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How is LSD tested quantitatively?

Solipsis

Bluelight Crew
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This is done for free in my country, but I must admit I don't know how it's done. Some sort of titration or by using a (UV?) spectrometer testing color intensity of a standard quantity of Ehrlich's applied to an extracted blotter hit? MS (as I imagine the normal spectrometry is inaccurate?
 
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I'd hazard a guess, by derivatization with Van Urk, Erlich's or Kovac's reagent, and measuring the intensity of fluorescence quite possibly in combination with fluorescein, which is helpful at least when testing plates containing colonies of Claviceps purpurea, or other Clavicipitalean fungi (although I myself have only worked with C.purpurea, and what I believe to be C.sulcata. Possibly also C.microcephala also, but the one most attention is paid to has been the most available one, C.purpurea. Wish I could find some C.paspali though, but I've never been lucky enough to see it in the wild

I don't have a UV or UV-VIS spectrophotometer yet, but even using blacklight exposure to part of the colony, if the agar medium is first doped with a bit of fluorescein before innoculation then once one's plates grow out there is a far, FAR better ability, even by eye to separate higher productive strains for the subsequent mutagenesis work, It gives much better contrast.
If I was using UV-VIS, I would add a standardized quantity of both fluorescein and van urk reagent to a sample containing no ergoloids, and then to a series of reference standards of increasing concentration, and measure the intensity of fluorescence, to obtain a calibration curve, and then subject the extracts, in the same volume of solvent that the samples for the calibration curve were done in.

Perhaps, although I'd have to read into this more, even addition of known reference quantities to the test sample, measuring the differential between the calibration quantity on its own, the sample on its own and the two together.
 
Titration is only effective with a pure compound -- most lysergamides yield similar color changes in reagent tests. The usual method of quantification is HPLC with mass spec. Measuring the area of the LSD peak allows the amount to be quantified.
 
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Well yes, I should have clarified that, I was referring to LSD itself.

5HT2a, could you perhaps go into UV/UV-VIS techniques, am I on the mark here?
 
I think some wavelengths - like UVA - can actually be detected by CMOS smartphone cameras. If you can properly calibrate it, a 5% error margin can be achieved. Not sure if it's the right wavelength (or if every proper wavelength emitted by LSD activated reagent including in the visible spectrum could be used), and also don't know if actual LSD must be used to calibrate since other indoles with added reagent may not have the same spectrum and emission at the measured wavelength; seems like it's necessary. But that would be amazing if a spectrometer is not strictly necessary to test blotter potency yourself if you can test a reference sample one time (quantitatively tested and used to calibrate under certain conditions you standardize: always the exact same lighting and placement etc).

(I have a UV light for checking counterfeit currency that I guess is 365 nm off the top of my head, meant for TLC... if it could be focused and if I had a cuvette - I wonder)

Yeah I noticed HPLC with MS is used for like tox screens on bodily fluids, but it seemed uncertain if that kind of separation is strictly necessary to analyze blotters.

I figured that since IIRC GC requires only very small amounts you could construct the analysis so that you use a certain fraction of the sample to ID with GC just like I think powders are usually tested (aren't they?) after some preliminary reagent checks, and do mass or UV-VIS with the rest of the sample. Is HPLC strictly necessary? Or is it routine and facile if the sample is quite pure? I don't remember having done HPLC or if it's like column chromatography but that was usually a lot of work which I would find amazing if it would be done on LSD samples.

DMT Nexus says the 2 method combinations used for quantative LSD testing is:
- GC / MS
- HPLC / UV-VIS

But only with partial reference: https://www.erowid.org/chemicals/lsd/lsd_article1.shtml

Which nicely provides the info that here in The Netherlands, HPLC was used which was occasionally confirmed with GC/MS. However it's quite possible that in the meantime the national service switched to another lab to facilitate this (DSM) - this switch happened, I mean it's possible or likely it happened since erowid posted that.

If you accept that HPLC cannot be done at home and just try quantitative testing via UV-VIS operating under the assumption that the primary active present is LSD (at least I virtually never got weird results other than some quantity of LSD present whether high or low, so for me it would be alright. It's unfortunate if impure crystal cannot be accounted for, only total lysergamides) maybe it could work. Am just curious - I can just submit a sample to the lab, but it would be a fun experiment to see if the result can be replicated. Question is: what is the best visible spectrum wavelength to measure from fluorescence? I guess first obtain a total spectrum with and without UV irradiation and compare emissions? Oh wait, it's mentioned there on the erowid page: 320 nm is where it's absorbed.

Seems that using Ehrlich's and the purple pink wavelengths and checking UV absorption are two separate ideas. I don't know - if a smartphone can be used at all - whether it is better to try and detect UV absorption or UV re-emission at an angle or just add an exact quantity of Ehrlich's and measure visible. The Ehrlich's idea introduces another order / degree that possibly makes it less linear or reliable than plain UV reaction - so if UV works that's the first thing to try.
 
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Actually it is not LSD I am interested in. It is ergovaline. And ergotamine, ergocristine, ergokryptines.
What about Van Urk's? I'm more familiar with it, never used Erlich's. And the combination of either Van Urk/fluorescein or Erlich's/fluorescein.

And yes phone-based specs using CDs as a diffraction grating, have been done. Can't remember if it was UV-VIS or IR though.

HPLC I would find tough. Would have better luck with mass spec than HPLC. (thats not sarcasm, although building a mass spec would be a terribly expensive project for me. Qualitative analysis is what I am after, less fussed about quantitative. That can be done with TLC, at least, for the purposes *I* have in mind. Which have absolutely bollocks to do with extracting anything off blotters, and I will have a good idea of what potentially is going to be in the mixtures. Depends on the strains of ergot. Or endophytes if I ever decide to fuck with the grass endophytes like Epichloe/Neotyphodium lolii. I don't need to know how pure LSD is on blotters. Because I don't HAVE blotters of acid, and I am not trying to find them. Its ergot strain chemotyping I am after. And ratios.

HPLC, I can't afford that. And I don't think I could make one easily. (note when I say easily, I have a somewhat different opinion of 'easily' and of how high I'd set my own personal bar as for trying and what can be accessed and made. I do have access to at least a half decent machine shop, lathe, for precision metal parts etc, One idea I've been somewhat toying with, mulling over, is the use of an orbitrap ion-trap mass spec.

https://en.wikipedia.org/wiki/Orbitrap

And a question-can calutrons be used for complex mass analysis of a set of possible compounds of known likely identity, in a mixture, and how would one process the signal? I know they are more used for isotopic separation, which in and of itself is something I'd love to have a crack at (no, not for refining plutonium, just any old element with a couple of natural radioisotopes and nonradioactive isotopes. Potassium, and 40K would do me just fine, especially as potassium is volatile enough and easy enough to melt and ionize, and easy to obtain as the metal.
 
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Is TLC then what you personally employ to double check progress of reactions in multi step synths? (or even less than multi)? I've been meaning to ask...

Tell me, what's the real difference with Van Urk's? :) Seems like just like Ehrlich's it's pDMAB with a strong acid, but Van Urk is apparently maybe even a tad more selective and sensitive

But you need some chromatography to separate those alkaloid mixtures into phases? Just a silica column then? Or do you really need zeolite for those bastard lysergolines?

Building that kind of equipment sounds mad, I like it!

Meanwhile I'm just sticking to the plan of using a microcontroller to fully regulate a grow hood for herbs and succulents and stuff.

(PS I checked if maybe boletes were popping up in a region where there has been absolute overkill in previous years, but no luck so far... I wonder what it's a good time for now. Also I have ergot but it's just a small old sample and I have no idea if it's basically long dead)
 
Got no experience with column chromatography. And at the moment basically, trying to get the buggers productive. Testing on a plate level. Culture the fungus, and once the plate has colonized then divide in half (sterile conditions of course) and test. Like I said, can't compare personal experience with Erlich's, not got any.

PM me about ergot if you wish. Will test my samples for life, and ive been really wanting to look for more.. Mine is from last year but have been climbing the gods damned styxfuckingbegotten bastard WALLS wanting to go out to my little spot and search. It was prolific as hell last year and the two before. Right now though I cannot go out of the house, bar short car-to-shop and back, getting a lift (can't drive, too poor, my family couldn't afford to support the driving lessons, and I had to stop, business making guitar parts on the lathe fell through, uneconomical in the end and couldn't keep it going.

The major problems with ergot (this applies to C.purpurea, I've not read up so much on C.paspali because I haven't GOT any Claviceps paspali:p I DO have C.purpurea, possibly some C.sulcata mixed in, a certain proportion of the sclerotiae have a very different shape to the rest, not rounded but of a distinctly triangular profile, and with three deep longitudinal grooves running from pole to pole. Which according to my copy of Kren and Kvac's book, 'The Genus Claviceps' (this is available on ebook format, I don't have a copy on this computer, did on my other one but the screen is busted. And I didn't download another copy, because I went and bought it in hardback, so I can for example, take it with me when going to pick up my meds from the pharmacy when my scripts are due, or if I need take a journey on public transport, et cetera, and for some light, easygoing material to read for those annoying nights where you just can't sleep and the only thing to do are either stay up playing videogames, stay up in the lab, burrowing away like a reagent-devouring blowfly maggot-stroke-Gollum chimaera created by some fiendish mad scientist's experimenting.

Boletes? what kinda boletes? in my ergot spot theres such a profusion of fungi, its a lovely walk down the naturalized, non-paved, woodland and tree-enclosed, uncivilized half of the towpath, there be fly agarics in profusion, to dry for the winter, and the coming year, and to cook like blushers (boil, toss water, boil, toss water, rinse in boiling water quickly and then cook however, similar to how idiots eat those false morels, something I wouldn't touch with a bargepole covered in osama bin laden's fermenting typhoid-dysenteric-haemosquirtings (like the hershey squirts, only much, much worse and with even less in common with even commercial milk chocolate, but fly agaric, oh yes, and blusher, the young Amanita muscaria are the best, they stay the firmest. And blusher, Amanita rubescens, they are NOT to be eaten raw, as they contain a thermolabile haemolytic poison called rubescenslysin, but if they are treated similar to preparing fly agaric for the table, twice parboiling well at least, and then cooking how desired, it leaches out and destabilizes the haemolysin, and they are a valuable early-appearing species, ofttimes popping up when there is little else wild and fungal to be munched.)

Saw the amathyst Laccaria, a relative of the deceiver (Laccaria laccata), the other year, never seen them before, but they are so beautiful. Weird tiny caps, with bulbous-based stems, thick at the bottom and growing steadily thinner, but best of all this really gorgeous shade of deep, vivid violet. Tried eating them, for they are indeed edible, but oh, oh mein gott! I fried a handful or two up to try with my haul of Boletus badius (of late, transferred to share a family with the red cracking bolete, now placed with it in Xerocomus), its the bay bolete, blues like a nun caught bonking her pet golden retriever, pores start pale, turn dull yellow, never red, very tasty, got me a few ceps, which were really good, as good as they ever are, and they always are yummy. Seen various Russula ( brittlegills, name needs no explaining, although all the tissue is brittle and snaps easily, they do not bleed milk, although the genus is closely tied to the genus Lactarius, the milk caps. Last year went to Delamere forrest, and found a whole load of Lactarius deliciosus under the pine woods, its a bright orange species, orange milk, slightly greening with age on the cap and bruises of the stem especially, sweetish taste, and they are a very, very sought after species, lovely roasted, not bad either if given a wee splotch of salted bugger. Real stuff not that shitty vegetable oil based spread, just a light sizzling in a frying pan until cooked through with brownedb ugger covering thefruit bodies.

Theres a lookalike to L.deliciosus though, Lactarius deterrimus, which looks very much like the saffron milkcap, L.deliciosus, grows similar habitat, open pine woods, much more tendency to age green, and less pitted, a more gracile build generally speaking than the saffrons, although otherwise similar in most respects, it too is edible, but often somewhat bitterish and apparently, although I've never found any, not nearly so good as the delectable saffron milkcaps.

This year, best finds so far, were two giant puffballs over at lymm dam. Had to scrabble down a steep, steep drop down a muddy as fuck messy embankment after hopping the fence, and stop myself falling into the dam by means of grabbing saplings and small trees one after the other, grab the puffballs and SOMEHOW, made it back up without getting a thorough soaking or eating my bodyweight in mud and leaf litter-D

Each was about a foot in diameter, quite a respectable weight to them, perfectly, perfectly formed and at that stage of growth where they are at their salty, succulent, scrumptious white all through and through best. Knocked up some batter by making a slurry of brown granary bread with various nuts and seeds in it in whipped egg and a pinch of salt, and some eggy bread over the top, then a wee bit of soy and worcestershire sauce, heated up a pan with some salted proper butter, melted, made sure it was ready to quickly sear, after first cutting the puffballs into 'steaks' about three quarters of an inch thick, covered in eggy bread batter, then fried in the worcestershire/bit of soy-sauce-doped butter.

And tried some too with a mixture of a wee dash of powdered cubeb (the seed of Piper cubeba, has a cold, camphoraceous taste, not entirely removed from that of cardamom but without anyt hint of its warmth, ) just a tiny dash of those, powdered in a spice grinder and added to powdered pink peppercorns, their sweetness mixed with a hint of camphoraceousness complimented the cubeb quite well

Fried again in the form of puffball steaks. Absolute heaven. I saved (unfortunately my old man threw the bulk of the dried part out) but not before I spiked the garden with buried chunks of spore mass all around. I'm hoping that the gleba from the two different puffballs will form hyphae and eventually, if I am lucky, pop up puffballs next year.

(another thing I am working on, is to seed the lawn with Hygrocybe calyptraeformis, one of the waxcaps, because I know where to find it, and that the waxcaps and H.calyptraeformis itself, grows in association, unusually for fungi I have observed at least, with mosses. Its a slender, very gracile waxcap with a thin-fleshed, bright pink cap, and a white, thin stipe. Rare, only recently been taken off the red list in the UK. I am wishing to propagate the species and seed it around, like a kind of jonny waxcapspore (so to speak)=D

If your interested in ergot, do PM me, I have much...knowledge to share, if you wish it;)

-Russula-species, some of them snackable some of them not (rule of thumb, if its hot, for the pot, its not, most mild ones are edible, save some with nasty ass smells although of course always ID. Charcoal burners are prolific sometimes there (R.cyanoxantha, or as my dad calls them, 'those grey Russulas'
 
On the topic of LSD testing - HPLC with UV detection can be more than enough to quantitate LSD without derivatization - assuming you have access to a standard of known purity and can make serial dilutions to generate a calibration curve. If you have the ability to measure fluorescence spectra of the peaks that's useful as well, but I would imagine that AL-LAD, PRO-LAD, ALD52 and so on are quite similar in fluorescence spectra to LSD, so the test is merely a diagnostic of ergot alkaloids being present)

GC-MS or HPLC-MS will give you a structural ID as well, but you are still limited to ballpark guesses if you don't know what the response of your detector setup is (area counts per nanogram of LSD).

Derivitization for GC/HPLC is usually conducted with something like BSTFA, and as a rule of thumb, increases the detector sensitivity and therefore lowers the detection limit. On modern mass spectrometers you can see LSD peaks without derivatization, so it's not strictly necessary, but it's good practice.

On the topic of lab equipment: wait around for deals and buy it used, IMO. If you build a mass spectrometer in your local hackerspace don't expect it to be practical to use for everyday analysis - and expect to spent a pretty penny on parts. Calutrons are ... the worst of the bunch. Relying on intense but homogenous magnetic fields and strong vacuums... that's well outside "hobby chemistry" to me.

For real though, you can piece together a HPLC from bits and bobs for no more than $1-2 grand I'd say. It may not be the shiniest, but functionally all you need are some pumps, valves, a column, and a detector, all of which are fairly abundant in the used lab equipment scene. (Think of all the biotech places that go tits up)

On the topic of column chromatography: Quit wasting time and silica and switch to dry column vacuum chromatography :) I've been using that recently and I must say, it makes the older, classical-chromatography trained chemist jealous of my seperatory powar. Even though I'm doing everything "wrong"!
 
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