Therapeutic properties, SOD and catecholase mimetic activities of novel ternary copper(II) complexes of the anti-inflammatory drug Fenoprofen with imidazole and caffeine

Article abstract of DOI:10.1016/j.poly.2011.12.005

The copper(II) ternary complexes of the non-steroidal anti-inflammatory drug Fenoprofen (Hfen) and the biologically relevant molecules imidazole (im) and caffeine (caf) as auxiliary ligands were investigated as novel anti-inflammatory agents. The new copper(II) complexes with formula [Cu(fen)₂(im)₂] (1) and Cu₂(fen) ₄(caf)₂ (2) were synthesized from the dinuclear complex [Cu₂(fen)₄(dmf)₂] and characterized by IR, UV-Vis, EPR spectral and elemental analysis. The molecular structure of complex 1 was determined by X-ray crystallography. Both complexes 1 and 2 present enhanced and prolongued anti-inflammatory properties against the parent drug calcium Fenoprofenate, Ca(fen)₂·2H₂O, with a better performance for complex 1. Ternary complexes are potential models for several mono and poly-nuclear metal enzymes. The measured superoxide dismutase (SOD) mimetic activities of the complexes indicated a higher SOD mimic activity for complex 2 (IC₅₀ of 0.24 μM) than complex 1 (IC₅₀ of 0.70 μM), and also than the native enzyme evaluated by the same method (IC₅₀ of 0.480 μM). The catecholase activity of the complexes toward the aerobic oxidation of 3,5-di-tert-butylcatechol (dtbc) onto 3,5-di-tert-butylquinone (dtbq) showed that both complexes have moderate catalytic oxidase activity.

Full text of DOI:10.1016/j.poly.2011.12.005

Polyhedron 34 (2012) 74–83
Contents lists available at SciVerse ScienceDirect
Polyhedron
journal homepage: www.elsevier.com/locate/poly
Therapeutic properties, SOD and catecholase mimetic activities of novel
ternary copper(II) complexes of the anti-inflammatory drug Fenoprofen with
imidazole and caffeine
a
b
c
d
Mariela A. Agotegaray a, , Mariana Dennehy , Mónica A. Boeris , María A. Grela , Robert A. Burrow ,
⇑
Oscar V. Quinzani a
a INQUISUR, Departamento de Química, Universidad Nacional del Sur, Avda. Alem 1253, B8000CPB, Bahía Blanca, Argentina
b Centro de Investigación y Desarrollo de Fármacos (CIDEF), Facultad de Ciencias Veterinarias, Universidad Nacional de La Pampa, Calle 5 y 116, General Pico, CP 6360, Argentina
c Departamento de Química, Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, Funes N° 3350, 7602AYL Mar del Plata, Argentina
d Laboratório de Materiais Inorgánicos, Departamento de Química, Universidade Federal de Santa María, 97105-900 Santa María, RS, Brazil
a r t i c l e i n f o
a b s t r a c t
Article history:
The copper(II) ternary complexes of the non-steroidal anti-inflammatory drug Fenoprofen (Hfen) and the
biologically relevant molecules imidazole (im) and caffeine (caf) as auxiliary ligands were investigated as
novel anti-inflammatory agents. The new copper(II) complexes with formula [Cu(fen)2(im)2] (1) and
Cu2(fen)4(caf)2 (2) were synthesized from the dinuclear complex [Cu2(fen)4(dmf)2] and characterized
by IR, UV–Vis, EPR spectral and elemental analysis. The molecular structure of complex 1 was determined
by X-ray crystallography. Both complexes 1 and 2 present enhanced and prolongued anti-inflammatory
properties against the parent drug calcium Fenoprofenate, Ca(fen)2ꢀ2H2O, with a better performance for
complex 1. Ternary complexes are potential models for several mono and poly-nuclear metal enzymes.
The measured superoxide dismutase (SOD) mimetic activities of the complexes indicated a higher SOD
Received 5 October 2011
Accepted 9 December 2011
Available online 24 December 2011
Keywords:
Copper
Fenoprofen
Anti-inflammatory
SOD
mimic activity for complex 2 (IC50 of 0.24
lM) than complex 1 (IC50 of 0.70
lM), and also than the native
Catecholase
enzyme evaluated by the same method (IC50 of 0.480
l
M). The catecholase activity of the complexes
toward the aerobic oxidation of 3,5-di-tert-butylcatechol (dtbc) onto 3,5-di-tert-butylquinone (dtbq)
showed that both complexes have moderate catalytic oxidase activity.
Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction
matory properties when compared to the uncomplexed parent
drug [15], however, this behavior has not been observed for some
A great amount of data about the medicinal properties of small
synthetic copper(II) complexes with carboxylato ligands recog-
nized as non-steroidal anti-inflammatory drugs (NSAIDs) have
been developed since the pioneering studies of Sorenson et al.
[1,2]. These studies of the anti-inflammatory properties of binary
and ternary copper(II) carboxylates were extended to other medic-
inal properties of this now wide family of compounds by several
research groups. Anti-inflammatory [3], antioxidant and superox-
ide scavenging [3–7], catalytic and catecholase [4], anticonvulsant
[8], DNA binding and nuclease [6,9–11], renal and gastrointestinal
toxicity [11] and antimicrobial [5,12] properties have been re-
ported for numerous copper(II)–NSAID complexes. A great part of
those studies have been organized in an excellent review by Lay
and co-workers [15].
AINEs [16]. One mechanism of action proposed for this enhanced
properties is that these complexes present antioxidant properties
due to their capacity to mimic the activity of the superoxide dis-
mutase enzyme [15]. Oxidative stress is implicated in pathophysi-
ological processes, such as inflammation, rheumatoid arthritis and
carcinogenesis [17]. The main species involved in the above men-
tioned processes is superoxide radical (O2ꢀꢁ), an important regula-
tor of cell death [18]. Superoxide dismutase, Cu,Zn-SOD is a
metalloprotein which catalyzes the scavenging of superoxide anion
O2ꢁ. Copper is present in the catalytic center and undergoes reduc-
tion–oxidation cycling during the dismutation of O2ꢁ. Although
zinc ion is not involved in this cycle, it facilitates the oxidation step
by maintaining the configuration of the active site [19].
The use of SOD enzyme as a diet supplement in patients pre-
senting inflammatory and oxidative disorders is limited not only
by costs, but also because of its high molecular weight and low sta-
bility. These facts make the research on SOD-mimic agents a rele-
vant issue.
It is known that many copper(II) complexes with non-steroidal
anti-inflammatory drugs (NSAIDs) present enhanced anti-inflam-
⇑
Corresponding author.
E-mail address: magotegaray@uns.edu.ar (M.A. Agotegaray).
0277-5387/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.poly.2011.12.005

M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
75
Some copper(II) complexes also present catechol oxidase mimic
2.3. Preparation of complexes
activity. Catechol oxidase is a dicopper enzyme and belongs to the
polyfenol oxidases [20]. It catalyzes the two-electron oxidation of
o-difenols to the corresponding quinones. The role of this enzyme
is related to the biological synthesis of polyfenolic pigments such
as melanins, which protect tissues against pathogens [20].
Fenoprofen (Scheme 1), which systematic name is 2-(3-phen-
oxyphenyl)propanoic acid, is a non-steroidal anti-inflammatory
drug employed in inflammatory pathologies such as rheumatoid
arthritis. It exerts its anti-inflammatory activity by inhibiting cic-
looxigenase enzyme, which participates in the inflammatory re-
sponse [21]. In a previous work we have presented the first
crystal structure of a dinuclear copper(II) complex of Fenoprofen
and a full characterization of its structure and biological properties
[22,23].
2.3.1. Bis[2-(3-fenoxyfenyl)-propionato]bis(imidazole)copper(II),
Cu(fen)2(im)2 (1)
A solution of 11.0 mg (0.160 mmol) of imidazole in 1 mL of ace-
tone was added to another solution containing 50.0 mg
(0.0400 mmol) of Cu2(fen)4(dmf)2 in acetone. The blue solution
was kept under stirring for about 30 min and then diffused with
acetonitrile. After a few days at 4 °C, the obtained blue crystals
were washed with acetonitrile and air dried. Yield: 28 mg (51%).
Suitable single crystals were selected for X-ray diffraction analysis.
Anal. Calc. for C36H34CuN4O6: C, 63.4; H, 4.9; N, 8.2%. Found: C,
62.4; H, 4.8; N, 8.3%. Several samples were used for the carbon con-
tent determination and reduced percentages were always
obtained.
Motivated by the interest that represent copper(II) complexes
of NSAIDs we continue our work in the synthesis of new Fenopro-
fen–copper(II) complexes. In this paper we present the first mono-
nuclear copper(II) complex of Fenoprofen with imidazole as
auxiliary ligand. The coordination chemistry of imidazoles has
been subjected to intensive studies because the interaction of his-
tidine residues and metal ions is relevant in many active biological
systems. Herein we also report the spectroscopic and biological
characterization of a new dinuclear Cu(II) complex of Fenoprofen
containing caffeine in its structure. Caffeine is a xanthine alkaloid
with stimulating activity. As part of the biological characterization,
we compare the anti-inflammatory properties of the synthesized
compounds with the uncomplexed commercial parent drug Feno-
profen calcium salt. The SOD-mimic activity is evaluated. Besides,
catechol oxidase mimic activity is also presented as model for
the catecholase function.
2.3.2. Tetrakis[l
-2-(3-fenoxyfenyl)-propionato-j2O:O0]bis[(1,3,7-
trimethyl-1H-purine-2,6(3H,7H)-dione)copper(II)], Cu2(fen)4(caf)2 (2)
The addition of a solution containing 50 mg (0.040 mmol) of
Cu2(fen)4(dmf)2 in 2 mL of acetone to 31 mg (0.160 mmol) of caf-
feine dissolved in 2 mL of hot ethanol under stirring, led to a result-
ing limpid, green solution. The slow diffusion of water resulted in
the formation of two layers of solvents. After a few days at 4 °C
green crystals were obtained, which were washed with water
and air dried. Yield: 34 mg (57%). The well-shaped crystals could
not be solved by X-ray crystallography due to molecular disorder.
Anal. Calc. for C76H72Cu2N8O16: C, 61.6; H, 4.9; N, 7.6%. Found: C,
61.3; H, 4.8; N, 7.7%.
2.4. X-ray crystallography
The X-ray data collection and processing for complex 1 were
performed on a Bruker APEX2/BIS/COSMO diffractometer by using
2. Experimental
graphite monochromated Mo
Ka radiation (k = 0.71073 Å) at
2.1. General remarks
100(2) K. Cell refinement was done with Bruker SAINT and data
reduction by Bruker SAINT/SADABS/XPREP. Programs used to solve
structure were SHELX97; programs used to refine structure were SHEL-
XL97. Images were created with Crystal Impact Diamond 3 molecu-
lar graphics [12].
All reagents were of commercial analytical quality and used
without further purification. The dinuclear complex Cu2(-
fen)4(dmf)2 was synthesized following the procedure published
elsewhere [8]. Chemical analyses of carbon, hydrogen and nitrogen
have been performed using a Thermo Electron Flash EA instrument
at UMYMFOR (FCEyN, UBA, Argentina).
2.5. Evaluation of anti-inflammatory properties
To evaluate the anti-inflammatory activity of complexes 1 and
2, carrageenan induced paw oedema assay was carried out as de-
scribed by Winter et al. [13]. Female mice of about 30 g were ran-
domly divided into three groups. Each group contained six mice
fasted for 18 h. Test animals were orally administered an aqueous
suspension of the copper(II) complexes under study (31 mg/kg for
complex 1 and 28 mg/kg for complex 2) and the calcium salt of
Fenoprofen (22 mg/kg). Each quantity of drug administered is
equivalent to 20 mg/kg of Fenoprofen. The vehicle alone was used
as excipient for the control group. The drugs were suspended in
carboxymethylcellulose at 0.1% and Tween 80 at 0.05% (1:1, v/v)
just before use. Drug and excipient were orally administered in
0.5 mL of the corresponding suspension to each animal one-hour
before inducing oedema in the left hind paw by sub-plantar injec-
tion of 0.05 ml of a 2% suspension of carrageenan.
2.2. Spectroscopic studies
IR spectra were recorded in KBr pellets and Nujol mulls with a
Nicolet Nexus FT-IR spectrophotometer in the range 400–
4000 cmꢁ1. Raman spectra of the solids, in the region between
3500 and 100 cmꢁ1, were obtained with a FRA 106 accessory
mounted in a Bruker IFS 66 FTIR instrument, using a 1064 nm exci-
tation line from an Nd-YAG laser. The UV–Vis spectra were regis-
tered on a GBC-Cintra 20 equipment in the region between 190
and 900 nm. CW EPR spectra of solid samples placed in quartz cap-
illary tubes were obtained, at room temperature, with a Bruker
ELEXSYS E500T A spectrometer, operating at X band (ꢂ9.8 GHz),
with a Bruker cavity ER 4102ST and a modulation frequency of
100 KHz. Measurements of g values were made relative to TEMPO
(g = 2.0051) [10,11].
The length of the paw was measured with a digital electronic
caliber ‘‘Caliper’’ (0.1 mm resolution) immediately before the injec-
tion of carrageenan and 3, 5, 7 and 9 h after the injection. The anti-
inflammatory effect of the test was expressed in terms of the per-
cent inhibition of oedema produced by each drug-treated group
CH3
O
COOH
and was calculated as [(
of the control group and
D
D
C ꢁ
D
T)/
D
C] ꢃ 100 being
DC the mean
T the mean of the test group. The data
Scheme 1. Structure of Fenoprofen (Hfen).
were expressed as mean SEM.

76
M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
Student’s t test has been applied in order to determine signifi-
ibrated with atmospheric oxygen. The mean value of three exper-
iments was determined using methanolic solution of the
cant differences between the mean responses of the treated and
control groups. The difference in the mean response between the
group treated with the complex and one treated with the Fenopro-
fen calcium salt (Aldrich) was also evaluated. Statistical signifi-
cance was set as P < 0.05.
The care and the handling of the animals were in accordance
with the internationally accepted standard Guide for the Care
and Use of Laboratory Animals as adopted and promulgated by
the National Institute of Health.
a
substrate without the complex as a blank. Concentrations were
evaluated from the absorbance measurements using the reported
extinction molar coefficient, e
400nm = 1900 L molꢁ1 cmꢁ1 for 3,5-
dtbq in methanol [17]. Time profiles were adjusted with a polyno-
mial function and the initial rates were evaluated as the derivative
of this function at t = 0–2 min. The same treatment was applied to
a methanolic solution of CuCl2 in order to obtain the same final
copper(II) concentration of the complex solution, and to evaluate
the catecholase activity of free cupric ions.
2.6. Superoxide dismutase activity assay
A kinetic treatment of the experimental data revealed a satura-
tion Michaelis-type mechanism for the reaction and then the
Michaelis–Menten approach was applied using a commercially
available software.
The SOD-mimic activity was determined by an indirect method
adapted from Starha et al. [14]. This method is based on the
competitive reaction of the tested compounds (1, 2, Fenoprofen
calcium salt and CuCl2) and XTT dye [2,3-bis(2-methoxy-4-nitro-
5-sulfofenyl)-2H-tetrazolium-5-carboxanilide natrium salt] (Al-
drich) with a saturated DMF solution of potassium superoxide,
KO2 (Aldrich). The interaction of XTT dye with superoxide anion
radical led to the formation of the XTT-formazane. Its concentra-
tion was determined by spectroscopic measurements at 480 nm.
The tested complexes, which acted as the scavengers for superox-
ide anion radicals, decreased the absorbance at 480 nm.
3. Results and discussion
The reaction of cupric chloride and calcium Fenoprofenate
yielded the dinuclear copper(II) Fenoprofenate in quantitative
amount [22]. Its reaction with a 1:4 and 1:2 molar proportion of
imidazole and caffeine bases, respectively, produces the mononu-
clear complex Cu(fen)2(im)2 (1) and the dinuclear species Cu2(-
fen)4(caf)2 (2). Both substances were soluble in acetone, DMF and
DMSO, scarcely soluble in methanol and ethanol, and insoluble in
acetonitrile and water. Selected crystals of complex 1 were struc-
turally studied by X-ray diffraction techniques (Scheme 2).
The required amounts of 1.00 ꢃ 10ꢁ3 mol Lꢁ1 DMF solutions of
the tested compounds were added to 1.0 ꢃ 10ꢁ2 M potassium
phosphate buffer (pH 7.4) to provide 0.125, 0.250, 0.500, 0.750,
1.00, 2.50, 5.00, 7.50 and 10.0 ꢃ 10ꢁ6 mol Lꢁ1 solutions. For the
cases of CuCl2 and Fenoprofen calcium salt the final copper(II) con-
centration as well as the concentration of Fenoprofenate anions
were the same than the acquired for complexes solutions in order
3.1. Description of the Cu(fen)2(im)2 (1) structure based upon X-ray
to compare results. Subsequently, 500
were added. The resulting solution was mixed thoroughly. The
reaction was started by the addition of 500 L of a saturated KO2
l
L of XTT dissolved in buffer
crystallography
l
The crystal data with structure refinements are given in Table 1.
The molecular structure of the complex Cu(fen)2(im)2 is shown in
Fig 1.
solution in DMF. In all cases, the final volume was 3.00 mL and five
samples of each concentration level of all the copper(II) complexes
were tested (n = 5). After 30 min of incubation at room tempera-
ture the absorbance at 480 nm was measured against a blank sam-
ple prepared without the XTT dye. The same procedure was used to
prepare the control sample without the tested copper(II) com-
plexes and the absorbance was measured against a solution con-
taining only XTT dye. The percentages of inhibition (% INH) were
The structure consists of discrete centrosymmetric mononu-
clear units in which the Cu(II) ions acquire distorted octahedral
environments. The almost trans-square plane coordination envi-
ronment in the equatorial plane is built by two closest oxygen
atoms (O(1)) and (O(1a)) atoms of different Fenoprofenate anions
and two nitrogen atoms (N(1)) of the neutral imidazole molecules.
The apical coordination sites of the copper centers are occupied by
other two weakly coordinating atoms (O(2)) and (O(2a)) of the car-
boxylate anions located at a greater distance. Selected bond dis-
tances and angles of the crystal structure are shown in Table 2.
The asymmetric chelate type coordination of Fenoprofenate anions
determined according to the formula (1 ꢁ Xsample/Xcon-
1/2
trol) ꢀ 100 (S2sample + S2
)
ꢀ 100% where X is the media of
control
the absorbance and S its standard deviation. The SOD-mimic activ-
ity was expressed as the IC50 value, calculated from the concentra-
tion-dependent curve of the inhibition of the absorbance at
480 nm at eight concentration levels. The IC50 values represent
the concentrations of the tested copper(II) complexes reducing
the XTT-formazane formation to 50%.
(
j2-O,O0) forces a small angle (c.a. 53°) between the square CuO2N2
plane and the O(2)-Cu-O(2) line. The coordination sphere of the
metallic center is similar to that observed for the other few struc-
turally resolved examples of mononuclear bis-imidazole adducts of
copper(II) carboxylates [24–27]. Comparing with the other Cu(II)-
AINE complex reported, the trans-Cu(salicylate)2(im)2 complex
[25], the Cu–O distances are longer (2.042 Å) and the Cu–N bond
lengths a little shorter (1.961 Å) than in the Fenoprofenate com-
plex 1. The Cu–O distances of the weakly coordinated carboxylate
oxygen atoms in the axial positions for the salicylate adduct
(2.87 Å) are also longer than those of the Fenoprofenate complex
1, suggesting a closer carboxylate–Cu(II) interaction in the later.
The observed distances and angles of the Fenoprofenate anions
are very similar (differences in the order of 0.03 Å and 3°, respec-
tively) to those reported for the dinuclear Cu2(fen)4(dmf)2 complex
[22]. Between the crystal packing forces there is only one intermo-
lecular hydrogen bridge between a imidazole proton (N(2)–H(2))
and a Fenoprofenate oxygen atom (O(2)b) with a total distance
d(N2–H2ꢀꢀꢀO2b) = 2.752(2) Å.
2.7. Catecholase mimetic activity assay
Kinetic experiments concerning the catechol oxidase activity of
complexes 1 and 2 related with the oxidation of 3,5-di-tert-butyl-
catechol (3,5-dtbc) (Aldrich) were monitored spectrophotometri-
cally by following the increase of the 3,5-di-tert-butyl-o-
benzoquinone (3,5-dtbq) characteristic absorption band at
400 nm. Data of thermostated solutions (25.0 0.5 °C) were col-
lected over 10 min. The experiment was carried out by the addition
of 1.50 mL of a freshly prepared (3,5-dtbc) solution in methanol
(0.25, 0.50, 1.0, 2.0, 4.0, 6.0, 8.0, 10.0 ꢃ 10ꢁ3 mol Lꢁ1) to a 1-cm-
path-lengh cell containing 1.50 mL of a methanol solution of the
copper complexes in order to obtain a final copper(II) concentra-
tion of 5.00 ꢃ 10ꢁ5 mol Lꢁ1. During the kinetic run, the reaction cell
was opened to the air to allow the solution to be continually equil-

M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
77
DMF/H2O
CuCl2 + Ca(fen)2
Cu2(fen)4(dmf)2
H
N
acetone/CH3CN
Cu2(fen)4(dmf)2
Cu2(fen)4(dmf)2
+
Cu(fen)2(im)2 (1)
N
O
CH3
N
H3C
acetone/H2O
N
+
Cu2(fen)4(caf)2 (2)
N
O
N
CH3
Scheme 2. Synthetic procedure used with copper(II) Fenoprofenates.
Table 1
The EPR spectra for the microcrystalline form of the dinuclear
Cu(II) complex 2 (Fig 2B), recorded at room temperature, resemble
those of previous reported Cu(II) Fenoprofenates [23,31] and other
tetracarboxylate-bridged dinuclear Cu(II) complexes with well
known paddle wheel ‘‘Cu(II)2–(RCOO)4’’ cages [32–34]. Assuming
Crystal data and structure refinement parameters for complex 1.
Empirical formula
Formula weight
T (K)
C36H34CuN4O6
682.22
100(2)
0.71073
Monoclinic
P21/c
k (Å)
that the zero field splitting (D) is significantly greater than hm,
Crystal system
Space group
Unit cell dimensions
a (Å)
E ffi 0 (axial symmetry), a singlet ground state (S = 0) and a ther-
mally populated triplet state (S = 1), we expect to observe three
10.1183(11)
12.881(2)
absorptions of the six
DM = 1 resonance fields [32]. Data obtained
b (Å)
for complex 2 are: Hz1 = 370, H\ = 4810, and Hz2 = 5990 G, which
are in good accordance to previous reports in similar Cu(II) dinu-
clear complexes [23,31,35]. The quality of the spectra does not al-
low a precise determination of the magnetic parameters. g-Values
(g\ = 2.11, gz = 2.48, and gav = 2.23) were calculated taking into ac-
count the simplified model for moderate interacting dinuclear
Cu(II) centers [32,33]. The calculated magnetic parameters are in
agreement with the previous reports for dinuclear copper Feno-
profenates (Table 3) and other dicopper(II)–tetracarboxylate com-
plexes [23,31,34,36–38].
c (Å)
12.3477(17)
90
94.943(4)
90
1603.3(4)
2
1.413
0.735
a
(°)
b (°)
c
(°)
V (Å3)
Z
Density (calculated) (Mg/m3)
l
(mmꢁ1
)
F(000)
710
Crystal size (mm3)
h (°)
0.23 ꢃ 0.18 ꢃ 0.18
2.96–30.14
Index ranges
ꢁ14 6 h 6 14, ꢁ18 6 k 6 14,
ꢁ17 6 l 6 16
17683
4708 [Rint = 0.0365]
99.3
Complexes 1 and 2 present two characteristic IR absorption
bands (1578/1391 and 1602/1409 cmꢁ1, respectively for 1 and 2)
Reflections collected
Independent reflections
Completeness of theta = 30.14°
(%)
Refinement method
Data/restrains/parameters
Goodness-of-fit (GOF) on F2
corresponding to the antisymmetric,
mas(COO), and symmetric,
ms(COO), stretching vibrations of the carboxylate groups of the
Fenoprofenate anions. Those bands are shifted against Ca(-
fen)2ꢀH2O (1560/1420 cmꢁ1) [23] indicating the coordination of
the carboxylate ligands to the copper(II) centers. The separation
Full-matrix least-squares on F2
4708/3/229
1.107
R1 = 0.0430, wR2 = 0.1004
R1 = 0.0607, wR2 = 0.1091
0.770 and ꢁ0.635
between mas(COO) and m
s(COO) bands (187 cmꢁ1) in complex 1 is
Final R indices [I > 2
R indices (all data)
r(I)]
consistent with carboxylate groups acting as unsymmetrical
bidentate ligands and comparable to those reported for mononu-
clear copper(II)–carboxylate–imidazole complexes having essen-
tially the CuN2O2 chromophore [34,39]. The great separation of
the carboxylate bands for complex 2 (193 cmꢁ1) is similar to those
previously reported for other Cu(II)-Fenoprofenates (Table 4) and
other binuclear copper(II) complexes in which the carboxylate ad-
ducts act as bridging bidentate ligands [40,41]. The coordination of
imidazole molecules to copper(II) in complex 1 is evidenced by the
presence of some characteristic bands in the vibrational spectra
Largest ꢁd3ifference peak and hole
(e Å
)
3.2. Description of the Cu(fen)2(im)2 (1) and Cu2(fen)4(caf)2 (2)
structures based upon the spectroscopic analysis
The EPR spectra of microcrystalline samples of complexes Cu(-
fen)2(im)2 (1) and Cu2(fen)4(caf)2 (2) confirm the stabilization of
Cu(II) centers in both substances and their mononuclear and li-
gand-bridged dinuclear molecular structures, respectively. The
room temperature EPR spectrum of complex 1 shown in Fig 2A
was analyzed using the freeware WinSim 2002 program [28]. The
simulation could be achieved with three nearly equal g-compo-
nents g1 = 2.112, g2 = 2.107 and g3 = 2.109 (gav = 2.11), which is
consistent with a pseudo-isotropic system (see Supplementary
information S1). The X-ray structural determination indicates the
presence of tetragonally elongated environments for the non-iso-
lated Cu(II) centers (Fig 1) and an expected dx2–y2 ground state
with g-values in the order: g|| ꢄ g\ > 2.0 [29,30]. The presence of
two 90° misaligned chromophores (50:50%) of the single copper(II)
centers in the crystalline solid (see Supplementary information S2)
produces the pseudo-isotropic behavior of the substance [27].
such as that at 3138 (
752 (
(CH)) cmꢁ1 shifted with respect to the free imidazole mol-
ecule [42]. The free caffeine molecule presents strong absorption
bands at 1699 ( (CN)), 1660 ( as(CO)) and 1542 (
s(CO)) cmꢁ1
m(NH)), 1330 (m(CN)), 1073 (d(CNC)) and
c
m
m
m
.
These bands appear in the IR spectrum of complex 2, shifted to
higher wavenumbers, 1708, 1666 and 1582 cmꢁ1, respectively,
indicating the coordination of the caffeine molecules to copper(II)
[43]. A stretching vibration (m(CC)) of the heterocyclic ring of caf-
feine, appearing at 1590 cmꢁ1 in the free ligand, shifts to a higher
Raman signal (1597 cmꢁ1) on complexation to the metal atom
[44].
The electronic spectra for the mononuclear complex 1 as Nujol
mull and in solution (Table 4) exhibit one weak, broad and asym-

78
M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
long wavelength transitions [47]. In Nujol mulls and in solutions
in acetone and DMSO another very weak band is observable
around 370 nm. This band corresponds to the Cu–OCO–Cu bridging
linkage in dinuclear copper(II) complexes [38,45] and its charge–
transfer character has been confirmed by theoretical calculations
[46]. The presence of this absorption in the spectra of complex 2
clearly indicates that paddle-wheel units of the solid complex are
still present in solution. Remarkably, this band is not present in
the spectra of methanolic solutions, indicating the rupture of the
paddle wheel ‘‘Cu(II)2–(RCOO)4’’ cages into mononuclear species,
maybe to Cu(fen)2(caf)(MeOH) molecules. This behavior has been
observed in methanol for other dinuclear Cu(II)–carboxylato com-
plexes [46,48]. Following the criteria of Prenesti et al. [49] the
highest energy location of the visible band of complex 1 compared
to complex 2 assures the former still forms Cu(fen)2(im)2 molecu-
lar units in methanolic solutions.
3.3. Anti-inflammatory properties
The results obtained in carrageenan induced paw oedema in
mice are presented in Table 5. Fenoprofen calcium salt presented
very significant differences with respect to the control at the fifth
hour from the beginning of the assay and significant differences
at the third and at the seventh hour. The mononuclear Cu(fe-
n)2(im)2 complex (1) presented a significant difference in reduc-
tion of oedema at the third hour of the experiment; meanwhile it
presented very significant differences at the fifth and seventh
hours when compared to the control group. These results indicate
that, when comparing complex 1 and the uncomplexed parent
drug Fenoprofen, even both present anti-inflammatory properties,
the complex shows a more sustained activity in time. As can be
seen in Fig 3, the inflammation inhibition percent approached for
complex 1 was significantly higher than that for Fenoprofen cal-
cium salt at the seventh and at the ninth hour of the assay
(51.3% and 32.0% for complex 1 versus 19.9% and 10.5% for Feno-
profen calcium salt, respectively), indicating a higher capacity of
the mononuclear complex to maintain the anti-inflammatory
activity until the end of the assay.
When analyzing the results obtained for the dinuclear Cu2(-
fen)4(caf)2 complex (2) the pattern for the inhibition of inflamma-
tion differs from the observed for the mononuclear complex 1.
When compared to the control, the dinuclear copper(II) complex
presented very significant differences until the seventh hour of
the carrageenan induced paw oedema. The percent of inhibition
of inflammation was higher for this compound than Fenoprofen
calcium salt and complex 1 until the seventh hour.
When comparing the complexes under study each other, it can
be seen from Fig 3 that the mononuclear copper(II) complex con-
taining Fenoprofenate anions and imidazole, presents a more sus-
tained anti-inflammatory activity in time, but on the other hand,
complex 2 containing caffeine is more effective as anti-inflamma-
tory agent at the first time of the experiment with excellent per-
centages of inhibition until the fifth hour.
It is also important the comparison of the anti-inflammatory
activity of the complexes here studied with the one for other Cu(II)
complexes with AINEs. In the study carried out by Dillon et al. [3] it
was evaluated the anti-inflammatory activity of the uncomplexed
parent drug Indomethacin and the dinuclear copper(II) complex
Cu2(Indo)4(dmf)2. It was found that the most significant results
were obtained at the third hour of the assay, so they only showed
the results for that time, finding that the copper(II) complex pre-
sents higher anti-inflammatory properties than Indomethacin.
Taking into account these results it is remarkable to mention that
we have found significant results until the ninth hour from the
inflammation induction for complex 1 and until the seventh hour
for complex 2, indicating a more sustained anti-inflammatory
Fig. 1. ORTEP view of the copper complex 1 with 50% probability of thermal
ellipsoids.
Table 2
Selected bond lengths (Å) and angles (°) for [Cu(fen)2(im)2] (1).
Bond lengths (Å)
Bond angles (°)
Cu1–O1
Cu1–O1a
Cu1–O2
Cu1–O2a
Cu1–N1
Cu(1)–N(1)a
O1–C1
1.9668(13)
1.9668(13)
2.733(3)
2.733(3)
1.9815(15)
1.9815(15)
1.273(2)
1.237(3)
1.542(3)
1.273(2)
1.237(3)
1.542(3)
O1a–Cu1–O1
O1a–Cu1–N1
O1–Cu1–N1
O1a–Cu1–N1a
O1–Cu1–N1a
N1–Cu1–N1a
O2–C1–O1
O2–C1–C2A
O2–Cu1–N1
O1–Cu1–O2
O1a–C1a–C2Aa
180.00(10)
90.48(6)
89.52(6)
89.52(6)
90.48(6)
180.00(10)
123.84(18)
114.86(19)
90.48(6)
53.25(5)
121.26(19)
O2–C1
C1–C2a
O2a–C1a
O1a–C1a
C1a–C2Aa
Hydrogen bonds
D–HꢀꢀꢀA
d(D–H)
0.88
d(HꢀꢀꢀA)
d(DꢀꢀꢀA)
N(2)–H(2)ꢀꢀꢀO(2)
1.87
2.752(2)
metric band around 660 nm and very intense ultraviolet bands.
The first band is identified with d-d transitions of the Cu(II) center
[38]. This band is due to the xz, yz ? x2 ꢁ y2 transitions with a
shoulder at lower energies originated in the z2 ? x2 ꢁ y2 transition.
It appears at the expected position for mononuclear copper(II) car-
boxylate adducts containing two imidazole molecules [34]. The
ultraviolet absorptions correspond to
p ?
p⁄ transitions of the aro-
matic rings of imidazole and Fenoprofenate moieties.
Complex 2 electronic spectra correspond to those reported for
dinuclear Cu(II)–carboxylate complexes [45,46], showing a weak
asymmetric band around 690 nm and intense unresolved ultravio-
let absorptions. The first, as in complex 1, is usually assigned to the
d ? d transitions of tetragonally distorted Cu(II) centers. New
experimental and theoretical data of the model complex Cu2(CH3-
COO)4ꢀ2H2O recently reported propose that not only copper(II)
orbitals but also carboxylate anions orbitals are involved in the

M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
79
A
B
100
300
500
700
250
310
370
430
Magnetic Field (mT)
Magnetic Field (mT)
Fig. 2. 298 K X-band (9.75 MHz) spectra of microcrystalline complexes 1 (A) and 2 (B). For A: central field: 335 mT, sweep width: 80 mT; microwave power: 0.6539 mW. For
B: central field: 400 mT, sweep width: 800 mT; microwave power: 43 mW.
Table 3
lyzed. Both substances were tested for 4 h employing the same
technique. The increase in percentage inhibition of paw oedema
for Diflunisal copper complex as compared to the drug was 6%,
9%, 11%, 12% and 11% at 30 min, 1, 2, 3 and 4 h after the carra-
geenan injection, respectively. For complex 1, the increase in per-
centage inhibition of paw oedema with respect to Fenoprofen
calcium salt was 32% and 22% at the seventh and ninth hour from
the carrageenan injection and 13%, 6% and 19% at third, fifth and
seventh hour of the assay for complex 2. These differences indicate
that both copper(II) Fenoprofenates here studied are very good and
promising anti-inflammatory agents when compared to other cop-
per(II) complexes with NSAIDs not only for the activity purchased
but also for their capacity to sustain the effect in time.
X-band EPR dataa of complex 2 and other dinuclear copper(II) Fenoprofenates.
Compounds
Hz1/G
H\/G
Hz2/G
g\
gz
Refs.
[Cu2(fen)4(dmf)2]
502
629
557
370
4590
4740
4710
4810
6080
6210
6080
5990
2.14
2.11
2.20
2.14
2.49
2.48
2.52
2.49
[23]
[31]
[31]
T.w.
[Cu2(fen)4(4,40-bipy)]n
[Cu2(fen)4(2,5-Me2pyz)]n
Cu2(fen)4(caf)2
a
At room temperature.
activity in time for both complexes. In another study reported by
Razi et al. [50] the anti-inflammatory properties of the NSAID
Diflunisal and the corresponding copper(II) complex were ana-
Table 4
Selected FTIR (cmꢁ1) and UV–Vis data [k(nm)/Molar absorptivity
e
(L molꢁ1 cmꢁ1)] of complexes 1 and 2 and other dinuclear copper(II) Fenoprofenates.
Compound
Infrared bands
as(COO)
UV–Vis bands
Nujol
m
ms(COO)
DMSO
Acetone
Methanol
Assig.a
[Cu(fen)2(im)2]
Cu2(fen)4(caf)2
1578
1602
1391
1409
660a
680
696/124
716/404
375/350
658/156
683/423
370/280
686/85
696/136
d–d
d–d
CTBb
d–d
[Cu2(fen)4(dmf)2] [21]
[Cu2(fen)4(pyz)]n [30]
[Cu2(fen)4(4,40-bipy)]n [30]
1612
1618
1626
1408
1409
1406
699
699
699
694/140
CTBb
d–d
704/419
385/210
705/491
380/245
CTBb
d–d
CTBb
a
See text for assignments.
CTB: charge transfer band.
b
Table 5
Anti-inflammatory activity in carrageenan-induced paw oedema in mice.
Groups
D
mm (mean SEM)a
3 h
5 h
7 h
9 h
Control
Fenoprofen calcium salt
Indomethacin
1.44 0.09
1.02 0.14*
1.28 0.08
1.70 0.08
0.98 0.15**
1.46 0.13
1.71 0.09
1.37 0.09*
1.34 0.10*
1.52 0.11
1.36 0.20
1.41 0.09
Complex 1
Complex 2
0.97 0.17*
0.83 0.13**
1.02 0.15**
0.88 0.13**
0.83 0.14**,#
1.05 0.17**
1.03 0.20
1.42 0.16
a
Mean values of the difference of the length of the hind paw with respect to the beginning of the assay.
*
p < 0.05 in comparison to control.
p < 0.01 in comparison to control.
p < 0.05 with respect to Fenoprofen calcium salt.
**
#

80
M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
Table 6
SOD mimic activity of the complexes under study and other Cu(II) complexes with
NSAIDs.
Compound
IC50
(
lM)
Refs.
Cu(fen)2(im)2 (1)a
0.70
0.24
0.41
0.23
0.65
0.74
T.w.
T.w.
[31]
[3]
[51]
[6]
Cu2(fen)4(caf)2 (2)a
a
Cu2(fen)4(dmf)2
b,c
Cu2(indo)4(dmf)2
b,d
Cu(sal)2(1,2-meim)2
b,e
Cu(sal)2(bzdh)2
Native Cu,Zn-SODb
Native Cu,Zn-SODa
0.0042
0.480
[52]
[14]
a
b
c
Compound evaluated by the XTT method.
Compound evaluated by the xantine–xantine oxidase/INT method.
Indo: indomethacinate.
sal: salicylate; meim: methylimidazole.
bzdh: benzimidazole.
d
e
complex or enzyme required to reduce 50% of the superoxide rad-
ical anion (O2ꢀꢁ). Fig 4 shows the percent inhibition of the forma-
tion of XTT-formazane against the increasing concentration of
the complexes under study. Table 6 presents the IC50 values of
complexes 1, 2 and other mononuclear and dinuclear complexes
evaluated by different techniques. Although sometimes the activity
of the complexes is not preserved across the different assays [7] we
still found that in this case the IC50 values obtained for the inves-
tigated complexes employing the XTT method are comparable to
that obtained with other methods. Both complexes investigated
in this work presented significant SOD-mimic activity.
Fig. 3. Inflammation inhibition percent approached by the complexes under study
and Fenoprofen calcium salt in carrageenan induced paw oedema in mice.
The SOD-mimic activity of imidazole containing complex 1 is
comparable to other complexes of Cu(II) with neutral imidazole
derivatives and other nitrogenated ligands. In this mononuclear
compound, as well as proposed for other Cu(II)–NꢀSꢁAIDs complexes
with the same structure, the site on Cu(II) for O2 binding would
be provided by the dissociation of one of the oxygen atoms from
the carboxylate anions or by the imidazole nitrogen atom. This dis-
sociation would facilitate the necessary geometrical changes to
permit the reaction course. The dinuclear copper complex Cu2(-
fen)4(caf)2 presents a SOD mimic activity comparable to the dinu-
clear copper(II) complex of Indomethacin with dmf as adduct,
which is considered to be an excellent SOD mimic compound [3].
In a previous work we have evaluated the SOD mimic activity of
the complex Cu2(fen)4(dmf)2 with an IC50 value of 0.411 lM [31].
The replacement of the axial dmf ligands by caffeine molecules in-
creases the activity for complex 2. The mechanism for scavenging
O2ꢀꢁ would consist, in this case, in the dissociation of one Fenoprof-
enate oxygen atom in the equatorial plane of a copper(II) ion. This
site would be occupied by superoxide and the nitrogen atom cor-
responding to the caffeine molecule would provide a more favor-
able environment for the reaction in comparison to the oxygen
atom of the dmf ligand in Cu2(fen)4(dmf)2. Remarkably, complex
2 presents higher SOD mimic activity than the native enzyme eval-
uated by the same method (Table 6).
3.5. Catechol oxidase mimetic activity
The ability of the complexes under study to oxidize catechols
was tested following the procedure previously reported [23]. In a
kinetic run a 2.50 ꢃ 10ꢁ3 mol Lꢁ1 methanolic solution of 3,5-DTBC
was reacted with a 5.00 ꢃ 10ꢁ5 mol Lꢁ1 methanolic solution of Cu(-
fen)2(im)2 (1) (a concentration of 2.50 ꢃ 10ꢁ5 mol Lꢁ1 was used for
Cu2(fen)4(caf)2 (2) in order to achieve the same conditions related
to copper concentration), under aerobic conditions and the course
of the reaction was followed by UV–Vis spectroscopy (Fig 5). Both
complexes show moderate catecholase activity.
Fig. 4. Plot of percentage of inhibition of XTT-formazane against the increase in
concentration of the complexes under study.
3.4. Superoxide dismutase activity assay
The superoxide dismutase mimetic activity of complexes 1 and
2 has been determined through the XTT method and is expressed
as the IC50 value, which represents the concentration (lM) of the

M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
81
Fig. 5. Plot of the evolution of the UV–Vis spectra of the oxidation reaction of 3,5-DTBC to 3,5-DTBQ in methanol at 25.0 °C, catalyzed by the [Cu(fen)2(im)2] complex (2). The
spectra were recorded at the beginning of reaction and after 5, 10, 20, 30, 60 min and corrected with the blank of reaction.
Table 7
Michaelis–Menten and enzymatic activity parameters for the catecholase activity of
copper(II) complexes.
a
b
Compound
KM
Vmax
Enzymatic Refs.
activityc
[Cu(fen)2(im)2] (1)
[Cu2(fen)4(caf)2] (2)
[Cu2(fen)4(dmf)2]
Cu2(L1)(H2O)(NO3)2
Cu2(Stryp)2(H2O)e
1.50 0.23 6.72 0.38 1.97
0.25 0.05 4.97 0.19 1.34
0.13 0.02 5.70 0.16 1.84
T.w.
T.w.
[23]
[54]
[56]
[55]
[57]
[3]
d
2.1
173
1800
53.4
37
n.r.
n.r.
n.r.
[Cu2(BPMP)(OAc)2][ClO4].H2Of 6.4
g
[Cu2(L2O)(CF3SO3)](CFSO3)2
[Cu(ibup)2(im)2]h
[Cu(valp)2(im)2]i
2.92
24.6
n.r.
n.r.
n.r.
n.r.
n.r.
n.r.k
n.r.
n.r.
n.r.
n.r.
0.12
0.75
2.6
1.3
0.220
[58]
[58]
[4]
[Cu2(valp)4]
[Cu2(nap)4]n
j
[Cu2(ibup)4(caf)2]
n.r.
[3]
a
b
c
d
e
f
In units 10ꢁ3 mol Lꢁ1
.
In units of 10ꢁ5 mol Lꢁ1 minꢁ1
Amount of 3,5-dtbq ( mol) produced by 1.0 mg of the complex.
.
l
L1: 2-formil-4-metil-6R-Iminometil-fenolato.
STryp: triptofane.
BPMP: 2,6-bis[bis(piridin-2-yl-methylamine)methyl]-4-methylfenolate.
L2: 1,3-bis{N,N-bis(2-[2-piridyl]ethyl}-amine-2-hidroxypropane.
Ibup: ibuprofenate.
Valp: valproate.
Nap: naproxenate.
n.r.: not reported.
g
h
i
j
k
this model indicates the existence of a first complexation equilib-
rium between the DTBC substrate and the active copper(II) species
in the methanol solution. The obtained M–M parameters are
shown in Table 7. As it was pointed out previously when discussing
the UV–Vis spectra the active copper(II) species present in the
methanolic solution could be mononuclear. The KM value indicates
that complex 1 presents lower affinity in bonding to the 3,5-dtbc
molecule and then it is a better catalyst than the complexes con-
taining dmf and caffeine as adducts. In Table 7 the catalytic behav-
ior of the new complexes has been compared with bibliographic
data of active dinuclear copper(II) complexes (as far as we know
M–M parameters have not been reported yet for mononuclear spe-
cies) the new compounds show greater affinity to the binding of
3,5-dtbc (lower KM values) than dinuclear copper(II) model com-
plexes of catechol oxidase activity and, by this way, a lower cata-
lytic performance [54–57]. For other Cu(II)–NSAIDs complexes
the catecholase mimetic activity was reported by Abuhijleh and
Fig. 6. Initial reaction rate (Vo) of the oxidation of dtbc to dtbq in methanol at
25.0 0.5 °C: (A) [Cu2(fen)4(caf)2] (2.50 ꢃ 10ꢁ5 mol Lꢁ1); (B) [Cu(fen)2(im)2]
(5.00 ꢃ 10ꢁ5 mol Lꢁ1).
The 3,5-DTBQ concentration versus time plots were fitted with
potential functions in order to recover the initial reaction rates
(Vo). The dependence of Vo on the 3,5-DTBC concentrations for
the oxidation reaction catalyzed by the copper(II) complexes (Fig
6) and the correlation between Voꢁ1 and [3,5-DTBC]ꢁ1 both indi-
cate the existence of a two step Michaelis type mechanism for
the reaction [53]. A complete kinetic study was carried out on
the basis of the Michaelis–Menten (M–M) model, originally devel-
oped for enzymatic reactions. The good accordance of data with

82
M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
co-workers as the enzymatic activity [4,33,57]. The new copper
Fenoprofenates showed good catecholase mimetic activity in com-
parison to other NSAIDs complexes.
Appendix A. Supplementary data
CCDC 831490 contains the supplementary crystallographic data
for 1. These data can be obtained free of charge via http://
www.ccdc.cam.ac.uk/conts/retrieving.html, or from the Cambridge
Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ,
UK; fax: (+44) 1223-336-033; or e-mail: deposit@ccdc.cam.ac.uk.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.poly.2011.12.005.
As reported previously, when methanolic solutions of CuCl2
were tested versus the 3,5-DTBC substrate, under the same condi-
tions than the complexes under study, there was no significant cat-
echolase mimetic activity for the solvated copper(II) ions [23]. This
may be indicative that in the methanolic solutions of the com-
plexes the active species still have the Fenoprofenates acting as li-
gands. In terms of the catalytic model for mononuclear copper(II)
compounds proposed by Abuhijleh and co-workers [59–61] the
low performance of the CuCl2 solutions could be due to the absence
of a proton scavenger for the production of dtbc2ꢁ anions.
References
[1] J.R.J. Sorenson, J. Med. Chem. 19 (1976) 135.
[2] J.R.J. Sorenson, Prog. Med. Chem. 26 (1989) 437.
[3] C.T. Dillon, T.W. Hambley, B.J. Kennedy, P.A. Lay, O. Zhou, N.M. Davies, J.R.
Biffin, H.L. Regtop, Chem. Res. Toxicol. 16 (2003) 28.
4. Conclusions
[4] A.L. Abuhijleh, J. Khalaf, Eur. J. Med. Chem. 45 (2010) 3811.
[5] T. Suksrichavalit, S. Prachayaittikul, C. Nantasenamat, C. Isarankura-Na-
Ayudhya, V. Prachayasittikul, Eur. J. Med. Chem. 44 (2009) 3259.
[6] M. Devereux, D. OShea, M. OConner, H. Grehan, G. Conner, M. McCann, G.
Rosair, F. Lyng, A. Kellet, M. Walsh, D. Egan, B. Thati, Polyhedron 26 (2007)
4073.
[7] D. Kovala-Demertzi, A. Galani, M. Demertzis, S. Skoulika, C. Kotoglou, J. Inorg.
Biochem. 98 (2004) 358.
[8] B. Viossat, F.T. Greenaway, G. Morgant, J.-C. Daran, N.-H. Dung, J.R.J. Sorenson, J.
Inorg. Biochem. 99 (2005) 355.
[9] F. Dimiza, F. Perdih, V. Tangoulis, I. Turel, D.P. Fessissoglou, G. Psomas, J. Inorg.
Biochem. 105 (2011) 476.
[10] K. Ghosh, P. Kumar, N. Tyagi, U.P. Singh, V. Aggarwal, Eur. J. Med. Chem. 45
(2010) 3770.
[11] A.M. Bonin, J.A. Yañez, C. Fukuda, X.W. Teng, C.T. Dillon, T.W. Hambley, P.A.
Lay, N.M. Davies, Cancer Chemother. Pharmacol. 66 (2010) 755.
[12] Diamond (version 3.2g), Cystal Impact, K. Brandenburg & M. Berndt GbR, Bonn,
Germany, 2011.
[13] C.A. Winter, E.A. Risley, G. Nuss, Proc. Soc. Exp. Bio. Med. 111 (1962) 544.
[14] P. Starha, Z. Travnicek, R. Herchel, I. Popa, P. Suchy, J. Vanco, J. Inorg. Biochem.
103 (2009) 432.
[15] J.E. Weder, C.T. Dillon, T.W. Hambley, B.J. Kennedy, P.A. Lay, J.R. Biffin, H.L.
Regtop, N.M. Davies, Coord. Chem. Rev. 232 (2002) 95.
[16] T.S. Oga, S.F. Taniguchi, R. Najjar, A.R. Souza, J. Inorg. Biochem. 41 (1991) 45.
[17] J.D. Scandalis (Ed.), Oxidative Stress and the Molecular Biology of Antioxidant
Defenses, Cold Spring Harbor Laboratory Press, Releigh, 1997.
[18] S.M. Cardoso, C. Pereira, C.R. Oliveira, Biochem. Biophys. Res. Commun. 246
(1998) 703.
[19] A.M. Ramadan, M.M. EI-Naggar, J. Inorg. Biochem. 63 (1996) 143.
[20] I.A. Koval, P. Gamez, C. Belle, K. Selmeczi, J. Reedijk, Chem. Soc. Rev. 35 (2006)
814.
[21] R. Nickander, W. Marshall, J.L. Emmerson, G.C. Todd, in: M.E. Goldberg (Ed.),
Pharmacological and Biochemical Properties of Drug Substances, Am. Pharm.
Assoc., Washington, 1977, pp. 183–213.
[22] M.A. Agotegaray, O.V. Quinzani, R. Faccio, C. Goyenola, A.W. Mombrú, Acta
Crystallogr., Sect. E 64 (2008) m1612.
Mixed-ligand ternary copper(II) complexes with the anti-
inflammatory drug Fenoprofen as primary ligand and the biologi-
cally relevant imidazole and caffeine molecules as auxiliary ligands
were synthesized and characterized by spectral methods. The crys-
tal structure of complex 1 was determined by X-ray diffraction
techniques. Complex 1 is built of mononuclear [Cu(fen)2(im)2]
units in which the copper centers have rhombically distorted octa-
hedral N2O2 + O2 coordination spheres. The dinuclear structure of
complex 2 was elucidated by EPR spectral analysis. The molecular
complex 2 is built of typical paddle-wheel binuclear ‘‘Cu2(fen)4’’
units with two caffeine molecules at the axial positions of the
Cu(II) centers. The carrageenan induced paw oedema in mice
experiments show that both complexes 1 and 2 present enhanced
and lasting anti-inflammatory properties against the parent drug
calcium Fenoprofenate, Ca(fen)2ꢀ2H2O. The imidazole containing
complex 1 has a similar performance than the caffeine containing
complex 2 in the first phase of the experiments (3–5 h) but greater
significant effects at long time exposure (7–9 h). By this way this
complex could be considered as a potential therapeutic agent in
the treatment of inflammatory diseases. Complex 1 has similar
but lower anti-inflammatory properties than the dinuclear com-
plex [Cu2(fen)4(dmf)2] [23] showing that there is not a clear corre-
lation between structure–anti-inflammatory effects due to the
unknown differences in the pharmacodinamic properties of these
copper(II) complexes. Both complexes show potent superoxide dis-
mutase (SOD) mimetic activities determined through the XTT
method. The measured SOD mimetic activities of the complexes
´
´
indicated a higher activity for complex 2 (IC50 of 0.24
complex 1 (IC50 of 0.70 M), and also than the native enzyme eval-
uated by the same method (IC50 of 0.480 M). The measured SOD
lM) than
[23] M.A. Agotegaray, M.A. Boeris, O.V. Quinzani, J. Braz. Chem. Soc. 21 (2010) 2294.
[24] R. Carballo, B. Covelo, E.M. Vazquez-López, E. García-Martínez, A. Castiñeiras,
C. Janiak, Z. Anorg. Allg. Chem. 631 (2005) 2006.
l
l
[25] A.L. Abuhijleh, C. Woods, Inorg. Chem. Commun. 4 (2001) 119.
[26] Y.-Y. Wang, Q. Shi, Q.-Z. Shi, Y.-C. Gaio, Z.-Y. Zhou, Polyhedron 18 (1999) 2009.
[27] H.A. Henriksson, Acta Crystallogr., Sect. B 33 (1977) 1947.
[28] D.R. Duling, J. Magn. Reson., Ser. B 104 (1994) 105.
[29] B.J. Hathaway, A new look at the stereochemistry and electronic properties of
complexes of the copper(II) ion, in: J. Emsley (Ed.), Complex Chemistry,
Structure and Bonding, vol. 57, Springer-Verlag, 1984, pp. 55–118.
[30] B. Kozlevcar, P. Segedin, Croat. Chem. Acta 81 (2008) 369.
[31] M.A. Agotegaray, M.A. Boeris, A. Diez, M.R. Prat, O.V. Quinzani, Dinuclear
copper(II) complexes of Fenoprofen with nitrogenated ligands: synthesis,
spectroscopic, thermal, and SOD-mimic activity studies, J. Coord. Chem. 64
(2011) 2246.
mimetic activity of both complexes 1 and 2 are in the range of
other copper(II)–NSAID compounds studied through the same
experimental procedures [6,30,49]. The SOD activity presented by
the complexes here studied permits its consideration as therapeu-
tic agents because of their anti-oxidant activity and the low molec-
ular weight in comparison to the native enzyme. The catecholase
activities of the complexes toward the aerobic oxidation of 3,5-
di-tert-butylcatechol (dtbc) onto 3,5-di-tert-butylquinone (dtbq)
showed that both complexes have moderate catalytic oxidase
activities.
[32] P.M. Selvakumar, E. Suresh, P.S. Subramanian, Inorg. Chim. Acta 361 (2008)
1503.
[33] C.J. Williams, H. Morris, J. Svorec, M. Valkova, M. Valko, J. Moncol, M. Manzur,
F. Valach, M. Melnik, J. Mol. Struct. 659 (2003) 53.
[34] A.L. Abuhijleh, J. Inorg. Biochem. 55 (1994) 255.
Acknowledgements
[35] J. Boonmak, S. Youngme, N. Chaichit, G.A. van Albada, J. Reedijk, Cryst. Growth
Des. 9 (2009) 3318.
[36] Y. Muto, M. Nakashima, T. Tokii, I. Suzuki, S. Ohba, O.W. Steward, M. Kato, Bull.
Chem. Soc. Jpn. 75 (2002) 511.
[37] A.L. Abuhijleh, J. Inorg. Biochem. 66 (1997) 167.
[38] J.L. Meier, C.E. Coughenor, J.A. Carlisle, G.O. Carlisle, Inorg. Chim. Acta 106
(1985) 159.
M.A.A., M.D. and O.V.Q. thank SGCyT-UNS for financial support
of Project M24/Q025. The authors would like to thank to Dr. Jorge
A. Güida (CEQUINOR, Facultad de Ciencias Exactas, Universidad
Nacional de La Plata, La Plata, Argentina) for register the Raman
spectra of the substances under study. M.A.A. is fellow of CONICET
(Consejo Nacional de Investigaciones Científicas y Técnicas).
[39] A.L. Abuhijleh, C. Woods, E. Bogas, G. LeGuenniou, Inorg. Chim. Acta 195 (1992)
67.

M.A. Agotegaray et al. / Polyhedron 34 (2012) 74–83
83
[40] J.E. Weder, T.W. Hambley, B.J. Kennedy, P.A. Lay, D. MacLachlan, R. Bramley,
C.D. Delfs, K.S. Murray, B. Moubaraki, B. Warwick, J.R. Biffin, H.L. Regtop, Inorg.
Chem. 38 (1999) 1736.
[51] A.L. Abuhijleh, J. Mol. Struct. 980 (2010) 201.
[52] Y.H. Zhou, H. Fu, W.X. Zhao, W.L. Chen, C.Y. Su, H. Sun, L.N. Ji, Z.W. Mao, Inorg.
Chem. 46 (2007) 734.
[41] C.D. Samara, D.P. Kessissoglou, G.E. Manoussakis, D. Mentzafos, A. Terzis, J.
Chem. Soc., Dalton Trans. (1990) 959.
[53] R. Schmid, V.N. Sapunov, Non Formal Kinetics, Verlag Chemie, Weinheim,
1982. pp. 39–44.
[42] W. Tatara, M.J. Wojcik, J. Lindgren, M. Probst, J. Phys. Chem. A 107 (2003) 7827.
[43] S. Gunasekar, G. Sankari, S. Ponnusamy, Spectrochim. Acta, Part A 61 (2005)
117.
[44] M. Melnik, M. Koman, T. Glowiak, Polyhedron 17 (1998) 1767.
[45] A.B.P. Lever, Inorganic Electronic Spectroscopy, second ed., Elsevier, New York,
1984. pp. 554–572, and 636–639.
[54] K.S. Banu, T. Chattopadhyay, A. Banerjee, S. Bhattacharya, E. Suresh, M. Nethaji,
E. Zangrando, D. Das, Inorg. Chem. 47 (2008) 7083.
[55] S.J. Smith, C.J. Noble, R.C. Palmer, G.R. Hanson, G. Schenk, L.R. Gahan, M.J. Riley,
J. Biol. Inorg. Chem. 13 (2008) 499.
[56] C.-T. Yang, M. Vetrichelvan, X. Yang, B. Moubaraki, K.S. Murray, J.J. Vittal,
Dalton Trans. (2004) 113.
[46] P. Kögerler, P.A.M. Williams, B.S. Parajón-Costa, E.J. Baran, L. Lezama, T. Rojo, A.
Müller, Inorg. Chim. Acta 268 (1998) 239.
[47] M. Kyuzou, W. Mori, J. Tanaka, Inorg. Chim. Acta 363 (2010) 930.
[48] N. Kumar, A.K. Suri, Proc. Indian Natl. Sci. Acad. A 46 (1980) 565.
[49] E. Prenesti, P.G. Daniele, S. Berto, S. Toso, Polyhedron 25 (2006) 2815. and
references therein.
[57] K. Selmeczi, M. Réglier, M. Giorgi, G. Speier, Coord. Chem. Rev. 245 (2003) 191.
[58] A.L. Abuhijleh, C. Woods, Inorg. Chim. Acta 209 (1993) 187.
[59] A.L. Abuhijleh, Inorg. Chem. Commun. 14 (2011) 759.
[60] J. Kaizer, J. Mol. Catal. A: Chem. 329 (2010) 71.
[61] I.A. Koval, K. Selmeczi, C. Belle, C. Philouze, E.S. Aman, I.G. Luneau, A.M.
Schuitema, M. Van Vliet, P. Gamez, O. Roubeau, M. Luken, B. Krebs, M. Lutz, A.L.
Spek, J.L. Pierre, J. Reedijk, Chem. Eur. J. 12 (2006) 6138.
[50] M.T. Razi, M. Ahmad, M.E. Rabbani, M.A. Saeed, Int. J. Agric. Biol. 5 (2003) 1.