## 3 Results and Discussion
### Good Survival during Storage at 0 and \(-\)20 \({}^{\circ}\)C  
For bacteriophage products (in pure form as powder/compressed into tablets or incorporated into food matrices) to be able to influence the gut microbiota and to achieve a therapeutic effect, their infectivity must not be lost during storage. For this purpose, the capsules were subjected to a storage experiment. The bacteriophage titers for the capsule variant composed of alginate and micellar casein during a storage period of 60 days at 23, 0, and \(-\)20 \({}^{\circ}\)C are exemplary shown in Table 3.  
A preliminary test (single determination) showed that storage at 23 \({}^{\circ}\)C reduces the titer by more than 3 log units within 60 days, which is unacceptable in terms of the quality characteristics and function of such a product. Therefore, this temperature is not recommended. At storage temperatures of 0 and \(-\)20 \({}^{\circ}\)C, the bacteriophage titers remain stable over 60 days. Accordingly, it would be sufficient to store the product at around 4 \({}^{\circ}\)C. It may be assumed that the bacteriophages remain active for a much longer period of time at cool temperatures; however, this needs to be investigated further. In most studies, a low storage temperature is considered beneficial. [PERSON] et al., stored air-dried extruded alginate/ whey protein isolate capsules containing _Salmonella_ bacteriophage Felix O1 or _Staphylococcus aureus_ bacteriophage K for 14 days. They found a reduction of 10-20% at 23 \({}^{\circ}\)C, but no reduction at 4 \({}^{\circ}\)C [88; 90]. [PERSON] et al., analyzed the storage stability of _Salmonella_ bacteriophages Felix O1 in spray-dried Eudragit S100(r)/trehalose capsules over a period of 90 days. They observed 1 log unit reduction at 23 \({}^{\circ}\)C, but only 0.2 log units reduction at 4 \({}^{\circ}\)C [76]. Within a time frame of 6 months at 4 \({}^{\circ}\)C, alginate/calcium carbonate encapsulated _Salmonella_ Typhimurium bacteriophages were reduced by around 25% [93]. [PERSON] et al., showed that _Pseudomonas_ bacteriophages encapsulated in trehalose/leucine capsules survived storage at 20 and 4 \({}^{\circ}\)C for one year [96]. Storage studies of bacteriophages at freezing conditions showed their persistence over longer periods. For example, _Lactococcus_ spp. bacteriophages (formerly referred to as _Streptococcus_ spp. bacteriophages) stored at \(-\)18 \({}^{\circ}\)C did not decrease significantly within 2.5 years [97]. \begin{table}
\begin{tabular}{c c c c} \hline \hline  & & **Log \(N_{C,t}\) (pfu g\({}^{-1}\))** & \\ \hline \(t\) (Days) & \(\theta\) = 23 \({}^{\circ}\)C * & \(\theta\) = 0 \({}^{\circ}\)C & \(\theta\) = \(-\)20 \({}^{\circ}\)C \\ \hline
0 & 7.19 \(\pm\) n.d. ** & 7.13 \(\pm\) 0.53 & 7.13 \(\pm\) 0.53 \\
15 & 6.37 \(\pm\) n.d. & 7.04 \(\pm\) 0.49 & 7.21 \(\pm\) 0.40 \\
30 & 4.91 \(\pm\) n.d. & 7.35 \(\pm\) 0.14 & 7.35 \(\pm\) 0.19 \\
60 & 3.43 \(\pm\) n.d. & 7.25 \(\pm\) 0.05 & 7.31 \(\pm\) 0.08 \\ \hline \hline \end{tabular} \({}^{\star}\) single determination. ** n.d.—not determined.  
\end{table} 
Table 3: Survival during storage at 23, 0, and \(-\)20 \({}^{\circ}\)C of bacteriophage K5 capsules composed of alginate and micellar casein. The limit of quantification was 103 pfu g\({}^{-1}\). Besides the low temperature, a dry ambient air is also beneficial. This is directly correlated to the humidity of the capsules themselves (Section 3.4), since they tend to absorb water from their surroundings. [PERSON] et al., could have taken this into account, as they vacuum packaged their particularly long-lasting capsules [96]. Additionally, the composition of the capsules is important for the survival of bacteriophages. For example, saccharides such as lactose or trehalose are known as efficient auxiliary materials [96,98]. In the end, the bacteriophage species should always be taken into account as well.