Lotus seed (Nelumbo nucifera) protein-derived calcium-binding peptides: Isolation, characterization, and osteogenic...

Natural protein-derived peptides are gaining attention for their potential in promoting health, particularly in nutraceutical formulations. In this study, calcium-binding peptides from lotus seed were produced and characterized using UV, FT-IR, Raman, and EDS, and SEM. The calcium-peptide (LSPIH-Ca) complex was subjected to its osteogenic effect in murine bone marrow-derived mesenchymal stem cells (D1 MSCs). Peptides from 4 % Alcalase® showed the highest calcium-binding capacity (49.14 μg/mg). Spectroscopy indicated calcium bound to LSPIH via amide (1537 cm^-1 to 1553 cm^-1), carboxyl, carbonyl, and carboxylate groups. Moreover, the LSPIH-Ca complex enhanced osteoblast differentiation, as evidenced by increased alkaline phosphatase activity and mineralization (123.29 % at 100 μg/mL). Underlying mechanism investigation revealed that the LSPIH-Ca complex stimulated osteogenesis by activating MAPK pathways and upregulating key osteogenic marker such as BMP-2/4, Runx2, ALP activity, COLI, and promoting mineralization. Furthermore, the LSPIH-Ca complex improved calcium transport than Ca²⁺ alone in Caco-2 cells (30.0 ± 2.01 μg/well within 120 min). Finally, this study was conducted to purify and identify calcium-binding peptides and the identified calcium-binding peptides composed of pentapeptide to nonapeptide. The findings of this study establish a theoretical foundation for the creation of innovative calcium nutraceutical additives aimed at enhancing calcium bioavailability and supporting health benefit.