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Bioinformatics tools were used to predict radical scavenging and metal chelating activities of peptides derived from abundant potato, seaweed, microbial, and spinach proteins. The antioxidant activity was evaluated in 5% oil-in-water emulsions (pH4) and best-performing peptides were tested in mayonnaise and compared with EDTA. Emulsion physical stability was intact. The peptide DDDNLVLPEVYDQD showed the highest protection against oxidation in both emulsions by retarding the formation of oxidation products and depletion of tocopherols during storage, but it was less efficient than EDTA when evaluated in mayonnaise. In low-fat emulsions, formation of hydroperoxides was reduced 4-folds after 5 days compared to control. The concentration effect of the peptide was confirmed in mayonnaise at the EDTA equimolar concentration. The second-best performing peptides were NNKWVPCLEFETEHGFVYREHH in emulsion and AGDWLIGDR in mayonnaise. In general, the peptide efficacy was higher in low-fat emulsions. Results demonstrated that peptide negative net charge was important for chelating activity.

Peptide emulsifiers derived from plant proteins are gathering growing interest as green and sustainable replacements for chemical additives in food. Potato (Solanum tuberosum) is one of the most important crops for both human consumption and industrial processing. Globally, the annual production of potato starch exceeds 3,000,000 MT with more than 200,000 MT of potato protein isolated as a side-stream, providing an enormous source of raw protein. The direct isolation of food-grade protein is in many cases regarded as cost-ineffective. Nevertheless, potato proteins may be a valuable source of functional peptides.Previously, we demonstrated amphiphilicity-based bioinformatic prediction of peptide emulsifiers embedded in potato proteins1,2. Amongst the predicted peptides, especially one (γ1), derived from the storage protein patatin, showed exceptional emulsifying activity in vitro. Although patatin is the most abundant protein in potatoes, it is not a single protein, but a family of highly homologous isoforms. Using bottom-up proteomics in combination with multiple sequence alignment and in silico digestion, we identified several γ1 variants. The variants consist of both full-length isoforms with single amino acid substitutions and tryptic variants/truncations from different patatin isoforms. The emulsifying activity of the γ1 variants, physical stability of the emulsions during storage, and interfacial properties were investigated3. Furthermore, the interfacial conformation of the peptides was investigated by SRCD and supplemented by NMR and benchtop CD for selected peptides in micellar model systems. Based on these results, we are able to i) evaluate the full potential of using γ1 variants as peptide emulsifiers in food; ii) provide novel insight on the structure/function relationship of amphipathic, α-helical peptide emulsifiers; iii) combine in vitro functional validation with in silico proteolysis to design a scalable and targeted enzymatic hydrolysis, resulting in a hydrolysate with improved emulsifying properties; and iv) apply the hydrolysates as stabilizers for encapsulation of fish oil for foods.

The demand for functional ingredients from natural and more sustainable sources is increasing. In recent years, peptides have been investigated for their functions including their antioxidant activity and ability to prevent oxidation in food. Previously, we have identified antioxidant peptides (4-30 AA residues) embedded in potato, seaweed, single cell, and spinach RuBisCO proteins using bioinformatics, validated their radical scavenging activity using in vitro assays (e.g. DPPH) and tested selected peptides’ ability to limit lipid oxidation in 5% fish oil-in-water emulsions at pH 7. In this work, we evaluated the effect of the same selected set of peptides (0.05 wt%) on the oxidative stability of iron-mediated (50 µM) 5% fish oil-in-water emulsions stabilized with 1 wt% of Tween 20 at pH 4. Results showed that all the peptides had antioxidant effect compared to control emulsions without any peptides based on the formation of hydroperoxides, consumption of tocopherols and the formation of secondary volatile oxidation compounds. Different peptides did not have any effect on the physical stability of the emulsions. Based on the emulsion study, the four best performing peptides from each protein source were selected and their effects on oxidative stability of fish oil enriched mayonnaise were investigated. Two control samples were included, one with EDTA, which is a commonly used synthetic antioxidant in mayonnaise, and another one without any addition of antioxidants . Increase in peroxide value and volatile compounds (e.g. 1-penten-3-ol, (E,E)-2,4-heptadienal, and (E,Z)-2,6-nonadienal) formed during the storage showed that none of the antioxidant peptides prevented lipid oxidation to the same extent as EDTA; however, potato peptide provided the best protection among all peptides followed by the seaweed peptide .

Poster for LAB13 (13th International Symposiumon Lactic Acid Bacteria)