Inhibition and Promotion of Heat-Induced Protein Modifications in Whey by Vitamins: Quantitative Profiling of 14 Modifications at 29 β-Lactoglobulin Binding Sites

Nonenzymatic post-translational modifications (nePTMs) induced by thermal processing of whey-based foods impair the protein quality and digestibility. This study evaluates the impact of 14 water- or fat-soluble vitamins on the formation of heat-induced nePTMs in whey proteins. Whey samples were fortified with vitamins and subjected to heat treatment. After Glu-C hydrolysis, 14 nePTMs at 29 binding sites in β-lactoglobulin were quantified by microflow-liquid chromatography-electrospray ionization tandem mass spectrometry in scheduled multiple reaction monitoring mode. The analysis included primary and advanced Maillard products, oxidation products, deamidation and deamination products. This comprehensive structure- and site-specific approach revealed that vitamin B2, folic acid, vitamin A, and vitamin B12 were the most effective inhibitors against nePTM formation. Vitamin K2, vitamin D3, retinoic acid, vitamin K1, and β-carotene demonstrated moderate inhibitory effects. In contrast, vitamin B1 and biotin tended to promote modifications, while vitamin C affected different reaction mechanisms in varying ways.