With the purpose of analyzing the size and composition of enzyme-unextractable proteins in differently heat-treated soybean meals, a selection of extractants was screened for their ability to extract these proteins from enzyme-unextractable residues. The largest effects were obtained with urea, urea plus β-mercaptoethanol, and dilute alkali; the latter extracted up to 87% of the enzyme-unextractable protein. Gel permeation chromatography indicated that a large proportion of the extracted material was of high molecular weight. However, the combined results from gel electrophoresis, LC-MS, and MALDI-ToF MS showed that the extracted protein material was composed of aggregated peptides. The largest aggregates were observed in the enzymatic residues originating from meals heat-treated at high humidity. Extracted aggregates were fully degraded upon subsequent proteolytic treatment. Chemicals/CAS: amino acid, 65072-01-7; mercaptoethanol, 60-24-2; protein, 67254-75-5; proteinase, 9001-92-7; soybean protein, 9010-10-0; urea, 57-13-6; Amino Acids; Carbohydrates; Endopeptidases, EC 3.4.-; Mercaptoethanol, 60-24-2; Peptides; Soybean Proteins; Urea, 57-13-6.