Recurrent phenotypic selection for low‐ and high‐saponin concentration in alfalfa (Medicago sativa L.) was practiced in each of six alfalfa varieties for up to five generations. In the last cycle of selection the low series did not differ significantly from one variety to another, and the saponin index averaged 0.34%. In the last cycle of the high series the saponin index averaged 3.14% and ranged from 1.12% for ‘Lahontan’ to 4.72% for ‘Uinta.‘ The saponin concentrations of leaves and stems were correlated (P < 0.01). Concentrations of saponin in first and second crops were also correlated (P < 0.01). There was a low correlation between saponinconcentration index and leaf percentage (P < 0.05). With the exception of high‐saponin Uinta, selection for saponin‐concentration did not significantly change the leaf percentage from that of the original variety. Changes in leaf percentage accounted for only a small part of the change in saponin concentrations. Yield and protein concentration were not affected by low‐ or high‐saponin selection.Evidently a shift occurred towards an increase of saponins that contain carboxylic acid‐type aglycones in the high‐saponin selection and towards saponins that do not contain carboxylic acid‐type aglycones in the low‐saponin selections. This conclusion was supported by 1) chromatographic separations, 2) bioassays of butanol extracts, and 3) bioassays with water extracts with added cholesterol. Apparently more toxic fractions of saponins were increased by selection for high saponin and decreased by selection for low saponin.