title: Rapid ultraperformance liquid chromatography-tandem mass spectrometry assay for a characteristic glycogen-derived tetrasaccharide in Pompe disease and other glycogen storage diseases.
abstract: BACKGROUND
  Urinary excretion of the tetrasaccharide 6-α-D-glucopyranosyl-maltotriose (Glc₄) is increased in various clinical conditions associated with increased turnover or storage of glycogen, making Glc₄ a potential biomarker for glycogen storage diseases (GSD). We developed an ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) assay to detect Glc₄ in urine without interference of the Glc₄ isomer maltotetraose (M₄).
  
  
  METHODS
  Urine samples, diluted in 0.1% ammonium hydroxide containing the internal standard acarbose, were filtered, and the filtrate was analyzed by UPLC-MS/MS.
  
  
  RESULTS
  We separated and quantified acarbose, M₄, and Glc₄ using the ion pairs m/z 644/161, 665/161, and 665/179, respectively. Response of Glc₄ was linear up to 1500 μmol/L and the limit of quantification was 2.8 μmol/L. Intra- and interassay CVs were 18.0% and 18.4% (10 μmol/L Glc₄), and 10.5% and 16.2% (200 μmol/L Glc₄). Glc₄ in control individuals (n = 116) decreased with increasing age from a mean value of 8.9 mmol/mol to 1.0 mmol/mol creatinine. M₄ was present in 5% of urine samples. Mean Glc₄ concentrations per age group in untreated patients with Pompe disease (GSD type II) (n = 66) were significantly higher, ranging from 39.4 to 10.3 mmol/mol creatinine (P < 0.001-0.005). The diagnostic sensitivity of Glc₄ for GSD-II was 98.5% and the diagnostic specificity 92%. Urine Glc₄ was also increased in GSD-III (8 of 9), GSD-IV (2 of 3) and GSD-IX (6 of 10) patients.
  
  
  CONCLUSIONS
  The UPLC-MS/MS assay of Glc₄ in urine was discriminative between Glc₄ and M₄ and confirmed the diagnosis in >98% of GSD-II cases.
authors.affiliation: Department of Clinical Genetics, Center for Lysosomal and Metabolic Diseases, Erasmus MC University Medical Center, Rotterdam, the Netherlands.
authors.forename: Wim
authors.initials: W
authors.lastname: Sluiter
authors.forename: Jeroen C
authors.initials: JC
authors.lastname: van den Bosch
authors.forename: Daphne A
authors.initials: DA
authors.lastname: Goudriaan
authors.forename: Carin M
authors.initials: CM
authors.lastname: van Gelder
authors.forename: Juna M
authors.initials: JM
authors.lastname: de Vries
authors.forename: Jan G M
authors.initials: JG
authors.lastname: Huijmans
authors.forename: Arnold J J
authors.initials: AJ
authors.lastname: Reuser
authors.forename: Ans T
authors.initials: AT
authors.lastname: van der Ploeg
authors.forename: George J G
authors.initials: GJ
authors.lastname: Ruijter
mesh_terms: D000293:Adolescent; D000328:Adult; D000367:Age Factors; D000368:Aged; D002648:Child; D002675:Child, Preschool; D002853:Chromatography, Liquid; D006003:Glycogen / Q000378:metabolism*; D006008:Glycogen Storage Disease / Q000652:urine*; D006009:Glycogen Storage Disease Type II / Q000652:urine; D006010:Glycogen Storage Disease Type III / Q000652:urine; D006011:Glycogen Storage Disease Type IV / Q000652:urine; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008320:Maltose / Q000031:analogs & derivatives / Q000652:urine; D008875:Middle Aged; D009844:Oligosaccharides / Q000652:urine*; D012016:Reference Values; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry; D055815:Young Adult
chemical_list: D009844:Oligosaccharides; D008320:Maltose; D006003:Glycogen; C009819:maltotetraose