## 2 Materials and Methods
### Encapsulation of Probiotics  
The encapsulation of the probiotics (_Lactobacillus casei_ bacteria) was performed according to the methodology described in our previously published article [10] and patent application P. 443812 [35]. Briefly, a pre-emulsion was prepared by mixing the encapsulating material (2% sodium alginate solution and bacterial suspension in a 4:1 ratio) with capric/caprylic triglycerides and 1% ECO-Tween 80. An aqueous calcium chloride solution was added drop by drop to the solution in order to cross-link the microspheres. For the purposes of separating the obtained microspheres from the microdispersion, a centrifugation process was then conducted (EBA 20 by Hettich Zentrifugen). The composition and production methodology of the microspheres have been precisely described in our previous studies [10]. For further tests, we selected samples of the microspheres without centrifugation (1); microspheres spun at 500 rpm (2) and 30,000 rpm (3); and a stable microdispersion (4). Assessment of the Viability of Free and Encapsulated Probiotic Cells Depending on the Influence of Temperature, pH, UV Radiation and Preservatives  
Aqueous solutions containing 0.9% sodium chloride (\(w/v\)) were prepared, and their pH level was adjusted to the values of 2.0, 5.5 and 9.0 with sterile 1 mol/L HCl or NaOH as well as the preservative solution--sodium benzoate combined with sodium salts of levulinic and anisic acid. In order to assess the survivability of bacteria in solutions with different pH values and the preservative solution, 1 g of encapsulated probiotics containing 9-11 log CFU of LAB cells was mixed with 9 mL of prepared solutions and incubated at room temperature (20 \({}^{\circ}\)C) for 15, 30 and 60 min. After incubation, the capsules were collected and put into 9 mL of 0.2 mol/L sterile sodium citrate (pH = 6). Cell viability was assessed through pour plate inoculation and the counting of bacterial cells grown on plates with MRS agar at 37 \({}^{\circ}\)C for 72 h under aerobic conditions [36]. A similar test was performed using free cells in order to compare the viability of free and encapsulated probiotic cells under similar conditions. A volume of 1 g of cell suspension with the live cell count amounting to 12.6 log CFU/g was mixed with 9 mL of the preservative solutions at various pH values. After incubation (15, 30 and 60 min), cell viability was assessed in line with the methodology described above. In the experiments related to the assessment of the influence of temperature and UV light, free and encapsulated probiotic cells were kept at 40 \({}^{\circ}\)C for 1 h and 96 h and at 50 \({}^{\circ}\)C for 10 min and were treated with UV light. The assessment of the impact of UV radiation on microcapsules was made according to the methodology proposed by [PERSON] et al. [37] with minor modifications. Samples (5 g) of each microencapsulated and free cell suspension (control) were placed on glass Petri dishes. The plates were then exposed to long-term UVB irradiation (311 nm) for 24 and 48 h. The UV sources were located 20 cm from each sample. Next, 1 g of encapsulated cells or 1 g of free cell suspension (initial density: 12.6 log CFU/g), stored under the conditions described above, was placed into 9 mL of 2 mol/L sterile sodium citrate (pH = 6) or 0.9% sodium chloride (\(w/v\)), respectively. After a series of dilutions, the viability of free and encapsulated cells was determined and compared. The same procedure was applied to microcapsules and free bacterial cells exposed to specific temperature conditions. The experiments were repeated two times, and the results were used to calculate the average of log reduction. LAB cell reduction was calculated as the difference between the initial number of free LAB cells or LAB cells contained in the microspheres (No) and the number of free LAB cells or LAB cells contained in the microspheres exposed to physicochemical factors for a specific time (Nt).