Effect of Fermented Red Ginseng Extract Enriched in Ginsenoside Rg3 on the Differentiation and Mineralization of Preosteoblastic MC3T3-E1 Cells

Abstract

In this study, red ginseng extract (RGE) was converted into high-content minor ginsenosides by fermenting with Bgp1 enzymes at 37°C for 5 days. Compared to the RGE, the minor ginsenoside contents were increased in fermented red ginseng extract (FRGE). Moreover, the amount of minor ginsenosides such as Rh1 (11%) and Rg2 (16%) was slightly augmented, while the level of Rg3 (33%) was significantly increased after bioconversion. Furthermore, we also examined and compared the effect of RGE and FRGE on the differentiation and mineralization of preosteoblastic MC3T3-E1 cells. Similarly, the level of mRNA expression of intracellular alkaline phosphatase (ALP) activity, type-1 collagen (Col-I) was also increased. Based on the comparison, it is clear that the FRGE has improved effects on bone formation and differentiation of preosteoblastic MC3T3-E1 cells.

Keywords: Panax ginseng; bone formation; differentiation; mineralization; osteoblast; β-glycosidase.

FIG. 1.

High-performance liquid chromatography (HPLC) analysis of the biotransformation of crude extract of ginsenoside by Bgp1. (a) Red ginseng extract (RGE), (b) fermented red ginseng extract (FRGE). From the HPLC analysis, it is concluded that Rg1 and Re are fully converted into other minor ginsenosides, whereas a high amount of Rg3 (33%) was identified compared to other minor ginsenosides (Rf, Rh1, and Rg2). Color images available online at www.liebertpub.com/jmf

FIG. 2.

Effect of FRGE on the growth of MC3T3-E1 cells (a), ALP (b), and (c) Col-I of osteoblastic MC3T3-E1 cells. Data were expressed as a percentage of control. *P<.05, **P<.01, control versus RGE, ^#P<.05, ^##P<.01 RGE versus FRGE.

FIG. 3.

Effect of FRGE on the growth of MC3T3-E1 cells. For mineralization, Alizarin Red S staining (a, b), MC3T3-E1 cells were exposed to a culture medium containing β-glycerophosphate and ascorbic acid for 24 days. **P<.01, control versus RGE, ^#P<.05, ^##P<.01, ^###P<.001 RGE versus FRGE. Color images available online at www.liebertpub.com/jmf

FIG. 4.

Effects of FRGE treated with β-glycosidase 1 (Bgp1) on ALP, BMP-2, Col-I, and on Runx-2 mRNA expression level in MC3T3-E1 cells were examined by reverse-transcriptase polymerase chain reaction. MC3T3-E1 cells were treated with (1, 10, and 100 μg/mL) or without FRGE fermented by β-glycosidase 1 (Bgp1) for 12 days. The GAPDH mRNA level was analyzed in the same samples as a reference gene. Each value is the mean±SEM of three to five independent experiments. **P<.01, ***P<.001 control versus RGE, ^##P<.01, ^###P<.001 RGE versus FRGE. Hence, 10 μg/mL provides a significant effect on ALP, BMP-2, Col-I, and on Runx-2 expression in MC3T3-E1 cells. BMP, bone morphogenetic protein; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.