Measuring the Effectiveness of Antioxidants

Electron Paramagnetic Resonance (EPR, ESR) spectroscopy is a unique tool to answer specific questions evolving radical processes in both research and development (R D) as well as quality control (QC). Key QC examples comprise the food irradiation control for customer care according to European Union (EU) norms and the flavour stability and shelf life assessment on fresh beer. For the first time, an automated DPPH-assay to measure the effectiveness of antioxidants by CW-EPR is reported. 

We have seen earlier that in the presence of an antioxidant the rate of hydroperoxide formation is related to the ratio of lipid concentration to that of the antioxidant concentration, which is dependent on the rate of inhibition reaction (4). The effectiveness of an antioxidant is generally based on the balance between the inhibition rate (k ) of reaction (4) and the transfer reactions ( ), (9) and (13). Therefore, the effect of antioxidants on hydroperoxide decomposition reactions (10) and (11) is an important property that needs to be evaluated. However, most studies of antioxidant actions measure initial events of lipid oxidation based on oxygen absorption, hydroperoxide formation, and peroxide values (Chapters 5 and 7). Very few studies have measured the effect of antioxidants on decomposition products of hydroperoxides, such as aldehydes and carbonyl compounds. Yet these volatile decomposition aldehydes are most relevant to the development of rancidity and to the ultimate quality and stability of food lipids. 

Table 4 The effect of antioxidants on induction time of oxidation of polypropylene determined at 150°C from chemiluminescence measurements...

Despite the threat to the equilibrium of oxidation/antioxidant defenses, OS was never measured in any of the epidemiological studies, but only in a few cases of acute or chronic diseases. Under these circumstances it is hard to draw any valid conclusions on the effect of antioxidants on health status. 

The effects of antioxidants on OT of SME by non-isothermal (conventional) DSC, static mode P-DSC, and dynamic mode P-DSC were investigated by Dunn (2006a), which is summarized in Table 1.15. Results from all three methods consistently showed that treating SME with antioxidants TBHQ and a-tocopherol increased OT with respect to untreated SME. Statistical comparison of P-DSC results with those from isothermal analysis of OSI at 60°C was facilitated by calculation of the corresponding response factors (defined ratios of OT of the sample to that of methyl oleate, and of OSI of the sample to that of methyl oleate). Data for the sample and reference material (methyl oleate) were measured under the same experimental conditions. Results showed the highest degree of correlation (P = 0.79) between dynamic-mode P-DSC and isothermal OSI analyses. 

There are many discrepancies in the literature regarding the amount of antioxidants in healthy subjects and the effects of antioxidants either in preventing LDL oxidation and cardiovascular diseases. Different results are obtained when different methods, with different sensibility and accuracy, are used for measurement of plasma antioxidants, and... 

Direct Measurements—UVNIS Techniques. The conjugated diene (CD) formed among the polyene hydroperoxide products that are formed as a result of oxidation of polyunsaturated fatty acids (PUFAs) have a UV absorbance that can be monitored to follow the progress of the oxidation. The effect of antioxidants on the suppressed rate of product formation can be followed with time. For example, conjugated dienes from oxidation of linoleate lipid molecules absorb at 234 nm and can be monitored directly , or else after HPLC separation (via normal phase or reverse phase j of the individual isomers. In order to use these findings to calculate the antioxidant activity of phenols and relate it to oxygen uptake studies (equations 7 and 14), one also has to make a correction to account for loss of absorbance due to loss (from decomposition) of hydroperoxides (equation 22) . 

Factors Determining the Effectiveness of Antioxidants. It was for many years a source of puzzlement to polymer chemists that the rating of antioxidants in polymers and in model hydrocarbons appeared to be very different. The widely used antioxidant BHT (I) is one of the most "efficient" antioxidants known for liquid hydrocarbons as measured by oxygen absorption but is virtually ineffective in rubbers... 

The intrinsic dielectric losses of pure polyethylene and polypropylene are very small at 4.2 K (i.e., 5 x 10 ) The higher values of tan 6 measured for commercially produced polyolefins are due to the presence of additives placed in the polymer during the manufacturing process to protect the polymer in its intended air environment. Early work by King and Thomas disclosed that the antioxidant may be one of the major sources of dielectric loss at temperatures of 6 to 8 K. A subsequent study of the effects of antioxidant on tan 6, carried out jointly by Battelle Columbus Laboratories (BCL), the National Bureau of Standards (NBS), and BNL, also showed that the 60 Hz loss tangent of polyethylene, in the region of 4 to 10 K, was strongly dependent upon both type and... 

Thermal stability can he determined in DSC by measuring the temperature of reaction or degradation under isothermal or scanned temperature conditions. Other conditions include an inert or an oxidative atmosphere at ambient or elevated temperature. It is also possible to study the effects of increased pressure using pressurized DSC cells. By varying the conditions, thermooxidative processes can be separated from thermodegradative processes. The induction time before reaction begins is taken as a measure of the stability The effect of antioxidants and stabilizers on this time may be readily studied. 

It has been observed that the effectiveness of antioxidants, as measured by OIT at high temperatures, may differ as a function of temperature. OIT test temperatures should preferably be close to actual use temperature. At ambient pressures, significant oxidation is often not detected until the polymer is above the melting point. At these temperatures, essential ingredients in the polymer formulation can be lost. Also, tests performed above the melting point cannot be extrapolated reliably to temperatures below the melting point. Pressure DSC suppresses volatilisation of additives and degradation by-products, an event which thermally competes with the oxidation exotherm. Moreover, the... 

The value of the induction period of the oxidation reaction (Ip) is a measure of antioxidant efficiency. Figure 10.1 shows the general trend of the effect of different antioxidants on oxidation kinetics of polyolefins. Obviously, oxidation progresses in the same manner as for unstabilized polymer when all the antioxidant has been consumed. In many cases, physical properties of the polymer change quite seriously even in the absence of oxidation during the induction period. However, it is necessary to know the effect of antioxidants on physical properties. 

The antioxidant activities of carotenoids and other phytochemicals in the human body can be measured, or at least estimated, by a variety of techniques, in vitro, in vivo or ex vivo (Krinsky, 2001). Many studies describe the use of ex vivo methods to measure the oxidisability of low-density lipoprotein (LDL) particles after dietary intervention with carotene-rich foods. However, the difficulty with this approach is that complex plant foods usually also contain other carotenoids, ascorbate, flavonoids, and other compounds that have antioxidant activity, and it is difficult to attribute the results to any particular class of compounds. One study, in which subjects were given additional fruits and vegetables, demonstrated an increase in the resistance of LDL to oxidation (Hininger et al., 1997), but two other showed no effect (Chopra et al, 1996 van het Hof et al., 1999). These differing outcomes may have been due to systematic differences in the experimental protocols or in the populations studied (Krinsky, 2001), but the results do indicate the complexity of the problem, and the hazards of generalising too readily about the putative benefits of dietary antioxidants. 

The terminology describing the action of antioxidants is unfortunately not clear. Terms such as antioxidant power , antioxidant effectiveness , antioxidant ability , antioxidant activity , and antioxidant capacity are often used interchangeably and without discrimination. Here we use the term antioxidant activity as meaning a measure of the rate of antioxidant action, and the term antioxidant capacity as meaning a measure of the extent of antioxidant action, i.e. the amount of radicals or intermediates and products produced during oxidation that are quenched by a given antioxidant. Thus antioxidant activity is related to the kinetics of the antioxidant action and antioxidant capacity to the stoichiometry. 

Measuring the relative oxidative stability of polymers is important. Measurements can be used to determine dependencies on structural and molecular weight/weight distribution or the effectiveness of an antioxidant, or to perhaps assess the amount present in a polymer sample, etc. The preferred and commonest method consists in raising the sample temperature to a predetermined level, while in an inert atmosphere, then switching the atmosphere to air or oxygen. The time to the onset of exothermic reaction is measured. 

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