The present study was carried out to screen the anti-quorum sensing property of the possible bioactive fractions and valuable secondary metabolites from the hexane extract of bark and leaves of Acacia ferruginea. Solvent extraction of the plant material using hexane followed by preparative thin layer separation and centrifugal thin layer separation of the extract using silica gel yields 12 and 5 fractions, respectively. Spectral characterization (UV, FT-IR) and anti-QS screening were done for the fractions. GC-MS was done for both nonpolar extracts and cyclograph fractions. The active moieties from bark and leaves fractions were established on the basis of UV, FT-IR and GC-MS. From the anti-QS screening studies, the active fractions were determined. A total of 17 fractions were obtained from the hexane extract of bark and leaves of Acacia ferruginea. Among these fractions, 5 fractions were shown to exhibit activity against Chromobacterium violaceum, while 2 fractions showed moderate activity.