Liquid Chromatographic Assay for a Glucose Tetrasaccharide, a Putative Biomarker for the Diagnosis of Pompe Disease

Abstract A HPLC method associated with butyl- p -aminobenzoate derivatization has been developed for the analysis of a tetraglucose oligomer, Glcα1-6Glcα1-4Glcα1-4Glc, designated Glc 4 , in biological fluids. This tetraglucose, normally excreted in the urine, has previously been shown to be elevated in a number of pathological conditions including Pompe disease (glycogen storage disease type II), which is caused by a deficiency of the lysosomal enzyme acid α-glucosidase. Concentrations of Glc 4 in both urine and plasma were established for the age ranges of 20 years, both in normal individuals and in a cohort of 21 patients with enzymatically confirmed Pompe disease. The Glc 4 concentration decreased with age in both groups, but all the patients had elevated Glc 4 levels compared with age-matched controls. Electrospray tandem mass spectrometry was employed to establish the homogeneity of the HPLC peak for Glc 4 and to investigate the identity of other unusual oligosaccharides excreted in patient urine. Our results demonstrate that this method is suitable for application in clinical laboratories to help establish the diagnosis of Pompe disease.