[magnetic111]

Which chromatography procedure should be used for small samples 10 mg and less?

[Furanone]

You need to supply more information about your samples than the sample size, most importantly what analytes you are wishing to measure.

A general rule of thumb is to always use gas chromatography if it is possible, meaning the analytes to be measured in the samples are thermally stable and volatile (or can be derivatized to be made volatile such that they can be carried by the gas phase). The reason for this rule of thumb is the unsurpassed resolution and sharp peak shape in GC. Combined with the flame ionization detector, the GC-FID is very sensitive and low maintenance instrument for most organic compounds. Identity of compounds can be predicted based on using the Kovats retention index (based on the column you have -- go to http://webbook.nist.gov/chemistry/) but true identification will be only possible using GC-MS (mass spec). If you know your sample though and have access to journal articles, this will show you what to expect for compounds and this makes it a lot easier. For sugars, organic acids and amino acids, MSTFA is the most powerful derivatizing agent and will make these compounds able to be analyzed by GC.

Otherwise, HPLC is your next step up. Here you need to find what will dissolve your compound (reverse phase for water/methanol/acetonitrile soluble compounds and normal phase for compounds soluble in more hydrophobic organic solvents). Also, you will need to select the right detector to use. Where there is conjugated dienes (such as phenolics, carotenoids or proteins due to tyrosine and tryptophan), a variable wavelngth or diode array detector may be used. Otherwise, an evaporative light scattering detector or refractive index detector (no gradient elution) may be required for things such as sugars or fats.

Finally, if you are working with large polymers, you will need to use size exclusion chromatography which can be classified as gel permeation chromatography (hydrophilic water soluble polymers) or gel filtration chromatography (hydrophobic organic soluble polymers), and then you will have to select your column based on the MW range of the compounds you wish to separate. Resolution with this technique is rather poor, so ofter it is just used to characterize your sample based on sMW size distribution rather than separating by individual compounds. If you have proteins, SDS-PAGE (electrophoresis) with quantification using densitometry will be far better for resolution of proteins.

[magnetic111]

Thanks Furanone. 

I was myself confused by this question.  This question appeared in one of my assignments.  No further sample information was supplied.

[Dan]

Preparative thin layer chromatography (a.k.a. PTLC, prep-TLC) is the easiest method for <10 mg of most small organic molecules, and does not require expensive equipment.

Prep-GC and prep-HPLC are not standard pieces of kit (in my experience) - they will of course outperform prep-TLC, but I thought it was worth mentioning.

[OC pro]

Why not column chromatography? Use a small glass column with 0,5cm diameter and fractions of ~10ml size.

[fledarmus]

In my previous lab I would have used a Chromatotron - a very convenient method of prep TLC where the plate is spinning and you collect the fractions as they come off the end, rather than developing the plate and trying to scrape off the fractions after it is developed. In my current lab, either a small (10 gm) flash column, or semi-prep HPLC depending on the compound to be purified. The "correct" answer for a university lab is probably prep TLC.

I presume you are trying to purify the 10 mg of material, not just get a chromatogram...

[Doc Oc]

I agree with OC Pro and fledarmus.  For very small amounts like that, I do a microscale flash column, which can be done by packing a 5 3/4" Pasteur pipet with silica and collecting fractions between 0.5 and 1 mL, depending on how good your separation is.

I've also used semiprep LC for this amount of material, though I don't think this is a common type of column for people to have.

[james_a]

Second/third vote for either a pipet column or for prep TLC. These are the fastest and cheapest options for purification at this scale. If you require publication-quality spectra, then prep HPLC would be the next direction to go.