Chitosan Stabilizes Curcumin against Hydrolysis and Potentiates Antimicrobial Activity against Pseudomonas aeruginosa in a pH-Dependent Manner.

Curcumin exhibits antimicrobial activity against Gram-negative bacteria but is limited by rapid hydrolytic degradation at near-neutral pH. We hypothesized that chitosan, a cationic polymer that independently destabilizes the Gram-negative outer membrane, could simultaneously stabilize curcumin and potentiate its antibacterial activity. Curcumin degradation kinetics in cation-adjusted Mueller-Hinton broth (35-85 °C) revealed a concentration-dependent transition in chitosan-mediated stabilization: low chitosan ratios (1:0.5-1:1) reduced degradation rates without altering activation energy (∼33-35 kJ/mol), consistent with kinetic shielding, whereas higher ratios (1:2-1:3) increased activation energy to 39.0-45.8 kJ/mol, indicating formation of a protective curcumin-chitosan complex. Complementary microplate kinetic spectroscopy at 35 °C across pH 6.6, 7.2, and 7.4 confirmed that chitosan-mediated stabilization is itself pH-dependent, extending curcumin's half-life by up to 10.8-fold at pH 7.2 (1:2 ratio) and 5.9-fold at pH 6.6 (1:3 ratio). Checkerboard synergy assays against six Pseudomonas aeruginosa isolates revealed pH-dependent potentiation consistent with synergy (Bliss Independence model), with mean Bliss Δ shifting from +0.112 at pH 6.6 to -0.019 at pH 7.4, tracking the ionization of chitosan's amine groups (pK a 6.2-6.5). At a 24-h end point, the 1:2 combination produced greater nucleic acid and protein leakage than either agent alone; however, a complementary short-duration kinetic assay (0-120 min) across three pH values indicated that this enhancement reflects chitosan-mediated stabilization extending curcumin's effective exposure duration rather than direct mechanistic synergy at the membrane. Together, these results identify a composite mechanism in which chitosan protects curcumin from hydrolysis while both agents contribute pH-dependent membrane disruption, with maximal antibacterial potentiation in the pH range characteristic of the cystic fibrosis airway.