Abstract: Background and Objective: The utilization of bacteriocin-producing lactic acid bacteria as defending traces or bacteriocins in variety of compounds as bio-preservatives to manage undesirable microorganism remains a predominant core of recognition of researches. The aim of this study was the partial characterization and determination of the potential antimicrobial activity of lactic acid bacteria Enteroccocus feacium (E. feacium) isolated from traditional fermented tomatoes, studies the combination effect of foods ingredients on bacteriocin activity and use some agro-industrial byproduct for bacteriocin production. Materials and Methods: The bacteriocin-producing LAB was identified by 16sDNA, agar well diffusion assay and agar spot test were used to determine the antimicrobial spectrum of E. faecium against fifteen indicators microorganisms. The bacteriocin was characterized by study the effect of enzymes, heat, pH, UV light, storage time, EDTA, citric acid, ascorbic acid, acetic acid, NaCl and sucrose on their activity. The MRS broth, whey, tomatoes wastes and oranges wastes were tested for their suitability for bacteriocin activity using the optical density method. Data were analyzed by one-way ANOVA and the data were expressed as mean±standard deviation. Results: Strain was identified by 16SDNA as Enteroccocus feacium and producing a bacteriocin spectrum with Gram negative and Gram positive bacteria and fungus. Its antimicrobial compound was determined as a proteinaceous substance, stable to heat 120°C and pH 3.0-6.0, storage temperatures -20 to 15°C and in presence of non-ionizing artificial ultraviolet, its shows a synergic effect with sucrose and NaCl compared to EDTA, citric acid, acetic acid and ascorbic acid. Replace the synthetic De Man Rogosa Sharpe broth using agro-industrial wastes such as whey and tomato wastes can be promote bacteriocin production and activity. Conclusion: The characterization of bacteriocin revealed interesting properties that justifies its importance regarding food safety and may positively have an impact on its potential application in the biological control of pathogenic and spoilage microorganisms and hence food protection. The study showed evidence of the fact that the type of medium and medium components can influence bacteriocin activity, which may be taken into consideration when developing food products and modifying their consumption acceptability properties including texture and taste perception.