title: Determination of total antioxidant capacity by oxygen radical absorbance capacity (ORAC) using fluorescein as the fluorescence probe: First Action 2012.23.
abstract: An improved method for the measurement of oxygen radical absorbance capacity (ORAC) was developed and validated using fluorescein (3',6'-dihydroxyspiro[isobenzofuran-1[3H], 9'[9H]-xanthen]-3-one) as a new fluorescence probe (ORAC(FL)). Randomly methylated beta-cyclodextrin (RMCD) was introduced as the water-solubility enhancer for lipophilic antioxidants. 7% RMCD (w/v) in 50% acetone-H2O mixture sufficiently solubilized vitamin E compounds and other lipophilic phenolic antioxidants in 75 mM phosphate buffer (pH 7.4). Results indicated that fluorescein shows excellent photostability under the plate reader conditions. This ORAC(FL) was validated through linearity, precision, accuracy, and ruggedness. The validation results demonstrated that the ORACFL assay is reliable and robust. The mean of intraday and interday CVs were <15%; for hydrophilic ORAC, LOD and LOQ are 5 and 6.25 microM, respectively; for lipophilic ORAC, LOD and LOQ are 6.25 and 12.5 microM, respectively. It is concluded that unlike other popular methods, the ORAC(FL) assay provides a direct measure of total antioxidant capacity against the peroxyl radicals.
authors.forename: Boxin
authors.initials: B
authors.lastname: Ou
authors.forename: Tony
authors.initials: T
authors.lastname: Chang
authors.forename: Dejian
authors.initials: D
authors.lastname: Huang
authors.forename: Ronald L
authors.initials: RL
authors.lastname: Prior
mesh_terms: D000096:Acetone / Q000737:chemistry; D000975:Antioxidants / Q000737:chemistry*; D019540:Area Under Curve; D002021:Buffers; D002839:Chromans / Q000737:chemistry; D019793:Fluorescein / Q000737:chemistry*; D005452:Fluoresceins / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D005609:Free Radicals / Q000737:chemistry; D006861:Hydrogen Peroxide / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000737:chemistry; D010100:Oxygen / Q000737:chemistry*; D010545:Peroxides / Q000737:chemistry; D017382:Reactive Oxygen Species / Q000737:chemistry; D015203:Reproducibility of Results; D012995:Solubility; D014867:Water / Q000737:chemistry; D047392:beta-Cyclodextrins / Q000737:chemistry
chemical_list: D000975:Antioxidants; D002021:Buffers; D002839:Chromans; C045076:Fenton's reagent; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005609:Free Radicals; D010545:Peroxides; D017382:Reactive Oxygen Species; D047392:beta-Cyclodextrins; C587846:fluorescein (3',6'-dihydroxyspiro(isobenzofuran-1(3H), 9'(9H)-xanthen)-3-one); D014867:Water; D000096:Acetone; C049375:perhydroxyl radical; D006861:Hydrogen Peroxide; D007501:Iron; C031215:betadex; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid; D010100:Oxygen; D019793:Fluorescein