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Indian Journal of Pharmacology
Medknow Publications on behalf of Indian Pharmacological Society
ISSN: 0253-7613 EISSN: 1998-3751
Vol. 36, Num. 5, 2004, pp. 312-313

Indian Journal of Pharmacology, Vol. 36, No. 5, October, 2004, pp. 312-313

Research Letter

Prevention of carbon tetrachloride-induced hepatotoxicity in rats by Adhatoda vasica leaves

Department of Pharmacology, Post Graduate Institute of Basic Medical Sciences, University of Calcutta, Kolkata
Correspondence Address:Department of Pharmacology, Post Graduate Institute of Basic Medical Sciences, University of Calcutta, Kolkata

Code Number: ph04104


The plant Adhatoda vasica Nees (AV) of the Acanthaceae family has been used for thousands of years in India. Extracts of the leaves of AV are extensively used in cough, asthma, bronchitis, tuberculosis, inflammation and allergy.[1],[2],[3] Several active constituents have also been isolated from different parts of AV.[4] Though the plant is traditionally used in the treatment of jaundice in Bengal, more evidence is needed to substantiate its pharmacological effects.

It is well known that the hepatotoxic effect of carbon tetrachloride is due to the oxidative damage by free radical generation, and antioxidant property is claimed to be one of the mechanisms of hepatoprotective drugs.[5] Therefore, the aim of the present study was to evaluate the antioxidant effect of AV in carbon tetrachloride (CCl4)-induced hepatotoxicity in rats.

Fresh leaves of AV were air-dried, powdered and soaked in 80% ethanol for 72 h. The extract was then filtered, concentrated under vacuum and lyophilized to obtain a solid mass (6.2%). The chemical constituents of the ethanolic extract were investigated. From preliminary phytochemical analysis it was found that the extract showed positive response for the presence of flavonoids, tannins, alkaloids, reducing sugars and saponins.[6] In atomic absorption spectrophotometric (AAS) analysis, the extract revealed the presence of Mg, Co, Cu, Mn and Cr in trace amounts.

The hepatoprotective study was carried out in adult male Wistar rats (150-175 g). They were housed in clean polypropylene cages and fed with commercial rat chow and water ad libitum. Permission from the institutional ethical committee for laboratory use of animals was duly obtained. A total of 30 animals were equally divided into 5 groups (n = 6 in each group). Group I (normal control) and Group II (CCl4-treated control) were given 0.9% saline (5 ml/kg, b.w.) for 9 days. Group III and Group IV were pretreated with AV (100 and 200 mg/kg, p.o., respectively) for 9 days, while Group V was pretreated with silymarin (25 mg/kg, p.o.) for 9 days. Liver damage was induced in these rats, except Group I, with 1:1 (v/v) mixture of CCl4 and olive oil (1ml/kg, s.c.) at Day 7 while, olive oil (0.5 ml/kg, s.c.) was injected to Group I.[7],[8] The doses of the test drug were selected on the basis of earlier work in our laboratory.[9] After 48 h of CCl4 treatment, i.e., on the 9th day the animals were sacrificed under anesthesia and blood was collected for the assay of serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT) and serum alkaline phosphatase (ALP).[8] The livers were removed immediately, washed with ice-cold saline and a 10% homogenate was prepared in phosphate buffer (pH 7.0). The homogenate was centrifuged at 3000 rpm for 15 min at 4oC and the supernatant was used[6] for the estimation of thiobarbituric acid reacting substances (TBARS), superoxide dismutase (SOD), catalase and reduced glutathione (GSH) and protein. The pieces of liver were preserved in 10% formaldehyde solution for histological study.[7] The results were statistically analyzed using one-way analysis of variance (ANOVA) followed by Student-Newman-Keuls test. P values<0.05 were considered significant.

It is well established that CCl4 is metabolized in the liver to the highly reactive trichloromethyl radical and this free radical leads to auto-oxidation of the fatty acids present in the cytoplasmic membrane phospholipids and causes functional and morphological changes in the cell membrane.[5] This is evidenced by an elevation of the serum marker enzymes namely SGOT, SGPT and ALP in CCl4-treated rats.[7],[8] Pretreatment with the test drug AV in both doses as well as pretreatment with standard drug silymarin significantly (P<0.01) reduced these liver enzyme levels dose dependently, showing that AV has hepatoprotective action [Table - 1]. Histopathological findings indicated that pretreatment with AV (100 and 200 mg/kg) offered protection to the hepatocytes from damage induced by CCl4, with mild fatty changes in the hepatic parenchymal cells, which corroborated the changes observed in the hepatic enzymes.

In our earlier work, it was reported that the antioxidant activity or inhibition of the generation of free radicals is important in the protection against CCl4-induced liver lesion.[7] At present, a significant elevation in the levels of end products of lipid peroxidation or MDA in the liver of rats treated with CCl4 was observed when compared to normal control [Table - 1]. Pretreatment with AV (100 and 200 mg/kg) as well as silymarin significantly (P<0.01) reversed these changes. AV also enhanced significantly (P<0.01) the protective enzymes, SOD and catalase when examined in the liver homogenate. SOD and catalase act as protective enzymes against lipid peroxidation in liver tissues. In the present study, CCl4 further depleted GSH concentration in the liver of rats [Table - 1], whereas, AV as also silymarin pretreatment reversed this effect (P<0.01). Flavonoids, tannins and microelements have been suggested to act as antioxidants and exert their antioxidant activity by scavenging the lipid peroxidation.[10] Therefore, the present work provides conclusive evidence for the hepatoprotective effect of AV against carbon tetrachloride-induced hepatotoxicity. The plausible mechanism of the hepatoprotective action of AV might be at least partly due to its antioxidant effect.


1.Dhuley JN. Antitussive effect of Adhatoda vasica extract on mechanical or chemical stimulation-induced coughing in animals. J Ethnopharmacol 1999;67:361-5.  Back to cited text no. 1  [PUBMED]  [FULLTEXT]
2.Grange JM, Snell NJ. Activity of bromhexine and ambroxol, semi-synthetic derivatives of vasicine from the Indian shrub Adhatoda vasica, against Mycobacterium tuberculosis in vitro. J Ethnopharmacol 1996;50:49-53.   Back to cited text no. 2  [PUBMED]  
3.Chakraborty A, Brantner AH. Study of alkaloids Adhatoda vasica Nees on their antiinflammatory activity. Phytother Res 2001;15:532-4.  Back to cited text no. 3  [PUBMED]  [FULLTEXT]
4.Wagner H. Search for new plant constituents with potential antiphlogistic and antiallergic activity. Planta Med 1989;55:235-41.  Back to cited text no. 4  [PUBMED]  
5.Recknagel RO. Carbon tetrachloride hepatotoxicity. Pharmacol Rev 1967;19:145-208.  Back to cited text no. 5  [PUBMED]  
6.Trease GE, Evans WC. In: Pharmacognosy. 12th Ed. Bailliere Tindall; ELBS Publication 1985.  Back to cited text no. 6    
7.Bhattacharyya D, Mukherjee R, Pandit S, Das N, Sur TK. Prevention of carbon tetrachloride induced hepatotoxicity in rats by Himoliv®, a polyherbal formulation. Indian J Pharmacol 2003;35:183-5.   Back to cited text no. 7    
8.Bhattacharyya D, Mukherjee R, Pandit S, Das N, Sur TK. Hepatoprotective effect of Himoliv®, a polyherbal formulation in rats. Indian J Physiol Pharmacol 2003;47:435-40.   Back to cited text no. 8    
9.Pandit S, Sur TK, Debnath PK, Bhattacharyya D. Hepatoprotective and antioxidant evaluation of leaves of Adhatoda vasica Nees. Proc. 89th Indian Science Congress, Part III. Lucknow, India 2002. p. 7.  Back to cited text no. 9    
10.Yuting C, Rongliang Z, Zhongjian J, Yong J. Flavonoids as superoxide scavengers and antioxidants. Free Radical Biol Med 1990;9:19-23.  Back to cited text no. 10    

Copyright 2004 - Indian Journal of Pharmacology

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