Polyvinylalcohol (PVA) is a hydrophilous and semi-crystallized. It has attracted muchrelevancy due to its pretty chemical endurance, fine thermal determination,decent physical specialities, wonderful bio-compatibility and cheapness.Gelatin (G) is a natural polymer and that are interesting materials forbiomedical applications. Electrospinning is a simple method that provides veryporous nanofiber production with high surface area. It is possible to producebiomedical, filtration, energy storage and protective materials by usingelectrospinning method. Benzalkonium chloride (BAC) is a kind of anti-microbialcationic surface-active agent, which has been pretty used in merchant wounddressings and has a powerful status toward Gr+ bacteria. In this study, nanofibers wereproduced from electrospinning of BAC, PVA and G containing solutions at variousconcentrations. The fibers of obtained nanofiber structure were uniform,continuous and intensive. The optimum parameters in terms of good mechanicaland antibacterial properties were determined. S2 sample, electrospun from 11.63% PVA + 1.0% G +1.0% BACcontaining solution, had the best morphological and mechanical properties dueto having the thinnest fiber diameter (51±13nm) and the highest verticalstrength (4.299MPa) and horizontal strength (4.058MPa). It also had antibacterialactivity against all the bacteria tested (E.coli,P.aeruginosa, B.subtilis and S.aureus). Due to owninggood mechanical and antibacterial properties, S2 sample can have many uses inmedical sector.