High-Throughput Methods To Assess Lipophilic and Hydrophilic Antioxidant Capacity of Food Extracts in Vitro

Abstract

Assays comprising three probes for different mechanisms of antioxidant activity in food products have been modified to allow better comparison of the contributions of the different mechanisms to antioxidant capacity (AOC). Incorporation of a common format for oxygen radical absorbance capacity (ORAC), ferric reducing antioxidant power (FRAP), and iron(II) chelating activity (ICA) assays using 96-well microplates provides a comprehensive and high-throughput assessment of the antioxidant capacity of food extracts. The methods have been optimized for aqueous extracts and validated in terms of limit of quantification (LoQ), linearity, and precision (repeatability and intermediate reproducibility). In addition, FRAP and ORAC assays have been validated to assess AOC for lipophilic extracts. The relative standard deviation of repeatability of the methods ranges from 1.2 to 6.9%, which is generally considered to be acceptable for analytical measurement of AOC by in vitro methods. Radical scavenging capacity, reducing capacity, and iron chelating properties of olive mill wastewaters (OMWW), oregano, and parsley were assessed using the validated methods. OMWW showed the highest radical scavenging and reducing capacities, determined by ORAC and FRAP assays, respectively, followed by oregano and parsley. The ability to chelate Fe (2+) was, in decreasing order of activity ( p > 0.05) parsley congruent with oregano > OMWW. Total phenol content, determined by the Folin-Ciocalteu method, correlated to the radical scavenging and reducing capacities of the samples but not to their chelating properties. Results showed that the optimized high-throughput methods provided a comprehensive and precise determination of the AOC of lipophilic and hydrophilic food extracts in vitro.

Stability of the total antioxidant capacity and total polyphenol content of 23 commercially available vegetable juices before and after in vitro digestion measured by FRAP, DPPH, ABTS and Folin–Ciocalteu methods

“…Where FRAP measures the reducing capacity of all the antioxidants in the mixture, FCR records only those classed as polyphenolic compounds (Huang et al, 2005). As a result previous researchers have often found very good linear correlations between FCR and one or more SET based assays such as FRAP or ABTS •+ (Jimenez-Alvarez et al, 2008;Molan, De & Meagher, 2009;Sreeramulu & Raghunath, 2010;Stratil, Klejdus & Kuban, 2006). The same relationship is observed in the current study, where a very strong positive linear correlation (r 2 = 0.96) is observed between FRAP and FCR.…”

Capacidade antioxidante total de óleos vegetais comestíveis: determinantes químicos e sua relação com a qualidade dos óleos

“…A localização dos antioxidantes em interfaces causa um fenômeno conhecido como paradoxo polar, no qual os antioxidantes polares são mais efetivos em meios apolares e vice-versa 13 . Consistentemente com essas complexas interações moleculares, Espín et al 14 6 tenham sugerido que apenas os ensaios de TEAC e ORAC são passíveis de adaptação para os compostos lipofílicos, houve adaptações de protocolos para outros ensaios 14,19 . No Quadro 1, destacam-se algumas das principais características e diferenças entre os ensaios mais utilizados para a determinação da capacidade antioxidante de óleos vegetais.…”

“…Recentemente, os resultados desse ensaio têm sido apresentados como equivalentes de trolox 10 . Devido aos potenciais redox comparáveis do Fe +3 -TPTZ (0,70 V) e do ABTS + (0,68 V), os compostos antioxidantes que reagem no ensaio do FRAP tendem a reagir de forma similar no ensaio da TEAC, gerando resultados que se correlacionam positivamente 19 .…”

“…Recentemente, Jimenez-Alverez et al 19 reportaram o uso de β-ciclodextrina aleatoriamente metilada para dissolver adequadamente a fração apolar de orégano e permitir a reação desse extrato com o reagente do FRAP. Entretanto, não foi possível localizar artigos publicados que reportassem a aplicação dessa abordagem analítica para a determinação da capacidade antioxidante de óleos vegetais.…”

Improvement and Interlaboratory Validation of the Lipophilic Oxygen Radical Absorbance Capacity: Determination of Antioxidant Capacities of Lipophilic Antioxidant Solutions and Food Extracts

“…11 In an earlier study of L-ORAC assays, 12 we revealed that the repeatability and intermediate precision of the original L-ORAC method 13 are inadequate, and thus we developed a modified method that provides satisfactory repeatability and intermediate precision in a single-laboratory validation study. Here, we conducted interlaboratory tests in accordance with an internationally harmonized protocol for the design of method-performance studies 14 to validate this improved L-ORAC method.…”