Investigating the impact of ultrasound-assisted extraction on the angiotensin-converting enzyme (ACE) inhibitory activity of chickpea (Cicer arietinum L.) peptides: In vitro, in silico, and acute toxicity study.

This study aimed to assess the isolation, purification, characterization and acute toxicity study of chickpea (Cicer arietinum L.) peptides with angiotensin-converting enzyme inhibitory (ACE-I) activity prepared through ultrasound-assisted hydrolysis by alcalase. The ultrasonicated chickpea protein hydrolysate (UCPH) was purified sequentially using ultrafiltration (UF) and fast protein liquid chromatography (FPLC). The <3 kDa fraction (UCPH-1) from UF, showing vigorous ACE-I activity (85.82 % ± 0.03, IC₅₀: 0.35 mg/mL), was further purified by FPLC. The most active FPLC fraction, F6 (95.76 % ± 0.22, IC₅₀: 0.20 mg/mL), was identified as the peptide MLRVYLR (UCPH-1-F6). This peptide remained stable (91.1 %) after gastrointestinal digestion and acted as a competitive ACE inhibitor. In addition, molecular docking showed MLRVYLR forms Pi-interactions and hydrogen bonds with ACE, and it exhibited no acute oral toxicity. Based on these results, the purified peptide from hydrolysed chickpea protein treated with ultrasound exhibits possible anti-hypertensive characteristics and could be utilised in functional foods, pharmaceuticals, and nutraceuticals.