*3.2.1. Total Radical Trapping Antioxidant Parameter (TRAP) assay*

This method is based on the measurement of the fluorescence of a molecular sample. Canadian researchers proposed this method to determine total peroxyl radical-trapping antioxidant capability of plasma [39]. They used a water-soluble azo compound, such as AAPH [2,2'-azobis-(2-amidinopropane)]. They measured the induction time electrochemically by measuring the time in which the antioxidants contained in the analysed sample prevent the capture of oxygen atoms. Four years later, DeLange and Glazer [40] proposed R-phycoerythrin in an induction measurement as a fluorescence indicator. They observed the time in which antiox‐ idants in the sample protect R-phycoerythrin from oxidation and compared it with the time by which Trolox, added at a known amount, prevented a decrease in fluorescence. The reaction kinetics curve was monitored at the excitation wavelength of 495 and emission wavelength of 575, and antioxidant properties were expressed as Trolox equivalent. However, the method is time-consuming and rather complicated, which makes it susceptible to considerable errors, especially in inter-laboratory studies. Another modification of the method was developed by Valkonen and Kuusi [41], who used dichlorofluorescein diacetate (CDFH-DA) as an indicator of oxidation progress. DCFH-DA is hydrolysed in the presence of AAPH to highly fluorescent dichlorofluoresceine (DCF). An increase in fluorescence is a sign that the antioxidant activity of the analysed sample is exhausted.
