Among the unique flora of the Amazonian Forest, numerous species have captured researcher’s attention for their therapeutic potential, demonstrating promising activities against infectious diseases. Notably, mycobacterial infections such as tuberculosis stand as a significant health concern in French Guiana. With a high incidence rate and the emergence of drug-resistant strains, there is an urgent need to explore therapeutic alternatives. This study delves into French Guiana’s biodiversity, to explore the chemical composition of 9 plant species, and uncover novel metabolites showcasing anti-mycobacterial properties. Drawing from the literature, preliminary results indicate interesting activities in the selected plant species. The use of a methodology combining recent computational metabolomics tools and a MS-based Molecular Network approach, facilitates structural dereplication and allows to work on large datasets, ensuring rapid and accurate characterization of metabolites of therapeutic interest. Using multiple analytical techniques, such as UHPLC-HRMS/MS and NMR, we conducted a comprehensive metabolomic analysis to profile the chemical composition of the plant extracts. Simultaneously, bioassays against multiple strains of mycobacteria were performed. The integration of bioassays results on a Molecular Network facilitated the identification of bioactive molecules, which were prioritized for further investigation and bioassay-guided purification. This multidisciplinary approach provides insights into the diverse chemical landscape of the studied plant extracts, and proposes a systematic approach to uncover novel anti-bacterial agents. This presentation will highlight the methodologies employed, initial findings, and the potential contribution of these plants in discovering therapeutic alternatives against mycobacterial infections.