A quantitative comparison of the extraction of protein fractions from wheat grain by different solvents, and of the polypeptide and amino acid composition of the alcohol-soluble proteins

Using several cultivars, whole milled wheat grain was first extracted with 0.5M NaCl to remove non‐protein N, albumins and globulins: the amount of protein subsequently extractable by either 70% ethanol, 55% propan‐2‐ol, or 50% propan‐1‐ol (with or without the addition of acetic acid), was then compared. All solvents were tested at 4, 20 and 60°C both with and without the addition of 2‐mercaptoethanol (2‐ME) as a reducing agent. Raising the temperature, including a reducing agent, and repeated extractions, all combined to maximise protein extractability; least was extracted by 70% ethanol at 4°C and most by 50% propan‐1‐ol containing acetic acid and 2‐ME at 60°C. The differing polypeptide compositions of these alcohol‐soluble fractions illustrated why N solubility alone is not a sufficient guide to protein extraction, e.g. 35% of grain N was extractable by either 70% ethanol at 60°C or 70% ethanol containing 2‐ME at 4°C, but high mol. wt polypeptides (>60000) were found only in the 60°C extract. In contrast, the complete range of alcohol‐soluble polypeptides was extracted by all solvent mixtures based on propan‐1‐ol, even at 4°C. Amino acid analyses of these alcohol‐soluble fractions confirmed the low levels of lysine in fractions free from high mol. wt polypeptides. More glycine was found in all fractions containing high mol. wt polypeptides, but only those extracted by propan‐1‐ol mixtures had an increased lysine content, the amount of which increased linearly as the extraction temperature was raised. Defatting the milled grain did not affect the extractability of alcohol‐soluble proteins, but the amount of protein soluble in 0.5M NaCl decreased. The protein rendered NaCl‐insoluble was not extracted by any of the alcoholic solvents tested, hence the N content of the residual material was increased. The inclusion of this denatured metabolic fraction in the residue will affect its amino composition. The problems arising from the retention of the classical nomenclature for Osborne‐type fractions are discussed.