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KMID : 1146120170030010020
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Journal of Mucopolysaccharidosis and Rare Diseases
2017 Volume.3 No. 1 p.20 ~ p.27
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Keratanase II Digestion Accompanied with a Liquid Chromatography/Tandem Mass Spectrometry for Urinary Keratan Sulfate Quantitative Analysis
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Chuang Chih-Kuang
Lin Hsiang-Yu
Wang Tuen-Jen
Huang Sung-Fa
Lin Shuan-Pei
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Abstract
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Purpose: Mucopolysaccharidosis IV (MPS IV) is a disease characterized by deficient activity of N-acetylgalactosamine-6-sulfatase (GALNS) causing excessive lysosomal storage of keratan sulfate (KS). The identification of the relevant disaccharide units of KS after keratanase II digestion followed by liquid chromatography/tandem mass spectrometry detection (LC-MS/MS) is validated and applicable for the preliminary diagnosis of MPS IV.
Methods: A total of 67 urine samples were collected and analyzed from 11 MPS IV patients comprising 10 MPS IVA and one MPS IVB patients, and 56 normal controls. Urinary glycosaminoglycan was first precipitated by the Alcian blue method followed by a digestion of keratanase II. The protonated species of the digested disaccharide products were detected by using multiple reaction monitoring experiment.
Results: One particular disaccharide of KS was selected. The transition mass-to-charge (m/z) of the parent ion and its daughter ion after collision was 462.0→97.0, whereas the chondrosine used as an internal standard in this assay was m/z 353.9→73.0. The results corresponded well with the two-dimensional electrophoresis method. The quantities of urinary KS were significantly raised in confirmed MPS IV patients when comparing with those of normal controls (170.2±81.1 vs. 4.06±1.92 μg/mL).
Conclusion: The LC?MS/MS method for MPS IVA determination is specific, sensitive, validated, and applicable for urinary KS quantification. This method can be used not only as a first-line biochemistry examination of MPS IVA, but also as an outcome survey after enzyme replacement therapy.
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KEYWORD
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Mucopolysaccharidosis IV, Keratan sulfate (KS), Keratanase, Liquid chromatography/tandem mass spectrometry (LC-MS/MS), Two-dimensional electrophoresis
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