Inhibitory compounds formation after liquid hot water (LHW) pretreatment of corn stover as an alternative to wood lignocellulosic feedstock for bioethanol production. Thus far, corn stover has been perceived as a promising lignocellulosic alternative to wood intended for bioethanol procurement, however it should be recognised also as a potential future component in a mixed biomass system. The aim of this research was to investigate the effect of applying different hydrothermal treatment conditions on the potential inhibitory compounds formation from corn stover. An analysis of selected inhibitory compounds formed after pretreatment performed at different temperatures (160°C, 175°C, 190°C and 205°C) was carried out. Furfural, simple sugars and lignin were some of the inhibitors examined with HPLC and UV-VIS spectrophotometric methods. Furthermore, the chemical composition of organic extracts obtained from native and LHW pretreated biomass was analyzed qualitatively with GC-MS method and inhibitory compounds like vanillin, sitosterol or syringol were detected. As a result of those investigations compared to enzymatic hydrolysis yield the temperature of 175°C was chosen as the most promising condition of corn stover LHW pretreatment in terms of the efficiency of the subsequent phases of bioethanol production.