I have a question for HPLC and LC/MS experts. I'm trying to develop a LC/MS (SIM mode) method for separation of coeluting impurities of synthetic peptide (double addition impurities). While most of the impurities are behaving well and can be easily motinored in the SIM mode, one impurity (molecular weight corresponds to addition of Glycine molecule, so +57) cannot be determined. There is a wide shoulder at the end of the peak. Through spiking, it was determined that it is not a +Gly impurity, but rather something else with very similar molecular weight that interferes with the analysis of +Gly impurity. Has anybody else encountered this problem - additional +57 peak?
There seems to be nothing in the synthetic procedure with the same MW.
Thanks,
Tatsiana
Addition of 57 Da to a peptide always suggests me carbamidomethylation. Typical modification used in proteomics profiling. After incubation with iodoacetamide reduced cysteine may easily be modified.
Other amino acids which can be modified:
K / H / D / E / S / T / Y
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