Pyrazole-based azo-metal(II) complexes as potential bioactive agents: synthesis, characterization, antimicrobial, anti-tuberculosis, and DNA interaction studies

Article abstract of DOI:10.1080/00958972.2019.1630613

In the present investigation, the bioactive azo-dye ligand 1,5-dimethyl-4-[(3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl)diazenyl]-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (L) and its transition metal complexes have been synthesized and characterized

Full text of DOI:10.1080/00958972.2019.1630613

Journal of Coordination Chemistry
ISSN: 0095-8972 (Print) 1029-0389 (Online) Journal homepage: https://www.tandfonline.com/loi/gcoo20
Pyrazole-based azo-metal(II) complexes
as potential bioactive agents: synthesis,
characterization, antimicrobial, anti-tuberculosis,
and DNA interaction studies
Mallikarjuna Niluvanji Matada & Keshavayya Jathi
To cite this article: Mallikarjuna Niluvanji Matada & Keshavayya Jathi (2019): Pyrazole-based
azo-metal(II) complexes as potential bioactive agents: synthesis, characterization, antimicrobial,
anti-tuberculosis, and DNA interaction studies, Journal of Coordination Chemistry, DOI:
10.1080/00958972.2019.1630613
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JOURNAL OF COORDINATION CHEMISTRY
https://doi.org/10.1080/00958972.2019.1630613
Pyrazole-based azo-metal(II) complexes as potential
bioactive agents: synthesis, characterization, antimicrobial,
anti-tuberculosis, and DNA interaction studies
Mallikarjuna Niluvanji Matada and Keshavayya Jathi
Department of PG Studies and Research in Chemistry, School of Chemical Sciences, Kuvempu
University, Jnana Sahyadri, Shankaraghatta, Karnataka, India
ABSTRACT
ARTICLE HISTORY
Received 29 January 2019
Accepted 26 May 2019
In the present investigation, the bioactive azo-dye ligand
1,5-dimethyl-4-[(3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-
yl)diazenyl]-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (L) and its tran-
sition metal complexes have been synthesized and characterized by
various physical and spectroscopic techniques to elucidate their
geometrical structures. The molar conductivity measurements con-
firmed the non-electrolytic nature of the complexes. EPR spectros-
copy indicated that the metal complexes are monomeric in nature
and exhibited octahedral geometry. The redox behavior of the cop-
per complex was studied by the cyclic voltammetric technique in
DMF solution and the complex showed well-established redox
behavior at a scan rate of 0.05 V sꢀ1. The antimicrobial activity of
the ligand and its metal complexes were screened against
Escherichia coli, Bacillus subtilis, Aspergillus niger, and Candida albi-
cans, and the results indicated increased activity after coordination
of the ligand to the metal ions. The metal complexes exhibited
enhanced antitubercular activity after chelation against M. tubercu-
losis. The DNA-binding experiments showed that the ligand and its
metal complexes showed effective binding properties through
intercalative mode against CT-DNA. All the synthesized molecules
showed partial cleavage of supercoiled plasmid pUC18 DNA.
KEYWORDS
Pyrazole; azo dyes; metal(II)
complexes; anti-
tuberculosis; DNA
interactions
CONTACT Keshavayya Jathi
jathikeshavayya1959@gmail.com
Department of PG Studies and Research in
Chemistry, School of Chemical Sciences, Kuvempu University, Jnana Sahyadri, Shankaraghatta, Karnataka, India
Supplemental data for this article can be accessed https://doi.org/10.1080/00958972.2019.1630613.
ß 2019 Informa UK Limited, trading as Taylor & Francis Group

2
M. N. MATADA AND K. JATHI
1. Introduction
The dyes and pigments playing a crucial role as coloring agents, particularly the azo-
dyes having conjugative or heterocyclic system, ruled the whole color industry for the
past four decades due to their versatile applications. Azo-dyes are the most important
class of organic compounds with -N¼N- chromophoric group and are generally
formed by conventional diazo-coupling reaction [1,2]. The study of azo-compounds
derived from heterocyclic amines is the fastest growing field because of their wide
range of potential applications in industrial, medicinal, analytical, and pharmaceutical
fields [3,4]. In the industrial field, they were used in optical storage devices, photo-
switches, dying of wool, fabric, and also in inkjet printers due to their good storability,
sensitivity, and thermal stability [5–8]. The synthesis and structural investigations on
coordination complexes of azo-dyes containing pyrazole ring have been widely
studied due to their excellent properties and were significantly used in the dyeing
industry as well as in pharmacological investigations [9]. Particularly the metal com-
plexes having pyrazole system exhibit potential biological properties, including anti-
microbial, anticancer, anti-inflammatory, and anti-tuberculosis activities [10–12].
Azo-components having fused pyrazole and its derivatives played an important role
and showed excellent pharmacological as well as good fabricating agents for cotton,
wool, and fibers [13–15]. Literature survey also revealed that some pyrazole derivatives
have been identified in preventing pathogenic infections such as anti-HIV, anti-inflam-
matory, protein kinase, anti-bacterial, and non-nucleoside HIV-1 reverse transcriptase
inhibitors. The metal(II) complexes of pyrazole-based azo-dyes constitute biologically
potent drugs that attracted much attention of bioinorganic chemists due to their large
ranging pharmacological activities [16,17]. Metwally et al. reported the synthesis of
novel disperse antipyrine based azo-dyes and studied their antitumor and antioxidant
activities [18]. Anitha and co-workers explored the synthesis and biological studies on
some metal complexes derived from azo Schiff base ligand having pyrazole ring [19].
Recently, other studies have reported on the synthesis and biological studies of novel
metal complexes derived from pyrazolone derivatives [20,21].
In view of the above findings and encouraged by our recent investigation on the devel-
opment of biologically potent coordination complexes derived from various heterocyclic
azo-dyes, in the present work we have synthesized some transition metal complexes
derived from 1,5-dimethyl-4-[(3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl)dia-
zenyl]-2-phenyl-1,2-dihydro-3H-pyrazol-3-one [22–25]. The different physicochemical tech-
niques were employed to deduce the geometrical structures of the synthesized metal
complexes. Further, the pharmacological potency of the ligand and its metal complexes
were examined by antimicrobial, antitubercular, DNA-binding, and cleavage experiments.
2. Experimental
2.1. Materials and methods
All reagents and solvents used for the present study were of analytical grade and
obtained from Sigma Aldrich Chemical Company, India. Melting points of the newly

JOURNAL OF COORDINATION CHEMISTRY
3
synthesized compounds were determined by electro-thermal apparatus using open
capillary tubes and are uncorrected. Elemental analysis was carried out on a Vario EL
III CHNS analyzer and metal contents in the complexes were estimated by the stand-
ard methods [26]. UV–visible spectra of the synthesized compounds were recorded on
basic Eppendorf Bio-Spectrometer in the wavelength range of 200–800 nm in DMSO
solution (10ꢀ6 M). Infrared spectra were recorded on a Perkin-Elmer Spectrum RX-IFTIR
1
instrument in the region 4000–400 cmꢀ1. The H and 13C NMR spectra were recorded
using a Bruker Avance II spectrometer and the chemical shifts were calibrated relative
to residual solvent signals. LC-MS spectra of the compounds were recorded on an
LCMS 2010, SHIMADZU mass analyzer. The molar conductivity measurements were
recorded on an ELICO CM-180 conductivity bridge in dry DMSO (10ꢀ6 M) using a dip-
type conductivity cell fitted with a platinum electrode. The magnetic susceptibility
measurements were recorded at room temperature on vibrating sample magnetom-
eter using Ni as the calibrant. EPR spectrum of the copper complex was recorded in
DMSO solution at 77 K on a JES-FA200 EPR spectrometer. The cyclic voltammetric
study on the Cu(II) complex was studied in DMF solution (10ꢀ4 M) on a CHI660E series
model electrochemical analyzer and cetyl trimethyl ammonium chloride
[C15H31(CH3)3NþCl–] was used as a supporting electrolyte. The surface morphology of
the synthesized metal complexes was studied by scanning electron microscopy (SEM)
and energy dispersive X-ray analysis (EDAX) (GEMINI, Ultra 55).
2.2. Synthesis of azo-dye ligand (L)
A well-stirred solution of 4-amino-1,5-dimethyl-2-phenyl-1,2-dihydro-3H-pyrazol-3-one
(1) (2.0 mmol) in 6 mL glacial acetic acid was cooled in an ice-bath and diazotized with
a solution of sodium nitrite (2.2 mmol) in 2 mL of H2SO4 and stirred at 0–5 ꢁC for 2 h.
Further, the cold diazonium salt solution was added to the well-cooled solution of 5-
methyl-2-phenyl-2,4-dihydro-3H-pyrazol-3-one (2) (2 mmol) in acetic acid (5 mL). The
resulting reaction mixture was stirred for another 2 h at 0–5 ꢁC maintaining the pH 5-6
by adding the required volume of saturated sodium carbonate solution. The crude
product was filtered off, washed with hot water, dried and recrystallized from ethanol.
The reaction was monitored by thin layer chromatography (Scheme 1).
2.2.1. Azo-dye ligand A: (L)
Yield: 74%; Elemental analysis calcd (%) for C21H20N6O2: C, 64.94; H, 5.19; N, 21.64;
found: C, 64.42; H, 5.12; N, 21.22. IR (cmꢀ1): v(N-H) 3431, v(Ar-H) 3060, v(C ¼ O) 1744,
1
1665, v(N ¼ N) 1344; H NMR (CDCl3, d ppm): 13.15 (s, 1H, hydrazone NH); 7.91–7.15
(m, 10H, Ar-H); 2.31 (s, 3H, -N-CH3); 2.54 and 3.51 (s, 6H, CH3). 13C NMR (CDCl3, d
ppm): 158.46, 157.49, 147.82, 143.34, 138.28, 134.13, 129.35, 128.85, 128.61, 127.28,
124.86, 124.24, 36.10, 11.81 and 11.49. MS (m/z, 70 eV): 389 (M þ 1)þ.
2.3. Synthesis of Cu(II), Co(II), Ni(II), Mn(II), and Zn(II) metal complexes
To the hot solution of 1,5-dimethyl-4-[(3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyra-
zol-4-yl)diazenyl]-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (0.00054 mol) (L) in 5 mL of

4
M. N. MATADA AND K. JATHI
Scheme 1. Synthetic root adopted for the synthesis of azo-dye ligand (L).
methanol was added a hot methanolic solution of respective metal chlorides
(0.0002 mol). The solution was refluxed on a water bath for about 5–6 h. The reaction
mixture was cooled to room temperature and poured into distilled water. The colored
solids separated and were collected by filtration, washed with sufficient quantity of
distilled water, and then with hot ethanol to remove any impurities. It was finally dried
in a vacuum over anhydrous CaCl2 in a desiccator.
2.3.1. Complex A1: [Cu(L)2]
Yield: 63%; Elemental analysis calcd (%) for C42H38CuN12O4: C, 60.17; H, 4.57; N, 20.05;
Cu, 7.58; found: C, 60.03; H, 4.80; N, 19.93; Cu, 7.51. IR (cmꢀ1): v(Ar-H) 3041, v(C ¼ O)
1648, 1606, v(N ¼ N) 1337, v(M-N) 437, v(M-O) 582; MS (m/z, 70 eV): 839 (M þ 1)þ;
Conductivity (Do, ohmꢀ1 cm2 molꢀ1) in DMSO: 22.
2.3.2. Complex A2: [Co(L)2]
Yield: 68%; Elemental Anal. Calcd (%) for C42H38CoN12O4: C, 60.50; H, 4.59; N, 20.16;
Co, 7.07; found: C, 60.36; H, 4.78; N, 20.11; Co, 6.59. IR (cmꢀ1): v(Ar-H) 3060, v(C ¼ O)
1613, 1587, v(N ¼ N) 1347, v(M-N) 436, v(M-O) 581; MS (m/z, 70 eV): 834 (M þ 1)þ;
Conductivity (Do, ohmꢀ1 cm2 molꢀ1) in DMSO: 14.
2.3.3. Complex A3: [Ni(L)2]
Yield: 73%; Elemental Anal. Calcd (%) for C42H38NiN12O4: C, 60.52; H, 4.60; N, 20.17; Ni,
7.04; found: C, 60.37; H, 4.83; N, 20.12; Ni, 7.66. IR (cmꢀ1): v(Ar-H) 3045, v(C ¼ O) 1614,
1586, v(N ¼ N) 1348, v(M-N) 436, v(M-O) 580; MS (m/z, 70 eV): 834 (M þ 1)þ;
Conductivity (Do, ohmꢀ1 cm2 molꢀ1) in DMSO: 16.
2.3.4. Complex A4: [Mn(L)2]
Yield: 74%; Elemental Anal. Calcd (%) for C42H38MnN12O4: C, 60.79; H, 4.62; N, 20.26;
Mn, 6.62; found: C, 60.65; H, 4.78; N, 20.19; Mn, 6.62. IR (cmꢀ1): v(Ar-H) 3044, v(C ¼ O)
1609, 1587, v(N ¼ N) 1339, v(M-N) 455, v(M-O) 579; MS (m/z, 70 eV): 831 (M þ 1)þ;
Conductivity (Do, ohmꢀ1 cm2 molꢀ1) in DMSO: 21.

JOURNAL OF COORDINATION CHEMISTRY
5
2.3.5. Complex A5: [Zn(L)2]
Yield: 62%; Elemental Anal. Calcd (%) for C42H38ZnN12O4: C, 60.04; H, 4.56; N, 20.00;
Zn, 7.69; found: C, 58.87; H, 4.79; N, 19.96; Zn, 7.77. IR (cmꢀ1): v(Ar-H) 3065, v(C ¼ O)
1610, 1588, v(N ¼ N) 1341, v(M-N) 463, v(M-O) 580; MS (m/z, 70 eV): 841 (M þ 1)þ;
Conductivity (Do, ohmꢀ1 cm2 molꢀ1) in DMSO: 19.
2.4. Pharmacological studies
2.4.1. Antibacterial screening
The antimicrobial activity of the newly synthesized compounds was evaluated against
Escherichia coli (ATCC 25922) and Bacillus subtilis (ATCC 19659) by agar disc diffusion
assay [27]. Briefly, a 24 and 48-h old culture of selected bacteria was mixed with sterile
physiological saline (0.9%) and the turbidity was adjusted to the standard inoculum of
Mac Farland scale 0.5 (106 colony forming suits (CFU) per mL). Petri plates containing
20 mL of Mueller Hinton Agar and Sabouraud dextrose agar was used for the antibac-
terial activity. The inoculum was spread on the surface of the solidified media and
Whatman No. 1 filter paper discs (5 mm in diameter) impregnated with the test com-
pound (20 mL/disc) were placed on the plates. Streptomycin was used as a positive
control for bacteria. A paper disc impregnated with DMSO was used as a negative
control. Plates inoculated with the bacteria were incubated for 24 h at 37 ꢁC. The diam-
eters of zone of inhibition were measured in millimeters by using an antibiotic zone
scale procured from Hi-media. All tests were performed in triplicate and the average
value was taken as the final reading.
2.4.2. Antifungal screening
Antifungal activity of the ligand and its metal complexes was investigated by poisoned
food technique [28] against Aspergillus niger (ATCC627) and Candida albicans (ATCC
10231). The test organisms were sub-cultured using potato dextrose agar medium.
The potato-dextrose-agar medium was sterilized by autoclave at 121 ꢁC (15 lb/sq. inch)
for 15 min. The Petri-plates, tubes, and flasks plugged with cotton were sterilized in an
autoclave at 121 ꢁC for 1 h. Into each sterilized Petri-plate (10 cm diameter), about
30 mL each of molten potato dextrose-agar medium inoculated with respective fungus
(5 mm disc of the fungus grown) was transferred, aseptically along with the solution
of the target compounds in DMSO. The Petri dishes were incubated at 28 ꢁC. The
diameter of the zone of inhibition was read with the help of an “antibiotic zone read-
er.” The experiments were performed in triplicate in order to minimize the errors. The
inhibition percentage (IP) of the A. niger and C. albicans mycelial growth was calcu-
lated using the following formula:
ꢀ
ꢁ
CꢀT
IP ¼
ꢂ 100
(1)
C
where IP is the percentage of inhibition and C is the average of three replicates of
mycelial growth (cm) of control Petri dishes and T is the average of three replicates of
mycelial growth (cm) of treated Petri dishes. Fluconazole was used as the positive con-
trol and DMSO was thus used as the negative control.

6
M. N. MATADA AND K. JATHI
2.4.3. Anti-tuberculosis activity
The ligand and its metal complexes were further screened for their anti-mycobacterial
activity against Mycobacterium tuberculosis (ATCC 27294) using Microplate Alamar Blue
assay (MABA). This methodology is non-toxic, uses a thermally stable reagent and
shows good correlation with proportional and BACTEC radiometric method [29,30].
Briefly, 200 mL of sterile deionized water was added to all outer perimeter wells of the
sterile 96-well plate to minimized evaporation of medium in the test wells during incu-
bation. The 96-well plate received 100 mL of the Middlebrook 7H9 broth and serial
dilution of compounds was made directly on the plate. The final concentration of the
compounds tested was 100 to 0.2 mg/mL. Plates were covered and sealed with Para
film and incubated at 37 ꢁC for five days. After incubation, 25 mL of freshly prepared
1:1 mixture of Almar blue reagent and 10% Tween-80 was added to the plate and
incubated for 24 h. The blue color in the well was interpreted as no bacterial growth,
and pink color was scored as growth. The minimum inhibitory concentration (MIC)
was defined as the lowest drug concentration which prevented the color change from
blue to pink. The test compounds were compared with standard drug Pyrazinamide
(MIC ¼ 3.125 lg/mL).
2.4.4. DNA-binding studies
The interaction between the newly synthesized azo-dye and its metal complexes with
calf-thymus DNA (CT-DNA) was investigated by binding studies as described in earlier
studies [31]. The DNA-binding experiment was carried out by electronic absorption
spectral titrations and the test samples of the synthesized compounds were prepared
in a mixture of DMSO and Tris-HCl buffer (5 mM Tris-HCl, 50 mM NaCl, pH 7.1) solution,
where the concentration of the compounds was kept constant. All the experiments
involving the interaction of the test compounds with CT-DNA were carried out in Tris-
HCl buffer solution. The CT-DNA used for the present study was procured from Merck
Chemical Company in the form of disodium salt and stored at 4 ꢁC. While conducting
the absorption spectral titrations concentration of the target compounds was kept
constant and the concentration of DNA was varied. The DNA solution was added to
both the test solution and reference solution in order to eliminate the absorbance of
DNA itself. The prepared test solutions having the synthesized compounds and the
DNA along with the reference solution were incubated for 2 min before recording the
absorption spectra. The electronic spectra were recorded for the test solutions with
increasing concentration of DNA in the range 200–800 nm. From the electronic spec-
tral data, the intrinsic binding constant (Kb) was determined by observing the changes
in the absorption band with increasing concentration of DNA and fitting the above
experimental results into the Stern-Volmer equation which is given by Eq. (2):
½
ꢃ
½
ꢃ
DNA
DNA
1
¼
þ
(2)
e ꢀ e
a
e ꢀ e
b
K e ꢀe
fꢃ
½
fꢃ
½
fꢃ
½
b
b
Here, [DNA] represents the concentration of DNA, ea the apparent molar absorption
coefficient, ef the molar absorption coefficient of the free compound and eb the molar
absorption coefficient of the compound bound to DNA. Further, a plot of [DNA]/(ef –
ea) versus [DNA] gave a slope equal to 1/(ef – ea) and a y-intercept of 1/(Kb(ef – ea)), and

JOURNAL OF COORDINATION CHEMISTRY
7
the intrinsic binding constant is the ratio of slope to the y-intercept which was
obtained from the above plot.
2.4.5. DNA-cleavage activity
DNA-cleavage activity of the synthesized compounds was studied against supercoiled
plasmid DNA pUC 18 as a target molecule by using agarose gel electrophoresis
method [32,33]. Briefly, 200 mg of agarose was dissolved in 25 mL of TAE buffer (4.84 g
Tris base, pH 8.0, 0.5 M EDTA) by boiling. When the gel attained approximately 55 ꢁC,
it was poured into the gel cassette fitted with a comb. The gel was allowed to solidify
and then carefully the comb was removed. The gel was placed in the electrophoresis
chamber flooded with TAE buffer. The target compounds were dissolved in freshly dis-
tilled DMSO (1 mg/mL). The test compounds were added separately to the isolated
pUC18 DNA (225 ng) and incubated for 2 h at 37 ꢁC. After incubation, 20 mL of DNA
sample (mixed with bromophenol blue dye at 1:1 ratio) was loaded carefully into the
wells, along with standard DNA marker and constant electricity of 50 V was passed for
around 45 min. The gel was removed and carefully stained with ethidium bromide
(ETBR) solution (10 mg/mL) for 10–15 min. The bands were observed under UV transil-
luminator (UVP, Germany) and photographed to determine the extent of DNA-cleav-
age, and the results were compared with those of a standard DNA-marker.
3. Results and discussion
All the newly synthesized metal complexes of the azo-dye ligand (L) are stable, non-
hygroscopic, colored and partially soluble in common organic solvents and completely
soluble in DMF and DMSO. Melting points of the neꢀw1ly synthesized complexes were
above 300 ꢁC. Molar conductance values (14–22 ohm cm2 molꢀ1) of the complexes
revealed that they are non-electrolytic in nature [34]. The analytical and physical data
of the azo-dye ligand (L) and its metal complexes are summarized in Table S1, which
is provided in the Supplementary Information. From the analytical data, the metal-to-
ligand ratio in all complexes was found to be 1:2. Elemental analysis of the synthe-
sized compounds is in good agreement with the proposed stoichiometry of
the complexes.
3.1. IR spectral data
The IR spectra of the ligand and its metal complexes were recorded in KBr pellets to
understand the coordination mode between the ligand and the corresponding metal
complexes. The main absorption bands of the azo dye ligand and its metal complexes
are presented in Table 1. The IR spectrum of the ligand exhibited a band at
3431 cmꢀ1 assigned to the N-H functional group of the hydrazo-form of the ligand. In
the IR spectra of the metal complexes, the disappearance of absorption band at
3431 cmꢀ1 indicates the loss of hydrogen of the NH group via enolization [35].
Further, it is observed that the enolic oxygen and azoic nitrogen are involved in
coordination with the metal ions as there appeared new bands at 584–579 and
463–436 cmꢀ1 corresponding to the formation of M-O and M-N bonds, respectively.

8
M. N. MATADA AND K. JATHI
Table 1. Comparative IR frequencies (cmꢀ1) of the azo-dye ligand (L) and its metal complexes.
Compound
v(N-H)
v(Ar-H)
v(C ¼ O)
v(N ¼ N)
v(M-N)
v(M-O)
A
L
3431
3060
3041
3060
3045
3044
3065
1744, 1666
1648
–
–
–
A1
A2
A3
A4
A5
[Cu(L)2]
[Co(L)2]
[Ni(L)2]
[Mn(L)2]
[Zn(L)2]
–
–
–
–
–
1337
1347
1348
1339
1341
437
436
436
455
463
582
581
580
579
580
1613
1614
1609
1610
However, the observed new absorption bands in the range of 1348–1337 cmꢀ1 are
due to –N ¼ N– group of the metal complexes which confirms the formation of com-
plexes in the azoic from of the ligand [36]. The strong-intensity band observed at
1744 cmꢀ1 in the ligand due to carbonyl function v(C ¼ O) is shifted to lower fre-
quency side and appeared in the range 1665–1606 cmꢀ1. This confirms the involve-
ment of oxygen atom of the carbonyl group in the complexation with the metal ions
[37]. Medium intensity bands observed in the region of 3041–3065 cmꢀ1 in all the
metal complexes are assigned to aromatic C–H stretching vibrations. Therefore, from
the above IR spectral data it is inferred that the ligand coordinated with the metal
ions in azo-enol form. Further, it is observed that the ligand behaves as tridentate and
forms the bond with the metal ions via azoic nitrogen, enolic oxygen and oxygen
atom of the carbonyl group (ONO).
3.2. Electronic spectra and magnetic susceptibility measurement
The electronic spectra of the ligand and its metal complexes were measured in DMSO
in the wavelength range 200–800 nm and the electronic absorption spectral data of
the azo-dye ligand (L) and its metal complexes are presented in Table 2. The azo-dye
ligand (L) exhibits electronic transitions with a band at 30769 cmꢀ1 assigned to p!p
ꢄ
transitions. The absorption band at 22573 cmꢀ1 corresponds to n!p transition asso-
ciated with the –N¼N– group of the azo-dye [38]. In all the synthesized complexes,
this band undergoes shift towards lower wavelengths due to the coordination of the
nitrogen atom of the azo-group with the metal ion via deprotonation.
ꢄ
The absorption spectra of the Cu(II) complex shows two bands each at 20703 and
2
2
2
29230 cmꢀ1 which are attributed to the electronic transition to B1g ! A1g (v1), B1g
2
2
2
! B2g (v2) and B1g ! Eg (v3). These bands and transitions indicate that Cu(II) has a
distorted octahedral geometry. Further, the observed magnetic moment value of the
Cu(II) complex is 1.85 BM and falls within the range observed for octahedral geom-
etry [39].
In the absorption spectrum of the Co(II) complex, three absorption bands appear
4
4
each at 15082, 18903 and 26737 cmꢀ1 which are assigned to T1g(F) ! T2g(F) (v1),
4
4
4
4T1g(F) ! A2g(F) (v2) and T1g(F) ! T2g(P) (v3) transitions, respectively [40]. These
transitions suggest an octahedral geometry for Co(II), which is well within the range of
literature values. This was further confirmed by the magnetic moment, which was
found to be 3.28 BM, and falls within the range of 2.8–3.5 BM, suggesting octahedral
geometry [41].
The Ni(II) complex showed two absorption bands each at 21097 and 27322 cmꢀ1
3
3
3
3
which are assigned to A2g ! T1g(F) (v2) and A2g(F) ! T1g(P) (v3) transitions. These

JOURNAL OF COORDINATION CHEMISTRY
9
Table 2. UV–vis spectral data of the azo-dye ligand (L) and its metal complexes.
Ligand/complexes
Wavenumber (cm–1
)
Assignments
meff (BM)
ꢄ
ꢄ
A
A1
L
30769, 22573
20703
p!p , n!p
–
1.85
[Cu(L)2]
2B1g
2B1g
!
2A1g
,
29230
!
2B2g
,
2B1g ! Eg
2
4
A2
[Co(L)2]
15082
18903
26737
21097
27322
21717
23860
26068
26472
31377
29552
4T1g (F) ! T2g (F),
3.28
4T1g(F) ! A2g (F),
4
4T1g (F) ! T2g (P)
4
A3
A4
[Ni(L)2]
3A2g
!
3T1g (F),
2.91
5.73
3A2g (F) ! 3T1g (P)
4
[Mn(L)2]
6A1g ! T1g (4G),
6A1g ! Eg (4G),
4
6A1g ! Eg (4G),
4
6A1g
!
4T1g (4P)
A5
[Zn(L)2]
LMCT (M!N)
Diamagnetic
transitions suggest an octahedral environment around the Ni(II) ion. Also, the mag-
netic moment value 2.91 BM was consistent with octahedral geometry around Ni(II)
ion [42].
The electronic absorption spectrum of the Mn(II) complex exhibited five absorption
bands each at 21717, 23860, 26068, 26472 and 31377 cmꢀ1 which are assigned to
4
6
4
6
4
6
4
6A1g ! T1g(4G) (v1), A1g ! Eg (4G) (v2), A1g ! Eg (4G) (v3) and A1g ! T1g (4P)
(v4) transitions. The electronic transitions together with a magnetic moment value 5.73
BM, which is close to the spin-only value (5.92 BM), suggests an octahedral geometry
for the Mn(II) complex [43,44].
The absorption spectrum of the Zn(II) complex showed broad absorption bands in
the region 29552–23098 cmꢀ1 corresponding to the ligand-to-metal charge-transfer
transitions (LMCT). Except this, the complex does not show any bands in the region
below 23098 cmꢀ1 in accordance with the d10 system of Zn(II) ion [45].
1
3.3. H and 13C NMR spectral data of the azo-dye ligand (L)
In the 1H NMR spectrum of the azo-dye ligand (Figure 1) measured in CDCl3, the
hydrazone proton appears as a singlet at 13.15 ppm (s, 1H, hydrazone NH) which cor-
responds to imine-NH proton resonance of the hydrazone form. We have also
observed that one absorption peak was observed in the IR spectrum of the ligand at
3431 cmꢀ1 corresponding to NH functional group, which confirms that the compound
exists in azo-hydrazo form as represented in Scheme 1. Our observation confirms the
azo-hydrazo tautomerism, which is in consistent with the reported literature [46]. The
ten aromatic protons resonated as multiplets in the region 7.91–7.15 ppm (m, 10H, Ar-
H). The signal appeared as a singlet at 2.31 ppm (s, 3H, N-CH3) due to the methyl
group attached to the antipyrine moiety. The signals appeared as a singlet at 2.54 and
3.51 ppm (s, 6H, CH3) corresponding to the six methyl protons.
The 13C NMR spectrum of the synthesized azo-dye was recorded at room tempera-
ture in a CDCl3 solvent. The carbonyl carbon of both the pyrazole rings resonated at
158.76 and 157.49 ppm, respectively. The carbon atoms attached on both sides of the
azo-chromophore appeared at 147.82 and 143.34 ppm. The signals observed at 138.28
and 134.13 ppm are due to the carbon atoms of the pyrazole ring attached to the

10
M. N. MATADA AND K. JATHI
Figure 1. Cyclic voltammogram of the Cu(II) complex of the azo-dye ligand (L).
methyl group. The ligand exhibited signals at 129.35, 128.85, 128.61, 127.28, 124.86
and 124.24 ppm, corresponding to carbon atoms of the phenyl ring attached to the
pyrazole moiety. The methyl carbon attached to the nitrogen atom of the pyrazole
ring resonated at 36.30 ppm and the carbon atoms of the methyl group in the pyra-
zole ring exhibited signals at 11.83 and 11.52 ppm, respectively. Therefore, from overall
1
observation, it is evident that the H and 13C NMR spectral data are in good agree-
ment with the proposed molecular structure of the ligand.
3.4. ESI-mass spectral data
The newly synthesized azo-dye ligand and its metal complexes were studied for their
mass spectral study. The mass spectrum of the ligand exhibited a molecular ion peak
equivalent to its molecular weight along with other fragment peaks. The mass spec-
trum of the azo-dye ligand exhibited M þ 1 peak at 389 (100%), which is also a base
peak. This on a simultaneous loss of C2H4 and CH10 radicals showed a peak for the
fragments at m/z 360 and 338, respectively. The mode of mass fragmentation of the
azo-dye (L) is consistent with its molecular formula and schematic representation is
presented in Scheme S2 (Supplementary Information).
In the mass spectrum of the Cu(II) complex, the molecular ion peak appeared at m/
z 839 corresponding to its molecular weight of the complex. The molecular ion under-
went simultaneous loss of C6H14 radical and exhibited a fragment peak at m/z 751.
This fragment ion on a loss of two cyanide and one hydroxyl radicals produced a frag-
ment peak recorded at 664. The fragment lost a part of the ligand with molecular for-
mula C8H3O2 and C8HN2 and gave peaks recorded at m/z 535 and 409, respectively.
This fragment ion further undergoes sequential loss of C12H14 and C3H2N4 radicals and
exhibited fragment peaks recorded at m/z 250 and 156, respectively. The schematic
representation of the proposed mode of the fragmentation of the Cu(II) complex is
shown in Scheme S3 (Supplementary Information), consistent with its molecu-
lar structure.
The mass spectrum of the Co(II) complex produced a molecular ion peak after sup-
plying the 70 eV energy and appeared at m/z 834 equivalent to its formula weight.

JOURNAL OF COORDINATION CHEMISTRY
11
The molecular ion underwent the simultaneous loss of C4H10 radical and gave a peak
at m/z 775. This on the loss of phenyl and hydrogen radical and exhibited a frag-
mented peak at m/z 697. The complex has undergone fragmentation with the elimin-
ation of C10H7N2O, C6H5, C5HN5O, C6H5, C2HN2O and C2HNO fragments and showed a
fragments peak at m/z 526, 449, 302, 225, 156 and 102, respectively. Further, the sche-
matic representation of the fragmentation pattern is depicted in Scheme S4
(Supplementary Information).
The mass spectrum of the Ni(II) complex showed the molecular ion peak at m/z 834
corresponding to its molecular formula. Also, the mass spectrum exhibits fragment
peaks which appeared at m/z 795, 704, 662, 527, 421, 302, 247 and 96 due to the
elimination of C2HN, C6H5N, C2HO, C10H16, C7H5N, C7H5NO, C2H4N2 and C4HN6O frag-
ments, respectively. In the mass spectrum of the Mn(II) complex the molecular ion
peak appeared at m/z 831 corresponding to its weight. This molecular ion further
underwent sequential fragmentation with the elimination of three methyl, phenyl,
C11H8N2O, C6H4N2O, methyl, NH, C3HN3O, C3N2O and Mn2þ fragments and exhibited
the fragment peaks at m/z 708, 523, 403, 387, 293, 214 and 157, respectively. Similarly,
the Zn(II) complex showed a molecular ion peak at m/z 841, this on a simultaneous
loss of C7H6, C5H12, C12H12, C6H4, C3N4O, CH2N3O and C3NO fragments and showed
fragment peaks at 751, 677, 520, 446, 336, 264 and 198, respectively. The tentative
mode of fragmentation of the Ni(II), Mn(II) and Zn(II) complexes is shown in Schemes
S5–S7 (Supplementary Information), respectively. From overall mass spectral fragmen-
tation, it is inferred that the obtained m/z values for the fragments are in good agree-
ment with the proposed molecular structures of all the metal complexes of the azo-
dye ligand (L) and confirm their stoichiometry as [M(L)2].
3.5. EPR spectral data
To understand the nature of the metal–ligand bonding and to determine the mag-
netic interaction in the metal complexes, the EPR spectra of the Cu(II) complex of the
azo-dye ligand was recorded in DMSO solution at 77 K. The axial symmetric parame-
ters such as g , g , gav and G were evaluated and depicted in Table 3. The room tem-
jj
?
perature EPR spectra of the copper complex exhibit axially symmetric parameters with
g
(2.14) > g (2.02) > g (2.0023), suggesting the unpaired electron predominantly
jj
?
resides in dx2-y2 orbital which is characteristic of the octahedral environment [47].
From the results, it is evident that the value of g , which is the measure of the cova-
jj
lent character of the metal–ligand bond, is less than 2.3, indicating the noticeable
covalent character in the metal–ligand bonding [48]. The band corresponding to
DMs¼ 2 transition has not appeared in the observed spectrum, ruling out any Cu–Cu
interaction and therefore the complex has mononuclear nature. The value of G a geo-
metric parameter to know the exchange interaction between the two metal centers
can be measured from the expression G ¼ (g – 2.0023)/(g – 2.0023). According to
jj
?
Hathaway, if the value is G > 4, it indicates the negligible interaction and if G < 4, con-
siderable interaction exists between the metal centers in a complex. In the present
case, the value of G is higher than 4 (G ¼ 5.71), suggesting the negligible exchange
interaction in a complex and ruled out the possibility of binuclear nature [49].

12
M. N. MATADA AND K. JATHI
Table 3. Spin Hamiltonian parameter calculated for the Cu(II) complex in DMSO at 77 K.
Compound
[Cu(L)2]
g
g
gav
G
a2
b2
c2
K
K
?
0.16
jj
?
jj
2.14
2.02
2.10
5.71
0.26
0.98
0.62
0.25
The molecular orbital coefficient a2 (covalent in-plane r-bonding), b2 (covalent in-
plane p-bonding) and c2 (out-plane p-bonding), a measure of metal–ligand bonding
were derived from the following expressions:
À
Á
A
3
7
?
a2 ¼
þ g ꢀ 2:0027 þ þ g ꢀ2:0027 þ 0:04
(3)
(4)
ð
Þ
?
jj
0:036
g ꢀ2:0027 E
ð
Þ
?
ð
jj
b2 ¼
2
Þ
ꢀ8k a
À
Á
g ꢀ2:0027 E
c2 ¼
(5)
2
ð
Þ
ꢀ2k a
where k ¼ –828 cmꢀ1 for free copper ion and E is the electronic transition energy
[50]. These parameters are more effective in determining the covalent or ionic charac-
ter of the metal–ligand bonding. The value of a2 is 0.5 and indicates the complete
covalent nature, and if it is 1, it may suggest the complete ionic character of the
bond. For the present copper complex, the value was found to be 0.26, confirming
that there is some covalent character in metal–ligand bonding. The other two factors
such as b2 and c2 (0.98 and 0.62, respectively) suggest the possible interaction
between the out-of-plane p-bonding, whereas in-plane p-bonding is ionic. Further, the
orbital reduction factors K and K , which are obtained from the relations (6) and (7),
jj
?
give the detailed information for the nature of bonding in the complex [51].
À
Á
K ¼ g ꢀ 2:0027 E=8ko
(6)
(7)
jj
jj
K ¼ g ꢀ 2:0027 E=2k
ð
Þ
?
?
o
According to the Hathaway expressions, if K ꢅ K , there is pure r-bonding, if K <
jj
?
jj
K , there is an in-plane p-bonding and if K > K , suggests the existence of out-of-
?
jj
?
plane p-bonding [52]. From Table 3, it is inferred that the values K (0.25) and K
jj
?
(0.16) satisfy the condition K < K and therefore there is relatively more contribution
jj
?
of an in-plane p-bonding as compared to out-of-plane p-bonding in the metal–ligand
p-bonding [53]. Therefore, from overall observation, it is concluded that the EPR spec-
tra of the Cu(II) complex provided significant importance as additional information to
the conclusion obtained on the basis of magnetic susceptibility and electronic spec-
tral studies.
3.6. Redox behavior of Cu(II) complex of the azo-dye ligand (L)
The copper complex was examined for its electrochemical behavior (redox properties)
by cyclic voltammetry (CV) at room temperature and cetyl trimethyl ammonium chlor-
ide [C15H31(CH3)3NþCl-] as supporting electrolyte. Cyclic voltammogram of the complex
was recorded by employing glassy carbon electrode, Ag/AgCl electrode and the plat-
inum wire as working, reference and auxiliary electrodes, respectively. The measure-
ment was carried out by using the 10ꢀ4 M solution of DMF in the potential range of

JOURNAL OF COORDINATION CHEMISTRY
13
Figure 2. SEM images of the metal complexes of the azo dye ligand (L).
–1.0 to þ1.0 V with a scan rate of 0.05 V sꢀ1. The graphical representation of the cyc-
lic voltammogram of the copper complex is depicted in Figure 1.
The CV profile of the complex showed well-established reduction and oxidation
peaks at Epc ¼ 287 mV and Epa ¼ 442 mV, respectively, corresponding to the redox
process of the metal complex and the azo-dye ligand (L). The separation between the
potential of the reduction and the oxidation peaks is the measure of the number of
electrons transferred in the electrode reaction. In the present case, the peak separation
(DE) has been found to be 155 mV at 0.05 Vꢀ1, suggesting quasi-reversible one elec-
tron transfer reaction [9].
3.7. Surface morphology (SEM and EDAX)
The SEM and EDAX images of the newly synthesized metal complexes that provide
extensive information regarding surface morphology and chemical composition of the
complexes were recorded. The metal complexes of the azo-dye ligand (L) exhibited
different SEM morphologies as depicted in Figure 2 and the SEM micrographs of all
the complexes showed homogeneous nature. The rod-shaped cylindrical structure was
observed for Cu(II) complex, irregular flake-like morphology, the broken flake-like
structure was observed in Ni(II) complex, the needle-shaped flower structure was
shown by Mn(II) complex and an irregular semi-flattened rod-like structure was seen
in Zn(II) complex. Further, the EDAX spectral data of the metal complexes confirmed
the presence of respective elements in their surfaces and their respective graphs are

14
M. N. MATADA AND K. JATHI
Figure 3. EDAX spectra of the metal complexes of the azo dye ligand (L).
shown in Figure 3. The EDAX graphs have indicated the existence of copper, cobalt,
nickel, manganese, and zinc ions in their structure along with C, N, O, and S atoms in
the respective complexes.
3.8. Pharmacological evaluation
3.8.1. Antibacterial activity
The azo-dye ligand and its metal complexes were screened for their antibacterial activ-
ity against E. coli and B. subtilis in DMSO by the well diffusion method and the results
of the activity along with the standard drug are displayed in Table 4. Streptomycin
was used as the standard drug. From the results, it is suggested that the Cu(II) and
Ni(II) complexes showed higher activity against both the bacterial strains whereas the

JOURNAL OF COORDINATION CHEMISTRY
15
Table 4. Antibacterial activity results of azodye ligand (L) and its metal complexes.
Zone of inhibition (mm)
E. coli
B. subtilis
Compounds
25 mg/mL
50 mg/mL
25 mg/mL
50 mg/mL
A
A1
A2
A3
A4
A5
1.1 0.6
1.5 02
1.3 0.22
1.8 0.1
1.2 0.25
1.3 0.22
1.9 0.25
1.5 0.8
1.8 0.5
1.8 0.23
2.1 0.30
1.8 0.2
1.4 0.23
2.5 0.28
1.3 0.3
2.0 0.12
1.4 0.1
1.9 0.24
1.5 0.4
1.5 0.1
2.0 0.32
1.7 0.5
2.4 0.12
1.8 0.4
2.4 0.29
1.6 0.3
2.3 0.4
2.6 0.35
Streptomycin
Table 5. Antifungal activity results of the azo dye ligand (L) and its metal complexes.
% of inhibition
A. flavus
C. albicans
Compounds
50 mg/mL
100 mg/mL
50 mg/mL
100 mg/mL
A
A1
A2
A3
A4
A5
25 0.70
33 0.35
44 0.28
46 0.14
46 0.71
32 0.15
37 0.16
58 0.26
65 0.15
70 0.22
74 0.33
82 0.13
65 0.23
78 0.17
21 0.42
25 0.29
55 0.62
39 0.14
35 0.13
23 0.31
39 0.16
57 0.37
66 0.25
64 0.37
78 0.15
70 0.73
68 0.72
81 0.14
Fluconazole
rest of the compounds exhibited moderate activity. Further, the observed activity in
the copper and nickel complexes over the ligand may be attributed to the change in
molecular structure as a result of coordination with the metal ions [54].
3.8.2. Antifungal activity
To provide a medicinal scope in the field of bioinorganic chemistry, consequently, the
synthesized ligand and its metal complexes have been evaluated for their antifungal
activity against two fungal strains, A. niger and C. albicans, by food poison method
and the results were tabulated in Table 5. The results of the antifungal activity reveal
that the Ni(II) and Mn(II) complexes show higher activity than the free ligand. The
increase in the antifungal activity of the metal complexes is due to the inhibition of
the multiplication process of the microbes by blocking their active sites. Such
increased activity on metal chelation can be explained on the basis of chelation theory
[55,56]. The chelation also increases the lipophilic character of the cell wall and the
interaction between the metal ion and lipid is favored. This may lead to the collapse
of the permeability of the cell wall, resulting in interference with the normal cell proc-
esses, while chelation is not the only factor for antimicrobial activity but also on sev-
eral aspects like the nature of metal ion and the ligand, geometry of metal complexes,
the lipophilicity and pharmacokinetic factors [57].
3.8.3. Anti-tuberculosis activity
The results of the antimicrobial studies encouraged us to investigate the preliminary
screening of the synthesized ligand and its metal complexes for their in vitro anti-
mycobacterium tuberculosis (TB) activity, expressed as MIC. The Mn(II) complex

16
M. N. MATADA AND K. JATHI
Table 6. Anti-tuberculosis activity results of the azo dye ligand (L) and its metal complexes.
Compounds 100 mg/mL 50 mg/mL 25 mg/mL 12.5 mg/mL 6.25 mg/mL 3.12 mg/mL 1.6 mg/mL 0.8 mg/mL
A
A1
A2
A3
A4
A5
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
N
N
N
N
S
N
S
S
S
N
N
N
N
N
N
S
N
N
N
N
N
N
S
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
Pyrazinamide
Ciprofloxacin
Streptomycin
S
S
S
N
N, not sensitive; S, sensitive.
showed highest MIC value of 12.5 mg/mL and all other complexes exhibited moderate
activity with MIC equal to 25 mg/mL as compared to the standard drugs, which may
be taken as lead compounds for the development of effective anti-TB drugs [58]. The
results obtained are presented in Table 6 and Figure S1 (Supplementary Information)
and compared with the available standard drugs pyrazinamide, streptomycin, and
ciprofloxacin.
3.8.4. DNA-binding experiment
From the recent studies on the binding abilities of the metal complexes with DNA
revealed that some of the drugs having anticancer properties destruct the DNA-repli-
cation by reducing the RNA production via binding with the nuclear DNA [59]. In add-
ition to this, the electronic spectroscopy becomes an advanced and reliable method
to determine the extent of binding of the complexes with DNA [60]. These binding
studies are effective tools in order to develop effective and novel metal-based drugs
that are targeted to DNA. The binding properties of the synthesized ligand and its
metal complexes were investigated against CT-DNA and the electronic absorption
spectra were recorded with the increasing concentration of CT-DNA in the region
300–800 nm and represented in Figure 4. During the experimental condition, while
increasing the concentration of CT-DNA, a significant hypochromic effect was observed
with a slight red-shift, suggesting stabilization of the double helical structure of DNA.
The red-shift in the absorption spectra of the metal complexes insights the interaction
of the lone pair of electrons of the ligand with CT-DNA. While increasing the concen-
tration of CT-DNA, the absorption bands exhibited better hypochromism of about
22–41% for all the compounds, which indicates significant binding with DNA with par-
tial intercalation. These absorption spectral characteristics suggest that there is a sig-
nificant interaction of the extensive conjugation and chromophores of metal chelates
with the base pairs of CT-DNA. The intrinsic binding constant (Kb) for all the molecules
were evaluated by using the Wolfe-Shimer equation (Eqn. 2), where the plot of [DNA]/
(ef – ea) versus [DNA] gave a slope equal to 1/(ef – ea) and a y-intercept of 1/(Kb(ef –
ea)), and the plots are displayed in Figure S2 (Supplementary Information). From the
above plot, we obtain the values of Kb and are summarized in Table 7. The Kb values
obtained for ligand, Cu(II), Co(II), Ni(II), Mn(II), and Zn(II) complexes were in the range
0.78–1.25 ꢂ 106 Mꢀ1 which is in accordance with the values obtained for known clas-
sical intercalators (for example, ethidium-DNA which has the value of 1.4 ꢂ 106
M
ꢀ1) [61–63].

JOURNAL OF COORDINATION CHEMISTRY
17
Figure 4. Electronic absorption spectra of azo dye ligand (L) and its metal complexes in a buffer
solution of pH 7.2 in the presence of increasing concentration of CT-DNA.
Table 7. Electronic absorption spectral features of the DNA binding experiments of the ligand
and its metal complexes.
kmax (nm)
Compounds
Free
Bound
Dk (nm)
Hypochromicity (%)
Kbꢂ106 (M–1
)
A
239
239
237
237
231
234
240
241
238
237
232
235
1
2
1
0
1
1
33
37
22
41
34
34
1.12
1.17
1.25
0.80
0.81
0.78
A1
A2
A3
A4
A5

18
M. N. MATADA AND K. JATHI
3.8.5. DNA-cleavage studies
To understand the mechanism of drug toxicity, DNA-cleavage studies play an import-
ant role to overcome the toxicity and design of novel drugs [64]. The interaction of
the newly synthesized ligand and its metal complexes with plasmid pUC18 DNA was
examined by gel electrophoresis technique. It is evident from the electrophoresis tech-
nique that the ligand and its metal complexes acted on the isolated DNA because of
a difference in molecular weight between the control DNA and the treated DNA sam-
ples. In the present investigation, all the tested compounds displayed partial cleavage
of Form I DNA. However, Form II DNA was cleaved completely by all the compounds
and it is important to note that the better DNA cleavage by meager significance could
be made upon increased concentrations of sample as Form I and Form II DNA band
intensity was diminished. On the basis of these results, it is apparent that the tested
compounds are potential inhibitors for pathogenic microorganisms. The gel picture
exhibiting the DNA cleavage is shown in Figure 5.
Figure 5. DNA cleavage activity on plasmid pUC18 DNA: M: Standard DNA, C: Control DNA
(untreated pUC 18), A1: azo-dye ligand, A2: Cu(II) complex, A3: Co(II) complex, A4: Ni(II) complex,
A5: Mn(II) complex and A6: Zn(II) complex
CH3
H3C
H3C
N
N
N
N
N
N
Ph
Ph
O
O
M
O
O
Ph
Ph
N
N
N
N
N
CH3
N
CH3
H3C
M: Cu(II), Co(II), Ni(II), Mn(II) and Zn(II)
Figure 6. Proposed molecular structure of the metal complexes of the azo-dye ligand (L).

JOURNAL OF COORDINATION CHEMISTRY
19
4. Conclusion
The present work mainly focuses on the design of bioactive transition metal(II) com-
plexes obtained from azo-dye ligand 1,5-dimethyl-4-[(3-methyl-5-oxo-1-phenyl-4,5-
dihydro-1H-pyrazol-4-yl)diazenyl]-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (L). The struc-
ture of the newly synthesized molecules was elucidated by various analytical and spec-
troscopic studies such as elemental analysis, FT-IR, electronic, NMR, mass, EPR, molar
conductivity, magnetic susceptibility measurements, SEM, and EDAX techniques. The
non-electrolytic nature of the complexes was confirmed by molar conductivity data. IR
data suggested the mode of bonding between the metal ion and the ligand through
the nitrogen atom of the azo group, enolic oxygen atom of the carbonyl functional
group. Based on the physicochemical evidence, the following structures were pro-
posed for the complexes (Figure 6). The Cu(II) complex showed well-established redox
behavior in DMF solution. SEM and EDAX studies revealed their homogeneous surface
morphology and the coordination of ligand with metal ions, respectively.
Antimicrobial studies of the metal complexes have significant activity than the corre-
sponding azo-dye ligand. The Cu(II), Ni(II) and Mn(II) complexes exhibited potentially
better activity against both bacterial and fungal strains. All the metal complexes
showed promising activity against M. tuberculosis as compared to the free ligand. The
metal chelates of the azo-dye ligand showed effective binding with CT-DNA through
intercalation mode. The cleavage properties of the synthesized molecules were studied
against supercoiled plasmid pUC18 DNA and all the compounds displayed partial
cleavage of pUC18 DNA.
Acknowledgements
Authors are thankful to the Department of Chemistry, Kuvempu University, Shivamogga for pro-
viding laboratory facility. Authors are also grateful to SAIF and CIL Panjab University for provid-
ing spectral data and Maratha Mandal’s Central research lab, Belagavi for anti-
tuberculosis results.
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