Isolamento, quantificação e avaliação das atividades leishmanicida e tripanocida de furanoditerpenos do oleorresina de Pterodon spp. Vogel (Fabaceae)

Abstract

Pterodon (Fabaceae) plant species known as white “sucupira” are widespread in Brazilian savannah. Their therapeutic use is mainly due to the analgesic and antiinflamatory properties, essentially attributed to the furanic diterpenes with vouacapanic skeleton, found in their fruits. Researches carried out with diterpenes from other plant species have shown promising activity against the causative agents of neglected diseases. It is necessary to implement a HPLC method for the achievement and quantification of these diterpenes, because the methods available to date are multi-step and laborious. This research aimed to isolate (chapter I) and quantify furanic diterpenes from sucupira’s fruit by HPLC (chapter II) and assess their in vitro leishmanicidal and trypanocidal activity (chapter III). First, the oil was fractionated through liquid-liquid partition, and so it was undergone to preparative reverse phase HPLC separation. Three compounds were isolated: two new furanic diterpenes: 6α-acetoxy- 7β,14,18-trihydroxyvouacapan and 6α-acetoxy-7β,14-dihydroxyvouacapan and methyl-6α- acetoxy-7β-hydroxyvouacapan-17β-oate (MAHV), as established by NMR and mass spectroscopy. MAHV, the major compound in the oil, and β-caryophyllene (Sigma®) were chosen as chemical markers for the quality control of the oil, by reverse phase HPLC. A method was developed and validated. Mobile phase consisted of a gradient of acetonitrile, acidified with phosphoric acid 0.2 % (v/v), methanol, and phosphoric acid 0.5 % (v/v) in water, at 1 mL/min. A C18 column at 30ºC was used. Chromatograms were recorded at 210 nm. MAHV was present in the sample at the concentration of 15.98% (w/w), and β- caryophyllene at 8.01% (w/w). The oil, the isolated MAHV and the compound methyl-6α-7β- dihydroxyvouacapan-17β-oate (MHV), a synthesized furanic diterpene also present at sucupira’s fruit, was assessed against the promastigote form of the Leishmania braziliensis e Leishmania amazonensis species, and against the epimastigote form of the parasite Trypanosoma cruzi, through the in vitro MTT colorimetric assay. The MAHV revealed to be more active than both the MHV and the oil. It showed an EC50 < 30 µg/mL at all of the assays. The results indicated the potential of the diterpenes of sucupira’s fruits in the control of neglected diseases, as leishmaniosis and trypanosomiasis. Further studies should be carried out to investigate the cytotoxicity of these compounds in mammalian cells.