Immobilization of alliinase and its application: Flow injection enzymatic analysis for alliin

Abstract

Alliinase was immobilized on chitosan microspheres by means of glutaraldehyde, and a flow-injection enzymatic analytical system was developed for determination of alliin based on the immobilized
alliinase and an ammonia gas electrode. The factors affecting the activity of the immobilized enzyme, such as glutaraldehyde concentration, cross-linking time and the amount of alliinase, were investigated. Results showed that the maximum of enzyme activity could be obtained at glutaraldehyde concentration of 4%, cross-linking time of 2 h and the amount of alliinase 20.2 u. The properties of the immobilized alliinase were also studied in detail. For the immobilized alliinase, the highest activity was allowed at pH of 7.0 and temperature at 35°C. Besides, the immobilized enzyme showed good thermal and pH stabilities. The flow-injection enzymatic analytical system based on the immobilized alliinase and an ammonia gas electrode provided linearity in the 1 × 10-5 to 1 × 10-3 mol/L alliin concentration
range and exhibited good repeatability and operational stability.