High throughput flow injection bioluminometric method for olive oil antioxidant capacity

Abstract

This paper describes a rapid flow injection automated method for the determination of olive oil total antioxidant capacity. The chemistry involved is the horseradish peroxidase (HRP) catalysed oxidation of luminol by hydrogen peroxide. Oxidation results in light emission (bioluminescence) that is enhanced using p-iodophenol sensitizer. Olive oil (0.7mL) is extracted with two 0.7mL aliquots of 80–20% (v/v) methanol–water solvent. A 17μL aliquot of the extract containing hydrophilic antioxidants is injected in a phosphate buffer channel that subsequently merges with a luminol–HRP–p-iodophenol reagent stream. Bioluminescence resulting after merging the mixture with a hydrogen peroxide stream is suppressed upon increasing antioxidants’ concentration resulting in negative peaks due to hydrogen peroxide consumption by antioxidants. The method has been optimized on (a) number of manifold channels, (b) flow rates, (c) coil length and (d) HRP, hydrogen peroxide and p-iodophenol concentrations. Detection limit is calculated at 1.5×10−7M gallic acid, linear range is between 1.0×10−6 and 1×10−4M and precision is better than 2.8% RSD (n=4). The fully automated method is achieving a rate of sampling equal 180 probes per hour. The proposed method is applied for the assessment of 50 extra-virgin olive oil samples of different Greek cultivars and regions.