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Hidden “Digestome”: Current Analytical Approaches Provide Incomplete Peptide Inventories of Food Digests

De Cicco, Maristella, Mamone, Gianfranco, Di Stasio, Luigia, Ferranti, Pasquale, Addeo, Francesco, Picariello, Gianluca
Journal of agricultural and food chemistry 2019 v.67 no.27 pp. 7775-7782
algorithms, allergenicity, beta-lactoglobulin, bioinformatics, crosslinking, disulfides, high performance liquid chromatography, ions, lactalbumin, polypeptides, proteomics, tandem mass spectrometry, trichloroacetic acid, whey protein
Analyzing an in vitro gastroduodenal digest of whey proteins by high-performance liquid chromatography (HPLC) coupled to high-resolution/high-sensitivity tandem mass spectrometry (MS/MS), we sought to evaluate if state-of-art peptidomics provide comprehensive peptide coverage of food “digestomes”. A multitude of small-sized peptides derived from both α-lactalbumin and β-lactoglobulin as well as disulfide cross-linked hetero-oligomers remained unassigned, even when the digests were compared before and after S–S reduction. The precipitation with 12% trichloroacetic acid demonstrated the occurrence of large-sized polypeptides that escaped the bioinformatic identification. The analysis of a HPLC–MS/MS run with different proteomic search engines generated dissimilar peptide subsets, thus emphasizing the demand of refined searching algorithms. Although the MS/MS fragmentation of monocharged ions with exclusion of non-peptide-interfering compounds enlarged the inventory of short peptides, the overall picture of the “digestome” was still incomplete. These findings raise relevant implications for the identification of possible food-derived bioactive peptides or allergenic determinants.