Dear Katharina Webhofer ,

I didn't encounter this myself, but I can try to speculate.

Your peptide might form an intractable aggregate or precipitate. This can happen if the peptide is not fully deprotected or if the cleavage conditions were not optimal.

If so, one potential solution could be to try cleaving the peptide again using different conditions, such as a higher concentration of TFA or adding a scavenger like 2-mercaptoethanol to the cleavage mixture. You could also try different methods of deprotection such as using a different deprotecting agent or adjusting the pH of the deprotection buffer. Additionally, you could try refluxing the peptide in a solvent that is known to be effective in dissolving peptides, such as acetic acid or acetonitrile. It could also be useful to purify the peptide by HPLC or recrystallization before further analysis. It's also important to check the structure of the peptide by mass spectrometry or NMR to confirm that the peptide is fully deprotected and that the deprotection process did not cause any structural changes or degradation. It's also worth checking the allyl ester is fully removed during the deprotection process. If not the ester could cause aggregation or insolubility. Keep in mind that the solution may involve a combination of several steps.

Hi Katharina,

have you tried HFIP als co-solvent/solvent? HFIP has a very high solubilizing potential. For a first insight: Article The Solubility of Peptide Intermediates in Organic Solvents....

Best,

Matthias

I've seen this before with short hydrophobic peptides. I was able to recover the product by adding water to the cleavage mixture (no ether), and then purifying by reverse phase chromatography. In aqueous solution my product was prone to aggregation into a thick gel. I minimized this via controlling pH and concentration.

Try DMF (dimethylformamide). Solubilsation can be slow, so leave it overnight.

My 'go-to' solvent for insoluble peptides is 50 % acetic acid.