## 2 Results
### Optimization of HS-SPME-GC \(\times\) GC-TOFMS Methodology
This can be explained by the fact that the use of NaCl favors the release of polar compounds occupying the binding sites on the fiber to the detriment of terpenes, which are compounds possessing a more non-polar nature [39]. This decrease is also reported in the literature and indicates that the increase in the amount of salt can indeed lead to a loss of fiber selectivity [23]. According to [PERSON] et al. 2011 [4], when we are dealing with complex matrices such as grapes and wines, sample dilution can affect the partition between the headspace and the sample. Consequently, the volatile profile can be altered, as competition between the analytes that are presented in the headspace [4]. To investigate the dynamics of headspace competition, we conducted sample dilution by adding 2 and 4 mL of H\({}_{2}\)O. A sample without dilution was used for comparison. The results obtained are presented in Figure 1c, and it is possible to observe that, using 2 mL of H\({}_{2}\)O, it was possible to  
Figure 1: Influence of the different amounts of (**a**) sample, (**b**) salt (NaCl), and (**c**) H\({}_{2}\)O in the number of terpene compounds of each chemical family. identify 45 compounds. Using 4 mL of H2O allowed for the identification of 25 compounds, while 41 compounds were identified in the undiluted sample. This result was relatively unexpected, since according to the literature [4], a more significant difference should have been observed between the undiluted sample and the sample with 2 mL of H2O, which allows us to infer that this dilution did not affect partitioning between the headspace and the sample. However, with a higher dilution of 4 mL, it was observed that the number of compounds found decreased, specially the sesquiterpenes. Nevertheless, the results show that the addition of 2 mL of H2O was the most promising procedure for the analysis of varietal volatile compounds. After optimizing the amount of the sample, salt, and H2O, the extraction time and temperature were optimized. In order to optimize the extraction time, the DVB/CAR/PDMS fiber was exposed in the headspace for 20, 40, and 60 min. The choice of extraction times was based on the information in the literature [4]. According to Figure 2a, it appears that 40 min of extraction is the optimal time, since it is possible to identify a greater number of compounds, and for this reason this was the time chosen for the implementation of the methodology. In addition, the extraction temperature was also optimized, since it affects the extraction efficiency of the volatile compounds. According to the literature [4], terpenes and C13-norisoprenoids show better results at higher extraction temperatures. To evaluate this condition, the samples were tested at 40, 50, and 60 \({}^{\circ}\)C, as according to [PERSON] et al. 2011 [4]. The results obtained are presented in Figure 2b. It can be observed that the best extraction temperature used was 60 \({}^{\circ}\)C, as this allowed for the identification of 43 varietal volatile compounds. Application of HS-SPME-GC \(\times\) GC-TOFMS Methodology for Analysis of Varietal Volatile Composition  
After the optimization steps, the global volatile signature of the grapes from Trinc, CS, Sy, Cast, and TB grapes was established. The contour plots of Trinc, CS, Sy, Cast, and TB grapes are shown in Figures S1, S2, S3, S4 and S5, respectively. Figure S6 shows a contour plot of a sample of grapes in the study, with the assignments of monotepenes, C13-norisoprenoids, and sesquiterpenes regions. As expected, the compounds are eluted from the chromatographic system according to their affinity to the column; thus, the monoterepenes (region (a)) come out first, followed by the C13-norisoprenoids (region (b)), and finally the sesquiterpenes (region (c)), C10, C13, and C15, respectively. Figure 2: Influence of the (**a**) extraction time and (**b**) extraction temperature on the number of terpen compounds of each chemical family.