Proteins constitute a crucial component of human nutrition, with meat acknowledged as the most superior source of protein. As a vital nutrient for human growth and health, meat ensures the provision of essential amino acids and micronutrients, while also playing a key role in energy metabolism regulatory processes (Jiménez-Colmenero, Carballo, & Cofrades, 2001). Despite these benefits, the cholesterol and high proportion of saturated to unsaturated fatty acids contained in meat are significantly associated with various diseases like type II diabetes, stroke, coronary heart disease, and particularly colon cancer (Biesalski, 2005).

As consumer health and food safety are critical concerns in the food industry, multiple approaches have been created to facilitate the reduction of meat consumption among consumers. One strategy is the innovation of structured products, which can serve as a replacement for meat and meat products and are consumable as meat analogues. Meat analogues are products that emulate meat in functionality, exhibiting product properties and sensory characteristics that are similar to those of meat (Kumar et al., 2017).

The production of meat analogues makes use of various plant proteins, with soy protein being the most commonly used. Soy is a widely consumed pulse that provides favorable nutritional and performance benefits, and is often used as a partial or complete replacement for meat due to its high nutrient content and lowered risk of cardiovascular disease. However, plant proteins, including soy protein, generally have weaker gelation capacity compared to animal proteins due to lower solubility and higher denaturation temperature, which limits their use in the food industry (Basak & Singhal, 2022). Therefore, it is important to investigate methods to improve the gelling capacity of soy protein for use as an ingredient in plant protein-based meat and egg analogs.

Two methods investigated for improving the gelling capacity of soy protein are cold plasma and pH-shifting. Atmospheric cold plasma is comprised of energetic species with a neutral charge, including free electrons, ions, photons, atoms, molecules, and free radicals (Q. Zhang et al., 2021). Previous studies have demonstrated that these reactive species can promote oxidative reactions in macromolecules and change their physicochemical properties (Sarangapani, Patange, Bourke, Keener, & Cullen, 2018). Reactive species generated by cold plasma can induce conformational and spatial structural changes in proteins (Han, Cheng, & Sun, 2019) through reaction with sulfur-containing amino acids (cysteine and methionine) and the aromatic rings of tryptophan, tyrosine, and phenylalanine. Additionally, cold plasma treatment can cause alterations in protein molecular weight and secondary structures, resulting in modification of physicochemical properties of these macromolecules (Venkataratnam, Sarangapani, Cahill, & Ryan, 2019).

Dong, Guo, Chen, Chen, Ji, Ran, et al. (2018) reported that the formation of crosslinking via disulfide bonds in cold plasma treated zein, resulted in improved mechanical properties of zein film. In addition, Ji, Dong, Han, Li, Chen, Li, et al. (2018) observed that cold plasma treatment can modify the solubility, emulsion stability, and water holding capacity of peanut protein. Mehr and Koocheki (2020) demonstrated that DBD plasma positively affects the interfacial and emulsifying properties of Grass pea protein isolate. Our recent study also suggested that the treatment of SPI with DBD plasma at 16, 18, and 20 kV for 5, 10, and 15 min can improve physicochemical properties of the protein compared with the untreated sample (Sharafodin & Soltanizadeh, 2022a). However, in that study, the pH-shifting treatment was not applied on the protein.

The pH-shifting method is a convenient approach for unfolding the structure of proteins in an extreme acid-base environment, which ultimately causes a conformational change in the globular protein resulting in the formation of a “molten globule” structure. Upon neutralization, the protein structure is refolded (J. Jiang, Xiong, & Chen, 2011). There are several studies that involve modifying both plant and animal proteins using the pH-shifting method. Zhu, Huang, Guo, and Chen (2021) indicated that pH-shifting can improve the gelling properties of pea protein isolate. Also, it was reported that the unfolding of SPI after alkaline pH-shifting improved its solubility, surface hydrophobicity, and emulsification (Yan, Xu, Zhang, Xie, Zhang, Jiang, et al., 2021). In particular, the application of pH-shifting has improved the foaming and emulsifying properties of pea and soy protein by increasing protein solubility and surface hydrophobicity. This phenomenon is attributed to partial protein unfolding, which results in a balance between hydrophilic and hydrophobic groups exposed on the protein surface (J. Jiang, Chen, & Xiong, 2009).

Karabulut, Kapoor, Yemis, and Feng (2024) indicated that the combination of high pressure homogenization and manothermosonication with pH-shifting can change the structure of hemp protein isolate and considerably improve its functional properties. The structural change of chickpea protein after ultrasonic assisted pH-shifting improved their interfacial properties and foaming capacities due to protein unfolding (Wang, Wang, Li, Wang, Xiang, Li, et al., 2022). Previous research has demonstrated the noteworthy impact of DBD plasma and pH-shifting on the unfolding of the three-dimensional configuration of globular proteins. Since protein unfolding plays a crucial role in the gelling properties of proteins, it is hypothesized that the combination of DBD plasma treatment and pH-shifting can potentially facilitate the formation of a gel with meat-like characteristics. The purpose of this study was to examine the viability of employing pH-shifting and cold plasma techniques to augment the gelation process of SPI. Moreover, aside from analyzing the mechanical properties of the resultant SPI gel, its functional attributes were also explored to determine its usability in meat products. The impact of these treatments on the mechanical properties of the resulting gel was investigated. If the physical properties of the gel met the criteria of animal protein-derived gels, value-added opportunities would exist for SPI in the food industry through the creation of meat analogues.