Preliminary acute toxicity levels of crude extracts of
It is indeed true that
The smallholder farmers in the South Western Agro-ecological Zone of Uganda (SWAEZ) use
Despite the plant and its compounds being used by the farmers, the acute toxicities of
2. Materials and methods
2.1. Plant seed collection and preparation
2.2. Acute toxicity study protocol
The ethical committee of Mbarara University of Science and Technology (MUST) approved the use of experimental animals for the experiments.
Swiss albino mice of 16-25 gr bodyweight from both sexes were selected and labelled using tail markers of different colours. The animals (n=30) were divided into five groups of six mice each and kept in different cages for easy observation.
The dose levels were determined after a preliminary acute toxicity trial, which had been carried out earlier. The doses rates were as shown in table 1.
The volumes of the drug doses administered to the mice were calculated using the following formula : Volume (ml) = body weight (kg) x dose levels (mg/kg)/stock drug concentration (mg/ml). The extracts were administered by gavage orally.
The derived dose levels for ethanolic extracts were 3,500, 5000, 6500 and 8,000 mg/kg and for aqueous extracts were 10,000, 12,000, 14,000 and 16,000 mg/ml.
Distilled water was used for extraction to prevent contamination of the extract. Both the aqueous and ethanolic extracts of
The Swiss albino mice of the same age were used for the study to minimize variation in the test results. The control group given 1 ml of distilled water was used for comparison with the groups given the plant extract.
3.1. Aqueous acute toxicity profile of
The aqueous acute toxicity profile of
It was observed that it would take 12043 mg/kg to kill 50% of the test animals (Graph 1). This gives a wide safety margin for the lethal dose of
Probit for aqueous
3.2. Toxicity signs observed with dose increase within the groups
There were notable side effects associated to higher doses of
3.3. Acute toxicity of 70% ethanolic extract of
The ethanolic acute toxicity profile of
3.4. Signs of acute toxicity observed
For the ethanolic extracts at a dosage of 3500mg/kg and 5000mg/kg, the following signs were observed: irritation shown by the use of forelimbs to scratch the areas of the mouth, gasping for air and dyspnoea. For aqueous extracts at a dosage of 6500mg/kg and 8000mg/kg there were observed twitching of GIT muscles, urination, dry mouth, reddening of lips and convulsions.
3.5. Comparison of key histo-pathological findings due to aqueous and ethanolic extracts
The key histopathological findings due to the effects of aqueous and 70% ethanolic extracts of
Probit for 70% ethanolic
3.6. Histopathology findings
|A −Brain (Control). The scale bar is 100µm||B −No significant effects on the brain at 10,000mg/kg. The scale bar is 100µm||C −Slight congestion of the meninges at 3500mg/kg. The scale bar is 100µm|
|D −Kidney (Control) Medulla. The scale bar is 100µm||E −Kidney renal haemorrhages, congestion and tissue degeneration at 12,000mg/kg. The scale bar is 100µm||F −Haemorrhages and congestion at 5,00mg/kg. The scale bar is 100µm|
|G −Lungs (Control). The scale bar is 100µm||H −Mild Lung emphysema noted at 10,000mg/kg. The scale bar is 100µm||I −Lungs haemorrhages and congestion at 5,000mg/kg. The scale bar is 100µm|
|J −Small intestine (Control). The scale bar is 100µm||K −Erosions of the small intestine at 14,000mg/kg. The scale bar is 100µm||L −Intestines villus sloughing off and mucosal erosion at 6,500mg/kg. The scale bar is 100µm|
|M −Liver (Control). The scale bar is 100µm||N −Hepatic degeneration and congestion of the liver at 14,000mg/kg. The scale bar is 100µm||O −Liver wide necrosis and tissue degeneration at 6,500mg/kg. The scale bar is 100µm|
|P −Heart (Control). The scale bar is 100µm||Q −Myocardial haemorrhages at 12,000mg/kg. The scale bar is 100µm||R −Wide spread haemorrhages on the myocardium at 5,000mg/kg. The scale bar is 100µm|
4.1. Lethal dose (LD50) levels
The lethal dose (LD50) of aqueous extracts of
Local smallholder farmers are using aqueous extracts of
The Organization for Economic Cooperation Development (OECD) Guidelines for the Testing of Chemicals  recommended that the maximum dose levels for any chemical compounds should not exceed 5000 mg/kg of the animal body weight. The ethanolic extraction of Capsicum annum has advantages compared to the aqueous because ethanol is better solvent for extraction of bioactive compounds.
The extracts derived from ethanolic solvents have lower LD50 of 5492mg/kg because ethanol is less polar. The ethanolic extracts have yet another advantage: the longer shelf life for use because of the preservative effects of alcohol on the extracts when compared to that of the water in aqueous extracts . It can be noted here that extracts, which seem to show high toxicity in many circumstances, have no traditional uses, which is in agreement with previous findings [1,8].
After administration of the higher extract dosages there were often observed notable clinical signs in test animals, due to the side effects of the extracts. Key clinical signs noted were irritation, evidenced by the use of fore limbs to scratch the areas of the mouth, gasping for air, dyspnoea, twitching of GIT muscle, urination, dry mouth, reddening of lips and convulsions. The effects of bioavailability of the drugs on major organs like the intestines, liver, lungs, kidneys and brain showed clear manifestations of the presence of the active substances. Bioavailability is the rate and extent to which the active drug ingredient is absorbed from a drug product and becomes available at the site of drug action. The effects of the active substances in the body of the mice could affect by the rate of disintegration of the drug product, dissolution of the drug in the fluids at the absorption site or the transfer of drug molecule across the membrane lining the gastrointestinal tract into the systemic circulation.
The key physiologic factors that could affect the availability of the active substances in the body include: variations in absorption power along GI tract, variations in pH of GI fluids, gastric emptying rate, intestinal motility, perfusion of GI tract, presystemic and first-pass metabolism, age, sex, weight and disease states. The interactions with other substances like food, fluid volume and other drugs or chemicals could also play an important role.
There exist some reports of seeds side effects by
The histopathological findings showed significant effects on brain, kidney, lungs, small intestines, liver and heart. Studies in India indicated that necropsy examination is paramount in linking the general and target organ specific toxic effects of phytomedicine . Many others are in agreement OECD [10-12]. Absence of any significant gross pathological lesions in treated rats and mice at the terminal sacrifice indicates the justifiable harmless nature of the phytomedicine .
Results of study indicated that the lethal dose (LD50) of aqueous extracts of
Conflict of interest
The authors declare here that there is no conflict of interest.
We profoundly thank the Belgian Technical Cooperation (BTC) for funding this research work. The vital roles of Dr.Vudriko Patrick, Mr. James Ndukui in laboratory services and coordination are appreciated. Dr.Nanyingi Mark and Dr. Sarah Nalule are recognized for their cordial advice and peer review.