I have injected a hydrophobic peptide into analytical C18 column and i have tried to cleaning the column applying a lot of protocols using acetonitrile, methanol, isopropanol, but in the blank profile still appear peptide signal (with significant intensity). Does anyone have experience using guanidine to remove hydrophobic peptides from C18 column?
Hi Lidia,
Would it be an option to add some protease (chymotrypsin, for example) with the appropriate buffer to the column to have that digest your hydrophobic peptide off?
Hope this helps!
Thanks for your answer!!
I don't think so, because the peptide i am working is short, it has 6 residues of amino acids and is very hydrophobic! The curious is that after cleaning with many solvents, including 2-propanol, peptide still appear in the blank. Do you have experience using guanidine to remove peptides like this? I want to elute with mobile phase containing guanidine, but i am affraid about clogging the column.
Try to modify your mobile phase with the addition of more strength solvent like THF. The increasing composition will elute your target. For a routine application, this approach will worsen the resolution thus reducing the alkyl chain length can improve the elution strategy. Choose C8 or C4 ligands for reproducible and carry over free analysis.
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