Abstract
A micro-flow enzyme system with a microdialysis probe is proposed for the amperometric detection of trace amounts of neurotransmitter L-glutamate released 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 sensitivity of L-glutamate as an on-line amplifier based on substrate recycling. 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-ascorbate, the adsorption of low molecular-weight proteins in a dialysate, and NADPH added to the carrier solution to initiate substrate recycling. By the 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 µmol dm–3. This method was applied to an in 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 KCl to demonstrate the reliability of the system.
Similar content being viewed by others
References
C. E. Jahr and R. A. Lester, Curr. Opin. Neurobial., 1992, 2, 270.
F. Dagani and E. D’Angelo, Funct. Neurol., 1992, 7, 315.
Y. Hu, K. M. Mitchell, F. N. Albahadily, E. K. Michaelis, and G. S. Wilson, Brain Res., 1994, 659, 117.
J. M. Cooper, P. L. Foreman, A. Glidle, T. W. Ling, and D. J. Pritchard, J. Electroanal. Chem., 1995, 388, 143.
W. G. Kuhr, V. L. Barrett, M. R. Gagnon, P. Hopper, and P. Pantano, Anal. Chem., 1993, 65, 617.
M. R. Ryan, J. P. Lowry, and R. D. O’Neill, Analyst, 1997, 122, 1419.
S. Cosnier, C. Innocent, L. Allien, S. Poitry, and M. Tsacopoulos, Anal. Chem., 1997, 69, 968.
T. Yao, S. Suzuki, H. Nishino, and T. Nakahara, Electroanalysis, 1995, 7, 1114.
D. Moscone, M. Pasini, and M. Mascini, Talanta, 1992, 39, 1039.
E. Zilkha, A. Koshy, T. P. Obrenovitch, H. P. Bennetto, and L. Symon, Anal. Lett., 1994, 27, 453.
O. Niwa, K. Torimitsu, M. Morita, P. Osborne, and K. Yamamoto, Anal. Chem., 1996, 68, 1865.
O. Niwa, T. Horiuchi, and K. Torimitsu, Biosensors Bioelectronics, 1997, 12, 311.
M. O. Berners, M. G. Boutelle, and M. Fillenz, Anal. Chem., 1994, 66, 2017.
A. S. Hazell, R. F. Butterworth, and A. M. Hakim, J. Neurochem., 1993, 61, 1153.
T. J. O’Shea, P. L. Weber, B. P. Bammel, C. E. Lunte, and M. R. Smyth, J. Chromatogr., 1992, 608, 189.
T. Yao, M. Satomura, and T. Nakahara, Electroanalysis, 1995, 7, 395.
S. C. Hartman, J. Biol. Chem., 1968, 243, 853.
J. S. Holcenberg, L. Ericsson, and J. Roberts, Biochemistry, 1978, 17, 411.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Yao, T., Nanjyo, Y. & Nishino, H. Micro-flow in vivo Analysis of l-Glutamate with an On-line Enzyme Amplifier Based on Substrate Recycling. ANAL. SCI. 17, 703–708 (2001). https://doi.org/10.2116/analsci.17.703
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.2116/analsci.17.703