Assays for Hydrophilic and Lipophilic Antioxidant Capacity (oxygen radical absorbance capacity (ORACFL)) of Plasma and Other Biological and Food Samples

• Published in: Journal of agricultural and food chemistry Vol. 51; no. 11; pp. 3273 - 3279
• Main Authors: Prior, Ronald L, Hoang, Ha, Gu, Liwei, Wu, Xianli, Bacchiocca, Mara, Howard, Luke, Hampsch-Woodill, Maureen, Huang, Dejian, Ou, Boxin, Jacob, Robert
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 21.05.2003
• Subjects: absorption; Adult; Aged; Amidines - chemistry; antioxidant activity; antioxidants; Antioxidants - analysis; Antioxidants - chemistry; apple juice; apples; assays; Beverages - analysis; blood serum; blueberries; Blueberry Plants - chemistry; Chemical Phenomena; Chemistry, Physical; colored varieties; cranberries; Diet; dried fruit; ethanol; extraction; Female; Food Analysis; free radical scavengers; free radicals; Fruit - chemistry; fruit juices; grape juice; hexane; Humans; hydrophilic interactions; hydrophobicity; methodology; Middle Aged; orange juice; oranges; Oxidation-Reduction; phenolic compounds; Plasma - chemistry; plums; Reactive Oxygen Species - chemistry; Reproducibility of Results; Sensitivity and Specificity; strawberries; water
• ISSN: 0021-8561; 1520-5118; 1520-5118
• DOI: 10.1021/jf0262256

Methods are described for the extraction and analysis of hydrophilic and lipophilic antioxidants, using modifications of the oxygen radical absorbing capacity (ORACFL) procedure. These methods provide, for the first time, the ability to obtain a measure of “total antioxidant capacity” in the protein free plasma, using the same peroxyl radical generator for both lipophilic and hydrophilic antioxidants. Separation of the lipophilic and hydrophilic antioxidant fractions from plasma was accomplished by extracting with hexane after adding water and ethanol to the plasma (hexane/plasma/ethanol/water, 4:1:2:1, v/v). Lipophilic and hydrophilic antioxidants were efficiently partitioned between hexane and aqueous solvents. Conditions for controlling temperature effects and decreasing assay variability using fluorescein as the fluorescent probe were validated in different laboratories. Incubation (37 °C for at least 30 min) of the buffer to which AAPH was dissolved was critical in decreasing assay variability. Lipophilic antioxidants represented 33.1 ± 1.5 and 38.2 ± 1.9% of the total antioxidant capacity of the protein free plasma in two independent studies of 6 and 10 subjects, respectively. Methods are described for application of the assay techniques to other types of biological and food samples. Keywords: ORACFL; phenolics; antioxidant; free radical; lipophilic; hydrophilic; blueberry; fruit juices