Strategieën voor efficiëntere extractie van functionele proteïnen uit aardappelsnippers en soja-okara

Abstract

A future-proof food system should make optimal use of the available resources. Recovering proteins from by-products of plant-based food processing can contribute to the growing global protein demand. However, the extractability of plant proteins is limited by various factors, such as the entrapment in cellular structures or preceding processing steps. Additionally, recovered proteins should be functional to allow their use in food applications. Against this background, this dissertation aimed to evaluate both single and combined strategies for improving the extraction of functional proteins from potato trimmings and soybean okara, two case studies relevant to the (Belgian) food industry. Potato trimmings are a by-product from potato fries production, and okara is the insoluble residue obtained during the production of soy-based drinks or tofu. Proteins in potato trimmings and okara were shown to have different inherent extractabilities. In potato trimmings, approximately 50% of the proteins were readily extractable, while 25% were aggregated, and the remaining 25% were physically inaccessible, likely trapped within cellular structures. The extractability of okara proteins was low, with 50% restricted by protein aggregation and the other 50% by physical inaccessibility. Therefore, a comparison of identical extraction processes between these by-products can provide insights into how protein extraction is influenced by differences in biomass composition and inherent protein extractability. Ultrasound-assisted protein extraction disrupted potato and soybean cells with increasing ultrasonication time. This increased the protein yield up to 98% for potato trimmings and up to 90% for okara. Short ultrasound treatments (2000 J/g fresh weight, about 5 min) were especially effective in solubilizing aggregated okara proteins, increasing the protein yield from <10% to about 50%. Potato trimming proteins obtained by extended ultrasonication showed a slower adsorption at the air-water interface due to induced protein aggregation, which was correlated to a reduced foamability. In contrast, increasing ultrasound times decreased the size of extracted okara protein aggregates, which positively affected their foaming properties. In general, okara proteins were bad foaming agents, likely due to their aggregated nature. In contrast, isolated potato trimming proteins had very good foaming properties. The mechanism behind foam stabilization of potato proteins, and its major fractions patatin and protease inhibitors, was therefore investigated more in-depth. The foaming properties of proteins isolated from potato trimmings were better than those of a blend of commercial potato proteins containing similar relative amounts of patatin and protease inhibitors. This was the case despite the fact that more proteins were insoluble in the isolates from potato trimmings than the commercial isolates. Likely, impurities also contributed to these superior foaming properties, given the lower purity of potato trimming protein isolates compared to the commercial isolates. Patatin and protease inhibitors were shown to both contribute considerably to the formation and stabilization of foams. Interestingly, a beneficial effect on the foam stability was found when patatin and protease inhibitors of various colloidal sizes were present. Generally, the importance of acknowledging and studying the heterogeneity in potato protein fractions for understanding their foaming properties was highlighted. Incubation of the by-products with cell wall degrading enzymes (at pH 5.0) increased the protein yield after subsequent alkaline extraction (at pH 9.0) from 57 to 76% for cellulase pre-treated potato trimmings, and from 17 to 35% for pectinase pre-treated okara. A short ultrasound treatment during the enzyme-assisted process could be used to increase the efficiency, but only when applied during the alkaline extraction step. When applied during the enzymatic incubation at pH 5.0, protein recoveries were not increased or even decreased, despite that the ultrasound treatment could induce additional cell disruption. This was attributed to induced protein aggregation when ultrasound was applied at pH values close to the protein isoelectric point. The combined enzyme pre-treatment and ultrasound-assisted alkaline extraction increased the yield up to 87% for potato trimmings and up to 54% for okara. The pectinase pre-treatment of okara allowed the extraction of a less aggregated protein fraction while the additional ultrasound treatment extracted more aggregated proteins. The molecular weight distribution of extracted potato proteins was similar regardless of the used process. Mechanical particle size reduction by ball milling more efficiently disrupted cellular structures in freeze-dried potato trimmings than in freeze-dried okara. However, for both matrices, the protein recovery at pH 9.0 was not remarkably increased despite the observed particle size reduction. The reason for this differed between the by-products. For potato trimmings, the ball milling treatment induced protein aggregation which reduced their solubility, achieving a net zero effect of the treatment on the protein yield. For okara, the inherent aggregated nature of the proteins limited their extractability, regardless of the level of cellular disruption. For both matrices, applying a short ultrasound treatment at the start of the alkaline extraction was effective in extracting the aggregated proteins. This resulted in consecutive increases in protein recovery with decreasing particle size, with increases up to 76% for potato trimmings and up to 64% for okara. The combined ball milling pre-treatment and ultrasound-assisted isolation process reduced the purity and foamability of the isolated potato trimming proteins. Extensive co-isolation of starchy compounds was avoided by adding starch degrading enzymes during protein isolation, leading to an improvement of the protein foaming properties compared to proteins isolated from potato trimmings that were not ball milled. This was due to a more efficient air-water interfacial adsorption, presumably due to the formation of soluble aggregates containing partially unfolded potato proteins. In general, potato proteins had good foaming properties regardless of the used process, and alterations were better understood by investigating changes in their structural and air-water interfacial properties. Okara protein isolates always had low purity and contained heavily aggregated proteins regardless of the used process, limiting their ability to stabilize gas cells in foams. To summarize, this dissertation led to several key conclusions. A first conclusion is that cell disruption alone does not guarantee increased protein extractability. The impact of the cell disruption method itself should be considered, as well as the inherent solubility of the proteins. A second conclusion is that rational combinations of treatments are more effective in improving the extraction efficiency compared to single treatments. Third, identical processes can have different effects depending on the substrate, both in terms of how they impact protein recovery as well as how they impact protein functionality. Therefore, knowledge on the extraction-limiting factors and inherent protein properties is required to tailor extraction processes to specific substrates.