The ion transfer reaction of 2-benzoylpyridine-thiosemicarbazone (HL), which has antimicrobial and antifungal properties and anticancer activity, has been studied to determine its lipophilicity by cyclic voltammetry at the water/1,2-dichloroethane (1,2-DCE) interface. The physicochemical parameters such as standard partition coefficient (lgP I ) and the standard Gibbs energy of transfer ( Δ G tr , I 0 , w → o ) of the protonated form of the ligand were measured as a function of pH in aqueous phase. The protonated form of the ligand exhibited reversible or quasi-reversible voltammograms at the 1,2-DCE in the range of pH 1–5. The protonation constants of the ligand, pK a1 and pK a2 , were determined spectrophotometrically and were found to be 12.14 and 3.24, respectively. The standard Gibbs energy of transfer ( Δ G tr , N 0 , w → o ) and the partition coefficient of neutral species (lgP N ) were also determined by the shake-flask method. The standard Gibbs energy of transfer of this compound across the water/1,2-DCE interface was evaluated as the quantitative measure of its lipophilicity. The difference between lgP I and lgP N was related to the degree of charge delocalization and was used to evaluate qualitatively the lipophilicity of the ligand.