The enzyme specific activity was defined and determined as previously described 1The specific activity of D-Lac and its various variants towards D-PL was evaluated in 25 mL triangular vials utilizing

The enzyme's specific activity was defined and determined in accordance with established protocols [1]. To evaluate the specific activity of D-Lac and its various variants towards D-PL, a reaction system consisting of 768 mM DL-PL, Tris-HCl buffer (500 mM, pH 7.5), and 200 uL of diluted purified enzyme was prepared in 25 mL triangular vials. The reaction was allowed to proceed for 20 minutes at 30°C, after which the enzyme was inactivated through boiling and the resulting product was analyzed via reversed-phase high-performance liquid chromatography. Samples were detected in a single layer using a state-of-the-art Waters HPLC system equipped with a Chiralcel IG-3 column (4.6 mm × 250 mm, 3 µm, Diacel, Tokyo, Japan)