Copper complexes of pyridyl–tetrazole ligands with pendant amide and hydrazide arms: synthesis, characterization, DNA-binding and antioxidant properties

Article abstract of DOI:10.1007/s11243-016-0047-2

Pyridyl–tetrazole ligands 2-(5-(pyridin-2-yl)-1H-tetrazol-1-yl)acetamide (L1), 2-(5-(pyridin-2-yl)-2H-tetrazol-2-yl)acetamide (L2), 2-(5-(pyridin-2-yl)-1H-tetrazol-1-yl)acetohydrazide (L3) and 2-(5-(pyridin-2-yl)-2H-tetrazol-2-yl)acetohydrazide (L4) have been prepared and coordinated with CuCl₂·2H₂O to furnish the corresponding complexes [Cu(L1)₂]–[Cu(L4)₂]. EPR spectra of the complexes are characteristic of square planar geometries, with nuclear hyperfine spin 3/2. DNA-binding studies using UV–Vis absorption spectroscopy, viscosity and thermal denature studies revealed that all of these complexes are avid binders of calf thymus DNA. The antioxidant properties of the free ligands and the Cu(II) complexes were investigated using the p-nitrosodimethyl aniline hydroxyl radical scavenging method, and [Cu(L4)₂] was found to show the highest activity.

Full text of DOI:10.1007/s11243-016-0047-2

Transition Met Chem
DOI 10.1007/s11243-016-0047-2
Copper complexes of pyridyl–tetrazole ligands with pendant
amide and hydrazide arms: synthesis, characterization,
DNA-binding and antioxidant properties
1
1
1
•
Santhosh Reddy Kasi Reddy
1
Phani Raja Kanuparthy
•
Surendrababu Manabolu Surya
•
Mustafa Shaik
Received: 31 December 2015 / Accepted: 24 March 2016
Ó Springer International Publishing Switzerland 2016
Abstract Pyridyl–tetrazole ligands 2-(5-(pyridin-2-yl)-
1
biochemical and pharmaceutical applications by virtue of
their high physiological activity and low toxicity [3].
Tetrazoles are important tools in synthetic organic chemistry
and also used as precursors of carbenes in flash pyrolysis [4].
The pharmacological applications of tetrazoles with gly-
cosidase inhibitory, antihypertensive, anti-inflammatory,
antibacterial, antifungal, analgesic, antinociceptive, anti-
cancer, anticonvulsant, antidiabetic, antiulcer and antitu-
bercular activities have been reported [5]. Tetrazoles play a
significant role in coordination chemistry as ligands, as
metabolically stable surrogates for carboxylic acid groups in
medical chemistry and as high-energy materials [6]. Tetra-
zole resembles the carboxylic group in its acidic character-
istics and is metabolically stable [7]. Applications of
5-substituted-1H-tetrazoles as lipophilic spacers have been
reported [8]. In addition, pyridines are associated with
diverse biological activities [9]. In continuation of our
ongoing research [10], we have prepared pyridyl tetrazoles
with pendant acetamide and acetohydrazide groups
H-tetrazol-1-yl)acetamide (L1), 2-(5-(pyridin-2-yl)-2H-
tetrazol-2-yl)acetamide (L2), 2-(5-(pyridin-2-yl)-1H-tetra-
zol-1-yl)acetohydrazide (L3) and 2-(5-(pyridin-2-yl)-2H-
tetrazol-2-yl)acetohydrazide (L4) have been prepared and
coordinated with CuCl Á2H O to furnish the corresponding
2
2
complexes [Cu(L1) ]–[Cu(L4) ]. EPR spectra of the
2
2
complexes are characteristic of square planar geometries,
with nuclear hyperfine spin 3/2. DNA-binding studies using
UV–Vis absorption spectroscopy, viscosity and thermal
denature studies revealed that all of these complexes are
avid binders of calf thymus DNA. The antioxidant prop-
erties of the free ligands and the Cu(II) complexes were
investigated using the p-nitrosodimethyl aniline hydroxyl
radical scavenging method, and [Cu(L4) ] was found to
2
show the highest activity.
Introduction
(Scheme 1) and investigated their copper(II) complexes
Tetrazoles exhibit various biological activities and act as a
pharmacophore for the carboxylate group [1]. Recently,
there has been a growing demand for pyridyl tetrazoles as
components of metal–organic frameworks (MOFs), which
have attracted much attention in the past decade as a result of
their interesting structural topographies as well as their
useful properties and applications, including storage of
gases, catalysis, drug delivery, magnetism and luminescence
(Scheme 2). The results of our investigations are reported in
this paper.
Experimental
Materials and measurements
[
2]. The metal complexes of tetrazoles find a wide range of
Picolinonitrile 1 was purchased from Sigma-Aldrich. The
solvents used in the synthesis of the ligands and their
complexes were distilled before use. All other chemicals
were of AR grade and were used without further purifica-
tion. Hydrazine hydrate is a hazardous compound, and its
MSDS data sheet was referred to prior to its use. All
melting points were obtained on an Elico instrument, India
&
Mustafa Shaik
mustaff_02@yahoo.co.in
1
Department of Chemistry, GITAM University Hyderabad
Campus, Hyderabad 502329, India
123

Transition Met Chem
Scheme 1 Synthesis of L1–L4.
Reagents and conditions:
a NaN , LiCl, NH Cl, DMF,
reflux, 10 h; b ethylbromo
acetate, DMF, 70 °C, 8 h; c,
d aq. NaOH (1 N), MeOH, r.t.,
O
H
N
b
OEt
N
N
a
N
3
4
N
N
N
N
N
+
h
N
N
N
CN
OEt
N
3
N
N
2
N
1
4
O
c
6
pyridine, (NH CO , dioxane,
h; e, f BOC anhydride,
g
d
4
)
2
3
O
r.t., 8 h; g, h hydrazine hydrate,
EtOH, 80 °C, 10 h
OH
N
N
N
N
N
N
N
N
N
N
N
N
O
OH
5
N
N
N
NH NH2
6
NH NH2
L4
O
e
N
O
N
L3
f
N
O
N
N
NH2
N
N
N
N
N
N
N
N
N
L1
N
NH2
L2
O
R
N
was washed successively with saturated NaHCO (50 ml)
3
R
and then water (40 ml 9 3) followed by brine solution
N
N
N
N
(40 ml). The organic layer was dried over MgSO and
4
N
N
N
N
N
N
N
N
filtered, and the solvent was evaporated under reduced
pressure to afford a gummy brown liquid, which was
purified by column chromatography using 17 % EtOAc in
hexane (V/V) to afford the esters 3 and 4.
Cu
Cu
N N
N
N
N
N
R
R
N
Ethyl 2-(5-(pyridin-2-yl)-1H-tetrazol-1-yl)acetate (3) [2]
Yellowish solid; Yield: 0.45 g (28 %). Yellow solid. M.p.
[
[
Cu(L1)2]
Cu(L3)2]
[Cu(L2)2]
[Cu(L4)2]
5
4–57 °C. Anal. Calc. for C H N O (233.23): % of C,
1
0 11 5 2
5
1.5; H, 4.7; N, 30.0. Found: % of C, 51.5; H, 4.7; N, 30.0.
1
H NMR (CDCl , 300 MHz): d 8.66 (d, 1H, J = 3.3 Hz),
Scheme 2 Structural formulae of the complexes. L1, L2, R =
CH N(O)NH ; L3, L4, R = –CH N(O)NHNH
3
–
2
2
2
2
8.43 (d, 1H, J = 8.1 Hz), 7.92 (dt, 1H, J = 7.8, 1.8 Hz),
7.48–7.41 (m, 1H), 5.75 (s, 2H), 4.19 (q, 2H, 6.9 Hz), 1.21
(t, 3H, J = 6.9 Hz) ppm.
(
Model MP96), and are uncorrected. Mass spectra were
Ethyl 2-(5-(pyridin-2-yl)-2H-tetrazol-2-yl)acetate (4) [2]
Yellowish solid Yield: 0.80 g (50 %). White solid. M.p.
obtained on a PeSciex API 2000 eV spectrometer and
1
13
Q1MSQ1/auto-injection mass spectra. H and C NMR
spectra were recorded on a Bruker Top Spin Instrument. IR
spectra were recorded using an Alpha T OPUS spectrom-
eter. Elemental analysis (C, H and N) was obtained using a
PerkinElmer 2400 elemental analyzer. Magnetic moments
were determined in the polycrystalline state on a PAR
model-155 vibrating sample magnetometer operating at
field strength of 2–8 kg. High-purity Ni metal (saturation
9
4–98 °C. Anal. Calc. for C H N O (233.23): % of C,
1
0 11 5 2
5
1.5; H, 4.7; N, 30.0. Found: % of C, 51.5; H, 4.7; N, 30.0.
1
H NMR (CDCl , 300 MHz): d 8.79 (d, 1H, J = 4.8 Hz),
3
8.28 (d, 1H, J = 7.8 Hz), 7.88 (td, 1H, J = 7.8, 1.8 Hz),
7.42 (m, 1H), 5.50 (s, 2H), 4.29 (q, 2H, J = 7.2 Hz), 1.29
(t, 3H, J = 7.2 Hz) ppm.
-
1
moment 55 emu g ) was used as standard. EPR spectra
were recorded on a Varian E-122 X-band spectrometer at
liquid nitrogen temperature in DMF.
General procedure for the synthesis of 5 and 6
To a methanolic (21 ml) solution of ester 3 or 4 (0.5 g,
2.14 mmol), 2 ml of aq. NaOH (1 N) was added and the
General procedure for the synthesis of 3 and 4
resulting solution was stirred for 6 h at room temperature,
resulting in the formation of a white precipitate. The
reaction was quenched with 3–4 drops of acetic acid, and
10 ml of ethyl acetate was added. The mixture was filtered
and the residue washed with ethyl acetate (20 ml); the
colorless residue was collected and dried in air.
To a solution of tetrazole 2 (1 g, 6.8 mmol) in DMF
(
15 ml) was added ethylbromo acetate (0.75 ml,
.8 mmol). The mixture was stirred for 8 h at 70 °C and
then diluted with ethyl acetate (50 ml). The organic layer
6
1
23

Transition Met Chem
2
-(5-(Pyridin-2-yl)-1H-tetrazole-1-yl)acetic acid (5):
Colorless solid; Yield: 400 mg (91 %). M.p. 284–287 °C.
General procedure for the synthesis of L3 and L4
Anal. Calc. for C H N O (205.17): % of C, 46.8; H, 3.4;
7 5 2
1
N, 34.1. Found: % of C, 46.8; H, 3.4; N, 34.1. H NMR
To an ethanolic solution (10 ml) of ester 3 or 4 (0.5 g,
2.14 mmol), hydrazine hydrate (0.2 ml, 4.28 mmol) was
added, and the resulting solution was stirred for 10 h at
80 °C. The solvent was removed under reduced pressure,
and the crude material was triturated with diethyl ether to
afford a colorless solid.
8
(
CDCl , 300 MHz): d 8.72 (t, 1H, J = 4.8,3.9 Hz), 8.12 (d,
H, J = 7.8 Hz), 8.03 (m, 1H), 7.58 (m, 1H), 5.45 (s, 2H)
3
ppm. C NMR (CDCl , 65 MHz): d 172.5, 153.1, 149.9,
3
1
1
3
1
43.2, 138.4, 126.3, 124.4, 52.9. ESI–MS: m/z 204
M - 1).
-(5-(Pyridin-2-yl)-2H-tetrazole-2-yl)acetic acid (6):
(
2-(5-(Pyridin-2-yl)-1H-tetrazol-1-yl)acetohydrazide (L3):
Colorless solid; Yield: 0.32 g (68 %). M.p. 185–187 °C. Anal.
2
Colorless solid; Yield: 0.38 g (86 %). M.p. 277–279 °C.
Anal. Calc. for C H N O (205.17): % of C, 46.8; H, 3.4;
1
N, 34.1. Found: % of C, 46.8; H, 3.4; N, 34.1. H NMR
Calc. for C H N O (219.20): % of C, 43.8; H, 4.1; N, 44.7.
8 9 7
1
Found: % of C, 43.8; H, 4.1; N, 44.7. H NMR (DMSO-D6,
300 MHz): d 9.48 (br-s, 1H–NH), 8.72 (d, 1H, J = 4.5 Hz),
8.30 (d, 1H, J = 12.9 Hz), 8.08 (dt, 1H, J = 7.8, 1.8 Hz), 7.62
(dd, 1H, J = 4.8, 1.8 Hz), 5.64 (s, 2H), 4.29 (br-s, 2H–NH2)
8
7 5 2
(
CDCl , 300 MHz): d 8.73 (d, 1H, J = 4.5 Hz), 8.11 (d,
H, J = 8.1 Hz), 7.98 (td, 1H, J = 7.5, 1.5 Hz), 7.52 (m,
1
H), 4.97 (s, 2H) ppm. C-NMR (CDCl , 75 MHz): d
3
1
1
1
5
3
13
ppm. C-NMR (DMSO-D6, 75 MHz): d 166.7, 152.6, 149.7,
3
72.19, 164.49, 150.15, 145.47, 139.25, 126.59, 123.58,
144.2, 138.2, 125.9, 123.7, 48.6 ppm; MS: m/z 220 (M ? 1).
2-(5-(Pyridin-2-yl)-2H-tetrazol-2-yl)acetohydrazide (L4):
Colorless solid; Yield: 0.32 g (68 %). M.p. 168–171 °C.
6.71 ppm. ESI–MS: m/z 204 (M - 1).
General procedure for the synthesis of L1 and L2
Anal. Calc. for C
8 9 7
H N O (219.20): % of C, 43.8; H, 4.1; N,
1
4.7. Found: % of C, 43.8; H, 4.1; N, 44.7. H NMR
4
To a solution of acid 5 or 6 (0.5 g, 2.44 mmol) in dioxane
(
(DMSO-D6, 300 MHz): d 10.8 (br-s, 1H–NH), 8.76 (d, 1H,
J = 4.5 Hz), 8.13 (d, 1H, J = 7.8 Hz), 8.02 (dt, 1H,
J = 7.5, 1.5 Hz), 7.57 (dd, 1H, J = 4.8, 1.0 Hz), 5.88 (s,
30 ml), a mixture of BOC anhydride (0.85 ml, 3.7 mmol)
and pyridine (0.24 ml, 2.93 mmol) was added at room
temperature. The mixture was stirred for 0.5 h, and then,
ammonium carbonate (281 mg, 2.93 mmol) was added,
and the resulting solution was stirred for 8 h. The reaction
mixture was filtered, and the filtrate was extracted with
ethyl acetate (30 ml). The organic layer was washed with
water (30 ml 9 3) followed by brine. The organic layer
was dried over Na SO and filtered, and the solvent was
1
3
2H), 3.56 (br-s, 2H–NH ) ppm. C-NMR (DMSO-D6,
2
75 MHz): d 166.5, 164.2, 150.4, 145.6, 139.3, 126.2, 123.3,
51.4 ppm; MS: m/z 220 (M ? 1).
General procedure for synthesis of the complexes
To a solution of L1 to L4 (0.487 mmol) in methanol (10 ml),
2
4
removed under reduced pressure to afford a crude product,
which was triturated with diethyl ether to furnish a color-
less solid.
CuCl
2
Á2H
2
O (0.244 mmol) was added, and the resulting solu-
tion was stirred for 2 h at 70 °C. The resulting mixture was
allowed to cool, and the solvent was evaporated slowly in the
hope of obtaining crystals. But after 7 days, the evaporation
afforded only amorphous green solids. These solids were trit-
urated with diethyl ether and used without further purification.
Ethyl
2-(5-(pyridin-2-yl)-1H-tetrazol-1-yl)acetamide
(
L1): Colorless solid; Yield: 0.25 g (50 %). M.p.
7–99 °C. Anal. Calc. for C H N O (204.19): % of C,
9
8
8 6
-
1
4
1
7.0; H, 3.9; N, 41.2. Found: % of C, 47.0; H, 3.9; N, 41.1.
[Cu(L1)
2938 (s), 2027 (s), 1636 (m), 1575 (m), 1400 (s), 1316 (s),
1294 (s), 1084 (s), 985 (s), 896 (s). Anal. calc. for C16
Cu (472): % of C, 40.7; H, 3.4; N, 35.6; Found: % of C,
40.3; H, 3.3; N, 35.5. ESI–MS: m/z 472 (M ? 1).
2
]: 189 mg, 0.4 mmol; IR (KBr, cm ): 3445 (w),
H NMR (CDCl , 300 MHz): d 8.73 (t, 1H, J = 4.2,
3
3
.3 Hz), 8.16 (d, 1H, J = 7.8 Hz), 8.05 (m, 1H), 8.01 (br.
H
16-
s, 1H), 7.59 (m, 2H), 6.14 (br. s, 1H), 6.13 (s, 2H) ppm.
3
N O
12 2
1
C-NMR (CDCl , 75 MHz): d 172.6, 153.3, 149.4, 143.6,
3
-
1
1
38.4, 126.3, 124.6, 53.1 ppm. MS: m/z 205 (M ? 1).
[Cu(L2)
(w), 2981 (s), 2067 (s), 1635 (m), 1568 (m), 1397 (s), 1252
(s), 1168 (s), 1053 (s), 789(s), C16 Cu (472): % of
2
]: 195 mg, 0.42 mmol; IR (KBr, cm ): 3448
Ethyl 2-(5-(pyridin-2-yl)-2H-tetrazol-2-yl)acetamide
L2): Colorless solid; Yield: 0.36 g, (72 %). M.p.
(
H N O
16 12 2
9
8–103 °C. Anal. Calc. for C H N O (204.19): % of C,
C, 40.7; H, 3.4; N, 35.6. Found: % of C, 40.3; H, 3.4; N,
8
8 6
4
1
7.0; H, 3.9; N, 41.2. Found: % of C, 47.0; H, 3.9; N, 41.1.
35.5. ESI–MS: m/z 472 (M ? 1).
-1
[Cu(L3) ]: 210 mg, 0.42 mmol; IR (KBr, cm ): 3413
2
H NMR (CDCl , 300 MHz): d 8.75 (t, 1H, J = 4.2,
3
3
.3 Hz), 8.15 (d, 1H, J = 7.8 Hz), 8.02 (td, 1H, J = 7.5,
(w), 3254 (w), 2033 (s), 1763 (m), 1637 (m), 1618 (m),
1455 (s), 1309 (s), 1294 (s), 1229 (s), 1110 (s), 1013 (s),
1
.5 Hz), 7.90 (br-s, 1H), 7.56 (m, 2H), 6.16 (br-s, 1H), 5.51
1
3
(
s, 2H) ppm. C-NMR (CDCl , 75 MHz): d 172.2, 164.5,
788 (s), 748 (m). Anal. calc. for C16H N14CuO (502): %
18 2
3
1
50.2, 145.5, 139.3, 126.6, 123.6, 56.7 ppm; MS: m/z 205
of C, 38.3; H, 3.6; N, 39.0. Found: % of C, 38.1; H, 3.5; N,
(
M ? 1).
39.0. ESI–MS: m/z 502 (M ? 1).
123

Transition Met Chem
-
1
[
Cu(L4) ]: 195 mg, 0.39 mmol; IR (KBr, cm ): 3453 (w),
2
2 on alkylation with ethylbromo acetate in dry DMF at
70 °C afforded regio-isomers 3 and 4 by alkylation at the
tetrazole N(1) or N(2) positions, respectively (Scheme 1).
The structures of the isomers were readily assigned by their
3
254 (w), 2042 (s), 1753 (m), 1628 (m), 1592 (m), 1438 (s), 1395
(
s), 1313 (s), 1170 (s), 1113 (s), 1013 (s), 786 (s), 728 (m). Anal.
calc. for C H N CuO (502): % of C, 38.3; H, 3.6; N, 39.0.
2
1
6
18 14
1
Found: % of C, 38.2; H, 3.5; N, 39.0. ESI–MS: m/z502 (M ? 1).
H NMR spectra. Compound 3 showed characteristic peaks
at d 5.75 (–N–CH –CO–), 4.19 (O–CH ) and 1.21(–CH ),
which confirmed the formation of an ethyl ester. Similarly,
2
2
3
Measurement of antioxidant activity
compound 4 showed characteristic peaks at d 5.5 (–N–
To a solution containing FeCl (0.1 mM, 0.5 ml), EDTA
3
CH –CO–), 4.29 (O–CH ) and 1.29 (–CH ). The ethyl
2 2 3
(
0.1 mM, 0.5 ml), ascorbic acid (0.1 mM, 0.5 ml), hydro-
esters were further confirmed by comparing the data with
reported compounds [2]. The ethyl ester derivatives 3 and 4
were hydrolyzed to acid derivatives 5 and 6, respectively,
gen peroxide (2 mM, 0.5 ml) and p-nitrosodimethyl aniline
H
0.01 mM, 0.5 ml) in phosphate buffer (p 7.4, 20 mM)
(
1
were added various concentrations of the test compounds
in distilled DMSO to produce a final volume of 3 ml. The
absorbance of the solution was then measured at 440 nm,
and the radical scavenging activity was calculated using the
following equation;
by treating with aqueous 1 N NaOH. The H NMR spectra
of 5 and 6 confirmed the absence of –OCH
upfield shifts of the N–CH peaks. The acid derivatives
CH peaks and
3
2
2
were further characterized by their mass spectra, which
showed M - 1 (negative mode) peaks at 204. The acid
derivatives 5 and 6 on treatment with ammonium carbon-
ate, BOC anhydride and pyridine in dry 1,4-dioxane
afforded the respective acetamide ligands L1 and L2. The
formation of L1 was confirmed by the observation of –CO–
p À NDA radical scavenging activity ð%Þ
½
AðsampleÞ À Aðstandardފ
¼
 100
ð1Þ
½
where A = Absorbance.
Aðsampleފ
NH signals at d 8.01 and 6.14 ppm as broad singlets;
2
similarly, L2 showed broad singlets at d 7.9 and 6.16 ppm.
The mass spectra of both L1 and L2 showed M ? H peaks
at 205.
DNA-binding experiments
The acetohydrazide ligands L3 and L4 were prepared by
reacting the ethylester derivatives 3 and 4 with hydrazine
All measurements with CT-DNA were taken in Tris–HCl
buffer 5 mM (pH 7.2), 50 mM NaCl. The UV absorbance
ratio of the DNA at 260/280 was 1.8–1.9, indicating that
the DNA was sufficiently free of protein [11]. The con-
centration of CT-DNA per nucleotide was determined from
the absorption intensity at 260 nm with the known
1
Scheme 1). The hydrazide L3 was characterized by H
NMR, which showed a characteristic –CO–NH peak as a
(
broad singlet at d 9.48 ppm. The hydrazide –NH peak
2
1
appeared as broad singlet at d 4.29 ppm. The H NMR
-
1
-1
spectrum of the hydrazide L4 showed a characteristic –
CO–NH peak at d 10.8 ppm as a broad singlet, while the
extinction coefficient of 6600 M cm . The absorption
titrations were performed by adding increasing amounts of
CT-DNA to a solution of the complex at a fixed concen-
tration contained in a quartz cell and recording the UV–Vis
spectrum after each addition. The absorption of CT-DNA
was subtracted by adding the same amount to the blank.
The data were then fitted to Eq. (2) to obtain the intrinsic
hydrazide –NH peak gave a broad singlet at d 3.56 ppm.
2
The mass spectra of both L3 and L4 showed M ? H peaks
at 220.
1
The H NMR spectra of all the compounds 3–6 and L1–
L4 all showed four signals corresponding to pyridyl pro-
1
3
tons. The C NMR chemical shift values for the tetrazole
quaternary carbon of each of these compounds were
observed at *153 ppm for the N(1)-isomers and at
d * 164 ppm for the N(2)-isomers.
binding constant, K [12].
b
À
Á
À
Á
À
Á
½
DNAŠ= e À e ¼ ½DNAŠ= e À e þ 1=K e À e
a
f
b
f
b
b
f
ð2Þ
The ligands L1–L4 were treated with CuCl Á2H O, in
2
2
where [DNA] is the molar concentration of CT-DNA, e , e
a
b
refluxing methanol using a 1:2 meta/ligand ratio to give the
corresponding complexes [Cu(L)2] (Scheme 2). The
physical properties of the complexes are given in Table 1.
The elemental analyses of all four complexes showed 1:2
and e are apparent, free and bound metal complex
f
extinction coefficients, respectively. Thus, K is the ratio of
b
the slope to the y-intercept.
(metal/ligand) compositions. All of these complexes have
Results and discussion
magnetic moment values in the range of 1.84–1.92 BM,
which is slightly higher than the spin-only values (1.73
9
Starting from the picolinonitrile 1, the literature procedure
was followed to give the tetrazole 2, whose analytical data
were comparable to the literature values [13, 14]. Tetrazole
l ) expected for a d copper(II) system [15].
eff
The ESI mass spectra of the complexes were used to
confirm their stoichiometries. The molecular ion peaks
1
23

Transition Met Chem
Table 1 Physical properties of the complexes
Table 4 EPR spectral assignments for the complexes at room
temperature
Complex
Color
Melting point (°C)
leff (BM)
Complexes
g
k
g
\
g
av
G
[Cu(L1)
[Cu(L2)
[Cu(L3)
[Cu(L4)
2
]
2
]
2
]
2
]
Green
Green
Green
Green
162–164
158–160
201–203
194–198
1.84
1.92
1.86
1.90
[
Cu(L1)
Cu(L3)
2
]
]
2.16
2.14
2.05
2.04
2.10
2.09
3.306
3.652
[
2
copper [18]. Additional peaks around 1395–1250 and
-
1
800–600 cm
are consistent with coordination of the
Table 2 Electronic spectral data for the complexes
pyridine ring to the metal. The relatively few bands
observed in the spectra of the complexes suggest that they
are symmetric in nature [19].
Complexes
MLCT (nm)
d–d (nm)
Cu(L1)
Cu(L2)
Cu(L3)
Cu(L4)
2
2
2
2
438 (8250)
430 (7400)
437 (8150)
445 (7840)
655 (145)
652 (130)
650 (140)
654 (150)
The solid-state EPR spectra of [Cu(L1) ] and [Cu(L3) ]
2
2
were recorded in the X-band region at 25 °C, and the data
are summarized in Table 4. [Cu(L1) ] and [Cu(L3) ]
exhibit g values of 2.16 and 2.18 and g\ values of 2.05
2
2
k
Extinction coefficients are given in the brackets
and 2.06, respectively. In both cases, it is clear that
g [ g [ 2.00, suggesting that the unpaired electron lies
k
predominantly in the d orbital, which is the characteristic
\
?
M ) of [Cu(L1) ] and [Cu(L2) ] were observed at m/
22
x-y
(
2
2
z = 472 and those for [Cu(L3) ] and [Cu(L4) ] at 502,
2
of square planar geometry in copper(II) complexes.
The geometric parameter G, which is a measure of the
exchange interaction between the copper centers in the
polycrystalline compound, was calculated using Eq. 3.
2
consistent with the proposed formulations.
The electronic spectra of these complexes recorded in
DMF are summarized in Table 2. The spectra showed bands
in the region of 430–450 nm with high molar extinction
À
Á
?
G ¼ ðgjj À 2:0023Þ= g À 2:0023
ð3Þ
-
1
-1
coefficients in the range of 8300–7300 M cm , which
are assigned to metal-to-ligand charge transfer (MLCT)
transitions (n - p*). The single broadband observed in the
region 655–650 nm is assigned to d–d transitions; these
bands have low intensities, with molar extinction coeffi-
According to Hathaway and Tomlinson [20], if G [ 4.0,
the exchange interaction is negligible because the local
tetragonal axes are aligned parallel or only slightly mis-
aligned. If G \ 4.0, exchange is considerable, and the local
tetragonal axes are misaligned. The G values of [Cu(L1)2]
-
1
-1
cients between 130 and 95 M cm . They are assigned to
2
2
E ? T electronic transitions, suggesting a symmetrical
and [Cu(L3) ] were obtained as 3.306 and 3.652, respec-
2
g
2g
square planar geometry for the copper(II) complexes [16,
7].
The IR spectral data for the complexes are summarized
in Table 3. The free ligands L1 and L2 both show a
tively, suggesting that there is no exchange interaction in
these copper(II) complexes.
1
DNA-binding studies
-
1
broadband in range 3550–3100 cm corresponding to –
NH of the amide/hydrazine group, plus amide peaks at
2
The binding interactions of the complexes with CT-DNA
were investigated by UV–Vis spectroscopy. The com-
plexes were stable in Tris buffer solution [21] (refer
Fig. 2S, supplementary information). The absorption
spectra of the copper complexes were compared with and
without CT-DNA at 400 nm (Fig. 1). The obtained UV–
Vis data and the binding constants of these complexes are
given in Table 5. Upon addition of increasing amounts of
-
1
681 and 1685 cm , respectively. The equivalent at 1622
1
-
1
and 1642 cm for the hydrazine ligands L3 and L4 is
observed at lower frequencies. All of these peaks are
shifted to lower frequencies in the copper complexes. The
-
1
peaks at 1626–1534 cm assigned to the pyridyl tetrazole
group are also shifted to lower frequency in the complexes,
indicating coordination of the pyridyl tetrazole rings with
Table 3 Selective I.R. bands
cm , of Cu(II) complexes with
tentative assignments
-1
Vibration
L1
[Cu(L1)
2
]
L2
[Cu(L2)
2
]
L3
[Cu(L3)
2
]
L4
2
[Cu(L4) ]
Amide/
1681
–
1636
–
1685
–
1635
–
–
–
–
–
Hydrazine
1622
1534
1471
1433
1618
1455
1309
1294
1642
1626
1508
1443
1628
1592
1438
1395
C=N (tetrazole)
C=C (tetrazole)
Py/M-Py
1617
1498
1392
1575
1400
1316
1606
1486
1334
1568
1397
1252
123

Transition Met Chem
It is evident from Table 5 that all four complexes bind to
DNA with high affinities such that the estimated binding
4
-1
constants are in the range 5.4 - 7.2 9 10 M . The Kb
values of these complexes are comparable with the repor-
ted values for redox active Cu(II) pyridine-based tetra-
zolo[1,5-a]pyrimidine complexes [22]. The intrinsic
binding constant for cis-platin, which shows a hyper-
chromic shift in the presence of CT-DNA, is reported as
4
-1
3
.20 10 M [23].
The interaction of the present complexes with DNA is
most likely via p-stacking interactions of the planar pyridyl
tetrazole rings. Binding of these complexes to CT-DNA by
external contact (electrostatic or groove binding) brought
about a bathochromic shift with hyperchromism [24].
Antioxidant activity
2
Fig. 1 UV–Vis absorption spectra of the copper complex [Cu(L3) ]
with and without CT-DNA in Tris buffer solution
The in vitro antioxidant activities of the pyridine tetrazole
derivatives and their complexes were evaluated using a
reported p-NDA radical scavenging method [25]. The free
radical scavenging activities of the compounds were eval-
uated through their ability to bleach p-NDA using ascorbic
acid as a reference standard. Potencies for the antioxidant
activities of the test compounds compared to ascorbic acid
as a reference are given in Table 6. In general, all the
present compounds were moderately more potent than the
reference. The complex [Cu(L4)2] showed highest
antioxidant activity.
Table 5 Interactions of the complexes with CT-DNA from UV–Vis
spectroscopy
-
1
Complex
kmax/nm
Dk/nm
H%
b
K /M
Free
Bound
4
4
4
4
[Cu(L1)
[Cu(L2)
[Cu(L3)
[Cu(L4)
2
]
2
]
2
]
2
]
434.0
438.0
440.0
439.0
433.5
437.4
437.5
437.5
0.5
0.6
1.8
1.5
7.5
4.5
10.5
8.0
6.2 9 10
5.4 9 10
7.2 9 10
6.4 9 10
H% = Hyperchromism; Dk = Bathochromic shift
Conclusions
CT-DNA, the UV–Vis absorption spectra of all four
complexes showed increase in absorbance, together with a
bathochromic shift (*0.5 nm) with hyperchromism with
respect to the control (0 ll DNA). The changes in absor-
bance values with increasing amounts of CT-DNA were
Pyridyl–tetrazole ligands with acetoamide and acetohy-
drazide pendant arms and their copper complexes were
synthesized and characterized. Physicochemical and spec-
tral studies suggest square planar copper(II) geometries.
All four complexes are avid binders of CT-DNA and also
have antioxidant properties. These ligands provide a useful
used to evaluate the intrinsic binding constant K (Table 5).
b
Table 6 p-NDA radical
Compound
% radical scavenging method concentrations mg/ml
scavenging activities of pyridine
tetrazole ligands and their
complexes
0.5 mg/ml
1 mg/ml
1.5 mg/ml
2 mg/ml
2.5 mg/ml
IC-50 mg/ml
L1
L2
L3
L4
28.47
25.36
23.8
40
32.43
30.45
37.38
40.49
40.06
38.7
39.28
32.67
37.84
40.34
44.16
39.26
36.35
10.16
80.45
39.84
33.87
40.04
44.73
44.23
42.13
39.05
3.3
40.72
35.91
40.66
42.91
44.24
45.29
39.31
42.29
85.46
3.7
5.24
3.43
5.76
4.3
[Cu(L1)
[Cu(L2)
[Cu(L3)
[Cu(L4)
2
2
2
2
]
]
]
]
39.6
37.8
36.26
59.54
32.3
4.04
8.38
2.815
0.64
36.04
54.64
56.78
Ascorbic acid
82.34
1
23

Transition Met Chem
template for further derivatization with a view to the
development of more potent and selective antioxidants and
DNA-binding agents.
10. Mustafa S, Rao BU, Surendrababu MS, Raju KK, Rao GN (2015)
Chem Biodivers 12:1516
1
1. Reichmann ME, Rice SA, Thomas CA, Doty P (1954) J Am
Chem Soc 76:3047
2. Wolfe A, Shimer GH, Meehan T (1987) Biochemistry 26:6392
1
Acknowledgments SM, MSB and KSR are thankful to the financial
assistances from DST with project No.SB/FT/CS-034/2012 (SERB,
New Delhi, India) and UGC with project F.No. 42-306/2013 (SR)
13. Bond AD, Fleming A, Gaire J, Kelleher F, McGinley J, McKee
V, Sheridan U (2012) Polyhedron 33:289
14. Fleming A, Kelleher F, Mahon MF, McGinley J, Prajapati V
(2005) Tetrahedron 61:7002
(
New Delhi, India).
1
1
1
5. Geary WJ (1971) Coord Chem Rev 7:81
6. Butler RN, Fleming AFM (1997) J Heterocycl Chem 34:691
7. Hathaway BJ (1987) In: Wilkinson G, Gillard RD, Mecleverty JA
(eds) Comprehensive coordination chemistry, vol 5. Pergamon
press, Oxford, p 583
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