Area delle Scienze 17 A, 43100 Parma, Italy
b
Istituto di Patologia Speciale Medica, University of Parma, 43100 Parma, Italy
c
Centro di Ricerca Interuniversitario per Diagnosi, Terapia e Prognosi di Tumori Umani, University of Milan, 20122 Milan, Italy
Received 27 April 2000; received in revised form 10 August 2000; accepted 16 August 2000

Abstract
Three new complexes of transition metals as copper, nickel and cobalt with 5-formyluracil thiosemicarbazone (H 3 ut) have been
synthesised and characterised by single-crystal X-ray diffraction. In all compounds the ligand behaves as SNO terdentate. In the copper
complex the coordination geometry is square pyramidal with the ligand lying on the basal plane and two water molecules that complete
the metal environment, the nickel compound is surrounded by six donor atoms (three of the ligand, two water oxygen atoms and a
chlorine atom) in an octahedral fashion, and cobalt also shows an octahedral geometry but determined only by two terdentate ligand
molecules. These three compounds have been tested on human leukemic cell lines K562 and CEM. The nickel and cobalt complexes have
demonstrated low activity in cell growth, while the copper complex that is more active has been tested also on a third leukemic human
cell line (U937), but it was not able to induce apoptosis on all cell lines. 2001 Elsevier Science B.V. All rights reserved.
Keywords: 5-Formyluracil thiosemicarbazone; Cu, Ni, Co Complexes; Apoptosis; X-ray structure

1. Introduction
Following our interest in the biological and chemical
properties of polydentate ligands belonging to the
thiosemicarbazone family and of their complexes with
potential antitumor properties [15], we have recently
studied the reaction of a novel ligand, 5-formyluracil
thiosemicarbazone (H 3 ut) with salts of transition metals
and reported in a recent paper the synthesis, characterisation and biological activity of three copper complexes with
this ligand [Cu(H 2 ut)(OH 2 )Cl]?2H 2 O, [Cu(H 3 ut)Cl 2 ]?
2H 2 O and [Cu(H 3 ut)(OH 2 )SO 4 ]?3H 2 O [4]. In the present
work we have extended the study to another copper
complex containing the nitrate group as counterion
[Cu(H 3 ut)(OH 2 ) 2 ](NO 3 ) 2. H 2 O (1) and observed the coordinative behaviour of the same ligand towards different
transition metals such as nickel and cobalt. Two complexes
*Corresponding author. Tel.: 139-521-905-432; fax: 139-521-905557.
E-mail address: chimic8@ipruniv.cce.unipr.it (M. Belicchi Ferrari).

have also been characterised by single-crystal X-ray

diffraction:
[Ni(H 3 ut)(OH 2 ) 2 Cl]Cl?H 2 O
(2)
and
[Co(H 2 ut) 2 ]0.5SO 4 ?2.5H 2 O (3).
In our previous studies we have observed that metal
complexes with thiosemicarbazones can interfere in the
complicated mechanisms that lead to a leukemic transformation [36]. The uncontrolled proliferation of the
leukemic clone, the arrest of maturation at the initial stage
of hemopoiesis and the slowing down of cell turnover are
the most important alterations that characterise the
pathological manifestation of leukemia. Molecules that
interfere with these mechanisms can contribute to the
elimination of neoplastic cells by inhibiting replication, by
inducing differentiation or by triggering apoptotic processes and, in this way, act as potentially very important
therapeutic substances [710].
Therefore for all the complexes under study, assays of
proliferation inhibition on human leukemic cell lines K562
and CEM were carried out in vitro. The copper complex
has demonstrated a higher activity in relation to the
inhibition of cell proliferation with respect to the cobalt

0162-0134 / 01 / $ see front matter 2001 Elsevier Science B.V. All rights reserved.
PII: S0162-0134( 00 )00181-1

170

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

and nickel complexes, therefore complex 1 has been tested

on a third human leukemic cell (U937). In the subsequent
apoptosis tests complex 1 was not able to induce selfregulated cell death.

2. Experimental

2.1. Chemical data

All reactants and solvents were reagent grade.
Thiosemicarbazide was purchased from Merck and 5formyluracil from Aldrich. Elemental analyses (C, H, N,
S) were performed on a Carlo-Erba Instruments CHNS-O
EA 1108 automatic equipment.

2.2. Preparations
2.2.1. H3 ut? H2 O
The 5-formyluracil thiosemicarbazone was prepared
using the procedure reported in Ref. [4].
2.2.2. [ Cu( H3 ut)( OH2 )2 ]( NO3 )2? H2 O (1)
The ligand (49 mg) was refluxed under N 2 for ca. 2 h in
H 2 O (40 ml) until dissolution was complete. To this
yellow solution under stirring Cu(NO 3 ) 2 ?3H 2 O (55 mg)
(ratio H 3 ut:metal51:1) dissolved in H 2 O (25 ml) at room
temperature was added slowly dropwise. To the green
mixture, diluted with water (10 ml), an HNO 3 (2N) and
KNO 3 saturated solution in H 2 O (25 ml) was added slowly
dropwise and refluxed for ca. 2 h under N 2 . During the
evaporation, the green-coloured solution became more
clear. Crystals were obtained by slow evaporation of the
solvent. Anal. found: C, 15.12; H, 3.15; N, 20.76; S,
7.55%. Calc. for C 6 H 13 CuN 7 O 11 S: C, 15.85; H, 2.88; N,
21.56; S, 7.05%.
2.2.3. [ Ni( H3 ut)( OH2 )2 Cl] Cl? H2 O (2)
A solution of NiCl 2 ?6H 2 O (55 mg; 0.234 mmol) in
water (40 ml) at room temperature was added dropwise to
a hot water solution of the ligand H 3 ut, (molar ratio51:1).
From the limpid reaction mixture (pH ca. 3), after slow
evaporation of the solvent, clear green crystals were
isolated. Anal. found: C, 18.56; H, 3.05; N, 17.46; S,
8.25%. Calc. for C 6 H 12 NiN 5 O 4 S: C, 18.98; H, 3.15; N,
18.43; S, 8.44%.
2.2.4. [ Co( H2 ut)2 ]0.5 SO4?2.5 H2 O (3)
A solution of CoCl 2 ?6H 2 O (22 mg; 0.0925 mmol) in
water (20 ml) was added to a hot water solution of the
ligand H 3 ut, (molar ratio51:1). The resultant solution was
muddy and pH ca. 6. After slow evaporation prismatic
brown crystals were isolated. The analytical data were

consistent with the formula [Co(H 2 ut) 2 ]0.5SO 4 ?2.5H 2 O

established by X-ray analysis. The formation of a Co(III)
complex with the SO 22
group as counterion, although the
4
reactant salt was CoCl 2 , could be due to a prolonged
contact with the atmosphere that induced the oxidation of
the metal and the ligand. Anal. found: C, 24.32; H, 2.45;
N, 23.55; S, 14.36%. Calc. for C 12 H 17 CoN 10 O 8.5 S 2.5 : C,
25.00; H, 2.97; N, 24.30; S, 13.90%.
The same compound was also isolated by reacting
CoSO 4 ?7H 2 O with the ligand in water solution.

2.3. Data collection and processing

Relevant data concerning data collection are summarised
in Table 1. All intensity data were collected by the u 22u
technique on a Siemens AED single-crystal computer
controlled diffractometer with Cu Ka radiation for 1 and 2
and with Mo Ka radiation for 3. The absorption correction
following Sheldricks method was applied for both complexes 1 and 2, while no correction was made for
compound 3 because of the small crystal dimensions. The
intensities were measured using a modified version [11] of
the profile analysis method of Lehmann and Larsen [12]
and were corrected for Lorentz polarisation effects.

2.4. Structure analysis and refinement

The structure for all three compounds was solved by
direct methods using SHELX86 [13] and SIR92 [14].
Successive Fourier syntheses allowed the assignment of
the atoms to the electron density peaks. Refinements were
carried out by least-squares cycles (SHELX97 [15] for
compound 1 and SHELX93 [16] for 2 and 3). Anisotropic
thermal parameters were used for all non hydrogen atoms
in compounds 1 and 2, while in complex 3 the atoms of the
sulphate ion, that are disordered on two positions, a water
molecule O3W, in special position on a 2-fold axis, a
second water molecule O2W and the uracil oxygen atom
O2A, that are distributed on two positions, (O2W and
O2W9; O2A and O2A9, respectively) were refined isotropically.
In compound 1 the hydrogen atoms, located on a
difference map, were isotropically refined, except those
belonging to water molecules.
For both complexes 2 and 3 all hydrogen atoms were
not refined. In particular in compound 3 some hydrogen
atoms were placed in calculated positions (H4A, H4N,
H4AN, H5N, H5AN). Atomic scattering factors were
taken from Ref. [17]. All calculations were performed on
an ENCORE 91 computer at the Centro di Studio per la
Strutturistica Diffrattometrica del C.N.R. (Parma). Molecular geometry calculations were carried out by using
computer program PARST [18] and the structure drawings
were made using ORTEP [19] and PLUTO [20].

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

171

Table 1
Experimental data for crystallographic analyses
Compound

Formula
Molecular weight
Space group

a (A)

b (A)

c (A)
a (8)
b (8)
g (8)
3)
V (A
Z
F (000)
Dcalc (Mg / m 3 )
m (cm 21 )

l (A)
Radiation
u range (8)
hkl ranges
Crystal size (mm)
No. meas. reflections
No. unique reflections
No. refined parameters
Max and min height in
23 )
final DF map (eA
R5SiFo u2uFc i / SuFo u
wR 2
Weights a
a
b

(1)

(2)

(3)

C 6 H 13 CuN 7 O 11 S
454.8
P1
10.866(2)
9.420(3)
8.337(1)
83.91(1)
83.80(1)
110.46(2)
785.3(3)
2
462
1.923
39.58
1.54178
Cu Ka
370
213 to 13; 211 to 11;
0 to 10
0.6130.5130.23
2892
2755
300
0.88 20.61

C 6 H 13 Cl 2 N 5 NiO 5 S
396.9
P2 1 / c
16.345(5)
12.967(4)
6.756(2)
90.0
92.40(2)
90.0
1430.6(8)
4
808
1.843
70.30
1.54178
Cu Ka
370
219 to 19; 0 to 15;
0 to 8
0.9430.2330.05
2695
1867
245
1.13 20.66

C 12 H 17 CoN 10 O 8.5 S 2.5

576.4
C2 / c
7.372(2)
26.709(5)
21.958(4)
90.0
90.49(2)
90.0
4323(1)
8
2352
1.771
11.03
0.71069
Mo Ka
330
28 to 8; 0 to 31;
0 to 26
0.1630.2030.23
3821
1536
342
0.83 20.88

0.052 b
0.190
1 / [s 2 (F 2o )1(0.1033P)2 1
0.31P]

0.068 b
0.224
1 / [s 2 (F 2o )1(0.1971P)2
10.0P]

0.072
0.358
1 / [s 2 (F 2o )1(0.1234P)2
149.44P]

P5(max(F 2o ,0)12F 2c ) / 3.
The data were corrected for absorption following Sheldricks method [15].

3. Biological data

3.1. Materials and methods

Complexes 1, 2 and 3 were stored dry at room temperature and dissolved in dimethyl sulfoxide (DMSO) just
before their use.

3.2. Cellular growth

All cells K562, CEM clone-13 and U937 were grown in
RPMI 1640 supplemented with 10% foetal calf serum
(FCS) and antibiotics (penicillin 100U / ml and streptomycin 100 mg / ml) and added with L-glutamine 2 mM.
The cells were seeded at 3310 5 ml 21 concentration and
treated with the above mentioned compounds added to the
culture medium prior to cell seeding, using DMSO as a
control at the percentage of 1.2%.On the fourth day cell
mortality was evaluated by the trypan blue exclusion
method and determined by using a haemocytometer.

3.3. Apoptosis assay

The cell lines were seeded at 8310 5 ml 21 in the

presence of compound 1 and employed in apoptosis assay

using the agarose gel 2% electrophoresis. To these cells
(2310 6 ) (washed with PBS at 2000 rpm for 10 min at
48C) 20 ml of a solution of EDTA 10 mM, TrisHCl 50
mM, pH 8.0, and 0.5% (w / v) sodium laurylsarkosinate
was added. The pellet was subsequently redissolved and
added of 2.5 ml of Proteinase K [4 mg / ml] to a final
concentration of 0.5 mg / ml. After 1 h at 508C 2.5 ml of
Rnase A [2 mg / ml] were added and then incubated for 1 h
at 508C. The whole sample was subsequently loaded into
the wells and a tension of 80 V was applied to evaluate the
typical apoptotic effects of DNA fragmentation.

4. Results

4.1. Effects on cell proliferation and apoptosis induction

Nickel and cobalt compounds, 2 and 3, respectively,
have shown no cell growth inhibition activity at concentrations up to 40 mg / ml both on K562 and CEM lines.
On the contrary compound 1, already at a concentration of
30 mg / ml significantly inhibits proliferation in cell lines
CEM and also on the more resistant K562 at the con-

172

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

Fig.
1. Comparison
between
the
effects
of
complexes
[Cu(H 3 ut)(OH 2 ) 2 ](NO 3 ) 2 ?H 2 O (1), [Ni(H 3 ut)(OH 2 ) 2 Cl]Cl?H 2 O (2)
[Co(H 2 ut) 2 ]0.5SO 4 ?2.5H 2 O (3) on CEM proliferation (above) and on
K562 proliferation (below) on the fourth day.

centration of 35 mg / ml. These data are reported in Fig. 1

as average results of three experiments performed in
duplicate. Because of its higher activity in relation to the
inhibition of cell proliferation with respect to the cobalt
and nickel complexes, complex 1 has been tested on a third
human leukemic cell line U937 showing an inhibition of
cell proliferation greater than 50% at the value of 30
mg / ml (Fig. 2).
This compound, given its inhibition action on cell
growth, has therefore been tested to check its capacity of
inducing apoptosis. It has been observed that the test
carried out gave negative results, also on cell line U937
that corresponds to one of the latest stages of maturation
and is therefore more susceptible to undergo apoptosis.

5. Discussion

5.1. X-ray studies

An ORTEP plot of compounds 1, 2 and 3 is shown in
Figs. 35, respectively. Final atomic parameters are given
in Tables 24. A comparison between the main interatomic distances and angles for all complexes is reported
in Table 5.

Fig. 2. Effects of copper complex 1 at different concentrations on three

cell lines CEM, K562 and U937 on the fourth day.

In compound 1 the copper atom is surrounded by five

donor atoms in a square pyramidal fashion (411). The
basal plane is made up from S, N and O atoms of the
ligand in its neutral form and one water molecule (O2W);
the apical position is occupied by a second water molecule
with the N3 atom in
(O1W). A very long contact (3.52 A)
position 22x, 2y, 12z can be considered as a completion
of the copper elongated bipyramidal coordination. Two
ionic nitrate groups and another water molecule complete
the environment of the complex. The metal atom lies 0.19
over the average basal plane towards the O1W atom.
A
The angle between the CuO1W bond and the normal to
the plane is 7.38. The five-member chelation ring presents
a f2 of 2238 corresponding to a twist conformation and
the six-member one is characterised by a f2 of 38 and a u
of 678, i.e., an intermediate conformation between envelope and boat [21]. Couples of hydrogen bonds between
O2W and O1W of a centrosymmetrical molecule (O2W
create a dimeric
H???O1W(22x, 2y, 22z)52.694(5) A)

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

173

Fig. 3. Perspective view of centrosymmetric [Cu(H 3 ut)(OH 2 ) 2 ] 21 cations connected by hydrogen bonds in a dimer-like pattern.

situation, and dimer-like groups are linked by nitrate group

N6O3O4O5 by means of the oxygen atom O5 (O2WH???
N1H???O5(22x,12y,12z)52.911(6)
O552.664(4) A,
to form chains of dimer-like complexes running along
A)
the z direction at x50. The second nitrate ion N7O6O7O8
and the non-coordinated water molecule, bridged between
the two nitrates, connect the chains through hydrogen

bondings such as N1(amine)H???O752.806(7) A,

and N4(uracil)H???
N2(hydrazine)H???O652.859(6) A
Differently from compound
O8(12x,2y,2z)52.729(4) A.
1 the two copper complexes previously reported [4]
contain the chlorine atom coordinated to the central atom
both in a square pyramidal fashion (411). In
[Cu(H 2 ut)(OH 2 )Cl]?2H 2 O the chlorine occupies the apical

Fig. 4. ORTEP drawing of [Ni(H 3 ut)(OH 2 ) 2 Cl] 1 cation with thermal ellipsoids at 50% probability level.

174

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

Fig. 5. ORTEP drawing of [Co(H 2 ut) 2 ] 1 cation with thermal ellipsoids at 50% probability level.

position; in [Cu(H 3 ut)Cl 2 ]?2H 2 O the second chlorine atom

lies in the basal plane of the pyramid and the organic
ligand is neutral. Some relevant structural data of these
complexes are reported together with those of complex 1
in Table 6. It can be observed that the ligand distances in
complex 1 result more similar to those present in
[Cu(H 3 ut)Cl 2 ]?2H 2 O as a consequence of the fact that the
ligand is in its neutral form.
In compound 2 the nickel atom has an octahedraldistorted environment determined by the SNO terdentate
ligand, in its neutral form, by two water molecules and by
one chlorine atom. The complex is therefore positively
charged (Fig. 4) and a second chlorine atom is the
counterion. The five-member chelation ring presents a f2
of 1388 corresponding to an envelope conformation and the
six-member one is characterised by a f2 of 2268 and a u
of 938, i.e. shows a twist boat conformation [21]. The
and NiN (2.023(5) A)
bonds are
NiS of 2.347(2) A
comparable with those found in other Ni(II) octahedral
is
species [5,22,23]. The NiO (uracil) bond (2.029(4) A)
of the same magnitude order of the sum of the Pauling
The NiCl distance conforms to
covalent radii, i.e. 2.05 A.
the values given in the literature, such as NiO2W [24,25].
Packing is characterised by molecule layers extending

along the b axis and linked by crystallization water

molecules, O3W, near the plane yz (O3W???O2W (22x,2
N1???O3W (x,2y,z21 / 2)52.878(7)
y,22z)52.748(7) A,
and by CH???O2(12x,y21 / 2,5 / 22z) interactions
A
A network of hydrogen bonds
(3.335(7) and 3.184(7) A).
involving also the coordinated water molecules, the coordinated chloride atom and the other chloride ion which is out
of the metal coordination sphere contribute to the crystal
packing stability.
In compound 1 as well as in 2, the neutral ligand
presents a little charge delocalization as shown by the
SC1 and C2N3 distances, typical values of double bond
1.702(6) and 1.293(7) A,

(1.702(4) and 1.297(4) A;

respectively) (Table 5).
In complex 3, the central cobalt atom has an octahedral
environment given by two monodeprotonated ligand molecules (Fig. 5). Structural analysis revealed also the presence of SO 22
anions. There are also three non-coordinated
4
water molecules: one in general position, the second in
special position on a binary symmetry axis with half s.o.f.,
and the third disordered on two positions. Two ligands
have a mer configuration with S and O atoms in cis
position and N atoms in trans position as already found in
other Co(III) octahedral complexes [26,27]. The sulphate

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

175

Table 2
Fractional atomic coordinates (310 4 ) and equivalent isotropic displace4
2 ) for non-hydrogens atoms with e.s.d.s in
ment parameters (310 A
parentheses for (1)a

Table 4
Atomic coordinates (310 4 ) and equivalent isotropic displacement param4
2 ) for non hydrogens atoms with e.s.d.s in parentheses for
eters (310 A
(3)a

Atom

X /a

Y /b

Z /c

Ueq

Atom

X /a

Y /b

Z /c

Ueq

Cu
S1
O1
O2
O3
O4
O5
O6
O7
O8
O1W
O2W
O3W
N1
N2
N3
N4
N5
N6
N7
C1
C2
C3
C4
C5
C6

9435.5(5)
9455.8(9)
9606(3)
8597(3)
13 651(7)
14 250(6)
12 200(4)
5355(5)
5693(6)
3951(3)
8116(3)
11 119(3)
4213(4)
7724(4)
7574(3)
7994(3)
7299(3)
9096(3)
13 379(4)
4989(4)
8163(4)
7378(3)
7718(3)
7002(4)
8341(3)
8840(3)

354.8(5)
2573.9(9)
21573(3)
26597(3)
4690(9)
5591(11)
4361(4)
21385(5)
879(5)
21014(3)
124(3)
1311(3)
2111(5)
2577(4)
204(3)
2714(3)
25921(3)
24086(3)
4864(4)
2526(4)
1743(4)
22183(4)
23304(4)
24829(4)
25617(4)
22899(4)

7170.8(5)
5816.6(10)
8023(3)
9654(3)
8913(6)
6447(9)
7296(6)
3455(5)
2235(7)
2014(4)
9644(3)
7905(4)
326(6)
3850(5)
4965(4)
5982(3)
7988(4)
8720(3)
7559(4)
2596(4)
4819(4)
6099(4)
7073(4)
7133(4)
8849(4)
7937(3)

229(15)
263(15)
273(17)
306(17)
1088(39)
1392(42)
686(22)
722(23)
988(27)
437(18)
325(17)
431(18)
675(22)
417(20)
267(17)
226(17)
264(18)
248(17)
375(19)
439(20)
256(18)
236(18)
224(18)
256(18)
243(18)
212(18)

Co
S1
S1A
O2
O2A
O2A9
O1
O1A
N1
N1A
N2
N2A
N3
N3A
N4
N4A
N5
N5A
C1
C1A
C2
C2A
C3
C3A
C4
C4A
C5
C5A
C6
C6A
S2
O2S
O1S
O3S
O1W
O2W
O3W
O4W

595.8(32)
2379(6)
3347(7)
2430(22)
26031(48)
26241(42)
1351(16)
21825(14)
21582(26)
5400(23)
2666(19)
2857(20)
226(18)
1238(20)
1598(23)
23894(28)
1813(20)
23936(23)
2889(24)
3884(28)
175(22)
323(26)
834(21)
21417(28)
1034(24)
22287(33)
1983(30)
24745(35)
1293(23)
22342(27)
69(21)
1696(21)
0
230(53)
5332(20)
8258(43)
5000
10905(70)

1295.9(6)
1094(1)
1382(1)
2358(5)
762(11)
562(9)
1455(3)
1247(3)
1749(5)
610(5)
2087(4)
412(4)
1973(4)
617(4)
2763(5)
124(7)
1915(4)
931(6)
1690(5)
752(5)
2347(5)
317(5)
2331(5)
417(5)
2763(5)
49(7)
2348(6)
565(9)
1879(5)
879(6)
3977(3)
3697(5)
4506(11)
4072(14)
1983(4)
1029(11)
4255(17)
264(18)

241.5(9)
2669(2)
2106(2)
2767(5)
1916(14)
1680(13)
1083(4)
594(4)
21459(6)
2337(6)
2561(5)
168(6)
31(5)
399(6)
1910(6)
1566(8)
1904(5)
1194(7)
2893(7)
285(7)
386(7)
708(8)
1009(7)
971(7)
1325(7)
1310(10)
2232(8)
1544(10)
1316(6)
902(7)
2620(5)
2415(6)
2500
3256(17)
1298(6)
2421(14)
2500
2279(24)

362(7)
442(15)
469(16)
898(66)
823(91)
558(69)
441(39)
384(35)
815(72)
550(60)
421(47)
443(50)
380(44)
403(48)
647(63)
747(75)
524(52)
593(62)
456(58)
456(66)
445(61)
436(61)
334(52)
525(69)
476(63)
674(81)
672(84)
785(101)
381(55)
425(65)
598(27)
922(45)
1471(105)
1280(127)
751(53)
929(88)
2466(198)
1980(209)

Ueq 5(1 / 3)S i S jUij a *i a *j a i a j .

Table 3
Atomic coordinates (310 4 ) and equivalent isotropic displacement param 2 ) for non hydrogens atoms with e.s.d.s in parentheses for
eters (310 4 A
a
(2)
Atom

X /a

Y /b

Z /c

Ueq

Ni
CL1
CL2
S1
O1
O2
O1W
O2W
O3W
N1
N2
N3
N4
N5
C1
C2
C3
C4
C5
C6

8122.8(5)
8669.3(9)
6642.0(9)
8928.9(9)
7351(2)
5156(3)
7447(3)
8877(3)
9870(3)
8442(3)
7551(3)
7303(3)
5095(3)
6247(3)
8273(3)
6581(3)
6236(3)
5469(3)
5469(3)
6662(3)

687.9(7)
764.0(12)
21420.7(12)
2721.4(11)
1841(3)
3527(4)
708(3)
1794(4)
21248(3)
22632(4)
21393(3)
2378(4)
1770(4)
2628(3)
21627(4)
2249(4)
779(4)
852(4)
2705(5)
1747(4)

9839.1(14)
13305.5(22)
15874.1(25)
8945.4(22)
10562(6)
12721(7)
6898(6)
8869(8)
13507(8)
9668(8)
10563(7)
10682(7)
12608(7)
11675(7)
9780(8)
11355(8)
11644(8)
12303(9)
12346(8)
11244(8)

266(32)
370(32)
393(32)
319(32)
298(33)
405(34)
329(34)
455(35)
420(34)
352(35)
297(34)
249(34)
323(34)
264(34)
258(35)
259(35)
283(36)
307(36)
288(36)
242(35)

Ueq 5 (1 / 3)S i S jUij a *i a *j a i a j .

Ueq 5 (1 / 3)S i S jUij a *i a *j a i a j .

ion is disordered on two positions related by a 2-fold

crystallographic axis laying on the plane defined by O1S,
O2S, O92S atoms and passing through O1S and in the
middle between O2S and O92S. The sulphur atom and O3S
are statistically distributed on two positions with a factor
of occupance of 0.5 (Fig. 6). The oxidation state III of the
metal atom is evidenced by the deprotonation of the two
ligand molecules, by the charge of SO 24 with half multiplicity because of its special position, and even by the
coordination distances (Table 5), with values similar to
those previously reported for many other Co(III) complexes [26,27]. The five-member chelation rings present a f2
of 1528 and 21728, respectively, for both ligand molecules, both corresponding to twist-envelope conformations,
while the six-member ones are characterised by a f2 of
21328 and 2218 and a u of 788 and 548 showing,
respectively, a boat and a half chair conformation [21]. In

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

176

Table 5
and angles (8) for 1, 2, and 3
Selected bond distances (A)

MS
MO1
MN3
MO1W
MO2W
MCL1
S1C1
O1C6
O2C5
N1C1
N2N3
N2C1
N3C2
N4C4
N4C5
N5C5
N5C6
C2C3
C3C4
C3C6
O1MN3
S1MN3
S1MO1
O1WMN3
O1MO1W
S1MO1W
O2WMN3
O2WMS1
O2WMO1
O2WMO1W
CL1MN3
CL1MS1
CL1MO1
CL1MO1W
CL1MO2W
O1CoN3
O1ACoN3
O1CoN3A
N3CoS1A
N3ACoS1
S1CoO1A
C1ACoO1
N3CoN3A
S1CoS1A
O1CoO1A
a
b

Compound 1

Compound 2

2.260(1)
1.956(3)
1.974(3)
2.325(3)
1.933(3)

2.347(2)
2.029(4)
2.023(5)
2.232(4)
2.017(5)
2.473(2)
1.702(6)
1.241(6)
1.214(7)
1.334(7)
1.380(7)
1.348(7)
1.293(7)
1.358(8)
1.372(8)
1.371(7)
1.366(7)
1.464(8)
1.351(8)
1.466(8)

1.702(4)
1.250(4)
1.211(5)
1.307(6)
1.370(5)
1.351(5)
1.297(4)
1.341(5)
1.367(5)
1.374(4)
1.362(5)
1.438(5)
1.366(5)
1.440(5)
91.8(1)
87.3(1)
170.6(1)
97.5(1)
87.3(1)
102.1(1)
165.9(1)
94.0(1)
84.8(1)
96.0(1)

90.6(2)
85.7(1)
175.7(1)
87.1(2)
85.2(2)
92.4(1)
175.9(2)
96.5(1)
87.1(2)
89.4(2)
88.7(1)
95.2(6)
86.9(1)
171.0(1)
94.5(2)

Compound 3
2.185(4)
1.970(10)
1.921(10)

2.184(5)a
1.955(10)
1.904(10)

1.71(1)
1.24(2)
1.22(2)
1.35(2)
1.41(2)
1.30(2)
1.27(2)
1.35(2)
1.34(2)
1.37(2)
1.35(2)
1.45(2)
1.35(2)
1.42(2)

1.73(1)
1.25(2)
b
1.31(2)
1.41(2)
1.31(2)
1.25(2)
1.33(3)
1.34(3)
1.38(3)
1.35(2)
1.44(3)
1.39(3)
1.42(2)

95.0(4)
86.4(4)
176.5(3)

95.2(5)
86.2(4)
176.4(3)

95.0(4)
86.6(5)
88.1(5)
92.2(4)
90.6(4)
92.9(3)
92.8(4)
176.6(4)
90.3(2)
84.0(4)

The second set of values are for the ligand with A labels.
respectively.
The O2A atom is disordered on two positions; the O2AC5 bond distances are 1.36(4) and 1.15(5) (A),

3, the two ligand molecules are both monodeprotonated

and this fact produces a negative charge which should be
delocalized in the thiosemicarbazone moieties. As a consequence of the structural disorder, the worse refinement
degree led to larger error values in bond distances and it is
therefore difficult to discuss the bond distances in any
detail. The disorder of the sulphate group, of the C5A
O2A bond of one uracil group and in many water
molecules is a further cause of lack in crystal regularity
that could generate differences in bond lengths.

Hydrogen bonds between the disordered sulphate and N

atoms of the uracil ring belonging to the same asymmetric
and N atoms of nearby
unit (N4H???O2S52.73(2) A)
molecules (N4AH???O1S (2x21 / 2, y21 / 2, 1 / 22z)5
determine the packing. Complexes are piled
2.76(2) A)
along the b direction joining ribbons along this axis at 1 / 4
and 3 / 4 of z. These ribbons are linked by contact

interactions between ligands (N1AH???N2A53.04(2) A).

Couples of water molecules along the ribbon contribute to
the hydrogen bonds network.

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

177

Table 6
Comparison of relevant structural data of complexes [Cu(H 2 ut)(OH 2 )Cl]?2H 2 O and [Cu(H 3 ut)Cl 2 ]?2H 2 O [4] and compound 1
[Cu(H 2 ut)(OH 2 )Cl]?2H 2 O

[Cu(H 3 ut)Cl 2 ]?2H 2 O

Compound 1

CuS1
CuO1
CuN3
CuO1W
CuO2W
CuCL1
CuCL2
S1C1
O1C6
O2C5
N1C1
N2N3
N2C1
N3C2
N4C4
N4C5
N5C5
N5C6
C2C3
C3C4
C3C6

2.240(2)
1.983(5)
1.962(6)
1.966(7)

2.267(3)
1.998(6)
1.986(7)

2.260(1)
1.956(3)
1.974(3)
2.325(3)
1.933(3)

2.731(2)

2.665(3)
2.256(3)
1.706(9)
1.244(10)
1.200(12)
1.325(12)
1.387(10)
1.332(12)
1.275(11)
1.321(12)
1.389(12)
1.373(12)
1.351(11)
1.413(12)
1.377(13)
1.454(11)

O1CuN3
S1CuN3
S1CuO1
O1WCuN3
O1CuO1W
S1CuO1W
CL1CuN3
CL1CuO1W
CL1CuO1
CL1CuS1
CL2CuN3
CL2CuO1
CL1CuCL2
S1CuCL2
O2WCuN3
O2WCuS1
O2WCuO1
O2WCuO1W

92.8(2)
86.1(2)
165.8(2)
166.9(3)
84.2(2)
93.6(2)
90.5(2)
102.2(2)
89.8(2)
104.4(1)

1.738(7)
1.252(7)
1.232(8)
1.342(10)
1.397(8)
1.308(8)
1.316(8)
1.336(9)
1.362(9)
1.364(8)
1.364(8)
1.446(9)
1.366(9)
1.433(10)

5.2. Biological assay

Summing up the data obtained up to now from these
uracil derivatives concerning their biological activity on
human leukemic cell lines K562, CEM and U937, as far as
their potentiality as antitumor drugs is concerned, it can be
stated the following:
(a) nickel and cobalt complexes tested on K562 and
CEM lines have no inhibiting activity on cell proliferation up to a concentration of 40 mg / ml beyond which
the amount of DMSO necessary for the solubilization
can interfere with the experimental data;
(b) even though copper complex 1 inhibits cell growth
more than 50% (cell lines CEM, K562 U937) and with a
lower concentration with respect to copper complexes

89.8(3)
86.3(2)
169.4(2)

1.702(4)
1.250(4)
1.211(5)
1.307(6)
1.370(5)
1.351(5)
1.297(4)
1.341(5)
1.367(5)
1.374(4)
1.362(5)
1.438(5)
1.366(5)
1.440(5)
91.8(1)
87.1(1)
170.6(1)
97.5(1)
87.3(1)
102.1(1)

92.0(2)
93.6(2)
96.3(1)
166.2(2)
89.9(2)
101.8(1)
91.5(1)
165.9(1)
94.0(1)
84.8(1)
96.0(1)

with the same ligand and different counterion reported

in a previous work [4], in this case it does not induce
apoptosis.
These results have confirmed that copper complexes are
generally active in inhibiting proliferation and, as already
quoted in previous works of ours, among a series of
homologue complexes, those containing copper are the
most effective [3,5].
According to this screening procedure it can be observed
that: (a) the lack of biological activity of nickel and cobalt
octahedral complexes 2 and 3 could be ascribed to the
presence of a complete coordination sphere [3,5]; and (b)
copper complex 1, cationic with formal charge 12 which,
in the solid state, contains two coordinated water molecules creating a dimer-like situation by couples of hydro-

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

178

Fig. 6. Projection of the structure of complex 3 on the (100) plane. The hydrogen atoms have been omitted for clarity.

gen bonds, is more active in the proliferation assays in

comparison with the monomeric neutral complex
[Cu(H 2 ut)(OH 2 )Cl]?2H 2 O [4]. Nevertheless this monomeric complex induces apoptosis (cell lines CEM and
K562), while complex 1 does not cause the typical DNA
fragmentation on the same cell lines and also on the U937
line. We can conclude by stating that these substances
behave differently on cells under study, probably because
they act on different mechanisms. Studies are now in
progress in order to understand which are the mechanisms
in which our compounds intervene (signal transduction,
DNA replication, etc.).

Acknowledgements
This work was supported by Italian MURST (COFIN
98) in the frame of the Project Pharmacological and
Diagnostic Properties of Metal Complexes (Coordinator
Professor G. Natile) and by grants from Italian Association
for Cancer Research (AIRC).

References
R.C. Chikate, P.B. Sonawane,
[1] D.X. West, A.E. Liberta, S.B. Padhye,
A.S. Kunshan, R.G. Yerande, Coord. Chem. Rev. 123 (1993) 49.
[2] R.W. Sidwell, J.H. Huffman, T.W. Schafer, C. Shipman, Chemotherapy 36 (1990) 58.
[3] M. Belicchi Ferrari, G. Gasparri Fava, P. Tarasconi, R. Albertini, S.
Pinelli, R. Starcich, J. Inorg. Biochem. 53 (1994) 13.
[4] M. Belicchi Ferrari, G. Gasparri Fava, E. Leporati, G. Pelosi, R.
Rossi, P. Tarasconi, R. Albertini, A. Bonati, P. Lunghi, S. Pinelli, J.
Inorg. Biochem. 70 (1998) 145.
`

`
[5] M.C. Rodrguez
Arguelles,
A. Sanchez,
M. Belicchi Ferrari, G.
Gasparri Fava, C. Pelizzi, G. Pelosi, R. Albertini, P. Lunghi, S.
Pinelli, J. Inorg. Biochem. 73 (1999) 7.
[6] J.S. Casas, A. Castineiras, M.C. Rodriguez-Arguelles, A. Sanchez, J.
Sordo, A. Vazquez, A. Lopez, S. Pinelli, P. Lunghi, P. Ciancianaini,
A. Bonati, P. DallAglio, R. Albertini, J. Inorg. Biochem. 76 (1999)
277.
[7] U. Testa, F. Grignani, P. Samoggia, C. Zanetti, R. Riccioni, F.L.
Coco, D. Diverio, N. Felli, C.G. Passerini, M. Grell, P.G. Pelicci, C.
Peschle, J. Clin. Invest. 101 (1998) 2278.
[8] U. Testa, F. Grignani, H.J. Hassan, D. Rogaia, R. Masciulli, V.
Gelmetti, R. Guerriero, G. Macioce, C. Liberatore, T. Barberi, G.
Mariani, P.G. Pelicci, C. Peschle, Leukemia 12 (1998) 563.
[9] C. Nervi, F.F. Ferrara, M. Fanelli, M.R. Rippo, B. Tomassini, P.F.

M. Belicchi Ferrari et al. / Journal of Inorganic Biochemistry 83 (2001) 169 179

[10]
[11]

[12]
[13]
[14]

[15]
[16]
[17]
[18]

Ferrucci, M. Ruthardt, V. Gelmetti, C. Gambacorti Passerini, D.

Diverio, F. Grignani, P.G. Pelicci, R. Testi, Blood 92 (1998) 2244.
K.E. Knudsen, E. Weber, K.C. Arden, W.K. Cavenee, J.R.
Feramisco, E.S. Knudsen, Oncogene 18 (1999) 5239.
D. Belletti, A. Cantoni, G. Pasquinelli, Gestione on line di diffrattometro a cristallo singolo Siemens AED con sistema IBM PS 2 / 30,
Internal Report 1 / 88, Centro di Studio per la Strutturistica Diffrattometrica del CNR (1988).
M.S. Lehmann, F.K. Larsen, Acta Crystallogr. Sect. A 30 (1974)
580.
G. Sheldrick, in: SHELXL 86, Crystallographic Computing 3,
Oxford University Press, London, 1985.
A. Altomare, G. Cascarano, C. Giacovazzo, A. Guagliardi, M.C.
Burla, G. Polidori, M. Camalli, SIR 92, J. Appl. Crystallogr. 27
(1994) 435.
G. Sheldrick, in: SHELXL 97 A Program for Structure Refinement,
University of Goettingen, Germany, 1997.
G. Sheldrick, in: SHELXL 93, A Program for Structure Refinement,
University of Goettingen, Germany, 1993.
International Tables for X-Ray Crystallography, Vol. 4, Kynoch
Press, Birmingham, UK, 1975.
M. Nardelli, PARST95. An update to PARST: a system of Fortran

[19]
[20]
[21]
[22]

[23]
[24]
[25]
[26]
[27]

179

routines for calculating molecular structure parameters from the

results of crystal structure analyses, J. Appl. Crystallogr. 28 (1995)
659.
C.K. Johnson, ORTEP Report ORNL 3794, Oak Ridge National
Laboratory, Oak Ridge, TN (1965).
W.D.S. Motherwell, in: PLUTO, University Chemical Laboratory,
Cambridge, 1976.
D. Cremer, J. A Pople, J. Am. Chem. Soc. 97 (1975) 1354.
`

M.C. Rodrguez
Arguelles,
M. Belicchi Ferrari, G. Gasparri Fava,
C. Pelizzi, G. Pelosi, R. Albertini, A. Bonati, P.P. DallAglio, P.
Lunghi, S. Pinelli, J. Inorg. Biochem. 66 (1997) 7.
F.H. Allen, O. Kennard, Chemical Design Automation News 8(1)
(1993) 1, 31.
A. Marzotto, D.A. Clemente, G. Valle, Acta Crystallogr. C49 (1993)
1252.
R. Cortes, M.I. Arriortra, T. Rojo, X. Solans, C. Miravitlles, D.
Beltran, Acta Crystallogr. C41 (1985) 1733.
M. Belicchi Ferrari, G. Gasparri Fava, M. Lanfranchi, C. Pelizzi, P.
Tarasconi, J. Chem. Soc. Dalton Trans. (1991) 1951.
`
M. Belicchi Ferrari, G. Gasparri Fava, G. Pelosi, M.C. Rodrguez

Arguelles,
P. Tarasconi, J. Chem. Soc. Dalton Trans. (1995) 3035.