2.10. Assay for Copper Chelating Activity

The copper-chelating activity was assessed by a spectrometric method using PCV [40]. An aqueous solution of 200 μM PCV was incubated at 25 °C for 20 min in the absence and presence of 200 μM CuSO₄ and a test substance, and the absorption spectrum was recorded using a Shimadzu UV-1650PC spectrophotometer. The copper chelating activity was evaluated by the absorbance at 632 nm (A₆₃₂) of the [Cu²⁺-PCV] complex.