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Measurements of the antioxidant capacity in human plasma

The human body, constantly exposed to free radicals, evolved a complex defence system, including compounds that prevent the formation of new radicals by removing peroxides, by dismutating superoxides, by chelating metal ions and compounds that inhibit the propagation of peroxidation (the so-called chain-breaking antioxidants) [4]. Dietary modulation of oxidative stress, however, involves mainly the chain-breaking antioxidants. Essential dietary antioxidants, including vitamin A, C and E are involved in oxidative stress control, as are carotene and carotenoids [1,2]. However, numerous non-essential dietary compounds such as phenolics are also considered antiatherosclerotic and anticarcinogenic and presumably play a role in controlling oxidative reactions in vivo [5]. While these antioxidants seem to be important physiologically, very little is known about the absorption, incorporation, metabolic fate and antioxidant action in humans. The concentration of each of these antioxidants and their relative antioxidant activity (that is the amount of free radicals trapped per mole), and their possible synergism contributes to the total antioxidant capacity. This parameter, therefore, represents a more integrated measure than the simple evaluation of the content of individual antioxidants, and includes all unmeasured or unmeasurable compounds. The methods available for this purpose take advantage of a constant peroxyl radical flow produced by thermal decomposition of an azocompound [8]. This flow leads to the oxidation of a target molecule, whose decay is a measure of the lipid peroxidation reaction and, indirectly, of the ability of plasma, body fluids or simple chemical solutions to break the reaction. Figure 1 resumes the method we used to assess the antioxidant capacity of plasma or antioxidant compounds [3]. A constant flow of peroxyl radicals is obtained by adding 4 mM AAPH to a reaction mixture consisting in 150 nM R-Phycoerythrin (R-PE) when plasma (or antioxidants) is added, a lag phase is produced. The plasma antioxidant capacity can be quantified, by standardizing the lag-phase induced by plasma with that induced by a known amount of an antioxidant compound (Trolox). The antioxidant potential produced by ascorbate, urate, tocopherol, and thiol groups accounted only for 67% of the total antioxidant capacity. The remaining 23% may be explained by other compounds present in plasma (carotenoids, retinol, bilirubin, creatinine, polyphenols) which could play an important antioxidant role. To evaluate whether dietary phenolics significantly contribute to systemic antioxidant defences we studied plasma antioxidant capacity in human beings before and after consumption of polyphenol-rich beverages (tea and wine). Black and green teas have a different phenolic pattern, as reported in Table 1. During black tea preparation, in fact, oxidation of polyphenols takes place, thus decreasing the levels of simple