Ginsenosides Rk1 Research Articles

Ginsenoside Rk3 Treats Corneal Injury Through the HMGB1/TLR4/NF-κB Pathway.

The cornea serves as a vital protective shield for the eye, safeguarding its intricate internal structures from external threats. Damage to the cornea compromises this protective function, triggering inflammation and potentially causing long-term harm. While ginsenoside Rk3 has demonstrated potential for repairing the corneal barrier and reducing inflammation, its effectiveness in treating corneal damage remains relatively unexplored. This comprehensive study uses both in vivo and in vitro models to investigate the therapeutic capabilities of ginsenoside Rk3. Using two models of corneal damage, a benzalkonium chloride-induced mouse model and a high osmolarity-induced human corneal epithelial cell model, we scrutinized the effects of ginsenoside Rk3 treatment. Our results showed that ginsenoside Rk3-treated mice manifested reduced corneal damage and inflammation compared with their untreated counterparts. Furthermore, mice treated with ginsenoside Rk3 exhibited an organized arrangement of corneal cells and diminished stromal layer thickness, indicating reparative properties of ginsenoside Rk3. Additionally, ginsenoside Rk3 increased the expression of tight junction proteins, suppressed inflammatory factors, and decreased HMGB1 protein expression, thereby modulating downstream signaling pathways. Collectively, our findings present compelling evidence that ginsenoside Rk3 is a promising therapeutic option for corneal injury. By repairing the corneal barrier, mitigating inflammation, and modulating specific protein levels, ginsenoside Rk3 opens new avenues for managing corneal damage.

Ginseng fruit rare saponins (GFRS) improved inflammatory response: In vitro and in vivo assessment

Inflammation is the body's protective immune response to tissue damage. Ginseng has a long history of medicinal use, and its active ingredient ginsenosides have anti-inflammatory effects. Ginseng fruit rare saponins (GFRS) is a transformation product of ginseng saponins and rich in a variety of rare saponins. We used HPLC-DAD method to study GFRS rare saponins with ginsenoside F4, R-Rg3, SRg3, Rk1, Rg6, Rg5, Rk3 and Rh4. However, there is no study on the use of GFRS to reduce skin inflammation. This study enriched the action pathway of GFRS through network pharmacology and revealed the anti-inflammatory effect of GFRS for the first time. In vitro experiments showed that GFRS could significantly reduce the release of NO in lipopolysaccharide (LPS) -induced RAW264.7 cells and HaCaT cells, and reduce the secretion and expression of inflammation-related factors Interleukin-6 (IL-6), Tumor necrosis factor-α (TNF-α) and Interleukin-17 A (IL-17 A), thereby reducing cell inflammatory damage. In the imiquimod (IMQ) -induced mouse inflammatory model, the therapeutic effect of GFRS on the pathogenesis of psoriasis-like dermatitis was studied. In vivo experiments showed that the skin erythema, scales, thickness and inflammatory infiltration of GFRS-treated mice were reduced, and the psoriasis area severity index score was significantly lower than that of IMQ group. GFRS restored IMQ-induced spleen size and reduced the secretion and expression of TNF-α, IL-6, Interferon-γ (IFN-γ) and IL-17 A in serum. In summary, our results demonstrate that GFRS alleviates IMQ-induced dermatitis symptoms, effectively reduces the secretion of inflammatory factors, and inhibits IL-17 A expression.

Ginsenoside Rk3 Regulates Tryptophan Metabolism along the Brain-Gut Axis by Targeting Tryptophan Hydroxylase and Remodeling the Intestinal Microenvironment to Alleviate Depressive-Like Behavior in Mice.

Depression is a neuropsychiatric disease that significantly impacts the physical and mental health of >300 million people worldwide and places a major burden on society. Ginsenosides are the main active ingredient in ginseng and have been proven to have various pharmacological effects on the nervous system. Herein, we investigated the antidepressant effect of ginsenoside Rk3 and its underlying mechanism in a murine model of depression. Rk3 significantly improved depression-like behavior in mice, ameliorated the disturbance of the hypothalamus-pituitary-adrenal axis, and alleviated neuronal damage in the hippocampus and prefrontal cortex of mice. Additionally, Rk3 improved the abnormal metabolism of tryptophan in brain tissue by targeting tryptophan hydroxylase, thereby reducing neuronal apoptosis and synaptic structural damage in the mouse hippocampus and prefrontal cortex. Furthermore, Rk3 reshaped the composition of the gut microbiota of mice and regulated intestinal tryptophan metabolism, which alleviated intestinal barrier damage. Thus, this study provides valuable insights into the role of Rk3 in the tryptophan metabolic cycle along the brain-gut axis, suggesting that Rk3 may have the potential for treating depression.

Increasing the Amounts of Bioactive Components in American Ginseng (Panax quinquefolium L.) Leaves Using Far-Infrared Irradiation.

Both the roots and leaves of American ginseng contain ginsenosides and polyphenols. The impact of thermal processing on enhancing the biological activities of the root by altering its component composition has been widely reported. However, the effects of far-infrared irradiation (FIR), an efficient heat treatment method, on the bioactive components of the leaves remain to be elucidated. In the present study, we investigated the effects of FIR heat treatment between 160 and 200 °C on the deglycosylation and dehydration rates of the bioactive components in American ginseng leaves. As the temperature was increased, the amounts of common ginsenosides decreased while those of rare ginsenosides increased. After FIR heat treatment of American ginseng leaves at an optimal 190 °C, the highest total polyphenolic content and kaempferol content were detected, the antioxidant activity was significantly enhanced, and the amounts of the rare ginsenosides F4, Rg6, Rh4, Rk3, Rk1, Rg3, and Rg5 were 41, 5, 37, 64, 222, 17, and 266 times higher than those in untreated leaves, respectively. Moreover, the radical scavenging rates for 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) and the reducing power of the treated leaf extracts were 2.17, 1.86, and 1.77 times higher, respectively. Hence, FIR heat treatment at 190 °C is an efficient method for producing beneficial bioactive components from American ginseng leaves.

Ginsenoside Rk3 Ameliorates Obesity-Induced Colitis by Modulating Lipid Metabolism in C57BL/6 Mice.

Lipid metabolism is closely related to obesity and its complications. Our previous study found that ginsenoside Rk3 (Rk3), a natural bioactive substance derived from ginseng, can effectively alleviate obesity-induced colitis, while its impact on the improvement of the lipid metabolism disorder remains unclear. Here, we demonstrated that Rk3 significantly alleviated inflammation, oxidative stress, and lipid dysregulation in high-fat diet-induced colitis C57BL/6 mice. The potential mechanism by which Rk3 mitigated colon inflammation in the context of obesity may involve the modulation of polyunsaturated fatty acid metabolism with specific attention to n-6 fatty acids, linoleic acid, and arachidonic acid. Rk3 intervention markedly reduced the production of pro-inflammatory factors (PGE2, PGD2, TXB2, HETE, and HODE) by inhibiting cyclooxygenase and lipoxygenase pathways, while enhancing the production of anti-inflammatory factors (EET and diHOME) via cytochrome P450 pathways. Our findings suggest that Rk3 is a potential anti-inflammatory natural drug that can improve obesity-induced intestinal inflammation by regulating lipid metabolism.

Effective constituents and protective effect of Mudan granules against Schwann cell injury

Ethnopharmacological relevanceDiabetic peripheral neuropathy (DPN) is the most common complication of diabetes. Mudan granules (MD) is a Chinese patent medicine for treating DPN, which is composed of nine Chinese medicinal herbs, including the radix of Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao or Astragalus membranaceus (Fisch.) Bge. (Huangqi in Chinese), rhizome of Corydalis yanhusuo W.T. Wang (Yanhusuo), radix and rhizome of Panax notoginseng (Burk.) F. H. Chen (Sanqi), radix of Paeonia lactiflora Pall. or Paeonia veitchii Lynch (Chishao), radix and rhizome of Salvia miltiorrhiza Bge. (Danshen), rhizome of Ligusticum chuanxiong Hort. (Chuanxiong), flowers of Carthamus tinctorius L. (Honghua), lignum of Caesalpinia sappan L. (Sumu), and caulis of Spatholobus suberectus Dunn (Jixueteng). MD was reported to have a protective effect on Schwann cell (SC) that is considered as an important therapeutic target of DPN. However, the constituents of MD have not been reported, and the effective constituents and protective pathways for MD against SC injury remain unclear. Aim of the studyThis study aimed to identify the constituents in MD, and to investigate the effective constituents and protective pathways of MD against high-glucose/lipid injury in SC. Materials and methodsThe chemical constituents of MD were identified using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS). Protective effect and effective constituents screening were performed in an in vitro SC injury model induced by high glucose and lipid levels. The protective pathways of MD and its effective constituents were investigated by western blotting assay of related proteins. ResultsA total of 136 constituents were identified in MD. MD downregulated the phosphorylation of extracellular-regulated protein kinases 1/2 (ERK1/2) and expression of cyclooxygenase-2 (COX-2) and upregulated the expression of sirtuin 2 (SIRT2). Seven effective constituents were screened out, including three from Sanqi [20(R)-ginsenoside Rh2, 20(S)-ginsenoside Rh2, and ginsenoside Rk3], one from Huangqi (astragaloside II), one from Danshen (danshensu), and two from Chuanxiong (chlorogenic and cryptochlorogenic acid). Six of the seven compounds, excluding danshensu, inhibited the phosphorylation of ERK1/2. Both astragaloside II and chlorogenic acid upregulated the expression of SIRT2, and cryptochlorogenic acid and danshensu downregulated the expression of COX-2. ConclusionsThe constituents of MD were firstly identified, and seven effective constituents were found. MD can protect SC against high-glucose and -lipid injury by downregulating ERK1/2 phosphorylation and COX-2 expression and upregulating SIRT2 expression. Seven effective constituents regulated the expression of these proteins. This study presented an important advance toward elucidating the chemical constituents, and the effective constituents and protective pathways of MD against high-glucose/lipid injury in SC, which is very helpful for investigating the action mechanism of MD on treating DPN, and could ultimately inform the development of effective quality control procedures for MD production.

Black ginseng: a novel medicine for treating heart failure.

Introduction: Black ginseng (BG) was processed by "steaming and drying" (generally nine times) repeatedly to produce "rare saponins" and secondary ginsenosides. Both ginseng (GS) and red ginseng (RG) were commonly used in treating heart failure (HF), and the latter was confirmed to be more potent, implying the presence of rare ginsenosides that contribute positively to the treatment of heart failure. Previous research indicated that rare ginsenosides are more abundant in BG than in RG. Consequently, this study aims to investigate the effects of BG and its components on HF to elucidate the active substances and their underlying mechanisms in the treatment of HF. Methods: The effects of BG and its fractions (water-eluted fraction (WEF), total saponin fraction (TSF), and alcohol-eluted fraction (AEF)) on rats with isoproterenol (ISO)-induced HF were explored, and steroids belonging to the hypothalamic-pituitary-adrenal (HPA) and hypothalamic-pituitary-gonadal (HPG) axes were determined quantitatively using the ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry (UPLC-QqQ-MS/MS) method. In addition, 16S rDNA sequencing was performed on the gut microbiota, followed by GC-MS analysis of short-chain fatty acids (SCFAs), and the biochemical indexes related to energy metabolism and the serum cyclic nucleotide system were also analyzed by ELISA. Results: Based on a thorough evaluation of energy metabolism and the endocrine system, it was observed that the effects of BG components on the hypothalamic-pituitary-thyroid (HPT) and HPA axes were more pronounced. Notably, the treatment efficacy of the low dose of the total saponin fraction (TSFL), water decoction (WD), and high dose of the polysaccharide fraction (PSFH) was superior based on pharmacodynamic indicators such as brain natriuretic peptide (BNP), creatine kinase (CK), and estradiol (E2)/T). Furthermore, the WD and BG components exhibited significant effects on androgens (T and androstenedione (A4)). The TSFL group exerts an anti-inflammatory effect by regulating Lactobacillus/Erysipelotrichales. The WD, PSFH, and TSFL may impact inflammatory cytokines through the gut microbiota (Lactobacillus/Erysipelotrichales) and their metabolites (acetate and butyrate), exerting an anti-inflammatory effect. Discussion: The BG and all its split components demonstrated varying levels of efficacy in alleviating HF, and TSF and PSF exhibited a significant protective effect on HF. The main active components in TSF were revealed to be ginsenosides Rk1, Rk3, 20-(S)-Rg3, and 20-(S)-Rh2 by the H9C2 cell experiment. The decoction of BG and its components exhibited a potent impact on androgen hormones, with an elevation trend. This phenomenon may be attributed to the activation of the eNOS-NO pathway through androgen regulation, thereby contributing to its anti-HF activities. The WD, PSFH, and TSFL may exert anti-inflammatory effects through the intestinal flora (Lactobacillaceae/Erysipelotrichaceae) and its metabolites (acetic acid and butyric acid), which affect the inflammatory factors. The different mechanisms of action of each component of HF also reflect the significance and necessity of the overall role of traditional Chinese medicine (TCM). Our research was the first to report that the E2/T is related to HF and can be used as an indicator to evaluate HF.

Ginsenoside RK3 promotes neurogenesis in Alzheimer's disease through activation of the CREB/BDNF pathway

Ethnopharmacological relevanceIn the ancient book “Shen Nong's Herbal Classic,” Panax ginseng CA Mey was believed to have multiple benefits, including calming nerves, improving cognitive function, and promoting longevity. Ginsenosides are the main active ingredients of ginseng. Ginsenoside RK3 (RK3), a rare ginsenoside extracted from ginseng, displays strong pharmacological potential. However, its effect on neurogenesis remains insufficiently investigated. Aim of the studyThis study aims to investigate whether RK3 improves learning and memory by promoting neurogenesis, and to explore the mechanism of RK3 action. Materials and methodsThe therapeutic effect of RK3 on learning and memory was determined by the Morris water maze (MWM) and novel object recognition test (NORT). The pathogenesis and protective effect of RK3 on primary neurons and animal models were detected by immunofluorescence and western blotting. Protein expression of cAMP response element-binding protein (CREB)/brain-derived neurotrophic factor (BDNF) signaling pathway was detected by western blotting. ResultsOur results showed that RK3 treatment significantly improved cognitive function in APPswe/PSEN1dE9 (APP/PS1) mice and C57BL/6 (C57) mice. RK3 promotes neurogenesis and synaptogenesis in the mouse hippocampus. In vitro, RK3 prevents Aβ-induced injury in primary cultured neurons and promotes the proliferation of PC12 as well as the expression of synapse-associated proteins. Mechanically, the positve role of RK3 on neurogenesis was combined with the activation of CREB/BDNF pathway. Inhibition of CREB/BDNF pathway attenuated the effect of RK3. ConclusionIn conclusion, this study demonstrated that RK3 promotes learning and cognition in APP/PS1 and C57 mice by promoting neurogenesis and synaptogenesis through the CREB/BDNF signaling pathway. Therefore, RK3 is expected to be further developed into a potential drug candidate for the treatment of Alzheimer's disease (AD).

Anti-prostate cancer components of black ginseng based on the "nine steaming and nine sun-drying" process

This study was to identify the main active components of black ginseng responsible for prostate cancer treatment. HPLC fingerprints of 10 batches of processed ginseng from the "nine steaming and nine sun-drying" procedure were established. MTT test was performed to evaluate the inhibitory activity of black ginseng on human prostate cancer (DU145) cells. The spectrum-effect relationship analysis was assessed by correlation coefficient and grey relational analysis, and the active ginsenosides were further validated by bioactivity assay. Results indicated the anti-prostate cancer activity of black ginseng increased until the 7th cycle times of steaming and sun-drying, and the primary anti-prostate cancer ingredients of black ginseng were disclosed to be ginsenosides Rg5, F4, Rh4, Rk1, R-Rg3, and S-Rg3, of which Rk3, Rg5, and F4 exerted the highest effects. It is for the first time that a spectrum-effect relationship based on the ingredient changes of the "Nine Steaming and Nine Sun-drying" process was used to identify the active anticancer constituents of ginseng systemically and could further the stronger anticancer activity of black ginseng as well as provide a theoretical basis for the quality evaluation of black ginseng.

Rational design of supramolecular self-assembly sensor for living cell imaging of HDAC1 and its application in high-throughput screening

Supramolecular chemistry offers new insights in bioimaging, but specific tracking of enzyme in living cells via supramolecular host-guest reporter pair remains challenging, largely due to the interference caused by the complex cellular environment on the binding between analytes and hosts. Here, by exploiting the principle of supramolecular tandem assay (STA) and the classic host-guest reporter pair (p-sulfonatocalix[4]arene (SC4A) and lucigenin (LCG)) and rationally designing artificial peptide library to screen sequence with high affinity of the target enzyme, we developed a “turn-on” fluorescent sensing system for intracellular imaging of histone deacetylase 1 (HDAC1), which is a potential therapeutic target for various diseases, including cancer, neurological, and cardiovascular diseases. Based on computational simulations and experimental validations, we verified that the deacetylated peptide by HDAC1 competed LCG, freeing it from the SC4A causing fluorescence increase. Enzyme kinetics experiments were further conducted to prove that this assay could detect HDAC1 specifically with high sensitivity (the LOD value is 0.015 μg/mL, ten times lower than the published method). This system was further applied for high-throughput screening of HDAC1 inhibitors over a natural compound library containing 147 compounds, resulting in the identification of a novel HDAC1 down-regulator (Ginsenoside RK3). Our results demonstrated the sensitivity and robustness of the assay system towards HDAC1. It should serve as a valuable tool for biochemical studies and drug screening.

Ginsenoside Rk3 modulates gut microbiota and regulates immune response of group 3 innate lymphoid cells to against colorectal tumorigenesis

The gut microbiota plays a pivotal role in the immunomodulatory and protumorigenic microenvironment of colorectal cancer (CRC). However, the effect of ginsenoside Rk3 (Rk3) on CRC and gut microbiota remains unclear. Therefore, the purpose of this study is to explore the potential effect of Rk3 on CRC from the perspective of gut microbiota and immune regulation. Our results reveal that treatment with Rk3 significantly suppresses the formation of colon tumors, repairs intestinal barrier damage, and regulates the gut microbiota imbalance caused by CRC, including enrichment of probiotics such as Akkermansia muciniphila and Barnesiella intestinihominis, and clearance of pathogenic Desulfovibrio. Subsequent metabolomics data demonstrate that Rk3 can modulate the metabolism of amino acids and bile acids, particularly by upregulating glutamine, which has the potential to regulate the immune response. Furthermore, we elucidate the regulatory effects of Rk3 on chemokines and inflammatory factors associated with group 3 innate lymphoid cells (ILC3s) and T helper 17 (Th17) signaling pathways, which inhibits the hyperactivation of the Janus kinase-signal transducer and activator of transcription 3 (JAK-STAT3) signaling pathway. These results indicate that Rk3 modulates gut microbiota, regulates ILC3s immune response, and inhibits the JAK-STAT3 signaling pathway to suppress the development of colon tumors. More importantly, the results of fecal microbiota transplantation suggest that the inhibitory effect of Rk3 on colon tumors and its regulation of ILC3 immune responses are mediated by the gut microbiota. In summary, these findings emphasize that Rk3 can be utilized as a regulator of the gut microbiota for the prevention and treatment of CRC.

Photoprotective Effects of Processed Ginseng Leaf Administration against UVB-Induced Skin Damage in Hairless Mice.

Although ginseng leaves contain a larger amount of ginsenosides than the roots, studies on the protective effect of oral administration of ginseng leaves against photoaging are lacking. Processed ginseng leaves (PGL) prepared by acid reaction to increase effective ginsenoside content showed higher levels of Rg3 (29.35 mg/g) and Rk1 (35.16 mg/g) than ginseng leaves (Rg3 (2.14 mg/g) and Rk1 (ND)), and ginsenosides Rg3 and Rk1 were evaluated as active ingredients that protected human keratinocytes against UVB-induced cell damage by increasing cell proliferation and decreasing matrix metalloproteinase (MMP)-2 and 9 secretion. Herein, the effect of oral PGL administration (50, 100, or 200 mg/kg, daily) against photoaging in HR-1 hairless mice was assessed by measuring wrinkle depth, epidermal thickness, and trans-epidermal water loss for 16 weeks. The PGL treatment group showed reduced skin wrinkles, inhibited MMP-2 and MMP-9 expression, and decreased IL-6 and cyclooxygenase-2 levels. These data suggest that oral PGL administration inhibits photoaging by inhibiting the expression of MMPs, which degrade collagen, and inhibiting cytokines, which induce inflammatory responses. These results reveal that ginseng leaves processed by acid reaction may serve as potential functional materials with anti-photoaging activities.

An innovative processing driven efficient transformation of rare ginsenosides enhances anti-platelet aggregation potency of notoginseng by integrated analyses of processing-(chemical) profiling-pharmacodynamics

Ethnopharmacological relevancePanax notoginseng (Burk.) F. H. Chen, a valuable Chinese herb medicine, shows a characteristic bi-directional regulation of hemostasis and activating blood circulation with ginsenosides as the predominant bioactive compounds and is a typical representative of “processing triggered heteropotency”. Aim of the studyProcessing triggered heteropotency, one of the unique theories and practices in traditional Chinese medicine, refers to that the processing will lead to change in physical and chemical properties, and eventually disparate efficacy of the crude drugs, yet the optimum process and underlying mechanism remains unclear. In this study, using Panax notoginseng (PN) as a representative sample, a processing-(chemical) profiling-pharmacodynamics (3-P) relationship was proposed to investigate the processing mechanism of PN. Materials and methodsFirstly, a temperature programmed steaming process was designed to evaluate the steaming triggered chemical transformation of triterpene saponins and the corresponding enhancement in anti-platelet aggregation activity. The steaming process was programed from the conventional 100 °C–150 °C in a time course of 0–12 h, aiming to achieve the maximized conversion of rare ginsenosides (RGs), and dynamic profile of ginsenosides were constructed by a UPLC-Q-TOF-MS/MS analysis. Then, a processing-(chemical) profiling-pharmacodynamics (3-P) relationship was assessed by using the grey relational analysis (GRA) and orthogonal projections to latent structures (OPLS), and validated by bioactive fraction of 140 °C steamed PN. Subsequently, the P2Y12-ligand binding affinity of potential candidates was analyzed by molecular docking. Finally, the dynamic changes of ginsenosides during steaming of SPN were quantitatively detected by UPLC-QQQ-MS/MS. ResultsA total of 48 differential ginsenosides were characterized and monitored including the primary and secondarily transformed saponins. The higher temperature steaming especially at 140 °C induces not only the predominant production of the RGs, but also the stronger anti-platelet aggregation activity. The 3-P relationship showed the fraction (3) of 140 °C steamed PN rich in RGs exhibits the most predominant efficacy, in which, a series of RGs including ginsenosides Rg5, Rk1, 20(S/R)-Rg3 were proven to be potent components. Molecular docking analysis suggested that ginsenosides Rg5 and Rk1 showed more strong interaction with the platelet P2Y12 receptor. Quantitative analysis found 140 °C-2h PN possessed highest contents of Rk1 and Rg5 and total RGs. ConclusionsThe integrated 3-P strategy uncovered the promising ginsenosides with anti-platelet effect, thereby revealing the material basis of PN steaming, which could provide a new enlightenment for the investigation of processing mechanism of traditional Chinese medicines.

The Content and Principle of the Rare Ginsenosides Produced from Gynostemma pentaphyllum after Heat Treatment.

Ginsenoside Rg3, Rk1, and Rg5, rare ginsenosides from Panax ginseng, have many pharmacological effects, which have attracted extensive attention. They can be obtained through the heat treatment of Gynostemma pentaphyllum. In this study, scanning electron microscopy (SEM) and thermal gravity-differential thermal gravity (TG-DTG) were employed to investigate this process and the content change in ginsenosides was analyzed using liquid chromatography-mass spectrometry (LC-MS). SEM and TG-DTG were used to compare the changes in the ginsenosides before and after treatment. In SEM, the presence of hydrogen bond rearrangement was indicated by the observed deformation of vascular bundles and ducts. The before-and-after changes in the peak patterns and peaks values in TG-DTG indicated that the content of different kinds of compounds produced changes, which all revealed that the formation of new saponins before and after the heat treatment was due to the breakage or rearrangement of chemical bonds. Additionally, the deformation of vascular bundles and vessels indicated the presence of hydrogen bond rearrangement. The glycosidic bond at the 20 positions could be cleaved by ginsenoside Rb3 to form ginsenoside Rd, which, in turn, gave rise to ginsenoside Rg3(S) and Rg3(R). They were further dehydrated to form ginsenoside Rk1 and Rg5. This transformation process occurs in a weak acidic environment provided by G. pentaphyllum itself, without the involvement of endogenous enzymes. In addition, the LC-MS analysis results showed that the content of ginsenoside Rb3 decreased from 2.25 mg/g to 1.80 mg/g, while the contents of ginsenoside Rk1 and Rg5 increased from 0.08 and 0.01 mg/g to 3.36 and 3.35 mg/g, respectively. Ginsenoside Rg3(S) and Rg3(R) were almost not detected in G. pentaphyllum, and the contents of them increased to 0.035 and 0.23 mg/g after heat treatment. Therefore, the rare ginsenosides Rg3(S), Rg3(R), Rk1, and Rg5 can be obtained from G. pentaphyllum via heat treatment.

Ginsenoside Rk3 Regulates Nonalcoholic Steatohepatitis by Modulation of Intestinal Flora and the PI3K/AKT Signaling Pathway in C57BL/6 Mice.

Non-alcoholic steatohepatitis (NASH) has become the most important reason of liver disease around the world and is predisposed to further progression to cirrhosis and hepatocellular carcinoma. Ginsenoside Rk3 has been reported to have a plenty of biological activities, including anti-apoptotic, anti-anemia, and protective effects against acute kidney injury. However, whether ginsenoside Rk3 can improve NASH has not been reported yet. Therefore, the purpose of this study is to investigate the protective effect of ginsenoside Rk3 against NASH and its mechanism of action. C57BL/6 mice were treated with different dosages of ginsenoside Rk3 after being established as a NASH model. Our results showed that Rk3 administration significantly improved liver inflammation, lipid deposition, and fibrosis caused by a high-fat-high-cholesterol (HFHC) diet and CCl4 injection in mice. Notably, ginsenoside Rk3 was discovered significantly to inhibit the PI3K/AKT signaling pathway. Additionally, treatment with ginsenoside Rk3 remarkably amended the abundance of short-chain fatty acids. These changes were associated with beneficial variations to the variety and composition of the intestinal microbiota. In conclusion, ginsenoside Rk3 ameliorates hepatic non-alcoholic lipid inflammation and triggers changes in the beneficial intestinal flora, helping to reveal host-microbe interactions. The outcomes of this study indicate that ginsenoside Rk3 is a promising drug candidate for the treatment of NASH.

Ginsenoside Rk3 suppresses U46619-induced human platelets aggregation through regulation of cAMP and PI3K/Akt pathway

Ginsenoside Rk3 suppresses U46619-induced human platelets aggregation through regulation of cAMP and PI3K/Akt pathway

Structural-Activity Relationship of Rare Ginsenosides from Red Ginseng in the Treatment of Alzheimer's Disease.

Rare ginsenosides are the major components of red ginseng. However, there has been little research into the relationship between the structure of ginsenosides and their anti-inflammatory activity. In this work, BV-2 cells induced by lipopolysaccharide (LPS) or nigericin, the anti-inflammatory activity of eight rare ginsenosides, and the target proteins expression of AD were compared. In addition, the Morris water maze test, HE staining, thioflavins staining, and urine metabonomics were used to evaluate the effect of Rh4 on AD mice. Our results showed that their configuration influences the anti-inflammatory activity of ginsenosides. Ginsenosides Rk1, Rg5, Rk3, and Rh4 have significant anti-inflammatory activity compared to ginsenosides S-Rh1, R-Rh1, S-Rg3, and R-Rg3. Ginsenosides S-Rh1 and S-Rg3 have more pronounced anti-inflammatory activity than ginsenosides R-Rh1 and R-Rg3, respectively. Furthermore, the two pairs of stereoisomeric ginsenosides can significantly reduce the level of NLRP3, caspase-1, and ASC in BV-2 cells. Interestingly, Rh4 can improve the learning ability of AD mice, improve cognitive impairment, reduce hippocampal neuronal apoptosis and Aβ deposition, and regulate AD-related pathways such as the tricarboxylic acid cycle and the sphingolipid metabolism. Our findings conclude that rare ginsenosides with a double bond have more anti-inflammatory activity than those without, and 20(S)-ginsenosides have more excellent anti-inflammatory activity than 20(R)-ginsenosides.

Anti-thrombotic effects of ginsenoside Rk3 by regulating cAMP and PI3K/MAPK pathway on human platelets

Background and objectiveThe ability to inhibit aggregation has been demonstrated with synthetically derived ginsenoside compounds G-Rp (1, 3, and 4) and ginsenosides naturally found in Panax ginseng 20(S)-Rg3, Rg6, F4, and Ro. Among these compounds, Rk3 (G-Rk3) from Panax ginseng needs to be further explored in order to reveal the mechanisms of action during inhibition. MethodologyOur study focused to investigate the action of G-Rk3 on agonist-stimulated human platelet aggregation, inhibition of platelet signaling molecules such as fibrinogen binding with integrin αIIbβ3 using flow cytometry, intracellular calcium mobilization, dense granule secretion, and thromboxane B2 secretion. In addition, we checked the regulation of phosphorylation on PI3K/MAPK pathway, and thrombin-induced clot retraction was also observed in platelets rich plasma. Key ResultsG-Rk3 significantly increased amounts of cyclic adenosine monophosphate (cAMP) and led to significant phosphorylation of cAMP-dependent kinase substrates vasodilator-stimulated phosphoprotein (VASP) and inositol 1,4,5-trisphosphate receptor (IP3R). In the presence of G-Rk3, dense tubular system Ca2+ was inhibited, and platelet activity was lowered by inactivating the integrin αIIb/β3 and reducing the binding of fibrinogen. Furthermore, the effect of G-Rk3 extended to the inhibition of MAPK and PI3K/Akt phosphorylation resulting in the reduced secretion of intracellular granules and reduced production of TXA2. Lastly, G-Rk3 inhibited platelet aggregation and thrombus formation via fibrin clot. Conclusions and implicationsThese results suggest that when dealing with cardiovascular diseases brought upon by faulty aggregation among platelets or through the formation of a thrombus, the G-Rk3 compound can play a role as an effective prophylactic or therapeutic agent.

Ginsenoside Rk3 is a novel PI3K/AKT-targeting therapeutics agent that regulates autophagy and apoptosis in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is the third leading cause of cancer death worldwide. Ginsenoside Rk3, an important and rare saponin in heat-treated ginseng, is generated from Rg1 and has a smaller molecular weight. However, the anti-HCC efficacy and mechanisms of ginsenoside Rk3 have not yet been characterized. Here, we investigated the mechanism by which ginsenoside Rk3, a tetracyclic triterpenoid rare ginsenoside, inhibits the growth of HCC. We first explored the possible potential targets of Rk3 through network pharmacology. Both in vitro (HepG2 and HCC-LM3 cells) and in vivo (primary liver cancer mice and HCC-LM3 subcutaneous tumor-bearing mice) studies revealed that Rk3 significantly inhibits the proliferation of HCC. Meanwhile, Rk3 blocked the cell cycle in HCC at the G1 phase and induced autophagy and apoptosis in HCC. Further proteomics and siRNA experiments showed that Rk3 regulates the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway to inhibit HCC growth, which was validated by molecular docking and surface plasmon resonance. In conclusion, we report the discovery that ginsenoside Rk3 binds to PI3K/AKT and promotes autophagy and apoptosis in HCC. Our data strongly support the translation of ginsenoside Rk3 into novel PI3K/AKT-targeting therapeutics for HCC treatment with low toxic side effects.

Antiviral Effect of Ginsenosides rk1 against Influenza a Virus Infection by Targeting the Hemagglutinin 1-Mediated Virus Attachment

Influenza A virus (IAV) infections have been a serious hazard to public health everywhere. With the growing concern of drug-resistant IAV strains, there is an urgent need for novel anti-IAV medications, especially those with alternative mechanisms of action. Hemagglutinin (HA), an IAV glycoprotein, plays critical roles in the early stage of virus infection, including receptor binding and membrane fusion, making it a good target for developing anti-IAV drugs. Panax ginseng is a widely used herb in traditional medicine with extensive biological effects in various disease models, and its extract was reported to show protection in IAV-infected mice. However, the main effective anti-IAV constituents in panax ginseng remain unclear. Here, we report that ginsenoside rk1 (G-rk1) and G-rg5, out of the 23 screened ginsenosides, exhibit significant antiviral effects against 3 different IAV subtypes (H1N1, H5N1, and H3N2) in vitro. Mechanistically, G-rk1 blocked IAV binding to sialic acid in a hemagglutination inhibition (HAI) assay and an indirect ELISA assay; more importantly, we showed that G-rk1 interacted with HA1 in a dose-dependent manner in a surface plasmon resonance (SPR) analysis. Furthermore, G-rk1 treatment by intranasal inoculation effectively reduced the weight loss and mortality of mice challenged with a lethal dose of influenza virus A/Puerto Rico/8/34 (PR8). In conclusion, our findings reveal for the first time that G-rk1 possesses potent anti-IAV effects in vitro and in vivo. We have also identified and characterized with a direct binding assay a novel ginseng-derived IAV HA1 inhibitor for the first time, which could present potential approaches to prevent and treat IAV infections.