## 4 Discussion
### Bioactive, Trepenoid, and Phenolic Profile Comparison Among Different Ecotypes of Burdock Roots
In this sense, the PCA indicated that the high bioactivity of the SPME extracts could be due to the content of caffeoylquinic acids, such as chlorogenic acid and 4-O-caffeoylquinic acid, as well as cyanophyllene oxide, spathulenol, and bisonwallocholanic acid. In fact, several authors have reported that caffeoylquinic acids, such as chlorogenic acid, have demonstrated a memory improvement that has been related to their high antioxidant capacity, in addition to their antibacterial, antiviral, and anticarcinogenic capacities [56]. The phenolic profile of burdock has been widely studied. Several authors have shown that most of the phenolic compounds found in _Arcium lappa_ L. are caffeoylquinic acids such as monocaffeoylquinic acids (chlorogenic acid and 1-O-caffeoylquinic acid), dicaffeoylquinic acids (1,5-O-dicaffeoylquinic acid, 3,4-O-dicaffeoylquinic acid/3,5-O-dicaffeoylquinic acid, and 1,3-O-dicaffeoylquinic acid (also known as cynarine)), and a dicaffeoylmuloylquinic acid [54; 55]. Recently, cynarine, which is one of the major compounds identified in the SPME extract, has shown an interesting capacity to inhibit neuronal degeneration reducing glutamate release, an excitatory neurotransmitter that, in excess, causes oxidative stress, neuroinflammation, mitochondrial dysfunction, and finally neuron death [57]. In addition, although arctin was found to have the lowest peak area of the phenolic compounds identified in this study, the extraction of this compound from burdock roots has been widely investigated for its antioxidant potential [58; 59; 60; 61]. The arctin content of plants is crucial because it serves as a precursor to arcigenin, a bioactive compound with a broader spectrum than arctin in pharmacological properties. Upon ingestion, arctin is metabolized by the gut microbiota into arcigenin, which exhibits enhanced biological activity, including antiviral, anticancer, and anti-inflammatory effects. Therefore, plants rich in arctin provide a natural source for generating arcigenin within the body, amplifying their therapeutic potential [58; 59; 60; 61]. Notably, caryophyllene has been shown to be neuroprotective in cases of cerebral ischemia and neuronal damage caused by oxidative stress, modulating glial activation and neuroinflammation [62]. In addition to the high antioxidant, anti-inflammatory, antiproliferative, and antimycobacterial activities demonstrated by spathulenol, this compound has shown interesting neuroprotective capacity by reducing ROS production and maintaining the mitochondrial integrity [63; 64]. Spathulenol and caryophyllene oxide have also been identified as components of essential oils obtained from plants of the Asteraceae family. In agreement with [PERSON] et al., caryophyllene oxide was identified as the major volatile compound detected in the burdock roots [65]. However, to our knowledge, this is the first time that spathulenol has been identified in burdock roots. Probably, these compounds were responsible for the high antioxidant and anticholinergic capacity of the SPME extract. Moreover, although sterols were not the primary contributors to the antioxidant and anticholinergic capacities of the extracts in this study, various authors have highlighted that stigmasterol is commonly found in burdock roots [66; 67]. In addition, the absence of quercetin in the burdock root extracts obtained with 2-MTHF in this study was noted, although its frequent detection in extracts obtained with methanol or ethanol-water mixtures in other studies may be attributed to differences in solvent polarity and extraction efficiency [68; 69]. This compound has been characterized by its high neuroprotective potential; however, the anti-cholinergic capacity of the extracts obtained in this study was not related to quercetin [70]. Quercetin, a highly polar flavonoid, is more efficiently extracted by methanol-water mixtures due to their higher polarity and stronger hydrogen-bonding capacity, which facilitates the solubilization of hydroxyl-rich compounds.