ESTUDO DOS METABÓLITOS SECUNDÁRIOS DAS FOLHAS DE Coussarea hydrangeifolia (Benth.) Müll.Arg. (RUBIACEAE) E AVALIAÇÃO DE SUAS ATIVIDADES CITOTÓXICAS

Abstract

The area of Natural Product Chemistry studies plants as a source of bioactive products. Among the most promising plant families regarding the diversity of bioactive secondary metabolites is Rubiaceae, for which several biological activities have been reported, including anticancer activity. Coussarea hydrangeifolia is a Rubiaceae species popularly known as quina-branca, whose tea made from its leaves have the ability to strengthen the organism. The literature reports only two studies on this species, however, with low information about its chemical composition and biological properties. Thus, the present work aimed at the phytochemical investigation of the leaves of C. hydrangeifolia collected in Campo Grande/MS and the evaluation of its cytotoxic potential against human leukemia tumor cell lines, named K562 and Jurkat. For this purpose, the leaves were extracted with 95% ethanol and the resulting extract (EFCH) was successively partitioned using hexane, dichloromethane and ethyl acetate, yielding the corresponding phases. Among them, the hexane and dichloromethane phases were selected to continue the chemical study by combining column chromatography techniques using silica gel 60 (70-230 and 230-400 mesh) and Sephadex LH-20®. The isolated substances were submitted to 1H and 13C NMR analysis (one- and two-dimensional). Such chromatographic fractionations resulted in the isolation and further structural elucidation of 26 secondary metabolites: a carotenoid, lutein (1); one diterpene E-phytol (4); two tocopherols, α and β-tocopherol (3 and 5); five steroids, stigmasterol (6), β-sitosterol (7), campesterol (8), glycosylated stigmasterol and glycosylated β-sitosterol (24 and 25); a glycosylated iridoid, asperuloside (26); and sixteen triterpenes, squalene (2), hederagenin (17), 4-epi-hederagenin (18), 3-epi-ursolic acid (9), 3-epi-olenolic acid (10), barbinervic acid (11), 3-epi-pomolic acid (12), 3α,19α-dihydroxyurs-12-en-24,28-dioic acid (13), rotungenic acid (15), 19α,24-dihydroxyurs-12-en-3-one-28-oic acid (16), 24-hydroxyursolic acid (19), 3α,24-dihydroxyurs-12-en-28-oic acid (20), rotundic acid (21), scutellaric acid (22), 3-epi-sparthodic acid (23) and hydrangifolic acid (14), where the latter has never been described. Among the isolated substances, only seven had already been reported in the genus Coussarea and five are being described for the first time in the Rubiaceae family in this work. In addition, the extract and phases showed promising activities in both cell lines in the cytotoxicity assays, with best result for the dichloromethane phase against the Jurkat, with IC50 of 73.98 µg/mL after 48 hours. The evaluation of the cytotoxic activity of the isolated triterpenes indicated that most compounds showed cytotoxic action, highlighting the activities of rotungenic acid (15) and 3β,24-dihydroxyurs-12-en-28-oic acid (19), which showed very significant activity against Jurkat cells, with IC50 values of 12.53 and 12.69 µg/mL after 24h, and 10.20 and 9.57 µg/mL after 48h, respectively. It was also possible to point out relevant observations regarding the structure-activity of triterpenes, such as: the ursane skeleton shows better activity, the β orientation of the hydroxyl group at C-3 increases activity, and the presence of a hydroxyl group at C-24 enhances cytotoxicity. Finally, these results contribute to a better understanding of the chemical constitution of the genus Coussarea and the species C. hydrangeifolia, which was shown to be a rich source of polyhydroxylated triterpenes and showed promising results of cytotoxic activity for tumor cells.

Keywords: Rubiaceae, Coussarea, triterpenes, cytotoxic activity, leukemia.