Tryptamine Turns Purple with Ehrlich Reagent

— by: earth, Sylvia, Fire, Jurek, and anonymous experts

Here’s a peek into how Erowid works with a network of drug-checking experts around the world work. Just another day at DrugsData. :]

On June 30, we published the test results for a sample of 1P-LSD blotter (dd10683), confirming the presence of 1P-LSD.

On July 12, Jurek from protestkit.eu, a Polish harm reduction and field reagent specialist, inquired about this sample, noting that the Ehrlich reagent photo showed an unexpected purple reaction. Jurek pointed out that 1P-LSD isn’t known to result in a purple color in the presence of Ehrlich reagent, helping to differentiate it from LSD-25, which does cause a purple color change with Ehrlich reagent.

We discussed this with our lab and learned that there was a small GC peak they had not initially reported in the results: inactive salts and inks on blotter do not always get reported due to DEA-imposed limitations.

Given the unexpected Ehrlich reaction, we published the spectrum for the unidentified chemical and added it to the results as a second chemical present in the sample.

A chemist in the Erowid Expert Network identified the unknown chemical as tryptamine, so we ordered a lab standard for tryptamine and found that it was a perfect match via GC/MS.

Further, DrugsData’s lab did side-by-side comparison in a ceramic well plate of lab standards for 1B-LSD, 1P-LSD, and LSD-25. The third of four wells is the ‘blank’ labeled MeCN (acetonitrile) which was the solvent used to dissolve each of the ergoloid standards (1B-LSD, 1P-LSD, LSD-25). Ehrlich reagent was applied to each, demonstrating that neither 1B-LSD nor 1P-LSD turn purple with Ehrlich, where LSD-25 does.

So the mystery of the the unexpected Ehlrich reaction for this 1P-LSD blotter is resolved, but the reason why someone added tryptamine to 1P-LSD blotter is still open. We all guess the goal was to be able to sell the 1P-LSD blotter as LSD-25, and that adding tryptamine to the 1P-LSD will result in reagent reactions consistent with LSD-25.

This is the first time Erowid has seen this type of adulteration of non-LSD ergoloids with the chemical tryptamine.

The image below is a link to a video of the reagent test:

Then, a photo of lab-grade tryptamine reacted with Ehrlich. A strong purple color:

“Intractable Byproduct” in 5-MeO-DMT Samples

Erowid’s DrugsData project recently tested two samples of 5-MeO-DMT that both contained an unidentified chemical. The first was dd10559, published Jun 08, 2021 and the second was dd10808, published July 19, 2021. Both samples were sold as 5-MeO-DMT and were reportedly sourced from the Netherlands to California. The unidentified chemical in the two samples appeared to be the same substance.

In June, one of our EEN experts (Eddee) proposed a possible identification for the chemical in the first sample, and we began consulting others in our network. Once we received the second sample, with apparently the same unidentified chemical, an outside expert weighed in with a slightly different proposed identification. We examined these more closely and with Eddee’s help, we think we’ve finalized our current opinion on the identity of the chemical dd10559-unid1 and dd10808-unid1:

6-methoxy-2-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole

This chemical is likely an unwanted byproduct resulting from imperfect synthesis of 5-MeO-DMT. Borax, one of Erowid’s main chemistry experts, proposed the name “N-methyl-Pinoline”, and Eddee proposed “N-methyl-5-Methoxytryptoline”.

Another expert pointed out that PubChem’s synonym list for this chemical includes 2-Methyl-6-methoxy-1,2,3,4-tetrahydro-beta-carboline (CAS# 6582-80-5), and cites the Japan Chemical Substance Dictionary for the CAS#. Isomer Design, a longtime supporter of the DrugsData project, refers to it as 6-MeO-2-Me-THβC.

We do not believe it has any common trivial name, and are currently settling on the name “N-methyl-Pinoline”. Its structure:

There were no published GC/MS graphs for this chemical. The identification is based on analysis of the fragmentation pattern, and on a 2020 paper, Synthesis and Characterization of 5‑MeO-DMT Succinate for Clinical Use, by Sherwood et al. Because of this paper, we originally considered calling this chemical “5-MeO-DMT Synthesis Byproduct A”, to parallel names given to unwanted synthesis products in other drugs.

The other proposed identification was the very similar compound 2,3,4,5-Tetrahydro-8-methoxy-2-methyl-1H-pyrido[4,3-b]indole (CAS# 41505-84-4).

The two proposed identified chemical structures have the same number of elements and chemical formula: C13 H16 N2 O ( C13H16N2O ). The only difference is which order the carbons are coming off the indole ring relative to the amine nitrogen.

This very technical image shows a comparison of the two chemicals, with our proposed ID in the upper panel and CAS 41505-84-4 in the bottom:

Now look again at the drawing of our identification and note the red line showing the bond between the indole ring and the methyl (carbon) coming off the nitrogen.

Finally, take a look at the structure of 5-MeO-DMT:

Imagine that the red line in the structure of N-methyl-Pinoline was broken where it connects to main rings. That substance, with that connection free, is 5-MeO-DMT. 5-MeO-DMT just happens to also have the same chemical formula: C13H18N2O.

Quoting from the Sherwood et al. 2020 paper: “Several small-scale attempts were initially evaluated with reaction monitoring by liquid chromatography-mass spectrometry (LCMS). Though product formation was evident, the reaction was plagued by challenges that would likely multiply at larger scales. The Pictet−Spengler reaction to the corresponding tryptoline (8) was difficult to suppress and removal of this structurally similar and possibly biologically active byproduct was challenging. Further optimization to Route 1 may be possible, but ultimately, the reaction was not recommended for further development.”

Their Scheme 1 Graphic shows what they label the “intractable byproduct”, which is the chemical we are proposing as the identification of the impurity in these two 5-MeO-DMT samples.

Thanks to everyone who participated in this: the submitter of the samples, Eddee, our anonymous experts, Borax, Sylvia, and the authors Sherwood AM, Claveau R, Lancelotta R, Kaylo KW, and Lenoch K for their excellent 2020 paper, which nailed down the reason for this unwanted contaminant in these synthetic 5-MeO-DMT samples.

Name Change from Levamisole to Tetramisole/Levamisole

Erowid’s DrugsData lab has recently changed from reporting simply “levamisole” to instead reporting “tetramisole/levamisole”. Tetramisole is the “real” primary name for this substance, a chemical that has two stereoisomers/enantiomers. For those unfamiliar, left (“lev”) and right (“dex”) isomers are only different in the same way that left- and right-handed gloves are different. They are also referred to as the “S-enantiomer” (lev) and the “R-enantiomer” (dex) of tetramisole. (Tetramisole refers to the “racemic” or mix of the S-enantiomer and R-enantiomer of the chemical.)

Many (though certainly not all) chemicals have this type of physical isomerism. An update in 2020 to the SWGDrug library that our lab uses as one of its sources changed the way it reports the name of the substance upon match. Neither the chief chemist at our lab nor any of our team remembers ever seeing the name “tetramisole” before December 2020. The technical distinction and update merits some further explanation.

The technical language can be pretty confusing. In some cases, the FDA allows commercial pharmaceuticals to use a shorthand, where they prepend “es” or “ar” (or “lev” and “dex”) to the front of a pharmaceutical name to denote an enantiomer-specific product. Examples include “armodafinil”, “eszopiclone”, and “escitalopram”.

Erowid’s DrugsData lab, using GC/MS testing, has no ability to isolate, separate, or identify which enantiomer is present, yet we’ve previously reported “levamisole” (the left or S-enantiomer) since first identifying this substance in cocaine samples in 2009 (see DrugsData Levamisole Results). Our lab’s techniques can’t distinguish the stereoisomer composition of any substance we analyze, e.g. a specific chemical identified in a single sample could be a 50/50 mix of R-enantiomer & S-enantiomer; 100% R-enantiomer and 0% S-enantiomer; 0% R-enantiomer and 100% S-enantiomer; or any ratio combination of the different physical isomers. This would be true for any lab using GC/MS for its testing process.

While DrugsData has been reporting “levamisole” since 2009, the question of enantiomers had not come up before. However, the “lev-” syllable should have been a giveaway. Sorry we didn’t identify this issue earlier!
In the last few years, research appears to indicate that most of the “levamisole” mixed with cocaine is actually the racemic tetramisole. In 2019, Madry et al. seemed to nail this issue down through analysis of hair samples from 627 cocaine users. The authors write: “Samples mainly contained racemic tetramisole (87.5%), only one sample contained levamisole only and two samples contained non-racemic [tetramisole].”

All of that, combined with our inability to test which spatial isomer we have in the samples we analyze, means that we’re going to switch to using “tetramisole” as the primary name for this substance, with “levamisole” being included so as to help avoid confusion due to the switch.

Reference:
Madry MM, Kraemer T, Baumgartner MR. “Cocaine adulteration with the anthelminthic tetramisole (levamisole/dexamisole): Long-term monitoring of its intake by chiral LC-MS/MS analysis of cocaine-positive hair samples”. Drug Test Anal. 2019 Mar;11(3):472-478. doi: 10.1002/dta.2505. Epub 2018 Oct 17. PMID: 30239147. Erowid Ref9448

Evolving the Approach to Reagents and LSD Gel Tabs

Drug checking is a complex and evolving area of research. In EcstasyData’s effort to show accurate findings to the public, we’re working with the unique conditions of each sample. Most recently, the lab has innovated in its handling of LSD gel tabs.

There’s LSD, and then there’s gel tab LSD

Since 2014, the year EcstasyData’s lab developed its procedure for a practical and time-efficient way to identify LSD using GC/MS, gel tabs have been infrequently submitted for analysis. The majority of LSD samples submitted to our lab use blotter paper as the carrier (the lab requires that all samples be dry, no liquid samples are accepted without prior arrangement), though it is Erowid Center’s opinion that most of the LSD currently in distribution is in liquid/solution form.

Prior to 2017, the rare gel tab sample would get refused by the lab’s main chemist, who at the time did not feel confident that these samples could be adequately analyzed for the presence of LSD.

Besides analyzing each sample using GC/MS, which is the analytical method EcstasyData uses to detect the presence of chemicals, the lab also tests samples with reagents. Reagent testing adds descriptive data that adds to the collective knowledge base for drug checking. (Reagent testing can’t positively identify chemicals.)

It turns out that gelatin as a medium makes reagent testing more complicated; the pH conditions required to dissolve the gel affect the reagent even when dried. For this reason, gel tabs do not react normally to field reagents such as Marquis or Ehrlich.

De-weirding reagent colors

Between 2017 and November 2018, five gel tab samples were analyzed by EcstasyData, with GC/MS showing that four of them were LSD. The Ehrlich reagent reactions for these four samples were atypical. LSD normally reacts to Ehrlich reagent by turning purple, but when Ehrlich was applied directly to the dry (or even wet) gelatin in these cases, the results were mixed, turning brown or brown-purple, or other atypical reactions.

The lab began working with the special needs of these samples, and in November 2018, they developed a sample-preparation procedure that allows Ehrlich reagent to show a typical positive (rule-in) response to LSD in gel tabs.

New process for gelatin

We are publishing Erowid Center / DDL’s new procedure that is being used to process dry-gelatin-tab dose units, for the historical record, and so that others can duplicate it and critique it.

The following is the procedure that was used to produce the photo shown for Sample 6813, the first sample treated in this way:

  1. Gel medium placed in small amount of water.
  2. Basify gel-water mixture with NaOH.
  3. Gel medium fully dissolves.
  4. Solvent (ethyl acetate) added to gel-water mixture.
  5. Solvent separated off and dropped onto ceramic well plate.
  6. Unheated evaporation of solvent until dry.
  7. Drop field reagents into wells, photograph.

This is the process that the lab will use to prep future gel tab samples for reagents. It will be interesting to see how other samples respond to it, and whether further tinkering with the process will be required.

Lab Drug Checking and Positional Isomers: 2-, 3-, or 4-MEC

This last week, a sample was submitted and analyzed through EcstasyData that we clearly established was one of the three main ring-positional isomers of Methylethylcathinone (aka MEC). However, we didn’t have the lab standards on hand for this chemical, because it is the first time we’ve run into it since the supplier of lab standards we order from has stocked the three positional isomers.

Based on library matches* alone, it was impossible to be certain whether the sample contained 2-MEC, 3-MEC, or 4-MEC. We’ve run into this issue of positional isomers a bunch of times before over the last sixteen years of operating our street drug analysis project.

Luckily, Cayman Chemicals is a really great source of lab standards for NPS (“new psychoactive substances” aka psychoactive research chemicals). So we ordered reference standards for 2-MEC, 3-MEC, and 4-MEC, to find out if our equipment and lab procedures could make use of having the actual verified isomers on hand, for the purpose of confirming whether sample #5682 contained one or more of these slightly different versions of the same parent compound.

We ordered the standards on September 5th and they arrived at Drug Detection Laboratories (the lab that EcstasyData contracts with) on the 7th. The amazing DDL lab team, working on Saturday, ran the standards through their GC/MS and were able to confirm that sample #5682 contains only 4-Methylethylcathinone and none of the other two positional isomers. Yay!

There are many psychoactive chemicals with positional isomers that are difficult to reliably differentiate using GC/MS or UV absorption, even with the proper standards on hand. We’ve spent a lot of time over the last few years seeking clarity in our analysis of fluorinated amphetamines (2-, 3-, or 4-Fluoroamphetamine aka 4-FA) and the *-APB chemicals. And we’ve not been entirely successful. In most cases, one of the positional isomers is easy to tell apart from the others, but the other versions overlap in complex ways by retention time or fragmentation patterns. For MEC, the differences in column retention times for each of the positional isomers make them easy to differentiate using DDL’s Agilent GC/MS.

On another note, we also ordered a lab standard for Benzyl fentanyl this week, and were able to confirm that sample #5667 contains only Benzyl fentanyl. Less impressive, since we were pretty certain that’s what it was to begin with, but the initial match was based on comparing against other published spectra and not our own lab’s confirmation using a known standard of the same substance.

by Earth & Sylvia

 


Notes

*Identifying “by library match” refers to the process of comparing images of GC/MS output for a given sample to images of GC/MS output for a verified reference standard. Because equipment and lab procedures vary, to double-check identification by library match, a lab can acquire its own sample of a reference standard (if one is available), run it through the lab’s equipment, and compare the resulting images to those of the submitted sample being analyzed.