A total of 54 natural origin compounds were evaluated for their activity in inhibiting the proliferation of glioma cells. Results showed that four Aesculus polyhydroxylated triterpenoid saponins (3-6), six Gleditsia triterpenoid saponins (7-12), and five phenolic compounds (43-46, 51) had dose-dependent activity suppressing the proliferation of both C6 and U251 cells. Structure-activity relationship analysis suggested that the acetyl group at C-28 for the Aesculus saponins and the monoterpenic acid moiety for the Gleditsia saponins could be critical for the activity of these active compounds. Aesculioside H (4), gleditsioside A (7), and feuric acid 3,4-dihydroxyphenethyl ester (FADPE, 46) were the three most active compounds from the different types of the active compounds and induced apoptosis and necrosis in glioma cells.