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The dichloromethane extracts of the root and leaf of Tephrosia vogelii Hook F. (Family: Papilionaceae) grown in Kenya, were subjected to antimicrobial assays against Staphylococcus aureus, Escherichia coli and Fusarium phoseolida . The minimum inhibition concentrations (MIC) ranged between 0.25 to 6.4 μg/ml for all the microorganisms tested. The crude root and leaf extracts were also toxic to the brine shrimps, organisms used as indicators of toxicity (LC50: 0.960; 0.958 μg/ml, respectively). Chromatographic separation led to the identification of (6as, 12as, 5's)-6a, 12a, 4',5'-tetra hydro-2,3-dimethoxy 5'-s- isopropenyl–furano (3',2': 8,9)-6H-rotoxen-12-one, (rotenone) in 9.75% yield, (roots), and 11.75% (leaves). Characterisation was achieved by use of spectroscopic techniques and comparison with rotenone standard and literature data. Novel HPLC conditions for purification of rotenoids were established. The MIC of rotenone against the tested microbes was found to be 5.2 and 1.0 μg/ml for Staphylococcus aureus and Escherichia col i respectively, while the LC50 against the brine shrimps was found to be 3.20 μg/ml. The findings indicate that the extracts of T. vogelii contain metabolites that have antimicrobial activity comparable to other medicinal plants. Keywords : Tephrosia vogelii , Rotenone, Staphylococcus aureus, Escherichiacoli, Fusarium phoseolida Journal of Agriculture, Science and Technology Vol. 8 (1) 2006: pp. 1-14