Avaliação da fração diclorometano de Libidibia ferrea (Fabales: Fabaceae) em estudos Pré-clínicos da Leishmaniose cutânea

Abstract

Among neglected tropical diseases, cutaneous Leishmaniasis is one that deserves more attention, as it occurs in more than 88 countries, with about 20 thousand new cases per year in the world. The application and therapeutic indication described by the Ministry of Health can often not reflect a regional reality, difficulties are observed from the invasive route of administration and the numerous adverse reactions, thus contributing to low adherence to treatment. Therefore, the search for therapeutic alternatives is necessary, one of which is the natural source of medicinal plants, which have been used since the dawn of humanity in order to combat different diseases. Among the several plant species with pharmacological potential, one of them is Libidibia ferrea, popularly known as jucá, with proven biological activities, including antileishmania. Its active fractions can be incorporated into the microemulsified system to increase efficacy in the treatment of cutaneous leishmaniasis (LC). This is considered an excellent vehicle for transporting different drug classes, due to its stability, control in the distribution of drugs and being very permeable, even allowing its administration by topical route. Thus, the objective of this work is to develop microemulsions containing the Dichloromethane (DCM) fraction of L. ferrea and evaluate its effect on hamsters infected with L. amazonensis, in addition to studying its chemical composition and testing the intileishmania activity in vitro and cytotoxicity of isolated substances, contributing to a more efficient treatment of LC, with reduced unwanted effects and painless administration. The DCM fraction was obtained from the fractionation of the methanolic extract of L. ferrea. Approximately 300 mg (Final concentration of the DCM fraction in the microemulsion) was incorporated into the microemulsion, which was subjected to physico-chemical characterization by the average droplet diameter, polydispersity index, zeta potential, refractive index and pH, before application to animals. After 30 days of treatment, the animals were euthanized and fragments of the lesions were collected to determine the parasitic burden and histopathological aspect. Another part of the obtained fraction was subjected to High Efficiency Liquid Chromatography for the isolation and the identification of the substances was done by Nuclear Magnetic Resonance analyzes. The samples of fractions and isolated substances were evaluated for antileishmania activity against the promastigote and amastigote forms of Leishmania amazonensis and the cytotoxic profile in macrophages of lineage J774 and peritoneal. The results obtained indicated that the microemulsion remained stable and promising in the treatment of LC in an animal model, with no statistical difference between the groups treated with the microemulsion and the standard medication (Glucantime®). From the Dichloromethane fraction, three substances were isolated, with methyl gallate and a derivative of phenylpropanoic acid being identified. The substances showed antileishmania activity against promastigote forms (between 71.5 to 17.4 µg.mL-1 in the 72h period) and without a cytotoxic profile. Therefore, these results allowed the identification of the major substances, which even show themselves to be active and when using the DCM fraction of L. ferrea incorporated in the microemulsion, due to possible synergistic action, showed effectiveness in the topical treatment, indicating a new proposal for the therapeutic protocol, complementary to standard LC pharmacotherapy.