| Abstract | We recently screened more than 900 plant extracts for anthelmintic potential using an egg-hatching inhibition procedure. Dried plant materials were ground to a particle size of no larger than 1 mm and a portion of 30 g was then transferred to an Erlenmeyer flask and extracted with 250 mL of methanol. The mixture was shaken for 1 h and filtered to derive the crude plant extract. The latter was suspended in 50 mL of Tetrahydrofuran (THF), stirred for 15 min and filtered. The THF solution was evaporated and the concentrated extract was dried by using a Savant SpeedVac Vaccum Concentrator System. The dried extract was weighed and dissolved in dimethyl sulfoxide (DMSO) before use. With a sucrose step gradient procedure, worm eggs were harvested from fresh feces of goats naturally infected with parasitic nematodes in which more than 90% were Haemonchus contortus. The eggs in phosphate buffered saline (PBS) were distributed into 96-well plates at 30-40 eggs/well in 98 μL. Two μL of plant extracts in DMSO was added to each well in duplicate and mixed. The final concentration of extracts in the wells was 400 μg/mL and untreated eggs in PBS plus 2 μL DMSO served as negative control. The test plates were kept in an incubator at 27°C for 48 h. Hatched larvae (dead or alive) and unhatched eggs (embryonated and non-embryonated) were then counted under a microscope at a 40 × magnification. The percentage of inhibition of egg hatching was calculated as: inhibition (%) = number of egg / (number of egg + number of larvae) × 100. We found that 16 extracts showed inhibitive effects (> 50%) at the concentration of 400 μg/mL. The extract of black pepper (Piper Nigrum L.) was particularly effective and examined further. The egg-hatching inhibition rate for the black pepper extract was 5, 11, 26, 70, 78, 86, 97, 96, and 98% at concentrations of 0, 3.9, 15.6, 31.3, 62.5, 125, 250, 500, and 1000 µg/mL, respectively. The results suggest that more work is needed to evaluate the anthelmintic efficacies of solvent extracts of black pepper and other plants. |
