Introduction

Recently the use of medicinal plants to cure and treat various forms of diseases and dysfunction is becoming increasingly popular and has received wild acceptance (Grubben and Denton, 2004). Several hundreds of plants have been examined for use in a wide variety and they are believed to play role in preventing the development of chronic diseases due to their anti-oxidants effects (Grubben and Denton, 2004). Herbal medicine also called botanical medicine or phytomedicine refers to using a plant’s seed, roots, leaves, bark or flowers for medicinal purposes. Herbalism has a long traditional use outside of conventional medicine. It is becoming more main-stream as improvement in analysis and quantity control along their advances in clinical research shows the value of herbal medicine in the treating and preventing diseases (Dos Santos-Neto et al., 2006).

Cassia alata is a plant commonly known as Senna alata or Acapulco. It is an annual or biennial shrubs that grows up to 2.5 meter in height, leaves are compound, saripinnate, leaflets oblong, obtuse, membranous flowers seen in long terminal spike, yellow and showy. Fruit green angulated dehiscent pods, contain numerous compress brown coloured ovate seeds. Its major active ingredient includes yellow chrysophanic acid, kaempterol-3-gentiobioside and flavonoid (Moriyama et al., 2003; Hart et al., 2008). Cassia alata has been shown to be effective against inflammation, ringworm, cough, intestinal worms, poisons, constipation, hemorrhoids, bronchitis, asthma and sexual weakness (Mohideen et al., 2005; Moriyama et al., 2003).

Carbon tetrachloride (CCL₄) is a hepatotoxin. Trichloromethyl radicals are generated from it in vivo. These radicals stimulate a sequence of reactions that culminate in the initiation of the peroxidation of membrane lipids and hence liver damage (Reinke et al., 1988). CCL₄-induced lipid peroxidation process provides a model for which Cassia alata can be assessed for antioxidant property since previous reports has shown that it posses flavonoids (Moriyama et al., 2003). The ability of the extract to exhibit antioxidant action was therefore investigated by administering it to rats prior to CCL₄ treatment. The effect was compared with that in extract free CCL₄– treated rats.

Material and Methods

Collection of plant materialFresh leaves of Cassia alata were collected from Ughelli, in Ughelli South Local Government Area of Delta State and were identified and authenticated at the Herbarium Unit of Forestry Research Institute of Nigeria (FRIN) at Ibadan, Oyo State.

Extraction of plant material

The method of extract as described by Agbafor et al. (1999) was adopted. 1kg of fresh leaves of Cassia alata were sliced extracted by percolation at room temperature with 500ml of 70% ethanol for 48hours. The leaf extract was concentrated under reduced pressure (bath temperature 50ºC) and the extract evaporated to dryness.

Experimental Animals

Twenty four male Wister rats (220–250g) used for this study were purchased from the Animal Unit, College of Medicine, Delta State University, Abraka, Nigeria. They were divided into four experimental groups of six rats per group. Members of each group were housed in a standard rat cage (Griffin and George Modular cage system, Model YSM 580 cage base and YSM 600 – 540 cage top) and allowed to acclimatize to laboratory condition for one week. All rats were allowed free access to drinking water and fed ad libitum with chow (product of Edo Feeds and Flour Mill (EFFM), Ewu Edo state, Nigeria)

Treatments

The rats were randomized into four groups of six animals each. Group I served as the normal control, which received 0.5 ml corn oil once for 5 days. Animals in group 3 and 4 were pre-treated with Cassia alata extract at doses of 2.5 ml/kg and 5.0 ml/kg body weight respectively for five days.  Following the last extract treatment, CCL₄ was dissolved in corn oil and given i.p to rats in group 2 and those in groups 3 and 4 once (on day 5) at a dose of 0.5 ml CCL₄/kg body weight in not more than 0.5 ml of corn as described by Obi et al., (1998).

Preparation of Serum Homogenate

The animals were fasted overnight and sacrificed on day 6. The blood was collected by heart puncture into tubes without anticoagulant to separate serum for various biochemical estimations. The serum was separated by centrifugation at 3000rpm for 10mins. The obtained supernatants were stored at 4°C until required for assay.

Biochemical Assay

Aspartate aminotransferase (AST) and alanine aminotransferase activities were quantified spectrophotometrically using the methods of Reitman et al (1957) as described by Schmidt and Schmidt, (1963). The reagents for the assays were already prepared and packaged in commercial kits supplied by Randox Laboratories, United Kingdom.

Statistical Analysis

The data were presented as mean ± SD. The mean values of the various treatment groups were compared using ANOVA with Dunnett’s post test GraphPad Prism version 4.00 for Windows. The significant level was set at P < 0.05.

Results and Discussion

In this study, serum ALT and AST activities were used to measure both CCL₄– induced hepatotoxicity and protection of Cassia alata extract against the effect of CCL₄ in rats. In agreement with the report of previous workers (Reinke et al., 1988), this results show that CCL₄ caused an elevation in the serum content of ALT and AST which is indicative of damage to the liver and other organ of the body (Reinke et al., 1988, Obi et al., 1998). Treatment of the rats with 2.5ml of Cassia alata / kg body weight for 5 days before 0.5ml/kg CCL₄ caused progressively reduced hepatotoxicity than with CCL₄ alone. This was further reduced at a higher dose of 5ml of Cassia alata / kg body weight (Table1).

Various mechanisms have been proposed for CCL₄-induced damage (Brattin et al., 1985) and one of them is that trichloromethyl radical produced from carbon tetrachloride by reductive dechlorination abstracts a hydrogen atom from a fatty acid to form chloroform and a lipid radical. The lipid radical may then react with molecular oxygen to initiate lipid peroxidation which is thought to ultimately cause the cytotoxic responds. There is a wide variety of naturally occurring compound that can protect the liver and other tissues from damage. Among these are antioxidants such as carotenes, tocopherol, catechols, flavonoids etc (Aruoma, 1994).

Although the biochemical mechanism of the protective effect of Cassia alata against CCL₄-induced damage was not examined in this current study, it is possible that in Wistar rats the extract may have acted directly or indirectly in protection the liver against damage. In addition, it may be an effective scavenger of the reactive metabolite, trichloromethyl radical (^*CCL₃).

Table 1: Effect of Cassia alata (C.a) extract on serum aspartate and alanine aminotransferase activity.

n= 5, values are expressed as mean ± SD. Values bearing the same superscript in a column do not differ significantly (p > 0.05) compared to control

*% change with respect to control group I

**% change with respect to CCL₄ treated group alone II

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