## 6 Influence of Ageing Factors on the Coumarin Contents of Spirit Beverages
### Ageing Time  
In the regulations of the European Union and other countries in which aged spirits are produced, a minimum ageing time is always defined. For those addressed in the present review, this requirement is specified in Section 2. The results of extensive research on this subject have clearly shown that the ageing stage requires a certain period of time for multiple physicochemical phenomena to occur, slowly and continuously, in order to confer specific characteristics to the distillates, contributing to the sensory fullness of the final products [14; 18; 19; 21; 190]. It is also recognised that these phenomena basically fall into two categories:  
* Additive--including the ones that introduce or give rise to new compounds in the distillate undergoing ageing, such as extraction of wood compounds and lignin hydroalcoholysis;
* Subtractive--encompassing the ones that remove or modify some constituents of the distillate undergoing ageing, such as evaporation of volatile compounds, adsorption/degradation by the charred surface of the barrel, sorption in wood, and oxidation and hydrolysis reactions, among other chemical transformations. Consequently, the chemical composition of the aged spirits evolved progressively over the ageing time, as a result of the balance between additive and subtractive phenomena. Concerning coumarins, some available data in the literature (for whiskey, wine spirit and cachaca) are shown in Figure 18.  
The results reveal that scopoletin, coumarin and 4-methylumbelliferone contents increase in the first years of ageing, which can be ascribed to the prevalence of addictive phenomena, the higher concentration gradient between the wood and the distillate (devoid of these phenolic compounds) being the most plausible driving force [212]. However, different patterns are observed depending on the compound, the kind of distillate, the wood, and their interaction. Interestingly, after many years of ageing, the scopoletin content in wine spirit decreased sharply (Figure 18c), which may reflect the predominance of subtractive phenomena over additive ones. This behaviour may contribute to reducing the bitter taste of the aged wine spirit, in which coumarins seem to be involved [22]. The formation of coumarin glycosides often occurs through the process of hydroxylation and isomerization of _trans_-hydroxycinamic acid and analogous molecules, a reaction that can also occur during the ageing stage [115]. Due to the glycosidic bond, coumarin glycosides taste bitter. Bitterness comes from sugar moieties connected to the coumarin core. In plant defence mechanisms, bitterness discourages herbivores and insects from overeating [213]. Figure 17: Average concentrations of coumarins in aged wine spirits according to the toasting level of the barrels; LT—light toasting, MT—medium toasting, HT—heavy toasting (adapted from Canas [19]). Bars marked with different letters denote significant differences (Fisher’s test; \(p<0.05\)). When the sugar moiety is removed, coumarins have a slightly bitter taste in their aglycone form [22]. In addition, [PERSON] et al. [22] monitored the evolution of scopoletin, umbelliferone, 4-methylumbelliferone, esculetin, coumarin and fraxetin in ten vintages of Cognac (between 1970 and 2015) from the same producer. The wine spirits were aged in 350 L used barrels made from oak wood, with five replicates. The outcomes show that scopoletin and aesculetin had higher concentrations in the oldest spirits (338 and 264 ug/L, respectively). Compared with the aforementioned previous work [37], the opposite behaviour found for scopoletin during ageing might be related to the ageing conditions and/or to the analytical method used. The coumarin concentration also tended to increase over time, but with some fluctuation. Regarding fraxetin, the evolution pattern was similar to a bell-shaped curve; lower concentrations were observed in the 2015 and 1973 samples (19 ug/L and 118 ug/L, respectively), and the highest concentration was found in the 1995 sample (204 ug/L).