Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175

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Spectrochimica Acta Part A: Molecular and

Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa

Tridentate hydrazone metal complexes derived from cephalexin

and 2-hydrazinopyridine: Synthesis, characterization and antibacterial
activity
J.R. Anacona ⇑, Maria Rincones
Department of Chemistry, Universidad de Oriente, Cumana 6101 Venezuela

h i g h l i g h t s g r a p h i c a l a b s t r a c t

 A new Schiff base using cephalexin Transition coordination compounds with a tridentate hydrazone ligand (HL) derived from the condensa-
antibiotic was prepared. tion of cephalexin antibiotic with 2-hydrazinopyridine were synthesized, characterized and screened for
 Cephalexin Schiff base transition antibacterial activity.
metal complexes were prepared.
 Magnetic, spectroscopic properties NH2
NH2
and antibacterial activity were NH S NH S
studied.
H2O
O N O N
N NH O
N
HN
NH2
O OH
O OH
N

a r t i c l e i n f o a b s t r a c t

Article history: Metal(II) coordination compounds of a tridentate hydrazone ligand (HL) derived from the condensation of
Received 23 May 2014 cephalexin antibiotic with 2-hydrazinopyridine were synthesized. The hydrazone ligand and mononu-
Received in revised form 17 November 2014 clear [ML(OAc)(H2O)] (M(II) = Mn, Co, Ni, Cu, Zn, Ag) complexes were characterized by several techniques,
Accepted 5 January 2015
including elemental and thermal analysis, molar conductance and magnetic susceptibility measure-
Available online 31 January 2015
ments, electronic, FT-IR, EPR and 1H NMR spectral studies. The cephalexin 2-pyridinylhydrazone ligand
HL behaves as a monoanionic tridentate NNO chelating agent. The biological applications of complexes
Keywords:
have been studied on three bacteria strains (Escherichia coli, Acinetobacter baumannii and Enterococcus fae-
Hydrazone metal complexes
Cephalexin 2-pyridinylhydrazone ligand
calis) by agar diffusion disc method.
Synthesis hydrazone containing cephalexin Ó 2015 Elsevier B.V. All rights reserved.
Magnetic and spectral studies
Antibacterial activity

Introduction structural studies on hydrazone ligands show the importance of

hydrazone complexes in various fields, including analytical and
In view of their applicability in various fields, hydrazones, a biological fields [1,2]. Many studies have been reported regarding
member of the Schiff base family with triatomic >C@N–NH– link- the biological activities of hydrazones, including their anticancer
age, takes the forefront position in the development of coordina- [3–5], antibacterial [6,7] and antidepressant activities [8].
tion chemistry. Reports on the synthesis, characterization and The b-lactam ring, has largely been recognized as a useful build-
ing block in the synthesis of biologically important compounds.
The activity of famous antibiotic classes such as the penicillins
⇑ Corresponding author. Tel./fax: +58 (293)4002405. and cephalosporins is attributed to the presence of a b-lactam ring
E-mail address: [email protected] (J.R. Anacona).

http://dx.doi.org/10.1016/j.saa.2015.01.009
1386-1425/Ó 2015 Elsevier B.V. All rights reserved.
170 J.R. Anacona, M. Rincones / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175

[9]. Unfortunately, the wide use of the antibiotics resulted in the Synthesis of 2-hydrazinopyridine
serious medical problem of drugs resistance and public health con-
cern [10,11]. Preparation of new synthetic derivatives of antibiotics One mmol liquid 2-chloropyridine was dissolved in 2.0 mL 85%
with novel mechanism of action has become an important task to hydrazine hydrate and methanol (5 mL) and the mixture was
cope with drug resistance problems. refluxed at 90 °C for 6 h upon cooling, a yellow solid was separated
Although hydrazones have extensively been studied and widely out and recrystallized with ethanol-DMF to get the pure 2-hydraz-
employed in coordination chemistry, rather less is known about inopyridine. Yield 60%.
hydrazone ligands containing b-lactam antibiotics. Continuing
with metal-based antibiotics studies in order to establish whether
Synthesis of cephalexin 2-pyridinyl hydrazone HL
complexation affects the pharmacological properties of the ligand
and to derive additional fundamental knowledge about antibiotic
To 1 mmol of cephalexin in 25 mL of hot methanol were added
action [12–16], we report here the isolation and characterization
1 mmol of 2-hydrazinopyridine. The solution was refluxed under
of metal(II) complexes containing a hydrazone ligand derived from
nitrogen atmosphere at 70 °C for 3 h to give a dark yellow precip-
the condensation of cephalexin antibiotic, first generation cephalo-
itate. This material was filtered off and washed with methanol and
sporin, with 2-hydrazinopyridine. The chemical structure of ceph-
ether, and dried under reduced pressure. The product was purified
alexin is shown in Fig. 1.
by recrystallization from the same solvent. Yield 54%.

Experimental
Synthesis of hydrazone metal complexes
Materials and methods
Metal(II) complexes were prepared by the same general
method. To a solution of 1 mmol of the metallic acetate in 20 mL
All necessary precautions were taken to exclude oxygen and
of water was slowly added with stirring a solution of 1 mmol of
moisture during the synthesis and handling of the compounds.
HL in 10 mL of ethanol. To this solution KOH (0.1% in methanol)
Analytical grade chemicals were used as received for all experi-
was added to adjust the pH of the solution at 7–8 and the mixture
ments. Fourier transform infrared (FTIR) spectra of the ligand and
was then refluxed for 4 h and colored precipitates formed. The
its metal complexes as KBr pellets were recorded in the spectral
metal(II) complexes of HL were separated from the reaction mix-
range 4000–400 cm1 with a Perkin-Elmer Series 2000 apparatus.
ture as amorphous solids and washed several times with water,
FT–IR spectra as polyethylene pellets were recorded between 450
methanol and ether and dried under reduced pressure at room
and 120 cm1 using a Bruker IFS 66V spectrophotometer. EPR spec-
temperature. Complexes were purified by recrystallization from
tra were recorded on a Bruker ECS 106 spectrometer operating in
dimethylsulfoxide/water mixture. Yield 65–78%,
the X-band (9.76 GHz). DPPH free radical was used as the g marker.
Measurements of d–d transitions in the visible and u.v. regions
were taken with a Perkin-Elmer spectrophotometer. The contents Antibacterial activity
of C, H, N and S were analyzed on a LECO CHNS 932 model micro-
analytical instrument. To determine the metal contents, the com- The antibacterial activity of hydrazone ligand and hydrazone
pounds were decomposed by wet digestion at 350 °C with metal complexes was tested against Enterococcus faecalis as a
sulfuric acid and hydrogen peroxide and determined by normal Gram-positive bacterium and Escherichia coli and Acinetobacter
complexometric titration procedures with standard EDTA solution baumannii as Gram-negative bacteria, according to a modified Kir-
[17]. The metal contents as well as the coordinated water were also by-Bauer disc diffusion method under standard conditions using
obtained from the TGA curves. Molecular weight determinations Mueller-Hinton agar medium, as previously reported [12–15].
were carried out by the Rast camphor method. Magnetic suscepti- The test compounds in DMSO solutions (5  102 M) were added
bilities were measured on a Johnson Matthey Magnetic Suscepti- dropwise (10 lL) to a 5 mm diameter filter paper disc placed at
bility balance at room temperature using HgCo(NCS)4 as the center of each agar plate.
calibrant. 1H NMR spectra were run at 80 MHz on a Varian spec-
trometer in DMSO against tetramethylsilane (TMS) as internal ref-
erence. Thermograms were recorded on a simultaneous thermal Results and discussion
analyzer, STA-6000 (Perkin Elmer) instrument at a heating rate of
10 °C min1 up to 800 °C. X-ray powder diffraction patterns for The ligand and the metal(II) complexes were isolated pure in
the studied complexes and final solid product of thermal decompo- very good yields and they are of various colors. The ligand, manga-
sition were recorded on a HZG 4 diffractometer. Measurements nese(II) and zinc(II) complexes are yellow, cobalt(II) complex is red
were taken over the range of 2h = 2–70° using Ni filtered CuKa wine, nickel(II) and copper(II) complexes are green, and silver(II) is
radiation. orange in color. All the complexes did not melt/decompose when
heated up to 158 °C. The synthetic route of HL ligand is given in
Scheme S1. The elemental analyses of the ligand and complexes
NH2 are contained in Table 1 and they agree well with a 1:1:1:1 metal:
ligand: acetate: coordinated water stoichiometry. Thus, the general
NH S formulae [ML(OAc)(H2O)] (M(II) = Mn, Co, Ni, Cu, Zn, Ag) have been
assigned to the complexes and they are very air stable solids at
room temperature without decomposition for a long time. The
O N complexes are insoluble in water and other common organic sol-
vents such as ethanol, chloroform, benzene, acetone, dichloro-
O
methane, DMF, acetonitrile and ether but soluble in DMSO.
Attempts to form complexes of a well-defined stoichiometry,
O OH
under the above-mentioned conditions, with chromium(III), iro-
n(II) and mercury(II) ions were unsuccessful. The molar conduc-
Fig. 1. Chemical structure of cephalexin. tance values measured in DMSO at room temperature vary from
 J.R. Anacona, M. Rincones / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175 171

Table 1
Analytical, thermoanalytical (TG) and physical data.

Compound Found (Calcd.)%

C N H S H2O M1 M2 Mol wt.
[(HL)] 57.6 19.2 4.4 7.2 427.0
[(C21H21N6O3S)] (57.7) (19.2) (4.8) (7.3) (437.5)
[MnL(OAc)(H2O)] 48.7 14.9 4.9 5.4 2.8 9.5 9.7 553.2
[Mn(C23H25N6O6S)] (48.6) (14.8) (4.4) (5.6) (3.2) (9.7) (9.7) (568.5)
[CoL(OAc)(H2O)] 48.5 14.9 4.8 5.3 3.5 10.2 10.5 560.3
[Co(C23H25N6O6S)] (48.3) (14.7) (4.4) (5.6) (3.1) (10.3) (10.3) (572.5)
[NiL(Oac)(H2O)] 48.7 14.8 4.2 5.5 2.9 10.4 10.1 561.4
[Ni(C23H25N6O6S)] (48.3) (14.7) (4.4) (5.6) (3.1) (10.3) (10.3) (572.3)
[CuL(OAc)(H2O)] 47.7 14.2 4.7 5.4 3.5 10.7 11.4 564.8
[Cu(C23H25N6O6S)] (47.9) (14.6) (4.4) (5.6) (3.1) (11.0) (11.0) (577.1)
[ZnL(OAc)(H2O)] 48.1 14.7 4.1 5.8 3.2 11.2 11.0 565.3
[Zn(C23H25N6O6S)] (47.7) (14.5) (4.4) (5.5) (3.1) (11.3) (11.3) (578.9)
[AgL(OAc)(H2O) 44.8 13.2 3.8 5.4 3.0 17.1 17.7 610.7
[Ag(C23H25N6O6S)] (44.5) (13.5) (4.1) (5.2) (2.9) (17.4) (17.4) (621.4)

M1 = complexometric analysis, M2 = thermal analysis.

10.4 to 12.8 S cm2 mol1, revealing the non-electrolytic nature of ring protons was also present. All the complexes are paramagnetic
the complexes [18]. with the exception of zinc(II) complex, therefore the 1H-NMR spec-
tra of the complexes could not be obtained Comparison of the 1H-
NMR spectrum of hydrazone ligand with that of the diamagnetic
Thermal analysis
zinc(II) complex, shows the absence of the proton signals assigned
to the COOH moiety of hydrazone ligand indicating deprotonation
By thermal analysis, information on their properties, nature of
and suggests the formation of COO-metal bond. No significant
intermediate and final products of their thermal decomposition
changes in 1H chemical shifts were observed for other atoms upon
can be obtained [19]. From TGA curves, the mass loss was calcu-
complexation suggesting that, in solution, the aromatic rings are
lated for the different steps and compared with those theoretically
not involved in stacking interactions. Owing to their low solubility
calculated for the suggested formulae based on the results of ele-
it was not possible to record satisfactory 13C-NMR spectrum for the
mental analyses as well as molar conductance measurements.
diamagnetic complex.
TGA indicated the formation of metal oxide as the end product
from which the metal content could be calculated and compared
with that obtained from analytical determination. Thermograms
Infrared spectra
of the hydrated metal complexes indicate endothermic decomposi-
tions in two steps and also reveal that the complexes are stable
The IR spectrum of the complexes were recorded down to the
with no hydration water and solvent molecules. The first step in
far-IR region of 200 cm1 and tentative band assignments of some
the 158–175 °C range is assigned to loss of coordinated water mol-
characteristic bands of hydrazone ligand were made by analogy
ecules (Table 1). The final decomposition step includes complete
with other related systems [21,22]. The main infrared spectral
evaporation of the ligand as well as formation of metal oxide as
bands of the hydrazone ligand and its metal complexes are pre-
final product from which the metal content was found to be in very
sented in Table 2. The lactam m(C@O) band appear at 1735 cm1
good agreement with the data obtained from complexometric
in the spectra of cephalexin. The IR spectrum of the hydrazone
analyses. Thus, the overall thermogravimetric results are consis-
ligand shows no absorption bands which can be assigned to lactam
tent with the formulation of these complexes. The solid residues
m(C@O) vibrational mode coming from cephalexin. The absence of
obtained during thermal decomposition of complexes are suitable
such absorption together with the appearance of a new band at
metal oxides: MnO, CoO, NiO, CuO and ZnO. Their compositions
1635 cm1 attributed to m(C@N–NH) vibrations, is consistent with
have been confirmed by X-ray diffraction measurement. The dif-
the product being the expected hydrazone ligand [23,24]. The
fraction patterns of obtained residues have been compared with
infrared spectra of the metal complexes display IR absorption
reference patterns.
bands in the 1620–1625 cm1 range which can be assigned to
the C@N stretching frequencies of the coordinated ligand (HL),
1
H-NMR strongly suggesting involvement of this group in coordination.
The m(N–N) of the hydrazone complexes are found at 1110 cm1.
The values of the chemical shifts obtained were similar to those The increase in the frequency of this band in the spectra of the
of Schiff base ligands reported in the literature [20]. In the 1H-NMR complexes, due to the increase in the bond strength, again con-
spectrum of hydrazone ligand single peaks attributed to methyl, firms the coordination via the azomethine nitrogen [25].
COOH and @N–NH groups appeared at 1.95, 10.1 and 11.2 ppm Disappearance of the stretching frequency at 1690 cm1
respectively. Three groups of double peaks given by N@C–CH and assigned to m(COOH) in the ligand and appearance in the com-
N–CH on the b-lactam ring and O@C–NH– appeared at 4.90, 5.45 plexes of new masym and msym modes of the (COO) group indicates
and 9.01 ppm, respectively. One group of four resonance signals that the hydrazone has reacted. Since the carboxylate group can
consistent with an AB system attributed to S-CH2 on the dihydro- coordinate to the metal ion in either bidentate or monodentate
thiazine ring, was observed in the 3.18–3.49 ppm region with cou- fashion, the ‘‘Dm criterion’’ [Dm = masym(COO)  msym(COO)] was
pling constant 16.9 Hz for JAB. Furthermore, coupling between NH2 employed to determine the coordination mode of the carboxylate
and the adjacent CH moieties could not be distinguished and a group. The hydrazone metal complexes exhibit strong bands corre-
broad single signal due to NH2 protons was observed at sponding to masym(COO) at 1600–1605 and 1580–1585 cm1, and
5.05 ppm. A multiplet in the range 6.70–7.90 ppm due to aromatic msym(COO) at 1420–1425 and 1370–1375 cm1. The Dm value of
172 J.R. Anacona, M. Rincones / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175

Table 2
Main vibrational wavenumbers of the metal complexes (cm1).

Compound mC@O mC@N mCOO mCOO Dm mCOO mCOO Dm

amide imino asym sym asym sym
Cephalexin 1657
HL 1656 1635
[MnL(OAc)(H2O)] 1656 1625 1600 1420 180 1580 1370 210
[CoL(OAc)(H2O)] 1653 1620 1600 1420 180 1585 1375 210
[NiL(OAc)(H2O)] 1653 1620 1605 1425 180 1585 1370 215
[CuL(OAc)(H2O)] 1658 1625 1605 1425 180 1580 1370 210
[ZnL(OAc)(H2O)] 1654 1620 1600 1425 175 1580 1375 205
[AgL(OAc)(H2O)] 1656 1620 1600 1425 175 1585 1375 210

175–180 and 205–215 cm1 in our complexes are very similar to When silver(I) acetate or nitrate was added to an aqueous sus-
that reported for the metal complexes with both bidentate l2- pension or to a 50% MeOH solution of the hydrazone ligand, per-
O,O0 and monodentate carboxylate groups respectively [26], thus formed under aerobic conditions, we observed an obvious color
indicating the carboxylate groups behave both as bidentate change from light yellow to dark orange. This observation suggests
[27,28] and monodentate ligands. the occurrence of oxidation process, (Ag(I) d10 ? Ag(II) d9 com-
The remaining carboxylate bands, namely c(COO), x(COO) and plex), induced by the oxygen in air at an observable and rather
q(COO), formerly at 785, 610 and 530 cm1, respectively, also rapid rate, depending on conditions of concentration and temper-
change as a result of coordination. Furthermore, the appearance ature. The starting Ag(I)-material does not yield an EPR spectrum,
of new bands in the 450–490 cm1 ranges attributed to m(M–N) however, its oxidation was followed by EPR spectroscopy which
stretching vibrations, observed in the spectra of the complexes showed the gradual gain of the signal assigned to the oxidised
(absent in the free ligand) provide evidence that the pyridine moi- product; on completion of the oxidation a EPR signal could be
ety can be bonded to the metal ion through the nitrogen atom. The appreciated. The silver(II) complex, isolated as [AgL(OAc)(H2O)],
metal(II) complexes also display bands in the 1590–1595 and is an orange solid which can be conveniently recrystallized from
1015–1025 cm1 ranges due to m(C@N) of coordinated pyridine water, alcohol or acetonitrile. The compound is paramagnetic, hav-
ring and pyridine ring breathing mode of vibration [29]. ing an effective magnetic moment of 2.10 lB characteristic of one
The bands in the 350–400 cm1 region observed in the com- unpaired electron. Thus the magnetic data supports a mononuclear
plexes, and absent in the free hydrazone ligand, are tentatively structure. In a control experiment it was shown that no EPR signal
assigned to m(M–O) vibrations. Medium intensity band appearing occurred when hydrazone ligand was employed in the absence of
in the 2830–2950 cm1 region corresponds to aliphatic m(C–H), silver(I) ions. The propensity of divalent silver to act as an oxidizing
while aromatic m(C–H) stretches appear in the 3000–3100 cm1 agent is well documented [32] and silver(II) complexes have been
region. A broad band centered at 3426 cm1 for the complex can rare and stable only when formed with ligands which could sur-
be assigned to the m(OH) stretch of coordinated water molecules. vive the strong oxidizing capabilities of silver(II). For the most part
These overall data suggest that the bonding sites are the pyridine these ligands have been nitrogen containing unsaturated heterocy-
nitrogen, azomethine nitrogen and carboxylate oxygen atoms are cles, such as pyridines and related compounds. Ligand oxidation
involved in coordination in the complexes and that the hydrazone did not occur in the above reaction mixtures as shown by IR and
behaves as a tridentate monoanionic NNO chelating agent. UV–Vis spectra, and the compound appears to be stable in the solid
state. The formation of this novel complex illustrates further the
Magnetic properties effect of the exceptionally strong ligand field exerted by the hydra-

zone ligand. This serves to raise antibonding level ðdx2 y2 Þ of the
Magnetic susceptibility is the degree of the magnetization of a Ag(I) ion of hydrazone complex to such a high energy that electron
material in response to a magnet. The method measures the Boltz- is easily removed.
mann occupation of all energy levels. From the molar magnetic The powder EPR spectrum of the orange product at room tem-
susceptibility values, corrected magnetic moments were calculated perature shows an axial spectrum with principal g-factor values at
using Pascal’s constants [30]. The magnitudes of the magnetic 2.17 and 2.04 for parallel and perpendicular tensors respectively,
moments for the paramagnetic complexes fall within the ranges and they coincide with other reported values for silver(II) com-
associated with spin-free high spin ions in octahedral fields. The plexes [33–35]. The experimental hyperfine structure observed is
manganese(II) complex has a magnetic moment value of 5.90 lB presented in Fig. 2. The spectrum is a superposition of the hyper-
which is typical of high spin d5 systems with five unpaired elec- fine doublet structure characteristic of the silver nuclear spin
trons and S = 5/2 ground state. The cobalt(II) complex has a mag- (I = 1/2) and the superhyperfine couplings due to 14N nuclei
netic moment of 4.60 lB which is a typical value of a d7 system (I = 1). The number of the superhyperfine lines due to azomethine
with three unpaired electrons indicating a quartet state in an octa- and pyridine nitrogen nuclei coordinated to the silver center is
hedral arrangement around the metal, as compared with the expected to be two sets of five (according to the known rule for
reported values for octahedral complexes of cobalt(II) (4.7– the number of the superhyperfine lines to be equal to 2nI + 1,
5.2 lB) [31]. The nickel(II) complex has a magnetic moment of where I is the nuclear magnetic moment and n is the number of
3.24 lB characteristic of two unpaired electrons and greater than the corresponding nuclei) with relative intensity of the superhy-
the spin-only value, presumably due to the orbital contribution perfine lines, 1:2:3:2:1. It could be even more complicated consid-
resulting from the transfer of an electron from the dx2 y2 orbital ering the presence of superhyperfine structure due to magnetic
to the dxy orbital. The complex therefore probably has distorted interaction in both, parallel and in perpendicular directions, with
octahedral geometry. At room temperature a magnetic moment the nuclei of silver, 107Ag, I = 1/2, 51.82% and 109Ag, I = 1/2,
of 1.9–2.2 lB is usually observed for mononuclear copper(II) com- 48.18%, which have the magnetic dipole moment of 0.1135 and
plexes, regardless of stereochemistry [31]. A magnetic moment of 0.1305, respectively. The EPR line has a width of 550 G. The line
2.10 lB is observed for the copper(II) compound in the solid state. width does not change very much with decreasing temperature
 J.R. Anacona, M. Rincones / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175 173

in the complexes [41]. The manganese(II) complex shows a very

broad absorption at 22,220–20,830 cm1 probably due to the coin-
cidence of charge transfer, d ? p⁄, L ? M and intraligand n ? p⁄
transitions [42,43]. The visible region spectrum of the cobalt(II)
complex indicates additional two bands at 25,640 and
20,000 cm1, attributed to metal–ligand charge and 4T1g(F) ?
4
A2g(F) (m2) transitions respectively, suggesting octahedral stereo-
chemistry around the metal ion [44]. The UV–Vis spectrum of the
nickel(II) complex presents two major absorptions maxima, at
25,000 and 18,182 cm1 due to d–d bands which may be assigned,
considering that the immediate coordination sphere of the metal is
Oh symmetry, to the spin allowed transitions 3A2g ? 3T1g(P) (m3)
and 3A2g ? 3T1g(F) (m2) respectively [45]. Ligand field parameters
(10Dq, B, b,) and m1 have been calculated for Ni(II) complex by using
m2, m3 and the procedure of Lever [41]. The results are satisfactory
owing to the ratio m2/m1 is found to be 1.57 as required for the octa-
hedral Ni(II) complexes [46]. The values of m1 = 11,570 cm1, the
magnetic moment (lspin only < lobs  lS+L) and the calculated ligand
field parameters of Dq = 1157 cm1, B = 565 cm1 and b = 0.54 are
also consistent with the octahedral geometry. The low value of
Racah parameter (B) for the complexes compared to the free ion
value indicates significant covalent character of the metal–ligand
bonds. The nephelauxetic ratio b also support covalent character
in this complex. The electronic spectrum of copper(II) complex
exhibits a broad band centered at 14,280 cm1 (e = 280.1 M1
cm1) assigned to the 2E2g ? 2T2g transition in a distorted octahe-
dral geometry around the copper(II) ion. A band at 24,400 cm1 is
assigned to charge-transfer, mainly of the L ? Cu type [41]. The
electronic spectrum of silver(II) complex presents a broad band at
25,000 cm1 assigned to charge transfer of the L ? Ag type. The
spectrum of the Zn(II) complex exhibits a strong intense charge
Fig. 2. EPR spectrum of [AgL(OAc)(H2O)] complex at room temperature.
transfer transition (LMCT) band at 27,000 cm1.

(down to 120 K) and should be the result of the dipolar and Coordination sites
exchange interactions between the silver ions. Superhyperfine
structure is also observed at both temperatures. The coordination chemistry of transition metal ions with ceftri-
From the g values of a d9 metal complex, we can obtain very axone [47], cefotaxime [48], cefepime [12] and ceftazidime [49]
important information about the structure of the complex. The antibiotics have been reported. In the present case, the hydrazone
geometric parameter G, which is a measure of the exchange inter- ligand containing cephalexin has a number of potential donor
action between the silver(II) centers in the polycrystalline com- atoms in various positions which can bind to the metal ions form-
pound, is calculated using the equation: G = (g||  2.0023)/ ing multinuclear chelates. From the data it appears that each metal
(g\  2.0023) for axial spectrum. If G < 4.0 considerable exchange ion lies in a distorted octahedron coordination sphere and the
interaction is indicated in the solid complex [36–39]. The G param- hydrazone would act as an efficient pseudo-encapsulating ligand,
eter has the value of 4.25 suggesting that there is no exchange with pyridine nitrogen, azomethine nitrogen and carboxylate oxy-
interaction between silver(II) centers. gen atoms, presumably bound to the octahedral ions. Thus, the
metal ions in the [ML(OAc)(H2O)] complexes containing one water
Electronic spectra molecule and acetate with bidentate mode at the vertices of an
octahedron are hexacoordinate. We have attempted to grow single
A long-term UV–Vis study was carried out to verify the stability crystals of the metal chelates but in no case have we had any suc-
of new complexes in DMSO solution. Compared with hydrazone cess, due to their insolubility in common organic solvents. The
ligand, it is significant to note that the absorption wavelengths of complexes only form amorphous materials as revealed by their
new complexes hardly varied for up to 1 month, meaning that XRD patterns. Up to now no crystal structures of hydrazone com-
new complexes were stable in DMSO solution. The electronic spec- plexes containing cephalexin have been reported. These studies
trum of the hydrazone ligand in DMSO solution showed three represent a contribution to future crystallographic analyses, which
broad bands at 32,250, 30,300 and 27,780 cm1. The former two are complicated by the difficulties in obtaining X-ray quality crys-
bands are due to the p–p⁄ transitions within the aromatic rings tals of cephalosporin derived complexes. Although crystal struc-
and remains almost unchanged in the spectra of metal complexes, ture of the complexes are not known, the coordination
while the third band, due to the p-p⁄ transition within the >C@N– environment of mononuclear complexes, may tentatively be pro-
NH– chromophore, is shifted to a longer wavelength as a conse- posed (Fig. 3).
quence of coordination when binding with the metal atom, con-
firming the formation of metal complexes and reflecting that Antibacterial activity
azomethine nitrogen is involved in coordination [40].
The UV–Vis spectra of the metal complexes in DMSO solutions Preliminary screening for antimicrobial activities of the stock
present absorption maxima attributable to the hydrazone ligand solutions were performed qualitatively using the disc diffusion
together with the absorptions, around 25,000–12,500 cm1, due assay. In vitro antimicrobial activities were measured from the
to ligand to metal charge transfer and d–d transitions of the metal diameter of clear inhibition zones caused by samples against the
174 J.R. Anacona, M. Rincones / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 141 (2015) 169–175

same bacteria and under the identical experimental conditions. As Table 3

assessed by color, the complexes remain intact during biological Antibacterial activity.

testing. In order to clarify role of DMSO any participating and Compound Zone of inhibition (mm)
metal(II) acetate salts in the biological screening, separate studies E.C. A.B. E.F.
were carried out with the solutions alone of DMSO and the free
Cephalexin 40.0 ± 1.0 50.0 ± 1.0 20.0 ± 1.5
metal salt and they have been found that they have no effect on [HL] 38.0 ± 1.0 50.0 ± 1.0 21.0 ± 1.0
the growth of any microorganisms taken. The antibacterial activity [MnL(Oac)(H2O)] 45.0 ± 1.0 52.0 ± 1.0 30.0 ± 1.0
of hydrazone ligand HL as well as its metal(II) complexes were [CoL(OAc)(H2O)] 45.0 ± 1.0 55.0 ± 1.0 50.0 ± 1.0
tested on against E. faecalis, E. coli and A. baumannii bacteria and [NiL(OAc)(H2O)] 15.0 ± 1.0 35.0 ± 1.0 5.0 ± 1.0
[CuL(OAc)(H2O)] 20.0 ± 1.5 40.0 ± 1.0 5.0 ± 1.0
compared to cephalexin used as standard.
[ZnL(OAc)(H2O)] 5.0 ± 1.0 10.0 ± 1.0 5.0 ± 1.0
The average results are shown in Table 3 where can be appreci- [AgL(OAc)(H2O)] 42.0 ± 1.0 52.0 ± 1.0 25.0 ± 1.0
ated that the hydrazone ligand and its metal complexes have dif-
E.C. Escherichia coli, A.B. Acinetobacter baumannii, E.F. Enterococcus faecalis. Values
ferent behavior compared with standard antibiotics against the
are the mean ± standard deviation of the mean.
same bacteria. Thus: (1) Cephalexin and the hydrazone ligand have
similar antibacterial activity. (2) Manganese(II), cobalt(II) and sil-
ver(II) complexes were found to have higher activity than the
established drug and HL against the bacteria strains studied under obtained results may highlight that the bactericidal activity of
the test conditions, showing that they have good activity as bacte- hydrazone metal complexes compared to hydrazone ligand and
ricides. (3) The antibacterial activity of nickel(II), copper(II) and cephalexin may reflect a different mechanistic pathway by which
zinc(II) complexes show to be less toxic than the reference drug they react with the PBP active sites to achieve formation of a stable
and hydrazone ligand. PBP-inhibitor adduct. The level of resistance to b-lactam metal
According to Tweedy’s theory [50], the enlarged activity of com- complexes is determined by the amount, nature and kinetic prop-
plexes compared to ligands may be attributed to increased delocal- erties of the PBPs.
isation of p electrons of ligand over the whole chelate ring.
Chelation reduces the polarity of metal ions and increases the lipo- Conclusions
philic character of the chelate, which subsequently favors its per-
meation through the lipid layers of the cell membrane and A hydrazone ligand derived from cephalexin and 2-hydrazino-
blocking the metal binding sites on enzymes of microorganism. pyridine and its transition metal complexes have been prepared.
However, in the present case, the in vitro antibacterial activities The hydrazone coordination to metal occurs through the pyri-
demonstrated that manganese(II), cobalt(II) and silver(II) com- dine-N, azomethine-N and carboxylate-O atoms. Manganese(II),
plexes have higher antimicrobial activity in comparison with that cobalt(II) and silver(II) complexes were found to have higher bac-
of the ligand HL, whereas nickel(II), copper(II) and zinc(II) com- tericidal activity than the uncomplexed cephalexin and the hydra-
plexes showed lower activity against the tested strains compared zone ligand against the bacteria strains, showing that they have a
to hydrazone ligand. Therefore, antimicrobial activity must be good activity as bactericides. The nickel(II), copper(II) and zinc(II)
influenced by other factors beyond membrane permeability. complexes showed to be less toxic than the reference drug and
The targets for b-lactam antibiotics are cell wall-synthesizing the hydrazone ligand. Apart of membrane permeability, antibacte-
enzymes (penicillin binding proteins, PBPs) which are found as rial activity of metal complexes depends mainly on the metal ion
both membrane-bound and cytoplasmic enzymes that catalyze and the type of microorganism.
cross-linking reactions. b-Lactam antibiotics, interfere with cell
wall synthesis by binding covalently to the PBPs catalytic site. PBPs Acknowledgements
are present in almost all bacteria, but they vary from species to
species differing in amount, molecular weight, affinity for b-lactam The authors express their sincere thanks to Comision de Inves-
antibiotics and enzymatic function (e.g., transpeptidase, carboxy- tigación from the Universidad de Oriente for financial support.
peptidase, or endopeptidase) [51].
The results in Table 3 can be understood considering that the
Appendix A. Supplementary data
enzyme probably serves primarily to hold catalytic groups or the
substrate in the proper positions and is possible to expect that
Supplementary data associated with this article can be found, in
hydrazone metal complexes may change the stereochemistry
the online version, at http://dx.doi.org/10.1016/j.saa.2015.01.009.
required in solvolytic reactions on an enzyme surface. The

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