Bioactivity-guided isolation of anti-inflammation flavonoids from the stems of Millettia dielsiana Harms
Graphical abstract
Introduction
Millettia dielsiana Harms (Family: Leguminosae) is a kind of semi-evergreen perennial woody vine plant and its stems have been widely used by the ethnic minorities (Yi, Miao and Yao) in China to treat muscle aches and pains, rheumatic arthritis and gynecological diseases, such as anemia and amenorrhea [1]. As part of our ongoing research program for the discovery of plant-derived anti-inflammation constituents, we found that the EtOAc extract of the stems of M. dielsiana showed strong inhibitory activity against the production of NO and TNF-α in RAW 264.7 cells. Based on the bioassay-guided fractionation of EtOAc extract, two new isoflavones along with nine known isoflavones were isolated from the bioactive fractions of EtOAc extract from stems of M. dielsiana. Millesianin C (1) showed excellent inhibitory activity towards NO and TNF-α. Their structures were characterized on the basis of spectroscopic methods including extensive 2D NMR and MS spectrometry.
Section snippets
General
Column chromatography was carried out on silica gel 60 (0.2–0.5 mm, 0.040–0.063 mm, Merck, Darmstadt, German). Medium-pressure liquid chromatography (MPLC) (Büchi, Flawil, Switzerland) was performed with a C-605 pump, a C-615 pump manager, a C-635 UV detector, a C-660 fraction collector and a Sepacore record 1.0 workstation. Counter-current chromatography was performed on a Midi-DE centrifuge (Dynamic Extraction, Slough, UK). HPLC was performed on a Waters Alliance 2695 separation module (Empower
Chemistry
Compound 9 was isolated as a white powder. The molecular formula was determined as C22H22O5 by HRESIMS of its [M + H]+ ion peak at m/z = 367.1545 (calcd. 366.1467). The UV spectrum absorption at 256 and 319 nm and 1H (δ 7.99 for H-2) and 13C (δ 152.01 for C-2) NMR spectra showed this compound to be an isoflavone type skeleton. The 1H NMR (Table 1) spectrum displayed ortho-coupled A2B2-type protons at δ 7.50 and 6.98 (each 2H, d, J = 8.4 Hz) assignable to H-2′, 6′ and H-3′, 5′, respectively of ring B
Conclusion
Based on the bioactivity-guided isolation, isoflavones (1–11) were separated from the bioactive fractions of Fr.5, Fr.6 and Fr.8 from EtOAc extract of M. dielsiana Harms, which were then tested for the inhibitory activity of NO production and TNF-α secretion in LPS-stimulated RAW 267.4 cells. Among the tested compounds, Millesianin C (1) with 2,2-dimethylpyran fragment, had the most potent anti-inflammatory effect decreasing NO production similar to that of dexamethasone and decreasing TNF-α
Acknowledgment
The work was supported by Guangdong Innovative Research Team Program (2011Y073), the National Science Foundation for Post-doctoral Scientists of China (2013M530403) and the National Natural Science Foundation of China (81373283). The authors have declared no conflict of interest.
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Both authors have contributed equally to the paper.