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In vitro and In vivo anti-inflamma-
tory activity of leaves of Symplocos 
cochinchinensis (Lour) Moore ssp 
laurina  

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Introduction  

Symplocos cochinchinensis Lour ssp laurina 
(Symplococaceae) otherwise known as  kabli-
vetti or Lodh tree is widely distributed in 
tropical, subtropical areas in Asia, Oceania and 
America. It is widely used in the treatment of 
various disorders like leprosy, tumors, diarrhea, 
dysentery, menorrhagia, inflammation and 
uterine disorders (Ali et al., 1990). Recently 
much attention have been paid for Symplocos 
species due to their diverse biological activity in 
treating various disorders like anti-HIV (Ishida 
et al., 2001), inhibitory activities against 
phosphodiesterase (Ahmad et al., 2003) and 
antitumor (Li et al., 2003) applications. 

The inflammatory response involves a complex 
array of enzyme activation, mediator release, 
fluid extravasations, cell migration, tissue 
breakdown and repair (Vane et al., 1995) which 

are aimed at host defense and usually activated 
in most disease condition. Currently much 
interest have been paid in the searching of 
medicinal plants with anti-inflammatory activity 
which may lead to the discovery of new 
therapeutic agent  that is not only used to 
suppress the inflammation but also used in 
diverse disease conditions where the 
inflammation response in amplifying the disease 
process. In this work the various extracts of S. 
cochinchnensis were studied for its in vitro and in 
vivo anti-inflammatory activities. 
 

Materials and Methods 

The fresh leaves of S. cochinchinensis were 
collected from Ooty in the month of August and 
it was authenticated by Prof. J. Jayaraman, 
Taxonomist, PARC, Chennai. A voucher speci-
men is deposited in the department for the 

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In vitro and In vivo anti-inflammatory activity of leaves of 
Symplocos cochinchinensis (Lour) Moore ssp laurina  
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The anti-inflammatory activities of different extracts of leaves 
Symplocos cochinchinensis Lour ssp laurina (Symplococaceae) were 
investigated for in vitro anti-inflammatory activity by human red 
blood cell membrane stabilization method. The methanol extract 
showed effective in vitro anti-inflammatory activity was screened for 
in vivo anti-inflammatory activity by carrageenan-induced paw edema 
in rat model. The potency of the extracts was compared with standard 
diclofenac (50 mg/kg). The methanol extract showed significant 
membrane stabilizing action on human red blood cell membrane and 
reduction of edema in carrageenan induced rat paw edema model. 

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Received:                               8 July 2008 
Accepted:                            28 July 2008 
Available Online:           1 August 2008  
 
DOI: 10.3329/bjp.v3i2.956 
 
 
Cite this article: 
Vadivu R, Lakshmi KS. In vitro and In 
vivo anti-inflammatory activity of 
leaves of Symplocos cochinchinensis 
(Lour) Moore ssp laurina. Bangladesh 
J Pharmacol. 2008; 3: 121-24. 

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future reference. The collected leaves were shade 
dried and coarsely powdered. The coarse 
powder was subjected to continuous extraction 
in a soxhlet apparatus using n-hexane, 
chloroform, ethyl acetate and methanol as 
solvents. The solvents were distilled under 
reduced pressure using rotary vacuum 
evaporator. The extracts were dissolved in 
distilled water using 1% CMC as suspending 
agent. 

Animals 

Wister male albino rats weighing 150-200 g were 
used for the in vivo anti-inflammatory studies. 
The animals were housed under standard 
conditions of temperature (23 ± 1°C), 12 hours 
light/dark cycle and fed with water ad libitium. 
Before performing the experiment the ethical 
clearance was obtained from institutional animal 
ethics committee. 

In vitro anti-inflammatory activity (Gandhisan et 
al., 1991): The human red blood cell membrane  
stabilization method was used for this study. 
The blood was collected from healthy human 
volunteer who was not taken any NSAIDS for 2 
weeks prior to the experiment and mixed with 
equal volume of Alsever solution (2% dextrose, 
0.8% sodium citrate, 0.5% citric acid and 0.4% 
NaCl) and centrifuged at 3,000 rpm. The packed 
cells were washed with isosaline and a 10% 
suspension was made. Various concentrations of 
extracts were prepared (250, 500 and 1,000 mcg/
mL) using distilled water and to each 
concentration 1 mL of phosphate buffer, 2 mL 
hyposaline and 0.5 mL of HRBC suspension 
were added. It is incubated at 37°C for 30 min 
and centrifuged at 3,000 rpm for 20 min. the 
hemoglobin content of the supernatant solution 
was estimated spectrophotometrically at 560 nm. 
Diclofenac (50 mcg/mL) was used as reference 
standard and a control was prepared omitting 
the extracts. 

Acute toxicity studies (Ecobichon, 1987) 

Acute toxicity study was carried out for 
methanol extract using Acute Toxic Class 
Method as described in OECD [Organization of 
Economic Co-operation and Development] 
Guidelines No. 423. The methanol extract was 
safe up to a dose of 2,000 mg/kg body weight so 
200 mg/kg and 400 mg/kg were used as 

moderate dose for the evaluation. 

In vivo anti-inflammatory activity (Winter et 
al., 1962) 

In vivo anti-inflammatory activity was studied 
for the methanolic extract by carrageenan 
induced paw edema model in rats. The Wister 
albino rats were divided in to 4 groups of 6 
animals each. Group I: served as control, Group 
II, III: received methanol extract at the dose of 
200 and 400 mg/kg and Group IV was served as 
standard (diclofenac 50 mg/kg). Carrageenan 
was injected in to the sub planter aponeurosis of 
the right hind paw of rats. An hour before 
carrageenan injection the animals were given 
with the extract, standard orally. The paw 
volumes were measured before and three hours 
after carrageenan administration by volume 
displacement method. 

Statistical analysis 

The data’s were expressed as mean ± SEM,  
statistical analysis was performed by one-way 
ANOVA followed by Tukey-Kramer multiple 
comparison test, p values <0.05 were considered 
as significant. Highest significant difference test 
performed with GraphPad instat software.  

 

Results and Discussion 

The n-hexane, chloroform, ethyl acetate and 
methanolic extracts of the leaves of S. 
cochinchinensis were studied for in vitro anti-
inflammatory activity by HRBC membrane 
stabilization method. Among all the extracts 
methanolic extract showed significant anti-
inflammatory activity in a concentration 
dependent manner. Methanol extract at a 
concentration of 1,000 mcg/ml showed 67% 
protection of HRBC in hypotonic solution. All 
the results were compared with standard 
diclofenac which showed 74% protection. 

The extracts exhibited membrane stabilization 
effect by inhibiting hypotonicity induced lyses of 
erythrocyte membrane. The erythrocyte 
membrane is analogous to the lysosomal 
membrane (Chou, 1997) and its stabilization 
implies that the extract may as well stabilize 
lysosomal membranes. Stabilization of 
lysosomal membrane is important in limiting the 

122 Bangladesh J Pharmacol 2008; 3: 121-124 



 

 

inflammatory response by preventing the release 
of lysosomal constituents of activated neutrophil 
such as bactericidal enzymes and proteases, 
which cause further tissue inflammation and 
damage upon extra cellular release (Murugasan, 
1981). Some of the NSAIDs are known to posses 
membrane stabilization properties which may 
contribute to the potency of their anti-
inflammatory effect. Though the exact 
mechanism of the membrane stabilization by the 
extract is not known yet, hypotonicity-induced 

hemolysis may arise from shrinkage of the cells 
due to osmotic loss of intracellular electrolyte 
and fluid components. The extract may inhibit 
the processes, which may stimulate or enhance 
the efflux of these intracellular components 
(Iwueke, 2006). 

Since methanol extract showed significant in-
vitro anti-inflammatory activity it was selected 
for the evaluation of in vivo anti-inflammatory 
activity by carrageenan induced paw edema 
model in rats. The extract showed significant 
anti-inflammatory activity (53%) at the dose of 
400 mg/ml while the standard diclofenac 
showed 75% inhibition of edema. On the basis of 
the above results it can be concluded that the 
methanol extract posses significant anti-
inflammatory activity studied by in vitro and in 
vivo models. The study also provides a strong 
evidence for the use of the leaves S. 
cochinchinensis in folkloric treatment as anti-
inflammatory agent. The activity may be due to 
the presence of one or more phytochemical 
constituents present in the extract further study 
is warranted, for isolation of the constituents 
responsible for the activity and also to explore 
the exact mechanism of action of the activity. 

 

References 
Ali M, Bhutani KK, Srivastava TN. Triterpenoids from 

Symplocos racemosa bark. Phytochemistry 1990; 29: 3601-04. 
http://dx.doi.org/10.1016/0031- 9422(90)85284-M 

Ahmad VU, Abbasi MA, Hussain H, Akhtar MN, Farooq U, 
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00075-X (PMid:12711144) 

Chou CT. The anti-inflammatory effect of Tripterygium wilfordii 
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11:2%3c152::AID-PTR45%3e3.0.CO;2-L 

Ecobichon DJ. The basis of toxicology testing. 2nd ed. New 
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Table I 

In vitro anti inflammatory activity of methanolic 
extract of symplocos cochinchinensis by HRBC 

membrane stabilization method 
Treatment Conc 

(mcg/mL) 
Absorbance 

(540 nm) 
%

Inhibition 
  

Control   -- 0.48 ± 0.012   -- 

n-Hexane  1,000 0.35 ± 0.03b 28.6 

  500 0.36 ± 0.0b 25.8 

  250 0.38 ± 0.005 23.4 

Chloroform  1,000 0.34 ± 0.003a 31.0 

  500 0.33 ± 0.04a 30.0 

  250 0.35 ± 0.005a 26.5 

Ethyl acetate  1,000 0.25 ± 0.001a 47.3 

  500 0.28 ± 0.007a 43.1 

  25 0.29 ± 0.005a 40.2 

Methanol  1,000 0.15 ± 0.001a 69.9 

  500 0.19 ± 0.002a 60.5 

  250 0.20 ± 0.001 58.1 

Diclofenac      50 0.13 ± 0.002a 73.9 

Values are expressed as mean ± SEM.  n = 6 animals in 
each group. ap<0.001; bp<0.01 compared to control group 

Table II 

In vivo anti inflammatory activity of methanolic 
extract of Symplocos cochinchinensis on caar-

ageenin- induced paw edema in rats 
Treatment Dose 

(mg/kg) 
Edema volume 

(mL) 
% Inhi-
bition 

Control   -- 0.72 ± 0.020   -- 

Methanolic 
extract 

200 0.48 ±  0.029a 33.3 

Methanolic 
extract 

400 0.315 ± 0.012a  53.4 

Diclofenac 250 0.180 ± 0.001a 75.0 

Values are expressed as mean ± SEM. n = 6 animals in 
each group. ap<0.001 compared to control group 

                                                                         Bangladesh J Pharmacol 2008; 3: 121-124                                                                                123 



 

 
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$XWKRU�,QIR 
Rajendran Vadivu (Principal contact) 
e-mail: r.vadivu@rediffmail.com 

 

124 Bangladesh J Pharmacol 2008; 3: 121-124 


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