That's some phenomenal research! I don't pretend to completely understand everything you did there, as that I have almost no biological training or knowledge, but I think that in general I'm a pretty intelligent dude, and have some ?'s that are directly related to your posted experiment, which perhaps you might be able to answer:

1: Where might I find phytic acid (hexaphosphoinositol) and how do I basifiy to a near-neutral pH (i.e. what basifying agent would be added)

2: What effect, if any, does the increased content of the idole alkaloids psilocybine and its demethylated derivative baeocystine as constrained by the hexaphosphoinositol on advanced mycelial development and the development of primordia and carpophores?

3: Is hexaphosphoinositol a toxic substance? Do the alkaloids need to be extracted from the phytic acid with methanol or can the amalgam be safely introduced to a human system?

4: Are there other phosphor "store houses" that might be easily adapted to the use of the layman in increasing alkaloid content in thallophytic plants?

I guess, in short, is there a "tek" for us home-growin yokels with no access to lab mills, flash dryers, and centrifuges?

Thanks for posting, Doc, I know that 99% of the posts here are well below your superior understanding of the biological processes at play here, but It's people like you and posts like yours that might knock it up a notch here at the shroomery!

Mr. G, I dont get involved usually with you big-wiggs, since I dont know shit about what came from who or where, but I do think that if you want to convince someone of something you need more than words, and rude foceful words will always work against your goal. Put up a website, I'd love to see the sources for your scoop on what b+ is.

Peace Out
S7!

------------------
"Keep your eyes on the road and your hands upon the wheel..."
JM

"Mr Majestyk's phytochemical manipulation of Psilocybe (Stropharia) cubensis strains"

I. Overview

Spores of a Central American strain of Psilocybe (Stropharia) cubensis
were germinated on a standard potato-dextrose agar medium. Several
dikaryotic strains were isolated and the most vigorous candidates chosen
for an experiment to ascertain what effect, if any, the addition of
phytic acid (hexaphosphoinositol) would have on the accumulation of the
indole alkaloid psilocybine and its demethylated derivative baeocystine.

Phytic acid is a substance commonly found in seeds and is a storehouse
for phosphate, an essential nutrient for the biosynthetic production of
O-phosphoryl-4-hydroxy-N,N-dimethyltryptamine
(psilocybine) and desmethyl psilocybine (baeocystine).

II. Procedure

Three genetically distinct, rhizomorphic strains were chosen and grown
in 1000 mL Fernbach flasks containing 550 mL liquid Emmon's medium.
Phytic acid (basified to a near-neutral pH) was added incrementally to
this medium in amounts ranging from 10-35 mg. The cultures were
incubated at 25C and aerated on a Gump rotary shaker (Model M) at 240
rpm. The resultant mycelial pellets, separated from the culture medium
by suction filtration on a Buchner funnel, were dried in a forced-air
drying oven at 45C for a minimum of 48 hrs and subsequently stored in a
dessicator over anhydrous calcium chloride. After determining the pH
values of the filtered culture medium, the filtrates were evaporated to
dryness in a flash evaporator at 40C under reduced pressure. The
residues were carefully extracted with 2-mL portions of absolute
methanol to dissolve the psilocybine (and baeocystine, if present); the
solutions were stored in a refrigerator until subjected to thin layer
chramatographic (TLC) analyses.

After drying, the mycelial pellets from the individual culture flasks
were weighed and then reduced to a #60 powder in a Wiley laboratory
mill. Ten mL of methanol was added to each sample of powdered material
in a 25-mL Erlenmyer flask. After shaking for eight hours, the resulting
slurry was centrifuged and the supernatant liquids decanted into a 50-mL
round bottom flask. The marc was treated in an identical manner two
additional times, after which the combined methanol extracts were
evaporated to a small volume (1-2 mL) in a rotary film evaporator at 40C
under reduced pressure. The concentrated extract was transferred to a
5-mL volumetric flask with a microdropper. Residual material in the
evaporation flask was removed by a methanol wash, added to the
volumetric flask, and the final volume adjusted to 5 mL. Extracts
prepared in this manner were labeled and refrigerated until personal
bioassay.

III. Chromatographic Results.

Thin layer chromatographic procedure was determined suitable for rapid,
sensitive detection/evaluation of the hydroxyindole metabolites. Weighed
portions (usually 50 mg)
of the dried, powdered mycelia were extracted by shaking with 5 mL
methanol in 12-mL glass-stoppered centrifuge tubes for 1 hr. The
mixtures were centrifuged and the clear supernatant solutions were
removed with pipets. The marcs were extracted with second 5-mL volumes
of methanol. Preliminary results detected:

1. Psilocybine in amounts ranging from 1.9% to 2.8% (by wt vol).
2. Baeocystine in amounts ranging from .64% to 1.2% (by wt vol).
3. Trace amounts, e.g., .010-.023% of a new monoethyl analog similar to
psilocine.

The amounts of psilocybine and baeocystine are the highest ever detected
in any species. The structure of the new monoethyl analog is still under
study; preliminary tests reveal a hydroxyindole compound with a
substituted ethyl group in the 4th or 5th position of the aromatic
(benzene) side chain.

IV. Personal Bioassay.

The psychedelic effects experienced after ingesting the extracts were
nearly identical to those of psilocine. However, the duration of the
trip was much longer--on the order of
14 hrs(!) This increase in activity is thought to be due to the amount
of baeocystine and the possible synergistic effect of baeocystine and
the new monoethyl analog.

Mr Majestyk, PhD

------------------
The real Mr. G

Workman is the the real deal.
Hoth out

------------------
Dogs fucked the Pope, no fault of mine.

Mr. G's ridiculous comment about needing a microscope to identify the B+ is analogous to someone needing a microscope to distinguish a cocker spaniel from a cat, or a Pleurotus ostreatus from an Amanita pantherina.

[This message has been edited by Mr Majestyk (edited April 30, 1999).]

rhubarb-

S7!

------------------
Dogs fucked the Pope, no fault of mine.

I've got a few honest question's about the colchicine treatment, this isn't intended to be inflammatory I'm just looking for some answers.

I've seen it stated that colchicine will not bind to fungal tubulins but I've seen passing references to colchicine binding sites on tubulins in Neurospora and Coprinus species. Can you give me any ref's that can clear up this contradiction?

In your experiments how did you confirm that the chromosomes were doubled? Did you do a count? If so, how many chromosomes were in each cell nucleus?

Thanks in advance for your answers.