Simultaneous Determination of Lactic Acid, Glutamic Acid and Glucose by HPLC Using an Enzyme Sensor Using Photocrosslinkable Poly (vinyl alcohol) Support as the Detector

Abstract

Lactate oxidase, glutamate oxidase and glucose oxidase were co-immobilized in a membrane through the photoreaction a poly (vinyl alcohol) bearing styrylpiridinium group (PVA-SbQ). The co-immobilized enzymes membrane was placed on the Pt anode of an electrochemical detector. The enzymes membrane sensor thus prepared was incorporated with analytical column in HPLC. Lactic acid, glutamic acid and glucose in the sample employed reacted with lactate oxidase, glutamate oxidase and glucose ox-id ase, respectively, to produce hydrogen peroxide, and the product was monitored electrochemically. Th e sensitivity of this detection system was nearly 100-fold greater than when using differential refrac tive index detection. The lower detection limit of each analyte was ca.70 ng (S/N= 3). The use of specific enzymatic reaction and electrochemical detector could simplify the pretreatments of the sample used. This method was successfully applied for the determination of lactic acid, glutamic acid and glucose in wine, sake, sour milk and sarad dressing.