Antioxidant potential and antimicrobial screening of some novel imidazole derivatives: Greenway efficient one pot synthesis

Article abstract of DOI:10.1007/s00044-011-9702-5

A series of substituted imidazoles have been synthesized under solvent-free condition by grinding 1,2-diketone, aromatic aldehyde, and ammonium acetate in the presence of molecular iodine as the catalyst. The short reaction time and easy workup make this

Full text of DOI:10.1007/s00044-011-9702-5

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2012) 21:1850–1860
DOI 10.1007/s00044-011-9702-5
ORIGINAL RESEARCH
Antioxidant potential and antimicrobial screening of some novel
imidazole derivatives: greenway efficient one pot synthesis
•
Jayaraman Jayabharathi Venugopal Thanikachalam
•
•
•
Nagarajan Rajendraprasath Kanagarathinam Saravanan
Marimuthu Venkatesh Perumal
Received: 3 February 2011 / Accepted: 9 June 2011 / Published online: 25 June 2011
ꢀ Springer Science+Business Media, LLC 2011
Abstract A series of substituted imidazoles have been
synthesized under solvent-free condition by grinding
1,2-diketone, aromatic aldehyde, and ammonium acetate in
the presence of molecular iodine as the catalyst. The short
reaction time and easy workup make this protocol practi-
cally and economically attractive and are characterized by
NMR spectra, X-ray, mass, and CHN analysis. Their
antioxidant potential were evaluated using different in vitro
antioxidant models namely, DPPH (1,1-diphenyl-2-picryl-
hydrazyl) radical, superoxide anion, and hydroxyl radical
scavenging activities. Their antibacterial screening against
Staphylococcus aureus, Escherichia coli, and Klbesiella
pneumoniae and antifungal activity against Aspergillus
niger, Aspergillus flavus, and Candida-6 were also evalu-
ated. Among all, dimethoxyphenyl substituent at N3 of the
imidazole derivatives exhibited the highest hydroxy and
superoxide anion radical scavenging activities, whereas
dimethoxyphenyl substituent at N3 and fluorophenyl at C2
of the imidazole derivatives exhibited the highest DPPH
radical scavenging activity.
Introduction
Organic solvents are high on the list of damaging chemi-
cals. In recent years, solid-state organic reactions have
caused great interest. Designing of ‘‘green’’ experimental
protocol is an enormous challenge to chemists to improve
the quality of the environment for present and future gen-
erations. Compounds with an imidazole ring system have
many pharmacological properties (Lambardino and Wise-
man, 1974; Maier et al., 1989). Though there are several
methods reported (Lantos et al., 1993; Zhang et al., 1996)
in the literature for the synthesis of imidazoles, they suffer
from one or more disadvantages such as harsh reaction
conditions, poor yields, prolonged time period, use of
hazardous, and often expensive acid catalysts. Recently,
molecular iodine received considerable attention as an
inexpensive, nontoxic, readily available catalyst for various
organic transformations, affording the corresponding
products in excellent yields with high selectivity. Owing to
numerous advantages associated with this eco-friendly
element, iodine has been explored as a powerful catalyst
for various organic transformations (Jianwei et al., 2004).
During the course of our studies toward the development of
new route to the synthesis of biologically active hetero-
cycles, we wish to report a simple and an efficient method
for the synthesis of substituted imidazoles.
Keywords Antioxidant activity ꢀ Antimicrobial
screening ꢀ Free radical scavenging activity ꢀ NMR ꢀ
X-ray
Free radicals, the partially reduced metabolites of
oxygen and nitrogen, are highly toxic and reactive. Free
radicals are linked with the majority of diseases like aging,
atherosclerosis, cancer, diabetes, liver cirrhosis, cardio-
vascular disorders, etc. (Gutteridgde 1995; Aruoma 1998).
The most common reactive oxygen species are superoxide
anion (O2•–), hydrogen peroxide (H2O2), peroxyl radical
(ROO•), and highly reactive hydroxyl radical (OH•). The
nitrogen-derived free radicals are nitric oxide (NO) and
J. Jayabharathi (&) ꢀ V. Thanikachalam ꢀ K. Saravanan ꢀ
M. V. Perumal
Department of Chemistry, Annamalai University,
Annamalainagar, Chidambaram 608002, Tamilnadu, India
e-mail: jtchalam2005@yahoo.co.in
N. Rajendraprasath
Department of Biochemistry, Annamalai University,
Annamalainagar, Chidambaram 608 002, Tamilnadu, India
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Med Chem Res (2012) 21:1850–1860
1851
peroxy nitrite anion (ONOO•). Oxidation process is one of
the most important routes for producing free radicals in
food, drugs and living systems. Antioxidants are the sub-
stances that, when present in low concentration, signifi-
cantly delay or reduce the oxidation of the substrate
(Halliwell, 2000). Antioxidants protect the body from
damaging oxidation reactions by reacting with free radi-
cals and other reactive oxygen species within the body and
hindering the process of oxidation. So diseases linked with
free radicals can be prevented by antioxidant therapy
which gained an immense importance.
Experimental
Materials and methods
General procedure for the synthesis of 2-aryl imidazole
derivatives
A mixture of aldehyde (1 mmol), 1,2-diketone (1 mmol),
ammonium acetate (2.5 mmol), and iodine (15 mol%)
were grined in a mortar at room temperature for 2 h. The
completion of the reaction was monitored by TLC and the
reaction mixture was treated with aqueous Na2S2O3, the
formed crude was purified by column chromatography
(Jayabharathi et al., 2009, 2010a, b; Saravanan et al., 2010)
(n-hexane:ethyl acetate (9:1) (Scheme 1).
Current research is now directed toward finding natu-
rally occurring antioxidants particularly of plant origin.
Currently available synthetic antioxidants like butylated
hydroxy anisole (BHA), butylated hydroxy toluene
(BHT), tertiary butylated hydroquinone and gallic acid
esters have been suspected to cause negative health
effects. BHA and BHT are suspected of being responsible
for liver toxicity and carcinogenesis (Branen, 1975;
Wichi, 1986; Grice, 1988). Hence, strong restrictions have
been placed on their application and there is a trend to
substitute them with naturally occurring antioxidants.
Traditionally used natural antioxidants from tea, wine,
fruits, vegetables, spices and medicine (e.g., rosemary and
sage) are already exploited commercially either as anti-
oxidant additives or as nutritional supplements (Schuler,
1990). Also many other plant species have been investi-
gated in search of novel antioxidants (Mantle et al., 2000;
Koleva et al., 2002; (Oke and Hamburger, 2002; Parejo
et al., 2003), but generally there is still a demand to find
more information concerning the antioxidant potential of
plant species. It has been mentioned that the antioxidant
activity of plants might be due to their phenolic com-
pounds (Cook and Samman, 1996). Flavonoids are a
group of polyphenolic compounds with known properties
which include free radical scavenging, inhibition of
hydrolytic and oxidative enzymes and anti-inflammatory
actions (Frautchy et al., 2001; Wang et al., 2006; Clavin
et al., 2007). The use of traditional medicine is wide-
spread and plants still present a large source of natural
antioxidants that might serve as leads for the development
of novel drugs. Many plant species have been investigated
in search for novel antioxidants in the recent time but still
the demand to find more novel antioxidant persists. When
compared with natural phytochemicals, it is possible to
synthesize more potent antioxidants through chemical
synthesis. Therefore, the objective of present study was a
development of new route to the synthesis of heterocycles
with antioxidant potential. These substituted imidazoles
were subjected to evaluate in vitro antioxidant activity
through determining different free radical scavenging
assay and their antimicrobial screening were also carried
out.
4,5-Dimethyl-1-(30,50-dimethoxyphenyl)-2-
(p-fluorophenyl)-1H-imidazole (dmdmpfpi)
Yield: 60%. 1H NMR (400 MHz, CDCl3): d 2.28 (s, 3H), 1.99
(s, 3H), 3.85 (s, 3H), 6.85–7.35 (aromatic protons). 13C NMR
(100 MHz, CDCl3): d 9.50, 12.69, 55.48, 114.71, 114.93,
115.14, 128.90, 129.80, 130.42, 133.21, 144.36, 159.45. Anal.
calcd. for C18H17N2OF: C, 72.95; H, 5.78; N, 9.45. Found: C,
72.24; H, 5.36; N, 8.98. MS: m/z 296.5, calcd. 296.35.
4,5-Dimethyl-1-(40-methoxyphenyl)-2-phenyl-1H-imidazole
(dmmppi)
1
Yield: 60%. H NMR (400 MHz, CDCl3): d 2.29 (s, 3H),
2.02 (s, 3H), 3.85 (s, 3H), 6.91–7.10 (aromatic protons).
13C NMR (100 MHz, CDCl3): d 9.39, 12.59, 55.32, 114.50,
125.51, 127.41, 127.87, 128.78, 130.48, 130.74, 133.12,
145.08, 159.23. Anal. calcd. for C18H18N2O: C, 77.67; H,
6.52; N, 10.06. Found: C, 77.14; H, 6.32; N, 9.87. MS: m/z
278.2, calcd. 278.36.
4,5-Dimethyl-1-(30,50-dimethoxyphenyl)-2-
(p-fluorophenyl)-1H-imidazole (dmdmpfpi)
1
Yield: 65%. H NMR (400 MHz, CDCl3): d 2.28 (s, 3H),
2.04 (s, 3H), 3.74 (s, 6H), 6.30–7.40 (aromatic protons).
13C NMR (100 MHz, CDCl3): d 9.40, 12.58, 55.44, 100.32,
106.25, 114.85, 115.06, 125.23, 127.00, 129.55, 133.38,
139.27, 143.98, 161.18. Anal. calcd. for C19H19N2O2F: C,
69.92; H, 5.87; N, 8.58. Found: C, 69.12; H, 5.37; N, 8.24.
MS: m/z 326.3, calcd. 326.37.
4,5-Dimethyl-1-(30,50-dimethoxyphenyl)-2-phenyl-1H-
imidazole (dmdmppi)
1
Yield: 65%. H NMR (400 MHz, CDCl3): d 2.29 (s, 3H),
2.05 (s, 3H), 3.73 (s, 6H), 6.32–7.43 (aromatic protons).
123

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Med Chem Res (2012) 21:1850–1860
Scheme 1 General schematic
representation of synthesis of
imidazole derivatives
13C NMR (100 MHz, CDCl3): d 9.36, 12.58, 55.32, 100.21,
106.20, 125.13, 127.45, 127.65, 127.83, 130.68, 133.34,
139.38, 144.79, 161.01. Anal. calcd. for C19H20N2O2: C,
73.93; H, 6.54; N, 9.08. Found: C, 73.23; H, 6.18; N, 8.79.
MS: m/z 308.3, calcd. 308.38.
fungi. The minimum inhibitory concentrations (MICs)
were recorded by visual observations after 24 h (for bac-
teria) and 72–96 h (for fungi) of incubation; ciprofloxacin
and Amphotericin B were used as standards.
In vitro antioxidant activity
In Vitro Antibacterial and Antifungal Activity
DPPH radical scavenging activity The stable free radical
DPPH method is an easy, rapid and sensitive way to survey
the antioxidant activity of a specific compound. The DPPH
radical scavenging activity of imidazole derivatives were
determined by the method of Baricevic et al., 2001.
The in vitro activities of imidazole derivatives were tested
in Sabourauds dextrose broth (Jayabharathi et al., 2007,
2008, 2010a, b) (SDB; Hi-media, Mumbai) for bacteria by
the two fold serial dilution method. The compounds were
dissolved in dimethylsulfoxide (DMSO) to obtain 1 mg/ml
stock solutions. Seeded broth (broth containing microbial
spores) was prepared in NB from 24-h-old bacterial cul-
tures were suspended in SDB. The colony forming units
(cfu) of the seeded broth were determined by plating
technique and adjusted in the range of 104–105 cfu/ml. The
final inoculum size was 105 cfu/ml for antibacterial assay
and 1.1–1.5 9 102 cfu/ml for antifungal assay, and testing
was performed at pH 7.4 0.2. Exactly 0.2 ml of the
solution of each test compound was added to 1.8 ml of
seeded broth to form the first dilution. 1 ml of this was
diluted with a further 1 ml of the seeded broth to give the
second dilution and so on till six such dilutions were
obtained. A set of assay tubes containing only seeded broth
was kept as control and similarly solvent controls were also
run simultaneously. The tubes were incubated in BOD
incubators at 37 1 ꢁC for bacteria and 28 1 ꢁC for
Superoxide radical scavenging activity Superoxide anion
radical scavenging activity of synthetic imidazole deriva-
tives were determined by the method of Nishimiki (Nish-
imiki et al., 1972). The assay was based on the oxidation of
Nicotinamide adenine dinucleotide (NADH) by phenazine
methosulphate (PMS) to liberate PMSred
.
Hydroxyl radical scavenging activity Hydroxyl radical
scavenging activity of synthetic imidazole derivatives were
determined by the reported method (Elizabeth and Rao,
1990).
Reducing power assay The Fe3? reducing power
of imidazole was determined by the method of Oyaizu
(1986).
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Med Chem Res (2012) 21:1850–1860
Results and discussion
1853
˚
from 1.47 to 1.36 A on excitation from S0 to S1 state
(Table 1). Since these imidazole derivatives are used as
fluorophore, they were tested for antioxidant and antimi-
crobial activities.
Photophysical properties of dmdmpfpi
Crystal growth of dmdmpfpi was carried out by Slow
Evaporation Solution Growth Technique (SEST) and the
crystalline imidazole derivative dmdmpfpi was a mono-
clinic crystal (Fig. 1) crystallizes in the space group p21/c,
There are different models available for evaluation of
antioxidant activities. The chemical complexity of com-
pounds could lead to scattered results, depending on the
test employed. Therefore, an approach with multiple assays
for evaluating the antioxidant potential of imidazole
derivatives would be more informative and even necessary.
In this study, different free radical scavenging activities
were measured and all the results were compared with
standard antioxidant.
˚
and cell has dimensions of a = 8.5132 (1) A, b = 9.5128
˚
˚
(2) A, c = 19.2610 (3) A, b = 96.798 (2). ORTEP dia-
gram (Gayathri et al., 2010) of dmdmpfpi shows that the
imidazole ring makes dihedral angles of 76.46 (7) and
40.68 (7)8 with the methoxyphenyl and fluorophenyl rings,
respectively. The dihedral angle between the two benzene
rings is 71.25 (6)8. The band appeared at 292.8 nm (kabs
)
In vitro antioxidant activity
and 364.4 nm (kem) is due to the p–p* transition. Since in
dmdmpfpi, n–p* and the p–p* transitions are in close
proximity, the low intensity n–p* transition is often com-
pletely overlaid by the intensive p–p* transition. The
UV–visible spectrum (Fig. 2) of dmdmpfpi was recorded
in different solvents such as non-hydroxy solvents and
hydroxy solvents, and it was observed that the absorption
maximum was red shifted in the polar aprotic solvents,
may be due to the presence of increased resonance inter-
action of the p-cloud of the phenyl ring attached to the
carbon of the imidazole ring with the lone pair of nitrogen
atom (N3) of the imidazole ring (Ren et al., 2003) and the
blue shift in polar protic solvent is due to hydrogen
bonding interactions with the lone pair on nitrogen atom and
thus inhibiting from resonance interaction with p-cloud of
phenyl ring. The resonance interaction increases if the lone
pair and the p-cloud are parallel to each other and to
understand the coplanarity, optimization of dmdmpfpi DFT
at B3LYP/6-31G(d,p) have been performed using Gaussian-
03 (Table 1). The observed dihedral angle from both XRD
[N3–C2–C21–C22(-137.68); N3–C2–C21–C26 (40.2) and
C5–N1–C11–C12 (84.28); C5–N1–C11–C16 (-94.78)] and
DFT [N3–C2–C21–C22 (-139.18); N3–C2–C21–C26
(39.0) and C5–N1–C11–C12 (71.38); C5–N1–C11–C16
(-84.18)] results strongly evidenced the existence of reso-
nance interaction of the p-cloud of the phenyl ring attached
to the carbon of the imidazole ring with the lone pair of
nitrogen (N3) of the imidazole ring and also it was found that
there is poor coplanarity between the p-methoxyphenyl ring
attached to the nitrogen with the imidazole nucleus (Fig. 3).
The observed fluorescence spectrum (Fig. 4) is broad and
more sensitive to changing the polarity of the solvents. This
is in argument with the fact that greater charge transfer takes
place from aromatic ring to imidazole nucleus in S1 state
than S0 state which is evidenced by the decrease in the
dihedral angle of N3–C2–C21–C22 and N3–C2–C21–C26
from 139.18 to 127.28 and 38.98 to 10.08, respectively.
Moreover, the reduction in the bond distances of C2–C21
DPPH radical scavenging activity
Compounds, at various concentrations ranging from 10 to
50 lM, were mixed in 1 mL of freshly prepared 0.5 mM
DPPH ethanolic solution and 2 mL of 0.1 M acetate buffer
at pH 5.5. The resulting solutions were then incubated at
37 ꢁC for 30 min and measured at 517 nm in a Shimadzu
UV-1601 spectrophotometer. DPPH• scavenging activities
of the imidazoles were calculated from the decrease in
absorbance from 517 nm in comparison with the negative
control (Fig. 5). IC50 value is the concentration of the
compound required to inhibit 50% of DPPH• production.
% of DPPHꢁscavenging ¼ ½A0 ꢂ A1= A0ꢃ ꢄ 100
Fig. 1 ORTEP diagram for dmmpfpi
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Med Chem Res (2012) 21:1850–1860
Fig. 3 Energy minimized modeling of dmmpfpi
Fig. 2 UV–Vis Spectrum of dmdmppi in various solvents
Table 1 Calculated parameters of dmdmpfpi in the ground and
excited states
Characteristics
dmdmpfpi
S0 state
E (k cal/mole)
lg (D)
0.00
4.17
Excited state DFT
E (k cal/mol)
ll (D)
1.00
10.02
Dihedral angle (ꢁ)
Ground state
(N3–C2–C21–C22)
(N3–C2–C21–C26)
Excited state
139.1
38.9
(N3–C2–C21–C22)
(N3-C2-C21–C26)
-127.2
10.0
˚
Bond length (A)
Fig. 4 Fluorescence spectrum of dmdmppi in various solvents
C2–C21 (Ground state)
C2–C21 (Excited state)
1.47
1.36
Superoxide anion scavenging assay
In the PMS/NADH-NBT system, superoxide anion is
generated using a non-enzymatic reaction of phenazine
methosulphate in the presence of NADH and molecular
oxygen (Robak and Gryglewski, 1998). It is well known
that superoxide anions damage biomacromolecules directly
or indirectly by forming H2O2, OH•, peroxylnitrite, or
where A0 was the absorbance of the control and A1 was the
absorbance in the presence of the sample of imidazole
derivatives. All the tested compounds showed DPPH rad-
ical quenching activity in a concentration dependent
manner, dmmpfpi showed maximum activity (IC50 =
11.11 1.74 lg/mL L).
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Med Chem Res (2012) 21:1850–1860
1855
Fig. 6 Scavenging potential of the imidazole derivatives at different
concentrations (lg/mL) on superoxide radicals generated by the PMS/
NADH system
Fig. 5 Radical scavenging potential of the imidazole derivatives by
DPPH method at different concentrations (lg/mL)
singlet oxygen during pathophysiologic events such as
ischemic-reperfusion injury. PMSred (phenazine methosul-
phate) convert oxidized nitro blue tetrazolium (NBToxi) to
the reduced form (NBTred), which formed a violet-colored
complex. The color formation indicates the generation of
superoxide anion, which was measured spectrophotomet-
rically at 560 nm. A decrease in the formation of color
after the addition of the antioxidant was a measure of its
superoxide-scavenging activity.
ascorbic acid. The generation of OH• is detected by its
ability to degrade deoxyribose to form products, which on
heating with thiobarbituric acid (TBA) form a pink-colored
chromogen. The addition of imidazole compound with
deoxyribose for OH• diminishes the color formation. The
incubation mixture in a total volume of 1 mL contained
0.1 mL of buffer, varying volumes of imdazoles (10, 20,
40, 60, 80, and 100 lM), 0.2 mL of 500 lM ferric chlo-
ride, 0.1 mL of 1 mM ascorbic acid, 0.1 L of 1 M EDTA,
0.1 mL of 10 mM H2O2, and 0.2 mL of 2-deoxyribose.
The contents were mixed thoroughly and incubated at room
temperature for 60 min. Then, 1 mL of 1% TBA (1 g in
100 mL of 0.05 N NaOH) and 1 mL of 28% trichloroacetic
acid (TCA) were added. All the tubes were kept in a
boiling water bath for 30 min. The absorbance of the
supernatant was observed at 535 nm with reagent blank
containing water in place of compounds. Decreased
absorbance of the reaction mixture indicates increased
hydroxyl radical scavenging activity. The percentage
scavenging was calculated as shown below:
The superoxide radical scavenging activities of active
imidazoles were evaluated based on their ability to quench
the superoxide radical generated from the PMS/NADH
reaction. 1 mL of NBT (100 lmol of NBT in 100 mM
phosphate buffer, pH 7.4), 1 mL of NADH (468 lmol in
100 mM phosphate buffer, pH 7.4) solution and varying
volume of imidazoles (10, 20, 40, 60, 80, and 100 lM)
were mixed well. The reaction was started by the addition
of 100 lL of PMS (60 lmol/100 mM phosphate buffer,
pH 7.4). The reaction mixture was incubated at 30 ꢁC for
15 min. The absorbance was measured at 560 nm in a
spectrophotometer. Incubation without the compounds was
used as blank. Decreased absorbance of the reaction mix-
ture indicates increased superoxide anion scavenging
activity (Fig. 6). The percentage scavenging was calculated
as shown below:
% of scavenging ½OHꢁꢃ ¼ ½A0 ꢂ A1= A0ꢃ ꢄ 100
where A0 was the absorbance of the control and A1 was the
absorbance in the presence of imidazole derivatives.
All the imidazoles suppressed hydroxyl radical mediated
deoxyribose degradation in a concentration-dependent
manner (Fig. 7). The hydroxyl radical is a highly potent
oxidant that reacts with almost all bio-molecules found in
living cells when it reacts with polyunsaturated fatty acid
moieties of cell membrane phospholipids, lipid hydroper-
oxides is produced (Valentao et al., 2002). Lipid hydro-
peroxide can be decomposed alkoxyl and peroxyl radicals
and numerous carbonyl products such as malondialdehyde
(MDA). The carbonyl products are responsible for DNA
damage, generation of cancer and aging-related diseases
(Okhawa et al., 1979). All the tested compounds showed
Â
Ã
% of scavenging Oꢁ2ꢂ ¼½A0 ꢂ A1= A0ꢃ ꢄ 100
where A0 was the absorbance of the control and A1 was the
absorbance in the presence of the sample of imidazole
derivatives. All the tested compounds showed superoxide
anion quenching activity in a concentration dependent
manner, dmmppi showed maximum activity (IC50 =
10.52 5.26 lg/mL).
Hydroxyl radical scavenging
In this assay, hydroxyl radical was produced by reduction
of H2O2 by the transition metal (iron) in the presence of
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Med Chem Res (2012) 21:1850–1860
superoxide anion quenching activity in a concentration
dependent manner, dmdmppi showed maximum activity
(IC50 = 34.48 3.44 lg/mL L).
(1%, w/v), followed by incubating at 50 ꢁC for 20 min.
The reaction was stopped by adding 2.5 mL of trichloro-
acetic acid (TCA) solution (10%) and then centrifuged at
8009g for 10 min. 2.5 mL of the supernatant was mixed
with 2.5 mL of distilled water and 0.5 mL of ferric chlo-
ride solution (0.1%, w/v) and the absorbance was measured
at 700 nm. Butylated hydroxyl toluene was used as refer-
ence standard. Higher absorbance of the reaction mixture
indicated greater reducing power.
Reducing power assay
The imidazole (2.5 mL) at various concentrations was
mixed with 2.5 mL of phosphate buffer (0.2 M, pH 6.6)
and 2.5 mL of potassium ferricyanide [K3Fe(CN)6]
The reducing power assay serves as a significant indi-
cator of potential antioxidant activity. Although, different
mechanism was proposed for the antioxidant activity such
Table 3 In vitro antibacterial activity of imidazole derivatives
Compound
Minimum inhibitory concentration (MIC) in lg/mL
S. aureus
E. Coli
K. pneumoniae
dmdmpfpi
dmmppi
50
50
50
25
50
25
25
50
50
100
100
50
dmmpfpi
dmdmppi
Fig. 7 Hydroxy radical scavenging potential of the imidazole
derivatives at different concentrations (lg/mL) on deoxyribose
degradation method
100
100
Ciprofloxacin 25
Table 2 IC50 values (lg/ml) of
Concentrations (lm) Ascorbic acid dmdmpfpi
dmmppi
dmmpfpi
dmdmppi
imidazole derivatives (1–4) and
standard antioxidants by
different free radical scavenging
methods
Hydroxyl radical method
10
24.26 2.14
41.29 3.14
48.57 3.7
35.63 1.99 35.63 3.98 39.08 1.99
40.22 1.99 40.22 1.99 48.27 1.99
47.12 1.99 43.67 1.99 43.67 1.99
57.47 5.26 48.27 3.44 48.27 3.44
55.17 3.44 50.57 5.26 50.57 5.26
54.02 1.99 52.87 1.99 52.87 1.99
34.48 3.44
39.08 1.99
47.12 1.99
54.02 3.98
55.17 3.44
58.62 3.44
4.14 0.14
20
40
60
65.32
5
80
70.4 4.9
78.46 5.97
2.72 0.12
100
IC50 value
4.18 0.16
4.50 0.15
3.84 0.0.11
Superoxide anion method
10
32.56 2.8
21.90 1.64 10.52 5.26 38.09 4.36
24.76 1.64 28.07 8.03 39.04 1.64
55.23 3.29 63.15 5.26 39.04 4.36
60.0 2.85 64.91 3.03 41.90 1.64
58.09 1.64 68.42 5.26 45.71 4.94
55.23 4.36 59.64 3.03 40.95 1.64
13.33 6.59
32.38 3.29
49.52 1.64
53.33 3.29
59.04 5.94
60.95 1.64
4.16 0.16
20
48.78 3.71
55.45 4.22
62.11 4.75
66.52 2.45
73.45 5.56
2.80 .22
40
60
80
100
IC50 value
4.16 0.14
3.74 0.12
5.20 .18
DPPH radical method
10
26.48 2.08
34.47 2.62
48.77 3.71
66.5 5.06
75.39 5.74
3.1 .24
12.12 3.03 14.14 3.49 11.11 1.74
19.19 1.74 28.28 1.74 20.20 1.74
42.42 3.03 59.59 3.49 46.46 3.49
57.57 3.03 56.56 1.74 64.64 1.74
56.56 1.74 55.55 1.74 68.68 1.74
17.17 1.74
20.20 1.74
52.52 3.49
57.57 3.03
55.55 1.74
3.80 0.12
20
30
40
50
IC50 value
3.98 0.18
3.64 0.12
3.46 0.16
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Med Chem Res (2012) 21:1850–1860
1857
as prevention of chain initiation, binding of transition-
metal ion catalysts, decomposition of peroxides, prevention
of continued hydrogen abstraction, reductive capacity and
radical scavenging (Gordon 1990). The substituted imida-
zoles showed concentration-dependant reductive effects.
The reducing properties are generally associated with the
presence of different reductants (Duh 1998). The moderate
reducing property was observed for all imidazole deriva-
tives. The antioxidant action of reluctant is based on the
breaking of the free radical chain by donating a hydrogen
atom. Reductones also react with certain precursors of
peroxide, thus preventing peroxide formation.
concentration dependent and the percentage of inhibition
was linearly increased with increasing concentration and
the superoxide anion radical scavenging ability was found
to be in the order dmdmppi [ dmmppi [ dmdmpf-
pi [ dmmpfpi. DPPH• with suitable reducing agents and
electron becomes paired off and the solution loses color
stoichiometrically with the number of electrons taken up.
Such reactivity has been employed to test the ability of
imidazole derivatives to act as free radical scavengers. The
scavenging ability of compounds on DPPH• were linearly
increased with increasing concentration and DPPH•-scav-
enging ability was found to be in the order dmmpfpi [
dmdmpfpi [ dmmppi [ dmdmppi. The calculated IC50
(inhibitory concentration) values of imidazole derivatives
are presented in Table 2.
The hydroxyl radical scavenging ability of imidazole
derivatives exhibits inhibition of OH• formation and
percentage of inhibition were linearly increased with
increasing concentration and the OH•-scavenging ability
was found to be in the order dmdmppi [ dmmp-
pi [ dmdmpfpi [ dmmpfpi. The superoxide anion scav-
enging ability of imidazole derivatives were found to be
From the observed results, it was concluded that the
imidazole derivatives showed potent-scavenging activities
and it is evident that dimethoxyphenyl at C3 of the imid-
azole ring (dmdmppi) has maximum OH• and superoxide
Plate 1 Antibacterial activity
of the imidazole derivatives
123

1858
Med Chem Res (2012) 21:1850–1860
Antimicrobial studies
anion radical scavenging activities when compared with
other imidazole derivatives. The low IC50 value of
dmdmppi may be due to the electron donating (?I effect)
ability exerted by the two methoxy substituents. The p-fluoro
phenyl group at C2 of the imidazole ring (dmmpfpi) has
maximum DPPH• radical scavenging activity when com-
pared with other imidazole derivatives and the low IC50
value of dmmpfpi may be due to the electron withdrawing
(-I effect) ability exerted by the fluoro substituent.
Antibacterial activity
All the synthesized imidazole derivatives were tested for their
antibacterial activity in vitro against Staphylococcus aureus,
Escherichia coli and Klebesiella pneumoniae. Ciprofloxacin
was used as reference drug the minimum inhibitory concen-
tration (MIC) values of which were furnished in Table 3
(Plate 1). Imidazole derivatives exerted antibacterial activity in
vitro at 25–100 lg/ml against all the tested strain. All com-
pounds exerted improved activity against all the tested strains.
Table 4 In vitro antifungal activity of imidazole derivatives
Antifungal activity
Compound
Minimum inhibitory concentration (MIC) in lg/mL
Candida-6
A. niger
A. flavus
The in vitro antifungal activity of imidazole derivatives
were examined against the fungal strains viz., Aspergillus
niger, Aspergillus flavus and Candida-6. Amphotericin-B
was used as a standard drug the minimum inhibitory con-
centration (MIC) values of which are furnished in Table 4
(Plate 2). Imidazole derivatives exhibited improved fungal
activity at 25–100 lg/ml against all the tested strains.
dmdmpfpi
dmmppi
50
25
25
25
50
100
25
25
50
50
25
25
25
25
dmmpfpi
dmdmppi
Amphotericin-B 12.5
Plate 2 Antifungal activity of
the imidazole derivatives
123

Med Chem Res (2012) 21:1850–1860
1859
Gayathri P, Jayabharathi J, Saravanan K, Thiruvalluvar A, Butcher RJ
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Conclusions
A series of substituted imidazoles have been synthesized
under solvent-free condition in the presence of molecular
iodine as the catalyst and characterized by NMR spectra,
X-ray, mass and CHN analysis. The novel imidazole
derivatives showed free radical scavenging activity when
tested in different models. Among all, dimethoxyphenyl
substituent at N3 of the imidazole derivatives exhibited the
highest hydroxy and superoxide anion radical scavenging
activities whereas dimethoxyphenyl substituent at N3 and
fluorophenyl at C2 of the imidazole derivatives exhibited
the highest DPPH radical scavenging activity. It is well
documented that free radicals are responsible for several
diseases. The present result confirms the free radical
scavenging activity of the imidazole derivatives and it can
be used for several diseases. A minute examination of in
vitro antibacterial and antifungal screening of imidazole
derivatives against the tested bacterial and fungal strains
provide a better structure activity and thus in future these
compounds may be used as templates to generate better
drugs to fight against bacterial and fungal infections.
Acknowledgments One of the authors, Dr. J. Jayabharathi, Asso-
ciate Professor in Chemistry, Annamalai University is thankful to the
Department of Science and Technology (No. SR/S1/IC-07/2007), the
University Grants Commission (F. No. 36-21/2008 (SR)) for pro-
viding fund to this research study, and Prof. C.H. Cheng, Chairman,
the National Science Council, the National Tsing Hua University for
her post-doctoral research study.
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