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Determination of 4-methylumbelliferone after separation from its conjugates by high-performance liquid
Abstract:
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.