Chitosan (CS, molecular weight (MW) 20.2 kDA, stability of 210 °C and degree of deacetylation (DD) 73.31%) was obtained by deacetylation of chitin extracted from shrimp (Litopenaeus vannamei) shell wastes. The encapsulation of the studied bioactive natural product, toussaintine A (TA) isolated from the leaves of Toussaintia orientalis, on a chitosan-tripolyphosphate (CS/TPP) nanoformulation was attained through ionotropic gelation. Characterization of pure CS, CS/TPP and TA-CS/TPP nanocomposites was carried out by FTIR and SEM. The encapsulation efficiency and loading capacity of the TA were 69.33 and 0.46%, respectively. The in vitro release kinetics established an initial release of 27% of TA in the initial six hours followed by a slow and maintained release up to 72 h. The in vivo antimycobacterial acitivities of both TA and TA-CS/TPP nanocomposites against Mycobacterium indicus pranii (MIP) employing Galleria mellonella larvae as an infection model were evaluated. TA-CS/TPP nanocomposite formulations exhibited remarkable effectiveness against MIP than free TAInternational Foundation for Science (IFS)