Control of the Oxygen Dependence of an Implantable Polymer/Enzyme Composite Biosensor for Glutamate

Abstract

Biosensors for glutamate (Glu) were fabricated from Teflon-coated Pt wire (cylinders and disks), modified with the enzyme glutamate oxidase (GluOx) and electrosynthesized polymer PPD, poly(o-phenylenediamine). The polymer/enzyme layer was deposited in two configurations: enzyme before polymer (GluOx/PPD) and enzyme after polymer (PPD/GluOx). These four biosensor designs were characterized in terms of response time, limit of detection, Michaelis-Menten parameters for Glu (J max and K M (Glu)), sensitivity to Glu in the linear response region, and dependence on oxygen concentration, K M (O 2 ). Analysis showed that the two polymer/enzyme configurations behaved similarly on both cylinders and disks. Although the two geometries showed different behaviors, these differences could be explained in terms of higher enzyme loading density on the disks; in many analyses, the four designs behaved like a single population with a range of GluOx loading. Enzyme loading was the key to controlling the K M (O 2 ) values of these first generation biosensors. The counterintuitive, and beneficial, behavior that biosensors with higher GluOx loading displayed a lower oxygen dependence was explained in terms of the effects of enzyme loading on the affinity of GluOx for its anionic substrate. Some differences between the properties of surface immobilized GluOx and glucose oxidase are highlighted.L-Glutamate (Glu) is the most widespread excitatory neurotransmitter in the mammalian central nervous system, 1 plays a major role in a broad range of brain functions, and has been implicated in a number of neurological disorders. 2 Indeed, the development of devices for Glu detection has become an important research area due to the value of monitoring this key amino acid in a number of complex matrixes, including food processing, 3,4 cell cultures, 5-7 tissue slices ex vivo, 8,9 and intact brain in vivo. [10][11][12][13][14][15] A number of designs of biosensors for Glu monitoring have been reported and, although Glu receptors 16 Brain Res. Bull. 2002, 58, 401-404. (10) Burmeister, J. J.; Gerhardt, G. A. Trends Anal. Chem. 2003, 22, 498-502. (11)

Contributions by a Novel Edge Effect to the Permselectivity of an Electrosynthesized Polymer for Microbiosensor Applications

“…These implantable biosensors must also be able to reject electrochemical interference by reducing agents present in the tissue, and a common strategy is the incorporation of a permselective membrane on the biosensor surface, such as electropolymerized PPD. 43,[75][76][77][78][79][80] This polymer is unusual 81 in that it displays very high permeability to the biosensor signal transduction molecule (H 2 O 2 , see…”

Ascorbic Acid Rejection Characteristics of Modified Platinum Electrodes: A Shelf Life Investigation

“…This is in stark contrast to the PPD layer grown on bare Pt which can be controlled more efficiently due to the self-limiting characteristic associated with the polymer during the electropolymerisation process. A number of groups report the thickness of PPD polymers to be between 10 and 30 nm [12,40,41]. Similarly, Nafion ® has been electrodeposited onto carbon electrode surfaces as a method of incorporating a uniform reproducible layer to ameliorate issues associated with dip coating and annealing [3,32,42].…”

Oxygen tolerance of an implantable polymer/enzyme composite glutamate biosensor displaying polycation-enhanced substrate sensitivity

“…The biosensor response reached a plateau at oxygen levels which depended on the concentration of Glu in the cell, but typically by 30-50 M O 2 . Non-linear regression analysis of the current was performed up to this plateau region to determine the relevant K M (O 2 ) value (McMahon et al, 2006a; see Section 2.3).…”