# Synthesis, Structure, and In Vitro Biological Evaluation of Semi-Synthetic Derivatives of Betulin
## 3 Results and Discussion
### Lipophilicity--Experimental and Theoretical Studies  
Lipophilicity is one of the most important physicochemical parameters related to the pharmacodynamic, pharmacokinetic, and toxic properties of medicinal substances. It is of key importance in the initial stage of developing new drugs and plays a role in binding ligands to proteins and determining the ADMET (absorption, distribution, metabolism, excretion, toxicity) characteristics that are the basis for determining the therapeutic dose. These properties affect the passive permeability of the membrane, which determines the drug absorption process after oral administration, as well as the possibility of access to intracellular areas and penetration into tissues. Determining theoretical and experimental lipophilicity values allows for the design of compounds characterized by appropriate potency obtained using a low dose of the drug [40]. Lipophilicity is most often presented as the decimal logarithm of the partition coefficient P (log P), which expresses the affinity of the active compound molecules for the organic and aqueous phases. Reversed-phase thin-layer chromatography (RP-TLC) is a commonly used chromatographic method for determining the lipophilicity of drug-like compounds. This method is characterized by the ability to analyze a large number of compounds in a relatively short time [41]. Chromatographic lipophilicity parameters R\({}_{\text{M0}}\) for \(\alpha\)-lipoic acid, betulin, 3-acetylbetulin **2**, 3,28-diacetyl-30-hydroxybetulin **3**, 28-acetylbetulin **5**, and the \(\alpha\)-lipoic esters of triterpenoids **6-10** were determined using the RP-TLC method. The value of the R\({}_{\text{M0}}\) parameter of the tested compounds was calculated from the retardation factor (R\({}_{\text{f}}\)) according to Equations (1) and (2) described in Section 2.2. R\({}_{\text{M0}}\) and b in Equation (2) represent the intercept and slope of the simple linear regression between acetone concentration (C) and R\({}_{\text{M}}\), respectively. The R\({}_{\text{M0}}\) and log P\({}_{\text{lit}}\) values of the reference substances were used to establish a calibration curve (Supplementary Materials, Table S1), based on which the experimental log P\({}_{\text{TLC}}\) values of \(\alpha\)-lipoic acid, betulin, and derivatives **2-3**, **5-10** were calculated. The experimentally determined values of lipophilicity (R\({}_{\text{M0}}\), log P\({}_{\text{TLC}}\)) are presented in Table 1.  
The values of log P\({}_{\rm TLC}\) of the \(\alpha\)-lipoic esters of triterpenoids **6-10** are in the range of 7.66-9.34. The most lipophilic compounds are diesters **7** and **9** with an acetyll and (1,2-dithiolan-3-yl)pentanoyl groups at the C3 and/or C28 position. The introduction of the (1,2-dithiolan-3-yl)pentanoyl moiety at the C30 position into the structure of 3,28-diacetyl-30-hydroxybetulin **3** (log P\({}_{\rm TLC}\) = 7.25) leads to an increase in the lipophilicity parameter of derivative **8** (log P\({}_{\rm TLC}\) = 8.12). The lowest lipophilic properties from the group of \(\alpha\)-lipoate esters were found for compound **6** (log P\({}_{\rm TLC}\) = 7.66), with a hydroxyl group at the C3 carbon atom. From six computer programs, in the case of the analog of compound **6** with a hydroxyl group at position C28 (derivative **10**), an increase in the log P\({}_{\rm TLC}\) value is visible (log P\({}_{\rm TLC}\) = 8.29). The theoretical lipophilicity values (log P) of the tested compounds were obtained (XLOGP2, MLOGP, ALOGPs, ALOGP, and XLOGP3) based on various computational algorithms. The values of log P ranged from 7.00 to 11.45 (Table 2). This indicates agreement with the results obtained from the RP-TLC method. For triterpenoids **6-10**, the lowest lipophilicity log P values were obtained using ALOGPs, while the highest log P values were obtained using XLOGP2 and XLOGP3. The last stage of the lipophilicity analysis concerned a comparison of the theoretical log \(p\) values of the tested compounds.