Comparative Mass Spectrometry Analysis and Immunomodulatory Effects of Casein Glycomacropeptide O-Glycans in Bovine and Caprine Whey Powder

Casein glycomacropeptide carries various O-glycan modifications, which, together with variations in the amino acid composition of the glycopeptide, may result in different biological activities. In this study, O-glycans of casein glycomacropeptide from bovine and caprine whey powder were qualitatively and quantitatively analyzed by LC-UV-ESI-MS/MS, and their immune activities and regulatory mechanisms were compared. O-Glycans' total content was 1.54 times higher in bovine than in caprine glycomacropeptide. The glycoform H1N1S2 (H: hexose; N: N-acetylgalactosamine; and S: N-acetylneuraminic acid) accounted for nearly 50% of total glycomacropeptide O-glycans in bovine milk but less than 20% in caprine milk. Bovine glycomacropeptide glycosylation promoted the immune activity of RAW264.7 cells, which may be linked to a higher content of disialylated O-glycans. Glycomacropeptide from both milk sources significantly upregulated the mRNA expression of IL-1α, TNF-α, and IL-10 in RAW264.7 cells and activated the MAPK immunomodulatory signaling pathway. This study demonstrates the possible use of casein glycomacropeptide as an immunomodulatory agent.