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Abstract: The determination of hydrogen peroxide (H2O2) and the evaluation of scavenging capacity against this species were performed using five colorimetric reactions, which were adapted to flow injection analysis. The reactions chosen were based on the oxidation of iodide (I− method), on the formation of titanium-peroxide complex (TiP method), on the formation of titanium-xylenol orange-peroxide complex (TiXoP method), on the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB method) and on the co-oxidation of phenol-4-sulfonic acid and 4-aminoantipyrine (PSA/4-AAP method). The operational conditions were studied in order to improve the sensitivity of each method. Concerning to the method sensitivity, the ranking order was TMB method>I− method>TiXoP method ∼PSA/4-AAP method>TiP method. All methods showed an excellent repeatability (RSD<2%) and, except for I− method, relative deviations from the reference method were <1.9%. The FIA manifolds were adapted to perform the determination of scavenging capacity against H2O2 and glutathione (GSH) was applied as model compound. TiP and TiXoP methods were not suitable as no inhibition or an increase of analytical signal was attained. PSA/4-AAP method was chosen for further application to dietary phenolics and pharmaceutical compounds, providing IC50 values for those compounds that are fast reacting antioxidants. [Copyright &y& Elsevier]

Abstract: Antioxidant capacity of food samples is usually assessed by different analytical methods, however the results attained even for the same method are strongly dependent on the selected reaction time and also on the standard compound used. To tackle this problem, we propose here a kinetic matching approach, associated to the conversion of results into equivalents of a common standard compound, as a universal way for expression of results. The methodology proposed was applied to methods based on different chemistries (Folin–Ciocalteu (F–C), CUPRAC, DPPH• and ABTS•+ assays) and red wines (n=40) were chosen as a model of complex food sample. For implementation of the kinetic matching approach, the standard phenolic mixture (caffeic acid, (+)-catechin, hesperetin, morin and (−)-epigallocatechin gallate) was chosen for calibration in F–C, CUPRAC and DPPH• assays, while tannic acid was suitable for ABTS•+ assay. Results showed that, for all methods, there was no statistical difference between results attained by the kinetic matching approach (after <10min of reaction) and that at endpoint conditions (after 60 to 300min). The repeatability and the reproducibility of the kinetic matching approach was <4.5%, for all antioxidant assays. The sample throughput increases from <18 (endpoint measurements) to >108h−1 using the proposed kinetic approach. Moreover, we have established here a way of converting results to equivalents of a common standard, providing values independent of its kinetic profile, by using the ratio between calibration sensitivities performed at endpoint conditions. [Copyright &y& Elsevier]

Abstract: In the present work, an automatic method based on multisyringe flow injection analysis (MSFIA) was developed for determination of scavenging capacity against H2O2 using a non-enzymatic fluorimetric assay for H2O2 detection based on the formation of europium-tetracycline-H2O2 complex. The MSFIA-fluorimetric methodology was developed to perform in-line the scavenging reaction of H2O2 prior to reaction of the remaining H2O2 with the fluorescent probe, using conditions close to those found in vivo regarding pH (6.9), temperature (37°C) and H2O2 concentration (25μM). This approach allowed the evaluation of scavenging capacity against H2O2 in a non-competitive (antioxidant+H2O2) or a competitive (antioxidant+H2O2 +biological target) scheme. Using the first strategy, IC50 values determined for the antioxidant compounds glutathione reduced (1191±46μM) and pyruvate (446±49μM) were lower than those obtained for biological targets such as cysteine (2616±182μM), taurine (359±38mM) and adenine (2224±214μM), indicating that reactivity towards H2O2 was higher for antioxidant compounds than for biological targets. However, when a competitive scheme was applied, the scavenging effectiveness against H2O2 depended on the biological molecule present, showing that antioxidant assessment should also take into consideration the concomitant reactivity of biological molecules or structures that are prone to oxidative damage. [Copyright &y& Elsevier]

Abstract: Redox reactions are the heart of numerous biochemical pathways found in cellular chemistry, generating reactive oxygen species (ROS) and reactive nitrogen species (RNS), that includes superoxide anion radical (O2 −), hydrogen peroxide (H2O2), hydroxyl radical (HOt oxygen (1O2), hypochlorite anion (OCl−), peroxynitrite anion (ONOO−) and nitric oxide radical (NOasurement of scavenging capacity against these reactive species presents new challenges, which can be met by flow injection analysis (FIA). In the present review several methods based on FIA and also on its predecessors computer-controlled techniques (sequential injection analysis, multisyringe flow injection analysis, multicommutated and multipumping flow systems) are critically discussed. The selectivity and applicability of the methodology, the generation and detection of the target reactive species, the benefits and limitations of automation when compared to batch methods are some of the issues addressed. [Copyright &y& Elsevier]

Abstract: The role and importance of antioxidants in different fields, ranging from physiology to food technology, have become evident in the past years, requiring adequate analytical methodologies. Therefore, the determination of antioxidant capacity as a routine or screening analysis fosters its automation. In this context, several flow injection methods based on scavenging of 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulphonate) radical cation (ABTS +) or 2,2-diphenyl-1-picrylhydrazyl radical (DDPHd on the determination of total reducing capacity have been proposed. The objective of the present review is to critically compare the different approaches, regarding their degree of automation, their performance vs. the respective batch procedure and its applicability to real samples. [Copyright &y& Elsevier]