Meglumine antimonate is the active of Glucantime® used for the treatment of leishmaniasis, a tropical disease caused by parasitic protozoa, and it is estimated that 12 million people worldwide are affected. This drug mainly contains Sb(V) under the form of an organic complex with N-methylglucamine (NMG). During the synthesis of this molecule, traces of Sb(III) may be present, also probably complexed. Due to the fact that Sb(III) is considered more toxic than Sb(V), it is important to evaluate the Sb(III) concentration in the drug samples. In the literature, very different concentrations for residual concentrations of Sb(III) in the drug ampoules are found. Therefore, to have a true insight of antimony speciation, two independent analytical methods were developed in this work. We used an anion exchange method coupled with inductively coupled plasma mass spectrometry (ICP-MS) which was cross-referenced with an electrochemistry method (differential pulse polarography (DPP)) that could be used for routine analysis on the production site. To obtain Sb species in detectable forms, the complexes between Sb species and NMG need to be broken. This was obtained by diluting samples in hydrochloric acid in deaerated conditions to avoid Sb redox reactions. For the two analytical methods, the HCl concentration was optimized to obtain simultaneously a complete destruction of the complexes as well as limited redox reactions for Sb(V) and Sb(III) released species. For high-performance liquid chromatography (HPLC)-ICP-MS, a dilution with 5 M HCl gives the better results. The side reaction is an oxidation of Sb(III) which can be limited by the removal of oxygen. When DPP is used, the major problem is the reduction of Sb(V) which is present in high amount in the samples. Working with 0.6 M HCl allows this problem to be minimized. When applied to different lots of Glucantime®, Sb(III) concentration values are in good agreement for the two analytical methods, with, for HPLC-ICP-MS, the advantage of the simultaneous detection of both Sb redox species.