Experimental and in silico approaches for the buffalo whey protein-folic acid complexation elucidation. Molecular changes impacting on protein structure and functionality.

Using a buffalo whey proteins concentrate (BWPC) as a nanocarrier of labile bioactive compounds as vitamins constitutes a very innovative approach with potential application in the food and nutraceutical industries. This work aims to deepen the knowledge of the phenomena occurring in the complexation process of vitamin B9 with BWPC, providing valuable information on the molecular and functional properties of complexes and intervening substances. For such purpose, analytical (SEC-FPLC, Fluorescence spectroscopy, FTIR, DLS, UV-vis spectroscopy) and in-silico methods (molecular docking) were performed to get complementary data. Five types of proteins were identified in the BWPC. Folic acid (FA) interacted with BWPC in buffer pH 7 through H-bonds and hydrophobic interactions, inducing conformational changes and modifying the secondary and tertiary protein structure. The resultant BWPC-FA complexes showed a size distribution in the nanoscale (100-150 nm) with no aggregation. Molecular docking showed that lactoferrin had the highest FA binding affinity. Complexation did not reduce the antioxidant activity of intervening substances. Indeed, the radical scavenging capacity of BWPC-FA was 20 % higher than single BWPC. The obtained results provide relevant data enabling the adding value of the main effluent of buffalo dairy industries.

Molecular interactions involved in the complexation process between buffalo whey proteins concentrate and folic acid.

Using buffalo whey proteins as carrier agents of sensitive molecules raises an interesting approach allowing adding value and minimizing the pollution impact of this by-product. In this context, this work aims to explore the molecular interactions between buffalo whey proteins concentrate (BWPC) and folic acid (FA). For this purpose, fluorescence, UV and FTIR analysis were performed on aqueous or solid dispersions of a buffalo whey protein concentrate (5 µM) (BWPC), with variable concentrations (0-20 µM) of FA. Fluorescence and absorption data were fitted by Stern-Volmer, Beckett, Förster resonance energy transfer, and sphere-of-action models (R2 > 0.9). Derived results suggest that BWPC strongly bind to FA through non-covalent interactions and form ground-state complexes. Additionally, BWPC improves the photostability of FA against UV radiation, and chemical denaturation negatively affects the binding properties. Obtained results encourage further studies of BWPC as carrier agents, which could promote innovative applications for this under-utilized proteins source.