## 4 Discussion  
Hidden beneath their protective extracellular matrices, multi-cellular microbial communities, referred to as biofilms, display elevated tolerance levels toward the immune system and medicinal measures. In the current work, we scrutinized the potential of octenidine dihydro-containing lozenges against five pathogenic biofilms of the oral cavity and throat formed by _S. aureus_, _S. pyogenes_, _P. aeruginosa_, _C. albicans_ and _A. actinomycetemcomitans_. These microorganisms differ with regard to the structure of their cell wall--_S. aureus_ and _S. pyogenes_ are Gram-positive, _Paeruginosa_ and _A. actinomycetemcomitans_ are Gram-negative bacteria and the cell wall of the _C.albicans_ fungus differs essentially from the bacterial cell walls as it is composed of (among others) glucans, chitin and glycoproteins. This wall-based differentiation of applied microorganisms is of a pivotal matter regarding the active substance of analyzed OCT lozenges, namely, octenidine dihydrochloride. This is so because the mechanism of action of this cationic antiseptic relies on its adhesion to the outermost microbial structures (cell walls, membranes) and their alteration to an extent which leads to their discontinuity, causing the subsequent leakage of the cytoplasm and, finally, the death of microbial cell, finally [14]. The spectrum of octenidine activity covers Gram-negative and Gram-positive bacteria (but not the spores) as well as fungi and specific types of viruses. Another rationale behind the choice of the five above-mentioned microbial species for analysis was the fact that all of them are causative factors for a variety of biofilm-based infections in the oral cavity and also (in the case of _S. aureus_, _S. pyogenes_, _P. aeruginosa_ or _C. albicans_) in the throat [15; 16; 17; 18]. The main goal of the present research was to investigate the impact of the specific type of formulation (lozenge) containing 2.6 mg of octenidine dihydrochloride on the drug's antibiofilm activity. It is generally accepted that, with regard to the hygiene of the oral cavity (understood as both prophylaxis or eradication of existing infection), solid carriers (such as lozenges) display significant advantages over liquid antiseptics [19]. This is because in the case of the last type of formulation mentioned (applied in a variety of so-called antiseptic mouth washes), the contact time of the bacteria to antiseptic substance rarely exceeds a few dozen seconds. After this time, the mouth wash needs to be removed from the oral cavity and its leftovers should be washed out with an abundance of water. On the contrary, solid carriers such as tablets or lozenges release the antiseptic into the oral cavity and subsequently to the throat over a more controlled, longer period (a dozen or so minutes, most often). Moreover, the antiseptic is naturally distributed by saliva throughout the oral cavity and streams down to the throat. The measured time period for the OCT lozenge to completely dissolve in the oral cavity was ca. 22 min, and such a time was established as the contact time (exposition time) in the experimental settings in this research utilizing biofilms pre-formed on HA or agar surfaces. Nevertheless, in the first line of experimentation, the standard microdilution method was used to check if the octenidine dihydro related from the OA maintains its antiseptic functions (Table 1). As a comparator, an octenidine-containing liquid antiseptic (OL) was applied. This antiseptic, being a commercially available product, also contains in its composition a phenoxyethanol (PHE) additive (of weak antimicrobial activity when applied as a stand-alone agent). However, PHE significantly boosts the antimicrobial activity of octenidine dihydrochloride [20]. Because the composition of OCT contains no PHE, it seemed rational to compare the activity of these two medicinal products. Interestingly, there were no significant differences in the observed MBC and MBEC values between OCT and OL in this experimental setting.