The electrochemical behavior of cefotaxime (CTX +) was investigated at the polarized macro-and micro-interface between two immiscible electrolyte solutions (ITIES) by cyclic voltammetry and alternating current voltammetry. Miniaturization was achieved with fused silica microcapillary tubing entrapped in a polymeric casing. Scanning electron microscopy (SEM) was employed for the fabricated LLI support characterization. Voltammetric investigation of CTX + at macro-and μ-ITIES allowed the determination of many physicochemical parameters, such as formal Galvani potential of the ion transfer reaction (Δ aq org ϕ), diffusion coefficients (D), formal free Gibbs energy of the ion transfer reaction (∆G ′aq → org), and water-1,2-dichloroethane partition coefficient (logP CTX+ water∕DCE). Additionally, based on the results obtained the analytical parameters including voltammetric sensitivity, limits of detection and the limits of quantification (in micromolar range) were calculated. The applicability of the developed procedures was verified in spiked still mineral and tap water samples.