Evaluation of glycosidic bond cleavage and formation of oxo groups in oxidized barley mixed-linkage β-glucans using tritium labelling

This study investigated the formation of oxo groups in pure solutions of barley mixed-linkage (1→3),(1→4)-β-d-glucans (MLGs) incubated in the presence of hydrogen peroxide or Fenton reaction-generated hydroxyl radicals (OH), and it gives a fingerprint of products obtained after enzymic and acidic hydrolysis of OH-attacked MLG. Hydroxyl radical, but not hydrogen peroxide, introduced a range of NaB³H₄-reducible functions into MLG chains. Driselase or lichenase digestion of NaB³H₄-reduced MLGs released a complex mixture of ³H-labelled products due to the presence of unusual and incompletely digestible residues in OH-attacked polysaccharide chains. Complete acid hydrolysis of OH-treated MLGs yielded a mixture of ³H-aldoses (mainly glucose, mannose, galactose and allose) deriving from random OH attack at positions 2, 3 or 4 to form glycosulose residues which were NaB³H₄-reducible to epimeric mixtures of ³H-aldose residues. Furthermore, the production of [³H]glucitol demonstrated the radical-mediated cleavage of mid-chain glucose residues to create new reducing termini. Oxidative scission of MLGs by hydroxyl radical caused a decrease in molecular weight of about 96%, which was partially inhibited by the addition of DMSO, an OH scavenger. The results can be a starting point for developing an assay to detect changes due to OH attack of MLGs in vivo, or during food processing and storage, based on the chemical or enzymic release of unusual sugar residues such as allose as diagnostic products after reduction/labelling treatment and partial purification (or selective digestion by lichenase) of polymeric material.