Esomeprazole is the most effective of the proton-pump inhibitors for the acid-related diseases and at first was examined for the electroanalytical purposes. The drug standard and as a content of injection powder was investigated by cyclic voltammetry (CV) and quantitatively determined using square wave voltammetry (SWV) via its electrooxidation at Au electrode in 0.05M NaHCO3. SWV showed a linear dependency of the anodic peak currents vs. esomeprazole standard concentrations in the range from 3.0 to 500μgmL−1 with the values of limit of detection (LOD) and limit of quantification (LOQ): 1.4 and 4.6μgmL−1, respectively. Using the constructed and validated calibration curve, the values of unknown esomeprazole concentrations in injection powder and in human serum spiked with standard were determined. Before the electrochemical oxidation, it was shown by atomic force microscopy (AFM) that the small esomeprazole islands formed inside holes were visible and their diameter was about 200nm attributed to physico-chemical characteristics of esomeprazole. After the electrochemical oxidation, the morphology of esomeprazole standard on Au surface was completely changed and composed of spherical particles in a diameter between 200 and 600nm. With esomeprazole suspended in human serum, the process of crystallization partly occurred in the form of spherical grains with the average size of these grains was about 4μm. The analysis at the macro level done by the optical microscopy (OM) confirmed this opinion. The study of esomeprazole degradation showed that at Au electrode, after 3h of cycling, a neglectable amount of the esomeprazole was changed. Using IrOx electrode under directed stress conditions, its almost complete degradation was realized after 3h confirmed by high performance liquid chromatography (HPLC). Total organic carbon (TOC) analysis showed that 95% of esomeprazole was mineralized. The HPLC and Liquid chromatography-mass spectrometry (LC-MS) study revealed the formation of 4-hydroxy omeprazole sulphide, 4-hydroxy omeprazole sulphone, esomeprazole sulphone and methylated esomeprazole. Display omitted •SWV method for esomeprazole determination using Au electrode was developed.•Method was applied for esomeprazole determination in human serum/injection powder.•Morphology of Au electrode surface during study was characterized by AFM and OM.•Esomeprazole degradation at IrOx electrode led to 95% of its mineralization.•Several degradation products were detected by HPLC and LCMS analysis.