Polysorbate 80 p80

The synthesis of the hydrochloride salt form of the cationic lipid 2,3-di (tetradecyloxy) propan-1-amine (D) was performed as described in the literature, with slight modifications of the laboratory protocol [17] . Niosomes were composed by modified reverse-phase evaporation approach [18] . Concisely, 5 mg (0.1% w/v) of the lipid was dispersed in 1 ml of dichloromethane (organic phase). Subsequently, 5 ml milliQ water containing 12.5 mg (0.25% w/v) poloxamer 188 (P) (Sigma-Aldrich, Madrid, Spain), 12.5 mg (0.25% w/v) polysorbate 80 (P80) (Sigma-Aldrich, Madrid, Spain) and 2.5 mg (0.05% w/v) chloroquine diphosphate (CQ) (Sigma-Aldrich, Madrid, Spain) were added to the organic phase. The emulsions were obtained by sonication of the mixture for 50 s at 45W (Branson Sonifier 250®, Danbury, USA). Dichloromethane was eliminated from emulsions by dissipation under magnetic stirring for 2 h, rendering the cationic niosomes in the aqueous medium. The molar ratios of both DPP80 and DPP80-CQ formulations were, 1.9/0.3/1.9 and 1.9/0.3/1.9/1, respectively.

T was obtained from Euma, Buenos Aires, Argentina (density of T was considered 0,915 g/ml). Soybean phosphatidylcholine (SPC, purity >90%) was a gift from Lipoid, Ludwigshafen, Germany. Polysorbate 80 (P80), molybdenum oxide, ascorbic acid, Sephadex G-25, 6-dodecanoyl-N, N-dimethyl-2-naphthylamine (laurdan), vancomycin, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and crystal violet were purchased from Sigma-Aldrich (St Louis, MO, USA). Roswell Park Memorial Institute (RPMI) 1640 medium was from Gibco (Buenos Aires, Argentina).
Fetal bovine serum (FBS), antibiotic/antimycotic solution (penicillin 10,000 IU/ml, streptomycin sulphate 10 mg/mL, amphotericin B 25 µg/ml), glutamine, and trypsin/ethylenediaminetetraacetic acid were from PAA Laboratories GmbH (Pasching, Austria). Tryptic soy broth (TSB) and bacteriological agar were acquired from Britania, Buenos Aires, Argentina. The other reagents were of analytic grade from Anedra (Buenos Aires, Argentina), Research AG (Tigre, Buenos Aires, Argentina).