DETERMINATION OF PROTEIN-GLUTATHIONE MIXED DISULFIDES IN WHEAT-FLOUR

An HPLC method was adapted for measurement of protein-glutathione mixed disulfides (PSSG) in flour and flour protein fractions. It involves extraction of unbound glutathione with perchloric acid (PCA), release of glutathione from PSSG by dithiothreitol (DTT) reduction, and separation and quantification of the carboxymethylated and dinitrophenylated peptide by HPLC on amino-bonded phase silica. PSSG levels ranged from 70 to 150 nmol/g in flours from different wheat cultivars. PSSG also varied among flour protein fractions obtained by Osborne fractionation. Glutenin had the highest value (897 nmol/g of protein) and albumin a somewhat lower value (796 nmol/g of protein). Levels in globulin and gliadin were considerably lower (239 and 76 nmol/g of, protein, respectively). SDS-unextractable glutenin macropolymer preparations also had high levels (1062 nmol/g of protein, cv. Glenlea; 1538 nmol/g of protein, cv. Bussard). Availability of this methodology for GSH, GSSG, and PSSG analysis provides a powerful tool for elucidating the importance of glutathione in flour and dough systems.