## 2 Methods for Determination of Lipophilicity
### Indirect Experimental Determination of Lipophilicity
#### 2.3.2 Reversed-Phase High-Performance Liquid Chromatography
\begin{table}
\begin{tabular}{c c c c} \hline \hline
**Analytes** & **Aim** & **Chromatographic Method** & **Chromatographic Conditions** & **Ref.** \\ \hline Structurally diverse & Estimate & & Column: 5–5.5 cm\(\times\)5 mm glass column containing \\ Structurally diverse & lipophilicity and & ILC/UV-Vis & EPC, PS–chol, and EPC–PS–PE liposomes & \\ drugs & correlate it with & (220 nm) & CNBr-Sepharose gel by avidin–biotin binding; MP: 0.01 M HEPES with NaCl (0.15 M\({}_{J}\)) at pH 7.4; FR: 0.3 mL/min & [192] \\  & absorption & & 0.3 mL/min \\ \hline  & & & Columns: Spherisorb ODS-2 (125 \(\times\) 4 mm, 5 \(\upmu\)m), \\  & & & IAM.PC.DD2 (100 \(\times\) 4.6 mm, 10 \(\upmu\)m), Purosphere \\ Fused azasiocytosine & Compare between & & RP-HPLC, IAM, \\   analogues & RP-HPLC, IAM, & IAM-HPLC, \\  & and MLC & MLC/UV-Vis & phosphate citrate buffer at pH 7.4 with Brij 35 \\  & & (0.15-0.075 M) and with isopropanol (7%); mixtures & of ACN (20–50\%) and 0.01 M phosphate citrate buffer at pH 7.4; FR: 0.1, 1 and 1.3 mL/min \\ \hline Series of & Study the effects & & Column: RP cyanopropyl Spherisorb (150 \(\times\) 4.6 \\  & of the variations & MLC/UV-Vis & mm, 5\(\upmu\)m); MP: 0.1 M phosphate buffer with SDS at \\  & in pH in MLC & determination & pH 3.0, 4.0, 5.0, 5.8, 6.4 and 7.0; FR: 1.35 mL/min \\ \hline Benzene derivatives & Study the & & Column: Nucleodur C18 Gravity (2 \(\times\) 125 mm, \\  & influence of & & 5 \(\upmu\)m); MP: HCl solution at pH 2 with different Brij 35/Triton X-100–ratios (0.05–0.30%); FR: 0.3 mL/min & [195] \\  & mixtures of Triton & & \\  & X-100 and Brij 35 & & 0.3 mL/min \\ \hline  & Estimate & & \\ Structurally diverse & lipophilicity and & & Column: Grace GraceSmart C18 (150 \(\times\) 2.1 mm, \\  & correlate it with & MLC/UV-Vis (210 & 3 \(\upmu\)m); MP: 0.05 M phosphate buffer at pH 7.4 with \\  & intestinal & & 0.01 M miltefosine; FR: 0.2 mL/min \\  & absorption and & & \\  & BBB & & \\ \hline Structurally diverse & Estimate & & Column: Kromasil octadecyl-silane C18 \\  & lipophilicity and & MLC/UV-Vis & (150 \(\times\) 4.6 mm, 5 \(\upmu\)m); MP: 0.05 M phosphate \\  & correlate it with & buffer at pH 6.5 and 7.4 with 0.04 M Brij35; FR: 0.5 & \\  & absorption & & mL/min \\ \hline Structurally diverse & Estimate & & Column: Spherisorb (15 cm \(\times\) 4.6 mm, 5 \(\upmu\)m); MP: 0.05 M phosphate \\  & lipophilicity and & MLC/UV-Vis (240 & buffer at pH 6.5 and 7.4 with 0.04 M Brij35; FR: 0.5 \\  & drying & & mL/min \\  & absorption & & mL/min \\  & absorption & & \\ \hline Structurally diverse & Estimate & & Column: Kromasil octadecyl-silane C18 \\  & lipophilicity and & MLC/UV-Vis (240 & (120 \(\times\) 4 mm, 5 \(\upmu\)m), Kromasil octadecyl-silane \\  & drying & & ODS-2 C18 (50\(\times\)4.6 mm, 5 \(\upmu\)m), Kromasil octadecyl-silane \\  & correlate it with & or 254 nm) & octadecyl-silane C18 (150 \(\times\)4.6 mm, 5 \(\upmu\)m); MP: 0.05 M phosphate buffer at pH 7.4 with \\  & absorption & & 0.04 M Brij35; FR: 1 and 1.5 mL/min \\ \hline  & Estimate & & \\ Anticonvulsant drugs & lipophilicity and & MLC/UV-Vis (220 & 5 \(\upmu\)m); MP: 0.05 M phosphate buffer at pH 7.4 with \\  & pharmacokinetic & & 0.04 M Brij35; FR: 1 mL/min \\  & parameters & & \\ \hline \hline \end{tabular}
\end{table} 
Table 1: _Cont._As an indirect measurement of lipophilicity, RP-HPLC does not directly provide a partition coefficient but rather a lipophilicity index which is correlated with the partition coefficient. The lipophilicity index is inferred by the assumption that compounds are retained in proportion to their nonaqueous-aqueous partition coefficient, with hydrophilic compounds eluted first and lipophilic compounds last, and it is expressed by the retention factor k [30].