Synthesis, analgesic, anti-inflammatory and antimicrobial studies of 2,4-dichloro-5-fluorophenyl containing thiazolotriazoles

Article abstract of DOI:10.1016/j.ejmech.2008.04.022

A series of 2,4-dichloro-5-fluorophenyl containing thiazolotriazoles (1) were synthesized starting from 3-(2,4-dichloro-5-fluorophenyl)-4H-1,2,4-triazole-3-thiol (5). The newly synthesized compounds were characterized by IR, ¹H NMR, mass and el

Full text of DOI:10.1016/j.ejmech.2008.04.022

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European Journal of Medicinal Chemistry 44 (2009) 827e833
http://www.elsevier.com/locate/ejmech
Original article
Synthesis, analgesic, anti-inflammatory and antimicrobial studies
of 2,4-dichloro-5-fluorophenyl containing thiazolotriazoles
Mari Sithambaram Karthikeyan1
Department of Chemistry, Mangalore University, Mangalagangothri 574199, India
Received 8 February 2008; received in revised form 15 April 2008; accepted 30 April 2008
Available online 10 May 2008
Dedicated to my beloved Professor B. Shivarama Holla on the occasion of his 62nd birthday.
Abstract
A series of 2,4-dichloro-5-fluorophenyl containing thiazolotriazoles (1) were synthesized starting from 3-(2,4-dichloro-5-fluorophenyl)-4H-
1
1,2,4-triazole-3-thiol (5). The newly synthesized compounds were characterized by IR, H NMR, mass and elemental analysis. Some of the
synthesized compounds were tested for their anti-inflammatory, analgesic and antimicrobial activities. Compounds 8c, 8d and 8h exhibited
excellent anti-inflammatory activity whereas compounds 8d, 8e and 8h showed excellent analgesic activity. Compounds 8a, 8cee, and 8h
exhibited good antibacterial activity and compounds 8c, 8e and 8h showed good antifungal activity.
Ó 2008 Elsevier Masson SAS. All rights reserved.
Keywords: 3-(2,4-Dichloro-5-fluorophenyl)-4H-1,2,4-triazole-5-thiol; PPA cyclization; Thiazolotriazole; Biological activities
1. Introduction
Ritonavir (antiretroviral drug) [10] and Bleomycine and
Tiazofurin (antineoplastic agents) [11]. Thiazole ring is an
important pharmacophore [12] and its coupling with other
rings could furnish new biologically active compounds. Thia-
zole containing compounds exhibit a wide range of biological
properties, such as anti-inflammatory, antimicrobial [13], anti-
tumor [14], anticonvulsant [15], cardiotonic [16], IMP dehy-
drogenase inhibiton [17], antithermic, analgesic [18],
anticancer [19], fungicidal and insecticidal [20].
1,2,4-Triazoles represent an overwhelming and rapid devel-
oping field in modern heterocyclic chemistry. From literature
it is predictable that, 1,2,4-triazoles represent important phar-
macophores, and play a vital role as medicinal agents. A
degree of respectability has been bestowed for 1,2,4-triazole
derivatives due to their wide range of biological activities
such as antibacterial, antifungal [21], antitubercular [22] and
anticancer [23]. Certain 1,2,4-triazoles also find applications
in the preparation of photographic plates, polymers, and as
analytical agents [24].
In recent years active research has been initiated on
halogen containing heterocycles, particularly fluorine and
chlorine. 2,4-Dichloro-5-fluoroacetophenone is used in the
synthesis of drugs like Ciprofloxacin and their analogues
[1]. Incorporation of fluorine or CF3 group into an organic
molecule largely enhances the pharmacological properties
[2e4] and also could lead to increased lipid solubility,
thereby enhancing the rates of absorption and transport of
drugs in vivo. The replacement of hydrogen or hydroxy group
by fluorine is a strategy widely used in drug development to
alter biological function [5].
Thiazoles are important class of heterocyclic compounds,
found in many potent biologically active molecules such as
Fentiazac, Meloxicam (anti-inflammatory agent) [6,7], Nizati-
dine (antiulcer agent) [8], Sulfatiazol (antimicrobial agent) [9],
Fluorine incorporated heterocycles, thiazoles and triazoles
displayed varied pharmacological properties. Since there
have been only few reports on dichlorofluorophenyl containing
E-mail address: karthims02@rediffmail.com
Present address: Syngene International Limited, Plot Nos. 2 and 3 C,
Unit-II, Bommansandra Industrial Area, Bangalore 560099, Karnataka, India.
1
0223-5234/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.04.022

828
M.S. Karthikeyan / European Journal of Medicinal Chemistry 44 (2009) 827e833
thiazolotriazoles, it was contemplated to synthesize them and
to pursue their anti-inflammatory, analgesic and antimicrobial
activities. The corresponding results are presented in this
paper.
4.2. Analgesic studies
All compounds were screened for analgesic activity by hot
plate test according to Eddy and Leimbach [30]. The analgesic
screening results revealed that compounds 8d, 8e and 8h
showed excellent analgesic activity whereas compounds 8a
and 8f showed moderate to good analgesic activity compared
with Pethidine. Compounds 8b, 8c, 8g showed lesser degree of
activity.
2. Chemistry
2,4-Dichloro-5-fluorobenzoyl hydrazide (3) was obtained
starting from 2,4-dichloro-5-fluoroacetophenone (1) according
to Refs. [25,26]. 3-(2,4-Dichloro-5-fluorophenyl)-4H-1,2,4-
triazole-5-thiol (5) was prepared according to Ref. [27] in
good yield. Substituted phenacyl bromides were prepared
according to Ref. [28]. When triazole (5) was treated with
substituted phenacyl bromides (6) in the presence of base in
ethanol medium yielded 2-[5-(2,4-dichloro-5-fluorophenyl)-
4H-1,2,4-triazol-3-yl]thio-1-(substituted phenyl) ethanone (7)
which on further cyclization in the presence of PPA yielded
a series of 2,4-dichloro-5-fluorophenyl containing thiazolotria-
zoles (8). The reaction sequence is outlined in Scheme 1.
4.3. Antibacterial studies
The newly prepared compounds were screened for their
antibacterial activity against Escherichia coli, Staphylococcus
aureus, Pseudomonas aeruginosa, Streptococcus pyogenes,
and Klebsiella pneumoniae (recultured) bacterial strains by
the disc diffusion method [31,32]. Compounds 8a, 8cee,
and 8h exhibited good antibacterial activity against all tested
bacterial strains. Compounds 8b, 8f and 8g showed lesser
degree of activity.
3. Results and discussion
4.4. Antifungal studies
In the IR spectrum of compound 8a, the aromatic CeH
stretching frequency was observed at 3098 cmꢀ1. The absorp-
tion band at 1618 cmꢀ1 is due to the presence of eC]Ne
stretch of the triazole and thiazole ring system. The CeF
stretching frequency was observed at 1098 cmꢀ1 and another
absorption band at 823 cmꢀ1 due the CeCl stretching.
The 1H NMR spectrum of 8a, a singlet at d 3.96 was due to
the OeCH3 protons. The four aromatic protons of p-anisyl
moiety resonated as two doublets at d 6.93 and 7.11
(J ¼ 8.2 Hz), respectively. The C3 (HeFmeta) and C6 (He
Fortho) protons of 2,4-dichloro-5-fluorophenyl moiety reso-
nated as a mutiplet between d 7.91 and 8.08. A characteristic
singlet was observed at d 8.19 due to the proton of the thiazole
ring. Thiazole proton resonated as a singlet between d 8.02 and
8.30 for compounds 8beh.
Further evidence for the formation of thiazolotriazole (8a)
was obtained by recording its mass spectrum. The mass spec-
trum of compound 8a showed molecular ion peak at m/z 393,
in conformity with the molecular formula C17H10Cl2FN3OS.
A peak at m/z 189 was due to formation of dichlorofluoroben-
zonitrile cation. The characterization data of thiazolotriazoles
(8aeh) are given in Table 1.
The newly prepared compounds were screened for their an-
tifungal activity against Aspergillus niger, Candida albicans,
Aspergillus fumigatus, Penicillium marneffei, and Trichophy-
ton mentagrophytes (recultured) by agar diffusion method
[33,34]. Compounds 8c, 8e and 8h showed good activity
against all tested fungal strains. Compounds 8a, 8b, 8d, 8f
and 8g showed lesser degree of activity.
5. Conclusion
Anti-inflammatory studies revealed that compounds with 4-
fluorophenyl, 4-chlorophenyl and 2,4-dichloro-5-fluorophenyl
at position 6 of the thiazolotriazole ring system showed excel-
lent anti-inflammatory activity. Analgesic activity studies
revealed that compounds with 4-chlorophenyl, 4-bromophenyl
and 2,4-dichloro-5-fluorophenyl exhibited good analgesic
activity. Compound bearing 4-fluorophenyl, 4-chlorophenyl
and 2,4-dichloro-5-fluorophenyl at position 6 of the thiazolo-
triazole ring system displayed excellent antibacterial activity.
Antifungal screening studies revealed that compounds with
4-chlorophenyl and 2,4-dichloro-5-fluorophenyl showed
excellent antifungal activity. These inferences are based on
preliminary tests, further study in these classes of compounds
are in progress.
4. Pharmacological studies
4.1. Anti-inflammatory studies
6. Experimental section
All compounds were screened for their anti-inflammatory
activity by carrageenan induced rat paw oedema (acute-
inflammation model) as described by Winter et al. [29]. The
anti-inflammatory results revealed that compounds 8c, 8d
and 8h exhibited excellent anti-inflammatory activity whereas
compound 8a showed good anti-inflammatory activity com-
pared to that of the standard drug, Indomethacin. Compounds
8b, 8eeg showed lesser degree of activity.
Melting points were determined by open capillary
method. The IR spectra (in KBr pellets) were recorded
on a Shimadzu FT-IR 157 spectrophotometer. 1H NMR
spectra were recorded (in CDCl3/DMSO-d6) on a Bruker
300/400 MHz NMR spectrometer using TMS as an internal
standard. The mass spectra were recorded on a MASPEC/
FAB mass spectrometer operating at 70 eV. The purity of

M.S. Karthikeyan / European Journal of Medicinal Chemistry 44 (2009) 827e833
829
O
O
F
F
CH3
OH
Haloform
reaction
Cl
Cl
Cl
Cl
2
1
1. Esterification
2. Hydrazinolysis
O
O
F
NH
NH2
F
NHNH2
NH
KSCN
Con. HCl
S
Cl
Cl
Cl
Cl
3
4
NaOH
N
Δ
O
N
F
Br
SH
+
N
H
R
Cl
Cl
6
5
KOH
EtOH
N
N
F
S
R
N
H
Cl
O
Cl
7
PPA
R
F
N
N
Cl
S
N
Cl
8
Where R = 4-OCH3, 4-CH3, 4-F, 4-Cl, 4-Br, 4-NO2, 2,4-Cl2 and 2,4-Cl2-5-F
Scheme 1.
the compounds was checked by thin layer chromatography
(TLC) on silica gel plates using a mixture of petroleum
ether and ethyl acetate (5:5). Iodine was used as visualiz-
ing agent.
6.1. General procedure for the preparation of
2-{[5-(2,4-dichloro-5-fluorophenyl)-4H-1,2,
4-triazol-3-yl]thio}-1-(substituted phenyl)
ethanone (7aeh)
2,4-Dichloro-5-fluorobenzoic acid (2) was obtained from
2,4-dichloro-5-fluoroacetophenone
(1)
according
to
To 0.01 mol of 5-(2,4-dichloro-5-fluorophenyl)-4H-1,2,4-
triazole-3-thiol (5), 0.01 mol of substituted phenacyl bromides
(6) were added in the presence of 0.015 mol of KOH in abso-
lute ethanol and refluxed for 5 h. The reaction mixture was
cooled and poured onto crushed ice. The resulting solid was
filtered, dried and recrystallized from a mixture of ethanol
Ref. [25]. 2,4-Dichloro-5-fluorobenzoyl hydrazide (3) was
prepared according to Ref. [26]. 5-(2,4-Dichloro-5-
fluorophenyl)-4H-1,2,4-triazole-3-thiol (5) was prepared
according to Ref. [27]. Substituted phenacyl bromides (6)
were prepared according to Ref. [28].

830
M.S. Karthikeyan / European Journal of Medicinal Chemistry 44 (2009) 827e833
Table 1
Characterization data of thiazolotriazoles (8aeh)
6.2.3. 6-(4-Chlorophenyl)-2-(2,4-dichloro-5-fluorophenyl)-
1,3-thiazolo[3,2-b]-1,2,4-triazole (8d)
C16H7Cl3FN3S; IR (KBr, n, cmꢀ1): 3090 (AreH str), 2965
(CeH str), 1098 (CeF str), 821 (CeCl str); 1H NMR
(400 MHz DMSO-d6) d: 7.51 (d, 2H, AreH, J ¼ 8.5 Hz),
7.56 (d, 1H, dichlorofluorophenyl protons, J ¼ 6.5 Hz), 7.94
(d, 1H, dichlorofluorophenyl protons, J ¼ 9.6 Hz), 8.15 (s,
1H, thiazole proton), 8.21 (d, 2H, AreH, J ¼ 8.5 Hz); FABMS
m/z: 399 (M þ 1, 36%).
Compd. No.
R
M.p. (ꢁC) Yield (%) Analysis (%)
Found (Calculated)
C
H
N
8a
8b
8c
8d
8e
8f
4-OCH3
4-CH3
4-F
180e82
130e32
207e09
167e69
184e86
138e40
189e91
82
70
68
78
86
65
70
72
51.52
2.41
10.42
(51.78) (2.54) (10.66)
53.59 2.51 10.93
(53.97) (2.65) (11.11)
49.92 1.68 10.80
(50.26) (1.83) (10.99)
47.87 1.65 10.36
(48.18) (1.76) (10.54)
43.07 1.41 9.26
(43.34) (1.58) (9.48)
46.62 1.54 13.45
(46.94) (1.71) (13.69)
44.08 1.14 9.52
(44.34) (1.39) (9.70)
42.16 1.02 9.18
(42.57) (1.11) (9.31)
6.2.4. 6-(4-Bromophenyl)-2-(2,4-dichloro-5-fluorophenyl)-
1,3-thiazolo[3,2-b]-1,2,4-triazole (8e)
4-Cl
C16H7BrCl2FN3S; IR (KBr, n, cmꢀ1): 3096 (AreH str),
4-Br
1
2977(CeH str), 1092 (CeF str), 821 (CeCl str); H NMR
4-NO2
2,4-Cl2
(400 MHz DMSO-d6) d: 7.80 (d, 2H, AreH, J ¼ 8.6 Hz),
8.01e8.05 (m, 2H, dichlorofluorophenyl protons), 8.12 (s,
1H, thiazole proton), 8.23 (d, 2H, AreH, J ¼ 8.6 Hz); FABMS
m/z: 444 (M þ 1, 42%).
8g
8h
2,4-Cl2-5-F 204e06
6.2.5. 2-(2,4-Dichloro-5-fluorophenyl)-6-(4-nitrophenyl)-
1,3-thiazolo[3,2-b]-1,2,4-triazole (8f)
C16H7Cl2FN4O2S; IR (KBr, n, cmꢀ1): 3089 (AreH str),
1
2965 (CeH str), 1110 (CeF str), 832 (CeCl str); H NMR
and dimethylformamide. The yields of compounds (7aeh)
were between 65 and 90%.
(400 MHz DMSO-d6) d: 8.03 (d, 1H, dichlorofluorophenyl
protons, J ¼ 6.8 Hz), 8.07 (d, 1H, dichlorofluorophenyl
protons, J ¼ 9.8 Hz), 8.28 (s, 1H, thiazole proton), 8.43 (d,
2H, AreH, J ¼ 8.5 Hz), 8.58 (d, 2H, AreH, J ¼ 8.5 Hz);
FABMS m/z: 410 (M þ 1, 25%).
6.2. General procedure for the preparation of 2-(2,4-
dichloro-5-fluorophenyl)-6-(4-substituted phenyl)-1,3-
thiazolo[3,2-b]-1,2,4-triazole (8aeh)
To 0.01 mol of 2-{[5-(2,4-dichloro-5-fluorophenyl)-4H-
1,2,4-triazol-3-yl]thio}-1-(substituted phenyl)ethanone (7),
w5 ml of PPA were added and refluxed at 120 ꢁC for 6 h.
The reaction mixture was cooled and poured onto crushed
ice and neutralized by adding NaHCO3. The resulting solid
was filtered, dried and recrystallized from a mixture of ethanol
and dimethylformamide.
6.2.6. 2-(2,4-Dichloro-5-fluorophenyl)-6-(2,4-dichloro-
phenyl)-1,3-thiazolo[3,2-b]-1,2,4-triazole (8g)
C16H6Cl4FN3S; IR (KBr, n, cmꢀ1): 3092(AreH str), 2985(Ce
1
H str), 1112 (CeF str), 827 (CeCl str); H NMR (400 MHz
DMSO-d6) d: 7.94e8.04 (m, 2H, dichlorofluorophenyl and
dichlorophenyl protons), 8.09 (s, 1H, thiazole proton), 8.12e
8.14 (m, 3H, dichlorofluorophenyl and dichlorophenyl protons).
6.2.1. 2-(2,4-Dichloro-5-fluorophenyl)-6-(4-methylphenyl)-
1,3-thiazolo[3,2-b]-1,2,4-triazole (8b)
6.2.7. 2,6-Bis(2,4-dichloro-5-fluorophenyl)[1,3]thia-
zolo[3,2-b][1,2,4]triazole (8h)
C17H10Cl2FN3S; IR (KBr, n, cmꢀ1): 3099 (AreH str),
2975 (CeH str), 1092 (CeF str), 832 (CeCl str); 1H
NMR (400 MHz DMSO-d6) d: 2.31 (s, 3H, CH3), 7.16
(d, 2H, AreH, J ¼ 8.5 Hz), 7.27 (d, 2H, AreH,
J ¼ 8.5 Hz), 7.86 (d, 1H, dichlorofluorophenyl protons,
J ¼ 6.5 Hz), 7.98 (d, 1H, dichlorofluorophenyl protons,
J ¼ 9.6 Hz), 8.15 (s, 1H, thiazole proton); FABMS m/z:
378 (M þ 1, 35%), 189 (dichlorofluorobenzonitrile cation,
25%).
C16H5Cl4F2N3S; IR (KBr, n, cmꢀ1): 3099 (AreH str), 2978
(CeH str), 1098 (CeF str), 821 (CeCl str); 1H NMR
Table 2
Anti-inflammatory activity data of thiazolotriazoles (8aeh)
Compd. No. Dose (mg/kg, p.o.) Increase in paw volume % Inhibition
in ml
of paw oedema
8a
8b
50
50
0.381 ꢂ 0.0016
0.653 ꢂ 0.0016
0.336 ꢂ 0.0023
0.37 ꢂ 0.0031
0.652 ꢂ 0.0020
0.578 ꢂ 0.0026
0.53 ꢂ 0.0028
0.26 ꢂ 0.0020
0.65 ꢂ 0.0096
0.24 ꢂ 0.0014
40.6
0
8c
8d
50
50
48.8
42.2
25
6.2.2. 2-(2,4-Dichloro-5-fluorophenyl)-6-(4-fluorophenyl)-
1,3-thiazolo[3,2-b]-1,2,4-triazole (8c)
8e
50
50
C16H7Cl2F2N3S; IR (KBr, n, cmꢀ1): 3092 (AreH str),
2972 (CeH str), 1112 (CeF str), 827 (CeCl str); 1H
NMR (400 MHz DMSO-d6) d: 7.43e7.47 (m, 2H, Are
H), 8.02e8.06 (m, 3H, AreH), 8.31e8.35 (m, 2H,
dichlorofluorophenyl protons). FABMS m/z: 383 (M þ 1,
25%).
8f
8g
10.9
17.2
59.4
e
50
50
8h
Control
Standarda
a
10 ml/kg
2
62.9
Indomethacin is used as the standard; N ¼ 6 in each group.

M.S. Karthikeyan / European Journal of Medicinal Chemistry 44 (2009) 827e833
831
Table 3
Analgesic activity data of thiazolotriazoles (8aeh)
1.5 mg/kg. The 3rd to 10th groups were administered the
test compounds (at a dose of 50 mg/kg suspended in 10 ml/
kg of 2% gum acacia) orally.
Compd. No.
Dose (mg/kg)
Time of reaction
to pain stimulus
After 30 min of treatment of test compounds, 0.1 ml of 1%
(w/v) carrageenan was injected in the subplantar region of the
left hind paw. The right paw served as a reference to non-
inflammed paw for comparison. The initial paw volume was
measured within 30 s of the injection. The relative increase
in paw volume was measured in control, standard and test
compounds at 1, 2 and 3 h after the carrageenan injection.
The difference between the two readings was taken as the
volume of oedema and the percentage inhibition by the drugs
was calculated using the formula,
at time (h) [s] ꢂ SEM
0
1
3
8a
8b
25
25
25
25
25
25
25
25
10
5
7.8
8.1
8.4
8.2
9.1
8.1
8.9
8.1
8.25
8.5
11.1
8.2
11.96
8.4
8.2
8c
8d
8.3
13.2
14.8
11.2
12.9
13.9
10.9
9.6
8e
8f
8g
8.96
8h
14.21
8.2
16.41
13.6
8.8
Control
Standarda
a
14.3
Percentage of oedema inhibition
Pethidine is used as the standard, N ¼ 6 in each group.
¼ 100 ꢀ ðVtest=VcontrolÞ ꢃ 100;
where Vcontrol ¼ volume of paw oedema in control group;
Vtest ¼ volume of paw oedema in the test compounds in treated
group. The results were expressed as % inhibition of oedema
over the untreated control group. Experiments were approved
by the Institutional ethics committee. The results of anti-
inflammatory studies are given in Table 2.
(400 MHz DMSO-d6) d: 7.94 (d, 1H, dichlorofluorophenyl
protons, J ¼ 9.8 Hz), 8.01 (d, 1H, dichlorofluorophenyl
protons, J ¼ 9.8 Hz), 8.09 (s, 1H, thiazole proton), 8.11e
8.14 (m, 2H, dichlorofluorophenyl protons); FABMS m/z:
452 (M þ 1, 30%).
7. Pharmacological assay
7.2. Analgesic assay
7.1. Anti-inflammatory assay
Albino mice of either sex with weight between 20 and 25 g
were used for analgesic study. The animals were divided in to
10 experimental groups each consisted of 6 animals. Gum
Acacia (2%) was administered to group 1. Group 2 received
Pethidine at a dose 5 mg/kg by intraperitoneal injection. Other
groups were given the test compounds at a dose 25 mg/kg
orally. The animals were housed and fed in laboratory kept
at constant temperature of 22 ꢁC under standard conditions
(12:12 h light-dark cycle, standard pellet diet, tap water). In
this test, reaction of mice to painful stimulus was measured.
Mice were placed on the metal plate heated to 55 ꢂ 0.4 ꢁC
and covered with a glass cylinder (25 cm high, 15 cm in
diameter). The time(s) elapsing to the first pain response
(licking or jumping) was determined by a stop watch and
then recorded as response latency, prior to 60, 180 min
following the p.o. administration of the investigated
Wister albino rats of either sex weighing 180e250 g were
used for the experiment. They were housed in clean polypro-
pylene cages and kept under room temperature (25 ꢁC),
relative humidity (60e70%) in 112 h light-dark cycle. The
animals were given standard laboratory diet and water ad
libitum. Food was withdrawn 12 h before and during experi-
mental hours.
The animals were divided into 10 groups each group
contained 6 animals. A mark was made on the hind paw
(left) just beyond the tibioetarsal junction, so that every
time the paw was dipped in the mercury column up to fixed
mark constant paw volume was ensured. The initial paw
volume of each rat was noted by plethysmometrically (Ugo
Basile, Italy). First group received normal saline and the
second group received Indomethacin orally at a dose of
Table 4
Antibacterial activity data of thiazolotriazoles (8aeh)
Compd. No.
Staphylococcus aureus
Escherichia coli
Pseudomonas aeruginosa
Klebsiella pneumoniae
Streptococcus pyogenes
8a
8b
21 (6.25)
15 (6.25)
18 (12.5)
26 (6.25)
21 (6.25)
12 (12.5)
14 (12.5)
23 (6.25)
23(6.25)
25 (6.25)
13 (12.5)
23 (6.25)
26 (6.25)
24 (6.25)
15 (6.25)
11 (12.5)
28 (6.25)
29 (6.25)
30 (6.25)
14 (6.25)
28 (6.25)
31 (6.25)
29 (6.25)
15 (6.25)
13 (12.5)
30 (6.25)
32 (6.25)
19 (6.25)
15 (6.25)
20 (6.25)
17 (6.25)
19 (6.25)
11 (12.5)
13 (12.5)
19 (6.25)
21 (6.25)
23 (6.25)
14 (12.5)
21 (6.25)
19 (6.25)
25 (6.25)
12 (12.5)
15 (6.25)
24 (6.25)
25 (6.25)
8c
8d
8e
8f
8g
8h
Standarda
MIC values are given in brackets. MIC (mg cmꢀ3) ¼ minimum inhibitory concentration, i.e. the lowest concentration to completely inhibit bacterial growth.
a
Ciprofloxacin is used as the standard, diameter zones of inhibitions in mm.

832
M.S. Karthikeyan / European Journal of Medicinal Chemistry 44 (2009) 827e833
Table 5
Antifungal activity of data of thiazolotriazoles (8aeh)
Compd. No.
Candida albicans
Aspergillus Niger
Aspergillus fumigatus
Trichophyton mentagrophytes
Penicillium marneffei
8a
8b
17 (6.25)
e
20 (6.25)
11 (12.5)
21 (6.25)
24 (6.25)
9 (25)
e
14 (12.5)
8 (25)
17 (6.25)
e
16 (6.25)
29 (6.25)
19 (6.25)
12 (12.5)
17 (6.25)
8 (25)
8c
8d
19 (6.25)
20 (6.25)
14 12.5)
16 (6.25)
10 (25)
18 (6.25)
20 (6.25)
17 (6.25)
19 (6.25)
e
21 (6.25)
19 (6.25)
10 (25)
8e
8f
8g
e
10 (25)
e
18 (6.25)
19 (6.25)
18 (6.25)
12 (12.5)
23 (6.25)
23 (6.25)
14 (12.5)
22 (12.5)
25 (6.25)
8h
Standarda
20 (6.25)
21 (6.25)
Diameter zones of inhibitions in mm. MIC values are given in brackets. MIC (mg cmꢀ3) ¼ minimum inhibitory concentration, i.e. the lowest concentration to com-
pletely inhibit fungal growth.
Amphotericin B is used as the standard; e indicates fungus is resistant to the compounds >100 mg cmꢀ3
a
.
compounds. Institutional ethics committee approved all the
experiments. The analgesic screening results are given in
Table 3.
days. The inhibition zones in diameter were measured and
compared with the controls. The Nutrient Broth, which con-
tained logarithmic serially two-fold diluted amount of test
compound and controls was inoculated with approximately
1.6 ꢃ 104e6 ꢃ 104 cfu cmꢀ3. The cultures were incubated
for 48 h at 35 ꢁC and the growth was monitored. The lowest
concentration (highest dilution) required to arrest the growth
of fungus was regarded as minimum inhibitory concentrations
(MIC). Amphotericin B was used as the standard drug. The di-
ameter of zone of inhibition and minimum inhibitory concen-
tration values are given in Table 5.
7.3. Antibacterial assay
A standard inoculum (1e2 ꢃ 107 cfu/cm3 0.5 McFarland
standards) was introduced onto the surface of sterile agar
plates, and a sterile glass spreader was used for even distribu-
tion of the inoculum. The discs measuring 6.25 mm in diame-
ter were prepared from Whatman no.1 filter paper and
sterilized by dry heat at 140 ꢁC for 1 h. The sterile disc previ-
ously soaked in a known concentration of the test compounds
were placed in nutrient agar medium. Solvent and growth
controls were kept. The plates were inverted and incubated
for 24 h at 37 ꢁC. The inhibition zones were measured and
compared with the controls. Minimum inhibitory concentra-
tion (MIC) was determined by broth dilution technique. The
Nutrient Broth, which contained logarithmic serially two-
fold diluted amount of test compound and controls were inoc-
ulated with approximately 5 ꢃ 105 cfu of actively dividing
bacteria cells. The cultures were incubated for 24 h at 37 ꢁC
and the growth was monitored visually and spectrophotomet-
rically. The lowest concentration (highest dilution) required
to arrest the growth of bacteria was regarded as minimum
inhibitory concentration (MIC). Ciprofloxacin was used as
a standard drug. The diameter of the zone of inhibition and
minimum inhibitory concentration values are given in Table 4.
Acknowledgement
The author is grateful to The Head of the NMR Research
Center, IISc e Bangalore, and CDRI e Lucknow for
1
providing H NMR, and mass spectral data. The author is
also grateful to Dr. N.S. Kumari and Dr. Anoop, Department
of Biochemistry, K.S. Hegde Medical Academy, Deralakatte
for providing screening results. M.S.K. acknowledges the
financial support (SRF) rendered by CSIR, New Delhi.
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