Chitosan nanoparticles (CNP) were synthesized via ionic gelation and used for the preparation of starch-based nanocomposite films containing different concentration of CNP (0, 5, 10, 15, 20% w/w). Antimicrobial properties of starch/CNP films was evaluated via in vitro (disc diffusion analysis) and in vivo (microbial count in wrapped cherry tomatoes) study. It was found that inhibitory zone of the 15 and 20% of starch/CNP films were clearly observed for all the tested bacteria including Bacillus cereus, Staphylococcus aureus, Escherichia coli and Salmonella typhimurium. In vivo study revealed that the starch/CNP film (15% w/w) was more efficient to inhibit the microbial growth in cherry tomatoes (7 × 102 CFU/g) compared to neat starch film (2.15 × 103 CFU/g) thus confirmed the potential application of the films as antimicrobial food packaging.