Design and synthesis of positional isomers of 1-alkyl-2-trifluoromethyl-5 or 6-substituted benzimidazoles and their antimicrobial activity

Article abstract of DOI:10.1007/s00044-012-0131-x

A series of novel positional isomers of 1-alkyl-2-trifluoromethyl benzimidazole derivatives 3a-j and 4a-j have been synthesized and screened for antibacterial activity against gram positive bacteria viz, Bacillus subtilis, Staphylococcus aureus and gram negative bacteria viz., Escherichia coli, Pseudomonas aeruginosa and antifungal activity against Candida albicans, Aspergillus niger, Rhizopus oryzae and Saccharomyces cerevisiae. Results indicate that compounds 3b, 4b were having promising antibacterial and antifungal activity.

Full text of DOI:10.1007/s00044-012-0131-x

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2013) 22:1229–1237
DOI 10.1007/s00044-012-0131-x
ORIGINAL RESEARCH
Design and synthesis of positional isomers of 1-alkyl-2-
trifluoromethyl-5 or 6-substituted benzimidazoles and their
antimicrobial activity
•
G. Sathaiah A. Ravi Kumar A. Chandra Shekhar
•
•
•
K. Raju P. Shanthan Rao B. Narsaiah
•
•
•
A. Raghuram Reddy D. Lakshmi B. Sridhar
•
Received: 7 March 2012 / Accepted: 25 May 2012 / Published online: 9 June 2012
ꢀ Springer Science+Business Media, LLC 2012
Abstract A series of novel positional isomers of 1-alkyl-
2-trifluoromethyl benzimidazole derivatives 3a–j and 4a–
j have been synthesized and screened for antibacterial
activity against gram positive bacteria viz, Bacillus subtilis,
Staphylococcus aureus and gram negative bacteria viz.,
Escherichia coli, Pseudomonas aeruginosa and antifungal
activity against Candida albicans, Aspergillus niger, Rhi-
zopus oryzae and Saccharomyces cerevisiae. Results indi-
cate that compounds 3b, 4b were having promising
antibacterial and antifungal activity.
activity as drugs (Denny et al., 1990) pesticides (Banks,
1979) and so on. In particular, trifluoromethyl benzimi-
dazoles are considered as antiparasitic (Gabriel et al., 2001;
Juan et al., 2002), antiprotozoal (Gabriel et al., 2006;
Fransisco et al., 2010) agents. Benzimidazole is the core
nucleus present in anthelmintic drugs such as albendazole
and mebendazole. Benzimidazoles derived copper com-
plexes (Farukh et al., 2005) also reported as antibacterial/
antifungal agents. The antimicrobial activity of imidazoles
and benzimidazoles have long been established as anti-
bacterial (Nakamura, 2005), fungicidal (Keith et al., 1978
and Maxwell and Broody, 1971) and as trichomonocidal
activities. The emergence of bacterial resistance to various
classes of antibiotics including b-lactum antibiotics, mac-
rolides, quinolones, became a major worldwide health
problem.
Keywords Benzimidazole Á Trifluoromethyl Á
Antifungal Á Antibacterial Á Rhizhopus oryzae
Introduction
In view of the great importance of antibacterial com-
pounds in animal and particularly human health, it can be
seen that there is a need to identify a new and novel anti-
bacterial agents. Our continued interest on the synthesis of
fluorinated heterocycles (Venkat Reddy et al., 2009;
Kurumurthy et al., 2011) promoted us to synthesize 2-tri-
fluoromethyl benzimidazoles system as a promising scaf-
fold for the development of new antimicrobial agents.
Synthesis of fluorinated heterocycles with positioning of
fluorine or trifluoromethyl group at a strategic position is
expected to enhance the activity due to its lipophilic nature
(Banks, 1982). Benzimidazoles are an important class of
potential organic molecules due to their wide range of
G. Sathaiah Á A. Ravi Kumar Á A. Chandra Shekhar Á K. Raju Á
P. Shanthan Rao (&) Á B. Narsaiah
Fluoroorganic Division, Indian Institute of Chemical
Technology, Tarnaka 500607, Hyderabad, India
e-mail: shanthanpp1954@gmail.com
Chemistry
The 2-trifluoromethyl/2-pentafluoroethyl benzimidazoles
2a–d were prepared from 4-nitro or 4-benzoyl o-phenylene
diamine on refluxing trifluoro acetic acid or pentafluoro
propionic acid for 30–60 min to furnish the corresponding
5(or 6)-nitro or benzoyl fluoroalkyl benzimidazoles in high
yield. The reaction pathway is mainly by selective attack of
B. Sridhar
Department of X-ray Crystallography, Indian Institute of
Chemical Technology, Tarnaka 500607, Hyderabad, India
A. Raghuram Reddy Á D. Lakshmi
UCPSc, Kakatiya University, Warangal, India
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Med Chem Res (2013) 22:1229–1237
amine nucleophile meta to nitro or benzoyl group on acid
carbonyl followed by cyclisation through participation of
amine para to nitro or benzoyl group. This is attributed to
the relatively high basicity of amine which participates
first to drive the reaction (Scheme 1). The compounds
2a–d were independently alkylated using alkyl halide by
conventional method at room temperature for 3–4 h, fur-
nished two positional isomers in each case 3a–j, 4a–j and
were separated by a long column of silica gel. The isomers
are characterized as 2-trifluoromethyl-6-nitro/benzoyl
benzimidazoles 3a–j and 2-trifluoromethyl-5-nitro/benzoyl
benzimidazoles 4a–j based on IR, 1H_NMR and mass
spectral data. The benzimidazole acidic proton is in equi-
librium between 1 and 3 positions as a result two positional
isomers 3 and 4 are formed during the alkylation. The
inorganic base abstracts the imidazole proton leads to the
generation of anion on nitrogen. The anion on nitrogen
para to electron withdrawing groups (–NO2, –COPh) may
be more stable by resonance effect, thus it is reflecting on
the formation of positional isomers 4 is slightly higher
yield comparing to the other isomer 3. The yield of the
products (3, 4) is in the ratio 2:3, respectively. Whereas,
reaction of 4-methyl-o-phenylene diamine in trifluoro
acetic acid followed by alkylation afforded products 3 and
4 in 1:1 ratio, supporting the earlier observation. When
alkylation reaction was carried out in sonicator, the reac-
tion time was drastically reduced to 10–15 min. The details
are outlined in Scheme 1 and products are tabulated in
Table 1.
proton of C4 appeared as singlet at d 8.81, C6 as doublet
at d 8.38 and for C7 as doublet at d 7.52. The presence of
N-alkyl para to nitro group in benzene promotes electron
density as a result shifting of aromatic protons to slightly
upfield is observed in compound 4a compared to com-
pound 3a. The same trend in chemical shift difference is
followed for all other compounds. The characterization of
each product is further supported by nuclear Overhauser
effect (NOE) experiments.
NOE studies
In order to confirm the structure of two isomers, a repre-
sentative example of compound 3a and 4a was analysed by
NOESY 1D NMR experiments. On irradiation of H7 pro-
ton in 3a resulted disappearance of H4 proton which away
from H7 proton and enhancement of H6 proton which is
close to H7 proton is observed. Similarly, on irradiation of
H7 proton of compound 4a resulted disappearance of H4
proton which is para to H7 and enhancement of H6 proton
close to H7 proton is observed. Thus, the characteristic
difference in NOE of compound 3a and 4a confirm the
structures.
In order to confirm the structure of isomers unambigu-
ously, one of the isomer was prepared through series of
reactions as shown in Scheme 2. The 2,4-dinitrochloro-
benzene was reacted with ethylamine to give 5. The
presence of two electron withdrawing groups at o- and
p-positions promote the substitution of chlorine with eth-
ylamine. The compound 5 on reduction in presence of
catalytic quantity of palladium on carbon and hydrazine
hydrate in refluxing ethanol furnished 2,4-diamino-N-ethyl
aniline 6. The latter on treatment with trifluoro acetic acid
In a specific example of compound 3a and 4a, a char-
acteristic chemical shift difference of aromatic protons in
NMR is observed. For compound 3a, proton at C4
appeared as doublet at d 7.95, C5 as doublet at d 8.34, and
for C7 as singlet at d 8.41, whereas for compound 4a,
R2
H
XCH2R2
N
NH2
NH2
N
Reflux
N
R1
R1
K2CO3/ DMF
or
SONICATOR
R1
R1COOH
N
N
N
R
R
R
R
2a-d
4a-j
1
3a-j R2
H
N
NH2
R2= -CH3,-CH2CH2Cl
-CH2CH2CH2Cl,-CN
NH2
O
OH
R1
OH
C
CH,-COOEt
N
H
N
H
N
R
R1
O
R1
R
R
H
H
X=Cl,Br,I
2a, R= -NO2, R1= -CF3
2b, R= -NO2, R1= -CF2CF3
2c, R= -COPh, R1= -CF3
2d, R= -COPh, R1= -CF2CF3
Scheme 1 The mechanistic aspects of synthesis of benzimidazoles
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Med Chem Res (2013) 22:1229–1237
1231
Table 1 Physical properties of
benzimidazoles
Compound
Yield (%) m.p (°C)
Compound
Yield (%)
60
m.p (°C)
Compound
no.
Compound
no.
N
N
3a
40
177–179
4a
125–127
CF3
N
N
O2N
CF3
O2N
N
N
3b
38
150–152
4b
62
80–85
CF3
O2N
3
Cl
Cl
N
N
3c
35
40
140–142
145–150
4c
65
60
70–74
93–95
CF3
4
N
O2N
CF3
4
Cl
N
O2N
N
N
N
3d
4d
CF3
O2N
N
N
CF3
N
O2N
N
N
3e
40
120–125
4e
60
75–80
CF3
O2N
N
N
CF3
CF3
F
O2N
EtOOC
N
3f
3g
3h
3i
34
32
32
30
120–123
85–89
4f
4g
4h
4i
66
68
68
70
75–80
80–82
90–92
90–92
CF3
N
N
N
O2N
EtOOC
O2N
Cl
N
F
4
F
CF3
N
N
O2N
4
4
Cl
F
CF3
N
O2N
EtOOC
N
N
124–126
120–122
F
F
CF3
N
N
O2N
F
F
CF3
EtOOC
O2N
EtOOC
N
N
CF3
O
N
N
CF3
O
COOEt
EtOOC
N
N
3j
27
75–80
4j
73
85–88
F
F
O
CF3
N
N
F
O
F
CF3
COOEt
under refluxing conditions gave cyclised product found
to be 1-ethyl-2-trifluoromethyl-5-trifluoroacetamido benz-
imidazole derivative 7. The compound 7 on hydrolysis in
refluxing 10 % aqueous sodium hydroxide solution fur-
nished 1-ethyl-2-trifluoromethyl-5-amino benzimidazole 8.
The reduction of 4a also afforded compound 8. The com-
pound 8 obtained in series of steps is compared with the
product obtained by reduction of 4a and found to be the
same, by super imposable IR and their mixed melting point
is not depressed.
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Med Chem Res (2013) 22:1229–1237
Cl
NHC2H5
NO2
NHC2H5
TFA
C2H5NH2
Rt, 15Min
Pd/C, NH2-NH2.H2O
EtOH,Reflux, 5hrs
Reflux,2hrs
NH2
O2N
NO2
O2N
H2N
5
6
N
N
N
N
N
Pd/C, NH2-NH2.H2O
10% Aq.NaOH
Reflux, 5-6hrs
CF3
CF3
CF3
EtOH,Reflux, 5hrs
O2N
N
H2N
F3COCHN
7
4a
8
Scheme 2 Chemical method for isomer identification
X-ray studies
The structures of one pair of isomers was established by
X-ray analysis. X-ray data for the compounds 3a and 4a
were collected at room temperature using a Bruker Smart
Apex CCD diffractometer with graphite monochromated
˚
MoKa radiation (k = 0.71073 A) with x-scan method.
Preliminary lattice parameters and orientation matrices
were obtained from four sets of frames. Unit cell dimen-
sions were determined using 3483 reflections in the range
of 3.03 \ h \ 23.82ꢁ for AE87 and 3780 reflections in the
range of 2.64 \ h \ 27.99ꢁ for AE88 integration and
scaling of intensity data were accomplished using SAINT
program (Bruker, 2001). The structure was solved by direct
methods using SHELXS97 (Sheldrick, 2008) and refine-
ment was carried out by full-matrix least-squares technique
using SHELXL97. Anisotropic displacement parameters
were included for all non-hydrogen atoms. All H atoms
were positioned geometrically and treated as riding on
Fig. 1 A view of AE87(3a), showing the atom-labelling scheme (1-
ethyl-2-trifluoromethyl-6-nitro benzimidazole)
˚
their parent C atoms [C–H = 0.93–0.97 A and Uiso(H) =
1.5 Ueq(C) for methyl H or 1.2 Ueq(c) for other H atoms].
The methyl groups were allowed to rotate but not to tip.
Displacement ellipsoids are drawn at the 30 % probability
level and H atoms are represented by circles of arbitrary
radii (Figs. 1, 2)
Antimicrobial activity
Antibacterial activity
Fig. 2 A view of AE87(4a), showing the atom-labelling scheme
(1-ethyl-2-trifluoromethyl-6-nitro benzimidazole)
The antibacterial activity of compounds 3a–j and 4a–
j were screened against different bacterial strains, viz.,
B. subtilis (NCIM 2921), S. aureus (NCIM 2079) and gram
negative bacteria viz., E. coli (NCIM 2068), P. aeruginosa
(NCIM 2957). Compounds 3c, 4c, 3f, 4f, 3h, 4h, 3j and 4j
showed moderate activity against bacterial strains. It is
found that the inhibition is concentration dependent as it
increases with higher concentration. In conclusion, 3b and
4b are the most active among the screened compounds.
The inhibitory zone diameters of the compounds are
compared with those obtained with 50 lg/ml of standard
ciprofloxacin. The details of each compound minimum
inhibitory concentration of each compound have been
tabulated in Table 2.
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Med Chem Res (2013) 22:1229–1237
1233
Table 2 Minimum inhibitory concentration of compounds 3a–j and
4a–j against various bacterial strains
Table 3 The zone of inhibition of compounds 3a–j and 4a–j in mm
against various fungal strains
Compound no. Gram positive
Gram negative
Compound no. C. albicans A. niger R. oryzae S. cerevisiae
B. subtilis S. aureus E. coli P. aeruginosa
3a
4a
3b
4b
3c
4c
3d
4d
3e
4e
3f
900
700
400
300
600
500
–
800
750
350
300
650
450
–
850
700
300
200
400
500
800
–
800
850
500
400
700
800
900
–
3a
4a
3b
4b
3c
4c
3d
4d
3e
4e
3f
–
–
–
–
–
–
–
–
300
350
700
750
–
350
300
800
800
–
400
200
–
450
400
–
600
–
800
–
–
–
600
700
450
400
800
700
900
800
400
500
700
700
650
600
500
550
700
650
800
850
700
800
900
1,000
700
700
700
600
800
700
–
700
750
600
600
750
750
900
900
500
600
700
800
–
–
–
–
–
–
–
–
–
–
–
–
4f
950
800
–
900
800
–
800
700
–
850
900
–
3g
4g
3h
4h
3i
4f
3g
4g
3h
4h
3i
–
–
–
–
950
400
450
850
800
–
800
–
–
–
–
900
–
800
–
4i
–
–
3j
4i
–
–
–
–
4j
3j
850
900
900
700
600
650
900
700
4j
Experimental
Antifungal activity
Melting points of all the compounds was recorded on
Casua-Siamia (VMP-AM) melting point apparatus and are
uncorrected. IR spectra were recorded on a Perkin-Elmer
FT-IR 240-C spectrophotometer using KBr optics. 1H
NMR spectra were recorded on Bruker AVANCE
300 MHz and INOVA 400 MHz in CDCl3 using TMS as
internal standard. NOE experiments carried out using
NOESY1D NMR. Electron impact (EI) and chemical ion-
ization mass spectra were recorded on a VG 7070 H
instrument at 70 eV. All the reaction was monitored by thin
layer chromatography (TLC) on precoated silica gel 60
The antifungal activity of compounds 3a–j and 4a–j were
screened against different fungal strains, viz., C. albicans
(MTCC 227), A. niger (MTCC 1344), R. oryzae (MTCC
262) and S. cerevisiae (MTCC 36). Compounds, 4c, 3f, 4f,
3i and 6i showed promising activity against most of the
fungi = 500 lg/ml. It is found that the inhibition is con-
centration dependent as it increases with higher concen-
tration. All other compounds were showing moderate
activity against all strains. In conclusion, 3b and 4b are the
most active among the screened compounds. The inhibitory
zone diameters of the compounds are compared with those
obtained with 100 lg/ml of standard fluconazole. The
details of minimum inhibitory concentration of each
compound have been tabulated in Table 3.
F
254(mesh); spots were visualized with UV light. Merk
silica gel (60–120 mesh) was used for column chroma-
tography. CHN analysis was recorded on a vario EL
analyzer.
Preparation of 5(6)-substituted-2-trifluoromethyl/
pentafluoroethyl-1H-benzimidazole (2a–d)
Conclusion
General procedure
A series of novel 2-trifluoromethyl-N-alkylated 6-nitro/
benzoyl benzimidazoles 3a–j and 2-trifluoromethyl-N-
alkylated 5-nitro/benzoyl benzimidazoles 4a–j were syn-
thesized, the two positional isomers are characterised and
evaluated for antimicrobial activity and promising com-
pounds 3b and 4b have been identified.
4-Substituted-o-phenylenediamine (6.5 mmol) and tri-
fluoroaceticacid/pentafluoro propionicacid (19.6 mmol) are
charged in a round-bottom flask and refluxed at 72 ꢁC
for 30–60 min. The reaction mixture was neutralized with
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Med Chem Res (2013) 22:1229–1237
aq. Ammonia, that solid precipitated, was separated by
filtration and collected. The collected solid was washed
with n-pentane and dried. The crude product was recrys-
tallised by ethylacetate–methanol mixture.
washed with n-pentane and dried. The positional isomers
were separated with long silica gel column chromatography.
1-Ethyl-6-nitro-2-trifluoromethyl-1H-benzimidazole
(3a)
5(6)-Nitro-2-trifluoromethyl-1H-benzimidazole (2a)
M.P: 177–179 ꢁC I.R: 1580 (–NO2), 1340 (–CF3).
1HNMR(CDCl3): d1.63 (t, 3H, J = 15.108); 4.50 (q, 2H,
J = 22.662); 7.95 (d, 1H, J = 9.065); 8.34 (d, 1H,
J = 9.065); 8.41 (s, 1H). Mass: 259 (M?, 100 %),
244(30 %), 229 (20 %), 201 (25 %). Anal. Calcd. for
C10H8F3N3O2: C, 46.34; H, 3.11; N, 16.21. Found: C,
46.33; H, 3.10; N, 16.21.
M.P: 108–110 ꢁC I.R: 3565.46(–NH), 1575 (–NO2), 1340
(–CF3). HNMR(CDCl3): d7.80 (d, 1H, J = 9.065); 8.29
(d, 1H, J = 9.065); 8.71 (s, 1H); 14.18 (broad s, 1H).
Mass: 231 (M?, 100 %), 201 (60 %), 69 (50 %). Anal.
Calcd. for C8H4F3N3O2: C, 41.57; H, 1.74; N, 18.18.
Found: C, 41.55; H, 1.73; N, 18.16.
1
5(6)-Nitro-2-(perfluoroethyl)-1H-benzimidazole (2b)
1-Ethyl-5-nitro-2-trifluoromethyl-1H-benzimidazole
(4a)
M.P: 80–82 ꢁC I.R: 1585 (–NO2). 1HNMR(CDCl3): d 7.91
(d, 1H, J = 9.065), 8.42 (d, 1H, J = 9.065); 8.90 (s, 1H);
10.41 (bs, 1H). Mass: 281 (M?, 100 %). Anal. Calcd. for
C9H4F5N3O2: C, 38.45; H, 1.43; N, 14.95 Found: C, 38.43;
H, 1.41; N, 14.25.
M.P: 125–127 ꢁC I.R: 1580 (–NO2), 1341 (–CF3). 1HNMR
(CDCl3): d 1.58 (t, 3H, J = 15.108); 4.49 (q, 2H,
J = 22.662); 7.52 (d, 1H, J = 9.065); 8.38 (d, 1H,
J = 9.065); 8.81 (s, 1H). Mass: 259 (M?, 100 %), 244
(20 %), 229 (30 %), 201 (25 %). Anal. Calcd. for
C10H8F3N3O2: C, 46.34; H, 3.11; N, 16.21. Found: C,
46.33; H, 3.10; N, 16.21.
Phenyl-(2-(trifluoromethyl)-3H-benzimidazole-5-
yl)methanone (2c)
1
M.P: 90–95 ꢁC I.R: 1710(–C=O), 1370 (–CF3). HNMR
1-(3-Chloropropyl)-6-nitro-2-trifluoromethyl-1H-
(CDCl3): d 7.60–7.62 (m, 2H); 7.81–7.83 (m, 4H);
7.89–7.99 (m, 1H); 8.21 (d, 1H, J = 9.065); 11.81 (broad s,
1H). Mass: 290(M?, 100 %). Anal. Calcd. for C15H9F3
N2O: C, 62.07; H, 3.13; N, 9.65. Found: C, 62.06; H, 3.12;
N, 9.64.
benzimidazole (3b)
M.P: 150–152 ꢁC I.R: 1585 (–NO2), 1340 (–CF3). 1HNMR
(CDCl3): 2.44 (dt, 2H); 3.60 (dt, 2H); 4.61 (t, 2H, J =
15.168); 8.01 (d, 1H,9.065); 8.31 (d, 1H,9.065); 8.52 (d,
1H). Mass: 308 (M?, 100 %), 310 (33 %), 252 (60 %), 254
(18 %). Anal. Calcd. for C11H9ClF3N3O2: C, 42.94; H,
2.95; N, 13.66. Found: C, 42.93; H, 2.94; N, 13.65.
Phenyl-(2-(perfluoroethyl)-3H-benzimidazole-5-yl)
methanone (2d)
1
M.P: 70–75 ꢁC I.R: 1705 (–C=O), 1390 (–CF3). HNMR
1-(3-Chloropropyl)-5-nitro-2-trifluoromethyl-1H-
(CDCl3): d 7.56–7.67 (m, 2H); 7.81 (t, 1H, J = 21.906);
7.90–7.92 (m, 4H); 8.20 (s, 1H); 12.61 (broad s, 1H). Mass:
341(M?, 100 %). Anal. Calcd. for C16H9F5N2O: C, 56.48;
H, 2.67; N, 8.23. Found: C, 56.46; H, 2.65; N, 8.22.
benzimidazole (4b)
M.P: 80–85 ꢁC I.R: 1585 (–NO2), 1350 (–CF3).
1HNMR(CDCl3): d2.41 (dt, 2H); 3.56 (dt, 2H); 4.60 (t, 2H,
J = 15.168); 7.65 (d, 1H,9.065); 8.34 (d, 1H,9.065); 8.72
(s, 1H). Mass: 308 (M?1, 100 %), 310 (33 %), 252 (60 %),
254 (18 %). Anal. Calcd. for C11H9ClF3N3O2: C, 42.94; H,
2.95; N, 13.66. Found: C, 42.93; H, 2.94; N, 13.65.
Preparation of 1-alkylated-5(6)-substituted-2-
trifluoromethyl/pentafluoroethyl-1H-benzimidazole
(3a–j and 4a–j)
General procedure
1-(4-Chlorobutyl)-6-nitro-2-trifluoromethyl-1H-
benzimidazole (3c)
5(6)-substituted-2-trifluoromethyl/pentafluoroethyl-1H-ben-
zimidazole(4.3 mmol), alkylhalide (5.2 mmol), potassium
carbonate (8.6 mmol) and DMF (5 ml) was charged into a
round-bottomed flask and stirred at room temperature for
3–4 h/on sonicator for 15 min. Then the reaction mixture was
poured in ice cold water, separated solid was filtered and
M.P: 140–142 ꢁC I.R: 1590 (–NO2), 1370 (–CF3). 1HNMR
(CDCl3): d 2.09 (t, 4H); 3.56 (dt, 2H, J = 12.086); 4.45
(dt, 2H, J = 15.108); 8.03 (d, 1H, J = 9.08); 8.34 (d, 1H,
J = 9.08); 8.52 (d, 1H). Mass: 321 (M?, 100 %), 323
(33 %), 244 (60 %), 198 (18 %). Anal. Calcd. for
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Med Chem Res (2013) 22:1229–1237
1235
Ethyl 2-(6-nitro-2-trifluoromethyl)-1H-benzimidazole-
C12H11ClF3N3O2: C, 44.80; H, 3.45; N, 13.66. Found: C,
44.68; H, 3.44; N, 13.65.
1-yl)acetate (3f)
M.P: 120–123 ꢁC I.R: 1745.80 (–COO–), 1580 (–NO2),
1375 (–CF3). 1HNMR(CDCl3): d 1.28 (t, 3H, 30.215); 4.37
(q, 2H, J = 21.096); 5.21 (s, 2H); 8.02 (d, 1H, J = 9.065);
8.40 (d, 1H, J = 9.065)8.40 (s, 1H) Mass: 318 (M?, 20 %),
290 (90 %), 244(100 %). Anal. Calcd. for C12H10F3N3O4:
C, 45.43; H, 3.18; N, 13.25. Found: C, 45.42; H, 3.19; N,
13.26.
1-(4-Chlorobutyl)-6-nitro-2-trifluoromethyl-1H-
benzimidazole (4c)
1
M.P: 70–74 ꢁC I.R: 1590 (–NO2), 1340 (–CF3). HNMR
(CDCl3): d 2.11–2.23 (m, 4H); 3.51 (t, 2H, J = 12.086);
4.45 (t, 2H, J = 15.108); 7.60 (d, 1H, J = 9.08); 8.34 (d,
1H, J = 9.08); 8.78 (s, 1H). Mass: 321 (M?, 35 %), 323
(12 %), 244 (65 %), 198 (30 %). Anal. Calcd. for
C12H11ClF3N3O2: C, 44.80; H, 3.45; N, 13.66. Found: C,
44.68; H, 3.44; N, 13.65.
Ethyl 2-(5-nitro-2-trifluoromethyl-1H-benzimidazole-
1-yl)acetate (4f)
M.P: 75–80 ꢁC I.R: 1737.54 (–COO–), 1585 (–NO2), 1370
(–CF3). HNMR(CDCl3): d 1.30 (t, 3H, 30.215); 4.31 (q,
2-(6-Nitro-2-trifluoromethyl-1H-benzimidazol-1-
1
yl)acetonitrile (3d)
2H, J = 21.096); 5.10 (s, 2H); 7.48 (d, 1H, J = 9.065);
8.39 (d, 1H, J = 9.065); 8.80 (s, 1H) Mass: 318 (M?,
25 %), 290 (90 %), 244 (100 %). Anal. Calcd. for
C12H10F3N3O4: C, 45.43; H, 3.18; N, 13.25. Found: C,
45.41; H, 3.19; N, 13.26.
M.P: 145–150 ꢁC I.R: 2210.68 (–CN), 1340 (–CF3).
1HNMR(CDCl3): d 5.30 (s, 2H); 8.09 (d, 1H, J = 9.065);
8.40 (d, 1H, J = 9.065) 8.52 (s, 1H). Mass: 270 (M?,
100 %), 240 (78 %), 69 (58 %). Anal. Calcd. for
C10H5F3N4O2: C, 44.46; H, 1.87; N, 20.74. Found: C,
44.44; H, 1.86; N, 20.75.
1-(4-Chlorobutyl)-6-nitro-2-(perfluoroethyl)-1H-
benzimidazole (3g)
2-(5-Nitro-2-trifluoromethyl-1H-benz]imidazole-1-
M.P: 85–89 ꢁC I.R: 1580 (–NO2), 1385 (–CF3).
1HNMR(CDCl3): d 1.94–2.07 (m, 4H); 3.58 (t, 2H,
J = 12.086); 4.49 (t, 2H, J = 15.108); 7.92 (d, 1H,
J = 9.065); 8.25 (d, 1H, J = 9.065); 8.40 (s, 1H). Mass:
372 (M?, 100 %), 374 (33 %). Anal. Calcd. for
C13H11ClF5N3O2: C, 42.01; H, 2.98; N, 11.31. Found: C,
42.02; H, 2.98; N, 11.30.
yl)acetonitrile (4d)
1
M.P: 93–95 ꢁC I.R: 2210 (–CN), 1345 (–CF3). HNMR
(CDCl3): d 5.30 (s, 2H); 7.71 (d, 1H, J = 9.357); 8.50 (d,
1H, J = 9.357) 8.90 (s, 1H). Mass: 270 (M?, 40 %), 240
(50 %), 69 (100 %). Anal. Calcd. for C10H5F3N4O2: C,
44.46; H, 1.87; N, 20.74. Found: C, 44.43; H, 1.85; N,
20.75.
1-(4-Chlrobutyl)-5-nitro-2-(perfluoroethyl)-1H-
benzimidazole (4g)
6-Nitro-1-(prop-2ynyl)-2-trifluoromethyl)-1H-
benzimidazole (3e)
1
M.P: 80–82 ꢁC I.R: 1590 (–NO2), 1380 (–CF3). HNMR
(CDCl3): d 2.00–2.10 (m, 4H); 3.61 (t, 2H, J = 12.086);
4.52 (t, 2H, J = 15.108); 7.60 (d, 1H, J = 9.065); 8.31 (d,
1H, J = 9.065); 8.80 (s, 1H). Mass: 372 (M?, 100 %), 374
(33 %). Anal. Calcd. for C13H11ClF5N3O2: C, 42.01; H,
2.98; N, 11.31. Found: C, 42.02; H, 2.99; N, 11.30.
M.P: 120–125 ꢁC I.R: 2100 (–C:C–), 1580 (–NO2), 1345
(–CF3). HNMR(CDCl3): d 0.89 (t, 1H, 8.309); 5.89 (d,
2H, J = 6.798); 7.98 (d, 1H, J = 9.065); 8.30 (d, 1H,
J = 9.065); 8.71 (s, 1H). Mass: 270 (M?, 100 %). Anal.
Calcd. for C11H6F3N3O2: C, 49.08; H, 2.26; N, 15.02.
Found: C, 49.06; H, 2.25; N, 15.01.
1
Ethyl-2-(6-nitro-2-(perfluoroethyl)-1H-benzimidazole-
1-yl)acetate (3h)
5-Nitro-1-(prop-2-ynyl)-2-trifluoromethyl-1H-
benzimidazole (4e)
M.P: 124–126 ꢁC I.R: 1754.45 (–COO–), 1590 (–NO2),
1385 (–CF3). 1HNMR(CDCl3): d 1.40 (t, 3H, J = 30.215);
4.31(q, 2H, J = 21.906); 5.20 (s, 2H); 8.09 (d, 1H,
J = 9.065); 8.41 (d, 1H, J = 9.065); 8.41 (s, 1H). Mass:
368 (M?, 100 %). Anal. Calcd. for C13H10F5N3O4: C,
42.52; H, 2.74; N, 11.44. Found: C, 42.51; H, 2.74; N,
11.45.
M.P: 75–80 ꢁC I.R: 2100 (–C:C–), 1580 (–NO2), 1345
(–CF3). HNMR(CDCl3): d 0.83 (t, 1H, 7.554); 5.75 (d,
2H, J = 6.798); 7.78 (d, 1H, J = 9.065); 8.30 (d, 1H,
J = 9.065); 8.72 (s, 1H). Mass: 270 (M?, 100 %). Anal.
Calcd. for C11H6F3N3O2: C, 49.08; H, 2.25; N, 15.01.
Found: C, 49.06; H, 2.26; N, 15.02.
1
123

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Med Chem Res (2013) 22:1229–1237
Ethyl-2-(5-nitro-2-(perfluoroethyl)-1H-benzimidazole-
Antibacterial activity
1-yl)acetate (4h)
Procedure Antibacterial activity of all the test compounds
was assayed systematically by plate method (National
Committee for Clinical Laboratory) against for non-patho-
genic strains of gram positive microorganisms such as
B. subtilis (NCIM 2921), S. aureus (NCIM 2079) and gram
negative bacteria E. coli (NCIM 2068), P. aeruginosa
(NCIM 2957) at different concentrations using DMSO as
solvent. The bacteria were sub-cultured on Mueller–Hinton
Agar medium. The test compound (standard concentrations
50, 100, 150, 200, 250, 300…1,000 lg/ml) was added to the
petri dishes (measuring 90 mm each side) containing 25-ml
nutrient agar medium, then standard inoculum. The test
compounds were introduced into the nutrient agar medium
by decreasing concentrations from 1 mg/ml. The plates were
incubated at 37 ꢁC for 18–20 h. The antibacterial activity
was evaluated by measuring the zone of inhibition sur-
rounding the discs using Vernier Calipers. Standard con-
centrations of ciprofloxacin (50 lg) served as positive
antibacterial control. Solvent control was also maintained
under similar conditions. All the tests were repeated three
times to minimize the error.
M.P: 90–92 ꢁC I.R: 1740.84 (–COO–), 1585 (–NO2), 1390
(–CF3). HNMR(CDCl3): d 1.31 (t, 3H, J = 30.215); 4.29
(q, 2H, J = 21.906); 5.11 (s, 2H); 7.45 (d, 1H, J = 9.065);
8.44 (d, 1H, J = 9.065); 8.80 (s, 1H). Mass: 368 (M?,
100 %). Anal. Calcd. for C13H10F5N3O4: C, 42.52; H, 2.74;
N, 11.44. Found C, 42.50; H, 2.73; N, 11.45.
1
Ethyl-2-(6-benzoyl-2-(trifluoromethyl)-1H-
benzo[d]imidazole-1-yl)acetate (3i)
M.P: 120–122 ꢁC I.R: 1752.64 (–COO–), 1705 (–CO–),
1370 (–CF3). 1HNMR(CDCl3): d 1.28 (t, 3H, J = 30.215);
4.22 (q, 2H, J = 21.096); 5.10 (s, 2H); 7.51 (t, 2H) 7.59 (d,
1H, J = 9.065); 7.70–7.82 (m, 3H); 7.92 (s, 1H); 7.99 (d,
1H, J = 9.065). Mass: 377 (M?, 100 %). Anal. Calcd. for
C19H15F3N2O3: C, 60.64; H, 4.02; N, 7.44. Found: C,
60.65; H, 4.02; N, 7.42.
Ethyl-2-(5-benzoyl-2-trifluoromethyl)-1H-
benzimidazole-1-yl)acetate (4i)
Antifungal activity
M.P: 90–92 ꢁC I.R: 1757.44 (–COO–), 1710 (–CO–), 1370
1
(–CF3). HNMR(CDCl3): d 1.26 (t, 3H, J = 30.215); 4.28
Procedure The test compounds were screened for anti-
fungal activity using paper disc method was performed
using Sabouraud dextrose broth. The organisms used are
C. albicans (MTCC 227), A. niger (MTCC 1344), R. ory-
zae (MTCC 262) and S. cerevisiae (MTCC 36). The
medium was poured into sterile petri dishes under aseptic
conditions in a laminar flow chamber. When the medium in
the plates solidified, 0.5 mL of the 24 h liquid cultures
containing 107 microorganisms/mL was inoculated. Filter
paper discs (5-mm diameter) saturated with solutions of
each test compound (concentrations: 50, 100, 150, 200,
250, 300…1,000 lg/mL were prepared using DMSO) were
placed on the indicated agar mediums. The incubation time
was 24 h at 37 ꢁC. Results were compared with control
(DMSO) and the standard i.e. fluconazole apart from the
test compounds. Inhibitory activity was measured (in mm)
as the diameter of the observed inhibition zones. All the
tests were repeated three times to minimize the error.
(q, 2H, J = 21.096); 5.03 (s, 2H); 7.40–7.51 (m, 3H) 7.56
(t, 1H); 7.78 (d, 1H, J = 9.065); 8.01 (d, 1H, J = 9.065);
8.28 (s, 1H). Mass: 377 (M?, 100 %). Anal. Calcd. for
C19H15F3N2O3: C, 60.64; H, 4.02; N, 7.44. Found: C,
60.65; H, 4.02; N, 7.43.
Ethyl-2-(6-benzoyl)-2-(perfluoroethyl)-1H-
benzimidazole-1-yl)acetate (3j)
M.P: 75–80 ꢁC I.R: 1749.95 (–COO–), 1370 (–CF3).
1HNMR(CDCl3): d 1.31 (t, 3H, J = 30.215); 4.23 (q, 2H,
J = 21.096); 5.09 (s, 2H); 7.35–7.47 (m, 3H) 7.70–7.83
(m, 3H); 7.85–7.96 (m, 2H). Mass: 427 (M?, 100 %). Anal.
Calcd. for C20H15F5N2O3: C, 56.34; H, 3.55; N, 6.57.
Found: C, 56.33; H, 3.56; N, 6.56.
Ethyl-2-(5-benzoyl)-2-(perfluoroethyl)-1H-
benzimidazole-1-yl)acetate (4j)
M.P: 85–88 ꢁC I.R: 1757.37 (–COO–), 1370 (–CF3).
1HNMR(CDCl3): d 1.26 (t, 3H, J = 30.215); 4.23 (q, 2H,
J = 21.096); 5.02 (s, 2H); 7.45–7.52 (m, 4H) 7.71 (d,
1H, J = 9.065); 8.02 (d, 1H, J = 9.065); 8.30 (s, 1H).
Mass: 427 (M?, 100 %). Anal. Calcd. for C20H15F5N2O3:
C, 56.34; H, 3.55; N, 6.57. Found: C, 56.32; H, 3.56; N,
6.56.
X-ray crystallography data
Crystal data (AE88) (3a)
C10H8F3N3O2, M = 259.19, triclinic, space group P1,
˚
˚
˚
a = 6.7539(11) A, b = 8.0267(13) A, c = 10.3766(16) A,
a = 77.694(2) ꢁ, b = 87.340(2) ꢁ, c = 79.188(2) ꢁ,
123

Med Chem Res (2013) 22:1229–1237
1237
3
V = 539.84(15) A ,
˚
Denny WA, Rewcastle GW, Bagulaey BC (1990) Potential antitumor
agents. 59. Structure–activity relationships for 2-phenylbenzimida-
zole-4-carboxamides, anew class of minimal-interacting agents
which may not act via topoisomerase-II. J Med Chem 33:814–819
Farukh A, Bhawana M, Shamin (2005) Synthesis, antibacterial,
antifungal activity & interaction of CT-DNA with a new
benzimidazole derived Cu(II) complex. Eur J Med Chem
40:1103
Fransisco HL, Alicia HC, Rafeal C, Gabriel NV, Olivia SA, Manuel
H, Libian GM (2010) Synthesis and biological activity of
2-(trifluoromethyl)-1H-benzimidazole derivatives against some
protozoa and Trichinella spiralis. Eur J Med Chem 45:3135
Gabriel NV, Roberto C, Aliica HC, Cilian G, Fransisco KL, Suan V,
Raual M, Rafeal C, Manuel H, Rafel C (2001) Synthesis and
antiparasitic activity of 2-(trifluoromethyl)-1H-benzimidazole
derivatives. Bioorg Med Chem Lett 11:1879
Z = 2,
Dcalcd = 1.595 mg m-3
,
T = 294(2) K, l = 0.148 mm-1, F(000) = 264, k =
˚
0.71073 A. Data collection yielded 5145 reflections result-
ing in 1892 unique, averaged reflection, 1725 with I [2r(I).
Full-matrix least-squares refinement led to a final R =
0.0594, wR = 0.1659 and GOF = 1.059. CCDC 828078
contains supplementary Crystallographic data for the
structure.
Crystal data (AE87)(4a)
C10H8F3N3O2, M = 259.19, monoclinic, space group P21/n,
˚
a = 12.152(3) A, b = 7.8740(19) A, c = 12.376(3) A,
˚
˚
3
Gabriel NV, de Maria MRV, Cilian G, Victor M, Lucia G, Alicia H,
Rafeal C, Fransisco HL (2006) Synthesis and antiprotozoal
activity of some 2-(trifluoromethyl)-1H-benzimidazole bioisos-
teres. Eur J Med Chem 41:135
˚
b = 113.387(4) ꢁ, V = 1086.9(5) A , Z = 4, Dcalcd
=
1.584 mg m-3, T = 294(2) K, l = 0.147 mm-1, F(000) =
˚
528, k = 0.71073 A. Data collection yielded 10099 reflec-
Juan V, Roberto C, Alicia HC, Lilian G, Fransisco HL, Gabriel NV,
Amparo T, Rafel C, Manuel H, Rafel C (2002) Synthesis and
antiparasitic activity of 1H-benzimidazole derivatives. Bioorg
Med Chem Lett 12:2221
Keith WAM, Allen CB, Sharon H, Richard EJ, Thomas RM (1978)
1-[4-(4-Chloro phenyl)-2-(2,6-di chloro phenyl thio)-n-butyl]-
1H-imidazole nitrate a new point antifungal agent. J Med Chem
21:840
tions resulting in 1916 unique, averaged reflection, 1664
with I [ 2r(I). Full-matrix least-squares refinement led to a
final R = 0.0455, wR = 0.1190 and GOF = 1.054. Inten-
sity data were measured on Bruker Smart Apex with CCD
area detector. CCDC 828077 contains supplementary crys-
tallographic data for the structure.
Kurumurthy C, Sambasiva Rao P, Veeraswamy B, Santosh Kumar G,
ShanthanRao P, Narsaiah B, Velatooru LR, Pamanji R, Venk-
ateswara Rao J (2011) Synthesis of novel alkyltriazole tagged
pyrido[2,3-d]pyrimidine derivatives and their anticancer activity.
Eur J Med Chem 46:3462–3468
Acknowledgments Authors are thankful to Dr. J. S. Yadav,
Director, IICT for his constant encouragement and also authors are
thankful to Council of Scientific and Industrial Research (CSIR),
India for the financial assistance in the form of SRF fellowship.
Maxwell WA, Brody G (1971) Antifungal activity of selected
benzimidazole compounds. Appl Microbiol 21:944
Nakamura Shoshiro (2005) Structure of azomycin, a new antibiotic.
Chem Pharm Bull 3:379
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