## 4 Discussion
pendula_ extract, our findings are in line with the existing literature. Chemical compounds in birch bark, such as betulin and other triterpenes, have demonstrated antibacterial properties and may inhibit the growth of various bacterial strains [109]. Polyphenolsfound across the _Betula_ L. genus include catechin, salidroside, rhododendrin, and platyphyloside, which are known for their antioxidant, anti-inflammatory, and antimicrobial properties [110]. The interaction extends beyond bacteria; extracts from birch bark have also shown antiviral and antifungal activity against various pathogenic strains [51]. The presence of salicin is a significant factor contributing to the widespread investigation and utilization of _Salix_ species extracts in therapeutic applications. Alicin, along with other phenolic compounds such as procyanidins, can inhibit bacterial growth through various mechanisms, including antioxidant activity [111]. However, other bioactive compounds present in willow bark may also play a significant role in its antibacterial activity. Research by [PERSON] et al. has identified quercetin glycosides, monomeric, dimeric, and trimeric flavan-3-ol derivatives, including B-type procyanidins, as well as cavoylquinone pseudodepsides, all of which may contribute to the observed antimicrobial effects [63]. Our study also revealed that the combination of plant extracts exhibiting antimicrobial activity in the initial screening phase can lead to enhanced inhibition of pathogen growth. All extract mixtures were prepared in a 1:1 (_w/w_) ratio, and the same total volume (100 \(\upmu\)L) was applied in each assay. This means that the individual extracts were present at half the concentration used in the single-extract tests. Therefore, any observed increase in antimicrobial activity in the combinations, despite the lower concentration of each extract, may indicate an increased effect. Notably, five of the six extract mixtures that demonstrated increased antibacterial effects contained at least one bark extract. A similar observation was made in the study by [PERSON] et al., where the combination of extracts from _Solanum torum_ and _Uncaria gambir_ significantly augmented their antibacterial activity against methicillin-resistant _S. aureus_ strains [112]. Similarly, [PERSON] et al., in their research, highlight that utilizing the synergistic effects of plant extracts may have potential therapeutic applications [113]. Individual extracts within the mixture may target distinct cellular mechanisms of the pathogen. For instance, one extract could disrupt the integrity of the bacterial cell membrane, thus facilitating the penetration of a second extract, which could act on other key metabolic processes, such as protein synthesis or DNA replication. Alternatively, one of the extracts may inhibit the pathogen's resistance mechanisms, such as efflux pumps, thus enhancing the effectiveness of the second component within the mixture [114; 115]. Several factors may explain the lack of confirmed antimicrobial activity for the seven extracts despite existing literature reports. The primary factors include the plant species, the type of material used, and the storage conditions of the raw material [116]. The method of extract preparation is also crucial. As noted by [PERSON] et al., methanolic extracts consistently exhibited higher antimicrobial activity compared to ethanolic or aqueous extracts. They attributed this difference to the solvent's ability to dissolve a greater number of bioactive compounds [117]. The selection of strains is equally important, as their origin can significantly influence the variation in response to the applied extracts. The limitations of our study include the use of a single extraction method (ethanol:water), which may not have captured the full spectrum of bioactive compounds. Additionally, the thermal processing of extracts at 65 \({}^{\circ}\)C could have led to the degradation of heat-sensitive compounds, potentially reducing their antibacterial activity.