The antiprotozoal effect of saponins varies according to both the structure of the sapogenin and the composition and linkage of the sugar moieties to the sapogenin. The effect of saponins on protozoa has been considered to be transient as it was thought that when saponins were deglycosilated to sapogenins in the rumen they became inactive; however, no studies have yet evaluated the antiprotozoal effect of sapogenins compared to their related saponins. The aims of this study were to evaluate the antiprotozoal effect of eighteen commercially available triterpenoid and steroid saponins and sapogenins in vitro, to investigate the effect of variations in the sugar moiety of related saponins and to compare different sapogenins bearing identical sugar moieties. Our results show that antiprotozoal activity is not an inherent feature of all saponins and that small variations in the structure of a compound can have a significant influence on their biological activity. Some sapogenins (20(S)-protopanaxatriol, asiatic acid and madecassic acid) inhibited protozoa activity to a greater extent than their corresponding saponins (Re and Rh1 and asiaticoside and madecassoside), thus the original hypothesis that the transient nature of the antiprotozoal action of saponins is due to the deglycosilation of saponins needs to be revisited.