Quantification of monosaccharides through multiple-reaction monitoring liquid chromatography/ mass spectrometry using an aminopropyl column

A simple, sensitive, and reproducible quantitative liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was designed for the simultaneous quantification of monosaccharides derived from glycoprotein and blood serum using a multiple-reaction monitoring (MRM) approach. Sialic acids and neutral monosaccharides were efficiently separated using an amino-bonded silica phase column. Neutral monosaccharide molecules were detected as their aldol acetate anion adducts [M+CH₃CO₂]^- using electrospray ionization in negative ion MRM mode, while sialic acids were detected as deprotonated ions [M-H]^-. The new method did not require a reduction step, and exhibited very high sensitivity to carbohydrates with limits of detection of 1 pg for the sugars studied. The linearity of the described approach spanned over three orders of magnitude (pg to ng). The method was validated for monosaccharides originating from N-linked glycans attached to glycoproteins and glycoproteins found in human blood serum. The method effectively quantified monosaccharides originating from as little as 1 μg of glycoprotein and 5 μL of blood serum. The method was robust, reproducible, and highly sensitive. It did not require reduction, derivatization or postcolumn addition of reagents.