A Sensitive Method for the Quantification of b-Glucuronidase Activity in Urine Using Capillary Electrophoresis with Fluorescence Detection
No Thumbnail Available
Authors
Wu, X.
Loganathan, D.
Linhardt, Robert J.
Issue Date
1998
Type
Article
Language
ENG
Keywords
Biology , Chemistry and chemical biology , Chemical and biological engineering , Biomedical engineering
Alternative Title
Abstract
Capillary electrophoresis (CE) with fluorescence detection was used to determine the concentration of 4-methylumbelliferone liberated from 4-methylumbelliferyl-beta-D-glucuronide by beta-glucuronidase. Enzyme substrate saturation kinetics were studied in buffer and the pH range for the enzyme reaction was optimized. A linear relationship of initial enzyme reaction velocity as a function of peak area of enzyme product was obtained for enzyme activity ranging from 1 to 100 units. The beta-glucuronidase activity in urine was next determined. Freshly collected urine samples were dialyzed, the retentate was incubated with 4-methylumbelliferyl-beta-D-glucuronide, boiled and centrifuged. The supernatant was separated by CE in an uncoated capillary with 0.1 M sodium acetate buffer by applying a voltage of 12 kV. The product of the enzymatic reaction, 4-methylumbelliferone, was detected by fluorescence, facilitating the determination of as little as one unit of beta-glucuronidase activity in a 0.5-h incubation time, with an error of less than +/-5%.
Description
Journal of Chromatography, 708, 61-66
Note : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
Note : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
Full Citation
A Sensitive Method for the Quantification of b-Glucuronidase Activity in Urine Using Capillary Electrophoresis with FluorescenceDetection, X. Wu, D. Loganathan, R.J. Linhardt, Journal of Chromatography,708,61-66, 1998.