Perhaps it would be best to still focus on maybe 5 liter cultures thus far.
In fact, what is the best "structure" for highest alkaloid levels?
Most all of them that I read use a Erlenmeyer flask on a shaker, and often it is relatively large compared to the culture, such as 50mls of culture to 250ml flask.

What sort of reaction vessels are you guys thinking would be good?

I have seen polypropylene 5 gallon buckets that hermetically, but it isn't like it would be easy to sterilize that, though, smaller polypropylene buckets could be obtained, perhaps?

They also sell polypropylene gallon jugs for water in certain places, but I get the idea that ventilation will become an issue, competing with batch size and autoclave-ability. Even one liter starts to seem a bit tricky to do...

Seems like it would be hard to do all of that under sterile conditions. Might be more ideal to have many small reaction vessels that can be autoclaved with the nutrient solution already, and than have them good to go for the next step, being inoculation.

Good points, I just don't like the idea of having unwanted bacteria/yeast in liquid cultures. However their growth would be really slow as you mentioned a lot of good points.
Though, it isn't quite the same as a bulk substrate for several reasons, as the mycelium here isn't going to do much more vegetative growth, as mycelium does in bulk substrates, in addition to it being liquid - liquid allows for contaminants to travel/spread through out the whole thing way faster.

Probably making it so it is "essentially" sterile might do the trick.

I used to use that for agar, than one time it turned brown and never solidifed, other times it had inhibited growth. It seems very hard to get it so the peroxide is destroyed without it destroying something else..- at least with agar, with this it could work much better.

Also it still requires heat. However perhaps adding an insoluble peroxide decomp. catalyst (MnO2?) might be worth considering.

I guess just a big 5 liter flask would do the trick for at least 1 liter of solution though, thats probably the best starting point I guess.. That can easily bring things up to boiling, and if it were pre-sterilized, you'd only really have to worry about endospores enter the solution than germinating.


Also could do stuff like boil it, incubate it, than boil it again - this causes the often temperature resistant endospores to germinate - going into a vegetative growth, and than kills those as they are susceptible to death by heat. I forget what this is called, something that starts with a "T"

Oh yep, called tyndallization!

Fractional sterilization or tyndallization is a method used to destroy bacteria and endospores in preparation of grain spawn(rye, wheat, millet, birdseed...) and agar, which requires no pressure cooker.
In this case, the jars fitted with a filter disc or a polyfill lid filter are boiled or steamed at 212°F (100°C) for 30 min in a pot with lid, three days in a row. Between the boiling steps the jars are kept warm, around 30°C(but room temperature will work too), to allow the remaining endospores to germinate.
The basic principle behind this method is that any resistant endospores will germinate after the first heating and therefore be susceptible to killing during the second and third heating.

Timetable of the tyndallization (=fractional sterilisation) process

1) Steam heating to 100 °C for 30 min
Vegetative cells are destroyed but endospores survive

2) Incubate at 30°C-37°C overnight
Most bacterial endospores germinate

3) Second heat treatment, 100 °C, 30 min
Germinated endospores are killed.

4) Second incubation at 30°C-37 °C overnight
Remaining endospores germinate

5) Third heat treatment, 100 °C, 60 min
Last remaining germinated endospores are killed

It might not need to be totally sterile, but I bet bacteria can out grow mycelium in a culture that is meant not for the mycelium to grow, but to produce alkaloids.
Could be a problem easily at day 10 at ideal temps and aeration.

Good idea with the 4/5 or whatever in addition to added bubbles - probably would be pretty easy to do with a fish air pump, a long tube of sterile cotton/polyfill, and an air bubbler.

In addition a magnetic stirrer might help every now and than to kick up mycelium and mix air bubbles better.

Imagine this:
[youtube]http://www.youtube.com/watch?v=n7cKbIxnTdA[/youtube]

Though, that much agitation isn't good I wouldn't think... Does anyone have papers on alkaloid content vs oxygen levels vs agitation, etc?

I know more oxygen the better, but the agitation might ruin it and you'd need to find a "happy" middle ground.
 

Interesting, so garicinol and those others (w51, w56) induce alkaloid formation?
I only had time to skim it, I'll look more into that later - but those seem unpractical to use I suppose.

I am very interested in this... I don't have the knowledge of biochemistry or genetics to make any conclusions from this research except that its a potential game changer... Can anyone clarify which compounds would work for our needs?  Keeping my ears percked up for this one...

From all the other ergot files I have, they always specify it in ug/ml - this doesn't seem too but it does say ug as a concentration. It makes sense to me that it would be ug/ml - being 50mg per liter, that isn't bad at all. Though, if you were to buy the pure chemical, it would be very expensive. Hopefully other more OTC/cheap chemicals, such as curcumin or EGCG work as well. Though, that might be wishful thinking.

Just as an emphasis:

Can anyone clarify which compounds would work for our needs?

Well, you keep it as simple as possible.

I am going to explore all the options and see what sort of gems can be found.. IMO it is important to look at everything and evaluate the situation.. plus - whats wrong with aiding everyone with tons of information?

While life never seems to work this way, perhaps any crude nutrient solution, some cheap 90% EGCG, and any wild strain or worse of a ergot culture could be mixed up to have some high yields of ergot alkaloids. You never know, and its all about the possibilities...
It isn't like arcemone knew about the garcinol, or even had all the references.

Perhaps just going the extra mile could really put a little umph in the whole ordeal...

I think the real probably would be to do this cheaply, and "semi-quantitative" - it would be very very hard to determine after a certain point which solution had the most ergot alkaloids in it using something as simple as the Van Urk...