I am trying to separate a small peptide (4 amino acids) synthesized using solid-phase synthesis after cleavage. I have tried dissolving the peptide in diethyl ether, but it did not percipitate from the cleavage reagent. What other methods can I use to effectively percipitate the peptide from the cleavage reagent and resin? Any suggestions or protocols would be greatly appreciated.
Hi Shima,
Thanks for the question. I had a hard time to precipitate short peptides. I just evaporate the cleavage reagents. Blowing air can evaporate it in the fume hood. Ideally, you can use rotavapor to evaporate as well. You can read my method in my paper here:
https://pubmed.ncbi.nlm.nih.gov/35406720/
However, you need to use only water 2.5% and triisopropylsilane 2.5% in TFA instead of trifluoroacetic acid (TFA), anisole, thioanisole, and dithiothreitol to cleave the peptides from the resin and to remove all the protecting groups. This is a great source for you to learn more.
https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/chemistry-and-synthesis/peptide-synthesis/fmoc-cleavage-deprotection
I hope it helps.
Thank you David for your kind answer. My issue is realating to the non-perciptation of the peptide in face to the diethyl ether. The cleavage cocktail is TFA/TIS/Water (95,2.5,2.5). How can percipitate and isolate peptide without diethyl ether?Would you please let me know your consideration? Thank you.
As I mentioned in my first answer, just evaporate cleavage reagents (TFA/TIS/Water) and your peptide will precipitate. Do not use diethyl ether. As Recarda mentioned, your peptide dissolves in diethyl ether. However, using derivating reactions to modify the peptide makes your work much harder.
To isolate the peptide, you will use chromatography techniques. In my paper, I have used HPLC to isolate my peptides. Please read my paper. I hope it helps.
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