Estudo químico e biológico de Duroia macrophylla Huber (Rubiaceae)

Abstract

The species of Rubiaceae revealed great diversity of secondary metabolites, which are responsible for a range of biological activities. Among these species is Duroia macrophylla Huber, endemic to the Amazon Rainforest, popularly known as “cabeça-de-urubú, apuruí ou puruí-grande-da-mata”. The absence of studies of plants of the genus Duroia and the absence of chemical studies and biological activity to D. macrophylla, the aim of this work was to isolate the chemical constituents and evaluate the extracts and compounds isolated on activities: antioxidant, toxicity against Artemia salina, antibacterial, antimycobacterial and antitumor. The plant material was collected two times, dried, grounded and extracted with dichloromethane or hexane and methanol. The extracts were subjected to phytochemical screening by comparative thin layer chromatography and to determine specific telltale signs of chemical classes. The extracts were tested as antioxidant, cytotoxic, antibacterial, antimycobacterial and antitumor, to increase the chances of obtaining active molecules and/or prototypes of drugs. The isolated compounds were identified by spectroscopic methods (1H, 13C and two-dimensional NMR) and mass spectrometry and essayed as antimicrobial and antitumor. All extracts showed signs of terpenes and only the dichloromethane extracts of leaves and branches of the 1st collection did not reveal with DPPH. The methanol extracts of both collections showed aromatic compounds. The presence of alkaloids was detected only in extracts from the branches of the 2nd collection. There were isolated and identified four substances of the extracts of the 1st collection: two triterpenes from dichloromethane extract of the leaves (oleanolic acid and ursolic acid), one chalcone from methanol extract of the leaves (4,4'- dihydroxy-3'-chalcone) and a phenolic acid from methanol extract of the branches (mmethoxy- p-hydroxy-benzoic acid). Were identified of the extracts of the 2nd collection, eight monoterpene indole alkaloids: 10-methoxy-ajmalicine, 11-methoxy-ajmalicine, 11-methoxy- 3-isoajmalicine, 9-methoxy-3-isoajmalicine, 9-methoxy-19-epi-3-isoajmalicine, 10-methoxy- 19-epi-3-isoajmalicine, 10-methoxy-3-isorauniticine and 10-methoxy-rauniticine. All compounds isolated in this study were described for the first time in the genus Duroia. The methanol extracts from leaves and branches in both collections showed a good antioxidant activity. In cytotoxic assay against A. salina only the methanol extract of the leaves (2nd collection) presented toxicity at lethal concentration (LC50) of 120 mg/mL. The extracts showed bacteriostatic activity against the bacteria Klebsiella pneumoniae, Flavobacterium corumnare, Salmonella enteridis and Pseudomonas aeruginosa. Of the substances tested only oleanolic acid showed antibacterial activity against Nocardia brasiliensis and Serratia marcescens, with a MIC of 500 mg/mL. Extracts subjected to antimycobacterial bioassay, the dichloromethane extract of the leaves (1st collection) showed better results against all strains of Mycobacterium tuberculosis with an MIC of 6.25 mg/mL for INHr strain, 25 mg/mL for the strain RMPr and ≤ 6.25 mg/ml for H37Rv strain. Only the alkaloids 10-methoxy-3- isorauniticine, the mixture of 9-methoxy-3-isoajmalicine with 9-methoxy-19-epi-3- isoajmalicine, 10-methoxy-3-isorauniticine and 10-methoxy-rauniticine tested against M. tuberculosis (strain INHr) showed better results rather those obtained from crude extracts. The extracts and alkaloids showed low cytotoxic potential on neoplastic cell lines: HCT116 (human colorectal carcinoma), MCF-7 (breast carcinoma), SK -Mel-19 (human melanoma) and on the non-neoplastic line: MRC-5 (human lung fibroblast). Keywords: alkaloids, antioxidant activity, antimycobacterial activity.