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BACKGROUND This study focused on the valorization of prickly pear (PP) fruit (Opuntia ficus‐indica) into vinegar by spontaneous surface fermentation on different starting matrices (with/without the addition of sucrose and with/without PP peel in the raw material). Different parameters were monitored during the fermentation process in terms of their physicochemical and biological properties. RESULTS Physicochemical and phytochemical analysis revealed significant differences depending on the starting matrix. An increase in total phenolic content (TPC) was observed for the majority of samples when transformed from PP juice into PP vinegar revealing the role of fermentation in enhancing the bioactive compounds content. Better antioxidant and antibacterial activity were detected for vinegar samples compared with the initial starting matrix. Using whole PP fruit resulted in better TPC and antioxidant activity; in contrast, sugar addition had no significant effect on any studied data. Analysis of variance, taking into account the four factors that were studied (matrix, variety, with/without peel, and with/without sugar), demonstrated that only the factor ‘presence or absence of the peel’ had a significant influence on the TPC values. CONCLUSION This study demonstrated that both whole PP fruit and PP juice could be used as new raw materials for vinegar production. © 2023 Society of Chemical Industry.