Effects of Direct Steam Injection Conditions on the Physicochemical and Bioactive Properties of Enzymatically Extracted Black Bean Proteins

ABSTRACT While the bioactivity of pulse protein hydrolysates is widely studied, the holistic impact of unit operations from flour to final powder remains unexamined. This study produced black bean hydrolysate powders using enzyme‐assisted extraction, direct steam injection (low temperature, LT: 130°C/15 s, or high temperature, HT: 145°C/4 s) and spray‐drying to evaluate their effects on bioactivity. Overall, the structural and physicochemical properties of the LT and HT samples were similar, with evidence of denaturation and aggregation through decreased intrinsic fluorescence intensity, increased surface hydrophobicity, and decreased surface charge at pH 4 compared to a non‐thermally treated, freeze‐dried control. Despite structural changes, protein solubility was similar (50%–51% at pH 4, 86%–94% at pH 7), as aggregates remained soluble in neutral conditions. Trypsin inhibitor activity was unaffected by thermal treatment (165–167 trypsin units inhibited/mg protein), but the in vitro protein digestibility of the LT (81%) and HT (83%) samples was significantly higher than the control (51%). Total phenolic content (58–63 mg GAE/g protein), antioxidant activities (ABTS/ORAC) (0.83–1.02 mg GAE/g protein), and antidiabetic activity (α‐glucosidase inhibition, 18%–19%) were mostly retained following direct steam injection and spray‐drying, but ACE‐inhibition activity decreased for HT (70% vs. 89%–93% for LT and control), signifying that antihypertensive peptides may have been degraded or irreversibly aggregated at higher processing temperatures. The results demonstrate that enzyme‐assisted extraction followed by industrially relevant thermal processing and drying is an effective method to generate bioactive black bean hydrolysate powders, supporting future commercialization of functional pulse ingredients for nutraceutical applications.