Selections in Du Puits alfalfa (Medicago sativa L. ) were evaluated for saponin content. The data obtained from the Trichoderma bioassay were converted to saponin values by regressions and standard curves. It was necessary to adjust the concentration of alfalfa extract in the bioassay so that the saponin content in alfalfa was estimated in that portion of the curve showing a marked response to saponin. A different regression was established for each group of low and high saponin plants of Du Puits alfalfa. The standard curve prepared with extract of Du Puits alfalfa used on the basis of its saponin content or the commercial purified saponin could be utilized for estimating the saponin content in Du Puits alfalfa, because the results were comparable to the values determined by chemical analysis. Attention should be given to the concentration of extract used when converting the bioassays to actual saponin content, because certain concentrations gave values more similar to the chemical analysis than others. Two cycles of selection for low and high saponin content in Du Puits alfalfa were involved in the tests. The results indicated that the saponin content in low saponin plants was lower and that the saponin content in the high saponin plants was higher than in the unselected plants. They also showed that average saponin values in low and high saponin and the unselected plants were similar between the second and the third cuttings, but different from the first cutting