## 3 Results and Discussion
### Chemistry
#### 3.1.2 Determination of Some Physicochemical Properties  
* Determination of electronic properties The UV/Vis absorption spectra of methanolic solutions (_c_ = 5.0 \(\times\) 10\({}^{-5}\) M) of **6a-g** were measured in the \(\lambda\) interval of 190-820 nm. Three absorption maxima (Table 1) were found for these solutions as follows: \(\lambda_{1}\) = 203.50-206.00, \(\lambda_{2\text{ (CH-T)}}\) = 238.50-243.00, and \(\lambda_{3}\) = 279.50-286.50 nm. This observation was in full agreement with [53], as the positions of particular absorption bands were typical for (substituted) phenylcarbamic acid derivatives. The observed bands were labeled properly as first local excitation (\(\lambda_{3}\)), charge-transfer (\(\lambda_{2\text{ (CH-T)}}\)), and second local excitation (\(\lambda_{1}\)) absorption maxima, respectively. Almost all substituents attached to the aromatic of a 4-(substituted phenyl)piperazine-1-ium-1-yl moiety, which carried non-bonding electrons, caused shifts in the \(\lambda_{1}\) and \(\lambda_{2}\) as well as \(\lambda_{3}\) absorption bands to longer wavelengths. Thus, bathochromic shifts at these absorption maxima were found for the compounds **6b-g** compared to those of **6a**, a molecule containing a 4-phenylpiperazine-1-ium-1-yl group. This trend was in agreement with conclusions already published [54]. In fact, exceptions were observed only for the molecule **6b** and its \(\lambda_{1}\) absorption band (equal values were find out) as well as **6g** and its \(\lambda_{3}\) absorption band, for which a slight hypsochromic shift was observed (Table 1). The log \(\varepsilon_{2\text{ (CH-T)}}\) parameter was the most sensitive to the differences in length, position, electronic, and steric properties of substituent(s) attached to a piperazin-1-ium-1,4-diyl moiety. Thus, when discussing SARs, special consideration was paid to log \(\varepsilon_{2\text{ (CH-T)}}\). Absorption intensity at this maximum increased slightly for the compounds containing bulky substituents (auxochromes) with strong EW properties (**6e**, **6f**) and molecules with a bulky diphenylmethyl moiety (**6g**). This behavior is called a hyperchromic shift [55]. On the other hand, a hypochromic shift [55] (i.e., decreased values of a given parameter) was observed for mono-substituted derivatives (Table 1) containing a 4'-CH\({}_{3}\), 4'-F or 4'-Cl substituent (**6b-d**). * Determination of lipohydphilic propertiesPathogenic _Mtb_ strains possess an external cell wall composed mainly of a covalently attached complex of a branched polysaccharide peptidoglycan, lipopolysaccharideascharides (arabinogalactan, lipoarabinomanan, and lipomannan), and highly lipophilic mycolic acids. These structures have important roles in the growth and integrity maintenance of the cell wall. In addition, _Mtb_ has an outer layer composed of an array of glycolipids, lipopolysaccharide, and lipids associated with this wall [56; 57; 58]. Thus, the lipophilicity is an important parameter that should definitely be taken into consideration in the design of novel anti-TB compounds to ensure their permeation via the cell wall of _Mtb_[59]. Computational methods (i.e., CLOGP, ALOGP, XLOGP2, or XLOGP3) are frequently used to calculate or predict the lipophilicity of various synthetic or natural antimyobacterial compounds [60; 61; 62; 63; 64; 65]. However, it would be questionable if such in silico approaches might be considered accurate in predicting the lipophilic behavior of the compounds **6a-g**. The structure of the molecules investigated in [60; 61; 62; 63; 64; 65] allowed for such predictions; however, particular predictors were not designed to evaluate salts, i.e., charged compounds as discussed in [31]. Taking into strong consideration this aspect, the lipohydrophilic features of **6a-g** were investigated by an experimental RP-HPLC technique. The retention behavior of a molecule in a reversed-phase (RP) chromatographic system is in very close connection with its lipophilicity commonly derived from \(\log k\)[66].