To date, no comparative analysis has been conducted regarding the effectiveness of cold plasma (CP) techniques—comprising both vacuum CP (VCP) and atmospheric CP (ACP)—high hydrostatic pressure (HHP), and thermal treatment (ThT) on protein denaturation prior to the electrospinning (ESP). This comparative evaluation is essential, as the current solution (ThT) to improving protein quality before ESP results in significant organoleptic consequences (e.g., color degradation and off-flavor) and nutrient loss. The expected impact of the study is to advance current knowledge of the denaturation dynamics of soy protein isolate (SPI) using green technologies, including VCP, ACP, and HHP. The main aim of the study was to investigate the suitability of green non-thermal technologies to replace inefficient conventional ThT. Thus, different properties of treated SPI samples, including solubility, surface hydrophobicity, sulfhydryl content, turbidity, secondary structure, viscosity and electrophoretic pattern were studied. While physicochemical properties, including solubility, secondary structure, and viscosity of SPI, were improved by CP techniques and HHP, ThT had an edge over non-ThTs in terms of denaturation before ESP. For example, ThT resulted in the highest SPI solubility (39%), sulfhydryl content (19%), turbidity value (5.19), and disordered structure (62.2%). Among non-ThTs, VCP dominated ACP and HHP in different characteristics, e.g., sulfhydryl content, viscosity, disordered structure, random coil content, and α-helix to β-sheet ratio. VCP preserved α-helix better due to oxidation-induced stability and reduced aggregation, while HHP increased β-sheet formation due to pressure-induced unfolding and aggregation. VCP generates reactive oxygen and nitrogen species, inducing mild amino acid oxidation that stabilizes α-helix via intra-molecular hydrogen bonding while preventing extensive unfolding.
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