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468 Medicinal Chemistry, 2009,5, 468-473
1573-4064/09 $55.00+.00 © 2009 Bentham Science Publishers Ltd.
Total Synthesis and Analgesic Activity of 6-Fluoroindan-1-acetic Acid and
its 3-Oxo Derivative
Hasina Yasmin1, Sharmistha Das1, Lutfun Nahar2, M. Mehedi Masud3, M. Shafikur Rahman3,
Suvash C. Roy4, M. Mukhlesur Rahman5, Simon Gibbons5, Joydeb K. Kundu6, Bidyut K. Datta6,
Sitesh C. Bachar6, A. K. Azad Chowdhury7 and Satyajit D. Sarker8,*
1Department of Pharmacy, State University of Bangladesh, Dhaka-1209, Bangladesh, 2Drug Discovery and Design Re-
search Division, Department of Pharmacy, School of Applied Sciences, University of Wolverhampton, City Campus
South, Wulfruna Street, Wolverhampton WV1 1LY, UK; 3Department of Pharmaceutical Chemistry, Faculty of Phar-
macy, University of Dhaka, Dhaka 1000, Bangladesh; 4Department of Pharmacy, University of Science & Technology
Chittagong, Foy’s Lake, Chittagong, Bangladesh; 5Centre for Pharmacognosy and Phytochemistry, The School of
Pharmacy, University of London, 29-39 Brunswick Square, London WC1N 1AX, UK; 6Department of Pharmaceutical
Technology, Faculty of Pharmacy, University of Dhaka, Dhaka 1000, Bangladesh; 7Department of Clinical Pharmacy
and Pharmacology, Faculty of Pharmacy, University of Dhaka, Dhaka 1000, Bangladesh; 8Department of Pharmacy,
School of Applied Sciences, University of Wolverhampton, MM Building, Molineux Street, Wolverhampton WV1 1SB,
UK
Abstract: 6-Fluoro-3-oxo-indan-1-acetic acid (5) and 6-fluoroindan-1-acetic acid (6) were conveniently synthesised from
3-fluorobenzaldehyde in four and five steps, respectively. The structures of these new compounds and two other interme-
diates, 3-fluorobenzylidine-bis-acetoacetate (2) and 3-fluoro--phenyl glutaric acid (3) were elucidated by spectroscopic
means, notably, HRMS, 1D and 2D NMR. The analgesic activity of compounds 5and 6 were assessed by the acetic acid
induced writhing in Swiss albino mice.
Key Words: 3-fluorobenzylidine-bis-acetoacetate, 3-fluoro--phenyl glutaric acid,6-fluoro-3-oxo-indan-1-acetic acid, 6-
fluoroindan-1-acetic acid, analgesic activity.
INTRODUCTION
Analgesic and anti-inflammatory drugs that are in use
today show considerable variations in potency, incidence of
side effects and individual patient responses. Historically,
aspirin has been the drug of choice as a general pain-killer
because of its better tolerance, low toxicity and low cost.
Similarly, ibuprofen is the most commonly used first line
analgesic and anti-inflammatory agent as it combines hood
efficacy with a low incidence of side effects [1,2]. However,
its analgesic efficacies are weaker than those of other non-
steroidal anti-inflammatory drugs (NSAIDs). Diclofenac is a
popular alternative as a first or second line analgesic. Indo-
methacin, which apparently has a superior anti-inflammatory
property, is associated with a higher incidence of side effects
and is often used as a third line agent. Azapropazone pro-
duces a similar effect to naproxen, but is associated with a
high incidence of gastrointestinal (GI) complications. The
most significant adverse effects of NSAIDs are GI bleeding
and perforation, which occur in approximately 1% of pa-
tients taking NSAIDs for long term [1]. Because of the major
limitations of existing NSAIDs it has become necessary to
develop more effective and less toxic new analgesic and anti-
inflammatory agents.
*Address correspondence to this author at the Department of Pharmacy,
School of Applied Sciences, University of Wolverhampton, MM Building,
Molineux Street, Wolverhampton WV1 1SB, UK;
E-mail: S.Sarker@wlv.ac.uk
Indan derivatives, particularly those with a carboxylic
acid functionality, possess anti-inflammatory property [2].
For example, 1H-indene-3-acetic acid-5-fluoro-2-methyl-1-
[4-(methylsulfinyl)-phenyl]methylene (Sulindac) and indan-
1,3-dione are well known anti-inflammatory agents [3]. Re-
cently, significant anti-inflammatory activity among a series
of substituted indan-1-carboxylic acids has been reported [4],
a number of methoxyindan-1-alkanoic acids have been syn-
thesised with considerable anti-inflammatory properties [5],
and indan derivatives with a halo-substituted indanyl group
have been found to possess analgesic and anti-inflammatory
properties [6-9]. It has been established that aromatic halo-
gen substitution could increase the analgesic and anti-
inflammatory potency and widen the margin of safety [9]. As
part of our continuing search for new, more effective analge-
sic and anti-inflammatory agents with little or no side-effects
[2, 9], we now report on the total synthesis of 6-fluoroindan-
1-acetic acid (6) from 3-fluorobenzaldehyde, and the as-
sessment of its analgesic activity along with the intermediate
6-fluoro-3-oxo-indan-1-acetic acid (5) using the acetic acid
induced writhing in Swiss albino mice.
CHEMISTRY AND ANALGESIC ACTIVITY
Synthesis and Spectroscopic Identification of 6-fluoro-3-
oxo-indan-1-acetic Acid (5) and 6-fluoro-indan-1-acetic
Acid (6)
6-Fluoroindan-1-acetic acid (6) was conveniently synthe-
sised from 3-fluorobenzaldehyde (1) in five steps with an
Total Synthesis and Analgesic Activity Medicinal Chemistry, 2009, Vol. 5, No. 5 469
overall 30.1% yield (Scheme 1). The four intermediates were
3-fluorobenzylidine-bis-acetoacetate (2), 3-fluoro--phenyl
glutaric acid (3), 3-fluorophenyl succinyl chloride (4) and 6-
fluoro-3-oxo-indan-1-acetic acid (5). The structures of com-
pounds 2, 3, 5 and 6 were elucidated unambiguously by
spectroscopic means, particularly by comprehensive 1D and
2D NMR analyses, e.g. 1H NMR, 13C NMR, 13C DEPT-135,
1H-1H COSY, 1H-1H NOESY, 1H-13C HMQC and 1H-13C
HMBC. The UV spectral analyses of 2, 3, 5 and 6 revealed
the presence of aromaticity in these molecules. The IR spec-
tra revealed the presence of ester and ketonic carbonyl func-
tionalities and a C-F in 2; a C-F and a carboxylic acid
(COOH) moieties in 3; a COOH, a ketonic carbonyl and a C-
F groups in 5; and a COOH and a C-F moieties in 6.
3-Fluorobenzaldehyde (1) was condensed with ethylcya-
noacetate in the presence of piperidine in 1:2 molar ratio
using the Knoevenagel reaction [10] to produce 3-
fluorobenzylidine-bis-acetoacetate (2) with an excellent yield
of 84.5% (Scheme 1). A CI-MS spectrum of 2revealed the
pseudomolecular ion [M+NH4]+ at m/z 384, and an HR-
EIMS showed the molecular ion [M]+ peak at 366.1479, cor-
responding to the molecular formula C19H23FO6. In the 1H
NMR spectrum of 2 (Table 1), there were signals for four
aromatic methines ( 7.04, 7.11, 7.17 and 7.30) correspond-
ing to a 1, 3-di-substituted benzene ring system, three other
methines ( 3.02-3.94), two methyls ( 2.88 / 2.90) assigna-
ble to two Me-CO- functionalities, and two ethoxy groups (
3.84 / 3.80 and 0.85 / 0.95) representing two -COOEt sub-
structure. The 13C NMR (Table 1) displayed signals corre-
sponding to 19 carbons including four aromatic methines (
114.2, 115.10, 124.1 and 130.8), two aromatic quaternary (
139.9 and 161.1), three methines ( 38.0, 42.6 and 43.0), two
ketonic carbonyl carbon (199.7 and 199.9), two ester car-
bonyl (169.8 and 169.9), two oxymethylene (64.3 and
64.4) and four methyl carbons (14.4, 14.4, 14.9 and 15.2).
The 1H and 13C NMR spectra confirmed the identity of 2 as
3-fluorobenzylidine-bis-acetoacetate.
Compound 2 was alkali hydrolysed to 3-fluoro--phenyl
glutaric acid (3; yield 75.3%) (Scheme 1). A CI-MS spec-
trum of 3revealed the pseudomolecular ion [M+NH4]+ at m/z
244, and an HR-EIMS showed the molecular ion [M]+ peak
at 226.0642, corresponding to the molecular formula
C11H11FO4. In the 1H and 13C NMR spectra (Table 1), in ad-
dition to the signals corresponding to a 1,3-disubstituted
benzene ring as in 2, there were signals for a methine (H
3.02-3.94, C 36.3), two methylene (H 3.90, 2.88 and 3.94,
2.90, C 37.2 and 37.3), and two carboxylic acid functionali-
ties (C 172.2 and 170.0). All these signals together with 2D
NMR data analyses confirmed the identity of compound 3 as
3-fluoro--phenyl glutaric acid.
The compound 3was cyclized to 3-fluoro-3-oxo-indan-
acetic acid (5; 62.7 %) via the formation of 3-fluoro--
phenyl glutaryl chloride (4), a moisture sensitive liquid mate-
rial (Scheme 1). Clemmensen reduction of 5 resulted in 6-
fluoroindan-1-acetic acid (6; 75.5 %). A CI-MS spectrum of
5 revealed the [M+NH4]+ ion at m/z 226, and an HR-EIMS
spectrum displayed [M]+ ion at m/z 208.0536 confirming the
molecular formula C11H9FO3. In the 1H NMR spectrum of 5
(Table 1), there were signals corresponding to a 1,3,6-
trisubstituted benzene ring ( 7.07, 7.13 and 7.37), a highly
deshielded signal ( 12.41) for -OH of a carboxylic acid
group, a methine ( 3.04) and two methylene ( 2.20 and
2.44; 3.92) groups. The 13C NMR (Table 1) exhibited signals
for 11 carbons including three aromatic methines ( 112.2,
114.9 and 123.9), three aromatic quaternary ( 130.4, 141.3
and 163.5), a methine ( 40.4), two methylene ( 37.2 and
38.9), and acid carbonyl (70.0) and a ketonic carbonyl car-
bon (196.9). Thus the identity of the new compound 5 was
confirmed as 6-fluoro-3-oxo-indan-1-acetic acid.
A CI-MS spectrum of 6displayed the pseudomolecular
ion [M+NH4]+ at m/z 212, and an HR-EIMS displayed the
molecular ion [M]+ at m/z 194.0743 corresponding to the
molecular formula C11H11FO2. In the 1H and 13C NMR spec-
tra of 6 (Table 1) were similar to those of 5 with the excep-
Scheme 1. Synthesis of compounds 2-6.
470 Medicinal Chemistry, 2009, Vol . 5, N o. 5 Yasm in e t a l.
tions that there were signals for an additional methylene
group (H 2.31 and 2.68, C 29.3), and there were no signal
for any ketone carbonyl carbon. All these signals together
with 2D NMR data analyses confirmed the identity of the
new compound 6 as 6-fluoroindan-1-acetic acid.
Analgesic Activity of 6-fluoro-3-oxo-indan-1-acetic Acid
(5) and 6-fluoro-indan-1-acetic Acid (6)
The analgesic activity of compounds 5and 6 were as-
sessed by the acetic acid induced writhing in Swiss albino
mice [2, 11], because the analgesic and the anti-inflam-
matory activity of various indan-1-acids and tetrazoles were
reported previously [5, 12-15]. The results of the current
study showed that the writhing induced by acetic acid was
significantly reduced by the test compounds in a dose de-
pendent manner (Table 2). While 6-fluoro-3-oxo-indan-1-
acetic acid (5) showed 23.9 and 32.3% (p < 0.0005) inhibi-
tions, respectively, at the doses of 25 and 50 mg/kg-body
weight, 6-fluoroindan-1-acetic acid (6) exhibited 37.7 and
46.7% (p < 0.0005) inhibitions, respectively, at the doses of
25 and 50 mg/kg-body weight. The analgesic activity of 5
and 6 were comparable to those of the positive controls, e.g.
aminopyrine with 47.9% (p < 0.00005) inhibition at 30
mg/kg body weight, indomethacin with 48.5% (p < 0.0005)
inhibition at 8 mg/kg body weight and diclofenac Na with
62.9% (p < 0.0005) inhibition at 10 mg/kg body weight.
None of the test compounds (5 and 6) displayed any signifi-
cant side effects at test doses. However, the b ehavioural pat-
tern of the mice was slightly affected, e.g. reduced move-
ment, head down, and increased respiration.
Table 1. 1H and 13C NMR Data of Compounds 2, 3, 5 and 6
Chemical Shifts
in ppm
1H NMR (Coupling Constant J in Hz) 13C NMR
Position
23 5
6
2356
1 - - 3.04 m
3.00 m
139.9 140.2 40.4 40.6
2 7.04 br d (2.0) 7.01 br d (2.0) 2.88 dd (21.5, 12.5)
2.42 dd (21.5, 6.5)
2.44 m
2.20 m
115.1 115.1 37.2 23.7
3- - -
2.68 m
2.31 m
161.1 161.9 196.9 29.3
4 7.11 dd (8.0, 2.0) 7.12 dd (8.0, 2.0) 7.13 d (8.0)
7.10 d (8.0)
114.2 114.0 123.9 124.0
5 7.30 dd (8.0) 7.32 dd (8.0) 7.37 dd (8.0, 2.2)
7.33 dd (8.0, 2.1)
130.8 130.8 112.0 110.9
6 7.17 dd (8.0, 2.0) 7.16 dd (8.0, 2.0) -
-
124.1 124.1 163.5 163.1
7 3.02-3.94* s 2.46 m 7.07 d (2.2)
7.04 d (2.1)
38.0 36.3 114.9 112.9
8 3.02-3.94* s 3.90 m
2.88 m -
-
42.6 37.2 141.3 138.4
8’ 3.02-3.94* s 3.94 m
2.90 m -
-
43.0 37.3 - -
9 3.02-3.94* s - -
-
169.9 172.0 130.4 132.3
9’ 3.02-3.94* s - -
-
169.8 172.2 - -
10 - - 3.92 d (6.8)
3.91 d (6.8)
199.9 - 38.9 39.0
10’ - - -
-
199.7 - -
11 2.88 s - 12.41 br s
12.41 br s
15.2 - 170.0 171.1
11’ 2.90 s - -
-
14.9 - - -
12 3.84 q (7.0) - -
-
64.4 - - -
12’ 3.80 q (7.0 - -
-
64.3 - - -
13 0.85 t (7.0) - -
-
14.4 - - -
13’ 0.95 t (7.0) - -
-
14.4 - - -
Spectra obtained in DMSO-d6
*Overlapped peaks.
Total Synthesis and Analgesic Activity Medicinal Chemistry, 2009, Vol. 5, No . 5 471
It is noteworthy that the absence of the ketonic carbonyl
functionality at C-3 in 6 increased the cyclopentane ring
flexibility, and contributed to the increased analgesic activity
of 6 measured by % inhibition of induced writhing in mice.
This phenomenon was previously observed with two similar
compounds, 6-fluoro-3-oxo-indan-1-carboxylic acid and 6-
fluoroindan-1-carboxylic acid [2]. It could also be noted that
an increase in carbon number, i.e. acetic acid (-CH2COOH as
in 5 and 6) instead of a carboxylic acid (-COOH) [2] func-
tionality, appeared to have increased the analgesic potency of
such compounds.
The writhing reflex in mice induced by acetic acid is a
sensitive procedure to assess the potential analgesic property
of drugs. It has been suggested that acetic acid acts by releas-
ing endogenous mediators which stimulate the nociceptive
neurons in mice [16]. Acetic acid is sensitive to cyclooxy-
genase inhibitors and has been used to study the effect of
analgesic agents that primarily inhibit the cyclooxygenase
involved in prostaglandin synthesis. Acetic acid is also sensi-
tive to NSAIDs and to narcotics and other centrally acting
drugs [2, 16]. Recently it has been found that the nociceptive
activity of acetic acid may be due to the release of cytokines,
such as TNF-, interleukin-1, and interleukin-8, by resident
peritoneal macrophages and mast cells [17]. In the light of
this report [17], it is reasonable to assume that the antinoci-
ceptive action showed by the compounds 5 and 6 in the ace-
tic acid induced writhing test was probably due to inhibition
of the release of TNF-, interleukin-1, and interleukin-8, by
resident peritoneal macrophages and mast cells.
EXPERIMENTAL SECTION
3.1. General
The chemicals and solvents used in various reactions
were purchased from Merck (Germany); BDH (India), or SD
Fine Chemicals (India). The melting points were determined
by using Adco Melting Point Apparatus and were uncor-
rected. Thin-layer chromatography was performed using
Kieselgel 60 F254 plates (Merck). The absorption maxima
(max) of all the newly synthesised compounds were deter-
mined in absolute methanol by using Genesis-2 spectropho-
tometer. By using 8010M FTIR spectrometer, the character-
istic absorption bands (max) of the newly synthesised com-
pounds were recorded on KBr disk. NMR spectra were re-
corded in DMSO-d6 on a Bruker AVANCE 500 MHz NMR
Spectrometer (500 MHz for 1H and 125 MHz for 13C) using
the residual solvent peaks as internal standard. MS analyses
were performed, on a Finnigan MAT95 spectrometer. HMBC
spectra were optimized for a long range JH-C of 9 Hz and
NOESY experiment was carried out with a mixing time of
0.4 s.
3.2. Synthesis of Compounds 2-6
3.2.1. Synthesis of 3-fluorobenzylidine-bis-acetoacetate (2)
3-Fluorobenzylidine-bis-acetoacetate (2) was prepared
from 3-fluorobenzaldehyde (1) (17.3 g; 0.18 mol) by con-
densation with ethyl acetoacetate (46.8 g; 0.36 mole) in the
presence of piperidine (3.5 mL) in anhydrous condition for
96 h at r.t. following the Knoevenagel reaction [10] (Scheme
1). On completion of the reaction a solid mass was obtained,
which was crushed in a mortar and pestle, washed with ether
to remove piperidine and filtered. The resulting solid was re-
crystallised from acetone-water as compound 3-fluoroben-
zylidine-bis-acetoacetate (2) (55.67 g; mp. 154-156o; yield
84.5%).
Crystalline solid, mp. 154-156 oC. UV (MeOH) max in
nm: 278. IR (KBr) max in cm-1: 1730 (COOEt), 1620
(COMe) and 1115 (C-F). 1H NMR (500 MHz, DMSO-d6)
and 13C NMR (125 MHz, DMSO-d6): Table 1. CI-MS m/z:
[M+NH4]+ 384. HR-EIMS m/z: [M]+ 366.1478 calcd.
366.1479 for C19H23FO6.
3.2.2. Synthesis of 3-fluoro--phenyl Glutaric Acid (3)
Compound 2 (54.9 g, 0.15 mol) was hydrolyzed in pres-
ence of 25% alcoholic solution of KOH (50 gm) by refluxing
for 2.5 h (Scheme 1). The alcohol was then distilled off un-
der reduced pressure, diluted with water, washed with chlo-
roform and neutralised by conc. HCl in cold condition with
constant stirring. The precipitation thus formed was filtered
and recrystallized from alcohol-water to afford 3-fluoro--
phenyl glutaric acid (3) as a crystalline solid (25.53 g; mp.
124-126o; yield 75.3% ).
Table 2. Analgesic Activity of 6-fluoro-3-oxo-indan-1-acetic Acid (5) and 6-fluoroindal-1-acetic Acid (6)
Test Compounds/Controls Group Do se (mg/kg body we ight) Total Number of Writhing Mean± SD % Inhibition
A 25 27, 17, 24, 18, 19, 22 21.17±3.53 23.93a
6-Fluoro-3-oxo-indan-1-acetic acid (5)
B 50 22, 14, 15, 18, 24, 20 18.83±3.57 32.34a
C 25 21, 15, 13, 24, 12, 19 17.33±4.35 37.73a
6-Fluoroindal-1-acetic acid (6)
D 50 14, 10, 23, 16, 16, 10 14.83±4.41 46.71a
Aminopyrine E 30 17, 08, 19, 20, 14, 09 14.50±4.64 47.89a
Indomethacin F 8 12, 14, 18, 09, 13, 20 14.33±3.68 48.50a
Diclofenac Na G 10 07, 14, 14, 05, 10, 12 10.33±3.39 62.88a
Saline H - 32, 30, 28, 25, 28, 24 27.83±2.73 -
a Probability values (calcu lated as compared to control using student’s t-test): < 0.0005.
All values are means of six mice.
472 Medicinal Chemistry, 2009, Vol . 5, N o. 5 Yasm in e t a l.
Crystalline solid, mp. 124-126 oC. UV (MeOH) max in
nm: 280. IR (KBr) max in cm-1: 1690 (COOH) and 1115 (C-
F). 1H NMR (500 MHz, DMSO-d6) and 13C NMR (125
MHz, DMSO-d6): Table 1. CI-MS m/z: [M+NH4]+ 244. HR-
EIMS m/z: [M]+ 226.0642 calcd. 226.0641 for C11H11FO4.
3.2.3. Synthesis of 3-fluorophenyl Succinyl Chloride (4)
Compound 3 (22.6 g; 0.1 mol) was converted to acyl-
chloride (4) by refluxing it with thionyl chloride (sp. gr.
1.631; 23.81 g; 0.2 mol) in benzene (dry, 100 mL) as solvent
for 1.5 h. The benzene and excess thionyl chloride were re-
moved in vaccuo to obtain 3-fluoro--phenyl gluteryl chlo-
ride (4) as a liquid which was used for the next step without
further purification or spectroscopic identification (Scheme
1).
3.2.4. Synthesis of 6-fluoro-3-oxo-indan-1-acetic Acid (5)
Anhydrous aluminium chloride (40.0 g, 0.3 mol) was
added portion-wise to the liquid (4) in a well stirred condi-
tion using CS2 (100 mL) as a solvent. The reaction mixture
was stirred for 2 h at r.t. (Scheme 1), and then decomposed
in ice-water mixture (300 mL). The solvent CS2was evapo-
rated in hot water bath. After cooling off the mixture, the
precipitates were filtered, washed thoroughly with water and
recrystallised from alcohol-water to yield 6-fluoro-3-oxo-
indan-1-acetic acid (5) (13.0 g; mp. 148-150˚; yield 62.68%).
Crystalline solid, mp. 148-150 oC. UV (MeOH) max in
nm: 280. IR (KBr) max in cm-1: 1680 (C=O), 1650 (COOH)
and 1115 (C-F). 1H NMR (500 MHz, DMSO-d6) and 13C
NMR (125 MHz, DMSO-d6): Table 1. CI-MS m/z: [M+NH4]+
226. HR-EIMS m/z: [M]+ 208.0534 calcd. 208.0536 for
C11H9 FO3.
3.2.5. Synthesis of 6-fluoroindan-1-acetic Acid (6)
6-Fluoroindan-1-acetic acid (6) was obtained from 6-
fluoro-3-oxo-indan-1-acetic acid (5) by Clemmensen reduc-
tion (Scheme 1). Compound 5 (10.4 g; 0.05 mol) was treated
with amalgamated zinc (50 g), water (50 mL), conc. HCl (40
mL) and immiscible solvent benzene (60 mL) by refluxing in
a water bath till (18 h) the reaction mixture gave no keto test.
The reduced product was separated in benzene layer and the
aqueous layer was extracted with benzene (203 = 60 mL)
and combin ed with the benzene layer, which was washed
with water and dried over anhydrous sodium sulphate. After
removal of the solvent in vaccuo, a brownish oily liquid was
obtained. Compound 6(7.3 g; mp 48-50°; yield 75.5%) was
obtained as a crystalline solid (from alcohol water) from this
oily liquid.
Crystalline solid, mp. 48-50 oC. UV (MeOH) max in nm:
280. IR (KBr) max in cm-1: 1650 (COOH) and 1115 (C-F).
1H NMR (500 MHz, DMSO-d6) and 13C NMR (125 MHz,
DMSO-d6): Table 1. CI-MS m/z: [M+NH4]+ 212. HR-EIMS
m/z: [M]+ 194.0742 calcd. 194.0743 for C11H11 FO2.
3.3. Assessment of analgesic activity
The analgesic activity of the compounds 5 and 6, and the
positive controls, aminopyrine (BDH, Germany), indo-
methacin (BDH, India) and diclofenac Na (BDH, Germany)
was studied by acetic acid induced writhing test as described
by Vogel and Vogel [11] with little modification.
3.3.1. Animals
Young Swiss albino mice aged 4-5 weeks weighed 20-25
g of either sex were used for the assessment of analgesic
activity. They were collected from the animal house of the
International Center for Diarrheal Diseases and Research,
Bangladesh (ICDDR,B), Mohakhali, Dhaka. The mice were
kept in groups of 6 in plastic polyvinyl cages (BIK indus-
tries, India) having dimensions of (28 22 13) cm3. The
animals were given standard mice feed delivered by ICDDR’B
and water ad libitum. They were kept in the laboratory envi-
ronment for seven days maintaining light and dark; were
fasted overnight and weighed before the experiment.
3.3.2. Test Compounds and Positive Controls
6-Fluoro-3-oxo-indan-1-acetic acid (5) and 6-fluoro-
indan-1-acetic acid (6) were weighed in 20 mg each and
taken into separate graduated test tubes. The compounds
were then dissolved in 2 mL of saline solution and a few
drops of 0.1N NaOH in saline. The pH of the solution was
adjusted to 7.4±0.2 by drop wise addition of 0.1N HCl in
saline. Then the final volumes of the solutions were adjusted
to 10 mL with saline water.
The solutions of the positive controls aminopyrine (BDH,
Germany), indomethacin (BDH, India) and diclofenac Na
(BDH, Germany) were prepared as follows. Each of these
drugs (5 mg) was dissolved separately in 2 mL of saline so-
lution and 2-3 drops of 0.1N NaOH in saline. The pH of the
solution was adjusted to 7.4±0.2 by drop wise addition of
0.1N HCl in saline. Finally, the volume was adjusted to 6
mL with saline.
3.3.3. Protocol
The mice were randomly divided into eight groups which
consisted of 6 mice in each group. Groups A and B received
the test compound 5, and groups C and D received the test
compound 6. All test compounds were administered orally
with a help of a feeding needle at doses of 25 and 50 mg/kg
body weight in the groups, respectively. Mice groups E, F
and G received positive controls, aminopyrine 30 mg/kg
body weight, diclofenac Na 10 mg/kg body weight and in-
domethacin 8 mg/kg body weight, respectively. Group H
was kept as negative control giving saline solution only. A
forty minutes interval was allowed to ensure proper absorp-
tion of the administered compounds. Then the writhing in-
ducing chemical, acetic acid solution (0.7%, 0.1 ml/10 g),
was administered intraperitoneally (i.p.) to each of the ani-
mals of a group. After an interval of ten minutes numbers of
writhing were counted for another 10 min. The average per-
cent decrease in writhing was calculated and comp ared
against the control (saline treated) group. Percent inhibition
was calculated using the following formula.
% Inhibition = [(Wc– Wt) / Wc] x 100
Where, Wc = Average writhing counted for control group;
Wt = Average writhing calculated for individual test group.
ACKNOWLEDGEMENTS
One of the authors (SCB) is grateful to the Ministry of
Science, Information, Communication and Technology,
Total Synthesis and Analgesic Activity Medicinal Chemistry, 2009, Vol. 5, No . 5 473
Government of Bangladesh, Dhaka for a grant (No. BTAJO-
PROMA/Sha-9/B.:ANU.:PRO.:/2007-2008/BS-74/118) to
perform some parts of the project. The mass spectral analy-
ses were carried out in the EPSRC National Mass Spec-
trometry Service Centre, Swansea, and the NMR spectros-
copy was performed in the School of Pharmacy, University
of London.
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Received: 27 April, 2009 Revised: 01 July,2009 Accepted: 01 July,2009
