Uniform stable chitosan nanoparticles (CS NPs) loaded with sodium ceftriaxone (CTX) were prepared using the ionic crosslinking method by applying a monovalent salt. To achieve NPs with a minimum polydispersity index (PDI), minimum size, and maximum entrapment efficiency of the drug (EE %), the effect of NaCl salt concentration and pH of CS solution was investigated using a 32-full factorial design. Then, the stability of CTX-loaded CS NPs over six-month storage was investigated. Surface charge density was calculated using Gouy-Chapman theory. Then, minimum inhibitory concentration and minimum bactericidal concentration of free CTX and salt-assisted CTX-loaded CS NPs were determined against Staphylococcus aureus and Escherichia coli as aerobic bacteria, and Bacteroides fragilis as anaerobic bacteria. Results suggested that optimized NP was 265 nm in size with a PDI value of 0.075, and high drug EE% of 80% with high stability over 6 months. Results of time-kill assays revealed that the salt-assisted CTX-loaded CS NPs could thoroughly kill the E. coli and B.fragilis bacterium after 8 h, and S.aureus bacterium after 10 h, while free CTX could kill all bacteria after 24 h. Finally, the salt-assisted CTX-loaded CS NPs provide uniform and stable NPs with more controlled release and higher antibacterial effect compared to free CTX.
