Square planar palladium(II) complexes of bipyridines: Synthesis, characterization, and biological studies

Article abstract of DOI:10.1080/00958972.2012.727207

A series of square planar mononuclear palladium(II) complexes, [Pd(L ⁿ)(OMe)₂] (where L ⁿ =4-(4-chlorophenyl)-6- phenyl-2,2′-bipyridine, 4,6-bis(4-chlorophenyl)-2,2′-bipyridine, 4-(4-bromophenyl)-6-(4-chlorophenyl)-2,2′-bipyridine, and 6-(4-chlorophenyl)-4-p-tolyl-2,2′-bipyridine), have been synthesized and characterized. Interaction of palladium complexes with DNA has been investigated by electronic absorption spectra, viscosity measurements and DNA melting temperature techniques. The metal complex-DNA interaction produced hypochromism and bathochromism, suggesting intercalative binding. Viscosity data support the intercalation mode of binding. Palladium(II) complexes promote cleavage of plasmid pUC19 DNA from the supercoiled form to the open circular form. Antibacterial activities of the complexes have also been evaluated against five pathogenic bacteria. The antimicrobial test results reveal that all compounds have good antibacterial activity against both Gram(ve) and Gram(+ve) bacterial species compared to the ligands and metal salt.

Full text of DOI:10.1080/00958972.2012.727207

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Square planar palladium(II)
complexes of bipyridines: synthesis,
characterization, and biological studies
Mohan N. Patel a , Promise A. Dosi a & Bhupesh S. Bhatt a
a Department of Chemistry, Sardar Patel University, Vallabh
Vidyanagar 388 120, Gujarat, India
Accepted author version posted online: 04 Sep 2012.Version of
record first published: 18 Sep 2012.
To cite this article: Mohan N. Patel , Promise A. Dosi & Bhupesh S. Bhatt (2012): Square planar
palladium(II) complexes of bipyridines: synthesis, characterization, and biological studies, Journal
of Coordination Chemistry, 65:21, 3833-3844
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Journal of Coordination Chemistry
Vol. 65, No. 21, 10 November 2012, 3833–3844
Square planar palladium(II) complexes of bipyridines: synthesis,
characterization, and biological studies
MOHAN N. PATEL*, PROMISE A. DOSI and BHUPESH S. BHATT
Department of Chemistry, Sardar Patel University,
Vallabh Vidyanagar 388 120, Gujarat, India
(Received 13 April 2012; in final form 30 July 2012)
A series of square planar mononuclear palladium(II) complexes, [Pd(Ln)(OMe)2] (where Ln ¼
4-(4-chlorophenyl)-6-phenyl-2,20-bipyridine, 4,6-bis(4-chlorophenyl)-2,20-bipyridine, 4-(4-bro-
mophenyl)-6-(4-chlorophenyl)-2,20-bipyridine, and 6-(4-chlorophenyl)-4-p-tolyl-2,20-bipyri-
dine), have been synthesized and characterized. Interaction of palladium complexes with
DNA has been investigated by electronic absorption spectra, viscosity measurements and DNA
melting temperature techniques. The metal complex-DNA interaction produced hypochromism
and bathochromism, suggesting intercalative binding. Viscosity data support the intercalation
mode of binding. Palladium(II) complexes promote cleavage of plasmid pUC19 DNA from the
supercoiled form to the open circular form. Antibacterial activities of the complexes have also
been evaluated against five pathogenic bacteria. The antimicrobial test results reveal that all
compounds have good antibacterial activity against both Gram(ꢀve) and Gram(þve) bacterial
species compared to the ligands and metal salt.
Keywords: Square planar; Palladium(II); Intercalation; pUC19; CT DNA
1. Introduction
Reactivity of palladium(II) complexes is of particular interest because these systems are
rapidly reacting prototypes for platinum(II) complexes [1]. Platinum compounds are the
most efficacious metal-based anticancer agents [2]. However, because of severe side
effects, drug resistance and the limited spectrum of tumors [2], there has been a search
for related complexes (e.g. Pd, Ru, and Au) that are able to improve effectiveness and
reduce side effects of platinum-based drugs [3, 4]. Special attention in this regard has
been given to palladium(II) complexes [5–8], since the coordination geometry and
complex forming processes of palladium(II) are very similar to those of platinum(II).
DNA intercalation is a reversible non-covalent interaction originally proposed by
Lerman [9] to account for the affinity of some small planar aromatic molecules for
double-stranded DNA. This interaction consists of insertion of a drug molecule
between two adjacent base pairs of the nucleic acid. Many workers [10] believe that
intercalation is an important intermediate state in mutagenesis and carcinogenesis
caused by a number of small molecules; it is also thought [11] that intercalation is part
*Corresponding author. Email: jeenen@gmail.com
Journal of Coordination Chemistry
ISSN 0095-8972 print/ISSN 1029-0389 online ß 2012 Taylor & Francis
http://dx.doi.org/10.1080/00958972.2012.727207

3834
M.N. Patel et al.
of the mode of action of some antibacterial and antineoplastic medicines. Because of
this, a large number of studies [12] on intercalation have been performed; studies on
DNA intercalation by square-planar complexes [13] have appeared, although system-
atic structural variations have not been reported.
Continuing our interest in metal complexes [14], we report synthesis and character-
ization of new Pd(II) complexes. The complexes have been tested for their antibacterial
activity against five microorganisms. All the complexes have been tested for their ability
to bind to calf-thymus (CT) DNA. The binding properties of the complexes with
CT DNA have been investigated with absorption titration, viscosity and DNA melting
temperature experiments. The cleavage properties of the complexes have been
performed using gel electrophoresis.
2. Experimental
2.1. Materials
Sodium tetrachloropalladate(II) was purchased from Chemport (India). p-Chloro
acetophenone, p-bromo acetophenone, p-methyl acetophenone, acetophenone, p-chloro
benzaldehyde, and 2-acetylpyridine were purchased from Spectrochem (Mumbai,
India). Agarose, EB, TAE (Tris-acetyl-EDTA), bromophenol blue and xylene cyanol
FF were purchased from Himedia (India). CT DNA was purchased from Sigma
Chemical Co. (India). Culture of pUC19 bacteria (MTCC 47) was purchased from the
Institute of Microbial Technology (Chandigarh, India).
2.2. Physical measurements
C, H, and N elemental analyses were performed with a VarioMICRO superuser
elementar analysensysteme GmbH. Infrared spectra were recorded on a Fourier
transform IR (FTIR) ABB Bomen MB 3000 spectrophotometer as KBr pellets from
.
4000 to 400 cmꢀ1 1H and 13C NMR spectra were recorded on a Bruker Avance
(400 MHz). Fast atomic bombardment mass spectra (FAB MS) were recorded on a Jeol
SX 102/Da–600 mass spectrophotometer/data system using Argon/Xenon (6 kV,
10 mA) as the FAB gas. The accelerating voltage was 10 kV and spectra were recorded
at room temperature. Electronic spectra were recorded on a UV–160 A UV-Vis
spectrophotometer, Shimadzu (Japan). Photo quantization of the gel after electropho-
resis was carried out on AlphaDigiDocTM RT. Version V.4.0.0 PC–Image software.
2.3. Synthesis of ligands
Substituted bipyridines, i.e. 4-(4-Xphenyl)-6-phenyl-2,20-bipyridines, were synthesized
by reacting 1-[2-oxo-2-(2-pyridyl)ethyl]pyridiniumiodide and 3-(4-Xphenyl)-1-phenyl-
prop-2-en-1-one [15]. The reactant 1-[2-oxo-2-(2-pyridyl)ethyl]pyridinium iodide was
prepared using 2-acetylpyridine, iodine, and pyridine [16], and 3-(4-Xphenyl)-1-
phenylprop-2-en-1-one (X ¼ chloro/bromo/methyl) was prepared via aldol condensa-
tion between substituted acetophenones and p-chloro benzaldehyde.

Square planar palladium(II)
3835
2.3.1. 4-(4-Chlorophenyl)-6-phenyl-2,20-bipyridine (L1). An excess of ammonium ace-
tate (approximately 10 equiv) was added to a mixture of 3-(4-chlorophenyl)-1-
phenylprop-2-en-1-one (2.88 mmol) and 1-[2-oxo-2-(2-pyridyl)ethyl]pyridinium iodide
(2.88 mmol) in methanol (20 mL). After refluxing for 5–7 h, the reaction mixture was
allowed to cool which led to the formation of greenish yellow needles of pure product.
The solid was filtered off, washed with cold methanol, dried under vacuum and further
recrystallized using hexane. A further crop of the desired product was obtained upon
refluxing the filtrate for another 5–6 h and subsequent workup.
Yield: 54%, m.p. 140ꢁC, mol. wt. 342.82. Anal. Calcd for C22H15ClN2 (%): C, 77.08;
H, 4.41; N, 8.17, found (%): C, 76.90; H, 4.47; N, 8.08. 1H NMR (CDCl3, 400 MHz): ꢀ
0
(ppm) 8.754–8.710 (complex, 2H, H3 ,6 ), 8.659 (s, 1H, H3), 8.220 (d, 2H, H2
0
000,6000
), 7.960
(s, 1H, H5), 7.910 (t, 1H, H4 ), 7.789 (d, 2H, H2 ,6 ), 7.578–7.474 (complex, 5H,
0
00 00
), 7.386 (t, 1H, H5 ), 13C NMR (CDCl3, 100 MHz): ꢀ (ppm) 157.43 (C6),
00 00
H3 ,5 ,3
0
000,4000,5000
0
155.86 (C2), 150.22 (C2 ), 149.09 (C4), 147.22 (C6 ), 139.28 (C1 ), 138.25 (C1 ), 137.08
0
0
000
0
000
00 00
00 00
000,5000
), 127.10 (C3 ,5 ),
000,6000
(C4 ), 129.92 (C4 ), 129.30 (C2
00
), 128.80 (C2 ,6 ), 128.55 (C3
0
124.08 (C4 ), 121.78 (C5 ), 118.40 (C3 ), 117.50 (C3), 102.75 (C5). FT-IR (KBr, cmꢀ1):
0
ꢁ(C–H), 3012 (m); ꢁ(C¼C), 1538 (s); ꢁ(C¼N), 1459 (s); ꢁ(C–H), 1047; ꢀ (C–H), 725 (s).
2.3.2. 4,6-Bis(4-chlorophenyl)-2,20-bipyridine (L2). The ligand was synthesized using
1,3-bis(4-chlorophenyl)prop-2-en-1-one. Yield: 51%, m.p. 149ꢁC, mol. wt. 377.27.
Anal. Calcd for C22H14Cl2N2 (%): C, 70.04; H, 3.74; N, 7.43, found (%): C, 69.96; H,
1
3.68; N, 7.36. H NMR (CDCl3, 400 MHz): ꢀ (ppm) 8.741 (dd, 1H, H3 ), 8.682–8.641
0
0
(complex, 2H, H6 ,5), 8.169 (d, 2H, H2
00 00
2H, H2 ,6 ), 7.531–7.510 (complex 4H H3 ,5 ,3
0
), 8.133–7.830 (complex, 2H, H3,4 ), 7.773 (d,
000,6000
), 7.413 (dt, 1H, H5 ), 13C NMR
00 00
0
000,5000
0
(CDCl3, 100 MHz): ꢀ (ppm) 156.51 (C6), 156.10 (C2), 155.97 (C2 ), 149.24 (C4), 149.09
0
(C6 ), 137.74 (C1 ), 137.10 (C4 ), 136.97 (C1 ), 135.34 (C4 ), 129.64 (C4 ), 129.28
00
0
000
000
00
00 00
00 00 0 0
000,6000
), 128.38 (C2 ,6 ), 124.04 (C5 ), 121.54 (C3 ),
000,5000
(C3
), 128.95 (C3 ,5 ), 128.53 (C2
117.97 (C3), 117.53 (C5), FT-IR (KBr, cmꢀ1): ꢁ(C–H), 3155 (m); ꢁ(C¼C), 1554 (s);
ꢁ(C¼N), 1457 (s); ꢁ(C–Cl), 1045; ꢀ (C–H), 717 (s).
2.3.3. 4-(4-Bromophenyl)-6-(4-chlorophenyl)-2,20-bipyridine (L3). The ligand was syn-
thesized using 3-(4-bromophenyl)-1-(4-chlorophenyl)prop-2-en-1-one. Yield: 59%, m.p.
156ꢁC, mol. wt. 421.72. Anal. Calcd for C22H14BrClN2 (%): C, 62.66; H, 3.35; N, 6.64,
found (%): C, 62.73; H, 3.29; N, 6.57. 1H NMR (CDCl3, 400 MHz): ꢀ (ppm) 8.732 (dd,
0
1H, H3 ), 8.662–8.616 (complex, 2H, H6 ,5), 8.148 (d, 2H, H2
0
00 00
), 7.870 (d, 2H, H2 ,6 ),
), 7.282 (s, 1H, H3), 7.249 (d, 2H,
000,6000
0
7.803–7.772 (complex, 2H, H4 ,5 ), 7.502 (d, 2H, H3
0
000,5000
13
00 00
H3 ,5 ). C NMR (CDCl3, 100 MHz): ꢀ (ppm) 156.43 (C6), 156.02 (C2), 155.94 (C2 ),
0
0
00
0
000
000
149.34 (C4), 149.10 (C6 ), 137.75 (C1 ), 136.91 (C4 ), 135.27 (C1 ), 134.68 (C4 ), 129.05
00 00
00 00 0
000,6000
), 123.97 (C2 ,6 ), 121.47 (C5 ),
000,5000
(C5), 128.97 (C3
0
), 128.92 (C3 ,5 ), 128.32 (C2
00
117.93 (C3 ), 117.55 (C3), 116.16 (C4 ), FT-IR (KBr, cmꢀ1): ꢁ(C–H), 3029 (m); ꢁ(C¼C),
1555 (s); ꢁ(C¼N), 1516 (s); ꢁ(C–Cl), 1049 (s); ꢁ(C–Br), 1025; ꢀ (C–H), 750.
2.3.4. 6-(4-Chlorophenyl)-4-p-tolyl-2,20-bipyridine (L4). The ligand was synthesized
using 1-(4-chlorophenyl)-3-p-tolylprop-2-en-1-one. Yield: 57%, m.p. 138ꢁC, mol. wt.
356.11. Anal. Calcd for C23H17ClN2 (%): C, 77.41; H, 4.80; N, 7.85, found (%): C,
1
77.35; H, 4.75; N, 7.79. H NMR (CDCl3, 400 MHz): ꢀ (ppm) 8.743–8.695 (complex,

3836
M.N. Patel et al.
0
2H, H3 ,6 ), 8.598 (s, 1H, H5), 8.122 (d, 1H, H2
0
0
), 7.930–7.870 (complex, 2H, H3,4 ),
000,6000
00 00
00 00
0
000,5000
7.775 (d, 2H, H2 ,6 ), 7.508 (d, 2H, H3
), 7.385–7.350 (complex, 3H, H3 ,5 ,5 ), 2.470
0
(s, 3H, methyl), 13C NMR (CDCl3, 100 MHz): ꢀ (ppm) 157.32 (C6), 156.31 (C2 ), 156.25
0
0
00
0
000
000
(C2 ), 149.02 (C4), 148.92 (C6 ), 139.24 (C1 ), 137.33 (C4 ), 136.92 (C1 ), 136.53 (C4 ),
0
00 00
00 00 0
000,6000
), 126.95 (C2 ,6 ), 123.87 (C5 ),
000,5000
135.14 (C3 ), 129.50 (C3
00
), 129.21 (C3 5 ), 128.53 (C2
121.56 (C4 ), 117.86 (C3), 116.96 (C5), 21.33 (methyl), FT-IR (KBr, cmꢀ1): ꢁ(C–H),
3063 (m); ꢁ(C¼C), 1574 (s); ꢁ(C¼N), 1535 (s); ꢁ(C–Cl), 1045 (s); ꢀ (C–H), 733 (m).
2.4. Synthesis of complexes
2.4.1. [Pd(L1)(OMe)2] (1). All the complexes were synthesized by following the
literature procedure [17]. 4-(4-Chlorophenyl)-6-phenyl-2,20-bipyridine (0.5 mmol) in
methanol and Na2PdCl4 (0.5 mmol) in distilled water (50 mL) were refluxed for 5 h.
Reaction mixture was allowed to cool to room temperature and filtered. The obtained
yellow product was washed with hot water and dried under vacuum. General reaction
scheme for the synthesis of complexes is given in scheme 1. Yield: 63.7%, m.p.: 205ꢁC,
anal. Calcd for C24H21ClN2O2Pd, m.w. (%): 511.31, elemental anal. Calcd (%):
C, 56.38; H, 4.14; N, 5.48; Pd, 20.81, found (%): C, 56.29; H, 4.08; N, 5.52; Pd, 20.73.
FT-IR (KBr, cmꢀ1): ꢁ(C–H) 3055 (m); ꢁ(C¼C) 1574 (s); ꢁ(C¼N) 1489 (s); ꢁ(OMe),
1250; ꢁ(C–Cl), 1040; ꢀ (C–H) 741 (s); ꢁ(M–O), 589; ꢁ(M–N) 545 (s).
2.4.2. [Pd(L2)(OMe)2] (2). Compound 2 was prepared using 4,6-bis(4-chlorophenyl)-
2,20-bipyridine. Yield: 63.5%, m.p.: 217ꢁC, anal. Calcd for C24H20Cl2N2O2Pd, m.w.
(%): 545.75, elemental anal. Calcd (%): C, 52.82; H, 3.69; N, 5.13; Pd, 19.50, found
(%): C, 52.88; H, 3.57; N, 5.08; Pd, 19.43. FT-IR (KBr, cmꢀ1): ꢁ(C–H) 3055 (m);
Scheme 1. General reaction scheme for synthesis of ligand and its Pd(II) complexes.

Square planar palladium(II)
3837
ꢁ(C¼C) 1582 (s); ꢁ(C¼N) 1481 (s); ꢁ(OMe), 1257; ꢁ(C–Cl), 1035; ꢀ (C–H) 733 (s);
ꢁ(M–O), 585; ꢁ(M–N) 543 (s).
2.4.3. [Pd(L3)(OMe)2] (3). Compound 3 was prepared using 4-(4-bromophenyl)-6-
(4-chlorophenyl)-2,20-bipyridine. Yield: 62.9%, m.p.: 227ꢁC, anal. Calcd for
C24H20BrClN2O2Pd, m.w. (%): 590.20, elemental anal. Calcd (%): C, 48.84; H, 3.42;
N, 4.75; Pd, 18.03, found (%): C, 48.88; H, 3.35; N, 4.68; Pd, 17.96. FT-IR (KBr,
cmꢀ1): ꢁ(C–H) 3055 (m); ꢁ(C¼C) 1605 (s); ꢁ(C¼N) 1543 (s); ꢁ(OMe), 1242; ꢁ(C–Cl),
1042; ꢁ(C–Br), 1022 ꢀ (C–H) 771 (s); ꢁ(M–O), 640; ꢁ(M–N), 550 (s).
2.4.4. [Pd(L4)(OMe)2] (4). Compound 4 was prepared using 6-(4-chlorophenyl)-4-p-
tolyl-2,20-bipyridine. Yield: 65.3%, m.p.: 217ꢁC, anal. Calcd for C25H23ClN2O2Pd,
m.w. (%): 525.34, elemental anal. Calcd (%): C, 57.16; H, 4.41; N, 5.33; Pd, 20.26,
found (%): C, 57.23; H, 4.35; N, 5.27; Pd, 20.19. FT-IR (KBr, cmꢀ1): ꢁ(C–H) 3055 (m);
ꢁ(C¼C), 1597 (s); ꢁ(C¼N) 1551 (s); ꢁ(OMe), 1227; 1043; ꢁ(C–Cl), 1043; ꢀ (C–H) 741 (s);
ꢁ(M–O), 594; ꢁ(M–N), 555 (s).
2.5. Antibacterial activity
The in-vitro antibacterial activities of metal salt, ligands, and palladium complexes were
tested against three Gram(ꢀve); Serratia marcescens (MTCC 7103), Escherichia coli
(MTCC 433), Pseudomonas aeruginosa (MTCC P09), and two Gram(þve);
Staphylococcus aureus (MTCC 3160), Bacillus subtilis (MTCC 7193), microorganisms.
Solution of the complexes was prepared in DMSO. The minimum inhibitory
concentration of each compound was defined as the lowest concentration exhibiting
no visible growth. Luria Broth was employed as the bacteria growth medium. Serial
concentrations of each of the tested compounds from 71 to 2800 mmol Lꢀ1 were
prepared for the determination of MIC and inoculated with five different microorgan-
isms. After incubation at 37ꢁC for 24 h, the solution without turbidity was determined
as the minimum inhibitory concentration.
2.6. Absorption titration
The UV absorbance at 260 and 280 nm of the CT DNA solution in 5 mmol Lꢀ1
Tris-HCl buffer (pH 7.2) gave a ratio of ꢂ1.9, indicating the DNA was free of protein.
The concentration of CT DNA was measured from the band intensity at 260 nm with
the known " value (6600 mol Lꢀ1 cmꢀ1). Absorption titration experiments were
performed by maintaining the palladium complex concentration constant at
20 mmol Lꢀ1 while varying the concentration of CT DNA within 50–150 mmol Lꢀ1
.
While measuring the absorption spectra, an equal quantity of CT DNA was added to
both the complex solution and the reference solution to eliminate the absorbance of CT
DNA itself. From the absorption data, the intrinsic binding constant Kb was
determined from a plot of [DNA]/("a ꢀ "f) versus [DNA] using the following equation:
½DNAꢃ
½DNAꢃ
1
¼
þ
ð"a ꢀ "fÞ ð"b ꢀ "fÞ Kbð"b ꢀ "fÞ

3838
M.N. Patel et al.
where [DNA] is the concentration of CT DNA in base pairs. The apparent absorption
coefficients "a, "f, and "b correspond to Aobsd./[Pd(II)], the extinction coefficient for
the free palladium(II) complex and extinction coefficient for the palladium(II) complex
in the fully bound form, respectively [18]. Kb is given by the ratio of slope to the
intercept.
2.7. DNA melting experiments
DNA melting experiments were carried out by monitoring the absorption intensity of
CT DNA (100 mmol Lꢀ1) at 260 nm at various temperatures, both in the absence and
presence of the palladium(II) complexes (20 mmol Lꢀ1). Measurements were carried out
using a Perkin–Elmer Lambda 35 spectrophotometer equipped with a Peltier
temperature-controlling programmer (PTP 6) (ꢄ0.1ꢁC) increasing the temperature of
the solution by 0.5ꢁCꢀ1
.
2.8. Viscosity measurements
Viscometric titrations were performed with a Cannon-Ubbelohde viscometer.
The viscometer was thermostated at 27ꢁC (ꢄ0.1) in a constant temperature bath.
The concentration of CT DNA was 200 mmol Lꢀ1 in NP and flow times were
measured with an automated timer; each sample was measured three times and an
average flow time was calculated. Data were presented as (ꢂ/ꢂ0)1/3 versus [complex]/
[DNA], where ꢂ is the viscosity of DNA in the presence of complex and ꢂ0
that of DNA alone. Viscosity values were calculated from the observed flowing
time of DNA-containing solutions (t) corrected for that of buffer alone (t0),
ꢂ ¼ (t ꢀ t0).
2.9. Gel electrophoresis technique
Gel electrophoresis of plasmid DNA (pUC19 DNA) was carried out in TE buffer with
15 mL reaction mixture containing 300 mg mLꢀ1 plasmid DNA (10 mmol Lꢀ1 Tris,
1 mmol Lꢀ1 EDTA, pH 8.0) and 150 mmol Lꢀ1 complex. Reactions proceeded for 3 h at
37ꢁC. All reactions were quenched by the addition of 5 mL loading buffer (40% sucrose,
0.2% bromophenol blue). The aliquots were loaded onto 1% agarose gel and
electrophoreses occurred at 50 V in 1X TAE buffer. Gel was stained with 0.5 mg m Lꢀ1
ethidium bromide and photographed on a UV illuminator. The percentage of each form
of DNA was quantified using AlphaDigiDocTM RT Version V.4.1.0 PC–Image
software. The degree of DNA cleavage activity was expressed as percentage cleavage of
SC-DNA according to the following equation:
ð% of SC ꢀ DNAÞ control ꢀ ð% of SC ꢀ DNAÞ sample
%DNA cleavage ¼
ð% of SC ꢀ DNAÞ control

Square planar palladium(II)
3839
3. Results and discussion
3.1. Electronic spectra and magnetic moments
In electronic spectra, the low-spin square-planar d8 metal complexes are expected to
give three spin allowed d–d transitions from the lower laying dz2 ! dxz, dyz, dz2 ! dxy,
and dz2 ! dx2–y2 [19].
In the synthesized palladium(II) complexes, only one d–d band is observed at
ꢂ438 nm, which is assigned to the dz2 ! dx2ꢀy2 transition, with the other two bands
masked by high intensity charge transfer bands. The bands at ꢂ280 and ꢂ260 nm
correspond to charge transfer transitions. All these bands point to low-spin complex of
the d8 system with square planar geometry. The synthesized complexes were diamagnetic.
3.2. IR spectra
Aromatic C–H stretching and out of plane bending vibrations give bands at 740 and
3050 cmꢀ1. Very strong bands at ꢂ1500 and ꢂ1550 cmꢀ1 for the C¼N and C¼C highest-
energy pyridine-ring vibration of the ligands are shifted by 30–40 cmꢀ1 to higher
frequencies after complexation. The band for OMe is observed at ꢂ1250 cmꢀ1 and
indicates OMe in the compound. The coordinate covalent bonding of the metal via
nitrogen of bipyridine is supported by a sharp band at ꢂ550, characteristic of Pd–N,
suggesting that nitrogen of pyridine is coordinated [20]. The characteristic band for Pd–O
is at ꢂ600 cmꢀ1, which suggests that the OMe is directly attached to palladium [21].
3.3. Mass spectrometry
FAB MS spectrum of 1 was obtained using m–nitro benzyl alcohol as matrix
(Supplementary material). Peaks at 136, 137, 154, 289, and 307 m/z values are of the
matrix. The molecular ion peak for complex is observed at 512 m/z (I) and peaks
corresponding to the fragments [M–OMe]þ, [M–2(OMe)]þ, and 4-(4-chlorophenyl)-6-
phenyl-2,20-bipyridine are observed at 481 (II), 448 (III), and 342 (IV) m/z, respectively.
The fragmentations of phenyl and p-chlorophenyl from 4-(4-chlorophenyl)-6-phenyl-
2,20-bipyridine shows peaks at 265 (V) and 231 (VI) m/z, respectively. The peak
corresponding to 2,20-bipyridine is observed at 154 (VII) m/z. Doublets exist at
512 : 514, 481 : 483, 448 : 450, and 265 : 267 m/z, which correspond to isotopic patterns of
chlorine. Moreover, peaks due to species containing the isotope of palladium were
easily detected at m/z: 508, 510, 511, 512, 514, and 516 for fragment I; 476, 478, 480,
481, 483, and 485 for fragment II; and 444, 446, 447, 448, 450, and 452 for fragment III
of 102Pd, 104Pd, 105Pd, 106Pd, 108Pd, and 110Pd, respectively (figure 1) [22]. The mass
fragmentation pattern is shown in ‘‘Supplementary material.’’
3.4. Antibacterial activity
A comparative study of the MIC values of the ligands and their complexes indicates
that the metal complexes show better activity against three Gram(ꢀve) and two
Gram(þve) microorganisms compared to the bidentate ligands and metal salts (table 1).
Complexes 1 and 2 show better antibacterial activity than 3 and 4. Complex 2 is more

3840
M.N. Patel et al.
Figure 1. Experimental and theoretical isotope patterns of FAB-MS for 1.
Table 1. Minimum inhibitory concentration of palladium complexes (mmol Lꢀ1).
Compounds
S. aureus
B. subtilis
S. marcescens
P. aeruginosa
E. coli
Na2PdCl4
2238.00
610
540
610
670
88
77
82
99
2490.00
705
510
595
715
89
74
84
104
2145.00
670
585
650
720
91
71
82
102
1940.00
645
490
625
720
92
79
81
98
2820.00
675
535
635
725
88
78
80
99
Ligand 1 (L1)
Ligand 2 (L2)
Ligand 3 (L3)
Ligand 4 (L4)
[Pd(L1)(OMe)2] (1)
[Pd(L2)(OMe)2] (2)
[Pd(L3)(OMe)2] (3)
[Pd(L4)(OMe)2] (4)
potent because of the presence of two electronegative chlorines in the bidentate ligand.
The increase in antibacterial activity of the metal chelate can be explained by chelation
theory [23]. Our complexes show higher antibacterial activity than Pd(II) complexes of
tetracycline, doxycycline and chlortetracycline (MIC ¼ 2.08–66.50 mmol Lꢀ1 against
E. Coli) reported by Guerra et al. [24], but the complexes are less active than
ciprofloxacin. Palladium complexes may deactivate various cellular enzymes, which
play a role in various metabolic pathways of organisms [25].
3.5. Absorption titration
Absorption titration is a common way to investigate the interactions of complexes with
DNA. We have used absorption titration method to monitor interaction of 1–4 with CT
DNA. Intercalation of a complex to DNA generally results in hypochromism and red
shift (bathochromism) (figure 2) of the absorption band due to strong ꢃ–ꢃ* stacking
interaction between the aromatic chromophore of the ligand and the base pairs of DNA
[14]. The extent of hypochromism, thus gives an estimate of the strength of
intercalation. The binding constants of the palladium complexes are 2.31 ꢅ 104,
4.76 ꢅ 104, 3.47 ꢅ 104, and 1.24 ꢅ 104 for 1, 2, 3, and 4, respectively. The complexes
show higher binding constant values than [Pd(en)(8-QO)]Cl (1.7 ꢅ 103 at 300 K and
1.6 ꢅ 103 at 310 K) reported by Mansouri-Torshizi et al. [26].
3.6. DNA denaturation experiments
The binding of molecules into the double helix is known to increase the melting
temperature. The melting temperature is the temperature at which the double helix

Square planar palladium(II)
3841
Figure 2. Absorption spectral traces of 1 with increasing amount of DNA in Tris-HCl buffer (50 mmol Lꢀ1
Tris-HCl, pH 7.2). [complex] ¼ 20 mmol Lꢀ1, [DNA] ¼ 50–150 mmol Lꢀ1 with incubation period of 15 min at
37ꢁC. Plots of [DNA]/("a ꢀ "f) vs. [DNA] for the titration of DNA with palladium(II) complexes.
Figure 3. DNA melting temperature curves of CT DNA and palladium complexes in Tris-HCl buffer
(50 mmol Lꢀ1 Tris-HCl, pH 7.2).
is denatured into single-stranded DNA. The extinction coefficient of DNA bases at
260 nm in the double-helical form is much lower than in the single-stranded form.
Hence, melting of the helix leads to an increase in absorption at this wavelength.
Thus, the transition temperature from helix to coil can be determined by
monitoring the absorbance of the DNA base at 260 nm as a function of
temperature.
The thermal denaturation experiment carried out for CT DNA in the absence of
Pd(II) complexes revealed a Tm of 74.2ꢁC under our experimental conditions (figure 3).
On addition of the complexes, Tm of DNA increased to 78.1, 79.1, 78.6, and 77.8 for 1,
2, 3, and 4, respectively, at a concentration ratio [DNA]/[Pd] of 5 : 1. The increase
(3.6–4.9) in Tm is comparable to that observed for classical intercalators [27]. Such
DNA melting temperature studies of some mixed-ligand palladium(II) complexes were
studied by Tercero et al. [28], who reported that the DTm of the complexes increased by
ꢂ9.3ꢁC and suggested the intercalative mode of binding.

3842
M.N. Patel et al.
Figure 4. Effect on relative viscosity of DNA under the inflꢀu1ence of increasing amount of ethidium bromide
and complexes at 27 ꢄ 0.1ꢁC in Tris-HCl buffer (50 mmol L Tris-HCl, pH 7.2).
3.7. Viscosity measurements
To investigate further, the binding modes of the complexes, viscosity measurements on
solutions of CT DNA incubated with the complexes were carried out. Because the
viscosity of a DNA solution is sensitive to the addition of drugs and metal complexes
bound by intercalation, we examined the effect on the specific relative viscosity of DNA
upon addition of the complexes. Since the relative specific viscosity (ꢂ/ꢂ0) (ꢂ and ꢂ0 are
the specific viscosities of DNA in the presence and absence of the complexes,
respectively) of DNA reflects the increase in contour length associated with separation
of DNA base pairs caused by intercalation, a classical intercalator such as ETBr could
cause a significant increase in viscosity of DNA solutions. In contrast, a partial and/or
non-classical intercalation could bend or kink DNA resulting in a decrease in its
effective length with a concomitant increase in its viscosity [29]. The plots of relative
viscosities with [Pd]/[DNA] are shown in figure 4. The relative viscosity of DNA
increases with increase in the concentration of the complexes but less compared to that
of potential classical intercalators, e.g. ethidium bromide. Similar results were obtained
for Pd(II) complexes reported by Ghosh et al. [30] and Nyarko et al. [31]. This is
consistent with the observed trend by other optical methods and suggests primarily
intercalation of the complex.
3.8. Cleavage of pUC19 plasmid DNA by palladium(II) complexes
Agarose gel electrophoresis is a convenient method to assess cleavage of DNA by metal
complexes to assess the factors affecting the nucleolytic efficiency of a complex and to
compare the nucleolytic property of different metal complexes. This has been done in
this study with the [Pd(Ln)(OMe)2] complexes. DNA cleavage experiments were
performed by agarose gel electrophoresis using plasmid pUC19 DNA (300 mmol Lꢀ1) in
10 mmol Lꢀ1 Tris/1 mmol Lꢀ1 NaCl buffer (pH 8) and the palladium(II) complexes
(figure 5). DNA cleavage data are given in table 2. The complexes show complete
conversion of the SC (form I) to its nicked-circular form (NC, form II). Control
experiments with DNA or palladium salt alone do not show any apparent conversion of
SC to its NC form. Such cleavage ability of plasmid DNA by Pd(II) complexes was
reported by Perez-Cabre et al. [32] and Yodoshi and Okabe [33].
´ ´

Square planar palladium(II)
3843
Figure 5. Photogenic view of interaction of pUC19 DNA (300 mg mLꢀ1) with palladium(II) complexes
(200 mmol Lꢀ1) using 1% agarose gel containing 0.5 mg mLꢀ1 ethidium bromide. All reactions were incubated
in TE buffer (pH 8) in a final volume of 15 mL for 3 h at 37ꢁC: Lane 1, DNA control; Lane 2, Na2PdCl4;
Lane 3, [Pd(clpbpy)(OMe)2] (1); Lane 4, [Pd(clcpbpy)(OMe)2] (2); Lane 5, [Pd(brcpbpy)(OMe)2] (3); and Lane
6, [Pd(clmpbpy)(OMe)2] (4).
Table 2. Gel electrophoresis data of Pd(II) complexes.
Compounds
SC (%)
OC (%)
Cleavage (%)
DNA control
DNA þ Na2PdCl4
DNA þ 1
88.2
70.8
29.1
17.4
23.1
36.9
11.8
20.8
70.8
82.6
76.1
43.1
–
19.77
67.00
80.27
73.80
58.16
DNA þ 2
DNA þ 3
DNA þ 4
4. Conclusion
Synthesis and characterization of four Pd(II) complexes with different bipyridines has
been achieved. All complexes were more bacteriostatic than bipyridines and metal salt
against five microorganisms. Binding behavior of the palladium chelates of bipyridines
was subtly but distinctly different, depending on the structure of the bipyridines. As the
electronegativity of the substituted group on bipyridine skeleton increases, the
interaction becomes stronger. Increasing order for the DNA interaction behavior of
synthesized complexes is 4 5 1 5 3 5 2 on the basis of Kb values, change in relative
viscosity, change in melting temperature and the plasmid cleavage study.
Acknowledgments
Authors thank Head, Department of Chemistry, Sardar Patel University, India, for
making it convenient to work in the laboratory and U.G.C. for providing financial
support under ‘‘UGC Research Fellowship for Meritorious Students’’ scheme.
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