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This study was intended to isolate and investigate endophytic fungi and its metabolite as an active biocontrol agent. Endophytic fungal secondary metabolites are considered as an effective and environmentally friendly alternative to synthetic chemicals for integrated pest management. In this study, different parts of medicinal plant Plectranthus amboinicus (Oregano) were selected to isolate twenty-three fungal endophytes. Out of twenty-three, one isolate demonstrates potent activities against cotton leaf worm Spodoptera litura (S. litura) an insecticides resistance pest. Identification of potent isolate was attained through the morphological, sequence similarity and phylogenetic analysis of the ITS regions and confirmed as Aspergillus sojae (A. sojae). The GCMS analysis reveals the metabolites of A. sojae were rich in 2-Furancarboxaldehyde and Levoglucosenone then other compounds. The FTIR peaks of A. sojae crude extract shows the major peaks that corresponding to NH, C–H, and OH stretching and minor peaks are C–H bend (meta) and acetylenic C–H bend. FTIR analysis supports the presence of functional groups in accordance with GCMS data as dominating compound. The bioassay results showed that A. sojae metabolite affects third and fourth instar larvae of S. litura. In addition, antibiotic quality of A. sojae metabolite tested against five selective bacteria. Among five different bacteria, the maximum zone of inhibition was displayed for Escherichia coli and Staphylococcus aureus in the higher concentration of 100 μg/mL. Our resultssuggest the importance of endophytic fungal metabolites to control the pesticide-resistant insect pests.