Synthesis and biological studies of a novel series of 4-(4-(1H-imidazol-1- yl)phenyl)-6-arylpyrimidin-2-amines

Article abstract of DOI:10.1007/s00044-013-0523-6

A novel series of eleven 4-(4-(1H-imidazol-1-yl)phenyl)-6-arylpyrimidin-2- amines has been prepared from synthesized 3-[4-(1H-imidazol-1-yl) phenyl]prop-2-en-1-ones and evaluated for phosphodiesterase (PDE) inhibition and antimicrobial activities. N-aryla

Full text of DOI:10.1007/s00044-013-0523-6

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-013-0523-6
ORIGINAL RESEARCH
Synthesis and biological studies of a novel series
of 4-(4-(1H-imidazol-1-yl)phenyl)-6-arylpyrimidin-2-amines
•
Mujahid Hussain Bukhari Matloob Ahmad
•
•
Tanvir Hussain Syed Umar Naveed Ahmad
•
Received: 14 August 2012 / Accepted: 29 January 2013
Ó Springer Science+Business Media New York 2013
Abstract A novel series of eleven 4-(4-(1H-imidazol-1-
yl)phenyl)-6-arylpyrimidin-2-amines has been prepared
from synthesized 3-[4-(1H-imidazol-1-yl) phenyl]prop-2-
en-1-ones and evaluated for phosphodiesterase (PDE)
inhibition and antimicrobial activities. N-arylation of
imidazole with 4-fluorobenzaldehyde using hexadecyl-
trimethylammonium bromide as catalyst gave 4-(1H-imi-
dazol-1-yl) benzaldehyde which on treatment with
substituted acetophenones yielded corresponding chalcones
(1a–1k). Each chalcone on further reaction with guanidine
hydrochloride resulted in title compounds (2a–2k). Pyrim-
idines thus synthesized were subjected to biological studies.
Some compounds showed marked activities in PDE inhibi-
tion and anti-bacterial and anti-fungal bioassays.
in cell function by regulating intracellular levels of cyclic
adenine monophosphate (cAMP) and cyclic guanine
monophosphate (cGMP). Theophylline and Papaverine
have historically been used as drugs and are known to be
weak inhibitors of PDE. The discovery of several isoen-
zyme families provides a pace for the development of
isoenzyme selective inhibitors for the treatment of various
ailments. The role of PDE3 inhibitors for congestive heart
failure (Barnes et al., 1988; Lugnier, 2006; Nicholson
et al., 1991), PDE4 inhibitors for inflammatory airways
(Muller et al., 1996; Torphy and Undem, 1991) and most
successfully, PDE5 inhibitors for erectile dysfunction
(Murray, 1993) is widely recognised. The structure of one
of the famous PDE5 inhibitors, Sildinafil and its analogues
is based on pyrimidine scaffold. (Toque et al., 2008)
Pyrimidine ring system is present in pyrimidine and
purine bases of DNA and RNA. In purines, both pyrimidine
as well as imidazole rings are fused together. Pyrimidine
derivatives are reported as highly potentially biologically
active compounds including antimicrobial (Ballell et al.,
2007; Rao et al., 2003), anticancer (Miyazaki et al., 2005),
anti-inflammatory and analgesic (Breault and Pease, 2000;
Venu et al., 2008; Zienab et al., 2011), anti-HCV (Cha-
makura et al., 2007), anti-HIV (Malik et al., 2006), anti-
oxidant (Biagi et al., 1996), anti-aging (Bbizhayev, 2006)
and several others. A variety of synthetic imidazole
derivatives themselves are extensively used as amoebicidal
(Metronidazole or FlagylÒ, Tinidazole, Timorazole), anti-
fungal (Emami et al., 2008) (Miconazole, Ketoconazole),
anti-thyroid (CarbizoleÒ), anti-ulcer (Omeprazole, Cimet-
idine, e.g. CimetÒ), anxiolytic (LoprazolamÒ) and several
other drugs available in the market. Recent research shows
that chalcones based on imidazole scaffold exhibit anti-
oxidant, anti-fungal and anti-leishmanial activities (Huss-
ain et al., 2009).
Keywords Arylpyrimidin-6-amine Á Phosphodiesterases Á
N-arylation Á Anti-fungal
Introduction
Phosphodiesterases (PDEs) form a unique class of enzymes
that hydrolyse cyclic nucleotides and thus play a vital role
This work is dedicated to our beloved teacher, the Late Professor Dr.
Hamid Latif Siddiqui, University of the Punjab, Lahore-Pakistan.
M. H. Bukhari (&) Á T. Hussain Á S. Umar Á N. Ahmad
Institute of Chemistry, University of the Punjab,
Lahore 54590, Pakistan
e-mail: mujahid_bk@yahoo.com
M. Ahmad
Department of Chemistry, Government College University,
Faisalabad 38000, Pakistan
123

Med Chem Res
As both the ring systems play a vital role in our life, we
planned to study their synergic effect as PDE inhibitors and
as anti-bacterial and anti-fungal agents.
analysis which was found in accordance to the calculated
values (Scheme 1) (Hussain et al., 2009).
Characterisation
Results and discussion
All the pyrimidines were characterized with the help of
NMR, IR and MS spectral studies and CHN analysis. The IR
spectra of each compound synthesized showed characteristic
absorptions for NH2, N–C, N=C and aromatic functionalities
present in each compound. Absorption at *1550 cm-1 is
characteristic for pyrimidine skeleton. A characteristic peak
Chemistry
Synthesis of 3-[4-(1H-imidazol-1-yl) phenyl]prop-2-
en-1-ones (1a–1k)
1
in the range of d 6.67–7.08 ppm in H-NMR as a broad
Imidazole was N-arylated with 4-fluorobenzaldehyde in the
presence of hexadecyltrimethylammonium bromide as a
catalyst. 4-(1H-Imidazol-1-yl) benzaldehyde thus synthe-
sized was then treated with various substituted acetophe-
nones using 10 % NaOH solution in MeOH to get the
corresponding chalcones (1a–1k).
singlet integrated for two protons was assigned to NH2 pro-
tons (Varga et al., 2003). A singlet was assigned to imidazole
H-2 at near d 7.15 ppm. Another singlet, in most cases was
differentiable in the aromatic region which may be assigned
to H-5 of pyrimidine ring. Table 1
Biological activities of compounds 2a–2k
Synthesis of 4-(4-(1H-imidazol-1-yl)phenyl)-6-
arylpyrimidin-2-amines (2a–2k)
Phosphodiesterase inhibition activity of compounds 2a–2k
Each chalcone was treated with guanidine hydrochloride
and 50 % aqueous KOH solution in EtOH at reflux tem-
perature followed by portion wise addition of 30 % H2O2
solution under the same conditions (Scheme 1) (Varga
et al., 2003). The reaction was frequently monitored
visualising TLC, and after completion of reaction, precip-
itates formed were filtered, dried and thoroughly washed
with cold MeOH and then cold water. The semi-pure
product was purified by column chromatography using
CHCl3/MeOH (4:1) solvent system. Each title compound
was characterized by spectral studies and elemental
It has been reported that Imazodan CI-914 (I), CI-930 (II),
and related compounds 4,5-dihydro-6-[4-(1H-imidazol-1-
yl)phenyl]-3(2H)-pyridazinones, have positive inotropic
activity(Bristol et al., 1984; Sircar et al., 1985). Series of
heterocyclic systems have been investigated for their ino-
tropic activity. These studies revealed the contribution of
the (IH-imidazol-1-yl)phenyl moiety for superior inotropic
activity in comparison with other more conventional aro-
matic substituents. Keeping in view this fact, we studied
the phosphodiesterase inhibition activity of the compounds.
The results are shown in Table 2.
Scheme 1 Synthesis of 4-(4-
DMF
N
N
≡ Ar1
CHO
N
(1H-imidazol-1-yl)phenyl)-6-
arylpyrimidin-2-amines (2a–2k)
from 3-[4-(1H-imidazol-1-yl)
phenyl]prop-2-en-1-ones
(1a–1k)
+
NH
F
CHO
CHO
(i)
Ar2 COCH3
(ii)
Ar1
Ar2
O
(iii)
(iv)
Ar1
Ar2
N
N
NH2
(1a-1k)
(2a-2k)
(i)
K2CO3/ C16H33(CH3)3N+Br- , 100°C
MeOH/10% NaOH/ room temp.
(NH2)2C= NH.HCl/ EtOH/ reflux
30% H2O2/ reflux
(ii)
(iii)
(iv)
123

Med Chem Res
alternata, Aspergillus flavus and Aspergillus fumigatus
procured from Biofertilizers and Biopesticide Laboratory,
Institute of Mycology & Plant Pathology, University of the
Punjab Lahore, Pakistan. The results are summarized in
Table 4.
Table 1 General description of 3-[4-(1H-imidazol-1-yl) phenyl]-
prop-2-en-1-ones (1a–1k) and 4-(4-(1H-imidazol-1-yl)phenyl)-6-
arylpyrimidin-2-amines (2a–2k)
Compounds Ar2
Compounds Ar2
1a
1b
1c
1d
1e
1f
4-chlorophenyl
2a
2b
2c
2d
4-chlorophenyl
4-bromophenyl
4-methoxyphenyl
3-methoxyphenyl
4-bromophenyl
4-methoxyphenyl
3-methoxyphenyl
3,4-dimethoxyphenyl
2,4-dichlorophenyl
4-fluorophenyl
Conclusion
3,4-dimethoxyphenyl 2e
In short, we evaluated 4-(4-(1H-imidazol-1-yl)phenyl)-6-
arylpyrimidin-2-amines (2a–2k) for their antimicrobial and
Phosphodiesterase Inhibition Activity. These compounds
are structural hybrids of two excellent bioactive heterocy-
cles i.e. imidazole and pyrimidine. Thus, it is proposed that
the title compounds may possess the biological activities of
parent ring systems and could be used as template for
further discoveries.
2,4-dichlorophenyl
4-fluorophenyl
2f
1g
1h
1i
2g
2,3,4-trichlorophenyl 2h
2,3,4-trichlorophenyl
2,5-dichlorophenyl
4-iodophenyl
2,5-dichlorophenyl
4-iodophenyl
Phenyl
2i
1j
2j
2k
1k
Phenyl
Table 2 Phosphodiesterase inhibition activity of compounds (2a–2k)
Compound
Conc. (mM)
% Inhibition
2a
2b
2c
2d
2e
2f
0.2
0.1
0.2
0.1
0.1
0.2
0.1
0.2
0.1
0.1
0.2
Negative
9.2
Experimental
Conversion of imidazole to 4-(4-(1H-imidazol-1-
yl)phenyl)-6-arylpyrimidin-2-amines
12.0
22.4
Negative
Negative
Negative
12.0
Procedure for synthesis of 4-(1H-imidazol-1-
yl)benzaldehyde
2g
2h
2i
A mixture of imidazole (3.40 g, 50 mmol), anhydrous
potassium carbonate (6.90 g, 50 mmol), 4-fluorobenzalde-
hyde (6.2 g, 50 mmol), hexadecyltri-n-butylphosphonium
bromide (10 mg) and dimethylformamide (30.0 mL) was
stirred for a period of 15 h at 100 °C. The contents were
poured onto ice cold water (100 mL) after cooling to room
temperature. Pale yellow precipitates obtained were fil-
tered, dried and crystallized from methanol. Yield: 78.3 %;
mp 151–153 °C (Hussain et al., 2009).
Negative
11.7
2j
2k
Negative
Anti-bacterial activities
The compounds (2a–2k) were subjected to anti-bacterial
studies. Some compounds exhibited moderate activity
against Bacillus subtillis, Escherichia coli and Staphylo-
coccus aureus. Particularly, compound 2a, 2h and 2j
showed significant activity against E. coli. Compound 2j
exhibited marked activity also against S. aureus and B.
subtillis. This compound with p-iodophenyl moiety may be
undertaken for further biological investigations. All the
compounds showed insignificant activity against Shigella
flexenari, Pseudomonas aeruginosa and Salmonella typhi.
The results of anti-bacterial activity are summarized in
Table 3.
Procedure for synthesis of 3-(4-(1H-imidazol-1-yl)
phenyl)prop-2-en-1-ones (1a–1k)
All chalcones were synthesized according to the procedure
which has already been reported by us (Hussain et al.,
2009). A solution of NaOH (40 %; 10.0 mL) was added
drop wise to a mixture of 4-(1H-imidazol-I-yl)benzalde-
hyde (10.0 mmol, 1.72 g), corresponding substituted ace-
tophenone (10.0 mmol) and methanol (50 mL) over a
period of 30–40 min with continuous stirring at ambient
temperature till completion of the reaction monitored fre-
quently by TLC. Precipitates thus obtained, were filtered,
washed with cold MeOH followed by cold water. Finally,
recrystallization from MeOH gave the title compound.
Antifungal activities
The synthesized compounds were tested for anti-fungal
activities against three fungal species viz. Alternaria
123

Med Chem Res
Table 3 Anti-bacterial activity
(zone of inhibition in mm) of
compounds (2a–2k)
Compound
Bacillus
subtillis
Escherichia
coli
Staphylococcus
aureus
Shigella
flexneri
Pseudomonas
aeruginosa
Salmonella
typhi
2a
12
13
14
_
18
12
12
13
15
14
16
18
12
20
11
35
_
11
12
11
_
_
_
_
_
_
_
_
_
_
_
_
_
25
2b
12
_
_
2c
11
12
10
_
2d
14
13
11
_
2e
_
_
2f
_
10
12
11
12
_
2g
_
_
2h
15
12
21
13
30
15
_
_
2i
_
2j
16
_
_
2k
_
_
Imipenem
30
30
31
of I hr. The ethanol was removed under reduced pressure in
a rotary evaporator and distiled water (*20 mL) was
added to the residue. The product was easily isolated as
precipitates and was washed repeatedly with pure water.
The still crude product was recrystallized from ethanol
and was dried finally in a vacuum desiccator over P2O5/
KOH.
Table 4 Anti-fungal activities of compounds (2a–2k)
Compounds
MIC (mg/mL)
Alternaria
alternata
Aspergillus
fumigatus
Aspergillus
flavus
2a
1.0
[1.0
[1.0
1.0
1.0
1.0
1.0
1.0
2b
Compound 2a: (52 %, Pale yellow crystals), m.
p. 252–254 °C. IR (KBr) mmax cm-1: 3328, 3205, 1690,
1546, 675; 1H NMR (400 MHz, DMSO-d6) d: 6.81 (2H, br.
s, NH2), 7.14 (1H, s, H-2 imidazole), 7.56–7.60 (3H, m,
ArH), 7.81–7.87 (4H, m, ArH), 8.27 (2H, d, J = 8.4 Hz,
Ar2H-3 ? Ar2H-5), 8.36–8.39 (2H, m, Ar2H-2 ? Ar2H-6).
MS m/z: 347.09 (M?). 13C NMR: 100.8, 115.5, 115.7,
120.9, 126.0, 126.2 126.4, 126.6, 128.9 130.2, 130.4,
131.6, 133.5, 134.0, 135.1, 135.3, 136.8, 164.4, 167.2.
Anal. Calc. for C19H14ClN5; C, 65.61; H, 4.06; N, 20.14:
Found: C, 65.64; H, 4.05; N, 20.15.
2c
1.0
[1.0
1.0
2d
[1.0
[1.0
[1.0
0.50
[1.0
[1.0
1.0
2e
[1.0
1.0
1.0
2f
[1.0
1.0
2g
1.0
2h
[1.0
[1.0
1.0
[1.0
[1.0
1.0
2i
2j
2k
[1.0
0.25
1.0
1.0
Mancozeb
0.25
0.25
4-(4-(1H-imidazol-1-yl)phenyl)-6-(4-
bromophenyl)pyrimidin-2-amine (2b)
A typical procedure for the synthesis of 4-(4-(1H-
imidazol-1-yl)phenyl)-6-arylpyrimidin-2-amines
(2a–2k)
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(4-bromophenyl)prop-2-en-1-one.
Compound 2b: (46 %, Yellow powder), m. p. 225–
227 °C, IR (KBr) mmax cm-1: 3310, 3195, 1680, 1532, 670.
1H NMR (400 MHz, DMSO-d6) d: 7.08 (2H, br. s, NH2),
7.26 (3H, d, J = 5.6 Hz), 7.50–7.58 (6H, m, ArH), 7.71
(1H, s, H-5), 8.21–8.25 (2H, m, ArH). 13C NMR: 101.6,
115.6, 115.8, 120.7, 122.9, 125.1, 125.4, 125.6, 125.8,
130.4, 133.3, 133.6, 133.9 134.6, 135.4, 136.5, 162.4,
164.1, 167.1. MS m/z: 391.04 (M?). Anal. calc. for
C19H14BrN5; C, 58.18; H, 3.60; N, 17.85; Found: C, 58.20;
H, 3.61; N, 17.82.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(4-
chlorophenyl)pyrimidin-2-amine (2a)
3-(4-(1H-Imidazol-1-yl) phenyl)-1-(4-chlorophenyl)prop-
2-en-1-one (2.8 g, 9.08 mmol), guanidine hydrochloride
(1.3 g, 1.5 mmol), ethanol (20 mL) and 50 % aqueous
KOH solution (4 mL,) were mixed together, then heated up
and stirred at reflux temperature for 1 h. Under the same
conditions, 30 % aqueous H2O2 (3.1 mL, 27.3 mmol) was
added to the above mixture in small portions over a period
123

Med Chem Res
4-(4-(1H-imidazol-1-yl)phenyl)-6-(4-methoxyphenyl)-
pyrimidin-2-amine (2c)
C21H19N5O2; C, 67.55; H, 5.13; N, 18.76; Found: C, 67.55;
H, 5.13; N, 18.75.
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(4-methphxyenyl)prop-2-en-1-one.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(2,4-dichlorophenyl)-
pyrimidin-2-amine (2f)
Compound 2c: (55 %, Whitish amorphous solid), m.
p. 211–213 °C. IR (KBr) mmax cm-1: 3380, 3086, 1645,
1545, 688. 1H NMR (400 MHz, DMSO-d6) d: 3.82 (3H, s,
OCH3), 6.78 (2H, br. s, NH2), 7.05 (1H, d, J = 6.8 Hz,
ArH), 7.16 (1H, s, Ar1H-2, imidazole), 7.77 (1H, s, H-5),
7.81–7.84 (4H, m, ArH), 7.89 (2H, d, J = 8.4 Hz, ArH),
8.03–8.07 (3H, m, ArH). 13C NMR: 23.5, 102.2, 115.5,
115.9, 118.5, 123.5, 124.0, 129.2 (2C), 130.1, 130.3, 131.2,
132.1, 133.1, 135.1, 135.6, 136.5, 161.2, 165.0, 168.5. MS
m/z: 343.14 (M?). Anal. calc. for C20H17N5O; C, 69.96; H,
4.99; N, 20.40; Found: C, 69.95; H, 4.98; N, 20.38.
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(2,4-dichlorophenyl)prop-2-en-1-one.
Compound 2f: (54 %, Yellowish solid), m. p. 224–
226 °C. IR (KBr) mmax cm-1: 3428, 3110, 1680, 1570, 655.
1H NMR (400 MHz, DMSO-d6) d: 6.89 (2H, br. s, NH2),
7.13 (1H, s, H-2 imi), 7.41 (1H, s, Ar2H-3), 7.56 (1H, d,
J = 7.6 Hz, Ar2H-5), 7.64 (1H, d, J = 8.0 Hz ArH),
7.77–7.85 (4H, m, ArH), 8.25 (2H, d, J = 8.0 Hz, Ar1H-
2 ? Ar1H-6), 8.37 (1H, s, H-5). 13C NMR: 102.9, 115.6,
115.9, 119.5, 124.4, 125.2, 129.3, 129.5, 130.8, 131.1, 131.9,
132.1, 135.1, 135.6, 136.5, 162.7, 164.2, 165.9, 168.1. MS
m/z: 381.05 (M?). Anal. calc. for C19H13Cl2N5; C, 59.70; H,
3.43; N, 18.32: Found: C, 59.72; H, 3.40; N, 18.35.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(3-methoxyphenyl)-
pyrimidin-2-amine (2d)
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(3-methphxyenyl)prop-2-en-1-one.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(4-fluorophenyl)-
pyrimidin-2-amine (2g)
Compound 2d: (44 %, Yellow amorphous solid), m.
p. 232–234 °C. IR (KBr) mmax cm-1. 3390, 3080, 1635,
1533, 690. 1H NMR (400 MHz, DMSO-d6) d: 3.84 (3H, s,
OCH3), 6.82 (2H, br. s, NH2), 7.15 (1H, d, J = 8.4 Hz,
ArH), 7.19 (1H, s, Ar1H-2, imidazole), 7.67 (1H, s, ArH),
7.76–7.80 (3H, m, ArH), 7.84 (1H, s, ArH), 7.93 (2H, d,
J = 8.4 Hz, ArH), 8.01–8.03 (3H, m, ArH). 13C NMR:
21.6, 101.2, 114.5, 114.9, 118.5, 123.8, 124.0, 128.2,
128.5, 129.2, 130.1, 132.1, 133.1, 135.1, 135.6, 136.5,
138.1, 162.2, 164.0, 167.1. MS m/z: 343.14 (M?). Anal.
calc. for C20H17N5O; C, 69.96; H, 4.99; N, 20.40; Found:
C, 69.95; H, 4.98; N, 20.38.
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(4-fluorophenyl)prop-2-en-1-one.
Compound 2g: (63 %, Yellow powder), m. p. 255–
256 °C. IR (KBr) mmax cm-1: 3446, 3100, 1670, 1580, 685.
1H NMR (400 MHz, DMSO-d6) d: 6.77 (2H, br. s, NH2),
7.14 (1H, s, H-2 imidazole), 7.35 (2H, m, Ar2H-3 ? Ar2H-5),
7.77–7.83 (3H, m, ArH), 7.87 (1H, s, H-5), 8.29–8.32 (2H, m,
ArH), 8.36–8.39 (3H, m, ArH). 13C NMR: 100.4, 115.6,
116.9, 118.5, 118.7, 120.4, 127.2, 127.4, 129.2, 130.1, 130.3,
131.2, 135.1, 135.6, 136.5, 163.2, 164.0, 165.6, 167.8. MS
m/z: 331.12 (M?). Anal. calc. for C19H14FN5; C, 68.87; H,
4.26; N, 21.14: Found: C, 68.91; H, 4.27; N, 21.11.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(3,4-dimethoxyphenyl)-
pyrimidin-2-amine (2e)
4-(4-(1H-imidazol-1-yl)phenyl)-6-(2,3,4-trichlorophenyl)-
pyrimidin-2-amine (2h)
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(3,4-dimethoxyphenyl)prop-2-en-1-one.
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(2,3,4-trichlorophenyl)prop-2-en-1-one.
Compound 2e: (55 %, Whitish amorphous solid), m.
p. 318–320 °C. IR (KBr) mmax cm-1: 3410, 3090, 1640,
1545, 692. 1H NMR (400 MHz, DMSO-d6) d: 3.83 (3H, s,
OCH3), 3.87 (3H, s, OCH3), 6.67 (2H, br. s, NH2), 7.08
(1H, d, Ar2H-5), 7.14 (1H, s, Ar1H-2, imidazole), 7.72 (1H,
s, H-5), 7.79–7.82 (3H, m, ArH), 7.85–7.88 (2H, m, ArH),
8.34–8.38 (3H, m, ArH). 13C NMR: 19.6, 21.5, 100.2,
115.5, 115.9, 118.5, 123.1, 123.3, 128.3, 128.5, 129.2,
129.9, 130.1, 131.1, 135.1, 135.6, 137.0, 139.1, 162.2,
164.1, 167.0. MS m/z: 373.15 (M?). Anal. calc. for
Compound 2h: (46 %, Bright yellow powder, m.
p. 306–308 °C. IR (KBr) mmax cm-1: 3428, 3110, 1680,
1570, 655. 1H NMR (400 MHz, DMSO-d6) d: 6.94 (2H, br.
s, NH2), 7.13 (1H, s, H-2 imidazole), 7.41 (1H, s, H-5),
7.56–7.65 (2H, m, ArH), 7.77–7.85 (3H, m, ArH), 8.27
(2H, d, J = 8.0 Hz, ArH), 8.37 (1H, d, J = 6.8 Hz, ArH).
13C NMR: 100.5, 115.5, 115.7, 119.4, 129.2, 129.3, 129.7,
130.5, 131.1, 132.1, 133.5, 134.6, 135.4, 135.6, 135.8,
138.6, 161.1, 162.2, 166.7. MS m/z: 415.02 (M?). Anal.
123

Med Chem Res
calc. for C19H12Cl3N5; C, 54.77; H, 2.90; N, 16.81; Found:
C, 54.80; H, 2.94; N, 16.80.
m/z: 313.13 (M?). Anal. calc. for C19H15N5; C, 72.83; H,
4.82; N, 22.35: Found: C, 72.83; H, 4.82; N, 22.38.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(2,5-dichlorophenyl)-
pyrimidin-2-amine (2i)
Phosphodiesterase inhibition assay
Activity against snake venom was determined by taking
0.66 mM bis-(p-nitrophenyl) phosphate (Sigma N-3002) as
substrate and 66 mM Tris–HCl buffer of pH 8.8, 60 mM
Mg-acetate with final concentration of 0.0001484 U/well of
enzyme using a microtitre plate assay. EDTA and Cystein
(Merck) were used as positive controls (IC50 = 274
0.007 lM, 748 0.015 lM, respectively). Enzyme activ-
ity was monitored by spectrophotometer at 410 nm after
30-min pre-incubation of the enzyme with test samples, at
37 °C on a microtitre plate reader (SpectraMax, Molecular
Devices) by following change in O.D/min (rate) of release
of p-nitrophenol from p-nitrophenyl phosphate. All assays
were conducted in triplicate (Goding et al., 1998; Johnson
et al., 2001).
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(2,5-dichlorophenyl)prop-2-en-1-one.
Compound 2i: (46 %, Whitish solid), m. p.[340 °C. IR
1
(KBr) mmax cm-1: 3300, 3195, 1670, 1565, 680. H NMR
(400 MHz, DMSO-d6) d: 6.88 (2H, br. s, NH2), 7.14 (1H, s,
H-2 imidazole), 7.48 (1H, s, H-5), 7.54–7.58 (3H, m, ArH),
7.77–7.85 (3H, m, ArH), 8.23 (2H, d, J = 8.4 Hz, ArH),
8.31 (1H, s, Ar2H-6). 13C NMR: 101.2, 116.3, 116.4, 121.3,
128.5, 128.6, 128.7, 129.9, 130.1, 130.4, 132.0, 132.6,
135.6, 135.8, 136.2, 138.1, 163.1, 164.0, 166.2. MS m/z:
381.05 (M?). Anal. calc. for C19H13Cl2N5; C, 59.70; H,
3.43; N, 18.32: Found: C, 59.72; H, 3.40; N, 18.35.
4-(4-(1H-imidazol-1-yl)phenyl)-6-(4-iodophenyl)-
pyrimidin-2-amine (2j)
Anti-bacterial activity
All the synthesized pyrimidines were tested against six
bacterial species viz. S. aureus, E. coli, B. subtilis, S. flexe-
nari, P. aeruginosa and S. typhi. Each compound (dissolved
in DMSO) was subjected to anti-bacterial screening for
determining the zone of inhibition by well diffusion method.
The Petri plates were inoculated in cultures of bacteria on
potato dextrose agar medium. Plates were incubated at
37 °C for 24 h for bacteria. After inoculation, the diameter
of clear zone of inhibition surrounding the sample was taken
as a measure of the inhibitory power of the sample against
the particular test organism. (Gaulejac et al., 1999)
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1-(4-iodophenyl)prop-2-en-1-one.
Compound 2j: (68 %, Off white powder) m.
p. 189–191 °C. IR (KBr) mmax cm-1: 3310, 3180, 1680, 1584,
685. 1H NMR (400 MHz, DMSO-d6) d: 6.79 (2H, br. s, NH2),
7.14 (1H, s, H-2 imidazole), 7.27 (1H, d, J = 8.5 Hz, ArH),
7.36 (1H, d, J = 8.0 Hz, ArH), 7.50 (2H, d, J = 8.0 Hz,
Ar1H-3 ? Ar1H-5), 7.70–7.74 (2H, m, Ar1H-2 ? Ar1H-6),
7.79–7.91 (2H, m, Ar2H-3 ? Ar2H-5), 8.22 (1H, s, H-5),
8.31–8.39 (2H, m, Ar2H-2 ? Ar2H-6). 13C NMR: 98.2, 102.1,
115.6, 115.8, 119.0, 128.2, 128.3, 129.3, 129.5, 130.3, 134.5,
135.3, 135.5, 137.8, 138.1, 138.3, 163.5, 165.1, 168.0. MS
m/z: 439.03 (M?). Anal. calc. for C19H14IN5; C, 51.95; H,
3.21; N, 15.94; Found: C, 51.97; H, 3.21; N, 15.92.
Antifungal activity
Three fungal species viz. A. alternata, A. flavus and
A. fumigatus were procured from Biofertilizers and Bio-
pesticide Laboratory, Institute of Mycology & Plant
Pathology, University of the Punjab, Lahore, Pakistan.
Pure cultures of A. alternata, A. flavus and A. fumigatus
were prepared in Petri plates using agar medium of malt
extract which was autoclaved at 121 °C for 60 min. Then it
was poured to plates and after its setting, media was
inoculated with all fungal strains and incubated for 5 days
at 37 °C to obtain pure cultures without any contamination.
2 % malt extract (ME) broth was autoclaved at 121 °C
for 30 min and cooled at room temperature. 6 mg of each
of the synthetic compounds was dissolved in 0.7 mL
dimethyl sulfoxide (DMSO). Appropriate quantity of ME
broth was added to make the volume 6 mL. Lower con-
centrations of these stock solutions (1 mg/mL) viz. 0.500,
0.250 and 0.125 mg/mL were prepared by double dilution.
4-(4-(1H-imidazol-1-yl)phenyl)-6-phenylpyrimidin-2-
amine (2k)
The experimental procedure was similar to that described
for compound 2a starting from 3-(4-(1H-imidazol-1-yl)
phenyl)-1phenylprop-2-en-1-one.
Compound 2k: (36 %, Cream coloured powder), m.p.
210–212 °C. IR (KBr) mmax cm-1: 3305, 3165, 1655, 1556,
1
674. H NMR (400 MHz, DMSO-d6) d: 7.08 (2H, br. s,
NH2), 7.27 (2H, m, ArH), 7.36 (2H, m, ArH),7.49–7.56 (4H,
m, ArH), 7.70 (1H, s, H-5), 8.21 (2H, m, Ar2H-3 ? Ar2H-5),
8.29 (2H, m, Ar2H-2 ? Ar2H-6). 13C NMR: 100.7, 115.3,
115.5, 120.5, 129.4, 129.6, 130.2(2C), 131.9, 132.0, 132.2,
132.4, 136.5, 136.7, 140.1, 140.4, 163.5, 164.1, 166.9. MS
123

Med Chem Res
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Control treatments were similarly prepared without the
addition of compounds. A commercial fungicide mancozeb
was used as a reference compound.
One-milliliter solution of each of the concentration of
the synthetic compounds, mancozeb and control was
poured into a sterilized 5 mL culture tube. Two drops
(0.01 mL) of suspension of spores/conidia of each of the
three test fungal species was added to each culture tube.
Culture tubes were completely closed with cotton plugs
and incubated at room temperature for 48 h. After 48 h,
tubes were observed for appearance of fungal mycelia. The
effectiveness of a compound was assessed in terms of
100 % inhibition of germination of spores. The minimum
inhibitory concentration (MIC) of each compound was
recorded.
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