Tailoring aqueous and enzymatic extraction processes for enhanced functionality of lentil proteins

Lentils are becoming an important source of plant-based proteins for applications in foods products. While previous research has explored the optimization of lentil protein extraction for yields and purity, functionality-driven extraction approaches have yet to be developed. Aqueous (AE) and enzyme-assisted extraction (EAE) were utilized to extract proteins from lentil flour. An initial screening of commercial proteases assessing the impacts of EAE on total protein extractability (TPE), solubility at pH 4.0, and in vitro protein digestibility (IVPD) demonstrated that EAE with Neutral Protease (NP) increased TPE (72 to 77%) and solubility (50 to 90%) compared to AE (pH 7, no enzyme). Therefore, EAE with NP was selected for subsequent optimization, yielding two possible enzyme loadings (0.5 and 0.87% w/w) that achieved similarly high extraction yields (79-81% TPE) compared to AE (73% TPE). Physicochemical analyses revealed that as the amount of enzyme increased, the isoelectric point decreased from pH 4.9 to 2.7, and the surface hydrophobicity significantly decreased. Differential scanning calorimetry demonstrated that proteolysis caused a downward shift in thermal transition temperatures and denaturation enthalpy, suggesting that EAE modified protein folding and thermal stability. EAE improved protein solubility at pH 4 (33% for AE vs. 73-95% for EAE), but decreased emulsifying capacity, with the use of 0.87% NP resulting in proteins unable to form emulsions at pH 4.0 or 7.0. Although EAE had minimal impact on foaming capacity, the use of 0.87% NP significantly increased foaming stability at pH 4.0 (71% for AE to 93%). Proteins produced with moderate enzyme concentration (0.5%) exhibited the most favorable overall functionality. Nevertheless, extracts produced using 0.87% enzyme could be useful in acidic food applications due to its enhanced solubility and foaming stability. These findings underscore the importance of judicious selection of enzyme loading in producing protein extracts with desired functional properties.