Azodicarbonamide, Hydrogen Peroxide, and l-Ascorbic Acid Aid in the Modification of Protein Texture During High-Moisture Meat Analog Extrusion Processing.

Protein texturization during high-moisture extrusion is a complex process. Functional additives, like azodicarbonamide (ADA), hydrogen peroxide (HO), and l-ascorbic acid (AA), play a critical role in modulating protein structure and texture during food processing applications. However, the specific effects of these additives on protein texture, microstructure, and functionality during high-moisture meat analog (HMMA) extrusion remain underexplored. This study aimed to understand how these additives at different concentrations (0.1%-2.0%) influence the texturization of wheat protein isolate (WPI) during extrusion at 45% and 60% moisture levels. WPI was selected due to its well-known reactivity and fibrous texturization capability. The extrudates were analyzed for their texture, microstructure, and physicochemical characteristics. Amino acid composition was determined using ninhydrin-based ion-exchange chromatography. At low concentrations, ADA, HO, and AA improved cross-linking and aggregation within the protein matrix, increasing hardness, chewiness, and cutting force. However, higher concentrations disrupted the protein network, yielding more porous and fragmented structures, evident from the observed scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) micrographs. Color (L*, a*, b*) shifted significantly with increasing additive concentration, with ADA resulting in brighter and yellower samples, whereas AA and HO had moderate effects. Amino acids mostly remained unchanged, but key residues, such as proline and lysine content, were slightly modified and correlated with the observed textural modifications. This study investigated how thermally reactive and oxidative additives influence protein cross-linking and aggregation during extrusion, and how these molecular-level changes govern fibrous texture formation in plant-based meat analogs. Potential ADA and HO residues post-extrusion underscore the need for future safety-focused quantification.