Screening of Korean Natural Products for Anti-Adipogenesis Properties and Isolation of Kaempferol-3-O-rutinoside as a Potent Anti-Adipogenetic Compound from Solidago virgaurea

Abstract

In this study, the anti-adipogenetic activity of 300 plant extracts was investigated using an Oil Red O staining assay in a 3T3-L1 cell line. Our results indicate that three plants, including the stem and leaf of Physalis angulata, the whole grass of Solidago virgaurea, and the root of Dioscorea nipponica, produced over 90% inhibition of adipogenesis. Kaempferol-3-O-rutinoside, which demonstrated a 48.2% inhibitory effect on adipogenesis without cytotoxicity, was isolated from the butanol layer of a water extract of S. virgaurea guided by the anti-adipogenesis assay in 3T3-L1. PPAR-γ and C/EBPα expression levels were determined using western blot, and our results indicate that kaempferol-3-O-rutinoside has a strong anti-adipogenic effect in 3T3-L1 cells through the suppression of increases in PPAR-γ and C/EBPα expression.

Keywords: Solidago virgaurea; kaempferol-3-O-rutinoside; obesity; screening.

Figure 1

Effect of the SV extracts on inhibition 3T3-L1 adipocyte differentiation. (A) The relative lipid content; (B) Oil Red O staining images. Stained triglyceride content was quantified by measuring absorbance at 540 nm. Con is control group; DM is differentiation media cells; SVMC is methyl chloride extract of S. virgaurea var. gigantea, SV70E is 70% ethanol extract of S. virgaurea var. gigantea; SVW is water extract of S. virgaurea var. gigantea. The concentration is 10 μg/ml; three independent experiments have been carried out; * p < 0.05 vs. DM; *** p < 0.005 vs. DM.

Figure 2

Effect of SVW solvent fractions on inhibition of 3T3-L1 adipocyte differentiation. (A) The relative lipid content; (B) Oil Red O staining images. Confluent 3T3-L1 preadipocyte were differentiated into adipocytes in medium with or without 10 μg/mL of SVW fractions for 8 day. Con is control group; DM is differentiation media cells; SVW-Ef is ethyl acetate fraction from S. virgaurea var. gigantea water extract; SVW-Bf is n-butanol fraction from S. virgaurea var. gigante water extract; SVW-Wf is water fraction from S. virgaurea var. gigantea water extract. Three independent experiments have been carried out; * p < 0.05 vs. DM; *** p < 0.005 vs. DM.

Figure 3

Effect of the SVW and the SVW-Bf on preadipocyte viability. 3T3-L1 preadipocytes were incubated with SVW and SVW-Bf at various concentrations (10, 50 and 100 μg/mL) for 24 h, 48 h, and 72 h. SVW is S. virgaurea var. gigantea water extracts; SVW-Bf is n-butanol fraction of S. virgaurea var. gigante water extract. Three independent experiments have been carried out.

Figure 4

Effect of Diaion HP-20 fractions of SVW-Bf in 3T3-L1. (A) The relative lipid content; (B) Oil Red O staining images. Confluent 3T3-L1 preadipocytes were differentiated into adipocytes in medium with or without different concentrations of SVW-Bf Diaion HP-20 fractions for 8 days. Three independent experiments have been carried out; * p < 0.05 vs. DM.

Figure 5

Molecular structures of (A) protocatechuic acid; (B) chlorogenic acid; and (C) kaempferol-3-O-rutinoside; (D) preadipocyte viability effect of protocatechuic acid, chlorogenic acid and kaempferol-3-O-rutinoside; relative lipid content and Oil Red O staining images of (E) protocatechuic acid; (F) chlorogenic acid; and (G) kaempferol-3-O-rutinoside. Three independent experiments have been carried out; * p < 0.05 vs. DM; ** p < 0.01 vs. DM.

Figure 6

Effect of kaempferol-3-O-rutinoside on PPAR-γ (A,C) and C/EBP-α (B,D) expression in 3T3-L1 cells. Three independent experiments have been carried out; * p < 0.05 vs. DM; ** p < 0.01 vs. DM. mL not ml.

Figure 7

Extraction and fractionation of SV. ^a Yield; ^b Inhibition % (concentrations of protocatechuic acid, chlorogenic acid, kaempferol-3-O-rutinoside are 50 μg/mL; concentrations of others are 10 μg/mL).