Synthesis, pharmacological and biological screening of some novel pyrimidine derivatives

Article abstract of DOI:10.1007/s00044-014-0914-3

In the present study a new series of 4-substituted phenyl-6-(pyridin-2-yl) pyrimidin-2-ol (1-9) and 4-substituted phenyl-6-(pyridin-2-yl)pyrimidin-2-thiol (10-18) have been synthesized by cyclizing 3-substituted phenyl-1-(pyridin-2-yl) prop-2-en-1-one wit

Full text of DOI:10.1007/s00044-014-0914-3

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-014-0914-3
ORIGINAL RESEARCH
Synthesis, pharmacological and biological screening of some novel
pyrimidine derivatives
•
K. Ishwar Bhat Abhishek Kumar
•
•
Muhammed Nisar Pankaj Kumar
Received: 25 July 2013 / Accepted: 9 January 2014
Ó Springer Science+Business Media New York 2014
Abstract In the present study a new series of 4-substi-
tuted phenyl-6-(pyridin-2-yl)pyrimidin-2-ol (1–9) and
4-substituted phenyl-6-(pyridin-2-yl)pyrimidin-2-thiol (10–
18) have been synthesized by cyclizing 3-substituted phe-
nyl-1-(pyridin-2-yl)prop-2-en-1-one with urea/thiourea in
the presence of NaOH as base (1a–18a). 3-Substituted
phenyl-1-(pyridin-2-yl)prop-2-en-1-one was prepared by
condensing 2-acetylpyridine with substituted benzaldehyde
in the presence of 20 % NaOH as base. The structures of
the newly synthesized compounds were characterized by
elemental analysis, IR, 1H NMR, 13C NMR, and mass
spectroscopic studies. Newly synthesized compounds were
screened for their anti-inflammatory, analgesic, cytotoxic,
and antitubercular activities studies. Few of the compounds
exhibited excellent anti-inflammatory activities compared
to standard drug diclofenac sodium. Some compounds have
shown moderate analgesic activities compared to standard
drug pentazocine. Also, some compounds have exhibited
moderate cytotoxic and antitubercular activities.
Introduction
Pyrimidine itself is not found in nature but substituted
pyrimidines and compounds containing the pyrimidine ring
are widely distributed in nature. As pyrimidine is a basic
nucleus in DNA and RNA, it has been found to be asso-
ciated with diverse biological activities. Pyrimidine deriv-
atives are well known for the wide spectrum of biological
activities such as antimicrobial (Mohamed et al., 2010),
antitumor (Linyi and Malhotra, 2012; Sherif et al., 2009),
analgesic (Nofal et al., 2009), anti-inflammatory (El-Gaz-
zar et al., 2007), antibacterial (Hilmy et al., 2010), anti-
fungal (Sun et al., 2011), antiplatelet (Bruno et al., 2011),
and antitubercular (Mohan et al., 2010) activities. Many of
the pyrimidine derivatives are reported to possess potential
CNS depressant properties (Shishoo et al., 1981). A num-
ber of nonsteroidal anti-inflammatory drugs (NSAIDs) are
available clinically to treat inflammatory disorders. The
main mechanism of anti-inflammatory action of NSAIDs is
by inhibition of prostaglandin synthesis. Cyclooxygenase
(COX; prostaglandin G/H synthase, EC 1.14.99.1) cata-
lyzes the first two steps in the biosynthesis of biological
mediator prostaglandin, and there lies a great interest in
COX-2 as a key therapeutic target for inflammation
(Rouzer and Marnett, 2009; McKellar et al., 2008; Jan-
kowski and Hunt, 2008). But side effects associated with
chronic use of these NSAIDs are gastrointestinal ulcer.
Apart from this, there is evidence to suggest that leuko-
trienes enhance gastric ulceration which limits the thera-
peutic utilization of these drugs (Osiri and Moreland, 1999;
Lombardino, 1985). Pyrimidine heterocycles possessing
hydroxyl group has a unique place in medicinal chemistry
and also plays a vital role in biological processes as well as
synthetic drugs. Prompted by these literatures, we planned
to synthesize some novel pyrimidine derivatives derived
Keywords Chalcones ꢀ Pyrimidines ꢀ Anti-inflammatory ꢀ
Analgesic ꢀ Cytotoxicity ꢀ Antitubercular
K. Ishwar Bhat (&) ꢀ A. Kumar ꢀ M. Nisar ꢀ P. Kumar
Department of Pharmaceutical Chemistry, NGSM Institute
of Pharmaceutical Sciences of Nitte University, Paneer,
Deralakatte, Mangalore 575 018, Karnataka, India
e-mail: bhatishwar@yahoo.co.in
A. Kumar
e-mail: abhi12bunty@gmail.com
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Med Chem Res
from chalcones and to explore their biological and phar-
macological activities.
9, and 11 containing 4-chlorophenyl, 3-hydroxyphenyl,
2-bromophenyl (pyrimidin-2-ol derivatives), and 4-fluoro-
phenyl (pyrimidin-2-thiol derivatives) exhibited significant
anti-inflammatory activities compared to the standard drug
diclofenac. The presence of pyrimidine moiety with elec-
tron-withdrawing groups like chloro, fluoro, and bromo
accounted for the anti-inflammatory activity.
Herein, the synthesis of some derivatives of some title
structure compounds containing pyridine moiety is repor-
ted in an attempt to significantly improve the biological
spectrum of pyrimidines. All the novel compounds were
evaluated for their anti-inflammatory, analgesic, cytotoxic,
and antitubercular activities.
Analgesic activity studies
Results and discussion
All the compounds were tested for their analgesic activities by
tail-immersion method (Ramabadran et al., 1989) and
exhibited percentage inhibition in the range of
12.40–147.74 % after 120 min. The compounds 1, 2, 3, 7, and
12 containing 4-chlorophenyl, 4-fluorophenyl, 3-hydroxy-
phenyl, 4-methoxyphenyl (pyrimidin-2-ol derivatives), and
3-hydroxyphenyl (pyrimidin-2-thiol derivatives) substituents
showed highly potent analgesic activities compared to the
standard drug pentazocine, whereas rest of the tested com-
poundsdidnotshowsignificantanalgesicactivityinthemodel
used. The presence of pyrimidine moiety with other substi-
tution and groups like fluoro, chloro, and hydroxy has
accounted for their analgesic activities.
Chemistry
Various substituted 3-substituted phenyl-1-(pyridin-2-
yl)prop-2-en-1-ones were prepared as reported in the liter-
ature (Babasaheb et al., 2010) by the reaction of 2-acetyl-
pyridine and substituted aromatic aldehydes in the presence
of 20 % NaOH as base. These compounds were character-
ized by TLC, melting point, and IR spectra that showed
characteristic absorption band at 1,670 cm-1 of CH=CH
group. The title compounds 4-substituted phenyl-6-(pyridin-
2-yl)pyrimidin-2-ol (1–9) and 4-substituted phenyl-6-(pyri-
din-2-yl)pyrimidin-2-thiol (10–18) derivatives were pre-
pared as reported in the literature (Munawar et al., 2008) by
cyclization of chalcones with urea/thiourea using ethanol in
the presence of NaOH as base. The infrared spectra of the
substituted pyrimidines 1–18 showing characteristic
absorption bands at 3,400–3,450 cm-1 were attributed to
OH and 2,500–2,550 cm-1 were attributed to SH group
which is absent in the intermediate chalcones. Similarly, the
1H NMR of the synthesized pyrimidines showed a broad
singlet at d 9.6–10.0 due to OH and at d 9.2–9.9 due to SH
Cytotoxic activity studies
The test compounds were subjected to in vitro cytotoxicity
against Ehrlich ascites carcinoma (EAC) cells using ‘‘try-
pan blue exclusion method’’ (Louis and Siegel, 2011). The
damaged cells are stained blue by trypan blue stain and can
be distinguished from viable cells. Compounds 10 and 11
containing 2-chlorophenyl (pyrimidin-2-ol derivatives) and
2-bromophenyl (pyrimidin-2-thiol derivatives) substituents
induced the greatest effect on EAC cells with an activity
more than 60 % at a concentration of 250 lg/ml. The
presence of pyrimidine moiety with other substitution and
groups like chloro and fluoro has accounted for their
cytotoxic activity.
1
group which was absent in the H NMR spectra of the
substituted chalcones. Hence, the formation of the title
compounds pyrimidines was confirmed and further estab-
lished by 13C NMR and mass spectra which are in accor-
dance with molecular formula.
Anti-inflammatory activity studies
Antitubercular activity studies
All the synthesized compounds were tested for their anti-
inflammatory activities using carrageenan-induced rat hind
paw edema method (Winter et al., 1962). Data of anti-
inflammatory activity were expressed as mean SEM,
and the student’s t test was applied to determine the sig-
nificance of the difference between the control group and
rats treated with the test compounds. The anti-inflamma-
tory activity of the newly synthesized compounds was
compared with the standard diclofenac sodium 13.5 mg/kg
body weight, showing 64.52 % inhibition of rat paw edema
whereas the tested compounds showed inhibition ranging
from 31.61 to 58.18 % after 120 min. The compounds 1, 3,
The test compounds were evaluated for their antitubercular
activities against M. tuberculosis using microplate alamar
blue assay (Chauca et al., 2007). A blue color in the well
was interpreted as no bacterial growth and pink color was
scored as growth. This indicates that the test compound has
a potent antitubercular activity in in vitro condition. Com-
pounds 6 and 9 containing 3-nitrophenyl and 2-bromophenyl
(pyrimidin-2-ol derivatives) substituents induced the greatest
effect on bacterial growth with an activity at a concentration
of 25 lg/ml. The presence of pyrimidine moiety with other
substitutions and groups like nitro and bromo has accounted
for their antitubercular activity.
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Med Chem Res
General procedure for the synthesis of 4-substituted
phenyl-6-(pyridin-2-yl)pyrimidin-2-ol and 2-thiol
(1–18)
Conclusion
Several 4-substituted phenyl-6-(pyridin-2-yl)pyrimidin-2-
ol and 4-substituted phenyl-6-(pyridin-2-yl)pyrimidin-2-
thiol were successfully synthesized in 57–76 % yields and
A mixture of 3-substituted phenyl-1-(pyridin-2-yl)prop-2-en-
1-one (1a–18a) (0.01 mol) and urea/thiourea was dissolved
in 20 ml ethanol in the presence of 4 ml 20 % NaOH and the
reaction mixture was refluxed for about 18 h. After the
completion of the reaction, the reaction mixture was poured
into crushed ice. The separated solid was filtered, washed
with cold water, and recrystallized from ethanol (Fig. 1).
1
are characterized by H NMR, mass spectrometry, and IR
studies. All the synthesized compounds were screened for
their anti-inflammatory, analgesic, cytotoxic, and antitu-
bercular activities. Anti-inflammatory activity was evalu-
ated by carrageenan-induced paw edema method.
Compounds 1, 3, 9, and 11 were found to be biologically
active whereas the remaining compounds showed poor
anti-inflammatory activities. Analgesic activity was eval-
uated by tail-immersion method. Compounds 1, 2, 3, 7, and
12 showed highly potent analgesic activities whereas rest
of the tested compounds did not show significant analgesic
activities. Cytotoxic activity was evaluated by trypan blue
exclusion method. Compounds 10 and 11 induced the
greatest effect on EAC cells with an activity more than
60 % at a concentration of 250 lg/ml. Antitubercular
activity was evaluated by microplate alamar blue assay.
Compounds 6 and 9 induced the greatest effect on bacterial
growth with an activity at a concentration of 25 lg/ml.
3-(2-Chlorophenyl)-1-(pyridin-2-yl)prop-2-en-1-one (1a)
Yellowish needles (EtOH). This compound was prepared
by condensing 2-acetylpyridine with substituted benzalde-
hyde in the presence of NaOH. It was obtained as a yellow
solid; mp 75–77 °C; UV (EtOH) kmax (log e) 270 (4.25)
nm; IR (KBr) mmax 1,506(Ar C=C str), 3,120(Ar C–H
bending), 1,690(C=O str), 2,900(aliphatic C–H str), 785(C–
Cl str), 1,680(Ar C=N str) cm-1 1H NMR (CDCl3,
;
400 MHz): d 6.9 (1H, d, J = 15.1 Hz, 3H), 7.1 (1H, d,
J = 15.1 Hz, 2H), d 7.2–7.6 (8H, m, Ar–H); 13C NMR
(CDCl3, 300 MHz): 187(C=O, C-1), 121.3, 145.1(CH=CH,
C-2, C-3), 134(C, C-20), 129.9, 129.3, 126.7, 127.8, 120.7,
137.3, 128, 149.7(8 CH aromatic, C-30, C-40, C-50, C-60 of
2-chlorophenyl, C-300, C-400, C-500, C-600 of pyridine), 133,
153.3(2C aromatic, C-10, C-200); MS: m/z: 245 (M?1);
Anal. calcd. for C14H11ClON: C, 68.70; H, 4.53; N, 5.73.
Found: C, 68.62; H, 4.51; N, 5.86.
Experimental
Materials and methods
Melting points were determined by open capillary and were
uncorrected. The IR spectra (in KBr pellets) were recorded
in Tensor 27 spectrophotometer, Bruker optik (Germany).
1H NMR spectra were recorded (CHCl3) on Bruker
(400Mz) spectrometer using TMS as an internal standard.
Chemical shift values are given in d scales. The mass
spectra were recorded using a Jeol-D-300 mass spectrom-
eter (70 eV), Shimadzu (Japan) by FAB. Elemental ana-
lysis was performed using Vairo Elementar Model, C, H, N
analyzer. The completion of the reaction was checked by
thin-layer chromatography (TLC) on silica gel-G-coated
plates using ethylacetate:chloroform (9:1) as the solvent
and observed in UV light. Commercial-grade solvents and
reagents were used without further purification.
3-(4-Fluorophenyl)-1-(pyridin-2-yl)prop-2-en-1-one (2a)
Green needles (EtOH). This compound was prepared by
condensing 2-acetylpyridine with substituted benzaldehyde
in the presence of NaOH. It was obtained as a green solid; mp
82–84 °C; UV (EtOH) kmax (log e) 275 (4.28) nm; IR (KBr)
mmax 1,502(Ar C=C str), 3,120(Ar C–H bending), 1,692(C=O
str), 2,900(aliphatic C–H str), 1,221(C–Br str), 1,680(Ar
C=N str) cm-1; 1H NMR (CDCl3, 400 MHz): d 6.9 (1H, d,
J = 15.1 Hz, H-3), 7.1 (1H, d, J = 15.1 Hz, H-2), d 7.2–7.5
(8H, m, Ar–H); 13C NMR (CDCl3, 300 MHz): 187(C=O,
C-1), 121.3, 145.1(CH=CH, C-2, C-3), 162.1(C, C-40), 130.4,
1150.4, 130.4, 130.8, 137.3, 128, 128, 149.7(8 CH aromatic,
C-2 , C-30, C-50, C-60 of 4-fluorophenyl, C-300, C-400, C-500,
C-600 of pyridine), 130.8, 153.3(2C aromatic, C-10, C-200);
MS: m/z: 228 (M?1); Anal. calcd. for C14H11FON: C, 73.66;
H, 4.86; N, 6.14. Found: C, 73.12; H, 4.88; N, 6.18.
General procedure for the synthesis of 3-substituted
phenyl-1-(pyridin-2-yl)prop-2-en-1-one (1a–18a)
A mixture of 2-acetylpyridine (0.01 mol) and substituted
benzaldehyde (0.01 mol) in absolute ethanol (20 ml) were
stirred together for 24 h in the presence of 20 % NaOH (3-
4 ml). The reaction mixture was poured into crushed ice
and acidified with HCl. The separated solid was filtered,
washed with water, and recrystallized from ethanol.
3-(3-Hydroxyphenyl)-1-(pyridin-2-yl)prop-2-en-1-one (3a)
Dark-brown needles(EtOH). Thiscompoundwaspreparedby
condensing 2-Acetylpyridine with substituted benzaldehyde
123

Med Chem Res
Fig. 1 Scheme for pyrimidine
derivatives
R
H
C
H
C
O
3
+
N
O
Substituted benzaldehyde
2-Acetyl pyridine
20% NaOH
C2H5OH
R
N
O
Chalcones (1a-18a)
O
S
C
(a)
(b)
C
H2N
NH2
H2N
NH2
R
N
N
N
Y: a=OH/b=SH
Y
Pyrimidine derivatives (1-18)
R= 4Cl 4F 3-OH 4-N(CH ) 4-CH 3-NO 4-OCH 4-CN 2Br 2-Cl
,
,
,
,
,
3
,
2
,
3
,
,
3 2
in the presence of NaOH. It was obtained as a dark-brown
solid;mp105–107 °C;UV(EtOH)kmax (loge)265(4.23)nm;
IR (KBr) mmax 1,509(Ar C=C str), 3,125(Ar C–H bending),
1,690(C=O str), 2,912(aliphatic C–H str), 3,420(O–H str),
1,680(Ar C=N str) cm-1; 1H NMR (CDCl3, 400 MHz): d 6.9
(1H, d, J = 15.1 Hz, H-3), 7.1 (1H, d, J = 15.2 Hz, H-2), d
7.2–7.6 (8H, m, Ar–H); 13C NMR (CDCl3, 300 MHz):
187(C=O, C-1), 121.3, 145.1(CH=CH, C-2, C-3), 162.1(C,
C-30), 117.6, 115.1, 130, 121, 120.7 137.3, 128, 149.7(8 CH
aromatic, C-20, C-40, C-50, C-60 of 3-hydroxyphenyl, C-300,
C-400, C-500, C-600 of pyridine), 135.4, 153.3(2C aromatic,
C-10, C-200); MS: m/z: 216 (M?); Anal. calcd. for C14H12O2N:
C, 74.31; H, 5.35; N, 6.19. Found: C, 74.21; H, 5.33; N, 6.23.
4-(2-Chlorophenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (1)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 2-chloro chalcones with urea. It was
obtained as a yellow solid; mp 128–130 °C; yield 72 %; IR
(KBr) mmax 1,687(C=N str), 3,058(aromatic C–H str),
1,589(aromatic C=C str), 778 (C–Cl str), 1,350(C–N str),
1
3,419(O–H str) cm-1; H NMR (CDCl3): d 7.51–7.92 (m,
123

Med Chem Res
9H, Ar–H), 9.73 (s, 1H, H-2); 13C NMR (CDCl3, 300 MHz):
159.6 (C, C-2), 132.6 (C, C-200), 93.4, 124.0, 128.9, 130.1,
130.6, 132.5, 134.0, 147.5, 149.7 (9 CH aromatic, C-5, C-50,
C-600, C-400, C-500, C-300, C-40, C-20, C-60), 129.9, 133, 155.1,
159.1 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 284
(M?2); Anal. calcd. for C15H11ClN3O: C, 63.27; H, 3.87; N,
14.76. Found: C, 63.19; H, 3.82; N, 14.81.
C-600, C-40, C-20, C-60), 125.3, 133, 155.1, 159.1 (4C aro-
matic, C-100, C-30, C-4, C-6); MS: m/z 280 M?); Anal.
calcd. for C17H17N4O: C, 69.62; H, 5.80; N, 19.11. Found:
C, 69.69; H, 5.72; N, 19.16.
4-(pyridin-2-yl)-6-p-tolyl-pyrimidin-2-ol (5)
Clay-colored needles (EtOH). This compound was
obtained by condensing substituted 4-methyl chalcones
with urea. It was obtained as a clay-colored solid; mp
86-88 °C; yield 70 %;IR (KBr) mmax 1,682(C=N str),
1,350(C–N aromatic), 3,105(Ar C–H str), 2,985(aliphatic
4-(4-Fluorophenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (2)
Green needles (EtOH). This compound was obtained by
condensing substituted 4-fluoro chalcones with urea. It was
obtained as a green solid; mp 99–102 °C; yield 75 %; IR
(KBr) mmax 1,680(C=N str), 3,058(Ar C–H str), 1,582(Ar
C=C str), 1,224(C–F str), 1,346(C–N str), 3,410(O–H str)
cm-1; 1H NMR (CDCl3): d 7.31–7.87 (m, 9H, Ar–H), 9.82
(s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 159.6 (C, C-2),
162.9 (C, C-400), 93.4, 116.0, 124.0, 130.6, 134.0, 147.5,
149.7 (9 CH aromatic, C-5, C-300, C-500, C-50, C-200, C-600,
C-40, C-20, C-60), 131.4, 133, 155.1, 159.1 (4C aromatic,
C-100, C-30, C-4, C-6); MS: m/z 268 (M?1); Anal. calcd.
for C15H11FN3O: C, 67.21; H, 4.11; N, 15.68. Found: C,
67.30; H, 4.08; N, 15.71.
1
CH str), 3,415(O–H str), 1,572(C=C str) cm-1; H NMR
(CDCl3): d 7.11–7.82 (m, 9H, Ar–H), 4.73 (s, 3H, H-400),
9.91 (s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 131.7 (C,
C-2), 132.6 (C, C-400), 93.4, 123.2, 124.0, 129.5, 134.0,
147.5, 149.7 (9 CH aromatic, C-5, C-200, C-600, C-300, C-500
C-50, C-40, C-20, C-60), 132.8, 133, 155.1, 159.1 (4C aro-
matic, C-100, C-30, C-4, C-6); MS: m/z 264 (M?); Anal.
calcd. for C16H14N3O: C, 72.73; H, 5.80; N, 15.91. Found:
C, 72.79; H, 5.39; N, 15.88.
4-(3-Nitrophenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (6)
4-(3-Hydroxyphenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (3)
Brown needles (EtOH). This compound was obtained by
condensing substituted 2-chloro chalcones with urea. It was
obtained as a brown solid; mp 154–156 °C; yield 68 %; IR
(KBr) mmax 1,691(C=N str), 1,354(C–N str), 3,095(Ar C–H
Dark-brown needles (EtOH). This compound was obtained
by condensing substituted 3-hydroxy chalcones with urea. It
was obtained as a dark-brown solid; mp 90–92 °C; yield
74 %; IR (KBr) mmax 1,683(C=N str), 3,045(Ar C–H str),
1,572(Ar C=C str), 1,350(C–N str), 3,425(O–H str),
1
str), 3,410(O–H str), 1,565(Ar C=C str) cm-1; H NMR
(CDCl3): d 7.12–7.65 (m, 9H, Ar–H), 9.87 (s, 1H, H-2);
13C NMR (CDCl3, 300 MHz): 159.6 (C, C-2), 148.4 (C,
C-300), 93.4, 103.5, 122.7, 123.9, 124.0, 129.5, 134.0,
147.5, 149.7 (9 CH aromatic, C-5, C-200, C-500, C-400, C-50,
C-600, C-40, C-20, C-60), 136.8, 133, 155.1, 159.1 (4C aro-
matic, C-100, C-30, C-4, C-6); MS: m/z 334 (M?); Anal.
calcd. for C15H11N4O3: C, 61.02; H, 3.73; N, 18.98. Found:
C, 61.12; H, 3.81; N, 18.91.
1
1,350(C–N str) cm-1; H NMR (CDCl3): d 7.43–7.91 (m,
9H, Ar–H), 9.83 (s, 2H, H-2); 13C NMR (CDCl3, 300 MHz):
159.6 (C, C-2), 138.9 (C, C-300), 93.4, 124.0, 124.5, 129,
129.1, 130.4, 134.0, 147.5, 149.7 (9 CH aromatic, C-5, C-50,
C-600, C-400 C-500, C-200, C-40, C-20, C-60), 132.9, 133, 155.1,
159.1 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 266
(M?); Anal. calcd. for C15H12N3O: C, 67.67; H, 4.51; N,
15.80. Found: C, 67.67; H, 4.62; N, 15.89.
4-(4-Methoxyphenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (7)
4-(4-(Dimethylamino)phenyl)-6-(pyridin-2-yl)pyrimidin-2-
White needles (EtOH). This compound was obtained by
condensing substituted 4-methoxy chalcones with urea. It
was obtained as a white solid; mp 80–82 °C; yield 76 %;
IR (KBr) mmax 1,682(C=N str), 1,350(C–N str), 3,090(Ar
C–H str), 3,415(O–H str), 1,560(Ar C=C str), 2,980(ali-
ol (4)
Dark-brown needles (EtOH). This compound was obtained
by condensing substituted 4-dimethylamino chalcones with
urea. It was obtained as a dark-brown solid; mp
145-147 °C; yield 60 %; IR (KBr) mmax 1,678(C=N str),
3,053(Ar C–H str), 1,560(Ar C=C str), 1,338(C–N str),
1
phatic C–H str) cm-1; H NMR (CDCl3): d 7.13–7.86 (m,
9H, Ar–H), 9.83 (s, 1H, H-2), 3.85 (s, 3H, H-400); 13C NMR
(CDCl3, 300 MHz): 159.6 (C, C-2), 132.6 (C, C-20), 93.4,
114.8, 124.0, 128.5, 134.0, 147.5, 149.7 (9 CH aromatic,
C-5, C-300, C-500, C-50, C-200, C-600, C-40, C-20, C-60), 128.1,
133, 155.1, 159.1 (4C aromatic, C-100, C-30, C-4, C-6); MS:
m/z 278 (M?1); Anal. calcd. for C16H14N3O2: C, 68.57; H,
5.01; N, 15.00. Found: C, 68.61; H, 5.09; N, 15.12.
1
2,955(aliphatic C–H str), 3,420(O–H str) cm-1; H NMR
(CDCl3): d 7.42–7.87 (m, 9H, Ar–H), 9.87 (s, 1H, H-2),
4.83 (s, 6H, H-400); 13C NMR (CDCl3, 300 MHz): 159.6
(C, C-2), 155.3 (C, C-400), 93.4, 112.7, 124.0, 128.4, 134.0,
147.5, 149.7 (9 CH aromatic, C-5, C-300, C-500, C-50, C-200,
123

Med Chem Res
4-(2-Hydroxy-6-(pyridin-2-yl)pyrimidin-
was obtained as a green solid, mp 106–108 °C, yield 70 %.
IR (KBr) mmax 1,690(C=N str), 3,055(Ar C–H str), 1,585(Ar
C=C str), 1,220(C–F str), 1,342(C–N str), 2,046(S–H str)
4-yl)benzonitrile (8)
cm-1 1H NMR (CDCI3): d 7.23–7.95 (m, 9H, Ar–H),
;
White needles (EtOH). This compound was obtained by
condensing substituted 4-cyano chalcones with urea. It was
obtained as a yellow solid; mp 150–152 °C; yield 61; IR
(KBr) mmax 1,690(C=N str), 1,360(C–N str), 3,082(Ar C–H
str), 3,420(O–H str), 1,555(Ar C=C str), 1,350(aliphatic C–
9.47(s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 180.2 (C,
C-2), 162.9 (C, C-400), 108.3, 116.0, 124.0, 130.6, 134.0,
147.5, 149.7 (9 CH aromatic, C-5, C-50, C-300, C-500, C-200,
C-600, C-40, C-20, C-60), 131.4, 133, 165, 161 (4C aromatic,
C-100, C-30, C-4, C-6); MS: m/z 284 (M?1); Anal. calcd.
for C15H11FN3S: C, 63.40; H, 3.87; N, 14.79. Found: C,
63.48; H, 3.92; N, 14.68.
1
N str) cm-1; H NMR (CDCl3): d 7.51–7.89 (m, 9H, Ar–
H), 9.94 (s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 159.6
(C, C-2), 112.6 (C, C-400), 93.4, 124.0, 126.1, 132.7, 134.0,
147.5, 149.7 (9 CH aromatic, C-5, C-50, C-200, C-600, C-300,
C-500, C-40, C-20, C-60), 140.1, 133, 155.1, 159.1 (4C aro-
matic, C-100, C-30, C-4, C-6); MS: m/z 289 (M?); Anal.
calcd. for C16H11N4O: C, 69.82; H, 4.00; N, 20.36. Found:
C, 69.91; H, 4.09; N, 20.425.
4-(3-Hydroxyphenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (12)
Dark-brown needles (EtOH). This compound was obtained
by condensing substituted 3-hydroxy chalcones with thio-
urea. It was obtained as a dark-brown solid; mp 120–122 °C;
yield 65 %; IR (KBr) mmax 1,679(C=N str), 3,052(Ar C–H
str), 1,572(Ar C=C str), 1,340(C–N str), 3,420(O–H str),
2,046(S–H str) cm-1; 1H NMR (CDCl3): d 7.23-7.95 (m, 9H,
4-(2-Bromophenyl)-6-(pyridin-2-yl)pyrimidin-2-ol (9)
Green needles (EtOH). This compound was obtained by
condensing substituted 2-bromo chalcones with urea. It
was obtained as a green solid; mp 110–112 °C; yield 69 %;
IR (KBr) mmax 1,678(C=N str), 1,365(C–N str), 3,082(Ar
C–H str), 3,425(O–H str) 1,562(Ar C=C str), 675 (C–Br
13
Ar–H), 9.86 (s, 1H, H-300), 9.47 (s, 1H, H-2); C NMR
(CDCl3, 300 MHz): 180.2 (C, C-2), 157.5 (C, C-300), 108.3,
115.9, 120.1, 124.0, 130.6, 134.0, 147.5, 149.7 (9 CH aro-
matic, C-5, C-200, C-400, C-600, C-50, C-500, C-40, C-20, C-60),
138.3, 133, 165, 161 (4C aromatic, C-100, C-30, C-4, C-6);
MS: m/z 282 (M?); Anal. calcd. for C15H12N3SO: C, 63.82;
H, 4.25; N, 1,489. Found: C, 63.89; H, 3.92; N, 14.68.
1
str) cm-1; H NMR (CDCl3): d 7.13–7.97 (m, 9H, Ar–H),
9.68 (s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 159.6 (C,
C-2), 132.6 (C, C-200), 93.4, 124.0, 128.2, 129.7, 130.9,
132.1, 134.0, 147.5 149.7 (9 CH aromatic, C-5, C-50, C-500,
C-600, C-400, C-300, C-40, C-20, C-60), 139.8, 133, 155.1,
159.1 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 330
(M?1); Anal. calcd. for C15H11ON3Br: C, 46.27; H, 3.34;
N, 12.77. Found: C, 46.32; H, 3.39; N, 12.82.
4-(4-(Dimethylamino)phenyl)-6-(pyridin-2-yl)pyrimidin-2-
thiol (13)
Dark-brown needles (EtOH). This compound was obtained by
condensing substituted 4-dimethylamino chalcones with thio-
urea. It was obtained as a dark-brown solid; mp 110–112 °C;
yield 58 %. IR (KBr) mmax 1,684(C=N str), 3,062(Ar C–H str),
1,569(Ar C=C str), 1,345(C–N str), 2,061(S–H str), 2,962(ali-
phatic C–H str) cm-1; 1H NMR (CDCl3): d 7.12–7.76 (m, 9H,
Ar–H), 9.97(s, 1H, H-2), 4.83 (s, 6H, H-400); 13C NMR (CDCl3,
300 MHz): 180.2 (C, C-2), 155.3 (C, C-400), 108.3, 112.7,
124.0, 128.4, 134.0, 147.5, 149.7 (9 CH aromatic, C-5, C-300,
C-500, C-50, C-200, C-600, C-40, C-20, C-60), 125.3, 133, 165, 161
(4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 309 (M?); Anal.
calcd. for C17H17N4S: C, 63.02; H, 5.50; N, 18.12. Found: C,
66.12; H, 5.59; N, 18.09.
4-(2-Chlorophenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (10)
Green needles (EtOH). This compound was obtained by
condensing substituted 2-chloro chalcones with thiourea. It
was obtained as a reddish solid; mp 87–89 °C; yield 74 %;
IR (KBr) mmax 1,690(C=N str), 1,370(C–N str), 2,052(S–H
str), 3,080(Ar C–H str), 1,572(Ar C=C str), 770(C–Cl str)
cm-1; 1H NMR (CDCl3): d 7.22–7.96 (m, 9H, Ar–H), 9.37
(s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 180.2 (C, C-2),
132.2 (C, C-200), 108.3, 124.0, 128.9, 130.1, 130.6, 132.5,
134.0, 147.5, 147.9 (9 CH aromatic, C-5, C-50, C-600, C-400,
C-500, C-300, C-40, C-20, C-60), 129.9, 133, 163.3, 161 (4C
aromatic, C-100, C-30, C-4, C-6); MS: m/z 302 (M?2);
Anal. calcd. for C15H11ClN3S: C, 59.90; H, 3.66; N, 13.98.
Found: C, 59.96; H, 3.71; N, 13.91.
4-(Pyridin-2-yl)-6-p-tolyl-pyrimidin-2-thiol (14)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 4-methyl chalcones with thiourea. It
was obtained as a yellow solid, mp 160–162 °C; yield 69 %;
IR (KBr) mmax 1,682(C=N str), 1,360(C–N str) 3,090(Ar C–
H str), 2,062(S–H str), 2,975(aliphatic C–H str), 1,570(Ar
4-(4-Fluorophenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (11)
Green needles (EtOH). This compound was obtained by
condensing substituted 4-fluoro chalcones with thiourea. It
123

Med Chem Res
C=C str) cm-1; 1H NMR (CDCl3): d 7.13–7.78 (m, 9H, Ar–
H),4.63 (s, 6H, H-400), 9.54 (s, 1H, H-2); 13C NMR (CDCl3,
300 MHz): 180.2 (C, C-2), 131.7 (C, C-400), 108.3, 123.2,
124.0, 129.5, 134.0, 147.5, 149.7 (9 CH aromatic, C-5, C-200,
C-600, C-50, C-300, C-500, C-40, C-20, C-60), 132.8, 133, 165,
161 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 280 (M?);
Anal. calcd. for C16H14N3S: C, 68.57; H, 5.00; N, 15.00.
Found: C, 68.51; H, 5.12; N, 15.13.
C-5, C-50, C-200, C-600, C-300, C-500, C-40, C-20, C-60), 140.1,
133, 165, 161 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z
291(M?); Anal. calcd. for C16H11N4S: C, 65.98; H, 3.78;
N, 19.24. Found: C, 65.88; H, 3.84; N, 19.36.
4-(2-Bromophenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (18)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 2-bromo chalcones with thiourea. It
was obtained as a green solid; mp 120–122 °C; yield 71 %;
IR (KBr) mmax 1,685(C=N str), 1,360(Ar C–H str), 2,058(S–
H str), 3,075(Ar C–H str), 1,562(Ar C=C str), 668(C–Br
4-(3-Nitrophenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (15)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 3-nitro chalcones with thiourea. It was
obtained as a black solid; mp 101–103 °C; yield 57 %; IR
(KBr)) mmax 1,685(C=N str), 1,350(C–N str), 3,090(Ar C–H
1
str) cm-1; H NMR (CDCl3): d 7.13–7.68 (m, 9H, Ar–H),
9.76 (s, 1H, H-2); 13C NMR (CDCl3, 300 MHz): 180.2 (C,
C-2), 120.2 (C, C-200), 108.2, 124.0, 128.2, 129.7, 130.9,
132.1, 134.0, 147.5, 149.7 (9 CH aromatic, C-5, C-50, C-500,
C-600, C-400, C-300, C-40, C-20, C-60), 139.8, 133, 163.3, 161
(4C aromatic, C-100, C-30, C-4, C-6); MS: m/z 345 (M?);
Anal. calcd. for C15H11N3SBr: C, 52.17; H, 3.19; N, 12.17.
Found: C, 52.24; H, 3. 23; N, 12.29.
1
str), 2,065(S–H str), 1,562(Ar C=C str) cm-1; H NMR
(CDCl3): d 7.13–7.73 (m, 9H, Ar–H), 9.46 (s, 1H, H-2); 13
C
NMR (CDCl3, 300 MHz): 180.2 (C, C-2), 148.4 (C, C-300),
103.5, 108.3, 122.7, 123.9 124.0, 129.5, 134.0, 147.5, 149.7 (9
CH aromatic, C-200, C-5, C-500, C-400, C-50, C-600, C-40, C-20,
C-60), 136.8, 133, 165, 161 (4C aromatic, C-100, C-30, C-4,
C-6); MS: m/z 311 (M?); Anal. calcd. for C15H14N4O2S: C,
57.32; H, 4.45; N, 17.83. Found: C, 57.19; H, 4.29; N, 17.80.
Anti-inflammatory activity
The anti-inflammatory activities of the test compounds was
carried out using carrageenan-induced rat paw edema model
according to Winter et al. (1962) by employing 1 % carra-
geenan solution as a phlogistic agent. Edema was induced in
the left hind paw of Wistar rats (150–200 g) of either sex by
the subplantar injection of 0.1 ml of 1 % carrageenan in
distilled water. Each group composed of six animals. The
animals which were bred in our laboratory were housed
under standard conditions and received a diet of commercial
food pellets and water ad libitum during the maintenance but
they were entirely fasted during the experiment period. Our
studies were conducted in accordance with recognized
guidelines on animal experimentation. The test compounds
were given intraperitoneally 30 min after carrageenan
injection. Diclofenac sodium was taken as the standard at a
dose of 10 mg/kg body weight (p.o). The rat paw volume was
measured after 1, 2, 3, and 4 h, respectively, after carra-
geenan injection by Plethysmometer. The difference
between the paw volume at 4 h and 0 h measurement was
calculated and taken as edema volume. Percentage inhibition
in the paw edema was calculated using the formula,
4-(4-Methoxyphenyl)-6-(pyridin-2-yl)pyrimidin-2-thiol (16)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 4-methoxy chalcones with thiourea.
It was obtained as a white solid; mp 90–92 °C; yield 70 %;
IR (KBr) mmax 1,690(C=N str), 1,345(C–N str), 2,054(S–H
str), 3,085(Ar C–H str), 1,554(Ar C=C str), 2,974(aliphatic
1
C–H str) cm-1; H NMR (CDCl3): d 7.23–7.95 (m, 9H,
13
Ar–H), 9.48 (s, 1H, H-2), 3.81 (s, 3H, H-400); C NMR
(CDCl3, 300 MHz): 180.2 (C, C-2), 160.6 (C, C-400), 108.3,
114.8, 124.0, 128.5, 134.0, 147.5, 149.7 (9 CH aromatic,
C-5, C-300, C-500, C-50, C-200, C-600, C-40, C-20, C-60), 128.1,
133, 165, 161 (4C aromatic, C-100, C-30, C-4, C-6); MS: m/z
296 (M?); Anal. calcd. for C16H14N3SO: C, 64.86; H,
4.73 N, 14.19. Found: C, 64.79; H, 4.69; N, 14.77.
4-(2-Mercapto-6-(pyridin-2-yl)pyrimidin-4-yl)benzonitrile
(17)
Yellow needles (EtOH). This compound was obtained by
condensing substituted 4-cyano chalcones with thiourea. It
was obtained as a yellow solid; mp 150–152 °C; yield
65 %; IR (KBr) mmax 1,690(C=N str), 1,358(Ar C–H str),
2,065(S–H str), 3,085(Ar C–H str), 1,575(Ar C=C str),
% Edema inhibition ¼ 100ð1 ꢁ Vt=VcÞ;
where Vt represents mean increase in paw volume of test
and Vc represents mean increase in paw volume of control.
1,345(aliphatic C–N str) cm-1
;
1H NMR (CDCl3): d
Statistical analysis
13
7.13–7.85 (m, 9H, Ar–H), 9.57 (s, 1H, H-2); C NMR
(CDCl3, 300 MHz): 180.2 (C, C-2), 112.6 (C, C-400), 108.3
124.0, 126.1, 132.7, 134.0, 147.5, 149.7 (9 CH aromatic,
All the experimental groups were composed of six animals.
Data obtained from animal experiments were expressed as
123

Med Chem Res
Table 1 Anti-inflammatory activities of compounds (1–18) by carrageenan-induced paw edema in rats
Treatment Dose Increase in the paw volume (ml)
(mg/kg)
1 h
2 h
3 h
4 h
0.3867 0.0223
Control
Vehicle 0.2827 0.0318
0.3716 0.0315
0.385 0.0209
Diclofenac 10
sodium
0.1083 0.0296** (61.69) 0.1217 0.0282** (67.24) 0.1362 0.0144** (64.62) 0.1372 0.0145** (64.52)
1
50
50
50
50
50
50
50
50
50
50
50
50
0.2067 0.0164 (29.13)
0.1817 0.1817* (37.70)
0.1927 0.0145** (48.84) 0.47 0.01** (50.23)
0.2805 0.0065 (31) 0.2667 0.0098 (30.72)
0.54 0.01** (57.33)
2
0.2633 0.0066** (31.61)
3
0.1817 0.00094* (37.70) 0.165 0.0076** (56.19)
0.1928 0.0263** (49.92) 0.1617 0.0256** (58.18)
4
0.1912 0.0013 (32.36)
0.1932 0.12 (31.65)
0.1617 0.11 (42.80)
0.2067 0.0164 (29.13)
0.1912 0.03 (32.36)
0.1927 0.0094* (31.83)
0.2013 0.0066 (45.82)
0.1987 0.021 (29.71)
0.165 0.063 (55.59)
0.2805 0.0065 (24.51)
0.2613 0.001 (29.68)
0.2517 0.020 (34.62)
0.22 0.066 (42.85)
0.1809 0.022 (53.01)
0.2667 0.0098 (30.72)
0.242 0.064 (37.14)
0.22 0.07 (43.10)
5
0.172 0.067 (55.52)
0.1617 0.023 (51.18)
0.172 0.067 (55.52)
0.231 0.038 (40.26)
0.162 0.0256** (58.03)
0.201 0.0016 (47.29)
6
7
8
9
0.1752 0.0076** (17.54) 0.188 0.0163** (51.16)
0.1928 0.0263** (49.92) 0.2667 0.0098 (30.72) 0.231 0.038 (40.10)
10
11
12
0.1912 0.02 (32.08)
0.1932 0.12 (31.65)
0.175 0.0076** (52.90)
0.1809 0.022 (51.31)
0.1912 0.0263** (50.33) 0.1617 0.0256** (58.18)
0.22 0.07 (42.85) 0.213 0.0064 (44.91)
All the values are expressed as mean SEM (n = 6)
* P \ 0.05 significant compared to control
** P \ 0.01 significant compared to control
mean SEM. The statistical significance of difference
between groups was assessed by means of analysis of
variance (ANOVA) followed by Dunnett’s test. The results
of the anti-inflammatory studies are presented in Table 1.
mean SEM, and percentage analgesic activity shown by
the tested compounds are presented in Table 2.
Cytotoxic activity
Analgesic activity
All the test compounds were studied for short-term in vitro
cytotoxicity against EAC cells. The tumor cells aspirated
from peritoneal cavity of tumor-bearing mice was washed
thrice with normal saline and checked for viability using
trypan blue exclusion method. The cell suspension (1
million cells in 0.1 ml) was added to tubes containing
various concentrations of the test compounds and volume
was made up to 1 ml using phosphate-buffered saline.
Control tubes contained only cell suspension. The assay
mixtures were incubated for 3 h at 37 °C and then percent
of dead cells was evaluated by trypan blue exclusion
method. The results of the cytotoxicity studies are pre-
sented in Table 3.
All the test compounds were tested for their analgesic
activities using tail-immersion method. The animals were
weighed and numbered with different identification marks.
Animals were divided into ten different groups containing
six of each. Albino rats of either sex weighing between 150
and 200 g were used for the experiment. pentazocine at the
dose 10 mg/kg body weight (oral) was used as standard
which showed percentage analgesic activity of 142.48 %.
The lower 5 cm of the tail of animals was marked and this
part of the tail was immersed in a cup of freshly filled water
at 55 °C. Within few seconds the rat reacts by withdrawing
the tail. A withdrawal timing of more than 6 s was regar-
ded as positive response. The reaction time of animals in
warm water at 0, 30, 60, 90, and 120 min after the treat-
ment was noted. Increase or decrease in reaction time of
the test substance was then compared to that of the stan-
dard-drug-treated and solvent-treated animals. The per-
centage increase or decrease in reaction time at each time
interval was calculated as a percentage increase in reaction
time = (Rt/Rc) - 1 9 100, where Rt is the mean reaction
time of the treated group; Rc is the mean reaction time of
the control group. All the values are expressed as
Antitubercular activity
The antitubercular activities of the test compounds were
assessed against Mycobacterium tuberculosis using
microplate alamar blue assay. This methodology is non-
toxic, uses a thermally stable reagent, and shows good
correlation with proportional and BACTEC radiometric
method. 200 ll of sterile deionized water was added to all
the outer-perimeter wells of sterile 96-well plate to mini-
mize evaporation of medium in the test wells during
123

Med Chem Res
Table 2 Analgesic activities of compounds (1–18) by tail-immersion method in rats
Treatment
Dose (mg/kg) Reaction time in sec at time (min)
0 min 30 min
2.42 0.01 2.49 0.02
60 min
90 min
120 min
Control
–
2.58 0.03
2.56 0.02
2.66 0.03
Pentazocine 10
2.32 0.01 4.92 0.02* (130.5) 8.14 0.02* (215.50) 8.19 0.01* (219.92) 6.45 0.02** (142.48)
2.17 0.05 3.82 0.01* (53.41) 6.26 0.01** (142.63) 6.99 0.04** (173.04) 5.43 0.02* (104.13)
1
50
50
50
50
50
50
50
50
50
2
2.66 0.02 3.27 0.14 (31.32) 4.12 0.05* (59.68)
2.64 0.03 3.69 0.02* (48.19) 4.12 0.06* (59.68)
6.32 0.06** (146.87) 5.9 0.04** (121.80)
6.88 0.03* (168.75) 6.25 0.03** (134.96)
3
4
2.03 0.04 2.31 0.03* (7.22)
2.47 0.03 3.88 0.01* (55.82) 7.52 0.03** (191.47) 7.18 0.04** (180.46) 6.59 0.03** (147.74)
2.82 0.06 3.12 0.04 (25.30) 3.10 0.06 (20.15) 4.26 0.03* (66.40) 2.33 0.03 (12.40)
2.34 0.02 3.66 0.02* (46.98) 5.87 0.03** (127.51) 7.58 0.02** (196.09) 3.80 0.01* (42.85)
2.12 0.02 2.42 0.03 (2.81) 2.77 0.03 (7.36) 3.69 0.02** (44.14) 3.05 0.03 (14.66)
2.55 0.03 4.23 0.03* (69.87) 7.45 0.03** (188.75) 7.21 0.03** (181.64) 6.2 0.01** (133.08)
3.2 0.04 (24.03)
3.4 0.02 (32.8)
2.26 0.03 (15.03)
7
8
9
10
12
All the values are expressed as mean SEM (n = 6)
*P \ 0.05 significant compared to control
** P \ 0.01 significant compared to control
Table 3 Cytotoxic activities of compounds (1–18) by trypan blue
exclusion method
were covered and sealed with parafilm and incubated at
37 °C for 5 days. After this, 25 ll of freshly prepared 1:1
mixture of alamar blue reagent and 10 % tween 80 was
added to the plate and incubated for 24 h. A blue color in
the well was interpreted as no bacterial growth and pink
color was interpreted as growth. The minimum inhibitory
concentration was defined as the lowest drug concentration
which prevented the color change from blue to pink. The
results of the antitubercular studies are presented in
Table 4.
Compounds No. of dead cells (%) at different concentrations (lg/ml)
50
100
200
250
Control
–
1
24
25
21
24
18
32
30
30
33
22
52
54
44
57
42
59
60
51
64
69
2
6
10
11
Acknowledgments The authors are thankful to the Head, SAIF,
1
Punjab University, Chandigarh for H NMR and Mass spectroscopic
analysis; and Mangalore University for elemental analysis. The
authors are highly thankful to Nitte University for providing all the
necessary research facilities and financial assistance.
Table 4 Antitubercular activities of compounds (1–18) by micro-
plate alamar blue assay
Compounds
Conc. (lg/ml)
References
100 50 25 12.5 6.25 3.125 1.6 0.8 0.4 0.2
Isoniazid
(standard)
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