Co-encapsulated urease and glucose oxidase in polyelectrolyte microcapsules for simultaneous glucose and urea determination.

BACKGROUND

Clinical monitoring of metabolic disorders requires stable tools for simultaneous analyte detection. However, conventional free-enzyme assays suffer from rapid degradation, poor operational stability, and single-use limitations. Herein, we report a novel bi-enzymatic diagnostic system based on the rational co-encapsulation of glucose oxidase (GOx) and urease within polyelectrolyte microcapsules (PMCs) for the simultaneous determination of glucose and urea.

RESULTS

Fabricated via layer-by-layer assembly, the PMCs achieved high enzyme loading (73 ± 2% for GOx and 93 ± 5% for urease). Co-encapsulation significantly reduced the GOx Michaelis constant from 10.1 to 2.5 mM. The system exhibited linear ranges of 0.5-3.0 mM (glucose) and 1-40 mM (urea), covering clinically relevant concentrations. It maintained performance for over 61 measurement cycles and a 27-day shelf-life at room temperature. Direct analysis of human saliva revealed that the PMCs effectively shield enzymes from matrix interferences, yielding significantly lower data dispersion and higher reproducibility than standard Clark-electrode and Berthelot reference methods.

SIGNIFICANCE: By combining multi-cycle reusability, room-temperature stability, and enhanced precision in raw biological fluids, this co-encapsulation strategy overcomes the fundamental trade-offs of traditional immobilization. The developed PMCs provide a robust, cost-effective, and comprehensive platform for point-of-care metabolic diagnostics and non-invasive monitoring, establishing a generalizable framework for future multi-enzymatic biosensing applications.