Amperometric determination of acetic acid with a trienzyme/poly(dimethylsiloxane)-bilayer-based sensor.

A trienzyme sensor for the amperometric determination of acetic acid was prepared by immobilizing acetate kinase (AK), pyruvate kinase (PK), and pyruvate oxidase (PyOx) on a poly(dimethylsiloxane) (PDMS)-coated electrode. AK catalyzes the phospho-transferring reaction between acetic acid and ATP to form ADP; PK, the phospho-transferring reaction between ADP and phosphoenolpyruvate to form pyruvic acid; and PyOx, the oxidation of pyruvic acid with oxygen. The oxygen consumption could be monitored by using the PDMS-coated electrode without interference from the PyOx reaction product, hydrogen peroxide. Thus, the concentration of acetic acid (5 microM-0.5 mM) could be determined from the decrease in the cathodic current at -0.4 V vs Ag/AgCl. This is the first example of a biosensor that can be used for the determination of acetic acid in ethanol-containing food samples. The acetate-sensing electrode could be used for more than one month.