I am rather hasty at the moment but stumbled onto this that might be of importance :

http://www.springerlink....ontent/q8w280384570n510/



I wonder if pectinases from wine shops could come handy...

Here is the full article. While it does not refer to seeds, the pectin contained is hydrolysable by pectinases. It is mentioned as having good gelling capacities and being able to form solutions with high viscosity.

The reason high viscosity troubled me is not only because it makes filtering difficult - i am afraid that high viscosity might interfere with alkaloid precipitation either in salt addition step ("manske" ) or while basifying the liquid.

Very interesting, pectinase could be a way to make extraction easier. Thanks!

And because untested theories are only wet dreams someone i know has decided to try a small experiment. What has been located so far is a pectinase called Depectil EasyCLAR 32 FCE and here is the manual describing its action: http://www.martinvialatt...FCE_2-045_GB_054_10_.pdf

It has hemicellulase, endo and exo poly galactonurase, pectin methylesterase, and pectin lyase actions.

I think that if pectins are the problem this will make short work of them. If anyone though care to propose another enzyme or a combination, please tell so.

Hmm.. cellulase might be useful to break down the cell walls???
This might reduce the need to use NaOH for this purpose as in the STB tek, especially nowadays when pre-powdered MHRB is banned here in this small flat country.

Just a brainfart...

Slightly out-off topic:

Yes propably hemicellulase and cellulase could help break cell walls - such enzymes along with "pectinases" are used in fruit beverage industries both to clarify the juice and also to extract the juice from fruits! Such enzymes are secreted by funghi (and if i am correct bacteria also) to break down cell walls and turn fruits and plants into a tasty (for them), easily digestable...snot!

What i do not know though is if ligninases could be more usefull, since bark and generally secondary growth is usually high in lignin. One could experiment here with incubating the bark with the enzymes of choice and then move on to other procedures. If i had to choose i would go for a ligninase, but i do not know how easily acquirable would this be. If one has access to hemicellulase or mixtures of it with other enzymes (even better if they are widely commercially available) now it would be a good time to try putting them to use.

On an unrelated sidenote, if the shift from "thoroughly powdered bark" to "unpowdered bark" causes problems for the "hobbyist" imagine what kinds of problems will unavailability of the bark will cause. Times might come where a simple STB of a high-yielding, easily accessible material will be a thing of the past, giving hardons to newbies and making "old folks" nostalgic to the point of tears. Till then, it would be clever if energy is spent looking into alternatives before one is forced to do so.


On topic:

300 grams of whole Peganum harmala seeds were boiled in approximately 1 liter of water (x3) for half an hour (time counting from boil start) and the almost three liters* of water obtained where reduced down to 1 liter. No acids where used in the procedure, alkaloids propably exist in their natural salt forms**.

This 1 liter was split to two 500 ml portions (jars), as it was, no filtering. When the temperature was measured to be around 30 C , 500 mg of EasyCLAR*** was added in one of the containers while the other one was left without addition of enzyme. Both jars were shaken vigorously, and will be left overnight in room temperature ( approximately 23-24 C).

Next, both solutions will be filtered through loosely packed cotton and also coffee filter**** if time allows.

* Seeds seem to have absorbed quite some water and pressing them does not seem to retrieve all the water.

** pH could play a role as well as temperature: While those enzymes act in an acidic enviroment, the experimented did not want to introduce a factor and acidify the water although i suppose it could be done , given that extremes are not reached. Enzyme addition also happened having in mind that the solution should not have a temperature that would deactivate the enzyme. 30 C, "seemed fine" given what the experimenter knows about enzymes.

*** Given the manual for the amount of liquid one has (500 ml), only 20 mg of EasyCLAR should be used (described as the "maximum" addition of enzyme, 4g to 100 l ). The experimenter chose a higher amount since availability of the product is not an issue (100g in possession) and since higher amount of enzyme will speed up the reaction up to a point. This point is the point where all the substrate is "bound" to enzyme molecules, at which further enzyme addition does not influence the rate of reaction.

**** The experimenter would rather do it with whatman filters under vacuum but at the time being this is not an option, plus this is a small "proof of concept" experiment rather that a "full extraction".

Further ideas: One could also let the seeds incubate in the enzyme before extracting. Maybe this could aid extraction and destroy pectins in one step. The only problem that could possibly be faced is more starch being liberated -still this is just a guess.

Will update once i receive more data on the subject at hand. All ideas are welcome, also thanks for making it a "sticky"!

Great work, thanks - I can't wait for the results!


Amylase in some spit could fix that

Seems like we might be onto something here.

The two jars were left overnight and compared today. The untreated one had sediment and the supernatant liquid remained "murky" looking. The treated one had more sediment on the bottom, but the supernatant looked less "murky" as if part of what made it "cloudy" had precipitated. Still, the liquid was not transparent it looked black like strong coffee.

It was chosen than only the supernatant will be filtered for the initial stage so as less clogging of the cotton filter takes place*. The treated jar here performed better , with better flow and almost no need to change the cotton , whereas the untreated one clogged twice and had to have to cotton filter replaced.

The resulting liquids were filtered through coffee filter paper (100 ml of each, to compare flow). Still the treated supernatant seemed to perform better. Both of them went slower compared to pure water, but the untreated one after a point started going painstakingly slow. The treated supernatant seemed to leave less precipitate on the filter paper.

*Trying to redissolve the sediment in acidic water did not work, meaning that it is insoluble to water, although i would advise that it should be washed since the water seems to have pulled something (part of the alkaloids?). If everything went as expected the greater quantity of the sediment could be attributed to pectate which is insoluble to water.

To conclude on what we have so far it seems that the seeds too might contain pectin or other soluble polysaccharides that can increase viscocity of the resulting brew. Pectinases (those used) seemed to clarify the brew, increase sedimentation making the supernatant clearer, with less suspended particles. Note that the experimenter tried the above as a quick experiment, the whole procedure might benefit from a longer incubation time.

A side idea : Has anyone tried concentrating peganum harmala tea to a minimum amount and adding copious amounts of organic solvent? If so , was there a precipitate and how could it be described (what where its properties?). From what i see online they seem to be insoluble in organic solvents, so that could serve as a second test or even further this sediment from organic solvent isolated and checked if it has gelling properties and if it can be digested by pectinases.

This sounds very promising. Pectinase is on my list of things to get!

Two small additions that might come usefull to anyone interested in the above methods or technical information about enzymes

Pectinase Database : Good information on pectinases and pectin like compounds. "know your beast, know how to defeat it"

BRENDA: Simply put, Sexy as enzymic activity! A gem ! BRENDA is "The Comprehensive Enzyme Information System". Give an enzyme name (or even better its EC number, check the product PDF i have supplied to see what i mean) and BRENDA will bring to you lots of information about the enzyme. At what temperature the enzyme works best? Is there an optimal pH? What is the reaction it catalyses and what are the products? What do different homologues of this enzyme do from different organisms? BRENDA has the answer, or at least tries to! NB: It is usefull if you know the origin of the enzyme you are using, for example mine are from Aspergilus sp., BRENDA has an option to search only for data of enzyme pertaining to a specific organism which can come really handy.

Also, whatever product you are about to buy, or if you are in the process of choosing, please see its technical brochure or MSDS to see what you are buying so you know what your product does.

That's a great resource. Top notch contribution, thanks again!

Enzymes like pectinase, ligninase and cellulase could make life easier for people with innocent little hobbies like plant extraction

To anyone concerned: the method that dealt with the Peganum mucilage was also tried in two tabs full of cactus snot. Success! It made short work of them! More information here.

So would this be this issue of my harmala extraction being extremely viscous? I let it sit overnight to settle, but it just settled in many different layers of precipitates. When I transferred it to a larger container (to add more lye-solution, I thought it wasn't based enough), it was the consistency of jell-o.... and the additional lye-water didn't help...

Where does one get these enzymes? healthfood store? Is that even the issue?

Possibly. It sounds like you might have reduced too much and maybe not filtered enough. After reducing I let it sit a few days to let the brown stuff settle on the bottom, which seems to clarify it some and help with filtering.

Take some of it and dilute it with plain water and see what happens. If nothing happens, add more water. Generally, a quart jar is enough to settle and decant crystals from 100-200g of seeds, with 0.5 to 1.5 inches of settled crystals. The first basing and water washes usually give very fluffy crystals that can take a day or so to settle out. If you're doing around a pound, you should be working with around a gallon of water. I'm not saying it can't be done with less water, but this is what I've seen in my experience.

Ok so you're saying to try taking it bit by bit and diluting, settling, and decanting it in a quart jar?

The extraction was maybe around 200 g of harmala, I didn't weigh it...

I shouldhave let it settle for a day or so after reducing... that would have helped, I just moved right along trying to get it moving along... which I guess is rather contrary to the Tao of Rue...

Yes, i think that the viscosity can be caused by pectin - thats what pectin does, that why they use it in marmelades! It does hurt to try enzymes, although note that if you add lye (increse pH) they might not work.

You can get the enzymes from winemaking shops, shops with supplies for wine makers and home brewers.

Interesting !
I battled with my precipitates some days ago ;
As said above, after freebasing, i got a multi layer stuff, hard to work with.
I found that sifoning what i can, then redisolving in huge quantity of water helped a lot. Instead of the liquid mud i got a more "sandy" precipitate.

But after re-salting for the Manske, i filtered and there was an horrible goo on all the filters. This goo was not soluble in hot acetic water, and didn't seem to "yield" much alk at all (judging by the color of the water ).
I didn't expected that much mess at that stage, especially after multiple filtration in first step.

All of this to say, i'm really happy to see i'm not the only one struggling and we all looking for nice solutions (literally)

Something of a footnote here, of perhaps limited utility due to its fortuitous and uncontrolled nature:
A while back during a repeated extraction process from syrian rue seeds there was an annoying gloopiness to the resulting tea. In order to minimise particulates in the brew, it was allowed to settle at room temperature for a number of days.

During the settling time the weather turned significantly warmer; this resulted in mould colonies developing on and in the brew; a lucky observation during filtering showed that the mould involved was producing a potent cellulase - the paper filter disintegrated rapidly during a filtering attempt!

The growth of the mould was stopped completely by acidification of the brew to 1% w/v with acetic acid. Subsequent filtration proceeded much more smoothly with the greatly reduced viscosity from the removal of the presumably cellulose-like pectins.

I would speculate that at least some of the pectins from Syrian rue seeds are hemicellulose-like and form a colloidal solution, and that these are amenable to hydrolysis by fungal cellulase.

I'm now suspicious of the notion that pectin, itself, is what is interfering with our extractions.
I recently tried to isolate the vasicine from based tea solids [its a bronchodilator and, covid] I was not immediately successful so I checked the literature. In mature Peganum seed nearly all of the vasicine is actually there as an alkaloid glycoside, vasicine hooked to a chain of two glucose units. This is 10% of the seed weight.
Vasicine glycoside could explain why we have a gunk in there that has properties of an alkaloid, pectin, and a saponin all in one.

Yes, you could well be right. I just filtered a crude Manske (finally, the one that got the mould "treatment") and the filtrate is still remarkably gloopy and holds a froth for a good long while

The collected crystals were redissolved in distilled water, mainly because I was too lazy to scrape them out of the funnel, and something is causing that solution to hold a foam as well - there's still a few bubbles nearly 24 hours after putting it through the Buchner.

I shall pay close attention to the viscosity and any apparent surfactant activity during the rest of the harmala recovery process.