Synthesis, structure elucidation, biological screening, molecular modeling and DNA binding of some Cu(II) chelates incorporating imines derived from amino acids

Article abstract of DOI:10.1016/j.molstruc.2015.09.039

Three tridentate Schiff bases amino acids were prepared by direct condensation of 3-methoxysalicylaldehyde (MS) or 4-diethylaminosalicylaldehyde (DS) with α-amino acid ligands [l-phenylalanine (P), l-histidine (H) and DL-tryptophan (T)]. The prepared Schi

Full text of DOI:10.1016/j.molstruc.2015.09.039

Accepted Manuscript
Synthesis, Structure Elucidation, Biological Screening, Molecular Modeling and DNA
binding of Some Cu(II) Chelates incorporating Imines Derived from Amino Acids
Laila H. Abdel-Rahman, Ahmed M. Abu-Dief, Mohammed Ismael, Mounir A.A.
Mohamed, Nahla Ali Hashem
PII:
S0022-2860(15)30281-7
10.1016/j.molstruc.2015.09.039
MOLSTR 21845
DOI:
Reference:
To appear in: Journal of Molecular Structure
Received Date: 30 June 2015
Revised Date: 7 September 2015
Accepted Date: 29 September 2015
Please cite this article as: L.H. Abdel-Rahman, A.M. Abu-Dief M. Ismael, M.A.A. Mohamed, N.A.
Hashem, Synthesis, Structure Elucidation, Biological Screening, Molecular Modeling and DNA binding
of Some Cu(II) Chelates incorporating Imines Derived from Amino Acids, Journal of Molecular Structure
(2015), doi: 10.1016/j.molstruc.2015.09.039.
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ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT
Synthesis, Structure Elucidation, Biological Screening, Molecular Modeling and DNA binding of Some Cu(II)
Chelates incorporating Imines Derived from Amino Acids
*
Laila H. Abdel-Rahman, Ahmed M. Abu-Dief Mohammed Ismael, Mounir A. A. Mohamed and Nahla Ali
,
Hashem
Chemistry Department, Faculty of Science, Sohag University, Sohag 82524, Egypt
Corresponding author: E- mail address: ahmed_benzoic@yahoo.com
Abstract:
Three tridentate Schiff bases amino acids were prepared by direct condensation of 3-methoxysalicylaldehyde
(MS) or 4-diethylaminosalicylaldehyde (DS) with α-amino acid ligands [L-phenylalanine (P), L-histidine (H) and DL-
1
tryptophan (T)]. The prepared Schiff bases amino acids were investigated by melting points, elemental analysis, H
1
3
NMR and C NMR, IR, UV –Vis spectra, conductivity and magnetic measurements analyses. Subsequently, copper
was introduced and Cu(II) complexes formed. These complexes were analyzed by thermal and elemental analyses and
further investigated by FT-IR and UV/Vis spectroscopies. The experimental results indicating that all Cu(II) complexes
contain hydrated water molecules (except DSPCu complex) and don't contain coordinated water molecules. The kinetic
and thermal parameters were extracted from the thermal data using Coast and Redfern method. The molar conductance
values of the Schiff base amino acid ligands and their Cu(II) complexes were relatively low, showing that these
compounds have non-electrolytic nature. Magnetic susceptibility measurements showed the diamagnetic nature of the
Schiff base amino acid ligands and paramagnetic nature of their complexes. Additionally, a spectrophotometric method
was determined to extract their stability constants. It was found that the complexes possess 1: 2 (M: L) stoichiometry.
The results suggested that 3-methoxysalicylaldehyde and 4-diethylaminosalicylaldehyde amino acid Schiff bases
behave as monobasic tridentate ONO ligands and coordinate Cu(II) ions in octahedral geometry according to the
general formula [Cu(HL) ].nH O. To further understanding the structural and electronic properties of these complexes,
2
2
Density Functional Theory (DFT) calculations were employed and provided a satisfactory description. The optimized
structures of MST Schiff base ligand and its complex were calculated using DFT. The antimicrobial activity of the
Schiff base ligands and their complexes were screened against some types of bacteria such as B. subtilis (+ve), E. coli
(-ve) and M. luteus (+ve) and some types of fungi such as A. niger, C. glabrata and S. cerevisiae. The results of these
studies indicated that the metal complexes exhibit a stronger antibacterial and antifungal efficiency compared to their
corresponding ligands. The complexes were screened for antiviral activity against a panel of DNA and RNA viruses.
Minimum cytotoxic and minimum virus inhibitory concentrations of these complexes were determined. The mode of
interaction between complexes and CT-DNA was monitored using absorption spectra, viscosity measurements and gel
electrophoreses.
Keywords: Amino acids, Imine, Molecular modeling, antimicrobial, antiviral, DNA
Introduction
The chemistry of metal-drug coordination compounds nowadays has a greater attention than before due to the
importance needs particularly in designing more biologically active drugs [1]. Metal ions are known to affect the action
of many drugs [2]. Several metal chelates are known to possess antimicrobial, antiviral and anticancer activity. In many
cases, metal chelates have been found to be more antimicrobial than the chelating agents themselves [2-4]. Schiff bases
ligands and their complexes have a variety of applications in clinical, analytical and industrial field [5].
1

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Among these, heterocyclic Schiff base ligands and their complexes do have significant interest because of
pharmacological properties [7]. Furthermore, the interaction of these complexes with DNA has gained much attention
due to their possible applications as new therapeutic agents [8]. Some drugs show increased activity when administered
as metal chalets and inhibit the growth of tumors [9]. The transition metal ions are responsible for the proper
functioning of different enzymes. Certain drugs play a vital role as bio-ligands in the biological systems. Also, nitrogen
bases such as (pyridine, pyrimidine and pyrazine) and amines such as (histamine, carbohydrates and different vitamins)
have a vital role as bio-ligands. Metal complexes of Schiff base phenolates with favorable cell membrane permeability
have been exploited in cancer multidrug resistance and used as antimalarial agents [10].
A number of diseases and their remedies are dependent on metabolism of inorganic constituents. The complexes of
copper with Schiff bases have wide applications in food and dye industries, catalysis, fungicidal, agrochemical,
antiradical activities and biological activities [11, 12].
Cu(II) complexes are also attractive since Cu(II) is known to play a significant role in naturally occurring
biological systems as well as a pharmacological agent. Copper is a biologically relevant element and many enzymes
that depend on copper for their activity have been identified. The metabolic conversions catalyzed by most of these
enzymes are oxidative.
This paper describes the synthesis of binary Cu(II) Schiff base amino acid complexes and characterization by
various physical methods. Moreover, antimicrobial studies of the investigated Cu(II) complexes were performed
against many types of bacteria, fungi and viruses. Moreover, the interaction between DNA and the synthesized Cu(II)
complexes was performed by using absorption spectra, viscosity measurements and gel electrophoresis. 3-
methoxysalicylaldehyde (MS) or 4-diethylaminosalicylaldehyde (DS) is the aldehyde that used in this investigation, the
amino acids are L-phenylalanine (P), L-histidine (H) and DL-tryptophan (T).
Materials
All chemicals reported here, such as 3-methoxysalicylaldehyde (C H O ) (MS), 4-diethylaminosalicylaldehyde
8
8
3
(
C H O N) (DS), amino acids[L-Phenylalanine (P), DL-Tryptophan (T), L-Histidine (H)], Calf thymus DNA (CT –
11 15 2
DNA), and the metal salt [copper acetate (Cu(CH COO) . H O), were purchased from Sigma-Aldrich and used as
3
2
2
received.
Characterization of the prepared Schiff base amino acid ligands and their complexes
Melting points for the isolated ligands and decomposition points for their complexes were monitored on a melting point
apparatus, Cimarec 3 Thermolque. The carbon, hydrogen, and nitrogen contents were determined on a Perkin Elmer
-1
(
2400) CHNS analyzer. IR spectra (4000 – 400 cm ) were recorded on Shimadzu FT-IR model 8101 spectrometer
1
13
using KBr pellets. HNMR and CNMR spectral measurements were determined by using a BRUKER, using DMSO
-1
as an internal reference. The TG/DT analyses were recorded on Shimdzu corporations 60 H at 10 degrees min . The
UV-Vis spectra were recorded on a PG spectrophotometer model T+80 at 298 K. Magnetic susceptibility
measurements of the metal complexes were done on a Gouy balance at room temperature using Hg[Co(SCN) ] as a
4
-1
calibrant. Molar conductance was measured on an Elico CM-180 conductometer using 1 mmol L solutions in DMF. A
HANNA 211 pH meter at 298 K equipped with a CL-51B combined electrode was used for pH measurements,
calibrated against standard buffers (pH 4.02 and 9.18) before measurements. Quantum chemical calculations for 3-
methoxysalicyaldehyde and tryptophan (MST) ligand structure were performed using MOPAC2000 [13] with
2

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WinMOPAC 2.0 [14] as a graphic interface. At the beginning, structure the starting material was optimized with the
eigenvector-following routine (EF) [15] using the semi-empirical PM3 method [16, 17]. To investigate the 3D structure
and molecular stability for the Cu-MST complex accurate density functional theory (DFT) calculations were done
using Gaussian 03 software package [18]. Calculations were carried out at DFT level of theory with pseudo potential
functions, 6-311G (p,d) [19] basis set for ligand atoms, and LANL2DZ [20] basis set with effective core potential
(ECP) for Cu ion. Antimicrobial screening was carried by using agar well diffusion. Viscosity measurements were
o
performed by using viscometer immersed in a thermo stated water bath maintained at 25 C. Gel electrophoresis was
visualized under UV a transilluminator and photographed with a Panasonic DMC-LZ5 Lumix Digital Camera.
Synthesis of Schiff base amino acids
To a stirred solution of 3-methoxysalicylaldehyde or 4-diethylaminosalicylaldehyde (5 mmol, 0.76 g, 0.995 g
respectively) in ethanol, 40 ml of the appropriate (L-Phenylalanine (P), DL-Tryptophan (T), L-Histidine (H)) (5 mmol,
0
.825 g, 1.045 g, 1.02 g respectively) was added. The resultant mixture in each case was heated under reflux for 2h.
The solid products were collected by filtration, washed with hot ethanol, dried at room temperature and finally stored in
an air tight sample vial for further use [21].
Complexes preparation
Aqueous solutions of the amino acids were prepared by dissolving (5 mmol, 0.825 g, 1.045 g, 1.02 g respectively
in 40 ml of aqueous-ethanol mixture) of each (P, T, H). Each of the solutions was mixed with
3
-methoxysalicylaldehyde (5 mmol, 0.76 g, 50 ml hot ethanol) or 4-diethylaminosalicylaldehyde (5 mmol, 0.995 g, 50
o
ml hot ethanol). Then the mixture was stirred at 70 C for 1 hour. Cu(II) acetate mono hydrate solution of (2.5 mmol,
0
.5 g in 40 ml aqueous-ethanol mixture) was added to the previous mixtures. The color changed from yellow to (green
for MSTCu, dark green for DSPCu and bright brown for DSHCu) after stirring at room temperature for 3 hour. The
obtained products were evaporated over night. The obtained solid products were filtered, washed with water, and dried
in vacuum over anhydrous CaCl [2, 3, 22]. The structures of the investigated Schiff base amino acid ligands and their
2
complexes are shown in Table 1.
Magnetic susceptibility measurements
The magnetic susceptibility measurement is one of the most useful methods available to the coordination chemists
for studying the electronic structure of a transition metal complex. It provides fundamental information about the
bonding and stereochemistry of the metal complexes [23]. The magnetic properties of the coordination compounds are
based on the effect of the ligands on the spectroscopic terms of the metal ions. The susceptibility per gram atom of a
paramagnetic metal ion in a particular compound is determined by measuring the molar susceptibility of the
compounds and applying diamagnetic corrections for the other ions or molecules in the compounds. The diamagnetic
corrections can be determined by various methods [24], however Pascal’s constants are more often used to calculate the
corrections. The magnetic susceptibility and moments can generally be calculated by using the following relationships
[
25].
µ
eff = 2.83 X T
M
Where ꢀ , the magnetic moment (in Bohr Magneton), B. M. where T = temperature (K).
eff
3

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-
χ χ
M = M – (diamag. corr.)
-
χM molar magnetic susceptibility before correction,
χ
molar magnetic susceptibility after correction.
M
Kinetic data for thermogarvimetric analysis (TGA) of the prepared complexes
The kinetic parameters of decomposition processes of complexes namely activation energy (E*), enthalpy (H*),
entropy (S*) and Gibbs free energy change of the decomposition (G*) were evaluated graphically by employing the
Coats-Redfern relation [26].
log [
=log [
(1− )] –
(1)
Where W is the mass loss at the completion the decomposition reaction, W is the mass loss up to temperature T, R is
∞
the gas constant and ɸ is the heating rate. Since 1-2RT/E* = 1, the plot of the left hand side of equation (1) against 1/T
would give straight line. E* was then determined from the slope and from the intercept, the Arrhenius constant, A, was
obtained. The other kinetic parameters; the entropy of activation (S*), enthalpy of activation (H*) and the free energy
change of activation (G*) were calculated using the following equation:
S* = 2.303 R log
(2)
H* = E* - RT
G* = H* - TS*
(3)
(4)
Where, (k) and (h) are the Boltzmann's and Planck's constant, respectively.
Antibacterial bioassay
All the prepared Schiff base amino acid ligands and their complexes were screened for their antibacterial activity
against two Gram-positive (Bacillus subtilis and Micrococcus luteus) and one Gram-negative (Escherichia coli)
bacterial strains using the agar well diffusion method [2, 3, 27]. The solution of complexes and ligands were prepared
by dissolving in DMSO at a concentration (15 and 30 mg/ml). Paper discs of Whatman No.1 were sterilized in an
autoclave and saturated with solution of metal complexes, the test organisms were grown on nutrient agar medium in
Petri plates. After that, holes were formed in the agar using a sterile crook borer and these holes were completely filled
o
with the test solutions. The Petri dishes were incubated for 24 h at 37 C. Ciprofloxacin was used as standards for
antibacterial activity.
Antifungal screening
Antifungal activities of all the synthesized Schiff base amino acid ligands and their complexes were studies
against three fungal cultures (A. niger, C. glabrata and S. cerevisiae) using disk diffusion method [2, 3, 28]. These
fungal species were isolated from the infected parts of the host plants, potato dextrose agar. The fungus strains were
directly mixed with potato dextrose agar and dispersed into the Petri dishes. The discs were soaked in DMSO in which
the test complexes were dissolved (concentration of ligands and complexes 15 and 30 mg/ml). Filter paper discs of 6
o
mm in size were saturated with solution of complexes. The plates were inverted and incubated at 35 C for 48 h. The
4

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plates were then observed and the diameters of the inhibition zones were measured. Amphotricine B was used as
standard for antifungal activity.
Antiviral activities of the prepared complexes
For Herps Simplex Virus (HSV): the Vero cell line was maintained in RPMI 1640 (Gibco, Tunisia) supplemented
to fetal bovine serum (10 %, v/v), L-Glutamin (2ꢁꢀM), penicillin (100 U/ꢀl), and streptomycin (100ꢁꢀg/ml). Cells were
incubated at 37 °C in a 5% CO humidified atmosphere. Coxsakievirus B Nancy strain (kindly provided by Pr. Bruno
2
3
Pozzetto, Laboratory of Bacteriology-Virology, Saint-Etienne, France) was propagated in Vero cells.
Confluent Vero cell cultures were treated with three non-cytotoxic concentrations of the submitted sample during
4
and after virus infection in two sets of experiments as follows: (1) 5 x 10 TCID50 of the virus was exposed to essential
oil for one hr at 37 °C. Then 100 ꢀl of the mixture were added to the cells cultured fluently in 96-well flat-bottom
microtiter plate (100 ꢀl); (2) Cells were treated with sample (100 ꢀl) for one hr at 37 °C. After one hour of incubation
4
at 37 °C, 5 x 10 TCID50 of the virus (100 ꢀl) were added.
All plates were incubated at CO -incubator for 48ꢁhrs. The viability of the infected and non-infected cells was
2
evaluated according to the absorbance values of formazan using the MTT inclusion assay, as described in cytotoxicity
assay. The percentage of protection was calculated as follows:
Percent protectionꢁ=ꢁ[(ODT) - (ODC) V] / [(ODC) M - (ODC) V]ꢁ×ꢁ100
Where (ODT), (ODC) V and (ODC) M are the absorbance of the test sample, the absorbance of the virus-infected
control (no compound) and absorbance of the mock-infected control (no virus and no compound), respectively. The 50
%
inhibition concentration (IC50) was calculated by regression curve analysis, which is defined as the concentration of
the essential oil that inhibits the viral infection by 50 %.
For Tobacco Mosaic Virus (TMV): plants grown in Magenta® vessels were ready for mechanical inoculation
with the virus within 2 weeks. Czech isolate of Turnip Tobacco mosaic virus (TMV) [29] maintained in the virus
collection at the Institute of Plant Molecular Biology was used for this study. Singular virus infection was checked by
sucrose density gradient during the virus purification, electron microscopy, preparation of antiserum and serology. The
inoculum was prepared from infected B. pekinensis cv. Manoko plants with mosaic symptoms 3 weeks after
inoculation. A leaf sample was homogenized in a mortar in a 1: 10 (w/v) ratio with 0.1 ꢀ phosphate buffer, pH = 7.0,
and filtered through 0.22 m sterile filters (Millipore). Mechanical inoculation of plants was conducted on the first 2
leaves in a flow box using a cotton pad soaked in inoculum mixed with carborundum powder.
Interaction of the prepared complexes with Calf Thymus DNA (CT-DNA)
ꢀ
Absorption spectral studies
Electronic absorption spectrum of the complex was recorded before and after addition of CT-DNA in the presence
of Tris-HCl buffer (pH 7.2). Different concentration of metal complexes was titrated with incremental amounts of CT-
DNA over the range (3 –30 ꢀM). The equilibrium binding constant (K ) values for the interaction of the complex with
b
CT-DNA were obtained from absorption spectral titration data using the following equation [2, 3, 30].
[
DNA]/ (ε - ε ) = [DNA]/ (ε - ε ) + 1/K (ε - ε )
a
f
b
f
b
b
f
5

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Where ε , ε and ε are the molar extinction coefficient observed for the charge transfer absorption at a given DNA
a
f
f
concentration, the extinction coefficient at the complex free in solution and the extinction coefficient of the complex
when fully bound to DNA respectively. K and [DNA] are the equilibrium binding constant and the concentration in
b
nucleotides respectively. A plot of [DNA]/ (ε - ε ) versus [DNA] gives K as the ratio of the slope to the intercept. The
a
f
b
standard Gibb´s free energy for DNA binding was calculated from the following relation.
o
ꢂG = -RT ln K
b
b
Where R, T and K are the general gas constant, the absolute temperature and the binding constant respectively [2, 3]
b
ꢀ
viscosity measurements
Various concentrations of complex (10–250 ꢀM) were added into the viscometer to give a specific mole-ratio
r (r = [complex] / [DNA]) while keeping the DNA concentration constant at (250 ꢀM). The control sample was carried
out on EB by using the same method. After thermal equilibration, the time of the solution's flowing through the
1
/3
capillary was determined by a digital stopwatch. The data were founded as (η/η0) versus the mole-ratio values, where
η and η are the viscosity of DNA in the presence and absence of complex, respectively. Viscosity values were
0
calculated from the observed flow time of DNA containing solutions (t) and corrected for buffer solution (t0),
η = (t - t )/t [2, 3, 22, 31].
0
0
ꢀ
Agarose gel electrophoresis
Solutions of DNA were freshly prepared before each experiment using doubly distilled water containing 0.1 M
Tris - HCl buffer, pH = 7.2. The concentration of DNA solutions was determined by UV-Vis spectrophotometer at 260
-1
-1
nm using a value of 6600 M cm for the absorption coefficient. The purity (freedom from bound protein) was assessed
from the ratio of the absorbencies at 260 nm and 280 nm. In general, the commercial DNA preparation was found to be
free of protein (A260 nm /A280 nm= 1.9) according to this criterion and no further purification was attempted. Cleavage
reactions were run between the metal complexes and DNA, and the prepared solutions were subjected to
electrophoresis on 1 % agarose gel prepared in TBE buffer (45 ꢀM Tris, 45 ꢀM boric acid and 1 ꢀM EDTA, pH 7.3).
Then (20 micron) of ethidium bromide was added to the above solution and mixed well. The gel was mounted into
electrophoretic tank; enough electrophoretic buffers were added to cover the gel. DNA sample (10 micron) and metal
o
complex (20 micron) were mixed with loading dye (bromophenol blue) and incubated for 30 min at 25 C, then loaded
into the well of the submerged gel using a micropipette. The electrophoresis was performed at a constant voltage
(
100 V) for about 1-2 h (until bromophenol blue had passed through 50% of the gel) in TBE buffer. The gel was
visualized under UV a transilluminator and photographed with a Panasonic DMC-LZ5 Lumix Digital Camera [2, 3,
2].
3
Results and discussion
Characterization of the prepared Schiff base amino acid ligands
1
13
H NMR and CNMR spectra of the prepared Schiff base amino acid ligands:
1
The HNMR spectrum (DMSO-d , ppm) of MST ligand show singlet signal at 13.6 for COOH proton
6
respectively, singlet signal at 8.2 for CH=N proton respectively, multiple signals at 7.4 – 6.6 for nine aromatic protons
respectively. Also they show singlet signal at 4.3 for OH proton respectively, singlet signal at 3.7 for CH aliphatic
proton respectively and singlet signal at 3.8 for three OCH protons respectively. Furthermore, they show doublet
3
6

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1
signals at 3.0 for three CH protons respectively and the HNMR spectrum of MST ligand showed singlet signal at 10.9
2
for NH proton respectively.
1
3
The CNMR spectrum of MST display the signals corresponding the different non-equivalent carbon atoms at
different values of δ as follows: at δ 171 ppm (COOH) due to carboxylic acid group in the amino acid, at δ 168 ppm
(CH=N) due to azomethine, at δ 165-70 ppm (14CH – Ar) due to carbon atoms of aromatic rings, at δ 58 ppm (OCH3)
due to carbon atom of methoxy group, at δ 52 ppm (CH) due to aliphatic carbon atom, at δ 29 ppm (CH ) due to
2
carbon atom of methyl group connected with aromatic ring
1
The HNMR spectrum (DMSO-d , ppm) of DSP and DSH ligands show singlet signal at 11.0 and 11.5 for
6
COOH proton respectively, singlet signal at 8.5 and 8.3 for CH=N proton respectively, multiple signals at 7.4 – 6.1 and
7
.5– 6.0 for five and eight aromatic protons respectively. Also they show singlet signal at 5.0 and 4.3 for OH proton
respectively, singlet signal at 3.4 and 3.5 for CH aliphatic proton respectively and quartet signals at 3.4 and 3.3 for four
CH - CH protons respectively. Furthermore, they show triplet signals at 1.1 and 0.9-1.2 for six 2CH -CH protons
2
2
3
3
2
1
respectively and the HNMR spectrum of DSH ligand showed singlet signals at 9.5 for NH proton respectively.
1
3
The CNMR spectrum of DSP and DSH display the signals corresponding the different non-equivalent carbon
atoms at different values of δ as follows : at δ 191 and 197 ppm (COOH) due to carboxylic acid group in the amino
acid, at δ 165 and 163 ppm (CH=N) due to azomethine, at δ 163- 99 and 158 ̶ 100 ppm (12CH – Ar) and (9CH – Ar)
due to aromatic carbon atoms, at δ 50 and 52 ppm (CH) due to aliphatic carbon atom, at δ 40 and 38 ppm (CH -N)
2
corresponding to carbon atom in diethyl group, at δ 39 and 35 ppm (CH -Ar) corresponding to carbon atom connected
2
with aromatic group, at δ 17 and 13 ppm (CH ) due to carbon atom in diethyl group.
3
Characterization of the prepared Schiff base amino acid ligands and their complexes
Microanalysis measurements
Results of empirical formula, Molecular weight, melting points (for ligands), decomposition points (for
complexes), elemental analysis and magnetic moment measurements are given in Table 2. The conductivity values with
-3
-3
-3
different concentrations in the range from (1.4 x 10 to 18.8 x 10 ) Mol dm are shown in Table 2. The molar
conductivity measurements were carried out in dimethylformamide (DMF). All the prepared Schiff base amino acid
ligands and their complexes were relatively low, indicating that these compounds have the non-electrolytic nature [2, 3,
2
2, 33]. The microanalysis results suggest that 3-methoxysalicylaldehyde or 4-diethylaminosalicylaldehyde amino acid
Schiff bases behave as monobasic tridentate ONO ligands and coordinate to Cu(II) in octahedral geometry according to
the general formula[Cu(HL) ].nH O. Magnetic susceptibility measurements values of the prepared Schiff base amino
2
2
acid ligands are diamagnetic and for their complexes are (1.8
complexes, Table 2 [2, 3, 22, 34].
̶ 2.2 B. M.) which suggests octahedral geometry of the
IR and Electronic spectra for the investigated compounds
Results of IR spectra are showed in Table 3. The wavelengths at maximum absorption band (λmax) and the molar
absorptivity (ε) of the different bands in the recorded spectra of the complexes (Fig. 1) are given in Table 4.
The IR spectra provide valuable information regarding the nature of functional group attached to the metal
atom. The IR spectrum of the free ligand is compared with the spectra of the complexes. The IR spectrum of the free
-1
Schiff base amino acid ligands exhibit a broad band around 3441-3389 cm , which is attributed to the stretching
7

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frequency of the aromatic hydroxyl substituent group, perturbed by intramolecular hydrogen bonding [O―H---N]
between phenolic hydrogen and azomethine nitrogen atoms [35, 36, 37]. Further, the appearance of the OH band
-1
around 3465-3395 cm in the spectra of the complexes with an increase in intensity indicates that the hydroxyl oxygen
is coordinated to the M(II) ion without proton displacement [38]. The ligands exhibit the characteristic (C=N) band in
-1
-1
the 1651– 1632 cm region, while the complexes the (C=N) were observed in the 1628-1610 cm region. The (C=N)
stretching frequency is shifted to a lower frequency, indicating that decrease in the (C=N) bond order due to the
coordinate bond of the metal with the imine nitrogen lone pair [39]. The ν(C - O phenolic) vibration of ligands are
-1
observed around 1293-1235 cm , which get shifting to lower or higher frequency region in the complexes indicating
-1
coordination of phenolic oxygen [40]. The ligands exhibit other two intense bands at (1430-1362), (1588-1572) cm
corresponding to symmetric stretching and asymmetric stretching frequencies of (COOH) group, respectively of the
organic ligand. On complexation symmetric and asymmetric bands were shifted to a higher frequency or remained
unaltered in the position of the ligands [41]. The difference between symmetry and asymmetry stretching vibration of
-
-
-1
COO group showed that amino acid Schiff bases coordinated through COO group [42]. At 695-688 cm and 542-569
-1
cm to ν(Cu-N) and ν(Cu-O) stretching, respectively [43, 44].
The electronic absorption spectra of the Schiff bases recorded in DMF is composed of three bands in the 200 -
8
00 nm region. The first band A appearing below 300 nm region can be assigned to the π-π* transition of the aromatic
rings. The second band B is observed within the wavelength range 315-431 nm is due to transition between the π-
orbital localized on the central azomethine (-CH = N-) bond [45] while the third band C located within the 363-379 nm
region can be ascribed to charge transfer within the entire Schiff base molecule. This band is observed in o-hydroxyl
Schiff bases and is based on strong intramolecular hydrogen bonding between the hydroxyl group of the salicylidene
and the azomethine nitrogen [46]. The charge transfer bands being more sensitive to solvent changes than bands
resulting from local transitions. A broad band from 507 to 630 nm, which indicating that band could be mainly
attributed to d → d transition in an octahedral structure of the prepared complexes [47, 48]
Thermogravimetric analysis (TGA)
o
−1
In the present investigation, heating rates were suitably controlled at 10 C min under nitrogen atmosphere,
o
and the weight loss was measured from the ambient temperature up to 750 C. The experimental results are given in
Table 5. Thermogravimetric analyses of the Schiff base amino acid complexes were used to: (i) get information about
the thermal stability of these new complexes, (ii) decide the water molecules (if present) are inside or outside the inner
coordination sphere of the central metal ion, and (iii) suggest a general scheme for thermal decomposition of these
complexes [49]. Thermal analyses providing important information regarding thermal stability of substances and thus
indirectly provide clues to the structure and composition of the substances. In thermogravimetry the change in mass of
the sample is recorded as a function of temperature. It provides the analyst with quantitative measurements of change in
weight associated with any transition. TG can directly record the loss in weight with time or temperature due to
o
dehydration or decomposition. Routine measurements can be made at temperature range from ambient to 750 C with
inert atmosphere [50]. Thermogravimetric analyses of synthesized complexes are observed in 3 steps, (i) a small weight
o
loss which is assigned to loss of lattice water molecules, except DSPCu complex is stable up to 113.1 C, indicating
that absence of lattice water molecules, (ii) maximum weight loss which is assigned to the loss of coordinated water
molecules, but in this paper all complexes don’t contain coordinated water molecules, (iii) gradual weight loss can be
assigned to complete decomposition of ligand moiety around the metal ion. Finally complexes are converted into their
metal ion.
8

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Kinetic Data for TGA of the prepared complexes
The data is tabulated in Table 6. The activation energy of decomposition was in the range 14.4-111.9 KJ /mol. The
high values of the activation energies reflect the thermal stability of the Complexes. The entropy of activation had
negative values in all the complexes, which indicating that the decomposition reactions proceed with a lower rate than
the normal one.
Determination of stoichiometry of the investigated complexes
The stoichiometry of the various complexes formed in solutions via the reaction of Cu(II) with the studied
ligands was determined by applying the spectrophotomertic molar ratio [51] and continuous variation methods [52] as
shown in Figs 2, 3. Maximum in the curve at Xligand =0.62-0.73 implicates a 1: 2 (metal ion to ligand) molecular
association.
The formation constants of the investigated complexes
Stability constant is equilibrium constant for the formation of a complex in solution. Stability constant is a
measure of the strength of the interaction between the reagents that come together to form the complex. There are two
main kinds of complexes, i) compounds formed by the interaction of a metal ion with a ligand and ii) supramolecular
complexes. There are many areas of application in chemistry, biology and medicine. The apparent formation constants
(
K ) of the studied Cu(II) Schiff base amino acid complexes formed in solution were determined from the
f
spectrophotometeric measurements using the continuous variation method [2, 3, 22] according to the following
relation:
Where A (A) and C are the absorbance of the maximum formation of the complex, the arbitrary chosen absorbance
m,
values on either sides of the absorbance pass and the initial concentration of the metal respectively. The obtained Kf
values are indicating that high stability of the prepared complexes. The values of K of the prepared complexes increase
f
in the following order: DSPCu > DSHCu > MSTCu. The values of the stability constant and Gibbs free energy of the
investigated Cu(II) Schiff base amino acid complexes are cited in Table 7. The negative values of Gibbs free energy
mean that the reaction is spontaneous and favorable.
Stability range of the investigated complexes
The pH profile (absorbance νs. pH) presented in Fig. 4 showed typical dissociation curves and a wide stability pH
range [(4-11for DSPCu and DSHCu) and (6-9 for MSTCu)] of the studied complexes. This means the formation of the
complex greatly stabilizes the Schiff base amino acid ligands. This pH range means the prepared complexes are more
favorable to different physiological reactions.
Molecular modeling
Figs. 5, 6 show the optimized structures and electronic configuration of MST and its Cu complex. MST has
numerous donor groups; however three of them are oriented toward each other. Those groups are the carboxylate
-
(
COO ), hydroxyl (OH) and imine group (-N=C).Optimized [Cu(MST) ] complex structure has local octahedral
2
geometry. The stability of this complex is due to the aromatic rings formed between the metal and donor atoms. To
9

ACCEPTED MANUSCRIPT
relative stability of the complex was evaluated by calculating the metal-ligand interaction energy (E
– Ereactants).
complex
According to the current DFT approximation this energy was found to be -39 kcal/ mol. Moreover quantum chemical
descriptors were also calculated to estimate the chemical reactivity of the complex. These descriptors include, the
highest occupied molecular orbital (HOMO), the lowest unoccupied molecular orbital (LUMO), the energy gap (ꢂE
=ELUMO-EHOMO), chemical hardness (η), electronic chemical potential (µ) and electrophilicity index (ϵ) of the prepared
Schiff base amino acid (MST) and its complex. Chemical Hardness (η) is associated with the reactivity of a chemical
system. The harder and less reactive the molecule corresponds to the larger η. Electronic chemical potential (µ) is a
measure of electronegativity of the molecule. The greater η, the less stable or more reactive is the complex.
Electrophilicity Index (ϵ), measures the capacity of a species to accept electrons [16]. Table 8 summarizes the
calculated descriptors, and reflects the high electrophilic capacity of the complex compared with the ligand itself. The
small value of η explains the intense color of the complex.
To further compare the theoretical calculations the experimental results, frequencies single point calculations at
same level of DFT approximation. From these calculations, IR vibrational spectrum for ligand and complex models
was generated (cf Table 3). The following table stats the calculated vibrations for the main function groups in ligand
-1
and complex models. The difference between the calculated and experimental values (~100 cm ) may be attributed to
the approximation method used.
Application of the prepared Schiff base amino acid ligands and their complexes
Antimicrobial studies
The measured zones of inhibition against the growth of various microorganisms are summarized in Table 9. The
antimicrobial screening results exhibited marked enhancement in activity on coordination with the Cu(II) ion against
more testing bacterial and fungal strains Figs. 7, 8. Furthermore, the activities of the prepared Schiff base amino acid
ligands and their complexes were reported in Table 10.
The morphology of the cell wall is key factor that influences the activity of antimicrobial agents. The increase
in biological activity of the metal chelates may be due to the effect of the metal ion on the normal cell process [53].
Chelation considerably reduce the polarity of the metal ion because of partial sharing of their positive charge with the
donor group and possible p-electron delocalization within the whole chelate ring system that is formed during
coordination. Chelation could enhance the lipophilic character of the central metal atom and hence increasing the
hydrophobic character and liposolubility of the complex favoring its permeation through the lipid layers of the cell
membrane. The antimicrobial activity of the three complexes can be referred to the increase of their lipophilic character
which in turn deactivates enzymes responsible for respiration processes and other cellular enzymes, which is playing a
vital role in various metabolic pathways of the tested microorganisms. Activities of the synthesized complexes were
confirmed by calculating the activity index according to the following relation:
Inhibition Zone of compound (mm)
Activity index (A) =
x 100
Inhibition Zone of standard drug (mm)
Complexes have more activity against Gram-positive bacteria than against Gram negative pathogens. The results
show that Gram-positive bacteria were inhibited more strongly than Gram-negative bacteria and that can be explained
by considering the structural features of both bacterial types. Gram-negative bacteria possessing an extra outer layer on
top of the peptidoglycan and this has been found to be highly impermeable. However, Gram-positive bacteria have
polysaccharides in their cell wall called teichoic acid, which is negatively charged and have facilitated the passage of
1
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ACCEPTED MANUSCRIPT
the positive metal ions [54]. An increase in lipophilicity of a metal complex enhances bacterial cell membrane
penetration and blocking of metal binding sites on enzymes.
Antiviral activities of the prepared complexes
In principle, a molecule can act as an anti-viral drug if it inhibits some stage of the virus replication cycle, without
being too toxic to the body's cells. The possible modes of action of anti-viral agents would include being able to:
1
2
3
4
5
. Inactivate extracellular virus particles.
. Prevent viral attachment and/or entry.
. Prevent replication of the viral genome.
. Prevent synthesis of specific viral protein(s).
. Prevent assembly or release of new infectious virions.
Antiviral activities of the prepared complexes were investigated by determination of minimum cytotoxic and
minimum inhibitory concentration which is tabulated in Table 11. The activities of the prepared complexes were higher
in the sequence: DSPCu > MSTCu > DSHCu. The results show that the DSHCu complex has the highest concentration
that inhibit the replication of a virus after overnight incubation and the highest concentration that toxic the virus.
Methodology for DNA binding analysis using absorption spectral studies
The application of electronic absorption spectroscopy in CT-DNA binding studies is one of the most important
techniques [55]. The DNA binding studies were characterized by absorbance maximum at 382 for MSTCu, 380 for
DSPCu and 368 for DSHCu. The addition of increasing higher concentration of DNA led to hypochromic in their
visible absorption spectra as a result of the formation of more stable complexes. The interaction of complexes with
DNA resulted in the decrease of absorption intensity accompanied by a shift towards lower wavelengths (2 to 14 nm),
which corresponds to 25.9 % to 38.5 % reduction (Hypochromism). The spectral changes were used to evaluate the
5
5
5
-1
intrinsic binding constant (K ), it observed 0.8 x 10 , 1.8 x 10 and 9.4 x 10 M for MSTCu, DSPCu and DSHCu,
b
which are intercalated into DNA base pairs [56].The complexes may bind with DNA through intercalative interaction,
which is brought by the π-π interaction between the complex which possesses an aromatic ring moiety in the ligands
and the aromatic heterocyclic bases of DNA. The electronic absorption spectra of the prepared complexes in the
absence and presence of different concentration of buffered CT-DNA are given in Table 12 and Fig. 9.
Methodology for DNA binding analysis using viscosity measurements
Viscosity measurements were also carried out to provide clues for a binding model between complexes and DNA.
In the presence of lower concentrations of complexes, no significant changes were observed in the relative viscosity of
DNA. However, at higher concentrations of complexes, the relative specific viscosity of DNA increased, but the
increase is less than that observed for the typical intercalator EB, indicating that intercalative as shown in Fig. 10. A
classical intercalation model results in lengthening of the DNA helix as base pairs to an increase of DNA viscosity [57,
5
8]. This observation suggests that the mode of DNA binding by complexes involved base pair intercalation.
Agarose gel electrophoresis of CT-DNA interaction with the investigated complexes
DNA binding studies are important for the rational design and construction of new and more efficient drugs
targeted to DNA. The molecules interact reversibly with double stranded DNA, primarily through three modes: (i)
1
1

ACCEPTED MANUSCRIPT
electrostatic interactions with negatively charged nucleic sugar–phosphate structure, (ii) binding interaction with two
groves of DNA double helix and (iii) intercalation between the stacked base pairs of native DNA [59].
The cleavage efficiency of complexes was compared to that of the control is due to their efficient DNA binding
ability control experiment using DNA alone does not show any significant cleavage of CT-DNA even on longer
exposure time. The variation in DNA–cleavage efficiency of ligands/transition metal complexes was due to their
difference in binding ability of ligands /complexes to DNA. The intensity of lanes Fig. 11 were higher in the sequence
DSHCu > DSPCu > MSTCu. It was clear concluded that as the complexes was observed to cleave the DNA, therefore
inhibits the growth of the pathogenic organism by cleaving the genome.
Conclusion
In this paper, coordination chemistry of Schiff base amino acid ligands, obtained from the reaction of 3–
methoxysalicylaldehyde (MS) or 4-diethylaminosalicylaldehyde (DS) with α-amino acids (L-phenylalanine (P), L-
histidine (H), DL-tryptophan (T)). The resulted Schiff base amino acid ligands are mono anionic tridentate ligands.
Cu(II) complexes have been synthesized by using the above Schiff base amino acid ligands. Results of physical
measurements showed that Cu(II) ion is coordinated by two phenolic oxygen atoms, two azomethine N atoms and two
carboxylate O atom to form octahedral complexes with general formula [Cu(HL) ].nH O. The prepared Schiff base
2
2
amino acid ligands and their complexes have non-electrolytic nature. Moreover, the obtained K values increased in the
f
following order DSPCu > DSHCu > MSTCu. The prepared complexes bind to DNA via an intercalative mode
according to the spectral, viscosities measurements and gel electrophoresis. Furthermore, antimicrobial and antiviral
for the prepared complexes were screened. These finding clearly indicated that transition metal based complexes have
many potential applications including new therapeutic reagents for diseases. In addition, these complexes showed better
therapeutic drug for antimicrobial treatment.
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List of Tables:
Table 1: The 2D structure of Schiff base amino acid ligands derived from 3-Methoxysalicylaldehyde (MS)
or 4-Diethylaminosalicylaldehyde (DS) and their complexes
Amino acids
Aldehyde
3
- MethoxySalicylaldehyde (MS)
MST
MSTCu
NH
HN
O
O
N
OH
Tryptophan (T)
N
OH
O
O
Cu
O
O
O
OH
N
O
OH
HN
4
-DiethylaminoSalicylaldehyde (DS)
DSP
DSPCu
N
N
N
OH
O
N
O
Phenylalanine (P)
Cu
N
OH
O
N
O
OH
O
OH
DSH
DSHCu
N
HN
N
N
HN
N
OH
Histidine (H)
N
O
O
Cu
N
OH
O
N
O
OH
N
O
OH
N
HN
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Table 2: Analytical and physical data of Schiff base amino acid ligands and their complexes
Comp.
Molecular
formula
-1
M. p. and
Decomp.
Elemental Analysis
found (calc.) %
ꢀeff
ᴧm, Ohm
2
-1
(
B. M.)
cm mol
o
(
M. wt.)
C H N O
4
( C)
C
H
N
MST
Diamagnetic
1.8
20.5
7.7
240
280
67.28
5.10
8.05
(8.18)
7.21
19
18
2
(338.0)
(67.31) (5.22)
59.49 4.68
(59.54) (4.74)
70.40 6.98
(70.52) (7.06)
64.73 6.20
(64.81) (6.29)
61.82 6.67
MSTCu C H O N Cu
3
8
37 9.5
4
(
764.5)
C H N O
3
(7.33)
8.11
DSP
Diamagnetic
1.8
8.2
160
20
24
2
(340.0)
(8.23)
7.55
DSPCu
C H O N Cu
4.9
>300
275
40
46
6
4
(
741.5)
C H N O
3
(7.59)
16.97
DSH
Diamagnetic
2.2
19.4
8.7
17
24
4
(330)
(61.91) (6.72) (17.02)
55.17 5.95 15.14
(55.22) (6.01) (15.21)
DSHCu C H O N Cu
>300
34
44
7
8
(739.5)
Table 3: IR spectral data of the investigated Schiff base amino acid ligands and their complexes.
Comp.
υ(OH)/H O
υSt (-C=N) υS(COO)
υA(COO)
1362 (m) 1588 (w)
1605 1655
1352 (m) 1566 (m)
1595 1535
υph(C-O) υ (Cu-N)
υ (Cu-O)
2
MST (found)
MST (calc.)
MSTCu (found)
MSTCu (calc.)
DSP
3389 (s)
3535
1651 (s)
1761
1293 (m)
1265
-
-
-
695 (m)
650
-
563 (m)
425
3395 (s)
3485
1628 (s)
1701
1224 (m)
1225
3441 (s)
3465 (s)
3438 (s)
3457 (s)
1632 (s)
1618 (s)
1640 (s)
1630 (s)
1405 (s) 1573 (s)
1380 (m) 1554 (w)
1430 (m) 1572 (w)
1415 (m) 1567 (m)
1235 (s)
1183 (w)
1280 (m)
1237 (m)
-
-
DSPCu
689 (m)
-
542 (m)
-
DSH
DSHCu
688 (w)
569 (w)
S = strong, m = medium, w = weak, ph = phenolic
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7

ACCEPTED MANUSCRIPT
3
-1
-1
Table 4: Molecular electronic spectra, λmax (nm) and εmax (dm mol cm ) of the prepared Schiff base amino acid
ligands and their complexes in DMF at 298 K against DMF as a blank.
Comp.
MST
Assignment
λmax(nm)
εmax
3
-1
-1
(
dm mol cm )
431
100
31
n→π*
n→π*
3
2
15
47
37
π→π*
MSTCu
614
104
438
650
75
d-d band
3
2
79
46
LMCT band
Intraligand band
n→π*
DSP
389
50
630
2
136
100
1192
1664
76
π→π*
DSPCu
d-d band
3
2
75
44
LMCT band
Intraligand band
n→π*
DSH
397
49
507
2
101
46
π→π*
DSHCu
d-d band
3
2
63
42
220
315
LMCT band
Intraligand band
Table 5: Thermal analysis of the prepared complexes
Complex
Temperature
Fragment loss %
Weight loss %
o
C
Molecular
formula
Molecular Found Calc.
weight
MSTCu
20.7-240.2
1.5H O
27.0
214.0
184.0
276.0
63.5
118.0
221.0
334.0
63.5
3.75
3.81
2
2
2
3
41.5-289.7 C H N O
2
26.99 27.06
23.75 24.07
36.97 36.39
12
10
2
90.9-340.5
81.8-527.5
>750
C H O
8 8 5
C H N O
18
16
2
Residu
DSPCu
Cu
8.23
8.31
113.1-209.3
C H N
15.14 15.23
28.67 28.80
45.90 45.95
8
8
2
3
09.3-339.3
39.3-456.8 C H N O
C H NO
12 15 3
20
23
2
3
Residu
DSHCu
>750
15.3-157.9
Cu
H O
2
8.49
2.35
8.53
2.41
18.0
1
57.9-396.1
C H N O
4
196.0
26.47 26.50
7
6
3
3
4
96.1-445.3
45.3-530.9
>750
C H NO
2
194.0
268.0
63.5
26.19 26.24
36.17 36.24
11
16
C H N
4
16
20
Residu
Cu
8.49
8.52
1
8

ACCEPTED MANUSCRIPT
Table 6: The kinetic and the thermodynamic data
Complex
Temp.
E*
A
Thermodynamic Parameters
o
-1
(
C)
KJ/mol
S
S* (J/mol)
H* (KJ/mol)
-1.8
G* (KJ/mol)
MSTCu
218.9
1.0
1.3
-240.3
-242.19
-243.7
-246.6
-242.1
-247.61
-250.8
-230.7
-241.8
-246.9
50.8
63.7
77.5
111.9
33.6
67.0
99.8
14.4
57.8
108.8
2
3
4
72.6
29.3
69.5
-2.3
-2.7
-3.9
DSPCu
DSHCu
143.6
0.5
1.0
0.6
1.2
-1.2
2
4
80.0
11.7
-2.3
-3.4
64.8
-0.5
3
13.0
55.8
-2.0
4
-3.8
≠
Table 7: The formation constant (K ), stability constant (pK) and Gibbs free energy (ꢂG ) values
f
of the prepared complexes in aqueous - ethanol at 298 K.
9
*
Complex
Type of
complex
10 Kf
pK
ꢂ G
-1
kJ mol
MSTCu
DSPCu
DSHCu
1: 2
1: 2
1: 2
1.1
2.5
1.8
20.8
21.6
21.3
-51.6
-53.6
-52.8
Table 8: The calculated HOMO, LUMO, energy gap (ꢂE), chemical hardness (µ), chemical reactivity (η)
and electrophilicity index (ϵ)
Comp.
HOMO LUMO
ꢂE
η
µ
ϵ
(ev)
(ev)
(ev)
(ev)
(ev)
(ev)
MST
-4.20
-1.18
-3.25
3.05
1.88
1.53
0.94
-2.69
-4.19
2.36
9.34
[
Cu-MST]
-5.13
1
9

ACCEPTED MANUSCRIPT
Table 9: Results of antimicrobial bioassay of the prepared Schiff base amino acid ligands and their complexes in
DMSO
Comp.
Inhibition Zone (mm)
Fungi
Bacteria
B. subtilis E. coli
15 30 15 30
A. niger C. glabrata
S. cerevisiae
M. luteus
Conc.(mg/ml)
MST
MSTCu
DSP
DSPCu
DSH
15 30
10
12 24
15 30
15 30
15 30
5
3
8
3
9
3
9
7
13
7
14
8
13
5
8
4
15
4
9
24
9
3
9
3
8
20
8
6
11
16 32
11
14 30
10
11 17
4
8
5
8
6
13 24
11 19
13
5
23
9
9
4
18
8
5
9
5
8
6
DSHCu
10 23
11 19
12
19
8
18
12 28
24 42
Ciprofloxacin
Amphotricine B
-
-
-
-
-
-
19
-
33
-
13 28
16 29
11 17
14 23
-
-
-
-
A. niger: Asperagillus niger, C. glabarta: Candida Glabarta, S. cerevisiae: Saccharomyces cerevisiae
B. subtilis: Bacillus subtilis, E. coli: Escherichia coli, M. luteus: Micrococcus luteus
Table 10: Results of activity index (%) for antimicrobial assay of the prepared Schiff base amino acid ligands and their
complexes
Activity index (%)
Comp.
Bacteria
Fungi
B. subtilis
E. coli
M. luteus
A. niger C. glabrata
S. cerevisiae
MST
27.3
72.7
27.3
69.7
27.3
57.6
28.6
71.4
28.6
64.3
28.6
64.3
26.2
76.2
26.2
71.4
23.8
66.7
34.5
82.8
27.6
82.2
31.0
82.8
41.2
76.5
41.2
82.4
47.1
76.5
34.8
73.9
34.8
82.6
34.8
78.3
MSTCu
DSP
DSPCu
DSH
DSHCu
Table 11: Antiviral activities of the prepared complexes
Minimum inhibitory concentration
Minimum cytotoxic
concentration
Compoud
Herpes
simplex
Herpes simplex
virus-1 TK-
VMW1837
Herpes simplex
virus-1 (KOS)
Tobacco Mosaic
Virus
(ꢀg/ml)
virus2- (G)
MSTCu
DSHCu
DSPCu
BVDU
ACG
43
84
102.7
> 400
1.2
86.2
> 400
1.3
38.9
> 400
10.2
0.64
0.64
0.32
16.60
98.30
64
1.45
> 400
> 400
> 100
0.0256
0.0768
0.0038
> 400
0.0768
0.0064
-
-
-
DHPG
2
0

ACCEPTED MANUSCRIPT
Table 12: Spectral parameters for DNA interaction with the prepared complexes
5
*
Complex
λmax free
nm)
620
82
634
80
498
68
λmax bound
ꢂn
Chromism
Type of
10 K
ꢂG
b
a
-1
3
-1
(
(nm)
(nm)
(%)
Chromism mol dm
kJ mol
-28.2
MSTCu
DSPCu
DSHCu
628
380
648
367
485
374
8
2
31.4
38.5
53.8
30.4
73.9
25.9
Hypo
Hypo
Hyper
Hypo
Hyper
Hypo
0.8
1.8
9.4
3
14
13
13
6
-30.2
-22.9
3
3
a
Chromism (%) = (Afree – Abound) / Afree
List of Figures:
3
3
2
1
0
0
.75
.00
.25
.50
.75
MSTCu
DSPCu
DSHCu
.00
2
00
400
600
800
Wavelength (nm)
-3
-3
-3
-3
Figure 1: Molecular electronic spectra of (1) [DSHCu] = 8.0 x 10 mol dm , (2) [DSPCu] = 1.4 x 10 mol dm ,
-3
-3
(
3) [MSTCu] = 5.0 x 10 mol dm .
2
1

ACCEPTED MANUSCRIPT
1.05
0.90
0.75
0.60
MSTCu
MSPCu
DSPCu
0.0 0.5 1.0 1.5 2.0 2.5 3.0
[L] / [M]
2+
Figure 2: Molar ratio plot for the prepared complexes in aqueous-ethanol mixture at [Cu ] = [MST] =
-2
-3
2+
-2
-3
2+
-2
5
.0 x 10 mol dm , [Cu ] = [DSP] = 2.8 x 10 mol dm and [Cu ] = [DSP] = 2.2 x 10 mol
-3
dm and 298 K
1
1
0
0
0
.5
.2
.9
.6
.3
MSTCu
MSPCu
DSTCu
0.30
0.45
0.60
0.75
0.90
[L] / [L] + [M]
2
+
Figure 3: Continuous variation plot for the prepared complexes in aqueous-ethanol mixture at [Cu ] = [DSH] =
-2
-3
2+
-2
-3
2+
-2
-3
2
.5 x 10 mol dm , [Cu ] = [DSP] = 1.7 x 10 mol dm and [Cu ] = [MST] = 5.0 x 10 mol dm
and 298 K.
2
2

ACCEPTED MANUSCRIPT
0.5
0.4
0.3
0.2
0.1
DSTCu
DSHCu
DSPCu
2
4
6
8
10
12
14
pH
-3
-3
-3
Figure 4: pH profile of the prepared complexes at [complex] = 1.0 x 10 mol dm (except [DSHCu] = 5.4 x 10 mol
-3
dm ) and 298 K.
Figure 5: Optimized structures of MST ligand, HOMO-LUMO molecular orbitals with the calculated energy gap
2
3

ACCEPTED MANUSCRIPT
Figure 6 Optimized structures of a) MST ligand and the octahedral Cu complex. b) HOMO-LUMO molecular orbitals
with the calculated energy gap.
Figure 7: Antifungal evaluation of the investigated Schiff base amino acids and their complexes against C. glabrata
fungi.
2
4

ACCEPTED MANUSCRIPT
Figure 8: Antibacterial evaluation of the investigated Schiff base amino acids and their complexes against E. coli
bacteria.
3.6
3.0
2.4
1.8
1.2
0.6
0.0
2
0
MSTCu
15
10
5
0
5
10
15
20
25
30
35
6
-3
1
0 [DNA], mol dm
MSTCu + DNA
300
400
500
600
700
Wavelength (nm)
-3
-3
-3
Figure 9: Spectral scans of the interaction of MSTCu complex (5.0 x 10 mol dm ) in 0.01 mol dm Tris buffer
(
pH 7.2, 298 K) with CT – DNA (from 3 - 30 ꢀm DNA). Plot of [DNA] / (ε - ε ) versus [DNA] for the
a f
titration of DNA with MSTCu complex.
2
5

ACCEPTED MANUSCRIPT
1
1
1
1
1
1
.5
.4
.3
.2
.1
.0
DSHCu
MSTCu
DSPCu
EB
0
.0 0.1 0.2 0.3 0.4 0.5 0.6
[Complex]/ [DNA]
Figure 10: The effect of increasing concentration of the synthesized complexes on the relative viscosities of DNA
at [DNA] = 0.5 mM , [complex] and [EB] = 25 -250 ꢀM and 298K.
Figure 11: DNA binding results of Schiff base amino acid Complexes based on gel electrophoresis. Lane 1: CT –
DNA, Lane 2: CT – DNA+DSHCu, Lane 3: CT – DNA+DSPCu, Lane 4: CT – DNA+MSTCu, Lane 5:
MSTCu.
2
6

ACCEPTED MANUSCRIPT
Highlights for review
1
2
3
- New Cu(II) Amino acid Schiff base chelates were
synthesized and characterized
- The prepared chelates were tested for their antimicrobial
and antiviral activities.
- DNA interaction of the prepared chelates was investigated