Small-angle neutron scattering characterization of liposomes for anti-tuberculosis inhaled therapy.

The present investigation studied the effects of two first-line anti-tuberculosis (TB) drugs, rifampicin (RIF) and isoniazid (INH), on the structure of multilamellar liposomes. Liposomes have been shown to be a promising system for inhaled therapy. (1) The study of liposome-drug interactions is essential, and small-angle neutron scattering (SANS) technique provides valuable and unique data about steric bilayer thickness, particle dispersion, number of lamellae and drug localization under physiological conditions. (2) Unloaded, single drug-loaded and co-loaded liposomes were prepared using different amounts of drugs by reverse phase evaporation method. Liposomal suspensions were prepared using D2O, in order to emphasize the contrast between the aqueous and the lipid/drug phases. The samples were characterized by dynamic light scattering, atomic force microscopy and finally by SANS technique (Rutherford Appleton Laboratory, U.K.). Neutron scattering curves were analyzed using a multi-shell spherical model of the fitting routine SASView 2.2.0. Liposomes have been shown to be physico-chemically stable during the experiments, efficiently drug-loaded, and able to control drug release. Dimensional analysis demonstrated that particle sizes are in the range of SANS dimensional detection. SANS curves exhibited Bragg peaks for all samples, confirming the multilamellar liposome structure. By fitting the data, significant differences among the samples have been highlighted. RIF-liposomes were less ordered than unloaded liposomes: a reduction of the lamellae number was observed and the periodicity of the lipid bilayers slightly increased with the increment of the drug loading, may be due to RIF interaction with phospholipid tails, which can destabilize liposome lamellarity, since RIF is a hydrophobic drug. In INH-liposomes, the drug payloads did not change vesicle structure, because INH is a hydrophilic drug. However, INH induced a change in the inter-bilayer periodical spacing, which could be compatible with the formation of drug-liposome complexes at the water-lipid interface. Finally, the RIF-INH co-loaded liposomes exhibited the same characteristics of unloaded liposomes, suggesting that INH and RIF together have a stabilizing effect on the structure. In fact, no destabilization and no changing in inter-bilayer periodical spacing were observed. In conclusion, SANS analysis provides fundamental information about drug-liposome interactions to comprehend the relation between system structure behaviour and its biological activity. Moreover, data suggest that the co-encapsulation of the two anti-TB drugs may have a synergic effect on liposome stability. 1. A. Elhissi, Curr. Pharm. Des., 2017, 23, 362-372. 2. PC. Lin, S. Lin, PC. Wang, R. Sridhar, Biotechnol. Adv., 2014, 32, 711– 726.