INTRODUCTION:
Inflammatory abnormalities underlie a vast variety of
human diseases. The immune system is often involved in inflammatory disorders
and is demonstrated in both allergic reactions and myopathies. Non-immune
diseases with inflammation include atherosclerosis and ischemic conditions
(Cortan et al., 1998). Due to the serious side effects produced by the
synthetic drugs a trend from the usage of synthetic drugs to herbal medicine is
observed among mankind which can be called ‘Return to Nature’. Antioxidant and
anti-inflammatory principles present in the natural resources are providing
enormous scope in herbal medicine. Therefore, in the recent years, research
interest is focused on phytochemicals that are derived from herbal sources in
view of their therapeutic benefits (Kamat et al., 2007). Terminalia catappa L.
is a Combretaceous plant whose
leaves are widely used as a folk medicine in Southeast Asia for the treatment
of dermatosis and hepatitis (Lin et al., 1997). This species is globally
distributed from Indo-Malaya to Australia. It is widely planted throughout the
tropics, especially along sandy seashores, for shade, ornamental purposes, and
edible nuts.
The phytochemicals of this plant include tannins
(puni-calagin, punicalin, terflavins A and B, tergallagin, ter-catain,
chebulagic acid, geranin, granatin B, corilagin) , flavanoids (isovitexin,
vitexin, isoorientin, rutin) and triterpinoids (ursolic acid, 2á, 3â,
23-trihydroxyurs-12-en-28 oic acid) (Ahmed et al.,2005). The Fruit of T. catappa contains corilagin,
brevifolin-carboxylic-acid, beta-carotene, cyanidin-3-glucoside, ellagic-acid,
gallic-acid, glucose, pentosans, tannin (Duke, 2008).In view of this, the
present study was designed to evaluate in vitro anti oxidant and anti
inflammatory potentials on the fruits of Terminalia
catappa L.
MATERIALS
AND METHODS:
Collection of plant material:
The fresh fruits of Terminalia catappa were collected from Mannargudi, Tamil Nadu.
Plant material was identified and authenticated in the department of CARISM,
SASTRA University, Thirumalaisamudhram, Tamil Nadu. The collected materials
were cleaned, pericarp shade dried and coarsely powdered.
Preparation of the extract:
50g of plant powder was dissolved in 250ml water and
incubated for 36 hrs. Then, it was filtered and the filtrate was boiled at 56◦C
up to semi solid consistency. Then the aqueous extract was used for the
evaluation of in vitro antioxidant & anti-inflammatory activities.
Assessment of in vitro anti-Inflammatory Activity
In vitro anti inflammatory activity was carried out
using various models such as HRBC Membrane Stabilization (Sakat et al., 2001),
Inhibition of protein Denaturation (Mizushima and Kobayashi, 1968) Proteinase
inhibitory activity (Oyedepo and Femurewas, 1995).
Assessment of in vitro Antioxidant
Activity:
In vitro anti oxidant activity was carried out using
DPPH radical scavenging activity (Brand et al., 1995) and inhibition of lipid
peroxidation (Yagi, 1978).
RESULTS
AND DISCUSSION:
Inflammation is a normal protective response to tissue
injury caused by physical trauma, noxious chemical or microbial agents. It is
the body response to inactivate or destroy the invading organisms, to remove
the irritants and set the stage for tissue repair. It is triggered by the
release of chemical mediators from injured tissue and migrating cells. The
commonly used drug for management of
inflammatory conditions are
non-steroidal anti- inflammatory drugs, which have several adverse effects
especially gastric irritation leading to formation of gastric ulcers (Tripathi,
2008). In the present study, initiatives
were taken to evaluate the anti inflammatory and antioxidative potentials of T. catappa fruits. Erythrocytes have been used
as a model by a number of scientists to investigate the interaction of drugs
with membranes (Sessa and Weisman, 1968; Lietman et al., 1976). The membrane stabilizing activity of red blood
cells (RBC) that are exhibited by some drugs is used for in vitro method for
assessing the anti-inflammatory activity of various herbal drugs. In the present study, 10 different
concentrations of the aqueous extract of T.
catappa fruits was evaluated for the
HRBC membrane stabilization activity. 50mg/ml of the extract was found to
stabilize the RBC membrane up to 87.32% (Table 1).
Table 1: HRBC Membrane
stabilization activity of aqueous extract of T. catappa fruits
|
S. No.
|
Concentration of T. catappa fruits (mg/ml)
|
% of
Stabilization
|
|
1
|
50.00
|
87.32±0.51
|
|
2
|
25.00
|
82.80±0.10
|
|
3
|
12.50
|
79.64±0.45
|
|
4
|
6.25
|
77.23±0.51
|
|
5
|
3.15
|
74.36±0.51
|
|
6
|
1.56
|
68.52±0.29
|
|
7
|
0.78
|
63.48±0.29
|
|
8
|
0.39
|
61.86±0.06
|
|
9
|
0.20
|
60.11±0.03
|
|
10
|
0.10
|
55.36±0.32
|
|
11
|
Aspirin(0.1mg/ml )
|
69.11±23.82
|
The extract exhibited membrane stabilization activity
in a dose dependent manner. Protein
denaturation is a process in which proteins lose their tertiary structure and
secondary structure as well as biological function. Denaturation of tissue
protein is one of the well-documented causes of inflammatory and arthritic
diseases. Production of auto antigens in certain arthritic diseases may be due
to denaturation of proteins. In vivo agents that can prevent protein
denaturation therefore would be worthwhile for anti-inflammatory drug
development (Umapathy et al., 2010).
Hence the ability of a plant extract to inhibit protein denaturation can be
studied to assess the anti inflammatory activity of the extract. In the present
study, the aqueous extract of T. catappa fruits
exhibited high degree of inhibition of protein denaturation (Table 2).
Table 2: Protein denaturation
inhibitory activity of aqueous extract of T.catappa
fruits
|
S.No.
|
Concentration of T.
catappa fruits (mg/ml)
|
%Inhibition
|
|
1
|
50.00
|
99.07±0.19
|
|
2
|
25.00
|
96.20±0.31
|
|
3
|
12.50
|
91.56±0.35
|
|
4
|
6.25
|
86.68±0.14
|
|
5
|
3.13
|
82.39±0.02
|
|
6
|
1.56
|
78.22±0.06
|
|
7
|
0.78
|
73.20±0.06
|
|
8
|
0.39
|
68.87±0.13
|
|
9
|
0.20
|
63.71±1.57
|
|
10
|
0.10
|
58.20±0.09
|
|
11
|
Aspirin(0.1mg/ml)
|
28.66±3.16
|
The result indicated that 50mg/ml of the extract
inhibited protein denaturation up to 99.07%. The activity was dose dependent.
Neutrophils are known to be a rich source of serine proteinase and are
localized at lysosomes. It was previously reported that leukocytes proteinase
play an important role in the development of tissue damage during inflammatory
reaction and significant level of protection was provided by proteinase
inhibition. The aqueous extract of T. catappa fruits exhibited significant
anti proteinase activity of different concentrations as shown in Table 3.
Table
3: Proteinase inhibitory activity of aqueous extract of T.
catappa fruits
|
S.No.
|
Concentration of T.catappa
fruits (mg/ml)
|
% Inhibition
|
|
1
|
50
|
99.14±0.12
|
|
2
|
25
|
97.73±0.18
|
|
3
|
12.5
|
96.81±0.28
|
|
4
|
6.25
|
95.26±0.11
|
|
5
|
3.13
|
93.35±0.60
|
|
6
|
1.56
|
91.38±0.07
|
|
7
|
0.78
|
85.29±0.23
|
|
8
|
0.39
|
73.82±1.08
|
|
9
|
0.2
|
65.61±0.23
|
|
10
|
0.1
|
60.18±1.31
|
|
11
|
Aspirin(0.1
mg/ml)
|
62.3±15.56
|
It showed maximum inhibition of 99% at 50mg/ml.
Aspirin showed the maximum inhibition 62.3% at 0.1mg/ml. The radical scavenging activity of different
extracts was tested using methanolic solution of the stable free radical DPPH.
A freshly prepared DPPH solution exhibits a deep purple color generally
fades/disappears when an antioxidant is present in the medium. Thus,
antioxidant molecule can quench DPPH free radicals (by providing hydrogen atom
or by electron transfer, conceivably via a free radical attack on the DPPH
molecule) and convert them to a colorless product (2,
2-diphenyl-1-picrylhydrazyl, or a substituted analogous hydrazine) resulting in
a decrease in absorbance at 518 nm (Yamaguchi et al., 2002). Table 4
reveals the free radical scavenging activity of the plant extract.
Table 4: DPPH Radical Scavenging activity of aqueous extract of T. catappa fruits
|
S.No.
|
Concentration of T.catappa
fruits (mg/ml)
|
% Inhibition
|
|
1
|
50
|
97.34±0.11
|
|
2
|
25
|
95.61±0.06
|
|
3
|
12.5
|
86.95±1.14
|
|
4
|
6.25
|
70.75±1.03
|
|
5
|
3.13
|
53.67±0.46
|
|
6
|
1.56
|
40.93±0.11
|
|
7
|
0.78
|
30.54±1.14
|
|
8
|
0.39
|
14.34±1.03
|
|
9
|
0.2
|
8.74±0.40
|
|
10
|
0.1
|
5.56±1.03
|
|
11
|
BHT(0.1mg/ml)
|
95.30±8.25
|
Table
5: Lipid peroxide Scavenging activity of
the aqueous extract of T. catappa fruits
|
S.No.
|
Concentration of T.catappa fruits (mg/ml)
|
%Inhibition
|
|
1
|
50
|
85.32±1.43
|
|
2
|
25
|
71.52±0.76
|
|
3
|
12.5
|
62.96±1.09
|
|
4
|
6.25
|
45.72±1.26
|
|
5
|
3.13
|
38.59±0.76
|
|
6
|
1.56
|
25.09±2.02
|
|
7
|
0.78
|
11.24±0.92
|
|
8
|
0.39
|
4.16±0.17
|
|
9
|
0.2
|
2.73±0.34
|
|
10
|
0.1
|
1.55±0.50
|
The extract exhibited radical scavenging activity of
97.34% at 50 mg/ml. Radical Scavengers may protect tissues from free radicals,
thereby from diseases such as cancer (Nakayama et al., 1998). Terminila catappa fruits demonstrated
DPPH scavenging activity in a concentration dependent manner. Free radical induced lipid peroxidation is
associated with a number of disease processes (Giugliano et al., 1996). The
aqueous extract of T.catappa fruits
was evaluated for lipid peroxide scavenging activity. The extract exhibited
peroxide scavenging activity of 85.3% at 50mg/ml and the potential of the
extract to scavenge peroxide radicals is found to be dose dependent. Present
paper deals with in vitro studies on anti-inflammatory and antioxidant
potentials of Terminalia catappa fruits.
Results obtained from the present study provide scientific evidences for the
use of this plant in folk medicine. Further, the present study suggests that
fruits of T. catappa could serve as a lead in the development of a novel herbal
anti-inflammatory and antioxidant agent.
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