• Journal of Microbiology and Biotechnology
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• 제27권7호
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• pp.1233-1241
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• 2017

The ginsenoside Rh2 has strong anti-cancer, anti-inflammatory, and anti-diabetic effects. However, the application of ginsenoside Rh2 is restricted because of the small amounts found in Korean white and red ginsengs. To enhance the production of ginsenoside Rh2-MIX (comprising 20(S)-Rh2, 20(R)-Rh2, Rk2, and Rh3 as a 10-g unit) with high specificity, yield, and purity, a new combination of enzymatic conversion using the commercial enzyme Viscozyme L followed by acid treatment was developed. Viscozyme L treatment at pH 5.0 and $50^{\circ}C$ was used initially to transform the major ginsenosides Rb1, Rb2, Rc, and Rd into ginsenoside F2, followed by acid-heat treatment using citric acid 2% (w/v) at pH 2.0 and $121^{\circ}C$ for 15 min. Scale-up production in a 10-L jar fermenter, using 60 g of the protopanaxadiol-type ginsenoside mixture from ginseng roots, produced 24 g of ginsenoside Rh2-MIX. Using 2 g of Rh2-MIX, 131 mg of 20(S)-Rh2, 58 mg of 20(R)-Rh2, 47 mg of Rk2, and 26 mg of Rh3 were obtained at over 98% chromatographic purity. Then, the anti-cancer effect of the four purified ginsenosides was investigated on B16F10, MDA-MB-231, and HuH-7 cell lines. As a result, these four rare ginsenosides markedly inhibited the growth of the cancer cell lines. These results suggested that rare ginsenoside Rh2-MIX could be exploited to prepare an anti-cancer supplement in the functional food and pharmaceutical industries.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 2006년도 춘계학술대회
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• pp.3-12
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• 2006

1 The present work was performed to investigate the effects of ginsenoside Rh2 on proliferation, cell cycle-regulation and differentiation of human leukemia HL-60 cells as well as the underlying mechanisms for these effects. 2 Ginsenoside Rh2 potently inhibited the proliferation of HL-60 cells in both a dose- and time-dependent manner with an $IC_{50}$, $20{\mu}M$. 3 DNA flow-cytometry indicated that ginsenoside Rh2 markedly induced a $G_1$ phase arrest of HL-60 cells. 4 Among the $G_1$ phase cell cycle-related proteins, the levels of cyclin-dependent kinase(CDK)4, 6 and cyclin D1, cyclin D2, cyclin D3 were reduced by ginsenoside Rh2, whereas the steadystate levels of CDK2 and cyclin E were unaffected. 5 The protein levels of a CDK inhibitor p16, $p21^{CIP1/WAF1}$ and $p27^{KIP1}$ were markedly increased by ginsenoside Rh2. 6 Ginsenoside Rh2 markedly enhanced the binding of $p21^{CIP1/WAF1}$ and $p27^{KIP1}$ with CDK2 and CDK6, resulting in the reduced activity of both kinases and the hypophosphorylation of Rb protein. 7 We furthermore suggest that ginsenoside Rh2 is a potent inducer of the differentiation of HL-60 cells, based on observations such as a reduction of the nitroblue tetrazolium level, an increase in the esterase activities and phagocytic activity, morphology changes, and the expression of CD11b, CD14, CD64 and CD66b surface antigens. 8 In conclusion, the onset of ginsenoside Rh2-induced the $G_0/G_1$ arrest of HL-60 cells prior to the differentiation is linked to a sharp up-regulation of the $p21^{CIP1/WAF1}$ level and a decrease in the CDK2, CDK4 and CDK6 activities. This is the first report demonstrating that ginsenoside Rh2 potently inhibits the proliferation of human promyelocytic HL-60 cells via the $G_1$ phase cell cycle arrest and differentiation induction.

• Archives of Pharmacal Research
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• 제27권8호
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• pp.834-839
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• 2004

In human neuroblastoma SK-N-BE(2) cells undergoing apoptotic death induced by ginsenos-ide Rh2, a dammarane glycoside that was isolated from Panax ginseng C. A. Meyer, caspase-1 and caspase-3 were activated. The expression of Bax was increased in the cells treated with ginsenoside Rh2, whereas Bcl-2 expression was not altered. Treatment with caspase-1 inhibi-tor, Ac-YVAD-CMK, or caspase-3 inhibitor, Z-DEVD-FMK, partially inhibited ginsenoside Rh2-induced cell death but almost suppressed the cleavage of the 116 kDa PARP into a 85 kDa fragment. When the levels of p53 were examined in this process, p53 accumulated rapidly in the cells treated early with ginsenoside Rh2. These results suggest that activation of caspase-1 and -3 and the up-regulation of Bax are required in order for apoptotic death of SK-N-BE(2) cells to be induced by ginsenoside Rh2, and p53 plays an important role in the pathways to promote apoptosis.

• Journal of Ginseng Research
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• 제38권1호
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• pp.16-21
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• 2014

Ginseng saponins exert various important pharmacological effects with regard to the control of many diseases, including cancer. In this study, the anticancer effect of ginsenosides on human cancer cells was investigated and compared. Among the tested compounds, ginsenoside-Rh2 displays the highest inhibitory effect on cell viability in HepG2 cells. Ginsenoside-Rh2, a ginseng saponin isolated from the root of Panax ginseng, has been suggested to have potential as an anticancer agent, but the underlying mechanisms remain elusive. In the present study, we have shown that cancer cells have differential sensitivity to ginsenoside-Rh2-induced apoptosis, raising questions regarding the specific mechanisms responsible for the discrepant sensitivity to ginsenoside-Rh2. In this study, we demonstrate that AMP-activated protein kinase (AMPK) is a survival factor under ginsenoside-Rh2 treatment in cancer cells. Cancer cells with acute responsiveness of AMPK display a relative resistance to ginsenoside-Rh2, but cotreatment with AMPK inhibitor resulted in a marked increase of ginsenoside-Rh2-induced apoptosis. We also observed that p38 MAPK (mitogen-activated protein kinase) acts as another survival factor under ginsenoside-Rh2 treatment, but there was no signaling crosstalk between AMPK and p38 MAPK, suggesting that combination with inhibitor of AMPK or p38 MAPK can augment the anticancer potential of ginsenoside Rh2.

• 약학회지
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• 제38권4호
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• pp.425-429
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• 1994

The genuine aglycone, 20(S)-protopanaxadiol, obtained from the leaves of Panax ginseng as a result of direct alkaline treatment was isolated and characterized by spectroscopic evidences. The study on the yield of genuine aglycone which is produced from the treatment of some kinds of alkali was carried out. $Ginsenoside-Rh_2$ was synthesized by conjugation of 2,3,4,6-tetra-O-acetyl-${\alpha}$-D-glucopyranosyl bromide to 20(S)-protopanaxadiol in the presence of silver carbonate and cadmium cabonate. The preparation of $ginsenoside-Rh_2$ by this method is a new one which the yield of this saponin can be improved in the mild condition.

• Journal of Ginseng Research
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• 제41권3호
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• pp.330-338
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• 2017

Background: Ginsenoside Rh2 (G-Rh2) is a ginseng saponin that is widely investigated because of its remarkable antitumor activity. However, the molecular mechanism by which (20S) G-Rh2 triggers its functions and how target animals avoid its cytotoxic action remains largely unknown. Methods: Phage display was used to screen the human targets of (20S) G-Rh2. Fluorescence spectroscopy and UV-visible absorption spectroscopy were used to confirm the interaction of candidate target proteins and (20S) G-Rh2. Molecular docking was utilized to calculate the estimated free energy of binding and to structurally visualize their interactions. MTT assay and immunoblotting were used to assess whether human serum albumin (HSA), bovine serum albumin (BSA), and bovine serum can reduce the cytotoxic activity of (20S) G-Rh2 in HepG2 cells. Results: In phage display, (20S) G-Rh2-beads and (20R) G-Rh2-beads were combined with numerous kinds of phages, and a total of 111 different human complementary DNAs (cDNA) were identified, including HSA which had the highest rate. The binding constant and number of binding site in the interaction between (20S)-Rh2 and HSA were $3.5{\times}10^5M^{-1}$ and 1, and those in the interaction between (20S) G-Rh2 and BSA were $1.4{\times}10^5M^{-1}$ and 1. The quenching mechanism is static quenching. HSA, BSA and bovine serum significantly reduced the proapoptotic effect of (20S) G-Rh2. Conclusion: HSA and BSA interact with (20S) G-Rh2. Serum inhibited the activity of (20S) G-Rh2 mainly due to the interaction between (20S) G-Rh2 and serum albumin (SA). This study proposes that HSA may enhance (20S) G-Rh2 water solubility, and thus might be used as nanoparticles in the (20S) G-Rh2 delivery process.

• Journal of Ginseng Research
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• 제30권3호
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• pp.95-99
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• 2006

When the inhibitory effect of ginsenoside (G) Re isolated from ginseng and its metabolites G-Rg1, G-F1, G-Rh1 and protopanaxatriol in mouse ear skin psoriasis stimulated by oxazolone was investigated, G-Re and its metabolites suppressed mouse ear swelling stimulated by oxazolone. Among these agents tested, G-Rh1 most potently suppressed ear swelling as well as mRNA expression of COX-2 and proinflammatory cytokines $IL-1{\beta},\;TNF-{\alpha}$ and $interferon-{\gamma}$. These findings suggest that G-Rh1 may improve chronic dermatitis and psoriasis.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 1998년도 Advances in Ginseng Research - Proceedings of the 7th International Symposium on Ginseng -
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• pp.244-252
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• 1998

Ginsenoside Rh, (G-Rh,) from Panax ginseng induced morphological features of apoptosis and DNA fragmentation as a biochemical marker of apoptosis confirmed by TUNEL reaction and agarose gel electrophoresis in human neuroblastoma SK-N-BE(2) and rat glioma C6Bu-1 cells During apoptosis by G-Rh2, protein kinase C (PKC) isoforms were analysed by immunoblotting. In SK-N-BE(2) cells, the levels of a, p and ${\gamma}$ subtypes were increased by undergoing apoptosis, while PKC e isoform increased early in treatment (3 h and 6 h). In addition, PKC s isoform gradually decreased during apoptosis by G-Rh2 and PKC $\theta$ isoform was detected in neither untreated- nor G-Rh1-treated SK-N-BE(2) cells (data not shown). However, no significant changes in the level of S and s isoforms were observed in C6Bu-1 cells undergoing apoptosis by G-Rh2. These results suggest that PKC subtypes may play differential roles in apoptotic signal pathways and their roles can be cell type-specific in apoptosis induced by G-Rh2.

• 한국식품과학회지
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• 제13권1호
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• pp.57-66
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• 1981

인삼주성분 사포닌 및 미량성분 사포닌의 단리법에 대하여 HPLC의 응용을 검토하여 이미 보고한 바 있다. 따라서 인삼미량성분인 $Ginsenoside-Rh_1$ 및 미지성분인 $Ginsenoside-Rh_2$의 단리법을 검토하였다. 저자등의 방법에 따라 홍삼의 70% 에탄올추출액(4 kg)에서 부탄올추출액을 얻어 에틸 아세테이트로 처리 (실온에서 3시간 및 $60^{\circ}C$에서 3 시간)하여 에틸 아세테이트 획분(29g)을 preparative HPLC인 prep LC/System-500을 사용하여 부분분획하여 각 획분을 analtical HPLC/ALC-201로 동정, ginsenoside-Rh group을 다시 에테르, 에틸 아세테이트 및 물로 정제한 후, semipreparative HPLC/ALC-201로 $ginsenoside-Rh_1$ 및 $-Rh_2$를 단리했다. 미지성분단리에 HPLC의 응용이 효과적임이 판명되었다.

• Archives of Pharmacal Research
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• 제23권5호
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• pp.518-524
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• 2000

We examined the modulation of protein kinase C (PKC) subtypes during apoptosis induced by ginsenoside Rh2 (G-Rh2) in human neuroblastoma SK-N-Bl(2) and rat glioma C6Bu-1 cells. Apoptosis induced by C-Rh2 in both cell lines was confirmed, as indicated by DNA fragmentation and in situ strand breaks, and characteristic morphological changes. During apoptosis induced by G-Rh2 in SK-N-BE(2) cells, PKC subtypes $\alpha$, $\beta$ and $\gamma$ were progressively increased with prolonged treatment, whereas PKC $\delta$ increased transiently at 3 and 6 h and PKC $\varepsilon$ was gradually down-regulated after 6 h following the treatment. On the other hand, PKC subtype $\beta$ markedly increased at 24 h when maximal apoptosis was achieved. In C6Bu-l cells, no significant changes in PKC subtypes $\alpha$, $\gamma$, $\delta$, $\varepsilon$ and $\beta$ were observed during apoptosis induced by G-Rh2. These results suggest the evidence for a possible role of PKC subtype in apoptosis induced by G-Rh2 in SK-N-BE(2) cells but not in C6Bu-1 cells, and raise the possibility that G-Rh2 may induce apoptosis via different pathways interacting with or without PKC in different cell types.