*2.2. Proposed Mechanism of Echinochrome A Oxidative Degradation*

Many bioactive natural and pharmaceutical compounds that are α-hydroxyketones such as oxolin, ascorbate, glyoxal, and cyclic ketones are susceptible to autooxidation [29]. It has been shown that α-hydroxyketones auto-oxidize under physiological conditions via the enediol tautomer [30,31]. One condition which favors the formation of the enediol is the presence of a vicinal carbonyl group. The equilibrium is generally displaced in favor of the more thermodynamically stable ketol tautomer. The autooxidation of such enediols, as well as tetrahydroxy-l,4-benzoquinone, ascorbate, and Ech A, has been shown to involve the generation of intermediates such as carbon-centered free radicals and ROS including the superoxide radical anion, hydroxyl radical, and hydrogen peroxide [23,30,32,33]. The mechanisms of the primary attack of triplet and singlet oxygen molecules on Ech A (**1**), the result of which is bis-*gem*-diol **2** formation, were described in detail in previous studies [23,34]. Here, according to the structures of the isolated products, we propose the scheme of the further Ech A oxidative degradation process (Figure 8).

**Figure 8.** Proposed scheme of the Ech A oxidative degradation process.

Pracht et al. showed that splitting of the aromatic system of phenolic substances occurs only if the first oxidation stage includes the formation of *o*-quinone [35]. Subsequent cleavage of the ring structure can occur between two keto groups, as in our case, for example, in compounds **3**, **4**, **4a**, and **4b**, which always presented together with compound **2** (Figures 3 and 4). It can be assumed that ring rupture in the primary oxidation product **2** with loss of H2O and CO2 led to the formation of phthalonic acid derivative **7**. Compound **7**, resulting from keto–enol tautomerism, gave echinolactone **11** upon dehydration in an acidic medium and heating during the evaporation process. Establishment of the structure of **11** using X-ray analysis played an important role in the elucidation of the structure of the labile compound **7**. Further sequential decarboxylation of **7** resulted in the formation of **8**, a benzoic acid derivative with an aldehyde group, and a benzoic acid derivative **9**. Decarboxylation of **9** and subsequent hydration and oxidation led to the formation of stable benzoquinone **10**.

Thus, it was shown that the oxidative destruction of Ech A did not affect the benzenoid fragment of its molecule. All transformations occurred only in the quinonoid ring with the formation of bis-*gem*-diol **2**, further oxidation of which occurred upon cleavage of the dihydroquinonoid ring and led to the formation of derivatives of phthalonic (**7**) and benzoic (**8**, **9**) acids, as well as benzoquinone **10**.
