Original Article

ISSN: 2230-7761

Hepatoprotective and antioxidant activity of the alcoholic extract of Ipomoea turpetnm against anti-TB drugs induced hepatotoxicity in rats.
*Gopi Sudheer Kumar J, Bala Krishna R.B2, Vijay Kumar G2, Ranganayakulu D.1 1 Sri Padmavathi School of Pharmacy, Tiruchanoor, Tirupathi, Andhra Pradesh, India. 2 A.K.R.G College of Pharmacy, Nallajerla, Andhra Pradesh, India. ABSTRACT
The hydroalcoholic root extract of Ipomoea turpethum Linn (Convolvulaceae) L. showed a significant dose dependent (200 mg, 400 mg/kg p.o) protective effect against isoniazid and rifampicin induced hepatotoxicity in albino rats. The degree of protection was measured by using biochemical parameters like serum glutamate pyruvate transaminases, serum glutamate oxaloacetate transaminases, alkaline phosphatase, total cholesterol, HDL- cholesterol, total protein,and total bilirubin. The plant extract prevented the toxic effects of combination of isoniazid and rifampicin (INH + RIF) on the above serum parameters. Histological studies and bio-chemical estimations also supported the hepatoprotective activity of the hydro-alcoholic root extract of Ipomoea turpethum L.

Corresponding Author, Gopi Sudheer Kumnar .J, M.Pharm.,

Assistant Professor of Pharmacology,

A.K.R.J College of Pharmacy, Nallajerla, Andhra Pradesh, India. Email: sudheerphrma1@gmail.com

KEY WORDS: Isoniazid, rifampicin, hepatic damage, hepatoprotection,

antioxidant parameters.

INTRODUCTION:
The liver is the major site of xenobiotic metabolism and excretion. Liver injury caused by xenobiotics such as toxic chemicals, drugs, herbal products, environmental chemicals and virus infiltration from ingestion or infection represents the leading cause of acute liver failure(1). The toxins absorbed from the intestinal tract gain access first to the liver resulting in a variety of liver ailments. Thus liver diseases remain one of the serious health problems (2). Hepatotoxicity has a considerable impact on health because many of the hepatic reactions induced by pharmaceutical preparations can be very severe(3). Drug-induced liver injury may account for as many as 10 percent of hepatitis cases in adults overall, 40 percent of hepatitis cases in adults over fifty years old, and 25 percent of cases of fulminant liver failure. In Western countries, paracetamol represents the first cause of all liver failures. But it accounts only for 25% - 40 % cases of fulminant hepatic failure(4). Whereas, antitubercular drugs being the second common cause of drug induced hepatotoxicity cause, the risk of hepatotoxicity based on data from four prospective Indian

studies was 11.5% compared with 4.3% in Western publications(5). Anti tubercular (AT) drugs are the commonest agents causing serious, clinically significant drug induced acute liver failure in India. Most commonly used AT drugs like isoniazid (INH), rifampicin (RIF) and pyrazinamide are hepatotoxic(6) (7). Ipomoea turpethum has been claimed to be useful in hepatotoxicity(8). Few reports are available supporting the hepatoprotective effect of Operculina turpethum white variety(9), but no scientific work was carried out to support the claims regarding hepatoprotective potential of Ipomoea turpethum black variety. Hence the present study aims to evaluate the hydro-alcoholic extract of root of Ipomoea turpethum for its hepatoprotective activity against INH and RIF induced hepatotoxicity in rats. MATERIALSAND METHODS Ipomoea turpethum was collected in and around Tiruchanoor and identified by Dr. N. Yasodamma, Prof. of Botany, Sri Venkateswara University, Tirupathi. Roots were separated, washed in water, chopped into pieces and then shade dried.The coarsely powdered root was extracted with ethanol: water (1:1) mixture on a reflex water

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bath for 3 hr. The cycle was repeated for three times. The extract was concentrated on rotary flash evaporator to semi solid consistency and then dried over a water bath (yield - 146.6 g/kg). Animals Wistar rats (150-200 g) body weight were obtained from Raghavendra enterprises, Bangalore, and they were housed under standard husbandry conditions, The experimental protocol was approved by the Institutional Animal Ethical Committee of Sri Padmavathi School of Pharmacy (1016/a/06/CPCSEA/005/2009). Drugs and Chemicals Silymarin and DTNB (5,5-di thio bis 2-nitro benzoic acid) were procured from Sigma-Aldrich Pvt. Ltd, (India) isoniazid and rifampicin procured from Lupin Ltd. India and all other chemicals and reagents used were of analytical grade, procured from SD fine chemicals Ltd, (India). SGPT, SGOT, alkaline phosphatase, total cholesterol and HDL, total bilirubin and Total Protein estimation kits were procured from Kamineni Life Sciences Pvt. Ltd. (India). Acute toxicity studies Acute oral toxicity study was performed as per OECD 423 guidelines (acute toxic class method). Wistar albino rats of either sex were selected randomly and divided into five groups (n = 2). The animals were fasted overnight and hydro-alcoholic extract in doses of 500, 1000, 2000, 4000 mg/kg body weight, were administered orally to II V groups. Group I which received vehicle (water) served as control. The animals were observed continuously for 2 hrs, and then intermittently for 6 hr and at the end of 24 hrs, the number of deaths was noted to determine LD50 of the extract (Annie et al., 2004). Animals were also observed for behavioral, neurological and autonomic profiles simultaneously(10). Hepatoprotective activity Hepatotoxicity was induced in the present study, using isoniazid (200 mg/kg, p.o) and rifampicin (200 mg/kg, p.o) for 28 days and silymarin (100 mg/kg, p.o) was used as the standard. Experimental animals were randomly divided into five groups, each group containg 6 animals and treated for 28 days, as follows. Group I received only distilled water, serves as normal. Group II was treated with INH (200 mg/kg, p.o) and rifampicin (200 mg/kg, p.o), serves as control. Group III received silymarin (100 mg/kg, p.o) along with INH (200 mg/kg, p.o) and rifampicin (200 mg/kg, p.o), serves as standard. Group IV and V were treated with INH (200 mg/

kg, p.o), rifampicin (200 mg/kg, p.o) and Ipomoea turpethum root extract at doses of 200 and 400 mg/kg, p.o. respectively for evaluating the hepatoprotective effect of Ipomoea turpethum. Collection of blood samples The blood samples were withdrawn on 0th, 7th, 14th, 21st and 28th day from the retrorbital venous plexus of rats without any coagulant for the separation of serum. After collecting the blood in eppendroff tubes kept aside for 1 hr at room temperature and then serum was isolated by centrifugation at 2000 rpm for 15 min and stored until analyzed for various biochemical parameters. Liver parameters At the end of the treatment schedule, weight of the animals was noted and sacrificed by cervical dislocation for excision of livers. One lobe of the liver was immediately processed for antioxidant parameters. Other lobe of the liver was stored in 10% formalin solution for histological preparations. Statistical Analysis All the data was expressed as mean S.E.M. Statistical significance between more than two groups was tested using one way ANOVA followed by the Tukey test using computer based fitting program (Prism graph pad.). Statistical significance was set accordingly. RESULTS The hydro-alcoholic root extract of Ipomoea turpethum was found to be safe since no animal died even at the maximum single dose of 4000 mg/kg when administered orally and the animals did not show any gross behavioral changes. Hence, 1/20 and 1/10 of maximum tested dose (4000 mg/kg) were selected for the present study. Rats treated with INH+RIF (G-II) showed a significant increase in SGPT and SGOT levels on 7th, 14th, 21st and 28th day, when compared to normal group (G-I). The group (III) rats treated with standard drug silymarin (100 mg/kg) showed a significant decrease in SGPT levels on 7th, 14th, 21st and 28th day, when compared to control (G-II). The groups (IV and V) receiving hydro-alcoholic root extract of Ipomoea turpethum (200 mg/kg and 400 mg/kg) shows a dose dependent decrease on SGPT levels on 7th, 14th, 21st and 28th day when compared to control group (G-II) (Table-1and 2). Administration of INH + RIF induced a significant increase in ALP levels on 7th, 14th, 21st and 28th day in control group (G-II) when compared to normal group (G-I) . On treatment with silymarin a significant decreases in ALP levels on 7th, 14th, 21st and 28th day in standard group

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(G-III) when compared to control group (G-II) was observed Groups (IV and V) treated with hydro-alcoholic root extract of Ipomoea turpethum induces a significant decreases in ALP levels on 7th, 14th, 21st and 28th day. The effect was dose dependent when compared to control group (G-II) (Table-3). A significant increase in total cholesterol and total bilurubin levels in INH + RIF treated group (G-II) on 7th, 14th, 21st and 28th day when compared to normal group (GI).Silymarin treated group (G-III) shows significant decreases in both total cholesterol and bilurubin levels on 7th, 14th, 21st and 28th day was observed when compared to control group (G-II). Hydro-alcoholic root extract of Ipomoea turpethum treated groups (IV and V) shows a significant decreases in total cholesterol levels on 7th, 14th, 21st and 28th day when compared to control group (GII) (Table-4 and 7). INH + RIF receiving groups (G-II) shows a significant decrease in HDL- cholesterol levels on 7th, 14th, 21st and

28 th day when compared to the normal group (G-I). Silymarin receiving group (G-III) shows significant decreases in HDL- cholesterol levels on 7th, 14th, 21st and 28th day when compared to control group (G-II).Hydroalcoholic root extract of Ipomoea turpethum treated groups (IV and V) shows a significant decreases in HDLcholesterol levels on 7th, 14th, 21st and 28th day in both doses has shown dose dependent when compared to control group (G-II) (Table 5). The groups (IV and V) receiving hydro-alcoholic root extract of Ipomoea turpethum (200 mg/kg and 400 mg/kg) showed a significant increase in total proteins levels on 7th, 14th, 21st and 28th day in both doses has shown dose dependent when compared to control group (G-II) (Table-6).Administration of INH + RIF induced a significant increase in total bilirubin levels on 7th, 14th, 21st and 28th day in control group (G-II) when compared to normal group (G-I). On treatment with silymarin induced a

Table-1: Effect of hydro-alcoholic root extract of Ipomoea turpethum on SGPT.

Group I II III IV V Treatment Normal Control INH+RIF +Sy INH+RIF +IPT1 INH+RIF + IPT2 0 day 172.67.05 157.46.94 154.33.96 165.63.18 172.8 4.16 7th day 174.4 16.51 286.3 16.91a 164.4 16.91b 216.2 19.31b 213.9 10.77b SGPT (IU/L) 14th day 175.3 16.84 317.5 13.51a 183.5 18.14b 254.3 14.82b 236.5 16.17b 21st day 176.8 14.23 323.0 16.27a 170.5 18.50b 240.3 14.16b 223.7 19.41b 28th day 172.7 17.30 341.5 16.91a 161.9 17.75b 250.7 15.69b 219.2 11.81b

Table-2: Effect of hydro-alcoholic root extract of Ipomoea turpethum on SGOT.

Group Treatment 0 day 7th day SGOT (IU/L) 14th day 21st day 28th day

I II III IV V

Normal Control INH+RIF +Sy INH+RIF +IPT1 INH+RIF + IPT2

151.8 6.10 160.2 4.54 154.6 7.78 180.3 6.42 170.2 6.11

154.1 13.19 300.0 6.06

a

152.0 16.252 367.1 14.075

a

153.5 13.145 370.0 18.05

a

155.215.81 380.2 15.41a 183.1 16.16b 251.8 15.97b 224.6 16.94b

120.5 6.46b 232.3 16.64b 201.0 14.90b

201.2 16.06b 280.3 16.18b 240.4 17.17b

181.0 16.11b 259.5 14.65b 230.7 17.13b

RIF-Rifampicin (200 mg/kg), SY- Silymarin (100 mg.kg), IPT1- Ipomoea turpethum (200 mg/kg), IPT2- pomoea turpethum (400 mg/kg), a = p < 0.001, when compared to normal animals,a = p < 0.001, when compared to control animal.

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significant decreases in total bilirubin levels on 7th, 14th, 21st and 28th day in standard group (G-III) when compared to control group (G-II). Groups (IV and V) treated with hydro-alcoholic root extract of Ipomoea turpethum induces a significant decreases in total bilirubin levels on 7th, 14th, 21st and 28th day both the doses has shown dose dependent when compared to control group (G-II) (Table7).

III. In vivo antioxidant parameters In the present study, various antioxidant parameters were assessed in the liver homogenate at the end of the study on 29th day.Administration of INH + RIF diminished the antioxidant status in liver by decreases the catalase, GSH, SOD levels and increases the LPO levels in control group (G-II) when compared to normal group (G-I) (Table 8).

Table 3-Effect of hydro-alcoholic root extract of Ipomoea turpethum on ALP

Group I II III IV V Treatment Normal Control INH+RIF +Sy INH+RIF+IPT1 INH+RIF+IPT2 ALP (IU/L) 0 day 172.1 8.84 153.812.05 175.9 4.52 161.8 8.46 51.7 3.52 7th day 175.9 15.72 242.012.88a 160.817.01c 201.110.27b 199.5 7.56b 14th day 173.117.21 320.313.27a 190.615.95c 279.5 17.14 250.113.84c 21st day 174.316.09 331.018.25a 186.817.79c 246.012.01c 240.815.79c 28th day 174.915.85 341.416.35a 182.616.00c 244.317.88c 234.611.54c

RIF-Rifampicin (200 mg/kg), SY- Silymarin (100 mg.kg), IPT1- Ipomoea turpethum (200 mg/kg), IPT2- pomoea
turpethum (400 mg/kg), a = p < 0.001, when compared to normal animals,a = p < 0.001, when compared to control animal. Standard group (G-II) treated with silymarin increases the antioxidant status in liver by increasing the SOD, catalase, GSH levels and decreases the LPO levels when compared to control group (G-II).Both doses of hydro-alcoholic extract of Ipomoea turpethum shows the significant increases in antioxidant status in liver by increasing the SOD, catalase, GSH levels and decreasing the levels of LPO has shown dose dependent when compared to control group (G-II) (Table 8)). DISCUSSION Hepatotoxicity of anti-TB drugs is a serious problem because it causes significant morbidity and mortality that requires modification of the drug regimen (11) (12). The increased risk of hepatotoxicity with INH and rifampicin (RIF) combination has been attributed to the interaction between the metabolism of isoniazid and rifampicin.

Table-4: Effect of hydro-alcoholic root extract of Ipomoea turpethum on total cholesterol

Group I II III IV V Treatment Normal Control INH+RIF + SY INH+RIF+IPT1 INH+RIF+IPT2 Total cholesterol (mg/dl)
0 day 7 day
th

14th day

21st day

28th day

63.20 5.973 72.29 6.104 71.84 6.218 57.11 6.597 86.13 8.462

64.02 8.43 90.15 9.87 40.536.06b 60.20 7.62 58.60 6.96

67.965.88 140.618.55
a

67.81 6.48 150.416.52

a

63.188.49 149.916.54a 55.49 6.58c 88.95 9.84c 70.99 5.92c

50.75 5.96c 80.65 8.15c 70.95 5.92c

60.56 7.58c 105.8 6.65b 88.11 6.94c

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Table-5: Effect of hydro-alcoholic root extract of Ipomoea turpethum on HDL-cholesterol

Group I II III IV V Treatment Normal Control INH+RIF + SY INH+RIF+IPT1 INH+RIF+IPT2 HDL- cholesterol (mg/dl) 0 day 42.40 5.60 58.79 6.14 56.46 5.25 59.18 5.22 51.68 5.33 7 day 45.06 7.67 21.34 5.85 36.02 7.22 32.03 7.93 34.19 7.98
th

14th day 49.4 7.35 26.216.49 31.538.14 35.897.32 31.198.22

21st day 52.12 6.61 27.43 5.85 35.47 6.72 33.02 6.53 35.62 5.99

28th day 51.07 .72 20.894.33a 51.406.02b 46.985.61b 50.446.87b

Table-6: Effect of hydro-alcoholic root extract of Ipomoea turpethum on total protein

Total protein (mg/dl) Group I II III IV V Treatment Normal Control INH+RIF + SY INH+RIF +IPT1 INH+RIF + IPT2 0 day 5.31 0.628 4.25 0.679 4.46 0.312 5.07 0.506 5.24 0.428 7 day 4.88 0.43 3.16 0.37 4.17 0.59 3.80 0.54 3.04 0.38
th

14th day 5.11 0.78 2.98 0.37 4.31 0.53 4.13 0.63 4.56 0.51

21st day 4.24 0.63 3.27 0.68 5.07 0.67 3.97 0.41 5.25 0.76

28th day 4.94 0.45 2.46 0.42a 5.92 0.56c 4.930.62b 6.05 0.46c

Table-7: Effect of hydro-alcoholic root extract of Ipomoea turpethum on total bilirubin

Group I II III IV V Treatment Normal Control INH+RIF + SY INH+RIF +IPT1 INH+RIF +IPT2 Total bilirubin (mg/dl) 0 day 0.830.05 1.06 0.14 0.98 0.16 0.78 0.10 0.97 0.10 7 day 0.94 0.06 3.24 0.10
a th

14th day 1.010.03 3.440.03

a

21st day 0.76 0.02 2.98 0.03

a

28th day 0.89 0.03 3.14 0.02a 0.98 0.04c 1.12 0.03c 0.87 0.03c

2.74 0.05c 2.91 0.06b 2.37 0.03c

1.970.04c 2.110.04c 2.030.03c

1.47 0.02c 1.85 0.04c 1.34 0.02c

RIF-Rifampicin (200 mg/kg), SY- Silymarin (100 mg.kg), IPT1- Ipomoea turpethum (200 mg/ kg), IPT2- pomoea turpethum (400 mg/kg), a = p < 0.001, when compared to normal animals, a = p < 0.001, when compared to control animals.

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The plasma half life of acetyl isoniazid (metabolite of INH) is shortened by RIF and is quickly converted to its active metabolites, which is related to the higher incidence of liver necrosis caused by INH and RMP in combination(13). There is no suitable drug for treating hepatotoxicity caused due to this combination. In regard of these herbs are implicated as potential hepatoprotective agents. Therefore, the present study attempts to study the hepatoprotective and antioxidant property of Ipomoea turpethum against anti-TB drug induced hepatotoxicity, in rats of wistar strain.Biochemical parameters of hepatotoxicity and oxidative stress were analyzed from serum and liver homogenates to assess the hepatoprotective activity. Further histopathological study was also carried out to confirm the pathological changes.The disturbance in the transport function of the hepatocytes as a result of hepatic injury causes the leakage of enzymes from cells due INH and RIF induced peroxidative damage or altered permeability of membrane (14) . Increased protein catabolism and urea formation that are seen in antitubercular drugs-induced hepatocellular damage and necrotic lesions in the hepatocytes may also be responsible for the increase of these amino tranferases activities in liver (15) (16). Similar increase in levels of SGPT and SGOT in the hepatic cells were also observed in the present study, when treated with combination of INH+RIF. In the present study, co-administration of hydro-alcoholic root extract of Ipomoea turpethum with INH+RIF significantly decreased the levels of these diagnostic marker enzymes (SGPT and SGOT) and the effect was observed to be dose dependent. Alkaline phosphatse was found to increase in the groupII animals treated with INH+RIF. ALP activity on endothelial cell surfaces is responsible for the conversion of adenosine nucleotides to adenosine, a potent vasodilator and anti-inflammatory mediator that results from injury. So, following injury, accumulation of interleukin-6 can lead to production of adenosine by alkaline phosphatase and subsequent protection from ischemic injury. This may be the reason for The increment in ALP in intoxicated rats due to liver cell necrosis(17). In the present study, co-administration of hydro-alcoholic root extract of Ipomoea turpethum with INH+RIF, decreased the levels of these ALP marker enzymes in the serum.Toxicity begins with the change in endoplasmic reticulum, which results in the loss of metabolic enzymes located in the intracellular structures (18). The toxic

metabolite hydrazine is produced, which further binds covalently to the macromolecule and causes peroxidative degradation of lipid membrane of the adipose tissue. In view of this, the reduction in levels of SGOT, SGPT and ALP caused by the Ipomoea turpethum is an indication of stabilization of plasma membrane as well as repair of hepatic tissue damage caused by INH+RIF. Similar mechanism for hepatoprotective action were proposed for Ginkgo biloba, Strychno spotatoru and Momordica dioica Roxb (19) (20) (21). In the present study, the levels of total cholesterol were higher in INH and RIF administered rats, indicating that the antitubercular drugs induce hypercholesterolemic condition. The increase in cholesterol levels in the liver might be due to increased uptake of LDL from the blood by the tissues(22). The abnormal cholesterol deposition is favored by the dangerous tendency of cholesterol to undergo passive exchange between the plasma lipoproteins and the cell membranes(23). Hence, protective HDL-cholesterol levels were reduced in the animals treated with INH+RIF. In the present study, co-administration with hydro-alcoholic root extract of Ipomoea turpethum reduced the elevation in the levels of total cholesterol induced by anti-TB drugs. The levels of protective HDL-cholesterol were also prominently increased when animals treated with Ipomoea turpethum,The probable mechanism responsible may be due to the decrease in the biosynthesis of cholesterol in the liver or by inhibiting enzymes responsible for the synthesis of cholesterol, by the chemical constituents of Ipomoea turpethum. This effect may also be responsible for an improvement in the serum HDL-levels.Tridox procumbens is also reported to have beneficial effect on lipid profile due to similar mechanism.Determination of serum bilirubin represents an index for the assessment of hepatic function and any abnormal increase in the levels of bilirubin in the serum indicate hepatobiliary disease and severe disturbance of hepatocellular function(24). The disaggregation of polyribosomal profiles induced by toxins is also associated with the inhibition of protein synthesis, which may be partially responsible for the fatty liver, probably not necrosis although it contributes to disabling of the cell(25). Hence, decrease in protein levels and increase in total bilirubin was observed in animals when treated with INH+RIF.In the present study, coadministration of hydro-alcoholic root extract of Ipomoea turpethum with INH+RIF increased the levels of these total proteins and decreased the total bilirubin levels in

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HISTOPATHOLOGICAL STUDIES

Normal

Control group

(INH+RIF+SY) Treated Group

(INH+RIF+SY) Treated Group

INH+RIF + IPT2 Treated. 16

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the serum.Cytochrome P450 is one of the liver enzymes, considered responsible for damage of hepatic cells (26). Ipomoea turpethum may inhibit these enzymes, thus enhance in the level of total proteins and decrease in the levels of total bilirubin. Thus the hepatoprotective action of Ipomoea turpethum may be mediated through the inhibition of UDP-sugar derivatives, enhancement of glycoprotein biosynthesis and stabilisation of cell membrane and inhibition of lipid accumulation by its hypolipidemic property, which are the few common mechanisms attributed for hepatoprotective activity for natural drugs (27). The results of histopathological

parameters also support the results of biochemical parameters and explain the hepatoprotective activity of Ipomoea turpethum. Isoniazid and rifampicin induced hepatitis is due to their biotransformation to reactive metabolites that are capable of binding to cellular macromolecules(28). The role of oxidative stress in the mechanism of isoniazid and rifampicin-induced hepatitis has been reported by Attri et al., (2000). Hence, effect of Ipomoea turpethum on oxidative stress induced by INH+RIF was studied. Lipid peroxidation is a common toxic phenomenon, regulated by the availability of substrate in the form of polyunsaturated fatty acids (PUFA).

Table-8: Effect of hydro-alcoholic root extract of Ipomoea turpethum on invivo antioxidant parameters
Group I II III IV V Treatment Normal Control INH+RIF + SY INH+RIF+IPT1 INH+RIF+IPT2 CAT (M/mg tissue) 8.20 0.27 2.66 0.32
*

GSH (M /mg tissue) 7.74 0.48 3.12 0.24

*

SOD (M /mgtissue) 25.50 1.30 12.92 0.43

*

LPO (M/mgtissue) 0.620 0.08 2.76 0.30* 1.27 0.18** 1.81 0.12 1.34 0.13**

5.43 0.32** 3.73 0.19 4.75 0.19**

6.42 0.21** 3.34 0.13 6.27 0.11**

21.76 0.69** 15.58 0.30 20.76 0.41**

RIF-Rifampicin (200 mg/kg), SY- Silymarin (100 mg.kg), IPT- Ipomoea turpethum (200 mg/kg), a = p < 0.001, when compared to normal animals, a = p < 0.001, when compared to control animals.

In the present study, free radicals formed either by the reaction of metabolites of INH+RIF with oxygen or by the interaction of superoxide radicals with H2O2, seem to initiate peroxidative degradation of membrane lipids and endoplasmic reticulum rich in polyunsaturated fatty acids. This leads to formation of lipid peroxides which in turn give products like MDA that cause loss of integrity of cell membrane and damage to hepatic tissue. Reduced glutathione is one of the most abundant non-enzymatic biological antioxidant present in the liver which scavenges reactive toxic metabolites of antitubercular drugs. Liver injury has been observed when GSH stores were markedly depleted(29). In our study, similar decrease in GSH was observed on administration of INH+RIF. It is known that SOD, CAT constitutes a mutually supportive team of antioxidant enzymes which provides a defense system against ROS. Catalase is an enzymatic

antioxidant, a heamoprotein which catalyses the reduction of hydrogen peroxide to water and oxygen and protects the tissue from highly reactive hydroxyl free radicals(30). In the present study, SOD and catalase decreased in INH+RIF treated animals may be due to an excessive formation of superoxide anions. Concomitant administration of hydro-alcoholic root extract of Ipomoea turpethum and INH+RIF effectively increased the GSH, SOD and CAT activities and also decreased the MDA levels which may be attributed to the scavenging of radicals by hydro-alcoholic root extract of Ipomoea turpethum resulting in protection of these enzymes. Triterpenes, present in the lupeol and betulin is known to quench the free radical by maintaing antioxidant levels(31). Thus, it can be suggested that significant

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antioxidant activity shown by Ipomoea turpethum may be due to presence of sitosterol, butulin and lupeol. Moreover, inhibitory activity of butulin and lupeol in free radical production could be related to the hepatoprotective effect. The present study indicates that the hydro alcoholic root extract of Ipomoea turpethum may be used as an effective hepatoprotective agent. Further studies on isolation and structural determination of active principles might be worthy. REFERENCES: 1. Thomas J Flynn, Martine S Ferguson. Multiend point mechanistic profiling of hepatotoxicants in HepG2/C3A human hepatoma cells and novel statistical approaches for development of a prediction model for acute hepatotoxicity. Toxicology in Vitro,22, 2008,1618-1631. 2. GowriShankar N L, Manavalan R, Venkappayya D, DavidRaj C. "Hepato protective and antioxidant effects of Commiphora berryi (Arn) Engl bark extract against CCl4-induced oxidative damage in rats". Food and Chemical Toxicology 46, 2008, 3182-3185. 3. Raul J Andrade, Mercedes Robles, Alejandra Fernandez Castaer, Susana Lopez Ortega Carmen Lopez Vega, Assessment of druginduced hepatotoxicity in clinical practice: A challenge for gastroenterologists, World J Gastroenterology ,13(3), 2007, 329-340. 4. James H Lewis, Drug induced liver disease. Advances in gastroenterology 84(5), 2000, 12751312. 5. Col AC An,VSM, Lt Col AK Seth, Lt Col M Paul, Lt Col P Puri. Risk factors of hepatotoxicity during anti-tuberculosistreatment.MJAFI,62,2000, 45 - 49. 6. Abhijit Chowdhury, Amal Santra, Koutilya Bhattacharjee, Subhadip Ghatak, Dhira Rani Saha, Gopal Krishna Dhali. "Mitochondrial oxidative stress and permeability transition in isoniazid and rifampicin induced liver injury in mice", Journal of Hepatology, 45,2006,117-126. 7. Pal R, Rana S V,Vaiphei K, Singh K, Is-oniazid rifampicin induced lipid changes in rats, Clinica Chimica Acta 389, 2008,55-60. 8. Kapoor L D Ayurvedic Medicinal plants, Delhi Publishers, 1st Edition, 2001, 222-223. 9. SureshKumar SV, Sujatha C, Syamala J, Naghasudha B, Mishra SH. "Protective effect of root extract of Operculina turpethum Linn.

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