## 3 Results and Discussion
### Volatile Organic Compounds  
Beer is a beverage rich in alcohols and esters, which are formed during alcoholic fermentation [36]. Higher alcohols, considered the most abundant aromatic compounds in beer, are produced from amino acids through the Ehrlich route or from carbohydrate metabolism [37]. Esters are formed by yeast via an enzyme-catalyzed reaction between acyl-CoA and higher alcohols. These compounds play a crucial role in the production of various beverages, such as wine, whiskey, and beer, due to their low perception threshold. Table 2 presented the normalized area of volatile compounds produced by the selected _S. cerevisiae_ strain, both single and co-inoculum fermentation with _Sc. pombe_ G18, under different experimental conditions. The volatile profiles of single fermentations showed significant differences, depending on the type of cereal used, compared to those with mixed-starter fermentations. Laboratory-scale fermentation using single inoculation was characterized by higher values across all volatile compound classes. The esters class was the most represented, with values varying from 3.78 (LSC-95029) to 0.82 (LSC-G18-9502) mg/L. Among the volatile compounds, esters were the most represented class in all beers, followed by alcohols. Among the various compounds, esters and alcohols play a crucial role in the organoleptic characteristics of alcoholic beverages [38]. Specifically, esters, such as isoamyl acetate, ethyl hexanoate, and ethyl octanoate (associated with banana, sweet, and sour apple notes, respectively), positively contributed to the aroma with fruity notes due to their low perception threshold [39; 40; 41]. Figure 1: Biplot of scores (**A**) and loadings (**B**) obtained with the application of PCA to alcohol, organic acid, and sugar contents along the first and second component. The total higher alcohol content ranged from 0.67 (LSC-9502) to 0.12 mg/L (CSC-G18-9502). When present in amounts below 300 mg/L, higher alcohols impart floral, pleasant, and refreshing notes and contribute a desirable warming character that adds complexity to the beer. However, higher concentrations result in a burning sensation and an alcohol or solvent aroma [42]. Based on the pairwise comparisons, the LSC samples had significantly higher contents of alcohols, while the sole hydrocarbon detected, i.e., styrene, was found only in CSC beers and in very low concentrations, regardless of the product formulation. Styrene can be found in many commercial wheat beers because of both thermal decarboxylation of cinnamic acid during the boiling step and enzymatic decarboxylation during fermentation. The absence of styrene from LSC beers could be explained by their higher aeration that facilitated the evaporation of this aromatic compound [43]. The eerpenes, norispoprenoids, and aldehydes/ketons were highly concentrated in LSC beers except for beers produced by sequential fermentation, where concentrations were similar in laboratory and industrial beers. The organic acids were highly concentrated in LSC beers, except for industrial beers produced by sequential fermentation. Regarding total volatile concentration, the beer samples were ranked in the following decreasing order: LSC-9502, CSC-9502, LSC-G18-9502, and CSC-G18-9502. To visualize the differences in volatile compounds depending on the different experimental conditions tested (raw materials, fermentation conditions, and type of production level), a PCA was carried out using the mean of the repetitions. Figure 2: Biplot of scores (**A**) and loadings (**B**) obtained with the application of PCA to the various classes of volatile compounds along the first and second component. 1, total hydrocarbons; 2, total alcohols; 3. Total terpen/norisoprenoids; 4, total aldehydes/ketons; 5, total esters.