## 3 Results and Discussion
### Yeast Analysis
This metric was chosen to enable an evaluation of filtration behavior within a realistic brewing framework. Figure 1: SEM images of dry yeast in its original state, before pitching (SEM parameters: HV 15.000 kV; det ETD; WD 5 mm). Yeast viability (percentage of live, metabolically active cells in a yeast population) was assessed after 60 h of fermentation at 20 \({}^{\circ}\)C, followed by a 48 h cold crash at 2.5 \({}^{\circ}\)C and an additional 24 h at 4 \({}^{\circ}\)C in bottles before sample processing. Sampling for yeast cell count and viability assessment was performed immediately prior to the onset of filtration, ensuring that the measured values accurately reflected the yeast load at the filter inlet. To eliminate sampling-related variability, all samples were homogenized before microscopic staining. Consequently, the 24 h storage in bottles prior to staining analysis did not influence the total cell count. The measured concentrations were 4.7 \(\times\) 10\({}^{6}\) cells/mL for LA-01, 5.8 \(\times\) 10\({}^{6}\) cells/mL for LoNa, and 6.2 \(\times\) 10\({}^{6}\) cells/mL for W-34/70. According to standard brewing research [11; 23], beers typically undergo cold maturation at 0 to \(-\)2 \({}^{\circ}\)C for 1 to 2 weeks to improve clarification, enhance filterability, and increase colloidal stability. During this phase, yeast concentrations are expected to decrease below 2 \(\times\) 10\({}^{6}\) cells/mL, with 5 \(\times\) 10\({}^{6}\) cells/mL considered the upper acceptable limit prior to filtration [23]. In the present study, the beers were subjected to a short cold-storage period of only 48 h at 2.5 \({}^{\circ}\)C. As a result, yeast concentrations remained slightly above the commonly recommended thresholds. Nevertheless, the observed values were considered acceptable for initiating the filtration trials, particularly given this study's focus on realistically simulating practical brewing conditions. The viability of the non-maltose-metabolizing yeasts was 55.6% for LA-01 and 53.4% for LoNa, compared to 88.4% for the bottom-fermenting yeast W-34/70. This reduced and varying viability observed in the specialized non-maltose-metabolizing yeasts may present challenges in brewing non-alcoholic beers. The differences in viability can be explained by initial damage and irregular yeast cell morphology (Figure 1) caused by the freeze-drying process. If yeast cells are not in a healthy condition at the outset, their long-term viability may be negatively affected, potentially impacting fermentation performance and product quality. Furthermore, limited sugar metabolism may lead to earlier yeast starvation and decreased viability due to the restricted availability of fermentable sugars. The literature shows that non-alcoholic mashing procedures typically yield higher maltose and maltotriose levels while limiting glucose and fructose concentrations to limit the ethanol content in the final non-alcoholic products [9]. The initial glucose concentration in wort significantly impacts sugar consumption, as glucose repression prevents yeast from metabolizing other sugars until glucose is depleted. High glucose levels, whether from crystalline or liquid sugar extract, can allow the yeast's enzymatic system to adapt to prioritize glucose. This adaptation reduces or halts maltose uptake, often resulting in incomplete fermentation [11]. Brewing processes after fermentation must be meticulously planned if yeast reuse or filtration is required. Yeast starvation can lead to autolysis, potentially releasing substances that negatively affect filtration [12; 13]. In contrast, the bottom-fermenting yeast (W-34/70), capable of metabolizing all low-molecular-weight sugars, exhibited higher viability (88.4%). This can be attributed to increased sugar availability in wort, supporting more robust metabolism during fermentation.