# Impact of Cork Closures on the Volatile Profile of Sparkling Wines during Bottle Aging
## Materials and Methods
### Statistical Analyses  
Multiple unpaired \(t\)-tests were applied to evaluate the differences in the levels of volatile compounds in sparkling wines sealed with 2D compared with MA at each post-bottling time. In addition, ordinary one-way analysis of variance (ANOVA) was computed to assess the differences in volatile concentrations between different post-bottling times. The concentration levels were considered significantly different for \(p\)-values \(<\) 0.05. All statistical analyses were performed using the software GraphPad Prism 9 (version 9.3.0, San Diego, CA, USA). The volatile composition of sparkling wines #1 and #2 was more affected by the type of closure at 24- and 42-months post-bottling, while only a qualitative change in camphor (only present in samples sealed with 2D, Figure 1 and Table 1) was detected in both sparkling wines at 12 months. Camphor is responsible for pleasant aromas--such as herbal, minty, and woody [13]--but the olfactory perception threshold in wine or wine model solution has not been reported so far. At 24 months post-bottling, sparkling wine #1 showed significantly higher levels of 3-hexen-1-ol and \(\beta\)-damascenone in samples sealed with 2D compared with MA and the presence of camphor only in 2D samples (Figure 1, Table 1). 3-hexen-1-ol is characterized by green and leafy odors and was present in concentrations below the olfactory perception threshold (<400 \(\mu\)g/L) reported for wine model solution [14], while \(\beta\)-damascenone is characterized by woody, floral, and herbal odors [14; 15], and was present above its olfactory perception threshold (0.05 \(\mu\)g/L) [14]. At 42 months, this sparkling wine showed significantly higher levels of ethyl isobutanoate, ethyl butanoate, and ethyl isovalerate in samples sealed with 2D, as well as significantly lower levels of 2-undecanone and the presence of camphor (Figure 1, Table 1). From these compounds, the three ethyl esters (ethyl isobutanoate, ethyl butanoate, and ethyl isovalerate), and 1-hexanol were present above their olfactory perception thresholds (Table 1) [14], and can contribute with fruity notes [13; 15] to the sparkling wine aroma. Interestingly, the levels of 1-hexanol, ethyl butanoate, ethyl isovalerate, 2-undecanone, \(\beta\)-damascenone, and camphor changed significantly with bottle aging, while the levels of 3-hexen-1-ol and ethyl isobutanoate were relatively constant over time (Table 1). \begin{table}
\begin{tabular}{l c c c c c c c c c c c c} \hline \hline \multirow{2}{*}{**ClustCompound**} & \multicolumn{3}{c}{**22 Months 1**} & \multicolumn{3}{c}{**22 Months 1**} & \multicolumn{3}{c}{**22 Months 1**} & \multicolumn{3}{c}{**22 Months 2**} & \multicolumn{3}{c}{**22 Months 2**} & \multicolumn{3}{c}{**22 Months 2**} & \multicolumn{3}{c}{**Oloratory perception**} \\  & **2D** & **MA** & \(p\) & **2D** & **MA** & \(p\) & **2D** & **MA** & \(p\) & **Months 2** & 2 & **Perception** & **Improved3** \\ \hline \hline Alcohols & & & & & & & & & & & & & \\ 3-Homm-I-ol & 342 \(\pm\) 6.2 & 31.0 \(\pm\) 5.3 & **m** & 30.1 \(\pm\) 40 & 22.8 \(\pm\) 2.0 & * & 25.4 \(\pm\) 18.0 & 20.6 \(\pm\) 18.1 & **m** & **m** & **Crown, leafy** & 400 \(\mu\)g/L \\ \(\log\)(g/L) & 1.1 \(\pm\) 0.03 & 1.0 \(\pm\) 0.1 & **m** & 22.1 \(\pm\) 1.0 & 21.6 \(\pm\) 0.3 & **m** & 13.7 \(\pm\) 0.5 & 55 \(\pm\) 2.7 & **-** & **-** & **Crown, leafy** & 8 \(\mu\)g/L \\ \(\log\)(g/L) & & & & & & & & & & & & \\ Ethylutona & 640 \(\pm\) 6.3 & 80.7 \(\pm\) 3.0 & **m** & 95.3 \(\pm\) 540 & 1523 \(\pm\) 52 & **m** & 201 \(\pm\) 29.0 & 81.7 \(\pm\) 38.7 & **-**  
In contrast, at 24 months post-bottling, sparkling wine #2 showed the consistent presence of camphor in samples sealed with 2D, as well as significantly higher levels of ethyl decanoate (Figure 2, Table 2). At 42 months, significantly higher levels of ethyl octanoate and a tendency for higher levels of ethyl decanoate were observed in samples sealed with 2D, along with the presence of camphor (Figure 2, Table 2).