Ginseng Allelopathy in the Rhizosphere

The active, therapeutic phytochemicals in the Panax spp. appear to be ginsenosides that consist of an aglycone base structure and glycosides (sugar molecules). These compounds can be referred to as triterpene glycosides, triterpene saponins and steroid saponins. The varied nomenclature comes from the multiple ways of defining the molecule. Saponins froth when shaken. And from the image below, ginsenoside aglycones contain a steroid backbone. Based on the numbers of carbons they can also be classified as a triterpene. 

Ginsensodie Steroid

So what role do these phytochemicals they play on the plants behalf? Saponins as class have anti-fungal properties and may act allelopathically (Carter et al., 1999). The sterol portion of the molecule appears to be inserted into and disrupt fungal membrane integrity by interacting with fungal sterols present.

Nicole et al. (2002) noted that American ginseng saponins inhibit in vitro growth of the fungus Trichoderma spp., but stimulate growth of Cylindrocarpon destructans.

In a followup study to  investigate the concentration of ginsensosides in rhizosphere, Nicol et al. (2003) collected ginsenosides from root associated soil several times between 1999-2002. They found that the concentration in the soil ranged 0.02 – 0.098%. They also collected root exudate from pot-grown ginseng over 22 days, using an exudate trapping system,  which yielded a concentration of 0.6% ginsenosides. We need to test whether this soil concentration level can be considered an active allelopathic level.

Additional evidence supporting the sterol disruption hypothesis can be found in the Pythiaceous fungi (especially Pythium spp. and Phytophthora spp.) that lack sterols in their  cell walls.  Growth of these fungi appear to be stimulated by both the presence of fungal sterols (ergosterol) and ginsenosides in the medium of in vitro studies. The authors suggested potential mechanisms include: 

The ginsenosides…

  • provide a carbon sink for the fungus.
  • alter fungal membranes in a positive manner.
  • act as a fungal growth hormone.

Future studies should look at the ratio of ginsensosides and their respective influence on fungi.

References:

  1. Carter, JP, Spink, J, Cannon, PF, Daniels, and Osbourn, AE. (1999) Isolation, Characterization, and Avenacin Sensitivity of a Diverse Collection of Cereal-Root- Colonizing Fungi.AppliedandEnvironmentalMicrobiology. 65(8): 3364–3372.
  2. Nicol, RW, Traquair, JA and , Bernards, MA. (2002) Ginsenosides as host resistance factors in American ginseng (Panaxquinquefolius).CanadianJournalofBotany. 80(5): 557-562.
  3. Nicol RW, Yousef L, Traquair JA, and Bernards MA. (2003) Ginsenosides stimulate the growth of soilborne pathogens of American ginseng.Phytochemistry. 64(1):257-64.

Rhizosphere Influence on Plant Medicine

Einjähriger Beifuß (Artemisia annua)
Artemisia annua
Wikipedia

Mycorrhization leads to nutrient and information flow, often in both directions. The plant root supplies sugars to the fungus, while the fungus induces Jasmonic Acid biosynthetic enzymes in the plant, leading to an increase in jasmonate ­ levels that enhance the accumulation of soluble sugars in plant root and the production  of plant root defense compounds.

From a research article,  the presence of mycorrhizal fungus, Glomus mosseae and nitrogen fixing Bacillus subtilis on the roots influenced the levels of plant biomass growth, and the yield of an important medicinally active phytochemical, artemisinin, from Artemisia annua L and used as an anti-malarial treatment.

Gabriele et al. (2016) investigated the effect of mycorrhizal soil inoculation of various Sangiovese wine grapes and found the presence of the fungus increased levels of 14 polyphenols compared to un-inoculated plants. Here the presence of symbiotic relations in the soil altered the phytochemical makeup of fruit.

So how are the plant roots attracting mycorrhizal symbionts? Plant produced flavanoid compounds accumulate at root tips/cap and make up a large portion of root exudate (the portion of the root sap excreted to the external environment). These phytochemicals are easily modified and their biosynthesis is triggered  by transcription factors, which suggests a role as elicited signal compounds – compounds that are made specifically in response to conversation from rhizosphere fungi and bacteria. Interestingly, their presence in the rhizosphere soil triggers mycorrhizal fungi to explore their surroundings (Hassan and Mathesius, 2012), perhaps increasing the likely hood of contact with plant roots.

Given the high price of American wild grown ginseng, the ecological influence on ginsenoside formation, and ultimately, the therapeutic value, points to optimizing the rhizosphere cross talk by way of forest farming.

The highest ginsenoside content occurs (from highest to lowest) in the root hairs > lateral roots > cortex > interior taproot (Li and Wardle, 2002), exactly where we should expect a chemical conversation to occur.

Within this class of compounds we designate as ginsenosides, two molecular forms are dominant, protopanaxadiols and protopanaxatriols. Data from two different papers (Zhu et al., 2004: Wang et al., 2010) compared levels of diols and triols in different species and sources of ginseng. American ginseng (Panax quinquefolia) had higher levels of the triols (especially Rg1) compare to Chinese ginseng (P. ginseng), which had higher levels of diols (especially Rb1  Rd).

Structures-of-ginsenosides-from-Panax-ginseng-Glc-glucose-Rha-rhamnose-Araf
Li, H, Lee, JH, and Ha, JM. (2008) Effective Purification of Ginsenosides from Cultured Wild Ginseng Roots, Red Ginseng, and White Ginseng with Macroporous Resins. Journal of Microbiology and Biotechnology. 18(11):1789-91. DOI: 10.4014/jmb.0800.192

Comparing wild grown versus cultivated plants within each species, a similar pattern emerged, with wild plants showing a higher concentration of triols (especially Rg1  Re), while cultivated plants had higher concentration of diols (especially RbRb2).

James, et al. (2013) investigated levels of diols and triols in wild sourced P. quinquefolia leaf and root  in a North Carolina collection, finding that there was no relationship between age and ginsenoside content. However total ginsensosides were higher in the leaf, as was Rb2 and Rd (diols), In the root tissue, Rb1(diol) and Rg1 (triol) was found to be higher.

This has implications for how we “farm” medicine and speaks to a long held tenet; complex interactions in native ecologies, including the soil,  produce medicinal plant crops that are more biologically active. Farm versus wild grown ginseng is only one example. What’s been your experience as a imbiber, herbalist, researcher, plant grower or manufacturer?

 

Lousewort

Watercolor painting of Pedicularis_bracteosa-2.
(Wikipedia)

Pedicularis bracteosa
P. canadensis
P. greenlandica
P. attollens

Ecology – Hemi-parasite

Molecular phylogeny more recently placed the genus Pedicularis in the Orobanchaceae, when formally it had been  in part of Scrophulariaceae.

Herbalist Michael Moore has written on the therapeutic uses Pedicularis spp., as has David Winston and 7Song.

The plant is an excellent skeletal muscle relaxant, with some of its specific indications as follows:

  • Adrenaline-stressed or nerve impinged muscles
  • Hypertonicity and muscular rigidity
  • Children with highly excited flight or fight response

I’ve created formula with Pedicularis for massage therapist and chiropractors to  increase “hold” of treatment. In particular, it combines well with other skeletal muscle relaxants include Black cohosh (Actaea racemosa), Kava kava (Piper methysticum) and Skullcap (Scutellaria).

Since it is a root parasite the plant can take up compounds from it’s host plant. Schnieder and Stermitz (1990) noted that several Pediculars.spp. uptake alkaloids from a variety of hosts: pyrrolizidine alkaloid senecionine from Senecio triangularis, anagyrine from Thermopsis montanaN-methylcytisine from  Thermopsis divaricarpa and quinolizidines from Lupinus argenteus.

For this reason it’s unclear which therapeutic compounds are made by the plant and which come from host, which can make the safety profile a little trickier to predict. The host compounds can even alter the pigment of Pedicularis flowers. Best to find it growing alone in its own stand, or rely on a highly skilled wildcrafter to help identify a good stand.

Experimentation and observational studies have shown that two hosts can be parasitized simultaneously. Such threesomes seem to improve the overall growth performance and survivability of the parasite.

This is a fascinating plant that requires the deft touch of an herbalist, with science providing interesting data on how plant parasites interact with their ecosystem.

 

 

 

 

Do Plant Roots Talk to Leaves?

Arabidopsis thaliana
Arabidopsis thaliana (Wikipedia)

Surrounded by material excreted (exudate) by their own root border cells, the growing root tips (apical region) of plants move through soil regions where important biological interactions occur with a community of soil microbes. This exudate not only helps define the soil microbiome (microbial community), but also changes the physical and chemical characteristics of rhizosphere soil.

Root-tip-tag
Root tip (100×) 1. Meristem 2. Columellae 3. Lateral part of the tip 4. Dead cells 5. Elongation zone (Photo: SuperManu – Clematis)

Hiltpold et al (2011) provided evidence of  systemic, volatile signals in maize roots in response to herbivore attack. From 2013 research on Arabidopsis suggests that soil microbes can alter plant leaf chemistry to inhibit insect feeding. They posited a role for microbial-derived volatile organic compounds acting as a deterrence signal, and noted the presence of Actinobacteria, Firmicutes and Proteobacteria in soil and within Arabidopsis root tissue.In a 2013 Tansley Review, Turnbull and Lopez-Cobello noted that despite localized cellular communication found in the root apical meristem, communication via vascular transport to the rest of the plant did not seem to occur. That left me wondering how plant roots communicated changes throughout the entire plant (systemic).

Those microbes are often associated with “soil odors”. On a sensorial level,  “smelling” the earth may help us appreciate the complex, unseen communication happening under foot.

Are All Plants Carnivores?

Fungal species of the Metarhizium genus colonize most land plants and help provide nitrogen to the plant root. The nitrogen source is unique – insects that the fungus has pathogenized and killed using enzymatic degradation of the insect’s shell.

200px-Metarhizium_anisopliae_infected_cockroach_(PLoS)
Insect infected with Metarhizium spp.

Mike Bidochka of Brock University investigated the phenomena by injecting labelled nitrogen into Galleria mellonella larvae (moth). They buried the larvae in soil and separated the larvae from either beans (Phaseolus vulgaris) or switchgrass (Panicum virgatum) plants using a screen with pores large enough for fungal mycelium to grow through but small enough to prevent plant root growth.

Fourteen days later, they found labelled nitrogen made up more than a quarter of nitrogen found in plant root tissue. Insects Larvae with labelled nitrogen not infected by the fungus did not act as nitrogen sources for the plant.

Good evidence for an ecosystem rich in biota, rather than one where selective human inputs alters it into a simpler set of relationships. In most cases, the soil environment becomes less sustainable.

Are you a Renegade?

The renegade, or wild thing, in my heart is a man that loves to play.

I was cheerfully reminded of my need for play at a men’s retreat, how a water balloon and squirt gun fight, with little or no rules, can lift my heart and mind. I’ve been in play deficit, probably for some time. My inner grown up has been in charge much too long.  Although I consider the craft of writing, both science and poetry, a form of play, rarely do I experience physical play. Working out is not really play, but it can be – see Go Animal.

My girls claim I play like a boy, a bit too wild. When I was playing pickup soccer, or actively studying martial arts or dance I played daily. Stuart Brown at the National Institute for Play presents research in a TED Talk from 2008. Take a minute and review your own life. Are you playing enough?

I do play using my mind quite frequently, my inventive creativity a strength. I firmly trust the concept of Beginners Mind, as espoused by Shunryu Suzuki. I find in mentoring herbalists and researchers the freedom to play is a great asset that few have learned. This is especially true in multi-disciplinary environments. Here’s an example I ran across recently about the power of trusting in play.

Justine Musk’s blog discusses the Innocentive problem solving website started by Alpheus Bingham, a vice president at Eli Lilly. He asked the public to solve some of the company’s hardest scientific problems by offering a financial reward to anyone who proposed a solution.

When solvers “rated the problem as outside their own domain”, Alpheus noted that they were more likely to stumble upon solutions. They were “bridging knowledge fields” – taking ideas from one domain and introducing them into a different domain. They reframed problems, combined and recombined ideas, and opened up new lines of thinking.

This message echoed my own experience. Pursue your passions outside the explicit area in which you “work”. Embrace your inner renegade. And don’t back down from throwing unusual ideas into the conversation. Have faith in your own unique perspective. You never know when creative people in your sphere of influence will “run with it”.

I want to give a shout out to my best bud, Chas Murray. Whenever we are together, we play. Let me share what I miss the most. He was living in Norfolk and when I’d visit we inevitably ended up at the beach. Our favorite game was to take a ball or a Frisbee and play catch with a twist. The  player without the ball ran into the surf at full speed and the thrower tried to lead them into a full layout just as a cresting wave arrived. You had to trust in what would unfold and that your commitment to the moment was a joyful act in and of itself!

Not knowing is the most intimate thing (Zen Master Jizu).

Plant Chemical Out Posts

Flower of Garden Strawberry (Fragaria ×ananassa)
Image via Wikipedia

The search for chemical mediators in plant root rhizosphere interactions with symbiotic and pathogenic organisms found in the soil continues to generate interesting research. Martha Hawes group at the University of Arizona reported on the role of sugars, proteins and small molecules found in root cap secretions – a mucilaginous mixture that covers the growing root tip and “converses” with the surrounding matrix of living organisms. The cap is rich in root border cells, which detach from the growing root tip. Curlango-Rivera et al (2010) provides us a bit more detail about which metabolites are biologically active. Neither sugars nor amino acids triggered root growth or border cell production. Transient exposure to biologically active concentration levels of the isoflavonoid pisatin, a phytoalexin, stimulated root border cell production but not root tip growth. I wonder if inhibition of root elongation may “reset” plant growth patterns as root border cells, acting as chemical sense organs, define the nature of the environment?

A second paper used histochemcial methods to profile root metabolites in plants from the Rose family (Hoffman et al., 2010). They found flavan-3-ol molecules in the root tip and border cells. Their findings suggest that the distribution of flavan-3-ols in Fragaria and Malus is under tight developmental control. These molecules are found in plants as catechin and epicatechin derivatives and in long chain (polymeric) form. They influence the taste and medicinal potential of green tea and wine, to name a few well-known plants. Previous researchers summarized their role in chelating toxic cations (metals) in the soil, establishing mycorrhizal interactions and priming plant root defense. This paper suggests a role in the transport of the long distance plant hormone auxin, which would link the chemical cross talk at root border cells with responses that occur in tissue distal to root tips. Hoffman’s research lacked a clear distinction of whether the monomeric or polymeric flavan-3-ol forms where the active species. This has plagued plant research for some time, since the analytical methods for detecting the polymeric forms have been crude and ineffective. All of their samples were from a botanical garden. I wonder if the flavan-3-ol profile would differ compared to native wild grown species?

References:

  1. Curlango-Rivera, G. et al. (2010) Plant Soil 332:267-275
  2. Hoffmann, T. et al. (2011) Plant Biology, 13: no. doi: 10.1111/j.1438-8677.2011.00462.x