Research The nexian phalaris breeding programme

[chlorophyll]

Hello everyone, new and naive member here


What is known about the grass fats? If they evaporate below the DMT boiling temperature maybe you could defat in an oven without petroleum ether.

I don’t have the knowledge to see if this idea is any good, nor the material and skills to test it myself. But who knows, maybe it can help smarter people…

 

[Transform]

Hello everyone, new and naive member here


What is known about the grass fats? If they evaporate below the DMT boiling temperature maybe you could defat in an oven without petroleum ether.

I don’t have the knowledge to see if this idea is any good, nor the material and skills to test it myself. But who knows, maybe it can help smarter people…

Hello and welcome!

That's a very big if, tbh. Evaporating fats without destroying DMT is a bit of a tall order. Your speculation needs to be tempered with hard technical data. Fortunately, you're in the right place for learning how to do that

 

[Grasshoppers]

Keep up the good work!

I am unfortunately not really available these days to help more in tlc analyzing and so on, hope my time schedule will become a little bit more open the next months.

I still have the identification key for phalaris seeds, to share with you!

Arundinacae is growing strong these days!

View attachment 99079

Thank you! I'm looking forward to the identification guide.
That P. arundinacea clone looks remarkably healthy. Hopefully, we can soon send you some of the clones scheduled for breeding for your research.


We've been utilizing densitometry for some time now and wanted to share insights into the relative alkaloid distribution among the accessions we're currently studying.

Quantification of N,N-DMT is depicted below:


Quantification of 5-MeO-DMT is depicted below:


The measurements exhibit a logarithmic scaling effect, where small values are nearly linear, but as the values increase, the distances between them compress, reflecting a non-linearity that intensifies at higher measurements, similar to a sigmoid function. Additionally, the values are normalized to 100%.

As illustrated, 5-MeO-DMT exhibits a more even distribution, while N,N-DMT shows a more uneven pattern.

It's important to mention that the accessions we're currently testing are not representative of wild accessions you might find randomly. These accessions were selected due to their higher-than-usual alkaloid content found in initial screenings.

 

[Grasshoppers]

Today, we report on two phenomena that we have encountered:

1. Several of our P. aquatica specimens displayed significant fluctuations in yield, while others remained relatively stable. Notably, a clone dominant in N,N-DMT experienced a sharp drop to 10% of its initial yield within just four weeks. This marked variability necessitates further investigation. However, the observation period has been too brief to draw any definitive conclusions. A more detailed report on these fluctuations is in progress, with the necessary data currently being collected. Understanding the precise triggers for these fluctuations and determining if they can be controlled is crucial. Once this is achieved, we will adjust the testing protocol to effectively select the optimal clones.
   
   
2. We observed that P. aquatica from mediterranean origin (USDA10) has prematurely flowered in central europe (USDA7). In contrast, mature plants from USDA9a (latin american origin) showed no signs of flowering under the same USDA7 conditions. This suggests significant intraspecific variation in thermoperiodism and photoperiodism for P. aquatica. Therefore, it may be necessary to develop different breeding lines for varying climate zones.

Consequently, our breeding efforts during this period will be limited and will primarily focus on testing fertilization methods. We will not be able to effectively select clones for breeding until we have a comprehensive understanding of the alkaloid fluctuations.

 

[SpectrumFarms]

Today, we report on two phenomena that we have encountered:

1. Several of our P. aquatica specimens displayed significant fluctuations in yield, while others remained relatively stable. Notably, a clone dominant in N,N-DMT experienced a sharp drop to 10% of its initial yield within just four weeks. This marked variability necessitates further investigation. However, the observation period has been too brief to draw any definitive conclusions. A more detailed report on these fluctuations is in progress, with the necessary data currently being collected. Understanding the precise triggers for these fluctuations and determining if they can be controlled is crucial. Once this is achieved, we will adjust the testing protocol to effectively select the optimal clones.
   
   
2. We observed that P. aquatica from mediterranean origin (USDA10) has prematurely flowered in central europe (USDA7). In contrast, mature plants from USDA9a (latin american origin) showed no signs of flowering under the same USDA7 conditions. This suggests significant intraspecific variation in thermoperiodism and photoperiodism for P. aquatica. Therefore, it may be necessary to develop different breeding lines for varying climate zones.

Consequently, our breeding efforts during this period will be limited and will primarily focus on testing fertilization methods. We will not be able to effectively select clones for breeding until we have a comprehensive understanding of the alkaloid fluctuations.

Just curious, are you micro-propagating the selected specimens to make the clones or are they cuttings? If I am understanding correctly you had a clone drift down 10% in yield in 4 weeks. In theory wouldn't a micro-propagated clone of the donor plant keep the yields relatively steady?

 

[Grasshoppers]

Just curious, are you micro-propagating the selected specimens to make the clones or are they cuttings?

We propagate them using cuttings. For Phalaris, it's relatively easy to cut rhizomes or small shoots.

If I am understanding correctly you had a clone drift down 10% in yield in 4 weeks.

Even worse, that clone's yield dropped by 90% to just 10% of its original yield in 4 weeks.

In theory wouldn't a micro-propagated clone of the donor plant keep the yields relatively steady?

These fluctuations seem unrelated to plant age. Without specific data, we can only speculate. For instance, Phalaris aquatica can probably yield high initially after dormancy, especially if it experienced drought stress. However, if the soil is nutrient-depleted, growth can halt, maybe causing a significant drop in yield. Other factors could also be at play, and the situation might differ for Phalaris arundinacea.

 

[Grasshoppers]

We recently conducted a series of tests on P. arundinacea samples from a swamp region in central Europe (USDA7).
The 17 samples did not contain detectable amounts of N,N-DMT, some samples contained 5-Meo-DMT, one of which was an outlier containing significant amounts of 5-Meo-DMT.





Relative 5-Meo-DMT concentration in 17 P. arundinacea Samples




Overall the cocktail is relatively similar amongst all samples but contains a wide range of different substances.


Example 1: 4 P. arundinacea samples. uv275 fluorescence. wet plate vs dry plate.

Example 2: P. arundinacea uv275 fluorescence wet vs dry, same four samples



Example 3: P. arundinacea iodine stain, same samples.


Compared to the profile seen in P arundinace the P. aquatica samples appeared relatively cleaner.
What still makes P. arundinacea interesting is its rapid growth. The isolated clone P. arundinacea outlier clone that has significant 5-Meo-DMT countent is currently undergoing propagation for further investigation.

 

[neurobloom]

Brilliant work as always! That arudinacea outlier holds great promise. Even if it's not a clean profile it still proves that wild sourced arudinacea is able to provide a practical yield we just have to find a clean profile strain which i think it's just a matter of time.

 

[Grasshoppers]

We conducted further testing on additional P. arundinacea specimens and initiated a retesting phase. The outlier sample, bxt002, once again demonstrated a high yield of 5-MeO-DMT in the retest. While we successfully induced 5-MeO-DMT synthesis in other P. arundinacea clones, their yields were significantly lower compared to bxt002.

Currently, bxt002 is being prepared for cloning and distribution among our members to facilitate further research. It's also worth noting that the yield in most specimens can be significantly influenced by environmental factors and the timing of harvest.

P. arundinacea exhibited a diverse range of alkaloids, not just 5-MeO-DMT, which sets it apart from the P. aquatica specimens we've studied so far. It remains to be seen whether this broader alkaloid profile is beneficial or detrimental when used for psychedelic purposes. Further investigation is planned to understand its potential effects.

 

[Grasshoppers]

[H3 id='Getting+to+Know+Each+Plant']Getting to Know Each Plant[/H3]
To fully understand each plant's properties, we conduct multiple TLC analyses throughout the growing season, along with a final bioassays to verify their psychedelic potential and ensure they are non-toxic. Unfortunately, there is currently no method for rapid phenotyping, meaning that each plant requires a significant investment of time and effort.

[H3 id='Introducing+a+Clone+Many+of+You+Will+Appreciate']Introducing a Clone Many of You Will Appreciate[/H3]
We've identified a clone with several desirable traits:

• Alkaloids: 5-MeO-DMT-dominant, potent, stable, and verified non-toxic through bioassays.
• Climate: Thrives in USDA Zone 7 climates and above.
• Appearance: Dark green leaves with blood-red sap and upright growth.
• Origin: Israel-Lebanon border region.

[H3 id='Discovering+Even+Higher-Yielding+Clones']Discovering Even Higher-Yielding Clones[/H3]
We've also identified other promising 5-MeO-DMT-dominant P. aquatica clones, many of which are still young and small. These clones will require further testing and bioassays before they can be considered for final approval.

[H3 id='Our+Most+Valuable+Seedling']Our Most Valuable Seedling[/H3]
N,N-DMT-dominant P. aquatica plants with yields sufficient for psychedelic use are exceedingly rare. After extensive efforts, we've found only one seedling with a peak yield that might already be usable. A long journey lies ahead to fully understand this precious seedling.

There are also a few older N,N-DMT-dominant clones with yields significantly above average. These have been cloned, distributed, and are currently under investigation.

[H3 id='Suspension+of+Seed+and+Clone+Distribution']Suspension of Seed and Clone Distribution[/H3]

• We cannot yet provide dosage guidelines for Phalaris aquatica, as certain plants may have alkaloid peaks far exceeding baseline levels. In some cases, less than 500mg of dried leaves could pose a risk of overdose to untrained users.
• Cultivating P. aquatica from seeds is more challenging than anticipated, with several failed attempts observed among novice growers.
• Similarly, extracting alkaloids from Phalaris requires specialized techniques, and we've seen numerous unsuccessful attempts by lay users, leading to frustration.

For these reasons, we have decided to limit our collaboration to experienced individuals with the necessary commitment and capability to contribute to the research process. Applications or requests from lay users will no longer be accepted.

 

[neurobloom]

This is a crude extract from 126g fresh from the cultivar Tanit (phalaris aquatica). 96g initial harvest on the 7th of September of first growth after break of summer dormancy (4 months of Mediterranean summer drought) and 30g regrowth harvest on the 10th of September. Standard acid base pulled with chloroform as per usual.

Didn't have an accurate scale to weigh but eyeballed it seems around 1g of crude freebase. Based on smoking bioassay is around 70% pure and 5-meo-dmt dominant. It had a bit of a Stony feel to it smoked but administered intranasally and sublingually it felt like pure 5-meo-dmt so whatever are the actives that contributes to the Stony qualities it does not seem to absorb intranasally or sublingually.

Performed a mini acid base cleaning step on this crude freebase and it went up to about 90% pure feel wise smoked and the Stony effect has completely disappeared. At around ~ 8mg vaporized on a piece of aluminum foil it felt like candy flipping (LSD + MDMA) pretty potent stuff and 5-meo-dmt like but the onset and peak are pretty smoother than what I would consider as pure 5-meo-dmt. The mini acid base has eliminated most of the Stony qualities to the extract but left some of the more desirable other components that I very much like.
TLC is coming up soon. Lacking an accurate scale really sucks but this is by far my best yield. No fertilizers were used just plain tap water.

 

[neurobloom]

What used to be unconfirmed hypothesis I read from the agronomic literature by CSIRO is now a proven fact. Summer dormant aquatica buds produce the highest yield once they start growing back in Mediterranean like climates. The longer and more severe the summer drought period the higher yielding the first growth after summer dormancy. This is a new fact never mentioned before in the entheogenic literature on phalaris.
I found this trait to be the most consistent throughout the years. My second conclusion is that water stress is more important than high nitrogen fertilization for yield. Shading didn't seem to have an impact on aquatica but it seems to work well for arudinacea. The grass/mystery puzzle keeps unfolding gradually.
I'll be posting next on phalaris extraction specifics and what improved yield for me so far.

 

[neurobloom]

Extraction wise phalaris seems to pose it's own challenges. The biggest two are emulsions and low yeilds from some kind of obstruction going on preventing alkaloids from migrating to the non polar layer. Defatting step at the acidic phase didn't seem to amend this problem.

What worked for me so far is to avoid reducing the crude tea too much before basing. A more dilute tea minimizes emulsions. Secondly I use the minimum amount of base necessary for freebasing the alkaloids. I simply add sodium hydroxide prills directly into the tea in small increments until I see a change of colour and smell suggesting the free base point has been attaind and stop adding at that point. Then I use an excess of chloroform (can be replaced with DCM or a mixture of naptha+DCM if you're buying HPLC grade DCM to save on cost). The excess solvent + minimum amount of base completely elinimiate emulsions... the layers seperates very cleanly. I think for highly fatty botanicals like phalaris saponification (reaction between strong base with fatty acids as in making soap) is unavoidable and the saponins formed breaks surface tension and works as a surfactant to create emulsion between the polar and non polar layer. Keeping the pH at the minimum level needed for freebasing helps avoid or delay the saponification reaction.

With this method I had zero emulsions and very minimum fine sedimentation formed. However this method has its own drawbacks: dirty crude extract...whatever substances that would normally react with sodium hydroxide to form sediment is now left un-reacted and free to migrate to the chloroform. Also some of the non desirable actives in phalaris that would normally be poorly soluble in chloroform and only partially pulled becomes fully extractable with the excess chloroform so the crude freebase pulled has added Stony like qualities which I don't prefer. My suggestion is to wait till you have harvested enough leaf material to obtain a crude extract amount large enough for a mini acid base cleaning step with minimum solvent to eliminate those non desirable actives.

Don't worry DCM like chloroform are highly efficient at dissolving tryptamines you'll not loose yield on the mini acid base if you use minimum solvent. The cleaned extract still preserves a good chunk of the full spectrum qualities of phalaris which I really prefer over single compound minus those non desirable Stony actives.
What I have learnt so far and could confirm is that phalaris is as good yielding as any other commonly used botanical tryptamine source and under certain controllable conditions and with the right strains can even be one of the best yielding botanical sources out there.

Making e-juice cartridges is now next on my to do list. I expect several grams of freebase in the next couple months with the expanded cultivation plot (100m2) this Will be a nice demonstration for the practicality of phalaris as a reliable botanical source.. I'll be documenting the process pretty soon. Seeds currently just started germinating and being transferred into seedling trays.

 

[chlorophyll]

Wow thanks for sharing your insights on emulsions Neurobloom. Bookmarked

I am interested in searching for other wild plants than phalaris, because I don't have a garden for cultivation, only beautiful forests around!
In order to not reinvent the wheel, I have a question on Grasshoppers TLC protocoll:
After defat, why do you make an acid base extraction and analyse the freebase, rather than just mix the plant material with acetic acid or acetone, concentrate and do TLC directly? The polarity of the solvent would have to be adjusted to seperate dmt tannate or acetate salt rather than freebase.
Is it because

1. DMT Tannate is potentially not the only naturally occuring form of DMT and you would miss other forms
2. Other substances dissolved in acetone or acetic acid are much more concentrated, therefore overcasting DMT spots
3. DCM is easier to concentrate losslessly because it evaporates easily and floats on amonia solution
4. Alkaloid salts don't seperate as well as freebases
5. You can't use indole fluorescence on salts
6. Something else
7. A combination of 1-6?

Thank you so much!

 

[Grasshoppers]

Hi chlorophyll,

Thanks for reaching out. The TLC protocol you found was functional at the time but has since been significantly improved and is no longer current.

Directly spotting alcoholic or acidic extracts onto TLC plates can yield some results, especially on non-fluorescent plates, where the intrinsic fluorescence of DMT under 275 nm UV light can be observed. However, the main issue with this method is contamination by unknown substances, which introduces a matrix effect. This results in spot distortion, with the extent of distortion depending on the specific contaminants present. As a result, direct extracts aren’t suitable for accurate quantification using densitometry.

Acid-base extraction, on the other hand, minimizes contaminants, producing much cleaner spots that can be reliably quantified. While alkaloid salts don’t always separate well on TLC, we’ve found that their fluorescence properties remain intact with the salts we’ve tested.

We use DCM as a broad-spectrum solvent to investigate alkaloids that are insoluble in petroleum ether, expanding the range of compounds we can study.

If you’re interested in joining the research, we can definitely support you. We can assist with plant identification, TLC procedures, and even provide you with a custom script designed for the identification and quantification of Phalaris alkaloids on TLC plates.

Feel free to send me a private message if you'd like to join our internal discussions for further details.

 

[aizoaceous]

The polarity of the solvent would have to be adjusted to seperate dmt tannate or acetate salt rather than freebase.

To emphasize from @Grasshoppers's response, to a first order no such adjustment is required. The purpose of the ammonia in our eluent is to keep the pH high enough that our alkaloids are always in neutral form, regardless of how they were spotted. The charged form might be retained differently, so we don't want a mixture. (All charged would also be okay, using an acidic eluent. That's less common for TLC, though very common for HPLC.)

If the alkaloids are spotted as freebase, then life is easy since they're already in the desired form. If they're spotted as salts--or, even worse, as a mixture with excess nonvolatile acid--then they effectively need to get freebased in situ. The ammonia isn't a perfectly strong buffer, so that may result in a significant local change in pH. That may result in a change in retention, which may result in bad spot shape to the extent that's not uniform across the spot.

The analytes might also interact with other substances in the sample (the matrix), affecting retention and also degrading spot shape. I'm optimistic that a TLC method could be developed without initial separation of the alkaloids, especially since the indole fluorescence lets us distinguish our analytes from most co-eluting interferences, and I use an analogous HPLC method myself. That's a more difficult task than with, though. It's also potentially fragile, sensitive to variation in the plants other than alkaloid content.

I am interested in searching for other wild plants than phalaris, because I don't have a garden for cultivation, only beautiful forests around!

Phalaris is easily grown indoors under lights, at least for experimentation. With good genetics, even a few square feet should also yield reasonable amounts for personal consumption. This is especially true in 5-MeO-DMT-dominant varieties, since the dose is so much lower (though note the greater danger). I'd strongly suggest growing or otherwise obtaining some known positive plants first, since most unfamiliar plants will be negative and you'll otherwise have no way to know if your method is working.

 

[Isanara]

However, a critical consideration arises: which solvents are optimal for selectively removing more polar constituents such as resins while preserving the integrity of alkaloid salts within the sample?

Alkanes?

 

[Isanara]

The longer and more severe the summer drought period the higher yielding the first growth after summer dormancy. This is a new fact never mentioned before in the entheogenic literature on phalaris.

It's in Trout's Ayahuasca book, online at Erowid.
Summer regrowth after a drought is more potent and stress increases that potency. If you haven't read it, it is a good resource that has aged rather well. Link below:

Erowid Online Books : "Ayahuasca: alkaloids, plants, and analogs" by Keeper of the Trout

Full text of 'Ayahuasca: alkaloids, plants & analogs' by Keeper of the Trout: title page

www.erowid.org

More specifically:

Erowid Online Books : "Ayahuasca: alkaloids, plants, and analogs" by Keeper of the Trout

Ayahuasca: alkaloids, plants & analogs by Keeper of the Trout (full text) title page

www.erowid.org

And a quote from a description of P. aquatica:

Its alkaloid concentration in winter and spring are comparable to cv. Australian but its summer growth can be much stronger. However, unless there is adequate summer rain, it does not produce foliage during the summer

Below another quote:

tryptamine production was highest in young chlorophyllaceous tissue.

Erowid Online Books : "Ayahuasca: alkaloids, plants, and analogs" by Keeper of the Trout

Ayahuasca: alkaloids, plants & analogs by Keeper of the Trout (full text) title page

www.erowid.org

Oram 1970 found great variations of alkaloids between the strains and also between sampling dates. [Samples were taken as the two uppermost (youngest) leaves from 20 sites in each plot.]

He determined that the total tryptamine levels in Seedmaster [DMT is main alkaloid] and Sirocco [5-MeO-DMT is main alkaloid] were approximately 5 times greater in Autumn (April) than in Winter (July and September).

Autumn had higher temperatures, higher light intensities, longer days and more moisture stress.

Above the study correlated increased potency with summer-like conditions, warmer, brighter, longer days and more drought.

As for the summer regrowth being potent, it's why you can collect fresh tips in summer and use serial boiling, by using a pot of water, boiling the tips for 15 minutes, remove them and add more fresh ones and repeat to get an alkaloid rich solution. One can process quite a bit of grass that way. This was reported previously at the old site by AlbertK.Lkoyd as well, if I recall correctly.

The technique of selecting the tender alkaloid rich material of new growth is widely used for many things like silver needle and white tea. It also works well for Phalaris.

The literature also shows alkaloid recovery from one plant may vary from year to year. Why is not clear to me.

 

[Grasshoppers]

The analytes might also interact with other substances in the sample (the matrix), affecting retention and also degrading spot shape. I'm optimistic that a TLC method could be developed without initial separation of the alkaloids, especially since the indole fluorescence lets us distinguish our analytes from most co-eluting interferences, and I use an analogous HPLC method myself. That's a more difficult task than with, though. It's also potentially fragile, sensitive to variation in the plants other than alkaloid content.

Here’s an example highlighting the issues that can arise when spotting the acidic extract directly onto TLC plates. Unfortunately, the plate was slightly damaged, but it still clearly shows contamination, along with irregular spots and tailing. On the left side, you’ll see a wet plate with blue N,N-DMT, while the right side shows a dry plate with green 5-MeO-DMT under UV at 275nm. While the quality isn’t quite sufficient for densitometry, it works well enough for qualitative analysis. I'm open to any suggestions on how we could refine this method!

I'd strongly suggest growing or otherwise obtaining some known positive plants first, since most unfamiliar plants will be negative and you'll otherwise have no way to know if your method is working.

Totally agree! We've got a 5-MeO-dominant release candidate ready, but we're still working on completing the bioassays in different climates. That process will take about 6 to 12 months.

It's in Trout's Ayahuasca book, online at Erowid.

Thanks for sharing the relevant literature! Yes, the fluctuations can be significant. We have some clones that show a relatively high baseline yield, but we still don’t have enough data to fully understand the relationship between baseline and peak yields. Interestingly, we haven’t observed much seasonal variation so far. However, peaks in alkaloid levels do pose a risk of overdose, especially for inexperienced users.

 

[Isanara]

Yes, a standardized approach preventing the ingestion of excessive doses is ideal.