Two commercial proteases (subtilisin-typed FNA from Bacillus amyloliquefaciens, and chymotrypsin-like NPP from Nocardiopsis prasina), porcine pepsin, porcine pancreatin having protease activity and their combinations were studied in vitro by LC-MS for their ability to digest soy protein isolate (SPI) under conditions close to those found in the stomach (pH 3.7) and small intestine (pH 6.5). The total number of peptides generated, and their size distribution were obtained under each set of the digestion conditions. These peptides were grouped according to their C-terminal amino acid (AA) residue (P1) and mass, based on which two concepts were proposed, i.e., Normalized Peptide Bond Cleavage Frequency (NPBCF) and Protease Substrate Broadness Index (PSBI). At pH 3.7, FNA+pepsin increased PSBI vs. pepsin alone by 2.7 and 4.9 percentage points (p.p.