The results of physical parameters of the prepared samples
The results of physical parameters of the prepared samples are summarized in Table 2. The equivalent diameter ranged from 1.09 to 1.14 mm, which is within the required size distribution of 0.8–1.25 mm. A slight increase in size is probably caused by increasing the thickness of the second coat. To determine the precise appearance of the carriers, SEM photos of the sample E2.5 were taken according to the methodology in the Section 2.8. From Fig. 1 it is obvious that the double coating is uniformly spread over the pellet core and remains intact during the analysis, which is a vital attribute to prevent leaching of reagents from pellets to a liquid media. Sphericity and Hausner ratio are parameters characterizing flow properties of carriers, which are important for filling the detection tubes. Sphericity can range from 0 to 1, where the sphericity between 0.8–1.0 means a sufficiently spherical shape . According to Ph. Eur. 9, Hausner ratio of 1.00–1.11 corresponds with an excellent flow character of particles. In both cases the tested samples achieved excellent results. Easy filling into detection tubes, occupying only a small volume by carrier particles, forming a compact layer and facilitating easier observation of the color transition could be expected for all the samples. Measured values of pycnometric density showed a slight decrease with the increasing thickness of the second coat. Eudragit® RL used for the preparation of the second layer is commonly intended for controlling the release of drugs by BKT140 through the coating. This polymer enables preparing homogeneous coatings without pores and it can be supposed that the used gas did not penetrate inside the pellets during the measurement. Fig. 1 confirms the theory of creating the second coating without pores. Thus the mentioned slight decrease probably means that the density of the second coat was lower than the density of the core and the first coat. Interparticular porosity indicates the amount of air gaps between the individual pellets and correlates with the flow properties of the carriers. The lower the values are, the less space the pellets occupy and create a more compact layer when filled into the detection tubes facilitating an easier detection. Interparticular porosity was almost similar for all samples varying from 40.35% up to 41.17%. Sufficient mechanical resistance was characterized by hardness and friability, which ensured a stable quality of the carriers during their filling into glass tubes, transportation and storage . The hardness of pellets was determined for non-coated pellet cores because of the possible coat rupture during the measurement of coated pellets. Obtained values of hardness (Table 2) were more than sufficient compared to other studies [, , ]. Pellets with the friability values of <1.7% were judged as mechanically acceptable . From this point of view, all the pellet samples exhibited excellent mechanical properties and the friability values were below 0.2% (Table 2). For the purpose of enzyme activity and inhibition, testing the standard Ellman’s reaction was utilized. Preparation of all used solutions is described in chapter 2.4 and the methodology of these tests in Section 2.5. The most important measured aspects were the time at which the pellets changed to an observable yellow color and the intensity of that color. For the reason of standardization of this process and colors, photos were taken at given time points (see Section 2.6). The color transition from a white to yellow/orange color is observable in Fig. 2. The colors were standardized via RGB values stated at the bottom of the figures. The visible color change occurred within 48 ± 3 s for E1.5, 58 ± 4 s for E2.5, 80 ± 2 s for E5.0 and 108 ± 5 s for E7.5 after adding the test solution to the moistened pellets during the test of enzyme activity. The highest color intensity was shown by sample E2.5, as is demonstrated in Fig. 2. This color even shifted from a standard yellow color to an almost orange color facilitating an easier visual detection. The thickness of the second coat significantly influenced the color intensity and speed of the detection. On the one hand, the thickness could not be too large; otherwise endothermy might have decreased the amount of the test solution which penetrated through the coat, and the overall speed of the reaction (samples E5.0 and E7.5). On the other hand, if the thickness of the coat was insufficient, the developed color would be indistinctive (sample E1.5). With respect to these reasons, the sample E2.5 showed the best results.