Absolute quantification of twelve oligosaccharides in human milk using a targeted mass spectrometry-based approach

Human milk oligosaccharides (HMOs) are beneficial to infants' growth. Currently, the challenges were the complex operation and weak separation which limited the accurate quantification of HMOs. Liquid chromatography-mass spectrometry (LC-MS/MS) under multi-reaction monitoring (MRM) mode was developed here to separate and quantitatively determine 12 oligosaccharides in human milk. The degree of polymerization of 12 oligosaccharides ranged from 3 to 6. Among 12 HMOS, four pairs of isomeric oligosaccharides (LNFPⅠand LNFPⅢ; LSTa、LSTb and LSTc; 2FL and 3FL; 3SL and 6SL) were successfully separated. Variations of HMOs among 61 individuals from Beijing, China were observed using the present method. Pairwise correlations analysis showed high correlation in abundance between LNDFH Ⅱand 6SL, LSTc and 6SL, LNDFH Ⅱ and LSTa, LSTa and LSTc, LSTa and 6SL. This method allows accurate and reliable quantitative determination of 12 oligosaccharides in human milk, and could be adopted in determining variations in HMOs among samples.