## 3 Results and Discussion
### Biological Assays
#### 3.2.2 In Vitro Cytotoxicity Screening  
Another goal of this research was to set in vitro the impact of **6a-g** on a liver cancer HepG2 cell line. The model was chosen since antimyobacterial agents have been included within a group of the top five pharmacodynamic classes associated with the highest reporting odds ratio (ROR) score. In attempt to set this ROR score, the frequency of hepatotoxicity was estimated at both the group and individual drug levels. The ROR score index describes the measure of such undesirable phenomenon [73]. The highest ROR scores with the highest risk of drug-induced liver injury were observed for the compounds belonging to antineoplastic, antiviral, analgesic, antibiotic, and antimyobacterial classes. The molecules **6a-g** were highly lipophilic as their log \(k_{\rm w}\) values from 4.1170 (**6a**) to 5.2184 (**6g**) indicated (Table 2). Potential disadvantages of highly lipophilic bedaquiline (**BDQ**), a very effective antimyobacterial agent (Figure 5), include the inhibition of a human Ether-a-go-go-related gene (hERG) potassium channel due to the concurrent risk of cardiac toxicity, possible phospholipidosis, or hepatic toxicity. The shortening or lengthening of its dimethylaminoethyl sidechain did not improve antimyobacterial activity, but increased cytotoxicity was observed as the lipophilicity of synthesized **BDQ** derivatives increased [74, 75]. The current nonlinear regression analyses explored the relationships between a semi-logarithmic plot of incubation concentration and percentages of absorbance relative to untreated controls, thus providing IC\({}_{50}\) values [63]. These IC\({}_{50}\) parameters (in uM units) describing the cytotoxic properties of **6a-g** varied from 22.18 (6g) to 83.08 uM (6a) as listed in Table 3. The most effective compounds (**6g** and **6e**) against _Mtb_ H\({}_{37}\)Ra showed IC\({}_{50}\) = 22.18 (6g) and 29.39 uM (6e). If accepting IC\({}_{50}\) > 25.00 uM as a \"very mild\" IC\({}_{50}\) criterion characterizing low toxic derivatives [76], the cytotoxicity of **6e** might be \"theoretically acceptable\". However, [PERSON] et al. (2020) stated that the molecules with IC\({}_{50}\)\(\leq\) 100.00 uM were relatively cytotoxic [77]. The INH standard drug showed a very limited ability to reduce the population of HepG2 liver cancer cells (IC\({}_{50}\) > 1000 uM) [63]. The limited safety of effective antimycobacterial agents due to the relatively low IC\({}_{50}\) values was the reason for not progressing further. [PERSON] et al. (2019) stated that molecules with IC\({}_{50}\)\(\geq\) 40.00 uM were safe [78]. Following the conclusions published in [76; 78], both **6d** (IC\({}_{50}\) = 43.35 uM) and 1-[hydroxypoyl-[(3-trifluoromethyl)phenyl]carbamoyloxy]-4-(3-trifluoromethylphenyl)piperazine-1-ium chloride (**6f;** IC\({}_{50}\) = 54.71 uM) might be \"theoretically safe\". In regard to confirming or refusing the validity of statements about the cytotoxic properties of the compounds **6d-g** based on the conclusions published in [76; 78], previous in vitro investigations [30; 79; 80] also contributed to the current quest. [PERSON] et al. (2021) evaluated the in vitro cytotoxic properties of a tamoxifen (TMX) reference drug against the HepG2 cell line. The authors described the efficiency of TMX with IC\({}_{50}\) = 19.56 uM [63]. Other anticancer drugs, such as afatinib, 5-fluorouracilacil, and sorafenibib, showed IC\({}_{50}\) = 5.40, 7.91, and 9.18 uM [79; 80], respectively. The compound **6g** also efficiently inhibited the proliferation in vitro of a human monocytic leukemia THP-1 cell line showing IC\({}_{50}\) = 5.91 uM as the concentration, which inhibited 50% of the proliferation. Moreover, lower cytotoxic activity of a cisplatin standard drug was even observed (IC\({}_{50}\) = 13.65 uM) in those experiments [30].