Avaliação anti-leishmania in vitro e in vivo de Libidibia ferrea e estudo de formulações tópicas para o tratamento da forma cutânea da Leishmaniose.

Abstract

The American Cutaneous Leishmaniasis (ACL) is a protozoonosis caused by parasites of the genus Leishmania, which is transmitted by the bite of the sand fly insect. The LTA, patients can present cutaneous form, with localized skin lesions that may progress to chronic lesions. The treatment of choice is pentavalent antimony and drugs of second choice are pentamidine and amphotericin B, but all have side effects and require parenteral administration. Natural products are presented as an alternative for the treatment of many diseases, and there are studies that demonstrate potential against Leishmania. In the Amazon region, Libidibia ferrea (Mart. Ex Tul) LP Queiroz stands out for its economic and pharmacological importance, due to anti-inflammatory and antimicrobial reported. This study aimed to evaluate the activity of extracts and fractions of L. ferrea against promastigotes and amastigotes of L. (L.) amazonensis and L. (V.) guyanensis also conducting its chemical characterization, and study the effect in topical formulations containing pentamidine, extracts and most promising fractions. And hexane extracts were prepared methanolic the epicarp, leaves and branches of L. ferrea, who underwent in vitro bioassays against promastigotes and amastigotes of L. (L.) amazonensis and L. (V) guyanensis, the counting method direct. The most active extracts were evaluated in vitro for cytotoxic activity in J774 macrophage lineage. The extracts and active fractions were analyzed by TLC assay, the quantized flavonoid and phenolic content, and analyzed by NMR and HPLC. The in vivo assays employed as an animal model hamsters infected with L. (L.) amazonensis. They were prepared and evaluated by topical application on lesions initially hydrating and anhydrous emulsions containing pentamidine isethionate (10%). The results of the bioassays showed that the methanolic extract from epicarp (EpMeOH) showed a higher activity against species of Leishmania, with EC50 15.01 µg/ml against L (L.) amazonensis. This extract had the highest concentration of phenolic (5.14%). A 2-DCM fraction also demonstrated activity with EC50 12.77 µg/mL against L. (L.) amazonensis. No cytotoxic activity was observed in macrophages for both samples. The moisturizing emulsions containing pentamidine were most effective, although viable parasites were detected and all groups after eight days of treatment. These moisturizing emulsions were used as a base for creams with addition of 2.5% of EpMeOH extract and evaluated in two areas with skin lesions - snout and paw. The groups treated with cream EpMeOH extract (2.5%) and placebo showed differences in lesion volume compared to the control group after 20 days of treatment, but showed the presence of parasites, with no statistical difference between the control group. These results suggest that the treatment time and extract concentration to be increased beyond the proportion of use of EpMeOH extract in a more hydrophilic formulation. The hydrogel EpMeOH was then evaluated for lesions on the snout of hamsters as well as cream Fr2. After 40 days of treatment, the groups treated with these topical formulations showed significant differences between the control group and between their respective placebos, as volume, clinical aspect of the lesion, and as the parasitic quantification. Minor inflammatory infiltrate was observed in the hydrogel EpMeOH. This study identified extracts promising in the treatment of experimental cutaneous leishmaniasis and supports pointing new opportunities and innovation serving as a basis for herbal formulations to treat this neglected disease.