**2.5.3 Tail-flick test**

This test as first described [28], and subsequently modified (Janssen et al, 1969; Asongalem et al 2004a) was used. Briefly, before treatment, the terminal (2 cm) of each rat tail was immersed in hot water contained in a 500ml beaker and maintained at 55 ± 1oC using a thermo-regulated hot plate (Ugo Basile, Socrel DS-35) and the time (in seconds) between the onset of stimulation and tail withdrawal was measured as the tail-flick latency. Twenty five rats that shows response within 0 – 4 s were selected and grouped into five (n = 5) for the study. Immediately after basal latency assessment, normal saline, reference drug (Pethidine hydrochloride; Bayer, England) or the plant extract were administered and the reaction time again recorded at 30 and 60min. after administration the extract. (50 – 200 mg/kg p.o) (Sanchez-Mateo et al, 2006).

Herbal Medicine in the Treatment of Malaria:

(MUST).

**3. Results** 

sun drying,

with appropriate Human Kits.

appropriate kit (Coulter ACT 5 diff Diluent's).

Men dominate the practices of traditional medicine.

*Vernonia amygdalina*: An Overview of Evidence and Pharmacology 173

Food and water intake were measured daily while the animal's body weights were taken preexposure and weekly during exposure. All animals were observed at least once daily for clinical signs (behavior such as lethargy, hyperactivity, depression and diarrhea), and once/week clinical observations were performed on each rat by removing it from its cage and examining it for changes in general health. On day 44, immediately prior to euthanasia, all animals were anesthetized with chloroform and bled via the descending aorta for hematology and clinical chemistry determination. Organs were dissected and weighed to determine absolute and relative weight. The blood for clinical chemistry was allowed to clot in microtainer separator tubes, centrifuged and sera collected and stored at -70 0C until performing the biochemical analyses. The biochemical and hematological parameter were analyzed in the (New Italian Laboratory). Mbarara University of Science and Technology

The collected plasma samples were analyzed using a (HumanStar 180 and Humalyze 2000, Germany) autoanalyzer. Sixteen biochemical parameters were studied; plasma sodium, potassium and chloride were also assayed with (Humalyzer 17410, Germany) autoanalyzer

Hematological parameters were analyzed using (Beckman and Coulter, USA) with the

In the study, a total of 90 healers from - sub-county from the three districts were interviewed. Most of them were members of traditional healers association in their district.

Most of the traditional healers interviewed (81 out of 90), indicated to have passed through a several routes to become healers. All of the 81 were raised in families with traditional healers, who had involved them in the healing process and they were familiar with the profession. After some time they started to practice on their own. All reported on the importance of the family environment of a traditional healer in the context of acquiring knowledge and experience by members of the family. Furthermore it was reported that, the entrance into practice through these routes is facilitated through training by an experienced healer or a family member, who decides when the apprentice is ready to become an independent healer or to take up the practice. Eight out of 81 of the traditional healers indicated to have been instructed through dreams by their ancestral spirits to take up the traditional healing practice. They were required to learn and observe traditional healing procedures as dictated by the spirits. Indeed, the ancestral spirits are considered to be supernaturally powerful and ignoring them is to invite punishment to an individual and or

her family. Of the 90 healers interviewed, two indicated to be self-taught healers.

The leaves are the most frequently used plant part (56.3%), the root and fruits are used about 30% and 8.5% respectively, and the less used plant part is the bark (5.3%). The majority of the remedies are prepared in the form of decoction of fresh leaves. In our study area people do usually not store remedies for prolonged period of time. When needed they go out and collect the plant and prepare the remedy from fresh or sun dried material. Powders are prepared by pounding the fresh plant part or the crushed plant material after

Decoction is the most frequent method way of remedy preparation (65%) followed by infusion (13%), which is used for the powders; the maceration (11%) is mostly used for the

## **2.6 Acute and sub-chronic toxicity of the herbs 2.6.1 Acute – toxicity test**

The intraperitoneal (i.p) acute toxicity of the extract was evaluated in Swiss albino mice using a slightly modified Lorke's method (Lorke, 1998). Briefly, this method involved the determination of LD50 value in biphasic manner. The animals were starved of feed but allowed access to water 24 h prior to the study. In the initial investigatory step (phase 1), a range of doses of the extract producing the toxic effects was established. This was done by intraperitoneal administration of geometric doses of the extract (10, 100, 1000, 1500 mg/kg) to four groups of mice (n = 4). Based on the results obtained, a phase 2 investigatory step was done by giving more specific doses (200, 400, 600, 800 mg/kg i.p) to four other groups of mice.

The mice were observed for 24 h for such behavioral signs as, excitement, dullness, ataxia or death. The LD50 was estimated from the geometric mean of the dose that caused 100% mortality and the dose which caused no lethality.

The same procedure was used in rats which received (1000, 2000, 3000, 5000 mg/kg oral) in phase 1 and (1500, 3000, 4000, 5000 mg/kg oral) in the second phase.
