Sensitive Electrochemical Assay Research Articles

Assay of enzymatic activity by electrochemical detection of reduced coenzyme

A highly sensitive electrochemical assay of the enzymatic activities of aqueous samples of lactate dehydrogenase, alcohol dehydrogenase, and malate dehydrogenase has been developed using an improved amperometric determination of NADH concentration in the test solution. An anode current sensitivity of 750 μA/mmol of NADH was obtained with a platinum-mesh electrode in an H cell modified to permit vigorous stirring of the anolyte. Fouling of the platinum anode was significantly decreased by working at a pH ≥ 8.1. The rate of increase in net anode current in substrate solutions containing as little as 2 × 10 −3 unit of enzyme/ml correlated well with the rate of change in absorbance at 340 nm for each sample. The reproducibility of the assay of enzyme activity was about ± 10%.