Simultaneous kinetic-based determination of fructose and ascorbate with a rotating bioreactor and amperometric detection: application to the analysis of food samples.

A recently introduced biosensor comprising a rotating bioreactor and a stationary platinum ring amperometric detector (Anal. Chem. 1993, 65, 636-639) has been utilized for the simultaneous determination of fructose and ascorbate. The approach has been successfully applied to the simultaneous determination of these analytes in fresh food samples. Hexacyanoferrate(II) is the monitored species at +0.380 V vs a Ag/AgCl, 3 M NaCl reference. The determination takes advantage of a fast chemical oxidation of ascorbate by hexacyanoferrate(III) ions and the subsequent slower production of hexacyanoferrate(II) in the D-fructose 5-dehydrogenase-catalyzed reaction between D-fructose and hexacyanoferrate(III) as acceptor. D-Fructose 5-dehydrogenase (EC 1.1.99.11) was incorporated into the rotating disk reactor in immobilized form and on controlled-pore glass via the glutaraldehyde attachment and cross-linking with bovine serum albumin. Sample/reagent processing was accomplished by programmed continuous-flow/stopped-flow/continuous-flow operation.