Investigação fitoquímica das folhas de Strychnos subcordata Spruce ex Benth. (Loganiaceae)

Abstract

The genus Strychnos L., the principal representative of the Loganiaceae family, is native to tropical and subtropical regions, encompassing 204 accepted species. This genus is globally recognized for its traditional use in the preparation of curare, a arrow poison utilized by indigenous peoples of South America, and stands out as a promising source of bioactive natural products. Although the phytochemical content of several Strychnos species has been extensively investigated, others remain largely unexplored, such as S. subcordata Spruce ex Benth., a liana species distributed throughout the Amazon region, traditionally used in the preparation of curare. Thus, the present study aimed to contribute to the phytochemical knowledge of the Strychnos genus in the Amazon region through the dereplication and/or isolation of substances from the leaves of S. subcordata. To achieve this, the leaves were subjected to infusion extraction, and the resulting aqueous extract was submited to solid-phase extraction (SPE) procedure. The obtained methanol fraction was analyzed by direct injection into a ion trap mass spectrometer (DI-MS), employing neutral loss scanning mode through data-dependent acquisition (DDA) for the recognition and dereplication of flavonoid glycosides. Subsequently, the methanol fraction was analyzed by high-performance liquid chromatography with ultraviolet detection (HPLC-UV) and high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS), utilizing six different columns (Luna C18(2), Kinetex C8, Luna PFP(2), Luna phenyl-hexyl, Luna CN, and Kinetex biphenyl). Based on these analyses, several offline single heart-cutting two-dimensional liquid chromatography (2D-LC) strategies were proposed to isolate the five major compounds from the methanol fraction of S. subcordata leaves, three of which could not be preliminarily identified by MS analyses. The combination of a C18 column in the first dimension (1D) and a biphenyl column in the second dimension (2D) was selected as the strategy for the isolation of a previously undescribed isoflavone, named subcordatine; two rare flavonoid glycosides (apigenin 6-C-[β-apiofuranosyl-(1→2)]-β-glucopyranoside and apigenin 7-methyl ether 6-C-[β-apiofuranosyl-(1→2)]-β-glucopyranoside); as well as the flavone glycosides isovitexin and swertisin. The structures of these compounds were elucidated by integrating manual interpretation of MS spectra and one-dimensional and two-dimensional nuclear magnetic resonance (1D and 2D NMR) data. In an effort to understand the column selectivity observed for the investigated compounds, a discussion was presented based on the solute-column interactions predicted by Snyder’s hydrophobic subtraction model (HSM). In particular, subcordatine exhibited greater retention on phenyl-type columns, especially on the biphenyl column, likely due to conformational similarities between isoflavones and the biphenyl groups of the stationary phase. These observations underscore the importance of specific molecular interactions in chromatographic selectivity and highlight the potential of 2D-LC methodologies in the offline heart-cutting mode for isolating structurally complex natural products. Furthermore, this study revealed S. subcordata as a promising source of bioactive substances, expanding the phytochemical knowledge of the Strychnos genus.