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Hepatoprotective effect of hydroethanolic extract of Hybanthus enneaspermus against Paracetamol induced liver damage in rats.

Introduction

Liver disease is a world wide problem. Conventional drugs used in the treatment of liver disease are sometimes inadequate and can have serious adverse effects. It is therefore, necessary to search for alternative drugs for the treatment of liver diseases to replace currently used drugs of doubtful efficacy and safety (1). Liver has a pivotal role in regulation of physiological process. It is involved in several vital functions such as metabolism, secretion and storage. Liver diseases are mainly caused by toxic chemicals (Certain antibiotics, chemotherapeutics, peroxidised oil, aflatoxins, CCl4, chlorinated hydrocarbons etc), excess consumption of alcohol, infections and autoimmune disorder. Most of the hepatotoxic chemicals damage liver cells by inducing lipid peroxidation and other oxidative damages in liver (2-5)

Hybanthus enneaspermus (L) F Muell (Violaceae) known as Lakshmisheshta, Padmavathi in Sanskrit, is an important plant in the Indian system of medicine. It grows 15-30 cm in height with many diffuse or ascending branches and is pubescent in nature. The plant is reported in ancient ayurvedic literature to cure conditions of "kapha and pitta", Urinary calculi,strangury, painful dysentery, vomiting, burning sensation, wandering of mind, urethral discharge, blood troubles, asthma, epilepsy, cough and to give tone to breast(6)

Scientific Classification:
Kingdom Plantae
Division Magnoliophyta
Order Malpighiales
Family Violaceae
Genus Hybanthus
Species H.enneaspermus


Materials and Methods

Plant Material

The whole plant of Hybanthus enneaspermus were collected from the paddy fields of Thanjavur during the month of July. The plant samples were identified and authenticated by Dr.G.V.S.Murthy, Scientist and Head, Botanical survey of India ( BSI/SRC/5/932), Coimbatore.

Animals

Male Wistar albino rats weighing between 150 - 220 gm were used for the study. The animals were obtained from animal house, IRT Perundurai Medical College, Erode, Tamilnadu, India. On arrival the animals were placed at random and allocated to treatment groups in polypropylene cages with paddy husk as bedding. Animals were housed at a temperature of 24[+ or -]2[degrees]C and relative humidity of 30 - 70 %. A 12:12 light: day cycle was followed. All animals were allowed to free access to water and fed with standard commercial pelleted rat chaw (M/s. Hindustan Lever Ltd, Mumbai). All the experimental procedures and protocols used in this study were reviewed by the Institutional Animal Ethics Committee (Regd no: 688/2/C-CPCSEA/ Proposal No: IAEC/NCP/01/2009-2010) and were in accordance with the guidelines of the CPCSEA.

Preparation of the extract

The coarse powdered plant was extracted with 70% Etoh by using soxhlet apparatus. The solvent were removed under reduced pressure to get semisolid mass and stored in refrigeration. The freeze dried material was weighed, dissolved in water and used for the study.

Phytochemical analysis

The extract was subjected to preliminary phytochemical analysis to identify the chemical constituents (7).

Acute Toxicity Studies

Acute toxicity studies were performed using hydroethanolic extract of Hybanthus enneaspermus according to OECD-423 guidelines.

Experimental procedure

The animals were divided in to 5 groups of 5 animals each. Group-I, which served as normal control, received normal saline (1ml/kg., p.o) for 7 days, Group - II served as hepatotoxicant control received saline for 7 days. Group - III, IV and V received silymarin (100mg/kg., p.o.), 200 and 400mg/kg of hydroethanolic extract of Hybanthus enneaspermus plant once daily for 7 days. On the fifth day, after the administration of the respective treatments, all the animals of groups II, III, IV and V were administered with paracetamol 2g/kg orally.

On 7th day, after 2h of respective treatments the blood was collected by sinus puncture under light ether anesthesia and serum was separated for various biochemical estimations. The activities of serum hepatic marker enzymes namely SGOT, SGPT,and Alkaline phosphatase (ALP) , and Total Bilirubin were assayed in serum using standard kits

Statistical analysis

The values were expressed as mean [+ or -] SEM. The statistical analysis was carried out by one way analysis of variance (ANOVA) values P<0.05 were considered significant.

Results and Discussion

Phytochemical analysis of the extract revealed the presence of alkaloids, flavonoids, saponins, triterpenes, cardiac glycosides and proteins. All the doses (5,50,300 and 200 mg/kg) of the hydroethanolic extract of Hybanthus enneaspermus employed for acute toxicity studies were found to be non toxic and it did not produce any mortality even at the highest dose( 2000mg/kg). The results of the hydroethanolic extract of Hybanthus enneaspermus on paracetamol treated rats are shown in Table I. The levels of hepatic marker enzymes SGOT, SGPT, ALP,and Total Bilirubin was elevated in paracetamol treated animals when compared to control groups. The hydroethanolic extract of Hybanthus enneaspermus and silymarin treatment decreases the levels SGOT,SGPT, ALP and Total Bilirubin when compared to Paracetamol treated rats.

The hydroethanolic extract of Hybanthus enneaspermus was investigated for its hepatoprotective effect. Various mechanisms may be associated with the liver damage. Paracetamol is eliminated mainly as sulfates and glucuronide. Only a small amount (5%) is metabolized via the cytochrome P450 enzyme system to the alkylating metabolite N-acetyl-p- benzoquinone imine (NAPQI) which is responsible for the toxic effect (8).The advancement in modern medicine notwithstanding, there are no synthetic drugs for the management of many liver disorders. In the absence of a reliable and effective hepatoprotective agent in modern medicine, a number of medicinal plant preparations have been recommended for the treatment of liver problems (9). In this study hydroethanolic extract of Hybanthus enneaspermus demonstrated significant liver protection against the hepatic injuries caused by the hepatotoxin.

It is evident that several phytoconstituents have the ability to induce microsomal enzymes either by accelerating the excretion of hepatotoxin or by inhibition of lipid peroxidation induced by it (10).A good number of naturally occurring compounds have been shown to protect liver and other organ from damage(11) .Phytoconstituents like flavonoids(12), triterpenes(13), saponins(14) and alkaloids (15) are known to possess hepatoprotective activities. Recently, total flavonoids were reported to protect animals from liver injury and liver damage (16,17). From this study it is concluded that the hydroethanolic extract of Hybanthus enneaspermus plant protects liver damage and could be used as an effective protector against Paracetamol induced hepatic damage.

Abbreviations
SGOT Serum glutamic oxaloacetic transaminase;
SGPT Serum glutamic pyruvic transaminase;
ALP Alkaline phospatase;
HEHE Hydroethanolic extract of Hybanthus enneaspermus


References

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V. Premalakshmi (1) and C. Deepika Thenmozhi (2)

(1) Asst. Professor, Department of Horticulture, Agricultural College and Research Institute, Madurai 625104, TamilNadu, India

E-mail: premavelu@rediffmail.com

(2) Research Scholar, Department of Biotechnology, Mother Teresa Womens University, Kodaikanal, TamilNadu, India.

E-mail: biotechmtwu@gmail.com
Table I: Effect of Hydroethanolic extract of Hybanthus enneaspermus
(HEHE) on biochemical parameters in paracetamol-induced hepatic
injury in rats.

Treatment SGOT U/I SGPT U/I

Control (Saline 1ml 65.67[+ or -]0.32 135.81[+ or -]0.35
p.o., Seven days)

Positive control
(Saline 1 ml p.o., 385.05[+ or -]0.45 408.07[+ or -]1.15
seven days) +PCM

Silymarin (100mg/kg 70.85[+ or -]0.61 * 139.89[+ or -]0.75 *
p.o.seven days) +
PCM

HEHE (200mg/kg p.o. 108.05[+ or -]0.94 * 178.04[+ or -]1.18 *
seven days) +PCM

HEHE (400mg/kg 89.08[+ or -]0.78 * 145.78[+ or -]0.45 *
p.o.seven days) +PCM

Treatment ALP U/I Total
 Bilirubin
 mg/dl

Control (Saline 1ml 127.57[+ or -]1.03 0.85[+ or -]0.01
p.o., Seven days)

Positive control
(Saline 1 ml p.o., 479.08[+ or -]1.57 3.97[+ or -]0.05
seven days) +PCM

Silymarin (100mg/kg 131.89[+ or -]0.70 * 1.81[+ or -]0.01 *
p.o.seven days) +
PCM

HEHE (200mg/kg p.o. 167.65[+ or -]0.83 * 2.87[+ or -]0.45 *
seven days) +PCM

HEHE (400mg/kg 145.04[+ or -]1.08 * 2.03[+ or -]0.04 *
p.o.seven days) +PCM

Values are in mean [+ or -] S.E.M (n = 5) * P < 0.01 vs PCM group.
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Author:Premalakshmi, V.; Thenmozhi, C. Deepika
Publication:International Journal of Biotechnology & Biochemistry
Date:May 1, 2011
Words:1742
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