Application of a Microplate-Based ORAC-Pyrogallol Red Assay for the Estimation of Antioxidant Capacity: First Action 2012.03

Abstract

A method was developed for microplate-based oxygen radicals absorbance capacity (ORAC) using pyrogallol red (PGR) as probe (ORAC-PGR). The method was evaluated for linearity, precision, and accuracy. In addition, the antioxidant capacity of commercial beverages, such as wines, fruit juices, and iced teas, was measured. Linearity of the area under the curve (AUC) versus Trolox concentration plots was [AUC = (845 +/- 110) + (23 +/- 2) [Trolox, microM]; R = 0.9961, n = 19]. Analyses showed better precision and accuracy at the highest Trolox concentration (40 microM) with RSD and recovery (REC) values of 1.7 and 101.0%, respectively. The method also showed good linearity for red wine [AUC = (787 +/- 77) + (690 +/- 60) [red wine, microL/mL]; R = 0.9926, n = 17], precision and accuracy with RSD values from 1.4 to 8.3%, and REC values that ranged from 89.7 to 103.8%. Red wines showed higher ORAC-PGR values than white wines, while the ORAC-PGR index of fruit juices and iced teas presented a wide range of results, from 0.6 to 21.6 mM of Trolox equivalents. Product-to-product variability was also observed for juices of the same fruit, showing the differences between brands on the ORAC-PGR index.

Antioxidant Activity/Capacity Measurement. 2. Hydrogen Atom Transfer (HAT)-Based, Mixed-Mode (Electron Transfer (ET)/HAT), and Lipid Peroxidation Assays

“…Automation indeed reduces analysis time and improves the efficiency of the assay in multiple analyses. Another method was developed for microplate-based ORAC using pyrogallol red (PGR) as probe with good linearity, precision, and accuracy . However, there are no published data on a manual ORAC assay.…”

“…Another method was developed for microplate-based ORAC using pyrogallol red (PGR) as probe with good linearity, precision, and accuracy. 26 However, there are no published data on a manual ORAC assay. The BIOOXYTECH AOP-490 microplate assay based on the reduction of Cu(II) to Cu(I) by the antioxidants in the oxygenated sample resulting in DNA cleavage was developed 27 and has not been directly compared to the original ORAC method.…”

Measurement of antioxidant activity

“…Even though use of c-phycocyanin probe resulted in higher ORAC values than fluorescein in general, the relative ORAC values and trend among different samples were similar for both probes. Ortiz et al (2012) investigated the use of pyrogallol red probe for ORAC assay, which showed good linearity, precision and accuracy. Guclu, Kibrislioglu, Ozyurek, and Apak (2014) reported ORAC values of liver and kidney samples measured using p-aminobenzoic acid (PABA) as the fluorescent probe.…”

Antioxidant Capacity of Anthocyanin Pigments

“…Pyrogallol reacts faster than fluorescein with RCOO* radicals, and its consumption does not present induction times, even in the presence of very reactive oxidants, with the exception of ascorbic acid. First action ORAC assay has been reported both with fluorescein [226] (dextracts from tea, blueberry and grape skins) and pyrogallol red [228] (red wine, fruit juices and iced teas).…”