http://www.omicsonline.org/open-access/gas-chromatographic-determination-of-amino-acids-and-polyamines-in-human-skin-samples-using-trifluoroacetylacetone-2157-7064.1000248.pdf
I am attempting to reproduce the protocols using trifluoroacetylacetone and ethyl or isobutyl chloroformate. I've been using just ethyl chloroformate and ethanol for doing GC of amino acids for some time. I find I need a 1701 column and GC/MS to assist with Serine, Threonine and it does not do arginine at all. Needless to say These papers really caught my eye for the ability to do arg and the simple chromatography on a db-5 and FID.
So far using their protocol I fail but adapting the chloroformate mix and matching the alkyl chloroformate and alcohol I get predominantly tradionional chloroformate derivatives without the schiff base on the amine from the FAA. Anyone else get the protocol to work? Right now I am shifting to focus on arginine with is reported to form a pyrimidinyl derivative when reacted with beta diketones.
So far I've tried over 2 dozen variations of their protocol. I've tried using a phosphate buffer instead of ammonium acetate, altered the solvent composition, used ethanol methanol, methyl ethyl and isobutyl chloroformate I've brought the solvent mix for the chloroformate derivatization more in line with other papers (their protocol has too much water and not enough pyridine) I've tried using high excesses of the trifluoroacetyl acetone (past papers used 2 parts ethanol to 1 part aqueous buffered sample) and increased the incubation time to as long as 4 hours. I can get chloroformate derivatives but no schiff bases.
I'm kind of sceptical. acetylacetone reacts with primary amines yet they show proline right in line with the other amino acids. I am wondering if because of the mismatch between alcohol and chloroformate they have methyl esters from the methanol and ethyl or isobutyl carbamates instead of schiff base esters.
I am now focusing on getting it to do arginine. It has been reported in the past that arginine condenses with 2,4 diones to form pyrimidyl derivates which I could hopefully then derivatized with ethyl chloroformate on the amine and acid.
Darn their chromatograms are so beautiful and on a db-5 column (I need to use a 1701 and a MS).
Hi I am just curious? did you figure it? I am working with Chloroformates too now
No I gave up on the trifluoroacety acetone. Those papers really excited me especially the good response and resolution and ability to do arginine. I am instead going to keep doing ethyl chloroformate derivatization. I just could absolutely not reproduce the results in those papers.
Hi, have you detect 20 amino acids now? can you tell me the protocol? I am analyzing amino acids now with MSTFA but I cannot separate proline and isoleucine peaks, and I cannot detect glutamine and arginine. I changed my mthod to Ethyl chloroformate and MSTFA （2 step derivatization) but also failed.
I still use ethyl chloroformate. I thought about treating the sample with hydrazine beforehand to decompose arginine to ornithine and quanting arginine as ornithine. MY best suggestion is either do HPLC or amino acid analyzer or Use the ECF for the nonpolars and get a cheap capillary electrophoresis to do arginine serine and threonine as those are the problem child amino acids on GC.
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