## 3 Results and Discussion  
### Fermentation Kinetics and Trend of Titratable Acidity during the Pilot-Scale Vinitification  
Pilot-scale viniification was undertaken using Cabernet Sauvignon must with LP in two inoculation modalities with SC (PreAF-LP and S-LP). Titratable acidity (TA, expressed as tartaric acid) of the grape must was monitored and recorded daily during fermentation to assess the ability of LP to induce biological acidification (Figure 1A). The trends in TA for D-SC, TA-SC and S-LP showed initial increases at the onset of fermentation, followed by a slight decrease from Day 2, then fluctuated slightly from Day 3 till Day 7, with average TA around 6.5, 7.7 and 7.6 g/L, respectively. Acidification kinetics of the bio-acidified S-LP wines were comparable to the chemically acidified TA-SC wine control (\(p\) = 0.2885, [PERSON]'s HSD post hoc analysis), but were significantly more pronounced compared to D-SC(\(p<0.05\), [PERSON]'s HSD post hoc analysis). Interestingly, TA of PreAF-LP increased steadily throughout fermentation, finally reaching 8.04 g/L by the time alcoholic fermentation completed, which was the highest amongst all treatments. Similarly, [PERSON] and [PERSON] (2014) observed a continuous decrease in pH from 3.9 (grape must) to 3.4 (resultant wine) over the course of fermentation reversely inoculated with LP. Together, these findings highlighted the effectiveness of using LP to correct the inadequate wine acidity in warm wine regions.  
Trends in yeast population during fermentation showed initial increase after inoculation, followed by reaching stationary phase from Day 2 onwards (Figure 1B). [PERSON]'s multiple comparisons test further indicated that the trends in viable yeast cell numbers were not significantly different between any two of the four batches (\(p>0.05\) in each situation). In terms of LP growth, viability drastically decreased for both LP inoculation modalities, dropping below \(10^{6}\) CFU/mL by Day 5 (Figure 1C). Such a decrease might be attributed to the inhibitory effect of the multi-stressor wine environment, especially ethanol and high wine acidity ([PERSON], 2016). Ethanol interacts with bacterial cell membranes, consequently perturbing membrane structure and cell function, whilst high acidity can impair ATPase activity and hexose metabolism in lactic acid bacteria ([PERSON], 2017). Of particular interest was the fact that bacterial numbers of PreAF-LP were 2-20-fold higher compared to those of S-LP throughout fermentation (\(p=0.0327\), Students' \(t\)-test), indicating the role of inoculation timing on bacterial viability. Differential yeast-bacteria interaction patterns between these two LP inoculation regimes may result in this phenomenon ([PERSON], 2016), but the corresponding analysis was beyond the scope of the current study.  
Compared to the control groups (D-SC and TA-SC), inoculation of LP induced a relatively slower fermentation onset (Figure 1D). Nonetheless, all tested batches fermented to dryness, with less than 4 g/L residual sugar when fermentation terminated. The fastest fermentation was displayed by D-SC (6 days), whilst the rest three batches took one day longer to deplete sugar (Figure 1D). Since very marginal difference was observed in yeast population amongst the four treatments (Figure 1B), delay in alcoholic fermentation in both the bio-acidified and chemical acidified treatments were likely to be the result of less efficient sugar metabolism by yeast compared to the D-SC control. Considering the  
Figure 1: Fermentation and acidification kinetics in the Cabernet Sauvignon pilot-scale vinicification. (**A**) Sugar consumption kinetics; (**B**) Yeast viability; (**C**) LP population; (**D**) Trends in total titratable acidity during fermentation. D-SC (), TA-SC (), S-LP (), PreAF-LP ().  
comparable fermentation duration between LP and TA-SC wines, yeast-bacteria interaction alone is unlikely to reduce the sugar metabolism efficiency of SC in LP treatments, but rather in combination with the inhibitory effect of increased TA [19].