In Vivo Vaginal Fungal Load Reduction After Treatment with Itraconazole-Loaded Polycaprolactone-Nanoparticles.

Itraconazole (ITZ) has a broad spectrum of action and is commonly used for the treatment of fungal infections. Topic administration of ITZ is a promising strategy to improve vulvovaginal candidiasis treatment, which can be further optimized by its encapsulation in nanoparticles to increase drug delivery and reduce ITZ toxicity. In this work, we designed polycaprolactone nanoparticles containing ITZ and evaluated in vivo the efficacy of this yet unexplored approach. Nanocapsules (ITZ-NC) and nanospheres (ITZ-NS) were obtained by nanoprecipitation. ITZ-NC presented encapsulation efficiency of 99%, mean diameter of 190 nm, PDI 0.1 and zeta potential of -15 mV. ITZ-NS showed encapsulation efficiency of 97%, mean diameter of 120 nm, PDI 0.1 and zeta potential of -10 mV. Both particles were efficiently freeze-dried using 10% trehalose + 10% sucrose. Nanoparticles were then incorporated in a viscous formulation for vaginal application in female Balb/C mice infected with Candida albicans. Fungal load was significantly reduced in infected animals after treatment with ITZ-NC but not with ITZ-NS, compared to animals treated with ITZ solution. Histological analysis showed a clear difference between vaginal tissues of ITZ-NC and ITZ-NS and ITZ solution-treated animals, which correlated with IL-1ß and TNF-α quantification. Animals treated with ITZ-NC showed reduced cytokine levels and healthy tissue characteristics, while animals treated with ITZ-NS and ITZ solution showed increased IL-1ß and TNF-α levels and typical tissue inflammation. Our results demonstrate the potential of ITZ-NC to improve the treatment of vulvovaginal candidiasis after topical application in the vagina, opening new perspectives for the treatment of this disease.

Photoprotective effect and acute oral systemic toxicity evaluation of the novel heterocyclic compound LQFM048.

The new heterocyclic derivative LQFM048 (3) (2,4,6-tris ((E)-ethyl 2-cyano-3-(4-hydroxy-3-methoxyphenyl)acrylate)-1,3,5-triazine) was originally designed through the molecular hybridization strategy from Uvinul® T 150 (1) and (E)-ethyl 2-cyano-3-(4hydroxy-3-methoxyphenyl)acrylate (2) sunscreens, using green chemistry approach. This compound was obtained in global yields (80%) and showed an interesting redox potential. In addition, it is thermally stable up to temperatures around 250°C. It was observed that LQFM048 (3) showed a low degradation after 150min of sunlight exposure at 39°C, whereas the extreme radiation conditions induced a considerable photodegradation of the LQFM048 (3), especially when irradiated by VIS and VIS+UVA. During the determination of sun protection factor, LQFM048 (3) showed interesting results, specially as in association with other photoprotective compounds and commercial sunscreen. Additionally, the compound (3) did not promote cytotoxicity for 3T3 fibroblasts. Moreover, it was not able to trigger acute oral systemic toxicity in mice, being classified as a compound with low acute toxicity hazard (2.000mg/kg>LD50<5.000mg/kg). Therefore, this compound synthesized using green chemistry approach is promising showing potential to development of a new sunscreen product with advantage of presenting redox potential, indicating antioxidant properties.

Liposomal Entrapment of 4-Nerolidylcatechol: Impact on Phospholipid Dynamics, Drug Stability and Bioactivity.

Liposomes containing 4-nerolidylcatechol (4-NC), the major metabolite isolated from Pothomorphe umbellata, were obtained and characterized. Influence of liposomal encapsulation on chemical stability of 4-NC and on cytotoxicity profile of this drug was evaluated. Soybean phosphatidylcholine liposomes were prepared by lipid film hydration followed by extrusion. Entrapment efficiency for 4-NC was approximately 92%. Mean diameter of liposomes was 100 nm with a polydispersity index below 0.13. Liposomal 4-NC (L4-NC) and free drug (F4-NC) were submitted to forced degradation assays, monitored by HPLC. Photodegradation assay followed ICH Guidelines, using a photostability chamber equipped with both UV and white light sources. Liposomal encapsulation was able to markedly reduce 4-NC degradation rates under all the conditions tested. L4-NC showed a half-live approximately 15% higher than F4-NC under light exposure. After 72 hours, acid and base hydrolysis of F4-NC lead to 13 and 16% of degradation, respectively. However, no degradation was observed in L4-NC. EPR spectra of liposomal membrane showed that greatest changes in membrane properties were obtained when 5-doxyl stearic acid was used as the spin label, indicating a marked decrease in the fluidity of the bilayer. Following incubation with K562 cells, 4-NC showed a concentration-dependent cytotoxicity profile, while L4-NC exhibited a time and concentration-dependent profile, consistent with a controlled drug release system. F4-NC induced extensive hemolysis under isotonic conditions; conversely liposomal encapsulation protected erythrocytes from 4-NC induced lysis. Liposomal 4-NC resulted in a hemocompatibility and stable formulation, representing a viable drug delivery system to further investigate in vivo performances of 4-NC in pre clinical studies.

Atividade citotóxica e antiangiogênica de Punica granatum L., Punicaceae / Cytotoxic and antiangiogenic activities of Punica granatum L., Punicaceae

Punica granatum L., Punicaceae, amplamente usada no Brasil, foi avaliada quanto ao seu potencial antitumoral in vitroe in vivo. Investigou-se in vitro a citotoxicidade do extrato etanólico do fruto e folha da P. granatumutilizando células K-562 e células do Tumor Ascítico de Ehrlich (TAE), pelos métodos de redução do MTT e exclusão do azul de tripano. Nos estudos in vivoavaliou-se o aumento da sobrevida de animais portadores do TAE e tratados, por via oral, com diferentes doses dos extratos etanólicos da P. granatum(12,5; 25; 50 e 100 mg/kg) por dez dias consecutivos. Além disso, nestes animais analisou-se o potencial de inibição tumoral e a atividade antiangiogênica da P. granatum. Os resultados dos estudos in vitrodemonstraram uma redução na viabilidade das células K-562 e do TAE, concentração-dependente, nos métodos investigados. Os resultados in vivo demonstraram aumento da sobrevida dos animais portadores do TAE tratados, de forma dose-dependente. Em paralelo, observou-se diminuição do número de células tumorais na cavidade peritoneal dos animais portadores e tratados. Além disto, os tratamentos empregados reduziram o padrão de vascularização da parede abdominal. Dessa forma, os dados apresentados revelaram que o extrato de P. granatumpossui atividade antitumoral in vitroe in vivo em paralelo a redução da angiogênese peritoneal.

Punica granatum L., a plant widely used in Brazil, was tested for its antitumor and antiangiogenic activities in vitroand in vivo. In this work, the in vitrocytotoxicity was evaluated using the K-562 cell line and Ehrlich ascites tumour cells, by MTT tetrazolium reduction test and the trypan blue exclusion test. In vivostudies investigated the increase in the survival time of Ehrlich tumour-bearing mice after treatment with different doses of Punica granatumL. ethanol extract (12.5; 25; 50 e and 100 mg/kg), by gavages, for ten consecutive days. In addition, we also investigated the tumour inhibition potential and antiangiogenic activity of this plant. In vitroresults demonstrated a decrease of K-562 and Ehrlich ascites tumour cells viability, with both methods used, in a dependent-manner concentration. In vivoresults showed a significant antitumor activity against Ehrlich ascites tumour growth, increasing survival time. In parallel, we detected a significant inhibition of the tumour growth, along with a decrease in the vascular pattern of the peritoneal wall. Thus, the data presented herein clearly showed that Punica granatum L. has antitumor and antiangiogenic activities.

Avaliação da toxicidade aguda e subaguda, em ratos, do extrato etanólico das folhas e do látex de Synadenium umbellatum Pax / Acute and subacute toxicity studies of the latex and of the ethanolic extract of the leaves of Synadenium umbellatum Pax in rats

O uso de plantas medicinais tem sido muito significativo nos últimos anos, sendo incentivado pela Organização Mundial de Saúde (OMS). Synadenium umbellatum Pax, Euphorbiacea (vulgo cola-nota, cancerola, milagrosa) tem o látex usado empiricamente como antitumoral e antiinflamatório. Por existir espécies tóxicas nesta família e visando à segurança no uso de extratos vegetais, tal estudo avaliou a toxicidade pré-clínica do látex e do extrato etanólico das folhas (EEF) de S. umbellatum, por via oral, em ratas Wistar. O estudo seguiu diretrizes do Guideline 423 (toxicidade aguda) e Guideline 407 (toxicidade subaguda) da OECD (Organisation for Economic Cooperation and Development). Na toxicidade aguda do látex e do EEF, não se observou letalidade nem alterações fisiológicas e comportamentais das ratas na dose de 2000 mg/kg, sendo praticamente atóxico. Porém, na análise histopatológica, o látex ocasionou congestão e infiltrado leucocitário nos rins, fígado e pulmões, efeitos não observados com o EEF. Na toxicidade subaguda, doses de 50, 100 e 200 mg/kg de EEF não produziram alterações dose-dependentes significativas nos parâmetros laboratoriais e fisiológicos, nem alterações macroscópicas e histopatológicas nos órgãos das ratas. Contudo, o uso crônico da planta S. umbellatum merece mais estudos.

The use of medicinal plants has been being very significant in the last years, being the use encouraged by WHO. Synadenium umbellatum Pax, Euphorbiacea (popularly known as cola-note, cancerola, miraculous) has the latex used empirically as anti-cancerous and anti-inflammatory. For there being toxic species in this family and aiming at the safety in the use of vegetable extracts, such study evaluated the pre-clinical toxicity of the latex and of the ethanolic extract of the leaves (EEL) of S. umbellatum, administrated by oral route, in Wistar female rats. The study followed OECD's Guidelines for test of acute toxicity (Guideline 423) and for subacute toxicity (Guideline 407). In the acute toxicity of latex and EEL, behavioral and physiological alterations were not observed neither animal's death in the dose level of 2,000 mg/kg. However, the latex caused congestion and leukocytes infiltration of the kidneys, liver and lungs, effects not observed with EEL. In the subacute toxicity, dose levels of 50, 100 and 200 mg/kg of EEL did not produced significant dose-dependent alterations in the lab results and no physiologic, macroscopic and hystopathological alterations. EEL of S. umbellatum is practically poisonless in acute exposure; already the latex can cause hystological damages. The chronic use of S. umbellatum needs more specific studies.