DESENVOLVIMENTO DE NANOPARTÍCULAS POLIMÉRICAS CONTENDO RESVERATROL E AVALIAÇÃO DA AÇÃO NEUROPROTETORA SOBRE UM MODELO EXPERIMENTAL DE DOENÇA DE PARKINSON

Abstract

This study aimed at developing polymeric nanoparticles containing resveratrol (RVT). This drug has several pharmacological uses, including antioxidant and neuroprotective effects. However, the therapeutic effect is reduced due to its physico-chemical characteristics of low solubility in aqueous media and an extensively metabolization in a short time which results in a low bioavailability and ineffective amount in its active sites. Poly(lactic acid) (PLA), PLA-poly(ethylene glycol) (PLA-PEG) and PLA-Polisorbate 80 (PLA-P80) nanoparticles containing resveratrol (RVT) were obtained by emulsion-solvent evaporation method. A factorial design was carried out in order to investigate the influence of the formulation (ratio of aqueous to organic phase, surfactant concentration and solvent composition) on diameter and drug encapsulation efficiency (EE%). The factorial design was used in order to provide the best final formulation. An analytical method using high performance liquid hromatography (HPLC) / photodiode array (PDA) detection was developed and validated for RVT determination in nanoparticles. The mobile phase consisted of methanol:water (51:49) flowed at 0.9 mL/min with PDA detector set at 306 nm. This method had a suitable specificity, linearity, precision, accuracy, limits of detection and quantification and robustness according to current guidelines. PLA, PLA-PEG and PLA-P80 nanoparticles showed mean diameter and EE% of 227.57 nm and 82.47%, 185.47 nm and 75.67%, and 174.7 nm and 76.88%, respectively. The in vitro release study showed that RVT was released more slowly from PLA nanoparticles than PLA-PEG nanoparticles. The in vitro release data were in best-fitted by the Higuchi kinetics for both nanoformulations and the Kossmeyer-Peppas coefficient indicated that anomalous transport was the main release mechanism for RVT. The nanoparticles applicability was analyzed by studying the antioxidant potential of RVT-loaded formulations regarding their ability to scavenge the radical (2,2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt) (ABTS+). All formulations kept a significant antioxidant potential. At the end of 72 hours, PLA and PLA-P80 nanoparticles hed similar IC50 close to 16 μM of RVT. On the other hand, the PLA-PEG nanoparticles led to IC50 values similar to the free drug, 12.761 ± 0.792 μM and 11.148 ± 0.344 μM, respectively. Furthermore, the neuroprotective potential of RVT-loaded PLA-P80 nanoparticles and free drug (20 mg/kg/day for 15 days, intraperitoneally) was investigated using an experimental model of Parkinson's Disease induced by the 1-metil-4-fenil-1,2,3,6-tetrahidropiridine (MPTP) using C57BL/6 mices. A neuroprotection was demonstrated by behavoral data, lipid peroxidation (TBARS) levels, and tyrosine hydroxylase protein (TH) levels in striatal tissue of mices using formulation PLA-P80_RVT. However, free drug provide no neuroproteccion. In summary, RVT-loaded formulation prepared with PLA, PLA-PEG and PLA-P80 have high encapsulation efficiency, small diameter and are able to keep the antioxidant activity of the drug. In addition, PLA-P80 nanoparticles provide a remarkable neuroprotective effect related to RVT.