## 3 Results and Discussion
### Stability in Simulated Gastric Fluid and Release in Simulated Intestinal Fluid
[PERSON] et al., investigated the survival of _Lactococcus lactis_ bacteriophage P008 encapsulated in extruded capsules in SGF at pH 2, either with alginate or with alginate and whey protein. The extruded alginate/whey protein capsules protected the bacteriophages over a period of 120 min, whereas the alginate capsules did not protect the bacteriophages sufficiently, because bacteriophages were no longer detectable after 30 min [33]. Similar results were obtained by [PERSON] et al., when comparing alginate capsules with and without whey protein [88]. [PERSON] et al., determined which concentration ratios of alginate and sodium caseinate must be present for _Salmonella_ bacteriophages Felix O1 to remain intact over a 120 min period in SGF at pH 2. For the optimal ratio (2.0% alginate and 2.69% sodium caseinate), the titer was reduced by 1.25 log units [36]. [PERSON] et al., encapsulated _Lactobacillus delbrueckii_ ssp. _bulgaricus_ cells (formerly referred to as _Lactobacillus bulgaricus_) into alginate/milk capsules with extrusion. They observed a proportionality between milk concentration and bacteriophage stability in SGF at pH 2.0 [89]. This effect, or the absence of any protein component, was particularly noticeable with Eudragit S100\({}^{\textregistered}\)/trehalose capsules by [PERSON] et al., which did not provide protection [76]. Again, the important role of a protein component as a bacteriophage protector was shown, analogous to the protection during spray drying, as already discussed in Section 3.4. Since the positively charged proteins (isoelectric point of milk proteins pl \(\approx\) 4.1-5.2) interact with negatively charged alginate molecules, they produce a dense network, preventing the penetration of acid molecules and ions into the core [90]. In addition, the milk proteins can contribute to the protection mechanism with their buffering capacity [91]. [PERSON] et al., produced extruded and air-dried alginate/whey protein capsules. Their _Salmonella_ bacteriophages Felix O1 were not within the capsules when incubated in SGF at pH 2.0. Scanning electron micrographs showed cracks and holes on the surface of these capsules, whereby the authors postulated that they allowed ions to move easily into the matrix [88]. However, the air-drying process in the previous study with a total time of 30 h is not directly comparable to the spray drying process of this study. Furthermore, cracks and holes do not seem to be present or problematic in the present experiments (Figure 5b). Just as important as protecting the capsules from acidic pH is the subsequent release of the bacteriophages. Most likely, the contact of the bacteriophages with the neutral environment (SIF) results in swelling of the polymer alginate and the proteins. The previously dense capsular matrix expands, causing the K5 bacteriophages to release from the matrix. In addition, pancreatic enzymes are able to diffuse into the matrix in the opposite direction and hydrolyze proteins inside the capsules [33; 92]. Compared with the existing literature, the release of the bacteriophages in this study was complete and extremely rapid. [PERSON] et al., found that the release of the extruded alginate/whey protein capsules was fast, within the first minute, and then slow but steadily increasing over the incubation period of 120 min. Furthermore, not all bacteriophages were fully released in the end [33]. [PERSON] et al., observed 60-80% released bacteriophages after 5 min, but did not observe a full release [36]. The extruded alginate/milk capsules with _Lactobacillus delbrueckii_ ssp. _bulgaricus_ cells from [PERSON] et al., were fully released after 60 min [89]. [PERSON] et al., reported complete release of _Salmonella_ bacteriophages Felix O1 from air-dried alginate/whey protein capsules after more than 100 min [88]. The rapid release of the K5 bacteriophages from the capsules could lead to the bacteriophages being released prematurely in the small intestine and, thus, not reaching their host bacterium in the colon, in view of in vivo conditions.