Antihepatotoxic and antioxidant defense potential of Mimosapudica

Abstract

Reactive Oxygen species (ROS) are believed to be responsible for pathogenesis of variousdiseases affecting tissues and the main organ, the Liver. Hence, in the present study, the extent of LipidPeroxidation (LPO) and ROS elimination and its defense mechanisms by the enzymic & non enzymicantioxidants in liver & serum was investigated. Hepatoxicity was manifested by significantly decreased(p<0.05) levels in the activities of the enzymic antioxidants such as Superoxide dismutase (SOD) Catalase(CAT), Glutathione peroxidase and the non enzymic antioxidants such as glutathione & Vitamin C in ratsinduced hepatic damage by ethanol Simultaneous administration of the leaf extract Mimosa pudica alongwith the toxin ethanol in rats showed a considerable protection against the toxin induced oxidative stressand liver damage as evidence by a significant increase (p<0.05) in antioxidant activities. The study revealsthat the co administration of Mimosa pudica aqueous extract significantly lowered the level of lipidperoxidation in alcohol fed mice.

Full text

International Journal of Drug Disco very, ISSN: 0975–4423, Volume 1, Issue 2, 2009, pp-01-04
Copyright © 2009, Bioinfo Publications, International Journal of Drug Discovery, ISSN: 0975–4423, Volume 1, Issue 2, 2009
Antihepatotoxic and antioxidant defense potential of Mimosa pudica
Nazeema T.H.1 and Brindha V.2*
1Department of Biochemistry, RVS College of Arts and Science, Coimbatore, Tamil Nadu, India
*2Department of Biochemistry, PSG College of Arts and Science, Coimbatore, Tamil Nadu, India,
brindhavenkatesh@ymail.com
Abstract- Reactive Oxygen species (ROS) are believed to be responsible for pathogenesis of various
diseases affecting tissues and the main organ, the Liver. Hence, in the present study, the extent of Lipid
Peroxidation (LPO) and ROS elimination and its defense mechanisms by the enzymic and non enzymic
antioxidants in liver and serum was investigated. Hepatoxicity was manifested by significantly decreased
(p<0.05) levels in the activities of the enzymic antioxidants such as Superoxide dismutase (SOD) Catalase
(CAT), Glutathione peroxidase and the non enzymic antioxidants such as glutathione and Vitamin C in rats
induced hepatic damage by ethanol. Simultaneous administration of the leaf extract Mimosa pudica along
with the toxin ethanol in rats showed a considerable protection against the toxin induced oxidative stress
and liver damage as evidence by a significant increase (p<0.05) in antioxidant activities. The study reveals
that the co administration of Mimosa pudica aqueous extract significantly lowered the level of lipid
peroxidation in alcohol fed mice.
Key words- Mimosa pudica, Hepatoxicity, antioxidants
Introduction
Liver damage due to consumption of alcohol may
be caused by oxygen radicals such as
Superoxide and hydroxyl radicals generated
during the metabolism of ethanol by the
microsomal oxidising system [1]. Antioxidants are
likely to provide beneficial effects on hepatocytes
desensitization against oxidant stress while
inhibiting primary mechanism for expression of
pro-inflammatory and cytotoxic mediators [2]. In
spite of tremendous scientific advancement in the
field of hepatology in recent years, liver problems
are on the rise. [3]. Ingested alcohol produces
striking metabolic imbalances in the liver and
leads to the formation of reactive oxygen species
[4] and free radial increase [5]. The free radical
produced during the metabolism of the drug is
considered to be responsible for alteration
induced in lipid peroxidation, enzymic and non
enzymic antioxidants [6]. Antioxidants stop the
free radical from forming in the first place or
interrupt an oxidizing chain reaction to minimize
the damage caused by free radicals [7].
Medicinal plants are considered to be an
important source of antioxidant compounds and
the therapeutic benefit of many medicinal plants
in often attributed to their antioxidant properties
[8]. Mimosa pudica is an annual shrub found
through out India. Stem and rachis are clothed
with prickles arising from bulbous bases.
Previous studies have shown that Mimosa pudica
cures congestion with flem, biliousness,
leucoderma and diseases of blood [9]. The
present study investigates Mimosa pudica used
in herbal medicine for their potential to scavenge
free radicals as consequence may be considered
as effective sources for combating oxidative
hepatic damage [10]. Although a number of
reports described the usefulness of this herb
against various disorders, no studies have been
published to date describing its beneficial role
against ethanol induced hepatotoxicity in rats. In
this study, the extent of liver damage was
assessed by determining the hepatic antioxidant
status by measuring the hepatic content of SOD,
GSH, CAT, Reduced glutathione Vitamin C and
TBARS expressed as malanoldialdehyde
equivalents.
Materials and methods
Animals
Male albino Wister rats (150 gm) were housed in
large spacious cages and were fed on standard
pellet diet (M/s Hindustan Lever Ltd.,) and water
ad libitum. The experimental procedures were
carried out in strict compliance with the Animal
Ethics committee’s rules and regulations of this
institute.
Plant material
The plant material was taxonomically identified
by Prof. Parthiban, Associate, Prof. (Forestry),
TNAU, Tamil Nadu, India.
Experimental design
The rats were divided into four groups (n = 4).
The ethanol dose for rats was calculated on the
basis of their body weight
Group I:-
Control – Normal Healthy Rats with control
vehicle i.e., distilled water.
Group II:-
Alcohol Control – Rats administered with 40%
alcohol orally.
Group III:-
Simultaneous Induction and Treatment – (Alcohol
+ Aqueous extract) Simultaneous administration
of 40 % alcohol and aqueous plant (Mimosa
pudica) extract (1 gm /kg body wt.) was given
orally.
Group IV:-
Aqueous Extract Control – Rats administered
with aqueous extract (1gm/kg body wt.) of the
plant Mimosa pudica orally.

Antihepatotoxic and antioxidant defense potential of Mimosa pudica
International Journal of Drug Disco very, ISSN: 0975–4423, Volume 1, Issue 2, 2009
2
Preparation of sample for antioxidant studies
After the end of the experimental period of 30
days the rats were deprived of food over night
and sacrificed by cervical dislocation. Blood was
collected and kept for 30 minutes without
disturbing. The blood clots were then centrifuged
for 15 minutes at 2000 rpm to separate the serum
and it is used for antioxidant analysis. The liver
was washed with ice cold tris buffered saline,
blotted dry and 10% homogenate was prepared
using tris buffered saline (pH 7.4) to follow lipid
peroxidation rate. 15% homogenate were
prepared using phosphate buffered saline (pH
7.0) in cold condition for the estimation of
antioxidants and 20% liver homogenate was
prepared using 5% trichloroacetic acid to
estimate reduced glutathione. The homogenate
was then centrifuged at 2000 rpm for 10 minutes
and the supernatant was used for the
experimental analysis.
Results
Antioxidant levels in Mimosa pudica
Enzymic antioxidants
The activity of the enzymic antioxidants such as
Superoxide dismutase, peroxidase, catalase and
polyphenol oxidase were found to be present in
the plant extract of Mimosa pudica. Superoxide
dismutase activity was found to be maximum in
the leaf sample followed by polyphenol oxidase.
Peroxidase activity was found to be less when
compared to Superoxide dismutase. Polyphenol
was found to be 5.3 X 10¯ ³ units as catechol
oxidase and laccase respectively [Table I]. When
different extracts of Mimosa pudica were
compared for free radical, scavenging activity
aqueous extract showed higher percentage of
inhibition (89.73 %) over ethanol, methanol,
chloroform and petroleum ether extract. Based
on the above results water extract of Mimosa
pudica was selected for its free radical
scavenging activity. Lipid peroxidation is a free
radical mediated process and acts as a potential
marker of susceptibility of early and irreversible
tissue damage. The antiperoxidative effect of
Mimosa pudica against alcohol toxicity is
illustrated in Table II and III; the levels of non-
enzymic antioxidants (total phenols, flavonoids,
GSH, vitamin C and vitamin E are shown (Tables
II and III). The levels of the effect of Mimosa
pudica (various extract sournces) on in vitro lipid
peroxidation in ethanol induced rats were shown
(Table IV). The hepatoprotective effect of Mimosa
pudica against alcohol toxicity is presented in
Table V. The levels of TBARS in liver were
elevated in alcoholic group when compared with
control group. Our study reveals that co
administration of Mimosa pudica aqueous extract
significantly lowered the level of lipid peroxidation
in alcohol fed mice.
In vitro inhibition of lipid peroxidation
Free radical scavenging activity of different
extracts of Mimosa pudica induced lipid
peroxidation is depicted in Table-II. When
different extracts of Mimosa pudica were
compared for free radical, scavenging activity
aqueous extract showed higher percentage of
inhibition (89.73 %) over ethanol, methanol,
chloroform and petroleum ether extract. Based
on the above results water extract of Mimosa
pudica was selected for its free radical
scavenging activity. Lipid peroxidation is a free
radical mediated process and acts as a potential
marker of susceptibility of early and irreversible
tissue damage. The antiperoxidative effect of
Mimosa pudica against alcohol toxicity is
illustrated in Table-III. The hepatoprotective effect
of Mimosa pudica against alcohol toxicity is
presented in Table III. The levels of TBARS in
liver were elevated in alcoholic group when
compared with control group. Our study reveals
that co administration of Mimosa pudica aqueous
extract significantly lowered the level of lipid
peroxidation in alcohol fed mice.
Discussion
The first line of cellular defense against free
radicals consists of catalase, super oxide
dismutase and peroxidase [11]. These enzymes
react directly with oxygen radicals to yield non –
radical products [12]. A significant reduction
(P<0.05) in the activity of antioxidant enzyme like
SOD and catalase were observed in rats
intoxicated with alcohol (Group II). But Co-
administration with Mimosa pudica extract
increases its activity. The enzyme antioxidant
levels were increased when alcohol fed rats were
treated with Mimosa pudica. Inhibition of SOD
and catalase activities could be due to
exhaustion or inactivation as a result of oxidative
stress [13]. The increase in the SOD and
Catalase activity by treatment with aqueous
extract of Mimosa pudica effectively eliminates
the super oxides and peroxides by alcohol
intoxication. Table IV depicts the activity of
Glutathione-S-transferase and Glutathione
peroxidase. Recent studies have indicated that
the cytosolic liver enzyme -glutathione S-
transferase ( GST) functions as an early and
sensitive indicator of hepatocyte damage caused
by various adverse conditions, including
hemorrhagic shock [14] ischemia and
reperfusion[15] liver transplant rejection [16] and
acetaminophen overdose [17]. Liver failure can
be manifested by the increased level of GST or
when GST remains persistently elevated [18].
Hepatocytes in severe alcoholic liver disease
coexpressed both alpha and pi class glutathione
S-transferase [19]. The activity of Glutathione –
S- transferase was considerably elevated in
alcohol group than in other groups. Severe liver
damage was observed on exposure to alcohol.

Nazeema TH and Brindha V
Copyright © 2009, Bioinfo Publications, International Journal of Drug Discovery, ISSN: 0975–4423, Volume 1, Issue 2, 2009
3
On administration of Mimosa pudica extract the
rat liver cells were protected from oxidative
damage. GPx and GR activities in haemolysate
decreased after 12 weeks of exposure to Chronic
ethanol treatment in mice [20]. The activity of
GPx was significantly decreased in the liver of
rats treated with alcohol. The decreased level of
GPx can result in increased level of free radical
[21] which induced lipid peroxidation [22]. Co-
administration of Mimosa pudica showed the
adaptive nature of the system against the
damage of super oxide and peroxide radical. This
agrees with the result of Omotuyi I. O. and
Oluyemi, who reported a decrease in Glutathione
peroxidase which requires selenium for its
optimum activity [23]. Selenium is decreased
during increased free radical formation [24]. GSH
antioxidant system consists of array of enzymic
and non enzymic pathway involved in
neutralisation of reactive free radical species [25].
Under conditions of increased oxidative stress to
cells, levels of GSH are usually reduced [26]. The
hepatoprotection was associated with the
significant increase in hepatic GSH status, as
indicated by substantial increase in tissue GSH
levels in Mimosa pudica treated rats.
From our finding it was observed that the activity
of Vitamin C and reduced Glutathione decreased
significantly in the liver tissues of animals
administered with alcohol when compared to the
control. Vitamins C and E are antioxidants that
scavenge for free radicals [27]. The level of
vitamin C and reduced glutathione was near
normal when Mimosa pudica extract was fed with
alcohol in rats. GSH, Vitamin E and C exist in
their inter convertible forms and participate in the
detoxification of the toxic reactive oxygen species
[28]. So the present result indicates the protective
potential offered by the Mimosa pudica extract
against hepatic dysfunction caused by alcohol.
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Antihepatotoxic and antioxidant defense potential of Mimosa pudica
International Journal of Drug Disco very, ISSN: 0975–4423, Volume 1, Issue 2, 2009
4
Table I - Levels of enzymic Antioxidants in Mimosa pudica
PPO 1 unit = Amount of catechol oxidase / laccase which transforms 1 units of dihydric phenol to
Quinone / minute
Peroxidase 1 unit = Change of absorbance / minute at 430 nm
SOD 1 unit = Amount of enzyme that gives
50% inhibition of the extent of NBT oxidation.
Table II - Levels of non-enzymic Antioxidants in Mimosa pudica
Table III- Levels of non-enzymic Antioxidants in Mimosa pudica
Table IV-Effect of Mimosa pudica on invitro lipid peroxidation in ethanol induced rats
Table V- effect of Mimosa pudica on lipid peroxidation and antioxidants in liver of rats
S.NO Parameters Group I Group II Group III Group IV
1
2
3
4
5
6
7
SOD units
CAT units
GPX units
GST units
GSH (nmoles/g tissue)
Vit C (mg/g tissue)
LPO (nmoles of MDA/g tissue)
7.626 ± 0.07
80.13 ± 2.43
4.36 ± 0.04
1.934 ± 0.02
55.13 ± 0.39
0.796 ± 0.03
1.18 ± 0.05
4.665 ± 0.04 a
55.18 ± 0.84 a
3.32 ± 0.07 a
2.518 ± 0.01 a
31.26 ± 4.50 a
0.31 ± 0.08 a
1.95 ± 4.36 a
6.269 ± 0.07 b
73.18 ± 0.70 b
3.536 ± 0.05 b
2.15± 0.07 b
48.08 ± 0.23 b
0.57 ± 0.11 b
1.45 ± 0.12 b
6.558 ± 0.02
84.23 ± 0.74
4.142 ± 0.09
1.811 ± 0.07
54.96 ± 0.24
0.647 ± 0.04
1.20 ± 0.042
(Values are mean ± S.D. from 6 animals in each group)
Statistical Comparison:
a = significant (p< 0.05) when Group II is compared with Group I
b = significant (p< 0.05) when Group III is compared with Group II
SOD - Units: 50 % inhibition of nitrite/min/mg/protein
Catalase - Units: n moles of H2O2 decomposed / min / mg protein
GPx - Units: n moles of GSH / min / mg protein
GST - Units: n moles of CDNB conjugated / min / mg protein
Enzymic Antioxidants
Poly phenol oxidase
units / g tissue (PPO)
Catechol
oxidase
Laccase
Superoxide
Dismutase
units/g tissue
(SOD)
Peroxidase
units/g tissue
5.3 X
10¯ ³
4.6 X 10¯ ³ 172.78 0.966
Non – Enzymic Antioxidants
Total
proteins
mg/g
tissue
Total Phenol
mg catechol/g
tissue
Flavonoids
mg/g
tissue
GSH
nmoles/g
tissue
92.34 27.67 0.346 210.12
Non – Enzymic Antioxidants
Vitamin E µg/g
tissue
Vitamin C
µg/g
tissue
Tannins,
mg/g
tissue
175.81 5.67 87.576
Plant Extract Percentage
inhibition
Aqueous
Alcohol
Methanol
Petroleum Ether
Chloroform
89.73 %
82.88 %
80.23 %
40.34 %
52.46 %