Gelatin nanoparticles, synthesized via desolvation, serve as effective carriers for anti-tuberculosis drugs in both single and combination therapies. Characterization through cutting-edge technology (dynamic light scattering, high-resolution transmission electron microscopy Field emission scanning electron microscopy) confirmed their structural features, with entrapment efficiencies ranging from 60 to 92%. Notably, isoniazid, streptomycin, and pyrazinamide exhibit sustained release of 35-42%, while rifampicin shows 6-13% release over 115 h at pH 7.4 in PBS buffer for both therapies. In rifampicin/isoniazid- and rifampicin/pyrazinamide-loaded gelatin microparticles, fractional inhibitory concentration indices were found to be 0.031, 0.022, 0.267, and 0.0003 g/mL for rifampicin, isoniazid, rifampicin, and pyrazinamide, respectively. Minimum inhibitor concentrations for isoniazid and streptomycin in single-drug therapy were 0.23 and 0.31 g/mL. Combination therapy exhibited superior anti-tuberculosis efficacy (indices < 0.5), suggesting a synergistic effect. These findings indicate promising prospects for sustained drug release and improved tuberculosis treatment through combination therapy using gelatin nanoparticles.