It is estimated that approximately one quarter of processed drugs contain plant extracts or active ingredients obtained from or modeled on plant substances (Tripathi and Tripathi, 2003, Samy et al., 2007). In recent times there is an increasing awareness and interest in medicinal plants and their preparations commonly known as herbal medicines (Steve et al., 2009). The major hindrance to the use of traditional herbal preparations is the lack of scientific and clinical data in support of better understanding of the efficacy and safety of the drugs (Sathya et al., 2009)
Sclerocarya birrea (A. Rich) Hochst., subspecies Caffra (family:Anacardiaceae) is an important food, commercial, cultural and ethnomedicinal plant in Africa (Ojewole et al., 2010). The tree is commonly found in semi-arid deciduous and savanna regions of sub-Saharan Africa (Borochov-Neori et al., 2008). It grows in wooded grasslands, riverine woodland areas and bush lands and is frequently associated with rocky hills. Its geographical distribution stretches from Gambia in West Africa across Nigeria and Cameroon in Central Africa, to Ethiopia and Sudan in East Africa (Ojewole et al., 2010). In Nigeria, the plant is commonly known as danya (Hausa), kemaa (Kanuri) and edere (fula-fulfulde). It is also known as maroola-plum (English) or marula (South Africa) (GRIN, 2009). The stem-bark, roots and leaves have been reported to possess medicinal and other properties in addition to the nutritional values of the fruit and seeds of the plant (Ojewole, 2004). Sclerocarya birrea is widely used in traditional medicine in Africa against hypertension, stomach or gastro-enteritis, cough and antihyperglycemia (Dimo et al., 2007). It has also been reported to have a large number of therapeutic properties and pharmacological activities (Ojewole et al., 2010). These include antifungal activities Runyoro et al. (2006), antihelminthic activity (McGaw et al., 2007), antiplasmodial activity (Gathirwa et al.. 2008), anti-inflammatory activity (Ojewole 2003a), hypoglycemic activity (Dimo et al., 2007), anticonvulsant activity (Ojewole, 2006a), anti-hypertensive activity (Ojewole, 2006b), anti-bacterial activity(Eloff, 2001) and antioxidant activity (Masoko et al., 2008). Even though S. birrea is an important source of nutrients and phytocompounds that play a role in protecting against a variety of ailments, there is no record in the medical literatures on the toxicity study of the acetone extract of its stem bark. The present study was therefore carried out to evaluate the safety of acetone extract of S. birrea by acute and sub-acute toxicity studies in Wistar rats.
2.0 Materials and methods
2.1 Collection and identification of plant material: Pieces of fresh stem bark of S. birrea were harvested from the Federal Polytechnic Staff Quarters, Bauchi, Bauchi State, Nigeria. This was identified and authenticated at the Department of Biological Sciences, Ahmadu Bello University, Zaria Herbarium and was found to have a voucher specimen number of 1071.
2.2 Preparation of plant extract: The fresh stem-bark of S. birrea was air dried, minced and powdered using laboratory mortar. 1 kg of stem back powder was extracted in 2.5 litres of acetone (JDH) using a Soxhlet extractor. This was filtered using a Whatman’s filter paper (24 cm). The filtrate was evaporated using a hot water bath and a total yield of 141.7 g was obtained.
2.3 Experimental animals: Thirty adult Wistar rats of both sexes (average weight of 120 g per group) were obtained from the Department of Human Anatomy, Ahmadu Bello University, Zaria, Kaduna State, Nigeria. They were kept in plastic cages and maintained under laboratory conditions of temperature and light with free access to food (standard pellet diet, Grand Cereal Ltd, Jos Pleateau State) and water. The experimental animals were acclimatized for two weeks after which they were divided into six groups of five animals each. The experimental protocol was in accordance with the OECD guidelines and care of the experimental animals was done according to the Ahmadu Bello University, Zaria, Nigeria guidelines for laboratory animal care and handling.
2.4 Acute toxicity study: Acute toxicity studies of acetone extract of S. birrea stem bark was carried out in Wistar rats using Organization for Economic Co-operation and Development (OECD) guideline 425 (OECD 425, 2008). In this study, all rats received a single oral dose of the test sample. Rats in Group I received 1 ml/kg of distilled water while doses of 2000 mg/kg and 5000 mg/kg of the extract were given using oral gavage to rats of Group II and Group III respectively. All the rats were observed for general behavioral changes, symptoms of toxicity and mortality after treatment for the first four (critical) hours, then over a period of 24 hours, thereafter daily for 14 days.
2.5 Sub-Acute Toxicity Studies: Sub-acute toxicity study (28-day repeated oral toxicity study) was carried out according to OECD 407 guidelines (OECD 407, 2008). Rats were divided into three groups of 5 animals each. The Group I received 1ml/kg distilled water and served as a control group whereas Group II and Group III 500 mg/kg and 1000 mg/kg of extract respectively orally via gavage. All the groups of rats were observed twice daily for mortality and morbidity till the completion of the experiment. All the animals were observed for clinical signs and the time of onset, duration of these symptoms, if any were recorded. Body weights of the rats in all groups were recorded once before the start of dosing, once weekly during the treatment period and finally on the day of sacrifice. At the end of the experiment (on 29th day), experimental animals were humanely killed under chloroform anaesthesia and blood collected via cardiac puncture into heparinized and non-heparinized tubes for hematological analysis and biochemical analysis.
2.6 Haematological parameters: The heparinized blood was used for the analysis of hematological parameters such as hemoglobin, packed cell volume (PCV) and white blood cell count were measured using an automated hematology analyzer (PE 6000).
2.7 Biochemical Parameters: The serum was separated from non-heparinized blood and the serum biochemical parameters including total cholesterol, creatinine, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), blood urea nitrogen (BUN), triglycerides, total cholesterol, albumin, bilirubin and total protein were analyzed using a digital colorimeter AC-14.
2.8 Histopathology: The liver and kidney were collected from all the animals for histopathological study. The collected organs were weighed and preserved in 10 % neutral buffered formalin, trimmed and a 5 ? thickness of tissue sections were stained with hematoxylin and eosin for histopathological study.