Synthesis of novel pyrazolyl tetrazoles as selective COX-2 inhibitors

Article abstract of DOI:10.1007/s00044-013-0500-0

A series of novel pyrazolyl tetrazoles were synthesized by introducing tetrazole moiety at the fourth position of 1,3-substituted pyrazole nucleus. Synthesis was carried out by cyclization of different pyrazolonitriles using sodium azide in the presence of triethylammonium chloride as phase transfer catalyst. The structures of the synthesized compounds were confirmed on the basis of physical and spectral data. Among the synthesized compounds, 4b and 4e displayed significant anti-inflammatory activity with no observable ulcerogenic effect when compared with diclofenac sodium. Furthermore, compounds 4b and 4e were found to have COX-2 selectivity with a ratio of 0.44 and 0.48, respectively.

Full text of DOI:10.1007/s00044-013-0500-0

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2013) 22:4886–4892
DOI 10.1007/s00044-013-0500-0
ORIGINAL RESEARCH
Synthesis of novel pyrazolyl tetrazoles as selective COX-2
inhibitors
•
Kolli Sri Swetha Ravula Parameshwar
•
•
B. Madhava Reddy V. Harinadha babu
Received: 13 October 2012 / Accepted: 16 January 2013 / Published online: 31 January 2013
Ó Springer Science+Business Media New York 2013
Abstract A series of novel pyrazolyl tetrazoles were syn-
thesized by introducing tetrazole moiety at the fourth position
of 1,3-substituted pyrazole nucleus. Synthesis was carried out
by cyclization of different pyrazolonitriles using sodium azide
in the presence of triethylammonium chloride as phase
transfer catalyst. The structures of the synthesized compounds
were confirmed on the basis of physical and spectral data.
Among the synthesized compounds, 4b and 4e displayed
significant anti-inflammatory activity with no observable
ulcerogenic effect when compared with diclofenac sodium.
Furthermore, compounds 4b and 4e were found to have
COX-2 selectivity with a ratio of 0.44 and 0.48, respectively.
COX-1 is predominantly expressed ubiquitously and con-
stitutively, and it serves a housekeeping role in stomach
and kidneys. By contrast, COX-2 is involved in the pro-
duction of prostaglandin, mediating pain and supporting
the inflammatory process. Gastric side effects associated
with classical NSAIDs are attributed to the nonselective
inhibition of the isoenzymes. Even the selective COX-2
inhibitors have been associated with the unexpected car-
diovascular adverse effects. Thus, development of a reli-
able and selective anti-inflammatory agent with a fewer
side effects is a major challenge to medicinal chemists.
Pyrazole template is considered as an important chemical
entity of various physiological significances and pharmaceu-
tical utility. Pyrazoles have drawn a greater attention due to
their wide range of therapeutic activities including anticon-
vulsant (Abdel et al., 2009), antidepressant (Abdel et al.,
2009), antitumor (Peng-cheng et al., 2010), (Christodoulou
et al., 2010), (Lin et al., 2007), antimicrobial (Bondock et al.,
2010), (Radi et al., 2010), ACE inhibitor (Menichini et al.,
2010), antiviral (Osma et al., 2009), anti-inflammatory
(Barsoum and Girgis, 2009), (Adnan et al., 2004) etc.
Keywords Pyrazoles ꢀ Tetrazoles ꢀ
Anti-inflammatory activity ꢀ Acute ulcerogenicity ꢀ
In vitro COX inhibitor study
Introduction
Currently marketed non steroidal anti-inflammatory drugs
(NSAIDs) have been associated with several side effects
like gastrointestinal mucosal damage, bleeding, intoler-
ance, renal toxicity, and hepatotoxicity. The pharmaco-
logical effects of NSAIDs occur due to inhibition of the
membrane enzyme called cyclooxygenase (COX-1 and
COX-2), which is involved in prostaglandin biosynthesis.
The well known selective COX-2 inhibitor, Celecoxib, and
non steroidal antiphlogistic drug, Lonazolac, are good exam-
ples of pyrazoles containing anti-inflammatory drugs. Accord-
ing to literature, tetrazole and its derivatives are known to
possess antibacterial (Adnan et al., 2004), anti-inflammatory
(Umarani et al., 2010), antifungal (Ram Shankar et al., 2004),
cyclo-oxygenase inhibitor (Navidopour et al., 2006), hypo-
glycemic (Gao et al., 2010), and anticancer (Gundugola et al.,
2010) activities.
Electronic supplementary material The online version of this
article (doi:10.1007/s00044-013-0500-0) contains supplementary
material, which is available to authorized users.
Our current strategyconsistsof designing a selective COX-
2 inhibitor with a gastric safety profile which may allow the
use of these new agents for long-term prophylactic use in
certain chronic inflammatory diseases. Since tetrazole is
bioisostere for –COOH group and is also metabolically more
K. S. Swetha ꢀ R. Parameshwar ꢀ B. M. Reddy ꢀ V. H. babu (&)
Medicinal Chemistry Division, G. Pulla Reddy College
of Pharmacy, Mehdipatnam, Hyderabad 500028, India
e-mail: parmi_pharma@yahoo.com
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Med Chem Res (2013) 22:4886–4892
4887
stable than the carboxylic acid group (Gupta et al., 1999), in
our present study, we made an attempt to synthesize title
compounds as per Scheme 1 by replacing–CH2COOH func-
tion in Lonazolac with tetrazole moiety in order to have better
and more potent compounds which are not reported so far.
recorded on Varian 400 MHz spectrometer using DMSO-d6
as solvent and TMS as an internal standard. 13C NMR was
recorded on Varian Gemini 400 MHz Spectrophotometer and
mass spectra on Agilent 6430 triple quadruple LC–MS sys-
tem. Thin layer chromatography was performed using
E.Merck 0.25 mm silica gel plates, and visualization of spots
was accomplished using UV light (256 nm).
Experimental
General procedure for the synthesis of 1-phenyl-3-
Melting points (oC) were determined on Analab melting point
apparatus by open capillary method and were uncorrected.
The IR spectra were recorded on Shimadzu FTIR spectrom-
eter using 1 % potassium bromide disks. 1H NMR was
substituted phenyl-1H-pyrazole-4-carbaldehyde (1a–i)
A mixture of substituted acetophenone (0.01 mol) and
phenyl hydrazine (0.01 mol) was taken in 15–20 ml of
Scheme 1 General procedure for the synthesis of 5-(1-phenyl-3-substituted phenyl-1H-pyrazol-4-yl)-2H-tetrazoles (4a–i)
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Med Chem Res (2013) 22:4886–4892
ethanol containing catalytic amounts of glacial acetic acid
and heated at 50–60 °C for 1 h till a precipitate of phenyl
hydrazone was obtained. The separated precipitate was fil-
tered and recrystallized from ethanol. To an ice-cold solution
of DMF (0.1 mol), phosphorus oxychloride (0.012 mol) was
added drop-wise while maintaining the temperature below
10 °C. To this mixture, ice-cold solution of phenyl hydra-
zone (0.01 mol) in DMF was added lots wise, and stirring
was continued under ice-cold condition for another half an
hour. The reaction mixture was brought to room temperature
and refluxed at 60–70 °C for 4–5 h. The reaction mixture
was then cooled and poured into crushed ice with stirring and
neutralized with aq. NaHCO3 solution. The solid obtained
was filtered and recrystallized from 90 % ethanol.
150.08; Mass: m/z 288.6 [M?]; Anal. Calcd. for C16H12N6
(288.6): C, 66.56; H, 4.20; N, 29.15; Found: C, 66.04; H,
4.68; N, 29.25.
5-[3-(4-Chlorophenyl)-1-phenyl-1H-pyrazol-4-yl]-2H-
tetrazole (4b)
Light brown powder; m.p.230–232 °C, yield, 66 %.
IR[KBr] cm-1: 3,421, 1,653, 1,608, 1,595, 748;1H NMR
(DMSO-d6): d 9.12 (s, 1H, NH in pyrazole ring), 7.96 (d,
2H, J = 7.85 Hz, Ar–H), 7.79 (d, 2H, J = 8.60 Hz, Ar–H),
7.60 (t, 2H, J = 7.82 Hz, Ar–H), 7.30 (d, 2H, J = 8.21 Hz,
Ar–H), 7.43(t, 1H, J = 7.43 Hz, Ar–H); 13C NMR(DMSO-
d6):d 119.45, 128.05, 129.11, 130.40, 130.43, 131.02,
131.50, 134.09, 139.33, 149.87; Mass: m/z 332.8 [M?];
Anal. Calcd. for C16H11ClN6 (332.8): C, 67.54; H, 4.67; N,
27.80. Found: C, 66.94; H, 4.68; N, 28.25.
General procedure for the synthesis of 1-phenyl-3-
substituted phenyl-1H-pyrazole-4-carbaldehyde
oxime (2a–i)
5-[3-(2-Chlorophenyl)-1-phenyl-1H-pyrazol-4-yl]-2H-
A mixture of compound 1 (0.01 mol), hydroxylamine
hydrochloride (0.02 mol), and sodium hydroxide (0.02 mol)
was taken in ethanol and heated to reflux for 2 h. The reac-
tion mixture was cooled, added to water, and neutralized
with 0.1 N HCl. The separated solid was filtered and re-
crystallized from 90 % ethanol.
tetrazole (4c)
Light brown powder; m.p.236–238 °C, yield, 62 %.
IR[KBr] cm-1: 3,424, 1,657, 1,602, 1,592, 738; H NMR
1
(DMSO-d6): d 9.16 (s, 1H, NH in pyrazole ring),7.92 (d, 2H,
J = 7.80 Hz, Ar–H), 7.70 (d, 2H, J = 8.64 Hz, Ar–H), 7.63
(t, 2H, J = 7.89 Hz, Ar–H), 7.36 (d, 2H, J = 8.20 Hz, Ar–
H), 7.42(t, 1H, J = 7.42 Hz, Ar–H);13CNMR(DMSO-d6):d
119.30, 128.16, 129.12, 130.53, 130.66, 131.21, 131.62,
134.16, 139.12, 149.93; Mass: m/z 332.8 [M?]; Anal. Calcd.
for C16H11ClN6 (332.8): C, 67.54; H, 4.67; N, 27.80. Found:
C, 66.92; H, 4.64; N, 28.26.
General procedure for the synthesis of 1-phenyl-3-
substituted phenyl-1H-pyrazole-4-carbonitrile (3a–i)
Compound 2 (0.01 mol) and acetic anhydride (0.01 mol)
were heated strongly for 1 h. The reaction mixture was
cooled to room temperature and added to crushed ice with
stirring. The separated solid was filtered and recrystallized
from 60 % ethanol.
5-[3-(4-Methylphenyl)-1-phenyl-1H-pyrazol-4-yl]-2H-
tetrazole (4d)
General procedure for the synthesis of 5-(1-phenyl-3-
Lightbrown powder; m.p. 220–222 °C, yield, 58 %. IR[KBr]
cm-1: 3,424, 2,932, 1,648, 1,596, 1,500; 1H NMR (DMSO-
d6): d 9.15 (s, 1H, NH in pyrazole ring), 8.13 (d, 2Ar–H,
J = 8.25 Hz),7.80 (d, 2Ar–H, J = 7.43 Hz), 7.62 (m, 3Ar–
H), 7.40 (m, 2Ar–H), 2.41 (s, 3H, CH3);13C NMR (DMSO-
d6):d 21.83, 118.96, 119.62, 120.42, 129.45, 130.12, 130.65,
131.44, 131.76, 133.87, 134.06, 139.46, 150.45; Mass: m/
z 302.3 [M?]; Anal. Calcd. for C17H14N6 (302.3): C, 67.54; H,
3.44; N, 26.04. Found: C, 67.94; H, 4.28; N, 26.85.
substituted phenyl-1H-pyrazol-4-yl)-2H-tetrazole (4a–i)
A mixture of carbonitrile 3 (0.01 mol), sodium azide
(0.02 mol), triethylamine hydrochloride (0.02 mol) was
taken in 15 ml of DMF and heated for reflux for 1–4 days by
monitoring continuously with thin layer chromatogra-
phy(TLC). The mixture was cooled and added to 50 ml of
water and acidified with 0.1 N HCl. The separated solid was
filtered and recrystallized from chloroform–ethanol mixture.
5-[3-(4-Methoxyphenyl)-1-phenyl-1H-pyrazol-4-yl]-2H-
5-(1,3-Diphenyl-1H-pyrazol-4-yl)-2H-tetrazole (4a)
tetrazole (4e)
Light brown powder; m.p. 210–212 °C, yield, 60 %. IR
[KBr] cm-1: 3,412, 1,656, 1,623, 1,596; 1H NMR (DMSO-
d6): d 9.14(s, 1H, NH in pyrazole ring), 7.05–8.02 (m, 9H,
Ar–H); 13C NMR (DMSO-d6):d 119.26, 120.12, 128.21,
129.05, 130.12, 130.28, 131.20, 131.66, 134.76, 139.42,
Yellow powder; m.p.240–242 °C, yield, 56 %. IR [KBr] cm-1
:
3,418, 1,651, 1,596, 1,500, 1,249, 1,180;1H NMR (DMSO-
d6): d 9.05(s, 1H, NH in pyrazole ring), 7.92 (d, 2Ar–H,
J = 7.85 Hz), 7.70 (d, 2Ar–H, J = 8.99 Hz), 7.62 (t,
2Ar–H, J = 8.09 Hz), 7.41(t, 1Ar–H, J = 7.43 Hz), 7.05
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Med Chem Res (2013) 22:4886–4892
4889
(d, 2Ar–H, J = 8.60 Hz), 3.82(s, 3H, OCH3); 13C
NMR(DMSO-d6):d 56.82, 119.12, 120.44, 120.68, 129.58,
130.42, 130.75, 131.76, 131.94, 134.92, 140.12, 148.62, 150.46
Mass: m/z 318.3 [M?]; Anal. Calcd. for C17H14N6O (318.3): C,
64.14; H, 4.43; N, 26.40. Found: C, 64.04; H, 4.88; N, 26.95.
d6):d 9.04(s,1H, NH in pyrazole ring), 7.02–8.06(m, 12H, Ar–
H); 13C NMR(DMSO-d6): 119.08, 120.20, 120.44, 120.86,
128.44, 129.36, 130.45, 130.86, 131.60, 131.86, 134.66,
139.32, 150.86; Mass: m/z 338.3 [M?]; Anal. Calcd. for
C20H14N6 (338.3): C, 70.99; H, 4.17; N, 24.84. Found: C,
71.24; H, 4.25; N, 24.65.
5-[3-(2-Methoxyphenyl)-1-phenyl-1H-pyrazol-4-yl]-
2H-tetrazole (4f)
Biological activity
Yellow powder; m.p.244–246 °C, yield, 60 % IR [KBr]
cm-1: 3,422, 1,649, 1,593, 1,506, 1,238, 1,162;1H NMR
(DMSO-d6): d 9.18(s, 1H, NH in pyrazolering), 8.16 (d, 2Ar–
H, J = 7.82 Hz), 7.84 (d, 2Ar–H, J = 8.96 Hz), 7.66 (t, 2Ar–
H, J = 8.12 Hz), 7.39 (t, 1Ar–H, J = 7.47 Hz) , 7.03 (d,
2Ar–H, J = 8.68 Hz), 3.86(s, 3H, OCH3); 13C NMR(DMSO-
d6):d 55.68,119.14,120.44,120.67, 129.68, 130.67, 130.86,
131.26, 131.98, 134.56, 139.66, 151.46 Mass: m/z 318.3
[M?]; Anal. Calcd. for C17H14N6O (318.3): C, 64.14; H, 4.43;
N, 26.40. Found: C, 64.10; H, 4.62; N, 26.66.
Anti-inflammatory activity
Adult albino rats of both sexes weighing between 120 and
150 g were used and divided into 11 groups of six animals
each. The control group was given 1 % aq. CMC. Dic-
lofenac sodium (20 mg/kg, standard) and test compounds
(4a–i, 20 mg/kg) in 1 % aq. CMC were administered orally
1 h before induction of inflammation. Induction of inflam-
mation was performed by s.c injection of 0.1 ml of freshly
prepared 1 % carrageenan in saline solution, into the sub-
plantar region of the right hind paw. The paw volume was
measured using the plethysmometer at 0.5, 1, 2, 3 h after the
carrageenan challenge. The average value of edema was
calculated by taking the average of six animals at different
hours. Percentage inhibition of edema was calculated for
each group with respect to the control group.
4-[1-Phenyl-4-(2H-tetrazol-5-yl)-1H-pyrazol-3-yl]
phenol (4g)
Brown powder; m.p.250–252 °C, yield, 55 %.IR[KBr]
1
cm-1: 3,442, 3,316, 1,657, 1,596, 1,519; H NMR (DMSO-
d6): d 9.14 (s, 1H, NH in pyrazole ring), 7.98 (d, 2H,
J = 7.85 Hz, Ar–H), 7.82 (d, 2H, J = 8.60 Hz, Ar–H), 7.76
(t, 2H, J = 7.82 Hz, Ar–H), 7.56 (d, 2H, J = 8.21 Hz, Ar–
H), 7.43 (t, 1H, J = 7.43 Hz, Ar–H); 13C NMR(DMSO-d6):d
119.23, 120.28, 120.88, 130.08, 130.66, 130.89, 131.22,
131.86, 134.12, 139.86, 146.12, 150.35; Mass: m/z 304.3
[M?]. Anal. Calcd. for C16H12N6O (304.30): C, 63.15; H,
4.17; N, 24.84. Found: C, 63.74; H, 4.98; N, 23.65.
Inflammation was expressed as the change in paw
volume
Edema ¼ Tt ꢁ T0
where
T0 Volume at ‘0’ h
Tt Volume at ‘t’ h
2-[1-Phenyl-4-(2H-tetrazol-5-yl)-1H-pyrazol-3-yl]
Percentage reduction ¼ ðV0 ꢁ VtÞ=V0 ꢂ 100
where,
phenol (4h)
Brown powder; m.p.254–256 °C, yield, 50 %.IR[KBr]
V0 Volume of the paw of control at time ‘t’
1
cm-1: 3,412, 3,386, 1,649, 1,598, 1,509; H NMR(DMSO-
Vt Volume of the paw of drug treated at time ‘t’
d6): d 9.12 (s, 1H, NH in pyrazole ring), 7.89 (d, 2H,
J = 7.88 Hz, Ar–H), 7.84 (d, 2H, J = 8.65 Hz, Ar–H), 7.72
(t, 2H, J = 7.86 Hz, Ar–H), 7.48 (d, 2H, J = 8.28 Hz, Ar–
H), 7.44 (t, 1H, J = 7.46 Hz, Ar–H);13C NMR(DMSO-d6):d
119.03,120.42, 120.58, 130.18, 130.86, 130.92, 131.42,
131.66, 134.32, 139.87, 148.22, 150.85; Mass: m/z 304.3
[M?]. Anal. Calcd. for C16H12N6O (304.3): C, 63.15; H,
4.17; N, 24.84. Found: C, 63.46; H, 4.84; N, 23.64.
Acute ulcerogenicity study
Young adult albino rats weighing 180–220 g were used for
the experiment, and animals were divided into groups of six
animals each. Rats were fasted 20 h before drug adminis-
tration. The test compounds, diclofenac sodium and cele-
coxib, were given orally in doses of 100, 50, and 50 mg/kg,
respectively, while the control group received only vehicle,
i.e., 1 % CMC. Rats were fasted for 2 h, allowed to feed for
2 h, and then fasted for another 20 h. Rats were given
another two doses in the second and third day. In the fourth
day, rats were sacrificed, the stomach was removed, opened
5-[3-(2-Naphthyl)-1-phenyl-1H-pyrazol-4-yl]-2H-
tetrazole (4i)
Lightbrown powder; m.p. 250–252 °C, yield, 55 %. IR[KBr]
1
cm-1: 3,426, 1,648, 1,596, 1,500, 1,443; H NMR(DMSO-
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Med Chem Res (2013) 22:4886–4892
along with the greater curvature, and rinsed with 0.9 %
saline solution. The number of mucosal damage (red spots)
was counted,and their severity (ulcerogenic severity) was
graded from 0 to 4 according to the following score
assignment. The following figures were calculated.
seconds and incubated for 5 min at 25 °C. After 5 min
incubation, 20 ll of the colorimetric substrate solution was
added to all the wells, followed by the addition of 20 ll of
arachidonic acid to all the wells. The plates were shaken
gently for few seconds and again incubated for 5 min at
25 °C. The absorbance of all the wells was read at 590 nm
using Thermo make Automatic Ex-Microplate Reader. The
COX inhibition activity (%) was calculated using the fol-
lowing formula:
–
–
–
% Incidence/10 = [no. of rats showing ulcer of any grade
divided by total number of rats in the group 9 100]/10.
Average number of ulcers: number of ulcers in the
group/total number of rats in the group.
COX inhibition activity ð%Þ ¼ T=C ꢂ 100
where T = absorbance of the inhibitor well at 590 nm, and
C = absorbance of the 100 initial activity wells without
inhibitor at 590 nm.
Average severity: R[each ulcer multiplied by its score
of severity]/number of ulcers in the group.
Ulcer index = the sum of above three figures
Score
Results and discussion
Normal (no injury)
Latent small red spot
Wide red spot
0
1
2
3
4
Chemistry
The synthetic methodology followed to obtain the target
compounds is outlined in Scheme 1. Cyclization of different
phenyl hydrazones by Vilsmeier–Haack reaction afforded
1-phenyl-3-substituted phenyl-1H-pyrazole-4-carbaldehydes
(1a–i), which on reaction with hydroxylamine hydrochloride
gave pyrazole carbaldehyde oximes (2a–i). The heating of
oximes with acetic anhydride at 140–150 °C gave pyrazol-
onitriles (3a–i). 1,3 Dipolar cyclization of compounds
(3a–i) with sodium azide in the presence of triethylammo-
nium chloride (acts as base and phase-transfer catalyst) gave
pyrazolyl tetrazoles (4a–i). During the formation of pyraz-
olyl tetrazoles, there was variation in reaction time from 1 to
4 days depending on the substituent present in the phenyl
ring. The conversion of pyrazolonitriles to tetrazoles was
confirmed through IR spectra which showed disappearance
of CN absorption band around 2,200–2,220 cm-1 and
appearance of new band at around 3,400–3,440 cm-1 due to
Slight injury
Severe injury
In vitro COX study
The COX inhibition assay (Syed Nasir et al., 2012) was
performed according to the procedure mentioned in the
‘Colorimetric COX (ovine) inhibitor Assay Kit’ supplied
by Cayman Chemical Company, MI, USA. While the
reaction mixture of 100 % initial activity wells contained
150 ll of assay buffer, 10 ll of heme, and 10 ll of relative
(COX-1or COX-2) enzyme solution, the reaction mixture
of inhibitor wells was made of 150 ll of assay buffer, 10 ll
of heme, 10 ll of enzyme (COX-1 or COX-2), and 10 ll of
the test samples. The plates were carefully shaken for few
1
N–H. H NMR spectrum of compound 4a showed a singlet
Table 1 Anti-inflammatory
Compound
Control
0.5 h
1 h
2 h
3 h
% Inhibition after 3 h
activity of diclofenac sodium
and synthesized compounds
evaluated by carrageenan-
induced paw edema
2.5 0.02 3.75 0.01 4.25 0.07 4.75 0.01
Diclofenac sodium 1.67 0.04 1.15 0.05 0.25 0.03 0.54 0.05 88
4a
4b
4c
4d
4e
4f
1.1 0.06 1.25 0.07 1.45 0.06
1.5 0.01 68
0.9 0.02 0.70 0.05 84
1.0 0.08 1.15 0.03 1.2 0.02 75
1.6 0.06
0.9 0.06
1.2 0.08
1.87 0.06 2.21 0.07 1.20 0.06 1.50 0.03 68
1.78 0.01 1.5 0.06 1.1 0.08 0.88 0.05 81
1.74 0.03 1.35 0.02 1.30 0.06 1.6 0.02 66
0.75 0.06 1.25 0.06 1.3 0.08 1.35 0.05 72
Each value represents
mean SE of six animals;
statistical analysis was done by
student’s t test
4g
4h
4i
0.8 0.01 1.15 0.04 1.25 0.05 1.30 0.08 73
1.78 0.01 2.12 0.04 1.84 0.05 1.45 0.08 70
* P \ 0.05 when compared to
control
Edema (ml) (Mean ES)*
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4891
at d 9.14 due to pyrazole proton, and signals for aromatic
protons appeared between d 7.05 and 8.02, thus confirming
the structure. Mass spectrum of compound 4a displayed a
molecular ion peak at m/z 288.6(M?). The structure was
further supported by 13C NMR, and the signals appearing in
the spectrum account for all the carbon atoms present in the
molecule 4a. Similarly, all other compounds were charac-
terized, and the data were given in the experimental section.
Table 3 COX-2/COX-1 ratio of compounds 4b, 4e and celecoxib
Compound
COX-2/COX-1
4b
0.44
0.48
0.02
4e
Celecoxib
Conclusion
Anti-inflammatory screening
Inthis investigation, ninenew pyrazolyltetrazoles(4a–i) were
synthesized by isosteric replacement of –CH2COOH function
in Lonazolac with tetrazole heterocycle, and the structures of
resulting compounds were characterized on the basis of
spectral data. All the compounds were screened for in vivo
anti-inflammatory studies. Among the nine compounds, 4b
and 4e exhibited most promising activity, and the activity was
comparable with the standard drug, diclofenac sodium.
Moreover, the same compounds 4b and 4e have less ulc-
erogenicity when compared with the diclofenac sodium. In
COX-2/COX-1 selectivity assay, the same compounds exhib-
ited more selectivity toward COX-2 enzyme.
In anti-inflammatory screening, among the nine tetrazole
derivatives, compounds 4b and 4e were found to have good
activity (above 80 % inhibition), which may be attributed
to the presence of p-chloro and p-methoxy substitutions on
phenyl ring at the third position of pyrazole nucleus, and
the results are shown in Table 1.
Ulcerogenicity study
Compounds (4b and 4e) which showed good anti-inflam-
matory activity in in vivo screening were further studied
for gastric ulcerogenic potential in rats. Ulcerogenic effect
was compared with classical NSAID, diclofenac sodium,
and a COX-2 selective inhibitor, celecoxib. The results
revealed that the test compounds were less ulcerogenic
than diclofenac sodium and similar to celecoxib in ulcero-
genic induction. Results are shown in Table 2.
Acknowledgments The authors are thankful to the management of
G. Pulla Reddy college of Pharmacy, Hyderabad, India for providing
facilities. Thanks is also due to University of Hyderabad, India for
providing spectral data and in vitro anti-inflammatory studies.
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Compounds 4b and 4e were evaluated for their selectivity
to inhibition of COX-2 and COX-1 isoenzymes using COX
(ovine) inhibitor screening assay kit. The ratio of IC50 of
COX-2 to IC50 of COX-1 (COX-2/COX-1) would suggest
the selectivity of the compound and hence its gastric lia-
bility. The COX-2/COX-1 ratio of compounds 4b and 4e
clearly indicated their selectivity toward COX-2 enzyme
with a ratio of 0.44 and 0.48, respectively. The results are
shown in Table 3.
Table 2 Ulcer index of synthesized compounds of the most active
compounds, diclofenac sodium and celecoxib
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Compound
%
Average no. of
ulcer
Average Ulcer
severity
Incidence/
10
index
Control
0
0
0
0
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Diclofenac 10
5.60
3.20
4.30
4.60
1.42
1.31
1.70
1.62
17.02
12.51
13.90
14.22
Celecoxib
8
8
8
4b
4e
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