Synthesis, antimicrobial, analgesic activity, and molecular docking studies of novel 1-(5,7-dichloro-1,3-benzoxazol-2-yl)-3-phenyl-1H-pyrazole-4- carbaldehyde derivatives

Article abstract of DOI:10.1007/s00044-013-0565-9

Condensation of 5,7-dichloro-2-hydrazino-1,3-benzoxazole 3 with different aromatic acetophenones in methanol using catalytic amount of glacial acetic acid afforded the corresponding 1-phenylethanone(5,7-dichloro-1,3-benzoxazol-2-yl) hydrazones 5a-e in good yield. The compounds 5a-e, when subjected to Vilsmeier-Haack reaction with POCl₃ in DMF yielded (5,7-dichloro-1,3-benzoxazol-2-yl)-3-phenyl-1H-pyrazole-4-carbaldehyde derivatives 6a-e. The structural assignments of the compounds 6a-e are based on their spectral data and elemental analysis. The obtained compounds were tested for antimicrobial and analgesic activities, and subjected to molecular docking studies with respect to antimicrobial activity. The compound 6b showed pronounced antimicrobial and analgesic activity and exhibited an interesting binding profile with very high receptor affinity.

Full text of DOI:10.1007/s00044-013-0565-9

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-013-0565-9
ORIGINAL RESEARCH
Synthesis, antimicrobial, analgesic activity, and molecular docking
studies of novel 1-(5,7-dichloro-1,3-benzoxazol-2-yl)-3-phenyl-1H-
pyrazole-4-carbaldehyde derivatives
•
N. D. Jayanna H. M. Vagdevi J. C. Dharshan
•
•
•
R. Raghavendra Sandeep B. Telkar
Received: 21 November 2012 / Accepted: 27 February 2013
Ó Springer Science+Business Media New York 2013
Abstract Condensation of 5,7-dichloro-2-hydrazino-1,3-
benzoxazole 3 with different aromatic acetophenones in
methanol using catalytic amount of glacial acetic acid
afforded the corresponding 1-phenylethanone(5,7-dichloro-
1,3-benzoxazol-2-yl)hydrazones 5a–e in good yield. The
compounds 5a–e, when subjected to Vilsmeier–Haack
reaction with POCl3 in DMF yielded (5,7-dichloro-1,3-
benzoxazol-2-yl)-3-phenyl-1H-pyrazole-4-carbaldehyde
derivatives 6a–e. The structural assignments of the com-
pounds 6a–e are based on their spectral data and elemental
analysis. The obtained compounds were tested for antimi-
crobial and analgesic activities, and subjected to molecular
docking studies with respect to antimicrobial activity. The
compound 6b showed pronounced antimicrobial and
analgesic activity and exhibited an interesting binding
profile with very high receptor affinity.
Introduction
Human beings have been in constant exposure to pathogens
for many millennia. Microbial infections are a growing
problem in contemporary medicine, yet only a few antimi-
crobial agents are used in clinical practice. Although several
antimycobacterial agents are currently available, there is a
critical need for the development of new and specific anti-
mycobacterial agents. Among the heterocycles, benzoxazole
derivatives constitute much importance in medicinal field
due to their broad range of biological activity (Jauhari et al.,
2008; Rida et al., 2005). In recent years, pyrazoles belong to
one of the most important classes of heterocyclic compounds,
which are very useful in medicinal field. Molecules of many
modern drugs, such as antiphlogistic, antidiabetic, analgesic,
and insectoacaricides containing pyrazole ring as structural
fragment (Elguero et al., 2002; Grapov, 1999). Introduction
of an aldehyde group to pyrazole molecule was expected to
insure intramolecular heterocyclizations with the formation
of fused or linearly linked heterocyclic compounds (Bratenko
et al., 2008). Considering the importance of benzoxazole and
pyrazoles, it was planned to synthesize new derivatives of
pyrazole moiety associated with benzoxazole and screened
for antimycobacterial and analgesic activities.
Keywords Dichlorobenzoxazole ꢀ Antimicrobial ꢀ
Analgesic activity ꢀ Vilsmeier–Haack ꢀ Molecular docking
N. D. Jayanna ꢀ H. M. Vagdevi (&)
Department of Chemistry, Sahyadri Science College
(Autonomous), Shimoga 577 203, India
e-mail: vagdevihm@gmail.com
The application of the Vilsmeier–Haack reagent (POCl3/
DMF) in the formylation to aromatic and hetero aromatic
substrates is well documented (Jones and Stanforth, 2001).
Many of these reactions have led to the novel and conve-
nient routes for the synthesis of various heterocyclic com-
pounds (Majo and Perumal, 1998). A notable example that
finds significant application in heterocyclic chemistry is the
synthesis of 4-formylpyrazole from the double formylation
of hydrazones with Vilsmeier–Haack reagent (Selvi and
Perumal, 2002). These observations provide the recent
developments in the simple synthesis of pyrazole derivatives
J. C. Dharshan
Department of PG Studies and Research in Industrial Chemistry,
Sir M.V. Govt. Science College, Bommanakatte, Bhadravathi
577451, India
R. Raghavendra
Department of PG Studies and Research in Biochemistry, Jnana
Sahyadri, Kuvempu University, Shimoga 577 451, India
S. B. Telkar
Department of PG Studies and Research in Biotechnology
and Bioinformatics, Jnana Sahyadri, Kuvempu University,
Shimoga 577 451, India
123

Med Chem Res
(Elguero and Katrizky, 1984) and study of their various
biological activities (Bratenko et al., 1999; Cottineau et al.,
2002; El-Emary and Bakhite, 1999; Rathelot et al., 2002),
prompted us to undertake the synthesis of pyrazole-4-carb-
aldehyde derivatives using Vilsmeier–Haack reagent.
antibacterial activity (Chamara et al., 1985; Milewski et al.,
1988).
Results and discussion
In the present work, novel derivatives of benzoxazole
linked to pyrazole moiety containing aldehyde group were
synthesized. The synthesized compounds exhibited encour-
aging antimicrobial and analgesic activities. Hence, the in
silico molecular docking studies of newly synthesized
compounds were carried out to predict the glucosamine-6-
phosphate synthase inhibitory activity. The enzyme, namely
Glucosamine-6-phosphate synthase (GlmS, GlcN-6-P syn-
thase, L-glutamine: D-fructose-6P amidotransferase, EC
2.6.1.16) also known under the trivial name of glucosamine-
6-phosphate synthase, is a new target in the determination of
Chemistry
The compound 3 is the intermediate compound for the
synthesis of compounds 6a–e. The compound 3 was
obtained by the reaction of 5,7-dichloro-2-(ethylthio)-1,3-
benzoxazole 2 with hydrazine hydrate (Scheme 1). The
compound 3 was synthesized already in another route
(Satyendra et al., 2011), but it is possible to synthesize by
direct method. The compounds 1-(5,7-dichloro-1,3-ben-
zoxazol-2-yl)-3-phenyl-1H-pyrazole-4-carbaldehyde
Scheme 1 Synthetic route for
compound 3
Cl
Cl
NH2
OH
N
CS2/KOH
MeOH
SH
O
1
Cl
Cl
Ethyl iodide
DMSO
Cl
Cl
N
S
O
N
O
NH2NH2.H2O
MeOH
NH
NH2
CH3
2
3
Cl
O
Cl
Scheme 2 Synthetic route for
compounds 6a–e
CH3
Cl
Cl
N
N
O
Ethanol
Reflux
NH
N
CH3
NH
+
O
NH2
R1
Cl
Cl
R
3
5a-e
R1
4a-e
DMF/POCl3
R
Cl
Cl
R1
N
N
O
N
R
6a-e
O
Compounds
R
R1
H
H
H
H
6a
6b
6c
6d
6e
OCH3
OH
CH3
Br
Cl
Cl
123

Med Chem Res
maintained at National College of Pharmacy Shimoga. The
In vitro antimicrobial activity was carried out against 24-h
culture of four bacterial strains, such as Staphylococcus
aureus, Bacillus subtilis (Gram positive), Salmonella typhi,
Escherichia coli (Gram negative). The two fungal strains,
such as Aspergillus niger and Candida albicans, were used
to determine antifungal activity. The compounds were
tested at 40 lg/mL concentration against both bacterial and
fungal strains. DMSO was used as vehicle. Ciprofloxacin
(40 lg/100 lL) and Fluconazole (40 lg/100 lL) were
used as standard drugs for the comparison of antibacterial
and antifungal activities, respectively. The zone of inhibi-
tion was compared with standard drug after 24 h of incu-
bation at 37 °C for antibacterial activity and 72 h at 25 °C
for antifungal activity. All the tested compounds were
considered to be equipotent since the values obtained were
very close to each other. The compounds 6b and 6d
exhibited higher antimicrobial activity as compared with
standard drugs. The results are recorded in Table 2.
Table 1 Physical data of compounds 6a–e
Compounds Molecular
formula
Molecular M.P. (°C) Yield (%)
weight
6a
6b
6c
6d
6e
C18H11Cl2N3O3
388.20
374.17
372.20
198–197
197–198
199–200
141–142
159–160
87
85
89
90
81
C
C
C
C
17H9Cl2N3O3
18H11Cl2N3O2
17H8BrCl2N3O2 439.80
17H7Cl4N3O2 427.06
derivatives 6a–e were synthesized using 5,7-dichloro-2-
hydrazino-1,3-benzoxazole 3 (Scheme 2). The structure of
newly synthesized compounds were confirmed by IR,1H
NMR, 13C NMR, LCMS, and elemental analysis. Broad
singlet at d 4.6 for two protons of –NH2 and another broad
1
singlet at d 9.3 for –NH proton (D2O exchangeable) in H
NMR evidences the formation of compound 3. For the
compounds 6a–e, the singlet obtained at d 10.05 in 1H NMR
and the strong stretching absorption at 1,682 cm-1 in IR
spectrum confirmed the formation of aldehyde group. Also,
the pyrazole ring –C=CH proton signal at d 9.49 in 1H NMR
evidences the formation of target molecules. The physical
data of the newly synthesized compounds are tabulated in
Table 1.
Minimum inhibitory concentrations (MIC)
The MIC of all the synthesized compounds was determined
by microdilution method (Harish et al., 2007, Raghavendra
and Neelagund, 2009). The respective clinical strain was
spread separately on the medium. The wells were created
using a stainless steel sterilized cork borer under aseptic
conditions. The synthesized compounds at different con-
centrations, viz., 10, 20, 30, 40, and 50 lg were dissolved
respectively in 25, 50, 75, 100, and 125 lL of DMSO and
later loaded into corresponding wells. The drugs Cipro-
floxacin (40 lg in 100 lL) and Fluconazole (40 lg
in100 lL) were used as standard for the comparison of
antibacterial and antifungal activities, respectively. The
results were recorded in mm and presented in Table 3.
Antimicrobial activity
The antimicrobial activity of the synthesized compounds
was carried out by agar well diffusion method (Sharath
et al., 2008; Raghavendra and Neelagund, 2009; Saundane
et al., 1998). The bacterial strains were collected from
patients with different infectious status who had not
administered any antibacterial drugs for at least two weeks
with the suggestions of an authorized physician in Kiran
diagnostic health Centre of Chitradurga, Karnataka state,
India. Fungal strains were procured from the culture
Table 2 Anti-microbial activity of compounds 6a–e
Zone of inhibition in (mm)
Compounds
Antibacterial strains
Antifungal strains
S. aureus
B. subtilis
S. typhi
E. coli
A. niger
C. albicans
6a
19 9.50
23 11.50
21 10.50
23 11.50
22 11.50
23 11.50
–
20 10.0
23 11.50
20 10.0
22 11.0
22 11.50
23 11.50
–
19 9.50
21 10.50
20 10.0
21 10.50
20 10.0
21 10.50
–
22 11.0
22 11.0
21 10.50
22 11.0
22 11. 0
22 11.0
–
20 10.0
22 11.0
19 9.50
22 11.0
22 11.0
–
20 10.0
22 11.0
19 9.50
23 11.50
22 11.0
–
6b
6c
6d
6e
Ciprofloxacin
Fluconazole
Control
22 11.0
0
23 11.50
0
0
0
0
0
The value of each constituents consisted of Mean SE of 3 replicates
Value is significantly different when p \ 0.05
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Med Chem Res
Table 3 Minimum Inhibitory Concentration (MIC) of compounds 6a–e
MIC (lg/lL)
Compounds
Antibacterial strains
Antifungal strains
S. aureus
B. subtilis
S. typhi
E. coli
A. niger
C. albicans
10–50 (lg)
6a
30 15
20 10
30 15
20 10
30 15
0
40 20
20 10
30 15
20 10
30 15
0
40 20
30 15
30 15
30 15
30 15
0
40 20
30 15
40 20
20 10
30 15
0
30 15
30 15
30 15
20 10
40 20
0
40 20
20 10
30 15
30 15
40 20
0
6b
6c
6d
6e
Control
The value of each constituents consisted of Mean SE of 3 replicates
Value is significantly different when p \ 0.05
Analgesic activity
ð1 ꢁ Vc=VtÞ ꢂ 100
Where Vt = Mean number of writhing in test animals
and Vc = Mean number of writhing in control.
The analgesic activity was performed by acetic acid-
induced writhing in mice (Vagdevi et al., 2001) and
approved by the Institutional Animal Ethical Committee
(IAEC), National College of Pharmacy, Shimoga. Five
groups of six male Swiss albino mice, each 25–35 g of
body weight, were used. Acetic acid (0.6 % dose = 10
mL/Kg of body weight) was injected intraperitoneally. The
number of writhes was counted for 20 min, after 5 min of
injection of acetic acid into each mice. This reading was
taken as control. Next day, same group of mice was used
for evaluating analgesic activity. Each group was admin-
istered orally with the suspension of test compound (0.1 %
Tween-80 solution) at a dose of 100 mg/kg body weight of
animal was given 1 h before injecting acetic acid. After
5 min of acetic acid injection, mice were observed for the
number of writhes for the duration of 20 min. The mean
value for each group was calculated and compared with
control. Acetyl salicylic acid was used as a standard drug
for the comparison of analgesic activity. The compounds
6b and 6d exhibited potent analgesic activity and the
results are recorded in Table 4. Percent protection was
calculated by the following formula:
In silico molecular docking studies
Considering the results obtained in in vitro study, it was
thought worthwhile to perform molecular docking studies,
screening the compounds to both in silico and in vitro
results. Considering GlcN-6-P synthase as the target
receptor, comparative and automated docking studies with
synthesized candidate, lead compound was performed to
determine the best in silico conformation. The Lamarckian
genetic algorithm was inculcated in the docking program.
Auto Dock 4.2 was employed to satisfy the purpose
(Morris et al., 2007). Figure 1 was considered as the native
crystal structure of GlcN-6-P synthase (X chain) in com-
plex with glucosamine-6-phosphate, obtained from Protein
Data Bank (http://www.pdb.org/pdb/home/home.do) with
˚
PDB ID 2VF5, which was resolved at 2.90 A by X-ray
diffraction (Mouilleron et al., 2008).
The docking receptor of GlcN-6-P with newly synthe-
sized candidate ligands exhibited well-established bonds
with one or more amino acids in the receptor active pocket.
The active pocket was considered to be the site, where
glucosamine-6-phosphate complexes in GlcN-6-P of 2VF5.
The active pocket consists of 12 amino acid residues as
Ala602, Val399, Ala400, Gly301, Thr302, Ser303, Cys300,
Gln348, Ser349, Thr352, Ser347, and Lys603 as shown in
Fig. 2. The synthesized ligand molecules having 2D
structure were converted to energy minimized 3D structure
and further used for in silico protein–ligand docking.
Seven molecules were docked; among them, five were
synthesized and two were standard drugs such as Cipro-
floxacin and Fluconazole, as antibacterial and antifungal,
respectively. The docked images of all the seven molecules
Tables 4 Analgesic activity of the synthesized compounds 6a–e
Compounds Dose
mg/kg
Mean no. of writhing
Before drug After drug
% Protection
6a
100
100
100
100
100
100
34.5 0.85 17.16 0.48 50.27
34.66 0.93 15.33 0.51 55.78
6b
6c
34.5 0.85
34.16 0.70
31.0 0.73
31.5 0.44
16.8 0.40 51.40
15.5 0.44 54.63
15.5 1.09 50.00
13.0 0.36 58.74
6d
6e
Standard
Values are in mean SEM, (Standard error mean)
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Med Chem Res
hydrogen bound to Thr352, compound 6b hydrogen bound
to Thr352 and Glu488. The compound 6d hydrogen bound
to Thr352, but the compounds 6c and 6e have no hydrogen
bonds. Among the synthesized compounds, 6b ligand
docked in the best of its conformation and it is shown in
Fig. 4.
Experimental section
Chemistry
Melting points were recorded on electro thermal melting
point apparatus and are uncorrected. 1H NMR and 13C
NMR spectra were recorded on Bruker 400 MHz spec-
trometer. Chemical shifts were shown in d values (ppm)
with tetramethylsilane (TMS) as an internal standard.
LC–MS were obtained using Atlantis DC18 (50 9 4.6) mm
column on Shimadzu, LCMS 2010A, Japan. The 0.1 %
HCOOH and methanol were used as mobile phase at the
flow rate of 1 mL/min. The FT-IR spectra of compounds
were taken in KBr pellets (100 mg) using Shimadzu Fourier
transformed infrared (FT-IR) spectrophotometer. Column
chromatography was performed using silica gel (60–120
mesh). Silica gel GF254 plates from Merck were used for
TLC and the spots were observed by UV light. The chem-
icals were purchased from Sigma Aldrich Co. and solvents
for column chromatography were of reagent grade and
purchased from commercial source.
Fig. 1 Crystal structure of X chain of GlcN-6-P synthase complex
with glucosamine-6-phosphate
Preparation of 5,7-dichloro-1,3-benzoxazole-2-thiol (1)
To a solution of methanol (50 mL), KOH (1.1 eq) was added
and stirred for 10 min followed by slow addition of CS2 at
room temperature. To the reaction mass 2-amino-4,
6-dichlorophenol was added on stirring. The reaction mass
was refluxed for 6 h on water bath. The completion of the
reaction was confirmed by TLC and reaction mass was poured
onto ice cold water, followed by the acidification with glacial
acetic acid (pH 6). The obtained solid was filtered, dried, and
recrystallized using ethanol.Yield(95 %), M.P. 198 °C.
Fig. 2 PDB sum’s ligplot results of 2VF5 (showing all 12 amino acid
residues of active pocket)
are presented in Fig. 3. The Table 5 showing binding
energy and inhibition constant of seven compounds
including both the standards. In silico studies revealed that
all the five synthesized molecules exhibited required
binding energy toward the target protein ranging from
-8.19 kJ/mol to -5.65 kJ/mol.
Preparation of 5,7-dichloro-2-(ethylthio)-1,3-
benzoxazole (2)
To a mixture of sodium hydroxide (10 mol) and compound
1 (10 mol) in DMSO (10 mL), ethyl iodide (10 mol) was
added drop wise. The reaction mass was stirred about 1 h
and the completion of the reaction was confirmed by TLC.
The reaction mass was poured onto ice cold water. The
obtained solid was filtered, dried, and recrystallized using
hexane.
The conformations of docked ligands are shown in
Fig. 3. Images a–g refer for compounds 6a, 6b, 6c, 6d, 6e,
Ciprofloxacin, and Fluconazole, respectively, representing
the best of their docked conformation. In compound 6a,
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Med Chem Res
Fig. 3 Ligands docked in best of its conformation
Yield (90 %), M.P. 38 °C, IR (KBr) cm-1: 2,974 (CH
alkyl), 1,608 (C=N), 1,491 (C=C), 756 (C–Cl); H NMR
Preparation of 5,7-dichloro-2-hydrazino-1,3-
benzoxazole (3)
1
(CHCl3) ppm: d 1.4 (t, 3H, CH3), d 3.26 (q, 2H, CH2), d 7.1
(s,1H, Ar H) d 7.4 (s,1H, Ar H), 13C NMR (CHCl3) ppm: d
124.85–144.4 (7C, sp2 carbon atoms), d 14.29 (CH2), d
13.22 (CH3), Analyses: C9H7Cl2NOS (248.1). m/z 248,
M?2 250, M?4 252, Required: C (43.56 %) H (2.84 %) N
(5.64 %) Found: C (43.54) H (2.82), N (5.63).
Compound 2 (10 mol) and hydrazine hydrate (15 mol) in
ethanol (15 mL) was taken in a round bottomed flask and
refluxed for 3 h. The completion of the reaction was con-
firmed by TLC and the reaction mixture was cooled and
filtered. The obtained solid was recrystallized from ethanol.
123

Med Chem Res
Tables 5 Binding Energy (kJ mol-1), Inhibition constant of com-
pounds 6a–e
Two drops of glacial acetic acid was added while stirring
and the mixture was refluxed for 3 h. The solid obtained on
cooling was filtered, dried, and recrystallized from ethanol.
Compounds 5b–e were synthesized by similar method.
Compounds
Binding energy (kJ mol-1
)
Inhibition constant lM
6a
6b
6c
6d
6e
-7.03
-8.19
-6.80
-7.22
-7.78
7.07
996.39
10.37
5.08
Preparation of 1-(5,7-dichloro-1,3-benzoxazol-2-yl)-
3-(4-methoxyphenyl)-1H-pyrazole-4-carbaldehyde (6a)
1.99
To the Vilsmeier–Haack reagent prepared from DMF
(10 mL) and POCl3 (1.1 mL, 0.0012 mol), hydrazone (5a)
(0.01 mol) was added and the reaction mixture was stirred
at 0–5 °C for 1 h and at 60 °C temperature for 3 h. The
reaction mass was poured to ice cold water, filtered, and
washed with water. The compound was dried and purified
using methanol to afford compounds 6a.
Ciprofloxacin -6.48
Fluconazole -5.65
17.93
72.72
Compounds 6b–e have been synthesized from similar
procedure.
1-(5,7-Dichloro-1,3-benzoxazol-2-yl)-3-(4-methoxy-
phenyl)-1H-pyrazole-4-carbaldehyde (6a) IR (KBr)
cm-1: 3,073 (OCH3), 1,684 (HC=O), 1,466 (C=C), 767
(C–Cl); 1H NMR (DMSO-d6) ppm: d 3.84 (s, 3H, OCH3), d
7.1 (d, 2H, Ar H, J = 8.88, 6.92), d 7.96 (d, 2H, Ar H,
J = 6.48, 2.52), d 7.76 (s, 1H, Ar H), d 7.97 (s, 1H, Ar H),
d 9.43 (s, 1H, pyrazole =CH), d 10.05 (s, 1H, CHO),13C
NMR (DMSO-d6): d 114–161.1 (16C, sp2 carbon atoms), d
187 (CHO), d 57 (OCH3), m/z 388.2, M?2 390, m?4 392,
Analyses: C18H11Cl2N3O3, Required: C (55.69 %) H
(2.86 %) N (10.82 %) Found: C (55.05 %) H (2.24 %) N
(10.87 %).
1-(5,7-Dichloro-1,3-benzoxazol-2-yl)-3-(4-hydroxy-
phenyl)-1H-pyrazole-4-carbaldehyde (6b) IR (KBr) cm-1
:
1
3,332 (OH), 1,680 (HC=O), 1,445 (C=C) 770 (C–Cl); H
NMR (DMSO-d6) ppm: d 6.9 (d, 2H, Ar H, J = 9.56,
9.52), d 7.7 (s, 1H, –OH), d 7.84 (d, 2H, Ar H, J = 8.64,
1.72), d 7.75 (s, 1H, Ar H), d 7.94 (s, 1H, Ar H), d 9.49
(s, 1H, pyrazole =CH), d 10 (s, 1H, CHO), 13C NMR
(DMSO-d6): d 109–159.0 (16C, sp2 carbon atoms), d 187
(CHO), m/z 374.17, M?2 376, M?4 378, Analyses:
C17H9Cl2N3O3, Required: C (54.57 %) H (2.42 %) N
(11.23 %) Found: C (54.52 %) H (2.41 %) N (11.22 %).
Fig. 4 Image showing the crevice of the portion of the target protein
GlcN-6-P and 6b ligand docked in best of its conformation
Yield (70 %), M.P. 220 °C, IR (KBr) cm-1: 3,349 (NH2),
3,283 (NH), 1,617 (C=N), 1,453 (C=C), 771 (C–Cl), H
1
NMR (DMSO-d6)ppm: d 4.6 (s, 2H, –NH2, D2O exchange-
able), d 9.3 (s,1H, –NH, D2O exchangeable), d 7.18 (s,1H, Ar
H) d 7.29 (s,1H, Ar H); 13C NMR (DMSO-d6): 122.9–143.0
(7C, sp2 carbon atoms), Analyses: C7H5Cl2N3O (218.04).
m/z218, M?2 220, M?4 222, Required: C (38.56 %) H (2.31 %)
N (19.27 %) Found: C (38.53 %) H (2.30 %) N (19.25 %).
1-(5,7-Dichloro-1,3-benzoxazol-2-yl)-3-(4-methoxy-
phenyl)-1H-pyrazole-4-carbaldehyde (6c) IR (KBr) cm-1
:
3,139(CH3), 1,681 (HC=O), 1,465 (C=C), 766 (C–Cl);
1H NMR (DMSO-d6) ppm: d 2.48 (s, 3H, –CH3), d 7.35
(d, 2H, Ar H, J = 8.04), d 7.87 (d, 2H, Ar H, J = 8.16), d
7.77 (s, 1H, Ar H), d 7.97 (s, 1H, Ar H), d 9.49 (s, 1H,
pyrazole =CH), d 10.06 (s, 1H, –CHO),13CNMR (DMSO-
d6): d 112–161.0 (16C, sp2 carbon atoms), d 187 (CHO), d
52 (CH3), m/z 372.2, M?2 374, M?4 376, Analyses:
Preparation of 1-(4-methoxyphenyl)ethanone(5,7-
dichloro-1,3-benzoxazol-2-yl)hydrazone (5a)
To a solution of compound 3 (10 mol) in absolute ethanol
(30 mL), p-methoxyacetophenone (10 mol) was added.
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Med Chem Res
´
˚
C18H11Cl2N3O2, Required: C (58.08 %) H (2.98 %) N
(11.29 %) Found: C (58.06 %) H (2.95 %) N (11.27 %).
and the grid center was set to 30.59, 15.822 A, and -3.497
for x, y, and z, respectively, which covered all the 12 amino
acid residues in the considered active pocket. Docking soft-
ware AutoDock 4.2 Program supplied with AutoGrid 4.0 and
AutoDock 4.0 was used to produce grid maps. The spacing
1-(5,7-Dichloro-1,3-benzoxazol-2-yl)-3-(4-bromophe-
nyl)-1H-pyrazole-4-carbaldehyde (6d) IR (KBr) cm-1
:
´
1,692 (HC=O), 1,445 (C=C), 767 (C–Cl); 1H NMR
(DMSO-d6) ppm: d 7.75 (d, 2H, Ar H,), d 7.99 (d, 2H, Ar
H,), d 7.5 (s, 1H, Ar H), d 8.1 (s, 1H, Ar H), d 9.56 (s, 1H,
pyrazole =CH), d 10.05 (s, 1H, –CHO), 13C NMR (DMSO-
d6): d 110–159.3.0 (16C, sp2 carbon atoms), 187 (CHO), m/
between grid points was 0.375 A. The Lamarckian Genetic
˚
Algorithm (LGA) was chosen to search for the best con-
formers. During the docking process, a maximum of 10
conformers was considered for each compound. All the
AutoDock docking runs were performed in Intel Centrino
Core2Duo CPU @ 2.20 GHz of IBM system origin with
2 GB DDR2 RAM. AutoDock 4.0 was compiled and run
under Windows XP operating system.
z
437.07, M?2 439.08, M?4 441: Analyses
C17H8BrCl2N3O2, Required: C (46.72 %) H (1.84 %) N
(9.61 %) Found: C (46.70 %) H (1.83 %) N (9.60 %).
1-(5,7-Dichloro-1,3-benzoxazol-2-yl)-3-(3,4-dichloro-
phenyl)-1H-pyrazole-4-carbaldehyde (6e) IR (KBr)
Conclusion
1
cm-1: 1,695 (HC=O), 1,461 (C=C), 767 (C–Cl); H NMR
(DMSO-d6) ppm: d 7.59 (d, 2H, Ar H), d 7.77 (s, 1H, Ar
H), d 7.84 (s, 1H, Ar H), d 7.98 (s, 1H, Ar H), d 9.57 (s, 1H,
pyrazole =CH), d 9.96 (s, 1H, –CHO),13C NMR (DMSO-
d6): d 113–160.5.0 (16C, sp2 carbon atoms), d 187 (CHO),
m/z 427.06, M?2 429, M?4 431, M?6 433, Analyses:
C17H7Cl4N3O2, Required: C (47.81 %) H (1.65 %) N
(9.84 %) Found: C (47.80 %) H (1.64 %) N (9.83 %).
This study reports the successful synthesis of the title
compounds in good yield and characterized by IR, 1H
NMR, 13C NMR, and Mass spectral analysis. The data
reported here clearly indicated that the synthesized com-
pounds have shown considerable antimicrobial activity.
Among the compounds, 6b and 6d were emerged as
potentially active antimicrobial as well as analgesic agents.
In molecular docking studies, compound 6b showed min-
imum binding energy. The compounds with minimum
binding energy are responsible for more active antimicro-
bial agent with respect to standard drugs. The best dock
conformation is one with least binding energy has the
highest affinity.
Molecular docking studies
The ligands were drawn in ChemDraw Ultra 6.0 (Chem
Office package) assigned with proper 2D orientation and the
structure of each compound was analyzed for connection
error in bond order. OSIRIS, an ADMET-based Java library
layer that provides reusable cheminformatics functionality, is
an entirely in-house-developed drug discovery informatics
system used to predict the total drug score via in silico (Bates
et al., 1966). Energy of the molecules was minimized using
Dundee PRODRG2 server (Chamara and Borowski, 1986).
The energy-minimized compounds were then read as input
for Auto Dock 4.2 to carry out the docking simulation
(Milewski et al., 1986). All heteroatoms were removed from
the 2VF5 pdb to make complex receptor free of any ligand
before docking. The Graphical User Interface program
‘‘Auto Dock Tools’’ was used to prepare, run, and analyze the
docking simulations. Kollman united atom charges, salvation
parameters, and polar hydrogens were added to the receptor
for the preparation of protein in docking simulations. In the
present study, the binding site was selected based on the
amino acid residues, which were involved in binding with
glucosamine-6-phosphate of GlcN-6-P synthase as obtained
from PDB with ID 2VF5. Therefore, the grid was centered at
the region including all the 12 amino acid residues (Ala602,
Val399, Ala400, Gly301, Thr302, Ser303, Cys300, Gln348,
Ser349, Thr352, Ser347, and Lys603) that surround active
Acknowledgments The author (N.D.J) is grateful to UGC for
providing Rajiv Gandhi Research Fellowship. The authors are
thankful to Director, IISc, Bangalore, for providing spectral data and
thankful to the Principal, Sahyadri Science College, Shimoga, for
providing laboratory facilities to carry out research work.
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