Variations in bovine milk oligosaccharides after calving using p-aminobenzoic ethyl ester closed-ring labeling and negative ion electrospray LC/MS/MS

Han-Ju Chien ^a, Yi-Chun Ma ^b, Yi-Feng Zheng ^a, Cheng-Yu Kuo ^a, Wei-Chen Wang ^a, Guor-Rong Her ^b, Chien-Chen Lai ^a,c,d,e,f,*

a Institute of Molecular Biology, National Chung Hsing University, Taichung, Taiwan

b Department of Chemistry, National Taiwan University, Taipei, Taiwan

c Advanced Plant Biotechnology Center, National Chung Hsing University, Taichung, Taiwan

d Graduate Institute of Chinese Medical Science, China Medical University, Taichung, Taiwan

e Ph.D. Program in Translational Medicine, National Chung Hsing University, Taichung, Taiwan

f Rong Hsing Research Center For Translational Medicine, National Chung Hsing University, Taichung, Taiwan

A strategy was proposed to analyze bovine milk oligosaccharides using p-aminobenzoic ethyl ester (ABEE) closed-ring labeling and C18 capillary liquid chromatography negative ion electrospray tandem mass spectrometry. Linkage specific fragment ions were used to identify oligosaccharide isomers. By constructing the mass chromatograms using linkage specific fragment ions, isomers were differentiated based on m/z values as well as temporal separation provided by liquid chromatography. In addition to disialyllactose and the single isomer lacto-N-neohexaose, four pairs of linkage isomers including 3’/6’-sialyllactose (3’/6’-SL), 3’/6’-sialyllactosamine (3’/6’-SLN), 3’/6’-sialylgalactosyl-lactose (3’/6’-SGL), and lacto-N-tetraose/lacto-N-neotetraose (LNT/LNnT) in bovine milk were investigated. Variations of bovine milk oligosaccharides in a lactation period of 72 hrs after calving were studied. Sialylated oligosaccharide was found to be distinctively more abundant in milk of the first 24 h, decreasing in successive milkings. For the first time, the variation of lacto-N-tetraose in bovine milk was reported.

Keywords: Bovine milk, Closed-ring labeling, Mass spectrometry, Oligosaccharides, p-aminobenzoic ethyl ester

https://doi.org/10.38212/2224-6614.3386