Mushroom Extraction & Questions

[Solipsis]

This week I've been working on a Cubensis mushroom extraction, I used half of the material to be extracted with methanol and the other half with 99.8% isopropanol.

I did 2 long pulls (24 hrs each) at room temp and each pull was followed by an extra elution wash of the mushroom material.

The methanol extract solution was noticeably darker in color, I don't know whether this is because of water present in the methanol which oxidized some indoles or because there is a more 'complete' extraction of principles.

I filtered using a Büchner funnel and big filters from a professional coffeemaking machine. I cut out round filter papers but some very fine particulate matter slipped through. Tried filtering through glassfilter at the end, but apparently using vacuum from an aspirator on a por. 4 glassfilter is too much too ask (Can someone confirm that for question #1?). So I filtered a final time through a double filter, the top one uncut so that nothing could circumvent it at the edges. This produced reasonably clear solutions.

Each is now evaporating in an oven dish at room temp, in my mushroom cultivation mini-greenhouse (it is what is called a Martha), which has automatic ventilation that goes through a small chimney.

Something weird happened when I was filtering: before discarding the mushroom material I pooled it in the Büchner funnel and used IPA as an eluens even though I was collecting the methanol extract at that point. When the IPA was added to the MeOH there was a lot of precipitate, a creamy white flaky material. The amount was way too much too be alkaloids (I am counting on about 300 mg of alkaloids max and this was just massive flaking) and besides, after checking it did not seem soluble upon adding IPA to just that precipitate which of course matches the other observations. I know the alkaloids are less soluble in IPA than in MeOH but not that insoluble...
It is possible this was the sugars that from what I understand can be extracted along, apparently they are insoluble in IPA? (^^ Question #2)

If that is true, it might be useful to know that while MeOH is the ideal extraction solvent, adding IPA at the end could crash out sugars which may result in a more pure product.

Then again, this is not always necessary, depending on the method / tek used... if you are recrystallizing anyway there would be no point.

I read this:
http://www.erowid.org/archive/rhodium/chemistry/psilocin.extraction.html

..and became curious about the apparently neat way to get a crystal product. Unfortunately I don't have any ether on hand, but that could be arranged.
OTOH, doing an extraction after all this would basically be doing the job twice, although working with crude extract is probably a lot quicker and more practical.

Other than that I'm very interesting in freeze-precip recrystallisation options... anyone? (Question #3)
I know that I could try recrystallizing from boiling water but it seems pretty tricky. Obviously freeze-precipitation with water is not an option.

P.S. I'm not looking for yield, but mainly for crystalline product.

 

[sekio]

If you lack analytical gear like TLC and a set of stains, you're working blind and should only consider this a novelty.

Methanol is pretty polar and will extract a lot more colored oxidised material than isopropanol will. The precipitate you noticed from adding IPR to the methanol extract was probably sugars or starches or some other polar component that was extracted and then crashed out of solution by adding isopropyl alcohol.

If psilocybin/psilocin or their salts have low solubility in isopropanol (I didn't check), maybe that's the precipitate. Sometimes compounds are soluble only in a pure solvent and adding a little bit of an "antisolvent" to the mix will cause them to precipitate.

You really need some method of figuring out what you end up with, beyond your five senses.

 

[MyExcuse]

I had a discussion about this very topic over at shroomery and I was informed that the most reliable method of cleaning up a crude psilocybe extract would be to use ion exchange resin.

http://www.shroomery.org/forums/showflat.php/Number/18083782#18083782

supposedly it's cheap and readily available. Also, there is a patent on extracting morphine from crude opium using ion exchange resin which could possibly be applied to this extraction process as well. I'll find it tomorrow.

Lots of info in that thread. If you happen to have good university access to journal articles, you could probably get more info than I could from the articles posted therein.

 

[Solipsis]

Thanks.

Yes I do have TLC plates and UV, but no stains (yet). I should probably make or buy Ehrlich's for this, don't think ninhydrin is quite necessary. Also I think checking for indoles is more selective here and thus better.
Water stains compounds involved here blue, I wonder if that alone would be a feasable indicator.
As a rule you are absolutely right about working blind, just thought that as with DMT extraction it is not quite the mystery what is happening compared to the average synthetic reaction.

Great tip myexcuse! Having a crude chromatography option would be very nice, not sure if it is like that. Can't access the shroomery thread though, searching for it only shows Amanita extraction discussion. But I will check out the uses of the material anyway.

I still wonder about freeze precipitation and other recrystallisation options. If they are no good I don't understand why.

 

[Abject]

Here are the shroomery posts:

Spoiler: long

well, 1-butanol is used as the major solvent in a tertiary solvent system for TLC separation of psilocin/psilocybin,
the others being ammonia and chloroform, iirc. ethyl acetate may be used in place of chloroform.
using a similar solvent system, one may be able to separate the alks from proteins in a column of silica.

alcohols in general aren't particularly selective, proteins will precipitate out of them, especially if the alcohol is cold

What do you think may be more selective?

Per merk, both are insoluble in chloroform, so ethyl acetate would be similar.

ahh you're correct. must've been thinking of another alkaloid
http://www.erowid.org/archive/rhodium/pdf/psilo.analysis.bigwood.pdf

1-butanol:acetic acid:water (12:3:5)

keep in mind, this is against a stationary phase. You could probably
apply the separation against some cellulose, like cotton balls.

I'll just post this here cuz it's relevant. (links to "Obtaining psilocybin and psilocin from fungal material. R Heim, A Hofmann , A Brack, H Kobel, R Cailleux. US Patent 3183172" You won't be able to download, pm me if you really want it is dated 1965)

The last post will have to be made in a second post as it has quotes in it.

 

[Abject]

I'm curious what the product would be like without chromatography? I would imagine the extra purification steps would yield a crude powder, but I could be wrong.

There are two interesting parts:

24.2 parts by weight of dried fruit bodies are thoroughly ground and extracted once with 300 parts by volume and then three times with 150 parts by volume of methanol, each time at room temperature for 30 minutes. The extracts are combined and evaporated to dryness under reduced pressure. The residue (6 parts by weight) is defatted by rubbing well four times 125 parts by volume of petroleum ether and three more times with 50 parts by volume of chloroform each time, the chloroform containing 10% of ethanol. The undissolved residue (59 parts by weight) is dissolved in 5 parts by volume of water. From this solution other products are precipitated by slowly adding 50 parts by volume of absolute ethanol so that the active substance accumulates in the solution. The purification is repeated two or three times in the same manner. The decanted solutions are combined and evaporated to dryness in vacuum.

1) 24.2 grams of mushroom are ground to a fine powder
2) Extract with 300ml methanol
4) Extract with 150ml methanol x3 times, each time for 30 minutes
5) Evaporate all extracts
6) Defat the residue with 125ml of ether x4 times
7) Defate the residue with 50ml of chloroform with 5ml ethanol x3 times
8) The residue is dissolved in 5ml of water
9) Add 50ml of ethanol and decant 4 times
10) Evaporate ethanol under vacuum

306 parts by weight of the dried fungal material are finely powdered and shaken 3 times with 500 parts by volume of chloroform each time and 3 more times with 500 parts by volume of chloroform containing 10% of ethanol. 2.8 parts by weight of inactive accompanying substance are thereby dissolved. The pre-extracted fungal material is thoroughly extracted once with 3000 parts by volume and 3 times with 1500 parts by volume of methanol each time. The combined extracts are evaporated to dryness under reduced pressure, a clear brown residue of 17.5 parts by weight remaining. In order to remove fatty impurities from this residue, it is taken up in 17.5 parts by weight of water and the suspension is extracted once with 500 parts by volume and twice with 250 parts by volume of petroleum ether each time. The petroleum ether solution contains 0.75 part by weight of inactive accompanying material. The residual aqueous solution is concentrated under reduced pressure to about 25 parts by volume and is then treated with 250 parts by volume of absolute ethanol, while being vigorously stirred. From the sticky precipitate so produced, the solution containing active material is separated by decantation. The precipitate is re-dissolved in a little water and treated with a tenfold quantity of absolute ethanol. This purification by precipitation is repeated twice with the residue. The solutions are combined and evaporated to dryness in vacuo. There remains a solid residue of 11.7 parts by weight containing the whole amount of active material.

1) Grind 306g of shrooms down to a fine powder
2) Mix and shake with 1,500ml Chloroform in 500ml increments (500ml x3)
3) Mix and shake with 1,500ml Chloroform mixed with 10% ethanol as in step 2
4) Mix the powder with 3,000ml Methanol
5) Mix the powder with 1,500ml Methanol 3 times (1,500ml x3)
6) Combine all extracts and evaporate
7) You'll be left with 17.5g of brown residue
8) Add 17.5ml of water to this material and mix it up
9) Extract with 500ml ether
10) Extract with 250ml ether two times
11) Combine and reduce 1,000ml ether down to 250ml ether
12) Add 250ml of absolute ethanol and stir vigorously
13) Decant the liquid leaving behind sticky residue
14) Dissolve sticky residue in a "little" water
15) Add 10x (the amount of water used) absolute ethanol
16) Repeat steps 13-15 twice
17) Evaporate all ethanol under vacuum


Somebody then replied with:
The first one will still leave an active goo.
The second one will not even be active.

Good luck, hope the info helps.

 

[sekio]

I think at that stage you might as well just be eating the mushrooms So much work. The crude extract would probably be mostly psilocybin and other polar stuff like sugars and amino acids.

I would imagine that there's plenty of procedures for getting psilocin and psilocybin out of the mushrooms in the literature. A simple methanol extraction is probably all you need. It would probably behoove you to work under a nitrogen atmosphere too, to limit oxidation of the indoles.

Still: if you want to have any sort of solid knowledge about your product you need to do a TLC or HPLC analysis.

 

[MyExcuse]

My postings in that thread were with the idea of extracting from P. Mexicana truffles. The quantity one can produce in small quarters, and with such little effort, is above anything that can be reliably stored for any substantial period of time. Extraction would be the best route of preserving and condensing the constituents.

With such an abundance of available starting material, I feel that a methanol extract would be a great starting point. However, if you do have such abundance of fungus available, why wouldn't you bother with the extra steps to purify your end product even at great loss?

If nothing more than for novelty.

I'm not familiar with ion-exchange resin's and their various properties. I'm lazy and like keeping things simple so I referenced an article related to morphine and the method used is similar to the one used to create Polish Heroin. Essentially they use ion-exchange resin to extract morphine from poppy straw. It involves highly-acidic ion-exchange resin and then extraction using ammonia. A hypothesis is, if you used the right solvents and adjusted the solution to the correct pH, you would be able to extract at least some of the psilocin. There is a specific range where psilocin is most soluble in solvents.

Solipsis, I personally keep an account on most of the major drug forums in full-standing (full viewing privileges) because you gain a greater collection of resources to gather from. If you don't have an active Shroomery account (I think you need like 30-50 posts), it's worth it.

 

[FunctionalOlfactio]

Solipsis,

Here is some additional info. on mushroom alkaloid extraction.
http://www.fanaticus.com/mycoalki.htm

 

[Solipsis]

Thank you, yes I have a Shroomery account and I am very familiar with the mycoalki site Although I have a hard time finding good instructions to go with those pics of crystals 'snowing' in the mother liquor. Even if I could get that to happen, it is probably important that I do a TLC and use synthetic psilocin as a reference. Because for all we know it is the sugars that are 'snowing'.
I don't have Rf values yet but they shouldn't be too hard to find.

Not entirely certain yet how I am going to do the purification but I have access to ethanol and other solvents. A chemist friend also said that sodium sulfate should work as a drying agent for alcohols, or better is to use 3Å mol sieves (4Å sieves are only júst able to absorb methanol). And assuming that methanol or ethanol are the ideal solvent to recrystallize.
The honey like product I have now would probably be more like hard caramel if it wasn't for the water residue.

A professor at the uni was asked about the matter and he claimed that the recrystallisation shouldn't be that hard at all, which leads me to believe that under the right circumstances good product could be made in a way similar to how DMT is commonly made.
Of course there are things to do or avoid because of the differences between indolol tryptamines and plain ones like DMT.
Seems like a good time to go hunting for psilocin / psilocybin solubility values for different solvents. :D And, I guess pick the one that has the biggest solubility difference when comparing them at different temperatures...

And @sekio: to answer your question / sentiments about the seemingly pointless work, yes this is about novelty and geekery.

 

[Abject]

My posts have disappeared
If something is glitching and this is a redundant posts please delete it

I was about to make a thread but boy am I glad I did a quick search

Is there a reason we were hypothesising methanol? I was quite a bit more unaware 3 years ago and would much prefer metho (denatured alcohol) over iso or methanol.
Winter has started where I live and I'm not that keen on drying all my finds this season. I've eaten everything I've found so far fresh cause we're only just starting but I'll have kilos of shrooms in pretty soon

 

[Solipsis]

You should still dry them if you want to extract, so as not to have any water present during the process / in your product.

Methanol is considered an ideal solvent for this extraction cause of the polarity and solubilities.

To freeze them for long term storage (hypothetically), again you should first dry them. I can't really think of a way around the drying, and honestly it's so easy lol... just lay them out properly to first do the air drying... then for the final drying enclose them in a box with desiccant.

You still do have posts in this thread. You lost very recent ones? Did you make them on mobile and try to edit perhaps?