Aggregation of peptides in soy protein isolate hydrolysates : the individual contributions of glycinin- and ß-conglycinin-derived peptides

Keywords: Soy proteins, glycinin, β-conglycinin, enzymatic hydrolysis, peptides, aggregation, gelation, identification, mass-spectrometry, mappingThe aim of the work presented in this thesis is to understand howlimited enzymatic hydrolysis can influence the aggregation behavior of soy protein material. This is performed by uncoupling the enzymatic degradation and the aggregation process. Subsequently, the regions in soy proteins from which the aggregating peptides originate were identified.Hydrolysates of soy protein isolates (SPI) were prepared using the protease subtilisin Carlsberg. The enzyme was inhibited when the desired degree of hydrolysis (DH) was reached. It was shown that with increasing DH, the pH at which a gel could be formed was increasing as well. The aggregation behavior of SPI, glycinin- and β-conglycinin-derived hydrolysates showed that glycinin-derived peptides are responsible for the aggregation of SPI-derived peptides. This was also found when glycinin was hydrolyzed with other enzymes (bromelain, papain, and chymotrypsin) using SPI, β-conglycinin, and bovine whey proteins as reference materials. Only hydrolysis of glycinin by trypsin did not result in strong aggregating peptides.Subsequently, a new method was developed, denoted accumulative-quantitative-peptide-mapping. This method reveals those regions in the parental protein from which the aggregating peptides originate. This method comprises a second hydrolysis of the aggregating peptides, followed by separation, identification, and quantification of individual peptides obtained. Quantification was performed based on absorbance at 214 nm, corrected for the calculated molar extinction coefficient based on the amino acid composition. This novel method revealed that mainly the basic polypeptide and that part of the acidic polypeptide close to the location of the disulfide bridge connecting the basic to the acidicpolypeptide,are the predominant regions of glycinin yielding aggregating peptides. These regions have a relative high hydrophobicity compared to that of the total protein. Upon hydrolysis the net hydrophobicity of the remaining glycinin is increasing, eventually resulting in aggregation and subsequent gelation.