Analysis of urinary oligosaccharides in lysosomal storage disorders by capillary high-performance anion-exchange chromatography-mass spectrometry

Abstract

Many lysosomal storage diseases are characterized by an increased urinary excretion of glycoconjugates and oligosaccharides that are characteristic for the underlying enzymatic defect. Here, we have used capillary high-performance anion-exchange chromatography (HPAEC) hyphenated to mass spectrometry to analyze free oligosaccharides from urine samples of patients suffering from the lysosomal storage disorders fucosidosis, α-mannosidosis, G(M1)-gangliosidosis, G(M2)-gangliosidosis, and sialidosis. Glycan fingerprints were registered, and the patterns of accumulated oligosaccharides were found to reflect the specific blockages of the catabolic pathway. Our analytical approach allowed structural analysis of the excreted oligosaccharides and revealed several previously unpublished oligosaccharides. In conclusion, using online coupling of HPAEC with mass spectrometric detection, our study provides characteristic urinary oligosaccharide fingerprints with diagnostic potential for lysosomal storage disorders.

Fig. 1

Separation of oligosaccharides in urine of a fucosidosis patient. H hexose, N N-acetylhexosamine, F fucose, S N-acetylneuraminic acid, BPC base peak chromatogram

Fig. 2

Positive-ion fragmentation mass spectrum of the monosodiated disaccharide HexNAc₁Fuc₁ (precursor ion at m/z 390.2) from urine of a fucosidosis patient. Red triangle = fucose, blue square = N-acetylglucosamine

Fig. 3

Histograms showing the relative abundance of the detected glycans in the urine sample (Table 1) of lysosomal storage disorders fucosidosis (a), α-mannosidosis (b, c, and d), G_M1-gangliosidosis (e), G_M2-gangliosidosis (f and g), and sialidosis (h). H or white circle = hexose, N or white square = N-acetylhexosamine, F or red triangle = fucose, S or purple diamond = N-acetylneuraminic acid, NeuGc = N-glycolylneuraminic acid, X = hexonic acid, SO ₃ = sulfate, yellow circle = galactose, green circle = mannose, blue square = N-acetylglucosamine

Fig. 3

Histograms showing the relative abundance of the detected glycans in the urine sample (Table 1) of lysosomal storage disorders fucosidosis (a), α-mannosidosis (b, c, and d), G_M1-gangliosidosis (e), G_M2-gangliosidosis (f and g), and sialidosis (h). H or white circle = hexose, N or white square = N-acetylhexosamine, F or red triangle = fucose, S or purple diamond = N-acetylneuraminic acid, NeuGc = N-glycolylneuraminic acid, X = hexonic acid, SO ₃ = sulfate, yellow circle = galactose, green circle = mannose, blue square = N-acetylglucosamine

Fig. 4

Positive-ion fragmentation mass spectrum of the monosodiated tetrasaccharide Man₃GlcNAc₁ (precursor ion at m/z 730.6) from urine of a α-mannosidosis patient. Green circle = mannose, blue square = N-acetylglucosamine

Fig. 5

Separation of oligosaccharides in urine of a G_M1-gangliosidosis patient. H hexose, N N-acetylhexosamine, BPC base peak chromatogram

Fig. 6

Positive-ion fragmentation mass spectrum of the disodiated diantennary oligosaccharide Hex₅HexNAc₃ (precursor ion at m/z 742.1) from urine of a G_M1-gangliosidosis patient. Yellow circle galactose, green circle mannose, blue square N-acetylglucosamine

Fig. 7

Separation of oligosaccharides in urine of a G_M2-gangliosisis patient. Fragment ion spectra of the species A and B is shown in Fig. 8. F fucose, H hexose, N N-acetylhexosamine, BPC base peak chromatogram

Fig. 8

Positive-ion fragmentation mass spectra of two isomeric monosodiated tetrasaccharides Hex₂HexNAc₂ (precursor ion at m/z 771.5) from urine of a G_M2-gangliosisis patient. The separation of A and B is shown in Fig. 7. Green circle mannose, blue square = N-acetylglucosamine