Immobilization of glutamate dehydrogenase on glass derivatives. A method for the assay of glutamates in real samples with simplex optimized automated FIA-system.
An enzymatic method for the determination of glutamic acid in food samples and pharmaceuticals is described. l-Glutamate dehydrogenase (GLDH) from beef liver was immobilized on isothiocyanate modified Controlled Pore Glass for the construction of a packed bed reactor. The NADH produced from the enzymic reaction was monitored fluorimetrically. The working curve is linear up to 200 mumol/l glutamate for an injection volume of 58 mul. The detection limit of the method is 0.3 mumol/l. The composite modified simplex was employed for the selection of the proper experimental conditions using an in-house flow injection manifold. Many interfering species and several amino acids were tested to verify the specificity of the enzyme reactor. The system works selectively for glutamic acid. The method is ideally suited to the assay of glutamic acid in a large number of samples because of its simplicity, stability and low cost. Forty-five samples per hour can be analyzed with a relative standard deviation better than 2%. The reactor is stable for a period of more than four months under specified storage conditions. The accuracy of the proposed method is tested by comparison of the results with those obtained by the official methods and the manufacturer's specifications for the analyzed samples. Good correlation was attained. Recovery experiments showed results between 97 and 104%.
Stalikas
C D
CD
Department of Chemistry, University of Ioannina, Ioannina 45110, Greece.
Karayannis
M I
MI
Tzouwara-Karayanni
S
S