Alternaria sesami is a major pathogen causing Alternaria leaf spot, prevalent in all the sesame growing areas of the world. Fungicidal effect of Mikania scandens plant extract was evaluated against Alternaria leaf spot disease in Sesamum orientale. Methodology includes hydrodistillation of M. scandens using clevenger apparatus followed by GC-MS, GC-FID spectrometer analysis, changes in the activity of phenylalanine ammonia lyase (PAL), peroxidase (POX), total phenolic content, and isozyme pattern of superoxide dismutase in sesame. Aqueous M. scandens leaf extract showed remarkable inhibition of mycelial growth. Treatment of healthy S.orientale with M. scandens aqueous extract followed by fungal inoculation induced significantly high phenylalanine ammonia lyase activity; the initiator enzyme of secondary metabolic pathway followed by a substantial increase in cytosolic and membrane-bound peroxidase activity. The isozyme profile of superoxide dismutase showed no significant changes. Phenolic compounds also showed an increased profile. Bioassays were conducted under field conditions for two successive growing seasons (2011 and 2012) at the experimental farm which resulted in a sufficient decrease in both disease incidence and disease severity compared with the control. Thus, application of M. scandens aqueous extract to sesame initiated a series of biochemical changes in the plants which are considered to be a part of the plant defense response. Further studies designed to assess the role of the extract in the molecular signal induction of resistance against pathogens are in progress.