Rapid measurement of transaminase activities using an amperometric l-glutamate-sensing electrode based on a glutamate oxidase–polyion complex-bilayer membrane

Abstract

An amperometric l-glutamate-sensing electrode was prepared by immobilizing glutamate oxidase (GlOx) on a polyion complex layer-modified electrode. First, a monolayer of 3-mercaptopropionic acid was made on the surface of a gold electrode by immersing it in an ethanol solution containing the modifier. Next, aqueous solutions of poly-l-lysine and poly(4-styrenesulfonate) were successively placed on the electrode surface and allowed to dry. Finally, a GlOx layer was formed on the poly-l-lysine/poly(4-styrenesulfonate)-complex layer by crosslinking the enzyme by the addition of a glutaraldehyde solution. The use of thin bilayer system with the inner, polyion complex membrane, which showed permselectivity based on the solute size with the molecular cut-off of ≈100, brought high performance characteristics to the l-glutamate-sensing electrode; it showed high sensitivity (detection limit, 20 nM), rapid response (100% response time, 3 s), low interferential level (the ratio of response for l-ascorbic acid to that for the same concentration of l-glutamic acid, 8×10−2), and high stability (usable for more than a month). The bilayer-based electrode was useful for the rapid measurement of glutamate–oxaloacetate transaminase (GOT) and glutamate–pyruvate transaminase (GPT) in serum sample: each transaminase (0.2–1000 U l−1) could be determined within 10 s.