From Preparation to Bioactivity: A Comparative Study on Preparation Methods and Characterization of Postbiotics.

Postbiotics are bioactive compounds derived from bacterial metabolism that offer various health benefits in food applications. This study aimed to compare three methods for preparing postbiotics: cell-free supernatant (CFS), thermal treatment, and enzymatic treatment with ultrasonication by using lactic acid bacteria (LAB): F2, AF1, and 50. The focus was on assessing cell viability and antimicrobial activity, followed by characterizing the CFS through additional analyses of antioxidant activity, total phenolic content (TPC), and the presence of phenolic and flavonoid compounds. The results demonstrated that both the CFS and thermal treatment methods effectively removed viable bacterial cells, whereas enzymatic treatment combined with ultrasonication exhibited lower efficacy. Antimicrobial activity varied based on both the bacterial strain and the preparation method. Specifically, F2 and AF1 exhibited higher antimicrobial activity when using the CFS method, whereas 50 showed greater efficacy after complete thermal treatment. Additionally, F2 demonstrated the highest antioxidant capacity and TPC, measuring 76.11 ± 6.21 mg/L and 125 ± 6.6 mg GAE/mL, respectively. This was correlated with elevated lactic acid production (17.2 ± 0.1 mg/mL) and acetic acid production (6.25 ± 0.07 mg/mL). 50 had the highest concentration of citric acid (1.47 ± 0.04 mg/mL), while AF1 recorded the highest level of succinic acid at 1.10 ± 0.02 mg/mL. These findings highlight the impact of preparation methods and strain-specific characteristics on the bioactivity of postbiotics, providing valuable insights for optimizing postbiotic production in food preservation applications.