Two hundred and seven extracts were prepared from sixty plants from South Africa and screened for <i>in vitro</i> activity against <i>Trypanosoma brucei rhodesiense, Trypanosoma cruzi, Leishmania donovani</i>, and <i>Plasmodium falciparum</i>. For the 21 extracts which inhibited the growth of one or more parasites with more than 95 % at 10 µg/mL, the IC₅₀ values against all four protozoal parasites and cytotoxic IC₅₀ values against L6 myoblasts were determined. Amongst the most notable results are the activities of <i>Psoralea pinnata</i> (IC₅₀ of 0.15 µg/mL), <i>Schkuhria pinnata</i> (2.04 µg/mL), and <i>Vernonia mespilifolia</i> (1.01 µg/mL) against <i>Trypansoma brucei rhodesiense.</i> HPLC-based activity profiling was used to identify the active constituents in the extracts, and the germacranolide sesquiterpene lactones schkuhrin I and II from <i>S. pinnata</i>, and cynaropicrin from <i>V. mespilifolia</i> were identified, with IC₅₀ values of 0.9, 1.5, and 0.23 µM, respectively.