Pleurotus ostreatus has potent antibacterial, antifungal, and antioxidant properties. In this study, bioactive compounds were extracted from Pleurotus ostreatus and screened against bacterial and fungal phytopathogens. Antibacterial activity was determined against four plant pathogenic bacterial species, viz. Erwinia carotovora, Pseudomonas syringae, Ralstonia solanacearum, and Xanthomonas axonopodis In antibacterial experiment, extract of n-hexane showed maximum activity as compared to other organic extracts. The zones of inhibition caused by n-hexane and ethyl acetate extracts on Xanthomonas axonopodis were 88.55 mm and 99.67 mm, respectively. n-hexane extract of Pleurotus ostreatus also showed the highest antifungal activity. As compared to controls, 25 mg/ml caused 76, 82, 82, 83, and 60% decreases in fungal biomass against, Alternaria alternata, Aspergillus flavus, Drechslera australiensis, Fusarium oxysporum and Macrophomina phaseolina, respectively. The most susceptible fungus observed in the present investigation was Fusarium oxysporum, although extracted fractions inhibited other test fungi as well. Gas Chromatography Mass Spectrometry (GC-MS) analysis was performed on n-hexane extract depicting the presence of 26 compounds. Out of these compounds, one compound identified as Toluene (Molecular weight= 92) exhibited maximum peak area as 91 % followed by another significant compound named as Aziridine 2-methyl-3-(1-methylethyl)- (Mol. Weight = 99) was identified only one time and show maximum hit of 84% showing peak area 56%. During this investigation, it was determined that the bioactivities observed were due to these compounds, which exploited that, Pleurotus ostreatus could be a potent biocontrol antagonist against the plant pathogens.