Antioxidants detection

Many methods including photometric, fluorimetric, chromatographic, and electrochemical methods have been used to detect the antioxidants so far. Recently, electrochemical methods have intensively been used for antioxidant detection. Among the electrochemical methods, the detection of antioxidant based on the direct redox transformation of cyt c has been studied over the decade. Since cyt c can act as an oxidant of superoxide, the superoxide level in solution can be detected as an oxidation current at the sensor electrode due to electron transfer from the radical via cyt c to the electrode. 

The principle of antioxidant detection is shown in Fig. 17.3. Superoxide was enzymatically produced and dismutated spontaneously to oxygen and H202. Under controlled conditions of superoxide generation such as air saturation of the buffer, optimal hypoxanthine concentration (100 pM) and XOD activity (50mU ml-1) a steady-state superoxide level could be obtained for several min (580-680 s). Since these steady-state superoxide concentrations can be detected by the cyt c-modified gold electrode, the antioxidate activity can be quantified from the response of the sensor electrode by the percentage of the current decrease. 

Enhanced chemiluminescent assay Luminol Amersham ECL Antioxidant Detection Pack Luminometric (induction time) W10... 

Flaxseed oils contain much lower amounts of tocopherols, half of the amount present in sunflower and canola oils and one-third of that present in soybean oil (Table 2). A lower content of these antioxidants makes these oils even more susceptible to oxidation. Gamma-tocopherol was found as the main tocopherol in flax oils, with a contribution of about 80% to the total amount. This makes flax oil comparable with soybean oil. Among unique antioxidants detected in flax oils was plasto-chromanol-8. This compound is a derivative of gamma tocotrienol with twice as long unsaturated side chain. Plastochromanol-8 was found to be a more efficient antioxidant than any tocopherols isomer (15). A low content of tocopherols in flaxseed did not make them more susceptible to oxidation experiments showed that milled flaxseed could be stored for 28 months at ambient temperatures without measurable changes in oxidation products. This can be attributed to the presence of antioxidants other than tocopherols in the seeds (16). 

Fig. 69.8 Reversed-phase HPLC of an aqueous infusion of F. deltoidea leaves with absorbance detection at 280 and 365 nm and online ABTS antioxidant detection at 720 nm [87]...

Unlike some of the other common test methods, the XRF technique does not provide any structural information about the antioxidants detected. In addition to the total concentration, chromatographic and infrared based methods can also provide information on the degree of antioxidant degradation products (such as the phosphate content, which is the oxidized form of phosphite antioxidants, normally created during polymer processing). The XRF technique also has limited utility when more than one phosphite or thioester antioxidant is present in the sample, because the total phosphorus or sulfur concentration does not provide the degree of distribution from each antioxidant. 

Gallate (and Other Antioxidant) Detection System Detection Limit Application Reference... 

We have shown that known reaction of luminol with peroxydisulphate at low luminol concentrations takes place in the regime of controlled generation of SO ion-radicals at spontaneous destruction of peroxydisulphate. The detection limit for various types of antioxidants in water using this reaction is varied from 10 to 10 M. It is possible also to determine some polluting admixtures present in the atmosphere. The reagent used is the mixture of the luminol, base and K S O, which, once prepai ed, could be used during a working day. 

In some cases, e.g. the detection of antioxidants [6], the plate is heated to 105 °C for 5 min after being sprayed and the still hot plate placed immediately in an ammonia-vapor chamber. The blue color of the tryptamine derivatives is also stabilized by spraying afterwards with a 5% methanolic ammonia solution [12]. 

The detection limit per chromatogram zone is 50—200ng for lipids [11], 200— 400 ng for antioxidants [3] and several ng for ascorbic acid. 

In the case of carbohydrates blue chromatogram zones are produced on a yellow background that slowly fades [2]. Steroids, vitamins, antioxidants, phenols and aromatic amines yield, sometimes even at room temperature, variously colored chromatogram zones [5]. -Blockers and laxatives also acquire various colors [7, 10]. The detection hmits are in the nanogram to microgram range [5]. 

In relation to consumer uses of possible concern for this CICAD, data from the Women s Environmental Network indicate that butyltin stabilizers have been detected in the non-woven polypropylene topsheet of babies nappies (diapers). It is possible that this could relate to the last of the three key uses described above, in that the topsheet could be of silicone-grafted polypropylene (or, as discussed below, the butyltin may be present because of its use as a catalyst in the production of an antioxidant in polyolefin films). 

The reaction of eq. 16.9 will regenerate the antioxidant Arj-OH at the expense of the antioxidant At2-OH. Despite the fact that such regeneration reactions are not simple electron transfer reactions, the rate of reactions like that of eq. 16.9 has been correlated with the E values for the respective Ar-0. Thermodynamic and kinetic effects have not been clearly separated for such hierarchies, but for a number of flavonoids the following pecking order was established in dimethyl formamid (DMF) by a combination of electrolysis for generating the a-tocopherol and the flavonoid phenoxyl radicals and electron spin resonance (ESR) spectroscopy for detection of these radicals (Jorgensen et al, 1999) ... 

Knowledge of the identity of phenolic compounds in food facilitates the analysis and discussion of potential antioxidant effects. Thus studies of phenolic compounds as antioxidants in food should usually by accompanied by the identification and quantification of the phenols. Reversed-phase HPLC combined with UV-VIS or electrochemical detection is the most common method for quantification of individual flavonoids and phenolic acids in foods (Merken and Beecher, 2000 Mattila and Kumpulainen, 2002), whereas HPLC combined with mass spectrometry has been used for identification of phenolic compounds (Justesen et al, 1998). Normal-phase HPLC combined with mass spectrometry has been used to identify monomeric and dimeric proanthocyanidins (Lazarus et al, 1999). Flavonoids are usually quantified as aglycones by HPLC, and samples containing flavonoid glycosides are therefore hydrolysed before analysis (Nuutila et al, 2002). 

DPPH- has an intense absorption maximum around 520 run (Yordanov and Christova, 1997), and antioxidant capacity and activity measured by the reduction of DPPH- are easily quantified by VIS-spectroscopy (Brand-Williams et al, 1995 Bondet et al, 1997, Espin et al, 2000). The stable radicals Fremy s salt (potassium nitrosodisulphonate) and galvinoxyl (2,6-di-tert-butyl-a-(3,5-di-tert-butyl-4-oxo-2,5-cyclohexadien-l-ylidene)-p-tolyloxy radical) have been used in a similar manner but with ESR detection, which can be used with samples that are not optically transparent (Gardner et al, 1998). 

The on-line measurement of reducing capacity can be performed with either a single or a series of electrochemical detectors, and linear correlations have been demonstrated between total antioxidative activities determined by the electrochemical detection and those determined by DPPH- reduction or by the ORAC assay (Guo et al, 1997 Peyrat-Maillard et al, 2000). The reducing capacity must also be quantified by post-column reactions, either with DPPH- or by the reduction of phosphomolybdenum complexes followed by UV-VIS-detection (Bandoniene and Murkovic, 2002 Cardenosa et al, 2002). A combination of HPLC and semi-automatic ORAC analysis has also been described (Caldwell, 2001). 

CARDENOSA R, MOHAMED R, PINEDA M and AGUILAR M (2002) On-line HPLC detection of tocopherols and other antioxidants through the formation of a phosphomolybdemun complex, /dgricihod Chem, 50, 3390-5. 

---