Analysis of bioactive constituents from the leaves of Amorpha fruticosa L

Abstract

Amorpha fruticosa L. is a Chinese folk medicine and rich in polyphenols. Fifteen known compounds were isolated and identified from the leaves of A. fruticosa L. They are tephrosin (1), 6a,12a-dehydrodeguelin (2), vitexin (3), afrormosin-7-O-β-d-glucopyranoside (4), 2″-O-α-l-rhamnopyranosyl isovitexin (5), rutin (6), chrysoeriol (7), 7-O-methylluteolin (8), trans-p-coumaric acid (9), 2-benzyl-4,6-dihydroxybenzoic acid-4-O-β-d-glucopyranoside (10), formononetin (11), quercetin (12), apigenin (13), β-sitosterol (14), and β-daucosterol (15). Compounds 3, 4, 5, and 7-9 were isolated from A. fruticosa L. for the first time. Cytotoxicity of individual compounds 3-10 and 90% ethanol extract against human cancer cell lines HCT116 and HepG2 were reported. The results suggested that compounds 7 and 8, and the crude extract exhibited inhibitory effects on human cancer cell line HCT116, at concentrations of 100 μg/mL, 5 μg/mL, and 25 μg/mL at <60% of cell viability rate, respectively. In addition, a valid high-performance liquid chromatography diode array detector method was established to quantitatively analyze compounds 1-12 in the leaves of A. fruticosa L., which was harvested at three different stages of maturity from May 20 to August 10, 2014. The results demonstrated that contents were greatly influenced by the maturity. Total amounts of the analytical constituents gradually increased from May 20 to August 10, with the values ranging from 10.86 mg/g to 18.84 mg/g, whereas bioactive compounds 7 and 8 presented the opposite variation trend. The results of this study may provide data for further study and comprehensive utilization of A. fruticosa L.

Keywords: Amorpha fruticosa L.; Leguminosae; cytotoxicity; phenolics; quantitative analysis.

Figure 1

Chemical structures of compounds 1–12 isolated from the leaves of A. fruticosa L.

Figure 2

Effect of compounds 3–10 and 90% ethanol crude extract on the viability of (A) HCT116 and (B) HepG2 cells. Cells were treated with 5 μg/mL, 10 μg/mL, 25 μg/mL, 50 μg/mL, or 100 μg/mL concentrations of the indicated compounds and 90% ethanol crude extract for 48 hours. Cell viability rate was then determined as described under the Materials and methods section.

Figure 3

HPLC chromatograms of solution of (A) standards and (B) the sample A. fruticosa L. leaves at 295 nm. Peaks: 1, tephrosin (47.28 minutes); 2, 6a,12a-dehydrodeguelin (52.06 minutes); 3, vitexin (12.86 minutes); 4, afrormosin-7-O-β-d-glucopyranoside (22.03 minutes); 5, 2″-O-α-l-rhamnopyranosyl isovitexin (18.81 minutes); 6, rutin (16.42 minutes); 7, chrysoeriol (43.04 minutes); 8, 7-O-methylluteolin (34.01 minutes); 9, trans-p-coumaric acid (12.53 minutes); 10, 2-benzyl-4,6-dihydroxybenzoic acid-4-O-β-d-glucopyranoside (6.46 minutes); 11, formononetin (44.08 minutes); and 12, quercetin (28.70 minutes). HPLC =high-performance liquid chromatography.

Figure 4

Content of compounds 1–12 in three different batches of A. fruticosa L. leaves.