Determination of 4-methylumbelliferone after separation from its conjugates by high-performance liquid chromatography. Application to lysosomal enzyme activity assays.
A high-performance liquid chromatographic method is described for the separation and estimation of 4-methylumbelliferone in the presence of its conjugates. The technique utilizes a simple, isocratic eluent and fluorometric detection. Percentages of 4-methylumbelliferone in the conjugates and fluorescences of the conjugates are reported. 4-Methylumbelliferone, liberated under currently used conditions for enzyme activity in urine and fibroblasts, can be measured by this procedure because these materials contain no substances interfering with the 4-methylumbelliferone peak. Applied to lysosomal enzymatic activity analyses, this procedure eliminates substrate background fluorescence. Determinations of eight lysosomal enzymes in urine and fibroblasts are presented in bar graph form.
D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004798:Enzymes / Q000652:urine*; D005347:Fibroblasts / Q000201:enzymology; D005470:Fluorometry; D006801:Humans; D006923:Hymecromone / Q000302:isolation & purification*; D008247:Lysosomes / Q000201:enzymology*; D014468:Umbelliferones / Q000302:isolation & purification*
D004798:Enzymes; D014468:Umbelliferones; D006923:Hymecromone