Quantitation of a urinary tetrasaccharide by gas chromatography and mass spectrometry

A gas chromatographic mass spectrometric method has been developed for the rapid determination of a urinary tetrasaccharide (alpha-D-Glc-(1 leads to 6)-alpha-D-Glc-(1 leads to 4)-alpha-D-Glc-(1 leads to 4)-D-Glc). The urine sample is first fractionated by gel chromatography. An appropriate internal standard is added to the pooled tri-pentasaccharide fraction, which is then reduced, methylated and fractionated by g.c. The identification of the tetrasaccharide derivative is based on the g.c. relative retention time and the mass spectrum of the reduced permethylated tetrasaccharide. The normal excretion rate was in the range of 0.1-2.5 mg per 24 hours. Greatly increased amounts (9.4-89.6 mg 24 h-1) were found in the urine of patients with glycogen storage disease type II and type III and in one patient with unclassified muscular disease. A moderate increase (3.6m6 mg 24 h-1) was observed in one patient with glycogen storage disease type VI, in two patients with Duchenne muscular dystrophy and in two other patients with unclassified muscular disease.