Eltrombopag olamine isprescribedfor chronic immune (idiopathic)thrombocytopenicpurpura (ITP). This work aims to investigate the formation of potential degradants ofthe drug and determine their toxicity in silico. A stability-indicating high performance liquid chromatography (HPLC) method was developed to separate six oxidative degradation impurities and three thermal degradation impurities employing the quality by design (QBD) approach. The degradation impurities were resolved with minimum resolution of 1.5 using a phenyl column with 0.1 % trifluoroacetic acid (TFA) and acetonitrile as the mobile phase and quantified at 245 nm. The structure and degradation pathway for the degradants was proposed by employing liquid chromatography with tandem mass spectrometry (LC–MS/MS), among the identified degradation pathways demethylation and decarboxylation are common reactions observed during oxidation resulted in majority of degradation products. All the degradation products are characterized with help of the daughter ions and product ion obtained upon LC–MS/MS analysis. The HPLC method parameters such as column temperature, flow rate, TFA concentration and organic concentration are identified as critical method attributes (CMA), a design of experiments (DOE) mediated design space was established through use of design experts. The resolution between sets of adjacent peaks was identified as a critical quality attribute; among the investigated CMAs, column temperature and flow rate significantly affected the resolution. Furthermore, the toxicology of the degradation products was predicted with the help of in silico TOPKAT analysis, the carcinogenicity of the impurities was discussed.