Dear Ciara,

You could try using ion-exchange resins, with the pH of your sample carefully adjusted so that it is below the pKa of the relevant group (i.e. for a cation-exchanger, ensure that the pH is above the relevant pKa for the amino group(s)).  You would still have, in this case, some chloride ions complexed with the counter ion from the resin (H?), so you may have to use both anion- and cation- exchangers to get rid of all of it.  You can also get mixed-mode resins that have both functionalities, but it might get a bit tricky in terms of avoiding adsorption of the amino acids.

You could also have a look at solvent extraction, or see if there is a solvent that will selectively precipitate your sample.  

It's not an easy one, so you'll probably have to do some development work on your compound to work it out.

Hope this helps.

Dave

Dear David,

Thank you so much for your answer! The main resins I have been looking at from different companies all seem to have the perfect properties until I examined pore size and all seem to exclude and end up eluting the amino acids from the samples. My amino acids are organic , but my buffer solution has both inorganic and organic salts so it becomes even more complicated to seperate the salts from the amino acids on this basis. There is extraction/ digestion techniques using nitric acid but these break the amino acid down whereas I need to retain the structure for quantification. I think this one will definitely require some more development work on the compound!

Could I ask what your peptide is and might be able to help more? Cheers Dave

use can carbonate resine

Hi David it is a selenium containing amino acid (0.9 kDA).

If the material has a reasonable degree of hydrophobicity, you should be able to retain it on a reverse-phase column.  Elute the salts with water, then elute the peptide with a gradient of acetonitrile or ethanol.  You can use C-18 SPE or "spin columns" (see links).

http://sampleprep.kinesis-usa.com/products-page/sample-preparation/kinesis-spe-columns-telos-c18-1g6ml-spe-columns/?application=SPE%20Columns&sorbent=&sorbent_mass=&reservoir_volume/format=&membrane=&undefined=&functional_group=&#.VLPRc6b6baQ

http://www.piercenet.com/product/c18-spin-columns

Hi James,

Thanks so much for that. I have been looking at C18 columns and contacted a number of companies, however my amino acids which are about 0.96kDa are too small to be retained on the columns.

Dear Ciara,

I wonder if you can exploit the selenium-containing nature of your compound in some way.  I think there are some proteins, such as I think some thioredoxins, that contain selenocysteine.  It might be worth spending some time and finding out if there are any protocols specific to selenocysteine-containing proteins.  You might be able to use any affinity resins that are used in these methods, on your amino acid.  Sorry I can't be more specific, but it might be worth a little look on the internet.

Cheers,

Dave

The size of the molecules has nothing to do with whether the compounds are retained on a C18 column - it's entirley a matter of Van Der Waals forces.  Small molecule drugs, even benzene and toluene, are adsorbed from water and eluted with a solvent gradient, and peptides are routinely separated. (See linked datasheets.) If anything, you might want to check that the pore size of the silica will accomodate your peptide. 

http://www.hamiltoncompany.com/downloads/PRP-C18_ProductDataSheet.pdf

http://www.sigmaaldrich.com/analytical-chromatography/hplc/columns/ascentis-express/peptide.html

Hi James, thanks for that.

I use a C18 column currently for HPLC detection of my amino acids which is fine. I have issue detecting in a high salt solution(HBSS) compared to when it is prepared in 0.1M HCL.  So I was looking for a desalting method to use to avoid interference pre HPLC analysis. The problem I was finding due to size, is that many of the desalting columns made a point of eluting such small amino acids classing them as contaminants only retaining the higher molecular weight components of the sample.  I can detect my amino acids successfully using an isocratic method for HPLC analysis when it is diluted in HCL but not HBSS. I have seen HPLC methods with a higher organic phase and gradient methods but the problem lies with using a large amounts of salts on the column which we want to avoid.

Pierce C18 Tips, 100µL bed

I am going to try these out. I will let you know how I get on.

Thanks all!

Hi Ciara,

I would like to know how you went with desalting your amino acids? I have a similar problem, but with organic acids instead.

I have exactly this same problem; i tried ZipTips with increased C18 bed volume and it didn't work at all. This problem is quite difficult so solve especially for small and very polar molecules. IE would be a way out only if you can work with bigger volumes and have enough of the material so you don't need to worry about dilution/ concentration factors, or even loosing some of the material. I tried also precipitation with various different solvents and solvents mixtures and I had no luck. I think we can only keep on looking. Good luck! 

TiO2 will bind cis diols. If you have this functionality on your targets then you could use this approach.