Mayaro virus is a neglected virus that causes a mild, dengue-like febrile syndrome characterized by fever, headache, rash, retro-orbital pain, vomiting, diarrhea, articular edemas, myalgia, and severe arthralgia, symptoms which may persist for months and become very debilitating. Though the virus is limited to forest areas and is most frequently transmitted by Haemagogus mosquitoes, Aedes mosquitoes can also transmit this virus and, therefore, it has the potential to spread to urban areas. This study focuses on the metabolic foot-printing of Vero cells infected with the Mayaro virus. Nuclear magnetic resonance combined with multivariate analytical methods and pattern recognition tools found that metabolic changes can be attributed to the effects of Mayaro virus infection on cell culture. The results suggest that several metabolite levels vary in infection conditions at different time points. There were important differences between the metabolic profile of non-infected and Mayaro-infected cells. These organic compounds are metabolites involved in the glycolysis pathway, the tricarboxylic acid cycle, the pentose phosphate pathway, and the oxidation pathway of fatty acids (via β-oxidation). This exometabolomic study has generated a biochemical profile reflecting the progressive cytopathological metabolic alterations induced by Mayaro virus replication in the cells and can contribute to the knowledge of the molecular mechanisms involved in viral pathogenesis.