Who wants to get into TLC?

[Fractalhead]

Hi ppl,

The purpose of this thread is to gauge what interest there might be amongst the community in obtaining materials for conducting thin layer chromatography (TLC) analyses.

For those of you who don't know, TLC is an analytical technique for the qualitative and semi-quantitative analysis of chemical mixtures (eg. essential oils, alkaloid extracts, biodiesel etc). You can get a great rundown on the basic TLC procedure here:

TLC Procedure

This technique could not only be used for quality control or to find new sources of known active compounds but coupled with chemical fractionation and bioassay, TLC could help us pinpoint new substances worthy of thorough chemical examination. Some of the beauties of the technique are its affordability, ease, rapidity and portability (highly applicable to analysis in the field).

If there is enough interest, I'll see what I can do about making TLC kits available for sale. These kits would basically consist of TLC plates, detection reagents, capillaries, pipettes, beakers and test tubes (for sample prep). Other components such as solvents, reagent sprayers, UV lamps etc are readily available throughout australia and it would be pointless me offering to provide them.

Please let me know if any of you are interested.

[Amulte]

i would be intesnly interested. i have been playing with common sedative herbs for a few years on and off, but have laked the equipment to go further with the extracts. and i imagine that there are a few that have been washed away or broken down before isolation. with the proper equipment i think there will be endless possabilites for new exracts and additives.

PM me if you have more info PLEASE. i have great intrest.

[Guest electro]

i was going to throw in with rev & ph for some chrom paper ... if you are making up kits i may well get one of these instead ...

i dont know about the others ..

id say there is definatley interest in this

[teonanacatl]

yeah im interested, depending on price to.

[gerbil]

I'm definately interested in giving it a go, kits would be great.

[Happy Cadaver]

yep, would be interested.

[Fractalhead]

Wow. That's already more of a response than I was expecting... I think if more people in this community were familiar with the technique and got into the habit of analysing everything they bioassay (including myself), we would all be getting, as T would say, "more scientific mileage" out of our bioassays. It really is quite easy once you have a little practice.

TLC plates are a little bit more expensive than chromo paper but the ability to get better separation of complex mixtures is definitely worth the investment.

Re price: I think I will try to keep the price of the kits down initially by including in them only what people will need in order to have a go at the technique and see if they like it without too much risk. That way, more people will be able to get into it. I think I should be able to put together a minimal kit for about $20-30 maybe with the option to buy extra plates. There will be a few things you need to source locally but most of these will probably already be around the home or down at the hardware store. You should be able to put together a nice little set up for under $50 including the kit.

Sounds like i'd better get to it. Thanks for the support guys. I'll assemble something and report back when I have something specific to offer.

BTW Darcy, that's a beautiful quote.

[ 19. March 2005, 10:15: Message edited by: Fractalhead ]

[Plant Pimp]

This is an excellent idea! I'd love to have a kit!

You say "solvents...are readily available". Where might they be obtained and which ones would you suggest? Since most traditional herbs are steeped in a tea, would water be suitable? I remember using hexane/acetylene mixtures (I think??) at different ratios to get various degrees of separation at an old work-study job. I haven't a clue where to find solvents like these. I thought in this day and age that solvents were tightly controlled to prevent things like meth labs?

[Auxin]

Not sure about canada but in that foreign country south of ya you can get toluene, xylene, 95% ethanol, 70-99% isopropanol, acetone, MEK, kerosene, turpentine, and VM&P naphtha quite easily... and for a bit more $ methanol from the auto stores (as gasoline drier)... never found hexane though

...and if your ok at chem its easy to make isopropyl acetate from vinegar, baking soda, a dead car battery, and 98-99% isopropanol.

If you like living dangerously hexane is the main component of gasoline- it can be distiled out (I strongly advise against it).

[Fractalhead]

Hi PP,

Um, yeah, what Auxin said. You can get a nice range of solvents relatively cheaply by using pure solvents from the hardware store or by distilling them out of impure solvents bought from the hardware store. This usually works out significantly cheaper than buying from chem suppliers. At least in Australia, individuals *can* buy most solvents and reagents but for many things you will need to provide photo ID and fill out an end-user declaration which basically states what you will use the chemical for and that you won't be passing it on to some low-life meth-head. Sourcing solvents from common 'household' products is also a good skill to pick up if you intend to fly around a lot with a TLC kit in your backpack since you can't take your solvents on the plane and you will be hard pressed to find a chem-supplier once you get well off the beaten track (where all the coolest plants are).

Unfortunately, most of the published solvent systems for the separation of our known compounds/extracts use solvents that are a bit tricky to source from domestic products. Therefore, it is important that new methods are developed using materials that are readily available throughout most of the world or can be brought on board a plane in a safe manner. This can certainly be done.

[Fractalhead]

Sorry PP,

I forgot to answer one of your questions. Standard silica gel TLC plates don't like water since it ruins the silica gel layer so with these sorts of plates you can't use water as the developing solvent.

As you point out, many herbs are prepared by steeping in water and one might argue that the most meaningful TLC results for those herbs might come from a water extract applied directly to the plate. However, even a little bit of water in your sample can screw with the separation. You would probably get just as much info about the content of the herb by using an alcohol to extract and applying the alcoholic extract to the plate. This will evaporate easily off the plate and then you can run your plate without a disturbed layer. Alternatively, you can pay a bit extra and get water resistant plates.

[teonanacatl]

we were doing some tlc the other day at uni with chlorophyll and found all our Rf values were different to the books cos they used different solvents, so my question is, is everyone going to use the same solvent which is highly unlikely because otherwise we would need either a pure sample to compare it to or a data base of Rf values for different solvent.

[Fractalhead]

While everyone is free to figure out their own methods by trial and error, someone may find that for a particular kind of extract or class of compounds that a particular system works really well. If this can be made from common solvents then it makes sense for other people to use this same system as it allows direct comparison between individual plates (and Rf values). However, even when the the same solvent system has apparently been used, slight variations in the qualities/proportions of solvents as well as local conditions and slightly variations in plates can cause general shifts in Rf that affect the Rfs of all the compounds by a similar amount. That's where it is very useful to run a mixture of known Rf reference compounds along side your unknown mixture. This might be as simple as a mixture of common food colouring dyes, as long as they are readily available and show a range of Rf values. The Rf of the unknown is then given as a relative Rf (ie. Rf of unknown/Rf of known). These results are usually much more reproducible between experiments and hence more diagnostic (especially when combined with a positive colour reaction).

A database of Rfs, relative Rfs and colour reactions is in the pipeline. I would also like to be able to offer a library of standard compounds/mixtures but this will take some time and obviously won't be practical for all compounds.

[Benzito]

Fractal,

I'm interested too!

very much so.

[Guest electro]

will alcohols like ethanol, methanol and acetone do as solvents for the plates ?

also does the solvent have to be different to the solvent used for the sample ... eg can one use an ethanolic extract of something and then "develop" it on a tlc plate using ethanol as the solvent ?

[ 24. March 2005, 16:50: Message edited by: electro ]

[teonanacatl]

when you put you sample on there you let it dry so providing you used a pretty clean solvent it will evaporate away. acetone seems pretty normal solvent to use.

[Amulte]

Electro - i dont mean to be a whinging perfectionist but acetone is not an alcohol. alcohol is a polar solvent where as acetone is di-polar and petreolium distilate is non-polar. just some useful extraction knowledge. the poles refered to in polar/dipolar/nonpolar relates to the metabolic structure of the compund. fairly trippy when you get into it. sorry again all i just love my chemistry.

FractalHead - how far off will the TLC kits be do ya think?

[Fractalhead]

I'm guessing i'll have the basic kits ready in about two weeks (i'm waiting on a few things).

[Guest electro]

im sure i remember it being referred to as a secondary or tertiary alcohol ...

a quick google says acetone is also known Allylic alcohol

*shrug* i could be wrong through - im certainly no chemist

[Amulte]

hmmm, if i remember rightly (im prolly wrong) but the consumption alcohol falls into the ethol where anyother is a result from molecular bonding with ethol. when i find my old bloody uni notebook ill know. been looking for a week now, dam huge shed.

what copst would be put on a TLC kit again, i know its up in the thread somewhere but shit it got long!

[teonanacatl]

acetone is actually 2-propanone, which is a ketone(sorry if you knew this)

[Fractalhead]

Amulte,

I haven't worked out how much the kits will be yet but the minimal kits will probably be about $20-30. These will include most of the bare essentials to have a go at the technique and become familiar with it. The rest of the bare essentials will need to be sourced locally. I will also provide the option of a more advanced kit and extra TLC plates.

[Amulte]

fractal - yes the advanced would be great just to let you know im keen for that. no probs for me the get the solvents :D :D :D

"...get by with a little help from me friends..."

teonanacatl - Yup your on the nose , i smell ya from here, nah i mean your right. but i didn't remember that, i was way off. found my booky :D

[Fractalhead]

Ok, I've put together something of a demo kit to give you all a rough idea of what the basic kits will be like. Here's an image:

The starter kits will contain:

1 large beaker

1 small beaker

watch glass/beaker cover

10 pyrex test tubes

10 glass pipettes + 2 pipette bulbs

10 glass micro-vials

1 small measuring cylinder

1 large measuring cylinder

10 4 x 10 cm Silica Gel G60 TLC plates (with 254 nm exciteable fluorescent indicator)

~100 glass micro-capillaries (for sample application)

This will all come packed in a plastic storage container to keep it all safe on its travels

** Note that some of the stuff that will be in the kit isn't shown in this picture. **

Firstly, there will be more TLC plates (ie. ten 4x10cm plates). You can cut these up as small as you like to make them go further while you practice. If you cut each 4x10 cm plate into five 4x2cm plates (which you could run in the small beaker) you will get 50 runs out of this kit. On each run, you might fit 3 or four samples so you are talking maybe 100-200 samples before you need to order more plates. Of course you may opt for more complete separation by using longer plates in the large beaker. There will also be an extra (larger) measuring cylinder for making up solvent systems and detection reagents. Also, there will be some filter paper for putting inside the development chamber (beaker) so as to saturate the chamber with solvent vapours.

The beakers can double as both development chambers for running the TLCs aswell as for preparing solvent systems and detection reagents (they also look cool on your kitchen shelf .

You will need to source solvents and things locally as these cannot be sent in the mail. Its amazing what you can do with a bit of acetone, vinegar, metho, ammonia etc.

There are a few highly recommended (though not absolutely essential) additional items that you should consider budgeting for when thinking about getting into this:

* One would be an aerosol propelled "paint" sprayer. You can get these from the big hardware stores and they basically consist of a glass jar with a aerosol can that screws onto the top and sprays the contents of the jar out as a fine mist. These are about $25 and are perfect for spraying detection reagents evenly onto the plates.

* Another thing would be a handheld UV blacklight. With a spectral peak at about 366 nm, these are awesome for showing up all sorts of cool fluorescent compounds without exciting the fluorescent indicator dye that is impregnated into the plates. You can get these lamps from Dickos or Tandy for about 10 or 20 bucks. If you can get your hands on a 254 nm UV light (CAREFUL: DANGEROUS FOR EYES AND SKIN) you can detect UV absorbant compounds as dark spots on the plate against a blue fluorescent background.

I've added everything up (including my time and effort) and worked out these kits will cost $60 + $9.50 p/h. I realise its a bit more than I first invisaged but I felt a few extra bits and pieces would go a long way.

Those of you who are interested should email me at: [email protected] and we can sort something out.

The Ethnowiki TLC Page will evolve rapidly in the near future so people interested in getting into TLC should check there regularly for new info.

Finally, a VERY important point I should make:

These kits are for conducting LEGAL research on LEGAL plants/substances only. While I am more than happy to offer support and advice with regards to conducting general types of TLC analyses once everyone is up and running... I will NOT reply to any emails or PMs asking me explicitly about analysing illicit substances.

[ 11. April 2005, 22:13: Message edited by: Fractalhead ]

[Fractalhead]

I forgot to mention, for those of you who don't need all the extra stuff, I can send just the plates and capillaries for $25 inc postage.