BACKGROUND Caylusea absyssinica, a plant used as vegetable and for medicinal purposes was selected for in vitro antibacterial evaluation in this study. The main aim of this study was to isolate compounds from the plant roots and evaluate their antibacterial activities on clinical bacterial test strains. METHODS Compounds from roots of Caylusea absyssinica (fresen) were identified based on observed spectral (1H-NMR, 13C-NMR and IR) data and physical properties (melting point) as well as reported literature. Disk diffusion method was employed to evaluate the antibacterial activities of the isolated compounds on four test bacterial strains namely, Staphylococcus aureus (ATCC25903), Escherichia coli (ATCC25722), Pseudomonas aeruginosa (DSMZ1117) and Salmonella thyphimurium (ATCC13311). RESULTS Two compounds, CA1 and CA2 were isolated from the methanol crude extract of the roots of Caylusea absyssinica (fresen). The compounds were identified as β-sitosterol and stigmasterol, respectively. Evaluation of antibacterial activities revealed that the compounds are active against all the bacterial strains in the experiment, showing inhibition zones ranging from 12 mm-15 mm by CA1 and 11 mm-18 mm by CA2 against the different test strains. However, the compounds were less active than the reference drug (Gentamycine), which showed minimum inhibition zone of 21 mm (Pseudomonas aeruginosa) and maximum of 28 mm (Escherichia coli) inhibition zone. CONCLUSIONS The isolation of the compounds is the first report from roots of Caylusea abyssinica and could be potential candidates for future antibacterial drug development programs.