This study evaluated the antifungal and antibacterial activities of the extracts and their fractions, essential oils, and major constituents of Piper arboreum tissues. The chemical compositions of P. arborem tissues and major constituent were identified using GC/MS, 1 H and 13 C NMR. All the botanic material tested proved to be specific bactericidal agents against Staphylococcus aureus, with MBC/MIC = 2, including the amide piperetine, the major compound isolated of P. arboreum for the first time. The essential oil of leaves showed antifungal activity against Microsporum gypseum and Epidermophyton floccosum, with MFC values of 312.5 and 125 µg/mL, respectively.