## 3 Results and Discussion
### Volatile Aroma Compounds
#### 3.2.8 Multivariate Statistical Analysis  
To better visualize the diversity in volatile aroma compound composition produced by the investigated non-_Saccharomyces_ yeasts in the early phase of grape must fermentation, and to extract the most useful variables for their differentiation, SLDA was applied on mean-centred data of a reduced dataset including 40 variables (volatile compound concentrations) with the highest _F_-ratios. This SLDA model correctly classified all the grape must samples according to yeast species and extracted the nine most useful variables for their differentiation (Figure 2), with rather high squared Mahalanobis distances from group centroids (numerical data not shown). All the samples (100%) were classified correctly after including only two variables already, ethyl propanoate and 2-phenethyl acetate, while the SLDA model further extracted butyric acid, dihydro-2-methyl-3(2H)-thiophenone, 3-methylbutanal, 3-buten-2-ol, decanoic acid, hexanoic acid, and isoamyl alcohol. Hierarchical clustering analysis performed on a reduced dataset including 40 variables (volatile compound concentrations) with the highest _F_-ratios, confirmed that each yeast species studied produced a distinct volatile profile in the early phase of fermentation (Figure 3). _Saccharomyces cerevisiae_ early ferment was separated from the musts inoculated by non-_Saccharomyces_ yeasts mostly by higher concentrations of propyl acetate, isobutyl acetate, butyric acid, decanoic acid, 2-phenylethanol, and benzaldehyde, and the lowest concentrations of compounds such as \(\beta\)-damascenone and dihydro-2-methyl-3(2H)-thiophenone. _Saccharomyces cerevisiae_ grape must was also characterized by lower concentrations of particular esters compared to the musts inoculated with non-_Saccharomyces_ycasts except _T. delbrueckii_, such as methyl octanoate, ethyl dodecanoate, isobutyl octanoate, isoamyl hexanoate, and ethyl 9-decenoate. _Torulaspora delbrueckii_ must contain the lowest concentrations of a large array of compounds and, at the same time, was distinguished by the highest concentrations of ethyl propanoate, 2-phenethyl propanoate, dihydro-2-methyl-3(2H)-thiophenone, and _trans_-3-hexen-1-ol, which resulted in the largest distance from the other species. _Pichia kluyveri_ stood out with increased levels of linalool, 3-buten-2-ol, \(\beta\)-damascenone and a whole range of acetate esters. The highest levels of \(\beta\)-pinene, hexanoic acid, and ethyl hexanoate were characteristic of the _S. pombe_ early ferment. Although evidently different and distant from each other, grape musts inoculated with _S. cerevisiae_ and _S. pombe_ were distinguished from the others by high octanoic acid and ethyl butyrate, and low _trans_-2-hexen-1-ol, 1-hexanol, and dihydro-2-methyl-3(2H)-thiophenone concentrations. Figure 2: Separation of Malvazija istarska grape musts in early fermentation according to inoculated yeast species defined by the first three discriminant functions (roots) obtained by forward stepwise discriminant analysis on the basis of volatile aroma compound composition. Projection of grape must samples is shown in sub-figures (**a**,**c**,**e**), while standardized coefficients of the variables (volatile aroma compounds) are shown in sub-figures (**b**,**d**,**f**). Abbreviations: SC—_Saccharomyces cerevisiae_; TD—_Toruspora delbrueckii_; MP—_Metschinkowia pulcherima_; PC—_Pichia kluyveri_; LT—_Lahancua thermotolaneus_; SP—_Schizosaccharomyces pombe_; Dih-2-meththio–dihydro-2-methyl-3(2H)-thiophenone; EthProp—ethyl propanoate; IsoamAlco—isoamyl alcohol; 2-PhenethAc—2-phenethyl acetate; HexAc—hexyl acetate; ButAcid—butanoic acid; DecAçid—decanoic acid; 3-Metbut—3-methylbutanal. Figure 3: Hierarchical clustering analysis performed using volatile aroma compound composition of Malvazija istarska grape must in early fermentation by _Saccharomyces cerevisiae_ and various non-_Saccharomyces_ yeasts. The rows in the heatmap represent compounds, and the columns indicate samples.