Protective effect of Pseudarthria viscida (L.) against Paracetamol induced hepatotoxicity

Adil Nafar A., Rince Rajan and Jose Padikkala*

Amala Cancer Research Centre, Amala Nagar, Thrissur-555, Kerala

*Corresponding Author E-mail: jpadikkala@rediffmail.com

ABSTRACT:

In this study we have evaluated the hepatoprotective activity of methanolic extract of Dasamoola plant Pseudarthria viscida L. (PVM) in rats toxicated with high dose of paracetamol. Animals were treated with two concentrations of PVM (200 mg/kg and 400 mg/kg) for seven consecutive days. After 24 hrs of paracetamol treatment animals were sacrificed; liver tissue and blood samples were collected. The level of protection was evaluated by measuring the serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), alkaline phosphatase (ALP) and bilirubin levels and superoxide dismutase (SOD), glutathione peroxidase (GPX), reduced glutathione (GSH) and malondialdehyde (MDA) levels in liver tissue homogenates. Levels of serum GOT, GPT, ALP and bilirubin were got reduced in PVM treated animals. PVM treatment was also found to be effective in plummeting paracetamol induced oxidative stress.

KEYWORDS: Pseudarthria viscida, paracetamol, hepatoprotection, liver enzymes.

INTRODUCTION:

Liver has essential role in detoxifying harmful substances entering the circulatory system. It has significant role in metabolism of fats, carbohydrates and proteins. Any variation in hepatic structure or function may result in portal hypertension, jaundice, and increased bleeding which may lead to metabolic changes that affect other organs too¹. Even though several drugs are available to treat liver dysfunctions, growing awareness over their side effects is changing the attention of modern science towards developing safer drugs from natural products. Pseudarthria viscida (L.) Wight and Arnis of Fabaceae family, is an important plant in the Ayurvedic systems of medicine. Its root is used as a nerve tonic and is also known for its anti-inflammatory, rejuvenating and cardio protective properties². Its root is reported to have found with both antioxidant and antiulcer properties³. PVM also exhibits anti-hypertensive⁴, antifungal⁵, antidiarrhoeal⁶ as well as wound healing⁷activities. The current study was designed to evaluate protective effect of P. viscida on paracetamol induced hepatotoxicity in Wistar rats.

MATERIALS AND METHODS:

Animals

Male Wistar Rats (220-240 g) were purchased from the Small Animal Breeding Station, Agricultural University, Mannuthy, Kerala. The animals were maintained under standardized environmental conditions (22-28^oC, 60-70% relative humidity, 12 hr dark/light cycle) and fed with standard rat feed (Sai Durga Feeds, Bangalore, India) and water ad libitum. All the animal experiments conducted in the present study had prior permission from Institutional Animal Ethics Committee (IAEC) and followed its guidelines.

Plant Material

P. viscida plants were collected from Peechi forest, Thrissur, Kerala and were identified and authenticated by Dr. N. Sasidharan, Taxonomist, NWFP – Division, Kerala Forest Research Institute, Peechi, Thrissur, Kerala. A voucher specimen was deposited in the Kerala Forest Research Institute herbarium for further reference.

Preparation of extract

Collected P. viscida plants were thoroughly washed, shade dried and made into fine powder. Extract preparation with 70% methanol was done using soxhlet apparatus. The extract was filtered and evaporated to dryness. The dried extract was re-dissolved in distilled water and used for further studies.

Table 1: Effect of Pseudartharia viscidia on serum SGOT, SGPT, ALP and Bilirubin against Paracetmol induce toxicity.

GROUP	SGOT(IU/L)	SGPT (IU/L)	ALP(IU/L)	BILIRUBIN(mg/dL)
NORMAL	173.71±10.1	189.52±8.55	202±11.59	0.28±0.04
CONTROL (Paracetmol)	536±14.09^a	432.21±11.73^a	422±17.131^a	0.56±0.003^a
PVM (200mg/kg)	242.5±14.5^c	378±5.09^c	313±24.22^c	0.48±0.1^c
PVM(400mg/kg)	225.6±17.23^c	265±8.02^c	247±20.97^c	0.37±0.04^c

Values are Mean ± SD for six animals in each group; (a) p<0.01, (b) p < 0.05 as compared to normal; (c) p<0.01, (d) p < 0.05 as compared to control

Values are Mean ±SD for six animals in each group; (a) p<0.01, (b) p < 0.05 as compared to normal; (c) p<0.01, (d) p < 0.05 as compared to control

Fig.1 Effect of PVM on liver SOD activity in paracetamol treated rats

Values are Mean ±SD for six animals in each group; (a) p<0.01, (b) p < 0.05 as compared to normal; (c) p<0.01, (d) p < 0.05 as compared to control.

Fig.2 Effect of PVM on liver GPx activity in paracetamol treated rats

Hepatoprotective study

Rats were divided into four groups with six in each group.

Group I: Normal without any treatment

Group II: Paracetamol + water (control group)

Group III: Paracetamol + PVM (200 mg/kg body weight)

Group IV: Paracetamol + PVM (400 mg/kg body weight)

Oral treatment of PVM was started seven days prior to paracetamol (2g/kg) administration. After 24 hrs of paracetamol administration all the animals were sacrificed. Blood was collected and serum separated was used to evaluate the activities of serum glutamate pyruvate transaminase (SGPT), serum glutamate oxaloacetate transaminase (SGOT), alkaline phosphatase (ALP), bilirubin and total proteins following standard procedures by using commercially available kits (Span Diagnostic Ltd).The 25 % liver homogenate in tris buffer (pH 7) was subjected to further analysis to evlauate the effect of PVM in protecting liver tissues from oxidative stress. Reduced glutathione (GSH) was determined by the method described by Moron et al. (1979)⁸. Glutathione peroxidase (GPx) was measured by the method described by Hafeman et al. (1974)⁹. Estimation of Superoxide dismutase (SOD) was done by the method of Mc Cord and Fridovich (1969)¹⁰. Lipid peroxidation was measured as n mols MDA/mg protein, following the method by Okhawa et al (1979)¹¹.

Statistical analysis

The values were presented as mean ± SD. Statistical evaluation of the data was done by one way ANOVA followed by Dunnet ‘t’ test. Results were considered statistically significant when p<0.05.

RESULTS:

The effect of PVM on serum GOT, GPT, ALP and Bilirubin in paracetamol toxicated animals in comparison to control group is summarized in Table.1. Serum GOT, GPT, ALP and Bilirubin levels were significantly lower (p< 0.01) in PVM treated groups even after getting exposed to higher dose of paracetamol. To the contrary Control group has higher level of Serum GOT, GPT, ALP and Bilirubin in comparison to Normal group (p< 0.01).

Figure. 1-4 represent the level of SOD, GPx, GSH and MDA levels in liver tissue homogenate of animals in different groups. Group IV showed improvement in SOD level (p< 0.05) when compared to Group II. Where as after administration of single dose of paracetamol there was a significant decrease in GPx and GSH levels also. This trend was reversed in Group III as well as group IV (p< 0.01). Although paracetamol increased the lipid peroxidation level PVM treatment had significantly decreased this effect (p< 0.01).

DISCUSSION:

Paracetamol, a widely used antipyretic agent, is mainly metabolized in liver. Its higher dosages could cause severe hepatic necrosis in experimental animals and humans¹². Electrophilic metabolite of paracetamol, N-acetylparabenzoquinonimine, conjugates with cellular glutathione (GSH) causing sharp deterioration in GSH level ¹³. This destruction further drastically increases the liver enzymes in serum. The liver enzyme level in serum becomes quantitative markers of the extent and type of hepatocellular damage¹⁴. Higher levels of serum GOT, GPT, ALP and Bilirubin (p< 0.01) in Paracetamol toxicated Group II showed that greater destruction of hepatocytes could have occurred in comparison to normal group (Group I) which has lesser levels of SGOT, SGPT, ALP and Bilirubin in their serum. PVM treated groups showed improvement in their SGOT, SGPT, ALP and Bilirubin levels (p< 0.01). Even though, it is evident that PVM is able to decrease the level of hepatocyte destruction in paracetamol treated groups (Group III and Group IV) further studies are required to explain the nature of the mechanism of action.

GSH a thiol-based antioxidant plays an important role in the cellular defense cascade against oxidative damage. It protects living system from heavy assault of reactive oxygen species¹⁵. GPx with the support of its cofactor GSH, catalyzes the reduction of hydrogen peroxide to water and oxygen, and thereby limit the possibility of formation of hydroxyl radical, a highly toxic reactive oxygen species ¹⁶. Deficiency of GSH leads to oxidant damage and increased level of lipid peroxidation which in turn leads to cell damage ¹⁷. SOD also play significant role by scavenging the dangerous superoxide free radicals¹⁸. Administration of paracetamol to Wistar rats increased the level of depletion of GSH in control group (p< 0.01). Similarly there was a significant level of decrease in SOD and GPx and a drastic increase in tissue MDA level (p< 0.01). Conversely the PVM treated animals showed improvement in their innate antioxidant system and a recovery of significant levels in GSH, SOD and GPx (p< 0.01).

CONCLUSION:

The current study suggest that PVM treatment shows improvement of innate anti-oxidant system which is evident from the higher level of SOD, GSH and GPx and a drastic decrease in MDA level. The decrease in serum SGOT, SGPT, ALP and Bilirubin level in PVM treated animals indicate that the plant extract is able to reduce the level of hepatocyte destruction caused by heavy dose of paracetamol.

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Received on 19.04.2012 Modified on 02.05.2012

Research J. Pharm. and Tech. 5(7): July 2012; Page 946-949