This study investigated the effects of gaseous ozone treatments at different concentrations and exposure times on the physicochemical and microbiological quality of beef (Musculus longissimus lumborum) during cold storage. Beef loin samples were divided into control and treatment groups, with ozone applied at 0.5-0.6 ppm for 10 or 20 minutes. Quality parameters, including pH, water activity, color (L*, a*, b*), lipid oxidation (TBARs), and microbial counts, were measured immediately after treatment and on days 7 and 14 of storage. Ozone treatment significantly reduced microbial loads, particularly mesophilic aerobic bacteria, Pseudomonas spp., and Enterobacterales, and maintained microbial counts at acceptable levels throughout storage. Although ozone exposure increased lipid oxidation, pH and water activity values remained within normal ranges, and color changes were moderate. These findings highlight ozone as a promising non-thermal preservation technology that can improve microbial safety and extend the shelf life of beef without leaving chemical residues.