Immobilization of glutamate oxidase on non-porous glass beads. Automated flow injection system for the assay of glutamic acid in food samples and pharmaceuticals

Abstract

An enzymic method is proposed for the determination of glutamic acid in food samples and pharmaceuticals. The method is useful in the range 0.01–0.5 mmol l^–1, with a detection limit of 0.005 mmol l^–1 for an injection volume of 52 µl. L-Glutamate oxidase from Streptomyces sp. X-119-6 was immobilized on non-porous glass beads, and the H₂O₂ produced was caused to react with Trinder's reagent. Various parameters were studied for the establishment of the optimum operating conditions for an in-house flow injection manifold with a spectrophotometric detector. Many interfering species and several amino acids were tested in order to verify the specificity of the enzyme reactor. Experimental data for interfering organic and inorganic substances are reported. The system works very selectively for glutamic acid and glutamates. The method is ideally suited to the assay of glutamates in a large number of samples within a single working day, as the frequency is 40 samples h^–1. The relative standard deviation is 1.8% for six assays. The immobilized reactor is stable for a long period under proper conditions of storage. The accuracy of the proposed method was tested by comparison of the results with those of the Association of Official Analytical Chemist's method and with the manufacturer's specifications for the analysed samples, where available. Good agreement was obtained. Recovery studies yielded results between 97 and 105%.