## 4 Discussion
Our in vitro simulated digestion study shows that the inclusion complex of propolis with HP-\(\beta\)-CD is the more convenient and efficient form to prepare the raw propolis sample for bioaccessibility studies of polyphenols. The conditions and protocol of the in vitro method described in this study were chosen based on the standardized COST INFOGEST method [29], which is widely applicable to both human and animal organisms. By simulating in vitro digestion in different foods or plant extracts, several simulation models for determining the bioaccessible content of polyphenols in IF were established, using centrifugation or dialysis membrane [21; 23; 35; 36]. Accordingly, we tested the propolis-HP-\(\beta\)-CD complex in the small intestinal phase on both models. In IF-C, the sample from the intestinal phase was centrifuged, and its supernatant represents the soluble compounds which can potentially be absorbed into the organism. In the dialysis model, the dialyzable fraction (Din) represents the amount of the sample that has passed through the semipermeable membrane and is available for absorption from the small intestine into the organism, i.e., it is bioavailable. On the other hand, the fraction outside the membrane (Dout) represents the amount of the sample that is further digested in the large intestine. As reported earlier by [PERSON] and [PERSON] [37], molecules of polyphenols are stable in acidic conditions and in this study the complex showed stability in GF compared to OF. Moreover, our study demonstrated that the propolis-HP-\(\beta\)-CD complex has good stability under OF conditions, most likely because hydrolysis by \(\alpha\)-amylase did not occur. In the IF, the polyphenols remained stable and no degradation occurred due to the influence of slightly alkaline pH (around 7.5-8.0), showing that the polyphenol molecules in the complex propolis-HP-\(\beta\)-CD complex have continuous stability during all three phases of GID (Tables 2-4). This finding was confirmed in previous study, reported by [PERSON] et al. [38], on olive pomace extract encapsulated with several cyclodextrin derivatives, in which bioaccessibility after small intestinal phase was also above 100%. Despite the relatively high values of bioaccessible polyphenols after IF-C, their dialyzability through the semipermeable membrane (Din fraction) turned out to be lower, and a considerable content of bioaccessible polyphenols was obtained in the Dout fraction (Tables 2-4). Wepresume that the impact on cyclodextrin ability to pass through biological membranes can be very variable. As a matter of fact, cyclodextrins enhance the oral bioavailability of poorly soluble compounds in the water, but in some cases, they have a limiting role [39; 40]. The bioaccessibility of TPs, TFFs, and TFDs in Vlp and Zg propolis samples after the intestinal phase with centrifugation has shown to be higher than in the dialysis phase by 0.73/1.38, 0.3/8.86, and 0.35/0.02 times, respectively (Figure 2). Moreover, the centrifugation model showed that the two propolis complexes had similar bioaccessibility profiles without statistical significance. Nevertheless, from the results of TFFs content in Din fraction (Table 3), the differences between Vlp and Zg propolis samples can be noticed. That indicates that the propolis origin might reflect their content to an extent. Even though our study gains valuable insights into polyphenols' bioavailability in propolis, the differences in the content of TFFs in propolis samples observed after gastric and intestinal digestion suggest the need for additional analyses. The result should be confirmed by further research conducted on a large number of propolis sampled from various continental areas. The presented results are credible since they were obtained by reliable, validated procedures and analyzed by acceptable statistical methods. The spectrophotometric methods enabled the quantification of the polyphenolic contents of the main polyphenolic groups in the poplar type of propolis during in vitro GID study.