I see that in these normal test tubes the precipitate forms a somewhat globular clump with liquid around the sides. You would find improved results using centrifuge tubes, with the tapered/conical base. As you probably already know!

Yes, thanks for the tip. I have 15ml test tubes with the tapered bottom. Have not tested them out yet though, they don't fit as snug as the ones pictured which that came with the cheapo $50 fuge.

Yeah, best to have a snug fit rather than a broken tube and potentially exploded centrifuge!

I always had loads of the tapered tubes when I was a kid, but no centrifuge. My dad brought them home from work for me. Shame he couldn't get away with bringing me a centrifuge, but really it would have been too heavy to fit on the back of his bike

i meant the tumeric yellow you mentioned, i could only find this which says the harmalol base is red but i couldn't find the reference to this so... maybe the tumeric yellow is in solution or as salt?
i also over basified the manske solution and still got the slime although it looks better now after 3 water washes.. haven't got the time and space to dry it yet
i'm not convinced with the new results i need to read them later when i am more focused.. anyway good work and thank you

Thanks for doing the extra basing test.

I did manske on the test-tube family from post #39 and cloudiness appeared in most test tubes, but the NaOH test tube did not show cloudiness. Also, when I did get cloudiness it was reddish, not yellow. Since this result is different than your's and Jees' (yellow haze even at very high pH), I have to say I don't know what the yellow slime is and that it is not simply the red/brown harmalol candidate we separated under high basic conditions in post #27 (where it becomes more soluble as shown in post #39).

The tumeric yellow comment is from here:

https://www.indofinechem...t-details.aspx?Sku=H-003

I see in your link that harmalol can also be red in certain conditions. There are lots of colors out there, my philosophy now is that anything red/yellow is interesting enough to work on.

I tried the NaOH extraction on rue seeds (because our harmalol candidate dissolved best in a strong base and FISHER had used KOH to isolate harmalol from rue seeds back in 1901). There was a strikingly fast reddening going into solution after dipping 50g of seeds in a hop nylon bag into 4% NaOH (picture below). Something is being pulled out of the seeds very aggressively (and I think harmine/harmaline should mostly stay on the seeds as they are not soluble in a strong base). Large color change within seconds/minutes and at room temperature. Next, I'm going to carefully pour off and acidify the dark red base extract with HCl (muriatic acid from the hardware store) to see if something interesting happens. Or does anyone already know what is causing this deep dark color when rue seeds go into a strong base? The standard harmine/harmaline extraction on these pre-based seeds will also be interesting (maybe gives normal yields and is cleaner with less redness during the extraction steps?).

so yeah the tumeric yellow is not necessarily in freebase form.. the site you linked doesn't specify but since they mention crystals then i suspect it's in salt form... i think the salt is tumeric yellow and the freebase is maybe red?
very interesting i never seen that dark color when basing rue tea, whatever is causing this it must be insoluble in acidic solution

I guess that the precipitae in the vinegar tube does not consist of harmala alkaloids of any kind. Their acetates are very soluble in dilute acetic acid. Any harmalol would therefore have to be in the supernatant liquid.

I agree it is not an acetate, but I think there is a chance the red centrifugate can be harmalol which never formed an acetate despite being in vinegar.

The way I rationalize it (aka could be lying to myself in my hopes and dreams that we are chasing harmalol despite trying to be objective), is that to form the acetate, harmalol needs to be pronated at the amine group, but for that the electropositive carboxylic O-H group needs to be overcome.

In short, I think harmalol is an inner salt and very high/low pH values are needed to bring it into solution (e.g. FISHER used KOH to isolate harmalol from rue seeds). I did test phosphoric acid after reading that Jees uses it and more of it seemed to stay in solution compared to vinegar (going by color), but I did not take a picture.

Muriatic acid test is tonight (outdoors)...

Public Note: Although muriatic acid can be bought at the hardware store it can be dangerous (produce corrosive vapor, react with metal, etc). No one should use it just because it is mentioned in this thread. Please stay informed and safe and make informed decisions after careful review of the official hazard and safety information.

Muriatic acid (HCl) test was interesting

1) Room temp extract of 50g of rue seeds using 4% NaOH was done. Pulls where done with ~150ml of base each time. Total of 9 pulls. Dramatic dark red color was observed in the first two to three base pulls. 9 total pulls where combined into 3 jars:



2) First jar (pulls 1-3) was pressure filtered with 4 stacked coffee filters then acidified with 32% HCl from the hardware store. Using 32% HCl is not trivial from a safety stand point. It is each person's responsibility to research this or any new chemichal and make their own decision on using it or not. Mods, please help if I'm stating anything improperly. I ended up acidifying outside with chem gloves (I bought the blue ones in honor of Jees), safety googles, close toe shoes, and a big fan blowing behind me (this is not a how-to guide for this chemichal and will not guarantee safety, I'm just documenting this experiment).

Here is a UV picture before the acidification started (pH 12.5)



After adding some HCl UV clouds of green (compatible with 534nm) start to appear.



Eventually what lool like crystals started to appear! pH was measure at 1 at this time, but the crystal candidates started appearing a bit before that.



3) Jar with Crystal candidate precipitates was put in the fridge to settle. Everything else was cleaned and washed carefully. HCl was stored outside in a plastic weather bin.

So what is a possible interpretation?

-The NaOH pulled the harmalol candidate by "grabbing" on to the -OH group. A strong base is needed because of the electronegative amine group.
-It does not seem like much harmine or harmaline where pulled by the base (expected) since there is no major precipitate in the strong base and yellow fluorescence was not observed in any of the pulls or during acidification. Also, while at pH 12.5, the NaOH pull was filtered with 4 stacked coffee filters to remove harmine/harmalol that still made it out of the rue seeds.
-As the pH was lowered by HCl the green harmalol fluorescence at (apparently) 534nm starts to appear (expected from the paper previously uploaded). Once the pH is very low, the harmalol can be pronated at the amine group, overcoming the electropositive -OH group. Once pronated it can combine with Cl (harmalol HCl candidate). This may work also with 10% phosphoric (which is a relatively strong acid concentration not tested yet, but 3% did not seem to work).

Since 150ml of 32% HCl where added to a 4% NaOH 450ml solution, it is the equivalent of 73g of salt for Cl- concentration and 26g for Na+ concentration. Usually for this solution (ended at 600ml), 120g of salt will be added for manske. Plan is to let the crystal candidates settle, filter, and add salt to see if more precipitates out. Also, need to acidify the other two jars with base pulls.

These are all just pie in the sky ideas right now. We need external analysis of the harmanol candidate. Until then this could be all wrong. However, laying out the experiments and my understanding so far for review/feedback/corrections. If anyone can repeat this even better.

It should be very easy to obtain a free base harmalol candidate. Simply do base pulls at pH of 12.5, filter, then reduce to pH of 10 to 5. Harmalol candidate will slowly precipitate out at the lower pH. Clean with repeated water rinses allowing for water precipitation. Any takers to try this?

Update: with a lot of difficulty some of the precipitate was recovered from the HCl solution in post (#49). I don't recommend anyone doing this - it was only for experimental reasons. Yield is non optimal (a lot was lost), so not reporting it.

Yellow crystals where isolated. These are harmalol HCl candidates (based on the color info that Sakkadelic found):



Standard manske did not yield any crystals (makes sense since we extracted with NaOH room temp and filtered the extract, which means harmaline and harmine should have been left behind in the seeds and/or filters).

More experiments and reports to follow. We will not know for sure what this is until we send it out.

Finally the slime dried! Leaving a nice pattern
2.4g after putting it in the oven on low for 15 mins, 2.4% that's a lot!

pitubo you say it might be protein, but according to this shouldn't protiens salt out?

Since we drink rue tea and this stuff must come from rue do you think it's safe to try some? Like a 100 mg, i've been taking frequently similar doses of the freebase i made so i would be able to tell if it's active. Is there anything to be concerned about that i am not thinking of?

I would advise starting with way less than that. Less than 1mg. Or, preferably, send a bit to Energy Control first so you have some idea of what it is.

That said, if this has been extracted from 100g of SR, 100mg would correspond to just over 4g of whole seeds.

Either way, I love the picture!

Cool!
What did you do before the oven dry? Is it settled decanted basified manske slime that you put in the oven? How do you deal with the base and salt, did some dry with it?

downwardsfromzero, yeah i'd send a sample if i could and the mystery would be resolved! and okay i'll start lower if i decide to

Loveall, when i did the last base i tried not to add too much NaOH, then i did 3 water washes each of them is like a 5x dilution, with the last one i let it sit after the decant for 2 more days and removing any water that shows, i guess the salt content would be minimal but the base might not be, i don't have PH paper or anything to measure PH, what i do is rub a drop of the water between my fingers if it feels slimy then i need more washes and when it feels like normal water i taste it and if it's sour then it still needs more washes i need to upgrade!

for drying i let it dry by itself in the dish and it took around 3 days but it wasn't fully dry so i put it in oven to finish the drying.. so yeah some base and salt will dry with it but i think it's just a little

i want to work more on it, get things and test it but i'm just not as enthusiastic about extractions or anything these days...

Hey Sakkadelic. Everything ok? You mention you are not enthusiastic, sending you some love.

Yeah, a pH meter would be helpful. It is even more mazing that you are such a great rue extractor and don't even own one!

Like you with the manske slime, I've been thinking of sampling the red harmalol candidate from NaOH extraction. Right now I'm trying to make it repeatably and cleanly and then send it out to endlessness if he will take it. Attached is what it looked like at the begining (after 20minites) during the last oven dry at low temp, but I think it was damaged somehow because it has lost it's pH 9 greenish/yellow fluorescence. I left it in the oven for many hours, maybe the harmalol candidate degrades in hot air over time? I've read before that harmalol degrades in air but had not had any issues with the candidate until this long oven dry. If this is an issue, then isolating it and keeping it active is going to be even more difficult (really need to dry it in HCl form).

By the way, whatever it is, about 2% of the rue mass turns into this.

Endlessness will run your samples for FREE.

https://www.dmt-nexus.me...aspx?g=posts&t=74123

edit:

ah, I saw your post in another thread so you know this already.

I think you may be onto something here, but really impossible to be definite without mass spec. Really looking forward to hearing about those results



aqueous NaOH will dissolve the harmalol by deprotonating the -OH group to give -O(-) ion, which is soluble in water. A strong base is needed because the pKa of a phenol is around 10, in the case of harmalol which is very electron rich aromatic, it may be upwards of around 11. The actual value may be documented somewhere.



the OH group is more electronegative than the amine. Part of why it holds onto its proton tighter, giving it that high pKa value. In this way it is zwitterionic, so its not easy to extract as a neutral, freebase molecule. At low pH you get exclusively the amine protonated salt, the hydrochloride salt precipitation arises out of a equlibrium effect, large excess of chloride ions, the harmala salt precipitates to lower the concentration of chloride in solution. This isn't a universal effect in alkaloid extraction, it works well in the case of harmaline and harmine, but not necessarily for other molecules. Probably has to do with the structure, fully aromatic, schiff base. So a good chance it will still work with harmalol.

Thank you Mindlusion! It is great to have a chemist with us in this thread! Definitely need the help.

Basic and Acid pKa are published here, based on ChemAxon calculations (which I just figured out anyone can do on the free demos!). Also, this claims that at the Acid and Basic pKa average (pH=7.96) the zwitterion will be mostly neutral. A free ChemAxon web simulation gives this for harmalol,



So minimun solubility is near pH = 8.8 which seems to match the centrifuge results from earlier and is a little higher than the simple pKa average. How accurate are these numerical demos? There is no way to input salt presence, so that seems like a limitation right there.

After an NaOH extract and filtering (5x stacked coffee filter), if the pH is brought down to 8.8, after a few days dark red/brown conglomerates that look just like dried blood crust start to form. They do not form if the solution is left at 12.5pH. They seem to be very similar to the dried crust from the oven dry centrifuged material in the earlier post.

Working on the oven dried platelets they seem pretty indestructible and I can't get them to fluores. They don't dissolve well in NaOH, IPA, or 32% HCl. I'm worried that the long oven dry did something bad to the harmalol candidate? I almost sent out the oven dried crust out for analysis but this concern stopped me. Could the crust be a stable matrix of OH and NH groups sharing a bond and forming a stable conglomerate? Here is a picture of the crust not dissolving in IPA. Is this a known behaviour?



Still need to test the precipitated platelets (not oven dried).

Also have tried to pull the candidate from the slightly basic solution (before the platelets form). FISHER used chloroform, which I don't have. I tried toluene, naphtha, and IPA. One solvent worked: IPA which was made immiscible by using salt. A beautiful fluorescence was picked up by the IPA. No platelets formed in this test tube over time (but that could also be due to precesnce of salt, need to test that separately). Below are pictures of the IPA picking up something.





I'm thinking about adding HCl to the IPA that picked up the fluorescence to see if harmalol HCl candidate would form, is this stupidly dangerous?

Anyway, I run experiments almost every day so guidance is welcome. For example, would dissolving the centrifugate harmalol candidate (does not come out as platelets) in HCl then allowing to dry outside be a good way to try to convert to stable harmalol HCl candidate?

PS: Harmalol code to input into the solubility demo below.

<cml><MDocument><MChemicalStruct><molecule molID="m1"><atomArray><atom y2="-0.8789191835816532" x2="-4.25436572983439" elementType="C" id="a1" /><atom y2="-0.5587351797223008" x2="-2.748018424704327" elementType="C" id="a2" /><atom y2="0.9058918553722368" x2="-2.2721322533669106" elementType="C" id="a3" /><atom y2="2.0503348866074224" x2="-3.302593387159554" elementType="C" id="a4" /><atom y2="1.7301508827480707" x2="-4.808940692289614" elementType="C" id="a5" /><atom y2="0.2655238476535341" x2="-5.284826863627032" elementType="C" id="a6" /><atom y2="-1.4639244682527068" x2="-1.5021322533669057" elementType="N" id="a7" lonePair="1" /><atom y2="-0.5587351797222957" x2="-0.25624608202948806" elementType="C" id="a8" /><atom y2="0.90589185537224" x2="-0.7321322533669099" elementType="C" id="a9" /><atom y2="-0.8789191835816432" x2="1.2501012231005748" elementType="C" id="a10" /><atom y2="0.2655238476535463" x2="2.2805623568932125" elementType="N" id="a11" lonePair="1" /><atom y2="1.7301508827480822" x2="1.8046761855557918" elementType="C" id="a12" /><atom y2="2.0503348866074287" x2="0.29832888042573024" elementType="C" id="a13" /><atom y2="-0.05466015620581555" x2="-6.791174168757093" elementType="O" id="a14" lonePair="2" /><atom y2="-2.3435462186761784" x2="1.725987394437996" elementType="C" id="a15" /></atomArray><bondArray><bond order="1" atomRefs2="a1 a2" /><bond order="2" atomRefs2="a2 a3" /><bond order="1" atomRefs2="a3 a4" /><bond order="2" atomRefs2="a4 a5" /><bond order="2" atomRefs2="a8 a9" /><bond order="1" atomRefs2="a3 a9" /><bond order="1" atomRefs2="a9 a13" /><bond order="1" atomRefs2="a8 a10" /><bond order="1" atomRefs2="a5 a6" /><bond order="2" atomRefs2="a1 a6" /><bond order="1" atomRefs2="a6 a14" /><bond order="1" atomRefs2="a7 a8" /><bond order="1" atomRefs2="a2 a7" /><bond order="1" atomRefs2="a10 a15" /><bond order="2" atomRefs2="a10 a11" /><bond order="1" atomRefs2="a11 a12" /><bond order="1" atomRefs2="a13 a12" /></bondArray></molecule></MChemicalStruct><MElectronContainer id="o1" occupation="0 0" radical="0"><MElectron atomRefs="m1.a7" difLoc="0.0 0.0 0.0" /><MElectron atomRefs="m1.a7" difLoc="0.0 0.0 0.0" /></MElectronContainer><MElectronContainer id="o2" occupation="0 0" radical="0"><MElectron atomRefs="m1.a11" difLoc="0.0 0.0 0.0" /><MElectron atomRefs="m1.a11" difLoc="0.0 0.0 0.0" /></MElectronContainer><MElectronContainer id="o3" occupation="0 0" radical="0"><MElectron atomRefs="m1.a14" difLoc="0.0 0.0 0.0" /><MElectron atomRefs="m1.a14" difLoc="0.0 0.0 0.0" /></MElectronContainer><MElectronContainer id="o4" occupation="0 0" radical="0"><MElectron atomRefs="m1.a14" difLoc="0.0 0.0 0.0" /><MElectron atomRefs="m1.a14" difLoc="0.0 0.0 0.0" /></MElectronContainer></MDocument></cml>

Still working on the dark red NaOH rue extract. According to idealized predictions (no salt or other alkaloid interactions) from here,



LogS definition here

So the simple strategy is to extract at pH>12 (using 4% NaOH) and filter. At pH 8.8 and after a wait there is some clumpy precipitation, not pursuing that at the moment (could be a neutral harmalol candidate). When acidified, solution turns orange/brown. Trying to salt out with NaCl (manske) yields a precipitate but it is not crystals, more like a deep orange mud. I collected this post manske mud and based/filtered again. Here is the resulting base,



This was acidified with phosphoric acid until after it changed color and then the pH measured (pH~2.8 ),



At this point there was about 100ml of solution. I decided to now salt with ammonium sulfate (instead of NaCl) to try to get something other than mud. Here is the solution right after adding 50g of ammonium sulfate (which sank to the bottom):



This was shaken. After the ammonium sulfate dissolved, pH went up to about 3.5. After a few minutes there was a surprise:



Something is floating at the top. Looks similar to the mud I got during NaCl salting but browner. I'll collect it later. Anyone know what it could be or why it would float this time? Im having difficulties getting salted crystals, any ideas welcome.

Update: Moved some of the separating liquid from the last jar in the last post to a test tube. Added equal volume of water. Mud went from floating to sinking (see attachment). Whatever is conglomerating has a density inside the range of the ammonium sulfate solution density window I guess.

The brown precipitate from the previous post was collected and dried. It resulted in a loose fine particle powder that was easily scraped up (harmalol sulfate candidate):



A small amount of this was redisolved in vinegar and it fluoresced green/yellow. It also and lost the green fluoresence when the pH was brought up with NaOH and turned a deeper red.

So to recap, we have found 3 candidates. From left to right (1) harmalol HCl candidates (post #50), (2) Harmalol sulfate candidate (this post), and (3) Free Harmalol candidate (post #55)



Previously (post #57) it was reported that the free harmalol candidate was "indestructible" in several solvents. However, under closer UV inspection the flakes gave off faint green luminescence indication very slow (but existing) dissolution:



After spending the night being stirred in a magnetic stir plate, almost all the flakes dissolved. Still some very small ones remain. Will keep on stirring and once/if they dissolve completely will add Zn and check for luminescent change (which would be a tetrahydroharmol candidate I believe). They flakes sure are resilient.

Finally, I started a new harmalol candidate extraction on 100g of rue in a nylon filter bag. Using 0.5% NaOH this time. Interestingly under UV one can see green harmalol candidate bleeding into the solution and moving downward. Once this green seed juice goes into the larger NaOH solution it looses it's color. Here is a picture of the situation:



However the nice deep red color is not comming up at this low NaOH concentration, so I brought it up to 2.5% for bulk extraction of the candidate.

I'm thinking of requesting Endlessness for GC analysis of the 3 harmalol candidates above unless someone can find any issues with the work so far. I don't want to waste the chance for a free analysis.