CHAPTER THREE
3.0 MATERIALS AND METHOD
3.1 MATERIALS Cannabis sativa plants were obtained with permission from the botanical gardens of Delta state university, Delta State. Other materials used include; wooden cages with iron netting, litter, (saw dust) animal feed and water, laboratory coat and gloves, dissecting kit and board, weighing balance, syringes, sample bottles, test tubes, 10% formal saline, spectrophotometer, pasteur pipette, micro pipette, cotton wool, chloroform timer, microscope, microtome, slide, cover slip, distilled water, embedding machine, slider warmer, tissue basket, wax bath, ethanol, paraffin block, all other equipment used were standard laboratory equipment.
3.2 RESEARCH POPULATION Twenty adult rats, Wistar breed weighing between 130g – 220g were procured from the Animal House of the Department of Physiology, Delta State University, Abraka. The wistar rats were housed in wire mesh cages under standard nutrition and environmental conditions to enable them acclimatize for two weeks before the start of the experiment.
3.3 RESEARCH DESIGN
3.3.1 GROUPING OF ANIMALS The wistar rats were divided into four (4) different groups, with five (5) wistar rats each, namely; • Group 1 (Control, which received no administration of cannabis sativa) • Group 2 (Coffee moderate dose (1200mg/kg)) • Group 3 (Low dose of cannabis sativa (900mg/kg)) • Group 4 (High dose of cannabis sativa 600mg/kg))
3.3.2 PREPARATION OF STOCK SOLUTION FOR Low dose (100mg/kg) 2000mg (2g) of cannabis extract was weighed with electronic weighing balance and dissolved with 200ml of 2% ethanol. This gave a concentration of 2000mg/200ml (10mg/ml). High dose (200mg/kg) 400mg (4g) of cannabis extract was weighed with electronic weighing balance and dissolved with200ml of 2% ethanol. This gave a concentration of 4000mg/200ml (20mg/ml).
3.4 ADMINISTRATION OF CANNABIS SATIVA Cannabis sativa was administered to experimental animals according to their body weight, such that animal weighing 200g, 150g, 170g received 2ml, 1.5ml and 1.7ml respectively. It was administered orally using oral canular
3.5 SAMPLE COLLECTION At the end of cannabis sativa administration, the wistar rats were anesthetized in a dessicator containing cotton wool soaked with chloroform. After they had attained deep anesthesia, they were brought out of the dessicator and a laparotomy was carried out (by making a V-shape incision in the abdominal region with the aid of a surgical scissors) and the visceral organs were exposed. Blood samples were collected via cardiac puncture
3.6 DETERMINATION OF BODY AND ORGAN WEIGHT The body weight of the experimental male wistar rats were initially taken after procurement, (before administration). The final weights of the wistar rats were determined before sacrificing and sample collection. Percentage weight gain was later calculated using the formula:
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