Bridelia micrantha commonly known as coastal golden leaf is a member of the family Phyllanthaceae. In preliminary studies, nine fractions, named F1- F9, were obtained by fractionating the crude methanol extract of the stem bark of Bridelia micrantha using column chromatographic techniques. The F6 fraction was found to be the most active when tested for the antibacterial activity. This study is thus aimed at investigating the effect of fractionation on antibacterial activity of F6 fraction. The F6 fraction was fractionated by adsorption chromatography on silica gel into eight sub-fractions designated F’1- F’8. A product was isolated from the dichloromethane/ methanol (10%) fraction and the structure was determined on the basis of spectroscopic data. The antibacterial activity of the F6 fraction, sub-fractions and the product was evaluated by broth microdilution method against two reference strains and eighteen clinical bacterial strains. The chemical analysis of F6 and three sub-fractions F’3, F’4 and F’5 was done using HPLC-MS. The fraction F6exhibited strong activity on all the tested bacteria with MIC values of 128 µg/ml on nine strains, including Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, and Salmonella sub-fractions F’4 and F’5 exhibited the best activities on all the tested bacteria with MIC values of 32 to 256 µg/ml. The chemical analysis by HPLC-MS of F6, F’3, F’4 and F’5 revealed the presence of almost 180 identified compounds from various classes of secondary metabolites including alkaloids, flavonoids, steroids and terpenoids. The product obtained although inactive was elucidated as Daucosterol. | Chemical composition and antibacterial activity of fractions from Bridelia micrantha stem bark Methanol extract