An enzymatic amplification Flow Injection Analysis (FIA) system for the sensitive determination of phenol

A highly sensitive, enzymatic FIA procedure was developed to determine phenol fluorimetrically. Tyrosinase was immobilized in a packed bed flow reactor. Upon contact with tyrosinase, phenol is oxidized to o-benzoquinone, which oxidizes ascorbic acid to dehydroascorbic acid producing catechol, which is enzymatically reoxidized to o-benzoquinone. Dehydroascorbic acid reacts with o-phenylenediamine forming a highly fluorescent product, which is excited at lambda(exc) = 345 nm and detected at lambda(em) = 410 nm. Dehydroascorbic acid was detected in the range between 0.5 and 100 mu M. The chemoenzymatic substrate recycling enhances the sensitivity of the detection of phenol and catechol with a detection limit of around 0.02 mu M in both cases. Amplification factors between 8 and 12 were estimated. Phenol and catechol can be determined in the approximately linear ranges between 0.1 and 2 mu M and between 0.02 and 2 mu M, respectively. It is possible to perform 20 phenol detections per hour by the automatic FIA procedure with high operational stability. (C) 1998 Elsevier Science S.A. All rights reserved.