Due to the raising requirements of drug quality, there is an increasing need for fast liquid separations of pharmaceutical substances with high efficiency and good resolution. The ultra-high pressure liquid chromatography (UHPLC) has been considered to meet this challenge. However, it was found that this fast method has also serious disadvantages. The range of applications of the UHPLC in the analysis of pharmaceutical substances and dosage forms is currently extensively discussed. In this study we investigated the consequences of the shortening of the analysis time of the liquid chromatographic method. Bosentan, a non-peptide antagonist of human endothelin receptors, was chosen as an example in this study, due to its therapeutic importance and lack of the reported analytical methods of the drug product. Two high-performance, reversed phase liquid chromatography methods with UV detection at 220 nm were developed for this purpose. Both methods were validated and the resulting performance characteristics were compared. The first separation (method A) was achieved on Kinetex column, (2.6 μ C18 1OOA, 150 x 4.6 mm), the second-fast (method B) employed Kinetex column, (1.7 μ XB-C18 100A 50 x 3.0 mm). Both methods were performed with a buffered mobile phase containing 0.1% of triethylamine in water brought to the pH 2.5 with phosphoric acid and methanol as the solvent A and acetonitrile as the solvent B. Gradient program was used and flow rate of 0.8 mL/min and 0.4 mL/min, for the methods A and B, respectively. The methods were validated according to the ICH guidelines for specificity, precision on the specified and LOQ limits, intermediate precision, accuracy, linearity (correlation co-efficient =0.999) and robustness. The robustness was confirned using four factors: the mobile phase pH, the flow rate of the mobile phase, column temperature and the second column of the same kind. The limits of detection and quantification were established as 0.0132 and 0.1505 μg/mL for methods A and B, respectively. Both validated methods complied with the acceptance criteria. The method B was 3.5 times faster than the method A, but the method A showed much better sensitivity. The resolution between compound B and bosentan was 3.39 and 1.75 for methods A and B, respectively. The lower sensitivity limits the use of Method B, especially in the analyses at low levels of active substances (e.g., bioanalysis, validation of the cleaning procedures) and makes method A more suitable for this purpose.