Redox hydrogel based bienzyme electrode for L-glutamate monitoring

Amperometric bienzyme electrodes based on coupled L-glutamate oxidase (GlO(x)) and horseradish peroxidase (HRP) were constructed for the direct monitoring of l-glutamate in a flow injection (FI)-system. The bienzyme electrodes were constructed by coating solid graphite rods with a premixed solution containing GlO(x) and HRP crosslinked with a redox polymer formed of poly(l-vinylimidazole) complexed with (osmium (4-4'-dimethylbpy)(2) Cl)(II.III). Poly(ethylene glycol) diglycidyl ether (PEGDGE) was used as the crosslinker and the modified electrodes were inserted as the working electrode in a conventional three electrode flow through amperometric cell operated at - 0.05 V versus Ag\AgCl (0.1 M KCI). The bienzyme electrode was optimized with regard to wire composition, Os-loading of the wires, enzyme ratios, coating procedure, flow rate, effect of poly(ethyleneimine) addition, etc. The optimized electrodes were characterized by a sensitivity of 88.36 +/- 0.14 mu A mM(-1) cm(-2), a detection limit of 0.3 mu M (calculated as three times the signal-to-noise ratio), a response time of less than 10 s and responded linearly between 0.3 and 250 mu M (linear regression coefficient = 0.999) with an operational stability of only 3% sensitivity loss during 8 h of continuous FI operation at a sample throughput of 30 injections h(-1). (C) 1999 Elsevier Science B.V. All rights reserved.