Effect of surface functionalization on vitamin D₂ bioavailability in fortified cereal and millet flours using Caco-2 cells

The study assessed the bioaccessibility and bioavailability of vitamin D2 in fortified cereals and millet flours using Caco-2 cells. Vitamin D2 was extracted from agro-waste of Agaricus bisporus stalks and added to wheat, rice, pearl millet, and finger millet flours through surface functionalization. The bioaccessibility of vitamin D2 was found to be 69.48%, 70.43%, 70.28%, and 71.34%, respectively, compared to 64.42% for the control vitamin D2. Notably, the cellular uptake of vitamin D2 from fortified flours (25–27%) was approximately 12% higher than that of the control vitamin D2 (18.6%). The improved bioaccessibility and cellular uptake are attributed to the ability of surface functionalization to facilitate the binding of vitamin D2 particles to the flour surface via Van der Waals forces, thereby protecting them from degradation. Cytotoxicity analysis using the MTT assay confirmed the safety of the fortified vitamin D2, with over 80% cell viability observed at concentrations ranging from 12.5 to 50 µg/mL. However, cell viability significantly decreased beyond 75 µg/mL, with the half-maximal inhibitory concentration (IC50) of 172.65 µg/mL. Furthermore, vitamin D2 fortified finger millet flour demonstrated a 20% improvement in calcium absorption (31.99%) compared to unfortified finger millet flour (26.67%), highlighting the role of vitamin D2 in enhancing calcium uptake. These findings highlight surface functionalization as an effective method for fortifying staple flours with vitamin D2, enhancing bioavailability and calcium absorption. This strategy offers a promising solution for combating vitamin D deficiency and improving nutrition in populations dependent on staple flours.