Therapeutic properties of Gynostemma pentaphyllum (jiaogulan)

A literature review

Aug 14, 2022

Last update:

24th August ‘22. (Jiaogulan tea, what does it taste like?)

Any extracts used in the following article are for non commercial research and educational purposes only and may be subject to copyright from their respective owners.

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Abstract

This Substack is a scientific literature review of research into the therapeutic properties of Gynostemma pentaphyllum, also known as the immortality herb, five-leaf ginseng, poor man's ginseng, miracle grass, fairy herb, sweet tea vine, gospel herb, and southern ginseng. A PubMed 10 year search for “Gynostemma pentaphyllum” returned 298 results. Some of the more cited research is presented here for review:

From a paper published in 2021, Rehan and Shafiullah conducted an in silico molecular docking binding analysis of 60 saponins with the COVID-19 main protease 6LU7 Mpro.

Although they found that 34 saponins were more effective than hydroxychloroquine, chloroquine or nelfinavir, as this was software based analysis the results need confirming in vitro and in vivo, but it is extremely promising research.

Okoye et al (2012) investigated the antiviral effects of extracts of G. pentaphyllum against yellow fever virus infectivity of chicken egg embryos and mice, and the percentage inhibition of viral induced hemagglutination (ie red blood cells clumping together). Results were somewhat encouraging, especially as even today there are still no specific anti-viral drugs to treat yellow fever.

From the same year, 2012, the same author Okoye worked with Nworu to study “Inhibition of HIV-1 lentiviral particles infectivity by Gynostemma pentaphyllum extracts in a viral vector- based assay”. As with the previous study they performed solvent extraction from leaf powder using ethyl ether (EG), methanol (MG) or water (AG).

Again the results from this in vitro research pointed to potential antiviral drugs that could be developed using extracts from the herb.

Sornpet et al (2017) investigated the antiviral activity of five Asian medicinal plant crude extracts against H5N1 avian influenza virus.

In 2020, Shaito et al published a comprehensive review of the ethnopharmacological therapeutic potentials and medicinal properties against cardiovascular diseases (CVDS) of four widely used plants: Ginseng, Ginkgo biloba, Ganoderma lucidum, and Gynostemma pentaphyllum.

This review is unusual, so far, in that it considers available clinical trials data as well as safety, toxicity and side effects.

(2012), Li et al extracted 2 acidic polysaccharides and tested against cancer cells in vivo and in vitro. After using alcohol to remove lipid they used water at 90°C for 2 hours, x 3, to extract from dried stem and leaf material.

Cancer types: Human chronic myeloid leukemia K-562, breast adenocarcinoma MCF-7, colon adenocarcinoma HT-29, hepatocellular carcinoma HepG2 and mouse melanoma B16 cell lines.

For in vivo studies they used tumor bearing mice and melanoma injected male rats, feeding them or injecting the polysaccharides and various statistically significant results are presented.

In 2010, Peng, Zhou and Zhang investigated the antitumor activities of dammarane triterpene saponins from a different species, Bacopa monniera.

I reference this study due to the commonality of the dammaranes with G. pentaphyllum.

This herb is also known as water hyssop, waterhyssop, brahmi, thyme-leafed gratiola, herb of grace, and Indian pennywort. Their in vivo studies in mice demonstrated tumor inhibition of up to 90%.

Moving on to 2016 and Li et al performed an in vitro study into the anticancer activity of a nonpolar fraction from G. pentaphyllum.

They provided evidence for significant anticancer activities from previously unreported, non-dammarane compounds. This provides further evidence of broad spectrum anti-tumor efficacy and the potential for developing novel anticancer agents.

Also from 2016, Li et al published a comprehensive review of the literature associated with G. pentaphyllum (GpM) and anti-cancer activities and mechanisms of action.

The most recent paper in this review of anti-cancer properties is from 2021, by Liu et al. They investigated how gypenosides of G. pentaphyllum can induce apoptosis of renal cancer tells through decreasing the phosphorylation level of Akt and mTOR in the PI3K/Akt/mTOR signaling pathway.

In 2010, Choi et al reported neuroprotective effects of ethanol extracts from G. pentaphyllum in a rat model of Parkinson’s disease.

And in 2018, Dong et al published research into how gypenosides reverse depressive behaviour by inhibiting hippocampal neuroinflammation.

Then in 2022, Wang et al conducted an in silico analysis to screen for compounds that can inhibit neurologically damaging signalling pathways associated with Alzeimer’s disease (AD).

From 2020, Wang et al conducted an in vitro investigation in to how dammarane-Type saponins from G. pentaphyllum can prevent hypoxia-induced neural injury through activation of ERK, Akt, and CREB pathways.

Hong et al (2018) conducted a biomedical investigation integrated with an in silico assay into how G. pentaphyllum can attenuate the progression of nonalcoholic liver disease in mice.

In 2017, Wong et al demonstrated how G. pentaphyllum saponins attenuate inflammation in vitro and in vivo by inhibition of NF-κB and STAT3 signaling. This is also another tumorigenic pathway and one of the key pathways mediated by cellular exposure to spike protein due to COVID-19 infection or transfection.

Back in 2006, Megalli, Davies and Roufogalis conducted an in vivo investigation using rats into how extracts of G. pentaphyllum can improve the serum ratios of cholesterols, triglyceride, blood sugar and the insulin resistance profile.

In 2020, Yin et al reported on ten new dammarane-type saponins with hypolipidemia activity from G. pentaphyllum herbal tea.

Huang et al (2022) published a review focusing on the prebiotic and therapeutic aspects of saponins and polysaccharides of jiaogulan tea, and the indirect anticancer effects of a healthy gut biome.

Bioavailability, contraindications, interactions with chemotherapeutic drugs and dosage recommendations are then considered, to conclude the review.

Discussion

Firstly, a hat tip to the many insightful comments by “The Post-Covid Pharmacist” who unfortunately doesn’t post but suggested a therapeutic containing extracts of G. pentaphyllum which ultimately led to this literature review.1

Gynostemma pentaphyllum, or jiaogulan is a dioecious, herbaceous climbing vine of the family Cucurbitaceae (cucumber or gourd family) widely distributed in South and East Asia as well as New Guinea. Its fruit is a small purple inedible gourd. It is a climbing vine, attaching itself to supports using tendrils. The serrated leaflets commonly grow in groups of five (as in G. pentaphyllum) although some species can have groups of three or seven leaflets. The plant is dioecious, meaning each plant exists either as male or female. Therefore, if seeds are desired, both a male and female plant must be grown.

G. pentaphyllum is one of about 17 species in the genus Gynostemma, including nine species endemic to China. However, G. pentaphyllum has a wide distribution outside of China, ranging from India and Bangladesh to Southeast Asia to Japan and Korea as well as to New Guinea. In China, it grows in forests, thickets, and roadsides on mountain slopes at elevations of 300–3,200 m (980–10,500 ft) above sea level.

Jiaogulan is a vine hardy to USDA zones 8-11 (frost tender) in which it may grow as a short lived perennial plant. It can be grown as an annual in most temperate climates, in well-drained soil with full sun. It does not grow well in cold climates with temperatures below freezing.

Constituents of G. pentaphyllum include sterols, saponin, flavonoids, and chlorophyll. Gypenosides have been extracted from its leaves. Some saponin compounds are the same as those found in ginseng roots. While there have been in vitro studies on toxicity, there have been no clinical trials, therefore no information is available about human toxicity.

The plant is used in folk medicine, typically as an herbal tea, but may be used as an alcohol extract or in dietary supplements. It has not seen widespread use in traditional Chinese medicine (TCM), being adopted only in the past 20 years, because it grows far from central China where TCM evolved; consequently, it was not included in the standard pharmacopoeia of the TCM system. Before then, it was a locally-known herb used primarily in mountainous regions of southern China and in northern Vietnam. It is described by the local inhabitants as the "immortality herb" (仙草, xiān cǎo), because a large number of elderly people within Guizhou Province reported consuming the plant regularly. In the European Union, jiaogulan is considered a novel food following a 2012 court ruling that prohibited its sale as food.2

WHAT IS GYNOSTEMMA/JIAOGULAN
Gynostemma, also known as Jiaogulan, is praised as “the herb of Immortality”. It is a climbing vine of the cucumber /gourd family. It is said to “have the most wide ranging benefits for human health and wellness of any plant yet discovered” (Dr.Jialiu Liu)
The oldest documented history of Jialgulan goes back to the Ming Dynasty (1368-1644) when it was harvested wild for food. A reknowed [sic] herbaslist [sic], Li Shi-Zhen has included this herb in his classical book Compendium of Materia Medica, describing its use for hematuria, edema and pain of the pharynx, heat and edema [sic] of the neck, tumors and trauma in 1578AD.
People in the provinces of Guizhou, Guangxi, and Sichuan in the mountains of south central China have been using jiaogulan (called xiancao by locals) as a general health tonic and rejuvenating elixier [sic]. It was largely unknown outside of these mountainous regions until the early 1970s.
A 1970s census in China showed a high percentage of centenarians, with low incidences of diseases that usually afflict the aging, in these regions. This started a research on jiaogulan’s possible antiaging properties by the Chinese government.
In 1972, The first study was conducted evaluating the therapeutic effects of the herb on 537 cases of chronic tracheo-bronchitis by the Research Group of Combined Traditional Chinese-Western Medicine of Qu Jing. Since then over 300 scientific papers have been published, and Gynostemma pentaphyllum has been included in the Dictionary of Chinese Materia Medica.
It was discovered by Japanese when they were researching on plants that could be used as sugar substitues [sic]. In 1977-78 Dr Masahiro Nagai discovered the saponins similar to those in ginseng, which were named gypenosides in Amachazuru, Japanese name for Jiaogulan. Over the next 10 years, another Japanese researcher, Dr Takemoto, along with his research group, identified 82 saponins in Amachazuru. Since then, 174 different gypenosides, 9 of whcich [sic] are also found in Panax, have been identified in Gynostemma pentaphyllum.
Jiaogulan’s health benefits are mostly due to these saponins. They give the adaptogenic and antioxidant properties as well as its ability to regulate Nitric Oxide production in the body.
The TCM qualities of jiaogulan are described as sweet, slightly bitter, neutral, warm, enhancing “yin” and supporting “yang,” and would be used to increase resistance to infection and to reduce inflammation.
ADAPTOGEN
If you are a regular reader here and fan of tonic herbs you will know how amazing adaptogens are. Let’s recap what they are.
Adaptogens are plants that can increase the body’s ability to adapt to physical, chemical, emotional, biological and environmental stressors. They do this by raising nonspecific resistance towards these stresses. Adaptogens support normal metabolism, immune, nervous and endocrine systems. They are intelligent herbs that can maintain body’s homeostasis.
For a plant to be qualified as an adaptogen it must meet these criteira [sic].
Non-toxic
Increase resistance to stress
Has normalising effect – bidirectional effect on functions.
Most of the time these adaptogenic qualities come from particular class of chemical compounds called triterpenoid saponins.
Gynostemma is often compared with Ginseng because nine of Gynostemma’s saponins are identical to those of Panax Ginseng. While Panax ginseng contains up to 28 saponins gynostemma is shown to contain at least 174 (in 2015). In fact Gynostemma has the widest spectrum of saponins of any known plant in the world by far.
Also high quality Ginseng can be more challenging to source and often not tolerated if the right variety was not chosen according to your individual consitution.3

Recommendations for use as a superior affordable substitute for quality dry ginseng is not without controversy, but misses the point that the range of different therapeutic compounds is as important as their relative concentrations in the source plant material:4

Hello, I received a leaflet with Thai Jiaogulan Herbal Tea, where, among other things, Jiaogulan identifies more than four times more effective saponins than ginseng root, and some healers perceive it and evaluate it as 10x greater. I know that with regard to the "perception of some healers" it is quite questionable, but you can send me a comparison of ginseng and the above tea, Your opinion? Alda.
Hello, Aldo!
This statement about Gynostemma pentaphyllum , jiao-ke-lan, is known. This is a specific lie. I have already mentioned the real state of affairs here - Gynostemma pentaphyllum - Jiaogulan - but the lie about gynostem is one of the so-called shameful lies that were often used in the politics of the 20th century. This lie is no deviation from reality, but from pure fiction - the opposite of reality. So you will not get the right opinion by mitigating my statement with the opponent's statement - you have to drop it completely into the wastebasket.
But before I go on I would like to point out that gynostema and the five-leaf is an amazing, important plant that is a unique alternative source of panaxosides, otherwise only available from ginseng. Gynostema is also a distinct healing plant that does not need to look like a ginseng as an older brother. It is not my goal to damage the reputation of this plant! In an article about gynostem I did not want to concentrate on this. But when we compare, then:
Quality dry ginseng ( Panax spp.) Contains about 20-24% panaxosid saponins. Dried leaf gynostemy contains 2.4% saponins, of which about one quarter is identical to ginseng panaxosides . Quantitatively, therefore, the gynostem drug contains 8 to 10 times less total saponins than ginseng and 30 to 40 times less panaxosides than ginseng. From a qualitative point of view, gynostema contains 10 species of panaxosides, while ginseng contains many tens.
Some people have decided that they can not write the truth - that gynostema contains 40x less panaxosides in dry matter than ginseng. Instead they created a story that contains "four times more saponins than ginseng", true in the sense that, in addition to about 0.6% of ginseng panaxosides, gynostema contains about three times as many (1.8%) of other non-ginsular gynosomic saponins. Therefore, they use the phrase "curative saponins". Do you understand? The people who created this story know the reality and use such a phrase to be able to defend themselves when they have been told otherwise. It was one of my friends in the casino: he told the croupier a double-minded sentence, and when he lost, he claimed the bet that he was thinking otherwise.
Yet gynostema has a unique position in the function of ginseng substitution (" southern ginseng "), because it contains panaxosides as the only plant outside the genus Panax . Gynostema is a type of pumpkin ( Cucurbitaceae ) and its saponins are obtained from leaves, so it is very cheap compared to real ginseng. This thermophilous plant can be well grown in ours (greenhouse, balcony or even apartment) . I would recommend it to grow for all seniors who can not afford to buy herbs.

Antiviral activity

Saponins (Latin "sapon", soap + “-in", one of), also selectively referred to as triterpene glycosides, are bitter-tasting usually toxic plant-derived organic chemicals that have a foamy quality when agitated in water. They are widely distributed but found particularly in soapwort (genus Saponaria), a flowering plant, and the soapbark tree (Quillaja saponaria). They are used in soaps, medicinals, fire extinguishers, speciously as dietary supplements, for synthesis of steroids, and in carbonated beverages (the head on a mug of root beer). Structurally, they are glycosides, sugars bonded to another organic molecule, usually a steroid or triterpene, a steroid building block. Saponins are both water and fat soluble, which gives them their useful soap properties. Some examples of these chemicals are glycyrrhizin, licorice flavoring; and quillaia (alt. quillaja), a bark extract used in beverages.”5

Protease
The most studied of the coronavirus enzymes is most definitely the protease 3CL (also known as MPro). Here shown with its substrate (protease from PDB:6Y2E and peptide TSAVLQSGFRK from SARS protease PDB:2Q6G).
Role
To save genetic space, many RNA viruses employ a strategy wherein different protein acting at the same stage of the infection cycle are produced as a single long peptide that gets cut up by a protease at given recognition sequences. In the case modelled here, between a glutamine and a serine.
(figure from Kiemer et al. 2004)
Activity
A protease works thanks to a catalytic triad: an acid hydrogen bonds to a base, which in turns deprotonates a nucleophile making it more active. In the case of viral protease the acid is part of the ligand, so it's a dyad. With 2019-nCoV (and SARS) histidine-163 is the base and cysteine-145 is the nucleophile.
The cysteine attacks the backbone ketone of the glutamine of the substrate. The tetrahedral hemiacetamide intermediate is highly unstable and the N-terminal half of the peptide leaves. The thioester between the enzyme and the intermediate is more stable but undergoes a substition reaction with water (hydrolysis) freeing the enzyme for another round of catalysis and releasing the second half of the substrate.
Inhibitors
Given the following:
it is a very essential protein, so inhibiting it will stop the infection
its substrate is large, which means it has an exposed active site to accommodate its substrate, so drug design is easier
its substrate is a long peptide, so it is easy to make peptidomimetic drugs.
It is clear why viral protease made good candidates. In fact, the crystallisation of the HIV protease was a big deal —appearing the Voet and Voet textbook and featuring in the book The Billion-Dollar Molecule and there are many HIV protease inhibitors on the market (NB. HIV protease has a diffent structure but works on the same principle).6

From a paper published in 2021, Rehan and Shafiullah conducted an in silico molecular docking binding analysis of 60 saponins with the COVID-19 main protease as above, 6LU7 Mpro.7

Highlights
Approximately 34 saponins are more effective on COVID-19 Mpro than hydroxychloroquine, chloroquine, and nelfinavir. 13 saponins exhibit high potency against COVID-19 Mpro due to more binding energies than 10 kcal/mol.
Tradition
Medicinal plants have been used for health care since ancient times worldwide in the traditional system of medicine such as Unani, Siddha, Ayurveda, and traditional Tibetan and Chinese medicine. Jiaogulan (Gynostemma pentaphyllum) is the source of the dammarane-type saponins, which was described in 1406 C.E. by Zhu Xiao in the ancient Chinese medicine classics Materia Medica for Famine as a survival food. Renshen (Panax ginseng) which was first recorded in Shennong's Classic of Materia Medica written between about 200 C.E. and 250 C.E. is a big source of saponins, which was used for medicinal purposes over 3,000 years ago. Various types of saponins such as hopane, lupane, and oleanane have been isolated from many parts such as leaves, roots, barks, stems, and rhizomes of various herbs. The main advantage of saponins is to protect plants against various pathogen attacks.
Abstract
Background:
Recently, the Chinese scientists Liu et al. demonstrated a crystallized form of severe acute respiratory syndrome coronavirus-2 main protease (Mpro), the best target of the drug, which was published in Nature in June 2020. Many components of herbs are determined as the potential inhibitors of coronavirus disease 2019 (COVID-19) Mpro such as quercetin, cirsimaritin, hispidulin, and flavonoids.
Methods:
Library of herb-based bioactive saponins are analyzed with 6LU7 Mpro using AutoDock tools 1.5.6, BIOVIA Discovery Studio 2017 R2, Chimera 1.13.1, and AutoDock Vina to evaluate their potency against COVID-19 Mpro. The conventional Western medicines, including hydroxychloroquine, chloroquine and nelfinavir, are used as positive controls for comparison.
Results:
Binding energies of 60 saponins with 6LU7 Mpro are obtained in which approximately 34 saponins are more effective on COVID-19 Mpro than hydroxychloroquine, chloroquine, and nelfinavir. 13 saponins exhibit high potency against COVID-19 Mpro due to more binding energies than 10 kcal/mol.
Conclusion:
Further research on all effective saponins is needed to evaluate the real medicinal potential against COVID-19.

Discussion
…Approximately, 34 diverse types of saponins were showing more binding affinity with COVID-19 Mpro than hydroxychloroquine, chloroquine, and nelfinavir, and their main original medical plant resources were shown in Table 4. 3-O-β-D-Xylopyranosyl-6-O-β-D-glucopyranosyl-16-O-β-D-glucopyranosyl-3β,6α,16β,24(S)-25-pentahydroxycycloartane is a cycloartane-type saponin found in Astragalus brachycalyx plant [30] (Table 4, entry 1). This saponin was showing an excellent binding affinity (−11.9 kcal/mol) with COVID-19 Mpro. Gypenoside J1 (Table 4, entry 7), gypenoside J2 (Table 4, entry 11), gypenoside J3 (Table 4, entry 12), and gypenoside LVII (Table 4, entry 14) are a dammarane-type saponin reported from Gynostemma pentaphyllum plant [35]. Gynostemma pentaphyllum plant is a big source of dammarane-type saponins, which showed an excellent binding affinity in the range from −10.4 to −9.7 kcal/mol. Schekwanglupaside A (Table 4, entry 19) and schekwanglupaside B (Table 4, entry 33) are lupane-type saponins found in the Schefflera kwangsiensis plant which showed a moderate binding affinity in the range from −9.1 to −8.1 kcal/mol [42]. Saponins are very important for human life which play a vital role in diverse biological activities.

The authors listed 40 saponins in total from various medicinal plants including dammarane-type ginsenoside Rg12 from Panax ginseng, and modelled binding with COVID-19 Mpro.

Thirteen highly potential saponins (binding affinity above −10 kcal/mol) in this study may show a better outcome in COVID-19. This method presented here can play a highly potential role in rapid drug discovery with a clinical trial against the COVID-19.

Ginsenoside Rg12, (*Panax ginseng*/roots), **docking affinity: -10.9 kcal/mol**

Some of the related dammarane-type saponins found in G pentaphyllum. It is encouraging to see such a mix of saponins, as this could make it harder for the virus to select mutations that are resistant to binding, and 3 of the 5 have excellent binding affinity:

Gypenoside J1, (*G. pentaphyllum*/aerial parts), **docking affinity: -10.2 kcal/mol**

Gypenoside J2, (*G. pentaphyllum*/aerial parts), **docking affinity: -10.4 kcal/mol**

Gypenoside J3, (*G. pentaphyllum*/aerial parts), **docking affinity: -10.2 kcal/mol**

Gypenoside LVII, (*G. pentaphyllum*/aerial parts), docking affinity: -9.7 kcal/mol

Gylongiposide I, (*G. pentaphyllum*/aerial parts), docking affinity: -8.9 kcal/mol

Conclusion
…Approximately, 34 diverse types of saponins were showing more binding affinity with COVID-19 Mpro than hydroxychloroquine, chloroquine, and nelfinavir and may act as COVID-19 Mpro inhibitors, among which 13 highly potential saponins have binding affinity above −10 kcal/mol. So, further research on medicinal plant-derived saponins and plant extract is necessary for the treatment of COVID-19.

Objectives: Yellow fever is a disease of significant public health concern with no known drug for treatment. There is therefore the need for a suitable antiviral drug against the disease. This has given rise to the present research into medicinal plants for suitable alternative antiviral drugs since most antiviral drugs known have side effects on the host cells.
Methods: The antiviral activity of the crude extracts of Gynostemma pentaphyllum on yellow fever virus was evaluated. The antiviral properties were determined against yellow fever virus using three methods: protection of chicken egg embryo against viral infectivity by the extracts, protection of mice against viral infectivity by the extracts and percentage inhibition of viral induced hemagglutination by the extracts. Phytochemical analysis of the extracts was also carried out.
Results: The phytochemical analysis of the extracts revealed the presence of saponins, alkaloids, glycosides, tannins, flavonoids, carbohydrates, reducing sugar, resins, acidic compounds, fats and oil and proteins. The egg embryo and mice protection studies against yellow fever viral infectivity showed that the extracts were able to give up to 100% protection to the embryonated eggs and mice and hence prevented egg/mice mortality. The extracts gave plausible percentage inhibitions of yellow fever virus in embryonated eggs up to 90%. It was observed that the antiviral effect decreased with decrease in concentration.
Conclusion: The research has shown that the plant Gynostemma pentaphyllum possesses potent antiviral potential and could serve as a possible source of lead antiviral drugs against yellow fever since the disease has no known drug for treatment.

Passaged virus with sterile phosphate buffered saline (PBS) acted as the positive control, PBS alone acted as the negative control and G. pentaphyllum extracts alone acted as toxicity controls.

They used 5 replicants of each.

The aqueous extract of Gynostemma pentaphyllum at the concentration of 200 mg/mL gave 100% protection to the embryonated eggs for the three inoculation modes (pre infection, at infection and post infection). At 20 mg/mL it gave 100%, 60% and 80% protection for pre infection, at infection and post infection respectively. The 2 mg/mL concentration gave 20%, 80% and 20% protection for pre infection, at infection and post infection respectively. The positive control (virus alone) showed the death of all eggs inoculated and therefore gave 0% protection to the embryonated eggs. For the toxicity control all eggs inoculated survived showing that the extracts were not toxic to the cells (Figure 1).

FIG. 1: Protection of embryonated eggs against yellow fever virus by **aqueous extract** of *Gynostemma pentaphyllum*

Methanolic extracts showed 20% less protection pre-infection, but much greater protection at infection and post infection:

FIG. 2: Protection of embryonated eggs against yellow fever virus by **methanolic extract** of *Gynostemma pentaphyllum*

FIG. 3: Protection of embryonated eggs against yellow fever virus by **ether extract** of *Gynostemma pentaphyllum*

Mice inoculation: Ten microlitres of different concentrations of methanolic extract of the plant were separately inoculated into five suckling mice less than one week old intraperitoneally.

The only mouse deaths were when extracts were administered post infection, but even then with a survival rate of 60% even at the lowest dose, vs a 0% survival rate from the positive control:

Discussion
The phytochemical screening of the extracts of Gynostemma pentaphyllum showed the presence of saponins, alkaloids, glycosides, tannins, flavonoids, carbohydrates, reducing sugar, resins and proteins. The presence of saponins, glycosides and flavonoids in the extracts of Gynostemma is consistent with other findings about the leaves of the plant (Xin 2004, Cui 1999) The leaves of Gynostemma pentaphyllum have been shown to contain more than 90 saponins and more than 100 dammarane type glycosides have been isolated and identified from it (Zhang 1993, Cui 1999). The extracts have shown varying degrees of antiviral activities against the yellow fever virus assayed. The antiviral activities may be attributed to the rich phytochemicals contained in the extracts since various studies have shown that phytochemicals such as tannins found in almost all plant parts cure or prevent a variety of viral infections (Serafini 1994, Nonaka 1990).
Flavonoids on the other hand have been shown to exhibit inhibitory effects against viruses including HIV and respiratory syncytial virus (Li 2000). Plant polysaccharides have also been shown to exhibit potent antiviral activities especially against enveloped viruses (Hosoya 1991, Premanathan 1990). In fact Abram et al (1993) attributed the medicinal properties of Gynostemma pentaphyllum to be mainly due to the presence of saponins in the plant.

The mice inoculation assay for the determination of the extracts’ protection of mice against viral infectivity showed that the extracts were able to prevent the symptoms of encephalitis and death in many of the mice and some of the extracts gave up to 100% protection to the mice. This result corroborates other research findings that showed the antiviral activities of this plant using animal models against such viruses as Epstein-Barr virus (EBV), Herpes Simplex virus (HSV-1) and HIV–AIDS virus (Lipipum 2003, Abram 1993). Nevertheless this is the first report of the antiviral activity of this plant against yellow fever virus.

The result of the embryo mortality clearly showed that the extracts were not toxic to the chicken embryonated eggs since all embryos used for toxicity control survived by the fifth day of the experiment. The extracts with antiviral effect showed activity in two subsequent dilutions of the maximum non toxic concentration. This is in line with the suggestion made by Vanden et al (1993) that the antiviral activity of crude plant extracts should be detectable in at least two subsequent dilutions of the maximum non toxic concentration to ensure that the activity is not directly correlated with the toxicity of the extracts.
It is interesting to note that the extracts had activity against the yellow fever virus at different times of inoculation (one hour pre infection, 0 h at infection and one hour post infection). The extracts that inhibited viral replication at 1 h pre infection might have acted on the viral entry step of the replication cycle and prevented the virus from attachment and further replication inside the host. Those extracts that inhibited the yellow fever virus at zero hour might have acted on the virus before attachment by mechanism of binding on the active site of the host cell blocking the virus from attaching to the host receptors or by binding on the active site of the virus. According to Vanden et al (1986) polyphenols act principally by binding to the virus and/or the protein of the host cell membrane thus arresting adsorption of the virus. The extracts that inhibited 1 h post infection must have inhibited a post entry step of the viral replication. The extracts gave up to 100% protection with the three modes of inoculation. Viral infectivity was also inhibited up to 90% by the extracts of the plant inoculated in three different modes. This shows that the three modes gave good inhibition of viral growth and protection on the embryonated eggs and mice and that no mode of inoculation worked better than the other. This suggests that the extracts could be used as both preventive and curative therapy. It was observed that the antiviral activity decreased with a decrease in concentration as the 200 mg/mL concentration showed the highest activity while 2 mg/mL showed the least activity.
Conclusion
The extracts of the plant Gynostemma pentaphyllum used in this study have shown credible antiviral activities against the yellow fever virus and could be recommended as a potential source for a yellow fever remedy.

No conflicts of interest were registered.

From the same year, 2012, the same author Okoye worked with Nworu to study “Inhibition of HIV-1 lentiviral particles infectivity by Gynostemma pentaphyllum extracts in a viral vector- based assay”9. As with the previous study they performed solvent extraction from leaf powder using ethyl ether (EG), methanol (MG) or water (AG).

Again the results from this in vitro research pointed to potential antiviral drugs that could be developed using extracts from the herb:

Three different extracts of Gynostemma pentaphyllum (Cucurbitaceae), a medicinal plant used for a variety of ailments in complementary and alternative medicine (CAM) including those caused by viral infections with claims of efficacy against HIV-1 infections were screened. These claims motivated the study in which the inhibition of viral vector infectivity of HeLa cells was assessed flow cytometrically by measuring the expression of green fluorescent protein (GFP) transgene incorporated in the lentiviral vector construct. An infectious VSV-G-pseudotyped, human immunodeficiency virus type 1-based, self-inactivating lentivirus vector particles were generated by transient co-transfection of the vector plasmid (pHIV-1 CSCG), with packaging plasmids encoding tat, rev, gag-pol (pCMV∆R8.2), a VSV-G expression plasmid (pHIT-G) and a secretory alkaline phosphate expression plasmid (pSEAP) all necessary for viral infectivity. The extracts studied were obtained by solvent extraction of the leaf powder of G. pentaphyllum with ethyl ether (EG), methanol (MG), and water (AG). The AG, MG and EG were all active against the HIV-1 lentiviral vector and inhibited the early events of the viral replication cycle on HeLa cells in a concentration-dependent manner with a IC50 of 6.21 µg AG/ml, 8.32 µg MG/ml and 5.8 µg EG/ml, respectively. The cytotoxicity of the extracts to HeLa cells evaluated in parallel by the 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay method showed TC50 values of 36.77 µg AG/ml, 38.68 µg MG/ml and 41.02 µg EG/ml with selectivity indices (SI) of 5.92, 4.65 and 7.02, respectively. The results of the study show that the extracts of G. pentaphyllum possess potent and selective anti-retroviral potentials and could serve as possible source of lead antiviral drugs against HIV.
Key words: Antiviral activity, antiviral screening, Gynostemma pentaphyllum, HIV-1, viral vector-based screening.

DISCUSSION
The need for the discovery of novel anti-viral agents is increasingly felt due to many shortcomings of the existing drugs, which includes their inherent cytotoxicity to the host cells, the ease with which resistance are developed and the emergence of new viral variants which are not susceptible. In this study, we used a novel vector-based assay technique to screen lipophilic and polar solvent extracts of the plant, G. pentaphyllum. A VSV-Gpseudotyped, human immunodeficiency virus type 1- based, self-inactivating lentivirus vector that expressed GFP under the control of cytomegalovirus promoter was constructed and used as viral infective particles on HeLa cells.

Transient supply of the packaging plasmids and the self-inactivation of the vector by deletion of the U3 region in the 31 -LTR ensured that the resulting lentiviral vector was only capable of a single round of replication, which makes the viral vectors comparatively safer than their parent wild retroviral virus. The infectious vector particles additionally differed from the wild-type HIV-1 virus in that they lacked some of the HIV-1 accessory genes such as nef, vif, vpu and vpr. Due to the transgene (in this case, green fluorescent protein) being integrated into the genome of target cells in the process of infection, vector infectivity in the presence or absence of various concentrations of the extracts is easily determined flow cytometrically as a function of the amount of GFP expression. In this system, GFP expression is driven by an internal CMV promoter and occurs after integration. The implication is that inhibitors of late stages in viral replication cycle such as inhibitors of new viral assemblage and inhibitors of budding are not detected by the screening assay used in the study.
The results of the study show that the extracts inhibited early events in viral replication cycle. Although HIV-1 has been shown to transduce both dividing and non-dividing cells (Mendoza et al., 1997; Merluzzi et al., 1990; Pannell, 1992), the cell cycle-dependence of productive HIV-1 and other retroviral infection is well established (De Clercq, 2000; Raff and Glover, 1988; Yang et al., 1999). Using the Rous sarcoma virus (an avian retrovirus) as a prototype retrovirus, it was shown that arrest of cells at the G0 phase resulted in failure of reverse transcription (and hence blockage of productive infection) (Zack et al., 1990; Esimone et al., 2007), while arrest of cells during the S phase did not affect the reverse transcription or integration process (Fritsch and Temin, 1977; Zack et al., 1990). The selectivity index of the G. pentaphyllum extracts (4.65 to 7.02) against the HIV-1 lentiviral vector is sufficiently large to ascertain that the antiviral effects are not simply due to the cytotoxicity of the extracts.
Phytochemical analysis of the extracts of G. pentaphyllum showed generally, the presence of saponins, alkaloids, glycosides, tannins, carbohydrates, flavonoids, resins, acidic compounds and proteins. These phytoconstituents were present in varying degrees in the different solvent extracts of G. pentaphyllum (Table 1). Although, the scope of the work is not yet enough to associate the antiviral activity of G. pentaphyllum extracts to a particular phyto-constituent, the antiviral activities may be attributed to some phytoconstituents contained in the extracts such as the tannins, which has been found in previous studies to prevent a variety of viral infections (Serafini et al., 1994, Nonaka et al., 1990). Tannins have also been demonstrated to inhibit viral reverse transcriptase (Nonaka et al., 1990). Similarly, flavonoids have been shown to exhibit inhibitory effects against viruses including HIV and respiratory syncytial virus (Li et al., 2000). G. pentaphyllum is also rich in saponins which have been reported to be responsible for most of the observed activities of the plant (Cui et al., 1999).

It could be even more efficacious against the wild type virus:

The pattern of inhibition of the HIV lentiviral vector suggests that the extracts either directly interacted with the vector particles inhibiting the envelop protein or that it interacted with some host cells-derived components of the viral particle which could be the lipid membrane derived from the cell or the cellular membrane proteins that are frequently incorporated in lentiviral particles during budding (Gould et al., 2003). In previous studies, correlations between anti-HIV-I vector activity and anti-wild type HIV-I activity have been demonstrated. In one of such studies, Steinstraesser et al. (2005) demonstrated in an assay that the porcine defensin (protegrin-I) showed more than three-fold higher activity against the wild type HIV-I than against the corresponding lentiviral vector. It was also shown that some antiviral medicinal plant extracts showed up to ten-fold higher activities against wild type lentiviruses as against the corresponding lentiviral vector particles (Esimone et al., 2005). This is very instructive and points to the reliability of the high through-put viral vector particles-based assay in the screening of putative substances for antiviral activities. It also suggested that the extracts screened in this study using the lentiviral HIV-I vector could possibly show higher antiviral activities when used against wild type HIV-I virus.

No conflicts of interest were declared.

Sornpet et al (2017) investigated the antiviral activity of five Asian medicinal plant crude extracts against H5N1 avian influenza virus.10

Although efficacious, as Curcuma longa (turmeric) and Kaempferia parviflora (Thai black ginger) had the strongest antiviral activity they didn’t select G. pentaphyllum for further analysis of its effects on cytokine expression. This may have been a questionable conclusion as they didn’t conduct in vivo bioavailability studies - which is very low for turmeric unless you supplement with piperine for example.11

Bioavailability of several saponins in G. pentaphyllum is much greater in comparison, as will be discussed later.

Abstract
Objective
To study the antiviral properties of the five Asian medicinal plants against in vitro infection by the highly pathogenic avian influenza virus (H5N1).
Methods
Crude extracts of Andrographis paniculata, Curcuma longa (C. longa), Gynostemma pentaphyllum, Kaempferia parviflora (K. parviflora), and Psidium guajava obtained by both water and ethanol extractions were investigated for their cytotoxicity in the Madin–Darby canine kidney cells. Thereafter, they were investigated in vitro for antiviral activity and cytokine response upon H5N1 virus infection.
Results
The results revealed that both water and ethanol extracts of all the five studied plants showed significant antiviral activity against H5N1 virus. Among these plants, C. longa and K. parviflora showed strong anti-H5N1 activity. Thus, they were selected for further studies on their cytokine response upon virus infection. It was found that ethanol and water crude extracts of C. longa and K. parviflora induced significant upregulation of TNF-α and IFN-β mRNA expressions, suggesting their roles in the inhibition of H5N1 virus replication.
Conclusions
To the best of the authors' knowledge, this study is among the earliest reports to illustrate the antiviral property of these Asian medicinal plants against the highly pathogenic avian H5N1 influenza virus. The results of this study shed light on alternative therapeutic sources for treatment of H5N1 influenza virus infection in the future.

Low cytotoxicity, which is excellent. Higher is better:

3. Results
3.1. Plant preparation and determination of its toxicity on MDCK cells
To investigate the anti-H5N1 virus activity of the five Asian medicinal plants, including A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava, crude extracts using water and ethanol extractions were obtained. The plant part used and the percentage yield obtained by the two different solvents are shown in Table 1. The cytotoxicity test on the MDCK cells of the ethanol extracts of A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava study reagents revealed that at high concentrations of 8.2 g/mL, 69.3 g/mL, 135.6 g/mL, 2.2 g/mL, and 90.7 g/mL, respectively, these ethanol extracts did not show any cytotoxic effect to the cells (Table 1). On the other hand, the water extracts of A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava at high concentrations of 380.3 g/mL, 142.3 g/mL, 468.2 g/mL, 438.4 g/mL, and 195.6 g/mL, respectively, showed no toxicity on the tested cells (Table 1). These concentrations of the extracts were, therefore, chosen for further studies on their antiviral activity in vitro.

Results using water extracts were still significant when compared to the DMSO control. Lower is better:

3.2. Screening of extracts for anti-H5N1 virus activity
The results of the antiviral test show that crude extracts obtained by water and ethanol extractions of A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava significantly (P < 0.05) inhibited H5N1 virus replication in MDCK cells when compared to the control group (Figure 1). Among these plants, inhibition of the H5N1 virus replication was predominantly observed as early as 24 hpi by the ethanol extract of C. longa (Figure 1). Furthermore, at 48–72 hpi, the water and the ethanol crude extracts of C. longa and K. parviflora showed significant antiviral activity (P < 0.05) against the H5N1 virus. The plants were, therefore, chosen for further investigation of their induction of cytokine response in the tested cells after H5N1 virus infection. Here, it needs to be emphasized that the antiviral property of A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava study reagents were not due to DMSO since the control group, the DMSO-containing medium, did not affect the virus growth kinetic (Figure 1).

Figure 1. The antiviral activity of *A. paniculata*, *C. longa*, *G. pentaphyllum*, *K. parviflora*, and *P. guajava* crude extracts on the H5N1 virus at the multiplicity of infection (MOI).

The five Asian medicinal plants used in this study were particularly chosen since there have been reports of their antiviral activity against herpes simplex virus type 1, Newcastle disease virus, hepatitis B virus, influenza H1N1 virus, human immunodeficiency virus (HIV), and human cytomegalovirus virus, exhibited especially by A. paniculata, G. pentaphyllum, C. longa, P. guajava, and K. parviflora crude extracts, respectively. Despite reports that these five medicinal plants inhibit pathogenic H5N1 influenza virus replication, the results of this study indicate that among these plants, only the crude extracts of C. longa and K. parviflora obtained by both water and ethanol extractions showed strong antiviral activity against the H5N1 virus.

In conclusion, the present study demonstrates that crude extracts of A. paniculata, C. longa, G. pentaphyllum, K. parviflora and P. guajava at appropriate concentrations potentially inhibit H5N1 virus replication in vitro. The results of this study may be useful for application of these five medicinal plants in the control and prevention of H5N1 virus in the future.
Conflict of interest statement
The authors declare that there is no conflict of interest.

Cardiovascular diseases

This review is unusual, so far, in that it considers available clinical trials data as well as safety, toxicity and side effects:

Figure 1
Pathological processes involved in the development and progression of CVDs. Several risk factors can predispose to CVDs. These can include hypertension, smoking, dyslipidemia stemming from an unhealthy diet, or endocrinopathies like diabetes mellitus, hypothyroidism, and aging. The risk factors can lead to pathological alterations most of which can be due to endothelial dysfunction or VSMC alterations. Endothelial dysfunction or VSMC alterations increase the risk of developing atherosclerosis and hypertension. Atherosclerosis and hypertension are themselves CVDs risk factors and enhancers for the development of other CVDs like myocardial infarction, coronary artery diseases, or stroke. VSMC, vascular smooth muscle cell; ECM, extracellular matrix; NO, nitric oxide; eNOS, endothelial nitric oxide synthase; iNOS, inducible nitric oxide synthase; Ox-LDL, oxidized low-density lipoprotein.

“Superoxide dismutase (SOD, EC 1.15.1.1) is an enzyme that alternately catalyzes the dismutation (or partitioning) of the superoxide (O−2) radical into ordinary molecular oxygen (O2) and hydrogen peroxide (H2O2). Superoxide is produced as a by-product of oxygen metabolism and, if not regulated, causes many types of cell damage. Hydrogen peroxide is also damaging and is degraded by other enzymes such as catalase. Thus, SOD is an important antioxidant defense in nearly all living cells exposed to oxygen.”13

Not just linked to atherosclerosis: “The phosphoinositide 3-kinase (PI3K) signaling pathway is one of the most frequently altered pathways in human cancer and has a critical role in driving tumor initiation and progression.”14

The TNF-α-induced NF-κB inflammatory pathway is also upregulated by both COVID-19 and its component spike protein, via transfection.15

“Lipid sensor peroxisome proliferator-activated receptor alpha (PPAR- α) is the master regulator of lipid metabolism.”16

Note the presence of quercetin, a potent therapeutic agent in its own right.17

G. pentaphyllum at the Bench: Mechanism of Action in CVDs
As already mentioned, antioxidants are important when it comes to the prevention of atherosclerosis. Four flavonoids—namely quercetin-3-O-(2″,6″-di-α-L-rhamnosyl)-β-D-galactopyranoside, quercetin-3-O-(2″,6″-di-α-L-rhamnosyl)-β-D-glucopyranoside, quercetin-3-O-(2″-α-L-rhamnosyl)-β-D-galactopyranoside, and quercetin-3-O-(2″-α-L-rhamnosyl)-β-D-glucopyranoside—with potent antioxidant effects against DPPH and OH free radicals, in vitro, were found in the extracts of G. pentaphyllum. These flavonoids also exhibited cytoprotection against AAPH-induced oxidative damage in pig kidney LLC-PK1 cells by suppressing the increase of MDA, and limiting the decrease of SOD and GSH (Lin et al., 2019). In another study, flavonoids from G. pentaphyllum were extracted and tested on human lung carcinoma A549 cells. It was found that the flavonoids protected A549 cells against hydrogen peroxide-induced oxidative damage by increasing the expression levels members of the endogenous antioxidant system including SOD, GSH, Nrf2, NQO1, and HO-1 (Wang et al., 2018). Another study evaluated the antioxidant potential of one G. pentaphyllum component, the phytoestrogen gypenoside XVII, it was found that the phytoestrogen alleviated atherosclerosis via the ERα-mediated PI3K/Akt pathway (Yang et al., 2017). A prior study evaluated the effects of gypenosides of G. pentaphyllum on hydrogen peroxide-induced oxidative damage in bovine pulmonary artery ECs. The gypenosides protected the ECs from oxidative injury further suggesting its potent antioxidant activity as well as its prospective use as a preventative supplement against atherosclerosis (Li and Lau, 1993).
Inflammation can contribute to the onset of atherosclerosis and other CVD risk factors, hence, reducing inflammation can act as a protective factor in CVDs. The gypenoside XLIX (Gyp-XLIX) from G. pentaphyllum has been studied for its anti-inflammatory properties. Gyp-XLIX inhibited LPS- and TNF-α-induced NF-κB activation in THP-1 monocytes and in HUVECs. Gyp-XLIX inhibition of NF-κB activation appears to be through a PPAR-α-dependent pathway (Huang et al., 2006). On the other hand, contradictory results were reported by Aktan et al., where G. pentaphyllum gypenosides attenuated NF-κB activation. In fact, the gypenosides extracted from G. pentaphyllum could suppress NO production by inhibiting iNOS activity and levels in murine macrophages. Gypenoside-mediated decrement of iNOS protein expression turned out to be mediated by the inhibition of NF-κB activation (Aktan et al., 2003). In a different study, Tanner et al. showed that G. pentaphyllum could elicit beneficial effects on vascular function by acting as an inducer of eNOS (Tanner et al., 1999).
Attenuating lipid accumulation may decrease CVDs incidence. A study assessed the role of ombuine, a dual agonist of PPAR-α and PPAR-δ/β in lipid metabolism. Ombuine or mbuin-3-O-β-d-glucopyranoside, a flavonoid from G. pentaphyllum, were applied to HepG2 cells. Ombuine-stimulated HepG2 cells had activated PPAR-α and PPAR-δ/β, transcription factors that enhance lipolysis. Ombuine-mediated activation of PPAR-α and PPAR-δ/β significantly reduced intracellular concentrations of triglyceride and cholesterol as well as decreased lipogenic gene expression witnessed as decreased levels of sterol regulatory element binding protein-1c and stearoyl-CoA desaturase-1. These findings further our understanding of how G. pentaphyllum may be involved in lipid metabolism (Malek et al., 2013).
In a study that evaluated the role of total flavonoids of G. pentaphyllum on apoptosis in cardiomyocytes of neonatal rats, it was found that hypoxia-reoxygenation (H/R)-cardiomyocytes had an increased protein expression of apoptosis-associated Fas/FasL genes. Flavonoids of G. pentaphyllum could protect cardiomyocytes against H/R injury by decreasing the production of TNF-α and downregulating the protein levels of Fas/FasL genes leading to inhibition of myocyte apoptosis (Le et al., 2007).

In a study involving Wistar rats, a mixture of Fermentum rubrum Hongqu and G. pentaphyllum gypenosides revealed that this had anti-atherosclerotic effects that were better than those from a statin, simvastatin:

In Vivo Preclinical Evaluations of G. pentaphyllum
Gypenosides are G. pentaphyllum key components with the ability to help prevent atherosclerosis. The anti-atherosclerotic effects of a mixture (HG) of Fermentum rubrum Hongqu and G. pentaphyllum gypenosides were investigated in Wistar rats. The study results revealed that the HG mixture had anti-atherosclerotic effects that were better than statin, simvastatin, treatment highlighting the anti-atherosclerotic potential of G. pentaphyllum (Gou et al., 2018). In addition, HG alleviated oxidative stress biomarkers by restoring antioxidant defense components and decreasing the serum levels of anti-inflammatory cytokines in male Sprague-Dawley rats with fatty liver disease. The HG mixture in this study displayed athero-protective characteristics (Gou et al., 2016).

“Ischemia or ischaemia is a restriction in blood supply to any tissues, muscle group, or organ of the body, causing a shortage of oxygen that is needed for cellular metabolism (to keep tissue alive). Ischemia is generally caused by problems with blood vessels, with resultant damage to or dysfunction of tissue i.e. hypoxia and microvascular dysfunction. It also means local hypoxia in a given part of a body sometimes resulting from constriction (such as vasoconstriction, thrombosis or embolism). Ischemia comprises not only insufficiency of oxygen, but also reduced availability of nutrients and inadequate removal of metabolic wastes. Ischemia can be partial (poor perfusion) or total blockage. The inadequate delivery of oxygenated blood to the organs must be resolved either by treating the cause of the inadequate delivery or reducing the oxygen demand of the system that needs it. For example, patients with myocardial ischemia have a decreased blood flow to the heart and are prescribed with medications that reduce chronotrophy and ionotrophy to meet the new level of blood delivery supplied by the stenosed so that it is adequate.”18

Emphasizing the reported G. pentaphyllum gypenoside anti-inflammatory and antioxidant capabilities, Yu et al. demonstrated their beneficial effects in an ischemia–reperfusion injury rat model where they were found to inhibit apoptosis. Ischemia–reperfusion injury has detrimental outcomes in CHD. Yu et al. found that in ischemia reperfusion injured-rats, the administration of gypenosides decreased apoptotic rates as well as improved cardiac function. Gypenosides inhibited ER-stress and apoptosis through the blockade of the CHOP pathway and the activation of PI3K/Akt pathway (Yu et al., 2016b). In a different study, it was demonstrated that pretreatment with gypenosides limited the infarct size and relieved ischemia reperfusion-induced pathological changes in the myocardium, also in an ischemia reperfusion injury rat model. Additionally, left ventricle function was preserved. Molecularly, gypenosides pre-treatment reduced oxidative stress and restored the antioxidant machinery in the myocardium. The cardio-protective effects were also evidenced by the preservation of mitochondrial function in myocytes. In this regard, the maintenance of the mitochondrial membrane integrity inhibited the release of cytochrome c from the mitochondria into the cytosol. This further demonstrated the role of gypenosides from G. pentaphyllum as a cytoprotective agent against acute myocardial infarction and reperfusion injury (Yu et al., 2016a).
As elucidated to earlier, diabetes is positively correlated with the development of CVDs. The effect of G. pentaphyllum extracts on fasting blood sugar levels in diabetic mice was assessed. It was found that the extracts had inhibitory effects on α-glucosidase activity while affecting the protein expression of GLUT2 which highlights its potential to manage diabetes (Wang et al., 2019).

Beneficial effects on serum triglycerides and high density to low density lipoprotein ratios were demonstrated, but without evidence of depletion of the antioxidant coenzyme Q1019 or the cardioprotective vitamin K220, that is paradoxically associated with long term statin use.

According to the hypothesis, HDL=“good cholesterol”, LDL= “bad cholesterol” as it is prone to inflammatory oxidation in arterial walls.21

Attenuating obesity, a risk factor for developing CVDs, may decrease CVD incidence. G. pentaphyllum is largely used for the management of diseases such as hyperlipidemia, fatty liver, and obesity in China. G. pentaphyllum was found to affect lipid metabolism and elicit anti-hyperlipidemic effects by elevating the levels of phosphatidylcholine and decreasing the levels of trimethylamine N-oxide in the plasma and liver of rats (Wang et al., 2013). In addition, Gauhar et al. studied the effects of heat processed ethanol extracts of G. pentaphyllum on obese mice. They found that this extract decreased obesity in ob/ob mice by activating the AMP-activated protein kinase (AMPK) pathway. This study suggested a possible mechanism for fat-loss as well as a potential for the use of G. pentaphyllum as a weight-loss supplement (Gauhar et al., 2012). Gypenosides anti-hyperlipidemic effects were examined in rats with poloxamer P407-induced hyperlipidemia. Gypenosides at 250 mg/kg of body weight was orally administered to hyperlipidemic rats. Four and 12 days of gypenosides administration reduced plasma triglycerides levels by 53% and 85%, respectively. Similarly, total cholesterol levels were decreased by 10% and 44%, respectively. Interestingly enough, results were similar to atorvastatin cholesterol-lowering statin drugs. Additionally, LDL levels were reduced and HDL levels were increased by gypenoside, which also reversed the poloxamer P407 inhibition of lipoprotein lipase activity. This shows a promising therapeutic potential of G. pentaphyllum for lowering high triglyceride and cholesterol levels during acute hyperlipidemia (Megalli et al., 2005). In accordance, extracts from the plant also decreased triglyceride levels and LDL levels in obese Zucher rats (Megalli et al., 2006). By employing a new extraction technique, Lee et al. described some biological activity of a G. pentaphyllum extract with a higher content of gypenoside L (1.8% w/w), gypenoside LI (1.4% w/w), and ginsenoside Rg3 (0.15% w/w) (Lee et al., 2019b). While HFD-fed mice showed significant clinical effects such as increases in body weight, fat mass, white adipose tissue, and adipocyte hypertrophy as compared to the control group, the GPE-treated group failed to show them. GPE treatment also reduced serum levels of triglyceride, total cholesterol, and LDL-cholesterol, without affecting HDL-cholesterol. Mechanistically, the clinically observed GPE effects appeared due to increased AMPK activation and suppressed adipogenesis by decreasing the mRNA levels of CCAAT/enhancer binding protein-α (C/EBPα), PPARγ, SREBP-1c, PPARγ coactivator-1α, fatty acid synthase, adipocyte protein 2, and sirtuin 1, and increased levels of carnitine palmitoyltransferase and hormone-sensitive lipase (Lee et al., 2019b).
G. pentaphyllum to the Clinic
In general, there are few human trials that addressed G. pentaphyllum extract therapeutic effects or safety. A search on clinicaltrials.gov shows that there are only four studies that use G. pentaphyllum extracts. One of the studies is on obese patients and three are on diabetes mellitus patients.
Actiponin, an extract of G. pentaphyllum, is a dietary supplement used for weight loss in obese individuals. During an interventional study, 80 randomized Korean participants took part in a double blind, parallel study for 12 weeks where the experimental group took Actiponin at a dose of 450 mg/day. The experimental group lost weight with no adverse effects, as compared to the placebo group (Park S. et al., 2014).
In another study, 1 mg/kg of the water extract of G. pentaphyllum was given to 44 patients with CVDs and 56 healthy individuals and the platelet aggregation was studied. It was revealed that the water extract elicited significant inhibition of the aggregation of platelets. This means that there is potential for the use of this supplement to prevent cardio-cerebrovascular diseases while being cautious not to give these supplements to patients who suffer from low platelet count or bleeding disorders (Juan and Shanzhang, 1995).
Some studies suggest there is a link between anxiety disorders and an increased risk of developing a CVD (Fan et al., 2008; Vogelzangs et al., 2010; Seldenrijk et al., 2011; Batelaan et al., 2014). For this reason, it is of interest to reduce anxiety in order to decrease the risk of developing CVD in predisposed individuals. It was shown that G. pentaphyllum ethanol extract had anti-anxiety effects on mice exposed to chronic stress (Choi H. et al., 2013). This finding was replicated in a double-blind, placebo-controlled clinical trial that had 72 healthy Korean individuals under chronic stressful conditions. Thirty-six participants were given 200 mg of G. pentaphyllum ethanol extract, twice a day for 8 weeks. The supplementation reduced the experimental group anxiety without any adverse drug effects suggesting its potential as a safe anti-anxiety supplement (Choi et al., 2019).

T2DM: Type 2 Diabetes Mellitus.

Although the proven pharmacological effects of G. pentaphyllum in in vitro studies and in vivo animal studies may not necessarily translate well into efficacy human subjects, there are positive studies. For instance, a set of studies conducted in T2DM patients supplemented with G. pentaphyllum tea showed improvements in insulin sensitivity and glycemia with no adverse side effects (Huyen et al., 2010; Huyen et al., 2013). In one of the trials, 24 drug-naïve T2DM patients were randomized to take either 6 g daily of Gynostemma pentaphyllum tea or placebo tea, during 1- week period. The authors measured FPG, insulin levels, and HbA1c levels before, during, and after the treatment. The study showed a prompt improvement of glycemia and insulin sensitivity, and suggested that Gynostemma pentaphyllum tea could be an effective, and safe approach to treat T2DM patients (Huyen et al., 2010). The same authors followed up with another study that used the same study design but enrolled only 16 drug-naïve T2DM patients. The authors measured the same parameters and came to the same conclusion that Gynostemma pentaphyllum tea exerted antidiabetic effects by improving insulin sensitivity (Huyen et al., 2013). These results were confirmed by another study where G. pentaphyllum was used together with sulfonylurea (Huyen et al., 2012). Thus far, the current data indicate that G. pentaphyllum is quite efficient at improving insulin sensitivity and blood sugar levels if administered solely and that its efficacy may be enhanced when combined with other medications.
Safety, Toxicity, and Side Effects of G. pentaphyllum
In a study evaluating the toxicity of G. pentaphyllum extract on female Sprague-Dawley rats, a single dose of up to 5000 mg/kg of the extract was given and subchronic toxicity tests were performed with 1000 mg/kg/day for 90 days. No rat death occurred nor did any signs of toxicity arise. Blood chemistry values, though statistically different from the control group, were within normal ranges in rats. Thus, no mortality nor abnormalities have risen from the G. pentaphyllum extract treatment (Chiranthanut et al., 2013). In addition, no toxicity or mortality was reported upon long-term administration of a dose up to 750 mg/kg body weight of G. pentaphyllum in rats (Attawish et al., 2004).
A Phase I clinical trial was conducted to evaluate the safety of G. pentaphyllum whereby three groups of healthy volunteers were administered 50, 200, and 400 mg twice daily with water extract of G. pentaphyllum for two months. No major immune adverse events such as significant changes in natural killer cell activities, number of CD3+, CD4+, and CD8+ cells, were reported. No biochemical parameters were significantly affected either. Such doses of G. pentaphyllum were deemed to be safe (Chavalittumrong et al., 2007). In another clinical trial, 537 bronchitic patients were treated three times a day with G. pentaphyllum (2.5–3 g, prepared as tablets or capsules). Adverse side effects that included vomiting, abdomen tension, diarrhea (or constipation), dizziness, blurred vision, and tinnitus effects were seen in a small number of patients. Notably, these symptoms were mild and did not stop the patients from taking the G. pentaphyllum extract (Razmovski-Naumovski et al., 2005). A very recent randomized, double-blind, placebo-controlled clinical trial utilizing G. pentaphyllum extract in 72 healthy adults revealed no adverse side effects of the ingestion of the ethanolic extract of G. pentaphyllum (Choi et al., 2019). Overall, consumption of G. pentaphyllum seems to be safe at the doses required to observe a therapeutic effect.

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Figure 6
Herbal therapies in the context of CVDs. Herbal preparations can exert protective effects by ameliorating the pathological effects exerted by CVDs risk factors. The herbal extracts can attenuate endothelial dysfunction and/or VSMC alterations by acting as, vasodilators, ROS scavengers, anti-oxidants, anti-inflammatory, anti-apoptotic, anti-hypertrophic, or anti-proliferative agents. This achieved through mechanisms that act in ECs only, VSMCs only, or through overlapping mechanism that act in both ECs and VSMCs. In ECs, herbal preparations can increase NO availability, decrease mitochondrial dysfunction and/or metabolic abnormalities as well as enhance angiogenesis. This can decrease the incidence of atherosclerosis and hypertension, which in return can decrease the risk of CVDs development. In VSMCs, the herbal extracts can modulate ECM deposition as well as cell migration, proliferation, and cell shape changes. VSMC, vascular smooth muscle cell; ECM, extracellular matrix; EC, endothelial cell; NO, nitric oxide; PPARY, peroxisome proliferator-activated receptor-gamma.

Conflict of Interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest

Anti-cancer effects

(2012), Li et al extracted 2 acidic polysaccharides and tested against cancer cells in vivo and in vitro22. After using alcohol to remove lipid they used water at 90°C for 2 hours, x 3, to extract from dried stem and leaf material.

Cancer types: Human chronic myeloid leukemia K-562, breast adenocarcinoma MCF-7, colon adenocarcinoma HT-29, hepatocellular carcinoma HepG2 and mouse melanoma B16 cell lines.

For in vivo studies they used tumor bearing mice and melanoma injected male rats, feeding them or injecting the polysaccharides.

Abstract
Two acidic polysaccharides (GP-B1 and GP-C1) were obtained from Gynostemma pentaphyllum. The molecular weights (Mw) of the two fractions were 79 kDa for GP-B1 and 126 kDa for GP-C1. GP-B1 was composed of Gal, Ara, Man, Rha, Xyl, Glc, GalA and GlcA in a molar ration of 3.5:3.2:0.6:0.9:0.3:0.5:0.6:0.4. GP-C1 consisted of Gal, Ara, Man, Rha, Glc, and GlcA in the proportions of 2.1:1.0:0.3:0.5:0.4:0.9. Among them, GP-B1 treatment had a significant inhibitory effect on the growth of melanoma B16 in vivo and in vitro. Meanwhile GP-B1 could increase the relative spleen weight and stimulate the splenocyte proliferation alone or combined with ConA. Moreover, GP-B1 treatment induced an evident increase in the level of serum TNF-α, IFN-γ, and IL-12 and a reduction for IL-10 production. These results indicate that the antitumor effects of GP-B1 are associated with immunostimulation.

Low cytotoxicity against normal mouse bone marrow cells:

Fig. 1. Inhibitory effect of acidic polysaccharide GP-B1 on the proliferation of normal mouse bone marrow cells. Data are presented as mean ± S.D (n = 3).

As shown in Table 3, the growth of melanoma B16 tumor in the model mice was significantly suppressed by GP-B1 with the inhibition rate of 46.9% and 62.5% at the dose of 50 and 100 mg/kg body weight, respectively. Meanwhile, tumor delay time of CTX and two GP-B1 treated groups had a noticeable prolong than that of model control (P < 0.01 or P < 0.001.). Supporting this observation, GP-B1 treatment decreased the average size of tumor volume by about 58% and 76% compared with that of negative control (Fig. 2). As expected, the positive control drug CTX obviously inhibited tumor growth and simultaneously decreased the spleen weight compared to the negative control group (P < 0.01). It is notable that the spleen index of the mice treated with the GP-B1 are higher than those of the model mice treated with or without CTX, which is close to the normal control group, suggesting that GP-B1 treatment caused no damage to the immunological function in tumor-bearing mice.

Fig. 2. Inhibitory effect of acidic polysaccharide GP-B1 on B16-induced melanoma tumor growth in C57BL/6 mice. Data are presented as mean ± S.D. values based on 10 mice in each group.

Concanavalin A (ConA) is a well-known T cell mitogen that can activate the immune system, recruit lymphocytes and elicit cytokine production. In addition to its mitogenic activity, ConA can induce programmed cell death via mitochondria-mediated apoptosis and autophagy.23

The results showed that GP-B1 could corporate [sic] with ConA to significantly augment ConA-induced lymphocyte proliferation in vitro. Simultaneously GP-B1 itself also exhibited a moderate stimulation effect on lymphocytes proliferation. However, ConA-stimulated splenocyte proliferations in the CTX-treated group were significantly lower than those of the model control (P < 0.05)

Higher is better as evidence of immunomodulation:

Fig. 3. Effect of acidic polysaccharide GP-B1 on lymphocyte proliferation in melanoma-B16-bearing mice. Data are presented as mean ± S.D. values based on 10 mice in each group. Significant differences compared to the negative control were evaluated using Student's t test: *P < 0.05 and **P < 0.01.

…we investigated the effect of GP-B1 on serum TNF-α, IFN-γ, IL-10 and IL-12 production in melanoma-B16-bearing mice by MTT assay. As shown in Fig. 4, the levels of TNF-α, IFN-γ, and IL-12 were significantly decreased in CTX-treated mice as compared to those of the model control (P < 0.05). However, GP-B1 markedly promoted TNF-α, IFN-γ and IL-12 production in serum of melanoma-B16-bearing mice at two doses (P < 0.05). In addition, IL-10, an immunosuppressive cytokine, decreased significantly after administered with GP-B1 compared with that of negative control (P < 0.05) at the dose of 100 mg/kg.

Fig. 4. Effect of acidic polysaccharide GP-B1 on serum TNF-α, IFN-γ, IL-10 and IL-12 production in melanoma-B16-bearing mice. Data are presented as mean ± S.D. values based on 10 mice in each group. Significant differences compared to the negative control were evaluated using Student's t test: *P < 0.05.

4. Conclusion
Previous reports had documented that G. pentaphyllum crude extract or water-soluble neutral polysaccharide has anticancer effect. In this investigation, one neutral polysaccharide (GP-I) and two acidic polysaccharides (GP-B1 and GP-C1) had been achieved by ion-exchange and gel-filtration chromatography. MTT assay proved that GP-B1 had a potent anti-proliferation activity on melanoma B16 cells, with the lowest IC50 value of 65.4 μg/ml and had no toxicity on the normal mouse bone marrow cells, even at the high dose of 400 μg/ml, indicating its potential application value in the therapy for cancer. Furthermore GP-B1 administrated to melanoma B16-bearing mice could obviously inhibit the tumor growth and improve the immune organ status.

In conclusion, GP-B1, an acidic polysaccharide, from G. pentaphyllum could not only significantly inhibit the growth of tumor in vitro and in vivo, but also remarkably increase splenocytes proliferation and the level of serum TNF-α, IFN-γ, IL-10 and IL-12 in tumor-bearing mice, which indicated that the GP-B1 could improve cellular immune response. The above results suggested that the antitumor activity of this acidic polysaccharide might be achieved by improving immune response, and GP-B1 could act as antitumor agent with immunomodulatory activity.

No conflict of interest statement was provided.

In 2010, Peng, Zhou and Zhang investigated the antitumor activities of dammarane triterpene saponins from a different species, Bacopa monniera.

I reference this study due to the commonality of the dammaranes with G. pentaphyllum.

This herb is also known as water hyssop, waterhyssop, brahmi, thyme-leafed gratiola, herb of grace, and Indian pennywort. Their in vivo studies in mice demonstrated tumor inhibition of up to 90%:24

Abstract
Bioassay-guided methods were used to test the antitumor activity of methanol extract of the whole plant of Bacopa monniera (L.) Wettst. and four different fractions (petroleum ether, CHCl3, EtOAc, and n-BuOH fractions) of the methanol extract. Among the five crude samples, n-BuOH fraction was noted to have the highest antitumor activity. The dammarane triterpene saponins isolated from n-BuOH fraction, bacopaside É (1) and bacopaside VII (3), had potential antitumor effect. 1 and 3 showed cytotoxicity of all the tested human tumor cell lines MDA-MB-231, SHG-44, HCT-8, A-549 and PC-3M in MTT assay in vitro, and showed 90.52 % and 84.13 % inhibition in mouse implanted with sarcoma S180 in vivo at the concentration of 50 μmol/kg, respectively. The remaining two compounds, bacopaside II (2) and bacopasaponin C (4) were found to be much less potent compared with 1 and 3. 1 and 3 significantly inhibited human breast cancer cell line MDA-MB-231 adhesion, migration and Matrigel invasion in vitro at the concentration of 50 μmol/L. Since no antitumor activities about the monomers from Bacopa monniera (L.) Wettst. have been reported, these results indicate that the mechanism of action of 1 and 3 needs further study.

In contrast, the FDA disagrees with any health claims made for dietary supplements made from B. monniera and declared them to be unproven and illegal.25

Research and regulation
Bacopa monnieri is used in Ayurvedic traditional medicine to improve memory and to treat various ailments. Reviews of preliminary research found that Bacopa monnieri may improve cognition, although the effect was measurable only after several weeks of use.
In 2019, the FDA issued warning letters to manufacturers of dietary supplements containing Bacopa monnieri that advertised health claims for treating or preventing stomach disease, Alzheimer's disease, hypoglycemia, blood pressure, and anxiety were unproven and illegal. The FDA stated that Bacopa monnieri products have not been approved for these or any medical purposes.

Moving on to 2016 and Li et al performed an in vitro study into the anticancer activity of a nonpolar fraction from G. pentaphyllum.26

“A nonpolar molecule has no separation of charge, so no positive or negative poles are formed. In other words, the electrical charges of nonpolar molecules are evenly distributed across the molecule. Nonpolar molecules tend to dissolve well in nonpolar solvents, which are frequently organic solvents.

In a polar molecule, one side of the molecule has a positive electrical charge and the other side has a negative electrical charge. Polar molecules tend to dissolve well in water and other polar solvents."2728

Abstract
Gynostemma pentaphyllum (Thunb.) Makino (GpM) has been widely used in traditional Chinese medicine (TCM) for the treatment of various diseases including cancer. Most previous studies have focused primarily on polar fractions of GpM for anticancer activities. In this study, a nonpolar fraction EA1.3A from GpM showed potent growth inhibitory activities against four cancer cell lines with IC50 ranging from 31.62 μg/mL to 38.02 μg/mL. Furthermore, EA1.3A also inhibited the growth of breast cancer cell MDA-MB-453 time-dependently, as well as its colony formation ability. EA1.3A induced apoptosis on MDA-MB-453 cells both dose-dependently and time-dependently as analyzed by flow cytometry and verified by western blotting analysis of apoptosis marker cleaved nuclear poly(ADP-ribose) polymerase (cPARP). Additionally, EA1.3A induced cell cycle arrest in G0/G1 phase. Chemical components analysis of EA1.3A by GC-MS revealed that this nonpolar fraction from GpM contains 10 compounds including four alkaloids, three organic esters, two terpenes, and one catechol substance, and all these compounds have not been reported in GpM. In summary, the nonpolar fraction EA1.3A from GpM inhibited cancer cell growth through induction of apoptosis and regulation of cell cycle progression. Our study shed light on new chemical bases for the anticancer activities of GpM and feasibilities to develop new anticancer agents from this widely used medicinal plant.

More than 210 compounds have been identified from GpM so far. Based on their chemical structures, these compounds could be grouped into saponin, sterol, flavonoid, and polysaccharide. Among these groups, saponin (named gypenoside, Gyp) is the largest family containing about 170 members and is reported as the major active ingredients of GpM. Other components of GpM, like flavonoids and polysaccharides, also exhibit various biological activities, while sterols are rarely reported.
In most phytochemical studies of GpM, the water soluble extracts, the ethanol or methanol extracts, or fractions eluted with polar solvent were focused on. Few studies regarding the nonpolar fractions of GpM have been carried out. Piao et al. extracted GpM with ethyl acetate, but, in the following steps of elution and isolation, polar solvents like ethanol, methanol, and water were applied, which indicated that the final compounds or ingredients were still polar.
To date, most of the reported anticancer components from GpM are polar fractions, like Gyps, flavonoids, and polysaccharides. In this work, we focused on the nonpolar fraction of GpM. After extracting with ethyl acetate, nonpolar solvents like n-hexane and trichloromethane were used as the elution solvents at the chromatographic separation step. The obtained fraction (EA1.3A) was thus nonpolar and showed potent anticancer effects in vitro. And chemical characterization of EA1.3A by GC-MS analysis found that compounds have not been reported in GpM previously which may contribute to its anticancer activities.

Figure 1
EA1.3A inhibits proliferation of cancer cell lines. (a) Dose effect of EA1.3A treatment (72 hours) on the proliferation of two breast cancer cell lines (MCF7 and MDA-MB-453), one colon cancer cell line (HCT116), one prostate cancer cell line (LNCaP), and one normal human lung fibroblast cell line (CCD-19Lu). The cell number at each EA1.3A concentration is represented as a percentage of control. Average values are from three independent experiments performed in duplicate (). (b) Time course of EA1.3A treatment (50 μg/mL) on the proliferation of breast cancer cell line MDA-MB-453. The cell number at each time point is represented as a percentage of control. Average values are from three independent experiments performed in duplicate (). Data are shown as mean ± SD.
Consistent with the dose effect of EA1.3A on cancer cell growth (Figure 1(a)), when treated with 50 μg/mL of EA1.3A, the growth of MDA-MB-453 cells was also inhibited in a time-dependent manner. After treatment for 24 hours, the relative growth rate of MDA-MB-453 cells was 88.34% and declined to 35.12% after 72 hours (Figure 1(b)). So EA1.3A inhibited the growth of breast cancer cell MDA-MB-453 both dose- and time-dependently. This was further confirmed by colony formation assay (Figure 2). Colony formation ability of MDA-MB-453 cell was significantly decreased after treatment with EA1.3A at 6.25 μg/mL and almost abolished at 50 μg/mL.

Figure 2
Colony formation of MDA-MB-453 cells with EA1.3A treatment. (a) Representative colony formation assay plates of MDA-MB-453 cells treated with EA1.3A at indicated concentrations for 12 days. (b) Quantification of colony number (). Data are shown as mean ± SD. values determined by Student’s t-test. *P<0.001; **P<0.0001.

4. Discussion
To date, most reported components of GpM with anticancer activities are polar fractions/compounds, like Gyps, flavonoids, and polysaccharides. Here, for the first time, we showed that the nonpolar fraction EA1.3A from GpM also has potent anticancer activities through regulation of cell cycle progression and induction of apoptosis. EA1.3A even exhibited greater anticancer activities than other fractions (data not shown) with IC50 values ranging from 31.62 μg/mL to 38.02 μg/mL against 4 cancer cell lines tested in this study which are also comparable with or more potent than reported polar fractions from GpM. For example, the IC50 values of Gyps were of 39.3 μg/mL on PC-3 cells [23], 112.39 μg/mL on Eca-109 cells, and 61.68 μg/mL on SW620 cells [36], the IC50 value of flavonoids was of 33.3 μg/mL on PC-3 cells [23], and the IC50 value of polysaccharide was of 65.4 μg/mL on B16 cells [26].
GC-MS analysis identified 10 compounds in EA1.3A and all of them have not been reported in GpM. It was not surprising to identify three organic esters, which are usually nonpolar, in fraction EA1.3A from GpM. The major component of EA1.3A was propanoic acid, 2-methylpropyl ester (isobutyl propionate), which is widely used in food and beverage industries as a rum flavor. Although there is no anticancer activity reported for this compound, the parent acid propionate inhibits cancer cell proliferation through multiple mechanisms. Alkaloids are widely distributed in plants characterized with basic nitrogen atoms and have a wide range of pharmacological activities. Although few alkaloids have been isolated from GpM, in the present study, we identified four alkaloids in EA1.3A. One of them, 2-methyl-7-phenylindole, has been found present in a variety of plants, and extract containing 2-methyl-7-phenylindole showed cytotoxicity effects. Two terpenes were identified in EA1.3A. One of them, β-carotene-3,3′-diol,5,8-epoxy-5,8-dihydro,(3S,3′R,5R,8S)- belongs to xanthophylls, one kind of carotenoids, which are widely presented in the leaves of most green plants. Xanthophylls are known for their antioxidant activities and have been found to reduce oxidative stress and prevent tumorigenesis. Phytol, a constituent of chlorophyll, is a precursor for the manufacture of synthetic forms of vitamin E and vitamin K1. Extensive studies have proven anticancer activities of phytol. The catechol substance, 3,5-di-tert-butylcatechol, was identified to be an effective inhibitor of the enzyme sarco/endoplasmic reticulum calcium ATPase (SERCA), a potential target for cancer chemotherapy, by virtual screens and confirmed by bioassays. These compounds may contribute to the anticancer activities of nonpolar fraction EA1/3A from GpM.
5. Conclusions
In summary, the rarely studied nonpolar fraction EA1/3A from GpM showed potent anticancer activities via induction of cell cycle arrest and apoptosis. Our study shed light on new chemical bases for the anticancer activities of this widely used medicinal herb. Further studies to isolate and identify pure compounds from this nonpolar fraction from GpM and evaluate the anticancer activities as well as the mechanisms of action of these compounds are suggested for developing novel anticancer agents.
Conflict of Interests
The authors confirm that this paper content has no conflict of interests.

Also from 2016, Li et al published a comprehensive review of the literature associated with G. pentaphyllum (GpM) and anti-cancer activities and mechanisms of action:29

Various extracts and fractions of GpM, as well as numerous pure compounds isolated from this herb exhibited inhibitory activity towards the proliferation of cancer cells in vitro and in vivo. Furthermore, the results of several clinical studies have shown that GpM formula could have potential curative effects on cancer. Multiple mechanisms of action have been proposed regarding the anti-cancer activities of GpM, including cell cycle arrest, apoptosis, inhibition of invasion and metastasis, inhibition of glycolysis and immunomodulating activities.

Gynostemma pentaphyllum (Thunb.) Makino (GpM) (Jiaogulan) has been widely used in Chinese medicine for the treatment of various diseases, including hepatitis, diabetes and cardiovascular disease. Modern medical research has shown that GpM exhibits a variety of pharmacological properties, including anti-inflammatory, antioxidative, lipid metabolism regulatory, antiproliferative, neuroprotective, and anxiolytic activities. GpM has consequently been widely used for the treatment of hepatitis, diabetes, cardiovascular disease and cancer. GpM is also widely used as a health supplement in beverages, biscuits, noodles, face washes and bath oils.

In vitro anti-cancer activities of GpM
The in vitro antiproliferative activities of some of the pure compounds and extracts isolated from GpM have been widely reported and the details of these materials are summarized in Table 3. Shi et al. obtained four dammarane-type triterpene saponins (compounds 3–6) from the aerial parts of GpM, which exhibited moderate cytotoxic activities in vitro against several human cancer cell lines, including HL-60 (human promyelocytic leukemia cells), Colon 205 (human colon cancer cells) and Du145 (human prostate carcinoma cells) cells. Yin et al. isolated nine dammarane saponins from the methanol extract of the aerial part of GpM, and found that compounds 7, 8 and 9 exhibited inhibitory activities towards the growth of SGC-7901 (stomach cancer cells) and BEL-74020 (hepatocellular carcinoma cells) at a concentration of 100 μM with percentage inhibition values of 21, 93 and 8 %, and 77, 92 and 40 %, respectively.
Almost all of the compounds and extracts isolated from GpM to date have be reported to exhibit noticeable antiproliferative activities with IC50 values ranging from 0.05 to 74.3 μg/mL (Table 3). Compound 16 exhibited potent antiproliferative activities against A549 human lung cancer cells and U87 glioblastoma cells with IC50 values of 0.05 and 0.25 μg/mL, respectively. Compound 15 showed antiproliferative activity against MDA-MB-435 human breast cancer cells with an IC50 value of 3.90 μg/mL, whereas the carotenoid fraction of GpM exhibited the strongest activities of all of the reported extracts with an IC50 value of 1.6 μg/mL against Hep3B human hepatocellular carcinoma cells.
The hydrolysates of the extracts of GpM have also been reported to exhibit anti-cancer activities, together with several other derivatives of the natural products found in GpM. For example, Chen et al. reported the synthesis of four sulfated derivatives of GPP2, which is a native polysaccharide isolated from GpM. One of the sulfated derivatives prepared by Chen (GPP2-s4) inhibited the growth of HepG2 human hepatocellular carcinoma cells by 46.4 ± 2.8 % at a concentration of 2000 μg/mL. Compared with GPP2, all four sulfated derivatives exhibited stronger antiproliferative activities against HeLa cervical cancer cells at concentrations as low as 100 μg/mL. GP-B1, which is an acidic polysaccharide derived from GpM, significantly inhibited the growth of B16 melanoma cells with an IC50 of 65.4 μg/mL with very little cytotoxicity against normal cells. Moreover, GP-B1 not only significantly inhibited the growth of cancer cells, but also improved cellular immune response by increasing levels of tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-10 (IL-10) and interleukin-12 (IL-12) observed in the serum of melanoma-B16-bearing mice.
In vivo anti-cancer activities of GpM
The in vivo anti-cancer activities of GpM are summarized in Table 4. Gyps led to significant reductions in the size of solid tumors in nude mice injected with SAS oral cancer cells. Gyps also promoted the survival of mice xenografted with WEHI-3 leukemia cells, which was accompanied by an increase in the number of megakaryocytes and reduced spleen weight in these animals, indicating an enhanced immune response. Similar anti-cancer activities have also been reported for Gyps in another leukemia mouse model. The intraperitoneal treatment of tumor-bearing mice with Gyps (5 or 20 mg/kg/day) for 4 weeks led to considerable decreases in the size and weight of their tumors without altering their body weight. Gyps also strongly suppressed tumor growth in mice bearing advanced S180 sarcoma, which was associated with an increase in the ratio of tumor necrosis area to tumor total area and lymphocyte/macrophage infiltration into the peripheral areas of tumors. This effect also led to an increase in the weight of the spleens of these animals, as well as increases in the quantity and size of their splenic white pulp. Gyps enhanced the anti-cancer effects of 5-fluorouracil in colorectal cancer cells and xenografts. Gyps have also been reported to inhibit tumorigenesis in a transgenic mouse models of cancer, such as the ApcMin/+ mouse model of intestinal neoplasia. Moreover, rats fed with a standardized extract of GpM did not show any mortal or toxic effects, highlighting the good safety profile of this material.

A polysaccharide from GpM inhibited the development of transplanted S180 sarcoma in a dose-dependent manner and increased the phagocytosis of macrophages, as well as increasing the production of NO, IL-1β and TNF-α from the peritoneal macrophages. The neutral polysaccharide fraction CGPP inhibited the growth of H22 hepatocarcinoma cells transplanted into ICR mice [20]. CGPP treatment also led to improvements in the body weight, spleen/thymus index and degree of splenocyte proliferation in tumor-bearing mice. Furthermore, CGPP treatment led to considerable increases in the levels of cytokines, such as IL-2, TNF-α and IFN-γ in tumor-bearing mice, as well as increases in the activity of natural killer (NK) cells and cytotoxic T lymphocytes (CTL). The tumor inhibitory and immunoregulatory effects of CGPP greatly increased the life span of H22 ascites in tumor-bearing mice.

Something not considered until now is perhaps the most important: clinical studies of patients with advanced malignant tumors who were treated with extracts of G. pentaphyllum:

Clinical anti-cancer studies on GpM
A clinical study was conducted in 1993 involving 59 patients with advanced malignant tumors to assess the effects of GpM. The results revealed that patient treated with a GpM formula showed cancer relapse and metastasis rates of 11.9 and 8.5 %, respectively, compared with values of 72.4 and 55.2 % in the control group. The results of this study also revealed that the T lymphocyte transformation rate and acid α-naphthyl acetate esterase (ANAE+) activity increased by 8.2 % following GpM treatment. The results of a separate 5-year observational study also showed that the treatment of cancer patients with GpM formula led to significant reductions in cancer relapse and metastasis rates, as well as reduced mortality and improved immune function in these patients. GpM has also been reported to enhance NK cell activity in breast cancer patients, and improve the immune function of cancer patients after chemotherapy, as demonstrated by increased T lymphocyte transformation rate and decreased IgG and IgM levels. Furthermore, GpM enhanced the immunological function of lung cancer patients after chemotherapy. The results of a recent study demonstrated that GpM formula can work in synergy with chemotherapy reagents. The clinical uses of GpM are summarized in Table 5
Mechanisms of action
Multiple mechanisms of action have been proposed regarding the anti-cancer activities of GpM, including cell cycle arrest, apoptosis induction, inhibition of invasion and metastasis, glycolysis inhibition and immunomodulation (Fig. 2).
Fig. 2
Mechanisms of action for the anti-cancer activities of GpM

Perspectives
Numerous studies have been published during the last four decades regarding the anti-cancer effects of GpM, including reports focused on (i) the isolation and characterization of its chemical components; (ii) the evaluation of its anti-cancer activities and mechanisms of action; and (iii) studies on its toxicity. Taken together, the results of these reports have demonstrated that GpM has a broad anti-cancer spectrum (against 30 cancer cell lines, Table 3) without any obvious inhibitory effect on normal cell proliferation. However, there are limitations associated with most of these studies.

The authors then suggest that for experimental and clinical consistency the standard preparation of Gyps needs to be unified, and experimental systems need to be closer to the clinical settings, eg by using genetically engineered mouse models instead of using immunodeficient mice.

Conclusion
In summary, GpM has been investigated extensively as a potent anti-cancer agent against many types of cancers both in vitro and in vivo. The general consensus from the literature is that GpM exerts its anti-cancer activities through multiple mechanisms, including cell cycle arrest, the induction of apoptosis, inhibition of invasion and metastasis, glycolysis inhibition and immunomodulation.

Competing interests
The authors declare that they have no competing interests.

The most recent paper in this review is from 2021, by Liu et al. They investigated how gypenosides of G. pentaphyllum can induce apoptosis of renal cancer tells through decreasing the phosphorylation level of Akt and mTOR in the previously discussed PI3K/Akt/mTOR signaling pathway.30

Protein phosphorylation is an important cellular regulatory mechanism as many enzymes and receptors are activated/deactivated by phosphorylation and dephosphorylation events, by means of kinases and phosphatases.31

Abstract
Ethnopharmacological relevance: Gynostemma pentaphyllum (Thunb.) Makino is a traditional medicine commonly used in China, East Asia and Southeast Asia. In clinic, it is mainly used for hyperlipidemia and antitumor. Its antitumor activity was first recorded in "Illustrated Catalogue of Plants". Gypenosides were the main active ingredients of G. pentaphyllum. The anticancer activity of gypenosides in vivo and in vitro had been widely reported. However, the mechanism of gypenosides in renal cell carcinoma (RCC) still unclear.
Aim of the study: In this study, we tried to investigate the active constituents from G. pentaphyllum and potential mechanisms in RCC treatment through network pharmacology and in vitro experiments.
Material/methods: Active compounds and their targets were evaluated and screened through TCMSP and Swiss Target Prediction database. Notably, nine preliminary screened components obtained from database were identified by LC-MS and LC-MS/MS. The targets associated with RCC were obtained from OMIM, TTD and GeneCards database. The PPI network and active component/target/pathway networks were constructed to identify the potential drug targets using String database and Cytoscape software. The functions and pathways of targets were analyzed through DAVID database. Finally, AutoDockTools 1.5.6 was used for molecular docking to assess the binding ability between compounds and targets. To support our prediction, we then explore the antitumor effect and mechanism of gypenosides by vitro experiments. CCK8 and flow cytometry were performed to evaluate cell death treated with gypenosides. Quantitative real-time PCR and Western blot were conducted to detect the changes of PI3K/AKT/mTOR signaling pathway.
Results: Nine saponins and 68 targets have been screened. The hub targets covered PIK3CA, VEGFA, STAT3, JAK2, CCND1 and MAPK3. Enrichment analysis showed that the pathways mainly contained PI3K/Akt/mTOR, HIF-1, TNF, JAK-STAT and MAPK signaling pathways. Gypenosides extracted from G. pentaphyllum showed strong activity against 786-O and Caki-1 cells, and cell apoptosis were detected through Annexin V/PI dual staining assay. RT-qPCR showed that gypenosides downregulated the levels of PIK3CA, Akt and mTOR in Caki-1 and 786-O cells. Mechanistically, gypenosides induced apoptosis of RCC cells through regulating PI3K/Akt/mTOR signaling pathway which was implemented though decreasing the phosphorylation level of Akt and mTOR.
Conclusions: Gypenosides induced apoptosis of RCC cells by modulating PI3K/Akt/mTOR signaling pathway.
Keywords: Gypenosides; Molecular docking; Network pharmacology; PI3K/Akt/mTOR pathway; Renal cell carcinoma.

Fig. 1. The detailed flow chart of the current study.

DMSO was used as a control.

3.9. Gypenoside regulated PI3K/Akt/mTOR pathway in RCC cells
Combined with the core targets and pathway prediction of network pharmacology, the apoptosis-inducing effect of gypenosides in RCC may be related to the PI3K/Akt/mTOR signaling. To test, we conducted RT-qPCR to ascertain the effects of gypenosides on this pathway. As shown in (Fig. 7a), gypenoside inhibited the mRNA levels of Akt, mTOR and PIK3CA of 786-O and Caki-1. We further determined the level of Akt, P-Akt, mTOR, P-mTOR using western bloting and observed that inhibited phosphorylation of Akt/mTOR and Akt in both 786-O and Caki-1 cells, the protein level of mTOR while upregulating in Caki-1. Similar results were observed in Western blot analysis, where gypenoside decreased expression of mTOR in 786-O as well as increased expression of mTOR in Caki-1 cells (Fig. 7b). These findings indicate that gypenosides may induce RCC apoptosis by inhibiting the PI3K/Akt/mTOR pathway.
Fig. 7. Gypenosides influenced the expression of relevant proteins. (a) mRNA expression of Akt, mTOR, PIK3CA in 786-O and Caki-1 cells after treated with gypenosides was measured with RT-qPCR, *p < 0.05. (b) Protein expression of Akt, mTOR, P-Akt, P-mTOR in 786-O and Caki-1 cells after treated with gypenosides. The histograms show quantified results of protein levels, which are adjust with corresponding ACTB protein level and expressed as folds of control (mean ± SD, n = 3). Differences between the treatment group and the control group are determined by one way AVOVA followed by post-hoc Dunnett's test, *p < 0.05.

While on the other side, although Chinese herbal medicine and its natural products provide new ideas for cancer treatment, the complexity of the compositions and the specific target genes or target proteins used for drug action are not clear. In the past, drug development mainly followed the “one drug, one gene, one disease” model (Casas et al., 2019), which was the main reason for the failure of 70% of new drugs in clinical trials. In fact, a variety of clinical chronic diseases such as tumors, cardiovascular and cerebrovascular diseases are multi-gene, multi-factorial diseases, and it is difficult to achieve significant therapeutic effects based on a single target. Therefore, the relationship between multiple targets and complex diseases has become a new model of drug development. In the present study, network pharmacology provides a research concept for drug development of gypenosides for the treatment of RCC.
In the research and development of natural medicines, the complexity of ingredients and the blindness of separation and extraction significantly increase the cost of antitumor drugs research and development. Additionally, The ADME/T properties of compounds play an important role in innovative drug transformation research. Therefore, due to the lack of ADME, effective drugs screened in vitro may not achieve the expected effects of in vivo. Because the complex composition of G. pentaphyllum, the ADME screening method was used in this study, to ensure more accurate and faster screening of active saponins.

The activation of PI3K/Akt/mTOR signal transduction in kidney cancer is closely related to poor prognosis.

Mechanistically, the mRNA level of PIK3CA, Akt and mTOR was Significantly lower in 786-O and Caki-1 cells treated with gypenosides than DMSO using RT-qPCR. Further, phosphorylation of Akt and mTOR expression was remarkably decreased by gypenosides treatment in cell models. Curiously, we did not find that the level of p-Akt protein in Caki-1 treated with gypenoside was reduced. A possible explanation for this fact was that PI3K may not be the only way to activate Akt.

5. Conclusion
In conclusion, nine active saponins of G. pentaphyllum could act on many targets and signaling pathways related to RCC by using the method of network pharmacological combined with molecular docking. Additionally, based on the results of network pharmacology predictions, we next explored how gypenosides can play an antitumor effect. This study proved the inhibitory effect of gypenosides on renal cancer cells. Mechanistically, gypenosides may induce apoptosis by reducing the phosphorylation of AKT and mTOR. The network pharmacological method developed and the experimental evaluations in this study pointed to a new window for laboratory research and clinical application of gypenosides for the treatment of RCC. Thus, gypenosides may be a natural resource of anticancer drug targeting the PI3K/Akt/mTOR pathway.

Declaration of competing interest
The authors declare no competing financial interests.

Neurological and other effects

In 2010, Choi et al reported neuroprotective effects of ethanol extracts from G. pentaphyllum in a rat model of Parkinson’s disease:32

…an oral administration of herbal ethanol extracts from Gynostemma pentaphyllum (GP-EX) (10 mg/kg and 30 mg/kg) starting on day 3 post-lesion for 28 days markedly ameliorated the reduction of TH-immunopositive neurons induced by 6-hydroxydopamine-lesioned rat brain from 40.2% to 67.4% and 75.8% in the substantia nigra. GP-EX administration (10 and 30 mg/kg) also recovered the levels of dopamine, 3,4-dihydroxyphenylacetic acid, homovanillic acid and norepinephrine in post-lesion striatum to 64.1% and 65.0%, 77.9% and 89.7%, 82.6% and 90.2%, and 88.1% and 89.2% of the control group. GP-EX at the given doses did not produce any sign of toxicity such as weight loss, diarrhea and vomiting in rats during the 28 day treatment period and four gypenoside derivatives, gynosaponin TN-1, gynosaponin TN-2, gypenoside XLV and gypenoside LXXIV were identified from GP-EX. These results suggest that GP-EX might be helpful in the prevention of Parkinson’s disease.
Keywords: Gynostemma pentaphyllum, 6-hydroxydopamine-lesioned rats, tyrosine hydroxylase, dopamine, Parkinson’s disease

In 2018, Dong et al published research into how gypenosides reverse depressive behaviour by inhibiting hippocampal neuroinflammation.33

Abstract
Gypenosides, a saponins extract isolated from the Gynostemma pentaphyllum plant, produces neuroprotective effects in the brain. Our previous studies have shown that hippocampal glucocorticoid receptor (GR)-brain-derived neurotrophic factor (BDNF)-TrkB signaling was involved in the antidepressant-like effects of gypenosides. It remains unknown whether gypenosides could alleviate neuroinflammation in depressive-like animals. The aim of the present study was to address this issue in chronic unpredictable mild stress (CUMS). Gypenosides was administrated for four weeks, followed by sucrose preference test and tail suspension test, which were performed to evaluate the effects of gypenosides. The results showed that gypenosides reversed both the decreased sucrose preference and increased immobility time in CUMS mice. In addition, gypenosides also attenuated the increase of pro-inflammatory cytokine levels in the hippocampus of CUMS animals. Furthermore, the activation of NF-κB, as well as its upstream mediators IKKα and IKKβ were inhibited by gypenosides. Last but not the least, CUMS promoted the activation of microglia, while gypenosides suppressed it according to the reduced number of iba1 positive cells. In conclusion, this study demonstrates that gypenosides exhibits the antidepressant-like effects in mice, which may be mediated by the inhibition of microglia and NF-κB signaling in the hippocampus.

In 2022, Wang et al conducted an in silico analysis to screen for compounds that can inhibit neurologically damaging signalling pathways associated with Alzeimer’s disease (AD):34

Consistent with our results, HIF-1 signaling pathway and foxo signaling pathway are involved in the occurrence and development of AD, which has been reported in previous studies. Inflammatory bowel disease (IBD) is considered to be closely related to pathogenesis of AD, based on studies of gut-brain axis. Additionally, vitamin A deficiency has been demonstrated in patients with AD, while vitamin A supplementation can alleviate the development of AD. More interestingly, our study suggests that the AD-associated pathways may relate to amoebiasis, African trypanosomiasis and malaria, which are all caused by parasitic infections and may invade the central nervous system. Existing studies have shown the existence of anti-parasitic infection and anti-AD drugs (such as GSK-3 inhibitors), which imply that the incidence of AD may relate to parasitic infection. To a certain extent, it shows the credibility and innovation of our conclusion.
Previous studies have shown that inflammatory cytokines are involved in AD. A recent study showed that elderly individuals with amyloid-beta deposition had higher levels of IL-1β and IL-6. In addition, it has been reported that higher level of Aβ42 can reduce endothelial NO synthase (eNOS, NOS3), cyclic GMP (cGMP) and protein kinase G (PKG) activity. This is consistent with our finding that GpM may prevent against AD by regulating the activity of NOS3 and the levels of IL-1β and IL-6. PON1 is a new factor associated with impaired cognition and may play a role in the development of AD. It is reported that EGFR is related to AD, and EGFR inhibitors can be used as burgeoning therapeutic strategy for AD. These data suggest that targeting on IL-1β, IL-6, NOS3, PON1 or EGFR may be effective in the treatment of AD, which supports our finding that proteins with good molecular docking are the important target proteins for neuroprotection of GpM in the treatment of AD.
In conclusion, based on network pharmacology and bioinformatics, this study illustrated the key targets and molecular mechanisms of GpM, which will provide instructive suggestions for the further study of GpM in the treatment of AD.

Conclusions
In this study, we applied network pharmacology and bioinformatics to analyze the therapeutic effect of GpM on AD. The active components and putative targets of GpM were explored and discussed systematically. Comparing putative targets of GpM with known AD related genes, constructing and analyzing CTP/PPI networks, 5 important proteins were identified, showing strong therapeutic potentials against AD. Through enrichment analysis, we further identified GpM as a promising drug with multiple components, targets and pathways for the treatment of AD. In addition, we demonstrated the feasibility of GpM in the treatment of AD by molecular docking. In conclusion, our findings provide a new idea for the neuroprotection of GpM and contribute to the development of GpM in the treatment of AD. However, data mining and analysis alone is not enough. In future, we will conduct experiments to confirm our points and provide more effective treatment measures for AD patients.

Abstract
Gynostemma pentaphyllum possesses neuroprotective bioactivity. However, the effect of gypenosides on hypoxia-induced neural damage remains obscure. In this study, Gyp, the active fraction extracted from G. pentaphyllum and its bioactive compounds as well as the underlying molecular mechanisms were investigated. Eighteen dammarane-type saponins were isolated from Gyp. The absolute configurations of six unreported compounds (13-18) were assessed via electron capture detection (ECD) analyses. The results of cell viability assay showed that Gyp and its bioactive compounds (13-16 and 18) effectively protected PC12 cells from hypoxia injury. Gyp pretreatment also improved mice spatial memory impairment caused by hypoxia exposure. At the molecular level, Gyp and its bioactive compounds could activate the signaling pathways of ERK, Akt, and CREB in vitro and in vivo. In summary, Gyp and its bioactive compounds could prevent hypoxia-induced injury via ERK, Akt, and CREB signaling pathways.
Keywords: Gynostemma pentaphyllum; dammarane-type saponins; hypoxia-induced neural injury

Figure 3. Effects of Gyp on the cell viability of PC12 cells under hypoxia condition. PC12 cells were pretreated with Gyp for 6 h and then treated with hypoxia (0.1% O2) for 24 h. The cell viability was determined by CCK8 assay. Results are presented as mean values ± S.E.; *p < 0.05, **p < 0.01.
Memory-Improving Effect of Gyp
The hippocampus, which plays a critical role in memory function, is one of the brain regions particularly vulnerable to hypoxic damage. To determine whether the Gyp administration could produce behavioral consequences, we then detected its effect on spatial memory by Morris water maze. Although Gyp treatment exhibited a slight effect on acquisition learning during the first 5 days of training (Figure 4A and B), it effectively protected against the spatial memory impairment, as described by increased quadrant entries and more distance in the target quadrant when compared with the HH group (Figure 4C and D). The swimming paths further confirmed that Gyp could prevent memory impairment induced by HH (Figure 4E). These results showed that HH alone produced an injury effect on the spatial memory of mice, which could be reversed by the Gyp administration.
Figure 4. Gyp administration reversed the memory impairment caused by HH. (A) Experimental schedule of Gyp administration, hypobaric hypoxia exposure, and Morris water maze test. The mice (10 mice in each group) were pretreated with Gyp at a dose of 50 mg/kg·day for 2 weeks and then exposed to hypobaric hypoxia (mimic high altitude 8000 m for 24 h). (B) Mean of the escape latencies to find the hidden platform across the four trials are shown for the 5 days during the acquisition training period. Ten mice were included in each group. (C and D) Mean number of platform crossings and the mean distance in the target quadrant during the probe test were evaluated. Values are presented as the mean ± SE (n = 10). (E) Representative tracking plots showing the swimming path during the probing test. The lower right circle indicates the platform location.

The current study suggested that Gyp and saponins 13–16 and 18 could prevent hypoxia-induced neural injury through activation of the ERK, Akt, and CREB pathways. The limited amounts of saponins 13–16 and 18 are unable to support the in vivo study on the antihypobaric [sic] hypoxia activity, which will be the future plan.

Hong et al (2018) conducted a biomedical investigation integrated with an in silico assay into how G. pentaphyllum can attenuate the progression of nonalcoholic liver disease in mice:36

Abstract
Nonalcoholic fatty liver disease (NAFLD) is the most common type of liver disease in developed countries. Oxidative stress plays a critical role in the progression of NAFLD. Modern pharmacological study and clinical trials have demonstrated the remarkable antioxidant activity of Gynostemma pentaphyllum (GP) in chronic liver disease. One aim of this study was to explore the potential protective effects and mechanisms of action of GP extract on NAFLD. The in vivo results showed that GP extract could alleviate fatty degeneration and haptic fibrosis in NAFLD mice. For exploring the hepatoprotective mechanisms of GP, we used network pharmacology to predict the potential active components of GP and their intracellular targets in NAFLD. Based on the network pharmacology results, we further utilized biomedical assays to validate this in silico prediction. The results showed that Gypenoside XL could upregulate the protein level of PPARα in NAFLD; the transcription level of several PPARα downstream target genes such as acyl-CoA oxidase (ACO) and carnitine palmitoyltransferase-1 (CPT-1) also increased after Gypenoside XL treatment. The overexpression of ACO and CPT-1 may involve the hepatoprotective effects of GP and Gypenoside XL on NAFLD by regulating mitochondrial fatty acid β-oxidation.

In 2017, Wong et al demonstrated how G. pentaphyllum saponins attenuate inflammation in vitro and in vivo by inhibition of NF-κB and STAT3 signaling.37 This is also another tumorigenic pathway and one of the key pathways mediated by cellular exposure to spike protein due to COVID-19 infection or transfection.38

Abstract
Recent advances in the development of anti-inflammatory agents have improved their therapeutic outcome in inflammatory bowel disease (IBD), however, the presence of side effects and limited effectiveness hinder their widespread use. Therefore, novel compounds with strong anti-inflammatory efficacy are still required. In this study, we investigated the anti-inflammatory effect and potential mechanisms of Gynostemma pentaphyllum (Thunb.) Makino saponins (GpS), a major component of the herbal medicine widely used in Asian countries. In in vitro studies, we demonstrated that GpS dose dependently suppressed activation of macrophages, one of the main effectors in IBD. GpS also suppressed cytokine production and the activation of NF-κB and STAT3 signaling in lipopolysaccharide-induced macrophages, without affecting their viability. Further in vivo studies demonstrated that GpS could ameliorate the weight loss, increased disease activity index, colon shortening and histological damage associated with dextran sulfate sodium (DSS)-induced colitis in mice. In agreement with results from our in vitro experiments, GpS suppressed cytokine production and activation of NF-κB and STAT3 signaling in the colons of DSS-induced mice. In this study, we present for the first time, evidence of the therapeutic effect of GpS in IBD, highlighting its potential as an effective therapeutic against the disease.
Keywords: anti-inflammation; colitis; gynostemma pentaphyllum saponins; inflammatory bowel disease; macrophages.

Abstract
Gynostemma pentaphyllum is a traditional Chinese medicine used for a variety of conditions, including elevated cholesterol. We have examined the pharmacological anti-hyperlipidemic and hypoglycemic effectiveness of Gynostemma pentaphyllum in the obese Zucker fatty diabetic rat model. After treatment for 4 days Gynostemma pentaphyllum 250 mg/kg reduced triglyceride (33%), total cholesterol, (13%) and low density lipoprotein cholesterol levels (33%). These effects were dose-dependent and maintained for at least 5 weeks. Chronic treatment for 3-5 weeks also reduced post-prandial hypertriglyceridemia induced by olive oil 10 mg/kg in the Zucker fatty rats but had no significant effect in lowering sucrose-induced hyperglycemia in Sprague-Dawley rats. A novel regulation by Gynostemma of glucose levels was also observed in the Zucker fatty rat model. In a glucose tolerance test in obese and lean Zucker rats pretreatment with Gynostemma pentaphyllum 250 mg/kg demonstrated glucose levels were significantly less 2 hours post challenge (20%) in the Gynostemma pentaphyllum obese rats compared to the control group. Gynostemma pentaphyllum did not significantly reduce glucose levels at 120 min in the lean strain, in contrast to the 20% decrease seen in the obese rat. In vitro, Gynostemma pentaphyllum inhibited alpha-glucosidase activity (50% inhibition at 42.8), which compared to acarbose (50% at 53.9 microg/mL). The improvement in glucose tolerance at 120 min by Gynostemma pentaphyllum in obese Zucker fatty rats but not lean rats suggests that it may improve insulin receptor sensitivity and together with the significant reduction of hypertriglyceridemia, cholesterol and low density lipoprotein cholesterol suggests that Gynostemma should be examined further by oral hypoglycemic/anti-hyperlipidemic therapy.

In 2020, Yin et al reported on ten new dammarane-type saponins with hypolipidemia activity from G. pentaphyllum herbal tea:40

Gynostemma pentaphyllum is a folk medicine and functional herbal tea, which has a good reputation for lowering blood lipid and blood pressure. It consists of numerous chemical components, such as saponins, vitamins, polysaccharides, flavonoids, and amino acids. Previous studies have shown that Gypenoside is a marker component in this plant, and its main chemical components have excellent anti-hyperlipidemia, anti-oxidative, anti-inflammatory, anti-tumor, and other biological activities. In view of its good pharmacological activity and edible value, the chemical composition aroused our research group’s interest. Consequently, chemical constituents of G. pentaphyllum, especially triterpene saponins, were systematically studied in this paper. The saponins of this plant were isolated and identified by column chromatography (CC) and preparative HPLC methods. As a result, based on the physicochemical properties and spectral data, 11 compounds (Figure 1) were obtained and their structures were determined. The 10 novel compounds were identified and named as yunnangypenosides A–J, followed by a known one, 3β, 20S-dihydroxydammar-24-ene-21-carboxylic acid 3-O-{[α-l-rhamnopyranosyl-(1→2)]-[β-d-glucopyranosyl-(1→3)]-β-d-glucopyranosyl}-21-O-[β-d-glucopyranosyl-(1→2)-β-d-glucopyranoside. Moreover, the toxicities of compounds 1–11 were detected using CCK-8 assay (Cell Counting Kit-8), and 11 compounds’ activities in lowering lipid by oil red O staining method in HepG-2 cells were also estimated. The results showed that compounds 1–11 had no obvious cytotoxicity, ten compounds (1, 3–11) were significant lipid lowering activity, with the exception of 2, and compound 8 showed the best hypolipidemia activity.
Figure 1. Chemical structures of compounds 1–11.

3.3. Extraction and Isolation
We dried the plants at 60 °C in the oven; next, the processed herbs of G. pentaphyllum (about 5000 g) were soaked in 60% C2H5OH/H2O solution (v/v) for three days, three times in total, and then filtered. The extracted solution was mixed and rotated to evaporate until no alcohol was present. We suspended the extract with water, and then extracted it with three different polar solvents (petroleum ether (PE) three times, ethyl acetate (EtOAc) five times, and five times with n-butanol).

4. Conclusions
G. pentaphyllum, as a kind of functional tea beverage commonly used by people, attracted the interest of our research group due to its lipid-lowering chemical activity. As far as we know, G. pentaphyllum is becoming more and more popular as a food and beverage. Therefore, much chemical analysis work was carried out for G. pentaphyllum, and researchers found a large number of Gynostemma saponins. However, no detailed chemical composition report was found on the plants we picked in Yunnan province, and some unknown antilipidemic active ingredients were not clarified. Based on the above reasons, we collected some samples from Yunnan Province, China, and carried out a systematic chemical separation with ethanol extract using the method of food chemical analysis. Interestingly, 10 previously undescribed dammaranne-type saponins (1–10) and one known compound (11) were obtained. To study the antilipidemic activity of these compounds, an oil red O staining assay was carried out to determine their bioactivities. Interestingly, these isolated compounds produced hypolipidemia activity in HepG-2 cells except for 2, while compound 8 exhibited the best hypolipidemia activity through the oil red O staining assay. This study provides some scientific evidence for people drinking G. pentaphyllum tea to reduce blood lipid levels, while it also provides new compounds which can be used to enrich the chemical composition of this functional herbal tea. We believe that our research will encourage further studies of the chemical composition and antilipidemic activity of G. pentaphyllum, leading to the development of a healthy tea food based on G. pentaphyllum.

Huang et al (2022) published a review focusing on the prebiotic and therapeutic aspects of saponins and polysaccharides of jiaogulan tea, and the indirect anticancer effects of a healthy gut biome:41

The anticancer abilities of jiaogulan, both direct and indirect, have been summarized in Figure 2.
FIGURE 2
Thematic presentation of the jiaogulan tea's anticancer effects. Through literature review, it is observed that different components of the jiaogulan tea possess anticancer properties that these compounds exert either directly or indirectly. Through indirect approach, jiaogulan tea's component exerts anticancer effects through the interface of the gut microbiota. Here, we summarize that jiaogulan promotes the growth of beneficial bacteria, particularly the short‐chain fatty acid (SCFA) producers. SCFAs eventually exert anticancer properties.

“Macrophages that encourage inflammation are called M1 macrophages, whereas those that decrease inflammation and encourage tissue repair are called M2 macrophages.”42

We and several other studies have demonstrated that GpS improves gut microbial composition by promoting the growth of beneficial bacteria and suppressing potential pathogens (Chen et al., 2015, 2016; Huang et al., 2017, 2018; Khan et al., 2019; Shen, Zhong et al., 2020). While evaluating the anticancer effects of GpS in a Apc Min/+ mouse model, GpS displayed a stimulating effect on the abundance of Lactococcus, Bifidobacterium, Lactobacillus, and short‐chain fatty acids (SCFAs) producing bacteria. However, the growth of potential pathogens, for example, Dysgonomonas spp., Helicobacter spp., sulfate‐reducing bacteria, were suppressed after GpS introduction to mouse gut (Chen et al., 2015, 2016; Huang et al., 2017; Khan et al., 2019; Liao et al., 2020) (Figure (Figure3).
FIGURE 3
Schematic presentation of GpS' anticancer effects through gut microbiota. (a) In colorectal cancer (CRC), the intestinal track is characterized by polyp formation, imbalanced gut microbiota, reduced mucus layer, suppressed population of goblet and Paneth cells, and inflamed immune milieu. (b) Treated CRC preclinical mouse model with GpS reinstates the inflamed mucosal immunity, promotes goblet and Paneth cell population – that results in mucus layer thickness and higher secretion of lysozyme. Most importantly, the gut microbial composition improves with the prevalence of SCFA producers. (c) At the subcellular level, GpS‐associated increase in SCFAs upregulates fatty acid‐sensing GPCRs that results in the suppression of histone deacetylases and PPARγ, which downstream inhibits PI3K/AKT oncogenic signaling pathways, as well STAT3 and Src. This graph is based on results published by Hsiao's group (Chen et al., 2016; Khan et al., 2019; Liao et al., 2020)
After noticing the stimulating effect of GpS in SCFAs producer in the mouse gut, we proved that GpS could increase the growth of Bifidobacterium animalis, Lactobacillus casei, and Lactobacillus reuteri (Liao et al., 2020). By gavaging B. animalis and butyrate (separately) to a cancer preclinical mouse model and noticing anticancer effects, we proved that the anticancer effect of GpS is partly through stimulating the growth of beneficial bacteria in the gut (Liao et al., 2020). The anticancer efficacy of GpS could be improved in the presence of polysaccharides. We confirmed the enhanced cancer‐preventive properties of GpS when applied in combination with Ganoderma lucidum (Lingzhi) polysaccharides. It was observed that GpS and polysaccharides from lucidum can greatly improve the inflamed gut barrier of Apc Min/+ mice by inhibiting polyp formation, changing colonic M1 to M2 macrophages, stopping the oncogenic signaling molecules, and increasing the E‐cadherin/N‐cadherin ratio (Khan et al., 2019).

CONFLICT OF INTEREST
The authors declare no competing financial interest.

Bioavailability, contraindications, interactions with chemotherapeutic drugs and dosage recommendations

In 2020 Ahmed at al investigated the caco-2 cell permeability of flavonoids and saponins from G. pentaphyllum as part of a study into bioavailability. Results varied according to the type of saponin. Other compounds were not assessed:43

The human epithelial cell line Caco-2 has been widely used as a model of the intestinal epithelial barrier.

If P-gp efflux pumps are later confirmed then taking with silymarin extracted from Milk thistle would be beneficial, as this can inhibit P-gp-mediated efflux in Caco-2 cells.44

The nine dammarane saponins selected for an assessment in the Caco-2 monolayer permeability assay in this study displayed Papp values in the range of 1.33 (±0.073) × 10–6 to 35.3 (±5.8) × 10–6 cm/s (Table 2 and Figure5). Gypenoside LVI (11), a saponin with four sugar units, two at C-3 and two at C-20 (Figure1), showed the highest Papp value of 35.3 (±5.8) × 10–6 cm/s, indicative of high to complete human oral bioavailability. The lowest Papp value of 1.33 (±0.073) × 10–6 cm/s was attributed to damulin A (15), an E1 elimination product of 11, and a saponin with two glucose units at C-3 (Figure1). Gypenoside XLVI (10), bearing three glucose units, two at C-3 and one at C-20, exhibited a Papp value of 15.7 (±1.16) × 10–6 cm/s, a twofold decrease in permeability compared to the four sugar bearing 11. As seen previously for the flavonoids, a higher degree of glycosylation also favors monolayer permeability of the saponins. Gypenosides LVI (11) and XLVI (10) are both glycosylated at two sites on the molecule and bear a total of four and three sugar units, respectively, and exhibit higher Papp values than all saponins investigated here.
Gypenoside L (12), with the C-3 disaccharide, showed high permeability with a Papp value of 10.7 (±2.09) × 10–6 cm/s. Interestingly, gypenoside LI (13), the 20R-epimer of 12, exhibited a 10-fold decrease in permeability (1.39 (±0.088) × 10–6 cm/s). This may suggest that the stereochemistry at C-20 (Figure1) can be a significant factor for determining the monolayer permeability of the saponins. Yixinoside B (14) exhibits the same C-20 stereochemistry as 12 and bears two glucose units attached through the hydroxy group at C-3 but is hydroxylated at position C-1 rather than at C-2 as seen in 12 and 13 (Figure1). This structural isomer of 12 exhibited a slightly higher Papp value (13.6 (±1.61) × 10–6 cm/s) than 12, suggesting that a saponin possessing a hydroxyl group at C-1 and S-configuration at C-20 is more permeable across Caco-2 monolayers than its structural or configurational isomers.
…A literature survey confirmed the lack of any report on the permeability of dammarane saponins. However, reports suggest that the other class of saponins, the steroidal saponins, can pass through Caco-2 monolayers. These saponins are actively transported by suitable transport proteins, such as sugar transporters, or can be effluxed across the monolayers by efflux pumps such as P-gp and MRP-2. We know that Caco-2 cells express active transporters including sugar transporters such as SGLT1 (responsible for the transport of glycosides), GLUT2 and GLUT5. Analysis of our permeability assay data in light of these literature reports suggests that the glycosylated saponins assessed here may be actively transported by sugar transporters across Caco-2 monolayers. The low permeability of saponins 13 and 15 may be attributed to the fact that these saponins are effluxed across Caco-2 monolayers by suitable efflux pumps such as P-gp and MRP-2. An inhibitory transport study can be used in the future to determine the specific transporters or efflux proteins that are involved.
Conclusions
G. pentaphyllum is an important medicinal herb with promising and clinically important biological activities. It also contains a plethora of structurally diverse natural products, any or all of which may be responsible for the activities observed for the extracts. Our study identifies the compounds most likely to be orally available from the complex mixture that is ingested as “the immortal tea”. The Caco-2 cell monolayer permeability assay on the flavonoids and saponins from G. pentaphyllum resulted in the identification of strong candidates for potential human oral bioavailability. The study indicated that kaempferol glycosides 2–5 exhibited moderate to high permeability across Caco-2 cell monolayers. A notable observation was that the Papp value was not significantly affected by the presence of different sugar units as galactose-containing compounds 2 and 4 showed similar Papp values to the glucose-containing compounds 3 and 5, respectively. The quercetin glycoside 7 also exhibited moderate permeability across Caco-2 cell monolayers, whereas other quercetin glycosides 8 and 9 with a higher degree of glycosylation exhibited high Caco-2 cell permeability. Quercetin glycosides exhibited higher permeability across Caco-2 monolayers than kaempferol and isorhamnetin glycosides. It was also observed that the permeability of the glycosides was dependent on the number of sugar units attached to the aglycone and the number of glycosylation sites with a higher degree of glycosylation facilitating the permeability across Caco-2 cell monolayers. Figure6 provides a summary of the key structural features that our data suggest are critical for enhanced permeability across Caco-2 cell monolayers and therefore for potential bioavailability.
Figure 6
Summary of the structural features of G. pentaphylum flavonoids and saponins that enhance permeability across Caco-2 cell monolayers. Key structural features highlighted in red.
A similar trend was observed for the highly glycosylated dammarane saponins, 10 and 11, where the permeability across Caco-2 cell monolayers was favored by a higher degree of glycosylation. Analysis of the structure–permeability correlations revealed that saponins with a hydroxyl group at C-1 and S-configuration at C-20 were more permeable across Caco-2 cell monolayers than the structural or configurational isomers. It was also observed that the compounds that result from the elimination of the C-20-OH group, compounds 15–18, showed low to moderate permeability across Caco-2 cell monolayers, whereas the precursor compounds, 10, 11, 12, and 14, exhibited high Caco-2 cell monolayer permeability, making them strong candidates for possible complete human oral bioavailability. Figure6 depicts these permeability-enhancing structural features highlighted in red. Based on the literature reports on the bioavailability of other classes of saponins and comparison with our results, we propose that the flavonoids and saponins with moderate to high permeability across Caco-2 cell monolayers are transported via active transport, possibly by sugar transporters. This study does not implicate the involvement of specific uptake transporters or efflux pumps. In the future, transport studies can be utilized to determine the specific transporters involved in the active transport of these compounds. This was the first bioavailability study of dammarane saponins and also of a number of flavonoids from G. pentaphyllum, and the results have definite significance in allowing future bioavailability assays to identify potential drugs for oral administration or structures for lead optimization.

As we have seen, there are hundreds of compounds in extracts of G. pentaphyllum, the vast majority of which are soluble in polar solvents like water or ethanol.

If to be taken as a tincture please consult the manufacturer or herbalist for specific advice as concentrations can vary.

Image result for gynostemma pentaphyllum tincture

Contraindications and safety data from various sources are now considered:

2-4 cups of tea a day prepared from dried leaves in hot water at around 80°C appears to be common advice. This is similar to the recommendations for preparing Artemisia annua tea.45

Shelf life of dried leaves or powder from manufacture is frequently quoted as 18-24 months.

From Healthline.com:46

Ginseng substitute?
Traditional Chinese medicine uses ginseng to treat stress, insomnia, colds, and flu. It’s also said to improve concentration and memory, physical stamina, and endurance. In Western medicine it’s used as a stimulant.
Jiaogulan advocates writing for the website jiaogulan.org say that it offers many of the same benefits as ginseng and can be used as a substitute for ginseng. It doesn’t contain many of the other chemical compounds found in ginseng, and it can’t be considered identical.

What to do if you want to try jiaogulan
Talk to your doctor first if you want to try jiaogulan as a complementary health approach. It’s best to use herbal medicines under the supervision of a doctor or someone trained in herbal medications. Information on the credentials and licensing of herbalists is available from the National Center for Complementary and Alternative Medicine.
There are no proven effective doses of the herb for adults or children. Herbalists usually recommend 2 to 4 cups of jiaogulan tea per day. Jiaogulan has few known negative side effects. In some people it causes nausea and increased bowel movements. In addition to tea it’s available as an extract and in pill form.

From Jiaogulan.org:47

Who should be using Jiaogulan as a supplement?
The honest answer is that we all should. This herb is many times stronger and more effective than Ginseng, and ongoing research indicates that it can provide the first line of defense against many diseases.
It will help us to better endure anything that may afflict us by increasing the body’s energy levels naturally. When our bodies have enough energy, they can fight off many of the diseases that we associate with modern day life.
This is what makes Jiaogulan so effective when it comes to combating diabetes, balancing cholesterol, reducing heart disease and increasing the brain’s dopamine levels to ward off Alzheimer’s.
Different Forms of Jiaogulan
There are different forms of treatment quality Jiaogulan available, but it is important that what ever form you use is derived from organic Jiaogulan. Not only is this more effective but it is also less likely to have been affected by pollutants such as heavy metals.
Jiaogulan tea is widely available, and can be bought online plus in Chinese stores. In China a product called tea-pills are also available, and they are fine for long term used.
Here in the West we can also find Jiaogulan available in capsule format which will allows you to control the dosage more effectively.
Jiaogulan should not at any time be seen as a “one off treatment.” As it is not associated with any harmful side effects, it can be used on an everyday basis no matter how old or young you are. Can it improve and strengthen your health? Anything that increases our energy levels to further our endurance can improve and strengthen our health, however, Jiaogulan does so more effectively.

Jiaogulan Preparation: Jioagulan is consumed as a tea, as capsules or simply raw or dried. Opinions vary on the method of preparation as a tea and seem to be a mater [sic] of personal preference. Some experts recommend treating jiaogulan similarly to a high quality green tea, i.e. brewing temperatures between 140 deg and 160 deg and brewing times under 2 minutes. Other’s prefer a much longer brewing time to maximize the extraction of the beneficial saponins.
Here are some links to videos on brewing jiaogulan tea;

From Dr.Axe:48

How to Use (Dosage)
How much jiaogulan should you take? While there isn’t a standard dosage that experts recommend, a general recommendation for adults is to consume between two and four cups of brewed gynostemma tea per day.
In two of the studies mentioned above, the dosages that were used to help promote metabolic health were around six grams of leaves per day (measured by dry weight).
To make jiaogulan tea, sleep the leaves in hot water for at least 10 minutes. According to the Indigo Herbs website, if you’re using dried gynostemma powder, add 1/2 to 3/4 teaspoon of powder per cup of hot water, let it infuse for 15 minutes and then drink up to three times per day (or as directed by your herbal practitioner).
If taking it in extract or pill form, read dosage directions for the specific product you’re using, since concentrations of jiaogulan vary from product to product. Also take note of other compounds and ingredients that may be combined with gynostemma for metabolic or heart health-promoting effects, such as berberine (a Chinese herb known for lowering blood sugar), quercetin (a flavonoid antioxidant) or vitamin C.
Some studies have found that for promoting heart and metabolic health, a daily dosage of about 10 milligrams of extract taken three times daily may be most effective and safe.
Risks and Side Effects
Is jiaogulan safe? Most studies have found there to be few jiaogulan side effects when it’s consumed in recommended amounts for several months. It seems safest to use it for up to four months before taking a break or consulting a doctor.
Some people have reported mild gynostemma side effects, including nausea and diarrhea. If you experience digestive issues when beginning to use this supplement, consider taking less or taking a break before beginning to use it again.
Gynostemma may not be safe for pregnant women or those with autoimmune diseases due to how it can impact the immune system. It should also be avoided by people with bleeding disorders and anyone taking medications to control blood clotting or that decrease the immune system.

From WebMD.com:49

Side Effects
When taken by mouth: Jiaogulan is possibly safe when the tea is used for up to 3 months, and when the extract is used for up to 4 months. The most common side effects are diarrhea and nausea. There isn't enough reliable information to know if jiaogulan is safe when used long-term.
Interactions
Moderate Interaction
Be cautious with this combination
Medications that decrease the immune system (Immunosuppressants) interacts with JIAOGULAN
Jiaogulan can increase the activity of the immune system. Some medications, such as those used after a transplant, decrease the activity of the immune system. Taking jiaogulan along with these medications might decrease the effects of these medications.
Medications that slow blood clotting (Anticoagulant / Antiplatelet drugs) interacts with JIAOGULAN
Jiaogulan might slow blood clotting. Taking jiaogulan along with medications that also slow blood clotting might increase the risk of bruising and bleeding.
Medications for diabetes (Antidiabetes drugs) interacts with JIAOGULAN
Jiaogulan might lower blood sugar levels. Taking jiaogulan along with diabetes medications might cause blood sugar to drop too low. Monitor your blood sugar closely.
Dosing
Jiaogulan extract has most often been used by adults in doses of 200-225 mg by mouth twice daily for up to 16 weeks. Speak with a healthcare provider to find out what dose might be best for a specific condition.

From “Chinese Medical Herbology and Pharmacology”:50

DOSAGE
5 to 12 grams in decoction, and 0.75 to 1.0 gram in
powder form.

CAUTIONS I CONTRAINDICATIONS
• Jiao Gu Lan may cause mild stomach discomfort when
taken as tea on an empty stomach.
• Use of Jiao Gu Lan has been associated with potential
side effects such as fatigue, lack of energy, dizziness, chest
congestion, mild fever, perspiration, sore throat, rash,
increased heartbeat, and increased respiration rate. 1
• Jiao Gu Lan may caused drowsiness and sedation.
Therefore, individuals who take this herb should exercise
caution if driving or operating heavy machinery.
CHEMICAL COMPOSITION
Gynoside, ginsenoside, rutin, ombuoside.
PHARMACOLOGICAL EFFECTS
• Immunostimulant: Jiao Gu Lan has been shown in animal studies to increase the weight of the spleen, the phagocytic activity of macrophages, and the number of T-lymphocytes and NK cells.
• Cardiovascular: Decoction of Jiao Gu Lan (20 to 40%) has demonstrated positive inotropic and negative chronotropic effects in rabbits. ln anesthetized cats, injection of Jiao Gu Lan at 50 mglkg has a reliable antihypertensive effect for up to 30 minutes.
• Antihyperlipidemic: Water extract of Jiao Gu Lan has demonstrated a marked effect to lower cholesterol and triglycerides in rats.
• Antineoplastic: According to laboratory studies, preparations of Jiao Gu Lan have demonstrated an inhibiting influence on various kinds of cancer cells, including cancers of the stomach, abdomen, uterus, liver, mouth, esophagus, pancreas, brain, lung, kidney, tongue, breast, and skin. There is also an increase in life expectancy in mice that received the herb, in comparison to the placebo group.
• Antiaging: Administration of Jiao Gu Lan is associated with prolonged life expectancy and delayed aging in animals. In one study, a group of old mice were divided into two groups. After 4 months, all mice in the control group died, while only 50% of the mice died in the herb group.
• Antiplatelet: Jiao Gu Lan inhibits platelet aggregation and thrombus formation in mice.
• Endocrine: Administration of Jiao Gu Lan is associated with an increase in plasma ACTH levels in rats.
• CNS suppressant: Extract of Jiao Gu Lan has demonstrated sedative, hypnotic and analgesic effects that last up to 7 hours in mice. lt prolonged sleeping time
induced by barbiturates and reversed the effects of mescaline. Lastly, Jiao Gu Lan at 100 to 150 mg/kg has a significant analgesic effect in mice.

HERB-DRUG INTERACTION
• Sedatives: Jiao Gu Lan has sedative, hypnotic and analgesic effects. It potentiates the sedative effect of barbiturates and reverses the effect of mescaline.
[Note: Many categories of drugs induce sedation, such as antihistamines, narcotic analgesics, barbiturates, benzodiazepines and many others.]
TOXICOLOGY
No abnormalities were reported via blood panel, or in the liver, kidney, heart, and testes when Jiao Gu Lan was given to mice at 4 glkglday for 90 days. In acute toxicology studies, the LD50 for oral ingestion of extract is 48.94 glkg, and
the LD 50 for intraperitoneal injection is 2,862 mglkg.

From “Management of Cancer with Chinese Medicine”:51

lao Gu Lan (Gynostemma Pentaphyllum)
Properties: bitter, cold.
Channels entered: Lung, Kidney.
Functions: disperses inflammation and relieves Toxicity, dispels Phlegm and stops coughing, supplements Deficiency, consolidates the Essence, and inhibits debility.
Indications: cough with expectoration of phlegm, wheezing due to chronic bronchitis, dream emission due to Kidney Deficiency.
Common dosage: 10-30g in a decoction, 3-6g as a powder.

Updated 24th August ‘22:

Jiaogulan tea, what does it taste like?

This can vary according to whether its loose leaf or powder, the age of the leaves at harvest and the drying process employed.

I used 1 x 5ml or about 1g of finely powdered Thai G. pentaphyllum in hot water at the “fisheye” boiling stage, 80-90°C and left it to infuse for 15 minutes.

As depicted it was like tasting a green tea but with warming earthy notes similar to ginseng, slight sweetness. You can easily get 2 cups out of 1 teabag. At higher strengths the bitterness comes out, and you get a slight aftertaste.

Once I’ve tried loose leaf tea I will update here.

Conclusion

An antioxidant adaptogen with antiviral, antitumor, lipid/glycaemic and nootropic effects, multiple in silico, in vitro and in vivo research papers provide significant experimental evidence to support many of the claimed health benefits, although it should be considered in its own right, not as a replacement for ginseng.

Further standardised clinical trials should also be conducted to confirm the findings we have accumulated to date from experimental research data.

Bioavailability varies from poor to excellent depending on the molecular makeup of its hundreds of compounds, but as it is of low toxicity with broad spectrum in efficacy and most of its compounds are water soluble tea is one of the most suitable ways to take it.

Taking with 80% silymarin extracted from Milk thistle as an adjunct may improve gut absorption further due to it acting as an inhibitor of the P-glycoprotein efflux pumps in enterocytes, but further research is warranted.

Taking the research findings into consideration as well as its use as a traditional Chinese medicine for hundreds if not thousands of years I would therefore unhesitatingly recommend adding G. pentaphyllum and its extracts to the list of therapeutics to take regularly or in response to a viral infection or to help treat many other health conditions as discussed.

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Disclaimer

This site is strictly an information website about potential therapeutic agents and a review of the current state of research. It does not advertise anything, provide medical advice, diagnosis or treatment. This site is not promoting any of these as potential treatments or offers any claims for efficacy. Its content is aimed at researchers, registered medical practitioners, nurses or pharmacists. This content is not intended to be a substitute for professional medical advice, diagnosis, or treatment. Always seek the advice of your physician or other qualified health provider with any questions you may have regarding a medical condition. Never disregard professional medical advice or delay in seeking it because of something you have read on this website. Always consult a qualified health provider before introducing or stopping any medications as any possible drug interactions or effects will need to be considered.

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