Two studies, addition of DET and Tryptamine to mushroom substrate

[yaesutom]

It looks to me from this study:

Addition of DET Gartz study (Oct. 1988)
http://www.geocities.com/radio879z/psilocybebio.pdf

And this study already on the shroomery,

Addition of Tryptamine Gartz study (March 1988):
http://www.shroomery.org/index.php/par/7953

That addition of DMT to the substrate may provide just as much potentiation / increase in psilocin as adding Tryptamine HCl.

Both studies show 5 flushes and up to 3.3% 4-hydroxy-DET/DMT out at the end.

Here's another image,

http://www.geocities.com/radio879z/psilocybes_enzymesteps.gif

I recently got ahold of a couple grams of Tryptamine freebase, and I do have mimosa bark and DMT, also a friend has access to analysis equipment..

So hopefully sometime soon i'll be able to innoculate several jars, some with tryptamine added, some with ground up mimosa bark added, a jar or two with nothing added..

Now, its easy to make the HCl salt of tryptamine, but just as easy to make the phosphate salt (I have some phosphoric acid), and the point of making a salt of Tryptamine (or DMT) is to make it water soluble right? so..

The results in Table 1 show that the enzyme systems in Psilocybe cubensis have a high hydroxylation and methalation capacity to convert added Tryptamine to psilocin. It is possible that a reduced amount of phosphate in the culture media decreased the bio-synthesis of psilocybin from psilocin in the media.

For psilocybin, for the phosphoryloxy group, where does it get that from? I would think possibly using phosphoric acid instead might result in more psilocybin production, I have IMed DMT phosphate solution before, adding 1/3 molar phosphoric acid (1 molecule of phosphoric acid can attach to 3 DMT molecules, enough to make it water soluble), but each psilocybin molecule has its own so I could add an equal molar amount of phosphoric acid per tryptamine or DMT molecule, i'm wondering what is the best ph, or a range for the ph that the substrate should or could be for proper growth?

I haven't read up on it but some of you might know, what effect does ph have on mushroom growth in general? How much phosphoric acid could I add to the substrate mix, to add more phosphoric acid but not let the ph drop too low?

It'll be a while til i could get some results, but from everything i've read it seems like adding either tryptamine or DMT to the substrate would probably produce super incredibly potent mushrooms, probably equally, but since Tryptamine is hard to obtain - mimosa root bark IS NOT hard to get and INCREDIBLY cheap!

In that study i linked to above (Oct. Gartz study) they added DET (also talks about adding NMT resulting in more baeocystin), got mushrooms with as much as around 3.3% of the dry weight of 4-HO-DET out (same percent as adding Tryptamine HCl), both studies show a chart with 5 flushes and similar % numbers.

Now i'm not sure why this hasn't 'caught on' yet, because people HAVE added DMT to the substrate and gotten super potent mushrooms out, and if its as easy as simply adding a little ground up mimosa hostilis root bark to your substrate i'd think once enough people started doing it, ...well who wouldn't want to do it?

Typically mimosa bark has 0.57% DMT by weight (average maybe.. often more), thats around 50mg per 10g bark, I got some info from someone who's added different amounts of DMT per jar and found that for a "pf style" jar, 30mg - 40mg produced more potent mushrooms and 50mg seemed to be the max in his experiments they would take, and adding 60mg+ didn't hurt anything but didn't add any more potency than 50mg - but the 50mg or more jars produced some mega godly potent shrooms!

So looking at those numbers lets say typical dried mushrooms contain 0.6% combined psilocin & psilocybin, so a 5 gram cubensis trip would be around 30mg psilocin/psilocybin (correct me if my numbers are off).

Now if you have dried cubensis with average 3.0% psilocin then 5 grams would be more like 150mg's.. 5x as potent, or lets say even 2.0% would be over 3x potency, which seems to be what people have been reporting from just adding DMT (at least 2x, up to around 5x potency).

The price of 1kg mimosa root bark can be had for as low as $100, enough for 100 "pf style" jars, costing $1 per jar to add 10g bark for such a huge potency increase..

Ok even if you didn't get 5x potency by just tossing in 10g bark, a dollar for 2x potency at the lowest, ain't too bad..

 

[Tryptamine*Dreamer]

This sounds interesting, thanks for the info. I'm going to try this with the mimosa root bark soon, sometime this month if possible. If it just doubled the potency of the mushrooms it would be well worth it.

I may also try it with tryptamine HCL but that will be a while off.

 

[fastandbulbous]

I've read bits about this and what really surprised me is the fact that the tryptamine-4-hydroxylase is so non-specific. If I recall correctly, they also used DoPT and MiPT as substrates and got the appropriate 4-hydroxy compound (can't remember if they said anything about getting 4-phosphoryloxy compound).

Might be quite a good method for preparing the 4-hydroxytryptamines that Shulgin said were difficult/impossible to prepare synthetically, such as 4-hydroxy DPT and 4-hydroxy DALT (enzymes make synthetic chemistry so much easier!)

 

[yaesutom]

I've read bits about this and what really surprised me is the fact that the tryptamine-4-hydroxylase is so non-specific. If I recall correctly, they also used DoPT and MiPT as substrates and got the appropriate 4-hydroxy compound (can't remember if they said anything about getting 4-phosphoryloxy compound).

Might be quite a good method for preparing the 4-hydroxytryptamines that Shulgin said were difficult/impossible to prepare synthetically, such as 4-hydroxy DPT and 4-hydroxy DALT (enzymes make synthetic chemistry so much easier!)

I've heard about mushrooms being grown and sold with DiPT added (there is a trip report on bluelight i remember seeing a while back, someone tripped off these supposedly), and a friend across the country said he thinks he does have some 4-HO (or maybe AcO don't remember)-DPT, although its not been analysed.. if that is what he has then 4-HO-DPT isn't too impressive - a very high oral dose and nothing spectacular (but i'm not convinced until its been analysed!).

Now what I couldn't get out of the 2nd Gartz paper, when they added DET, and got mostly 4-HO-DET out (and a little 4-PO-DET), i'm too lazy to find the exact quote but it said something about no detectable levels of psilocin/psilocybin found.. or no more than normal? Well whatever it said I couldnt figure out if they meant that adding enough DET resulted in ONLY 4-HO/PO-DET out, and stopped all psilocin/psilocybin production, or if there was still a minor amount of psilocin/cybin present?

It would be really fucking useful if simply adding enough N-whatever, N-whatever-tryptamine results in 4-HO-whatever out and stops psilocin/cybin production totally then you wouldnt have to worry about the minor amounts of those mixed in with whatever your trying to make/get out of it.

----

I'm thinking I should innocuate a bunch of jars, but have one jar where basically I just toss in small amounts of most of the tryptamines I have, any idea what the enzymes would do with AMT? Well, i'd toss some in anyway, also some 5-MeO tryptamines to see if any kind of 4-HO-5-MeO-whataver comes out, could just put small amounts of several in the same jar (or a couple jars) and see what shows up in an analysis..

 

[fastandbulbous]

Well whatever it said I couldnt figure out if they meant that adding enough DET resulted in ONLY 4-HO/PO-DET out, and stopped all psilocin/psilocybin production, or if there was still a minor amount of psilocin/cybin present?

I'd imagine that it would reduce/halt synthesis of psilocin/psilocybin as the DET added to the culture would be competing with any DMT synthesized by the fungus to be a substrate for the hydroxylase enzyme. The concn of DET added in this way is going to be many times that of DMT formed as part of the biosynthetic pathway, so (according to the principals of enzyme kinetics) is going to 'take up' much more of the hydroxylase enzyme.

With compounds like AMT & 5-methoxytryptamine there's always a big possibility that they'll act as competative inhibitors of the enzyme so that you end up with wery little of anything.

 

[mumintrol]

tryptamine-4-hydroxylase is so non-specific

Fungal enzymes are non-specific as a rule, and have other interesting features

sorry can't give quotation - lost the djvu file... it was in russian anyway

 

[yaesutom]

well I wonder how easily DALT can be made, if i could obtain some.. i'd wait until i perfect my growing ability first, but i'd innoculate a jar with DALT added so I can try out 4-HO-DALT . I think Shulgin said he expected 4-AcO-DALT to be one of the fastest orally active psychedelics, well it would be nice to see how good of a psychedelic 4-ho-dalt is, with the help of my mushroom enzyme friends.

 

[TheDEA.org]

what really surprised me is the fact that the tryptamine-4-hydroxylase is so non-specific.

Given that it's attacking the indole ring, it's likely that the indole is the key shape in the enzyme-substrate fit. The ethylaminewhatever tail may be able to flop around as it sees fit in most cases.

I'd imagine that it would reduce/halt synthesis of psilocin/psilocybin as the DET added to the culture would be competing with any DMT synthesized by the fungus to be a substrate for the hydroxylase enzyme.

It's possible that the 4-hydroxylase enzyme isn't the rate-limitting step, in which case the extent of the inhibition of psilocin production by adding DET might not be dramatic. If the researchers found little or no 4-hydroxyDMT or DMT when supplementing with DET, that would suggest that inhibition of the psilocin pathway was occurring further upstream (at the tryptophan decarboxylase or methyltransferase steps) rather than simply through competition for the indole 4-hydroxylase enzyme.

​

 

[fastandbulbous]

^ Even so, if the growing medium contains DET the concn of DET in the fungal cytoplasm is going to be a lot higher than that of any DMT produced via the psilocin pathway and as such the higher concn of DET will mean that the majority of the 4-hydroxylations are going to be of DET. This would occur whether or not the hydroxylase enzyme was the rate limiting step. If there isn't a specificity issue to take into account, the enzyme kinetics say that reaction rate is dependant upon the substrate concn and because there's only a finite amount of the enzyme, DET is going to be the major substrate (a bit like the mechanism behind giving people ethanol to treat methanol poisonong - the higher concn of ethanol prevents the methanol from being metabolized to the toxic metabolites)

 

[yaesutom]

Ahh! Check this out.... from Ask Dr. Shulgin,

http://www.cognitiveliberty.org/shulgin/blg/

4-Hydroxy-5-methoxy-N,N-dimethyltryptamine would be a fascinating compound to explore. The reason it's not in TIKHAL is that it is virtually unknown. The only report of it in the chemical literature was a paper published by Marc Julia's group at the Pasteur Institute in 1965. They reported the synthesis and physical properties of the compound but to my knowledge it has never been explored in any way. The synthesis is quite a frightening thing. It starts with ortho-vanillin and takes approximately 10 steps to get to the 4,5-HO-MeO-DMT. I'm not surprised that no one has pursued the compound.

However there is a very interesting study that took place in Leipzig about 15 years ago. Jochen Gartz, a mushroom explorer whom I know quite well, has done some fascinating studies with Psilocybe species by raising them on solid media containing strange tryptamines that are alien to the mushroom. Apparently the enzymes that are responsible for the 4-hydroxy group of psilocin are indifferent to what it is they choose to 4-hydroxylate. He has taken things like DPT or DIPT and put them in the growth media and the fruiting bodies that came out contain 4-hydroxy-DPT or 4-hydroxy-DIPT instead of psilocin. In fact, he has a patent on the process. These active compounds are made by the mushroom so they really are natural and yet they never have been observed in nature. I'll give you even odds that if you put spores of a psilocybe species on cow droppings loaded with 5-MeO-DMT you would come out with mushrooms containing 4,5-HO-MeO-DMT. This way you avoid a 10 step synthesis by growing a psychoactive mushroom that contains no illegal drug.

INTERESTING! . So being that the spores are legal almost everywhere, if this is true.. that adding enough of some other tryptamine stops psilocin/cybin production, technically you could legally grow mushrooms containing these things using cubensis spores possibly?

Well... i'm going to go through my box of tryptamines and see what I have, I only have a little bit of 5-MeO-DMT left its the HCl salt and i'd give it all up in the name of science (and possibly testing out a hopefully active new compound..). I am out of 5-MeO-MiPT.. I do have a little bit of AMT, but also 5-MeO-AMT.. and some others..

Maybe I will do some testing first, with as many jars as possible (I REALLY hope some others out there will help me along with this research! its fun shit..), maybe one jar with just tiny amounts of many tryptamines tossed in hoping to get a bunch of detectable amounts of 4-ho-whatever's out, so I can hopefully see what works and what doesn't.

VERY facinating! If the mycellium will take something like 5-MeO-whatever and add a 4-HO group to it, and they turn out active.. or take AMT and produce 4-HO-AMT.. well there's a lot of easily made compounds anyone could "synth up" at home out of their RC collection..

I would love it if some other people who had more mushroom growing experience would get on this, I only grew some once, if anyone has any tips on how to speed up the process, make it simpler (and/or fastest / easiest way to fruit them, maybe smaller jars, and what substrate to use, a good cubensis strain. etc etc) let me know - I will practice on just the ones with added tryptamine and mimosa bark first, then adding (mostly pure) DMT, get a good method down before trying other tryptamines so hopefully I don't fuck up any (or many!) with things like bacteria etc.

Oh, found that dipt'ed shrooms trip report:
http://www.bluelight.ru/vb/showthread.php?s=&threadid=215910&r=4

Okay i am not a big writer and am writing this a year after growing and a week after tripping but i will try to give you some information witch might or might not interest the myco- or tripto-files reading this.
The idea behind this experiment came after reading the statement of A. Shulgin in his book Tihkal under 4-oh-det that maybe the mushroom was a "x in, 4-oh-x out"-machine, and the papers by J.Garz.
I had grown some cubensis using rye. I took one of my jars of completely collonated rye and added 100mg dipt to a small amount of soil and vermucellite. I had done this because in the earlyer stage of rye colonization i would normally loose some jars to infection. After a certain amount of time cubensis mushrooms started to appear. Unluckily i could harvest just a small amount of mushrooms before the rest of the culture got infected. It is with these mushrooms i did my experiment. I had given some to a friend who had reported eating just a little amount. But since the effects did not develop within 15 minutes (and he had taken a small dose) he concluded that he had not had an 4-oh-dipt effect.

Setting: a cottage in sweden
Substances: half a B-100 pill, half a C-1000 pill, about 2mg of deprenyl (held under the tung), half a 40mg inderal pill, four OO capsules of mushroom powder with about 3,1 grams of mushroom powder total, half a 50mg tofisopam tablet.

Chronology:

11:00 or 12:00 : had eaten breakfast of pasta from the dinner the day before

13:50 : Decide to take the mushrooms today. And take some b-vit, c-vit (both time release), inderal, and deprenyl. I take the first two almost every day. The deprenyl i take two or three times per week. I should probably have left the deprenyl out. But after an earlyer experience where i had purposefully abstained from deprenyl for 10 days, so as to not mess with my first taste of Methylone, and then found the effect of Methylone (180mg) to be almost completely absent i decided that i should make a point of taking my "medicine"that day (someone else taking 1 mg every day had had no problems with Methylone, and i had read a comment of one guy who had much less effect from mdma when taking 5mg deprenyl three days prior, but not when taking directly prior to mdma). Of course Methylone and mdma are no triptamines, but hey, i did what i did anyway. The inderal was to combat the panic wich sometimes comes over me when taking a psychedelic these days. I did not want the panic to spiral out of control, and thought of it as a possibility to overcome that panic just as stage fright can be overcome by taking inderal once or a few times before an event, and then by positive learning making the use of the substance unneccesary. The inderal impacted my trip but i think it also did do what i intended.

14:00 : I drop the first of four capsules. pack a bag with some provisions and cd's. Put on "bitches brew" by Miles Davis and leave the cottage to have a walk. Short panic, what have i done? Is this what i want? But i go my way and think no more of it. After about ten or more minutes of walking i have an alert that something is happening, but it does not develop.

14:15 : I took another capsule of mushroom powder. Since 4-oh-dipt seems to be a fast and hard hitter i had planned to space the four capsules 15 minutes apart. If the mushrooms tuned out to be more potent than normal mushrooms (wich would mean there would at least be some 4-oh-dipt along with psilocybin, i had read of up to 3% strength with triptamine/DET fed mushrooms) i would have some time to find out before taking a full dose. The swedish scenery was beautiful with little fields with cows forrest, and a little stream running threw. I decide to lay down on a natural stone wall and enjoy the view. No more than the alert before.

14:35 : Take another capsule. I am surprised not to feel any more than i do. Normally i would feel something within half an hour of taking mushrooms. Maybe a year of storage might have decreased potency quite a bit.

14:40 : What the fuck, i drop the last capsule and head back to the cabin. The scenery makes me think this would be a good place to take acid. Probably because it reminds me of switzerland where i had taken my first day time acid.

Ok now the chronology gets lost. I can just estimate.
When coming back to the cabin (probably about ten or fifteen minutes after taking the last capsule) i sit down in the garden and look at the lake and the trees surrounding me. I start to feel the trip. It feels different than a mushroom trip. I come to the conclusion that this is : more purple than mushrooms... and think the description is quite funny. Hi hi..
When looking at my surroundings the purpleness reminds me of 2-ct-7 (probably because the package(blue mystic) was blue/purple) and also reminds me of it and other substances because it feels "chemical" (not dirty just not organic). I do not feel a connection with the trees, nature seems uninteresting ( in contrast to mushrooms, lsd, or san pedro). I decide to close my eyes and check it out. A swirling...*snip*

 

[TheDEA.org]

^ Even so, if the growing medium contains DET the concn of DET in the fungal cytoplasm is going to be a lot higher than that of any DMT produced via the psilocin pathway and as such the higher concn of DET will mean that the majority of the 4-hydroxylations are going to be of DET.

Yes, I guess my main point was that if DET were simply out-competing the DMT for access to the hydroxylase, there could still be a build-up of unhydroxylated DMT. If such wasn't observed, that would be suggestive of inhibition of psilocin production through a feedback loop targeting part of the DMT pathway.

Then again, there's no reason why DET couldn't also competitively inhibit the earlier steps of the pathway as well by acting as a substrate (just not being altered by them.)

Too many questions, not enough data. What brave soul will test Shulgin's idea by throwing a 5-methoxytryptamine at his mushrooms? :D

 

[fastandbulbous]

^ Ah, right - got what you mean. Yes they (fungal enzymes) seem so non-specific that 4-hydroxy DET could well inhibit the N-methylating enzyme (is it one dependant upon methionine as the methyl donator?). I suppose that if it is that non-specific it would be possible to add ethionine (S-ethyl cysteine) to the growing medium and end up with 4-hydroxy DET without ever having to add DET (which is good considering that DET is scarce due to its controlled status).

 

[yaesutom]

^^^

hopefully me, but don't hold back yourself!

Anyway...


If I added plenty of potassium phosphate like this, into my mimosa/DMT jars (or tryptamine jars), you think i'll get more psilocybin out? Would this be correct (good way to add more phosphate)?

 

[fastandbulbous]

^ You will not get more psilocybin out in terms of the absolute amount you'll get, but the ratio of psilocybin to psilocin will be increased. Just don't add too much potassium dihydrogen phosphate as it'll alter the pH of the growth medium (solns of said salt have a pH well below pH7 if I remember correctly)

 

[yaesutom]

Yeah i'd do some sort of half calculation figure out some range of expected psilocin output by guess and really it wouldnt take much of this stuff to add anyway, i'd only want to add about enough to be available for each psilocin molecule, which isn't much and probably won't alter the ph much either, but i might fool around with this in a couple jars with mimosa added, just to see if more psilocybin can be made (a more stable super shroom) / one with a little added one with more etc.

On chemfinder.com says the ph is around 4-5, all I did was add some KOH to water, and add phosphoric acid until the ph was around there, i'll pour some out and let it evaporate later today.

 

[yaesutom]

Heh.. alright, picked up some jars tonight, got some rye berries, some old spores, haven't done this in years but i'll try innoculating a jar with 4% honey 96% water, use that for a mycellium source and suck some of that up to innoculate some jars of just rye/mimosa bark first to make sure I got everything straight as far as substrate/water etc goes..

THEN.. the most facinating part of this to me is when i'll add different tryptamines to jars to see what comes out . I'm 99% sure that simply adding mimosa bark alone will produce some amazingly potent mushrooms, but i'm more interested now in throwing some 5-MeO-DMT into a jar, if that works to get 4-HO-5... then maybe some 5-MeO-MiPT/DiPT/DET/DPT.. etc.

THEN.. (or at the same time with different jar(s) ) AMT, i'm guessing not just 4-HO-AMT will come out but some methylation might occur, 4-HO-a,N-DMT / 4-HO-a,N-TMT. If the above 5-MeO's do work i'll try adding 5-MeO-AMT also w00t w00t!

Also i'll add potassium phosphate in varying amounts first to the mimosa/DMT jars to see if more 4-phosphoryloxy-.. 's come out, - I have a bunch of that in a baggie which I just dumped into a jar with water and checked the ph, its about neutral so I don't think it would affect the growth that much / at all especially adding such small amounts.

I can't find which Tihkal entry but somewhere Shulgin says psilocybin might have to convert first into psilocin before entering the brain - but the acetate ester doesn't, something about the phosphoryloxy group that won't allow it to cross the BBB?

Well we know that 4-AcO-MiPT for example is a much different drug than 4-HO-MiPT, if it was/is possible for 4-phosphoryloxy-MiPT to get into the brain like the acetoxy then i'd assume it would be another different drug. Any idea's? ... could toss some MiPT into the substrate and enough potassium phosphate possibly getting a good amount of 4-PO-MiPT out.. etc..

--edit: Forgot to ask how easily something like DALT could be made? Heard it can be made from tryptamine..

 

[fastandbulbous]

Well we know that 4-AcO-MiPT for example is a much different drug than 4-HO-MiPT, if it was/is possible for 4-phosphoryloxy-MiPT to get into the brain like the acetoxy then i'd assume it would be another different drug. Any idea's? ... could toss some MiPT into the substrate and enough potassium phosphate possibly getting a good amount of 4-PO-MiPT out.. etc..

--edit: Forgot to ask how easily something like DALT could be made? Heard it can be made from tryptamine..

I think 4-phosphoryloxy MiPT wouldn't cross the BBB because the phosphate ester has too many ionizable hydrogen ions left (which massively reduces lipid solubility). I've got a feeling that the 4-acetoxy ester does cross the BBB more efficiently than the free OH group as the free OH can ionize to a phenoxide type ion - the increased polarity reducing lipid solubility. I think it also make a difference in that (from what I've heard) 4-acetoxy esters have a much smoother coming-up period, the free OH compounds being characterized by a stop-start nature (producing a 'jerky' and more anxiety prone coming-up).


DALT is very easy to produce from tryptamine base, but in keeping with BL rules, I'll PM that to you

 

[evlove]

I am certainly all for this type of research, but I really wonder about bioassays in which the user claims to be able to discriminate between a potential psychedelic combination of 4-HO-DMT + let's say DIPT vs a combination of these two plus 4-HO-DIPT vs 4-HO-DMT on its own. Lots of variables and each makes you trip, making conclusions pretty difficult without additional analysis.

 

[yaesutom]

^^^ Oh I intend to have samples from all this sent to a lab and have an analysis if you missed that, i'm not quite sure what your saying, but i'm also tripping

 

[Psychedelics_r_best]

How would you sterilize the root bark before adding it to the substrate to avoid contamination?
Since heat would break down the dmt wouldnt there be no way to sterilize the root bark?