Phytochemical investigation of Cassia italica Mill. (Fabaceae) aerial parts collected from Saudi Arabia gave nine compounds: β-sitosterol (1), (22E)-3--hydroxycycloart-22-en-24-one (2), uvaol (3), daucosterol (4), methyl 3,4-dihydroxybenzoate (5), emodin (6), 4-hydroxyphenyl-O-β-D-glucopyranoside (7), aloin B (8), and rutin (9). Their structures were established by extensive spectroscopic study, in addition to co-chromatography with authentic samples. The total MeOH extract (TME), different fractions, and compounds 1-9 were assessed for their antimicrobial effects using agar plate diffusion assay. The TME, EtOAc fraction, and compounds 5-9 showed significant antimicrobial activity against the tested microbial strains.