On-line screening method for antioxidants by liquid chromatography with chemiluminescence detection Original Research Article

A novel screening method for individual antioxidants in complex matrices was developed. The determination of antioxidants is based on the decrease of chemiluminescence (CL) intensity derived from luminol and superoxide anion radical (O−⋅2) which is generated from the enzyme reaction of xanthine oxidase with hypoxanthine. The constant CL intensity, that is recorded as background (baseline) is required for an accurate and precise detection before injection of sample solutions containing antioxidants. Each of the antioxidant separated by the liquid chromatography (LC) column was observed as a negative peak corresponding to its antioxidative potential. Some authentic antioxidants, i.e. ascorbic acid, isoascorbic acid, glutathione, (+)-catechin, (−)-epicatechin, and (−)-epigallocatechin gallate, seemed to possess scavenging activity for O−⋅2. However, the potential was different for each antioxidant and relatively stronger activity was found in the flavonoids. The detection limits (signal-to-noise ratio of two) of (+)-catechin and (−)-epicatechin with on-line LC-CL were 4.4 and 4.3 pmol, respectively.