## 3 Results and Discussion
### Antimicrobial Activity
MBC and MFC values at 0.05% \(v/v\) for the oregano essential oil which, according to GS-MS analysis, contained 63% carvacrol, was directly comparable to the MBC and MFC value at 0.03 % \(v/v\) of pure (98% \(v/v\)) carvacrol used as a control. Identification of carvacrol as the major antimicrobial compound in oregano EO might be used to form a classification system of antimicrobial activity exerted by oregano EOs of even different chemical compositions. However, other compounds present in oregano EO might negatively affect antimicrobial activity. A recent study on the antibacterial activity against _Staphylococcus aureus_ demonstrated that thymol and carvacrol present in oregano EO exhibit antagonism [72]. Thymol and carvacrol ratio might vary in oregano essential oils [73], so it is imperative to conduct further antimicrobial studies on oregano EOs of different chemical composition. Despite the fact that wage EO in this study has exerted lower antibacterial activity (MBC 0.35% \(v/v\)) compared to oregano EO, a recent study has demonstrated increased activity against _Streptococcus pyogenes_ biofilm formation [74]. Furthermore, a clinical study showed that an implemented sage extract reduced the colony number of _S. mutans_ in dental plaque in 11-14 year old children [75]. Bacterial species like _S. mutans_ adhere to the primary colonizers, mainly _Streptococcus sanguus_ and _Actinomyces viscosus_, by cell-to-cell interactions, leading to biofilm formation on teeth [2]. Therefore, sage EO might be an essential compound in mouthwash solutions. Pennyroyal essential oil in this study exerted lower antibacterial activity at 0.45% \(v/v\) than antifungal activity against _C. albicans_ (0.25% \(v/v\)). Pennyroyal is an herb belonging to the Mentha genus. These herbs are widely used in EO production of high commercial value, exceeding USD 400 million. Moreover, Mentha species are used as additives in many spice mixtures used in foods because of their aroma and flavor [76]. The antimicrobial activity exerted by Mentha Eos, in combination with their intense aroma and flavor, make them attractive constituents in oral hygiene products. Nevertheless, according to European Medicines Agency (EMA), there is a maximum daily intake for pulegone, which is the major substance of pennyroyal essential oil (62.1%) [77]. Therefore, the implementation of pennyroyal EO should be limited, and the maximum daily intake should be taken into account in commercial product development. Although hydrosols are considered distillation byproducts [18,78], they exert some biological properties. Hydrosols are chemically different regarding the volatile compounds compared to EOs [18]. In this study, GS-MS analysis demonstrated that the main oregano HS volatile component is carvacrol, which exerts strong antimicrobial activity [60]. This finding implies that oregano hydrosol might be an ingredient of oral hygiene products exerting antimicrobial activity. Hydrools from sage and pennyroyals have not demonstrated any antibacterial or antifungal activity. However, they contain active volatile components, as depicted in the GS-MS analysis. A common usage of HSs in cosmetics is to replace water by adding active ingredients [18]. In the case of a mouthwash solution produced by natural ingredients, hydrosols might have the same application. HSs could make more palatable the flavor of the product, and might increase its antioxidant activity. In that respect, the application of hydrosols from sage, thyme, and peppermint as mouthwash ingredients clearly reduced mucositis in patients undergoing chemotherapy in a randomized controlled pilot study [79]. Nevertheless, further studies (including in vivo studies and clinical trials) must be carried out in order to fully elucidate the spectra and the mechanisms of antimicrobial activity exerted by EOs and their respective hydrosols distilled from a plethora of herbs grown in regions characterized by high plant diversity and certain (micro)climates (especially xerothermic) in order to develop commercial products.