Psilocybe growth and extraction.

[shienar]

Hey everyone,

Im about to try something a little bit strange. Im going to try and use spores extract from a mushroom to grow on agar, grain, liquid in a fermentation. This is the first part i was wondering if any of you have tried this and can recommend a media which is suited. Im quite experienced at fermenting organism and marine fungii for natural products so i was looking for a few pointers as to what is should try to optimise it on.

The second part is not so easy.

Im wondering about the general extraction methods people have used. Also whether anyone has a UV spectrum of the targets in this guys.

Thanks any advice would be helpful and ill report back in a few weeks.

Cheers

 

[Beenhead]

Seems like if you dont have access to an autoclave you would have a load of a time keeping contamination down. Are you talking about cultureing spores on Agarose? And the fermentaion, what are you hoping to get out of this?

 

[shienar]

Its not equipment thats the problem i have autoclaves, temp control rooms and access to a 500 lt fermenter but im trying to extract the active metabolites from a few strains of mushrooms, but ill be trying to grow it on a media that i have due to the fact that i cant grow mushrooms but have access to spores.

Thanks for the reply

 

[vecktor]

you might get more joy posting this question over at the shroomery.

I cannot help more without more info, are you running a suspended mycelial culture, are you planning to run on stream monitoring of the metabolites? I assume you don't plan to use UV to identify the metabolites, the UV spectra are available but are pretty useless for identification.
the density of the myceliumm in suspended culture is very low far lower then in mushroom fruiting bodies so the absolute levels of psilocybin is usually also low.

I assume you are not growing mushrooms because of a legal restriction however you should remember many countries ban fungal material containing psilocin etc, and a mycelial culture would be a fungal material.

 

[Carsick]

I remember seeing an article about something very similar on shroomery a while back.

 

[Beenhead]

Theres TLC and HPLC;
for purification/ID

For LSD, I believe the correct diluents were Hexane and Chloroform.

Absorption maximum: uv max (methanol): 220, 267, 290 nm (log e 4.6, 3.8, 3.6)
for Psilocybin

Absorption maximum: uv max: 222, 260, 267, 283, 293 nm (log e 4.6, 3.7, 3.8, 3.7, 3.6)
for Psilocin

What are you going to use for Purifying the metabolites? HPLC would work.. the metabolites you want shouldbe in just a couple fractions that absorb in those wavelengths. Im sure you know how to use HPLC though!

ANd if you can find the Rf Factor for your Wanteds, you can use TLC to purify them, just cut out the paper where your compound migrates too, we use SDS PAge Gels for this purpose, we run the gel and when the protein reaches the isoelectirc point you stain with Comassie Blue, and cut out the band. Not what its intended for but it works!

I dont know much about fermentation, though I grow E coli, for Immunoprecipitation in liquid media: Tryptone, and a few other things. I could look up the protocol if that would be of any help, though it is for bacteria. I dont work with Fungi!

 

[mumbles7]

You can find some pretty detailed info on growing cube cultures, supplementing in tryptamine and extracting the desired alkaloids in Otto Snow's LSD-25 & Tryptamine Syntheses.

 

[shienar]

Excellent.

My current plan is to do a MA liquid fermentation then C18 the LON and take the mycellia to a acetone extraction and a ethyl acetate partition trying to get all my material into under 4 mls to mega prep then c18 recover the relavent fractions.

I personnally hate hexane so i think methanol chloroform should work (its what i normally use for silica column and tlc plates, 10meoh/90chcl3)

What i dont know is how to translate "Absorption maximum: uv max (methanol): 220, 267, 290 nm (log e 4.6, 3.8, 3.6)
for Psilocybin

Absorption maximum: uv max: 222, 260, 267, 283, 293 nm (log e 4.6, 3.7, 3.8, 3.7, 3.6)
for Psilocin" into what it actually looks like. Ive only been working in chemistry for about 18 months.

Thanks for the input