Chitosan-trimethoprim injectable hydrogel for anti-biofilm and chronic wound healing.

Schiff-base hydrogels are widely used as drug depots. However, their application in antibacterial wound healing is rarely explored. To fill this gap, we developed an injectable chitosan (CS) hydrogel physically loaded with trimethoprim (TMP) within a Schiff-base crosslinked network. The key novelty lies in the covalent crosslinking between aldehyde-terminated PEG and chitosan, which enables pH-responsive release of physically loaded TMP while maintaining injectability and self-healing properties. The hydrogel was characterized by SEM, and its antibacterial activity and wound healing capacity were evaluated in vitro and in vivo. Results showed ≥99.95% inhibition against Staphylococcus aureus (S. aureus) and 52.23% against methicillin-resistant S. aureus (MRSA) within 24 h. The hydrogel disrupted preformed biofilms, reducing biofilm biomass by approximately 46-54% across S. aureus, Escherichia coli (E. coli), and mixed-culture models. Drug release exhibited pH-dependent behavior, with cumulative release reaching 95.9% at pH 5.3, 60.21% at pH 6.4, and 41.12% at pH 7.4 at 48 h. DHFR activity in bacterial lysate dropped from 668.0 U/L to 84.8 U/L. Nucleic acid and protein leakage were 4509 μg/mL and 688 μg/mL, respectively. In vivo, the treatment group maintained body weight (23.7 g) versus controls, with only 10% residual wound area by day 7. This hydrogel integrates pH-responsive release, potent antibiofilm activity, and pro-repair function, offering a new strategy for chronic infected wounds.