Ascorbic acid, i.e., vitamin C, is a well-known essential nutrient, and has attracted considerable attention as a new candidate for cancer therapy. Previously, R2c consisting of silicon tetra- tert -butylphthalocyanine (SiPc) and two 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) radicals has been encapsulated into the hydrophobic cavity of dimeric bovine serum albumin (BSA), i.e., R2c@(BSA) 2 , and the system was found to behave as a highly sensitive and selective fluorescent probe for detecting ascorbic acid not only in aqueous solutions but also in vivo . In this study, kinetics of the reaction of R2c@(BSA) 2 with ascorbic acid have been studied based on the temporal evolution of fluorescence. Global fitting of the concentration dependence using the Runge-Kutta method revealed the existence of stepwise two proton-coupled electron transfer processes. The rate constants for the reactions with ascorbic acid ( k AA ) and ascorbate radical ( k AR ) were 3 × 10 and 1 × 10 5 min −1 M −1 , respectively, suggesting that the reaction with ascorbate radical was much faster than that with ascorbic acid. These results were further corroborated by theoretical calculations of the Gibbs free energy differences and by spin statistical factors. The analysis presented herein will aid in understanding the two proton-coupled electron transfer processes in the reaction with ascorbic acid.