Biofilm forming pathogens are among the major causes of hospital-acquired infections and are not much affected by antibiotic treatment. Consequently, novel agents and therapeutics are required urgently that possess antibacterial and antibiofilm activities. This study analyzed two bacteriocins from Lactobacillus plantarum subsp. argentoratensis SJ33 strain for their antibacterial and antibiofilm activity as well as cytotoxic properties. BacF1 and BacF2 showed broad spectrum activity against both Gram-positive (Listeria monocytogenes, Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa, Escherichia coli) bacteria. Significant bactericidal action was also observed on S. aureus cells by pore formation. Additionally, bacteriocins disrupted biofilms formed by S. aureus and P. aeruginosa which were shown by crystal violet staining assay and visualized by fluorescence as well as scanning electron microscopy. Quantitative Real-Time PCR study revealed changes in gene expression of biofilm formation in S. aureus (ica) and P. aeruginosa (pelA, psl, rhlA). Cytotoxicity of bacteriocins was further analyzed on normal mammalian cells and Caenorhabditis elegans. Notably, bacteriocins showed no major effect on HEK-293 cell line and enhanced the survival of S. aureus infected HEK-293 cells. Similarly, no cytotoxic effect was visible on C. elegans even after treatment with higher concentration than MIC at different time intervals.