Bioassays, using guppies as a test animal, were conducted onTephrosia leaf extracts of seven plants obtained by four different extraction methods. The total rotenoids recovered by acetone extraction of fresh leaves appeared to have slightly more effective piscicidal properties than rotenoids recovered by acetone extraction of oven-dried leaves. These differences, although not statistically significant, proved the validity of using fresh leaf extraction for the evaluation of rotenoid content inTephrosia vogelii leaves. Individual plants varied significantly with regard to their toxicological effectiveness, thereby demonstrating that the proportion of rotenone vs. other rotenoids varies amongTephrosia plants. ATephrosia breeding program cannot rely solely on the red-color test as a measure of rotenone equivalents but must be supplemented with periodic bioassays to assess the pesticidal effectiveness of the breeding material.