Multi-enzyme electrodes for the determination of starch by flow injection

Abstract

A tri-enzyme electrode has been developed for determining starch in a flowing stream based on amperometric monitoring of hydrogen peroxide at a potential of +600 mV versus a silver-sliver chloride reference electrode. The nylon-based starch-sensing membranes (over a platinum electrode) were prepared from an enzyme cocktail containing various ratios of amyloglucosidase (AMG), mutarotase (MUT) and glucose oxidase (GO). The best starch-sensing membrane (to give the type A electrode) was made from an enzyme cocktail of AMG-MUT-GO (2000 + 100 + 100 U; where 1 U = 16.67 nanokatals), i.e., containing a high level of AMG. In this system, starch samples were first incubated with soluble α-amylase (α-AMY)(1000 U) for 1 h at room temperature prior to analysis with the tri-enzyme electrode. Attempts were also made to immobilise α-AMY on to nylon net, either alone or as a component of a four-membrane starch-enzyme electrode but starch signals were weak compared with those generated by starch pre-treated with soluble α-AMY. This system, associated with the type A starch electrode, not only exhibited a calibration of wide linear range (1 × 10^–4–0.1% m/v starch) but also showed promising operational properties. It has excellent thermal stability over the range 30–70 °C.