Design, synthesis, and PASS-assisted evaluation of novel 2-substituted benzimidazole derivatives as potent anthelmintics

Article abstract of DOI:10.1007/s00044-013-0856-1

A series of novel 2-substituted benzimidazole analogs has been designed and synthesized by connecting the benzimidazole nucleus with variedly substituted chalcone moieties through an amino linker. The designed analogs were predicted for their biological activity profile through the computer software PASS. The compounds were predicted to have potent anthelmintic activity. These were synthesized and the activity of each compound was evaluated experimentally at the concentrations of 0.1, 0.2, and 0.5% in terms of mortality time and paralysis time for the helminthes. The experimentally observed activity was found to comply with the PASS predicted activity. All compounds showed dose-dependent activities. The compounds with an electron releasing group at the para position on phenyl ring in the chalcone moiety (8 and 9) were the most active in comparison to those bearing electron withdrawing groups. The corresponding ortho analogs (4 and 5) also revealed good paralytic and lethal activities. The higher activities of 8 and 9 may be attributed to the favorable electronic interactions of the electron releasing groups present at para position of the phenyl ring. Comparative analysis of the Lipinski's parameters and the activities of the compounds revealed all the compounds to comply with the Lipinski's rule of five. Further an optimum hydrophilicity and total polar surface area in the range of 65-80 of the molecule are required for the potent activity, but Molar refractance is not found to have any significant role in determining the anthelmintic activity. Springer Science+Business Media 2013.

Full text of DOI:10.1007/s00044-013-0856-1

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2014) 23:2690–2697
DOI 10.1007/s00044-013-0856-1
ORIGINAL RESEARCH
Design, synthesis, and PASS-assisted evaluation of novel
2-substituted benzimidazole derivatives as potent anthelmintics
•
•
•
Kamalpriya Garg Yogita Bansal Gulshan Bansal
R. K. Goel
Received: 29 March 2013 / Accepted: 17 October 2013 / Published online: 31 October 2013
ꢀ Springer Science+Business Media New York 2013
Abstract A series of novel 2-substituted benzimidazole
analogs has been designed and synthesized by connecting
the benzimidazole nucleus with variedly substituted chal-
cone moieties through an amino linker. The designed
analogs were predicted for their biological activity profile
through the computer software PASS. The compounds
were predicted to have potent anthelmintic activity. These
were synthesized and the activity of each compound was
evaluated experimentally at the concentrations of 0.1, 0.2,
and 0.5 % in terms of mortality time and paralysis time for
the helminthes. The experimentally observed activity was
found to comply with the PASS predicted activity. All
compounds showed dose-dependent activities. The com-
pounds with an electron releasing group at the para posi-
tion on phenyl ring in the chalcone moiety (8 and 9) were
the most active in comparison to those bearing electron
withdrawing groups. The corresponding ortho analogs (4
and 5) also revealed good paralytic and lethal activities.
The higher activities of 8 and 9 may be attributed to the
favorable electronic interactions of the electron releasing
groups present at para position of the phenyl ring. Com-
parative analysis of the Lipinski’s parameters and the
activities of the compounds revealed all the compounds to
comply with the Lipinski’s rule of five. Further an optimum
hydrophilicity and total polar surface area in the range of
65–80 of the molecule are required for the potent activity,
but Molar refractance is not found to have any significant
role in determining the anthelmintic activity.
Keywords Benzimidazole ꢀ Chalcones ꢀ Lipinski ꢀ
Pharmacophore ꢀ Helminthes ꢀ PASS
Introduction
Benzimidazoles nucleus has received worldwide attention
for the development of new molecules of therapeutic
interest due to its presence in numerous bioactive mole-
cules. Many drugs derived from benzimidazole belonging
to different categories of pharmacological activities include
omeprazole, enviradine, candesartan, bendamustine,
astemizole, albendazole, mebendazole, thiabendazole, and
fenbendazole (Bansal and Silakari, 2012). An elaborate
survey of the literature has revealed the importance of
2-position of benzimidazole for a majority of the thera-
peutic activities and a substituted amino group is proved to
be the most versatile substituent at this position (Navrocka,
1996; Mor et al., 2004; Caroti et al., 1989; Akul et al.,
2011; Nawrocka et al., 2004). Chalcones (1,3-diaryl-2-
propen-1-ones) are open chain flavonoids, distributed
widely in fruits, vegetables, spices, tea, and soy-based
foodstuff. These also possess many useful pharmacological
activities such as anti-inflammatory, antimicrobial, anti-
fungal, antioxidant, anthelmintic, antitumor, and anticancer
(Settimo et al., 1991; Zdzislawa, 2007; Di Carlo et al.,
1999). These activities are attributed to the a,b-unsaturated
keto group owing to its propensity to be attacked by the
nucleophilic groups present in proteins. Coupling of
benzimidazole and chalcone moieties has produced com-
pounds (Fig. 1) exhibiting antitumor, anti-depressant,
antimicrobial activities (Dimmock et al., 2005; Mishra
et al., 2001; Shaharyar et al., 2010; Babu and Selvakumar,
2012; Mathew et al., 2012). Hence, based upon the che-
motherapeutic importance of 2-aminobenzimidazoles and
K. Garg ꢀ Y. Bansal (&) ꢀ G. Bansal ꢀ R. K. Goel
Department of Pharmaceutical Sciences and Drug Research,
Punjabi University, Patiala 147 002, India
e-mail: yogitabansalp@rediffmail.com
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Fig. 1 Benzimidazole–chalcones as important therapeutic agents
a,b-unsaturated keto group of chalcone, novel compounds
have been designed by coupling the two vital moieties
through an amino linker in the present study (Fig. 1). The
three components of the designed compounds, i.e., benz-
imidazole nucleus, guanidine, and chalcone moiety are
responsible for varied activities and hence these are
expected to exhibit wide spectra of pharmacological
activities. The compounds were synthesized and predicted
for their pharmacological activities through a computer-
aided program, prediction of activity spectrum of sub-
stances (PASS). The most probable activity predicted for
the synthesized compounds was evaluated experimentally.
ascertained by appearance of Amide-I bands of medium
intensity in the range of 1,670–1,700 cm-1 in IR spectra
and confirmed by accurate mass of quasimolecular ion
(M?H?) in the ?ESI-TOF mass spectrum of each com-
pound. Finally, elementary composition of each compound
was confirmed through calculation of molecular formula
corresponding to mass of its (M?H)? ion using elemental
composition calculator. The calculated molecular formula
of (M?H)? ion of each compound was found to be same as
the actual one.
Predicted biological activities of benzimidazole
derivatives by PASS
Results and discussion
PASS predicts the pharmacological effects and different
side effects of a molecule based on the analysis of struc-
ture–activity relationships for the training set of database
currently including more than 46,000 drugs, drug candi-
dates, and lead compounds having biological activities
determined experimentally. Two probabilities are calcu-
lated for each activity, i.e., probability of compound being
active (Pa) and probability of compound being inactive
(Pi). Being probabilities, the Pa and Pi values vary from
0.000 to 1.000 and in general, Pa ? Pi \ 1, since these
probabilities are calculated independently. The PASS pre-
dictions can be interpreted and used in a flexible manner:
(i) only activities with Pa [ Pi are considered as possible
for a particular compound; (ii) If Pa [ 0.7, the chance to
find the activity experimentally is high; (iii) If
0.5 \ Pa \ 0.7, the chance to find the activity experi-
mentally is less, but the compound is probably not so
similar to known pharmaceutical agents; and (iv) If
Pa \ 0.5, the chance to find the activity experimentally is
even less, but the chance to find a structurally new
Chemistry
A series of benzimidazole–chalcone conjugates was
designed and synthesized based on the importance of
2-aminobenzimidazole and chalcones toward varied bio-
logical activities. The target compounds were synthesized
through a three step process (Fig. 2). In the first step,
2-aminobenzimidazole (1) was prepared from o-phenyl-
enediamine and cyanogen bromide as reported by Leonard
et al. (1947). In the second step, the 1 was acetylated using
acetic anhydride by stirring at 5–10 ꢁC in chloroform to
form 2-acetamidobenzimidazole (2). Finally, 2 was cou-
pled with variedly substituted benzaldehydes to produce
the target compounds (3–11) through conventional reflux-
ing as well as microwave-assisted method (Wang et al.,
2007). Structures of the target compounds were confirmed
1
through IR, H NMR, 13C NMR, and high resolution mass
spectral (HRMS) data. In general, the structure was
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Med Chem Res (2014) 23:2690–2697
Fig. 2 Synthetic scheme for 2-substituted benzimidazole derivatives. (i) CNBr, methanol, stirring, 24 h, NH4OH; (ii) acetic anhydride, CHCl3,
stirring, 5–10 ꢁC; (iii) ArCHO, KOH, microwave (2 min, 170 W)/conventional (reflux, 5 h)
Table 1 PASS predicted activity spectrum of the target compounds
Compound
Anthelmintic activity
Antineoplastic activity
Leukotriene antagonist
Cytokinine modulator
Pa
Pi
Pa
Pi
Pa
Pi
Pa
Pi
3
0.818
0.833
0.836
0.698
–
0.003
0.023
0.003
0.005
–
0.792
0.809
0.679
0.703
0.531
0.640
0.679
0.587
0.686
0.015
0.011
0.042
0.031
0.029
0.050
0.042
0.074
0.021
0.692
0.726
0.746
–
0.04
0.029
0.025
–
0.591
0.049
4
0.556
0.063
5
–
–
6
–
–
7
–
–
–
–
8
0.850
0.836
0.776
–
0.003
0.003
0.018
–
0.754
0.75
–
0.022
0.021
–
–
–
9
0.636
0.032
10
11
–
–
–
–
–
–
compound, i.e., novel chemical entity (NCE ) increases
(Poroikov et al., 2000; Marawaha et al., 2007). Recently,
an internet version of PASS has been made available at the
PASS developer’s web site. The accuracy of prediction is
reported to be as high as 85 %. The predicted activity
spectrum of the synthesized compounds (Table 1) revealed
that most of the compounds possess high probability of
showing anthelmintic and antineoplastic activities. The
compounds bearing no or electron releasing groups
(hydroxyl and methoxy) on the chalconic phenyl ring were
predicted to have higher anthelmintic activity than the
compounds bearing electron withdrawing groups (chloro
and nitro).
dependent paralytic as well as lethal activities and the
maximum anthelmintic activity was observed at 0.5 % of
the test compounds (Table 2; Figs. 3, 4). The activity was
in concordance with their PASS predicted scores that
support the applicability of the PASS in reliable prediction
of the activity spectrum of the compounds. The 8 and 9 (p-
hydroxyl and p-methoxy derivatives) were the most active
compounds from the series and slightly less active than
albendazole at all concentrations. The corresponding ortho
analogs (4 and 5) also revealed good paralytic and cidal
activities. On the contrary, the chloro and nitro analogs in
the series (6, 7, 10, and 11) were found to be the least
active of all even at 0.5 % concentration. It revealed that
the higher activities of 8 and 9 may be attributed to the
favorable electronic interactions of the electron releasing
groups present at para position of the phenyl ring. Com-
parative analysis of the anthelmintics activities of the
compounds and the Lipinski’s parameters for a bioactive
drug (Tables 2, 3, respectively) has revealed that all cal-
culated parameters comply with the Lipinski’s rule of five
Anthelmintic activity
The earthworms are used to study the anthelmintic activity
because of their resemblance, both anatomically and
physiologically, with the intestinal roundworm parasites in
human beings. All test compounds exhibited concentration-
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Table 2 Anthelmintic activity of the compounds at concentrations of 0.1, 0.2, and 0.5 % w/v
Compound
Time of paralysis (min)
Time of death (min)
0.1
0.1
0.2
0.5
0.2
0.5
Albz.
8.8 0.3
17.3 0.1
14.1 0.2
13.8 0.1
14.4 0.1
13.7 0.2
11.3 0.1
10.5 0.1
13.1 0.2
14.3 0.1
[25
7.6 0.2
14.2 0.1
10.8 0.2
11.8 0.1
12.3 0.1
12.8 0.1
10.7 0.1
9.2 0.1
12.2 0.2
11.1 0.1
[25
6.1 0.3
11.3 0.1
8.9 0.2
9.5 0.1
10.3 0.1
12.3 0.1
8.5 0.1
8.7 0.1
11.7 0.2
10.5 0.1
[25
12.7 0.3
18.9 0.1
16.3 0.1
14.6 0.1
16.3 0.1
16.4 0.1
12.8 0.2
12.0 0.2
15.5 0.2
16.1 0.2
[25
9.8 0.4
15.9 0.2
12.9 0.3
12.4 0.1
14.4 0.1
14.9 0.2
11.9 0.2
11.7 0.2
14.6 0.2
13.3 0.1
[25
7.0 0.3
13.2 0.1
10.1 0.1
10.3 0.1
11.2 0.1
14.3 0.1
10.2 0.1
10.7 0.1
14.3 0.1
12.3 0.1
[25
3
4
5
6
7
8
9
10
11
Control
Fig. 3 Time of paralysis of
compounds 3–11. Each value is
expressed as mean SD
(n = 6). Statistical significant
difference was obtained at
p \ 0.05, a, b, and c as
compared to Albendazole
(Albz.) at the respective
concentration
20
18
16
14
12
10
8
0.10%
0.20%
0.50%
a
a
a
b
a
a
a
b
b
b
c
c
a
b
c
b
b
a
c
b
c
c
6
4
2
0
Albz.
3
4
5
6
7
8
9
10
11
Target compounds
Fig. 4 Time of death of
compounds 3–11. Each value is
expressed as mean SD
(n = 6). Statistical significant
difference was obtained at
p \ 0.05, a, b, and c as
compared to Albendazole
(Albz.) at the respective
concentration
20
18
16
14
12
10
8
a
0.10%
0.20%
0.50%
a
a
a
a
b
a
b
b
c
c
b
c
c
b
c
a
a
b
b
c
c
c
c
6
4
2
0
Albz.
3
4
5
6
7
8
9
10
11
Target compounds
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Table 3 TPSA, molar refractance, and calculated Lipinski’s parameters for the test compounds
Compound
TPSAa
MRb
Log Pc
MWd
nONe
nOHNHf
nviolationsg
nrotbh
3
57.78
78.01
67.02
103.60
57.78
78.01
67.02
103.60
57.78
77.94
79.63
84.40
NDi
3.28
3.04
3.11
3.01
3.73
2.80
3.34
3.24
3.96
263.30
279.30
293.33
308.30
297.75
279.30
293.33
308.30
297.75
4
5
5
7
4
5
5
7
4
2
3
2
2
2
3
2
2
2
0
0
0
0
0
0
0
0
0
3
3
4
4
3
3
4
4
3
4
5
6
7
82.75
79.63
84.40
NDi
8
9
10
11
a
82.75
Total polar surface area (hydrophilicity)
Molar refractance (cm3/mol)
Calculated lipophilicity
b
c
d
e
f
Molecular weight
Number of hydrogen bond acceptor
Number of hydrogen bond donors
g
h
i
Number of violations from Lipinski’s rule of five
Number of rotatable bonds
Not determined/calculated
(Lipinski et al., 2001), i.e., a molecular weight of 500 or
less, a log P not more than 5, number of hydrogen bond
donor sites not exceeding five, and number of hydrogen
bond acceptor sites not exceeding ten. However, a wide
variability in the total polar surface area (TPSA), molar
refractance (MR, a measure of steric factor), and log
P values of the compounds have indicated that though an
optimum hydrophilicity and a TPSA in the range of 65–80
of the molecule may be required for the potent activity but
MR does not play any significant role in determining the
anthelmintic activity.
Materials and methods
The reactions requiring anhydrous conditions were con-
ducted in flame-dried apparatus. Melting points were
recorded on electronic melting point apparatus (Perfit,
India). The microwave-assisted reactions were carried out
in a microwave oven (IFB23SCI, 850 W). IR spectra were
recorded on IR Spectrophotometer (Perkin Elmer Fx) as
1
KBr disks. H NMR and 13C NMR spectra were recorded
on Bruker Avance II NMR spectrometer (400 MHz), using
CDCl3 as solvent and tetramethylsilane (TMS) as internal
1
standard. In H NMR, chemical shifts were reported in d
values with number of protons, multiplicities (s—singlet,
m—multiplet, dd—double doublet, br—broad), and cou-
pling constants (J) in Hertz (Hz). The mass spectra were
recorded on Bruker Daltonics microTOF instrument using
electrospray ionization in positive and/or negative polarity
mode. Each compound was purified by crystallization and
purity of all synthesized compounds was ascertained by
TLC using pre-coated Silica gel G plates (Merck, Mumbai,
India) using chloroform:methanol (96:4) as developing
solvent system. The dried developed plates were visualized
in UV chamber at both short and long wavelengths as well
as by exposure to iodine vapors in a tightly closed cham-
ber. Earth worms (Pheretima posthuma) of nearly equal
size (6 1 cm, in expanded form) were procured from the
vermicompost (Jagatpur, Chandigarh, India) and were
selected randomly for the present study. The species of
earthworms was authenticated from the Department of
Zoology, Punjabi University, Patiala (India).
Conclusion
The ABC series of benzimidazole derivatives bearing
variedly substituted chalcone moieties have been synthe-
sized and evaluated through PASS software for predicting
the activity spectrum of the compound. All compounds
were predicted to have potent anthelmintic activity which
was found to comply with the activity observed experi-
mentally. The SAR designed from the in silico as well as
experimental study reveals that the electron releasing
groups increases the anthelmintic activity. On the contrary,
an electron withdrawing group at both ortho as well as
para position of the phenyl ring incurs lesser activity.
Further, the activity is higher for the analog bearing the
group at para position on phenyl ring in the chalcone
moiety. All compounds have been found to comply with
the Lipinski’s rule of five for a bioactive drug.
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Chemistry
2695
mixture were poured into crushed ice and acidified with
dilute HCl. The solid separated was filtered and recrystal-
lised from ethanol to obtain the pure product.
2-Aminobenzimidazole (1)
2-[3-Phenylacrylamido]-benzimidazole (3) Yield: 62 %
(Method A), 56 % (Method B); m.p. 118–120 ꢁC; IR:
3500–3400 (N–H str.), 3048 (C–H, Ar, str.), 2880 (C–H,
Ali, str.), 1685 (C=O str.), 1642, 1453 (C….C str.), 922 (Ar
C–H bending); 1H NMR (400 MHz, CDCl3): d 7.9–7.8 (br
s, 1H, NH), 7.5–7.4 (m, 2H, ArH), 7.3–7.1 (m, 8H, ArH,
CH=CH), 6.9–6.8 (m, 1H, CH=CH), 5.0 (br s, 1H, NH).
13C NMR (400 MHz, CDCl3): d 117.4 (C-2, C-5), 121.2
(C-8), 124.3 (C-3, C-4), 127.6 (C-11, C-15), 129.3 (C-13),
130.2 (C-12, C-14), 136.1 (C-10), 138.5 (C-1, C-6), 142.4
(C-7), 143.5 (C-9), 166.2 (C=O). HRMS (?ESI): m/
z 264.1151 (C16H14N3O?).
A mixture of 3.2 g (30 mM) of cyanogen bromide and
2.1 g (20 mM) of o-phenylenediamine in aqueous metha-
nol was stirred at room temperature for 24 h. Thereafter,
the solution was heated on water bath to remove the
methanol. The solution was cooled and basified with
ammonia. The crude precipitates obtained were recrystal-
lized from ethanol–water. Yield 88 %; m.p. 234 ꢁC; IR:
3500–3400 (N–H str.), 3116 (C–H, Ar, str.), 1636 (C=N
str.), 1483 (N–H scissoring), 1451 (C=C str.), 1239 (C–N
1
str.), 846 (N–H wagging), 719 (Ar–H bending); H NMR
(400 MHz, CDCl3): d 7.2 (dd, 2H, J = 5.8, 3 Hz, ArH),
6.9 (dd, 2H, J = 5.8, 3 Hz, ArH), 5.4 (br s, 2H, NH2), 4.8
(br s, 1H, NH). 13C NMR (400 MHz, CDCl3): d 117.2 (C-
2, C-5), 123.9 (C-3, C-4), 139.1 (C-1, C-6), 142.5 (C-7).
HRMS (?ESI): m/z 134.0729 (C7H8N3?).
2-[3-(2-Hydroxyphenyl)acrylamido]-benzimidazole (4)
Yield: 69 % (Method A), 65 % (Method B); m.p.
290–294 ꢁC; IR: 3619 (O–H str), 3500–3400 (N–H str.),
3052 (C–H, Ar, str.), 2883 (C–H, Ali, str.), 1680 (C=O
str.), 1631, 1450 (C….C str.), 901 (C–H bending); 1H NMR
(400 MHz, CDCl3): d 7.9–7.8 (br s, 1H, NH), 7.8–7.5 (m,
3H, ArH, CH=CH), 7.4–7.3 (m, 3H, ArH), 7.2–6.9 (m, 2H,
ArH), 6.6–6.5 (m, 3H, ArH, CH=CH), 5.0 (br s, 1H, NH).
13C NMR (400 MHz, CDCl3): d 117.2 (C-2, C-5), 120.5
(C-8), 122.3 (C-14), 123.6 (C-10), 124.1 (C-3, C-4), 128.7
(C-15), 130.7 (C-13), 138.7 (C-1, C-6), 156.7 (C-11), 142.8
(C-7, C-9), 165.8 (C=O). HRMS (?ESI): m/z 280.1198
(C16H14N3O2?).
2-Acetylamino benzimidazole (2)
The 1 (1.33 g, 0.01 M) was dissolved in 15 mL of chlo-
roform and 50 mL of pyridine. Acetic anhydride (1.02 g,
0.01 M) was added dropwise in the reaction mixture with
constant stirring over 4 h at 5–10 ꢁC. The solvent was
recovered on rotary vacuum evaporator, the contents were
dried and recrystallised from ethanol to give pure com-
pound. Yield 52 %; m.p. 280–282 ꢁC; IR: 3500–3400 (N–
H str.), 3052 (C–H, Ar, str.), 2920 (C–H, Ali, str.), 1685
(C=O str.), 1473 (N–H scissoring), 1458 (C=C str.), 1316
(C–H bend), 1234 (C–N str.); 1H NMR (400 MHz, CDCl3):
d 8.0 (s, 1H, NH), 7.5–7.4 (m, 2H, ArH), 7.3–7.1 (m, 2H,
ArH), 5.1 (br s, 1H, NH), 2.5-2.1 (s, 3H, CH). 13C NMR
(400 MHz, CDCl3): d 19.4 (CH3) 117.1 (C-2, C-5), 124.2
(C-3, C-4), 138.6 (C-1, C-6), 143.1 (C-7), 170.2 (C=O).
HRMS (?ESI): m/z 176.0838 (C9H10N3O?).
2-[3-(2-Methoxyphenyl)acrylamido]-benzimidazole (5)
Yield: 60 % (Method A), 67 % (Method B); m.p. 129 ꢁC;
IR: 3500–3400 (N–H str.), 3053 (C–H, Ar, str.), 2872 (C–
H, Ali, str.), 1685 (C=O str.), 1422 (C=C str.), 1218 (C–O
1
str.), 927 (Ar–H bending); H NMR (400 MHz, CDCl3): d
7.9–7.8 (br s, 1H, NH), 7.7–7.6 (m, 2H, ArH), 7.5 (m, 1H,
CH=CH), 7.3–7.1 (m, 5H, ArH, CH=CH), 6.9-6.7 (m, 2H,
ArH), 5.0 (br s, 1H, NH), 3.6 (s, 3H, OCH3). 13C NMR
(400 MHz, CDCl3): d 57.1 (OCH3), 115.7 (C-12), 116.9
(C-2, C-5), 121.1 (C-9), 122.3 (C-10, C-14), 124.2 (C-3,
C-4), 128.7 (C-15), 129.6 (C-13), 138.9 (C-1, C-6), 142.2
(C-7, C-9), 161.6 (C-11), 165.2 (C=O). HRMS (?ESI): m/
z 294.1261 (C17H16N3O2?).
Target compounds (3–11)
Conventional method (Method A) The 2 (1.75 g, 0.01 M)
was dissolved in ethanol (30 mL) and aqueous solution of
KOH (2 %, 5 mL). The aromatic aldehyde (0.01 M) was
added and the reaction mixture was refluxed for 5 h. The
solvent was recovered on rotary vacuum evaporator, the
contents were poured onto crushed ice and acidified with
dilute HCl. The solid separated was filtered and recrystal-
lised from ethanol to obtain the pure product.
2-[3-(2-Nitrophenyl)acrylamido]-benzimidazole (6) Yield:
82 % (Method A), 78 % (Method B); m.p. 122–125 ꢁC; IR:
3500–3400 (N–H str.), 3078 (C–H, Ar, str.), 2875 (C–H, Ali,
str.), 1685 (C=O str.), 1516,1341 (O….N….O str.), 1646,1429
(C….C str.), 924 (C–H bending); 1H NMR (400 MHz,
CDCl3): d 8.0–7.8 (m, 4H, NH, ArH, CH=CH), 7.7–7.6 (m,
2H, ArH), 7.5–7.4 (m, 5H, ArH), 5.0 (br s, 1H, NH). 13C
NMR (400 MHz, CDCl3): d 117.3 (C-2, C-5), 120.7 (C-8),
121.4 (C-10), 124.2 (C-3, C-4), 125.4 (C-12), 128.6 (C-15),
129.9 (C-13), 136.3 (C-14), 139.1 (C-1, C-6), 142.9 (C-7,
C-9), 148.2 (C-11), 166.2 (C=O). HRMS (?ESI): m/z
309.1003 (C16H13N4O3?).
Microwave irradiation method (Method B) The mixture
of 2 (1.75 g, 0.01 M) and aromatic aldehyde (0.01 M) in
aqueous solution of KOH (2 %, 5 mL) was irradiated in
microwave for 2 min at 20 % power output. The reaction
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Med Chem Res (2014) 23:2690–2697
2-[3-(2-Chlorophenyl)acrylamido]-benzimidazole (7)
CDCl3): d 8.0–7.9 (br s, 1H, NH), 7.7–7.6 (m, 3H, ArH,
CH=CH), 7.3–7.1 (m, 6H, ArH), 6.9–6.8 (m, 1H, CH=CH),
5.0 (br s, 1H, NH). 13C NMR (400 MHz, CDCl3): d 116.9
(C-2, C-5), 121.3 (C-8), 124.3 (C-3, C-4), 128.8 (C-11,
C-15), 130.2 (C-12, C-14), 134.5 (C-10, C-13), 138.7 (C-1,
C-6), 142.3 (C-7, C-9), 165.1 (C=O). HRMS (?ESI): m/
z 298.0756 (C16H13N3OCl?).
Yield: 65 % (Method A), 72 % (Method B); m.p.
111–115 ꢁC; IR: 3500–3400 (N–H str.), 3054 (C–H, Ar,
str.), 2886 (C–H, Ali, str.), 1682 (C=O str.), 1652, 1425
(C….C str.), 930 (C–H bending); 1H NMR (400 MHz,
CDCl3): d 8.0–7.9 (br s, 1H, NH), 7.7–7.6 (m, 3H, ArH,
CH=CH), 7.3–7.1 (m, 7H, ArH, CH=CH), 5.0 (br s, 1H,
NH). 13C NMR (400 MHz, CDCl3): d 116.8 (C-2, C-5),
120.9 (C-8), 124.1(C-3, C-4), 127.6 (C-14), 128.5 (C-15),
130.1 (C-12, C-13), 132.8 (C-11), 136.7 (C-10), 138.5(C-1,
C-6), 143.2 (C-7, C-9), 165.3 (C=O). HRMS (?ESI): m/
z 298.0759 (C16H13N3OCl?).
PASS prediction of the target compounds
Biological activities of the target compounds were pre-
dicted by a computer software PASS. The activities pre-
dicted for each target compound (3–11) having Pa [ 0.5
are listed in Table 1. The Lipinski’s parameters were cal-
culated (Table 3) through Molinspiration software (Lipin-
ski et al., 2001).
2-[3-(4-Hydroxyphenyl)acrylamido]-benzimidazole (8)
Yield: 75 % (Method A), 68 % (Method B); m.p.
256–260 ꢁC; IR: 3610 (Free O–H str.), 3500–3400 (N–H
str.), 3035 (C–H, Ar, str.), 2887 (C–H, Ali, str.), 1680
1
(C=O str.), 1636,1421 (C….C str.), 923 (C–H bending); H
NMR (400 MHz, CDCl3): d 8.0–7.9 (br s, 1H, NH), 7.7-7.6
(m, 3H, ArH, CH=CH), 7.3–7.1 (m, 4H, ArH), 6.9–6.8 (m,
ArH, CH=CH), 5.0 (br s, 1H, NH). 13C NMR (400 MHz,
CDCl3): d 116.9 (C-2, C-5, C-12, C-14), 121.3 (C-8), 123.8
(C-3, C-4), 128.4 (C-10, C-11, C-15), 138.7 (C-1, C-6),
143.5 (C-7, C-9), 157.9 (C-13), 165.4 (C=O). HRMS
(?ESI): m/z 280.1195 (C16H14N3O2?).
Anthelmintic activity
The anthelmintic activity of each compound was evaluated
on Indian adult earthworms (Pheretima posthuma) col-
lected from moist soil and cleaned by washing with normal
saline. The activity was evaluated using the method
described in various reports (Ajaiyeoba et al., 2001; Dash
et al., 2002; Shivkumar and Kumar, 2003; Kosalge and
Fursule, 2009). Briefly, the worms were divided into dif-
ferent groups containing six earthworms in each group and
washed with the normal saline. All the target compounds
were evaluated at three concentrations (0.1, 0.2, and 0.5 %
w/v) taking albendazole as standard drug. The helminthes
were placed in solutions of each compound at each con-
centration individually in Petri plates. The time taken for
worms to become motionless was noted as paralysis time,
whereas the time taken for death of the worm was noted as
lethal time. To ascertain death, each worm was frequently
subjected to external stimuli that stimulate and induce
movement in earth worms, if alive. The time taken for
complete paralysis and death was recorded. The mean
paralysis time and mean lethal time were calculated for
each compound at each concentration.
2-[3-(4-Methoxyphenyl)acrylamido]-benzimidazole (9)
Yield: 67 % (Method A), 60 % (Method B); m.p.
196–200 ꢁC; IR: 3500–3400 (N–H str.), 3047 (C–H, Ar,
str.), 2870 (C–H, Ali, str.), 1682 (C=O str.), 1643, 1420
(C….C str.), 927 (C–H bending); 1H NMR (400 MHz,
CDCl3): d 7.9–7.8 (br s, 1H, NH), 7.7–7.6 (m, 2H, ArH),
7.5 (m, 1H, CH=CH), 7.3–7.1 (m, 4H, ArH), 6.9–6.7 (m,
3H, ArH, CH=CH), 5.0 (br s, 1H, NH), 3.41 (m, 3H,
OCH3). 13C NMR (400 MHz, CDCl3): d 57.4 (OCH3),
115.2 (C-12, C-14), 117.4 (C-2, C-5), 121.4 (C-8), 124.3
(C-3, C-4), 128.3 (C-10, C-11, C-15), 138.8 (C-1, C-6),
142.9 (C-7, C-9), 162.7 (C-13), 166.1 (C=O). HRMS
(?ESI): m/z 294.1256 (C17H16N3O2?).
2-[3-(4-Nitrophenyl)acrylamido]-benzimidazole (10) Yield:
73 % (Method A), 85 % (Method B); m.p. 226–230 ꢁC; IR:
3500–3400 (N–H str.), 3048 (C–H, Ar, str.), 2874 (C–H, Ali,
str.), 1680 (C=O str.), 1523,1336 (O….N….O str.), 1640, 1449
(C….C str.), 927 (C–H bending); 1H NMR (400 MHz, CDCl3):
d 8.0–7.8 (m, 5H, NH, ArH), 7.7–7.4 (m, 4H, ArH, CH=CH),
7.4–7.3 (m, 2H, ArH), 5.0 (br s, 1H, NH). 13C NMR
(400 MHz, CDCl3): d 116.9 (C-2, C-5), 120.7 (C-8), 124.5 (C-
3, C-4), 126.4 (C-12, C-14), 128.2 (C-11, C-15), 139.2 (C-1,
C-6), 141.9 (C-10), 142.8 (C-7, C-9), 149.3 (C-13), 165.3
(C=O). HRMS (?ESI): m/z 309.0997 (C16H13N4O3?).
Statistical analysis
Results are expressed as mean SD. The statistical sig-
nificance of the observed data was determined by one-way
analysis of variance (ANOVA) followed by Tukey’s test
for the anthelmintic activity of the synthesized compounds
and the results were reported to be statistically significant
at p \ 0.05.
2-[3-(4-Chlorophenyl)acrylamido]-benzimidazole (11)
Yield: 76 % (Method A), 70 % (Method B); m.p.
276–280 ꢁC; IR: 3500–3400 (N–H str.), 3050 (C–H, Ar,
str.), 2873 (C–H, Ali, str.), 1680 (C=O str.), 1652,1423
(C….C str.), 936 (C–H bending); 1H NMR (400 MHz,
Acknowledgments The authors are thankful to Punjabi University,
Patiala for providing the infrastructure and financial assistance, to All
India Council for Technical Education (AICTE), New Delhi for
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Med Chem Res (2014) 23:2690–2697
2697
fellowship to one of the authors (Kamalpriya Garg), to Crystal
Pharmaceuticals (Ambala, India) for albendazole as a generous gift
sample, and to molsinpiration.com for free access to the software for
calculation of the Lipinski’s parameters. Authors acknowledge the
free PASS software license provided by Dr. V.V. Poroikov and
IBMC, Russian Academy of Medical Sciences, Moscow.
and bioactivity score was done by software available at website
http://www.molinspiration.com/
Marawaha A, Goel RK, Mahajan MP (2007) PASS-predicted design,
synthesis and biological evaluation of cyclic nitrones as
nootropics. Bioorg Med Chem Lett 17:5251–5255
Mathew B, Suresh J, Anbazhagan S (2012) Development of novel
(1H) benzimidazole bearing pyrimidine-trione based MAO-A
inhibitors: synthesis, docking studies and antidepressant activity.
J Saudi Chem Soc. doi:10.1016/j.jscs.2012.09.015
Mishra L, Sinha R, Itokawa H, Kenneth FB, Tachibana Y, Nakanishi
Y, Kilgore N, Lee KH (2001) Anti-HIV and cytotoxic activities
of Ru(II)/Ru(III) polypyridyl complexes containing 2,6-(20-
Benzimidazolyl)-pyridine/chalcone as co-ligand. Biorg Med
Chem. 9:1607–1617
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