Sonja Jakovetić Tanasković

Soy protein concentrate (SPC) often has limited food applications due to the loss of its functional properties under harsh industrial processing. This study explored the effects of exposure time to high-intensity ultrasound (HUS) on the structural properties of SPC to assess the potential of a single protein for multiple bakery applications. HUS treatment modified SPC free sulfhydryl group content (4.81 ± 0.03 to 1.47 ± 0.01 µmol/g_protein) and hydrophobicity (34.17 ± 0.02 to 30.56 ± 0.03 µgBPB/mg_protein) and promoted the formation of soluble and insoluble aggregates, especially with longer exposure times, as evidenced by SDS-PAGE. According to Raman analysis, SPC exposed to 0.5 min HUS exhibited an α-helical content of 33.52 ± 1.58% and β-sheet content of 56.80 ± 4.40%, while the tyrosine doublet (I₈₅₀/I₈₃₀) ratio was associated with dough stability and indicated intermolecular hydrogen bonding within the dough matrix. Water absorption capacity was improved upon addition of HUS-exposed SPC samples, to 58.4 ± 0.71%, compared with 52.6 ± 0.85% of SPC-enriched dough. These changes accelerated dough development time and enhanced amylase activity, resulting in a dough with desirable viscosity. HUS-exposed samples with higher α-helix content and solubility, decreased water syneresis, and hydrophobic SPC formed stabile complexes with hydrophobic regions of the amylose chain, both leading to reduced starch retrogradation (1.551 ± 0.13 to 0.855 ± 0.04). Overall, this study showed that by controlling the HUS treatment time, protein structure can be tailored for its use in diverse bakery applications, further enhancing the commercial value of protein concentrates.

In this study, the effects of spinach and pumpkin leaf proteins, as well as the addition of soy protein, on dough rheology were investigated. Changes in rheological properties were attributed to the distinct interactions of proteins, which differ in their functional characteristics—such as solubility and water/oil binding capacity, and structural features, including hydrophobicity and sulfhydryl group content. The dough water absorption capacity increased from 52.6 ± 0.21% to 57.6 ± 0.33%, 55.5 ± 0.42%, and 53.1 ± 0.12% when soybean, spinach, and pumpkin leaf proteins were incorporated into wheat flour at 5% w / w , respectively. In addition to improved water absorption, changes in dough development time, dough stability, viscosity, and the shelf life of the final product were observed as a result of the incorporation of these plant proteins. A higher content of free sulfhydryl groups observed in pumpkin (49.07 ± 1.03 µmol/g protein) and spinach leaf proteins (58.19 ± 0.49 µmol/g protein) was associated with a noticeable reduction in dough development time and stability compared to soy protein, where a considerably lower free sulfhydryl content (6.21 ± 0.21 µmol/g protein) was measured. The formation of a dough with higher mixing stability was attributed to hydrophobic interactions between soy proteins and similarly hydrophobic gluten. Significantly higher polyphenol content and biological activity were exhibited by dough enriched with pumpkin and spinach leaf proteins compared to dough made with wheat flour and fortified with soy protein. In addition, the essential amino acid profile of the enriched dough was notably improved through the addition of these plant‐based proteins to wheat flour.

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The recovery of valuable nutritional compounds, like proteins, from waste streams and by-products is a key strategy for enhancing production sustainability and opening up new market potential. This research aimed to use high-intensity ultrasound as an innovative technique to extract the soluble proteins from the pumpkin leaves. The impact of various sonication amplitudes and duration periods on protein yield, functional properties, antioxidant qualities, and structural characteristics, were studied. Utilization of ultrasound technology significantly increased the yield of pumpkin leaf protein by up to 40%—six times higher than maceration. The ultrasound extraction provided a RuBisCO-rich protein fraction with high radical scavenging and chelating activities, especially at 40% amplitude. Cavitation modified the tertiary and secondary structures of leaf proteins: the amount of α-helix changed based on amplitude (12.3–37.7%), the amount of random coil increased to 20.4%, and the amount of β-turn reduced from 31 to 18.6%. The alteration of the protein fluorescence spectrum (blue shift in spectrum) provides further evidence that ultrasound alters the proteins’ molecular structure in comparation with maceration; the maximum tryptophan fluorescence intensity decreased from 22.000 to 17.096. The hydrophobicity values of 76.8–101.5 were substantially higher than the maceration value of 53.4, indicating that ultrasound improved the hydrophobicity of protein surfaces. Ultrasound resulted in a significant increase in solubility in an acidic environment with the increase in sonication amplitude. A 2.4-fold increase in solubility at pH 2 becomes apparent (20% amplitude; 43.1%) versus maceration (18.2%). The emulsifying ability decreases from 6.62 to 5.13 m²/g once the sonication amplitude increases by 20–70%. By combining the ultrasound periods and amplitudes, it is possible to create high-value protein leaf extracts with improved properties which can find real application as food additives and dietary supplements.

Research background. The development of a novel process for the production of xylooligosaccharides (XOS) based on the 4R concept is made possible by the integration of numerous techniques, especially enzymatic modification together with the physical pretreatment of renewable materials. This study aims to integrate the use of agricultural wastes for the production of xylanase by a new strain of Penicillium sp. and value-added products, XOS. Experimental approach. For the production of xylanase, a solid-state fermentation was performed using wheat bran as substrate. To obtain the most active crude extract of xylanase, the time frame of cultivation was first adjusted. Then, the downstream process for xylanase purification was developed by combining different membrane separation units with size exclusion chromatography. Further characterisation included determination of the optimal pH and temperature, determination of the molecular mass of the purified xylanase and analysis of kinetic parameters. Subsequently, the hydrolytic ability of the partially purified xylanase in the hydrolysis of alkali-extracted hemicellulose from soybean hulls was investigated. Results and conclusions. Our results show that Penicillium rubens produced extracellular xylanase at a yield of 21 U/g during solid-state fermentation. Using two ultrafiltration membranes of 10 and 3 kDa in combination with size exclusion chromatography, a yield of 49 % and 13-fold purification of xylanase was achieved. The purified xylanase (35 kDa) cleaved linear bonds β-(1→4) in beechwood xylan at a maximum rate of 0.64 μmol/(min·mg) and a Michaelis constant of 44 mg/mL. At pH=6 and 45 °C, the purified xylanase showed its maximum activity. The xylanase produced showed a high ability to hydrolyse the hemicellulose fraction isolated from soybean hulls, as confirmed by thin-layer chromatography. In the hydrothermally pretreated hemicellulose hydrolysate, the content of XOS with different degrees of polymerisation was detected, while in the non-pretreated hemicellulose hydrolysate, the content of xylotriose and glucose was confirmed. Novelty and scientific contribution. Future research focusing on the creation of new enzymatic pathways for use in processes to convert renewable materials into value-added products can draw on our findings.

In this study, we assessed the effects of microwave irradiation of wheat gluten proteins as a pretreatment performed in a microwave reactor that could accurately control process parameters as a function of power and temperature, as well as comparing it with conventional heat treatment. The aim was to identify suitable combinations of partial enzymatic hydrolysis and microwave pretreatment parameters to produce gluten hydrolysates with reduced allergenicity and conserved techno-functional features for food application. FTIR analysis, and total and reactive SH group contents confirmed that the microwave-controlled heating can significantly change the secondary structure and conformation of gluten protein. The microwave treatment had the largest effect at 200 W and 100 °C, at which the content of gluten has been reduced by about 2.5-fold. The microwave pretreatment also accelerated the enzymatic hydrolysis of gluten, changing the kinetic profile. The apparent hydrolysis rate constants (k₂) were 1.00, 3.68, 3.48, 4.64 and 4.17 min⁻¹ for untreated gluten, and those pretreated with microwave power of 200, 400, 600 and 800 W, respectively. Compared to the heat treatment, it appeared that microwave specific non-thermal effects had a significant influence on the gluten structure and allergenicity and, in combination with the enzymatic hydrolysis, ultimately yielded protein hydrolysates with enhanced antioxidant and functional properties.