Abstract: A simple, rapid continuous-flow method with gas chromatography detection for the simultaneous determination of natural and synthetic antioxidants (α-tocopherol, α-tocopheryl acetate, 2,6-di-tert-butyl-p-hydroxytoluene, tert-butyl-4-hydroxyanisole and tert-Bu hydroquinone) is proposed. A solid phase extractor is coupled to the flow system to isolate antioxidants from the sample matrix. During extraction, analytes are released from ~70% of the triglycerides present in fats and oils by using XAD-7 sorbent; after selective elution with 400 µL of 2-propanol, only ~0.7-0.8% of total triglycerides originally present in the sample remain in the final ext. XAD-7 adsorbs 70-95% of the antioxidants except BHT of which only ~30% is adsorbed. The proposed method is considered an effective alternative to earlier procedures.

Abstract: Cyclic voltammetry has been used to quantify antioxidants at a carbon electrode in white wines diluted to 10% and red wines diluted to 0.25% in a model wine solution (12% ethanol, 0.033 M tartaric acid adjusted to pH 3.6). The first peak near 400 mV (versus Ag/AgCl) was due to phenolic compounds with an ortho-diphenol group or to gallic acid, while a small feature at 300 mV was seen in red wines containing higher levels of myricetins (triphenol on the flavonoid B-ring), and a peak or shoulder at 470 mV was ascribed to quercetin glycosides. The integral of the current to 500 mV was then used as a measure of the lower oxidation potential phenolics, reported as gallic acid equivalents, and compared to traditional measures of total phenols in wines. A peak at 640 mV in red wines was associated with malvidin anthocyanins and was smaller in an older red wine sample, while further oxidation current beyond 700 mV prominent in white wines was due to meta-diphenol or isolated phenol groups and other oxidizable compounds. Simulated voltammograms constructed from high-pressure liquid chromatography results for six monomeric phenolics were in good agreement with the experimental voltammograms for the white wines, while a higher proportion of the current for the red wine samples was due to oligomeric phenolics.

Abstract: In order to evaluate the actual antioxidant features of foods, beverages and also plasma from patients, a number of assays have been developed in the last few years to determine the so called total antioxidant activity (TAA), intended as the cumulative capacity of a biological sample to scavenge free radicals. Most of the assays partially failed in obtaining a good reproducibility when using plasma because it is composed of a large number of substances, some of which are present at very high concentrations and possess masking features. For these reasons we have improved the widely known ABTS method by means of a FIA system where both temperature and dispersion of sample and reagent were strictly controlled. We found that temperature may be a critical aspect in the measurement of plasma TAA whilst its influence may be less important in the assay of non-complex biological samples. We demonstrated that also the reaction time may be critical, depending on the nature of the substance employed. Data confirmed the high TAA of a methylsalicylate-containing mouthrinse as well as the negligible TAA offered by the chlorhexidine containing one. White wines (Verdicchio) also displayed interesting TAA values. The improved method was useful to screen rapidly, without dilution, with very limited handling of the sample and with high repeatability the TAA of plasma in addition to chemical products, beverages and non-complex biological mixtures.

Abstract: Antioxidant activity and/or the inhibition of lipoxygenases may contribute to the cell and tissue protective properties of flavonoids. Soybean lipoxygenase-1 (LOX-1), is a useful paradigm for lipoxygenases isolated from different sources, and can be used to devise assays of oxygenase inhibitors and antioxidants. Here we describe the voltammetric detection of methylene blue (MB) to assay for antioxidant and lipoxygenase inhibition activity. MB is formed by the catalytic action of hydroperoxide, produced by soybean lipoxygenase reaction on linoleic acid, on benzoyl-leuco methylene blue (BLMB). Every compound which blocks either the enzymatic reaction, lipoxygenase inhibitors, or the oxidation of BLMB, antioxidants, can thus be detected using this method. The assay involves reaction for a period of one hour of linoleic acid, lipoxygenase, BLMB and inhibitor, followed by injection into a flow-injection system and detection by square-wave voltammetry of the methylene blue produced. The problem of irreproducibility induced by adsorption at the solid platinum working electrode is alleviated using the method of alternate injections of control and assay solutions. Assays of several known lipoxygenase inhibitors were undertaken. All of these inhibitors were also, however, shown to be antioxidants. Extension of the assay to the rapid evaluation of novel therapeutic antioxidants and lipoxygenase inhibitors is proposed.