T. Yao et al., Micro-flow in vivo analysis of L-glutamate with an on-line enzyme amplifier based on substrate recycling, ANAL SCI, 17(6), 2001, pp. 703-708
A micro-flow enzyme system with a microdialysis probe is proposed for the a
mperometric detection of trace amounts of neurotransmitter L-glutamate rele
ased from rat brain cells. The L-glutamate oxidase (EC 1.4.3.11)/glutamate
dehydrogenase (EC 1.4.1.4) coimmobilized reactor was used to enhance the se
nsitivity of L-glutamate as an on-line amplifier based on substrate recycli
ng. A poly(1,2-diaminobenzene) film-coated platinum electrode was also used
to selectively detect only the hydrogen peroxide generated into a upstream
enzyme reactor, without interference from oxidizable species, such as L-as
corbate, the adsorption of low molecular-weight proteins in a dialysate, an
d NADPH added to the carrier solution to initiate substrate recycling. By t
he present in vivo system, L-glutamate was selectively assayed with about a
600-fold increase in sensitivity compared with the unamplified responses.
The detection limit was 0.08 mu mol dm(-3). This method was applied to an i
n vivo assay of L-glutamate in the extracellular space of rat brain; also,
monitoring of the L-glutamate level changed after a continuous stimulation
of KCI to demonstrate the reliability of the system.