HPLC ANALYSIS OF URIDINE-DIPHOSPHATE SUGARS - DECREASED CONCENTRATIONS OF URIDINE-DIPHOSPHATE GALACTOSE IN ERYTHROCYTES AND CULTURED SKIN FIBROBLASTS FROM CLASSICAL GALACTOSEMIA PATIENTS

A high performance liquid chromatography (HPLC) method has been develo ped for the measurement of uridine diphosphate galactose (UDPGal) and uridine diphosphate glucose (UDPGlc) in erythrocytes and cultured skin fibroblasts of normal controls and galactosemia patients. The method incorporates an internal standard, UDPxylose, and alkaline phosphatase for the removal of nucleoside phosphates in the regions of UDPGal and UDPGlc in the chromatographic system, UDPGal and UDPGlc were separate d on dual Dionex Carbo Pac PA-1 anion exchange columns. Samples derive d from galactosemia patients without any detectable erythrocyte galact ose-1-phosphate uridyl transferase (GALT) activity (GG), had in compar ison to the controls significantly lower levels of UDPGal in both eryt hrocytes and cultured skin fibroblasts (P = 0.0001), while galactosemi a patients with detectable GALT activity (GV) had normal levels of UDP Gal. GG patients on oral uridine therapy had normal levels of UDPGal. These findings are consistent with our previous reports using an enzym atic method, In terms of absolute values, the values for both UDPGal a nd UDPGlc in erythrocytes by HPLC analysis were lower than those repor ted with the enzymatic method; however, in cultured skin fibroblasts, the values were similar with the two methods.