Characterization of heparin-chitosan polyelectrolyte complexes for lung delivery

Heparin is an interesting drug for the treatment of inflammatory lung diseases but needs an efficient formulation strategy to reach the deep lung. To this purpose, heparin-chitosan polyelectrolyte nanocomplexes (PECs) were prepared and characterized. To gain a thorough understanding of their mechanisms of association and of the effect of ionic strength and concentration on heparin release, isothermal titration calorimetry (ITC), a powerful technique for evaluating the thermodynamics of molecular interactions, was applied [1]. High and low molecular weight heparins were investigated (HMWH,17,361 Da and LMWH, 6,839 Da) in association with chitosan (CS, 150,000 Da, DD 95%). Polymeric solutions were prepared in 0.1 M acetate buffer pH 4.1 and filtered (0.22 μm mPES). A 3.5 mg/mL of LMWH or HMWH solution was added dropwise to a 0.5 mg/mL CS solution at 25°C under magnetic stirring, in a 1:7 volume ratio. The resulting nanocomplexes were characterized in terms of size by dynamic light scattering (DLS), surface charge by Z-potential analysis, and polymers encapsulation efficiency by colorimetric assays (dimethylmethylene blue assay for heparin and Reactive Red 4 for chitosan). The formation of complexes was studied by ITC, by titrating LMWH or HMWH into CS, at 25 °C. Particle size and surface charge were 331 ± 168.8 nm and 42.8 ± 5.6 mV for LMWH/CS, or 342.1 ± 142.7 nm and 42.5 ± 4.9 mV for HMWH/CS. The quantitation revealed that all heparin was encapsulated in the complexes while CS involved was 89.7 ± 0.9 % for the LMWH system and 89.9 ± 0.6 % for HMWH system. ITC measurements revealed strongly enthalpy-driven interactions with nearly constant heat release over a wide range of molar ratios, which might suggest cooperative multivalent complex formation. Thermograms exhibited steep curves, with the final injections reversing from an exothermic to endothermic trend, consistent with dilution. Overall, despite the high energy involved, the signal is close to instrument’s detection limits, allowing only a qualitative interpretation of data as a strong electrostatic interaction between polyelectrolytes. These ITC measurements therefore represent a first step toward elucidating the interaction mechanism between these polyelectrolytes and assessing how ionic strength and concentration could influence heparin release. Literature: [1] I. D. Lima Cavalcanti, Int. J. Pharm. 2023, 641, 123063.