LC-MS/MS-based non-isotopically paired labeling (NIPL) strategy for the qualification and quantification of monosaccharides

• 03 March 2026

Investigation into monosaccharides is critical for studies of oligosaccharides structure and function in biological
processes. However, monosaccharides quantification is still challenge due to their isomeric structure and high
hydrophilic properties. Besides, it was difficult to obtain isotopic internal standards (IS) of each monosaccharide
in complex matrixes. Herein, we developed a novel strategy for the qualification and quantification of monosaccharides
in urine using two structure analogs 1-(4-methylphenyl)-3-methyl-5-pyrazolone (MPMP) and1-
phenyl-3-methyl-5-pyrazolone (PMP) as non-isotopically paired labeling (NIPL) reagents by liquid
chromatograph-tandem mass spectrometry (LC-MS/MS). The derivatized monosaccharides by NIPL method not
only had sufficient retention time differences on reversed-phase column, but also exhibited predominant product
ion pairs (m/z 189 & m/z 175) in the multiple reaction monitoring (MRM) mode. In this method, PMP labeled
standards were adopted as one-to-one internal standards (ISs). 12 urinary monosaccharides were successfully
determined and the linear ranges expanded five orders of magnitude with limit of quantification (LOQ) varied
from 0.09 ng mL

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