Mechanistic characterization of iron-catalyzed oxidation of polysorbate 80: The role of ferrous iron, hydrogen peroxide, and superoxide.

We investigated the role of individual radical species during Fe-catalyzed oxidation of PS80. Solutions containing 1 gL^-1 PS80 (0.1 % w/v) in 10 mM acetate buffer (pH 6) were exposed to various amounts of either Fe(II) or Fe(III), hydrogen peroxide (H₂O₂), and various enzymes or antioxidants. PS80 oxidation was measured using a fluorescence micelle assay (FMA) alongside LC-MS. Hydrogen peroxide inhibited PS80 oxidation in the presence of Fe(II) but promoted oxidation in the presence of Fe(III). Furthermore, Ferrostatin-1 (Fer-1), an antioxidant which is known to preferentially react with alkoxy radicals, inhibited PS80 oxidation in the presence of Fe(II). Superoxide dismutase (SOD) partially inhibited PS80 oxidation in the presence of either Fe(II) or Fe(III), suggesting that superoxide plays a role in both cases. Ferryl species (Fe^IV=O) or hydroxyl radicals (HO•), produced by the Fenton reaction, do not play a major role in the oxidation of PS80. Rather, oxidation was initiated by the reaction of both Fe(II) and Fe(III) with pre-existing lipid hydroperoxides on PS80, as well as via superoxide.