The aim of this study was to characterize lactic acid bacteria (LAB) isolated from cocoa mucilage and beef and evaluate their inhibitory effect in vitro against pathogenic bacteria, as well as determine their effect on beef quality. For the antagonist assay, 11 strains of LAB were selected and tested against pathogenic strains of Escherichia coli and Salmonella sp. The pathogenic bacteria were cultured in a medium, and a previously reactivated LAB bacterial pellet was added. After incubation, halos were observed around the bacterial colonies of the pathogenic strains, indicating inhibition by the LAB. It was identified that the LAB strains used belonged to the genus Lactobacillus, and the CCN-5 strain showed high percentages of inhibition against Salmonella sp. (58.33%) and E. coli (59%). The effectiveness of LAB application methods (immersion, injection, and spraying) did not present statistical differences. Furthermore, no significant changes in the physicochemical characteristics of beef were observed after the application of LAB. The results obtained demonstrate the potential of cocoa mucilage, as a biological control agent through LAB application, for beef biopreservation due to its ability to inhibit the growth of pathogenic bacteria.