Abstract: In order to solve the problems of the antimicrobial peptide Temporin-GHaR that stability is poor and the hemolytic toxicity is high, in the report, the preparation design technology was used to construct a thermosensitive quaternary composite hydrogel of carboxymethyl chitosan/hydroxypropyl methylcellulose/poloxamer loaded with Temporin-GHaR (CHPP/GHaR). The response surface method was used to optimize the formulation, the obtained hydrogel with a porous fiber network structure exhibited shear-thinning, self-healing, and injectability, and the swelling rate was 161.5%. The drug release tests showed a sustained release of 55% within 40 hours with low burst release risk. Biocompatibility evaluation test confirmed that the hydrogel loaded with 200 μmol·L^-1 GHaR had only 2.2% hemolysis rate and no significant cytotoxicity to keratinocytes and umbilical vein endothelial cells. In vitro antimicrobial experiments demonstrated that CHPP/GHaR achieved above 99% inhibition rates against methicillin-resistant Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, the inhibition rates of which were more than 10 times higher than that of the control. Our findings provide a new candidate material for the treatment of the resistant bacterial. infections.