## 3 Results and Discussion
### Cell Cytotoxicity Activity of T. canadensis Seed and Cone EOS  
In order to determine the viability of skin fibroblasts, keratinocytes, and melanoma cells (A375 and C32) after 24 h of treatment with five concentrations of essential oils (0.012, 0.025, 0.05, 0.1, and 0.2 \(\upmu\)L/mL), the MTT assay was used. As shown in Figure 2, _Tsuga_ seed is less cytotoxic than _Tsuga_ cone, and its IC\({}_{50}\) values are higher than 0.2 \(\upmu\)L/mL in all tested cell lines. Our previous studies on essential oils from seeds of various species of the _Abies_ genus belonging, like _Tsuga canadensis_, to the _Pinaceae_ family showed higher cytotoxicity in fibroblasts (IC\({}_{50}\) values about 0.11 \(\upmu\)L/mL) than _Tsuga_ seed [1; 2]. Similar to a previous study based on some _Picea EOs_, where cone EO exhibited stronger antimicrobial and cytotoxic effects compared to seed EO [14], the cone EO of _T. canadensis_ demonstrated significant cytotoxicity, particularly against keratinocytes and melanoma cell lines (A375 and C32), with IC\({}_{50}\) values as low as 0.104 \(\pm\) 0.015 \(\upmu\)L/mL. In contrast, _T. canadensis_ seed EO displayed minimal effects on cell viability, resembling the milder impact observed for _A. concolor_[9] and _P. pungens_[14]. Interestingly, IC\({}_{50}\) values for _Tsuga_ cone in both melanoma cells lines are significantly lower than in fibroblasts and amounted to 0.155 \(\pm\) 0.022 \(\upmu\)L/mL in A375 cells and 0.104 \(\pm\) 0.015 \(\upmu\)L/mL in C32 cells. When compared to standard chemotherapeutic agents, such as cisplatin and paclitaxel, this cytotoxic effect is significant. For instance, the Genomics of Drug Sensitivity in Cancer (GDSC) database reports that cisplatin exhibits IC\({}_{50}\) values in various cancer cell lines ranging from 0.177 to 1.02 \(\upmu\)M. Similarly, paclitaxel shows IC\({}_{50}\) values between 0.00126 and 12.1 \(\upmu\)M across different cell lines [25]. These comparisons suggest that the cytotoxic potency of _T. canadensis_ cone EO is comparable to these established chemotherapeutic agents. In contrast, viability of fibroblasts treated with 0.2 \(\upmu\)L/mL _Tsuga_ cone decreased only by 24%. However, the viability of keratinocytes is reduced by _Tsuga_ cone similarly to viability of melanoma cells, and its IC\({}_{50}\) value was 0.134 \(\pm\) 0.019 \(\upmu\)L/mL. The inclusion of fibroblasts in the study allowed us to understand that OEs used as, for example, fragrance ingredients in cosmetics can be used without significant damage to healthy cells. The study showed that cone EO reduced fibroblast viability by only 24% at the highest tested concentration, indicating its relatively low toxicity to healthy cells compared to melanoma cells. These results reinforce the importance of cone EO, especially as a potential source of bioactive compounds or for possible applications as additional substances as anticancer drug development. The radar plot (Supplementary Figure S2) highlights the potent cytotoxicity of _T. canadensis_ cone EO against melanoma cell lines (IC50: 0.104 \(\pm\) 0.015 \(\upmu\)L/mL for C32 cells) compared to milder effects observed in seed EO and related species.