Comparative glycation extent and structure-function differences between casein and whey protein

Casein (CN) and whey protein (WP) were compared as substrates for chitooligosaccharide (COS) glycation under a controlled Maillard-reaction framework. Protein:COS ratio, reaction time, temperature, and initial pH were screened using the degree of glycation (DG) as the primary index. The conjugates were characterized by SEM, UV spectroscopy, FTIR, particle size, and ζ-potential, and their solubility, foaming/emulsifying properties, and radical-scavenging responses were evaluated. Both systems showed maximum glycation at protein:COS (w/w) = 1:4 during ratio screening, but CN reached a higher value than WP (40.09% vs 30.74%). CN also reached its optimal window under milder conditions (120 min, 85 °C, pH 9) than WP (150 min, 95 °C, pH 10). Glycation reduced particle size from 372.00 to 277.27 nm for CN and from 299.93 to 248.27 nm for WP, while WP-COS showed a marked decrease in the absolute ζ-potential value from 39.43 to 20.17 mV. Functionally, CN-COS markedly improved solubility at pH 5 (55.34% vs 18.10% for CN), whereas WP-COS showed the highest foaming capacity. In a proof-of-concept Lactobacillus rhamnosus GG (LGG) test, CN-COS-LGG showed higher encapsulation efficiency and better survival retention during storage and simulated gastrointestinal exposure than CN-LGG. These findings identify protein-dependent glycation windows and support CN-COS as a promising casein-based probiotic carrier candidate.