Synthesis and biological evaluation of benzo[d]imidazolyl chromeno[2,3-d]pyrimidinones

Article abstract of DOI:10.1007/s00044-013-0674-5

A series of benzo[d]imidazolyl chromeno[2,3-d]pyrimidnones were described. The key intermediate 2-cyano-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)acetamide (3) is obtained by reacting 5-amino-2-mercaptobenzimidazole (1) with ethyl cyanoacetate (2). Compound 3 on reaction with substituted salicylaldehydes afforded 2-imino-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H-chromene-3- carboxamides (5) in good yields. Compounds 5 on condensation with formalin furnished the title compounds viz., 3-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H- chromeno[2,3-d]pyrimidin-4(3H)-ones (7). All the synthesized compounds were screened for their anti-microbial and anti-oxidant activities.

Full text of DOI:10.1007/s00044-013-0674-5

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-013-0674-5
ORIGINAL RESEARCH
Synthesis and biological evaluation of benzo[d]imidazolyl
chromeno[2,3-d]pyrimidinones
•
Anisetti Ravindernath Malladi Srinivas Reddy
•
Vodela Sunil
Received: 9 January 2013 / Accepted: 6 June 2013
Ó Springer Science+Business Media New York 2013
Abstract A series of benzo[d]imidazolyl chromeno[2,
3-d]pyrimidnones were described. The key intermediate
2-cyano-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)acetamide
(3) is obtained by reacting 5-amino-2-mercaptobenzimida-
zole (1) with ethyl cyanoacetate (2). Compound 3 on reaction
with substituted salicylaldehydes afforded 2-imino-N-(2-
mercapto-1H-benzo[d]imidazol-5-yl)-2H-chromene-3-car-
boxamides (5) in good yields. Compounds 5 on condensation
with formalin furnished the title compounds viz., 3-(2-mer-
capto-1H-benzo[d]imidazol-5-yl)-2H-chromeno[2,3-d]pyr-
imidin-4(3H)-ones (7). All the synthesized compounds were
screened for their anti-microbial and anti-oxidant activities.
have been identified, and the presence of the chromeno-based
structure has been associated with the capacity to prevent
diseases (O’Kennedy and Thornes, 1997). Synthetic analogs
have been developed over the years, some of them displaying
remarkable effects as pharmaceuticals (Borges et al., 2005),
including anti-fungal (Tangmouo et al., 2006), anti-microbial
(Kitamura et al., 2006), molluscidial (Abdelrazek et al.,
2007), anti-coagulant, spasmolytic, diuretic, anti-cancer, and
anti-anaphylactic characteristics (Singh et al., 1996). Pyrim-
idines and its fused derivatives play an essential role in several
biological processes and chemical and pharmacological
importance. In particular, pyrimidine nucleus can be found in
a broad variety of anti-bacterial and anti-tumor agents as well
as in agrochemical and veterinary products (Ismali et al.,
2008; El-Gaby et al., 2006; Prikazchikova et al., 1975; Brown
et al., 1984).
Keywords
Benzo[d]imidazolyl chromeno[2,3-d]pyrimidinones ꢀ
Condensation ꢀ Anti-bacterial activity ꢀ
Anti-fungal activity ꢀ Anti-oxidant activity
Benzimidazole derivatives are also reported to possess
analgesic (Sondhi et al., 2002), anti-helminthic (Hazelton
et al., 1995; Labaw and Webb, 1981), anti-inflammatory (Ito
et al., 1982), anti-microbial, anti-arthritic, anti-bacterial,
anti-tumor, and anti-HIV activities (Rao et al., 2002). This in
view of diverse therapeutic activity of chromenes, pyrimi-
dines, and benzimidazoles and in the course of our search for
an effective biomimetic model compounds (Ravindernath
and Srinivas Reddy, 2012), chromeno[2,3-d]pyrimidines,
which can be regarded as a 5-deazaflavin analog, was
selected as a target molecule (Fig. 1; Scheme 1).
Introduction
Chromeno derivatives are an important class of compounds,
widelypresent in plants, including ediblevegetablesandfruits
(Curini et al., 2006). Numerous bioactive natural products
A. Ravindernath (&)
University College of Pharmacy, Osmania University,
Hyderabad 500007, AP, India
e-mail: ravindernathanisetti@gmail.com
M. S. Reddy
Experimental
Department of Pharmaceutical Chemistry, St. Peter’s Institute
of Pharmaceutical Sciences, Vidyanagar,
Warangal 506001, AP, India
Materials and methods
V. Sunil
Department of Chemistry, Talla Padmavathi College of
Engineering, Kazipet 506003, AP, India
All the melting points were determined on Cintex melting
point apparatus and are uncorrected. Analytical TLC was
123

Med Chem Res
O
N
refluxed for 5 h. After the completion of the reaction
(monitored by TLC), reaction mixture was cooled and the
solid obtained was filtered off and recrystallized from ethyl
acetate to give benzimidazolyl cyanoacetamide 3.
O
N
R
R
N
N
N
R
O
O
O
5-deazaflavin
5
-deaza-10-oxaflavin
2-Cyano-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)acetam-
ide (3) m.p. 116–118 °C, Yield, 78 %; IR (KBr) cm-1
:
Fig. 1 Biologically active chromeno[2,3-d]pyrimidines as design
templates
3363, 3176, 2471, 1618. 1H NMR (CDCl3) d (ppm): 3.58 (s,
1H, SH) 3.80 (s, 2H, CH2CN), 4.98 (s, 1H, NH, D2O
exchangeable), 6.93 (s, 1H), 7.14 (d, 1H), 7.18 (d, 1H), 8.58
(s, 1H, CONH, D2O exchangeable). ESI–MS m/z 233
[M?H]?. 13C NMR (75 MHz, CDCl3): 23.90, 108.22,
115.48, 123.74, 125.71, 134.57, 135.29, 139.20, 168.24,
168.45. Anal. Calcd. for C10H8N4OS: C, 51.71; H, 3.47; N,
24.12. Found: C, 51.65; H, 3.50; N, 24.07 %.
performed on Merck precoated 60 F254 silica gel plates.
Visualization was done by exposure to iodine vapor. IR
spectra (KBr pellet) were recorded on a Perkin-Elmer BX
series FT-IR spectrometer. 1H NMR spectra were recorded
on a Varian Gemini 300 MHz spectrometer. 13C NMR
spectra were recorded on a Bruker 75 MHz spectrometer.
Chemical shift values are given in ppm (d) with tetramethyl
silane as an internal standard. Mass spectral measurements
were carried out by EI method on a Joel JMC-300 spec-
trometer at 70 eV. Elemental analyses were performed on a
Carlo Erba 106 and Perkin-Elmer model 240 analyzers.
General procedure for the synthesis of benzimidazolyl
chromene-3-carboxamides (5a–e) To a solution of com-
pound 2 (1 mmol) in absolute ethanol (20 mL) containing
piperidine (0.5 mL), substituted salicylaldehydes
4
Synthesis
(1 mmol) was added. The reaction mixture was heated
under reflux for 3 h and then allowed to cool. The pre-
cipitate obtained on pouring the reaction mixture into
crushed ice was filtered off, washed with ethanol, dried,
and recrystallized from ethyl acetate to afford 5. Spectral
data of each compound are given below.
General procedure for the synthesis of 2-cyano-N-(2-mer-
capto-1H-benzo[d]imidazol-5-yl)acetamide (3) A mix-
ture of 5-amino-2-mercapto benzimidazole 1 (1 mmol) and
ethyl cyanoacetate 2 (1 mmol) in ethanol (15 mL) were
CHO
H
O
N
Ethanol
N
OH
NH2
N
CN
HS
+
R
CN
H C O
+
5
2
HS
N
H
O
N
H
R
4
3
2
1
Ethanol
Piperidine
O
H
N
N
HCHO
+
HS
HN
O
R
N
H
R
6
5
Ethanol
O
N
N
HS
N
H
N
O
R
R
7
R= H, OCH3 , Br, Cl
R' = Br
Scheme 1 Synthesis of benzimidazolyl chromeno[2,3-d]pyrimidin-4(3H)-ones (7a–e)
123

Med Chem Res
2-Imino-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H-
7,8-Dibromo-2-imino-N-(2-mercapto-1H-benzo[d]imidazol-
chromene-3-carboxamide (5a) m.p. 168–170 °C Yield,
5-yl)-2H-chromene-3-carboxamide (5e) m.p. 177–179 °C.
1
1
79 %; IR (KBr) cm-1: 3310, 3250, 3200, 1770. H NMR
Yield, 75 %; IR (KBr) cm-1: 3311, 3232, 3201, 1770. H
(CDCl3) d (ppm): 4.42 (s, 1H, SH), 6.24 (s, 1H, =CH),
6.98–7.40 (m, 7H, Ar–H), 8.62 (s, 1H, NH, D2O
exchangeable), 9.46 (s, 1H, =NH), 10.20 (s, 1H, CONH,
D2O exchangeable). ESI–MS m/z 337 [M?H]?. 13C NMR
(75 MHz, CDCl3) d (ppm): 107.37, 113.81, 115.20,
115.57, 119.01, 121.67, 123.72, 127.98, 130.16, 132.65,
134.54, 138.90, 140.45, 141.54, 155.23, 162.67, 164.10.
Anal. calcd for C17H12N4O2S: C, 60.70; H, 3.60; N, 16.66;
Found C, 60.76; H, 3.58; N, 16.68 %.
NMR (CDCl3) d (ppm): 4.20 (s, 1H, SH), 6.10 (s, 1H,
=CH), 6.91–7.81 (m, 5H, Ar–H), 8.30 (s, 1H, NH, D2O
exchangeable), 9.21 (s, 1H, =NH), 10.00 (s, 1H, CONH,
D2O exchangeable). ESI–MS m/z 495 [M?H]?. 13C NMR
(75 MHz, CDCl3) d (ppm): 107.96, 114.21, 116.20,
116.71, 119.88, 120.99, 122.72, 126.98, 131.16, 133.15,
134.86, 137.90, 141.45, 142.24, 154.99, 163.47, 164.89.
Anal. calcd for C17H10Br2N4O2S: C, 41.32; H, 2.04; N,
11.34; Found C, 41.37; H, 2.10; N, 11.38 %.
2-Imino-N-(2-mercapto-1H-benzo[d]imidazol-5-yl)-7-meth-
General procedure for the synthesis of benzimidazolyl
chromeno[2,3-d]pyrimidin-4(3H)-ones (7a–e) A mixture
of compound 5 (1 mmol) and formalin 6 (37 % 1 mmol) was
refluxed in ethanol (15 mL) for 4 h. The reaction mixture
after cooling to room temperature was poured into ice-cold
water. The separated solid was filtered and recrystallized
from benzene to get pure compounds 7a–e. Spectral data of
each compound are given below.
oxy-2H-chromene-3-carboxamide (5b) m.p. 151–153 °C.
1
Yield, 75 %; IR (KBr) cm-1: 3210, 3125, 3053, 1677. H
NMR (CDCl3) d (ppm): 3.63 (s, 3H, OCH3), 4.38 (s, 1H,
SH), 6.18 (s, 1H, =CH), 6.87–7.61 (m, 6H, Ar–H), 8.32 (s,
1H, NH, D2O exchangeable), 9.48 (s, 1H, =NH), 10.12 (s,
1H, CONH, D2O exchangeable). ESI–MS m/z 367
[M?H]?. 13C NMR (75 MHz, CDCl3) d (ppm): 63.14,
106.96, 114.15, 114.50, 115.07, 119.01, 120.67, 122.42,
126.98, 129.96, 131.85, 133.94, 137.99, 141.05, 142.04,
154.85, 163.32, 164.19. Anal. calcd for C18H14N4O3S: C,
59.01; H, 3.85; N, 15.29; Found C, 59.05; H, 3.81; N,
15.31 %.
3-(2-Mercapto-1H-benzo[d]imidazol-5-yl)-2H-chromeno[2,
3-d]pyrimidin-4(3H)-one (7a) m.p. 203–205 °C. Yield,
1
77 %; IR (KBr) cm-1: 3375 (NH), 1631 (C=O). H NMR
(CDCl3) d (ppm): 4.50 (s, 2H, CH2), 4.56 (s, 1H, SH), 6.51
(s, 1H, =CH), 6.81–7.66 (m, 7H, Ar–H), 8.13 (s, 1H, NH,
D2O exchangeable). ESI–MS m/z 349 [M?H]?. 13C NMR
(75 MHz, CDCl3) d (ppm): 61.91, 106.45, 115.37, 115.92,
116.53, 121.75, 122.17, 122.54, 127.42, 129.93, 131.53,
133.78, 138.30, 142.16, 142.72, 154.63, 162.67, 164.53.
Anal. calcd for C18H12N4O2S: C, 62.06; H, 3.47; N, 16.08;
Found C, 62.01; H, 3.40; N, 16.14 %.
7-Bromo-2-imino-N-(2-mercapto-1H-benzo[d]imidazol-5-
yl)-2H-chromene-3-carboxamide (5c) m.p. 163–165 °C
1
Yield, 81 %; IR (KBr) cm-1: 3330, 3231, 3225, 1770. H
NMR (CDCl3) d (ppm): 4.36 (s, 1H, SH), 6.23 (S, 1H,
=CH), 6.97–7.20 (m, 6H, Ar–H), 8.22 (s, 1H, NH, D2O
exchangeable), 9.37 (s, 1H, =NH), 10.01 (s, 1H, CONH,
D2O exchangeable). ESI–MS m/z 415 [M?H]?. 13C NMR
(75 MHz, CDCl3) d (ppm):108.01, 114.21, 114.90, 115.01,
120.21, 122.07, 122.92, 126.98, 131.16, 133.05, 133.94,
139.01, 139.95, 140.84, 154.93, 163.67, 163.99. Anal.
calcd for C17H11BrN4O2S: C, 49.17; H, 2.67; N, 13.49;
Found C, 49.21; H, 2.61; N, 13.54 %.
3-(2-Mercapto-1H-benzo[d]imidazol-5-yl)-8-methoxy-2H-
chromeno[2,3-d]pyrimidin-4(3H)-one (7b) m.p. 210–
214 °C. Yield, 79 %; IR (KBr) cm-1: 3250, 1609.1H NMR
(CDCl3) d (ppm): 3.63 (s, 3H, OCH3), 4.22 (s, 1H, SH),
4.55 (s, 2H, CH2), 6.11 (s, 1H, =CH), 6.92–7.30 (m, 6H,
Ar–H), 8.18 (s, 1H, NH, D2O exchangeable). ESI–MS m/z
379 [M?H]?. 13C NMR (75 MHz, CDCl3) d (ppm): 62.11,
63.56, 107.15, 114.98, 116.22, 117.43, 120.98, 121.77,
123.24, 127.98, 128.33, 132.83, 133.08, 137.87, 142.93,
143.55, 154.88, 163.07, 164.96. Anal. calcd for C19H14N4
O3S: C, 60.31; H, 3.73; N, 14.81; Found C, 60.22; H, 3.78;
N, 14.86 %.
7-Chloro-2-imino-N-(2-mercapto-1H-benzo[d]imidazol-5-
yl)-2H-chromene-3-carboxamide (5d) m.p. 170–172 °C.
1
Yield, 84 %; IR (KBr) cm-1: 3353, 3240, 3215, 1737. H
NMR (CDCl3) d (ppm): 4.20 (s, 1H, SH), 6.11 (s, 1H,
=CH), 6.82–7.71 (m, 6H, Ar–H), 8.43 (s, 1H, NH, D2O
exchangeable), 9.40 (s, 1H, =NH),10.18 (s, 1H, CONH,
D2O exchangeable). ESI–MS m/z 371 [M?H]?. 13C NMR
(75 MHz, CDCl3) d (ppm): 106.96, 112.90, 114.80,
115.01, 120.01, 122.45, 122.85, 127.02, 131.08, 133.32,
134.98, 137.89, 141.55, 142.54, 154.83, 162.94, 163.90.
Anal. calcd for C17H11ClN4O2S: C, 55.06; H, 2.99; N,
15.11; Found C, 55.10; H, 2.94; N, 15.15 %.
8-Bromo-3-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H-
chromeno[2,3-d]pyrimidin-4(3H)-one (7c) m.p. 218–
222 °C. Yield, 74 %; IR (KBr) cm-1: 3215, 1730. 1H
NMR (CDCl3) d (ppm): 4.33 (s, 2H, CH2), 4.62 (s, 1H,
SH), 6.51 (s, 1H, =CH), 6.34–7.22 (m, 6H, Ar–H), 8.21 (s,
1H, NH, D2O exchangeable). ESI–MS m/z 427 [M?H]?.
123

Med Chem Res
13C NMR (75 MHz, CDCl3) d (ppm): 62.11, 107.35,
116.07, 116.92, 117.23, 122.35, 123.57, 123.98, 127.42,
130.63, 132.65, 134.48, 138.99, 143.11, 143.82, 153.58,
163.27, 164.85. Anal. calcd for C18H11BrN4O2S: C, 50.60;
H, 2.59; N, 13.11; Found C, 50.63; H, 2.55; N, 13.15 %.
volumeof0.5 mLofovernightculturewasaddedinallthetest
tube which was incubated at 37 °C for 24 h. After 24 h, these
tubes were measured for turbidity.
Anti-fungal assay
8-Chloro-3-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H-
chromeno[2,3-d]pyrimidin-4(3H)-one (7d) m.p. 220–
224 °C. Yield, 79 %; IR (KBr) cm-1: 3225, 1740. 1H
NMR (CDCl3) d (ppm): 4.31 (s, 2H, CH2), 4.68 (s, 1H,
SH), 6.44 (s, 1H, =CH), 6.45–7.28 (m, 6H, Ar–H), 8.24 (s,
1H, NH, D2O exchangeable).ESI–MS m/z 383 [M?H]?.
13C NMR (75 MHz, CDCl3) d (ppm): 62.56, 105.99,
116.37, 116.97, 117.32, 120.99, 121.47, 123.24, 128.02,
129.01, 132.33, 134.48, 138.86, 141.93, 142.01, 153.63,
161.85, 164.53. Anal. calcd for C18H11ClN4O2S: C, 56.47;
H, 2.90; N, 14.64; Found C, 56.52; H, 2.94; N, 14.58 %.
The target compounds 7a–e were also evaluated for their anti-
fungal activity against Candida albicans and Aspergillus
niger in acetone by agar cup bioassay method (Margery
Linday, 1962), using fluconazole as the standard drug. The
MIC was recorded in lg mL-1. The readymade potato dex-
trose agar media (Himedia, 39 g) was suspended in distilled
water (100 mL) and heated until it dissolved completely. The
medium and Petri dishes were autoclaved at a pressure of
15 lb inc-2 for 20 min. The medium 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
(week-old) culture of test organism was inoculated and uni-
formly spread over the agar surface with a sterile L-shaped
rod. Solutions were prepared by dissolving plant extract in
acetone (100 lg mL-1). Agar inoculation cups were scooped
outwith6 mmsterilecorkborerandthelidsofthedisheswere
replaced. To each cup, 100 lg mL-1 of test solution was
added. Controls were maintained with acetone and clotrima-
zole (100 lg mL-1). The treated and the control were kept at
room temperature for 72–96 h. Inhibition zones were mea-
sured and the diameter was calculated in millimeters. Three to
four replicates were maintained for each treatment.
8,9-Dibromo-3-(2-mercapto-1H-benzo[d]imidazol-5-yl)-2H-
chromeno[2,3-d]pyrimidin-4(3H)-one (7e) m.p. 250–
254 °C. Yield, 82 %; IR (KBr) cm-1: 3245, 1760. 1H
NMR (CDCl3) d (ppm): 4.25 (s, 2H, CH2), 4.62 (s, 1H,
SH), 6.38 (s, 1H, =CH), 6.51–7.66 (m, 5H, Ar–H), 8.30 (s,
1H, NH, D2O exchangeable). ESI–MS m/z 507 [M?H]?.
13C NMR (75 MHz, CDCl3) d (ppm): 60.99, 107.35,
114.89, 116.22, 116.89, 120.75, 122.78, 123.44, 126.12,
128.63, 132.23, 133.87, 137.75, 141.76, 143.27, 153.73,
161.34, 162.99. Anal. calcd for C18H10Br2N4O2S: C, 42.71;
H, 1.99; N, 11.07; Found C, 42.75; H, 1.95; N, 11.11 %.
The structures of all newly synthesized compounds were
Anti-oxidant activity
1
confirmed by analytical and spectral data (IR, H NMR,
and MS).
For the evaluation of anti-oxidant activity, we have used a
stable free radical a,a-diphenyl-b-picryl hydrazyl (DPPH), at
the concentration of 0.2 mM in methanol (Ranjit et al., 2010).
To 0.1 mL of test compound (at different concentrations),
1.5 mL of methanol and 0.5 mL of DPPH solution were
added, mixed thoroughly and absorbance (OD) was read at
517 nm against the blank. The % reduction of free radical
concentration (OD) with different concentration of test com-
pounds was calculated and was compared with standard,
ascorbic acid. The results were expressed as IC50 values (the
concentration of test required to scavenge 50 % free radicals).
Pharmacological screening
Anti-bacterial assay
The newly synthesized compounds 7a–e were evaluated for
their in vitro anti-bacterial activity against Gram-positive
bacteria viz., Bacillus subtilis, Bacillus cereus and Gram-
negative bacteria viz. Micrococcus luteus, Entamoeba Coli at
100 lg mL-1 concentration. The in vitro anti-bacterial
activity of the tested compounds was assessed by minimum
inhibitory concentration (MIC) using broth dilution method
(NCCLS, 1982). Ciprofloxacin was used as standard com-
parison. The readymade nutrient broth media (Himedia, 24 g)
was suspended in distilled water (100 mL) and heated until it
dissolved completely. The medium and test tubes were auto-
claved at a pressure of 15 lb inc-2 for 20 min. A set of ster-
ilized test tubes with nutrient broth medium was capped with
cotton plugs. The test compound was dissolved in DMSO and
a concentration of 250 lg mL-1 of the test compounds was
added in the first test tube, which was serially diluted. A fixed
Results and discussion
Synthesis
The key intermediate, 2-cyano-N-(2-mercapto-1H-benzo[d]
imidazol-5-yl)acetamide 3 required for synthesis of target
compounds was obtained by reacting 5-amino-2-mercapto
benzimidazole 1 with ethyl cyanoacetate 2 in refluxing
ethanol. IR spectrum of benzimidazolyl cyanoacetamide 3
123

Med Chem Res
showed absorption bands 3363, 3176, 2471, and
1618 cm-1 benzimidazole-NH, NH, CN, and C=O func-
bacteria viz., M. luteus, E. Coli at 100 lg mL-1 concen-
tration. The results of anti-bacterial screening (Table 1)
reveal that the compounds 7a–e displayed a better activity
and more active than the standard Ciprofloxacin. Com-
pound 7b possessing methoxy group as substituent on the
benzene ring showed a better activity. Compound 7a
exhibited least activity because it has no substituent on the
benzene ring. However, the degree of inhibition varied
both with the test compound and with the bacteria used.
The title compounds 7a–e were also evaluated for their
anti-fungal activity against C. albicans and A. niger in
acetone by agar cup bioassay method (19), using clotrim-
azole as standard drug. The anti-fungal activity results
(Table 2) indicate that compounds 7a–e are significantly
toxic toward all the fungi under investigation. Compound
7c possessing bromo substituent on benzene ring are highly
toxic toward all the fungi. However, the degree or spore
germination inhibition varied with the test compound as
well as with the fungi under investigation.
1
tional group stretching vibrations, respectively. H NMR
spectra of 3 showed singlet at d 3.80 due to CH2 proton,
broad singlet at d 8.58 CONH proton. The mass spectrum
of 3 is in conformity with the structure by displaying the
molecular ion peak at [M?H]? m/z 233.
Cyclocondensation of benzimidazolyl cyanoacetamide 3
with substituted salicylaldehydes 4 in boiling absolute
ethanol containing a catalytic amount of piperidine affor-
ded the chromeno derivatives viz., 2-imino-N-(2-mercapto-
1H-benzo[d]imidazol-5-yl)-2H-chromene-3-carboxamides
5 in good yields (75–85 %). The scope and generality of
this reaction is illustrated by reacting with benzimidazolyl
cyanoacetamide and different substituted salicylaldehydes
afforded their corresponding derivatives 5a–e without any
difficulty. In the IR spectra of compounds 5 exhibited three
NH stretching absorptions at 3310, 3250, 3200, and amide
carbonyl at 1770 cm-1 confirm the cyclization. Further
support was obtained from 1H NMR spectra of 5, in which
benzimidazole-NH, imine-NH, and amide-NH proton sig-
nals appeared as singlets at d 8.62, 9.46, and 10.20,
respectively, which are D2O exchangeable. The mass
spectrum of 5a is in conformity with the assigned structure,
which displayed the molecular ion peak at [M?H]? m/z
337 corresponding to its molecular formula. Elemental
analyses satisfactorily confirmed elemental composition
and purity of the synthesized compounds.
Anti-oxidant activity
The anti-oxidant DPPH free radical scavenging activity of
all the synthesized compounds 7a–e were performed using
DPPH method is shown in Table 3. The IC50 values of all
the compounds 7a–e were found between 12.62 and
56.45 lM, with anti-oxidant activity. In the series,
Compounds 5 on heating with formalin in ethanol affor-
ded the corresponding 3-(2-mercapto-1H-benzo [d]imida-
zol-5-yl)-2H-chromeno[2,3-d]pyrimidin-4(3H)-ones 7 in
good yields. The IR spectra of compounds 5 showed the
disappearance of amide-NH and imine-NH functional group
stretching absorption bands, present in its precursor 5 at 3310
and 3250 cm-1, indicating the evidence for cyclization and
formation of title compound 5. In 1H NMR spectra of 5, the
absence of imine-NH and amide-NH proton signals (present
in its precursor 5 at d 9.46 and 10.20) and appearance of a
new signal as a singlet at d 4.50 is due to –N–CH2–N– pro-
tons clearly confirms the product formation 7. Moreover, the
mass spectrum of 7a showed the molecular ion peak at m/z
349 [M?H]?, which well agrees with the molecular weight
of the proposed structure. C, H, N analyses are also in full
conformity with the assigned structures 7a–e.
Table 1 Anti-bacterial activity of benzimidazolyl chromeno[2,3-
d]pyrimidin-4(3H)-ones (7a–e) (minimum inhibitory concentration)
Compound
B. subtilis
B. cereus
M. luteus
E. coli
7a
21
18
23
20
26
24
23
20
20
26
21
24
18
17
19
24
18
25
22
19
26
20
23
25
7b
7c
7d
7e
Ciprofloxacin
Negative control (acetone)––no activity; concentration 100 lg mL-1
Table 2 Anti-fungal activity of benzimidazolyl chromeno[2,3-
d]pyrimidin-4(3H)-ones (7a–e) (zone of inhibition in mm)
Compound
C. albicans
A. niger
7a
22
22
36
20
24
24
24
23
27
28
20
18
Biological screening
7b
7c
Anti-microbial activity
7d
7e
The newly synthesized compounds 7a–e were evaluated for
their in vitro anti-bacterial activity against Gram-positive
bacteria viz., B. subtilis, B. cereus and Gram-negative
Fluconazole
Negative control (acetone)––no activity; concentration 100 lg mL-1
123

Med Chem Res
thieno[2,3-b]quinoline, quinolino[30,20:4,5] thieno[3,2-d]pyrimi-
dine and pyrido[20,30:4,5]thieno[2,3-b]quinoline derivatives.
Phosphorus Sulfur Silicon 181:279
Table 3 Anti-oxidant activity of benzimidazolyl chromeno[2,3-
d]pyrimidin-4(3H)-ones (7a–e)
Compound
R
R0
IC50 (lL)
Hazelton JC, lldon B, Suschitzky H, Wolley LH (1995) Synthesis of
polysubstituted o-phenylenediamines and their conversion into
heterocycles, particularly 2-substituted benzimidazoles with
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Ito H, Kagaya T, Fukuda K, Nose T (1982) Pharmacological studies
of a new non-steroidal anti-inflammatory drug: 2-(5-ethylpyri-
din-2-yl)benzimidazole. Arzneim Forsch 32:49
7a
H
H
H
H
H
Br
–
56.45
45.20
21.36
29.46
12.62
8.64
7b
OCH3
Br
7c
7d
Cl
7e
Br
Ascorbic acid
–
Kitamura ROS, Romoff P, Young MCM, Kato MJ, Lago JHG (2006)
Chromenes from Peperomia serpens (Sw.) Loudon (Piperaceae).
Phytochemistry 67:2398
Labaw CS, Webb RL (1981) N-Phenyl-N’-cyano-o-phenylisourea. US
Patent 4285878. Chem Abstr. (1981) 95, 168837
compounds 7c and 7e possessing bromo and dibromo
substituent’s on the benzene ring showed better activity
against DPPH free radicals.
Margery Linday E (1962) Practical introduction to microbiology. E &
F.N. Spon Ltd, London, p 177
National Committee for Clinical Laboratory Standards (NCCLS)
(1982) Method for dilution antimicrobial susceptibility tests for
bacteria, which grows aerobically. National Committee for
Clinical Laboratory Standards, Ltd., Villanova, p 242
O’Kennedy P, Thornes RD (eds) (1997) Coumarins: biology,
applications and mode of action. Wiley, Chichester
Prikazchikova LP, Khutova BM, ladimirtsev IF, Boldyrev IV,
Zhuravskaya NI (1975) Stimulating activity of mono- and
diketone pyrimidine derivatives. Fiziol Akt Veshchestva. 7:84.
Chem. Abstr. 83. 127346m
Ranjit K, Rao GK, Pai PNS (2010) Synthesis and biological
evaluation of N1-[(3Z)-5-substituted-2-oxo-1,2-dihydro-3H-in-
dol-3-ylidene]-5H-dibenzo[b, f]azepine-5-carbo hydrazides. Int J
Biol Chem 4:19
Rao A, Chimirri E, Montorte C (2002) Synthesis and anti-HIV
activity of 1-(2,6-difluorophenyl)-1H,3H-thiazolo[3,4-a]benzi-
midazolestructurally-related 1,2-substituted benzimidazoles. Il
Farmaco 57:819
Conclusion
In conclusion, we have demonstrated a simple, facile
convenient, and efficient synthesis of benzo[d]imidazolyl
chromeno[2,3-d]pyrimidones from commercially available
materials and screened for their anti-bacterial, anti-fungal,
and anti-oxidant activities. All the synthesized compounds
7a–e exhibited good anti-bacterial, anti-fungal, and anti-
oxidant activities.
Acknowledgments The authors are thankful to chairman St. Peter’s
Institute of Pharmaceutical Sciences, Vidhyanagar, Hanamkonda,
India, for the facilities and to Director, Indian Institute of Chemical
Technology, Hyderabad, India, for the spectral facility.
Ravindernath A, Srinivas Reddy M (2012) Synthesis, antimicrobial,
anti-inflammatory and antioxidant activity of novel Spiro
(imidazo[40,50:4,50]benzo[1,2-e][1,4] thiazepine)-9,30-indolines.
J Sulfur Chem 3:363
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