Amrit Pal Singh

Anil K Gangwar

N Venkat Rao

Nishant S Katiyar

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IJRPC 2015, 5(2), 322-326                  Nishant S Katiyar et al.                  ISSN: 22312781 
 
322 
INTERNATIONAL JOURNAL OF RESEARCH IN PHARMACY AND CHEMISTRY 
 
Available online at www.ijrpc.com 
 
EVALUATION OF CARRAGEENAN INDUCED ANTI-
INFLAMMATORY ACTIVITY OF STEM EXTRACTS OF 
CUSCUTA REFLEXA (ROXB) IN RATS 
Nishant S Katiyar1*, Amrit Pal Singh1, Anil K Gangwar2 and N. Venkat Rao3 
1 Sanjay College of Pharmacy, Mathura, Uttar Pradesh- 281 406, India. 
2Department of Pharmacy, Kanpur Institute of Technology,  
Kanpur- 208 001, Uttar Pradesh, India. 
3Department of Pharmacology, V.L.College of Pharmacy, Raichur,  
Karnataka-584 103, Bangalore, India. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
INTRODUCTION 
Inflammation, a local response of living 
mammalian tissues to injury, is a body defense 
reaction to eliminate or limit the spread of 
injurious agent. There are various components 
to an inflammatory reaction that can contribute 
to the associated symptoms and tissue injury. 
Edema formation, leukocyte infiltration and 
granuloma formation represent such 
components of inflammation1. 
Herbal medicines derived from plant extracts 
are being increasingly utilized to treat a wide 
variety of clinical diseases, though relatively 
little knowledge about their mode of action is 
available. There is a growing interest in the 
pharmacological evaluation of various plants 
used in Indian traditional systems of medicine. 
Thus, the present investigation was carried out 
to evaluate the anti-inflammatory potential of 
C. reflexa in experimental animal models. 
 C. reflexa, commonly known as Amarbel is a 
leafless, delicate yellow coloured total stem 
parasite, belonging to the plant family 
Convolvulaceae. The tiny white flowers appear 
in bunches and the fruit are pea shaped and 
seeds are black in colour2. It is found 
throughout India. The plant is acrid; bitter; 
astringent , aphrodisiac, alternative, tonic and 
useful in diseases of the eye and of the heart, 
also used in biliousness and in “kapha;3  
The herb has a bitter sharp taste; used as 
expectorant, carminative, tonic, anthelmintic, 
diuretic, blood purifier and lessens 
inflammation. It is also useful in jaundice, pain 
in the muscles and joints, headache, paralysis 
and in lumbago. 
It was reported that decoction prepared with 
stem is useful in constipation, flatulence, liver 
complaints and bilious affections. The seeds 
have a bitter bad taste; sedative, 
emmenagogue, diuretic; useful in diseases of 
the liver and the spleen, chronic fevers, 
griping. hiccough; used to purify the blood and 
cleanse the bowels; the infusion is given in 
opthalmia, the decoction in biliousness as a 
purgative (Unani). The plant is purgative, it is 
Research Article 
ABSTRACT 
Alcoholic (AESCR) and aqueous (AQESCR) extracts of stem of Cuscuta reflexa were evaluated for their 
anti-inflammatory activity in carrageenan induced paw edema model in rats, and compared to the 
activity of standard drug, Ibuprofen. These extracts were given orally in a concentration of 100, 200 
and 400 mg/kg bd.wt. before carrageenan injection. Both the extracts with medium and higher doses 
i.e. 200 mg/kg and 400 mg/kg have reduced oedema volume by 47.27%, 72.72% and 57.72%, 80.00% 
respectively at 5th h as compared to standard drug Ibuprofen 96.36%. Thus the present study revealed 
that the selected extracts of C. reflexa exhibited a significant anti-inflammatory activity in carrageenan 
induced paw oedema model in rats. 
 
Keywords: C reflexa, stem extracts, carrageenan, ibuprofen, anti-inflammatory activity. 
 
IJRPC 2015, 5(2), 322-326                  Nishant S Katiyar et al.                  ISSN: 22312781 
 
323 
used externally against itch and internally to 
protract fevers. The stems are specially useful 
in bilious disorders 2,3,4. 
On the basis of these common uses of this 
plant in traditional folk medicine, it is planned 
to evaluate the anti-inflammatory effect of 
AESCR and AQESCR in carrageenan induced 
inflammatory model in rats. 
 
MATERIALS AND METHODS 
Plant material 
Stems of C. reflexa collected in the month of 
May were identified by a botanist Prof. V. 
Hemanth Kumar, V.L. College of Pharmacy, 
Raichur and dried in shade at room 
temperature then subjected to size reduction 
to a fine powder with the help of mixer grinder. 
 
Chemicals 
Ibuprofen was obtained as a gift sample from 
Cipla, Mumbai, India. Carrageenan was 
purchased from HiMedia Laboretories, 
Mumbai, India.   
Animals:  
Albino rats (Wistar strain) of either sex 
weighing between 150-200 g and Albino mice 
16-25g were procured from National Centre for 
Laboratory Animal sciences, C/0 
Sri.Venkateswara Enterprises, Bengaluru for 
experimental purpose. Then the animals were 
acclimatized for 7 days under standard 
husbandry condition. i.e. 
Room temperature                -          26 ± 20 C   
Relative humidity                  -           45-55% 
Light/ dark cycle                   -           12:12 h  
The animals were fed with a synthetic 
standard diet from Amrut Laboratories & 
Pranav Agro Industries Ltd. Sangli. Water was 
allowed ad libitum under strict hygienic 
conditions. All animal studies were performed 
in accordance to the guidelines of CPCSEA, 
No. 425 with registration number 
557/02/c/CPCSEA and Institutional Animal 
Ethical Committee (IAEC) of V.L. College of 
Pharmacy, Raichur (Karnataka) and all the 
procedures were followed as per rules and 
regulations. 
 
Preparation of extracts5 
Preparation of alcoholic extract 
The stem powder was packed in a soxhlet 
apparatus and extracted with 95% alcohol for 
18 h. Appearance of colourless solvent in the 
siphon tube was taken as the termination of 
extraction. The extract was then transferred 
into the previously weighed empty beaker and 
evaporated to a thick paste on the water bath, 
maintained at 50oC. The extract was finally air 
dried thoroughly to remove all traces of the 
solvent. 
 
Preparation of aqueous extract 
About 100 g of powder was taken in a round 
bottom flask (2000 ml) and macerated with 
500 ml of distilled water with 10 ml of 
chloroform used as a preservative for 7 days 
with occasional shaking for every hour in a 
closed vessel. Then the marc was removed by 
filtering the extract and the extract was 
concentrated on a water bath maintained at 
50oC .  
These two extracts were stored in airtight 
containers in a refrigerator below 40C and 
were examined for their colour and 
consistency. Their percentage yield was 
calculated with reference to air-dried powder 
sample used for the extraction. 
 
Toxicity studies 
The acute toxicity of AESCR and AQESCR 
was determined by using albino mice of either 
sex (16-20 g), maintained under standard 
husbandry conditions. The animals were 
fasted for 3 h prior to the experiment and were 
administered with single dose of individual 
extracts and observed for the mortality upto 48 
h study period (Short term toxicity). Based on 
the short-term toxicity profile, the next dose of 
the individual extracts was determined as per 
OECD guidelines No. 425. A maximum dose 
of 2000 mg/kg bd. wt was tested for its 
mortality.  From the LD50 doses of each 
extracts 1/20,1/10 and 1/5 doses were 
selected and considered as low, medium and 
high dose respectively6. 
  
Carrageenan induced paw oedema model7,8 
Albino rats of either sex weighing 125 – 150 g 
were selected and were maintained on 
standard pellet diet and free access to water. 
The animals were divided into 8 groups each 
containing 6 animals. The groups were treated 
as follows:  
Group A   - Toxicant control 
(Carrageenan 1% w/v,0.1 ml s.c) 
Group B - Standard (Ibuprofen 
40 mg/kg, p.o) 
Group C - AESCR (low dose 100 
mg/kg, p.o) 
Group D -          AESCR (medium dose 
200 mg/kg, p.o) 
Group E -           AESCR (high dose 
400 mg/kg, p.o) 
Group F - AQESCR (low dose 
100 mg/kg, p.o) 
Group G - AQESCR (medium 
dose 200 mg/kg, p.o) 
Group H - AQESCR (high dose 
400 mg/kg, p.o) 
Initial paw volume of individual rats (right paw) 
was noted at ‘0’ h. One hour after the 
administration of vehicle, extracts, standard 
IJRPC 2015, 5(2), 322-326                  Nishant S Katiyar et al.                  ISSN: 22312781 
 
324 
drug, all the rats were injected with 0.1 ml of 
histamine (0.1%) in normal saline in the sub-
plantar region of the right hind paw and the left 
hind paw was served as reference. 
Immediately thereafter the oedema volume of 
the injected paws were measured 
plethysmographically at fixed time intervals 
i.e., 3rd h and 5th h. 
 The difference between paw volumes of the 
treated animals was measured and the mean 
oedema volume was calculated. Percentage 
reduction in oedema volume was calculated by 
using the formula,    
 
 
 Where    
 Vo = Paw volume of the control at time‘t’.    
 Vt = Paw Volume of the drug treated at time‘t’. 
 
Stastical analysis 
All the recorded results are expressed as 
mean ± SEM from 6 animals. Statistical 
difference in mean was analyzed by using 
one-way ANOVA (analysis of variance) 
followed by Post hoc test (Dunnett’s ‘t’ test). 
P< 0.05*, 0.01** and 0.001*** were considered 
statistically significant. 
 
RESULT 
The AESCR and AQESCR with three dose 
levels tested i.e. 100, 200 and 400 mg/kg had 
exhibited a significant reduction in paw 
oedema volume in carrageenan induced paw 
oedema (acute) model in rats. Results are 
tabulated in (Table 1). Ibuprofen (40 mg/kg) is 
used as standard reference and it has reduced 
paw oedema volume by 93.61% at 5th h. 
AESCR and AQESCR with medium and 
higher doses i.e. 200 mg/kg and 400 mg/kg 
have reduced oedema volume by 38.29%, 
53.19% and 48.93%, 61.70% respectively at 
5th hr. (Figure 1). 
 
DISCUSSION  
Inflammation has different phases; the first 
phase is caused by an increase in vascular 
permeability, the second one by infiltration of 
leukocytes and the third one by granuloma 
formation9. Histamine is one of the important 
inflammatory mediators and it is potent 
vasodilators substance and increases the 
vascular permeability10,. Inflammatory 
diseases affecting majority of the peoples, are 
very common and are known to be as oldest 
diseases as that of mankind. No substantial 
progress has been achieved till today for their 
permanent cure. Anti-inflammatory activity was 
investigated in acute models of inflammation in 
rats as the inflammation was induced by sub-
plantar injection of carrageenan. It was 
reported that carrageenan administration 
causes release of various mediators like 
histamine, serotonin (initial phase), kinins 
(middle phase) and PG (final phase) that play 
an important role in the development of 
inflammation11. AESCR and AQESCR have 
inhibited the initial, middle and final phases 
suggesting that the extracts can block the 
mediators like histamine, kinins and PGs. In 
histamine induced paw edema model AESCR 
and AQESCR exhibited significant inhibitory 
action against histamine induced paw edema 
and this indicates that these extracts exhibited 
their anti-inflammatory action by means of 
inhibiting the synthesis, release or action of 
various inflammatory mediators like kinins and 
histamine.12 The study showed that the 
AESCR and AQESCR effectively suppressed 
the oedema produced by the carrageenan, 
which is indicate that the extracts exhibit its 
anti-inflammatory action by means of either 
inhibiting the synthesis, release or action of 
inflammatory mediators viz. histamine, 
serotonin and prostaglandin’s might be 
involved in the inflammation. 
Phytoconstituents reported for the anti-
inflammatory activity are glycosides, 
flavonoids, sterols and saponins present in 
both the extracts hence these can be 
accounted for its anti-inflammatory activity13. 
The AQESCR was found to possess relatively 
better anti-inflammatory activity than AESCR. 
 
CONCLUSION  
The pharmacological studies on different parts 
of the plant C. reflexa revealed a significant 
antibacterial5, anti-steroidogenic, 
psychopharmacological activity.  
The preliminary phytochemical analysis of the 
AESCR and AQESCR  revealed the presence 
of carbohydrates, sterols, flavonoids, 
glycosides, fixed oils fats, saponins and 
alkaloids.   
From the studies it can be concluded that 
AESCR and AQESCR (200, 400 mg/kg) have 
shown significant anti-inflammatory activity 
against carrageenan induced paw edema 
model in rats (acute model). The AQESCR 
was found to be more potent than AESCR 
which is confirmed by its higher percentage 
reduction in paw oedema volume than the 
other in carrageenan induced paw oedema 
(acute) model in rats. 
 
 
 
ACKNOWLEDGEMENT 
The Authors are thankful to the principal and 
management of V L College of Pharmacy, 
IJRPC 2015, 5(2), 322-326                  Nishant S Katiyar et al.                  ISSN: 22312781 
 
325 
Raichur, Karnataka for providing the facilities to carry out this research work. 
 
 
 
Table 1: Anti-inflammatory effect of stem extracts of C. reflexa  
carrageenan induced   (acute) paw oedema model in rats 
Group %ROV Treatment Dose mg/kg at 3rd h Paw oedema (ml) %ROV at 5th h Paw oedema (ml) 
1 
- Control 10 mL 0.49±0.009 - 0.47±0.004 
2 
93.61 
Standard 
(Ibuprofen) 40 0.08±0.005** 83.67 0.03±0.003** 
3 
25.33 AESCR 100 0.04±0.006** 18.36 0.35±0.0011** 
4 
38.29 AESCR 200 0.34±0.006** 30.61 0.29±0.004** 
5 
53.19 AESCR 400 0.28±0.008** 42.85 0.22±0.007** 
6 
34.04 AQESCR 100 0.36±0.006** 26.53 0.31±0.003** 
7 
48.93 AQESCR 200 0.29±0.006** 40.81 0.24±0.006** 
8 
61.70 AQESCR 400 0.23±0.006** 53.06 0.17±0.006** 
   n = 6,  Significant at P < 0.05*,  0.01**, and 0.001***,   ns = not significant 
    AESCR – Alcoholic extract of stem of C. reflexa,   AQESCR – Aqueous extract of stem of C. reflexa 
    % ROV - % reduction of oedema volume 
 
 
 
 
Fig. 1: 
 
           
 
 
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EVALUATION OF CARRAGEENAN INDUCED ANTI- INFLAMMATORY ACTIVITY OF STEM EXTRACTS OF CUSCUTA REFLEXA (ROXB) IN RATS

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EVALUATION OF CARRAGEENAN INDUCED ANTI- INFLAMMATORY ACTIVITY OF STEM EXTRACTS OF CUSCUTA REFLEXA (ROXB) IN RATS

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