New tridentate azo-azomethines and their copper(II) complexes: Synthesis, solvent effect on tautomerism, electrochemical and biological studies

Article abstract of DOI:10.1016/j.molstruc.2015.04.043

In this study, three azo-azomethines and their copper(II) complexes were prepared and characterized by analytical and spectroscopic methods. The complexes prepared were found to be mononuclear and the chelation of the ligands to the copper(II) ions occurs

Full text of DOI:10.1016/j.molstruc.2015.04.043

Journal of Molecular Structure 1096 (2015) 64–73
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Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
New tridentate azo–azomethines and their copper(II) complexes:
Synthesis, solvent effect on tautomerism, electrochemical and
biological studies
Munire Sarigul a, Pervin Deveci b, Muhammet Kose a, Ugur Arslan c, Hatice Türk Dagi c,
Mukerrem Kurtoglu a,
⇑
a
_
Department of Chemistry, Kahramanmaraßs Sütçü Imam University, Kahramanmaraßs 46050, Turkey
b Department of Chemistry, Selçuk University, Konya 58140, Turkey
c Department of Clinical Microbiology, Medical Faculty, Selcuk University, Konya 58140, Turkey
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
ꢀ
ꢀ
Three azo–azomethines and their
copper(II) complexes are synthesized.
The new compounds were
characterized by IR, UV–Vis., 1H and
13C NMR, mass spectrometry and
elemental analysis.
ꢀ
ꢀ
ꢀ
The tautomeric behaviors of the azo–
azomethines in solution were
examined by UV–Vis. study.
The redox behaviors of the
compounds were investigated by
cyclic voltammetry.
The antibacterial activity of the
compounds was also determined.
a r t i c l e i n f o
a b s t r a c t
Article history:
In this study, three azo–azomethines and their copper(II) complexes were prepared and characterized by
analytical and spectroscopic methods. The complexes prepared were found to be mononuclear and the
chelation of the ligands to the copper(II) ions occurs through two phenolic oxygens and a nitrogen atom
of the azomethine group of the ligand. The tautomeric behaviors of the azo–azomethines in solution were
studied by UV–Vis. spectra in three organic solvents with different polarity (CHCl3, DMSO and DMF) at
room temperature. The redox behaviors of the azo–azomethines and their Cu(II) complexes were inves-
tigated by cyclic voltammetry (CV) in DMSO solution containing 0.1 M tetrabutylammonium tetrafluo-
roborate (TBATFB) as supporting electrolyte. Additionally, the antibacterial activity was also evaluated
by the broth microdilution methods against Staphylococcus aureus, Enterococcus faecalis, Escherichia coli,
Klebsiella pneumoniae and Pseudomonas aeruginosa. The compounds were found to be less effective
against all bacteria tested than two reference antibiotics (ampicillin and gentamicin).
Received 23 February 2015
Received in revised form 27 April 2015
Accepted 28 April 2015
Available online 6 May 2015
Keywords:
Azo–azomethines
Tautomer
Copper(II) complexes
Solvent effect
Cyclic voltammetry
Antibacterial activity
Ó 2015 Elsevier B.V. All rights reserved.
Introduction
have advanced applications in organic synthesis and high technol-
ogy areas such as laser, liquid crystalline displays and ink-jet print-
Azo compounds are versatile molecules that find widespread
applications as dyes and pigments in textile industry. They also
ers [1–3]. Azo dyes are an important class of the organic
photoactive materials, due to their excellent optical switching
properties [4–8]. In addition, the azo dyes and their metal com-
plexes are involved in many biological reactions such as inhibition
of DNA, RNA and biological activity against bacteria and fungi
⇑
Corresponding author. Tel.: +90 344 280 1450; fax: +90 344 280 1352.
E-mail addresses: kurtoglu01@gmail.com, mkurtoglu@ksu.edu.tr (M. Kurtoglu).
http://dx.doi.org/10.1016/j.molstruc.2015.04.043
0022-2860/Ó 2015 Elsevier B.V. All rights reserved.

M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
65
[9,10]. They are increasingly used in textile, leather and plastic
industries [11]. These compounds have a great ability to coordinate
with many metal ions and form stable complexes. The coordina-
tion compounds of azo–azomethine ligands are also widely used
in medicine, for corrosion prevention, metal recovery as well as
to treat nuclear wastes [12]. Furthermore, since the azo com-
pounds have a good thermal stability, the azo–azomethine com-
pounds find important applications in the optical data storage as
recording layer on digital versatile disk-recordable [13]. Schiff
bases derived from azo chromophore group containing aromatic
aldehydes and their metal complexes have been extensively
studied [14] because of their interesting physical and chemical
properties [15] and potentially useful biological activities [16].
Azo–azomethine ligands contain both azo and azomethine groups.
Azo groups possess excellent donor properties and play an
important role in coordination chemistry [14,17,18].
Spectrometer. Mass spectra were recorded on a Thermo Fisher
Exactive + Triversa Nanomate mass spectrometer. The IR spectra
were obtained (4000–400 cmꢁ1) using a Perkin Elmer spectrum
400 FTIR spectrophotometer (resolution; 0.5–4 cmꢁ1). Electronic
absorption spectra were recorded on a T80-UV–Vis. Spectrometer
PG Instruments Ltd. Melting points were obtained with
Electrothermal LDT 9200 Apparatus in open capillaries.
a
All the electrochemical experiments were performed using a
Gamry Reference 600 workstation (Gamry, Pennsylvania, USA)
electrochemical analyzer (Model 600C series) equipped with BAS
C3 cell stand. The working electrode was a bare glassy carbon
(GC) disk (BAS Model MF-2012) with a geometric area of 0.027
cm2. The reference electrode was Ag/Ag+ (0.01 M) in nonaqueous
media, and the counter electrode was a Pt wire.
Electrochemical studies
Proton induced tautomerism has an importance in various
fields of chemistry and particularly in biochemistry [19].
o-Hydroxy Schiff bases exhibit selfisomerisation via intramolecular
proton transfer and they have potential applications in higher
energy radiation detectors and memory storage devices [20].
They may exist in two tautomeric forms (enol and keto forms) in
both solutions and the solid state and tautomerisms affects their
photo-physical and photochemical properties [21].
Copper is an essential trace element are involved in a number of
metabolic activities [22]. Copper complexes containing Schiff bases
have attracted particular interest due to their potential antitumor
activity and interaction with DNA [23,24]. Therefore, investigations
on copper complexes are becoming more prominent in the
research area of bioinorganic chemistry [25,26].
In our laboratory, we have been investigated the synthesis,
spectral, structural and biological properties of new azo
chromophore group containing ligands and their transition metal
complexes [15,18,27–30]. In this study, novel azo–azomethine
compounds, L1H2–L3H2, with ONO donor set were synthesized
via condensation reaction of 2-hydroxy-3-methoxy-5-[(E)-
phenyldiazenyl]benzaldehyde with 2-amino-4-(X) phenol (X = H,
Me and Cl). The new azo–azomethines were characterized by ana-
lytical and spectroscopic methods such as elemental analysis,
FT-IR, UV–Vis., mass, 1H and 13C NMR spectroscopy. Mononuclear
copper(II) complexes of the azo–azomethine ligands were also pre-
pared. To examine the tautomeric behavior of the azo–azomethine
compounds L1H2–L3H2 in solution, electronic spectra in three
organic solvents with different polarity (CHCl3, DMSO and DMF)
were performed at room temperature. In addition, the electro-
chemical properties and biological activities of the synthesized
compounds were investigated.
GC electrodes were prepared by first polishing them with fine
wet emery papers grain size 4000 (Buehler, Lake Bluff, IL, USA) fol-
lowed by a 0.1 and 0.05 lm alumina slurry on a polishing pad
(Buehler, Lake Bluff, IL, USA), to give them a mirror-like appear-
ance. The electrodes were sonicated for 5 min in water and in
50:50 (v/v) isopropyl alcohol and acetonitrile (IPA + MeCN) solu-
tion purified over activated carbon. Before the electrochemical
experiments, the electrodes were dried with an argon gas stream
and the solutions were purged with pure argon gas (99.999%) at
least for 10 min and an argon atmosphere was maintained over
the solution during experiments. Electrochemical studies of the
prepared azo–azomethines were performed in
a solution of
1 mM of all the compounds in 0.1 M TBATFB in DMSO versus an
Ag/Ag+ (0.01 M) reference electrode using CV with a scan rate of
100 mV sꢁ1 between 0 and ꢁ2.3 V.
Determination of antibacterial activity
The minimal inhibitory concentration (MIC) were determined
by broth microdilution method according to Clinical and
Laboratory Standarts Instıtute (CLSI) guidelines (2011) in
Mueller–Hinton broth (MHB) (Becton Dickinson, Sparks, MD) with
an inoculum of approximately 5 ꢂ 105 colony-forming units
(CFU)/mL [32]. The in vitro antibacterial activity of the substances
was evaluated against standart strains; Staphylococcus aureus ATCC
(American Type Culture Collection) 29213, Enterococcus faecalis
ATCC 29212, Escherichia coli ATCC 25922, Klebsiella pneumoniae
ATCC 700603 and Pseudomonas aeruginosa ATCC 27853. All the
synthesized complexes were weighted (10.24 mg) and dissolved
in DMSO (10 mL) to prepare the stock solutions. The serial dilution
from 256 to 0.25
lg/mL were made in a 96-wells microplate. To
Experimental
each well 100 L of a bacterial suspension, obtained from a 24 h
l
culture, containing ꢃ5 ꢂ 105 CFU/mL was added. The plate was
incubated at 35 °C for 24 h. The data were reported as MICs, the
lowest concentration of antibiotic and complexes inhibiting visible
growth after 24 h of incubation at 35 °C. For quality control of the
method ampicillin (IE Ulugay, Turkey) and gentamicin (IE Ulugay,
Turkey) were tested as antimicrobial agents. These experiments
were carried out in duplicate.
Reagents
Chemicals and solvents used were of analytical reagent grades
and used without any further purification. Copper(II) acetate
monohydrate was purchased from Merck and used without purifi-
cation.
Aniline,
2-aminophenol,
2-amino-4-methylphenol,
2-amino-4-chlorophenol, and 2-hydroxy-3-methoxybenzaldehyde
were purchased from the Aldrich Chemical Company and
2-hydroxy-3-methoxy-5-[(E)-phenyldiazenyl]benzaldehyde was
prepared according to a published procedure [31].
Preparation of azo–azomethines (L1H2, L2H2ꢄ1/2H2O and L3H2)
Azo–azomethines (L1H2, L2H2ꢄ1/2H2O and L3H2) were synthe-
sized by addition of corresponding substituted 2-aminophenols
(2-aminophenol, 0.109 g, 1 mmol; 2-amino-p-cresol, 0.123 g,
1 mmol; 2-amino-4-chlorophenol, 1 mmol, 0.143 g) in MeOH
Apparatus
Carbon, hydrogen and nitrogen elemental analyses were per-
formed with a model CHNS-932 (LECO) elemental analyzer. NMR
spectra were performed using a Bruker Avance III HD 600 MHz.
(10 mL) to a methanolic solution of 2-hydroxy-3-methoxy-
5-[(E)-phenyldiazenyl]benzaldehyde (0.256 g, 1 mmol). The mix-
tures were stirred for about 3 h at room temperature and the

66
M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
colored precipiates were collected, washed with cold methanol and
dried in air.
salt [Cu(CH3COO)2ꢄH2O] (1 mmol), was added dropwise. The reac-
tion mixture was refluxed for 3 h. After cooling, green colored pre-
cipitates from the mixtures were separated out and washed with
cold MeOH and dried in air.
L1H2, (1); yield: 0.21 g, 61%, color: orange. M.p.: 285–286 °C.
Analysis Calc. for C20H17N3O3 (347.367): C, 69.15; H, 4.93 N,
12.10%. Found: C, 69.36; H, 5.02; N, 11.95. ESI-MS in MeOH (m/z
(rel. intensity) assignment, positive ion): 347.36 (55%) [M]+.
NMR: 1H (d6-DMSO as solvent, d in ppm,), 15.30 (b, 1H, ANHA),
10.46 (b, 1H, Ph-OH), 9.24 (s, 1H, @CH-N), 7.85 (s, 1H,
aromatic-H), 7.83–7.82 (d, 2H, aromatic-H), 7.69–7.67 (d, 1H,
aromatic-H), 7.58–7.55 (t, 2H, aromatic-H), 7.49–7.48
(d, 1H, aromatic-H), 7.46–7.45 (s, 1H, aromatic-H), 7.21–7.18 (t,
1H, aromatic-H), 7.05–7.03 (d, 1H, aromatic-H), 6.99–6.96 (t, 1H,
aromatic-H), 3.85 (s, 3H, OCH3). 13C NMR (d6-DMSO as solvent, d
in ppm): 167.87 (@CHAN), 158.33 and 152.63 (PhAOH or Ph@O),
152.32, 149.89, 141.84, 130.47, 129.80, 129.07, 128.85, 128.76,
122.415, 120.35, 118.49, 116.86, 115.57, 102.92 (aromatic C
atoms), 55.90 (OCH3). IR (KBr, cmꢁ1): 3356–2925 (OH), 3061
(ArAH), 2992, 2831 (aliphatic CAH), 1635 (C@O), 1613 (C@N),
1590 (CAC aromatic), 1541 (AN@NA), 1273 (CAO), 1127
(CAOAC).
[CuL1(CH3OH)], (4); yield: 0.083 g, 65%, color: green.
M.p. > 250 °C. Analysis Calc. for C21H19CuN3O4 (440.939 g/mol): C,
57.20; H, 4.34 N, 9.53%. Found: C, 57.97 H, 3.877; N, 10.00%.
ESI-MS in MeOH (m/z (rel. intensity) assignment, positive ion):
441.00 (40%) [M + 1]+. FT-IR (KBr, cmꢁ1): 3338–2816 (OH), 3056
(ArAH), 2957, 2839 (aliphatic CAH), 1601 (C@N), 1583 (CAC aro-
matic), 1539 (AN@NA), 1270 (CAO), 1129 (CAOAC), 982 (CAO
of MeOH), 868 (CuAOHMe), 511 (CuAO), 427 (CuAN).
[CuL2(H2O)], (5); yield: 0.111 g, 91%, color: green. M.p. > 250 °C.
Analysis Calc. for C21H19CuN3O4 (440.939 g/mol): C, 57.20; H, 4.34;
N, 9.53%. Found: C, 56.73; H, 4.048; N, 9.636%. ESI-MS in MeOH
(m/z (rel. intensity) assignment, positive ion): 440.94 (30%)
[M + H]+. FT-IR (KBr, cmꢁ1): 2915, 2835 (aliphatic CAH), 1595
(C@N), 1590 (CAC aromatic), 1538 (AN@NA), 1128 (CAOAC),
870 (CuAOH2), 514 (CuAO), 421 (CuAN).
[CuL3(H2O)], (6); yield: 0.119 g, 98%, color: green. M.p. > 250 °C.
Analysis Calc. for C20H16ClCuN3O4 (461.358 g/mol): C, 52.07; H,
3.50; N, 9.11%. Found: C, 54.08; H, 3.39; N, 9.40%. ESI-MS in
MeOH (m/z (rel. intensity) assignment, positive ion): 461.36 (5%)
[M-1]+. FT-IR (KBr, cmꢁ1): 3060 (ArAH), 2994, 2835 (aliphatic
CAH), 1602 (C@N), 1580 (CAC aromatic), 1538 (AN@NA), 1255
(CAO), 1129 (CAOAC), 871 (CuAOH2), 511 (CuAO), 439 (CuAN).
L2H2ꢄ1/2H2O, (2); yield: 0.247 g, 68%, color: dark red. M.p.: 242–
243 °C. Analysis Calc. for C21H20N3O3ꢄ1/2H2O (370.40 g/mol): C,
68.09; H, 5.44; N, 11.34%. Found: C, 68.77; H, 5.518; N, 11.41%.
ESI-MS in MeOH (m/z (rel. intensity) assignment, positive ion):
373.40 (20%) [M + 2]+. NMR: 1H (d6-DMSO as solvent, d in ppm,),
15.31–15.29 (d, 1H, ANHA), 10.23 (s, 1H, Ph-OH), 9.23–9.22
(d, 1H, @CHAN), 7.84–7.83 (d, 2H, aromatic-H), 7.81
(s, 1H, aromatic-H), 7.57–7.54 (t, 2H, aromatic-H), 7.53
(s, 1H, aromatic-H), 7.48–7.46 (t, 1H, aromatic-H), 7.45 (s, 1H,
aromatic-H), 7.01–7.00. (d, 1H, aromatic-H), 6.93–6.91 (d, 1H,
aromatic-H), 3.84 (s, 3H, OCH3), 2.30 (s, 3H, PhACH3). 13C NMR
(d6-DMSO as solvent, d in ppm): 168.52 (@CHAN), 157.87 and
152.64 (PhAOH or Ph@O), 152.46, 147.54, 141.675, 130.41,
129.80, 129.35, 129.27, 129.015, 128.35, 122.40, 118.57, 116.70,
115.40, 102.75 (aromatic C atoms), 55.88 (OCH3), 20.79 (PhAC). IR
(KBr, cmꢁ1): 3338–2816 (OH), 3061 (ArAH), 2967 (aliphatic CAH),
1633 (C@O), 1612 (C@N), 1590 (CAC aromatic), 1537 (AN@NA),
1263 (CAO), 1122 (CAOAC).
Results and discussion
Synthesis
The azo–azomethine ligands L1H2–L3H2 were synthesized by
the condensation reaction of equimolar quantities of
2-aminophenol, or its two derivatives with 2-hydroxy-3-
methoxy-5-[(E)-phenyldiazenyl]benzaldehyde in MeOH. The com-
pounds L1H2–L3H2 were obtained in yields 61, 68 and 88%, respec-
tively. The purity of the ligands were confirmed by TLC technique
and C, H, N elemental analyses. Mononuclear Cu(II) complexes
[CuL1(CH3OH)] (4), [CuL2(H2O)] (5) and [CuL3(H2O)] (6) were pre-
pared by the reaction of one equivalent of the ligands with one
equivalent Cu(OAc)2ꢄH2O according to the following equation:
L3H2, (3); yield: 0.335 g, 88%, color: dark red. M.p.: 255–256 °C.
Analysis Calc. for C20H16ClN3O3 (381.812 g/mol): C, 62.91; H, 4.22;
N, 11.01%. Found: C, 63.18; H, 4.30; N, 10.98%. ESI-MS in MeOH
(m/z (rel. intensity) assignment, positive ion): 383.81 (30%)
[M + 2]+. NMR: 1H (d6-DMSO as solvent, d in ppm,), 14.98–14.97
(b, 1H, ANHA), 10.60 (s, 1H, Ph-OH), 9.25 (s, 1H, @CH-N), 7.86–
7.85 (d, 2H, aromatic-H), 7.84 (s, 1H, aromatic-H), 7.75 (s, 1H,
aromatic-H), 7.76–7.75 (s, 1H, aromatic-H), 7.59–7.56 (t, 2H,
aromatic-H), 7.51–7.49 (t, 2H, aromatic-H), 7.23–7.21 (dd, 1H,
aromatic-H), 7.03–7.01 (d, 1H, aromatic-H), 3.87 (s, 3H, OCH3).
13C NMR (d6-DMSO as solvent, d in ppm): 164.85 (@CHAN),
160.06 and 152.54 (Ph-OH or Ph@O), 151.64, 149.31, 142.66,
131.73, 130.77, 129.85, 128.15, 127.33, 123.85, 122.52, 118.64,
118.18, 116.46, 109.18, 103.81 (aromatic C atoms), 56.05 (OCH3).
FT-IR (KBr, cmꢁ1): 3330–2843 (OH), 3060 (ArAH), 2835 (aliphatic
CAH), 1611 (C@N), 1587 (CAC aromatic), 1539 (AN@NA), 1257
(CAO), 1123 (CAOAC).
MeOHꢁCHCl3
ðCH3COOÞ Cu:H2O þ LxH2
½CuLxðROHÞꢅ
ƒƒƒƒƒƒƒƒƒ!
2
Reflux
ðx ¼ 1; R ¼ H or CH3 and x ¼ 2; 3 R ¼ HÞ
The copper(II) complexes derived from azo–azomethine ligands
are green colored, stable toward air. The complexes are insoluble
in water and some common organic solvents but soluble in
dimethylformamide and dimethylsulfoxide. Single crystals of the
new ligands synthesised and their copper(II) chelates could not be
isolated from organic solvents, and so no definite structures are
available. However, structures of the tridentate ligands and their
Cu(II) complexes were proposed by the analytical and spectroscopic
data as shown in Scheme 1, and Fig. 1, respectively. The possible
tautomers of synthesized ligands are shown in Fig. 2. The physical
characteristics and microanalytical data of the ligands and their
copper(II) complexes are given in experimental section. The struc-
tures of the obtained ligands and their respective copper(II) com-
plexes were elucidated by elemental analysis, and infrared, mass,
and ultraviolet–visible spectroscopy. Microanalytical data for the
ligands and their copper(II) complexes are in good agreement with
theoretical values. The spectroscopic and analytical measurements
showed that the complexes have general formulae of [CuL(X)],
where L = dianionic azo–azomethines, X = MeOH (L2H2ꢄ1/2H2O)
and H2O (L1H2 or L3H2). The Cu(II):L ratio was found to be 1:1 for
Preparation of the copper complexes
The general procedure for the preparation of the copper(II)
complexes
is
as
follows:
To
a
solution
of
the
2-{(E)-[(2-hydroxyphenyl)imino]methyl}-6-methoxy-4-[(E)-phen-
yldiazenyl]phenol [L1H2], 2-{(E)-[(2-hydroxy-5-methylphenyl)
imino]methyl}-6-methoxy-4-[(E)-phenyldiazenyl]phenol, [L2H2ꢄ1/
2H2O],
2-{(E)-[(5-chloro-2-hydroxyphenyl)
imino]methyl}-6-
methoxy-4-[(E)-phenyldiazenyl]phenol [L3H2], (1 mmol) dissolved
in CHCl3 (15 mL) heating at 80 °C, methanolic solution of copper

M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
67
NH2
N+
(i )
N
(ii)
O
OH
O
N
N
HO
N
OH
N
(v)
(iv )
(iii)
OH
N
OH
N
O
O
Cl
H
HO
OH
N
O
N
N
N
.1/2H2O
L1H2
( )
L3H2
( )
N
1
3
L2H2
( )
2
Scheme 1. The general preparation route of azo–azomethines. (i) NaNO2, HCl, 0 °C, (ii) 2-hydroxy-3-methoxybenzaldehyde, (iii) 2-aminophenol, (iv) 2-amino-p-cresol, and
(v) 2-amino-4-chlorophenol.
which are assigned to
are indicative of keto-amine tautomer in solid state. However, the
compound L3H2 do not exhibit a
(C@O) vibration suggesting the
m(C@O) vibrations, respectively. These bands
R1
O
O
H
O
m
O
enol-imine tautomer in the solid state. Previous X-ray crystallo-
graphic studies on o-hydroxy Schiff bases derived from salicylalde-
hyde and 2-hydroxy anilines indicated that both enol-imine and
keto-amine forms may exist in solid state [33–36]. The azo group
Cu
N
N
N
m
(N@N) stretching was observed at 1541, 1537 and 1539 cmꢁ1
for the compounds L1H2–L3H2, respectively.
H
X-ray crystallographic study of N-(2-hydroxy-5-chlorophenyl)
salicylaldimine derived from salicylaldeyhde and 5-chloro-2-
hydroxyaniline showed that both tautomeric forms (keto and enol
forms) are present in the solid state. This shows that there is a
quick shift between two tautomeric forms suggesting the low
energy difference between these two forms. Ledbetter proposed
that the second o-hydroxy group in the aniline ring of o-hydroxy
Schiff bases exhibits stronger keto-tautomerism due to this second
hydroxy group [37].
R
R = H, R1 = H (4); R = CH3, R1 = H (5); R = Cl, R1 = H (6)
Fig. 1. The proposed structure of the mononuclear Cu(II) complexes.
all three complexes. The chelation of the ligands to the copper(II)
ions occurs through the phenolic oxygen atoms and the nitrogen
atom of the ligand. ESI mass spectra for the synthesized azo–
azomethines and their Cu(II) complexes exhibit several fragmenta-
tion signals, however molecular ion peaks were observed with lower
intensities.
Upon complexation with the Cu(II) ion, m
(C@O) vibrations in L1H2
and L2H2ꢄ1/2H2O disappeared in the spectra suggesting the imine
form in the complexes. In the FT-IR spectra of the complexes, aꢁz1ome-
thine
m
(C@N) bands shifted to lower values 1601 cm
for
FT-IR spectra
[CuL1(CH3OH)], 1595 cmꢁ1 for [CuL2(H2O)] and 1602 cmꢁ1 for
[CuL3(H2O)] suggesting coordination of the imine nitrogen to the
metal center. The absence of the band due to phenolic OH group in
the spectra of Cu(II) complexes confirm the coordination of ligands
to the metal via phenolate oxygen atom. The vibrations of the
CuAO and CAN bonds are located at the low wavenumbers of
The characteristic FT-IR absorption bands of the azo–azome-
thine compounds and their mononuclear Cu(II) complexes were
determined and the data are given in experimental section. FT-IR
spectra of L1H2 and its Cu(II) complex [CuL1(CH3OH)] (4) are given
in Figs. 3 and 4 and S1–S4, respectively. The solid state FT-IR spec-
tra of azo–azomethines showed phenolic OH band in 3360–
2900 cmꢁ1 range. A strong sharp absorption band at 1613 cmꢁ1
for L1H2, 1612 cmꢁ1 for L2H2ꢄ1/2H2O, and 1611 cmꢁ1 for L3H2 in
the spectra of the azo–azomethines can be assigned to the
400–650 cmꢁ1
.
NMR spectra
m
(C@N) stretching. In addition to CH@N stretchings, compounds
The 1H and 13C NMR spectra of azo–azomethines L1H2–L3H2
were recorded in d6-dimethylsulfoxide (DMSO) solution using
L1H2 and L2H2ꢄ1/2H2O exhibits a band at 1635 and 1633 cmꢁ1

68
M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
O
R
O
H
N
N
N
R
N
N
N
H
OH
O
O
H
O
enol-imine
keto-amine
O
O
H
O
N
N
N
H
R
hydrazone-imine
Fig. 2. The possible tautomers of synthesized azo–azomethines (where R = H for L1H2, R = CH3 for L3H2 and R = Cl for L3H2.
93.9
92
90
88
86
84
82
80
78
76
74
72
70
68
66
64
62
60
58
56
54
52
50
48
46
44
1708.21
804.13
826.18
785.86
553.09
496.57
514.63
923.66
2831.72
1441.96
855.01
459.23
2992.02
975.66 874.88
537.56
470.89
%T
422.57
1072.04
1027.62
996.44
618.93
1306.41
770.33
1464.15
1635.07
1613.63
1361.31
744.62
1293.57
1273.86
1158.54
1215.42
1541.00
1107.32
688.00
1390.76
1182.40
1127.73
42
40.5
4000.0
3600
3200
2800
2400
2000
1800
1600
1400
1200
1000
800
600
400.0
cm-1
Fig. 3. FT-IR spectrum of L1H2.
tetramethylsilane (TMS) as an internal standard. The 1H NMR and
13C NMR data of the azo–azomethines are collected in experimen-
tal section. The 1H NMR and 13C NMR spectra of L2H2 are shown in
Figs. 5 and 6 and rest of the NMR spectra are given in the supple-
mentary file (Figs. S5–S8). The 1H NMR spectra of L1H2–L3H2 dis-
play a singlet signal at d 3.85, 3.80 and 3.87 ppm with an
integration equivalent to three hydrogens corresponding to the
methoxy (OCH3) group. In the 1H NMR spectrum of L1H2, a singlet
at d 2.30 ppm was assigned to the protons of methyl group
(PhACH3). The 1H NMR spectra of L1H2–L3H2 exhibited broad sig-
nals for the phenolic proton in the aniline ring in the range of d
10.23–10.60 ppm. The azo–azomethines also exhibit signals at d
14.97–15.30 and 9.22–9.25 ppm ranges assigned to ANHA and
@CHAN protons, respectively. In the spectrum of L1H2, doublet sig-
nals of ANHA and @CHAN protons were observed at d 9.22–9.23
and 15.29–15.30 ppm as shown in Fig. 5. These chemical shifts sug-
gests that compounds L1H2–L3H2 favor keto–amine tautomer in
DMSO solution that are in agreement with the results of related

M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
69
96.0
94
92
90
88
86
84
82
80
78
76
74
72
70
68
66
64
62
60
58
56
54
2100.58
1729.46
2957.78
955.47
918.94
545.08
1071.43
1181.02
1004.84
982.47
658.55
621.77
699.63
1409.16
%T
738.75
847.76
830.40
868.97
588.54
1464.55
1539.38
1110.29
686.31
1440.23
1482.65
1601.70
1583.69
1270.47
1259.45
1389.32
1361.97
1289.79
753.45
518.03
1129.69
51.8
4000.0
3600
3200
2800
2400
2000
1800
1600
1400
1200
1000
800
600
400.0
cm-1
Fig. 4. FT-IR spectrum of [CuL1(CH3OH)] (4).
Fig. 5. The 1H NMR spectrum of L2H2 azo–azomethine.

70
M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
Fig. 6. The 13C NMR spectrum of L2H2 azo–azomethine.
compounds [38,39]. These chemical shifts were also supported by
UV–Vis. spectra in DMSO solution.
DMSO and DMF) at room temperature. UV–Vis. spectra of
L2H2ꢄ1/2H2O and L3H2 are shown in Figs. 7 and 8 and electronic
data are illustrated in Table 1. The absorption spectra of azo–
azomethines are similar and exhibit two absorpiton maximums
in the solvents studied CHCl3, DMSO and DMF. Maximum absorp-
tion bands observed in the range of 320–420 nm were assigned to
The 13C NMR spectra of azo–azomethine compounds L1H2–L3H2
exhibit consistent carbon atom signals with their proposed
structures. In the 13C NMR spectra of compounds L1H2–L3H2
display signals at d 55.90, 55.875 and 56.05 ppm assigned to the
carbon atoms of the methoxy groups (OCH3), respectively. The
the
p– p–
p⁄ transitions of the aromatic rings. The p⁄ transitions
methyl carbon (PhACH3) signal in L2H2 was observed at
d
shifted to higher wavelength values (batochromic effect) with
the increase in polarity. The second band was observed in the
range of 450–550 nm. The previous tautomeric studies on
o-hydroxy Schiff bases suggested that a band above 400 nm is
indicative of existence of keto-amine tautomer in solution [39].
20.79 ppm. In 13C NMR spectra of L1H2–L3H2, the signals at d
167.865 and 168.521 and 164.848 ppm was assigned to the carbon
atom of the C@O group, respectively. The signal belong to carbon
atom phenolic group (CAOH) were seen at d 158.33, 157.87,
160.055 ppm for L1H2–L3H2, respectively. Rest of the carbon atom
signals were observed in the range of d 152–102 ppm for all three
compounds.
UV–Vis. spectra
There are several different spectroscopic techniques including
FT-IR, UV–Vis., NMR and X-ray crystallography techniques to eval-
uate tautomeric species of o-hydroxy Schiff bases both in solutions
and solid state [40–42]. UV–Vis. spectroscopy is very useful tool for
studying tautomerism in Schiff bases involving a hydroxyl group in
ortho position to the azomethine group. The previous reports indi-
cated that o-hydroxy Schiff bases with azo group (AN@NA) may
exist in keto and/or enol forms in solid state [43] and solvent
media [10]. There are several factors influencing the tautomeric
equilibrium including substitute groups in the molecule, solvent
polarity and temperature [39].
To examine the tautomeric behavior of the azo–azomethine
compounds L1H2–L3H2 in solution, electronic spectra were per-
formed in three organic solvents with different polarity (CHCl3,
Fig. 7. UV–Vis. spectra of L2H2ꢄ1/2H2O ligand in solvents DMSO, DMF and CHCl3.

M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
71
Table 1
The keto-amine band intensity increases with the increase in the
solvent polarity. In CHCl3 solution, the intensity of the
keto-amine band is lower than in DMSO and DMF for all three
compounds. The electronic absorption spectra of the azo–
azomethine compounds at different volume ratios of the applied
pair solvents DMSO/H2O were also measured. The absorption
curves of L2H2ꢄ1/2H2O in DMSO/H2O mixtures are shown in
Fig. 9. In DMSO/H2O mixture, the absorption bands shifted to lower
wavelengths (hypsochromic effect) and absorption values
(hypochromic effect).
UV–Vis. absorption bands for the synthesized azo–azomethines in various solvents
[kmax (nm), absorbance].
Compounds
CHCl3
DMSO
DMF
L1H2 (1)
365(2.389),
482(0.257)
360(2.181),
475(0.518)
365(2.278),
480(0.269)
–
382(2.389),
482(0.257)
385(2.411),
473(1.956)
390(2.17),
475(2.007)
–
365(2.063),
475(1.307)
380(1.607),
475(1.178)
365(2.12),
470(1.091)
420(1.167)
415(2.654)
435(2.868)
L2H2ꢄ1/2H2O (2)
L3H2 (3)
[CuL1(MeOH)] (4)
[CuL2(H2O)] (5)
[CuL3(H2O)] (6)
–
–
–
–
Absorption spectra for Cu(II) complexes were also recorded in
DMF solution. The complexes show only one absorption band in
the range of 330–500 nm assigned to the
p–
p⁄ and M ? L charge
transitions. The keto-amine band in the range of 450–550 nm
observed for ligands disappeared in the spectra of the complexes.
In the spectra, no metal induced d-d transitions were observed in
the solvents and concentrations studies. UV–Vis. spectrum of the
complex [CuL1(CH3OH)] (4) is shown in Fig. 10.
Electrochemistry
Electrochemical behaviors of the new azo–azomethines
(L1H2–L3H2) and their copper(II) metal complexes were investi-
gated using CV technique in DMSO solution containing 0.1 M
TBATFB in the range from 0 to ꢁ2.3 V.
For all azo–azomethine ligands studied, cyclic voltammograms
at 100 mV sꢁ1 consist of two cathodic peaks in the range
ꢁ1.45–(ꢁ1.58) V and ꢁ2.05–(ꢁ2.12); no anodic wave occurs in
the reverse scan. This behavior was observed for a wide range of
scan rates from 25 to 1000 mV sꢁ1. Hence, such a reduction process
should correspond to a totally irreversible electron transfer. The CV
curves for L1H2 and [CuL1(MeOH)] are shown in Fig. 11 and electro-
chemical data of all these compounds are given in Table 2. It is
believed that the reduction peaks correspond to the AN@NA
(azo) and AC@NA (imino) groups of the organic ligands in solution
[44]. The electrochemical properties of Cu(II) metal complexes,
have been studied in order to consider spectral and structural
changes accompanying electron transfer. The cyclic voltammo-
gram of the complex [CuL1(MeOH)], displays three reduction
waves in potentials ꢁ0.83, ꢁ1.19 and ꢁ1.59 V. The peak at
Epc = ꢁ0.83 V and Epc = ꢁ1.19 V is characteristic for CuII ? CuI and
CuI ? Cu0 reductions, respectively [45]. The irreversible reduction
wave at ꢁ1.59 V is related to reduction of L1H2 ligand [46,47]. In
other complexes, the cyclic voltammetric CuII ? CuI reduction pro-
cess was observed at ꢁ0.72 and ꢁ0.80 V, for [CuL2(H2O)] and
[CuL3(H2O)], respectively. For Cu(II) complexes, the absence of
the reduction peak of the imine group indicates that, owing to
Fig. 9. UV–Vis. spectra of L2H2ꢄ1/2H2O ligand in DMSO-H2O mixtures.
Fig. 10. UV–Vis. spectra of the ligand L1H2 and its Cu(II) complex in DMF.
the lack of transferable hydroxylic protons, the reduction poten-
tials of the dianionic ligands have been shifted beyond the lower
limit of the potential interval considered in the experimental mea-
surements [48].
Antibacterial activity
MIC values of the synthesised azo–azomethines and their
mononuclear copper complexes against bacteria were between 8
and 128
(8 g/mL) against E. faecalis strain but this value is higher than the
reference antibiotics (ampicillin and gentamicin). [CuL1(MeOH)]
and L1H2 substances had the highest MIC values (128
g/mL)
l
g/mL. The ligand L3H2 showed the lowest MIC value
l
Fig. 8. UV–Vis. spectra of L3H2 ligand in various solvents.
l

72
M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
Fig. 11. Cyclic voltammograms recorded for the reduction of 1.0 mM L1H2 (a) and [CuL1(MeOH)] (b) at a GC electrode in DMSO containing 0.1 M TBATFB in the range from 0 to
ꢁ2.3 V. Scan rate is 100 mV sꢁ1
.
Table 2
(32 lg/mL) against all Gram negative bacteria than the other sub-
stances. The complexes [CuL2(H2O)] (5) [CuL3(H2O)] (6) exhibit
more biological activity than their corresponding free ligands
against E. coli, P. aeruginosa and K. pneumoniae under identical
experimental conditions. The possible cause of the increased biolog-
ical activity of the metal chelates compared to that of the free ligand
may be explained in terms of Tweedy’s chelation theory [49].
According to this theory, the polarity of the central metal ion is
reduced by chelation via partial sharing of positive charge of metal
ion with the donor atoms and possible p-electron delocalization
within the whole chelate ring. In addition, the lipophilic character
of the central metal atom is increased by chelation which subse-
quently favors its permeation through the lipid layer of the cell
membrane [49–51].
Cyclic voltammetric parameters for the azo–azomethines and their copper(II)
complexes.
Compounds
Epc (V); ligand
reductions
Epc (V); CuII ? CuI Epc (V); CuI ? Cu0
L1H2(1)
ꢁ1.48, ꢁ2.08
ꢁ1.45, ꢁ2.05 V
ꢁ1.58, ꢁ2.12
–
–
L2H2ꢄ1/2H2O (2)
–
–
L3H2 (3)
–
–
[CuL1(MeOH)] (4) ꢁ1.59
ꢁ0.83
ꢁ0.72
ꢁ0.80
ꢁ1.19
ꢁ1.28
ꢁ1.14
[CuL2(H2O)] (5)
[CuL3(H2O)] (6)
ꢁ1.63
ꢁ1.62
Epc indicates the cathodic peak potential for irreversible reduction processes.
against K. pneumoniae and P. aeruginosa strains and this value is
higher than gentamicin but is equal to that of ampicillin. These sub-
stances had the same MIC values (128
lg/mL) against E. coli and
Conclusion
higher than gentamicin and ampicillin. Also S. aureus and E. faecalis
which are Gram positive bacteria were more sensitive to these sub-
The azo–azomethines L1H2–L3H2 and their copper(II) complexes
were synthesized and characterized by spectroscopic and analyti-
cal methods. The analytical data show that the metal ligand stoi-
chiometry in all these copper(II) complexes is 1:1. The spectral
data show that the ligands act as tridentate coordinating through
nitrogen atom of the azomethine and two oxygen atoms of hydro-
xyl groups of the ligands. Proton induced tautomerism for L1H2–
L3H2 was investigated by 1H, 13C NMR, FT-IR and UV–Vis. spectral
methods. Existence of keto-amine tautomer for ligands L1H2–
L3H2 were confirmed by an absorption band in the range of 450–
550 nm in solutions (CHCl3, DMSO and DMF). It is evident from
the cyclic voltammogram of the tridentate ligands and the
copper(II) complexes that all the compounds are electroactive over
stances [CuL1(MeOH)] (64 g/mL) and L1H2 (32
l lg/mL)] but higher
than reference antibiotics. The results are given in Table 3. All sub-
stances had the higher MIC values against Gram negative bacteria
than Gram positive bacteria. L1H2 and [CuL1(MeOH)] substances
had the highest MIC values against all bacteria but the MIC values
for Gram negative bacteria (128
bacteria. L1H2 substance showed a lower MIC value (32
[CuL1(MeOH)] (64
g/mL) against S. aureus and E. faecalis.
L2H2ꢄ1/2H2O and L3H2 had the same MIC value (64 g/mL) against
all Gram negative bacteria but while L2H2ꢄ1/2H2O had the same
l
g/mL) higher than Gram positive
l
g/mL) than
l
l
MIC value (16 lg/mL) against all Gram positive bacteria.
[CuL2(H2O)] and [CuL3(H2O)] substances had the lower MIC value
Table 3
Antibacterial activity of the prepared azo–azomethines and their copper(II) complexes as MICs values (lg/mL).
Compounds
E. coli ATCC 25922
P. aeruginosa ATCC 27853
K. pneumoniae ATCC 700603
S. aureus ATCC 29213
E. faecalis ATCC 29212
L1H2 (1)
128
64
64
128
32
32
8
128
64
64
128
32
32
128
64
64
128
32
32
32
16
16
64
16
16
0.5
1
32
16
8
64
16
16
2
L2H2ꢄ1/2H2O (2)
L3H2 (3)
[CuL1(MeOH)] (4)
[CuL2(H2O)] (5)
[CuL3(H2O)] (6)
Ampicillin
128
0.5
128
1
Gentamicin
1
4

M. Sarigul et al. / Journal of Molecular Structure 1096 (2015) 64–73
73
[21] Ç. Albayrak, G. Kastas, M. Odabasoglu, O. Büyükgüngör, J. Mol. Struct. 1000
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a range from 0.0 to ꢁ2.3 V in DMSO solvent. MIC values of azo–
azomethines against bacteria were between at 8 and 128 g/mL.
The synthesized compounds were found to exhibit lower MIC
l
values against Gram positive bacteria than Gram negative bacteria.
[24] N.M. Hosny, M.A. Hussien, F.M. Radwan, N. Nawar, Spectrochim. Acta A 132
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Appendix A. Supplementary material
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the online version, at http://dx.doi.org/10.1016/j.molstruc.2015.04.
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