Harmine, harmaline and THH from Syrian Rue. Verification and finetuning of the VDS-protocols

[downwardsfromzero]

[...]
The hydrogen ions in (acidic) water are positively charged. They would need to be balanced out with the addition of a corresponding number of electrons. Where would these electrons come from? Reducing agents provide electrons...

Well, my tap water, even filtered tap water I believe would have both positive and negative ions in it, various minerals, etc., right? Who's to say that using distilled water plus zinc or other minerals of your choosing is more effective or efficient than using tap or well water, which likely has various elements from both the ground and the pipes?

I mean heck, a quick google search shows that common "galvanized" pipes are coated with zinc. Not to mention all the trace minerals in ground water.

Yes, the water has to have a balanced amount of positively charged (cations) and negatively charged particles. This does not mean that any of them are able to give up electrons to a protonated harmaline molecule.

It's important to make the distinction between dissolved metallic ions and metals in isolated elemental form. Once the (e.g.) zinc metal has dissolved it no longer has any spare electrons to give up, likewise the other trace dissolved metallic minerals - "trace" also being significant here because the electrons have to be supplied in a corresponding ('stoichiometric') quantity to that of the amount of harmaline being reduced.

Before this gets too complicated, I'll emphasize again that the electrons necessary for the reduction to take place have to come from somewhere sensible. There are plenty of substances in plant material which could fulfil this role, especially if metallic ions of variable valency are present.

Some metals are capable of giving up variable numbers of electrons, like iron, which can typically have a charge of +2 (bivalent) or +3 (tervalent). Bivalent iron in solution is fairly rapidly oxidised to tervalent iron on contact with oxygen from the air, making it an unlikely candidate for THH reduction unless it were acting in a catalytic way by transferring electrons from another reducing substance ('antioxidant') such as ascorbate, flavonoids or tannins.

It's quite conceivable that even in an earthenware vessel, if it were made of a sufficiently iron-rich clay there might be enough iron present to catalyse the THH reduction by transferring electrons from the aforementioned 'antioxidant' substances. Antioxidant = reductant. Prolonged boiling of plant material is, therefore, very different from boiling pure harmaline in pure water.

As for galvanised pipes - the zinc is on the outside, far away from your reaction vessel!

 

[Jagube]

Is my understanding correct?:

Even when basing with ammonia, some of the metal salts (dissolved or not) may get trapped in the precipitating freebase THH crystals. Adding ammonia slowly and under continuous stirring minimizes the extent of it, but doesn't eliminate it completely.
If one wants extra peace of mind, a simple solution to that is to redissolve the freebase in vinegar and re-base it with ammonia.

 

[Jagube]

Is my understanding correct?:

Even when basing with ammonia, some of the metal salts (dissolved or not) may get trapped in the precipitating freebase THH crystals. Adding ammonia slowly and under continuous stirring minimizes the extent of it, but doesn't eliminate it completely.
If one wants extra peace of mind, a simple remedy is to redissolve the freebase in vinegar and re-base it with ammonia. That way, if let's say 0.1% metals get trapped per precipitation, the re-precipitation will reduce the contamination to (0.1%)^2 = 0.0001%.

And a couple of questions:

1. What should the pH be after complete reduction? Last time I didn't use enough vinegar and the pH was about neutral, which resulted in an incomplete reduction due to insufficient acid. This time I added more vinegar and after 10 hours it was 5.5. Presumably that's ok, as it indicates there is still unreacted acetic acid in solution, so the initial amount of acetic acid was sufficient? In GordoTEK's video he ends up with 3.8, but it doesn't have to be that low?

2. Post-reduction, can I neutralize the unreacted acetic acid with NaOH, or at least up to a certain level (e.g. pH 6), before the harmine starts precipitating, and then switch to ammonia? The motivation is to save ammonia. Or would that affect the metal salts' solubility?

 

[blue.magic]

Even when basing with ammonia, some of the metal salts (dissolved or not) may get trapped in the precipitating freebase THH crystals. Adding ammonia slowly and under continuous stirring minimizes the extent of it, but doesn't eliminate it completely.

I can confirm the zinc contamination is almost completely removed by filtering the freebase THH and washing it with dilute ammonia.

Here the heavy metal test strip shows high concentration of Zn(+2) ions in the reaction mixture (the numbers are ppb concentration):

However, after washing THH and redissolving in HCl, the liquid is no longer contaminated with zinc. It's hard to compare the colours but the strip colour remained the same so I assume the zinc contamination is below 10 ppb:

1. What should the pH be after complete reduction? Last time I didn't use enough vinegar and the pH was about neutral, which resulted in an incomplete reduction due to insufficient acid. This time I added more vinegar and after 10 hours it was 5.5. Presumably that's ok, as it indicates there is still unreacted acetic acid in solution, so the initial amount of acetic acid was sufficient? In GordoTEK's video he ends up with 3.8, but it doesn't have to be that low?

I the VDS paper, they used 2 g harmaline base (9.3 mmol) and 100 mL of 7% acetic acid (approx. 116 mmol) so it is about 12 times molar excess of acetic acid.

I think the pH is not as important, only that excess acid is present so that the reaction will proceed reasonably fast without heating.

2. Post-reduction, can I neutralize the unreacted acetic acid with NaOH, or at least up to a certain level (e.g. pH 6), before the harmine starts precipitating, and then switch to ammonia? The motivation is to save ammonia. Or would that affect the metal salts' solubility?

I would not do it as sodium ions could skew the pH readings (sodium acetate is basic at higher concentration) - although this might not be a problem at small concentrations.

To save ammonia, I guess you can precipitate everything, filter, redissolve in just enough acid. The problem is that the precipitate is quite hard to filter because of all the zinc co-precipitating. It's also harder to redissolve the zinc and it takes excess acid. So maybe this is not a good idea...

It would be interesting to optimize the amount of acid used in the reduction. Maybe even something like 4 times molar excess will suffice. Separating unreacted harmaline and THH will tell how the reduction goes after X hours of reacting.

 

[Jagube]

I can confirm the zinc contamination is almost completely removed by filtering the freebase THH and washing it with dilute ammonia.

Thanks for carrying out the tests and posting the results.

However, you redissolved the washed THH in HCl. Dissolving THH in HCl can result in the THH precipitating as THH HCl (like in Manske precipitation) when the HCl concentration is high enough. A freebase THH crystal with metal compounds trapped inside it, when placed in a concentrated HCl solution, could theoretically turn into *solid* THH HCl on the crystal's outer surface and prevent the HCl from penetrating deeper into the freebase and dissolving it, which would keep the metal contamination trapped inside.

I the VDS paper, they used 2 g harmaline base (9.3 mmol) and 100 mL of 7% acetic acid (approx. 116 mmol) so it is about 12 times molar excess of acetic acid.

I've used less vinegar and ended up with incomplete reductions. That sounds like a lot of vinegar!
I like to work with smaller volumes and what seems to have worked for me in the past is adding a small amount of HCl to bring the pH down without increasing the volume too much.

I think the pH does matter, because this reduction reaction is about donating protons to the harmaline molecule to form THH, so the higher the concentration of H+ ions in solution, the better. Therefore strong acids (like HCl) should be better than weak ones (like acetic).
The reaction in this case is 2HCl + Zn -> H2 + ZnCl2, and the H2 binds with the harmaline.

We don't want the HCl concentration to be too high though, as we don't want the harmaline HCl to precipitate, and both harmaline and THH may be destroyed by HCl.

 

[blue.magic]

However, you redissolved the washed THH in HCl. Dissolving THH in HCl can result in the THH precipitating as THH HCl (like in Manske precipitation) when the HCl concentration is high enough. A freebase THH crystal with metal compounds trapped inside it, when placed in a concentrated HCl solution, could theoretically turn into *solid* THH HCl on the crystal's outer surface and prevent the HCl from penetrating deeper into the freebase and dissolving it, which would keep the metal contamination trapped inside.

Yes this is what happened as I stupidly dissolved THH in a tiny volume of HCl (about 50 mL) - it started precipitating quickly so I increated the volume and heated the solution to keep everything dissolved.

I measured the heavy metals after everything was dissolved. There is a chance of trapping some metal in the precipitate, so redissolving THH and reprecipitating is probably a good practice. I found this also removed some discoloration in my case, so it definitely helps with removing trapped stuff.

I probably should have used acetic acid to redissolve THH, because of the solublity issues you mentioned.

Yes I agree that acetic acid concentration is important. That's one thing that can be optimized - using just enough to convert sth. like 98% harmaline in say 5 hours

There are so many tiny little secrets that are all essential for success...

 

[tregar]

The thing to do would be to probably study the method used by the Santo Daime to brew Ayahuasca, as Dr. Callaway mentioned that all of their brews contained near zero mg of harmaline (see pic attached), whatever they are doing, they are managing to reduce any and all remaining harmaline in the brew to thh.

Professor8 managed to reduce using vitamin C (see his threads here from years ago), and Dr. Shulgin mentioned in the "entheogen review" many years ago that vitamin C is a hydrogen donor.

2005: Many brews below are 100ml or 3.3oz, simply multiply each figure x 100 to get the mg amount for those brews, examples:

entry #1 from UDV is 183mg tetrahydroharmine, 9mg harmaline, 172mg harmine.
entry #22 from Santo Daime is 168mg tetrahydroharmine, 0mg harmaline, 198mg harmine.
entry #29 from Shuar Indian is 163mg tetrahydroharmine, 0mg harmaline, 180mg harmine.

 

[Jees]

...Professor8 managed to reduce using vitamin C (see his threads here from years ago),

Did he? I only found this post were nothing was checked at all. Maybe you have another link were some proof was offered? Thank you.

and Dr. Shulgin mentioned in the "entheogen review" many years ago that vitamin C is a hydrogen donor.

I'm no chemist or maybe I have something skewed but are not all acids hydrogen donors by definition? We cook rue often in acidy pots with a ton of harmaline surviving. So I suspect "being a hydrogen donor" is not enough to sustain a reduction to THH.

Respectfully,
Jees

Edit: or has it to do with the difference between a hydrogen atom donor (vit C) or hydrogen-ion donor (acid)?

 

[tregar]

Yes, that's the post Jees. I tried it once boiling with pure citric acid in high amounts for 3 hours, and still no color change from green to blue. I don't think it works, perhaps with 12 hours boiling, but who want's to boil harmaline for 12 hours?

Checking color change is easy, simply put an ear cleaner tip in the solution, then swab the back of a paper plate with the swab, then put the plate under blacklight, if it glows blue then you have THH. Other substances that are said to glow blue: LSD, psilocybin, etc.

Substances like Ayahuasca, LSD, mescaline, mushrooms, etc. have been show in studies to activate the right side of the brain--the part involved with creativity, euphoria, visualization, empathy. They seriously light up that side of the brain like a neon light.

The world is moving in the direction of the Left Brain: technology and science. What the world needs is to move in the direction of Right Brain development: empathy, spirituality, connectedness. Compounds like Caapi could be said to improve emotional intelligence. Is this caapi a smart-nutrient for the right side of the brain? you be the judge. I certainly think it is.

 

[Jagube]

I'm no chemist or maybe I have something skewed but are not all acids hydrogen donors by definition? We cook rue often in acidy pots with a ton of harmaline surviving. So I suspect "being a hydrogen donor" is not enough to sustain a reduction to THH.
[..]

Edit: or has it to do with the difference between a hydrogen atom donor (vit C) or hydrogen-ion donor (acid)?

See the chemistry of harmaline reduction:

When acetic acid and zinc react (one molecule of zinc and two molecules of acetic acid) to form zinc acetate, H2 gas is released and the harmaline accepts it.

The reduction requires a reducing agent and an acid. The acid is a proton donor, the reducing agent is something that oxidizes readily. The reduction of one thing always needs to be balanced by the oxidation of something else.

My understanding (and please correct me if I'm wrong) is that even though ascorbic acid is both an acid and a reducing agent, it can't act in both roles in one reaction, because it won't react with itself.

 

[tregar]

Yes! that makes sense, thanks for the explanation Jagube.

 

[Jees]

Thx Jagube for that link, Gordotek said many things spot on and I learned some.
His precipitation of harmine at pH 8, while that happens in VDS around 7,2 illustrates nicely what we have been telling a lot, namely you cannot pin point a pH level because it is dependent on factors. The shape of the curve (the pH depression) on the other hand does not lie and is reliable.
The curve shift lower when there is low salt 'contam' e.g. separating harmine from DHH by using min acid to dissolve. Adding base there will be less salt. Doing that I could drop out harmine even lower than 7.
The curve shift higher with more salt contam e.g. reducing DHH to THH, then there is more acid needed. One needs more base during precipitation thus more salt forming. Gordotek saw his harmine falling out at 8.
We suppose all probes used were calibrated nicely to start with of course.

It is the first time I saw someone mentioning pH depressions outside the VDS papers, and he also confirms that the second pH depression (the one for THH in this case) is more a plateau than a real pH dip. He called it pH "stuck".

Also this illustrates that if pH depressions occur (it happens when the conditions are met) then the good ole pKa point of 8,75 as a separation point is not valid.

 

[blue.magic]

See the chemistry of harmaline reduction:

The GordoTek's THH is tan in colour. Mine is bright white (the HCl salt is slightly off white or yellowish). Which color is the correct one?

I thought the tan color is due to lighting but in the end the filter paper containing THH is white and the content is really tan colored.

Could it be due to some harmine/harmaline still in the mix? He started with mixed harmalas so maybe the product is contaminated (we know some harmaline stays in solution even at high pH, not much but possibly enough to discolor THH).

 

[Jagube]

The GordoTek's THH is tan in colour. Mine is bright white (the HCl salt is slightly off white or yellowish). Which color is the correct one?

Maybe it's less pure. I've never obtained white THH, but I've had anything from golden to off-white, depending on purity.

 

[downwardsfromzero]

Backtracking to vit C again:

I'm no chemist or maybe I have something skewed but are not all acids hydrogen donors by definition? We cook rue often in acidy pots with a ton of harmaline surviving. So I suspect "being a hydrogen donor" is not enough to sustain a reduction to THH.
[..]

Edit: or has it to do with the difference between a hydrogen atom donor (vit C) or hydrogen-ion donor (acid)?

See the chemistry of harmaline reduction:

When acetic acid and zinc react (one molecule of zinc and two molecules of acetic acid) to form zinc acetate, H2 gas is released and the harmaline accepts it.

The reduction requires a reducing agent and an acid. The acid is a proton donor, the reducing agent is something that oxidizes readily. The reduction of one thing always needs to be balanced by the oxidation of something else.

My understanding (and please correct me if I'm wrong) is that even though ascorbic acid is both an acid and a reducing agent, it can't act in both roles in one reaction, because it won't react with itself.

Ascorbic acid is a bit different from the typical organic carboxylic acid. It has what is known as an 'endiol' structure that is responsible for both its acidity and its electron-donating properties. So, loosely speaking, it can donate both protons and electrons 'at the same time'. Attached is a somewhat simplified illustration of how that works (minus the substrate that accepts the protons and electrons).
[N.B. 'electron donor' is another way of saying 'reducing agent'.]

This in no way is intended to demonstrate proof that ascorbic acid will reduce harmaline to THH, nor that dehydroascorbic acid (on the right) will oxidise harmaline to harmine (especially not in the absence of other biomolecules and/or transition metal ions) but I think it helps piece together the outlines of a possibility.

 

[tregar]

Very interesting, thanks downwardsfromzero.

 

[GordoTEK]

The GordoTek's THH is tan in colour. Mine is bright white (the HCl salt is slightly off white or yellowish). Which color is the correct one?

In the video (I did mention this by the way) I do not separate the harmine from the THH, so yes, you are looking at a mixed product there (I did this partly out of laziness, because the result is about a 50/50 mix of harmine and THH which is how I like to use them anyway, so no need to separate if you are just going to combine them again later in the same ratio). Also my starting mix of harmine/harmaline was a little on the darker side as I did not do the excessive manske's that some people do to get it to the highest possible levels of purity (I don't think this is necessary, and it results in loss of the desired compounds because of the inefficiencies involved in each iteration). You can see exactly what the mix looked like before doing the reduction here (the pre-reduction video that shows the harmine/harmaline extraction).

To answer the earlier question about how much vinegar to use - yes, you can get away with using much less than I used in the video. I also included a calculation in the video (sorry I know its hard to read, I did a lousy job on the text color there):

You might be able to get away with using a 2x factor for excess, haven't tried, but I used considerably more than I needed to.

Likewise, the calculation for zinc is:

 

[Jagube]

Is harmine really darker than harmaline and THH, or is it just that at the pH at which harmine precipitates most impurities co-precipitate with it, so by the time the harmaline starts precipitating, the supernatant is already relatively clean?

I've had harmine anywhere from yellowish to dark brown, depending on the degree of purification, and when I have separated it from harmaline, the harmaline has always been a lighter color, and post-reduction, the resulting THH even lighter. This leads me to believe that pure harmine, pure harmaline and pure THH are all white (or maybe just off-white), but products coming from P. harmala extractions are usually far from pure, hence the color gradient from harmine -> harmaline -> THH.

 

[GordoTEK]

Yes, I do believe for all 3, the lab grade versions would all be pure white. In this thread Endlessness says:

White. So is pure harmaline. I think the browner color is neither of those, it's some other impurity. If you do several cycles of precipitation you should be able to get it white, but its not necessary. Three rounds of precipitations should already be enough to have a pretty pure product that you can accurately dose.

I agree with his comments. Syrian Rue is a powerful dye, kind of like food coloring- trace amounts of the dye compounds will give you those tans and browns but it is not detrimental to the targeted alkaloids (indeed many use a full spectrum extract and claim benefits over purists but I don’t agree with that, I want purity but there is no need to take it to extremes, you will just lower your yields substantially with no added benefits).

 

[blue.magic]

Thanks, Gordo, for the explanation. Now it makes sense.

I found a prolonged vigorous stirring (over 10-12 hours) inctroduces a reddish/brown substance and the loss of yield. Some people said this could be "harmala red" or some oxidation or polymerisation product.

Because of that, I do the reductions in a closed container with a bubbler (a simple airlock for alcohol ferments will do). Inert atmosphere is not needed as the container fills with hydrogen anyway.

So far I did eight or so separations with varying conditions and harmine is always brown, harmaline light brown or tan and tetrahydroharmine yellowish, off-white or white.