Crystal structure and solution chemistry of the cytotoxic complex 1,2-dichloro(o-phenanthroline)gold(III) chloride

Article abstract of DOI:10.1016/S0020-1693(00)00299-1

The crystal structure of the cytotoxic complex 1,2-dichloro(o-phenanthroline)gold(III) chloride ([AuphenCl₂]Cl) has been solved through single crystal X-ray diffraction methods. The complex is square planar and exhibits a quite regular geometry. Crystals of the compound belong to the space group P2₁/n with a = 12.632(5), b = 16.916(3), c = 12.902(6) ?, β = 91.31(3)° and Z = 8. The coordination of the two gold(III) ions in the asymmetric unit is completed by two chloride ions at 2.972(3) and 3.043(3) ?, respectively, forming a distorted square pyramid. The behavior in solution of [AuphenCl₂]Cl was further analyzed through ¹H NMR spectroscopy. Results point out that the [Au(III)phen]³⁺ molecular fragment is stable in solution for several hours, even under physiological conditions, whereas the two chloride ligands are released within approximately 30 min after dissolution in the buffer, at 25°C. The gold(III) chromophore is easily and quickly reduced by addition of stoichiometric amounts of sodium ascorbate; metallic gold is formed and free phenanthroline liberates. The implications of these findings for the biological properties of the [Au(III)phen]³⁺ species are discussed.

Full text of DOI:10.1016/S0020-1693(00)00299-1

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Inorganica Chimica Acta 311 (2000) 1–5
Crystal structure and solution chemistry of the cytotoxic complex
1,2-dichloro(o-phenanthroline)gold(III) chloride
F. Abbate a, P. Orioli a,*, B. Bruni a, G. Marcon b, L. Messori a
a Department of Chemistry, Uni6ersity of Florence, 6ia Gino Capponi 7, 50121 Florence, Italy
b CIRCMSB, Unit of Florence, Florence, Italy
Received 29 May 2000; accepted 24 July 2000
Abstract
The crystal structure of the cytotoxic complex 1,2-dichloro(o-phenanthroline)gold(III) chloride ([AuphenCl2]Cl) has been solved
through single crystal X-ray diffraction methods. The complex is square planar and exhibits a quite regular geometry. Crystals of
,
the compound belong to the space group P21/n with a=12.632(5), b=16.916(3), c=12.902(6) A, i=91.31(3)° and Z=8. The
,
coordination of the two gold(III) ions in the asymmetric unit is completed by two chloride ions at 2.972(3) and 3.043(3) A,
1
respectively, forming a distorted square pyramid. The behavior in solution of [AuphenCl2]Cl was further analyzed through H
NMR spectroscopy. Results point out that the [Au(III)phen]3+ molecular fragment is stable in solution for several hours, even
under physiological conditions, whereas the two chloride ligands are released within approximately 30 min after dissolution in the
buffer, at 25°C. The gold(III) chromophore is easily and quickly reduced by addition of stoichiometric amounts of sodium
ascorbate; metallic gold is formed and free phenanthroline liberates. The implications of these findings for the biological properties
of the [Au(III)phen]3+ species are discussed. © 2000 Elsevier Science B.V. All rights reserved.
Keywords: Gold complexes; Phenanthroline; X-ray crystal structures
1. Introduction
complex [AuphenCl2]Cl is highly cytotoxic toward the
A2780 tumor cell line either sensitive or resistant to
cisplatin; some aspects of the solution chemistry of
[AuphenCl2]Cl were also described [7]. To further eluci-
date the relationships existing between the biological
activity and the chemical properties of [AuphenCl2]Cl,
we report here the crystal structure of this complex and
describe in more detail its solution behavior. In particu-
lar, our interest focuses on the identification of the
dominant species existing in solution, under physiologi-
cal conditions. The chemical and biological properties
of [AuphenCl2]Cl are critically compared to those of
similar metallo-phenanthroline complexes.
For the last 5 years new interest has focused on
gold(III) complexes as possible cytotoxic and antitumor
agents [1–3]. Indeed, a few gold(III) complexes were
shown to be sufficiently stable in buffered solutions,
owing to stabilization effects played by the ligands, and
to exhibit interesting cytotoxic properties when tested in
vitro on selected tumor cell lines [1]. For instance
Buckley et al. reported that the complex Audamp shows
promising antitumor properties both in vitro and in
vivo [4]. Henderson et al. prepared and characterized a
number of organogold(III) complexes that showed in-
teresting antibacterial, antitumor and antifungal prop-
erties [5]. Three representative gold(III) polyamine
complexes were recently investigated in our laboratory,
and two of them turned out to exhibit relevant cyto-
toxic properties [6]. Moreover, we have shown that the
2. Experimental
2.1. Synthesis of [AuphenCl2]Cl
* Corresponding author. Tel.: +39-055-275 7554; fax: +39-055-
275 7555.
E-mail address: orioli@risc1.cerm.unifi.it (P. Orioli).
[AuphenCl2]Cl was prepared by addition of
phenantroline·HCl to a HAuCl4 solution at a 1:1 stoi-
0020-1693/00/$ - see front matter © 2000 Elsevier Science B.V. All rights reserved.
PII: S0020-1693(00)00299-1

2
F. Abbate et al. / Inorganica Chimica Acta 311 (2000) 1–5
chiometry according to the procedure reported in Ref.
[8] (using ethanol as solvent). The obtained product was
checked by elemental analysis; the purity of the com-
pound was higher than 95%. Further evidence for the
correct identification of the obtained compound is pro-
vided by the electronic and mass spectra.
freshly prepared, concentrated water solutions of
[AuphenCl2]Cl to the reference buffer (50 mM phos-
phate, 4 mM NaCl, pH 7.4 at 25°C) and monitoring
the electronic spectra of the resulting mixtures over 72
h. The stability of the chromophore in water was also
investigated. Potentiometric measurements of chloride
release were performed by using a chloride selective
electrode interfaced to a Hanna potentiometer. The
apparatus was calibrated with sodium chloride solu-
tions of known concentration.
2.2. X-ray structure determination
Crystals of [AuphenCl2]Cl were obtained from the
mother liquor of the reaction. X-ray diffraction data
were collected on a single crystal X-ray Enraf-Nonius
CAD4 diffractometer using the Ka radiation of Mo
1
2.4. H NMR studies
Solution 1H NMR spectra of [AuphenCl2]Cl were
recorded on a Bruker Avance spectrometer operating at
600 MHz. Solutions (1.0 mM) were prepared in D2O
and in 50 mM PO43−, 100 mM NaCl, pH 7.4 buffer.
The spectra were recorded immediately after dissolution
and over a period of 1–3 h.
,
(u=0.71073 A) at room temperature (r.t.). Cell
parameters were obtained from 25 well-centered reflec-
tions by using least-square refinement in the range
85q515; relevant crystal data are reported in Table
1. The absorption correction was done by using the
PSISCAN procedure. The structure was solved by using
direct methods and successive difference Fourier syn-
thesis; an alternative approach was also used starting
with Patterson method. All non-hydrogen atoms were
refined anisotropically and the hydrogen atoms were
placed in calculated positions. Calculations were carried
on with the SIR-97 [9] and SHELX-97 [10] programs.
3. Results and discussion
3.1. Crystal structure of [AuphenCl2]Cl
The crystal structure of [Au(phen)Cl2]Cl shows the
presence of two molecules of the complex in the asym-
metric unit (Fig. 1). Analogous bond lengths and angles
in the two molecules show only differences within few
standard deviations. In both molecules the gold(III)
ions show a slightly distorted square planar coordina-
tion formed by two chloride ions and the two nitrogen
atoms of the bidentate 1,10-N,N%-phenanthroline lig-
and. Bond lengths and angles around the gold(III) ions
and in the phenanthroline ligands are in agreement with
values from the literature (Tables 2 and 3) [11]. The
least squares planes through the coordination planes
and the phenanthroline ligands show deviations from
planarity within a few standard deviations. The angles
between the coordination and the phenanthroline
planes are 4.4° and 1.1°, respectively, for the two
complex molecules. The two molecules are not totally
parallel, indeed the angle between the mean plane of
the two molecules is 4.5°.
2.3. Solution studies
The absorption spectra in the UV–Vis region were
recorded on a Perkin–Elmer Bio 20 spectrophotometer
operating at r.t. The gold(III) complex [AuphenCl2]Cl
is soluble in water (10−2 M solutions are easily ob-
tained); electronic spectra were recorded on freshly
prepared buffered solutions at r.t. The hydrolysis exper-
iments were carried out by adding small amounts of
Table 1
Crystallographic data for [Au(phen)Cl2]Cl
Formula
M
Space group
Unit cell parameters
C12H8N2Cl3Au1
483.5
P21/n
,
a (A)
12.632(5)
16.916(3)
12.902(6)
91.31(3)
2756.21(1.75)
8
2.33
1872
11.25
0.71069
2–25
54
4823 (unique reflections)
0.0415 for 3861 Fo\4|(Fo)
0.0571 for all 4823 data
,
b (A)
In both molecules the gold(III) ions are additionally
,
c (A)
,
i (°)
coordinated by chloride ions at 2.972(3) and 3.043(3) A,
3
,
V (A )
Z
respectively, for the two molecules, forming the apices
of distorted square pyramids. The sixth positions are
Dcalc (g cm−3
F(000)
)
,
filled by a chloride ion at 3.54 A for one gold(III) ion
v (Mo Ka) (mm−1
)
and by an aromatic ring of a phenanthroline ligand for
the other gold(III) ion. The chloride ions at the apices
of the square pyramids are hydrogen bonded to the
,
u (A)
qmin–qmax
Number of atoms
Independent reflections
R1
R1
,
water molecules (Cl···O=3.12 and 3.15 A).
The square pyramidal or tetragonal bipyramidal
stereochemistries attained with secondary bonds are
quite common for gold(III) complexes as it can be seen

F. Abbate et al. / Inorganica Chimica Acta 311 (2000) 1–5
3
,
a distance of 2.58 A from the gold(III) ion. The differ-
ence between the stereochemistries of the two com-
plexes is undoubtedly attributable to the steric
hindrance of the bulky methyl groups which do not
allow the coplanarity of the dmphen ligand with the
chloride ions.
3.2. Solution chemistry of [AuphenCl2]Cl
[AuphenCl2]Cl is pretty soluble in water and within a
standard physiological buffer; freshly prepared solu-
tions of [AuphenCl2]Cl are bright yellow. As previously
reported, the electronic spectrum of [AuphenCl2]Cl in
water and in the buffer is characterized by a number of
intense bands in the visible falling between 300 and 450
nm [7], assigned as N-to-gold(III) charge transfer
Table 2
,
Selected bond lengths (A) of the two molecules in the asymmetric unit
for [Au(phen)Cl2]Cl
Au1–Cl1
Au1–Cl2
Au1–Cl3
Au1–N1
Au1–N2
N1–C1
N1–C2
N2–C11
N2–C12
C1–C12
C1–C5
C2–C3
C3–C4
C4–C5
C5–C6
C6–C7
C7–C8
C8–C9
2.972(3)
2.266(3)
2.263(3)
2.033(8)
2.056(8)
1.38(1)
1.34(1)
1.30(1)
1.37(1)
1.42(1)
1.41(1)
1.40(1)
1.35(2)
1.41(1)
1.41(1)
1.37(2)
1.45(1)
1.39(2)
1.40(1)
1.36(2)
1.41(1)
Au1%–Cl1%
Au1%–Cl2%
Au1%–Cl3%
Au1%–N1%
Au1%–N2%
N1%–C1%
N1%–C2%
N2%–C11%
N2%–C12%
C1%–C12%
C1%–C5%
C2%–C3%
C3%–C4%
C4%–C5%
C5%–C6%
C6%–C7%
C7%–C8%
C8%–C9%
3.043(3)
2.256(3)
2.268(3)
2.044(8)
2.045(8)
1.37(1)
1.32(1)
1.34(1)
1.38(1)
1.40(1)
1.40(1)
1.39(2)
1.37(2)
1.40(2)
1.46(2)
1.34(1)
1.43(1)
1.40(1)
1.39(1)
1.38(2)
1.38(2)
C8–C12
C9–C10
C10–C11
C8%–C12%
C9%–C10%
C10%–C11%
Table 3
Fig. 1. Crystal structure of [AuphenCl2]Cl.
Selected angles (°) of the two molecules in the asymmetric unit for
[Au(phen)Cl2]Cl
by comparison with the analogous structure of
[Auphen(CN)2]Br [11]. In the paper, which describes
this structure, the authors make an interesting survey of
102 square planar gold(III) complexes and find that
nearly 15% of them display secondary coordination.
It is also interesting to compare the present structure
with that of trichloro-(2,9-dimethyl-1,10-phenanthro-
line)gold(III) [12]. In this structure the gold(III) ion
shows a distorted square pyramidal coordination, but
the three chloride ions lie in the coordination plane
with one nitrogen atom of the 2,9-dimethyl-1,10-
phenanthroline (dmphen) ligand, whereas the second
nitrogen atom of the ligand is in the apical position at
Cl1–Au1–Cl2
Cl1–Au1–Cl3
Cl1–Au1–N2
Cl1–Au1–N1
Cl2–Au1–Cl3
Cl2–Au1–N1
Cl2–Au1–N2
Cl3–Au1–N1
Cl3–Au1–N2
Au1–N1–C1
Au1–N1–C2
Au1–N2–C12
Au1–N2–C11
N1–Au1–N2
93.1(1)
97.0(1)
90.7(2)
86.7(2)
89.5(1)
94.2(2)
174.5(2)
174.6(2)
94.0(2)
112.1(6)
129.0(7)
111.1(6)
127.6(7)
82.0(3)
Cl1%–Au1%–Cl2%
Cl1%–Au1%–Cl3%
Cl1%–Au1%–N2%
Cl1%–Au1%–N1%
Cl2%–Au1%–Cl3%
Cl2%–Au1%–N1%
Cl2%–Au1%–N2%
Cl3%–Au1%–N1%
Cl3%–Au1%–N2%
Au1%–N1%–C1%
Au1%–N1%–C2%
Au1%–N2%–C12%
Au1%–N2%–C11%
N1%–Au1%–N2%
92.9(1)
91.1(0)
89.7(2)
91.6(2)
90.1(1)
93.6(2)
175.2(2)
175.2(2)
93.9(2)
110.6(6)
129.4(7)
111.2(6)
128.4(7)
82.3(3)

4
F. Abbate et al. / Inorganica Chimica Acta 311 (2000) 1–5
containing gold in the oxidation state +3 is present
either in water or in the buffer.
Potentiometric experiments were carried out, with the
aid of
a chloride-selective electrode, to establish
whether the dominant species in solution still contains
gold(III)-coordinated chlorides. Potentiometric profiles
are shown in Fig. 2. Notably, a progressive large de-
crease of the electrode potential is observed within
approximately 80 min after dissolution, indicating an
increase of chloride concentration. The total change of
potential equals 40 mV: such a value roughly corre-
sponds to an increase in chloride concentration from
ꢀ1 to ꢀ2.8 mM. From analysis of this data it is
inferred that the [AuphenCl2]+ species slowly releases
both gold-coordinated Cl groups. The hydrolysis pro-
cess is clearly biphasic, as it was observed also for
[Au(Oac)2(damp) [13], with the fast phase lasting ap-
proximately within 15 min and the slow phase reaching
completion within ꢀ60 min. It is straightforward to
assign the fast phase to the release of the first Cl group
and the slow phase to the release of the second Cl
group.
Fig. 2. Potentiometric profiles of a freshly prepared [AuphenCl2]Cl
solution. The graph shows the values of the electrode potentials of a
chloride selective electrode against time. Experimental conditions: 1
mM [AuphenCl2]Cl was dissolved in a buffer 50 mM phosphate, pH
7.4 at 25°C and the electrode potential was monitored continuously
for 3 h.
The solution behavior of [AuphenCl2]Cl was further
1
analyzed by H NMR spectroscopy. Spectra are shown
in Fig. 3. Upon dissolving [AuphenCl2]Cl in the refer-
ence physiological buffer (50 mM phosphate, 100 mM
NaCl, pH 7.4) several signals, of variable intensity, are
observed in the aromatic region, that are suggestive of
the simultaneous presence of at least three species under
slow exchange conditions (Fig. 3(a)). A significant spec-
tral simplification then takes places within 2 h after
dissolution (at r.t.) indicating that one species becomes
1
dominant (Fig. 3(b)). Careful analysis of the H NMR
1
spectra allows assignment of the H NMR signals to
three well distinct species most likely corresponding to
the dichloro species [AuphenCl2]+, the monochloro
species [Auphen(OH)Cl]+ and the dihydroxo species
[Auphen(OH)2]+. Signals are attributed to each species
by analysis of the time dependent changes in intensity
1
and are labeled accordingly. Notably H NMR signals
attributed to the [AuphenCl2]+ and [Auphen(OH)Cl]+
species decrease progressively in intensity with time
whereas signals assigned to the [Auphen(OH)2]+ species
become dominant after approximately 30 min. Later
1
on, the spectrum remains stable for hours. The final H
1
Fig. 3. 600 MHz H NMR spectra of [AuphenCl2]Cl in the buffer (50
NMR spectrum corresponding to the end-point of the
kinetic process shows that, under the experimental con-
ditions employed, an equilibrium is reached where the
dihydroxo species is largely predominant but signals of
the monochloro and dichloro species are still observed.
The effects of the addition of the reducing agent
sodium ascorbate on the spectral properties of
[AuphenCl2]Cl were also analyzed. The electronic spec-
tra show immediate disappearance of the characteristic
LMCT transitions suggesting that sodium ascorbate
mM phosphate, 100 mM NaCl, pH 7.4) at mixing (a) and after 2 h
(b); the individual species are labeled as follows: [AuphenCl2]+ (ꢀ),
[Auphen(OH)Cl]+ (ꢁ), [Auphen(OH)2]+ (ꢂ).
bands. Notably the visible spectra of [AuphenCl2]Cl, in
water and in the buffer, do not change appreciably with
time when monitored over a time period of 24 h,
implying that the [Au(phen)]3+ chromophore is stable.
These results point out that the same chromophore

F. Abbate et al. / Inorganica Chimica Acta 311 (2000) 1–5
5
Acknowledgements
causes immediate and complete reduction of the gold(III)
1
center; accordingly the H NMR spectra show that the
1H NMR signals of [AuphenCl2]Cl suddenly disappear.
It is inferred that, upon reduction, the complex breaks
down and free phenanthroline is released. In nice agree-
MURST is acknowledged for financial support in the
frame of the project ‘Pharmacological and diagnostic
aspects of metal complexes’. The Cassa di Risparmio di
Firenze is gratefully acknowledged for a generous
grant. We thank Professor Marco Mascini, Dr. Ilaria
Palchetti and Dr. Costanza Landi for helping with
chloride selective potentiometric measurements.
1
ment with this hypothesis, the H NMR signals of free
phenanthroline are newly detected after reduction;
simultaneously formation of characteristic Cassius por-
pora (colloidal gold) is observed.
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[AuphenCl2]Cl exhibits relevant cytotoxic properties
toward cultured human tumor cells. Single crystal X-
ray diffraction studies demonstrate that [AuphenCl2]Cl
is a classical square planar gold(III) complex. Solution
studies show that the molecular fragment [Au(phen)]3+
is relatively stable under physiological conditions
whereas the chloride groups are slowly released after
dissolution in the buffer. The predominant species exist-
ing in solution under physiological conditions is likely
to be [Auphen(OH)2]+; this species may interact with
biomolecules through two mechanisms: (i) formation of
monofunctional or bifunctional coordination bonds at
the two (labile) cis positions on the gold(III) center; (ii)
intercalation of the phenanthroline moiety. Neverthe-
less, it must be remembered that the complex investi-
gated is promptly reduced by a relatively mild reducing
agent such as ascorbate, implying that the biological
properties of [AuphenCl2]Cl might derive at least par-
tially from free phenanthroline. Indeed, previous stud-
ies showed that the free ligand phenanthroline is
endowed with significant cytotoxic properties [7]. Fur-
ther studies are now needed to identify unambiguously
the species that are actually responsible for the cyto-
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