Quantification of Amino Acids Released During Fermentation

Amino acids released during fermentation were estimated as the amount of soluble protein in trichloroacetic acid (TCA) following the method described by Asensio-Grau et al. (2020 (link)) and Gallego et al. (2021 (link)). Samples (100 mg) were mixed with TCA solution to a final concentration of 12% and incubated at 4 °C for 15 min. Then, samples were centrifuged (Eppendorf MiniSpin Plus) at 4200g-force for 10 min. The supernatant was diluted with 50 mM EDTA and 8 M UREA buffer (pH 10), and the absorbance was measured by ultraviolet spectrophotometry (Helios Zeta UV/Vis, Thermo Scientific) at 280 nm. A calibration line was used for quantification using tyrosine as standard. Results are expressed as grams of soluble protein in TCA per 100 g of protein.