β-lactoglobulin/Linoleic Acid Complexes – Formation, Binding Stoichiometry and Cytotoxic Capability
Résumé
β-lactoglobulin (β-lg) is a major whey protein in bovine milk, which has the ability to bind hydrophobic ligands such as fatty acids. It is well known that the whey protein alpha-lactalbumin also forms a stable complexe with fatty acid, which is cytotoxic to a wide variety of tumour cells. The aim of this project was to investigate the binding stoichiometry of β-lg and linoleic acid (LA; cis,cis-9,12-octadecadienoic acid) and investigate the impact of this complex on viability of intestinal cancer cells in vitro. The interaction between β-lg and fatty acids was studied by HPLC and GC. Complexes with a binding stoichiometry of 1, 2 and 3 LA:β-lg were produced. The effect of the complexes on cell viability was measured by exposing the human colorectal adenocarcinoma epithelial cell line, Caco-2, to various concentrations of complexes from 0 to 72 hours. LA on its own exhibited a cell viability effect at 58 M as measured by MTS assays. A cytotoxic effect at 35 M was measured by an impedance-based biosensing microelectronic real-time cell analyzer system (XCELLigence RTCA system, Roche Diagnostics). Neither the β-lg (0 to 75 μM) nor β-lg/LA complexes (LA concentration from 0 to 200 μM) had an adverse effect on Caco-2 cells. In vivo it is assumed that β-lg gradually releases LA during gut transit which, in turn, may modulate fatty acid uptake by the gut and/or have a satiating effect. Future work will investigate fatty acid uptake and satiety signaling in the gut in response to β-lg/LA complexes.