High-performance liquid chromatography is the most powerful and the most frequently used technique for saponin determination due to the fact that it can deal effectively with non-volatile, highly polar compounds. It has been used extensively for determination of both aglycones and intact saponins. The separations are performed usually on normal (silica gel) and reversed-phase (C8, C18) columns, of which C18 has been definitely preferred. The detection at lower wavelengths, however, limits the selection of solvents and the gradients that can be used. Since acetonitrile gives much lower absorption at lower wavelengths than methanol, the selection of acetonitrile–water gradients is the mode of choice.