In vitro and in vivo degradation of glucose oxidase enzyme used for an implantable glucose biosensor.
BACKGROUND
The degradation of the glucose oxidase (GOD) enzyme, commonly used in the construction of glucose sensors has been of concern for scientists for decades. Many researchers have found that GOD deactivates over time, mostly due to H2O2 oxidation. This decay can lead to the eventual failure of the sensor. However, these findings are controversial, because other researchers did not find this degradation.


METHODS
The goal of this study was twofold. The first goal was to evaluate the in vitro and in vivo stability of two commercially available GOD enzymes and the second goal was to evaluate Nafion as a protective coating of GOD. Crosslinked GOD samples were sandwiched between two 10-microm pore polycarbonate membranes (Nafion coated or uncoated) and placed in custom designed Lexan chambers. Chambers were then exposed to a total of five different environments: Dulbecco's Modified Eagle Medium (DMEM) or phosphate buffered saline (PBS) with and without a 5.6-mM glucose concentration, as well as the subcutaneous in vivo environment of 12 rats. After a period of up to 4 weeks, chambers were retrieved, opened, and tested for enzyme activity using a three-electrode system.


RESULTS
Enzyme activity showed only a slight decrease when exposed to DMEM and PBS without glucose. A more dramatic decrease in activity was observed in enzymes exposed to PBS and DMEM with 5.6 mM glucose. The in vivo environment also caused a significant decrease in enzyme activity, but the decrease was lower than for the in vitro environment with glucose conditions.


CONCLUSIONS
The presence of glucose in vitro and in vivo led to the production of H2O2, suggesting this to be the main agent responsible for enzyme degradation. The use of a Nafion coating did not provide any additional protection.
Valdes
T I
TI
University of Connecticut Health Center, Center for Biomaterials & Surgical Research Center, Farmington 06030-1615, USA.
Moussy
F
F
D000818:Animals; D015374:Biosensing Techniques; D001786:Blood Glucose / Q000032:analysis*; D002021:Buffers; D004795:Enzyme Stability; D004867:Equipment Design; D005949:Glucose Oxidase; D006801:Humans; D006861:Hydrogen Peroxide; D007332:Insulin Infusion Systems; D051381:Rats; D017207:Rats, Sprague-Dawley; D012044:Regression Analysis; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D001786:Blood Glucose; D002021:Buffers; D006861:Hydrogen Peroxide; D005949:Glucose Oxidase