Due to this problem of resistance against antibiotics, attention is now being shifted towards biologically active components isolated from plant species communing used as herbal medicine, as they may produce a new potent source of antibacterial and antifungal activities. 3 Therefore, it was vital to seek a natural drug that could overcome these disease conditions without causing any other adverse effects. 4 Annona squamosa Linn is a shrub that belongs to the family Annonaceae. 5 It is a perennial tree with thin, oblong-lanceolate, alternate leaves. It is abundantly native to tropical America, the Caribbean, and Indomalaya. 5 The fruits, leaves, bark, and root possess a wide variety of medicinal and nutritional value as it contains vitamin C, amino acid, thiamine, riboflavin, potassium, calcium, and dietary fibers. 5 A. squamosa exhibits several biological activities like hypoglycemic, 6 antimicrobial, 7 anticancer, 8 analgesic, 9 and antioxidant. 10 The leaf extract of A. squamosa L significantly reduce the average testicular-index, that happened due to the testicular weight shrinkage in animals. 11 The phytoconstituents like aporphine and annonaine alkaloids, flavonoids, and glycosides were identified from the leaves and bark exhibited many biological activities. 5 The antimicrobial activity of the methanolic leaf extract of A. squamosa was performed against Gram-positive and Gram-negative species like B. subtilis, S. aureus, P. aeruginosa, and E. coli, using the agar disc diffusion method. 12 However, the anti-microbial effect of n-hexane extract from the leaves of the plant is not performed yet. Hence the present study was designed to evaluate the antimicrobial activity and to investigate the compounds by GC-MS analysis and; further, an in-silico docking study to support its claim.