Potentiometric biosensor for L-ascorbic acid based on ascorbate oxidase ofnatural source immobilized on ethylene-vinylacetate membrane

The development of a potentiometric biosensor for L-ascorbic acid based on ascorbate oxidase is described. The enzyme was extracted of the epicarp of Cucumis sativus L. fruit and immobilized in a graphite/epoxy electrode by o cclusion in a poly(ethylene-co-vinyl acetate) matrix (EVA 40% m/m). The ele ctrode showed a sub-Nerntian response (slope of 50.3+/-0.6 mV), between 8.0 x10(-6) and 4.5x10(-4) mol l(-1) for ascorbate, when 0.1 mol l(-1) KH2PO4 b uffer at pH 5.0 and 25.0 degrees C was employed. The biosensor was continuo usly tested, during 15 days (about 300 analyses), and no decrease in the re sponse was observed, for vitamin C determination in pharmaceuticals samples , without any previous treatment. The biosensor was stable for at least 1 m onth when stored in a refrigerator. The relative standard deviation was abo ut 4% compared to the spectrophotometric method. The response time was abou t 5 min at room temperature. (C) 1999 Elsevier Science B.V. All rights rese rved.