Method for quantifying trifluoroacetic acid in peptides by 1 H -19 F NMR spectral correlation

Widely used in peptide synthesis and purification, trifluoroacetic acid (TFA) can affect properties of corresponding salts, which are challenging to characterize without weighing samples. We report a rapid, non-destructive NMR method to quantify TFA without sample weighing by correlating 1 H and 19 F integrations using a fluorinated alcohol as internal references. A fluorinated dipeptide model was first used to benchmark HFIP and TFE as probes in D 2 O at different temperatures. Different 1 H integration strategies were studied for optimal analysis of TFA content. Correlation of 1 H and 19 F signal integration was subsequently applied on a set of linear and cyclic peptides possessing up to four potential positive charges. The method was shown to quantify effectively TFA offering utility for the assessment of peptide salt composition without sample destruction.