Ozonated olive oils have emerged as a promising alternative for natural antimicrobial agents in the food industry due to their potential to inhibit microbial growth. However, the stability and effectiveness of these oils under different storage conditions has not been thoroughly explored. This study examines the changes in the physicochemical properties and antimicrobial activity of ozonated virgin olive oil (VOO) and pomace olive oil (POO), stored at 4 °C and 20 °C for 6 months. The peroxide index (PI), acidity index (AI), iodine value (IV), and viscosity (V) were analyzed, along with their antimicrobial activity against Escherichia coli (STCC 45), Pseudomonas aeruginosa (STCC109), and Staphylococcus aureus (STCC 239). The results showed that both oils underwent changes in their physicochemical properties and antimicrobial activity over time. The PI initially increased up to day 30, with VOO reaching a peak value of 741.44 ± 32.16 meq O₂/kg and POO reaching 1067.23 ± 56.56 meq O₂/kg, but after this point, it began to decrease in both oils and at both temperatures (4 °C and 20 °C). The acidity index (AI) increased over time, particularly in POO, which reached a final value of 6.32 ± 0.14 mg KOH/g. Both oils showed a reduction in iodine value (IV), and an increase in viscosity (V) over time. In terms of antimicrobial activity, P. aeruginosa remained stable with an average inhibition zone of 9.41 ± 0.23 mm, while E. coli showed the greatest increase in activity over time, reaching 21.31 ± 4.01 mm in POO at 20 °C. On the other hand, S. aureus exhibited the highest average antimicrobial activity, with a mean inhibition diameter of 14.49 ± 0.36 mm, and the largest inhibition zone of Ø = 18.97 ± 1.46 mm observed after 180 days of storage. A Spearman correlation analysis revealed a strong positive relationship (ρ > 0.85, p < 0.05) among PI, AI, and the antimicrobial activity with storage duration. This study provides novel insights into the stability of ozonated oils, offering valuable perspectives for their application in the food industry, especially using pomace olive oil, a key by-product in olive oil production.

Ozonated vegetable oils are increasingly recognized as bioactive agents with both antimicrobial and functional properties, attracting growing interest for their potential applications in food systems. This review critically synthesizes current knowledge on their chemical transformations, physicochemical properties, antimicrobial and functional properties, their toxicological and safety considerations and the implications of their regulatory classification based on 84 studies published between 2005 and 2026. Evidence shows that antimicrobial performance increases with oxidation level, but significant variability in ozonation conditions and analytical methods limits comparability and hinders standardization. Despite encouraging in vitro results and early applications in food matrices and packaging systems, major gaps remain regarding toxicology, sensory quality, regulatory classification, and real-world functionality. Overall, ozonated vegetable oils hold notable potential as antimicrobial and functional tools, yet further research is required to ensure their safe and practical implementation in food systems.