Abstract
The soybean cold-insoluble protein was hydrolyzed with pepsin and the hydrolyzate was dialyzed. The diffusate was submitted to gel permeation chromatography of Sephadex G-10 and thin layer chromatography of silica gel G. A ninhydrin-negative bitter peptide was detected by chlorine-starch-iodine test and was purified by paper electrophoresis and by rechromatography on the silica gel. Azeotropic HCl hydrolysis of the peptide gave equal molar ratio of Ala, Glu, Gly, Ile, Leu, Phe, Ser and Val. N-Terminal residue was composed of pyrrolidone carboxylic acid, which was tentatively identified in comparison with the authentic sample by paper electrophoresis and thin layer chromatography. Hy-drazinolysis of the peptide, followed by the 2, 4-dinitrophenylation, produced α, α, γ-tri-DNP-glutamic acid-α, γ-dihydrazide and DNP-pyrrolidone carboxylic acid hydrazide, also supporting the N-terminal structure. Hydrolysis of the peptide by carboxypeptidase A gave Leu, Val, Phe, Ile, Ala, etc. in the order of the liberation rate. As the peptide fragments remaining in this hydrolyzate were detected pyrrolidone carboxyl-Gly•OH, pyrrolidone carboxyl-Gly-Ser•OH, pyrrolidone carboxyl-Gly-Ser-Ala•OH, etc. In conclusion, the structure of this peptide was proposed as: pyrrolidone carboxyl-Gly-Ser-Ala-Ile-Phe-Val-Leu•OH. Quantitative information about the total amount of pyrrolidone carboxyl residue contained in the peptic hydrolyzate of the soybean protein and sensory study on the bitterness of the peptide as referred by standard solution of phenylthiourea, were demonstrated.
