Discovery of novel phenoxyacetic acid derivatives as antimycobacterial agents

Article abstract of DOI:10.1016/j.bmc.2007.01.006

A series of 2-{4-[1-amino (thioxo) methyl-5-(substituted phenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid and 2-{4-[1-carbamoyl-5-(substituted phenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid were synthesized and the in vit

Full text of DOI:10.1016/j.bmc.2007.01.006

Bioorganic & Medicinal Chemistry 15 (2007) 1896–1902
Discovery of novel phenoxyacetic acid derivatives
as antimycobacterial agents
Mohamed Ashraf Ali and Mohammad Shaharyar*
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Jamia Hamdard University,
Hamdard Nagar, New Delhi 110062, India
Received 8 November 2006; revised 5 January 2007; accepted 6 January 2007
Available online 10 January 2007
Abstract—A series of 2-{4-[1-amino (thioxo) methyl-5-(substituted phenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid and 2-{4-[1-carbamoyl-5-(substituted phenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid were synthesized
and the in vitro activity of the synthesized compounds against Mycobacterium tuberculosis H37Rv (MTB) and INH-resistant
M. tuberculosis (INHR-MTB) was studied. Among the synthesized compounds, compound (3f) 2{-[4-(1-carbamoyl-5-(chlorophe-
nyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid was found to be the most active against M. tuberculosis H37Rv
(MTB) and INH-resistant M. tuberculosis (INHR-MTB) with minimum inhibitory concentration of 0.06 lg/ml.
ꢀ 2007 Elsevier Ltd. All rights reserved.
1. Introduction
time. The synthesis of novel pyrazoline derivatives and
investigation of their chemical and biological behavior
have gained more importance in recent decades for bio-
logical, medicinal, and agricultural reason. Living
organism finds difficulty in construction of NAN bonds
that limits the natural abundance of compounds having
such bonds. Pyrazoline and their derivatives, a class of
compounds containing the NAN bond, exhibit a wide
range of biological activities. The current work describes
the synthesis of novel pyrazoline moiety with encourag-
ing antimycobacterial activity against M. tuberculosis
H37Rv and INH-resistant M. tuberculosis (INHR-
MTB).
Mycobacterium tuberculosis is the primary cause of mor-
tality due to an infectious disease in the world today.
Mycobacteria are ubiquitous organisms that are becom-
ing increasingly important intracellular pathogen that
establishes an infection in oxygen-rich macrophage of
the lung.1 The emergence of AIDS, decline of socioeco-
nomic standards, and a reduced emphasis on tuberculo-
sis control programs contribute to the disease’s
resurgence in industrialized countries.2 Resistance of
M. tuberculosis strains to antimycobacterial agents is
an increasing problem worldwide.3–5 However, powerful
new anti-TB drugs with new mechanism of action have
not been developed in the last 40 years. In spite of severe
toxicity on repeated dosing isoniazid (INH) is still con-
sidered to be a first-line drug for chemotherapy of tuber-
culosis.6 Literature survey reveals pyrazoline derivatives
are active against many Mycobacteria.7–9 The current
work describes the synthesis of novel pyrazoline moiety
with encouraging antimycobacterial activity against
M. tuberculosis H37Rv and INH-resistant M. tuberculo-
sis (INHR-MTB). The chemistry of heterocyclic com-
pounds has been an interesting field of study for long
2. Results and discussion
2.1. Chemistry
The synthesis of phenoxyacetic acid derivatives has been
carried out following the steps shown in the Scheme 1.
In the initial step, chalcones (2a–k) were synthesized
by condensing 2-(4-formyl-2-methoxyphenoxy)acetic
acid with appropriate acetophenone in dilute ethanolic
potassium hydroxide solution by Claisen–Schmidt con-
densation. The compounds (3a–k) and (4a–k) were syn-
thesized by reacting chalcones with thiosemicarbazide
and semicarbazide in glacial acetic acid (reaction time
varies from 4 to 7 hrs). The purity of the compounds
was checked by single-spot TLC, and the compounds
Keywords: Mycobacterial agents; Phenoxy acetic acid.
*
Corresponding author. Tel.: +91 9899452373; fax: +91 11
26059663; e-mail: yarmsy@rediffmail.com
0968-0896/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.01.006

M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
1897
CH3
O
A
CHO
CH3
a
O
OH
+
b
R1-COCH3
HO
R1
O
O
O
O
1
O
2a-k
CH
3
O
OH
O
O
S
N
N
3a-k
H N
2
R
1
CH3
O
B
CHO
CH3
a
O
OH
+
b
R1-COCH3
HO
R1
O
O
O
O
1
O
2a-k
CH
3
O
OH
O
O
O
N
N
4a-k
H N
2
R
1
Scheme 1. Protocol for synthesis. (A) Reagents: (a) KOH/EtOH; (b) NH2CSNHNH2/CH3COOH. (B) Reagents: (a) KOH/EtOH; (b)
NH2CONHNH2/CH3COOH.
1
of this study were identified by spectral data (IR, H
NMR, and mass). Spectral data of all the newly synthe-
sized compounds were in full agreement with proposed
structures. In general, Infra Red Spectra (IR) revealed
COOH, OH, C@O, C@N, CAN and C@S peak at
3317, 3212, 1682, 1540, 1320, and 1130 cmÀ1, respective-
ly. In the Nuclear Magnetic Resonance Spectra (1H
NMR) the signals of the respective protons of the pre-
pared titled compounds were verified on the basis of
their chemical shifts, multiplicities, and coupling
constants. The spectra showed a singlet at d 3.0 ppm
2.2. Antimycobacterial activity
All newly synthesized compounds were screened for
their antimycobacterial activity against M. tuberculosis
H37Rv using BACTEC-460 radiometric system and
INH-resistant M. tuberculosis (INHR-MTB) using agar
dilution method and MICs of the compounds are
reported in Tables 1 and 2 with standard drug INH
for comparison. Among the 22 newly synthesized
compounds seven compounds were found to be the most
active compounds with minimum inhibitory concentra-
tion of less than 1 lg/ml. The newly synthesized
compounds, compound 2-{4-[1-amino(thioxo)methyl-5-(4-
chlorophenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxy-
phenoxy}acetic acid (3f) was found to be the most
active agent against M. tuberculosis H37Rv (MTB) and
INH-resistant M. tuberculosis (INHR-MTB) with mini-
mum inhibitory concentration of 0.06 lg/ml. Compounds
with electron-withdrawing group substituted analogues
showed more activity.
corresponding to methylene group; singlet at
d
3.4 ppm corresponding to C4 methylene group; singlet
at d 3.8 ppm corresponding to methoxy group; singlet
at d 6.0 ppm corresponding to C5 CH group; singlet at
d 7.13 ppm corresponding to NH2 and multiplet at d
7.2–8.3 ppm for aromatic proton; singlet at d 8.8 ppm
corresponding to OH group; singlet at d 10.83 ppm
corresponding to COOH group. The elemental analysis
results were within 0.4% of the theoretical values.

1898
M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
Table 1. Physical constants and antimycobacterial activity of the synthesized compounds
CH
3
O
OH
O
O
S
N
N
3a-k
H N
2
R
1
Compound
R
Yield (%)
Mp (ꢁC)
Mol. formula
C19H19N3O4S
Mol. wt
(MIC) lg/ml
MTBa
MTBb
3a
3b
3c
3d
3e
3f
Phenyl–
94
78
66
80
82
86
94
86
80
78
65
—
119
166
110
140
138
186
152
146
194
212
112
—
385.43
401.43
430.43
403.42
399.46
419.88
419.88
400.45
399.46
401.43
430.43
—
6.25
0.20
6.32
0.96
6.15
0.06
0.12
6.25
0.20
6.25
0.96
6.15
0.06
0.12
12.5
6.25
0.58
3.12
1.36
4-Hydroxy phenyl–
4-Nitro phenyl–
4-Fluoro phenyl–
4-Methyl phenyl–
4-Chloro phenyl–
2-Chloro phenyl–
4-Amino phenyl
Benzyl
C
19H19N3O5S
C19H18N4O6S
19H18N3O4SF
C20H21N3O4S
C
C19H18N3O4SCl
C19H18N3O4SCl
3g
3h
3i
C
19H20N4O4S
12.5
C20H21N3O4S
C19H19N3O5S
C19H18N4O6S
—
6.25
0.58
6.25
0.06
3j
2-Hydroxyphenyl
3-Nitro phenyl–
—
3k
INH
a Mycobacterium tuberculosis H37Rv.
b INH-resistant Mycobacterium tuberculosis.
Table 2. Physical constants and antimycobacterial activity of the synthesized compounds
CH
3
O
OH
O
O
O
N
N
4a-k
H N
2
R
1
Compound
R
Yield (%)
Mp (ꢁC)
Mol. formula
Mol. wt
(MIC) lg/ml
MTBa
MTBb
4a
4b
4c
4d
4e
4f
Phenyl–
86
94
86
80
78
65
66
80
82
86
94
—
119
166
110
140
138
186
152
146
194
212
112
—
C19H19N3O5
C19H19N3O6
369.37
385.37
414.37
387.36
383.40
403.81
403.81
384.38
383.40
385.37
414.37
—
6.25
1.23
6.25
3.16
4.12
0.13
1.26
5.32
6.25
0.58
6.25
0.06
6.12
1.23
6.25
3.25
6.25
0.13
1.26
5.42
6.25
0.58
6.25
1.36
4-Hydroxy phenyl–
4-Nitro phenyl–
4-Fluoro phenyl–
4-Methyl phenyl–
4-Chloro phenyl–
2-Chloro phenyl–
4-Amino phenyl–
Benzyl
C
19H18N4O7
C19H18N3O5F
C20H21N3O5
C19H18N3O4Cl
4g
4h
4i
C
C
20H17N3O4 Cl
19H20N4O5
C20H22N3O5
C19H19N3O6
C19H18N4O7
4j
2-Hydroxyphenyl
3-Nitro phenyl–
—
4k
INH
—
a Mycobacterium tuberculosis H37Rv.
b INH-resistant Mycobacterium tuberculosis.
Among the 2-{4-[1-amino(thioxo)methyl-5-(substituted
phenyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}-
acetic acid (3a-k) and 2-{4-(1-carbamoyl-5-(substituted
phenyl)-4,5-dihydro-1H-3-pyrazolyl)-2-methoxyphenoxy}-
acetic acid (4a–k) substituent with electron-withdrawing
group, such as chlorine (3f) and (3g), hydroxy (3b) en-
hanced the antimycobacterial activity. However the elec-
tron withdrawing group such as 4-fluoro phenyl and

M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
1899
nitrophenyl-substituted analogue produced moderate
inhibitory activity. On the other hand, the electron-do-
nating group substituted analogue showed low to mod-
erate inhibitory activity. Instead of (C@O) substitution
at N-1 (C@S) group substitution in pyrazoline analogue
improves the antitubercular activity. These reports
clearly showed that the increases in the presence of N-
1 (C@S) group substitution with 4-chlorophenyl substi-
tution at C-5 pyrazoline (3a–k) derivatives 3b, 3g, and
3f cause remarkable improvement in antimycobacterial
activity. All the newly synthesized compounds were test-
ed for cytotoxicity (1C50) in VERO cells at concentra-
tions 62.5 lg/ml or 10 times. After 72 hrs exposure,
viability was assessed on the basis of cellular conversion
of MTT into a formazan product using the Promega
Cell Titer 96 non-radioactive cell proliferation method.
Most of the active compounds were found to be non-
toxic up to 62.5 lg/ml. Screening of the antitubercular
activity of these novel phenoxy acetic acid derivatives
identified compounds endowed with antimycobacterial
activity, exhibiting MIC values less than 0.5 lg/ml. It
is conceivable that derivatives showing more potency,
selectivity, and low toxicity of these derivatives make
them excellent leads for synthesizing novel derivatives
for antimycobacterial activity, can be further modified
to exhibit better potency than the standard drugs. Fur-
ther studies to acquire more information about Quanti-
tative Structure–Activity Relationships (QSAR) are in
progress in our laboratory. The pyrazoline derivatives
discovered in this study may provide valuable therapeu-
tic intervention for the treatment of antitubercular
diseases.
3.1.1. 2-{2-Methoxy-4-[(Z)-3-oxo-3-phenyl-1-propenyl]phen-
oxy}acetic acid (2a). IR (KBr) cmÀ1: 3210 (OH), 1682
(C@O), 3040 (CH). 1H NMR (DMSO-d6) ppm: 10.1
(1H, s, COOH), 6.8–7.9 (7H, m, aromatic), 6.7, 7.07
(2· 1H, dd, –CH@CH, J = 9, 9 Hz), 3.7 (3H, s,
OCH3), 2.8 (2H, s, CH2).
3.1.2. 2-{4-[(Z)-3-(4-hydroxyphenyl)-3-oxo-1-propenyl]-
2-methoxyphenoxy}acetic acid (2b). IR (KBr) cmÀ1
:
3202 (OH), 1680 (C@O), 3042 (CH). 1H NMR (DMSO-
d6) ppm: 10.1(1H, s, COOH), 8.8 (1H, s, OH), 6.8-7.82
(7H, m, aromatic), 6.7, 7.07 (2· 1H, dd, –CH@CH,
J = 9, 9 Hz), 3.8 (3H, s, OCH3), 2.7 (2H, s, CH2).
3.1.3. 2-{2-Methoxy-4-[(Z)-3-(4-nitrophenyl)-3-oxo-1-prop-
enyl]phenoxy}acetic acid (2c). IR (KBr) cmÀ1: 3232
1
(OH), 1680 (C@O), 3044 (CH); H NMR (DMSO-d6)
ppm: 10.1 (1H, s, COOH), 8.8 (1H, s, OH),
6.8–8.26 (8H, m, aromatic), 6.7, 7.07 (2· 1H, dd,
–CH@CH, J = 9, 8.7 Hz), 3.8 (3H, s, OCH3), 2.9 (2H,
s, CH2).
3.1.4. 2-{4-[(Z)-3-(4-fluorophenyl)-3-oxo-1-propenyl]-2-meth-
oxyphenoxy}acetic acid (2d). IR (KBr) cmÀ1: 3216 (OH),
1
1680 (C@O), 3040 (CH); H NMR (DMSO-d6) ppm:
10.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.8–7.78 (7H,
m, aromatic), 6.7, 7.07 (2· 1H, dd, –CH@CH, J = 9,
8.9 Hz), 3.8 (3H, s, OCH3), 2.9 (2H, s, CH2).
3.1.5. 2-{2-Methoxy-4-[(Z)-3-(4-methylphenyl)-3-oxo-1-prop-
enyl]phenoxy}acetic acid (2e). IR (KBr) cmÀ1: 3200
1
(OH), 1680 (C@O), 3040 (CH); H NMR(DMSO-d6)
ppm: 10.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.8–7.92
(7H, m, aromatic), 6.7, 7.07 (2· 1H, dd, –CH@CH,
J = 9, 9 Hz), 3.9 (3H, s, OCH3), 2.2 (2H, s, CH2), 2.4
(3H, s, CH3).
3. Experimental
All the chemicals were supplied by E. Merck (Germa-
ny) and S.D. Fine Chemicals (India). Melting points
were determined by open tube capillary method and
are uncorrected. Purity of the compounds was checked
on thin-layer chromatography (TLC) plates (silica gel
G) in the solvent system toluene–ethyl formate–formic
acid (5:4:1) and benzene–methanol (8:2), the spots
were located under iodine vapors or UV light. IR
spectra were obtained on a Perkin-Elmer 1720 FT-
3.1.6. 2-{4-[(Z)-3-(4-chlorophenyl)-3-oxo-1-propenyl]-2-
:
3200(OH), 1680 (C@O), 3040 (CH). H NMR(DMSO-
d6 ppm): 10.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.8–
7.86 (7H, m, aromatic), 6.7, 7.07 (2· 1H, dd, –CH@CH,
J = 9, 9 Hz), 3.8 (3H, s, OCH3), 2.9 (2H, s, CH2).
methoxyphenoxy}acetic acid (2f). IR (KBr) cmÀ1
1
3.1.7. 2-{4-[(Z)-3-(2-chlorophenyl)-3-oxo-1-propenyl]-2-
methoxyphenoxy}acetic acid (2g). IR (KBr) cmÀ1: 3200
1
IR spectrometer (KBr Pellets). H NMR spectra were
1
recorded on a Bruker AC 300 MHz spectrometer
using TMS as internal standard in DMSO/CDCl3.
Mass spectra were recorded on a Bruker Esquire
LCMS using ESI.
(OH), 1680 (C@O), 3040 (CH); H NMR (DMSO-d6)
ppm: 10.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.8–7.78
(7H, m, aromatic), 6.7, 7.07 (2· 1H, dd, –CH@CH,
J = 9, 9 Hz), 3.8 (3H, s, OCH3), 2.9 (2H, s, CH2).
3.1. General procedure for synthesis of chalcones
3.1.8. 2-{4-[(Z)-3-(4-aminophenyl)-3-oxo-1-propenyl]-2-meth-
oxyphenoxy}acetic acid (2h). IR (KBr) cmÀ1: 3200 (OH),
1680 (C@O), 3040 (CH); H NMR (DMSO-d6) ppm:
10.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.6–7.23 (7H,
m, aromatic), 5.12 (2H, s, NH2), 6.7, 7.07(2· 1H, dd,
–CH@CH, J = 9, 9 Hz), 3.8 (3H, s, OCH3), 2.9 (2H, s,
CH2).
1
2-(4-Formyl-2-methoxyphenoxy)acetic acid (0.01 mole)
was dissolved in ethanol. Then, a solution of potassium
hydroxide (30%, 5 ml) and suitable acetophenone in
10 ml of Petroleum ether was added to the resulting
solution with continuous stirring. The resulting solution
was allowed to stand overnight. After 4 hrs stirring then
the solution was poured into ice-cold water and then
neutralized with HCl. The solid separate was filtered
off, dried, and purified by ethanol affording (2a–k) in
64–95% yield.
3.1.9. 2-{2-methoxy-4-[(Z)-3-oxo-4-phenyl-1-butenyl]phen-
oxy}acetic acid (2i). IR (KBr) cmÀ1: 3200 (OH), 1680
(C@O), 3040 (CH); 1H NMR (DMSO-d6) ppm:
10.1(1H, s, COOH), 8.8 (1H, s, OH), 6.8–7.70 (7H, m,

1900
M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
aromatic), 5.12 (2H, s, NH2), 6.02, 7.27 (2· 1H, dd,
–CH@CH, J = 9, 9 Hz), 3.8 (3H, s, OCH3), 2.9 (2H, s,
CH2), 2.4 (2H, s, CH2).
d6) ppm: 10.87 (1H, s, COOH), 7.21–7.51 (7H, m, aro-
matic), 6.86 (2H, s, NH2), 6.0 (1H, s, CH), 3.4 (3H, s,
OCH3), 3.0 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2): ESI-MS: m/z: 403 (M+).
3.1.10. 2-{4-[(Z)-3-(2-hydroxyphenyl)-3-oxo-1-propenyl]-
2-methoxyphenoxy}acetic acid (2j). IR (KBr) cmÀ1: 3200
(OH), 1680 (C@O), 3040 (CH); H NMR(DMSO-d6)
ppm: 12.1 (1H, s, COOH), 8.8 (1H, s, OH), 6.8–7.70
(7H, m, aromatic), 5.12 (2H, s, NH2), 6.02, 7.27 (2·
1H, dd, –CH@CH, J = 6, 6 Hz), 3.8 (3H, s, OCH3),
2.9 (2H, s, CH2), 2.4 (2H, s, CH2).
3.2.5. 2-{4-[1-Amino (thioxo) methyl-5-(4-methylphenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3e). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1590 (C@N), 1320 (CAN) 1130 (C@S); 1H NMR
(DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.21–7.51 (7H,
m, aromatic), 6.86 (2H, s, NH2), 6.0 (1H, s, CH), 3.4
(3H, s, OCH3) 3.2 (3H, s, CH3), 2.7 (2H, s, CH2, C4 meth-
ylene), 2.7 (2H, s, CH2): ESI-MS: m/z: 400 (M+1).
1
3.1.11. 2-{2-Methoxy-4-[(Z)-3-(3-nitrophenyl)-3-oxo-1-
propenyl] phenoxy}acetic acid (2k). IR (KBr) cmÀ1
:
3200 (OH), 1680 (C@O), 3040 (CH). 1H NMR
(DMSO-d6 ppm): 12.1 (1H, s, COOH), 8.8 (1H, s,
OH), 6.8–8.82 (7H, m, aromatic), 5.12 (2H, s, NH2),
6.02, 7.27 (2· 1H, dd, –CH@CH, J = 9, 9 Hz), 3.8
(3H, s, OCH3), 2.9 (2H, s, CH2), 2.4 (2H, s, CH2).
3.2.6. 2-{4-[1-Amino (thioxo) methyl-5-(4-chlorophenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3f). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1
1590 C@N), 1320 (CAN) 1130 (C@S), 776(CACl); H
NMR(DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.21–
7.51 (7H, m, aromatic), 6.86 (2H, s, NH2), 6.0 (1H, s,
CH), 3.4 (3H, s, OCH3), 3.1 (2H, s, CH2, C4 methylene),
2.7 (2H, s, CH2): ESI-MS: m/z: 419 (M+).
3.2. General procedure for the synthesis of 2-{4-[1-amino
(thioxo) methyl-5-(substituted phenyl)-4,5-dihydro-1H-3-
pyrazolyl]-2-methoxyphenoxy}acetic acid (3a–k)
3.2.7. 2-{4-[1-Amino (thioxo) methyl-5-(2-chlorophenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3g). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
To a solution of (0.01 mole) chalcones (2a–k) in ethanol
(20 ml) was added thiosemicarbazide (0.02 mole) in gla-
cial acetic acid and the reaction mixture was refluxed for
4 hrs. The reaction mixture was cooled and then poured
onto crushed ice. Then solid mass separated out was fil-
tered, washed with water, and purified by ethanol (3a–k).
1
1590(C@N), 1320 (CAN) 1130 (C@S), 776(CACl); H
NMR (DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.21–
7.51 (7H, m, aromatic), 6.86 (2H, s, NH2), 6.0 (1H, s,
CH), 3.4 (3H, s, OCH3), 3.1 (2H, s, CH2, C4 methylene),
2.7 (2H, s, CH2): ESI-MS: m/z: 420 (M+1).
3.2.1. 2-{4-[1-Amino (thioxo) methyl-5-phenyl-4,5-dihy-
dro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (3a).
IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590
3.2.8. 2-{4-[5-(4-Aminophenyl)-1-amino (thioxo) methyl-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy} acetic
acid (3h). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1590 (C@N), 1320 (CAN) 1130 (C@S); 1H NMR
(DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.21–7.51 (7H,
m, aromatic), 6.86 (2H, s, NH2), 6.82 (2H, s, NH2) 6.0
(1H, s, CH), 3.4 (3H, s, OCH3), 3.1 (2H, s, CH2, C4 meth-
ylene), 2.7 (2H, s, CH2): ESI-MS: m/z: 401 (M+1).
1
(C@N), 1320 (CAN) 1130 (C@S); H NMR (DMSO-
d6) ppm: 10.87 (1H, s, COOH), 6.83–7.51 (8H, m, aro-
matic), 5.34 (2H, s, NH2), 5.82 (1H, s, CH), 3.8 (3H,
s, OCH3), 3.3 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2): ESI-MS: m/z: 385 (M+).
3.2.2. 2-{4-[1-Amino(thioxo)methyl-5-(4-hydroxyphenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3b). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1590 (C@N), 1320 (CAN) 1130 (C@S); 1H NMR
(DMSO-d6) ppm: 10.82 (1H, s, COOH), 9.2 (1H, s,
OH), 7.21–7.51 (7H, m, aromatic), 5.4 (2H, s, NH2),
5.8 (1H, s, CH), 3.4 (3H, s, OCH3), 3.2 (2H, s, CH2,
C4 methylene), 2.7 (2H, s, CH2): ESI-MS: m/z: 401
(M+).
3.2.9. 2-{4-[1-Amino (thioxo) methyl-5-benzyl-4,5-dihy-
dro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (3i).
IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590
1
(C@N), 1320 (CAN) 1130 (C@S); H NMR (DMSO-
d6) ppm: 10.87 (1H, s, COOH), 7.21–7.51 (7H, m, aro-
matic), 6.86 (2H, s, NH2), 6.0 (1H, s, CH), 3.4 (3H, s,
OCH3), 3.1 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2), 2.1(2H, s, benzylCH2): ESI-MS: m/z: 399 (M+).
3.2.3. 2-{4-[1-Amino (thioxo) methyl-5-(4-nitrophenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3c). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1590 (C@N), 1320 (CAN) 1130 (C@S); 1H NMR
(DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.41–8.22
(7H, m, aromatic), 6.86 (2H, s, NH2), 6.0 (1H, s, CH),
3.9 (3H, s, OCH3), 3.1 (2H, s, CH2, C4 methylene), 2.7
(2H, s, CH2): ESI-MS: m/z: 431 (M+1).
3.2.10. 2-{4-[1-Amino (thioxo) methyl-5-(2-hydroxyphe-
nyl)-4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}-
acetic acid (3j). IR (KBr) cmÀ1: 3320 (NH2), 3307
1
(COOH), 1590 (C@N), 1320 (CAN) 1130 (C@S); H
NMR (DMSO-d6) ppm: 10.87 (1H, s, COOH), 9.4 (1H,
s, OH), 7.21–7.51 (7H, m, aromatic), 6.86 (2H, s, NH2),
6.0 (1H, s, CH), 3.4 (3H, s, OCH3), 3.1 (2H, s, CH2, C4
methylene), 2.7 (2H, s, CH2): ESI-MS: m/z: 401 (M+).
3.2.4. 2-{4-[1-Amino(thioxo)methyl-5-(4-fluorophenyl)-4,5-
dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid
(3d). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590
3.2.11. 2-{4-[1-Amino (thioxo) methyl-5-(3-nitrophenyl)-
4,5-dihydro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic
acid (3k). IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH),
1590 (C@N), 1320 (CAN) 1130 (C@S); 1H NMR
1
(C@N), 1320 (CAN) 1130 (C@S); H NMR (DMSO-

M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
1901
1
(DMSO-d6) ppm: 10.87 (1H, s, COOH), 7.21–8.34 (7H,
m, aromatic), 6.86 (2H, s, NH2), 6.0 (1H, s, CH), 3.4
(3H, s, OCH3), 3.1(2H, s, CH2, C4 methylene), 2.7
(2H, s, CH2): ESI-MS: m/z: 430 (M+).
1320 (CAN), 776(CACl); H NMR (DMSO-d6) ppm:
10.87 (1H, s, COOH), 7.21–7.51 (7H, m, aromatic),
6.21 (2H, s, NH2), 4.87 (1H, s, CH), 3.4 (3H, s,
OCH3), 3.1 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2): ESI-MS: m/z: 403 (M+).
3.3. General procedure for synthesis of 2-{4-(1-carbamo-
yl-5-(substituted phenyl)-4,5-dihydro-1H-3-pyrazolyl)-2-
methoxyphenoxy}acetic acid (4a–k)
3.3.7. 2-{4-[1-Carbamoyl-5-(2-chlorophenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4g). IR
(KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N),
1
To a solution of (0.01 mole) chalcone (2a–k) in ethanol
(20 ml) was added semicarbazide (0.02 mole) in glacial
acetic acid and the reaction mixture was refluxed for
4 hrs.The reaction mixture was cooled and then poured
onto crushed ice. Then solid mass separated out was fil-
tered, washed with water, and purified by ethanol (4a–k).
1320 (CAN), 776(CACl); H NMR (DMSO-d6) ppm:
10.87 (1H, s, COOH), 7.21–7.51 (7H, m, aromatic),
6.12 (2H, s, NH2), 4.83 (1H, s, CH), 3.4 (3H, s,
OCH3), 3.1 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2): ESI-MS: m/z: 403 (M+).
3.3.8. 2-{4-[5-(4-Aminophenyl)-1-carbamoyl-4,5-dihydro-
1 H-3-pyrazolyl]-2-methoxy phenox y}acetic acid (4 h).
IR (KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590
3.3.1. 2-{4-(1-carbamoyl-5-phenyl-4,5-dihydro-1H-3-pyr-
azolyl)-2-methoxyphenoxy}acetic acid (4a). IR (KBr)
cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N), 1320
(CAN); 1H NMR (DMSO-d6) ppm: 10.87 (1H, s,
COOH), 6.83–7.51 (8H, m, aromatic), 6.12 (2H, s,
NH2), 4.81 (1H, s, CH), 3.8 (3H, s, OCH3), 3.3 (2H,
s, CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-MS: m/z:
369 (M+).
1
(C@N), 1320 (CAN); H NMR (DMSO-d6)ppm: 10.87
(1 H, s, COOH), 7.21–7.51 (7H, m, aromatic), 6.12
(2H, s, 2· NH2), 6.0 (1H, s, CH), 3.4 (3H, s, OCH3),
3.1 (2H, s, CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-
MS: m/z: 385 (M+1).
3.3.9. 2-{4-(5-Benzyl-1-carbamoyl-4,5-dihydro-1H-3-pyr-
azolyl)-2-methoxyphenoxy}acetic acid (4i). IR (KBr)
cmÀ1: 3320 (NH2), 3307 (COOH), 590 (C@N), 1320
(CAN); 1H NMR (DMSO-d6) ppm: 10.87 (1H, s,
COOH), 7.21–7.51 (7H, m, aromatic), 6.14 (2H, s,
NH2), 4.83 (1H, s, CH), 3.4 (3H, s, OCH3), 3.1 (2H,
s, CH2, C4 methylene), 2.7 (2H, s, CH2), 2.1 (2H, s,
benzylCH2): ESI-MS: m/z: 383 (M+).
3.3.2. 2-{4-[1-Carbamoyl-5-(4-hydroxyphenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4b). IR
(KBr) cmÀ1: 3322 (NH2), 3307 (COOH), 1590 (C@N),
1
1320 (CAN); H NMR (DMSO-d6) ppm: 10.82 (1H, s,
COOH), 9.2 (1H, s, OH), 7.21–7.51(7H, m, aromatic),
5.98 (2H, s, NH2), 4.85 (1H, s,CH), 3.4 (3H, s, OCH3),
3.2(2H, s, CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-
MS: m/z: 385 (M+).
3.3.10. 2-{4-[1-Carbamoyl-5-(2-hydroxyphenyl)-4,5-dihy-
dro-1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4j).
3.3.3. 2-{4-[1-Carbamoyl-5-(4-nitrophenyl)-4,5-dihydro-1H-
3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4c). IR
(KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N),
IR (KBr) cmÀ1
:
3310 (NH2), 3317 (COOH),
15,800 (C@N), 1320 (CAN); 1H NMR (DMSO-d6)
ppm: 10.87 (1H, s, COOH), 9.4 (1H, s, OH),
7.21–7.51 (7H, m, aromatic), 6.16 (2H, s, NH2),
4.83 (1H, s, CH), 3.4 (3H, s, OCH3), 3.1 (2H, s,
CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-MS:
m/z: 385 (M+).
1
1320 (CAN); H NMR (DMSO-d6) ppm: 10.84 (1H, s,
COOH), 7.41–8.22 (7H, m, aromatic), 6.14 (2H,
s, NH2), 4.86 (1H, s, CH), 3.9 (3H, s, OCH3), 3.1 (2H,
s, CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-MS: m/z:
415 (M+1).
3.3.4. 2-{4-[1-Carbamoyl-5-(4-fluorophenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4d). IR
(KBr) cmÀ1: 3310 (NH2), 3317 (COOH), 1590 (C@N),
3.3.11. 2-{4-[1-Carbamoyl-5-(3-nitrophenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4k). IR
(KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N),
1
1
1320 (CAN); H NMR (DMSO-d6) ppm: 10.85 (1H, s,
1320 (CAN); H NMR(DMSO-d6) ppm: 10.82 (1H, s,
COOH), 7.21–7.51 (7H, m, aromatic), 6.14 (2H, s, NH2),
4.84 (1H, s, CH), 3.4 (3H, s, OCH3), 3.0 (2H, s, CH2, C4
methylene), 2.7 (2H, s, CH2): ESI-MS: m/z: 387 (M+).
COOH), 7.21–8.34 (7H, m, aromatic), 6.16 (2H,
s, NH2), 4.86 (1H, s, CH), 3.4 (3H, s, OCH3), 3.1 (2H,
s, CH2, C4 methylene), 2.7 (2H, s, CH2): ESI-MS: m/z:
414 (M+).
3.3.5. 2-{4-[1-Carbamoyl-5-(4-methylphenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4e). IR
(KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N),
3.4. Biology
1
1320 (CAN); H NMR (DMSO-d6) ppm: 10.87 (1H,
The primary screening was conducted at a concentra-
tion of 6.25 lg/ml (or molar equivalent of highest
molecular weight compound in a series of congeners)
against M. tuberculosis H37Rv (ATCC27294) and
INH-resistant M. tuberculosis in BACTEC 12B medi-
um using the BACTEC 460 radiometric system.10,11
Compound demonstrating at least 90% inhibition in
the primary screen is re-examined at lower concentra-
tion (MIC) in broth micro dilution assay with Almar
s, COOH), 7.21–7.51 (7H, m, aromatic), 6.00 (2H, s,
NH2), 4.81 (1H, s, CH), 3.4 (3H, s, OCH3), 3.2 (3H,
s, CH3), 2.7 (2H, s, CH2, C4 methylene), 2.7 (2H, s,
CH2): ESI-MS: m/z: 384 (M+1).
3.3.6. 2-{4-[1-Carbamoyl-5-(4-chlorophenyl)-4,5-dihydro-
1H-3-pyrazolyl]-2-methoxyphenoxy}acetic acid (4f). IR
(KBr) cmÀ1: 3320 (NH2), 3307 (COOH), 1590 (C@N),

1902
M. A. Ali, M. Shaharyar / Bioorg. Med. Chem. 15 (2007) 1896–1902
blue. The MIC was defined as the lowest concentra-
tion inhibiting 99% of the inoculum. Concurrent with
the determination of MICs, compounds were tested
for cytotoxicity (IC50) in VERO cells at concentration
equal to and greater than the MIC for M. tuberculosis
H37Rv and INH-resistant M. tuberculosis. After 72 h
exposure, viability was assessed on the basis of cellular
conversion of MTT into a formazan product using the
Promega cell Titer 96 non-radioactive cell proliferation
assay.12
References and notes
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3. Sbarbaro, J. A. Chest 1997, 111, 1149.
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Osahan, S. S.; Ebrahimzadeh, A.; Frieden, T. R. Arch.
Intern. Med 1997, 157, 531.
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H. Pneumologie. 1996, 50, 21, 1987.
6. Kucukguzel, S. G.; Rollas, S. Farmaco 2002, 57, 583.
7. Shaharyar, M.; Siddiqui, A. A.; Ali, M. A.; Sriram, D.;
Yogeeswari, P. Bioorg. Med. Chem. Lett. 2006, 16, 4571.
8. Nauduri, D.; Reddy, G. B. Chem. Pharm. Bull. (Tokyo)
1998, 46, 1254.
9. Santilli, A. A.; Kim, D. H.; Gregory, F. J. J. Pharm. Sci.
1975, 64, 1057.
10. Collins, L.; Frazblau, S. G. Antimicrob. Agents. Chemo-
ther. 1997, 41, 1004.
11. Gundersen, L. L.; Nissen-Meyer, J.; Spilsberg, B. J. Med.
Chem. 2002, 45, 1383.
12. Heifets, L. B.; Flory, M. A.; Lindholm-Levy, P. J.
Antimicrob. Agents. Chemother. 1989, 33, 1252.
Acknowledgments
The author (M. Shaharyar) expresses thanks to Uni-
versity Grant Commission, New Delhi, India, for the
research award, we thank the Tuberculosis Antimicro-
bial Acquisition and Coordinating Facility (TAACF),
National Institute of Allergy and Infectious Diseases
Southern Research Institute/GW Long Hansen’s Dis-
ease Center, Colorado State University Birmingham,
Alabama, USA, for the in vitro antimycobacterial
screening.