OBTENÇÃO E AVALIAÇÃO DA EFICÁCIA E TOXICIDADE IN VITRO DE NANOPARTÍCULAS DE BLENDAS DE PLA-PEG CONTENDO ANFOTERICINA B

Abstract

Amphotericin B (AmB) is the drug of choice for treating of systemic fungal infections, although its toxicity limitates its use. Many nanotechnology based formulations (liposomes and nanoemulsions) have been evaluated, whereas some of them are already been commercialized. The objective of these formulations is the reduction of AmB toxic effects, but due to the strong interaction between the drug and the formulations lipids, they do not present satisfactory efficacy. In this work, it was developed poly(D-L-lactide)- polyethyleneglycol (PLA-PEG) blend nanoparticles with variable molecular weight of PEG (2, 10 or 20 kDa) to encapsulate AmB and evaluate its in vitro efficacy over strains of Candida sp and in vitro toxicity over erythrocytes. Nanoparticles were prepared by the emulsification/solvent evaporation technique and the size, size distribution and polydispersity index were assessed by light scattering technique. Hemolysis test was performed to evaluate the protective caracter of the nanoparticles on erythrocytes from the lysis caused by AmB. In order to evaluate encapsulation efficiency and AmB in vitro release profile from nanoparticles, it was developed and validated a method by high performance liquid chromatography (HPLC) to quantify the drug. The validation process was performed considering the following parameters: selectivity, linearity, precision, accuracy, robustness, limit of detection (LOD) and limit of quantitation (LOQ). Reliability of the methodology was assured in every test. Encapsulation efficiency of AmB in nanoparticles of PLA-PEG blends was more than 68.9%. The mean particle size of the nanoparticles was 241 nm. The in vitro release experiment has demonstrated a burst effect in the first 24 h, followed by a sustained release of approximately 29.5% of AmB over 26 day of observation. Nanoparticles were able to protect erythrocytes from the lysis caused by AmB in the first 12 hours of observation, and did not modify therapeutic efficacy, considering sustained release profile. The formulations in study decreased AmB toxicity over erythrocytes, maintained in vitro efficacy and promoted sustained release of AmB.