New Isoflavonoids from the extract of Rhynchosia precatoria (Humb. & Bonpl. ex Willd.) DC. and their antimycobacterial activity

Abstract

Ethnopharmacology relevance: The evaluation of the antimycobacterial activity of extracts of medicinal plants used by Mayos against tuberculosis and respiratory problems, allowed the identification of Rhynchosia precatoria (Humb. & Bonpl. ex Willd.) DC (Fabaceae) as the best candidate to find new antimycobacterial compounds.

Aim of the study: To isolate and characterize the compounds of R. precatoria responsible for the inhibitory and bactericidal activity against Mycobacterium tuberculosis H37Rv and Mycobacterium smegmatis ATCC 700084. To determine antimycobacterial synergistic effect of pure compounds and their selectivity index towards Vero cells.

Materials and methods: A total of six flavonoids were purified by silica gel column chromatography. Structural elucidation of the isolated compounds was achieved by using 1D and 2D NMR spectroscopy techniques. The configuration at the C-3 chiral center was established by quantum mechanical calculation of the electronic circular dichroism (ECD) spectrum. In vitro inhibitory and bactericidal activity against M. tuberculosis and M. smegmatis were determined with the redox indicator Alamar Blue (resazurin). Synergy was determined by X/Y quotient. Cytotoxicity was measured by MTT assay.

Results: The isolated compounds were identified as precatorin A (1), precatorin B (2), precatorin C (3), lupinifolin (4), cajanone (5) and lupinifolinol (6). Compounds 1-3 are new. Compounds 1 to 5 inhibited the growth of M. tuberculosis (MIC ≥31.25µg/mL); compounds 1, 2, 4 and 5 killed the bacteria (MBC ≥31.25µg/mL) and also inhibited M. smegmatis (MIC ≥125µg/mL), while 1 and 4 also resulted bactericidal (MBC ≥125µg/mL). Compounds 4 and 5 presented synergistic effect (X/Y quotient value <0.5) at a concentration of 1/2 MIC of each compound in the combination. Cytotoxicity in murine macrophages (RAW 264.7 cells) gave IC₅₀ values of 13.3-46.98µM, for compounds 1-5.

Conclusions: In this work we isolated two new isoflavanones (1 and 2), and one new isoflavone (3) with a weak antimycobacterial activity. The (3R) absolute configuration was assigned to 1 by computational analysis of its ECD spectrum and to 2 and 5 by similarity of their ECD spectra with that of 1. We are also reporting by first time, activity against virulent strain of M. tuberculosis for compounds 4 and 5 and their antimycobacterial synergistic effect.

Keywords: Absolute configuration; Fabaceae; Isoflavonoids; Mycobacterium tuberculosis; Precatorins; Rhynchosia precatoria.