Hello! Long time reader, first time poster.

I have been making extracts of peganum harmala for about 7-8 years now, and in the last couple years I have gone the extra step of separating and crystallizing the individual alkaloids, and I have some test results I could use some extra input on.

My extraction procedure: Make P harmala tea (usually 3kg of seeds at a time in a giant stainless steel cooking pot (from academy, normally used for boiling the hell out of dead animals). Precipitate with lab grade NaOH, redissolve with distilled vinegar, filter through lab paper with vacuum, NaOH precip, repeat 3-4 times. Redissolve in vinegar, filter, get it up to boiling temperature, add salt (Manske), let xtals crash out over a day, filter and dry, redissolve in vinegar, heat&salt&crash again, filter, add base until the brownish alkaloids fall out, filter those out, add more base to get the orangish portion (a large portion of the alkaloids, believed to be harmine), filter those out add more base till the precipitate begins to turn white, filter that portion out, then crank the pH up to 13 and white (with maybe a hint of pink) precipitate falls out (the 2nd largest portion, believed to be harmaline). I then take the individual harmine and harmaline portions and run through a couple more Manske cycles, save some as HCl, and base precip the rest to get the freebases.

I have been working with HPTLC plates (looking for fluorescent compound in wild mushrooms) for the last few months, and decided to run my harmala extracts through a plate to see how well the separation works. Here are the results.

1. The un-separated alkaloids (full spectrum after Manske). Surprisingly, this only had 2 spots on it.
2 & 3 harmaline reduced with zinc to produce THH.
4. Orange harmine fraction.
5. White harmaline fraction.

2& 3 have a spot with a lower rf than any of the others, but I don't know the rf of THH so I can't say for sure that it worked.

I think that the purple spots are harmine (based on the fact that the harmaline sample has a very light purple spot, and the harmine sample has a bigger/brighter spot).

Questions: Has anyone done this and identified the individual spots? Does anyone have the relative rf values for the different harmalas? Why do the separated alkaloids have an additional spot in the middle, while the mixed alkaloids don't? The separated alkaloids were made from the mixed alkaloids, so perhaps something that was created during the separation process? Whatever it is, the zinc reaction made it disappear because lines 2 & 3 were made by mixing #5 with zinc.

Any input would be much appreciated. I am trying to improve my separation technique but don't have access to HPLC or MS. I will hopefully be running GC (no MS) next week and will post the results. Thanks!

By the way, I don't know how to get the images to show in the post so I just attached them as files. Sorry for the inconvenience

Oh and why wont it let me download my own image to see if it uploaded properly? What do I have to do to get sufficient privileges to view my own images?

Hey!

Yeah the top blue/purple spot is harmine, the other bottom spot is harmaline, and the spots that appeared below harmaline are THH. Here`s a relevant thread:

https://www.dmt-nexus.me...aspx?g=posts&t=35984

If you want a better separation of harmine vs harmaline, you need to control the pH more (or maybe you did but didnt describe it). The best is to precipitate at pH 8.75 to get most your harmine with very little harmaline. Then go up to 12 to get all your harmaline with a significant amount of harmine. Redissolve your harmine/harmaline and repeat process, getting most of the leftover harmine precipitating it with 8.75 and getting your purer harmaline raising the pH after that.

As for your zinc reduction, how exactly did you do it? Maybe you could use more zinc and acid, maybe leave it reacting for longer, or mixing more.. ?

Regarding the middle spot on your lanes 4,5, thats the biggest mistery for me. If it wasnt there in the crude manske extract, why would it be there later? Must be some artifact, but what? I`d try redissovling and reprecipitating them both and testing again, see what you come out with. I`d also try repeating the whole experiment but separating harmine and harmaline as mentioned above. Not sure what else you can test but its late and my mind isnt working so well now.

Thanks for sharing this btw, feel free to share more TLC tests you do in the future, harmalas or otherwise, its always interesting to see

Hi TreehouseChemist!

Thanks for posting your interesting research. Did you run the plates with freebase or with HCl (or other) salt? What solvent did you use?

Awesome! Thanks for the info, I have another 8 kilos of seeds on the way, I will be doing more tests and posting the results. I am not sure what the middle alkaloid in the last two are either, I will see if I can scrape it off and get an IR reading next week.

The THH was made by dissolving about a gram and a half of harmaline in vinegar, filtering in to a small erlenmeyer, and then adding about a tsp or two of zinc powder, swirling by hand and letting react until no more gas bubbles formed on top of the zinc. I will try the THH method again next week too (going out of town for the weekend).

The plates were run by dissolving 10mg of each sample (as freebase) in 10ml of 50:50 acetone:methanol, spotting 5μl on the left plate, and 1-2μl on the right plate, and running with ethyl acetate:methanol:ammonia 70:10:3 as recommended by this article http://www.ncbi.nlm.nih.gov/pubmed/18980138