Aghel, N., Rashidi, I., Mombeini, A. (2010). Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice. Iranian Journal of Pharmaceutical Research, Volume 6(Number 4), 285-290. doi: 10.22037/ijpr.2010.734
N Aghel; I Rashidi; A Mombeini. "Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice". Iranian Journal of Pharmaceutical Research, Volume 6, Number 4, 2010, 285-290. doi: 10.22037/ijpr.2010.734
Aghel, N., Rashidi, I., Mombeini, A. (2010). 'Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice', Iranian Journal of Pharmaceutical Research, Volume 6(Number 4), pp. 285-290. doi: 10.22037/ijpr.2010.734
Aghel, N., Rashidi, I., Mombeini, A. Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice. Iranian Journal of Pharmaceutical Research, 2010; Volume 6(Number 4): 285-290. doi: 10.22037/ijpr.2010.734
Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice
Many hepatoprotective herbal preparations have been recommended in alternative systems of medicine for the treatment of hepatic disorders. No systematic study has been done on protective efficacy of Capparis spinosa (Capparidaceae) to treat hepatic diseases. Protective action of C. spinosa ethanolic root bark extract was evaluated in this study in an animal model of hepatotoxicity, which was induced by carbon tetrachloride. Healthy male mice (30-35 g body weight, 6-8 week old) were divided into 7 groups. Group 1 was normal control group; Group 2, the hepatotoxic group was given CCl4; Group 3 was administered olive oil (vehicle); Groups 4-6 received different doses of ethanolic root bark extract (100, 200 & 400 mg/kg) with CCl4; Group 7 was administered overdose of the extract (800 mg/kg). The parameters studied were alanine transaminase and aspartate transaminase activities and duration of sleep. The hepatoprotective activity was also supported by histopathological studies of liver tissue. Results of the biochemical studies of blood samples of CCl4 treated animals showed significant increase in the levels of serum enzyme activities, reflecting the liver injury caused by CCl4. Whereas blood samples from the animals treated with ethanolic root bark extracts showed significant decrease in the levels of serum markers, indicating the protection of hepatic cells. The results revealed that ethanolic root bark extract of C. spinosa could afford significant dose-dependent protection against CCl4 induced hepatocellular injury.
Acute and Subchronic Toxicity?of Teucrium polium Total Extract in Rats
Iranian Journal of Pharmaceutical Research
(2007), 6 (4): 285-290
Received: March 2006
Accepted: September 2006
Copyright ? 2007 by School of Pharmacy Shaheed Beheshti University of Medical Sciences and Health Services
Original Article
Hepatoprotective Activity of Capparis spinosa Root Bark Against
CCl4 Induced Hepatic Damage in Mice
Nasrin Aghela*, Iran Rashidib and Amir Mombeinia
aDepartment of Pharmacognosy, School of Pharmacy, Ahwaz Joundishapour Medical
Sciences University, Ahwaz, Iran. bDepartment of Pathology, School of Medicine,
Ahwaz Joundishapour Medical Sciences University, Ahwaz, Iran.
Abstract
Many hepatoprotective herbal preparations have been recommended in alternative
systems of medicine for the treatment of hepatic disorders. No systematic study
has been done on protective efficacy of Capparis spinosa (Capparidaceae) to
treat hepatic diseases. Protective action of C. spinosa ethanolic root bark
extract was evaluated in this study in an animal model of hepatotoxicity, which
was induced by carbon tetrachloride.
Healthy male mice (30-35 g body weight, 6-8 week old) were divided into 7
groups. Group 1 was normal control group; Group 2, the hepatotoxic group was
given CCl4; Group 3 was administered olive oil (vehicle); Groups 4-6 received
different doses of ethanolic root bark extract (100, 200 & 400 mg/kg) with CCl4;
Group 7 was administered overdose of the extract (800 mg/kg). The parameters
studied were alanine transaminase and aspartate transaminase activities and
duration of sleep. The hepatoprotective activity was also supported by
histopathological studies of liver tissue.
Results of the biochemical studies of blood samples of CCl4 treated animals
showed significant increase in the levels of serum enzyme activities, reflecting
the liver injury caused by CCl4. Whereas blood samples from the animals treated
with ethanolic root bark extracts showed significant decrease in the levels of
serum markers, indicating the protection of hepatic cells. The results revealed
that ethanolic root bark extract of C. spinosa could afford significant
dose-dependent protection against CCl4 induced hepatocellular injury.
Capparidaceae are a medium-sized family of approximately 40-45 genera and
700-900 species, whose members present considerable diversity in habit, fruit,
and floral features (1-3). Capparidaceae are pantropical in distribution, being
most conspicuous in tropical seasonally dry habitants (4). Capparis spinosa
(caper), a winter-deciduous perennial shrub, is a consistent floristic element
of Mediterranean ecosystems, growing from May to October, i.e. entirely during
the prolonged summer drought. Capers, a useful and beautiful plant in the
Capparidaceae, can today be found growing wild all over Mediterranean, and are
frequently cultivated (e.g., in France, Spain, Italy and Algeria; furthermore,
Iran, Cyprus and Greece produce significant amounts of the plant); their origin
is, though, supposed in the dry areas of Western or Central Asia.
The caper plant is well known for the culinary properties of the caper, the
immature flower buds which have been pickled in vinegar or preserved in granular
salt. They have long been used in recipes of salads, pasta, meat, sauces and
garnishes to add a pungent spicy flavor and aroma to food. The caper had other
uses prior to its use in cooking. The first recorded use of the caper bush was
for medicinal purposes in 2000 BC by the Sumerians. It has been suggested that
Capers have been used or are still being used in reducing flatulence, in the
treatment of rheumatism, anemia and gout. Further medical uses include ingesting
for improving liver functions, as diuretics, kidney disinfectants (5). Infusions
and decoctions from caper root bark have been traditionally used for dropsy,
anemia, arthritis and gout (6, 7). The root-bark is analgesic, anthelmintic,
antihaemorrhoidal, aperient, depurative, diuretic, emmenagogue, expectorant,
hepatoprotective, tonic and vasoconstrictive (7-9). Externally, it is used to
treat skin conditions, capillary weakness and easy bruising (10). The bark is
harvested in the autumn and dried for later use (6).
Liver, an important organ actively involved in metabolic functions, is a
frequent target of a number of toxicants (11). The principal cause of carbon
tetrachloride CCl4 induced hepatic damage is lipid peroxidation and decreased
activities of antioxidant enzymes and generation of free radicals (12, 13). The
resulting hepatic injury was characterized by leakage of cellular enzymes into
the blood stream and by centrilobular necrosis (14).
Presently, the use of herbal medicines for prevention and control of chronic
liver diseases is in the focus of attention for the physicians, pharmaceutical
manufacturers and patients; the reasons for such shift toward the use of herbals
include the expensive cost of conventional drugs, adverse drug reactions, and
their inefficacy.
Many hepatoprotective herbal preparations have been recommended in alternative
systems of medicine for the treatment of hepatic disorders. No systematic study
has been done on protective efficacy of Capparis spinosa to treat hepatic
diseases. Therefore, the protective action of Capparis spinosa root bark extract
was evaluated in an animal model of hepatotoxicity induced by carbon
tetrachloride.
Experimental
Plant extract
The roots of Capparis spinosa were collected in July 2005 from Shoosh, province
of Khuzestan, south-west of Iran. The plant was authenticated by Department of
Botany, Faculty of Agriculture, Shahid Chamran University, Ahwaz, Iran and the
voucher specimen has been deposited in the herbarium of the Department of
Pharmacognosy, Faculty of Pharmacy, Ahwaz Joundishapour Medical Sciences
University. Ahwaz, Iran. The collected roots were washed and the bark was peeled
off and then air-dried at shade and room temperature.
Hydroalcoholic extract was prepared by using the soxhlet method. The coarsely
powdered plant barks (200 g) were extracted with ethanol (80% v/v in water) for
5 h. After filtration using Whatman No.1 filter paper, the ethanolic extract was
evaporated in vaccum below 50 ?C. The yield of evaporation and solvent removal
of hydroalcoholic extract of Capparis spinosa was 7.35% w/w, which was stored in
refrigerator for further use.
Experimental animals
Healthy, 6-8 week old male mice (30-35g) were obtained from Razi Vaccine & Serum
Research Institute, Karaj, Iran. Animals were maintained on a standard
laboratory diet. Food and water were given ad libitum. They were housed in
standard stainless-steel cages at a 12-h cycle of light and dark. Room
temperature was kept at 24?2?C and humidity maintained at 50%. All the chemicals
used were of the analytical grade from standard companies.
Treatment of animals
Mice were randomly divided into 7 groups with 7 animals in each group. Group 1
served as negative control and was administered a single daily dose of 0.2 ml of
normal saline by oral gavage. Group 2, the positive control was given daily
carbon tetrachloride (0.2 mg/kg/0.2 ml in olive oil) orally. Group 3 was
administered a single daily dose of 0.2 ml olive oil (vehicle) orally. The drug
control groups (4, 5 and 6) were given the plant extracts orally in doses of
100, 200 and 400 mg/kg/0.2 ml (in normal saline), respectively, one hour after
the administration of carbon tetrachloride (0.2 mg/kg/0.2ml in olive oil), for
four days (15, 16). To evaluate the effects of the root bark extract overdose on
the studying factors, the 7th group was selected, which received 800 mg/kg daily
of the extract. In the 5th day, all animals were given sodium thiopental (25
mg/kg/0.2 ml) intraperitoneally and the effects of extracts on CCl4-induced
prolongation of thiopental sleeping time were studied (17). All animals were
sacrificed in the 6th day, blood samples were collected and serum was separated.
The liver was excised, fixed in 10% buffered formalin for histopathological
assessment of liver damage.
Assessment of liver damage
Liver damage was assessed by the estimation of serum activities of alanine
aminotransferase (ALT) and asparate aminotransferase (AST) using commercially
available test kits. Histopathological assessment of liver damage was done by
studying Haematoxylin and Eosin (H&E) stained slides of liver tissue, including
cell necrosis, fatty change, infiltration of kupffer cells and lymphocytes.
Statistical analysis
Data were analyzed by one-way analysis of variance (ANOVA), followed by the
Dunnett test for multiple comparisons using SPSS software and p<0.05 was
regarded as significant.
Results and Discussion
The results of hepatoprotective effects of Capparis spinosa extracts on
CCl4-intoxicated mice are shown in Table 1. In the CCl4-treated control, serum
ALT and AST were increased (191.80 and 239.40 U/ml, respectively), whereas these
values showed 114.00 and 143.20 U/ml in normal saline group, respectively. In
contrast, serum ALT and AST in the groups treated with 100, 200 and 400 mg/kg of
root bark extracts decreased significantly (p<0.05) in a dose dependent manner
toward normalization. Treatment with 400 mg/kg of root bark extract showed
highly significant activity. There was no significant changes in the activities
of serum ALT and AST in the overdose group, means the plant root bark extract is
not hepatotoxic itself.
The results of Capparis spinosa root bark extract on the thiopental-induced
sleeping time in mice are presented in Table 2. They indicated that CCl4
produced significant increases in thiopental induced sleeping time compared to
the control groups (1 and 3). The treatment with different doses of root bark
extracts, especially with 400 mg/kg, resulted in decreased in thiopental induced
sleeping time compared to the group received CCl4 only.
Histophathological studies also provided supportive evidence for biochemical
analysis. Histology of the liver section of normal control animals (group 1)
showed normal hepatic cells each with well preserved cytoplasm, prominent
nucleus and nucleolus and well brought out central vein (Figure 1). The liver
sections of CCl4-intoxicated mice showed massive fatty changes, necrosis,
ballooning degeneration and broad infiltration of the lymphocytes and Kupffer
cells around the central vein and the loss of cellular boundaries (Figure 2).
The extract toxicity control group (group 7), showed the normal parenchymal
architecture with cords of hepatocytes, portal tracts and central veins without
noticeable alterations compared to the normal saline control group. The toxin
mediated changes in livers of mice pre-treated with Capparis spinosa root bark
extracts in different doses one hour after the administration of CCl4 were less
intensity than those observed in the livers of carbon tetrachloride treated
mice. The histological architecture of liver sections of mice treated with 400
mg/kg plant extract showed normal lobular pattern with a mild degree of fatty
change, necrosis and lymphocyte infiltration almost comparable to the normal
control (Figure 3).
The present study showed for the first time, Capparis spinosa root bark extract
possess hepatoprotective activity as evidenced by the significant inhibition in
the elevated levels of serum enzyme activities induced by CCl4. Capparis spinosa
root bark extract given orally (100, 200 & 400 mg/kg) once daily for 4 days
showed dose dependent hepatoprotective activity. However, highly significant
effect was seen with 400 mg/kg body weight against CCl4 induced hepatic damage.
It is well established that CCl4 induces hepatotoxicity by metabolic activation,
therefore it selectively causes toxicity in liver cells maintaining semi-normal
metabolic function. CCl4 is bio-transformed by the cytochrome P450 system in the
endoplasmic reticulum to produce trichloromethyl free radical (٭CCl3). This free
radical then combined with cellular lipids and proteins in the presence of
oxygen to form a trichloromethyl peroxyl radical, which may attack lipids on the
membrane of endoplasmic reticulum faster than trichloromethyl free radical.
Thus, trichloromethylperoxyl free radical leads to elicit lipid peroxidation,
the destruction of Ca2+ homeostasis, and finally, results in cell death (18).
Many compounds known to be beneficial against carbon tetrachloride-mediated
liver injury exert their protective action by toxin-mediated lipid peroxidation
either via a decreased production of CCl4 derived free radicals or through the
antioxidant activity of the protective agents themselves (19). The mechanism by
which Capparis spinosa exert its protective action against CCl4 induced
alternations in the liver may be (attributed to) due to the antioxidant effect
of the plant extract; but the suggestion needs to be more expleted.
In conclusion, the results indicated that under the present experimental
conditions, hydroalcoholic extract of Capparis spinosa root bark showed
hepatoprotective effects against CCl4 induced liver damage in mice.
Acknowledgement
The authors would like to thank Dr. H. Kalantari for his valuable comments and
helps.
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