Synthesis and in vitro microbiological evaluation of novel 4-aryl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidinones

Article abstract of DOI:10.1016/j.ejmech.2009.09.018

Seven 4-aryl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidin-2(1H)-ones 4a-g and 4-phenyl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidin-2(1H)-thione 4h have been synthesized by a one-pot cyclocondensation of aldehydes, isopropyl acetoacetate and u

Full text of DOI:10.1016/j.ejmech.2009.09.018

European Journal of Medicinal Chemistry 45 (2010) 367–371
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Synthesis and in vitro microbiological evaluation of novel
4-aryl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidinones
*
S. Chitra, D. Devanathan, K. Pandiarajan
Department of Chemistry, Annamalai University, Annamalainagar 608 002, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Seven 4-aryl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidin-2(1H)-ones 4a–g and 4-phenyl-5-
isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidin-2(1H)-thione 4h have been synthesized by a one-
pot cyclocondensation of aldehydes, isopropyl acetoacetate and urea/thiourea in ethanol by using
strontium chloride hexahydrate as the catalyst. All the compounds were screened for their antibacterial
activity against Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and
Salmonella typhi and antifungal activity against Candida albicans, Aspergillus flavus, Rhizopus and Mucor.
Compounds 4b, 4c, 4f, 4g exhibited excellent in vitro antibacterial activity against Staphylococcus aureus,
Salmonella typhi and Pseudomonas aeruginosa and potent in vitro antifungal activity against Candida
albicans, Rhizopus and Mucor. Compound 4f with a nitro group at the para position of the 4-aryl group and
4g with a fluorine at the para position of the 4-aryl group showed more activity than the standard drugs.
Ó 2009 Elsevier Masson SAS. All rights reserved.
Received 2 August 2008
Received in revised form
23 July 2009
Accepted 10 September 2009
Available online 16 September 2009
Keywords:
Dihydropyrimidinones
Antibacterial activity
Antifungal activity
Strontium chloride hexahydrate
1. Introduction
A number of synthetic procedures, reported in the recent years
for preparing dihydropyrimidinones (DHPMs), involve these
3,4-Dihydropyrimidin-2(1H)-ones, named Biginelli compounds,
represent a heterocyclic system of remarkable pharmacological
interest. In the past decades, a broad range of biological effects
including antiviral, antitumor, antibacterial and anti-inflammatory
activities have been described for these compounds [1]. In recent
years, much research has been focused on the synthesis of dihy-
dropyrimidinones (DHPMs), which are important compounds due
to their therapeutic and pharmacological properties. For example,
they can serve as the integral of several calcium channel blockers
reagents and catalysis by protic acids such as H2SO4 adsorbed on
silica gel [8], H3BO3 [9], KHSO4 and PEG-SO3H [10], a variety of Lewis
acids (VCl3 [11], CeCl3$7H2O [12], Cu(OTf)2 [13], LiClO4 [14], LiBr [15]
and Zn(NH2SO3)2 [16]) or recoverable catalysts (KSF clay [17], ion
exchange resins [18], amberlyst-15 [19], heteropolyacids [20,21] and
the poly(4-vinylpyridine-co-divinylbenzene)-Cu(II) complex [22]).
However, in spite of their potential use, some of these methods
involve expensive reagents, strong acidic conditions, long reaction
times, high temperatures, stoichiometric amounts of catalysts and
environmental pollution and give unsatisfactory yields [23].
We wish to report, here, a simple, but effective procedure for
Biginelli’s three component cyclocondensation, producing high
yields of 3,4-dihydropyrimidin-2(1H)-ones byemploying SrCl2.6H2O
as catalyst.
[2], antihypertensive agents [3],
a-1a-antagonists [4] and neuro-
peptide Y (NPY) antagonists [5]. Dihydropyrimidin-2(1H)-ones
have been found as potential antioxidant agents [6]. The aim of this
study is to evaluate the biological activities of some new dihy-
dropyrimidin-2(1H)-ones.
3. Results and discussion
2. Chemistry
3.1. Synthesis
In 1893, Italian Chemist Pietro Biginelli synthesized 3,4-dihy-
dropyrimidin-2(1H)-ones (DHPMs) by heating a mixture of the three
4-Aryl-5-isopropoxycarbonyl-6-methyl-3,4-dihydropyrimidin-
2(1H)-ones 4a–g and 4-phenyl-5-isopropoxycarbonyl-6-methyl-
3,4-dihydropyrimidin-2(1H)-thione 4h were synthesized by heat-
ing a mixture of aldehyde (10 mmol), isopropyl acetoacetate
(10 mmol) and urea/thiourea (15 mmol) in ethanol (20 ml) at 40 ꢀC
in the presence of SrCl2$6H2O (1 mmol, 10 mol%). The reaction
sequence is shown in Scheme 1.
components (aldehyde, b-ketoester and urea) in ethanol containing
a catalytic amount of HCl [7]. However, this procedure suffers from
low yields, long reaction times and harsh reaction conditions.
* Corresponding author. Tel.: þ91 9994469980.
E-mail address: profkpchem@yahoo.co.in (K. Pandiarajan).
0223-5234/$ – see front matter Ó 2009 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2009.09.018

368
S. Chitra et al. / European Journal of Medicinal Chemistry 45 (2010) 367–371
R
CHO
CH3
O
O
O
X
H3C
H3C
SrCl2.6H2O
ethanol,
refluxed
C
O
H
NH
CH3
+
H2N
H
CO
+
NH2
H3C
H3C
R
H3C
X
N
H
4a h
R
X
4a
b
;
;
;
;
;
;
H
O
O
O
4-Cl
2-Cl
4-CH3
4-OCH3
4-NO2
c
d
O
O
O
e
f
g
h
;
;
4-F
H
O
S
Scheme 1. Synthesis of DHPMs catalyzed by SrCl2$6H2O.
Elemental analysis has been done for all these compounds. The
H-3. However, H-3 appeared only as a singlet due to poor resolution
of its signal. In the p-fluoro compound 4g the signal due to H-30 and
H-50 appeared as a triplet due to coupling with 19F.
molecular mass of 4a was confirmed by its mass spectrum. The
physical data are given in Table 1. It is seen that the reaction
proceeds with high yields in all cases.
In the 1H NMR spectrum of 4a there was a singlet at
d
9.15 ppm
which could be attributed to H-1. There was a singlet at
d 7.69 ppm,
which could be attributed to H-3. The aromatic protons appeared as
a multiplet in the range 7.32–7.21 ppm. There was a doublet at
3.2. Analysis of spectra
The compounds were characterized by using IR, 1H NMR and 13
NMR spectra. For 4a the NMR signals were assigned unambiguously
using HMBC spectrum. For 4b the NMR signals were assigned using
HSQC spectrum.
The numbering of atoms in the heterocyclic and aromatic rings
is shown in Fig. 1. The protons are numbered accordingly. Thus, the
proton at C-4 is denoted as H-4. The protons on the nitrogen atoms
N-1 and N-3 are denoted as H-1 and H-3, respectively.
C
d
5.11 ppm, with a J value of 3 Hz, due to the benzylic proton H-4.
There was a multiplet at 4.79 ppm, corresponding to one proton,
due to the methine proton of the isopropyl group. There was
a singlet at 2.23 ppm, corresponding to three protons, due to the
methyl protons at C-6. There were two doublets at 1.14 and
d
d
d
0.96 ppm, each corresponding to three protons, due to the two
diastereotopic methyl protons of the isopropyl group.
The 1H NMR spectra of 4b–h were similar to that of 4a except for
the aromatic protons. The signals were assigned by comparing with
4a. The signals for the aromatic protons were assigned based on
known substituent effects [24].
3.2.1. IR spectra
In all cases two separate sharp bands were observed for the two
different NH-stretching vibrations. One band was observed in the
range 3235–3250 cmꢁ1. The other band was observed in the range
3.2.3. 13C NMR spectra
The observed 13C chemical shifts (
d, ppm) of 4a are as follows:
3120–3170 cmꢁ1
.
164.8 (ester carbonyl carbon), 152.0 (C-6), 148.1 (C-2), 145.0 (C-10),
128.3 (C-30, C-50), 127.2 (C-40), 126.3 (C-20, C-60), 99.5 (C-5), 66.3
(isopropyl methine carbon), 54.0 (C-4), 21.7 (isopropyl methyl
carbon), 21.4 (other isopropyl carbon), 17.7 (methyl carbon at C-6).
All these assignments were made unambiguously based on the
observed correlations in the HMBC spectrum.
The ester carbonyl stretching frequency was observed in the range
1700–1730 cmꢁ1. In compounds 4a–g the amide carbonyl stretching
frequency was observed in the range 1645–1650 cmꢁ1. In 4h the band
due to C]S stretching vibration was observed at 1596 cmꢁ1
.
3.2.2. 1H NMR spectra
In all cases the benzylic proton appeared as a doublet with a J
value of about 3–4 Hz. This is due to the coupling between H-4 and
13C NMR spectra of 4b–g were similar to that of 4a except for the
aromatic carbons. The 13C NMR spectrum of 4h was similar to that
Table 1
Physical data for compounds 4a–h.
Compd.
R
X
Time (hrs)
Molecular formula
M.p. (ꢀC)
Yield (%)
Analysis (%) found (calculated)
C
H
N
4a
4b
4c
4d
4e
4f
H
O
O
O
O
O
O
O
S
2
2
2
2
2
4
2
2
C15H18N2O3
C15H17ClN2O3
C15H17ClN2O3
C16H20N2O3
C16H20N2O4
C15H17N3O5
C15H17FN2O3
C15H18N2O2S
222–223
232–233
224–226
236–238
242–244
196–197
165–166
202–204
96
94
96
92
94
90
94
90
65.80 (65.69)
58.48 (58.35)
58.36 (58.35)
66.60 (66.66)
62.91 (63.15)
56.50 (56.42)
61.55 (61.64)
62.19 (62.06)
6.58 (6.56)
5.49 (5.51)
5.52 (5.51)
6.93 (6.94)
6.59 (6.57)
5.33 (5.32)
5.83 (5.82)
6.18 (6.20)
10.25 (10.21)
9.05 (9.07)
9.04 (9.07)
9.70 (9.72)
9.24 (9.21)
13.15 (13.16)
9.55 (9.58)
9.64 (9.65)
4-Cl
2-Cl
4-CH3
4-OCH3
4-NO2
4-F
4g
4h
H

S. Chitra et al. / European Journal of Medicinal Chemistry 45 (2010) 367–371
369
Table 3
R
In vitro antifungal activity of compounds 4a–h.
Compd.
Minimum inhibitory concentration (MIC) in m
g mlꢁ1
4
'
Candida albicans
Aspergillus flavus
Rhizopus
Mucor
5
'
3
2
'
'
4a
4b
4c
4d
4e
4f
4g
4h
100
25
25
50
50
12.5
12.5
50
–
200
25
50
50
50
12.5
12.5
100
25
200
50
50
50
50
100
100
200
100
25
25
100
50
6
'
1
'
CH3
O
C
12.5
H3C
25
50
50
4
H
C
O
Amphotericin B
25
5
6
3 NH
‘–’ No inhibition even at a higher concentration of 200 m .
g mlꢁ1
H3C
2
1
H3C
X
N
H
P. aeruginosa. It exhibits antibacterial activity against the other
tested organisms only at 200
g mlꢁ1
m
.
However, 4b, in which the hydrogen at the para position of the
Fig. 1. Target compound numbering.
aryl moiety is replaced by chlorine, shows activity against all the
tested organisms in the range of 25–50 m .
g mlꢁ1
Compound 4c, which has the chlorine in the ortho position, has
the same activity as 4b, which has the chlorine in the para position.
Indeed the compounds 4b–g, bearing a substituent in the aryl
group, are more active than the parent compound 4a. Compound
4a and its sulphur analogue 4h have the same activity against E. coli.
However, 4h is more active than 4a against all the other tested
organisms.
of 4a except for the position of C-2. The signals were assigned by
comparing with 4a and based on known effects of the aromatic
substituents [25]. For 4b the assignments were confirmed using the
correlations in the HSQC spectrum.
In the case of 4g coupling with 19F was observed for the aromatic
carbons except C-10.
Only 4f which has a nitro group at the para position and 4g
which has a fluorine group in the para position are more active than
the reference drug Ciprofloxacin.
3.3. Biological activity
All the compounds were screened for their antibacterial activity
against Staphylococcus aureus, Escherichia coli, Klebsiella pneumo-
niae, Pseudomonas aeruginosa and Salmonella typhi and antifungal
activity against Candida albicans, Aspergillus flavus, Rhizopus and
Mucor. Compounds 4b–g with various stubstituents in the aromatic
ring will be useful in understanding the influence of steric and
electronic effects on the biological activity. Comparison of 4a and
4h will tell about the influence of replacement of oxygen by sulphur
on the biological activity.
3.3.2. Antifungal activity
For evaluating antifungal activity Amphotericin B was used as
the standard drug. The observed minimum inhibitory concentra-
tions (MIC) are given in Table 3.
It is seen that 4a shows no activity against A. flavus even at
a concentration of 200
activity against all the other tested fungi in the range of 100–
200
g mlꢁ1
m
g mlꢁ1. However, it shows antifungal
m
.
Compounds 4b–g which have substituents in the 4-aryl group
are more active than the parent compound 4a against all the tested
fungi. The o-chloro compound 4c is less active than the p-chloro
compound 4b against Rhizopus. However, against all the tested
fungi, both 4b and 4c have the same activity. Compound 4h, the
sulphur analogue of 4a, is more active than 4a against all the tested
fungi. Compounds 4f and 4g are more active than the standard drug
Amphotericin B against all the tested organisms.
3.3.1. Antibacterial activity
For evaluating antibacterial activity Ciprofloxacin was used as
the standard drug. The observed minimum inhibitory concentra-
tions (MIC) are given in Table 2. In general all the synthesized
compounds exert a wide range of modest antibacterial activity in
vitro against the tested organisms.
Compound 4a without any substituent in the aryl moiety
exhibits antibacterial activity in vitro at 100 m
g mlꢁ1 against
3.3.3. Influence of aromatic substituents
The results suggest that the antibacterial and antifungal activi-
ties are markedly influenced by the aromatic substituents. Thus, 4b,
4c, 4f and 4g with electron-withdrawing substituents in the
aromatic ring, show greater antibacterial activity than the other
four compounds against all the tested organisms. Also compounds
4f and 4g show greater antifungal activity than all the other
compounds against all the tested organisms. The aromatic
substituents in 4b, 4c, 4f and 4g have positive values [26] for the
Table 2
In vitro antibacterial activity of compounds 4a–h.
Compd.
Minimum inhibitory concentration (MIC) in
m
g mlꢁ1
Staphylococcus Escherichia Klebsiella Pseudomonas Salmonella
aureus
coli
pneumoniae aeruginosa
typhi
4a
4b
4c
4d
4e
4f
200
25
25
100
50
200
50
50
100
100
200
50
50
100
100
100
25
25
50
50
200
25
25
100
50
12.5
12.5
100
50
Hammett substituent constant sp
.
The 4-fluoro compound 4g is the most active compound though
the sp value of F is less than that of the other substituents. Probably
the 4-fluoro compound is able interact with the microorganisms
more effectively due to some intermolecular hydrogen bonding.
Fluorine has the greatest ability to form hydrogen bonding and
hence 4g is more active than the other compounds.
12.5
12.5
50
12.5
12.5
12.5
12.5
50
4g
4h
25
200
25
25
100
25
Ciprofloxacin 25
12.5

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S. Chitra et al. / European Journal of Medicinal Chemistry 45 (2010) 367–371
4. Experimental
66.2 (CH carbon of the isopropyl group), 54.0 (C-4), 21.7 and 21.3
(CH3 carbon of the isopropyl group), 20.6 (CH3 carbon at C-40), 17.7
(CH3 carbon at C-6).
4.1. Chemistry
Melting points were determined in open capillaries and are
uncorrected. 1H and 13C NMR spectra were recorded on a Bruker
AMX 400 spectrometer operating at 400.13 MHz for 1H and
100.62 MHz for 13C in DMSO-d6. HMBC and HSQC spectra were
recorded on a Bruker DRX 500 NMR spectrometer using standard
parameters. The number of data points was 1 K. For recording 1D
NMR spectra solutions were prepared by dissolving about 10 mg of
the compound in 0.5 ml of the solvent and 2D NMR spectra solu-
tions were prepared by dissolving about 50 mg of the compound in
0.5 ml the solvent. IR spectra were recorded in KBr discs on an
Avatar (300 FT-IR) Thermo Nicolet spectrometer. CHN analyses were
performed on an Elementar Vario EL III analyzer. The mass spectrum
of 4a was obtained on a JEOL SX-102 mass spectrometer. Thin layer
chromatography was performed on silica gel G (Merck). Column
chromatography was performed with silica gel (100–200 mesh).
4.2.4. 5-Isopropoxycarbonyl-4-(40-methoxyphenyl)-6-methyl-3,4-
dihydropyrimidin-2(1H)-one 4e
IR (KBr,
n
cmꢁ1): 3243, 3114, 2980, 2929, 1723, 1649, 1463, 1382,
1282, 1178, 788; 1H NMR (
d, DMSO-d6): 8.93 (s, 1H, H-1), 7.34 (s, 1H,
H-3), 7.21 (d, 2H, H-20, H-60), 6.80 (d, 2H, H-30, H-50), 5.21 (s,1H, H-4),
4.88(m,1H, OCH–(CH3)2), 3.76 (s, 3H, OCH3 at C-40), 2.29 (s, 3H, CH3 at
C-6), 1.19 (d, 3H, OCH–(CH3)2), 1.03 (d, 3H, OCH–(CH3)2); 13C NMR (
d,
DMSO-d6): 164.9 (COO),158.3 (C-40),152.5 (C-6),147.0 (C-2),137.0 (C-
10),127.4 (C-20, C-60),113.1 (C-30, C-50),100.1 (C-5), 66.2 (CH carbon of
the isopropyl group), 54.7 (C-4), 53.8 (OCH3 carbon at C-40), 21.6 and
21.3 (CH3 carbon of the isopropyl group), 17.7 (CH3 carbon at C-6).
4.2.5. 5-Isopropoxycarbonyl-6-methyl-4-(40-nitrophenyl)-3,
4-dihydropyrimidin-2(1H)-one 4f
IR (KBr,
n
cmꢁ1): 3247, 3124, 2981, 2945, 1726, 1645, 1464, 1381,
1292, 1180, 780; 1H NMR (
d
, DMSO-d6): 9.27 (s, 1H, H-1), 7.86 (s, 1H,
4.2. Preparation of DHPMs
H-3), 8.19 (d, 2H, H-30, H-50), 7.55 (d, 2H, H-20, H-60), 5.36 (s, 1H,
H-4), 4.88 (m, 1H, OCH–(CH3)2), 2.33 (s, 3H, CH3 at C-6), 1.20 (d, 3H,
A 250 ml round bottomed flask, fitted with a reflux condenser,
was charged with isopropyl acetoacetate (10 mmol), aldehyde
(10 mmol), urea/thiourea (15 mmol), SrCl2$6H2O (1 mmol, 10 mol%)
and EtOH (20 ml). The mixture was heated at 40 ꢀC and the progress
of the reaction was monitored by TLC. After completion of the
reaction (about 2–4 h) the solution was cooled to room temperature
and poured into crushed ice. The resultant solid product was
collected on a Buchner funnel under suction and purified by column
chromatography on silica gel. Elution with 3:2 (benzene:ethyl
acetate) gave the product in the pure form.
OCH–(CH3)2), 1.03 (d, 3H, OCH–(CH3)2); 13C NMR (
d, DMSO-d6):
165.0 (COO), 152.5 (C-40), 152.3 (C-6), 149.3 (C-2), 147.0 (C-10), 128.0
(C-20, C-60), 123.8 (C-30, C-50), 99.0 (C-5), 67.1 (CH carbon of the
isopropyl group), 54.3 (C-4), 22.1 and 21.8 (CH3 carbon of the iso-
propyl group), 18.2 (CH3 carbon at C-6).
4.2.6. 5-Isopropoxycarbonyl-4-(40-fluorophenyl)-6-methyl-3,
4-dihydropyrimidin-2(1H)-one 4g
IR (KBr,
n
cmꢁ1): 3247, 3122, 2981, 2934, 1703, 1649, 1463, 1380,
1291, 1161, 792; 1H NMR (
d
, DMSO-d6): 9.01 (s, 1H, H-1), 7.46 (s, 1H,
H-3), 7.30 (t, 2H, H-20, H-60), 6.99 (t, 2H, H-30, H-50), 5.25 (s,1H, H-4),
4.88 (m, 1H, OCH–(CH3)2), 2.31 (s, 3H, CH3 at C-6), 1.20 (d, 3H, OCH–
4.2.1. 5-Isopropoxycarbonyl-4-(40-chlorophenyl)-6-methyl-3,4-
dihydropyrimidin-2(1H)-one 4b
(CH3)2), 1.02 (d, 3H, OCH–(CH3)2); 13C NMR (
d, DMSO-d6): 164.7
IR (KBr,
n
cmꢁ1): 3249, 3121, 2983, 2934, 1722, 1648, 1462, 1382,
(COO), 161.5 (C-40), 152.3 (C-6), 147.6 (C-2), 140.7 (C-10), 128.0 (C-20,
C-60), 114.5 (C-30, C-50), 99.6 (C-5), 66.2 (CH carbon of the isopropyl
-group), 53.8 (C-4), 21.6 and 21.2 (CH3 carbon of the isopropyl
group), 17.7 (CH3 carbon at C-6).
1291, 1186, 785; 1H NMR (
d, DMSO-d6): 9.11 (s, 1H, H–1), 7.62 (s, 1H,
H-3), 7.30 (s, 4H, Ar–H), 5.21 (d, 1H, H-4), 4.87 (m, 1H, OCH–(CH3)2),
2.29 (s, 3H, CH3 at C-6), 1.19 (d, 3H, OCH–(CH3)2), 1.03 (d, 3H, OCH–
(CH3)2); 13C NMR (
d, DMSO-d6): 164.6 (COO), 152.2 (C-6), 148.0
(C-2), 143.5 (C-10), 132.0 (C-40), 127.9 (C-20, C-30, C-50, C-60), 99.2
(C-5), 66.2 (CH carbon of the isopropyl group), 53.7 (C-4), 21.6 and
21.3 (CH3 carbon of the isopropyl group), 17.8 (CH3 carbon at C-6).
4.2.7. 5-Isopropoxycarbonyl-6-methyl-4-phenyl-3,
4-dihydropyrimidin-2(1H)-thione 4h
IR (KBr,
n
cmꢁ1): 3235, 3170, 2983, 2930, 1703, 1596, 1478, 1379,
1275, 1195, 760, 700; 1H NMR (
d, DMSO-d6): 9.92 (s, 1H, H-1), 9.34 (s,
4.2.2. 5-Isopropoxycarbonyl-4-(20-chlorophenyl)-6-methyl-3,
1H, H-3), 7.35–7.25 (m, 5H, Ar–H), 5.29 (d,1H, H-4), 4.92 (m,1H, OCH–
(CH3)2), 2.36 (s, 3H, CH3 at C-6), 1.21 (d, 3H, OCH–(CH3)2), 1.04 (d, 3H,
4-dihydropyrimidin-2(1H)-one 4c
IR (KBr,
n
cmꢁ1): 3249, 3111, 2979, 2934, 1706, 1650, 1464, 1379,
OCH–(CH3)2); 13C NMR (
d, DMSO-d6): 174.1 (C]S),164.6 (COO),144.1
1285, 1177, 753; 1H NMR (
d, DMSO-d6): 9.14 (s, 1H, H-1), 7.87 (s, 1H,
(C-6), 143.3 (C-10), 128.0 (C-30, C-50), 127.2 (C-40), 126.5 (C-20, C-60),
101.3 (C-5), 66.7 (CH carbon of the isopropyl group), 54.6 (C-4), 21.5
and 21.2 (CH3 carbon of the isopropyl group),17.2 (CH3 carbon at C-6).
H-3), 7.35–7.18 (m, 4H, Ar–H), 5.76 (s, 1H, H-4), 4.81 (m, 1H, OCH–
(CH3)2), 2.37 (s, 3H, CH3 at C-6),1.15 (d, 3H, OCH–(CH3)2), 0.84 (d, 3H,
OCH–(CH3)2); 13C NMR (
d, DMSO-d6): 165.0 (COO), 152.2 (C-6), 149.1
(C-2), 141.6 (C-10), 132.4 (C-20), 129.4 (C-40), 129.0 and 128.8 (C-30,
C-60),127.5 (C-50), 98.7 (C-5), 66.5 (CH carbon of the isopropyl group),
52.0 (C-4), 22.0 and 21.4 (CH3 carbon of the isopropyl group), 18.0
(CH3 carbon at C-6).
4.3. Evaluation of antibacterial activity
The in vitro antibacterial activity of the compounds was tested
in nutrient broth (NB, Hi-media, Mumbai) for bacteria by twofold
serial dilution method [27]. The test compounds were dissolved in
dimethyl sulphoxide (DMSO) to obtain 1 mg/ml stock solutions.
Seeded broth (broth containing microbial spores) were prepared in
NB from 24 hrs old bacterial cultures on nutrient agar (Hi-media,
Mumbai) at 37 ꢂ 1 ꢀC. The colony forming units (cfu) of the seeded
broth were determined by the plating technique and adjusted in
the range of 104–105 cfu/ml. For antibacterial assay the final ino-
culum size was 105 cfu/ml. Testing was performed at 7.4 ꢂ 0.2.
Exactly 0.2 ml of the solution of test compound was added to 1.8 ml
of seeded broth to form the first dilution. One ml of this was diluted
4.2.3. 5-Isopropoxycarbonyl-4-(40-methylphenyl)-6-methyl-3,
4-dihydropyrimidin-2(1H)-one 4d
IR (KBr,
n
cmꢁ1): 3238, 3113, 2976, 2934, 1701, 1645, 1462, 1379,
1288, 1178, 809; 1H NMR (
d
, DMSO-d6): 9.04 (s, 1H, H-1), 7.54 (s, 1H,
H-3), 7.16 (d, 2H, H-20, H-60), 7.08 (d, 2H, H-30, H-50), 5.18 (d, 1H,
H-4), 4.85 (m, 1H, OCH–(CH3)2), 2.29 (s, 3H, CH3 at C-6), 2.27 (s, 3H,
CH3 at C-40), 1.19 (d, 3H, OCH–(CH3)2), 1.03 (d, 3H, OCH–(CH3)2); 13C
NMR (
d, DMSO-d6): 164.8 (COO), 152.5 (C-6), 147.3 (C-2), 141.8
(C-10), 136.2 (C-40), 128.5 (C-30, C-50), 126.2 (C-20, C-60), 100.0 (C-5),

S. Chitra et al. / European Journal of Medicinal Chemistry 45 (2010) 367–371
371
[2] (a) G.C. Rovnyak, S.D. Kinball, B. Beyar, G. Cucinotta, J.D. Dimarco,
J.Z. Gougoutas, A. Hedberg, M. Malley, J.P. McCarthy, R. Zhang, S. Moreland,
J. Med. Chem. 38 (1995) 119–129;
with a further 1 ml of the seeded broth to give the second dilution
and so on until six such dilutions were obtained. A set of assay
tubes containing only seeded broth were kept as controls and
likewise solvent controls were also run simultaneously. The tubes
were incubated in biochemical oxygen demand (BOD) incubators at
37 ꢂ 1 ꢀC for bacteria. The minimum inhibiting concentrations
(MICS) were recorded by visual observations after 24 h. Cipro-
floxacin was used as a standard for the antibacterial study.
(b) K.S. Aswal, G.C. Rovnyak, S.D. Kinball, D.M. Floyd, S. Moreland, B.N. Swanson,
J.Z. Gougoutas, J. Schwartz, K.M. Smillie, M.F. Mallay, J. Med. Chem. 33 (1990)
2629–2635.
[3] (a) K.S. Atwal, B.N. Swanson, S.E. Unger, D.M. Floyd, S. Moreland, A. Hedberg,
B.C. O’Reilly, J. Med. Chem. 34 (1991) 806–811;
(b) G.J. Grover, S. Dzwonczyk, D.M. McMullen, C.S. Normadinam, P.G. Sleph,
S.J. Moreland, J. Cardiovasc. Pharmacol. 26 (1995) 289–294.
[4] (a) D. Nagarathnam, W.C. Wong, S.W. Miao, M.A. Patance, C. Gluchowski, PCT
Int. Appl. WO 97 17 (1997) 969;
4.4. Evaluation of antifungal activity
(b) D.R. Sidler, R.D. Larsen, M. Chartrain, N. Ikemoto, C.M. Roberg, C.S. Taylor,
W. Li, G.F. Bills, PCT Int. WO 99 07 (1999) 695.
[5] M.A. Bruce, G.S. Pointdexter, G. Johnson, PCT Int. Appl. WO 98 33 (1998) 791.
[6] H.A. Stefani, C.B. Oliveira, R.B. Almeida, C.M.P. Pereira, R.C. Braga, R. Cella,
V.C. Borges, L. Savegnago, C.W. Nogueira, Eur. J. Med. Chem. 41 (2006)
513–518.
[7] P. Biginelli, Gazz. Chim. Ital. 23 (1893) 360.
[8] P. Salehi, M. Dabiri, M.A. Zolfigol, L.A.B. Fard, Tetrahedron Lett. 44 (2003)
2889–2891.
[9] S. Tu, F. Fang, C. Miao, H. Jiang, Y. Feng, D. Shi, X. Wang, Tetrahedron Lett. 44
(2003) 6153–6155.
[10] X. Wang, Z. Quan, F. Wang, M. Wang, Z. Zhang, Z. Li, Synth. Commun. 36 (2006)
451–456.
[11] G. Sabitha, G.S.K. Reddy, K.B. Reddy, J.S. Yadav, Tetrahedron Lett. 44 (2003)
6497–6499.
[12] D. Subhas Bose, L. Fatima, H.B. Mereyala, J. Org. Chem. 68 (2003) 587–590.
[13] A.S. Paraskar, G.K. Dewkar, A. Sudalai, Tetrahedron Lett. 44 (2003)
3305–3308.
The in vitro antifungal activity of the compounds was tested in
Sabouraud’s dextrose broth (SDB, Hi-media, Mumbai) for fungi by
twofold serial dilution method [27]. The test compounds were
dissolved in dimethyl sulphoxide (DMSO) to obtain 1 mg/ml stock
solutions. Seeded broth (broth containing microbial spores) was
prepared by suspending 24 h to 7-days old Sabouraud’s agar slant
cultures in SDB. The colony forming units (cfu) of the seeded broth
were determined by the plating technique and adjusted in the
range of 104–105 cfu/ml. For antifungal assay the final inoculum
size was 1.1–1.5 ꢃ102 cfu/ml. Testing was carried out by the same
procedure as done for antibacterial studies except the temperature
which was maintained at 28 ꢂ 1 ꢀC for about 72–96 h of incubation.
Amphotericin B was used as a standard for the antifungal study.
[14] J.S. Yadav, B.V.S. Reddy, R. Srinivas, C. Venugopal, T. Ramalingam, Synthesis 9
(2001) 1341–1345.
[15] G. Maiti, P. Kundu, C. Guin, Tetrahedron Lett. 44 (2003) 2757–2758.
[16] M. Zhang, Y. Li, Synth. Commun. 36 (2006) 835–841.
[17] A.K. Mitra, K. Banerjee, Synlett (2003) 1509–1511.
[18] A. Dondoni, A. Massi, Tetrahedron Lett. 42 (2001) 7975–7978.
[19] J.S. Yadav, B.V.S. Reddy, E.J. Reddy, T. Ramalingam, J. Chem. Res. Synop.
7 (2000) 354–355.
Acknowledgement
The authors are thankful to SIF, Indian Institute of Science,
Bangalore for recording all the NMR spectra. The authors are
thankful to SAIF, Central Drug Research Institute, Lucknow and
CECRI, Council of Scientific and Industrial Research, Karaikudi,
Tamil Nadu for providing elemental analysis data. The authors are
also thankful to Faculty of Medicine, Department of Microbiology,
Annamalai University for carrying out the antimicrobial activities.
[20] M.M. Heravi, F.F. Bakhtiari, F.F. Barmoharram, Catal. Commun.
373–376.
7 (2006)
[21] E. Rafiee, F. Shahbazi, J. Mol. Catal. A 250 (2006) 57–61.
[22] R.V. Yarapathi, S. Kurva, S. Tammishetti, Catal. Commun. 5 (2004) 511–513.
[23] M.M. Amini, A. Shaabani, A. Bazgir, Catal. Commun 7 (2006) 843–847.
[24] R.M. Silverstein, F.X. Webster, Spectrometric Identification of Organic
Compounds, sixth ed. Wiley, New York, 1998, p. 209.
[25] R.M. Silverstein, F.X. Webster, Spectrometric Identification of Organic
Compounds, sixth ed. Wiley, New York, 1998, p. 229.
[26] J. March, Advanced Organic Chemistry, fourth ed. John Wiley & Sons Pvt. Ltd.,
Singapore, 2005, p. 280.
[27] M.H. Dhar, M.M. Dhar, B.N. Dhawan, B.N. Mehrotra, C. Ray, Indian J. Exp. Biol. 6
(1968) 232–236.
References
[1] (a) C.O. Kappe, Tetrahedron 49 (1993) 6937–6963;
(b) C.O. Kappe, Eur. J. Med. Chem. 35 (2000) 1043–1052;
(c) C.O. Kappe, Acc. Chem. Res. 33 (2000) 879–888.