Mixed-phase weak anion-exchange/reversed-phase LC-MS/MS for analysis of nucleotide sugars in human fibroblasts.

Autor: Rahm M; Department of Neurology, Donders Institute for Brain, Cognition, and Behavior, Radboud University Medical Center, Geert Grooteplein 10, 6525 GA, Nijmegen, The Netherlands., Kwast H; Translational Metabolic Laboratory (TML), Department of Human Genetics, Radboud University Medical Center, Geert Groote Plein Zuid 10, 6525 GA, Nijmegen, The Netherlands., Wessels HJCT; Translational Metabolic Laboratory (TML), Department of Human Genetics, Radboud University Medical Center, Geert Groote Plein Zuid 10, 6525 GA, Nijmegen, The Netherlands., Noga MJ; Laboratory of Clinical Genetics, Inborn Errors of Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands., Lefeber DJ; Department of Neurology, Donders Institute for Brain, Cognition, and Behavior, Radboud University Medical Center, Geert Grooteplein 10, 6525 GA, Nijmegen, The Netherlands. dirk.lefeber@radboudumc.nl.; Translational Metabolic Laboratory (TML), Department of Human Genetics, Radboud University Medical Center, Geert Groote Plein Zuid 10, 6525 GA, Nijmegen, The Netherlands. dirk.lefeber@radboudumc.nl.
Jazyk: angličtina
Zdroj: Analytical and bioanalytical chemistry [Anal Bioanal Chem] 2024 Jun; Vol. 416 (15), pp. 3595-3604. Date of Electronic Publication: 2024 Apr 27.
DOI: 10.1007/s00216-024-05313-w
Abstrakt: Nucleotide sugars (NS) fulfil important roles in all living organisms and in humans, related defects result in severe clinical syndromes. NS can be seen as the "activated" sugars used for biosynthesis of a wide range of glycoconjugates and serve as substrates themselves for the synthesis of other nucleotide sugars. NS analysis is complicated by the presence of multiple stereoisomers without diagnostic transition ions, therefore requiring separation by liquid chromatography. In this paper, we explored weak anion-exchange/reversed-phase chromatography on a hybrid column for the separation of 17 nucleotide sugars that can occur in humans. A robust and reproducible method was established with intra- and inter-day coefficients of variation below 10% and a linear range spanning three orders of magnitude. Application to patient fibroblasts with genetic defects in mannose-1-phosphate guanylyltransferase beta, CDP-L-ribitol pyrophosphorylase A, and UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase showed abnormal levels of guanosine-5'-diphosphate-α-D-mannose (GDP-Man), cytidine-5'-diphosphate-L-ribitol (CDP-ribitol), and cytidine-5'-monophosphate-N-acetyl-β-D-neuraminic acid (CMP-Neu5Ac), respectively, in consonance with expectations based on the diagnosis. In conclusion, a novel, semi-quantitative method was established for the analysis of nucleotide sugars that can be applied to diagnose several genetic glycosylation disorders in fibroblasts and beyond.
(© 2024. The Author(s).)
Databáze: MEDLINE
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