Enzyme-modified Pt nanoelectrodes for glutamate detection

Abstract

We present here a glutamate oxidase (GluOx)-modified platinum (Pt) nanoelectrode with a planar geometry for glutamate detection. The Pt nanoelectrode was characterized using electrochemistry and scanning electron microscopy (SEM). The radius of the Pt nanoelectrode measured using SEM is ∼210 nm. GluOx-modified Pt nanoelectrodes were generated by dip coating GluOx on the Pt nanoelectrode in a solution of 0.9% (wt%) bovine serum albumin (BSA), 0.126% (wt%) glutaraldehyde, and 100 U mL⁻¹ GluOx. An increase in current was observed at +0.7 V vs. Ag/AgCl/1 M KCl with adding increasing concentrations of glutamate. Two-sample t-test results showed that there is a significant difference for current at +0.7 V between the blank and the added lowest glutamate concentration, as well as between adjacent glutamate concentrations, confirming that the increase in current is related to the increased glutamate concentration. The experimental current–concentration curve of glutamate detection fitted well to the theoretical Michaelis–Menten curve. At the low concentration range (50 μM to 200 μM), a linear relationship between the current and glutamate concentration was observed. The Michaelis–Menten constants of I_max and K_m were calculated to be 1.093 pA and 0.227 mM, respectively. Biosensor efficiency (the ratio of glutamate sensitivity to H₂O₂ sensitivity) is calculated to be 57.9%. Enz_act (I_max/H₂O₂ sensitivity, an indicator of the amount of enzyme loaded on the electrode) of the GluOx-modified Pt nanoelectrode is 0.243 mM. We further compared the sensitivity of a GluOx-modified Pt nanoelectrode with a GluOx-modified carbon fiber microelectrode (7 μm diameter and a sensing length of ∼350 μm). Glutamate detection on the GluOx-modified carbon fiber microelectrode fitted well to a Michaelis–Menten like response. Based on the fitting, the GluOx-modified carbon fiber microelectrode exhibited an I_max of 0.689 nA and a K_m of 301.2 μM towards glutamate detection. The best linear range of glutamate detection on the GluOx-modified carbon fiber microelectrode is from 50 μM to 150 μM glutamate. The GluOx-modified carbon fiber microelectrode exhibited a higher potential requirement for glutamate detection compared to the GluOx-modified Pt nanoelectrode.