Potentiometric biosensor for l-ascorbic acid based on ascorbate oxidase of natural source immobilized on ethylene–vinylacetate membrane Original Research Article

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 occlusion in a poly(ethylene-co-vinyl acetate) matrix (EVA 40% m/m). The electrode showed a sub-Nerntian response (slope of 50.3±0.6 mV), between 8.0×10−6 and 4.5×10−4 mol l−1 for ascorbate, when 0.1 mol l−1 KH2PO4 buffer at pH 5.0 and 25.0°C was employed. The biosensor was continuously tested, during 15 days (about 300 analyses), and no decrease in the response was observed, for vitamin C determination in pharmaceuticals samples, without any previous treatment. The biosensor was stable for at least 1 month when stored in a refrigerator. The relative standard deviation was about 4% compared to the spectrophotometric method. The response time was about 5 min at room temperature.