Graphite-Teflon composite bienzyme electrodes for the determination of l-lactate: Application to food samples

Abstract

A bienzyme amperometric graphite-Teflon composite biosensor, in which lactate oxidase (LOD) and peroxidase, together with the mediator ferrocene, are incorporated into the electrode matrix, was developed for the determination of l-lactate in food samples such as wine and yogurt by using both batch- and flow-injection modes. This bienzyme electrode was fabricated by simple physical inclusion of the enzymes and the mediator in the bulk of the graphite-Teflon matrix. A Teflon content of 70%, an applied potential of 0.00 V, and a pH of 7.4 were employed as working conditions. The composite bioelectrode exhibited long-term operation because of the renewability of its surface by polishing. Reproducible amperometric responses were achieved with different electrodes fabricated from different composite matrices, and no significant loss of the enzyme activity occurred after 6 months of storage at 4°C. Detection limits for l-lactate of 1.4 and 0.9 μM were obtained by batch amperometry in stirred solutions and flow-injection with amperometric detection, respectively. An interferences study with different substances which may be present in wine and yogurt together with l-lactic acid demonstrated very good selectivity for the determination of this analyte. The bienzyme composite electrode was applied to the determination of l-lactic acid in red wine and shaken yogurt, and the methods were validated by comparing these results with those obtained by applying a recommended reference method.