*2.8. Determination of Fe (II) Ion Chelation*

Chelation of iron ions by hydrolysates was estimated by the method of Xu et al. [14] with modifications. A 250 μL sample was mixed with 1250 μL H2O and 110 μL 1 mM FeCl2. After 2 min, 1 mL of 500 μM ferrozine (Sigma, 160601) aqueous solution was added and the mixture was allowed to react for 10 min. The absorbance of ferrous iron–ferrozine complex was measured spectrophotometrically at λ = 562 nm. A known concentration of FeCl2 (0–20 μg) was used to generate a standard curve, and the ability to chelate iron ions was expressed as μg Fe2+/mg protein.
