A Strategy for Annotating the Human Milk Glycome

TLDR

Oligosaccharides are abundant bioactive molecules in human milk that, despite lacking direct nutritive value, are thought to support intestinal microflora and mucosal immunity. The study aims to resolve the unknown exact composition of these oligosaccharides so their biological functions can be understood. Researchers isolated oligosaccharides from milk lipids and proteins and used HPLC‑Chip/MS, MALDI‑FT ICR MS, and tandem MS with exoglycosidase digestion to determine composition and distinguish structural isomers. Orthogonal TOF MS identified about 200 distinct oligosaccharide species, and HPLC‑Chip/MS profiling revealed compositional variation among women, establishing a routine identification method. Keywords: Human milk oligosaccharides; milk glycome; HPLC‑Chip/MS technology; infrared multiphoton dissociation; exoglycosidases; matrix‑assisted laser desorption/ionization Fourier transform mass spectrometry.

Abstract

Oligosaccharides in human milk represent a group of bioactive molecules that have evolved to be an abundant and diverse component of human milk, even though they have no direct nutritive value to the infant. A recent hypothesis proposes that they could be substrates for the development of the intestinal microflora and the mucosal immune system. The inability to determine the exact composition of these oligosaccharides limits research and the ability to understand their biological functions. Oligosaccharides isolated from the lipids and proteins of individual human milk samples were analyzed by a combination of techniques including microchip liquid chromatography mass spectrometry (HPLC-Chip/MS) and matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FT ICR MS). Accurate mass measurements obtained using an orthogonal time-of-flight (o-TOF) mass spectrometry provided oligosaccharide composition for approximately 200 individual molecular species. Comparison of HPLC-Chip/MS profiles from five different women revealed variations in milk oligosaccharide compositions. HPLC-Chip/MS profiling provides a method for routinely identifying milk oligosaccharides. Tandem MS in combination with exoglycosidase digestion provides unambiguous differentiation of structural isomers. Keywords: Human milk oligosaccharides; milk glycome; HPLC-Chip/MS technology; infrared multiphoton dissociation; exoglycosidases; matrix-assisted laser desorption/ionization Fourier transform mass spectrometry