## 4 Discussion
Esters and amides of ferulic and caffe acid have shown the ability to inhibit HIV-1 reverse transcriptase [68; 69]. Next, the recent literature suggests that ERK, a member of the mitogen-activated protein kinase (MAPK) family of serine/threonine protein kinases, plays a key role in cell survival and is essential for antioxidant response by polyphenols [70]. The cellular activation of ERK by the ketone **44** as an extracellular stimulus is similar to previous reports indicating ERK activation by phenolic acids and their derivatives [18; 21; 36; 67]. As ERK also plays a central role in morphogenesis and physiological homeostasis of _Drosophila_, its probable activation by ketone **44** indicates organized functional changes and improved tissue homeostasis [71]. Further, on metabolic front our simulation showed that ketone **44** exhibited strongest metabolic uptake among the analyzed polyphenol analogs. This supports the potent in vivo activity of ketone **44** in _Drosophila_. This study demonstrates that the antioxidant effect of phenolic acids does not only depend on the presence of the catechol moiety. Interestingly, the replacement of the carboxylic acid functional group by the ketone function increases the antioxidant effect of the molecule. Unlike the ketone analog, the amide and ether analogs were less active. The effect of ketone **44** can be attributed to the favorable interactions as predicted by molecular docking. Increased lipophilicity is probably an important factor to consider since ketone **44** is three times more lipophilic than caffeic acid. Compared to CAPE, ketone **44** is more stable since it is less conducive to hydrolysis. Investigations of the position of the 3,4-dihydroxyds in ketone **44** may have an effect on its activity. These arguments agree with a recent study in which the amide derivative of CAPE was ineffective against antioxidant transcription factors [72]. Di-hydroxylated analogs at 2.5 positions have demonstrated interesting anticancer effects [32; 72]. The effect of increased lipophilicity, as well as the presence of the unsaturated \(\alpha\)-\(\beta\) unsaturated ketone versus a simple ketone, can be as important. Our study shows mechanisms underlying the antioxidant potential of selected phenolic acids; however, several future investigations can be suggested. Activation of the Nrf2 pathway is dependent on the endogenous inhibitor of Nrf2, Keep1, function as an evolutionarily conserved intracellular defense mechanism to counteract oxidative stress [47; 73]. Moreover, activation of antioxidant enzymes depends on the spatial distribution of both Nrf2 and Keep1 [48]. Therefore, future experiments can be targeted to understand the impact of the novel phenolic acid derivatives on keep1, the main intracellular regulator of Nrf2 activation. Interestingly, our pilot experiment did show the nuclear migration of Nrf2, thus, supporting further experiments on intricate cellular antioxidant mechanisms (Figure S4). Most investigated Nrf2 activators function as non-specific covalent modification of thiol groups Keep1 cysteine residues. Owing to its potent antioxidant activity, we expect ketone **44** to inhibit Keep1-Nrf2 binding, leading to nuclear translocation of Nrf2 _vitro_ (Figure S5). The lack of comparative analysis of ketone **44** with small-molecule Nrf2 activators such as dimethylfumarate, curcumin, quercetin, and sulforaphane hinders a broader understanding of its efficacy. The novel compounds are also warranted investigations on their ability to induce epigenetic changes in histones, owing to Nrf2-ERK activation. It is vital to note that despite the lack of downstream SOD activation in vivo (Figure 6D), Nrf2 activation did result in increased HO-1 and GPx levels, indicating a SOD independent but wider antioxidant spectrum of the tested compounds in vivo. It is well known that certain polyphenols exhibit pro-oxidant activities at higher concentrations depending on the cellular microenvironment [74].