Abstract
The term ginseng refers to the dried roots of several plants belonging to the genus Panax of the Araliaceae family. The 3 major commercial ginsengs are Panax notoginseng (Burk.) F.H. Chen (Notoginseng), P. ginseng C.A. Meyer (Ginseng), and P. quinquefolius L. (American ginseng), which have been used as herbal medicines. Over 18,000 papers on ginsengs have been published on the basis of their structural diversity and biological activities. Many reviews have summarized the phytochemistry, pharmacology, and clinical use of ginsengs, but the structure-activity relationship (SAR) of ginsenosides from ginsengs in autophagy is unavailable. Herein, we review the structural diversity of ginsenosides, especially the ones in notoginseng, and the SAR in autophagic activity is discussed in detail.
Panax notoginseng (Burk.) F.H. Chen (notoginseng), P. ginseng C.A. Meyer (Asian ginseng), and P. quinquefolius L. (American ginseng) are the 3 major commercial ginsengs worldwide. Among them, P. notoginseng is mainly cultivated in southwest China. Panax ginseng is widespread in northeast China, Korea, and Japan, whereas P. quinquefolius is native to south Canada and northern United States, but is nowadays cultivated in northeast China, Korea, and Japan. These 3 major commercial ginsengs show different drug properties according to the traditional Chinese medicine theory. American ginseng is adept at nourishing Yin, and ginseng can tonify Yang, while notoginseng is quite mild.
The emergence of complex diseases is promoting a change from one-target to multi-target drugs and traditional Chinese herbs are ideal models. 1 Panax notoginseng, P. ginseng, and P. quinquefolius are the best selected and refined herbs used to treat some complex diseases. Dammarane-type ginsenosides are the main biological ingredients of the 3 commercial ginsengs. However, ocotillol (OCT)- and oleanane-type oligoglycosides, saponins in ginseng and American ginseng, respectively, have not been reported in notoginseng. The dammarane-type saponins from ginsengs are classified into protopanaxadiol (PPD) and protopanaxatriol (PPT) types, as well as C-17 side-chain varied types. PPD and PPT are transformed into C-17 side-chain compounds such as ginsenosides Rg3, Rg5, Rk1, Rh1, Rh4, and Rk3 after steaming, heating, or microwave processing. 2 Pharmacological research proved that these 3 major ginsengs may adjust autophagy and play a role in various disease treatments such as cancer, neurodegeneration, autoimmune diseases and liver disorders. 3 -6
Autophagy is an evolutionarily conserved catabolic process whereby the cytoplasmic contents of a cell are sequestered within autophagosomes through a lysosome-dependent pathway. 7 Increasing evidence shows that this process is of great importance in a wide range of complex diseases, including neurodegenerative disorders, hepatic fibrosis, and aging autoimmune diseases. 8 -10
Some publications previously reported that ginsenosides in ginsengs played a role in various disease treatments by adjusting autophagy. Herein, we review the data reported in the literature from 1960 to date, paying attention to the structural diversity of ginsenosides from ginsengs, especially concerning their structure-activity relationship (SAR) in autophagy. The results showed that although similar in structures, different types of ginsenosides showed distinct effects on autophagy. Ginsenoside Rb1, a PPD-type, should be an autophagy inhibitor, while ginsenoside Rg1, a PPT-type, should be an autophagy promoter. Previous studies may support the view that ginsengs show different drug properties due to their different effects on autophagy.
Moreover, most of the transformed saponins bearing lower numbers of sugar moieties could promote autophagy. They showed stronger anticancer activities in comparison with their prototype compounds. However, since the biological data of some ginsenosides are not available and some are either not consistent or even contradictory, further studies on autophagy of ginsenosides are necessary. It would be sensible to modify ginsenosides to produce new drugs targeting autophagy.
Structural Diversity of Ginsenosides
Ginsenosides are responsible for diverse pharmacological properties ascribed to ginsengs, which have attracted considerable attention. 11 To date, more than 280 saponins have been isolated and identified from different parts of ginsengs, including the root, rhizome, stem, leaf, flower, flower bud, fruit, seed, and flower pedicel. 12 -20 Some transformed saponins were isolated from the steamed root or leaf of ginsengs. 21 -25 In general, the ginsenosides are classified into dammarane, oleanane, and OCT types (Figure 1). Dammarane-type compounds with either a PPD or PPT aglycon are the main chemical constituents of the 3 ginsengs (Figure 1). However, the OCT type is most common in P. quinquefolius, whereas the oleanane type is most common in P. ginseng. Few OCT and oleanane types have been found in P. notoginseng. Furthermore, the diverse ginsenosides vary in the type and number of monosaccharide residues, as well as their mutual linkages. Stereoisomerism and C-17 side-chain variation in the saponins also contribute to the structural diversity of the ginsenosides (Figure 2). However, the ring system of the ginsenosides is very stable. To date, no ginsenoside with a ring rearrangement has been reported.
Comparative HPLC profiles of the 3 commercial ginsengs.
Skeletons and structures of main ginsenosides isolated from ginsengs (PPD, protopanaxadiol; PPT, protopanaxatriol).
Based on the aforementioned structural differentiation of the ginsenosides, the dammarane-type can be divided into 3 groups: PPD, PPT, and C-17 side-chain varied types (Figure 2). The PPD and PPT types have hydroxyl groups attached to C-3, C-12, and C-20. Compared with PPD, the PPT type has an extra hydroxyl group at C-6. Furthermore, PPD has sugar units linked to either C-3 or C-20, while PPT has saccharides attached to either C-6 or C-20. Glucose (Glc), rhamnose (Rha), xylose (Xyl), and arabinose (pyran and furan-Ara) are the 5 major monosaccharides found in the ginsenosides. Moreover, many of the saponins from ginsengs possess acylated sugar moieties, especially those from Asian ginseng.
The contents of ginsenosides in the roots and other parts of 3 ginsengs are totally different. 26 The content of total saponins of notoginseng (8%) is higher than that of Asian ginseng (6%) and American ginseng (4%) (Figure 3). Usually, the 5 main ginsenosides, notoginsenoside R1 and Rb1, Rg1, Rd, and Re, make up more than 90% of the total ginsenosides content in notoginseng.
Transformed ginsenosides in steamed ginsengs and possible transformation rules.
A Diversity of Ginsenosides After Steaming Ginsengs
Both the raw and processed forms of ginsengs have been used traditionally.
The raw ginsengs are prepared by air-drying after harvest. The processed notoginseng prepared by either steaming or heating is called steamed notoginseng. Asian ginseng prepared in the same way is named as red ginseng. However, few steamed American ginsengs have been reported. Traditionally, raw notoginseng is used to invigorate the circulation of blood as a hemostatic, traumatic, and cardiovascular medicine. Steamed notoginseng, as a tonic medicine, is used to nourish the blood and increase the production of various blood cells in anemic conditions. Raw, red, and black Asian ginsengs showed different biological activities. Due to the obvious degradation and oxidation of prototype saponins during the steaming process, the ginsenosides of steamed ginsengs are totally different from those of the untreated ones. It is reported that after 8 hours steaming at 120°C, the content of 5 major saponins of notoginseng, notoginsenoside R1 and ginsenosides Rg1, Re, Rb1, and Rd, decreased considerably. The content of less polar ginsenosides increased, for example, ginsenosides 20(R/S)-Rh1, Rk3, Rh4, 20(S/R)-Rg3, Rk1, and Rg5. 27 Those 8 saponins are either minor or trace components of raw notoginseng. Moreover, many acetylated ginsenosides (quinquenoside R1 and ginsenosides Rs1 and Rs2) are relatively abundant in red ginseng due to the inactivation of the deacetylating enzyme during steaming. Significant contents of the less polar ginsenosides (F4, F5, Rk1, Rg2, Rg5, Rg6, 20(R)-Rs3, 20(S)-Rs3, and Rs4) are observed in red ginseng. 11
The transformed ginsenosides can be produced via the hydrolysis of sugar moieties and subsequent dehydration or oxidation or cleavage of the side chain at C-17. The structural changes in ginsenosides after steaming are shown in Figure 3. To date, over 60 transformed saponins have been reported in steamed notoginseng. 21,28-30 Herein, we take transformed saponins of steamed notoginseng as an example to explain the transformation progress of ginsenosides during steaming. The C-20 sugar residue is easy to eliminate during steaming, while the C-3 sugar is difficult to hydrolyze. Consequently, ginsenosides Rb1, Rd, and (R/S) Rg3 are produced via the hydrolysis of the C-20 sugar unit. Rg3 is further dehydrated to produce 2 isomers of the double bond at either C-20/21 or C-20/22, namely Rk1 and Rg5, respectively. Accordingly, (R/S) Rh1 is transformed from the PPT group of ginsenosides (R1, Rg1, and Re). Rh1 is further dehydrated to 2 geometric isomers namely Rk3 and Rh4. Subsequently, the ginsenosides are transformed to degradation products due to the cyclization, dehydration, and oxidative cleavage of the side chain at C-17. It is noted that the epimerides of 20(S) and 20(R)-ginsenoside were produced after hydrolysis of the glycosyl unit at C-20. Moreover, the emergence of some positional isomers of the double bond at C-20/21 or C-20/22, cis/trans isomers of the double bond at C-20/22, and stereoisomers at 24(S, R) and 25(S, R) is attributable to the dehydration of the hydroxyl at C-20 and selective oxidation of the double bond. In conclusion, the diversity of ginsenosides in steamed ginsengs is mainly due to stereoisomerism.
The saponins found in steamed ginsengs displayed moderate potential for enhancing the neurite outgrowth of NGF-mediated PC12 cells and AChE inhibitory activities, properties not shown by the major compounds in the raw roots. 21,29,30 Furthermore, the major saponins, ginsenosides Rg3, Rk1, and Rg5, being rare in raw ginsengs, were identified as potential neurogenic and anticancer molecules. 31 It is suggested that many of the transformed compounds generated in steamed ginsengs have a structural diversity and biological activity in the central nervous system, as well as having anticancer properties.
Autophagy and Possible Mechanism
Autophagy
Autophagy is a regulated and evolutionarily conserved catabolic process by which damaged organelles or superfluous proteins are delivered to lysosomes for clearance. 32 The (dys)regulation of autophagy is implicated in the pathogenesis of various diseases such as cancer, 33 neurodegeneration, 8 autoimmune diseases, 34 and liver disorders. 35 Ginsenosides, as the major bioactive constituents of ginsengs, are used for the treatment of cardiovascular diseases and have anticancer, anti-inflammatory, and neuroprotective activities by regulating the autophagy signaling pathway (Table 1).
Ginsenosides Acting on Autophagy and Their Potential Structure-Activity Relationship in Autophagy.
PPD, protopanaxadiol; PPT, protopanaxatriol.
Action on Cardiovascular and Cerebral-Vascular Diseases
Autophagy, related to the occurrence of atherosclerosis and other cardiovascular diseases, might be an attractive target for new cardiovascular therapies. However, autophagy is a double-edged sword in cardiovascular disease, acting in either beneficial or maladaptive ways depending on the context. Ginsengs have been used to treat atherosclerosis and other cardiovascular diseases for a long time. 32 It was reported that ginsenoside Rb1 inhibited autophagy through regulation of Rho/ROCK and PI3K/mTOR pathways in a pressure-overload heart failure rat model. 36 On the contrary, ginsenoside Rg1 could promote autophagy and thus inhibit apoptosis via AMPK/mTOR signaling in serum deprivation macrophages. 52,55 Rg1 and Rb1 are 2 major compounds in ginsengs, but the ratios of Rg1/Rb1 are different among the 3 ginsengs. 26 The aforementioned data suggested that Rb1 should be an autophagy inhibitor and Rg1 an autophagy promoter. The 3 ginsengs showed different effects on autophagy due to their different ratios of Rg1/Rb1.
Anticancer Activities
Autophagy is regarded as a potential target for anticancer therapy due to its complicated functions in different stages of cancer. 56 Ginsenoside Rb1, one of the major metabolites of ginsengs, shows anticancer properties by inducing autophagic cell death and apoptosis via AMPK/JNK pathway activation. 57 Previous experimental results suggested that the transformed saponins in ginsengs showed better anticancer activities than the protosaponins. For example, both ginsenosides 20(S)-Rg3 and Rh2 are transformed saponins from ginsenoside Rb1 in steamed ginsengs. Ginsenoside 20(S)-Rg3 exhibited anticancer activity against various cancers through the induction of autophagy-mediated inhibition of cancer invasive progression. 40,58,59 Ginsenoside Rh2 inhibited hepatocellular carcinoma by influence on autophagy. 60 Ginsenoside Rk1 is an effective anticancer regimen in HepG2 cells due to autophagy inhibition. 46 Ginsenoside compound K (CK), identified as a major ginsenoside metabolite in urine and blood (Figure 4), induced autophagy and apoptosis via the generation of ROS and the activation of c-Jun NH2-terminal kinase in human colon cancer cells. 49 Recent results revealed a cooperative function of CK and TRAIL against colon cancer through autophagy-dependent and autophagy-independent (p53-CHOP pathway) DR5 upregulation. 61 KG-135, a standardized formulation enriched with transformed ginsenosides, including Rk1, Rg3, and Rg5, has shown obvious inhibition of various types of cancer cells by activating potentially extrinsic FOXO3a/FasL/caspase-8 and intrinsic caspase-9 apoptotic pathways by blockade of autophagy. 62 It is noted that ginsenoside F2, one major ingredient of the aerial part of notoginseng, showed antiproliferative activity against breast cancer stem cells (CSCs) by initiating an autophagic progression. 51
Metabolized products of ginsenosides and their possible transformation rules.
Neurodegenerative Disorders
It is interesting that saponins from raw and steamed ginsengs could act on neurodegenerative disorders through autophagy. Ginsenoside Rg1 is a specific promoter of autophagy and inhibitor of apoptosis 63 and is considered valuable for neuronal survival. For example, the ginsenoside Rg1 protected microglial cells against oxidative damage induced by 10 µM tert-Bu hydroperoxide through decreasing autophagic activation and apoptosis. 64 Ginsenoside Rg1 protected against sepsis-associated encephalopathy through beclin 1-independent autophagy in mice. 65 Moreover, the combination of astragaloside IV and Rg1 resisted autophagic injury in PC12 cells induced by oxygen glucose deprivation/reoxygenation associated with the PI3K I/Akt/mTOR and PI3K III/Becline-1/Bcl-2 signaling pathways. 66 Inhibition of autophagy is involved in the neuroprotective effects of ginsenoside Rb1 on spinal cord injury. 37 Ginsenoside Rb1 may also protect against neuronal death caused by ischemic insults by inhibition of autophagy via activation of the PI3K/Akt pathway. 58 Ginsenoside Rg3, a transformed saponin, could attenuate prion peptide (106-126)-induced neurotoxicity and mitochondrial damage via autophagy flux, which could be a possible therapeutic agent in neurodegenerative disorders, including prion diseases. 67 CK, the metabolite of ginsenoside of ginsengs, promotes β-amyloid peptide clearance in primary astrocytes via autophagy enhancement. 50 Moreover, CK may inhibit autophagy-mediated apoptosis through activation of the PI3K-Akt signaling pathway thus protecting against ischemia/reperfusion injury. 48 Furthermore, gypenoside XVII is a promising candidate for the prevention of AD due to facilitation of the autophagy-dependent removal of Aβ through TFEB activation in cellular and rodent models of AD. 68 Gypenoside XVII could also attenuate Aβ25-35-induced parallel autophagic and apoptotic cell death through the estrogen receptor-dependent activation of Nrf2/ARE pathways. 69
Renoprotection
Ginsenoside Rg1 is beneficial for alleviation of podocyte injury through inhibition of angiotensin II-induced autophagy via the AMPK/mTOR/PI3K pathway. 53 It could inhibit renal injury induced by aldosterone possibly through reducing the phosphorylation of AMPK and preserving mTOR activity. 70 Notoginsenoside R1, one of the typical saponins of notoginseng, could protect podocytes by activating the PI3K/Akt/mTOR pathway. 71
Other Disorders
Ginsenoside Rb2 could alleviate hepatic lipid accumulation by restoring autophagy via the induction of sirt1 and activation of AMPK, resulting in improved nonalcoholic fatty liver disease and glucose tolerance. 72
Structure-Activity Relationship of Ginsenosides From Notoginseng in Autophagy
Effects of Different Skeleton Types on Autophagy
To date, the ginsenosides of ginsengs are mainly divided into 3 skeleton types: dammarane, OCT, and oleanane. The dammarane type includes PPD, PPT, and C-17 side-chain varied types. It is reported that dammarane-type saponins affected autophagy, while the effects of OCT-type and oleanane-type oligoglycosides on autophagy were unavailable, except for pseudoginsenoside F11, a major OCT-type saponin in American ginseng. 54 Moreover, PPT-type saponins should be autophagy inhibitors, for example, Rb1. 36,37,39 However, the transformed PPT-type ginsenosides, such as Rg3 58 and Rh2, were autophagy promoters. 73 The PPD-type oligoglycosides were autophagy promoters, for example, Rg1 and its transformed ginsenoside, such as Rh4. 74 It is a regret that few publications have reported the C-17 side-chain varied type saponins with an effect on autophagy, although some of them showed promising biological activity. 29 An SAR analysis of C-17 side-chain varied type saponins is unavailable.
Stereoselectivity of 20(S) and 20(R)
The transformed dammarane-type saponins from the steamed ginsengs feature C-20 epimers. It was reported that ginsenosides Rg3 and Rh2, the major components of red ginseng and steamed notoginseng, induced apoptotic cell death in a stereoisomer-specific fashion. The ginsenosides 20(S) Rg3 and Rh2 may promote cell death in a dose-dependent manner due to downregulation of Bcl2 and upregulation of Fas gene expression, resulting in apoptosis of HepG2 cells with poly adenosine diphosphate ribose polymerase cleavage. The results suggested that the 20(S) forms of Rg3 and Rh2 possess more potent anticancer activities with autophagy than their 20(R) forms. 42 It is regretful that studies on the effect of other stereoisomers on autophagy are unavailable.
Effects of Sugar Chain on Autophagy
Some ginsenosides and their metabolites affected autophagy. Either the metabolites of ginsenosides or transformed saponins by heating or steaming showed more potent anticancer activity in autophagy than their prototype. For example, ginsenoside F2 (F2) and ginseng compound K (CK) are metabolites of ginsenoside Rb1 (Figure 4). Both of them showed better antitumor activity than that of ginsenoside Rb1. 61 Ginsenoside F2 (F2), transformed by the human intestinal bacterial enzyme after oral ingestion of ginsenoside Rb1, induced protective autophagy and apoptotic cell death in breast CSCs through the upregulation of p53. 51,75 Ginseng compound K (CK), a major metabolite of several PPD-type ginsenosides, for example, Rb1, Rb2, and Rc, acts as a potent tumor suppressor of human melanoma cells by inducing autophagic cell death and apoptosis. 57,76,77 The transformed saponins of steamed ginsengs also showed promising anticancer activity, such as ginsenosides Rg3, 40,58 Rh2, 42,78 Rh4, Rg5, 79 and Rk1. 47 Among them, ginsenosides Rg3, Rh2, Rg5, and Rk1 were transformed from Rb1, while Rh4, Rh1, and Rk3 were produced from Rg1. 47 Compared with their prototype compounds, the transformed saponins had less sugar moieties and showed potential anticancer activity. The results suggested that the sugar chain should be the key pharmacophore of ginsenosides against cancer cells.
Conclusion
Herein, we reviewed the ginsenosides of ginsengs and discussed their potential SAR in autophagy. Although different ginsengs have similar ginsenosides, the ratios of the major ginsenosides are different. Most transformed ginsenosides with chemical diversity at the C-17 side chain were reported from steamed ginsengs. However, chemical studies on other parts of ginsengs, except for the root, are inadequate. In addition, some analytical data published were not self-consistent and some chemical constituents were not identified carefully. It is important to make further investigation and comparative analysis of the different parts of ginsengs, especially those of notoginseng.
Based on previous publications, our SAR analysis suggested that the skeleton type, stereo configuration of C-20, and the sugar chain of the saponins are related to autophagic activity. Rb1, PPD-type saponins of ginsengs, should be autophagy inhibitors, while Rg1, PPT-type saponins, could be autophagy promoters. The 20(S)-ginsenosides possess more potent anticancer activity in autophagy than their respective 20(R)-forms. With a decrease in the number of sugar moieties, most transformed saponins as autophagy promoters showed more potential anticancer activities than the prototype compounds. The effects of C-17 side-chain varied compounds on autophagy are unavailable, as well as those of OCT and oleanane saponins. Moreover, the aforementioned data suggested that most of the diverse ginsenoside derivatives were isolated from steamed ginsengs. However, some autophagic activity of the transformed ginsenosides is inconsistent, unavailable, or self-contradictory. It is imperative to make further efforts to study the effect of ginsenoside derivatives on autophagy in order to optimize the SAR. Moreover, although many publications reported on the mechanisms of most ginsenosides, the target proteins of ginsenosides are still not clear. It is urgent to investigate the target proteins of the ginsenosides from ginsengs.
Footnotes
Acknowledgments
The authors are grateful to Dr San-Jun Zhao of the School of Life Sciences, Yunnan Normal University, for the manuscript revision. This work was supported by 2016BC013 (project from Yunnan Provincial Science and Technology Department) and program for innovative research team (in science and technology) in universities of Yunnan Province.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The author(s) declared no financial support for the research, authorship, and/or publication of this article.