Zn(II) coordination compounds derived from 4-acyl pyrazolones and 1,10 phenanthroline: Syntheses, crystal structures, spectral analysis and DNA binding studies

Article abstract of DOI:10.1016/j.poly.2013.07.018

A new series of three Zn(II) coordination complexes [Zn(MCPMPAC) ₂H₂O] = complex 1, [Zn(PMPAC)₂H₂-O] = complex 2 and [Zn(PMPAC)₂(phen)] = complex 3 (MCPMPAC = 4-acyl-3-methyl-1-(3-chlorophenyl) pyrazolone-5-one, PMPAC = 4-acyl-3-methyl-1-phenyl pyrazolone-5-one, Phen=1,10 phenanthroline) has been synthesized and characterized. The structural features of synthesized complexes were determined by metal estimation, molar conductivity, IR, UV-Vis, NMR and single crystal X-ray study. The conductivity data confirm the non-electrolytic nature of the complexes. The single crystal analyses of the complexes show that the Zn(II) ion is five-coordinated with water molecule at axial position in case of 1 and 2 whereas, six-coordinated with phenanthroline ligand in case of 3. Binding of the synthesized complexes with calf thymus DNA (CT-DNA) was studied by spectroscopic methods and viscosity measurements. Experimental results suggest that the zinc complexes have the ability to form adducts with DNA and to distort the double helix by changing the base stacking.

Full text of DOI:10.1016/j.poly.2013.07.018

Accepted Manuscript
Zn(II) coordination compounds derived from 4-acyl pyrazolones and 1, 10 phe‐
nanthroline: Syntheses, crystal structures, spectral analysis and DNA binding
studies
R.N. Jadeja, Mitesh Chhatrola, Vivek K. Gupta
PII:
S0277-5387(13)00543-3
DOI:
http://dx.doi.org/10.1016/j.poly.2013.07.018
POLY 10249
Reference:
To appear in:
Polyhedron
Received Date:
Accepted Date:
20 April 2013
11 July 2013
Please cite this article as: R.N. Jadeja, M. Chhatrola, V.K. Gupta, Zn(II) coordination compounds derived from 4-
acyl pyrazolones and 1, 10 phenanthroline: Syntheses, crystal structures, spectral analysis and DNA binding studies,
Polyhedron (2013), doi: http://dx.doi.org/10.1016/j.poly.2013.07.018
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Zn(II) coordination compounds derived from 4-acyl pyrazolones and 1, 10
phenanthroline: Syntheses, crystal structures, spectral analysis and DNA
binding studies
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R. N. Jadejaa*, Mitesh Chhatrolaa, Vivek K. Guptab
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aDepartment of Chemistry, Faculty of Science, The M. S. University of Baroda, Vadodara 390 002, India
bPost-Graduate Department of Physics & Electronics, University of Jammu, Jammu Tawi 180 006, India
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*E-mail- rajendra_jadeja@yahoo.com, Tel: +91-265-2795552 ext 30
Abstract
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A new series of three Zn(II) coordination complexes [Zn(MCPMPAC)2H2O] =
complex 1, [Zn(PMPAC)2H2O] = complex 2 and [Zn(PMPAC)2(phen)] = complex 3
(MCPMPAC = 4-acyl-3-methyl-1-(3-chlorophenyl) pyrazolone-5-one, PMPAC = 4-acyl-3-
methyl-1-phenyl pyrazolone-5-one, Phen = 1, 10 phenanthroline) has been synthesized and
characterized. The structural features of synthesized complexes were determined by metal
estimation, molar conductivity, IR, UV–Visible, NMR and single crystal X-ray study. The
conductivity data confirm the non-electrolytic nature of the complexes. The single crystal
analyses of the complexes show that the Zn(II) ion is five-coordinated with water molecule at
axial position in case of 1 and 2 whereas, six-coordinated with phenanthroline ligand in case
of 3. Binding of the synthesized complexes with calf thymus DNA (CT-DNA) was studied by
spectroscopic methods and viscosity measurements. Experimental results suggest that the
zinc complexes have the ability to form adducts with DNA and to distort the double helix by
changing the base stacking.
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Keywords: 4-propionyl pyrazolone, Zn(II) complex, Crystal structure, DNA binding study
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1. Introduction
Structural investigation of the binding modes of the ligands in ternary transition metal
complexes is of great interest due to their effect on the topologies and propagation of
extended coordination compounds [1]. This area of research has evolved rapidly in recent
years as the coordination compounds may have interesting properties and applications, e.g.,
DNA Binding, Cleavage and other biological applications [2]. On the other hand, studying
the interactions of metal ions with drugs and biologically active ligands is important to
investigate the potential for synergistic activity between the metal and the drugs as well as
understanding the toxic side effects of synthetic drugs which may, in part, be arising due to
these interactions [3]. Pyrazolone nucleus, which is very useful pharmacological ingredient,
especially to the class of non-steroidal anti-inflammatory drugs (NSAIDs) is used in the
treatment of arthritis and other musculoskeletal joint disorders and ear preparations [4].
Coordination chemistry of 4-acyl pyrazolones has been previously investigated where it has
been used as a blocker for coordination compounds and for synthesis of biologically active
complexes [5]. Their metal complexes have been found to display catalytic performance,
biological activity and photochromic properties. The focus of our research on transition metal
complexes with pyrazolone derivatives is due to the theoretical and practical significance of
these compounds. A number of pyrazolone derivatives show biological activity, as a
consequence, some are commercial products or compounds in the phase of activity
evaluation. As part of our investigations on metal pyrazolone systems, we have previously
examined the ligation behavior of various 4-acyl 3-methyl-1-phenyl-pyrazolone-5-one and
their derivatives with different metal ions like Ca, V, Cr, Mn, Fe, Ni, Cu [6-12].
In this work, we have extended our study to the synthesis of binary and mixed-ligand
complexes of Zn(II) metal with 4-propionyl-pyrazol-5-one ligands and chelating
heteroaromatic N-donor ligand 1, 10 phenanthroline. We here in report the synthesis,
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characterization and crystal structures of three zinc acyl pyrazolone complexes, in which the
components are 4-propionyl-pyrazolone ligands and 1, 10 phenanthroline (phen).
2. Experimental
2.1. Materials
The reagents and chemicals of analytical reagent grade were procured from
commercial sources. Solvents used for electrochemical and spectroscopic studies were
purified using standard procedures [13]. 1, 4 dioxane was obtained from E. Merck (India)
Ltd. Calcium hydroxide, zinc acetate and 1, 10 phenanthroline were obtained from LOBA
Chem. Pvt. Ltd., Mumbai and used as supplied. Absolute alcohol was obtained from Baroda
Chem. Industry Ltd. and was used after distillation. Methanol was obtained from
Spectrochem. Mumbai, India and was used after distillation. CT-DNA (Calf Thymus DNA)
was purchased from Sigma Aldrich. Ethidium bromide (EB) was obtained from Hi-media
laboratories Pvt. Ltd., Mumbai.
2.2. Methods
The synthesized compounds were characterized using different techniques.
Infrared
spectra (4000-400 cm-1, KBr discs) of the samples were recorded on a model RX 1 FTIR
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1H NMR spectra of the ligands were recorded with Bruker
Perkin-Elmer spectrophotometer.
AV 400 MHz using CDCl3 as a solvent and TMS as an internal reference. 1H NMR spectra of
1
complex 3 was recorded with Bruker AV 400 MHz using CDCl3 as a solvent, H NMR
spectra of complexes 1 and 2 were recorded with Bruker AV 400 MHz using DMSO as a
solvent and
Mass spectra of the ligands were recorded on Trace GC ultra DSQ II.
nm were
The electronic spectra (in DMF at room temperature) in the range of 400-800
recorded on a model Perkin Elmer Lambda 35 UV-VIS spectrophotometer. Fluorescence
spectra were recorded on a model JASCO, FP-6300 fluorescence spectrophotometer. Molar
conductivity of 1 0 - 3 M solution o f the complexes in DMF was measured at room
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temperature with a model Elico CM 180 digital direct reading deluxe digital conductivity
meter. Zinc content was determined by gravimetric analysis after decomposing the complexes
with HNO3.
2.3. Synthesis of ligands
2.3.1. 4-propionyl-3-methyl-1-phenyl pyrazolone-5-one (PMPAC)
3-methyl-1-phenyl pyrazolone-5-one (0.1 mol, 17.4gm) was dissolved in hot dioxane
(80 cm3) in a flask equipped with a stirrer, separating funnel and reflux condenser. Calcium
hydroxide (0.2 mol, 14.81gm) was added to this solution, followed by acetyl chloride (0.1
mol, 8.684 cm3) added drop wise with precaution, as this reaction was exothermic. During
this addition the whole mass was converted into a thick paste. After the complete addition,
the reaction mixture was refluxed for half an hour and then it was poured into dilute
hydrochloric acid (2 M, 200 cm3). The colored crystals (PMPAC) thus obtained were
separated by filtration and recrystallized from an acidified methanol–water mixture
(HCl:MeOH:H2O = 1:80:19). M.P. 90ºC, Yield 80.85 %, Molecular formula:C13H14N2O2
(calc.M.W. 230.26); Mass: m/z = 230.02 [C13H14N2O2, MIP]+, 231.26 [C13H14N2O2 (m+1)
Peak]+, 200.62 [C11H8N2O2 Base peak]+, 201.31 [C11H9N2O2]+, 82.70 [C3H3N2O]+, 137.20
[C7H10N2O]+, 68.90 [C5H5]+, 56.99 [C4H8]+; IR (KBr, cm-1): 1563(s) (C=N, cyclic), 1635 (m)
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(C=O, Pyrazolone ring), 1651 (m) (C=O, Propionyl group); H NMR (CDCl3): 1.24-1.28 (t)
(3H, Propionyl C-CH3), 2.48 (s) (3H, Pyrazolone C-CH3), 2.77-2.82 (q) (2H Propionyl-CH2-
), 7.27-7.85 (m) (5H Phenyl), 11.695 (s) (1H, Phenolic –OH).
2.3.2. 4-propionyl-3-methyl-1-(3-chlorophenyl) pyrazolone-5-one (MCPMPAC)
It was analogously prepared from 3-methyl-1-(3-chlorophenyl) pyrazolone-5-one. The
colored crystals thus obtained were separated by filtration and recrystallized from an acidified
methanol–water mixture (HCl: MeOH: H2O = 1:80:19). M.P. 86°C, Yield 69.6 %, Molecular
formula C13H13ClN2O2 (calc.M.W. 264.70); Mass: m/z = 264.08 [C13H13ClN2O2, MIP]+,
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266.08 [C13H13ClN2O2 (m+2) Peak]+, 263 [C13H13ClN2O2 (m-1) Peak]+, 236.15
[C11H9ClN2O2]+, 85.07 [C3H4N2O]+, 96.81 [C4H4N2O]+, 68.93 [C5H8]+, 219.21
[C12H14N2O2]+, 56.92 [C4H8, Base peak]+, 125.15 [C6H8N2O]+, 42.96 [C3H6]+; IR (KBr, cm-
1): 1563(s) (C=N, cyclic), 1634 (m) (C=O, Pyrazolone ring), 1651 (m) (C=O, Propionyl
group); 1H NMR (CDCl3): 1.25-1.29 (t) (3H, Propionyl C-CH3), 2.48 (s) (3H, Pyrazolone C-
CH3), 2.77-2.82 (q) (2H, Propionyl –CH2-), 7.24-7.93 (m) (4H, Phenyl group), 11.65 (s) (1H,
Phenolic –OH).
Synthesis of ligands is summarized in scheme 1.
2.4. Syntheses of Zn(II) complexes
2.4.1. Synthesis of 1
The complex was synthesized by the following method. The metal salt Zinc-acetate
[Zn(CH3COO)2.2H2O] (0.219 gm, 0.001 mmol) was dissolved in methanol and the solution
was added to a cool methanolic solution of the ligand (MCPMPAC) (0.528gm, 0.002 mmol).
After complete addition, the mixture was refluxed about for 4 hours. The reaction mixture
was then filtered and was washed with Et2O and water and dried in air. It was recrystallized
from DMF at RT. M.P. 210’C, Yield 79.2 %, Molecular formula: C26H26Cl2N4O5Zn
(calc.M.W. 632.87); IR (KBr, cm-1): 3109 (co-ordinated H2O), 1615 (C=O Propionyl group),
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1555 (C=N cyclic), 1485 (C-O, pyrazolone ring), 511 (Zn-O); H NMR (CDCl3): 1.063-
1.026 (t) (6 H, C-CH3 Propionyl group), 2.703-2.648 (q) (4H,-CH2-CH3 Propionyl group ),
2.300 (s) (6H, -CH3 methyl group), Benzene ring 7.140-7.121 (d) (2H), 7.362-7.322 (t) (2H)
7.882-7.861 (d) (2H), 8.161 (s) (2H).
2.4.2. Synthesis of 2
It was analogously prepared from Zn(CH3COO)2.2H2O (0.219 gm, 0.001 mmol) and
PMPAC (0.460gm, 0.002 mmol). It was recrystallized from DMF at RT. M.P.170-175’C,
Yield. 69.33 %, Molecular formula: C26H32N4O5Zn (calc.M.W. 545.965); IR (KBr, cm-1):
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3060 (co-ordinated H2O), 1615 (C=O Propionyl group), 1505 (C=N cyclic), 1416 (C-O bond
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pyrazolone ring), 510 (Zn-O); H NMR (CDCl3): 1.035-1.072 (t) (3 H, C-CH3 Propionyl
group), 2.634-2.689 (q) (2H,-CH2-CH3 Propionyl group ), 2.317 (s) (3H, -CH3 methyl group),
Benzene ring 7.971-7.951 (d) (2H), 7.346-7.307 (t) (2H) 7.123-7.086 (t) (1H).
2.4.3. Synthesis of 3
The complex was synthesized by the following method. The metal salt Zinc- acetate
(Zn(CH3COO)2.2H2O) (0.219 gm, 0.001 mmol) was dissolved in methanol and the solution
was added to a hot methanolic solution of the ligand (PMPAC) (0.460gm, 0.002 mmol) and
1, 10 phenanthroline (0.198 gm, 0.001 mmol). After complete addition, little amount of
sodium acetate was added and the mixture was refluxed about for 5 hours and after some time
crystalline solid was obtained. The mixture was then filtered and washed with hot water and
dried in air. It was recrystallized from DMF. M.P. 230ºC, Yield 61.42%, Molecular formula:
C38H38N6O4Zn (cacl.M.W. 708.15); IR (KBr, cm-1): 1635 (m) (C=O Propionyl group), 1555
(s) (C=N cyc lic), 1485 (C-O, pyrazolone ring), 511 (Zn-O), 422 (Zn-N); 1H NMR (CDCl3):
1.003-0.965 (t) (3H, C-CH3 Propionyl group), 2.611-2.556 (q) (2H,-CH2-CH3 Propionyl
group), 2.367 (s) (3H, -CH3 methyl group), Benzene ring 7.226-7.188 (t) (2H), 7.047-7.029
(t) (1H), 9.247-9.239 (d) (1H), 8.479-8.454 (d) (1H), Phenanthroline ring 7.953-7.846 (m)
(4H).
Synthesis of complexes is summarized in scheme 2.
2.5. Crystallography
Crystals having good morphology were chosen for three-dimensional intensity data
collection. X-ray intensity data of the complexes were collected at room temperature on
Bruker CCD area-detector diffractometer equipped with graphite monochromated MoKα
radiation (α=0.71073 Å). The crystals used for data collection was of suitable dimensions
0.30x0.20 x0.20 mm. The unit cell parameters were determined by least-squares refinement.
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Data were corrected for Lorentz, polarization and multi-scan absorption correction [14]. The
structures were solved by direct methods using SHELXS97 [15]. All non-hydrogen atoms of
the molecules were located in the best E-map. Full-matrix least-squares refinement was
carried out using SHELXL97 [15]. Hydrogen atoms were placed at geometrically fixed
positions and allowed to ride on the corresponding non-H atoms with C-H = 0.93-0.96Å, and
Uiso=1.5 Ueq of the attached C atom for methyl H atoms and 1.2 Ueq for other H atoms. An
ORTEP [16] view of the complexes with atomic labeling is shown in Fig. 1. The geometry of
the molecules has been calculated using the software PLATON [17] and PARST [18]. The
crystallographic data for the complexes are summarized in Table 1. The important bond
lengths and bond angles of the complexes are listed in Table 2.
2.6. DNA binding
All of the experiments involving the binding of complexes with CT-DNA were
carried out in double distilled water with trisodium citrate (Tris, 15 mM) and sodium chloride
(150 mM) and adjusted to pH 7.05 with hydrochloric acid. The DMF solution of the complex
was used throughout the study. The concentration of CT-DNA per nucleotide was estimated
from its known extinction coefficient at 260 nm (6600 M-1 cm-1) [19]. Solutions of CT-DNA
in tris buffer gave a ratio of UV absorbance at 260 and 280 nm (A260/A280) 1.8-1.9 indicating
that the DNA was sufficiently free of protein. Absorption titration experiments were
performed by maintaining a constant metal complex concentration (5 μM), while gradually
increasing the concentration of DNA (0−75 μM). While measuring the absorption spectra, an
equal amount of DNA was added to both the test solution and the reference solution to
eliminate the absorbance of DNA itself.
The data were then fit to eq 1 [20] to obtain intrinsic binding constant Kb.
…… (1)
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Where, [DNA] is the concentration of DNA in base pairs,
is the extinction
a
coefficient observed for the MLCT absorption band at the given DNA concentration, f is the
extinction coefficient of the complex free in solution, and b is the extinction coefficient of the
complex when fully bound to DNA. A plot of [DNA]/[ - ] versus [DNA] gave a slope 1/[ -
a
f
a
f] and Y intercept equal to (1/Kb)[ - ], respectively. The intrinsic binding constant Kb is the
b
f
ration of slope to intercept [20].
Competitive studies of compound with ethidium bromide (EB) have been investigated
with fluorescence spectroscopy in order to examine whether the compound can displace EB
from its CT DNA−EB complex. The CT DNA−EB complex was prepared by adding 3.3 μM
EB and 4.2 μM CT-DNA in buffer (150 mM NaCl and 15 mM trisodium citrate at pH 7.05).
The intercalating effect of the compound with the DNA−EB complex was studied by adding
a certain amount of a solution of the complex step by step (0-370 µM) into the solution of the
DNA−EB complex. The influence of the addition of each complex to the DNA−EB complex
solution has been obtained by recording the variation of fluorescence emission spectra. The
emission spectra were monitored by keeping the excitation of the test compound at 546 nm
and the emission was monitored in the range of 550-750 nm. The emission was observed at
610 nm.
Commonly, fluorescence quenching can be described by the following Stern−Volmer
equation (eq 2) [21].
…… (2)
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Where F0 and F are the steady-state fluorescence intensities in the absence and
presence of quencher, respectively, Ksv is the Stern-Volmer quenching constant, obtained
from the slope of the plot F0/F versus [Complex] and [Q] is the total concentration of
quencher.
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Viscosity experiments were carried out by using an auto viscometer (SCHOOT AVS
◦
350), immersed in a thermostated water bath with the temperature setting at 30 1 C for 15
min. DNA samples with an approximate average length of 200 base pairs were prepared by
sonication in order to minimize complexities arising from DNA flexibility [22]. The
concentration changes of the Zn(II) complexes were realized by adding different volumes of
Zn(II) complex stock solution. Flow time was measured with a digital stopwatch, and each
sample was measured triply, and an average flow time was obtained. Data were presented as
(η/η ) 1/3 versus the mole ratio of Zn(II) complex to DNA. Where η is the viscosity of DNA in
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the presence of complex, and η is the viscosity of DNA alone.
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3. Results and discussion
The pyrazolone derivatives were prepared by refluxing an appropriate amount of
respective 3-methyl-pyrazolone-5-one with the Propionyl chloride. The structures of the
synthesized ligands were established with the help of IR, NMR, and Mass spectra. All
spectral data are agreed well with the acyl pyrazolone ligand structures. All Zn(II) complexes
were prepared by using the metal salt with the corresponding ligands in molar ratio of
metal:ligand as 1:2 as obtainable in the following reaction.
M·(CH3COO)2·nH2O + 2L → ML2 + nH2O + nCH3COOH
where, M= Zn(II)
All these complexes are colorless, air and moisture free amorphous solids. They are
insoluble in common organic solvents and only soluble in DMF and DMSO. Molar
conductance values of the soluble complexes in DMF (10-3M solution at 25 ◦C) indicate that
the complexes have molar ratio of metal:ligand as 1:2. The lesser molar conductance values
7.00 for 1, 5.00 for 2 and 15.00 for 3 ohm−1 cm2 mol−1 indicate that they are all
nonelectrolytic in nature [23]. The elemental analyses data concur well with the planned
formulae and also recognized the [ML2 (H2O)] composition for 1 and 2 complexes and [ML2
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(Phen)] composition for 3 complex. All the complexes were characterized by FT-IR, NMR,
Single crystal analyses and UV–Visible spectroscopic techniques. Analytical data of the
synthesized complexes and ligands are given in section 2.
3.1. Spectroscopy
The IR spectral studies provide valuable information regarding the coordinating sites
of ligand. The IR spectra of the complexes were compared with that of the free ligand to
determine the changes that might have taken place during the complexation. A comparative
study of the IR spectra of ligands and its metal complexes reveals that certain bands are
common and therefore, only important bands, which have been either shifted or newly
appeared, are discussed. In the free ligand, a medium-intensity band at 3011cm-1 which
assigned to enol ν(OH) of β-diketone tautomer is absent in complex. A band at 1626 cm-1 in
the free ligand is allocated as ν(C=O) of pyrazolone-ring transfers to 1485 cm-1 in the
complex [24]. The band at 1651cm-1 in Propionyl group changes to 1615 cm-1 in the
complex. IR spectral evidence, therefore, suggests that the enolic proton of ligand is replaced
by Zn(II) in the complex. FT-IR spectra of ligands exhibits the bands within the range 1550–
1565 cm-1 which can be assign to v(C=N) (cyclic). The complexes show the absorption
within 3000–3150 cm-1 which can be assigned to the coordinated water molecules. The
complexes show bands at 510 cm-1 and 422 cm-1 which are due to the v(Zn-O) and v(Zn-N),
respectively [25]. The IR spectral data are shown in section 2.
The 1H spectra of both the ligands were recorded in CDCl3 at room temperature. The
signals due to one methyl group appeared as singlet in the range 2.38–2.47 ppm. In the
aromatic region, a few doublets and in few cases some overlapping doublets/multiplets are
observed in the range 7.25–7.99 ppm in all the ligands. These doublets/multiplets are due to
aryl protons of benzene ring. However, a singlet at 11.695 ppm may be attributed to the
1
phenolic -OH of the acyl pyrazolone ligand. The H NMR spectral data are presented in
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section 2. The H NMR spectrum of ligand PMPAC is shown in Fig. S6. The H NMR
spectrum of Zn(II) complexes of 1 and 2 were recorded in deuterated DMSO, and 1H NMR
spectrum of Zn(II) complexes of 3 was recorded in CDCl3 at room temperature shows signals
consistent with the proposed structure. The multiplets around 7.0–7.9 ppm are assigned to
aromatic protons, and propionyl protons of –CH2- group at 2.556-2.611(q) ppm. The peak for
phenolic –OH of the acylpyrazolone ligand is absent in the zinc(II) complexes which
confirms deprotonation on complexation. The 1H NMR spectral data are presented in section
2. The 1H NMR spectrum of complex 3 is shown in Fig. S10.
In the 13C NMR spectra of the ligands, the carbon atoms of the methyl groups appear
in the range 15.72–21.02 ppm. The carbon atoms of the one benzene rings exhibit signals in
the range 118.13–147.76 ppm. Eleven signals were observed for ligand PMPAC. Some of the
signals might be overlapped as indicated by the intensity of a few signals. The other ligand
MCPMPAC, where one of the benzene rings is m-substituted, show thirteen signals, as
expected because of the loss of symmetry. All the protons and carbons were found as to be in
their expected region. The 13C NMR spectral data are shown in section 2. The 13C NMR
spectrum of ligand PMPAC is shown in Fig. S11.
The mass spectra of both bidentate pyrazolone ligands are in good agreement with
proposed structures. The mass spectra of synthesized ligands were recorded and the obtained
molecular ion peaks confirmed the proposed formula. Melting point of each ligand is high, as
a result of this; the mass spectra were carried out by EI. The electronic impact mass spectra of
PMPAC shows a molecular ion peak at m/z = 229.84 [C13H14N2O2]+ with a relative intensity
near to 100%, which is equivalent to its molecular weight. The electronic impact mass spectra
of MCPMPAC shows a molecular ion peak at m/z = 264.34 [C13H13ClN2O2]+ with a relative
intensity near to 40.98%. The other peaks appeared in the mass spectra (abundance range 1–
100%) are attributed to the fragmentation of ligand molecules obtained from the rupture of
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different bonds inside the molecule. The mass spectral assignments for both the ligands are
shown in the section 2. The mass spectral data of ligand PMPAC is shown in Fig. S13.
3.2. Crystal structure description
3.2.1. Complex 1
The molecular structure and the atom labeling scheme of the complex 1 is shown in
Fig. 1. The main bond distances and angles are listed in Table 2.
As shown in Fig. 1, the Zn(II) ion is pentacoordinated by four oxygen atoms of two
acyl pyrazolone ligands and one oxygen atom of water molecule. The two oxygen donors of
both pyrazolone molecules occupy the basal sites of the pyramidal structure, whereas the
water molecule binds the zinc at the apical site. In this complex, the incorporation of the
solvent into the coordination sphere increases the coordination number to five and the
geometry around the metal is now distorted trigonal bipyramidal.
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In the complex, the zinc atom occupies a distorted trigonal bipyramidal environment
(TBP), formed by four oxygen atoms [O(1), O(2), O(1_i), O(2_i)] supplied by two
pyrazolone ligands (Symmetry code: (i) -x, y, 0.5-z). The fifth coordination site is occupied
by one oxygen atom of water molecule. The Zn–O distances of the basal plan are Zn(1)-
O(1)= 1.9934(16) Å and Zn(1)-O(2)= 2.0158(17) Å, whereas the axial distance of Zn(1)-O(3)
is 2.011(3)Å. The longest distance does not correspond to the terminal Zn(1)-O(3), but
instead to one of the basal Zn(1)-O(2) distance of the pyrazolone molecule.
All atoms of the chelate and pyrazolone rings lie almost in the same plane. One water
molecule connects with the neighbouring pyrazolone-ring N(1) through intermolecular
hydrogen bonds [O(3)–H(3)……N(1_i)]. H-bonding interactions play an important role in
forming the supramolecular structure by self-assembly and stabilizing. The Hydrogen
bonding geometry parameters of the complex are shown in Table 3.
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In this complex the chlorine molecules are disordered at two positions. At C(8),
occupancy of H(8) is 0.06 and Cl(1A) 0.94 (total 1). Similarly at C(10) occupancy of H(10) is
0.94 and Cl(1B), 0.06 (total 1). We have taken whole molecule except chlorine as part 1 and
chlorine as part 2.
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3.2.2. Complex 2
The structure of the complex 2 is illustrated in Fig. 1. Selected bond lengths
and bond angles are listed in Table 2. Complex 2 is five-coordinate Zn(II) with one water
molecule and four oxygens of two bidentate acylpyrazolonates. The coordination geometry
may be described as slightly distorted trigonal bipyramidal. Zn exists in the location of the
centre of symmetry. Four oxygens [O(16), O(16_i), O(17) and O(17_i)] from two PMPAC-
composed equatorial planes, while one water molecule is axial. The angle of O(17)–Zn(1)–
O(17_i) is 174.91(9)°. The angles of the O(17)–Zn(1)–O(16), O(17_i)–Zn(1)–O(16),
O(16_i)–Zn(1)–O(16), O(17)–Zn(1)–O(1) and O(16)–Zn(1)–O(1) are 89.83(6)º, 88.43(6)º,
139.98(10)º, 92.55(4)º and 110.01(5)º, respectively (Symmetry code: (i) -x, y, 0.5-z).
Moreover, the C(3)–N(2) bond length is close to the C–N double bond length, confirming
that the keto form of the ligand tautomerizes to the enol form.
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Zn(II) and the coordinated oxygens constitute two six-chelate rings which have the
boat configuration. All atoms of the chelate and pyrazolone rings lie almost in the same
plane. One water molecule connects with the neighbouring pyrazolone-ring N(2) through
intermolecular hydrogen bonds O(1)–H(1)……N(2_i). H-bonding interactions play an
important role in forming the supramolecular structure by self-assembly and stabilizing. The
Hydrogen bonding geometry parameters of the complex are also shown in Table 3.
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3.2.3. Complex 3
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The molecular structure of complex 3 together with the atom-numbering scheme is
illustrated in Fig. 1. Important bond lengths and angles are listed in Table 2. The X-ray
analysis revealed that the complex was a 6-coordinate mononuclear Zn(II) complex with two
nitrogen atoms of the phenanthroline molecule and four oxygen atoms of two bidentate
pyrazolonates. Taking into consideration that it usually adopts an octahedral conformation
because of the large ligand field stabilization energy, this configuration is quite ideal. The
coordination plane around Zn(1) ion is composed of O(1), O(2) and N(16) atoms with normal
bond distances of Zn(1)–O(1): 2.1123(14)Å, Zn(1)–O(2): 2.0347(13)Å and Zn(1)–N(16):
2.1908(16)Å. In the equatorial plane, the trans angles in O(1)–Zn(1)–O(1_i) and O(2)–Zn(1)–
N(16) are around 180º, while 90.35(6)º for O(2)–Zn(1)–O(1_i), 91.45(6)º for O(2_i)–Zn(1)–
N(16), 97.70(6)º for O(1)–Zn(1)–N(16_i) and 84.86(6)º for the angle O(1)–Zn(1)–N(16), add
up to 364.36(6)º. Bond angles show that the coordination geometry around the zinc ion in the
complex is distorted octahedral. Atoms O(1), O(2), O(1_i) and N(16) of the ligand molecules
occupy the equatorial positions around the metal centre. Atoms N(16_i) and O(2_i) of the
ligands occupy axial positions creating a octahedral geometry around the central metal.
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In the complex, the bond length of O(2)–C(3) and O(1)–C(13) are 1.272(2) and
1.245(3) Å, respectively, which are shorter than 1.43 Å for a C–O single bond and longer
than 1.22 Å for a C=O double bond length. Moreover, the C(3)–N(2) bond length is close to
the C–N double bond length, confirming that the keto form of the ligand tautomerizes to the
enol form. All atoms of the complex and the pyrazolone rings are not in the same plane. As a
result, the complex is a non-planar molecule. As this complex has no solvent molecule, H-
bonding is absent in this complex. In Complex 3 also, methyl group is disordered over two
sets of sites in a 0.75(2):0.25(2) ratio.
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3.3. DNA binding studies
Many workers [26, 27] were of the opinion that the two major binding modes of
interaction of substrate with DNA were (a) the intercalation: wherein the metal complexes
squeeze in between the double helix through hydrogen bonding and (b) the covalent binding:
where two major sites are available for the metal ion to interact with the DNA, one being the
electron donor groups of the bases, more preferably at the guanine N-7 and the other the
phosphate moieties of the ribose phosphate backbone.
3.3.1. Electronic absorption titration
The application of electronic absorption spectroscopy in DNA binding studies is one
of the most useful techniques carried out at 25 2ºC. The binding of the metal complexes to
DNA helix is often characterized through absorption spectral titration, followed by the
changes in the absorbance and shift in the wavelength. The electronic spectra of Zn(II)
complexes titrated with DNA are given in Fig. 2. Hyperchromism and hypochromism are the
spectral changes typical of a metal complexes association with the DNA helix [20].
Absorption titration experiments were performed by maintaining the metal complex
concentration as constant at 5 μM while varying the concentration of the DNA within 0–75
µM. Upon the addition of DNA, interesting changes in the absorbencies of the d–d transition
absorption bands of the complexes were observed. The observed hyperchromism for all the
complexes unambiguously revealed the active participation of pyrazolone moieties in
association with the DNA. However, the lack of red shift suggests that the binding mode of
both the complexes was not classical intercalation. Because of the bulky structure of the
complexes, the aromatic rings cannot completely intercalate. Therefore, the observed spectral
changes were rationalized in terms of partial intercalation.
To further illustrate the DNA binding strength, the intrinsic binding constant Kb was
determined from the non-linear DNA/Ea-Ef vs [DNA] for all three complexes (1, 2 & 3) and
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they were found to be 1.24 × 105 M−1, 0.52 ×105 M−1 and 0.88 × 105 M-1, respectively.
Binding constants of these complexes were lower in comparison to those observed for typical
classical intercalators (ethidium–DNA, 1.4 × 106M−1) [21]. The diminution of the intrinsic
binding constant s could be explained by the steric constraints imposed by the ligand
framework and thus encouraging a partial intercalation binding mode for the complexes. Our
results are consistent with earlier reports on preferential binding to DNA in the Zn complexes
[28].
Fluorescence spectral technique using the emission intensity of ethidium bromide
(EB) bound to DNA has been used to determine the binding propensity of the complexes. No
luminescence was observed for the complex solution, either with or without the presence of
DNA. So, the binding of complex and DNA cannot be directly presented in the emission
spectra. Ethidium bromide (EB) is a weak fluorophore, but its emission intensity in the
presence of DNA can be greatly enhanced because of its strong intercalation between the
adjacent DNA base pairs. EB, a planar aromatic heterocyclic dye intercalates non specifically
into the DNA which causes it to fluoresce strongly.
If the complexes can intercalate into DNA, the binding sites of DNA available for
EB will be decreased, and hence the fluorescence intensity of EB will be quenched [29]. In
our experiments, as depicted in Fig. 3 for complexes 1, 2 and 3, the fluorescence intensity of
EB show a remarkable decreasing trend with the increasing concentration of the complexes,
indicating that some EB molecules are released from EB-DNA complex after an exchange
with the complexes, which result in the fluorescence quenching of EB. This may be due eiter
to the metal complex competing with EB for the DNA-binding sites thus displacing the EB
(whose fluorescence is enhanced upon DNA binding) or it should be a more direct quenching
interaction on the DNA itself. We assume the reduction of the emission intensity of EB on
increasing the complex concentration could be caused due to the displacement of the DNA
16

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bound EB by the Zn(II) complexes. Such a quenched fluorescence behavior of EB bound to
DNA caused by the interaction between Zn(II) complexes and DNA is also found in other
zinc complexes [30].
The fluorescence quenching curve of EB bound to DNA by the Zn (II) complexes
are shown in Fig. 4. The ratio of the slope to the intercept obtained by plotting F0/F versus
[Complex] yielded the value of Ksv corresponding to the three complexes as 3.21×103 M-1,
0.86×103 M-1 and 0.91×103 M-1, respectively. Values of Ksv suggested that the complex 1
showed higher quenching efficiency than the other complexes 2 and 3. Further, the figures
also show that the ratio of quenching of the intensities in all three complexes is different,
reflecting more binding of complex 1 with DNA to leach out more number of EB molecules
originally bound to DNA than that of the complexes 2 and 3. Our results are consistent with
earlier reports on preferential binding to DNA in the Zn(II) complexes [31–32]. All these
results showed clearly that the complex 1 possesses strong tendency to bind with DNA which
is consistent with the viscosity results.
In addition to spectroscopic titrations, viscosity measurements were carried out to
provide further information on the nature of the interaction between the complex and DNA.
A classical intercalation model demands that the DNA helix lengthens as base pairs are
separated to accommodate the bound ligand leading to an increase of the DNA viscosity. In
contrast, semi-intercalation of a ligand could bend or kink the DNA helix, and thus reduce its
effective length and, concomitantly, its viscosity. A classical intercalation mode causes a
significant increase in viscosity of DNA due to an increase in separation of base pairs at
intercalation sites and hence an increase in overall DNA length [33].
The changes in the relative viscosity of rod-like DNA in the presence of Zn(II)
complexes are shown in Fig. 5. The viscosity of DNA increases greatly with increasing
concentration of complex, which is similar to that of the proven intercalator EB [34]. This
17

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observation suggests that the principal mode of DNA binding by complexes involve base-pair
intercalation, with one ligand intercalating into the base pairs and the other ligand being left
outside the helix [35]. The results obtained in this study are consistence with similar studies
done by others [36-38]. The intercalative interaction with DNA is related to the molecular
structure, as in these complexes there is a little distorted plane that may lead to the weak
intercalative mode. This result also parallels the pronounced emission enhancement of the
complexes, and is comparable with the proven classical intercalator EB. On the basis of the
viscosity results, the complexes bind with DNA through the intercalation mode [7].
Conclusion
The 4-acyl pyrazolone ligands have been prepared by the acylation of pyrazolone
derivatives with propionyl chloride. They have been characterized by IR, 1H NMR, 13C NMR
and mass spectrometry. Binary and mixed-ligand Zn(II) complexes of these ligands have
been synthesized and characterized by metal estimation, molar conductance, IR, 1H NMR and
single crystal X-ray study. The data shows that they have composition of the type [ZnL2.H2O]
and [ZnL2(Phen)]. The X-ray analyses and NMR spectra of the complexes show that the
Zn(II) ion center in [ZnL2(Phen)] is six-coordinated while the Zn(II) ion center in [ZnL2.H2O]
is five-coordinated. The lower molar conductance values of the complexes reveals that these
complexes are non electrolytes. The DNA-binding properties of the synthesized complexes
have been examined by absorption spectroscopy, fluorescence spectroscopy and viscosity
measurements. Evidences have suggested that the complexes could interact with DNA via
partial intercalative mode. Furthermore, in the EB competition absorption titration assay, the
binding constants for the complexes, Ksv are obtained, which are 3.21×103 M-1 for 1,
0.86×103 M-1 for 2 and 0.91×103 M-1 for 3, respectively. The results indicate that complex 1
has a greater DNA affinity than 2 and 3. Moreover, complex 1 can strongly bind to DNA
through intercalation, while 2 and 3 binds to DNA in a partial intercalative mode.
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Acknowledgement
The authors are thankful to University Grants Commission, New Delhi for the
financial assistance to this work in terms of major research project [Project F.37-376/2009
12.01.2010(SR)] to RNJ. They are also thankful Head, Department to of Chemistry for
providing necessary facilities required carrying out this work.
Supporting information
CIF files for the X-ray crystal structures have been deposited with the
Cambridge Crystallographic Data Center (CCDC, 931119-1, 910379-2, 929473-3). Copies of
this information can be obtained free of charge at www.ccdc.cam.ac.uk/conts/retrievel.html
or from the CCDC, 12 Union Road, Cambridge CB2 1EZ, UK; fax : +44-1223/336-033.
email: deposit@ccdc.ac.uk.
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523
Where R = H, Cl
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526
527
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529
530
Scheme 1. Synthesis of acyl Pyrazolone ligands
23

531
532
Where, (1):R = Cl (2):R = H
C H
2
5
C H
2
5
N
N
H C
3
H C
3
CH
3
O
O
O
N
N
O
Zn
Methanol
N
O
N
O
Zinc acetate
C H
2
5
1,10 phenanthroline
N
N
Reflux at 60-70 'C For 6 h.
[CH COONa]
3
533
534
Scheme 2. Synthesis of complexes
535
536
537
538
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539
540
(A)
(B)
541
542
(C)
543
544
545
546
Fig. 1. ORTEP view of the Complex 1(A), Complex 2(B) and Complex 3(C) showing atom-
labelling scheme. Displacement ellipsoids are drawn at the 50% probability level and H
atoms are shown as small spheres of arbitrary radii.
547
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548
549
550
551
Fig. 2. Electronic spectra of complexes (1) Complex 1, (2) Complex 2 and (3) Complex 3 in
DMF in the absence and presence of CT-DNA. Arrow shows the absorbance changes upon
increasing DNA concentrations.
552
553
554
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555
556
557
558
559
560
Fig. 3. Emission spectra of EB bound to DNA in the absence and presence of complexes (1)
Complex 1, (2) Complex 2 and (3) Complex 3. The arrow shows the intensity changes on
increasing the complex concentration.
27

561
562
Fig. 4. Fluorescence quenching curve of EB bound to DNA by (1) Complex 1, (2) Complex 2
and (3) Complex 3.
563
564
565
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566
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568
569
570
571
572
573
574
575
576
577
578
579
580
581
Fig. 5. Effect of increasing amounts of (1) Complex 1, (2) Complex 2 and (3) Complex 3 on
the relative viscosities of CT-DNA at 30.0 0.1 ºC. [DNA] = 3 mM, [Complex]/[DNA] = 0,
0.0015, 0.0031, 0.0047, 0.0062, 0.0078, 0.0094 respectively.
29