## 3 Results and Discussion
### Evolution of the Concentrations of Cu and SO\({}_{2}\) during Wine Production
The addition of SO\({}_{2}\) to grape juice/must did not significantly (\(p\leq 0.05\)) impact the total or free Cu concentration for either Chardonnay or Shiraz throughout the production process (Tables 1 and 2). SO\({}_{2}\) has been reported to lead to the degradation of polysulfanes to generate H\({}_{2}\)S [47], but in this study, the results did not reflect a significant (\(p\leq 0.05\)) decrease in free Cu and increase in bound Cu for wines when the SO\({}_{2}\) was added prior to fermentation. The SO\({}_{2}\)-treated Shiraz musts (M) had particularly high total SO\({}_{2}\) concentrations with poor precision, which was due to the difficulty of mixing and taking a representative sample from the must. As expected, free SO\({}_{2}\) was largely absent in all the wine samples after fermentation (PAF), regardless of any SO\({}_{2}\) addition to the grape juice/must (Tables 1 and 2). In the Shiraz wines, the total SO\({}_{2}\) concentrations were lower than those for Chardonnay, which was presumably due to more rapid oxidative losses in the must and/or due to the adsorption onto solid materials. After 15 months of aging (15 Mo), both Chardonnay and Shiraz wines showed lower total SO\({}_{2}\) concentrations than at bottling (PB-post SO\({}_{2}\)). Impact of Cu or SO\({}_{2}\) Addition to Juice/Must on Total Concentrations of Low Molecular Weight Sulfur Compounds (LMWSCs) in Wine  
The method used to measure the LMWSCs in wine involved the saturation of wine with 300 g/L of NaCl and incubation for 30 min at 45 \({}^{\circ}\)C [37]. For the majority of LMWSCs, this method is recognized as measuring the total concentrations, including the forms bound to metals as well as non-metal bound forms. The exception is hydrogen sulfide/sulfide, for which the pretreatment is regarded as being insufficient to release all sulfide from metal complexes, particularly Cu. In this instance, the measured sulfide concentration represents the sulfide in wine that is not bound to metals and sulfide that can be released from metal complexes based on the pretreatment [48]. Of the measured LMWSCs, only H\({}_{2}\)S and MeSH were significantly (\(p\leq 0.05\)) influenced by the Cu and SO\({}_{2}\) treatments (Figure 1, Table S2). For Chardonnay, after fermentation (PAF), a trend to increased H\({}_{2}\)S concentration with higher Cu addition to the grape juice (J) was evident (Figure 1A). This is consistent with Cu being toxic for yeast [49], which consequently up-regulates the expression of sulfur metabolism related genes [50] to convert Cu to the less toxic sulfide-bound form. After the cold stabilization (PCold for Chardonnay), the concentrations of H\({}_{2}\)S dropped to below its odor threshold of 1.1 \(\mu\)g/L [37] in all of the Chardonnay treatments. This may have been a consequence of the oxidative wine production process [42], the removal of some colloidal sulfide (e.g., complexed to Cu) during cracking, and volatile H\({}_{2}\)S losses. It is interesting that the decrease in bound Cu (i.e., sulfide bound) with bentonite treatments (Table 1) did not show equivalent decreases in H\({}_{2}\)S (Figure 1A). The lack of change in H\({}_{2}\)S concentrations with bentonite treatments may be due to the incomplete dissociation of all bound sulfide from Cu during the LMWSCs quantification [48]. After 15 months of bottle aging (15 Mo), the Cu addition treatments had significantly (\(p\leq 0.05\)) increased H\({}_{2}\)S concentrations in the Chardonnay wines. However, the bentonite-treated low Cu addition samples (LCu) showed significantly (\(p\leq 0.05\)) lower H\({}_{2}\)S concentrations after 15 months of bottle aging compared to the equivalent samples without bentonite treatment (Figure 1A). Cu has the ability to sequester sulfide when low oxygen conditions are reached in wine [46; 51; 52]. Therefore, the higher measured H\({}_{2}\)S concentrations in the non-bentonite aged wines may have resulted from the free Cu in the finished wines sequestering sulfide during bottle aging.