R. Vijay Amirtharaj*, V. Suresh and R. Senthil Kumar
Department of Pharmaceutical chemistry,
JKKMMRF College of Pharmacy, Ethirmedu, Komarapalayam 638183, Namakkal Dt, Tamilnadu, India.
ABSTRACT:
The methanol
extract of aerial part of Clerodendrum inerme were investigated for anti-inflammatory and
analgesic at the dose 200 mg/kg, body weight. The experimental paradigms used
were carrageenan, induced pedal edema for anti-inflammatory
activity, acetic acid induced writhing methods were used to assess
analgesic activity. In acute phase inflammation, a maximum inhibition 60.17% (P
< 0.01) was noted at the dose of 200 mg/kg after 3 h of treatment with
methanol extract of Clerodendrum inerme (MECI) in carrageenan, induced pedal edema
respectively. The extract also produced significant (P < 0.01)
analgesic activity in both paradigms. This study exhibits that the methanol
extracts of aerial part of Clerodendrum inerme possess anti-inflammatory and analgesic
activities.
KEYWORDS: Clerodendrum inerme, Anti-inflammatory, Analgesic
INTRODUCTION:
Inflammation or phlogosis is a pathophysiological
response of living tissue to injuries that leads to the lo-cal accumulation of
plasmatic fluid and blood cells. Although it is a defense mechanism, the
complex events and mediators involved in the inflammatory reaction can be
induced, maintain or aggravate many diseases1. However, studies have
been continuing on inflammatory diseases and the side effects of the currently
available anti-inflammatory drugs pose a major problem during their clinical
use2. Therefore, development of newer and more powerful
anti-inflammatory drugs with lesser side effects is necessary.
Clerodendrum inerme (L.) Gaertn. (Family: Verbenaceae) commonly
known as Lanji, Sankupi, Batraj (Hindi), it is found through-out the India near the
sea, Ceylon3. The plant extract of Clerodendrum inerme are traditionally used for the
treatment of skin diseases and antifungal4. It has also been found
to posses multiple therapeutic properties like hepatoproductive5,
anticarcinogenic6, antilipidperoxidative7 and anti-viral8.
However, no work
has been reported on the anti-inflammatory effects on acute and chronic phases
of inflammation and analgesic of Clerodendrum inerme. Keeping this in view, the present study has
been undertaken to investigate the anti-inflammatory and analgesic potential of
methanol extract of Clerodendrum inerme (MECI)
in experimental animal models.
MATERIALS AND
METHODS:
The plant grows
in all textures of mildly acid to alkaline soil. Annual rainfall in the areas
where Clerodendrum inerme
grows in Puerto Rico ranges from 250 mm to 750 mm. The plant Clerodendrum inerme was
collected from Pichavaram, Cuddalore
dt. of
Tamilnadu, India.
The plant
material was taxonomically identified by Mr. G.V.S. Murthy, Join Director,
Govt. of Tamilnadu, Ministry of Environment and
forest, Botanical Survey of India, South circle, Lowely
Road, Coimbadore-641003. A Voucher specimen (No.BSI/SC/5/23/09-10/TECH.228) has
been preserved in our laboratory. The aerial part were dried under shade and
then powdered with a mechanical grinder and stored in an airtight container.
The dried powder material of the aerial part was extracted with methanol (Yield
12.7%), in a soxhlet apparatus. Phytochemical
screening of the ex-tracts revealed the presence of alkaloids, saponins, flavonoids, triterpenes, tannins and steroids.
Swiss albino
mice of both sex weighing between (18-22 g) and Albino Wistar
rats of the either sex (180-200 g) were used for the present study. They were
maintained under standard environmental conditions and were fed with standard
pellet diet supplied by Venkateshwara Enterprises,
Bangalore, India, and water Ad libitum.
Carrageenan (S.
D. Fine Chemicals Limited, Bombay), Acetic acid were used in the present study
and indomethacin (Re-con, Bangalore), were used as
the standard drugs.
Carrageenan-induced
paw oedema in rats. Male wistar rats were divided
into 5 groups and each group consists of 6 animals. Paw swelling was induced by
sub-plantar injection of 0.1 ml 1% sterile carrageenan in saline into the right
hind paw. Group-I served as control which received the vehicle (5% gum acacia
solution, 5 ml/kg/p.o.). group-II
received the standard drug (Indomethacin, 10 mg/kg/p.o.). Group-III, IV and V were received the plant extract
1,2 and 3 at the dose of 200 mg/kg/p.o.).
All the treatments were given 60 minutes prior to carrageenan injection. The
inflammation was quantified by measuring the volume displaced by the paw, using
a plethysmometer at time 0, 1, 2 and 3 hours after
carrageenan administration. The paw volume was determined and the percentage
inhibition of edema was calculated in comparison to the control group. Inhibition = (1-Vt/Vc)
X 100 Where Vt
= volume of test group, Vc =
volume of control group9-11.
MECI
at the dose of 200 mg/kg and combination of above doses of extract with the
standard drug 10 mg/kg (Acetic acid induced writhing response in mice) were
administered to five groups of six mice in each paradigm.
Acetic acid-induced writhing test. Swiss
albino mice were divided into 5 groups and each group consists of 6 animals. Nociception was induced by an intraperitoneal
injection of 0.6% acetic acid (0.1 ml/10 g of body weight). Group-I served as
control which received the vehicle (5% gum acacia solution, 5 ml/kg/p.o.). Group-II received the standard drug (Indomethacin, 10 mg/kg/p.o.).
Group-III, IV and V were received the plant extract 1, 2 and 3 at the dose of
200 mg/kg/p.o.). All the treatments were given 60
minutes prior to acetic acid injection. The number of abdominal writhes (full
extension of both hind paws) was cumulatively counted every 5 minutes over a
period of 20 minutes. The analgesic activity was expressed as percentage of
inhibition of abdominal writhes.% inhibition = (C-T/C)
X 100 where, C = number of writhes
produced by the control group and T = number
of writhes produced by the test group.12,13.
The experimental results
were expressed as the mean ± S.E.M. Data were assessed by the method of
analysis of ANOVA followed by student's t-test. P value of < 0.01 was
considered as statistically significant.
The
anti-inflammatory potential of MECI (200 mg/kg) against various experimental
animal models exhibited significant (P < 0.01) anti-inflammatory activity.
The effects of MECI and indomethacin on the
inflammation induced by carrageenan, are summarized in Fig 1, table-1.
As shown in Fig 1, MECI
showed maximum inhibition of 60.17% at the dose of 200 mg/kg after 3 h of
treatment in carrageenan induced paw oedema, whereas
the standard drug (Indomethacin 10 mg/kg) showed
62.34% of inhibition (P < 0.01). The effect of MECI on acute (carrageenan)
and chronic phase of inflammation, a maximum inhibition 60.17% was noted at the
dose of 200 mg/kg when compared with standard drug.
Table.1: Effects of MECI and indomethacin on carrageenan induced rat paw oedema
|
Design of treatment |
Drug |
Paw volume (in ml) |
||||
|
0 h |
1 h |
2 h |
3h |
Percentage inhibition |
||
|
Control |
5% gum acacia (5ml/kg/p.o.) |
0.54 ± 0.01 |
1.47±0.07 |
1.99±0.05** |
2.31±0.06** |
- |
|
Standard |
Indomethacin (10mg/kg/p.o.) |
0.56±0.011 |
0.63±0.01** |
0.85±0.01** |
0.87±0.01** |
62.34 |
|
Methanol Extract |
200mg/kg/p.o. |
0.54±0.005 |
0.75±0.15** |
0.86±0.006** |
0.92±0.1** |
60.17 |
aThe results given are mean±S.E.M.;
number of animal used (n=6), bP < 0.01
Experimental groups were compared with control
Table.2. Effects of MECI and indomethacin on writhing induced by acetic acid in mice
|
Design of treatment |
Drug |
Number of writhes in 20 minutes |
% inhibition |
|
Control |
5% gum acacia (5ml/kg/p.o.) |
34.3 ± 1.05** |
- |
|
Standard |
Indomethacin (10mg/kg/p.o.) |
13.83 ± 0.60** |
59.68 |
|
Methanol
Extract |
200mg/kg/p.o. |
14.5
± 0.43** |
57.73 |
aThe results given are mean±S.E.M.;
number of animal used (n=6), bP < 0.01
Experimental groups were compared with control
Acetic acid-induced writhing in mice:
The results presented
in Table 2, shows that MECI the doses of 200 mg/kg and indomethacin
(10mg/kg) exhibited significant (P < 0.01) inhibition of the control writhes
at the rate of 57.73%, when compared to that of control. In addition, MECI at
the above mentioned dose potentiated the analgesic activity of indomethacin shown by further decreasing the writhing
response when given in combination. Fig-2.
Test for acute toxicity: The aerial part
extract was found to be non toxic upto doses of 1.6
g/kg and did not cause any death of the animals tested.
The
MECI was evaluated for its anti-inflammatory activity in acute and chronic
models. A significant (P < 0.01) anti-inflammatory activity was observed for
MECI in carrageenan, induced oedema.
Carrageenan
induced rat paw oedema has been used as an
inflammation model in order to investigate the anti-inflammatory effect of drug14.
There are two phases of carrageenan induced inflammatory reaction: early or
first phase and later or second phase. It has been proposed that early phase
results from histamine, serotonin and bradykinin
liberation while late phase is associated with the release of prostaglandin15.
In carrageenan induced paw oedema the MECI showed
maxi-mum inhibition of 60.17% at the dose of 200 mg/kg after 3 h of drug
treatment.
From
these results, it is suggested that antioedematogenic
effects of the MECI on carrageenan induced oedema may
be related to inhibition of inflammation mediator formation.
In
order to distinguish between the central and peripheral analgesic action of
MECI, acetic acid induced writhing responses in mice were used to examine the
effect. This method is not only simple and reliable but also affords rapid
evaluation of peripheral type of analgesic action. In this test, the animals
react with characteristic stretching behavior, which is called writhing. It was
found that MECI significantly (P < 0.01) inhibited the acetic acid induced
writhing response and potentiated the analgesic activity of indomethacin
as well. The abdominal constriction is related to the sensitization of nociceptive receptors to prostaglandins. It is therefore
possible that MEC produced analgesic effect may be probably due to the
inhibition of synthesis or action of prostaglandins.
REFERENCES:
1.
Sosa S, Balick, MJ, Arvigo R,
Esposito RG, Pizza C, Altinier G,et al. A Screening of the topical Anti-inflammatory activity
of some Central American plants. J Ethanopharmacol
2002;8:211-215
2.
Kayaalp SO. Medical pharmacology, in terms of rational
treatment (Rasyonel tedavi yonunden tibbi farmakoloji), Ankara: Hacettepe Tas Ltd.Sti;
1998.
3.
Kirtikar KR, Basu BD. Indian medicinal plants. Vol II, 2 ed. Dehradum,
India Bishen Mahendra Pal
Singh. 1975. p. 1945-1946.
4.
The wealth of India, A Dictionary of Indian Raw materials and industrial
products. , 2004: P; 66-70, 2006: P; 205-207.
5. Gopal N
, Sengottuvelu S. Hepatoprotective activity of Clerodendrum inerme against CCl4 induced hepatic injury in rats. Fitoterapia. 2008 Jan;79(1):24-6.6.
Manoharan S, Kavitha K, Senthil N, Renju GL, Evaluation of anticarcinogenic
effects of Clerodendron inerme on
7,12-dimethylbenz(a) anthracene-induced hamster buccal pouch carcinogenesis. Singapore Med J. 2006 Dec;47(12):1038-43.
7.
Renju GL, Manoharan S, Balakrishnan S, Senthil N, Chemopreventive
and antilipidperoxidative potential of Clerodendron inerme (L) Gaertn in 7,12-dimethylbenz(a)anthracene
induced skin carcinogenesis in Swiss albino mice. Pak J Biol
Sci. 2007 May 1;10(9):1465-70.
8.
Olivieri F, Prasad V,
Valbonesi P, Srivastava S, Ghosal-Chowdhury P, Barbieri L, Bolognesi A, Stirpe F A
systemic antiviral resistance-inducing protein isolated from Clerodendrum inerme Gaertn. is a polynucleotide:adenosine
glycosidase (ribosome-inactivating protein), FEBS Lett.
1996 Nov 4;396(2-3):132-4.
9.
Winter, C.A., Risley, E.A., Nuss, G.W. Carrageenan induced
edema in hind paw of the rat as an assay for anti-inflammatory drugs. Proc.
Soc. Exp. Biol. Med., 1962, 111; 544-52
10.
Winter CA, Poster CC. Effect of alteration in side chain up on
anti-inflammatory and liver glycogen activities in hydrocortisone ester. J Amer Pharmacol Soc 1957;46:515-519.
11.
Suleyman H, Demirezer LO, Kuruuzum A, Banoglu ZN, Gocer F, Ozbakir G, et al.
Anti-inflammatory effect of the aqueous extract from Rumex
patientia L, roots. J Ethanopharmacol
1999;65:141-148.
12.
Nakamura, H, Shimoda, A, Ishii, K, Kadokawa, T. Central and peripheral analgesic action of
non-acidic non-steroidal anti-inflammatory drugs in mice and rats. Achieves
International Pharmacodynamic Therapy, 1986, 282;
16-25.
13.
Turner RA. In analgesics: Screening Methods in Pharmacology Turner R, Ebborn P, eds. Academic Press, New York: 1965.p.100
14.
El-Shenawy SM, Abdel-Salam OM, Baiuomy AR, El-Batran S, Arbid MS. Studies on the Anti-inflammatory and anti-nociceptive effects of melatonin in the rat. Pharmacol Res 2002;46:235-243.
15.
Ogonowski AA, May WS, Moor AB, Barret
LT, Bryant CL, Pollock SH. Antiinflammatory and
analgesic activity of an inhibitor of neuropeptide amidation. J Pharm Exp Ther 1997;280:846-853.
Received
on 05.12.2009
Accepted on 17.03.2010
© A&V Publication all right reserved
Research Journal of Pharmacognosy and Phytochemistry.
2(5): Sept.-Oct. 2010, 421-424