Abstract: Cheese production with mixed adjunct cultures of two or more microorganisms has gained significant attention due to their advantages over single-strain adjunct cultures. The aim of the current study was to assess the effects of mixed probiotic adjunct cultures on the physicochemical indicators, textural properties, free amino acid contents, short-chain fatty acid profile, and microbial load of Cheddar cheese during 90 days of ripening. Cheddar cheese was prepared with adjunct cultures of Pediococcus acidilactici AS185 and/or Kluyveromyces marxianus JM2, and Cheddar cheese made with a commercial cheese starter served as a control. The results showed that the addition of the mixed adjunct culture did not influence the chemical composition of Cheddar cheese. However, the pH of the experimental cheese was found to be higher than that of the control. Notably, after 90 days of ripening, the counts of P. acidilactici AS185 and K. marxianus JM2 in the mixed adjunct culture group were higher than those in the single adjunct culture group. The textural parameters of the cheese were not significantly affected by the adjunct cultures during the ripening process. The mixed adjunct culture resulted in an increase in the levels of certain individual amino acids as well as an elevation in the total content of free amino acids. Cheese with the addition of P. acidilactici AS185 and K. marxianus JM2 as adjunct cultures contained higher levels of short-chain fatty acids, especially acetic acid and butyric acid. These findings provide a basis for the application of mixed adjunct cultures in cheese production, which helps enrich the flavor and improve the texture of cheese.

Key words: Pediococcus acidilactici; Kluyveromyces marxianus; probiotic cheese; Cheddar cheese; adjunct starter