In this study, the synthesis, spectral characterization and in vitro antimicrobial activity of two new metallodendrimers based on the PAMAM dendrimer modified with four 1,8-naphthalimide units are described. The chemical structure of metallodendrimers has been analyzed by FT-IR, UV-Vis, fluorescence, NMR spectroscopy, MS and SEM analysis. The antimicrobial activity of the new compounds has been tested in vitro against different model pathogenic microorganisms. The results showed good inhibitory activity of the compounds better expressed against the tested Gram-positive bacteria and yeasts. Cotton fabrics were treated with each of the dendrimers, and their morphology, antimicrobial activity and biofilm inhibiting potential were investigated. Both metallodendrimers showed stronger antimicrobial and biofilm inhibiting abilities compared to the initial dendrimer ligand.