## 3 Results and Discussion
Interestingly, the results obtained using PYR as a probe contrast with the ORAC-FL index of Trp, Gly-Trp, Trp-Gly, and Gly-Trp-Gly, which did not show differences (2.5 \(\pm\) 0.3, 2.4 \(\pm\) 0.2, 2.6 \(\pm\) 0.2, and 2.4 \(\pm\) 0.3, respectively), reflecting a similar protection of FL mediated by the additives [22]. Thus, comparing the AC of these peptides using PYR and FL as probes clearly demonstrates the dependence of competitive assays with the probe employed, with the differences explained by complex mechanisms involving, as mentioned, reactions towards ROO\({}^{\bullet}\), and RO\({}^{\bullet}\), as well as reactions of secondary radicals of the probes and Trp. For example, reactions involving hydroperoxides could be present, since these species have shown to extend the original damage on proteins inflicted by oxidants [23]. Similarly, comparison of results with systems using PGR as a probe should also denote important differences, since this target molecule has shown to be protected only for highly reactive derivatives. Consumption of 5 \(\upmu\)M PGR mediated by 10 mM AAPH was not affected for PYR, reflecting the different reactivity of both probes (data not shown). To explore if the protective pattern presented in Figure 2, Panel A, can be extended to larger peptides, we assessed the consumption of PYR in the presence of three peptides:  
Figure 2: Effect of peptides on the consumption of PYR triggered by ROO\({}^{\bullet}\). Solutions of 5 \(\upmu\)M PYR were incubated in the absence (black) or presence of 1 mM free Trp and peptides. Panel (**A**): Free Trp (red), Gly-Trp (orange), Trp-Gly (green), Gly-Trp-Gly (olive). Panel (**B**): Glu-Val-Trp-Lys-Ser-Ala-Glu (cyan), Ser-Val-Trp-Ile-Gly-Gly-Ser-Ile (blue), and Asn-Trp-Asp-Asp-Met (magenta). F/F\({}_{0}\) = normalized fluorescence intensity. Asn-Trp-Asp-Asp-Met, Glu-Val-Trp-Lys-Ser-Ala-Glu, and Ser-Val-Trp-Ile-Gly-Gly-Ser-Ile. As expected, under our experimental conditions, these peptides lack secondary structures, displaying a random conformation, according to circular dichroism measurements (Supplementary Figure S2). Similarly to Gly-Trp, a low protection of PYR, without lag times in the kinetics, was observed in the presence of 1 mM Asn-Trp-Asp-Asp-Met (Figure 2, Panel B). As Met is also susceptible to oxidation by ROO\({}^{\bullet}\)[24], the consumption of 5 mM PYR elicited by ROO\({}^{\bullet}\) was assessed in the presence of 5 mM free Met. Similar values of R\({}_{0}\) that control experiments were determined, showing a lack of PYR protection given by Met (Supplementary Figure S3). In contrast to the kinetics registered in the presence of the pentapeptide (Asn-Trp-Asp-Asp-Met), Glu-Val-Trp-Lys-Ser-Ala-Glu and Ser-Val-Trp-Ile-Gly-Gly-Ser-Ile (hepta- and octa- peptides, respectively) inhibited the consumption of PYR, generating lag times in the kinetic profiles. These results demonstrated an effect of the chemical structure of peptides on the mechanisms involved in their AC. Analogously to shorter peptides, reactions involving intermediates or oxidation products were probably involved; however, since lag times were detected in the presence of the larger species (hepta- and octa-peptides), it is reasonable to infer that their AC was Influenced by a PYR-repairing mechanism. Remarkably, the length of the lag times was dependent on the peptide studied; while the lag time produced by 1 mM Glu-Val-Trp-Lys-Ser-Ala-Glu was -5 min, the lag time generated by the same concentration of Ser-Val-Trp-Ile-Gly-Gly-Ser-Ile was close to 11 min. If the lag times are explained by the direct reaction of peptides with ROO\({}^{\bullet}\), these values would imply that 4 and 8.8 mM of ROO\({}^{\bullet}\) were neutralized by the hepta- and octa-peptides, respectively. By contrast, if Reaction (8) explains the observed lag times, these values would reflect the total amount of PYR\({}^{\bullet}\) generated within the system (assuming a low extent of termination reactions of PYR\({}^{\bullet}\)).