Regioselective degradation of [beta] 1,3 glucan by ferrous ion and hydrogen peroxide (Fenton oxidation)

Abstract

Many species use Fe+2 and H2O2 to oxidize a wide variety of compounds to simpler molecules. Both pathogen killing by leukocytes (neutrophils and lymphocytes) and degradation of cellulose by brown rot fungi rely on excretion of Fe+2 ions and H2O2, the Fenton reagent. To elucidate the mechanism of Fenton oxidation of carbohydrates, β1,3 glucan (laminaran), a major fungal wall polysaccharide, was oxidized using a molar ratio of monomer/Fe+2/H2O2 of 10:1:1 (primarily). We labeled the reaction products and profiled them as fluorescent-labeled molecules in polyacrylamide gels and as hydrophobic-tagged molecules using reverse phase liquid chromatography/mass spectrometry (HPLC/MS).Sub-stoichiometric concentrations of Fe+2 and H2O2 fragmented laminaran into smaller molecules containing carbonyl and carboxylic acid groups visible on fluorescent-labeled carbohydrate polyacrylamide gel electrophoresis. HPLC/MS analysis of glucan fragments showed masses consistent with six classes of molecules: aldoses, dialdoses, uronic acids, hexosuloses, aldonic acids, and hexulosonic acids.The results were consistent with published mechanisms where hydrogen radical (H•) abstraction from a C–H or O–H bond begins a cascade of reactions leading to 1) C–C bond cleavage to produce aldose/dialdose pairs; 2) oxo-group (O = ) addition to produce uronic and aldonic acids; 3) hydroxyl group (HO-) addition to produce gluconolactone and hexosuloses; and 4) hexulosonic acids. Most products resulted from regioselective H• abstractions characteristic of oxidations by ferryl-oxo ion [(FeO)+2] or perferryl-oxo ion [(FeO)+3] in close contact with specific positions in the glycan. Therefore, oxidations initiated by regioselectively-bound Fe ions were the predominant initiators of polysaccharide degradations.