In addition to peroxyl radicals the most found radical

In addition to peroxyl radicals, the most found radical forms in biological systems, some of assays took into account other free radicals such as superoxide anion, hydroxyl, hydrogen peroxide, singlet oxygen and peroxynitrite. ORAC procedure was also modified using different radical sources such as reactive oxygen species, superoxide anion, hydroxyl radical, singlet oxygen and reactive nitrite species and the sum of the antioxidant capacity measured by these six radicals was described as ORAC Multiple Radicals (ORACMR) (Mullen et al., 2011, Ou et al., 2002, Soung et al., 2004, Zhang et al., 2009). However, it was found that these assays were inconvenient to determine the antioxidant capacity of foods since some of them was not suitable to quantify nonenzymatic antioxidants, some of them was difficult to apply and was not practical to use in routine analysis. Later on, another method based on the absorbance of a stable but non-biological ABTS+ radical cation was developed and trolox equivalent antioxidant capacity (TEAC) of plasma or foods was determined as its mechanism represented schematically in Table 3 (Brand-Williams et al., 1995, Miller et al., 1993). It was noticed that adding a sample to the reaction medium before the radical (ABTS+) formed could be a drawback of this assay as antioxidants could react with oxidizing agent leading to overestimation of antioxidant capacity. Therefore, the assay was modified by adding the sample into the reaction medium after generation of ABTS+ to prevent the overestimation of antioxidant capacity of a food sample. The ABTS assay was used to screen the antioxidant capacity of both lipophilic and hydrophilic antioxidants, including flavonoids, hydroxycinnamates, carotenoids, and plasma antioxidants (Re et al., 1999). On the other hand, it was criticized that the ABTS+ radical was not found in human body and recognized as a non-physiological assay. Based on a similar approach, the antioxidant capacity of a compound was measured using a hydrophilic radical N,N-dimethyl-p-phenylenediamine (DMPD) instead of ABTS radical. In this assay, DMPD+ radical cation was generated in the presence of an oxidant (e.g. ferric chloride) at acidic condition and decolorization of DMPD+ radical cation due to the transfer of hydrogen Lovastatin from an antioxidant to radical was monitored spectrophotometrically (Table 3) (Fogliano, Verde, Randazzo, & Ritieni, 1999). Another spectrophotometric method was used to determine the total phenols content by Folin-Ciocalteu Reagent (FCR). Although this assay was initially discovered for the protein analysis (Folin & Ciocalteu, 1927), then it was adapted to total phenol assay based on electron transfer during reacting phenolic compounds with FCR under basic condition (Singleton, Orthofer, & Lamuela-Raventos, 1999). Several studies applied to observe the lineer correlation between total phenol content and antioxidant capacity of a sample (Velioglu, Mazza, Gao, & Oomah, 1998). However, this assay has been criticized since FCR is nonspecific to phenolic compounds leading to be reduced by other non-phenolic compounds (Huang, Ou, & Prior, 2005). Benzie and Strain (1996) developed a new method based on the electron transfer reaction mechanism called Ferric Reducing Antioxidant Power (FRAP) and determined the reducing capability in plasma at first, and then in extracts or food (Benzie and Strain, 1996, Bravo et al., 2007, Liu et al., 2008). On the other hand, it was revealed that FRAP assay caused an underestimation in antioxidant capacity because of its incapability to measure the radical quenching power of compounds such as thiols and proteins. Therefore, the combination of FRAP assay with other methods measuring the radical quenching ability was thought to be important to determine the dominant reaction mechanism of antioxidant compounds (Prior et al., 2005). Bravo et al. (2007) showed a very good correlation between antioxidant capacity measurement performed with FRAP and ABTS method and concluded that the mate leaves antioxidants exert their activity through redox-based reactions. Furthermore, it was emphasized that the recommended time for FRAP protocol might not be enough to measure all antioxidants, as ferric ion reacted with antioxidant compounds slowly. FRAP assay was conducted in non-physiological acidic condition. Pérez-Burillo et al. (2018) et al. developed an alternative method in order to measure the general reducing capacity of samples instead of ferric reducing capacity using indigo carmine as a redox indicator in physiological condition. It was demonstrated that carmine was considered the fore candidate as a redox indicator to measure the antioxidant capacity due to the fact that it did not suffer any interference during the absorbance readings (Pérez-Burillo et al., 2018). Apak et al. (2004) also introduced a method called Copper Reducing Antioxidant Capacity (CUPRAC) that overcome the disadvantages of FRAP assay, due to the redox chemistry and faster kinetic potential of copper ion. In addition, all types of antioxidants including the thiols and proteins could be detected more selectively than FRAP since common interferences in FRAP such as sugars, citric acid could not react with copper and not oxidized in CUPRAC approach (Prior et al., 2005).