*2.2. Capsule Formation*

Schematic presentation of the capsule formation process is shown in Figure 1.

**Figure 1.** Capsule formation scheme.

The preparation method of the film-forming mixtures and films was described in detail in the previously published work [5,6]. Shortly, the mixture of components (Table 1) was stirred at 80 ◦C for 2 h, which was followed by deaeration under vacuum. Afterwards the mixture was casted on a glass plate using a plate coating device (Camag TLC Plate Coater, Camag, Muttenz, Switzerland) with a height of fluid layer of 1500 μm. After drying the thickness of the film was around 600 μm.


**Table 1.** Compositions of the films.

1 non-modified film (reference); 2 modified binary (GA); and ternary (GAC) polymer films.

The capsules were prepared using GAC composition (Table 1), by placing 2 pieces of the film (immediately after casting) in a steel form for suppositories. After closing the form, the resulting reservoirs were filled with: (a) MCT oil, (b) PEG 400 or (c) cetearyl alcohol. For a better visual identification of a disintegration test endpoint, the filling material was colored with small amount of a hydrophilic or lipophilic dye. Afterwards, the filling orifice was manually closed with a strip of a film, and to ensure good sealing the capsules were placed for 5 min at 60 ◦C. Finally, the capsules were stored and dried at ambient temperature and of 15–25% RH for at least 24 h. The measured moisture content in the capsules was around 2.5% (Radwag WPS210S Moisture Analyzer, Radwag, Radom, Poland).

#### *2.3. Microscopic Imaging*

The imaging of samples was performed with use of a scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), confocal Raman microscopy and optical microscopy.

The observation of film samples was performed before and after submersion in 0.1 M HCl at 37 ◦C, under constant stirring for 2 h (similar to the procedure of swelling test described in our previous work [5]). The films after submersion in acid were frozen in a liquid nitrogen and freeze-dried for 24 h. The investigation was performed with Jeol 7900F SEM (Jeol, Tokyo, Japan), Nikon Ti-E/A1 + CLSM (Nikon, Tokyo, Japan) and WITec Alpha 300 Access Raman microscope equipped with 785 nm laser (WITec, Ulm, Germany).

The imaging of the lab-manufactured capsules was performed using Phenom Pure SEM (Phenom World, Eindhoven, the Netherlands), and Nikon Eclipse 50i optical microscope (Nikon, Tokyo, Japan).

## *2.4. Gas Permeability*

The films GEL, GA and GAC (Table 1) were subjected to oxygen permeability tests, performed with an coulometric detector technique according to method ASTM F 1927-14. The equipment used was OX-TRAN 2-20 (Mocon, Minneapolis, MN, USA). The investigated surface was 50 cm2.

#### *2.5. Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D)*

QCM-D was employed to investigate the a ffinity of CAP latex particles present in Aquacoat CPD to gelatin. The preparation step comprised coating of the gold-plated quartz crystal sensor with branched polyethyleneimine (PEI), then spin-coating the sensor with 1% gelatin solution (5 s at 2500 rpm and low acceleration, followed by 60 s at 8000 rpm and high acceleration). Afterwards the sensor was dried at 60 ◦C for 20 min. The gelatin on the sensor was subjected to crosslinking by submersion in 1.5% formaldehyde solution, in order to prevent it from dissolving in aqueous conditions. Finally, the sensor was dried at 60 ◦C for 60 min.

The sensors were mounted in a Qsense equipment (Qsense, Västra Frölunda, Sweden). A deionized water (at 25 ◦C) was flushed over the sensors until a stable baseline was obtained. Then the diluted (0.1%) Aquacoat CPD at 25 ◦C was pumped through the cells, and the changes in fundamental frequency overtones of the crystal were registered. After stabilization of the system, the cells were once again pumped with deionized water to remove all the substances that were not bound to the film.

Additionally, to assess the surface structure and stability of the gelatin films obtained in situ on the sensors, the Atomic Force Microscopy (AFM) was performed with NTEGRA Prima setup (NT-MDT Spectrum Instruments, Moscow, Russia), with a silicon probe (spring constant of 40 N m<sup>−</sup><sup>1</sup> and resonant frequency of 300 kHz) (Tap 300AI-G, Budget Sensors, Sofia, Bulgaria). The images were analyzed using a Gwyddion software (Version 2.55, Free Software Foundation, Boston, MA, USA).

#### *2.6. Disintegration Time*

Disintegration time test of the capsules filled with PEG-400, MCT oil or cetearyl alcohol, was performed. The test was performed using: (a) a tablet disintegration tester ED-2SAPO (Electrolab, Mumbai, India); (b) a paddle dissolution apparatus DT800 (Erweka, Langen, Germany), with a capsule placed in a steel sinker (the stirring rate was 50 rpm). The capsules were tested for 120 min in 0.1 M HCl, followed by pH 6.8 phosphate bu ffer until disintegration.

#### *2.7. Drug Release Test*

The study was performed using a vertical di ffusion cell (Enhancer cell, Erweka, Langen, Germany) and a paddle dissolution apparatus DT800 (Erweka, Langen, Germany) equipped with a built-in autosampler. The stirring rates of 50, 100 and 150 rpm were used. The enhancer cell with the mounted modified gelatin film is shown in Figure 2.

The film selected for the test was gelatin + Aquacoat + carrageenan (GAC), the same as for disintegration tests. The di ffusion cell was filled with 2.5 mL of a 1% diclofenac solution in PEG 400 (the amount of diclofenac sodium was 25 mg). Then the investigated film (cut to a circle of 3 cm in diameter) was carefully placed on the top of the solution and secured with a sealing ring and a screw cap; the active surface was 4.15 cm2. The test was performed in 900 mL of 0.1 M HCl for 120 min followed by 900 mL phosphate bu ffer pH 6.8 for 60 min.

Sampling of the acceptor fluid was performed every 15 min in the acid phase, and every 5 min in the bu ffer phase. Quantification of diclofenac was performed spectrophotometrically at 276 nm wavelength. The study was performed in triplicates.

**Figure 2.** The enhancer cell with gelatin + Aquacoat + carrageenan (GAC) film.
