Development of a New Free Radical Absorption Capacity Assay Method for Antioxidants: Aroxyl Radical Absorption Capacity (ARAC)

Abstract

A new free radical absorption capacity assay method is proposed with use of an aroxyl radical (2,6-di-tert-butyl-4-(4'-methoxyphenyl)phenoxyl radical) and stopped-flow spectroscopy and is named the aroxyl radical absorption capacity (ARAC) assay method. The free radical absorption capacity (ARAC value) of each tocopherol was determined through measurement of the radical-scavenging rate constant in ethanol. The ARAC value could also be evaluated through measurement of the half-life of the aroxyl radical during the scavenging reaction. For the estimation of the free radical absorption capacity, the aroxyl radical was more suitable than the DPPH radical, galvinoxyl, and p-nitrophenyl nitronyl nitroxide. The ARAC value in tocopherols showed the same tendency as the free radical absorption capacities reported previously, and the tendency was independent of an oxygen radical participating in the scavenging reaction and of a medium surrounding the tocopherol and oxygen radical. The ARAC value can be directly connected to the free radical-scavenging rate constant, and the ARAC method has the advantage of treating a stable and isolable radical (aroxyl radical) in a user-friendly organic solvent (ethanol). The ARAC method was also successfully applied to a palm oil extract. Accordingly, the ARAC method would be useful in free radical absorption capacity assay of antioxidative reagents and foods.

Kinetic Study of the Scavenging Reaction of the Aroxyl Radical by Eight Kinds of Vegetable Oils in Solution

“…Antioxidant activity of food extracts has been measured using several different methods such as (1) TRAP (total radical‐trapping antioxidant parameter), (2) TEAC (trolox equivalent antioxidant capacity), (3) FRAP (ferric‐ion‐reducing antioxidant power), (4) ORAC (oxygen radical absorption capacity), and (5) SOAC (singlet oxygen absorption capacity) assay methods. Stable free radicals such as (6) DPPH• (2,2‐diphenyl‐1‐picrylhydrazyl), (7) galvinoxyl•, and (8) ABTS + • (2,2′‐azinobis(3‐ethylbenzothiazoline‐6‐sulfonic acid)) radicals are generally used to evaluate free radical scavenging activity of natural AOH and food extracts (Espin, Soler‐Rivas, & Wichers, ; Foti, ; Kawashima, Ohkubo, & Fukuzumi, ; Nagaoka, Nagai, Fujii, Ouchi, & Mukai, ; Naqvi, Li, Melø, & Webster, ).…”

“…Recently, a kinetic study of the scavenging reaction of the 2,6‐di‐ t ‐butyl‐4‐(4‐methoxyphenyl)phenoxyl (abbreviated as aroxyl [ArO•]) radical (Fig. ) with natural AOH, including α‐, β‐, γ‐, and δ‐Tocs and four tocopherol derivatives, was performed in ethanol solution at 25 °C (Nagaoka et al, ). The second‐order rate constants ( k s ) for the reaction of AOH with ArO• were measured using stopped‐flow spectrophotometry (reaction (1)). …”

“…The second‐order rate constants ( k s ( f ) and k s ( t 1/2 )) were determined by analyzing the first‐order rate constant ( f ) and the half‐life ( t 1/2 ) of the decay curve of ArO•, respectively, and were found to be in good agreement with each other. From the results, a new assay method that can quantify the aroxyl radical absorption capacity (ARAC) of phenolic AOH was proposed (Nagaoka et al, ). The ARAC assay method was applied to food extracts.…”

“…The ARAC assay method was applied to food extracts. However, the examples of food extracts investigated have been limited to a palm oil extract (Nagaoka et al, ) and seven rice bran extracts (Mukai et al, ).…”

Tunneling effect in vitamin E recycling by green tea

“…40,41 The ARAC method was an alternative to ORAC method (oxygenradical absorbing capacity method), 42 which was most widely used to assess antioxidant capacities against free radicals but whose validity was regrettably doubted by U.S. Department of Agriculture in 2012. 40,41 The ARAC method was an alternative to ORAC method (oxygenradical absorbing capacity method), 42 which was most widely used to assess antioxidant capacities against free radicals but whose validity was regrettably doubted by U.S. Department of Agriculture in 2012.…”